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Sample records for von legionella pneumophila

  1. Growth regulation of Legionella Pneumophila in biofilms and amoebae; Wachstumsregulation von Legionella Pneumophila in Biofilmen und Amoeben

    Energy Technology Data Exchange (ETDEWEB)

    Hilbi, H.

    2006-07-01

    This final report for the Swiss Federal Office of Energy (SFOE) presents the results of studies made on the regulation of the growth of Legionella Pneumophila bacteria in biofilms and amoebae. In a first project, the formation of biofilms by Legionella Pneumophila bacteria was analysed in static and dynamic systems using a complex growth medium. Under static and dynamic clinical and environmental conditions, the adherence of the biofilms on polystyrene tissue was studied. This was also examined under dynamic flow conditions. In a second part of the project, the regulation of growth of Legionella Pneumophila in amoebae was examined in that changes were made to the genome of the bacteria. The importance of the work for the de-activation of Legionella Pneumophila bacteria in biofilms is noted in the conclusions of the report.

  2. Pneumonia a Legionella pneumophila.

    Directory of Open Access Journals (Sweden)

    E. Álvares

    1998-03-01

    Full Text Available RESUMO: A Legionella pneumophila é uma bacteria comum do ambiente e um agente causal importante de pneumonias graves da comunidade. O diagnóstico em fase precoce da doença, nem sempre é fácil.Procedemos a um estudo retrospectivo, com o objectivo de caracterizar os doentes intemados na Unidade de Cuidados lntensivos Respiratórios (UCIR, com o diagnóstico de pneumonia a Legionella pneumophila (pLp, durante dez anos (1986-1996. Foram analisados doze processos clinicos que repre-sentavam 6,7% das pneumonias da comunidade intemados nesse perfodo. Comparam-se também as pneumonias a Legionella pneumophila com as outras pneumonias da comunidade, durante o mesmo perído de internamento. Onze doentes eram do sexo masculino e uma do sexo feminino, comidade média de 49,6±11, 9 anos. O TISS foi de 25,8±9,5, o APACHE II de 23,4±6,5 e o APS de 20,3±6,5. Apenas 1 doente não necessitou de ventilação medcânica, tendo sido o tempo médio de ventilação de 11,5±12,5 dias (min: 1 e máx: 44. Quatro pneumonias ocorreram fora da época sazonal habitual e só quatro doentes tinham história epidemiológica sugestiva. Apenas um doente, não tinha factores de risco. A confirmação diagnóstica foi feita por serologia e imunonuorescência directa das secreçõs brônquicas. Em relação às complicaçõoes, 9 doentes tiveram disfunção hepática, 7 insuficiência renal, 1 endocardite e 1 desenvolveu SDRA. A septicemia noscomial, ocorreu em 5 doentes. Faleceram 58,3% dos doentes que tinham índices de gravidade mais elevados.Os doentes com pLp, apresentavam maior gravidade que se traduziu por uma maior necessidade de suporte ventilatorio. A mortalidade embora mais elevada nos doentes com pLp (58,3% versus 31,8%, não foi significativamente diferente nos dois grupos quando consideramos apenas os doentes ventilados.Os autores concluem que a pneumonia da comunidade causada por Legionella pneumophila, é uma situaçã o grave que se acompanha de

  3. Legionella pneumophila in commercial bottled mineral water.

    NARCIS (Netherlands)

    Klont, R.R.; Rijs, A.J.M.M.; Warris, A.; Sturm, P.D.J.; Melchers, W.J.G.; Verweij, P.E.

    2006-01-01

    Sixty-eight commercial bottled mineral waters (64 brands, 68 different 'best-before dates') were tested for the presence of bacteria and fungi. Six samples were Legionella antigen positive and six were Legionella pneumophila PCR positive. Two samples were both Legionella antigen and L. pneumophila

  4. Hartmannella vermiformis inhibition of Legionella pneumophila cultivability

    Science.gov (United States)

    Hartmannella vermiformis is frequently isolated from drinking water (DW) and is permissive to Legionella pneumophila intracellular replication. Thus, H. vermiformis may play an important role in the growth and survival potential of such environmental pathogens. In this study, Pag...

  5. Biofilms: The Stronghold of Legionella pneumophila

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    Mena Abdel-Nour

    2013-10-01

    Full Text Available Legionellosis is mostly caused by Legionella pneumophila and is defined as a severe respiratory illness with a case fatality rate ranging from 5% to 80%. L. pneumophila is ubiquitous in natural and anthropogenic water systems. L. pneumophila is transmitted by inhalation of contaminated aerosols produced by a variety of devices. While L. pneumophila replicates within environmental protozoa, colonization and persistence in its natural environment are also mediated by biofilm formation and colonization within multispecies microbial communities. There is now evidence that some legionellosis outbreaks are correlated with the presence of biofilms. Thus, preventing biofilm formation appears as one of the strategies to reduce water system contamination. However, we lack information about the chemical and biophysical conditions, as well as the molecular mechanisms that allow the production of biofilms by L. pneumophila. Here, we discuss the molecular basis of biofilm formation by L. pneumophila and the roles of other microbial species in L. pneumophila biofilm colonization. In addition, we discuss the protective roles of biofilms against current L. pneumophila sanitation strategies along with the initial data available on the regulation of L. pneumophila biofilm formation.

  6. Detection of cell-associated or soluble antigens of Legionella pneumophila serogroups 1 to 6, Legionella bozemanii, Legionella dumoffii, Legionella gormanii, and Legionella micdadei by staphylococcal coagglutination tests.

    OpenAIRE

    Wilkinson, H W; Fikes, B J

    1981-01-01

    Current methods used for the detection of whole-cell isolates of Legionella or for the detection of Legionella soluble antigens are technically impractical for many clinical laboratories. The purpose of this study was to explore practical alternatives. The results showed that whole cell isolates of Legionella pneumophila serogroups 1 to 6, Legionella bozemanii, Legionella dumoffii, Legionella gormanii, and Legionella micdadei were identified specifically by a simple slide agglutination test o...

  7. Fatal coinfection with Legionella pneumophila serogroup 8 and Aspergillus fumigatus.

    Science.gov (United States)

    Guillouzouic, Aurélie; Bemer, Pascale; Gay-Andrieu, Françoise; Bretonnière, Cédric; Lepelletier, Didier; Mahé, Pierre-Joachim; Villers, Daniel; Jarraud, Sophie; Reynaud, Alain; Corvec, Stéphane

    2008-02-01

    Legionella pneumophila is an important cause of community-acquired and nosocomial pneumonia. We report on a patient who simultaneously developed L. pneumophila serogroup 8 pneumonia and Aspergillus fumigatus lung abscesses. Despite appropriate treatments, Aspergillus disease progressed with metastasis. Coinfections caused by L. pneumophila and A. fumigatus remain exceptional. In apparently immunocompetent patients, corticosteroid therapy is a key risk factor for aspergillosis.

  8. Ciliate Paramecium is a natural reservoir of Legionella pneumophila

    Science.gov (United States)

    Watanabe, Kenta; Nakao, Ryo; Fujishima, Masahiro; Tachibana, Masato; Shimizu, Takashi; Watarai, Masahisa

    2016-04-01

    Legionella pneumophila, the causative agent of Legionnaires’ disease, replicates within alveolar macrophages and free-living amoebae. However, the lifestyle of L. pneumophila in the environment remains largely unknown. Here we established a novel natural host model of L. pneumophila endosymbiosis using the ciliate Paramecium caudatum. We also identified Legionella endosymbiosis-modulating factor A (LefA), which contributes to the change in life stage from endosymbiosis to host lysis, enabling escape to the environment. We isolated L. pneumophila strains from the environment, and they exhibited cytotoxicity toward P. caudatum and induced host lysis. Acidification of the Legionella-containing vacuole (LCV) was inhibited, and enlarged LCVs including numerous bacteria were observed in P. caudatum infected with L. pneumophila. An isogenic L. pneumophila lefA mutant exhibited decreased cytotoxicity toward P. caudatum and impaired the modification of LCVs, resulting in the establishment of endosymbiosis between them. Our results suggest that L. pneumophila may have a mechanism to switch their endosymbiosis in protistan hosts in the environment.

  9. Molecular Evolution of the dotA Gene in Legionella pneumophila

    OpenAIRE

    Ko, Kwan Soo; Hong, Seong Karp; Lee, Hae Kyung; Park, Mi-Yeoun; Kook, Yoon-Hoh

    2003-01-01

    The molecular evolution of dotA, which is related to the virulence of Legionella pneumophila, was investigated by comparing the sequences of 15 reference strains (serogroups 1 to 15). It was found that dotA has a complex mosaic structure. The whole dotA gene of Legionella pneumophila subsp. pneumophila serogroups 2, 6, and 12 has been transferred from Legionella pneumophila subsp. fraseri. A discrepancy was found between the trees inferred from the nucleotide and deduced amino acid sequences ...

  10. Genetic typing in a cluster of Legionella pneumophila infections

    NARCIS (Netherlands)

    van Ketel, R. J.; de Wever, B.

    1989-01-01

    Legionella pneumophila strains isolated from six patients, three air-conditioning- and cooling tower-derived strains, and three hot water supply-derived strains were analyzed by three genetic typing methods. The results of the whole-cell DNA restriction endonuclease analysis and the restriction

  11. Legionella pneumophila Seropositivity-Associated Factors in Latvian Blood Donors

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    Olga Valciņa

    2015-12-01

    Full Text Available Continuous environmental exposure of humans to Legionella may induce immune responses and generation of antibodies. The aim of this study was to investigate the seroprevalence of Legionella pneumophila serogroups (SG 1–6 in the general healthy population and identify the associated host-related and environmental risk factors. L. pneumophila SG 1–6 seroprevalence among a total of 2007 blood samples collected from healthy donors was 4.8%. Seroprevalence was higher in women (5.9% than men (3.3% and in areas with a larger number of inhabitants, ranging from 3.5% in rural regions to 6.8% in the capital, Riga. Blood samples from inhabitants of apartment buildings tested positive for L. pneumophila in more cases (5.8% compared to those from inhabitants of single-family homes (2.7%. Residents of buildings with a municipal hot water supply system were more likely to be seropositive for L. pneumophila (OR = 3.16, 95% CI 1.26–7.91. Previous episodes of fever were additionally identified as a risk factor (OR = 2.42, 95% CI 1.43–4.1. In conclusion, centralized hot water supply, female gender and previous episodes of fever were determined as the main factors associated with L. pneumophila seropositivity in our study population.

  12. Legionella pneumophila Pathogenesis in the Galleria mellonella Infection Model

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    Harding, Clare R.; Schroeder, Gunnar N.; Reynolds, Stuart; Kosta, Artemis; Collins, James W.; Mousnier, Aurélie

    2012-01-01

    Legionella pneumophila is a facultative intracellular human pathogen and the etiological agent of severe pneumonia known as Legionnaires' disease. Its virulence depends on protein secretion systems, in particular, the Dot/Icm type IV secretion system (T4SS), which is essential to establish a replication-permissive vacuole in macrophages. The analysis of the role of these systems and their substrates for pathogenesis requires easy-to-use models which approximate human infection. We examined the effectiveness of the larvae of the wax moth Galleria mellonella as a new model for L. pneumophila infection. We found that the L. pneumophila strains 130b, Paris, and JR32 caused mortality of the G. mellonella larvae that was strain, infectious dose, growth phase, and T4SS dependent. Wild-type L. pneumophila persisted and replicated within the larvae, whereas T4SS mutants were rapidly cleared. L. pneumophila strain Lp02, which is attenuated in the absence of thymidine but has a functional T4SS, resisted clearance in G. mellonella up to 18 h postinfection without inducing mortality. Immunofluorescence and transmission electron microscopy revealed that L. pneumophila resided within insect hemocytes in a vacuole that ultrastructurally resembled the Legionella-containing vacuole (LCV) observed in macrophages. The vacuole was decorated with the T4SS effector and LCV marker SidC. Infection caused severe damage to the insect organs and triggered immune responses, including activation of the phenoloxidase cascade leading to melanization, nodule formation, and upregulation of antimicrobial peptides. Taken together, these results suggest that G. mellonella provides an effective model to investigate the interaction between L. pneumophila and the host. PMID:22645286

  13. Virulence properties of the Legionella pneumophila cell envelope

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    Olga eShevchuk

    2011-04-01

    Full Text Available The bacterial envelope plays a crucial role in the pathogenesis of infectious diseases. In this review, we summarize the current knowledge of the structure and molecular composition of the Legionella pneumophila cell envelope. We describe LPS biosynthesis and the biological activities of membrane and periplasmic proteins and discuss their decisive functions during the pathogen-host interaction. In addition to adherence, invasion and intracellular survival of L. pneumophila, special emphasis is laid on iron acquisition, detoxification, key elicitors of the immune response and the diverse functions of outer membrane vesicles. The critical analysis of the literature reveals that the dynamics and phenotypic plasticity of the Legionella cell surface during the different metabolic stages requires more attention in the future.

  14. Genetic typing in a cluster of Legionella pneumophila infections.

    OpenAIRE

    van Ketel, R J; de Wever, B

    1989-01-01

    Legionella pneumophila strains isolated from six patients, three air-conditioning- and cooling tower-derived strains, and three hot water supply-derived strains were analyzed by three genetic typing methods. The results of the whole-cell DNA restriction endonuclease analysis and the restriction patterns based on genes coding for rRNA correlated with each other and demonstrated that the patient isolates were indistinguishable from the air-conditioning- and cooling tower-derived isolates but di...

  15. Environmental surveillance of Legionella pneumophila in two Italian hospitals

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    Marina Tesauro

    2010-01-01

    Full Text Available The aim of this study was to identify the most effective disinfection protocol to reduce the presence of Legionella pneumophila in the water system of two Italian hospitals. From 2004 to 2009, 271 samplings of hot water were carried out in 11 hospital units to detect the presence of L. pneumophila. Additionally, water samples collected from one boiler outlet and the hot water recirculation were tested. From 2004 to 2009, L. pneumophila was present in 37% of the samples. Of these, 68.3% and 18.8% were positive for serogroups 2-14 and 1, respectively. Furthermore, 12.9% of the samples were positive for both serogroups. Finally, a maximal count of 10(4 CFU/L was measured in the most distal sites. To reduce L. pneumophila colonization, a two-year long hyperchlorination (2004-2006 was carried out. Moreover, from June 2005 until now, continuous maintenance of boilers and tanks, substitution of the shower heads and increase of the boiler outlet temperature to 60 ºC were performed. All these treatments led to a marked reduction of L. pneumophila colonization in the short but not in the medium-long term. Only the use of chlorine dioxide led, after four years, to a reduction of the loads of L. pneumophila to values below 100 CFU/L. However, in the distal sites a persistent degree of colonization (maximum value 700 CFU/L, average 600 CFU/L was observed probably due to the presence of L. pneumophila in the stagnant water in dead legs. In conclusion, data show that long-term chlorination of hot water sources together with carefully aimed maintenance of water pipes can lead to an effective reduction of L. pneumophila concentration in hospital water systems.

  16. Isolation, identification, characterization and antibiotic sensitivity profile of pathogenic Legionella pneumophila isolates from different water sources

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    Kannan Subbaram

    2017-05-01

    Conclusions: Serious and fatal L. pneumophila infections may be transmitted through water. Legionella can survive under various conditions in various water sources. L. pneumophila is the important pathogen causing human disease. Great challenge prevails to health care professionals because these Legionellae acquired antibiotic resistance to many routinely prescribed antibiotics.

  17. Nucleotide sequence analysis of the Legionella micdadei mip gene, encoding a 30-kilodalton analog of the Legionella pneumophila Mip protein

    DEFF Research Database (Denmark)

    Bangsborg, Jette Marie; Cianciotto, N P; Hindersson, P

    1991-01-01

    After the demonstration of analogs of the Legionella pneumophila macrophage infectivity potentiator (Mip) protein in other Legionella species, the Legionella micdadei mip gene was cloned and expressed in Escherichia coli. DNA sequence analysis of the L. micdadei mip gene contained in the plasmid p...... homology with the mip-like genes of several Legionella species. Furthermore, amino acid sequence comparisons revealed significant homology to two eukaryotic proteins with isomerase activity (FK506-binding proteins)....

  18. Severe Pneumonia Caused by Legionella pneumophila: Differential Diagnosis and Therapeutic Considerations.

    Science.gov (United States)

    Chahin, Abdullah; Opal, Steven M

    2017-03-01

    Severe legionella pneumonia poses a diagnostic challenge and requires early intervention. Legionnaire's disease can have several presenting signs, symptoms, and laboratory abnormalities that suggest that Legionella pneumophila is the pathogen, but none of these are sufficient to distinguish L pneumophila pneumonia from other respiratory pathogens. L pneumophila is primarily an intracellular pathogen and needs treatment with antibiotics that efficiently enter the intracellular space. Copyright © 2016 Elsevier Inc. All rights reserved.

  19. Legionella pneumophila Arthritis: use of medium specific for Mycobacteria for isolation of L. pneumophila in culture of articular fluid specimens.

    Science.gov (United States)

    Bemer, Pascale; Leautez, Sophie; Ninin, Emmanuelle; Jarraud, Sophie; Raffi, François; Drugeon, Henri

    2002-07-01

    We report the first case, to our knowledge, of acute purulent arthritis due to Legionella pneumophila in an immunosuppressed patient. L. pneumophila was isolated from samples of blood and articular fluid cultured with use of medium specific for mycobacteria (Bactec 13A medium).

  20. Legionella pneumophila pangenome reveals strain-specific virulence factors

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    Peris-Bondia Francesc

    2010-03-01

    Full Text Available Abstract Background Legionella pneumophila subsp. pneumophila is a gram-negative γ-Proteobacterium and the causative agent of Legionnaires' disease, a form of epidemic pneumonia. It has a water-related life cycle. In industrialized cities L. pneumophila is commonly encountered in refrigeration towers and water pipes. Infection is always via infected aerosols to humans. Although many efforts have been made to eradicate Legionella from buildings, it still contaminates the water systems. The town of Alcoy (Valencian Region, Spain has had recurrent outbreaks since 1999. The strain "Alcoy 2300/99" is a particularly persistent and recurrent strain that was isolated during one of the most significant outbreaks between the years 1999-2000. Results We have sequenced the genome of the particularly persistent L. pneumophila strain Alcoy 2300/99 and have compared it with four previously sequenced strains known as Philadelphia (USA, Lens (France, Paris (France and Corby (England. Pangenome analysis facilitated the identification of strain-specific features, as well as some that are shared by two or more strains. We identified: (1 three islands related to anti-drug resistance systems; (2 a system for transport and secretion of heavy metals; (3 three systems related to DNA transfer; (4 two CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats systems, known to provide resistance against phage infections, one similar in the Lens and Alcoy strains, and another specific to the Paris strain; and (5 seven islands of phage-related proteins, five of which seem to be strain-specific and two shared. Conclusions The dispensable genome disclosed by the pangenomic analysis seems to be a reservoir of new traits that have mainly been acquired by horizontal gene transfer and could confer evolutionary advantages over strains lacking them.

  1. [Analysis of prevalence and variability of Legionella pneumophila and Legionella spp. strains on the basis of study of allelic profiles].

    Science.gov (United States)

    Voronina, O L; Kunda, M S; Bitkina, V V; Karpova, T I; Romanenko, V V; Durasova, A L; Tartakovskiĭ, I S

    2009-01-01

    To analyze prevalence and variability of Legionella strains isolated in town Verkhnaya Pyshma located in Sverdlovsk region during prophylactic surveillance of potentially dangerous water objects in 2007 - 2008. Sequencing of mip gene was conducted for identification of species of Legionella. Multi-locus sequence typing was used for describing of allelic profiles of Legionella pneumophila strains. Five firstly identified on Russian territory strains of Legionella species were deposited in institute's collection. Sixty-three strains of L. pneumophila belonging to 28 sequence types were characterized. Relation between strains isolated in industrial building and from water supply system was demonstrated. Observations made on the basis of study of L. pneumophila strains isolated from cooling stacks of industrial plants confirmed potential danger of these objects as a source of dissemination of Legionella infection.

  2. Exploring the Legionella pneumophila positivity rate in hotel water samples from Antalya, Turkey.

    Science.gov (United States)

    Sepin Özen, Nevgün; Tuğlu Ataman, Şenay; Emek, Mestan

    2017-05-01

    The genus Legionella is a fastidious Gram-negative bacteria widely distributed in natural waters and man made water supply systems. Legionella pneumophila is the aetiological agent of approximately 90% of reported Legionellosis cases, and serogroup 1 is the most frequent cause of infections. Legionnaires' disease is often associated with travel and continues to be a public health concern at present. The correct water management quality practices and rapid methods for analyzing Legionella species in environmental water is a key point for the prevention of Legionnaires' disease outbreaks. This study aimed to evaluate the positivity rates and serotyping of Legionella species from water samples in the region of Antalya, Turkey, which is an important tourism center. During January-December 2010, a total of 1403 samples of water that were collected from various hotels (n = 56) located in Antalya were investigated for Legionella pneumophila. All samples were screened for L. pneumophila by culture method according to "ISO 11731-2" criteria. The culture positive Legionella strains were serologically identified by latex agglutination test. A total of 142 Legionella pneumophila isolates were recovered from 21 (37.5%) of 56 hotels. The total frequency of L. pneumophila isolation from water samples was found as 10.1%. Serological typing of 142 Legionella isolates by latex agglutination test revealed that strains belonging to L. pneumophila serogroups 2-14 predominated in the examined samples (85%), while strains of L. pneumophila serogroup 1 were less numerous (15%). According to our knowledge, our study with the greatest number of water samples from Turkey demonstrates that L. pneumophila serogroups 2-14 is the most common isolate. Rapid isolation of L. pneumophila from environmental water samples is essential for the investigation of travel related outbreaks and the possible resources. Further studies are needed to have epidemiological data and to determine the types of L

  3. Detection and Quantification of Legionella pneumophila from Water Systems in Kuwait Residential Facilities

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    Qadreyah A. Al-Matawah

    2012-01-01

    Full Text Available The prevalence of Legionella pneumophilia in water systems of residential facilities in Kuwait was performed during the period from November 2007 to November 2011. A total of 204 water samples collected from faucets and showerheads in bathrooms (n = 82, taps in kitchens (n = 51, and water tanks (n = 71, from different locations of residential facilities in Kuwait were screened for Legionella pneumophila by the standard culture method and by real time polymerase chain reaction (RT-PCR. Out of the 204 samples, 89 (43.6% samples were positive for Legionella spp., 48 (23.5% samples were detected by the standard culture method, and 85 (41.7% were detected by RT-PCR. Of the culture positive Legionella samples, counts ranged between 10 to 2250 CFU/L. Serological typing of 48 Legionella isolates revealed that 6 (12.5% of these isolates belonged to Legionella pneumophila serogroup 1, 37 (77.1% isolates to Legionella pneumophila serogroup 3, and 1 isolate each (2.1% belonged to serogroups 4, 7, and 10. The minimum inhibitory concentration (MICs of the 46 environmental L. pneumophila isolates against the 10 antimicrobials commonly used for Legionella infection treatments were determined. Rifampicin was found to be the most active against L. pneumophila serogroups isolates in vitro.

  4. Epidemiology and Ecology of Opportunistic Premise Plumbing Pathogens: Legionella pneumophila, Mycobacterium avium, and Pseudomonas aeruginosa

    Science.gov (United States)

    BACKGROUND: Legionella pneumophila, Mycobacterium avium, and Pseudomonas aeruginosa are opportunistic premise plumbing pathogens (OPPPs) that persist and grow in household plumbing, habitats they share with humans. Infections caused by these OPPPs involve individuals with preexis...

  5. Occurrence of Opportunistic Pathogens Legionella pneumophila and non-tuberculous mycobacteria in hospital plumbing systems

    Science.gov (United States)

    Occurrence of Opportunistic Pathogens Legionella pneumophila and non-tuberculous mycobacteria in hospital plumbing systems Jill Hoelle, Michael Coughlin, Elizabeth Sotkiewicz, Jingrang Lu, Stacy Pfaller, Mark Rodgers, and Hodon Ryu U.S. Environmental Protection Agency, Cincinnati...

  6. Effect of Legionella pneumophila cytotoxic protease on human neutrophil and monocyte function

    DEFF Research Database (Denmark)

    Rechnitzer, C; Kharazmi, A

    1992-01-01

    The extracellular metalloprotease of Legionella pneumophila, also called tissue-destructive protease or major secretory protein, has been proposed as one of the virulence factors of this organism. Considering the decisive role played by the phagocytic cells in host defense against Legionella infe....... pneumophila protease on neutrophil chemotaxis and on the listericidal activity of human neutrophils and monocytes demonstrated in this study provides evidence for a role of this enzyme in the pathogenesis of Legionnaires' disease....

  7. Spatial distribution of Legionella pneumophila MLVA-genotypes in a drinking water system.

    Science.gov (United States)

    Rodríguez-Martínez, Sarah; Sharaby, Yehonatan; Pecellín, Marina; Brettar, Ingrid; Höfle, Manfred; Halpern, Malka

    2015-06-15

    Bacteria of the genus Legionella cause water-based infections, resulting in severe pneumonia. To improve our knowledge about Legionella spp. ecology, its prevalence and its relationships with environmental factors were studied. Seasonal samples were taken from both water and biofilm at seven sampling points of a small drinking water distribution system in Israel. Representative isolates were obtained from each sample and identified to the species level. Legionella pneumophila was further determined to the serotype and genotype level. High resolution genotyping of L. pneumophila isolates was achieved by Multiple-Locus Variable number of tandem repeat Analysis (MLVA). Within the studied water system, Legionella plate counts were higher in summer and highly variable even between adjacent sampling points. Legionella was present in six out of the seven selected sampling points, with counts ranging from 1.0 × 10(1) to 5.8 × 10(3) cfu/l. Water counts were significantly higher in points where Legionella was present in biofilms. The main fraction of the isolated Legionella was L. pneumophila serogroup 1. Serogroup 3 and Legionella sainthelensis were also isolated. Legionella counts were positively correlated with heterotrophic plate counts at 37 °C and negatively correlated with chlorine. Five MLVA-genotypes of L. pneumophila were identified at different buildings of the sampled area. The presence of a specific genotype, "MLVA-genotype 4", consistently co-occurred with high Legionella counts and seemed to "trigger" high Legionella counts in cold water. Our hypothesis is that both the presence of L. pneumophila in biofilm and the presence of specific genotypes, may indicate and/or even lead to high Legionella concentration in water. This observation deserves further studies in a broad range of drinking water systems to assess its potential for general use in drinking water monitoring and management. Copyright © 2015 Elsevier Ltd. All rights reserved.

  8. Characterization of an acetyltransferase that detoxifies aromatic chemicals in Legionella pneumophila

    DEFF Research Database (Denmark)

    Kubiak, Xavier Jean Philippe; Dervins-Ravault, Delphine; Pluvinage, Benjamin

    2012-01-01

    Legionella pneumophila is an opportunistic pathogen and the causative agent of Legionnaires' disease. Despite being exposed to many chemical compounds in its natural and man-made habitats (natural aquatic biotopes and man-made water systems), L. pneumophila is able to adapt and survive in these e...

  9. A Multiplex PCR for Detection of Mycoplasma pneumoniae, Chlamydophila pneumoniae, Legionella pneumophila, and Bordetella pertussis in Clinical Specimens

    National Research Council Canada - National Science Library

    McDonough, E. A; Barrozo, C. P; Russell, K. L; Metzgar, D

    2005-01-01

    A multiplex PCR was developed that is capable of detecting four of the most important bacterial agents of atypical pneumophia, Mycaplasma pneumoniae, Chlamydophia pneumoniae, Legionella pneumophila...

  10. Preferential colonization and release of Legionella pneumophila from mature drinking water biofilms grown on copper versus unplasticized polyvinylchloride coupons

    Science.gov (United States)

    Legionella persistence and amplification in premise drinking water systems is a known contributor to legionellosis outbreaks, especially in the presence of suitable eukaryotic hosts. Here we examined Legionella pneumophila behavior within drinking water biofilms grown on copper ...

  11. A comparison of assays measuring the viability of Legionella pneumophila after treatment with copper and silver ions

    Science.gov (United States)

    Background: The relatively high prevalence of Legionella pneumophila in premise plumbing systems has been widely reported. Published reports indicate Legionella has a comparatively high resistance to chlorine and moreover has the ability to grow in phagocytic amoeba which could p...

  12. Identification of vacuoles containing extraintestinal differentiated forms of Legionella pneumophila in colonized Caenorhabditis elegans soil nematodes

    Science.gov (United States)

    Hellinga, Jacqueline R; Garduño, Rafael A; Kormish, Jay D; Tanner, Jennifer R; Khan, Deirdre; Buchko, Kristyn; Jimenez, Celine; Pinette, Mathieu M; Brassinga, Ann Karen C

    2015-01-01

    Legionella pneumophila, a causative agent of Legionnaires’ disease, is a facultative intracellular parasite of freshwater protozoa. Legionella pneumophila features a unique developmental network that involves several developmental forms including the infectious cyst forms. Reservoirs of L. pneumophila include natural and man-made freshwater systems; however, recent studies have shown that isolates of L. pneumophila can also be obtained directly from garden potting soil suggesting the presence of an additional reservoir. A previous study employing the metazoan Caenorhabditis elegans, a member of the Rhabditidae family of free-living soil nematodes, demonstrated that the intestinal lumen can be colonized with L. pneumophila. While both replicative forms and differentiated forms were observed in C. elegans, these morphologically distinct forms were initially observed to be restricted to the intestinal lumen. Using live DIC imaging coupled with focused transmission electron microscopy analyses, we report here that L. pneumophila is able to invade and establish Legionella-containing vacuoles (LCVs) in the intestinal cells. In addition, LCVs containing replicative and differentiated cyst forms were observed in the pseudocoelomic cavity and gonadal tissue of nematodes colonized with L. pneumophila. Furthermore, establishment of LCVs in the gonadal tissue was Dot/Icm dependent and required the presence of the endocytic factor RME-1 to gain access to maturing oocytes. Our findings are novel as this is the first report, to our knowledge, of extraintestinal LCVs containing L. pneumophila cyst forms in C. elegans tissues, highlighting the potential of soil-dwelling nematodes as an alternate environmental reservoir for L. pneumophila. PMID:26131925

  13. Widespread molecular detection of Legionella pneumophila Serogroup 1 in cold water taps across the United States

    Science.gov (United States)

    In the United States 3,522 cases of legionellosis were reported to the Center for Disease Control and Prevention in 2009. Of these reports, it is estimated that 84% are caused by the microorganism Legionella pneumophila Serogroup (Sg) 1. Legionella spp. have been isolated and r...

  14. Legionella pneumophila secretes a mitochondrial carrier protein during infection.

    Directory of Open Access Journals (Sweden)

    Pavel Dolezal

    2012-01-01

    Full Text Available The Mitochondrial Carrier Family (MCF is a signature group of integral membrane proteins that transport metabolites across the mitochondrial inner membrane in eukaryotes. MCF proteins are characterized by six transmembrane segments that assemble to form a highly-selective channel for metabolite transport. We discovered a novel MCF member, termed Legionellanucleotide carrier Protein (LncP, encoded in the genome of Legionella pneumophila, the causative agent of Legionnaire's disease. LncP was secreted via the bacterial Dot/Icm type IV secretion system into macrophages and assembled in the mitochondrial inner membrane. In a yeast cellular system, LncP induced a dominant-negative phenotype that was rescued by deleting an endogenous ATP carrier. Substrate transport studies on purified LncP reconstituted in liposomes revealed that it catalyzes unidirectional transport and exchange of ATP transport across membranes, thereby supporting a role for LncP as an ATP transporter. A hidden Markov model revealed further MCF proteins in the intracellular pathogens, Legionella longbeachae and Neorickettsia sennetsu, thereby challenging the notion that MCF proteins exist exclusively in eukaryotic organisms.

  15. Lymphoid cell blastogenesis as an in vitro indicator of cellular immunity to Legionella pneumophila antigens.

    OpenAIRE

    Friedman, F; Widen, R; Klein, T; Friedman, H

    1984-01-01

    The lymphocyte blastogenic transformation assay was adapted to study responsiveness of lymphoid cells from animals and humans to Legionella pneumophila antigens in vitro. Spleen cells from guinea pigs after active immunization with Legionella vaccine, but not from normal animals, responded by blast cell transformation when stimulated in vitro with killed Legionella whole-cell vaccine, sonic extracts thereof, or a purified somatic antigen. The response was dose dependent. Similar lymphocyte bl...

  16. [EFFECT OF TEMPERATURE ON THE VIABILITY OF PLANKTON CELLS AND MODEL LEGIONELLA PNEUMOPHILA BIOFILMS IN WATER].

    Science.gov (United States)

    Tartakovsky, I S; Karpoval, T I; Gruzdeva, O A; Marinenko, O V; Dronina, Yu E

    2015-01-01

    Study the effect of water temperature from 40 to 70 degrees C on viability of plankton forms and model Legionella pneumophila under experimental conditions. Monospecies legionella biofilms, obtained in plates for enzyme immunoassay during 96 hours at 28 degrees C, and water suspension of BCYE agar cultivated cells of L. pneumophila at a concentration of 10(3) - 10(5) CFU per liter were used in the study for evaluation of bactericidal effect of temperature on various legionella forms. Analysis of effects of various temperature regimens on plankton forms and model legionella biofilms has shown that at a temperature range from 50 to 60 degrees C a significant reduction of quantity of viable legionella cells occurs. Model legionella biofilms have partially conserved viability at a temperature of 60 degrees C and only exposition to a temperature of 70 degrees C resulted in death of legionella biofilms and plankton forms of bacteria. A dependence of viability conservation of legionella from the initial concentration of the causative agent in water and duration of exposition at varying temperature was shown. Short-term heating at a temperature of at least 70 degrees C has the most pronounced bactericidal effect on plankton forms and model L. pneumophila biofilms under experimental conditions. Such temperature regimen could be used as one of the prophylaxis approaches during maintenance of especially dangerous water system and, fist of all, systems of hot water supply.

  17. Dynamics of Legionella spp. and bacterial populations during the proliferation of L. pneumophila in a cooling tower facility.

    Science.gov (United States)

    Wéry, Nathalie; Bru-Adan, Valérie; Minervini, Céline; Delgénes, Jean-Philippe; Garrelly, Laurent; Godon, Jean-Jacques

    2008-05-01

    The dynamics of Legionella spp. and of dominant bacteria were investigated in water from a cooling tower plant over a 9-month period which included several weeks when Legionella pneumophila proliferated. The structural diversity of both the bacteria and the Legionella spp. was monitored by a fingerprint technique, single-strand conformation polymorphism, and Legionella spp. and L. pneumophila were quantified by real-time quantitative PCR. The structure of the bacterial community did not change over time, but it was perturbed periodically by chemical treatment or biofilm detachment. In contrast, the structure of the Legionella sp. population changed in different periods, its dynamics at times showing stability but also a rapid major shift during the proliferation of L. pneumophila in July. The dynamics of the Legionella spp. and of dominant bacteria were not correlated. In particular, no change in the bacterial community structure was observed during the proliferation of L. pneumophila. Legionella spp. present in the cooling tower system were identified by cloning and sequencing of 16S rRNA genes. A high diversity of Legionella spp. was observed before proliferation, including L. lytica, L. fallonii, and other Legionella-like amoebal pathogen types, along with as-yet-undescribed species. During the proliferation of L. pneumophila, Legionella sp. diversity decreased significantly, L. fallonii and L. pneumophila being the main species recovered.

  18. Molecular characterization of Legionella pneumophila-induced interleukin-8 expression in T cells

    Directory of Open Access Journals (Sweden)

    Mukaida Naofumi

    2010-01-01

    Full Text Available Abstract Background Legionella pneumophila is the causative agent of human Legionnaire's disease. During infection, the bacterium invades macrophages and lung epithelial cells, and replicates intracellularly. However, little is known about its interaction with T cells. We investigated the ability of L. pneumophila to infect and stimulate the production of interleukin-8 (IL-8 in T cells. The objective of this study was to assess whether L. pneumophila interferes with the immune system by interacting and infecting T cells. Results Wild-type L. pneumophila and flagellin-deficient Legionella, but not L. pneumophila lacking a functional type IV secretion system Dot/Icm, replicated in T cells. On the other hand, wild-type L. pneumophila and Dot/Icm-deficient Legionella, but not flagellin-deficient Legionella or heat-killed Legionella induced IL-8 expression. L. pneumophila activated an IL-8 promoter through the NF-κB and AP-1 binding regions. Wild-type L. pneumophila but not flagellin-deficient Legionella activated NF-κB, p38 mitogen-activated protein kinase (MAPK, Jun N-terminal kinase (JNK, and transforming growth factor β-associated kinase 1 (TAK1. Transfection of dominant negative mutants of IκBα, IκB kinase, NF-κB-inducing kinase, TAK1, MyD88, and p38 MAPK inhibited L. pneumophila-induced IL-8 activation. Inhibitors of NF-κB, p38 MAPK, and JNK blocked L. pneumophila-induced IL-8 expression. In addition, c-Jun, JunD, cyclic AMP response element binding protein, and activating transcription factor 1, which are substrates of p38 MAPK and JNK, bound to the AP-1 site of the IL-8 promoter. Conclusions Taken together, L. pneumophila induced a flagellin-dependent activation of TAK1, p38 MAPK, and JNK, as well as NF-κB and AP-1, which resulted in IL-8 production in human T cells, presumably contributing to the immune response in Legionnaire's disease.

  19. Legionella pneumophila-Derived Outer Membrane Vesicles Promote Bacterial Replication in Macrophages.

    Directory of Open Access Journals (Sweden)

    Anna Lena Jung

    2016-04-01

    Full Text Available The formation and release of outer membrane vesicles (OMVs is a phenomenon of Gram-negative bacteria. This includes Legionella pneumophila (L. pneumophila, a causative agent of severe pneumonia. Upon its transmission into the lung, L. pneumophila primarily infects and replicates within macrophages. Here, we analyzed the influence of L. pneumophila OMVs on macrophages. To this end, differentiated THP-1 cells were incubated with increasing doses of Legionella OMVs, leading to a TLR2-dependent classical activation of macrophages with the release of pro-inflammatory cytokines. Inhibition of TLR2 and NF-κB signaling reduced the induction of pro-inflammatory cytokines. Furthermore, treatment of THP-1 cells with OMVs prior to infection reduced replication of L. pneumophila in THP-1 cells. Blocking of TLR2 activation or heat denaturation of OMVs restored bacterial replication in the first 24 h of infection. With prolonged infection-time, OMV pre-treated macrophages became more permissive for bacterial replication than untreated cells and showed increased numbers of Legionella-containing vacuoles and reduced pro-inflammatory cytokine induction. Additionally, miRNA-146a was found to be transcriptionally induced by OMVs and to facilitate bacterial replication. Accordingly, IRAK-1, one of miRNA-146a's targets, showed prolonged activation-dependent degradation, which rendered THP-1 cells more permissive for Legionella replication. In conclusion, L. pneumophila OMVs are initially potent pro-inflammatory stimulators of macrophages, acting via TLR2, IRAK-1, and NF-κB, while at later time points, OMVs facilitate L. pneumophila replication by miR-146a-dependent IRAK-1 suppression. OMVs might thereby promote spreading of L. pneumophila in the host.

  20. The Legionella pneumophila Collagen-Like Protein Mediates Sedimentation, Autoaggregation, and Pathogen-Phagocyte Interactions

    Science.gov (United States)

    Abdel-Nour, Mena; Duncan, Carla; Prashar, Akriti; Rao, Chitong; Ginevra, Christophe; Jarraud, Sophie; Low, Donald E.; Ensminger, Alexander W.; Terebiznik, Mauricio R.

    2014-01-01

    Although only partially understood, multicellular behavior is relatively common in bacterial pathogens. Bacterial aggregates can resist various host defenses and colonize their environment more efficiently than planktonic cells. For the waterborne pathogen Legionella pneumophila, little is known about the roles of autoaggregation or the parameters which allow cell-cell interactions to occur. Here, we determined the endogenous and exogenous factors sufficient to allow autoaggregation to take place in L. pneumophila. We show that isolates from Legionella species which do not produce the Legionella collagen-like protein (Lcl) are deficient in autoaggregation. Targeted deletion of the Lcl-encoding gene (lpg2644) and the addition of Lcl ligands impair the autoaggregation of L. pneumophila. In addition, Lcl-induced autoaggregation requires divalent cations. Escherichia coli producing surface-exposed Lcl is able to autoaggregate and shows increased biofilm production. We also demonstrate that L. pneumophila infection of Acanthamoeba castellanii and Hartmanella vermiformis is potentiated under conditions which promote Lcl dependent autoaggregation. Overall, this study shows that L. pneumophila is capable of autoaggregating in a process that is mediated by Lcl in a divalent-cation-dependent manner. It also reveals that Lcl potentiates the ability of L. pneumophila to come in contact, attach, and infect amoebae. PMID:24334670

  1. Legionella pneumophila Carbonic Anhydrases: Underexplored Antibacterial Drug Targets

    Directory of Open Access Journals (Sweden)

    Claudiu T. Supuran

    2016-06-01

    Full Text Available Carbonic anhydrases (CAs, EC 4.2.1.1 are metalloenzymes which catalyze the hydration of carbon dioxide to bicarbonate and protons. Many pathogenic bacteria encode such enzymes belonging to the α-, β-, and/or γ-CA families. In the last decade, enzymes from some of these pathogens, including Legionella pneumophila, have been cloned and characterized in detail. These enzymes were shown to be efficient catalysts for CO2 hydration, with kcat values in the range of (3.4–8.3 × 105 s−1 and kcat/KM values of (4.7–8.5 × 107 M−1·s−1. In vitro inhibition studies with various classes of inhibitors, such as anions, sulfonamides and sulfamates, were also reported for the two β-CAs from this pathogen, LpCA1 and LpCA2. Inorganic anions were millimolar inhibitors, whereas diethyldithiocarbamate, sulfamate, sulfamide, phenylboronic acid, and phenylarsonic acid were micromolar ones. The best LpCA1 inhibitors were aminobenzolamide and structurally similar sulfonylated aromatic sulfonamides, as well as acetazolamide and ethoxzolamide (KIs in the range of 40.3–90.5 nM. The best LpCA2 inhibitors belonged to the same class of sulfonylated sulfonamides, together with acetazolamide, methazolamide, and dichlorophenamide (KIs in the range of 25.2–88.5 nM. Considering such preliminary results, the two bacterial CAs from this pathogen represent promising yet underexplored targets for obtaining antibacterials devoid of the resistance problems common to most of the clinically used antibiotics, but further studies are needed to validate them in vivo as drug targets.

  2. Viable Legionella pneumophila bacteria in natural soil and rainwater puddles

    NARCIS (Netherlands)

    van Heijnsbergen, E.; de Roda Husman, A. M.; Lodder, W. J.; Bouwknegt, M.; Docters van Leeuwen, A. E.; Bruin, J. P.; Euser, S. M.; den Boer, J. W.; Schalk, J. A C

    Aims: For the majority of sporadic Legionnaires' disease cases the source of infection remains unknown. Infection may possible result from exposure to Legionella bacteria in sources that are not yet considered in outbreak investigations. Therefore, potential sources of pathogenic Legionella

  3. [Construction and expression of eucaryotic recombinant plasmid of Legionella pneumophila mip gene].

    Science.gov (United States)

    Wang, Tao; Chen, Jian-Ping; Li, Hong; Lei, Zhang; Tao, Da-Chang; Yang, Chun-Lei

    2005-11-01

    To construct recombinant plasmid of Legionella pneumophila mip gene and detect its expression in NIH3T3 cells. mip gene of Legionella pneumophila was amplified by PCR. The amplified DNA was ligated to pcDNA3.1(+) vector. The recombinant plasmid was named pcDNA3.1-mip. NIH3T3 cell was transfected by recombinant plasmid pcDNA3.1-mip with Lipofection strategy. Transient and stable products of mip gene were detected by immunofluorescence and Western-blot. It was found that there was high green fluorescence on the cell membrane and inside the cell. It showed that NIH3T3 cell was transfected by pcDNA3.1-mip successfully. Rabbit serum antibody of Legionella pneumophila detected the NIH3T3 cell transfected with pcDNA3.1-mip. There was the protein in relative molecular weight 24 X 10(3), whereas no evidence for the protein in NIH3T3 cell transfected with pcDNA3.1(+) was seen. The protein expression of mip gene was shown. We have successfully constructed the recombinant plasmid of Legionella pneumophila mip gene and detected the relative molecular weight 24 X 10(3) Mip protein in NIH3T3 cells.

  4. Chlamydia trachomatis contains a protein similar to the Legionella pneumophila mip gene product

    DEFF Research Database (Denmark)

    Lundemose, AG; Birkelund, Svend; Fey, SJ

    1991-01-01

    A 27kDa Chlamydia trachomatis L2 protein was characterized by the use of monoclonal antibodies and by two-dimensional gel electrophoresis. The protein was shown to be located in the membrane of reticulate bodies as well as elementary bodies. Its synthesis could be detected from 10 hours post-infe...... potentiator (mip) gene of Legionella pneumophila....

  5. Structural and functional studies of a Cu+-ATPase from Legionella pneumophila

    DEFF Research Database (Denmark)

    Mattle, Daniel

    During his studies, Daniel Mattle explored the copper(I) export mechanism of a P-type Cu+ ATPase from Legionella pneumophila – a homologue to the human Cu+ ATPases. Cu+ ATPases are responsible for the homeostatic control of the physiological relevant – but toxic – copper(I) cations. To assess...

  6. Travel-associated infections caused by unusual serogroups of Legionella pneumophila identified using Legionella BIOCHIP slides in Turkey and Iraq.

    Science.gov (United States)

    Kocazeybek, Bekir S; Yuksel, Pelin; Keskin, Dilek; Sheikh, Suhail; Habip, Zafer; Yavuzer, Serap Sahin; Caliskan, Reyhan; Altun, Yagız Meric; Kuskucu, Mert; Cengiz, Mahir; Dinc, Harika Oyku; Karakullukcu, Asiye; Ergin, Sevgi; Saribas, Suat; Yilmaz, Nail; Tokman, Hrisi Bahar

    2016-01-01

    Although Legionella pneumophila serogroup 1 is the common disease causing serogroup, rare serogroups can also may cause legionellosis. A 54-year-old male patient (index case) reported that he had been on a religious trip (for visiting, tomb of Ali, which is important for Shias) to Iraq with a large group (50 shia pilgrims from Kars city of Turkey) two weeks prior to admission. Due to civil war, the hotel where the patient stayed in Iraq lacked proper hygiene. A large number of people in the travel group were experiencing the same symptoms. Other five cases were 2 males (ages; 50, 45) and 3 females including the wife of the index case (ages; 50, 28, 27). The detection of L. pneumophila IgG and IgM was performed by anti-L. pneumophila Indirect Immunofluorescent IgM, IgG kit. Legionella 1 biochip/verification BIOCHIP slides were used for serogrouping in Euroimmun AG, Leubeck, Germany. In index case, L. pneumophila IgM was positive with a titer of 1/32 titer. IgG was negative with a 1/100 titer. Another case (28 year old female), had clinical symptoms identical to the index case. L. pneumophila IgM and IgG were positive with titers of 1/64 and 1/100, respectively. These two cases were diagnosed with Legionnaires' disease caused by L. pneumophila serogroup 12 (index case) and female (28-year-old) by serogroup 11. The other 4 cases were diagnosed with possible Pontiac fever caused by L. pneumophila serogroups 14 (wife of the index case), 4, and 6 whereas the serogroup of L. pneumophila detected in 27 years old female case could not be identified. A major limitation of this work is the absence of genotyping and the serogroup difference between index case and his wife who shared the same hotel. We suggest that this serogroup difference may be caused by (for men and women) sitting separately in Islamic rules. On the other hand, the movement of people in the context of mutual visits between countries or neighboring countries for tourism-related (i.e., for religious events

  7. Detection limits of Legionella pneumophila in environmental samples after co-culture with Acanthamoeba polyphaga

    Science.gov (United States)

    2013-01-01

    Background The efficiency of recovery and the detection limit of Legionella after co-culture with Acanthamoeba polyphaga are not known and so far no investigations have been carried out to determine the efficiency of the recovery of Legionella spp. by co-culture and compare it with that of conventional culturing methods. This study aimed to assess the detection limits of co-culture compared to culture for Legionella pneumophila in compost and air samples. Compost and air samples were spiked with known concentrations of L. pneumophila. Direct culturing and co-culture with amoebae were used in parallel to isolate L. pneumophila and recovery standard curves for both methods were produced for each sample. Results The co-culture proved to be more sensitive than the reference method, detecting 102-103 L. pneumophila cells in 1 g of spiked compost or 1 m3 of spiked air, as compared to 105-106 cells in 1 g of spiked compost and 1 m3 of spiked air. Conclusions Co-culture with amoebae is a useful, sensitive and reliable technique to enrich L. pneumophila in environmental samples that contain only low amounts of bacterial cells. PMID:23442526

  8. A bacterial protein promotes the recognition of the Legionella pneumophila vacuole by autophagy.

    Science.gov (United States)

    Khweek, Arwa A; Caution, Kyle; Akhter, Anwari; Abdulrahman, Basant A; Tazi, Mia; Hassan, Hoda; Majumdar, Neal; Doran, Andrew; Guirado, Evelyn; Schlesinger, Larry S; Shuman, Howard; Amer, Amal O

    2013-05-01

    Legionella pneumophila (L. pneumophila) is an intracellular bacterium of human alveolar macrophages that causes Legionnaires' disease. In contrast to humans, most inbred mouse strains are restrictive to L. pneumophila replication. We demonstrate that autophagy targets L. pneumophila vacuoles to lysosomes and that this process requires ubiquitination of L. pneumophila vacuoles and the subsequent binding of the autophagic adaptor p62/SQSTM1 to ubiquitinated vacuoles. The L. pneumophila legA9 encodes for an ankyrin-containing protein with unknown role. We show that the legA9 mutant replicate in WT mice and their bone marrow-derived macrophages. This is the first L. pneumophila mutant to be found to replicate in WT bone marrow-derived macrophages other than the Fla mutant. Less legA9 mutant-containing vacuoles acquired ubiquitin labeling and p62/SQSTM1 staining, evading autophagy uptake and avoiding lysosomal fusion. Thus, we describe a bacterial protein that targets the L. pneumophila-containing vacuole for autophagy uptake. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Galleria mellonella apolipophorin III - an apolipoprotein with anti-Legionella pneumophila activity.

    Science.gov (United States)

    Zdybicka-Barabas, Agnieszka; Palusińska-Szysz, Marta; Gruszecki, Wiesław I; Mak, Paweł; Cytryńska, Małgorzata

    2014-10-01

    The greater wax moth Galleria mellonella has been exploited worldwide as an alternative model host for studying pathogenicity and virulence factors of different pathogens, including Legionella pneumophila, a causative agent of a severe form of pneumonia called Legionnaires' disease. An important role in the insect immune response against invading pathogens is played by apolipophorin III (apoLp-III), a lipid- and pathogen associated molecular pattern-binding protein able to inhibit growth of some Gram-negative bacteria, including Legionella dumoffii. In the present study, anti-L. pneumophila activity of G. mellonella apoLp-III and the effects of the interaction of this protein with L. pneumophila cells are demonstrated. Alterations in the bacteria cell surface occurring upon apoLp-III treatment, revealed by Fourier transform infrared (FTIR) spectroscopy and atomic force microscopy, are also documented. ApoLp-III interactions with purified L. pneumophila LPS, an essential virulence factor of the bacteria, were analysed using electrophoresis and immunoblotting with anti-apoLp-III antibodies. Moreover, FTIR spectroscopy was used to gain detailed information on the type of conformational changes in L. pneumophila LPS and G. mellonella apoLp-III induced by their mutual interactions. The results indicate that apoLp-III binding to components of bacterial cell envelope, including LPS, may be responsible for anti-L. pneumophila activity of G. mellonella apoLp-III. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. The Legionella pneumophila GIG operon responds to gold and copper in planktonic and biofilm cultures.

    Directory of Open Access Journals (Sweden)

    Kathleen Jwanoswki

    Full Text Available Legionella pneumophila contaminates man-made water systems and creates numerous exposure risks for Legionnaires' Disease. Because copper/silver ionization is commonly used to control L. pneumophila, its mechanisms of metal response and detoxification are of significant interest. Here we describe an L. pneumophila operon with significant similarity to the GIG operon of Cupriavidus metallidurans. The Legionella GIG operon is present in a subset of strains and has been acquired as part of the ICE-βox 65-kB integrative conjugative element. We assessed GIG promoter activity following exposure of L. pneumophila to multiple concentrations of HAuCl4, CuSO4 and AgNO3. At 37°C, control stationary phase cultures exhibited GIG promoter activity. This activity increased significantly in response to 20 and 50uM HAuCl4 and CuSO4 but not in response to AgNO3. Conversely, at 26°C, cultures exhibited decreased promoter response to copper. GIG promoter activity was also induced by HAuCl4 or CuSO4 during early biofilm establishment at both temperatures. When an L. pneumophila GIG promoter construct was transformed into E. coli DH5α, cultures showed baseline expression levels that did not increase following metal addition. Analysis of L. pneumophila transcriptional regulatory mutants suggested that GIG up-regulation in the presence of metal ions may be influenced by the stationary phase sigma factor, RpoS.

  11. The Legionella pneumophila GIG operon responds to gold and copper in planktonic and biofilm cultures.

    Science.gov (United States)

    Jwanoswki, Kathleen; Wells, Christina; Bruce, Terri; Rutt, Jennifer; Banks, Tabitha; McNealy, Tamara L

    2017-01-01

    Legionella pneumophila contaminates man-made water systems and creates numerous exposure risks for Legionnaires' Disease. Because copper/silver ionization is commonly used to control L. pneumophila, its mechanisms of metal response and detoxification are of significant interest. Here we describe an L. pneumophila operon with significant similarity to the GIG operon of Cupriavidus metallidurans. The Legionella GIG operon is present in a subset of strains and has been acquired as part of the ICE-βox 65-kB integrative conjugative element. We assessed GIG promoter activity following exposure of L. pneumophila to multiple concentrations of HAuCl4, CuSO4 and AgNO3. At 37°C, control stationary phase cultures exhibited GIG promoter activity. This activity increased significantly in response to 20 and 50uM HAuCl4 and CuSO4 but not in response to AgNO3. Conversely, at 26°C, cultures exhibited decreased promoter response to copper. GIG promoter activity was also induced by HAuCl4 or CuSO4 during early biofilm establishment at both temperatures. When an L. pneumophila GIG promoter construct was transformed into E. coli DH5α, cultures showed baseline expression levels that did not increase following metal addition. Analysis of L. pneumophila transcriptional regulatory mutants suggested that GIG up-regulation in the presence of metal ions may be influenced by the stationary phase sigma factor, RpoS.

  12. AUTOMATED DEAD-END ULTRAFILTRATION FOR ENHANCED SURVEILLANCE OF LEGIONELLA 2 PNEUMOPHILA AND LEGIONELLA SPP. IN COOLING TOWER WATERS

    Energy Technology Data Exchange (ETDEWEB)

    Brigmon, R.; Leskinen, S.; Kearns, E.; Jones, W.; Miller, R.; Betivas, C.; Kingsley, M.; Lim, D.

    2011-10-10

    Detection of Legionella pneumophila in cooling towers and domestic hot water systems involves concentration by centrifugation or membrane filtration prior to inoculation onto growth media or analysis using techniques such as PCR or immunoassays. The Portable Multi-use Automated Concentration System (PMACS) was designed for concentrating microorganisms from large volumes of water in the field and was assessed for enhancing surveillance of L. pneumophila at the Savannah River Site, SC. PMACS samples (100 L; n = 28) were collected from six towers between August 2010 and April 2011 with grab samples (500 ml; n = 56) being collected before and after each PMACS sample. All samples were analyzed for the presence of L. pneumophila by direct fluorescence immunoassay (DFA) using FITC-labeled monoclonal antibodies targeting serogroups 1, 2, 4 and 6. QPCR was utilized for detection of Legionella spp. in the same samples. Counts of L. pneumophila from DFA and of Legionella spp. from qPCR were normalized to cells/L tower water. Concentrations were similar between grab and PMACS samples collected throughout the study by DFA analysis (P = 0.4461; repeated measures ANOVA). The same trend was observed with qPCR. However, PMACS concentration proved advantageous over membrane filtration by providing larger volume, more representative samples of the cooling tower environment, which led to reduced variability among sampling events and increasing the probability of detection of low level targets. These data highlight the utility of the PMACS for enhanced surveillance of L. pneumophila by providing improved sampling of the cooling tower environment.

  13. First Case of Legionnaire's Disease Caused by Legionella anisa in Spain and the Limitations on the Diagnosis of Legionella non-pneumophila Infections.

    Directory of Open Access Journals (Sweden)

    Lucianna Vaccaro

    Full Text Available Legionnaires' disease is a severe form of pneumonia, with worldwide relevance, caused by Legionella spp. Approximately 90% of all cases of legionellosis are caused by Legionella pneumophila, but other species can also be responsible for this infection. These bacteria are transmitted by inhalation of aerosols or aspiration of contaminated water. In Spain, environmental studies have demonstrated the presence of Legionella non-pneumophila species in drinking water treatment plants and water distribution networks. Aware that this evidence indicates a risk factor and the lack of routine assays designed to detect simultaneously diverse Legionella species, we analyzed 210 urine samples from patients presenting clinical manifestations of pneumonia using a semi-nested PCR for partial amplification of the 16S rDNA gene of Legionella and a diagnostic method used in hospitals for Legionella antigen detection. In this study, we detected a total of 15 cases of legionellosis (7.1% and the first case of Legionnaires' disease caused by L. anisa in Spain. While the conventional method used in hospitals could only detect four cases (1.9% produced by L. pneumophila serogroup 1, using PCR, the following species were identified: Legionella spp. (10/15, L. pneumophila (4/15 and L. anisa (1/15. These results suggest the need to change hospital diagnostic strategies regarding the identification of Legionella species associated with this disease. Therefore, the detection of Legionella DNA by PCR in urine samples seems to be a suitable alternative method for a sensitive, accurate and rapid diagnosis of Legionella pneumonia, caused by L. pneumophila and also for L. non-pneumophila species.

  14. Assessment of antibiotic susceptibility of Legionella pneumophila isolated from water systems in Poland.

    Science.gov (United States)

    Sikora, Agnieszka; Gładysz, Iwona; Kozioł-Montewka, Maria; Wójtowicz-Bobin, Małgorzata; Stańczak, Tomasz; Matuszewska, Renata; Krogulska, Bożena

    2017-03-21

    Several studies have reported therapy failures in patients with legionnaires'disease; however, antimicrobial resistance of clinical and environmental isolates of Legionella spp. has not yet been documented. Routine susceptibility testing of Legionella spp. is not recommended because of difficulties in determining standard minimal inhibitory concentration values. The purpose of this study was to analyze the antimicrobial susceptibility of Legionella pneumophila. strains isolated from a water supply system. Twenty-eight isolates of L. pneumophila (16 - L. pneumophila SG 1, 12 - L. pneumophila SG 2-14) obtained from water systems in public buildings in Poland were tested. Susceptibility testing was performed using the E-test method. The tested antibiotic were azithromycin, ciprofloxacin, and rifampicin. The medium used for the susceptibility testing was BCYE-, a special medium for Legionella cultivation. Among the tested strains, L. pneumophila was the only one resistant to azithromycin. It was a strain of L. pneumophila SG 2-14 isolated from the water system in a sanitorium. All isolates were found to be sensitive to ciprofloxacin and rifampicin. However, the azithromycin-resistant strain exhibited higher ciprofloxacin and rifampicin MIC (1.5 μg/ml, and 0.19 μg/ml, respectively). The MIC50 for azithromycin, ciprofloxacin, and rifampicin were 0,032, 0,125, and 0,003 μg/ml, respectively. The MIC90 for azithromycin, ciprofloxacin, and rifampicin were 0,032, 0,125, and 0,003 μg/ml, respectively. Azithromycin resistance was found in one strain of L. pneumophila SG 2-14, but the resistance mechanism is unknown and needs further study. It is possible that therapeutic failures in Legionnaires' disease may be associated with bacterial resistance which should be taken into account. The antibiotic sensitivity testing described in this study could be helpful in detecting the resistance of clinical L. pneumophila isolates. Ciprofloxacin and rifampicin have good in vitro

  15. Application of ozonation process for the removal of Legionella pneumophila from water

    Directory of Open Access Journals (Sweden)

    Mohammad Safaee

    2015-09-01

    Full Text Available Background: Legionella pneumophila mortality and morbidity is a health concern worldwide. Due to the role of water in transmission of Legionenlla, several techniques have been used for water disinfection. This research was aimed to analyze the efficacy of ozonation process and the effects of bacterial density, contact time and pH on the removal of Legionella pneumophila from water. Methods: Legionella pneumophila was isolated from hospital water line and spiked into sterile drinking water with 300, 700 and 1000 CFU/ml densities. Ozonation was conducted within 1 L batch glass reactor with injection of 5 mg/h and contact time of 5 to 30 minutes at pH = 5, 7 and 9. Legionella culture was performed in supplemented BCYE containing GVPC and thermal treatment. After ozonation, the developed colonies were identified via biochemical and morphological tests. Results: In pH =5, the contact time 25 min and pH= 7 as well as the contact time 30 min, increase of legionella density from 300 to 1000 CFU/ml led to the reduction of removal efficiency from 100 to 87% and 100 to 82%, respectively. In pH=9 and contact time 20 min with the same bacterial density, 300 to 1000 CFU/ml, the disinfection efficacy was decreased from 100 to 91.5 %. Conclusion: Ozonation is an appropriate technique for elimination of legionella from water. The increased bacterial density led to the reduction of removal efficiency. The lowest and highest performance rates were obtained in pH=7 and 9, respectively.

  16. Legionella pneumophila persists within biofilms formed by Klebsiella pneumoniae, Flavobacterium sp., and Pseudomonas fluorescens under dynamic flow conditions.

    Directory of Open Access Journals (Sweden)

    Catherine R Stewart

    Full Text Available Legionella pneumophila, the agent of Legionnaires' disease pneumonia, is transmitted to humans following the inhalation of contaminated water droplets. In aquatic systems, L. pneumophila survives much of time within multi-organismal biofilms. Therefore, we examined the ability of L. pneumophila (clinical isolate 130 b to persist within biofilms formed by various types of aquatic bacteria, using a bioreactor with flow, steel surfaces, and low-nutrient conditions. L. pneumophila was able to intercalate into and persist within a biofilm formed by Klebsiella pneumoniae, Flavobacterium sp. or Pseudomonas fluorescens. The levels of L. pneumophila within these biofilms were as much as 4 × 10(4 CFU per cm(2 of steel coupon and lasted for at least 12 days. These data document that K. pneumoniae, Flavobacterium sp., and P. fluorescens can promote the presence of L. pneumophila in dynamic biofilms. In contrast to these results, L. pneumophila 130 b did not persist within a biofilm formed by Pseudomonas aeruginosa, confirming that some bacteria are permissive for Legionella colonization whereas others are antagonistic. In addition to colonizing certain mono-species biofilms, L. pneumophila 130 b persisted within a two-species biofilm formed by K. pneumoniae and Flavobacterium sp. Interestingly, the legionellae were also able to colonize a two-species biofilm formed by K. pneumoniae and P. aeruginosa, demonstrating that a species that is permissive for L. pneumophila can override the inhibitory effect(s of a non-permissive species.

  17. Impact of drinking water conditions and copper materials on downstream biofilm microbial communities and legionella pneumophila colonization

    Science.gov (United States)

    Legionella pneumophila, the medically important species within the genus Legionella, is a concern in engineered water systems. Its ability to amplify within free-living amoebae is well documented, but its interactions/ecology within the microbial community of drinking water biofi...

  18. Hyperoxia accelerates Fas-mediated signaling and apoptosis in the lungs of Legionella pneumophila pneumonia

    Directory of Open Access Journals (Sweden)

    Tanabe Yoshinari

    2011-04-01

    Full Text Available Abstract Background Oxygen supplementation is commonly given to the patients with severe pneumonia including Legionella disease. Recent data suggested that apoptosis may play an important role, not only in the pathogenesis of Legionella pneumonia, but also in oxygen-induced tissue damage. In the present study, the lethal sensitivity to Legionella pneumonia were compared in the setting of hyperoxia between wild-type and Fas-deficient mice. Findings C57BL/6 mice and B6.MRL-Faslpr mice characterized with Fas-deficiency were used in this study. After intratracheal administration of L. pneumophila, mice were kept in hyperoxic conditions (85-90% O2 conc. in an airtight chamber for 3 days. Bone-marrow derived macrophages infected with L. pneumophila were also kept in hyperoxic conditions. Caspase activity and cytokine production were determined by using commercially available kits. Smaller increases of several apoptosis markers, such as caspase-3 and -8, were demonstrated in Fas-deficient mice, even though the bacterial burdens in Fas-deficient and wild type mice were similar. Bone-marrow derived macrophages from Fas-deficient mice were shown to be more resistant to Legionella-induced cytotoxicity than those from wild-type mice under hyperoxia. Conclusions These results demonstrated that Fas-mediated signaling and apoptosis may be a crucial factor in the pathogenesis of Legionella pneumonia in the setting of hyperoxia.

  19. Non-opsonic phagocytosis of Legionella pneumophila by macrophages is mediated by phosphatidylinositol 3-kinase.

    Directory of Open Access Journals (Sweden)

    Souvenir D Tachado

    Full Text Available BACKGROUND: Legionella pneumophila, is an intracellular pathogen that causes Legionnaires' disease in humans, a potentially lethal pneumonia. L. pneumophila has the ability to enter and replicate in the host and is essential for pathogenesis. METHODOLOGY/PRINCIPAL FINDINGS: Phagocytosis was measured by cell invasion assays. Construction of PI3K mutant by PCR cloning and expression of dominant negative mutant was detected by Western blot. PI3K activity was measured by 32P labeling and detection of phospholipids products by thin layer chromatography. Infection of macrophages with virulent L. pneumophila stimulated the formation of phosphatidylinositol 3-phosphate (PIP3, a phosphorylated lipid product of PI3K whereas two structurally distinct phosphatidylinositol 3 kinase (PI3K inhibitors, wortmannin and LY294002, reduced L. pneumophila entry into macrophages in a dose-dependent fashion. Furthermore, PI3K activation led to Akt stimulation, a serine/threonine kinase, which was also inhibited by wortmannin and LY294002. In contrast, PI3K and protein kinase B (PKB/Akt activities were lower in macrophages infected with an avirulent bacterial strain. Only virulent L. pneumophila increased lipid kinase activity present in immunoprecipitates of the p85alpha subunit of class I PI3K and tyrosine phosphorylated proteins. In addition, macrophages expressing a specific dominant negative mutant of PI3K reduced L. pneumophila entry into these cells. CONCLUSION/SIGNIFICANCE: Entry of L. pneumophila is mediated by PI3K/Akt signaling pathway. These results suggest an important role for PI3K and Akt in the L. pneumophila infection process. They point to possible novel strategies for undermining L. pneumophila host uptake and reducing pathogenesis of Legionnaires' disease.

  20. Computed tomographic features of 23 sporadic cases with Legionella pneumophila pneumonia

    Energy Technology Data Exchange (ETDEWEB)

    Yu Hui [Department of Respiratory Diseases, Shanghai Pneumology Hospital, Tongji University, Shanghai (China); Higa, Futoshi; Hibiya, Kenji; Furugen, Makoto [Department of Medicine and Therapeutics, Control and Prevention of Infectious Diseases (First Department of Internal Medicine), Faculty of Medicine, University of the Ryukyus, Okinawa (Japan); Sato, Yoko [Tomishiro Chuo Hospital, Okinawa (Japan); Shinzato, Takashi [Nakagami General Hospital, Okinawa (Japan); Haranaga, Shusaku; Yara, Satomi; Tateyama, Masao [Department of Medicine and Therapeutics, Control and Prevention of Infectious Diseases (First Department of Internal Medicine), Faculty of Medicine, University of the Ryukyus, Okinawa (Japan); Fujita, Jiro, E-mail: fujita@med.u-ryukyu.ac.j [Department of Medicine and Therapeutics, Control and Prevention of Infectious Diseases (First Department of Internal Medicine), Faculty of Medicine, University of the Ryukyus, Okinawa (Japan); Li, Huiping [Department of Respiratory Diseases, Shanghai Pneumology Hospital, Tongji University, Shanghai (China)

    2010-06-15

    Objective: To describe the chest computed tomographic (CT) findings of Legionella pneumophila pneumonia. Methods: CT scans obtained from 23 sporadic cases of L. pneumophila pneumonia were retrospectively reviewed. Chest CT findings were analyzed with regard to the patterns and distributions of pulmonary abnormalities. We also analyzed the histopathology of lungs from guinea pigs with experimentally induced L. pneumophila pneumonia. Results: Consolidation and ground-glass opacity (GGO) were the main findings of CT scans in L. pneumophila pneumonia. The distribution of opacities was categorized as non-segmental (n = 20) and segmental (n = 4). Non-segmental distribution may follow an onset of segmental distribution. Pleural effusion was observed in 14 (58.3%) patients, of which 13 were accompanied with non-segmental distribution. Abscess formation was observed in only one immunocompromised patient. In the animal pneumonia model, the lesions comprised of terminal bronchioles, alveolar spaces, and interstitia. Small bacilli were observed to be contained by many macrophages within the alveoli. Conclusion: Non-segmental distribution was significantly more frequent than segmental distribution in L. pneumophila pneumonia. It is possible that L. pneumophila infection initially results in segmental pneumonia, which progresses to typical non-segmental distribution.

  1. Comparative study of community-acquired pneumonia caused by Streptococcus pneumoniae, Legionella pneumophila or Chlamydia pneumoniae.

    Science.gov (United States)

    Sopena, Nieves; Pedro-Botet, Maria Luisa; Sabrià, Miquel; García-Parés, Delia; Reynaga, Esteban; García-Nuñez, Marian

    2004-01-01

    The objective of this study was to compare epidemiological data and clinical presentation of community-acquired pneumonia (CAP) caused by Streptococcus pneumoniae, Legionella pneumophila or Chlamydia pneumoniae. From May 1994 to February 1996, 157 patients with S. pneumoniae (n = 68), L. pneumophila (n = 48) and C. pneumoniae (n = 41) pneumonia with definitive diagnosis, were prospectively studied. The following comparisons showed differences at a level of at least p pneumoniae pneumonia had more frequently underlying diseases (HIV infection and neoplasm) and those with C. pneumoniae pneumonia were older and had a higher frequency of chronic obstructive pulmonary disease (COPD), while L. pneumophila pneumonia prevailed in patients without comorbidity, but with alcohol intake. Presentation with cough and expectoration were significantly more frequent in patients with S. pneumoniae or C. pneumoniae pneumonia, while headache, diarrhoea and no response to betalactam antibiotics prevailed in L. pneumophila pneumonia. However, duration of symptoms > or = 7 d was more frequent in C. pneumoniae pneumonia. Patients with CAP caused by L. pneumophila presented hyponatraemia and an increase in CK more frequently, while AST elevation prevailed in L. pneumophila and C. pneumoniae pneumonia. In conclusion, some risk factors and clinical characteristics of patients with CAP may help to broaden empirical therapy against atypical pathogens until rapid diagnostic tests are available.

  2. The Identification of a Legionella pneumophila Toxin with in vivo Lethality

    Science.gov (United States)

    1980-01-01

    they cause 9. Baine WB, Rasheed JK, Maca HW, Kaufmann similar clinical illnesses. We have AF: Endotoxin activity associated with the Legionnaires... Maca HW, Kaufmann organisms that are lethal for AKR/J AF: Hemolytic activity of plasma and urine from rabbits experimentally infected with Le-mice in...disease. 11. Muller HE: Proteolytic action of Legionella pneumophila on human serum proteins. Infect Immun 27:51-53, 1980. 12. Friedman RL, Iglewski BH

  3. Detection of Legionella, L. pneumophila and Mycobacterium Avium Complex (MAC along Potable Water Distribution Pipelines

    Directory of Open Access Journals (Sweden)

    Harriet Whiley

    2014-07-01

    Full Text Available Inhalation of potable water presents a potential route of exposure to opportunistic pathogens and hence warrants significant public health concern. This study used qPCR to detect opportunistic pathogens Legionella spp., L. pneumophila and MAC at multiple points along two potable water distribution pipelines. One used chlorine disinfection and the other chloramine disinfection. Samples were collected four times over the year to provide seasonal variation and the chlorine or chloramine residual was measured during collection. Legionella spp., L. pneumophila and MAC were detected in both distribution systems throughout the year and were all detected at a maximum concentration of 103 copies/mL in the chlorine disinfected system and 106, 103 and 104 copies/mL respectively in the chloramine disinfected system. The concentrations of these opportunistic pathogens were primarily controlled throughout the distribution network through the maintenance of disinfection residuals. At a dead-end and when the disinfection residual was not maintained significant (p < 0.05 increases in concentration were observed when compared to the concentration measured closest to the processing plant in the same pipeline and sampling period. Total coliforms were not present in any water sample collected. This study demonstrates the ability of Legionella spp., L. pneumophila and MAC to survive the potable water disinfection process and highlights the need for greater measures to control these organisms along the distribution pipeline and at point of use.

  4. Detection of Legionella, L. pneumophila and Mycobacterium avium complex (MAC) along potable water distribution pipelines.

    Science.gov (United States)

    Whiley, Harriet; Keegan, Alexandra; Fallowfield, Howard; Bentham, Richard

    2014-07-18

    Inhalation of potable water presents a potential route of exposure to opportunistic pathogens and hence warrants significant public health concern. This study used qPCR to detect opportunistic pathogens Legionella spp., L. pneumophila and MAC at multiple points along two potable water distribution pipelines. One used chlorine disinfection and the other chloramine disinfection. Samples were collected four times over the year to provide seasonal variation and the chlorine or chloramine residual was measured during collection. Legionella spp., L. pneumophila and MAC were detected in both distribution systems throughout the year and were all detected at a maximum concentration of 103 copies/mL in the chlorine disinfected system and 106, 103 and 104 copies/mL respectively in the chloramine disinfected system. The concentrations of these opportunistic pathogens were primarily controlled throughout the distribution network through the maintenance of disinfection residuals. At a dead-end and when the disinfection residual was not maintained significant (p coliforms were not present in any water sample collected. This study demonstrates the ability of Legionella spp., L. pneumophila and MAC to survive the potable water disinfection process and highlights the need for greater measures to control these organisms along the distribution pipeline and at point of use.

  5. [Construction and Expression of Vector with mip/flaA Advantages Epitope Genes of Legionella pneumophila].

    Science.gov (United States)

    He, Jin-lei; Peng, Ran; Zhang, Jun-rung; Yu Ze-ying; Li, Na-li-sha; Gong, Yan-ju; Chen, Ying; Chen, Da-li; Chen, Jian-ping

    2016-01-01

    To generate and express fusion vector with mip/flaA advantages epitope genes of Legionella pneumophila by select mip and flaA advantages epitope genes for future research on Legionella pneumophila protein vaccine. Following analysis of secondary structure and surface properties such as: physical and chemical properties, hydropathy, plasticity, antigen index and extracellular domain of Mip and FlaA proteins by bioinformatics methods, the region which active epitope may exist was selected as advantages epitope region. Then, the recombinant plasmid pET-mip, pET-flaA and pET-mip/flaA with advantages epitope genes were constructed by PCR amplification and T4 ligase connection, and induced the expression in E. coli. Many potential antigenic epitopes in Mip and FlaA were identified, and the selected advantages epitope regions were cloned and expressed successfully. Moreover, the mip/flaA two advantages associated epitope fusion proteins were also successfully expressed. DNA Star software and Expasy online analysis system can successfully predict antigenic epitopes for Legionella pneumophila Mip and FlaA. And prokaryotic expression vector pET-mip/flaA with advantages epitope genes has been successfully constructed and efficiently expressed.

  6. Isolation and identification of Legionella pneumophila from drinking water in Basra governorate, Iraq.

    Science.gov (United States)

    Al-Sulami, A A; Al-Taee, A M R; Yehyazarian, A A

    2013-11-01

    This study in Iraq investigated the occurrence of Legionella. pneumophila in different drinking-water sources in Basra governorate as well as the susceptibility of isolates to several antibiotics. A total of 222 water samples were collected in 2008-2009: 49 samples from water purification plants (at entry points, from precipitation tanks, from filtration tanks and at exit points), 127 samples of tap water; and 46 samples from tankers and plants supplying water by reverse osmosis. The findings confirmed the presence of L. pneumophila in sources of crude water, in general drinking water supplies and drinking water tankers. Of 258 isolates 77.1% were serotype 1 and 22.9% serotypes 2-15. All examined isolates displayed drug resistance, particularly to ampicillin, but were 100% susceptible to doxycycline. The prevalence of L. pneumophila, especially serogroup 1, is a strong indicator of unsuitability of drinking water and requires appropriate action.

  7. Legionella pneumophila type IV effectors hijack the transcription and translation machinery of the host cell.

    Science.gov (United States)

    Rolando, Monica; Buchrieser, Carmen

    2014-12-01

    Intracellular bacterial pathogens modulate the host response to persist and replicate inside a eukaryotic cell and cause disease. Legionella pneumophila, the causative agent of Legionnaires' disease, is present in freshwater environments and represents one of these pathogens. During coevolution with protozoan cells, L. pneumophila has acquired highly sophisticated and diverse strategies to hijack host cell processes. It secretes hundreds of effectors into the host cell, and these manipulate host signaling pathways and key cellular processes. Recently it has been shown that L. pneumophila is also able to alter the transcription and translation machinery of the host and to exploit epigenetic mechanisms in the cells it resides in to counteract host responses. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. Legionella pneumophila serogroup 5 infection in the presence of multiple environmental contamination. The importance of a bacteriological diagnosis

    Directory of Open Access Journals (Sweden)

    Maria Teresa Montagna

    2007-03-01

    Full Text Available Legionella pneumophila is a pathogen that causes severe pneumonia in humans; L. pneumophila serogroup 1 accounts for at least 90% of infections. This is not linked to an environmental predominance of Legionella pneumophila 1, but may be due to a greater virulence of the strain. L. pneumophila sg 5 has also been reported, albeit less frequently, to be a cause of the disease. We report a case of L. pneumophila sg 5 occurring in a large hospital in southern Italy (Apulia region, where both L. pneumophila sg 1 and sg 5 were detected in the water supply; the nosocomial origin was demonstrated by molecular subtyping (PFGE. An environmental investigation, performed immediately after diagnosis of the case of legionellosis, identified a ow L. pneumophila sg 5 contamination level. Our experience highlights that in hospital, risk assessment, in rder to institute control measures for Legionella, should be carried out not only in response to a case of the disease and/or in risk wards only, as described in the Italian Guidelines, but periodically in every ward. The present study confirms that, although in the community L. pneumophila sg 1 is the most frequent strain isolated in both outbreaks and isolated cases, in hospital other serogroups and species may often cause infection because of the high susceptibility of the hosts.

  9. Side-polished fiber immunosensor based on surface plasmon resonance for detection of Legionella pneumophila

    Science.gov (United States)

    Tsao, Yu-Chia; Yang, Yi-Wen; Tsai, Woo-Hu; Yan, Tsong-Rong

    2008-02-01

    Side-polished fiber immunosensor based on surface plasmon resonance (SPR) onto self-assembled protein A layer was proposed for the detection of Legionella pneumophila. A self-assembled protein A layer on gold (Au) surface was fabricated by adsorbing a mixture of 11-mercaptoundecanoic acid (MUA) and activated by N-Ethyl-N'-(3-dimethylaminopropyl) carbodiimide/ N-Hydroxysuccinimide (EDC/NHS). The formation of self-assembled protein A and gold layer on side-polished surface and the binding of antibody and antigen in series were confirmed by SPR response on spectrum. The binding protein A layer can improve the sensitivity, which indirectly supports the configurations that antibody layer is immobilized on the binding protein A layer with a well-ordered orientation. The surface morphology analyses of self-assembled protein A layer on Au substrate and monoclonal antibody against L. pneumophila immobilized on protein A were demonstrated by SPR dip shifts on optical spectrum analyzer. The SPR fiber immunosensor for detection of L. pneumophila was developed and the detection limit was 10 CFU/ml with the SPR dip shift in wavelength from 1070 to 1105nm. The current fabrication technique of a SPR immunosensor using optical fiber for the detection of Legionella pneumophila could be applied to construct other biosensor.

  10. Invasion of Eukaryotic Cells by Legionella Pneumophila: A Common Strategy for all Hosts?

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    Paul S Hoffman

    1997-01-01

    Full Text Available Legionella pneumophila is an environmental micro-organism capable of producing an acute lobar pneumonia, commonly referred to as Legionnaires’ disease, in susceptible humans. Legionellae are ubiquitous in aquatic environments, where they survive in biofilms or intracellularly in various protozoans. Susceptible humans become infected by breathing aerosols laden with the bacteria. The target cell for human infection is the alveolar macrophage, in which the bacteria abrogate phagolysosomal fusion. The remarkable ability of L pneumophila to infect a wide range of eukaryotic cells suggests a common strategy that exploits very fundamental cellular processes. The bacteria enter host cells via coiling phagocytosis and quickly subvert organelle trafficking events, leading to formation of a replicative phagosome in which the bacteria multiply. Vegetative growth continues for 8 to 10 h, after which the bacteria develop into a short, highly motile form called the ‘mature form’. The mature form exhibits a thickening of the cell wall, stains red with the Gimenez stain, and is between 10 and 100 times more infectious than agar-grown bacteria. Following host cell lysis, the released bacteria infect other host cells, in which the mature form differentiates into a Gimenez-negative vegetative form, and the cycle begins anew. Virulence of L pneumophila is considered to be multifactorial, and there is growing evidence for both stage specific and sequential gene expression. Thus, L pneumophila may be a good model system for dissecting events associated with the host-parasite interactions.

  11. Post-translational modifications are key players of the Legionella pneumophila infection strategy

    Science.gov (United States)

    Michard, Céline; Doublet, Patricia

    2015-01-01

    Post-translational modifications (PTMs) are widely used by eukaryotes to control the enzymatic activity, localization or stability of their proteins. Traditionally, it was believed that the broad biochemical diversity of the PTMs is restricted to eukaryotic cells, which exploit it in extensive networks to fine-tune various and complex cellular functions. During the last decade, the advanced detection methods of PTMs and functional studies of the host–pathogen relationships highlight that bacteria have also developed a large arsenal of PTMs, particularly to subvert host cell pathways to their benefit. Legionella pneumophila, the etiological agent of the severe pneumonia legionellosis, is the paradigm of highly adapted intravacuolar pathogens that have set up sophisticated biochemical strategies. Among them, L. pneumophila has evolved eukaryotic-like and rare/novel PTMs to hijack host cell processes. Here, we review recent progress about the diversity of PTMs catalyzed by Legionella: ubiquitination, prenylation, phosphorylation, glycosylation, methylation, AMPylation, and de-AMPylation, phosphocholination, and de-phosphocholination. We focus on the host cell pathways targeted by the bacteria catalyzed PTMs and we stress the importance of the PTMs in the Legionella infection strategy. Finally, we highlight that the discovery of these PTMs undoubtedly made significant breakthroughs on the molecular basis of Legionella pathogenesis but also lead the way in improving our knowledge of the eukaryotic PTMs and complex cellular processes that are associated to. PMID:25713573

  12. Survey of Legionella pneumophila among pneumonia patients at ...

    African Journals Online (AJOL)

    Objective: To determine the occurrence of L. pneumophila among pneumonia patients at Kenyatta National Hospital and any association with possible risk factors. Design: A cross- sectional descriptive study. Setting: The study was conducted from March to June 2007, at the medical ward of Kenyatta National Hospital.

  13. Master manipulators: an update on Legionella pneumophila Icm/Dot translocated substrates and their host targets

    Science.gov (United States)

    Isaac, Dervla T; Isberg, Ralph

    2014-01-01

    Macrophages are the front line of immune defense against invading microbes. Microbes, however, have evolved numerous and diverse mechanisms to thwart these host immune defenses and thrive intracellularly. Legionella pneumophila, a Gram-negative pathogen of amoebal and mammalian phagocytes, is one such microbe. In humans, it causes a potentially fatal pneumonia referred to as Legionnaires' disease. Armed with the Icm/Dot type IV secretion system, which is required for virulence, and approximately 300 translocated proteins, Legionella is able to enter host cells, direct the biogenesis of its own vacuolar compartment, and establish a replicative niche, where it grows to high levels before lysing the host cell. Efforts to understand the pathogenesis of this bacterium have focused on characterizing the molecular activities of its many effectors. In this article, we highlight recent strides that have been made in understanding how Legionella effectors mediate host-pathogen interactions. PMID:24762308

  14. Severe Community-acquired Pneumonia Due to Legionella pneumophila Serogroup 6

    Directory of Open Access Journals (Sweden)

    Chung-Yu Chen

    2006-01-01

    Full Text Available Legionella pneumophila is a common cause of sporadic community-acquired pneumonia, but culture-proven legionellosis is rarely diagnosed. There is no laboratory test for Legionnaires' disease that can detect all patients with the disease. Culture is the standard diagnostic method and should be initiated as soon as possible in suspected cases. We describe a rare case of community-acquired pneumonia caused by L. pneumophila serogroup 6. A 77-year-old man was admitted to a tertiary care hospital because of high fever, productive cough, and progressive dyspnea. Chest radiography showed bilateral pneumonia, which led to respiratory failure necessitating mechanical ventilatory support. Despite antibiotic therapy, his condition continued to deteriorate and acute renal failure also developed. Urine was negative for L. pneumophila. Culture of the sputum yielded L. pneumophila serogroup 6, although there was no elevation of the serum antibody titer. Pneumonia resolved gradually and he was extubated after treatment with levofloxacin followed by erythromycin. L. pneumophila other than serogroup 1 should be included in the differential diagnosis of patients with suspected atypical community-acquired pneumonia.

  15. The transcriptome of Legionella pneumophila-infected human monocyte-derived macrophages.

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    Christopher T D Price

    Full Text Available Legionella pneumophila is an intracellular bacterial pathogen that invades and replicates within alveolar macrophages through injection of ∼ 300 effector proteins by its Dot/Icm type IV translocation apparatus. The bona fide F-box protein, AnkB, is a nutritional virulence effector that triggers macrophages to generate a surplus of amino acids, which is essential for intravacuolar proliferation. Therefore, the ankB mutant represents a novel genetic tool to determine the transcriptional response of human monocyte-derived macrophages (hMDMs to actively replicating L. pneumophila.Here, we utilized total human gene microarrays to determine the global transcriptional response of hMDMs to infection by wild type or the ankB mutant of L. pneumophila. The transcriptomes of hMDMs infected with either actively proliferating wild type or non-replicative ankB mutant bacteria were remarkably similar. The transcriptome of infected hMDMs was predominated by up-regulation of inflammatory pathways (IL-10 anti-inflammatory, interferon signaling and amphoterin signaling, anti-apoptosis, and down-regulation of protein synthesis pathways. In addition, L. pneumophila modulated diverse metabolic pathways, particularly those associated with bio-active lipid metabolism, and SLC amino acid transporters expression.Taken together, the hMDM transcriptional response to L. pneumophila is independent of intra-vacuolar replication of the bacteria and primarily involves modulation of the immune response and metabolic as well as nutritional pathways.

  16. Inhibitors for the bacterial ectonucleotidase Lp1NTPDase from Legionella pneumophila.

    Science.gov (United States)

    Fiene, Amelie; Baqi, Younis; Malik, Enas M; Newton, Patrice; Li, Wenjin; Lee, Sang-Yong; Hartland, Elizabeth L; Müller, Christa E

    2016-09-15

    Legionella pneumophila is an aerobic, Gram-negative bacterium of the genus Legionella, which constitutes the major causative agent of Legionnaires' disease. Recently a nucleoside triphosphate diphosphohydrolase (NTPDase) from L. pneumophila was identified and termed Lp1NTPDase; it was found to be a structural and functional homolog of mammalian NTPDases catalyzing the hydrolysis of ATP to ADP and ADP to AMP. Its activity is believed to contribute to the virulence of Legionella pneumophila. Therefore Lp1NTPDase inhibitors are considered as novel antibacterial drugs. However, only weakly potent compounds are available so far. In the present study, a capillary electrophoresis (CE)-based enzyme assay for monitoring the Lp1NTPDase activity was established. The enzymatic reaction was performed in a test tube followed by separation of substrate and products by CE and subsequent quantification by UV analysis. After kinetic characterization of the enzyme, a series of 1-amino-4-ar(alk)ylamino-2-sulfoanthraquinone derivatives structurally related to the anthraquinone dye Reactive Blue 2, a non-selective ecto-NTPDase inhibitor, was investigated for inhibitory activity on Lp1NTPDase using the CE-based enzyme assay. Derivatives bearing a large lipophilic substituent (e.g., fused aromatic rings) in the 4-position of the 1-amino-2-sulfoanthraquinone showed the highest inhibitory activity. Compounds with IC50 values in the low micromolar range were identified. The most potent inhibitor was 1-amino-4-[phenanthrene-9-yl-amino]-9,10-dioxo-9,10-dihydroanthracene-2-sulfonate (28, PSB-16131), with an IC50-value of 4.24μM. It represents the most potent Lp1NTPDase inhibitor described to date. These findings may serve as a starting point for further optimization. Lp1NTPDase inhibition provides a novel approach for the (immuno)therapy of Legionella infections. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. Dendrimers and polyamino-phenolic ligands: activity of new molecules against Legionella pneumophila biofilms.

    Directory of Open Access Journals (Sweden)

    Elisa eAndreozzi

    2016-03-01

    Full Text Available Legionnaires’ disease is a potentially fatal pneumonia caused by Legionella pneumophila, an aquatic bacterium often found within the biofilm niche. In man-made water systems microbial biofilms increase the resistance of legionella to disinfection, posing a significant threat to public health. Disinfection methods currently used in water systems have been shown to be ineffective against legionella over the long-term, allowing recolonization by the biofilm-protected microorganisms. In this study, the anti-biofilm activity of previously fabricated polyamino-phenolic ligands and polyamidoamine dendrimers was investigated against legionella mono-species and multi-species biofilms formed by L. pneumophila in association with other bacteria that can be found in tap water (Aeromonas hydrophila, Pseudomonas aeruginosa, Escherichia coli, Klebsiella pneumoniae. Bacterial ability to form biofilms was verified using a crystal violet colorimetric assay and testing cell viability by real-time quantitative PCR and Plate Count assay. The concentration of the chemicals tested as anti-biofilm agents was chosen based on cytotoxicity assays: the highest non-cytotoxic chemical concentration was used for biofilm inhibition assays, with dendrimer concentration ten-fold higher than polyamino-phenolic ligands. While Macrophen and Double Macrophen were the most active substances among polyamino-phenolic ligands, dendrimers were overall two-fold more effective than all other compounds with a reduction up to 85% and 73% of legionella and multi-species biofilms, respectively. Chemical interaction with matrix molecules is hypothesized, based on SEM images and considering the low or absent anti-microbial activity on planktonic bacteria showed by flow cytometry. These data suggest that the studied compounds, especially dendrimers, could be considered as novel molecules in the design of research projects aimed at the development of efficacious anti-biofilm disinfection

  18. Legionella pneumophila catalase-peroxidases are required for proper trafficking and growth in primary macrophages.

    Science.gov (United States)

    Bandyopadhyay, Purnima; Byrne, Brenda; Chan, Yolande; Swanson, Michele S; Steinman, Howard M

    2003-08-01

    Legionella pneumophila, a parasite of aquatic amoebae and pathogen of pulmonary macrophages, replicates intracellularly, utilizing a type IV secretion system to subvert the trafficking of Legionella-containing phagosomes. Defense against host-derived reactive oxygen species has been proposed as critical for intracellular replication. Virulence traits of null mutants in katA and katB, encoding the two Legionella catalase-peroxidases, were analyzed to evaluate the hypothesis that L. pneumophila must decompose hydrogen peroxide to establish a replication niche in macrophages. Phagosomes containing katA or katB mutant Legionella colocalize with LAMP-1, a late endosomal-lysosomal marker, at twice the frequency of those of wild-type strain JR32 and show a decreased frequency of bacterial replication, in similarity to phenotypes of mutants with mutations in dotA and dotB, encoding components of the Type IV secretion system. Quantitative similarity of the katA/B phenotypes indicates that each contributes to virulence traits largely independently of intracellular compartmentalization (KatA in the periplasm and KatB in the cytosol). These data support a model in which KatA and KatB maintain a critically low level of H(2)O(2) compatible with proper phagosome trafficking mediated by the type IV secretion apparatus. During these studies, we observed that dotA and dotB mutations in wild-type strain Lp02 had no effect on intracellular multiplication in the amoeba Acanthamoeba castellanii, indicating that certain dotA/B functions in Lp02 are dispensable in that experimental model. We also observed that wild-type JR32, unlike Lp02, shows minimal contact-dependent cytotoxicity, suggesting that cytotoxicity of JR32 is not a prerequisite for formation of replication-competent Legionella phagosomes in macrophages.

  19. Dot/Icm Effector Translocation by Legionella longbeachae Creates a Replicative Vacuole Similar to That of Legionella pneumophila despite Translocation of Distinct Effector Repertoires.

    Science.gov (United States)

    Wood, Rebecca E; Newton, Patrice; Latomanski, Eleanor A; Newton, Hayley J

    2015-10-01

    Legionella organisms are environmental bacteria and accidental human pathogens that can cause severe pneumonia, termed Legionnaires' disease. These bacteria replicate within a pathogen-derived vacuole termed the Legionella-containing vacuole (LCV). Our understanding of the development and dynamics of this vacuole is based on extensive analysis of Legionella pneumophila. Here, we have characterized the Legionella longbeachae replicative vacuole (longbeachae-LCV) and demonstrated that, despite important genomic differences, key features of the replicative LCV are comparable to those of the LCV of L. pneumophila (pneumophila-LCV). We constructed a Dot/Icm-deficient strain by deleting dotB and demonstrated the inability of this mutant to replicate inside THP-1 cells. L. longbeachae does not enter THP-1 cells as efficiently as L. pneumophila, and this is reflected in the observation that translocation of BlaM-RalFLLO (where RalFLLO is the L. longbeachae homologue of RalF) into THP-1 cells by the L. longbeachae Dot/Icm system is less efficient than that by L. pneumophila. This difference is negated in A549 cells where L. longbeachae and L. pneumophila infect with similar entry dynamics. A β-lactamase assay was employed to demonstrate the translocation of a novel family of proteins, the Rab-like effector (Rle) proteins. Immunofluorescence analysis confirmed that these proteins enter the host cell during infection and display distinct subcellular localizations, with RleA and RleC present on the longbeachae-LCV. We observed that the host Rab GTPase, Rab1, and the v-SNARE Sec22b are also recruited to the longbeachae-LCV during the early stages of infection, coinciding with the LCV avoiding endocytic maturation. These studies further our understanding of the L. longbeachae replicative vacuole, highlighting phenotypic similarities to the vacuole of L. pneumophila as well as unique aspects of LCV biology. Copyright © 2015, American Society for Microbiology. All Rights

  20. Rapid on-site monitoring of Legionella pneumophila in cooling tower water using a portable microfluidic system.

    Science.gov (United States)

    Yamaguchi, Nobuyasu; Tokunaga, Yusuke; Goto, Satoko; Fujii, Yudai; Banno, Fumiya; Edagawa, Akiko

    2017-06-08

    Legionnaires' disease, predominantly caused by the bacterium Legionella pneumophila, has increased in prevalence worldwide. The most common mode of transmission of Legionella is inhalation of contaminated aerosols, such as those generated by cooling towers. Simple, rapid and accurate methods to enumerate L. pneumophila are required to prevent the spread of this organism. Here, we applied a microfluidic device for on-chip fluorescent staining and semi-automated counting of L. pneumophila in cooling tower water. We also constructed a portable system for rapid on-site monitoring and used it to enumerate target bacterial cells rapidly flowing in the microchannel. A fluorescently-labelled polyclonal antibody was used for the selective detection of L. pneumophila serogroup 1 in the samples. The counts of L. pneumophila in cooling tower water obtained using the system and fluorescence microscopy were similar. The detection limit of the system was 10 4  cells/ml, but lower numbers of L. pneumophila cells (10 1 to 10 3  cells/ml) could be detected following concentration of 0.5-3 L of the water sample by filtration. Our technique is rapid to perform (1.5 h), semi-automated (on-chip staining and counting), and portable for on-site measurement, and it may therefore be effective in the initial screening of Legionella contamination in freshwater.

  1. Monitoring of Legionella pneumophila viability after chlorine dioxide treatment using flow cytometry.

    Science.gov (United States)

    Mustapha, Pascale; Epalle, Thibaut; Allegra, Séverine; Girardot, Françoise; Garraud, Olivier; Riffard, Serge

    2015-04-01

    The viability of three Legionella pneumophila strains was monitored after chlorine dioxide (ClO2) treatment using a flow cytometric assay. Suspensions of L. pneumophila cells were submitted to increasing concentrations of ClO2. Culturable cells were still detected when using 4 mg/L, but could no longer be detected after exposure to 6 mg/L of ClO2, although viable but not culturable (VBNC) cells were found after exposure to 4-5 mg/L of ClO2. When testing whether these VBNC were infective, two of the strains were resuscitated after co-culture with Acanthamoeba polyphaga, but neither of them could infect macrophage-like cells. Copyright © 2015 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  2. Recombinant Mip-PilE-FlaA dominant epitopes vaccine candidate against Legionella pneumophila.

    Science.gov (United States)

    He, Jinlei; Huang, Fan; Chen, Han; Chen, Qiwei; Zhang, Junrong; Li, Jiao; Chen, Dali; Chen, Jianping

    2017-06-01

    Legionella pneumophila is the main causative agent of Legionnaires' disease, which is a severe multi-system disease with pneumonia as the primary manifestation. We designed a recombinant Mip-PilE-FlaA dominant epitopes vaccine against Legionella pneumophila to prevent the disease and evaluated its immunogenicity and protective immunity. The protein structures of Mip, PilE and FlaA were analyzed using a computer, and the gene sequences of the dominant epitopes of the three proteins were selected to construct and optimize the vaccine. The optimized mip, pilE, flaA and recombinant mip-pilE-flaA gene sequences were cloned, expressed and purified. The purified proteins were used as dominant epitopes vaccines to immunize BALB/c mice and determine the protective immunity and immunogenicity of these purified proteins. The identification confirmed that the recombinant mip-pilE-flaA was successfully cloned and expressed. ELISA revealed that the Mip-PilE-FlaA group produced the highest IgG response, and this protein may considerably improve the production of some cytokines in BALB/c mice. Histopathology analyses of lungs from mice immunized with Mip-PilE-FlaA revealed a certain protective effect. Our work demonstrated that the recombinant dominant epitopes of Mip-PilE-FlaA exhibited strong immunogenicity and immune protection, and this protein may be an efficient epitopes vaccine candidate against Legionella pneumophila. Copyright © 2017 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.

  3. Molecular evolution of key genes for type II secretion in Legionella pneumophila.

    Science.gov (United States)

    Costa, Joana; d'Avó, Ana Filipa; da Costa, Milton S; Veríssimo, António

    2012-08-01

    Given the role of type II protein secretion system (T2S) in the ecology and pathogenesis of Legionella pneumophila, it is possible that this system is a target for adaptive evolution. The population genetic structure of L.pneumophila was inferred from the partial sequences of rpoB and from the complete sequence of three T2S structural components (lspD, lspE and pilD) and from two T2S effectors critical for intracellular infection of protozoa (proA and srnA) of 37 strains isolated from natural and man-made environments and disease-related from worldwide sources. A phylogenetic analysis was obtained for the concatenated alignment and for each individual locus. Seven main groups were identified containing the same L.pneumophila strains, suggesting an ancient divergence for each cluster and indicating that the operating selective pressures have equally affected the evolution of the five genes. Although linkage disequilibrium analysis indicate a clonal nature for population structure in this sample, our results indicate that recombination is a common phenomenon among T2S-related genes on this species, as 24 of the 37 L.pneumophila isolates contained at least one locus in which recombination was identified. Furthermore, neutral selection acting on the analysed T2S-related genes emerged as a clear result, namely on T2S effectors, ProA and SrnA, indicating that they are probably implicated in conserved virulence mechanisms through legionellae hosts. © 2011 Society for Applied Microbiology and Blackwell Publishing Ltd.

  4. Promotion and Rescue of Intracellular Brucella neotomae Replication during Coinfection with Legionella pneumophila.

    Science.gov (United States)

    Kang, Yoon-Suk; Kirby, James E

    2017-05-01

    We established a new Brucella neotomae in vitro model system for study of type IV secretion system-dependent (T4SS) pathogenesis in the Brucella genus. Importantly, B. neotomae is a rodent pathogen, and unlike B. abortus , B. melitensis , and B. suis , B. neotomae has not been observed to infect humans. It therefore can be handled more facilely using biosafety level 2 practices. More particularly, using a series of novel fluorescent protein and lux operon reporter systems to differentially label pathogens and track intracellular replication, we confirmed T4SS-dependent intracellular growth of B. neotomae in macrophage cell lines. Furthermore, B. neotomae exhibited early endosomal (LAMP-1) and late endoplasmic reticulum (calreticulin)-associated phagosome maturation. These findings recapitulate prior observations for human-pathogenic Brucella spp. In addition, during coinfection experiments with Legionella pneumophila , we found that defective intracellular replication of a B. neotomae T4SS virB4 mutant was rescued and baseline levels of intracellular replication of wild-type B. neotomae were significantly stimulated by coinfection with wild-type but not T4SS mutant L. pneumophila Using confocal microscopy, it was determined that intracellular colocalization of B. neotomae and L. pneumophila was required for rescue and that colocalization came at a cost to L. pneumophila fitness. These findings were not completely expected based on known temporal and qualitative differences in the intracellular life cycles of these two pathogens. Taken together, we have developed a new system for studying in vitro Brucella pathogenesis and found a remarkable T4SS-dependent interplay between Brucella and Legionella during macrophage coinfection. Copyright © 2017 American Society for Microbiology.

  5. Phase-variable Expression of Lipopolysaccharide Contributes to the Virulence of Legionella pneumophila

    OpenAIRE

    Lüneberg, Edeltraud; Zähringer, Ulrich; Knirel, Yuriy A; Steinmann, Dorothee; Hartmann, Maike; Steinmetz, Ivo; Rohde, Manfred; Köhl, Jörg; Frosch, Matthias

    1998-01-01

    With the aid of monoclonal antibody (mAb) 2625, raised against the lipopolysaccharide (LPS) of Legionella pneumophila serogroup 1, subgroup OLDA, we isolated mutant 811 from the virulent wild-type strain RC1. This mutant was not reactive with mAb 2625 and exhibited an unstable phenotype, since we observed an in vitro and in vivo switch of mutant 811 to the mAb 2625–positive phenotype, thus restoring the wild-type LPS. Bactericidal assays revealed that mutant 811 was lysed by serum complement ...

  6. Disulfide Bond Oxidoreductase DsbA2 of Legionella pneumophila Exhibits Protein Disulfide Isomerase Activity

    OpenAIRE

    Kpadeh, Zegbeh Z.; Jameson-Lee, Max; Anthony J. Yeh; Chertihin, Olga; Shumilin, Igor A.; Dey, Rafik; Day, Shandra R.; Hoffman, Paul S.

    2013-01-01

    The extracytoplasmic assembly of the Dot/Icm type IVb secretion system (T4SS) of Legionella pneumophila is dependent on correct disulfide bond (DSB) formation catalyzed by a novel and essential disulfide bond oxidoreductase DsbA2 and not by DsbA1, a second nonessential DSB oxidoreductase. DsbA2, which is widely distributed in the microbial world, is phylogenetically distinct from the canonical DsbA oxidase and the DsbC protein disulfide isomerase (PDI)/reductase of Escherichia coli. Here we s...

  7. Legionella pneumophila infection of Drosophila S2 cells induces only minor changes in mitochondrial dynamics.

    Directory of Open Access Journals (Sweden)

    Elizabeth Wen Sun

    Full Text Available During infection of cells by Legionella pneumophila, the bacterium secretes a large number of effector proteins into the host cell cytoplasm, allowing it to alter many cellular processes and make the vacuole and the host cell into more hospitable environments for bacterial replication. One major change induced by infection is the recruitment of ER-derived vesicles to the surface of the vacuole, where they fuse with the vacuole membrane and prevent it from becoming an acidified, degradative compartment. However, the recruitment of mitochondria to the region of the vacuole has also been suggested by ultrastructural studies. In order to test this idea in a controlled and quantitative experimental system, and to lay the groundwork for a genome-wide screen for factors involved in mitochondrial recruitment, we examined the behavior of mitochondria during the early stages of Legionella pneumophila infection of Drosophila S2 cells. We found that the density of mitochondria near vacuoles formed by infection with wild type Legionella was not different from that found in dotA(- mutant-infected cells during the first 4 hours after infection. We then examined 4 parameters of mitochondrial motility in infected cells: velocity of movement, duty cycle of movement, directional persistence and net direction. In the 4 hours following infection, most of these measures were indistinguishable between wild type and dotA(-.infection. However, wild type Legionella did induce a modest shift in the velocity distribution toward faster movement compared dotA(- infection, and a small downward shift in the duty cycle distribution. In addition, wild type infection produced mitochondrial movement that was biased in the direction of the bacterial vacuole relative to dotA-, although not enough to cause a significant accumulation within 10 um of the vacuole. We conclude that in this host cell, mitochondria are not strongly recruited to the vacuole, nor is their motility

  8. AMPylation Is Critical for Rab1 Localization to Vacuoles Containing Legionella pneumophila

    Science.gov (United States)

    Hardiman, Camille A.; Roy, Craig R.

    2014-01-01

    ABSTRACT Legionella pneumophila is an intracellular pathogen that resides within a membrane-bound compartment that is derived from vesicles exiting the endoplasmic reticulum (ER). To create this compartment, these bacteria use a type IV secretion system to deliver effector proteins that subvert host cell functions. Several Legionella effector proteins modulate the function of the host protein Rab1, which is a GTPase that is recruited to the Legionella-containing vacuole (LCV). Here, we examined which of the Rab1-directed enzymatic activities displayed by Legionella effectors are important for localizing the Rab1 protein to the LCV membrane. The guanine nucleotide exchange factor (GEF) domain in the effector protein DrrA (SidM) was essential for Rab1 recruitment to the LCV and Rab1 AMPylation by the nucleotidyltransferase domain in DrrA was important for Rab1 retention. Legionella organisms producing mutant DrrA proteins that were severely attenuated for GEF activity in vitro retained the ability to localize Rab1 to the LCV. Rab1 localization to the LCV mediated by these GEF-defective mutants required AMPylation. Importantly, we found that efficient localization of Rab1 to the LCV occurred when Rab1 GEF activity and Rab1 AMPylation activity were provided by separate proteins. Rab1 phosphocholination (PCylation) by the effector protein AnkX, however, was unable to substitute for Rab1 AMPylation. Lastly, the defect in Rab1 localization to the LCV in AMPylation-deficient strains of Legionella was partially suppressed if the GTPase-activating protein (GAP) LepB was eliminated. Thus, our data indicate that AMPylation of Rab1 is an effective strategy to maintain this GTPase on the LCV membrane. PMID:24520063

  9. Identification of host cytosolic sensors and bacterial factors regulating the type I interferon response to Legionella pneumophila.

    Directory of Open Access Journals (Sweden)

    Kathryn M Monroe

    2009-11-01

    Full Text Available Legionella pneumophila is a gram-negative bacterial pathogen that replicates in host macrophages and causes a severe pneumonia called Legionnaires' Disease. The innate immune response to L. pneumophila remains poorly understood. Here we focused on identifying host and bacterial factors involved in the production of type I interferons (IFN in response to L. pneumophila. It was previously suggested that the delivery of L. pneumophila DNA to the host cell cytosol is the primary signal that induces the type I IFN response. However, our data are not easily reconciled with this model. We provide genetic evidence that two RNA-sensing proteins, RIG-I and MDA5, participate in the IFN response to L. pneumophila. Importantly, these sensors do not seem to be required for the IFN response to L. pneumophila DNA, whereas we found that RIG-I was required for the response to L. pneumophila RNA. Thus, we hypothesize that bacterial RNA, or perhaps an induced host RNA, is the primary stimulus inducing the IFN response to L. pneumophila. Our study also identified a secreted effector protein, SdhA, as a key suppressor of the IFN response to L. pneumophila. Although viral suppressors of cytosolic RNA-sensing pathways have been previously identified, analogous bacterial factors have not been described. Thus, our results provide new insights into the molecular mechanisms by which an intracellular bacterial pathogen activates and also represses innate immune responses.

  10. A STUDY ON LEGIONELLA PNEUMOPHILA, WATER CHEMISTRY, AND ATMOSPHERIC CONDITIONS IN COOLING TOWERS AT THE SAVANNAH RIVER SITE

    Energy Technology Data Exchange (ETDEWEB)

    Smith, C.; Brigmon, R.

    2009-10-20

    Legionnaires disease is a pneumonia caused by the inhalation of the bacterium Legionella pneumophila. The majority of illnesses have been associated with cooling towers since these devices can harbor and disseminate the bacterium in the aerosolized mist generated by these systems. Historically, Savannah River Site (SRS) cooling towers have had occurrences of elevated levels of Legionella in all seasons of the year and in patterns that are difficult to predict. Since elevated Legionella in cooling tower water are a potential health concern a question has been raised as to the best control methodology. In this work we analyze available chemical, biological, and atmospheric data to determine the best method or key parameter for control. The SRS 4Q Industrial Hygiene Manual, 4Q-1203, 1 - G Cooling Tower Operation and the SRNL Legionella Sampling Program, states that 'Participation in the SRNL Legionella Sampling Program is MANDATORY for all operating cooling towers'. The resulting reports include L. pneumophila concentration information in cells/L. L. pneumophila concentrations >10{sup 7} cells/L are considered elevated and unsafe so action must be taken to reduce these densities. These remedial actions typically include increase biocide addition or 'shocking'. Sometimes additional actions are required if the problem persists including increase tower maintenance (e.g. cleaning). Evaluation of 14 SRS cooling towers, seven water quality parameters, and five Legionella serogroups over a three-plus year time frame demonstrated that cooling tower water Legionella densities varied widely though out this time period. In fact there was no one common consistent significant variable across all towers. The significant factors that did show up most frequently were related to suspended particulates, conductivity, pH, and dissolved oxygen, not chlorine or bromine as might be expected. Analyses of atmospheric data showed that there were more frequent significant

  11. Genome-scale identification of Legionella pneumophila effectors using a machine learning approach.

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    David Burstein

    2009-07-01

    Full Text Available A large number of highly pathogenic bacteria utilize secretion systems to translocate effector proteins into host cells. Using these effectors, the bacteria subvert host cell processes during infection. Legionella pneumophila translocates effectors via the Icm/Dot type-IV secretion system and to date, approximately 100 effectors have been identified by various experimental and computational techniques. Effector identification is a critical first step towards the understanding of the pathogenesis system in L. pneumophila as well as in other bacterial pathogens. Here, we formulate the task of effector identification as a classification problem: each L. pneumophila open reading frame (ORF was classified as either effector or not. We computationally defined a set of features that best distinguish effectors from non-effectors. These features cover a wide range of characteristics including taxonomical dispersion, regulatory data, genomic organization, similarity to eukaryotic proteomes and more. Machine learning algorithms utilizing these features were then applied to classify all the ORFs within the L. pneumophila genome. Using this approach we were able to predict and experimentally validate 40 new effectors, reaching a success rate of above 90%. Increasing the number of validated effectors to around 140, we were able to gain novel insights into their characteristics. Effectors were found to have low G+C content, supporting the hypothesis that a large number of effectors originate via horizontal gene transfer, probably from their protozoan host. In addition, effectors were found to cluster in specific genomic regions. Finally, we were able to provide a novel description of the C-terminal translocation signal required for effector translocation by the Icm/Dot secretion system. To conclude, we have discovered 40 novel L. pneumophila effectors, predicted over a hundred additional highly probable effectors, and shown the applicability of machine

  12. Interaction of legionella pneumophila and helicobacter pylori with bacterial species isolated from drinking water biofilms

    Directory of Open Access Journals (Sweden)

    Azevedo Nuno F

    2011-03-01

    Full Text Available Abstract Background It is well established that Legionella pneumophila is a waterborne pathogen; by contrast, the mode of Helicobacter pylori transmission remains unknown but water seems to play an important role. This work aims to study the influence of five microorganisms isolated from drinking water biofilms on the survival and integration of both of these pathogens into biofilms. Results Firstly, both pathogens were studied for auto- and co-aggregation with the species isolated from drinking water; subsequently the formation of mono and dual-species biofilms by L. pneumophila or H. pylori with the same microorganisms was investigated. Neither auto- nor co-aggregation was observed between the microorganisms tested. For biofilm studies, sessile cells were quantified in terms of total cells by SYTO 9 staining, viable L. pneumophila or H. pylori cells were quantified using 16 S rRNA-specific peptide nucleic acid (PNA probes and cultivable cells by standard culture techniques. Acidovorax sp. and Sphingomonas sp. appeared to have an antagonistic effect on L. pneumophila cultivability but not on the viability (as assessed by rRNA content using the PNA probe, possibly leading to the formation of viable but noncultivable (VBNC cells, whereas Mycobacterium chelonae increased the cultivability of this pathogen. The results obtained for H. pylori showed that M. chelonae and Sphingomonas sp. help this pathogen to maintain cultivability for at least 24 hours. Conclusions It appears that M. chelonae may have an important role in the survival of both pathogens in drinking water. This work also suggests that the presence of some microorganisms can decrease the cultivability of L. pneumophila but not the viability which indicates that the presence of autochthonous microorganisms can lead to misleading results when the safety of water is assessed by cultivable methods alone.

  13. Contamination of Tap Water with Pseudomonas aeruginosa, Legionella pneumophila, and Escherichia coli in Guilan, Iran

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    Masoumeh Ahmadi Jalali Moghadam

    2016-12-01

    Full Text Available Background: River and underground waters are main sources of tap water in Guilan, Iran. Overlandwastes move into rivers during periods of heavy or extended rain that is very common in the area.Pseudomonas aeruginosa, Legionella pneumophila, and Escherichia coli are main human pathogenswith water source. This study is designed to determine the load of these bacteria in main water suppliesof the area.Methods: Samples were collected directly into sterile containers, concentrated by centrifuge,inoculated in enrichment medium and incubated for 3-4 days. DNA was extracted by using commercialkit. Several rounds of PCR was performed to search P. aeroginosa, integron I, Metallo-β-lactamasesgene, L. pneumophila, mip gene, and E. coli.Results: About 92.0% of the samples showed bacterial contamination as revealed by PCR with primersof 16S rRNAgene, 9.5% of the samples had L. pneumophila, and 11,1% had Pseudomonas aeruginosa,but Escherichia coli was not detected. We found the mip gene in 66.6% of the samples with L.pneumophila. Metallo-β-lactamasesgene was found in 11.1% of all samples. We also found Integrin 1in 28.5% of the samples with P. aeruginosa.Conclusion: This study indicates that in spite of chlorination, total bacterial contamination of potwaters in the area is high and contamination with L. pneumophila and P. aeroginosa is considerable. Itmight be related to the biofilm formation and the growth of water microflora. It seems that free residualchlorine is ineffective. We suggest a more effective decontamination procedure based on moderntechnology.

  14. Free-living protozoa in drinking water supplies: community composition and role as hosts for Legionella pneumophila

    NARCIS (Netherlands)

    Valster, R.M.

    2011-01-01

    Free-living protozoa in drinking water supplies: community composition and role as hosts for Legionella pneumophila
    Free-living protozoa, which feed on bacteria, play an important role in the communities of microorganisms and invertebrates in drinking water supplies and in (warm) tap water

  15. Severe Legionella Pneumophila Infection in an Immunocompetent Patient: A Success Story 300 Kilometers Away.

    Science.gov (United States)

    Jacob, Miguel; Ramos, Helena C; Morgado, Bruno

    2016-12-21

    The most significant outbreak of Legionella pneumophila, or Legionnaires' Disease, ever registered in Portugal occurred in 2014, and was considered one of the largest in European history. This relatively rare infection has a dire prognosis if not timely identified and correctly treated, presenting with a high lethality rate. We describe a case of infection by Legionella pneumophila in a previously healthy individual during an outbreak that originated 300 kilometers away from our hospital. The patient presented to the Emergency Department and after an initial assessment, was admitted to the Intensive Care Unit (ICU). He underwent supportive treatment with invasive mechanical ventilation and antibiotic therapy, having been discharged with functional improvement 21 days after admission. During follow-up, the patient presented well without residual clinical or radiological findings. Prompt management following established guidelines allowed for the appropriate treatment and a favorable prognosis. This case serves as a reminder that early management is important, healthy individuals without known risk factors may present with severe infection, and there is the possibility for individual cases of Legionellosis to present far from the outbreak source.

  16. The metal efflux island of Legionella pneumophila is not required for survival in macrophages and amoebas.

    Science.gov (United States)

    Kim, Eun-Hae; Charpentier, Xavier; Torres-Urquidy, Oscar; McEvoy, Megan M; Rensing, Christopher

    2009-12-01

    Legionella pneumophila is an intracellular pathogen causing pneumonia-like disease in humans. A 43-kb putative heavy metal efflux gene island was found on the L. pneumophila genome. Large Legionella deletion strains of the metal efflux genes were tested in human THP-1-derived macrophages and amoebal Acanthamoeba castellanii cells and were able to survive and replicate similar to the wild type, suggesting that they do not play a significant role within the intracellular environment. Examination of the sequence of this genomic island revealed that some genes were not accurately annotated and there were no known metal-responsive regulators encoded in this region. Therefore, functional roles of these metal resistance genes were tested by conducting metal resistance assays. Individual genes were cloned in an expression vector and expressed in an appropriate metal-sensitive Escherichia coli background with varying concentrations of the tested metal. Of the 11 efflux systems, a role was determined only for one. A Cu(I)-translocating P(IB)-type ATPase was shown to be encoded by lpg1024. This gene, termed copA, complemented a copper-sensitive (Delta copA) E. coli strain in trans and was able to confer copper resistance.

  17. Molecular typing of Legionella pneumophila from air-conditioning cooling waters using mip gene, SBT, and FAFLP methods.

    Science.gov (United States)

    Gong, Xiangli; Li, Juntao; Zhang, Ying; Hou, Shuiping; Qu, Pinghua; Yang, Zhicong; Chen, Shouyi

    2017-08-01

    Legionella spp. are important waterborne pathogens. Molecular typing has become an important method for outbreaks investigations and source tracking of Legionnaires. In a survey program conducted by the Guangzhou Center for Disease Control and Prevention, multiple serotypes Legionella pneumophila (L. pneumophila) were isolated from waters in air-conditioning cooling towers in urban Guangzhou region, China between 2008 and 2011. Three genotyping methods, mip (macrophage infectivity potentiator) genotyping, SBT (sequence-based typing), and FAFLP (fluorescent amplified fragment length polymorphism analysis) were used to type these waterborne L. pneumophila isolates. The three methods were capable of typing all the 134 isolates and a reference strain of L. pneumophila (ATCC33153), with discriminatory indices of 0.7034, 0.9218, and 0.9376, for the mip, SBT, and FAFLP methods respectively. Among the 9 serotypes of the 134 isolates, 10, 50, and 34 molecular types were detected by the mip, SBT, and FAFLP methods respectively. The mip genotyping and SBT typing are more feasible for inter-laboratory results sharing and comparison of different types of L. pneumophila. The SBT and FAFLP typing methods were rapid with higher discriminatory abilities. Combinations of two or more of the typing methods enables more accurate typing of Legionella isolates for outbreak investigations and source tracking of Legionnaires. Copyright © 2017 Elsevier B.V. All rights reserved.

  18. [Investigation of Legionella pneumophila seropositivity in the professional long distance drivers as a risky occupation].

    Science.gov (United States)

    Polat, Yusuf; Ergin, Cağri; Kaleli, Ilknur; Pinar, Ahmet

    2007-04-01

    Contaminated water sources, reservoirs and systems such as evaporative condensers of air-conditioners are known to be the main transmission routes of Legionella spp. which are ubiquitous aquatic bacteria. By virtue of this point the aim of this study was to investigate the rate of Legionella pneumophila seropositivity in a profession considered as risky due to the direct and prolonged exposure to air-conditioning and air-circulating systems. For this purpose, in the period of February-August 2004 a total of 79 male subjects (63 were bus drivers and 16 were driver assistants) who were continously travelling to two different route (South part as hot climate and Middle/North parts as cold climate of Turkey) from Denizli province coach station (a province located in internal Aegian where accepted as crossroads), were included to the study. The mean age and mean working duration of bus drivers were 43.0 +/- 1.1 years and 20.0 +/- 1.1 years, respectively, while these values were 22.5 +/- 0.9 years and 4.0 +/- 0.6 years, respectively, for the drivers' assistants. The serum samples collected from the subjects were screened by a commercial indirect immunofluorescence method (Euroimmun, Germany) using L. pneumophila serogrup 1-14 antigens for the presence of specific antibodies. Additionally, air-conditioners' moisture exhaust samples of the busses in which seropositive subjects travelling with have been examined by culture and 5S rRNA gene targeted polymerase chain reaction (PCR) methods, for the presence of Legionella spp. The overall L. pneumophila seropositivity rate was detected as 15.2% (12/79). This rate was 19% (12/63) for bus drivers while all of the drivers' assistants were found seronegative. The seropositivity rate was found statistically higher in the personnel who were travelling to the hot climates (10/36, 27.8%) than those who travel to cold climates (2/43, 4.6%), (p distance bus drivers were chronically exposed to this pathogen and this may be considered as

  19. Virulence factor rtx in Legionella pneumophila, evidence suggesting it is a modular multifunctional protein

    Directory of Open Access Journals (Sweden)

    Pelaz Carmen

    2008-01-01

    Full Text Available Abstract Background The repeats in toxin (Rtx are an important pathogenicity factor involved in host cells invasion of Legionella pneumophila and other pathogenic bacteria. Its role in escaping the host immune system and cytotoxic activity is well known. Its repeated motives and modularity make Rtx a multifunctional factor in pathogenicity. Results The comparative analysis of rtx gene among 6 strains of L. pneumophila showed modularity in their structures. Among compared genomes, the N-terminal region of the protein presents highly dissimilar repeats with functionally similar domains. On the contrary, the C-terminal region is maintained with a fashionable modular configuration, which gives support to its proposed role in adhesion and pore formation. Despite the variability of rtx among the considered strains, the flanking genes are maintained in synteny and similarity. Conclusion In contrast to the extracellular bacteria Vibrio cholerae, in which the rtx gene is highly conserved and flanking genes have lost synteny and similarity, the gene region coding for the Rtx toxin in the intracellular pathogen L. pneumophila shows a rapid evolution. Changes in the rtx could play a role in pathogenicity. The interplay of the Rtx toxin with host membranes might lead to the evolution of new variants that are able to escape host cell defences.

  20. Post-translational modifications of host proteins by Legionella pneumophila: a sophisticated survival strategy.

    Science.gov (United States)

    Rolando, Monica; Buchrieser, Carmen

    2012-03-01

    Eukaryotic proteins are tightly regulated by post-translational modifications, leading to a very subtle degree of regulation in time and space. Pathogen-mediated post-translational modifications are key strategies to modulate host factors by targeting central signaling pathways in the host cell. Legionella pneumophila, an intracellular pathogen that coevolved with protozoan hosts, encodes a large arsenal of secreted effectors conferring the ability to evade host cellular defenses and to manipulate them to promote invasion and intracellular replication. Conservation of many signaling pathways of protozoa in human macrophages confers the ability of L. pneumophila to infect humans, causing a severe pneumonia called legionnaires' disease. Most of the secreted proteins are delivered by the Dot/Icm type IV secretion system and several of these have been shown to act on different cellular pathways critical for infection. Moreover, multiple effectors target a single host function to orchestrate bacterial survival. In this review, we focus on those effectors in the repertoire of L. pneumophila proteins that target key cellular pathways by specific post-translational modifications.

  1. A conserved OmpA-like protein in Legionella pneumophila required for efficient intracellular replication.

    Science.gov (United States)

    Goodwin, Ian P; Kumova, Ogan K; Ninio, Shira

    2016-08-01

    The OmpA-like protein domain has been associated with peptidoglycan-binding proteins, and is often found in virulence factors of bacterial pathogens. The intracellular pathogen Legionella pneumophila encodes for six proteins that contain the OmpA-like domain, among them the highly conserved uncharacterized protein we named CmpA. Here we set out to characterize the CmpA protein and determine its contribution to intracellular survival of L. pneumophila Secondary structure analysis suggests that CmpA is an inner membrane protein with a peptidoglycan-binding domain at the C-teminus. A cmpA mutant was able to replicate normally in broth, but failed to compete with an isogenic wild-type strain in an intracellular growth competition assay. The cmpA mutant also displayed significant intracellular growth defects in both the protozoan host Acanthamoeba castellanii and in primary bone marrow-derived macrophages, where uptake into the cells was also impaired. The cmpA phenotypes were completely restored upon expression of CmpA in trans The data presented here establish CmpA as a novel virulence factor of L. pneumophila that is required for efficient intracellular replication in both mammalian and protozoan hosts. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  2. Role of biofilm roughness and hydrodynamic conditions in Legionella pneumophila adhesion to and detachment from simulated drinking water biofilms.

    Science.gov (United States)

    Shen, Yun; Monroy, Guillermo L; Derlon, Nicolas; Janjaroen, Dao; Huang, Conghui; Morgenroth, Eberhard; Boppart, Stephen A; Ashbolt, Nicholas J; Liu, Wen-Tso; Nguyen, Thanh H

    2015-04-07

    Biofilms in drinking water distribution systems (DWDS) could exacerbate the persistence and associated risks of pathogenic Legionella pneumophila (L. pneumophila), thus raising human health concerns. However, mechanisms controlling adhesion and subsequent detachment of L. pneumophila associated with biofilms remain unclear. We determined the connection between L. pneumophila adhesion and subsequent detachment with biofilm physical structure characterization using optical coherence tomography (OCT) imaging technique. Analysis of the OCT images of multispecies biofilms grown under low nutrient condition up to 34 weeks revealed the lack of biofilm deformation even when these biofilms were exposed to flow velocity of 0.7 m/s, typical flow for DWDS. L. pneumophila adhesion on these biofilm under low flow velocity (0.007 m/s) positively correlated with biofilm roughness due to enlarged biofilm surface area and local flow conditions created by roughness asperities. The preadhered L. pneumophila on selected rough and smooth biofilms were found to detach when these biofilms were subjected to higher flow velocity. At the flow velocity of 0.1 and 0.3 m/s, the ratio of detached cell from the smooth biofilm surface was from 1.3 to 1.4 times higher than that from the rough biofilm surface, presumably because of the low shear stress zones near roughness asperities. This study determined that physical structure and local hydrodynamics control L. pneumophila adhesion to and detachment from simulated drinking water biofilm, thus it is the first step toward reducing the risk of L. pneumophila exposure and subsequent infections.

  3. Serotyping, ribotyping, PCR-mediated ribosomal 16S-23S spacer analysis and arbitrarily primed PCR for epidemiological studies on Legionella pneumophila

    NARCIS (Netherlands)

    A.F. van Belkum (Alex); H. Maas (Hugo); H.A. Verbrugh (Henri); N. van Leeuwen (N.)

    1996-01-01

    textabstractFifty clinical and environmental isolates of Legionella pneumophila were typed serologically and by DNA fingerprinting using arbitrarily primed polymerase chain reaction (AP-PCR). Furthermore, variability in and around ribosomal operons was assessed by conventional ribotyping and

  4. Convective Mixing in Distal Pipes Exacerbates Legionella pneumophila Growth in Hot Water Plumbing

    Directory of Open Access Journals (Sweden)

    William J. Rhoads

    2016-03-01

    Full Text Available Legionella pneumophila is known to proliferate in hot water plumbing systems, but little is known about the specific physicochemical factors that contribute to its regrowth. Here, L. pneumophila trends were examined in controlled, replicated pilot-scale hot water systems with continuous recirculation lines subject to two water heater settings (40 °C and 58 °C and three distal tap water use frequencies (high, medium, and low with two pipe configurations (oriented upward to promote convective mixing with the recirculating line and downward to prevent it. Water heater temperature setting determined where L. pneumophila regrowth occurred in each system, with an increase of up to 4.4 log gene copies/mL in the 40 °C system tank and recirculating line relative to influent water compared to only 2.5 log gene copies/mL regrowth in the 58 °C system. Distal pipes without convective mixing cooled to room temperature (23–24 °C during periods of no water use, but pipes with convective mixing equilibrated to 30.5 °C in the 40 °C system and 38.8 °C in the 58 °C system. Corresponding with known temperature effects on L. pneumophila growth and enhanced delivery of nutrients, distal pipes with convective mixing had on average 0.2 log more gene copies/mL in the 40 °C system and 0.8 log more gene copies/mL in the 58 °C system. Importantly, this work demonstrated the potential for thermal control strategies to be undermined by distal taps in general, and convective mixing in particular.

  5. First report of Legionella pneumophila in car cabin air filters. Are these a potential exposure pathway for professional drivers?

    Science.gov (United States)

    Alexandropoulou, Ioanna G; Konstantinidis, Theocharis G; Parasidis, Theodoros A; Nikolaidis, Christos; Panopoulou, Maria; Constantinidis, Theodoros C

    2013-12-01

    Recent findings have identified professional drivers as being at an increased risk of Legionnaires' disease. Our hypothesis was that used car cabin air filters represent a reservoir of Legionella bacteria, and thus a potential pathway for contamination. We analysed used cabin air filters from various types of car. The filters were analysed by culture and by molecular methods. Our findings indicated that almost a third of air filters were colonized with Legionella pneumophila. Here, we present the first finding of Legionella spp. in used car cabin air filters. Further investigations are needed in order to confirm this exposure pathway. The presence of Legionella bacteria in used cabin air filters may have been an unknown source of infection until now.

  6. SPR based immunosensor for detection of Legionella pneumophila in water samples

    Science.gov (United States)

    Enrico, De Lorenzis; Manera, Maria G.; Montagna, Giovanni; Cimaglia, Fabio; Chiesa, Maurizio; Poltronieri, Palmiro; Santino, Angelo; Rella, Roberto

    2013-05-01

    Detection of legionellae by water sampling is an important factor in epidemiological investigations of Legionnaires' disease and its prevention. To avoid labor-intensive problems with conventional methods, an alternative, highly sensitive and simple method is proposed for detecting L. pneumophila in aqueous samples. A compact Surface Plasmon Resonance (SPR) instrumentation prototype, provided with proper microfluidics tools, is built. The developed immunosensor is capable of dynamically following the binding between antigens and the corresponding antibody molecules immobilized on the SPR sensor surface. A proper immobilization strategy is used in this work that makes use of an important efficient step aimed at the orientation of antibodies onto the sensor surface. The feasibility of the integration of SPR-based biosensing setups with microfluidic technologies, resulting in a low-cost and portable biosensor is demonstrated.

  7. A PCR-Based Method for Monitoring Legionella pneumophila in Water Samples Detects Viable but Noncultivable Legionellae That Can Recover Their Cultivability▿

    Science.gov (United States)

    Dusserre, Eric; Ginevra, Christophe; Hallier-Soulier, Sylvie; Vandenesch, François; Festoc, Gabriel; Etienne, Jerome; Jarraud, Sophie; Molmeret, Maëlle

    2008-01-01

    Legionella pneumophila is the causative agent of Legionnaires' disease. This bacterium is ubiquitous in aqueous environments and uses amoebae as an intracellular replicative niche. Real-time PCR has been developed for rapid detection of Legionella DNA in water samples. In addition to culturable bacteria, this method may also detect dead and viable but noncultivable (VBNC) legionellae. In order to understand the significance of positive PCR results in this setting, we prepared water samples containing known concentrations of L. pneumophila and analyzed them comparatively by means of conventional culture, real-time PCR, viability labeling, and immunodetection (solid-phase cytometry). We also examined the influence of chlorination on the results of the four methods. The different techniques yielded similar results for nonchlorinated water samples but not for chlorinated samples. After treatment for 24 h with 0.5 and 1 ppm chlorine, all cultures were negative, PCR and immunodetection showed about 106 genome units and bacteria/ml, and total-viable-count (TVC) labeling detected 105 and 102 metabolically active bacteria/ml, respectively. Thus, PCR also detected bacteria that were VBNC. The recoverability of VBNC forms was confirmed by 5 days of coculture with Acanthamoeba polyphaga. Therefore, some TVC-positive bacteria were potentially infective. These data show that L. pneumophila PCR detects not only culturable bacteria but also VBNC forms and dead bacterial DNA at low chlorine concentrations. PMID:18515476

  8. Controlled evaluation of copper-silver ionization in eradicating Legionella pneumophila from a hospital water distribution system.

    Science.gov (United States)

    Liu, Z; Stout, J E; Tedesco, L; Boldin, M; Hwang, C; Diven, W F; Yu, V L

    1994-04-01

    A controlled evaluation was made of the efficacy of copper-silver ionization in eradicating Legionella pneumophila from a hospital water supply. Copper-silver ionization units were installed on the hot water recirculation line of one building with water fixtures positive for Legionella species. Another building with the same water supply served as a control. Legionella species persisted within the system when copper and silver concentrations were copper and silver concentrations were > 0.4 and > 0.04 ppm, respectively, there was a significant decrease in Legionella species colonization, but the percentage of water fixtures positive for organisms was unchanged in the control building. When the ionization unit was inactivated, water fixtures continued to be free of Legionella species for 2 additional months. Copper-silver ionization can eradicate L. pneumophila in a water distribution system. The advantages of copper-silver ionization include relatively low cost, straightforward installation, easy maintenance, nontoxic by-products and the presence of a disinfecting residual.

  9. Removal of bacteria Legionella pneumophila, Escherichia coli, and Bacillus subtilis by (super)cavitation.

    Science.gov (United States)

    Šarc, Andrej; Kosel, Janez; Stopar, David; Oder, Martina; Dular, Matevž

    2018-04-01

    In sufficient concentrations, the pathogenic bacteria L. pneumophila can cause a respiratory illness that is known as the "Legionnaires" disease. Moreover, toxic Shiga strains of bacteria E. coli can cause life-threatening hemolytic-uremic syndrome. Because of the recent restrictions imposed on the usage of chlorine, outbreaks of these two bacterial species have become more common. In this study we have developed a novel rotation generator and its effectiveness against bacteria Legionella pneumophila and Escherichia coli was tested for various types of hydrodynamic cavitation (attached steady cavitation, developed unsteady cavitation and supercavitation). The results show that the supercavitation was the only effective form of cavitation. It enabled more than 3 logs reductions for both bacterial species and was also effective against a more persistent Gram positive bacteria, B. subtilis. The deactivation mechanism is at present unknown. It is proposed that when bacterial cells enter a supercavitation cavity, an immediate pressure drop occurs and this results in bursting of the cellular membrane. The new rotation generator that induced supercavitation proved to be economically and microbiologically far more effective than the classical Venturi section (super)cavitation. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. Essential roles and regulation of the Legionella pneumophila collagen-like adhesin during biofilm formation.

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    Julia Mallegol

    Full Text Available Legionellosis is mostly caused by Legionella pneumophila (Lp and is defined by a severe respiratory illness with a case fatality rate ranging from 5 to 80%. In a previous study, we showed that a glycosaminoglycan (GAG-binding adhesin of Lp, named Lcl, is produced during legionellosis and is unique to the L. pneumophila species. Importantly, a mutant depleted in Lcl (Δlpg2644 is impaired in adhesion to GAGs and epithelial cells and in biofilm formation. Here, we examine the molecular function(s of Lcl and the transcriptional regulation of its encoding gene during different stages of the biofilm development. We show that the collagen repeats and the C-terminal domains of Lcl are crucial for the production of biofilm. We present evidence that Lcl is involved in the early step of surface attachment but also in intercellular interactions. Furthermore, we address the relationship between Lcl gene regulation during biofilm formation and quorum sensing (QS. In a static biofilm assay, we show that Lcl is differentially regulated during growth phases and biofilm formation. Moreover, we show that the transcriptional regulation of lpg2644, mediated by a prototype of QS signaling homoserine lactone (3OC12-HSL, may play a role during the biofilm development. Thus, transcriptional down-regulation of lpg2644 may facilitate the dispersion of Lp to reinitiate biofilm colonization on a distal surface.

  11. Comprehensive identification of protein substrates of the Dot/Icm type IV transporter of Legionella pneumophila.

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    Wenhan Zhu

    2011-03-01

    Full Text Available A large number of proteins transferred by the Legionella pneumophila Dot/Icm system have been identified by various strategies. With no exceptions, these strategies are based on one or more characteristics associated with the tested proteins. Given the high level of diversity exhibited by the identified proteins, it is possible that some substrates have been missed in these screenings. In this study, we took a systematic method to survey the L. pneumophila genome by testing hypothetical orfs larger than 300 base pairs for Dot/Icm-dependent translocation. 798 of the 832 analyzed orfs were successfully fused to the carboxyl end of β-lactamase. The transfer of the fusions into mammalian cells was determined using the β-lactamase reporter substrate CCF4-AM. These efforts led to the identification of 164 proteins positive in translocation. Among these, 70 proteins are novel substrates of the Dot/Icm system. These results brought the total number of experimentally confirmed Dot/Icm substrates to 275. Sequence analysis of the C-termini of these identified proteins revealed that Lpg2844, which contains few features known to be important for Dot/Icm-dependent protein transfer can be translocated at a high efficiency. Thus, our efforts have identified a large number of novel substrates of the Dot/Icm system and have revealed the diverse features recognizable by this protein transporter.

  12. Inhibition of host cell translation elongation by Legionella pneumophila blocks the host cell unfolded protein response.

    Science.gov (United States)

    Hempstead, Andrew D; Isberg, Ralph R

    2015-12-08

    Cells of the innate immune system recognize bacterial pathogens by detecting common microbial patterns as well as pathogen-specific activities. One system that responds to these stimuli is the IRE1 branch of the unfolded protein response (UPR), a sensor of endoplasmic reticulum (ER) stress. Activation of IRE1, in the context of Toll-like receptor (TLR) signaling, induces strong proinflammatory cytokine induction. We show here that Legionella pneumophila, an intravacuolar pathogen that replicates in an ER-associated compartment, blocks activation of the IRE1 pathway despite presenting pathogen products that stimulate this response. L. pneumophila TLR ligands induced the splicing of mRNA encoding XBP1s, the main target of IRE1 activity. L. pneumophila was able to inhibit both chemical and bacterial induction of XBP1 splicing via bacterial translocated proteins that interfere with host protein translation. A strain lacking five translocated translation elongation inhibitors was unable to block XBP1 splicing, but this could be rescued by expression of a single such inhibitor, consistent with limitation of the response by translation elongation inhibitors. Chemical inhibition of translation elongation blocked pattern recognition receptor-mediated XBP1 splicing, mimicking the effects of the bacterial translation inhibitors. In contrast, host cell-promoted inhibition of translation initiation in response to the pathogen was ineffective in blocking XBP1 splicing, demonstrating the need for the elongation inhibitors for protection from the UPR. The inhibition of host translation elongation may be a common strategy used by pathogens to limit the innate immune response by interfering with signaling via the UPR.

  13. Reassessing the role of DotF in the Legionella pneumophila type IV secretion system.

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    Molly C Sutherland

    Full Text Available Legionella pneumophila, the causative agent of a severe pneumonia termed Legionnaires' Disease, survives and replicates within both protozoan hosts and human alveolar macrophages. Intracellular survival is dependent upon secretion of a plethora of protein effectors that function to form a replicative vacuole, evade the endocytic pathway and subvert host immune defenses. Export of these factors requires a type IV secretion system (T4SS called Dot/Icm that is composed of twenty-seven proteins. This report focuses on the DotF protein, which was previously postulated to have several different functions, one of which centered on binding Dot/Icm substrates. In this report, we examined if DotF functions as the T4SS inner membrane receptor for Dot/Icm substrates. Although we were able to recapitulate the previously published bacterial two-hybrid interaction between DotF and several substrates, the interaction was not dependent on the Dot/Icm substrates' signal sequences as predicted for a substrate:receptor interaction. In addition, binding did not require the cytoplasmic domain of DotF, which was anticipated to be involved in recognizing substrates in the cytoplasm. Finally, inactivation of dotF did not abolish intracellular growth of L. pneumophila or translocation of substrates, two phenotypes dependent on the T4SS receptor. These data strongly suggest that DotF does not act as the major receptor for Dot/Icm substrates and therefore likely performs an accessory function within the core-transmembrane subcomplex of the L. pneumophila Dot/Icm type IV secretion system.

  14. Induction of caspase 3 activation by multiple Legionella pneumophila Dot/Icm substrates.

    Science.gov (United States)

    Zhu, Wenhan; Hammad, Loubna A; Hsu, Fosheng; Mao, Yuxin; Luo, Zhao-Qing

    2013-11-01

    The intracellular pathogen Legionella pneumophila is able to strike a balance between the death and survival of the host cell during infection. Despite the presence of high level of active caspase 3, the executioner caspase of apoptotic cell death, infected permissive macrophages are markedly resistant to exogenous apoptotic stimuli. Several bacterial molecules capable of promoting the cell survival pathways have been identified, but proteins involved in the activation of caspase 3 remain unknown. To study the mechanism of L. pneumophila-mediated caspase 3 activation, we tested all known Dot/Icm substrates for their ability to activate caspase 3. Five effectors capable of causing caspase 3 activation upon transient expression were identified. Among these, by using its ability to activate caspase 3 by inducing the release of cytochrome c from the mitochondria, we demonstrated that VipD is a phospholipase A2, which hydrolyses phosphatidylethanolamine (PE) and phosphocholine (PC) on the mitochondrial membrane in a manner that appears to require host cofactor(s). The lipase activity leads to the production of free fatty acids and 2-lysophospholipids, which destabilize the mitochondrial membrane and may contribute to the release of cytochrome c and the subsequent caspase 3 activation. Furthermore, we found that whereas it is not detectably defectively in caspase 3 activation in permissive cells, amutant lacking all of these five genes is less potent in inducing apoptosis in dendritic cells. Our results reveal that activation of host cell death pathways by L. pneumophila is a result of the effects of multiple bacterial proteins with diverse biochemical functions. © 2013 John Wiley & Sons Ltd.

  15. The membrane protein LasM Promotes the Culturability of Legionella pneumophila in Water

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    Laam Li

    2016-09-01

    Full Text Available The water-borne pathogen Legionella pneumophila (Lp strongly expresses the lpg1659 gene in water. This gene encodes a hypothetical protein predicted to be a membrane protein using in silico analysis. While no conserved domains were identified in Lpg1659, similar proteins are found in many Legionella species and other aquatic bacteria. RT-qPCR showed that lpg1659 is positively regulated by the alternative sigma factor RpoS, which is essential for Lp to survive in water. These observations suggest an important role of this novel protein in the survival of Lp in water. Deletion of lpg1659 did not affect cell morphology, membrane integrity or tolerance to high temperature. Moreover, lpg1659 was dispensable for growth of Lp in rich medium, and during infection of the amoeba Acanthamoeba castellanii and of THP-1 human macrophages. However, deletion of lpg1659 resulted in an early loss of culturability in water, while over-expression of this gene promoted the culturability of Lp. Therefore, these results suggest that lpg1659 is required for Lp to maintain culturability, and possibly long-term survival, in water. Since the loss of culturability observed in the absence of Lpg1659 was complemented by the addition of trace metals into water, this membrane protein is likely a transporter for acquiring essential trace metal for maintaining culturability in water and potentially in other metal-deprived conditions. Given its role in the survival of Lp in water, Lpg1659 was named LasM for Legionella aquatic survival membrane protein.

  16. Treatment of a Legionella pneumophila-colonized water distribution system using copper-silver ionization and continuous chlorination.

    Science.gov (United States)

    Biurrun, A; Caballero, L; Pelaz, C; León, E; Gago, A

    1999-06-01

    The detection in April 1997 of a case of nosocomial legionellosis in our hospital led to the discovery that both our hot- and cold-water circuits were heavily colonized with Legionella pneumophila. Conventional methods for eradication of the organisms were unsuccessful, so a copper-silver (Cu-Ag) ionization system and a continuous chlorination system were installed. Five months later, the number of colonized sites decreased from an initial 58.3% to 16.7%.

  17. [Rate and level of contamination of potentially dangerous water supply facilities in Moscow region by Legionella pneumophila].

    Science.gov (United States)

    Tartakovskiĭ, I S; Novokshonova, I V; Sadretdinova, O V; Gruzdeva, O A; Aliapkina, Iu S; Karpova, T I; Dronina, Iu E; Fokina, V G; Shustrova, N M

    2010-01-01

    To assess rate and level of contamination by Legionella pneumophila of cooling water systems in industrial facilities as well as hot water supply systems of administrative buildings in Moscow region. Cooling water systems of 8 industrial facilities and hot water supply systems of 12 administrative buildings or complexes located in Moscow or Moscow region were examined. Samples of water, washes and biofilms were studied by bacteriologic methods and RT-PCR. Results. Significant level of contamination of water systems by L. pneumophila was revealed in examined objects. Rate of contamination of cooling water systems in industrial facilities was 70%. The agent was detected in stagnant, end-capped, and rarely used segments of all hot water supply systems during decrease of water temperature to 36-52 degrees C. Visual detection of natural biofilms on the object correlated with high concentration of L. pneumophila in water samples. In some cases, associations of L. pneumophila with Pseudomonas aeruginosa were detected, including water samples from supply systems of 2 healthcare facilities. Obtained results confirm the importance of implementation of modem concept of legionellosis prevention in our country, based on regular quantitative monitoring for Legionella in potentially dangerous water objects and conduction of preventative measures if contamination exceeds acceptable level.

  18. Influence of pH on bioactivity of cinnamon oil against Legionella pneumophila and its disinfection efficacy in hot springs.

    Science.gov (United States)

    Chang, Ching-Wen; Chang, Wei-Lung; Chang, Shang-Tzen

    2008-12-01

    Cinnamon oil extracted from leaves of Cinnamomum osmophloeum has recently been proved as a promising antibacterial agent against Legionella pneumophila, an etiological agent of human pneumonia known as Legionnaires' disease. However, the pH effects on the efficacy of cinnamon oil against L. pneumophila and its applicability to recreational spring water remain unknown. We therefore determined the bactericidal activity of cinnamon oil at pH 3-10 in phosphate-buffered saline (PBS) and in four kinds of springs with various conductivity (259-5595 micros cm(-1)) and pH (2.1-7.7) levels. Results show L. pneumophila cells were more susceptible to cinnamon oil at pH 8-10 than at pH 4-6 in PBS, which became more evident as increasing contact time from 10 to 60 min. An increase in concentration of cinnamon oil and contact time significantly increased the anti-L. pneumophila activity (Pcinnamon oil at 300-750 microg ml(-1), with the highest inactivation in alkaline hydrogen carbonate spring. The great bioactivity of cinnamon oil demonstrates its potential to be used to control Legionella growth in recreational spring water and possibly other niches generally at basic pH, e.g., cooling towers.

  19. Legionella pneumophila serogroup 3 pneumonia in a patient with low-grade 4 non-Hodgkin lymphoma: a case report

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    Bistoni Francesco

    2011-08-01

    Full Text Available Abstract Introduction Nosocomial legionellosis has generally been described in immunodepressed patients, but Legionella pneumophila serogroup 3 has rarely been identified as the causative agent. Case presentation We report the case of nosocomial L. pneumophila serogroup 3 pneumonia in a 70-year-old Caucasian man with non-Hodgkin lymphoma. Diagnosis was carried out by culture and real-time polymerase chain reaction of bronchoalveolar lavage fluid. The results of a urinary antigen test were negative. A hospital environmental investigation revealed that the hospital water system was highly colonized by L. pneumophila serogroups 3, 4, and 8. The hospital team involved in the prevention of infections was informed, long-term control measures to reduce the environmental bacterial load were adopted, and clinical monitoring of legionellosis occurrence in high-risk patients was performed. No further cases of Legionella pneumonia have been observed so far. Conclusions In this report, we describe a case of legionellosis caused by L. pneumophila serogroup 3, which is not usually a causative agent of nosocomial infection. Our research confirms the importance of carrying out cultures of respiratory secretions to diagnose legionellosis and highlights the limited value of the urinary antigen test for hospital infections, especially in immunocompromised patients. It also indicates that, to reduce the bacterial load and prevent nosocomial legionellosis, appropriate control measures should be implemented with systematic monitoring of hospital water systems.

  20. Legionella pneumophila serogroup 3 pneumonia in a patient with low-grade 4 non-Hodgkin lymphoma: a case report.

    Science.gov (United States)

    Mencacci, Antonella; Corbucci, Cristina; Castellani, Alessio; Furno, Paolo; Bistoni, Francesco; Vecchiarelli, Anna

    2011-08-17

    Nosocomial legionellosis has generally been described in immunodepressed patients, but Legionella pneumophila serogroup 3 has rarely been identified as the causative agent. We report the case of nosocomial L. pneumophila serogroup 3 pneumonia in a 70-year-old Caucasian man with non-Hodgkin lymphoma. Diagnosis was carried out by culture and real-time polymerase chain reaction of bronchoalveolar lavage fluid. The results of a urinary antigen test were negative. A hospital environmental investigation revealed that the hospital water system was highly colonized by L. pneumophila serogroups 3, 4, and 8. The hospital team involved in the prevention of infections was informed, long-term control measures to reduce the environmental bacterial load were adopted, and clinical monitoring of legionellosis occurrence in high-risk patients was performed. No further cases of Legionella pneumonia have been observed so far. In this report, we describe a case of legionellosis caused by L. pneumophila serogroup 3, which is not usually a causative agent of nosocomial infection. Our research confirms the importance of carrying out cultures of respiratory secretions to diagnose legionellosis and highlights the limited value of the urinary antigen test for hospital infections, especially in immunocompromised patients. It also indicates that, to reduce the bacterial load and prevent nosocomial legionellosis, appropriate control measures should be implemented with systematic monitoring of hospital water systems.

  1. A Legionella pneumophila effector protein encoded in a region of genomic plasticity binds to Dot/Icm-modified vacuoles.

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    Shira Ninio

    2009-01-01

    Full Text Available Legionella pneumophila is an opportunistic pathogen that can cause a severe pneumonia called Legionnaires' disease. In the environment, L. pneumophila is found in fresh water reservoirs in a large spectrum of environmental conditions, where the bacteria are able to replicate within a variety of protozoan hosts. To survive within eukaryotic cells, L. pneumophila require a type IV secretion system, designated Dot/Icm, that delivers bacterial effector proteins into the host cell cytoplasm. In recent years, a number of Dot/Icm substrate proteins have been identified; however, the function of most of these proteins remains unknown, and it is unclear why the bacterium maintains such a large repertoire of effectors to promote its survival. Here we investigate a region of the L. pneumophila chromosome that displays a high degree of plasticity among four sequenced L. pneumophila strains. Analysis of GC content suggests that several genes encoded in this region were acquired through horizontal gene transfer. Protein translocation studies establish that this region of genomic plasticity encodes for multiple Dot/Icm effectors. Ectopic expression studies in mammalian cells indicate that one of these substrates, a protein called PieA, has unique effector activities. PieA is an effector that can alter lysosome morphology and associates specifically with vacuoles that support L. pneumophila replication. It was determined that the association of PieA with vacuoles containing L. pneumophila requires modifications to the vacuole mediated by other Dot/Icm effectors. Thus, the localization properties of PieA reveal that the Dot/Icm system has the ability to spatially and temporally control the association of an effector with vacuoles containing L. pneumophila through activities mediated by other effector proteins.

  2. Legionella pneumophila: the paradox of a highly sensitive opportunistic waterborne pathogen able to persist in the environment

    Directory of Open Access Journals (Sweden)

    Jean-Marc eBerjeaud

    2016-04-01

    Full Text Available Legionella pneumophila, the major causative agent of Legionnaires’ disease, is found in freshwater environments in close association with free-living amoebae and multispecies biofilms, leading to persistence, spread, biocide resistance, and elevated virulence of the bacterium. Indeed, legionellosis outbreaks are mainly due to the ability of this bacterium to colonize and persist in water facilities, despite harsh physical and chemical treatments. However, these treatments are not totally efficient and, after a lag period, L. pneumophila may be able to quickly re-colonize these systems. Several natural compounds (biosurfactants, antimicrobial peptides… with anti-Legionella properties have recently been described in the literature, highlighting their specific activities against this pathogen. In this review, we first consider this hallmark of Legionella to resist killing, in regard to its biofilm or host-associated life style. Then, we focus more accurately on natural anti-Legionella molecules described so far, which could provide new eco-friendly and alternative ways to struggle against this important pathogen in plumbing.

  3. Interactive effects of temperature, organic carbon, and pipe material on microbiota composition and Legionella pneumophila in hot water plumbing systems.

    Science.gov (United States)

    Proctor, Caitlin R; Dai, Dongjuan; Edwards, Marc A; Pruden, Amy

    2017-10-04

    Several biotic and abiotic factors have been reported to influence the proliferation of microbes, including Legionella pneumophila, in hot water premise plumbing systems, but their combined effects have not been systematically evaluated. Here, we utilize simulated household water heaters to examine the effects of stepwise increases in temperature (32-53 °C), pipe material (copper vs. cross-linked polyethylene (PEX)), and influent assimilable organic carbon (0-700 μg/L) on opportunistic pathogen gene copy numbers and the microbiota composition, as determined by quantitative polymerase chain reaction and 16S rRNA gene amplicon sequencing. Temperature had an overarching influence on both the microbiota composition and L. pneumophila numbers. L. pneumophila peaked at 41 °C in the presence of PEX (1.58 × 10 5 gene copies/mL). At 53 °C, L. pneumophila was not detected. Several operational taxonomic units (OTUs) persisted across all conditions, accounting for 50% of the microbiota composition from 32 to 49 °C and 20% at 53 °C. Pipe material most strongly influenced microbiota composition at lower temperatures, driven by five to six OTUs enriched with each material. Copper pipes supported less L. pneumophila than PEX pipes (mean 2.5 log 10 lower) at temperatures ≤ 41 °C, but showed no difference in total bacterial numbers. Differences between pipe materials diminished with elevated temperature, probably resulting from decreased release of copper ions. At temperatures ≤ 45 °C, influent assimilable organic carbon correlated well with total bacterial numbers, but not with L. pneumophila numbers. At 53 °C, PEX pipes leached organic carbon, reducing the importance of dosed organic carbon. L. pneumophila numbers correlated with a Legionella OTU and a Methylophilus OTU identified by amplicon sequencing. Temperature was the most effective factor for the control of L. pneumophila, while microbiota composition shifted with each stepwise temperature

  4. Water heater temperature set point and water use patterns influence Legionella pneumophila and associated microorganisms at the tap.

    Science.gov (United States)

    Rhoads, William J; Ji, Pan; Pruden, Amy; Edwards, Marc A

    2015-12-01

    Lowering water heater temperature set points and using less drinking water are common approaches to conserving water and energy; yet, there are discrepancies in past literature regarding the effects of water heater temperature and water use patterns on the occurrence of opportunistic pathogens, in particular Legionella pneumophila. Our objective was to conduct a controlled, replicated pilot-scale investigation to address this knowledge gap using continuously recirculating water heaters to examine five water heater set points (39-58 °C) under three water use conditions. We hypothesized that L. pneumophila levels at the tap depend on the collective influence of water heater temperature, flow frequency, and the resident plumbing ecology. We confirmed temperature setting to be a critical factor in suppressing L. pneumophila growth both in continuously recirculating hot water lines and at distal taps. For example, at 51 °C, planktonic L. pneumophila in recirculating lines was reduced by a factor of 28.7 compared to 39 °C and was prevented from re-colonizing biofilm. However, L. pneumophila still persisted up to 58 °C, with evidence that it was growing under the conditions of this study. Further, exposure to 51 °C water in a low-use tap appeared to optimally select for L. pneumophila (e.g., 125 times greater numbers than in high-use taps). We subsequently explored relationships among L. pneumophila and other ecologically relevant microbes, noting that elevated temperature did not have a general disinfecting effect in terms of total bacterial numbers. We documented the relationship between L. pneumophila and Legionella spp., and noted several instances of correlations with Vermamoeba vermiformis, and generally found that there is a dynamic relationship with this amoeba host over the range of temperatures and water use frequencies examined. Our study provides a new window of understanding into the microbial ecology of potable hot water systems and helps to resolve

  5. Neutrophil and Alveolar Macrophage-Mediated Innate Immune Control of Legionella pneumophila Lung Infection via TNF and ROS.

    Directory of Open Access Journals (Sweden)

    Pascal Ziltener

    2016-04-01

    Full Text Available Legionella pneumophila is a facultative intracellular bacterium that lives in aquatic environments where it parasitizes amoeba. However, upon inhalation of contaminated aerosols it can infect and replicate in human alveolar macrophages, which can result in Legionnaires' disease, a severe form of pneumonia. Upon experimental airway infection of mice, L. pneumophila is rapidly controlled by innate immune mechanisms. Here we identified, on a cell-type specific level, the key innate effector functions responsible for rapid control of infection. In addition to the well-characterized NLRC4-NAIP5 flagellin recognition pathway, tumor necrosis factor (TNF and reactive oxygen species (ROS are also essential for effective innate immune control of L. pneumophila. While ROS are essential for the bactericidal activity of neutrophils, alveolar macrophages (AM rely on neutrophil and monocyte-derived TNF signaling via TNFR1 to restrict bacterial replication. This TNF-mediated antibacterial mechanism depends on the acidification of lysosomes and their fusion with L. pneumophila containing vacuoles (LCVs, as well as caspases with a minor contribution from cysteine-type cathepsins or calpains, and is independent of NLRC4, caspase-1, caspase-11 and NOX2. This study highlights the differential utilization of innate effector pathways to curtail intracellular bacterial replication in specific host cells upon L. pneumophila airway infection.

  6. Neutrophil and Alveolar Macrophage-Mediated Innate Immune Control of Legionella pneumophila Lung Infection via TNF and ROS

    Science.gov (United States)

    Ziltener, Pascal; Reinheckel, Thomas; Oxenius, Annette

    2016-01-01

    Legionella pneumophila is a facultative intracellular bacterium that lives in aquatic environments where it parasitizes amoeba. However, upon inhalation of contaminated aerosols it can infect and replicate in human alveolar macrophages, which can result in Legionnaires’ disease, a severe form of pneumonia. Upon experimental airway infection of mice, L. pneumophila is rapidly controlled by innate immune mechanisms. Here we identified, on a cell-type specific level, the key innate effector functions responsible for rapid control of infection. In addition to the well-characterized NLRC4-NAIP5 flagellin recognition pathway, tumor necrosis factor (TNF) and reactive oxygen species (ROS) are also essential for effective innate immune control of L. pneumophila. While ROS are essential for the bactericidal activity of neutrophils, alveolar macrophages (AM) rely on neutrophil and monocyte-derived TNF signaling via TNFR1 to restrict bacterial replication. This TNF-mediated antibacterial mechanism depends on the acidification of lysosomes and their fusion with L. pneumophila containing vacuoles (LCVs), as well as caspases with a minor contribution from cysteine-type cathepsins or calpains, and is independent of NLRC4, caspase-1, caspase-11 and NOX2. This study highlights the differential utilization of innate effector pathways to curtail intracellular bacterial replication in specific host cells upon L. pneumophila airway infection. PMID:27105352

  7. Detection of Legionella pneumophila antigen in urine samples by the BinaxNOW immunochromatographic assay and comparison with both Binax Legionella Urinary Enzyme Immunoassay (EIA) and Biotest Legionella Urin Antigen EIA.

    Science.gov (United States)

    Helbig, J H; Uldum, S A; Lück, P C; Harrison, T G

    2001-06-01

    The new BinaxNOW Immunochromatographic (ICT) Assay for the detection of Legionella pneumophila antigens was used to test 535 urine specimens from patients with and without Legionnaires' disease. The specificity, calculated by testing 112 samples from patients with pneumonia of aetiologies other than Legionella infection, and 167 urine specimens from urinary tract infections, was found to be 97.1% if the manufacturer's guidelines were followed. However, it was determined that the 'false positive' results characterised by very weak bands could be discounted by re-examination of the results at 60 min, yielding a specificity of 100%. With this minor modification of the procedure applied to examination of urine samples from 117 patients with legionellosis confirmed by isolation of L. pneumophila and 70 patients who had seroconverted to L. pneumophila serogroup 1, sensitivity was calculated to be 79.7%. In comparison, the sensitivities of the Binax Urinary Antigen Enzyme Immunoassay (EIA) and Biotest Urin Antigen EIA were estimated to be 79.1 and 83.4%, respectively. Eleven cases (5.9%) were positive by BinaxNOW assay but negative by Binax or Biotest EIA, or both. The sensitivities of all assays increased to c. 94% if only diagnosis of cases confirmed by isolation of serogroup 1 L. pneumophila was considered, although the sensitivity for infections caused by L. pneumophila serogroup 1 monoclonal antibody (MAb) subgroup Bellingham was significantly lower than for other MAb subgroups. The Biotest EIA recognised 10 (45%) of the 22 cases not caused by L. pneumophila serogroup 1, whereas the two Binax kits detected only three each. The ICT assay BinaxNOW can be recommended as a rapid specific test for the diagnosis of Legionnaires' diseases caused by L. pneumophila serogroup 1, although very weak bands should be interpreted cautiously.

  8. Sequencing illustrates the transcriptional response of Legionella pneumophila during infection and identifies seventy novel small non-coding RNAs.

    LENUS (Irish Health Repository)

    Weissenmayer, Barbara A

    2011-01-01

    Second generation sequencing has prompted a number of groups to re-interrogate the transcriptomes of several bacterial and archaeal species. One of the central findings has been the identification of complex networks of small non-coding RNAs that play central roles in transcriptional regulation in all growth conditions and for the pathogen\\'s interaction with and survival within host cells. Legionella pneumophila is a gram-negative facultative intracellular human pathogen with a distinct biphasic lifestyle. One of its primary environmental hosts in the free-living amoeba Acanthamoeba castellanii and its infection by L. pneumophila mimics that seen in human macrophages. Here we present analysis of strand specific sequencing of the transcriptional response of L. pneumophila during exponential and post-exponential broth growth and during the replicative and transmissive phase of infection inside A. castellanii. We extend previous microarray based studies as well as uncovering evidence of a complex regulatory architecture underpinned by numerous non-coding RNAs. Over seventy new non-coding RNAs could be identified; many of them appear to be strain specific and in configurations not previously reported. We discover a family of non-coding RNAs preferentially expressed during infection conditions and identify a second copy of 6S RNA in L. pneumophila. We show that the newly discovered putative 6S RNA as well as a number of other non-coding RNAs show evidence for antisense transcription. The nature and extent of the non-coding RNAs and their expression patterns suggests that these may well play central roles in the regulation of Legionella spp. specific traits and offer clues as to how L. pneumophila adapts to its intracellular niche. The expression profiles outlined in the study have been deposited into Genbank\\'s Gene Expression Omnibus (GEO) database under the series accession GSE27232.

  9. Structural Basis for Rab1 De-AMPylation by the Legionella pneumophila Effector SidD

    Science.gov (United States)

    Neunuebel, M. Ramona; Pallara, Chiara; Brady, Jacqueline; Kinch, Lisa N.; Fernández-Recio, Juan; Rojas, Adriana L.; Machner, Matthias P.; Hierro, Aitor

    2013-01-01

    The covalent attachment of adenosine monophosphate (AMP) to proteins, a process called AMPylation (adenylylation), has recently emerged as a novel theme in microbial pathogenesis. Although several AMPylating enzymes have been characterized, the only known virulence protein with de-AMPylation activity is SidD from the human pathogen Legionella pneumophila. SidD de-AMPylates mammalian Rab1, a small GTPase involved in secretory vesicle transport, thereby targeting the host protein for inactivation. The molecular mechanisms underlying Rab1 recognition and de-AMPylation by SidD are unclear. Here, we report the crystal structure of the catalytic region of SidD at 1.6 Å resolution. The structure reveals a phosphatase-like fold with additional structural elements not present in generic PP2C-type phosphatases. The catalytic pocket contains a binuclear metal-binding site characteristic of hydrolytic metalloenzymes, with strong dependency on magnesium ions. Subsequent docking and molecular dynamics simulations between SidD and Rab1 revealed the interface contacts and the energetic contribution of key residues to the interaction. In conjunction with an extensive structure-based mutational analysis, we provide in vivo and in vitro evidence for a remarkable adaptation of SidD to its host cell target Rab1 which explains how this effector confers specificity to the reaction it catalyses. PMID:23696742

  10. Structural basis for Rab1 de-AMPylation by the Legionella pneumophila effector SidD.

    Directory of Open Access Journals (Sweden)

    Yang Chen

    Full Text Available The covalent attachment of adenosine monophosphate (AMP to proteins, a process called AMPylation (adenylylation, has recently emerged as a novel theme in microbial pathogenesis. Although several AMPylating enzymes have been characterized, the only known virulence protein with de-AMPylation activity is SidD from the human pathogen Legionella pneumophila. SidD de-AMPylates mammalian Rab1, a small GTPase involved in secretory vesicle transport, thereby targeting the host protein for inactivation. The molecular mechanisms underlying Rab1 recognition and de-AMPylation by SidD are unclear. Here, we report the crystal structure of the catalytic region of SidD at 1.6 Å resolution. The structure reveals a phosphatase-like fold with additional structural elements not present in generic PP2C-type phosphatases. The catalytic pocket contains a binuclear metal-binding site characteristic of hydrolytic metalloenzymes, with strong dependency on magnesium ions. Subsequent docking and molecular dynamics simulations between SidD and Rab1 revealed the interface contacts and the energetic contribution of key residues to the interaction. In conjunction with an extensive structure-based mutational analysis, we provide in vivo and in vitro evidence for a remarkable adaptation of SidD to its host cell target Rab1 which explains how this effector confers specificity to the reaction it catalyses.

  11. Applications of Legionella pneumophila protein FLA and PILE in serological diagnosis.

    Science.gov (United States)

    Liu, Huiyu; Liu, Yang; Yang, Zhiwei; Cao, Xiuqin

    2014-01-01

    Serological testing for antibody against Legionella pneumophila (LP) is often the primary method of screening for possible Lp infections. This study is an attempt to use the Lp protein FLA and PILE as coating antigen for ELISA. First, the coding sequences of the two proteins were cloned into the pET32a(+) vectors. Then the two proteins were produced and purified. The best working concentration of the two proteins as coating antigen was screened separately by criss-cross serial-dilution analysis. Then an ELISA method was developed for Lp antibody detection. The Lp antibody level of 120 serums was detected by both this new ELISA method and the DRG ELISA kit. The results suggest that protein FLA and PILE as coating antigen of ELISA to detect Lp antibody in serum shows better specificity and positive rate (5%, 3.4%) than the DRG ELISA kit. The new ELISA has a high referential value for Legionnaires' disease (LD) diagnosis and provided valuable information for the development of a serodiagnosis Kit for LD.

  12. Identification of Conserved ABC Importers Necessary for Intracellular Survival of Legionella pneumophila in Multiple Hosts

    Directory of Open Access Journals (Sweden)

    Amrita Lama

    2017-11-01

    Full Text Available It is established that the human pathogen Legionella pneumophila becomes significantly augmented for infection of macrophages after intracellular growth in amoebae when compared to like-strains cultivated in laboratory media. Based on this observation, we reasoned that the most critical virulence determinants of L.p. are expressed by responding to stimuli generated by the protozoan host specifically; a process we term “protozoan-priming.” We sought to identify L.p. virulence factors that were required for replication in amoebae in order to highlight the genes necessary for production of the most infectious form of the bacterium. Using a transposon mutagenesis screen, we successfully identified 12 insertions that produced bacteria severely attenuated for growth in amoebae, while retaining a functional Dot/Icm type IVb secretion system. Seven of these insertion mutants were found dispensable for growth in macrophages, revealing attractive therapeutic targets that reside upstream of the pathogen-human interface. Two candidates identified, lpg0730 and lpg0122 were required for survival and replication in amoebae and macrophage host cells. Both genes are conserved among numerous important human pathogenic bacteria that can persist or replicate in amoebae. Each gene encodes a component of an ATP binding cassette (ABC transport complex of unknown function. We demonstrate the lpg0730 ortholog in Francisella tularensis subsp. novicida to be essential for colonization of both protozoan and mammalian host cells, highlighting conserved survival mechanisms employed by bacteria that utilize protozoa as an environmental reservoir for replication.

  13. LegC3, an Effector Protein from Legionella pneumophila, Inhibits Homotypic Yeast Vacuole Fusion In Vivo and In Vitro

    Science.gov (United States)

    Bennett, Terry L.; Kraft, Shannon M.; Reaves, Barbara J.; Mima, Joji; O’Brien, Kevin M.; Starai, Vincent J.

    2013-01-01

    During infection, the intracellular pathogenic bacterium Legionella pneumophila causes an extensive remodeling of host membrane trafficking pathways, both in the construction of a replication-competent vacuole comprised of ER-derived vesicles and plasma membrane components, and in the inhibition of normal phagosome:endosome/lysosome fusion pathways. Here, we identify the LegC3 secreted effector protein from L. pneumophila as able to inhibit a SNARE- and Rab GTPase-dependent membrane fusion pathway in vitro, the homotypic fusion of yeast vacuoles (lysosomes). This vacuole fusion inhibition appeared to be specific, as similar secreted coiled-coiled domain containing proteins from L. pneumophila, LegC7/YlfA and LegC2/YlfB, did not inhibit vacuole fusion. The LegC3-mediated fusion inhibition was reversible by a yeast cytosolic extract, as well as by a purified soluble SNARE, Vam7p. LegC3 blocked the formation of trans-SNARE complexes during vacuole fusion, although we did not detect a direct interaction of LegC3 with the vacuolar SNARE protein complexes required for fusion. Additionally, LegC3 was incapable of inhibiting a defined synthetic model of vacuolar SNARE-driven membrane fusion, further suggesting that LegC3 does not directly inhibit the activity of vacuolar SNAREs, HOPS complex, or Sec17p/18p during membrane fusion. LegC3 is likely utilized by Legionella to modulate eukaryotic membrane fusion events during pathogenesis. PMID:23437241

  14. Biofilm Composition and Threshold Concentration for Growth of Legionella pneumophila on Surfaces Exposed to Flowing Warm Tap Water without Disinfectant.

    Science.gov (United States)

    van der Kooij, Dick; Bakker, Geo L; Italiaander, Ronald; Veenendaal, Harm R; Wullings, Bart A

    2017-03-01

    Legionella pneumophila in potable water installations poses a potential health risk, but quantitative information about its replication in biofilms in relation to water quality is scarce. Therefore, biofilm formation on the surfaces of glass and chlorinated polyvinyl chloride (CPVC) in contact with tap water at 34 to 39°C was investigated under controlled hydraulic conditions in a model system inoculated with biofilm-grown L. pneumophila The biofilm on glass (average steady-state concentration, 23 ± 9 pg ATP cm-2) exposed to treated aerobic groundwater (0.3 mg C liter-1; 1 μg assimilable organic carbon [AOC] liter-1) did not support growth of the organism, which also disappeared from the biofilm on CPVC (49 ± 9 pg ATP cm-2) after initial growth. L. pneumophila attained a level of 4.3 log CFU cm-2 in the biofilms on glass (1,055 ± 225 pg ATP cm-2) and CPVC (2,755 ± 460 pg ATP cm-2) exposed to treated anaerobic groundwater (7.9 mg C liter-1; 10 μg AOC liter-1). An elevated biofilm concentration and growth of L. pneumophila were also observed with tap water from the laboratory. The Betaproteobacteria Piscinibacter and Methyloversatilis and amoeba-resisting Alphaproteobacteria predominated in the clones and isolates retrieved from the biofilms. In the biofilms, the Legionella colony count correlated significantly with the total cell count (TCC), heterotrophic plate count, ATP concentration, and presence of Vermamoeba vermiformis This amoeba was rarely detected at biofilm concentrations of water-associated disease outbreaks reported in the United States. The organism proliferates in biofilms on surfaces exposed to warm water in engineered freshwater installations. An investigation with a test system supplied with different types of warm drinking water without disinfectant under controlled hydraulic conditions showed that treated aerobic groundwater (0.3 mg liter-1 of organic carbon) induced a low biofilm concentration that supported no or very limited growth of L

  15. Energy Conservation and the Promotion of Legionella pneumophila Growth: The Probable Role of Heat Exchangers in a Nosocomial Outbreak.

    Science.gov (United States)

    Bédard, Emilie; Lévesque, Simon; Martin, Philippe; Pinsonneault, Linda; Paranjape, Kiran; Lalancette, Cindy; Dolcé, Charles-Éric; Villion, Manuela; Valiquette, Louis; Faucher, Sébastien P; Prévost, Michèle

    2016-12-01

    OBJECTIVE To determine the source of a Legionella pneumophila serogroup 5 nosocomial outbreak and the role of the heat exchanger installed on the hot water system within the previous year. SETTING A 400-bed tertiary care university hospital in Sherbrooke, Canada. METHODS Hot water samples were collected and cultured for L. pneumophila from 25 taps (baths and sinks) within wing A and 9 taps in wing B. Biofilm (5) and 2 L water samples (3) were collected within the heat exchangers for L. pneumophila culture and detection of protists. Sequence-based typing was performed on strain DNA extracts and pulsed-field gel electrophoresis patterns were analyzed. RESULTS Following 2 cases of hospital-acquired legionellosis, the hot water system investigation revealed a large proportion of L. pneumophila serogroup 5 positive taps (22/25 in wing A and 5/9 in wing B). High positivity was also detected in the heat exchanger of wing A in water samples (3/3) and swabs from the heat exchanger (4/5). The outbreak genotyping investigation identified the hot water system as the source of infections. Genotyping results revealed that all isolated environmental strains harbored the same related pulsed-field gel electrophoresis pattern and sequence-based type. CONCLUSIONS Two cases of hospital-acquired legionellosis occurred in the year following the installation of a heat exchanger to preheat hospital hot water. No cases were reported previously, although the same L. pneumophila strain was isolated from the hot water system in 1995. The heat exchanger promoted L. pneumophila growth and may have contributed to confirmed clinical cases. Infect. Control Hosp. Epidemiol. 2016;1475-1480.

  16. Neumonía por Legionella pneumophila: Experiencia en un Hospital Universitario de Buenos Aires Neumonia due to Legionella pneumophila. Experience gathered in a University Hospital in Buenos Aires

    Directory of Open Access Journals (Sweden)

    Carlos M. Luna

    2004-04-01

    Full Text Available La enfermedad de los legionarios es una causa de neumonía adquirida en la comunidad (NAC reconocida en todo el mundo. En Latinoamérica su incidencia es desconocida. En este estudio se analizó a 9 pacientes con NAC por Legionella pneumophila atendidos entre 1997 y 2001 en el Hospital de Clínicas José de San Martín de la Universidad de Buenos Aires. Se registraron datos de antecedentes, enfermedad actual, contactos, exposición laboral, examen físico, pruebas de laboratorio y uso previo de antibióticos, y se tomó en cuenta la presencia de criterios de gravedad. Nueve pacientes presentaron diagnóstico de NAC por Legionella, ninguno refirió antecedentes de viajes recientes; cuatro de ellos debieron ser internados en unidades de cuidado intensivo. Siete pacientes tenían antecedentes de tabaquismo, 4 tenían EPOC y un paciente linfoma no-Hodgkin. Nuestra casuística corrobora la baja especificidad de la clínica y estudios complementarios para predecir esta etiología. El aislamiento de Legionella es dificultoso, la seroconversión permite el diagnóstico retrospectivo y requiere plazos prolongados y el antígeno urinario aporta un diagnóstico inmediato. Cuando la legionelosis aparece en casos aislados, como ocurriría en Argentina, si no se piensa en esta etiología no se llegará al diagnóstico. Legionella pneumophila es un patógeno de NAC en nuestro medio, debe buscarse mejor, particularmente en pacientes graves, inmunodeprimidos y en fumadores con enfermedad pulmonar obstructiva crónica (EPOC.Legionnaires’ disease is a well recognized cause of community acquired pneumonia (CAP all around the world. In Latin America its incidence remains unknown. This study analyzed a cohort of 9 patients with CAP due to Legionella pneumophila observed from 1997 to 2001, in the Hospital de Clínicas José de San Martín, University of Buenos Aires. Clinical history included recent illnesses, work exposure, physical exam, prior antibiotic use and

  17. Characterization of unrelated clinical Legionella pneumophila isolates in Catalonia by monoclonal subgrouping and sequence-based typing.

    Science.gov (United States)

    Garcia-Nuñez, Marian; Quero, Sara; Pedro-Botet, Maria Lluisa; Barrabeig, Irene; Avarez, Josep; Campoy, Irene; Sala, Maria Rosa; Parraga-Niño, Noemí; Minguell, Sofia; Caylà, Joan; Mateu, Lourdes; Sabria, Miquel

    2016-07-01

    To characterize the genetic diversity of unrelated Legionella pneumophila clinical isolates in Catalonia and compare with other European regions. 95 unrelated isolates were analyzed using monoclonal antibodies and sequence-based typing, 1989-2013. The isolates showed a high diversity (IOD 0.964) with a predominance of some profiles (ST37-Phialdelphia, ST23-Philadelphia and ST1-OLDA). All regions had predominant sequence types (STs) that differed between regions, and only 3% of STs were shared between the three regions. L. pneumophila clinical isolates from Catalonia presented a high diversity and can be used in epidemiological surveillance studies. The heterogeneous predominance of STs between European regions suggested a relationship between geographical distribution and virulence of some STs.

  18. Seasonal distribution of Legionella spp. and L. pneumophila in a river in Taiwan evaluated with culture-confirmed and direct DNA extraction methods

    Science.gov (United States)

    Tung, Min-Che; Chang, Tien-Yu; Hsu, Bing-Mu; Shen, Shu-Min; Huang, Jen-Te; Kao, Po-Min; Chiu, Yi-Chou; Fan, Cheng-Wei; Huang, Yu-Li

    2013-07-01

    In this study, we evaluated the presence and amount of Legionella in along a river in Taiwan, and the relations between seasonal distribution of Legionella spp. and geographic characteristics in the watershed were also evaluated. Water samples were pre-treated and analyzed with culture-confirmed and direct DNA extraction methods. For culture-confirmed method, water samples were cultivated through a series of selective media, and candidate colonies were confirmed by PCR. For direct DNA extraction method, direct DNA extraction was performed from pre-treated water samples. The DNA extracts were analyzed with PCR and DNA sequence analysis for species determination, quantitative PCR (qPCR) was performed to quantify Legionella concentration in the water sample. In all, 150 water samples were included in this study, with 73 (48.6%) water samples detected with Legionella spp., and 17 with L. pneumophila. Over 80% Legionella spp. detections were through direct DNA extraction method, but more than 80% L. pneumophila detections were through culture-confirmed method. While detection of Legionella spp. was done with two methods, positive results were found through only one method. Legionella spp. was detected in all seasons with detection rate ranging between 34.3-58.8% and seasonal average concentration from 1.9 × 102 to 7.1 × 103 CFU/L. Most of the L. pneumophila detections were from samples collected in fall (38.2%) and summer (6.0%), which also coincided with increased cases of Legionellosis reported through Center of Disease Control in Taiwan. The high prevalence and concentration of Legionella spp. and L. pneumophila in the surface waters should be further evaluated for potential health risks.

  19. Crystal Structure of a Legionella pneumophila Ecto -Triphosphate Diphosphohydrolase, A Structural and Functional Homolog of the Eukaryotic NTPDases

    Energy Technology Data Exchange (ETDEWEB)

    Vivian, Julian P.; Riedmaier, Patrice; Ge, Honghua; Le Nours, Jérôme; Sansom, Fiona M.; Wilce, Matthew C.J.; Byres, Emma; Dias, Manisha; Schmidberger, Jason W.; Cowan, Peter J.; d' Apice, Anthony J.F.; Hartland, Elizabeth L.; Rossjohn, Jamie; Beddoe, Travis (Monash); (Melbourne)

    2010-04-19

    Many pathogenic bacteria have sophisticated mechanisms to interfere with the mammalian immune response. These include the disruption of host extracellular ATP levels that, in humans, is tightly regulated by the nucleoside triphosphate diphosphohydrolase family (NTPDases). NTPDases are found almost exclusively in eukaryotes, the notable exception being their presence in some pathogenic prokaryotes. To address the function of bacterial NTPDases, we describe the structures of an NTPDase from the pathogen Legionella pneumophila (Lpg1905/Lp1NTPDase) in its apo state and in complex with the ATP analog AMPPNP and the subtype-specific NTPDase inhibitor ARL 67156. Lp1NTPDase is structurally and catalytically related to eukaryotic NTPDases and the structure provides a basis for NTPDase-specific inhibition. Furthermore, we demonstrate that the activity of Lp1NTPDase correlates directly with intracellular replication of Legionella within macrophages. Collectively, these findings provide insight into the mechanism of this enzyme and highlight its role in host-pathogen interactions.

  20. Incubation of premise plumbing water samples on Buffered Charcoal Yeast Extract agar at elevated temperature and pH selects for Legionella pneumophila.

    Science.gov (United States)

    Veenendaal, Harm R; Brouwer-Hanzens, Anke J; van der Kooij, Dick

    2017-10-15

    Worldwide, over 90% of the notified cases of Legionnaires' disease are caused by Legionella pneumophila. However, the standard culture medium for the detection of Legionella in environmental water samples, Buffered Charcoal Yeast Extract (BCYE) agar of pH 6.9 ± 0.4 with or without antimicrobial agents incubated at 36 ± 1 °C, supports the growth of a large diversity of Legionella species. BCYE agar of elevated pH or/and incubation at elevated temperature gave strongly reduced recoveries of most of 26 L. non-pneumophila spp. tested, but not of L. pneumophila. BCYE agar of pH 7.3 ± 0.1, incubated at 40 ± 0.5 °C (BCYE pH 7.3/40 °C) was tested for selective enumeration of L. pneumophila. Of the L. non-pneumophila spp. tested, only L. adelaidensis and L. londiniensis multiplied under these conditions. The colony counts on BCYE pH 7.3/40 °C of a L. pneumophila serogroup 1 strain cultured in tap water did not differ significantly from those on BCYE pH 6.9/36 °C when directly plated and after membrane filtration and showed repeatability's of 13-14%. By using membrane filtration L. pneumophila was detected in 58 (54%) of 107 Legionella-positive water samples from premise plumbing systems under one or both of these culture conditions. The L. pneumophila colony counts (log-transformed) on BCYE pH 7.3/40 °C were strongly related (r2 = 0.87) to those on BCYE pH 6.9/36 °C, but differed significantly (p < 0.05) by a mean of - 0.12 ± 0.30 logs. L. non-pneumophila spp. were detected only on BCYE pH 6.9/36 °C in 49 (46%) of the samples. Hence, BCYE pH 7.3/40 °C can facilitate the enumeration of L. pneumophila and their isolation from premise plumbing systems with culturable L. non-pneumophila spp., some of which, e.g. L. anisa, can be present in high numbers. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. Brote epidémico de neumonías por Legionella pneumophila en niños cubanos

    Directory of Open Access Journals (Sweden)

    Roberto Razón Behar

    2002-09-01

    Full Text Available La Legionella pneumophila es uno de los patógenos responsable de neumonías atípicas, a través de la inhalación de aerosoles o aspiración de líquidos infectados. Se detectó un brote epidémico de neumonías por Legionella, originado por la aspiración de agua contaminada de una piscina en un grupo de niños cubanos. El agente causal se identificó en 5 de 9 pacientes, por la técnica de inmunofluorescencia indirecta en muestras de sueros pareados. Los síntomas y signos más frecuentes fueron malestar general, anorexia, astenia, fiebre persistente de 39 °C a 40 °C (103 °F a 105 °F, mialgias, cefaleas, náuseas, vómitos, dolor abdominal, diarreas, tos húmeda, dolor torácico y polipnea. Durante el desarrollo de la enfermedad, el tratamiento antibiótico fue empírico (incluyendo los macrólidos, por no tener confirmado el diagnóstico. Todos los pacientes evolucionaron satisfactoriamente. Se reportó un brote epidémico de neumonías por Legionella en niños por primera vez en Cuba, lo cual tiene importancia clínica y epidemiológica.The legionella pneumophila is one of the pathogens responsible for atypic pneumonias by the inhalation of aerosols or aspiration of infected liquids. An epidemic outbreak of pneumonias caused by Legionella was detected among a group of Cuban children. It was originated by the aspiration of contaminated water in a swimming pool. The causal agent was identified in 5 of 9 patients by using the indirect immunofluorescence technique in samples of matched sera. The most frequent symptoms and signs were malaise, anorexia, asthenia, persistent fever from 39°C to 40°C (103° F to 105° F, myalgias, headache, nauseas, vomits, abdominal pain, diarrheas, moist cough, thoracic pain and polypnoea. The antibiotic treatment was empiric (including the macrolides during the development of the disease, since the diagnosis was not confirmed. The patients’ evolution was satisfactory. An epidemic outbreak of pneumonias

  2. Genome Analysis of Legionella pneumophila Strains Using a Mixed-Genome Microarray

    NARCIS (Netherlands)

    Euser, S.M.; Nagelkerke, N.J.; Schuren, F.; Jansen, R.; Boer, J.W. den

    2012-01-01

    Background: Legionella, the causative agent for Legionnaires' disease, is ubiquitous in both natural and man-made aquatic environments. The distribution of Legionella genotypes within clinical strains is significantly different from that found in environmental strains. Developing novel genotypic

  3. Comparison of pulsed corona plasma and pulsed electric fields for the decontamination of water containing Legionella pneumophila as model organism.

    Science.gov (United States)

    Banaschik, Robert; Burchhardt, Gerhard; Zocher, Katja; Hammerschmidt, Sven; Kolb, Juergen F; Weltmann, Klaus-Dieter

    2016-12-01

    Pulsed corona plasma and pulsed electric fields were assessed for their capacity to kill Legionella pneumophila in water. Electrical parameters such as in particular dissipated energy were equal for both treatments. This was accomplished by changing the polarity of the applied high voltage pulses in a coaxial electrode geometry resulting in the generation of corona plasma or an electric field. For corona plasma, generated by high voltage pulses with peak voltages of +80kV, Legionella were completely killed, corresponding to a log-reduction of 5.4 (CFU/ml) after a treatment time of 12.5min. For the application of pulsed electric fields from peak voltages of -80kV a survival of log 2.54 (CFU/ml) was still detectable after this treatment time. Scanning electron microscopy images of L. pneumophila showed rupture of cells after plasma treatment. In contrast, the morphology of bacteria seems to be intact after application of pulsed electric fields. The more efficient killing for the same energy input observed for pulsed corona plasma is likely due to induced chemical processes and the generation of reactive species as indicated by the evolution of hydrogen peroxide. This suggests that the higher efficacy and efficiency of pulsed corona plasma is primarily associated with the combined effect of the applied electric fields and the promoted reaction chemistry. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. Type IV secretion-dependent activation of host MAP kinases induces an increased proinflammatory cytokine response to Legionella pneumophila.

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    Sunny Shin

    2008-11-01

    Full Text Available The immune system must discriminate between pathogenic and nonpathogenic microbes in order to initiate an appropriate response. Toll-like receptors (TLRs detect microbial components common to both pathogenic and nonpathogenic bacteria, whereas Nod-like receptors (NLRs sense microbial components introduced into the host cytosol by the specialized secretion systems or pore-forming toxins of bacterial pathogens. The host signaling pathways that respond to bacterial secretion systems remain poorly understood. Infection with the pathogen Legionella pneumophila, which utilizes a type IV secretion system (T4SS, induced an increased proinflammatory cytokine response compared to avirulent bacteria in which the T4SS was inactivated. This enhanced response involved NF-kappaB activation by TLR signaling as well as Nod1 and Nod2 detection of type IV secretion. Furthermore, a TLR- and RIP2-independent pathway leading to p38 and SAPK/JNK MAPK activation was found to play an equally important role in the host response to virulent L. pneumophila. Activation of this MAPK pathway was T4SS-dependent and coordinated with TLR signaling to mount a robust proinflammatory cytokine response to virulent L. pneumophila. These findings define a previously uncharacterized host response to bacterial type IV secretion that activates MAPK signaling and demonstrate that coincident detection of multiple bacterial components enables immune discrimination between virulent and avirulent bacteria.

  5. Type II Secretion Substrates of Legionella pneumophila Translocate Out of the Pathogen-Occupied Vacuole via a Semipermeable Membrane

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    Hilary K. Truchan

    2017-06-01

    Full Text Available Legionella pneumophila replicates in macrophages in a host-derived phagosome, termed the Legionella-containing vacuole (LCV. While the translocation of type IV secretion (T4S effectors into the macrophage cytosol is well established, the location of type II secretion (T2S substrates in the infected host cell is unknown. Here, we show that the T2S substrate ProA, a metalloprotease, translocates into the cytosol of human macrophages, where it associates with the LCV membrane (LCVM. Translocation is detected as early as 10 h postinoculation (p.i., which is approximately the midpoint of the intracellular life cycle. However, it is detected as early as 6 h p.i. if ProA is hyperexpressed, indicating that translocation depends on the timing of ProA expression and that any other factors necessary for translocation are in place by that time point. Translocation occurs with all L. pneumophila strains tested and in amoebae, natural hosts for L. pneumophila. It was absent in murine bone marrow-derived macrophages and murine macrophage cell lines. The ChiA chitinase also associated with the cytoplasmic face of the LCVM at 6 h p.i. and in a T2S-dependent manner. Galectin-3 and galectin-8, eukaryotic proteins whose localization is influenced by damage to host membranes, appeared within the LCV of infected human but not murine macrophages beginning at 6 h p.i. Thus, we hypothesize that ProA and ChiA are first secreted into the vacuolar lumen by the activity of the T2S and subsequently traffic into the macrophage cytosol via a novel mechanism that involves a semipermeable LCVM.

  6. Legionella pneumophila in bronchoalveolar lavage samples of patients suffering from severe respiratory infections: Role of age, sex and history of smoking in the prevalence of bacterium

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    Faradonbeh Fatemeh Alaei

    2015-01-01

    Full Text Available Introduction. Legionella pneumophila is the most commonly detected cause of legionellosis, which is an acute respiratory tract infection with high morbidity and mortality rates. Objective. The aim of this study was to investigate the prevalence rate of L. pneumophila in bronchoalveolar lavages and study the role of sex, age and history of smoking as risk factors for susceptibility to the bacterium. Methods. One hundred bronchoalveolar lavage samples were collected from the Iranian health centers and immediately transferred to laboratory. The samples were cultured and those that were L. pneumophila positive were subjected to PCR method with respect to the 16S rRNA gene. Results. Twelve out of 100 samples were positive for L. pneumophila (12%. Patients older than 70 years had the highest incidence of L. pneumophila (17.77%. Prevalence of L. pneumophila in male and female patients was 14.81% and 8.69%, respectively. Total incidence of L. pneumophila in patients with and without history of smoking was 18% and 6%, respectively. There were significant differences in the incidence of bacterium between groups of our study. Conclusion. Sex, age and history of smoking are predominant risk factors for the occurrence of L. pneumophila. However, more studies should be undertaken to confirm these results.

  7. Studio del comportamento di Acanthamoeba. polyphaga in presenza di Legionella pneumophila e di altri batteri ad habitat acquatico

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    M. Bondi

    2003-05-01

    Full Text Available Le amebe a vita libera sono state oggetto di diversi studi negli ultimi anni, non solo per le loro potenzialità patogene nei confronti dell’uomo, ma anche per l’importante ruolo che svolgono in natura, dove agiscono come predatori in grado di controllare le popolazioni batteriche. Alcuni degli organismi fagocitati però possono evitare la lisi fagosomiale e mantenere la loro condizione vitale a livello intracellulare, divenendo endosimbionti. Le amebe fungono così da riserva per questi batteri, proteggendoli da difficili condizioni extracellulari e provvedendo ad un ambiente consono alla loro replicazione. Tale tipo di interazione è particolarmente studiata in Legionella pneumophila, dal momento che l’ampia diffusione di questo germe, nonché la sua virulenza, pare siano fortemente influenzate dalla capacità di parassitare protozoi appartenenti ai generi Acanthamoeba, Naegleria e Balamuthia. Al fine di ottenere maggiori informazioni sui fattori favorenti o inibenti lo sviluppo di questi protozoi, è stato studiato il comportamento di un ceppo di Acanthamoeba polyphaga coltivato, in solido e in liquido, in associazione con L. pneumophila ed altri batteri ad habitat acquatico (Pseudomonas, Aeromonas, Achromobacter, Burkholderia. Su tappeti di cellule batteriche allestiti in Non Nutrient Agar (NNA, A.polyphaga si è mostrata in grado di moltiplicarsi utilizzando come nutrimento tutti i ceppi testati, nonostante alcuni, come Burkholderia cepacia SSV6 e Achromobacter xylox SS28, risultino più idonei al suo sviluppo. In piastre a pozzetti addizionate di acqua condottata autoclavata, il protozoo ha mostrato una buona capacità di sopravvivenza, non risultando inoltre influenzato dalla presenza di legionella o dei batteri acquatici testati. Dal momento che, fra i batteri descritti come capaci di vita intra-amebica, sono inclusi patogeni quali Chlamydia, Legionella, Listeria e Rickettsiae, risulta necessario riconsiderare la rilevanza clinica

  8. Proteomic analysis of growth phase-dependent expression of Legionella pneumophila proteins which involves regulation of bacterial virulence traits.

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    Tsuyoshi Hayashi

    Full Text Available Legionella pneumophila, which is a causative pathogen of Legionnaires' disease, expresses its virulent traits in response to growth conditions. In particular, it is known to become virulent at a post-exponential phase in vitro culture. In this study, we performed a proteomic analysis of differences in expression between the exponential phase and post-exponential phase to identify candidates associated with L. pneumophila virulence using 2-Dimentional Fluorescence Difference Gel Electrophoresis (2D-DIGE combined with Matrix-Assisted Laser Desorption/Ionization-Mass Spectrometry (MALDI-TOF-MS. Of 68 identified proteins that significantly differed in expression between the two growth phases, 64 were up-regulated at a post-exponential phase. The up-regulated proteins included enzymes related to glycolysis, ketone body biogenesis and poly-3-hydroxybutyrate (PHB biogenesis, suggesting that L. pneumophila may utilize sugars and lipids as energy sources, when amino acids become scarce. Proteins related to motility (flagella components and twitching motility-associated proteins were also up-regulated, predicting that they enhance infectivity of the bacteria in host cells under certain conditions. Furthermore, 9 up-regulated proteins of unknown function were found. Two of them were identified as novel bacterial factors associated with hemolysis of sheep red blood cells (SRBCs. Another 2 were found to be translocated into macrophages via the Icm/Dot type IV secretion apparatus as effector candidates in a reporter assay with Bordetella pertussis adenylate cyclase. The study will be helpful for virulent analysis of L. pneumophila from the viewpoint of physiological or metabolic modulation dependent on growth phase.

  9. Multiplex polymerase chain reaction of genetic markers for detection of potentially pathogenic environmental Legionella pneumophila isolates

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    Arvind Valavane

    2017-01-01

    Interpretation & conclusions: The findings of this study showed the presence of gene markers of pathogenic potential of the bacterium L. pneumophila. However, the genomic constitution of the environmental isolates should be correlated with clinical isolates to prove their pathogenic potential. Rapid diagnostic methods such as multiplex PCR reported here, for elucidating gene markers, could help in future epidemiological studies of bacterium L. pneumophila.

  10. A new ELISA method for serological diagnosis of Legionella pneumophila: use of five purified proteins, FLA, MOMP, MIP, IP, and PILE, as diagnostic antigen.

    Science.gov (United States)

    Sun, Hui; Ma, Haifeng; Liu, Li; Cao, Xiuqin; Yang, Zhiwei

    2015-01-01

    Legionella pneumophila plays an important role in human infection. Commercial ELISA kits commonly used, which take Legionella pneumophila whole-cell protein as the coating antigen, often have cross-reactivity among serogroups or species. In this study, five Legionella pneumophila proteins FLA, MOMP, MIP, IP, and PILE were purified and further applied in serological diagnosis of Legionella pneumophila infections compared with R & D Legionella ELISA kits. The five recombinant plasmids pET-fla, pET-momp, pET-mip, pET-ip, and pET-pile were transformed into E. coli BL21 and then induced them with IPTG. The expression products were analyzed by SDS-PAGE and purified by affinity chromatography. Indirect enzyme-linked immunosorbent assays (ELISAs) were established with the five purified proteins FLA, MOMP, MIP, IP, and PILE altogether as the coating antigen and tested for the presence of IgG, IgM, and IgA antibody independently from 50 positive sera and 40 negative sera, compared with R & D IgG, IgM, and IgA Legionella ELISA kits. The FLA protein about 42 kDa in size, the MOMP protein about 45 kDa, the MIP protein about 40 kDa, the IP protein about 46 kDa, and the PILE protein about 35.7 kDa were separately expressed and purified. Compared with R & D IgG, IgM, and IgA Legionella ELISA kit, the outcome of indirect ELISAs set up with the five purified proteins showed that for IgG the sensitivity was 90.4%, the specificity was 97.4%, the area under ROC curve was 0.939, the kappa value was 0.865, the 95% confidence interval was 0.883 - 0.995. For IgM the sensitivity was 91.8%, the specificity was 95.1%, the area under ROC curve was 0.935, the kappa value was 0.866, the 95% confidence interval was 0.876 - 0.994. For IgA the sensitivity was 93.6%, the specificity was 95.3%, the area under ROC curve was 0.945, the kappa value was 0.889, the 95% confidence interval was 0.890 - 0.999. The proteins FLA, MOMP, MIP, IP, and PILE were successfully expressed and purified, and they seemed

  11. Rainfall is a risk factor for sporadic cases of Legionella pneumophila pneumonia.

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    Carolina Garcia-Vidal

    Full Text Available It is not known whether rainfall increases the risk of sporadic cases of Legionella pneumonia. We sought to test this hypothesis in a prospective observational cohort study of non-immunosuppressed adults hospitalized for community-acquired pneumonia (1995-2011. Cases with Legionella pneumonia were compared with those with non-Legionella pneumonia. Using daily rainfall data obtained from the regional meteorological service we examined patterns of rainfall over the days prior to admission in each study group. Of 4168 patients, 231 (5.5% had Legionella pneumonia. The diagnosis was based on one or more of the following: sputum (41 cases, antigenuria (206 and serology (98. Daily rainfall average was 0.556 liters/m(2 in the Legionella pneumonia group vs. 0.328 liters/m(2 for non-Legionella pneumonia cases (p = 0.04. A ROC curve was plotted to compare the incidence of Legionella pneumonia and the weighted median rainfall. The cut-off point was 0.42 (AUC 0.54. Patients who were admitted to hospital with a prior weighted median rainfall higher than 0.42 were more likely to have Legionella pneumonia (OR 1.35; 95% CI 1.02-1.78; p = .03. Spearman Rho correlations revealed a relationship between Legionella pneumonia and rainfall average during each two-week reporting period (0.14; p = 0.003. No relationship was found between rainfall average and non-Legionella pneumonia cases (-0.06; p = 0.24. As a conclusion, rainfall is a significant risk factor for sporadic Legionella pneumonia. Physicians should carefully consider Legionella pneumonia when selecting diagnostic tests and antimicrobial therapy for patients presenting with CAP after periods of rainfall.

  12. Molecular epidemiology of Legionella pneumophila serogroup 1 isolates following long-term chlorine dioxide treatment in a university hospital water system.

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    Casini, B; Valentini, P; Baggiani, A; Torracca, F; Frateschi, S; Nelli, L Ceccherini; Privitera, G

    2008-06-01

    This paper describes the results of a five-year monitoring programme applied to the water distribution system of the University Hospital of Pisa (Italy). The purpose of the programme was to evaluate the efficacy of an integrated water safety plan in controlling Legionella spp. colonisation of the potable water system. The impact of the safety plan on the ecology of legionella in the water network was evaluated by studying the genetic variability and the chlorine susceptibility of the strains isolated prior to, and throughout, the application of continuous chlorine dioxide treatment. After 45 months of water hyperchlorination, Legionella spp. were still present but the positive supply points were reduced by 79.4%. The samples exceeding 10(3)cfu/L were reduced by 83.8% and the mean counts showed a decrease of 94.6%. The majority of the isolates belonged to Legionella pneumophila serogroup 1 (overall positivity rate: 161/423; 38%). Molecular typing was performed on 61 isolates (37.9% of the positive samples) selected on spatial and temporal criteria. This revealed the circulation and the persistence in the hospital environment of three prevalent types of L. pneumophila Wadsworth, demonstrating allelic and electrophoretic characteristic profiles and different chlorine susceptibility. Two of these, one predominant and pre-dating the sanitation regimen, and one other isolated after three years of water treatment, were chlorine tolerant. Despite the ineffectiveness of chlorine dioxide in eradicating L. pneumophila, the risk management plan adopted appeared to discourage further cases of nosocomial legionellosis.

  13. Assessment of antibiotic susceptibility of Legionella pneumophila isolated from water systems in Poland

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    Agnieszka Sikora

    2017-03-01

    Azithromycin resistance was found in one strain of L. pneumophila SG 2–14, but the resistance mechanism is unknown and needs further study. It is possible that therapeutic failures in Legionnaires’ disease may be associated with bacterial resistance which should be taken into account. The antibiotic sensitivity testing described in this study could be helpful in detecting the resistance of clinical L. pneumophila isolates. Ciprofloxacin and rifampicin have good in vitro activity against environmental L. pneumophila SG 1 and SG 2–14 in Poland.

  14. UV-A photocatalytic treatment of Legionella pneumophila bacteria contaminated airflows through three-dimensional solid foam structured photocatalytic reactors.

    Science.gov (United States)

    Josset, Sébastien; Hajiesmaili, Shabnam; Begin, Dominique; Edouard, David; Pham-Huu, Cuong; Lett, Marie-Claire; Keller, Nicolas; Keller, Valérie

    2010-03-15

    A 3D-structured photocatalytic media was designed for allowing a tubular reactor to work in a traversing-flow mode at low pressure drops with a strong increase in the surface area-to-volume ratio inside the reactor. A protective polysiloxane coating was performed for protecting a structured polyurethane foam and anchoring the active TiO(2) particles. Filled with the 3D-structured solid foam supporting TiO(2) photocatalyst, the reactor could thus take advantages from the static mixer effect and from the low pressure drop resulting from the reticulated foam support. Very efficient decontamination levels towards airborne Legionella pneumophila bacteria were reached in a single-pass test mode. (c) 2009 Elsevier B.V. All rights reserved.

  15. UV-A photocatalytic treatment of Legionella pneumophila bacteria contaminated airflows through three-dimensional solid foam structured photocatalytic reactors

    Energy Technology Data Exchange (ETDEWEB)

    Josset, Sebastien; Hajiesmaili, Shabnam; Begin, Dominique; Edouard, David; Pham-Huu, Cuong [Laboratoire des Materiaux, Surfaces et Procedes pour la Catalyse (LMSPC), European Laboratory for Catalysis and Surface Sciences (ELCASS), CNRS, Strasbourg University, 25 rue Becquerel 67087 Strasbourg (France); Lett, Marie-Claire [Laboratoire de Genetique Moleculaire, Genomique, Microbiologie, CNRS, Strasbourg University, 28, rue Goethe 67083 Strasbourg Cedex (France); Keller, Nicolas, E-mail: nkeller@chimie.u-strasbg.fr [Laboratoire des Materiaux, Surfaces et Procedes pour la Catalyse (LMSPC), European Laboratory for Catalysis and Surface Sciences (ELCASS), CNRS, Strasbourg University, 25 rue Becquerel 67087 Strasbourg (France); Keller, Valerie [Laboratoire des Materiaux, Surfaces et Procedes pour la Catalyse (LMSPC), European Laboratory for Catalysis and Surface Sciences (ELCASS), CNRS, Strasbourg University, 25 rue Becquerel 67087 Strasbourg (France)

    2010-03-15

    A 3D-structured photocatalytic media was designed for allowing a tubular reactor to work in a traversing-flow mode at low pressure drops with a strong increase in the surface area-to-volume ratio inside the reactor. A protective polysiloxane coating was performed for protecting a structured polyurethane foam and anchoring the active TiO{sub 2} particles. Filled with the 3D-structured solid foam supporting TiO{sub 2} photocatalyst, the reactor could thus take advantages from the static mixer effect and from the low pressure drop resulting from the reticulated foam support. Very efficient decontamination levels towards airborne Legionella pneumophila bacteria were reached in a single-pass test mode.

  16. Structure and Function of Interacting IcmR-IcmQ Domains from a Type IVb Secretion System in Legionella pneumophila

    Energy Technology Data Exchange (ETDEWEB)

    Raychaudhury, S.; Farelli, J; Montminy, T; Matthews, M; Menetret, J; Dumenil, G; Roy, C; Head, J; Isberg, R; Akey, C

    2009-01-01

    During infection, Legionella pneumophila creates a replication vacuole within eukaryotic cells and this requires a Type IVb secretion system (T4bSS). IcmQ plays a critical role in the translocase and associates with IcmR. In this paper, we show that the N-terminal domain of IcmQ (Qn) mediates self-dimerization, whereas the C-terminal domain with a basic linker promotes membrane association. In addition, the binding of IcmR to IcmQ prevents self-dimerization and also blocks membrane permeabilization. However, IcmR does not completely block membrane binding by IcmQ. We then determined crystal structures of Qn with the interacting region of IcmR. In this complex, each protein forms an ?-helical hairpin within a parallel four-helix bundle. The amphipathic nature of helices in Qn suggests two possible models for membrane permeabilization by IcmQ. The Rm-Qn structure also suggests how IcmR-like proteins in other L. pneumophila species may interact with their IcmQ partners.

  17. Investigation of plasma-functionalized multiwalled carbon nanotube film and its application of DNA sensor for Legionella pneumophila detection.

    Science.gov (United States)

    Park, Eun Jin; Lee, Jun-Yong; Kim, Jun Hyup; Lee, Cheol Jin; Kim, H Stanley; Min, Nam Ki

    2010-08-15

    A novel multiwall carbon nanotube (MWCNT) electrode functionalized with oxygen plasma treatment was prepared and characterized, and its DNA sensing ability for Legionella pneumophila (L. pneumophila) detection was examined using electrochemical measurement. A well-patterned MWCNT working electrode (WE) on a Pt track was fabricated using photolithography, transfer methods and an etching technique. The MWCNT WE was functionalized by oxygen plasma treatment prior to applying for DNA sensor. The surface morphology of the plasma-functionalized MWCNT (pf-MWCNT) WEs were observed by scanning electron microscope (SEM) and the change of chemical composition was characterized by X-ray photoelectron spectroscopy (XPS), and electrochemical measurements were performed using CV with ferricyanide/ferrocyanide redox couple. Effective areas of working electrodes were calculated to be 0.00453 cm(2) for pristine MWCNT electrode and 0.00747-0.00874 cm(2) for pf-MWCNT electrodes with different plasma treatment times. Differential pulse voltammetry (DPV) was carried out in methylene blue solution for DNA sensing. The pf-MWCNT based DNA sensor was successfully operated in a target concentration range of 10 pM to 100 nM and had a lower detection limit than a pristine MWCNT based DNA sensor. Copyright 2010 Elsevier B.V. All rights reserved.

  18. Electrochemical Characterization of O2 Plasma Functionalized Multi-Walled Carbon Nanotube Electrode for Legionella pneumophila DNA Sensor

    Science.gov (United States)

    Park, Eun Jin; Lee, Jun-Yong; Hyup Kim, Jun; Kug Kim, Sun; Lee, Cheol Jin; Min, Nam Ki

    2010-08-01

    An electrochemical DNA sensor for Legionella pneumophila detection was constructed using O2 plasma functionalized multi-walled carbon nanotube (MWCNT) film as a working electrode (WE). The cyclic voltammetry (CV) results revealed that the electrocatalytic activity of plasma functionalized MWCNT (pf-MWCNT) significantly changed depending on O2 plasma treatment time due to some oxygen containing functional groups on the pf-MWCNT surface. Scanning electron microscope (SEM) images and X-ray photoelectron spectroscopy (XPS) spectra were also presented the changes of their surface morphologies and oxygen composition before and after plasma treatment. From a comparison study, it was found that the pf-MWCNT WEs had higher electrocatalytic activity and more capability of probe DNA immobilization: therefore, electrochemical signal changes by probe DNA immobilization and hybridization on pf-MWCNT WEs were larger than on Au WEs. The pf-MWCNT based DNA sensor was able to detect a concentration range of 10 pM-100 nM of target DNA to detect L. pneumophila.

  19. Natural transformation occurs independently of the essential actin-like MreB cytoskeleton in Legionella pneumophila.

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    Juan, Pierre-Alexandre; Attaiech, Laetitia; Charpentier, Xavier

    2015-11-03

    Natural transformation is the process by which bacteria can actively take up and integrate exogenous DNA thereby providing a source of genetic diversity. Under specific growth conditions the coordinated expression of several genes--a situation referred to as "competence"--allows bacteria to assemble a highly processive and dedicated system that can import high molecular weight DNA. Within the cell these large imported DNA molecules are protected from degradation and brought to the chromosome for recombination. Here, we report elevated expression of mreB during competence in the Gram-negative pathogen Legionella pneumophila. Interestingly a similar observation had previously been reported in the distantly-related Gram-positive organism Bacillus subtilis. MreB is often viewed as the bacterial actin homolog contributing to bacterial morphogenesis by coordinating peptidoglycan-synthesising complexes. In addition MreB is increasingly found to be involved in a growing number of processes including chromosome segregation and motor-driven motility. Using genetic and pharmacological approaches, we examined the possible role of MreB during natural transformation in L. pneumophila. Our data show that natural transformation does not require MreB dynamics and exclude a direct role of MreB filaments in the transport of foreign DNA and its recombination in the chromosome.

  20. Phenylalanine hydroxylase from Legionella pneumophila is a thermostable enzyme with a major functional role in pyomelanin synthesis.

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    Marte I Flydal

    Full Text Available Legionella pneumophila is a pathogenic bacterium that can cause Legionnaires' disease and other non-pneumonic infections in humans. This bacterium produces a pyomelanin pigment, a potential virulence factor with ferric reductase activity. In this work, we have investigated the role of phenylalanine hydroxylase from L. pneumophila (lpPAH, the product of the phhA gene, in the synthesis of the pyomelanin pigment and the growth of the bacterium in defined compositions.Comparative studies of wild-type and phhA mutant corroborate that lpPAH provides the excess tyrosine for pigment synthesis. phhA and letA (gacA appear transcriptionally linked when bacteria were grown in buffered yeast extract medium at 37°C. phhA is expressed in L. pneumophila growing in macrophages. We also cloned and characterized lpPAH, which showed many characteristics of other PAHs studied so far, including Fe(II requirement for activity. However, it also showed many particular properties such as dimerization, a high conformational thermal stability, with a midpoint denaturation temperature (T(m = 79 ± 0.5°C, a high specific activity at 37°C (10.2 ± 0.3 µmol L-Tyr/mg/min and low affinity for the substrate (K(m (L-Phe = 735 ± 50 µM.lpPAH has a major functional role in the synthesis of pyomelanin and promotes growth in low-tyrosine media. The high thermal stability of lpPAH might reflect the adaptation of the enzyme to withstand relatively high survival temperatures.

  1. Short-Term and Long-Term Survival and Virulence of Legionella pneumophila in the Defined Freshwater Medium Fraquil.

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    Nilmini Mendis

    Full Text Available Legionella pneumophila (Lp is the etiological agent responsible for Legionnaires' disease, a potentially fatal pulmonary infection. Lp lives and multiplies inside protozoa in a variety of natural and man-made water systems prior to human infection. Fraquil, a defined freshwater medium, was used as a highly reproducible medium to study the behaviour of Lp in water. Adopting a reductionist approach, Fraquil was used to study the impact of temperature, pH and trace metal levels on the survival and subsequent intracellular multiplication of Lp in Acanthamoeba castellanii, a freshwater protozoan and a natural host of Legionella. We show that temperature has a significant impact on the short- and long-term survival of Lp, but that the bacterium retains intracellular multiplication potential for over six months in Fraquil. Moreover, incubation in Fraquil at pH 4.0 resulted in a rapid decline in colony forming units, but was not detrimental to intracellular multiplication. In contrast, variations in trace metal concentrations had no impact on either survival or intracellular multiplication in amoeba. Our data show that Lp is a resilient bacterium in the water environment, remaining infectious to host cells after six months under the nutrient-deprived conditions of Fraquil.

  2. Gezondheidsaspecten van Legionella in water

    NARCIS (Netherlands)

    Schets FM; de Roda Husman AM; MGB

    2004-01-01

    Legionella pneumophila is een veroorzaker van ernstige longontsteking bij mensen. Legionella bacterien blijken overal in waterige milieus aanwezig te zijn. Met name in kunstmatige waterige milieus kunnen mensen blootgesteld worden aan aerosolen waarin Legionella aanwezig is. Dit beknopte

  3. Identification and characterization of genes, encoding the 3-hydroxybutyrate dehydrogenase and a putative lipase, in an avirulent spontaneous Legionella pneumophila serogroup 6 mutant.

    Science.gov (United States)

    Scaturro, Maria; Barello, Cristina; Giusti, Melania De; Fontana, Stefano; Pinci, Federica; Giuffrida, Maria Gabriella; Ricci, Maria Luisa

    2015-04-01

    Legionella pneumophila is a pathogen widespread in aquatic environment, able to multiply both within amoebae and human macrophages. The aim of this study was to identify genes differently expressed in a spontaneous avirulent Legionella pneumophila serogroup 6 mutant, named Vir-, respect the parental strain (Vir+), and to determine their role in the loss of virulence. Protein profiles revealed some differences in Vir- proteomic maps, and among the identified proteins the undetectable 3-hydroxybutyrate dehydrogenase (BdhA) and a down-produced lipase. Both Legionella enzymes were studied before and were here further characterized at genetic level. A significant down-regulation of both genes was observed in Vir- at the transcriptional level, but the use of defined mutants demonstrated that they did not affect the intracellular multiplication. A mutant (MS1) showed an accumulation of poly-3-hydroxybutyrate (PHB) granules suggesting a role of bdhA gene in its degradation process. The lipase deduced amino acid sequence revealed a catalytic triad, typical of the 'lipase box' characteristic of PHB de-polymerase enzymes, that let us suppose a possible involvement of lipase in the PHB granule degradation process. Our results revealed unexpected alterations in secondary metabolic pathways possibly linking the loss of virulence to Legionella lack of energy sources. © 2014 APMIS. Published by John Wiley & Sons Ltd.

  4. The Legionella pneumophila IcmSW complex interacts with multiple Dot/Icm effectors to facilitate type IV translocation.

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    Eric D Cambronne

    2007-12-01

    Full Text Available Many gram-negative pathogens use a type IV secretion system (T4SS to deliver effector proteins into eukaryotic host cells. The fidelity of protein translocation depends on the efficient recognition of effector proteins by the T4SS. Legionella pneumophila delivers a large number of effector proteins into eukaryotic cells using the Dot/Icm T4SS. How the Dot/Icm system is able to recognize and control the delivery of effectors is poorly understood. Recent studies suggest that the IcmS and IcmW proteins interact to form a stable complex that facilitates translocation of effector proteins by the Dot/Icm system by an unknown mechanism. Here we demonstrate that the IcmSW complex is necessary for the productive translocation of multiple Dot/Icm effector proteins. Effector proteins that were able to bind IcmSW in vitro required icmS and icmW for efficient translocation into eukaryotic cells during L. pneumophila infection. We identified regions in the effector protein SidG involved in icmSW-dependent translocation. Although the full-length SidG protein was translocated by an icmSW-dependent mechanism, deletion of amino terminal regions in the SidG protein resulted in icmSW-independent translocation, indicating that the IcmSW complex is not contributing directly to recognition of effector proteins by the Dot/Icm system. Biochemical and genetic studies showed that the IcmSW complex interacts with a central region of the SidG protein. The IcmSW interaction resulted in a conformational change in the SidG protein as determined by differences in protease sensitivity in vitro. These data suggest that IcmSW binding to effectors could enhance effector protein delivery by mediating a conformational change that facilitates T4SS recognition of a translocation domain located in the carboxyl region of the effector protein.

  5. Efficacy of thermal treatment and copper-silver ionization for controlling Legionella pneumophila in high-volume hot water plumbing systems in hospitals.

    Science.gov (United States)

    Mietzner, S; Schwille, R C; Farley, A; Wald, E R; Ge, J H; States, S J; Libert, T; Wadowsky, R M; Miuetzner, S

    1997-12-01

    Thermal treatment and copper-silver ionization are often used for controlling Legionella pneumophila in high-volume hospital plumbing systems, although the comparative efficacies of these measures in high-volume systems are unknown. Thermal treatment of a hot water circuit was accomplished by flushing hot water (> 60 degrees C) through distal fixtures for 10 minutes. Copper-silver ionization was conducted in three circuits by installing units into return lines immediately upstream from hot water tanks. Recovery rates of L. pneumophila were monitored by culturing swab samples from faucets. Concentrations of copper and silver in water samples were determined by atomic absorption spectrophotometry. Four heat-flush treatments failed to provide long-term control of L. pneumophila. In contrast, ionization treatment reduced the rate of recovery of L. pneumophila from 108 faucets from 72% to 2% within 1 month and maintained effective control for at least 22 months. Only three samples (1.9%) of hot water from faucets exceeded Environmental Protection Agency standards for silver, and none exceeded the standards for copper. Of 24 samples obtained from hot water tanks, 42% and 50% exceeded the silver and copper standards, respectively. Copper-silver ionization effectively controls L. pneumophila in high-volume plumbing systems and is superior to thermal treatment; however, high concentrations of copper and silver can accumulate at the bottom of hot water tanks.

  6. Effect of Legionella pneumophila sonicate on killing of Listeria monocytogenes by human polymorphonuclear neutrophils and monocytes

    DEFF Research Database (Denmark)

    Rechnitzer, C; Bangsborg, Jette Marie; Shand, G H

    1993-01-01

    will be exposed to bacterial components, either expressed on the surface of the organisms or released in the environment upon cell lysis. In this study, we have investigated the effect of water-soluble bacterial components present in L. pneumophila sonicate on the phagocytosis and bactericidal activity of human...... polymorphonuclear neutrophils and monocytes. Preincubation of neutrophils with L. pneumophila sonicate did not affect phagocytosis of L. monocytogenes, whereas Listeria killing was significantly inhibited at sonicate concentrations of 1 and 2 mg/ml. The phenol phase of a phenol-water extraction, containing most...... of the lipopolysaccharide (LPS), had no inhibitory effect on the listericidal activity of neutrophils. Killing of Listeria by monocytes was inhibited in a similar manner. The inhibitory activity was mainly recovered in the sonicate fraction above 100 kDa, suggesting that components organized in larger molecular complexes...

  7. Biosynthesis of CMP-N,N'-diacetyllegionaminic acid from UDP-N,N'-diacetylbacillosamine in Legionella pneumophila.

    Science.gov (United States)

    Glaze, Pavel A; Watson, David C; Young, N Martin; Tanner, Martin E

    2008-03-11

    Legionaminic acid is a nine-carbon alpha-keto acid that is similar in structure to other members of the sialic acid family that includes neuraminic acid and pseudaminic acid. It is found as a component of the lipopolysaccharide in several bacterial species and is perhaps best known for its presence in the O-antigen of the causative agent of Legionnaires' disease, Legionella pneumophila. In this work, the enzymes responsible for the biosynthesis and activation of N, N'-diacetyllegionaminic acid are identified for the first time. A cluster of three L. pneumophila genes bearing homology to known sialic acid biosynthetic genes ( neuA,B,C) were cloned and overexpressed in Escherichia coli. The NeuC homologue was found to be a hydrolyzing UDP- N, N'-diacetylbacillosamine 2-epimerase that converts UDP- N, N'-diacetylbacillosamine into 2,4-diacetamido-2,4,6-trideoxymannose and UDP. Stereochemical and isotopic labeling studies showed that the enzyme utilizes a mechanism involving an initial anti elimination of UDP to form a glycal intermediate and a subsequent syn addition of water to generate product. This is similar to the hydrolyzing UDP- N-acetylglucosamine 2-epimerase (NeuC) of sialic acid biosynthesis, but the L. pneumophila enzyme would not accept UDP-GlcNAc as an alternate substrate. The NeuB homologue was found to be a N, N'-diacetyllegionaminic acid synthase that condenses 2,4-diacetamido-2,4,6-trideoxymannose with phosphoenolpyruvate (PEP), although the in vitro activity of the recombinant enzyme (isolated as a MalE fusion protein) was very low. The synthase activity was dependent on the presence of a divalent metal ion, and the reaction proceeded via a C-O bond cleavage process, similar to the reactions catalyzed by the sialic acid and pseudaminic acid synthases. Finally, the NeuA homologue was shown to possess the CMP- N, N'-diacetyllegionaminic acid synthetase activity that generates the activated form of legionaminic acid used in lipopolysaccharide

  8. Detection of Legionella pneumophila in water and biofilm samples by culture and molecular methods from man-made systems in São Paulo - Brazil Detecção de Legionella pneumophila por métodos de cultivo e moleculares em sistemas artificiais de climatização de ambientes interiores em São Paulo - Brasil

    Directory of Open Access Journals (Sweden)

    Fábio R.S. Carvalho

    2007-12-01

    Full Text Available Legionella pneumophila is a pathogenic bacteria associated to aquatic habitat of natural and artificial environments. Clinical cases of legionellosis have been reported in Brazil but there is a lack of information about the incidence and concentration of this bacterium in environmental sources. Thus, the present study was designed to evaluate the occurrence of legionellae in São Paulo city, Brazil, using different methods of detection and identification. Sixty-seven water and biofilm samples from natural reservoirs and man-made systems were collected and analyzed for the presence of Legionella spp by culturing onto a selective medium, coculture in axenic free-living amoebae and direct fluorescent antibody (DFA assay. Results showed that freshwater of reservoirs did not contain legionellae, Legionella pneumophila was isolated from man-made systems, with predominance of Legionella pneumophila serogroup 1 strains. Although there was no statistical difference among the proposed detection methods, the plate culture method yielded a higher number of L. pneumophila positive samples, followed by amoebic coculture procedure and direct fluorescent antibody assay. Results of PCR and sequencing reactions revealed that application of macrophage infectivity potentiator gene as a molecular marker was an important tool for the identification of environmental isolates of L. pneumophila. The agreement among the three detection methods-when all methods yielded similar results- and the prevalence of a single Legionella species in the sampled man-made systems could suggest that the occurrence of this bacterium had been influenced by the higher concentration of metallic ions dissociated in water of those systems than in natural reservoirs. Thus, the results of this study revealed that the water of man-made systems in Sao Paulo may serve as a reservoir for L. pneumophila and other microorganism, including free-living protozoans.Legionella pneumophila é uma bact

  9. A multicenter evaluation of genotypic methods for the epidemiologic typing of Legionella pneumophila serogroup 1

    DEFF Research Database (Denmark)

    Fry, Norman K.; Alexiou-Daniel, Stella; Bangsborg, Jette Marie

    1999-01-01

    length polymorphism analysis, restriction endonuclease analysis, pulsed-field gel electrophoresis (PFGE), PCR using arbitrary/repeat sequence primers (AP-, AP/rep-PCR), and amplified fragment length polymorphism (AFLP) analysis. Results were analyzed visually or using gel analysis software. Each method...... isolates (114) of L. pneumophila sg 1 comprising one epidemiologically 'unrelated' (79) and one 'related' panel of isolates (35) were sent to 12 laboratories in 11 European countries. Analysis was undertaken in each laboratory using one or more of the following methods: ribotyping, restriction fragment...

  10. MALDI-TOF MS analysis as a useful tool for an identification of Legionella pneumophila, a facultatively pathogenic bacterium interacting with free-living amoebae: A case study from water supply system of hospitals in Bratislava (Slovakia).

    Science.gov (United States)

    Trnková, Katarína; Kotrbancová, Martina; Špaleková, Margita; Fulová, Miriam; Boledovičová, Jana; Vesteg, Matej

    2017-12-07

    Legionellae, i.e. Legionella pneumophila, are human bacterial hydrophilic facultative pathogens causing pneumonia (Legionnaires' disease). Free-living amoebae (FLA) can serve as natural hosts and thus as reservoirs of many amoebae-resistant bacteria. An encysted amoeba can contribute to the resistance of intracellular L. pneumophila to various chemical and physical treatments. Humans can be infected by droplets containing bacteria from an environmental source or human-made devices such as shower heads, bathtubs, air-conditioning units or whirlpools. In this study, we were investigating the presence of FLA and L. pneumophila in plumbing systems of healthcare facilities in Bratislava (Slovakia) by standard diagnostic methods, while the presence of L. pneumophila was verified also by MALDI-TOF MS (matrix-assisted laser desorption/ionization time-of-flight mass spectrometry) analysis. The results showed the occurrence of L. pneumophila and FLA in 62.26% and 66.4% of samples taken from four paediatric clinics, respectively. Both standard methods and MALDI-TOF MS showed comparable results and they can be successfully applied for the identification of L. pneumophila strains in environmental samples. Our approach could be useful for further monitoring, prevention and decreasing risk of Legionella infection also in other hospitals. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Prevalence of [i]Legionella pneumophila[/i] in water distribution systems in hospitals and public buildings of the Lublin region of eastern Poland

    Directory of Open Access Journals (Sweden)

    Agnieszka Sikora

    2015-05-01

    Full Text Available Objective. The aim of this study was to determine the prevalence of [i]L. pneumophila[/i] in water supply systems, hospitals and public buildings in the Lublin region of eastern Poland. Material and methods. The study was carried out in 26 different objects in the Lublin region. The number of [i]Legionella[/i] bacteria in water samples was determined by the membrane filtration method and/or by surface inoculation in accordance with the standards. Results. The study showed the presence of[i] L. pneumophila[/i] in 166 hot water samples (74.77%. In 34.33% (n=57 of water samples the count of tested bacteria exceeded the acceptable level of >100 CFU/100 ml. Of the samples where an acceptable level of bacteria was exceeded, 49 samples had an average level of [i]L. pneumophila[/i] (100–1,000 CFU/100 ml, and the level in 8 samples was high (>1,000 CFU/100 ml. Conclusions. The water samples collected form the hot water supply system of hospitals and public buildings showed exceeded counts of[i] L. pneumophila[/i], indicating the risk of infection. The constant monitoring of water distribution systems is an important element of the control of infections caused by these organisms.

  12. Three Antagonistic Cyclic di-GMP-Catabolizing Enzymes Promote Differential Dot/Icm Effector Delivery and Intracellular Survival at the Early Steps of Legionella pneumophila Infection

    Science.gov (United States)

    Allombert, Julie; Lazzaroni, Jean-Claude; Baïlo, Nathalie; Gilbert, Christophe; Charpentier, Xavier; Doublet, Patricia

    2014-01-01

    Legionella pneumophila is an intracellular pathogen which replicates within protozoan cells and can accidently infect alveolar macrophages, causing an acute pneumonia in humans. The second messenger cyclic di-GMP (c-di-GMP) has been shown to play key roles in the regulation of various bacterial processes, including virulence. While investigating the function of the 22 potential c-di-GMP-metabolizing enzymes of the L. pneumophila Lens strain, we found three that directly contribute to its ability to infect both protozoan and mammalian cells. These three enzymes display diguanylate cyclase (Lpl0780), phosphodiesterase (Lpl1118), and bifunctional diguanylate cyclase/phosphodiesterase (Lpl0922) activities, which are all required for the survival and intracellular replication of L. pneumophila. Mutants with deletions of the corresponding genes are efficiently taken up by phagocytic cells but are partially defective for the escape of the Legionella-containing vacuole (LCV) from the host degradative endocytic pathway and result in lower survival. In addition, Lpl1118 is required for efficient endoplasmic reticulum recruitment to the LCV. Trafficking and biogenesis of the LCV are dependent upon the orchestrated actions of several type 4 secretion system Dot/Icm effectors proteins, which exhibit differentially altered translocation in the three mutants. While translocation of some effectors remained unchanged, others appeared over- and undertranslocated. A general translocation offset of the large repertoire of Dot/Icm effectors may be responsible for the observed defects in the trafficking and biogenesis of the LCV. Our results suggest that L. pneumophila uses cyclic di-GMP signaling to fine-tune effector delivery and ensure effective evasion of the host degradative pathways and establishment of a replicative vacuole. PMID:24379287

  13. Biochemical and Functional Analyses of the Mip Protein: Influence of the N-Terminal Half and of Peptidylprolyl Isomerase Activity on the Virulence of Legionella pneumophila

    OpenAIRE

    Köhler, Rolf; Fanghänel, Jörg; König, Bettina; Lüneberg, Edeltraud; Frosch, Matthias; Rahfeld, Jens-Ulrich; Hilgenfeld, Rolf; Fischer, Gunter; Hacker, Jörg; Steinert, Michael

    2003-01-01

    The virulence factor Mip (macrophage infectivity potentiator) contributes to the intracellular survival of Legionella pneumophila, the causative agent of Legionnaires' disease. The protein consists of two domains that are connected via a very long α-helix (A. Riboldi-Tunnicliffe et al., Nat. Struct. Biol. 8:779-783, 2001). The fold of the C-terminal domain (residues 100 to 213) is closely related to human FK506-binding protein (FKBP12), and like FKBP12, Mip exhibits peptidylprolyl cis/trans i...

  14. Legionella pneumophila infection presenting as headache, confusion and dysarthria in a human immunodeficiency virus-1 (HIV-1 positive patient: case report

    Directory of Open Access Journals (Sweden)

    Robbins Nathaniel M

    2012-09-01

    Full Text Available Abstract Background Legionella pneumophila is a common cause of community-acquired pneumonia. Central nervous system dysfunction is common, and diagnosis in the absence of pulmonary symptoms can be challenging. Here we describe an atypical clinical presentation of Legionella infection in a patient with HIV who was found to have an unusual neuroradiologic lesion that further served to obscure the diagnosis. This is the first such description in a patient with Legionellosis and HIV coinfection. Case presentation A 43 year-old HIV positive man presented to our hospital with dysarthria, fevers, headache, and altered mental status. Initial work-up revealed pneumonia and a lesion of the splenium of the corpus callosum on magnetic resonance imaging. He was subsequently diagnosed with Legionella pneumonia and treated with complete symptom resolution. Conclusions Neurologic abnormalities are frequent in Legionellosis, but the diagnosis may be difficult in the absence of overt respiratory symptoms and in the presence of HIV coinfection. A high index of suspicion and early initiation of empiric antibiotics is imperative since early treatment may help prevent long-term sequelae. Neuroimaging abnormalities, though rare, can help the physician narrow down the diagnosis and avoid unnecessary invasive testing. Future studies should aim to elucidate the as yet unknown role of neuroimaging in the diagnoses and prognostication of Legionellosis, as well as the interaction between Legionella infection and HIV.

  15. Designation of the European Working Group on Legionella Infection (EWGLI) amplified fragment length polymorphism types of Legionella pneumophila serogroup 1 and results of intercentre proficiency testing Using a standard protocol

    DEFF Research Database (Denmark)

    Fry, N K; Bangsborg, Jette Marie; Bergmans, A

    2002-01-01

    (recorded as AFLP type 001-016 or untypeable) was determined by participants with reference to these 16 AFLP types, either visually or using gel analysis software where available, and reported to the coordinating centre. Nine of the 12 strains, including an epidemiologically related pair and two pairs......The utility of amplified fragment length polymorphism (AFLP) analysis as a genotyping method for the epidemiological typing of Legionella pneumophila serogroup 1 has been previously demonstrated. This study (i). reports recommendations for the designation of the European Working Group on Legionella...... Infections (EWGLI) AFLP types, (ii). describes the EWGLI AFLP types identified for the 130 strains in the EWGLI culture collection, and (iii). reports the results of a newly introduced international programme of proficiency testing. Following preliminary analysis of 20 epidemiologically unrelated isolates...

  16. Comparison of sputum and nasopharyngeal swab specimens for molecular diagnosis of Mycoplasma pneumoniae, Chlamydophila pneumoniae, and Legionella pneumophila.

    Science.gov (United States)

    Cho, Min-Chul; Kim, Hyewon; An, Dongheui; Lee, Miyoung; Noh, Shin-Ae; Kim, Mi-Na; Chong, Young Pil; Woo, Jun Hee

    2012-03-01

    Differentiation of atypical pathogens is important for community-acquired pneumonia (CAP). In this study, we compared sputum and nasopharyngeal swabs (NPS) for use in detection of Mycoplasma pneumoniae (MP), Chlamydophila pneumoniae (CP), and Legionella pneumophila (LP), using Seeplex PneumoBacter ACE Detection Assay (PneumoBacter; Seegene). Sputum and NPS specimens were collected from patients in 15 hospitals. DNA was extracted from sputum using QIAamp DNA Stool Mini Kit (Qiagen) and from NPS using easyMAG (bioMérieux). Both types of specimens were evaluated by multiplex PCR using PneumoBacter. To determine the diagnostic performance of this assay, sputum samples were also tested using BD ProbeTec ET Atypical Pneumonia Assay (APA; Becton Dickinson). Among 217 sputum and NPS, 20 (9.2%), 2 (0.9%), and 0 sputum were positive for MP, LP, and CP, respectively, whereas 8 (3.7%) NPS were positive for MP. The sputum APA test yielded 186, 206, and 204 interpretable results for MP, LP, and CP, respectively. Of these, 21 (11.3%) were positive for MP, 2 (1.0%) were positive for LP, and 0 samples were positive for CP. Compared to APA, the sensitivity and specificity of the sputum assay for MP were 95.2% and 100.0%, respectively, whereas for the NPS assay, these were 38.1% and 93.9%. Sputum testing was more sensitive than NPS testing (P=0.002). For LP and CP diagnosis, PneumoBacter and APA tests agreed 100%. Specimen type is crucial and sputum is preferred over NPS for simultaneous detection of MP, LP, and CP using multiplex PCR in CAP.

  17. A Unique cis-Encoded Small Noncoding RNA Is Regulating Legionella pneumophila Hfq Expression in a Life Cycle-Dependent Manner

    Directory of Open Access Journals (Sweden)

    Giulia Oliva

    2017-01-01

    Full Text Available Legionella pneumophila is an environmental bacterium that parasitizes protozoa, but it may also infect humans, thereby causing a severe pneumonia called Legionnaires’ disease. To cycle between the environment and a eukaryotic host, L. pneumophila is regulating the expression of virulence factors in a life cycle-dependent manner: replicating bacteria do not express virulence factors, whereas transmissive bacteria are highly motile and infective. Here we show that Hfq is an important regulator in this network. Hfq is highly expressed in transmissive bacteria but is expressed at very low levels in replicating bacteria. A L. pneumophila hfq deletion mutant exhibits reduced abilities to infect and multiply in Acanthamoeba castellanii at environmental temperatures. The life cycle-dependent regulation of Hfq expression depends on a unique cis-encoded small RNA named Anti-hfq that is transcribed antisense of the hfq transcript and overlaps its 5′ untranslated region. The Anti-hfq sRNA is highly expressed only in replicating L. pneumophila where it regulates hfq expression through binding to the complementary regions of the hfq transcripts. This results in reduced Hfq protein levels in exponentially growing cells. Both the small noncoding RNA (sRNA and hfq mRNA are bound and stabilized by the Hfq protein, likely leading to the cleavage of the RNA duplex by the endoribonuclease RNase III. In contrast, after the switch to transmissive bacteria, the sRNA is not expressed, allowing now an efficient expression of the hfq gene and consequently Hfq. Our results place Hfq and its newly identified sRNA anti-hfq in the center of the regulatory network governing L. pneumophila differentiation from nonvirulent to virulent bacteria.

  18. [Legionella pneumophila eukaryotic-like effector LegK3 inhibits growth of Saccharomyces cerevisiae and modulates its vesicle trafficking pathway].

    Science.gov (United States)

    Wang, Jiaming; Li, Xianghui; Chen, Aifeng; Lu, Yongjun

    2014-04-04

    To study biochemical functions of the Legionella pneumophila eukaryotic-like effector protein LegK3, the budding yeast Saccharomyces cerevisiae was used as an alternative host in which growth defect induced by the ectopic expression of LegK3 was assessed. Using genomic DNA of the L. pneumophila strain Lp02 as template, we respectively amplified and inserted the ORF sequences of legK3, ralF or lidA into the plasmid pESC-HK to yield the ectopic-expression plasmids. Then, the recombination plasmids were transformed into the yeast strain W301-1A. With 2% -galactose induction, growth defect and carboxypeptidase Y (CPY) delay were determined simultaneously. In parallel, total yeast proteins before or after induction were extracted and subjected to Immunoblot assay. For detecting the expression of effector proteins or determining CPY delay, anti-c-myc or anti-PGK/anti-CPY antibodies were utilized respectively. The expression of LegK3 resulted in visible growth defect in yeast cells, together with obvious retard in CPY processing. L. pneumophila eukaryotic-like effector LegK3 might target and interfere with the vesicle-trafficking pathways, thereby to inhibit the growth and division of host cells.

  19. Characterization of the icmH and icmF genes required for Legionella pneumophila intracellular growth, genes that are present in many bacteria associated with eukaryotic cells.

    Science.gov (United States)

    Zusman, Tal; Feldman, Michal; Halperin, Einat; Segal, Gil

    2004-06-01

    Legionella pneumophila, the causative agent of Legionnaires' disease, replicates intracellularly within a specialized phagosome of mammalian and protozoan host cells, and the Icm/Dot type IV secretion system has been shown to be essential for this process. Unlike all the other known Icm/Dot proteins, the IcmF protein, which was described before, and the IcmH protein, which is characterized here, have homologous proteins in many bacteria (such as Yersinia pestis, Salmonella enterica, Rhizobium leguminosarum, and Vibrio cholerae), all of which associate with eukaryotic cells. Here, we have characterized the L. pneumophila icmH and icmF genes and found that both genes are present in 16 different Legionella species examined. The icmH and icmF genes were found to be absolutely required for intracellular multiplication in Acanthamoeba castellanii and partially required for intracellular growth in HL-60-derived human macrophages, for immediate cytotoxicity, and for salt sensitivity. Mutagenesis of the predicted ATP/GTP binding site of IcmF revealed that the site is partially required for intracellular growth in A. castellanii. Analysis of the regulatory region of the icmH and icmF genes, which were found to be cotranscribed, revealed that it contains at least two regulatory elements. In addition, an icmH::lacZ fusion was shown to be activated during stationary phase in a LetA- and RelA-dependent manner. Our results indicate that although the icmH and icmF genes probably have a different evolutionary origin than the rest of the icm/dot genes, they are part of the icm/dot system and are required for L. pneumophila pathogenesis.

  20. Influence of Sampling Season and Sampling Protocol on Detection of Legionella Pneumophila Contamination in Hot Water / Paraugu Ņemšanas Sezonalitātes Un Paraugu Ņemšanas Metodes Ietekme Uz Legionella Pneumophila Kontaminācijas Noteikšanu Karstajš Ūdenī

    Directory of Open Access Journals (Sweden)

    Pūle Daina

    2016-08-01

    Full Text Available Legionella pneumophila is an environmental pathogen of engineered water systems that can cause different forms of legionellosis - from mild fever to potentially lethal pneumonia. Low concentrations of legionellae in natural habitats can increase markedly in engineered hot water systems where water temperatures are below 55 °C. In the current study, we aimed to investigate the influence of sampling season, hot water temperature and sampling protocol on occurrence of L. pneumophila. A total of 120 hot water samples from 20 apartment buildings were collected in two sampling periods - winter 2014 (n = 60 and summer 2015 (n = 60. Significantly higher occurrence of L. pneumophila was observed in summer 2015. Significant differences in temperature for negative and positive samples were not observed, which can be explained by low water temperatures at the point of water consumption. Temperature above 55 °C was observed only once, for all other sampling events it ranged from 14 °C to 53 °C.

  1. Gezondheidsaspecten van Legionella in water

    NARCIS (Netherlands)

    Schets FM; Roda Husman AM de; MGB

    2004-01-01

    Legionella pneumophila is a cause of severe pneumonia in humans. Legionella bacteria are found in all freshwater environments. Particularly in artificial environments people may be exposed to Legionella containing aerosols. In this brief overview of the state of the art of Legionella research it

  2. Planktonic replication is essential for biofilm formation by Legionella pneumophila in a complex medium under static and dynamic flow conditions

    DEFF Research Database (Denmark)

    Mampel, J.; Spirig, T.; Weber, S.S.

    2006-01-01

    within biofilms in the absence of protozoa. In this study, we analyzed surface adherence of and biofilm formation by L. pneumophila in a rich medium that supported axenic replication. Biofilm formation by the virulent L. pneumophila strain JR32 and by clinical and environmental isolates was analyzed...... formed biofilms in the inverse system if the medium was exchanged twice a day. However, after addition of Acanthamoeba castellanii, the bacteria proliferated and adhered to surfaces. Sessile (surface-attached) and planktonic (free-swimming) L. pneumophila expressed beta-galactosidase activity to similar......, and no sizeable patches of clonally growing bacteria were observed. Our findings indicate that biofilm formation by L. pneumophila in a rich medium is due to growth of planktonic bacteria rather than to growth of sessile bacteria. In agreement with this conclusion, GFP-labeled L. pneumophila initially adhered...

  3. Betekenis van Legionella-soorten voor preventiebeleid van leidingwaterinstallaties

    NARCIS (Netherlands)

    Versteegh JFM; Brandsema PS; Lodder WJ; de Roda Husman AM; Schalk JAC; van der Aa NGFM; IMG; LZO; EPI

    2009-01-01

    Het RIVM adviseert om de huidige normstelling voor het preventiebeleid van Legionella te handhaven en niet uitsluitend op Legionella pneumophila te richten. Als andere Legionella-soorten worden aangetroffen kan er ook groei van Legionella pneumophila optreden. Als er dan geen maatregelen worden

  4. Genomic investigation of a suspected outbreak of Legionella pneumophila ST82 reveals undetected heterogeneity by the present gold-standard methods, Denmark, July to November 2014.

    Science.gov (United States)

    Schjørring, Susanne; Stegger, Marc; Kjelsø, Charlotte; Lilje, Berit; Bangsborg, Jette M; Petersen, Randi F; David, Sophia; Uldum, Søren A

    2017-06-22

    Between July and November 2014, 15 community-acquired cases of Legionnaires´ disease (LD), including four with Legionella pneumophila serogroup 1 sequence type (ST) 82, were diagnosed in Northern Zealand, Denmark. An outbreak was suspected. No ST82 isolates were found in environmental samples and no external source was established. Four putative-outbreak ST82 isolates were retrospectively subjected to whole genome sequencing (WGS) followed by phylogenetic analyses with epidemiologically unrelated ST82 sequences. The four putative-outbreak ST82 sequences fell into two clades, the two clades were separated by ca 1,700 single nt polymorphisms (SNP)s when recombination regions were included but only by 12 to 21 SNPs when these were removed. A single putative-outbreak ST82 isolate sequence segregated in the first clade. The other three clustered in the second clade, where all included sequences had Denmark dating back as early as 2011. The study confirms that recombination plays a major role in L. pneumophila evolution. On the other hand, strains belonging to the same ST can have only few SNP differences despite being sampled over both large timespans and geographic distances. These are two important factors to consider in outbreak investigations. This article is copyright of The Authors, 2017.

  5. Fusion of Legionella pneumophila outer membrane vesicles with eukaryotic membrane systems is a mechanism to deliver pathogen factors to host cell membranes.

    Science.gov (United States)

    Jäger, Jens; Keese, Susanne; Roessle, Manfred; Steinert, Michael; Schromm, Andra B

    2015-05-01

    The formation and release of outer membrane vesicles (OMVs) is a phenomenon observed in many bacteria, including Legionella pneumophila. During infection, this human pathogen primarily invades alveolar macrophages and replicates within a unique membrane-bound compartment termed Legionella-containing vacuole. In the current study, we analysed the membrane architecture of L. pneumophila OMVs by small-angle X-ray scattering and biophysically characterized OMV membranes. We investigated the interaction of L. pneumophila OMVs with model membranes by Förster resonance energy transfer and Fourier transform infrared spectroscopy. These experiments demonstrated the incorporation of OMV membrane material into liposomes composed of different eukaryotic phospholipids, revealing an endogenous property of OMVs to fuse with eukaryotic membranes. Cellular co-incubation experiments showed a dose- and time-dependent binding of fluorophore-labelled OMVs to macrophages. Trypan blue quenching experiments disclosed a rapid internalization of OMVs into macrophages at 37 and 4 °C. Purified OMVs induced tumour necrosis factor-α production in human macrophages at concentrations starting at 300 ng ml(-1). Experiments on HEK293-TLR2 and TLR4/MD-2 cell lines demonstrated a dominance of TLR2-dependent signalling pathways. In summary, we demonstrate binding, internalization and biological activity of L. pneumophila OMVs on human macrophages. Our data support OMV membrane fusion as a mechanism for the remote delivery of virulence factors to host cells. © 2014 John Wiley & Sons Ltd.

  6. Negative effect of high pH on biocidal efficacy of copper and silver ions in controlling Legionella pneumophila.

    Science.gov (United States)

    Lin, Yu-Sen E; Vidic, Radisav D; Stout, Janet E; Yu, Victor L

    2002-06-01

    Copper-silver (Cu-Ag) ionization has effectively controlled Legionella spp. in the hot water systems of numerous hospitals. However, it was ineffective at controlling Legionella in one Ohio hospital despite the confirmation of adequate total concentrations of copper and silver ions. The pH of the water at this hospital was found to be 8.5 to 9.0. The purpose of this study was to investigate the impact of pH and other water quality parameters, including alkalinity (HCO3-), hardness (Ca2+ and Mg2+), and amount of dissolved organic carbon (DOC), on the control of Legionella by Cu-Ag ionization. Initial concentrations of Legionella and copper and silver ions used in batch experiments were 3 x 10(6) CFU/ml and 0.4 and 0.08 mg/liter, respectively. Changes in bicarbonate ion concentration (50, 100, and 150 mg/liter), water hardness (Ca2+ at 50 and 100 mg/liter; Mg2+ at 40 and 80 mg/liter), and level of DOC (0.5 and 2 mg/liter) had no significant impact on the efficacy of copper and silver ions in killing Legionella at a neutral pH. When the pH was elevated to 9 in these experiments, copper ions achieved only a 10-fold reduction in the number of Legionella organisms in 24 h, compared to a millionfold decrease at pH 7.0. Silver ions were able to achieve a millionfold reduction in 24 h at all ranges of water quality parameters tested. Precipitation of insoluble copper complexes was observed at a pH above 6.0. These results suggest that pH may be an important factor in the efficacy of copper-silver ionization in controlling Legionella in water systems.

  7. Differential growth of Legionella pneumophila strains within a range of amoebae at various temperatures associated with in-premise plumbing

    Science.gov (United States)

    The potential effect of in-premise plumbing temperatures (24, 32, 37 and 41 °C) on the growth of five different L. pneumophila strains within free-living amoebae (Acanthamoeba polyphaga, Hartmannella vermiformis and Naegleria fowleri) was examined. Compared to controls only fed E...

  8. Prevalence of Legionella pneumophila in water distribution systems in hospitals and public buildings of the Lublin region of eastern Poland

    Directory of Open Access Journals (Sweden)

    Agnieszka Sikora

    2015-05-01

    The water samples collected form the hot water supply system of hospitals and public buildings showed exceeded counts of L. pneumophila, indicating the risk of infection. The constant monitoring of water distribution systems is an important element of the control of infections caused by these organisms.

  9. Untersuchung zur Besiedlung der Warmwassersysteme von Ein- und Zweifamilienhäusern mit Legionellen

    OpenAIRE

    Stanke, J. (Juliane)

    2005-01-01

    In dieser Arbeit wurden 198 Warmwassersysteme von Ein- und Zweifamilienhäusern auf Legionellen untersucht. Zusätzlich wurden die Proben auf Kupfer und Zink analysiert. Die Untersuchungen ergaben bei zehn Proben (5% aller Proben) positive Nachweise von Legionella pneumophila. Es zeigte sich ein deutlicher Zusammenhang zwischen der Temperaturführung des Warmwassers und der Anzahl koloniebildender Einheiten von Legionellen.So wurden Legionellen ausschließlich aus Wasserproben mit einer Temperatu...

  10. Negative Effect of High pH on Biocidal Efficacy of Copper and Silver Ions in Controlling Legionella pneumophila

    OpenAIRE

    Lin, Yu-sen E.; Vidic, Radisav D.; Stout, Janet E.; Yu, Victor L.

    2002-01-01

    Copper-silver (Cu-Ag) ionization has effectively controlled Legionella spp. in the hot water systems of numerous hospitals. However, it was ineffective at controlling Legionella in one Ohio hospital despite the confirmation of adequate total concentrations of copper and silver ions. The pH of the water at this hospital was found to be 8.5 to 9.0. The purpose of this study was to investigate the impact of pH and other water quality parameters, including alkalinity (HCO3−), hardness (Ca2+ and M...

  11. Identification of Legionella pneumophila effectors regulated by the LetAS-RsmYZ-CsrA regulatory cascade, many of which modulate vesicular trafficking.

    Science.gov (United States)

    Nevo, Oded; Zusman, Tal; Rasis, Michal; Lifshitz, Ziv; Segal, Gil

    2014-02-01

    Legionella pneumophila, the causative agent of Legionnaires' disease, is an intracellular human pathogen that utilizes the Icm/Dot type IVB secretion system to translocate a large repertoire of effectors into host cells. To find coregulated effectors, we performed a bioinformatic genomic screen with the aim of identifying effector-encoding genes containing putative CsrA regulatory elements. The regulation of these genes by the LetAS-RsmYZ-CsrA regulatory cascade was experimentally validated by examining their levels of expression in deletion mutants of relevant regulators and by site-directed mutagenesis of the putative CsrA sites. These analyses resulted in the identification of 26 effector-encoding genes regulated by the LetAS-RsmYZ-CsrA regulatory cascade, all of which were expressed at higher levels during the stationary phase. To determine if any of these effectors is involved in modulating the secretory pathway, they were overexpressed in wild-type yeast as well as in a yeast sec22 deletion mutant, which encodes an R-SNARE that participates in the endoplasmic reticulum (ER)-Golgi trafficking. This examination identified many novel LetAS-RsmYZ-CsrA regulated effectors which are involved in this process. To further characterize the role of these 26 effectors in vesicular trafficking, they were examined in yeast arf and arl deletion mutants, which encode small GTPases that regulate ER-Golgi trafficking. This analysis revealed that the effectors examined manipulate different processes of the secretory pathway. Collectively, our results demonstrate that several of the L. pneumophila effectors which are coregulated in the bacterial cell are involved in the modulation of the same eukaryotic pathway.

  12. Aerosolization of respirable droplets from a domestic spa pool and the use of MS-2 coliphage and Pseudomonas aeruginosa as markers for Legionella pneumophila.

    Science.gov (United States)

    Moore, Ginny; Hewitt, Matthew; Stevenson, David; Walker, Jimmy T; Bennett, Allan M

    2015-01-01

    Legionnaires' disease can result when droplets or aerosols containing legionella bacteria are inhaled and deposited in the lungs. A number of outbreaks have been associated with the use of a spa pool where aeration, a high water temperature, and a large and variable organic load make disinfectant levels difficult to maintain. Spa pool ownership is increasing, and the aim of this study, using two surrogate organisms (MS-2 coliphage and Pseudomonas aeruginosa [a natural contaminant]), was to assess the potential risk to domestic users when disinfection fails. A representative "entry level" domestic spa pool was installed in an outdoor courtyard. The manufacturer's instructions for spa pool maintenance were not followed. A cyclone sampler was used to sample the aerosols released from the spa pool with and without activation of the air injection system. Samples were taken at increasing heights and distances from the pool. An aerodynamic particle sizer was used to measure the water droplet size distribution at each sample point. When the air injection system was inactivated, neither surrogate organism was recovered from the air. On activation of the air injection system, the mean mass of droplets within the respirable range (10 cm above the water line) was 36.8 μg cm(-3). This corresponded to a mean air concentration of P. aeruginosa of 350 CFU m(-3). From extrapolation from animal data, the estimated risk of infection from aerosols contaminated with similar concentrations of Legionella pneumophila was 0.76 (males) and 0.65 (females). At 1 m above and/or beyond the pool, the mean aerosol mass decreased to 0.04 μg cm(-3) and corresponded to a 100-fold reduction in mean microbial air concentration. The estimated risk of infection at this distance was negligible. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  13. Legionella pneumophila Type IV Effectors YlfA and YlfB Are SNARE-Like Proteins that Form Homo- and Heteromeric Complexes and Enhance the Efficiency of Vacuole Remodeling.

    Science.gov (United States)

    Campodonico, Eva M; Roy, Craig R; Ninio, Shira

    2016-01-01

    Legionella pneumophila is a Gram-negative bacterium that can colonize both freshwater protozoa and human alveolar macrophages, the latter infection resulting in Legionnaires' disease. The intracellular lifecycle of L. pneumophila requires extensive manipulation of its host cell, which is carried out by effector proteins that are translocated into the host cell through the Dot/Icm type IV secretion system. This study focuses on a pair of highly similar type IV substrates called YlfA/LegC7 and YlfB/LegC2 that were initially identified in a screen for proteins that cause growth inhibition in yeast. Analysis of truncation mutants revealed that the hydrophobic residues in the Ylf amino termini were required for localization of each protein to the membranes of host cells. Central and carboxy terminal coiled coil domains were found to mediate binding of YlfA and YlfB to themselves and to each other. In vivo, a ΔylfA ΔylfB double mutant strain of L. pneumophila was shown to be defective in establishing a vacuole that supports bacterial replication. This phenotype was subsequently correlated with a decrease in the association of endoplasmic reticulum (ER)-derived vesicles with vacuoles containing ΔylfA ΔylfB mutant bacteria. These data suggest that the Ylf proteins are membrane-associated effectors that enhance remodeling of the L. pneumophila -containing vacuole by promoting association and possibly fusion of ER-derived membrane vesicles with the bacterial compartment.

  14. Legionella pneumophila Type IV Effectors YlfA and YlfB Are SNARE-Like Proteins that Form Homo- and Heteromeric Complexes and Enhance the Efficiency of Vacuole Remodeling.

    Directory of Open Access Journals (Sweden)

    Eva M Campodonico

    Full Text Available Legionella pneumophila is a Gram-negative bacterium that can colonize both freshwater protozoa and human alveolar macrophages, the latter infection resulting in Legionnaires' disease. The intracellular lifecycle of L. pneumophila requires extensive manipulation of its host cell, which is carried out by effector proteins that are translocated into the host cell through the Dot/Icm type IV secretion system. This study focuses on a pair of highly similar type IV substrates called YlfA/LegC7 and YlfB/LegC2 that were initially identified in a screen for proteins that cause growth inhibition in yeast. Analysis of truncation mutants revealed that the hydrophobic residues in the Ylf amino termini were required for localization of each protein to the membranes of host cells. Central and carboxy terminal coiled coil domains were found to mediate binding of YlfA and YlfB to themselves and to each other. In vivo, a ΔylfA ΔylfB double mutant strain of L. pneumophila was shown to be defective in establishing a vacuole that supports bacterial replication. This phenotype was subsequently correlated with a decrease in the association of endoplasmic reticulum (ER-derived vesicles with vacuoles containing ΔylfA ΔylfB mutant bacteria. These data suggest that the Ylf proteins are membrane-associated effectors that enhance remodeling of the L. pneumophila -containing vacuole by promoting association and possibly fusion of ER-derived membrane vesicles with the bacterial compartment.

  15. DsbA2 (27-kDa Com1-Like Protein) of Legionella pneumophila Catalyses Extracytoplasmic Disulfide-Bond Formation in Proteins Including the Dot/Icm Type IV Secretion System

    OpenAIRE

    Jameson-Lee, Max; Rafael A Garduno; Hoffman, Paul S.

    2011-01-01

    In Gram negative bacteria, thiol oxidoreductases catalyze the formation of disulfide bonds (DSB) in extracytoplasmic proteins. In this study, we sought to identify DSB-forming proteins required for assembly of macromolecular structures in Legionella pneumophila. Here we describe two DSB forming proteins, one annotated as dsbA1 and the other annotated as a 27-kDa outer membrane protein similar to Com1 of Coxiella burnetii, which we designate as dsbA2. Both proteins are predicted to be periplas...

  16. Indispensable role for the eukaryotic-like ankyrin domains of the ankyrin B effector of Legionella pneumophila within macrophages and amoebae.

    Science.gov (United States)

    Price, Christopher T D; Al-Khodor, Souhaila; Al-Quadan, Tasneem; Abu Kwaik, Yousef

    2010-05-01

    The Dot/Icm-translocated ankyrin B (AnkB) effector of Legionella pneumophila exhibits molecular mimicry of eukaryotic F-box proteins and is essential for intracellular replication in macrophages and protozoa. In addition to two eukaryotic-like ankyrin (ANK) domains, AnkB harbors a conserved eukaryotic F-box domain, which is involved in polyubiquitination of proteins throughout the eukaryotic kingdom. We have recently shown that the F-box domain of the AnkB effector is essential for decoration of the Legionella-containing vacuole (LCV) with polyubiquitinated proteins within macrophages and protozoan hosts. To decipher the role of the two ANK domains in the function of AnkB, we have constructed in-frame deletion of either or both of the ANK domain-encoding regions (ankB Delta A1, ankB Delta A2, and ankB Delta A1A2) to trans-complement the ankB null mutant. Deletion of the ANK domains results in defects in intracellular proliferation and decoration of the LCV with polyubiquitinated proteins. Export of the truncated variants of AnkB was reduced, and this may account for the observed defects. However, while full-length AnkB ectopically expressed in mammalian cells trans-rescues the ankB null mutant for intracellular proliferation, ectopic expression of AnkB Delta A1, AnkB Delta A2, and AnkB Delta A1A2 fails to trans-rescue the ankB null mutant. Importantly, ectopically expressed full-length AnkB is targeted to the host cell plasma membrane, where it recruits polyubiquitinated proteins. In contrast, AnkB Delta A1, AnkB Delta A2, and AnkB Delta A1A2 are diffusely distributed throughout the cytosol and fail to recruit polyubiquitinated proteins. We conclude that the two eukaryotic-like ANK domains of AnkB are essential for intracellular proliferation, for targeting AnkB to the host membranes, and for decoration of the LCV with polyubiquitinated proteins.

  17. Life Stage-specific Proteomes of Legionella pneumophila Reveal a Highly Differential Abundance of Virulence-associated Dot/Icm effectors*

    Science.gov (United States)

    Aurass, Philipp; Gerlach, Thomas; Becher, Dörte; Voigt, Birgit; Karste, Susanne; Bernhardt, Jörg; Riedel, Katharina; Hecker, Michael; Flieger, Antje

    2016-01-01

    Major differences in the transcriptional program underlying the phenotypic switch between exponential and post-exponential growth of Legionella pneumophila were formerly described characterizing important alterations in infection capacity. Additionally, a third state is known where the bacteria transform in a viable but nonculturable state under stress, such as starvation. We here describe phase-related proteomic changes in exponential phase (E), postexponential phase (PE) bacteria, and unculturable microcosms (UNC) containing viable but nonculturable state cells, and identify phase-specific proteins. We present data on different bacterial subproteomes of E and PE, such as soluble whole cell proteins, outer membrane-associated proteins, and extracellular proteins. In total, 1368 different proteins were identified, 922 were quantified and 397 showed differential abundance in E/PE. The quantified subproteomes of soluble whole cell proteins, outer membrane-associated proteins, and extracellular proteins; 841, 55, and 77 proteins, respectively, were visualized in Voronoi treemaps. 95 proteins were quantified exclusively in E, such as cell division proteins MreC, FtsN, FtsA, and ZipA; 33 exclusively in PE, such as motility-related proteins of flagellum biogenesis FlgE, FlgK, and FliA; and 9 exclusively in unculturable microcosms soluble whole cell proteins, such as hypothetical, as well as transport/binding-, and metabolism-related proteins. A high frequency of differentially abundant or phase-exclusive proteins was observed among the 91 quantified effectors of the major virulence-associated protein secretion system Dot/Icm (> 60%). 24 were E-exclusive, such as LepA/B, YlfA, MavG, Lpg2271, and 13 were PE-exclusive, such as RalF, VipD, Lem10. The growth phase-related specific abundance of a subset of Dot/Icm virulence effectors was confirmed by means of Western blotting. We therefore conclude that many effectors are predominantly abundant at either E or PE which suggests

  18. [Use of UV rays for the disinfection of water. III. UV sensitivity of Legionella pneumophila of different ages in cold and warm drinking water].

    Science.gov (United States)

    Martiny, H; Seidel, K; Rüden, H

    1989-05-01

    In drinking water the sensitivity of L. pneumophila serogroup 1 type Philadelphia and L. pneumophila serogroup 5 type Dallas were studied with a flowthrough u.v. light treatment apparatus. By washing purified cells from broth cultures were used as inoculum directly ( = young cultures) or kept 2-3 weeks in drinking water in the dark ( = old cultures). A decrease of 99,9999% was found after u.v. treatment by 19 mWs/cm2 for young cultures and by 15 mWs/cm2 for old cultures. Reductions of 99,9999% were obtained by 16 mWs/cm2 in cold drinking water (13-16 degrees C) and by 13 mWs/cm2 in warm drinking water (45-47 degrees C). L. pneumophila serogroup 1 and L. pneumophila serogroup 5 show a very similar susceptibility to u.v.-irradiation. Reductions of 99,9999% were obtained by 14 mWs/cm2 and 15 mWs/cm2, respectively. Thus L. pneumophila-suspensions proved to be more sensitive to u.v.-irradiation than E. coli oder E. faecium in earlier experiments.

  19. Monochloramine and chlorine dioxide for controlling Legionella pneumophila contamination: biocide levels and disinfection by-product formation in hospital water networks.

    Science.gov (United States)

    Marchesi, Isabella; Ferranti, Greta; Bargellini, Annalisa; Marchegiano, Patrizia; Predieri, Guerrino; Stout, Janet E; Borella, Paola

    2013-12-01

    Legionella colonization in hospital hot water distribution networks was evaluated following 36 months of continuous treatment with monochloramine and compared with chlorine dioxide. Nitrite, nitrate, chlorite, chlorate, bromide, trihalomethanes and haloacetic acids as well as the biocide concentration at sampled points were measured. Only 8/84 samples treated with monochloramine were found contaminated and after the first 8 months of treatment no Legionella was isolated. Chlorine dioxide was associated with a strong reduction in Legionella contamination compared to pre-treatment, but differences according to the device were observed. Monochloramine between 2 and 3 mg l(-1) and chlorine dioxide between 0.50 and 0.70 mg l(-1) were needed to control Legionella colonization. Comparing no- and post-flush samples, a higher frequency of no-flush positive samples was noted using chlorine dioxide, suggesting an increased risk for patients when they open the tap. No increase in chlorite levels and no water nitrification occurred by using monochloramine. Chlorite at levels exceeding the limit requested for drinking water was measured when chlorine dioxide was applied. In conclusion, we highlight that continuous injection of monochloramine should be considered as an effective alternative to chlorine dioxide in controlling legionellae contamination inside hospital water distribution systems.

  20. Legionella infections in cyclosporine-immunosuppressed cardiac transplants

    National Research Council Canada - National Science Library

    Favor, A; Frazier, O H; Cooley, D A; Okereke, O U; Radovancevic, B; Powers, P; Chandler, L

    1985-01-01

    ... of the care of transplant patients. One such opportunistic Organism, Legionella pneumophila, was responsible for four episodes of infection in three of our patients who survived due to better management of immunosuppression, together...

  1. Legionella Infection in Cyclosporine-Immunosuppressed Cardiac Transplants

    National Research Council Canada - National Science Library

    Favor, Arsenio; Frazier, O.H; Cooley, Denton A; Okereke, O.U. John; Radovancevic, Branislav; Powers, Penny; Chandler, Linda

    1985-01-01

    ... of the care of transplant patients. One such opportunistic Organism, Legionella pneumophila, was responsible for four episodes of infection in three of our patients who survived due to better management of immunosuppression, together...

  2. Survey of Legionella spp. in Mud Spring Recreation Area

    Science.gov (United States)

    Hsu, B.-M.; Ma, P.-H.; Su, I.-Z.; Chen, N.-S.

    2009-04-01

    Legionella genera are parasites of FLA, and intracellular bacterial replication within the FLA plays a major role in the transmission of disease. At least 13 FLA species—including Acanthamoeba spp., Naegleria spp., and Hartmannella spp.—support intracellular bacterial replication. In the study, Legionellae were detected with microbial culture or by direct DNA extraction and analysis from concentrated water samples or cultured free-living amoebae, combined with molecular methods that allow the taxonomic identification of these pathogens. The water samples were taken from a mud spring recreation area located in a mud-rock-formation area in southern Taiwan. Legionella were detected in 15 of the 34 samples (44.1%). Four of the 34 samples analyzed by Legionella culture were positive for Legionella, five of 34 were positive for Legionella when analyzed by direct DNA extraction and analysis, and 11 of 34 were positive for amoebae-resistant Legionella when analyzed by FLA culture. Ten samples were shown to be positive for Legionella by one analysis method and five samples were shown to be positive by two analysis methods. However, Legionella was detected in no sample by all three analysis methods. This suggests that the three analysis methods should be used together to detect Legionella in aquatic environments. In this study, L. pneumophila serotype 6 coexisted with A. polyphaga, and two uncultured Legionella spp. coexisted with either H. vermiformis or N. australiensis. Of the unnamed Legionella genotypes detected in six FLA culture samples, three were closely related to L. waltersii and the other three were closely related to L. pneumophila serotype 6. Legionella pneumophila serotype 6, L. drancourtii, and L. waltersii are noted endosymbionts of FLA and are categorized as pathogenic bacteria. This is significant for human health because these Legionella exist within FLA and thus come into contact with typically immunocompromised people.

  3. Legionella risk assessment in cruise ships and ferries

    Directory of Open Access Journals (Sweden)

    Pasqualina Laganà

    2017-06-01

    Legionella pneumophila sg 1 was isolated from the samples of shower and tap water in 7 (70% of the 10 ferries examined, and in 3 (33% of the 6 cruise ships examined, and L. pneumophila sg 2–14 in 8 (80% and 1 (16.7% of these ships, respectively. No Legionella contamination was found in whirlpool baths, air and ice samples. In conclusion, the data obtained confirm higher levels of Legionella contamination in local ferries and cruise ships, underlining the need to adopt corrective actions more specific for these smaller vessels.

  4. Identification of mip-like genes in the genus Legionella

    DEFF Research Database (Denmark)

    Cianciotto, N P; Bangsborg, Jette Marie; Eisenstein, B I

    1990-01-01

    The mip gene of Legionella pneumophila serogroup 1 strain AA100 encodes a 24-kilodalton surface protein (Mip) and enhances the abilities of L. pneumophila to parasitize human macrophages and to cause pneumonia in experimental animals. To determine whether this virulence factor is conserved...... in the genus Legionella, a large panel of Legionella strains was examined by Southern hybridization and immunoblot analyses for the presence and expression of mip-related sequences. Strains representing all 14 serogroups of L. pneumophila contained a mip gene and expressed a 24-kilodalton Mip protein. Although...... the isolates of the 29 other Legionella species did not hybridize with mip DNA probes under high-stringency conditions, they did so at reduced stringency. In support of the notion that these strains possess mip-like genes, these species each expressed a protein (24 to 31 kilodaltons in size) that reacted...

  5. Antibioticakeuze bij een Legionella-infectie

    NARCIS (Netherlands)

    de Vries, P.A.M.; van der Werf, T.S.; Manson, W L; Zijlstra, J.G,

    2005-01-01

    Legionella pneumophila is an intracellularly-growing microorganism and the causative agent of Legionnaires' disease; this disease owes its name to the epidemic among American war veterans in Philadelphia in 1976. The analysis ofthe epidemic in Philadelphia revealed--retrospectively--that unlike

  6. Atypical presentation of Legionella pneumonia among patients with underlying cancer: A fifteen-year review.

    Science.gov (United States)

    del Castillo, Maria; Lucca, Anabella; Plodkowski, Andrew; Huang, Yao-Ting; Kaplan, Janice; Gilhuley, Kathleen; Babady, N Esther; Seo, Susan K; Kamboj, Mini

    2016-01-01

    Immunocompromised patients, especially those receiving treatment with corticosteroids and cytotoxic chemotherapy are at increased risk for developing Legionella pneumonia. The aim of this study was to determine clinical and radiographic characteristics of pulmonary infection due to Legionella in persons undergoing treatment for cancer and stem cell transplant (SCT) recipients. Retrospective review of Legionella cases at MSKCC over a fifteen-year study period from January 1999 and December 2013. Cases were identified by review of microbiology records. During the study period, 40 cases of Legionella infection were identified; nine among these were due to non-pneumophila species. Most cases occurred during the summer. The majority [8/9, (89%)] of patients with non-pneumophila infection had underlying hematologic malignancy, compared to 18/31 (58%) with Legionella pneumophila infections. Radiographic findings were varied-nodular infiltrates mimicking invasive fungal infection were seen only among patients with hematologic malignancy and hematopoietic stem cell transplant (SCT) recipients and were frequently associated with non-pneumophila infections (50% vs 16%; P = 0.0594). All cases of nodular Legionella pneumonia were found incidentally or had an indolent clinical course. Legionella should be considered in the differential diagnosis of nodular lung lesions in immunocompromised patients, especially those with hematologic malignancy and SCT recipients. Most cases of nodular disease due to Legionella are associated with non-pneumophila infections. Copyright © 2015 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

  7. The case of malignancy mimicking legionella pneumonia

    Directory of Open Access Journals (Sweden)

    Ali Karakuş

    2013-09-01

    Full Text Available Legionella pneumophila is a bacterium, which can grow inwater pipe networks and climate systems. Contaminationoccurs by aspiration of infected water or aerosol inhalation.It is usually presented with fever, bradycardia, andchange in mental status, hyponatremia, elevation of liverenzymes and deterioration of renal function. The definitediagnosis is established by detection of the antigens andcultivating in the culture medium. Also, malign lung tumorscan encounter with the same clinical findings, so lungcancer should be remembered in differential diagnosis.The patient hospitalized for the Legionella pneumophiladue to the physical examination and laboratory findingsduring the first evaluation in the emergency department.However, further examinations pointed to the cancer. Weaimed to emphasize the probability of malignant tumorsin terms of hyponatremia, increase in the liver enzymes,and failure in the renal functions, which were usually experiencedin emergency unit. J Clin Exp Invest 2013; 4(3: 390-392Key words: Legionella pneumophila, pneumonia, lung malignancy

  8. A new oligonucleotide microarray for detection of pathogenic and non-pathogenic Legionella spp.

    Science.gov (United States)

    Cao, Boyang; Liu, Xiangqian; Yu, Xiang; Chen, Min; Feng, Lu; Wang, Lei

    2014-01-01

    Legionella pneumophila has been recognized as the major cause of legionellosis since the discovery of the deadly disease. Legionella spp. other than L. pneumophila were later found to be responsible to many non-pneumophila infections. The non-L. pneumophila infections are likely under-detected because of a lack of effective diagnosis. In this report, we have sequenced the 16S-23S rRNA gene internal transcribed spacer (ITS) of 10 Legionella species and subspecies, including L. anisa, L. bozemanii, L. dumoffii, L. fairfieldensis, L. gormanii, L. jordanis, L. maceachernii, L. micdadei, L. pneumophila subspp. fraseri and L. pneumophila subspp. pasculleii, and developed a rapid oligonucleotide microarray detection technique accordingly to identify 12 most common Legionella spp., which consist of 11 pathogenic species of L. anisa, L. bozemanii, L. dumoffii, L. gormanii, L. jordanis, L. longbeachae, L. maceachernii, L. micdadei, and L. pneumophila (including subspp. pneumophila, subspp. fraseri, and subspp. pasculleii) and one non-pathogenic species, L. fairfieldensis. Twenty-nine probes that reproducibly detected multiple Legionella species with high specificity were included in the array. A total of 52 strains, including 30 target pathogens and 22 non-target bacteria, were used to verify the oligonucleotide microarray assay. The sensitivity of the detection was at 1.0 ng with genomic DNA or 13 CFU/100 mL with Legionella cultures. The microarray detected seven samples of air conditioner-condensed water with 100% accuracy, validating the technique as a promising method for applications in basic microbiology, clinical diagnosis, food safety, and epidemiological surveillance. The phylogenetic study based on the ITS has also revealed that the non-pathogenic L. fairfieldensis is the closest to L. pneumophila than the nine other pathogenic Legionella spp.

  9. Multiplex real-time PCR assay for Legionella species.

    Science.gov (United States)

    Kim, Seung Min; Jeong, Yoojung; Sohn, Jang Wook; Kim, Min Ja

    2015-12-01

    Legionella pneumophila serogroup 1 (sg1) accounts for the majority of infections in humans, but other Legionella species are also associated with human disease. In this study, a new SYBR Green I-based multiplex real-time PCR assay in a single reaction was developed to allow the rapid detection and differentiation of Legionella species by targeting specific gene sequences. Candidate target genes were selected, and primer sets were designed by referring to comparative genomic hybridization data of Legionella species. The Legionella species-specific groES primer set successfully detected all 30 Legionella strains tested. The xcpX and rfbA primers specifically detected L. pneumophila sg1-15 and L. pneumophila sg1, respectively. In addition, this assay was validated by testing clinical samples and isolates. In conclusion, this novel multiplex real-time PCR assay might be a useful diagnostic tool for the rapid detection and differentiation of Legionella species in both clinical and epidemiological studies. Copyright © 2015 Elsevier Ltd. All rights reserved.

  10. Presence of Legionella and Free-Living Amoebae in Composts and Bioaerosols from Composting Facilities

    Science.gov (United States)

    Conza, Lisa; Pagani, Simona Casati; Gaia, Valeria

    2013-01-01

    Several species of Legionella cause Legionnaires’ disease (LD). Infection may occur through inhalation of Legionella or amoebal vesicles. The reservoirs of Legionella are water, soil, potting soil and compost. Some species of free-living amoebae (FLA) that are naturally present in water and soil were described as hosts for Legionella. This study aimed to understand whether or not the composting facilities could be sources of community-acquired Legionella infections after development of bioaerosols containing Legionella or FLA. We looked for the presence of Legionella (by co-culture) and FLA (by culture) in composts and bioaerosols collected at four composting facilities located in southern Switzerland. We investigated the association between the presence of Legionella and compost and air parameters and presence of FLA. Legionella spp. (including L. pneumophila) were detected in 69.3% (61/88) of the composts and FLA (mainly Acanthamoeba, Vermamoeba, Naegleria and Stenamoeba) in 92.0% (81/88). L. pneumophila and L. bozemanii were most frequently isolated. FLA as potential host for Legionella spp. were isolated from 40.9% (36/88) of the composts in all facilities. In Legionella-positive samples the temperature of compost was significantly lower (P = 0.012) than in Legionella-negative samples. Of 47 bioaerosol samples, 19.1% (9/47) were positive for FLA and 10.6% (5/47) for L. pneumophila. Composts (62.8%) were positive for Legionella and FLA contemporaneously, but both microorganisms were never detected simultaneously in bioaerosols. Compost can release bioaerosol containing FLA or Legionella and could represent a source of infection of community-acquired Legionella infections for workers and nearby residents. PMID:23844174

  11. Cross-reactive Legionella antigens and the antibody response during infection

    DEFF Research Database (Denmark)

    Bangsborg, Jette Marie; Shand, G; Pearlman, E

    1991-01-01

    In order to define cross-reactive Legionella antigens suitable for diagnostic purposes, we investigated sonicate antigens from two Legionella species, including two serogroups of L. pneumophila. The antigens were reacted with heterologous and homologous rabbit antisera in Western blot. Sera from...

  12. A comparison of assays measuring the viability of Legionella ...

    Science.gov (United States)

    Background: The relatively high prevalence of Legionella pneumophila in premise plumbing systems has been widely reported. Published reports indicate Legionella has a comparatively high resistance to chlorine and moreover has the ability to grow in phagocytic amoeba which could provide additional protection in chlorinated drinking water distribution systems. Copper-Silver (Cu-Ag) ionization treatment systems are commercially available for use in large building water systems to help control the risks from Legionella bacteria. The objectives of this study were to develop and optimize Legionella viability assays and use them to investigate the viability of Legionella bacteria after exposure to water treated with coppper and silver ions. Methods: Log phase L. pneumophila cells were used in all experiments and were generated by incubation at 35C for 48 hours in buffered yeast extract broth. Viability assays used included plating on buffered charcoal yeast extract agar to determine the number of culturable cells and treating cells with propidium monoazide (PMA) or ethidium monoazide (EMA) followed by quantitative PCR targeting mip gene of L. pneumophila. The qPCR viability assays were optimized using L. pneumophila inactivated by heat treatment at 65C for 60 min. The effectiveness of Cu-Ag ionization treatment was studied by inoculating L. pneumonia at 105 CFU/mL in water collected directly from a building water system that employed this technology and incubat

  13. Occurrence of Legionella in UK household showers.

    Science.gov (United States)

    Collins, Samuel; Stevenson, David; Bennett, Allan; Walker, Jimmy

    2017-04-01

    Household water systems have been proposed as a source of sporadic, community acquired Legionnaires' disease. Showers represent a frequently used aerosol generating device in the domestic setting yet little is known about the occurrence of Legionella spp. in these systems. This study has investigated the prevalence of Legionella spp. by culture and qPCR in UK household showers. Ninety nine showers from 82 separate properties in the South of England were sampled. Clinically relevant Legionella spp. were isolated by culture in 8% of shower water samples representing 6% of households. Legionella pneumophila sg1 ST59 was isolated from two showers in one property and air sampling demonstrated its presence in the aerosol state. A further 31% of showers were positive by Legionella spp. qPCR. By multi-variable binomial regression modelling Legionella spp. qPCR positivity was associated with the age of the property (p=0.02), the age of the shower (p=0.01) and the frequency of use (p=0.09). The concentration of Legionella spp. detected by qPCR was shown to decrease with increased frequency of use (p=0.04) and more frequent showerhead cleaning (p=0.05). There was no association between Legionella spp. qPCR positivity and the cold water supply or the showerhead material (p=0.65 and p=0.71, respectively). Household showers may be important reservoirs of clinically significant Legionella and should be considered in source investigations. Simple public health advice may help to mitigate the risk of Legionella exposure in the domestic shower environment. Crown Copyright © 2016. Published by Elsevier GmbH. All rights reserved.

  14. Surveillance of parasitic Legionella in surface waters by using immunomagnetic separation and amoebae enrichment.

    Science.gov (United States)

    Hsu, Tsui-Kang; Wu, Shu-Fen; Hsu, Bing-Mu; Kao, Po-Min; Tao, Chi-Wei; Shen, Shu-Min; Ji, Wen-Tsai; Huang, Wen-Chien; Fan, Cheng-Wei

    2015-01-01

    Free-living amoebae (FLA) are potential reservoirs of Legionella in aquatic environments. However, the parasitic relationship between various Legionella and amoebae remains unclear. In this study, surface water samples were gathered from two rivers for evaluating parasitic Legionella. Warmer water temperature is critical to the existence of Legionella. This result suggests that amoebae may be helpful in maintaining Legionella in natural environments because warmer temperatures could enhance parasitisation of Legionella in amoebae. We next used immunomagnetic separation (IMS) to identify extracellular Legionella and remove most free Legionella before detecting the parasitic ones in selectively enriched amoebae. Legionella pneumophila was detected in all the approaches, confirming that the pathogen is a facultative amoebae parasite. By contrast, two obligate amoebae parasites, Legionella-like amoebal pathogens (LLAPs) 8 and 9, were detected only in enriched amoebae. However, several uncultured Legionella were detected only in the extracellular samples. Because the presence of potential hosts, namely Vermamoeba vermiformis, Acanthamoeba spp. and Naegleria gruberi, was confirmed in the samples that contained intracellular Legionella, uncultured Legionella may survive independently of amoebae. Immunomagnetic separation and amoebae enrichment may have referential value for detecting parasitic Legionella in surface waters.

  15. Intra-Amoeba Multiplication Induces Chemotaxis and Biofilm Colonization and Formation for Legionella

    Science.gov (United States)

    Bigot, Renaud; Bertaux, Joanne; Frere, Jacques; Berjeaud, Jean-Marc

    2013-01-01

    Legionella pneumophila, a facultative intracellular bacterium, is the causative agent of legionellosis. In the environment this pathogenic bacterium colonizes the biofilms as well as amoebae, which provide a rich environment for the replication of Legionella. When seeded on pre-formed biofilms, L. pneumophila was able to establish and survive and was only found at the surface of the biofilms. Different phenotypes were observed when the L. pneumophila, used to implement pre-formed biofilms or to form mono-species biofilms, were cultivated in a laboratory culture broth or had grown intracellulary within the amoeba. Indeed, the bacteria, which developed within the amoeba, formed clusters when deposited on a solid surface. Moreover, our results demonstrate that multiplication inside the amoeba increased the capacity of L. pneumophila to produce polysaccharides and therefore enhanced its capacity to establish biofilms. Finally, it was shown that the clusters formed by L. pneumophila were probably related to the secretion of a chemotaxis molecular agent. PMID:24205008

  16. Legionella prevalence and risk of legionellosis in Japanese households.

    Science.gov (United States)

    Kuroki, T; Watanabe, Y; Teranishi, H; Izumiyama, S; Amemura-Maekawa, J; Kura, F

    2017-05-01

    This study determined the occurrence of legionellae in private houses for which there were no available data on aquatic environments other than the water supply system. From June 2013 to November 2014, we collected 138 water and 90 swab samples from aquatic environments in 19 houses. Legionella DNA was detected via a loop-mediated isothermal amplification assay in 66 (47·8%) water and 17 (18·9%) swab samples. High Legionella DNA detection rates were observed in water samples from washing machines and aquariums. Legionella spp. was isolated from 9 (6·5%) water and 3 (3·3%) swab samples. Legionella pneumophila SG 1 was detected from the outlet water of a bathtub spout and a bath sponge. Use of amoebic co-culture effectively increased legionellae and Legionella DNA detection rates from all sample types. A logistic regression analysis revealed that the heterotrophic plate count was significantly related to Legionella contamination. Our findings indicate that there is a risk of legionellosis from exposure to Legionella spp. in a variety of aquatic environments in residential houses. Control measures for legionellae in houses should include frequent cleaning and disinfecting to reduce heterotrophic bacteria in water and, where possible, preventing aerosolization from aquatic environments.

  17. Different distribution patterns of ten virulence genes in Legionella reference strains and strains isolated from environmental water and patients.

    Science.gov (United States)

    Zhan, Xiao-Yong; Hu, Chao-Hui; Zhu, Qing-Yi

    2016-04-01

    Virulence genes are distinct regions of DNA which are present in the genome of pathogenic bacteria and absent in nonpathogenic strains of the same or related species. Virulence genes are frequently associated with bacterial pathogenicity in genus Legionella. In the present study, an assay was performed to detect ten virulence genes, including iraA, iraB, lvrA, lvrB, lvhD, cpxR, cpxA, dotA, icmC and icmD in different pathogenicity islands of 47 Legionella reference strains, 235 environmental strains isolated from water, and 4 clinical strains isolated from the lung tissue of pneumonia patients. The distribution frequencies of these genes in reference or/and environmental L. pneumophila strains were much higher than those in reference non-L. pneumophila or/and environmental non-L. pneumophila strains, respectively. L. pneumophila clinical strains also maintained higher frequencies of these genes compared to four other types of Legionella strains. Distribution frequencies of these genes in reference L. pneumophila strains were similar to those in environmental L. pneumophila strains. In contrast, environmental non-L. pneumophila maintained higher frequencies of these genes compared to those found in reference non-L. pneumophila strains. This study illustrates the association of virulence genes with Legionella pathogenicity and reveals the possible virulence evolution of non-L. pneumophia strains isolated from environmental water.

  18. Occurrence and Control of Legionella in Recycled Water Systems

    Science.gov (United States)

    Jjemba, Patrick K.; Johnson, William; Bukhari, Zia; LeChevallier, Mark W.

    2015-01-01

    Legionella pneumophila is on the United States Environmental Protection Agency (USEPA) Candidate Contaminant list (CCL) as an important pathogen. It is commonly encountered in recycled water and is typically associated with amoeba, notably Naegleria fowleri (also on the CCL) and Acanthamoeba sp. No legionellosis outbreak has been linked to recycled water and it is important for the industry to proactively keep things that way. A review was conducted examine the occurrence of Legionella and its protozoa symbionts in recycled water with the aim of developing a risk management strategy. The review considered the intricate ecological relationships between Legionella and protozoa, methods for detecting both symbionts, and the efficacy of various disinfectants. PMID:26140674

  19. Effector glycosyltransferases in Legionella

    Directory of Open Access Journals (Sweden)

    Yury eBelyi

    2011-04-01

    Full Text Available Legionella causes severe pneumonia in humans. The pathogen produces an array of effectors, which interfere with host cell functions. Among them are the glucosyltransferases Lgt1, Lgt2 and Lgt3 from L. pneumophila. Lgt1 and Lgt2 are produced predominately in the post-exponential phase of bacterial growth, while synthesis of Lgt3 is induced mainly in the lag-phase before intracellular replication of bacteria starts. Lgt glucosyltransferases are structurally similar to clostridial glucosylating toxins. The enzymes use UDP-glucose as a donor substrate and modify eukaryotic elongation factor eEF1A at serine-53. This modification results in inhibition of protein synthesis and death of target cells. In addition to Lgts, Legionella genomes disclose several genes, coding for effector proteins likely to possess glycosyltransferase activities, including SetA, which influences vesicular trafficking in the yeast model system and displays tropism for late endosomal/lysosomal compartments of mammalian cells. This review mainly discusses recent results on the structure-function relationship of Lgt glucosyltransferases.

  20. Metabolism of the vacuolar pathogen Legionella and implications for virulence

    Directory of Open Access Journals (Sweden)

    Christian eManske

    2014-09-01

    Full Text Available Legionella pneumophila is a ubiquitous environmental bacterium that thrives in fresh water habitats, either as planktonic form or as part of biofilms. The bacteria also grow intracellularly in free-living protozoa as well as in mammalian alveolar macrophages, thus triggering a potentially fatal pneumonia called Legionnaires’ disease. To establish its intracellular niche termed the Legionella-containing vacuole (LCV, L. pneumophila employs a type IV secretion system and translocates ~300 different effector proteins into host cells. The pathogen switches between two distinct forms to grow in its extra- or intracellular niches: transmissive bacteria are virulent for phagocytes, and replicative bacteria multiply within their hosts. The switch between these forms is regulated by different metabolic cues that signal conditions favorable for replication or transmission, respectively, causing a tight link between metabolism and virulence of the bacteria.Amino acids represent the prime carbon and energy source of extra- or intracellularly growing L. pneumophila. Yet, the genome sequences of several Legionella spp. as well as transcriptome and proteome data and metabolism studies indicate that the bacteria possess broad catabolic capacities and also utilize carbohydrates such as glucose. Accordingly, L. pneumophila mutant strains lacking catabolic genes show intracellular growth defects, and thus, intracellular metabolism and virulence of the pathogen are intimately connected. In this review we will summarize recent findings on the extra- and intracellular metabolism of L. pneumophila using genetic, biochemical and cellular microbial approaches. Recent progress in this field sheds light on the complex interplay between metabolism, differentiation and virulence of the pathogen.

  1. Metabolism of the vacuolar pathogen Legionella and implications for virulence.

    Science.gov (United States)

    Manske, Christian; Hilbi, Hubert

    2014-01-01

    Legionella pneumophila is a ubiquitous environmental bacterium that thrives in fresh water habitats, either as planktonic form or as part of biofilms. The bacteria also grow intracellularly in free-living protozoa as well as in mammalian alveolar macrophages, thus triggering a potentially fatal pneumonia called "Legionnaires' disease." To establish its intracellular niche termed the "Legionella-containing vacuole" (LCV), L. pneumophila employs a type IV secretion system and translocates ~300 different "effector" proteins into host cells. The pathogen switches between two distinct forms to grow in its extra- or intracellular niches: transmissive bacteria are virulent for phagocytes, and replicative bacteria multiply within their hosts. The switch between these forms is regulated by different metabolic cues that signal conditions favorable for replication or transmission, respectively, causing a tight link between metabolism and virulence of the bacteria. Amino acids represent the prime carbon and energy source of extra- or intracellularly growing L. pneumophila. Yet, the genome sequences of several Legionella spp. as well as transcriptome and proteome data and metabolism studies indicate that the bacteria possess broad catabolic capacities and also utilize carbohydrates such as glucose. Accordingly, L. pneumophila mutant strains lacking catabolic genes show intracellular growth defects, and thus, intracellular metabolism and virulence of the pathogen are intimately connected. In this review we will summarize recent findings on the extra- and intracellular metabolism of L. pneumophila using genetic, biochemical and cellular microbial approaches. Recent progress in this field sheds light on the complex interplay between metabolism, differentiation and virulence of the pathogen.

  2. Detection of Legionella Contamination in Tabriz Hospitals by PCR Assay

    Science.gov (United States)

    Ghotaslou, Reza; Yeganeh Sefidan, Fatemeh; Akhi, Mohammad Taghi; Soroush, Mohammad Hussein; Hejazi, Mohammad Saeid

    2013-01-01

    Purpose: The present study was designed to evaluate the occurrence of Legionella contamination in the tap water of Tabriz hospitals, Azerbaijan, Iran. Methods: One hundred and forty water samples from diverse water supply systems of 17 hospitals were collected and analyzed for the presence of Legionella spp. by PCR assay. Results: In this study, 10 of 140 (7.1%) samples were positive for Legionella which L. pneumophila was detected in 4 (2.85%) water samples. Conclusion: In conclusion, hospital potable systems are the primary reservoirs for Legionnaires’ disease. This study concludes that Legionella spp. are present in aquatic hospitals environment of Tabriz. Due to the serious risk of infections, it is better to make efforts to eliminate Legionella spp. in water supplies. PMID:24312825

  3. Hospital washbasin water: risk of Legionella-contaminated aerosol inhalation.

    Science.gov (United States)

    Cassier, P; Landelle, C; Reyrolle, M; Nicolle, M C; Slimani, S; Etienne, J; Vanhems, P; Jarraud, S

    2013-12-01

    The contamination of aerosols by washbasin water colonized by Legionella in a hospital was evaluated. Aerosol samples were collected by two impingement technologies. Legionella was never detected by culture in all the (aerosol) samples. However, 45% (18/40) of aerosol samples were positive for Legionella spp. by polymerase chain reaction, with measurable concentrations in 10% of samples (4/40). Moreover, immunoassay detected Legionella pneumophila serogroup 1 and L. anisa, and potentially viable bacteria were seen on viability testing. These data suggest that colonized hospital washbasins could represent risks of exposure to Legionella aerosol inhalation, especially by immunocompromised patients. Copyright © 2013 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.

  4. Detection of Legionella Contamination in Tabriz Hospitals by PCR Assay

    Directory of Open Access Journals (Sweden)

    Mohammad Saeid Hejazi

    2013-02-01

    Full Text Available Purpose: The present study was designed to evaluate the occurrence of Legionella contamination in the tap water of Tabriz hospitals, Azerbaijan, Iran. Methods: One hundred and forty water samples from diverse water supply systems of 17 hospitals were collected and analyzed for the presence of Legionella spp. by PCR assay. Results: In this study, 10 of 140 (7.1% samples were positive for Legionella which L. pneumophila was detected in 4 (2.85% water samples. Conclusion: In conclusion, hospital potable systems are the primary reservoirs for Legionnaires’ disease. This study concludes that Legionella spp. are present in aquatic hospitals environment of Tabriz. Due to the serious risk of infections, it is better to make efforts to eliminate Legionella spp. in water supplies.

  5. A case of nosocomial Legionella pneumonia associated with a contaminated hospital cooling tower.

    Science.gov (United States)

    Osawa, Kayo; Shigemura, Katsumi; Abe, Yasuhisa; Jikimoto, Takumi; Yoshida, Hiroyuki; Fujisawa, Masato; Arakawa, Soichi

    2014-01-01

    We report the epidemiological investigation of a nosocomial pneumonia case due to Legionella pneumophila linked to a contaminated hospital cooling tower in an immune-compromised patient. A 73-year-old female patient was diagnosed with nosocomial Legionella pneumonia proven by a culture of L. pneumophila serogroup 1 from bronchoalveolar lavage fluid. Two strains isolated from the patient and two strains isolated from two cooling towers were found to be identical using repetitive-sequence-based-PCR with a 95% probability. This Legionella pneumonia case might be caused by aerosol from cooling towers on the roof of the hospital building which was contaminated by L. pneumophila. We increased up the temperature of hot water supply appropriately for prevention of Legionella breeding in an environment of patients' living. On the other hand, as the maintenance of cooling tower, we increased the frequency of Legionella culture tests from twice a year to three times a year. In addition, we introduced an automated disinfectants insertion machine and added one antiseptic reagent (BALSTER ST-40 N, Tohzai Chemical Industry Co., Ltd., Kawasaki, Japan) after this Legionella disease, and thereafter, we have no additional cases of Legionella disease or detection of Legionella spp. from the cooling tower or hot water supply. This case demonstrates the importance of detecting the infection source and carrying out environmental maintenance in cooperation with the infection control team. Copyright © 2013 Japanese Society of Chemotherapy and the Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

  6. Characterization of the tail-specific protease (Tsp) from Legionella.

    Science.gov (United States)

    Lawrence, Amba; K Nicholls, Simon; H Stansfield, Scott; M Huston, Wilhelmina

    2014-01-01

    Bacterial tail-specific proteases (Tsps) have been attributed a wide variety of functions including intracellular virulence, cell wall morphology, proteolytic signal cascades and stress response. This study tested the hypothesis that Tsp has a key function for the transmissive form of Legionella pneumophila. A tsp mutant was generated in Legionella pneumophila 130b and the characteristics of this strain and the isogenic wild-type were examined using a range of growth and proteomic analyses. Recombinant Tsp protein was also produced and analyzed. The L. pneumophila tsp mutant showed no defect in growth on rich media or during thermo-osmotic stress conditions. In addition, no defects in cellular morphology were observed when the cells were examined using transmission electron microscopy. Purified recombinant Tsp was found to be an active protease with a narrow substrate range. Proteome analysis using iTRAQ (5% coverage of the proteome) found that, of those proteins detected, only 5 had different levels in the tsp mutant compared to the wild type. ACP (Acyl Carrier Protein), which has a key role for Legionella differentiation to the infectious form, was reduced in the tsp mutant; however, tsp(-) was able to infect and replicate inside macrophages to the same extent as the wild type. Combined, these data demonstrate that Tsp is a protease but is not essential for Legionella growth or cell infection. Thus, Tsp may have functional redundancy in Legionella.

  7. Exploring anti-bacterial compounds against intracellular Legionella.

    Directory of Open Access Journals (Sweden)

    Christopher F Harrison

    Full Text Available Legionella pneumophila is a ubiquitous fresh-water bacterium which reproduces within its erstwhile predators, environmental amoeba, by subverting the normal pathway of phagocytosis and degradation. The molecular mechanisms which confer resistance to amoeba are apparently conserved and also allow replication within macrophages. Thus, L. pneumophila can act as an 'accidental' human pathogen and cause a severe pneumonia known as Legionnaires' disease. The intracellular localisation of L. pneumophila protects it from some antibiotics, and this fact must be taken into account to develop new anti-bacterial compounds. In addition, the intracellular lifestyle of L. pneumophila may render the bacteria susceptible to compounds diminishing bacterial virulence and decreasing intracellular survival and replication of this pathogen. The development of a single infection cycle intracellular replication assay using GFP-producing L. pneumophila and Acanthamoebacastellanii amoeba is reported here. This fluorescence-based assay allows for continuous monitoring of intracellular replication rates, revealing the effect of bacterial gene deletions or drug treatment. To examine how perturbations of the host cell affect L. pneumophila replication, several known host-targeting compounds were tested, including modulators of cytoskeletal dynamics, vesicle scission and Ras GTPase localisation. Our results reveal a hitherto unrealized potential antibiotic property of the β-lactone-based Ras depalmitoylation inhibitor palmostatin M, but not the closely related inhibitor palmostatin B. Further characterisation indicated that this compound caused specific growth inhibition of Legionella and Mycobacterium species, suggesting that it may act on a common bacterial target.

  8. Population structure of Legionella spp. from environmental samples in Gabon, 2013.

    Science.gov (United States)

    Ehrhardt, Jonas; Alabi, Abraham S; Kuczius, Thorsten; Tsombeng, Francis Foguim; Becker, Karsten; Kremsner, Peter G; Schaumburg, Frieder; Esen, Meral

    2015-07-01

    Aquatic environments are the most important source for Legionella spp. infections such as Legionnaires' disease and Pontiac fever. The reservoirs of Legionella spp. are mostly unclear in sub-Saharan Africa. The aim of this study, conducted in 2013, was to identify geographical areas of an increased risk for exposure to Legionella spp., and to describe the population structure of Legionella spp. from different water sources in a cross-sectional study in Gabon. Fresh water samples (n = 200) were cultured on Legionella selective agar; species were confirmed by MALDI-TOF, a Legionella pneumophila specific real-time PCR and 16S RNA gene sequencing. Serogroups were identified by agglutination test. The population structure was assessed by multilocus sequence typing (MLST). Legionella spp. isolates (n = 29) were frequently found in the hospital setting particularly in hot water systems. Open water bodies (i.e. rivers, lakes) were not contaminated with Legionella spp. Isolated L. pneumophila mainly belonged to serogroups 2-14 (n = 19) and MLST sequence type ST1, ST75 (and related STs) and ST1911. In conclusion, hospitalized patients might have an increased risk to become infected with Legionella spp. in the studied areas in Gabon, particularly if they have risk factors such as comorbidities. Both broadly extended (ST1, ST75) and local lineages (ST1911) were present in our setting. Copyright © 2015 Elsevier B.V. All rights reserved.

  9. Influenceof antimicrobial therapy on the sensitivity of legionella PCR - three case report

    Directory of Open Access Journals (Sweden)

    Renato Eržen

    2007-02-01

    Full Text Available Background: L. pneumophila is a possible causative agent of severe community and hospital acquired pneumonia. Available diagnostic tests have a lot of shortcomings. Detection of Legionella DNA with the PCR technique is a promissing new method allowing diagnosis within clinically useful time.Methods: Three cases of severe Legionella pneumonia were followed with the PCR technique, urinary antigen and serologic methods.Results: Diagnostic tests have different sensitivity and specificity and also different shortcomings. Serologic tests are unable to provide diagnosis within clinically useful time. Urinary antigen is specific just for L. pneumophila serotype 1. Antimicrobial treatment has impact on the sensitivity of Legionella PCR.Conclusions: Detection of L. pneumophila infection with the PCR technique in addition to urinary antigen testing is likely to be the best diagnostic tool for the detection of all species within a time frame that will affect clinical management.

  10. Effect of Common Drinking Water Disinfectants, Chlorine and Heat, on Free Legionella and Amoebae-Associated Legionella

    Science.gov (United States)

    Cervero-Aragó, Sílvia; Rodríguez-Martínez, Sarah; Puertas-Bennasar, Antoni; Araujo, Rosa M.

    2015-01-01

    Chlorine and thermal treatments are the most commonly used procedures to control and prevent Legionella proliferation in drinking water systems of large buildings. However, cases of legionellosis still occur in facilities with treated water. The purpose of this work was to model the effect of temperature and free chlorine applied in similar exposure conditions as in drinking water systems on five Legionella spp. strains and two amoebal strains of the genera Acanthamoeba. Inactivation models obtained were used to determine the effectiveness of the treatments applied which resulted more effective against Legionella than Acanthamoeba, especially those in cystic stages. Furthermore, to determine the influence of the relationship between L. pneumophila and Acanthamoeba spp. on the treatment effectiveness, inactivation models of the bacteria-associated amoeba were also constructed and compared to the models obtained for the free living bacteria state. The Legionella-amoeba association did not change the inactivation models, but it reduced the effectiveness of the treatments applied. Remarkably, at the lowest free chlorine concentration, 0.5 mg L-1, as well as at the lowest temperatures, 50°C and 55°C, the influence of the Legionella-amoeba associate state was the strongest in reducing the effectiveness of the treatments compared to the free Legionella state. Therefore, the association established between L. pneumophila and amoebae in the water systems indicate an increased health risk in proximal areas of the system (close to the tap) where lower free chlorine concentrations and lower temperatures are commonly observed. PMID:26241039

  11. Effect of Common Drinking Water Disinfectants, Chlorine and Heat, on Free Legionella and Amoebae-Associated Legionella.

    Science.gov (United States)

    Cervero-Aragó, Sílvia; Rodríguez-Martínez, Sarah; Puertas-Bennasar, Antoni; Araujo, Rosa M

    2015-01-01

    Chlorine and thermal treatments are the most commonly used procedures to control and prevent Legionella proliferation in drinking water systems of large buildings. However, cases of legionellosis still occur in facilities with treated water. The purpose of this work was to model the effect of temperature and free chlorine applied in similar exposure conditions as in drinking water systems on five Legionella spp. strains and two amoebal strains of the genera Acanthamoeba. Inactivation models obtained were used to determine the effectiveness of the treatments applied which resulted more effective against Legionella than Acanthamoeba, especially those in cystic stages. Furthermore, to determine the influence of the relationship between L. pneumophila and Acanthamoeba spp. on the treatment effectiveness, inactivation models of the bacteria-associated amoeba were also constructed and compared to the models obtained for the free living bacteria state. The Legionella-amoeba association did not change the inactivation models, but it reduced the effectiveness of the treatments applied. Remarkably, at the lowest free chlorine concentration, 0.5 mg L-1, as well as at the lowest temperatures, 50°C and 55°C, the influence of the Legionella-amoeba associate state was the strongest in reducing the effectiveness of the treatments compared to the free Legionella state. Therefore, the association established between L. pneumophila and amoebae in the water systems indicate an increased health risk in proximal areas of the system (close to the tap) where lower free chlorine concentrations and lower temperatures are commonly observed.

  12. Legionella clemsonensis sp. nov.: a green fluorescing Legionella strain from a patient with pneumonia.

    Science.gov (United States)

    Palmer, Allison; Painter, Joseph; Hassler, Hayley; Richards, Vincent P; Bruce, Terri; Morrison, Shatavia; Brown, Ellen; Kozak-Muiznieks, Natalia A; Lucas, Claressa; McNealy, Tamara L

    2016-10-01

    A novel Legionella species was identified based on sequencing, cellular fatty acid analysis, biochemical reactions, and biofilm characterization. Strain D5610 was originally isolated from the bronchial wash of a patient in Ohio, USA. The bacteria were gram-negative, rod-shaped, and exhibited green fluorescence under long wave UV light. Phylogenetic analysis and fatty acid composition revealed a distinct separation within the genus. The strain grows between 26-45°C and forms biofilms equivalent to L. pneumophila Philadelphia 1. These characteristics suggest that this isolate is a novel Legionella species, for which the name Legionella clemsonensis sp nov. is proposed. © 2016 The Societies and John Wiley & Sons Australia, Ltd.

  13. Prolonged effect of two combined methods for Legionella disinfection in a hospital water system.

    Science.gov (United States)

    Casari, E; Ferrario, A; Montanelli, A

    2007-01-01

    This study was conducted in our 650 bed general hospital, which is situated on the southern outskirts of Milan (Italy). After a first nosocomial case of pneumonia (caused by Legionella pneumophila serogroup 1), we first used a conventional method (heat shock) without success. To solve the problem we then tried a copper-silver ionization system combined with a chlorine dioxide device. During the four years after the installation of these two systems we recorded a significant (p ionization system, combined with a chlorine dioxide device, is a highly promising method for the control of Legionella pneumophila in a hospital water distribution system.

  14. Recreational Vehicle Water Tanks as a Possible Source for Legionella Infections

    Directory of Open Access Journals (Sweden)

    Christine M. Litwin

    2013-01-01

    Full Text Available We investigated recreational vehicle (RV water reservoirs in response to a case of pneumonia in which Legionella pneumophila was cultured both from the patient and a RV reservoir in which he travelled. Water samples processed and cultured at the CDC according to standard protocol were positive for Legionella spp. in 4/17 (24% faucets, 1/11 (9% water tanks from 4/20 (20% RVs from three different campsites. Legionella spp. that were isolated included L. pneumophila (serogroups 1 and 6, L. anisa, L. feeleii, and L. quateriensis. Environmental controls from the potable water of the three campsites were culture-negative. A survey of maintenance practices by the RV users at the campsites revealed that chlorine disinfection of the water tanks was rarely performed. To prevent the possibility of Legionella infections, RV owners should implement regular chlorine disinfection of their water tanks and follow the recommended maintenance guidelines according to their owner's manuals.

  15. Legionella: a major opportunistic pathogen in transplant recipients.

    Science.gov (United States)

    Chow, J W; Yu, V L

    1998-06-01

    Legionella have a predilection for infecting immunocompromised patients, and transplant recipients have the highest risk. Legionella spp have been the most common cause of nosocomial pneumonia among transplant recipients at selected medical centers. Diagnosis is dependent on the ability of the clinical microbiology laboratory to isolate the organism by culture; therefore, the disease is easily overlooked. The mode of transmission of Legionella pneumophila is likely aspiration in transplant recipients. Clinical manifestations are similar to that of other bacterial pneumonias, although diarrhea is often prominent. The quinolone antibiotics (especially ciprofloxacin) are the antibiotics of choice because, unlike the macrolides or rifampin, they do not interact with the immunosuppressive agents used to counter rejection. Prevention of nosocomial legionellosis involves disinfection of the hospital's potable water system. Effective disinfection methods include superheat and flush or copper-silver ionization; hyperchlorination is no longer recommended. Routine culture surveillance directed at the hospital water supply for Legionella is mandatory in hospitals caring for transplant patients.

  16. Reduction of Legionella spp. in Water and in Soil by a Citrus Plant Extract Vapor

    Science.gov (United States)

    Kurzbach, Elena; Score, Jodie; Tejpal, Jyoti; Chi Tangyie, George; Phillips, Carol

    2014-01-01

    Legionnaires' disease is a severe form of pneumonia caused by Legionella spp., organisms often isolated from environmental sources, including soil and water. Legionella spp. are capable of replicating intracellularly within free-living protozoa, and once this has occurred, Legionella is particularly resistant to disinfectants. Citrus essential oil (EO) vapors are effective antimicrobials against a range of microorganisms, with reductions of 5 log cells ml−1 on a variety of surfaces. The aim of this investigation was to assess the efficacy of a citrus EO vapor against Legionella spp. in water and in soil systems. Reductions of viable cells of Legionella pneumophila, Legionella longbeachae, Legionella bozemanii, and an intra-amoebal culture of Legionella pneumophila (water system only) were assessed in soil and in water after exposure to a citrus EO vapor at concentrations ranging from 3.75 mg/liter air to 15g/liter air. Antimicrobial efficacy via different delivery systems (passive and active sintering of the vapor) was determined in water, and gas chromatography-mass spectrometry (GC-MS) analysis of the antimicrobial components (linalool, citral, and β-pinene) was conducted. There was up to a 5-log cells ml−1 reduction in Legionella spp. in soil after exposure to the citrus EO vapors (15 mg/liter air). The most susceptible strain in water was L. pneumophila, with a 4-log cells ml−1 reduction after 24 h via sintering (15 g/liter air). Sintering the vapor through water increased the presence of the antimicrobial components, with a 61% increase of linalool. Therefore, the appropriate method of delivery of an antimicrobial citrus EO vapor may go some way in controlling Legionella spp. from environmental sources. PMID:25063652

  17. Method modification of the Legipid® Legionella fast detection test kit.

    Science.gov (United States)

    Albalat, Guillermo Rodríguez; Broch, Begoña Bedrina; Bono, Marisa Jiménez

    2014-01-01

    Legipid(®) Legionella Fast Detection is a test based on combined magnetic immunocapture and enzyme-immunoassay (CEIA) for the detection of Legionella in water. The test is based on the use of anti-Legionella antibodies immobilized on magnetic microspheres. Target microorganism is preconcentrated by filtration. Immunomagnetic analysis is applied on these preconcentrated water samples in a final test portion of 9 mL. The test kit was certified by the AOAC Research Institute as Performance Tested Method(SM) (PTM) No. 111101 in a PTM validation which certifies the performance claims of the test method in comparison to the ISO reference method 11731-1998 and the revision 11731-2004 "Water Quality: Detection and Enumeration of Legionella pneumophila" in potable water, industrial water, and waste water. The modification of this test kit has been approved. The modification includes increasing the target analyte from L. pneumophila to Legionella species and adding an optical reader to the test method. In this study, 71 strains of Legionella spp. other than L. pneumophila were tested to determine its reactivity with the kit based on CEIA. All the strains of Legionella spp. tested by the CEIA test were confirmed positive by reference standard method ISO 11731. This test (PTM 111101) has been modified to include a final optical reading. A methods comparison study was conducted to demonstrate the equivalence of this modification to the reference culture method. Two water matrixes were analyzed. Results show no statistically detectable difference between the test method and the reference culture method for the enumeration of Legionella spp. The relative level of detection was 93 CFU/volume examined (LOD50). For optical reading, the LOD was 40 CFU/volume examined and the LOQ was 60 CFU/volume examined. Results showed that the test Legipid Legionella Fast Detection is equivalent to the reference culture method for the enumeration of Legionella spp.

  18. Reduction of Legionella spp. in water and in soil by a citrus plant extract vapor.

    Science.gov (United States)

    Laird, Katie; Kurzbach, Elena; Score, Jodie; Tejpal, Jyoti; Chi Tangyie, George; Phillips, Carol

    2014-10-01

    Legionnaires' disease is a severe form of pneumonia caused by Legionella spp., organisms often isolated from environmental sources, including soil and water. Legionella spp. are capable of replicating intracellularly within free-living protozoa, and once this has occurred, Legionella is particularly resistant to disinfectants. Citrus essential oil (EO) vapors are effective antimicrobials against a range of microorganisms, with reductions of 5 log cells ml(-1) on a variety of surfaces. The aim of this investigation was to assess the efficacy of a citrus EO vapor against Legionella spp. in water and in soil systems. Reductions of viable cells of Legionella pneumophila, Legionella longbeachae, Legionella bozemanii, and an intra-amoebal culture of Legionella pneumophila (water system only) were assessed in soil and in water after exposure to a citrus EO vapor at concentrations ranging from 3.75 mg/liter air to 15g/liter air. Antimicrobial efficacy via different delivery systems (passive and active sintering of the vapor) was determined in water, and gas chromatography-mass spectrometry (GC-MS) analysis of the antimicrobial components (linalool, citral, and β-pinene) was conducted. There was up to a 5-log cells ml(-1) reduction in Legionella spp. in soil after exposure to the citrus EO vapors (15 mg/liter air). The most susceptible strain in water was L. pneumophila, with a 4-log cells ml(-1) reduction after 24 h via sintering (15 g/liter air). Sintering the vapor through water increased the presence of the antimicrobial components, with a 61% increase of linalool. Therefore, the appropriate method of delivery of an antimicrobial citrus EO vapor may go some way in controlling Legionella spp. from environmental sources. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  19. Legionella Persistence in Manufactured Water Systems: Pasteurization Potentially Selecting for Thermal Tolerance

    Directory of Open Access Journals (Sweden)

    Harriet Whiley

    2017-07-01

    Full Text Available Legionella is an opportunistic waterborne pathogen of increasing public health significance. Pasteurization, otherwise known as super-heat and flush (increasing water temperature to above 70°C and flushing all outlets, has been identified as an important mechanism for the disinfection of Legionella in manufactured water systems. However, several studies have reported that this procedure was ineffective at remediating water distribution systems as Legionella was able to maintain long term persistent contamination. Up to 25% of L. pneumophila cells survived heat treatment of 70°C, but all of these were in a viable but non-culturable state. This demonstrates the limitations of the culture method of Legionella detection currently used to evaluate disinfection protocols. In addition, it has been demonstrated that pasteurization and nutrient starvation can select for thermal tolerant strains, where L. pneumophila was consistently identified as having greater thermal tolerance compared to other Legionella species. This review demonstrates that further research is needed to investigate the effectiveness of pasteurization as a disinfection method. In particular, it focuses on the potential for pasteurization to select for thermal tolerant L. pneumophila strains which, as the primary causative agent of Legionnaires disease, have greater public health significance compared to other Legionella species.

  20. Detection of airborne Legionella while showering using liquid impingement and fluorescent in situ hybridization (FISH).

    Science.gov (United States)

    Deloge-Abarkan, Magali; Ha, Thi-Lan; Robine, Enric; Zmirou-Navier, Denis; Mathieu, Laurence

    2007-01-01

    Aerosols of water contaminated with Legionella bacteria constitute the only mode of exposure for humans. However, the prevention strategy against this pathogenic bacteria risk is managed through the survey of water contamination. No relationship linked the Legionella bacteria water concentration and their airborne abundance. Therefore, new approaches in the field of the metrological aspects of Legionella bioaerosols are required. This study was aimed at testing the main principles for bioaerosol collection (solid impaction, liquid impingement and filtration) and the in situ hybridization (FISH) method, both in laboratory and field assays, with the intention of applying such methodologies for airborne Legionella bacteria detection while showering. An aerosolization chamber was developed to generate controlled and reproducible L. pneumophila aerosols. This tool allowed the identification of the liquid impingement method as the most appropriate one for collecting airborne Legionella bacteria. The culturable fraction of airborne L. pneumophila recovered with the liquid impingement principle was 4 and 700 times higher compared to the impaction and filtration techniques, respectively. Moreover, the concentrations of airborne L. pneumophila in the impinger fluid were on average 7.0 x 10(5) FISH-cells m(-3) air with the fluorescent in situ hybridization (FISH) method versus 9.0 x 10(4) CFU m(-3) air with the culture method. These results, recorded under well-controlled conditions, were confirmed during the field experiments performed on aerosols generated by hot water showers in health institutions. This new approach may provide a more accurate characterization of aerobiocontamination by Legionella bacteria.

  1. A preliminary assessment of the occupational risk of acquiring Legionnaires' disease for people working in telephone manholes, a new workplace environment for Legionella growth.

    Science.gov (United States)

    Ricci, Maria Luisa; Fontana, Stefano; Bella, Antonino; Gaggioli, A; Cascella, R; Cassone, Antonio; Scaturro, Maria

    2010-09-01

    Telephone manholes (TMs) are underground wells, used in Italy by the telecommunication companies to locate telephone networks. Following a fatal case of Legionnaires' disease (LD), acquired during working activity in a TM, we investigated whether Legionella was present in TMs and could be a risk for manhole workers (MWs). Three hundred fifty-three environmental samples were collected from 100 TMs, and serum antibody titer against Legionella pneumophila and Legionella species non-pneumophila was determined from both MWs and control non-manhole workers. L pneumophila and Legionella species non-pneumophila were detected in 28% of water samples, in 8% of the biofilm, and in 6% of sediment matrices taken in TMs, in a concentration range of 10(2) to 10(4) colony-forming units/L. No Legionella was found in TM air samples. Although there was a statistically significant difference (P = .027) in antibody titer to L pneumophila serogroup 1 (Lp1) between MWs and non-manhole workers, a multivariate logistic regression analysis showed a significant association between antibody against Lp1 and both age group and the practice of aquatic sports. Although further investigations will be performed to quantify the risk of acquiring legionellosis, this preliminary study demonstrates for the first time the presence of Legionella, including human pathogenic species, in a working environment such as TM.

  2. A multicenter evaluation of the Biotest legionella urinary antigen EIA

    DEFF Research Database (Denmark)

    Harrison, Timothy; Uldum, Søren; Alexiou-Daniel, Stella

    1998-01-01

    OBJECTIVES: To undertake a multicenter study to evaluate the Biotest legionella urinary antigen enzyme immunoassay (EIA) performance against those EIAs already in use in 14 European laboratories. METHODS: Each laboratory examined urine specimens from appropriate patients using both their current...... in the laboratories' current EIAs, and in 94.6% of those specimens which were positive for Legionella pneumophila serogroup 1. CONCLUSION: The Biotest EIA is simple to use and specific and the results obtained in different laboratories show excellent agreement. The assay compares well existing EIAs, at least for L...

  3. Detection of Legionella by cultivation and quantitative real-time polymerase chain reaction in biological waste water treatment plants in Norway.

    Science.gov (United States)

    Lund, Vidar; Fonahn, Wenche; Pettersen, Jens Erik; Caugant, Dominique A; Ask, Eirik; Nysaeter, Ase

    2014-09-01

    Cases of Legionnaires' disease associated with biological treatment plants (BTPs) have been reported in six countries between 1997 and 2010. However, knowledge about the occurrence of Legionella in BTPs is scarce. Hence, we undertook a qualitative and quantitative screening for Legionella in BTPs treating waste water from municipalities and industries in Norway, to assess the transmission potential of Legionella from these installations. Thirty-three plants from different industries were sampled four times within 1 year. By cultivation, 21 (16%) of 130 analyses were positive for Legionella species and 12 (9%) of 130 analyses were positive for Legionella pneumophila. By quantitative real-time polymerase chain reaction (PCR), 433 (99%) of 437 analyses were positive for Legionella species and 218 (46%) of 470 analyses were positive for L. pneumophila. This survey indicates that PCR could be the preferable method for detection of Legionella in samples from BTPs. Sequence types of L. pneumophila associated with outbreaks in Norway were not identified from the BTPs. We showed that a waste water treatment plant with an aeration basin can produce high concentrations of Legionella. Therefore, these plants should be considered as a possible source of community-acquired Legionella infections.

  4. Legionella - A threat to groundwater: Pathogen transport in recharge basin.

    Science.gov (United States)

    McBurnett, Lauren R; Holt, Nathan T; Alum, Absar; Abbaszadegan, Morteza

    2018-04-15

    This study elucidates the potential risk posed by Legionella during aquifer recharge practices. Experiments were conducted using pilot-scale column simulating infiltration of bacterial surrogate and pathogen, E. coli and Legionella pneumophila, under central Arizona recharge basin conditions. A column was packed with a loamy sand media collected from a recharge basin and was fitted with six sampling ports at soil depths of 15, 30, 60, 92, 122cm and acclimated for a month with tertiary treated wastewater. Transport of Legionella appeared to be delayed compared to E. coli. The breakthrough of E. coli and Legionella at 122cm depth occurred at 3 and 24h, respectively. Slow transport of Legionella is consistent with its pleomorphic nature and variation in size and shape under low nutrient conditions. Legionella persisted for a longer time in the column, but at lower concentrations. Given the novel results of this study, the transport of Legionella into groundwater aquifers can occur through engineering recharge basin conditions creating a potential public health risk. Copyright © 2017. Published by Elsevier B.V.

  5. Legionella detection and subgrouping in water air-conditioning cooling tower systems in Kuwait.

    Science.gov (United States)

    Al-Matawah, Qadreyah; Al-Zenki, Sameer; Al-Azmi, Ahmad; Al-Waalan, Tahani; Al-Salameen, Fadila; Hejji, Ahmad Ben

    2015-07-01

    The main aim of the study was to test for the presence of Legionnaires' disease-causing microorganisms in air-conditioned buildings in Kuwait using molecular technologies. For this purpose, 547 samples were collected from 38 cooling towers for the analysis of Legionella pneumophila. These samples included those from water (n = 178), air (n = 231), and swabs (n = 138). Out of the 547 samples, 226 (41%) samples were presumptive positive for L. pneumophila, with L. pneumophila viable counts in the positive water samples ranging from 1 to 88 CFU/ml. Of the Legionella culture-positive samples, 204 isolates were examined by latex agglutination. These isolates were predominately identified as L. pneumophila serogroup (sg) 2-14. Using the Dresden panel of monoclonal antibodies, 74 representatives isolates were further serogrouped. Results showed that 51% of the isolates belonged to serogroup 7 followed by 1 (18%) and 3 (18%). Serogroups 4 (4%) and 10 (7%) were isolated at a lower frequency, and two isolates could not be assigned to a serogroup. These results indicate the wide prevalence of L. pneumophila serogroup 7 as the predominant serogroup at the selected sampling sites. Furthermore, the 74 L. pneumophila (sg1 = 13; sg3 = 13; sg4 = 3; sg7 = 38; sg10 = 5; sgX = 2) isolates were genotyped using the seven gene protocol sequence-based typing (SBT) scheme developed by the European Working Group for Legionella Infections (EWGLI). The results show that Legionella isolates were discriminated into nine distinct sequence typing (ST) profiles, five of which were new to the SBT database of EWGLI. Additionally, all of the ST1 serogroup 1 isolates were of the OLDA/Oxford subgroup. These baseline data will form the basis for the development of a Legionella environmental surveillance program and used for future epidemiological investigations.

  6. Characterization of a Legionella micdadei mip mutant

    DEFF Research Database (Denmark)

    O'Connell, W A; Bangsborg, Jette Marie; Cianciotto, N P

    1995-01-01

    The pathogenesis of Legionella micdadei is dependent upon its ability to infect alveolar phagocytes. To better understand the basis of intracellular infection by this organism, we examined the importance of its Mip surface protein. In Legionella pneumophila, Mip promotes infection of both human m...... Mip. Although unimpaired in its ability to grow in bacteriologic media, this Mip mutant was defective in its capacity to infect U937 cells, a human macrophage-like cell line. Most significantly, the Mip- organism displayed a 24-fold reduction in survivability immediately after its entry...... into the phagocyte. Similarly, the mutant was less able to parasitize Hartmannella amoebae. Taken together, these data argue that Mip specifically potentiates intracellular growth by L. micdadei....

  7. Effects of Disinfection on Legionella spp., Eukarya, and Biofilms in a Hot Water System

    Science.gov (United States)

    Moletta-Denat, Marina; Frère, Jacques; Onillon, Séverine; Trouilhé, Marie-Cécile; Robine, Enric

    2012-01-01

    Legionella species are frequently detected in hot water systems, attached to the surface as a biofilm. In this work, the dynamics of Legionella spp. and diverse bacteria and eukarya associated together in the biofilm, coming from a pilot scale 1 system simulating a real hot water system, were investigated throughout 6 months after two successive heat shock treatments followed by three successive chemical treatments. Community structure was assessed by a fingerprint technique, single-strand conformation polymorphism (SSCP). In addition, the diversity and dynamics of Legionella and eukarya were investigated by small-subunit (SSU) ribosomal cloning and sequencing. Our results showed that pathogenic Legionella species remained after the heat shock and chemical treatments (Legionella pneumophila and Legionella anisa, respectively). The biofilm was not removed, and the bacterial community structure was transitorily affected by the treatments. Moreover, several amoebae had been detected in the biofilm before treatments (Thecamoebae sp., Vannella sp., and Hartmanella vermiformis) and after the first heat shock treatment, but only H. vermiformis remained. However, another protozoan affiliated with Alveolata, which is known as a host cell for Legionella, dominated the eukaryal species after the second heat shock and chemical treatment tests. Therefore, effective Legionella disinfection may be dependent on the elimination of these important microbial components. We suggest that eradicating Legionella in hot water networks requires better study of bacterial and eukaryal species associated with Legionella in biofilms. PMID:22820326

  8. Seasonal variation of Legionella in Taiwan's reservoir and its relationships with environmental factors.

    Science.gov (United States)

    Kao, Po-Min; Hsu, Bing-Mu; Chang, Tien-Yu; Hsu, Tsui-Kang; Tzeng, Kai-Jiun; Huang, Yu-Li

    2015-04-01

    In this study, the presence of Legionella in major water reservoirs of Taiwan was examined with respect to seasonal variation, geographical variation, and water quality parameters using TaqMan real-time qPCR. Water samples were collected quarterly at 19 reservoirs in Taiwan between November 2012 and August 2013. The detection rate for Legionella was 35.5% (27/76), and Legionella was detected in all seasons. The Legionella concentration was relatively high in spring and summer, reaching 3.86 × 10(8) and 7.35 × 10(8) cells/L, respectively. By sampling the area, Legionella was detected at a higher proportion in reservoirs in the northern and southern areas, and the difference was consistent in all seasons. Significant association was found between detection of Legionella and various water quality parameters, including conductivity, chlorophyll a, and dissolved oxygen (Mann-Whitney U test, P Legionella detection with pH (P = 0.030, R = -0.497) and dissolved oxygen (P = 0.007, R = -0.596) in fall and positive correlation with Carlson's trophic state index (P = 0.049, R = 0.457) in spring. The identified species included Legionella pneumophila and Legionella drancourtii. The detection of Legionella in reservoirs was indicative of a potential public health risk and should be further evaluated.

  9. Legionella on board trains: effectiveness of environmental surveillance and decontamination.

    Science.gov (United States)

    Quaranta, Gianluigi; Vincenti, Sara; Ferriero, Anna Maria; Boninti, Federica; Sezzatini, Romina; Turnaturi, Cinzia; Gliubizzi, Maria Daniela; Munafò, Elio; Ceccarelli, Gianluca; Causarano, Carmelo; Accorsi, Massimo; Del Nord, Pasquale; Ricciardi, Walter; Laurenti, Patrizia

    2012-08-07

    Legionella pneumophila is increasingly recognised as a significant cause of sporadic and epidemic community-acquired and nosocomial pneumonia. Many studies describe the frequency and severity of Legionella spp. contamination in spa pools, natural pools, hotels and ships, but there is no study analysing the environmental monitoring of Legionella on board trains. The aims of the present study were to conduct periodic and precise environmental surveillance of Legionella spp. in water systems and water tanks that supply the toilet systems on trains, to assess the degree of contamination of such structures and to determine the effectiveness of decontamination. A comparative pre-post ecological study was conducted from September 2006 to January 2011. A total of 1,245 water samples were collected from plumbing and toilet water tanks on passenger trains. The prevalence proportion of all positive samples was calculated. The unpaired t-test was performed to evaluate statistically significant differences between the mean load values before and after the decontamination procedures; statistical significance was set at p ≤ 0.05. In the pre-decontamination period, 58% of the water samples were positive for Legionella. Only Legionella pneumophila was identified: 55.84% were serogroup 1, 19.03% were serogroups 2-14 and 25.13% contained both serogroups. The mean bacterial load value was 2.14 × 10(3) CFU/L. During the post-decontamination period, 42.75% of water samples were positive for Legionella spp.; 98.76% were positive for Legionella pneumophila: 74.06% contained serogroup 1, 16.32% contained serogroups 2-14 and 9.62% contained both. The mean bacterial load in the post-decontamination period was 1.72 × 10(3) CFU/L. According to the t-test, there was a statistically significant decrease in total bacterial load until approximately one and a half year after beginning the decontamination programme (p = 0.0097). This study indicates that systematic environmental surveillance

  10. Legionella on board trains: effectiveness of environmental surveillance and decontamination

    Directory of Open Access Journals (Sweden)

    Quaranta Gianluigi

    2012-08-01

    Full Text Available Abstract Background Legionella pneumophila is increasingly recognised as a significant cause of sporadic and epidemic community-acquired and nosocomial pneumonia. Many studies describe the frequency and severity of Legionella spp. contamination in spa pools, natural pools, hotels and ships, but there is no study analysing the environmental monitoring of Legionella on board trains. The aims of the present study were to conduct periodic and precise environmental surveillance of Legionella spp. in water systems and water tanks that supply the toilet systems on trains, to assess the degree of contamination of such structures and to determine the effectiveness of decontamination. Methods A comparative pre-post ecological study was conducted from September 2006 to January 2011. A total of 1,245 water samples were collected from plumbing and toilet water tanks on passenger trains. The prevalence proportion of all positive samples was calculated. The unpaired t-test was performed to evaluate statistically significant differences between the mean load values before and after the decontamination procedures; statistical significance was set at p ≤ 0.05. Results In the pre-decontamination period, 58% of the water samples were positive for Legionella. Only Legionella pneumophila was identified: 55.84% were serogroup 1, 19.03% were serogroups 2–14 and 25.13% contained both serogroups. The mean bacterial load value was 2.14 × 103 CFU/L. During the post-decontamination period, 42.75% of water samples were positive for Legionella spp.; 98.76% were positive for Legionella pneumophila: 74.06% contained serogroup 1, 16.32% contained serogroups 2–14 and 9.62% contained both. The mean bacterial load in the post-decontamination period was 1.72 × 103 CFU/L. According to the t-test, there was a statistically significant decrease in total bacterial load until approximately one and a half year after beginning the decontamination programme (p

  11. Legionella risk assessment in cruise ships and ferries.

    Science.gov (United States)

    Laganà, Pasqualina; Gambuzza, Maria Elsa; Delia, Santi

    2017-06-12

    Introduction. The increasing development of marine traffic has led to a rise in the incidence of legionellosis among travellers. It occurs in similar environments, especially closed and crowded, and aboard ships Legionella survives and multiplies easily in water pipes, spreading into the environment through air conditioning systems and water distribution points. Although in recent years in the construction of cruise ships preventive measures aimed at curbing the proliferation of Legionella (design, materials, focus on the operation and maintenance of the water system), have been taken account, little or no attention has been paid to small ships which, in many cases, are old and not well maintained. Objective. The aim of the study was to evaluate the frequency and severity of Legionella contamination in ferries and cruise ships in order to adopt more specific control measures. Materials and method. A prevalence study was carried out on 10 ferries and 6 cruise ships docking or in transit across the port of Messina (Sicily, Italy). Water and air samples collected from many critical points were tested for qualitative and quantitative identification of Legionella. Results and conclusions. Legionella pneumophila sg 1 was isolated from the samples of shower and tap water in 7 (70%) of the 10 ferries examined, and in 3 (33%) of the 6 cruise ships examined, and L. pneumophila sg 2-14 in 8 (80%) and 1 (16.7%) of these ships, respectively. No Legionella contamination was found in whirlpool baths, air and ice samples. In conclusion, the data obtained confirm higher levels of Legionella contamination in local ferries and cruise ships, underlining the need to adopt corrective actions more specific for these smaller vessels.

  12. Legionella pnömonisini taklit eden malignite olgusu

    OpenAIRE

    Ali Karakuş; Ersin Şükrü Erden; Cenk Babayiğit; Eyüp Büyükkaya; Mehmet Mustafa Akın; Muhammet Murat Çelik; Veyis Taşın

    2013-01-01

    Legionella pneumophila is a bacterium, which can grow inwater pipe networks and climate systems. Contaminationoccurs by aspiration of infected water or aerosol inhalation.It is usually presented with fever, bradycardia, andchange in mental status, hyponatremia, elevation of liverenzymes and deterioration of renal function. The definitediagnosis is established by detection of the antigens andcultivating in the culture medium. Also, malign lung tumorscan encounter with the same clinical finding...

  13. Riesgos por Legionella, Prevención y Control

    OpenAIRE

    Martín Romero, Magdalena

    2014-01-01

    Para un buen control de riesgos por Legionella pneumophila, es importante conocer con detalle esta bacteria, los efectos que causa en el hombre, las principales fuentes de contagio y los medios para evitarlo. Este es el objetivo del presente trabajo. La legionelosis a pesar de ser percibida como una enfermedad infecciosa potencialmente erradicable, se puede controlar con medidas higiénico- sanitarias en las instalaciones implicadas.

  14. Legionella species diversity and dynamics from surface reservoir to tap water: from cold adaptation to thermophily.

    Science.gov (United States)

    Lesnik, René; Brettar, Ingrid; Höfle, Manfred G

    2016-05-01

    Water samples of the Drinking Water Supply System (DWSS) of the city of Braunschweig were analysed for its Legionella species composition using genus-specific PCR amplicons and single-strand conformation polymorphism (SSCP) fingerprint analyses based on 16S rRNA genes. These analyses comprised the whole supply chain including raw water, treatment process and large-scale storage, and a seasonal study of finished drinking water sampled monthly from cold and hot tap water. Treatment of raw water had a major impact on Legionella species by reducing their diversity and abundances. The Legionella species composition of the tap water was highly distinct from that of both source waters. In cold water, 8-14 different phylotypes of Legionella (PTLs) were observed per sample with relative abundances ranging from >1% to 53%. In hot water, L. pneumophila was present during all seasons at high relative abundances (8-40%) accompanied by 5-14 other PTLs of which 6 PTLs were in common with cold water. This thermophilic Legionella community, including L. pneumophila, was able to grow in the hot water above 50 °C. Such thermophilic Legionella populations are of general relevance for drinking water management and public health, but also for the ecology and evolution of the genus Legionella.

  15. Tracking Legionella in air generated from a biological treatment plant: a case study of the outbreak of legionellosis in Norway

    Science.gov (United States)

    Blatny, Janet M.; Olsen, Jaran S.; Andreassen, Øyvind; Waagen, Viggo; Reif, Bjørn Anders P.

    2011-05-01

    Two outbreaks of legionellosis occurred in the Sarpsborg/Fredrikstad region southeast of Norway in 2005 and 2008 where more than 60 exposed individuals were infected and 10 case patients died. The air scrubber at Borregaard, a wood-based chemical factory, was identified as the outbreak source. High concentration levels of Legionella species, including the etiological agent L. pneumophila SG1 was found in the aeration ponds, which belongs to Borregaard's biological treatment plant. Results showed that these ponds were able to generate Legionella-containing aerosols that were transported by the wind as such aerosols were measured up to 200 meters downwind of the pond. Our studies did not detect L. pneumophila SG1 isolates, only L. pneumophila SG4 during the air sampling measurement campaign. Furthermore, the operational conditions of the air scrubber proved to be harsh for Legionella growth as the outbreak L. pneumophila strains were not able to grow at 45ºC and pH8 (conditions during the outbreaks). These results, together, lead us to suggest that the aeration pond should be regarded as the primary amplifier and disseminator of Legionella and L. pneumophila and thereby most likely being the outbreak source.

  16. [Presence of Legionella spp. in household drinking water reservoirs in Resistencia, Chaco, Argentina. Preliminary report].

    Science.gov (United States)

    Lösch, Liliana S; Merino, Luis A

    Legionella spp. is an environmental bacterium that can survive in a wide range of physicochemical conditions and may colonize distribution systems of drinking water and storage tanks. Legionella pneumophila is the major waterborne pathogen that can cause 90% of Legionnaires' disease cases. The aim of this study was to detect the presence of Legionella spp. in household drinking water tanks in the city of Resistencia, Chaco. The detection of Legionella in water samples was performed by culture methods as set out in ISO 11731:1998. Thirty two water samples were analyzed and Legionella spp. was recovered in 12 (37.5%) of them. The monitoring of this microorganism in drinking water is the first step towards addressing the control of its spread to susceptible hosts. Copyright © 2016 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

  17. Automobile windshield washer fluid: A potential source of transmission for Legionella

    Energy Technology Data Exchange (ETDEWEB)

    Schwake, David Otto [School of Life Sciences, Arizona State University, P.O. Box 875306, Tempe, AZ 85287 (United States); Alum, Absar [School of Sustainable Engineering and the Built Environment, Arizona State University, P.O. Box 875306, Tempe, AZ 85287 (United States); Abbaszadegan, Morteza, E-mail: abbaszadegan@asu.edu [School of Sustainable Engineering and the Built Environment, Arizona State University, P.O. Box 875306, Tempe, AZ 85287 (United States)

    2015-09-01

    Epidemiological evidence suggesting driving cars to be a risk factor for legionellosis has prompted public health studies to investigate vehicle windshield washer fluid as a novel transmission source of this disease. The goal of the current study was to investigate whether or not windshield washer fluid could serve as a potential source of transmission for Legionella. A wide variation in the survival of L. pneumophila was observed when incubated in different washer fluids at 25 and 37 °C, however, one brand tested supported Legionella survival similar to or greater than sterilized deionized water. In addition, 1 L of tap water contained in a washer fluid reservoir was able to support population growth and survival of Legionella for several months. In a field study examining the windshield washer fluid of 12 elementary school buses, Legionella were detected from 84% of samples at a high concentration of 8.1 × 10{sup 4} CFU/mL. Culturable cells were also detected in aerosolized washer fluid during washer fluid spray. By demonstrating survival in certain windshield washer fluids, growth within washer fluid reservoirs, and the presence of viable cells in bus washer fluid spray, we have provided evidence suggesting the potential for a novel route of Legionella exposure. - Highlights: • L. pneumophila survival in one brand of washer fluid and sterilized deionized water were similar. • Legionella population was maintained in tap water for several months. • Culturable Legionella were detected in 10 of 12 school bus washer fluid reservoirs sampled. • Legionella concentrations up to 8.1 × 104 CFU/mL were detected in school bus washer fluid. • L. pneumophila was detected in washer fluid reservoirs and aerosolized washer fluid.

  18. High prevalence of Legionella in non-passenger merchant vessels.

    Science.gov (United States)

    Collins, S L; Stevenson, D; Mentasti, M; Shaw, A; Johnson, A; Crossley, L; Willis, C

    2017-03-01

    There is a paucity of information on the risk from potable water in non-passenger merchant vessels (NPMVs) particularly with regard to Legionella and other bacteria. This retrospective study examined water samples from 550 NPMVs docked in eight UK ports. A total of 1027 samples from 412 NPMVs were examined for total aerobic colony counts (ACC), coliforms, Escherichia coli and enterococci; 41% of samples yielded ACC above the action level (>1 × 103 c.f.u./ml) and 4·5% contained actionable levels (>1 c.f.u./100 ml) of faecal indicator bacteria. Eight hundred and three samples from 360 NPMVs were cultured specifically for Legionella and 58% of vessels proved positive for these organisms with 27% of samples showing levels greater than the UK upper action limit of 1 × 103 c.f.u./l. Cabin showers (49%) and hospital shower (45%) were frequently positive. A subset of 106 samples was analysed by quantitative polymerase chain reaction for Legionella and identified a further 11 Legionella-positive NPMVs, returning a negative predictive value of 100%. There was no correlation between NPMV age or size and any microbial parameters (P > 0·05). Legionella pneumophila serogroup 1 was isolated from 46% of NPMVs and sequence-based typing of 17 isolates revealed four sequence types (STs) previously associated with human disease. These data raise significant concerns regarding the management of microbial and Legionella risks on board NPMVs and suggest that better guidance and compliance are required to improve control.

  19. Minimum inhibitory concentration distribution in environmental Legionella spp. isolates.

    Science.gov (United States)

    Sandalakis, Vassilios; Chochlakis, Dimosthenis; Goniotakis, Ioannis; Tselentis, Yannis; Psaroulaki, Anna

    2014-12-01

    In Greece standard tests are performed in the watering and cooling systems of hotels' units either as part of the surveillance scheme or following human infection. The purpose of this study was to establish the minimum inhibitory concentration (MIC) distributions of environmental Legionella isolates for six antimicrobials commonly used for the treatment of Legionella infections, by MIC-test methodology. Water samples were collected from 2004 to 2011 from 124 hotels from the four prefectures of Crete (Greece). Sixty-eight (68) Legionella isolates, comprising L. pneumophila serogroups 1, 2, 3, 5, 6, 8, 12, 13, 15, L. anisa, L. rubrilucens, L. maceachernii, L. quinlivanii, L. oakridgensis, and L. taurinensis, were included in the study. MIC-tests were performed on buffered charcoal yeast extract with α-ketoglutarate, L-cysteine, and ferric pyrophosphate. The MICs were read after 2 days of incubation at 36 ± 1 °C at 2.5% CO2. A large distribution in MICs was recorded for each species and each antibiotic tested. Rifampicin proved to be the most potent antibiotic regardless of the Legionella spp.; tetracycline appeared to have the least activity on our environmental isolates. The MIC-test approach is an easy, although not so cost-effective, way to determine MICs in Legionella spp. These data should be kept in mind especially since these Legionella species may cause human disease.

  20. Strategies for the reduction of Legionella in biological treatment systems.

    Science.gov (United States)

    Nogueira, R; Utecht, K-U; Exner, M; Verstraete, W; Rosenwinkel, K-H

    A community-wide outbreak of Legionnaire's disease occurred in Warstein, Germany, in August 2013. The epidemic strain, Legionella pneumophila Serogruppe 1, was isolated from an industrial wastewater stream entering the municipal wastewater treatment plant (WWTP) in Wartein, the WWTP itself, the river Wäster and air/water samples from an industrial cooling system 3 km downstream of the WWTP. The present study investigated the effect of physical-chemical disinfection methods on the reduction of the concentration of Legionella in the biological treatment and in the treated effluent entering the river Wäster. Additionally, to gain insight into the factors that promote the growth of Legionella in biological systems, growth experiments were made with different substrates and temperatures. The dosage rates of silver micro-particles, hydrogen peroxide, chlorine dioxide and ozone and pH stress to the activated sludge were not able to decrease the number of culturable Legionella spp. in the effluent. Nevertheless, the UV treatment of secondary treated effluent reduced Legionella spp. on average by 1.6-3.4 log units. Laboratory-scale experiments and full-scale measurements suggested that the aerobic treatment of warm wastewater (30-35 °C) rich in organic nitrogen (protein) is a possible source of Legionella infection.

  1. Occurrence and Control of Legionella in Recycled Water Systems

    Directory of Open Access Journals (Sweden)

    Patrick K. Jjemba

    2015-07-01

    Full Text Available Legionella pneumophila is on the United States Environmental Protection Agency (USEPA Candidate Contaminant list (CCL as an important pathogen. It is commonly encountered in recycled water and is typically associated with amoeba, notably Naegleria fowleri (also on the CCL and Acanthamoeba sp. No legionellosis outbreak has been linked to recycled water and it is important for the industry to proactively keep things that way. A review was conducted examine the occurrence of Legionella and its protozoa symbionts in recycled water with the aim of developing a risk management strategy. The review considered the intricate ecological relationships between Legionella and protozoa, methods for detecting both symbionts, and the efficacy of various disinfectants.

  2. Approach to determine the diversity of Legionella species by nested PCR-DGGE in aquatic environments.

    Science.gov (United States)

    Huang, Wen-Chien; Tsai, Hsin-Chi; Tao, Chi-Wei; Chen, Jung-Sheng; Shih, Yi-Jia; Kao, Po-Min; Huang, Tung-Yi; Hsu, Bing-Mu

    2017-01-01

    In this study, we describe a nested PCR-DGGE strategy to detect Legionella communities from river water samples. The nearly full-length 16S rRNA gene was amplified using bacterial primer in the first step. After, the amplicons were employed as DNA templates in the second PCR using Legionella specific primer. The third round of gene amplification was conducted to gain PCR fragments apposite for DGGE analysis. Then the total numbers of amplified genes were observed in DGGE bands of products gained with primers specific for the diversity of Legionella species. The DGGE patterns are thus potential for a high-throughput preliminary determination of aquatic environmental Legionella species before sequencing. Comparative DNA sequence analysis of excised DGGE unique band patterns showed the identity of the Legionella community members, including a reference profile with two pathogenic species of Legionella strains. In addition, only members of Legionella pneumophila and uncultured Legionella sp. were detected. Development of three step nested PCR-DGGE tactic is seen as a useful method for studying the diversity of Legionella community. The method is rapid and provided sequence information for phylogenetic analysis.

  3. Legionella bacteria in combustion air humidifiers; Legionella i luftuppfuktare foer foerbraenningsluft

    Energy Technology Data Exchange (ETDEWEB)

    Jeppesen, Jessica; Hansson, Helen; Cederfeldt, Ola; Axby, Fredrik

    2007-10-15

    had positive test results for legionella, three of them contained Legionella pneumophila serogroup 1. In the second test round, two of the plants had positive test results but only one of them was verified to L. pneumophila sg 1. Both the fuel gas analyses showed negative results for legionella bacteria and the level of heterotrophic bacteria was low or nonexistent. In this report the occurrence of legionella bacteria in combustion air humidifiers has mainly been treated as a working environmental risk. A risk assessment for the plant with the highest amount of legionella bacteria shows that the growth of legionella primary should be minimized/eliminated. As a compliment, personal safety devices such as respiratory protection should be used during revision work in water tanks or while cleaning the water system, i.e. when the risk for water aerosols is greater than during normal operation of the plant. The employer is responsible to inform all personal who might become infected with legionella. There should also be routines for reporting as well as routines for both proactive and reactive measures. From the test results the conclusion that legionella bacteria occur in combustion air humidifiers can be drawn. In addition legionella bacteria can occur in all three studied types of air humidifiers; rotating-, lamella- and scrubber air humidifiers. It is difficult to predict how great a risk it is that legionella bacteria will occur in the different types of air humidifiers. The risk of legionella bacteria transferring from the water systems to outgoing fuel gas varies in the different types of air humidifiers. It is obvious that the risk of transferring legionella bacteria to the fuel gas is greater in rotating air humidifiers and in lamella air humidifiers where both fuel gas and air is washed with water from the same water system. There is also a risk of transferring legionella bacteria to fuel gas in scrubber air humidifiers where the air humidifier and the condenser

  4. Legionella-protozoa-nematode interactions in aquatic biofilms and influence of Mip on Caenorhabditis elegans colonization.

    Science.gov (United States)

    Rasch, Janine; Krüger, Stefanie; Fontvieille, Dominique; Ünal, Can M; Michel, Rolf; Labrosse, Aurélie; Steinert, Michael

    2016-09-01

    Legionella pneumophila, the causative agent of Legionnaireś disease, is naturally found in aquatic habitats. The intracellular life cycle within protozoa pre-adapted the "accidental" human pathogen to also infect human professional phagocytes like alveolar macrophages. Previous studies employing the model organism Caenorhabditis elegans suggest that also nematodes might serve as a natural host for L. pneumophila. Here, we report for the first time from a natural co-habitation of L. pneumophila and environmental nematode species within biofilms of a warm water spring. In addition, we identified the protozoan species Oxytricha bifaria, Stylonychia mytilus, Ciliophrya sp. which have never been described as potential interaction partners of L. pneumophila before. Modeling and dissection of the Legionella-protozoa-nematode interaction revealed that C. elegans ruptures Legionella-infected amoebal cells and by this means incorporate the pathogen. Further infection studies revealed that the macrophage infectivity potentiator (Mip) protein of L. pneumophila, which is known to bind collagen IV during human lung infection, promotes the colonization of the intestinal tract of L4 larvae of C. elegans and negatively influences the life span of the worms. The Mip-negative L. pneumophila mutant exhibited a 32-fold reduced colonization rate of the nematodes after 48h when compared to the wild-type strain. Taken together, these studies suggest that nematodes may serve as natural hosts for L. pneumophila, promote their persistence and dissemination in the environment, and co-evolutionarily pre-adapt the pathogen for interactions with extracellular constituents of human lung tissue. Copyright © 2016 Elsevier GmbH. All rights reserved.

  5. Effect of UV irradiation (253.7 nm) on free Legionella and Legionella associated with its amoebae hosts.

    Science.gov (United States)

    Cervero-Aragó, Sílvia; Sommer, Regina; Araujo, Rosa M

    2014-12-15

    Water systems are the primary reservoir for Legionella spp., where the bacteria live in association with other microorganisms, such as free-living amoebae. A wide range of disinfection treatments have been studied to control and prevent Legionella colonization but few of them were performed considering its relation with protozoa. In this study, the effectiveness of UV irradiation (253.7 nm) using low-pressure lamps was investigated as a disinfection method for Legionella and amoebae under controlled laboratory conditions. UV treatments were applied to 5 strains of Legionella spp., 4 strains of free-living amoeba of the genera Acanthamoeba and Vermamoeba, treating separately trophozoites and cysts, and to two different co-cultures of Legionella pneumophila with the Acanthamoeba strains. No significant differences in the UV inactivation behavior were observed among Legionella strains tested which were 3 logs reduced for fluences around 45 J/m(2). UV irradiation was less effective against free-living amoebae; which in some cases required up to 990 J/m(2) to obtain the same population reduction. UV treatment was more effective against trophozoites compared to cysts; moreover, inactivation patterns were clearly different between the genus Acanthamoeba and Vermamoeba. For the first time data about Vermamoeba vermiformis UV inactivation has been reported in a study. Finally, the results showed that the association of L. pneumophila with free-living amoebae decreases the effectiveness of UV irradiation against the bacteria in a range of 1.5-2 fold. That fact demonstrates that the relations established between different microorganisms in the water systems can modify the effectiveness of the UV treatments applied. Copyright © 2014 Elsevier Ltd. All rights reserved.

  6. Technologies for Legionella Control

    Science.gov (United States)

    Presentation includes a review of new Office of Water document entitled "Technologies for Legionella Control in Premise Plumbing Systems", and discussion on ORD research projects involving Legionella and disinfection.

  7. Annual variations and effects of temperature on Legionella spp. and other potential opportunistic pathogens in a bathroom.

    Science.gov (United States)

    Lu, Jingrang; Buse, Helen; Struewing, Ian; Zhao, Amy; Lytle, Darren; Ashbolt, Nicholas

    2017-01-01

    Opportunistic pathogens (OPs) in drinking water, like Legionella spp., mycobacteria, Pseudomonas aeruginosa, and free-living amobae (FLA) are a risk to human health, due to their post-treatment growth in water systems. To assess and manage these risks, it is necessary to understand their variations and environmental conditions for the water routinely used. We sampled premise tap (N cold = 26, N hot = 26) and shower (N shower = 26) waters in a bathroom and compared water temperatures to levels of OPs via qPCR and identified Legionella spp. by 16S ribosomal RNA (rRNA) gene sequencing. The overall occurrence and cell equivalent quantities (CE L-1) of Mycobacterium spp. were highest (100 %, 1.4 × 105), followed by Vermamoeba vermiformis (91 %, 493), Legionella spp. (59 %, 146), P. aeruginosa (14 %, 10), and Acanthamoeba spp. (5 %, 6). There were significant variations of OP's occurrence and quantities, and water temperatures were associated with their variations, especially for Mycobacterium spp., Legionella spp., and V. vermiformis. The peaks observed for Legionella, mainly consisted of Legionella pneumophila sg1 or Legionella anisa, occurred in the temperature ranged from 19 to 49 °C, while Mycobacterium spp. and V. vermiformis not only co-occurred with Legionella spp. but also trended to increase with increasing temperatures. There were higher densities of Mycobacterium in first than second draw water samples, indicating their release from faucet/showerhead biofilm. Legionella spp. were mostly at detectable levels and mainly consisted of L. pneumophila, L. anisa, Legionella donaldsonii, Legionella tunisiensis, and an unknown drinking water isolate based on sequence analysis. Results from this study suggested potential health risks caused by opportunistic pathogens when exposed to warm shower water with low chlorine residue and the use of Mycobacterium spp. as an indicator of premise pipe biofilm and the control management of those potential

  8. Real-time PCR to supplement gold-standard culture-based detection of Legionella in environmental samples.

    Science.gov (United States)

    Collins, S; Jorgensen, F; Willis, C; Walker, J

    2015-10-01

    Culture remains the gold-standard for the enumeration of environmental Legionella. However, it has several drawbacks including long incubation and poor sensitivity, causing delays in response times to outbreaks of Legionnaires' disease. This study aimed to validate real-time PCR assays to quantify Legionella species (ssrA gene), Legionella pneumophila (mip gene) and Leg. pneumophila serogroup-1 (wzm gene) to support culture-based detection in a frontline public health laboratory. Each qPCR assay had 100% specificity, excellent sensitivity (5 GU/reaction) and reproducibility. Comparison of the assays to culture-based enumeration of Legionella from 200 environmental samples showed that they had a negative predictive value of 100%. Thirty eight samples were positive for Legionella species by culture and qPCR. One hundred samples were negative by both methods, whereas 62 samples were negative by culture but positive by qPCR. The average log10 increase between culture and qPCR for Legionella spp. and Leg. pneumophila was 0·72 (P = 0·0002) and 0·51 (P = 0·006), respectively. The qPCR assays can be conducted on the same 1 l water sample as culture thus can be used as a supplementary technique to screen out negative samples and allow more rapid indication of positive samples. The assay could prove informative in public health investigations to identify or rule out sources of Legionella as well as to specifically identify Leg. pneumophila serogroup 1 in a timely manner not possible with culture. © 2015 The Society for Applied Microbiology.

  9. From Many Hosts, One Accidental Pathogen: The Diverse Protozoan Hosts of Legionella

    Directory of Open Access Journals (Sweden)

    David K. Boamah

    2017-11-01

    Full Text Available The 1976 outbreak of Legionnaires' disease led to the discovery of the intracellular bacterial pathogen Legionella pneumophila. Given their impact on human health, Legionella species and the mechanisms responsible for their replication within host cells are often studied in alveolar macrophages, the primary human cell type associated with disease. Despite the potential severity of individual cases of disease, Legionella are not spread from person-to-person. Thus, from the pathogen's perspective, interactions with human cells are accidents of time and space—evolutionary dead ends with no impact on Legionella's long-term survival or pathogenic trajectory. To understand Legionella as a pathogen is to understand its interaction with its natural hosts: the polyphyletic protozoa, a group of unicellular eukaryotes with a staggering amount of evolutionary diversity. While much remains to be understood about these enigmatic hosts, we summarize the current state of knowledge concerning Legionella's natural host range, the diversity of Legionella-protozoa interactions, the factors influencing these interactions, the importance of avoiding the generalization of protozoan-bacterial interactions based on a limited number of model hosts and the central role of protozoa to the biology, evolution, and persistence of Legionella in the environment.

  10. Detection of Legionella spp. from Domestic Water in the Prefecture of Arta, Greece

    Directory of Open Access Journals (Sweden)

    Dimitra Dimitriadi

    2014-01-01

    Full Text Available The aim of this research was the isolation of Legionella spp. from domestic water supply networks in the Prefecture of Arta. A total of 100 water samples, from 25 houses, were collected. Half of the samples concerned the cold water and half the hot water supply. Purpose was to detect colonization of the water networks with Legionella spp. >500 cfu/L by using the method of filtration (ISO 11731. Out of 100 samples, 6 samples from 3 houses were positive for Legionella spp. Legionella pneumophila serogroup 2–14 was isolated in 5 of 6 samples, whereas in the sixth sample Legionella anisa was identified. Only three of the samples had residual chloride over 0.2 mg/L, rate which is necessary for potable water, according to the Greek hygienic practice. Concerning the temperature of hot water, the mean temperature of the negative for Legionella samples was higher compared to the mean temperature of the positive for Legionella samples (49.9°C versus 45.5°C. It is estimated that there is risk of infection through the use of showers. The low concentration of chloride and the temperature, which was found within the limits favorable to developing Legionella spp. (20–45°C, provide fertile ground for proliferation of the bacteria.

  11. Legionella species and serogroups in Malaysian water cooling towers: identification by latex agglutination and PCR-DNA sequencing of isolates.

    Science.gov (United States)

    Yong, Stacey Foong Yee; Goh, Fen-Ning; Ngeow, Yun Fong

    2010-03-01

    In this study, we investigated the distribution of Legionella species in water cooling towers located in different parts of Malaysia to obtain information that may inform public health policies for the prevention of legionellosis. A total of 20 water samples were collected from 11 cooling towers located in three different states in east, west and south Malaysia. The samples were concentrated by filtration and treated with an acid buffer before plating on to BCYE agar. Legionella viable counts in these samples ranged from 100 to 2,000 CFU ml(-1); 28 isolates from the 24 samples were examined by latex agglutination as well as 16S rRNA and rpoB PCR-DNA sequencing. These isolates were identified as Legionella pneumophila serogroup 1 (35.7%), L. pneumophila serogroup 2-14 (39%), L. pneumophila non-groupable (10.7%), L. busanensis, L. gormanii, L. anisa and L. gresilensis. L. pneumophila was clearly the predominant species at all sampling sites. Repeat sampling from the same cooling tower and testing different colonies from the same water sample showed concurrent colonization by different serogroups and different species of Legionella in some of the cooling towers.

  12. Comparative and functional genomics of Legionella identified eukaryotic like proteins as key players in host-pathogen interactions

    Directory of Open Access Journals (Sweden)

    Laura eGomez-Valero

    2011-10-01

    Full Text Available Although best known for its ability to cause severe pneumonia in people whose immune defenses are weakened, Legionella pneumophila and Legionella longbeachae are two species of a large genus of bacteria that are ubiquitous in nature, where they parasitize protozoa. Adaptation to the host environment and exploitation of host cell functions are critical for the success of these intracellular pathogens. The establishment and publication of the complete genome sequences of L. pneumophila and L. longbeachae isolates paved the way for major breakthroughs in understanding the biology of these organisms. In this review we present the knowledge gained from the analyses and comparison of the complete genome sequences of different L. pneumophila and L. longbeachae strains. Emphasis is given on putative virulence and Legionella life cycle related functions, such as the identification of an extended array of eukaryotic-like proteins, many of which have been shown to modulate host cell functions to the pathogen's advantage. Surprisingly, many of the eukaryotic domain proteins identified in L. pneumophila as well as many substrates of the Dot/Icm type IV secretion system essential for intracellular replication are different between these two species, although they cause the same disease. Finally, evolutionary aspects regarding the eukaryotic like proteins in Legionella are discussed.

  13. Legionella prevalence and risk of legionellosis in Hungarian hospitals.

    Science.gov (United States)

    Barna, Zsófia; Kádár, Mihály; Kálmán, Emese; Róka, Eszter; Szax, Anita Sch; Vargha, Márta

    2015-12-01

    Nosocomial legionellosis is a growing concern worldwide. In Hungary, about 20% of the reported cases are health-care associated, but in the absence of legal regulation, environmental monitoring of Legionella is not routinely performed in hospitals. In the present study, 23 hospitals were investigated. The hot water distribution system was colonized by Legionella in over 90%; counts generally exceeded the public health limit value. Hot water temperature was critically low in all systems (Legionella prevalence (OR = 28.0, 27.3, 27.7, respectively). Risk management interventions (including thermal or chemical disinfection) were only efficient if the system operation was optimized. Though the risk factors were similar, in those hospitals where nosocomial legionellosis was reported, Legionella counts and the proportion of L. pneumophila sg 1 isolates were significantly higher. The results of environmental prevalence of legionellae in hospitals suggest that the incidence of nosocomial legionellosis is likely to be underreported. The observed colonization rates call for the introduction of a mandatory environmental monitoring scheme.

  14. Legionella pneumonia associated with severe acute respiratory distress syndrome and diffuse alveolar hemorrhage - A rare association

    Directory of Open Access Journals (Sweden)

    Muhammad Kashif

    2017-01-01

    Full Text Available Legionella pneumophila is a common, usually underreported and undiagnosed cause of community acquired pneumonia which can lead to significant morbidity and mortality. Diffuse alveolar hemorrhage rarely have been associated with legionella infection. We present a 61-year-old man with hypertension, diabetes mellitus and obesity admitted with severe acute respiratory distress syndrome. He was found to have Legionella pneumonia with associated diffuse alveolar hemorrhage diagnosed with bronchoscopic sequential bronchoalveolar lavage. He was successfully managed with antibiotics, lung protective strategies and intravenous pulse dose steroids. This patient highlights the unusual association of Legionella infection and diffuse alveolar hemorrhage. Additionally, the case re-enforces the need for early and aggressive evaluation and management of patients presenting with pneumonia and progressive hypoxia despite adequate treatment.

  15. Uncovering the Legionella genus effector repertoire - strength in diversity and numbers

    Science.gov (United States)

    Burstein, David; Amaro, Francisco; Zusman, Tal; Lifshitz, Ziv; Cohen, Ofir; Gilbert, Jack A; Pupko, Tal; Shuman, Howard A; Segal, Gil

    2016-01-01

    Infection by the human pathogen Legionella pneumophila relies on the translocation of ~300 virulence proteins, termed effectors, which manipulate host-cell processes. However, almost no information exists regarding effectors in other Legionella pathogens. Here we sequenced, assembled and characterized the genomes of 38 Legionella species, and predicted their effector repertoire using a previously validated machine-learning approach. This analysis revealed a treasure trove of 5,885 predicted effectors. The effector repertoire of different Legionella species was found to be largely non-overlapping, and only seven core-effectors were shared among all species studied. Species-specific effectors had atypically low GC content, suggesting exogenous acquisition, possibly from their natural protozoan hosts. Furthermore, we detected numerous novel conserved effector domains, and discovered new domain combinations, which allowed inferring yet undescribed effector functions. The effector collection and network of domain architectures described here can serve as a roadmap for future studies of effector function and evolution. PMID:26752266

  16. Genome Dynamics in Legionella: The Basis of Versatility and Adaptation to Intracellular Replication

    Science.gov (United States)

    Gomez-Valero, Laura; Buchrieser, Carmen

    2013-01-01

    Legionella pneumophila is a bacterial pathogen present in aquatic environments that can cause a severe pneumonia called Legionnaires’ disease. Soon after its recognition, it was shown that Legionella replicates inside amoeba, suggesting that bacteria replicating in environmental protozoa are able to exploit conserved signaling pathways in human phagocytic cells. Comparative, evolutionary, and functional genomics suggests that the Legionella–amoeba interaction has shaped this pathogen more than previously thought. A complex evolutionary scenario involving mobile genetic elements, type IV secretion systems, and horizontal gene transfer among Legionella, amoeba, and other organisms seems to take place. This long-lasting coevolution led to the development of very sophisticated virulence strategies and a high level of temporal and spatial fine-tuning of bacteria host–cell interactions. We will discuss current knowledge of the evolution of virulence of Legionella from a genomics perspective and propose our vision of the emergence of this human pathogen from the environment. PMID:23732852

  17. Characterization of aerosols containing Legionella generated upon nebulization

    Science.gov (United States)

    Allegra, Séverine; Leclerc, Lara; Massard, Pierre André; Girardot, Françoise; Riffard, Serge; Pourchez, Jérémie

    2016-09-01

    Legionella pneumophila is, by far, the species most frequently associated with Legionnaires’ disease (LD). Human infection occurs almost exclusively by aerosol inhalation which places the bacteria in juxtaposition with alveolar macrophages. LD risk management is based on controlling water quality by applying standardized procedures. However, to gain a better understanding of the real risk of exposure, there is a need (i) to investigate under which conditions Legionella may be aerosolized and (ii) to quantify bacterial deposition into the respiratory tract upon nebulization. In this study, we used an original experimental set-up that enables the generation of aerosol particles containing L. pneumophila under various conditions. Using flow cytometry in combination with qPCR and culture, we determined (i) the size of the aerosols and (ii) the concentration of viable Legionella forms that may reach the thoracic region. We determined that the 0.26-2.5 μm aerosol size range represents 7% of initial bacterial suspension. Among the viable forms, 0.7% of initial viable bacterial suspension may reach the pulmonary alveoli. In conclusion, these deposition profiles can be used to standardize the size of inoculum injected in any type of respiratory tract model to obtain new insights into the dose response for LD.

  18. Legionella pneumophila: Virulent and Avirulent Interaction with Acanthamoeba castellanii.

    Science.gov (United States)

    1993-08-01

    something else to look forward to the rc•t of my life. Finally, this work is dedicated to the memory of my mother, Dolores Anne Ravine, whose life shall...water, sea water, industrial cooling water, swimming pools, dental units, dialysis units and contact lens cases. Additionally, it can be recovered

  19. Inter-kingdom signaling by the Legionella Quorum Sensing Molecule LAI-1 modulates cell migration through an IQGAP1-Cdc42-ARHGEF9-dependent pathway

    OpenAIRE

    Sylvia Simon; Ursula Schell; Natalie Heuer; Dominik Hager; Albers, Michael F.; Jan Matthias; Felix Fahrnbauer; Dirk Trauner; Ludwig Eichinger; Christian Hedberg; Hubert Hilbi

    2015-01-01

    Small molecule signaling promotes the communication between bacteria as well as between bacteria and eukaryotes. The opportunistic pathogenic bacterium Legionella pneumophila employs LAI-1 (3-hydroxypentadecane-4-one) for bacterial cell-cell communication. LAI-1 is produced and detected by the Lqs (Legionella quorum sensing) system, which regulates a variety of processes including natural competence for DNA uptake and pathogen-host cell interactions. In this study, we analyze the role of LAI-...

  20. Rash, disseminated intravascular coagulation and legionella: Episode 10 and a rewind into the past

    Directory of Open Access Journals (Sweden)

    Prashanth M. Thalanayar

    2015-01-01

    Full Text Available Legionella pneumophila is the most common cause of legionellosis and is one of the organisms causing atypical pneumonia. We report the presentation of disseminated intravascular coagulation (DIC and skin rash in a single case of severe Legionella pneumonia. The unique clinical presentation of a diffuse rash diagnosed as purpura fulminans and the unpredictable variations encountered during the diagnostic work-up of the case make this write-up crucial. This article synthesizes all reported cases of L. pneumonia associated with cutaneous manifestations as well as cases presenting with DIC. Furthermore, this manuscript illustrates the correlation between cutaneous and coagulopathic manifestations, and morbidity and mortality from L. pneumonia.

  1. Legionella spp. in UK composts--a potential public health issue?

    Science.gov (United States)

    Currie, S L; Beattie, T K; Knapp, C W; Lindsay, D S J

    2014-04-01

    Over the past 5 years, a number of cases of legionellosis in Scotland have been associated with compost use; however, studies investigating sources of infection other than water systems remain limited. This study delivers the first comprehensive survey of composts commonly available in the UK for the presence of Legionella species. Twenty-two store-bought composts, one green-waste compost and one home-made compost were tested for Legionella by culture methods on BCYE-α medium, and the findings were confirmed by macrophage infectivity potentiator (mip) speciation. Twenty-two of the samples were retested after an enrichment period of 8 weeks. In total, 15 of 24 composts tested positive for Legionella species, a higher level of contamination than previously seen in Europe. Two isolates of Legionella pneumophila were identified, and Legionella longbeachae serogroup 1 was found to be one of the most commonly isolated species. L. longbeachae infection would not be detected by routine Legionella urinary antigen assay, so such testing should not be used as the sole diagnostic technique in atypical pneumonia cases, particularly where there is an association with compost use. The occurrence of Legionella in over half of the samples tested indicates that compost could pose a public health risk. The addition of general hygiene warnings to compost packages may be beneficial in protecting public health. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

  2. Evaluation of Legionella Air Contamination in Healthcare Facilities by Different Sampling Methods: An Italian Multicenter Study

    Directory of Open Access Journals (Sweden)

    Maria Teresa Montagna

    2017-06-01

    Full Text Available Healthcare facilities (HF represent an at-risk environment for legionellosis transmission occurring after inhalation of contaminated aerosols. In general, the control of water is preferred to that of air because, to date, there are no standardized sampling protocols. Legionella air contamination was investigated in the bathrooms of 11 HF by active sampling (Surface Air System and Coriolis®μ and passive sampling using settling plates. During the 8-hour sampling, hot tap water was sampled three times. All air samples were evaluated using culture-based methods, whereas liquid samples collected using the Coriolis®μ were also analyzed by real-time PCR. Legionella presence in the air and water was then compared by sequence-based typing (SBT methods. Air contamination was found in four HF (36.4% by at least one of the culturable methods. The culturable investigation by Coriolis®μ did not yield Legionella in any enrolled HF. However, molecular investigation using Coriolis®μ resulted in eight HF testing positive for Legionella in the air. Comparison of Legionella air and water contamination indicated that Legionella water concentration could be predictive of its presence in the air. Furthermore, a molecular study of 12 L. pneumophila strains confirmed a match between the Legionella strains from air and water samples by SBT for three out of four HF that tested positive for Legionella by at least one of the culturable methods. Overall, our study shows that Legionella air detection cannot replace water sampling because the absence of microorganisms from the air does not necessarily represent their absence from water; nevertheless, air sampling may provide useful information for risk assessment. The liquid impingement technique appears to have the greatest capacity for collecting airborne Legionella if combined with molecular investigations.

  3. Evaluation of Legionella Air Contamination in Healthcare Facilities by Different Sampling Methods: An Italian Multicenter Study.

    Science.gov (United States)

    Montagna, Maria Teresa; De Giglio, Osvalda; Cristina, Maria Luisa; Napoli, Christian; Pacifico, Claudia; Agodi, Antonella; Baldovin, Tatjana; Casini, Beatrice; Coniglio, Maria Anna; D'Errico, Marcello Mario; Delia, Santi Antonino; Deriu, Maria Grazia; Guida, Marco; Laganà, Pasqualina; Liguori, Giorgio; Moro, Matteo; Mura, Ida; Pennino, Francesca; Privitera, Gaetano; Romano Spica, Vincenzo; Sembeni, Silvia; Spagnolo, Anna Maria; Tardivo, Stefano; Torre, Ida; Valeriani, Federica; Albertini, Roberto; Pasquarella, Cesira

    2017-06-22

    Healthcare facilities (HF) represent an at-risk environment for legionellosis transmission occurring after inhalation of contaminated aerosols. In general, the control of water is preferred to that of air because, to date, there are no standardized sampling protocols. Legionella air contamination was investigated in the bathrooms of 11 HF by active sampling (Surface Air System and Coriolis(®)μ) and passive sampling using settling plates. During the 8-hour sampling, hot tap water was sampled three times. All air samples were evaluated using culture-based methods, whereas liquid samples collected using the Coriolis(®)μ were also analyzed by real-time PCR. Legionella presence in the air and water was then compared by sequence-based typing (SBT) methods. Air contamination was found in four HF (36.4%) by at least one of the culturable methods. The culturable investigation by Coriolis(®)μ did not yield Legionella in any enrolled HF. However, molecular investigation using Coriolis(®)μ resulted in eight HF testing positive for Legionella in the air. Comparison of Legionella air and water contamination indicated that Legionella water concentration could be predictive of its presence in the air. Furthermore, a molecular study of 12 L. pneumophila strains confirmed a match between the Legionella strains from air and water samples by SBT for three out of four HF that tested positive for Legionella by at least one of the culturable methods. Overall, our study shows that Legionella air detection cannot replace water sampling because the absence of microorganisms from the air does not necessarily represent their absence from water; nevertheless, air sampling may provide useful information for risk assessment. The liquid impingement technique appears to have the greatest capacity for collecting airborne Legionella if combined with molecular investigations.

  4. Use of copper-silver ionization for the control of legionellae in alkaline environments at health care facilities.

    Science.gov (United States)

    Dziewulski, David M; Ingles, Erin; Codru, Neculai; Strepelis, John; Schoonmaker-Bopp, Dianna

    2015-09-01

    There are multiple treatment options for the control of legionellae in premise hot water systems. Water chemistry plays a role in the efficacy of these treatments and should be considered when selecting a treatment. This study demonstrated the efficacy of copper-silver ionization (CSI) under alkaline water conditions in 2 health care facilities. Monitoring for copper (Cu) and silver (Ag) ions was performed, and the corresponding percentage of positive Legionella cultures was monitored. Low Legionella colony forming units (CFU), with a mean 10(3) CFU/100 mL) to consistently <30% (38 CFU/100 mL). Control of legionellae in premise water systems may be a complex process requiring long-term assessments for adequate control. This work found that CSI could be successful in controlling Legionella under alkaline water conditions, and the evidence suggests that Ag ions are responsible for the control of Legionella pneumophila 1, L pneumophila 6, and L anisa. Copyright © 2015 Association for Professionals in Infection Control and Epidemiology, Inc. Published by Elsevier Inc. All rights reserved.

  5. Presence of Legionella spp. in Hot Water Networks of Different Italian Residential Buildings: A Three-Year Survey

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    Michele Totaro

    2017-10-01

    Full Text Available Although the European reports highlight an increase in community-acquired Legionnaires’ disease cases, the risk of Legionella spp. in private houses is underestimated. In Pisa (Italy we performed a three-year survey on Legionella presence in 121 buildings with an independent hot water production (IB; 64 buildings with a central hot water production (CB; and 35 buildings with a solar thermal system for hot water production (TB. From all the 220 buildings Legionella spp. was researched in two hot water samples collected either at the recirculation point or on the first floor and on the last floor, while the potable water quality was analysed in three cold water samples collected at the inlet from the aqueduct network, at the exit from the autoclave, and at the most remote tap. Legionella pneumophila sg1, Legionella pneumophila sg2–16, and non-pneumophila Legionella species were detected in 26% of the hot water networks, mostly in CB and TB. In these buildings we detected correlations between the presence of Legionella and the total chlorine concentration decrease and/or the increase of the temperature. Cold water resulted free from microbiological hazards, with the exception of Serratia liquefaciens and Enterobacter cloacae isolated at the exit from two different autoclaves. We observed an increase in total microbial counts at 22 °C and 37 °C between the samples collected at the most remote taps compared to the ones collected at the inlet from the aqueduct. The study highlights a condition of potential risk for susceptible categories of population and supports the need for measures of risk assessment and control.

  6. Fatal pneumonia by Legionella in a farmer with hypersensitivity pneumonitis Neumonía por Legionella con desenlace fatal en un granjero con neumonitis por hipersensibilidad

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    Vega García López

    2009-12-01

    Full Text Available Background: The retrospective investigation of a fatal sporadic Legionnaires' disease identified an unknown case of occupational hypersensitivity pneumonitis in a swine breeder. Methods: Chest high-resolution computed tomography, bronchoalveolar lavage, lung biopsy, arterial gasometry, pulmonary function tests and autopsy were performed. It was studied the presence of Legionella by serology and risk water samples were analyzed to identify the Legionella's source. Results: HP and Legionella pneumophila pneumonia diagnostics were confirmed. Lung fibrosis, a restrictive functional pattern, decreased diffusion, hypoxemia and bronchoalveolar lavage lymphocytosis were evidenced. Legionella´s source was detected in a shower and a positive serology in the patient. Autopsy verified pulmonary fibrosis and the septic shock leaded to Legionella causing the death. Conclusions: Chronic cough and pulmonary infiltrates in a farmer should suspect the presence of hypersensitivity pneumonitis. Later diagnosis carries a worse prognosis, the offending antigens exposure can't be avoided and fibrotic stage enhanced opportunity infection disease.Introducción: La investigación retrospectiva sobre un fallecimiento aislado por Legionelosis, hizo aflorar un caso de neumonitis por hipersensibilidad en un granjero cuidador de cerdos. Métodos: Se realizaron las siguientes pruebas: tomografía axial computerizada de alta resolución, lavado broncoalveolar, biopsia pulmonar, gasometría arterial, pruebas de función respiratoria y autopsia. Se estudió la presencia de Legionella por serología y se analizaron las muestras de fuentes de riesgo para identificar el foco de Legionella. Resultados: El estudio confirmó los diagnósticos de neumonitis por hipersensibilidad y neumonía por Legionella pneumophila. Las pruebas realizadas objetivaron la fibrosis pulmonar, un patrón respiratorio funcional restrictivo, un descenso de la difusión pulmonar, hipoxemia y la presencia de

  7. Analysis of the Legionella longbeachae genome and transcriptome uncovers unique strategies to cause Legionnaires' disease.

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    Christel Cazalet

    2010-02-01

    Full Text Available Legionella pneumophila and L. longbeachae are two species of a large genus of bacteria that are ubiquitous in nature. L. pneumophila is mainly found in natural and artificial water circuits while L. longbeachae is mainly present in soil. Under the appropriate conditions both species are human pathogens, capable of causing a severe form of pneumonia termed Legionnaires' disease. Here we report the sequencing and analysis of four L. longbeachae genomes, one complete genome sequence of L. longbeachae strain NSW150 serogroup (Sg 1, and three draft genome sequences another belonging to Sg1 and two to Sg2. The genome organization and gene content of the four L. longbeachae genomes are highly conserved, indicating strong pressure for niche adaptation. Analysis and comparison of L. longbeachae strain NSW150 with L. pneumophila revealed common but also unexpected features specific to this pathogen. The interaction with host cells shows distinct features from L. pneumophila, as L. longbeachae possesses a unique repertoire of putative Dot/Icm type IV secretion system substrates, eukaryotic-like and eukaryotic domain proteins, and encodes additional secretion systems. However, analysis of the ability of a dotA mutant of L. longbeachae NSW150 to replicate in the Acanthamoeba castellanii and in a mouse lung infection model showed that the Dot/Icm type IV secretion system is also essential for the virulence of L. longbeachae. In contrast to L. pneumophila, L. longbeachae does not encode flagella, thereby providing a possible explanation for differences in mouse susceptibility to infection between the two pathogens. Furthermore, transcriptome analysis revealed that L. longbeachae has a less pronounced biphasic life cycle as compared to L. pneumophila, and genome analysis and electron microscopy suggested that L. longbeachae is encapsulated. These species-specific differences may account for the different environmental niches and disease epidemiology of these

  8. Amoebae in domestic water systems: resistance to disinfection treatments and implication in Legionella persistence.

    Science.gov (United States)

    Thomas, V; Bouchez, T; Nicolas, V; Robert, S; Loret, J F; Lévi, Y

    2004-01-01

    Monitoring of microbial changes during and after application of various disinfection treatments in a model domestic water system. A pilot-scale domestic water system consisting of seven galvanized steel re-circulation loops and copper dead legs was constructed. Culture techniques, confocal laser scanning microscopy after fluorescent in situ hybridization and viability staining with the BacLight LIVE/DEAD kit were used for planktonic and biofilm flora monitoring. Before starting the treatments, the system was highly contaminated with Legionella pneumophila and biofilm populations mainly consisted of beta-proteobacteria. In the water and the biofilm of the loops, continuous application of chlorine dioxide (0.5 mg l(-1)), or chlorine (2.5 mg l(-1)) were very effective in reducing the microbial flora, including L. pneumophila. Heterotrophic bacteria, although strongly reduced, were still detectable after ozone application (0.5 mg l(-1)), whereas with monochloramine (0.5 mg l(-1)) and copper-silver ionization (0.8/0.02 mg l(-1)), the contamination remained significantly higher. Monochloramine and copper-silver did not remove the biofilm. During copper-silver application, Legionella re-growth was observed. Only chlorine dioxide led to detectable effects in the dead leg. Amoebae could not be eliminated, and after interrupting the treatments, L. pneumophila quickly recovered their initial levels, in all cases. Chlorine dioxide, applied as a continuous treatment, was identified in this study as the most efficient for controlling L. pneumophila in a domestic water system. Chlorine dioxide showed a longer residual activity, leading to improved performance in the dead leg. Amoebae resisted to all the treatments applied and probably acted as reservoirs for L. pneumophila, allowing a quick re-colonization of the system once the treatments were interrupted. Control of microbial contamination requires maintenance of a constant disinfectant residual throughout the water system

  9. Mistaken identity: Legionella micdadei appearing as acid-fast bacilli on lung biopsy of a hematopoietic stem cell transplant patient.

    Science.gov (United States)

    Waldron, P R; Martin, B A; Ho, D Y

    2015-02-01

    Legionella micdadei is a potential cause of invasive lung infections in immunocompromised hosts. On biopsy specimens, it can appear as an acid-fast bacillus (AFB) and can be mistaken for a member of genus Mycobacterium. As Legionella requires selective media to grow in culture, and the commonly used, commercially available urine antigen test for Legionella only detects Legionella pneumophila serogroup-1, but not L. micdadei, it is important to consider this organism in the differential diagnosis for AFB in immunocompromised hosts. We report a case of L. micdadei infection, which was initially treated empirically for non-tuberculous mycobacteria based on AFB staining of biopsy tissue before the final diagnosis was made. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  10. Legionella in habitations

    DEFF Research Database (Denmark)

    Krøjgaard, Louise Hjelmar

    and the results from the PhD work the following subjects were addressed: a) prevalence of Legionella in habitations, b) validation of the use of qPCR in risk assessment in hot water systems, c) clarifying risk factors mainly associated with Legionella in habitations, and d) discussion of interventions which could......, it was difficult to interpret the specific amount. In samples collected from the first flush from empty apartments, culture and qPCR were inconclusive. The literature studies showed that Legionella is widely dispersed in habitations all over the world, including in Denmark. Different major risk factors were...

  11. Antibiotic susceptibility of Legionella strains isolated from public water sources in Macau and Guangzhou.

    Science.gov (United States)

    Xiong, Lina; Yan, He; Shi, Lei; Mo, Ziyao

    2016-12-01

    The purpose of this study was to investigate the susceptibility of waterborne strains of Legionella to eight antimicrobials commonly used in legionellosis therapy. The minimum inhibitory concentrations (MICs) of 66 environmental Legionella strains, isolated from fountains and cooling towers of public facilities (hotels, schools, and shopping malls) in Macau and Guangzhou, were tested using the microdilution method in buffered yeast extract broth. The MIC50/MIC90 values for erythromycin, cefotaxime (CTX), doxycycline (DOC), minocycline (MIN), azithromycin, ciprofloxacin, levofloxacin (LEV), and moxifloxacin were 0.125/0.5 mg/L, 4/8 mg/L, 8/16 mg/L, 4/8 mg/L, 0.125/0.5 mg/L, 0.031/0.031 mg/L, 0.031/0.031 mg/L, and 0.031/0.062 mg/L, respectively. Legionella isolates were inhibited by either low concentrations of macrolides and fluoroquinolones, or high concentrations of CTX and tetracycline drugs. LEV was the most effective drug against different Legionella species and serogroups of L. pneumophila isolates. The latter were inhibited in decreasing order by MIN > CTX >DOC, while non-L. pneumophila isolates were inhibited by CTX> MIN >DOC. In this study, we evaluated drug resistance of pathogenic bacteria from the environment. This may help predict the emergence of drug resistance, improve patient outcomes, and reduce hospitalization costs.

  12. Multiple regression as a preventive tool for determining the risk of Legionella spp.

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    Enrique Gea-Izquierdo

    2012-04-01

    Full Text Available To determine the interrelationship between health & hygiene conditions for prevention of legionellosis, the compositionof materials used in water distribution systems, the water origin and Legionella pneumophila risk. Material and methods. Include adescriptive study and multiple regression analysis on a sample of golf course sprinkler irrigation systems (n=31 pertaining to hotelslocated on the Costa del Sol (Malaga, Spain. The study was carried out in 2009. Results. Presented a significant lineal relation, withall the independent variables contributing significantly (p<0.05 to the model’s fit. The relationship between water type and the risk ofLegionella, as well as the material composition and the latter, is lineal and positive. In contrast, the relationship between health-hygieneconditions and Legionella risk is lineal and negative. Conclusion. The characterization of Legionella pneumophila concentration, asdefined by the risk in water and through use of the predictive method, can contribute to the consideration of new influence variables inthe development of the agent, resulting in improved control and prevention of the disease.

  13. Interactive Effects of Corrosion, Copper, and Chloramines on Legionella and Mycobacteria in Hot Water Plumbing.

    Science.gov (United States)

    Rhoads, William J; Pruden, Amy; Edwards, Marc A

    2017-06-20

    Complexities associated with drinking water plumbing systems can result in undesirable interactions among plumbing components that undermine engineering controls for opportunistic pathogens (OPs). In this study, we examine the effects of plumbing system materials and two commonly applied disinfectants, copper and chloramines, on water chemistry and the growth of Legionella and mycobacteria across a transect of bench- and pilot-scale hot water experiments carried out with the same municipal water supply. We discovered that copper released from corrosion of plumbing materials can initiate evolution of >1100 times more hydrogen (H2) from water heater sacrificial anode rods than does presence of copper dosed as soluble cupric ions. H2 is a favorable electron donor for autotrophs and causes fixation of organic carbon that could serve as a nutrient for OPs. Dosed cupric ions acted as a disinfectant in stratified stagnant pipes, inhibiting culturable Legionella and biofilm formation, but promoted Legionella growth in pipes subject to convective mixing. This difference was presumably due to continuous delivery of nutrients to biofilm on the pipes under convective mixing conditions. Chloramines eliminated culturable Legionella and prevented L. pneumophila from recolonizing biofilms, but M. avium gene numbers increased by 0.14-0.76 logs in the bulk water and were unaffected in the biofilm. This study provides practical confirmation of past discrepancies in the literature regarding the variable effects of copper on Legionella growth, and confirms prior reports of trade-offs between Legionella and mycobacteria if chloramines are applied as secondary disinfectant residual.

  14. Legionella cardiaca sp. nov., isolated from a case of native valve endocarditis in a human heart

    Science.gov (United States)

    Pearce, Meghan M.; Theodoropoulos, Nicole; Mandel, Mark J.; Brown, Ellen; Reed, Kurt D.

    2012-01-01

    A Gram-negative, rod-shaped bacterium, designated H63T, was isolated from aortic valve tissue of a patient with native valve endocarditis. 16S rRNA gene sequencing revealed that H63T belongs to the genus Legionella, with its closest neighbours being the type strains of Legionella brunensis (98.8 % similarity), L. londiniensis (97.0 %), L. jordanis (96.8 %), L. erythra (96.2 %), L. dresdenensis (96.0 %) and L. rubrilucens, L. feeleii, L. pneumophila and L. birminghamensis (95.7 %). DNA–DNA hybridization studies yielded values of Legionella. H63T was distinguishable from its neighbours based on it being positive for hippurate hydrolysis. H63T was further differentiated by its inability to grow on BCYE agar at 17 °C, its poor growth on low-iron medium and the absence of sliding motility. Also, H63T did not react with antisera generated from type strains of Legionella species. H63T replicated within macrophages. It also grew in mouse lungs, inducing histopathological evidence of pneumonia and dissemination to the spleen. Together, these results confirm that H63T represents a novel, pathogenic Legionella species, for which the name Legionella cardiaca sp. nov. is proposed. The type strain is H63T ( = ATCC BAA-2315T  = DSM 25049T  = JCM 17854T). PMID:22286905

  15. Free-living amoebae (FLA) co-occurring with legionellae in industrial waters☆

    Science.gov (United States)

    Scheikl, Ute; Sommer, Regina; Kirschner, Alexander; Rameder, Alexandra; Schrammel, Barbara; Zweimüller, Irene; Wesner, Wolfgang; Hinker, Manfred; Walochnik, Julia

    2014-01-01

    Legionella pneumophila is known as the causative agent of Legionnaires’ disease and free-living amoebae (FLA) can serve as vehicles for legionellae. The aim of this study was to screen industrial waters for the occurrence of FLA and their co-occurrence with legionellae. A total of 201 water samples, including 129 cooling waters and 72 process waters, and 30 cooling lubricants were included in the study. Treated waters were screened periodically, pre and post treatment. Altogether, 72.6% of the water samples were positive for FLA, acanthamoebae being most prevalent (in 23.9% of the samples) followed by Vermamoeba vermiformis (19.4%). Only one cooling lubricant was positive (Acanthamoeba genotype T4). Legionella spp. were detected in 34.8% of the water samples and in 15% in high concentrations (>1000 CFU/100 ml). Altogether, 81.4% of the Legionella-positive samples were positive for FLA by standard methods. By applying a highly sensitive nested PCR to a representative set of random samples it was revealed that Legionella spp. always co-occurred with Acanthamoeba spp. Although the addition of disinfectants did influence amoebal density and diversity, treated waters showed no difference concerning FLA in the interphases of disinfection. It appears that FLA can re-colonize treated waters within a short period of time. PMID:25062389

  16. Free-living amoebae (FLA) co-occurring with legionellae in industrial waters.

    Science.gov (United States)

    Scheikl, Ute; Sommer, Regina; Kirschner, Alexander; Rameder, Alexandra; Schrammel, Barbara; Zweimüller, Irene; Wesner, Wolfgang; Hinker, Manfred; Walochnik, Julia

    2014-08-01

    Legionella pneumophila is known as the causative agent of Legionnaires' disease and free-living amoebae (FLA) can serve as vehicles for legionellae. The aim of this study was to screen industrial waters for the occurrence of FLA and their co-occurrence with legionellae. A total of 201 water samples, including 129 cooling waters and 72 process waters, and 30 cooling lubricants were included in the study. Treated waters were screened periodically, pre and post treatment. Altogether, 72.6% of the water samples were positive for FLA, acanthamoebae being most prevalent (in 23.9% of the samples) followed by Vermamoeba vermiformis (19.4%). Only one cooling lubricant was positive (Acanthamoeba genotype T4). Legionella spp. were detected in 34.8% of the water samples and in 15% in high concentrations (>1000 CFU/100 ml). Altogether, 81.4% of the Legionella-positive samples were positive for FLA by standard methods. By applying a highly sensitive nested PCR to a representative set of random samples it was revealed that Legionella spp. always co-occurred with Acanthamoeba spp. Although the addition of disinfectants did influence amoebal density and diversity, treated waters showed no difference concerning FLA in the interphases of disinfection. It appears that FLA can re-colonize treated waters within a short period of time. Copyright © 2014 The Authors. Published by Elsevier GmbH.. All rights reserved.

  17. Legionella pneumonia appeared during hospitalization in a patient with hematological malignancy confirmed by sputum culture after negative urine antigen test.

    Science.gov (United States)

    Hase, Ryota; Miyoshi, Kazuyasu; Matsuura, Yasuhiro; Endo, Yasunobu; Nakamura, Masaki; Otsuka, Yoshihito

    2018-01-17

    Legionella pneumophila is recognized as a common causative organism for community-acquired pneumonia, but it is rarely a causative organism for hospital-acquired pneumonia, except in cases of hospital outbreak. Recently, most of the Legionella cases have been diagnosed using the urine antigen test. However, this test can reliably detect only L. pneumophila serogroup 1. Here we report a 63-year-old male patient who was recently diagnosed with acute leukemia and treated with chemotherapy and who developed pneumonia on hospital day 8 during the nadir phase. He was later diagnosed with Legionella pneumonia by culture despite a negative urine antigen test. This case suggests that Legionella pneumonia is an important differential diagnosis for pneumonia in inpatients in the early phase of hospitalization and that when Legionella infection is clinically suspected, culture using selective media or molecular tests should be performed even if the urine antigen test is negative. Copyright © 2017 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

  18. Prevalence of Legionella in premise plumbing in Hungary.

    Science.gov (United States)

    Barna, Zsófia; Kádár, Mihály; Kálmán, Emese; Scheirich Szax, Anita; Vargha, Márta

    2016-03-01

    Legionella is one of the emerging concerns of water quality in built water environments. Premise plumbing systems are among the recognised sources of infection. In the present study, colonisation of hot water networks in health care facilities, schools, hotels, private residences, office and industrial buildings was investigated. Data was analysed in connection with building and premise plumbing characteristics. Over 60% of all buildings were colonised by Legionella; counts were over 1000 CFU/L in 49%. The most prevalent type was Legionella pneumophila serogroup 2-14, isolated from 75% of the positive samples. Centrally produced hot water was found to be a key risk factor (46% of the samples were positive vs. 16% in individual systems); within this group the type of the building was less relevant. Colonisation levels in schools were similar to hotels or hospitals, representing a previously underestimated risk setting. Systems supplied by water from deep groundwater sources were significantly less likely to be colonised than more vulnerable sources (bank-wall filtration, surface water abstraction or karstic water; 28% vs. 51% positive), regardless of the type of treatment applied, including the presence of disinfection. The aggravating effect of larger, more complex and older buildings on colonisation was also confirmed. The present study represents the first baseline analysis, pre-empting regulation or monitoring requirements for Legionella. The prevalence of legionellae and the identified risk factors are indicative for other settings lacking targeted interventions. The statistically confirmed risk factors can serve as indicators for preliminary risk assessment and the prioritisation of interventions. Copyright © 2015 Elsevier Ltd. All rights reserved.

  19. A Legionella Effector Disrupts Host Cytoskeletal Structure by Cleaving Actin.

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    Yao Liu

    2017-01-01

    Full Text Available Legionella pneumophila, the etiological agent of Legionnaires' disease, replicates intracellularly in protozoan and human hosts. Successful colonization and replication of this pathogen in host cells requires the Dot/Icm type IVB secretion system, which translocates approximately 300 effector proteins into the host cell to modulate various cellular processes. In this study, we identified RavK as a Dot/Icm substrate that targets the host cytoskeleton and reduces actin filament abundance in mammalian cells upon ectopic expression. RavK harbors an H95EXXH99 motif associated with diverse metalloproteases, which is essential for the inhibition of yeast growth and for the induction of cell rounding in HEK293T cells. We demonstrate that the actin protein itself is the cellular target of RavK and that this effector cleaves actin at a site between residues Thr351 and Phe352. Importantly, RavK-mediated actin cleavage also occurs during L. pneumophila infection. Cleavage by RavK abolishes the ability of actin to form polymers. Furthermore, an F352A mutation renders actin resistant to RavK-mediated cleavage; expression of the mutant in mammalian cells suppresses the cell rounding phenotype caused by RavK, further establishing that actin is the physiological substrate of RavK. Thus, L. pneumophila exploits components of the host cytoskeleton by multiple effectors with distinct mechanisms, highlighting the importance of modulating cellular processes governed by the actin cytoskeleton in the intracellular life cycle of this pathogen.

  20. High prevalence, genetic diversity and intracellular growth ability of Legionella in hot spring environments.

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    Tian Qin

    Full Text Available BACKGROUND: Legionella is the causative agent of Legionnaires' disease, and hot springs are a major source of outbreaks of this disease. It is important from a public health perspective to survey hot spring environments for the presence of Legionella. METHODS: Prospective surveillance of the extent of Legionella pollution was conducted at three hot spring recreational areas in Beijing, China in 2011. Pulsed-field gel electrophoresis (PFGE and sequence-based typing (SBT were used to describe the genetic polymorphism of isolates. The intracellular growth ability of the isolates was determined by interacting with J774 cells and plating the dilutions onto BCYE agar plates. RESULTS: Overall, 51.9% of spring water samples showed Legionella-positive, and their concentrations ranged from 1 CFU/liter to 2,218 CFU/liter. The positive rates of Legionella were significantly associated with a free chlorine concentration of ≥0.2 mg/L, urea concentration of ≥0.05 mg/L, total microbial counts of ≥400 CFU/ml and total coliform of ≥3 MPN/L (p<0.01. The Legionella concentrations were significantly associated with sample temperature, pH, total microbial counts and total coliform (p<0.01. Legionella pneumophila was the most frequently isolated species (98.9%, and the isolated serogroups included serogroups 3 (25.3%, 6 (23.4%, 5 (19.2%, 1 (18.5%, 2 (10.2%, 8 (0.4%, 10 (0.8%, 9 (1.9% and 12 (0.4%. Two hundred and twenty-eight isolates were analyzed by PFGE and 62 different patterns were obtained. Fifty-seven L. pneumophila isolates were selected for SBT analysis and divided into 35 different sequence types with 5 main clonal groups. All the 57 isolates had high intracellular growth ability. CONCLUSIONS: Our results demonstrated high prevalence and genetic polymorphism of Legionella in springs in Beijing, China, and the SBT and intracellular growth assay results suggested that the Legionella isolates of hot spring environments were pathogenic. Improved control

  1. High Prevalence, Genetic Diversity and Intracellular Growth Ability of Legionella in Hot Spring Environments

    Science.gov (United States)

    Zhou, Haijian; Wang, Huanxin; Xu, Ying; Zhao, Mingqiang; Guan, Hong; Li, Machao; Shao, Zhujun

    2013-01-01

    Background Legionella is the causative agent of Legionnaires' disease, and hot springs are a major source of outbreaks of this disease. It is important from a public health perspective to survey hot spring environments for the presence of Legionella. Methods Prospective surveillance of the extent of Legionella pollution was conducted at three hot spring recreational areas in Beijing, China in 2011. Pulsed-field gel electrophoresis (PFGE) and sequence-based typing (SBT) were used to describe the genetic polymorphism of isolates. The intracellular growth ability of the isolates was determined by interacting with J774 cells and plating the dilutions onto BCYE agar plates. Results Overall, 51.9% of spring water samples showed Legionella-positive, and their concentrations ranged from 1 CFU/liter to 2,218 CFU/liter. The positive rates of Legionella were significantly associated with a free chlorine concentration of ≥0.2 mg/L, urea concentration of ≥0.05 mg/L, total microbial counts of ≥400 CFU/ml and total coliform of ≥3 MPN/L (pLegionella concentrations were significantly associated with sample temperature, pH, total microbial counts and total coliform (pLegionella pneumophila was the most frequently isolated species (98.9%), and the isolated serogroups included serogroups 3 (25.3%), 6 (23.4%), 5 (19.2%), 1 (18.5%), 2 (10.2%), 8 (0.4%), 10 (0.8%), 9 (1.9%) and 12 (0.4%). Two hundred and twenty-eight isolates were analyzed by PFGE and 62 different patterns were obtained. Fifty-seven L. pneumophila isolates were selected for SBT analysis and divided into 35 different sequence types with 5 main clonal groups. All the 57 isolates had high intracellular growth ability. Conclusions Our results demonstrated high prevalence and genetic polymorphism of Legionella in springs in Beijing, China, and the SBT and intracellular growth assay results suggested that the Legionella isolates of hot spring environments were pathogenic. Improved control and prevention strategies are

  2. Control of Legionella Contamination and Risk of Corrosion in Hospital Water Networks following Various Disinfection Procedures.

    Science.gov (United States)

    Marchesi, Isabella; Ferranti, Greta; Mansi, Antonella; Marcelloni, Anna M; Proietto, Anna R; Saini, Navneet; Borella, Paola; Bargellini, Annalisa

    2016-05-15

    Physical and chemical disinfection methods have been proposed with the aim of controlling Legionella water contamination. To date, the most effective procedures for reducing bacterial contamination have not yet been defined. The aim of this study was to assess the long-term effectiveness of various disinfection procedures in order to reduce both culturable and nonculturable (NC) legionellae in different hospital water networks treated with heat, chlorine dioxide, monochloramine, and hydrogen peroxide. The temperature levels and biocide concentrations that proved to give reliable results were analyzed. In order to study the possible effects on the water pipes, we verified the extent of corrosion on experimental coupons after applying each method for 6 months. The percentage of positive points was at its lowest after treatment with monochloramine, followed by chlorine dioxide, hydrogen peroxide, and hyperthermia. Different selections of Legionella spp. were observed, as networks treated with chlorine-based disinfectants were contaminated mainly by Legionella pneumophila serogroup 1, hyperthermia was associated with serogroups 2 to 14, and hydrogen peroxide treatment was associated mainly with non-pneumophila species. NC cells were detected only in heat-treated waters, and also when the temperature was approximately 60°C. The corrosion rates of the coupons were within a satisfactory limit for water networks, but the morphologies differed. We confirm here that chemical disinfection controls Legionella colonization more effectively than hyperthermia does. Monochloramine was the most effective treatment, while hydrogen peroxide may be a promising alternative to chlorine-based disinfectants due to its ability to select for other, less virulent or nonpathogenic species. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  3. [Legionella contamination risk factors in non-circulating hot spring water].

    Science.gov (United States)

    Karasudani, Tatsuya; Kuroki, Toshiro; Otani, Katsumi; Yamaguchi, Seiichi; Sasaki, Mie; Saito, Shioko; Fujita, Masahiro; Sugiyama, Kanji; Nakajima, Hiroshi; Murakami, Koichi; Taguri, Toshitsugu; Kuramoto, Tsuyoshi; Kura, Fumiaki; Yagita, Kenji; Izumiyama, Shinji; Amemura-Maekawa, Junko; Yamazaki, Toshio; Agata, Kunio; Inouye, Hiroo

    2009-01-01

    We examined water from 182 non-circulating hot spring bathing facilities in Japan for possible Legionella occurrence from June 2005 to December 2006, finding Legionella-positive cultures in 119 (29.5%) of 403 samples. Legionellae occurrence was most prevalent in bathtub water (39.4%), followed by storage tank water (23.8%), water from faucets at the bathtub edge (22.3%), and source-spring water (8.3%), indicating no statistically significant difference, in the number of legionellae, having an overall mean of 66 CFU/100mL. The maximum number of legionellae in water increased as water was sampled downstream:180 CFU/100 mL from source spring, 670 from storage tanks, 4,000 from inlet faucets, and 6,800 from bathtubs. The majority--85.7%--of isolated species were identified as L. pneumophila : L. pneumophila serogroup (SG) 1 in 22%, SG 5 in 21%, and SG 6 in 22% of positive samples. Multivariate logistic regression models used to determine the characteristics of facilities and sanitary management associated with Legionella contamination indicated that legionellae was prevalent in bathtub water under conditions where it was isolated from inlet faucet/pouring gate water (odds ratio [OR] = 6.98, 95% confidence interval [CI] = 2.14 to 22.8). Risk of occurrence was also high when the bathtub volume exceeded 5 m3 (OR = 2.74, 95% CI = 1.28 to 5.89). Legionellae occurrence was significantly reduced when the bathing water pH was lower than 6.0 (OR = 0.12, 95% CI = 0.02 to 0.63). Similarly, occurrence was rare in inlet faucet water or the upper part of the plumbing system for which pH was lower than 6.0 (OR = 0.06, 95% CI = 0.01 to 0.48), and when the water temperature was maintained at 55 degrees C or more (OR = 0.10, 95% CI = 0.01 to 0.77). We also examined the occurrence of amoeba, Mycobacterium spp., Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus in water samples.

  4. Legionella longbeachae and Endocarditis

    OpenAIRE

    Leggieri, Nicola; Gouriet, Frédérique; Thuny, Frank; Habib, Gilbert; Raoult, Didier; Casalta, Jean-Paul

    2012-01-01

    We report a case of infectious endocarditis attributable to Legionella longbeachae. L. longbeachae is usually associated with lung infections. It is commonly found in composted waste wood products. L. longbeachae should be regarded as an agent of infectious endocarditis, notably in the context of gardening involving handling of potting soils.

  5. Beyond Rab GTPases Legionella activates the small GTPase Ran to promote microtubule polymerization, pathogen vacuole motility, and infection.

    Science.gov (United States)

    Hilbi, Hubert; Rothmeier, Eva; Hoffmann, Christine; Harrison, Christopher F

    2014-01-01

    Legionella spp. are amoebae-resistant environmental bacteria that replicate in free-living protozoa in a distinct compartment, the Legionella-containing vacuole (LCV). Upon transmission of Legionella pneumophila to the lung, the pathogens employ an evolutionarily conserved mechanism to grow in LCVs within alveolar macrophages, thus triggering a severe pneumonia termed Legionnaires' disease. LCV formation is a complex and robust process, which requires the bacterial Icm/Dot type IV secretion system and involves the amazing number of 300 different translocated effector proteins. LCVs interact with the host cell's endosomal and secretory vesicle trafficking pathway. Accordingly, in a proteomics approach as many as 12 small Rab GTPases implicated in endosomal and secretory vesicle trafficking were identified and validated as LCV components. Moreover, the small GTPase Ran and its effector protein RanBP1 have been found to decorate the pathogen vacuole. Ran regulates nucleo-cytoplasmic transport, spindle assembly, and cytokinesis, as well as the organization of non-centrosomal microtubules. In L. pneumophila-infected amoebae or macrophages, Ran and RanBP1 localize to LCVs, and the small GTPase is activated by the Icm/Dot substrate LegG1. Ran activation by LegG1 leads to microtubule stabilization and promotes intracellular pathogen vacuole motility and bacterial growth, as well as chemotaxis and migration of Legionella-infected cells.

  6. Effect of disinfectant, water age, and pipe material on occurrence and persistence of Legionella, mycobacteria, Pseudomonas aeruginosa, and two amoebas.

    Science.gov (United States)

    Wang, Hong; Masters, Sheldon; Hong, Yanjuan; Stallings, Jonathan; Falkinham, Joseph O; Edwards, Marc A; Pruden, Amy

    2012-11-06

    Opportunistic pathogens represent a unique challenge because they establish and grow within drinking water systems, yet the factors stimulating their proliferation are largely unknown. The purpose of this study was to examine the influence of pipe materials, disinfectant type, and water age on occurrence and persistence of three opportunistic pathogens (Legionella pneumophila, Mycobacterium avium, and Pseudomonas aeruginosa), broader genera (Legionella and mycobacteria), and two amoeba hosts (Acanthamoeba spp. and Hartmanella vermiformis). Triplicate simulated distribution systems (SDSs) compared iron, cement, and PVC pipe materials fed either chlorinated or chloraminated tap water and were sampled at water ages ranging from 1 day to 5.7 days. Quantitative polymerase chain reaction quantified gene copies of target microorganisms in both biofilm and bulk water. Legionella, mycobacteria, P. aeruginosa, and both amoebas naturally colonized the six SDSs, but L. pneumophila and M. avium were not detected. Disinfectant type and dose was observed to have the strongest influence on the microbiota. Disinfectant decay was noted with water age, particularly in chloraminated SDSs (due to nitrification), generally resulting in increased microbial detection frequencies and densities with water age. The influence of pipe material became apparent at water ages corresponding to low disinfectant residual. Each target microbe appeared to display a distinct response to disinfectant type, pipe materials, water age, and their interactions. Differences between the first and the second samplings (e.g., appearance of Legionella, reduction in P. aeruginosa and Acanthamoeba) suggest a temporally dynamic drinking water microbial community.

  7. Bacterial community dynamics in a cooling tower with emphasis on pathogenic bacteria and Legionella species using universal and genus-specific deep sequencing.

    Science.gov (United States)

    Pereira, Rui P A; Peplies, Jörg; Höfle, Manfred G; Brettar, Ingrid

    2017-10-01

    Cooling towers are the major source of outbreaks of legionellosis in Europe and worldwide. These outbreaks are mostly associated with Legionella species, primarily L. pneumophila, and its surveillance in cooling tower environments is of high relevance to public health. In this study, a combined NGS-based approach was used to study the whole bacterial community, specific waterborne and water-based bacterial pathogens, especially Legionella species, targeting the 16S rRNA gene. This approach was applied to water from a cooling tower obtained by monthly sampling during two years. The studied cooling tower was an open circuit cooling tower with lamellar cooling situated in Braunschweig, Germany. A highly diverse bacterial community was observed with 808 genera including 25 potentially pathogenic taxa using universal 16S rRNA primers. Sphingomonas and Legionella were the most abundant pathogenic genera. By applying genus-specific primers for Legionella, a diverse community with 85 phylotypes, and a representative core community with substantial temporal heterogeneity was observed. A high percentage of sequences (65%) could not be affiliated to an acknowledged species. L. pneumophila was part of the core community and the most abundant Legionella species reinforcing the importance of cooling towers as its environmental reservoir. Major temperature shifts (>10 °C) were the key environmental factor triggering the reduction or dominance of the Legionella species in the Legionella community dynamics. In addition, interventions by chlorine dioxide had a strong impact on the Legionella community composition but not on the whole bacterial community. Overall, the presented results demonstrated the value of a combined NGS approach for the molecular monitoring and surveillance of health related pathogens in man-made freshwater systems. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. Applicability assessment of ceramic microbeads coated with hydroxyapatite-binding silver/titanium dioxide ceramic composite earthplus™ to the eradication of Legionella in rainwater storage tanks for household use

    Science.gov (United States)

    Oana, Kozue; Kobayashi, Michiko; Yamaki, Dai; Sakurada, Tsukasa; Nagano, Noriyuki; Kawakami, Yoshiyuki

    2015-01-01

    Water environments appear to be the habitats of Legionella species. Legionellosis is considered as a preventable illness because bacterial reservoirs can be controlled and removed. Roof-harvested rainwater has attracted significant attention not only as a groundwater recharge but also as a potential alternative source of nonpotable water. We successfully developed ceramic microbeads coated with hydroxyapatite-binding silver/titanium dioxide ceramic composite earthplus™ using the thermal spraying method. The ceramic microbeads were demonstrated to have bactericidal activities against not only Legionella but also coliform and heterotrophic bacteria. Immersing the ceramic microbeads in household rainwater storage tanks was demonstrated to yield the favorable eradication of Legionella organisms. Not only rapid-acting but also long-lasting bactericidal activities of the ceramic microbead were exhibited against Legionella pneumophila. However, time-dependent attenuation of the bactericidal activities against Legionella were also noted in the sustainability appraisal experiment. Therefore, the problems to be overcome surely remain in constantly managing the Legionella-pollution by means of immersing the ceramic microbeads. The results of our investigation apparently indicate that the earthplus™-coated ceramic microbeads would become the favorable tool for Legionella measures in household rainwater storage tanks, which may become the natural reservoir for Legionella species. Our investigation would justify further research and data collection to obtain more reliable procedures to microbiologically regulate the Legionella in rainwater storage tanks. PMID:26346201

  9. Applicability assessment of ceramic microbeads coated with hydroxyapatite-binding silver/titanium dioxide ceramic composite earthplus™ to the eradication of Legionella in rainwater storage tanks for household use.

    Science.gov (United States)

    Oana, Kozue; Kobayashi, Michiko; Yamaki, Dai; Sakurada, Tsukasa; Nagano, Noriyuki; Kawakami, Yoshiyuki

    2015-01-01

    Water environments appear to be the habitats of Legionella species. Legionellosis is considered as a preventable illness because bacterial reservoirs can be controlled and removed. Roof-harvested rainwater has attracted significant attention not only as a groundwater recharge but also as a potential alternative source of nonpotable water. We successfully developed ceramic microbeads coated with hydroxyapatite-binding silver/titanium dioxide ceramic composite earthplus™ using the thermal spraying method. The ceramic microbeads were demonstrated to have bactericidal activities against not only Legionella but also coliform and heterotrophic bacteria. Immersing the ceramic microbeads in household rainwater storage tanks was demonstrated to yield the favorable eradication of Legionella organisms. Not only rapid-acting but also long-lasting bactericidal activities of the ceramic microbead were exhibited against Legionella pneumophila. However, time-dependent attenuation of the bactericidal activities against Legionella were also noted in the sustainability appraisal experiment. Therefore, the problems to be overcome surely remain in constantly managing the Legionella-pollution by means of immersing the ceramic microbeads. The results of our investigation apparently indicate that the earthplus™-coated ceramic microbeads would become the favorable tool for Legionella measures in household rainwater storage tanks, which may become the natural reservoir for Legionella species. Our investigation would justify further research and data collection to obtain more reliable procedures to microbiologically regulate the Legionella in rainwater storage tanks.

  10. Legionella bozemanae synthesizes phosphatidylcholine from exogenous choline.

    Science.gov (United States)

    Palusinska-Szysz, Marta; Janczarek, Monika; Kalitynski, Rafal; Dawidowicz, Andrzej L; Russa, Ryszard

    2011-02-20

    The phospholipid class and fatty acid composition of Legionella bozemanae were determined using thin-layer chromatography, gas-liquid chromatography, and matrix-assisted laser desorption ionization-time of flight mass spectrometry. Phosphatidylcholine, phosphatidylethanolamine, and diphosphatidylglycerol were the predominant phospholipids, while phosphatidyl-N-monomethylethanolamine, phosphatidylglycerol, and phosphatidyl-N,N-dimethylethanolamine were present at low concentrations. With the use of the LC/MS technique, PC16:0/15:0, PC17:/15:0, and PE16:1/15:0 were shown to be the dominant phospholipid constituents, which may be taxonomically significant. Two independent phosphatidylcholine synthesis pathways (the three-step methylation and the one-step CDP-choline pathway) were present and functional in L. bozemanae. In the genome of L. bozemanae, genes encoding two potential phosphatidylcholine forming enzymes, phospholipid N-methyl transferase (PmtA) and phosphatidylcholine synthase (Pcs), homologous to L. longbeachae, L. drancourtii, and L. pneumophila pmtA and pcs genes were identified. Genes pmtA and pcs from L. bozemanae were sequenced and analyzed on nucleotide and amino acid levels. Bacteria grown on an artificial medium with labelled choline synthesized phosphatidylcholine predominantly via the phosphatidylcholine synthase pathway, which indicates that L. bozemanae phosphatidylcholine, similarly as in other bacteria associated with eukaryotes, is an important determinant of host-microbe interactions. Copyright © 2010 Elsevier GmbH. All rights reserved.

  11. Recirculation, stagnation and ventilation: The 2014 legionella episode

    Science.gov (United States)

    Russo, Ana; Soares, Pedro M. M.; Gouveia, Célia M.; Cardoso, Rita M.; Trigo, Ricardo M.

    2017-04-01

    Legionella transmission through the atmosphere is unusual, but not unprecedented. A scientific paper published in 2006 reports a surge in Pas-de-Calais, France, in which 86 people have been infected by bacteria released by a cooling tower more than 6 km away [3]. Similarly, in Norway, in 2005, there was another case where contamination spread beyond 10 km, although more concentrated within a radius of 1 km from an industrial unit [2]. An unprecedented large Legionella outbreak occurred in November 2014 nearby Lisbon, Portugal. As of 7 November 2014, 375 individuals become hill and 12 died infected by the Legionella pneumophila bacteria, contracted by inhalation of steam droplets of contaminated water (aerosols). These droplets are so small that can carry the bacteria directly to the lungs, depositing it in the alveoli. One way of studying the propagation of legionella episodes is through the use of aerosol dispersion models. However, such approaches often require detailed 3D high resolution wind data over the region, which isn't often available for long periods. The likely impact of wind on legionella transmission can also be understood based on the analysis of special types of flow conditions such as stagnation, recirculation and ventilation [1, 4]. The Allwine and Whiteman (AW) approach constitutes a straightforward method to assess the assimilative and dispersal capacities of different airsheds [1,4], as it only requires hourly wind components. Thus, it has the advantage of not needing surface and upper air meteorological observations and a previous knowledge of the atmospheric transport and dispersion conditions. The objective of this study is to analyze if the legionella outbreak event which took place in November 2014 had extreme potential recirculation and/or stagnation characteristics. In order to accomplish the proposed objective, the AW approach was applied for a hindcast time-series covering the affected area (1989-2007) and then for an independent

  12. Intermittent use of copper-silver ionization for Legionella control in water distribution systems: a potential option in buildings housing individuals at low risk of infection.

    Science.gov (United States)

    Liu, Z; Stout, J E; Boldin, M; Rugh, J; Diven, W F; Yu, V L

    1998-01-01

    One copper-silver ionization system was sequentially installed onto the hot-water recirculation lines of two hospital buildings colonized with Legionella pneumophila, serogroup 1. A third building with the same water supply and also colonized with Legionella served as a control. Four weeks after activation of the system, distal site positivity for Legionella in the first test building dropped to zero. After operating for 16 weeks, the system was disconnected and installed onto the second test building. Twelve weeks of disinfection reduced the distal site positivity for Legionella in the second test building to zero. Legionella recolonization did not occur in the first test building for 6-12 weeks and in the second test building for 8-12 weeks after inactivation of the system. The control building remained Legionella-positive throughout the experimental period. A significantly higher copper concentration was found in the biofilm taken from a sampling device than in that from water. This is likely to be the reason that the copper-silver ionization system had the residual effect of preventing early recolonization. Our study raises the possibility that one copper-silver unit could be rotated among several buildings to maintain a Legionella-free environment. Such an approach may be cost-effective for buildings housing individuals at low risk for contracting legionnaires' disease.

  13. Microbiological diagnosis and molecular typing of Legionella strains during an outbreak of legionellosis in Southern Germany.

    Science.gov (United States)

    Essig, Andreas; von Baum, Heike; Gonser, Theodor; Haerter, Georg; Lück, Christian

    2016-02-01

    An explosive outbreak of Legionnaires' disease with 64 reported cases occurred in Ulm/Neu-Ulm in the South of Germany in December 2009/January 2010 caused by Legionella (L.) pneumophila serogroup 1, monoclonal (mAb) subtype Knoxville, sequence type (ST) 62. Here we present the clinical microbiological results from 51 patients who were diagnosed at the University hospital of Ulm, the results of the environmental investigations and of molecular typing of patients and environmental strains. All 50 patients from whom urine specimens were available were positive for L. pneumophila antigen when an enzyme-linked immunosorbent assay (EIA) was used following concentration of those urine samples that tested initially negative. The sensitivity of the BinaxNow rapid immunographic assay (ICA), after 15 min reading and after 60 min reading were 70% and 84%, respectively. Direct typing confirmed the monoclonal subtype Knoxville in 5 out of 8 concentrated urine samples. Real time PCR testing of respiratory tract specimens for L. pneumophila was positive in 15 out of 25 (60%) patients. Direct nested sequence based typing (nSBT) in some of these samples allowed partial confirmation of ST62. L. pneumophila serogroup 1, monoclonal subtype Knoxville ST62, defined as the epidemic strain was isolated from 8 out of 31 outbreak patients (26%) and from one cooling tower confirming it as the most likely source of the outbreak. While rapid detection of Legionella antigenuria was crucial for the recognition and management of the outbreak, culture and molecular typing of the strains from patients and environmental specimens was the clue for the rapid identification of the source of infection. Copyright © 2016 Elsevier GmbH. All rights reserved.

  14. Prevalence study of Legionella spp. contamination in ferries and cruise ships

    Directory of Open Access Journals (Sweden)

    Masia Maria

    2006-04-01

    Full Text Available Abstract Background In the last years, international traffic volume has significantly increased, raising the risk for acquisition of infectious diseases. Among travel-associated infections, increased incidence of legionellosis has been reported among travellers. Aim of our study was: to describe the frequency and severity of Legionella spp. contamination in ferries and cruise ships; to compare the levels of contamination with those indicated by the Italian ministerial guidelines for control and prevention of legionellosis, in order to assess health risks and to adopt control measures. Method A prevalence study was carried out on 9 ships docked at the seaports of northern Sardinia in 2004. Water samples were collected from critical sites: passenger cabins, crew cabins, kitchens, coffee bars, rooms of the central air conditioning system. It was performed a qualitative and quantitative identification of Legionella spp. and a chemical, physical and bacteriological analysis of water samples. Results Forty-two percent (38/90 water samples were contaminated by Legionella spp.. Positive samples were mainly drawn from showers (24/44, washbasins (10/22. L. pneumophila was isolated in 42/44 samples (95.5%, followed by L. micdadei (4.5%. Strains were identified as L. pneumophila serogroup 6 (45.2%; 19 samples, 2–14 (42.9%, 5 (7.1% and 3 (4.8%. Legionella spp. load was high; 77.8% of the water samples contained > 104 CFU/L. Low residual free chlorine concentration (0–0,2 mg/L was associated to a contamination of the 50% of the water samples. Conclusion Legionella is an ubiquitous bacterium that could create problems for public health. We identified Legionella spp. in 6/7 ferries. Microbial load was predominantly high (> 104 CFU/L or ranging from 103 to 104 CFU/L. It is matter of concern when passengers are subjects at risk because of Legionella spp. is an opportunist that can survive in freshwater systems; high bacterial load might be an important

  15. Legionella cardiaca sp. nov., isolated from a case of native valve endocarditis in a human heart.

    Science.gov (United States)

    Pearce, Meghan M; Theodoropoulos, Nicole; Mandel, Mark J; Brown, Ellen; Reed, Kurt D; Cianciotto, Nicholas P

    2012-12-01

    A Gram-negative, rod-shaped bacterium, designated H63(T), was isolated from aortic valve tissue of a patient with native valve endocarditis. 16S rRNA gene sequencing revealed that H63(T) belongs to the genus Legionella, with its closest neighbours being the type strains of Legionella brunensis (98.8% similarity), L. londiniensis (97.0%), L. jordanis (96.8%), L. erythra (96.2%), L. dresdenensis (96.0%) and L. rubrilucens, L. feeleii, L. pneumophila and L. birminghamensis (95.7%). DNA-DNA hybridization studies yielded values of cardiaca sp. nov. is proposed. The type strain is H63(T) ( = ATCC BAA-2315(T)  = DSM 25049(T)  = JCM 17854(T)).

  16. The machinery at endoplasmic reticulum-plasma membrane contact sites contributes to spatial regulation of multiple Legionella effector proteins.

    Directory of Open Access Journals (Sweden)

    Andree Hubber

    2014-07-01

    Full Text Available The Dot/Icm system of the intracellular pathogen Legionella pneumophila has the capacity to deliver over 270 effector proteins into host cells during infection. Important questions remain as to spatial and temporal mechanisms used to regulate such a large array of virulence determinants after they have been delivered into host cells. Here we investigated several L. pneumophila effector proteins that contain a conserved phosphatidylinositol-4-phosphate (PI4P-binding domain first described in the effector DrrA (SidM. This PI4P binding domain was essential for the localization of effectors to the early L. pneumophila-containing vacuole (LCV, and DrrA-mediated recruitment of Rab1 to the LCV required PI4P-binding activity. It was found that the host cell machinery that regulates sites of contact between the plasma membrane (PM and the endoplasmic reticulum (ER modulates PI4P dynamics on the LCV to control localization of these effectors. Specifically, phosphatidylinositol-4-kinase IIIα (PI4KIIIα was important for generating a PI4P signature that enabled L. pneumophila effectors to localize to the PM-derived vacuole, and the ER-associated phosphatase Sac1 was involved in metabolizing the PI4P on the vacuole to promote the dissociation of effectors. A defect in L. pneumophila replication in macrophages deficient in PI4KIIIα was observed, highlighting that a PM-derived PI4P signature is critical for biogenesis of a vacuole that supports intracellular multiplication of L. pneumophila. These data indicate that PI4P metabolism by enzymes controlling PM-ER contact sites regulate the association of L. pneumophila effectors to coordinate early stages of vacuole biogenesis.

  17. Legionella control in the water system of antiquated hospital buildings by shock and continuous hyperchlorination: 5 years experience

    Science.gov (United States)

    2014-01-01

    Background To control the presence of Legionella in an old hospital water system, an integrated strategy of water disinfection-filtration was implemented in the university hospital Umberto I in Rome. Methods Due to antiquated buildings, hospital water system design and hospital extension (38 buildings), shock hyperchlorination (sodium hypochlorite, 20–50 ppm of free chlorine at distal points for 1–2 h) followed by continuous hyperchlorination (0.5-1.0 mg/L at distal points) were adopted, and microbiological and chemical monitoring of the water supply was carried out in the university hospital (December 2006-December 2011). Results Overall, 1308 samples of cold 45°C (17.8%) water were collected, determining residual free chlorine (0.43 ± 0.44 mg/L), pH (7.43 ± 0.29) and trihalomethanes (8.97 ± 18.56 μg/L). Legionella was isolated in 102 (9.8%) out of 1.041 water samples without filters (L. pneumophila sg 1 17.6%, L. pneumophila sg 2–14 28.4%, L. non pneumophila 53.9%), and in none of the 267 samples with filters. Legionella was recovered in 23 buildings out of 38 and 29 samples (28.4%) exceeded 103 cfu/L. When considering the disinfection treatment Legionella was isolated: before shock hyperchlorination (21.1%), 15 days after shock hyperchlorination (7.8%), 30 days after shock hyperchlorination (3.5%), during continuous hyperchlorination (5.5%) and without continuous hyperchlorination (27.3%). Continuous hyperchlorination following the shock treatment achieved >70% reduction of positive samples, whereas no continuous hyperchlorination after shock treatment was more frequently associated to Legionella isolation (OR 6.41; 95% CI 3.10–13.26; p Legionella isolation were: residual free chlorine Legionella reduction, but effective chlorine levels (>0.5 < 1.0 mg/L) deteriorated water quality (organoleptic and chemical). However, shock and continuous hyperchlorination remains a valid-term option in old buildings with no water system rational design, managing

  18. Detectiemethoden voor legionella in water

    NARCIS (Netherlands)

    Schalk M; de Roda Husman AM; LZO; cib

    2011-01-01

    Het RIVM heeft een overzicht gemaakt van bestaande en nieuwe methoden om legionella in water aan te tonen. Dit is gemaakt in opdracht van de VROM Inspectie, die inzicht wil hebben in de kenmerken en toepassingsmogelijkheden van bestaande en nieuwe methoden.
    Legionella komt overal in het

  19. Colonization of Legionella species in Turkish baths in hotels in Alanya, Turkey.

    Science.gov (United States)

    Erdogan, Haluk; Arslan, Hande

    2015-05-01

    This study evaluated the prevalence of Legionella species in water samples collected from Turkish baths in hotels in Alanya, Turkey, from August 2003 to September 2013. Water samples were collected in 100-mL sterile containers and then concentrated by filtration. Heat treatment was used to eliminate other microorganisms from the samples, which were then spread on Legionella-selective-buffered charcoal yeast extract alpha (BCYE-α) agar and on BCYE-α agar supplemented with glycine, vancomycin, polymyxin, and cycloheximide. Cysteine-dependent colonies were identified by latex agglutination. In total, 135 samples from 52 hotels with Turkish baths were evaluated. Legionella species were identified in 11/52 (21.2%) hotels and 18/135 (13.3%) samples. The most frequently isolated species was Legionella pneumophila, with most isolates belonging to serogroups 6 (55.6%) and 1 (22.2%). The colony count was 1000 CFU mL(-1) in three samples. These findings suggest that the hot water systems of Turkish baths in hotels must be viewed as a possible source of travel-associated Legionnaires' disease, and preventative measures should be put in place.

  20. Current and Emerging Legionella Diagnostics for Laboratory and Outbreak Investigations

    Science.gov (United States)

    Mercante, Jeffrey W.

    2015-01-01

    SUMMARY Legionnaires' disease (LD) is an often severe and potentially fatal form of bacterial pneumonia caused by an extensive list of Legionella species. These ubiquitous freshwater and soil inhabitants cause human respiratory disease when amplified in man-made water or cooling systems and their aerosols expose a susceptible population. Treatment of sporadic cases and rapid control of LD outbreaks benefit from swift diagnosis in concert with discriminatory bacterial typing for immediate epidemiological responses. Traditional culture and serology were instrumental in describing disease incidence early in its history; currently, diagnosis of LD relies almost solely on the urinary antigen test, which captures only the dominant species and serogroup, Legionella pneumophila serogroup 1 (Lp1). This has created a diagnostic “blind spot” for LD caused by non-Lp1 strains. This review focuses on historic, current, and emerging technologies that hold promise for increasing LD diagnostic efficiency and detection rates as part of a coherent testing regimen. The importance of cooperation between epidemiologists and laboratorians for a rapid outbreak response is also illustrated in field investigations conducted by the CDC with state and local authorities. Finally, challenges facing health care professionals, building managers, and the public health community in combating LD are highlighted, and potential solutions are discussed. PMID:25567224

  1. Experimental human-like model to assess the part of viable Legionella reaching the thoracic region after nebulization.

    Directory of Open Access Journals (Sweden)

    Jérémie Pourchez

    Full Text Available The incidence of Legionnaires' disease (LD in European countries and the USA has been constantly increasing since 1998. Infection of humans occurs through aerosol inhalation. To bridge the existing gap between the concentration of Legionella in a water network and the deposition of bacteria within the thoracic region (assessment of the number of viable Legionella, we validated a model mimicking realistic exposure through the use of (i recent technology for aerosol generation and (ii a 3D replicate of the human upper respiratory tract. The model's sensitivity was determined by monitoring the deposition of (i aerosolized water and Tc99m radio-aerosol as controls, and (ii bioaerosols generated from both Escherichia coli and Legionella pneumophila sg 1 suspensions. The numbers of viable Legionella prior to and after nebulization were provided by culture, flow cytometry and qPCR. This study was designed to obtain more realistic data on aerosol inhalation (vs. animal experimentation and deposition at the thoracic region in the context of LD. Upon nebulization, 40% and 48% of the initial Legionella inoculum was made of cultivable and non-cultivable cells, respectively; 0.7% of both populations reached the filter holder mimicking the thoracic region in this setup. These results are in agreement with experimental data based on quantitative microbial risk assessment methods and bring new methods that may be useful for preventing LD.

  2. Controlling Legionella in hospital water systems: experience with the superheat-and-flush method and copper-silver ionization.

    Science.gov (United States)

    Stout, J E; Lin, Y S; Goetz, A M; Muder, R R

    1998-12-01

    To evaluate the effect of copper-silver ionization on Legionella colonization and nosocomial legionnaires' disease and to compare the efficacy of metal ions versus the superheat-and-flush method of disinfection. Prospective determination over a 36-month period of copper and silver ion concentrations in the recirculating hot-water system, Legionella colonization of the hospital water distribution system, and cases of nosocomial legionnaires' disease. Retrospective comparison of results with the previous 13 years, during which the superheat-and-flush method was used. The Pittsburgh Veterans' Affairs Health Care System (University Drive Division) acute-care hospital. Three copper-silver ionization systems were installed on the hot-water distribution system in November 1994. The average number of cases of legionnaires' disease per year and the percentage of distal sites positive for Legionella pneumophila for the superheat-and-flush method versus the copper-silver ionization method was six cases with 15% positivity versus two cases with 4% positivity, respectively. The reduction in Legionella colonization after copper-silver ionization was significant (Pcopper and silver ion concentrations (mg/L) were 0.29 and 0.054 from hot-water tanks, and 0.17 and 0.04 from distal outlets, respectively. We conclude that a properly maintained and monitored copper-silver ionization system was more effective than the superheat-and-flush method for reducing the recovery of Legionella from the hospital water distribution system.

  3. Simultaneous detection of Legionella species and L. anisa, L. bozemanii, L. longbeachae and L. micdadei using conserved primers and multiple probes in a multiplex real-time PCR assay.

    Science.gov (United States)

    Cross, Kristen E; Mercante, Jeffrey W; Benitez, Alvaro J; Brown, Ellen W; Diaz, Maureen H; Winchell, Jonas M

    2016-07-01

    Legionnaires' disease is a severe respiratory disease that is estimated to cause between 8,000 and 18,000 hospitalizations each year, though the exact burden is unknown due to under-utilization of diagnostic testing. Although Legionella pneumophila is the most common species detected in clinical cases (80-90%), other species have also been reported to cause disease. However, little is known about Legionnaires' disease caused by these non-pneumophila species. We designed a multiplex real-time PCR assay for detection of all Legionella spp. and simultaneous specific identification of four clinically-relevant Legionella species, L. anisa, L. bozemanii, L. longbeachae, and L. micdadei, using 5'-hydrolysis probe real-time PCR. The analytical sensitivity for detection of nucleic acid from each target species was ≤50fg per reaction. We demonstrated the utility of this assay in spiked human sputum specimens. This assay could serve as a tool for understanding the scope and impact of non-pneumophila Legionella species in human disease. Published by Elsevier Inc.

  4. Legionella contamination in hot water systems of hospitals, nursing homes, hotels, factories and spas in Tuscany-Italy

    Directory of Open Access Journals (Sweden)

    Antonella Lo Nostro

    2011-03-01

    Full Text Available

    Abstract Following the report of many cases of Legionnaires’ disease associated with accommodation facilities such as hotels, spas, workplaces, hospitals and nursing homes, we verified if Legionella pneumophila and Legionella spp. were present in some of those structures in Tuscany, in order to estimate the species and serogroups in circulation. Legionella pneumophila serogroup 1 (30.9% was the most frequently isolated species along with serogroups 3 (16.1% and 6 (13.3%; these three serogroups are identified, in literature, as those most responsible for Legionnaires’ disease (LD. Studying all analyzed structures, we found some parts of the water system where Legionella concentration was higher than 103CFU/L, indicated, in Italy, as the maximum admitted concentration value above which a decontamination treatment is necessary when one or more cases of healthcare-acquired Legionnaires’ disease are observed. Moreover disinfection is recommended in any case when counts exceed 104CFU/L.
    Consequently, in order to prevent cases of Legionnaires’ disease, a continuous surveillance of the water
    systems of all accommodation facilities is necessary, with particular attention to hospitals and nursing
    homes where immunocompromised patients lodge, so as to promptly estimate the presence of the pathogen and consequently plan the most suitable intervention activities. We concluded that, in any structure, a continuous surveillance and disinfecting treatment of water systems is necessary. Moreover, after any disinfection treatment the temperature of the hot water flowing in the system must be necessarily maintained near 51°C in order to minimize the probability of recontamination from Legionella and limit the
    risk of LD in consumers.

  5. MultipleLegionella pneumophilaeffector virulence phenotypes revealed through high-throughput analysis of targeted mutant libraries.

    Science.gov (United States)

    Shames, Stephanie R; Liu, Luying; Havey, James C; Schofield, Whitman B; Goodman, Andrew L; Roy, Craig R

    2017-11-28

    Legionella pneumophila is the causative agent of a severe pneumonia called Legionnaires' disease. A single strain of L. pneumophila encodes a repertoire of over 300 different effector proteins that are delivered into host cells by the Dot/Icm type IV secretion system during infection. The large number of L. pneumophila effectors has been a limiting factor in assessing the importance of individual effectors for virulence. Here, a transposon insertion sequencing technology called INSeq was used to analyze replication of a pool of effector mutants in parallel both in a mouse model of infection and in cultured host cells. Loss-of-function mutations in genes encoding effector proteins resulted in host-specific or broad virulence phenotypes. Screen results were validated for several effector mutants displaying different virulence phenotypes using genetic complementation studies and infection assays. Specifically, loss-of-function mutations in the gene encoding LegC4 resulted in enhanced L. pneumophila in the lungs of infected mice but not within cultured host cells, which indicates LegC4 augments bacterial clearance by the host immune system. The effector proteins RavY and Lpg2505 were important for efficient replication within both mammalian and protozoan hosts. Further analysis of Lpg2505 revealed that this protein functions as a metaeffector that counteracts host cytotoxicity displayed by the effector protein SidI. Thus, this study identified a large cohort of effectors that contribute to L. pneumophila virulence positively or negatively and has demonstrated regulation of effector protein activities by cognate metaeffectors as being critical for host pathogenesis.

  6. The Legionella micdadei flagellin

    DEFF Research Database (Denmark)

    Bangsborg, Jette Marie; Hindersson, P; Shand, G

    1995-01-01

    carried a L. micdadei DNA insert of 5 kb, containing ca 95% of the fla gene. The complete DNA sequence of the L. micdadei fla gene was obtained by combining sequence data from pHI5588 with results using a polymerase chain reaction-based system for genome walking (vectorette PCR). The L. micdadei fla gene...... shared a high degree of homology with other flagellin genes in the amino- and carboxy termini, whereas the central region was found to be nonconserved. The fla sequence will facilitate the cloning of Fla proteins from other Legionella species and the study of flagella in the pathogenesis of Legionnaires...

  7. Combined use of real-time PCR and nested sequence-based typing in survey of human Legionella infection.

    Science.gov (United States)

    Qin, T; Zhou, H; Ren, H; Shi, W; Jin, H; Jiang, X; Xu, Y; Zhou, M; Li, J; Wang, J; Shao, Z; Xu, X

    2016-07-01

    Legionnaires' disease (LD) is a globally distributed systemic infectious disease. The burden of LD in many regions is still unclear, especially in Asian countries including China. A survey of Legionella infection using real-time PCR and nested sequence-based typing (SBT) was performed in two hospitals in Shanghai, China. A total of 265 bronchoalveolar lavage fluid (BALF) specimens were collected from hospital A between January 2012 and December 2013, and 359 sputum specimens were collected from hospital B throughout 2012. A total of 71 specimens were positive for Legionella according to real-time PCR focusing on the 5S rRNA gene. Seventy of these specimens were identified as Legionella pneumophila as a result of real-time PCR amplification of the dotA gene. Results of nested SBT revealed high genetic polymorphism in these L. pneumophila and ST1 was the predominant sequence type. These data revealed that the burden of LD in China is much greater than that recognized previously, and real-time PCR may be a suitable monitoring technology for LD in large sample surveys in regions lacking the economic and technical resources to perform other methods, such as urinary antigen tests and culture methods.

  8. PCR method for the rapid detection and discrimination of Legionella spp. based on the amplification of pcs, pmtA, and 16S rRNA genes.

    Science.gov (United States)

    Janczarek, Monika; Palusińska-Szysz, Marta

    2016-05-01

    Legionella bacteria are organisms of public health interest due to their ability to cause pneumonia (Legionnaires' disease) in susceptible humans and their ubiquitous presence in water supply systems. Rapid diagnosis of Legionnaires' disease allows the use of therapy specific for the disease. L. pneumophila serogroup 1 is the most common cause of infection acquired in community and hospital environments. The non-L. pneumophila infections are likely under-detected because of a lack of effective diagnosis. In this work, simplex and duplex PCR assays with the use of new molecular markers pcs and pmtA involved in phosphatidylcholine synthesis were specified for rapid and cost-efficient identification and distinguishing Legionella species. The sets of primers developed were found to be sensitive and specific for reliable detection of Legionella belonging to the eight most clinically relevant species. Among these, four primer sets I, II, VI, and VII used for duplex-PCRs proved to have the highest identification power and reliability in the detection of the bacteria. Application of this PCR-based method should improve detection of Legionella spp. in both clinical and environmental settings and facilitate molecular typing of these organisms.

  9. Molecular Survey of the Occurrence of Legionella spp., Mycobacterium spp., Pseudomonas aeruginosa, and Amoeba Hosts in Two Chloraminated Drinking Water Distribution Systems

    Science.gov (United States)

    Wang, Hong; Edwards, Marc; Falkinham, Joseph O.

    2012-01-01

    The spread of opportunistic pathogens via public water systems is of growing concern. The purpose of this study was to identify patterns of occurrence among three opportunistic pathogens (Legionella pneumophila, Mycobacterium avium, and Pseudomonas aeruginosa) relative to biotic and abiotic factors in two representative chloraminated drinking water distribution systems using culture-independent methods. Generally, a high occurrence of Legionella (≥69.0%) and mycobacteria (100%), lower occurrence of L. pneumophila (≤20%) and M. avium (≤33.3%), and rare detection of Pseudomonas aeruginosa (≤13.3%) were observed in both systems according to quantitative PCR. Also, Hartmanella vermiformis was more prevalent than Acanthamoeba, both of which are known hosts for opportunistic pathogen amplification, the latter itself containing pathogenic members. Three-minute flushing served to distinguish distribution system water from plumbing in buildings (i.e., premise plumbing water) and resulted in reduced numbers of copies of Legionella, mycobacteria, H. vermiformis, and 16S rRNA genes (P Legionella and H. vermiformis, were noted, emphasizing potential microbial ecological relationships. Overall, the results provide insight into factors that may aid in controlling opportunistic pathogen proliferation in real-world water systems. PMID:22752174

  10. Prevalence and distribution of Legionella spp in potable water systems in Germany, risk factors associated with contamination, and effectiveness of thermal disinfection.

    Science.gov (United States)

    Kruse, Eva-Brigitta; Wehner, Arno; Wisplinghoff, Hilmar

    2016-04-01

    Worldwide, Legionella spp are a common cause of community-acquired pneumonia. Potable water systems are a main reservoir; however, exposure in the community is unknown. Water samples from 718 buildings in Germany were collected. Possible risk factors were prospectively recorded. All samples were tested for Legionella spp using cultural microbiologic methods. Samples were assigned to 1 of 5 levels of contamination. Statistical analysis was performed to determine the influence of risk factors for contamination and, in a subgroup of buildings, for unsuccessful thermal disinfection. In total, 4,482 water samples from 718 different water supply systems were analyzed. In 233 buildings (32.7%), Legionella spp were identified, 148 (63.5%) of which had a medium or higher level of contamination. The most common species was Legionella pneumophila (94%). Contamination was strongly associated with temperature in the circulation, but not with the size of the building, time of the year, or transport time to the laboratory. Thermal disinfection was successful in fewer than half of the buildings. There is relevant exposure to Legionella spp in the community. Water systems are not always up to current technical standards. Although microbiological risk assessment remains a challenge, there is a case for monitoring for Legionella spp outside of hospitals. Copyright © 2016 Association for Professionals in Infection Control and Epidemiology, Inc. Published by Elsevier Inc. All rights reserved.

  11. Analysis of the genetic diversity of Legionella by sequencing the 23S-5S ribosomal intergenic spacer region: from phylogeny to direct identification of isolates at the species level from clinical specimens.

    Science.gov (United States)

    Grattard, Florence; Ginevra, Christophe; Riffard, Serge; Ros, Alain; Jarraud, Sophie; Etienne, Jerome; Pozzetto, Bruno

    2006-01-01

    This study focuses on the interest of the hypervariable 23S-5S ribosomal intergenic spacer region (ISR) of the genus Legionella to analyze the phylogenic diversity of Legionella at the species and subspecies levels and to identify isolates directly from clinical specimens. The method, using a real-time PCR assay with a single primer pair followed by sequencing, was able to identify correctly 49 reference strains of Legionella belonging to 37 different species, including those implicated in human infections, and to clearly differentiate the three subspecies of L. pneumophila. Based on sequence similarities, the 23S-5S ISR sequences were much more variable than the rpoB and mip sequences (P5S ISR method was able to cluster Legionella species in accordance with phenotypic traits, such as autofluorescence or fatty acid membrane composition. Using maximum parsimony methods, the rpoB and 23S-5S ISR data sets were shown to be incongruent (P5S ISR and the mip data sets were found to be congruent (P=0.313), suggesting the interest of combining these two regions to demonstrate phylogenetic links between Legionella species. This molecular assay was shown able to both detect Legionella DNA directly in respiratory specimens from patients exhibiting a Legionella infection and provide accurate identification of the bacterium at the species level in the tested specimens. These properties open a wide range of applications to the 23S-5S ISR sequencing method, from taxonomic analyses to clinical and epidemiological investigations.

  12. Induction of caspase 3 activation by multiple L. pneumophila Dot/Icm substrates

    Science.gov (United States)

    Zhu, Wenhan; Hammad, Loubna A.; Hsu, FoSheng; Mao, Yuxin; Luo, Zhao-Qing

    2013-01-01

    The intracellular pathogen Legionella pneumophila is able to strike a balance between the death and survival of the host cell during infection. Despite the presence of high level of active caspase-3, the executioner caspase of apoptotic cell death, infected permissive macrophages are markedly resistant to exogenous apoptotic stimuli. Several bacterial molecules capable of promoting the cell survival pathways have been identified, but proteins involved in the activation of caspase-3 remain unknown. To study the mechanism of L. pneumophila-mediated caspase-3 activation, we tested all known Dot/Icm substrates for their ability to activate caspase-3. Five effectors capable of causing caspase-3 activation upon transient expression were identified. Among these, by using its ability to activate caspase-3 by inducing the release of cytochrome c from the mitochondria, we demonstrated that VipD is a phospholipase A2, which hydrolyzes phosphatidylethanolamine (PE) and phosphocholine (PC) on the mitochondrial membrane in a manner that appears to require host co-factor(s). The lipase activity leads to the production of free fatty acids and 2-lysophospholipids, which destabilize the mitochondrial membrane and may contribute to the release of cytochrome c and the subsequent caspase 3 activation. Furthermore, we found that whereas it is not detectably defectively in caspase 3 activation in permissive cells, a mutant lacking all of these five genes is less potent in inducing apoptosis in dendritic cells. Our results reveal that activation of host cell death pathways by L. pneumophila is a result of the effects of multiple bacterial proteins with diverse biochemical functions. PMID:23773455

  13. Prevalence and Molecular Characteristics of Waterborne Pathogen Legionella in Industrial Cooling Tower Environments

    Directory of Open Access Journals (Sweden)

    Lijie Li

    2015-10-01

    Full Text Available Cooling towers are a source of Legionnaires’ disease. It is important from a public health perspective to survey industrial cooling towers for the presence of Legionella. Prospective surveillance of the extent of Legionella pollution was conducted at factories in Shijiazhuang, China between March 2011 and September 2012. Overall, 35.7% of 255 industrial cooling tower water samples showed Legionella-positive, and their concentrations ranged from 100 Colony-Forming Units (CFU/liter to 88,000 CFU/liter, with an average concentration of 9100 CFU/liter. A total of 121 isolates were obtained. All isolates were L. pneumophila, and the isolated serogroups included serogroups 1 (68 isolates, 56.2%, 6 (25, 20.7%, 5 (12, 9.9%, 8 (8, 6.6%, 3 (6, 5.0% and 9 (2, 1.6%. All 121 isolates were analyzed by pulsed-field gel electrophoresis (PFGE and 64 different patterns were obtained. All 121 isolates were analyzed sequence-based typing (SBT, a full 7-allele profile was obtained from 117 isolates. One hundred and seventeen isolates were divided into 49 sequence types. Two virulence genes, lvh and rtxA, are analyzed by polymerase chain reaction (PCR. 92.6% (112/121 and 98.3% (119/121 isolates carried lvh and rtxA respectively and 90.9% (110/121 of tested isolates carried both genes. Our results demonstrated high prevalence and genetic polymorphism of L. pneumophila in industrial cooling tower environments in Shijiazhang, China, and the SBT and virulence gene PCR results suggested that the isolates were pathogenic. Improved control and prevention strategies are urgently needed.

  14. Legionella Effector AnkX Disrupts Host Cell Endocytic Recycling in a Phosphocholination-Dependent Manner

    Directory of Open Access Journals (Sweden)

    Samual C. Allgood

    2017-09-01

    Full Text Available The facultative intracellular bacterium Legionella pneumophila proliferates within amoebae and human alveolar macrophages, and it is the causative agent of Legionnaires' disease, a life-threatening pneumonia. Within host cells, L. pneumophila establishes a replicative haven by delivering numerous effector proteins into the host cytosol, many of which target membrane trafficking by manipulating the function of Rab GTPases. The Legionella effector AnkX is a phosphocholine transferase that covalently modifies host Rab1 and Rab35. However, a detailed understanding of the biological consequence of Rab GTPase phosphocholination remains elusive. Here, we broaden the understanding of AnkX function by presenting three lines of evidence that it interferes with host endocytic recycling. First, using immunogold transmission electron microscopy, we determined that GFP-tagged AnkX ectopically produced in mammalian cells localizes at the plasma membrane and tubular membrane compartments, sites consistent with targeting the endocytic recycling pathway. Furthermore, the C-terminal region of AnkX was responsible for association with the plasma membrane, and we determined that this region was also able to bind the phosphoinositide lipids PI(3P and PI(4P in vitro. Second, we observed that mCherry-AnkX co-localized with Rab35, a regulator of recycling endocytosis and with major histocompatibility class I protein (MHC-I, a key immunoregulatory protein whose recycling from and back to the plasma membrane is Rab35-dependent. Third, we report that during infection of macrophages, AnkX is responsible for the disruption of endocytic recycling of transferrin, and AnkX's phosphocholination activity is critical for this function. These results support the hypothesis that AnkX targets endocytic recycling during host cell infection. Finally, we have demonstrated that the phosphocholination activity of AnkX is also critical for inhibiting fusion of the Legionella

  15. Molecular Detection of Legionella spp. and their associations with Mycobacterium spp., Pseudomonas aeruginosa and amoeba hosts in a drinking water distribution system.

    Science.gov (United States)

    Lu, J; Struewing, I; Vereen, E; Kirby, A E; Levy, K; Moe, C; Ashbolt, N

    2016-02-01

    This study investigated waterborne opportunistic pathogens (OPs) including potential hosts, and evaluated the use of Legionella spp. for indicating microbial water quality for OPs within a full-scale operating drinking water distribution system (DWDS). To investigate the occurrence of specific microbial pathogens within a major city DWDS we examined large volume (90 l drinking water) ultrafiltration (UF) concentrates collected from six sites between February, 2012 and June, 2013. The detection frequency and concentration estimates by qPCR were: Legionella spp. (57%/85 cell equivalent, CE l(-1) ), Mycobacterium spp. (88%/324 CE l(-1) ), Pseudomonas aeruginosa (24%/2 CE l(-1) ), Vermamoeba vermiformis (24%/2 CE l(-1) ) and Acanthamoeba spp. (42%/5 cyst equivalent, CE l(-1) ). There was no detection of the following microorganisms: human faecal indicator Bacteroides (HF183), Salmonella enterica, Campylobacter spp., Escherichia coli O157:H7, Giardia intestinalis, Cryptosporidium spp. or Naegleria fowleri. There were significant correlations between the qPCR signals of Legionella spp. and Mycobacterium spp., and their potential hosts V. vermiformis and Acanthamoeba spp. Sequencing of Legionella spp. demonstrated limited diversity, with most sequences coming from two dominant groups, of which the larger dominant group was an unidentified species. Other known species including Legionella pneumophila were detected, but at low frequency. The densities of Legionella spp. and Mycobacterium spp. were generally higher (17 and 324 folds, respectively) for distal sites relative to the entry point to the DWDS. Legionella spp. occurred, had significant growth and were strongly associated with free-living amoebae (FLA) and Mycobacterium spp., suggesting that Legionella spp. could provide a useful DWDS monitoring role to indicate potential conditions for non-faecal OPs. The results provide insight into microbial pathogen detection that may aid in the monitoring of microbial water

  16. Detection of Legionella by quantitative-polymerase chain reaction (qPCR) for monitoring and risk assessment

    DEFF Research Database (Denmark)

    Krøjgaard, Louise H.; Krogfelt, Karen A.; Albrechtsen, Hans-Jorgen

    2011-01-01

    Background: Culture and quantitative polymerase chain reaction (qPCR) assays for the detection of Legionella were compared on samples from a residential area before and after two interventions. A total of 84 samples were collected from shower hoses and taps as first flush samples and at constant...... temperature. Samples were grouped according to the origin of the sample, a) circulation water b) water from empty apartments c) water from shower hoses. The aims were to investigate the usefulness of qPCR compared to culture for monitoring remedial actions for elimination of Legionella bacteria and as a tool...... detection limit (> 10 CFU/L) to 1.6*10(6) CFU/L. In circulating water and in first flush water from shower hoses, culture and qPCR showed the same tendencies. The overall correlation between the bacteria number detected by culture and the two developed qPCR assays (L. spp and L. pneumophila) was relatively...

  17. Anti-Legionella dumoffii Activity of Galleria mellonella Defensin and Apolipophorin III

    Directory of Open Access Journals (Sweden)

    Małgorzata Cytryńska

    2012-12-01

    Full Text Available The gram-negative bacterium Legionella dumoffii is, beside Legionella pneumophila, an etiological agent of Legionnaires’ disease, an atypical form of pneumonia. The aim of this study was to determine the antimicrobial activity of Galleria mellonella defense polypeptides against L. dumoffii. The extract of immune hemolymph, containing a mixture of defense peptides and proteins, exhibited a dose-dependent bactericidal effect on L. dumoffii. The bacterium appeared sensitive to a main component of the hemolymph extract, apolipophorin III, as well as to a defense peptide, Galleria defensin, used at the concentrations 0.4 mg/mL and 40 μg/mL, respectively. L. dumoffii cells cultured in the presence of choline were more susceptible to both defense factors analyzed. A transmission electron microscopy study of bacterial cells demonstrated that Galleria defensin and apolipophorin III induced irreversible cell wall damage and strong intracellular alterations, i.e., increased vacuolization, cytoplasm condensation and the appearance of electron-white spaces in electron micrographs. Our findings suggest that insects, such as G. mellonella, with their great diversity of antimicrobial factors, can serve as a rich source of compounds for the testing of Legionella susceptibility to defense-related peptides and proteins.

  18. Anti-Legionella dumoffii Activity of Galleria mellonella Defensin and Apolipophorin III

    Science.gov (United States)

    Palusińska-Szysz, Marta; Zdybicka-Barabas, Agnieszka; Pawlikowska-Pawlęga, Bożena; Mak, Pawel; Cytryńska, Małgorzata

    2012-01-01

    The gram-negative bacterium Legionella dumoffii is, beside Legionella pneumophila, an etiological agent of Legionnaires’ disease, an atypical form of pneumonia. The aim of this study was to determine the antimicrobial activity of Galleria mellonella defense polypeptides against L. dumoffii. The extract of immune hemolymph, containing a mixture of defense peptides and proteins, exhibited a dose-dependent bactericidal effect on L. dumoffii. The bacterium appeared sensitive to a main component of the hemolymph extract, apolipophorin III, as well as to a defense peptide, Galleria defensin, used at the concentrations 0.4 mg/mL and 40 μg/mL, respectively. L. dumoffii cells cultured in the presence of choline were more susceptible to both defense factors analyzed. A transmission electron microscopy study of bacterial cells demonstrated that Galleria defensin and apolipophorin III induced irreversible cell wall damage and strong intracellular alterations, i.e., increased vacuolization, cytoplasm condensation and the appearance of electron-white spaces in electron micrographs. Our findings suggest that insects, such as G. mellonella, with their great diversity of antimicrobial factors, can serve as a rich source of compounds for the testing of Legionella susceptibility to defense-related peptides and proteins. PMID:23235329

  19. Isolation of Legionella species/serogroups from water cooling systems compared with potable water systems in Spanish healthcare facilities.

    Science.gov (United States)

    Rivera, J-M; Aguilar, L; Granizo, J J; Vos-Arenilla, A; Giménez, M-J; Aguiar, J-M; Prieto, J

    2007-12-01

    Surveillance of Legionella spp. in hospital water systems was performed in forty-four inpatient healthcare facilities in Spain during 2005-2006. A total of 2,341 samples were collected: 470 from cooling systems (cooling towers) and 1,871 from potable water systems. The latter included 211 from cold-water tanks and 260 from hot-water tanks, totalling 471 from central water reservoirs 136 from showers, 1,172 from unfiltered taps and 92 from filtered taps, totalling 1,400 from peripheral points. Temperature, chlorine levels and the presence of Legionella spp. were determined. In all, 373 (15.9%) samples yielded Legionella spp. Significantly higher isolation rates were obtained from cooling towers (23.8%) versus cold- and hot-water tanks (approximately 4.7%), due to the significantly higher number of samples positive for serogroup 1 (19.4 vs 0.9-3.5%). In potable water systems, no differences were found between central water tanks and showers, but significant differences in isolation rates between central water tanks and unfiltered taps were observed (4.7 vs 19.6%) due to differences in non-serogroup 1 L. pneumophila. Filters significantly decreased isolation rates of these serotypes (11 vs 0%). Some seasonal differences were noted, with higher isolation rates in summer for legionella serogroup 1 in cooling systems and for L. pneumophila serogroups 2-14 in potable water systems. In regression models, higher temperatures were associated with colonisation in cooling systems, while lower chlorine levels were associated with colonisation in potable water systems.

  20. Modelling characteristics to predict Legionella contamination risk - Surveillance of drinking water plumbing systems and identification of risk areas.

    Science.gov (United States)

    Völker, Sebastian; Schreiber, Christiane; Kistemann, Thomas

    2016-01-01

    For the surveillance of drinking water plumbing systems (DWPS) and the identification of risk factors, there is a need for an early estimation of the risk of Legionella contamination within a building, using efficient and assessable parameters to estimate hazards and to prioritize risks. The precision, accuracy and effectiveness of ways of estimating the risk of higher Legionella numbers (temperature, stagnation, pipe materials, etc.) have only rarely been empirically assessed in practice, although there is a broad consensus about the impact of these risk factors. We collected n = 807 drinking water samples from 9 buildings which had had Legionella spp. occurrences of >100 CFU/100mL within the last 12 months, and tested for Legionella spp., L. pneumophila, HPC 20°C and 36°C (culture-based). Each building was sampled for 6 months under standard operating conditions in the DWPS. We discovered high variability (up to 4 log(10) steps) in the presence of Legionella spp. (CFU/100 mL) within all buildings over a half year period as well as over the course of a day. Occurrences were significantly correlated with temperature, pipe length measures, and stagnation. Logistic regression modelling revealed three parameters (temperature after flushing until no significant changes in temperatures can be obtained, stagnation (low withdrawal, qualitatively assessed), pipe length proportion) to be the best predictors of Legionella contamination (>100 CFU/100 mL) at single outlets (precision = 66.7%; accuracy = 72.1%; F(0.5) score = 0.59). Copyright © 2015 Elsevier GmbH. All rights reserved.

  1. Legionella control in the water system of antiquated hospital buildings by shock and continuous hyperchlorination: 5 years experience.

    Science.gov (United States)

    Orsi, Giovanni Battista; Vitali, Matteo; Marinelli, Lucia; Ciorba, Veronica; Tufi, Daniela; Del Cimmuto, Angela; Ursillo, Paolo; Fabiani, Massimo; De Santis, Susi; Protano, Carmela; Marzuillo, Carolina; De Giusti, Maria

    2014-07-16

    To control the presence of Legionella in an old hospital water system, an integrated strategy of water disinfection-filtration was implemented in the university hospital Umberto I in Rome. Due to antiquated buildings, hospital water system design and hospital extension (38 buildings), shock hyperchlorination (sodium hypochlorite, 20-50 ppm of free chlorine at distal points for 1-2 h) followed by continuous hyperchlorination (0.5-1.0 mg/L at distal points) were adopted, and microbiological and chemical monitoring of the water supply was carried out in the university hospital (December 2006-December 2011). Overall, 1308 samples of cold 37.7%) and hot >45°C (17.8%) water were collected, determining residual free chlorine (0.43 ± 0.44 mg/L), pH (7.43 ± 0.29) and trihalomethanes (8.97 ± 18.56 μg/L). Legionella was isolated in 102 (9.8%) out of 1.041 water samples without filters (L. pneumophila sg 1 17.6%, L. pneumophila sg 2-14 28.4%, L. non pneumophila 53.9%), and in none of the 267 samples with filters. Legionella was recovered in 23 buildings out of 38 and 29 samples (28.4%) exceeded 103 cfu/L. When considering the disinfection treatment Legionella was isolated: before shock hyperchlorination (21.1%), 15 days after shock hyperchlorination (7.8%), 30 days after shock hyperchlorination (3.5%), during continuous hyperchlorination (5.5%) and without continuous hyperchlorination (27.3%). Continuous hyperchlorination following the shock treatment achieved >70% reduction of positive samples, whereas no continuous hyperchlorination after shock treatment was more frequently associated to Legionella isolation (OR 6.41; 95% CI 3.10-13.26; p chlorine 37 - 123.2; p chlorine levels (>0.5 < 1.0 mg/L) deteriorated water quality (organoleptic and chemical). However, shock and continuous hyperchlorination remains a valid-term option in old buildings with no water system rational design, managing problems due to hospital extension and absence of a proper hot water recirculation

  2. DIAGNOSTICO SEROLÓGICO DE NEUMONIA POR LEGIONELLA. INCIDENCIA EN UN PERIODO DE TRES AÑOS EN EL AREA SANITARIA OESTE DE VALLADOLID

    Directory of Open Access Journals (Sweden)

    M.A. Mazón

    2005-12-01

    Full Text Available ABSTRACT:Introduction: Legionella species are Gram-negative bacilli. There are described for the first time in 70's associated to the Legionnarie's disease. Presently, more than 34 species of Legionella have been identified, 20 of which have been found pathogenic for the man. The clinical manifestations of Legionella infections are primarily respiratory. The most common presentation is acute pneumonia, which varies in severity from mild illness that does not require hospitalization to fatal pneumonia.Material and Methods: The results corresponding to the determination of antibodies to L. pneumophila (Gull Laboratories by indirect inmunofluorescence (IFI have been analysed and reviewed. This conventional inmunofluorescent test measures antibodies versus a pool of 1, 2, 3, 4, 5, 6, 7 and 8 serogroups. During the 3 years of surveillance period 1249 determinations by IFI were performed in our Hospital. The determination of urinary antigen of Legionella for the qualitative detection of the antigen to L. pneumophila serogroup 1 was made in almost all studied samples by using a fast inmunochromatographical technique (Binax NOW.Results and Discussion: Of the 1249 cases of nosocomial pneumonia (diagnosed by a positive result of IFI Legionella test recorded during the period, 12 cases met the criteria for infection with L. pneumophila serogroup 1. Later, clinical histories were reviewed and it was observed that the ages of the confirmed cases oscillated between the 40 and 87 years, with an average of age of 40, of which 8 were men and 4 women. From the point of view of factors of risk, 6 patients were smokers. Clinically, the displayed symptoms were general malaise, fever high and pattern of pulmonary infiltration in all the cases, migraine in 3 cases, diarrhoea in 5 cases and arthralgias and muscle aches in 3 cases. The analytical parameters in common were high VSG and leukocytosis. Hyponatremia 1/1024, what it demonstrates a clear seroconversion. In

  3. Longitudinal Evaluation of the Efficacy of Heat Treatment Procedures against Legionella spp. in Hospital Water Systems by Using a Flow Cytometric Assay▿

    Science.gov (United States)

    Allegra, Severine; Grattard, Florence; Girardot, Françoise; Riffard, Serge; Pozzetto, Bruno; Berthelot, Philippe

    2011-01-01

    Legionella spp. are frequently isolated in hospital water systems. Heat shock (30 min at 70°C) is recommended by the World Health Organization to control its multiplication. The aim of the study was to evaluate retrospectively the efficacy of heat treatments by using a flow cytometry assay (FCA) able to identify viable but nonculturable (VBNC) cells. The study included Legionella strains (L. pneumophila [3 clusters] and L. anisa [1 cluster]) isolated from four hot water circuits of different hospital buildings in Saint-Etienne, France, during a 20-year prospective surveillance. The strains recovered from the different circuits were not epidemiologically related, but the strains isolated within a same circuit over time exhibited an identical genotypic profile. After an in vitro treatment of 30 min at 70°C, the mean percentage of viable cells and VBNC cells varied from 4.6% to 71.7%. The in vitro differences in heat sensitivity were in agreement with the observed efficacy of preventive and corrective heating measures used to control water contamination. These results suggest that Legionella strains can become heat resistant after heating treatments for a long time and that flow cytometry could be helpful to check the efficacy of heat treatments on Legionella spp. and to optimize the decontamination processes applied to water systems for the control of Legionella proliferation. PMID:21183641

  4. Occurrence of Legionella in hot water systems of single-family residences in suburbs of two German cities with special reference to solar and district heating.

    Science.gov (United States)

    Mathys, Werner; Stanke, Juliane; Harmuth, Margarita; Junge-Mathys, Elisabeth

    2008-03-01

    A total of 452 samples from hot water systems of randomly selected single family residences in the suburbs of two German cities were analysed for the occurrence of Legionella. Technical data were documented using a standardized questionnaire to evaluate possible factors promoting the growth of the bacterium in these small plumbing systems. All houses were supplied with treated groundwater from public water works. Drinking water quality was within the limits specified in the German regulations for drinking water and the water was not chlorinated. The results showed that plumbing systems in private houses that provided hot water from instantaneous water heaters were free of Legionella compared with a prevalence of 12% in houses with storage tanks and recirculating hot water where maximum counts of Legionella reached 100,000 CFU/100ml. The presence of L. pneumophila accounted for 93.9% of all Legionella positive specimens of which 71.8% belonged to serogroup 1. The volume of the storage tank, interrupting circulation for several hours daily and intermittently raising hot water temperatures to >60 degrees C had no influence on Legionella counts. Plumbing systems with copper pipes were more frequently contaminated than those made of synthetic materials or galvanized steel. An inhibitory effect due to copper was not present. Newly constructed systems (district heating systems were colonized by Legionella. Their significantly lower hot water temperature is thought to be the key factor leading to intensified growth of Legionella. Although hot water systems using solar energy to supplement conventional hot water supplies operate at temperatures 3 degrees C lower than conventional systems, this technique does not seem to promote proliferation of the bacterium. Our data show convincingly that the temperature of the hot water is probably the most important or perhaps the only determinant factor for multiplication of Legionella. Water with a temperature below 46 degrees C was

  5. Investigation of Legionella Contamination in Bath Water Samples by Culture, Amoebic Co-Culture, and Real-Time Quantitative PCR Methods

    Directory of Open Access Journals (Sweden)

    Akiko Edagawa

    2015-10-01

    Full Text Available We investigated Legionella contamination in bath water samples, collected from 68 bathing facilities in Japan, by culture, culture with amoebic co-culture, real-time quantitative PCR (qPCR, and real-time qPCR with amoebic co-culture. Using the conventional culture method, Legionella pneumophila was detected in 11 samples (11/68, 16.2%. Contrary to our expectation, the culture method with the amoebic co-culture technique did not increase the detection rate of Legionella (4/68, 5.9%. In contrast, a combination of the amoebic co-culture technique followed by qPCR successfully increased the detection rate (57/68, 83.8% compared with real-time qPCR alone (46/68, 67.6%. Using real-time qPCR after culture with amoebic co-culture, more than 10-fold higher bacterial numbers were observed in 30 samples (30/68, 44.1% compared with the same samples without co-culture. On the other hand, higher bacterial numbers were not observed after propagation by amoebae in 32 samples (32/68, 47.1%. Legionella was not detected in the remaining six samples (6/68, 8.8%, irrespective of the method. These results suggest that application of the amoebic co-culture technique prior to real-time qPCR may be useful for the sensitive detection of Legionella from bath water samples. Furthermore, a combination of amoebic co-culture and real-time qPCR might be useful to detect viable and virulent Legionella because their ability to invade and multiply within free-living amoebae is considered to correlate with their pathogenicity for humans. This is the first report evaluating the efficacy of the amoebic co-culture technique for detecting Legionella in bath water samples.

  6. Caspase-1 but Not Caspase-11 Is Required for NLRC4-Mediated Pyroptosis and Restriction of Infection by Flagellated Legionella Species in Mouse Macrophages and In Vivo.

    Science.gov (United States)

    Cerqueira, Daiane M; Pereira, Marcelo S F; Silva, Alexandre L N; Cunha, Larissa D; Zamboni, Dario S

    2015-09-01

    Gram-negative bacteria from the Legionella genus are intracellular pathogens that cause a severe form of pneumonia called Legionnaires' disease. The bacteria replicate intracellularly in macrophages, and the restriction of bacterial replication by these cells is critical for host resistance. The activation of the NAIP5/NLRC4 inflammasome, which is readily triggered in response to bacterial flagellin, is essential for the restriction of bacterial replication in murine macrophages. Once activated, this inflammasome induces pore formation and pyroptosis and facilitates the restriction of bacterial replication in macrophages. Because investigations related to the NLRC4-mediated restriction of Legionella replication were performed using mice double deficient for caspase-1 and caspase-11, we assessed the participation of caspase-1 and caspase-11 in the functions of the NLRC4 inflammasome and the restriction of Legionella replication in macrophages and in vivo. By using several species of Legionella and mice singly deficient for caspase-1 or caspase-11, we demonstrated that caspase-1 but not caspase-11 was required for pore formation, pyroptosis, and restriction of Legionella replication in macrophages and in vivo. By generating F1 mice in a mixed 129 × C57BL/6 background deficient (129 × Casp-11(-/-) ) or sufficient (129 × C57BL/6) for caspase-11 expression, we found that caspase-11 was dispensable for the restriction of Legionella pneumophila replication in macrophages and in vivo. Thus, although caspase-11 participates in flagellin-independent noncanonical activation of the NLRP3 inflammasome, it is dispensable for the activities of the NLRC4 inflammasome. In contrast, functional caspase-1 is necessary and sufficient to trigger flagellin/NLRC4-mediated restriction of Legionella spp. infection in macrophages and in vivo. Copyright © 2015 by The American Association of Immunologists, Inc.

  7. Investigation of Legionella Contamination in Bath Water Samples by Culture, Amoebic Co-Culture, and Real-Time Quantitative PCR Methods.

    Science.gov (United States)

    Edagawa, Akiko; Kimura, Akio; Kawabuchi-Kurata, Takako; Adachi, Shinichi; Furuhata, Katsunori; Miyamoto, Hiroshi

    2015-10-19

    We investigated Legionella contamination in bath water samples, collected from 68 bathing facilities in Japan, by culture, culture with amoebic co-culture, real-time quantitative PCR (qPCR), and real-time qPCR with amoebic co-culture. Using the conventional culture method, Legionella pneumophila was detected in 11 samples (11/68, 16.2%). Contrary to our expectation, the culture method with the amoebic co-culture technique did not increase the detection rate of Legionella (4/68, 5.9%). In contrast, a combination of the amoebic co-culture technique followed by qPCR successfully increased the detection rate (57/68, 83.8%) compared with real-time qPCR alone (46/68, 67.6%). Using real-time qPCR after culture with amoebic co-culture, more than 10-fold higher bacterial numbers were observed in 30 samples (30/68, 44.1%) compared with the same samples without co-culture. On the other hand, higher bacterial numbers were not observed after propagation by amoebae in 32 samples (32/68, 47.1%). Legionella was not detected in the remaining six samples (6/68, 8.8%), irrespective of the method. These results suggest that application of the amoebic co-culture technique prior to real-time qPCR may be useful for the sensitive detection of Legionella from bath water samples. Furthermore, a combination of amoebic co-culture and real-time qPCR might be useful to detect viable and virulent Legionella because their ability to invade and multiply within free-living amoebae is considered to correlate with their pathogenicity for humans. This is the first report evaluating the efficacy of the amoebic co-culture technique for detecting Legionella in bath water samples.

  8. Legionella spp. isolation and quantification from greywater

    OpenAIRE

    Rodríguez-Martínez, Sara; Blanky, Marina; Friedler, Eran; Halpern, Malka

    2015-01-01

    Legionella, an opportunistic human pathogen whose natural environment is water, is transmitted to humans through inhalation of contaminated aerosols. Legionella has been isolated from a high diversity of water types. Due its importance as a pathogen, two ISO protocols have been developed for its monitoring. However, these two protocols are not suitable for analyzing Legionella in greywater (GW). GW is domestic wastewater excluding the inputs from toilets and kitchen. It can serve as an altern...

  9. Exposure to synthetic greywater inhibits amoebae encystation and alters expression of Legionella pneumophila virulence genes

    Science.gov (United States)

    Water conservation efforts have focused on greywater (GW) usage, especially for applications that do not require potable water quality. However, there is a need to better understand environmental pathogens and their free-living amoebae (FLA) hosts within GW. Using synthetic gre...

  10. Legionella pneumophila transcriptional response following exposure to CuO nanoparticles

    Science.gov (United States)

    Copper ions are an effective antimicrobial agent used to control Legionnaires’ disease and Pontiac fever arising from institutional drinking water systems. Here we present data on an alternative bactericidal agent, CuO nanoparticles (CuO-NPs), and test its efficacy at three conce...

  11. Inter-kingdom Signaling by the Legionella Quorum Sensing Molecule LAI-1 Modulates Cell Migration through an IQGAP1-Cdc42-ARHGEF9-Dependent Pathway.

    Directory of Open Access Journals (Sweden)

    Sylvia Simon

    2015-12-01

    Full Text Available Small molecule signaling promotes the communication between bacteria as well as between bacteria and eukaryotes. The opportunistic pathogenic bacterium Legionella pneumophila employs LAI-1 (3-hydroxypentadecane-4-one for bacterial cell-cell communication. LAI-1 is produced and detected by the Lqs (Legionella quorum sensing system, which regulates a variety of processes including natural competence for DNA uptake and pathogen-host cell interactions. In this study, we analyze the role of LAI-1 in inter-kingdom signaling. L. pneumophila lacking the autoinducer synthase LqsA no longer impeded the migration of infected cells, and the defect was complemented by plasmid-borne lqsA. Synthetic LAI-1 dose-dependently inhibited cell migration, without affecting bacterial uptake or cytotoxicity. The forward migration index but not the velocity of LAI-1-treated cells was reduced, and the cell cytoskeleton appeared destabilized. LAI-1-dependent inhibition of cell migration involved the scaffold protein IQGAP1, the small GTPase Cdc42 as well as the Cdc42-specific guanine nucleotide exchange factor ARHGEF9, but not other modulators of Cdc42, or RhoA, Rac1 or Ran GTPase. Upon treatment with LAI-1, Cdc42 was inactivated and IQGAP1 redistributed to the cell cortex regardless of whether Cdc42 was present or not. Furthermore, LAI-1 reversed the inhibition of cell migration by L. pneumophila, suggesting that the compound and the bacteria antagonistically target host signaling pathway(s. Collectively, the results indicate that the L. pneumophila quorum sensing compound LAI-1 modulates migration of eukaryotic cells through a signaling pathway involving IQGAP1, Cdc42 and ARHGEF9.

  12. Inter-kingdom Signaling by the Legionella Quorum Sensing Molecule LAI-1 Modulates Cell Migration through an IQGAP1-Cdc42-ARHGEF9-Dependent Pathway.

    Science.gov (United States)

    Simon, Sylvia; Schell, Ursula; Heuer, Natalie; Hager, Dominik; Albers, Michael F; Matthias, Jan; Fahrnbauer, Felix; Trauner, Dirk; Eichinger, Ludwig; Hedberg, Christian; Hilbi, Hubert

    2015-12-01

    Small molecule signaling promotes the communication between bacteria as well as between bacteria and eukaryotes. The opportunistic pathogenic bacterium Legionella pneumophila employs LAI-1 (3-hydroxypentadecane-4-one) for bacterial cell-cell communication. LAI-1 is produced and detected by the Lqs (Legionella quorum sensing) system, which regulates a variety of processes including natural competence for DNA uptake and pathogen-host cell interactions. In this study, we analyze the role of LAI-1 in inter-kingdom signaling. L. pneumophila lacking the autoinducer synthase LqsA no longer impeded the migration of infected cells, and the defect was complemented by plasmid-borne lqsA. Synthetic LAI-1 dose-dependently inhibited cell migration, without affecting bacterial uptake or cytotoxicity. The forward migration index but not the velocity of LAI-1-treated cells was reduced, and the cell cytoskeleton appeared destabilized. LAI-1-dependent inhibition of cell migration involved the scaffold protein IQGAP1, the small GTPase Cdc42 as well as the Cdc42-specific guanine nucleotide exchange factor ARHGEF9, but not other modulators of Cdc42, or RhoA, Rac1 or Ran GTPase. Upon treatment with LAI-1, Cdc42 was inactivated and IQGAP1 redistributed to the cell cortex regardless of whether Cdc42 was present or not. Furthermore, LAI-1 reversed the inhibition of cell migration by L. pneumophila, suggesting that the compound and the bacteria antagonistically target host signaling pathway(s). Collectively, the results indicate that the L. pneumophila quorum sensing compound LAI-1 modulates migration of eukaryotic cells through a signaling pathway involving IQGAP1, Cdc42 and ARHGEF9.

  13. Development of a genus-specific next generation sequencing approach for sensitive and quantitative determination of the Legionella microbiome in freshwater systems.

    Science.gov (United States)

    Pereira, Rui P A; Peplies, Jörg; Brettar, Ingrid; Höfle, Manfred G

    2017-03-31

    Next Generation Sequencing (NGS) has revolutionized the analysis of natural and man-made microbial communities by using universal primers for bacteria in a PCR based approach targeting the 16S rRNA gene. In our study we narrowed primer specificity to a single, monophyletic genus because for many questions in microbiology only a specific part of the whole microbiome is of interest. We have chosen the genus Legionella, comprising more than 20 pathogenic species, due to its high relevance for water-based respiratory infections. A new NGS-based approach was designed by sequencing 16S rRNA gene amplicons specific for the genus Legionella using the Illumina MiSeq technology. This approach was validated and applied to a set of representative freshwater samples. Our results revealed that the generated libraries presented a low average raw error rate per base (95%) and very good repeatability. Only in samples in which the gammabacterial clade SAR86 was present more than 1% non-Legionella sequences were observed. Next-generation sequencing read counts did not reveal considerable amplification/sequencing biases and showed a sensitive as well as precise quantification of L. pneumophila along a dilution range using a spiked-in, certified genome standard. The genome standard and a mock community consisting of six different Legionella species demonstrated that the developed NGS approach was quantitative and specific at the level of individual species, including L. pneumophila. The sensitivity of our genus-specific approach was at least one order of magnitude higher compared to the universal NGS approach. Comparison of quantification by real-time PCR showed consistency with the NGS data. Overall, our NGS approach can determine the quantitative abundances of Legionella species, i. e. the complete Legionella microbiome, without the need for species-specific primers. The developed NGS approach provides a new molecular surveillance tool to monitor all Legionella species in qualitative

  14. [Evaluation of a Legionella outbreak emerged in a recently opening hotel].

    Science.gov (United States)

    Erdoğan, Haluk; Arslan, Hande

    2013-04-01

    samples could not be obtained from the cases, the diagnosis were not confirmed by culture but by urinary antigen test. Besides high antibody titer in single serum sample was accepted as a diagnostic marker. Additionally 26 cases who accommodated in the same hotel and presented with high fever without pneumonia were treated in the outpatient clinics of our hospital. Urinary antigen test was performed in 11 of those patients to confirm the prediagnosis of pontiac fever, however all were found negative. Likewise convalescent phase sera for the confirmation of the diagnosis by seroconversion could not be obtained since they all were foreign tourists. Investigation of water sources of the hotel revealed that the municipal drinking water network had not been connected yet and the hotel supplied water from groundwater sources. The analysis of multiple samples from multiple sites of hotel's water system indicated that the water temperature was between 35-45°C and the iron level was beyond the acceptable limits (245 µg/L) recommended for drinking water in the regulation guides. These properties were considered as the factors that enhanced the growth and survival of Legionella species. Water samples were cultivated on BCYE-_ (Buffered Charcoal Yeast Extract a-Ketoglutarate) and GVPC (Glycine-Vancomycin-Polymyxin-Cycloheximide) agar plates and 11 out of a total 13 samples yielded Legionella spp. growth. All isolates were identified as L.pneumophila serogroup 1 by specific antisera. Legionella decontamination of hotel's water system was managed by implementation of hyperchlorination method as well as superheating (> 60°C) of water. The hotel was not closed during the outbreak and cultures of water samples obtained for one year later did not yield any Legionella spp. growth. This outbreak emphasized that hotel residents are at risk for acquiring LD in the presence of a colonized water system, even in a newly constructed building. In conclusion, effective control and decontamination

  15. Extracorporele life support bij Legionella-pneumonie

    NARCIS (Netherlands)

    Koekkoek, W A C; Savelkoul, C; Wijnandts, P R; Platenkamp, M; Donker, D W|info:eu-repo/dai/nl/413646386; Tjan, D H T

    2016-01-01

    BACKGROUND: Legionella species cause 5% of all community acquired pneumonias. However, Legionella pneumonia results relatively often in admission to the intensive care unit (ICU). A significant complication is the development of acute respiratory distress syndrome (ARDS). The ICU mortality rate for

  16. Legionella spp. isolation and quantification from greywater.

    Science.gov (United States)

    Rodríguez-Martínez, Sara; Blanky, Marina; Friedler, Eran; Halpern, Malka

    2015-01-01

    Legionella, an opportunistic human pathogen whose natural environment is water, is transmitted to humans through inhalation of contaminated aerosols. Legionella has been isolated from a high diversity of water types. Due its importance as a pathogen, two ISO protocols have been developed for its monitoring. However, these two protocols are not suitable for analyzing Legionella in greywater (GW). GW is domestic wastewater excluding the inputs from toilets and kitchen. It can serve as an alternative water source, mainly for toilet flushing and garden irrigation; both producing aerosols that can cause a risk for Legionella infection. Hence, before reuse, GW has to be treated and its quality needs to be monitored. The difficulty of Legionella isolation from GW strives in the very high load of contaminant bacteria. Here we describe a modification of the ISO protocol 11731:1998 that enables the isolation and quantification of Legionella from GW samples. The following modifications were made:•To enable isolation of Legionella from greywater, a pre-filtration step that removes coarse matter is recommended.•Legionella can be isolated after a combined acid-thermic treatment that eliminates the high load of contaminant bacteria in the sample.

  17. Confirmed and Potential Sources of Legionella Reviewed

    NARCIS (Netherlands)

    van Heijnsbergen, Eri; Schalk, Johanna A C; Euser, Sjoerd M; Brandsema, Petra S; den Boer, Jeroen W; de Roda Husman, Ana Maria|info:eu-repo/dai/nl/139498281

    2015-01-01

    Legionella bacteria are ubiquitous in natural matrices and man-made systems. However, it is not always clear if these reservoirs can act as source of infection resulting in cases of Legionnaires' disease. This review provides an overview of reservoirs of Legionella reported in the literature, other

  18. Combined pericarditis and pneumonia caused by Legionella infection

    DEFF Research Database (Denmark)

    Svendsen, J H; Jønsson, V; Niebuhr, U

    1987-01-01

    During a one year period acute pericarditis was diagnosed in 16 consecutive patients without acute infarction or malignancy. In two of these patients with both pericarditis and pneumonia Legionella infection was present. One case was caused by Legionella longbeachae and the other by both Legionella...... longbeachae and Legionella jordanis. When pericarditis is associated with pneumonia Legionella infection should be sought so that effective treatment with erythromycin may be started early....

  19. Disinfection of water distribution systems for Legionella.

    Science.gov (United States)

    Lin, Y S; Stout, J E; Yu, V L; Vidic, R D

    1998-06-01

    Hospital-acquired legionnaires' disease arises from the presence of Legionella in hospital water systems. Legionella not only persists in hot water tanks but is also found in the biofilm throughout the entire water distribution system. Conditions within water systems that promote Legionella colonization include water temperature, configuration and age of the hot water tank, physicochemical constituents of the water, plumbing materials, and commensal microflora. Hospital-acquired legionnaires' disease has been prevented by instituting control measures directed at the water distribution system. These include superheat-and-flush, copper/silver ionization, ultraviolet light, instantaneous heating systems, and hyperchlorination. Each of the above disinfection methods has been proven to be effective in the short-term, but long-term efficacy has been difficult due to limitations associated with each method. The complexities of Legionella disinfection, including advantages and disadvantages of each method, are reviewed. A successful Legionella prevention program requires cooperation and communication among hospital administrative personnel, engineers, and infection control staff. Routine environmental surveillance cultures for Legionella are the critical component for successful long-term disinfection. Culture results document the efficacy of the disinfection method and alert the hospital staff to consider Legionella in hospitalized patients with pneumonia.

  20. Functional Analysis of the Alternative Sigma-28 Factor FliA and Its Anti-Sigma Factor FlgM of the Nonflagellated Legionella Species L. oakridgensis.

    Science.gov (United States)

    Tlapák, Hana; Rydzewski, Kerstin; Schulz, Tino; Weschka, Dennis; Schunder, Eva; Heuner, Klaus

    2017-06-01

    Legionella oakridgensis causes Legionnaires' disease but is known to be less virulent than Legionella pneumophilaL. oakridgensis is one of the Legionella species that is nonflagellated. The genes of the flagellar regulon are absent, except those encoding the alternative sigma-28 factor (FliA) and its anti-sigma-28 factor (FlgM). Similar to L. oakridgensis, Legionella adelaidensis and Legionella londiniensis, located in the same phylogenetic clade, have no flagellar regulon, although both are positive for fliA and flgM Here, we investigated the role and function of both genes to better understand the role of FliA, the positive regulator of flagellin expression, in nonflagellated strains. We demonstrated that the FliA gene of L. oakridgensis encodes a functional sigma-28 factor that enables the transcription start from the sigma-28-dependent promoter site. The investigations have shown that FliA is necessary for full fitness of L. oakridgensis Interestingly, expression of FliA-dependent genes depends on the growth phase and temperature, as already shown for L. pneumophila strains that are flagellated. In addition, we demonstrated that FlgM is a negative regulator of FliA-dependent gene expression. FlgM seems to be degraded in a growth-phase- and temperature-dependent manner, instead of being exported into the medium as reported for most bacteria. The degradation of FlgM leads to an increase of FliA activity.IMPORTANCE A less virulent Legionella species, L. oakridgensis, causes Legionnaires' disease and is known to not have flagella, even though L. oakridgensis has the regulator of flagellin expression (FliA). This protein has been shown to be involved in the expression of virulence factors. Thus, the strain was chosen for use in this investigation to search for FliA target genes and to identify putative virulence factors of L. oakridgensis One of the five major target genes of FliA identified here encodes the anti-FliA sigma factor FlgM. Interestingly, in contrast to

  1. Legionella spp. and legionellosis in southeastern Italy: disease epidemiology and environmental surveillance in community and health care facilities

    Science.gov (United States)

    2010-01-01

    Background Following the publication of the Italian Guidelines for the control and prevention of legionellosis an environmental and clinical surveillance has been carried out in Southeastern Italy. The aim of the study is to identify the risk factors for the disease, so allowing better programming of the necessary prevention measures. Methods During the period January 2000 - December 2009 the environmental surveillance was carried out by water sampling of 129 health care facilities (73 public and 56 private hospitals) and 533 buildings within the community (63 private apartments, 305 hotels, 19 offices, 4 churches, 116 gyms, 3 swimming pools and 23 schools). Water sampling and microbiological analysis were carried out following the Italian Guidelines. From January 2005, all facilities were subject to risk analysis through the use of a standardized report; the results were classified as good (G), medium (M) and bad (B). As well, all the clinical surveillance forms for legionellosis, which must be compiled by physicians and sent to the Regional Centre for Epidemiology (OER), were analyzed. Results Legionella spp. was found in 102 (79.1%) health care facilities and in 238 (44.7%) community buildings. The percentages for the contamination levels 10,000 cfu/L were respectively 33.1%, 53.4% and 13.5% for samples from health care facilities and 33.5%, 43.3% and 23.2% for samples from the community. Both in hospital and community environments, Legionella pneumophila serogroup (L. pn sg) 2-14 was the most frequently isolate (respectively 54.8% and 40.8% of positive samples), followed by L. pn sg 1 (respectively 31.3% and 33%). The study showed a significant association between M or B score at the risk analysis and Legionella spp. positive microbiological test results (p Legionella spp. Furthermore, the laboratory diagnosis of legionellosis cannot be excluded only on the basis of a single negative test: some patients were positive to only one of the diagnostic tests. PMID

  2. Legionella (Legionnaires' Disease and Pontiac Fever): Prevention

    Science.gov (United States)

    ... Program for Laboratories For Media Water System Maintenance Water Management Toolkit Identify Buildings at Increased Risk Considerations When Working with Legionella Consultants Surveillance & Reporting Resources Materials Guidelines, Standards, and Laws Publications Related Links ...

  3. Legionella (Legionnaires' Disease and Pontiac Fever): Diagnosis

    Science.gov (United States)

    ... Program for Laboratories For Media Water System Maintenance Water Management Toolkit Identify Buildings at Increased Risk Considerations When Working with Legionella Consultants Surveillance & Reporting Resources Materials Guidelines, Standards, and Laws Publications Related Links ...

  4. Confirmed and Potential Sources of Legionella Reviewed.

    Science.gov (United States)

    van Heijnsbergen, Eri; Schalk, Johanna A C; Euser, Sjoerd M; Brandsema, Petra S; den Boer, Jeroen W; de Roda Husman, Ana Maria

    2015-04-21

    Legionella bacteria are ubiquitous in natural matrices and man-made systems. However, it is not always clear if these reservoirs can act as source of infection resulting in cases of Legionnaires' disease. This review provides an overview of reservoirs of Legionella reported in the literature, other than drinking water distribution systems. Levels of evidence were developed to discriminate between potential and confirmed sources of Legionella. A total of 17 systems and matrices could be classified as confirmed sources of Legionella. Many other man-made systems or natural matrices were not classified as a confirmed source, since either no patients were linked to these reservoirs or the supporting evidence was weak. However, these systems or matrices could play an important role in the transmission of infectious Legionella bacteria; they might not yet be considered in source investigations, resulting in an underestimation of their importance. To optimize source investigations it is important to have knowledge about all the (potential) sources of Legionella. Further research is needed to unravel what the contribution is of each confirmed source, and possibly also potential sources, to the LD disease burden.

  5. Legionella on board trains: effectiveness of environmental surveillance and decontamination

    National Research Council Canada - National Science Library

    Quaranta, Gianluigi; Vincenti, Sara; Ferriero, Anna Maria; Boninti, Federica; Sezzatini, Romina; Turnaturi, Cinzia; Gliubizzi, Maria Daniela; Munafò, Elio; Ceccarelli, Gianluca; Causarano, Carmelo; Accorsi, Massimo; Del Nord, Pasquale; Ricciardi, Walter; Laurenti, Patrizia

    2012-01-01

    .... Many studies describe the frequency and severity of Legionella spp. contamination in spa pools, natural pools, hotels and ships, but there is no study analysing the environmental monitoring of Legionella on board trains...

  6. Combined pericarditis and pneumonia caused by Legionella infection

    DEFF Research Database (Denmark)

    Svendsen, Jesper Hastrup; Jønsson, V; Niebuhr, U

    1987-01-01

    During a one year period acute pericarditis was diagnosed in 16 consecutive patients without acute infarction or malignancy. In two of these patients with both pericarditis and pneumonia Legionella infection was present. One case was caused by Legionella longbeachae and the other by both Legionel...... longbeachae and Legionella jordanis. When pericarditis is associated with pneumonia Legionella infection should be sought so that effective treatment with erythromycin may be started early....

  7. [The clinical value of urinary antigen detection of Legionella pneumonia].

    Science.gov (United States)

    Jiang, Luxi; Chen, Yu; Xia, Shuyue; Ma, Jiangwei; Zhao, Hongwen; Lu, Ye; Tao, Sixu; Zhao, Li

    2015-01-01

    To investigate the clinical value of urinary antigen detection of Legionella, and to describe the clinical characteristics of Legionella pneumonia. Patients with suspected Legionella pneumonia were enrolled from the Respiratory departments of 3 tertiary hospitals in Shenyang during May 2011 to November 2013. Urinary Legionella antigen was detected for all the enrolled patients. Bacterial culture, polymerase chain reaction (PCR) for Legionella, and double Legionella antibody detection in sera were performed for each patient whose urinary antigen was positive. Patients confirmed to have Legionella pneumonia were pooled and analyzed. Totally 13 cases presenting with pneumonia were positive for Legionella by the urinary antigen method, and in one of them Legionella strain was isolated from the secretion of lower respiratory tract. PCR detection was performed in 8 patients, and 4 of them were positive. Legionella antibody detection was performed in 12 patients, and 7 of them were positive. Nine patients had a history of exposure to Legionella high-risk environments. The characteristics of the cases with Legionella pneumonia were as follows: characteristic orange sputum in 4 patients, digestive symptoms in 6, neurologic disorders in 8, hyponatremia in 10, hypoxia with oxygenation index 130) in 8 patients . Chest CT scan showed bilateral involvement in 6, ground-glass opacity combined with consolidation in 11, and moderate pleural effusion in 11 patients. Cavity and reversed halo sign were found in one case, respectively. All of the patients received fluoroquinolone treatment, and 11 patients recovered completely while 2 died of multiple organ dysfunction syndrome, one of them was complicated with secondary infection. Detection of urinary antigen of Legionella is very useful in the diagnosis of Legionella pneumonia. Attention should be paid to exposure history to the high-risk environments and multiple organ impairment when Legionella infection is suspected. Orange sputum

  8. Characterization of Legionella Species from Watersheds in British Columbia, Canada

    Science.gov (United States)

    Peabody, Michael A.; Caravas, Jason A.; Morrison, Shatavia S.; Mercante, Jeffrey W.; Prystajecky, Natalie A.; Raphael, Brian H.

    2017-01-01

    ABSTRACT Legionella spp. present in some human-made water systems can cause Legionnaires’ disease in susceptible individuals. Although legionellae have been isolated from the natural environment, variations in the organism’s abundance over time and its relationship to aquatic microbiota are poorly understood. Here, we investigated the presence and diversity of legionellae through 16S rRNA gene amplicon and metagenomic sequencing of DNA from isolates collected from seven sites in three watersheds with varied land uses over a period of 1 year. Legionella spp. were found in all watersheds and sampling sites, comprising up to 2.1% of the bacterial community composition. The relative abundance of Legionella tended to be higher in pristine sites than in sites affected by agricultural activity. The relative abundance levels of Amoebozoa, some of which are natural hosts of legionellae, were similarly higher in pristine sites. Compared to other bacterial genera detected, Legionella had both the highest richness and highest alpha diversity. Our findings indicate that a highly diverse population of legionellae may be found in a variety of natural aquatic sources. Further characterization of these diverse natural populations of Legionella will help inform prevention and control efforts aimed at reducing the risk of Legionella colonization of built environments, which could ultimately decrease the risk of human disease. IMPORTANCE Many species of Legionella can cause Legionnaires’ disease, a significant cause of bacterial pneumonia. Legionella in human-made water systems such as cooling towers and building plumbing systems are the primary sources of Legionnaires’ disease outbreaks. In this temporal study of natural aquatic environments, Legionella relative abundance was shown to vary in watersheds associated with different land uses. Analysis of the Legionella sequences detected at these sites revealed highly diverse populations that included potentially novel

  9. Multifaceted effects of synthetic TLR2 ligand and Legionella pneumophilia on Treg-mediated suppression of T cell activation

    Directory of Open Access Journals (Sweden)

    Sutmuller Roger PM

    2011-03-01

    Full Text Available Abstract Background Regulatory T cells (Treg play a crucial role in maintaining immune homeostasis and self-tolerance. The immune suppressive effects of Tregs should however be limited in case effective immunity is required against pathogens or cancer cells. We previously found that the Toll-like receptor 2 (TLR2 agonist, Pam3CysSK4, directly stimulated Tregs to expand and temporarily abrogate their suppressive capabilities. In this study, we evaluate the effect of Pam3CysSK4 and Legionella pneumophila, a natural TLR2 containing infectious agent, on effector T (Teff cells and dendritic cells (DCs individually and in co-cultures with Tregs. Results TLR2 agonists can directly provide a co-stimulatory signal inducing enhanced proliferation and cytokine production of naive CD4+ Teff cells. With respect to cytokine production, DCs appear to be most sensitive to low amounts of TLR agonists. Using wild type and TLR2-deficient cells in Treg suppression assays, we accordingly show that all cells (e.g. Treg, Teff cells and DCs contributed to overcome Treg-mediated suppression of Teff cell proliferation. Furthermore, while TLR2-stimulated Tregs readily lost their ability to suppress Teff cell proliferation, cytokine production by Teff cells was still suppressed. Similar results were obtained upon stimulation with TLR2 ligand containing bacteria, Legionella pneumophila. Conclusions These findings indicate that both synthetic and natural TLR2 agonists affect DCs, Teff cells and Treg directly, resulting in multi-modal modulation of Treg-mediated suppression of Teff cells. Moreover, Treg-mediated suppression of Teff cell proliferation is functionally distinct from suppression of cytokine secretion.

  10. Molecular survey of occurrence and quantity of Legionella spp., Mycobacterium spp., Pseudomonas aeruginosa and amoeba hosts in municipal drinking water storage tank sediments.

    Science.gov (United States)

    Lu, J; Struewing, I; Yelton, S; Ashbolt, N

    2015-07-01

    To examine the occurrence and quantity of potential pathogens and an indicator of microbial contamination in the sediments of municipal drinking water storage tanks (MDWSTs), given the absence of such data across the United States. Sediment samples (87 MDWST) from eighteen locations across ten states of the United States were collected and assayed by qPCR for a range of potential enteric and opportunistic microbial pathogens and a sewage-associated Bacteroides marker. Potential opportunistic pathogens dominated, with the highest detection of occurrence (per cent positive detection; average cell equivalence (CE)) being Mycobacterium spp. (88·9%; 6·7 ± 8·5 × 10(4) CE g(-1) ), followed by Legionella spp. (66·7%; 5·2 ± 5·9 × 10(3) CE g(-1) ), Pseudomonas aeruginosa (22·2%; 250 ± 880 CE g(-1) ) and Acanthamoeba spp. (38·9%; 53 ± 70 CE g(-1) ), with no detected Naegleria fowleri. Most enteric pathogens (Campylobacter jejuni, Escherichia coli 0157:H7, Salmonella enterica, Cryptosporidium parvum and Giardia duodenalis) were not detected, except for a trace signal for Campylobacter spp. There was significant correlation between the qPCR signals of Legionella spp. and Acanthamoeba spp. (R(2) = 0·61, n = 87, P = 0·0001). Diverse Legionella spp. including Leg. pneumophila, Leg. pneumophila sg1 and Leg. anisa were identified, each of which might cause legionellosis. These results imply that potential opportunistic pathogens are common within MDWST sediments and could act as a source of microbial contamination, but need downstream growth to be of potential concern. The results imply that opportunistic pathogen risks may need to be managed by regular tank cleaning or other management practices. 2015 The Society for Applied Microbiology.

  11. Epidemiology of severe pneumonia caused by Legionella longbeachae, Mycoplasma pneumoniae, and Chlamydia pneumoniae: 1-year, population-based surveillance for severe pneumonia in Thailand.

    Science.gov (United States)

    Phares, Christina R; Wangroongsarb, Piyada; Chantra, Somrak; Paveenkitiporn, Wantana; Tondella, Maria-Lucia; Benson, Robert F; Thacker, W Lanier; Fields, Barry S; Moore, Matthew R; Fischer, Julie; Dowell, Scott F; Olsen, Sonja J

    2007-12-15

    Legionella species, Mycoplasma pneumoniae, and Chlamydia pneumoniae are recognized as important causes of pneumonia in high-income countries, but their significance in middle-income countries, such as Thailand, is unknown. Population-based surveillance identified inpatient 3489 cases of clinically-defined pneumonia in a rural Thai province for 1 year. Patients who had a chest radiograph performed (for 2059 cases of pneumonia) were enrolled in an etiology study (which included 755 cases of pneumonia among 738 patients). Paired serum, nasopharyngeal swab, and urine specimens were obtained for diagnostic immunologic and molecular tests. Patients aged pneumonia due to Legionella longbeachae requiring hospitalization was 5-29 cases per 100,000 population. No case of Legionella pneumophila pneumonia was observed. The definite C. pneumoniae pneumonia incidence was 3-23 cases per 100,000 population; rates were highest among patients aged or=70 years (23-201 cases per 100,000 population). M. pneumoniae pneumonia had a similar age distribution, with an overall incidence of 6-44 cases per 100,000 population. These pathogens were associated with 15% of all cases of pneumonia. A nonsignificantly higher proportion of patients with pneumonia associated with L. longbeachae, compared with patients with pneumonia associated with M. pneumoniae or C. pneumoniae, required supplemental oxygen or mechanical ventilation (45% vs. 18%; Ppneumonia, only 15% received antibiotics with activity against the associated pathogen. M. pneumoniae, C. pneumoniae, and L. longbeachae, but not L. pneumophila, are frequently associated with severe pneumonia in rural Thailand. Few patients receive antibiotics that cover atypical pathogens.

  12. [i]Legionella spp[/i]., amoebae and not-fermenting Gram negative bacteria in an Italian university hospital water system

    Directory of Open Access Journals (Sweden)

    Pasqualina Laganà

    2014-09-01

    Full Text Available [b]Introduction. [/b]In hospital and other health care facilities, contamination of water systems by potentially infectious microorganisms, such as bacteria, viruses and protozoa, is a source of nosocomial infections, which may originate fromcolonization of water pipes, cooling towers, spa pools, taps, showers and water supplies. [b]Objective. [/b]The study focuses on the occurrence of [i]Legionella spp.[/i], free-living amoebae and non-fermenting Gram-negative microorganisms in a University hospital water system located in the town of Messina (Sicily, Italy, which had never been examined previously. Materials and Methods. From January 2008 – March 2009, hot tap water samples were collected from 10 wards.[i] Legionella spp[/i]. recovered on selective culture medium were identified by microagglutination latex test; free-living amoebae were cultured using [i]Escherichia coli [/i]as a food source. Non-fermenting Gram negative microorganisms were identified by API 20 NE strips. [b]Results.[/b] [i]Legionella spp.[/i] were found in 33.33% of the samples. [i]L. pneumophila[/i] serogroup 1 was recovered from the Laboratory Diagnostic and Anaesthesia-Neurology Wards, with a peak of 3.5 × 10[sup]4[/sup] cfu/L in May 2008. [i]L. pneumophila[/i] serogroups 2–14 were found in the Othorhinolaryngology, Pathologic Anatomy, Paediatrics and Surgery Wards, and peaked (4 × 10[sup]4[/sup] cfu/L in April 2008. Pseudomonadaceae and Hyphomycetes were also detected. Legionella spp. were recovered from samples positive for non-pathogenic amoebae [i]Hartmannella spp[/i]. [b]Conclusion.[/b] This first study of a Messina hospital water system suggested potential health risks related to the detection of [i]Hartmannella spp[/i]., as reservoirs for[i] Legionella spp.[/i], and Pseudomonas aeruginosa, a Gram negative non-fermenting bacterium frequently causing nosocomial pneumonia. The urgent need for monitoring programmes and prevention measures to ensure hospital water

  13. Structural basis for the recruitment and activation of the Legionella phospholipase VipD by the host GTPase Rab5

    Science.gov (United States)

    Lucas, María; Gaspar, Andrew H.; Pallara, Chiara; Rojas, Adriana Lucely; Fernández-Recio, Juan; Machner, Matthias P.; Hierro, Aitor

    2014-01-01

    A challenge for microbial pathogens is to assure that their translocated effector proteins target only the correct host cell compartment during infection. The Legionella pneumophila effector vacuolar protein sorting inhibitor protein D (VipD) localizes to early endosomal membranes and alters their lipid and protein composition, thereby protecting the pathogen from endosomal fusion. This process requires the phospholipase A1 (PLA1) activity of VipD that is triggered specifically on VipD binding to the host cell GTPase Rab5, a key regulator of endosomes. Here, we present the crystal structure of VipD in complex with constitutively active Rab5 and reveal the molecular mechanism underlying PLA1 activation. An active site-obstructing loop that originates from the C-terminal domain of VipD is repositioned on Rab5 binding, thereby exposing the catalytic pocket within the N-terminal PLA1 domain. Substitution of amino acid residues located within the VipD–Rab5 interface prevented Rab5 binding and PLA1 activation and caused a failure of VipD mutant proteins to target to Rab5-enriched endosomal structures within cells. Experimental and computational analyses confirmed an extended VipD-binding interface on Rab5, explaining why this L. pneumophila effector can compete with cellular ligands for Rab5 binding. Together, our data explain how the catalytic activity of a microbial effector can be precisely linked to its subcellular localization. PMID:25114243

  14. Relationships between free-living protozoa, cultivable Legionella spp., and water quality characteristics in three drinking water supplies in the Caribbean.

    Science.gov (United States)

    Valster, Rinske M; Wullings, Bart A; van den Berg, Riemsdijk; van der Kooij, Dick

    2011-10-01

    The study whose results are presented here aimed at identifying free-living protozoa (FLP) and conditions favoring the growth of these organisms and cultivable Legionella spp. in drinking water supplies in a tropical region. Treated and distributed water (±30°C) of the water supplies of three Caribbean islands were sampled and investigated with molecular techniques, based on the 18S rRNA gene. The protozoan host Hartmannella vermiformis and cultivable Legionella pneumophila were observed in all three supplies. Operational taxonomic units (OTUs) with the highest similarity to the potential or candidate hosts Acanthamoeba spp., Echinamoeba exundans, E. thermarum, and an Neoparamoeba sp. were detected as well. In total, 59 OTUs of FLP were identified. The estimated protozoan richness did not differ significantly between the three supplies. In supply CA-1, the concentration of H. vermiformis correlated with the concentration of Legionella spp. and clones related to Amoebozoa predominated (82%) in the protozoan community. These observations, the low turbidity (water. The absence of H. vermiformis in most samples from supply CA-3 suggests that growth of this protozoan is limited at ATP concentrations of <1 ng liter(-1).

  15. Relationships between Free-Living Protozoa, Cultivable Legionella spp., and Water Quality Characteristics in Three Drinking Water Supplies in the Caribbean▿†

    Science.gov (United States)

    Valster, Rinske M.; Wullings, Bart A.; van den Berg, Riemsdijk; van der Kooij, Dick

    2011-01-01

    The study whose results are presented here aimed at identifying free-living protozoa (FLP) and conditions favoring the growth of these organisms and cultivable Legionella spp. in drinking water supplies in a tropical region. Treated and distributed water (±30°C) of the water supplies of three Caribbean islands were sampled and investigated with molecular techniques, based on the 18S rRNA gene. The protozoan host Hartmannella vermiformis and cultivable Legionella pneumophila were observed in all three supplies. Operational taxonomic units (OTUs) with the highest similarity to the potential or candidate hosts Acanthamoeba spp., Echinamoeba exundans, E. thermarum, and an Neoparamoeba sp. were detected as well. In total, 59 OTUs of FLP were identified. The estimated protozoan richness did not differ significantly between the three supplies. In supply CA-1, the concentration of H. vermiformis correlated with the concentration of Legionella spp. and clones related to Amoebozoa predominated (82%) in the protozoan community. These observations, the low turbidity (water. The absence of H. vermiformis in most samples from supply CA-3 suggests that growth of this protozoan is limited at ATP concentrations of <1 ng liter−1. PMID:21873489

  16. Risk assessment and monitoring of Legionella by culture and q-PCR in a newly built block of flats associated with a small outbreak of legionnaires´ disease

    DEFF Research Database (Denmark)

    Krøjgaard, Louise Hjelmar; Krogfelt, Karen A.; Albrechtsen, Hans-Jørgen

    2010-01-01

    ) and permanent changes of the flow and temperature were conducted to overcome the high concentrations of Legionella in the water. Water samples (104 in total, both cold and warm water) from taps and shower hoses were collected and cultivated. The samples consisted of: A samples: the first one litre, B samples......: one litre collected after flushing until constant temperature and water from shower hoses at a temperature of 38ºC. The survey pointed at important risk factors 1) low temperature of the hot water in some of the most distant taps 2) low flow of the water circulating system because of too small pipe......-100. The samples will be investigated by an in-house q-PCR (Quantitative Real Time PCR). The q-PCR is a Taq-Man based assay with 5S primers detecting Legionella spp and mip primers detecting Legionella pneumophila. The two Legionella q-PCR assays have been validated according to the Afnor standard (T...

  17. Aerobiology of the built environment: Synergy between Legionella and fungi.

    Science.gov (United States)

    Alum, Absar; Isaacs, Galahad Zachariah

    2016-09-02

    The modern built environment (BE) design creates unique ecological niches ideal for the survival and mutual interaction of microbial communities. This investigation focused on the synergistic relations between Legionella and the fungal species commonly found in BEs and the impact of these synergistic relationships on the survival and transmission of Legionella. A field study was conducted to identify the types and concentrations of fungi in BEs. The fungal isolates purified from BEs were cocultured with Legionella to study their synergistic association. Cocultured Legionella cells were aerosolized in an air-tight chamber to evaluate the efficacy of ultraviolet (UV) to inactivate these cells. Aspergillus, Alternaria, and Cladosporium were the most common fungi detected in samples that tested positive for Legionella. After coculturing, Legionella cells were detected inside fungal hyphae. The microscopic observations of Legionella internalization in fungal hyphae were confirmed by molecular analyses. UV disinfection of the aerosolized Legionella cells that were cocultured with fungi indicated that fungal spores and propagules act as a shield against UV radiation. The shield effect of fungal spores on Legionella cells was quantified at >2.5 log10. This study provides the first evidence, to our knowledge, of Legionella cell presence inside fungi detected in an indoor environment. This symbiotic relationship with fungi results in longer survival of Legionella under ambient conditions and provides protection against UV rays. Copyright © 2016. Published by Elsevier Inc.

  18. Structural and Functional Studies of Fatty Acyl Adenylate Ligases from E. coli and L. pneumophila

    Energy Technology Data Exchange (ETDEWEB)

    Z Zhang; R Zhou; J Sauder; P Tonge; S Burley; S Swaminathan

    2011-12-31

    Fatty acyl-AMP ligase (FAAL) is a new member of a family of adenylate-forming enzymes that were recently discovered in Mycobacterium tuberculosis. They are similar in sequence to fatty acyl-coenzyme A (CoA) ligases (FACLs). However, while FACLs perform a two-step catalytic reaction, AMP ligation followed by CoA ligation using ATP and CoA as cofactors, FAALs produce only the acyl adenylate and are unable to perform the second step. We report X-ray crystal structures of full-length FAAL from Escherichia coli (EcFAAL) and FAAL from Legionella pneumophila (LpFAAL) bound to acyl adenylate, determined at resolution limits of 3.0 and 1.85 {angstrom}, respectively. The structures share a larger N-terminal domain and a smaller C-terminal domain, which together resemble the previously determined structures of FAAL and FACL proteins. Our two structures occur in quite different conformations. EcFAAL adopts the adenylate-forming conformation typical of FACLs, whereas LpFAAL exhibits a unique intermediate conformation. Both EcFAAL and LpFAAL have insertion motifs that distinguish them from the FACLs. Structures of EcFAAL and LpFAAL reveal detailed interactions between this insertion motif and the interdomain hinge region and with the C-terminal domain. We suggest that the insertion motifs support sufficient interdomain motions to allow substrate binding and product release during acyl adenylate formation, but they preclude CoA binding, thereby preventing CoA ligation.

  19. Occurrence of Legionella in technological water and studies of the total number of bacteria and fungi in indoor air at workplaces where water aerosol is generated

    Directory of Open Access Journals (Sweden)

    Bożena Krogulska

    2014-06-01

    Full Text Available Background: The aim of the study was to confirm the fact that technological water is a significant carrier of Legionella, a potential serious threat to the health of operators of mechanical devices generating contaminated water aerosol. Material and Methods: Microbiological analyses of water and indoor air were conducted in 8 different production facilities involved in mechanical processing of glass. The study covered 81 samples of water collected from technical water systems. Isolation of Legionella and the determination of total number of microorganisms were processed according to PN-EN ISO 11731-2:2008E and PN-EN ISO 6222:2004P, respectively. Air samples were collected using air samplers and total numbers of bacteria and fungi were determined. Results: The studies of process water, indicated the presence of Legionella in 27.2% of collected samples. These bacteria were present in both closed and open process water reservoirs at 10 cfu/100 ml to 2.9×104 cfu/100 ml. The count of other associated bacteria exceeded 103 cfu/ml. All strains isolated from Legionella-positive samples were identified as L. pneumophila SG 2-14. In 5 of 8 studied production facilities an increased total number of aerial bacteria and fungi was observed in samples collected in close vicinity of aerosol source. Conclusions: To reduce the number of microorganisms in water it is required to introduce technological water quality monitoring and procedures for the cleaning and disinfecting of mechanical devices generating water aerosol. Med Pr 2014;65(3:325–334

  20. Legionella spp. in dental unit waterlines.

    Science.gov (United States)

    Sedlata Juraskova, E; Sedlackova, H; Janska, J; Holy, O; Lalova, I; Matouskova, I

    2017-01-01

    To determine the current presence of Legionella spp. in the output water of dental unit waterlines (DUWLs) and examine its mitigation by disinfection at the Institute of Dentistry and Oral Sciences, Faculty of Medicine and Dentistry, Palacky University Olomouc and University Hospital Olomouc. The first stage of our survey involved collecting samples of DUWL output water from 50 dental chair units (DCUs), and 2 samples of the incoming potable water. In October 2015, a one-time disinfection (1 % Stabimed) of DUWLs was conducted. This was followed by collecting 10 control samples (survey stage 2). From the total of 50 samples (survey stage 1), 18 samples (36.0 %) tested positive for Legionella spp. Following the disinfection, nine of the ten samples no longer showed any presence of Legionella. Based on culture results, the one-time disinfection (1 % Stabimed) was effective. We are unable to comment on the duration of positive effect of disinfection on the occurrence of Legionella spp. in the outlet water. It was a one-time survey (Tab. 2, Ref. 32).

  1. Kunnen luchtwassers legionella verspreiden naar de omgeving?

    NARCIS (Netherlands)

    Bartels, A.A.; Schalk, J.A.C.; Melse, R.W.

    2013-01-01

    Industrieën en veehouderijen gebruiken luchtwassers om ongewenste chemische of organische stoffen, gassen of geuren te verwijderen uit lucht of gas. Onder bepaalde condities (temperatuur en zuurgraad), kan legionella in bepaalde typen natte luchtwassers uitgroeien. Als waternevel het systeem kan

  2. Counting Legionella cells within single amoeba host cells

    Science.gov (United States)

    Here we present the first attempt to quantify L. pneumophila cell numbers within individual amoebae hosts that may be released into engineered water systems. The maximum numbers of culturable L. pneumophila cells grown within Acanthamoeba polyphaga and Naegleria fowleri were 134...

  3. High Prevalence and Genetic Polymorphisms of Legionella in Natural and Man-Made Aquatic Environments in Wenzhou, China.

    Science.gov (United States)

    Zhang, Leyi; Li, Yi; Wang, Xin; Shangguan, Zhihui; Zhou, Haijian; Wu, Yuejin; Wang, Lianghuai; Ren, Hongyu; Hu, Yun; Lin, Meifen; Qin, Tian

    2017-02-24

    Natural and engineered water systems are the main sources of Legionnaires' disease. It is essential from a public health perspective to survey water environments for the existence of Legionella. To analyze the main serogroups, genotypes and pathogenicity of the pathogen, a stratified sampling method was adopted to collect water samples randomly from shower water, cooling tower water, and local public hot springs in Wenzhou, China. Suspected strains were isolated from concentrated water samples. Serum agglutination assay and real-time PCR (Polymerase chain reaction) were used to identify L. pneumophila. Sequence-based typing (SBT) and pulsed-field gel electrophoresis (PFGE) were used to elucidate the genetic polymorphisms in the collected isolates. The intracellular growth ability of the isolates was determined through their interaction with J774 cells and plating them onto BCYE (Buffered Charcoal Yeast Extract) agar plates. Overall, 25.56% (46/180) of water samples were Legionella-positive; fifty-two strains were isolated and two kinds of serogroups were co-detected from six water samples from 2015 to 2016. Bacterial concentrations ranged from 20 CFU/100 mL to 10,720 CFU/100 mL. In detail, the Legionella-positive rates of shower water, cooling tower water and hot springs water were 15.45%, 13.33%, and 62.5%, respectively. The main serogroups were LP1 (30.69%) and LP3 (28.85%) and all strains carried the dot gene. Among them, 52 isolates and another 10 former isolates were analyzed by PFGE. Nineteen distinct patterns were observed in 52 strains isolated from 2015 to 2016 with three patterns being observed in 10 strains isolated from 2009 to 2014. Seventy-three strains containing 52 from this study and 21 former isolates were selected for SBT analysis and divided into 25 different sequence types in 4 main clonal groups belonging to 4 homomorphic types. Ten strains were chosen to show their abilities to grow and multiply in J744 cells. Taken together, our results

  4. A Multiplex PCR for Detection of Mycoplasma pneumoniae, Chlamydophila pneumoniae, Legionella pneumophila, and Bordetella pertussis in Clinical Specimens

    National Research Council Canada - National Science Library

    McDonough, E. A; Barrozo, C. P; Russell, K. L; Metzgar, D

    2005-01-01

    ..., and Bordetella pertussis in uncultured patient specimens. These organisms cause similar symptomologies and are often not diagnosed because they are difficult to identify with classical methods such as culture and serology...

  5. Incidence and risk factors of Legionella pneumophila pneumonia during anti-tumor necrosis factor therapy: a prospective French study.

    Science.gov (United States)

    Lanternier, Fanny; Tubach, Florence; Ravaud, Philppe; Salmon, Dominique; Dellamonica, Pierre; Bretagne, Stephane; Couret, Marie; Bouvard, Beatrice; Debandt, Michel; Gueit, Isabelle; Gendre, Jean-Pierre; Leone, Jean; Nicolas, Nathalie; Che, Dider; Mariette, Xavier; Lortholary, Olivier

    2013-09-01

    Our objective was to describe the incidence and risk factors of legionellosis associated with tumor necrosis factor (TNF)-α antagonist use. From February 1, 2004, to January 31, 2007, we prospectively collected all cases of legionellosis among French patients receiving TNF-α antagonists in the Research Axed on Tolerance of Biotherapies (RATIO) national registry. We conducted an incidence study with the French population as a reference and a case-control analysis with four control subjects receiving TNF-α antagonists per case of legionellosis. Twenty-seven cases of legionellosis were reported. The overall annual incidence rate of legionellosis for patients receiving TNF-α antagonists, adjusted for age and sex, was 46.7 (95% CI, 0.0-125.7) per 100,000 patient-years. The overall standardized incidence ratio (SIR) was 13.1 (95% CI, 9.0-19.1; P < .0001) and was higher for patients receiving infliximab (SIR, 15.3 [95% CI, 8.5-27.6; P < .0001]) or adalimumab (SIR, 37.7 [95% CI, 21.9-64.9; P < .0001]) than etanercept (SIR, 3.0 [95% CI, 1.00-9.2; P = .06]). In the case-control analysis, exposure to adalimumab (OR, 8.7 [95% CI, 2.1-35.1]) or infliximab (OR, 9.2 [95% CI, 1.9-45.4]) vs etanercept was an independent risk factor for legionellosis. The incidence rate of legionellosis for patients receiving TNF-α antagonists is high, and the risk is higher for patients receiving anti-TNF-α monoclonal antibodies than soluble TNF-receptor therapy. In case of pneumonia occurring during TNF-α antagonist therapy, specific urine antigen detection should be performed and antibiotic therapy should cover legionellosis. ClinicalTrials.gov; No.: NCT00224562; URL: www.clinicaltrials.gov.

  6. Assessment of intercentre reproducibility and epidemiological concordance of Legionella pneumophila serogroup 1 genotyping by amplified fragment length polymorphism analysis

    DEFF Research Database (Denmark)

    Fry, N K; Bangsborg, Jette Marie; Bernander, S

    2000-01-01

    centre following a previously determined standard protocol. Results were analysed by the participants, using gel analysis software where available, and submitted to the coordinating centre. The coordinating centre reanalysed all results visually and selected data-sets with gel analysis software. Data...... analysis by participants yielded reproducibility (R) values of 0.20-1.00 and epidemiological concordance (E) values of 0.11-1.00, with 6 to 34 types. Following visual analysis by the coordinating centre, R=0.78-1.00, and E=0.67-1.00, with 10-20 types. Analysis of three data-sets by the coordinating centre.......00) and epidemiologically concordant (E=1.00), with good discrimination. The electropherograms generated are amenable to computer-aided analysis, but strict adherence to a previously defined laboratory protocol is required. Following designation of representative type strains and patterns, this method will be adopted...

  7. Surface-Associated Heat Shock Proteins of Legionella pneumophila and Helicobacter pylori: Roles in Pathogenesis and Immunity

    Directory of Open Access Journals (Sweden)

    P. S. Hoffman

    1999-01-01

    are defective in localizing Hsp60 onto their surface and are reduced approximately 1000-fold in their invasiveness towards HeLa cells. For the ulcer-causing bacterium Helicobacter pylori, surfaceassociated Hsp60 and Hsp70 mediate attachment to gastric epithelial cells. The increased expression of these Hsps following acid shock correlates with both increased association with and inflammation of the gastric mucosa. A role for Hsps in colonization, mucosal infection and in promoting inflammation is discussed. Infect. Dis. Obstet. Gynecol. 7:58–63, 1999.

  8. UV-C inactivation of Legionella rubrilucens

    Directory of Open Access Journals (Sweden)

    Schmid, Julian

    2017-04-01

    Full Text Available Background: Despite the great health significance of , there is only little information on their UV sensitivity. Besides only has been investigated so far. Methods: In this study has been spread on buffered charcoal yeast extract agar and irradiated with the 254 nm UV-C emission of a mercury vapor lamp. The disinfection success is measured by colony counting after incubation and comparison of the number of colonies on irradiated and unirradiated reference agar plates.Results: The average log-reduction dose is 1.08 mJ/cm for free which is at the lower end of the so far published Legionella log-reduction values, but all three species show similar sensitivities. Conclusion: The log-reduction dose of legionellae in amoebae has not been investigated, but with the observed high UV-C sensitivity for free , the idea of a future point-of-use disinfection by small UV-C LEDs in water-taps or shower heads appears to be realistic, even if legionellae are more resistant in amoebae.

  9. Comparative analysis of virulence traits between a Legionella feeleii strain implicated in Pontiac fever and a strain that caused Legionnaires' disease.

    Science.gov (United States)

    Wang, Changle; Saito, Mitsumasa; Tanaka, Tamami; Amako, Kazunobu; Yoshida, Shin-ichi

    2015-12-01

    Legionella strains of the same species and serogroup are known to cause Legionnaires' disease (a potentially fatal atypical pneumonia) or Pontiac fever (a mild, flu-like disease), but the bacterial factors that define these dramatic differences in pathology have not been elucidated. To gain a better understanding of these factors, we compared the characteristics of Legionella feeleii strains that were isolated from either a sample of freshwater implicated in an outbreak of Pontiac fever (ATCC 35072, serogroup 1, LfPF), or a patient with Legionnaires' disease (ATCC 38549, serogroup 2, LfLD). Growth of LfPF and LfLD in BYE broth was slower than the positive control, Legionella pneumophila strain JR32. However, LfLD grew faster than LfPF at 42 °C. After in vitro infection to J774 murine or U937 human macrophage cell lines and A549 human lung epithelial cell line, LfLD showed a higher cell infection rate, stronger internalization by host cells, and greater cytotoxicity than that of LfPF. Large amounts of IL-6 and IL-8 were secreted by human host cells after infection with LfLD, but not with LfPF. LfLD possessed mono-polar flagellum while LfPF was unflagellated. When LfLD was cultured at 25, 30 and 37 °C, the bacteria had higher motility rate at lower temperatures. Based on our results, this is the first study that showed distinct characteristics between LfPF and LfLD, which may give important leads in elucidating differences in their virulence. Copyright © 2015 Elsevier Ltd. All rights reserved.

  10. Survey of Legionella Species Found in Thai Soil

    Directory of Open Access Journals (Sweden)

    Tatiana C. Travis

    2012-01-01

    Full Text Available Members of the Gram-negative genus Legionella are typically found in freshwater environments, with the exception of L. longbeachae, which is present in composts and potting mixes. When contaminated aerosols are inhaled, legionellosis may result, typically as either the more serious pneumonia Legionnaires’ disease or the less severe flu-like illness Pontiac fever. It is presumed that all species of the genus Legionella are capable of causing disease in humans. As a followup to a prior clinical study of legionellosis in rural Thailand, indigenous soil samples were collected proximal to cases’ homes and workplaces and tested for the presence of legionellae by culture. We obtained 115 isolates from 22/39 soil samples and used sequence-based methods to identify 12 known species of Legionella represented by 87 isolates.

  11. Legionella (Legionnaires' Disease and Pontiac Fever): Fast Facts

    Science.gov (United States)

    ... Program for Laboratories For Media Water System Maintenance Water Management Toolkit Identify Buildings at Increased Risk Considerations When Working with Legionella Consultants Surveillance & Reporting Resources Materials Guidelines, Standards, and Laws Publications Related Links ...

  12. Legionella (Legionnaires' Disease and Pontiac Fever): Treatment and Complications

    Science.gov (United States)

    ... Program for Laboratories For Media Water System Maintenance Water Management Toolkit Identify Buildings at Increased Risk Considerations When Working with Legionella Consultants Surveillance & Reporting Resources Materials Guidelines, Standards, and Laws Publications File Formats ...

  13. Legionella (Legionnaires' Disease and Pontiac Fever): Signs and Symptoms

    Science.gov (United States)

    ... Program for Laboratories For Media Water System Maintenance Water Management Toolkit Identify Buildings at Increased Risk Considerations When Working with Legionella Consultants Surveillance & Reporting Resources Materials Guidelines, Standards, and Laws Publications Related Links ...

  14. Legionella (Legionnaires' Disease and Pontiac Fever): History and Disease Patterns

    Science.gov (United States)

    ... Program for Laboratories For Media Water System Maintenance Water Management Toolkit Identify Buildings at Increased Risk Considerations When Working with Legionella Consultants Surveillance & Reporting Resources Materials Guidelines, Standards, and Laws Publications Related Links ...

  15. Legionella (Legionnaires' Disease and Pontiac Fever): Causes and Transmission

    Science.gov (United States)

    ... Program for Laboratories For Media Water System Maintenance Water Management Toolkit Identify Buildings at Increased Risk Considerations When Working with Legionella Consultants Surveillance & Reporting Resources Materials Guidelines, Standards, and Laws Publications Related Links ...

  16. A Culture-Proven Case of Community-Acquired Legionella Pneumonia Apparently Classified as Nosocomial: Diagnostic and Public Health Implications

    Directory of Open Access Journals (Sweden)

    Annalisa Bargellini

    2013-01-01

    Full Text Available We report a case of Legionella pneumonia in a 78-year-old patient affected by cerebellar haemangioblastoma continuously hospitalised for 24 days prior to the onset of overt symptoms. According to the established case definition, this woman should have been definitely classified as a nosocomial case (patient spending all of the ten days in hospital before onset of symptoms. Water samples from the oncology ward were negative, notably the patient’s room and the oxygen bubbler, and the revision of the case history induced us to verify possible contamination in water samples collected at home. We found that the clinical strain had identical rep-PCR fingerprint of L. pneumophila serogroup 1 isolated at home. The description of this culture-proven case of Legionnaires’ disease has major clinical, legal, and public health consequences as the complexity of hospitalised patients poses limitations to the rule-of-thumb surveillance definition of nosocomial pneumonia based on 2–10-day incubation period.

  17. Trend of Legionella colonization in hospital water supply.

    Science.gov (United States)

    D'Alessandro, D; Fabiani, M; Cerquetani, F; Orsi, G B

    2015-01-01

    In many nosocomial Legionella outbreaks water distribution systems are the most frequent source of infection. Considering the hospital waterline old age, an investigation on colonization by Legionella spp was carried out in order to evaluate the pipeline system weaknesses and to implement environmental preventive measures. From 2004 to 2010, overall 97 samples from the water line were collected. The samples were analyzed according to the italian Legionella spp standard methods; water temperature, pH and residual free chlorine were determined at the time of collection. X2 test, exact-test and t-test were used to compare proportions and means. Overall 28 samples (23.7%) were positive for Legionella spp, and five of them (17.9%) exceeded the threshold level >104 cfu/L. The number of positive samples varied along the years, showing a significant increasing trend (X2 for trend = 11.5; pLegionella spp by comparison to negative ones showed a lower free chlorine concentration (0.08 mg/L vs 0.15 mg/L) and a higher water temperature (46.1° vs 42.7°). Actually the percentage of positive samples decreased significantly with the increasing in free chlorine in the water (X2 for trend = 8.53; pLegionella. All hospital buildings were colonized by Legionella spp, although 80% of samples >104 cfu/L occurred in the C-building. No cases of nosocomial legionellosis were reported during the study period. Hospital water system showed a diffuse colonization by Legionella spp, although the degree of contamination reached the threshold level (>104 cfu/L) only in a small percentage of samples, showing a substantial effectiveness of the control measures applied.

  18. von Willebrand Disease

    Science.gov (United States)

    ... for type 1 von Willebrand disease is called desmopressin. It causes a temporary increase in the von ... injection or by being sniffed into the nose. Desmopressin may also help some people with type 2 ...

  19. Interaktive Visualisierung von Strukturmechaniksimulationen

    OpenAIRE

    Sommer, Ove

    2003-01-01

    A) Motivation und Problemstellung Der Computer ist heute eines der wichtigsten Hilfsmittel beim Entwurf und der Entwicklung von Fahrzeugen. Zunächst wurde er von Konstrukteuren für das Computer Aided Design (CAD) von virtuellen Fahrzeugmodellen eingesetzt. Inzwischen ist er in vielen anderen Bereichen der Fahrzeugentwicklung unentbehrlich geworden: der Entwicklungszyklus von Automobilen ist durch die Computer-gestützte numerische Simulation substanziell verkürzt worden. An virtuellen Prot...

  20. Métodos analíticos para el estudio de Legionella Methods for Legionella detection

    Directory of Open Access Journals (Sweden)

    Carmen Pelaz Antolín

    2006-12-01

    Full Text Available Los ensayos para la determinación de Legionella en muestras de agua son uno de los aspectos contemplados en la legislación española sobre prevención de legionelosis. La periodicidad de estos ensayos en función del tipo de instalación, los laboratorios que los realizan, y las acciones correctoras que derivan de ellos en función de los recuentos bacterianos son acciones incluidas en los planes de mantenimiento preventivo de las instalaciones consideradas de riesgo (Real Decreto 865/2003. La comparación de nuestra legislación con otras legislaciones o recomendaciones adoptadas en otros paises (Reino Unido, Francia, Australia, América permite conocer nuestro grado de exigencia en relación a algunos de los parámetros contemplados. El cultivo de la bacteria es el método de referencia para la detección de Legionella en muestras de agua y existen varios ensayos normalizados, como los estándares ISO 11731/98 y 2004 y NF T 90- 431/2003 (AFNOR. Para ayudar a la interpretación de los resultados, los ensayos deben reflejar el estándar en el que se basan y el límite de detección del método, que no debe ser superior a 100 ufc/L. Además, los laboratorios que realizan estos ensayos deben estar acreditados por nuestra entidad de acreditación ENAC. En los últimos años se han desarrollado métodos rápidos de detección de la bacteria basados en la amplificación de ADN cromosómico en muestras de agua mediante reacciones de PCR. El desarrollo científico de estos métodos va por delante del desarrollo reglamentario, y los ensayos de PCR no deben desplazar a los ensayos de cultivo en cumplimiento de las normativas vigentes, sino que deben complementarlo.Assays for Legionella detection in water samples are one of the aspects included in the Spanish legislation on prevention of Legionnaires ́ disease. The frequency of these assays, laboratories that carry out them, and the required actions that derive from them, regarding colony counts, are

  1. Legionella control in water systems using copper and silver ion generation systems

    OpenAIRE

    Bedford, Birgitta

    2012-01-01

    Legionella can cause human disease which can be fatal. Routine monitoring for Legionella in water systems is not recommended by UK authorities. Evidence of the efficacy of control modalities against Legionella in these water systems is, therefore, not available. Although studies have been conducted with copper and silver ionization on its efficacy against Legionella and on its value in reducing hospital-acquired legionellosis, little evidence of its efficacy is available fro...

  2. Clinical predictors for Legionella in patients presenting with community-acquired pneumonia to the emergency department

    Directory of Open Access Journals (Sweden)

    Frei Reno

    2009-01-01

    Full Text Available Abstract Background Legionella species cause severe forms of pneumonia with high mortality and complication rates. Accurate clinical predictors to assess the likelihood of Legionella community-acquired pneumonia (CAP in patients presenting to the emergency department are lacking. Methods We retrospectively compared clinical and laboratory data of 82 consecutive patients with Legionella CAP with 368 consecutive patients with non-Legionella CAP included in two studies at the same institution. Results In multivariate logistic regression analysis we identified six parameters, namely high body temperature (OR 1.67, p Legionella CAP. Using optimal cut off values of these six parameters, we calculated a diagnostic score for Legionella CAP. The median score was significantly higher in Legionella CAP as compared to patients without Legionella (4 (IQR 3–4 vs 2 (IQR 1–2, p Legionella pneumonia. Conversely, of the 73 patients (16% with ≥4 points, 66% of patients had Legionella CAP. Conclusion Six clinical and laboratory parameters embedded in a simple diagnostic score accurately identified patients with Legionella CAP. If validated in future studies, this score might aid in the management of suspected Legionella CAP.

  3. Implementation of a Legionella Ordinance for Multifamily Housing, Garland, Texas.

    Science.gov (United States)

    Whitney, Ellen A; Blake, Sarah; Berkelman, Ruth L

    The incidence of legionellosis has sharply increased in the United States as a result of contaminated water systems. Jurisdictions across the country are considering whether to develop and implement regulations to protect individuals against Legionnaires' disease with its associated high morbidity and mortality. This article sheds light on the implementation and effectiveness of a 2005 citywide Legionella testing mandate of multifamily housing cooling towers in Garland, Texas. This ordinance has been in place for more than 10 years and represents the first of its kind in the United States to mandate routine testing of cooling towers for Legionella in multifamily housing. We utilized a mix of both qualitative and quantitative methods to explore the development, adoption, and implementation of the ordinance. Phone interviews were conducted with individuals from the City of Garland Health Department and apartment managers. Quantitative data included public health surveillance data on legionellosis. Barriers and facilitators of implementation, number and percentage of cooling towers from multifamily housing units that tested positive for Legionella by year, and number of legionellosis cases by year in Garland, Texas. Study outcomes highlight key themes that facilitated the successful implementation of the Legionella testing mandate, including the importance of timing, leadership support, stakeholder engagement, and education and outreach. The number of contaminated cooling towers was reduced over time. Mandatory monitoring for legionella in a local jurisdiction may result in reduced risk of legionellosis from cooling towers through raising awareness and education of building owners and managers about the need to prevent, detect, and remediate legionella contamination in their building water systems. Garland, Texas, broke new ground in the United States in moving toward primary prevention of legionellosis. The ordinance may be useful both in serving to educate and

  4. Legionella saoudiensis sp. nov., isolated from a sewage water sample.

    Science.gov (United States)

    Bajrai, Leena Hussein; Azhar, Esam Ibraheem; Yasir, Muhammad; Jardot, Priscilla; Barrassi, Lina; Raoult, Didier; La Scola, Bernard; Pagnier, Isabelle

    2016-11-01

    A Gram-stain-negative, bacilli-shaped bacterial strain, LS-1T, was isolated from a sewage water sample collected in Jeddah, Saudi Arabia. The taxonomic position of strain LS-1T was investigated using a polyphasic taxonomic approach. Phylogenetic analysis based on 16S rRNA gene sequences and those of four other genes indicated that strain LS-1T belongs to the genus Legionella in the family Legionellaceae. Regarding the 16S rRNA gene, the most closely related species are Legionella rowbothamii LLAP-6T (98.6 %) and Legionella lytica L2T (98.5 %). The mip gene sequence of strain LS-1T showed 94 % sequence similarity with that of L. lytica L2T and 93 % similarity with that of L. rowbothamii LLAP-6T. Strain LS-1T grew optimally at a temperature of 32 °C on a buffered charcoal yeast extract (BCYE) agar plate in a 5 % CO2 atmosphere and had a flagellum. The combined phylogenetic, phenotypic and genomic sequence data suggest that strain LS-1T represents a novel species of the genus Legionella, for which the name Legionella saoudiensis sp. nov. is proposed. The type strain is LS-1T (=DSM 101682T=CSUR P2101T).

  5. Lernen von und bei Max von Laue

    Science.gov (United States)

    Thiessen, P. A.

    Erinnerungen an die Entstehungsgeschichte und die Frühzeiten der Röntgenstrukturanalyse.Translated AbstractLearning at Max von LaueRemembrances of the historical rise and the early times of X-ray structure analysis.

  6. Legionella jamestowniensis fatal pneumonia in an immunosuppressed man.

    Science.gov (United States)

    Edelstein, Paul H

    2017-01-01

    A fatal case of Legionnaires' disease caused by Legionella jamestowniensis is reported in a severely immunocompromised patient with metastatic hepatocellular carcinoma, and liver and kidney transplants. L. jamestowniensis was cultured from two separate respiratory tract specimens and a PCR test for Legionella species was also positive from the same specimens. This is apparently the first reported case of human infection caused by L. jamestowniensis. Copyright © 2016 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

  7. The Legionella Kinase LegK2 Targets the ARP2/3 Complex To Inhibit Actin Nucleation on Phagosomes and Allow Bacterial Evasion of the Late Endocytic Pathway

    Science.gov (United States)

    Michard, Céline; Sperandio, Daniel; Baïlo, Nathalie; Pizarro-Cerdá, Javier; LeClaire, Lawrence; Chadeau-Argaud, Elise; Pombo-Grégoire, Isabel; Hervet, Eva; Vianney, Anne; Gilbert, Christophe; Faure, Mathias; Cossart, Pascale

    2015-01-01

    ABSTRACT Legionella pneumophila, the etiological agent of legionellosis, replicates within phagocytic cells. Crucial to biogenesis of the replicative vacuole is the Dot/Icm type 4 secretion system, which translocates a large number of effectors into the host cell cytosol. Among them is LegK2, a protein kinase that plays a key role in Legionella infection. Here, we identified the actin nucleator ARP2/3 complex as a target of LegK2. LegK2 phosphorylates the ARPC1B and ARP3 subunits of the ARP2/3 complex. LegK2-dependent ARP2/3 phosphorylation triggers global actin cytoskeleton remodeling in cells, and it impairs actin tail formation by Listeria monocytogenes, a well-known ARP2/3-dependent process. During infection, LegK2 is addressed to the Legionella-containing vacuole surface and inhibits actin polymerization on the phagosome, as revealed by legK2 gene inactivation. Consequently, LegK2 prevents late endosome/lysosome association with the phagosome and finally contributes to remodeling of the bacterium-containing phagosome into a replicative niche. The inhibition of actin polymerization by LegK2 and its effect on endosome trafficking are ARP2/3 dependent since it can be phenocopied by a specific chemical inhibitor of the ARP2/3 complex. Thus, LegK2-ARP2/3 interplay highlights an original mechanism of bacterial virulence with an unexpected role in local actin remodeling that allows bacteria to control vesicle trafficking in order to escape host defenses. PMID:25944859

  8. Onderzoek naar aanwezigheid van legionella in biologische luchtwassers bij stallen

    NARCIS (Netherlands)

    Melse, R.W.; Schalk, J.A.C.; Bartels, A.A.

    2015-01-01

    In an earlier study it was concluded, based on literature and interviews, that biotrickling filters might be responsible for airborne transmission of Legionella bacteria. Risk factors in this type of systems are recirculating of trickling water with a pH close to neutral and the use of spray nozzles

  9. Kinetic analysis of Legionella inactivation using ozone in wastewater.

    Science.gov (United States)

    Li, Jun; Li, Kunquan; Zhou, Yan; Li, Xuebin; Tao, Tao

    2017-02-01

    Legionella inactivation using ozone was studied in wastewater using kinetic analysis and modeling. The experimental results indicate that the relationship between the ozone concentration, germ concentration, and chemical oxygen demand (COD) can be used to predict variations in germ and COD concentrations. The ozone reaction with COD and inactivation of Legionella occurred simultaneously, but the reaction with COD likely occurred at a higher rate than the inactivation, as COD is more easily oxidized by ozone than Legionella. Higher initial COD concentrations resulted in a lower inactivation rate and higher lnN/N0. Higher temperature led to a higher inactivation efficiency. The relationship of the initial O3 concentration and Legionella inactivation rate was not linear, and thus, the Ct value required for a 99.99% reduction was not constant. The initial O3 concentration was more important than the contact time, and a reduction of the initial O3 concentration could not be compensated by increasing the contact time. The Ct values were compared over a narrow range of initial concentrations; the Ct values could only be contrasted when the initial O3 concentrations were very similar. A higher initial O3 concentration led to a higher inflection point value for the lnN/N0 vs C0t curve. Energy consumption using a plasma corona was lower than when using boron-doped diamond electrodes. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Comparing methods of determining Legionella spp. in complex water matrices.

    Science.gov (United States)

    Díaz-Flores, Álvaro; Montero, Juan Carlos; Castro, Francisco Javier; Alejandres, Eva María; Bayón, Carmen; Solís, Inmaculada; Fernández-Lafuente, Roberto; Rodríguez, Guillermo

    2015-04-29

    Legionella testing conducted at environmental laboratories plays an essential role in assessing the risk of disease transmission associated with water systems. However, drawbacks of culture-based methodology used for Legionella enumeration can have great impact on the results and interpretation which together can lead to underestimation of the actual risk. Up to 20% of the samples analysed by these laboratories produced inconclusive results, making effective risk management impossible. Overgrowth of competing microbiota was reported as an important factor for culture failure. For quantitative polymerase chain reaction (qPCR), the interpretation of the results from the environmental samples still remains a challenge. Inhibitors may cause up to 10% of inconclusive results. This study compared a quantitative method based on immunomagnetic separation (IMS method) with culture and qPCR, as a new approach to routine monitoring of Legionella. First, pilot studies evaluated the recovery and detectability of Legionella spp using an IMS method, in the presence of microbiota and biocides. The IMS method results were not affected by microbiota while culture counts were significantly reduced (1.4 log) or negative in the same samples. Damage by biocides of viable Legionella was detected by the IMS method. Secondly, a total of 65 water samples were assayed by all three techniques (culture, qPCR and the IMS method). Of these, 27 (41.5%) were recorded as positive by at least one test. Legionella spp was detected by culture in 7 (25.9%) of the 27 samples. Eighteen (66.7%) of the 27 samples were positive by the IMS method, thirteen of them reporting counts below 10(3) colony forming units per liter (CFU l(-1)), six presented interfering microbiota and three presented PCR inhibition. Of the 65 water samples, 24 presented interfering microbiota by culture and 8 presented partial or complete inhibition of the PCR reaction. So the rate of inconclusive results of culture and PCR was 36

  11. Environmental Legionella spp. collected in urban test sites of South East Queensland, Australia, are virulent to human macrophages in vitro.

    Science.gov (United States)

    Lawrence, Amba; Eglezos, Sofroni; Huston, Wilhelmina

    2016-01-01

    Legionellae are frequent contaminants of potable water supplies, resulting in sporadic infections and occasional outbreaks. Isolates of Legionella were collected from urban test sites within South East Queensland and evaluated for their virulence potential in vitro. Two strains (from the species Legionella londiniensis and Legionella quinlivanii) were demonstrated to have the ability to infect human macrophages, while a strain from the species Legionella anisa did not maintain an infection over the same time course. This suggests that the spectrum of urban environmentally associated Legionella with potential to cause human disease might be greater than currently considered. Copyright © 2015 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  12. Conventional and Alternative Disinfection Methods of Legionella in Water Distribution Systems – Review

    Directory of Open Access Journals (Sweden)

    Pūle Daina

    2016-12-01

    Full Text Available Prevalence of Legionella in drinking water distribution systems is a widespread problem. Outbreaks of Legionella caused diseases occur despite various disinfectants are used in order to control Legionella. Conventional methods like thermal disinfection, silver/copper ionization, ultraviolet irradiation or chlorine-based disinfection have not been effective in the long term for control of biofilm bacteria. Therefore, research to develop more effective disinfection methods is still necessary.

  13. A Cluster of Legionella-Associated Pneumonia Cases in a Population of Military Recruits

    Science.gov (United States)

    2007-06-01

    Naval Health Research Center A Cluster of Legionella -Associated Pneumonia Cases in a Population of Military Recruits E. A. McDonough...0095-1137/07/$08.000 doi:10.1128/JCM.02359-06 A Cluster of Legionella -Associated Pneumonia Cases in a Population of Military Recruits Erin A...17 April 2007 A Legionella cluster was identified through retrospective PCR analysis of 240 throat swab samples from X-ray-confirmed pneumonia cases

  14. Legionellosis and Lung Abscesses: Contribution of Legionella Quantitative Real-Time PCR to an Adapted Followup

    Directory of Open Access Journals (Sweden)

    G. Descours

    2013-01-01

    Full Text Available We report a case of severe Legionnaires' disease (LD complicated by a lung abscess in an immunocompetent patient who required ECMO therapy and thoracic surgery. The results of repeated Legionella quantitative real-time PCR performed on both sera and respiratory samples correlated with the LD severity and the poor clinical outcome. Moreover, the PCR allowed for the detection of Legionella DNA in the lung abscess specimen, which was negative when cultured for Legionella. This case report provides a logical basis for further investigations to examine whether the Legionella quantitative PCR could improve the assessment of LD severity and constitute a prognostic marker.

  15. von Willebrand Disease

    Science.gov (United States)

    ... the condition. For example, the school nurse, teacher, daycare provider, coach, or any leader of afterschool activities ... MedlinePlus) Von Willebrand Disease (MedlinePlus) Building 31 31 Center Drive Bethesda, MD 20892 Learn more about getting ...

  16. Carl von Clausewitz

    DEFF Research Database (Denmark)

    Højrup, Thomas

    2009-01-01

    Forskningsbaseret præsentation af Carl von Clausewitz' krigsteori og dens betydning for statsbegrebet og udviklingen af teori om statsdannelse og statssystem. Krigsbegrebets betydning for politikbegrebet ekspliciteres, og de indre relationer imellem de to begreber demonstreres med konkrete eksemp...

  17. Ellis Von Creveld Syndrome

    Directory of Open Access Journals (Sweden)

    Afshar H

    1999-01-01

    Full Text Available One patient with Ellis Von Creveld syndrome contains: dwarfism, congenital heart"ndisease, ectodermal dysplasia, polyductyly, an abnormally wide labial frenum and maxillary"nmolars with single root.

  18. Estudo comparativo da apresentação clínica da pneumonia a Legionella e outras pneumonias adquiridas na comuuidade

    Directory of Open Access Journals (Sweden)

    Nieves Sopena

    1998-11-01

    Full Text Available RESUMO: A Legionella pneumophila (LP é considerada em muitos estudos como uma das três causas mais comuns de pneumonia adquirida na comunidade (2,4 e a segunda causa em termos de gravidade.O principal objectivo do presente estudo realizado no Hospital da Universidade Autónoma de Barcelona, foi comparar os diferentes aspectos clínicos, biológicos e radiológicos da Pneumonia a Legionella pneumophilla e das outras Pneumonias adquiridas oa comunidade (PAC, de modo a auxiliar o diagnóstico precoce da pneumonia a LP.Foi realizado um estudo prospectivo de 392 doentes com PAC. Procedeu-se à análise comparativa dos aspectos epidemiológicos (hospitalizações ou viagens recentes, residência próximo de escavações ou trabalhos de constrção, contacto com animais, aves, demográficos (idade e sexo, clinicos (febre, tosse, expectoração, toracalgia, dispneia, cefaleias, confusão mental, dor aboominal, náuseas, vómitos, diarreia, artromialgias, dias de evolução, antibioterapia prévia, patologiaassociada, analiticos (leucocitose, natrémia­Na, creatina Kinase-CK, aspartato aminotransferase­AST e radiológicos. Estes aspectos foram estudados em 48 doentes com PAC por LP e 125 doentes com PAC de outra etiologia (68 por Streptococcus pneumoniae, 41 por Clamydia pneumoniae, 5 por Mycoplasma pneumoniae, 4 por Coxiella burnetii, 3 por Pseudomonas aeruginosa, 2 por Haemophilus injluenzae e 2 por Nocardia.A análise unifactorial mostrou que a PAC por LP foi mais frequente hos doentes de meia idade, do sexo masculino, em saúde aparente, mas com hábitos alcoó1icos acentuados, relativamente aos doentes com PAC de outra etioçogia. Também a auseocia de resposta aos fármacos beta-lactamicos prévios, cefaleias, diarreia, hiponatrémia grave e a elevação dos níveis de creatina Kinase sérica (CK foram mais frequentes na PAC por LP, enquanto que a tosse, expectoração e a toracalgia., foram mais frequeotes na pneumonia bacteriana de outra

  19. Contamination of water reservoirs to Legionella in khorramabad hospitals

    Directory of Open Access Journals (Sweden)

    seyed hamed Mirhossaini

    2009-04-01

    Full Text Available Background: Legionella is a negative aquatic bacterium and one of the most common nosocomial pathogen. Hospital environment in case of growth, aerosol transmission system and endangered individuals are the high potential location for growth and prevalence of this agent. The suitable temperature in water reservoirs and water distribution system empowered the growth of this bactria. The purpose of this investigation is the study of legionella presence in khorramabad water distribution system. Materials and Methods: Sampling performed with fifteen-day periods of each cold and hot hospital water reservoirs and also cold and hot water taps in those hospital wards which have more pathogens. Each of samples concentrate high vulnerable membrane and from each sample 2 plates were cultured with BCYE and GVPC optional culture media and the growth of bacteria in third and seventh and tenth days were controlled and registered. Results: From 240 samples of five Khorramabad hospitals 41.7 percent of the samples were positive. The percent of positive samples of Ashayer, Tamin ejtemaee, Tohid, and Asalian were respectively 68.8, 45.5, 33.3, 9.1 and 36.4 percent and the residual mean chlorine of samples were respectively 0.38, 0.52, 0.46, 0.82 and 0.62mg/l. The most positive samples related to hot shower and the lowest value related to cold water taps. Conclusion: In spite of the fact that all hospitals used treated water, but from 240 collected samples, 100 samples in different sections of hospital were positive these results show direct relation between residual chlorine value and presence of legionella, by the manner that in 0.6 mg/l and higher values of residual chlorine none of samples were positive. So usually the residual chlorine value in water distribution system is not enough to legionella against.

  20. Chronic endocarditis due to Legionella anisa: a first case difficult to diagnose

    Directory of Open Access Journals (Sweden)

    F. Compain

    2015-11-01

    Full Text Available Endocarditis due to Legionella spp. is uncommon but presumably underestimated given the prevalence of Legionellae in the environment. We report a first and unusual case of chronic native valve endocarditis due to L. anisa and advocate that the diagnosis of endocarditis be made collaboratively between the cardiologist, surgeon, microbiologist and pathologist.

  1. The role of biofilms and protozoa in Legionella pathogenesis: implications for drinking water

    Science.gov (United States)

    Current models to study Legionella pathogenesis include the use of primary macrophages and monocyte cell lines, various free-living protozoan species and murine models of pneumonia. However, there are very few studies of Legionella spp. pathogenesis aimed at associating the role ...

  2. The E. coli immunosorbent as used in serodiagnosis of Legionella infections studied by crossed immunoelectrophoresis

    DEFF Research Database (Denmark)

    Bangsborg, Jette Marie; Friis-Møller, A; Rechnitzer, C

    1988-01-01

    In this study we investigated an immunosorbent, E. coli blocking fluid (BF), proposed for use in the Legionella Indirect Fluorescent Antibody Test (IFA). With crossed immunoelectrophoresis (CIE) of clinically relevant Legionella species, only one heat-stable antigen (no. 1) cross...

  3. The presence and growth of Legionella species in thermostatic shower mixer taps: an exploratory field study

    NARCIS (Netherlands)

    Joost van Hoof; P.W.J.J. van der Wielen; L. Hornstra; E. van der Blom; O.W.W. Nuijten

    2014-01-01

    Legislation in the Netherlands requires routine analysis of drinking water samples for cultivable Legionella species from high-priority installations. A field study was conducted to investigate the presence of Legionella species in thermostatic shower mixer taps. Water samples and the interior of

  4. Community-acquired pneumonia by Legionella pneumophila. Do we need to include new recommendations for inflammatory bowel disease patients under immunomodulators?

    National Research Council Canada - National Science Library

    Fernández Llamas, Tania; Sánchez Torres, Antonio; Egea Valenzuela, Juan

    2016-01-01

    .... Following this and other similar cases reported in literature, we propose to include recommendations to prevent this infection in patients starting treatment with immunosuppressant drugs through...

  5. Von Braun Fountain

    Science.gov (United States)

    1999-01-01

    A fountain representing a rocket launch was dedicated in the Von Braun courtyard outside of Building 4200 at Marshall Space Flight Center during the weekend celebrating the 30th arniversary of the Apollo 11 lunar landing. On hand for the festivities were many of the Saturn and Apollo astronauts.

  6. Rhizostomeen von Manila

    NARCIS (Netherlands)

    Stiasny, G.

    1924-01-01

    Die hier beschriebene kleine Scyphomedusen-Sammlung wurde von Herrn Director P. B. Sivickis, Dept. of Zoology, University of Philippines, Manila, dem Rijksmuseum van Natuurlijke Historie in Leiden überwiesen. Das Material wurde im December 1922 in Manila-bay gefischt und befindet sich in bestem

  7. Karman, Prof. Theodore von

    Indian Academy of Sciences (India)

    Home; Fellowship. Fellow Profile. Elected: 1961 Honorary. Karman, Prof. Theodore von. Date of birth: 11 May 1881. Date of death: 6 May 1963. YouTube; Twitter; Facebook; Blog. Academy News. IAS Logo. Theory Of Evolution. Posted on 23 January 2018. Joint Statement by the Three Science Academies of India on the ...

  8. Schmetterlinge von Madeira

    NARCIS (Netherlands)

    Martin, K.

    1941-01-01

    Wer auf einer ozeanischen Insel Schmetterlinge sammeln will, darf seine Erwartungen nicht zu hoch spannen; aber dennoch waren meine Frau und ich sehr enttäuscht, als uns auf einer Reise nach Madeira von einer dort angesessenen, gebildeten Dame gesagt wurde, sie habe auf der Insel niemals einen

  9. El hospital ante un brote prolongado de legionelosis The hospital faced with a prolonged Legionella outbreak

    Directory of Open Access Journals (Sweden)

    Juan Antonio Fernández

    2004-08-01

    Full Text Available Objetivo: Describir los recursos asistenciales utilizados en pacientes hospitalizados por neumonía por Legionella durante un brote ocurrido en Alcoy y compararlos con los empleados en otras neumonías. Métodos y resultados: Mediante un protocolo, se analiza y compara a 177 pacientes con neumonía por Legionella con 180 pacientes ingresados por otras neumonías, y se describe su tratamiento y los recursos empleados. La claritromicina fue el antibiótico más utilizado en ambos grupos. La estancia hospitalaria fue similar, pero los requerimientos de ventilación mecánica y cuidados intensivos fueron superiores para Legionella. La hospitalización domiciliaria se utilizó con éxito en un 15,6% de los pacientes con Legionella y en un 11,3% de los demás. El uso de oxigenoterapia domiciliaria al alta fue menor para Legionella (7,8% que para el resto de neumonías (16,7%. Conclusiones: Se destaca la mayor necesidad de ventilación mecánica en la neumonía por Legionella y los buenos resultados de la hospitalización domiciliaria dentro de los nuevos sistemas de gestión.Objective: To describe the health resources used in patients hospitalized with Legionella pneumonia during an outbreak of Legionnaire's disease in Alcoy (Spain, and to compare them with those used in other forms of pneumonia. Methods and results: Using a clinical protocol, 177 Legionella pneumonia patients were compared with 180 patients hospitalized for other types of pneumonia. Data on therapy and the resources used were collected. The most common antibiotic treatment in both groups was clarithromycin, but intensive care and mechanical ventilation requirements were greater in Legionella pneumonia. Home-based hospital care was successfully used in 15.6% of patients with Legionella pneumonia and in 11.3% of those with other types of pneumonia. Home oxygen therapy after discharge was less frequent in the Legionella pneumonia group (7.8% than in the group with non-Legionella

  10. NCBI nr-aa BLAST: CBRC-XTRO-01-3137 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-XTRO-01-3137 ref|YP_096446.1| florfenicol efflux pump [Legionella pneumophila subsp. pneumophila... str. Philadelphia 1] gb|AAU28499.1| florfenicol efflux pump [Legionella pneumophila subsp. pneumophila str. Philadelphia 1] YP_096446.1 0.36 29% ...

  11. NCBI nr-aa BLAST: CBRC-CREM-01-1332 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-CREM-01-1332 ref|YP_094231.1| chloramphenicol resistance protein [Legionella pneumophila... subsp. pneumophila str. Philadelphia 1] gb|AAU26284.1| chloramphenicol resistance protein [Legionella pneumophila... subsp. pneumophila str. Philadelphia 1] YP_094231.1 2e-30 36% ...

  12. NCBI nr-aa BLAST: CBRC-HSAP-08-0029 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-HSAP-08-0029 ref|YP_096248.1| nitric oxide reductase, subunit B [Legionella pneumophila... subsp. pneumophila str. Philadelphia 1] gb|AAU28301.1| nitric oxide reductase, subunit B [Legionella pneumophila subsp. pneumophila str. Philadelphia 1] YP_096248.1 8.0 21% ...

  13. NCBI nr-aa BLAST: CBRC-SARA-01-1999 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-SARA-01-1999 ref|YP_095353.1| drug resistance transporter, Bcr/CflA [Legionella pneumophila... subsp. pneumophila str. Philadelphia 1] gb|AAU27406.1| drug resistance transporter, Bcr/CflA [Legionella pneumophila... subsp. pneumophila str. Philadelphia 1] YP_095353.1 0.29 21% ...

  14. Influence of climate and geography on the occurrence of Legionella and amoebae in composting facilities.

    Science.gov (United States)

    Conza, Lisa; Casati Pagani, Simona; Gaia, Valeria

    2014-11-24

    The incidence of Legionnaires' disease (LD) in southern Switzerland is three times higher than in northern Switzerland. Climatic and geographic factors may be potential causes for this difference.We studied the prevalence of Legionella and free-living amoebae (FLA) in compost and bioaerosol in two Swiss regions to understand the role of climate and geography in the transmission of LD. We also tried to investigate whether or not compost storage duration would influence the composition of Legionella and FLA communities. A larger proportion of compost heaps in facilities from southern Switzerland harbor more diverse Legionella compared to the north (P=0.0146). FLA were isolated from composts in northern facilities at slightly higher rates (88.2% vs. 69.2%) and at lower rates from bioaerosols (6.3% vs. 13%) than in southern Switzerland. The diversity of FLA was higher in northern than in southern Switzerland (80% vs. 65%). A general decrease in the presence and variety of species was observed with decreasing compost storage time length. A discriminant model showed that values of vapour pressure, relative humidity and temperature distinguish the two regions, which were also characterised by different contamination rates by FLA and Legionella. The duration of outdoor storage may favour contamination of the compost by Legionella, and may increase the number and isolation of Legionella naturally occurring in compost. The climate in the south seems to favour higher Legionella contamination of compost heaps: this could explain the higher incidence of LD in southern Switzerland.

  15. Legionella shows a diverse secondary metabolism dependent on a broad spectrum Sfp-type phosphopantetheinyl transferase

    Directory of Open Access Journals (Sweden)

    Nicholas J. Tobias

    2016-11-01

    Full Text Available Several members of the genus Legionella cause Legionnaires’ disease, a potentially debilitating form of pneumonia. Studies frequently focus on the abundant number of virulence factors present in this genus. However, what is often overlooked is the role of secondary metabolites from Legionella. Following whole genome sequencing, we assembled and annotated the Legionella parisiensis DSM 19216 genome. Together with 14 other members of the Legionella, we performed comparative genomics and analysed the secondary metabolite potential of each strain. We found that Legionella contains a huge variety of biosynthetic gene clusters (BGCs that are potentially making a significant number of novel natural products with undefined function. Surprisingly, only a single Sfp-like phosphopantetheinyl transferase is found in all Legionella strains analyzed that might be responsible for the activation of all carrier proteins in primary (fatty acid biosynthesis and secondary metabolism (polyketide and non-ribosomal peptide synthesis. Using conserved active site motifs, we predict some novel compounds that are probably involved in cell-cell communication, differing to known communication systems. We identify several gene clusters, which may represent novel signaling mechanisms and demonstrate the natural product potential of Legionella.

  16. Existence and control of Legionella bacteria in building water systems: A review.

    Science.gov (United States)

    Springston, John P; Yocavitch, Liana

    2017-02-01

    Legionellae are waterborne bacteria which are capable of causing potentially fatal Legionnaires' disease (LD), as well as Pontiac Fever. Public concern about Legionella exploded following the 1976 outbreak at the American Legion conference in Philadelphia, where 221 attendees contracted pneumonia and 34 died. Since that time, a variety of different control methods and strategies have been developed and implemented in an effort to eradicate Legionella from building water systems. Despite these efforts, the incidence of LD has been steadily increasing in the U.S. for more than a decade. Public health and occupational hygiene professionals have maintained an active debate regarding best practices for management and control of Legionella. Professional opinion remains divided with respect to the relative merits of performing routine sampling for Legionella, vs. the passive, reactive approach that has been largely embraced by public health officials and facility owners. Given the potential risks and ramifications associated with waiting to assess systems for Legionella until after disease has been identified and confirmed, a proactive approach of periodic testing for Legionella, along with proper water treatment, is the best approach to avoiding large-scale disease outbreaks.

  17. Sequence analysis of the Legionella micdadei groELS operon

    DEFF Research Database (Denmark)

    Hindersson, P; Høiby, N; Bangsborg, Jette Marie

    1991-01-01

    A 2.7 kb DNA fragment encoding the 60 kDa common antigen (CA) and a 13 kDa protein of Legionella micdadei was sequenced. Two open reading frames of 57,677 and 10,456 Da were identified, corresponding to the heat shock proteins GroEL and GroES, respectively. Typical -35, -10, and Shine-Dalgarno heat......, Western blot analysis with an L. micdadei specific anti-groEL antibody did not reveal a significant increase in the amount of the GroEL protein during heat shock in L. micdadei or in the recombinant E. coli expressing L. micdadei GroEL....

  18. Kommunikative Konstitution von Organisationen

    DEFF Research Database (Denmark)

    Schoeneborn, Dennis; Wehmeier, Stefan

    2014-01-01

    Beiträge zur Unternehmenskommunikation behandeln in der Regel (strategisch) geplante interne oder externe Kommunikation. Der vorliegende Beitrag stellt einen alternativen Theorieansatz vor, der aktuell im nordamerikanischen Forschungsgebiet „Organizational Communication“ vorherrschend ist....... zunehmende Aufmerksamkeit erhält Der sogenannten „CCO-Perspektive“ („Communicative Constitution of Organizations“) zufolge bestehen Unternehmen aus einer Vielzahl von internen und externen Kommunikationspraktiken, die nur in bedingtem Maße strategisch steuerbar sind. Zugleich sind es eben diese...

  19. Kulturelle Aspekte von Textsorten

    Directory of Open Access Journals (Sweden)

    Siegfried Heusinger

    1998-12-01

    Full Text Available Ich fasse meine Überlegungen zu kulturellen Aspekten von Textsorten zusammen: 1. Mit unseren Kulturverständnis ist die terminologisierte Wortgruppe "interkulturelle Kommunikation" nicht zu begreifen als Kommunikation zwischen Menschen verschiedener kultureller Bindungen, sondern sie ist nicht zu begreifen als Kommunikation zwischen Menschen verschiedener kultureller Bindungen, sondern sie ist ethnisch wie auch ethisch angepasstes sprachlichkommunikatives Verhalten an kulturelle Traditionen der jeweils für den Kommunkations- akt vereinbarten Sprache. 2. Das Adjektiv "interkulturell" referiert auf idiomatisierte Wendungen im zwischenmenschlichen Kontakt wie auch auf kulturell gebundene Unterschiede in der Benennungsmotivation (z.B. anloben/österr./, vereidigen /dt./ 3. Das Adjektiv "interkulturell" ist ebenfalls zu beziehen auf den Austausch von Erfahrungen und Ideen, auf den Dialog von Kulturen. 4. Textsorten existieren in ihrer kulturellen in ihrer kulturellen Tradition, wobei das Kulturspezifische vornehmlich in traditionellen (aber entwicklungsoffenen Formulierungs- und Gestaltungsmustern zu suchen iost. Andere Textsortencharakteristika wie die situative Einbettung, die kommunikative Zwecksetzung, die thematische Bindung und die Komposition haben offenbar kaum eine kulturspezifische Prägung. 5. Im Trend zur Internationalisierung der Textsorten erweisen sich die kulturell geprägten Textmustervor allem zwischen angrenzenden Nationen häufig als affin.

  20. Nutzerorientiertes Management von materiellen und immateriellen Informationsobjekten

    OpenAIRE

    Hübsch, Chris

    2001-01-01

    Schaffung einer stabilen, erweiterbaren und skalierbaren Infrastruktur für die Bereitstellung von Diensten im Umfeld von Bibliotheken und ähnlichen wissensanbietenden Einrichtungen unter Verwendung von XML-RPC und Python.

  1. Results from the National Legionella Outbreak Detection Program, the Netherlands, 2002-2012.

    Science.gov (United States)

    Den Boer, Jeroen W; Euser, Sjoerd M; Brandsema, Petra; Reijnen, Linda; Bruin, Jacob P

    2015-07-01

    In 2002, the National Legionella Outbreak Detection Program was implemented in the Netherlands to detect and eliminate potential sources of organisms that cause Legionnaires' disease (LD). During 2002-2012, a total of 1,991 patients with LD were reported, and 1,484 source investigations were performed. Of those sources investigated, 24.7% were positive for Legionella spp. For 266 patients with LD, 105 cluster locations were identified. A genotype match was made between a strain detected in 41 patients and a strain from a source location. Despite the systematic approach used by the program, most sources of LD infections during 2002-2012 remained undiscovered. Explorative studies are needed to identify yet undiscovered reservoirs and transmission routes for Legionella bacteria, and improved laboratory techniques are needed to detect Legionella spp. in clinical samples with a high background of microbial flora (such as soil).

  2. Cloning and expression of the Legionella micdadei "common antigen" in Escherichia coli

    DEFF Research Database (Denmark)

    Bangsborg, Jette Marie; Collins, M T; Høiby, N

    1989-01-01

    To study individual Legionella antigens, a Legionella micdadei genomic library in Escherichia coli SC181 was established. Partially Sau3A digested L. micdadei DNA fragments (15-25 kilobase pairs (kb] were cloned into the tetracycline resistance gene of the cosmid vector pHC79. Four thousand...... ampicillin resistant recombinants were obtained; seven hundred were screened for expression of Legionella antigens in Western blot analysis with a polyspecific E. coli-absorbed anti-L. micdadei rabbit antibody. One of the positive clones expressed a 60 kilodalton (K) antigen, which reacted strongly...... will provide important information with respect to genetic vs. antigenic relatedness among Legionellae and other Gram-negative species, as well as to CA structure and possible function....

  3. Longitudinale studie naar de aanwezigheid van legionella en amoeben in drinkwaterinstallaties

    NARCIS (Netherlands)

    Schalk JAC; Redeker S; Docters van Leeuwen AE; Lodder WJ; de Roda Husman AM; LZO; cib

    2012-01-01

    In Nederland wordt bij prioritaire instellingen met collectieve leidingwaterinstallaties, zoals ziekenhuizen en verzorgingstehuizen, twee keer per jaar gecontroleerd of legionella in de waterleidingen aanwezig is. De concentratie legionellabacteriën in het water moet wettelijk lager zijn dan 100

  4. Impact of a risk management plan on Legionella contamination of dental unit water.

    Science.gov (United States)

    Leoni, Erica; Dallolio, Laura; Stagni, Francesca; Sanna, Tiziana; D'Alessandro, Giovanni; Piana, Gabriela

    2015-02-23

    The study aimed to assess the prevalence of Legionella spp. in dental unit waterlines of a dental clinic and to verify whether the microbiological parameters used as indicators of water quality were correlated with Legionella contamination. A risk management plan was subsequently implemented in the dental health care setting, in order to verify whether the adopted disinfection protocols were effective in preventing Legionella colonization. The water delivered from syringes and turbines of 63 dental units operating in a dental clinic, was monitored for counts of the heterotrophic bacteria P. aeruginosa and Legionella spp. (22 °C and 37 °C). At baseline, output water from dental units continuously treated with disinfection products was more compliant with the recommended standards than untreated and periodically treated water. However, continuous disinfection was still not able to prevent contamination by Legionella and P. aeruginosa. Legionella was isolated from 36.4%, 24.3% and 53.3% of samples from untreated, periodically and continuously treated waterlines, respectively. The standard microbiological parameters used as indicators of water quality proved to be unreliable as predictors of the presence of Legionella, whose source was identified as the tap water used to supply the dental units. The adoption of control measures, including the use of deionized water in supplying the dental unit waterlines and the application of a combined protocol of continuous and periodic disinfection, with different active products for the different devices, resulted in good control of Legionella contamination. The efficacy of the measures adopted was mainly linked to the strict adherence to the planned protocols, which placed particular stress on staff training and ongoing environmental monitoring.

  5. Identification of Legionella in the Hot Water Supply of a General Hospital in Isfahan

    OpenAIRE

    H Movahedian Attar; M R Shahmansouri; A A Neshat; M Fazeli

    2004-01-01

    Background: Legionella is a gram negative, aerobic, and sporeless bacterium which is readily found in ventilation systems, cooling towers, hot water distribution systems, bathrooms, swimming pools, and fountains. Legionella is implicated in the legionnaires’ and Pontiac fever diseases. Hospitals are common habitats for the bacterium, where the bacterial growths are amply found and that provide the most likely places for susceptible people to contract the diseases. Given the importance of hosp...

  6. Cerebellar involvement that occurred during treatment of Legionella pneumonia: A case report

    Directory of Open Access Journals (Sweden)

    Ozlem Alici

    2013-06-01

    Full Text Available Legionnaires’ disease can appear with different levels of severity. A case of a previously healthy lady with communityacquiredpneumonia who progressed to severe acute respiratory distress syndrome and developed cerebellar dysfunctionis reported. In patients presenting with neurological symptoms after an episode of pneumonia, Legionella infectionshould be considered. J Microbiol Infect Dis 2013; 3(2: 83-85Key words: Legionella, cerebellar dysfunction, dysarthria, ataxia

  7. Legionella Risk Management and Control in Potable Water Systems: Argument for the Abolishment of Routine Testing.

    Science.gov (United States)

    Whiley, Harriet

    2016-12-24

    Legionella is an opportunistic pathogen of public health significance. One of the main sources of Legionella is potable water systems. As a consequence of aging populations there is an increasing demographic considered at high risk for Legionellosis and, as such, a review of the guidelines is required. Worldwide, Legionella has been detected from many potable water sources, suggesting it is ubiquitous in this environment. Previous studies have identified the limitations of the current standard method for Legionella detection and the high possibility of it returning both false negative and false positive results. There is also huge variability in Legionella test results for the same water sample when conducted at different laboratories. However, many guidelines still recommend the testing of water systems. This commentary argues for the removal of routine Legionella monitoring from all water distribution guidelines. This procedure is financially consuming and false negatives may result in managers being over-confident with a system or a control mechanism. Instead, the presence of the pathogen should be assumed and focus spent on managing appropriate control measures and protecting high-risk population groups.

  8. Legionella detection by culture and qPCR: Comparing apples and oranges.

    Science.gov (United States)

    Whiley, Harriet; Taylor, Michael

    2016-01-01

    Legionella spp. are the causative agent of Legionnaire's disease and an opportunistic pathogen of significant public health concern. Identification and quantification from environmental sources is crucial for identifying outbreak origins and providing sufficient information for risk assessment and disease prevention. Currently there are a range of methods for Legionella spp. quantification from environmental sources, but the two most widely used and accepted are culture and real-time polymerase chain reaction (qPCR). This paper provides a review of these two methods and outlines their advantages and limitations. Studies from the last 10 years which have concurrently used culture and qPCR to quantify Legionella spp. from environmental sources have been compiled. 26/28 studies detected Legionella at a higher rate using qPCR compared to culture, whilst only one study detected equivalent levels of Legionella spp. using both qPCR and culture. Aggregating the environmental samples from all 28 studies, 2856/3967 (72%) tested positive for the presence of Legionella spp. using qPCR and 1331/3967 (34%) using culture. The lack of correlation between methods highlights the need to develop an acceptable standardized method for quantification that is sufficient for risk assessment and management of this human pathogen.

  9. Occurrence of Legionella in wastewater treatment plants linked to wastewater characteristics.

    Science.gov (United States)

    Caicedo, C; Beutel, S; Scheper, T; Rosenwinkel, K H; Nogueira, R

    2016-08-01

    In recent years, the occurrence of Legionella in wastewater treatment plants (WWTP) has often been reported. However, until now there is limited knowledge about the factors that promote Legionella's growth in such systems. The aim of this study was to investigate the chemical wastewater parameters that might be correlated to the concentration of Legionella spp. in WWTP receiving industrial effluents. For this purpose, samples were collected at different processes in three WWTP. In 100 % of the samples taken from the activated sludge tanks Legionella spp. were detected at varying concentrations (4.8 to 5.6 log GU/mL) by the quantitative real-time polymerase chain reaction method, but not by the culture method. Statistical analysis with various parameters yielded positive correlations of Legionella spp. concentration with particulate chemical oxygen demand, Kjeldahl nitrogen and protein concentration. Amino acids were quantified in wastewater and activated sludge samples at concentrations that may not support the growth of Legionella, suggesting that in activated sludge tanks this bacterium multiplied in protozoan hosts.

  10. Identification of Legionella from clinically diagnosed pneumonia patients and environmental samples.

    Science.gov (United States)

    Jahan, R; Tarafder, S; Saleh, A A; Miah, M R A

    2015-04-01

    Legionnaires' disease is a multisystem disease with life-threatening acute and severe form of pneumonia which is responsible for 2-9% pneumonia with high mortality. Eighty six respiratory tract samples and urine were collected from clinically diagnosed pneumonia patients and 12 water samples were collected from different environment. Identification of Legionella was done by culture and Polymerase Chain Reaction (PCR) of respiratory tract samples and environmental samples and Legionella Antigen (Ag) in urine was detected by Immunochromatographic test (ICT). Legionella was identified from 4 (4.65%) clinically diagnosed pneumonia patients of which 1(1.16%) case was culture positive, 1(1.16%) case was urine ICT positive and PCR was positive in all four cases. Of the 12 water samples tested, 4 (33.33%) samples were Legionella positive by PCR but culture results of these samples were negative. Identification of Legionella should be done by PCR in parallel with culture and urine ICT. Detection of Legionella in environmental samples is also needed to explore possible links between the water sources and disease transmission in population.

  11. Incidence of Legionella and heterotrophic bacteria in household rainwater tanks in Azumino, Nagano prefecture, Japan.

    Science.gov (United States)

    Kobayashi, Michiko; Oana, Kozue; Kawakami, Yoshiyuki

    2014-01-01

    Many administrative agencies in Japan are encouraging installation of household rainwater-storage tanks for more effective use of natural rainwater. Water samples were collected periodically from 43 rainwater tanks from 40 households and tested for the presence of Legionella species and the extent of heterotrophic bacteria in Azumino city, Nagano prefecture, Japan. PCR assays indicated the presence of Legionella spp. in 12 (30%) of the 43 tank water samples. Attempts were made to identify correlations between PCR positive samples, topography, pH, chemical oxygen demand (COD), atmospheric temperature and the numbers of heterotrophic bacteria. Between June and October, 2012, the numbers of heterotrophic bacteria in rainwater tanks and the values of COD positively correlated with the presence of Legionella species. In most of the Legionella-positive cases, heterotrophic bacterial cell counts were >10(4) CFU/mL. Moreover, Legionella species were less frequently detected when the COD value was >5 mg KMnO(4)/L. Therefore, at least in Azumino, Japan between June and October 2012, both heterotrophic bacterial counts and COD values may be considered index parameters for the presence of Legionella cells in rainwater tanks. Much more accumulation of such data is needed to verify the accuracy of these findings. © 2013 The Societies and Wiley Publishing Asia Pty Ltd.

  12. Frequency of legionella contamination in conditional & water distribution systems of Tehran hospitals

    Directory of Open Access Journals (Sweden)

    Davod Esmaieli

    2008-09-01

    Full Text Available Background: Legionella species are ubiquitous in natural aquatic environments, capable of existing in waters with varied temperatures, PH levels, and nutrient and oxygen contents. Of 49 known legionella species, 20 species have been linked to pneumonia in humans. Contamination by legionella has occurred in the distribution systems of many hospitals. Aerosol-generating systems such as faucets, showerheads, cooling towers, and nebulizers are responsible for their transmission from water to air. Methods: A total of 113 water samples were gathered from different wards of 32 hospitals in different geographical regions of Tehran city. These samples were concentrated by filtration, treated with the acid and temperature buffers, and isolated on a BCYE agar culture medium. Results: A total of 22 hospitals out of 33 (26.5% were contaminated by legionella species, and 30 samples (26.5% out of 113 were positive. Chlorine concentration and pH level of the water samples were 0.18-2.2 mg/l and 6.6-7.6, respectively. Conclusion: The high rate of waste water contamination in Tehran hospitals with Legionella indicates the resistance of this microorganism to chlorine and other disinfectants, or inadequate disinfection process, representing the insufficiency of the current decontamination of hospital water distribution system. Thus identifying legionella species and their controlling in water distribution system of hospitals is of great importance.

  13. Molecular characterization of viable Legionella spp. in cooling tower water samples by combined use of ethidium monoazide and PCR.

    Science.gov (United States)

    Inoue, Hiroaki; Fujimura, Reiko; Agata, Kunio; Ohta, Hiroyuki

    2015-01-01

    Viable Legionella spp. in environmental water samples were characterized phylogenetically by a clone library analysis combining the use of ethidium monoazide and quantitative PCR. To examine the diversity of Legionella spp., six cooling tower water samples and three bath water samples were collected and analyzed. A total of 617 clones were analyzed for their 16S rRNA gene sequences and classified into 99 operational taxonomic units (OTUs). The majority of OTUs were not clustered with currently described Legionella spp., suggesting the wide diversity of not-yet-cultured Legionella groups harbored in cooling tower water environments.

  14. Insufficient knowledge on legionella control; Kennis legionellabeheersing ontbreekt nogal eens

    Energy Technology Data Exchange (ETDEWEB)

    Kordes, B. [Kordes Advies, Wijhe (Netherlands); Nuijten, O. [Isso, Rotterdam (Netherlands)

    2011-05-15

    After the lethal incident in the cooling tower of Amsterdam in 2006, legislation has been adapted. Parties that operate a wet cooling tower need to take precautions to prevent risks for the surroundings and to prevent unusual events or limit these to the best possible extent. Moreover, compliance with the Activities Decree (article 3.16a on legionella prevention in the Environmental legislation) is required. [Dutch] Na het incident met dodelijke afloop in de koeltoren in Amsterdam in 2006 is de wetgeving aangepast. Degene die een natte koeltoren beheert moet risico's voor de omgeving en ongewone voorvallen voorkomen of deze zoveel mogelijk beperken. Ook moet ten minste aan Activiteitenbesluit (artikel 3.16a over legionellapreventie in de Milieuwetgeving) worden voldaan.

  15. John von Neumann Birthday Centennial

    Energy Technology Data Exchange (ETDEWEB)

    Grcar, Joseph F.

    2004-11-12

    In celebration of John von Neumann's 100th birthday, a series of four lectures were presented on the evening of February 10, 2003 during the SIAM Conference on Computational Science and Engineering in San Diego. The venue was appropriate because von Neumann spent much of the later part of his life, in the 1950's, as an unofficial ambassador for computational science. He was then the only senior American scientist who had experience with the new computers (digital, electronic, and programmable) and a vision of their future importance. No doubt he would have relished the chance to attend a meeting such as this. The first speaker, William Aspray, described the ''interesting times'' during which computers were invented. His remarks were based on his history [1] of this period in von Neumann's life. We were honored to have John von Neumann's daughter, Marina von Neumann-Whitman, as our second speaker. Other accounts of von Neumann's life can be found in books by two of his colleagues [2] and [3]. Our third speaker, Peter Lax, provided both mathematical and international perspectives on John von Neumann's career. Finally, Pete Stewart spoke about von Neumann's numerical error analysis [4] in the context of later work; this talk did not lend itself to transcription, but readers may consult the historical notes in [5]. Our thanks to all the speakers for a remarkable evening. We are grateful to the DOE Applied Mathematical Sciences (AMS) program for partially supporting these lectures. Thanks are also due to SIAM and William Kolata, to our emcee, Gene Golub, to Paul Saylor for recording and editing, and to Barbara Lytle for the transcriptions. More about von Neumann's work can be learned from the recent American Mathematical Society proceedings [6].

  16. Alejandro Von Humboldt

    Directory of Open Access Journals (Sweden)

    Gerardo Paz Otero

    1965-09-01

    Full Text Available Diverso fue y sigue siéndolo, el destino de los dos hermanos Humboldt, Guillermo y Alejandro. Sino que se inicia con el nacimiento: Guillermo, el mayor, nace el 22 de junio de 1767 en Potsdam, residencia de los emperadores prusianos, la ciudad de los palacios imperiales, el imperio del militarismo germano; su cuna se meció cerca al palacete de Sans-Soussi, donde Federico el Grande forja ba el poderío de Prusia, cultivaba las ciencias y las artes, anfitrionaba a los intelectuales de Europa, y era "vasallo espiritual de Volta ire", según la aguda frase de Goethe. Alejandro viene al mundo dos años después (14 de septiembre de 1769 en Berlín, en la casa burguesa de la Jagerstrasse (calle del cazador que su madre Elizabeth von Humboldt heredara de su primer esposo.

  17. Der Einfluss von Phlorizin auf den programmierten Zelltod von Erythrozyten

    OpenAIRE

    Meier, Anja Doris

    2013-01-01

    Erythrozyten sind in der Lage, suizidalen Zelltod zu betreiben. Von einigen Substanzen ist bereits bekannt, dass sie die sogenannte Eryptose beeinflussen. Beispiele dafür sind Quecksilber, Monensin, Benzethonium und Oridonin, welche zu den Stimulatoren der Eryptose gehören. Blebbistatin, Koffein und Endothelin, gehören zu den Inhibitoren. In dieser Studie sollte untersucht werden, welchen Einfluss Phlorizin auf den Ablauf des suizidalen Zelltodes von Erythrozyten hat. Es wurde belegt, dass si...

  18. Fast label-free detection of Legionella spp. in biofilms by applying immunomagnetic beads and Raman spectroscopy.

    Science.gov (United States)

    Kusić, Dragana; Rösch, Petra; Popp, Jürgen

    2016-03-01

    Legionellae colonize biofilms, can form a biofilm by itself and multiply intracellularly within the protozoa commonly found in water distribution systems. Approximately half of the known species are pathogenic and have been connected to severe multisystem Legionnaires' disease. The detection methods for Legionella spp. in water samples are still based on cultivation, which is time consuming due to the slow growth of this bacterium. Here, we developed a cultivation-independent, label-free and fast detection method for legionellae in a biofilm matrix based on the Raman spectroscopic analysis of isolated single cells via immunomagnetic separation (IMS). A database comprising the Raman spectra of single bacterial cells captured and separated from the biofilms formed by each species was used to build the identification method based on a support vector machine (SVM) discriminative classifier. The complete method allows the detection of Legionella spp. in 100 min. Cross-reactivity of Legionella spp. specific immunomagnetic beads to the other studied genera was tested, where only small cell amounts of Pseudomonas aeruginosa, Klebsiella pneumoniae and Escherichia coli compared to the initial number of cells were isolated by the immunobeads. Nevertheless, the Raman spectra collected from isolated non-targeted bacteria were well-discriminated from the Raman spectra collected from isolated Legionella cells, whereby the Raman spectra of the independent dataset of Legionella strains were assigned with an accuracy of 98.6%. In addition, Raman spectroscopy was also used to differentiate between isolated Legionella species. Copyright © 2016 Elsevier GmbH. All rights reserved.

  19. Evaluation of a new monochloramine generation system for controlling Legionella in building hot water systems.

    Science.gov (United States)

    Duda, Scott; Kandiah, Sheena; Stout, Janet E; Baron, Julianne L; Yassin, Mohamed; Fabrizio, Marie; Ferrelli, Juliet; Hariri, Rahman; Wagener, Marilyn M; Goepfert, John; Bond, James; Hannigan, Joseph; Rogers, Denzil

    2014-11-01

    To evaluate the efficacy of a new monochloramine generation system for control of Legionella in a hospital hot water distribution system. A 495-bed tertiary care hospital in Pittsburgh, Pennsylvania. The hospital has 12 floors covering approximately 78,000 m(2). The hospital hot water system was monitored for a total of 29 months, including a 5-month baseline sampling period prior to installation of the monochloramine system and 24 months of surveillance after system installation (postdisinfection period). Water samples were collected for microbiological analysis (Legionella species, Pseudomonas aeruginosa, Stenotrophomonas maltophilia, Acinetobacter species, nitrifying bacteria, heterotrophic plate count [HPC] bacteria, and nontuberculous mycobacteria). Chemical parameters monitored during the investigation included monochloramine, chlorine (free and total), nitrate, nitrite, total ammonia, copper, silver, lead, and pH. A significant reduction in Legionella distal site positivity was observed between the pre- and postdisinfection periods, with positivity decreasing from an average of 53% (baseline) to an average of 9% after monochloramine application (Pevaluation in the United States of a commercially available monochloramine system installed on a hospital hot water system for Legionella disinfection, and it demonstrated a significant reduction in Legionella colonization. Significant increases in microbial populations or other negative effects previously associated with monochloramine use in large municipal cold water systems were not observed.

  20. Ten Questions Concerning the Aerosolization and Transmission of Legionella in the Built Environment.

    Science.gov (United States)

    Prussin, Aaron J; Schwake, David Otto; Marr, Linsey C

    2017-10-01

    Legionella is a genus of pathogenic Gram-negative bacteria responsible for a serious disease known as legionellosis, which is transmitted via inhalation of this pathogen in aerosol form. There are two forms of legionellosis: Legionnaires' disease, which causes pneumonia-like symptoms, and Pontiac fever, which causes influenza-like symptoms. Legionella can be aerosolized from various water sources in the built environment including showers, faucets, hot tubs/swimming pools, cooling towers, and fountains. Incidence of the disease is higher in the summertime, possibly because of increased use of cooling towers for air conditioning systems and differences in water chemistry when outdoor temperatures are higher. Although there have been decades of research related to Legionella transmission, many knowledge gaps remain. While conventional wisdom suggests that showering is an important source of exposure in buildings, existing measurements do not provide strong support for this idea. There has been limited research on the potential for Legionella transmission through heating, ventilation, and air conditioning (HVAC) systems. Epidemiological data suggest a large proportion of legionellosis cases go unreported, as most people who are infected do not seek medical attention. Additionally, controlled laboratory studies examining water-to-air transfer and source tracking are still needed. Herein, we discuss ten questions that spotlight current knowledge about Legionella transmission in the built environment, engineering controls that might prevent future disease outbreaks, and future research that is needed to advance understanding of transmission and control of legionellosis.

  1. [Legionella spp. contamination in indoor air: preliminary results of an Italian multicenter study].

    Science.gov (United States)

    Montagna, Maria Teresa; De Giglio, Osvalda; Napoli, Christian; Cannova, Lucia; Cristina, Maria Luisa; Deriu, Maria Grazia; Delia, Santi Antonino; Giuliano, Ada; Guida, Marco; Laganà, Pasqualina; Liguori, Giorgio; Mura, Ida; Pennino, Francesca; Rossini, Angelo; Tardivo, Stefano; Torre, Ida; Torregrossa, Maria Valeria; Villafrate, Maria Rosaria; Albertini, Roberto; Pasquarella, Cesira

    2014-01-01

    To propose a standardized protocol for the evaluation of Legionella contamination in air. A bathroom having a Legionella contamination in water >1,000 cfu/l was selected in 10 different healthcare facilities. Air contamination was assessed by active (Surface Air System, SAS) and passive (Index of Microbial Air, IMA) sampling for 8 hours, about 1 m away from the floor and 50 cm from the tap water. Two hundred liters of air were sampled by SAS every 12 min, after flushing water for 2 min. The IMA value was calculated as the mean value of colony forming units/16 plates exposed during sampling (2 plates/hour). Water contamination was evaluated at T0, after 4 and 8 hours, according to the standard methods. Air contamination by Legionella was found in three healthcare facilities (one with active and two with passive sampling), showing a concomitant tap water contamination (median=40,000; range 1,100-43,000 cfu/l). The remaining seven hospitals isolated Legionella spp. exclusively from water samples (median=8,000; range 1,200-70,000 cfu/l). Our data suggest that environmental Legionella contamination cannot be assessed only through the air sampling, even in the presence of an important water contamination.

  2. Effects of holding time and measurement error on culturing Legionella in environmental water samples.

    Science.gov (United States)

    Flanders, W Dana; Kirkland, Kimberly H; Shelton, Brian G

    2014-10-01

    Outbreaks of Legionnaires' disease require environmental testing of water samples from potentially implicated building water systems to identify the source of exposure. A previous study reports a large impact on Legionella sample results due to shipping and delays in sample processing. Specifically, this same study, without accounting for measurement error, reports more than half of shipped samples tested had Legionella levels that arbitrarily changed up or down by one or more logs, and the authors attribute this result to shipping time. Accordingly, we conducted a study to determine the effects of sample holding/shipping time on Legionella sample results while taking into account measurement error, which has previously not been addressed. We analyzed 159 samples, each split into 16 aliquots, of which one-half (8) were processed promptly after collection. The remaining half (8) were processed the following day to assess impact of holding/shipping time. A total of 2544 samples were analyzed including replicates. After accounting for inherent measurement error, we found that the effect of holding time on observed Legionella counts was small and should have no practical impact on interpretation of results. Holding samples increased the root mean squared error by only about 3-8%. Notably, for only one of 159 samples, did the average of the 8 replicate counts change by 1 log. Thus, our findings do not support the hypothesis of frequent, significant (≥= 1 log10 unit) Legionella colony count changes due to holding. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

  3. The importance of different laboratory methods in Legionella diagnosis in medical care.

    Science.gov (United States)

    Mathys, W; Junge, E; Sobek-Pfeiffer, C; Bösenberg, H; von Eiff, M

    1988-11-01

    Legionellae an infection by contaminated water is thought to be respo environment of patients with a high risk of infection (e.g. after ki lantation, immunosuppression) prophylactic measures (rising the tempe warm water to 60 degrees C) should be combined with bacteriological c s to be taken into account, however, that only by taking a couple of he same outlet during a period of time a colonization of central inst ms (sediments, storage tanks) can be discovered. In patients with acu a of unknown origin serological tests of Legionella antibodies are no c. Antibody titers of not infected and with Legionella infected patie gnificant difference. Well suited for the diagnosis of a legionellosi f the Direct Immunofluorescent Antibody Assay (DFA) in invasively sam . In immunocompromised pneumonia patients 18.3% of bronchoalveolar 6.0% of bronchoalveolar aspirates are Legionella positive. RETURN/proces t F3/ext F4/can F5/nxt F6/ins F7/up F8/dwn F9/fin lavages and 16.0% of bronchoalveolar aspirates are Legionella positive.

  4. Einsatz von Nickel-Katalysatoren zum Cracken von Teerprodukten aus der Niederdruckaufkohlung von Stahl mit Ethin

    OpenAIRE

    Mbadinga Mouanda, Gelase

    2009-01-01

    Die Niederdruckaufkohlung von Stahl ist ein modernes Verfahren, das zur Einsatzhärtung von Stahlbauteilen dient. Bei Temperaturen zwischen 900 und 1050 °C und Drücken unter 50 mbar werden Bauteile durch heterogene Pyrolyse von Ethin in einem Ofen einsatzgehärtet. Das während der Pyrolyse erzeugte Abgas enthält aromatische Kohlenwasserstoffe und kann katalytisch behandelt werden. Der Katalysator wird durch Rußablagerung desaktiviert und muss zyklisch durch Verbrennung mit Luft regeneriert werd...

  5. Living with von Willebrand Disease

    Science.gov (United States)

    ... the condition. For example, the school nurse, teacher, daycare provider, coach, or any leader of afterschool activities ... MedlinePlus) Von Willebrand Disease (MedlinePlus) Building 31 31 Center Drive Bethesda, MD 20892 Learn more about getting ...

  6. Ueber einige Coelenterata von Australien

    NARCIS (Netherlands)

    Stiasny, G.

    1931-01-01

    I. Hydromedusen. *Olindias singularis Browne. II. Scyphomedusen. *Atolla wyvillei Haeckel. *Linuche unguiculata O. Swartz. Pelagia noctiluca Péron u. Lesueur. Cyanea capillata var. annaskala von Lendenfeld. Netrostoma coerulescens Maas. *Versura anadyomene (Maas) Mayer. Catostylus mosaicus L.

  7. Kleben von Kunststoff mit Metall

    CERN Document Server

    Brockmann, W; Käufer, H

    1989-01-01

    Das Buch behandelt das Kleben von Kunststoffen mit Metallen in einer fur den Praktiker verstandlichen und umsetzbaren Form. Es leitet zu Klebeverfahren an, die optimale Ergebnisse hinsichtlich Qualitat, Dauerhaftigkeit und Wirtschaftlichkeit liefern.

  8. Detection of Legionella species in environmental water by the quantitative PCR method in combination with ethidium monoazide treatment.

    Science.gov (United States)

    Inoue, Hiroaki; Takama, Tomoko; Yoshizaki, Miwa; Agata, Kunio

    2015-01-01

    We detected Legionella species in 111 bath water samples and 95 cooling tower water samples by using a combination of conventional plate culture, quantitative polymerase chain reaction (qPCR) and qPCR combined with ethidium monoazide treatment (EMA-qPCR) methods. In the case of bath water samples, Legionella spp. were detected in 30 samples by plate culture, in 85 samples by qPCR, and in 49 samples by EMA-qPCR. Of 81 samples determined to be Legionella-negative by plate culture, 56 and 23 samples were positive by qPCR and EMA-qPCR, respectively. Therefore, EMA treatment decreased the number of Legionella-positive bath water samples detected by qPCR. In contrast, EMA treatment had no effect on cooling tower water samples. We therefore expect that EMA-qPCR is a useful method for the rapid detection of viable Legionella spp. from bath water samples.

  9. Fluorido-Komplexe von Technetium

    OpenAIRE

    Mariappan Balasekaran, Samundeeswari

    2013-01-01

    Zusammenfassung Diese Dissertationsschrift befasst sich mit der Synthese und Charakterisierung neuer Technetiumfluoride mit dem Metall in den Oxidationsstufen “+1”, “+2”, “+4” und “+6”. Im ersten Kapitel wird über die Isolierung von unterschiedlichen Salzen von Fluoridonitridotechnetaten(VI) entweder aus Nitridotechnetium(VI)-säure oder aus Pertechnetat durch den Einsatz geeigneter Reduktionsmittel berichtet. Das Cäsiumsalz dieser Verbindung bildet einen oxido-verbrückten, dimeren Kompl...

  10. Effiziente chemoenzymatische Synthese von dhydroartemisinaldehyd

    OpenAIRE

    Demiray, Melodi; Tang, Xiaoping; Wirth, Thomas; Faraldos, Juan A.; Allemann, Rudolf K.

    2017-01-01

    Artemisinin aus der Pflanze Artemisia annua ist das wirkungsvollste Arzneimittel zur Behandlung von Malaria. Die Sesquiterpen-Cyclase Amorphadien-Synthase, ein Cytochrom-abhängiges CYP450 und eine Aldehyd-Reduktase wandeln in der Pflanze Farnesyl-Diphosphat (FDP) in Dihydroartemisinaldehyd (DHAAl) um, welches ein Schlüsselzwischenprodukt in der Biosynthese von Artemisinin und eine halbsynthetische Vorstufe in der chemischen Synthese des Arzneimittels ist. Hier berichten wir über einen chemoen...

  11. Review of Various Solutions for avoiding critical levels of Legionella Bacteria in Domestic Hot Water System

    DEFF Research Database (Denmark)

    Yang, Xiaochen; Li, Hongwei; Svendsen, Svend

    2013-01-01

    Low temperature district heating (DH) is designed as 55/25oC for supply/return temperature to fulfill the low energy demand of future buildings. However, to secure the safety of domestic hot water, the supply temperature has to be kept around 60oC to avoid the existence of legionella, which...... reproduces rapidly at the temperature around 25oC- 45 oC. After several outbreaks of pheumonia and fever caused by legionella bacteria, most countries require 60 oC in the network and 50-55 oC at the faucets with periodic flush by hot water above 60 oC as disinfection solution. That makes obstacles of low...... temperature DH implementation. Therefore, effective solution of legionella bacteria is in urgent demand. To select optimal disinfection treatments for certain cases which are quite different in dimension or purpose of use, various methods were reviewed, including shock hyperchlorination, super heating...

  12. Colonization by Legionella spp. of water networks in residential buildings of the Province of Pisa, Italy.

    Science.gov (United States)

    Baggiani, A; Casini, B; Totaro, M; Aquino, F; Valentini, P; Bruni, B; Porretta, A; Casalini, F; Miccoli, M; Privitera, G

    2015-01-01

    Despite the increase of community acquired cases of legionellosis in Italy over the last years, the Italian guidelines do not give indications for prevention and control of Legionella in the hot water networks (or centralized conditioning systems) of residential buildings. We performed a survey on eight medium sized apartment buildings in the Pisa district to assess the prevalence of Legionella spp. in the water network and the respondance to drinking water requisites at the point of use, according to the Italian norms. For each building two hot water and three cold water samples (located at water entrance from the aqueduct network into the building pipework, at the exit from pressure autoclave, and at a remote tap) were collected. Legionella was detected in 20% of residential buildings, mostly in those with a central hot water production system. The study highlights a condition of potential risk for susceptible population subgroups and supports the need for measures of risk assessment and control.

  13. Legionella thermalis sp. nov., isolated from hot spring water in Tokyo, Japan.

    Science.gov (United States)

    Ishizaki, Naoto; Sogawa, Kazuyuki; Inoue, Hiroaki; Agata, Kunio; Edagawa, Akiko; Miyamoto, Hiroshi; Fukuyama, Masafumi; Furuhata, Katsunori

    2016-03-01

    Strain L-47(T) of a novel bacterial species belonging to the genus Legionella was isolated from a sample of hot spring water from Tokyo, Japan. The 16S rRNA gene sequences (1477 bp) of this strain (accession number AB899895) had less than 95.0% identity with other Legionella species. The dominant fatty acids of strain L-47(T) were a15:0 (29.6%) and the major ubiquinone was Q-12 (71.1%). It had a guanine-plus-cytosine content of 41.5 mol%. The taxonomic description of Legionella thermalis sp. nov. is proposed to be type strain L-47(T) (JCM 30970(T)  = KCTC 42799(T)). © 2016 The Societies and John Wiley & Sons Australia, Ltd.

  14. Viability-qPCR for detecting Legionella: Comparison of two assays based on different amplicon lengths.

    Science.gov (United States)

    Ditommaso, Savina; Giacomuzzi, Monica; Ricciardi, Elisa; Zotti, Carla M

    2015-08-01

    Two different real-time quantitative PCR (PMA-qPCR) assays were applied for quantification of Legionella spp. by targeting a long amplicon (approx 400 bp) of 16S rRNA gene and a short amplicon (approx. 100 bp) of 5S rRNA gene. Purified DNA extracts from pure cultures of Legionella spp. and from environmental water samples were quantified. Application of the two assays to quantify Legionella in artificially contaminated water achieved that both assays were able to detect Legionella over a linear range of 10 to 10(5) cells ml(-1). A statistical analysis of the standard curves showed that both assays were linear with a good correlation coefficient (R(2) = 0.99) between the Ct and the copy number. Amplification with the reference assay was the most effective for detecting low copy numbers (1 bacterium per PCR mixture). Using selective quantification of viable Legionella by the PMA-qPCR method we obtained a greater inhibition of the amplification of the 400-bp 16S gene fragment (Δlog(10) = 3.74 ± 0.39 log(10) GU ml(-1)). A complete inhibition of the PCR signal was obtained when heat-killed cells in a concentration below 1 × 10(5) cells ml(-1) were pretreated with PMA. Analysing short amplicon sizes led to only 2.08 log reductions in the Legionella dead-cell signal. When we tested environmental water samples, the two qPCR assays were in good agreement according to the kappa index (0.741). Applying qPCR combined with PMA treatment, we also obtained a good agreement (kappa index 0.615). The comparison of quantitative results shows that both assays yielded the same quantification sensitivity (mean log = 4.59 vs mean log = 4.31). Copyright © 2015 Elsevier Ltd. All rights reserved.

  15. Legionnaire's disease kills in Norway; 5 doede i legionella epidemi

    Energy Technology Data Exchange (ETDEWEB)

    Kristensen, Tore

    2001-07-01

    In Norway in 2001, five people died of legionnaires' disease. A water fountain in a lake in the centre of the city of Stavanger was suspected of being the source of infection. New instances of the disease were reported some time after the fountain was closed down, however, and after renewed search legionella bacteria were discovered in the cooling tower of one of the hotels of the city. The legionella bacteria is normally found in water and moist soil but in insignificant numbers. The article discusses what happened in Norway and gives some characteristics of the bacteria and the disease.

  16. Detection of Legionella bozemanae, a New Cause of Septic Arthritis, by PCR Followed by Specific Culture

    DEFF Research Database (Denmark)

    Just, Søren Andreas; Knudsen, John Bonde; Uldum, Søren Anker

    2012-01-01

    Legionella bozemanae is a rare isolate in clinical specimens. We describe a case of joint infection due to L. bozemanae in an immunocompromised patient with dermatomyositis. Without the use of PCR screening or culture on specialized medium, the organism would not have been detected.......Legionella bozemanae is a rare isolate in clinical specimens. We describe a case of joint infection due to L. bozemanae in an immunocompromised patient with dermatomyositis. Without the use of PCR screening or culture on specialized medium, the organism would not have been detected....

  17. Identification of Legionella in the Hot Water Supply of a General Hospital in Isfahan

    Directory of Open Access Journals (Sweden)

    H Movahedian Attar

    2004-12-01

    Full Text Available Background: Legionella is a gram negative, aerobic, and sporeless bacterium which is readily found in ventilation systems, cooling towers, hot water distribution systems, bathrooms, swimming pools, and fountains. Legionella is implicated in the legionnaires’ and Pontiac fever diseases. Hospitals are common habitats for the bacterium, where the bacterial growths are amply found and that provide the most likely places for susceptible people to contract the diseases. Given the importance of hospitals in this regard, this survey was carried out in a General Hospital in Isfahan. Methods: For the purposes of this study, a total of 30 samples