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Sample records for leishmania infantum vector

  1. Lutzomyia migonei is a permissive vector competent for Leishmania infantum.

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    Guimarães, Vanessa Cristina Fitipaldi Veloso; Pruzinova, Katerina; Sadlova, Jovana; Volfova, Vera; Myskova, Jitka; Filho, Sinval Pinto Brandão; Volf, Petr

    2016-03-17

    Leishmania infantum is the most widespread etiological agent of visceral leishmaniasis (VL) in the world, with significant mortality rates in human cases. In Latin America, this parasite is primarily transmitted by Lutzomyia longipalpis, but the role of Lutzomyia migonei as a potential vector for this protozoan has been discussed. Laboratory and field investigations have contributed to this hypothesis; however, proof of the vector competence of L. migonei has not yet been provided. In this study, we evaluate for the first time the susceptibility of L. migonei to L. infantum. Females of laboratory-reared L. migonei were fed through a chick-skin membrane on rabbit blood containing L. infantum promastigotes, dissected at 1, 5 and 8 days post-infection (PI) and checked microscopically for the presence, intensity and localisation of Leishmania infections. In addition, morphometric analysis of L. infantum promastigotes was performed. High infection rates of both L. infantum strains tested were observed in L. migonei, with colonisation of the stomodeal valve already on day 5 PI. At the late-stage infection, most L. migonei females had their cardia and stomodeal valve colonised by high numbers of parasites, and no significant differences were found compared to the development in L. longipalpis. Metacyclic forms were found in all parasite-vector combinations since day 5 PI. We propose that Lutzomyia migonei belongs to sand fly species permissive to various Leishmania spp. Here we demonstrate that L. migonei is highly susceptible to the development of L. infantum. This, together with its known anthropophily, abundance in VL foci and natural infection by L. infantum, constitute important evidence that L. migonei is another vector of this parasite in Latin America.

  2. Vector Competence of Lutzomyia cruzi Naturally Demonstrated for Leishmania infantum and Suspected for Leishmania amazonensis.

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    de Oliveira, Everton Falcão; Oshiro, Elisa Teruya; Fernandes, Wagner Souza; Ferreira, Alda Maria Teixeira; de Oliveira, Alessandra Gutierrez; Galati, Eunice Aparecida Bianchi

    2017-01-11

    Corumbá city is one of the oldest visceral leishmaniasis-endemic foci in the state of Mato Grosso do Sul, Brazil, where the transmission of Leishmania infantum has been attributed to Lutzomyia cruzi Aiming at investigating the parameters of the vectorial capacity of Lu. cruzi for L. infantum, a project was undertaken in this city. Among these parameters, vector competence was investigated and the results obtained are reported herein. Of the 12 hamsters exposed to feed wild-caught female sandflies, two developed infection with L. infantum and surprisingly, one with Leishmania amazonensis In addition, hamsters with L. infantum infection were bitten only by females of Lu. cruzi, whereas the hamster infected with L. amazonensis was bitten by 124 Lu. cruzi females and one of Evandromyia corumbaensis Although there is a strong suspicion regarding the competence of Lu. cruzi in transmitting L. amazonensis naturally, it was not demonstrated. © The American Society of Tropical Medicine and Hygiene.

  3. Asymptomatic dogs are highly competent to transmit Leishmania (Leishmania) infantum chagasi to the natural vector.

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    Laurenti, Márcia Dalastra; Rossi, Claudio Nazaretian; da Matta, Vânia Lúcia Ribeiro; Tomokane, Thaise Yumie; Corbett, Carlos Eduardo Pereira; Secundino, Nágila Francinete Costa; Pimenta, Paulo Filemon Paulocci; Marcondes, Mary

    2013-09-23

    We evaluated the ability of dogs naturally infected with Leishmania (Leishmania) infantum chagasi to transfer the parasite to the vector and the factors associated with transmission. Thirty-eight infected dogs were confirmed to be infected by direct observation of Leishmania in lymph node smears. Dogs were grouped according to external clinical signs and laboratory data into symptomatic (n=24) and asymptomatic (n=14) animals. All dogs were sedated and submitted to xenodiagnosis with F1-laboratory-reared Lutzomyia longipalpis. After blood digestion, sand flies were dissected and examined for the presence of promastigotes. Following canine euthanasia, fragments of skin, lymph nodes, and spleen were collected and processed using immunohistochemistry to evaluate tissue parasitism. Specific antibodies were detected using an enzyme-linked immunosorbent assay. Antibody levels were found to be higher in symptomatic dogs compared to asymptomatic dogs (p=0.0396). Both groups presented amastigotes in lymph nodes, while skin parasitism was observed in only 58.3% of symptomatic and in 35.7% of asymptomatic dogs. Parasites were visualized in the spleens of 66.7% and 71.4% of symptomatic and asymptomatic dogs, respectively. Parasite load varied from mild to intense, and was not significantly different between groups. All asymptomatic dogs except for one (93%) were competent to transmit Leishmania to the vector, including eight (61.5%) without skin parasitism. Sixteen symptomatic animals (67%) infected sand flies; six (37.5%) showed no amastigotes in the skin. Skin parasitism was not crucial for the ability to infect Lutzomyia longipalpis but the presence of Leishmania in lymph nodes was significantly related to a positive xenodiagnosis. Additionally, a higher proportion of infected vectors that fed on asymptomatic dogs was observed (p=0.0494). Clinical severity was inversely correlated with the infection rate of sand flies (p=0.027) and was directly correlated with antibody

  4. Could Phlebotomus mascittii play a role as a natural vector for Leishmania infantum? New data.

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    Obwaller, Adelheid G; Karakus, Mehmet; Poeppl, Wolfgang; Töz, Seray; Özbel, Yusuf; Aspöck, Horst; Walochnik, Julia

    2016-08-19

    The occurrence of phlebotomine sand flies in Central Europe was questioned until they were recorded for the first time in Germany in 1999, and ten years later also in Austria. The aim of this study was to investigate sand flies collected in Austria for their carrier status of Leishmania spp. From 2012 to 2013 field studies were conducted in eastern Austria. Altogether, 22 individuals of sand flies were found, all morphologically identified as Phlebotomus (Transphlebotomus) mascittii Grassi, 1908. Twelve non-engorged female specimens with no visible remnants of a blood meal in their bodies were individually investigated for Leishmania spp. by ITS-1 real-time PCR. One out of these was positive for Leishmania, identified as Leishmania infantum by DNA sequencing. This finding suggests that L. infantum is not excreted by P. mascittii and possibly can establish an infection within P. mascittii. Interestingly, an asymptomatic dog living on the farm where this sand fly had been caught was also Leishmania-positive. This study provides new data on the suspected vector capacity of P. mascittii, being the northernmost sand fly species in Europe and in most central European regions the only sand fly species found. Proven vector capacity of P. mascittii for Leishmania spp. would be of significant medico-veterinary importance, not only with respect to expanding sand fly populations in Central Europe related to global warming, but also in the light of globalization and increasing movements of humans.

  5. The Gut Microbiome of the Vector Lutzomyia longipalpis Is Essential for Survival of Leishmania infantum.

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    Kelly, Patrick H; Bahr, Sarah M; Serafim, Tiago D; Ajami, Nadim J; Petrosino, Joseph F; Meneses, Claudio; Kirby, John R; Valenzuela, Jesus G; Kamhawi, Shaden; Wilson, Mary E

    2017-01-17

    The vector-borne disease leishmaniasis, caused by Leishmania species protozoa, is transmitted to humans by phlebotomine sand flies. Development of Leishmania to infective metacyclic promastigotes in the insect gut, a process termed metacyclogenesis, is an essential prerequisite for transmission. Based on the hypothesis that vector gut microbiota influence the development of virulent parasites, we sequenced midgut microbiomes in the sand fly Lutzomyia longipalpis with or without Leishmania infantum infection. Sucrose-fed sand flies contained a highly diverse, stable midgut microbiome. Blood feeding caused a decrease in microbial richness that eventually recovered. However, bacterial richness progressively decreased in L. infantum-infected sand flies. Acetobacteraceae spp. became dominant and numbers of Pseudomonadaceae spp. diminished coordinately as the parasite underwent metacyclogenesis and parasite numbers increased. Importantly, antibiotic-mediated perturbation of the midgut microbiome rendered sand flies unable to support parasite growth and metacyclogenesis. Together, these data suggest that the sand fly midgut microbiome is a critical factor for Leishmania growth and differentiation to its infective state prior to disease transmission. Leishmania infantum, a parasitic protozoan causing fatal visceral leishmaniasis, is transmitted to humans through the bite of the sand fly Lutzomyia longipalpis Development of the parasite to its virulent metacyclic state occurs in the sand fly gut. In this study, the microbiota within the Lu. longipalpis midgut was delineated by 16S ribosomal DNA (rDNA) sequencing, revealing a highly diverse community composition that lost diversity as parasites developed to their metacyclic state and increased in abundance in infected flies. Perturbing sand fly gut microbiota with an antibiotic cocktail, which alone had no effect on either the parasite or the fly, arrested both the development of virulent parasites and parasite expansion

  6. Leishmania infantum nicotinamidase is required for late-stage development in its natural sand fly vector, Phlebotomus perniciosus

    OpenAIRE

    Gazanion, Elodie; Seblova, V.; Votypka, J.; Vergnes, Baptiste; Garcia, Deborah; Volf, P.; Sereno, Denis

    2012-01-01

    Leishmania infantum nicotinamidase, encoded by the Lipnc1 gene, converts nicotinamide into nicotinic acid to ensure Nicotinamide-Adenine-Dinucleotide (NAD(+)) biosynthesis. We were curious to explore the role of this enzyme during L infantum development in its natural sand fly vector, Phlebotomus perniciosus (Diptera, Phlebotominae), using null mutants with a deleted Lipnc1 gene. The null mutants developed as well as the wild type L infantum at the early time points post their ingestion withi...

  7. Molecular identification of Lutzomyia migonei (Diptera: Psychodidae) as a potential vector for Leishmania infantum (Kinetoplastida: Trypanosomatidae).

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    Rodrigues, Ana Caroline Moura; Melo, Luciana Magalhães; Magalhães, Rafaela Damasceno; de Moraes, Nélio Batista; de Souza Júnior, Antônio Domingos; Bevilaqua, Claudia Maria Leal

    2016-04-15

    Visceral leishmaniasis (VL) in Brazil is caused by the protozoan Leishmania infantum. This parasite is transmitted by the bite of a female sand fly. The most important sand fly species in VL transmission is Lutzomyia longipalpis. In Fortaleza, the capital of Ceará State, Brazil, the simultaneous occurrence of Lutzomyia migonei and L. longipalpis was detected in localities where VL transmission is observed. The purpose of this study was to determine conclusively if L. migonei can be found naturally infected with L. infantum in key focus in Fortaleza. Using a CDC traps we performed phlebotomine capture during one year. External morphological features and qPCR targeting species-specific gene sequences of Lutzomyia species were used to identify the female phlebotomine sand flies. The molecular identification of the Leishmania species was performed using qPCR targeting species-specific gene sequences of L. infantum and Leishmania braziliensis. The males L. migonei abundance was higher in the rainy season. Humidity and rainfall positively correlated with males L. migonei abundance, while temperature showed a negative correlation. The correlation between the density of L. migonei female with rainfall, relative air humidity, and temperature were not statistically significant. According to the molecular data produced by qPCR amplifications, three positive sand flies were identified as L. longipalpis, and one was identified as L. migonei. The infection rate was 0.35% and 0.18%, respectively. The parasite load was 32,492±2572 L. infantum in L. migonei while the L. longipalpis had parasite loads between 2,444,964.6±116,000 and 6,287,130±124,277. Our findings confirm L. migonei as a potential vector of VL in Fortaleza at a molecular level. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. Increased transmission potential of Leishmania major/Leishmania infantum hybrids

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    Volf, Petr; Benkova, Ivana; Myskova, Jitka; Sadlova, Jovana; Campino, Lenea; Ravel, Christophe

    2007-01-01

    Development of Leishmania infantum/Leishmania major hybrids was studied in two sand fly species. In Phlebotomus papatasi, which supported development of L. major but not L. infantum, the hybrids produced heavy late-stage infections with high numbers of metacyclic promastigotes. In the permissive vector Lutzomyia longipalpis, all Leishmania strains included in this study developed well. Hybrids were found to express L. major lipophosphoglycan, apparently enabling them to survive in P. papatasi...

  9. Leishmania infantum nicotinamidase is required for late-stage development in its natural sand fly vector, Phlebotomus perniciosus.

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    Gazanion, Elodie; Seblova, Veronika; Votypka, Jan; Vergnes, Baptiste; Garcia, Déborah; Volf, Petr; Sereno, Denis

    2012-04-01

    Leishmania infantum nicotinamidase, encoded by the Lipnc1 gene, converts nicotinamide into nicotinicacid to ensure Nicotinamide–Adenine–Dinucleotide (NAD+) biosynthesis. We were curious to explore the role of this enzyme during L. infantum development in its natural sand fly vector, Phlebotomus perniciosus (Diptera, Phlebotominae), using null mutants with a deleted Lipnc1 gene. The null mutants developed as well as the wild type L. infantum at the early time points post their ingestion within the bloodmeal. In contrast, once the blood meal digestion was completed, the null mutants were unable to develop further and establish late-stage infections. Data highlight the importance of the nicotinamide degradation pathway for Leishmania development in sand flies. They indicate that the endogenous nicotinamidase is essential for Leishmania development in the sand fly after the blood meal has been digested and the remnants defecated.

  10. Seasonal Dynamics of Phlebotomine Sand Fly Species Proven Vectors of Mediterranean Leishmaniasis Caused by Leishmania infantum.

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    Bulent Alten

    2016-02-01

    Full Text Available The recent geographical expansion of phlebotomine vectors of Leishmania infantum in the Mediterranean subregion has been attributed to ongoing climate changes. At these latitudes, the activity of sand flies is typically seasonal; because seasonal phenomena are also sensitive to general variations in climate, current phenological data sets can provide a baseline for continuing investigations on sand fly population dynamics that may impact on future scenarios of leishmaniasis transmission. With this aim, in 2011-2013 a consortium of partners from eight Mediterranean countries carried out entomological investigations in sites where L. infantum transmission was recently reported.A common protocol for sand fly collection included monthly captures by CDC light traps, complemented by sticky traps in most of the sites. Collections were replicated for more than one season in order to reduce the effects of local weather events. In each site, the trapping effort was left unchanged throughout the survey to legitimate inter-seasonal comparisons. Data from 99,000 collected specimens were analyzed, resulting in the description of seasonal dynamics of 56,000 sand flies belonging to L. infantum vector species throughout a wide geographical area, namely P. perniciosus (Portugal, Spain and Italy, P. ariasi (France, P. neglectus (Greece, P. tobbi (Cyprus and Turkey, P. balcanicus and P. kandelakii (Georgia. Time of sand fly appearance/disappearance in collections differed between sites, and seasonal densities showed variations in each site. Significant correlations were found between latitude/mean annual temperature of sites and i the first month of sand fly appearance, that ranged from early April to the first half of June; ii the type of density trend, varying from a single peak in July/August to multiple peaks increasing in magnitude from May through September. A 3-modal trend, recorded for P. tobbi in Cyprus, represents a novel finding for a L. infantum vector

  11. Seasonal Dynamics of Phlebotomine Sand Fly Species Proven Vectors of Mediterranean Leishmaniasis Caused by Leishmania infantum.

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    Alten, Bulent; Maia, Carla; Afonso, Maria Odete; Campino, Lenea; Jiménez, Maribel; González, Estela; Molina, Ricardo; Bañuls, Anne Laure; Prudhomme, Jorian; Vergnes, Baptiste; Toty, Celine; Cassan, Cécile; Rahola, Nil; Thierry, Magali; Sereno, Denis; Bongiorno, Gioia; Bianchi, Riccardo; Khoury, Cristina; Tsirigotakis, Nikolaos; Dokianakis, Emmanouil; Antoniou, Maria; Christodoulou, Vasiliki; Mazeris, Apostolos; Karakus, Mehmet; Ozbel, Yusuf; Arserim, Suha K; Erisoz Kasap, Ozge; Gunay, Filiz; Oguz, Gizem; Kaynas, Sinan; Tsertsvadze, Nikoloz; Tskhvaradze, Lamzira; Giorgobiani, Ekaterina; Gramiccia, Marina; Volf, Petr; Gradoni, Luigi

    2016-02-01

    The recent geographical expansion of phlebotomine vectors of Leishmania infantum in the Mediterranean subregion has been attributed to ongoing climate changes. At these latitudes, the activity of sand flies is typically seasonal; because seasonal phenomena are also sensitive to general variations in climate, current phenological data sets can provide a baseline for continuing investigations on sand fly population dynamics that may impact on future scenarios of leishmaniasis transmission. With this aim, in 2011-2013 a consortium of partners from eight Mediterranean countries carried out entomological investigations in sites where L. infantum transmission was recently reported. A common protocol for sand fly collection included monthly captures by CDC light traps, complemented by sticky traps in most of the sites. Collections were replicated for more than one season in order to reduce the effects of local weather events. In each site, the trapping effort was left unchanged throughout the survey to legitimate inter-seasonal comparisons. Data from 99,000 collected specimens were analyzed, resulting in the description of seasonal dynamics of 56,000 sand flies belonging to L. infantum vector species throughout a wide geographical area, namely P. perniciosus (Portugal, Spain and Italy), P. ariasi (France), P. neglectus (Greece), P. tobbi (Cyprus and Turkey), P. balcanicus and P. kandelakii (Georgia). Time of sand fly appearance/disappearance in collections differed between sites, and seasonal densities showed variations in each site. Significant correlations were found between latitude/mean annual temperature of sites and i) the first month of sand fly appearance, that ranged from early April to the first half of June; ii) the type of density trend, varying from a single peak in July/August to multiple peaks increasing in magnitude from May through September. A 3-modal trend, recorded for P. tobbi in Cyprus, represents a novel finding for a L. infantum vector. Adults

  12. Canine visceral leishmaniasis in the metropolitan area of São Paulo: Pintomyia fischeri as potential vector of Leishmania infantum

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    Galvis-Ovallos Fredy

    2017-01-01

    Full Text Available American visceral leishmaniasis is a zoonosis caused by Leishmania infantum and transmitted mainly by Lutzomyia longipalpis. However, canine cases have been reported in the absence of this species in the Greater São Paulo region, where Pintomyia fischeri and Migonemyia migonei are the predominant species. This raises the suspicion that they could be acting as vectors. Therefore, this study sought to investigate specific vector capacity parameters of these species and to compare them with those of Lu. longipalpis s.l. Among these parameters the blood feeding rate, the survival, and the susceptibility to the development of Le. infantum were evaluated for the three species, and the attractiveness of dogs to Pi. fischeri and Mg. migonei was evaluated. The estimated interval between blood meals was shorter for Lu. longipalpis s.l, followed by Pi. fischeri and Mg. migonei. The infection rate with Le. infantum flagellates in Lu. longipalpis was 9.8%, in Pi. fischeri 4.8%, and in Mg. migonei nil. The respective infective life expectancies (days of Lu. longipalpis, Mg. migonei, and Pi. fischeri were 2.4, 1.94, and 1.68. Both Pi. fischeri and Mg. migonei were captured in the kennel with a predominance (95% of Pi. fischeri. Considering the great attractiveness of dogs to Pi. fischeri, its susceptibility to infection by Le. infantum, infective life expectancies, and predominance in Greater São Paulo, this study presents evidence of Pi. fischeri as a potential vector of this parasite in the region.

  13. Incrimination of Phlebotomus kandelakii and Phlebotomus balcanicus as vectors of Leishmania infantum in Tbilisi, Georgia.

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    Ekaterina Giorgobiani

    Full Text Available A survey of potential vector sand flies was conducted in the neighboring suburban communities of Vake and Mtatsminda districts in an active focus of visceral Leishmaniasis (VL in Tbilisi, Georgia. Using light and sticky-paper traps, 1,266 male and 1,179 female sand flies were collected during 2006-2008. Five Phlebotomus species of three subgenera were collected: Phlebotomus balcanicus Theodor and Phlebotomus halepensis Theodor of the subgenus Adlerius; Phlebotomus kandelakii Shchurenkova and Phlebotomus wenyoni Adler and Theodor of the subgenus Larroussius; Phlebotomus sergenti Perfil'ev of the subgenus Paraphlebotomus. Phlebotomus sergenti (35.1% predominated in Vake, followed by P. kandelakii (33.5%, P. balcanicus (18.9%, P. halepensis (12.2%, and P. wenyoni (0.3%. In Mtatsminda, P. kandelakii (76.8% comprised over three fourths of collected sand flies, followed by P. sergenti (12.6%, P. balcanicus (5.8%, P. halepensis (3.7%, and P. wenyoni (1.1%. The sand fly season in Georgia is exceptionally short beginning in early June, peaking in July and August, then declining to zero in early September. Of 659 female sand flies examined for Leishmania, 12 (1.8% specimens without traces of blood were infected including 10 of 535 P. kandelakii (1.9% and two of 40 P. balcanicus (5.0%. Six isolates were successfully cultured and characterized as Leishmania by PCR. Three isolates from P. kandelakii (2 and P. balcanicus (1 were further identified as L. infantum using sequence alignment of the 70 kDa heat-shock protein gene. Importantly, the sand fly isolates showed a high percent identity (99.8%-99.9% to human and dog isolates from the same focus, incriminating the two sand fly species as vectors. Blood meal analysis showed that P. kandelakii preferentially feeds on dogs (76% but also feeds on humans. The abundance, infection rate and feeding behavior of P. kandelakii and the infection rate in P. balcanicus establish these species as vectors in the Tbilisi

  14. Incrimination of Phlebotomus kandelakii and Phlebotomus balcanicus as vectors of Leishmania infantum in Tbilisi, Georgia.

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    Giorgobiani, Ekaterina; Lawyer, Phillip G; Babuadze, Giorgi; Dolidze, Nato; Jochim, Ryan C; Tskhvaradze, Lamzira; Kikaleishvili, Konstantin; Kamhawi, Shaden

    2012-01-01

    A survey of potential vector sand flies was conducted in the neighboring suburban communities of Vake and Mtatsminda districts in an active focus of visceral Leishmaniasis (VL) in Tbilisi, Georgia. Using light and sticky-paper traps, 1,266 male and 1,179 female sand flies were collected during 2006-2008. Five Phlebotomus species of three subgenera were collected: Phlebotomus balcanicus Theodor and Phlebotomus halepensis Theodor of the subgenus Adlerius; Phlebotomus kandelakii Shchurenkova and Phlebotomus wenyoni Adler and Theodor of the subgenus Larroussius; Phlebotomus sergenti Perfil'ev of the subgenus Paraphlebotomus. Phlebotomus sergenti (35.1%) predominated in Vake, followed by P. kandelakii (33.5%), P. balcanicus (18.9%), P. halepensis (12.2%), and P. wenyoni (0.3%). In Mtatsminda, P. kandelakii (76.8%) comprised over three fourths of collected sand flies, followed by P. sergenti (12.6%), P. balcanicus (5.8%), P. halepensis (3.7%), and P. wenyoni (1.1%). The sand fly season in Georgia is exceptionally short beginning in early June, peaking in July and August, then declining to zero in early September. Of 659 female sand flies examined for Leishmania, 12 (1.8%) specimens without traces of blood were infected including 10 of 535 P. kandelakii (1.9%) and two of 40 P. balcanicus (5.0%). Six isolates were successfully cultured and characterized as Leishmania by PCR. Three isolates from P. kandelakii (2) and P. balcanicus (1) were further identified as L. infantum using sequence alignment of the 70 kDa heat-shock protein gene. Importantly, the sand fly isolates showed a high percent identity (99.8%-99.9%) to human and dog isolates from the same focus, incriminating the two sand fly species as vectors. Blood meal analysis showed that P. kandelakii preferentially feeds on dogs (76%) but also feeds on humans. The abundance, infection rate and feeding behavior of P. kandelakii and the infection rate in P. balcanicus establish these species as vectors in the Tbilisi VL

  15. Epidemiologic relationship between Toscana virus infection and Leishmania infantum due to common exposure to Phlebotomus perniciosus sandfly vector.

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    Bichaud, Laurence; Souris, Marc; Mary, Charles; Ninove, Laëtitia; Thirion, Laurence; Piarroux, Raphaël P; Piarroux, Renaud; De Lamballerie, Xavier; Charrel, Rémi N

    2011-09-01

    Sand flies are recognised vectors of parasites in the genus Leishmania and a number of arthropod-borne viruses, in particular viruses within the genus Phlebovirus, family Bunyaviridae. In southern France, Toscana phlebovirus (TOSV) is recognized as a prominent cause of summer meningitis. Since Leishmania and TOSV have a common vector (Phlebotomus perniciosus), an epidemiologic link has been assumed for a long time. However, there is no scientific evidence of such a link between human leishmaniosis and phleboviral infections. To identify a possible link, we investigated the presence and distribution of antibodies against these two microorganisms (i) in individuals and (ii) at a spatial level in the city of Marseille (south-eastern France). Five hundred sera were selected randomly in the biobank of the Department of Parasitology of the Public Hospitals of Marseille. All sera were previously tested for IgG against Leishmania by Western Blotting, and TOSV IgG were detected by indirect immunofluorescence. The seropositivity rates were 21.4% for TOSV and 28% for Leishmania. Statistical analysis demonstrated that seropositivity for one pathogen was significantly associated with seropositivity to the other pathogen. This result provided the first robust evidence for the existence of an epidemiological relationship between Leishmania infantum and TOSV. Addresses of tested patients were geolocalized and integrated into Geographical Information System software, in order to test spatial relationship between the two pathogens. Spatial analysis did not allow to identify (i) specific patterns for the spatial distribution of positive serological results for TOSV or Leishmania, and (ii) a spatial relationship between Leishmania and TOSV positive serological results. This may reflect the fact that the sample studied was not powerful enough to demonstrate either a spatial clustering or co-location, i.e. that the actual risk exposure area is smaller than the mean of distance between

  16. Epidemiologic relationship between Toscana virus infection and Leishmania infantum due to common exposure to Phlebotomus perniciosus sandfly vector.

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    Laurence Bichaud

    2011-09-01

    Full Text Available Sand flies are recognised vectors of parasites in the genus Leishmania and a number of arthropod-borne viruses, in particular viruses within the genus Phlebovirus, family Bunyaviridae. In southern France, Toscana phlebovirus (TOSV is recognized as a prominent cause of summer meningitis. Since Leishmania and TOSV have a common vector (Phlebotomus perniciosus, an epidemiologic link has been assumed for a long time. However, there is no scientific evidence of such a link between human leishmaniosis and phleboviral infections. To identify a possible link, we investigated the presence and distribution of antibodies against these two microorganisms (i in individuals and (ii at a spatial level in the city of Marseille (south-eastern France. Five hundred sera were selected randomly in the biobank of the Department of Parasitology of the Public Hospitals of Marseille. All sera were previously tested for IgG against Leishmania by Western Blotting, and TOSV IgG were detected by indirect immunofluorescence. The seropositivity rates were 21.4% for TOSV and 28% for Leishmania. Statistical analysis demonstrated that seropositivity for one pathogen was significantly associated with seropositivity to the other pathogen. This result provided the first robust evidence for the existence of an epidemiological relationship between Leishmania infantum and TOSV. Addresses of tested patients were geolocalized and integrated into Geographical Information System software, in order to test spatial relationship between the two pathogens. Spatial analysis did not allow to identify (i specific patterns for the spatial distribution of positive serological results for TOSV or Leishmania, and (ii a spatial relationship between Leishmania and TOSV positive serological results. This may reflect the fact that the sample studied was not powerful enough to demonstrate either a spatial clustering or co-location, i.e. that the actual risk exposure area is smaller than the mean of

  17. Discovery of markers of exposure specific to bites of Lutzomyia longipalpis, the vector of Leishmania infantum chagasi in Latin America.

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    Clarissa Teixeira

    2010-03-01

    Full Text Available Sand flies deliver Leishmania parasites to a host alongside salivary molecules that affect infection outcomes. Though some proteins are immunogenic and have potential as markers of vector exposure, their identity and vector specificity remain elusive.We screened human, dog, and fox sera from endemic areas of visceral leishmaniasis to identify potential markers of specific exposure to saliva of Lutzomyia longipalpis. Human and dog sera were further tested against additional sand fly species. Recombinant proteins of nine transcripts encoding secreted salivary molecules of Lu. longipalpis were produced, purified, and tested for antigenicity and specificity. Use of recombinant proteins corresponding to immunogenic molecules in Lu. longipalpis saliva identified LJM17 and LJM11 as potential markers of exposure. LJM17 was recognized by human, dog, and fox sera; LJM11 by humans and dogs. Notably, LJM17 and LJM11 were specifically recognized by humans exposed to Lu. longipalpis but not by individuals exposed to Lu. intermedia.Salivary recombinant proteins are of value as markers of vector exposure. In humans, LJM17 and LJM11 emerged as potential markers of specific exposure to Lu. longipalpis, the vector of Leishmania infantum chagasi in Latin America. In dogs, LJM17, LJM11, LJL13, LJL23, and LJL143 emerged as potential markers of sand fly exposure. Testing these recombinant proteins in large scale studies will validate their usefulness as specific markers of Lu. longipalpis exposure in humans and of sand fly exposure in dogs.

  18. Discovery of markers of exposure specific to bites of Lutzomyia longipalpis, the vector of Leishmania infantum chagasi in Latin America.

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    Teixeira, Clarissa; Gomes, Regis; Collin, Nicolas; Reynoso, David; Jochim, Ryan; Oliveira, Fabiano; Seitz, Amy; Elnaiem, Dia-Eldin; Caldas, Arlene; de Souza, Ana Paula; Brodskyn, Cláudia I; de Oliveira, Camila Indiani; Mendonca, Ivete; Costa, Carlos H N; Volf, Petr; Barral, Aldina; Kamhawi, Shaden; Valenzuela, Jesus G

    2010-03-23

    Sand flies deliver Leishmania parasites to a host alongside salivary molecules that affect infection outcomes. Though some proteins are immunogenic and have potential as markers of vector exposure, their identity and vector specificity remain elusive. We screened human, dog, and fox sera from endemic areas of visceral leishmaniasis to identify potential markers of specific exposure to saliva of Lutzomyia longipalpis. Human and dog sera were further tested against additional sand fly species. Recombinant proteins of nine transcripts encoding secreted salivary molecules of Lu. longipalpis were produced, purified, and tested for antigenicity and specificity. Use of recombinant proteins corresponding to immunogenic molecules in Lu. longipalpis saliva identified LJM17 and LJM11 as potential markers of exposure. LJM17 was recognized by human, dog, and fox sera; LJM11 by humans and dogs. Notably, LJM17 and LJM11 were specifically recognized by humans exposed to Lu. longipalpis but not by individuals exposed to Lu. intermedia. Salivary recombinant proteins are of value as markers of vector exposure. In humans, LJM17 and LJM11 emerged as potential markers of specific exposure to Lu. longipalpis, the vector of Leishmania infantum chagasi in Latin America. In dogs, LJM17, LJM11, LJL13, LJL23, and LJL143 emerged as potential markers of sand fly exposure. Testing these recombinant proteins in large scale studies will validate their usefulness as specific markers of Lu. longipalpis exposure in humans and of sand fly exposure in dogs.

  19. Experimental transmission of Leishmania infantum by two major vectors: a comparison between a viscerotropic and a dermotropic strain.

    Science.gov (United States)

    Maia, Carla; Seblova, Veronika; Sadlova, Jovana; Votypka, Jan; Volf, Petr

    2011-06-01

    We quantified Leishmania infantum parasites transmitted by natural vectors for the first time. Both L. infantum strains studied, dermotropic CUK3 and viscerotropic IMT373, developed well in Phlebotomus perniciosus and Lutzomyia longipalpis. They produced heavy late-stage infection and colonized the stomodeal valve, which is a prerequisite for successful transmission. Infected sand fly females, and especially those that transmit parasites, feed significantly longer on the host (1.5-1.8 times) than non-transmitting females. Quantitative PCR revealed that P. perniciosus harboured more CUK3 strain parasites, while in L. longipalpis the intensity of infection was higher for the IMT373 strain. However, in both sand fly species the parasite load transmitted was higher for the strain with dermal tropism (CUK3). All but one sand fly female infected by the IMT373 strain transmitted less than 600 promastigotes; in contrast, 29% of L. longipalpis and 14% of P. perniciosus infected with the CUK3 strain transmitted more than 1000 parasites. The parasite number transmitted by individual sand flies ranged from 4 up to 4.19×10(4) promastigotes; thus, the maximal natural dose found was still about 250 times lower than the experimental challenge dose used in previous studies. This finding emphasizes the importance of determining the natural infective dose for the development of an accurate experimental model useful for the evaluation of new drugs and vaccines.

  20. Feeding preferences of Lutzomyia longipalpis (Diptera: Psychodidae), the sand fly vector, for Leishmania infantum (Kinetoplastida: Trypanosomatidae).

    Science.gov (United States)

    Macedo-Silva, Virgínia P; Martins, Daniella R A; De Queiroz, Paula Vivianne Souza; Pinheiro, Marcos Paulo G; Freire, Caio C M; Queiroz, José W; Dupnik, Kathryn M; Pearson, Richard D; Wilson, Mary E; Jeronimo, Selma M B; Ximenes, Maria De Fátima F M

    2014-01-01

    Leishmania infantum, the causative agent of visceral leishmaniasis (VL) in Brazil, is spread mostly by the bite of the sand fly Lutzomyia longipalpis (Lutz & Neiva). We trapped sand flies in endemic neighborhoods near Natal, Brazil, where cases of human and dog VL were documented. Amplification of species-specific cytochrome b (Cyt b) genes by polymerase chain reaction revealed that sand flies from rural and periurban areas harbored blood from different sources. The most common source ofbloodmeal was human, but blood from dog, chicken, and armadillo was also present. We tested the preference for a source of bloodmeal experimentally by feeding L. longipalpis F1 with blood from different animals. There were significant differences between the proportion of flies engorged and number of eggs laid among flies fed on different sources, varying from 8.4 to 19 (P < 0.0001). Blood from guinea pig or horse was best to support sand fly oviposition, but human blood also supported sand fly oviposition well. No sand flies fed on cats, and sand flies feeding on the opossum Monodelphis domestica Wagner produced no eggs. These data support the hypothesis that L. longipalpis is an eclectic feeder, and humans are an important source of blood for this sand fly species in periurban areas of Brazil.

  1. Predicted altitudinal shifts and reduced spatial distribution of Leishmania infantum vector species under climate change scenarios in Colombia.

    Science.gov (United States)

    González, Camila; Paz, Andrea; Ferro, Cristina

    2014-01-01

    Visceral leishmaniasis (VL) is caused by the trypanosomatid parasite Leishmania infantum (=Leishmania chagasi), and is epidemiologically relevant due to its wide geographic distribution, the number of annual cases reported and the increase in its co-infection with HIV. Two vector species have been incriminated in the Americas: Lutzomyia longipalpis and Lutzomyia evansi. In Colombia, L. longipalpis is distributed along the Magdalena River Valley while L. evansi is only found in the northern part of the Country. Regarding the epidemiology of the disease, in Colombia the incidence of VL has decreased over the last few years without any intervention being implemented. Additionally, changes in transmission cycles have been reported with urban transmission occurring in the Caribbean Coast. In Europe and North America climate change seems to be driving a latitudinal shift of leishmaniasis transmission. Here, we explored the spatial distribution of the two known vector species of L. infantum in Colombia and projected its future distribution into climate change scenarios to establish the expansion potential of the disease. An updated database including L. longipalpis and L. evansi collection records from Colombia was compiled. Ecological niche models were performed for each species using the Maxent software and 13 Worldclim bioclimatic coverages. Projections were made for the pessimistic CSIRO A2 scenario, which predicts the higher increase in temperature due to non-emission reduction, and the optimistic Hadley B2 Scenario predicting the minimum increase in temperature. The database contained 23 records for L. evansi and 39 records for L. longipalpis, distributed along the Magdalena River Valley and the Caribbean Coast, where the potential distribution areas of both species were also predicted by Maxent. Climate change projections showed a general overall reduction in the spatial distribution of the two vector species, promoting a shift in altitudinal distribution for L

  2. New Insights Into the Transmissibility of Leishmania infantum From Dogs to Sand Flies: Experimental Vector-Transmission Reveals Persistent Parasite Depots at Bite Sites.

    Science.gov (United States)

    Aslan, Hamide; Oliveira, Fabiano; Meneses, Claudio; Castrovinci, Philip; Gomes, Regis; Teixeira, Clarissa; Derenge, Candace A; Orandle, Marlene; Gradoni, Luigi; Oliva, Gaetano; Fischer, Laurent; Valenzuela, Jesus G; Kamhawi, Shaden

    2016-06-01

    Canine leishmaniasis (CanL) is a chronic fatal disease of dogs and a major source of human infection through propagation of parasites in vectors. Here, we infected 8 beagles through multiple experimental vector transmissions with Leishmania infantum-infected Lutzomyia longipalpis. CanL clinical signs varied, although live parasites were recovered from all dog spleens. Splenic parasite burdens correlated positively with Leishmania-specific interleukin 10 levels, negatively with Leishmania-specific interferon γ and interleukin 2 levels, and negatively with Leishmania skin test reactivity. A key finding was parasite persistence for 6 months in lesions observed at the bite sites in all dogs. These recrudesced following a second transmission performed at a distal site. Notably, sand flies efficiently acquired parasites after feeding on lesions at the primary bite site. In this study, controlled vector transmissions identify a potentially unappreciated role for skin at infectious bite sites in dogs with CanL, providing a new perspective regarding the mechanism of Leishmania transmissibility to vector sand flies. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America.

  3. The current status of phlebotomine sand flies in Albania and incrimination of Phlebotomus neglectus (Diptera, Psychodidae) as the main vector of Leishmania infantum.

    Science.gov (United States)

    Velo, Enkelejda; Bongiorno, Gioia; Kadriaj, Perparim; Myrseli, Teita; Crilly, James; Lika, Aldin; Mersini, Kujtim; Di Muccio, Trentina; Bino, Silvia; Gramiccia, Marina; Gradoni, Luigi; Maroli, Michele

    2017-01-01

    The incidence of visceral leishmaniasis (VL) in Albania is higher than in other countries of southern Europe, however the role of local sand fly species in the transmission of Leishmania infantum was not addressed conclusively. In 2006, a country-wide collection of sand flies performed in 14 sites selected based on recent occurrence of VL cases showed that Phlebotomus neglectus was by far the most prevalent species (95.6%). Furthermore, 15% of pools made from 422 P. neglectus females tested positive for Leishmania sp. genomic DNA. In the same year, Culicoides trapping was performed for bluetongue disease surveillance in 91 sites of southern Albania, targeting livestock farms regardless recent occurrence of VL in the surveyed areas. In 35 sites where sand flies were collected along with midges, Phlebotomus perfiliewi was the most prevalent among the Phlebotomus species identified, however search for leishmanial DNA in females of this species was unsuccessful. In 2011, sand flies were trapped in 4 sites of north Albania characterized by high VL incidence, and females were dissected to search for Leishmania infections. Both P. neglectus and P. tobbi were collected at high densities. Two positive specimens were detected from a sample of 64 P. neglectus trapped in one site (3.1%). Parasites were successfully cultured from one specimen and characterized as belonging to Leishmania infantum zymodeme MON-1, the only zymodeme so far identified as the agent of human and canine leishmaniasis in the country. Altogether our studies indicate that P. neglectus is the main leishmaniasis vector in Albania.

  4. Leishmania infantum proteophosphoglycans regurgitated by the bite of its natural sand fly vector, Lutzomyia longipalpis, promote parasite establishment in mouse skin and skin-distant tissues.

    Science.gov (United States)

    Rogers, Matthew Edward; Corware, Karina; Müller, Ingrid; Bates, Paul Andrew

    2010-10-01

    We demonstrate that a proteophosphoglycan-rich gel secreted by Leishmania infantum inside the midgut of Lutzomyia longipalpis sand flies (promastigote secretory gel) is regurgitated along with an average dose of 500 L. infantum metacyclic promastigotes per infected bite. Using both low (10³) and high (10⁵) doses of parasites in the ears of BALB/c mice we show that the infections benefit from the presence of vector saliva and parasite gel in the skin. However, chronic infection of the spleen was only enhanced in high dose co-infections with gel. These results provide the framework for a more natural experimental model of visceral leishmaniasis. Copyright © 2010. Published by Elsevier SAS.

  5. Distribución de los vectores de Leishmania infantum (Kinetoplastida: Trypanosomatidae en Colombia

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    Camila González

    2006-10-01

    Full Text Available Introducción. Debido a la importancia que tiene la vigilancia entomológica como principalmedida de control en el manejo de la leishmaniasis visceral, es necesario contar con informaciónactualizada acerca de la distribución y ecología de los insectos involucrados en la transmisiónpara optimizar las estrategias de prevención. Objetivo. Presentar la distribución actualizada geo-referenciada de L. longipalpis y L. evansi,vectores de los parásitos que causan leishmaniasis visceral en Colombia, teniendo en cuentala asociación de los insectos con su hábitat. Materiales y métodos. Los registros de distribución se obtuvieron a partir de los ejemplaresrecolectados en Colombia desde 1967. La información obtenida se organizó en una base dedatos a partir de la cual se tomaron las localidades que, posteriormente, fueron sometidas aanálisis geográficos por medio de Arc View que se utilizaron para realizar los mapas dedistribución. Resultados. Para L. longipalpis se obtuvieron 40 localidades todas distribuidas a lo largo delvalle del río Magdalena: Alto (24, Medio (11 y Bajo (5 Magdalena. L. evansi fue registradoen 19 localidades también ubicadas en el mismo valle: cinco en el Magdalena Medio y 14 elMagdalena Bajo. Conclusiones. Ambas especies demostraron una consistente asociación con regionesclasificadas principalmente como bosque seco tropical según las zonas de vida de Holdridgelo que confirma el riesgo epidemiológico de leishmaniasis visceral en estas áreas.Palabras clave: Psychodidae, leishmaniasis visceral, Colombia.

  6. Serological tests fail to discriminate dogs with visceral leishmaniasis that transmit Leishmania infantum to the vector Lutzomyia longipalpis.

    Science.gov (United States)

    Mendonça, Ivete Lopes de; Batista, Joilson Ferreira; Werneck, Guilherme Loureiro; Soares, Maria Regiane Araújo; Costa, Dorcas Lamounier; Costa, Carlos Henrique Nery

    2017-01-01

    The control of reservoirs for Leishmania infantum -induced zoonotic visceral leishmaniasis requires the identification of dogs posing a population risk. Here, we assessed the performance of several assays to identify Lutzomyia longipalpis infectious dogs. We evaluated 99 dogs that were positive for visceral leishmaniasis based on parasite identification. Serological analyses were performed using an enzyme-linked immunosorbent assay, immunofluorescence antibody tests in 1:40 and 1:80 dilutions, rapid dual path platform tests, immunochromatographic assay with a recombinant rK39 antigen, fast agglutination screening tests, and direct agglutination tests. We also performed PCR to analyze peripheral blood and xenodiagnosis. Forty-six dogs infected at least one L. longipalpis specimen. Although the serological test sensitivities were above 85% for detecting L. longipalpis infectious dogs, none showed a satisfactory performance, as both specificity (0.06 to 13%) and the area under the receiver operating characteristic curve (45 to 53%) were low. The PCR results were also weak, with a sensitivity of 30%, specificity of 72%, and an area under the receiver operating characteristic curve of 51%. The infected L. longipalpis proportion was higher among asymptomatic dogs than symptomatic dogs. Among the symptomatic dogs, those with ulceration-free skin diseases were more infectious, with an odds ratio of 9.3 (confidence interval of 1.10 - 428.5). The larger the number of insects fed, the greater the detected infectiousness. Our study supports the imperative to develop novel technologies for identifying the infectious dogs that transmit L. infantum for the benefit of public health.

  7. The midgut transcriptome of Phlebotomus (Larroussius) perniciosus, a vector of Leishmania infantum: comparison of sugar fed and blood fed sand flies.

    Science.gov (United States)

    Dostálová, Anna; Votýpka, Jan; Favreau, Amanda J; Barbian, Kent D; Volf, Petr; Valenzuela, Jesus G; Jochim, Ryan C

    2011-05-10

    Parasite-vector interactions are fundamental in the transmission of vector-borne diseases such as leishmaniasis. Leishmania development in the vector sand fly is confined to the digestive tract, where sand fly midgut molecules interact with the parasites. In this work we sequenced and analyzed two midgut-specific cDNA libraries from sugar fed and blood fed female Phlebotomus perniciosus and compared the transcript expression profiles. A total of 4111 high quality sequences were obtained from the two libraries and assembled into 370 contigs and 1085 singletons. Molecules with putative roles in blood meal digestion, peritrophic matrix formation, immunity and response to oxidative stress were identified, including proteins that were not previously reported in sand flies. These molecules were evaluated relative to other published sand fly transcripts. Comparative analysis of the two libraries revealed transcripts differentially expressed in response to blood feeding. Molecules up regulated by blood feeding include a putative peritrophin (PperPer1), two chymotrypsin-like proteins (PperChym1 and PperChym2), a putative trypsin (PperTryp3) and four putative microvillar proteins (PperMVP1, 2, 4 and 5). Additionally, several transcripts were more abundant in the sugar fed midgut, such as two putative trypsins (PperTryp1 and PperTryp2), a chymotrypsin (PperChym3) and a microvillar protein (PperMVP3). We performed a detailed temporal expression profile analysis of the putative trypsin transcripts using qPCR and confirmed the expression of blood-induced and blood-repressed trypsins. Trypsin expression was measured in Leishmania infantum-infected and uninfected sand flies, which identified the L. infantum-induced down regulation of PperTryp3 at 24 hours post-blood meal. This midgut tissue-specific transcriptome provides insight into the molecules expressed in the midgut of P. perniciosus, an important vector of visceral leishmaniasis in the Old World. Through the comparative

  8. Leishmania infantum Genetic Diversity and Lutzomyia longipalpis Mitochondrial Haplotypes in Brazil

    OpenAIRE

    Ribolla, Paulo Eduardo Martins; Gushi, Letícia Tsieme; Pires e Cruz, Maria do Socorro; Costa, Carlos Henrique Nery; Costa, Dorcas Lamounier; Lima Júnior, Manoel Sebastião da Costa; Dorval, Maria Elizabeth Moraes Cavalheiros; Gutierrez de Oliveira, Alessandra; da Cunha Santos, Mirella Ferreira; Fonseca Camargo-Neves, Vera Lúcia; Fortaleza, Carlos Magno Castello Branco; Alonso, Diego Peres

    2016-01-01

    Leishmania infantum is the etiological agent of visceral leishmaniasis (VL) in the Americas with domestic dogs being its major reservoir hosts. The main VL vector is the sandfly Lutzomyia longipalpis, while other Lutzomyia species may play a role in disease transmission. Although the genetic structure of L. infantum populations has been widely evaluated, only a few studies have addressed this subject coupled to the genetic structure of the respective sandfly vectors. In this study, we analyze...

  9. New microsatellite markers for multi-scale genetic studies on Phlebotomus ariasi Tonnoir, vector of Leishmania infantum in the Mediterranean area.

    Science.gov (United States)

    Prudhomme, Jorian; Toty, Céline; Kasap, Ozge Erisoz; Rahola, Nil; Vergnes, Baptiste; Maia, Carla; Campino, Lenea; Antoniou, Maria; Jimenez, Maribel; Molina, Ricardo; Cannet, Arnaud; Alten, Bulent; Sereno, Denis; Bañuls, Anne-Laure

    2015-02-01

    The population structure of Phlebotomus ariasi, a proven vector of Leishmania infantum in the Mediterranean area, is still poorly understood. Previously, only two microsatellite loci had been developed to study the population genetics of this species. Herein we use these loci and determined fourteen novel microsatellite loci, useful for the characterization of P. ariasi populations. These loci were tested on three populations of P. ariasi, two from France and one from Portugal. In addition, the usefulness of these markers was also evaluated on seven other sandfly species. We show, that for P. ariasi, 15 of the 16 loci selected were polymorphic, with a mean of 4.25 alleles and an observed heterozygosity of 0.299. Within the P. ariasi population of France, 11 loci were polymorphic, with an average of 2.44 alleles and an observed heterozygosity of 0.2177. The fixation index was moderate among the French populations but high between French and Portuguese populations. In addition, eight loci were also found to be amplifiable in six other Phlebotomus species. These results demonstrate the usefulness of this new set of microsatellite loci for population structure and molecular ecology studies of P. ariasi at various spatial scales, but also of other sandfly species. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. Leishmania infantum Genetic Diversity and Lutzomyia longipalpis Mitochondrial Haplotypes in Brazil.

    Science.gov (United States)

    Ribolla, Paulo Eduardo Martins; Gushi, Letícia Tsieme; Pires E Cruz, Maria do Socorro; Costa, Carlos Henrique Nery; Costa, Dorcas Lamounier; Lima Júnior, Manoel Sebastião da Costa; Dorval, Maria Elizabeth Moraes Cavalheiros; Gutierrez de Oliveira, Alessandra; da Cunha Santos, Mirella Ferreira; Fonseca Camargo-Neves, Vera Lúcia; Fortaleza, Carlos Magno Castello Branco; Alonso, Diego Peres

    2016-01-01

    Leishmania infantum is the etiological agent of visceral leishmaniasis (VL) in the Americas with domestic dogs being its major reservoir hosts. The main VL vector is the sandfly Lutzomyia longipalpis, while other Lutzomyia species may play a role in disease transmission. Although the genetic structure of L. infantum populations has been widely evaluated, only a few studies have addressed this subject coupled to the genetic structure of the respective sandfly vectors. In this study, we analyzed the population structure of L. infantum in three major VL endemic areas in Brazil and associated it with Lutzomyia longipalpis geographic structure.

  11. Parasite load in the blood and skin of dogs naturally infected by Leishmania infantum is correlated with their capacity to infect sand fly vectors.

    Science.gov (United States)

    Borja, Lairton Souza; Sousa, Orlando Marcos Farias de; Solcà, Manuela da Silva; Bastos, Leila Andrade; Bordoni, Marcelo; Magalhães, Jairo Torres; Larangeira, Daniela Farias; Barrouin-Melo, Stella Maria; Fraga, Deborah Bittencourt Mothé; Veras, Patrícia Sampaio Tavares

    2016-10-15

    The sand fly Lutzomyia longipalpis is primarily responsible for the transmission of visceral leishmaniasis (VL) in the New World, and dogs are considered to be the main urban reservoir of this disease. In order to improve the efficacy of control measures, it is essential to assess the transmission capacity of Leishmania infantum to the sand fly vector by naturally infected dogs. The present study investigated the existence of correlations between canine clinical presentation and the intensity of parasite load in the blood, skin and spleen of naturally infected dogs. In addition, we also attempted to establish correlations between the intensity of parasite load in canine tissue and the parasite load detected in sandflies five days after feeding on naturally infected dogs. A total of 23 dogs were examined and classified according to clinical manifestation of canine VL. Blood samples, splenic aspirate and skin biopsies were collected and parasite DNA was quantified by qPCR. Canine capacity to infect Lu. longipalpis with parasites was evaluated by xenodiagnosis and parasite loads were measured five days after feeding. No significant differences were observed with respect to canine clinical manifestation and the parasite loads detected in the blood, skin and spleen samples obtained from naturally infected dogs. Regardless of clinical canine visceral leishmaniasis (CVL) presentation and the degree of parasite burden, almost half of the dogs successfully infected sandflies with parasites, albeit to a low number of sandflies with correspondingly low parasite loads. Parasite loads in both canine blood and skin were shown to be positively correlated with the canine infectiousness to the sand fly vector, and positive correlations were also observed with respect to these tissues and the sand fly infection rate, as well as the parasite load detected in sandflies following xenodiagnosis. In conclusion, this indicates that parasite loads in both blood and skin can function as

  12. Gentamicin-attenuated Leishmania infantum: a clinicopathological study in dogs.

    Science.gov (United States)

    Daneshvar, Hamid; Molaei, Mohammad M; Afshar, Reza Malekpour; Kamiabi, Hosein; Burchmore, Richard; Hagan, Paul; Phillips, R Stephen

    2009-05-15

    The clinicopathological changes following infection with an attenuated line of Leishmania infantum (L. infantum H-line) were evaluated in mixed breed dogs. Two groups of dogs were infected intravenously (i.v.) or intradermally (i.d.) with L. infantum H-line and two control groups were infected i.v. or i.d. with L. infantum wild-type (L. infantum WT). None of the dogs, which were infected i.v. or i.d. with L. infantum H-line, showed any abnormalities during the observation period. In contrast, two out of three dogs, which were infected i.v. with L. infantum WT, developed clinical signs of disease. In addition, no histopathological changes were seen in the liver and spleen of the dogs infected with the attenuated line of parasite, whereas the histopathological changes in the two dogs infected i.v. with L. infantum WT were severe in form and manifested by infiltration of high numbers of inflammatory cells. No promastigotes were found in cultures set up from spleens and livers of dogs infected with L. infantum H-line at 12 months post-infection, whereas promastigotes were seen in the spleen and liver cultures from 2 dogs infected i.v. with L. infantum WT. Serum levels of total IgG anti-Leishmania antibody were raised in all dogs. The antibody level in the serum of dogs infected i.v. with L. infantum WT was higher than that in dogs infected with L. infantum H-line. These results show no clinicopathological abnormalities in the dogs infected with gentamicin-attenuated L. infantum H-line. Moreover, L. infantum H-line induced IgG anti-Leishmania antibody in the dogs.

  13. Spread of Leishmania infantum in Europe with dog travelling.

    Science.gov (United States)

    Maia, Carla; Cardoso, Luís

    2015-09-30

    Leishmania infantum is the etiological agent of canine leishmaniosis (CanL) in Europe, where it is endemic in the Mediterranean region, with dogs being considered the major reservoir of the parasite for humans and other mammalian hosts. The main transmission mode of Leishmania is by the bite of infected phlebotomine sand fly insects (genus Phlebotomus), which are the only proven vectors of this zoonotic protozoan. Less common, non-vectorial transmission between dogs include infection through transfused blood products from infected donors, transplacental and venereal transmission. CanL has exhibited an expansion to new locations in Europe, mainly northwards, either by territorial contiguity, often in association with global warming that favours vectorial transmission, or by the long-distance importation of infected dogs. The increasing incidence of CanL in countries where the disease is not endemic is challenging owners, veterinarians and government authorities. Most infected dogs in these new areas have been relocated from or travelled with their owners to endemic regions, but in some cases transmission might have also been autochthonous. In the absence of prophylactic measures, the introduction of infected dogs in areas previously free of endemic CanL but which have competent sand fly vectors can result in a potential persistence of L. infantum. The spread of L. infantum in Europe is reviewed with a focus on transmission, epidemiology and geographic distribution of endemic and non-endemic CanL, infection and disease in humans and animal hosts other than dogs, together with prevention and additional control strategies. Copyright © 2015 Elsevier B.V. All rights reserved.

  14. Experimental mixed infection of Leishmania (Leishmania) amazonensis and Leishmania (L.) infantum in hamsters (Mesocricetus auratus).

    Science.gov (United States)

    DE Lima Celeste, Jordanna Luíza; Venuto Moura, Ana Paula; França-Silva, João Carlos; Matos DE Sousa, Gabriela; Oliveira Silva, Soraia; Norma Melo, Maria; Luiz Tafuri, Wagner; Carvalho Souza, Carolina; Monteiro DE Andrade, Hélida

    2017-08-01

    In South America, visceral leishmaniasis is frequently caused by Leishmania infantum and, at an unknown frequency, by Leishmania amazonensis. Therefore, mixed infections with these organisms are possible. Mixed infections might affect the clinical course, immune response, diagnosis, treatment and epidemiology of the disease. Here we describe the clinical course of mixed infections with L. amazonensis and L. infantum in a hamster model. We show that mixed infections are associated with more severe clinical disease than infection with L. amazonensis or L. infantum alone. In spleens with mixed infections, L. infantum outcompeted L. amazonensis in the tissue, but not in culture from tissue. We found increased levels of IgG in animals infected with L. infantum. Although more than 30 bands were revealed in a Western blot, the highest immunogenicity was observed with proteins having molecular masses of 95 and 90 kDa, whereas proteins with molecular masses of lower than 50 kDa were reactive frequently with serum from hamsters infected with L. amazonensis, and proteins with molecular masses of 80 and 70 kDa were reactive only with serum from hamsters infected with L. infantum. This finding has important implications regarding the biology of Leishmania and humoral immune responses to infections with these organisms.

  15. Canine antibody response to Phlebotomus perniciosus bites negatively correlates with the risk of Leishmania infantum transmission.

    Directory of Open Access Journals (Sweden)

    Michaela Vlkova

    2011-10-01

    Full Text Available BACKGROUND: Phlebotomine sand flies are blood-sucking insects that can transmit Leishmania parasites. Hosts bitten by sand flies develop an immune response against sand fly salivary antigens. Specific anti-saliva IgG indicate the exposure to the vector and may also help to estimate the risk of Leishmania spp. transmission. In this study, we examined the canine antibody response against the saliva of Phlebotomus perniciosus, the main vector of Leishmania infantum in the Mediterranean Basin, and characterized salivary antigens of this sand fly species. METHODOLOGY/PRINCIPAL FINDINGS: Sera of dogs bitten by P. perniciosus under experimental conditions and dogs naturally exposed to sand flies in a L. infantum focus were tested by ELISA for the presence of anti-P. perniciosus antibodies. Antibody levels positively correlated with the number of blood-fed P. perniciosus females. In naturally exposed dogs the increase of specific IgG, IgG1 and IgG2 was observed during sand fly season. Importantly, Leishmania-positive dogs revealed significantly lower anti-P. perniciosus IgG2 compared to Leishmania-negative ones. Major P. perniciosus antigens were identified by western blot and mass spectrometry as yellow proteins, apyrases and antigen 5-related proteins. CONCLUSIONS: Results suggest that monitoring canine antibody response to sand fly saliva in endemic foci could estimate the risk of L. infantum transmission. It may also help to control canine leishmaniasis by evaluating the effectiveness of anti-vector campaigns. Data from the field study where dogs from the Italian focus of L. infantum were naturally exposed to P. perniciosus bites indicates that the levels of anti-P. perniciosus saliva IgG2 negatively correlate with the risk of Leishmania transmission. Thus, specific IgG2 response is suggested as a risk marker of L. infantum transmission for dogs.

  16. Natural canine infection by Leishmania infantum and Leishmania amazonensis and their implications for disease control

    Directory of Open Access Journals (Sweden)

    Letícia da Cruz Sanches

    Full Text Available Abstract Leishmaniasis is a major public health problem worldwide. Because Leishmania can adapt to new hosts or vectors, knowledge concerning the current etiological agent in dogs is important in endemic areas. This study aimed to identify the Leishmania species detected in 103 samples of peripheral blood from dogs that were naturally infected with these protozoa. The diagnosis of leishmaniasis was determined through parasitological examination, the indirect enzyme-linked immunosorbent assay (ELISA and the polymerase chain reaction (PCR. The Leishmania species were identified by means of PCR-restriction fragment length polymorphism (PCR-RFLP. The samples were subjected to PCR using oligonucleotide primers that amplify the intergenic region ITS1 of the rRNA gene in order to identify the species. The amplified DNA was digested using the restriction enzyme HaeIII. A restriction profile identical to L. amazonensis was shown in 77/103 samples and the profile was similar to L. infantum in 17/103. However, a mixed profile was shown in 9/103 samples, which impeded species identification. In conclusion, the infection in these dogs was predominantly due to L. amazonensis, thus indicating that diagnosing of cases of canine leishmaniasis needs to be reexamined, since the causative agent identified is not restricted to L. infantum.

  17. Rattus norvegicus (Rodentia: Muridae Infected by Leishmania (Leishmania infantum (syn. Le. chagasi in Brazil

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    Fabiana de Oliveira Lara-Silva

    2014-01-01

    Full Text Available In the present study we surveyed the fauna of phlebotomine sand flies and small mammals in peridomestic areas from a Brazilian municipality where the American cutaneous leishmaniasis (ACL is endemic. A total of 608 female phlebotomine sand flies were captured during nine months in 2009 and 2010. Seven different species were represented with 60% of them being Lutzomyia intermedia and Lu. whitmani, both incriminated vectors of ACL. Lu. longipalpis, a proven vector of visceral leishmaniasis (VL was also captured at high proportion (12.8%. Genomic DNA analysis of 136 species-specific pools of female sand flies followed by molecular genotyping showed the presence of Leishmania infantum DNA in two pools of Lu. longipalpis. The same Leishmania species was found in one blood sample from Rattus norvegicus among 119 blood and tissue samples analysed. This is the first report of Le. infantum in R. norvegicus in the Americas and suggests a possible role for this rodent species in the zoonotic cycle of VL. Our study coincided with the reemergence of VL in Governador Valadares.

  18. First evidence of autochthonous cases of Leishmania (Leishmania) infantum in horse (Equus caballus) in the Americas and mixed infection of Leishmania infantum and Leishmania (Viannia) braziliensis.

    Science.gov (United States)

    Soares, Isabel R; Silva, Soraia O; Moreira, Filipe Moraghi; Prado, Luan Gavião; Fantini, Priscila; Maranhão, Renata de Pino Albuquerque; da Silva Filho, José Monteiro; Melo, Maria Norma; Palhares, Maristela S

    2013-11-08

    This study reports the first evidence of infection by Leishmania infantum in Equus caballus in Americas and the first mixed infection of L. infantum/Leishmania braziliensis on this mammalian species in the world. The diagnoses was based on presence of parasites in lesions and bone marrow aspirates, their identification by using specific primers for L. infantum and L. braziliensis complexes and also serological methods IFAT and ELISA. The analysis of the PCR products suggested mixed infection in three animals. Further studies involving equine leishmaniasis are carrying out in order to clarify the dynamic of Leishmania sp. in this mammalian specie and their role in the transmission of those parasites in urban endemic area of Belo Horizonte, Minas Gerais State, Brazil. Copyright © 2013 Elsevier B.V. All rights reserved.

  19. FIRST REPORT OF CUTANEOUS LEISHMANIASIS CAUSED BY Leishmania (Leishmania infantum chagasi IN AN URBAN AREA OF RIO DE JANEIRO, BRAZIL

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    Marcelo Rosandiski LYRA

    2015-10-01

    Full Text Available SUMMARY American tegumentary leishmaniasis (ATL is an infectious disease caused by protozoa of the genus Leishmania, and transmitted by sandflies. In the state of Rio de Janeiro, almost all of the cases of American tegumentary leishmaniasis (ATL are caused by Leishmania (Viannia braziliensis, while cases of visceral leishmaniasis (VL are caused by Leishmania (Leishmania infantum chagasi. The resurgence of autochthonous VL cases in Rio de Janeiro is related to the geographic expansion of the vector Lutzomyia longipalpis and its ability to adapt to urban areas. We report the first case of leishmaniasis with exclusively cutaneous manifestations caused by L. (L. infantum chagasi in an urban area of Rio de Janeiro. An eighty-one-year-old woman presented three pleomorphic skin lesions that were not associated with systemic symptoms or visceromegalies. Multilocus enzyme electrophoresis identified L. (L. infantum chagasi, but direct smear and PCR of bone narrow were negative for Leishmania sp. (suggesting exclusively cutaneous involvement. We discuss the different dermatological presentations of viscerotropic leishmaniasis of the New and Old World, and the clinical and epidemiological importance of the case. Etiologic diagnosis of ATL based upon exclusive clinical criteria may lead to incorrect conclusions. We should be aware of the constant changes in epidemiological patterns related to leishmaniases.

  20. Spatial and temporal changes in Lutzomyia longipalpis abundance, a Leishmania infantum vector in an urban area in northeastern Argentina

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    Fernández, María Soledad; Santini, María Soledad; Cavia, Regino; Sandoval, Adolfo Enrique; Pérez, Adriana Alicia; Acardi, Soraya; Salomón, Oscar Daniel

    2013-01-01

    This study aimed to analyse changes in the spatial distribution of Lutzomyia longipalpis in Posadas, an urban area located in northeastern Argentina. Data were obtained during the summer of 2007 and 2009 through two entomological surveys of peridomiciles distributed around the city. The abundance distribution pattern for 2009 was computed and compared with the previous pattern obtained in 2007, when the first human visceral leishmaniasis cases were reported in the city. Vector abundance was also examined in relation to micro and macrohabitat characteristics. In 2007 and 2009, Lu. longipalpis was distributed among 41.5% and 31% of the households in the study area, respectively. In both years, the abundance rates at most of the trapping sites were below 30 Lu. longipalpis per trap per night; however, for areas exhibiting 30-60 Lu. longipalpis and more than 60 Lu. longipalpis, the areas increased in both size and number from 2007-2009. Lu. longipalpis was more abundant in areas with a higher tree and bush cover (a macrohabitat characteristic) and in peridomiciles with accumulated unused material (a microhabitat characteristic). These results will help to prioritise and focus control efforts by defining which peridomiciles display a potentially high abundance of Lu. longipalpis. PMID:24271040

  1. Spatial and temporal changes in Lutzomyia longipalpis abundance, a Leishmania infantum vector in an urban area in northeastern Argentina.

    Science.gov (United States)

    Fernández, María Soledad; Santini, María Soledad; Cavia, Regino; Sandoval, Adolfo Enrique; Pérez, Adriana Alicia; Acardi, Soraya; Salomón, Oscar Daniel

    2013-11-01

    This study aimed to analyse changes in the spatial distribution of Lutzomyia longipalpis in Posadas, an urban area located in northeastern Argentina. Data were obtained during the summer of 2007 and 2009 through two entomological surveys of peridomiciles distributed around the city. The abundance distribution pattern for 2009 was computed and compared with the previous pattern obtained in 2007, when the first human visceral leishmaniasis cases were reported in the city. Vector abundance was also examined in relation to micro and macrohabitat characteristics. In 2007 and 2009, Lu. longipalpis was distributed among 41.5% and 31% of the households in the study area, respectively. In both years, the abundance rates at most of the trapping sites were below 30 Lu. longipalpis per trap per night; however, for areas exhibiting 30-60 Lu. longipalpis and more than 60 Lu. longipalpis, the areas increased in both size and number from 2007-2009. Lu. longipalpis was more abundant in areas with a higher tree and bush cover (a macrohabitat characteristic) and in peridomiciles with accumulated unused material (a microhabitat characteristic). These results will help to prioritise and focus control efforts by defining which peridomiciles display a potentially high abundance of Lu. longipalpis.

  2. Spatial and temporal changes in Lutzomyia longipalpis abundance, a Leishmania infantum vector in an urban area in northeastern Argentina

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    Maria Soledad Fernandez

    2013-11-01

    Full Text Available This study aimed to analyse changes in the spatial distribution of Lutzomyia longipalpis in Posadas, an urban area located in northeastern Argentina. Data were obtained during the summer of 2007 and 2009 through two entomological surveys of peridomiciles distributed around the city. The abundance distribution pattern for 2009 was computed and compared with the previous pattern obtained in 2007, when the first human visceral leishmaniasis cases were reported in the city. Vector abundance was also examined in relation to micro and macrohabitat characteristics. In 2007 and 2009, Lu. longipalpis was distributed among 41.5% and 31% of the households in the study area, respectively. In both years, the abundance rates at most of the trapping sites were below 30 Lu. longipalpis per trap per night; however, for areas exhibiting 30-60 Lu. longipalpis and more than 60 Lu. longipalpis, the areas increased in both size and number from 2007-2009. Lu. longipalpis was more abundant in areas with a higher tree and bush cover (a macrohabitat characteristic and in peridomiciles with accumulated unused material (a microhabitat characteristic. These results will help to prioritise and focus control efforts by defining which peridomiciles display a potentially high abundance of Lu. longipalpis.

  3. An investigation on vertical transmission of Leishmania infantum in experimentally infected dogs and assessment of offspring's infectiousness potential by xenodiagnosis.

    Science.gov (United States)

    Ben Slimane, T; Chouihi, E; Ben Hadj Ahmed, S; Chelbi, I; Barhoumi, W; Cherni, S; Zoghlami, Z; Gharbi, M; Zhioua, E

    2014-12-15

    Dogs are the main reservoir host of Leishmania infantum, etiologic agent of human visceral leishmaniasis (HVL) and canine visceral leishmaniasis (CVL). Transmission of L. infantum to humans and dogs is mainly through the bite of infected sand flies. In the Western Mediterranean basin, Phlebotomus perniciosus is the main vector of L. infantum. However, occasional vertical transmission of L. infantum has been reported. This study investigated L. infantum vertical transmission in offspring of experimentally infected dogs. Among 14 surviving puppies from three female beagle dogs that developed CVL following an experimental infection with L. infantum, one was tested positive by indirect immunofluorescence antibody test, by PCR and by xenodiagnosis with a high parasite burden in the spleen at 14 months old. None of the remaining puppies were tested positive for L. infantum. These findings strongly suggest that infected puppies following vertical transmission can sustain infection and contribute in infecting sand flies with L. infantum. Any strategy for controlling CVL should take into consideration the vertical transmission of L. infantum. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. Little evidence of seasonal variation of natural infection by Leishmania infantum in dogs in Spain.

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    Fernández-Bellon, H; Solano-Gallego, L; Rodríguez-Cortés, A; Ferrer, L; Gallego, M; Alberola, J; Ramis, A

    2008-08-01

    Leishmania infantum, the etiological agent of canine leishmaniosis in the Mediterranean region, is vectored by Phlebotomus spp sandflies, which are active during the warmer months of the year. In order to determine whether seasonality in transmission induces seasonal changes in the prevalence of infection by L. infantum and of parasite-specific immune response, two groups of dogs, one in February (n=37) and another in October (n=42), were studied. Clinical signs compatible with leishmaniosis, as well as presence of microscopic skin lesions in the muzzle were recorded for all dogs. Assays were also performed for detection of L. infantum parasites in muzzle skin samples (PCR, immunohistochemistry and culture), specific serum antibodies (ELISA), and specific lymphocyte proliferation and interferon-gamma production. Although prevalence of non-specific clinical signs increased significantly after the sandfly season, this was not the case for Leishmania-specific markers: positivity by PCR (24% vs. 21%) or immunohistochemistry (3% vs. 2%) of muzzle skin samples, as well as lymphocyte proliferation (59% vs. 50%) or interferon-gamma production (21% vs. 27%) were similar in February and in October. Only prevalence of positive specific antibody titers increased noticeably in October (8% vs. 20%), although this was not statistically significant. Overall, the sandfly season did not have a marked impact on the prevalence L. infantum infection or parasite-specific immune responses analyzed in this study.

  5. Quantification of Leishmania infantum DNA in females, eggs and larvae of Rhipicephalus sanguineus

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    Otranto Domenico

    2011-04-01

    Full Text Available Background Leishmania infantum is a widespread parasite that affects dogs and humans worldwide. It is transmitted primarily by phlebotomine sand flies, but recently there has been much discussion on the role of the brown dog tick, Rhipicephalus sanguineus, as a potential vector for this protozoan. Recent laboratory and field investigations have contributed to this hypothesis, but a proof of the vector capacity of R. sanguineus has yet to be provided. Following a recent study suggesting that L. infantum passes transovarially from the female tick to her progeny the current study provides new evidence of the transovarial transmission of L. infantum in R. sanguineus. Methods Engorged females of R. sanguineus were collected from the environment in a dog shelter of southern Italy, where canine leishmaniosis is endemic. In the laboratory, 97 females that successfully laid eggs, their eggs and the originated larvae were subjected to DNA extraction and then tested by a TaqMan-based real time PCR targeting a fragment of the kinetoplast DNA (kDNA of L. infantum. Results and conclusions L. infantum kDNA was detected in engorged females, their eggs and originating larvae, with a parasite load ranging from 1.8 × 10-4 to 10.0 × 100. Certainly, the current study provides further evidence on the passage of L. infantum from R. sanguineus females to their offspring. The observation of promastigote forms in larvae is necessary to definitively confirm this hypothesis, which would raise interesting questions about the possible role of ticks in the maintenance of L. infantum infection among dogs in certain areas.

  6. The prevalence of canine Leishmania infantum infection in western China detected by PCR and serological tests

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    Chen Hai-Tang

    2011-05-01

    Full Text Available Abstract Background Canine leishmaniasis (CanL is endemic in western China, resulting in important public health problem. It is essential to evaluate the prevalence of canine Leishmania infantum infection for designing control policy. In the present study we report for the first time prevalence of Leishmania infection in dogs living in Jiuzhaigou County (Sichuan Provence, China, which is not only an important endemic area of CanL but also a tourism scenic spot, detected by PCR, ELISA and dipstick test. The results could provide key information for designing control programs against canine and human leishmaniasis. In addition, the complete sequence of the Leishmania isolate from Sichuan Province has not been reported to date and we present the sequences of 116 base-pair (bp fragment of the conserved region in the minicircle kinetoplast DNA (kDNA and the results of phylogenetic analyses based on the sequence of the amplified fragment. Results The proportion of dogs infected with Leishmania in Jiuzhaigou County was 36.79%, 9.43%, and 51.88% detected by ELISA, dipstick test, and PCR, respectively. The ELISA and PCR tests were more sensitive than dipstick test. The PCR method is the most sensitive way to detect dogs infected with Leishmania parasites. The total positive rate for infected dogs in the area was 59.43% by the three methods. The PCR products of 116-bp fragment amplified from the kDNA conserved region of dog blood samples and laboratory maintained L. infantum were DNA sequenced and the variation of the sequences was observed. The phylogenetic tree based on the sequences of 116-bp fragment reveals that L. infantum is more genetically related to visceralizing species L. donovani than to the Leishmania species associated with cutaneous disease. Conclusions More than half of dogs living in the endemic Jiuzhaigou County were infected by L. infantum. Control measures, such as treatment or eradication of infected dogs, or prohibition of

  7. Methods of Control of the Leishmania infantum Dog Reservoir: State of the Art

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    Michele Podaliri Vulpiani

    2011-01-01

    Full Text Available Leishmania infantum is a protozoan parasite causing severe vector-borne visceral diseases both in humans and dogs. The latter are the most important natural reservoir and therefore should be the main target of control measures. The real efficacy of seropositive dogs culling as a direct control method is still debated, and the new sensitivity of large part of population considers ethically unacceptable this kind of approach. Treatment of infectious dogs with one of the available therapeutic protocols is recommendable as it allows to reduce parasite burdens and therefore the possibility of transmission of Leishmania infantum to vectors. Vaccination has been proven to be a very effective control tool, but the absence of a commonly recognized diagnostic method able to distinguish vaccinate from seropositive individuals is still an important limit. Concerning indirect control methods, a number of studies have demonstrated the efficacy of topical insecticides treatment (collars, spot-on, and sprays in reducing incidence and prevalence of L. infantum. Also, the reduction of the odds of seroconversion in humans in endemic areas has been reported after the application of indirect control measures on dogs. The contemporary use of direct and indirect methods is even more effective in reducing seroprevalence in dogs.

  8. First evidence of intraclonal genetic exchange in trypanosomatids using two Leishmania infantum fluorescent transgenic clones.

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    Estefanía Calvo-Álvarez

    2014-09-01

    Full Text Available The mode of reproduction in Leishmania spp has been argued to be essentially clonal. However, recent data (genetic analysis of populations and co-infections in sand flies have proposed the existence of a non-obligate sexual cycle in the extracellular stage of the parasite within the sand fly vector. In this article we propose the existence of intraclonal genetic exchange in the natural vector of Leishmania infantum.We have developed transgenic L. infantum lines expressing drug resistance markers linked to green and red fluorescent reporters. We hypothesized whether those cells with identical genotype can recognize each other and mate. Both types of markers were successfully exchanged within the sand fly midgut of the natural vector Phlebotomus perniciosus when individuals from these species were fed with a mixture of parental clones. Using the yellow phenotype and drug resistance markers, we provide evidence for genetic exchange in L. infantum. The hybrid progeny appeared to be triploid based on DNA content analysis. The hybrid clone analyzed was stable throughout the complete parasite life cycle. The progress of infections by the hybrid clone in BALB/c mice caused a reduction in parasite loads in both spleen and liver, and provided weight values similar to those obtained with uninfected mice. Spleen arginase activity was also significantly reduced relative to parental strains.A L. infantum hybrid lineage was obtained from intraclonal genetic exchange within the midgut of the natural vector, suggesting the ability of this parasite to recognize the same genotype and mate. The yellow hybrid progeny is stable throughout the whole parasite life cycle but with a slower virulence, which correlates well with the lower arginase activity detected both in vitro and in vivo infections.

  9. Leishmania infantum and Leishmania braziliensis: differences and similarities to evade the innate immune system

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    Sarah Athayde Couto Falcão

    2016-08-01

    Full Text Available Visceral Leishmaniasis is a severe form of the disease, caused by Leishmania infantum in the New World. Patients present an anergic immune response that favors parasite establishment and spreading through tissues like bone marrow and liver. On the other hand, Leishmania braziliensis causes localized cutaneous lesions, which can be self healing in some individuals. Interactions between host and parasite are essential to understand disease pathogenesis and progression. In this context, dendritic cells (DCs act as essential bridges that connect innate and adaptive immune responses. In this way, the aim of this study was to compare the effects of these two Leishmania species, in some aspects of human dendritic cells biology to better understanding of the evasion mechanisms of Leishmania from host innate immune response. To do so, DCs were obtained from monocytes from whole peripheral blood’s healthy volunteers donors and infected with L. infantum or L. braziliensis for 24 hours. We observed similar rates of infection (around 40% as well as parasite burden for both Leishmania species. Concerning surface molecules, we observed that both parasites induced CD86 expression when DCs were infected for 24h. On the other hand, we detected a lower surface expression of CD209 in the presence of both L. braziliensis and L. infantum, but only the last one promoted the survival of dendritic cells after 24 hours. Therefore, DCs infected by both Leishmania species showed a higher expression of CD86 and a decrease of CD209 expression, suggesting that both enter DCs through CD209 molecule. However, only L. infantum had the ability to inhibit DC apoptotic death, as an evasion mechanism that enables its spreading to organs like bone marrow and liver. Lastly, L. braziliensis was more silent parasite, once it did not inhibit DC apoptosis in our in vitro model.

  10. Differentiation and Gene Flow among European Populations of Leishmania infantum MON-1

    Science.gov (United States)

    Kuhls, Katrin; Chicharro, Carmen; Cañavate, Carmen; Cortes, Sofia; Campino, Lenea; Haralambous, Christos; Soteriadou, Ketty; Pratlong, Francine; Dedet, Jean-Pierre; Mauricio, Isabel; Miles, Michael; Schaar, Matthias; Ochsenreither, Sebastian; Radtke, Oliver A.; Schönian, Gabriele

    2008-01-01

    Background Leishmania infantum is the causative agent of visceral and cutaneous leishmaniasis in the Mediterranean region, South America, and China. MON-1 L. infantum is the predominating zymodeme in all endemic regions, both in humans and dogs, the reservoir host. In order to answer important epidemiological questions it is essential to discriminate strains of MON-1. Methodology/Principal Findings We have used a set of 14 microsatellite markers to analyse 141 strains of L. infantum mainly from Spain, Portugal, and Greece of which 107 strains were typed by MLEE as MON-1. The highly variable microsatellites have the potential to discriminate MON-1 strains from other L. infantum zymodemes and even within MON-1 strains. Model- and distance-based analysis detected a considerable amount of structure within European L. infantum. Two major monophyletic groups—MON-1 and non-MON-1—could be distinguished, with non-MON-1 being more polymorphic. Strains of MON-98, 77, and 108 were always part of the MON-1 group. Among MON-1, three geographically determined and genetically differentiated populations could be identified: (1) Greece; (2) Spain islands–Majorca/Ibiza; (3) mainland Portugal/Spain. All four populations showed a predominantly clonal structure; however, there are indications of occasional recombination events and gene flow even between MON-1 and non-MON-1. Sand fly vectors seem to play an important role in sustaining genetic diversity. No correlation was observed between Leishmania genotypes, host specificity, and clinical manifestation. In the case of relapse/re-infection, only re-infections by a strain with a different MLMT profile can be unequivocally identified, since not all strains have individual MLMT profiles. Conclusion In the present study for the first time several key epidemiological questions could be addressed for the MON-1 zymodeme, because of the high discriminatory power of microsatellite markers, thus creating a basis for further epidemiological

  11. Differentiation and gene flow among European populations of Leishmania infantum MON-1.

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    Katrin Kuhls

    2008-07-01

    Full Text Available Leishmania infantum is the causative agent of visceral and cutaneous leishmaniasis in the Mediterranean region, South America, and China. MON-1 L. infantum is the predominating zymodeme in all endemic regions, both in humans and dogs, the reservoir host. In order to answer important epidemiological questions it is essential to discriminate strains of MON-1.We have used a set of 14 microsatellite markers to analyse 141 strains of L. infantum mainly from Spain, Portugal, and Greece of which 107 strains were typed by MLEE as MON-1. The highly variable microsatellites have the potential to discriminate MON-1 strains from other L. infantum zymodemes and even within MON-1 strains. Model- and distance-based analysis detected a considerable amount of structure within European L. infantum. Two major monophyletic groups-MON-1 and non-MON-1-could be distinguished, with non-MON-1 being more polymorphic. Strains of MON-98, 77, and 108 were always part of the MON-1 group. Among MON-1, three geographically determined and genetically differentiated populations could be identified: (1 Greece; (2 Spain islands-Majorca/Ibiza; (3 mainland Portugal/Spain. All four populations showed a predominantly clonal structure; however, there are indications of occasional recombination events and gene flow even between MON-1 and non-MON-1. Sand fly vectors seem to play an important role in sustaining genetic diversity. No correlation was observed between Leishmania genotypes, host specificity, and clinical manifestation. In the case of relapse/re-infection, only re-infections by a strain with a different MLMT profile can be unequivocally identified, since not all strains have individual MLMT profiles.In the present study for the first time several key epidemiological questions could be addressed for the MON-1 zymodeme, because of the high discriminatory power of microsatellite markers, thus creating a basis for further epidemiological investigations.

  12. Transplacental transmission of Leishmania infantum as a means for continued disease incidence in North America.

    Science.gov (United States)

    Boggiatto, Paola Mercedes; Gibson-Corley, Katherine Nicole; Metz, Kyle; Gallup, Jack Michael; Hostetter, Jesse Michael; Mullin, Kathleen; Petersen, Christine Anne

    2011-04-12

    Dogs are the predominant domestic reservoir for human L. infantum infection. Zoonotic visceral leishmaniasis (ZVL) is an emerging problem in some U.S. dog breeds, with an annual quantitative PCR prevalence of greater than 20% within an at-risk Foxhound population. Although classically Leishmania is transmitted by infected sand flies and phlebotomine sand flies exist in the United States, means of ongoing L. infantum transmission in U.S. dogs is currently unknown. Possibilities include vertical (transplacental/transmammary) and horizontal/venereal transmission. Several reports have indicated that endemic ZVL may be transmitted vertically. Our aims for this present study were to establish whether vertical/transplacental transmission was occurring in this population of Leishmania-infected US dogs and determine the effect that this means of transmission has on immune recognition of Leishmania. A pregnant L. infantum-infected dam donated to Iowa State University gave birth in-house to 12 pups. Eight pups humanely euthanized at the time of birth and four pups and the dam humanely euthanized three months post-partum were studied via L. infantum-kinetoplast specific quantitative PCR (kqPCR), gross and histopathological assessment and CD4+ T cell proliferation assay. This novel report describes disseminated L. infantum parasites as identified by kqPCR in 8 day old pups born to a naturally-infected, seropositive U.S. dog with no travel history. This is the first report of vertical transmission of L. infantum in naturally-infected dogs in North America, emphasizing that this novel means of transmission could possibly sustain infection within populations. Evidence that vertical transmission of ZVL may be a driving force for ongoing disease in an otherwise non-endemic region has significant implications on current control strategies for ZVL, as at present parasite elimination efforts in endemic areas are largely focused on vector-borne transmission between canines and people

  13. Transplacental transmission of Leishmania infantum as a means for continued disease incidence in North America.

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    Paola Mercedes Boggiatto

    2011-04-01

    Full Text Available Dogs are the predominant domestic reservoir for human L. infantum infection. Zoonotic visceral leishmaniasis (ZVL is an emerging problem in some U.S. dog breeds, with an annual quantitative PCR prevalence of greater than 20% within an at-risk Foxhound population. Although classically Leishmania is transmitted by infected sand flies and phlebotomine sand flies exist in the United States, means of ongoing L. infantum transmission in U.S. dogs is currently unknown. Possibilities include vertical (transplacental/transmammary and horizontal/venereal transmission. Several reports have indicated that endemic ZVL may be transmitted vertically.Our aims for this present study were to establish whether vertical/transplacental transmission was occurring in this population of Leishmania-infected US dogs and determine the effect that this means of transmission has on immune recognition of Leishmania.A pregnant L. infantum-infected dam donated to Iowa State University gave birth in-house to 12 pups. Eight pups humanely euthanized at the time of birth and four pups and the dam humanely euthanized three months post-partum were studied via L. infantum-kinetoplast specific quantitative PCR (kqPCR, gross and histopathological assessment and CD4+ T cell proliferation assay.This novel report describes disseminated L. infantum parasites as identified by kqPCR in 8 day old pups born to a naturally-infected, seropositive U.S. dog with no travel history. This is the first report of vertical transmission of L. infantum in naturally-infected dogs in North America, emphasizing that this novel means of transmission could possibly sustain infection within populations.Evidence that vertical transmission of ZVL may be a driving force for ongoing disease in an otherwise non-endemic region has significant implications on current control strategies for ZVL, as at present parasite elimination efforts in endemic areas are largely focused on vector-borne transmission between canines

  14. Disseminated feline leishmaniosis due to Leishmania infantum in Southern France.

    Science.gov (United States)

    Ozon, C; Marty, P; Pratlong, F; Breton, C; Blein, M; Lelièvre, A; Haas, P

    1998-02-28

    A fortuitously discovered case of feline leishmaniosis is reported. The parasites were found in the skin and the bone marrow of a domestic female cat that spontaneously died after a few weeks of evolution. Serological tests for FeLV, FIV and PIF virus detection gave negative results. By using Western blot serology, a characteristic pattern of leishmaniosis was obtained and by performing an isoenzyme electrophoresis, a Leishmania infantum MON-1 strain was identified. The same zymodeme is implicated in most of the canine and human leishmaniosis in Southern Europe. A study on the prevalence of asymptomatic feline leismaniosis is foreseen.

  15. Effects of nitro-heterocyclic derivatives against Leishmania (Leishmania) infantum promastigotes and intracellular amastigotes.

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    Petri e Silva, Simone Carolina Soares; Palace-Berl, Fanny; Tavares, Leoberto Costa; Soares, Sandra Regina Castro; Lindoso, José Angelo Lauletta

    2016-04-01

    Leishmaniasis is an overlooked tropical disease affecting approximately 1 million people in several countries. Clinical manifestation depends on the interaction between Leishmania and the host's immune response. Currently available treatment options for leishmaniasis are limited and induce severe side effects. In this research, we tested nitro-heterocyclic compounds (BSF series) as a new alternative against Leishmania. Its activity was measured in Leishmania (Leishmania) infantum promastigotes and intracellular amastigotes using MTT colorimetric assay. Additionally, we assessed the phosphatidylserine exposure by promastigotes, measured by flow cytometry, as well as nitric oxide production, measured by Griess' method. The nitro-heterocyclic compounds (BSF series) showed activity against L. (L.) infantum promastigotes, inducting the phosphatidylserine exposition by promastigotes, decreasing intracellular amastigotes and increasing oxide nitric production. The selectivity index was more prominent to Leishmania than to macrophages. Compared to amphotericin b, our compounds presented higher IC50, however the selectivity index was more specific to parasite than to amphotericin b. In conclusion, these nitro-heterocyclic compounds showed to be promising as an anti-Leishmania drug, in in vitro studies. Copyright © 2016 Elsevier Inc. All rights reserved.

  16. Transmissibility of Leishmania infantum from maned wolves (Chrysocyon brachyurus) and bush dogs (Speothos venaticus) to Lutzomyia longipalpis.

    Science.gov (United States)

    Mol, Juliana P S; Soave, Semíramis A; Turchetti, Andréia P; Pinheiro, Guilherme R G; Pessanha, Angela T; Malta, Marcelo C C; Tinoco, Herlandes P; Figueiredo, Luiza A; Gontijo, Nelder F; Paixão, Tatiane A; Fujiwara, Ricardo T; Santos, Renato L

    2015-09-15

    Leishmania (Leishmania) infantum is the cause of visceral leishmaniasis in the Americas. The disease is transmitted mostly through the bite of the invertebrate vector, the phlebotomine Lutzomyia longipalpis in the New World. Although the domestic dog is considered the most important reservoir of the disease, other mammalian, including wildlife, are susceptible to infection. The goal of this study was to perform xenodiagnosis to evaluate the capacity of naturally infected maned wolves (Chrysocyon brachyurus) and bush dogs (Speothos venaticus) to transmit Leishmania infantum to female sand flies (L. longipalpis). Xenodiagnoses were performed in February and August, 2013, when 77.7% (three maned wolves and four bush dogs) or 100% of the animals were positive, respectively. However, parasite loads in the engorged sand flies was low (longipalpis, although the parasite loads in engorged phlebotomines exposed to these animals were very low. Copyright © 2015 Elsevier B.V. All rights reserved.

  17. Presence of sandflies infected with Leishmania infantum and Massilia virus in the Marseille urban area.

    Science.gov (United States)

    Faucher, B; Bichaud, L; Charrel, R; Mary, C; Izri, A; de Lamballerie, X; Piarroux, R

    2014-05-01

    Leishmaniasis is considered a rural disease in Europe. However, circumstantial evidence has indicated urban transmission of leishmaniasis and phleboviruses in the urban area of Marseille, France. To investigate this urban transmission, sandflies were trapped in 33 locations in the urban area (horse farms, public gardens and a residential area). Sandflies were always captured: 87.8% were Phlebotomus perniciosus, a vector of Leishmania infantum and Toscana and Massilia viruses. RT-PCR and cell culture inoculation identified the Massilia virus in 2/99 pools of sandflies, and PCR identified Leishmania in 5/99. No dual infection was observed, but both pathogens were detected in samples from the same trapping site. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

  18. Pathology of dogs in Campo Grande, MS, Brazil naturally co-infected with Leishmania infantum and Ehrlichia canis

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    Gisele Braziliano Andrade

    2014-12-01

    Full Text Available Different parasites that commonly occur concomitantly can influence one another, sometimes with unpredictable effects. We evaluated pathological aspects of dogs naturally co-infected with Leishmania infantum and Ehrlichia canis. The health status of the dogs was investigated based on histopathological, hematological and biochemical analyses of 21 animals infected solely with L. infantum and 22 dogs co- infected with L. infantum and E. canis. The skin of both groups showed chronic, predominantly lymphohistioplasmacytic inflammatory reaction. The plasmacytosis in the lymphoid tissues was likely related with the hypergammaglobulinemia detected in all the dogs. The disorganization of extracellular matrix found in the reticular dermis of the inguinal region and ear, characterized by the substitution of thick collagen fibers for thin fibers, was attributed to the degree of inflammatory reaction, irrespective of the presence of parasites. In addition, the histopathological analysis revealed that twice as many dogs in the co-infected group presented Leishmania amastigotes in the ear skin than those infected solely with Leishmania, increasing the possibility of becoming infected through sand fly vectors. Our findings highlight the fact that the health of dogs infected concomitantly with L. infantum and E. canis is severely compromised due to their high levels of total plasma protein, globulins, alkaline phosphatase and creatine kinase, and severe anemia.

  19. Prevalence of antibodies to Leishmania infantum and Toxoplasma gondii in horses from the north of Portugal

    Science.gov (United States)

    Background Leishmania infantum and Toxoplasma gondii are protozoa with zoonotic and economic importance. Prevalences of antibodies to these agents were assessed in 173 horses from the north of Portugal. Findings Antibodies to L. infantum were detected by the direct agglutination test (DAT); seven (...

  20. Prevalence of antibodies to Leishmania infantum and Toxoplasma gondii in horses from the north of Portugal

    NARCIS (Netherlands)

    Lopes, Ana Patrícia; Sousa, Susana; Dubey, Jp; Ribeiro, Ana J.; Silvestre, Ricardo; Cotovio, Mário; Schallig, Henk Dfh; Cardoso, Luís; Cordeiro-da-Silva, Anabela

    2013-01-01

    Leishmania infantum and Toxoplasma gondii are protozoa with zoonotic and economic importance. Prevalences of antibodies to these agents were assessed in 173 horses from the north of Portugal. Antibodies to L. infantum were detected by the direct agglutination test (DAT); seven (4.0%) horses were

  1. Low seroprevalence of Leishmania infantum infection in cats from northern Portugal based on DAT and ELISA

    NARCIS (Netherlands)

    Cardoso, Luís; Lopes, Ana Patrícia; Sherry, Kate; Schallig, Henk; Solano-Gallego, Laia

    2010-01-01

    Cats have been considered playing a role in the epidemiology of leishmaniasis caused by Leishmania infantum, an endemic zoonosis in countries of the Mediterranean basin. The present study assessed the prevalence of antibodies to L. infantum in 316 domestic cats from northern Portugal, by means of

  2. Tyrosine aminotransferase from Leishmania infantum: A new drug target candidate

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    Miguel Angel Moreno

    2014-12-01

    Full Text Available Leishmania infantum is the etiological agent of zoonotic visceral leishmaniasis in the Mediterranean basin. The disease is fatal without treatment, which has been based on antimonial pentavalents for more than 60 years. Due to resistances, relapses and toxicity to current treatment, the development of new drugs is required. The structure of the L. infantum tyrosine aminotransferase (LiTAT has been recently solved showing important differences with the mammalian orthologue. The characterization of LiTAT is reported herein. This enzyme is cytoplasmic and is over-expressed in the more infective stages and nitric oxide resistant parasites. Unlike the mammalian TAT, LiTAT is able to use ketomethiobutyrate as co-substrate. The pharmacophore model of LiTAT with this specific co-substrate is described herein. This may allow the identification of new inhibitors present in the databases. All the data obtained support that LiTAT is a good target candidate for the development of new anti-leishmanial drugs.

  3. Thrichomys laurentius (Rodentia; Echimyidae as a putative reservoir of Leishmania infantum and L. braziliensis: patterns of experimental infection.

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    André Luiz Rodrigues Roque

    Full Text Available The importance of the genus Thrichomys in the retention of infection and transmission of Leishmania species is supported by previous studies that describe an ancient interaction between caviomorphs and trypanosomatids and report the natural infection of Thrichomys spp. Moreover, these rodents are widely dispersed in Brazil and recognized as important hosts of other tripanosomatids. Our main purpose was to evaluate the putative role of Thrichomys laurentius in the retention of infection and amplification of the transmission cycle of Leishmania infantum and L. braziliensis. Male and female T. laurentius (n = 24 born in captivity were evaluated for the retention of infection with these Leishmania species and followed up by parasitological, serological, hematological, biochemical, histological, and molecular assays for 3, 6, 9, or 12 months post infection (mpi. T. laurentius showed its competence as maintenance host for the two inoculated Leishmania species. Four aspects should be highlighted: (i re-isolation of parasites 12 mpi; (ii the low parasitic burden displayed by T. laurentius tissues; (iii the early onset and maintenance of humoral response, and (iv the similar pattern of infection by the two Leishmania species. Both Leishmania species demonstrated the ability to invade and maintain itself in viscera and skin of T. laurentius, and no rodent displayed any lesion, histological changes, or clinical evidence of infection. We also wish to point out the irrelevance of the adjective dermotropic or viscerotropic to qualify L. braziliensis and L. infantum, respectively, when these species are hosted by nonhuman hosts. Our data suggest that T. laurentius may act at least as a maintenance host of both tested Leishmania species since it maintained long-lasting infections. Moreover, it cannot be discarded that Leishmania spp. infection in free-ranging T. laurentius could result in higher parasite burden due the more stressing conditions in the wild

  4. In vitro anti-leishmania evaluation of nickel complexes with a triazolopyrimidine derivative against Leishmania infantum and Leishmania braziliensis.

    Science.gov (United States)

    Ramírez-Macías, Inmaculada; Maldonado, Carmen R; Marín, Clotilde; Olmo, Francisco; Gutiérrez-Sánchez, Ramón; Rosales, María J; Quirós, Miguel; Salas, Juan M; Sánchez-Moreno, Manuel

    2012-07-01

    Studies on the anti-proliferative activity in vitro of seven ternary nickel (II) complexes with a triazolopyrimidine derivative and different aliphatic or aromatic amines as auxiliary ligands against promastigote and amastigote forms of Leishmania infantum and Leishmania braziliensis have been carried out. These compounds are not toxic for the host cells and two of them are effective at lower concentrations than the reference drug used in the present study (Glucantime). In general, the in vitro growth rate of Leishmania spp. was reduced, its capacity to infect cells was negatively affected and the multiplication of the amastigotes decreased. Ultrastructural analysis and metabolism excretion studies were executed in order to propose a possible mechanism for the action of the assayed compounds. Our results show that the potential mechanism is at the level of organelles membranes, either by direct action on the microtubules or by their disorganization, leading to vacuolization, degradation and ultimately cell death. Copyright © 2012 Elsevier Inc. All rights reserved.

  5. Different host complement systems and their interactions with saliva from Lutzomyia longipalpis (Diptera, Psychodidae and Leishmania infantum promastigotes.

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    Antonio Ferreira Mendes-Sousa

    Full Text Available BACKGROUND: Lutzomyia longipalpis is the vector of Leishmania infantum in the New World, and its saliva inhibits classical and alternative human complement system pathways. This inhibition is important in protecting the insect´s midgut from damage by the complement. L. longipalpis is a promiscuous blood feeder and must be protected against its host's complement. The objective of this study was to investigate the action of salivary complement inhibitors on the sera of different host species, such as dogs, guinea pigs, rats and chickens, at a pH of 7.4 (normal blood pH and 8.15 (the midgut pH immediately after a blood meal. We also investigated the role of the chicken complement system in Leishmania clearance in the presence and absence of vector saliva. RESULTS: The saliva was capable of inhibiting classical pathways in dogs, guinea pigs and rats at both pHs. The alternative pathway was not inhibited except in dogs at a pH of 8.15. The chicken classical pathway was inhibited only by high concentrations of saliva and it was better inhibited by the midgut contents of sand flies. Neither the saliva nor the midgut contents had any effect on the avian alternative pathway. Fowl sera killed L. infantum promastigotes, even at a low concentration (2%, and the addition of L. longipalpis saliva did not protect the parasites. The high body temperature of chickens (40°C had no effect on Leishmania viability during our assays. CONCLUSION: Salivary inhibitors act in a species-specific manner. It is important to determine their effects in the natural hosts of Leishmania infantum because they act on canid and rodent complements but not on chickens (which do not harbour the parasite. Moreover, we concluded that the avian complement system is the probable mechanism through which chickens eliminate Leishmania and that their high body temperature does not influence this parasite.

  6. Different host complement systems and their interactions with saliva from Lutzomyia longipalpis (Diptera, Psychodidae) and Leishmania infantum promastigotes.

    Science.gov (United States)

    Mendes-Sousa, Antonio Ferreira; Nascimento, Alexandre Alves Sousa; Queiroz, Daniel Costa; Vale, Vladimir Fazito; Fujiwara, Ricardo Toshio; Araújo, Ricardo Nascimento; Pereira, Marcos Horácio; Gontijo, Nelder Figueiredo

    2013-01-01

    Lutzomyia longipalpis is the vector of Leishmania infantum in the New World, and its saliva inhibits classical and alternative human complement system pathways. This inhibition is important in protecting the insect´s midgut from damage by the complement. L. longipalpis is a promiscuous blood feeder and must be protected against its host's complement. The objective of this study was to investigate the action of salivary complement inhibitors on the sera of different host species, such as dogs, guinea pigs, rats and chickens, at a pH of 7.4 (normal blood pH) and 8.15 (the midgut pH immediately after a blood meal). We also investigated the role of the chicken complement system in Leishmania clearance in the presence and absence of vector saliva. The saliva was capable of inhibiting classical pathways in dogs, guinea pigs and rats at both pHs. The alternative pathway was not inhibited except in dogs at a pH of 8.15. The chicken classical pathway was inhibited only by high concentrations of saliva and it was better inhibited by the midgut contents of sand flies. Neither the saliva nor the midgut contents had any effect on the avian alternative pathway. Fowl sera killed L. infantum promastigotes, even at a low concentration (2%), and the addition of L. longipalpis saliva did not protect the parasites. The high body temperature of chickens (40°C) had no effect on Leishmania viability during our assays. Salivary inhibitors act in a species-specific manner. It is important to determine their effects in the natural hosts of Leishmania infantum because they act on canid and rodent complements but not on chickens (which do not harbour the parasite). Moreover, we concluded that the avian complement system is the probable mechanism through which chickens eliminate Leishmania and that their high body temperature does not influence this parasite.

  7. Influence of the Microenvironment in the Transcriptome of Leishmania infantum Promastigotes: Sand Fly versus Culture.

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    Pedro J Alcolea

    2016-05-01

    Full Text Available Zoonotic visceral leishmaniasis is a vector-borne disease caused by Leishmania infantum in the Mediterranean Basin, where domestic dogs and wild canids are the main reservoirs. The promastigote stage replicates and develops within the gut of blood-sucking phlebotomine sand flies. Mature promastigotes are injected in the dermis of the mammalian host and differentiate into the amastigote stage within parasitophorous vacuoles of phagocytic cells. The major vector of L. infantum in Spain is Phlebotomus perniciosus. Promastigotes are routinely axenized and cultured to mimic in vitro the conditions inside the insect gut, which allows for most molecular, cellular, immunological and therapeutical studies otherwise inviable. Culture passages are known to decrease infectivity, which is restored by passage through laboratory animals. The most appropriate source of promastigotes is the gut of the vector host but isolation of the parasite is technically challenging. In fact, this option is not viable unless small samples are sufficient for downstream applications like promastigote cultures and nucleic acid amplification. In this study, in vitro infectivity and differential gene expression have been studied in cultured promastigotes at the stationary phase and in promastigotes isolated from the stomodeal valve of the sand fly P. perniciosus. About 20 ng RNA per sample could be isolated. Each sample contained L. infantum promastigotes from 20 sand flies. RNA was successfully amplified and processed for shotgun genome microarray hybridization analysis. Most differentially regulated genes are involved in regulation of gene expression, intracellular signaling, amino acid metabolism and biosynthesis of surface molecules. Interestingly, meta-analysis by hierarchical clustering supports that up-regulation of 22.4% of the differentially regulated genes is specifically enhanced by the microenvironment (i.e. sand fly gut or culture. The correlation between cultured and

  8. Leishmania (Leishmania infantum chagasi em canídeos silvestres mantidos em cativeiro, no Estado de Mato Grosso Leishmania (Leishmania infantum chagasi in wild canids kept in captivity in the State of Mato Grosso

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    Nely Pinheiro Souza

    2010-06-01

    Full Text Available INTRODUÇÃO: Leishmaniose visceral é uma zoonose que acomete diversos mamíferos tendo os canídeos domésticos como principais reservatórios em ambiente urbano. A presente nota descreve a infecção de canídeos silvestres por Leishmania (Leishmania infantum chagasi mantidos em cativeiro no Estado de Mato Grosso, Brasil. MÉTODOS: De seis raposas (Cerdocyon thous e um cachorro vinagre (Spheotos venaticus, foram coletadas amostras de pele, medula óssea e linfonodo para detecção e caracterização de Leishmania sp pela técnica de PCR-RFLP. RESULTADOS: Todos as animais pesquisados apresentaram-se positivos para Leishmania (L. infantum chagasi. CONCLUSÕES: Destaca-se a importância de monitoramento adequado dos mesmos, além do maior controle desta enfermidade já que estes animais estão em ambientes de recreação pública.INTRODUCTION: Visceral leishmaniasis is a zoonosis that affects many mammals, and domestic canids are the main reservoirs in urban environments. This note describes infection by Leishmania (Leishmania infantum chagasi among wild canids kept in captivity in the State of Mato Grosso, Brazil. METHODS: Skin, bone marrow and lymph node samples were collected from six crab-eating foxes (Cerdocyon thous and one bush dog (Spheotos venaticus, in order to detect and characterize Leishmania using the PCR-RFLP technique. RESULTS: All the animals studied were positive for Leishmania (L. infantum chagasi. CONCLUSIONS: This study highlights the importance of adequate monitoring of these animals, as well as greater control of this disease, given that these animals are in a public recreation environment.

  9. Transplacental Transmission of Leishmania infantum as a Means for Continued Disease Incidence in North America

    OpenAIRE

    Boggiatto, Paola Mercedes; Gibson-Corley, Katherine Nicole; Metz, Kyle; Gallup, Jack Michael; Hostetter, Jesse Michael; Mullin, Kathleen; Petersen, Christine Anne

    2011-01-01

    Background Dogs are the predominant domestic reservoir for human L. infantum infection. Zoonotic visceral leishmaniasis (ZVL) is an emerging problem in some U.S. dog breeds, with an annual quantitative PCR prevalence of greater than 20% within an at-risk Foxhound population. Although classically Leishmania is transmitted by infected sand flies and phlebotomine sand flies exist in the United States, means of ongoing L. infantum transmission in U.S. dogs is currently unknown. Possibilities incl...

  10. A comparison of molecular markers to detect Lutzomyia longipalpis naturally infected with Leishmania (Leishmania infantum

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    Kárita Cláudia Freitas-Lidani

    2014-07-01

    Full Text Available The aim of the present study was to detect natural infection by Leishmania (Leishmania infantum in Lutzomyia longipalpis captured in Barcarena, state of Pará, Brazil, through the use of three primer sets. With this approach, it is unnecessary to previously dissect the sandfly specimens. DNA of 280 Lu. longipalpis female specimens were extracted from the whole insects. PCR primers for kinetoplast minicircle DNA (kDNA, the mini-exon gene and the small subunit ribosomal RNA (SSU-rRNA gene of Leishmania were used, generating fragments of 400 bp, 780 bp and 603 bp, respectively. Infection by the parasite was found with the kDNA primer in 8.6% of the cases, with the mini-exon gene primer in 7.1% of the cases and with the SSU-rRNA gene primer in 5.3% of the cases. These data show the importance of polymerase chain reaction as a tool for investigating the molecular epidemiology of visceral leishmaniasis by estimating the risk of disease transmission in endemic areas, with the kDNA primer representing the most reliable marker for the parasite.

  11. Leishmania chagasi/infantum : further investigations on Leishmania tropisms in atypical cutaneous and visceral leishmaniasis foci in Central America

    NARCIS (Netherlands)

    Campos Ponce, M.; Ponce, C.; Ponce, E; Maingon, R.D.

    2005-01-01

    In Central America, apparently genetically identical Leishmania chagasi/infantum parasites cause cutaneous (CL) and visceral leishmaniasis (VL), the latter being more frequent in young children. The present study investigated if there were pathology-related differences in virulence between Honduran

  12. Leishmania chagasi/infantum: further investigations on Leishmania tropisms in atypical cutaneous and visceral leishmaniasis foci in Central America.

    NARCIS (Netherlands)

    Campos Ponce, M.; Ponce, C.; Ponce, E.; Maingon, R.D.

    2005-01-01

    In Central America, apparently genetically identical Leishmania chagasi/infantum parasites cause cutaneous (CL) and visceral leishmaniasis (VL), the latter being more frequent in young children. The present study investigated if there were pathology-related differences in virulence between Honduran

  13. The use of kDNA minicircle subclass relative abundance to differentiate between Leishmania (L.) infantum and Leishmania (L.) amazonensis.

    Science.gov (United States)

    Ceccarelli, Marcello; Galluzzi, Luca; Diotallevi, Aurora; Andreoni, Francesca; Fowler, Hailie; Petersen, Christine; Vitale, Fabrizio; Magnani, Mauro

    2017-05-16

    Leishmaniasis is a neglected disease caused by many Leishmania species, belonging to subgenera Leishmania (Leishmania) and Leishmania (Viannia). Several qPCR-based molecular diagnostic approaches have been reported for detection and quantification of Leishmania species. Many of these approaches use the kinetoplast DNA (kDNA) minicircles as the target sequence. These assays had potential cross-species amplification, due to sequence similarity between Leishmania species. Previous works demonstrated discrimination between L. (Leishmania) and L. (Viannia) by SYBR green-based qPCR assays designed on kDNA, followed by melting or high-resolution melt (HRM) analysis. Importantly, these approaches cannot fully distinguish L. (L.) infantum from L. (L.) amazonensis, which can coexist in the same geographical area. DNA from 18 strains/isolates of L. (L.) infantum, L. (L.) amazonensis, L. (V.) braziliensis, L. (V.) panamensis, L. (V.) guyanensis, and 62 clinical samples from L. (L.) infantum-infected dogs were amplified by a previously developed qPCR (qPCR-ML) and subjected to HRM analysis; selected PCR products were sequenced using an ABI PRISM 310 Genetic Analyzer. Based on the obtained sequences, a new SYBR-green qPCR assay (qPCR-ama) intended to amplify a minicircle subclass more abundant in L. (L.) amazonensis was designed. The qPCR-ML followed by HRM analysis did not allow discrimination between L. (L.) amazonensis and L. (L.) infantum in 53.4% of cases. Hence, the novel SYBR green-based qPCR (qPCR-ama) has been tested. This assay achieved a detection limit of 0.1 pg of parasite DNA in samples spiked with host DNA and did not show cross amplification with Trypanosoma cruzi or host DNA. Although the qPCR-ama also amplified L. (L.) infantum strains, the C q values were dramatically increased compared to qPCR-ML. Therefore, the combined analysis of C q values from qPCR-ML and qPCR-ama allowed to distinguish L. (L.) infantum and L. (L.) amazonensis in 100% of tested samples

  14. Natural infection of Didelphis aurita (Mammalia: Marsupialia with Leishmania infantum in Brazil

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    Carreira João Carlos

    2012-06-01

    Full Text Available Abstract Background The opossum Didelphis have been considered as natural hosts of Leishmania parasites in the New World, suggesting an important role in the epidemiology of Visceral Leishmaniasis (VL. Among six extant species that belong to the genus Didelphis, only two (D. marsupialis and D. albiventris, have been mentioned as natural hosts of Leishmania infantum in Brazil and Colombia. In the present paper, it is reported for the first time, the observation of intracellular parasites (amastigotes in tissues of Didelphis aurita naturally infected with Leishmania infantum in Brazil. We also discuss some aspects associated to the relationship between L. infantum and the geographical distribution of some species of the genus Didelphis. Methods The opossums studied were caught by wire traps (Tomahawk in Barra de Guaratiba, a peri-urban area in Rio de Janeiro. The opossums were killed with an overdose of Thiopental sodium.At necropsy, macroscopic alterations were examined and samples from liver, spleen, lymph nodes, ear, abdominal skin, scent glands and bone marrow were collected for parasitological and molecular diagnoses. Results Forty-eight opossums were captured in an AVL endemic region, 30 being caught in a mangrove area and eighteen animals in a forest area near to some residential-yards. Among the thirty opossums trapped in the mangrove area, all of them were negative by both imprint and sera samples assayed on Dipstick Tests, that is a test based on a combination of protein-A colloidal gold conjugate and rk39 Leishmania antigen to detect anti-Leishmania antibody in serum or plasma. At the macroscopic examination one out of eighteen opossums, caught close to the forest, presented alterations compatible with spleen hypertrophy and three were positive by Dipstick Tests (16.6% and presented amastigotes in the spleen and in one of them, the parasites were also observed in a submandibular lymph node. Leishmania infantum infections were confirmed

  15. Natural infection of Didelphis aurita (Mammalia: Marsupialia) with Leishmania infantum in Brazil.

    Science.gov (United States)

    Carreira, João Carlos Araujo; da Silva, Alba Valéria Machado; de Pita Pereira, Daniela; Brazil, Reginaldo Peçanha

    2012-06-07

    The opossum Didelphis have been considered as natural hosts of Leishmania parasites in the New World, suggesting an important role in the epidemiology of Visceral Leishmaniasis (VL). Among six extant species that belong to the genus Didelphis, only two (D. marsupialis and D. albiventris), have been mentioned as natural hosts of Leishmania infantum in Brazil and Colombia. In the present paper, it is reported for the first time, the observation of intracellular parasites (amastigotes) in tissues of Didelphis aurita naturally infected with Leishmania infantum in Brazil. We also discuss some aspects associated to the relationship between L. infantum and the geographical distribution of some species of the genus Didelphis. The opossums studied were caught by wire traps (Tomahawk) in Barra de Guaratiba, a peri-urban area in Rio de Janeiro. The opossums were killed with an overdose of Thiopental sodium.At necropsy, macroscopic alterations were examined and samples from liver, spleen, lymph nodes, ear, abdominal skin, scent glands and bone marrow were collected for parasitological and molecular diagnoses. Forty-eight opossums were captured in an AVL endemic region, 30 being caught in a mangrove area and eighteen animals in a forest area near to some residential-yards. Among the thirty opossums trapped in the mangrove area, all of them were negative by both imprint and sera samples assayed on Dipstick Tests, that is a test based on a combination of protein-A colloidal gold conjugate and rk39 Leishmania antigen to detect anti-Leishmania antibody in serum or plasma. At the macroscopic examination one out of eighteen opossums, caught close to the forest, presented alterations compatible with spleen hypertrophy and three were positive by Dipstick Tests (16.6%) and presented amastigotes in the spleen and in one of them, the parasites were also observed in a submandibular lymph node. Leishmania infantum infections were confirmed through dot blot hybridization using a L. infantum

  16. Low Seroprevalence of Leishmania infantum and Toxoplasma gondii in the Horse Population in Israel

    NARCIS (Netherlands)

    Aharonson-Raz, Karin; Baneth, Gad; Lopes, Ana Patrícia; Brancal, Hugo; Schallig, Henk; Cardoso, Luís; Steinman, Amir

    2015-01-01

    A cross-sectional investigation was done on the seroprevalence of Leishmania infantum and Toxoplasma gondii infection among apparently healthy horses in Israel. This survey included 383 horses distributed in 22 farms throughout Israel during the years 2011-2013. Serum samples were tested for the

  17. Kennel dogs as sentinels of Leishmania infantum, Toxoplasma gondii and Neospora caninum in Majorca Island, Spain

    Science.gov (United States)

    Kennel dogs can serve as sentinels and/or reservoirs of diseases of veterinary and zoonotic interest because they have often roamed free and lived outdoors, being exposed to pathogens. We tested for evidence of infection with three protozoans, Leishmania infantum, Toxoplasma gondii and Neospora cani...

  18. Low Seroprevalence of Leishmania infantum and Toxoplasma gondii in the Horse Population in Israel.

    Science.gov (United States)

    Aharonson-Raz, Karin; Baneth, Gad; Lopes, Ana Patrícia; Brancal, Hugo; Schallig, Henk; Cardoso, Luís; Steinman, Amir

    2015-12-01

    A cross-sectional investigation was done on the seroprevalence of Leishmania infantum and Toxoplasma gondii infection among apparently healthy horses in Israel. This survey included 383 horses distributed in 22 farms throughout Israel during the years 2011-2013. Serum samples were tested for the presence of immunoglobulin G (IgG) antibodies using the direct agglutination test (DAT) specific to Leishmania and by the modified agglutination test (MAT) for the presence of IgG antibodies to T. gondii. Low seroprevalences were detected for both L. infantum and T. gondii in the horse population in Israel; of the 338 horses tested, 6 (1.4%) were found to be seropositive for L. infantum and 11 (2.5%) for T. gondii, with no significant association between seroprevalence and demographic/environmental factors. An ongoing geographical expansion of L. infantum, previously reported in humans and dogs in Israel, was also supported by our results in horses. Here we present evidence of exposure of horses to L. infantum and T. gondii in Israel. Continuous seroprevalence surveillance in horses, such as the one performed in this study, might further elucidate the eco-epidemiology of these two important zoonotic parasites in this country.

  19. Detection of Leishmania infantum DNA in hamsters infested with ticks collected from naturally infected dogs

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    Valter dos Anjos Almeida

    2016-12-01

    Full Text Available ABSTRACT. Almeida V. dos A., da Hora T.N., Leça Júnior N.F., Carvalho F.S., da Silva A.L., Wenceslau A.A., Albuquerque G.R. & Silva F.L. Detection of Leishmania infantum DNA in hamsters infested with ticks collected from naturally infected dogs. [Detecção do DNA de Leishmania infantum em hamsters infestados com carrapatos coletados de cães naturalmente infectados.] Revista Brasileira de Medicina Veterinária, 38(4:329-333, 2016. Departamento de Ciências Agrárias e Ambientais, Universidade Estadual de Santa Cruz, Campus Soane Nazaré de Andrade, Hospital Veterinário, Km 16, Rodovia Jorge Amado, Ilhéus, BA 45662-900, Brasil. E-mail: fabiana.lessa@gmail.com The aim of this study was to investigate the role of Rhipicephalus sanguineus, the brown dog tick, in the transmission of Leishmania infantum. To accomplish this, we used 24 adult golden hamsters of both genders, and divided them into two groups: a control group (n = 4 and an experimental group (n = 20. The animals from the experimental group were infested with ticks obtained from dogs naturally infected with L. infantum. Hamsters of the control group were not infested and were maintained at the same conditions, as the infested animals. After three months of observation, animals were euthanized and they were posted to obtain samples of their blood, spleen, liver, lymph nodes, and skin. These samples were then processed by histopathology, immunohistochemistry, and polymerase chain reaction (PCR. Fourteen hamsters (70% of the experimental group tested PCR-positive for L. infantum DNA in samples of buffy coat. The results of this study indicated that R. sanguineus ticks can transmit some forms or parts of L. infantum to parasitized hamsters.

  20. Canine cutaneous leishmaniasis caused by neotropical Leishmania infantum despite of systemic disease: A case report.

    Science.gov (United States)

    Cavalcanti, Amanda; Lobo, Rogério; Cupolillo, Elisa; Bustamante, Fábio; Porrozzi, Renato

    2012-12-01

    Visceral leishmaniasis is an anthropozoonosis caused by a protozoan Leishmania infantum (syn. Leishmania chagasi). Here, we report a typical case of canine cutaneous leishmaniasis due to L. infantum infection without any other systemic symptom in one dog in the city of Rio de Janeiro, Brazil. A mongrel female dog was admitted in a veterinary clinic with reports of chronic wounds in the body. Physical examination revealed erosive lesions in the limbs, nasal ulcers, presence of ectoparasites and seborrheic dermatitis. Blood samples and fragments of healthy and injured skin were collected. The complete hemogram revealed aregenerative normocytic normochromic anemia and erythrocyte rouleaux, and biochemical analysis revealed normal renal and hepatic functions. Cytology of the muzzle and skin lesions suggested pyogranulomatous inflammatory process. The histopathology of a skin fragment was performed and revealed suspicion of protozoa accompanied by necrotizing dermatitis. The diagnosis of leishmaniasis was accomplished by positive serology, isolation of Leishmania from the skin lesion, and also by molecular test (PCR targeting the conserved region of Leishmania kDNA). Culture was positive for damaged skin samples. PCR targeting a fragment of Leishmania hsp70 gene was performed employing DNA extracted from damaged skin. RFLP of the amplified hsp70 fragment identified the parasite as L. infantum, instead of Leishmania braziliensis, the main agent of cutaneous leishmaniasis in Rio de Janeiro. Characterization of isolated promastigotes by five different enzymatic systems confirmed the species identification of the etiological agent. Serology was positive by ELISA and rapid test. This case warns to the suspicion of viscerotropic Leishmania in cases of chronic skin lesions and brings the discussion of the mechanisms involved in the parasite tissue tropism. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  1. Serum chemistry reference values for the common genet (Genetta genetta): variations associated with Leishmania infantum infection.

    Science.gov (United States)

    Millán, Javier; Chirife, Andrea D; Altet, Laura

    2015-03-01

    The role of wildlife in the epidemiology of leishmaniosis in under debate, and determining whether infection with Leishmania infantum causes illness in wild carnivores is important to determine its potential role as a reservoir. To provide for the first time serum biochemistry reference values for the common genet (Genetta genetta), and to determine variations associated with L. infantum infection. Twenty-five serum biochemistry parameters were determined in 22 wild-caught genets. Blood samples were analyzed for L. infantum DNA by means of real-time polymerase chain reaction (PCR). Two female genets were positive for L. infantum DNA but did not show any external clinical sign upon physical examination. Among other variations in the biochemistry values of these genets, one presented a higher concentration of gamma-globulins and cholesterol, whereas the other genet presented increased creatinine, bilirubin, and chloride levels when compared to uninfected females. Sex-related differences in some parameters were also reported. Infection with L. infantum may sometimes be accompanied by abnormal serum biochemistry in wild carnivores. Clinical disease may occur in L. infantum-infected wild carnivores. This has implications in the epidemiology of leishmaniosis. In addition, the data provided here would also be useful as reference values for researchers or rehabilitators working with the common genet.

  2. Seroprevalence of dogs to Leishmania infantum antigens in the territory of Crna Gora

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    Pavićević Zorica

    2014-01-01

    Full Text Available Due to expansion of vector borne diseases and their importance for human health we have investigated the prevalence of antibodies to Leishmania infantum antigens in serum samples of dogs. Leishmaniasis is a disease caused by protozoan parasites of the genus Leishmania. According to the World Health Organization the disease is a constant threat to 350 million people in 88 countries, provided that most Mediterranean countries belong to the area designated as endemic. Since Crna Gora is a country of Mediterranean basin, it is important to present the results of the investigation on the presence of antibodies on L. infantum antigens in blood serum of dogs, considering that dogs can be reservoirs and source of infection for humans. In the period from 2006. to 2010., there were tested 433 serum samples of dogs with clinical symptoms and 68 serum samples of dogs without symptoms of the disease. Those were dogs with visible symptoms of the disease such as alopecia, weight loss and lymphadenomegalia. In 66% of cases, there was known hematological profile with predominant hypochromic anemia and neutrophilic leukocytosis. As mentioned above, the investigation included also 68 asymptomatic dogs in good condition, out of which 45 were service dogs in use in military and police as well as 23 dogs from the asylum. By Indirect immunofluorescence test or Indirect Fluorescent Antibody Technique (IF-AT and Direct Agglutination Test (DAT, there was determined the presence of specific antibodies to L. infantum antigens. Positive findings in the serum dilution 1:160 was deter­mined in 258 samples (59.58% in the group of dogs with clinical symptoms, and in 5.9% in the tested asymptomatic dogs. A part of the territory of Crna Gora with its average annual air temperature of 14.6 - 17.8°C and mediterranean climate (long, warm and dry summer and mild, rainy winter has all the prerequisites for the development and maintenance of the infection. The confirmed presence of

  3. Transmission of Leishmania infantum in the Canine Leishmaniasis Focus of Mont-Rolland, Senegal: Ecological, Parasitological and Molecular Evidence for a Possible Role of Sergentomyia Sand Flies.

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    Massila Wagué Senghor

    2016-11-01

    Full Text Available Leishmania (L. infantum is the causative agent in an endemic focus of canine leishmaniasis in the Mont-Rolland district (Thiès, Senegal. In this area, the transmission cycle is well established and more than 30% of dogs and 20% of humans are seropositive for L. infantum. However, the sand fly species involved in L. infantum transmission cycle are still unknown. Between 2007 and 2010, 3654 sand flies were collected from different environments (indoor, peridomestic, farming and sylvatic areas to identify the main L. infantum vector(s. Nine sand fly species were identified. The Phlebotomus genus (n = 54 specimens; Phlebotomus (Ph duboscqi and Phlebotomus (Ph. rodhaini was markedly under-represented in comparison to the Sergentomyia genus (n = 3600 specimens; Sergentomyia (Se adleri, Se. clydei, Se. antennata, Se. buxtoni, Se. dubia, Se. schwetzi and Se. magna. Se. dubia and Se. schwetzi were the dominant species indoor and in peridomestic environments, near humans and dogs. Blood-meal analysis indicated their anthropophilic behavior. Some Se. schwetzi specimens fed also on dogs. The dissection of females in the field allowed isolating L. infantum from sand flies of the Sergentomyia genus (0.4% of Se. dubia and 0.79% of Se. schwetzi females. It is worth noting that one Se. dubia female not engorged and not gravid revealed highly motile metacyclic of L. infantum in the anterior part of the midgut. PCR-based diagnosis and sequencing targeting Leishmania kinetoplast DNA (kDNA highlighted a high rate of L. infantum-positive females (5.38% of Se. dubia, 4.19% of Se. schwetzi and 3.64% of Se. magna. More than 2% of these positive females were unfed, suggesting the parasite survival after blood-meal digestion or egg laying. L. infantum prevalence in Se. schwetzi was associated with its seroprevalence in dogs and humans and L. infantum prevalence in Se. dubia was associated with its seroprevalence in humans. These evidences altogether strongly suggest that

  4. Diagnosis of Leishmania infantum infection by Polymerase Chain Reaction in wild mammals

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    Mayara C. Lombardi

    2014-12-01

    Full Text Available Visceral leishmaniasis is a chronic infectious disease caused by Leishmania infantum (synonym: Leishmania chagasi and transmitted by the sandfly Lutzomyia longipalpis in Brazil. It is an endemic zoonosis in several regions of the country, including Belo Horizonte (State of Minas Gerais. In urban areas, the domestic dog is susceptible and considered the most important animal reservoir. However, L. infantum has been previously diagnosed in other species, including captive primates and canids. This study aimed to evaluate the presence of the agent DNA in captive animals as well as some free ranging animals from the Zoo-Botanical Foundation of Belo Horizonte by Polymerase Chain Reaction. Eighty one blood samples from primates, carnivores, ruminants, edentates, marsupial, and a monogastric herbivore were analyzed. Three primates Alouatta guariba (brown howler monkey, and two canids Speothos venaticus (bush dog were positive, demonstrating the importance of leishmaniasis control in endemic areas for preservation of wildlife species in captivity.

  5. Natural infection of Lutzomyia neivai and Lutzomyia sallesi (Diptera: Psychodidae) by Leishmania infantum chagasi in Brazil.

    Science.gov (United States)

    Saraiva, Lara; Carvalho, Gustavo M L; Gontijo, Célia M F; Quaresma, Patrícia F; Lima, Ana C V M R; Falcão, Alda L; Andrade Filho, José D

    2009-09-01

    Natural infections with Leishmania were found in females of the phlebotomine sand flies Lutzomyia neivai (Pinto) (= Nyssomyia neivai) and Lutzomyia sallesi (Galvão & Coutinho) (= Evandromyia sallesi) (Diptera: Psychodidae) from Lassance, in the Brazilian state of Minas Gerais. Promastigotes were found in the pyloric region of the former species and in the abdominal midgut of the latter species. Insects found to be infected by microscopic examination were macerated in saline solution and inoculated into hamsters. Subsequent analysis by polymerase chain reaction-restriction fragment length polymorphism revealed both isolates to belong to the species Leishmania infantum chagasi Cunha & Chagas.

  6. Prostaglandin E2/leukotriene B4 balance induced by Lutzomyia longipalpis saliva favors Leishmania infantum infection.

    Science.gov (United States)

    Araújo-Santos, Théo; Prates, Deboraci Brito; França-Costa, Jaqueline; Luz, Nívea F; Andrade, Bruno B; Miranda, José Carlos; Brodskyn, Claudia I; Barral, Aldina; Bozza, Patrícia T; Borges, Valéria Matos

    2014-12-20

    Eicosanoids and sand fly saliva have a critical role in the Leishmania infection. Here, we evaluated the effect of Lutzomyia longipalpis salivary gland sonicate (SGS) on neutrophil and monocyte recruitment and activation of eicosanoid production in a murine model of inflammation. C57BL/6 mice were inoculated intraperitonealy with Lutzomyia longipalpis SGS or Leishmania infantum or both, followed by analyses of cell recruitment, parasite load and eicosanoid production. Intraperitoneal injection of Lutzomyia longipalpis SGS together with Leishmania infantum induced an early increased parasite viability in monocytes and neutrophils. L. longipalpis SGS increased prostaglandin E2 (PGE2), but reduced leukotriene B4 (LTB4) production ex vivo in peritoneal leukocytes. In addition, the pharmacological inhibition of cyclooxygenase 2 (COX-2) with NS-398 decreased parasite viability inside macrophages during Leishmania infection in the presence of L. longipalpis SGS arguing that PGE2 production is associated with diminished parasite killing. These findings indicate that L. longipalpis SGS is a critical factor driving immune evasion of Leishmania through modulation of PGE2/LTB4 axis, which may represent an important mechanism on establishment of the infection.

  7. Molecular detection of Leishmania infantum and Leishmania tropica in rodent species from endemic cutaneous leishmaniasis areas in Morocco.

    Science.gov (United States)

    Echchakery, Mohamed; Chicharro, Carmen; Boussaa, Samia; Nieto, Javier; Carrillo, Eugenia; Sheila, Ortega; Moreno, Javier; Boumezzough, Ali

    2017-10-02

    Leishmaniasis remains a major public health problem in African nations, including Morocco, where little is known about the vertebrate reservoirs involved in the causal parasites' transmission cycles. The present study investigates the role of rodent species as potential reservoirs of Leishmania spp. in central Morocco, where both L. tropica and L. infantum have been reported. Rodents were caught from 22 sites in central Morocco, by using Sherman metal traps, and identified morphologically. For each specimen, genomic DNA was extracted from different tissues using the Speed Tools DNA extraction Kit. Then, samples were PCR-analyzed, targeting the SSU rRNA gene to detect Leishmania spp. DNA, followed by amplification of the internal transcribed spacer 1 (ITS1) and its sequencing to identify the species. A total of 197 rodents belonging to ten species were captured and identified: Rattus rattus (40.61%), Mus musculus (25.38%), Apodemus sylvaticus (8.63%), Mus spretus (7.11%), Meriones shawi (5.58%), Rattus norvegicus (4.57%), Meriones libycus (3.05%), Mastomys erythroleucus (2.03%), Gerbillus campestris (2.03%) and Lemniscomys barbarus (1.01%). Molecular analysis revealed the presence of Leishmania species in 18 specimens: six R. rattus (out of 80 captured; 7.5%), 11 M. musculus (out of 50 captured; 22%), and one R. norvegicus (out of 9 captured; 11.11%). To the best of our knowledge, L. infantum and L. tropica were identified in rodent species for the first time in Morocco. These findings suggest that rodent species may be involved in L. infantum and L. tropica transmission cycles in this country but that further studies are needed to confirm their role as reservoirs of Leishmania species in Morocco.

  8. Detection of Leishmania infantum in animals and their ectoparasites by conventional PCR and real time PCR.

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    de Morais, Rayana Carla Silva; Gonçalves, Suênia da Cunha; Costa, Pietra Lemos; da Silva, Kamila Gaudêncio; da Silva, Fernando José; Silva, Rômulo Pessoa E; de Brito, Maria Edileuza Felinto; Brandão-Filho, Sinval Pinto; Dantas-Torres, Filipe; de Paiva-Cavalcanti, Milena

    2013-04-01

    Visceral leishmaniosis (VL) is a parasitic disease caused by Leishmania infantum, which is primarily transmitted by phlebotomine sandflies. However, there has been much speculation on the role of other arthropods in the transmission of VL. Thus, the aim of this study was to assess the presence of L. infantum in cats, dogs and their ectoparasites in a VL-endemic area in northeastern Brazil. DNA was extracted from blood samples and ectoparasites, tested by conventional PCR (cPCR) and quantitative real time PCR (qPCR) targeting the L. infantum kinetoplast DNA. A total of 280 blood samples (from five cats and 275 dogs) and 117 ectoparasites from dogs were collected. Animals were apparently healthy and not previously tested by serological or molecular diagnostic methods. Overall, 213 (76.1 %) animals and 51 (43.6 %) ectoparasites were positive to L. infantum, with mean parasite loads of 795.2, 31.9 and 9.1 fg in dogs, cats and ectoparasites, respectively. Concerning the positivity between dogs and their ectoparasites, 32 (15.3 %) positive dogs were parasitized by positive ectoparasites. The overall concordance between the PCR protocols used was 59.2 %, with qPCR being more efficient than cPCR; 34.1 % of all positive samples were exclusively positive by qPCR. The high number of positive animals and ectoparasites also indicates that they could serve as sentinels or indicators of the circulation of L. infantum in risk areas.

  9. In Vitro Study on effects of Amiodarone and Ketoconazole on Leishmania infantum

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    Mohammad Hosein Razi Jalali

    2014-09-01

    Full Text Available Background & objectives: The leishmaniases are considered among the major infectious diseases affecting public health in several regions. There are many chemical agents which are effective in treatment of visceral leishmaniasis. But, overall treatment of visceral leishmaniasis is often difficult. Thus, identification of new chemotherapeutic agents is important for treatment of disease. Since targeting of the ergosterol synthesis pathway of Leishmania may be useful therapeutically, the aim of this study was to investigate the effect of alone or in combination of amiodarone and ketoconazole on Leishmania infantum.   Methods : To obtain logarithmic promastigotes of L. infantum, the parasites were cultured in BHI medium with FCS 10% together with antibiotics of penicillin and streptomycin and incubated at 24° C. Amastigote forms were obtained in BHI medium supplemented with 20% FCS at pH of 5.5 which incubated in 37° C. L.infantum susceptibility to amiodarone and ketoconazole was evaluated by proliferation of parasites in the absence or presence of these drugs with MTT assay. For evaluation of antiproliferative synergism against promastigotes and axenic amastigotes, fractional inhibitory concentrations (FIC were calculated. An isobologram curve was constructed too.   Results: Amiodarone produced a marked reduction in the viability of L.infantum promastigotes and axenic amastigotes. On the other hand ketoconazole induced a dose dependent effect on the parasites proliferation for promastigotes and axenic amastigotes. When the drugs were used in combination, the results indicated clear synergistic as shown by a concave isobologram and FIC value.   Conclusion: The present study represents the evidence that the combination of amiodarone plus ketoconazole acts synergistically in controlling L. infantume in vitro. It is possible that amiodarone could be used in combination with ketoconazole to combat infection at low doses, thus reducing its side

  10. Leishmania infantum Asparagine Synthetase A Is Dispensable for Parasites Survival and Infectivity.

    Science.gov (United States)

    Faria, Joana; Loureiro, Inês; Santarém, Nuno; Macedo-Ribeiro, Sandra; Tavares, Joana; Cordeiro-da-Silva, Anabela

    2016-01-01

    A growing interest in asparagine (Asn) metabolism has currently been observed in cancer and infection fields. Asparagine synthetase (AS) is responsible for the conversion of aspartate into Asn in an ATP-dependent manner, using ammonia or glutamine as a nitrogen source. There are two structurally distinct AS: the strictly ammonia dependent, type A, and the type B, which preferably uses glutamine. Absent in humans and present in trypanosomatids, AS-A was worthy of exploring as a potential drug target candidate. Appealingly, it was reported that AS-A was essential in Leishmania donovani, making it a promising drug target. In the work herein we demonstrate that Leishmania infantum AS-A, similarly to Trypanosoma spp. and L. donovani, is able to use both ammonia and glutamine as nitrogen donors. Moreover, we have successfully generated LiASA null mutants by targeted gene replacement in L. infantum, and these parasites do not display any significant growth or infectivity defect. Indeed, a severe impairment of in vitro growth was only observed when null mutants were cultured in asparagine limiting conditions. Altogether our results demonstrate that despite being important under asparagine limitation, LiAS-A is not essential for parasite survival, growth or infectivity in normal in vitro and in vivo conditions. Therefore we exclude AS-A as a suitable drug target against L. infantum parasites.

  11. Leishmania infantum Asparagine Synthetase A Is Dispensable for Parasites Survival and Infectivity.

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    Joana Faria

    2016-01-01

    Full Text Available A growing interest in asparagine (Asn metabolism has currently been observed in cancer and infection fields. Asparagine synthetase (AS is responsible for the conversion of aspartate into Asn in an ATP-dependent manner, using ammonia or glutamine as a nitrogen source. There are two structurally distinct AS: the strictly ammonia dependent, type A, and the type B, which preferably uses glutamine. Absent in humans and present in trypanosomatids, AS-A was worthy of exploring as a potential drug target candidate. Appealingly, it was reported that AS-A was essential in Leishmania donovani, making it a promising drug target. In the work herein we demonstrate that Leishmania infantum AS-A, similarly to Trypanosoma spp. and L. donovani, is able to use both ammonia and glutamine as nitrogen donors. Moreover, we have successfully generated LiASA null mutants by targeted gene replacement in L. infantum, and these parasites do not display any significant growth or infectivity defect. Indeed, a severe impairment of in vitro growth was only observed when null mutants were cultured in asparagine limiting conditions. Altogether our results demonstrate that despite being important under asparagine limitation, LiAS-A is not essential for parasite survival, growth or infectivity in normal in vitro and in vivo conditions. Therefore we exclude AS-A as a suitable drug target against L. infantum parasites.

  12. Association between Leishmania infantum DNA in the hair of dogs and their infectiousness to Lutzomyia longipalpis.

    Science.gov (United States)

    de Sousa Gonçalves, Rafaela; Franke, Carlos R; Magalhães-Junior, Jairo T; Souza, Bárbara M P S; Solcà, Manuela S; Larangeira, Daniela F; Barrouin-Melo, Stella Maria

    2016-12-15

    Diagnosis of infection with Leishmania infantum by DNA detection in the hair has been recently demonstrated in dogs and wild animals. Our objective was to investigate if polymerase chain reaction (PCR) in hair might be used to identify infectious dogs. Thus, we assessed the infectiousness to Lutzomyia longipalpis by xenodiagnosis in comparison with the detection of L. infantum DNA by PCR in the hair, and with serology for anti-Leishmania IgG by ELISA in 15 positive dogs for L. infantum infection. Eight healthy dogs were included as negative controls. Among the 15 infected dogs, 13 were found positive in the ELISA (87%), 12 were PCR positive in the hair (80%), and 10 were positive in xenodiagnosis (67%). Positivity in the hair was associated with positivity in spleen (p=0.0003), seropositivity for antibodies (p=0.0006) and parasite transmission to L. longipalpis (p=0.0028). Considering the benefits to animal welfare and feasibility of hair sampling method, studies in larger and more diverse populations of naturally infected dogs from endemic areas should be conducted to evaluate the sensitivity, specificity, and predictive values of PCR using hair as a possible biomarker of infectiousness in dogs. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. Immunopathological Features of Canine Myocarditis Associated with Leishmania infantum Infection

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    Alessandro Costagliola

    2016-01-01

    Full Text Available Myocarditis associated with infectious diseases may occur in dogs, including those caused by the protozoa Neospora caninum, Trypanosoma cruzi, Babesia canis, and Hepatozoon canis. However, although cardiac disease due to Leishmania infection has also been documented, the immunopathological features of myocarditis have not been reported so far. The aim of this study was to examine the types of cellular infiltrates and expression of MHC classes I and II in myocardial samples obtained at necropsy from 15 dogs with an established intravitam diagnosis of visceral leishmaniasis. Pathological features of myocardium were characterized by hyaline degeneration of cardiomyocytes, necrosis, and infiltration of mononuclear inflammatory cells consisting of lymphocytes and macrophages, sometimes with perivascular pattern; fibrosis was also present in various degrees. Immunophenotyping of inflammatory cells was performed by immunohistochemistry on cryostat sections obtained from the heart of the infected dogs. The predominant leukocyte population was CD8+ with a fewer number of CD4+ cells. Many cardiomyocytes expressed MHC classes I and II on the sarcolemma. Leishmania amastigote forms were not detected within macrophages or any other cell of the examined samples. Our study provided evidence that myocarditis in canine visceral leishmaniasis might be related to immunological alterations associated with Leishmania infection.

  14. Mitochondrial Proteomics of Antimony and Miltefosine Resistant Leishmania infantum

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    Isabel M. Vincent

    2015-10-01

    Full Text Available Antimony (SbIII and miltefosine (MIL are important drugs for the treatment of Leishmania parasite infections. The mitochondrion is likely to play a central role in SbIII and MIL induced cell death in this parasite. Enriched mitochondrial samples from Leishmania promastigotes selected step by step for in vitro resistance to SbIII and MIL were subjected to differential proteomic analysis. A shared decrease in both mutants in the levels of pyruvate dehydrogenase, dihydrolipoamide dehydrogenase, and isocitrate dehydrogenase was observed, as well as a differential abundance in two calcium-binding proteins and the unique dynamin-1-like protein of the parasite. Both mutants presented a shared increase in the succinyl-CoA:3-ketoacid-coenzyme A transferase and the abundance of numerous hypothetical proteins was also altered in both mutants. In general, the proteomic changes observed in the MIL mutant were less pronounced than in the SbIII mutant, probably due to the early appearance of a mutation in the miltefosine transporter abrogating the need for a strong mitochondrial adaptation. This study is the first analysis of the Leishmania mitochondrial proteome and offers powerful insights into the adaptations to this organelle during SbIII and MIL drug resistance.

  15. Natural hybrid of Leishmania infantum/L. donovani: development in Phlebotomus tobbi, P. perniciosus and Lutzomyia longipalpis and comparison with non-hybrid strains differing in tissue tropism.

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    Seblova, Veronika; Myskova, Jitka; Hlavacova, Jana; Votypka, Jan; Antoniou, Maria; Volf, Petr

    2015-11-25

    Infection caused by parasites from L. donovani complex can manifest as a serious visceral disease or a self-healing milder cutaneous form. The different tropism and pathology in humans is caused by the interaction between parasites, host and vector determinants but the mechanisms are not well understood. In Cukurova region in Turkey we previously identified a major focus of cutaneous leishmaniasis caused by L. donovani/infantum hybrids (CUK strain) and isolated this parasite from the locally abundant sand fly, Phlebotomus tobbi. Here, we present the first experimental study with P. tobbi. We tested the susceptibility of this species to various Leishmania under laboratory conditions, characterized glycoproteins in the P. tobbi midgut putatively involved in parasite-vector interaction and compared the development of the CUK strain in the sand fly with one other dermotropic and three viscerotropic strains belonging to the L. donovani complex. Females of laboratory reared P. tobbi, P. perniciosus and Lutzomyia longipalpis were infected using membrane feeding on rabbit blood containing promastigotes of various Leishmania species with different tropisms. The individual guts were checked microscopically for presence and localization of Leishmania parasites; the number of parasites was assessed more precisely by qPCR. In addition, glycosylation of midgut proteins of P. tobbi was studied by lectin blotting of midgut lysate with lectins specific for terminal sugars of N-type and O-type glycans. High infection rates, heavy parasite loads and late-stage infection with colonization of the stomodeal valve were observed in P. tobbi infected by Leishmania major or L. infantum CUK hybrid. In parallel, lectin blotting revealed the presence of O-glycosylated proteins in the P. tobbi midgut. In P. perniciosus and L. longipalpis all five Leishmania strains tested developed well. In both vectors, significantly higher parasite numbers were detected by qPCR for dermotropic L. donovani

  16. Purification, crystallization and preliminary X-ray diffraction analysis of the glyoxalase II from Leishmania infantum

    International Nuclear Information System (INIS)

    Trincão, José; Sousa Silva, Marta; Barata, Lídia; Bonifácio, Cecília; Carvalho, Sandra; Tomás, Ana Maria; Ferreira, António E. N.; Cordeiro, Carlos; Ponces Freire, Ana; Romão, Maria João

    2006-01-01

    A glyoxalase II from L. infantum was cloned, purified and crystallized and its structure was solved by X-ray crystallography. In trypanosomatids, trypanothione replaces glutathione in all glutathione-dependent processes. Of the two enzymes involved in the glyoxalase pathway, glyoxalase I and glyoxalase II, the latter shows absolute specificity towards trypanothione thioester, making this enzyme an excellent model to understand the molecular basis of trypanothione binding. Cloned glyoxalase II from Leishmania infantum was overexpressed in Escherichia coli, purified and crystallized. Crystals belong to space group C222 1 (unit-cell parameters a = 65.6, b = 88.3, c = 85.2 Å) and diffract beyond 2.15 Å using synchrotron radiation. The structure was solved by molecular replacement using the human glyoxalase II structure as a search model. These results, together with future detailed kinetic characterization using lactoyltrypanothione, should shed light on the evolutionary selection of trypanothione instead of glutathione by trypano-somatids

  17. Cytokine and Phenotypic Cell Profiles of Leishmania infantum Infection in the Dog

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    Carla Maia

    2012-01-01

    Full Text Available Leishmaniasis has reemerged in recent years showing a wider geographic distribution and increased global incidence of human and canine disease than previously known. Dogs are the main domestic/peridomestic reservoir hosts of zoonotic visceral leishmaniasis caused by Leishmania infantum. Since the evolution of leishmaniasis and clinical appearance is a consequence of complex interactions between the parasite and host immune response, a profound knowledge about the immune profile developed in dog's infection is crucial for vaccine and immunomodulatory therapy design. The main goal of this paper is to compile the recent advances made on cytokine and phenotypic cell profiles in different tissues and organs of dogs infected with L. infantum. This paper also stressed that the knowledge of the immune responses developed, namely, in liver, lymph node, and spleen is very limited. All data emphasizes that more research on canine leishmaniasis is necessary for the development of new and efficacious tools to control zoonotic leishmaniasis.

  18. Detection of antibodies against Leishmania infantum in cats (Felis catus from the State of Pernambuco, Brazil

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    Rita de Cássia Nascimento Silva

    2014-01-01

    Full Text Available Introduction: Little information is available concerning infection by Leishmania infantum in cats. Therefore, the aim of this study was to perform a serological study in domestic cats. Methods: Serum samples (n=153 obtained from animals living in the Cities of Recife and Petrolina, State of Pernambuco, Brazil, were tested by ELISA/S7® (Biogene. Results: Anti-L. infantum antibodies were detected in 3.9% (6/153 of the cats. All seroreagent animals were from Petrolina. Conclusions: These results serve as an important alert, and future studies are needed to better understand the possible role of cats in the epidemiology of visceral leishmaniasis (VL in this area.

  19. Protection mediated by chemokine CXCL10 in BALB/c mice infected by Leishmania infantum

    Science.gov (United States)

    Figueiredo, Webertty Mayk Eufrásio; Viana, Sayonara de Melo; Alves, Dorotheia Teixeira; Guerra, Priscila Valera; Coêlho, Zirlane Castelo Branco; Barbosa, Helene Santos; Teixeira, Maria Jania

    2017-01-01

    BACKGROUND Visceral leishmaniasis (VL) caused by Leishmania infantum is characterised by the loss of the ability of the host to generate an effective immune response. Chemokines have a direct involvement in the pathogenesis of leishmaniasis, causing a rapid change in the expression of these molecules during infection by Leishmania. OBJECTIVES Herein, it was investigated the role of CXCL10 in controlling infection by L. infantum. METHODS RAW 264.7 macrophages were infected with L. infantum in vitro and treated or not with CXCL10 (25, 50 and 100 ng/mL). Parasite load, as well as nitric oxide (NO), IL-4 and IL-10 production were assessed at 24 and 48 h after infection. In vivo, BALB/c mice were infected and treated or not with CXCL10 (5 μg/kg) at one, three and seven days of infection. Parasite load, IFN-g, IL-4, TGF-β and IL-10 were evaluated one, seven and 23 days post treatment. FINDINGS In vitro, CXCL10 reduced parasitic load, not dependent on NO, and inhibited IL-10 and IL-4 secretion. In vivo, CXCL10 was able to reduce the parasite load in both liver and spleen, four weeks after infection, representing a higher decrease in the number of parasites in these organs, also induced IFN-γ at day 23 after treatment, correlating with the decrease in parasite load, and reduced IL-10 and TGF-β. MAIN CONCLUSIONS This study suggests a partial protective role of CXCL10 against L. infantum, mediated by IFN-g, not dependent on NO, and with suppression of IL-10 and TGF-β. These data may provide information for the development of new approaches for future therapeutic interventions for VL. PMID:28767981

  20. Leishmania infantum, Dirofilaria spp. and other endoparasite infections in kennel dogs in central Italy

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    Sauda, Federica; Malandrucco, Livia; Macrì, Gladia; Scarpulla, Manuela; De Liberato, Claudio; Terracciano, Giuliana; Fichi, Gianluca; Berrilli, Federica; Perrucci, Stefania

    2018-01-01

    Prevalence and risk factors of Leishmania infantum, Dirofilaria spp. and other potentially zoonotic or canine-specific endoparasite infections were assessed in 639 kennel dogs from central Italy. To this end, individual blood and fecal samples were examined using parasitological, immunological and molecular techniques. The presence of compatible clinical pictures, as well as age and gender were considered as putative risks factors. To evaluate risk factors, multivariable analysis with logistic regression and univariable analysis with a Chi square test and a Fischer’s exact test were performed. Overall, 52.6% of dogs (95% CI 48.6-56.5) were found positive, while 39.6% of dogs (95% CI 35.8-43.5) were infected by potentially zoonotic species. Leishmania infantum and Dirofilaria repens showed prevalences of 2.5% (95% CI 1.5-4.1) and 2.8% (95% CI 1.7-4.5), respectively. The prevalence of cardiorespiratory parasites was 7.8% (95% CI 5.9-10.3) and included the species Angiostrongylus vasorum, Eucoleus aerophilus, Eucoleus boehmi and D. immitis; the latter showed a prevalence of 0.2% (95% CI 0.001-1). Intestinal parasites were significantly prevalent (38.8%, 95% CI 35-42.7) and they consisted mainly of species of major zoonotic concern, including ancylostomatids, Toxocara canis, Giardia duodenalis, Dipylidium caninum, Taeniidae, Strongyloides stercoralis and Cryptosporidium parvum. Endoparasites were significantly prevalent in clinically suspected dogs. Leishmania infantum and cardiorespiratory nematodes were prevalent in older dogs, while intestinal parasites were prevalent in younger dogs. Results show high dog and public health risks in kennels in central Italy, and suggest the need for more effective control measures. PMID:29388550

  1. Leishmania infantum, Dirofilaria spp. and other endoparasite infections in kennel dogs in central Italy

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    Sauda Federica

    2018-01-01

    Full Text Available Prevalence and risk factors of Leishmania infantum, Dirofilaria spp. and other potentially zoonotic or canine-specific endoparasite infections were assessed in 639 kennel dogs from central Italy. To this end, individual blood and fecal samples were examined using parasitological, immunological and molecular techniques. The presence of compatible clinical pictures, as well as age and gender were considered as putative risks factors. To evaluate risk factors, multivariable analysis with logistic regression and univariable analysis with a Chi square test and a Fischer’s exact test were performed. Overall, 52.6% of dogs (95% CI 48.6-56.5 were found positive, while 39.6% of dogs (95% CI 35.8-43.5 were infected by potentially zoonotic species. Leishmania infantum and Dirofilaria repens showed prevalences of 2.5% (95% CI 1.5-4.1 and 2.8% (95% CI 1.7-4.5, respectively. The prevalence of cardiorespiratory parasites was 7.8% (95% CI 5.9-10.3 and included the species Angiostrongylus vasorum, Eucoleus aerophilus, Eucoleus boehmi and D. immitis; the latter showed a prevalence of 0.2% (95% CI 0.001-1. Intestinal parasites were significantly prevalent (38.8%, 95% CI 35-42.7 and they consisted mainly of species of major zoonotic concern, including ancylostomatids, Toxocara canis, Giardia duodenalis, Dipylidium caninum, Taeniidae, Strongyloides stercoralis and Cryptosporidium parvum. Endoparasites were significantly prevalent in clinically suspected dogs. Leishmania infantum and cardiorespiratory nematodes were prevalent in older dogs, while intestinal parasites were prevalent in younger dogs. Results show high dog and public health risks in kennels in central Italy, and suggest the need for more effective control measures.

  2. Quantification of Leishmania infantum DNA in the bone marrow, lymph node and spleen of dogs.

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    Ramos, Rafael Antonio Nascimento; Ramos, Carlos Alberto do Nascimento; Santos, Edna Michelly de Sá; de Araújo, Flábio Ribeiro; de Carvalho, Gílcia Aparecida; Faustino, Maria Aparecida da Gloria; Alves, Leucio Câmara

    2013-01-01

    The aim of the present study was to quantify the parasite load of Leishmania infantum in dogs using real-time PCR (qPCR). Bone marrow, lymph node and spleen samples were taken from 24 dogs serologically positive for L. infantum that had been put down by the official epidemiological surveillance service. According to the clinical signs the dogs were classified as asymptomatic or symptomatic. After DNA extraction, the samples were subjected to qPCR to detect and quantify L. infantum DNA. Out of the 24 dogs, 12.5% (3/24) were classified as asymptomatic and 87.5% (21/24) as symptomatic. Real-time PCR detected L. infantum DNA in all the animals, in at least one biological sample. In particular, 100% of bone marrow and lymph node scored positive, whereas in spleen, the presence of DNA was detected in 95.9% (23/24). In addition, out of 24 animals, 15 were microscopically positive to amastigote forms of L. infantum in bone marrow. No statistical significant difference was found in the overall mean quantity of DNA among the different biological samples (P = 0.518). Considering each organ separately, there was 100% positivity in bone marrow and lymph nodes, while among the spleen samples, 95.9% (23/24) were positive. Regarding the different clinical groups, the overall mean parasite load varied significantly (P = 0.022). According to the results obtained, it was not possible determine which biological sample was most suitable tissue for the diagnosis, based only on the parasite load. Therefore, other characteristics such as convenience and easily of obtaining samples should be taken into consideration.

  3. Experimental infection and transmission of Leishmania by Lutzomyia cruzi (Diptera: Psychodidae: Aspects of the ecology of parasite-vector interactions.

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    Everton Falcão de Oliveira

    2017-02-01

    Full Text Available Several parameters should be addressed before incriminating a vector for Leishmania transmission. Those may include its ability to become infected by the same Leishmania species found in humans, the degree of attractiveness for reservoirs and humans and capacity to sustain parasite infection under laboratory conditions. This study evaluated the vectorial capacity of Lutzomyia cruzi for Leishmania infantum and gathered information on its ability to harbor L. amazonensis. Laboratory-reared Lu. cruzi were infected experimentally by feeding them on dogs infected naturally with L. infantum and hamsters infected with L. amazonensis. Sand fly attractiveness to dogs and humans was determined using wild caught insects. The expected daily survival of infected Lu. cruzi, the duration of the gonotrophic cycle, and the extrinsic incubation period were also investigated for both parasites. Vector competence was investigated for both Leishmania species. The mean proportion of female sand flies that fed on hosts was 0.40. For L. infantum and L. amazonensis, Lu. cruzi had experimental infection rates of 10.55% and 41.56%, respectively. The extrinsic incubation period was 3 days for both Leishmania species, regardless of the host. Survival expectancy of females infected with L. infantum and L. amazonensis after completing the gonotrophic cycle was 1.32 and 0.43, respectively. There was no association between L. infantum infection and sand fly longevity, but L. amazonensis-infected flies had significantly greater survival probabilities. Furthermore, egg-laying was significantly detrimental to survival. Lu. cruzi was found to be highly attracted to both dogs and humans. After a bloodmeal on experimentally infected hosts, both parasites were able to survive and develop late-stage infections in Lu. cruzi. However, transmission was demonstrated only for L. amazonensis-infected sand flies. In conclusion, Lu. cruzi fulfilled several of the requirements of vectorial

  4. Experimental infection and transmission of Leishmania by Lutzomyia cruzi (Diptera: Psychodidae): Aspects of the ecology of parasite-vector interactions.

    Science.gov (United States)

    Falcão de Oliveira, Everton; Oshiro, Elisa Teruya; Fernandes, Wagner de Souza; Murat, Paula Guerra; Medeiros, Márcio José de; Souza, Alda Izabel; Oliveira, Alessandra Gutierrez de; Galati, Eunice Aparecida Bianchi

    2017-02-01

    Several parameters should be addressed before incriminating a vector for Leishmania transmission. Those may include its ability to become infected by the same Leishmania species found in humans, the degree of attractiveness for reservoirs and humans and capacity to sustain parasite infection under laboratory conditions. This study evaluated the vectorial capacity of Lutzomyia cruzi for Leishmania infantum and gathered information on its ability to harbor L. amazonensis. Laboratory-reared Lu. cruzi were infected experimentally by feeding them on dogs infected naturally with L. infantum and hamsters infected with L. amazonensis. Sand fly attractiveness to dogs and humans was determined using wild caught insects. The expected daily survival of infected Lu. cruzi, the duration of the gonotrophic cycle, and the extrinsic incubation period were also investigated for both parasites. Vector competence was investigated for both Leishmania species. The mean proportion of female sand flies that fed on hosts was 0.40. For L. infantum and L. amazonensis, Lu. cruzi had experimental infection rates of 10.55% and 41.56%, respectively. The extrinsic incubation period was 3 days for both Leishmania species, regardless of the host. Survival expectancy of females infected with L. infantum and L. amazonensis after completing the gonotrophic cycle was 1.32 and 0.43, respectively. There was no association between L. infantum infection and sand fly longevity, but L. amazonensis-infected flies had significantly greater survival probabilities. Furthermore, egg-laying was significantly detrimental to survival. Lu. cruzi was found to be highly attracted to both dogs and humans. After a bloodmeal on experimentally infected hosts, both parasites were able to survive and develop late-stage infections in Lu. cruzi. However, transmission was demonstrated only for L. amazonensis-infected sand flies. In conclusion, Lu. cruzi fulfilled several of the requirements of vectorial capacity for L. infantum

  5. Leishmania infantum infection in dogs from maroon communities in the Eastern Amazon

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    Andréia Ferreira da Silva

    Full Text Available ABSTRACT: This study was designed to detect L. infantum infection in dogs and to evaluate the factors associated with canine visceral leishmaniasis in the maroon communities of Menino Jesus de Petimandeua and Itaboca in the municipality of Inhangapi, Pará, Brazil. Whole blood and intact skin samples were collected from 143 dogs, and a questionnaire was applied. L. infantum DNA was detected by polymerase chain reaction (PCR using primers RV1 and RV2. Collection sites were georeferenced to obtain a spatial distribution of the residences visited and infected dogs. L. infantum DNA was detected in 8.4% (12/143 of the skin samples and in 1.4% (2/143 of the blood samples. On the risk map, three clusters were observed in Itaboca and one was observed in Menino Jesus de Petimandeua. We observed that most of the inhabitants in these maroon communities live close to forested areas and do not use protection against insect vectors. The presence of canine reservoirs of L. infantum associated to environment characteristics (preserved forests and deforested areas and habits of dog owners (living near forested areas and not using any protection against insects may favor the transmission of L. infantum in the studied areas.

  6. First insights into the genetic diversity and origin of Leishmania infantum in Mont Rolland (Thiès region, Senegal).

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    Cassan, Cecile; Dione, Michel Mainack; Dereure, Jacques; Diedhiou, Souleymane; Bucheton, Bruno; Hide, Mallorie; Kako, Caroline; Gaye, Oumar; Senghor, Massila; Niang, Abdoul Aziz; Bañuls, Anne-Laure; Faye, Babacar

    2016-06-01

    Visceral leishmaniasis is not endemic in West Africa. However, high seroprevalence of Leishmania infantum infection (one of the Leishmania species that cause visceral leishmaniasis) was detected in dogs and humans in the Mont Rolland community (close to Thiès, Senegal), despite the lack of reports concerning human clinical cases. Our aim was to genetically characterize this L. infantum population and identify its origin. We thus conducted seven field surveys in 25 villages of the Mont Rolland community between 2005 and 2009 and blood samples were collected from 205 dogs. Serological testing indicated that 92 dogs (44.9%) were positive for Leishmania infection. L. infantum was identified as the cause of infection. Analysis of 29 L. infantum isolates from these dogs by multilocus microsatellite typing and multilocus sequence typing indicated that this population had very limited genetic diversity, low level of heterozygosity and only seven different genotypes (79.3% of all isolates had the same genotype). Multilocus sequence typing showed that the Mont Rolland isolates clustered with strains from the Mediterranean basin and were separated from East African and Asian strains. Therefore, our data suggest a quite recent and unique introduction into Senegal of a L. infantum strain from the Mediterranean basin. Copyright © 2016 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  7. Evidence of Leishmania infantum Infection in Rabbits (Oryctolagus cuniculus in a Natural Area in Madrid, Spain

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    Nerea García

    2014-01-01

    Full Text Available Leishmaniasis is one of the most important neglected zoonosis and remains endemic in at least 88 developing countries in the world. In addition, anthropogenic environmental changes in urban areas are leading to its emergency world wide. Zoonotic leishmaniasis control might only be achieved by an integrated approach targeting both the human host and the animal reservoirs, which in certain sylvatic cycles are yet to be identified. Recently, hares have been pointed out as competent reservoirs of Leishmania infantum in Spain, but the role of other lagomorphs has not been clarified. Here, 69 rabbits (Oryctolagus cuniculus from a natural area in Madrid in which a high density was present were analyzed using indirect (immunofluorescence antibody test, IFAT and direct (PCR, culture techniques. Fifty-seven (82.6% of the animals were positive to at least one technique, with IFAT yielding the highest proportion of positive samples. L. infantum was isolated in 13% animals demonstrating the occurrence of infection in this setting. Our results suggest that rabbits could play a role of competent reservoir of L. infantum and demonstrate that the prevalence of infection is high in the analyzed area.

  8. Seroprevalence rates of antibodies againstLeishmania infantum and other protozoan and rickettsial parasites in dogs

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    Silvana de Cássia Paulan

    Full Text Available Canine visceral leishmaniasis (CVL is caused by the protozoan Leishmania infantum, which infects dogs and humans in many regions of Brazil. The present study involved an indirect fluorescent antibody test (IFAT to analyze L. infantum,Ehrlichia spp., Babesia canis,Toxoplasma gondii and Neospora caninuminfection rates in serum samples from 93 dogs in a rural settlement in Ilha Solteira, SP, Brazil. The seroprevalence rates of anti-L. infantum, anti-Ehrlichia, anti-B. canis, anti-T. gondii and anti-N. caninum antibodies were 37.6%, 75.3%, 72%, 47.3% and 6.4%, respectively. In addition to IFAT, direct microscopic examination of popliteal lymph node aspirates revealed 26.9% of CVL positive dogs. Serological tests revealed that 17.2% of the dogs were seropositive for a single parasite, 29% for two parasites, 33% for three, 16.1% for four, and 1.1% for five parasites, while 3.2% were seronegative for five parasites. The presence of antibodies against these parasites in serum samples from dogs confirmed their exposure to these parasites in this rural area. Because of the potential zoonotic risk of these diseases, mainly leishmaniasis, ehrlichiosis and toxoplasmosis, special attention should focus on programs for the improvement of diagnostic assays and control measures against these parasites.

  9. Ultrastructural Analysis of Leishmania infantum chagasi Promastigotes Forms Treated In Vitro with Usnic Acid

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    João S. B. da Luz

    2015-01-01

    Full Text Available Leishmaniasis is considered by the World Health Organization as one of the infectious parasitic diseases endemic of great relevance and a global public health problem. Pentavalent antimonials used for treatment of this disease are limited and new phytochemicals emerge as an alternative to existing treatments, due to the low toxicity and cost reduction. Usnic acid is uniquely found in lichens and is especially abundant in genera such as Alectoria, Cladonia, Evernia, Lecanora, Ramalina, and Usnea. Usnic acid has been shown to exhibit antiviral, antiprotozoal, antiproliferative, anti-inflammatory, and analgesic activity. The aim of this study was to evaluate the antileishmanial activity of usnic acid on Leishmania infantum chagasi promastigotes and the occurrence of drug-induced ultrastructural damage in the parasite. Usnic acid was effective against the promastigote forms (IC50 = 18.30 ± 2.00 µg/mL. Structural and ultrastructural aspects of parasite were analyzed. Morphological alterations were observed as blebs in cell membrane and shapes given off, increasing the number of cytoplasmic vacuoles, and cellular and mitochondrial swelling, with loss of cell polarity. We concluded that the usnic acid presented antileishmanial activity against promastigote forms of Leishmania infantum chagasi and structural and ultrastructural analysis reinforces its cytotoxicity. Further, in vitro studies are warranted to further evaluate this potential.

  10. A serological study of Leishmania infantum in dogs of Khorasan Razavi province, Iran.

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    Sabzevari, S; Razmi, G R; Naghibi, A; Khoshnegah, J

    2013-10-01

    Leishmania infantum is the principal agent of visceral leishmaniosis (kala-azar) in all areas of Iran. The disease is endemic in Fars, Ardabil, East Azerbaijan and Bushehr provinces while it is sporadic in other parts of the country. Dog as a member of canidae family is the main animal reservoir host of kala-azar. The present study was carried out to determine the sero-prevalence of canine leishmaniosis in Mashhad, the capital city of Khorasan Razavi Province, in the north east of Iran. Two-hundred and nine dogs in two different classes including stray and sheepdogs were selected for this cross-sectional study. The blood samples were collected and tested by indirect immunofluorescent antibody test for detection of anti-Leishmania infantum antibodies. The results showed that 16/209 (7.6 %) of blood samples were positive. The sero-prevalence rates in stray and sheepdogs were 11/94 (11.7 %) and 5/115 (4.3 %), respectively (p < 0.05). Based on the results obtained, it is concluded that visceral leishmaniosis is distributed among dogs of this area, also sero-prevalence of canine leishmaniosis in stray dogs higher than sheepdogs.

  11. SYBR green-based detection of Leishmania infantum DNA using peripheral blood samples.

    Science.gov (United States)

    Ghasemian, Mehrdad; Gharavi, Mohammad Javad; Akhlaghi, Lame; Mohebali, Mehdi; Meamar, Ahmad Reza; Aryan, Ehsan; Oormazdi, Hormozd; Ghayour, Zahra

    2016-03-01

    Parasitological methods for the diagnosis of visceral leishmaniasis (VL) require invasive sampling procedures. The aim of this study was to detect Leishmania infantum (L. infantum) DNA by real time-PCR method in peripheral blood of symptomatic VL patient and compared its performance with nested PCR, an established molecular method with very high diagnostic indices. 47 parasitologically confirmed VL patients diagnosed by direct agglutination test (DAT > 3200), bone marrow aspiration and presented characteristic clinical features (fever, hepatosplenomegaly, and anemia) and 40 controls (non-endemic healthy control-30, Malaria-2, Toxoplasma gondii-2, Mycobacterium tuberculosis-2, HBV-1, HCV-1, HSV-1 and CMV-1) were enrolled in this study. SYBR-green based real time-PCR and nested PCR was performed to amplify the Kinetoplast DNA minicircle gene using the DNA extracted from Buffy coat. From among 47 patients, 45 (95.7 %) were positive by both nested-PCR and real time-PCR. These results indicate that real time-PCR was not only as sensitive as a nested-PCR assay for detection of Leishmania kDNA in clinical sample, but also more rapid. The advantage of real time-PCR based methods over nested-PCR is simple to perform, more faster in which nested-PCR requires post-PCR processing and reducing contamination risk.

  12. Ability of immunodiagnostic tests to differentiate between dogs naturally infected with Leishmania infantum and Leishmune(®)-vaccinated dogs

    NARCIS (Netherlands)

    Ribeiro, R. A. N.; Teixeira-Neto, R. G.; Belo, V. S.; Ferreira, E. C.; Schallig, H. D. F. H.; Silva, E. S.

    2015-01-01

    Visceral leishmaniasis (VL) is a serious chronic disease with a lethality rate of up to 10 % in humans. In urban areas of Brazil, dogs are the main reservoirs of the etiological agent (Leishmania infantum) of VL, and the Brazilian Ministry of Health recommends the euthanasia of animals that are

  13. Leishmania infantum Exoproducts Inhibit Human Invariant NKT Cell Expansion and Activation

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    Renata Belo

    2017-06-01

    Full Text Available Leishmania infantum is one of the major parasite species associated with visceral leishmaniasis, a severe form of the disease that can become lethal if untreated. This obligate intracellular parasite has developed diverse strategies to escape the host immune response, such as exoproducts (Exo carrying a wide range of molecules, including parasite virulence factors, which are potentially implicated in early stages of infection. Herein, we report that L. infantum Exo and its two fractions composed of extracellular vesicles (EVs and vesicle-depleted-exoproducts (VDEs inhibit human peripheral blood invariant natural killer T (iNKT cell expansion in response to their specific ligand, the glycolipid α-GalactosylCeramide (α-GalCer, as well as their capacity to promptly produce IL-4 and IFNγ. Using plate-bound CD1d and α-GalCer, we found that Exo, EV, and VDE fractions reduced iNKT cell activation in a dose-dependent manner, suggesting that they prevented α-GalCer presentation by CD1d molecules. This direct effect on CD1d was confirmed by the observation that CD1d:α-GalCer complex formation was impaired in the presence of Exo, EV, and VDE fractions. Furthermore, lipid extracts from the three compounds mimicked the inhibition of iNKT cell activation. These lipid components of L. infantum exoproducts, including EV and VDE fractions, might compete for CD1-binding sites, thus blocking iNKT cell activation. Overall, our results provide evidence for a novel strategy through which L. infantum can evade immune responses of mammalian host cells by preventing iNKT lymphocytes from recognizing glycolipids in a TCR-dependent manner.

  14. First occurrence of an autochthonous canine case of Leishmania (Leishmania infantum chagasi in the municipality of Campinas, State of São Paulo, Brazil

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    Elisa San Martin Mouriz Savani

    2011-08-01

    Full Text Available An autochthonous case of visceral leishmaniasis is reported in a dog (Canis familiaris as an apparently natural infection in a non-endemic area. DNA obtained from spleen and liver samples produced the expected fragment in a Leishmania-specific rDNA-based nested-PCR assay. The PCR product, a 490 bp fragment, was sequenced and the nucleotide sequence was identical to that of Leishmania (Leishmania infantum chagasi. These results are surprising since no autochthonous human or canine cases of visceral leishmaniasis have ever been reported in this municipality. This case suggests that natural transmission of this disease is occurring in this area.

  15. Clinical and parasitological protection in a Leishmania infantum-macaque model vaccinated with adenovirus and the recombinant A2 antigen.

    Science.gov (United States)

    Grimaldi, Gabriel; Teva, Antonio; Porrozzi, Renato; Pinto, Marcelo A; Marchevsky, Renato S; Rocha, Maria Gabrielle L; Dutra, Miriam S; Bruña-Romero, Oscar; Fernandes, Ana-Paula; Gazzinelli, Ricardo T

    2014-06-01

    Visceral leishmaniasis (VL) is a severe vector-born disease of humans and dogs caused by Leishmania donovani complex parasites. Approximately 0.2 to 0.4 million new human VL cases occur annually worldwide. In the new world, these alarming numbers are primarily due to the impracticality of current control methods based on vector reduction and dog euthanasia. Thus, a prophylactic vaccine appears to be essential for VL control. The current efforts to develop an efficacious vaccine include the use of animal models that are as close to human VL. We have previously reported a L. infantum-macaque infection model that is reliable to determine which vaccine candidates are most worthy for further development. Among the few amastigote antigens tested so far, one of specific interest is the recombinant A2 (rA2) protein that protects against experimental L. infantum infections in mice and dogs. Primates were vaccinated using three rA2-based prime-boost immunization regimes: three doses of rA2 plus recombinant human interleukin-12 (rhIL-12) adsorbed in alum (rA2/rhIL-12/alum); two doses of non-replicative adenovirus recombinant vector encoding A2 (Ad5-A2) followed by two boosts with rA2/rhIL-12/alum (Ad5-A2+rA2/rhIL12/alum); and plasmid DNA encoding A2 gene (DNA-A2) boosted with two doses of Ad5-A2 (DNA-A2+Ad5-A2). Primates received a subsequent infectious challenge with L. infantum. Vaccines, apart from being safe, were immunogenic as animals responded with increased pre-challenge production of anti-A2-specific IgG antibodies, though with some variability in the response, depending on the vaccine formulation/protocol. The relative parasite load in the liver was significantly lower in immunized macaques as compared to controls. Protection correlated with hepatic granuloma resolution, and reduction of clinical symptoms, particularly when primates were vaccinated with the Ad5-A2+rA2/rhIL12/alum protocol. The remarkable clinical protection induced by A2 in an animal model that is

  16. Clinical and parasitological protection in a Leishmania infantum-macaque model vaccinated with adenovirus and the recombinant A2 antigen.

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    Gabriel Grimaldi

    2014-06-01

    Full Text Available BACKGROUND: Visceral leishmaniasis (VL is a severe vector-born disease of humans and dogs caused by Leishmania donovani complex parasites. Approximately 0.2 to 0.4 million new human VL cases occur annually worldwide. In the new world, these alarming numbers are primarily due to the impracticality of current control methods based on vector reduction and dog euthanasia. Thus, a prophylactic vaccine appears to be essential for VL control. The current efforts to develop an efficacious vaccine include the use of animal models that are as close to human VL. We have previously reported a L. infantum-macaque infection model that is reliable to determine which vaccine candidates are most worthy for further development. Among the few amastigote antigens tested so far, one of specific interest is the recombinant A2 (rA2 protein that protects against experimental L. infantum infections in mice and dogs. METHODOLOGY/PRINCIPAL FINDINGS: Primates were vaccinated using three rA2-based prime-boost immunization regimes: three doses of rA2 plus recombinant human interleukin-12 (rhIL-12 adsorbed in alum (rA2/rhIL-12/alum; two doses of non-replicative adenovirus recombinant vector encoding A2 (Ad5-A2 followed by two boosts with rA2/rhIL-12/alum (Ad5-A2+rA2/rhIL12/alum; and plasmid DNA encoding A2 gene (DNA-A2 boosted with two doses of Ad5-A2 (DNA-A2+Ad5-A2. Primates received a subsequent infectious challenge with L. infantum. Vaccines, apart from being safe, were immunogenic as animals responded with increased pre-challenge production of anti-A2-specific IgG antibodies, though with some variability in the response, depending on the vaccine formulation/protocol. The relative parasite load in the liver was significantly lower in immunized macaques as compared to controls. Protection correlated with hepatic granuloma resolution, and reduction of clinical symptoms, particularly when primates were vaccinated with the Ad5-A2+rA2/rhIL12/alum protocol. CONCLUSIONS

  17. [Ecoepidemiology of leishmaniases in Syria. 2--Presence, in dogs, of Leishmania infantum Nicolle and Leishmania tropica (Wright) (Kinetoplastida-Trypanonomatidae)].

    Science.gov (United States)

    Dereure, J; Rioux, J A; Khiami, A; Pratlong, F; Périères, J; Martini, A

    1991-01-01

    In an ecoepidemiological study in the Syrian foci of human visceral (VL) and cutaneous (CL) leishmaniasis, domestic dogs infected with Leishmania infantum MON-1 and L. tropica MON-76 were found for the first time. In Syria canine leishmaniasis caused by L. infantum, occurs in the humid and sub-humid western belt from the coastal zone to the nearby mountain ranges. Sporadis cases of human VL occur in this area. Canine CL, caused by L. tropica was found in the semi-arid zone in a village with a high human infection rate. The infection causes small papules or crusted ulcers on the nose or muzzle. These observations are relevant to control. In human VL foci it is recommended not to control the dog population, unless this is linked with other activities, such as rabies control. Control of dogs in the CL areas is not justified because of the small size of the lesions and the rarity of the parasites. Indeed the role of the dog as the "true" reservoir host is questionable. Human CL infection is best controlled by active case detection and specific treatment.

  18. Histopathological and parasitological study of the gastrointestinal tract of dogs naturally infected with Leishmania infantum

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    Pinto Aldair JW

    2011-12-01

    Full Text Available Abstract Background The aim of this study was to provide a systematic pathological and parasitological overview of the gastrointestinal tract (GIT, including the stomach, duodenum, jejunum, ileum, caecum and colon, of dogs naturally infected with Leishmania. Methods Twenty mongrel dogs naturally infected with Leishmania (Leishmania infantum and obtained from the Control Zoonosis Center of the Municipality of Ribeirão das Neves, Belo Horizonte Metropolitan area, Minas Gerais (MG state, Brazil, were analyzed. The dogs were divided into two groups: Group 1 comprised nine clinically normal dogs and group 2 comprised 11 clinically affected dogs. After necropsy, one sample was collected from each GIT segment, namely the stomach, duodenum, jejunum, ileum, caecum and colon. Furthermore, paraffin-embedded samples were used for histological and parasitological (immunohistochemistry evaluation and a morphometrical study were carried out to determine the parasite load (immunolabeled amastigote forms of Leishmania. The Friedman and the Mann Whitney tests were used for statistical analysis. The Friedman test was used to analyze each segment of the GIT within each group of dogs and the Mann Whitney test was used to compare the GIT segments between clinically unaffected and affected dogs. Results The infected dogs had an increased number of macrophages, plasma cells and lymphocytes, but lesions were generally mild. Parasite distribution in the GIT was evident in all intestinal segments and layers of the intestinal wall (mucosal, muscular and submucosal irrespective of the clinical status of the dogs. However, the parasite load was statistically higher in the caecum and colon than in other segments of the GIT. Conclusion The high parasite burden evident throughout the GIT mucosa with only mild pathological alterations led us to consider whether Leishmania gains an advantage from the intestinal immunoregulatory response (immunological tolerance.

  19. Infectivity to Phlebotomus perniciosus of dogs naturally parasitized with Leishmania infantum after different treatments

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    Descalzo Miguel A

    2011-04-01

    Full Text Available Abstract Background In Europe most dogs with clinical leishmaniosis are treated with leishmanicides, typically antimonials combined with allopurinol and good clinical recovery is observed in a high number of these dogs. Through xenodiagnosis, the capacity of a treated animal to infect the vector of the disease under treatment is assessed as a measure of the chemotherapeutic efficacy of the drug used. The objective of the present study was to evaluate through direct xenodiagnosis the infectivity to Phlebotomus perniciosus of dogs naturally parasitized with Leishmania infantum after treatment, and to follow the clinical and parasite course of disease. Thirty two dogs with clinical leishmaniosis were assigned to one of three treatment groups: meglumine antimoniate plus allopurinol (Group A, meglumine antimoniate (Group B or allopurinol (Group C. During the study, the dogs were examined before treatment (Day 0 and bimonthly thereafter until Day 180 (six months post-treatment onset. Results The three groups were scored over time according to the effects of treatment on clinical signs and clinical-pathological variables. Significant differences in clinical scores were observed between Group A and the other two groups, indicating the combined treatment was the most effective. After treatment, bone marrow cultures were positive for the parasite in 30.8% of dogs in some of the check ups (3 or 25% in Group A, 1 or 11.1% in Group B, and 4 or 80% in Group C. Our xenodiagnosis experiments revealed that 15.4% of treated dogs were still able to infect sand flies at some point after treatment (2 dogs or 16.6% in Group A, 2 or 22.2% in Group B and none in Group C. Only 7.7% of the entire study population could infect sand flies as from the second month post-treatment onset. Conclusion The three treatment regimens tested significantly reduced the infectivity of dogs towards sand flies, thus diminishing the epidemiological risks of treated dogs both for human

  20. Leishmania infantum HSP70-II null mutant as candidate vaccine against leishmaniasis: a preliminary evaluation

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    Fresno Manuel

    2011-07-01

    Full Text Available Abstract Background Visceral leishmaniasis is the most severe form of leishmaniasis and no effective vaccine exists. The use of live attenuated vaccines is emerging as a promising vaccination strategy. Results In this study, we tested the ability of a Leishmania infantum deletion mutant, lacking both HSP70-II alleles (ΔHSP70-II, to provide protection against Leishmania infection in the L. major-BALB/c infection model. Administration of the mutant line by either intraperitoneal, intravenous or subcutaneous route invariably leads to the production of high levels of NO and the development in mice of type 1 immune responses, as determined by analysis of anti-Leishmania IgG subclasses. In addition, we have shown that ΔHSP70-II would be a safe live vaccine as immunodeficient SCID mice, and hamsters (Mesocricetus auratus, infected with mutant parasites did not develop any sign of pathology. Conclusions The results suggest that the ΔHSP70-II mutant is a promising and safe vaccine, but further studies in more appropriate animal models (hamsters and dogs are needed to appraise whether this attenuate mutant would be useful as vaccine against visceral leishmaniasis.

  1. Leishmania infantum and Epstein-Barr virus co-infection in a patient with hemophagocytosis

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    Zied Gaifer

    2017-01-01

    Full Text Available The authors describe a rare case of a 27-year old previously healthy male presenting with high grade fever, pancytopenia, hepatosplenomegaly, high levels of ferritin and triglyceride, suggesting a diagnosis of hemophagocytic lymphohistiocytosis (HLH syndrome. Other investigations showed a positive Leishmania infantum serology and high Epstein-Barr virus (EBV viremia. The diagnosis of a visceral leishmaniasis was confirmed by bone morrow biopsy, which showed Leishman-Donovan bodies and evidence of HLH. The patient received liposomal amphotericin B and he had a complete resolution of his symptoms and clearance of EBV viremia. This case of HLH associated with visceral leishmaniasis and EBV co-infection raises the question about the significance of EBV in patients with HLH. The treatment of actual etiological agent can lead to complete cure while using current recommend chemotherapy for HLH-related EBV in a patient with hidden infection may have deleterious effects.

  2. Antileishmanial activity and tubulin polymerization inhibition of podophyllotoxin derivatives on Leishmania infantum

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    José Miguel Escudero-Martínez

    2017-12-01

    Full Text Available Leishmania microtubules play an important role not only in cell division, but also in keeping the shape of the parasite and motility of its free-living stages. Microtubules result from the self-assembly of alpha and beta tubulins, two phylogenetically conserved and very abundant eukaryotic proteins in kinetoplastids. The colchicine binding domain has inspired the discovery and development of several drugs currently in clinical use against parasites. However, this domain is less conserved in kinetoplastids and may be selectively targeted by new compounds. This report shows the antileishmanial effect of several series of compounds (53, derived from podophyllotoxin (a natural cyclolignan isolated from rhizomes of Podophyllum spp. and podophyllic aldehyde, on a transgenic, fluorescence-emitting strain of Leishmania infantum. These compounds were tested on both promastigotes and amastigote-infected mouse splenocytes, and in mammalian – mouse non-infected splenocytes and liver HepG2 cells – in order to determine selective indexes of the drugs. Results obtained with podophyllotoxin derivatives showed that the hydroxyl group at position C-7α was a structural requisite to kill the parasites. On regards podophyllic aldehyde, derivatives with C9-aldehyde group integrated into a bicyclic heterostructure displayed more potent antileishmanial effects and were relatively safe for host cells. Docking studies of podophyllotoxin and podophyllic aldehyde derivatives showed that these compounds share a similar pattern of interaction at the colchicine site of Leishmania tubulin, thus pointing to a common mechanism of action. However, the results obtained suggested that despite tubulin is a remarkable target against leishmaniasis, there is a poor correlation between inhibition of tubulin polymerization and antileishmanial effect of many of the compounds tested, fact that points to alternative pathways to kill the parasites. Keywords: Leishmania, Tubulin, DNA

  3. Nuclear DNA polymerase beta from Leishmania infantum. Cloning, molecular analysis and developmental regulation

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    Taladriz, Soraya; Hanke, Tobias; Ramiro, María J.; García-Díaz, Miguel; Lacoba, Mario García de; Blanco, Luis; Larraga, Vicente

    2001-01-01

    We have identified a novel polymerase beta (Pol β)-like enzyme from Leishmania infantum, a parasite protozoon causing disease in humans. This protein, named Li Pol β, shows a nuclear localization that contrasts with the mitochondrial localization of Pol β from Crithidia fasciculata, a closely related parasite, the only polymerase β described so far in Trypanosomatidae. Li Pol β, that belongs to the DNA polymerase X family, displays an evolutionarily conserved Pol β-type DNA polymerase core, in which most of the key residues involved in DNA binding, nucleotide binding, dRPase and polymerization catalysis are conserved. In agreement with this, Li Pol β, overproduced in Escherichia coli, displayed intrinsic DNA polymerase activity. Cell synchronization experiments showed a correlation between both Li Pol β mRNA and protein levels along the parasite cell cycle. Analysis of these parameters at the different growth phases of the parasite, from the proliferative (non-infective) logarithmic phase to the non-dividing (highly infectious) stationary phase, showed high levels of Li Pol β at the infective phase of the parasite. The data suggest a role of Li Pol β in base excision repair in L.infantum, a parasite usually affected by oxygen stress environments into the macrophage host cells. PMID:11557814

  4. In Vitro Antiparasitic and Apoptotic Effects of Antimony Sulfide Nanoparticles on Leishmania infantum

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    Saied Soflaei

    2012-01-01

    Full Text Available Visceral leishmaniasis is one of the most important sever diseases in tropical and subtropical countries. In the present study the effects of antimony sulfide nanoparticles on Leishmania infantum in vitro were evaluated. Antimony sulfide NPs (Sb2S5 were synthesized by biological method from Serratia marcescens bacteria. Then the cytotoxicity effects of different concentrations (5, 10, 25, 50, and 100 μg/mL of this nanoparticle were assessed on promastigote and amastigote stages of L. infantum. MTT method was used for verification results of promastigote assay. Finally, the percentages of apoptotic, necrotic, and viable cells were determined by flow cytometry. The results indicated the positive effectiveness of antimony sulfide NPs on proliferation of promastigote form. The IC50 (50% inhibitory concentration of antimony sulfide NPs on promastigotes was calculated 50 μg/mL. The cytotoxicity effect was dose-dependent means by increasing the concentration of antimony sulfide NPs, the cytotoxicity curve was raised and the viability curve of the parasite dropped simultaneously. Moreover, the IC50 of antimony sulfide NPs on amastigote stage was calculated 25 μg/mL. On the other hand, however, antimony sulfide NPs have a low cytotoxicity effect on uninfected macrophages but it can induce apoptosis in promastigote stage at 3 of 4 concentrations.

  5. A diagnostic assay based on variable intergenic region distinguishes between Leishmania donovani and Leishmania infantum

    Czech Academy of Sciences Publication Activity Database

    Chocholová, Eva; Jirků, Milan; Lukeš, Julius

    2008-01-01

    Roč. 55, č. 1 (2008), s. 75-78 ISSN 0015-5683 R&D Projects: GA MŠk LC07032; GA MŠk 2B06129 Institutional research plan: CEZ:AV0Z60220518 Keywords : Leishmania * assay * diagnosis Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.307, year: 2008

  6. Genomic confirmation of hybridisation and recent inbreeding in a vector-isolated Leishmania population.

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    Rogers, Matthew B; Downing, Tim; Smith, Barbara A; Imamura, Hideo; Sanders, Mandy; Svobodova, Milena; Volf, Petr; Berriman, Matthew; Cotton, James A; Smith, Deborah F

    2014-01-01

    Although asexual reproduction via clonal propagation has been proposed as the principal reproductive mechanism across parasitic protozoa of the Leishmania genus, sexual recombination has long been suspected, based on hybrid marker profiles detected in field isolates from different geographical locations. The recent experimental demonstration of a sexual cycle in Leishmania within sand flies has confirmed the occurrence of hybridisation, but knowledge of the parasite life cycle in the wild still remains limited. Here, we use whole genome sequencing to investigate the frequency of sexual reproduction in Leishmania, by sequencing the genomes of 11 Leishmania infantum isolates from sand flies and 1 patient isolate in a focus of cutaneous leishmaniasis in the Çukurova province of southeast Turkey. This is the first genome-wide examination of a vector-isolated population of Leishmania parasites. A genome-wide pattern of patchy heterozygosity and SNP density was observed both within individual strains and across the whole group. Comparisons with other Leishmania donovani complex genome sequences suggest that these isolates are derived from a single cross of two diverse strains with subsequent recombination within the population. This interpretation is supported by a statistical model of the genomic variability for each strain compared to the L. infantum reference genome strain as well as genome-wide scans for recombination within the population. Further analysis of these heterozygous blocks indicates that the two parents were phylogenetically distinct. Patterns of linkage disequilibrium indicate that this population reproduced primarily clonally following the original hybridisation event, but that some recombination also occurred. This observation allowed us to estimate the relative rates of sexual and asexual reproduction within this population, to our knowledge the first quantitative estimate of these events during the Leishmania life cycle.

  7. Detection of Leishmania infantum in naturally infected Lutzomyia longipalpis (Diptera: Psychodidae: Phlebotominae) and Canis familiaris in Misiones, Argentina: the first report of a PCR-RFLP and sequencing-based confirmation assay.

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    Acardi, Soraya Alejandra; Liotta, Domingo Javier; Santini, María Soledad; Romagosa, Carlo Mariano; Salomón, Oscar Daniel

    2010-09-01

    In this study, a genotypification of Leishmania was performed using polimerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and sequencing techniques to identify species of Leishmania parasites in phlebotomine sand flies and dogs naturally infected. Between January-February of 2009, CDC light traps were used to collect insect samples from 13 capture sites in the municipality of Posadas, which is located in the province of Misiones of Argentina. Sand flies identified as Lutzomyia longipalpis were grouped into 28 separate pools for molecular biological analysis. Canine samples were taken from lymph node aspirates of two symptomatic stray animals that had been positively diagnosed with canine visceral leishmaniasis. One vector pool of 10 sand flies (1 out of the 28 pools tested) and both of the canine samples tested positively for Leishmania infantum by PCR and RFLP analysis. PCR products were confirmed by sequencing and showed a maximum identity with L. infantum. Given that infection was detected in one out of the 28 pools and that at least one infected insect was infected, it was possible to infer an infection rate at least of 0.47% for Lu. longipalpis among the analyzed samples. These results contribute to incriminate Lu. longipalpis as the vector of L. infantum in the municipality of Posadas, where cases of the disease in humans and dogs have been reported since 2005.

  8. Mimotope-based vaccines of Leishmania infantum antigens and their protective efficacy against visceral leishmaniasis.

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    Lourena Emanuele Costa

    Full Text Available BACKGROUND: The development of cost-effective prophylactic strategies to prevent leishmaniasis has become a high-priority. The present study has used the phage display technology to identify new immunogens, which were evaluated as vaccines in the murine model of visceral leishmaniasis (VL. Epitope-based immunogens, represented by phage-fused peptides that mimic Leishmania infantum antigens, were selected according to their affinity to antibodies from asymptomatic and symptomatic VL dogs' sera. METHODOLOGY/MAIN FINDINGS: Twenty phage clones were selected after three selection cycles, and were evaluated by means of in vitro assays of the immune stimulation of spleen cells derived from naive and chronically infected with L. infantum BALB/c mice. Clones that were able to induce specific Th1 immune response, represented by high levels of IFN-γ and low levels of IL-4 were selected, and based on their selectivity and specificity, two clones, namely B10 and C01, were further employed in the vaccination protocols. BALB/c mice vaccinated with clones plus saponin showed both a high and specific production of IFN-γ, IL-12, and GM-CSF after in vitro stimulation with individual clones or L. infantum extracts. Additionally, these animals, when compared to control groups (saline, saponin, wild-type phage plus saponin, or non-relevant phage clone plus saponin, showed significant reductions in the parasite burden in the liver, spleen, bone marrow, and paws' draining lymph nodes. Protection was associated with an IL-12-dependent production of IFN-γ, mainly by CD8+ T cells, against parasite proteins. These animals also presented decreased parasite-mediated IL-4 and IL-10 responses, and increased levels of parasite-specific IgG2a antibodies. CONCLUSIONS/SIGNIFICANCE: This study describes two phage clones that mimic L. infantum antigens, which were directly used as immunogens in vaccines and presented Th1-type immune responses, and that significantly reduced the

  9. Leishmania infantum EndoG is an endo/exo-nuclease essential for parasite survival.

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    Eva Rico

    Full Text Available EndoG, a member of the DNA/RNA non-specific ββα-metal family of nucleases, has been demonstrated to be present in many organisms, including Trypanosomatids. This nuclease participates in the apoptotic program in these parasites by migrating from the mitochondrion to the nucleus, where it takes part in the degradation of genomic DNA that characterizes this process. We now demonstrate that Leishmania infantum EndoG (LiEndoG is an endo-exonuclease that has a preferential 5' exonuclease activity on linear DNA. Regardless of its role during apoptotic cell death, this enzyme seems to be necessary during normal development of the parasites as indicated by the reduced growth rates observed in LiEndoG hemi-knockouts and their poor infectivity in differentiated THP-1 cells. The pro-life role of this protein is also corroborated by the higher survival rates of parasites that over-express this protein after treatment with the LiEndoG inhibitor Lei49. Taken together, our results demonstrate that this enzyme plays essential roles in both survival and death of Leishmania parasites.

  10. Phlebotomine sand fly survey in the focus of leishmaniasis in Madrid, Spain (2012-2014): seasonal dynamics, Leishmania infantum infection rates and blood meal preferences.

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    González, Estela; Jiménez, Maribel; Hernández, Sonia; Martín-Martín, Inés; Molina, Ricardo

    2017-08-01

    An unusual increase of human leishmaniasis cases due to Leishmania infantum is occurring in an urban area of southwestern Madrid, Spain, since 2010. Entomological surveys have shown that Phlebotomus perniciosus is the only potential vector. Direct xenodiagnosis in hares (Lepus granatensis) and rabbits (Oryctolagus cuniculus) collected in the focus area proved that they can transmit parasites to colonized P. perniciosus. Isolates were characterized as L. infantum. The aim of the present work was to conduct a comprehensive study of sand flies in the outbreak area, with special emphasis on P. perniciosus. Entomological surveys were done from June to October 2012-2014 in 4 stations located close to the affected area. Twenty sticky traps (ST) and two CDC light traps (LT) were monthly placed during two consecutive days in every station. LT were replaced every morning. Sand fly infection rates were determined by dissecting females collected with LT. Molecular procedures applied to study blood meal preferences and to detect L. infantum were performed for a better understanding of the epidemiology of the outbreak. A total of 45,127 specimens belonging to 4 sand fly species were collected: P. perniciosus (75.34%), Sergentomyia minuta (24.65%), Phlebotomus sergenti (0.005%) and Phlebotomus papatasi (0.005%). No Phlebotomus ariasi were captured. From 3203 P. perniciosus female dissected, 117 were infected with flagellates (3.7%). Furthermore, 13.31% and 7.78% of blood-fed and unfed female sand flies, respectively, were found infected with L. infantum by PCR. The highest rates of infected P. perniciosus were detected at the end of the transmission periods. Regarding to blood meal preferences, hares and rabbits were preferred, although human, cat and dog blood were also found. This entomological study highlights the exceptional nature of the Leishmania outbreak occurring in southwestern Madrid, Spain. It is confirmed that P. perniciosus is the only vector in the affected area

  11. New alkenyl derivative from Piper malacophyllum and analogues: Antiparasitic activity against Trypanosoma cruzi and Leishmania infantum.

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    Varela, Marina T; Lima, Marta L; Galuppo, Mariana K; Tempone, Andre G; de Oliveira, Alberto; Lago, João Henrique G; Fernandes, João Paulo S

    2017-11-01

    Alkylphenols isolated from Piper malacophyllum (Piperaceae), gibbilimbols A and B, showed interesting activity against the parasites Trypanosoma cruzi and Leishmania infantum. In continuation to our previous work, a new natural product from the essential oil of the leaves of P. malacophyllum was isolated, the 5-[(3E)-oct-3-en-1-il]-1,3-benzodioxole, and also a new set of five compounds was prepared. The antiparasitic activity of the natural product was evaluated in vitro against these parasites, indicating potential against the promastigote/trypomastigote/amastigote forms (IC 50 32-83 μm) of the parasites and low toxicity (CC 50  > 200 μm) to mammalian cells. The results obtained to the synthetic compounds indicated that the new derivatives maintained the promising antiparasitic activity, but the cytotoxicity was considerably lowered. The amine derivative LINS03011 displayed the most potent IC 50 values (13.3 and 16.7 μm) against amastigotes of T. cruzi and L. infantum, respectively, indicating comparable activity to the phenolic prototype LINS03003, with threefold decreased (CC 50 73.5 μm) cytotoxicity, leading the selectivity index (SI) towards the parasites up to 24.5. In counterpart, LINS03011 has not shown membrane disruptor activity in SYTOX Green model. In summary, this new set showed the hydroxyl is not essential for the antiparasitic activity, and its substitution could decrease the toxicity to mammalian cells. © 2017 John Wiley & Sons A/S.

  12. In vitro antileishmanial activity of fisetin flavonoid via inhibition of glutathione biosynthesis and arginase activity in Leishmania infantum.

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    Adinehbeigi, Keivan; Razi Jalali, Mohammad Hossein; Shahriari, Ali; Bahrami, Somayeh

    2017-06-01

    With the increasing emergence of drug resistant Leishmania sp. in recent years, combination therapy has been considered as a useful way to treat and control of Leishmaniasis. The present study was designed to evaluate the antileishmanial effects of the fisetin alone and combination of fisetin plus Meglumine antimoniate (Fi-MA) against Leishmania infantum. The IC50 values for fisetin were obtained 0.283 and 0.102 μM against promastigotes and amastigote forms, respectively. Meglumine antimoniate (MA, Glucantime) as control drug also revealed IC50 values of 0.247 and 0.105 μM for promastigotes and amastigotes of L. infantum, respectively. In order to determine the mode of action of fisetin and Meglumine antimoniate (MA, Glucantime), the activities of arginase (ARG), catalase (CAT), glutathione peroxidase (GPx) and superoxide dismutase (SOD) were measured. Moreover, intracellular glutathione (GSH) and nitric oxide (NO) levels in L. infantum-infected macrophages and L. infantum promastigotes which were treated with IC50 concentrations of fisetin, MA and Fi-MA were investigated. Our results showed that MA decreased CAT and SOD activity and increased NO levels in L. infantum-infected macrophages. In promastigotes, MA inhibited parasite SOD activity and reduced parasite NO production. The decreased levels of most of the antioxidant enzymes, accompanying by the raised level of NO in treated macrophages with MA, were observed to regain their normal profiles due to Fi-MA treatment. Furthermore, fisetin could prevent the growth of promastigotes by inhibition of ARG activity and reduction of GSH levels and NO production. In conclusion, these findings showed that fisetin improves MA side effects.

  13. In vitro activity of the beta-carboline alkaloids harmane, harmine, and harmaline toward parasites of the species Leishmania infantum.

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    Di Giorgio, C; Delmas, F; Ollivier, E; Elias, R; Balansard, G; Timon-David, P

    2004-01-01

    Harmane, harmine, and harmaline were investigated for their in vitro antileishmanial activity toward parasites of the species Leishmania infantum. Harmane and Harmine displayed a moderate antiproliferative activity toward human monocytes and exerted a weak antileishmanial activity toward both the promastigote and the amastigote forms of the parasite. Their mechanism of action on the promastigote form of the parasite involved interactions with DNA metabolism leading to an accumulation of parasites in the S-G(2)M phases of the cell-cycle. Harmaline, at the contrary, was deprived from toxicity toward human cells and Leishmania promastigotes, however it exerted a strong antileishmanial activity toward the intracellular amastigote form of the parasite. This property was shown to partly result from the capacity of the molecule to prevent parasite internalization within macrophages by inhibiting Leishmania PKC activity.

  14. Infección de fibroblastos de piel de animales con distinto grado de susceptibilidad a Leishmania infantum y Leishmania mexicana (Kinetoplastida: Trypanosomatidae

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    Miguel Angel Minero

    2004-03-01

    Full Text Available En este estudio se presenta un modelo in vitro de cultivo de fibroblastos de piel de hámster, ratón y rata hecho con el propósito de determinar diferencias en cuanto a la susceptibilidad a la infección por dos especies del género Leishmania (Kinetoplastida: Trypanosomatidae. Se realizó además un estudio ultraestructural por microscopía electrónica de transmisión con el fin de establecer si las formas intracelulares observadas correspondían a multiplicación interna o a fagocitosis múltiple. Se estudió la multiplicación de los parásitos en los fibroblastos de las tres especies de roedores infectados tanto por Leishmania infantum como por L. mexicana (cepa OCR y las diferencias entre las tres fueron estadísticamente significativas (pInfection and multiplication of Leishmania infantum and L. mexicana inside of skin fibroblasts from hamsters, mice and rats was achieved. This process was demonstrated either by counting parasites inside the stained cells or by electronic microscopy studies. In addition multiplication rate differences in the cells from these rodent species were determined, for L. infantum as well as for L. mexicana. Parasite development in hamsters and mice fibroblasts was evident but there was not multiplication in rat cells showing that apparently they are refractory to Leishmania infection. These results suggest that the parasite affinity for each animal, as well as any intracellular environment resistance, could involve genetic factors in the parasite multiplication. On the other hand, presence of amastigote multiplication inside of parasitophorus vacuole, showed by electronic microscopy images, probes a true parasite transformation. Therefore it is suggested that fibroblasts could work as host cells for parasite survival and permanency in the infected animals

  15. Seroepidemiology of Leishmania infantum in dogs in the city of Porto Alegre, Rio Grande do Sul, Brazil

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    Mariana Caetano Teixeira

    2016-12-01

    Full Text Available Leishmaniosis are zoonoses that present several clinical manifestations in humans and have dogs as their main reservoir in the urban environment. Visceral leishmaniasis (VL is the most severe form of the parasitosis and has been increasing in Brazil, despite the actions of public health agencies. Until 2002, the State of Rio Grande do Sul (RS was considered free of human and canine leishmaniasis. The first human case of cutaneous leishmaniasis in RS was recorded in 2003. In 2009, the first autochthonous cases of human VL and canine visceral leishmaniasis (CVL were confirmed in São Borja, RS, and the occurrence of the insect vector was recorded for the first time in the state. In 2010, the first confirmed case of CVL was reported and seropositive dogs were identified in the city of Porto Alegre, RS. Given the importance of this zoonosis and the difficulties of a reliable diagnosis in dogs, this study aimed to identify epidemiological aspects of CVL in dogs in an area of Porto Alegre where cases of the disease have been reported. A total of 300 blood samples were collected from dogs in this area, which were then tested by the methods of RT-DPP® and ELISA for diagnosis of Leishmania infantum. An epidemiological questionnaire was completed by dog owners, containing aspects related to care of the animals, characteristics of their environment, and their living conditions. We observed that 83% (250/300 of the studied dogs were of mixed breed, 58% (175/300 were female, 78% (238/300 slept outdoors, and 61% (183/300 shared their living quarters with other species. Clinically, we observed that 90% (270/300 of the animals were infested by ectoparasites, 70% (210/300 had dermatopathies, 24% (72/300 presented weight loss and anorexia, and 22% (65/300 had ocular disorders. The results of the two serological tests were 100% concordant for the three seropositive samples (1%, and the remaining 297 (99% were negative for both tests. We conclude that despite the

  16. Case report of canine co-infection with Leishmania infantum and Ehrlichia canis

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    Stefanovska Jovana

    2011-05-01

    Full Text Available Canine leishmaniasis (CanL due to Leishmania infantum and canine monocytic ehrilichiosis (CME due to Ehrlichia canis are common diseases with zoonotic potential in the Mediterranean area. Their prevalence in R. Macedonia as a neighboring Mediterranean county is expected. In both diseases similar clinical symptoms can be manifested in dogs such as: lethargy, anorexia, weight loss, epistaxis, fever, pale mucous membranes, enlarged lymph nodes, splenomegaly, ocular signs. This case report present an atypical case of 11 year old female Samoyed with starting single clinical symptom epistaxys. Initial diagnostic procedures revealed the presence only of CanL, which was diagnosed using indirect immunofluorescence method and ELISA. First laboratory findings showed normal hematological and renal profiles. Dog was put on a treatment with Allopurinol (20mg/kg, p/o for at least 9 months. Termination of the therapy after 6 months brought a numerous clinical symptoms involving weakness, dehydration, pale mucous membranes lost pupilar reflex, uremic breath and biochemical parameters revealed a renal failure. Using a commercial ELISA kit Ehrlichia canis as a co infection was diagnosed. Most probably the second infectious agent was induced in the past 6 months, causing more severe pathological effects than CanL infection alone.

  17. In vitro anti-Leishmania infantum activity of essential oil from Piper angustifolium

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    Lauriane S.S. Bosquiroli

    Full Text Available Abstract Piper angustifolium Lam., Piperaceae, popularly known as "matito", "pimenta-de-macaco", "pimenta-longa" or "jagurandi" in Brazil, has been commonly used in the treatment of cutaneous leishmaniasis-associated lesions, but there are few studies on the activity against visceral leishmaniasis-associated species. This study demonstrates the first in vitro antileishmanial activity of the P. angustifolium essential oil, of which the phytochemical profile showed the presence of sesquiterpenes and monoterpenes. The main compounds were spathulenol (23.8% and caryophyllene oxide (13.1%. P. angustifolium essential oil was highly active [the half maximum inhibitory concentration = 1.43 μg/ml] against intracellular amastigotes of Leishmania infantum, the etiological agent of visceral leishmaniasis in the New and Old World. Activity was obtained 24 h after addition of the oil (6.25–50 μg/ml, with a reduction of 100% in the infection index at concentrations of 25 and 50 μg/ml. P. angustifolium essential oil showed low cytotoxicity for mammalian fibroblasts and macrophages (the half maximum inhibitory concentration values of 31.67 and 48.22 μg/ml, respectively, and it was 33 and 22 times more toxic to amastigotes than to mammalian cells, as indicated by selectivity indexes. The results demonstrated that P. angustifolium essential oil is a promising alternative for the study of potential drugs for visceral leishmaniasis.

  18. Use of a Recombinant Cysteine Proteinase from Leishmania (Leishmania) infantum chagasi for the Immunotherapy of Canine Visceral Leishmaniasis

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    Ferreira, Josie Haydée Lima; Silva, Lucilene dos Santos; Longo-Maugéri, Ieda Maria; Katz, Simone; Barbiéri, Clara Lúcia

    2014-01-01

    Background A recombinant cysteine proteinase from Leishmania (Leishmania) infantum chagasi (rLdccys1) was previously shown to induce protective immune responses against murine and canine visceral leishmaniasis. These findings encouraged us to use rLdccys1 in the immunotherapy of naturally infected dogs from Teresina, Piauí, a region of high incidence of visceral leishmaniasis in Brazil. Methodology/Principal Findings Thirty naturally infected mongrel dogs displaying clinical signs of visceral leishmaniasis were randomly divided in three groups: one group received three doses of rLdccys1 in combination with the adjuvant Propionibacterium acnes at one month interval between each dose; a second group received three doses of P. acnes alone; a third group received saline. The main findings were: 1) dogs that received rLdccys1 with P. acnes did not display increase of the following clinical signs: weight loss, alopecia, onychogryphosis, cachexia, anorexia, apathy, skin lesions, hyperkeratosis, ocular secretion, and enlarged lymph nodes; they also exhibited a significant reduction in the spleen parasite load in comparison to the control dogs; 2) rLdccys1-treated dogs exhibited a significant delayed type cutaneous hypersensitivity elicited by the recombinant antigen, as well as high IgG2 serum titers and low IgG1 serum titers; sera from rLdccys1-treated dogs also contained high IFN-γ and low IL-10 concentrations; 3) control dogs exhibited all of the clinical signs of visceral leishmaniasis and had low serum IgG2 and IFN-γ levels and high concentrations of IgG1 and IL-10; 4) all of the dogs treated with rLdccys1 were alive 12 months after treatment, whereas dogs which received either saline or P. acnes alone died within 3 to 7 months. Conclusions/Significance These findings illustrate the potential use of rLdccys1 as an additional tool for the immunotherapy of canine visceral leishmaniasis and support further studies designed to improve the efficacy of this recombinant

  19. Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of glyoxalase I from Leishmania infantum

    International Nuclear Information System (INIS)

    Barata, Lídia; Sousa Silva, Marta; Schuldt, Linda; Costa, Gonçalo da; Tomás, Ana M.; Ferreira, António E. N.; Weiss, Manfred S.; Ponces Freire, Ana; Cordeiro, Carlos

    2010-01-01

    Glyoxalase I from L. infantum was cloned, overexpressed in E. coli, purified and crystallized. Glyoxalase I (GLO1) is the first of the two glyoxalase-pathway enzymes. It catalyzes the formation of S-d-lactoyltrypanothione from the non-enzymatically formed hemithioacetal of methylglyoxal and reduced trypanothione. In order to understand its substrate binding and catalytic mechanism, GLO1 from Leishmania infantum was cloned, overexpressed in Escherichia coli, purified and crystallized. Two crystal forms were obtained: a cube-shaped form and a rod-shaped form. While the cube-shaped form did not diffract X-rays at all, the rod-shaped form exhibited diffraction to about 2.0 Å resolution. The crystals belonged to space group P2 1 2 1 2, with unit-cell parameters a = 130.03, b = 148.51, c = 50.63 Å and three dimers of the enzyme per asymmetric unit

  20. Molecular detection of Leishmania infection in sand flies in border line of Iran-Turkmenistan: restricted and permissive vectors.

    Science.gov (United States)

    Bakhshi, H; Oshaghi, M A; Abai, M R; Rassi, Y; Akhavan, A A; Sheikh, Z; Mohtarami, F; Saidi, Z; Mirzajani, H; Anjomruz, M

    2013-10-01

    A molecular study was carried out to incriminate sand fly vectors of cutaneous leishmaniasis (CL) in rural areas of Sarakhs district, Khorassane-Razavi Province, northeastern Iran, in 2011. Sand flies of Sergentomyia with three species and Phlebotomus with six species respectively comprised 73.3% and 26.7% of the specimens. Phlebotomus papatasi was the most common Phlebotomine species in outdoor and indoor resting places. Leishmania infection was found at least in 17 (22%) specimens including Ph. papatasi (n=9 pool samples), Phlebotomus caucasicus (n=6), Phlebotomus alexandri (n=1), and Sergentomyia sintoni (n=1). The parasites were found comprised Leishmania major (n=5), Leishmania turanica (n=10), and Leishmania gerbilli (n=4). Infection of Ph. papatasi with both L. major and L. turanica supporting the new suggestion indicating that it is not restricted only with L. major. Circulation of L. major by Ph. alexandri, and both L. gerbilli and L. turanica by Ph. caucasicus, in addition to previous data indicating the ability of Ph. alexandri to circulate Leishmania infantum and Leishmania donovani, and Ph. caucasicus to circulate L. major, suggests that these two species can be permissive vectors. The results suggest that Ph. papatasi and Ph. alexandri are the primary and secondary vectors of CL where circulating L. major between human and reservoirs, whereas Ph. caucasicus is circulating L. turanica and L. gerbilli between the rodents in the region. Copyright © 2013 Elsevier Inc. All rights reserved.

  1. Occurrence of Leishmania infantum in the central nervous system of naturally infected dogs: Parasite load, viability, co-infections and histological alterations.

    Science.gov (United States)

    Oliveira, Valéria da Costa; Boechat, Viviane Cardoso; Mendes Junior, Artur Augusto Velho; Madeira, Maria de Fátima; Ferreira, Luiz Claudio; Figueiredo, Fabiano Borges; Campos, Monique Paiva; de Carvalho Rodrigues, Francisco das Chagas; Carvalhaes de Oliveira, Raquel de Vasconcellos; Amendoeira, Maria Regina Reis; Menezes, Rodrigo Caldas

    2017-01-01

    Zoonotic visceral leishmaniasis is caused by the protozoan Leishmania infantum and little is known about the occurrence and pathogenesis of this parasite in the CNS. The aims of this study were to evaluate the occurrence, viability and load of L. infantum in the CNS, and to identify the neurological histological alterations associated with this protozoan and its co-infections in naturally infected dogs. Forty-eight Leishmania-seropositive dogs from which L. infantum was isolated after necropsy were examined. Cerebrospinal fluid (CSF) samples were analyzed by parasitological culture, quantitative real-time PCR (qPCR) and the rapid immunochromatographic Dual Path Platform test. Brain, spinal cord and spleen samples were submitted to parasitological culture, qPCR, and histological techniques. Additionally, anti-Toxoplasma gondii and anti-Ehrlichia canis antibodies in serum and distemper virus antigens in CSF were investigated. None of the dogs showed neurological signs. All dogs tested positive for L. infantum in the CNS. Viable forms of L. infantum were isolated from CSF, brain and spinal cord in 25% of the dogs. Anti-L. infantum antibodies were detected in CSF in 61% of 36 dogs. Inflammatory histological alterations were observed in the CNS of 31% of the animals; of these, 66% were seropositive for E. canis and/or T. gondii. Amastigote forms were associated with granulomatous non-suppurative encephalomyelitis in a dog without evidence of co-infections. The highest frequency of L. infantum DNA was observed in the brain (98%), followed by the spinal cord (96%), spleen (95%), and CSF (50%). The highest L. infantum load in CNS was found in the spinal cord. These results demonstrate that L. infantum can cross the blood-brain barrier, spread through CSF, and cause active infection in the entire CNS of dogs. Additionally, L. infantum can cause inflammation in the CNS that can lead to neurological signs with progression of the disease.

  2. Occurrence of Leishmania infantum in the central nervous system of naturally infected dogs: Parasite load, viability, co-infections and histological alterations.

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    Valéria da Costa Oliveira

    Full Text Available Zoonotic visceral leishmaniasis is caused by the protozoan Leishmania infantum and little is known about the occurrence and pathogenesis of this parasite in the CNS. The aims of this study were to evaluate the occurrence, viability and load of L. infantum in the CNS, and to identify the neurological histological alterations associated with this protozoan and its co-infections in naturally infected dogs. Forty-eight Leishmania-seropositive dogs from which L. infantum was isolated after necropsy were examined. Cerebrospinal fluid (CSF samples were analyzed by parasitological culture, quantitative real-time PCR (qPCR and the rapid immunochromatographic Dual Path Platform test. Brain, spinal cord and spleen samples were submitted to parasitological culture, qPCR, and histological techniques. Additionally, anti-Toxoplasma gondii and anti-Ehrlichia canis antibodies in serum and distemper virus antigens in CSF were investigated. None of the dogs showed neurological signs. All dogs tested positive for L. infantum in the CNS. Viable forms of L. infantum were isolated from CSF, brain and spinal cord in 25% of the dogs. Anti-L. infantum antibodies were detected in CSF in 61% of 36 dogs. Inflammatory histological alterations were observed in the CNS of 31% of the animals; of these, 66% were seropositive for E. canis and/or T. gondii. Amastigote forms were associated with granulomatous non-suppurative encephalomyelitis in a dog without evidence of co-infections. The highest frequency of L. infantum DNA was observed in the brain (98%, followed by the spinal cord (96%, spleen (95%, and CSF (50%. The highest L. infantum load in CNS was found in the spinal cord. These results demonstrate that L. infantum can cross the blood-brain barrier, spread through CSF, and cause active infection in the entire CNS of dogs. Additionally, L. infantum can cause inflammation in the CNS that can lead to neurological signs with progression of the disease.

  3. Leishmania infantum ecto-nucleoside triphosphate diphosphohydrolase-2 is an apyrase involved in macrophage infection and expressed in infected dogs.

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    Raphael De Souza Vasconcellos

    2014-11-01

    Full Text Available Visceral leishmaniasis is an important tropical disease, and Leishmania infantum chagasi (synonym of Leishmania infantum is the main pathogenic agent of visceral leishmaniasis in the New World. Recently, ecto-nucleoside triphosphate diphosphohydrolases (E-NTPDases were identified as enablers of infection and virulence factors in many pathogens. Two putative E-NTPDases (∼70 kDa and ∼45 kDa have been found in the L. infantum genome. Here, we studied the ∼45 kDa E-NTPDase from L. infantum chagasi to describe its natural occurrence, biochemical characteristics and influence on macrophage infection.We used live L. infantum chagasi to demonstrate its natural ecto-nucleotidase activity. We then isolated, cloned and expressed recombinant rLicNTPDase-2 in bacterial system. The recombinant rLicNTPDase-2 hydrolyzed a wide variety of triphosphate and diphosphate nucleotides (GTP> GDP  =  UDP> ADP> UTP  =  ATP in the presence of calcium or magnesium. In addition, rLicNTPDase-2 showed stable activity over a pH range of 6.0 to 9.0 and was partially inhibited by ARL67156 and suramin. Microscopic analyses revealed the presence of this protein on cell surfaces, vesicles, flagellae, flagellar pockets, kinetoplasts, mitochondria and nuclei. The blockade of E-NTPDases using antibodies and competition led to lower levels of parasite adhesion and infection of macrophages. Furthermore, immunohistochemistry showed the expression of E-NTPDases in amastigotes in the lymph nodes of naturally infected dogs from an area of endemic visceral leishmaniasis.In this work, we cloned, expressed and characterized the NTPDase-2 from L. infantum chagasi and demonstrated that it functions as a genuine enzyme from the E-NTPDase/CD39 family. We showed that E-NTPDases are present on the surface of promastigotes and in other intracellular locations. We showed, for the first time, the broad expression of LicNTPDases in naturally infected dogs. Additionally, the blockade of

  4. Leishmania infantum Ecto-Nucleoside Triphosphate Diphosphohydrolase-2 is an Apyrase Involved in Macrophage Infection and Expressed in Infected Dogs

    Science.gov (United States)

    Vasconcellos, Raphael De Souza; Mariotini-Moura, Christiane; Gomes, Rodrigo Saar; Serafim, Tiago Donatelli; Firmino, Rafaela de Cássia; Silva e Bastos, Matheus; de Castro, Felipe Freitas; de Oliveira, Claudia Miranda; Borges-Pereira, Lucas; de Souza, Anna Cláudia Alves; de Souza, Ronny Francisco; Gómez, Gabriel Andres Tafur; Pinheiro, Aimara da Costa; Maciel, Talles Eduardo Ferreira; Silva-Júnior, Abelardo; Bressan, Gustavo Costa; Almeida, Márcia Rogéria; Baqui, Munira Muhammad Abdel; Afonso, Luís Carlos Crocco; Fietto, Juliana Lopes Rangel

    2014-01-01

    Background Visceral leishmaniasis is an important tropical disease, and Leishmania infantum chagasi (synonym of Leishmania infantum) is the main pathogenic agent of visceral leishmaniasis in the New World. Recently, ecto-nucleoside triphosphate diphosphohydrolases (E-NTPDases) were identified as enablers of infection and virulence factors in many pathogens. Two putative E-NTPDases (∼70 kDa and ∼45 kDa) have been found in the L. infantum genome. Here, we studied the ∼45 kDa E-NTPDase from L. infantum chagasi to describe its natural occurrence, biochemical characteristics and influence on macrophage infection. Methodology/Principal Findings We used live L. infantum chagasi to demonstrate its natural ecto-nucleotidase activity. We then isolated, cloned and expressed recombinant rLicNTPDase-2 in bacterial system. The recombinant rLicNTPDase-2 hydrolyzed a wide variety of triphosphate and diphosphate nucleotides (GTP> GDP  =  UDP> ADP> UTP  =  ATP) in the presence of calcium or magnesium. In addition, rLicNTPDase-2 showed stable activity over a pH range of 6.0 to 9.0 and was partially inhibited by ARL67156 and suramin. Microscopic analyses revealed the presence of this protein on cell surfaces, vesicles, flagellae, flagellar pockets, kinetoplasts, mitochondria and nuclei. The blockade of E-NTPDases using antibodies and competition led to lower levels of parasite adhesion and infection of macrophages. Furthermore, immunohistochemistry showed the expression of E-NTPDases in amastigotes in the lymph nodes of naturally infected dogs from an area of endemic visceral leishmaniasis. Conclusions/Significance In this work, we cloned, expressed and characterized the NTPDase-2 from L. infantum chagasi and demonstrated that it functions as a genuine enzyme from the E-NTPDase/CD39 family. We showed that E-NTPDases are present on the surface of promastigotes and in other intracellular locations. We showed, for the first time, the broad expression of LicNTPDases in

  5. F1 Domain of the Leishmania (Leishmania donovani Nucleoside Hydrolase Promotes a Th1 Response in Leishmania (Leishmania infantum Cured Patients and in Asymptomatic Individuals Living in an Endemic Area of Leishmaniasis

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    Eugenia Carrillo

    2017-07-01

    Full Text Available The Leishmania (Leishmania donovani nucleoside hydrolase NH36 is the main antigen of the Leishmune® vaccine and one of the promising candidates for vaccination against visceral leishmaniasis. The antigenicity of the N-terminal (F1, the central (F2, or the C-terminal recombinant domain (F3 of NH36 was evaluated using peripheral blood mononuclear cells (PBMC from individuals infected with L. (L. infantum from an endemic area of visceral leishmaniasis of Spain. Both NH36 and F1 domains significantly increased the PBMC proliferation stimulation index of cured patients and infected asymptomatic individuals compared to healthy controls. Moreover, F1 induced a 19% higher proliferative response than NH36 in asymptomatic exposed subjects. In addition, in patients cured from visceral leishmaniasis, proliferation in response to NH36 and F1 was accompanied by a significant increase of IFN-γ and TNF-α secretion, which was 42–43% higher, in response to F1 than to NH36. The interleukin 17 (IL-17 secretion was stronger in asymptomatic subjects, in response to F1, as well as in cured cutaneous leishmaniasis after NH36 stimulation. While no IL-10 secretion was determined by F1, a granzyme B increase was detected in supernatants from cured patients after stimulation with either NH36 or F1. These data demonstrate that F1 is the domain of NH36 that induces a recall cellular response in individuals with acquired resistance to the infection by L. (L. infantum. In addition, F1 and NH36 discriminated the IgG3 humoral response in patients with active visceral leishmaniasis due to L. (L. donovani (Ethiopia and L. (L. infantum (Spain from that of endemic and non-endemic area controls. NH36 showed higher reactivity with sera from L. (L. donovani-infected individuals, indicating species specificity. We conclude that the F1 domain, previously characterized as an inducer of the Th1 and Th17 responses in cured/exposed patients infected with L. (L. infantum chagasi, may

  6. Monocyte Chemotactic Protein 1 in Plasma from Soluble Leishmania Antigen-Stimulated Whole Blood as a Potential Biomarker of the Cellular Immune Response to Leishmania infantum

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    Ana V. Ibarra-Meneses

    2017-09-01

    Full Text Available New biomarkers are needed to identify asymptomatic Leishmania infection as well as immunity following vaccination or treatment. With the aim of finding a robust biomarker to assess an effective cellular immune response, monocyte chemotactic protein 1 (MCP-1 was examined in plasma from soluble Leishmania antigen (SLA-stimulated whole blood collected from subjects living in a Leishmania infantum-endemic area. MCP-1, expressed 110 times more strongly than IL-2, identified 87.5% of asymptomatic subjects and verified some asymptomatic subjects close to the cutoff. MCP-1 was also significantly elevated in all patients cured of visceral leishmaniasis (VL, unlike IL-2, indicating the specific memory response generated against Leishmania. These results show MCP-1 to be a robust candidate biomarker of immunity that could be used as a marker of cure and to both select and follow the population in vaccine phase I–III human clinical trials with developed rapid, easy-to-use field tools.

  7. Decreased antimony uptake and overexpression of genes of thiol metabolism are associated with drug resistance in a canine isolate of Leishmania infantum

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    Verónica Gómez Pérez

    2016-08-01

    Full Text Available Visceral leishmaniasis (VL caused by the protozoan parasite Leishmania infantum, is one of the most important zoonotic diseases affecting dogs and humans in the Mediterranean area. The presence of infected dogs as the main reservoir host of L. infantum is regarded as the most significant risk for potential human infection. We have studied the susceptibility profile to antimony and other anti-leishmania drugs (amphotericin B, miltefosine, paromomycin in Leishmania infantum isolates extracted from a dog before and after two therapeutic interventions with meglumine antimoniate (subcutaneous Glucantime®, 100 mg/kg/day for 28 days. After the therapeutic intervention, these parasites were significantly less susceptible to antimony than pretreatment isolate, presenting a resistance index of 6-fold to SbIII for promastigotes and >3-fold to SbIII and 3-fold to SbV for intracellular amastigotes. The susceptibility profile of this resistant L. infantum line is related to a decreased antimony uptake due to lower aquaglyceroporin-1 expression levels. Additionally, other mechanisms including an increase in thiols and overexpression of enzymes involved in thiol metabolism, such as ornithine decarboxylase, trypanothione reductase, mitochondrial tryparedoxin and mitochondrial tryparedoxin peroxidase, could contribute to the resistance as antimony detoxification mechanisms. A major contribution of this study in a canine L. infantum isolate is to find an antimony-resistant mechanism similar to that previously described in other human clinical isolates.

  8. Cytokine responses to novel antigens in a peri-urban population in Brazil exposed to Leishmania infantum chagasi.

    Science.gov (United States)

    Stober, Carmel B; Jeronimo, Selma M B; Pontes, Nubia N; Miller, E Nancy; Blackwell, Jenefer M

    2012-10-01

    Visceral leishmaniasis (VL) is fatal if untreated, and there are no vaccines for this disease. High levels of CD4-derived interferon-γ (IFN-γ) in the presence of low levels of interleukin-10 (IL-10) predicts vaccine success. Tumor necrosis factor-α (TNF-α) is also important in this process. We characterized human immune responses in three groups exposed to Leishmania infantum chagasi in Brazil: 1) drug-cured VL patients (recovered VL); 2) asymptomatic persons with positive Leishmania-specific delayed-type hypersensitivity skin reactions (DTH+); and 3) DTH-negative household contacts. Magnitude of DTH correlated with crude Leishmania antigen-driven IFN-γ, TNF-α, and IL-5, but not IL-10. DTH+ persons showed equivalent levels of IFN-γ, but higher levels of IL-10, to tryparedoxin peroxidase and Leishmania homolog of receptor for activated C kinase compared with recovered VL patients. The IFN-γ:IL-10 and TNF-α:IL-10 ratios were higher in recovered VL patients than in DTH+ persons. Seven of 11 novel candidates (R71, L37, N52, L302.06, M18, J41, and M22) elicited cytokine responses (36-71% of responders) in recovered VL patients and DTH+ persons. This result confirmed their putative status as cross-species vaccine/immunotherapeutic candidates.

  9. After infection with Leishmania infantum, Golden Hamsters (Mesocricetus auratus) become more attractive to female sand flies (Lutzomyia longipalpis).

    Science.gov (United States)

    Nevatte, T M; Ward, R D; Sedda, L; Hamilton, J G C

    2017-07-21

    In Brazil, human and canine visceral leishmaniasis is caused by infection with Leishmania infantum, a Protist parasite transmitted by blood-feeding female Lutzomyia longipalpis sand flies. The objective of this study was to determine if the odour of hamsters, infected with Le. infantum, was more attractive than the odour of the same hamsters, before they were infected. The attractiveness of odour collected from individual hamsters (n = 13), before they were infected, was compared in a longitudinal study, with the attractiveness of the odour of the same hamster in a Y-tube olfactometer bioassay, at a late stage of infection. The odour of six of the golden hamsters was significantly more attractive to 50% of the female sand flies at the end of infection compared to before infection and the odour of four of the golden hamsters was significantly more attractive to 75% of the female sand flies at the end of infection. These results strongly indicate that hamsters infected with Le. infantum become significantly more attractive to a greater proportion of female sand flies as the infection progresses.

  10. Inaccuracy of Enzyme-Linked Immunosorbent Assay Using Soluble and Recombinant Antigens to Detect Asymptomatic Infection by Leishmania infantum

    Science.gov (United States)

    Moreno, Elizabeth Castro; Gonçalves, Andréa Vieira; Chaves, Anderson Vieira; Melo, Maria Norma; Lambertucci, José Roberto; Andrade, Antero Silva Ribeiro; Negrão-Corrêa, Deborah; Antunes, Carlos Mauricio de Figueiredo; Carneiro, Mariângela

    2009-01-01

    Background One of the most important drawbacks in visceral leishmaniasis (VL) population studies is the difficulty of diagnosing asymptomatic carriers. The aim of this study, conducted in an urban area in the Southeast of Brazil, was to evaluate the performance of serology to identify asymptomatic VL infection in participants selected from a cohort with a two-year follow-up period. Methodology Blood samples were collected in 2001 from 136 cohort participants (97 positive and 39 negatives, PCR/hybridization carried out in 1999). They were clinically evaluated and none had progressed to disease from their asymptomatic state. As controls, blood samples from 22 control individuals and 8 patients with kala-azar were collected. Two molecular biology techniques (reference tests) were performed: PCR with Leishmania-generic primer followed by hybridization using L. infantum probe, and PCR with specific primer to L. donovani complex. Plasma samples were tested by ELISA using three different antigens: L. infantum and L. amazonensis crude antigens, and rK39 recombinant protein. Accuracy of the serological tests was evaluated using sensitivity, specificity, likelihood ratio and ROC curve. Findings The presence of Leishmania was confirmed, by molecular techniques, in all kala-azar patients and in 117 (86%) of the 136 cohort participants. Kala-azar patients showed high reactivity in ELISAs, whereas asymptomatic individuals presented low reactivity against the antigens tested. When compared to molecular techniques, the L. amazonensis and L. infantum antigens showed higher sensitivity (49.6% and 41.0%, respectively) than rK39 (26.5%); however, the specificity of rK39 was higher (73.7%) than L. amazonensis (52.6%) and L. infantum antigens (36.8%). Moreover, there was low agreement among the different antigens used (kappa<0.10). Conclusions Serological tests were inaccurate for diagnosing asymptomatic infections compared to molecular methods; this could lead to misclassification bias

  11. Substrate Preferences and Catalytic Parameters Determined by Structural Characteristics of Sterol 14[alpha]-Demethylase (CYP51) from Leishmania infantum

    Energy Technology Data Exchange (ETDEWEB)

    Hargrove, Tatiana Y.; Wawrzak, Zdzislaw; Liu, Jialin; Nes, W. David; Waterman, Michael R.; Lepesheva, Galina I. (Vanderbilt); (TTU); (NWU)

    2012-05-14

    Leishmaniasis is a major health problem that affects populations of {approx}90 countries worldwide, with no vaccine and only a few moderately effective drugs. Here we report the structure/function characterization of sterol 14{alpha}-demethylase (CYP51) from Leishmania infantum. The enzyme catalyzes removal of the 14{alpha}-methyl group from sterol precursors. The reaction is essential for membrane biogenesis and therefore has great potential to become a target for antileishmanial chemotherapy. Although L. infantum CYP51 prefers C4-monomethylated sterol substrates such as C4-norlanosterol and obtusifoliol (V{sub max} of {approx}10 and 8 min{sup -1}, respectively), it is also found to 14{alpha}-demethylate C4-dimethylated lanosterol (V{sub max} = 0.9 min{sup -1}) and C4-desmethylated 14{alpha}-methylzymosterol (V{sub max} = 1.9 min{sup -1}). Binding parameters with six sterols were tested, with K{sub d} values ranging from 0.25 to 1.4 {mu}m. Thus, L. infantum CYP51 is the first example of a plant-like sterol 14{alpha}-demethylase, where requirements toward the composition of the C4 atom substituents are not strict, indicative of possible branching in the postsqualene portion of sterol biosynthesis in the parasite. Comparative analysis of three CYP51 substrate binding cavities (Trypanosoma brucei, Trypanosoma cruzi, and L. infantum) suggests that substrate preferences of plant- and fungal-like protozoan CYP51s largely depend on the differences in the enzyme active site topology. These minor structural differences are also likely to underlie CYP51 catalytic rates and drug susceptibility and can be used to design potent and specific inhibitors.

  12. Host modulation by a parasite: how Leishmania infantum modifies the intestinal environment of Lutzomyia longipalpis to favor its development.

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    Vania Cristina Santos

    Full Text Available Some reports have described the interference of Leishmania on sand flies physiology, and such behavior most likely evolved to favor the development and transmission of the parasite. Most of these studies showed that Leishmania could modulate the level of proteases in the midgut after an infective blood meal, and decreased proteolytic activity is indeed beneficial for the development of promastigotes in the gut of sand flies. In the present study, we performed a detailed investigation of the intestinal pH in Lutzomyia longipalpis females naturally infected with Leishmania infantum and investigated the production of trypsin by these insects using different approaches. Our results allowed us to propose a mechanism by which these parasites interfere with the physiology of L. longipalpis to decrease the production of proteolytic enzymes. According to our hypothesis L. infantum promastigotes indirectly interfere with the production of trypsin by modulating the mechanism that controls the intestinal pH via the action of a yet non-identified substance released by promastigote forms inside the midgut. This substance is not an acid, whose action would be restrict on to release H+ to the medium, but is a substance that is able to interfere with midgut physiology through a mechanism involving pH control. According to our hypothesis, as the pH decreases, the proteolytic enzymes efficiency is also reduced, leading to a decline in the supply of amino acids to the enterocytes: this decline reduces the stimulus for protease production because it is regulated by the supply of amino acids, thus leading to a delay in digestion.

  13. Validity and reliability of enzyme immunoassays using Leishmania major or L. infantum antigens for the diagnosis of canine visceral leishmaniasis in Brazil.

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    Mauro Maciel de Arruda

    Full Text Available BACKGROUND: American visceral leishmaniasis is caused by the protozoan Leishmania infantum. Dogs are the main reservoirs in the domestic transmission cycle. The limited accuracy of diagnostic tests for canine leishmaniasis may contribute to the lack of impact of control measures recommended by the Brazilian Ministry of Health. The objective of this study was to estimate the accuracy of two enzyme-linked immunosorbent assays employing L. major or L. infantum antigens and their reliability between three laboratories of different levels of complexity. METHODS: A validation study of ELISA techniques using L. major or L. infantum antigens was conducted. Direct visualization of the parasite in hematoxylin/eosin-stained histopathological sections, immunohistochemistry, and isolation of the parasite in culture.were used as gold standard. An animal that was positive in at least one of the tests was defined as infected with L. infantum. Serum samples collected from 1,425 dogs were analyzed. Samples were separated in three aliquots and tested in three different laboratories. Sensitivity, specificity and the area under de ROC curve were calculated and the reliability was evaluated between the participant laboratories. RESULTS: The sensitivity was 91.8% and 89.8% for the L. major and L. infantum assays, respectively. The specificity was 83.75% and 82.7% for the L. major and L. infantum assays, respectively. The area under de ROC curve was 0.920 and 0.898 for L. major and L. infantum, respectively. The mean intraclass correlation coefficients between laboratories ranged from 0.890 to 0.948 when L. major was used as antigen, and from 0.818 to 0.879 when L. infantum was used. INTERPRETATION: ELISA tests using L. major or L. infantum antigens have similar accuracy and reliability. Our results do not support the substitution of the L. major antigen of the ELISA test currently used for the diagnosis of canine visceral leishmaniasis in Brazil.

  14. Visceral leishmaniasis caused by Leishmania infantum in a Spanish patient in Argentina: What is the origin of the infection? Case report

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    Salomón Oscar D

    2004-06-01

    Full Text Available Abstract Background The question "Where have you been?" is a common one asked by doctors in Northern Europe and America when faced with clinical symptoms not typical of their country. This question must also arise in the clinics of developing countries in which non-autochthonous cases such as the one described here can appear. Important outbreaks of Leishmania infantum have been recorded in the last decade in several Latin American countries but its presence has not yet been recorded in Argentina. We report the first case of visceral leishmaniasis owing to L. infantum in this country. Case presentation A 71-year-old Spanish woman who has been living in Mendoza, Argentina, during the last 40 years presented with a history of high fever and shivering, anemia, leukopenia and splenomegaly over two years. Argentinian doctors did not suspect visceral leishmaniasis even when the histological analysis revealed the presence of "intracytoplasmatic spheroid particles compatible with fungal or parasitic infection". After a serious deterioration in her health, she was taken to Spain where she was evaluated and visceral leishmaniasis was established. Specific identification of the parasite was done by PCR-ELISA, isoenzyme electrophoresis and RAPD-PCR. Conclusion We would like to point out that: i cases such as the one described here, which appear in non-endemic areas, can pass unnoticed by the clinical physician. ii in countries in which these introduced cases reside, in-depth parasitological studies are required into vectors and possible reservoirs to rule out the rare case of local infection and, once infection has taken place, to ensure that this does not spread by anthroponotic transmission or a competent reservoir.

  15. Comparative real-time kinetic analysis of human complement killing of Leishmania infantum promastigotes derived from axenic culture or from Phlebotomus perniciosus.

    Science.gov (United States)

    Moreno, Inmaculada; Molina, Ricardo; Toraño, Alfredo; Laurin, Edurne; García, Esther; Domínguez, Mercedes

    2007-01-01

    Although Leishmania metacyclic promastigotes are generally considered resistant to human complement, studies of in vitro-cultured axenic stationary promastigotes using serum concentrations that approximate physiological plasma conditions indicate complement sensitivity. Natural Leishmania infection is caused by sand fly-inoculated promastigotes, whose complement resistance has not been analyzed systematically. We compared Leishmania susceptibility to human complement in L. infantum promastigotes derived from in vitro cultures and from sand flies. Phlebotomus perniciosus sand flies were fed with axenic promastigotes, L. infantum-infected U-937 cells, or spleen cells from L. infantum-infected hamsters. On selected days post-feeding, flies were dissected and promastigotes isolated; in addition, axenic promastigotes were obtained from culture at equivalent days of growth. In near-physiological serum concentration and temperature conditions, measurement of real-time kinetics of propidium iodide uptake showed that approximately 90% of axenic- and sand fly-derived promastigotes were rapidly killed by complement. We found no substantial differences between promastigotes from axenic culture, those isolated from flies on different post-feeding days, or those generated in flies fed with distinct inocula. The results indicate that Leishmania susceptibility to human complement is independent of promastigote developmental stage in the sand fly mid-gut and in axenic culture.

  16. Clinical forms of canine visceral Leishmaniasis in naturally Leishmania infantum-infected dogs and related myelogram and hemogram changes.

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    Roney de Carvalho Nicolato

    Full Text Available Hematological analysis has limited applications for disease diagnosis in Leishmania infantum-infected dogs, but it can be very important in evaluating the clinical forms of the disease and in understanding the evolution of canine visceral leishmaniasis (CVL pathogenesis. Recently, we demonstrated that alterations in leucopoiesis and erythropoiesis are related to clinical status and bone marrow parasite density in dogs naturally infected by L. infantum. To further characterize these alterations, we evaluated the association between the hematological parameters in bone marrow and peripheral blood alterations in groups of L. infantum-infected dogs: asymptomatic I (AD-I: serum negative/PCR+, asymptomatic II (AD-II: serum positive, oligosymptomatic (OD, and symptomatic (SD. Results were compared with those from noninfected dogs (NID. The SD group was found to present a decrease in erythropoietic lineage with concomitant reductions in erythrocytes, hemoglobin, and hematocrit parameters, resulting in anemia. The SD group also had increased neutrophils and precursors and decreased band eosinophils and eosinophils, leading to peripheral blood leucopenia. In the AD-II group, lymphocytosis occurred in both the peripheral blood and the bone marrow compartments. The SD group exhibited lymphocytosis in the bone marrow, with lymphopenia in the peripheral blood. In contrast, the AD-I group, showed no significant changes suggestive of CVL, presenting normal counts in bone marrow and peripheral blood. Our results showed for the first time that important changes in hematopoiesis and hematological parameters occur during ongoing CVL in naturally infected dogs, mainly in symptomatic disease. Taken together, our results based on myelogram and hemogram parameters enable better understanding of the pathogenesis of the anemia, lymphocytosis, and lymphopenia, as well as the leucopenia (eosinopenia and monocytopenia, that contribute to CVL prognosis.

  17. Increased thiol levels in antimony-resistant Leishmania infantum isolated from treatment-refractory visceral leishmaniasis in Brazil.

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    Magalhães, Lucas S; Bomfim, Lays Gs; Mota, Sthefanne G; Cruz, Geydson S; Corrêa, Cristiane B; Tanajura, Diego M; Lipscomb, Michael W; Borges, Valéria M; Jesus, Amélia R de; Almeida, Roque P de; Moura, Tatiana R de

    2018-02-01

    BACKGROUND Treatment-refractory visceral leishmaniasis (VL) has become an important problem in many countries. OBJECTIVES We evaluated the antimony-resistance mechanisms of Leishmania infantum isolated from VL patients refractory or responsive to treatment with pentavalent antimony. METHODS Strains isolated from antimony-refractory patients (in vitro antimony-resistant isolates) and antimony-responsive patients (in vitro antimony-sensitive isolates) were examined. Morphological changes were evaluated by transmission electron microscopy after trivalent antimony exposure. P-glycoprotein (P-gp) efflux pump activity was evaluated using the pump-specific inhibitor verapamil hydrochloride, and the role of thiol in trivalent antimony resistance was investigated using the enzymatic inhibitor L-buthionine sulfoximine. FINDINGS Antimony treatment induced fewer alterations in the cellular structure of L. infantum resistant isolates than in that of sensitive isolates. P-gp efflux activity was not involved in antimony resistance in these isolates. Importantly, the resistant isolates contained higher levels of thiol compared to the sensitive isolates, and inhibition of thiol synthesis in the resistant isolates recovered their sensitivity to trivalent antimony treatment, and enhanced the production of reactive oxygen species in promastigotes exposed to the drug. MAIN CONCLUSIONS Our results demonstrate that isolates from patients with antimony-refractory VL exhibited higher thiol levels than antimony-sensitive isolates. This indicates that redox metabolism plays an important role in the antimony-resistance of New World VL isolates.

  18. Interleukin-27 Early Impacts Leishmania infantum Infection in Mice and Correlates with Active Visceral Disease in Humans

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    Begoña Pérez-Cabezas

    2016-11-01

    Full Text Available The complexity of Leishmania-host interactions, one of the main leishmaniasis issues, is yet to be fully understood. We detected elevated IL-27 plasma levels in European patients with active visceral disease caused by Leishmania infantum, which returned to basal levels after successful treatment, suggesting this cytokine as a probable infection mediator. We further addressed this hypothesis recurring to two classical susceptible visceral leishmaniasis mouse models. BALB/c, but not C57BL/6 mice, showed increased IL-27 systemic levels after infection, which was associated with an upregulation of IL-27p28 expression by dendritic cells and higher parasite burdens. Neutralization of IL-27 in acutely infected BALB/c led to decreased parasite burdens and a transient increase in IFN-γ+ splenic T cells, while administration of IL-27 to C57BL/6 promoted a local anti-inflammatory cytokine response at the site of infection and increased parasite loads. Overall we show that, as in humans, BALB/c IL-27 systemic levels are infection-dependently upregulated and may favor parasite installation by controlling inflammation.

  19. Dynamics and Predictive Potential of Antibodies against Insect-Derived Recombinant Leishmania infantum Proteins during Chemotherapy of Naturally Infected Dogs

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    Todolí, Felicitat; Galindo, Inmaculada; Gómez-Sebastián, Silvia; Pérez-Filgueira, Mariano; Escribano, José M.; Alberola, Jordi; Rodríguez-Cortés, Alhelí

    2010-01-01

    A predictive marker for the success treatment of canine leishmaniasis is required for the application of a more rational therapy protocol, which must improve the probability of cure and reduce Leishmania resistance to drugs. We investigated the dynamics and predictive value of antibodies against insect-derived recombinant L. infantum proteins rKMPII and rTRYP by using an enzyme-linked immunosorbent assay with retrospective serum samples from 36 dogs during treatment of canine leishmaniasis. In the entire group of dogs, concentrations of antibodies against rKMPII and rTRYP significantly decreased earlier than concentrations of antibodies against crude total Leishmania antigen (one versus six months), which suggested that the dynamics of antibodies against recombinant proteins may be useful for assessing clinical improvement after treatment. Interestingly, decreases in antibody concentrations against rKMPII occurred earlier in disease-free dogs than in dogs that remain clinically ill one year after beginning of treatment, which suggested that these antibodies may be useful for predicting disease-free survival one year after the beginning of therapy against canine leishmaniasis. PMID:20439957

  20. The Effect of Removing Potentially Infectious Dogs on the Numbers of Canine Leishmania infantum Infections in an Endemic Area with High Transmission Rates

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    Grimaldi, Gabriel; Teva, Antonio; Santos, Claudiney B.; Ferreira, Adelson L.; Falqueto, Aloísio

    2012-01-01

    To assess the effect of the rapid removal of potentially infectious dogs on the prevalence and incidence of canine infections, a prospective study was undertaken in an area endemic for Leishmania infantum. We used serological testing based on the rapid DPP rK28 fusion protein chromatographic immunoassay for this dog screening-and-culling intervention trial. The outcome was evaluated by measuring seropositivity and sero-conversion/-reversion rates for canine infection. Our estimates indicated ...

  1. Leishmaniosis due to Leishmania infantum in a FIV and FeIV positive cat with a squamous cell carcinoma diagnosed with histological, serological and isoenzymatic methods

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    Grevot A.

    2005-09-01

    Full Text Available Leishmaniosis caused by Leishmania infantum is an endemic zoonosis present in the Mediterranean area. Canidae (dog and fox constitute the main reservoir hosts for the parasite, whilst wild rodents or the cat can be carriers of the protozoan and are considered as secondary potential reservoirs. This paper describes a case of disseminated feline leishmaniosis with cutaneous (ulcerative, visceral (spleen and lymph nodes and blood involvement in a FIV-FelV positive cat. The microscopic identification of the Leishmania infection was initially made on a skin biopsy of the temporal area, where a squamous cell carcinoma was diagnosed. The diagnosis of the disease was achieved by several serological techniques (ELISA, IFAT and Western-blot. The strain was obtained by blood culture, characterized by electrophoresis of isoenzymes and identified as Leishmania infantum zymodeme MON-1. Since the infection due to L. infantum is a zoonosis, the potential feline reservoir should be more investigated. Serological analysis by Western blot on domestic cats provides a useful tool. In veterinary practice, feline leishmaniosis should be systematically included in the differential diagnosis when compatible cutaneous lesions are present, especially in the endemic areas of canine leishmaniosis.

  2. Survey of Wild and Domestic Mammals for Infection with Leishmania infantum following an Outbreak of Desert Zoonotic Visceral Leishmaniasis in Jiashi, People's Republic of China.

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    Chun-Hua Gao

    Full Text Available In 2008 and 2009, an outbreak of desert-subtype zoonotic visceral leishmaniasis occurred in Jiashi county, Xinjiang, China. So far, no animal reservoir has been identified for this type of visceral leishmaniasis. Therefore, we surveyed the most common mammals (wild and domestic for Leishmania infections during the outbreak in 2008 and 2009 in order to identify the source of the visceral leishmaniasis in this region. Spleen, liver, bone marrow and blood samples collected from 86 wood mice (Apodemus sylvaticus, 61midday jirds (Meriones meridianus and 27 Yarkand hares (Lepus yarkandensis were tested for the presence of Leishmania by microscopy, culture and PCR. All of the animals were found to be negative for Leishmania infections; On the other hand, Leishmania DNA was detected in blood samples collected from livestock reared in the outbreak area: 30.36% (17/56 of sheep, 21.57% (11/51 of goats, 17.78% (8/45 of cattle, and 21.62 (8/37 of donkeys were positive for Leishmania DNA by PCR. The amplified kDNA sequences from the livestock samples matched Leishmania DNA sequences isolated from patients with visceral leishmaniasis in the study area. We suggest that these domestic mammals are a possible reservoir host for Leishmania infantum in the outbreak area.

  3. Seasonality of sand flies (Diptera: Psychodidae) and Leishmania DNA detection in vector species in an area with endemic visceral leishmaniasis.

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    Saraiva, Lara; Leite, Camila Gonçalves; Lima, Ana Cristina Vianna Mariano da Rocha; Carvalho, Luiz Otávio Alves de; Pereira, Agnes Antônia Sampaio; Rugani, Jerônimo Marteleto Nunes; Rego, Felipe Dutra; Gontijo, Célia Maria Ferreira; Andrade, José Dilermando

    2017-04-01

    Leishmaniases are a serious health problem in southeast Brazil, including the city of Belo Horizonte (BH), Minas Gerais state (MG), where there are high rates of incidence and mortality due to visceral leishmaniases. BH is divided into nine sanitary districts (SD) of which one, the Venda Nova SD, was selected for this study because it has high rates of positivity for canine leishmaniasis and high incidence of human leishmaniasis. This study aimed to survey the sand fly fauna in Venda Nova SD from August 2011 to July 2013 and perform a descriptive analysis of the vector population. The sampling was carried out using automatic HP light traps at all covered areas of the Venda Nova SD, in a total of eighteen light traps. Sampled specimens were identified following Galati (2003), and females were submitted to molecular techniques for the detection and identification of Leishmania DNA. A simple environmental description was done for it area and Kernel estimation was used to infer vector density for each study site. A total of 2,427 sand fly specimens belonging to eight species and five genera were collected of which 95.3% were Lutzomyia longipalpis. The seasonal variation curve was delineated by this species. Lu. longipalpis was the most abundant at all collection points and in all months of the study, and exhibited a natural infection rate of 1.01% for Leishmania infantum and 1.77% for Leishmania braziliensis. The results show the presence and adaptation of Lu. longipalpis to the anthropic environment of BH and reinforces its role as the main vector of L. infantum in the region.

  4. Seasonality of sand flies (Diptera: Psychodidae) and Leishmania DNA detection in vector species in an area with endemic visceral leishmaniasis

    Science.gov (United States)

    Saraiva, Lara; Leite, Camila Gonçalves; Lima, Ana Cristina Vianna Mariano da Rocha; de Carvalho, Luiz Otávio Alves; Pereira, Agnes Antônia Sampaio; Rugani, Jerônimo Marteleto Nunes; Rego, Felipe Dutra; Gontijo, Célia Maria Ferreira; Andrade, José Dilermando

    2017-01-01

    BACKGROUND Leishmaniases are a serious health problem in southeast Brazil, including the city of Belo Horizonte (BH), Minas Gerais state (MG), where there are high rates of incidence and mortality due to visceral leishmaniases. BH is divided into nine sanitary districts (SD) of which one, the Venda Nova SD, was selected for this study because it has high rates of positivity for canine leishmaniasis and high incidence of human leishmaniasis. OBJECTIVES This study aimed to survey the sand fly fauna in Venda Nova SD from August 2011 to July 2013 and perform a descriptive analysis of the vector population. METHODS The sampling was carried out using automatic HP light traps at all covered areas of the Venda Nova SD, in a total of eighteen light traps. Sampled specimens were identified following Galati (2003), and females were submitted to molecular techniques for the detection and identification of Leishmania DNA. A simple environmental description was done for it area and Kernel estimation was used to infer vector density for each study site. FINDINGS A total of 2,427 sand fly specimens belonging to eight species and five genera were collected of which 95.3% were Lutzomyia longipalpis. The seasonal variation curve was delineated by this species. Lu. longipalpis was the most abundant at all collection points and in all months of the study, and exhibited a natural infection rate of 1.01% for Leishmania infantum and 1.77% for Leishmania braziliensis. MAIN CONCLUSIONS The results show the presence and adaptation of Lu. longipalpis to the anthropic environment of BH and reinforces its role as the main vector of L. infantum in the region. PMID:28327794

  5. A novel Leishmania infantum nuclear phosphoprotein Lepp12 which stimulates IL1-beta synthesis in THP-1 transfectants

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    Mograbi Baharia

    2003-04-01

    Full Text Available Abstract Background We report cloning and characterization of a novel Leishmania infantum protein which we termed Lepp12, and we examine its possible implication in the interference with intramacrophage signaling pathways. Results The protein Lepp12 contains 87 amino acid sequence and exhibits 5 potential phosphorylation sites by protein kinase C (PKC. Recombinant GST-Lepp12 is phosphorylated in vitro by exogenous PKC and by PKC-like activities present in promastigote and in the myelomonocytic THP-1 cell line, indicating that at least one phosphorylation site is functional on the recombinant Lepp12. The natural Lepp12 protein is present in L. infantum promastigotes, as evidenced using specific anti-Lepp12 antibodies produced by immunopurification from acute phase VL patient sera. Interestingly, human patient sera are strongly reactive with GST-Lepp12, demonstrating immunogenic properties of Lepp12 in man, but no immune response to Lepp12 is detectable in experimentally infected animals. When isolated from promastigotes, Lepp12 migrates as two species of apparent MW of 18.3 kDa (major and 14 kDa (minor, localizes in the nuclear fraction and appears constitutively phosphorylated. Natural Lepp12 is phosphorylable in vitro by both exogenous PKC and PKC-like activity present in THP-1 extracts. The intracellular Lepp12 transfected into THP-1 cells activates these cells to produce IL-1beta and induces an enhancing effect on PMA stimulated IL-1beta synthesis, as demonstrated using GST-Lepp12 transfectants. Conclusions Together these results indicate that Lepp12 represents a substrate for PKC or other PKC-like activities present in the promastigote form and the host cell and therefore may interfere with signal transduction pathways involving PKC.

  6. Cyclobenzaprine Raises ROS Levels in Leishmania infantum and Reduces Parasite Burden in Infected Mice.

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    Edézio Ferreira Cunha-Júnior

    2017-01-01

    Full Text Available The leishmanicidal action of tricyclic antidepressants has been studied and evidences have pointed that their action is linked to inhibition of trypanothione reductase, a key enzyme in the redox metabolism of pathogenic trypanosomes. Cyclobenzaprine (CBP is a tricyclic structurally related to the antidepressant amitriptyline, differing only by the presence of a double bond in the central ring. This paper describes the effect of CBP in experimental visceral leishmaniasis, its inhibitory effect in trypanothione reductase and the potential immunomodulatory activity.In vitro antileishmanial activity was determined in promastigotes and in L. infantum-infected macrophages. For in vivo studies, L. infantum-infected BALB/c mice were treated with CBP by oral gavage for five days and the parasite load was estimated. Trypanothione reductase activity was assessed in the soluble fraction of promastigotes of L. infantum. For evaluation of cytokines, L. infantum-infected macrophages were co-cultured with BALB/c splenocytes and treated with CBP for 48 h. The supernatant was analyzed for IL-6, IL-10, MCP-1, IFN-γ and TNF-α. CBP demonstrated an IC50 of 14.5±1.1μM and an IC90 of 74.5±1.2 μM in promastigotes and an IC50 of 12.6±1.05 μM and an IC90 of 28.7±1.3 μM in intracellular amastigotes. CBP also reduced the parasite load in L. infantum-infected mice by 40.4±10.3% and 66.7±10.5% in spleen at 24.64 and 49.28 mg/kg, respectively and by 85.6±5.0 and 89.3±4.8% in liver at 24.64 and 49.28mg/kg, after a short-term treatment. CBP inhibited the trypanothione reductase activity with a Ki of 86 ± 7.7 μM and increased the ROS production in promastigotes. CBP inhibited in 53% the production of IL-6 in infected macrophages co-culture.To the best of our knowledge, this study is the first report of the in vivo antileishmanial activity of the FDA-approved drug CBP. Modulation of immune response and induction of oxidative stress in parasite seem to contribute to

  7. Cross-species genetic exchange between visceral and cutaneous strains of Leishmania in the sand fly vector.

    Science.gov (United States)

    Romano, Audrey; Inbar, Ehud; Debrabant, Alain; Charmoy, Melanie; Lawyer, Phillip; Ribeiro-Gomes, Flavia; Barhoumi, Mourad; Grigg, Michael; Shaik, Jahangheer; Dobson, Deborah; Beverley, Stephen M; Sacks, David L

    2014-11-25

    Genetic exchange between Leishmania major strains during their development in the sand fly vector has been experimentally shown. To investigate the possibility of genetic exchange between different Leishmania species, a cutaneous strain of L. major and a visceral strain of Leishmania infantum, each bearing a different drug-resistant marker, were used to coinfect Lutzomyia longipalpis sand flies. Eleven double-drug-resistant progeny clones, each the product of an independent mating event, were generated and submitted to genotype and phenotype analyses. The analysis of multiple allelic markers across the genome suggested that each progeny clone inherited at least one full set of chromosomes from each parent, with loss of heterozygosity at some loci, and uniparental retention of maxicircle kinetoplast DNA. Hybrids with DNA contents of approximately 2n, 3n, and 4n were observed. In vivo studies revealed clear differences in the ability of the hybrids to produce pathology in the skin or to disseminate to and grow in the viscera, suggesting polymorphisms and differential inheritance of the gene(s) controlling these traits. The studies, to our knowledge, represent the first experimental confirmation of cross-species mating in Leishmania, opening the way toward genetic linkage analysis of important traits and providing strong evidence that genetic exchange is responsible for the generation of the mixed-species genotypes observed in natural populations.

  8. Situación epidemiológica y de los factores de riesgo de transmisión de Leishmania infantum en España

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    Berta Suárez Rodríguez

    2012-01-01

    Full Text Available En España la leishmaniasis es una zoonosis endémica presente en la mayor parte del territorio peninsular e Islas Baleares. El parásito que se detecta es la Leishmania infantum y el principal reservorio es el perro. Los vectores competentes para transmitir el parásito son los flebotomos, de los que existen dos especies distribuidas por todo el territorio peninsular e Islas Baleares. En este artículo se revisa la situación actual y el comportamiento de la leishmaniasis en España. Se analiza y compara la información sobre la enfermedad procedente de las altas hospitalarias y de la Red Nacional de Vigilancia Epidemiológica así como los factores de riesgo identificados en estudios anteriores que facilitan la transmisión. Desde 2009 está teniendo lugar un brote en el suroeste de la Comunidad de Madrid, destacando el elevado número de casos, cerca de 400 y la detección de un nuevo reservorio, la liebre. Las condiciones que pueden incrementar la incidencia de la leishmaniasis están asociadas al aumento del número de vectores infectados, al de la densidad del flebotomo, al de hospedadores o la aparición de nuevos reservorios en un área geográfica. Esta infección tiene una dinámica de transmisión compleja y para responder oportunamente a los brotes debe estar previamente desarrollado un plan de acción integral y multisectorial en el que participen todas las instituciones involucradas. Ante un brote, es importante estudiar los factores que condicionan la transmisión del lugar y adaptar el plan de acción a sus características específicas.

  9. Clinical management of canine leishmaniosis versus human leishmaniasis due to Leishmania infantum: Putting "One Health" principles into practice.

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    Miró, Guadalupe; López-Vélez, Rogelio

    2018-04-30

    The initiative One World, "One Health" tries to rapidly detect emerging or reemerging human and animal infectious diseases and prevent epidemiological situations such as deforestation, some agricultural practices or the appearance of new foci of leishmaniosis due to Leishmania infantum with alternative reservoirs. With this objective in mind, we here consider leishmaniosis in the Mediterranean basin and compare its current clinical management from two perspectives: that of a veterinarian specialized in infectious and parasitic diseases, and that of a physician specialized in infectious tropical diseases. We thus prepared a list of 10 key questions from epidemiology to control of the disease in both species: dogs and humans. This issue requires a concise and clear response to help animal and human health clinicians to improve their clinical management and understanding of this important zoonosis. Our ultimate aim is to update and bring together the information available backed by sound scientific evidence. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

  10. Serum Cytokine Responses over the Entire Clinical-Immunological Spectrum of Human Leishmania (L. infantum chagasi Infection

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    Patrícia Karla Ramos

    2016-01-01

    Full Text Available The clinical-immunological spectrum of human Leishmania (L. infantum chagasi infection in Amazonian Brazil was recently reviewed based on clinical, DTH, and IFAT (IgG evaluations that identified five profiles: three asymptomatic (asymptomatic infection, AI; subclinical resistant infection, SRI; and indeterminate initial infection, III and two symptomatic (symptomatic infection, SI; American visceral leishmaniasis, AVL; and subclinical oligosymptomatic infection, SOI. TNF-α, IL-4, IL-6, and IL-10 serum cytokines were analyzed using multiplexed Cytometric Bead Array in 161 samples from endemic areas in the Brazilian Amazon: SI [AVL] (21 cases, III (49, SRI (19, SOI (12, AI (36, and a control group [CG] (24. The highest IL-6 serum levels were observed in the SI profile (AVL; higher IL-10 serum levels were observed in SI than in SOI or CG and in AI and III than in SOI; higher TNF-α serum levels were seen in SI than in CG. Positive correlations were found between IL-6 and IL-10 serum levels in the SI and III profiles and between IL-6 and TNF-α and between IL-4 and TNF-α in the III profile. These results provide strong evidence for associating IL-6 and IL-10 with the immunopathogenesis of AVL and help clarify the role of these cytokines in the infection spectrum.

  11. Prevalence of Dirofilaria immitis, Ehrlichia canis, Borrelia burgdorferi sensu lato, Anaplasma spp. and Leishmania infantum in apparently healthy and CVBD-suspect dogs in Portugal - a national serological study

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    Cardoso Luís

    2012-03-01

    Full Text Available Abstract Background Canine vector-borne diseases (CVBDs are caused by a wide range of pathogens transmitted to dogs by arthropods including ticks and insects. Many CVBD-agents are of zoonotic concern, with dogs potentially serving as reservoirs and sentinels for human infections. The present study aimed at assessing the seroprevalence of infection with or exposure to Dirofilaria immitis, Ehrlichia canis, Borrelia burgdorferi sensu lato, Anaplasma spp. and Leishmania infantum in dogs in Portugal. Methods Based on 120 veterinary medical centres from all the regions of mainland and insular Portugal, 557 apparently healthy and 628 CVBD-suspect dogs were sampled. Serum, plasma or whole blood was tested for qualitative detection of D. immitis antigen and antibodies to E. canis, B. burgdorferi s. l., Anaplasma spp. and L. infantum with two commercial in-clinic enzyme-linked immunosorbent assay kits. Odds ratios (OR were calculated by logistic regression analysis to identify independent risk factors of exposure to the vector-borne agents. Results Total positivity levels to D. immitis, E. canis, B. burgdorferi, Anaplasma spp., L. infantum, one or more agents and mixed agents were 3.6%, 4.1%, 0.2%, 4.5%, 4.3%, 14.0% and 2.0% in the healthy group, and 8.9%, 16.4%, 0.5%, 9.2%, 25.2%, 46.3% and 11.6% in the clinically suspect group, respectively. Non-use of ectoparasiticides was a risk factor for positivity to one or more agents both in the apparently healthy (OR = 2.1 and CVBD-suspect (OR = 1.5 dogs. Seropositivity to L. infantum (OR = 7.6, E. canis (OR = 4.1 and D. immitis (OR = 2.4 were identified as risk factors for the presence of clinical signs compatible with CVBDs. Positivity to mixed agents was not found to be a risk factor for disease. Conclusions Dogs in Portugal are at risk of becoming infected with vector-borne pathogens, some of which are of zoonotic concern. CVBDs should be considered by practitioners and prophylactic measures must be put in

  12. Leishmania development in sand flies: parasite-vector interactions overview.

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    Dostálová, Anna; Volf, Petr

    2012-12-03

    Leishmaniases are vector-borne parasitic diseases with 0.9 - 1.4 million new human cases each year worldwide. In the vectorial part of the life-cycle, Leishmania development is confined to the digestive tract. During the first few days after blood feeding, natural barriers to Leishmania development include secreted proteolytic enzymes, the peritrophic matrix surrounding the ingested blood meal and sand fly immune reactions. As the blood digestion proceeds, parasites need to bind to the midgut epithelium to avoid being excreted with the blood remnant. This binding is strictly stage-dependent as it is a property of nectomonad and leptomonad forms only. While the attachment in specific vectors (P. papatasi, P. duboscqi and P. sergenti) involves lipophosphoglycan (LPG), this Leishmania molecule is not required for parasite attachment in other sand fly species experimentally permissive for various Leishmania. During late-stage infections, large numbers of parasites accumulate in the anterior midgut and produce filamentous proteophosphoglycan creating a gel-like plug physically obstructing the gut. The parasites attached to the stomodeal valve cause damage to the chitin lining and epithelial cells of the valve, interfering with its function and facilitating reflux of parasites from the midgut. Transformation to metacyclic stages highly infective for the vertebrate host is the other prerequisite for effective transmission. Here, we review the current state of knowledge of molecular interactions occurring in all these distinct phases of parasite colonization of the sand fly gut, highlighting recent discoveries in the field.

  13. Prevalence of antibodies to Trypanosoma cruzi, Leishmania infantum, Encephalitozoon cuniculi, Sarcocystis neurona, and Neospora caninum in Capybara, Hydrochoerus hydrochaeris, from São Paulo State, Brazil.

    Science.gov (United States)

    Valadas, Samantha; Gennari, Solange Maria; Yai, Lucia Eiko Oishi; Rosypal, Alexa C; Lindsay, David S

    2010-06-01

    Little is known about the importance of capybara, Hydrochoerus hydrochaeris, as reservoirs for parasites of zoonotic or veterinary importance. Sera from 63 capybaras, from 6 counties in the state of São Paulo, Brazil, were examined for antibodies to Trypanosoma cruzi, Leishmania infantum, Encephalitozoon cuniculi, Sarcocystis neurona, and Neospora caninum using an indirect immunofluorescent antibody test. Five (8%) of the 63 capybaras had antibodies to T. cruzi epimastigotes. None of the samples from capybara reacted positively with L. infantum promastigotes or with spores of E. cuniculi . Two (3%) of the serum samples were positive for antibodies to S. neurona merozoites, and 2 (3%) of the serum samples were positive for antibodies to N. caninum tachyzoites. A serum sample from 1 capybara was positive for antibodies to both T. cruzi and N. caninum. None of the remaining 62 samples reacted with more than 1 parasite.

  14. Human immunodeficiency virus/Leishmania infantum in the first foci of urban American visceral leishmaniasis: clinical presentation from 1994 to 2010

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    Iúri Paz Lima

    2013-04-01

    Full Text Available INTRODUCTION: Human immunodeficiency virus (HIV coinfection with Leishmania infantum or Leishmania donovani, the agents of visceral leishmaniasis (or kala-azar, has become a fatal public health problem in the tropics where kala-azar is endemic. METHODS: The clinical presentation of patients with HIV and L. infantum coinfection is described using two unique databases that together produce the largest case series of patients with kala-azar infected with HIV in South America. First, a retrospective study paired the list of all patients with kala-azar from 1994 to 2004 with another of all patients with HIV/AIDS from the reference hospital for both diseases in the City of Teresina, State of Piauí, Brazil. Beginning in 2005 through to 2010 this information was prospectively collected at the moment of hospitalization. RESULTS: During the study, 256 admissions related to 224 patients with HIV/L. infantum coinfection were registered and most of them were males between 20-40 years of age. Most of the 224 patients were males between 20-40 years of age. HIV contraction was principally sexual. The most common symptoms and signs were pallor, fever, asthenia and hepatosplenomegaly. 16.8% of the cohort died. The primary risk factors associated to death were kidney or respiratory failure, somnolence, hemorrhagic manifestations and a syndrome of systemic inflammation. The diagnosis of HIV and kala-azar was made simultaneously in 124 patients. CONCLUSIONS: The urban association between HIV and kala-azar coinfection in South America is worrisome due to difficulty in establishing the diagnosis and higher mortality among the coinfected then those with either disease independently. HIV/L. infantum coinfection exhibits some singular characteristics and due to its higher mortality it requires immediate assistance to patients and greater research on appropriate combination therapy.

  15. Evaluation of two modified culture media for Leishmania infantum cultivation versus different culture media.

    Science.gov (United States)

    Castelli, Germano; Galante, Antonella; Lo Verde, Vincenza; Migliazzo, Antonella; Reale, Stefano; Lupo, Tiziana; Piazza, Maria; Vitale, Fabrizio; Bruno, Federica

    2014-04-01

    The aim of this study is to improve the cultivation of Leishmania promastigotes without the use of common, semisolid culture media such as Evans' modified Tobie's medium (EMTM), liquid RPMI 1640, and Peptone-yeast extract medium (P-Y). Although EMTM medium permits the growth of a high number of parasites, it is technically difficult to prepare as it requires the use of fresh rabbit blood from animals bred on farms, while RPMI 1640 and P-Y show lower growth rates than the EMTM. There is, therefore, a need to develop new blood-free and time-saving culture systems. The aim of this paper is to propose new modified microbiological media, named RPMI-PY and Tobie-PY, to isolate Leishmania and cultivate parasites for research and diagnostic purposes. This study compares classic culture media to the new media, RPMI-PY and Tobie-PY, and demonstrates that the new media have superior performance in terms of time and parasitic load. The growth rate of the parasite was significantly higher at 24, 48, and 72 hr cultivation, based on counts using Bürker's chambers, when compared to classic media. This study was carried out at the National References Centre for Leishmaniasis (C.Re.Na.L.) where the isolation procedures are conducted daily from a number of different biological matrices.

  16. The current status of phlebotomine sandflies (Diptera: Psychodidae) in Tunisia and their role on Leishmania transmission: A review

    OpenAIRE

    Ahmed Tabbabi; Sajida Sboui; Jabeur Daaboub

    2017-01-01

    In Tunisia, the epidemiological situation of leishmaniasis is characterized by the coexistence in a rather circumscribed territory (165000 km2, including the Sahara) of 4 forms of leishmaniasis caused by 3 species: Leishmania infantum, Leishmania major and Leishmania tropica (L. tropica) (synonymous Leishmania killicki). One of the factors determining the clinical, epidemiological and immunological diversity of leishmanioses is certainly the existence of a vector-parasite specificity of of...

  17. A ready-to-use duplex qPCR to detect Leishmania infantum DNA in naturally infected dogs.

    Science.gov (United States)

    Rampazzo, Rita de Cássia Pontello; Solcà, Manuela da Silva; Santos, Liliane Celestino Sales; Pereira, Lais de Novaes; Guedes, José Carlos Oliveira; Veras, Patrícia Sampaio Tavares; Fraga, Deborah Bittencourt Mothé; Krieger, Marco Aurélio; Costa, Alexandre Dias Tavares

    2017-11-15

    Canine visceral leishmaniasis (CVL) is a systemic disease caused by Leishmania infantum. A precise CVL diagnosis would allow for a faster and more specific treatment. Quantitative PCR (qPCR) is a sensitive and specific technique that can diagnose CVL and also monitor parasite load in the animal during the course of the infection or treatment. The aim of this study was to develop a ready-to-use (gelified and freezer-free) duplex qPCR for the identification of infected animals. We combined a new qPCR protocol that detects the canine 18S rRNA gene with an existing protocol for L. infantum kDNA detection, creating a duplex qPCR. This duplex method was then developed into a ready-to-use format. The performance of the duplex and singleplex reactions were compared in the traditional format (liquid and freezer-stored). Furthermore, the duplex qPCR performance was compared between the ready-to-use and traditional formats. The singleplex and new duplex qPCR exhibited the same detection limit in the traditional format (0.1 parasites/reaction). The ready-to-use format showed a detection limit of 1 parasite/reaction without affecting the reaction efficiency. The performance of the new qPCR protocol in the two formats was assessed using canine tissue samples from 82 dogs in an endemic CVL area that were previously characterized by standard serological and parasitological protocols. Splenic aspirates provided a higher rate of positivity (92.9%) followed by skin (50%) and blood (35.7%). The reported detection limits were observed for all tissues studied. Our results show that the amplification of L. infantum kDNA and canine DNA in a single tube, using either the traditional or ready-to-use format, exhibited the same diagnostic performance as amplification of the parasite kDNA alone. The detection of the host gene strengthens the qPCR results by confirming the presence and quality of DNA in the samples and the absence of polymerase inhibitors. The ready-to-use duplex qPCR format

  18. Population genetics of Leishmania infantum in Israel and the Palestinian Authority through microsatellite analysis.

    Science.gov (United States)

    Amro, Ahmad; Schönian, Gabriele; Al-Sharabati, Mohamed Barakat; Azmi, Kifaya; Nasereddin, Abedelmajeed; Abdeen, Ziad; Schnur, Lionel F; Baneth, Gad; Jaffe, Charles L; Kuhls, Katrin

    2009-04-01

    Multilocus microsatellite typing (MLMT) was used to investigate the genetic variation among 44 Israeli and Palestinian strains of L. infantum isolated from infected dogs and human cases to determine their population structure and to compare them with strains isolated from different European countries. Most of the Israeli and Palestinian strains had their own individual MLMT profiles; a few shared the same profile. A Bayesian model-based approach and phylogenetic reconstructions based on genetic distances inferred two main populations that were significantly different from the European strains: population A, containing 16 strains from places in the West Bank and 11 strains from central Israel;and population B, containing 7 strains from northern Israel, 9 from central Israel, and one Palestinian strain from the Jenin District.Geographically distributed sub-populations were detected within population B. These results demonstrate similar disease dynamics in Israel and the Palestinian Authority. The re-emergence of VL in the case of population A is more likely owing to increased dog and human contact with sylvatic cycles of parasitic infection rather than to recent introduction from the older foci of northern Israel. The latter scenario could be true for population B found in few foci of Central Israel. (c) 2009 Elsevier Masson SAS. All rights reserved.

  19. Molecular detection of Leishmania infection due to Leishmania major and Leishmania turanica in the vectors and reservoir host in Iran.

    Science.gov (United States)

    Rassi, Yavar; Oshaghi, Mohammad Ali; Azani, Sadegh Mohammadi; Abaie, Mohammad Reza; Rafizadeh, Sina; Mohebai, Mehdi; Mohtarami, Fatemeh; Zeinali, Mohammad kazem

    2011-02-01

    An epidemiological study was carried out on the vectors and reservoirs of cutaneous leishmaniasis in rural areas of Damghan district, Semnan province, central Iran, during 2008-2009. Totally, 6110 sand flies were collected using sticky papers and were subjected to molecular methods for detection of Leishmania parasite. Phlebotomus papatasi Scopoli was the common species in outdoor and indoor resting places. Polymerase chain reaction technique showed that 24 out of 218 P. papatasi (11%) and 4 out of 62 Phlebotomus caucasicus Marzinovskyi (6.5%) were positive for parasites Leishmania major Yakimoff and Schokhor. Twenty-one rodent reservoir hosts captured using Sherman traps were identified as Rhombomys opimus Lichtenstein (95%) and Meriones libycus Lichtenstein (5%). Microscopic investigation on blood smear of the animals for amastigote parasites revealed 8 (40%) rodents infected with R. opimus. L. major infection in these animals was then confirmed by polymerase chain reaction against internal transcribed spacer ribosomal DNA (rDNA) loci of the parasite followed by restriction fragment length polymorphism. Further, sequence analysis of 297 bp of ITS1-rDNA loci revealed the presence of L. major and Leishmania turanica in P. papatasi, and L. major in R. opimus. This is the first molecular report of L. major infection in both vectors (P. papatasi and P. caucasicus) and reservoir host (R. opimus) in this region. The results indicated that P. papatas was the primary vector of the disease and circulating the parasite between human and reservoirs, and P. caucasicus could be considered as a secondary vector. Further, our study showed that R. opimus is the most important host reservoir for maintenance of the parasite source in the area.

  20. LiZIP3 is a cellular zinc transporter that mediates the tightly regulated import of zinc in Leishmania infantum parasites

    Science.gov (United States)

    Carvalho, Sandra; da Silva, Rosa Barreira; Shawki, Ali; Castro, Helena; Lamy, Márcia; Eide, David; Costa, Vítor; Mackenzie, Bryan; Tomás, Ana M.

    2016-01-01

    Summary Cellular zinc homeostasis ensures that the intracellular concentration of this element is kept within limits that enable its participation in critical physiological processes without exerting toxic effects. We report here the identification and characterization of the first mediator of zinc homeostasis in Leishmania infantum, LiZIP3, a member of the ZIP family of divalent metal-ion transporters. The zinc transporter activity of LiZIP3 was first disclosed by its capacity to rescue the growth of Saccharomyces cerevisiae strains deficient in zinc acquisition. Subsequent expression of LiZIP3 in Xenopus laevis oocytes was shown to stimulate the uptake of a broad range of metal ions, among which Zn2+ was the preferred LiZIP3 substrate (K0.5 ≈ 0.1 μM). Evidence that LiZIP3 functions as a zinc importer in L. infantum came from the observations that the protein locates to the cell membrane and that its overexpression leads to augmented zinc internalization. Importantly, expression and cell-surface location of LiZIP3 are lost when parasites face high zinc bioavailability. LiZIP3 decline in response to zinc is regulated at the mRNA level in a process involving (a) short-lived protein(s). Collectively, our data reveal that LiZIP3 enables L. infantum to acquire zinc in a highly regulated manner, hence contributing to zinc homeostasis. PMID:25644708

  1. HP-Lattice QSAR for dynein proteins: experimental proteomics (2D-electrophoresis, mass spectrometry) and theoretic study of a Leishmania infantum sequence.

    Science.gov (United States)

    Dea-Ayuela, María Auxiliadora; Pérez-Castillo, Yunierkis; Meneses-Marcel, Alfredo; Ubeira, Florencio M; Bolas-Fernández, Francisco; Chou, Kuo-Chen; González-Díaz, Humberto

    2008-08-15

    The toxicity and inefficacy of actual organic drugs against Leishmaniosis justify research projects to find new molecular targets in Leishmania species including Leishmania infantum (L. infantum) and Leishmaniamajor (L. major), both important pathogens. In this sense, quantitative structure-activity relationship (QSAR) methods, which are very useful in Bioorganic and Medicinal Chemistry to discover small-sized drugs, may help to identify not only new drugs but also new drug targets, if we apply them to proteins. Dyneins are important proteins of these parasites governing fundamental processes such as cilia and flagella motion, nuclear migration, organization of the mitotic splinde, and chromosome separation during mitosis. However, despite the interest for them as potential drug targets, so far there has been no report whatsoever on dyneins with QSAR techniques. To the best of our knowledge, we report here the first QSAR for dynein proteins. We used as input the Spectral Moments of a Markov matrix associated to the HP-Lattice Network of the protein sequence. The data contain 411 protein sequences of different species selected by ClustalX to develop a QSAR that correctly discriminates on average between 92.75% and 92.51% of dyneins and other proteins in four different train and cross-validation datasets. We also report a combined experimental and theoretic study of a new dynein sequence in order to illustrate the utility of the model to search for potential drug targets with a practical example. First, we carried out a 2D-electrophoresis analysis of L. infantum biological samples. Next, we excised from 2D-E gels one spot of interest belonging to an unknown protein or protein fragment in the region Mdata base with the highest similarity score to the MS of the protein isolated from L. infantum. We used the QSAR model to predict the new sequence as dynein with probability of 99.99% without relying upon alignment. In order to confirm the previous function annotation we

  2. The Seminested PCR Based Detection of Leishmania infantum Infection in Asymptomatic Dogs in a New Endemic Focus of Visceral Leishmaniasis in Iran

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    Y Rassi

    2007-06-01

    Full Text Available Visceral Leishmaniasis (Kala-azar is a serious health problem in some northern and south western parts of Iran. The incidence of kala-azar caused by Leishmania infantum has recently increased in Nourabad-Mamassani district of Fars Province, in the south of the country. This study was designed to determine the role of asymptomatic dogs as host reservoir of L. infantum in this new formed focus and detection of prevalence of infection near them. A total of 20 as¬ymptomatic stray and sheep dogs were randomly sampled. The Buffy coat layer of their peripheral blood was used for DNA extraction and PCR. A species specific seminested PCR was used for DNA amplification using LINR4, LIN17 and LIN19 primers. These primers amplified variable area of the minicircle kDNA of Leishmania parasites. Of the 20 sampled dogs checked for leishmanial kDNA, six (30% were found naturally infected. It is concluded that, dogs (Canis familiaris even if asympto¬matic, is considered as the domestic host reservoir of kala-azar in this endemic focus.

  3. Serological and molecular survey of Leishmania parasites in apparently healthy dogs in the West Bank, Palestine

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    Hamarsheh Omar

    2012-08-01

    Full Text Available Abstract Background Canine visceral leishmaniasis (CVL is caused by Leishmania infantum in all Mediterranean countries. The Leishmania parasite is transmitted by the bite of a corresponding sand fly vector and primarily maintained in nature by wild and domestic reservoirs, including dogs, foxes and jackals. Infected dogs are the primary reservoir host in endemic regions and are the most significant risk disposing humans to infection. The present study aimed at assessing the prevalence of infection with Leishmania and identification of Leishmania infantum in domestic dogs in the West Bank, Palestine. Methods The infection rate among domestic dogs collected from seven districts in the Palestinian West Bank was investigated by examination of parasites in culture from the buffy coat using serological and molecular methods; based on ELISA, internal transcribed spacer 1 (ITS1 and cysteine protease (CPB PCR. Results Out of 215 dogs examined for Leishmania, 36 (16.7% were positive in at least one method. Twenty three animals (11.5% were positive for Leishmania DNA, whereas, ELISA and culture revealed 16 (7.5%, and 4 (1.5% respectively. CPB-PCR on one of three culture-positive isolates revealed Leishmania infantum as the causative agent for Leishmania infection in dogs. Conclusions Our study showed that canine leishmania infection is prevalent with varying degrees in all the seven studied districts in Palestine despite the absence of human VL cases in 4 of these districts. The causative agent was confirmed to be Leishmania infantum.

  4. Molecular diagnosis of canine visceral leishmaniasis: a comparative study of three methods using skin and spleen from dogs with natural Leishmania infantum infection.

    Science.gov (United States)

    Reis, Levi Eduardo Soares; Coura-Vital, Wendel; Roatt, Bruno Mendes; Bouillet, Leoneide Érica Maduro; Ker, Henrique Gama; Fortes de Brito, Rory Cristiane; Resende, Daniela de Melo; Carneiro, Mariângela; Giunchetti, Rodolfo Cordeiro; Marques, Marcos José; Carneiro, Cláudia Martins; Reis, Alexandre Barbosa

    2013-11-08

    Polymerase chain reaction (PCR) and its variations represent highly sensitive and specific methods for Leishmania DNA detection and subsequent canine visceral leishmaniasis (CVL) diagnosis. The aim of this work was to compare three different molecular diagnosis techniques (conventional PCR [cPCR], seminested PCR [snPCR], and quantitative PCR [qPCR]) in samples of skin and spleen from 60 seropositive dogs by immunofluorescence antibody test and enzyme-linked immunosorbent assay. Parasitological analysis was conducted by culture of bone marrow aspirate and optical microscopic assessment of ear skin and spleen samples stained with Giemsa, the standard tests for CVL diagnosis. The primers L150/L152 and LINR4/LIN17/LIN19 were used to amplify the conserved region of the Leishmania kDNA minicircle in the cPCR, and snPCR and qPCR were performed using the DNA polymerase gene (DNA pol α) primers from Leishmania infantum. The parasitological analysis revealed parasites in 61.7% of the samples. Sensitivities were 89.2%, 86.5%, and 97.3% in the skin and 81.1%, 94.6%, and 100.0% in spleen samples used for cPCR, snPCR, and qPCR, respectively. We demonstrated that the qPCR method was the best technique to detect L. infantum in both skin and spleen samples. However, we recommend the use of skin due to the high sensitivity and sampling being less invasive. Copyright © 2013 Elsevier B.V. All rights reserved.

  5. Homeopathic medicines cause Th1 predominance and induce spleen and megakaryocytes changes in BALB/c mice infected with Leishmania infantum.

    Science.gov (United States)

    Cajueiro, Ana Paula Bacellar; Goma, Ester Puna; Dos Santos, Hilton Antônio Mata; Almeida Rodrigues, Igor; Toma, Helena Keiko; Araújo, Silvana Marques; Bonamin, Leoni Villano; Gomes, Nelson Brêtas de Noronha; Castelo-Branco, Morgana Teixeira Lima; de Souza Dias, Edilma Paraguai; Dos Santos Pyrrho, Alexandre; Holandino, Carla

    2017-07-01

    The prevalence of Th1/Th2 response, spleen changes and megakaryocytes were investigated in BALB/c mice (n=138) infected with Leishmania infantum, and treated with Leishmania infantum 30× (10 -30 ) biotherapy - BioLi30×. We performed controlled experiments using 8-to-12-week-old mice, infected with 5×10 7 L. infantum promastigotes, divided into eight groups: G1 (healthy), G2 (infected with L. infantum), G3 (BioLi30× pre-treated), G4 (BioLi30× pre/post-treated), G5 (BioLi30× post-treated), G6 (Water 30× post-treated), G7 (Antimonium crudum 30× post-treated) and G8 (Glucantime® post-treated). G3-G7 groups were orally treated with their respective drugs diluted in filtered water (1:10), and G8 received Glucantime® (0.6mg/100µl of PBS), intraperitoneally. Spleen fragments were submitted to double blind histopathological evaluation and the number of megakaryocytes was counted. Besides, animals' serum was measured after 49days of infection, and cytokines (IFN-γ, IL-4, IL-10, IL-12), as well as the Th1/Th2 correlation (IFN-γ/IL-4 and IFN-γ/IL-10), were analyzed. Spleen histological parameters were classified as: healthy appearance (G1); discreet (G3-G7), moderate (G2) and moderate to severe (G8) white pulp hyperplasia; proliferation of megakaryocytes (G2-G8), and intense disruption (G2-G8). All groups, except for G7, showed higher percentages of megakaryocytes per field ranging from 87% to 15%, when compared to healthy animals (G1). Th1 predominance in IFN-γ/IL-4 ratio (comparing to G2) was detected in G4, G5, G6 and G7. Finally, pre/post (BioLi30x) and post-treatment (Antimonium crudum 30x) presented reduction of megakaryocytes/spleen changes due to immunomodulation animal process, controlling the infection process, probably by the Th1 cytokine predominance. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. A minimally invasive approach to spleen histopathology in dogs: A new method for follow-up studies of spleen changes in the course of Leishmania infantum infection.

    Science.gov (United States)

    Santos, Silvana Ornelas; Fontes, Jonathan L M; Laranjeira, Daniela F; Vassallo, José; Barrouin-Melo, Stella Maria; Dos-Santos, Washington L C

    2016-10-01

    Severe forms of zoonotic visceral leishmaniosis (ZVL) are associated with disruption of the spleen structure. However, the study of spleen histology requires splenectomy or necropsy. In this work, we present a minimally invasive cell-block technique for studying spleen tissue histology in dogs with ZVL. We examined 13 dogs with and seven dogs without Leishmania infantum infection. The dogs with Leishmania infection had a lower frequency of lymphoid follicles (2/13, Fisher's test, Pdogs (5/7 exhibiting lymphoid follicles and a plasma cell score of 1). The dogs with Leishmania infection also presented with granulomas (8/13) and infected macrophages (5/13). These differences in the histological presentations of spleen tissue from infected and uninfected dogs corresponded to changes observed in conventional histology. Hence, the cell-block technique described here may be used in the follow-up care and study of dogs with ZVL and other diseases in both clinical practice and research. Copyright © 2016 Elsevier Ltd. All rights reserved.

  7. [Natural infection with Leishmania infantum chagasi in Lutzomyia (Lutzomyia) longipalpis (Diptera: Psychodidae) sandflies captured in the municipality of Janaúba, State of Minas Gerais, Brazil].

    Science.gov (United States)

    Michalsky, Erika Monteiro; Guedes, Karla de Sena; Lara e Silva, Fabiana de Oliveira; França-Silva, João Carlos; Dias, Consuelo Latorre Fortes; Barata, Ricardo Andrade; Dias, Edelberto Santos

    2011-01-01

    Visceral leishmaniasis has been notified in nearly all states of Brazil, and particularly in the north of Minas Gerais, where the disease is endemic. The aim of this study was to detect natural infection of Lutzomyia longipalpis and, through the PCR/RFLP technique, identify Leishmania species found in sandflies in the municipality of Janaúba. Using light traps, 1,550 females of L. longipalpis were caught and grouped into pools of 10 specimens to be subjected to DNA extraction and amplification, by means of generic PCR and cacophony. Out of the 155 pools, six were positive for Leishmania sp., and thus the infection rate in the municipality was 3.9%. Through PCR/RFLP, the digestion pattern among the positive samples was found to be similar to that of the reference strain of Leishmania chagasi (MHOM/BR/74/PP75). The detection of natural infection associated with studies on the epidemiology of visceral leishmaniasis suggests that L. longipalpis is involved in transmission of L. infantum chagasi in Janaúba, particularly in areas of intense transmission of visceral leishmaniasis.

  8. Leishmania major glycosylation mutants require phosphoglycans (lpg2- but not lipophosphoglycan (lpg1- for survival in permissive sand fly vectors.

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    Anna Svárovská

    2010-01-01

    Full Text Available Sand fly species able to support the survival of the protozoan parasite Leishmania have been classified as permissive or specific, based upon their ability to support a wide or limited range of strains and/or species. Studies of a limited number of fly/parasite species combinations have implicated parasite surface molecules in this process and here we provide further evidence in support of this proposal. We investigated the role of lipophosphoglycan (LPG and other phosphoglycans (PGs in sand fly survival, using Leishmania major mutants deficient in LPG (lpg1(-, and the phosphoglycan (PG-deficient mutant lpg2(-. The sand fly species used were the permissive species Phlebotomus perniciosus and P. argentipes, and the specific vector P. duboscqi, a species resistant to L. infantum development.The lpg2(- mutants did not survive well in any of the three sand fly species, suggesting that phosphoglycans and/or other LPG2-dependent molecules are required for parasite development. In vitro, all three L. major lines were equally resistant to proteolytic activity of bovine trypsin, suggesting that sand fly-specific hydrolytic proteases or other factors are the reason for the early lpg2(- parasite killing. The lpg1(- mutants developed late-stage infections in two permissive species, P. perniciosus and P. argentipes, where their infection rates and intensities of infections were comparable to the wild type (WT parasites. In contrast, in P. duboscqi the lpg1(- mutants developed significantly worse than the WT parasites.In combination with previous studies, the data establish clearly that LPG is not required for Leishmania survival in permissive species P. perniciosus and P. argentipes but plays an important role in the specific vector P. duboscqi. With regard to PGs other than LPG, the data prove the importance of LPG2-related molecules for survival of L. major in the three sand fly species tested.

  9. Evaluation of a novel dried blood spot collection device (HemaSpot™) to test blood samples collected from dogs for antibodies to Leishmania infantum.

    Science.gov (United States)

    Rosypal, Alexa C; Pick, Leanne D; Hernandez, Jaime O Esquivel; Lindsay, David S

    2014-09-15

    Collection of blood samples from veterinary and wildlife patients is often challenging because the samples have to be collected on farm or in the wild under various environmental conditions. This poses many technical problems associated with venipuncture materials, their safe use and disposal, transportation and processing of collected samples. Dried blood spot (DBS) sample collection techniques offer a simple and practical alternative to traditional blood collection methods to obtain blood samples from animals for parasite antibody evaluation. The DBS collection devices are compact, simple to use, and are particularly useful for large number of samples. Additionally, DBS samples take up less space and they are easier to transport than traditional venipuncture-collected blood samples. Visceral leishmaniasis (VL) is a potentially fatal parasitic disease of dogs and humans and it is frequently diagnosed by antibody tests. Immunochromatographic tests (ICT) for antibodies to Leishmania infantum are commercially available for dogs and they produce qualitative results in minutes. Measurement of canine antibodies to L. infantum with the ICT using traditional venipuncture has been validated previously, but the use of DBS samples has not been evaluated using this method. The purpose of the present study was to determine the ability of DBS samples to detect antibodies to L. infantum in dogs using a commercial ICT assay. One hundred plasma samples from dogs experimentally infected with the LIVT-1 strain of L. infantum were collected by venipuncture and frozen. Individual samples were thawed, and then 80 μl plasma (2 drops) was aliquotted onto the 8-spoked disk pad on individual DBS sample collection devices (HemaSpot™, Spot-On Sciences, Austin, TX), dried, and stored in the dark at room temperature. After one month and six months, respectively, 2 spokes of the 8 spokes of the disk pad of each DBS sample were removed and eluted in 200 μl PBS. The eluate was used to test

  10. Leishmania attachment in permissive vectors and the role of sand fly midgut proteins in parasite-vector interaction

    OpenAIRE

    Dostálová, Anna

    2012-01-01

    of PhD. thesis named "Leishmania attachment in permissive vectors and the role of sand fly midgut proteins in parasite-vector interaction", Anna Dostálová, 2011 This thesis focuses on the development of protozoan parasites of the genus Leishmania in their insect vectors, sand flies. It sums up results of three projects I was involved in during my PhD studies. Main emphasis was put on permissive sand fly species that support development of various species of Leishmania. Using a novel method of...

  11. Use of an Interferon Gamma Release Assay (IGRA) to test T-cell responsiveness to soluble Leishmania infantum antigen in whole blood of dogs from endemic areas.

    Science.gov (United States)

    Zribi, Lilia; El-Goulli, Amel F; Ben-Abid, Meriem; Gharbi, Mohamed; Ben-Sghaier, Ines; Boufaden, Imed; Aoun, Karim; Bouratbine, Aïda

    2017-11-15

    Interferon-Gamma (IFN-γ) Release Assays (IGRAs) are easy tests that allow rapid screening of primed memory T-cells immunity in response to antigen. The aim of this study was to use IGRA to assess IFN-γ release in response to Soluble Leishmania infantum antigen (SLA) in whole blood of dogs living in endemic area of visceral leishmaniasis and to interpret IGRA results according to clinical examination, specific anti-Leishmania humoral response and presence of L. infantum DNA in blood. The study was carried out on 56 dogs living in greater Tunis area. Physical examination, quantitative serology and PCR on blood were used to characterize dogs' status in relation to Leishmania infection and disease. IGRA consisted on testing by ELISA for IFN-γ-secretion in whole blood after a 20-h challenge with SLA. PBS and Phytohemagglutinin (PHA) stimulations were used as controls. Four groups of dogs were characterized: 31 were negative by both serology and PCR, two had doubtful serology, 10 presented no to mild clinical signs but low antibodies levels and 13 were affected by Canine Leishmaniasis (CanL). In seronegative dogs, IGRA was little contributory in 4 puppies (age dogs (median age=72months, IQR: 45-84 months) that didn't respond to PHA stimulation, IGRA was negative in 19 and positive in three animals with lymph node enlargement. In dogs with doubtful serology, IGRA was positive in one dog and negative in the other. In infected dogs with no to mild clinical signs, one dog exhibited high level of IFN-γ in absence of antigenic stimulation and all the other were positive by IGRA. CanL dogs showed variable IGRA results. Negative IGRAs (n=4) were shown in animals with the highest parasitic burden whereas positive IGRAs (n=5) were shown in dogs with negative PCR or low parasitic load. The 4 remaining dogs either didn't respond to PHA (n=2) or showed non-specific secretion in PBS tube (n=2). The results of this study showed that IGRA is a useful new tool that can assess

  12. Improved proteolytic stability and potent activity against Leishmania infantum trypanothione reductase of α/β-peptide foldamers conjugated to cell-penetrating peptides.

    Science.gov (United States)

    de Lucio, Héctor; Gamo, Ana María; Ruiz-Santaquiteria, Marta; de Castro, Sonia; Sánchez-Murcia, Pedro A; Toro, Miguel A; Gutiérrez, Kilian Jesús; Gago, Federico; Jiménez-Ruiz, Antonio; Camarasa, María-José; Velázquez, Sonsoles

    2017-11-10

    The objective of the current study was to enhance the proteolytic stability of peptide-based inhibitors that target critical protein-protein interactions at the dimerization interface of Leishmania infantum trypanothione reductase (Li-TryR) using a backbone modification strategy. To achieve this goal we carried out the synthesis, proteolytic stability studies and biological evaluation of a small library of α/β 3 -peptide foldamers of different length (from 9-mers to 13-mers) and different α→β substitution patterns related to prototype linear α-peptides. We show that several 13-residue α/β 3 -peptide foldamers retain inhibitory potency against the enzyme (in both activity and dimerization assays) while they are far less susceptible to proteolytic degradation than an analogous α-peptide. The strong dependence of the binding affinities for Li-TryR on the length of the α,β-peptides is supported by theoretical calculations on conformational ensembles of the resulting complexes. The conjugation of the most proteolytically stable α/β-peptide with oligoarginines results in a molecule with potent activity against L. infantum promastigotes and amastigotes. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  13. Genes that encodes NAGT, MIF1 and MIF2 are not virulence factors for kala-azar caused by Leishmania infantum

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    Bruno Guedes Alcoforado Aguiar

    2014-10-01

    Full Text Available Introduction Kala-azar is a disease resulting from infection by Leishmania donovani and Leishmania infantum. Most patients with the disease exhibit prolonged fever, wasting, anemia and hepatosplenomegaly without complications. However, some patients develop severe disease with hemorrhagic manifestations, bacterial infections, jaundice, and edema dyspnea, among other symptoms, followed by death. Among the parasite molecules that might influence the disease severity are the macrophage migration inhibitory factor-like proteins (MIF1 and MIF2 and N-acetylglucosamine-1-phosphotransferase (NAGT, which act in the first step of protein N-glycosylation. This study aimed to determine whether MIF1, MIF2 and NAGT are virulence factors for severe kala-azar. Methods To determine the parasite genotype in kala-azar patients from Northeastern Brazil, we sequenced the NAGT genes of L. infantum from 68 patients as well as the MIF1 and MIF2 genes from 76 different subjects with diverse clinical manifestations. After polymerase chain reaction (PCR, the fragments were sequenced, followed by polymorphism identification. Results The nucleotide sequencing of the 144 amplicons revealed the absence of genetic variability of the NAGT, MIF1 and MIF2 genes between the isolates. The conservation of these genes suggests that the clinical variability of kala-azar does not depend upon these genes. Additionally, this conservation suggests that these genes may be critical for parasite survival. Conclusions NAGT, MIF1 and MIF2 do not alter the severity of kala-azar. NAGT, MIF1 and MIF2 are highly conserved among different isolates of identical species and exhibit potential for use in phylogenetic inferences or molecular diagnosis.

  14. Serum Removal from Culture Induces Growth Arrest, Ploidy Alteration, Decrease in Infectivity and Differential Expression of Crucial Genes in Leishmania infantum Promastigotes.

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    Pedro J Alcolea

    Full Text Available Leishmania infantum is one of the species responsible for visceral leishmaniasis. This species is distributed basically in the Mediterranean basin. A recent outbreak in humans has been reported in Spain. Axenic cultures are performed for most procedures with Leishmania spp. promastigotes. This model is stable and reproducible and mimics the conditions of the gut of the sand fly host, which is the natural environment of promastigote development. Culture media are undefined because they contain mammalian serum, which is a rich source of complex lipids and proteins. Serum deprivation slows down the growth kinetics and therefore, yield in biomass. In fact, we have confirmed that the growth rate decreases, as well as infectivity. Ploidy is also affected. Regarding the transcriptome, a high-throughput approach has revealed a low differential expression rate but important differentially regulated genes. The most remarkable profiles are: up-regulation of the GINS Psf3, the fatty acyl-CoA synthase (FAS1, the glyoxylase I (GLO1, the hydrophilic surface protein B (HASPB, the methylmalonyl-CoA epimerase (MMCE and an amastin gene; and down-regulation of the gPEPCK and the arginase. Implications for metabolic adaptations, differentiation and infectivity are discussed herein.

  15. Serum Removal from Culture Induces Growth Arrest, Ploidy Alteration, Decrease in Infectivity and Differential Expression of Crucial Genes in Leishmania infantum Promastigotes.

    Science.gov (United States)

    Alcolea, Pedro J; Alonso, Ana; Moreno-Izquierdo, Miguel A; Degayón, María A; Moreno, Inmaculada; Larraga, Vicente

    2016-01-01

    Leishmania infantum is one of the species responsible for visceral leishmaniasis. This species is distributed basically in the Mediterranean basin. A recent outbreak in humans has been reported in Spain. Axenic cultures are performed for most procedures with Leishmania spp. promastigotes. This model is stable and reproducible and mimics the conditions of the gut of the sand fly host, which is the natural environment of promastigote development. Culture media are undefined because they contain mammalian serum, which is a rich source of complex lipids and proteins. Serum deprivation slows down the growth kinetics and therefore, yield in biomass. In fact, we have confirmed that the growth rate decreases, as well as infectivity. Ploidy is also affected. Regarding the transcriptome, a high-throughput approach has revealed a low differential expression rate but important differentially regulated genes. The most remarkable profiles are: up-regulation of the GINS Psf3, the fatty acyl-CoA synthase (FAS1), the glyoxylase I (GLO1), the hydrophilic surface protein B (HASPB), the methylmalonyl-CoA epimerase (MMCE) and an amastin gene; and down-regulation of the gPEPCK and the arginase. Implications for metabolic adaptations, differentiation and infectivity are discussed herein.

  16. The inflammatory cytokine effect of Pam3CSK4 TLR2 agonist alone or in combination with Leishmania infantum antigen on ex-vivo whole blood from sick and resistant dogs

    OpenAIRE

    Mart?nez-Orellana, Pamela; Quirola-Amores, Paulina; Montserrat-Sangr?, Sara; Ordeix, Laura; Llull, Joan; ?lvarez-Fern?ndez, Alejandra; Solano-Gallego, Laia

    2017-01-01

    Background A wide spectrum of clinical manifestations and immune responses exist in canine L. infantum infection. Ibizan hounds are more ?resistant? to disease than other dog breeds. Recognition of pathogen-associated molecule patterns by toll like receptors (TLRs) rapidly triggers a variety of anti-microbial immune responses through the induction of pro-inflammatory cytokines such as TNF-? and IL-6 which may play an important role in controlling Leishmania infection. The main objective of th...

  17. Les phlébotomes (Diptera-Psychodidae de l'île de Chypre.II - Presence de Leishmania (Leishmania infantum Nicolle, 1908 (zymodeme MON 1 chez Phlebotomus (Larroussius tobbi Adler et Theodor, 1930

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    Léger N.

    2000-06-01

    Full Text Available Dans le foyer leishmanien cypriote, les auteurs ont disséqué 2910 femelles de phlébotomes appartenant à 11 espèces : Phlebotomus papatasi, P. sergenti, P. jacusieli, P. alexandri, P. tobbi, P. galilaeus, P. mascittii, P. economidesi, Sergentomyia fallax, S. minuta et S. azizi . Les deux larroussius (P. galilaeus et P. tobbi sont les espèces les plus abondantes. Elles représentent plus de 60 % des captures réalisées avec des pièges CDC. Des promastigotes ont été observées chez un seul spécimen appartenant à l'espèce P. tobbi. Elles ont été mises en culture puis identifiées selon la méthode isoenzymatique. La souche de Leishmania isolée appartient à l'espèce Leishmania infantum, zymodème MON 1. Le même zymodème a été aussi isolé et identifié chez quatre chiens de l'île. En l'absence des vecteurs habituels de L. infantum dans l'est méditerranéen (P. neglectus ef P. syriacus et en raison de sa répartition à Chypre, P. tobbi constitue vraisemblablement un bon vecteur local. Sa faible anthropophilie expliquerait peut-être le très faible nombre de cas humains. Le rôle de P. galilaeus dans la transmission de la leishmaniose à Chypre reste à préciser.

  18. Humoral and In Vivo Cellular Immunity against the Raw Insect-Derived Recombinant Leishmania infantum Antigens KMPII, TRYP, LACK, and papLe22 in Dogs from an Endemic Area

    Science.gov (United States)

    Todolí, Felicitat; Solano-Gallego, Laia; de Juan, Rafael; Morell, Pere; del Carmen Núñez, Maria; Lasa, Rodrigo; Gómez-Sebastián, Silvia; Escribano, José M.; Alberola, Jordi; Rodríguez-Cortés, Alhelí

    2010-01-01

    Leishmania infantum causes visceral leishmaniasis, a severe zoonotic and systemic disease that is fatal if left untreated. Identification of the antigens involved in Leishmania-specific protective immune response is a research priority for the development of effective control measures. For this purpose, we evaluated, in 27 dogs from an enzootic zone, specific humoral and cellular immune response by delayed-type hypersensitivity (DTH) skin test both against total L. infantum antigen and the raw Trichoplusia ni insect-derived kinetoplastid membrane protein-11 (rKMPII), tryparedoxin peroxidase (rTRYP), Leishmania homologue of receptors for activated C kinase (rLACK), and 22-kDa potentially aggravating protein of Leishmania (rpapLe22) antigens from this parasite. rTRYP induced the highest number of positive DTH responses (55% of leishmanin skin test [LST]-positive dogs), showing that TRYP antigen is an important T cell immunogen, and it could be a promising vaccine candidate against this disease. When TRYP-DTH and KMPII-DTH tests were evaluated in parallel, 82% of LST-positive dogs were detected, suggesting that both antigens could be considered as components of a standardized DTH immunodiagnostic tool for dogs. PMID:21118936

  19. Canine Skin and Conjunctival Swab Samples for the Detection and Quantification of Leishmania infantum DNA in an Endemic Urban Area in Brazil

    Science.gov (United States)

    de Almeida Ferreira, Sidney; Leite, Rodrigo Souza; Ituassu, Leonardo Trindade; Almeida, Gregório Guilherme; Souza, Daniel Menezes; Fujiwara, Ricardo Toshio; de Andrade, Antero Silva Ribeiro; Melo, Maria Norma

    2012-01-01

    Background We evaluated kDNA PCR/hybridization and quantitative real-time PCR (qPCR) targeting the gene of DNA polymerase of Leishmania infantum for CVL diagnosis and assessment of parasite load in clinical samples obtained invasively and non-invasively. Methodology/Principal Findings Eighty naturally infected dogs from an endemic urban area in Brazil were used. Animals were divided into two groups based on the presence or absence of CVL clinical sings. Skin biopsies, bone marrow, blood and conjunctival swabs samples were collected and submitted to L. infantum DNA detection. In addition, anti-Leishmania antibody titers were measured by Immunofluorescence antibody test. The symptomatic dogs had increased titers compared to asymptomatic dogs (P = 0.025). The frequencies of positive results obtained by kDNA PCR/hybridization for asymptomatic and symptomatic dogs, respectively, were as follows: right conjunctiva, 77.5% and 95.0%; left conjunctiva, 75.0% and 87.5%; skin, 45.0% and 75.0%; bone marrow, 50.0% and 77.5%; and blood, 27.5% and 22.5%. In both groups, the parasite load in the skin samples was the highest (P<0.0001). The parasite loads in the conjunctival swab and bone marrow samples were statistically equivalent within each group. The parasite burden in conjunctival swabs was higher in the dogs with clinical signs than in asymptomatic dogs (P = 0.028). This same relationship was also observed in the bone marrow samples (P = 0.002). No differences in amastigotes load in the skin were detected between the groups. Conclusions The conjunctival swab is a suitable clinical sample for qualitative molecular diagnosis of CVL. The highest parasite burdens were detected in skin regardless of the presence of VL-associated clinical signs. The qPCR results emphasized the role of dogs, particularly asymptomatic dogs, as reservoirs for CVL because of the high cutaneous parasite loads. These results may help to explain the maintenance of high transmission rates and

  20. Alterações eletromiográficas e histopatológicas da musculatura de cães naturalmente infectados por Leishmania infantum

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    Gisela Cristiane Ferraro

    2012-10-01

    Full Text Available O objetivo do presente estudo foi avaliar as alterações eletromiográficas e histopatológicas de músculos estriados esqueléticos de cães naturalmente infectados por Leishmania infantum. Foram selecionados 25 cães adultos, sem raça definida, com diagnósticos parasitológico, molecular e sorológico estabelecidos para a infecção. Os músculos avaliados foram: tríceps braquial, extensorcarpo radial, bíceps femoral e gastrocnêmio. Um cão possuía problemas locomotores, com paresia de membros posteriores associada à intensa atrofia muscular. Vinte e três (92% apresentavam algum tipo de alteração muscular, sendo que em 22(88% tais alterações foram identificadas diretamente pela eletromiografia. Mesmo sem sinais clínicos, em dez cães (40% foram evidenciadas alterações eletromiográficas e histopatológicas. Antígenos de Leishmania foram identificados na musculatura de quatro (16% cães. Os resultados eletromiográficos indicaram a ocorrência de polimiosite crônica em 13 (52% cães, presença de músculos com inflamação tanto aguda quanto crônica em quatro (16%, miopatia aguda em dois (8%, e ausência de alterações eletromiográficas em três (12%. As alterações histopatológicas mais frequentemente observadas foram degeneração e necrose de miofibras e presença de infiltrado inflamatório verificadas em 12 (48% cães. Outras alterações, quando comparado com as amostras de cães normais, foram do tamanho de grupos de fibras musculares em 15 (60% e fibrose peri ou endomisial em 14(56% animais. As alterações observadas no presente estudo permitiram concluir que mesmo na ausência de sinais clínicos de comprometimento muscular, a maior parte dos cães infectados por L. infantum apresenta polimiosite crônica.

  1. Seroprevalence rates of antibodies against Leishmania infantum and other protozoan and rickettsial parasites in dogs Soroprevalência de anticorpos contra Leishmania infantum e outras espécies de protozoários e rickettsia em cães

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    Silvana de Cássia Paulan

    2013-03-01

    Full Text Available Canine visceral leishmaniasis (CVL is caused by the protozoan Leishmania infantum, which infects dogs and humans in many regions of Brazil. The present study involved an indirect fluorescent antibody test (IFAT to analyze L. infantum, Ehrlichia spp., Babesia canis, Toxoplasma gondii and Neospora caninum infection rates in serum samples from 93 dogs in a rural settlement in Ilha Solteira, SP, Brazil. The seroprevalence rates of anti-L. infantum, anti-Ehrlichia, anti-B. canis, anti-T. gondii and anti-N. caninum antibodies were 37.6%, 75.3%, 72%, 47.3% and 6.4%, respectively. In addition to IFAT, direct microscopic examination of popliteal lymph node aspirates revealed 26.9% of CVL positive dogs. Serological tests revealed that 17.2% of the dogs were seropositive for a single parasite, 29% for two parasites, 33% for three, 16.1% for four, and 1.1% for five parasites, while 3.2% were seronegative for five parasites. The presence of antibodies against these parasites in serum samples from dogs confirmed their exposure to these parasites in this rural area. Because of the potential zoonotic risk of these diseases, mainly leishmaniasis, ehrlichiosis and toxoplasmosis, special attention should focus on programs for the improvement of diagnostic assays and control measures against these parasites.Leishmaniose Visceral Canina (LVC é causada pelo protozoário Leishmania infantum, podendo infectar cães e humanos em várias regiões do Brasil. O presente estudo teve por objetivo realizar a reação de imunofluorescência indireta (RIFI para analisar os índices de infecção parasitária para L. infantum, Ehrlichia spp., Babesia canis, Toxoplasma gondii e Neospora caninum, em 93 amostras de soro de cães de um assentamento rural no município de Ilha Solteira, SP, Brasil. A taxa de soroprevalência de cães com anticorpos anti-L. infantum, anti-Ehrlichia, anti-B. canis, anti-T. gondii e anti-N. caninum foi de 37,6%, 75,3%, 72%, 47,3% e 6

  2. Parasite load induces progressive spleen architecture breakage and impairs cytokine mRNA expression in Leishmania infantum-naturally infected dogs.

    Science.gov (United States)

    Cavalcanti, Amanda S; Ribeiro-Alves, Marcelo; Pereira, Luiza de O R; Mestre, Gustavo Leandro; Ferreira, Anna Beatriz Robottom; Morgado, Fernanda N; Boité, Mariana C; Cupolillo, Elisa; Moraes, Milton O; Porrozzi, Renato

    2015-01-01

    Canine Visceral Leishmaniasis (CVL) shares many aspects with the human disease and dogs are considered the main urban reservoir of L. infantum in zoonotic VL. Infected dogs develop progressive disease with a large clinical spectrum. A complex balance between the parasite and the genetic/immunological background of the host are decisive for infection evolution and clinical outcome. This study comprised 92 Leishmania infected mongrel dogs of various ages from Mato Grosso, Brazil. Spleen samples were collected for determining parasite load, humoral response, cytokine mRNA expression and histopathology alterations. By real-time PCR for the ssrRNA Leishmania gene, two groups were defined; a low (lowP, n = 46) and a high parasite load groups (highP, n = 42). When comparing these groups, results show variable individual humoral immune response with higher specific IgG production in infected animals but with a notable difference in CVL rapid test optical densities (DPP) between highP and lowP groups. Splenic architecture disruption was characterized by disorganization of white pulp, more evident in animals with high parasitism. All cytokine transcripts in spleen were less expressed in highP than lowP groups with a large heterogeneous variation in response. Individual correlation analysis between cytokine expression and parasite load revealed a negative correlation for both pro-inflammatory cytokines: IFNγ, IL-12, IL-6; and anti-inflammatory cytokines: IL-10 and TGFβ. TNF showed the best negative correlation (r2 = 0.231; pdogs with high parasite load associated with a structural modification in the splenic lymphoid micro-architecture. We also discuss the possible mechanism responsible for the uncontrolled parasite growth and clinical outcome.

  3. The midgut transcriptome of Lutzomyia longipalpis: comparative analysis of cDNA libraries from sugar-fed, blood-fed, post-digested and Leishmania infantum chagasi-infected sand flies

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    Elnaiem Dia-Eldin

    2008-01-01

    Full Text Available Abstract Background In the life cycle of Leishmania within the alimentary canal of sand flies the parasites have to survive the hostile environment of blood meal digestion, escape the blood bolus and attach to the midgut epithelium before differentiating into the infective metacyclic stages. The molecular interactions between the Leishmania parasites and the gut of the sand fly are poorly understood. In the present work we sequenced five cDNA libraries constructed from midgut tissue from the sand fly Lutzomyia longipalpis and analyzed the transcripts present following sugar feeding, blood feeding and after the blood meal has been processed and excreted, both in the presence and absence of Leishmania infantum chagasi. Results Comparative analysis of the transcripts from sugar-fed and blood-fed cDNA libraries resulted in the identification of transcripts differentially expressed during blood feeding. This included upregulated transcripts such as four distinct microvillar-like proteins (LuloMVP1, 2, 4 and 5, two peritrophin like proteins, a trypsin like protein (Lltryp1, two chymotrypsin like proteins (LuloChym1A and 2 and an unknown protein. Downregulated transcripts by blood feeding were a microvillar-like protein (LuloMVP3, a trypsin like protein (Lltryp2 and an astacin-like metalloprotease (LuloAstacin. Furthermore, a comparative analysis between blood-fed and Leishmania infected midgut cDNA libraries resulted in the identification of the transcripts that were differentially expressed due to the presence of Leishmania in the gut of the sand fly. This included down regulated transcripts such as four microvillar-like proteins (LuloMVP1,2, 4 and 5, a Chymotrypsin (LuloChym1A and a carboxypeptidase (LuloCpepA1, among others. Upregulated midgut transcripts in the presence of Leishmania were a peritrophin like protein (LuloPer1, a trypsin-like protein (Lltryp2 and an unknown protein. Conclusion This transcriptome analysis represents the largest set

  4. Activity of cycloartane-type triterpenes and sterols isolated from Musa paradisiaca fruit peel against Leishmania infantum chagasi.

    Science.gov (United States)

    Silva, A A S; Morais, S M; Falcão, M J C; Vieira, I G P; Ribeiro, L M; Viana, S M; Teixeira, M J; Barreto, F S; Carvalho, C A; Cardoso, R P A; Andrade-Junior, H F

    2014-09-25

    The aim of the study was to evaluate in vitro the antileishmanial activity of triterpenes and sterols isolated from Musa paradisiaca (banana) fruit peel used traditionally to treat leishmaniasis. The compounds were isolated from the ethanolic extract of the peel of the banana fruit by column chromatography. The chemical structure of compounds was determined by (1)H and (13)C - nuclear magnetic resonance spectroscopy. The cytotoxicity was measured in RAW 264.7 cells and LLC-MK2. Leishmanicidal activity against L. infantum chagasi promastigotes was performed by the MTT colorimetric method and activity against amastigotes was assayed in mammalian cells using in situ ELISA method. Five compounds were identified, consisting of three triterpenes: cycloeucalenone, 31-norcyclolaudenone and 24-methylene-cicloartanol and a mixture of two sterols: beta-sitosterol and stigmasterol. With the exception of cycloeucalenone, all compounds showed statistically similar activity against promastigote to pentamidine. While, acting against amastigotes, excluding 31-norcyclolaudenone, other compounds showed activity similar to amphotericin B. All compounds showed low cytotoxicity in mammalian cells. This study partially confirms the use of Musa paradisiaca in folk medicine against leishmaniasis. Further in vivo studies are necessary to evaluate the efficacy. Copyright © 2014 Elsevier GmbH. All rights reserved.

  5. Production of a conjugate between the rK346 antigen from Leishmania infantum and the horseradish peroxidase C for the detection of rK346 antibodies.

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    Rengifo-González, Juan; Medina-Mora, Yollyseth; Silva-Barrios, Sasha; Márquez-Contreras, María Elizabeth; Tibisay Ruiz, María; Cáceres, Ana J; Concepción, Juan Luis; Quiñones, Wilfredo

    2016-06-01

    It was designed and characterized a reporter system to be captured by an- tibodies bound to ELISA plates. The system was designed with the rK346 from Leishmania infantum, a highly antigenic and specific protein. The rK346 was coupled to the horseradish peroxidase C (HRPc) from Armoracia rusticana using glutaraldehyde or sulfo-SMCC. Gluta- raldehyde conjugation was performed in two steps. Separation of conjugates was carried out using a Sepharose S-200 in size exclusion chromatography (SEC); fractions were analyzed via HRPc activity and through ELISA plates sensitized with polyclonal anti-rK346 IgG puri- fied from rabbit serum. A heterogeneous population of conjugates rK346-HRPc was obtained with molecular weights ranging between 109.7 ± 16.5 to 67.6 ± 10.1 kDa; with rK346-HRPe stoichiometries of 1:2; 2:1; 3:1; and 2:2. Conjugation using sulfo-SMCC was carried out first by introducing -SH groups onto the HRPc using the SATA reagent and the antigen was modi- fied with sulfo-SMCC during 45 min. Separation and analysis of conjugates was performed similarly as with glutaraldehyde, resulting in a heterogeneous population of conjugates rK346- HRPc with molecular weights between 150.5 ± 22.6 to 80.0 ± 12.0 kDa; with rK346-HRPC stoichiometries of 2:1; 1:2; 2:2; and 1:3, with an increased conjugation efficiency in compari- son with glutaraldehyde. This enables sulfo-SMCC to be used as a potential reagent for cou- pling the antigen to the HRPc, to design an economic, specific and easy method to apply as a reporter system, available to assess individuals at risk and/or at early and late stages of visceral leishmaniasis.

  6. Leishmania Surveillance and Diagnostic Capability in Support of the Joint Biological Agent Identification and Diagnostic System (JBAIDS) and Leishmania Vector Surveillance

    Science.gov (United States)

    2013-02-07

    01-10-09 to 07-02-13 ’+. I II L~ J.\\NU :::OU~ Ill L~ :la. l-UI’I I 11J.\\l- I NUIVI~~I1 LEISHMANIA SURVEILLANCE AND DIAGNOSTIC CAPABILITY IN None...SUPPORT OF THE JOINT BIOLOGICAL AGENT IDENTIFICATION AND :lD. l:JI1J.\\NI NUIVI~~I1 DIAGNOSTIC SYSTEM (JBAIDS) None . ./ LEISHMANIA VECTOR...Field Station at Kisumu completed project activities through a resource sharing arrangement with the 59th MDW. Testing of the Leishmania epidemiology

  7. Sandflies (Diptera: Psychodidae associated with opossum nests at urban sites in southeastern Brazil: a risk factor for urban and periurban zoonotic Leishmania transmission?

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    Andre Antonio Cutolo

    2014-06-01

    Full Text Available Sandflies associated with opossum nests are reported for the first time in the yards of residences located in the urban area of the municipality of Monte Mor, situated in the metropolitan region of Campinas, state of São Paulo, Brazil. Eleven specimens of Evandromyia cortelezzii and one of Evandromyia lenti were captured in two Didelphis albiventris nests. Ev. cortelezzii is considered a secondary vector species for the transmission of Leishmania (Viannia braziliensis and Leishmania (Leishmania infantum in the Neotropics. This association may contribute to the introduction, establishment and maintenance of urban and periurban zoonotic transmission outbreaks of Leishmania and should therefore be investigated further.

  8. Field trial of efficacy of the Leish-tec® vaccine against canine leishmaniasis caused by Leishmania infantum in an endemic area with high transmission rates.

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    Gabriel Grimaldi

    Full Text Available Because domestic dogs are reservoir hosts for visceral leishmaniasis (VL in Brazil, one of the approaches used to reduce human disease incidence is to cull infected dogs. However, the results of controlled intervention trials based on serological screening of dogs and killing of seropositive animals are equivocal. A prophylactic vaccine to protect dogs from being infectious to the sand fly vector could be an effective strategy to provide sustained control. Here, we investigated whether a currently licensed commercial subunit rA2 protein-saponin vaccine (Leish-tec® had an additional effect to dog culling on reducing the canine infectious populations.This prospective study was conducted in an L. infantum highly endemic area of southeast Brazil. At the onset of the intervention, all of the eligible dogs received through subcutaneous route a three-dose vaccine course at 21-day intervals and a booster on month 12. For the purpose of comparison, newly recruited healthy dogs were included as the exposed control group. To ascertain vaccine-induced protection, dogs were screened on clinical and serological criteria every 6 months for a 2-year follow-up period. Antibody-based tests and histopathological examination of post-mortem tissue specimens from euthanized animals were used as a marker of infection. The standardized vaccine regime, apart from being safe, was immunogenic as immunized animals responded with a pronounced production of anti-A2-specific IgG antibodies. It should be noted the mean seroconversion time for infection obtained among immunized exposed dogs (~ 18 months, which was twice as high as that for unvaccinated ones (~ 9 months. After two transmission cycles completed, the cumulative incidence of infection did differ significantly (P = 0.016 between the vaccinated (27% and unvaccinated (42% dogs. However, the expected efficacy for the vaccine in inducing clinical protection was not evident since 43% of vaccine recipients developed

  9. Molecular screening of Leishmania spp. infection and bloodmeals in sandflies from a leishmaniasis focus in southwestern Turkey.

    Science.gov (United States)

    Karaku Ş, M; Pekağ Irba Ş, M; Demir, S; Eren, H; Töz, S; Özbel, Y

    2017-06-01

    Leishmaniasis is an arthropod-borne disease that affects approximately 2 million people worldwide annually. The aims of this study were to detect the presence of Leishmania (Kinetoplastida: Trypanosomatidae) DNA and the feeding preferences of probable vector species in an endemic focus of Leishmania infantum in Turkey. Entomological sampling was performed in August and October 2015 in Aydın province, where cases of human and canine leishmaniasis have been reported previously. A total of 1059 sandfly specimens comprising nine species belonging to two genera, Phlebotomus and Sergentomyia (both: Diptera: Psychodidae), and five subgenera of the Phlebotomus genus (Phlebotomus, Paraphlebotomus, Larroussius, Adlerius and Transphlebotomus) were collected in five villages. Among all Phlebotomus specimens, Phlebotomus neglectus (39%) was noted as the most abundant species, followed by Phlebotomus tobbi (18%). Leishmania DNA was detected in pools from P. neglectus, P. tobbi and Sergentomyia dentata by kDNA polymerase chain reaction (PCR). Leishmania DNA from Phlebotomus specimens was identified as L. infantum, but Leishmania DNA from Sergentomyia spp. could not be identified to species level by ITS-1 real-time PCR. The detection of Leishmania DNA in wild-caught P. neglectus and the high percentage (24.2%) of human DNA in engorged specimens suggests that P. neglectus is probably an important vector species for L. infantum in Aydın province. © 2016 The Royal Entomological Society.

  10. Insect vectors of Leishmania: distribution, physiology and their control.

    Science.gov (United States)

    Sharma, Umakant; Singh, Sarman

    2008-12-01

    Leishmaniasis is a deadly vector-borne disease that causes significant morbidity and mortality in Africa, Asia, Latin America and Mediterranean regions. The causative agent of leishmaniasis is transmitted from man to man by a tiny insect called sandfly. Approximately, 600 species of sandflies are known but only 10% of these act as disease vectors. Further, only 30 species of these are important from public health point. Fauna of Indian sub-zone is represented by 46 species, of these, 11 belong to Phlebotomine species and 35 to Sergentomyia species. Phlebotomus argentipes is the proven vector of kala-azar or visceral leishmaniasis in India. This review gives an insight into the insect vectors of human leishmaniasis, their geographical distribution, recent taxonomic classification, habitat, and different control measures including indoor residual spraying (IRS), insecticide-treated bednets (ITNs), environmental management, biological control, and emerging resistance to DDT. Role of satellite remote sensing for early prediction of the disease by identifying the sandflygenic conditions cannot be undermined. The article also underlines the importance of synthetic pheromones which can be used in near future for the control of these vectors.

  11. Integrating ribosomal promoter vectors that offer a choice of constitutive expression profiles in Leishmania donovani.

    Science.gov (United States)

    Soysa, Radika; Tran, Khoa D; Ullman, Buddy; Yates, Phillip A

    2015-12-01

    We have designed a novel series of integrating ribosomal RNA promoter vectors with five incrementally different constitutive expression profiles, covering a 250-fold range. Differential expression was achieved by placing different combinations of synthetic or leishmanial DNA sequences upstream and downstream of the transgene coding sequence in order to modulate pre-mRNA processing efficiency and mRNA stability, respectively. All of the vectors have extensive multiple cloning sites, and versions are available for producing N- or C- terminal GFP fusions at each of the possible relative expression levels. In addition, the modular configuration of the vectors allows drug resistance cassettes and other components to be readily exchanged. In toto, these vectors should be useful additions to the toolkit available for molecular and genetic studies of Leishmania donovani. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Demonstration of genetic exchange during cyclical development of Leishmania in the sand fly vector.

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    Akopyants, Natalia S; Kimblin, Nicola; Secundino, Nagila; Patrick, Rachel; Peters, Nathan; Lawyer, Phillip; Dobson, Deborah E; Beverley, Stephen M; Sacks, David L

    2009-04-10

    Genetic exchange has not been shown to be a mechanism underlying the extensive diversity of Leishmania parasites. We report here evidence that the invertebrate stages of Leishmania are capable of having a sexual cycle consistent with a meiotic process like that described for African trypanosomes. Hybrid progeny were generated that bore full genomic complements from both parents, but kinetoplast DNA maxicircles from one parent. Mating occurred only in the sand fly vector, and hybrids were transmitted to the mammalian host by sand fly bite. Genetic exchange likely contributes to phenotypic diversity in natural populations, and analysis of hybrid progeny will be useful for positional cloning of the genes controlling traits such as virulence, tissue tropism, and drug resistance.

  13. Visualisation of Leishmania donovani fluorescent hybrids during early stage development in the sand fly vector.

    Science.gov (United States)

    Sadlova, Jovana; Yeo, Matthew; Seblova, Veronika; Lewis, Michael D; Mauricio, Isabel; Volf, Petr; Miles, Michael A

    2011-01-01

    The Leishmania protozoan parasites cause devastating human diseases. Leishmania have been considered to replicate clonally, without genetic exchange. However, an accumulation of evidence indicates that there are inter-specific and intra-specific hybrids among natural populations. The first and so far only experimental proof of genetic exchange was obtained in 2009 when double drug resistant Leishmania major hybrids were produced by co-infecting sand flies with two strains carrying different drug resistance markers. However, the location and timing of hybridisation events in sand flies has not been described. Here we have co-infected Phlebotomus perniciosus and Lutzomyia longipalpis with transgenic promastigotes of Leishmania donovani strains carrying hygromycin or neomycin resistance genes and red or green fluorescent markers. Fed females were dissected at different times post bloodmeal (PBM) and examined by fluorescent microscopy or fluorescent activated cell sorting (FACS) followed by confocal microscopy. In mixed infections strains LEM3804 and Gebre-1 reached the cardia and stomodeal valves more rapidly than strains LEM4265 and LV9. Hybrids unequivocally expressing both red and green fluorescence were seen in single flies of both vectors tested, co-infected with LEM4265 and Gebre-1. The hybrids were present as short (procyclic) promastigotes 2 days PBM in the semi-digested blood in the endoperitrophic space. Recovery of a clearly co-expressing hybrid was also achieved by FACS. However, hybrids could not sustain growth in vitro. For the first time, we observed L. donovani hybrids in the sand fly vector, 2 days PBM and described the morphological stages involved. Fluorescence microscopy in combination with FACS allows visualisation and recovery of the progeny of experimental crosses but on this occasion the hybrids were not viable in vitro. Nevertheless, genetic exchange in L. donovani has profound epidemiological significance, because it facilitates the emergence

  14. Transcription of Toll-Like Receptors 2, 3, 4 and 9, FoxP3 and Th17 Cytokines in a Susceptible Experimental Model of Canine Leishmania infantum Infection

    Science.gov (United States)

    Hosein, Shazia; Rodríguez-Cortés, Alhelí; Blake, Damer P.; Allenspach, Karin; Alberola, Jordi; Solano-Gallego, Laia

    2015-01-01

    Canine leishmaniosis (CanL) due to Leishmania infantum is a chronic zoonotic systemic disease resulting from complex interactions between protozoa and the canine immune system. Toll-like receptors (TLRs) are essential components of the innate immune system and facilitate the early detection of many infections. However, the role of TLRs in CanL remains unknown and information describing TLR transcription during infection is extremely scarce. The aim of this research project was to investigate the impact of L. infantum infection on canine TLR transcription using a susceptible model. The objectives of this study were to evaluate transcription of TLRs 2, 3, 4 and 9 by means of quantitative reverse transcription polymerase chain reaction (qRT-PCR) in skin, spleen, lymph node and liver in the presence or absence of experimental L. infantum infection in Beagle dogs. These findings were compared with clinical and serological data, parasite densities in infected tissues and transcription of IL-17, IL-22 and FoxP3 in different tissues in non-infected dogs (n = 10), and at six months (n = 24) and 15 months (n = 7) post infection. Results revealed significant down regulation of transcription with disease progression in lymph node samples for TLR3, TLR4, TLR9, IL-17, IL-22 and FoxP3. In spleen samples, significant down regulation of transcription was seen in TLR4 and IL-22 when both infected groups were compared with controls. In liver samples, down regulation of transcription was evident with disease progression for IL-22. In the skin, upregulation was seen only for TLR9 and FoxP3 in the early stages of infection. Subtle changes or down regulation in TLR transcription, Th17 cytokines and FoxP3 are indicative of the silent establishment of infection that Leishmania is renowned for. These observations provide new insights about TLR transcription, Th17 cytokines and Foxp3 in the liver, spleen, lymph node and skin in CanL and highlight possible markers of disease susceptibility in

  15. Leishmaniasis in Turkey: Visceral and cutaneous leishmaniasis caused by Leishmania donovani in Turkey

    NARCIS (Netherlands)

    Özbilgin, Ahmet; Harman, Mehmet; Karakuş, Mehmet; Bart, Aldert; Töz, Seray; Kurt, Özgür; Çavuş, İbrahim; Polat, Erdal; Gündüz, Cumhur; van Gool, Tom; Özbel, Yusuf

    2017-01-01

    In Turkey, the main causative agents are Leishmania tropica (L. tropica) and Leishmania infantum (L. infantum) for cutaneous leishmaniasis (CL) and L. infantum for visceral leishmaniasis (VL). In this study, we investigated leishmaniasis cases caused by L. donovani and established animal models for

  16. Current Knowledge of Leishmania Vectors in Mexico: How Geographic Distributions of Species Relate to Transmission Areas

    Science.gov (United States)

    González, Camila; Rebollar-Téllez, Eduardo A.; Ibáñez-Bernal, Sergio; Becker-Fauser, Ingeborg; Martínez-Meyer, Enrique; Peterson, A. Townsend; Sánchez-Cordero, Víctor

    2011-01-01

    Leishmaniases are a group of vector-borne diseases with different clinical manifestations caused by parasites transmitted by sand fly vectors. In Mexico, the sand fly Lutzomyia olmeca olmeca is the only vector proven to transmit the parasite Leishmania mexicana to humans, which causes leishmaniasis. Other vector species with potential medical importance have been obtained, but their geographic distributions and relation to transmission areas have never been assessed. We modeled the ecological niches of nine sand fly species and projected niches to estimate potential distributions by using known occurrences, environmental coverages, and the algorithms GARP and Maxent. All vector species were distributed in areas with known recurrent transmission, except for Lu. diabolica, which appeared to be related only to areas of occasional transmission in northern Mexico. The distribution of Lu. o. olmeca does not overlap with all reported cutaneous leishmaniasis cases, suggesting that Lu. cruciata and Lu. shannoni are likely also involved as primary vectors in those areas. Our study provides useful information of potential risk areas of leishmaniasis transmission in Mexico. PMID:22049037

  17. The Transcriptome of Leishmania major Developmental Stages in Their Natural Sand Fly Vector.

    Science.gov (United States)

    Inbar, Ehud; Hughitt, V Keith; Dillon, Laura A L; Ghosh, Kashinath; El-Sayed, Najib M; Sacks, David L

    2017-04-04

    The life cycle of the Leishmania parasite in the sand fly vector involves differentiation into several distinctive forms, each thought to represent an adaptation to specific microenvironments in the midgut of the fly. Based on transcriptome sequencing (RNA-Seq) results, we describe the first high-resolution analysis of the transcriptome dynamics of four distinct stages of Leishmania major as they develop in a natural vector, Phlebotomus duboscqi The early transformation from tissue amastigotes to procyclic promastigotes in the blood-fed midgut was accompanied by the greatest number of differentially expressed genes, including the downregulation of amastins, and upregulation of multiple cell surface proteins, sugar and amino acid transporters, and genes related to glucose metabolism and cell cycle progression. The global changes accompanying post-blood meal differentiation of procyclic promastigotes to the nectomonad and metacyclic stages were less extensive, though each displayed a unique signature. The transcriptome of nectomonads, which has not been studied previously, revealed changes consistent with cell cycle arrest and the upregulation of genes associated with starvation and stress, including autophagic pathways of protein recycling. Maturation to the infective, metacyclic stage was accompanied by changes suggesting preadaptation to the intracellular environment of the mammalian host, demonstrated by the amastigote-like profiles of surface proteins and metabolism-related genes. Finally, a direct comparison between sand fly-derived and culture-derived metacyclics revealed a reassuring similarity between the two forms, with the in vivo forms distinguished mainly by a stronger upregulation of transcripts associated with nutrient stress. IMPORTANCE The life cycle of Leishmania parasites in the sand fly vector includes their growth and development as morphologically distinct forms of extracellular promastigotes found within the different microenvironments of the

  18. Persistence of phlebotomine Leishmania vectors in urban sites of Catania (Sicily, Italy).

    Science.gov (United States)

    Lisi, Oscar; D'Urso, Vera; Vaccalluzzo, Valerio; Bongiorno, Gioia; Khoury, Cristina; Severini, Francesco; Di Muccio, Trentina; Gramiccia, Marina; Gradoni, Luigi; Maroli, Michele

    2014-12-09

    Pioneering research on "Mediterranean Kala-Azar" carried out by Adler and Theodor early in the past century (~1930s) had identified Catania city (Sicily) as a major focus of the disease nowadays known as zoonotic visceral leishmaniasis (VL). Despite the fact that disease in both humans and dogs has continued to be highly prevalent in the Catania province up to the present times, research on Leishmania vectors in this urban focus dates back to that distant period. This study aimed to evaluate the persistence and current composition of the sand fly fauna in urban environments of Catania in recent years, 2006 and 2013. In 2006 fifty-one suitable collecting sites were identified within 44 sub-units of a grid drawn to include the urban Catania area. In 2013 the survey was restricted to four of the most productive and representative sites resulting from the 2006 survey. In both periods 3 collections per month were performed using standard sticky traps set for 3 days in wall holes/cavities along public roads, from the end of April through December. 43/51 sites (84.3%) were found positive for sand flies. The 2006 collections accounted for a total of 4341 specimens including six species. Among competent Leishmania vector species, P. perniciosus was the most prevalent (36.5%) being identified in all sand fly-positive sites, with significant abundance in those of the old city centre. Other species of interest were P. sergenti (2.5%) and P. neglectus (1.5%). The 2013 survey produced 1130 sand flies, of which 39.5% were P. perniciosus, 1.6% P. sergenti and 0.7% P. neglectus. A search for Leishmania DNA in a small sample of 72 P. perniciosus females revealed 11% infection prevalence. Our findings from an old urban focus of leishmaniasis demonstrate that phlebotomine sand flies have adapted fairly well to the drastic environmental changes that have occurred in cities of the Western world in the past century and still represent a potential risk for Leishmania transmission.

  19. Molecular detection and identification of Leishmania infection in naturally infected sand flies in a focus of cutaneous leishmaniasis in northern Morocco.

    Science.gov (United States)

    Es-Sette, Nargys; Ajaoud, Malika; Laamrani-Idrissi, Abderrahman; Mellouki, Fouad; Lemrani, Meryem

    2014-07-02

    Cutaneous leishmaniasis is an infectious disease caused by various species of the flagellate protozoan Leishmania. During the past 20 years, cutaneous leishmaniasis has emerged as a major public health threat in Morocco. The main objective of this study was to study the occurrence of Leishmania infection in vectors and to identify sand fly blood meal sources in an endemic locality of cutaneous leishmaniasis within Sefrou province, where the vectors of leishmaniasis were still unknown. 2650 sand flies were collected using CDC miniature light traps and identified morphologically. The identified sand flies were tested for Leishmania infection by nested PCR. The source of blood meal of 10 freshly engorged females: 6 Phlebotomus longicuspis and 4 Phlebotomus sergenti, was determined using the Cyt b sequence. The collected sand flies consisted of 10 species, seven of which belonged to the genus Phlebotomus and three to the genus Sergentomyia. The most abundant species was P. longicuspis, accounting for 72% of the total sand flies collected. In females of three P. longicuspis and four P. sergenti, Leishmania infantum and Leishmania tropica DNA was detected, respectively.The source of blood meal of engorged females showed that all sand flies tested fed on humans. We report for the first time the natural infection of P. longicuspis with L. infantum in Morocco. The high frequency of this species in this region, in addition to its anthropophilic character make P. longicuspis the putative vector of L. infantum in this cutaneous leishmaniasis focus where L. tropica is confirmed as the causative agent of the disease and P. sergenti as its vector. The presence of L. infantum, and its presumed vector in this area, makes this a site of high risk of visceral leishmaniasis, mostly because of the proximity of a focus of human and canine visceral leishmaniasis.

  20. Sergentomyia (Neophlebotomus) gemmea, a potential vector of Leishmania siamensis in southern Thailand.

    Science.gov (United States)

    Kanjanopas, Kobkan; Siripattanapipong, Suradej; Ninsaeng, Ubolrat; Hitakarun, Atitaya; Jitkaew, Somnat; Kaewtaphaya, Preecha; Tan-ariya, Peerapan; Mungthin, Mathirut; Charoenwong, Chetsuda; Leelayoova, Saovanee

    2013-07-19

    Leishmaniasis, caused by Leishmania siamensis, is an emerging disease in Thailand. Although reported cases have been increasing, epidemiological information of the disease including host and vector aspects is not clearly known. This study was a preliminary survey of the potential vector of L. siamensis in an affected area of leishmaniasis, Trang Province, southern Thailand. The collection of sandflies was performed around the area where a case of leishmaniasis was reported using CDC light traps. Species of sandfly were identified based on morphological characteristics according to Lewis's key. PCR amplification and sequencing of the heat shock protein 70 gene (hsp70) was used to identify L. siamensis DNA in sandflies. A total of 146 male and female sandflies were collected in the affected areas. Of 71 female sandflies, four species were identified, i.e., Sergentomyia (Neophlebotomus) gemmea, S. (Neophlebotomus) iyengari, S. (Parrotomyia) barraudi and Phlebotomus (Anaphlebotomus) stantoni. Among these species, S. (Neophlebotomus) gemmea was the most predominant species in all areas. DNA of L. siamensis was identified in S. (Neophlebotomus) gemmea. Nucleotide sequences of PCR products using DNA extracted from S. (Neophlebotomus) gemmea showed 99.8% identity to L. siamensis. S. (Neophlebotomus) gemmea might be a potential vector of L. siamensis in an affected area, Trang Province, southern Thailand. However further studies are needed to prove whether these sandflies can be natural vectors of leishmaniasis.

  1. Infection parameters in the sand fly vector that predict transmission of Leishmania major.

    Science.gov (United States)

    Stamper, Lisa W; Patrick, Rachel L; Fay, Michael P; Lawyer, Phillip G; Elnaiem, Dia-Eldin A; Secundino, Nagila; Debrabant, Alain; Sacks, David L; Peters, Nathan C

    2011-08-01

    To identify parameters of Leishmania infection within a population of infected sand flies that reliably predict subsequent transmission to the mammalian host, we sampled groups of infected flies and compared infection intensity and degree of metacyclogenesis with the frequency of transmission. The percentage of parasites within the midgut that were metacyclic promastigotes had the highest correlation with the frequency of transmission. Meta-analysis of multiple transmission experiments allowed us to establish a percent-metacyclic "cutoff" value that predicted transmission competence. Sand fly infections initiated with variable doses of parasites resulted in correspondingly altered percentages of metacyclic promastigotes, resulting in altered transmission frequency and disease severity. Lastly, alteration of sand fly oviposition status and environmental conditions at the time of transmission also influenced transmission frequency. These observations have implications for transmission of Leishmania by the sand fly vector in both the laboratory and in nature, including how the number of organisms acquired by the sand fly from an infection reservoir may influence the clinical outcome of infection following transmission by bite.

  2. Complete mitochondrial genome of Lutzomyia (Nyssomyia) umbratilis (Diptera: Psychodidae), the main vector of Leishmania guyanensis.

    Science.gov (United States)

    Kocher, Arthur; Gantier, Jean-Charles; Holota, Hélène; Jeziorski, Céline; Coissac, Eric; Bañuls, Anne-Laure; Girod, Romain; Gaborit, Pascal; Murienne, Jérôme

    2016-11-01

    The nearly complete mitochondrial genome of Lutzomyia umbratilis Ward & Fraiha, 1977 (Psychodidae: Phlebotominae), considered as the main vector of Leishmania guyanensis, is presented. The sequencing has been performed on an Illumina Hiseq 2500 platform, with a genome skimming strategy. The full nuclear ribosomal RNA segment was also assembled. The mitogenome of L. umbratilis was determined to be at least 15,717 bp-long and presents an architecture found in many mitogenomes of insect (13 protein-coding genes, 22 transfer RNAs, two ribosomal RNAs, and one non-coding region also referred as the control region). The control region contains a large repeated element of c. 370 bp and a poly-AT region of unknown length. This is the first mitogenome of Psychodidae to be described.

  3. The role of gallery forests in maintaining Phlebotominae populations: potential Leishmania spp. vectors in the Brazilian savanna

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    Tâmara Dias Oliveira Machado

    Full Text Available BACKGROUND Knowledge on synanthropic phlebotomines and their natural infection by Leishmania is necessary for the identification of potential areas for leishmaniasis occurrence. OBJECTIVE To analyse the occurrence of Phlebotominae in gallery forests and household units (HUs in the city of Palmas and to determine the rate of natural infection by trypanosomatids. METHODS Gallery forests and adjacent household areas were sampled on July (dry season and November (rainy season in 2014. The total sampling effort was 960 HP light traps and eight Shannon traps. Trypanosomatids were detected in Phlebotominae females through the amplification of the SSU rDNA region, and the positive samples were used in ITS1-PCR. Trypanosomatid species were identified using sequencing. FINDINGS A total of 1,527 sand flies representing 30 species were captured in which 949 (28 spp. and 578 (22 spp. were registered in July and November, respectively. In July, more specimens were captured in the gallery forests than in the HUs, and Nyssomyia whitmani was particularly frequent. In November, most of the specimens were found in the HUs, and again, Ny. whitmani was the predominant species. Lutzomyia longipalpis was commonly found in domestic areas, while Bichromomyia flaviscutellata was most frequent in gallery forests. Molecular analysis of 154 pools of females (752 specimens identified Leishmania amazonensis, L. infantum, and Crithidia fasciculata in Ny. whitmani, as well as L. amazonensis in Lu. longipalpis, Trypanosoma sp. and L. amazonensis in Pintomyia christenseni, and L. amazonensis in both Psathyromyia hermanlenti and Evandromyia walkeri. MAIN CONCLUSIONS These results show the importance of gallery forests in maintaining Phlebotominae populations in the dry month, as well as their frequent occurrence in household units in the rainy month. This is the first study to identify Leishmania, Trypanosoma, and Crithidia species in Phlebotominae collected in Palmas, Tocantins

  4. The role of gallery forests in maintaining Phlebotominae populations: potential Leishmania spp. vectors in the Brazilian savanna.

    Science.gov (United States)

    Machado, Tâmara Dias Oliveira; Minuzzi-Souza, Thaís Tâmara Castro; Ferreira, Tauana de Sousa; Freire, Luciana Pereira; Timbó, Renata Velôzo; Vital, Tamires Emanuele; Nitz, Nadjar; Silva, Mariana Neiva; Santos, Alcinei de Souza; Sales, Nathyla Morgana Cunha; Obara, Marcos Takashi; Andrade, Andrey José de; Gurgel-Gonçalves, Rodrigo

    2017-10-01

    Knowledge on synanthropic phlebotomines and their natural infection by Leishmania is necessary for the identification of potential areas for leishmaniasis occurrence. To analyse the occurrence of Phlebotominae in gallery forests and household units (HUs) in the city of Palmas and to determine the rate of natural infection by trypanosomatids. Gallery forests and adjacent household areas were sampled on July (dry season) and November (rainy season) in 2014. The total sampling effort was 960 HP light traps and eight Shannon traps. Trypanosomatids were detected in Phlebotominae females through the amplification of the SSU rDNA region, and the positive samples were used in ITS1-PCR. Trypanosomatid species were identified using sequencing. A total of 1,527 sand flies representing 30 species were captured in which 949 (28 spp.) and 578 (22 spp.) were registered in July and November, respectively. In July, more specimens were captured in the gallery forests than in the HUs, and Nyssomyia whitmani was particularly frequent. In November, most of the specimens were found in the HUs, and again, Ny. whitmani was the predominant species. Lutzomyia longipalpis was commonly found in domestic areas, while Bichromomyia flaviscutellata was most frequent in gallery forests. Molecular analysis of 154 pools of females (752 specimens) identified Leishmania amazonensis, L. infantum, and Crithidia fasciculata in Ny. whitmani, as well as L. amazonensis in Lu. longipalpis, Trypanosoma sp. and L. amazonensis in Pintomyia christenseni, and L. amazonensis in both Psathyromyia hermanlenti and Evandromyia walkeri. These results show the importance of gallery forests in maintaining Phlebotominae populations in the dry month, as well as their frequent occurrence in household units in the rainy month. This is the first study to identify Leishmania, Trypanosoma, and Crithidia species in Phlebotominae collected in Palmas, Tocantins, Brazil.

  5. Ecological Niche Modelling Predicts Southward Expansion of Lutzomyia (Nyssomyia flaviscutellata (Diptera: Psychodidae: Phlebotominae, Vector of Leishmania (Leishmania amazonensis in South America, under Climate Change.

    Directory of Open Access Journals (Sweden)

    Bruno M Carvalho

    Full Text Available Vector borne diseases are susceptible to climate change because distributions and densities of many vectors are climate driven. The Amazon region is endemic for cutaneous leishmaniasis and is predicted to be severely impacted by climate change. Recent records suggest that the distributions of Lutzomyia (Nyssomyia flaviscutellata and the parasite it transmits, Leishmania (Leishmania amazonensis, are expanding southward, possibly due to climate change, and sometimes associated with new human infection cases. We define the vector's climatic niche and explore future projections under climate change scenarios. Vector occurrence records were compiled from the literature, museum collections and Brazilian Health Departments. Six bioclimatic variables were used as predictors in six ecological niche model algorithms (BIOCLIM, DOMAIN, MaxEnt, GARP, logistic regression and Random Forest. Projections for 2050 used 17 general circulation models in two greenhouse gas representative concentration pathways: "stabilization" and "high increase". Ensemble models and consensus maps were produced by overlapping binary predictions. Final model outputs showed good performance and significance. The use of species absence data substantially improved model performance. Currently, L. flaviscutellata is widely distributed in the Amazon region, with records in the Atlantic Forest and savannah regions of Central Brazil. Future projections indicate expansion of the climatically suitable area for the vector in both scenarios, towards higher latitudes and elevations. L. flaviscutellata is likely to find increasingly suitable conditions for its expansion into areas where human population size and density are much larger than they are in its current locations. If environmental conditions change as predicted, the range of the vector is likely to expand to southeastern and central-southern Brazil, eastern Paraguay and further into the Amazonian areas of Bolivia, Peru, Ecuador

  6. Ecological Niche Modelling Predicts Southward Expansion of Lutzomyia (Nyssomyia) flaviscutellata (Diptera: Psychodidae: Phlebotominae), Vector of Leishmania (Leishmania) amazonensis in South America, under Climate Change.

    Science.gov (United States)

    Carvalho, Bruno M; Rangel, Elizabeth F; Ready, Paul D; Vale, Mariana M

    2015-01-01

    Vector borne diseases are susceptible to climate change because distributions and densities of many vectors are climate driven. The Amazon region is endemic for cutaneous leishmaniasis and is predicted to be severely impacted by climate change. Recent records suggest that the distributions of Lutzomyia (Nyssomyia) flaviscutellata and the parasite it transmits, Leishmania (Leishmania) amazonensis, are expanding southward, possibly due to climate change, and sometimes associated with new human infection cases. We define the vector's climatic niche and explore future projections under climate change scenarios. Vector occurrence records were compiled from the literature, museum collections and Brazilian Health Departments. Six bioclimatic variables were used as predictors in six ecological niche model algorithms (BIOCLIM, DOMAIN, MaxEnt, GARP, logistic regression and Random Forest). Projections for 2050 used 17 general circulation models in two greenhouse gas representative concentration pathways: "stabilization" and "high increase". Ensemble models and consensus maps were produced by overlapping binary predictions. Final model outputs showed good performance and significance. The use of species absence data substantially improved model performance. Currently, L. flaviscutellata is widely distributed in the Amazon region, with records in the Atlantic Forest and savannah regions of Central Brazil. Future projections indicate expansion of the climatically suitable area for the vector in both scenarios, towards higher latitudes and elevations. L. flaviscutellata is likely to find increasingly suitable conditions for its expansion into areas where human population size and density are much larger than they are in its current locations. If environmental conditions change as predicted, the range of the vector is likely to expand to southeastern and central-southern Brazil, eastern Paraguay and further into the Amazonian areas of Bolivia, Peru, Ecuador, Colombia and Venezuela

  7. The flagellar protein FLAG1/SMP1 is a candidate for Leishmania-sand fly interaction.

    Science.gov (United States)

    Di-Blasi, Tatiana; Lobo, Amanda R; Nascimento, Luanda M; Córdova-Rojas, Jose L; Pestana, Karen; Marín-Villa, Marcel; Tempone, Antonio J; Telleria, Erich L; Ramalho-Ortigão, Marcelo; McMahon-Pratt, Diane; Traub-Csekö, Yara M

    2015-03-01

    Leishmaniasis is a serious problem that affects mostly poor countries. Various species of Leishmania are the agents of the disease, which take different clinical manifestations. The parasite is transmitted by sandflies, predominantly from the Phlebotomus genus in the Old World and Lutzomyia in the New World. During development in the gut, Leishmania must survive various challenges, which include avoiding being expelled with blood remnants after digestion. It is believed that attachment to the gut epithelium is a necessary step for vector infection, and molecules from parasites and sand flies have been implicated in this attachment. In previous work, monoclonal antibodies were produced against Leishmania. Among these an antibody was obtained against Leishmania braziliensis flagella, which blocked the attachment of Leishmania panamensis flagella to Phlebotomus papatasi guts. The protein recognized by this antibody was identified and named FLAG1, and the complete FLAG1 gene sequence was obtained. This protein was later independently identified as a small, myristoylated protein and called SMP1, so from now on it will be denominated FLAG1/SMP1. The FLAG1/SMP1 gene is expressed in all developmental stages of the parasite, but has higher expression in promastigotes. The anti-FLAG1/SMP1 antibody recognized the flagellum of all Leishmania species tested and generated the expected band by western blots. This antibody was used in attachment and infection blocking experiments. Using the New World vector Lutzomyia longipalpis and Leishmania infantum chagasi, no inhibition of attachment ex vivo or infection in vivo was seen. On the other hand, when the Old World vectors P. papatasi and Leishmania major were used, a significant decrease of both attachment and infection were seen in the presence of the antibody. We propose that FLAG1/SMP1 is involved in the attachment/infection of Leishmania in the strict vector P. papatasi and not the permissive vector L. longipalpis.

  8. Lutzomyia umbratilis, the main vector of Leishmania guyanensis, represents a novel species complex?

    Science.gov (United States)

    Scarpassa, Vera Margarete; Alencar, Ronildo Baiatone

    2012-01-01

    Lutzomyia umbratilis is an important Leishmania guyanensis vector in South America. Previous studies have suggested differences in the vector competence between L. umbratilis populations situated on opposite banks of the Amazonas and Negro Rivers in the central Amazonian Brazil region, likely indicating a species complex. However, few studies have been performed on these populations and the taxonomic status of L. umbratilis remains unclear. Phylogeographic structure was estimated for six L. umbratilis samples from the central Amazonian region in Brazil by analyzing mtDNA using 1181 bp of the COI gene to assess whether the populations on opposite banks of these rivers consist of incipient or distinct species. The genetic diversity was fairly high and the results revealed two distinct clades ( = lineages) with 1% sequence divergence. Clade I consisted of four samples from the left bank of the Amazonas and Negro Rivers, whereas clade II comprised two samples from the right bank of Negro River. No haplotypes were shared between samples of two clades. Samples within clades exhibited low to moderate genetic differentiation (F(ST) = -0.0390-0.1841), whereas samples between clades exhibited very high differentiation (F(ST) = 0.7100-0.8497) and fixed differences. These lineages have diverged approximately 0.22 Mya in the middle Pleistocene. Demographic expansion was detected for the lineages I and II approximately 30,448 and 15,859 years ago, respectively, in the late Pleistocene. The two genetic lineages may represent an advanced speciation stage suggestive of incipient or distinct species within L. umbratilis. These findings suggest that the Amazonas and Negro Rivers may be acting as effective barriers, thus preventing gene flow between populations on opposite sides. Such findings have important implications for epidemiological studies, especially those related to vector competence and anthropophily, and for vector control strategies. In addition, L

  9. First molecular detection of Leishmania tarentolae-like DNA in Sergentomyia minuta in Spain.

    Science.gov (United States)

    Bravo-Barriga, Daniel; Parreira, Ricardo; Maia, Carla; Blanco-Ciudad, Juan; Afonso, Maria Odete; Frontera, Eva; Campino, Lenea; Pérez-Martín, Juan Enrique; Serrano Aguilera, Francisco Javier; Reina, David

    2016-03-01

    Phlebotomine sand flies (Diptera, Psychodidae) are vectors of multiple Leishmania species, among which Leishmania infantum stands out as a being frequently pathogenic to humans and dogs in Mediterranean countries. In this study, Sergentomyia minuta sand flies were collected using CDC miniature light traps in different 431 biotopes from Southwest Spain. A total of 114 females were tested for the presence of Leishmania DNA by targeting ITS-1 and cyt-B sequences by PCR. Leishmania DNA was detected in one S. minuta. Characterization of the obtained DNA sequences by phylogenetic analyses revealed close relatedness with Leishmania tarentolae Wenyon, 1921 as well as with both human and canine pathogenic strains of Asian origin (China), previously described as Leishmania sp. To our knowledge, this is the first report of phlebotomine sand flies naturally infected with L. tarentolae-like in Spain. The possible infection of sand flies with novel Leishmania species should be taken into consideration in epidemiological studies of vector species in areas where leishmaniosis is endemic.

  10. The inflammatory cytokine effect of Pam3CSK4 TLR2 agonist alone or in combination with Leishmania infantum antigen on ex-vivo whole blood from sick and resistant dogs.

    Science.gov (United States)

    Martínez-Orellana, Pamela; Quirola-Amores, Paulina; Montserrat-Sangrà, Sara; Ordeix, Laura; Llull, Joan; Álvarez-Fernández, Alejandra; Solano-Gallego, Laia

    2017-03-13

    A wide spectrum of clinical manifestations and immune responses exist in canine L. infantum infection. Ibizan hounds are more "resistant" to disease than other dog breeds. Recognition of pathogen-associated molecule patterns by toll like receptors (TLRs) rapidly triggers a variety of anti-microbial immune responses through the induction of pro-inflammatory cytokines such as TNF-α and IL-6 which may play an important role in controlling Leishmania infection. The main objective of this study was to investigate and compare the effect of a TLR2 agonist (TLR2a) alone or in combination with L. infantum antigen (LSA) on ex vivo whole blood cytokine production from healthy seronegative IFN-γ non-producer dogs from an area of low in canine leishmaniosis endemicity (n = 11); sick seropositive dogs with low production of IFN-γ (n = 17) and healthy seronegative or low positive Ibizan hounds with a predominant IFN-γ production (n = 21) from a highly endemic area. Whole blood was stimulated with medium alone (Ø), LSA, concanavalin A, TLR2 (Pam3CSK4) receptor agonist (Ø + TLR2a) and TLR2a and LSA (LSA + TLR2a) for 48 h. Supernatants were harvested for measurement of canine TNF-α and IL-6 cytokines by ELISA. A significant increase of TNF-α was found in the supernatants of stimulated blood from all groups (Ø + TLR2a and LSA + TLR2a) when compared with medium alone. A similar pattern was observed for IL-6. Interestingly, a significant increase of TNF-α production was only observed when stimulation with LSA + TLR2a was compared with TLR2a alone in Ibizan hounds. A significant increase of TNF-α production was observed with stimulation of LSA + TLR2a when compared with LSA in all groups. Significantly higher concentrations of TNF-α and IL-6 were detected in Ibizan hounds, especially for the Ø + TLR2a and LSA + TLR2a treatments compared with other groups. This study demonstrated that TLR2a alone enhances the production of the

  11. Structure-based domain assignment in Leishmania infantum EndoG: characterization of a pH-dependent regulatory switch and a C-terminal extension that largely dictates DNA substrate preferences.

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    Oliva, Cristina; Sánchez-Murcia, Pedro A; Rico, Eva; Bravo, Ana; Menéndez, Margarita; Gago, Federico; Jiménez-Ruiz, Antonio

    2017-09-06

    Mitochondrial endonuclease G from Leishmania infantum (LiEndoG) participates in the degradation of double-stranded DNA (dsDNA) during parasite cell death and is catalytically inactive at a pH of 8.0 or above. The presence, in the primary sequence, of an acidic amino acid-rich insertion exclusive to trypanosomatids and its spatial position in a homology-built model of LiEndoG led us to postulate that this peptide stretch might act as a pH sensor for self-inhibition. We found that a LiEndoG variant lacking residues 145-180 is indeed far more active than its wild-type counterpart at pH values >7.0. In addition, we discovered that (i) LiEndoG exists as a homodimer, (ii) replacement of Ser211 in the active-site SRGH motif with the canonical aspartate from the DRGH motif of other nucleases leads to a catalytically deficient enzyme, (iii) the activity of the S211D variant can be restored upon the concomitant replacement of Ala247 with Arg and (iv) a C-terminal extension is responsible for the observed preferential cleavage of single-stranded DNA (ssDNA) and ssDNA-dsDNA junctions. Taken together, our results support the view that LiEndoG is a multidomain molecular machine whose nuclease activity can be subtly modulated or even abrogated through architectural changes brought about by environmental conditions and interaction with other binding partners. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  12. Serological cross-reactivity of Trypanosoma cruzi, Ehrlichia canis, Toxoplasma gondii, Neospora caninum and Babesia canis to Leishmania infantum chagasi tests in dogs

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    Maurício Franco Zanette

    2014-01-01

    Full Text Available Introduction: The aim of this study was to evaluate the serological cross-reactivity between Leishmania sp. and other canine pathogens. Methods: Positive serum samples for Ehrlichia canis, Babesia canis, Toxoplasma gondii, Neospora caninum and Trypanosoma cruzi were tested using three serological methods enzyme linked immunosorbent assay (ELISA, indirect immunofluorescent antibody test (IFAT and Kalazar Detect™, for canine visceral leishmaniasis. Results: Of the 57 dog samples tested, 24 (42.1% tested positive using one of the three serological methods: 10/57 (17.5% for ELISA, 11/57 (19.3% for IFAT and 3/57 (5.3% for Kalazar Detect™. Conclusions: Our results demonstrated that the presence of other infectious agents may lead to cross-reactivity on leishmaniasis serological tests.

  13. Molecular identification of vectors of Leishmania in Colombia: mitochondrial introgression in the Lutzomyia townsendi series.

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    Testa, J M; Montoya-Lerma, J; Cadena, H; Oviedo, M; Ready, P D

    2002-12-01

    The identity of the sandfly vectors of Leishmania braziliensis in Valle del Cauca Department, Colombia, was originally given as Lutzomyia townsendi, but then changed to L. youngi, another member of the L. townsendi series (Verrucarum group) with isomorphic females. To identify members of this series in Valle del Cauca, we analyzed the nuclear gene elongation factor-alpha (EF-alpha) and the mitochondrial gene cytochrome b (Cyt b). DNA sequences from the L. verrucarum series (L. columbiana, L. evansi and L. ovallesi) were used as outgroups. Flies from two locations on the western cordillera of the Andes were identified as L. townsendi s.s., according to male morphology and distinctive gene lineages. In the third location, on the central cordillera of the Andes, most specimens were identified as belonging to a geographical population of L. youngi, according to male morphology, an EF-alpha lineage shared with L. youngi from the Venezuelan-type locality, and a distinctive Cyt b sub-lineage. All other specimens were identified as L. youngi with the introgressed Cyt b sequences of L. townsendi. Such interspecific introgression implies that vectorial traits and ecological associations may no longer be viewed as fixed properties of different morphospecies.

  14. Transcriptional Profiling of Immune-Related Genes in Leishmania infantum-Infected Mice: Identification of Potential Biomarkers of Infection and Progression of Disease

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    Eduardo Ontoria

    2018-06-01

    Full Text Available Leishmania spp. is a protozoan parasite that affects millions of people around the world. At present, there is no effective vaccine to prevent leishmaniases in humans. A major limitation in vaccine development is the lack of precise understanding of the particular immunological mechanisms that allow parasite survival in the host. The parasite-host cell interaction induces dramatic changes in transcriptome patterns in both organisms, therefore, a detailed analysis of gene expression in infected tissues will contribute to the evaluation of drug and vaccine candidates, the identification of potential biomarkers, and the understanding of the immunological pathways that lead to protection or progression of disease. In this large-scale analysis, differential expression of 112 immune-related genes has been analyzed using high-throughput qPCR in spleens of infected and naïve Balb/c mice at four different time points. This analysis revealed that early response against Leishmania infection is characterized by the upregulation of Th1 markers and M1-macrophage activation molecules such as Ifng, Stat1, Cxcl9, Cxcl10, Ccr5, Cxcr3, Xcl1, and Ccl3. This activation doesn't protect spleen from infection, since parasitic burden rises along time. This marked difference in gene expression between infected and control mice disappears during intermediate stages of infection, probably related to the strong anti-inflammatory and immunosuppresory signals that are activated early upon infection (Ctla4 or remain activated throughout the experiment (Il18bp. The overexpression of these Th1/M1 markers is restored later in the chronic phase (8 wpi, suggesting the generation of a classical “protective response” against leishmaniasis. Nonetheless, the parasitic burden rockets at this timepoint. This apparent contradiction can be explained by the generation of a regulatory immune response characterized by overexpression of Ifng, Tnfa, Il10, and downregulation Il4 that

  15. First Human Cases of Leishmania (Viannia) lainsoni Infection and a Search for the Vector Sand Flies in Ecuador.

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    Kato, Hirotomo; Bone, Abdon E; Mimori, Tatsuyuki; Hashiguchi, Kazue; Shiguango, Gonzalo F; Gonzales, Silvio V; Velez, Lenin N; Guevara, Angel G; Gomez, Eduardo A; Hashiguchi, Yoshihisa

    2016-05-01

    An epidemiological study of leishmaniasis was performed in Amazonian areas of Ecuador since little information on the prevalent Leishmania and sand fly species responsible for the transmission is available. Of 33 clinical specimens from patients with cutaneous leishmaniasis (CL), causative parasites were identified in 25 samples based on cytochrome b gene analysis. As reported previously, Leishmania (Viannia) guyanensis and L. (V.) braziliensis were among the causative agents identified. In addition, L. (V.) lainsoni, for which infection is reported in Brazil, Bolivia, Peru, Suriname, and French Guiana, was identified in patients with CL from geographically separate areas in the Ecuadorian Amazon, corroborating the notion that L. (V.) lainsoni is widely distributed in South America. Sand flies were surveyed around the area where a patient with L. (V.) lainsoni was suspected to have been infected. However, natural infection of sand flies by L. (V.) lainsoni was not detected. Further extensive vector searches are necessary to define the transmission cycle of L. (V.) lainsoni in Ecuador.

  16. Lutzomyia longipalpis saliva or salivary protein LJM19 protects against Leishmania braziliensis and the saliva of its vector, Lutzomyia intermedia.

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    Natalia M Tavares

    Full Text Available BACKGROUND: Leishmania transmission occurs in the presence of insect saliva. Immunity to Phlebotomus papatasi or Lutzomyia longipalpis saliva or salivary components confers protection against an infection by Leishmania in the presence of the homologous saliva. However, immunization with Lutzomyia intermedia saliva did not protect mice against Leishmania braziliensis plus Lu. intermedia saliva. In the present study, we have studied whether the immunization with Lu. longipalpis saliva or a DNA plasmid coding for LJM19 salivary protein would be protective against L. braziliensis infection in the presence of Lu. intermedia saliva, the natural vector for L. braziliensis. METHODOLOGY/PRINCIPAL FINDINGS: Immunization with Lu. longipalpis saliva or with LJM19 DNA plasmid induced a Delayed-Type Hypersensitivity (DTH response against Lu. longipalpis as well as against a Lu. intermedia saliva challenge. Immunized and unimmunized control hamsters were then intradermally infected in the ears with L. braziliensis in the presence of Lu. longipalpis or Lu. intermedia saliva. Animals immunized with Lu. longipalpis saliva exhibited smaller lesion sizes as well as reduced disease burdens both at lesion site and in the draining lymph nodes. These alterations were associated with a significant decrease in the expression levels of IL-10 and TGF-β. Animals immunized with LJM19 DNA plasmid presented similar findings in protection and immune response and additionally increased IFN-γ expression. CONCLUSIONS/SIGNIFICANCE: Immunization with Lu. longipalpis saliva or with a DNA plasmid coding LJM19 salivary protein induced protection in hamsters challenged with L. braziliensis plus Lu. intermedia saliva. These findings point out an important role of immune response against saliva components, suggesting the possibility to develop a vaccine using a single component of Lu. longipalpis saliva to generate protection against different species of Leishmania, even those

  17. Assessing the importance of four sandfly species (Diptera: Psychodidae) as vectors of Leishmania mexicana in Campeche, Mexico.

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    Pech-May, A; Peraza-Herrera, G; Moo-Llanes, D A; Escobedo-Ortegón, J; Berzunza-Cruz, M; Becker-Fauser, I; Montes DE Oca-Aguilar, A C; Rebollar-Téllez, E A

    2016-09-01

    Localized cutaneous leishmaniasis represents a public health problem in many areas of Mexico, especially in the Yucatan Peninsula. An understanding of vector ecology and bionomics is of great importance in evaluations of the transmission dynamics of Leishmania parasites. A field study was conducted in the county of Calakmul, state of Campeche, during the period from November 2006 to March 2007. Phlebotomine sandfly vectors were sampled using Centers for Disease Control light traps, baited Disney traps and Shannon traps. A total of 3374 specimens were captured in the two villages of Once de Mayo (93.8%) and Arroyo Negro (6.1%). In Once de Mayo, the most abundant species were Psathyromyia shannoni, Lutzomyia cruciata, Bichromomyia olmeca olmeca and Psychodopygus panamensis (all: Diptera: Psychodidae). The Shannon trap was by far the most efficient method of collection. The infection rate, as determined by Leishmania mexicana-specific polymerase chain reaction, was 0.3% in Once de Mayo and infected sandflies included Psy. panamensis, B. o. olmeca and Psa. shannoni. There were significant differences in human biting rates across sandfly species and month of sampling. Ecological niche modelling analyses showed an overall overlap of 39.1% for the four species in the whole state of Campeche. In addition, the finding of nine vector-reservoir pairs indicates a potential interaction. The roles of the various sandfly vectors in Calakmul are discussed. © 2016 The Royal Entomological Society.

  18. Phenotypic and genotypic variations among three allopatric populations of Lutzomyia umbratilis, main vector of Leishmania guyanensis.

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    de Souza Freitas, Moises Thiago; Ríos-Velasquez, Claudia Maria; Costa, César Raimundo Lima; Figueirêdo, Carlos Alberto Santiago; Aragão, Nádia Consuelo; da Silva, Lidiane Gomes; de Aragão Batista, Marcus Vinicius; Balbino, Teresa Cristina Leal; Pessoa, Felipe Arley Costa; de Queiroz Balbino, Valdir

    2015-09-04

    In South America, Lutzomyia umbratilis is the main vector of Leishmania guyanensis, one of the species involved in the transmission of American tegumentary leishmaniasis. In Brazil, L. umbratilis has been recorded in the Amazon region, and in the state of Pernambuco, Northeastern region, where an isolated population has been identified. This study assessed the phylogeographic structure and size and shape differences of the wing of three Brazilian populations. Samples of L. umbratilis were collected from Rio Preto da Eva (north of the Amazon River, Amazonas), from Manacapuru (south of the Amazon River), and from the isolated population in Recife, Pernambuco state. These samples were processed to obtain sequences of the Cytochrome Oxidase I mitochondrial gene. Geometrics morphometry analysis of the right wing shape of the three populations was made using discriminate canonical analysis. Phylogenetic analysis revealed the presence of two distinct monophyletic clades: one clade comprised of the Recife and Rio Preto da Eva samples, and the other clade comprised of the Manacapuru samples. Comparing the Manacapuru population with the Recife and Rio Preto da Eva populations generated high indices of interpopulational divergence. Geometric morphometry analysis indicated two distinct groups between the studied populations. Canonical variate analysis of wing shape indicated that Rio Preto da Eva population is significantly closer to Recife population, and both populations were genetically distant from Manacapuru. The polymorphic sites and geometric morphometry analysis indicate that the distance, lack of continuity and environmental differences have not modified the ancestral relationship between Recife and Rio Preto da Eva populations. The genetic and morphological similarities shared by the Recife and Rio Preto da Eva populations suggest that these populations are more closely related evolutionarily. These results confirm the existence of an L. umbratilis species complex in

  19. Evaluation of four molecular methods to detect Leishmania infection in dogs.

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    Albuquerque, Andreia; Campino, Lenea; Cardoso, Luís; Cortes, Sofia

    2017-03-13

    Canine leishmaniasis, a zoonotic disease caused by Leishmania infantum vectored by phlebotomine sand flies, is considered a relevant veterinary and public health problem in various countries, namely in the Mediterranean basin and Brazil, where dogs are considered the main reservoir hosts. Not only diseased dogs but also those subclinically infected play a relevant role in the transmission of L. infantum to vectors; therefore, early diagnosis is essential, under both a clinical and an epidemiological perspective. Molecular tools can be a more accurate and sensitive approach for diagnosis, with a wide range of protocols currently in use. The aim of the present report was to compare four PCR based protocols for the diagnosis of canine Leishmania infection in a cohort of dogs from the Douro region, Portugal. A total of 229 bone marrow samples were collected from dogs living in the Douro region, an endemic region for leishmaniasis. Four PCR protocols were evaluated for Leishmania DNA detection in canine samples, three single (ITS1-PCR, MC-PCR and Uni21/Lmj4-PCR) and one nested (nested SSU rRNA-PCR). Two of the protocols were based on nuclear targets and the other two on kinetoplastid targets. The higher overall percentage of infected dogs was detected with the nested SSU rRNA-PCR (37.6%), which also was able to detect Leishmania DNA in a higher number of samples from apparently healthy dogs (25.3%). The ITS1-PCR presented the lowest level of Leishmania detection. Nested SSU rRNA-PCR is an appropriate method to detect Leishmania infection in dogs. Accurate and early diagnosis in clinically suspect as well as apparently healthy dogs is essential, in order to treat and protect animals and public health and contribute to the control and awareness of the disease.

  20. El "Status" taxonomico de leishmania garnhami, indicado por su patron de desarrollo en el Vector The taxonomic status of Leishmania garnhami, indicated by its development pattern in the vector

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    N. Añez

    1985-03-01

    Full Text Available Bajo condiciones experimentales, se estabelece el de desarrollo primario de Leishmania garnhami em su vector Lutzomyia townsendi, siguiendo disecciones diarias entre el Iro y 12mo dia post-imnfección. Se demuestra que L. garnhami puede lograr el estabelecimiento inicial de la infección en cualquiera de las regiones del intestino y en los túbulos de Malpighi del insecto vector. Un comportamiento similar fue observado en ejemplares de Lu. longipalpis. Se discute la susceptibilidad de Lu. townsendi a las infecciones por L. mexicana, L. braziliensis y L. garnhami. Se concluye que L. garnhami, a juzgar por su comportamiento en el vector, debe considerarse distinta a las especies del complejo L.mexicana y que su patrón de desarrollo difiere del de las especies de la Sección Suprapylaria, ya que invaden el triângulo pilórico, y del de la Peripylaria por su invasión a los túbulos de Malpighi.Under experimental conditions, the developmental pattern of Leishmania garnhami in its vector Lutzomyia townsendi, was established following daily dissections between the 1st and the 12th day post-infection. L. garnhami succeeded in establishing initial infections in any of the gut regions and in the Malpighian tubules of infected sandflies. Similar behavior was observed in infected Lu. longipalpis. The distribution of L. garnhami in the digestive tract of infected flies, is different to that observed in species of the L. mexicana and L. braziliensis complexes. The susceptibility of Lu. townsendi to infection by L. mexicana, L. braziliensis and L. garnhami is discussed. It is concluded that L. garnhami, according to its behaviour in the vector, must be considered different from the l. mexicana complex, and that its developmental pattern differs from that of the Suprapylaria, because it invades the hind triangle and also from the species of the Peripylaria by its invasion of the Malpighian tubules.

  1. The mating competence of geographically diverse Leishmania major strains in their natural and unnatural sand fly vectors.

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    Inbar, Ehud; Akopyants, Natalia S; Charmoy, Melanie; Romano, Audrey; Lawyer, Phillip; Elnaiem, Dia-Eldin A; Kauffmann, Florence; Barhoumi, Mourad; Grigg, Michael; Owens, Katherine; Fay, Michael; Dobson, Deborah E; Shaik, Jahangheer; Beverley, Stephen M; Sacks, David

    2013-01-01

    Invertebrate stages of Leishmania are capable of genetic exchange during their extracellular growth and development in the sand fly vector. Here we explore two variables: the ability of diverse L. major strains from across its natural range to undergo mating in pairwise tests; and the timing of the appearance of hybrids and their developmental stage associations within both natural (Phlebotomus duboscqi) and unnatural (Lutzomyia longipalpis) sand fly vectors. Following co-infection of flies with parental lines bearing independent drug markers, doubly-drug resistant hybrid progeny were selected, from which 96 clonal lines were analyzed for DNA content and genotyped for parent alleles at 4-6 unlinked nuclear loci as well as the maxicircle DNA. As seen previously, the majority of hybrids showed '2n' DNA contents, but with a significant number of '3n' and one '4n' offspring. In the natural vector, 97% of the nuclear loci showed both parental alleles; however, 3% (4/150) showed only one parental allele. In the unnatural vector, the frequency of uniparental inheritance rose to 10% (27/275). We attribute this to loss of heterozygosity after mating, most likely arising from aneuploidy which is both common and temporally variable in Leishmania. As seen previously, only uniparental inheritance of maxicircle kDNA was observed. Hybrids were recovered at similar efficiencies in all pairwise crosses tested, suggesting that L. major lacks detectable 'mating types' that limit free genetic exchange. In the natural vector, comparisons of the timing of hybrid formation with the presence of developmental stages suggest nectomonads as the most likely sexually competent stage, with hybrids emerging well before the first appearance of metacyclic promastigotes. These studies provide an important perspective on the prevalence of genetic exchange in natural populations of L. major and a guide for experimental studies to understand the biology of mating.

  2. A Poly(Lactic-co-Glycolic) Acid Nanovaccine Based on Chimeric Peptides from Different Leishmania infantum Proteins Induces Dendritic Cells Maturation and Promotes Peptide-Specific IFNγ-Producing CD8+ T Cells Essential for the Protection against Experimental Visceral Leishmaniasis.

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    Athanasiou, Evita; Agallou, Maria; Tastsoglou, Spyros; Kammona, Olga; Hatzigeorgiou, Artemis; Kiparissides, Costas; Karagouni, Evdokia

    2017-01-01

    Visceral leishmaniasis, caused by Leishmania ( L .) donovani and L. infantum protozoan parasites, can provoke overwhelming and protracted epidemics, with high case-fatality rates. An effective vaccine against the disease must rely on the generation of a strong and long-lasting T cell immunity, mediated by CD4 + T H1 and CD8 + T cells. Multi-epitope peptide-based vaccine development is manifesting as the new era of vaccination strategies against Leishmania infection. In this study, we designed chimeric peptides containing HLA-restricted epitopes from three immunogenic L. infantum proteins (cysteine peptidase A, histone H1, and kinetoplastid membrane protein 11), in order to be encapsulated in poly(lactic- co -glycolic) acid nanoparticles with or without the adjuvant monophosphoryl lipid A (MPLA) or surface modification with an octapeptide targeting the tumor necrosis factor receptor II. We aimed to construct differentially functionalized peptide-based nanovaccine candidates and investigate their capacity to stimulate the immunomodulatory properties of dendritic cells (DCs), which are critical regulators of adaptive immunity generated upon vaccination. According to our results, DCs stimulation with the peptide-based nanovaccine candidates with MPLA incorporation or surface modification induced an enhanced maturation profile with prominent IL-12 production, promoting allogeneic T cell proliferation and intracellular production of IFNγ by CD4 + and CD8 + T cell subsets. In addition, DCs stimulated with the peptide-based nanovaccine candidate with MPLA incorporation exhibited a robust transcriptional activation, characterized by upregulated genes indicative of vaccine-driven DCs differentiation toward type 1 phenotype. Immunization of HLA A2.1 transgenic mice with this peptide-based nanovaccine candidate induced peptide-specific IFNγ-producing CD8 + T cells and conferred significant protection against L. infantum infection. Concluding, our findings supported that

  3. A putative ATP/GTP binding protein affects Leishmania mexicana growth in insect vectors and vertebrate hosts

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    Hlaváčová, Jana; Zimmer, Sara L.; Butenko, Anzhelika; Podešvová, Lucie; Leštinová, Tereza; Lukeš, Julius; Kostygov, Alexei; Votýpka, Jan; Volf, Petr

    2017-01-01

    Background Leishmania virulence factors responsible for the complicated epidemiology of the various leishmaniases remain mainly unidentified. This study is a characterization of a gene previously identified as upregulated in two of three overlapping datasets containing putative factors important for Leishmania’s ability to establish mammalian intracellular infection and to colonize the gut of an insect vector. Methodology/Principal findings The investigated gene encodes ATP/GTP binding motif-containing protein related to Leishmania development 1 (ALD1), a cytosolic protein that contains a cryptic ATP/GTP binding P-loop. We compared differentiation, growth rates, and infective abilities of wild-type and ALD1 null mutant cell lines of L. mexicana. Loss of ALD1 results in retarded growth kinetics but not defects in differentiation in axenic culture. Similarly, when mice and the sand fly vector were infected with the ALD1 null mutant, the primary difference in infection and colonization phenotype relative to wild type was an inability to achieve maximal host pathogenicity. While ability of the ALD1 null mutant cells to infect macrophages in vitro was not affected, replication within macrophages was clearly curtailed. Conclusions/Significance L. mexicana ALD1, encoding a protein with no assigned functional domains or motifs, was identified utilizing multiple comparative analyses with the related and often experimentally overlooked monoxenous flagellates. We found that it plays a role in Leishmania infection and colonization in vitro and in vivo. Results suggest that ALD1 functions in L. mexicana’s general metabolic network, rather than function in specific aspect of virulence as anticipated from the compared datasets. This result validates our comparative genomics approach for finding relevant factors, yet highlights the importance of quality laboratory-based analysis of genes tagged by these methods. PMID:28742133

  4. KSAC, a defined Leishmania antigen, plus adjuvant protects against the virulence of L. major transmitted by its natural vector Phlebotomus duboscqi.

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    Regis Gomes

    Full Text Available Recombinant KSAC and L110f are promising Leishmania vaccine candidates. Both antigens formulated in stable emulsions (SE with the natural TLR4 agonist MPL® and L110f with the synthetic TLR4 agonist GLA in SE protected BALB/c mice against L. major infection following needle challenge. Considering the virulence of vector-transmitted Leishmania infections, we vaccinated BALB/c mice with either KSAC+GLA-SE or L110f+GLA-SE to assess protection against L. major transmitted via its vector Phlebotomus duboscqi.Mice receiving the KSAC or L110f vaccines were challenged by needle or L. major-infected sand flies. Weekly disease progression and terminal parasite loads were determined. Immunological responses to KSAC, L110f, or soluble Leishmania antigen (SLA were assessed throughout vaccination, three and twelve weeks after immunization, and one week post-challenge.Following sand fly challenge, KSAC-vaccinated mice were protected while L110f-vaccinated animals showed partial protection. Protection correlated with the ability of SLA to induce IFN-γ-producing CD4(+CD62L(lowCCR7(low effector memory T cells pre- and post-sand fly challenge.This study demonstrates the protective efficacy of KSAC+GLA-SE against sand fly challenge; the importance of vector-transmitted challenge in evaluating vaccine candidates against Leishmania infection; and the necessity of a rapid potent Th1 response against Leishmania to attain true protection.

  5. Importance of individual analysis of environmental and climatic factors affecting the density of Leishmania vectors living in the same geographical area: the example of Phlebotomus ariasi and P. perniciosus in northeast Spain

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    Cristina Ballart

    2014-05-01

    Full Text Available The aim of the present study was to determine the role of specific environmental and climatic factors affecting the distribution and density of Phlebotomus ariasi and P. perniciosus, the proven vectors for Leishmania infantum in Spain. An entomological study was carried out in July 2006 in the province of Lleida with sticky traps set in their diurnal resting places at altitudes ranging from 86 to 1,755 m above the mean sea level (339 sites were sampled. Bivariate analysis revealed that factors such as altitude, bioclimatic zone, temperature, precipitation, sampling site (site relative to settlement, site situation, site category, wall vegetation, particular environment (in this case a natural park, general environment, adjacent natural vegetation and land cover were significantly associated with sand fly densities. The multivariate model for P. perniciosus revealed that its density was affected by site and land cover. Specifically, paved driveways correlated negatively with vector density (Incidence Risk Ratio (IRR: 0.41 and arable land cover correlated positively (IRR: 4.59. In the case of P. ariasi, a significant correlation was observed with the altitude and bioclimatic zone, with density increasing at >800 m above the mean sea level (IRR: 3.40 and decreasing in the meso-Mediterranean bioclimatic zone (IRR: 0.08. Both species were mostly found in agricultural and forest areas far from domestic environments. However, the two species correlated differently with altitude, bio-climate, vegetation, temperature and precipitation, which emphasises the importance of their individual analysis in studies regarding risk of leishmaniasis transmission.

  6. Vectors of Leishmania braziliensis in the Petén, Guatemala.

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    Rowton, E; de Mata, M; Rizzo, N; Navin, T; Porter, C

    1991-12-01

    During a 1-year study, 13 species of sand fly were collected in bite-landing collections on human attractants in Tikal, Guatemala. Using isoenzyme analysis, Leishmania braziliensis was identified among isolates from Lutzomyia ovallesi, Lu. panamensis, and Lu. ylephiletor. Lutzomyia ovallesi, Lu. shannoni, and Lu. cruciata were found with flagellates whose isoenzyme patterns matched unidentified flagellates isolated from a patient with mucosal lesions.

  7. Ground ULV and thermal fog applications against Phlebotomine sand fly vectors of Leishmania in a hot arid environment in western Kenya

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    Phlebotomine sand fly vectors of Leishmania continue to threaten US military operations in Africa, Southwest Asia, and the Middle East. Ultra-low volume (ULV) and/or thermal fog pesticide dispersal are potentially effective against sand flies, but operational guidance is thinly based on mosquito con...

  8. Infective stages of Leishmania in the sandfly vector and some observations on the mechanism of transmission Formas infectante de Leishmania no vetor flebotomíneo e algumas observações sobre o mecanismo de transmissão

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    Ralph Lainson

    1987-09-01

    Full Text Available Infective stages of Leishmania (Leishmania amazonensis, capable of producing amastigote infections in hamster skin, were shown to be present in the experimentally infected sandfly vector Lutzomyia flaviscutellata 15, 25, 40, 49, 70, 96 and 120 hours after the flies had received their infective blood-meal. Similarly, infective stages of Leishmania (L. chagasi were demonstrated in the experimentally infected vector Lu. longipalpis examined 38, 50, 63, 87, 110, 135, 171 and 221 hours following the infective blood-meal, by the intraperitoneal inoculation of the flagellates into hamsters. The question of whether or not transmission by the bite of the sandfly is dependent on the presence of [quot ]metacyclic[quot ] promastigotes in the mouthparts of the vector is discussed.Foi demonstrado através de infecção experimental, que estágios infectivos de Leishmania (L. amazonensis, capazes de produzir infecção na pele do hamster, encontram-se presentes no vetor flebotomíneo Lutzomyia flaviscutellata 15, 25, 40, 49, 70, 96 e 120 horas após o inseto ter recebido sua refeição sangüínea infectiva. Da mesma maneira, foi comprovada a presença de estágios infectivos de L. (L. chagasi em exemplares do vetor Lu. longipalpis, examinados 38, 50, 63, 87, 110, 135, 171 e 221 horas após o repasto sangüíneo infectivo - através da inoculação em hamster por via intraperitoneal dos flagelados obtidos desses fle botomíneos. A questão sobre a transmissão do gênero Leishmania pelo flebotomíneo ser ou não dependente da presença de promastigotos "metacíclios" na proboscis do vetor, é discutida.

  9. First Human Cases of Leishmania (Viannia) lainsoni Infection and a Search for the Vector Sand Flies in Ecuador

    Science.gov (United States)

    Kato, Hirotomo; Bone, Abdon E.; Mimori, Tatsuyuki; Hashiguchi, Kazue; Shiguango, Gonzalo F.; Gonzales, Silvio V.; Velez, Lenin N.; Guevara, Angel G.; Gomez, Eduardo A.; Hashiguchi, Yoshihisa

    2016-01-01

    An epidemiological study of leishmaniasis was performed in Amazonian areas of Ecuador since little information on the prevalent Leishmania and sand fly species responsible for the transmission is available. Of 33 clinical specimens from patients with cutaneous leishmaniasis (CL), causative parasites were identified in 25 samples based on cytochrome b gene analysis. As reported previously, Leishmania (Viannia) guyanensis and L. (V.) braziliensis were among the causative agents identified. In addition, L. (V.) lainsoni, for which infection is reported in Brazil, Bolivia, Peru, Suriname, and French Guiana, was identified in patients with CL from geographically separate areas in the Ecuadorian Amazon, corroborating the notion that L. (V.) lainsoni is widely distributed in South America. Sand flies were surveyed around the area where a patient with L. (V.) lainsoni was suspected to have been infected. However, natural infection of sand flies by L. (V.) lainsoni was not detected. Further extensive vector searches are necessary to define the transmission cycle of L. (V.) lainsoni in Ecuador. PMID:27191391

  10. First Human Cases of Leishmania (Viannia lainsoni Infection and a Search for the Vector Sand Flies in Ecuador.

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    Hirotomo Kato

    2016-05-01

    Full Text Available An epidemiological study of leishmaniasis was performed in Amazonian areas of Ecuador since little information on the prevalent Leishmania and sand fly species responsible for the transmission is available. Of 33 clinical specimens from patients with cutaneous leishmaniasis (CL, causative parasites were identified in 25 samples based on cytochrome b gene analysis. As reported previously, Leishmania (Viannia guyanensis and L. (V. braziliensis were among the causative agents identified. In addition, L. (V. lainsoni, for which infection is reported in Brazil, Bolivia, Peru, Suriname, and French Guiana, was identified in patients with CL from geographically separate areas in the Ecuadorian Amazon, corroborating the notion that L. (V. lainsoni is widely distributed in South America. Sand flies were surveyed around the area where a patient with L. (V. lainsoni was suspected to have been infected. However, natural infection of sand flies by L. (V. lainsoni was not detected. Further extensive vector searches are necessary to define the transmission cycle of L. (V. lainsoni in Ecuador.

  11. Increased Transmissibility of Leishmania donovani From the Mammalian Host to Vector Sand Flies After Multiple Exposures to Sand Fly Bites.

    Science.gov (United States)

    Valverde, Joanna G; Paun, Andrea; Inbar, Ehud; Romano, Audrey; Lewis, Michael; Ghosh, Kashinath; Sacks, David

    2017-04-15

    Patients with active visceral leishmaniasis are important reservoirs in the anthroponotic transmission cycle of Leishmania donovani. The role of the blood or skin as a source of infection to sand flies remains unclear, and the possible effect of multiple exposures to fly bites on transmissibility has not been addressed. L. donovani-infected hamsters underwent xenodiagnoses with Lutzomyia longipalpis on the same or different sites on the abdomen on 2 consecutive days or by artificial feeding on the skin or blood. The transmission of L. donovani from sick hamsters to flies was surprisingly low (mean, 24% of fed flies). New flies fed on the same site acquired significantly more infections (mean, 61%; P Leishmania donovani. Using the hamster model of visceral disease, we demonstrate that prior exposure to bites of uninfected sand flies potentiates their ability to transmit infection to the vector. Published by Oxford University Press for the Infectious Diseases Society of America 2017. This work is written by (a) US Government employee(s) and is in the public domain in the US.

  12. Leishmania (infantum) chagasi in canine urinary sediment

    OpenAIRE

    Mendonça, Ivete Lopes de; Batista, Joilson Ferreira; Alves, Leucio Camara

    2015-01-01

    Canine visceral leishmaniasis (CVL) is difficult to diagnosis, mainly due to the presence of asymptomatic animals, the diversity of clinical symptoms and the difficulty in obtaining diagnostic evidence of high sensitivity and specificity. The purpose of this study was to diagnose CVL in urinary sediment of 70 dogs of different breeds, sexes and ages from the veterinary hospital of the Federal University of Piauí and Zoonosis Control Center of Teresina, Brazil. The serological tests were TR DP...

  13. Antigenicity of Leishmania-Activated C-Kinase Antigen (LACK in Human Peripheral Blood Mononuclear Cells, and Protective Effect of Prime-Boost Vaccination With pCI-neo-LACK Plus Attenuated LACK-Expressing Vaccinia Viruses in Hamsters

    Directory of Open Access Journals (Sweden)

    Laura Fernández

    2018-04-01

    Full Text Available Leishmania-activated C-kinase antigen (LACK is a highly conserved protein among Leishmania species and is considered a viable vaccine candidate for human leishmaniasis. In animal models, prime-boost vaccination with LACK-expressing plasmids plus attenuated vaccinia viruses (modified vaccinia Ankara [MVA] and mutant M65 expressing LACK, has been shown to protect against cutaneous leishmaniasis (CL. Further, LACK demonstrated to induce the production of protective cytokines in patients with active CL or cured visceral leishmaniasis, as well as in asymptomatic individuals from endemic areas. However, whether LACK is capable to trigger cytokine release by peripheral blood mononuclear cells from patients cured of CL due to Leishmania infantum (L. infantum or induce protection in L. infantum-infected hamsters [visceral leishmaniasis (VL model], has not yet been analyzed. The present work examines the ex vivo immunogenicity of LACK in cured VL and CL patients, and asymptomatic subjects from an L. infantum area. It also evaluates the vaccine potential of LACK against L. infantum infection in hamsters, in a protocol of priming with plasmid pCI-neo-LACK (DNA-LACK followed by a booster with the poxvirus vectors MVA-LACK or M65-LACK. LACK-stimulated PBMC from both asymptomatic and cured subjects responded by producing IFN-γ, TNF-α, and granzyme B (Th1-type response. Further, 78% of PBMC samples that responded to soluble Leishmania antigen showed IFN-γ secretion following stimulation with LACK. In hamsters, the protocol of DNA-LACK prime/MVA-LACK or M65-LACK virus boost vaccination significantly reduced the amount of Leishmania DNA in the liver and bone marrow, with no differences recorded between the use of MVA or M65 virus vector options. In summary, the Th1-type and cytotoxic responses elicited by LACK in PBMC from human subjects infected with L. infantum, and the parasite protective effect of prime/boost vaccination in hamsters with DNA

  14. Stage-specific adhesion of Leishmania promastigotes to sand fly midguts assessed using an improved comparative binding assay.

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    Raymond Wilson

    2010-09-01

    Full Text Available The binding of Leishmania promastigotes to the midgut epithelium is regarded as an essential part of the life-cycle in the sand fly vector, enabling the parasites to persist beyond the initial blood meal phase and establish the infection. However, the precise nature of the promastigote stage(s that mediate binding is not fully understood.To address this issue we have developed an in vitro gut binding assay in which two promastigote populations are labelled with different fluorescent dyes and compete for binding to dissected sand fly midguts. Binding of procyclic, nectomonad, leptomonad and metacyclic promastigotes of Leishmania infantum and L. mexicana to the midguts of blood-fed, female Lutzomyia longipalpis was investigated. The results show that procyclic and metacyclic promastigotes do not bind to the midgut epithelium in significant numbers, whereas nectomonad and leptomonad promastigotes both bind strongly and in similar numbers. The assay was then used to compare the binding of a range of different parasite species (L. infantum, L. mexicana, L. braziliensis, L. major, L. tropica to guts dissected from various sand flies (Lu. longipalpis, Phlebotomus papatasi, P. sergenti. The results of these comparisons were in many cases in line with expectations, the natural parasite binding most effectively to its natural vector, and no examples were found where a parasite was unable to bind to its natural vector. However, there were interesting exceptions: L. major and L. tropica being able to bind to Lu. longipalpis better than L. infantum; L. braziliensis was able to bind to P. papatasi as well as L. major; and significant binding of L. major to P. sergenti and L. tropica to P. papatasi was observed.The results demonstrate that Leishmania gut binding is strictly stage-dependent, is a property of those forms found in the middle phase of development (nectomonad and leptomonad forms, but is absent in the early blood meal and final stages (procyclic

  15. Molecular taxonomy of the two Leishmania vectors Lutzomyia umbratilis and Lutzomyia anduzei (Diptera: Psychodidae) from the Brazilian Amazon.

    Science.gov (United States)

    Scarpassa, Vera Margarete; Alencar, Ronildo Baiatone

    2013-09-11

    Lutzomyia umbratilis (a probable species complex) is the main vector of Leishmania guyanensis in the northern region of Brazil. Lutzomyia anduzei has been implicated as a secondary vector of this parasite. These species are closely related and exhibit high morphological similarity in the adult stage; therefore, they have been wrongly identified, both in the past and in the present. This shows the need for employing integrated taxonomy. With the aim of gathering information on the molecular taxonomy and evolutionary relationships of these two vectors, 118 sequences of 663 base pairs (barcode region of the mitochondrial DNA cytochrome oxidase I - COI) were generated from 72 L. umbratilis and 46 L. anduzei individuals captured, respectively, in six and five localities of the Brazilian Amazon. The efficiency of the barcode region to differentiate the L. umbratilis lineages I and II was also evaluated. The data were analyzed using the pairwise genetic distances matrix and the Neighbor-Joining (NJ) tree, both based on the Kimura Two Parameter (K2P) evolutionary model. The analyses resulted in 67 haplotypes: 32 for L. umbratilis and 35 for L. anduzei. The mean intra-specific genetic distance was 0.008 (0.002 to 0.010 for L. umbratilis; 0.008 to 0.014 for L. anduzei), whereas the mean interspecific genetic distance was 0.044 (0.041 to 0.046), supporting the barcoding gap. Between the L. umbratilis lineages I and II, it was 0.009 to 0.010. The NJ tree analysis strongly supported monophyletic clades for both L. umbratilis and L. anduzei, whereas the L. umbratilis lineages I and II formed two poorly supported monophyletic subclades. The barcode region clearly separated the two species and may therefore constitute a valuable tool in the identification of the sand fly vectors of Leishmania in endemic leishmaniasis areas. However, the barcode region had not enough power to separate the two lineages of L. umbratilis, likely reflecting incipient species that have not yet reached

  16. DSFL database: A hub of target proteins of Leishmania sp. to combat leishmaniasis

    Directory of Open Access Journals (Sweden)

    Ameer Khusro

    2017-07-01

    Full Text Available Leishmaniasis is a vector-borne chronic infectious tropical dermal disease caused by the protozoa parasite of the genus Leishmania that causes high mortality globally. Among three different clinical forms of leishmaniasis, visceral leishmaniasis (VL or kala-azar is a systemic public health disease with high morbidity and mortality in developing countries, caused by Leishmania donovani, Leishmania infantum or Leishmania chagasi. Unfortunately, there is no vaccine available till date for the treatment of leishmaniasis. On the other hand, the therapeutics approved to treat this fatal disease is expensive, toxic, and associated with serious side effects. Furthermore, the emergence of drug-resistant Leishmania parasites in most endemic countries due to the incessant utilization of existing drugs is a major concern at present. Drug Search for Leishmaniasis (DSFL is a unique database that involves 50 crystallized target proteins of varied Leishmania sp. in order to develop new drugs in future by interacting several antiparasitic compounds or molecules with specific protein through computational tools. The structure of target protein from different Leishmania sp. is available in this database. In this review, we spotlighted not only the current global status of leishmaniasis in brief but also detailed information about target proteins of various Leishmania sp. available in DSFL. DSFL has created a new expectation for mankind in order to combat leishmaniasis by targeting parasitic proteins and commence a new era to get rid of drug resistance parasites. The database will substantiate to be a worthwhile project for further development of new, non-toxic, and cost-effective antileishmanial drugs as targeted therapies using in vitro/in vivo assays.

  17. PCR evaluation of selected vector-borne pathogens in dogs with pericardial effusion.

    Science.gov (United States)

    Tabar, M-D; Movilla, R; Serrano, L; Altet, L; Francino, O; Roura, X

    2018-04-01

    To investigate evidence for selected vector-borne pathogen infections in dogs with pericardial effusion living in a Mediterranean area in which several canine vector-borne diseases are endemic. Archived EDTA blood (n=68) and pericardial fluid samples (n=58) from dogs with pericardial effusion (n=68) were included. Dogs without pericardial effusion examined for other reasons were included as controls (n=60). Pericardial effusion was classified as neoplastic in 40 dogs, idiopathic in 23 dogs and of unknown aetiology in 5 dogs. Real-time PCR was performed for Leishmania infantum, Ehrlichia/Anaplasma species, Hepatozoon canis, Babesia species, Rickettsia species and Bartonella species, and sequencing of PCR products from positive samples was used to confirm species specificity. Vector-borne pathogens were found in 18 dogs: 16 of 68 dogs with pericardial effusion (23·5%) and two of 60 control dogs (3·3%). Positive dogs demonstrated DNA of Leishmania infantum (n=7), Anaplasma platys (n=2, one dog coinfected with Leishmania infantum), Babesia canis (n=5), Babesia gibsoni (n=3) and Hepatozoon canis (n=2). Vector-borne pathogens were more commonly detected among dogs with pericardial effusion than controls (P=0·001). There was no relationship between aetiology of the pericardial effusion and evidence of vector-borne pathogens (P=0·932). Vector-borne pathogens are often detected in dogs with pericardial effusion and require further investigation, especially in dogs with idiopathic pericardial effusion. PCR can provide additional information about the potential role of vector-borne pathogens in dogs with pericardial effusion living in endemic areas. © 2018 British Small Animal Veterinary Association.

  18. Assessment of nuclear and mitochondrial genes in precise identification and analysis of genetic polymorphisms for the evaluation of Leishmania parasites.

    Science.gov (United States)

    Fotouhi-Ardakani, Reza; Dabiri, Shahriar; Ajdari, Soheila; Alimohammadian, Mohammad Hossein; AlaeeNovin, Elnaz; Taleshi, Neda; Parvizi, Parviz

    2016-12-01

    The polymorphism and genetic diversity of Leishmania genus has status under discussion depending on many items such as nuclear and/or mitochondrial genes, molecular tools, Leishmania species, geographical origin, condition of micro-environment of Leishmania parasites and isolation of Leishmania from clinical samples, reservoir host and vectors. The genetic variation of Leishmania species (L. major, L. tropica, L. tarentolae, L. mexicana, L. infantum) were analyzed and compared using mitochondrial (COII and Cyt b) and nuclear (nagt, ITS-rDNA and HSP70) genes. The role of each enzymatic (COII, Cyt b and nagt) or housekeeping (ITS-rDNA, HSP70) gene was employed for accurate identification of Leishmania parasites. After DNA extractions and amplifying of native, natural and reference strains of Leishmania parasites, polymerase chain reaction (PCR) products were sequenced and evaluation of genetic proximity and phylogenetic analysis were performed using MEGA6 and DnaSP5 software. Among the 72 sequences of the five genes, the number of polymorphic sites was significantly lower as compared to the monomorphic sites. Of the 72 sequences, 54 new haplotypes (five genes) of Leishmania species were submitted in GenBank (Access number: KU680818 - KU680871). Four genes had a remarkable number of informative sites (P=0.00), except HSP70 maybe because of its microsatellite regions. The non-synonymous (dN) variants of nagt gene were more than that of other expression genes (47.4%). The synonymous (dS)/dN ratio in three expression genes showed a significant variation between five Leishmania species (P=0.001). The highest and lowest levels of haplotype diversity were observed in L. tropica (81.35%) and L. major (28.38%) populations, respectively. Tajima's D index analyses showed that Cyt b gene in L. tropica species was significantly negative (Tajima's D=-2.2, PLeishmania parasites. Copyright © 2016 Elsevier B.V. All rights reserved.

  19. Genetic diversity and population structure in the Leishmania guyanensis vector Lutzomyia anduzei (Diptera, Psychodidae) from the Brazilian Amazon.

    Science.gov (United States)

    Scarpassa, Vera Margarete; Figueiredo, Adrya da Silva; Alencar, Ronildo Baiatone

    2015-04-01

    Lutzomyia (Nyssomyia) anduzei has been recognized as a secondary vector of Leishmania guyanensis in the Brazilian Amazon region. Since L. anduzei is anthropophilic, co-exists in areas of high leishmaniasis transmission and has been found infected with L. guyanensis, the understanding of the vector population structure and of the process responsible for it is paramount to the vector management and control efforts. In this study we analyzed 74 and 67 sequences of the COI and Cytb loci, respectively, from mitochondrial DNA, aiming to estimate the intra-population genetic variability and population structure in six L. anduzei samples from the Brazilian Amazon region. For COI, we found 58 haplotypes, low to high (FST=0.0310-0.4128) and significant (P=0.0033) genetic structure, and reduced gene flow among populations. The haplotype network yielded many reticulations that likely resulted from hypervariability in the locus. For Cytb, we observed 27 haplotypes, low to moderate (FST=0.0077-0.1954) and nonsignificant (P>0.05) genetic structure for the majority of comparisons and extensive gene flow among populations, in line with the haplotypes network data. AMOVA analysis indicated that most of the variation occurred within populations (83.41%, 90.94%); nevertheless, there were significant differences (ΦST=0.0906-0.1659; P=0.00098; P=0.00000) among them for both loci. The Mantel test showed that the genetic structure is not associated to an isolation-by-distance (IBD) model in either of both loci. These data suggest that L. anduzei is genetically very diverse. The genetic structure lacking IBD may be due to adaptation to local habitats and the low dispersal capacity of the sandflies, and both could lead to population fragmentation and geographic isolation. These findings have important implications for epidemiology, surveillance and vector control and may be a first step in understanding the evolutionary history of this species. Copyright © 2015 Elsevier B.V. All rights

  20. Leishmania major infection in a dog with cutaneous manifestations.

    Science.gov (United States)

    Baneth, Gad; Nachum-Biala, Yaarit; Shabat Simon, Maytal; Brenner, Ori; Gaier, Sarit; Rojas, Alicia; Yasur-Landau, Daniel

    2016-05-10

    Leishmania major is a main cause of cutaneous leishmaniasis in humans in an area that stretches from India through Central Asia, the Middle East, to North and West Africa. In Israel, it is a common infection of humans with rodents as the reservoir hosts and Phlebotomus papatasi as its sand fly vector. A 6 months old spayed female mixed breed dog was referred to the Hebrew University Veterinary Teaching Hospital with a large ulcerative dermal lesion on the muzzle, and lesions in the foot pads and left hind leg. Histopathology of a skin biopsy found chronic lymphohistiocytic dermatitis with the presence of Leishmania spp. amastigotes in the muzzle. Physical examination indicated that the dog was overall in a good clinical condition and the main findings were the skin lesions and enlarged prescapular lymph nodes. Complete blood count and serum biochemistry profile were within reference ranges. Serology by ELISA was positive for Leishmania spp. and PCR of the prescapular lymph node was positive by an ITS1 region PCR-high resolution melt analysis. However, the melt curve and subsequent DNA sequencing indicated that infection was caused by L. major and not L. infantum, which is the main causative agent of canine leishmaniosis in the Mediterranean region. DNA was extracted from the paraffin embedded muzzle biopsy and PCR with sequencing also indicated L. major. The dog's young age and the absence of hyperglobulinemia and anemia were not typical of L. infantum infection. The dog was treated with allopurinol and the skin lesions improved and later disappeared when the dog was re-evaluated. This is the first molecularly-confirmed case of L. major infection in a dog. Two previous reports of L. major in dogs originated from Saudi-Arabia and Egypt in 1985 and 1987 were confirmed by enzymatic biochemical techniques. Serology for L. infantum was positive probably due to the well documented serological cross-reactivity between Leishmania spp. Although dogs and wild carnivores are

  1. [Presence of infected vectors of Leishmania (V.) panamensis within dwellings in two endemic foci in the foothill of the middle Magdalena valley, western Boyacá, Colombia].

    Science.gov (United States)

    Santamaría, Erika; Ponce, Nubia; Zipa, Yaneth; Ferro, Cristina

    2006-10-01

    Case records of leishmaniasis of the years 1997 to 2003 of the department of Boyaca showed that since the year 2000 the department experienced an unusual rise in the incidence of cutaneous leishmaniasis that might correspond to an epidemic outbreak in the western region of the department. Age and gender distribution of cases supported a domestic transmission. This research was designed to identify the vectors of cutaneous leishmaniasis in the municipalities of Otanche and Pauna through their presence in dwellings and their natural infection with the same species of Leishmania isolated from patients. Sampling of sand flies was done with CDC traps in and around dwellings. Samples from patients and pooled females of the most abundant species of Lutzomyia were used to identify the parasite by PCR. Monoclonal antibody typing was also used to confirm the identification of the parasite in samples from patients. In both municipalities L. trapidoi was the most abundant anthropophilic species of Lutzomyia indoors and around dwellings. L. hartmanni and L. yuilli were also abundant in Otanche, and L. gomezi and L. panamensis in Pauna. Leishmania (V) panamensis was identified both in patients and in the sand flies: L. yuilli, L. gomezi and L. panamensis.. Our findings prove the presence of infected vectors of Leishmania panamensis within dwellings in the towns of Otanche and Pauna in Western Boyacá. Since L. trapidoi was the most abundant species, it may be considered as the principal vector of Leishmania (V.) panamensis. The evidence of transmission within human dwellings warrants vector control at least in this environment.

  2. Detection and molecular identification of leishmania RNA virus (LRV) in Iranian Leishmania species.

    Science.gov (United States)

    Hajjaran, Homa; Mahdi, Maryam; Mohebali, Mehdi; Samimi-Rad, Katayoun; Ataei-Pirkooh, Angila; Kazemi-Rad, Elham; Naddaf, Saied Reza; Raoofian, Reza

    2016-12-01

    Leishmania RNA virus (LRV) was first detected in members of the subgenus Leishmania (Viannia), and later, the virulence and metastasis of the New World species were attributed to this virus. The data on the presence of LRV in Old World species are confined to Leishmania major and a few Leishmania aethiopica isolates. The aim of this study was to survey the presence of LRV in various Iranian Leishmania species originating from patients and animal reservoir hosts. Genomic nucleic acids were extracted from 50 cultured isolates belonging to the species Leishmania major, Leishmania tropica, and Leishmania infantum. A partial sequence of the viral RNA-dependent RNA polymerase (RdRp) gene was amplified, sequenced and compared with appropriate sequences from the GenBank database. We detected the virus in two parasite specimens: an isolate of L. infantum derived from a visceral leishmaniasis (VL) patient who was unresponsive to meglumine antimoniate treatment, and an L. major isolate originating from a great gerbil, Rhombomys opimus. The Iranian LRV sequences showed the highest similarities to an Old World L. major LRV2 and were genetically distant from LRV1 isolates detected in New World Leishmania parasites. We could not attribute treatment failure in VL patient to the presence of LRV due to the limited number of specimens analyzed. Further studies with inclusion of more clinical samples are required to elucidate the potential role of LRVs in pathogenesis or treatment failure of Old World leishmaniasis.

  3. Molecular Diversity between Salivary Proteins from New World and Old World Sand Flies with Emphasis on Bichromomyia olmeca, the Sand Fly Vector of Leishmania mexicana in Mesoamerica.

    Science.gov (United States)

    Abdeladhim, Maha; V Coutinho-Abreu, Iliano; Townsend, Shannon; Pasos-Pinto, Silvia; Sanchez, Laura; Rasouli, Manoochehr; B Guimaraes-Costa, Anderson; Aslan, Hamide; Francischetti, Ivo M B; Oliveira, Fabiano; Becker, Ingeborg; Kamhawi, Shaden; Ribeiro, Jose M C; Jochim, Ryan C; Valenzuela, Jesus G

    2016-07-01

    Sand fly saliva has been shown to have proteins with potent biological activities, salivary proteins that can be used as biomarkers of vector exposure, and salivary proteins that are candidate vaccines against different forms of leishmaniasis. Sand fly salivary gland transcriptomic approach has contributed significantly to the identification and characterization of many of these salivary proteins from important Leishmania vectors; however, sand fly vectors in some regions of the world are still neglected, as Bichromomyia olmeca (formerly known as Lutzomyia olmeca olmeca), a proven vector of Leishmania mexicana in Mexico and Central America. Despite the importance of this vector in transmitting Leishmania parasite in Mesoamerica there is no information on the repertoire of B. olmeca salivary proteins and their relationship to salivary proteins from other sand fly species. A cDNA library of the salivary glands of wild-caught B. olmeca was constructed, sequenced, and analyzed. We identified transcripts encoding for novel salivary proteins from this sand fly species and performed a comparative analysis between B. olmeca salivary proteins and those from other sand fly species. With this new information we present an updated catalog of the salivary proteins specific to New World sand flies and salivary proteins common to all sand fly species. We also report in this work the anti-Factor Xa activity of Lofaxin, a salivary anticoagulant protein present in this sand fly species. This study provides information on the first transcriptome of a sand fly from Mesoamerica and adds information to the limited repertoire of salivary transcriptomes from the Americas. This comparative analysis also shows a fast degree of evolution in salivary proteins from New World sand flies as compared with Old World sand flies.

  4. Lulo cell line derived from Lutzomyia longipalpis (Diptera: Psychodidae): a novel model to assay Leishmania spp. and vector interaction.

    Science.gov (United States)

    Côrtes, Luzia Mc; Silva, Roger Mm; Pereira, Bernardo As; Guerra, Camila; Zapata, Angela C; Bello, Felio J; Finkelstein, Léa C; Madeira, Maria F; Brazil, Reginaldo P; Côrte-Real, Suzana; Alves, Carlos R

    2011-11-14

    Leishmania (Vianna) braziliensis, Leishmania (Leishmania) amazonensis and Leishmania (Leishmania) chagasi are important parasites in the scenario of leishmaniasis in Brazil. During the life cycle of these parasites, the promastigote forms adhere to the midgut epithelial microvillii of phlebotomine insects to avoid being secreted along with digestive products. Lulo cells are a potential model that will help to understand the features of this adhesion phenomenon. Here, we analyze the interaction between Leishmania spp. promastigotes and Lulo cells in vitro, specifically focusing on adhesion events occurring between three Leishmania species and this cell line. Confluent monolayers of Lulo cells were incubated with promastigotes and adhesion was assessed using both light microscopy and scanning electron microscopy. The results indicate that species from the subgenera Leishmania and Viannia have great potential to adhere to Lulo cells. The highest adherence rate was observed for L. (L.) chagasi after 24 h of incubation with Lulo cells (27.3 ± 1.8% of cells with adhered promastigotes), followed by L. (L.) amazonensis (16.0 ± 0.7%) and L. (V.) braziliensis (3.0 ± 0.7%), both after 48 h. In the ultrastructural analysis, promastigote adherence was also assessed by scanning electron microscopy, showing that, for parasites from both subgenera, adhesion occurs by both the body and the flagellum. The interaction of Lulo cells with Leishmania (L.) chagasi showed the participation of cytoplasmic projections from the former closely associating the parasites with the cells. We present evidence that Lulo cells can be useful in studies of insect-parasite interactions for Leishmania species.

  5. Design and Validation of Real-Time PCR: Quantitative Diagnosis of Common Leishmania Species in Iran.

    Science.gov (United States)

    Fekri Soofi Abadi, Maryam; Dabiri, Shahriar; Fotouhi Ardakani, Reza; Fani Malaki, Lina; Amirpoor Rostami, Sahar; Ziasistani, Mahsa; Dabiri, Donya

    2016-07-01

    Design and validation of Real-time PCR on the protected gene region ITS2 to quantify the parasite load in common leishmania (L) species. Probe and primer were designed from the ITS2 region between the rRNA genes with minimum gene variation in three common leishmania species followed by a Real-time PCR using the Taq man probe method in the form of absolute quantification. A series of different concentrations of leishmania were analyzed. After the purified PCR product was successfully placed in a PTG19-T plasmid vector, specialized ITS2 region was cloned in this plasmid. In the last phase, the cloned gene was transferred to the Ecoli.Top10F bacteria. The standard plasmid was provided in 10(7) to 10(1) copies/rxn concentrations. The specification and clinical sensitivity of the data was analyzed using inter and intra scales. The probe and primer were designed using three species, including L. infantum, L. major, and L.tropica. Seven concentrations of purified parasite in culture media showed that the selected region for quantifying the parasite is suitable. Clinical and analytical specificity and sensitivity were both 100%, respectively. The Taq man method for the ITS2 region in leishmania is one the most sensitive diagnostic test for identifying the parasite load and is suggested as a tool for fast identification and quantification of species.

  6. Phylogenetic position of Leishmania isolates from Khyber Pakhtunkhwa province of Pakistan.

    Science.gov (United States)

    Khan, Nazma Habib; Messenger, Louisa A; Wahid, Sobia; Sutherland, Colin J

    2016-08-01

    Several species of the genus Leishmania are causative agents of cutaneous leishmaniasis in Pakistan. This study aimed to determine phylogenetic placement of Leishmania species causing cutaneous leishmaniasis in Khyber Pakhtunkhwa province, Pakistan (34 Leishmania tropica, 3 Leishmania infantum), in-relation to species from other geographical areas using gene sequences encoding cytochrome b (cytb) and internal transcribed spacer 2 (its2). Based on cytochrome b sequence analysis, L. tropica strains from Pakistan and other geographical regions were differentiated into two genotype groups, A and B. Within the province, five distinct L. tropica genotypes were recognized; two in group A, three in group B. Two L. infantum isolates from the province were closely associated with both Afro-Eurasian and American species of the Leishmania donovani complex, including Leishmania chagasi, L. infantum and L. donovani from Sudan and Ethiopia; while a third L. infantum isolate could not be differentiated from visceralizing Kenyan and Indian L. donovani. We observed apposite phylogenetic placement of CL-causing L. tropica and L. infantum from Khyber Pakhtunkhwa. Affinities ascribed to Leishmania spp. From the region are valuable in tracing potential importation of leishmaniasis. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

  7. Identification of six New World Leishmania species through the implementation of a High-Resolution Melting (HRM) genotyping assay.

    Science.gov (United States)

    Hernández, Carolina; Alvarez, Catalina; González, Camila; Ayala, Martha Stella; León, Cielo Maritza; Ramírez, Juan David

    2014-11-14

    Leishmaniases are tropical zoonotic diseases, caused by parasites from the genus Leishmania. New World (NW) species are related to sylvatic cycles although urbanization processes have been reported in some South American Countries such as Colombia. This eco-epidemiological complexity imposes a challenge to the detection of circulating parasite species, not only related to human cases but also infecting vectors and reservoirs. Currently, no harmonized methods have been deployed to discriminate the NW Leishmania species. Herein, we conducted a systematic and mechanistic High-Resolution Melting (HRM) assay targeted to HSP70 and ITS1. Specific primers were designed that coupled with a HRM analyses permitted to discriminate six NW Leishmania species. In order to validate the herein described algorithm, we included 35 natural isolates obtained from human cases, insect vectors and mammals. Our genotyping assay allowed the correct assignment of the six NW Leishmania species (L. mexicana, L. infantum (chagasi), L. amazonensis, L. panamensis, L. guyanensis and L. braziliensis) based on reference strains. When the algorithm was applied to a set of well-characterized strains by means of PCR-RFLP, MLEE and monoclonal antibodies (MA) we observed a tailored concordance between the HRM and PCR-RFLP/MLEE/MA (KI = 1.0). Additionally, we tested the limit of detection for the HRM method showing that this is able to detect at least 10 equivalent-parasites per mL. This is a rapid and reliable method to conduct molecular epidemiology and host-parasite association studies in endemic areas.

  8. Diversity patterns, Leishmania DNA detection, and bloodmeal identification of Phlebotominae sand flies in villages in northern Colombia.

    Science.gov (United States)

    González, Camila; León, Cielo; Paz, Andrea; López, Marla; Molina, Gisell; Toro, Diana; Ortiz, Mario; Cordovez, Juan Manuel; Atencia, María Claudia; Aguilera, Germán; Tovar, Catalina

    2018-01-01

    Leishmaniases are neglected tropical diseases exhibiting complex transmission cycles due to the number of parasite species circulating, sand fly species acting as vectors and infected mammals, including humans, which are defined in the New World as accidental hosts. However, current transmission scenarios are changing, and the disease is no longer exclusively related to forested areas but urban transmission foci occur, involving some species of domestic animals as suspected reservoirs. The aim of this study was to determine the transmission cycles in urban environments by evaluating sand fly diversity, detection of Leishmania DNA, and bloodmeal sources through intra and peridomestic collections. The study was carried out in Colombia, in 13 municipalities of Cordoba department, implementing a methodology that could be further used for the evaluation of vector-borne diseases in villages or towns. Our sampling design included 24 houses randomly selected in each of 15 villages distributed in 13 municipalities, which were sampled in two seasons in 2015 and 2016. Sand flies were collected using CDC light traps placed in intra and peridomestic habitats. In addition to the morphological identification, molecular identification through DNA barcodes was also performed. A total of 19,743 sand flies were collected and 13,848 of them (10,268 females and 3,580 males) were used in molecular procedures. Circulation of two known parasite species-Leishmania infantum and Leishmania panamensis was confirmed. Blood source analyses showed that sand flies fed on humans, particularly in the case of the known L. infantum vector, P. evansi; further analyses are advised to evaluate the reservoirs involved in parasite transmission. Our sampling design allowed us to evaluate potential transmission cycles on a department scale, by defining suspected vector species, parasite species present in different municipalities and feeding habits.

  9. Habitats of the sandfly vectors of Leishmania tropica and L. major in a mixed focus of cutaneous leishmaniasis in southeast Tunisia.

    Science.gov (United States)

    Tabbabi, Ahmed; Ghrab, Jamila; Aoun, Karim; Ready, Paul Donald; Bouratbine, Aïda

    2011-08-01

    From 2009 to 2010, 3129 sandflies were caught in CDC light traps placed in various habitats in Ghomrassen, Tataouine governorate, southeast Tunisia, a mixed focus of human cutaneous leishmaniasis caused by Leishmania tropica and Leishmania major. Species diversity was quantified in anthropogenic, semi-anthropogenic and semi-natural locations. Sandflies were identified according to morphological characters and also by the comparative sequence analysis of a fragment of the mitochondrial cytochrome b gene to distinguish between two putative local vectors of L. tropica, namely Phlebotomus chabaudi and Phlebotomus riouxi. The lowest sandfly diversities were found in L. major sites, where the incriminated vector P. papatasi predominated in the burrows of the rodent reservoir hosts (Meriones) as well as inside and outside houses of human cases. In L. tropica sites, the incriminated peri-domestic vector Phlebotomus sergenti was the most abundant species inside houses, whereas P. riouxi or P. chabaudi was the dominant species in the semi-natural rocky habitats favoured by the putative rodent reservoir, Ctenodactylus gundi. All specimens of P. chabaudi identified molecularly had the diagnostic cytochrome b characters of P. riouxi, indicating either that the latter represents only a geographical variant of P. chabaudi or that these two species may sometimes hybridize. Copyright © 2011 Elsevier B.V. All rights reserved.

  10. Molecular characterization of sandflies and Leishmania detection in main vector of zoonotic cutaneous leishmaniasis in Abarkouh district of Yazd province, Iran.

    Science.gov (United States)

    Jafari, R; Najafzadeh, N; Sedaghat, M M; Parvizi, P

    2013-10-01

    To assess molecular characterization, distribution, seasonal activities of sandfly species and Leishmania parasites infecting them for this zoonotic cutaneous leishmaniasis focus. The collections were carried out in 2009-2011 using CDC traps, Sticky Papers and manual aspirator in and around the villages in Abarkouh district. Individual sandflies were characterized by PCR amplification and sequencing of fragments of their mitochondrial cytochrome b gene. Leishmania parasite infections within sandflies were performed by targeting Cyt b, ITS-rDNA, k-DNA and microsatellite genes. The PCR assays detected only Leishmania major (L. major). All infections (30) were found in the abundant and widespread vector Phlebotomus papatasi (P. papatasi). Small numbers of other sandfly species were also screened for infections, but none was found. Sergentomyia sintoni and P. papatasi were the predominant members in all locations of this district and in all habitats throughout the trapping season. Only five other sandfly species were found, namely Phlebotomus ansari, Phlebotomus caucasicus, Phlebotomus sergenti, Sergentomyia dentata and Sergentomyia merviney. In the current survey, the only infections detected are of L. major in females of P. papatasi (30 out of 190). The rates of infection of P. papatasi by L. major are not significantly different in compare with other locations in Iran with no diversity of parasite strains. Zoonotic cutaneous leishmaniasis may have emerged only recently in Abarkouh district, and the reason may well be the instability of the transmission cycles there. Copyright © 2013 Hainan Medical College. Published by Elsevier B.V. All rights reserved.

  11. Surveillance for antibodies to Leishmania spp. in dogs from Sri Lanka and India

    Science.gov (United States)

    The global distribution of leishmaniasis is rapidly expanding into new geographic regions. Dogs are the primary reservoir hosts for human visceral leishmaniasis (VL) caused by infection with Leishmania infantum. Natural infections with other Leishmania species can occur in dogs, but their role as re...

  12. Serological and molecular survey of Leishmania infection in dogs from Luanda, Angola

    NARCIS (Netherlands)

    Vilhena, Hugo; Granada, Sara; Oliveira, Ana Cristina; Schallig, Henk D. F. H.; Nachum-Biala, Yaarit; Cardoso, Luís; Baneth, Gad

    2014-01-01

    Canine leishmaniosis (CanL) due to Leishmania infantum is a global zoonosis endemic in more than 70 countries in Europe, North Africa, Asia and America; however, data on this infection is scarce from southern Africa. The aim of this study was to survey dogs in Luanda, Angola, for Leishmania

  13. Development of a dipstick assay for detection of Leishmania-specific canine antibodies

    NARCIS (Netherlands)

    Schallig, Henk D. F. H.; Cardoso, Luís; Hommers, Marieke; Kroon, Nel; Belling, Guus; Rodrigues, Manuela; Semião-Santos, Saul J.; Vetter, Hans

    2004-01-01

    A dipstick assay, based on Leishmania infantum antigen, for the rapid detection of Leishmania-specific antibodies in canine serum samples was developed and evaluated. After determination of optimal dipstick test conditions, test performance was compared with two existing serological tests, i.e., the

  14. Canine vector-borne diseases in Brazil

    Science.gov (United States)

    Dantas-Torres, Filipe

    2008-01-01

    Canine vector-borne diseases (CVBDs) are highly prevalent in Brazil and represent a challenge to veterinarians and public health workers, since some diseases are of great zoonotic potential. Dogs are affected by many protozoa (e.g., Babesia vogeli, Leishmania infantum, and Trypanosoma cruzi), bacteria (e.g., Anaplasma platys and Ehrlichia canis), and helminths (e.g., Dirofilaria immitis and Dipylidium caninum) that are transmitted by a diverse range of arthropod vectors, including ticks, fleas, lice, triatomines, mosquitoes, tabanids, and phlebotomine sand flies. This article focuses on several aspects (etiology, transmission, distribution, prevalence, risk factors, diagnosis, control, prevention, and public health significance) of CVBDs in Brazil and discusses research gaps to be addressed in future studies. PMID:18691408

  15. Canine vector-borne diseases in Brazil

    Directory of Open Access Journals (Sweden)

    Dantas-Torres Filipe

    2008-08-01

    Full Text Available Abstract Canine vector-borne diseases (CVBDs are highly prevalent in Brazil and represent a challenge to veterinarians and public health workers, since some diseases are of great zoonotic potential. Dogs are affected by many protozoa (e.g., Babesia vogeli, Leishmania infantum, and Trypanosoma cruzi, bacteria (e.g., Anaplasma platys and Ehrlichia canis, and helminths (e.g., Dirofilaria immitis and Dipylidium caninum that are transmitted by a diverse range of arthropod vectors, including ticks, fleas, lice, triatomines, mosquitoes, tabanids, and phlebotomine sand flies. This article focuses on several aspects (etiology, transmission, distribution, prevalence, risk factors, diagnosis, control, prevention, and public health significance of CVBDs in Brazil and discusses research gaps to be addressed in future studies.

  16. Molecular detection and identification of Leishmania spp. in naturally infected Phlebotomus tobbi and Sergentomyia dentata in a focus of human and canine leishmaniasis in western Turkey.

    Science.gov (United States)

    Özbel, Yusuf; Karakuş, Mehmet; Arserim, Suha K; Kalkan, Şaban Orçun; Töz, Seray

    2016-03-01

    Human visceral leishmaniasis (VL) is reported from 38 provinces of Turkey and dogs are accepted as main reservoir hosts. Kuşadası town, belonging to Aydın province and located in western part of Turkey, is endemic for human and canine visceral leishmaniasis caused by Leishmania infantum MON1 and MON98. In this study, phlebotomine survey was conducted to determine the vector sand fly species and to identify sand fly blood meal sources. In August and September 2012, 1027 sand fly specimens were caught using CDC light traps. Eight Phlebotomus and two Sergentomyia species with the dominancy of Phlebotomus tobbi (61.34%) were detected. A total of 622 female sand flies (571 Phlebotomus; 51 Sergentomyia) were checked for Leishmania infection by direct dissection of the midgut. The half of the midgut content was inoculated into NNN culture for isolation of the parasite. Leishmania species-specific ITS1 real time PCR, conventional PCR assays of ITS1 and hsp70 genes and subsequent sequencing were performed from extracted DNAs. A region of cytochrome b (cyt-b) gene of vertebrates based PCR was used to determine the source of blood meal of sand flies. In microscopical examinations, two female specimens (0.32%) were found naturally infected with high number and different stages of promastigotes. No growth was observed in NNN culture but Leishmania DNA was obtained from both specimens. First positive specimen was identified as P. tobbi and L. infantum DNA was detected. Second specimen was Sergentomyia dentata, but Leishmania DNA could not be identified on species level. A total of 16 blood-fed female P. tobbi specimens were used for blood meal analysis and eight, three and one specimens were positive for human, dog and mouse, respectively. This is the first detection of Leishmania promastigotes using microscopical examination in P. tobbi and S. dentata in human and canine visceral leishmaniasis endemic area in western part of Turkey. Our results indicate that, (i) P. tobbi is

  17. Efecto del tipo de sangre en la supervivencia y fecundidad del flebotomino Lutzomyia ovallesi Ortiz (Diptera: Psychodidae vector de Leishmania

    Directory of Open Access Journals (Sweden)

    Pedro Noguera

    2006-10-01

    Full Text Available Introducción. El potencial reproductivo de los flebotominos depende de varios factores, unode los cuales es el tipo de hospedador disponible como fuente sanguinea, este es importanteen determinar su capacidad de servir como vectores. Objetivo. Se estudia el efecto de la fuente de alimentación sanguínea sobre varios parámetrosbiológicos de L. ovallesi en condiciones de laboratorio. Materiales y métodos. Se utilizaron hembras de dos días de edad de L. ovallesi de colonia,alimentadas artificialmente a repleción usando membrana de pollo, con sangre de sietehospedadores vertebrados, caballo, perro, vaca, gallina, chivo, cochino y humano. Se determinóel tiempo de vida, tiempo de oviposición, tiempo de digestión sanguínea, número de huevospuestos, número de huevos retenidos y número de huevos totales. Resultados. Los resultados muestran la influencia de la fuente sanguínea sobre diferentesparámetros biológicos de L. ovallesi estudiados. Los resultados demuestran que con la sangrede gallina se obtienen mayor tiempo de oviposición (5,88 días, digestión más rápida (3,34días, mayor número de huevos retenidos (10,20 huevos por hembra y mayor fecundidad(30,80 huevos por hembra en comparación con los otros tipos de sangre. La sangre más satisfactoria fue la de gallina seguida, en orden descendente, por las de chivo, vaca, cochino,humano, perro y caballo. Conclusión. Los datos muestran que la sangre de gallina es la mejor fuente sanguínea entérminos bio-ecológicos para L. ovallesi, y la sangre de caballo la menos adecuada. Losresultados contribuyen al entendimiento de los mecanismos que influyen en las condicionesde cría en el laboratorio del flebotomino L. ovallesi y también de cómo ciertos factores de ladieta en los adultos afectan el potencial biológico y que podrían tener importante consecuenciasen la transmisión de Leishmania.

  18. Canine leishmaniosis caused by Leishmania major and Leishmania tropica: comparative findings and serology.

    Science.gov (United States)

    Baneth, Gad; Yasur-Landau, Daniel; Gilad, Matan; Nachum-Biala, Yaarit

    2017-03-13

    Infection and clinical disease associated with Leishmania major and Leishmania tropica, two common agents of human cutaneous leishmaniosis, have rarely been reported in dogs. This study describes dogs infected with these Leishmania spp. prevalent in the Middle East and North Africa, and compares the serological response of dogs infected with Leishmania infantum, L. major or L. tropica to whole promastigote antigen enzyme-linked immunosorbent assay (ELISA) of each species and to rK39 dipstick. Leishmania major infection in a 5-month-old male dog was associated with alopecic and ulcerative periocular and limb skin lesions which responded to allopurinol treatment. Infection was detected by skin and blood polymerase chain reaction (PCR) and confirmed by DNA sequencing but the dog was seronegative. Leishmania tropica infection was detected in a 3-month-old female dog co-infected with Babesia vogeli and Anaplasma platys and with no skin lesions. PCR and DNA sequencing of the blood and parasite culture were positive for L. tropica. Sera from 11 dogs infected with L. infantum, L. major or L. tropica were reactive with all three Leishmania spp. antigens except for sera from a dog with L. major infection. No significant differences were found between reactivity of dog sera to the antigen of the infecting species, or to the other Leishmania spp. antigens. Sera from dogs infected with L. infantum and L. tropica were positive with the rK39 antigen kit, while dogs with L. major infection were seronegative. Skin lesions in L. major infected dogs from this study and previous reports (n = 2) were ulcerative and located on the muzzle, feet and foot pads and not associated with generalized lymphadenomegaly and splenomegaly. In previous L. tropica infections, skin lesions were proliferative mucocutaneous in young dogs (n = 2), or associated with widespread dermatitis, lymphadenomegaly and splenomegaly in older dogs with similarity to L. infantum infection (n = 2). This

  19. The sandfly Lutzomyia longipalpis LL5 embryonic cell line has active Toll and Imd pathways and shows immune responses to bacteria, yeast and Leishmania.

    Science.gov (United States)

    Tinoco-Nunes, Bruno; Telleria, Erich Loza; da Silva-Neves, Monique; Marques, Christiane; Azevedo-Brito, Daisy Aline; Pitaluga, André Nóbrega; Traub-Csekö, Yara Maria

    2016-04-20

    Lutzomyia longipalpis is the main vector of visceral leishmaniasis in Latin America. Sandfly immune responses are poorly understood. In previous work we showed that these vector insects respond to bacterial infections by modulating a defensin gene expression and activate the Imd pathway in response to Leishmania infection. Aspects of innate immune pathways in insects (including mosquito vectors of human diseases) have been revealed by studying insect cell lines, and we have previously demonstrated antiviral responses in the L. longipalpis embryonic cell line LL5. The expression patterns of antimicrobial peptides (AMPs) and transcription factors were evaluated after silencing the repressors of the Toll pathway (cactus) and Imd pathway (caspar). AMPs and transcription factor expression patterns were also evaluated after challenge with heat-killed bacteria, heat-killed yeast, or live Leishmania. These studies showed that LL5 cells have active Toll and Imd pathways, since they displayed an increased expression of AMP genes following silencing of the repressors cactus and caspar, respectively. These pathways were also activated by challenges with bacteria, yeast and Leishmania infantum chagasi. We demonstrated that L. longipalpis LL5 embryonic cells respond to immune stimuli and are therefore a good model to study the immunological pathways of this important vector of leishmaniasis.

  20. Study on coinfecting vector-borne pathogens in dogs and ticks in Rio Grande do Norte, Brazil

    Directory of Open Access Journals (Sweden)

    Luiz Ricardo Gonçalves

    Full Text Available Since dogs presenting several vector borne diseases can show none or nonspecific clinical signs depending on the phase of infection, the assessment of the particular agents involved is mandatory. The present study aimed to investigate the presence of Babesia spp., Ehrlichia spp., Anaplasma spp., Hepatozoon spp. and Leishmania spp. in blood samples and ticks, collected from two dogs from Rio Grande do Norte showing suggestive tick-borne disease by using molecular techniques. DNA of E. canis, H. canis and L. infantum were detected in blood samples and R. sanguineus ticks collected from dogs. Among all samples analyzed, two showed the presence of multiple infections with E. canis, H. canis and L. infantum chagasi. Here we highlighted the need for molecular differential diagnosis in dogs showing nonspecific clinical signs.

  1. Predicting the risk of an endemic focus of Leishmania tropica becoming established in South-Western Europe through the presence of its main vector, Phlebotomus sergenti Parrot, 1917.

    Science.gov (United States)

    Barón, S D; Morillas-Márquez, F; Morales-Yuste, M; Díaz-Sáez, V; Gállego, M; Molina, R; Martín-Sánchez, J

    2013-09-01

    The aim of the study was the construction of risk maps for exposure to Phlebotomus sergenti, the main vector of Leishmania tropica, with a view to identifying hot spots for the potential establishment of this parasite in the southwest of Europe. Data were collected on the presence/absence of this vector and the ecological and climatic characteristics of 662 sampling sites located in the southeast, centre and northeast of the Iberian Peninsula (south-western Europe). The environmental factors associated with the distribution of P. sergenti were determined. The best predictors for the presence of this dipteran were ‘altitude’, ‘land use’, ‘land surface temperature’, ‘aspect’, ‘adjacent land cover’, ‘absence of vegetation in wall’ and the ‘absence of PVC pipes in the drainage holes of retaining walls’. Risk maps for exposure to the vector were drawn up based on these variables. The validation of the predictive risk model confirmed its usefulness in the detection of areas with a high risk of P. sergenti being present. These locations represent potential hot spots for an autochthonous focus of L. tropica becoming established. The risk maps produced for P. sergenti presence revealed several areas in the centre and south of the Iberian Peninsula to be the most prone to this process, which would make it possible for the disease to enter south-western Europe.

  2. Genetic structure and divergence in populations of Lutzomyia cruciata, a phlebotomine sand fly (Diptera: Psychodidae) vector of Leishmania mexicana in southeastern Mexico.

    Science.gov (United States)

    Pech-May, Angélica; Marina, Carlos F; Vázquez-Domínguez, Ella; Berzunza-Cruz, Miriam; Rebollar-Téllez, Eduardo A; Narváez-Zapata, José A; Moo-Llanes, David; Ibáñez-Bernal, Sergio; Ramsey, Janine M; Becker, Ingeborg

    2013-06-01

    The low dispersal capacity of sand flies could lead to population isolation due to geographic barriers, climate variation, or to population fragmentation associated with specific local habitats due to landscape modification. The phlebotomine sand fly Lutzomyia cruciata has a wide distribution throughout Mexico and is a vector of Leishmania mexicana in the southeast. The aim of this study was to evaluate the genetic diversity, structure, and divergence within and among populations of Lu. cruciata in the state of Chiapas, and to infer the intra-specific phylogeny using the 3' end of the mitochondrial cytochrome b gene. We analyzed 62 sequences from four Lu. cruciata populations and found 26 haplotypes, high genetic differentiation and restricted gene flow among populations (Fst=0.416, Nm=0.701, p<0.001). The highest diversity values were recorded in populations from Loma Bonita and Guadalupe Miramar. Three lineages (100% bootstrap and 7% overall divergence) were identified using a maximum likelihood phylogenetic analysis which showed high genetic divergence (17.2-22.7%). A minimum spanning haplotype network also supported separation into three lineages. Genetic structure and divergence within and among Lu. cruciata populations are hence affected by geographic heterogeneity and evolutionary background. Data obtained in the present study suggest that Lu. cruciata in the state of Chiapas consists of at least three lineages. Such findings may have implications for vector capacity and hence for vector control strategies. Copyright © 2013 Elsevier B.V. All rights reserved.

  3. First human cases of Leishmania (Viannia) naiffi infection in Ecuador and identification of its suspected vector species.

    Science.gov (United States)

    Kato, Hirotomo; Calvopiña, Manuel; Criollo, Hipatia; Hashiguchi, Yoshihisa

    2013-12-01

    Epidemiological surveillance of leishmaniasis was conducted in a northern Amazonian region of Ecuador, in which cutaneous leishmaniasis cases were recently reported. Sand flies were captured in the military training camp, and the natural infection of sand flies by Leishmania species was examined. Out of 334 female sand flies dissected, the natural infection by flagellates was microscopically detected in 3.9% of Lutzomyia yuilli yuilli and 3.7% of Lutzomyia tortura, and the parasite species were identified as Endotrypanum and Leishmania (Viannia) naiffi, respectively. After the sand fly surveillance, specimens from cutaneous leishmaniasis (CL) patients considered to have acquired the infection in the training camp area were obtained, and the infected parasite species were identified as L. (V.) naiffi. The present study reported first cases of CL caused by L. (V.) naiffi infection in Ecuador. In addition, a high ratio of infection of Lu. tortura by L. (V.) naiffi in the same area strongly suggested that Lu. tortura is responsible for the transmission of L. (V.) naiffi in this area. Copyright © 2013 Elsevier B.V. All rights reserved.

  4. The salivary glands of two sand fly vectors of Leishmania: Lutzomyia migonei (França) and Lutzomyia ovallesi (Ortiz)(Diptera: Psychodidae).

    Science.gov (United States)

    Nieves, Elsa; Buelvas, Neudo; Rondón, Maritza; González, Néstor

    2010-01-01

    Leishmaniasis is a vector-borne disease transmitted by the intradermal inoculation of Leishmania (Kinetoplastida: Trypanosomatidae) promastigotes together with saliva during the bite of an infected sand fly. The salivary glands were compared from two vector species, Lutzomyia ovallesi (Ortiz,1952) and Lutzomyia migonei (França,1920) (Diptera: Psychodidae). Protein profiles by SDS PAGE of salivary glands were compared among species as well as their development at several times post feeding. First, mice were immunized to salivary proteins by exposure to biting by L. ovallesi and of L. migonei. Antibodies in these mice against salivary gland-specific proteins were evaluated by immunoblotting. No apparent change was revealed in the kinetic expression of salivary proteins induced by the different physiological states post feeding. Qualitative and quantitative variations were detected in16-18 polypeptides with molecular weights ranging from 6 to 180 kDa. Species-specific proteins were demonstrated for L. migonei and L. ovallesi. In addition, antibodies against salivary gland specific proteins were found in mice immunized by the saliva of both species. Basic information was obtained concerning the nature of salivary gland proteins of L. migonei and L. ovallesi. This information helps to elucidate the role of salivary proteins and their potential as effective tools in screening risk factors in human and other vertebrate hosts.

  5. Direct detection of Leishmania from clinical samples.

    Science.gov (United States)

    Waitumbi, John N; Bast, Joshua; Nyakoe, Nancy; Magiri, Charles; Quintana, Miguel; Takhampunya, Ratree; Schuster, Anthony L; Van de Wyngaerde, Marshall T; McAvin, James C; Coleman, Russell E

    2017-01-01

    The ability to rapidly and accurately diagnose leishmaniasis is a military priority. Testing was conducted to evaluate diagnostic sensitivity and specificity of field-expedient Leishmania genus and visceral Leishmania specific dual-fluorogenic, hydrolysis probe (TaqMan), polymerase chain reaction assays previously established for use in vector surveillance. Blood samples of patients with confirmed visceral leishmaniasis and controls without the disease from Baringo District, Kenya, were tested. Leishmania genus assay sensitivity was 100% (14/14) and specificity was 84% (16/19). Visceral Leishmania assay sensitivity was 93% (13/14) and specificity 80% (4/5). Cutaneous leishmaniasis (CL) skin scrapes of patients from Honduras were also evaluated. Leishmania genus assay sensitivity was 100% (10/10). Visceral Leishmania assay specificity was 100% (10/10) from cutaneous leishmaniasis samples; no fluorescence above background was reported. These results show promise in a rapid, sensitive, and specific method for Leishmania direct detection from clinical samples.

  6. Towards multilocus sequence typing of the Leishmania donovani complex: Resolving genotypes and haplotypes for five polymorphic metabolic enzymes (ASAT, GPI, NH1, NH2, PGD)

    Czech Academy of Sciences Publication Activity Database

    Mauricio, I. L.; Yeo, M.; Baghaei, M.; Doto, D.; Pratlong, F.; Zemanová, Eva; Dedet, J.-P.; Lukeš, Julius; Miles, M. A.

    2006-01-01

    Roč. 36, č. 7 (2006), s. 757-769 ISSN 0020-7519 Grant - others:European Comission(EU) QLK2-CT-2001-01810 Institutional research plan: CEZ:AV0Z60220518 Keywords : Leishmania donovani * Leishmania infantum * multilocus sequence typing Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.337, year: 2006

  7. Differential Activation of Human Keratinocytes by Leishmania Species Causing Localized or Disseminated Disease.

    Science.gov (United States)

    Scorza, Breanna M; Wacker, Mark A; Messingham, Kelly; Kim, Peter; Klingelhutz, Aloysius; Fairley, Janet; Wilson, Mary E

    2017-10-01

    All Leishmania species parasites are introduced into mammalian skin through a sand fly bite, but different species cause distinct clinical outcomes. Mouse studies suggest that early responses are critical determinants of subsequent adaptive immunity in leishmaniasis, yet few studies address the role of keratinocytes, the most abundant cell in the epidermis. We hypothesized that Leishmania infection causes keratinocytes to produce immunomodulatory factors that influence the outcome of infection. Incubation of primary or immortalized human keratinocytes with Leishmania infantum or Leishmania major, which cause visceral or cutaneous leishmaniasis, respectively, elicited dramatically different responses. Keratinocytes incubated with L. infantum significantly increased expression of proinflammatory genes for IL-6, IL-8, tumor necrosis factor, and IL-1B, whereas keratinocytes exposed to several L. major isolates did not. Furthermore, keratinocyte-monocyte co-incubation studies across a 4 µM semipermeable membrane suggested that L. infantum-exposed keratinocytes release soluble factors that enhance monocyte control of intracellular L. infantum replication (P Leishmania species that may affect the course of disease. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  8. Caracteres moleculares para la determinación taxonómica de tres especies de Lutzomyia (Diptera: Psychodidae, vectores potenciales de Leishmania presentes en el valle de Aburrá, Colombia Molecular characters for the taxonomic determination of three species of Lutzomyia (Diptera: Psychodidae, potential Leishmania vectors found in the Aburrá valley, Colombia

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    Alveiro Pérez-Doria

    2008-12-01

    Full Text Available En Colombia están registradas 143 especies de Lutzomyia França, pero menos del 7% de éstas se encuentran incriminadas como vectores de Leishmania spp. Debido a la alta semejanza morfológica de algunas especies vectoras con otras no vectoras, se necesitan caracteres taxonómicos alternativos para identificar correctamente los flebotomíneos de cada zona geográfica del país. Con este objetivo, en el presente trabajo se secuenció el extremo 3' del gen mitocondrial que codifica para la proteína citocromo b en tres vectores potenciales de Leishmania presentes en el valle de Aburrá, Colombia, Lutzomyia hartmanni (Fairchild y Hertig, L. columbiana (Ristorcelli y Van Ty y L. tihuiliensis Le Pont, Torrez-Espejo y Dujardin. A partir del alineamiento múltiple de nucleótidos se determinaron los sitios polimórficos, las distancias genéticas pareadas netas (p y la entropía. Las secuencias de nucleótidos fueron trasladadas a aminoácidos para estimar el número de sustituciones sinónimas y no sinónimas. En el alineamiento múltiple de 321 nucleótidos del gen citocromo b de L. columbiana, L. hartmanni y L. tihuiliensis se detectaron 83 sustituciones. En la secuencia parcial de la proteína se encontraron 18 reemplazos de aminoácidos. Las distancias genéticas interespecíficas fluctuaron en un rango mínimo de 0,137 entre L. tihuiliensis y L. columbiana, y un máximo de 0,215 entre L. columbiana y L. hartmanni. Los polimorfismos detectados en la secuencia de nucleótidos del gen y de aminoácidos de la proteína constituyen caracteres moleculares potencialmente útiles para la determinación taxonómica de estas especies de flebotomíneos.To date, 143 species of Lutzomyia França are recorded in Colombia, but less than 7% is incriminated in the transmission of Leishmania spp. Alternative taxonomic characters are necessary to correctly identify the particular sand fly fauna in each Colombian region, and the separation of morphologically

  9. Prevalence and Geographic Distribution of Vector-Borne Pathogens in Apparently Healthy Dogs in Croatia.

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    Mrljak, Vladimir; Kuleš, Josipa; Mihaljević, Željko; Torti, Marin; Gotić, Jelena; Crnogaj, Martina; Živičnjak, Tatjana; Mayer, Iva; Šmit, Iva; Bhide, Mangesh; Barić Rafaj, Renata

    2017-06-01

    Vector-borne pathogens (VBPs) are a group of globally extended and quickly spreading pathogens that are transmitted by various arthropod vectors. The aim of the present study was to investigate the seroprevalence against Babesia canis, Anaplasma phagocytophilum, Borrelia burgdorferi sensu lato, Leishmania infantum, Dirofilaria immitis, and Ehrlichia canis in dogs in Croatia. We investigated 435 randomly selected apparently healthy dogs in 13 different locations of Croatia for antibodies to B. canis by indirect immunofluorescence using a commercial IFA IgG Antibody Kit. All samples were also tested for qualitative detection of D. immitis antigen and for antibodies to A. phagocytophilum, B. burgdorferi sensu lato, L. infantum, and E. canis with two point-of-care assays. Overall, 112 dogs (25.74%, 95% confidence interval [CI] 21.70-30.12) were serologically positive for one or more of the pathogens. B. canis was the most prevalent pathogen (20.00%, 95% CI 16.34-24.07), followed by A. phagocytophilum (6.21%, 95% CI 4.12-8.90), L. infantum, (1.38%, 95% CI 0.51-2.97), and B. burgdorferi sensu lato (0.69%, 95% CI 0.01-2.00). The lowest seroprevalence was for D. immitis and E. canis (0.46%, 95% CI 0.01-1.65). Coinfection was determined in 12 dogs (2.76%, 95% CI 1.43-4.77), of which 10 were positive to two pathogens (7 with B. canis and A. phagocytophilum and 1 B. canis with B. burgdorferi sensu lato or L. infantum or E. canis). One dog was positive to three pathogens and another dog to four pathogens. Seroprevalence for babesia was age, breed, and lifestyle/use dependent. Purebred dogs had almost half the chance of developing disease than crossbred (OR = 0.58, p Croatia. Some of these VBPs are zoonotic and represent a potential risk to public health.

  10. Genetic diversity of the mitochondrial cytochrome b gene in Lutzomyia spp., with special reference to Lutzomyia peruensis, a main vector of Leishmania (Viannia) peruviana in the Peruvian Andes.

    Science.gov (United States)

    Yamamoto, Kento; Cáceres, Abraham G; Gomez, Eduardo A; Mimori, Tatsuyuki; Iwata, Hiroyuki; Korenaga, Masataka; Sakurai, Tatsuya; Katakura, Ken; Hashiguchi, Yoshihisa; Kato, Hirotomo

    2013-05-01

    The genetic divergence caused by genetic drift and/or selection is suggested to affect the vectorial capacity and insecticide susceptibility of sand flies, as well as other arthropods. In the present study, cytochrome b (cyt b) gene sequences were determined in 13 species circulating in Peru to establish a basis for analysis of the genetic structure, and the intraspecific genetic diversity was assessed in the Lutzomyia (Lu.) peruensis, a main vector species of Leishmania (Viannia) peruviana in Peruvian Andes. Analysis of intraspecific genetic diversity in the cyt b gene sequences from 36 Lu. peruensis identified 3 highly polymorphic sites in the middle region of the gene. Haplotype and gene network analyses were performed on the cyt b gene sequences of 130 Lu. peruensis in 9 Andean areas from 3 Departments (Ancash, Lima and La Libertad). The results showed that the populations of La Libertad were highly polymorphic and that their haplotypes were distinct from those of Ancash and Lima, where dominant haplotypes were observed, suggesting that a population bottleneck may have occurred in Ancash and Lima, but not in La Libertad. The present study indicated that the middle region of the cyt b gene is useful for the analysis of genetic structure in sand fly populations. Copyright © 2013 Elsevier B.V. All rights reserved.

  11. Cryopreservation of Leishmania Species in Manisa Province.

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    Çavuş, İbrahim; Ocak, Fulya; Kaya, Tuğba; Özbilgin, Ahmet

    2017-09-01

    It was aimed to assess the success of the cryopreservation process which is carried out in order to preserve the genetic material and the virulence of the Leishmania species that are an important health problem in our region. Leishmania tropica, L. infantum, L. major, and L. donovani strains in Novy-MacNeal-Nicolle (NNN) medium in MCBU were used. Promastigotes cultured in the NNN medium were transferred to RPMI 1640 medium; promastigotes in the logarithmic phase were washed three times with PBS, and 15% dimethylsulfoxide (DMSO) was added. Leishmania species were transferred to 12 separate tubes. The tubes were stored at -86°C for one night by placing them in Coolcell boxes. The tubes were transferred into a liquid nitrogen tank. One cryotube per Leishmania strain is thawed monthly and cultured in NNN medium. For the duration of study it was observed that each Leishmania isolate preserved 60-65% of their viability and entered the logarithmic phase on the 7th day following the inoculation in the NNN medium. Abnormalities in the structures and movements of the promastigotes were not observed in microscopic examinations. The following conclusions were made: cryopreservation is important for studies planned related to leishmaniasis and cryopreservation with DMSO is successful.

  12. Species-specific ecological niche modelling predicts different range contractions for Lutzomyia intermedia and a related vector of Leishmania braziliensis following climate change in South America.

    Science.gov (United States)

    McIntyre, Shannon; Rangel, Elizabeth F; Ready, Paul D; Carvalho, Bruno M

    2017-03-24

    Before 1996 the phlebotomine sand fly Lutzomyia neivai was usually treated as a synonym of the morphologically similar Lutzomyia intermedia, which has long been considered a vector of Leishmania braziliensis, the causative agent of much cutaneous leishmaniasis in South America. This report investigates the likely range changes of both sand fly species in response to a stabilisation climate change scenario (RCP4.5) and a high greenhouse gas emissions one (RCP8.5). Ecological niche modelling was used to identify areas of South America with climates currently suitable for each species, and then the future distributions of these climates were predicted based on climate change scenarios. Compared with the previous ecological niche model of L. intermedia (sensu lato) produced using the GARP algorithm in 2003, the current investigation modelled the two species separately, making use of verified presence records and additional records after 2001. Also, the new ensemble approach employed ecological niche modelling algorithms (including Maximum Entropy, Random Forests and Support Vector Machines) that have been widely adopted since 2003 and perform better than GARP, as well as using a more recent climate change model (HadGEM2) considered to have better performance at higher resolution than the earlier one (HadCM2). Lutzomyia intermedia was shown to be the more tropical of the two species, with its climatic niche defined by higher annual mean temperatures and lower temperature seasonality, in contrast to the more subtropical L. neivai. These different latitudinal ranges explain the two species' predicted responses to climate change by 2050, with L. intermedia mostly contracting its range (except perhaps in northeast Brazil) and L. neivai mostly shifting its range southwards in Brazil and Argentina. This contradicts the findings of the 2003 report, which predicted more range expansion. The different findings can be explained by the improved data sets and modelling methods. Our

  13. Analysis of sandflies (Diptera: Psychodidae) in Barra do Garças, State of Mato Grosso, Brazil, and the influence of environmental variables on the vector density of Lutzomyia longipalpis (Lutz & Neiva, 1912).

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    Queiroz, Mirian Francisca Martins; Varjão, Jane Ramos; Moraes, Sinara Cristina de; Salcedo, Gladys Elena

    2012-06-01

    Leishmaniasis is an infectious and parasitic zoonotic, non-contagious, vector-borne disease caused by protozoa of the genus Leishmania. In Brazil, the major vector of Leishmania (Leishmania) infantum chagasi (Cunha & Chagas, 1934) is Lutzomyia longipalpis. Barra do Garças, State of Mato Grosso, was designated as a priority area by the Brazilian Ministry of Health for american visceral leishmaniasis, and it is important to identify the vector species present in this municipality. Our objective was to raise sandflies and study the influence of environmental variables on the vector density of Lutzomyia longipalpis. We performed entomological monitoring in 3 districts using Centers for Disease Control and Prevention (CDC) light traps and recorded human cases of american visceral leishmaniasis in the city. We calculated the relative frequency and richness of sandflies and applied a transfer function model to the vector density correlate with relative humidity. The sandfly population was composed of 2 genera and 27 species, totaling 8,097 individuals. Monitoring identified Lutzomyia longipalpis (44%), followed by Lutzomyia lenti (18.9%), Lutzomyia whitmani (13.9%), Lutzomyia carmelinoi (9.1%), Lutzomyia evandroi (5.1%), Lutzomyia termitophila (3.3%), Lutzomyia sordellii (1.9%), and 20 other species (Lutzomyia longipalpis showed occurrence peaks during the rainy season; there was a temporal correlation with humidity, but not with frequency or temperature. The presence of Lutzomyia longipalpis in the urban area of Barra do Garças underscores the changing disease profile, which was previously restricted to the wild environment.

  14. Canine babesiosis in northern Portugal and molecular characterization of vector-borne co-infections

    Science.gov (United States)

    2010-01-01

    Background Protozoa and bacteria transmitted by arthropods, including ticks and phlebotomine sand flies, may cause a wide range of canine vector-borne diseases. Dogs can be simultaneously or sequentially infected with multiple pathogens. Canine babesiosis caused by Babesia canis canis and Babesia canis vogeli is known to occur in Portugal. This study assessed, by means of blood smear examination, PCR and DNA nucleotide sequencing, the presence of Babesia spp. and co-infecting agents Leishmania, Anaplasma/Ehrlichia and Hepatozoon in 45 dogs from northern Portugal clinically suspected of babesiosis. Results Forty-four dogs (98%) had infection with B. canis canis and one with B. canis vogeli. Co-infections were detected in nine animals (20%). Eight dogs were found infected with two vector-borne agents: six with B. canis canis and Leishmania infantum; one with B. canis canis and Ehrlichia canis; and one with B. canis canis and Hepatozoon canis. Another dog was infected with three vector-borne pathogens: B. canis vogeli, E. canis and L. infantum. Overall, L. infantum was found in seven (16%), E. canis in two (4%), and H. canis in one (2%) out of the 45 dogs with babesiosis. Almost 90% of the 45 cases of canine babesiosis were diagnosed in the colder months of October (18%), November (27%), December (20%), February (13%) and March (9%). Co-infections were detected in February, March, April, May, October and November. Twenty-two (50%) out of 44 dogs infected with B. canis were found infested by ticks including Dermacentor spp., Ixodes spp. and Rhipicephalus sanguineus. Mortality (9%) included two co-infected dogs that died spontaneously and two with single infections that were euthanized. Conclusions Babesia canis canis is the main etiological agent of canine babesiosis in northern Portugal. A higher sensitivity of Babesia spp. detection was obtained with PCR assays, compared to the observation of blood smears. Twenty percent of the dogs were co-infected with L. infantum

  15. Canine babesiosis in northern Portugal and molecular characterization of vector-borne co-infections

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    Machado João

    2010-04-01

    Full Text Available Abstract Background Protozoa and bacteria transmitted by arthropods, including ticks and phlebotomine sand flies, may cause a wide range of canine vector-borne diseases. Dogs can be simultaneously or sequentially infected with multiple pathogens. Canine babesiosis caused by Babesia canis canis and Babesia canis vogeli is known to occur in Portugal. This study assessed, by means of blood smear examination, PCR and DNA nucleotide sequencing, the presence of Babesia spp. and co-infecting agents Leishmania, Anaplasma/Ehrlichia and Hepatozoon in 45 dogs from northern Portugal clinically suspected of babesiosis. Results Forty-four dogs (98% had infection with B. canis canis and one with B. canis vogeli. Co-infections were detected in nine animals (20%. Eight dogs were found infected with two vector-borne agents: six with B. canis canis and Leishmania infantum; one with B. canis canis and Ehrlichia canis; and one with B. canis canis and Hepatozoon canis. Another dog was infected with three vector-borne pathogens: B. canis vogeli, E. canis and L. infantum. Overall, L. infantum was found in seven (16%, E. canis in two (4%, and H. canis in one (2% out of the 45 dogs with babesiosis. Almost 90% of the 45 cases of canine babesiosis were diagnosed in the colder months of October (18%, November (27%, December (20%, February (13% and March (9%. Co-infections were detected in February, March, April, May, October and November. Twenty-two (50% out of 44 dogs infected with B. canis were found infested by ticks including Dermacentor spp., Ixodes spp. and Rhipicephalus sanguineus. Mortality (9% included two co-infected dogs that died spontaneously and two with single infections that were euthanized. Conclusions Babesia canis canis is the main etiological agent of canine babesiosis in northern Portugal. A higher sensitivity of Babesia spp. detection was obtained with PCR assays, compared to the observation of blood smears. Twenty percent of the dogs were co

  16. Serological survey of Leishmania infection in dogs from the municipality of Peso da Régua (Alto Douro, Portugal) using the direct agglutination test (DAT) and fast agglutination screening test (FAST)

    NARCIS (Netherlands)

    Cardoso, Luís; Schallig, Henk D. F. H.; Neto, Francisco; Kroon, Nel; Rodrigues, Manuela

    2004-01-01

    Leishmaniasis caused by Leishmania infantum is a prevalent disease in dogs and humans. A serological survey of Leishmania infection in dogs was carried out in the endemic region of Alto Douro (north Portugal). Two hundred and ninety-four dogs from the municipality of Peso da Régua were examined for

  17. Vaccine development against Leishmania donovani

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    Amrita eDas

    2012-05-01

    Full Text Available Visceral leishmaniasis (VL caused by Leishmania donovani and Leishmania infantum/ chagasi represents the second most challenging infectious disease worldwide, affecting nearly 500,000 people and 60,000 deaths annually. Zoonotic VL (ZVL caused by L. infantum is re-emergent canid zoonoses which represents a complex epidemiological cycle in New world where domestic dogs serve as reservoir host responsible for potentially fatal human infection where dog culling is the only control measure for eliminating reservoir host. Lifelong immunity in human against reinfection has motivated several attempts in developing prophylactic vaccines against the disease but very few have progressed beyond experimental stage. Absence of any licensed vaccine along with high toxicity and increasing resistance to the current chemotherapeutic drugs has further complicated the situation in endemic regions of the world. Advances in vaccinology, including recombinant proteins, novel antigen-delivery systems/adjuvants, heterologous prime-boost regimens and strategies for intracellular antigen presentation, have contributed to recent advances in vaccine development against VL. Attempts to develop an effective vaccine for use in domestic dogs in areas of canine VL should be pursued for preventing human infection. Studies in animal models and human patients have revealed the pathogenic mechanisms of disease progression and features of protective immunity. This review will summarize the accumulated knowledge about pathogenesis, immune response and prerequisites for protective immunity against human VL. Authors will discuss promising vaccine targets, their developmental status and future prospects in a quest for rational vaccine development against VL. In addition, several challenges such as safety issues, a renewed and coordinated commitment to basic research, preclinical studies and trial design will be addressed to overcome the problems faced in developing effective vaccines

  18. The population structure of Lutzomyia verrucarum (Diptera: Psycodidae), a Bartonella bacilliformis and Leishmania peruviana vector in Peru.

    Science.gov (United States)

    Cohnstaedt, Lee W; Caceres, Abraham G; Beati, Lorenza; Munstermann, Leonard E

    2012-01-01

    The population genetic structure of Lutzomyia verrucarum (Townsend), a sand fly disease vector of Carrion's disease and cutaneous leishmaniasis in the Peruvian Andes, was characterized by sequencing 653 bp of cytochrome b and 1,125 bp of the NADH dehydrogenase subunit 4 genes of its mitochondrial genome. DNA sequence variation within and between valleys was compared in a sample of 220 sand flies from three valleys (Purisima, Huaylas, and Conchucos) and five departments (Amazonas, Cajamarca, Piura, Lima, and Huancavelica). Gene network and phylogenetic analyses indicated a high similarity of haplotypes collected within a single valley (0-0.52% nucleotide divergence). Flies from each valley had unique genotypes not shared with specimens from other valleys or from more distant regions (0.8-3.1% nucleotide divergence). Mountain ranges and geographic distance appear to have impeded migration (N(m) = < 0.18) between valleys and separated populations into discrete genetic units.

  19. Comparison of LAMP and PCR for molecular mass screening of sand flies for Leishmania martiniquensis infection.

    Science.gov (United States)

    Tiwananthagorn, Saruda; Kato, Hirotomo; Yeewa, Ranchana; Muengpan, Amontip; Polseela, Raxsina; Leelayoova, Saovanee

    2017-02-01

    Leishmaniasis caused by Leishmania martiniquensis infection has been reported in human and domestic animals of Martinique Island, Germany, Switzerland, USA, Myanmar and Thailand. The peculiar clinical features of disseminated cutaneous and visceral forms co-existence render the urgent need of specific diagnostic tool to identify the natural sand fly vectors for effective prevention and control strategies. Loop-mediated isothermal amplification (LAMP) of 18S rRNA gene as well as polymerase chain reaction (PCR) of minicircle kinetoplast DNA gene (PCR-mkDNA) have never been applied to detect L. martiniquensis and L. siamensis in sand fly vectors. The present study was aimed to validate malachite green-LAMP (MG-LAMP) and PCR-mkDNA techniques to detect L. martiniquensis in sand fly vectors, compared with the conventional PCR of internal transcribed spacer 1 (PCR-ITS1). We compared the validity of LAMP of 18S rRNA gene and PCR-mkDNA, to PCR-ITS1 in simulation model of L. martiniquensis infection in Sergentomyia gemmea sand flies. Attributable to the sensitivity and specificity, PCR-mkDNA was consecutively applied to detect L. martiniquensis in 380 female sand fly individuals captured in the newly identified affected region of Lamphun Province, Thailand. Results showed that PCR-mkDNA could detect at least one promastigote per sand fly, which was 10-time superior to LAMP and PCR-ITS1. In addition, PCR-mkDNA was more specific, able to differentiate L. martiniquensis from other viscerotropic Leishmania species, such as L. siamensis, L. (L.) donovani, and L. (L.) infantum. Consecutively, mass screening of L. martiniquensis in 380 female sand fly individuals by PCR-mkDNA was implemented in a new affected area of Thailand where a patient with leishmaniasis/HIV co-infection resides; however Leishmania DNA was undetected. In conclusion, PCR-mkDNA is a promising tool for molecular mass screening of L. martiniquensis infection in outbreak areas where several species of Leishmania

  20. Comparison of LAMP and PCR for molecular mass screening of sand flies for Leishmania martiniquensis infection

    Science.gov (United States)

    Tiwananthagorn, Saruda; Kato, Hirotomo; Yeewa, Ranchana; Muengpan, Amontip; Polseela, Raxsina; Leelayoova, Saovanee

    2017-01-01

    BACKGROUND Leishmaniasis caused by Leishmania martiniquensis infection has been reported in human and domestic animals of Martinique Island, Germany, Switzerland, USA, Myanmar and Thailand. The peculiar clinical features of disseminated cutaneous and visceral forms co-existence render the urgent need of specific diagnostic tool to identify the natural sand fly vectors for effective prevention and control strategies. Loop-mediated isothermal amplification (LAMP) of 18S rRNA gene as well as polymerase chain reaction (PCR) of minicircle kinetoplast DNA gene (PCR-mkDNA) have never been applied to detect L. martiniquensis and L. siamensis in sand fly vectors. OBJECTIVE The present study was aimed to validate malachite green-LAMP (MG-LAMP) and PCR-mkDNA techniques to detect L. martiniquensis in sand fly vectors, compared with the conventional PCR of internal transcribed spacer 1 (PCR-ITS1). METHODS We compared the validity of LAMP of 18S rRNA gene and PCR-mkDNA, to PCR-ITS1 in simulation model of L. martiniquensis infection in Sergentomyia gemmea sand flies. Attributable to the sensitivity and specificity, PCR-mkDNA was consecutively applied to detect L. martiniquensis in 380 female sand fly individuals captured in the newly identified affected region of Lamphun Province, Thailand. FINDINGS AND MAIN CONCLUSIONS Results showed that PCR-mkDNA could detect at least one promastigote per sand fly, which was 10-time superior to LAMP and PCR-ITS1. In addition, PCR-mkDNA was more specific, able to differentiate L. martiniquensis from other viscerotropic Leishmania species, such as L. siamensis, L. (L.) donovani, and L. (L.) infantum. Consecutively, mass screening of L. martiniquensis in 380 female sand fly individuals by PCR-mkDNA was implemented in a new affected area of Thailand where a patient with leishmaniasis/HIV co-infection resides; however Leishmania DNA was undetected. In conclusion, PCR-mkDNA is a promising tool for molecular mass screening of L. martiniquensis

  1. Detection and characterization of Leishmania (Leishmania and Leishmania (Viannia by SYBR green-based real-time PCR and high resolution melt analysis targeting kinetoplast minicircle DNA.

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    Marcello Ceccarelli

    Full Text Available Leishmaniasis is a neglected disease with a broad clinical spectrum which includes asymptomatic infection. A thorough diagnosis, able to distinguish and quantify Leishmania parasites in a clinical sample, constitutes a key step in choosing an appropriate therapy, making an accurate prognosis and performing epidemiological studies. Several molecular techniques have been shown to be effective in the diagnosis of leishmaniasis. In particular, a number of PCR methods have been developed on various target DNA sequences including kinetoplast minicircle constant regions. The first aim of this study was to develop a SYBR green-based qPCR assay for Leishmania (Leishmania infantum detection and quantification, using kinetoplast minicircle constant region as target. To this end, two assays were compared: the first used previously published primer pairs (qPCR1, whereas the second used a nested primer pairs generating a shorter PCR product (qPCR2. The second aim of this study was to evaluate the possibility to discriminate among subgenera Leishmania (Leishmania and Leishmania (Viannia using the qPCR2 assay followed by melting or High Resolution Melt (HRM analysis. Both assays used in this study showed good sensitivity and specificity, and a good correlation with standard IFAT methods in 62 canine clinical samples. However, the qPCR2 assay allowed to discriminate between Leishmania (Leishmania and Leishmania (Viannia subgenera through melting or HRM analysis. In addition to developing assays, we investigated the number and genetic variability of kinetoplast minicircles in the Leishmania (L. infantum WHO international reference strain (MHOM/TN/80/IPT1, highlighting the presence of minicircle subclasses and sequence heterogeneity. Specifically, the kinetoplast minicircle number per cell was estimated to be 26,566±1,192, while the subclass of minicircles amplifiable by qPCR2 was estimated to be 1,263±115. This heterogeneity, also observed in canine clinical

  2. Validación de la PCR en la detección de parásitos de Leishmania (Viannia spp. en Lutzomyia (Diptera: Psychodidae como herramienta en la definición de especies vectores.

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    Erika Santamaría

    2005-06-01

    Full Text Available lntroducción. En leishmaniasis se acepta que la reacción en cadena de la polimerasa (PCR ha simplificado el proceso de incriminación vectorial. Sin embargo, pocas veces se ha determinado la sensibilidad y la especificidad de cada PCR en la detección y la identificación del parasito en los flebótomos. Objetivo. Evaluar la aplicabilidad de la reacción en cadena de la polimerasa (PCR, basada en los iniciadores 81 y 82, en la detecci6n e identificación de parasitos de Leishmania (Viannia en insectos vectores enteros sin disecar. Metodología. Se determinó la sensibilidad y la especificidad de la PCR empleando diluciones de cultivo de parasitos de Leishmania spp. Se estableció el número máximo de hembras de Lutzomyia que pueden ser procesadas a la vez sin disminuir la sensibilidad de la PCR, procesando el ADN de grupos de una a cinco hembras de Lutzomyia en presencia del ADN de las diluciones de cultivo de parasitos. Además, se comparó en grupos de flebótomos infectados experimentalmente, la sensibilidad de esta PCR en la detección de infección por Leishmania (Viannia frente al método de busqueda de flagelados por disección del insecto y examen microscópico. Resultados. La PCR detectó desde un parasito de Leishmania (Viannia y permiti6 el procesamiento de hasta tres insectos enteros sin alterar la sensibilidad. Los porcentajes de infección experimental detectados con las dos tecnicas fueron similares, 33,3% con la PCR y 30% con el examen microscópico. Además, se confirmó que los iniciadores 81 y 82 son especificos para especies del subgenero Leishmania (Viannia. Conclusión. Los resultados obtenidos demuestran la sensibilidad y la especificidad de esta PCR y permiten recomendar su uso en la determinaci6n de infección natural con parasitos de Leishmania (Viannia en poblaciones silvestres de flebótomos.

  3. High density of Leishmania major and rarity of other mammals' Leishmania in zoonotic cutaneous leishmaniasis foci, Iran.

    Science.gov (United States)

    Bordbar, Ali; Parvizi, Parviz

    2014-03-01

    Only Leishmania major is well known as a causative agent of zoonotic cutaneous leishmaniasis (ZCL) in Iran. Our objective was to find Leishmania parasites circulating in reservoir hosts, sand flies and human simultaneously. Sand flies, rodents and prepared smears of humans were sampled. DNA of Leishmania parasites was extracted, and two fragments of ITS-rDNA gene amplified by PCR. RFLP and sequencing were employed to identify Leishmania parasites. Leishmania major and L. turanica were identified unequivocally by targeting and sequencing ITS-rDNA from humans, rodents and sand flies. The new Leishmania species close to gerbilli (GenBank Accession Nos. EF413076; EF413087) was discovered only in sand flies. Based on parasite detection of ITS-rDNA in main and potential reservoir hosts and vectors and humans, we conclude that at least two Leishmania species are common in the Turkmen Sahra ZCL focus. Phylogenetic analysis proved that the new Leishmania is closely related to Leishmania mammal parasites (Leishmania major, Leishmania turanica, Leishmania gerbilli). Its role as a principal agent of ZCL is unknown because it was found only in sand flies. Our findings shed new light on the transmission cycles of several Leishmania parasites in sand flies, reservoir hosts and humans. © 2014 John Wiley & Sons Ltd.

  4. Presence of Putative Male-Produced Sex Pheromone in Lutzomyia cruciata (Diptera: Psychodidae), Vector of Leishmania mexicana.

    Science.gov (United States)

    Serrano, A K; Rojas, J C; Cruz-López, L C; Malo, Edi A; Mikery, O F; Castillo, A

    2016-11-01

    Lutzomyia cruciata (Coquillet) is a vector of cutaneous leishmaniasis in Mexico and Central America. However, several aspects of its ecology and behavior are unknown, including whether a male pheromone partially mediates the sexual behavior of this sand fly. In this study, we evaluated the behavioral response of females to male abdominal extracts in a Y-tube olfactometer. The volatile compounds from male abdominal extracts were identified by gas chromatography-mass spectrometry and compared with those of female abdominal extracts. Finally, the disseminating structures of the putative sex pheromone were examined by scanning electron microscopy in the male abdomen. Females were more attracted to male abdominal extract than to the hexane control, suggesting the presence of male-produced sex pheromone. The male abdominal extracts were characterized by the presence of 12 sesquiterpene compounds. The major component, an unknown sesquiterpene with an abundance of 60%, had a mass spectrum with molecular ion of m/z 262. In contrast, the abdominal female extracts contained saturated fatty acids. Finally, we detected the presence of small "papules" with a mammiform morphology distributed on the abdominal surface of tergites IV-VII of male Lu. cruciata These structures are not present in females. We conclude that Lu. cruciata males likely produce a pheromone involved in attracting or courting females. © The Authors 2016. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  5. Genetic variability in biochemical characters of Brazilian field populations of the Leishmania vector, Lutzomyia longipalpis (Diptera: Psychodidae).

    Science.gov (United States)

    Mukhopadhyay, J; Ghosh, K; Rangel, E F; Munstermann, L E

    1998-12-01

    The phlebotomine sand fly Lutzomyia longipalpis is the insect vector of visceral leishmaniasis, a protozoan disease of increasing incidence and distribution in Central and South America. Electrophoretic allele frequencies of 15 enzyme loci were compared among the L. longipalpis populations selected across its distribution range in Brazil. The mean heterozygosity of two colonized geographic strains (one each from Colombia and Brazil) were 6% and 13% respectively, with 1.6-1.9 alleles detected per locus. In contrast, among the seven widely separated field populations, the mean heterozygosity ranged from 11% to 16% with 2.1-2.9 alleles per locus. No locus was recovered that was diagnostic for any of the field populations. Allelic frequency differences among five field strains from the Amazon basin and eastern coastal Brazil were very low, with Nei's genetic distances of less than 0.01 separating them. The two inland and southerly samples from Minas Gerais (Lapinha) and Bahia (Jacobina) states were more distinctive with genetic distances of 0.024-0.038 and 0.038-0.059, respectively, when compared with the five other samples. These differences were the consequence of several high frequency alleles (glycerol-3-phosphate dehydrogenase [Gpd1.69] and phosphoglucomutase [Pgm1.69]) relatively uncommon in other strains. The low genetic distances, absence of diagnostic loci, and the distribution of genes in geographic space indicate L. longipalpis of Brazil to be a single, but genetically heterogeneous, polymorphic species.

  6. Molecular Characterization of Leishmania Parasites in Giemsa-Stained Slides from Cases of Human Cutaneous and Visceral Leishmaniasis, Eastern Algeria.

    Science.gov (United States)

    Beldi, Nadia; Mansouri, Roukaya; Bettaieb, Jihene; Yaacoub, Alia; Souguir Omrani, Hejer; Saadi Ben Aoun, Yusr; Saadni, Farida; Guizani, Ikram; Guerbouj, Souheila

    2017-06-01

    In Algeria, visceral leishmaniasis (VL) is due to Leishmania (L.) infantum, while three cutaneous forms (CL) are caused by Leishmania major, Leishmania tropica and Leishmania infantum. In this study, the use of Giemsa-stained slides was evaluated with two PCR techniques, in Eastern Algeria. A total of 136 samples corresponding to 100 CL smears (skin scrapings) and 36 VL slides (bone marrow aspirates) collected from 2008 to 2014 were tested. Upon DNA extraction, two PCRs were used to amplify the ribosomal Internal Transcribed Spacer 1 (ITS1) and mini-exon genes. Amplified products were digested (PCR-RFLP) and profiles analyzed for Leishmania species identification. A statistical analysis was also performed. ITS1-PCR was found significantly more sensitive than mini-exon-PCR (77.95% positives vs. 67.65%; p = 0.001). Comparison of PCR positivity showed statistically significant differences between old and recently prepared slides suggesting a better use of recent slides in PCR analyses. For species identification, PCR-restriction fragment length polymorphism (RFLP) results of ITS1 and mini-exon were concordant. L. infantum was identified from VL cases and L. infantum, L. major, and L. tropica from CL ones. According to geographical origin, L. infantum was found in North-Eastern provinces, while L. major was distributed from the North to the Center-East of Algeria. Interestingly, two L. tropica samples were identified in Annaba, located far North-East Algeria. Distribution of leishmaniasis in Eastern parts of Algeria, besides finding of L. tropica in the far North, is in this study described for the first time using molecular tools, thus confirming the usefulness of slides for PCR identification of Leishmania parasites in retrospective epidemiological investigations.

  7. Leishmania hijacking of the macrophage intracellular compartments.

    Science.gov (United States)

    Liévin-Le Moal, Vanessa; Loiseau, Philippe M

    2016-02-01

    Leishmania spp., transmitted to humans by the bite of the sandfly vector, are responsible for the three major forms of leishmaniasis, cutaneous, diffuse mucocutaneous and visceral. Leishmania spp. interact with membrane receptors of neutrophils and macrophages. In macrophages, the parasite is internalized within a parasitophorous vacuole and engages in a particular intracellular lifestyle in which the flagellated, motile Leishmania promastigote metacyclic form differentiates into non-motile, metacyclic amastigote form. This phenomenon is induced by Leishmania-triggered events leading to the fusion of the parasitophorous vacuole with vesicular members of the host cell endocytic pathway including recycling endosomes, late endosomes and the endoplasmic reticulum. Maturation of the parasitophorous vacuole leads to the intracellular proliferation of the Leishmania amastigote forms by acquisition of host cell nutrients while escaping host defense responses. © 2015 FEBS.

  8. Phylogenetic analysis of HSP70 and cyt b gene sequences for Chinese Leishmania isolates and ultrastructural characteristics of Chinese Leishmania sp.

    Science.gov (United States)

    Yuan, Dongmei; Qin, Hanxiao; Zhang, Jianguo; Liao, Lin; Chen, Qiwei; Chen, Dali; Chen, Jianping

    2017-02-01

    Leishmaniasis is a worldwide epidemic disease caused by the genus Leishmania, which is still endemic in the west and northwest areas of China. Some viewpoints of the traditional taxonomy of Chinese Leishmania have been challenged by recent phylogenetic researches based on different molecular markers. However, the taxonomic positions and phylogenetic relationships of Chinese Leishmania isolates remain controversial, which need for more data and further analysis. In this study, the heat shock protein 70 (HSP70) gene and cytochrome b (cyt b) gene were used for phylogenetic analysis of Chinese Leishmania isolates from patients, dogs, gerbils, and sand flies in different geographic origins. Besides, for the interesting Leishmania sp. in China, the ultrastructure of three Chinese Leishmania sp. strains (MHOM/CN/90/SC10H2, SD, GL) were observed by transmission electron microscopy. Bayesian trees from HSP70 and cyt b congruently indicated that the 14 Chinese Leishmania isolates belong to three Leishmania species including L. donovani complex, L. gerbilli, and L. (Sauroleishmania) sp. Their identity further confirmed that the undescribed Leishmania species causing visceral Leishmaniasis (VL) in China is closely related to L. tarentolae. The phylogenetic results from HSP70 also suggested the classification of subspecies within L. donovani complex: KXG-918, KXG-927, KXG-Liu, KXG-Xu, 9044, SC6, and KXG-65 belong to L. donovani; Cy, WenChuan, and 801 were proposed to be L. infantum. Through transmission electron microscopy, unexpectedly, the Golgi apparatus were not observed in SC10H2, SD, and GL, which was similar to previous reports of reptilian Leishmania. The statistical analysis of microtubule counts separated SC10H2, SD, and GL as one group from any other reference strain (L. donovani MHOM/IN/80/DD8; L. tropica MHOM/SU/74/K27; L. gerbilli MRHO/CN/60/GERBILLI). The ultrastructural characteristics of Leishmania sp. partly lend support to the phylogenetic inference that

  9. Anti-Leishmania activity of new ruthenium(II) complexes: Effect on parasite-host interaction.

    Science.gov (United States)

    Costa, Mônica S; Gonçalves, Yasmim G; Nunes, Débora C O; Napolitano, Danielle R; Maia, Pedro I S; Rodrigues, Renata S; Rodrigues, Veridiana M; Von Poelhsitz, Gustavo; Yoneyama, Kelly A G

    2017-10-01

    Leishmaniasis is a parasitic disease caused by protozoa of the genus Leishmania. The many complications presented by the current treatment - including high toxicity, high cost and parasite resistance - make the development of new therapeutic agents indispensable. The present study aims to evaluate the anti-Leishmania potential of new ruthenium(II) complexes, cis‑[Ru II (η 2 -O 2 CR)(dppm) 2 ]PF 6 , with dppm=bis(diphenylphosphino)methane and R=4-butylbenzoate (bbato) 1, 4-(methylthio)benzoate (mtbato) 2 and 3-hydroxy-4-methoxybenzoate (hmxbato) 3, in promastigote cytotoxicity and their effect on parasite-host interaction. The cytotoxicity of complexes was analyzed by MTT assay against Leishmania (Leishmania) amazonensis, Leishmania (Viannia) braziliensis, Leishmania (Leishmania) infantum promastigotes and the murine macrophage (RAW 264.7). The effect of complexes on parasite-host interaction was evaluated by in vitro infectivity assay performed in the presence of two different concentrations of each complex: the promastigote IC 50 value and the concentration nontoxic to 90% of RAW 264.7 macrophages. Complexes 1-3 exhibited potent cytotoxic activity against all Leishmania species assayed. The IC 50 values ranged from 7.52-12.59μM (complex 1); 0.70-3.28μM (complex 2) and 0.52-1.75μM (complex 3). All complexes significantly inhibited the infectivity index at both tested concentrations. The infectivity inhibitions ranged from 37 to 85%. Interestingly, the infectivity inhibitions due to complex action did not differ significantly at either of the tested concentrations, except for the complex 1 against Leishmania (Leishmania) infantum. The infectivity inhibitions resulted from reductions in both percentage of infected macrophages and number of parasites per macrophage. Taken together the results suggest remarkable leishmanicidal activity in vitro by these new ruthenium(II) complexes. Copyright © 2017 Elsevier Inc. All rights reserved.

  10. Canine and feline vector-borne diseases in Italy: current situation and perspectives

    Directory of Open Access Journals (Sweden)

    Dantas-Torres Filipe

    2010-01-01

    Full Text Available Abstract In Italy, dogs and cats are at risk of becoming infected by different vector-borne pathogens, including protozoa, bacteria, and helminths. Ticks, fleas, phlebotomine sand flies, and mosquitoes are recognized vectors of pathogens affecting cats and dogs, some of which (e.g., Anaplasma phagocytophilum, Borrelia burgdorferi, Dipylidium caninum, Leishmania infantum, Dirofilaria immitis, and Dirofilaria repens are of zoonotic concern. Recent studies have highlighted the potential of fleas as vectors of pathogens of zoonotic relevance (e.g., Rickettsia felis in this country. While some arthropod vectors (e.g., ticks and fleas are present in certain Italian regions throughout the year, others (e.g., phlebotomine sand flies are most active during the summer season. Accordingly, control strategies, such as those relying on the systematic use of acaricides and insecticides, should be planned on the basis of the ecology of both vectors and pathogens in different geographical areas in order to improve their effectiveness in reducing the risk of infection by vector-borne pathogens. This article reviews the current situation and perspectives of canine and feline vector-borne diseases in Italy.

  11. Lutzomyia gasparviannai Martins, Godoy & Silva, 1962, probable vector of Leishmania mexicana ssp. in Viana municipality, Espírito Santo State, Brazil

    OpenAIRE

    Falqueto,Aloísio; Grimaldi Júnior,Gabriel; Sessa,Paulo Augusto; Varejão,José Benedito Malta; Deane,Leonidas M.

    1985-01-01

    Dos flebótomos atraídos pelo Proechimys iheringi numa área onde esse roedor foi achado naturalmente infectado por Leishmania mexicana ssp., 98,1% foram Lutzomyia gasparviannai, o que sugere que essa espécie não antropofílica seja o transmissor entre os roedores mas não habitualmente ao homem.

  12. Eurasian golden jackal as host of canine vector-borne protists.

    Science.gov (United States)

    Mitková, Barbora; Hrazdilová, Kristýna; D'Amico, Gianluca; Duscher, Georg Gerhard; Suchentrunk, Franz; Forejtek, Pavel; Gherman, Călin Mircea; Matei, Ioana Adriana; Ionică, Angela Monica; Daskalaki, Aikaterini Alexandra; Mihalca, Andrei Daniel; Votýpka, Jan; Hulva, Pavel; Modrý, David

    2017-04-14

    Jackals are medium-sized canids from the wolf-like clade, exhibiting a unique combination of ancestral morphotypes, broad trophic niches, and close phylogenetic relationships with the wolf and dog. Thus, they represent a potential host of several pathogens with diverse transmission routes. Recently, populations of the Eurasian golden jackal Canis aureus have expanded into the Western Palaearctic, including most of Europe. The aim of our study was to examine Eurasian golden jackals from Romania, Czech Republic and Austria for a wide spectrum of vector-borne protists and to evaluate the role of this species as a reservoir of disease for domestic dogs and/or humans. Diagnostic polymerase chain reaction (PCR) DNA amplifications revealed 70% of jackals to be positive for Hepatozoon, 12.5% positive for piroplasms, and one individual positive for Leishmania infantum. Phylogenetic analyses of partial 18S rDNA sequences invariably placed sequenced isolates of Hepatozoon into the H. canis clade. For piroplasms, both the 18S and cox1 sequences obtained confirmed the presence of Babesia canis and "Theileria annae" in 5 and 2 individuals, respectively, providing the first records of these two piroplasmids in Eurasian golden jackals. A single animal from Dolj County (Romania) was PCR-positive for L. infantum, as confirmed also by sequencing of ITS1-5.8S. Apparently, expanding populations of jackals can play a significant role in spreading and maintaining new Babesia canis foci in Central Europe. The role of jackals in the epidemiology of "Theileria annae" and H. canis is probably similar to that of red foxes and should be taken into account in further research on these parasites. Also the presence of L. infantum deserves attention. Our study confirms that once established, the populations of Eurasian golden jackals constitute natural reservoirs for many canine vector-borne diseases, analogous to the role of the coyotes in North America.

  13. Detection of Leishmania spp. in Bats from an Area of Brazil Endemic for Visceral Leishmaniasis.

    Science.gov (United States)

    de Rezende, M B; Herrera, H M; Carvalho, C M E; Carvalho Anjos, E A; Ramos, C A N; de Araújo, F R; Torres, J M; de Oliveira, C E

    2017-12-01

    The multihost parasites Leishmania spp. infect a broad range of wild mammalian species including bats. Several species of bats have adapted to a variety of food resources and shelters in urban areas. This study aimed to detect Leishmania spp. DNA in bats present in forest fragments located in metropolitan areas endemic for leishmaniasis in Campo Grande, Mato Grosso do Sul (MS), Brazil. Blood samples were obtained from 80 individuals, including eight species of Phyllostomidae and one species of Vespertilionidae. Thirty of the 80 bats were positive for Leishmania spp. using conventional PCR, all belonging to the family Phyllostomidae. Eighteen samples tested by real-time PCR (qPCR) using specific primers for the kDNA of Leishmania infantum were positive. To the best of our knowledge, this is the first report detecting Leishmania spp. in Platyrrhinus incarum in addition to being the first reported detection of L. infantum in the bat species Phyllostomus discolor, Platyrrhinus lineatus, Artibeus planirostris and Artibeus lituratus. Our results show that bats can host Leishmania spp. in areas endemic for leishmaniasis, which must be taken into account in disease control operations by public health authorities. © 2017 Blackwell Verlag GmbH.

  14. Cloning and characterization of a V-ATPase subunit C from the American visceral leishmaniasis vector Lutzomyia longipalpis modulated during development and blood ingestion

    Directory of Open Access Journals (Sweden)

    JM Ramalho-Ortigão

    2007-06-01

    Full Text Available Visceral leishmaniasis (VL is a serious tropical disease that affects approximately 500 thousand people worldwide every year. In the Americas, VL is caused by the parasite Leishmania (Leishmania infantum chagasi mainly transmitted by the bite of the sand fly vector Lutzomyia longipalpis. Despite recent advances in the study of interaction between Leishmania and sand flies, very little is known about sand fly protein expression profiles. Understanding how the expression of proteins may be affected by blood feeding and/or presence of parasite in the vector's midgut might allow us to devise new strategies for controlling the spread of leishmaniasis. In this work, we report the characterization of a vacuolar ATPase subunit C from L. longipalpis by screening of a midgut cDNA library with a 220 bp fragment identified by means of differential display reverse transcriptase-polymerase chain reaction analysis. The expression of the gene varies along insect development and is upregulated in males and bloodfed L. longipalpis, compared to unfed flies.

  15. Serological and molecular survey of Leishmania infection in dogs from Luanda, Angola.

    Science.gov (United States)

    Vilhena, Hugo; Granada, Sara; Oliveira, Ana Cristina; Schallig, Henk D F H; Nachum-Biala, Yaarit; Cardoso, Luís; Baneth, Gad

    2014-03-24

    Canine leishmaniosis (CanL) due to Leishmania infantum is a global zoonosis endemic in more than 70 countries in Europe, North Africa, Asia and America; however, data on this infection is scarce from southern Africa. The aim of this study was to survey dogs in Luanda, Angola, for Leishmania infection. One hundred-and-three dogs presented to a veterinary medical centre in Luanda were serologically and molecularly assessed for Leishmania with the direct agglutination test (DAT) and polymerase chain reaction (PCR). Two dogs were seropositive, with DAT titres of 800 and ≥6400; the latter was also found to be PCR-positive and confirmed to be infected with L. infantum by DNA sequence analysis. No other dog was found to be PCR-positive. The first dog had been imported from Portugal, but the latter had never left Angola (neither had its parents), strongly suggesting an autochthonous infection. Although other cases of CanL have previously been described in the country, this is the first reported study of canine Leishmania infection at the population level, as well as the first report on the molecular characterization of L. infantum in dogs from Angola.

  16. Monocyte-Derived Signals Activate Human Natural Killer Cells in Response to Leishmania Parasites

    Science.gov (United States)

    Messlinger, Helena; Sebald, Heidi; Heger, Lukas; Dudziak, Diana; Bogdan, Christian; Schleicher, Ulrike

    2018-01-01

    Activated natural killer (NK) cells release interferon (IFN)-γ, which is crucial for the control of intracellular pathogens such as Leishmania. In contrast to experimental murine leishmaniasis, the human NK cell response to Leishmania is still poorly characterized. Here, we investigated the interaction of human blood NK cells with promastigotes of different Leishmania species (Leishmania major, Leishmania mexicana, Leishmania infantum, and Leishmania donovani). When peripheral blood mononuclear cells or purified NK cells and monocytes (all derived from healthy blood donors from Germany without a history of leishmaniasis) were exposed to promastigotes, NK cells showed increased surface expression of the activation marker CD69. The extent of this effect varied depending on the Leishmania species; differences between dermotropic and viscerotropic L. infantum strains were not observed. Upregulation of CD69 required direct contact between monocytes and Leishmania and was partly inhibitable by anti-interleukin (IL)-18. Unexpectedly, IL-18 was undetectable in most of the supernatants (SNs) of monocyte/parasite cocultures. Confocal fluorescence microscopy of non-permeabilized cells revealed that Leishmania-infected monocytes trans-presented IL-18 to NK cells. Native, but not heat-treated SNs of monocyte/Leishmania cocultures also induced CD69 on NK cells, indicating the involvement of a soluble heat-labile factor other than IL-18. A role for the NK cell-activating cytokines IL-1β, IL-2, IL-12, IL-15, IL-21, and IFN-α/β was excluded. The increase of CD69 was not paralleled by NK cell IFN-γ production or enhanced cytotoxicity. However, prior exposure of NK cells to Leishmania parasites synergistically increased their IFN-γ release in response to IL-12, which was dependent on endogenous IL-18. CD1c+ dendritic cells were identified as possible source of Leishmania-induced IL-12. Finally, we observed that direct contact between Leishmania and NK cells reduced the

  17. Monocyte-Derived Signals Activate Human Natural Killer Cells in Response to Leishmania Parasites

    Directory of Open Access Journals (Sweden)

    Helena Messlinger

    2018-01-01

    Full Text Available Activated natural killer (NK cells release interferon (IFN-γ, which is crucial for the control of intracellular pathogens such as Leishmania. In contrast to experimental murine leishmaniasis, the human NK cell response to Leishmania is still poorly characterized. Here, we investigated the interaction of human blood NK cells with promastigotes of different Leishmania species (Leishmania major, Leishmania mexicana, Leishmania infantum, and Leishmania donovani. When peripheral blood mononuclear cells or purified NK cells and monocytes (all derived from healthy blood donors from Germany without a history of leishmaniasis were exposed to promastigotes, NK cells showed increased surface expression of the activation marker CD69. The extent of this effect varied depending on the Leishmania species; differences between dermotropic and viscerotropic L. infantum strains were not observed. Upregulation of CD69 required direct contact between monocytes and Leishmania and was partly inhibitable by anti-interleukin (IL-18. Unexpectedly, IL-18 was undetectable in most of the supernatants (SNs of monocyte/parasite cocultures. Confocal fluorescence microscopy of non-permeabilized cells revealed that Leishmania-infected monocytes trans-presented IL-18 to NK cells. Native, but not heat-treated SNs of monocyte/Leishmania cocultures also induced CD69 on NK cells, indicating the involvement of a soluble heat-labile factor other than IL-18. A role for the NK cell-activating cytokines IL-1β, IL-2, IL-12, IL-15, IL-21, and IFN-α/β was excluded. The increase of CD69 was not paralleled by NK cell IFN-γ production or enhanced cytotoxicity. However, prior exposure of NK cells to Leishmania parasites synergistically increased their IFN-γ release in response to IL-12, which was dependent on endogenous IL-18. CD1c+ dendritic cells were identified as possible source of Leishmania-induced IL-12. Finally, we observed that direct contact between Leishmania and NK cells

  18. Experimental Acquisition, Development, and Transmission of Leishmania tropica by Phlebotomus duboscqi

    Science.gov (United States)

    2013-01-01

    ac ta t ropica Experimental acquisition, development, and transmission of Leishmania tropica by Phlebotomus duboscqi Hanafi A. Hanafi, El...August 2012 Accepted 2 September 2012 Available online 10 September 2012 Keywords: Phlebotomus duboscqi Leishmania tropica Transmission Vector...competency a b s t r a c t We report experimental infection and transmission of Leishmania tropica (Wright), by the blood-feeding sand

  19. Vectors

    DEFF Research Database (Denmark)

    Boeriis, Morten; van Leeuwen, Theo

    2017-01-01

    should be taken into account in discussing ‘reactions’, which Kress and van Leeuwen link only to eyeline vectors. Finally, the question can be raised as to whether actions are always realized by vectors. Drawing on a re-reading of Rudolf Arnheim’s account of vectors, these issues are outlined......This article revisits the concept of vectors, which, in Kress and van Leeuwen’s Reading Images (2006), plays a crucial role in distinguishing between ‘narrative’, action-oriented processes and ‘conceptual’, state-oriented processes. The use of this concept in image analysis has usually focused...

  20. Association of pro-inflammatory cytokines and iron regulatory protein 2 (IRP2 with Leishmania burden in canine visceral leishmaniasis.

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    Paulo Ricardo Porfírio do Nascimento

    Full Text Available Leishmania infantum infection in humans and dogs can evolve with a wide range of clinical presentations, varying from asymptomatic infections to visceral leishmaniasis. We hypothesized that the immune response elicited by L. infantum infection could modulate whether the host will remain asymptomatic or progress to disease. A total of 44 dogs naturally infected with L. infantum were studied. Leishmania burden was estimated in the blood and spleen by qPCR. The expression of IFN-γ, TNF-α, IL-10 and Iron Regulatory Protein 2 (IRP2 were determined in the spleen by quantitative PCR. Sera cytokines were evaluated by ELISA. Dogs were grouped in quartiles according parasite burden. Increased expression of IFN-γ and TNF-α was associated with reduced Leishmania burden, whereas increased IL-10 and IRP2 expressions were associated with higher Leishmania load. Increased plasma albumin and IFN-γ expression explained 22.8% of the decrease in parasite burden in the spleen. These data confirm that lower IFN-γ response and higher IL-10 correlated with increased parasite load and severity of the visceral leishmaniasis in dogs. The balance between the branches of immune response and the intracellular iron availability could determine, in part, the course of Leishmania infection.

  1. Lutzomyia gasparviannai Martins, Godoy & Silva, 1962, probable vector of Leishmania mexicana ssp. in Viana municipality, Espírito Santo State, Brazil

    Directory of Open Access Journals (Sweden)

    Aloísio Falqueto

    1985-12-01

    Full Text Available Dos flebótomos atraídos pelo Proechimys iheringi numa área onde esse roedor foi achado naturalmente infectado por Leishmania mexicana ssp., 98,1% foram Lutzomyia gasparviannai, o que sugere que essa espécie não antropofílica seja o transmissor entre os roedores mas não habitualmente ao homem.

  2. Leishmania spp. Epidemiology of Canine Leishmaniasis in the Yucatan Peninsula

    Directory of Open Access Journals (Sweden)

    A. López-Céspedes

    2012-01-01

    Full Text Available Canine Leishmaniasis is widespread in various Mexican states, where different species of Leishmania have been isolated from dogs. In the present study, we describe the detection of L. braziliensis, L. infantum, and L. mexicana in serum of dogs from the states of Yucatan and Quintana Roo in the Yucatan Peninsula (Mexico. A total of 412 sera were analyzed by ELISA using the total extract of the parasite and the iron superoxide dismutase excreted by different trypanosomatids as antigens. We found the prevalence of L. braziliensis to be 7.52%, L. infantum to be 6.07%, and L. mexicana to be 20.63%, in the dog population studied. The results obtained with ELISA using iron superoxide dismutase as the antigen were confirmed by western blot analysis with its greater sensitivity, and the agreement between the two techniques was very high.

  3. Leishmania spp. epidemiology of canine leishmaniasis in the Yucatan Peninsula.

    Science.gov (United States)

    López-Céspedes, A; Longoni, S S; Sauri-Arceo, C H; Sánchez-Moreno, M; Rodríguez-Vivas, R I; Escobedo-Ortegón, F J; Barrera-Pérez, M A; Bolio-González, M E; Marín, C

    2012-01-01

    Canine Leishmaniasis is widespread in various Mexican states, where different species of Leishmania have been isolated from dogs. In the present study, we describe the detection of L. braziliensis, L. infantum, and L. mexicana in serum of dogs from the states of Yucatan and Quintana Roo in the Yucatan Peninsula (Mexico). A total of 412 sera were analyzed by ELISA using the total extract of the parasite and the iron superoxide dismutase excreted by different trypanosomatids as antigens. We found the prevalence of L. braziliensis to be 7.52%, L. infantum to be 6.07%, and L. mexicana to be 20.63%, in the dog population studied. The results obtained with ELISA using iron superoxide dismutase as the antigen were confirmed by western blot analysis with its greater sensitivity, and the agreement between the two techniques was very high.

  4. Identification of Tunisian Leishmania spp. by PCR amplification of cysteine proteinase B (cpb) genes and phylogenetic analysis.

    Science.gov (United States)

    Chaouch, Melek; Fathallah-Mili, Akila; Driss, Mehdi; Lahmadi, Ramzi; Ayari, Chiraz; Guizani, Ikram; Ben Said, Moncef; Benabderrazak, Souha

    2013-03-01

    Discrimination of the Old World Leishmania parasites is important for diagnosis and epidemiological studies of leishmaniasis. We have developed PCR assays that allow the discrimination between Leishmania major, Leishmania tropica and Leishmania infantum Tunisian species. The identification was performed by a simple PCR targeting cysteine protease B (cpb) gene copies. These PCR can be a routine molecular biology tools for discrimination of Leishmania spp. from different geographical origins and different clinical forms. Our assays can be an informative source for cpb gene studying concerning drug, diagnostics and vaccine research. The PCR products of the cpb gene and the N-acetylglucosamine-1-phosphate transferase (nagt) Leishmania gene were sequenced and aligned. Phylogenetic trees of Leishmania based cpb and nagt sequences are close in topology and present the classic distribution of Leishmania in the Old World. The phylogenetic analysis has enabled the characterization and identification of different strains, using both multicopy (cpb) and single copy (nagt) genes. Indeed, the cpb phylogenetic analysis allowed us to identify the Tunisian Leishmania killicki species, and a group which gathers the least evolved isolates of the Leishmania donovani complex, that was originated from East Africa. This clustering confirms the African origin for the visceralizing species of the L. donovani complex. Copyright © 2012 Elsevier B.V. All rights reserved.

  5. Identificación de una nueva proteína en Leishmania (Viannia peruviana

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    Maxy De los Santos

    1998-01-01

    Full Text Available El análisis de la secuencia nucleotídica y aminoacídica de un clon de la biblioteca de expresión en fago λgt11 de Leishmania (Viannia peruviana, estableció identidad parcial con los genes de las proteínas acídicas ribosomales P2 de Leishmania (Leishmania infantum. Este hallazgo unido a ciertos dominios geonómicos conservados, sugeridos de la comparación de 14 secuencias de otras proteínas P1 eucarióticas, confirman que la secuencia del inserto de clon codifica la proteína acídica ribosomal P1 de L. (V. peruviana denominada LpP1. Este es el primer reporte sobre este tipo de proteína en el género Leishmania.

  6. Molecular and serological detection of tick-borne pathogens in dogs from an area endemic for Leishmania infantumin Mato Grosso do Sul, Brazil

    Directory of Open Access Journals (Sweden)

    Keyla Carstens Marques de Sousa

    Full Text Available Tick-borne pathogens affect a wide range of vertebrate hosts. To identify tick-borne pathogens among dogs from Campo Grande, MS, Brazil testing seropositive for Leishmania infantum (syn. L. chagasi, a serological and molecular study was conducted to detectEhrlichia canis, Anaplasma platys and Babesia vogeli in 60 serum and spleen samples. A confirmatory diagnosis ofL. infantum based on serological and molecular assays was also performed, as was sequence alignment and phylogenetic analysis to assess the identity of the parasite species infecting these animals. IgG antibodies toEhrlichia spp., B. vogeli and L. infantum were found, respectively, in 39 (65%, 49 (81.6% and 60 (100% of the sampled dogs. Twenty-seven (45%, fifty-four (90%, fifty-three (88.3%, two (3.3% and one (1.6% dog were positive, respectively, forE. canis, Leishmania spp., Leishmania donovani complex, Babesia sp. and Anaplasma sp. in PCR assays. After sequencing, the amplicons showed 99% of identity with E. canis, B. vogeli, A. platys andLeishmania chagasi isolates. The findings of this study indicate that L. infantum-seropositive dogs from Campo Grande are exposed to multiple tick-borne pathogens, which should therefore be included in the differential diagnosis of dogs with clinical suspicion of leishmaniasis.

  7. Effect of Leishmania spp infection on the survival, life expectancy, fecundity and fertility of Lutzomyia longipalpiss.l. and Lutzomyia pseudolongipalpis

    Directory of Open Access Journals (Sweden)

    Irma Fatima Agrela

    2015-08-01

    Full Text Available We evaluated the effects of Leishmaniaspp infection on several population parameters of Lutzomyia longipalpis sensu lato andLutzomyia pseudolongipalpis, vectors of visceral leishmaniasis in Venezuela, under experimental conditions during the first post-feeding period. Females of both species were allowed to feed and engorge on a suspension of fresh washed human red blood cells in foetal calf serum. These blood cells were either non-infected or infected with one of the fourLeishmaniaspp strains and were offered through a chicken skin membrane. The longevity, life expectancy and the fecundity of uninfected flies were similar in both species, but the fertility was significantly lower in uninfected Lu. longipalpis females. In all cases, the infection of Lu. longipalpis and Lu. pseudolongipalpis by the Leishmaniastrains resulted in significant detrimental effects, which exerted a fitness cost expressed by reduced survival and life expectancy, as well as decreased fertility and fecundity compared with the control groups. Nevertheless, differences in these parameters were observed between these vector species depending on whether they were infected with the autochthonous Venezuelan Leishmania infantum strain (NESA or the Brazilian reference strain (PP75. The experimental data obtained agree with field data on the natural infection of these vector species and the significance of this scenario is discussed.

  8. An experimental protocol for the establishment of dogs with long-term cellular immune reactions to Leishmania antigens

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    Márcia Cristina Aquino Teixeira

    2011-03-01

    Full Text Available Domestic dogs are considered to be the main reservoirs of zoonotic visceral leishmaniasis. In this work, we evaluated a protocol to induce Leishmania infantum/Leishmania chagasi-specific cellular and humoral immune responses in dogs, which consisted of two injections of Leishmania promastigote lysate followed by a subcutaneous inoculation of viable promastigotes. The primary objective was to establish a canine experimental model to provide positive controls for testing immune responses to Leishmania in laboratory conditions. After inoculation of viable promastigotes, specific proliferative responses of peripheral blood mononuclear cells (PBMCs to either Leishmania lysate or recombinant proteins, the in vitro production of interferon-γ by antigen-stimulated PBMCs and a significant increase in circulating levels of anti-Leishmania antibodies were observed. The immunized dogs also displayed positive delayed-type hypersensitivity reactions to Leishmania crude antigens and to purified recombinant proteins. An important finding that supports the suitability of the dogs as positive controls is that they remained healthy for the entire observation period, i.e., more than seven years after infection. Following the Leishmania antigen lysate injections, the infection of dogs by the subcutaneous route appears to induce a sustained cellular immune response, leading to an asymptomatic infection. This provides a useful model for both the selection of immunogenic Leishmania antigens and for immunobiological studies on their possible immunoprotective activities.

  9. Progress towards a Leishmania vaccine.

    Science.gov (United States)

    Tabbara, Khaled S

    2006-07-01

    Leishmaniasis is a vector-born protozoan disease. Approximately 12 million individuals are affected worldwide with an estimated annual incidence of 1.5-2 million. Two clinical manifestations are recognized, cutaneous, and visceral, both of which are common in the Middle East. In both forms, infection is chronic, with potential deformities, persistence following cure, and lifelong risk of reactivation. Attempts to develop an effective human Leishmania vaccine have not yet succeeded. Leishmanization, a crude form of live vaccination historically originated in this part of the world. Experimental vaccination has been extensively studied in model animals in the past 2 decades. In this review, major human killed vaccine trials are surveyed, and modern trends in Leishmania vaccine development, including subunit vaccines, naked DNA vaccines, and transmission blocking vaccines are explored. Recent findings of a link between persistence of live parasites, and maintenance of long-term immunity suggest live vaccination with attenuated strains, as a future vaccination strategy.

  10. Phylogenomic reconstruction supports supercontinent origins for Leishmania.

    Science.gov (United States)

    Harkins, Kelly M; Schwartz, Rachel S; Cartwright, Reed A; Stone, Anne C

    2016-03-01

    Leishmania, a genus of parasites transmitted to human hosts and mammalian/reptilian reservoirs by an insect vector, is the causative agent of the human disease complex leishmaniasis. The evolutionary relationships within the genus Leishmania and its origins are the source of ongoing debate, reflected in conflicting phylogenetic and biogeographic reconstructions. This study employs a recently described bioinformatics method, SISRS, to identify over 200,000 informative sites across the genome from newly sequenced and publicly available Leishmania data. This dataset is used to reconstruct the evolutionary relationships of this genus. Additionally, we constructed a large multi-gene dataset, using it to reconstruct the phylogeny and estimate divergence dates for species. We conclude that the genus Leishmania evolved at least 90-100 million years ago, supporting a modified version of the Multiple Origins hypothesis that we call the Supercontinent hypothesis. According to this scenario, separate Leishmania clades emerged prior to, and during, the breakup of Gondwana. Additionally, we confirm that reptile-infecting Leishmania are derived from mammalian forms and that the species that infect porcupines and sloths form a clade long separated from other species. Finally, we firmly place the guinea-pig infecting species, Leishmaniaenriettii, the globally dispersed Leishmaniasiamensis, and the newly identified Australian species from a kangaroo, as sibling species whose distribution arises from the ancient connection between Australia, Antarctica, and South America. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. Involvement of Leishmania donovani major surface glycoprotein ...

    Indian Academy of Sciences (India)

    The major surface glycoprotein gp63 of the kinetoplastid protozoal parasite Leishmania is implicated as a ligand mediating uptake of the parasite into, and survival within, the host macrophage. By expressing gp63 antisense RNA from an episomal vector in L. donovani promastigotes, gp63-deficient transfectants were ...

  12. The proliferation potential of promastigotes of the main Leishmania species of the old world in NNN culture medium prepared using blood of four different mammals.

    Science.gov (United States)

    Ladopoulos, Theodoros; Ntais, Pantelis; Tsirigotakis, Nikolaos; Dokianakis, Emmanouil; Antoniou, Maria

    2015-10-01

    The efficacy of the in vitro cultivation of promastigotes of four Leishmania spp. was tested in the biphasic Novy-MacNeal-Nicolle (NNN) medium prepared using blood from different animals (horse, donkey, goat and sheep). The aim was to test which NNN preparation gave the best yield in the shortest time for different parasite species, in order to obtain a large crop of promastigotes for experimental work and for antigen preparation. Promastigotes of Leishmania infantum, Leishmania donovani, Leishmania tropica and Leishmania major, the four main parasite species occurring in the old world, were defrosted from -80 °C and placed, at equal numbers, in the 4 different NNN preparations. At the end of the 7th day, the NNN medium using horse blood produced the greatest number of promastigotes for all Leishmania spp. tested, whilst goat blood proved the poorest medium, providing culture results only for L. infantum. This finding may be explained by the fact that Leishmania is a nicotinamide adenine dinucleotide (NAD) auxotroph and horse erythrocytes support NAD-dependent microorganisms. Copyright © 2015 Elsevier Inc. All rights reserved.

  13. Intestinal parasites and vector-borne pathogens in stray and free-roaming cats living in continental and insular Greece

    Science.gov (United States)

    Diakou, Anastasia; Di Cesare, Angela; Accettura, Paolo Matteo; Barros, Luciano; Iorio, Raffaella; Paoletti, Barbara; Frangipane di Regalbono, Antonio; Halos, Lénaïg; Beugnet, Frederic; Traversa, Donato

    2017-01-01

    This survey investigated the distribution of various intestinal parasites and vector-borne pathogens in stray and free-roaming cats living in four regions of Greece. A total number of one hundred and fifty cats living in three Islands (Crete, Mykonos and Skopelos) and in Athens municipality was established as a realistic aim to be accomplished in the study areas. All cats were examined with different microscopic, serological and molecular assays aiming at evaluating the occurrence of intestinal parasites, and exposure to or presence of vector-borne infections. A total of 135 cats (90%) was positive for one or more parasites and/or pathogens transmitted by ectoparasites. Forty-four (29.3%) cats were positive for one single infection, while 91 (60.7%) for more than one pathogen. A high number of (n. 53) multiple infections caused by feline intestinal and vector-borne agents including at least one zoonotic pathogen was detected. Among them, the most frequently recorded helminths were roundworms (Toxocara cati, 24%) and Dipylidium caninum (2%), while a high number of examined animals (58.8%) had seroreaction for Bartonella spp., followed by Rickettsia spp. (43.2%) and Leishmania infantum (6.1%). DNA-based assays revealed the zoonotic arthropod-borne organisms Bartonella henselae, Bartonella clarridgeiae, Rickettsia spp., and L. infantum. These results show that free-ranging cats living in areas of Greece under examination may be exposed to a plethora of internal parasites and vector-borne pathogens, some of them potentially able to infect humans. Therefore, epidemiological vigilance and appropriate control measures are crucial for the prevention and control of these infections and to minimize the risk of infection for people. PMID:28141857

  14. Intestinal parasites and vector-borne pathogens in stray and free-roaming cats living in continental and insular Greece.

    Directory of Open Access Journals (Sweden)

    Anastasia Diakou

    2017-01-01

    Full Text Available This survey investigated the distribution of various intestinal parasites and vector-borne pathogens in stray and free-roaming cats living in four regions of Greece. A total number of one hundred and fifty cats living in three Islands (Crete, Mykonos and Skopelos and in Athens municipality was established as a realistic aim to be accomplished in the study areas. All cats were examined with different microscopic, serological and molecular assays aiming at evaluating the occurrence of intestinal parasites, and exposure to or presence of vector-borne infections. A total of 135 cats (90% was positive for one or more parasites and/or pathogens transmitted by ectoparasites. Forty-four (29.3% cats were positive for one single infection, while 91 (60.7% for more than one pathogen. A high number of (n. 53 multiple infections caused by feline intestinal and vector-borne agents including at least one zoonotic pathogen was detected. Among them, the most frequently recorded helminths were roundworms (Toxocara cati, 24% and Dipylidium caninum (2%, while a high number of examined animals (58.8% had seroreaction for Bartonella spp., followed by Rickettsia spp. (43.2% and Leishmania infantum (6.1%. DNA-based assays revealed the zoonotic arthropod-borne organisms Bartonella henselae, Bartonella clarridgeiae, Rickettsia spp., and L. infantum. These results show that free-ranging cats living in areas of Greece under examination may be exposed to a plethora of internal parasites and vector-borne pathogens, some of them potentially able to infect humans. Therefore, epidemiological vigilance and appropriate control measures are crucial for the prevention and control of these infections and to minimize the risk of infection for people.

  15. Intestinal parasites and vector-borne pathogens in stray and free-roaming cats living in continental and insular Greece.

    Science.gov (United States)

    Diakou, Anastasia; Di Cesare, Angela; Accettura, Paolo Matteo; Barros, Luciano; Iorio, Raffaella; Paoletti, Barbara; Frangipane di Regalbono, Antonio; Halos, Lénaïg; Beugnet, Frederic; Traversa, Donato

    2017-01-01

    This survey investigated the distribution of various intestinal parasites and vector-borne pathogens in stray and free-roaming cats living in four regions of Greece. A total number of one hundred and fifty cats living in three Islands (Crete, Mykonos and Skopelos) and in Athens municipality was established as a realistic aim to be accomplished in the study areas. All cats were examined with different microscopic, serological and molecular assays aiming at evaluating the occurrence of intestinal parasites, and exposure to or presence of vector-borne infections. A total of 135 cats (90%) was positive for one or more parasites and/or pathogens transmitted by ectoparasites. Forty-four (29.3%) cats were positive for one single infection, while 91 (60.7%) for more than one pathogen. A high number of (n. 53) multiple infections caused by feline intestinal and vector-borne agents including at least one zoonotic pathogen was detected. Among them, the most frequently recorded helminths were roundworms (Toxocara cati, 24%) and Dipylidium caninum (2%), while a high number of examined animals (58.8%) had seroreaction for Bartonella spp., followed by Rickettsia spp. (43.2%) and Leishmania infantum (6.1%). DNA-based assays revealed the zoonotic arthropod-borne organisms Bartonella henselae, Bartonella clarridgeiae, Rickettsia spp., and L. infantum. These results show that free-ranging cats living in areas of Greece under examination may be exposed to a plethora of internal parasites and vector-borne pathogens, some of them potentially able to infect humans. Therefore, epidemiological vigilance and appropriate control measures are crucial for the prevention and control of these infections and to minimize the risk of infection for people.

  16. Caracteres moleculares para la determinación taxonómica de tres especies de Lutzomyia (Diptera: Psychodidae, vectores potenciales de Leishmania presentes en el valle de Aburrá, Colombia

    Directory of Open Access Journals (Sweden)

    Alveiro PÉREZ-DORIA

    2008-01-01

    Full Text Available En Colombia están registradas 143 especies de Lutzomyia França, pero menos del 7% de éstas se encuentran incriminadas como vectores de Leishmania spp. Debido a la alta semejanza morfológica de algunas especies vectoras con otras no vectoras, se necesitan caracteres taxonómicos alternativos para identificar correctamente los flebotomíneos de cada zona geográfica del país. Con este objetivo, en el presente trabajo se secuenció el extremo 3’ del gen mitocondrial que codifica para la proteína citocromo b en tres vectores potenciales de Leishmania presentes en el valle de Aburrá, Colombia, Lutzomyia hartmanni (Fairchild y Hertig, L. columbiana (Ristorcelli y Van Ty y L. tihuiliensis Le Pont, Torrez-Espejo y Dujardin. A partir del alineamiento múltiple de nucleótidos se determinaron los sitios polimórficos, las distancias genéticas pareadas netas (p y la entropía. Las secuencias de nucleótidos fueron trasladadas a aminoácidos para estimar el número de sustituciones sinónimas y no sinónimas. En el alineamiento múltiple de 321 nucleótidos del gen citocromo b de L. columbiana, L. hartmanni y L. tihuiliensis se detectaron 83 sustituciones. En la secuencia parcial de la proteína se encontraron 18 reemplazos de aminoácidos. Las distancias genéticas interespecíficas fluctuaron en un rango mínimo de 0,137 entre L. tihuiliensis y L. columbiana, y un máximo de 0,215 entre L. columbiana y L. hartmanni. Los polimorfismos detectados en la secuencia de nucleótidos del gen y de aminoácidos de la proteína constituyen caracteres moleculares potencialmente útiles para la determinación taxonómica de estas especies de flebotomíneos.

  17. Detection of Leishmania spp in silvatic mammals and isolation of Leishmania (Viannia braziliensis from Rattus rattus in an endemic area for leishmaniasis in Minas Gerais State, Brazil.

    Directory of Open Access Journals (Sweden)

    Agnes Antônia Sampaio Pereira

    Full Text Available Knowledge of potential reservoirs of Leishmania spp. in an anthropic environment is important so that surveillance and control measures can be implemented. The aim of this study was to investigate the infection by Leishmania in small mammals in an area located in Minas Gerais, Brazil, that undergoes changes in its natural environment and presents autochthonous human cases of cutaneous leishmaniasis (CL and visceral leishmaniasis (VL. For the capture of the animals, Sherman and Tomahawk traps were used and distributed in the peridomicile of houses with reports of autochthonous cases of CL or VL. Six catches were carried out on two consecutive nights with intervals of two months during one year and samples of spleen, liver, tail skin, ear skin and bone marrow of the animals were obtained. Parasitological and molecular methods were used to detect the infection. Identification of the Leishmania species was performed by PCR RFLPhsp70. Twenty five animals of four species were captured: ten Rattus rattus, nine Didelphis albiventris, five Cerradomys subflavus and one Marmosops incanus. In the PCR-hsp70, five animals were positive (20%. The Leishmania species identified in PCR-RFLPhsp70 were: Leishmania braziliensis in D. albiventris (2, C. subflavus (1 and R. rattus (1 and Leishmania infantum in R. rattus (1. The highest positivity rate for L. braziliensis was obtained in the liver samples. The spleen was the only tissue positive for L. infantum. It was isolated in culture medium L. braziliensis from two samples (liver and spleen of R. rattus. This is the first record of isolation of L. braziliensis from R. rattus in the southeastern region of Brazil. These results are relevant to the knowledge of the epidemiology of leishmaniasis in the region, mainly in the investigation of the presence of hosts and possible reservoirs of the parasite.

  18. Cutaneous leishmaniasis in the central provinces of Hama and Edlib in Syria: Vector identification and parasite typing.

    Science.gov (United States)

    Haddad, Nabil; Saliba, Hanadi; Altawil, Atef; Villinsky, Jeffrey; Al-Nahhas, Samar

    2015-10-12

    Cutaneous leishmaniasis is a disease transmitted by sand fly bites. This disease is highly prevalent in Syria where Leishmania major and Leishmania tropica are the known aetiological agents. In 2011, more than 58,000 cases were reported in the country by the Ministry of Health. The central region of the country harbors 20 % of the reported cases. However, the epidemiology of the disease in this area is not well understood. An epidemiological survey was conducted in 2010 to identity the circulating parasite and the sand fly vector in the central provinces of Edlib and Hama. Sand fly specimens were collected using CDC light traps and identified morphologically. Total DNA was extracted from the abdomens of female specimens and from Giemsa-stained skin lesion smears of 80 patients. Leishmania parasites were first identified by sequencing the ITS1 gene amplicons. Then polymorphism analysis was performed using the RFLP technique. A total of 2142 sand flies were collected. They belonged to eight species, among which Phlebotomus sergenti and Phlebotomus papatasi were the most predominant. L. tropica ITS1 gene was amplified from two pools of P. sergenti specimens and from skin smears of cutaneous leishmaniasis patients. This suggests that P. sergenti is the potential vector species in the study area. The digestion profiles of the obtained amplicons by TaqI restriction enzyme were identical for all analysed L. tropica parasites. Moreover, L. infantum ITS1 gene was amplified from two pools of Phlebotomus tobbi in the relatively humid zone of Edlib. L. tropica is confirmed to be the aetiological agent of cutaneous leishmaniasis cases in the central provinces. RFLP technique failed to show any genetic heterogeneity in the ITS1 gene among the tested parasites. The molecular detection of this parasite in human skin smears and in P. sergenti supports the vector status of this species in the study area. The detection of L. infantum in P. tobbi specimens indicates a potential

  19. Molecular Identification of Leishmania spp. in Sand Flies (Diptera: Psychodidae: Phlebotominae) in the Lençóis Maranhenses National Park, Brazil.

    Science.gov (United States)

    Pereira-Filho, Adalberto Alves; Fonteles, Raquel Silva; Bandeira, Maria da Conceição Abreu; Moraes, Jorge Luiz Pinto; Rebêlo, José Manuel Macário; Melo, Maria Norma

    2018-02-20

    Sand flies are very common in the region of Lençóis Maranhenses National Park, an important tourist attraction in Brazil. However, the role of some species and their relative importance locally in Leishmania Ross 1903 transmission is unclear. The objective of this study was to identify Leishmania infection in phlebotomine sand flies collected around the Lençóis Maranhenses National Park, an important conservation area and popular international/national tourist destination with a high incidence of leishmaniasis. Sand flies were collected in peridomiciliary areas on the tourist route from September 2012 to August 2013. The captured females were subjected to molecular analyses for the detection of Leishmania DNA. Sand flies were infected with four Leishmania species: Leishmania (Viannia) braziliensis (Vianna, 1911) was found in Lutzomyia whitmani (Antunes and Coutinho, 1939) (2.1%) and Lutzomyia longipalpis (Lutz and Neiva, 1912) (1.7%); Leishmania (Leishmania) infantum (Nicole, 1908) infected Lutzomyia wellcomei (Fraiha, Shaw, and Lainson, 1971) (20%), Lutzomyia sordellii (Shannon and Del Ponte, 1927) (4.3%), Lu. longipalpis (3.7%), and Lu. whitmani (0.8%); Leishmania (Leishmania) amazonensis (Lainson & Shaw, 1972) was found in Lu. whitmani (0.58%), while Leishmania (Viannia) lainsoni infected Lutzomyia evandroi (Costa Lima and Antunes, 1936) (3.4%), Lu. longipalpis (1.06%), and Lu. whitmani (0.29%). The occurrence of these parasites requires control measures to reduce the incidence of cutaneous leishmaniasis and to contain a possible epidemic of visceral leishmaniasis, the most severe form of the disease.

  20. Genetic structure and evolution of the Leishmania genus in Africa and Eurasia: what does MLSA tell us.

    Science.gov (United States)

    El Baidouri, Fouad; Diancourt, Laure; Berry, Vincent; Chevenet, François; Pratlong, Francine; Marty, Pierre; Ravel, Christophe

    2013-01-01

    Leishmaniasis is a complex parasitic disease from a taxonomic, clinical and epidemiological point of view. The role of genetic exchanges has been questioned for over twenty years and their recent experimental demonstration along with the identification of interspecific hybrids in natura has revived this debate. After arguing that genetic exchanges were exceptional and did not contribute to Leishmania evolution, it is currently proposed that interspecific exchanges could be a major driving force for rapid adaptation to new reservoirs and vectors, expansion into new parasitic cycles and adaptation to new life conditions. To assess the existence of gene flows between species during evolution we used MLSA-based (MultiLocus Sequence Analysis) approach to analyze 222 Leishmania strains from Africa and Eurasia to accurately represent the genetic diversity of this genus. We observed a remarkable congruence of the phylogenetic signal and identified seven genetic clusters that include mainly independent lineages which are accumulating divergences without any sign of recent interspecific recombination. From a taxonomic point of view, the strong genetic structuration of the different species does not question the current classification, except for species that cause visceral forms of leishmaniasis (L. donovani, L. infantum and L. archibaldi). Although these taxa cause specific clinical forms of the disease and are maintained through different parasitic cycles, they are not clearly distinct and form a continuum, in line with the concept of species complex already suggested for this group thirty years ago. These results should have practical consequences concerning the molecular identification of parasites and the subsequent therapeutic management of the disease.

  1. Genetic Structure and Evolution of the Leishmania Genus in Africa and Eurasia: What Does MLSA Tell Us

    Science.gov (United States)

    El Baidouri, Fouad; Diancourt, Laure; Berry, Vincent; Chevenet, François; Pratlong, Francine; Marty, Pierre; Ravel, Christophe

    2013-01-01

    Leishmaniasis is a complex parasitic disease from a taxonomic, clinical and epidemiological point of view. The role of genetic exchanges has been questioned for over twenty years and their recent experimental demonstration along with the identification of interspecific hybrids in natura has revived this debate. After arguing that genetic exchanges were exceptional and did not contribute to Leishmania evolution, it is currently proposed that interspecific exchanges could be a major driving force for rapid adaptation to new reservoirs and vectors, expansion into new parasitic cycles and adaptation to new life conditions. To assess the existence of gene flows between species during evolution we used MLSA-based (MultiLocus Sequence Analysis) approach to analyze 222 Leishmania strains from Africa and Eurasia to accurately represent the genetic diversity of this genus. We observed a remarkable congruence of the phylogenetic signal and identified seven genetic clusters that include mainly independent lineages which are accumulating divergences without any sign of recent interspecific recombination. From a taxonomic point of view, the strong genetic structuration of the different species does not question the current classification, except for species that cause visceral forms of leishmaniasis (L. donovani, L. infantum and L. archibaldi). Although these taxa cause specific clinical forms of the disease and are maintained through different parasitic cycles, they are not clearly distinct and form a continuum, in line with the concept of species complex already suggested for this group thirty years ago. These results should have practical consequences concerning the molecular identification of parasites and the subsequent therapeutic management of the disease. PMID:23785530

  2. Estudio en condiciones de laboratorio de los ciclos de vida de Lutzomyia torvida y Lutzomyia longiflocosa (Diptera: Psychodidae posibles vectores de Leishmania braziliensis en la zona cafetera colombiana

    Directory of Open Access Journals (Sweden)

    Marisol Neira

    1998-12-01

    Full Text Available En el presente estudio, se hizo un seguimiento diario en condiciones de laboratorio del ciclo de vida de Lutzomyia torvida y Lutzomyia longiflocosa con el propósito de mejorar el conocimiento de la biología de estas dos especies del grupo verrucarum, serie townsendi. Cada una de ellas es la especie antropofílica más abundante en dos focos diferentes de Leishmania braziliensis localizados en el centro del país sobre la cordillera oriental. L. torvida fue capturada en Reventones (Cundinamarca y L. longiflocosa en Tello (Huila con cebo humano y trampa Shannon entre febrero de 1996 y marzo de 1997, principalmente en la época seca. La postura promedio de las hembras capturadas fue de 25,8 para L. torvida y 27,6 huevos para L. longiflocosa. El tiempo promedio de duración del ciclo de vida de huevo a adulto fue de 96,8 días para L. torvida y de 93,8 días para L. longiflocosa. Los mayores porcentajes de pérdida de individuos para las dos especies se presentaron en la fase de huevo y cuarto estadio larval. Aunque 54,8% de los insectos de L. torvida y 72% de L. longiflocosa lograron llegar a la fase adulta, fue dificil alimentar las hembras sobre hámster porque éstas se resistieron a picar.

  3. Lutzomyia (Pintomyia) fischeri (Diptera: Psychodidae: Phlebotominae), a probable vector of American cutaneous leishmaniasis: detection of natural infection by Leishmania (Viannia) DNA in specimens from the municipality of Porto Alegre (RS), Brazil, using multiplex PCR assay.

    Science.gov (United States)

    Pita-Pereira, Daniela de; Souza, Getúlio D; Pereira, Thaís de Araújo; Zwetsch, Adriana; Britto, Constança; Rangel, Elizabeth F

    2011-12-01

    In order to determine natural Leishmania (Viannia) infection in Lutzomyia (Pintomyia) fischeri, a multiplex PCR methodology coupled to non-isotopic hybridization was adopted for the analysis of sand fly samples collected by CDC light traps in an endemic area of American Cutaneous Leishmaniasis (ACL) in the periurban region of the municipality of Porto Alegre, Rio Grande do Sul State, Brazil. We analyzed by PCR methodology 560 specimens of Lutzomyia (Pintomyia) fischeri (520 females and 40 males). The wild sand flies were grouped into 56 pools (52 females and 4 males) of 10 each, and positive results were detected in 2 of the 52 female pools, representing a minimum infection rate of 0.38% based on the presence of at least 1 infected insect in the pool. This result associated with some local evidence such as anthopophily, spatial distribution in accordance with the transmission area and human case incidence, suggests that L. (P.)fischeri may be considered as a secondary vector of ACL in the studied locality. Copyright © 2011 Elsevier B.V. All rights reserved.

  4. Cell migration induced by Leishmania (Leishmania) amazonensis, Leishmania (Leishmania) major and Leishmania (Viannia) braziliensis into the peritoneal cavity of BALB/c mice

    OpenAIRE

    DT Wakimoto; KV Gaspareto; TGV Silveira; MVC Lonardoni; SMA Aristides

    2010-01-01

    In American cutaneous leishmaniasis, the initial infection phase is characterized by recruitment of neutrophils and monocytes. The migration of these cells in response to the presence of Leishmania in the peritoneum of affected animals remains unclear. The objective of this study was to investigate cell migration to the peritoneum of BALB/c mice after infection with Leishmania (Leishmania) amazonensis, Leishmania (Viannia) braziliensis and Leishmania (Leishmania) major. Initially, Leishmania ...

  5. Molecular Detection of Leishmania DNA in Wild-Caught Phlebotomine Sand Flies (Diptera: Psychodidae) From a Cave in the State of Minas Gerais, Brazil.

    Science.gov (United States)

    Carvalho, G M L; Brazil, R P; Rêgo, F D; Ramos, M C N F; Zenóbio, A P L A; Andrade Filho, J D

    2017-01-01

    Leishmania spp. are distributed throughout the world, and different species are associated with varying degrees of disease severity. In Brazil, Leishmania transmission involves several species of phlebotomine sand flies that are closely associated with different parasites and reservoirs, and thereby giving rise to different transmission cycles. Infection occurs during the bloodmeals of sand flies obtained from a variety of wild and domestic animals, and sometimes from humans. The present study focused on detection of Leishmania DNA in phlebotomine sand flies from a cave in the state of Minas Gerais. Detection of Leishmania in female sand flies was performed with ITS1 PCR-RFLP (internal transcribed spacer 1) using HaeIII enzyme and genetic sequencing for SSUrRNA target. The survey of Leishmania DNA was carried out on 232 pools and the parasite DNA was detected in four: one pool of Lutzomyia cavernicola (Costa Lima, 1932), infected with Le. infantum (ITS1 PCR-RFLP), two pools of Evandromyia sallesi (Galvão & Coutinho, 1939), both infected with Leishmania braziliensis complex (SSUrRNA genetic sequencing analysis), and one pool of Sciopemyia sordellii (Shannon & Del Ponte, 1927), infected with subgenus Leishmania (SSUrRNA genetic sequencing analysis). The present study identified the species for Leishmania DNA detected in four pools of sand flies, all of which were captured inside the cave. These results represent the first molecular detection of Lu cavernicola with Le infantum DNA, Sc sordellii with subgenus Leishmania DNA, and Ev sallesi with Leishmania braziliensis complex DNA. The infection rate in females captured for this study was 0.17%. © The Authors 2016. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  6. Bacterial and protozoal agents of feline vector-borne diseases in domestic and stray cats from southern Portugal

    Science.gov (United States)

    2014-01-01

    Background Feline vector-borne diseases (FVBD) have emerged in recent years, showing a wider geographic distribution and increased global prevalence. In addition to their veterinary importance, domestic cats play a central role in the transmission cycles of some FVBD agents by acting as reservoirs and sentinels, a circumstance that requires a One Health approach. The aim of the present work was to molecularly detect feline vector-borne bacteria and protozoa with veterinary and zoonotic importance, and to assess associated risk factors in cats from southern Portugal. Methods Six hundred and forty-nine cats (320 domestic and 329 stray), from veterinary medical centres and animal shelters in southern Portugal, were studied. Anaplasma spp./Ehrlichia spp., Babesia spp., Bartonella spp., Borrelia burgdorferi sensu lato, Hepatozoon spp. and Leishmania spp. infections were evaluated by polymerase chain reaction (PCR) in blood samples. Results One hundred and ninety-four (29.9%) cats were PCR-positive to at least one of the tested genera or complex of FVBD agents. Sixty-four (9.9%) cats were positive to Leishmania spp., 56 (8.6%) to Hepatozoon spp., 43 (6.6%) to Babesia spp., 35 (5.4%) to Anaplasma spp./Ehrlichia spp., 19 (2.9%) to Bartonella spp. and 14 (2.2%) to B. burgdorferi s.l. Thirty-three (5.1%) cats were positive to two (n = 29) or three (n = 4) genera/complex. Babesia vogeli, Bartonella clarridgeiae, Bartonella henselae, Ehrlichia canis, Hepatozoon felis and Leishmania infantum were identified by DNA sequencing. Conclusions The occurrence of FVBD agents in southern Portugal, some of them with zoonotic character, emphasizes the need to alert the veterinary community, owners and public health authorities for the risk of infection. Control measures should be implemented to prevent the infection of cats, other vertebrate hosts and people. PMID:24655431

  7. Imported and travelling dogs as carriers of canine vector-borne pathogens in Germany

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    Lorentz Susanne

    2010-04-01

    Full Text Available Abstract Background With the import of pets and pets taken abroad, arthropod-borne diseases have increased in frequency in German veterinary practices. This is reflected by 4,681 dogs that have been either travelled to or relocated from endemic areas to Germany. The case history of these dogs and the laboratory findings have been compared with samples collected from 331 dogs living in an endemic area in Portugal. The various pathogens and the seroprevalences were examined to determine the occurrence of, and thus infection risk, for vector-borne pathogens in popular travel destinations. Results 4,681 dogs were examined serological for Leishmania infantum, Babesia canis and Ehrlichia canis. Buffy coats were detected for Hepatozoon canis and blood samples were examined for microfilariae via the Knott's test. The samples were sent in from animal welfare organizations or private persons via veterinary clinics. Upon individual requests, dogs were additionally examined serological for Anaplasma phagocytophilum, Borrelia burgdorferi and Rickettsia conorii. Overall B. canis was the most prevalent pathogen detected by antibody titers (23.4%, followed by L. infantum (12.2% and E. canis (10.1%. Microfilariae were detected in 7.7% and H. canis in 2.7% of the examined dogs. In 332/1862 dogs A. phagocytophilum, in 64/212 B. burgdorferi and in 20/58 R. conorii was detected. Of the 4,681 dogs, in total 4,226 were imported to Germany from endemic areas. Eighty seven dogs joined their owners for a vacation abroad. In comparison to the laboratory data from Germany, we examined 331 dogs from Portugal. The prevalence of antibodies/pathogens we detected was: 62.8% to R. conorii, 58% to B. canis, 30.5% to A. phagocytophilum, 24.8% to E. canis, 21.1% to H. canis (via PCR, 9.1% to L. infantum and 5.3% to microfilariae. Conclusions The examination of 4,681 dogs living in Germany showed pathogens like L. infantum that are non-endemic in Germany. Furthermore, the German

  8. In Vitro Inhibitory Effect of Berberis vulgaris (Berberidaceae) and Its Main Component, Berberine against Different Leishmania Species.

    Science.gov (United States)

    Mahmoudvand, Hossein; Sharififar, Fariba; Sharifi, Iraj; Ezatpour, Behrouz; Fasihi Harandi, Majid; Makki, Mahsa Sadat; Zia-Ali, Naser; Jahanbakhsh, Sareh

    2014-03-01

    Leishmaniasis has been identified as a major public health problem in tropical and sub-tropical countries. The present study was aimed to investigate antileishmanial effects of various extracts of Berberis vulgaris also its active compoenent, berberine against Leishmania tropica and L. infantum species on in vitro experiments. In this study in vitro antileishmanial activity of various extracts of B. vulgaris also its active compoenent, berberine against promastigote and amastigote stages of L. tropica and L. infantum was evaluated, using MTT assay and in a macrophage model, respectively. Furthermore, infectivity rate and cytotoxicity effects of B. vulgaris and berberine in murine macrophage cells were investigated. The findings of optical density (OD) and IC50 indicated that B. vulgaris particulary berberine significantly (P<0.05) inhibited the growth rate of promastigote stage of L.tropica and L.infantum in comparison to meglumine antimoniate (MA). In addition, B. vulgaris and berberine significantly (P<0.05) decreased the mean number of amastigotes in each macrophage as compared with positive control. In the evaluation of cytotoxicity effects, it could be observed that berberine as compared with B. vulgaris exhibited more cytotoxicity against murine macrophages. Results also showed that when parasites were pre-incubated with B. vulgaris their ability to infect murine macrophages was significantly decreased. B.vulgaris particularly berberine exhibited potent in vitro leishmanicidal effects against L. tropica and L.infantum. Further works are required to evaluate the antileishmanial effects of B.vulgaris on Leishmania species using clinical settings.

  9. Ciclo de vida y colonización de Lutzomyia ovallesi (Diptera: Psychodidae, vector de Leishmania spp. en América Latina

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    Olga L. Cabrera

    1999-09-01

    Full Text Available Lutzomyia ovallesi (Diptera: Psychodidae, presente en un foco de Leishmania (V braziliensis en Reventones, Cundinamarca, reg- ión ubicada en el centro de Colombia. se crió y mantuvo por catorce generaciones bajo condiciones de laboratorio con una temperatura diaria entre 22-28 ""C y una humedad relativa entre 90 y 98%. Después de la emergencia, se llevaron machos y hembras a vasos de poliestireno (transparente y se mantuvieron en una nevera de icopor. La duración de cada uno de los estadios del ciclo de vida observado en las crías individuales fue: huevo 9-12 días (X = 10, 42, DE = 0,86; larva 17-11 días (X=9,9, DE=3,06; larva 11 7-13 días (X=9,5, DE= 3,06; larva III 4-12 (X=7,4, DE=1,50; larva IV 9-19 días (X=15,16, DE=2,66 y pupa 8-16 días (X=11, DE=1,89. El tiempo promedio para el desarrollo total del ciclo biológico fue de 63,36 días. La longevidad para las hembras presentó un rango entre 5 y 10 días (X=9,36 y para los machos entre 4 y 13 días (X=7,44. Con respecto a la productividad registrada a partir de un promedio de 30 hembras de Lutzomyia por vaso de cría, se obtuvo una producción mínima de 260 flebótomos en la quinta generación y una máxima de 4.51 1 en la décimotercera generación.

  10. Molecular crosstalks in Leishmania-sandfly-host relationships

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    Volf P.

    2008-09-01

    Full Text Available Sandflies (Diptera: Phlebotominae are vectors of Leishmania parasites, causative agents of important human and animal diseases with diverse manifestations. This review summarizes present knowledge about the vectorial part of Leishmania life cycle and parasite transmission to the vertebrate host. Particularly, it focuses on molecules that determine the establishment of parasite infection in sandfly midgut. It describes the concept of specific versus permissive sandfly vectors, explains the epidemiological consequences of broad susceptibility of permissive sandflies and demonstrates that genetic exchange may positively affect Leishmania fitness in the vector. Last but not least, the review describes recent knowledge about circulating antibodies produced by hosts in response to sandfly bites. Studies on specificity and kinetics of antibody response revealed that anti-saliva IgG could be used as a marker of host exposure to sandflies, i.e. as a useful tool for evaluation of vector control.

  11. Asymptomatic Leishmania Infected Children: A Seroprevalence and Molecular Survey in a Rural Area of Fars Province, Southern Iran

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    Akram Layegh Gigloo

    2018-01-01

    Full Text Available The current study aimed to evaluate the seroprevalence of visceral leishmaniasis in asymptomatic healthy children in a rural area of Fars province, Southern Iran. Blood samples were taken from 617 asymptomatic healthy children and serum samples along with buffy coat were separated from the blood. The serum samples were assessed for antibodies against Leishmania infantum by an indirect ELISA and the buffy coats were tested for the presence of L. infantum DNA by molecular method. Of the 617 recruited children, 297 (48.1% were female and 317 (51.4% were male. Anti-Leishmania antibodies were detected in 17 (2.8% of the children. From those 17 seropositive cases, 5 (29.4% were male and 12 (70.6% cases were female. Children aged 5–8 years had the highest seroprevalence rate; however, no associations were found between seropositivity to Leishmania and gender or age of the children. Moreover, L. infantum DNA was detected in buffy coat of 8 (1.3% of 617 children. Three of the PCR-positive cases were seropositive whereas 14 of seropositive subjects (82.3% were PCR-negative. Findings of the current study revealed a considerable subclinical leishmanial infection in children in the studied rural area in the south of Iran. Results of the current study could be used for surveillance, prevention, and control of VL in the area.

  12. Sand fly-Leishmania interactions: long relationships are not necessarily easy

    OpenAIRE

    Ramalho-Ortigao, Marcelo; Saraiva, Elvira M.; Traub-Csekö, Yara M.

    2010-01-01

    Sand fly and Leishmania are one of the best studied vector-parasite models. Much is known about the development of these parasites within the sand fly, and how transmission to a suitable vertebrate host takes place. Various molecules secreted by the vector assist the establishment of the infection in a vertebrate, and changes to the vector are promoted by the parasites in order to facilitate or enhance transmission. Despite a generally accepted view that sand flies and Leishmania are also one...

  13. IFN-γ, IL-2, IP-10, and MIG as Biomarkers of Exposure to Leishmania spp., and of Cure in Human Visceral Leishmaniasis

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    Ana V. Ibarra-Meneses

    2017-05-01

    Full Text Available New biomarkers are needed for monitoring the effectiveness of treatment for visceral leishmaniasis (VL. They might also improve the detection of the asymptomatic population in Leishmania-endemic areas. This paper examines the IL-2, IFN-γ, IFN-γ-induced protein 10 (IP-10, and monokine-induced-by-IFN-γ (MIG levels in whole blood—stimulated in vitro with soluble Leishmania antigen (SLA—taken from asymptomatic individuals and patients treated for VL living in a post-outbreak (Leishmania infantum area in Spain, and in an endemic (Leishmania donovani area of Bangladesh. IP-10 was found to be an accurate global marker of asymptomatic subjects with positive cellular/humoral tests, while MIG was found to be a better marker of contact with L. donovani than IL-2 but no for those with L. infantum. Determining IP-10, MIG, and IFN-γ levels proved useful in monitoring the cellular immune response following treatment for active disease caused by L. infantum.

  14. Natural infection of bats with Leishmania in Ethiopia.

    Science.gov (United States)

    Kassahun, Aysheshm; Sadlova, Jovana; Benda, Petr; Kostalova, Tatiana; Warburg, Alon; Hailu, Asrat; Baneth, Gad; Volf, Petr; Votypka, Jan

    2015-10-01

    The leishmaniases, a group of diseases with a worldwide-distribution, are caused by different species of Leishmania parasites. Both cutaneous and visceral leishmaniasis remain important public health problems in Ethiopia. Epidemiological cycles of these protozoans involve various sand fly (Diptera: Psychodidae) vectors and mammalian hosts, including humans. In recent years, Leishmania infections in bats have been reported in the New World countries endemic to leishmaniasis. The aim of this study was to survey natural Leishmania infection in bats collected from various regions of Ethiopia. Total DNA was isolated from spleens of 163 bats belonging to 23 species and 18 genera. Leishmania infection was detected by real-time (RT) PCR targeting a kinetoplast (k) DNA and internal transcribed spacer one (ITS1) gene of the parasite. Detection was confirmed by sequencing of the PCR products. Leishmania kDNA was detected in eight (4.9%) bats; four of them had been captured in the Aba-Roba and Awash-Methara regions that are endemic for leishmaniasis, while the other four specimens originated from non-endemic localities of Metu, Bedele and Masha. Leishmania isolates from two bats were confirmed by ITS1 PCR to be Leishmania tropica and Leishmania major, isolated from two individual bats, Cardioderma cor and Nycteris hispida, respectively. These results represent the first confirmed observation of natural infection of bats with the Old World Leishmania. Hence, bats should be considered putative hosts of Leishmania spp. affecting humans with a significant role in the transmission. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

  15. Arginase activity of Leishmania isolated from patients with cutaneous leishmaniasis.

    Science.gov (United States)

    Badirzadeh, A; Taheri, T; Abedi-Astaneh, F; Taslimi, Y; Abdossamadi, Z; Montakhab-Yeganeh, H; Aghashahi, M; Niyyati, M; Rafati, S

    2017-09-01

    Cutaneous leishmaniasis (CL) is one of the most important vector-borne parasitic diseases, highly endemic in Iran, and its prevalence is increasing all over the country. Arginase (ARG) activity in isolated Leishmania parasites from CL patients is yet to be explored. This study aimed to compare the ARG activity of isolated Leishmania promastigotes from CL patients with a standard strain of Leishmania major and its influences on the disease pathogenesis. We recruited 16 confirmed CL patients from Qom Province, in central Iran; after detection of Leishmania species using PCR-RFLP, we assessed the levels of ARG in the isolated promastigotes and determined the parasites' growth rate. Only L. major was identified from CL patients. The level of ARG activity in the isolated Leishmania promastigotes from CL patients was significantly higher than that obtained from the standard strain of L. major. No significant correlations between ARG activity and lesion size, number or duration were observed; in contrast, a significant negative correlation was seen between ARG level and Leishmania' growth rate. The obtained results suggest that increased ARG expression and activity in the isolated Leishmania promastigotes might contribute to the higher parasite infectivity and play a major role in the pathogenicity of the CL. © 2017 John Wiley & Sons Ltd.

  16. Molecular identification of Leishmania species in Taybad district, Iran

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    Salehi Ghodratollah

    2014-09-01

    Full Text Available Objective: To identify Leishmania species in patients with cutaneous leishmaniasis in the city of Taybad in Razavi Khorasan Province from April 2012 to March 2013. Methods: Among 52 persons who referred to Health Center of Taybad with suspected skin lesions, stained slide smears of 35 patients showed positive result for Leishmania. Also polymerase chain reaction assay performed using specific kDNA primers. Data of patients were analyzed with SPSS. Results: Of 35 positive smears for Leishmania, 21 (60% belonged to males and 14 (40% belonged to females. Polymerase chain reaction bands were observed in all 35 samples of which 31 (88.6% samples showed Leishmania tropica and 4 (11.4% showed Leishmania major. The highest infected age group was 11-20 years old. Conclusions: Both anthroponotic cutaneous leishmaniasis and zoonotic cutaneous leishmaniasis are present in Taybad. Leishmania tropica is the dominant causative species for anthroponotic cutaneous leishmaniasis. Further study is recommended to discover probable reservoir and vector for Leishmania major in Taybad.

  17. First Molecular Characterization of Leishmania Species Causing Visceral Leishmaniasis among Children in Yemen.

    Science.gov (United States)

    Mahdy, Mohammed A K; Al-Mekhlafi, Abdulsalam M; Abdul-Ghani, Rashad; Saif-Ali, Reyadh; Al-Mekhlafi, Hesham M; Al-Eryani, Samira M; Lim, Yvonne A L; Mahmud, Rohela

    2016-01-01

    Visceral leishmaniasis (VL) is a debilitating, often fatal disease caused by Leishmania donovani complex; however, it is a neglected tropical disease. L. donovani complex comprises two closely related species, L. donovani that is mostly anthroponotic and L. infantum that is zoonotic. Differentiation between these two species is critical due to the differences in their epidemiology and pathology. However, they cannot be differentiated morphologically, and their speciation using isoenzyme-based methods poses a difficult task and may be unreliable. Molecular characterization is now the most reliable method to differentiate between them and to determine their phylogenetic relationships. The present study aims to characterize Leishmania species isolated from bone marrows of Yemeni pediatric patients using sequence analysis of the ribosomal internal transcribed spacer-1 (ITS1) gene. Out of 41 isolates from Giemsa-stained bone marrow smears, 25 isolates were successfully amplified by nested polymerase chain reaction and sequenced in both directions. Phylogenetic analysis using neighbor joining method placed all study isolates in one cluster with L. donovani complex (99% bootstrap). The analysis of ITS1 for microsatellite repeat numbers identified L. infantum in 11 isolates and L. donovani in 14 isolates. These data suggest the possibility of both anthroponotic and zoonotic transmission of VL-causing Leishmania species in Yemen. Exploring the possible animal reservoir hosts is therefore needed for effective control to be achieved.

  18. Identification and biochemical characterization of Leishmania strains isolated in Peru, Mexico, and Spain.

    Science.gov (United States)

    Rodríguez-González, Isabel; Marín, Clotilde; Vargas, Franklin; Córdova, Ofelia; Barrera, Mario; Gutiérrez-Sánchez, Ramón; Alunda, Jose María; Sánchez-Moreno, Manuel

    2006-01-01

    Eight Leishmania promastigotes were isolated from different geographical areas: three (LP1, LP2, and LP3) from the provincial department La Libertad and the fourth (LP4) from the department of Cajamarca (northern Peru); another three (LM1, LM2, and LM3) in the province of Campeche (Mexico); and the last (LS1) from a clinical case of a dog in Madrid (Spain). The isolates were characterized by carbohydrate cell-surface residues using agglutinations with four purified lectins, by isoenzyme analysis using different isoenzymes, by analysis of kinetoplast DNA (kDNA) restriction fragment length polymorphism using four different restriction endonucleases and by the final metabolite patterns after in vitro culture. These isolates were compared with four reference strains and typified as: Leishmania (Leishmania) donovani, two strains of L. (L.) infantum, and one species of L. (Viania) peruviana. According to our results and the statistical study, the Peruvian isolates represent three different strains: one would be L. (V.) peruviana, another the strain isolated in Cajamarca (LP4) and the third would include the three strains from the department of La Libertad (LP1, LP2, and LP3), these latter three isolates being phylogenetically closer to the reference strain L. (L.) donovani. Meanwhile, the three isolates from Mexico form a group with close phylogenetic relationships to each other. The isolate from Spain belongs to the species L. (L.) infantum. Thus, a close correlation was drawn between the identity of each strain and its geographical origin.

  19. Recombinant antigens from Phlebotomus perniciosus saliva as markers of canine exposure to visceral leishmaniases vector.

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    Jan Drahota

    Full Text Available BACKGROUND: Phlebotomus perniciosus is the main vector in the western Mediterranean area of the protozoan parasite Leishmania infantum, the causative agent of canine and human visceral leishmaniases. Infected dogs serve as a reservoir of the disease, and therefore measuring the exposure of dogs to sand fly bites is important for estimating the risk of L. infantum transmission. In bitten hosts, sand fly saliva elicits a specific antibody response that reflects the intensity of sand fly exposure. As screening of specific anti-saliva antibodies is limited by the availability of salivary gland homogenates, utilization of recombinant salivary proteins is a promising alternative. In this manuscript we show for the first time the use of recombinant salivary proteins as a functional tool for detecting P. perniciosus bites in dogs. METHODOLOGY/PRINCIPAL FINDINGS: The reactivity of six bacterially-expressed recombinant salivary proteins of P. perniciosus, yellow-related protein rSP03B, apyrases rSP01B and rSP01, antigen 5-related rSP07, ParSP25-like protein rSP08 and D7-related protein rSP04, were tested with sera of mice and dogs experimentally bitten by this sand fly using immunoblots and ELISA. In the immunoblots, both mice and canine sera gave positive reactions with yellow-related protein, both apyrases and ParSP25-like protein. A similar reaction for recombinant salivary proteins was observed by ELISA, with the reactivity of yellow-related protein and apyrases significantly correlated with the antibody response of mice and dogs against the whole salivary gland homogenate. CONCLUSIONS/SIGNIFICANCE: Three recombinant salivary antigens of P. perniciosus, yellow-related protein rSP03B and the apyrases rSP01B and rSP01, were identified as the best candidates for evaluating the exposure of mice and dogs to P. perniciosus bites. Utilization of these proteins, or their combination, would be beneficial for screening canine sera in endemic areas of visceral

  20. Zoonotic intestinal parasites and vector-borne pathogens in Italian shelter and kennel dogs.

    Science.gov (United States)

    Traversa, Donato; Di Cesare, Angela; Simonato, Giulia; Cassini, Rudi; Merola, Carmine; Diakou, Anastasia; Halos, Lénaïg; Beugnet, Frederic; Frangipane di Regalbono, Antonio

    2017-04-01

    This study investigated the presence of zoonotic parasites and vector-borne pathogens in dogs housed in kennels and shelters from four sites of Italy. A total of 150 adoptable dogs was examined with different microscopic, serological and molecular methods. Overall 129 dogs (86%) were positive for one or more parasites and/or pathogens transmitted by ectoparasites. Forty-eight (32%) were positive for one infection, while 81 (54%) for more than one pathogen. The most common zoonotic helminths recorded were hookworms, roundworms and Capillaria aerophila, followed by mosquito-borne Dirofilaria spp. and Dipylidium caninum. One hundred and thirteen (77.9%), 6 (4.1%) and 2 (1.4%) dogs were positive for Rickettsia spp., Leishmania infantum and Anaplasma spp., respectively. The results show that dogs living in rescue facilities from the studied areas may be infected by many zoonotic internal parasites and vector-borne pathogens, and that control measures should be implemented. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. A atividade da azitromicina contra a Leishmania (Viannia) braziliensis e a Leishmania (Leishmania) amazonensis no modelo golden hamster

    OpenAIRE

    Sinagra, Ángel; Luna, Concepción; Abraham, David; Iannella, Maria del Carmen; Riarte, Adelina; Krolewiecki, Alejandro J.

    2007-01-01

    New therapeutic alternatives against leishmaniasis remain a priority. The activity of azithromycin against Leishmania (Leishmania) major has been previously demonstrated. Different responses among species of Leishmania make species-specific drug screening necessary. The activity of azithromycin against Leishmania (Viannia) braziliensis and Leishmania (Leishmania) amazonensis was evaluated in golden hamsters infected through footpad injections of metacyclic promastigotes, and compared with unt...

  2. The activity of azithromycin against Leishmania (Viannia) braziliensis and Leishmania (Leishmania) amazonensis in the golden hamster model

    OpenAIRE

    Sinagra,Ángel; Luna,Concepción; Abraham,David; Iannella,Maria del Carmen; Riarte,Adelina; Krolewiecki,Alejandro J.

    2007-01-01

    New therapeutic alternatives against leishmaniasis remain a priority. The activity of azithromycin against Leishmania (Leishmania) major has been previously demonstrated. Different responses among species of Leishmania make species-specific drug screening necessary. The activity of azithromycin against Leishmania (Viannia) braziliensis and Leishmania (Leishmania) amazonensis was evaluated in golden hamsters infected through footpad injections of metacyclic promastigotes, and compared with unt...

  3. Novel features of a PIWI-like protein homolog in the parasitic protozoan Leishmania.

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    Prasad K Padmanabhan

    Full Text Available In contrast to nearly all eukaryotes, the Old World Leishmania species L. infantum and L. major lack the bona fide RNAi machinery genes. Interestingly, both Leishmania genomes code for an atypical Argonaute-like protein that possesses a PIWI domain but lacks the PAZ domain found in Argonautes from RNAi proficient organisms. Using sub-cellular fractionation and confocal fluorescence microscopy, we show that unlike other eukaryotes, the PIWI-like protein is mainly localized in the single mitochondrion in Leishmania. To predict PIWI function, we generated a knockout mutant for the PIWI gene in both L. infantum (Lin and L. major species by double-targeted gene replacement. Depletion of PIWI has no effect on the viability of insect promastigote forms but leads to an important growth defect of the mammalian amastigote lifestage in vitro and significantly delays disease pathology in mice, consistent with a higher expression of the PIWI transcript in amastigotes. Moreover, amastigotes lacking PIWI display a higher sensitivity to apoptosis inducing agents than wild type parasites, suggesting that PIWI may be a sensor for apoptotic stimuli. Furthermore, a whole-genome DNA microarray analysis revealed that loss of LinPIWI in Leishmania amastigotes affects mostly the expression of specific subsets of developmentally regulated genes. Several transcripts encoding surface and membrane-bound proteins were found downregulated in the LinPIWI((-/- mutant whereas all histone transcripts were upregulated in the null mutant, supporting the possibility that PIWI plays a direct or indirect role in the stability of these transcripts. Although our data suggest that PIWI is not involved in the biogenesis or the stability of small noncoding RNAs, additional studies are required to gain further insights into the role of this protein on RNA regulation and amastigote development in Leishmania.

  4. Cytokine Release Assays as Tests for Exposure to Leishmania, and for Confirming Cure from Leishmaniasis, in Solid Organ Transplant Recipients.

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    Eugenia Carrillo

    Full Text Available Spain has one of the world's largest pools of organ donors and is a global leader in terms of the number of transplants it performs. The current outbreak of leishmaniasis in Fuenlabrada (in the southwest of the region of Madrid, Spain has involved 600 clinical cases since late 2009 (prevalence 0.2%. It may therefore be wise to monitor the town's transplanted population for Leishmania infantum; its members are immunosuppressed and at greater risk of infection and relapse following treatment. The present work examines the use of cytokine release assays to determine the prevalence of Leishmania infection in this population, and to confirm recovery following treatment for visceral leishmaniasis (VL. The humoral and cellular immune responses to L. infantum were characterized in 63 solid organ transplant (SOT recipients from Fuenlabrada, 57 of whom reported no previous episode of VL (NVL subjects, and six of whom had been cured of VL (CVL subjects. Seventeen subjects (12 NVL and 5 CVL showed a patent lymphoproliferative response to soluble Leishmania antigen (SLA. Stimulation of peripheral blood mononuclear cell cultures and of whole blood with SLA led to the production of different combinations of cytokines that might serve to confirm Leishmania infection or recovery from VL and help prevent cured patients from relapsing into this serious condition.

  5. Detection and identification of Leishmania spp.: application of two hsp70-based PCR-RFLP protocols to clinical samples from the New World.

    Science.gov (United States)

    Montalvo, Ana M; Fraga, Jorge; Tirado, Dídier; Blandón, Gustavo; Alba, Annia; Van der Auwera, Gert; Vélez, Iván Darío; Muskus, Carlos

    2017-07-01

    Leishmaniasis is highly prevalent in New World countries, where several methods are available for detection and identification of Leishmania spp. Two hsp70-based PCR protocols (PCR-N and PCR-F) and their corresponding restriction fragment length polymorphisms (RFLP) were applied for detection and identification of Leishmania spp. in clinical samples recruited in Colombia, Guatemala, and Honduras. A total of 93 cases were studied. The samples were classified into positive or suspected of leishmaniasis according to parasitological criteria. Molecular amplification of two different hsp70 gene fragments and further RFLP analysis for identification of Leishmania species was done. The detection in parasitologically positive samples was higher using PCR-N than PCR-F. In the total of samples studied, the main species identified were Leishmania panamensis, Leishmania braziliensis, and Leishmania infantum (chagasi). Although RFLP-N was more efficient for the identification, RFLP-F is necessary for discrimination between L. panamensis and Leishmania guyanesis, of great importance in Colombia. Unexpectedly, one sample from this country revealed an RFLP pattern corresponding to Leishmania naiffi. Both molecular variants are applicable for the study of clinical samples originated in Colombia, Honduras, and Guatemala. Choosing the better tool for each setting depends on the species circulating. More studies are needed to confirm the presence of L. naiffi in Colombian territory.

  6. Leishmania infection and blood food sources of phlebotomines in an area of Brazil endemic for visceral and tegumentary leishmaniasis

    Science.gov (United States)

    Guimarães-e-Silva, Antônia Suely; Silva, Soraia de Oliveira; Ribeiro da Silva, Rosa Cristina; Pinheiro, Valéria Cristina Soares; Rebêlo, José Manuel Macário; Melo, Maria Norma

    2017-01-01

    The aims of the study were to determine the blood feeding preferences of sandflies and to identify species of Leishmania that infected phlebotomines in Caxias, Maranhão, Brazil, an area that is highly endemic for leishmaniasis. Sandflies were captured in light traps located in the peridomiciliary environments of randomly selected houses in urban and rural settings between 1800 and 0600 hours on new moon days between March 2013 and February 2015. DNA extracts from 982 engorged female sandflies were submitted to fragment length polymorphism analysis to identify infecting species of Leishmania, and blood sources were identified for 778 of these specimens. Infection by Leishmania infantum was detected in Lutzomyia longipalpis, Lu. whitmani and Lu. termitophila; L. infantum/L. braziliensis in Lu. longipalpis, Lu. whitmani and Lu. trinidadensis; L. shawi in Lu. longipalpis; L. mexicana in Lu. longipalpis; L. braziliensis in Lu. longipalpis and Lu. whitmani; L. guyanensis in Lu. longipalpis and Lu. termitophila; L. amazonensis in Lu. longipalpis and L. lainsoni or L. naiffi in Lu. longipalpis, while Lu. longipalpis and Lu. trinidadensis were infected with unidentified Leishmania sp. Blood sources were identified in 573 individual phlebotomines and the preferred hosts were, in decreasing order, chicken, dog, rodent and human with lower preferences for pig, horse, opossum and cattle. Lu. longipalpis and Lu. whitmani performed mixed feeding on man, dog and rodent, while Lu. longipalpis was the most opportunistic species, feeding on the blood of all hosts surveyed, but preferably on dog/chicken, dog/rodent and rodent/chicken. Our findings reveal the concomitant circulation of Leishmania species that cause visceral leishmaniasis and tegumentary leishmaniasis in the study area, and explain the occurrence of autochthonous human cases of both clinical forms of leishmaniasis in Caxias, Maranhão. The results support our hypothesis that, in the municipality of Caxias, transmission

  7. Experimental treatment with sodium stibogluconate of hamsters infected with Leishmania (Leishmania) chagasi and Leishmania (Leishmania) amazonensis Tratamento experimental com stibogluconato de sódio em hamsters infectados com Leishmania (Leishmania) chagasi e Leishmania (Leishmania) amazonensis

    OpenAIRE

    Elizabeth M. de Figueiredo; Jaime Costa e Silva; Reginaldo P. Brazil

    1999-01-01

    The present paper reports the experimental treatment of hamsters infected with Leishmania chagasi and Leishmania amazonensis with sodium stibogluconate (20mg/kg/day x 20 days). Only with L. chagasi did the treatment result in the complete elimination of parasites from the spleen. However, no parasitological cure was achieved in hamsters infected with L. amazonensis.O presente trabalho é um relato do tratamento experimental de hamsters infectado com Leishmania chagasi e Leishmania amazonensis ...

  8. Detection, molecular typing and phylogenetic analysis of Leishmania isolated from cases of leishmaniasis among Syrian refugees in Lebanon

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    Tamara Salloum

    2016-06-01

    Two molecular typing methods of 39 FFPE Leishmania isolates were used: the ITS1-PCR RFLP and the nested ITS1-5.8S rDNA gene amplification followed by sequencing and phylogenetic analysis. The efficiency of these two techniques in Leishmania identification was compared and the phylogenetic relationships among these isolates were illustrated based on the neighbor-joining (NJ method. The results were statistically correlated with the parasitic index (PI. The DNA storage in formalin-fixed paraffin embedded (FFPE tissues was assessed as well. The parasites identified were all L. tropica as determined by both techniques. ITS1-5.8S rDNA gene based typing proved to be more sensitive in the detection of parasites (positive in 69.2% of the isolates as opposed to the ITS1-PCR RFLP method that was successful in identifying L. tropica in only 43.6% of the isolates. Sequencing and phylogenetic analysis revealed high levels of heterogeneity. A statistically significant correlation was observed between PI and the results of the nested ITS1-5.8S rDNA gene PCR. Genotyping at the species level is essential for monitoring the relative frequency of CL in the Mediterranean area that is correlated to three different Leishmania species (Leishmania infantum, Leishmania major and L. tropica, each characterized by distinct epidemiological features. The obtained results highlight the need to find a universally accepted diagnostic tool for Leishmania typing.

  9. Trichophoromyia auraensis is a putative vector

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    Carolina Bioni Garcia Teles

    Full Text Available The sandfly Trichophoromyia auraensis has recently evolved as a proven vector of Leishmania (Viannia endemic to state of Acre in the north of Brazil. This note is intended to propose a correction in the report of the first occurrence of natural infection of Leishmania (Viannia in this species. We and the other scientific groups reinforced that Tr. auraensis is a possible vector involved in the transmission of American cutaneous leishmaniasis in Acre, Brazil.

  10. Variabilidad genética en Lutzomyia ( verrucarum evansi (Núñez-Tovar, 1924, vector de Leishmaniosis visceral americana

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    Charles Porter

    2001-04-01

    Full Text Available

    Lutzomyia evansi (Núñez-Tovar, 1924, Lutzomyia longipalpis
    (Lutz y Neiva, 1912 y Lutzomyia cruzi (Mangabeira, 1938, son los
    vectores de Leishmania infantum Nicolle, 1908, en el neotrópico. Lu. evansi ha sido incriminada como vector en zonas rurales de la Costa Caribe Colombiana, y algunas zonas de Venezuela y Nicaragua. A pesar de que esta especie reviste gran importancia en Salud Pública, no existen a la fecha estudios sobre su variabilidad genética, desconociéndose si existe o no flujo genético entre las poblaciones rurales y urbanas, endémicas y no endémicas de leishmaniosis visceral (LV. Con base en los genes mitocondriales Citocromo b, RNA de transferencia para Serina, subunidades uno y cuatro de la NADH deshidrogenasa, se estudió la variabilidad genética entre las distintas poblaciones de Lu. evansi en la Costa Caribe, incluyendo la población
    geográficamente aislada de Isla Fuerte, y una población de Venezuela.

     

  11. Identification and phylogenetic relationship of Iranian strains of various Leishmania species isolated from cutaneous and visceral cases of leishmaniasis based on N-acetylglucosamine-1-phosphate transferase gene.

    Science.gov (United States)

    Hajjaran, Homa; Mohebali, Mehdi; Teimouri, Aref; Oshaghi, Mohammad Ali; Mirjalali, Hamed; Kazemi-Rad, Elham; Shiee, Mohammad Reza; Naddaf, Saied Reza

    2014-08-01

    The identity of Iranian Leishmania species has been resolved to some extent by some genetic markers. In this study, based on N-acetylglucosamine-1-phosphate transferase (nagt) gene, we further elucidated the identity and phylogeny of the prevalent species in this country. DNAs of 121 isolates belonging to cutaneous leishmaniasis (CL) patients, canine visceral leishmaniasis (CVL) cases, and Rhombomys opimus rodents were amplified by targeting a partial sequence of nagt gene. All the amplicons were analyzed with restriction fragment length polymorphism (RFLP) using Acc1 enzyme, and 49 amplicons representing different reservoir hosts were sequenced and aligned with similar sequences from GenBank database. The RFLP analysis revealed that 41 CL patients were infected Leishmania tropica and 36 with Leishmania major. Among 10 CVL isolates, 6 were identified as Leishmania infantum and 4 as L. tropica. Amongst 34 rodents' isolates, 11 and 23 isolates exhibited patterns similar to those of L. major, and L. tropica/Leishmania turanica, respectively. The sequencing results from all CL patients, CVL cases, and 4 reservoir rodents were in agreement with RFLP analysis and showed 99-100% homologies with the registered species of L. major, L. tropica, and L. infantum from Turkey, Tunisia, Iraq and Israel. Of the 7 rodent isolates exhibiting RFLP patterns similar to L. tropica/L. turanica, 3 exhibited the highest homologies (99-100%) with L. turanica and 4 with Leishmania gerbilli. The 49 nagt DNA sequences were grouped into five clusters representing L. major, L. tropica, L. infantum, L. turanica and L. gerbilli species, encompassing 19 haplotypes. No correlation was observed between intraspecies divergence and geographic distribution of haplotypes. The L. tropica haplotypes exhibited more homologies with those of L. infantum than L. major (97.2% vs. 96.9%), a probable indication to the potential ability of L. tropica to visceralize. Characterization of Iranian Leishmania isolates

  12. Genetic Diversity and Phylogenetic Analysis of the Iranian Leishmania Parasites Based on HSP70 Gene PCR-RFLP and Sequence Analysis.

    Science.gov (United States)

    Nemati, Sara; Fazaeli, Asghar; Hajjaran, Homa; Khamesipour, Ali; Anbaran, Mohsen Falahati; Bozorgomid, Arezoo; Zarei, Fatah

    2017-08-01

    Despite the broad distribution of leishmaniasis among Iranians and animals across the country, little is known about the genetic characteristics of the causative agents. Applying both HSP70 PCR-RFLP and sequence analyses, this study aimed to evaluate the genetic diversity and phylogenetic relationships among Leishmania spp. isolated from Iranian endemic foci and available reference strains. A total of 36 Leishmania isolates from almost all districts across the country were genetically analyzed for the HSP70 gene using both PCR-RFLP and sequence analysis. The original HSP70 gene sequences were aligned along with homologous Leishmania sequences retrieved from NCBI, and subjected to the phylogenetic analysis. Basic parameters of genetic diversity were also estimated. The HSP70 PCR-RFLP presented 3 different electrophoretic patterns, with no further intraspecific variation, corresponding to 3 Leishmania species available in the country, L. tropica, L. major, and L. infantum. Phylogenetic analyses presented 5 major clades, corresponding to 5 species complexes. Iranian lineages, including L. major, L. tropica, and L. infantum, were distributed among 3 complexes L. major, L. tropica, and L. donovani. However, within the L. major and L. donovani species complexes, the HSP70 phylogeny was not able to distinguish clearly between the L. major and L. turanica isolates, and between the L. infantum, L. donovani, and L. chagasi isolates, respectively. Our results indicated that both HSP70 PCR-RFLP and sequence analyses are medically applicable tools for identification of Leishmania species in Iranian patients. However, the reduced genetic diversity of the target gene makes it inevitable that its phylogeny only resolves the major groups, namely, the species complexes.

  13. Gluconeogenesis in Leishmania mexicana

    Science.gov (United States)

    Rodriguez-Contreras, Dayana; Hamilton, Nicklas

    2014-01-01

    Gluconeogenesis is an active pathway in Leishmania amastigotes and is essential for their survival within the mammalian cells. However, our knowledge about this pathway in trypanosomatids is very limited. We investigated the role of glycerol kinase (GK), phosphoenolpyruvate carboxykinase (PEPCK), and pyruvate phosphate dikinase (PPDK) in gluconeogenesis by generating the respective Leishmania mexicana Δgk, Δpepck, and Δppdk null mutants. Our results demonstrated that indeed GK, PEPCK, and PPDK are key players in the gluconeogenesis pathway in Leishmania, although stage-specific differences in their contribution to this pathway were found. GK participates in the entry of glycerol in promastigotes and amastigotes; PEPCK participates in the entry of aspartate in promastigotes, and PPDK is involved in the entry of alanine in amastigotes. Furthermore, the majority of alanine enters into the pathway via decarboxylation of pyruvate in promastigotes, whereas pathway redundancy is suggested for the entry of aspartate in amastigotes. Interestingly, we also found that l-lactate, an abundant glucogenic precursor in mammals, was used by Leishmania amastigotes to synthesize mannogen, entering the pathway through PPDK. On the basis of these new results, we propose a revision in the current model of gluconeogenesis in Leishmania, emphasizing the differences between amastigotes and promastigotes. This work underlines the importance of studying the trypanosomatid intracellular life cycle stages to gain a better understanding of the pathologies caused in humans. PMID:25288791

  14. Computational prediction of protein-protein interactions in Leishmania predicted proteomes.

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    Antonio M Rezende

    Full Text Available The Trypanosomatids parasites Leishmania braziliensis, Leishmania major and Leishmania infantum are important human pathogens. Despite of years of study and genome availability, effective vaccine has not been developed yet, and the chemotherapy is highly toxic. Therefore, it is clear just interdisciplinary integrated studies will have success in trying to search new targets for developing of vaccines and drugs. An essential part of this rationale is related to protein-protein interaction network (PPI study which can provide a better understanding of complex protein interactions in biological system. Thus, we modeled PPIs for Trypanosomatids through computational methods using sequence comparison against public database of protein or domain interaction for interaction prediction (Interolog Mapping and developed a dedicated combined system score to address the predictions robustness. The confidence evaluation of network prediction approach was addressed using gold standard positive and negative datasets and the AUC value obtained was 0.94. As result, 39,420, 43,531 and 45,235 interactions were predicted for L. braziliensis, L. major and L. infantum respectively. For each predicted network the top 20 proteins were ranked by MCC topological index. In addition, information related with immunological potential, degree of protein sequence conservation among orthologs and degree of identity compared to proteins of potential parasite hosts was integrated. This information integration provides a better understanding and usefulness of the predicted networks that can be valuable to select new potential biological targets for drug and vaccine development. Network modularity which is a key when one is interested in destabilizing the PPIs for drug or vaccine purposes along with multiple alignments of the predicted PPIs were performed revealing patterns associated with protein turnover. In addition, around 50% of hypothetical protein present in the networks

  15. Coinfection of Leishmania chagasi with Toxoplasma gondii, Feline Immunodeficiency Virus (FIV) and Feline Leukemia Virus (FeLV) in cats from an endemic area of zoonotic visceral leishmaniasis.

    Science.gov (United States)

    Sobrinho, Ludmila Silva Vicente; Rossi, Cláudio Nazaretian; Vides, Juliana Peloi; Braga, Eveline Tozzi; Gomes, Ana Amélia Domingues; de Lima, Valéria Marçal Félix; Perri, Sílvia Helena Venturoli; Generoso, Diego; Langoni, Hélio; Leutenegger, Christian; Biondo, Alexander Welker; Laurenti, Márcia Dalastra; Marcondes, Mary

    2012-06-08

    The aim of the present study was to determine the coinfection of Leishmania sp. with Toxoplasma gondii, Feline Immunodeficiency Virus (FIV) and Feline Leukemia Virus (FeLV) in a population of cats from an endemic area for zoonotic visceral leishmaniasis. An overall 66/302 (21.85%) cats were found positive for Leishmania sp., with infection determined by direct parasitological examination in 30/302 (9.93%), by serology in 46/302 (15.23%) and by both in 10/302 (3.31%) cats. Real time PCR followed by amplicon sequencing successfully confirmed Leishmania infantum (syn Leishmania chagasi) infection. Out of the Leishmania infected cats, coinfection with FIV was observed in 12/66 (18.18%), with T. gondii in 17/66 (25.75%) and with both agents in 5/66 (7.58%) cats. FeLV was found only in a single adult cat with no Leishmania infection. A positive association was observed in coinfection of Leishmania and FIV (p0.05). In conclusion, cats living in endemic areas of visceral leishmaniasis are significantly more likely to be coinfected with FIV, which may present confounding clinical signs and therefore cats in such areas should be always carefully screened for coinfections. Copyright © 2012 Elsevier B.V. All rights reserved.

  16. An effective in vitro and in vivo antileishmanial activity and mechanism of action of 8-hydroxyquinoline against Leishmania species causing visceral and tegumentary leishmaniasis.

    Science.gov (United States)

    Costa Duarte, Mariana; dos Reis Lage, Letícia Martins; Lage, Daniela Pagliara; Mesquita, Juliana Tonini; Salles, Beatriz Cristina Silveira; Lavorato, Stefânia Neiva; Menezes-Souza, Daniel; Roatt, Bruno Mendes; Alves, Ricardo José; Tavares, Carlos Alberto Pereira; Tempone, André Gustavo; Coelho, Eduardo Antonio Ferraz

    2016-02-15

    The development of new therapeutic strategies to treat leishmaniasis has become a priority. In the present study, the antileishmanial activity of 8-hydroxyquinoline (8-HQN) was investigated against in vitro promastigotes and in vivo intra-macrophage amastigotes of three Leishmania species: Leishmania amazonensis, Leishmania infantum and Leishmania braziliensis. Studies were performed to establish the 50% Leishmania inhibitory concentration (IC50) of 8-HQN, as well as its 50% cytotoxic concentration (CC50) on murine macrophages and in human red blood cells. The inhibition of macrophages infection was also evaluated using parasites that were pre-treated with 8-HQN. The effects of this compound on nitric oxide (NO) production and in the mitochondrial membrane potential were also evaluated. Finally, the therapeutic efficacy of 8-HQN was assessed in a known murine model, L. amazonensis-chronically infected BALB/c mice. Our results showed that 8-HQN was effective against promastigote and amastigote stages of all tested Leishmania species, presenting a selectivity index of 328.0, 62.0 and 47.0 for L. amazonensis, L. infantum and L. braziliensis, respectively. It was effective in treating infected macrophages, as well as in preventing the infection of these cells using pre-treated parasites. In addition, 8-HQN caused an alteration in the mitochondrial membrane potential of the parasites. When administered at 10mg/kg body weight/day by subcutaneous route, this product was effective in reducing the lesion diameter, as well as the parasite load in evaluated tissues and organs of infected animals. The results showed the in vitro and in vivo efficacy of 8-HQN against three different Leishmania species causing tegumentary and/or visceral leishmaniasis, and it could well be used for future therapeutic optimization studies to treat leishmaniasis. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. The efficacy of 2-nitrovinylfuran derivatives against Leishmania in vitro and in vivo

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    Sergio Sifontes-Rodríguez

    2015-04-01

    Full Text Available Despite recent advances in the treatment of some forms of leishmaniasis, the available drugs are still far from ideal due to inefficacy, parasite resistance, toxicity and cost. The wide-spectrum antimicrobial activity of 2-nitrovinylfuran compounds has been described, as has their activity against Trichomonas vaginalis and other protozoa. Thus, the aim of this study was to test the antileishmanial activities of six 2-nitrovinylfurans in vitro and in a murine model of leishmaniasis. Minimum parasiticide concentration (MPC and 50% inhibitory concentration (IC50 values for these compounds against the promastigotes of Leishmania amazonensis, Leishmania infantum and Leishmania braziliensis were determined, as were the efficacies of two selected compounds in an experimental model of cutaneous leishmaniasis (CL caused by L. amazonensis in BALB/c mice. All of the compounds were active against the promastigotes of the three Leishmania species tested. IC50 and MPC values were in the ranges of 0.8-4.7 µM and 1.7-32 µM, respectively. The compounds 2-bromo-5-(2-bromo-2-nitrovinyl-furan (furvina and 2-bromo-5-(2-methyl-2-nitrovinyl-furan (UC245 also reduced lesion growth in vivo at a magnitude comparable to or higher than that achieved by amphotericin B treatment. The results demonstrate the potential of this class of compounds as antileishmanial agents and support the clinical testing of Dermofural(r (a furvina-containing antifungal ointment for the treatment of CL.

  18. In-silico Leishmania Target Selectivity of Antiparasitic Terpenoids

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    Ifedayo Victor Ogungbe

    2013-07-01

    Full Text Available Neglected Tropical Diseases (NTDs, like leishmaniasis, are major causes of mortality in resource-limited countries. The mortality associated with these diseases is largely due to fragile healthcare systems, lack of access to medicines, and resistance by the parasites to the few available drugs. Many antiparasitic plant-derived isoprenoids have been reported, and many of them have good in vitro activity against various forms of Leishmania spp. In this work, potential Leishmania biochemical targets of antiparasitic isoprenoids were studied in silico. Antiparasitic monoterpenoids selectively docked to L. infantum nicotinamidase, L. major uridine diphosphate-glucose pyrophosphorylase and methionyl t-RNA synthetase. The two protein targets selectively targeted by germacranolide sesquiterpenoids were L. major methionyl t-RNA synthetase and dihydroorotate dehydrogenase. Diterpenoids generally favored docking to L. mexicana glycerol-3-phosphate dehydrogenase. Limonoids also showed some selectivity for L. mexicana glycerol-3-phosphate dehydrogenase and L. major dihydroorotate dehydrogenase while withanolides docked more selectively with L. major uridine diphosphate-glucose pyrophosphorylase. The selectivity of the different classes of antiparasitic compounds for the protein targets considered in this work can be explored in fragment- and/or structure-based drug design towards the development of leads for new antileishmanial drugs.

  19. Further support for a palaearctic origin of Leishmania

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    Sara F Kerr

    2000-08-01

    Full Text Available The fossil record and systematics of murid rodents, reservoirs of zoonotic cutaneous leishmaniasis in the Palaearctic, Oriental, African, Nearctic and Neotropical, strongly support a Palaearctic origin of Leishmania. The fossil record and systematics of phlebotomine sand flies reinforce this idea. Interpretations of molecular data that place the origin of Leishmania in the Neotropical are inconsistent with the natural histories of reservoirs and vectors. The evolutionary pattern of New World rats (Sigmodontinae indicates that they may be the most important reservoirs of zoonotic cutaneous leishmaniasis throughout their range.

  20. The past, present, and future of Leishmania genomics and transcriptomics

    Science.gov (United States)

    Cantacessi, Cinzia; Dantas-Torres, Filipe; Nolan, Matthew J.; Otranto, Domenico

    2015-01-01

    It has been nearly 10 years since the completion of the first entire genome sequence of a Leishmania parasite. Genomic and transcriptomic analyses have advanced our understanding of the biology of Leishmania, and shed new light on the complex interactions occurring within the parasite–host–vector triangle. Here, we review these advances and examine potential avenues for translation of these discoveries into treatment and control programs. In addition, we argue for a strong need to explore how disease in dogs relates to that in humans, and how an improved understanding in line with the ‘One Health’ concept may open new avenues for the control of these devastating diseases. PMID:25638444

  1. Hepatozoon canis and Leishmania spp. coinfection in dogs diagnosed with visceral leishmaniasis

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    Fernanda Nazaré Morgado

    Full Text Available Abstract This study describes the occurrence of dogs naturally co-infected with Hepatozoon canis and two Leishmania species: L. infantum or L. braziliensis. Four dogs serologically diagnosed with Visceral Leishmaniasis were euthanized. Liver and spleen samples were collected for histopathological analysis and DNA isolation. H. canis meronts were observed in tissues from all four dogs. H. canis infection was confirmed by PCR followed by sequencing of a fragment of 18S rRNA gene. Leishmania detection and typing was confirmed by ITS1' PCR-RFLP and parasite burden was calculated using ssrRNA quantitative qPCR. A DPP - Dual Path platform test was performed. One out (Dog #2 of four animals was asymptomatic. Dogs #1 and #4 were infected by L. infantum and were DPP test positive. Dogs #2 and #3 were infected by L. braziliensis and were DPP test negative. Furthermore, visceral dissemination was observed in Dogs #2 and #3, since L. braziliensis was detected in liver and spleen samples. The visceral dissemination of L. braziliensis associated with systemic signs suggested that this co-infection could influence the parasite burden and disease progression.

  2. Hepatozoon canis and Leishmania spp. coinfection in dogs diagnosed with visceral leishmaniasis.

    Science.gov (United States)

    Morgado, Fernanda Nazaré; Cavalcanti, Amanda Dos Santos; Miranda, Luisa Helena de; O'Dwyer, Lúcia Helena; Silva, Maria Regina Lucas da; Menezes, Rodrigo Caldas; Andrade da Silva, Aurea Virgínia; Boité, Mariana Côrtes; Cupolillo, Elisa; Porrozzi, Renato

    2016-01-01

    This study describes the occurrence of dogs naturally co-infected with Hepatozoon canis and two Leishmania species: L. infantum or L. braziliensis. Four dogs serologically diagnosed with Visceral Leishmaniasis were euthanized. Liver and spleen samples were collected for histopathological analysis and DNA isolation. H. canis meronts were observed in tissues from all four dogs. H. canis infection was confirmed by PCR followed by sequencing of a fragment of 18S rRNA gene. Leishmania detection and typing was confirmed by ITS1' PCR-RFLP and parasite burden was calculated using ssrRNA quantitative qPCR. A DPP - Dual Path platform test was performed. One out (Dog #2) of four animals was asymptomatic. Dogs #1 and #4 were infected by L. infantum and were DPP test positive. Dogs #2 and #3 were infected by L. braziliensis and were DPP test negative. Furthermore, visceral dissemination was observed in Dogs #2 and #3, since L. braziliensis was detected in liver and spleen samples. The visceral dissemination of L. braziliensis associated with systemic signs suggested that this co-infection could influence the parasite burden and disease progression.

  3. Arrabidaea chica Hexanic Extract Induces Mitochondrion Damage and Peptidase Inhibition on Leishmania spp.

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    Igor A. Rodrigues

    2014-01-01

    Full Text Available Currently available leishmaniasis treatments are limited due to severe side effects. Arrabidaea chica is a medicinal plant used in Brazil against several diseases. In this study, we investigated the effects of 5 fractions obtained from the crude hexanic extract of A. chica against Leishmania amazonensis and L. infantum, as well as on the interaction of these parasites with host cells. Promastigotes were treated with several concentrations of the fractions obtained from A. chica for determination of their minimum inhibitory concentration (MIC. In addition, the effect of the most active fraction (B2 on parasite’s ultrastructure was analyzed by transmission electron microscopy. To evaluate the inhibitory activity of B2 fraction on Leishmania peptidases, parasites lysates were treated with the inhibitory and subinhibitory concentrations of the B2 fraction. The minimum inhibitory concentration of B2 fraction was 37.2 and 18.6 μg/mL for L. amazonensis and L. infantum, respectively. Important ultrastructural alterations as mitochondrial swelling with loss of matrix content and the presence of vesicles inside this organelle were observed in treated parasites. Moreover, B2 fraction was able to completely inhibit the peptidase activity of promastigotes at pH 5.5. The results presented here further support the use of A. chica as an interesting source of antileishmanial agents.

  4. Detection of Leishmania parasites in the blood of patients with isolated cutaneous leishmaniasis.

    Science.gov (United States)

    Nakkash-Chmaisse, Hania; Makki, Raja; Nahhas, Georges; Knio, Khouzama; Nuwayri-Salti, Nuha

    2011-07-01

    The consequences of the spread of Leishmania parasites to the blood from lesions in patients with cutaneous leishmaniasis are numerous. To assess the magnitude of this invasion we conducted the present study on patients referred to the American University of Beirut Medical Center for cutaneous leishmaniasis. Patients referred for the management of cutaneous leishmaniasis were included in the study. Skin and blood cultures for Leishmania were taken from these patients. One hundred sixty-two patients were proven to have cutaneous leishmaniasis by pathology; 52% were males and 44% females (gender information was missing for 4%). Patient age ranged from 5 months to 70 years. None of the patients had received treatment for Leishmania. We obtained parasite isolates from 85 patients (52.5%), proven by cultures from skin and blood/blood components. Interestingly, the parasite was isolated in the blood and blood components of 50 patients (30.9%). Isoenzyme analysis confirmed the fact that the organisms in blood and skin were the same; from the 28 isolates that were positive in both skin and blood, eight isolates were Leishmania major and two were Leishmania tropica. The remaining isolates, whether positive in both blood and skin or in either of these tissues, skin or blood and its products, were Leishmania infantum sensu lato. In the current study, the detection rate of parasites in the blood of patients with cutaneous leishmaniasis was high. This illustrates the invasive characteristic of the parasite that has escaped the skin. Testing should be considered in areas other than Lebanon, especially around the Mediterranean basin. Whether these findings support the administration of systemic treatment for cutaneous leishmaniasis or not needs to be confirmed in larger prospective studies. Crown Copyright © 2011. Published by Elsevier Ltd. All rights reserved.

  5. In Vitro Inhibitory Effect of Berberis vulgaris (Berberidaceae and Its Main Component, Berberine against Different Leishmania Species.

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    Hossein Mahmoudvand

    2014-03-01

    Full Text Available Leishmaniasis has been identified as a major public health problem in tropical and sub-tropical countries. The present study was aimed to investigate antileishmanial effects of various extracts of Berberis vulgaris also its active compoenent, berberine against Leishmania tropica and L. infantum species on in vitro experiments.In this study in vitro antileishmanial activity of various extracts of B. vulgaris also its active compoenent, berberine against promastigote and amastigote stages of L. tropica and L. infantum was evaluated, using MTT assay and in a macrophage model, respectively. Furthermore, infectivity rate and cytotoxicity effects of B. vulgaris and berberine in murine macrophage cells were investigated.The findings of optical density (OD and IC50 indicated that B. vulgaris particulary berberine significantly (P<0.05 inhibited the growth rate of promastigote stage of L.tropica and L.infantum in comparison to meglumine antimoniate (MA. In addition, B. vulgaris and berberine significantly (P<0.05 decreased the mean number of amastigotes in each macrophage as compared with positive control. In the evaluation of cytotoxicity effects, it could be observed that berberine as compared with B. vulgaris exhibited more cytotoxicity against murine macrophages. Results also showed that when parasites were pre-incubated with B. vulgaris their ability to infect murine macrophages was significantly decreased.B.vulgaris particularly berberine exhibited potent in vitro leishmanicidal effects against L. tropica and L.infantum. Further works are required to evaluate the antileishmanial effects of B.vulgaris on Leishmania species using clinical settings.

  6. Failure of Miltefosine Treatment in Two Dogs with Natural Leishmania infantum Infection

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    Daniela Proverbio

    2014-01-01

    Full Text Available Two dogs, with naturally acquired canine leishmaniasis, were treated orally with miltefosine (2 mg/kg q 24 hr and allopurinol (10 mg/kg q 12 hr for 28 days. Both dogs showed good initial response to therapy, with reduction in clinical signs and improvement of clinicopathological changes. However, in both dogs, clinical and clinicopathological abnormalities recurred 150 days after initial treatment and a second course of miltefosine and allopurinol was administered. One dog failed to respond to the 2nd cycle of miltefosine treatment and the other dog responded initially but suffered an early relapse. Treatment with meglumine antimoniate (100 mg/kg q 24 hr for a minimum of 4 weeks was then started in both dogs. Both dogs showed rapid clinical and clinicopathological improvement and to date they have not received further treatment for 420 and 270 days, respectively. In view of the low number of antileishmanial drugs available and the fact that some of these are used in human as well as veterinary medicine, it is of paramount importance that drug resistance is monitored and documented.

  7. In vitro anti-Leishmania infantum activity of essential oil from Piper angustifolium

    OpenAIRE

    Bosquiroli, Lauriane S.S.; Demarque, Daniel P.; Rizk, Yasmin S.; Cunha, Marillin C.; Marques, Maria Carolina S.; Matos, Maria de Fátima C.; Kadri, Mônica C.T.; Carollo, Carlos A.; Arruda, Carla C.P.

    2015-01-01

    Abstract Piper angustifolium Lam., Piperaceae, popularly known as "matito", "pimenta-de-macaco", "pimenta-longa" or "jagurandi" in Brazil, has been commonly used in the treatment of cutaneous leishmaniasis-associated lesions, but there are few studies on the activity against visceral leishmaniasis-associated species. This study demonstrates the first in vitro antileishmanial activity of the P. angustifolium essential oil, of which the phytochemical profile showed the presence of sesquiterpene...

  8. Localized mucosal leishmaniasis caused by Leishmania infantum mimicking cancer in the rhinolaryngeal region

    Directory of Open Access Journals (Sweden)

    Fernando Cobo

    2016-09-01

    Full Text Available The clinical, microbiological, and histopathological findings of six patients with mucosal leishmaniasis are reported. Five of these patients were Spanish with no history of travel abroad, while the other was from Bolivia but had lived in Spain for more than 5 years. Two patients had no underlying disease, while the other four had several other medical conditions. Lesions were located in the nose in three patients and in the larynx in the other three. Symptoms included difficulty in swallowing, nasal obstruction, dysphonia, and polypoid lesions mimicking cancer. The diagnosis was based on the identification of parasites, or on PCR assay or culture. Five patients were treated with liposomal amphotericin B and the other with antimonial compounds.

  9. The ability of T2/B4 primers to detect Leishmania infantum among ...

    African Journals Online (AJOL)

    SERVER

    2008-04-03

    Apr 3, 2008 ... Reverse Primer (5`-CCT CTC TTT TTT CNC TGT GC-3`). (Schönian et al., 1996),. B. Forward Primer (RV1) (5`-GTG GGG GAG GGG CGT TCT-3`) and Reverse Primer (RV2) (5`-ATT TTA CAC CAA CCC CCA GTT-. 3`) (Lachaud et al., 2002; Reale et al., 1999),. C. Forward Primer (T2) (5`-CGG CTT CGC ...

  10. Molecular characterization and functional analysis of pteridine reductase in wild-type and antimony-resistant Leishmania lines.

    Science.gov (United States)

    de Souza Moreira, Douglas; Ferreira, Rafael Fernandes; Murta, Silvane M F

    2016-01-01

    Pteridine reductase (PTR1) is an NADPH-dependent reductase that participates in the salvage of pteridines, which are essential to maintain growth of Leishmania. In this study, we performed the molecular characterization of ptr1 gene in wild-type (WTS) and SbIII-resistant (SbR) lines from Leishmania guyanensis (Lg), Leishmania amazonensis (La), Leishmania braziliensis (Lb) and Leishmania infantum (Li), evaluating the chromosomal location, mRNA levels of the ptr1 gene and PTR1 protein expression. PFGE results showed that the ptr1 gene is located in a 797 kb chromosomal band in all Leishmania lines analyzed. Interestingly, an additional chromosomal band of 1070 kb was observed only in LbSbR line. Northern blot results showed that the levels of ptr1 mRNA are increased in the LgSbR, LaSbR and LbSbR lines. Western blot assays using the polyclonal anti-LmPTR1 antibody demonstrated that PTR1 protein is more expressed in the LgSbR, LaSbR and LbSbR lines compared to their respective WTS counterparts. Nevertheless, no difference in the level of mRNA and protein was observed between the LiWTS and LiSbR lines. Functional analysis of PTR1 enzyme was performed to determine whether the overexpression of ptr1 gene in the WTS L. braziliensis and L. infantum lines would change the SbIII-resistance phenotype of transfected parasites. Western blot results showed that the expression level of PTR1 protein was increased in the transfected parasites compared to the non-transfected ones. IC50 analysis revealed that the overexpression of ptr1 gene in the WTS L. braziliensis line increased 2-fold the SbIII-resistance phenotype compared to the non-transfected counterpart. Furthermore, the overexpression of ptr1 gene in the WTS L. infantum line did not change the SbIII-resistance phenotype. These results suggest that the PTR1 enzyme may be implicated in the SbIII-resistance phenotype in L. braziliensis line. Copyright © 2015 Elsevier Inc. All rights reserved.

  11. High Resolution Melting Analysis Targeting hsp70 as a Fast and Efficient Method for the Discrimination of Leishmania Species.

    Science.gov (United States)

    Zampieri, Ricardo Andrade; Laranjeira-Silva, Maria Fernanda; Muxel, Sandra Marcia; Stocco de Lima, Ana Carolina; Shaw, Jeffrey Jon; Floeter-Winter, Lucile Maria

    2016-02-01

    Protozoan parasites of the genus Leishmania cause a large spectrum of clinical manifestations known as Leishmaniases. These diseases are increasingly important public health problems in many countries both within and outside endemic regions. Thus, an accurate differential diagnosis is extremely relevant for understanding epidemiological profiles and for the administration of the best therapeutic protocol. Exploring the High Resolution Melting (HRM) dissociation profiles of two amplicons using real time polymerase chain reaction (real-time PCR) targeting heat-shock protein 70 coding gene (hsp70) revealed differences that allowed the discrimination of genomic DNA samples of eight Leishmania species found in the Americas, including Leishmania (Leishmania) infantum chagasi, L. (L.) amazonensis, L. (L.) mexicana, L. (Viannia) lainsoni, L. (V.) braziliensis, L. (V.) guyanensis, L. (V.) naiffi and L. (V.) shawi, and three species found in Eurasia and Africa, including L. (L.) tropica, L. (L.) donovani and L. (L.) major. In addition, we tested DNA samples obtained from standard promastigote culture, naturally infected phlebotomines, experimentally infected mice and clinical human samples to validate the proposed protocol. HRM analysis of hsp70 amplicons is a fast and robust strategy that allowed for the detection and discrimination of all Leishmania species responsible for the Leishmaniases in Brazil and Eurasia/Africa with high sensitivity and accuracy. This method could detect less than one parasite per reaction, even in the presence of host DNA.

  12. High Resolution Melting Analysis Targeting hsp70 as a Fast and Efficient Method for the Discrimination of Leishmania Species.

    Directory of Open Access Journals (Sweden)

    Ricardo Andrade Zampieri

    2016-02-01

    Full Text Available Protozoan parasites of the genus Leishmania cause a large spectrum of clinical manifestations known as Leishmaniases. These diseases are increasingly important public health problems in many countries both within and outside endemic regions. Thus, an accurate differential diagnosis is extremely relevant for understanding epidemiological profiles and for the administration of the best therapeutic protocol.Exploring the High Resolution Melting (HRM dissociation profiles of two amplicons using real time polymerase chain reaction (real-time PCR targeting heat-shock protein 70 coding gene (hsp70 revealed differences that allowed the discrimination of genomic DNA samples of eight Leishmania species found in the Americas, including Leishmania (Leishmania infantum chagasi, L. (L. amazonensis, L. (L. mexicana, L. (Viannia lainsoni, L. (V. braziliensis, L. (V. guyanensis, L. (V. naiffi and L. (V. shawi, and three species found in Eurasia and Africa, including L. (L. tropica, L. (L. donovani and L. (L. major. In addition, we tested DNA samples obtained from standard promastigote culture, naturally infected phlebotomines, experimentally infected mice and clinical human samples to validate the proposed protocol.HRM analysis of hsp70 amplicons is a fast and robust strategy that allowed for the detection and discrimination of all Leishmania species responsible for the Leishmaniases in Brazil and Eurasia/Africa with high sensitivity and accuracy. This method could detect less than one parasite per reaction, even in the presence of host DNA.

  13. A Historical Overview of the Classification, Evolution, and Dispersion of Leishmania Parasites and Sandflies.

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    Mohammad Akhoundi

    2016-03-01

    Full Text Available The aim of this study is to describe the major evolutionary historical events among Leishmania, sandflies, and the associated animal reservoirs in detail, in accordance with the geographical evolution of the Earth, which has not been previously discussed on a large scale.Leishmania and sandfly classification has always been a controversial matter, and the increasing number of species currently described further complicates this issue. Despite several hypotheses on the origin, evolution, and distribution of Leishmania and sandflies in the Old and New World, no consistent agreement exists regarding dissemination of the actors that play roles in leishmaniasis. For this purpose, we present here three centuries of research on sandflies and Leishmania descriptions, as well as a complete description of Leishmania and sandfly fossils and the emergence date of each Leishmania and sandfly group during different geographical periods, from 550 million years ago until now. We discuss critically the different approaches that were used for Leishmana and sandfly classification and their synonymies, proposing an updated classification for each species of Leishmania and sandfly. We update information on the current distribution and dispersion of different species of Leishmania (53, sandflies (more than 800 at genus or subgenus level, and animal reservoirs in each of the following geographical ecozones: Palearctic, Nearctic, Neotropic, Afrotropical, Oriental, Malagasy, and Australian. We propose an updated list of the potential and proven sandfly vectors for each Leishmania species in the Old and New World. Finally, we address a classical question about digenetic Leishmania evolution: which was the first host, a vertebrate or an invertebrate?We propose an updated view of events that have played important roles in the geographical dispersion of sandflies, in relation to both the Leishmania species they transmit and the animal reservoirs of the parasites.

  14. A Historical Overview of the Classification, Evolution, and Dispersion of Leishmania Parasites and Sandflies

    Science.gov (United States)

    Akhoundi, Mohammad; Kuhls, Katrin; Cannet, Arnaud; Votýpka, Jan; Marty, Pierre; Delaunay, Pascal; Sereno, Denis

    2016-01-01

    Background The aim of this study is to describe the major evolutionary historical events among Leishmania, sandflies, and the associated animal reservoirs in detail, in accordance with the geographical evolution of the Earth, which has not been previously discussed on a large scale. Methodology and Principal Findings Leishmania and sandfly classification has always been a controversial matter, and the increasing number of species currently described further complicates this issue. Despite several hypotheses on the origin, evolution, and distribution of Leishmania and sandflies in the Old and New World, no consistent agreement exists regarding dissemination of the actors that play roles in leishmaniasis. For this purpose, we present here three centuries of research on sandflies and Leishmania descriptions, as well as a complete description of Leishmania and sandfly fossils and the emergence date of each Leishmania and sandfly group during different geographical periods, from 550 million years ago until now. We discuss critically the different approaches that were used for Leishmana and sandfly classification and their synonymies, proposing an updated classification for each species of Leishmania and sandfly. We update information on the current distribution and dispersion of different species of Leishmania (53), sandflies (more than 800 at genus or subgenus level), and animal reservoirs in each of the following geographical ecozones: Palearctic, Nearctic, Neotropic, Afrotropical, Oriental, Malagasy, and Australian. We propose an updated list of the potential and proven sandfly vectors for each Leishmania species in the Old and New World. Finally, we address a classical question about digenetic Leishmania evolution: which was the first host, a vertebrate or an invertebrate? Conclusions and Significance We propose an updated view of events that have played important roles in the geographical dispersion of sandflies, in relation to both the Leishmania species they

  15. A Historical Overview of the Classification, Evolution, and Dispersion of Leishmania Parasites and Sandflies.

    Science.gov (United States)

    Akhoundi, Mohammad; Kuhls, Katrin; Cannet, Arnaud; Votýpka, Jan; Marty, Pierre; Delaunay, Pascal; Sereno, Denis

    2016-03-01

    The aim of this study is to describe the major evolutionary historical events among Leishmania, sandflies, and the associated animal reservoirs in detail, in accordance with the geographical evolution of the Earth, which has not been previously discussed on a large scale. Leishmania and sandfly classification has always been a controversial matter, and the increasing number of species currently described further complicates this issue. Despite several hypotheses on the origin, evolution, and distribution of Leishmania and sandflies in the Old and New World, no consistent agreement exists regarding dissemination of the actors that play roles in leishmaniasis. For this purpose, we present here three centuries of research on sandflies and Leishmania descriptions, as well as a complete description of Leishmania and sandfly fossils and the emergence date of each Leishmania and sandfly group during different geographical periods, from 550 million years ago until now. We discuss critically the different approaches that were used for Leishmana and sandfly classification and their synonymies, proposing an updated classification for each species of Leishmania and sandfly. We update information on the current distribution and dispersion of different species of Leishmania (53), sandflies (more than 800 at genus or subgenus level), and animal reservoirs in each of the following geographical ecozones: Palearctic, Nearctic, Neotropic, Afrotropical, Oriental, Malagasy, and Australian. We propose an updated list of the potential and proven sandfly vectors for each Leishmania species in the Old and New World. Finally, we address a classical question about digenetic Leishmania evolution: which was the first host, a vertebrate or an invertebrate? We propose an updated view of events that have played important roles in the geographical dispersion of sandflies, in relation to both the Leishmania species they transmit and the animal reservoirs of the parasites.

  16. Leishmania (L.) mexicana Infected Bats in Mexico: Novel Potential Reservoirs

    Science.gov (United States)

    Berzunza-Cruz, Miriam; Rodríguez-Moreno, Ángel; Gutiérrez-Granados, Gabriel; González-Salazar, Constantino; Stephens, Christopher R.; Hidalgo-Mihart, Mircea; Marina, Carlos F.; Rebollar-Téllez, Eduardo A.; Bailón-Martínez, Dulce; Balcells, Cristina Domingo; Ibarra-Cerdeña, Carlos N.; Sánchez-Cordero, Víctor; Becker, Ingeborg

    2015-01-01

    Leishmania (Leishmania) mexicana causes cutaneous leishmaniasis, an endemic zoonosis affecting a growing number of patients in the southeastern states of Mexico. Some foci are found in shade-grown cocoa and coffee plantations, or near perennial forests that provide rich breeding grounds for the sand fly vectors, but also harbor a variety of bat species that live off the abundant fruits provided by these shade-giving trees. The close proximity between sand flies and bats makes their interaction feasible, yet bats infected with Leishmania (L.) mexicana have not been reported. Here we analyzed 420 bats from six states of Mexico that had reported patients with leishmaniasis. Tissues of bats, including skin, heart, liver and/or spleen were screened by PCR for Leishmania (L.) mexicana DNA. We found that 41 bats (9.77%), belonging to 13 species, showed positive PCR results in various tissues. The infected tissues showed no evidence of macroscopic lesions. Of the infected bats, 12 species were frugivorous, insectivorous or nectarivorous, and only one species was sanguivorous (Desmodus rotundus), and most of them belonged to the family Phyllostomidae. The eco-region where most of the infected bats were caught is the Gulf Coastal Plain of Chiapas and Tabasco. Through experimental infections of two Tadarida brasiliensis bats in captivity, we show that this species can harbor viable, infective Leishmania (L.) mexicana parasites that are capable of infecting BALB/c mice. We conclude that various species of bats belonging to the family Phyllostomidae are possible reservoir hosts for Leishmania (L.) mexicana, if it can be shown that such bats are infective for the sand fly vector. Further studies are needed to determine how these bats become infected, how long the parasite remains viable inside these potential hosts and whether they are infective to sand flies to fully evaluate their impact on disease epidemiology. PMID:25629729

  17. Leishmania (L. mexicana infected bats in Mexico: novel potential reservoirs.

    Directory of Open Access Journals (Sweden)

    Miriam Berzunza-Cruz

    2015-01-01

    Full Text Available Leishmania (Leishmania mexicana causes cutaneous leishmaniasis, an endemic zoonosis affecting a growing number of patients in the southeastern states of Mexico. Some foci are found in shade-grown cocoa and coffee plantations, or near perennial forests that provide rich breeding grounds for the sand fly vectors, but also harbor a variety of bat species that live off the abundant fruits provided by these shade-giving trees. The close proximity between sand flies and bats makes their interaction feasible, yet bats infected with Leishmania (L. mexicana have not been reported. Here we analyzed 420 bats from six states of Mexico that had reported patients with leishmaniasis. Tissues of bats, including skin, heart, liver and/or spleen were screened by PCR for Leishmania (L. mexicana DNA. We found that 41 bats (9.77%, belonging to 13 species, showed positive PCR results in various tissues. The infected tissues showed no evidence of macroscopic lesions. Of the infected bats, 12 species were frugivorous, insectivorous or nectarivorous, and only one species was sanguivorous (Desmodus rotundus, and most of them belonged to the family Phyllostomidae. The eco-region where most of the infected bats were caught is the Gulf Coastal Plain of Chiapas and Tabasco. Through experimental infections of two Tadarida brasiliensis bats in captivity, we show that this species can harbor viable, infective Leishmania (L. mexicana parasites that are capable of infecting BALB/c mice. We conclude that various species of bats belonging to the family Phyllostomidae are possible reservoir hosts for Leishmania (L. mexicana, if it can be shown that such bats are infective for the sand fly vector. Further studies are needed to determine how these bats become infected, how long the parasite remains viable inside these potential hosts and whether they are infective to sand flies to fully evaluate their impact on disease epidemiology.

  18. Impact of phlebotomine sand flies on U.S. military operations at Tallil Air Base, Iraq: 4. Detection and identification of leishmania parasites in sand flies.

    Science.gov (United States)

    Coleman, Russell E; Hochberg, Lisa P; Swanson, Katherine I; Lee, John S; McAvin, James C; Moulton, John K; Eddington, David O; Groebner, Jennifer L; O'Guinn, Monica L; Putnam, John L

    2009-05-01

    Sand flies collected between April 2003 and November 2004 at Tallil Air Base, Iraq, were evaluated for the presence of Leishmania parasites using a combination of a real-time Leishmania-generic polymerase chain reaction (PCR) assay and sequencing of a 360-bp fragment of the glucose-6-phosphate-isomerase (GPI) gene. A total of 2,505 pools containing 26,574 sand flies were tested using the real-time PCR assay. Leishmania DNA was initially detected in 536 pools; however, after extensive retesting with the real-time PCR assay, a total of 456 pools were considered positive and 80 were considered indeterminate. A total of 532 samples were evaluated for Leishmania GPI by sequencing, to include 439 PCR-positive samples, 80 PCR-indeterminate samples, and 13 PCR-negative samples. Leishmania GPI was detected in 284 samples that were sequenced, to include 281 (64%) of the PCR-positive samples and 3 (4%) of the PCR-indeterminate samples. Of the 284 sequences identified as Leishmania, 261 (91.9%) were L. tarentolae, 18 (6.3%) were L. donovani-complex parasites, 3 (1.1%) were L. tropica, and 2 were similar to both L. major and L. tropica. Minimum field infection rates were 0.09% for L. donovani-complex parasites, 0.02% for L. tropica, and 0.01% for the L. major/tropica-like parasite. Subsequent sequencing of a 600-bp region of the "Hyper" gene of 12 of the L. donovani-complex parasites showed that all 12 parasites were L. infantum. These data suggest that L. infantum was the primary leishmanial threat to U.S. military personnel deployed to Tallil Air Base. The implications of these findings are discussed.

  19. Phlebotomus sergenti in a Cutaneous Leishmaniasis Focus in Azilal Province (High Atlas, Morocco): Molecular Detection and Genotyping of Leishmania tropica, and Feeding Behavior

    OpenAIRE

    Ajaoud , Malika; Es-Sette , Nargys; Charrel , Rémi N.; Laamrani-Idrissi , Abderahmane; Nhammi , Haddou; Riyad , Myriam; Lemrani , Meryem

    2015-01-01

    International audience; Background Phlebotomus (Paraphlebotomus) sergenti is at least one of the confirmed vectors for the transmission of cutaneous leishmaniasis caused by Leishmania tropica and distributed widely in Morocco. This form of leishmaniasis is considered largely as anthroponotic, although dogs were found infected with Leishmania tropica, suggestive of zoonosis in some rural areas. Methodology and Findings This survey aimed at (i) studying the presence of Leishmania in field caugh...

  20. Development and evaluation of zinc phthalocyanine nanoemulsions for use in photodynamic therapy for Leishmania spp.

    Science.gov (United States)

    Betzler de Oliveira de Siqueira, Luciana; da Silva Cardoso, Verônica; Almeida Rodrigues, Igor; Lúcia Vazquez-Villa, Ana; Pereira dos Santos, Elisabete; da Costa Leal Ribeiro Guimarães, Bruno; dos Santos Cerqueira Coutinho, Cristal; Vermelho, Alane Beatriz; Ricci Junior, Eduardo

    2017-02-01

    Photodynamic therapy (PDT) combines light with photosensitizers (PS) for production of reactive oxygen species (ROS) that can kill infectious microorganisms such as bacteria, fungi and protozoa. The application of nanotechnology has enabled the advancement of PDT because many PS are insoluble in water, necessitating a nanocarrier as a physiologically acceptable carrier. Nanoemulsions are efficient nanocarriers for solubilizing liposoluble drugs, like the PS, in water. Cutaneous (CL) and mucocutaneous leishmaniasis (ML) are caused by different species of the genus Leishmania, transmitted to humans by sandfly bites. Parasites are hosted in skin macrophages producing ulcerative lesions. Thus, a topical treatment, effective and inexpensive, for CL and ML is preferable to systemic interventions. There are topical treatments like paromomycin and amphotericin B, but they have many local side effects or a very high cost, limiting their use. This work aimed to develop a zinc phthalocyanine (photosensitizer) oil-in-water nanoemulsion, essential clove oil and polymeric surfactant (Pluronic® F127) for the formulation of a topical delivery system for use in PDT against Leishmania amazonensis and Leishmania infantum. The nanoemulsion was produced by a high-energy method and characterized by size, polydispersity, morphology, pH, content and stability studies. The toxicity in the dark and the photobiological activity of the formulations were evaluated in vitro for Leishmania and macrophages. The formulation presented was pH compatible with topical use, approximately 30 nm in size, with a polydispersity index ≤0.1 and remained stable at room and refrigerator temperature during the stability study (60 days). The zinc phthalocyanine nanoemulsion is effective in PDT against Leishmania spp.; use against skin infections can be a future application of this topical formulation, avoiding the use of oral or injectable medications, decreasing systemic adverse effects.

  1. Evaluation of Live Recombinant Nonpathogenic Leishmania tarentolae Expressing Cysteine Proteinase and A2 Genes as a Candidate Vaccine against Experimental Canine Visceral Leishmaniasis

    Science.gov (United States)

    Shahbazi, Mehdi; Zahedifard, Farnaz; Taheri, Tahereh; Taslimi, Yasaman; Jamshidi, Shahram; Shirian, Sadegh; Mahdavi, Niousha; Hassankhani, Mehdi; Daneshbod, Yahya; Zarkesh-Esfahani, Sayyed Hamid; Papadopoulou, Barbara; Rafati, Sima

    2015-01-01

    Canine Visceral Leishmaniasis (CVL) is a major veterinary and public health problem caused by Leishmania infantum (L. infantum) in many endemic countries. It is a severe chronic disease with generalized parasite spread to the reticuloendothelial system, such as spleen, liver and bone marrow and is often fatal when left untreated. Control of VL in dogs would dramatically decrease infection pressure of L. infantum for humans, since dogs are the main domestic reservoir. In the past decade, various subunits and DNA antigens have been identified as potential vaccine candidates in experimental animal models, but none has been approved for human use so far. In this study, we vaccinated outbreed dogs with a prime-boost regimen based on recombinant L. tarentolae expressing the L. donovani A2 antigen along with cysteine proteinase genes (CPA and CPB without its unusual C-terminal extension (CPB-CTE) and evaluated its immunogenicity and protective immunity against L. infantum infectious challenge. We showed that vaccinated animals produced significantly higher levels of IgG2, but not IgG1, and also IFN-γ and TNF-α, but low IL-10 levels, before and after challenge as compared to control animals. Protection in dogs was also correlated with a strong DTH response and low parasite burden in the vaccinated group. Altogether, immunization with recombinant L. tarentolae A2-CPA-CPB-CTE was proven to be immunogenic and induced partial protection in dogs, hence representing a promising live vaccine candidate against CVL. PMID:26197085

  2. Evaluation of Live Recombinant Nonpathogenic Leishmania tarentolae Expressing Cysteine Proteinase and A2 Genes as a Candidate Vaccine against Experimental Canine Visceral Leishmaniasis.

    Directory of Open Access Journals (Sweden)

    Mehdi Shahbazi

    Full Text Available Canine Visceral Leishmaniasis (CVL is a major veterinary and public health problem caused by Leishmania infantum (L. infantum in many endemic countries. It is a severe chronic disease with generalized parasite spread to the reticuloendothelial system, such as spleen, liver and bone marrow and is often fatal when left untreated. Control of VL in dogs would dramatically decrease infection pressure of L. infantum for humans, since dogs are the main domestic reservoir. In the past decade, various subunits and DNA antigens have been identified as potential vaccine candidates in experimental animal models, but none has been approved for human use so far. In this study, we vaccinated outbreed dogs with a prime-boost regimen based on recombinant L. tarentolae expressing the L. donovani A2 antigen along with cysteine proteinase genes (CPA and CPB without its unusual C-terminal extension (CPB-CTE and evaluated its immunogenicity and protective immunity against L. infantum infectious challenge. We showed that vaccinated animals produced significantly higher levels of IgG2, but not IgG1, and also IFN-γ and TNF-α, but low IL-10 levels, before and after challenge as compared to control animals. Protection in dogs was also correlated with a strong DTH response and low parasite burden in the vaccinated group. Altogether, immunization with recombinant L. tarentolae A2-CPA-CPB-CTE was proven to be immunogenic and induced partial protection in dogs, hence representing a promising live vaccine candidate against CVL.

  3. Susceptibility of spiny rats (Proechimys semispinosus to Leishmania (Viannia panamensis and Leishmania (Leishmania chagasi

    Directory of Open Access Journals (Sweden)

    BL Travi

    2002-09-01

    Full Text Available The role of Proechimys semispinosus as reservoir of Leishmania (Viannia panamensis on the Colombian Pacific coast was experimentally evaluated. The susceptibility to L. chagasi also was assessed to determine the utility of this rodent as a model for studying reservoir characteristics in the laboratory. Wild-caught animals were screened for natural trypanosomatid infections, and negative individuals were inoculated intradermally (ID in the snout or feet with 10(7 promastigotes of L. panamensis. L. chagasi was inoculated intracardially (10(7 promastigotes or ID in the ear (10(8 promastigotes. PCR-hybridization showed that 15% of 33 spiny rats were naturally infected with L. Viannia sp. Animals experimentally infected with L. panamensis developed non-ulcerated lesions that disappeared by the 7th week post-infection (p.i. and became more resistant upon reinfection. Infectivity to sand flies was low (1/20-1/48 infected/fed flies and transient, and both culture and PCR-hybridization showed that L. panamensis was cleared by the 13th week p.i. Animals inoculated with L. chagasi became subclinically infected and were non-infective to sand flies. Transient infectivity to vectors of spiny rats infected with L. panamensis, combined with population characteristics, e.g., abundance, exploitation of degraded habitats and high reproductive rates, could make them epidemiologically suitable reservoirs.

  4. Efficacy of Recombinant Canine Distemper Virus Expressing Leishmania Antigen against Leishmania Challenge in Dogs.

    Directory of Open Access Journals (Sweden)

    Ryuichi Miura

    Full Text Available Canine distemper virus (CDV vaccination confers long-term protection against CDV reinfection. To investigate the utility of CDV as a polyvalent vaccine vector for Leishmania, we generated recombinant CDVs, based on an avirulent Yanaka strain, that expressed Leishmania antigens: LACK, TSA, or LmSTI1 (rCDV-LACK, rCDV-TSA, and rCDV-LmSTI1, respectively. Dogs immunized with rCDV-LACK were protected against challenge with lethal doses of virulent CDV, in the same way as the parental Yanaka strain. To evaluate the protective effects of the recombinant CDVs against cutaneous leishmaniasis in dogs, dogs were immunized with one recombinant CDV or a cocktail of three recombinant CDVs, before intradermal challenge (in the ears with infective-stage promastigotes of Leishmania major. Unvaccinated dogs showed increased nodules with ulcer formation after 3 weeks, whereas dogs immunized with rCDV-LACK showed markedly smaller nodules without ulceration. Although the rCDV-TSA- and rCDV-LmSTI1-immunized dogs showed little protection against L. major, the cocktail of three recombinant CDVs more effectively suppressed the progression of nodule formation than immunization with rCDV-LACK alone. These results indicate that recombinant CDV is suitable for use as a polyvalent live attenuated vaccine for protection against both CDV and L. major infections in dogs.

  5. Efficacy of Recombinant Canine Distemper Virus Expressing Leishmania Antigen against Leishmania Challenge in Dogs.

    Science.gov (United States)

    Miura, Ryuichi; Kooriyama, Takanori; Yoneda, Misako; Takenaka, Akiko; Doki, Miho; Goto, Yasuyuki; Sanjoba, Chizu; Endo, Yasuyuki; Fujiyuki, Tomoko; Sugai, Akihiro; Tsukiyama-Kohara, Kyoko; Matsumoto, Yoshitsugu; Sato, Hiroki; Kai, Chieko

    2015-01-01

    Canine distemper virus (CDV) vaccination confers long-term protection against CDV reinfection. To investigate the utility of CDV as a polyvalent vaccine vector for Leishmania, we generated recombinant CDVs, based on an avirulent Yanaka strain, that expressed Leishmania antigens: LACK, TSA, or LmSTI1 (rCDV-LACK, rCDV-TSA, and rCDV-LmSTI1, respectively). Dogs immunized with rCDV-LACK were protected against challenge with lethal doses of virulent CDV, in the same way as the parental Yanaka strain. To evaluate the protective effects of the recombinant CDVs against cutaneous leishmaniasis in dogs, dogs were immunized with one recombinant CDV or a cocktail of three recombinant CDVs, before intradermal challenge (in the ears) with infective-stage promastigotes of Leishmania major. Unvaccinated dogs showed increased nodules with ulcer formation after 3 weeks, whereas dogs immunized with rCDV-LACK showed markedly smaller nodules without ulceration. Although the rCDV-TSA- and rCDV-LmSTI1-immunized dogs showed little protection against L. major, the cocktail of three recombinant CDVs more effectively suppressed the progression of nodule formation than immunization with rCDV-LACK alone. These results indicate that recombinant CDV is suitable for use as a polyvalent live attenuated vaccine for protection against both CDV and L. major infections in dogs.

  6. The Leishmania nicotinamidase is essential for NAD+ production and parasite proliferation.

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    Gazanion, E; Garcia, D; Silvestre, R; Gérard, C; Guichou, J F; Labesse, G; Seveno, M; Cordeiro-Da-Silva, A; Ouaissi, A; Sereno, D; Vergnes, B

    2011-10-01

    NAD+ is a central cofactor that plays important roles in cellular metabolism and energy production in all living cells. Genomics-based reconstruction of NAD+ metabolism revealed that Leishmania protozoan parasites are NAD+ auxotrophs. Consequently, these parasites require assimilating NAD+ precursors (nicotinamide, nicotinic acid, nicotinamide riboside) from their host environment to synthesize NAD+ by a salvage pathway. Nicotinamidase is a key enzyme of this salvage pathway that catalyses conversion of nicotinamide (NAm) to nicotinic acid (Na), and that is absent in higher eukaryotes. We present here the biochemical and functional characterizations of the Leishmania infantum nicotinamidase (LiPNC1). Generation of Lipnc1 null mutants leads to a decrease in NAD+ content, associated with a metabolic shutdown-like phenotype with an extensive lag phase of growth. Both phenotypes could be rescued by an add-back construct or by addition of exogenous Na. In addition, Lipnc1 null mutants were unable to establish a sustained infection in a murine experimental model. Altogether, these results illustrate that NAD+ homeostasis is a fundamental component of Leishmania biology and virulence, and that NAm constitutes its main NAD+ source in the mammalian host. The crystal structure of LiPNC1 we solved allows now the design of rational inhibitors against this new promising therapeutic target. © 2011 Blackwell Publishing Ltd.

  7. Leishmania major infection in a dog with cutaneous manifestations

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    Baneth, Gad; Nachum-Biala, Yaarit; Shabat Simon, Maytal; Brenner, Ori; Gaier, Sarit; Rojas, Alicia; Yasur-Landau, Daniel

    2016-01-01

    Background Leishmania major is a main cause of cutaneous leishmaniasis in humans in an area that stretches from India through Central Asia, the Middle East, to North and West Africa. In Israel, it is a common infection of humans with rodents as the reservoir hosts and Phlebotomus papatasi as its sand fly vector. Findings A 6?months old spayed female mixed breed dog was referred to the Hebrew University Veterinary Teaching Hospital with a large ulcerative dermal lesion on the muzzle, and lesio...

  8. Integrated mapping of establishment risk for emerging vector-borne infections: a case study of canine leishmaniasis in southwest France.

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    Nienke Hartemink

    Full Text Available BACKGROUND: Zoonotic visceral leishmaniasis is endemic in the Mediterranean Basin, where the dog is the main reservoir host. The disease's causative agent, Leishmania infantum, is transmitted by blood-feeding female sandflies. This paper reports an integrative study of canine leishmaniasis in a region of France spanning the southwest Massif Central and the northeast Pyrenees, where the vectors are the sandflies Phlebotomus ariasi and P. perniciosus. METHODS: Sandflies were sampled in 2005 using sticky traps placed uniformly over an area of approximately 100 by 150 km. High- and low-resolution satellite data for the area were combined to construct a model of the sandfly data, which was then used to predict sandfly abundance throughout the area on a pixel by pixel basis (resolution of c. 1 km. Using literature- and expert-derived estimates of other variables and parameters, a spatially explicit R(0 map for leishmaniasis was constructed within a Geographical Information System. R(0 is a measure of the risk of establishment of a disease in an area, and it also correlates with the amount of control needed to stop transmission. CONCLUSIONS: To our knowledge, this is the first analysis that combines a vector abundance prediction model, based on remotely-sensed variables measured at different levels of spatial resolution, with a fully mechanistic process-based temperature-dependent R(0 model. The resulting maps should be considered as proofs-of-principle rather than as ready-to-use risk maps, since validation is currently not possible. The described approach, based on integrating several modeling methods, provides a useful new set of tools for the study of the risk of outbreaks of vector-borne diseases.

  9. Effects of trans-stilbene and terphenyl compounds on different strains of Leishmania and on cytokines production from infected macrophages.

    Science.gov (United States)

    Bruno, Federica; Castelli, Germano; Vitale, Fabrizio; Giacomini, Elisa; Roberti, Marinella; Colomba, Claudia; Cascio, Antonio; Tolomeo, Manlio

    2018-01-01

    Most of the antileishmanial modern therapies are not satisfactory due to high toxicity or emergence of resistance and high cost of treatment. Previously, we observed that two compounds of a small library of trans-stilbene and terphenyl derivatives, ST18 and TR4, presented the best activity and safety profiles against Leishmania infantum promastigotes and amastigotes. In the present study we evaluated the effects of ST18 and the TR4 in 6 different species of Leishmania and the modifications induced by these two compounds in the production of 8 different cytokines from infected macrophages. We observed that TR4 was potently active in all Leishmania species tested in the study showing a leishmanicidal activity higher than that of ST18 and meglumine antimoniate in the most of the species. Moreover, TR4 was able to decrease the levels of IL-10, a cytokine able to render the host macrophage inactive allowing the persistence of parasites inside its phagolysosome, and increase the levels of IL-1β, a cytokine important for host resistance to Leishmania infection by inducible iNOS-mediated production of NO, and IL-18, a cytokine implicated in the development of Th1-type immune response. Copyright © 2017 Elsevier Inc. All rights reserved.

  10. Characterization of Leishmania Parasites Isolated From Kala- azar Patients in Kohgiloyeh and Boyerahmad, Using Semi-Nested PCR

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    B Sarkari

    2006-04-01

    Full Text Available ABSTRACT: Introduction & Objective: Visceral leishmaniasis (VL is a disease commonly known as Kala-azar caused by protozoan parasites of the genus Leishmania including L. donovani, L. infantum and L. chagasi. VL is sporadic in many areas of Iran and is endemic in a few provinces such as Fars, Azarbayjan, Bushehr, Ardabil and Qom. VL has been reported from some areas of Kohgiloyeh and Boyerahmad and this study aimed to characterize the causative agent of VL in this region. Materials & Methods: Bone marrow sample was obtained from 6 VL patients from children department in Imam Sajad hospital in Yasuj. DNA was extracted from the obtained samples and was checked by semi-nested PCR to determine the species of the parasite. To do that, a segment of minicircle kinetoplast DNA was amplified, using LINR4 and LIN17 primers. Products of PCR were evaluated by electrophoresis, using 1.5% agarose and stained with ethidium bromide. Results: Parasitologically examination of bone marrow smears demonstrated amastigotes form of the parasite in the samples. For mass cultivation, isolated parasites were cultured in diphasic NNN followed by RPMI 1640 media. All the samples produced a 720 bp band in PCR assay. The isolates were compared with referent strains and it was revealed that all the isolates were L. infantum. Conclusion: Findings of this study demonstrated that the causative agent of VL in Kohgiloyeh and Boyerahmad was L. infantum. Further study is needed to explore other aspects of VL in this region.

  11. Assessment of PCR in the detection of Leishmania spp in experimentally infected individual phlebotomine sandflies (Diptera: Psychodidae: Phlebotominae

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    MICHALSKY Érika M.

    2002-01-01

    Full Text Available DNA amplification by the polymerase chain reaction (PCR was applied in the investigation of the presence of Leishmania (Kinetoplastida: Trypanosomatidae parasites in single phlebotomine sandflies. Three phlebotomine/parasite pairs were used: Lutzomyia longipalpis/Leishmania chagasi, Lutzomyia migonei/Leishmania amazonensis and Lutzomyia migonei/Leishmania braziliensis, all of them incriminated in the transmission of visceral or cutaneous leishmaniasis. DNA extraction was performed with whole insects, with no need of previous digestive tract dissection or pooling specimens. The presence of either mouse blood in the digestive tract of the sandflies or the digestive tract itself did not interfere in the PCR. Infection by as few as 10 Leishmania sp. per individual were sufficient for DNA amplification with genus-specific primers. Using primers for L. braziliensis and L. mexicana complexes, respectively, it was possible to discriminate between L. braziliensis and L. amazonensis in experimentally infected vectors (L. migonei.

  12. Innate Immunity against Leishmania Infections

    Science.gov (United States)

    Gurung, Prajwal; Kanneganti, Thirumala-Devi

    2015-01-01

    Leishmaniasis is a major health problem that affects more than 300 million people throughout the world. The morbidity associated with the disease causes serious economic burden in Leishmania endemic regions. Despite the morbidity and economic burden associated with Leishmaniasis, this disease rarely gets noticed and is still categorized under neglected tropical diseases. The lack of research combined with the ability of Leishmania to evade immune recognition has rendered our efforts to design therapeutic treatments or vaccines challenging. Herein, we review the literature on Leishmania from innate immune perspective and discuss potential problems as well as solutions and future directions that could aid in identifying novel therapeutic targets to eliminate this parasite. PMID:26249747

  13. Molecular Epidemiology for Vector Research on Leishmaniasis

    Science.gov (United States)

    Kato, Hirotomo; Gomez, Eduardo A; Cáceres, Abraham G; Uezato, Hiroshi; Mimori, Tatsuyuki; Hashiguchi, Yoshihisa

    2010-01-01

    Leishmaniasis is a protozoan disease caused by the genus Leishmania transmitted by female phlebotomine sand flies. Surveillance of the prevalence of Leishmania and responsive vector species in endemic and surrounding areas is important for predicting the risk and expansion of the disease. Molecular biological methods are now widely applied to epidemiological studies of infectious diseases including leishmaniasis. These techniques are used to detect natural infections of sand fly vectors with Leishmania protozoa and are becoming powerful tools due to their sensitivity and specificity. Recently, genetic analyses have been performed on sand fly species and genotyping using PCR-RFLP has been applied to the sand fly taxonomy. In addition, a molecular mass screening method has been established that enables both sand fly species and natural leishmanial infections to be identified simultaneously in hundreds of sand flies with limited effort. This paper reviews recent advances in the study of sand flies, vectors of leishmaniasis, using molecular biological approaches. PMID:20617005

  14. Molecular Epidemiology for Vector Research on Leishmaniasis

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    Hirotomo Kato

    2010-03-01

    Full Text Available Leishmaniasis is a protozoan disease caused by the genus Leishmania transmitted by female phlebotomine sand flies. Surveillance of the prevalence of Leishmania and responsive vector species in endemic and surrounding areas is important for predicting the risk and expansion of the disease. Molecular biological methods are now widely applied to epidemiological studies of infectious diseases including leishmaniasis. These techniques are used to detect natural infections of sand fly vectors with Leishmania protozoa and are becoming powerful tools due to their sensitivity and specificity. Recently, genetic analyses have been performed on sand fly species and genotyping using PCR-RFLP has been applied to the sand fly taxonomy. In addition, a molecular mass screening method has been established that enables both sand fly species and natural leishmanial infections to be identified simultaneously in hundreds of sand flies with limited effort. This paper reviews recent advances in the study of sand flies, vectors of leishmaniasis, using molecular biological approaches.

  15. Molecular epidemiology for vector research on leishmaniasis.

    Science.gov (United States)

    Kato, Hirotomo; Gomez, Eduardo A; Cáceres, Abraham G; Uezato, Hiroshi; Mimori, Tatsuyuki; Hashiguchi, Yoshihisa

    2010-03-01

    Leishmaniasis is a protozoan disease caused by the genus Leishmania transmitted by female phlebotomine sand flies. Surveillance of the prevalence of Leishmania and responsive vector species in endemic and surrounding areas is important for predicting the risk and expansion of the disease. Molecular biological methods are now widely applied to epidemiological studies of infectious diseases including leishmaniasis. These techniques are used to detect natural infections of sand fly vectors with Leishmania protozoa and are becoming powerful tools due to their sensitivity and specificity. Recently, genetic analyses have been performed on sand fly species and genotyping using PCR-RFLP has been applied to the sand fly taxonomy. In addition, a molecular mass screening method has been established that enables both sand fly species and natural leishmanial infections to be identified simultaneously in hundreds of sand flies with limited effort. This paper reviews recent advances in the study of sand flies, vectors of leishmaniasis, using molecular biological approaches.

  16. Analysis of the Antigenic and Prophylactic Properties of the Leishmania Translation Initiation Factors eIF2 and eIF2B in Natural and Experimental Leishmaniasis

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    Esther Garde

    2018-04-01

    Full Text Available Different members of intracellular protein families are recognized by the immune system of the vertebrate host infected by parasites of the genus Leishmania. Here, we have analyzed the antigenic and immunogenic properties of the Leishmania eIF2 and eIF2B translation initiation factors. An in silico search in Leishmania infantum sequence databases allowed the identification of the genes encoding the α, β, and γ subunits and the α, β, and δ subunits of the putative Leishmania orthologs of the eukaryotic initiation factors F2 (LieIF2 or F2B (LieIF2B, respectively. The antigenicity of these factors was analyzed by ELISA using recombinant versions of the different subunits. Antibodies against the different LieIF2 and LieIF2B subunits were found in the sera from human and canine visceral leishmaniasis patients, and also in the sera from hamsters experimentally infected with L. infantum. In L. infantum (BALB/c and Leishmania major (BALB/c or C57BL/6 challenged mice, a moderate humoral response against these protein factors was detected. Remarkably, these proteins elicited an IL-10 production by splenocytes derived from infected mice independently of the Leishmania species employed for experimental challenge. When DNA vaccines based on the expression of the LieIF2 or LieIF2B subunit encoding genes were administered in mice, an antigen-specific secretion of IFN-γ and IL-10 cytokines was observed. Furthermore, a partial protection against murine CL development due to L. major infection was generated in the vaccinated mice. Also, in this work we show that the LieIF2α subunit and the LieIF2Bβ and δ subunits have the capacity to stimulate IL-10 secretion by spleen cells from naïve mice. B-lymphocytes were identified as the major producers of this anti-inflammatory cytokine. Taking into account the data found in this study, it may be hypothesized that these proteins act as virulence factors implicated in the induction of humoral responses as well

  17. Analysis of the Antigenic and Prophylactic Properties of the Leishmania Translation Initiation Factors eIF2 and eIF2B in Natural and Experimental Leishmaniasis.

    Science.gov (United States)

    Garde, Esther; Ramírez, Laura; Corvo, Laura; Solana, José C; Martín, M Elena; González, Víctor M; Gómez-Nieto, Carlos; Barral, Aldina; Barral-Netto, Manoel; Requena, José M; Iborra, Salvador; Soto, Manuel

    2018-01-01

    Different members of intracellular protein families are recognized by the immune system of the vertebrate host infected by parasites of the genus Leishmania . Here, we have analyzed the antigenic and immunogenic properties of the Leishmania eIF2 and eIF2B translation initiation factors. An in silico search in Leishmania infantum sequence databases allowed the identification of the genes encoding the α, β, and γ subunits and the α, β, and δ subunits of the putative Leishmania orthologs of the eukaryotic initiation factors F2 (LieIF2) or F2B (LieIF2B), respectively. The antigenicity of these factors was analyzed by ELISA using recombinant versions of the different subunits. Antibodies against the different LieIF2 and LieIF2B subunits were found in the sera from human and canine visceral leishmaniasis patients, and also in the sera from hamsters experimentally infected with L. infantum . In L. infantum (BALB/c) and Leishmania major (BALB/c or C57BL/6) challenged mice, a moderate humoral response against these protein factors was detected. Remarkably, these proteins elicited an IL-10 production by splenocytes derived from infected mice independently of the Leishmania species employed for experimental challenge. When DNA vaccines based on the expression of the LieIF2 or LieIF2B subunit encoding genes were administered in mice, an antigen-specific secretion of IFN-γ and IL-10 cytokines was observed. Furthermore, a partial protection against murine CL development due to L. major infection was generated in the vaccinated mice. Also, in this work we show that the LieIF2α subunit and the LieIF2Bβ and δ subunits have the capacity to stimulate IL-10 secretion by spleen cells from naïve mice. B-lymphocytes were identified as the major producers of this anti-inflammatory cytokine. Taking into account the data found in this study, it may be hypothesized that these proteins act as virulence factors implicated in the induction of humoral responses as well as in the

  18. Sand fly captures with Disney traps in area of occurrence of Leishmania (Leishmania) amazonensis in the state of Mato Grosso do Sul, mid-western Brazil.

    Science.gov (United States)

    Dorval, Maria Elizabeth Cavalheiros; Alves, Tulia Peixoto; Cristaldo, Geucira; Rocha, Hilda Carlos da; Alves, Murilo Andrade; Oshiro, Elisa Teruya; Oliveira, Alessandra Gutierrez de; Brazil, Reginaldo Peçanha; Galati, Eunice Aparecida Bianchi; Cunha, Rivaldo Venancio da

    2010-01-01

    The work was conducted to study phlebotomine fauna (Diptera: Psychodidae) and aspects of American cutaneous leishmaniasis transmission in a forested area where Leishmania (Leishmania) amazonensis occurs, situated in the municipality of Bela Vista, State of Mato Grosso do Sul, Brazil. The captures were conducted with modified Disney traps, using hamster (Mesocricetus auratus) as bait, from May 2004 to January 2006. Ten species of phlebotomine sandflies were captured: Brumptomyia avellari, Brumptomyia brumpti, Bichromomyia flaviscutellata, Evandromyia bourrouli, Evandromyia lenti, Lutzomyia longipalpis, Psathyromyia campograndensis, Psathyromyia punctigeniculata, Psathyromyia shannoni and Sciopemyia sordellii. The two predominant species were Ev bourrouli (57.3%) and Bi flaviscutellata (41.4%), present at all sampling sites. Two of the 36 hamsters used as bait presented natural infection with Leishmania. The parasite was identified as Leishmania (Leishmania) amazonensis. Analysis of the results revealed the efficiency of Disney traps for capturing Bichromomyia flaviscutellata and the simultaneous presence of both vector and the Leishmania species transmitted by the same can be considered a predictive factor of the occurrence of leishmaniasis outbreaks for the human population that occupies the location.

  19. Molecular and serological detection of Leishmania spp. in captive wild animals from Ilha Solteira, SP, Brazil Detecção sorológica e molecular de Leishmania spp. em animais selvagens do zoológico de Ilha Solteira, SP, Brasil

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    Márcia Mariza Gomes Jusi

    2011-09-01

    Full Text Available Leishmaniasis is a zoonotic disease that affects 12 million people worldwide. Several mammalian species can serve as a reservoir for this disease. Dogs are the main reservoir for visceral leishmaniasis in urban areas, which has become a serious public health concern in Brazil. The aim of this study was to evaluate the presence of Leishmania spp. in captive wild animals from Ilha Solteira, São Paulo, Brazil. Blood and various tissues samples were collected from animals of five different species: Speothos venaticus, Chrysocyon brachyurus, Cerdocyon thous, Pseudalopex vetulus, and Procyon cancrivorus. Antibodies against Leishmania spp. were detected in three wild canids by indirect fluorescent antibody test (IFAT and enzyme-linked immunosorbent assay (ELISA. PCR analyses of blood and bone marrow from all animals were negative, but Leishmania DNA was found in the tissues and skin of seropositive animals. Positive PCR samples were also positive for Leishmania donovani complex. Analysis of sequenced PCR products showed similarities with different regions of Leishmania (Leishmania infantum and Leishmania (Leishmania chagasi kinetoplastids. Measures to control visceral leishmaniasis in wild animals kept in Brazilian zoos should be established, as no disease control programs are currently available.Leishmaniose é uma doença zoonótica que afeta cerca de 12 milhões de pessoas no mundo todo. Várias espécies mamíferas podem servir de reservatório para a doença. Os cães são considerados os principais reservatórios para a leishmaniose visceral em áreas urbanas, o que tem se tornado um sério problema de saúde pública no Brasil. O objetivo deste trabalho foi avaliar a presença de Leishmania spp. em animais selvagens mantidos no zoológico de Ilha Solteira, São Paulo, Brasil. Foram coletados amostras de sangue e tecidos de cinco espécies diferentes: Speothos venaticus, Chrysocyon brachyurus, Cerdocyon thous, Pseudalopex vetulus, e Procyon

  20. Genome sequencing of the lizard parasite Leishmania tarentolae reveals loss of genes associated to the intracellular stage of human pathogenic species

    Science.gov (United States)

    Raymond, Frédéric; Boisvert, Sébastien; Roy, Gaétan; Ritt, Jean-François; Légaré, Danielle; Isnard, Amandine; Stanke, Mario; Olivier, Martin; Tremblay, Michel J.; Papadopoulou, Barbara; Ouellette, Marc; Corbeil, Jacques

    2012-01-01

    The Leishmania tarentolae Parrot-TarII strain genome sequence was resolved to an average 16-fold mean coverage by next-generation DNA sequencing technologies. This is the first non-pathogenic to humans kinetoplastid protozoan genome to be described thus providing an opportunity for comparison with the completed genomes of pathogenic Leishmania species. A high synteny was observed between all sequenced Leishmania species. A limited number of chromosomal regions diverged between L. tarentolae and L. infantum, while remaining syntenic to L. major. Globally, >90% of the L. tarentolae gene content was shared with the other Leishmania species. We identified 95 predicted coding sequences unique to L. tarentolae and 250 genes that were absent from L. tarentolae. Interestingly, many of the latter genes were expressed in the intracellular amastigote stage of pathogenic species. In addition, genes coding for products involved in antioxidant defence or participating in vesicular-mediated protein transport were underrepresented in L. tarentolae. In contrast to other Leishmania genomes, two gene families were expanded in L. tarentolae, namely the zinc metallo-peptidase surface glycoprotein GP63 and the promastigote surface antigen PSA31C. Overall, L. tarentolae's gene content appears better adapted to the promastigote insect stage rather than the amastigote mammalian stage. PMID:21998295

  1. Differentiation of Leishmania (Viannia) panamensis and Leishmania (V.) guyanensis using BccI for hsp70 PCR-RFLP.

    Science.gov (United States)

    Montalvo Alvarez, Ana Margarita; Nodarse, Jorge Fraga; Goodridge, Ivón Montano; Fidalgo, Lianet Monzote; Marin, Marcel; Van Der Auwera, Gert; Dujardin, Jean-Claude; Bernal, Iván Darío Velez; Muskus, Carlos

    2010-05-01

    Leishmania panamensis and Leishmania guyanensis are two species of the subgenus Viannia that are genetically very similar. Both parasites are usually associated with cutaneous leishmaniasis, but also have the potential to cause the mucocutaneous form of the disease. In addition, the study of foci and consequently the identification of vectors and probable reservoirs involved in transmission require a correct differentiation between both species, which is important at epidemiological level. We explored the possibility of identifying these species by using restriction fragment length polymorphisms (RFLP) in the gene coding for heat-shock protein 70 (hsp70). Previously, an hsp70 PCR-RFLP assay proved to be very effective in differentiating other Leishmania species when HaeIII is used as restriction enzyme. Based on hsp70 sequences analysis, BccI was found to generate species-specific fragments that can easily be recognized by agarose gel electrophoresis. Using the analysis of biopsies, scrapings, and parasite isolates previously grouped in a cluster comprising both L. panamensis and L. guyanensis, we showed that our approach allowed differentiation of both entities. This offers the possibility not only for identification of parasites in biological samples, but also to apply molecular epidemiology in certain countries of the New World, where several Leishmania species could coexist. Copyright 2009 Royal Society of Tropical Medicine and Hygiene. Published by Elsevier Ltd. All rights reserved.

  2. Molecular detection and identification of Leishmania infection in naturally infected sand flies in a focus of cutaneous leishmaniasis in northern Morocco

    OpenAIRE

    Es-Sette, Nargys; Ajaoud, Malika; Laamrani-Idrissi, Abderrahman; Mellouki, Fouad; Lemrani, Meryem

    2014-01-01

    Background Cutaneous leishmaniasis is an infectious disease caused by various species of the flagellate protozoan Leishmania. During the past 20 years, cutaneous leishmaniasis has emerged as a major public health threat in Morocco. The main objective of this study was to study the occurrence of Leishmania infection in vectors and to identify sand fly blood meal sources in an endemic locality of cutaneous leishmaniasis within Sefrou province, where the vectors of leishmaniasis were still unkno...

  3. Investigation of Double-Band Electrophoretic Pattern of ITS-rDNA Region in Iranian Isolates of Leishmania Tropica

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    MA Ghatee

    2013-06-01

    Full Text Available Background: Leishmania tropica is a genetically divergent species. Amplification of entire internal tran­scribed spacer (ITS region of L. tropica isolates obtained from Bam district, one of the well known focus of anthroponotic cutaneous leishmaniasis ACL( in Iran, revealed a double-band pat­tern in agarose gel electrophoresis. This study explains how this pattern occurs.Methods: Twenty seven L. tropica smear preparations were collected from Bam district, south east Iran, and eight L. major and one L. infantum smear preparations were gathered from Shiraz, south west Iran. Furthermore one L. major and one L. infantum cultured standard strains were tested using entire ITS-PCR to survey their electrophoretic pattern. The ITS sequences of L. tropica, L. major, and L. infantum already deposited in GenBank were analyzed. Analysis of GenBank sequences of L. tropica revealed two groups of sequences based on length size, one group having a 100 bp gap. Therefore, a new re­verse primer namely LITS-MG was designed to exclude this gap in PCR products.Results: Whole ITS fragment amplification resulted in a double-band pattern in all L. tropica cases, while a sharp single band was observed for L. infantum and L. major isolates. This result was correspond­ing to the result obtained from in silico analysis of GenBank sequences. Use of LITS-MG primer was expectedly resulted in a single band including ITS1, 5.8s and partial ITS2 product for L. tropica which is appropriate for following molecular studies such as sequencing or restriction analysis.Conclusion: Sequences analysis of GenBank L. tropica sequences and following practical laboratory tests revealed at least two alleles in L. tropica which were confirmed in Bam isolates. This especial double-band pattern is because of a 100 bp fragment difference within ITS-rDNA alleles

  4. Characterization of a midgut mucin-like glycoconjugate of Lutzomyia longipalpis with a potential role in Leishmania attachment.

    Science.gov (United States)

    Myšková, Jitka; Dostálová, Anna; Pěničková, Lucie; Halada, Petr; Bates