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Sample records for genotype diversity detected

  1. Nested PCR for detection and genotyping of Ehrlichia ruminantium: use in genetic diversity analysis.

    Science.gov (United States)

    Martinez, Dominique; Vachiéry, Nathalie; Stachurski, Frederic; Kandassamy, Yane; Raliniaina, Modestine; Aprelon, Rosalie; Gueye, Arona

    2004-10-01

    Ehrlichia ruminantium, the agent of cowdriosis transmitted by Amblyomma ticks, presents an extensive genetic and antigenic diversity of key importance for vaccine formulation. Two means of nested polymerase chain reaction (PCR) targeting were developed to conduct molecular epidemiology studies in the Caribbean and Africa. The first used a conserved DNA fragment for detection of the pathogen in animals and vectors, and the second relied on the polymorphic map1 gene for genotyping. As compared to a PCR, the nested PCR showed a 2-Log10 improvement of sensitivity and allowed amplification from ticks, blood, brain, and lungs from infected animals, providing a more accurate picture of the tick infection rate. In Guadeloupe, this rate reached 36% (N = 212) instead of 1.7% (N = 224), as previously estimated. Genetic typing was done by restriction fragment length polymorphism or sequencing of map1 amplification products. Molecular epidemiology studies conducted in field sites selected for vaccination trials with inactivated vaccine, revealed the circulation of genetically divergent strains in limited geographical areas. It is known, then, that genetic clustering based on map1 has no predictive value regarding the protective value of a given strain against a new strain. However, tracing the strains by this technique revealed the extent of E. ruminantium diversity that one can expect in a given region, and the method allows differentiation between an inadequate immune response and the challenge by a breakthrough strain on animals dying despite vaccination. Up to now, genetic typing does not avoid cross-protection studies, which were conducted in parallel, although on a more limited scale. The importance of pathogen diversity studies for optimization of vaccine design is discussed as well as the research for new polymorphic genes. These genes may allow better predictions on cross-protection, given the recent completion of the sequence of the full genome of two E. ruminantium

  2. Generalist dinoflagellate endosymbionts and host genotype diversity detected from mesophotic (67-100 m depths coral Leptoseris

    Directory of Open Access Journals (Sweden)

    Kahng Samuel E

    2009-09-01

    Full Text Available Abstract Background Mesophotic corals (light-dependent corals in the deepest half of the photic zone at depths of 30 - 150 m provide a unique opportunity to study the limits of the interactions between corals and endosymbiotic dinoflagellates in the genus Symbiodinium. We sampled Leptoseris spp. in Hawaii via manned submersibles across a depth range of 67 - 100 m. Both the host and Symbiodinium communities were genotyped, using a non-coding region of the mitochondrial ND5 intron (NAD5 and the nuclear ribosomal internal transcribed spacer region 2 (ITS2, respectively. Results Coral colonies harbored endosymbiotic communities dominated by previously identified shallow water Symbiodinium ITS2 types (C1_ AF333515, C1c_ AY239364, C27_ AY239379, and C1b_ AY239363 and exhibited genetic variability at mitochondrial NAD5. Conclusion This is one of the first studies to examine genetic diversity in corals and their endosymbiotic dinoflagellates sampled at the limits of the depth and light gradients for hermatypic corals. The results reveal that these corals associate with generalist endosymbiont types commonly found in shallow water corals and implies that the composition of the Symbiodinium community (based on ITS2 alone is not responsible for the dominance and broad depth distribution of Leptoseris spp. The level of genetic diversity detected in the coral NAD5 suggests that there is undescribed taxonomic diversity in the genus Leptoseris from Hawaii.

  3. Generalist dinoflagellate endosymbionts and host genotype diversity detected from mesophotic (67-100 m depths) coral Leptoseris

    Science.gov (United States)

    Chan, Yvonne L; Pochon, Xavier; Fisher, Marla A; Wagner, Daniel; Concepcion, Gregory T; Kahng, Samuel E; Toonen, Robert J; Gates, Ruth D

    2009-01-01

    Background Mesophotic corals (light-dependent corals in the deepest half of the photic zone at depths of 30 - 150 m) provide a unique opportunity to study the limits of the interactions between corals and endosymbiotic dinoflagellates in the genus Symbiodinium. We sampled Leptoseris spp. in Hawaii via manned submersibles across a depth range of 67 - 100 m. Both the host and Symbiodinium communities were genotyped, using a non-coding region of the mitochondrial ND5 intron (NAD5) and the nuclear ribosomal internal transcribed spacer region 2 (ITS2), respectively. Results Coral colonies harbored endosymbiotic communities dominated by previously identified shallow water Symbiodinium ITS2 types (C1_ AF333515, C1c_ AY239364, C27_ AY239379, and C1b_ AY239363) and exhibited genetic variability at mitochondrial NAD5. Conclusion This is one of the first studies to examine genetic diversity in corals and their endosymbiotic dinoflagellates sampled at the limits of the depth and light gradients for hermatypic corals. The results reveal that these corals associate with generalist endosymbiont types commonly found in shallow water corals and implies that the composition of the Symbiodinium community (based on ITS2) alone is not responsible for the dominance and broad depth distribution of Leptoseris spp. The level of genetic diversity detected in the coral NAD5 suggests that there is undescribed taxonomic diversity in the genus Leptoseris from Hawaii. PMID:19747389

  4. Generalist dinoflagellate endosymbionts and host genotype diversity detected from mesophotic (67-100 m depths) coral Leptoseris.

    Science.gov (United States)

    Chan, Yvonne L; Pochon, Xavier; Fisher, Marla A; Wagner, Daniel; Concepcion, Gregory T; Kahng, Samuel E; Toonen, Robert J; Gates, Ruth D

    2009-09-11

    Mesophotic corals (light-dependent corals in the deepest half of the photic zone at depths of 30-150 m) provide a unique opportunity to study the limits of the interactions between corals and endosymbiotic dinoflagellates in the genus Symbiodinium. We sampled Leptoseris spp. in Hawaii via manned submersibles across a depth range of 67-100 m. Both the host and Symbiodinium communities were genotyped, using a non-coding region of the mitochondrial ND5 intron (NAD5) and the nuclear ribosomal internal transcribed spacer region 2 (ITS2), respectively. Coral colonies harbored endosymbiotic communities dominated by previously identified shallow water Symbiodinium ITS2 types (C1_ AF333515, C1c_ AY239364, C27_ AY239379, and C1b_ AY239363) and exhibited genetic variability at mitochondrial NAD5. This is one of the first studies to examine genetic diversity in corals and their endosymbiotic dinoflagellates sampled at the limits of the depth and light gradients for hermatypic corals. The results reveal that these corals associate with generalist endosymbiont types commonly found in shallow water corals and implies that the composition of the Symbiodinium community (based on ITS2) alone is not responsible for the dominance and broad depth distribution of Leptoseris spp. The level of genetic diversity detected in the coral NAD5 suggests that there is undescribed taxonomic diversity in the genus Leptoseris from Hawaii.

  5. Avian nephritis virus (ANV) on Brazilian chickens farms: circulating genotypes and intra-genotypic diversity.

    Science.gov (United States)

    Espinoza, Luis Luna; Beserra, Laila A R; Soares, Rodrigo M; Gregori, Fabio

    2016-12-01

    Avian nephritis virus (ANV), which belongs to the family Astroviridae, is associated with different clinical manifestations (including enteric disorders). Despite being frequently found in the avian industry worldwide, information regarding genetic features of these viruses in Brazil is scarce. Therefore, sixty fecal sample pools (5-6 birds of the same flock), representing 60 poultry farms from six Brazilian States, were screened using an astrovirus-specific hemi-nested-PCR assay targeting the conserved ORF1b gene, followed by nucleotide sequencing of amplified products. PCR and phylogenetic analysis confirmed the detection of 21 positive samples to ANV (35 %). In order to investigate the genetic diversity represented by these viruses, amplification, cloning and phylogenetic analysis of the deduced amino acid sequence of ORF2 gene were attempted. Eight samples were successfully cloned (generating 32 clones in total) and sequenced. Based on phylogenetic analysis of ORF2, sequences defined in this study were classified into three genotypes: genotype 5, which has already been described in birds, and two other novel genotypes, tentatively named genotype 8 and 9, all of which occurred in single or mixed infections. Moreover, high intra-genotypic diversity and co-circulation of distinct strains in a same host population were observed. This study revealed the presence of new strains of ANV in Brazilian poultry and their circulation in commercial chicken flocks.

  6. Genetic diversity in soybean genotypes with resistance to Heterodera glycines

    Directory of Open Access Journals (Sweden)

    Ana Paula Oliveira Nogueira

    2011-01-01

    Full Text Available The purpose of this study was to analyze the genetic diversity among soybean genotypes inoculated with Heteroderaglycines race 3. The experiments were conducted in a greenhouse. In two performance tests of morphological characteristics andresistance to the pathogen, 27 soybean genotypes were assessed. The coefficient of genotypic determination was estimated by themethod of analysis of variance and the genetic diversity analyzed based on dendrograms and optimization method. The estimatedcoefficients of determination indicated a predominantly genetic origin of the genotypic differences in the traits. The genetic variabilitywas maintained in the superior genotypes, which can be used in breeding programs for resistance to soybean cyst nematode

  7. Genotypic Diversity of Escherichia coli in the Water and Soil of Tropical Watersheds in Hawaii ▿

    OpenAIRE

    Goto, Dustin K.; Yan, Tao

    2011-01-01

    High levels of Escherichia coli were frequently detected in tropical soils in Hawaii, which present important environmental sources of E. coli to water bodies. This study systematically examined E. coli isolates from water and soil of several watersheds in Hawaii and observed high overall genotypic diversity (35.5% unique genotypes). In the Manoa watershed, fewer than 9.3% of the observed E. coli genotypes in water and 6.6% in soil were shared between different sampling sites, suggesting the ...

  8. Protein landmarks for diversity assessment in wheat genotypes

    African Journals Online (AJOL)

    jai ganesha

    1Department of Molecular Biology and Biotechnology, MPUAT Udaipur, Rajasthan, India. ... attributes exhibited the importance of seed storage as a marker system. .... were monomorphic for all genotypes. ..... bands, percent polymorphism Nei's gene diversity (h), Shannon's Information index (I), total genetic diversity.

  9. Phenotypic and Genotypic Diversity of Salmonella in Finishing Swine.

    Science.gov (United States)

    Pires, Alda F A; Funk, Julie A; Habing, Greg G; Bolin, Carole

    2016-04-01

    Salmonella enterica (nontyphoidal) is one of the major causes of foodborne diseases in the United States and worldwide. Molecular typing methods are significant tools used to better understand the transmission and ecology of Salmonella in order to implement pre-harvest control measures. The objectives of this study were to describe the Salmonella genotypes, the distribution of isolate subtypes from different ecological niches (i.e., barn environment, nursery, and individual pigs) and their evolution over time in a longitudinal study conducted in three finishing sites (housing pigs from 10 weeks of age until slaughter at 24-26 weeks of age). Among the 107 Salmonella isolates submitted for pulsed-field gel electrophoresis (PFGE) analysis, there were 25 distinct subtypes. PFGE genotyping results were consistent with the serotype findings. A large number of distinguishable PFGE patterns (i.e., within the same serovar) were observed and different combinations of subtypes were identified within and across sites and cohorts. New subtypes may result of the introduction of new strains, genetic changes, or ongoing transmission of evolved strains within the production system. The same subtypes were detected intermittently during the study period, which suggests the persistence of indistinguishable subtypes in this production system. In addition, this study suggests persistence of the same subtype over several cohorts of pigs and potential residual contamination from the barn. Factors affecting adaptation and transmission of Salmonella within and among ecological systems (e.g., finishing pigs, nursery, and environment) should be further investigated. Understanding genotypic diversity of Salmonella in different ecological niches during pre-harvest may contribute to the development of more targeted and cost effective control programs during nursery and finishing phases.

  10. Tree species, tree genotypes and tree genotypic diversity levels affect microbe-mediated soil ecosystem functions in a subtropical forest

    OpenAIRE

    Purahong, Witoon; Durka, Walter; Fischer, Markus; Dommert, Sven; Schöps, Ricardo; Buscot, François; Wubet, Tesfaye

    2016-01-01

    Tree species identity and tree genotypes contribute to the shaping of soil microbial communities. However, knowledge about how these two factors influence soil ecosystem functions is still lacking. Furthermore, in forest ecosystems tree genotypes co-occur and interact with each other, thus the effects of tree genotypic diversity on soil ecosystem functions merit attention. Here we investigated the effects of tree species, tree genotypes and genotypic diversity levels, alongside soil physicoch...

  11. High genotypic diversity found among population of Phytophthora infestans collected in Estonia.

    Science.gov (United States)

    Runno-Paurson, Eve; Kiiker, Riinu; Joutsjoki, Tiina; Hannukkala, Asko

    2016-03-01

    Potato late blight, caused by the oomycete pathogen Phytophthora infestans, is one of the most important diseases of potato worldwide. This is the first study characterising Estonian P. infestans population using the SSR marker genotyping method. 70 P. infestans isolates collected during the growing season in 2004 from eight potato fields in three different regions of Estonia were characterised with nine polymorphic SSR markers. A1 and A2 mating type isolates were detected from every studied field indicating the high potential for sexual reproduction, which raises the genotypic diversity in P. infestans population. Results revealed highly diverse P. infestans population in Estonia resembling the Northern European populations. Most of the multilocus genotypes were detected only once among the collected isolates. Subpopulations collected from different geographical regions of Estonia showed no differentiation from each other but instead formed one highly diverse group.

  12. The potential of plant viruses to promote genotypic diversity via genotype x environment interactions

    DEFF Research Database (Denmark)

    van Mölken, Tamara; Stuefer, Josef F.

    2011-01-01

    † Background and Aims Genotype by environment (G × E) interactions are important for the long-term persistence of plant species in heterogeneous environments. It has often been suggested that disease is a key factor for the maintenance of genotypic diversity in plant populations. However, empirical...... evidence for this contention is scarce. Here virus infection is proposed as a possible candidate for maintaining genotypic diversity in their host plants. † Methods The effects of White clover mosaic virus (WClMV) on the performance and development of different Trifolium repens genotypes were analysed...... and the G × E interactions were examined with respect to genotypespecific plant responses to WClMV infection. Thus, the environment is defined as the presence or absence of the virus. † Key Results WClMV had a negative effect on plant performance as shown by a decrease in biomass and number of ramets...

  13. Genotypic diversity of S. mutans in dental biofilm formed in situ under sugar stress exposure.

    Science.gov (United States)

    Arthur, Rodrigo Alex; Tabchoury, Cínthia Pereira Machado; Mattos-Graner, Renata de Oliveira; Del Bel Cury, Altair A; Paes Leme, Adriana Franco; Vale, Gláuber Campos; Cury, Jaime Aparecido

    2007-01-01

    In situ dental biofilm composition under sugar exposure is well known, but sugar effect on the genotypic diversity of S. mutans in dental biofilm has not been explored. This study evaluated S. mutans genotypic diversity in dental biofilm formed in situ under frequent exposure to sucrose and its monosaccharide constituents (glucose and fructose). Saliva of 7 volunteers was collected for isolation of S. mutans and the same volunteers wore intraoral palatal appliances, containing enamel slabs, which were submitted to the following treatments: distilled and deionized water (negative control), 10% glucose + 10% fructose (fermentable carbohydrates) solution or 20% sucrose (fermentable and EPS inductor) solution, 8x/day. After 3, 7 and 14 days, the biofilms were collected and S. mutans colonies were isolated. Arbitrarily primed polymerase chain reaction (AP-PCR) of S. mutans showed that salivary genotypes were also detected in almost all biofilm samples, independently of the treatment, and seemed to reflect those genotypes present at higher proportion in biofilms. In addition to the salivary genotypes, others were found in biofilms but in lower proportions and were distinct among treatment. The data suggest that the in situ model seems to be useful to evaluate genotypic diversity of S. mutans, but, under the tested conditions, it was not possible to clearly show that specific genotypes were selected in the biofilm due to the stress induced by sucrose metabolism or simple fermentation of its monosaccharides.

  14. Genetic diversity of some chili (Capsicum annuum L. genotypes

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    M.J. Hasan

    2014-06-01

    Full Text Available A study on genetic diversity was conducted with 54 Chili (Capsicum annuum L. genotypes through Mohalanobis’s D2 and principal component analysis for twelve quantitative characters viz. plant height, number of secondary branch/plant, canopy breadth , days to first flowering, days to 50% flowering, fruits/plant, 5 fruits weight, fruit length, fruit diameter, seeds/fruit, 1000 seed weight and yield/plant were taken into consideration. Cluster analysis was used for grouping of 54 chili genotypes and the genotypes were fallen into seven clusters. Cluster II had maximum (13 and cluster III had the minimum number (1 of genotypes. The highest inter-cluster distance was observed between cluster I and III and the lowest between cluster II and VII. The characters yield/plant, canopy breadth, secondary branches/plant, plant height and seeds/fruit contributed most for divergence in the studied genotypes. Considering group distance, mean performance and variability the inter genotypic crosses between cluster I and cluster III, cluster III and cluster VI, cluster II and cluster III and cluster III and cluster VII may be suggested to use for future hybridization program.

  15. The uses of AFLP for detecting DNA polymorphism, genotype identification and genetic diversity between yeasts isolated from Mexican agave-distilled beverages and from grape musts.

    Science.gov (United States)

    Flores Berrios, E P; Alba González, J F; Arrizon Gaviño, J P; Romano, P; Capece, A; Gschaedler Mathis, A

    2005-01-01

    The objectives were to determine the variability and to compare the genetic diversity obtained using amplified fragment length polymorphism (AFLP) markers in analyses of wine, tequila, mezcal, sotol and raicilla yeasts. A molecular characterization of yeasts isolated from Mexican agave musts, has been performed by AFLP marker analysis, using reference wine strains from Italian and South African regions. A direct co-relation between genetic profile, origin and fermentation process of strains was found especially in strains isolated from agave must. In addition, unique molecular markers were obtained for all the strains using six combination primers, confirming the discriminatory power of AFLP markers. This is the first report of molecular characterization between yeasts isolated from different Mexican traditional agave-distilled beverages, which shows high genetic differences with respect to wine strains.

  16. Tree species, tree genotypes and tree genotypic diversity levels affect microbe-mediated soil ecosystem functions in a subtropical forest

    Science.gov (United States)

    Purahong, Witoon; Durka, Walter; Fischer, Markus; Dommert, Sven; Schöps, Ricardo; Buscot, François; Wubet, Tesfaye

    2016-11-01

    Tree species identity and tree genotypes contribute to the shaping of soil microbial communities. However, knowledge about how these two factors influence soil ecosystem functions is still lacking. Furthermore, in forest ecosystems tree genotypes co-occur and interact with each other, thus the effects of tree genotypic diversity on soil ecosystem functions merit attention. Here we investigated the effects of tree species, tree genotypes and genotypic diversity levels, alongside soil physicochemical properties, on the overall and specific soil enzyme activity patterns. Our results indicate that tree species identity, tree genotypes and genotypic diversity level have significant influences on overall and specific soil enzyme activity patterns. These three factors influence soil enzyme patterns partly through effects on soil physicochemical properties and substrate quality. Variance partitioning showed that tree species identity, genotypic diversity level, pH and water content all together explained ~30% variations in the overall patterns of soil enzymes. However, we also found that the responses of soil ecosystem functions to tree genotypes and genotypic diversity are complex, being dependent on tree species identity and controlled by multiple factors. Our study highlights the important of inter- and intra-specific variations in tree species in shaping soil ecosystem functions in a subtropical forest.

  17. Tree species, tree genotypes and tree genotypic diversity levels affect microbe-mediated soil ecosystem functions in a subtropical forest

    Science.gov (United States)

    Purahong, Witoon; Durka, Walter; Fischer, Markus; Dommert, Sven; Schöps, Ricardo; Buscot, François; Wubet, Tesfaye

    2016-01-01

    Tree species identity and tree genotypes contribute to the shaping of soil microbial communities. However, knowledge about how these two factors influence soil ecosystem functions is still lacking. Furthermore, in forest ecosystems tree genotypes co-occur and interact with each other, thus the effects of tree genotypic diversity on soil ecosystem functions merit attention. Here we investigated the effects of tree species, tree genotypes and genotypic diversity levels, alongside soil physicochemical properties, on the overall and specific soil enzyme activity patterns. Our results indicate that tree species identity, tree genotypes and genotypic diversity level have significant influences on overall and specific soil enzyme activity patterns. These three factors influence soil enzyme patterns partly through effects on soil physicochemical properties and substrate quality. Variance partitioning showed that tree species identity, genotypic diversity level, pH and water content all together explained ~30% variations in the overall patterns of soil enzymes. However, we also found that the responses of soil ecosystem functions to tree genotypes and genotypic diversity are complex, being dependent on tree species identity and controlled by multiple factors. Our study highlights the important of inter- and intra-specific variations in tree species in shaping soil ecosystem functions in a subtropical forest. PMID:27857198

  18. Mycobacterium genotypes in pulmonary tuberculosis infections and their detection by trained African giant pouched rats.

    Science.gov (United States)

    Mgode, Georgies F; Cohen-Bacrie, Stéphan; Bedotto, Marielle; Weetjens, Bart J; Cox, Christophe; Jubitana, Maureen; Kuipers, Dian; Machang'u, Robert S; Kazwala, Rudovick; Mfinanga, Sayoki G; Kaufmann, Stefan H E; Drancourt, Michel

    2015-02-01

    Tuberculosis (TB) diagnosis in low-income countries is mainly done by microscopy. Hence, little is known about the diversity of Mycobacterium spp. in TB infections. Different genotypes or lineages of Mycobacterium tuberculosis vary in virulence and induce different inflammatory and immune responses. Trained Cricetomys rats show a potential for rapid diagnosis of TB. They detect over 28 % of smear-negative, culture-positive TB. However, it is unknown whether these rats can equally detect sputa from patients infected with different genotypes of M. tuberculosis. A 4-month prospective study on diversity of Mycobacterium spp. was conducted in Dar es Salaam, Tanzania. 252 sputa from 161 subjects were cultured on Lowenstein-Jensen medium and thereafter tested by rats. Mycobacterial isolates were subjected to molecular identification and multispacer sequence typing (MST) to determine species and genotypes. A total of 34 Mycobacterium spp. isolates consisting of 32 M. tuberculosis, 1 M. avium subsp. hominissuis and 1 M. intracellulare were obtained. MST analyses of 26 M. tuberculosis isolates yielded 10 distinct MST genotypes, including 3 new genotypes with two clusters of related patterns not grouped by geographic areas. Genotype MST-67, shared by one-third of M. tuberculosis isolates, was associated with the Mwananyamala clinic. This study shows that diverse M. tuberculosis genotypes (n = 10) occur in Dar es Salaam and trained rats detect 80 % of the genotypes. Sputa with two M. tuberculosis genotypes (20 %), M. avium hominissuis and M. intracellulare were not detected. Therefore, rats detect sputa with different M. tuberculosis genotypes and can be used to detect TB in resource-poor countries.

  19. Genotypic diversity of anogenital human papillomavirus in women attending cervical cancer screening in Harare, Zimbabwe.

    Science.gov (United States)

    Dube Mandishora, Racheal S; Christiansen, Irene K; Chin'ombe, Nyasha; Duri, Kerina; Ngara, Bernard; Rounge, Trine B; Meisal, Roger; Ambur, Ole H; Palefsky, Joel M; Stray-Pedersen, Babill; Chirenje, Zvavahera M

    2017-09-01

    Although anogenital cancers have been on a gradual rise in developing countries in the past few decades, they have been understudied. The objective was to investigate genotypic diversity of anogenital HPV amongst women reporting for routine cervical cancer screening in Harare in Zimbabwe. A cross-sectional study that enrolled 144 women ≥18 years from a cervical cancer-screening clinic was performed. Each woman provided a self-collected cervico-vaginal swab (VS) and a clinician-collected anal swab (CCAS). HIV testing was offered and cervical cytology was performed. Both VS and CCAS samples were HPV genotyped, using amplicon sequencing of the L1 gene region with Illumina technology. Mean age of the women was 39.9 (range 18-83 years, SD ± 11.0). HPV prevalence was 72% (104/144) in VS and 48% (69/144) in CCAS. The most common genotypes detected in both VS and CCAS were HPV18, HPV52, and HPV16. Sixty two percent of the subjects had multiple genotypic HPV infections. The odds of being HPV-positive among HIV-infected women were higher than in HIV-negative women in both the vagina and the anus (CCAS OR = 4.8; CI 2.4-9.8, P < 0.001) and (VS OR = 2.9; CI 1.3-6.4, P = 0.005). High HPV prevalence and diverse genotypes were detected in both the vagina and anus. Anal oncogenic HPV infection was common. HPV 52 was one of the most common oncogenic genotypes in both the vagina and anus. HIV co-infection played a significant role in the prevalence of HPV. These data have implications for design of primary and secondary programs for prevention of anogenital cancer in Zimbabwe. © 2017 Wiley Periodicals, Inc.

  20. Genotypic diversity and virulence factors of Streptococcus mutans in caries-free and caries-active individuals

    OpenAIRE

    Miula Portelinha Braga; Augusta Piovesan; Natália Valarini; Sandra Mara Maciel; Flaviana Bombarda de Andrade; Regina Célia Poli-Frederico

    2013-01-01

    The objective of this study was to analyze the genotypic diversity, frequency of serotypes and the detection of mutacins from Streptococcus mutans isolates in caries-free and caries-active individuals.A total of 260 S. mutans isolated from 28 individuals with and without dental caries were subjected to AP-PCR and PCR screening of glucosyltransferase B, mutacin and serotype genes, which showed the presence of. 70 different genotypes. There was no statistically significant association between t...

  1. [Genotypic Diversity of Wolbachia pipientis in Native and Invasive Harmonia axyridis Pall., 1773 (Coleoptera, Coccinellidae) Populations].

    Science.gov (United States)

    Goryacheva, I I; Blekhman, A V; Andrianov, B V; Gorelova, T V; Zakharov, I A

    2015-08-01

    The distribution and variability of reproductive symbiotic Wolbachia pipientis bacteria were studied in seven native and six invasive H. axyridis populations. Wolbachia-infected individuals were found in two invasive and two native populations. We demonstrated for the first time an infection of invasive H. axyridis populations with Wolbachia. Two new molecular forms of Wolbachia were detected by a system of multilocus typing. The supergroup A Wolbachia was found for the first time in H. axyridis. The detected genotypic diversity of Wolbachia indicates repeated and independent infection events in the evolutionary past of H. axyridis.

  2. Genetic diversity of the genotype VII Newcastle disease virus: identification of a novel VIIj sub-genotype.

    Science.gov (United States)

    Xue, Cong; Cong, Yanlong; Yin, Renfu; Sun, Yixue; Ding, Chan; Yu, Shengqing; Liu, Xiufan; Hu, Shunlin; Qian, Jing; Yuan, Qianliang; Yang, Mingxi; Wang, Chunfeng; Ding, Zhuang

    2017-02-01

    Newcastle disease (ND) is a highly contagious disease of poultry caused by Newcastle disease virus (NDV). Multiple genotypes of NDV have been circulating worldwide and NDV is continuously evolving, resulting into more diversity. Of multiple viral genotypes, VII is particularly important given that it had been associated with most recent ND outbreaks worldwide. In this study, an epidemiological investigation performed in northeastern China during 2014-2015 showed that 11 genotype VII isolates amounted to 55 percent in a total number of NDV isolates. Therefore, to evaluate the genetic diversity worldwide and epidemiological distribution in China of genotype VII NDV, a phylogenetic analysis based on the 1255 complete F gene sequences showed that VII is the most predominant genotype worldwide. A further detailed characterization on genotype VII was conducted based on the 477 complete F gene sequences from 11 isolates and 466 reference viruses available in GenBank. The results demonstrated that VII can be further divided into 8 sub-genotypes (VIIb, VIId-VIIj), indicating its complex genetic diversity. It is worthy of note that the isolation rate of VIIj is increasing recently. It emphasizes the necessity to pay close attention to the epidemiological dynamic of genotype VII NDV and highlights the importance of vaccination program.

  3. French Jura flor yeasts: genotype and technological diversity.

    Science.gov (United States)

    Charpentier, Claudine; Colin, Anne; Alais, Anne; Legras, Jean-Luc

    2009-03-01

    Fifty-four Saccharomyces cerevisiae strains were isolated from Jura "Vin Jaune" velum and characterized by conventional physiological and molecular tests including ITS RFLP and sequence analysis, karyotyping and inter delta typing. ITS RFLP and sequence revealed a specific group of related strains different from the specific profile of Sherry flor yeast caused by a 24 bp deletion in the ITS1 region described by Esteve-Zarzoso et al. (Antonie Van Leeuwenhoek 85:151-158, 2004). Interdelta typing, the most discriminative method, revealed a high diversity of Jura flor yeast strains and gathered strains in clusters unequally shared between the northern and southern part of the Jura vineyard. The assessment of phenotypic diversity among the isolated strains was investigated for three wine metabolites (ethanal, acetic acid, and sotolon) from micro scale velum tests. Except at an early stage of ageing, the production of these metabolites was not correlated to the five genetic groups obtained by interdelta typing, but correlated to the cellar where strains had been isolated. The different strains isolated in a cellar produced mostly one type of velum (thin or thick, grey or white); but thin and grey velums, recognized as responsible for high quality wines, were obtained more frequently for one of the five groups of delta genotypes.

  4. Genotypic diversity and virulence factors of Streptococcus mutans in caries-free and caries-active individuals

    Directory of Open Access Journals (Sweden)

    Miula Portelinha Braga

    2013-04-01

    Full Text Available The objective of this study was to analyze the genotypic diversity, frequency of serotypes and the detection of mutacins from Streptococcus mutans isolates in caries-free and caries-active individuals.A total of 260 S. mutans isolated from 28 individuals with and without dental caries were subjected to AP-PCR and PCR screening of glucosyltransferase B, mutacin and serotype genes, which showed the presence of. 70 different genotypes. There was no statistically significant association between the presence of genes for serotypes and mutacins with dental caries. However, there was a statistically significant and a strong association between the higher genotypic diversity in the subjects with caries (r = 0.72, p = 0.001. There was an increase in the number of genotypes with increasing age (p <0.01.

  5. Inter-varietal interactions among plants in genotypically diverse mixtures tend to decrease herbivore performance.

    Science.gov (United States)

    Grettenberger, Ian M; Tooker, John F

    2016-09-01

    Much research has explored the effects of plant species diversity on herbivore populations, but far less has considered effects of plant genotypic diversity, or how abiotic stressors, like drought, can modify effects. Mechanisms by which plant genotypic diversity affects herbivore populations remain largely unresolved. We used greenhouse studies with a model system of wheat (Triticum aestivum L.) and bird cherry-oat aphid (Rhopalosiphum padi L.) to determine whether the genotypic diversity of a plant's neighborhood influences performance and fitness of herbivores on a focal plant and if drought changes the influence of neighborhood diversity. Taken across all varieties we tested, plant-plant interactions in diverse neighborhoods reduced aphid performance and generated associational resistance, although effects on aphids depended on variety identity. In diverse mixtures, drought stress greatly diminished the genotypic diversity-driven reduction in aphid performance. Neighborhood diversity influenced mother aphid size, and appeared to partially explain how plant-plant interactions reduced the number of offspring produced in mixtures. Plant size did not mediate effects on aphid performance, although neighborhood diversity reduced plant mass across varieties and watering treatments. Our results suggest inter-varietal interactions in genotypic mixtures can affect herbivore performance in the absence of herbivore movement and that abiotic stress may diminish any effects. Accounting for how neighborhood diversity influences resistance of an individual plant to herbivores will help aid development of mixtures of varieties for managing insect pests and clarify the role of plant genotypic diversity in ecosystems.

  6. Genotype-by-temperature interactions may help to maintain clonal diversity in asterionella formosa (Bacillariophyceae)

    NARCIS (Netherlands)

    Gsell, A.S.; Domis, L.N.D.; Przytulska-Bartosiewicz, A.; Mooij, W.M.; Donk, van E.; Ibelings, B.W.

    2012-01-01

    Marine and freshwater phytoplankton populations often show large clonal diversity, which is in disagreement with clonal selection of the most vigorous genotype(s). Temporal fluctuation in selection pressures in variable environments is a leading explanation for maintenance of such genetic diversity.

  7. Genotypic diversity of root and shoot characteristics of

    Directory of Open Access Journals (Sweden)

    ali ganjali

    2009-06-01

    Full Text Available Root and shoot characteristics of chickpea (Cicer arietinum L. genotypes are believed to be important in drought tolerance. There is a little information about the response of genotypes root growth in hydroponics and greenhouse culture, also the relationships between root size and drought tolerance. This study was conducted to observe whether genotypes differ in root size, and to see that root size is associated with drought tolerance during early vegetative growth. We found significant differences (p0.01 in root dry weight, total root length, tap root length, root area, leaf dry weight, leaf area and shoot biomass per plant among 30 genotypes of chickpea grown in hydroponics culture for three weeks. Each of these parameters correlated with all others, positively. Among 30 genotypes, 10 genotypes with different root sizes were selected and were grown in a greenhouse in sand culture experiment under drought stress (FC %30 for three weeks. There were not linear or non-linear significant correlations between root characters in hydroponics and greenhouse environments. It seems that environmental factors are dominant on genetic factors in seedling stage and so, the expression of genotypics potential for root growth characteristics of genotypes are different in hydroponic and greenhouse conditions. In this study, the selection of genotypes with vigorous roots system in hydroponic condition did not lead to genotypes with the same root characters in greenhouse environment. The genotype×drought interactions for root characters of chickpea seedlings in 30 days were not significant (p

  8. Genetic diversity analysis of Gossypium arboreum germplasm accessions using genotyping-by-sequencing.

    Science.gov (United States)

    Li, Ruijuan; Erpelding, John E

    2016-10-01

    The diploid cotton species Gossypium arboreum possesses many favorable agronomic traits such as drought tolerance and disease resistance, which can be utilized in the development of improved upland cotton cultivars. The USDA National Plant Germplasm System maintains more than 1600 G. arboreum accessions. Little information is available on the genetic diversity of the collection thereby limiting the utilization of this cotton species. The genetic diversity and population structure of the G. arboreum germplasm collection were assessed by genotyping-by-sequencing of 375 accessions. Using genome-wide single nucleotide polymorphism sequence data, two major clusters were inferred with 302 accessions in Cluster 1, 64 accessions in Cluster 2, and nine accessions unassigned due to their nearly equal membership to each cluster. These two clusters were further evaluated independently resulting in the identification of two sub-clusters for the 302 Cluster 1 accessions and three sub-clusters for the 64 Cluster 2 accessions. Low to moderate genetic diversity between clusters and sub-clusters were observed indicating a narrow genetic base. Cluster 2 accessions were more genetically diverse and the majority of the accessions in this cluster were landraces. In contrast, Cluster 1 is composed of varieties or breeding lines more recently added to the collection. The majority of the accessions had kinship values ranging from 0.6 to 0.8. Eight pairs of accessions were identified as potential redundancies due to their high kinship relatedness. The genetic diversity and genotype data from this study are essential to enhance germplasm utilization to identify genetically diverse accessions for the detection of quantitative trait loci associated with important traits that would benefit upland cotton improvement.

  9. Genetic diversity analysis of highly incomplete SNP genotype data with imputations: an empirical assessment.

    Science.gov (United States)

    Fu, Yong-Bi

    2014-03-13

    Genotyping by sequencing (GBS) recently has emerged as a promising genomic approach for assessing genetic diversity on a genome-wide scale. However, concerns are not lacking about the uniquely large unbalance in GBS genotype data. Although some genotype imputation has been proposed to infer missing observations, little is known about the reliability of a genetic diversity analysis of GBS data, with up to 90% of observations missing. Here we performed an empirical assessment of accuracy in genetic diversity analysis of highly incomplete single nucleotide polymorphism genotypes with imputations. Three large single-nucleotide polymorphism genotype data sets for corn, wheat, and rice were acquired, and missing data with up to 90% of missing observations were randomly generated and then imputed for missing genotypes with three map-independent imputation methods. Estimating heterozygosity and inbreeding coefficient from original, missing, and imputed data revealed variable patterns of bias from assessed levels of missingness and genotype imputation, but the estimation biases were smaller for missing data without genotype imputation. The estimates of genetic differentiation were rather robust up to 90% of missing observations but became substantially biased when missing genotypes were imputed. The estimates of topology accuracy for four representative samples of interested groups generally were reduced with increased levels of missing genotypes. Probabilistic principal component analysis based imputation performed better in terms of topology accuracy than those analyses of missing data without genotype imputation. These findings are not only significant for understanding the reliability of the genetic diversity analysis with respect to large missing data and genotype imputation but also are instructive for performing a proper genetic diversity analysis of highly incomplete GBS or other genotype data.

  10. Diverse sapovirus genotypes identified in children hospitalised with gastroenteritis in selected regions of South Africa.

    Science.gov (United States)

    Murray, Tanya Y; Nadan, Sandrama; Page, Nicola A; Taylor, Maureen B

    2016-03-01

    Sapoviruses (SaVs) are recognised as causative agents of gastroenteritis worldwide. However, data on the genetic diversity of this virus in Africa are lacking, particularly in the form of current long-term studies. To determine the genetic diversity of SaVs in children hospitalised with gastroenteritis in South Africa (SA). From April 2009 to December 2013, SaVs were characterised from stool specimens from children hospitalised with gastroenteritis in four provinces of SA. Fourteen different SaV genotypes were identified from the 221 strains that were characterised. Genogroup (G) IV predominated overall and was detected in 24% (53/221) of specimens. The other identified genotypes included six belonging to GI (GI.1-GI.3, GI.5, GI.6, and GI.7) and seven belonging to GII (GII.1-GII.7). This study has provided the first comprehensive data on the genetic diversity of SaVs in a clinical setting in SA, contributing to the global knowledge of this virus. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. [Genotypic diversity of Streptococcus sobrinus in 3 to 4-year-old children suffering with severe early childhood caries].

    Science.gov (United States)

    Qin, Xiu-rong; Zhou, Qiong; Qin, Man

    2009-06-01

    To evaluate the genotypic diversity of Streptococcus sobrinus (Ss) between children suffering with severe early childhood caries (S-ECC) and caries-free children by arbitrarily primed-polymerase chain reaction (AP-PCR). A total of 178 children aged from 42 to 54 months were recruited from 14 urban kindergartens. The S-ECC group contained 87 children with more than 5 decayed teeth, and the control group was composed of 91 caries-free children. Stimulated whole saliva was collected by chewing paraffin. All mutans streptococcus isolates were subcultured, biochemically characterised and identified by PCR as Streptococcus mutans (Sm) and Ss. Then the Ss isolates were genotyped by AP-PCR. The frequency of Ss detection was 18% in S-ECC children, which was significantly higher than 3% in caries-free children (P < 0.01). Twenty-two distinct genotypes of Ss were identified from 53 clinical isolates. In S-ECC group, one to three genotypes of Ss were detected in each saliva sample. Only one genotype of Ss was detected in all the caries-free children. One genotype of Ss were shared by three S-ECC children. The genotypes of isolates in S-ECC group were relate to decayed-missing-filled teeth (r = 0.50, P < 0. 05). The rate of Ss detection was significantly higher in S-ECC children than in caries-free children. Isolates of Ss displayed genetic polymorphism. The multi-genotypes of Ss was related to differences in caries susceptibility. Strains of Ss with same genotype were present in unrelated subjects.

  12. Mixed genotype transmission bodies and virions contribute to the maintenance of diversity in an insect virus

    Science.gov (United States)

    Clavijo, Gabriel; Williams, Trevor; Muñoz, Delia; Caballero, Primitivo; López-Ferber, Miguel

    2010-01-01

    An insect nucleopolyhedrovirus naturally survives as a mixture of at least nine genotypes. Infection by multiple genotypes results in the production of virus occlusion bodies (OBs) with greater pathogenicity than those of any genotype alone. We tested the hypothesis that each OB contains a genotypically diverse population of virions. Few insects died following inoculation with an experimental two-genotype mixture at a dose of one OB per insect, but a high proportion of multiple infections were observed (50%), which differed significantly from the frequencies predicted by a non-associated transmission model in which genotypes are segregated into distinct OBs. By contrast, insects that consumed multiple OBs experienced higher mortality and infection frequencies did not differ significantly from those of the non-associated model. Inoculation with genotypically complex wild-type OBs indicated that genotypes tend to be transmitted in association, rather than as independent entities, irrespective of dose. To examine the hypothesis that virions may themselves be genotypically heterogeneous, cell culture plaques derived from individual virions were analysed to reveal that one-third of virions was of mixed genotype, irrespective of the genotypic composition of the OBs. We conclude that co-occlusion of genotypically distinct virions in each OB is an adaptive mechanism that favours the maintenance of virus diversity during insect-to-insect transmission. PMID:19939845

  13. Non-additive effects of genotypic diversity increase floral abundance and abundance of floral visitors.

    Directory of Open Access Journals (Sweden)

    Mark A Genung

    Full Text Available BACKGROUND: In the emerging field of community and ecosystem genetics, genetic variation and diversity in dominant plant species have been shown to play fundamental roles in maintaining biodiversity and ecosystem function. However, the importance of intraspecific genetic variation and diversity to floral abundance and pollinator visitation has received little attention. METHODOLOGY/PRINCIPAL FINDINGS: Using an experimental common garden that manipulated genotypic diversity (the number of distinct genotypes per plot of Solidago altissima, we document that genotypic diversity of a dominant plant can indirectly influence flower visitor abundance. Across two years, we found that 1 plant genotype explained 45% and 92% of the variation in flower visitor abundance in 2007 and 2008, respectively; and 2 plant genotypic diversity had a positive and non-additive effect on floral abundance and the abundance of flower visitors, as plots established with multiple genotypes produced 25% more flowers and received 45% more flower visits than would be expected under an additive model. CONCLUSIONS/SIGNIFICANCE: These results provide evidence that declines in genotypic diversity may be an important but little considered factor for understanding plant-pollinator dynamics, with implications for the global decline in pollinators due to reduced plant diversity in both agricultural and natural ecosystems.

  14. Genotypic diversity effects on the performance of Taraxacum officinale populations increase with time and environmental favorability.

    Directory of Open Access Journals (Sweden)

    Emily B M Drummond

    Full Text Available Within-population genetic diversity influences many ecological processes, but few studies have examined how environmental conditions may impact these short-term diversity effects. Over four growing seasons, we followed experimental populations of a clonal, ubiquitous weed, Taraxacum officinale, with different numbers of genotypes in relatively favorable fallow field and unfavorable mowed lawn environmental treatments. Population performance (measured as total leaf area, seed production or biomass clearly and consistently increased with diversity, and this effect became stronger over the course of the experiment. Diversity effects were stronger, and with different underlying mechanisms, in the fallow field versus the mowed lawn. Large genotypes dominated in the fallow field driving overyielding (via positive selection effects, whereas in the mowed lawn, where performance was limited by regular disturbance, there was evidence for complementarity among genotypes (with one compact genotype in particular performing better in mixture than monoculture. Hence, we predict stronger genotypic diversity effects in environments where intense intraspecific competition enhances genotypic differences. Our four-year field experiment plus seedling establishment trials indicate that genotypic diversity effects have far-reaching and context-dependent consequences across generations.

  15. Genotypic diversity effects on the performance of Taraxacum officinale populations increase with time and environmental favorability.

    Science.gov (United States)

    Drummond, Emily B M; Vellend, Mark

    2012-01-01

    Within-population genetic diversity influences many ecological processes, but few studies have examined how environmental conditions may impact these short-term diversity effects. Over four growing seasons, we followed experimental populations of a clonal, ubiquitous weed, Taraxacum officinale, with different numbers of genotypes in relatively favorable fallow field and unfavorable mowed lawn environmental treatments. Population performance (measured as total leaf area, seed production or biomass) clearly and consistently increased with diversity, and this effect became stronger over the course of the experiment. Diversity effects were stronger, and with different underlying mechanisms, in the fallow field versus the mowed lawn. Large genotypes dominated in the fallow field driving overyielding (via positive selection effects), whereas in the mowed lawn, where performance was limited by regular disturbance, there was evidence for complementarity among genotypes (with one compact genotype in particular performing better in mixture than monoculture). Hence, we predict stronger genotypic diversity effects in environments where intense intraspecific competition enhances genotypic differences. Our four-year field experiment plus seedling establishment trials indicate that genotypic diversity effects have far-reaching and context-dependent consequences across generations.

  16. Prevalence of single nucleotide polymorphism among 27 diverse alfalfa genotypes as assessed by transcriptome sequencing

    Directory of Open Access Journals (Sweden)

    Li Xuehui

    2012-10-01

    Full Text Available Abstract Background Alfalfa, a perennial, outcrossing species, is a widely planted forage legume producing highly nutritious biomass. Currently, improvement of cultivated alfalfa mainly relies on recurrent phenotypic selection. Marker assisted breeding strategies can enhance alfalfa improvement efforts, particularly if many genome-wide markers are available. Transcriptome sequencing enables efficient high-throughput discovery of single nucleotide polymorphism (SNP markers for a complex polyploid species. Result The transcriptomes of 27 alfalfa genotypes, including elite breeding genotypes, parents of mapping populations, and unimproved wild genotypes, were sequenced using an Illumina Genome Analyzer IIx. De novo assembly of quality-filtered 72-bp reads generated 25,183 contigs with a total length of 26.8 Mbp and an average length of 1,065 bp, with an average read depth of 55.9-fold for each genotype. Overall, 21,954 (87.2% of the 25,183 contigs represented 14,878 unique protein accessions. Gene ontology (GO analysis suggested that a broad diversity of genes was represented in the resulting sequences. The realignment of individual reads to the contigs enabled the detection of 872,384 SNPs and 31,760 InDels. High resolution melting (HRM analysis was used to validate 91% of 192 putative SNPs identified by sequencing. Both allelic variants at about 95% of SNP sites identified among five wild, unimproved genotypes are still present in cultivated alfalfa, and all four US breeding programs also contain a high proportion of these SNPs. Thus, little evidence exists among this dataset for loss of significant DNA sequence diversity from either domestication or breeding of alfalfa. Structure analysis indicated that individuals from the subspecies falcata, the diploid subspecies caerulea, and the tetraploid subspecies sativa (cultivated tetraploid alfalfa were clearly separated. Conclusion We used transcriptome sequencing to discover large numbers of SNPs

  17. Diversity and genetic stability in banana genotypes in a breeding program using inter simple sequence repeats (ISSR) markers.

    Science.gov (United States)

    Silva, A V C; Nascimento, A L S; Vitória, M F; Rabbani, A R C; Soares, A N R; Lédo, A S

    2017-02-23

    Banana (Musa spp) is a fruit species frequently cultivated and consumed worldwide. Molecular markers are important for estimating genetic diversity in germplasm and between genotypes in breeding programs. The objective of this study was to analyze the genetic diversity of 21 banana genotypes (FHIA 23, PA42-44, Maçã, Pacovan Ken, Bucaneiro, YB42-47, Grand Naine, Tropical, FHIA 18, PA94-01, YB42-17, Enxerto, Japira, Pacovã, Prata-Anã, Maravilha, PV79-34, Caipira, Princesa, Garantida, and Thap Maeo), by using inter-simple sequence repeat (ISSR) markers. Material was generated from the banana breeding program of Embrapa Cassava & Fruits and evaluated at Embrapa Coastal Tablelands. The 12 primers used in this study generated 97.5% polymorphism. Four clusters were identified among the different genotypes studied, and the sum of the first two principal components was 48.91%. From the Unweighted Pair Group Method using Arithmetic averages (UPGMA) dendrogram, it was possible to identify two main clusters and subclusters. Two genotypes (Garantida and Thap Maeo) remained isolated from the others, both in the UPGMA clustering and in the principal cordinate analysis (PCoA). Using ISSR markers, we could analyze the genetic diversity of the studied material and state that these markers were efficient at detecting sufficient polymorphism to estimate the genetic variability in banana genotypes.

  18. Are Tree Species Diversity and Genotypic Diversity Effects on Insect Herbivores Mediated by Ants?

    Science.gov (United States)

    Campos-Navarrete, María José; Abdala-Roberts, Luis; Munguía-Rosas, Miguel A.; Parra-Tabla, Víctor

    2015-01-01

    Plant diversity can influence predators and omnivores and such effects may in turn influence herbivores and plants. However, evidence for these ecological feedbacks is rare. We evaluated if the effects of tree species (SD) and genotypic diversity (GD) on the abundance of different guilds of insect herbivores associated with big-leaf mahogany (Swietenia macrophylla) were contingent upon the protective effects of ants tending extra-floral nectaries of this species. This study was conducted within a larger experiment consisting of mahogany monocultures and species polycultures of four species and –within each of these two plot types– mahogany was represented by either one or four maternal families. We selected 24 plots spanning these treatment combinations, 10 mahogany plants/plot, and within each plot experimentally reduced ant abundance on half of the selected plants, and surveyed ant and herbivore abundance. There were positive effects of SD on generalist leaf-chewers and sap-feeders, but for the latter group this effect depended on the ant reduction treatment: SD positively influenced sap-feeders under ambient ant abundance but had no effect when ant abundance was reduced; at the same time, ants had negative effects on sap feeders in monoculture but no effect in polyculture. In contrast, SD did not influence specialist stem-borers or leaf-miners and this effect was not contingent upon ant reduction. Finally, GD did not influence any of the herbivore guilds studied, and such effects did not depend on the ant treatment. Overall, we show that tree species diversity influenced interactions between a focal plant species (mahogany) and ants, and that such effects in turn mediated plant diversity effects on some (sap-feeders) but not all the herbivores guilds studied. Our results suggest that the observed patterns are dependent on the combined effects of herbivore identity, diet breadth, and the source of plant diversity. PMID:26241962

  19. Are Tree Species Diversity and Genotypic Diversity Effects on Insect Herbivores Mediated by Ants?

    Science.gov (United States)

    Campos-Navarrete, María José; Abdala-Roberts, Luis; Munguía-Rosas, Miguel A; Parra-Tabla, Víctor

    2015-01-01

    Plant diversity can influence predators and omnivores and such effects may in turn influence herbivores and plants. However, evidence for these ecological feedbacks is rare. We evaluated if the effects of tree species (SD) and genotypic diversity (GD) on the abundance of different guilds of insect herbivores associated with big-leaf mahogany (Swietenia macrophylla) were contingent upon the protective effects of ants tending extra-floral nectaries of this species. This study was conducted within a larger experiment consisting of mahogany monocultures and species polycultures of four species and -within each of these two plot types- mahogany was represented by either one or four maternal families. We selected 24 plots spanning these treatment combinations, 10 mahogany plants/plot, and within each plot experimentally reduced ant abundance on half of the selected plants, and surveyed ant and herbivore abundance. There were positive effects of SD on generalist leaf-chewers and sap-feeders, but for the latter group this effect depended on the ant reduction treatment: SD positively influenced sap-feeders under ambient ant abundance but had no effect when ant abundance was reduced; at the same time, ants had negative effects on sap feeders in monoculture but no effect in polyculture. In contrast, SD did not influence specialist stem-borers or leaf-miners and this effect was not contingent upon ant reduction. Finally, GD did not influence any of the herbivore guilds studied, and such effects did not depend on the ant treatment. Overall, we show that tree species diversity influenced interactions between a focal plant species (mahogany) and ants, and that such effects in turn mediated plant diversity effects on some (sap-feeders) but not all the herbivores guilds studied. Our results suggest that the observed patterns are dependent on the combined effects of herbivore identity, diet breadth, and the source of plant diversity.

  20. Are Tree Species Diversity and Genotypic Diversity Effects on Insect Herbivores Mediated by Ants?

    Directory of Open Access Journals (Sweden)

    María José Campos-Navarrete

    Full Text Available Plant diversity can influence predators and omnivores and such effects may in turn influence herbivores and plants. However, evidence for these ecological feedbacks is rare. We evaluated if the effects of tree species (SD and genotypic diversity (GD on the abundance of different guilds of insect herbivores associated with big-leaf mahogany (Swietenia macrophylla were contingent upon the protective effects of ants tending extra-floral nectaries of this species. This study was conducted within a larger experiment consisting of mahogany monocultures and species polycultures of four species and -within each of these two plot types- mahogany was represented by either one or four maternal families. We selected 24 plots spanning these treatment combinations, 10 mahogany plants/plot, and within each plot experimentally reduced ant abundance on half of the selected plants, and surveyed ant and herbivore abundance. There were positive effects of SD on generalist leaf-chewers and sap-feeders, but for the latter group this effect depended on the ant reduction treatment: SD positively influenced sap-feeders under ambient ant abundance but had no effect when ant abundance was reduced; at the same time, ants had negative effects on sap feeders in monoculture but no effect in polyculture. In contrast, SD did not influence specialist stem-borers or leaf-miners and this effect was not contingent upon ant reduction. Finally, GD did not influence any of the herbivore guilds studied, and such effects did not depend on the ant treatment. Overall, we show that tree species diversity influenced interactions between a focal plant species (mahogany and ants, and that such effects in turn mediated plant diversity effects on some (sap-feeders but not all the herbivores guilds studied. Our results suggest that the observed patterns are dependent on the combined effects of herbivore identity, diet breadth, and the source of plant diversity.

  1. Morphological and sequence-related amplified polymorphism-based molecular diversity of local and exotic wheat genotypes.

    Science.gov (United States)

    Abdelkhalik, S M; Salem, A K M; Abdelaziz, A R; Ammar, M H

    2016-04-28

    Assessing genetic diversity is a prerequisite for the genetic improvement of wheat. Molecular markers offer accurate and reproducible means for assessing genetic diversity. Field performance and sequence-related amplified polymorphism (SRAP)-based assessment of molecular diversity was carried out on a set of 10 local and introduced bread wheat (Triticum sativum L.) genotypes grown in the middle arid region of Saudi Arabia. The results revealed highly significant differences among the studied phenological traits and revealed a significant amount of genetic diversity across the tested genotypes. The overall performance revealed the superiority of KSU 102 in terms of yield and its components, with a yield potential of 8.7 tons/ha. Highly significant and positive correlations were observed among grain yield and biological yield, and also, spike length and spike weight. Thirteen SRAP primer combinations successfully amplified 954 fragments. The total number of genetic loci analyzed was 312. The overall polymorphism ratio was 99.67%, ranging from 98 to 100%. The polymorphic information content values ranged from 0.67 for ME11 x EM5 to 0.97 for ME9 x EM4 and ME11 x EM6, respectively. The wheat genotypes were clustered based on their genetic constitution and origin. The results demonstrate the power of SRAP primers for detecting molecular diversity and for varietal discrimination. The results show that high levels of genetic diversity exist, and suggest the potential of the tested materials for wheat crop improvement in the arid central region of Saudi Arabia.

  2. HCV genotyping from NGS short reads and its application in genotype detection from HCV mixed infected plasma.

    Science.gov (United States)

    Qiu, Ping; Stevens, Richard; Wei, Bo; Lahser, Fred; Howe, Anita Y M; Klappenbach, Joel A; Marton, Matthew J

    2015-01-01

    Genotyping of hepatitis C virus (HCV) plays an important role in the treatment of HCV. As new genotype-specific treatment options become available, it has become increasingly important to have accurate HCV genotype and subtype information to ensure that the most appropriate treatment regimen is selected. Most current genotyping methods are unable to detect mixed genotypes from two or more HCV infections. Next generation sequencing (NGS) allows for rapid and low cost mass sequencing of viral genomes and provides an opportunity to probe the viral population from a single host. In this paper, the possibility of using short NGS reads for direct HCV genotyping without genome assembly was evaluated. We surveyed the publicly-available genetic content of three HCV drug target regions (NS3, NS5A, NS5B) in terms of whether these genes contained genotype-specific regions that could predict genotype. Six genotypes and 38 subtypes were included in this study. An automated phylogenetic analysis based HCV genotyping method was implemented and used to assess different HCV target gene regions. Candidate regions of 250-bp each were found for all three genes that have enough genetic information to predict HCV genotypes/subtypes. Validation using public datasets shows 100% genotyping accuracy. To test whether these 250-bp regions were sufficient to identify mixed genotypes, we developed a random primer-based method to sequence HCV plasma samples containing mixtures of two HCV genotypes in different ratios. We were able to determine the genotypes without ambiguity and to quantify the ratio of the abundances of the mixed genotypes in the samples. These data provide a proof-of-concept that this random primed, NGS-based short-read genotyping approach does not need prior information about the viral population and is capable of detecting mixed viral infection.

  3. HCV genotyping from NGS short reads and its application in genotype detection from HCV mixed infected plasma.

    Directory of Open Access Journals (Sweden)

    Ping Qiu

    Full Text Available Genotyping of hepatitis C virus (HCV plays an important role in the treatment of HCV. As new genotype-specific treatment options become available, it has become increasingly important to have accurate HCV genotype and subtype information to ensure that the most appropriate treatment regimen is selected. Most current genotyping methods are unable to detect mixed genotypes from two or more HCV infections. Next generation sequencing (NGS allows for rapid and low cost mass sequencing of viral genomes and provides an opportunity to probe the viral population from a single host. In this paper, the possibility of using short NGS reads for direct HCV genotyping without genome assembly was evaluated. We surveyed the publicly-available genetic content of three HCV drug target regions (NS3, NS5A, NS5B in terms of whether these genes contained genotype-specific regions that could predict genotype. Six genotypes and 38 subtypes were included in this study. An automated phylogenetic analysis based HCV genotyping method was implemented and used to assess different HCV target gene regions. Candidate regions of 250-bp each were found for all three genes that have enough genetic information to predict HCV genotypes/subtypes. Validation using public datasets shows 100% genotyping accuracy. To test whether these 250-bp regions were sufficient to identify mixed genotypes, we developed a random primer-based method to sequence HCV plasma samples containing mixtures of two HCV genotypes in different ratios. We were able to determine the genotypes without ambiguity and to quantify the ratio of the abundances of the mixed genotypes in the samples. These data provide a proof-of-concept that this random primed, NGS-based short-read genotyping approach does not need prior information about the viral population and is capable of detecting mixed viral infection.

  4. Genetic diversity in palmyrah genotypes using morphological and molecular markers

    Directory of Open Access Journals (Sweden)

    V.Ponnuswami

    2010-07-01

    Full Text Available Palms are woody monocotyledons in the family Arecaceae which is placed in the order Arecales. Slow and tall growing,hardy and non branching, dioecious and perennial in nature, palmyrah palm has no distinguishing features to identify sex,stature and high neera yielding types until flowering age of about 12 to 15 years. Under these circumstances molecularmarkers can be effectively utilized to diagnose and select a genotype. Twenty palmyrah accessions were analysed usingRAPD and ISSR markers. In RAPD analysis, a total of 57 bands were obtained, among them 43 were polymorphic and restof them were monomorphic. Amplification size ranged between 250 and 3200 bp. UPGMA based cluster diagram showedthat all 20 different genotypes were grouped into four different clusters based on the stature, sex and high neera yieldingtypes. The distance matrix between genotypes showed an average distance range from 0.54 to 0.91 with a mean of 0.70. Atotal of 130 ISSR markers were scored, of which 65 were polymorphic, equivalent to 47.94% polymorphism. These markerswere used to estimate the genetic similarity among accessions using Jaccard’s similarity coefficient, with similarity valuesranging from 71.6 to 95.7%. The average number of markers produced per primer was 6.11. For each of the 21 ISSRprimers PIC value ranged between 0 and 0.46. Cluster analysis based on ISSR data grouped the 20 palmyrah accessions intotwo major clusters. PCA based on ISSR data clearly distinguished genotypes similar to the results of cluster analysis.

  5. The potential of plant viruses to promote genotypic diversity via genotype x environment interactions

    DEFF Research Database (Denmark)

    van Mölken, Tamara; Stuefer, Josef F.

    2011-01-01

    for WClMV to provoke differential selection on T. repens genotypes, which may lead to negative frequency-dependent selection in host populations. †Conclusions The apparent G × E interaction and evident repercussions for relative fitness reported in this study stress the importance of viruses...

  6. Genotypic diversity of Streptococcus mutans and Streptococcus sobrinus in 3-4-year-old children with severe caries or without caries.

    Science.gov (United States)

    Zhou, Qiong; Qin, Xiurong; Qin, Man; Ge, Lihong

    2011-11-01

    BACKGROUND.  The genotypic diversity of both Streptococcus mutans and Streptococcus sobrinus in children with different caries experience remains unclear. AIM.  To investigate the genotypic diversity of S. mutans and S. sobrinus in children with severe early childhood caries (SECC) and in caries-free (CF) children. METHODS.  Stimulated saliva of 87 SECC and 91 CF children aged 3-4 years was collected and submitted to cultivation, and MS colonies were enumerated. The genomic fingerprint analysis of S. mutans and S. sobrinus was carried out using AP-PCR. RESULTS.  One to five genotypes of S. mutans were colonized in an oral cavity of SECC and CF children; 85.5% SECC children and 57.9% CF children harboured more than one genotype of S. mutans. One to three genotypes of S. sobrinus were detected from each SECC child; 31.25% SECC children harboured more than one genotype of S. sobrinus. And one genotype was colonized in each CF child. S. mutans isolates from different individuals displayed distinctive DNA fingerprints. CONCLUSIONS.  DNA fingerprints of S. mutans and S. sobrinus isolates from 3- to 4-year-old children displayed genetic polymorphism, and S. mutans has greater genetic diversity than S. sobrinus. SECC children harboured more genotypes of S. mutans and S. sobrinus than CF children. © 2011 The Authors. International Journal of Paediatric Dentistry © 2011 BSPD, IAPD and Blackwell Publishing Ltd.

  7. Evidence for coral range expansion accompanied by reduced diversity of Symbiodinium genotypes

    Science.gov (United States)

    Grupstra, Carsten G. B.; Coma, Rafel; Ribes, Marta; Leydet, Karine Posbic; Parkinson, John Everett; McDonald, Kelly; Catllà, Marc; Voolstra, Christian R.; Hellberg, Michael E.; Coffroth, Mary Alice

    2017-09-01

    Zooxanthellate corals are threatened by climate change but may be able to escape increasing temperatures by colonizing higher latitudes. To determine the effect of host range expansion on symbiont genetic diversity, we examined genetic variation among populations of Symbiodinium psygmophilum associated with Oculina patagonica, a range-expanding coral that acquires its symbionts through horizontal transmission. We optimized five microsatellite primer pairs for S. psygmophilum and tested them on Oculina spp. samples from the western North Atlantic and the Mediterranean. We then used them to compare symbiont genotype diversity between an Iberian core and an expansion front population of O. patagonica. Only one multilocus S. psygmophilum genotype was identified at the expansion front, and it was shared with the core population, which harbored seven multilocus genotypes. This pattern suggests that O. patagonica range expansion is accompanied by reduced symbiont genetic diversity, possibly due to limited dispersal of symbionts or local selection.

  8. Evidence for coral range expansion accompanied by reduced diversity of Symbiodinium genotypes

    KAUST Repository

    Grupstra, Carsten G. B.

    2017-05-15

    Zooxanthellate corals are threatened by climate change but may be able to escape increasing temperatures by colonizing higher latitudes. To determine the effect of host range expansion on symbiont genetic diversity, we examined genetic variation among populations of Symbiodinium psygmophilum associated with Oculina patagonica, a range-expanding coral that acquires its symbionts through horizontal transmission. We optimized five microsatellite primer pairs for S. psygmophilum and tested them on Oculina spp. samples from the western North Atlantic and the Mediterranean. We then used them to compare symbiont genotype diversity between an Iberian core and an expansion front population of O. patagonica. Only one multilocus S. psygmophilum genotype was identified at the expansion front, and it was shared with the core population, which harbored seven multilocus genotypes. This pattern suggests that O. patagonica range expansion is accompanied by reduced symbiont genetic diversity, possibly due to limited dispersal of symbionts or local selection.

  9. Analysis of genotype diversity and evolution of Dengue virus serotype 2 using complete genomes

    Science.gov (United States)

    Waman, Vaishali P.; Kolekar, Pandurang; Ramtirthkar, Mukund R.; Kale, Mohan M.

    2016-01-01

    Background Dengue is one of the most common arboviral diseases prevalent worldwide and is caused by Dengue viruses (genus Flavivirus, family Flaviviridae). There are four serotypes of Dengue Virus (DENV-1 to DENV-4), each of which is further subdivided into distinct genotypes. DENV-2 is frequently associated with severe dengue infections and epidemics. DENV-2 consists of six genotypes such as Asian/American, Asian I, Asian II, Cosmopolitan, American and sylvatic. Comparative genomic study was carried out to infer population structure of DENV-2 and to analyze the role of evolutionary and spatiotemporal factors in emergence of diversifying lineages. Methods Complete genome sequences of 990 strains of DENV-2 were analyzed using Bayesian-based population genetics and phylogenetic approaches to infer genetically distinct lineages. The role of spatiotemporal factors, genetic recombination and selection pressure in the evolution of DENV-2 is examined using the sequence-based bioinformatics approaches. Results DENV-2 genetic structure is complex and consists of fifteen subpopulations/lineages. The Asian/American genotype is observed to be diversified into seven lineages. The Asian I, Cosmopolitan and sylvatic genotypes were found to be subdivided into two lineages, each. The populations of American and Asian II genotypes were observed to be homogeneous. Significant evidence of episodic positive selection was observed in all the genes, except NS4A. Positive selection operational on a few codons in envelope gene confers antigenic and lineage diversity in the American strains of Asian/American genotype. Selection on codons of non-structural genes was observed to impact diversification of lineages in Asian I, cosmopolitan and sylvatic genotypes. Evidence of intra/inter-genotype recombination was obtained and the uncertainty in classification of recombinant strains was resolved using the population genetics approach. Discussion Complete genome-based analysis revealed that the

  10. Analysis of genotype diversity and evolution of Dengue virus serotype 2 using complete genomes

    Directory of Open Access Journals (Sweden)

    Vaishali P. Waman

    2016-08-01

    Full Text Available Background Dengue is one of the most common arboviral diseases prevalent worldwide and is caused by Dengue viruses (genus Flavivirus, family Flaviviridae. There are four serotypes of Dengue Virus (DENV-1 to DENV-4, each of which is further subdivided into distinct genotypes. DENV-2 is frequently associated with severe dengue infections and epidemics. DENV-2 consists of six genotypes such as Asian/American, Asian I, Asian II, Cosmopolitan, American and sylvatic. Comparative genomic study was carried out to infer population structure of DENV-2 and to analyze the role of evolutionary and spatiotemporal factors in emergence of diversifying lineages. Methods Complete genome sequences of 990 strains of DENV-2 were analyzed using Bayesian-based population genetics and phylogenetic approaches to infer genetically distinct lineages. The role of spatiotemporal factors, genetic recombination and selection pressure in the evolution of DENV-2 is examined using the sequence-based bioinformatics approaches. Results DENV-2 genetic structure is complex and consists of fifteen subpopulations/lineages. The Asian/American genotype is observed to be diversified into seven lineages. The Asian I, Cosmopolitan and sylvatic genotypes were found to be subdivided into two lineages, each. The populations of American and Asian II genotypes were observed to be homogeneous. Significant evidence of episodic positive selection was observed in all the genes, except NS4A. Positive selection operational on a few codons in envelope gene confers antigenic and lineage diversity in the American strains of Asian/American genotype. Selection on codons of non-structural genes was observed to impact diversification of lineages in Asian I, cosmopolitan and sylvatic genotypes. Evidence of intra/inter-genotype recombination was obtained and the uncertainty in classification of recombinant strains was resolved using the population genetics approach. Discussion Complete genome-based analysis

  11. Detection of genotype recycling fraud in U.S. immigrants.

    Science.gov (United States)

    Wenk, Robert E

    2011-01-01

    Relationship testing laboratories provide genetic evidence to support or refute claims of kinship between U.S. citizen petitioners and potential immigrant beneficiaries. One female beneficiary presented a male amelogenin type and alleles at 15 autosomal loci that were identical to an alleged brother's. Laboratory records showed that her alleged father had petitioned to have 15 children emigrate from Ghana. The petitioner's 15 paternity indices exceeded 10⁵, but the children shared only four short tandem repeat (STR) profiles, suggesting fraudulent reuse of genotypes in this alleged pedigree (AP). To determine the extent of this "genotype recycling," I examined the laboratory's 555 APs from Ghana and 532 control APs from Nigeria. Seventeen Ghanaian APs (3.1%) but no Nigerian APs showed genotype recycling. Of 90 tested people in the 17 APs, 56 shared identical STR profiles with others in their AP. Of these 56 people, 10 were petitioners with unexpectedly high parentage indices. Seven of 56 had amelogenin types that disagreed with their declared genders. Database searches for identical multilocus genotypes in allegedly different people would best detect this fraud.

  12. Diversity of genotypes in CTX-M-producing Klebsiella pneumoniae isolated in different hospitals in Brazil

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    Thiago Pavoni Gomes Chagas

    2011-10-01

    Full Text Available OBJECTIVE: The present study was undertaken to characterize CTX-M ESBL-producing Klebsiella pneumoniae collected from hospitals in different cities of Brazil. MATERIAL AND METHODS: Eighty-five K. pneumoniae strains isolated from hospitalized patients in six different hospitals of three cities of Brazil were analyzed. ESBL production was confirmed by the standard double-disk synergy test and the Etest®. The MIC50 and MIC90 for ESBL-producing isolates were determined by the Etest® method. The antimicrobial susceptibilities of bacterial isolates were determined using the agar diffusion method according to the CLSI. Screening for blaTEM, blaSHV, blaCTX-M genes and class 1 integron was performed by PCR amplification. To determine the genomic diversity of CTX-M-producers, isolates were analyzed by macrorestriction profile analysis following PFGE. RESULTS AND DISCUSSION: Seventy-one K. pneumoniae isolates were ESBL-producing. PCR and sequencing experiments detected 38 CTX-M-producing K. pneumoniae belonged to groups CTX-M 1, CTX-M 2, CTX-M 8 and CTX-M 9. The association of different types ESBL (CTX-M, SHV and TEM was frequent. All K. pneumoniae isolates carried class 1 integron. PFGE analysis revealed thirty-one clonal types among CTX-M-producing isolates. The data presented herein illustrate the diversity of genotypes of CTX-M producing K. pneumoniae among Brazilians hospitals.

  13. Diversity of genotypes in CTX-M-producing Klebsiella pneumoniae isolated in different hospitals in Brazil

    Directory of Open Access Journals (Sweden)

    Thiago Pavoni Gomes Chagas

    Full Text Available OBJECTIVE: The present study was undertaken to characterize CTX-M ESBL-producing Klebsiella pneumoniae collected from hospitals in different cities of Brazil. MATERIAL AND METHODS: Eighty-five K. pneumoniae strains isolated from hospitalized patients in six different hospitals of three cities of Brazil were analyzed. ESBL production was confirmed by the standard double-disk synergy test and the Etest®. The MIC50 and MIC90 for ESBL-producing isolates were determined by the Etest® method. The antimicrobial susceptibilities of bacterial isolates were determined using the agar diffusion method according to the CLSI. Screening for blaTEM, blaSHV, blaCTX-M genes and class 1 integron was performed by PCR amplification. To determine the genomic diversity of CTX-M-producers, isolates were analyzed by macrorestriction profile analysis following PFGE. RESULTS AND DISCUSSION: Seventy-one K. pneumoniae isolates were ESBL-producing. PCR and sequencing experiments detected 38 CTX-M-producing K. pneumoniae belonged to groups CTX-M 1, CTX-M 2, CTX-M 8 and CTX-M 9. The association of different types ESBL (CTX-M, SHV and TEM was frequent. All K. pneumoniae isolates carried class 1 integron. PFGE analysis revealed thirty-one clonal types among CTX-M-producing isolates. The data presented herein illustrate the diversity of genotypes of CTX-M producing K. pneumoniae among Brazilians hospitals.

  14. Phenotypic and Genotypic Diversity of Thermophilic Campylobacter spp. in Commercial Turkey Flocks: A Longitudinal Study

    Science.gov (United States)

    Kashoma, Isaac P.; Kumar, Anand; Sanad, Yasser M.; Gebreyes, Wondwossen; Kazwala, Rudovick R.; Garabed, Rebecca

    2014-01-01

    Abstract Poultry are recognized as a main reservoir of Campylobacter spp. However, longitudinal studies investigating the persistence of Campylobacter on commercial meat turkeys are rare. The objectives of this study were to determine the prevalence, antimicrobial susceptibility, and persistence of genotypically related strains of Campylobacter spp. recovered from three commercial turkey farms in Ohio belonging to a single producer. Eight hundred ten samples were collected from birds aged 1 week to slaughter, consisting of 750 fecal droppings and 60 ceca at slaughter. Overall Campylobacter prevalence was 55.9%. Multiplex polymerase chain reaction (PCR) confirmed 72.3% of all isolates as C. coli, 5.3% as C. jejuni, 10.6% as both, and 11.9% as other Campylobacter spp. PCR restriction fragment length polymorphism of the flaA gene subtyping detected 70 types—62 for C. coli and 8 for C. jejuni isolates—with most (80%) of flaA-types constituting farm homogeneous groups. Multilocus sequence typing of 99 selected Campylobacter isolates resulted in 23 sequence types (STs), consisting of 8 STs for C. jejuni and 15 STs for C. coli isolates. Six novel STs—four for C. jejuni and two—for C. coli, were detected. In a subset of isolates (n=98) tested for antimicrobial resistance, the most common resistance was to tetracycline (95%), followed by azithromycin (43%), while 42% and 18% of the isolates were resistant to ciprofloxacin and erythromycin, respectively. All isolates were susceptible to florfenicol. C. coli isolates displayed a higher proportion of resistance than C. jejuni to most antimicrobials. This study highlights the high prevalence, genotypic diversity, and antimicrobial resistance of Campylobacter spp. in commercial turkey from farm to slaughter. PMID:25184688

  15. Type-specific PCR assays for Babesia bovis msa-1 genotypes in Asia: Revisiting the genetic diversity in Sri Lanka, Mongolia, and Vietnam.

    Science.gov (United States)

    Liyanagunawardena, Nilukshi; Sivakumar, Thillaiampalam; Kothalawala, Hemal; Silva, Seekkuge Susil Priyantha; Battsetseg, Badgar; Lan, Dinh Thi Bich; Inoue, Noboru; Igarashi, Ikuo; Yokoyama, Naoaki

    2016-01-01

    Babesia bovis is the most virulent Babesia organism, resulting in a high mortality rate in cattle. The genetic diversity of B. bovis merozoite surface antigens (MSAs), such as MSA-1, MSA-2b, and MSA-2c, might be linked to altered immune profiles in the host animals. The present study aimed to develop type-specific PCR assays for Asian msa-1 genotypes, thereby re-analyzing the genetic diversity of msa-1 in Sri Lanka, Mongolia, and Vietnam. Specific primers were designed for nine Asian msa-1 genotypes, which had been detected based on the phylogeny constructed using msa-1 gene sequences retrieved from the GenBank database. Specificity of the type-specific PCR assays was confirmed using plasmids containing the inserts of msa-1 gene fragments that represent Asian genotypes. Furthermore, no amplicons were observed by these PCR assays when DNA samples of Babesia bigemina, Babesia ovata, Theileria annulata, Theileria orientalis, Trypanosoma evansi, Trypanosoma theileri, Anaplasma marginale, and Anaplasma bovis, and non-infected bovine blood were analyzed. In total, 109 B. bovis-positive blood DNA samples sourced from Sri Lanka (44 cattle), Mongolia (26 cattle), and Vietnam (23 cattle and 16 water buffaloes) were then screened by the type-specific PCR assays. The sequences derived from all of the PCR amplicons were phylogenetically analyzed. Out of 109 DNA samples, 23 (20 from cattle and 3 from water buffaloes) were positive for at least one genotype. In agreement with previous studies, five and four different genotypes were detected among the DNA samples from Sri Lanka and Vietnam, respectively. In contrast, four genotypes, including three novel genotypes, were detected from Mongolia. Five DNA samples were found to be co-infected with multiple genotypes. The sequences of the PCR amplicons clustered phylogenetically within the corresponding clades. These findings indicated that the type-specific PCR assays described herein are useful for the determination of genotypic

  16. Detection, Diversity, and Population Dynamics of Waterborne Phytophthora ramorum Populations.

    Science.gov (United States)

    Eyre, C A; Garbelotto, M

    2015-01-01

    Sudden oak death, the tree disease caused by Phytophthora ramorum, has significant environmental and economic impacts on natural forests on the U.S. west coast, plantations in the United Kingdom, and in the worldwide nursery trade. Stream baiting is vital for monitoring and early detection of the pathogen in high-risk areas and is performed routinely; however, little is known about the nature of water-borne P. ramorum populations. Two drainages in an infested California forest were monitored intensively using stream-baiting for 2 years between 2009 and 2011. Pathogen presence was determined both by isolation and polymerase chain reaction (PCR) from symptomatic bait leaves. Isolates were analyzed using simple sequence repeats to study population dynamics and genetic structure through time. Isolation was successful primarily only during spring conditions, while PCR extended the period of pathogen detection to most of the year. Water populations were extremely diverse, and changed between seasons and years. A few abundant genotypes dominated the water during conditions considered optimal for aerial populations, and matched those dominant in aerial populations. Temporal patterns of genotypic diversification and evenness were identical among aerial, soil, and water populations, indicating that all three substrates are part of the same epidemiological cycle, strongly influenced by rainfall and sporulation on leaves. However, there was structuring between substrates, likely arising due to reduced selection pressure in the water. Additionally, water populations showed wholesale mixing of genotypes without the evident spatial autocorrelation present in leaf and soil populations.

  17. Serotype and genotype diversity and hatchery transmission of Campylobacter jejuni in commercial poultry flocks

    DEFF Research Database (Denmark)

    Petersen, L.; Nielsen, E.M.; On, Stephen L.W.

    2001-01-01

    We investigated the genotype and serotype diversity of Campylobacter coli and C jejuni in two parent flocks of adult hens and their offspring over two rotations in order to evaluate the role of hatchery mediated transmission and/or vertical transmission of campylobacters in broiler flocks. In tot...

  18. Genotyping and genetic diversity of Arcobacter butzleri by amplified fragment length polymorphism (AFLP) analysis

    DEFF Research Database (Denmark)

    On, Stephen L.W.; Atabay, H.I.; Amisu, K.O.

    2004-01-01

    Aims: To investigate the potential of amplified fragment length polymorphism (AFLP) profiling for genotyping Arcobacter butzleri and to obtain further data on the genetic diversity of this organism. Methods and Results: Seventy-three isolates of Danish, British, Turkish, Swedish, Nigerian and Nor...

  19. Influence of plant species on population dynamics, genotypic diversity and antibiotic production by indigenous Pseudomonas spp

    NARCIS (Netherlands)

    Bergsma-Vlami, M.; Prins, M.E.; Raaijmakers, J.M.

    2005-01-01

    The population dynamics, genotypic diversity and activity of naturally-occurring 2,4-diacetylphloroglucinol (DAPG)-producing Pseudomonas spp. was investigated for four plant species (wheat, sugar beet, potato, lily) grown in two different soils. All four plant species tested, except lily and in some

  20. A High-throughput Genomic Tool: Diversity Array Technology Complementary for Rice Genotyping

    Institute of Scientific and Technical Information of China (English)

    Yong Xie; Kenneth McNally; Cheng-Yun Li; Hei Leung; You-Yong Zhu

    2006-01-01

    Diversity array technology (DArTTM) was a genotyping tool characterized gel-independent and high throughput.The main purpose of present study is to validate DArT for rice (Oryza sativa L.)genotyping in a high throughput manner. Technically, the main objective was to generate a rice general purpose gene pool, and optimize this genomic tool in order to evaluate rice germplasm genetic diversity. To achieve this, firstly, a generalpurpose DArT array was developed. Ten representatives from 24 varieties were hybridized with the general-purpose array to determine the informativeness of the clones printed on the array. The informative 1 152 clones were re-arrayed on a slide and used to fingerprint 17 of 24 germplasms. Hybridizing targets prepared from the germplasm to be assayed to the DNA array gave DNA fingerprints of germplasms. Raw data were normalized and transformed into binary data, which were then analyzed by using NTSYSpc (Numerical taxonomy system for cluster and ordination analysis, v. 2.02j) software package. The graphically displayed dendrogram derived from the array experimental data was matched with simple Sequence repeats genotyping outline and varieties' pedigree deviation of the different varieties. Considering DArT is a sequence-independent genotyping approach, it will be applied in studies of the genetic diversity and the gene mapping of diverse of organisms, especially for those crops with less-developed molecular markers.

  1. Genetic Diversity Assessment and Identification of New Sour Cherry Genotypes Using Intersimple Sequence Repeat Markers

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    Roghayeh Najafzadeh

    2014-01-01

    Full Text Available Iran is one of the chief origins of subgenus Cerasus germplasm. In this study, the genetic variation of new Iranian sour cherries (which had such superior growth characteristics and fruit quality as to be considered for the introduction of new cultivars was investigated and identified using 23 intersimple sequence repeat (ISSR markers. Results indicated a high level of polymorphism of the genotypes based on these markers. According to these results, primers tested in this study specially ISSR-4, ISSR-6, ISSR-13, ISSR-14, ISSR-16, and ISSR-19 produced good and various levels of amplifications which can be effectively used in genetic studies of the sour cherry. The genetic similarity among genotypes showed a high diversity among the genotypes. Cluster analysis separated improved cultivars from promising Iranian genotypes, and the PCoA supported the cluster analysis results. Since the Iranian genotypes were superior to the improved cultivars and were separated from them in most groups, these genotypes can be considered as distinct genotypes for further evaluations in the framework of breeding programs and new cultivar identification in cherries. Results also confirmed that ISSR is a reliable DNA marker that can be used for exact genetic studies and in sour cherry breeding programs.

  2. Evaluation of genotypic diversity of Streptococcus mutans using distinct arbitrary primers

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    Cínthia Pereira Machado Tabchoury

    2008-12-01

    Full Text Available Streptococcus mutans has been considered one of the main etiological agents of dental caries and the genotypic diversity rather than its salivary counts may be considered as a virulence factor of this bacterium. For genotyping with polymerase chain reaction (PCR with arbitrary primers, several primers have been used in order to improve complexity and specificity of amplicon patterns. Thus, the aim of this study was to evaluate the degree of agreement of genotypic identification among AP-PCR reactions performed with 5 distinct arbitrary primers of S. mutans isolated from saliva. Stimulated saliva was collected from 11 adult volunteers for isolation of S. mutans, and a total of 88 isolates were genotyped with arbitrary primers OPA 02, 03, 05, 13 and 18. Fourteen distinct genotypes were identified in the saliva samples. Most volunteers (9 out of 11 presented only one genotype. The results of the present study suggest that primers OPA 02, 03, 05 and 13 were suitable for genotypic identification of S. mutans isolates of saliva from adult volunteers.

  3. Geographically diverse Australian isolates of Melissococcus pluton exhibit minimal genotypic diversity by restriction endonuclease analysis.

    Science.gov (United States)

    Djordjevic, S P; Smith, L A; Forbes, W A; Hornitzky, M A

    1999-04-15

    Melissococcus pluton, the causative agent of European foulbrood is an economically significant disease of honey bees (Apis mellifera) across most regions of the world and is prevalent throughout most states of Australia. 49 Isolates of M. pluton recovered from diseased colonies or honey samples in New South Wales, Queensland, South Australia, Tasmania and Victoria were compared using SDS-PAGE, Western immunoblotting and restriction endonuclease analyses. DNA profiles of all 49 geographically diverse isolates showed remarkably similar AluI profiles although four isolates (one each from Queensland, South Australia, New South Wales and Victoria) displayed minor profile variations compared to AluI patterns of all other isolates. DNA from a subset of the 49 Australian and three isolates from the United Kingdom were digested separately with the restriction endonucleases CfoI, RsaI and DraI. Restriction endonuclease fragment patterns generated using these enzymes were also similar although minor variations were noted. SDS-PAGE of whole cell proteins from 13 of the 49 isolates from different states of Australia, including the four isolates which displayed minor profile variations (AluI) produced indistinguishable patterns. Major immunoreactive proteins of approximate molecular masses of 21, 24, 28, 30, 36, 40, 44, 56, 60, 71, 79 and 95 kDa were observed in immunoblots of whole cell lysates of 22 of the 49 isolates and reacted with rabbit hyperimmune antibodies raised against M. pluton whole cells. Neither SDS-PAGE or immunoblotting was capable of distinguishing differences between geographically diverse isolates of M. pluton. Collectively these data confirm that Australian isolates of M. pluton are genetically homogeneous and that this species may be clonal. Plasmid DNA was not detected in whole cell DNA profiles of any isolate resolved using agarose gel electrophoresis.

  4. First insight into the genotypic diversity of clinical Mycobacterium tuberculosis isolates from Gansu Province, China.

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    Jie Liu

    Full Text Available BACKGROUND: Investigations of Mycobacterium tuberculosis genetic diversity in China have indicated a significant regional distribution. The aim of this study was to characterize the genotypes of clinical M. tuberculosis isolates obtained from Gansu, which has a special geographic location in China. METHODOLOGY/PRINCIPAL FINDINGS: A total of 467 clinical M. tuberculosis strains isolated in Gansu Province were genotyped by 15-locus mycobacterial interspersed repetitive units-variable number tandem repeats (MIRU-VNTR and spoligotyping. The results showed that 445 isolates belonged to six known spoligotype lineages, whereas 22 isolates were unknown. The Beijing genotype was the most prevalent (87.58%, n = 409, while the shared type 1 was the dominant genotype (80.94%, n = 378. The second most common lineage was the T lineage, with 25 isolates (5.35%, followed by the H lineage with 5 isolates (1.07%, the MANU family (0.64%, 3 isolates, the U family (0.43%, 2 isolates and the CAS lineage with 1 isolate (0.21%. By using the VNTR15China method, we observed 15 groups and 228 genotypes among the 467 isolates. We found no association between the five larger groups (including the Beijing genotype and sex, age, or treatment status, and there was no noticeable difference in the group analysis in different areas. In the present study, seven of the 15 MIRU-VNTR loci were highly or moderately discriminative according to their Hunter-Gaston discriminatory index. CONCLUSIONS/SIGNIFICANCE: The Beijing genotype is the predominant genotype in Gansu province. We confirm that VNTR15China is suitable for typing Beijing strains in China and that it has a better discriminatory power than spoligotyping. Therefore, the use of both methods is the most suitable for genotyping analysis of M. tuberculosis.

  5. Genetic diversity of Mycobacterium tuberculosis from Guadalajara, Mexico and identification of a rare multidrug resistant Beijing genotype.

    Science.gov (United States)

    Flores-Treviño, Samantha; Morfín-Otero, Rayo; Rodríguez-Noriega, Eduardo; González-Díaz, Esteban; Pérez-Gómez, Héctor R; Bocanegra-García, Virgilio; Vera-Cabrera, Lucio; Garza-González, Elvira

    2015-01-01

    Determining the genetic diversity of M. tuberculosis strains allows identification of the distinct Mycobacterium tuberculosis genotypes responsible for tuberculosis in different regions. Several studies have reported the genetic diversity of M. tuberculosis strains in Mexico, but little information is available from the state of Jalisco. Therefore, the aim of this study was to determine the genetic diversity of Mycobacterium tuberculosis clinical isolates from Western Mexico. Sixty-eight M. tuberculosis isolates were tested for susceptibility to first-line drugs using manual Mycobacteria Growth Indicator Tube method and genotyped using spoligotyping and IS6110-restriction fragment length polymorphism (RFLP) pattern analyses. Forty-seven (69.1%) isolates were grouped into 10 clusters and 21 isolates displayed single patterns by spoligotyping. Three of the 21 single patterns corresponded to orphan patterns in the SITVITWEB database, and 1 new type that contained 2 isolates was created. The most prevalent lineages were T (38.2%), Haarlem (17.7%), LAM (17.7%), X (7.4%), S (5.9%), EAI (1.5%) and Beijing (1.5%). Six (12.8%) of the clustered isolates were MDR, and type 406 of the Beijing family was among the MDR isolates. Seventeen (26.2%) isolates were grouped into 8 clusters and 48 isolates displayed single patterns by IS6110-RFLP. Combination of IS6110-RFLP and spoligotyping reduced the clustering rate to 20.0%. The results show that T, Haarlem, and LAM are predominant lineages among clinical isolates of M. tuberculosis in Guadalajara, Mexico. Clustering rates indicated low transmission of MDR strains. We detected a rare Beijing genotype, SIT406, which was a highly resistant strain. This is the first report of this Beijing genotype in Latin America.

  6. Diversity of Enterocytozoon bieneusi genotypes among small rodents in southwestern Poland.

    Science.gov (United States)

    Perec-Matysiak, Agnieszka; Buńkowska-Gawlik, Katarzyna; Kváč, Martin; Sak, Bohumil; Hildebrand, Joanna; Leśniańska, Kinga

    2015-12-15

    Diversity of Enterocytozoon bieneusi genotypes in wild small rodent populations still remains incomplete and only few molecular studies have been conducted among these hosts. Therefore, the aim of this study was to determine whether small rodents, i.e., Apodemus agrarius, Apodemus flavicollis, Mus musculus and Myodes glareolus act as hosts of E. bieneusi and can play an important role in spore spreading in the environment of south-western Poland. Molecular analyses were conducted to determine pathogen genotypes. A total of 191 fecal and 251 spleen samples collected from 311 rodent individuals were examined for the occurrence of E. bieneusi by PCR amplifying ITS gene. The overall prevalence of E. bieneusi in rodent samples was 38.9%. The nucleotide sequences of ITS region of E. bieneusi revealed the presence a total of 12 genotypes with two being already known, i.e., D and gorilla 1 genotypes. The remaining ten are novel genotypes (WR1-WR10) which segregated into three groups in a neighbor joining phylogeny. This study reports for the first time E. bieneusi occurrence in wild living rodents in Poland and shows extensive genetic diversity within E. bieneusi isolates of rodent origin.

  7. Emergence of new genotype and diversity of Theileria orientalis parasites from bovines in India.

    Science.gov (United States)

    George, Neena; Bhandari, Vasundhra; Reddy, D Peddi; Sharma, Paresh

    2015-12-01

    Bovine theileriosis is a serious threat to livestock worldwide. Uncertainty around species prevalence, antigenic diversity and genotypes of strains make it difficult to assess the impact of this parasite and to provide necessary treatment. We aimed to characterize genotypic diversity, phylogeny and prevalence of Theileria orientalis parasites from the states of Telangana and Andhra Pradesh, India by collecting bovine blood samples from the major districts of the two states. Bioinformatic analysis identified antigenic diversity among the prevalent parasite strains using major piroplasm surface protein (MPSP) gene. Our study revealed a prevalence rate of 4.8% (n=41/862) of T. orientalis parasites in bovine animals and a new genotype of T. orientalis parasite which was not previously reported in India. The emergence of these new genotypes could be an explanation for the frequent outbreaks of bovine theileriosis. Further, whole genome sequencing of T. orientalis strains will help to elucidate the genetic factors relevant for transmissibility and virulence as well as vaccine and new drug development.

  8. Genetic diversity analysis of rice (Oryza sativa genotypes for seedling characters under saline - alkaline condition

    Directory of Open Access Journals (Sweden)

    K Seetharam, S.Thirumeni, K.Paramasivam, S.Nadaradjan

    2013-03-01

    Full Text Available Rice is life for Asians as it provides 43 per cent calorie requirement for more than 70 per cent of the population. Theproduction is often limited by salinity. Understanding of physiological and genetic mechanisms is necessary for a breedingprogramme to improve crop performance under environmental stresses. Thirty rice genotypes pre-germinated in salinealkalinewater (pH-9.60; EC-10.0; SAR-54.32; RSC- 11.51 were placed in plastic cups filled with sterile soil and the stresswas imposed upto 21 days. Genetic diversity was estimated based on the observations recorded on germination per centage,vigor index, shoot length, root length, seedling length, root/shoot ratio, seeding dry weight, Na+/K+ ratio. The genotypeswere grouped in to five clusters based on the Euclidean coefficient which ranged between 2.09(CSR10 X CSR 13 and76.29 (IWP X Chettiviruppu. Cluster II was largest (22 genotypes followed by cluster I (4 genotypes. Genotypes groupedunder cluster I showed low Na+/K+ ratio which is an important physiological trait for salinity tolerance. Cluster V (MI 48 &IWP grouped the susceptible genotypes which had high Na+/K+ ratio. The hybrids thus developed from the genotypes ofcluster I & V may express high magnitude of transgressive segregants.

  9. Genetic Diversity Assessment Across Different Genotypes Of Mungbean And Urdbean Using Molecular Markers

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    Ashwini Narasimhan , B.R.Patil and S. Datta, M. Kaashyap

    2010-07-01

    Full Text Available Pulses compliment the daily diet of Indians along with cereals. They are rich in proteins with satisfactory proportion ofcarbohydrates. Mungbean, Vigna radiata and Urd bean, Vigna mungo are the important grain legume crops in agriculture,particularly in India. MYMV (Mungbean Yellow vein Mosaic Virus is a virus transmitted by whitefly, Bemesia tabaci, themost serious disease of Mungbean and Urdbean. In this study, six each of MYMV resistant and susceptible genotypes inMungbean and Urbean respectively were selected for the diversity analysis using molecular markers. Twenty four RGAprimers from cowpea were used to screen the twenty four genotypes. Dendrogram generated clearly indicated two bigclusters at 15% similarity. All mungbean genotypes made one cluster (cluster A except PS16, which was included in othercluster made by Urdbean genotypes (cluster B. Cluster A contained eleven genotypes while cluster B contained thirteengenotypes. Cluster A and B were further classified into two sub clusters namely A1 and A2, B1 and B2 respectively. A1consisted of seven genotypes of which five were resistant (PANT MUNG 1, PANT MUNG 5, HUM 12, PUSA 9531, HUM1 and two were susceptible (TARM 2, KOPERGAON 3, while A2 comprised of remaining four genotypes in which threewere susceptible (TAP 7, SML 134 and SML 668, and one (AKM 8803 was resistant. Further, it was found that fourmungbean resistant genotypes of A1 namely Pant Mung1, Pant Mung5, HUM 12, and PUSA 9531 made one cluster at 55%similarity. Cluster B, again was subdivided into B1 and B2. B1 consisted a single genotype which was a cross between IPU99-25* SPS5 while, B2 consisted of the rest of the twelve genotypes. It was interesting to see that two resistant (IPU 02-33and IPU 6-02 and two susceptible (LBG 20 and T9 genotypes of Urd bean made separate cluster with a similarity of 99 percent and which indicated that though genotypes are differing at resistant locus, they are highly similar at all other loci.

  10. HIV type 1 genetic diversity and genotypic drug susceptibility in the Republic of Moldova.

    Science.gov (United States)

    Pandrea, I; Descamps, D; Collin, G; Robertson, D L; Damond, F; Dimitrienco, V; Gheorghita, S; Pecec, M; Simon, F; Brun-Vézinet, F; Apetrei, C

    2001-09-01

    HIV-1 genetic diversity and, for the first time, genotypic drug susceptibility was investigated for strains circulating in the Republic of Moldova (of the former Soviet Union). Eighty-three samples from adults recently infected by intravenous drug use (IDU) (n = 60), heterosexual contact (n = 8), and from blood donors (n = 15) that tested positive from 1997 to 1998, and originating from different regions of Moldova were serotyped. By group-specific and subtype-specific peptide ELISA, patients were infected by serotype A (n = 65), serotype B (n = 1), or were nontypable (n = 17). Heteroduplex mobility assay (HMA) confirmed 11 subtype A and the one subtype B infection. Analyses of pol and env sequences for six of the IDUs confirmed that they were infected with subtype A strain. These strains clustered tightly with subtype A strains isolated from the former Soviet Union in phylogenetic analysis. No mutations associated with drug resistance were detected. The Republic of Moldova is culturally more closely related to Romania (where subtype F dominates the epidemic), but depends economically on Russia (where subtype A is established among IDUs). Thus, our results suggest that the spread of HIV in this region is driven by drug networks rather than being due to cultural similarities.

  11. Genotypic diversity and cariogenicity of Candida albicans from children with early childhood caries and caries-free children.

    Science.gov (United States)

    Qiu, Rongmin; Li, Wenqing; Lin, Yan; Yu, Dongsheng; Zhao, Wei

    2015-11-17

    The genotypic diversity and cariogenicity of C. albicans from the dental plaque of children are poorly understood. This study aimed to explore the genotypic diversity and cariogenicity of C. albicans from children with early childhood caries and caries-free children. Dental plaque samples from 238 children with early childhood caries and from 125 caries-free children were collected for C. albicans isolation. A PCR method based on 25S rDNA was used to analyze C. albicans genotypes, and the strains with different genotypes were tested with regard to acidogenicity and aciduricity. Among 129 C. albicans isolates, 79 (61.2 %) belonged to genotype A. The distribution frequency of genotypes A and C or genotypes B and C showed no significant difference between children with early childhood caries and caries-free children (p = 0.178 and 0.148), whereas genotypes A and B exhibited significantly different distributions (p = 0.010). No significant differences in aciduricity were found among the three genotypes, but the acidogenicity of genotypes B and C differed significantly from that of genotype A at pH 4.0. The genotypic distribution of C. albicans is associated with the caries experience of children, and the genotype may be related to its acidogenicity at pH 4.0.

  12. Distribution and diversity of hepatitis B virus genotypes in Yunnan, China.

    Science.gov (United States)

    Wang, Binghui; Feng, Yue; Li, Zheng; Duan, Haiping; Zhao, Ting; Zhang, Amei; Liu, Li; Baloch, Zulqarnain; Xia, Xueshan

    2014-10-01

    Hepatitis B virus (HBV) is one of the most prevalent pathogens in the world, and infection with this virus is a serious threat for public health. Yunnan is considered as an important endemic center for blood-borne viruses such as human immunodeficiency virus and hepatitis C virus, in China. However, the distribution and diversity of HBV subgenotypes remain unclear in Yunnan province. In the current study, HBV positive samples were collected from different prefectures of Yunnan province and their molecular epidemiological characters were determined. Phylogenetic analysis on the pre-S/S gene (865 bps) showed the prevalence of four HBV genotypes, including genotype B (24 cases, 33.3%), genotype C (45 cases, 62.5%), genotype I (two cases, 2.78%) and C/D recombinants (one case, 1.39%). The most prevalent genotypes B and C could be sub classified into subgenotype B2 and C1, C2, C5, and C7, respectively. Clusters of subgenotype B2 and C2 consisted of strains from China and other East Asian countries, while subgenotype C1, C5, and C7 and genotype I formed a cluster together with strains from Southeast Asia. Using Bayesian inference from phylogenetic, HBV genotypes B and C were estimated to have originated in 1860s and 1910s with an evolutionary rate of 3.26 and 8.01 × 10(-4) substitutions/site/year, respectively. These findings indicate that the distribution of HBV genotypes in Yunnan was influenced by strains from the rest of China and the neighboring countries.

  13. Genotypic diversity of Lactobacillus sanfranciscensis strains isolated from French organic sourdoughs.

    Science.gov (United States)

    Lhomme, Emilie; Onno, Bernard; Chuat, Victoria; Durand, Karine; Orain, Servane; Valence, Florence; Dousset, Xavier; Jacques, Marie-Agnès

    2016-06-01

    Lactobacillus sanfranciscensis is the predominant key lactic acid bacterium in traditionally fermented sourdoughs. Despite its prevalence, sourdough and their related breads could be different regarding their physicochemical and sensorial characteristics. The intraspecific diversity of L. sanfranciscensis might explain these observations. Fifty-nine strains isolated from French sourdoughs were typed by a polyphasic approach including Multilocus Sequence Typing (MLST) and Pulsed-field Gel Electrophoresis (PFGE), in order to study their genotypic diversity. MLST scheme can be reduced from six to four gene fragments (gdh, gyrA, nox and pta) without a major loss of discrimination between strains. The genes mapA and pgmA are not good candidates for inclusion in an MLST scheme to type L. sanfranciscensis strains, as they could not be amplified for a set of 18 strains among the 59 studied. This method revealed 20 sequence types (STs). Of these, 19 STs were grouped in one clonal complex, showing a strong relatedness between these strains. PFGE using SmaI discriminated 41 pulsotypes and so distinguished isolates better than the MLST scheme. Both genotypic methods indicate a low diversity between strains isolated from the same sourdough and a higher diversity between strains isolated from different sourdoughs, suggesting an influence of baker practices and/or environmental conditions on the selection of strains. The use of these two methods targeting genetic variations gives an optimal genotypic characterization of L.sanfranciscensis strains.

  14. Genotypic and phenotypic diversity in populations of plant-probiotic Pseudomonas spp. colonizing roots

    Science.gov (United States)

    Picard, Christine; Bosco, Marco

    2008-01-01

    Several soil microorganisms colonizing roots are known to naturally promote the health of plants by controlling a range of plant pathogens, including bacteria, fungi, and nematodes. The use of theses antagonistic microorganisms, recently named plant-probiotics, to control plant-pathogenic fungi is receiving increasing attention, as they may represent a sustainable alternative to chemical pesticides. Many years of research on plant-probiotic microorganisms (PPM) have indicated that fluorescent pseudomonads producing antimicrobial compounds are largely involved in the suppression of the most widespread soilborne pathogens. Phenotype and genotype analysis of plant-probiotic fluorescent pseudomonads (PFP) have shown considerable genetic variation among these types of strains. Such variability plays an important role in the rhizosphere competence and the biocontrol ability of PFP strains. Understanding the mechanisms by which genotypic and phenotypic diversity occurs in natural populations of PFP could be exploited to choose those agricultural practices which best exploit the indigenous PFP populations, or to isolate new plant-probiotic strains for using them as inoculants. A number of different methods have been used to study diversity within PFP populations. Because different resolutions of the existing microbial diversity can be revealed depending on the approach used, this review first describes the most important methods used for the assessment of fluorescent Pseudomonas diversity. Then, we focus on recent data relating how differences in genotypic and phenotypic diversity within PFP communities can be attributed to geographic location, climate, soil type, soil management regime, and interactions with other soil microorganisms and host plants. It becomes evident that plant-related parameters exert the strongest influence on the genotypic and phenotypic variations in PFP populations.

  15. Genetic diversity analysis among male and female Jojoba genotypes employing gene targeted molecular markers, start codon targeted (SCoT) polymorphism and CAAT box-derived polymorphism (CBDP) markers.

    Science.gov (United States)

    Heikrujam, Monika; Kumar, Jatin; Agrawal, Veena

    2015-09-01

    To detect genetic variations among different Simmondsia chinensis genotypes, two gene targeted markers, start codon targeted (SCoT) polymorphism and CAAT box-derived polymorphism (CBDP) were employed in terms of their informativeness and efficiency in analyzing genetic relationships among different genotypes. A total of 15 SCoT and 17 CBDP primers detected genetic polymorphism among 39 Jojoba genotypes (22 females and 17 males). Comparatively, CBDP markers proved to be more effective than SCoT markers in terms of percentage polymorphism as the former detecting an average of 53.4% and the latter as 49.4%. The Polymorphic information content (PIC) value and marker index (MI) of CBPD were 0.43 and 1.10, respectively which were higher than those of SCoT where the respective values of PIC and MI were 0.38 and 1.09. While comparing male and female genotype populations, the former showed higher variation in respect of polymorphic percentage and PIC, MI and Rp values over female populations. Nei's diversity (h) and Shannon index (I) were calculated for each genotype and found that the genotype "MS F" (in both markers) was highly diverse and genotypes "Q104 F" (SCoT) and "82-18 F" (CBDP) were least diverse among the female genotype populations. Among male genotypes, "32 M" (CBDP) and "MS M" (SCoT) revealed highest h and I values while "58-5 M" (both markers) was the least diverse. Jaccard's similarity co-efficient of SCoT markers ranged from 0.733 to 0.922 in female genotypes and 0.941 to 0.746 in male genotype population. Likewise, CBDP data analysis also revealed similarity ranging from 0.751 to 0.958 within female genotypes and 0.754 to 0.976 within male genotype populations thereby, indicating genetically diverse Jojoba population. Employing the NTSYS (Numerical taxonomy and multivariate analysis system) Version 2.1 software, both the markers generated dendrograms which revealed that all the Jojoba genotypes were clustered into two major groups, one group consisting of

  16. Genetic diversity analysis among male and female Jojoba genotypes employing gene targeted molecular markers, start codon targeted (SCoT polymorphism and CAAT box-derived polymorphism (CBDP markers

    Directory of Open Access Journals (Sweden)

    Monika Heikrujam

    2015-09-01

    Full Text Available To detect genetic variations among different Simmondsia chinensis genotypes, two gene targeted markers, start codon targeted (SCoT polymorphism and CAAT box-derived polymorphism (CBDP were employed in terms of their informativeness and efficiency in analyzing genetic relationships among different genotypes. A total of 15 SCoT and 17 CBDP primers detected genetic polymorphism among 39 Jojoba genotypes (22 females and 17 males. Comparatively, CBDP markers proved to be more effective than SCoT markers in terms of percentage polymorphism as the former detecting an average of 53.4% and the latter as 49.4%. The Polymorphic information content (PIC value and marker index (MI of CBPD were 0.43 and 1.10, respectively which were higher than those of SCoT where the respective values of PIC and MI were 0.38 and 1.09. While comparing male and female genotype populations, the former showed higher variation in respect of polymorphic percentage and PIC, MI and Rp values over female populations. Nei's diversity (h and Shannon index (I were calculated for each genotype and found that the genotype “MS F” (in both markers was highly diverse and genotypes “Q104 F” (SCoT and “82–18 F” (CBDP were least diverse among the female genotype populations. Among male genotypes, “32 M” (CBDP and “MS M” (SCoT revealed highest h and I values while “58-5 M” (both markers was the least diverse. Jaccard's similarity co-efficient of SCoT markers ranged from 0.733 to 0.922 in female genotypes and 0.941 to 0.746 in male genotype population. Likewise, CBDP data analysis also revealed similarity ranging from 0.751 to 0.958 within female genotypes and 0.754 to 0.976 within male genotype populations thereby, indicating genetically diverse Jojoba population. Employing the NTSYS (Numerical taxonomy and multivariate analysis system Version 2.1 software, both the markers generated dendrograms which revealed that all the Jojoba genotypes were clustered into two major

  17. Combined use of phenotypic and genotypic information in sampling animalsfor genotyping in detection of quantitative trait loci

    DEFF Research Database (Denmark)

    Ansari-Mahyari, S; Berg, P

    2008-01-01

    Conventional selective genotyping which is using the extreme phenotypes (EP) was compared with alternative criteria to find the most informative animals for genotyping with respects to mapping quantitative trait loci (QTL). Alternative sampling strategies were based on minimizing the sampling error...... of the estimated QTL effect (MinERR) and maximizing likelihood ratio test (MaxLRT) using both phenotypic and genotypic information. In comparison, animals were randomly genotyped either within or across families. One hundred data sets were simulated each with 30 half-sib families and 120 daughters per family....... The strategies were compared in these datasets with respect to estimated effect and position of a QTL within a previously defined genomic region at genotyping 10, 20 or 30% of the animals. Combined linkage disequilibrium linkage analysis (LDLA) was applied in a variance component approach. Power to detect QTL...

  18. Genotypic diversity of polyomaviruses circulating among kidney transplant recipients in Kuwait.

    Science.gov (United States)

    Chehadeh, Wassim; Nampoory, Mangalathillam Raman

    2013-09-01

    BK virus (BKV) and JC virus (JCV) are human polyomaviruses that cause asymptomatic latent infections. Under immunosuppression, BKV-associated nephropathy has been documented in Kuwait and elsewhere. Even though different BKV and JCV genotypes with distinct geographical distribution have been described, the genotype of polyomavirus detected in Kuwait is still unknown. The aim of this study was to determine the genotypes of BKV and JCV detected in renal transplant recipients. The detection of polyomavirus DNA was carried out in serum and urine samples of 200 post-transplant recipients during a 1-year follow-up period. Fifty-one (25.5%) post-transplant recipients were tested positive for polyomavirus DNA by semi-nested PCR. JCV DNA could be detected in 29 (57%) patients, and BKV DNA in 22 (43%) patients. In two renal transplant recipients, both BKV and JCV were detected. According to the Bayesian phylogenetic analysis of polyomavirus VP1 sequences, the majority of detected BKV sequences were most closely related to genotypes I and IV, whereas the majority of JCV sequences were most closely related to genotype 3. Polyomavirus VP1 sequences showed strong stability for up to 12 months in most patients; however, in one patient, an amino acid substitution in the BKV VP1 protein was identified over time. The results suggest a close relationship of BKV sequences with the Asian and European strains, and of JCV sequences with the African strains. Long follow-up studies are needed to investigate the association of polyomavirus polymorphism or genotypic shift with the development of nephropathy.

  19. Performance assessment of the GenoType MTBDRplus test and DNA sequencing in detection of multidrug-resistant Mycobacterium tuberculosis.

    Science.gov (United States)

    Huang, Wei-Lun; Chen, Huang-Yau; Kuo, Yuh-Min; Jou, Ruwen

    2009-08-01

    To facilitate the management of multidrug-resistant (MDR) tuberculosis, two nucleic acid sequence-based methods, the GenoType MTBDRplus test and DNA sequencing, were assessed for the rapid detection of drug-resistant Mycobacterium tuberculosis for the first time in the Asia-Pacific region. The performances of these two assays in detecting the presence of rifampin (rifampicin) (RIF) and isoniazid (INH) resistance-associated mutations in the rpoB, katG, inhA regulatory region, inhA, and oxyR-ahpC genes were compared to that of a conventional agar proportion drug susceptibility test. A total of 242 MDR and 30 pansusceptible M. tuberculosis isolates were evaluated in this study. The sensitivities obtained for RIF-resistant detection by the GenoType MTBDRplus test and by resistance gene sequencing were 95.5% and 97.9%, respectively. The sensitivities for INH resistance detection by the GenoType MTBDRplus test and by resistance gene sequencing were 81.8% and 93.4%, respectively. Together, the sensitivity for MDR tuberculosis detection was 78.5% with the GenoType MTBDRplus test and 91.3% by resistance gene sequencing. The specificity for RIF resistance, INH resistance, and MDR detection was 100% by both methods. The GenoType MTBDRplus test has the advantage of a short turnaround time for drug-resistant M. tuberculosis detection. Overall, the two assays performed equally well in detecting RIF resistance (P = 0.13). However, DNA sequencing demonstrated superior performance in detecting INH resistance (P tuberculosis (P GenoType MTBDRplus test, especially for different geographic areas with genetically diverse M. tuberculosis strains.

  20. Genotypic diversity and migration patterns of Phytophthora infestans in the Nordic countries.

    Science.gov (United States)

    Sjöholm, Lina; Andersson, Björn; Högberg, Nils; Widmark, Anna-Karin; Yuen, Jonathan

    2013-10-01

    In this study we investigated the genotypic diversity and the migration patterns of Phytophthora infestans in the Nordic countries. Isolates of P. infestans from outbreaks in 43 fields sampled in 2008 were collected using stratified sampling with country, field, and disease foci as the different strata. Microsatellites were used as markers to determine the genotypic variation in the sampled material. The results show a high genotypic variation of P. infestans in the Nordic countries with most of the genotypes found only once among the collected isolates. The major part of the genotypic variation was observed within the fields, with low differentiation between the fields. The observed low association of alleles among loci is consistent with frequent sexual reproduction of P. infestans in the Nordic countries. Coalescence analyses did not support a single common population for the four countries, thus indicating some degree of geographic differentiation. The analyses of migration patterns showed differing levels of gene flow among the Nordic countries. No correlation between migration rates and geographical distance could be seen. This could be explained by different degrees of genetic similarity between the pathogen populations in the different countries.

  1. Molecular diversity of brinjal (Solanum melongena L. and S. aethiopicum L. genotypes revealed by SSR markers

    Directory of Open Access Journals (Sweden)

    Abdul Majid Ansari, and Y. V. Singh

    2014-12-01

    Full Text Available In the present study, simple sequence repeat (SSR markers were used to study the genetic diversity among 14 genotypes of brinjal. A total of 14 polymorphic SSR primer pairs were used. Amplification of genomic DNA of 14 genotypes yielded 50 fragments, of which 43 were polymorphic. A clear cut differentiation was exhibited among the genotypes. The range of similarity coefficient varied from 17.8% [between S. aethiopicum L. (2n=2x=24 and Pant Rituraj (S. melongena L., 2n=2x=24] to 94.1% [between PB-71 and NDB-1] followed by 88.9% [between SMB-115 and KS-331] and 88.6% [between BARI and PB-67]. SAHN cluster analysis using UPGMA method separated the genotypes into six cluster groups. Solanum aethiopicum and PB-67 were positioned as single genotype in separate groups i.e., cluster-I & II, SMB-115 and KS-331 in cluster-III, BARI, PB-66 and Pant Rituraj in cluster-IV, WB-1, PB-4, PB-70 and LC-7 in cluster-V and PB-71, Pant Samrat and NDB-1 in cluster-VI. Morphological characters viz., shape, size and peel colour of brinjal fruits and plant type showed a positive relationship with the DNA based molecular analysis through SSR markers.

  2. GENETIC DIVERSITY OF S3 MAIZE GENOTYPES RESISTANT TO DOWNY MILDEW BASED ON SSR MARKERS

    Directory of Open Access Journals (Sweden)

    Amran Muis

    2016-02-01

    Full Text Available The compulsory requirement for releasing new high yielding maize varieties is resistance to downy mildew. The study aimed to determine the level of homozygosity, genetic diversity, and  genetic distance of 30 S3 genotypes of maize. Number of primers to be used were 30 polymorphic SSR loci which are distributed over the entire maize genomes. The S3 genotypes used were resistant to downy mildew with homozygosity level of >80%, genetic distance between the test and tester strains >0.7, and anthesis silking interval (ASI between inbred lines and tester lines was maximum 3 days. The results showed that 30 SSR primers used were spread evenly across the maize genomes which were manifested in the representation of SSR loci on each chromosome of a total of 10 chromosomes. The levels of polymorphism ranged from 0.13 to 0.78, an average of 0.51, and the number of alleles ranged from 2 to 8 alleles per SSR locus, an average of 4 alleles per SSR locus. The size of nucleotides in each locus also varied from 70 to 553 bp. Cophenetic correlation value (r at 0.67 indicated that the Unweighted Pair-Group Method Using Arithmetic Averages (UPGMA was less reliable for differentiating genotypes in five groups. Of the total of 30 genotypes analyzed, 17 genotypes had homozygosity level of >80% so it can be included in the hybrid assembly program.

  3. Real-time PCR assay for rapid detection of epidemiologically and clinically significant Mycobacterium tuberculosis Beijing genotype isolates.

    Science.gov (United States)

    Mokrousov, Igor; Vyazovaya, Anna; Zhuravlev, Viacheslav; Otten, Tatiana; Millet, Julie; Jiao, Wei-Wei; Shen, A-Dong; Rastogi, Nalin; Vishnevsky, Boris; Narvskaya, Olga

    2014-05-01

    Mycobacterium tuberculosis Beijing genotype strains are rapidly disseminating, frequently hypervirulent, and multidrug resistant. Here, we describe a method for their rapid detection by real-time PCR that targets the specific IS6110 insertion in the dnaA-dnaN genome region. The method was evaluated with a geographically and genetically diverse collection representing areas in East Asia and the former Soviet Union in which the Beijing genotype is endemic and epidemic (i.e., major foci of its global propagation) and with clinical specimens.

  4. Diversity of human astrovirus genotypes circulating in children with acute gastroenteritis in Thailand during 2000-2011.

    Science.gov (United States)

    Malasao, Rungnapa; Khamrin, Pattara; Chaimongkol, Natthawan; Ushijima, Hiroshi; Maneekarn, Niwat

    2012-11-01

    Human astrovirus (HAstV) is one of the causative agents of acute gastroenteritis in children worldwide. The objective of this study was to elucidate the molecular epidemiology and genotypic diversity of HAstV circulating in pediatric patients admitted to hospital with diarrhea in Thailand during the year 2000-2011, except for 2004, 2006, and 2009. A total of 1,022 fecal specimens were tested for HAstV by reverse transcription polymerase chain reaction (RT-PCR). HAstV was detected at 1.4% (14 of 1,022). All HAstV strains detected in this study were characterized further by nucleotide sequencing and phylogenetic analysis. Analysis of 348 bp partial capsid nucleotide sequences revealed that HAstV strains detected were HAstV-1 (1a, 1b, and 1d) (8 strains), HAstV-2 (2c) (3 strains), HAstV-3 (1 strain), and HAstV-5 (2 strains). HAstV-1, the most predominant genotype was detected initially in 2002 and circulated continuously up to 2011. HAstV-2 was detected in year 2001, and 2007 and grouped into a 2c lineage. HAstV-3 was found only in 2000 and HAstV-5 was found in the year 2001. The findings indicate that a wide variety of HAstV strains continue to circulate in children admitted to hospital with acute gastroenteritis in Thailand over a decade. The data provide an epidemiological overview of HAstV infection and HAstV genotype distribution in Thailand.

  5. DESCRIPTIVE AND DISCRIMINATORY SIGNIFICANCE OF POD PHENOTYPIC TRAITS FOR DIVERSITY ANALYSIS OF COCOA GENOTYPES

    Directory of Open Access Journals (Sweden)

    Daniel B. Adewale

    2013-12-01

    Full Text Available Intra-specific genetic diversity analysis precedes crop breeding proposal for species improvement. Sixteen and twenty-four parental and hybrid cocoa genotypes were respectively laid out in a randomized complete block design of six replications at Ibadan, Nigeria. A sampling unit of fifteen uniformly ripe pods was collected for assessment from each plot. Six quantitative data from the pods were subjected to statistical analysis. Highly significant (P < 0.0001 variability existed among the 40 genotypes. Range of performance of the genotypes were: pod weight (0.43 – 0.86kg, pod length (15.9 – 27.96cm, pod girth (21.51 – 34.07cm, pod thickness (1.26 – 5.71cm, number of beans per pod (20 - 51 and bean weight per pod (0.017 - 0.41kg. Positive and significant (P < 0.001 correlation existed between pod weight and length, pod girth and bean number/pod. The mean Gower genetic distance among the 40 genotypes was 0.228; the least (0.023 existed between G25 and G30 while the highest (0.529 was between G17 and G35. The first three principal component axes explained 73% of the total variation. Three distinct groups emerged from the Ward clustering technique. Significant (P<0.05 intra and inter cluster variability existed in the study. High genetic diversity lies within the studied population. Pod traits were important descriptors for cocoa genotypes classification.

  6. Analysis of population structure and genetic diversity of Egyptian and exotic rice (Oryza sativa L.) genotypes.

    Science.gov (United States)

    Salem, Khaled F M; Sallam, Ahmed

    2016-01-01

    Understanding the population structure and genetic diversity is a very important goal to improve the economic value of crops. In rice, a loss of genetic diversity in the last few centuries is observed. To address this challenge, a set of 22 lines from three different regions - India (two), and Philippines (six), and Egypt (14) - were used to assess the genetic diversity and the features of population structure. These genotypes were analyzed using 106 SSR alleles that showed a clear polymorphism among the lines. Genetic diversity was estimated based on the number of different alleles, polymorphism information content (PIC), and gene diversity. A total of 106 SSR alleles was identified from the 23 SSR loci and used to study the population structure and carry out a cluster analysis. All SSR loci showed a wide range of the number of different alleles extended from two (one loci) to seven alleles (three loci). Five and eight loci showed high PIC and gene diversity (≥0.70), respectively. The results of population structure are in agreement with cluster analysis results. Both analyses revealed two different subpopulations (G1 and G2) with different genetic properties in number of private alleles, number of different alleles (Na), number of effective alleles (Ne), expected heterozygosity (He), and Shannon's Information Index (SII). Our findings indicate that five SSR loci (RM 111, RM 307, RM 22, RM 19, and RM 271) could be used in breeding programs to enhance the marker-assisted selection through QTL mapping and association studies. A high genetic diversity found between genotypes which can be exploited to improve and produce rice cultivars for important traits (e.g. high agronomic features and tolerance to biotic or/and abiotic stresses).

  7. Detection and Genotyping of Leuconostoc spp. in a Sausage Processing Plant.

    Science.gov (United States)

    Padilla-Frausto, J J; Cepeda-Marquez, L G; Salgado, L M; Iturriaga, M H; Arvizu-Medrano, S M

    2015-12-01

    Some Leuconostoc spp. have the ability to produce slime and undesirable compounds in cooked sausage. The objectives of this research were to identify Leuconostoc sources in a Vienna-type sausage processing plant and to evaluate the genetic diversity of the isolated strains. Three hundred and two samples of sausage batter, sausages during processing, spoiled sausage, equipment surfaces, chilling brine, workers' gloves and aprons, and used casings were collected (March to November 2008 and February to April 2010) from a sausage processing plant. Lactic acid bacteria (LAB) were quantified, and Leuconostoc were detected using PCR. Strains were isolated and identified in Leuconostoc-positive samples. Leuconostoc strains were genotyped using randomly amplified polymorphic DNA and pulsed-field gel electrophoresis. LAB content of nonspoiled and spoiled sausage ranged from sausages showed the same genotype. One L. lactis genotype included strains isolated from spoiled sausages analyzed in April 2008 and March to April 2010. Equipment and conveyor belts constitute Leuconostoc contamination sources. Leuconostoc persistence in the sausage processing environment and in the final product suggests the existence of microbial reservoirs, possibly on equipment surfaces.

  8. High recombination frequency creates genotypic diversity in colonies of the leaf-cutting ant Acromyrmex echinatior.

    Science.gov (United States)

    Sirviö, A; Gadau, J; Rueppell, O; Lamatsch, D; Boomsma, J J; Pamilo, P; Page, R E

    2006-09-01

    Honeybees are known to have genetically diverse colonies because queens mate with many males and the recombination rate is extremely high. Genetic diversity among social insect workers has been hypothesized to improve general performance of large and complex colonies, but this idea has not been tested in other social insects. Here, we present a linkage map and an estimate of the recombination rate for Acromyrmex echinatior, a leaf-cutting ant that resembles the honeybee in having multiple mating of queens and colonies of approximately the same size. A map of 145 AFLP markers in 22 linkage groups yielded a total recombinational size of 2076 cM and an inferred recombination rate of 161 kb cM(-1) (or 6.2 cM Mb(-1)). This estimate is lower than in the honeybee but, as far as the mapping criteria can be compared, higher than in any other insect mapped so far. Earlier studies on A. echinatior have demonstrated that variation in division of labour and pathogen resistance has a genetic component and that genotypic diversity among workers may thus give colonies of this leaf-cutting ant a functional advantage. The present result is therefore consistent with the hypothesis that complex social life can select for an increased recombination rate through effects on genotypic diversity and colony performance.

  9. Positive effects of plant genotypic and species diversity on anti-herbivore defenses in a tropical tree species.

    Directory of Open Access Journals (Sweden)

    Xoaquín Moreira

    Full Text Available Despite increasing evidence that plant intra- and inter-specific diversity increases primary productivity, and that such effect may in turn cascade up to influence herbivores, there is little information about plant diversity effects on plant anti-herbivore defenses, the relative importance of different sources of plant diversity, and the mechanisms for such effects. For example, increased plant growth at high diversity may lead to reduced investment in defenses via growth-defense trade-offs. Alternatively, positive effects of plant diversity on plant growth may lead to increased herbivore abundance which in turn leads to a greater investment in plant defenses. The magnitude of trait variation underlying diversity effects is usually greater among species than among genotypes within a given species, so plant species diversity effects on resource use by producers as well as on higher trophic levels should be stronger than genotypic diversity effects. Here we compared the relative importance of plant genotypic and species diversity on anti-herbivore defenses and whether such effects are mediated indirectly via diversity effects on plant growth and/or herbivore damage. To this end, we performed a large-scale field experiment where we manipulated genotypic diversity of big-leaf mahogany (Swietenia macrophylla and tree species diversity, and measured effects on mahogany growth, damage by the stem-boring specialist caterpillar Hypsipyla grandella, and defensive traits (polyphenolics and condensed tannins in stem and leaves. We found that both forms of plant diversity had positive effects on stem (but not leaf defenses. However, neither source of diversity influenced mahogany growth, and diversity effects on defenses were not mediated by either growth-defense trade-offs or changes in stem-borer damage. Although the mechanism(s of diversity effects on plant defenses are yet to be determined, our study is one of the few to test for and show producer

  10. Genetic diversity analysis among pigeonpea genotypes adapted to South American regions based on microsatellite markers

    Directory of Open Access Journals (Sweden)

    Adna Cristina Barbosa de Sousa

    2011-08-01

    Full Text Available The pigeonpea [Cajanus cajan (L Millspaugh] is one of the most important perennial legume crops utilized in the food, fodder, soil conservation, crop-livestock integrated systems, reclaiming of degraded pastures and symbiotic nitrogen fixation. Microsatellite markers were used to estimate the genetic diversity of 77 pigeonpea genotypes selected from the germplasm collections at Embrapa Cattle-Southeast and, to evaluate their transferability to Phaseolus vulgaris and Vigna unguiculata species. The number of alleles per locus ranged from 2 to12, with an average of 5.1 alleles. The PIC values ranged from 0.11 to 0.80 (average 0.49 and the D values from 0.23 to 0.91 (average 0.58. The averages of observed and expected heterozygosity were 0.25 and 0.47, respectively, showing a deficit in heterozygosity. A model-based Bayesian approach implemented in the software STRUCTURE was used to assign genotypes into clusters. A dendrogram was constructed based on the modified Roger's genetic distances using a neighbor-joining method (NJ. A total of four clusters were assembled by STRUCTURE and a strong tendency of correspondence between the Bayesian clusters in the NJ tree was observed. The genetic distance ranged from 0.09 to 0.62 (average 0.37, showing a low genetic diversity in the pigeonpea genotypes. Transferability of pigeonpea-specific microsatellites revealed a cross-amplification and the presence of polymorphic alleles in P. vulgaris and V. unguiculata.

  11. Diversity analysis in Indian genotypes of linseed (Linum usitatissimum L.) using AFLP markers.

    Science.gov (United States)

    Chandrawati; Maurya, Ramanuj; Singh, P K; Ranade, S A; Yadav, Hemant Kumar

    2014-10-01

    AFLP fingerprinting of 45 Indian genotypes of linseed was carried out to determine the genetic relationship among them. Sixteen primer combinations produced 1142 fragments with 1129 as polymorphic and 13 as monomorphic fragments. Polymorphic fragments varied from 44 (E-ACA/M-CTA) to 94 (E-AGC/M-CAC) with an average of 70.6 fragments per primer combination. The frequency of polymorphism varied from 93.7% to 100% with an average of 98.8% across all the genotypes. The PIC value ranged from 0.19 to 0.31 with an average of 0.23 per primer combination. The primer pair E-AGC/M-CAC showed the maximum PIC value (0.31) followed by E-AGC/M-CAG (0.29), E-AAC/M-CAG (0.26) and E-AGC/M-CTA (0.25). Resolving power (RP) and marker index (MI) varied from 13.73 to 43.50 and 8.81 to 28.91 respectively. The Jaccard's similarity coefficient varied from 0.16 to 0.57 with an average of 0.26 ± 0.05. The maximum genetic similarities (57%) were detected between genotypes Him Alsi-1 and Him Alsi-2, followed by Him Alsi-1 and GS41 and GS41 and LC-54. The genotypes R-552, Himani, RKY-14, Meera, Indira Alsi-32 and Suyog were found to be more divergent genotypes. The NJ clustering grouped all the 45 genotypes into three major clusters. In general the genotypes of cluster III had high oil content and those of cluster I had low oil content. At the population level, within population variance was much higher than between populations variance. Copyright © 2014 Elsevier B.V. All rights reserved.

  12. Genetic Diversity Among Historical Olive (Olea europaea L.) Genotypes from Southern Anatolia Based on SSR Markers.

    Science.gov (United States)

    Sakar, Ebru; Unver, Hulya; Ercisli, Sezai

    2016-12-01

    Olive (Olea europaea) is an ancient and important crop in both olive oil production and table use. It is important to identify the genetic diversity of olive genetic resources for cultivar development and evaluation of olive germplasm. In the study, 14 microsatellite markers (UDO4, UDO8, UDO9, UDO11, UDO12, UDO22, UDO24, UDO26, UDO28, DCA9, DCA11, DCA13, DCA15, and DCA18) were used to assess the genetic variation on 76 olive (Olea europaea L.) genotypes from Mardin province together with 6 well-known Turkish and 4 well-known foreign reference cultivars. All microsatellite markers showed polymorphism and the number of alleles varied between 9 and 22, with an average of 14.57. The most informative loci were DCA 11 (22 alleles) and DCA 9 (21 alleles). Dendrogram based on genetic distances was constructed for the 86 olive genotypes/cultivars, which revealed the existence of different clusters. The high genetic similarity was evident between Bakırkire2 and Zinnar5 (0.74) genotypes, while the most genetically divergent genotypes were Gürmeşe5 and Yedikardeşler2 (0.19). It was concluded that there was abundant SSR polymorphism in olive germplasm in southern Anatolia in Turkey and could be important for future breeding activities.

  13. Human population structure detection via multilocus genotype clustering

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    Starmer Joshua

    2007-06-01

    Full Text Available Abstract Background We describe a hierarchical clustering algorithm for using Single Nucleotide Polymorphism (SNP genetic data to assign individuals to populations. The method does not assume Hardy-Weinberg equilibrium and linkage equilibrium among loci in sample population individuals. Results We show that the algorithm can assign sample individuals highly accurately to their corresponding ethnic groups in our tests using HapMap SNP data and it is also robust to admixed populations when tested with Perlegen SNP data. Moreover, it can detect fine-scale population structure as subtle as that between Chinese and Japanese by using genome-wide high-diversity SNP loci. Conclusion The algorithm provides an alternative approach to the popular STRUCTURE program, especially for fine-scale population structure detection in genome-wide association studies. This is the first successful separation of Chinese and Japanese samples using random SNP loci with high statistical support.

  14. Genotypic Diversity and Virulence Traits of Streptococcus mutans Isolated from Carious Dentin after Partial Caries Removal and Sealing

    Science.gov (United States)

    Damé-Teixeira, Nailê; Arthur, Rodrigo Alex; Parolo, Clarissa Cavalcanti Fatturi

    2014-01-01

    The aim of this study was to compare the genotypic diversity and virulence traits of Streptococcus mutans isolated from carious dentin before and after partial dentin caries removal (PDR) and sealing. Carious dentin samples were obtained three months before and after the PDR and cavity sealing. Up to seven isolates of each morphological type of S. mutans were selected and strain identity was confirmed using gtfB primer. Genotyping was performed by arbitrary primer-PCR (AP-PCR). Acidogenesis and acidurance of the genotypes were evaluated as virulence traits. A paired t-test and a Wilcoxon test were used to compare the virulence of genotypes. A total of 48 representative S. mutans isolates were genotyped (31 before and 17 after the sealing). At least one of the genotypes found before the sealing was also found on dentin after the sealing. The number of genotypes found before the sealing ranged from 2 to 3 and after the sealing from 1 to 2 genotypes. No difference was observed in the acidogenesis and acidurance between genotypes isolated before and after the sealing. In conclusion, genotypic diversity of S. mutans decreased after the PDR and sealing, but the virulence traits of S. mutans remained unchangeable. PMID:24578618

  15. Specificity of the Linear Array HPV Genotyping Test for detecting human papillomavirus genotype 52 (HPV-52)

    OpenAIRE

    Kocjan, Boštjan; Poljak, Mario; Oštrbenk, Anja

    2015-01-01

    Introduction: HPV-52 is one of the most frequent human papillomavirus (HPV) genotypes causing significant cervical pathology. The most widely used HPV genotyping assay, the Roche Linear Array HPV Genotyping Test (Linear Array), is unable to identify HPV- 52 status in samples containing HPV-33, HPV-35, and/or HPV-58. Methods: Linear Array HPV-52 analytical specificity was established by testing 100 specimens reactive with the Linear Array HPV- 33/35/52/58 cross-reactive probe, but not with the...

  16. Detection and genotyping of enteroviruses in cerebrospinal fluid in patients in Victoria, Australia, 2007-2013.

    Science.gov (United States)

    Papadakis, Georgina; Chibo, Doris; Druce, Julian; Catton, Michael; Birch, Chris

    2014-09-01

    Genotyping by VP1 fragment polymerase chain reaction (PCR) and nucleic acid sequencing to detect enterovirus (EV) genotypes was performed directly on 729 EV PCR positive cerebrospinal fluid (CSF) samples collected between 2007 and 2012 from Victorian hospital inpatients. The overall genotype identification rate from CSF-positive material was 43%. The four most common genotypes identified were Echovirus 6 (24%), Echovirus 30 (17%), Echovirus 25 (10%), and Coxsackievirus A9 (10%), together comprising 61% of all EVs typed. The seasonal distribution of all EVs identified followed the recognized pattern of mainly summer epidemics. Three of the four predominant genotypes were present in each of the 6 years in which the study was conducted, with 20 other EV genotypes also detected, often in only a single year. Genotyping of EVs directly in CSF is faster, simpler and more sensitive than traditional virus neutralization assays performed on EV positive samples. © 2014 Wiley Periodicals, Inc.

  17. Circulation of Coxiella burnetii in a Naturally Infected Flock of Dairy Sheep: Shedding Dynamics, Environmental Contamination, and Genotype Diversity.

    Science.gov (United States)

    Joulié, A; Laroucau, K; Bailly, X; Prigent, M; Gasqui, P; Lepetitcolin, E; Blanchard, B; Rousset, E; Sidi-Boumedine, K; Jourdain, E

    2015-10-01

    Q fever is a worldwide zoonosis caused by Coxiella burnetii. Domestic ruminants are considered to be the main reservoir. Sheep, in particular, may frequently cause outbreaks in humans. Because within-flock circulation data are essential to implementing optimal management strategies, we performed a follow-up study of a naturally infected flock of dairy sheep. We aimed to (i) describe C. burnetii shedding dynamics by sampling vaginal mucus, feces, and milk, (ii) assess circulating strain diversity, and (iii) quantify barn environmental contamination. For 8 months, we sampled vaginal mucus and feces every 3 weeks from aborting and nonaborting ewes (n=11 and n=26, respectively); for lactating females, milk was obtained as well. We also sampled vaginal mucus from nine ewe lambs. Dust and air samples were collected every 3 and 6 weeks, respectively. All samples were screened using real-time PCR, and strongly positive samples were further analyzed using quantitative PCR. Vaginal and fecal samples with sufficient bacterial burdens were then genotyped by multiple-locus variable-number tandem-repeat analysis (MLVA) using 17 markers. C. burnetii burdens were higher in vaginal mucus and feces than in milk, and they peaked in the first 3 weeks postabortion or postpartum. Primiparous females and aborting females tended to shed C. burnetii longer and have higher bacterial burdens than nonaborting and multiparous females. Six genotype clusters were identified; they were independent of abortion status, and within-individual genotype diversity was observed. C. burnetii was also detected in air and dust samples. Further studies should determine whether the within-flock circulation dynamics observed here are generalizable.

  18. Helicobacter pylori cag pathogenicity island genotype diversity within the gastric niche of a single host.

    Science.gov (United States)

    Matteo, Mario José; Granados, Gabriela; Pérez, Cecilia Valeria; Olmos, Martín; Sanchez, Cristian; Catalano, Mariana

    2007-05-01

    cag pathogenicity island (PAI) integrity was investigated in isolates from multiple biopsies recovered from 40 patients in an attempt to determine the co-existence of a varying cagPAI-positive to cagPAI-negative ratio in a single host. Six biopsies were obtained from each patient during the same endoscopic session. cagPAI analysis included amplification of seven loci (cagA, cagE, cagG, cagM, cagT, HP0527 and HP0524) and the left end of cagII (LEC). Absence of the island was confirmed by empty-site PCR. lspA-glmM RFLP and random amplified polymorphic DNA PCR were used for strain delineation. The number of biopsies with Helicobacter pylori-positive culture ranged from three to six per patient and a total of 218 isolates were recovered. Mixed infection was only found in two patients. Nearly one-third of the 40 patients harboured isolates with an intact cagPAI in all niches, another third of the isolates were empty-site-positive in all niches, whilst the remaining third of the isolates had a disrupted cagPAI in all or at least one of the niches. Co-existence of variants of the same strain with different cagPAI genotypes was observed in one-quarter of patients. The variations in cagPAI genotype included co-existence of: diverse cagPAI deletions in different niches, variants with intact and with partially deleted islands, variants with empty-site-positive and with partially deleted cagPAIs, and variants with an intact cagPAI and with empty-site-positive. Half of the patients with different cagPAI genotypes harboured an intact cagPAI in at least one niche. Co-existence of diverse genotypes of putative virulence factors in a single host must be considered when drawing a correlation with clinical presentation.

  19. Genotypic and phenotypic diversity of the noncapsulated Haemophilus influenzae: adaptation and pathogenesis in the human airways.

    Science.gov (United States)

    Garmendia, Junkal; Martí-Lliteras, Pau; Moleres, Javier; Puig, Carmen; Bengoechea, José A

    2012-12-01

    The human respiratory tract contains a highly adapted microbiota including commensal and opportunistic pathogens. Noncapsulated or nontypable Haemophilus influenzae (NTHi) is a human-restricted member of the normal airway microbiota in healthy carriers and an opportunistic pathogen in immunocompromised individuals. The duality of NTHi as a colonizer and as a symptomatic infectious agent is closely related to its adaptation to the host, which in turn greatly relies on the genetic plasticity of the bacterium and is facilitated by its condition as a natural competent. The variable genotype of NTHi accounts for its heterogeneous gene expression and variable phenotype, leading to differential host-pathogen interplay among isolates. Here we review our current knowledge of NTHi diversity in terms of genotype, gene expression, antigenic variation, and the phenotypes associated with colonization and pathogenesis. The potential benefits of NTHi diversity studies discussed herein include the unraveling of pathogenicity clues, the generation of tools to predict virulence from genomic data, and the exploitation of a unique natural system for the continuous monitoring of long-term bacterial evolution in human airways exposed to noxious agents. Finally, we highlight the challenge of monitoring both the pathogen and the host in longitudinal studies, and of applying comparative genomics to clarify the meaning of the vast NTHi genetic diversity and its translation to virulence phenotypes.

  20. [Investigation on HPV DNA detection and genotyping practices used in French laboratories in 2009].

    Science.gov (United States)

    Heard, Isabelle; Favre, Michel; Fihman, Valentine

    2011-01-01

    The French National reference Laboratory for Human papillomavirus (HPV) performed in 2009 a national study in order to review the methods used to detect and identify HR HPV genotypes in microbiology laboratories. Results from this study show a great diversity in volumes of samples treated in laboratories. Among clinical indications, the most frequent is a result of ASC-US at a Pap smear. This indication in the only one covered by the National Public Insurance System and is mostly performed in laboratories from private sector. Other indications mainly correspond to research programs and are performed in public Hospitals. This study allowed also to review the adequacy between the liquid based cytology samples and the assays used for direct detection of HR HPV or identification of the genotypes present in the sample. The right tests were not carried in the right solution storage according to the recommendations from different HPV testing assays. National recommendations should be elaborated in order to improve the performance of the test used.

  1. A study of Mycobacterium tuberculosis genotypic diversity & drug resistance mutations in Varanasi, north India

    Directory of Open Access Journals (Sweden)

    Anamika Gupta

    2014-01-01

    Full Text Available Background & objectives: One-fifth of the world′s new tuberculosis (TB cases and two-thirds of cases in the South East Asian region occur in India. Molecular typing of Mycobacterium tuberculosis isolates has greatly facilitated to understand the transmission of TB. This study was aimed to investigate the molecular epidemiology of M. tuberculosis genotypes in Varanasi, north India, and their association with clinical presentation among patients with pulmonary TB. Methods: M. tuberculosis isolates from 104 TB patients attending a tertiary referral hospital of north India were screened for susceptibility to isoniazid (INH, rifampicin (RIF, ethambutol (EMB and streptomycin (STR by proportion method and multiplex-allele-specific-polymerase chain reaction (MAS-PCR. These were genotyped by spoligotyping. The spoligotype patterns were compared with those in the international SITVIT2 spoligotyping database. Results: Eighty three of 104 isolates were distributed in 38 SITs, of which SIT3366 was newly created within the present study. The mass of ongoing transmission with MDR-TB isolates in Varanasi, northern India, was linked to Beijing genotype followed by the CAS1_Delhi lineage. HIV-seropositive patients had a significantly higher proportion of clustered isolates than HIV-seronegative patients and compared with the wild type(wt isolates, the isolates with katG315Thr mutation were considerably more likely to be clustered. Interpretation & conclusions: This study gives an insight into the M. tuberculosis genetic biodiversity in Varanasi, north India, the predominant spoligotypes and their impact on disease transmission. In this region of north India, TB is caused by a wide diversity of spoligotypes with predominance of four genotype lineages: Beijing, CAS, EAI and T. The Beijing genotype was the most frequent single spoligotype and strongly associated with multi drug resistant (MDR-TB isolates. These findings may have important implications for control

  2. Genotypic Diversity Is Associated with Clinical Outcome and Phenotype in Cryptococcal Meningitis across Southern Africa.

    Science.gov (United States)

    Beale, Mathew A; Sabiiti, Wilber; Robertson, Emma J; Fuentes-Cabrejo, Karen M; O'Hanlon, Simon J; Jarvis, Joseph N; Loyse, Angela; Meintjes, Graeme; Harrison, Thomas S; May, Robin C; Fisher, Matthew C; Bicanic, Tihana

    2015-01-01

    Cryptococcal meningitis is a major cause of mortality throughout the developing world, yet little is known about the genetic markers underlying Cryptococcal virulence and patient outcome. We studied a cohort of 230 Cryptococcus neoformans (Cn) isolates from HIV-positive South African clinical trial patients with detailed clinical follow-up using multi-locus sequence typing and in vitro phenotypic virulence assays, correlating these data with clinical and fungal markers of disease in the patient. South African Cn displayed high levels of genetic diversity and locus variability compared to globally distributed types, and we identified 50 sequence types grouped within the main molecular types VNI, VNII and VNB, with 72% of isolates typed into one of seven 'high frequency' sequence types. Spatial analysis of patients' cryptococcal genotype was not shown to be clustered geographically, which might argue against recent local acquisition and in favour of reactivation of latent infection. Through comparison of MLST genotyping data with clinical parameters, we found a relationship between genetic lineage and clinical outcome, with patients infected with the VNB lineage having significantly worse survival (n=8, HR 3.35, CI 1.51-7.20, p=0.003), and this was maintained even after adjustment for known prognostic indicators and treatment regimen. Comparison of fungal genotype with in vitro phenotype (phagocytosis, laccase activity and CSF survival) performed on a subset of 89 isolates revealed evidence of lineage-associated virulence phenotype, with the VNII lineage displaying increased laccase activity (p=0.001) and ex vivo CSF survival (p=0.0001). These findings show that Cryptococcus neoformans is a phenotypically heterogeneous pathogen, and that lineage plays an important role in cryptococcal virulence during human infection. Furthermore, a detailed understanding of the genetic diversity in Southern Africa will support further investigation into how genetic diversity is

  3. Evaluation of genotypic variation in leaf photosynthetic rate and its associated factors by using rice diversity research set of germplasm.

    Science.gov (United States)

    Kanemura, Tomomi; Homma, Koki; Ohsumi, Akihiro; Shiraiwa, Tatsuhiko; Horie, Takeshi

    2007-10-01

    In order to evaluate genotypic variation, we measured leaf photosynthetic rate (Pn) and its associated factors for the rice diversity research set of germplasm (RDRS) selected from the Genebank in National Institute of Agrobiological Sciences (NIAS). Pn showed large genotypic variation from 11.9 to 32.1 micromol m(-2 )s(-1). The variation in stomatal conductance to CO2 (Gs) explained about 50% of that in Pn, while that in nitrogen concentration (N) in leaves explained about 35%. The genotype group which mainly consists of aus type indica tended to have higher Gs, and the genotype group which corresponds to japonica had a higher nitrogen concentration (N) in leaves. The relationships of Pn with Gs and N were not significantly different among genotype groups, suggesting photosynthetic efficiencies are similar among genotype groups.

  4. Genetic diversity among Juglans regia L. genotypes assessed by morphological traits and microsatellite markers

    Energy Technology Data Exchange (ETDEWEB)

    Mahmoodi, R.; Rahmani, F.; Rezaee, R.

    2013-06-01

    In this study, genetic diversity was assayed among 16 accessions and five cultivars of Persian walnut (Juglans regia L.) using morphological traits and nine simple sequence repeat (SSR) markers. Samples were collected from Agriculture Research Center of Urmia city (North West Iran). Study on important morphological traits revealed genetic similarity of -0.6 to 0.99 based on CORR coefficient. The microsatellite marker system produced 34 alleles in range of 160-290 bp. The minimum (2) and maximum (7) number of alleles were obtained from WGA71 and WGA202 genetic loci, respectively. The mean number of alleles per locus was 4.25. Jaccards similarity coefficient ranged from 0.13 to 0.76. The results of this paper indicate high diversity among these genotypes which could be used for breeding management. (Author) 28 refs.

  5. Genetic diversity among Juglans regia L. genotypes assessed by morphological traits and microsatellite markers

    Directory of Open Access Journals (Sweden)

    R. Mahmoodi

    2013-05-01

    Full Text Available In this study, genetic diversity was assayed among 16 accessions and five cultivars of Persian walnut (Juglans regia L. using morphological traits and nine simple sequence repeat (SSR markers. Samples were collected from Agriculture Research Center of Urmia city (North West Iran. Study on important morphological traits revealed genetic similarity of -0.6 to 0.99 based on CORR coefficient. The microsatellite marker system produced 34 alleles in range of 160-290 bp. The minimum (2 and maximum (7 number of alleles were obtained from WGA71 and WGA202 genetic loci, respectively. The mean number of alleles per locus was 4.25. Jaccard’s similarity coefficient ranged from 0.13 to 0.76. The results of this paper indicate high diversity among these genotypes which could be used for breeding management.

  6. High diversity of hepatitis B virus genotypes in Panamanian blood donors: a molecular analysis of new variants.

    Directory of Open Access Journals (Sweden)

    Alexander A Martínez

    Full Text Available Hepatitis B Virus (HBV is an infectious agent that causes more than half of the cases of liver disease and cancer in the world. Globally there are around 250 million people chronically infected with this virus. Despite 16% of the cases of liver disease in Central America are caused by HBV, the information regarding its genetic diversity, genotypes and circulation is scarce. The purpose of this study was to evaluate the genetic variability of the HBV genotypes from HBV-DNA positive samples obtained from screening blood donors at the Social Security System of Panama and to estimate its possible origin. From 59,696 blood donors tested for HBV infection during 2010-2012, there were 74 HBV-DNA positive subjects. Analysis of the partial PreS2-S region of 27 sequences shows that 21% of the infections were caused by genotype A, 3% by genotype D and 76% by genotype F. In addition, we were able to confirm circulation of six sub-genotypes A1, A2, A3, D4, F3, F1 and a proposed new sub-genotype denominated F5pan. We found a confinement of sub-genotypes F1 and F5pan to the western area of Panama. The tMRCA analysis suggests a simultaneous circulation of previously described sub-genotypes rather than recent introductions of the Panamanian sub-genotypes in the country. Moreover, these results highlight the need of more intensive research of the HBV strains circulating in the region at the molecular level. In conclusion, Panama has a high HBV genotype diversity that includes a new proposed sub-genotype, an elevated number of PreCore-Core mutations, and confinement of these variants in a specific geographical location.

  7. Molecular diversity of native rhizobia trapped by five field pea genotypes in Indian soils.

    Science.gov (United States)

    Wadhwa, K; Dudeja, S S; Yadav, R K

    2011-02-01

    Five pea cultivars; HFP 4, HVP 3-5, HFP 9426, Jayanti and Hariyal, being grown in CCS Haryana Agricultural University farm were used to isolate native rhizobia. Selected 54 rhizobia, from all cultivars, were authenticated as rhizobia by plant infectivity test. Along with nodulation, symbiotic effectiveness in terms of symbiotic ratios showed wide range of effectiveness of pea rhizobia from 1.11 to 5.0. DNA of all the 54 rhizobia was extracted and amplified by PCR, using ERIC and 16S rDNA primers. Dendrogram based on ERIC profiles of these 54 rhizobia showed the formation of 13 subclusters at 80% level of similarity. Dendrogram based on RFLP of 16S rDNA by three restriction endonucleases; Msp I, Csp 6I and Rsa I; also formed 13 subclusters at 80% level of similarity. However, positioning of subclusters was different from that of ERIC based dendrogram. Majority of the isolates i.e. 64.8% by ERIC profiles and 44.4% by RFLP of 16S rDNA formed one cluster. Isolates from same nodule were not 100% similar. Considering each cluster representing a rhizobial genotype, both techniques used to assess molecular diversity indicated the presence of 13 genotypes of field pea rhizobia in CCS Haryana Agricultural University farm soil. Two pea rhizobial genotypes were able to nodulate all the five pea cultivars. Furthermore, high strain richness index (0.43-0.5) of field pea rhizobia was observed by both the techniques.

  8. Lactobacillus buchneri genotyping on the basis of clustered regularly interspaced short palindromic repeat (CRISPR) locus diversity.

    Science.gov (United States)

    Briner, Alexandra E; Barrangou, Rodolphe

    2014-02-01

    Clustered regularly interspaced short palindromic repeats (CRISPR) in combination with associated sequences (cas) constitute the CRISPR-Cas immune system, which uptakes DNA from invasive genetic elements as novel "spacers" that provide a genetic record of immunization events. We investigated the potential of CRISPR-based genotyping of Lactobacillus buchneri, a species relevant for commercial silage, bioethanol, and vegetable fermentations. Upon investigating the occurrence and diversity of CRISPR-Cas systems in Lactobacillus buchneri genomes, we observed a ubiquitous occurrence of CRISPR arrays containing a 36-nucleotide (nt) type II-A CRISPR locus adjacent to four cas genes, including the universal cas1 and cas2 genes and the type II signature gene cas9. Comparative analysis of CRISPR spacer content in 26 L. buchneri pickle fermentation isolates associated with spoilage revealed 10 unique locus genotypes that contained between 9 and 29 variable spacers. We observed a set of conserved spacers at the ancestral end, reflecting a common origin, as well as leader-end polymorphisms, reflecting recent divergence. Some of these spacers showed perfect identity with phage sequences, and many spacers showed homology to Lactobacillus plasmid sequences. Following a comparative analysis of sequences immediately flanking protospacers that matched CRISPR spacers, we identified a novel putative protospacer-adjacent motif (PAM), 5'-AAAA-3'. Overall, these findings suggest that type II-A CRISPR-Cas systems are valuable for genotyping of L. buchneri.

  9. Sequential reassortments underlie diverse influenza H7N9 genotypes in China.

    Science.gov (United States)

    Wu, Aiping; Su, Chunhu; Wang, Dayan; Peng, Yousong; Liu, Mi; Hua, Sha; Li, Tianxian; Gao, George F; Tang, Hong; Chen, Jianzhu; Liu, Xiufan; Shu, Yuelong; Peng, Daxin; Jiang, Taijiao

    2013-10-16

    Initial genetic characterizations have suggested that the influenza A (H7N9) viruses responsible for the current outbreak in China are novel reassortants. However, little is known about the pathways of their evolution and, in particular, the generation of diverse viral genotypes. Here we report an in-depth evolutionary analysis of whole-genome sequence data of 45 H7N9 and 42 H9N2 viruses isolated from humans, poultry, and wild birds during recent influenza surveillance efforts in China. Our analysis shows that the H7N9 viruses were generated by at least two steps of sequential reassortments involving distinct H9N2 donor viruses in different hosts. The first reassortment likely occurred in wild birds and the second in domestic birds in east China in early 2012. Our study identifies the pathways for the generation of diverse H7N9 genotypes in China and highlights the importance of monitoring multiple sources for effective surveillance of potential influenza outbreaks.

  10. Genotypic diversity and phenotypic traits of Streptococcus mutans isolates and their relation to severity of early childhood caries.

    Science.gov (United States)

    Valdez, Remberto Marcelo Argandoña; Duque, Cristiane; Caiaffa, Karina Sampaio; Dos Santos, Vanessa Rodrigues; Loesch, Maria Luiza de Aguiar; Colombo, Natalia Helena; Arthur, Rodrigo Alex; Negrini, Thais de Cássia; Boriollo, Marcelo Fabiano Gomes; Delbem, Alberto Carlos Botazzo

    2017-07-14

    Early childhood caries (ECC) is an aggressive condition that can affect teeth of young children. This study aimed to evaluate genotypic diversity and phenotypic traits of S. mutans isolated from dental biofilms of children with different caries status in comparison with caries free (CF) children. Streptococcus mutans strains were isolated from supragingival biofilm samples of CF, ECC and severe-ECC (S-ECC) children and genotyped by arbitrary-primer polymerase chain reaction - AP-PCR. S. mutans genotypes were tested for their ability to reduce the suspension pH through glycolysis, to tolerate extreme acid challenge and by their ability to form biofilm. Response variables were analyzed by ANOVA/Tukey or Kruskal-Wallis/Mann-Whitney tests at a 5% of significance. There was an increase in the prevalence of Streptococcus mutans in biofilms with the severity of dental caries. No differences in genotypic diversity and in acidogenicity of genotypes were found among CF, ECC and S-ECC children. S mutans strains with genotypes more characteristic for ECC and S-ECC children formed more biofilms than those identified in CF children. The strains isolated from S-ECC children were highly acid tolerant. Although S. mutans genotypic diversity was similar among the groups of children, phenotypic traits of S. mutans, especially the acid tolerance response, could explain the severity of early childhood caries.

  11. Predomination and new genotypes of Enterocytozoon bieneusi in captive nonhuman primates in zoos in China: high genetic diversity and zoonotic significance.

    Science.gov (United States)

    Karim, Md Robiul; Dong, Haiju; Li, Tongyi; Yu, Fuchang; Li, Dezhong; Zhang, Longxian; Li, Junqiang; Wang, Rongjun; Li, Shouyi; Li, Xiaofeng; Rume, Farzana Islam; Ning, Changshen

    2015-01-01

    To appreciate the genetic diversity and zoonotic implications of Enterocytozoon bieneusi in nonhuman primates (NHPs) in zoos, we genotyped E. bieneusi in captive NHPs in seven zoos located at six major cities in China, using ribosomal internal transcribed spacer (ITS)-based PCR and sequence analyses. A total of 496 fecal specimens from 36 NHP species under nine families were analyzed and E. bieneusi was detected in 148 (29.8%) specimens of 25 NHP species from six families, including Cercopithecidae (28.7%), Cebidae (38.0%), Aotidae (75.0%), Lemuridae (26.0%), Hylobatidae (50.0%) and Hominidae (16.2%) (P = 0.0605). The infection rates were 29.0%, 15.2%, 18.2%, 37.3%, 29.2%, 37.7% and 44.8% in Shijiazhuang Zoo, Wuhan Zoo, Taiyuan Zoo, Changsha Wild Animal Zoo, Beijing Zoo, Shanghai Zoo and Shanghai Wild Animal Park, respectively (P = 0.0146). A total of 25 ITS genotypes were found: 14 known (D, O, EbpC, EbpA, Type IV, Henan-IV, BEB6, BEB4, Peru8, PigEBITS5, EbpD, CM1, CM4 and CS-1) and 11 new (CM8 to CM18). Genotype D was the most prevalent one (40/148), followed by CM4 (20/148), CM1 (15/148), O (13/148), CM16 (13/148), EbpC (11/148). Of them, genotypes D, EbpC, CM4 and O were widely distributed in NHPs (seen in 9 to 12 species) whereas genotypes CM1 and CM16 were restricted to one to three NHP species. In phylogenetic analysis, 20 genotypes (121/148, 81.8%), excluding genotypes BEB4, BEB6, CM9, CM4 and CM18, belonged to group 1 with zoonotic potential. New genotype CM9 clustered in group 2 with BEB4 and BEB6. The remaining two genotypes CM4 and CM18 formed new cluster (group 9) in between two other genotypic clusters found in primates. The findings of high diversity in E. bieneusi genotypes and their zoonotic potentiality concluded the importance of captive NHPs as reservoir hosts for human microsporidiosis.

  12. Predomination and New Genotypes of Enterocytozoon bieneusi in Captive Nonhuman Primates in Zoos in China: High Genetic Diversity and Zoonotic Significance

    Science.gov (United States)

    Karim, Md Robiul; Dong, Haiju; Li, Tongyi; Yu, Fuchang; Li, Dezhong; Zhang, Longxian; Li, Junqiang; Wang, Rongjun; Li, Shouyi; Li, Xiaofeng; Rume, Farzana Islam; Ning, Changshen

    2015-01-01

    To appreciate the genetic diversity and zoonotic implications of Enterocytozoon bieneusi in nonhuman primates (NHPs) in zoos, we genotyped E. bieneusi in captive NHPs in seven zoos located at six major cities in China, using ribosomal internal transcribed spacer (ITS)-based PCR and sequence analyses. A total of 496 fecal specimens from 36 NHP species under nine families were analyzed and E. bieneusi was detected in 148 (29.8%) specimens of 25 NHP species from six families, including Cercopithecidae (28.7%), Cebidae (38.0%), Aotidae (75.0%), Lemuridae (26.0%), Hylobatidae (50.0%) and Hominidae (16.2%) (P = 0.0605). The infection rates were 29.0%, 15.2%, 18.2%, 37.3%, 29.2%, 37.7% and 44.8% in Shijiazhuang Zoo, Wuhan Zoo, Taiyuan Zoo, Changsha Wild Animal Zoo, Beijing Zoo, Shanghai Zoo and Shanghai Wild Animal Park, respectively (P = 0.0146). A total of 25 ITS genotypes were found: 14 known (D, O, EbpC, EbpA, Type IV, Henan-IV, BEB6, BEB4, Peru8, PigEBITS5, EbpD, CM1, CM4 and CS-1) and 11 new (CM8 to CM18). Genotype D was the most prevalent one (40/148), followed by CM4 (20/148), CM1 (15/148), O (13/148), CM16 (13/148), EbpC (11/148). Of them, genotypes D, EbpC, CM4 and O were widely distributed in NHPs (seen in 9 to 12 species) whereas genotypes CM1 and CM16 were restricted to one to three NHP species. In phylogenetic analysis, 20 genotypes (121/148, 81.8%), excluding genotypes BEB4, BEB6, CM9, CM4 and CM18, belonged to group 1 with zoonotic potential. New genotype CM9 clustered in group 2 with BEB4 and BEB6. The remaining two genotypes CM4 and CM18 formed new cluster (group 9) in between two other genotypic clusters found in primates. The findings of high diversity in E. bieneusi genotypes and their zoonotic potentiality concluded the importance of captive NHPs as reservoir hosts for human microsporidiosis. PMID:25705879

  13. Predomination and new genotypes of Enterocytozoon bieneusi in captive nonhuman primates in zoos in China: high genetic diversity and zoonotic significance.

    Directory of Open Access Journals (Sweden)

    Md Robiul Karim

    Full Text Available To appreciate the genetic diversity and zoonotic implications of Enterocytozoon bieneusi in nonhuman primates (NHPs in zoos, we genotyped E. bieneusi in captive NHPs in seven zoos located at six major cities in China, using ribosomal internal transcribed spacer (ITS-based PCR and sequence analyses. A total of 496 fecal specimens from 36 NHP species under nine families were analyzed and E. bieneusi was detected in 148 (29.8% specimens of 25 NHP species from six families, including Cercopithecidae (28.7%, Cebidae (38.0%, Aotidae (75.0%, Lemuridae (26.0%, Hylobatidae (50.0% and Hominidae (16.2% (P = 0.0605. The infection rates were 29.0%, 15.2%, 18.2%, 37.3%, 29.2%, 37.7% and 44.8% in Shijiazhuang Zoo, Wuhan Zoo, Taiyuan Zoo, Changsha Wild Animal Zoo, Beijing Zoo, Shanghai Zoo and Shanghai Wild Animal Park, respectively (P = 0.0146. A total of 25 ITS genotypes were found: 14 known (D, O, EbpC, EbpA, Type IV, Henan-IV, BEB6, BEB4, Peru8, PigEBITS5, EbpD, CM1, CM4 and CS-1 and 11 new (CM8 to CM18. Genotype D was the most prevalent one (40/148, followed by CM4 (20/148, CM1 (15/148, O (13/148, CM16 (13/148, EbpC (11/148. Of them, genotypes D, EbpC, CM4 and O were widely distributed in NHPs (seen in 9 to 12 species whereas genotypes CM1 and CM16 were restricted to one to three NHP species. In phylogenetic analysis, 20 genotypes (121/148, 81.8%, excluding genotypes BEB4, BEB6, CM9, CM4 and CM18, belonged to group 1 with zoonotic potential. New genotype CM9 clustered in group 2 with BEB4 and BEB6. The remaining two genotypes CM4 and CM18 formed new cluster (group 9 in between two other genotypic clusters found in primates. The findings of high diversity in E. bieneusi genotypes and their zoonotic potentiality concluded the importance of captive NHPs as reservoir hosts for human microsporidiosis.

  14. Physiological, genomic and transcriptional diversity in responses to boron deficiency in rapeseed genotypes

    Science.gov (United States)

    Hua, Yingpeng; Zhou, Ting; Ding, Guangda; Yang, Qingyong; Shi, Lei; Xu, Fangsen

    2016-01-01

    Allotetraploid rapeseed (Brassica napus L. AnAnCnCn, 2n=4x=38) is highly susceptible to boron (B) deficiency, a widespread limiting factor that causes severe losses in seed yield. The genetic variation in the sensitivity to B deficiency found in rapeseed genotypes emphasizes the complex response architecture. In this research, a B-inefficient genotype, ‘Westar 10’ (‘W10’), responded to B deficiencies during vegetative and reproductive development with an over-accumulation of reactive oxygen species, severe lipid peroxidation, evident plasmolysis, abnormal floral organogenesis, and widespread sterility compared to a B-efficient genotype, ‘Qingyou 10’ (‘QY10’). Whole-genome re-sequencing (WGS) of ‘QY10’ and ‘W10’ revealed a total of 1 605 747 single nucleotide polymorphisms and 218 755 insertions/deletions unevenly distributed across the allotetraploid rapeseed genome (~1130Mb). Digital gene expression (DGE) profiling identified more genes related to B transporters, antioxidant enzymes, and the maintenance of cell walls and membranes with higher transcript levels in the roots of ‘QY10’ than in ‘W10’ under B deficiency. Furthermore, based on WGS and bulked segregant analysis of the doubled haploid (DH) line pools derived from ‘QY10’ and ‘W10’, two significant quantitative trait loci (QTLs) for B efficiency were characterized on chromosome C2, and DGE-assisted QTL-seq analyses then identified a nodulin 26-like intrinsic protein gene and an ATP-binding cassette (ABC) transporter gene as the corresponding candidates regulating B efficiency. This research facilitates a more comprehensive understanding of the differential physiological and transcriptional responses to B deficiency and abundant genetic diversity in rapeseed genotypes, and the DGE-assisted QTL-seq analyses provide novel insights regarding the rapid dissection of quantitative trait genes in plant species with complex genomes. PMID:27639094

  15. The molecular basis of beta-thalassemia intermedia in southern China: genotypic heterogeneity and phenotypic diversity

    Directory of Open Access Journals (Sweden)

    Sun Manna

    2010-02-01

    Full Text Available Abstract Background The clinical syndrome of thalassemia intermedia (TI results from the β-globin genotypes in combination with factors to produce fetal haemoglobin (HbF and/or co-inheritance of α-thalassemia. However, very little is currently known of the molecular basis of Chinese TI patients. Methods We systematically analyzed and characterized β-globin genotypes, α-thalassemia determinants, and known primary genetic modifiers linked to the production of HbF and the aggravation of α/β imbalance in 117 Chinese TI patients. Genotype-phenotype correlations were analyzed based on retrospective clinical observations. Results A total of 117 TI patients were divided into two major groups, namely heterozygous β-thalassemia (n = 20 in which 14 were characterized as having a mild TI with the Hb levels of 68-95 g/L except for five co-inherited αααanti-3.7 triplication and one carried a dominant mutation; and β-thalassemia homozygotes or compound heterozygotes for β-thalassemia and other β-globin defects in which the β+-thalassemia mutation was the most common (49/97, hemoglobin E (HbE variants was second (27/97, and deletional hereditary persistence of fetal hemoglobin (HPFH or δβ-thalassemia was third (11/97. Two novel mutations, Term CD+32(A→C and Cap+39(C→T, have been detected. Conclusions Chinese TI patients showed considerable heterogeneity, both phenotypically and genotypically. The clinical outcomes of our TI patients were mostly explained by the genotypes linked to the β- and α-globin gene cluster. However, for a group of 14 patients (13 β0/βN and 1 β+/βN with known heterozygous mutations of β-thalassemia and three with homozygous β-thalassemia (β0/β0, the existence of other causative genetic determinants is remaining to be molecularly defined.

  16. Genotypic diversity and virulence traits of streptococcus sobrinus isolated from caries-free children and children suffering severe early childhood caries.

    Science.gov (United States)

    Qin, Xiu Rong; Zhou, Qiong; Qin, Man

    2013-01-01

    To evaluate the genotypic diversity and some virulence traits of Streptococcus sobrinus (S. sobrinus) isolated from caries-free children and children suffering severe early childhood caries (SECC). S. sobrinus isolated from stimulated whole saliva samples of 91 caries-free children and 87 SECC children were subcultured, identified by polymerase chain reaction and genotyped by arbitrarily primed polymerase chain reaction. Polysaccharide synthesis ability, acidogenicity, aciduricity and the adherence ability of these S. sobrinus isolates were measured. The frequency of S. sobrinus detection was 18.39% (16/87) in SECC children, which was significantly higher than that (3.30%, 3/91) in caries-free children. One to three different genotypes of S. sobrinus were detected in each SECC child. Only one genotype was colonised in each caries-free child. In SECC children, the production of water-insoluble glucan (WIG) was positively correlated with the ability of S. sobrinus adhering to a glass surface. The presence of S. sobrinus could be a risk factor for high caries activity in severe early childhood caries. The multi-genotypes could be related to different caries suceptibility. Water-insoluble glucan plays an important role in the adherence and accumulation of S. sobrinus on tooth surfaces.

  17. Genetic diversity of porcine sapoviruses, kobuviruses, and astroviruses in asymptomatic pigs: an emerging new sapovirus GIII genotype.

    Science.gov (United States)

    Dufkova, Lucie; Scigalkova, Ivana; Moutelikova, Romana; Malenovska, Hana; Prodelalova, Jana

    2013-03-01

    Small, non-enveloped RNA viruses belonging to the genera Sapovirus, Kobuvirus, and Mamastrovirus are usually associated with gastroenteritis in humans and animals. These enteric pathogens are considered potential zoonotic agents. In this study, the prevalence and genetic diversity of sapoviruses (SaVs), kobuviruses (KoVs), and astroviruses (AstVs) in asymptomatic pigs were investigated using a PCR approach. KoV was found to be the most prevalent virus (87.3 %), followed by AstV (34.2 %) and SaV (10.2 %). Interestingly, the intra- and inter-cluster distances between porcine SaV capsid sequences revealed one strain (P38/11/CZ) that formed a new genotype within genogroup III of porcine SaVs, and it is tentatively called "P38/11-like" genotype. Moreover, this is the first report of porcine kobuvirus detection on Czech pig farms. The high prevalence rate of gastroenteritis-producing viruses in clinically healthy pigs represents a continuous source of infection of pigs, and possibly to humans.

  18. Genotypic and phenotypic diversity does not affect productivity and drought response in competitive stands of Trifolium repens

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    Heidrun eHuber

    2016-03-01

    Full Text Available Clonal plants can form dense canopies in which plants of different genetic origin are competing for the uptake of essential resources. The competitive relationships among these clones are likely to be affected by extreme environmental conditions, such as prolonged drought spells, which are predicted to occur more frequently due to global climate change. This, in turn, may alter characteristics of the ecological system and its associated functioning.We hypothesized that the relative success of individual clones will depend on the size of the ramets as ramets with larger leaves and longer petioles (large ramets were predicted to have a competitive advantage in terms of increased light interception over smaller-sized ramets. Under drier conditions the relative performances of genotypes were expected to change leading to a change in genotype ranking. We also hypothesized that increased genotypic and phenotypic diversity will increase stand performance and resistance to drought. These hypotheses and the mechanisms responsible for shifts in competitive relationships were investigated by subjecting genotypes of the important pasture legume Trifolium repens to competition with either genetically identical clones, genetically different but similarly sized clones, or genetically as well as morphologically different clones under well-watered and dry conditions.Competitive relationships were affected by ramet size with large genotypes outperforming small genotypes in diverse stands in terms of biomass production. However, large genotypes also produced relatively fewer ramets than small genotypes and could not benefit in terms of clonal reproduction from competing with smaller genotypes, indicating that evolutionary shifts in genotype composition will depend on whether ramet size or ramet number is under selection. In contrast to our hypotheses, diversity did not increase stand performance under different selection regimes and genotype ranking was hardly

  19. A robust, simple genotyping-by-sequencing (GBS approach for high diversity species.

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    Robert J Elshire

    Full Text Available Advances in next generation technologies have driven the costs of DNA sequencing down to the point that genotyping-by-sequencing (GBS is now feasible for high diversity, large genome species. Here, we report a procedure for constructing GBS libraries based on reducing genome complexity with restriction enzymes (REs. This approach is simple, quick, extremely specific, highly reproducible, and may reach important regions of the genome that are inaccessible to sequence capture approaches. By using methylation-sensitive REs, repetitive regions of genomes can be avoided and lower copy regions targeted with two to three fold higher efficiency. This tremendously simplifies computationally challenging alignment problems in species with high levels of genetic diversity. The GBS procedure is demonstrated with maize (IBM and barley (Oregon Wolfe Barley recombinant inbred populations where roughly 200,000 and 25,000 sequence tags were mapped, respectively. An advantage in species like barley that lack a complete genome sequence is that a reference map need only be developed around the restriction sites, and this can be done in the process of sample genotyping. In such cases, the consensus of the read clusters across the sequence tagged sites becomes the reference. Alternatively, for kinship analyses in the absence of a reference genome, the sequence tags can simply be treated as dominant markers. Future application of GBS to breeding, conservation, and global species and population surveys may allow plant breeders to conduct genomic selection on a novel germplasm or species without first having to develop any prior molecular tools, or conservation biologists to determine population structure without prior knowledge of the genome or diversity in the species.

  20. Black Locust (Robinia pseudoacacia L. Root Cuttings: Diversity and Identity Revealed by SSR Genotyping: A Case Study

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    Maria Emilia Malvolti

    2015-10-01

    Full Text Available Background and Purpose: Black locust (Robinia pseudoacacia L. is a valuable species native to North America and today widely planted throughout the world for biomass production. In Hungary, where Robinia has great importance in the forest management, the clones have been selected for plantations on good, medium and poor quality sites. To conserve the identity, superior clones are vegetatively propagated by root cuttings. At times the collection of root cuttings can cause uncertainty for clonal identity because of the overlap of roots from neighboring plants. This can occur especially when the repository is damaged from severe environmental accidents and the planting layout has been lost. The aim of this study has been to verify by molecular markers the diversity or identity of black locust clones by root cuttings harvested in a damaged trial. Materials and Methods: Root cuttings of 91 clones belonging to five cultivars were collected in a trial severely damaged by storms and flooding periods. The obtained plantlets were analyzed with nine microsatellite (SSR markers and the genetic identity/diversity within and among the plants was tested using the software GenAlEx version 6. Results: Multilocus genotypes (MLG and the Paetkau’s assignation test (1985 revealed genetic variability among the samples: the analyzed plantlets were grouped in four classes instead of the five expected. In addition, 6 unique genotypes have been detected. Conclusions: This study remarks problems that may arise during the harvest of Robinia’s root cuttings, especially when the planting layout has been confused. Molecular analyses can be successfully used to control the germplasm before its sale as guaranty for nurseries, farmers and stakeholders.

  1. Diverse Molecular Genotypes of Mycobacterium tuberculosis Complex Isolates Circulating in the Free State, South Africa

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    Anneke Van der Spoel van Dijk

    2016-01-01

    Full Text Available Tuberculosis is a serious public health concern especially in Africa and Asia. Studies describing strain diversity are lacking in the Free State region of South Africa. The aim of the study was to describe the diversity of Mycobacterium tuberculosis (M. tuberculosis strain families in the Free State province of South Africa. A total of 86 M. tuberculosis isolates were genotyped using spoligotyping. A 12-locus mycobacterial interspersed repetitive units-variable-number tandem repeats (MIRU-VNTRs typing was used to further characterize the resulting spoligotyping clusters. SITVITWEB identified 49 different patterns with allocation to six lineages including Latin-American-Mediterranean (LAM (18 isolates, T (14 isolates, Beijing (five isolates, S (six isolates, Haarlem (one isolate, and X (five isolates, while 37 (43.0% orphans were identified. Eight clusters included 37 isolates with identical spoligotypes (2 to 13/cluster. MIRU-VNTR typing further differentiated three spoligotyping clusters: SIT1/Beijing/MIT17, SIT33/LAM3/MIT213, and confirmed one SIT34/S/MIT311. In addition, SpolDB3/RIM assignment of the orphan strains resulted in a further 10 LAM and 13 T families. In total, LAM (28 isolates and T (27 isolates cause 63% of the individual cases of MTB in our study. The Free State has a highly diverse TB population with LAM being predominant. Further studies with inclusion of multidrug-resistant strains with larger sample size are warranted.

  2. Detection of bovine leukemia virus and identification of its genotype in Mongolian cattle.

    Science.gov (United States)

    Ochirkhuu, Nyamsuren; Konnai, Satoru; Odbileg, Raadan; Nishimori, Asami; Okagawa, Tomohiro; Murata, Shiro; Ohashi, Kazuhiko

    2016-04-01

    Epidemiological studies have indicated that bovine leukemia virus (BLV) infection is globally distributed. However, no information regarding the disease and genetic diversity of the virus in the cattle of Mongolia is currently available. In this study, the prevalence of BLV was assessed using PCR, and the genetic diversity was analyzed through DNA sequencing. Of the 517 samples tested, 20 positives were identified. Phylogenetic analysis showed that six, one, and four isolates were classified into genotype 4, 7, and 1, respectively. Most isolates were clustered with isolates from Eastern Europe and Russia. This study is the first to investigate the BLV genotype in Mongolia.

  3. Study of Mycobacterium tuberculosis Complex Genotypic Diversity in Malaysia Reveals a Predominance of Ancestral East-African-Indian Lineage with a Malaysia-Specific Signature: e114832

    National Research Council Canada - National Science Library

    Fazli Ismail; David Couvin; Izzah Farakhin; Zaidah Abdul Rahman; Nalin Rastogi; Siti Suraiya

    2014-01-01

    .... Objectives This study intended a first assessment of spoligotyping-based MTBC genotypic diversity in Malaysia followed by a comparison of strains with those prevailing in neighboring countries...

  4. DNA fingerprinting and diversity analysis in Aus genotypes using microsatellite markers

    Directory of Open Access Journals (Sweden)

    MD. MONIRUL ISLAM

    2015-08-01

    Full Text Available DNA fingerprinting and genetic diversity of 94 Aus (6 BRRI released Aus variety and 88 local Aus landraces genotypes were carried out to protect the Aus landraces from biopiracy. A total of 91 microsatellite markers were tested for screening the genotypes. Among 91 amplified products, 56% have polymorphic bands giving 195 alleles. The number of alleles per locus ranged from four (RM25 and RM147 to twenty seven (RM519, where average allele number was 9.76. The Polymorphism Information Contents (PIC lied between 0.455 (RM5 to 0.934 (RM519. Most robust marker was found RM519 since it provided the highest PIC value (0.934. Pair-wise genetic dissimilarity co-efficient showed the lowest genetic dissimilarity was found BRRI dhan42 and BRRI dhan43 and the highest genetic dissimilarity was found local landraces each other. Here it is shown that most Aus landraces is recognized to have broad genetic base. Thus it is recommended to use these landraces for future breeding program or include new and untouched local landraces to incorporate new genes and broaden genetic base.

  5. Rare serotype occurrence and PFGE genotypic diversity of Streptococcus agalactiae isolated from tilapia in China.

    Science.gov (United States)

    Li, Liping; Wang, Rui; Liang, Wanwen; Gan, Xi; Huang, Ting; Huang, Yan; Li, Jian; Shi, Yunliang; Chen, Ming; Luo, Honglin

    2013-12-27

    Previously, we reported 10 PEGE types of 85 tilapia Streptococcus agalactiae (GBS), which shifted from Streptococcus iniae in China, by using PEGE method. Presently, larger and more representative tilapia GBS were isolated, for the first time in China, to characterize their serotypes and genetic diversities more precisely than had done before. 168 GBS strains were distributed in five provinces of China, in which Guangdong, Guangxi and Hainan were the major ones, holding 36.9% (62/168), 37.5% (63/168) and 19.6% (33/168), respectively. Serotypes, Ia, Ib and III, were observed in these strains and the most predominant one was Ia (95.2%), which mainly distributed in Guangdong, Guangxi and Hainan. Ia initially occurred in 2009, it shoot up to 32.1% in 2010, but decreased to 16.1% in 2011 before went up to 45.2% in 2012. Ib sporadically occurred during 2007-2011, III only occurred in 2012. 14 different PFGE types, including 4 new types (N, O, P and Q), were observed, in which B, D, F and G were the predominant types, holding 83.9% (141/168) of the total GBS strains. Ia corresponded to 11 PFGE types (A-H, N-P), in which type D predominated (51%). Ib represented 3 genotypes (I, J and Q) and III harbored only 2 genotypes (N and F). Type N and F synchronously presented in Ia and III. In summary, the genetic diversity of tilapia GBS varied by serotypes and changed with geographical locations and years. Although Ia still predominated, new rare serotype III already occurred in China.

  6. Detection of genotype-specific Ehrlichia canis exposure in Brazilian dogs by TRP36 peptide ELISA.

    Science.gov (United States)

    Aguiar, Daniel M; Zhang, Xiaofeng; Braga, Isis A; Taques, Isis I G G; McBride, Jere W

    2016-02-01

    We recently characterized a novel genotype of Ehrlichia canis based on the tandem repeat (TR) sequence of the TRP36 gene in Brazil. The TR amino acid sequence of the Brazilian (Br) genotype (ASVVPEAE) was divergent from the previously described US genotype (TEDSVSAPA) of E. canis. In this study, we developed an ELISA based on TRP36 TR synthetic peptides from both Br and US E. canis TRP36 genotypes to serologically detect and distinguish infections caused by these genotypes. Sera from 30 Brazilian dogs naturally infected with E. canis, sera from dogs experimentally infected E. canis (Jake and Cuiabá #1 strains) and E. chaffeensis (Arkansas strain) and 12 seronegative E. canis dogs were evaluated. Fifteen naturally infected Brazilian dogs had antibodies that reacted with the US TRP36 (n=9) or Br TRP36 (n=6) only, and 13 dogs had antibodies that reacted with both TPR36 peptides suggesting that these dogs were exposed to both genotypes. Most dogs (n=28) had antibodies that reacted with the highly conserved E. canis TRP19 peptide; however, two dogs had antibodies to E. canis TRP19, but did not have TRP36 antibodies, raising the possibility that another novel TRP36 genotype is circulating in Brazil. Our results demonstrate that synthetic peptides based on the TR region of E. canis TRP36 can be used to serologically distinguish infections or identify coinfections by different genotypes, and to determine the seroprevalence of various E. canis genotypes in Brazil.

  7. Genotypic diversity and environmental stability of starch physicochemical properties in the USDA rice mini-core collection.

    Science.gov (United States)

    Li, Kehu; Bao, Jinsong; Corke, Harold; Sun, Mei

    2017-04-15

    The USDA rice mini-core collection consists of 217 accessions representative of a world-wide germplam bank. We investigated its genotypic diversity in starch physicochemical properties and the effects of genotype, environment and G×E interaction in this study. High genotypic diversity was found in all 18 measured starch quality traits in the mini-core rice in two location-years in China. Genotype, environment and G×E effects on these traits were analysed using 115 common accessions successfully produced in both environments. Thermal properties (To, Tp and Tc) were very stable whereas most other traits differed significantly between environments. However, when these accessions were divided into five subgroups based on amylose content, environment was found to have differential effects. G×E interaction also played a significant role in determining the starch traits. These findings will provide guidance for selection from the diverse genotypes in the USDA mini-core collection for cultivation and for developing cultivars with desired cooking and eating quality. Copyright © 2016 Elsevier Ltd. All rights reserved.

  8. Genotypic diversity of stress response in Lactobacillus plantarum, Lactobacillus paraplantarum and Lactobacillus pentosus.

    Science.gov (United States)

    Ricciardi, Annamaria; Parente, Eugenio; Guidone, Angela; Ianniello, Rocco Gerardo; Zotta, Teresa; Abu Sayem, S M; Varcamonti, Mario

    2012-07-02

    Lactobacillus plantarum, Lactobacillus pentosus and Lactobacillus paraplantarum are three closely related species which are widespread in food and non-food environments, and are important as starter bacteria or probiotics. In order to evaluate the phenotypic diversity of stress tolerance in the L. plantarum group and the ability to mount an adaptive heat shock response, the survival of exponential and stationary phase and of heat adapted exponential phase cells of six L. plantarum subsp. plantarum, one L. plantarum subsp. argentoratensis, one L. pentosus and two L. paraplantarum strains selected in a previous work upon exposure to oxidative, heat, detergent, starvation and acid stresses was compared to that of the L. plantarum WCFS1 strain. Furthermore, to evaluate the genotypic diversity in stress response genes, ten genes (encoding for chaperones DnaK, GroES and GroEL, regulators CtsR, HrcA and CcpA, ATPases/proteases ClpL, ClpP, ClpX and protease FtsH) were amplified using primers derived from the WCFS1 genome sequence and submitted to restriction with one or two endonucleases. The results were compared by univariate and multivariate statistical methods. In addition, the amplicons for hrcA and ctsR were sequenced and compared by multiple sequence alignment and polymorphism analysis. Although there was evidence of a generalized stress response in the stationary phase, with increase of oxidative, heat, and, to a lesser extent, starvation stress tolerance, and for adaptive heat stress response, with increased tolerance to heat, acid and detergent, different growth phases and adaptation patterns were found. Principal component analysis showed that while heat, acid and detergent stresses respond similarly to growth phase and adaptation, tolerance to oxidative and starvation stresses implies completely unrelated mechanisms. A dendrogram obtained using the data from multilocus restriction typing (MLRT) of stress response genes clearly separated two groups of L

  9. [Hepatitis B virus genotype E infection in Turkey: the detection of the first case].

    Science.gov (United States)

    Sayan, Murat; Sanlıdağ, Tamer; Akçalı, Sinem; Arıkan, Ayşe

    2014-10-01

    Hepatitis B virus (HBV) infection is a global major health problem. Currently, 10 genotypes (A-J) of hepatitis B virus (HBV) are identified based on the nucleic acid sequence heterogeneity, and these genotypes have been shown to have distinct geographic distribution. Reports of the previous studies indicated that the genotype D is the predominant type among hepatitis B patients in different regions of Turkey. However, recent studies indicated that other HBV genotypes are also seen with an increasing rate. Although epidemiological and clinical information on genotype E infection is currently limited, it is known that genotype E infection is common in West and Central Africa. In this report, the first case of HBV genotype E infection in Turkey was presented. A 22-year-old Nigerian male employee who resided in Manisa for five years was admitted to Celal Bayar University Hospital Manisa, Turkey, for his routine check-up. Since HBsAg was found positive, other HBV markers were tested with a repeated serum sample. Laboratory findings were as follows; HBsAg (+), anti-HBs (-), HBeAg (-), anti-HBe (+), anti-HBc (+), anti-HCV (-), anti-HIV (-), ALT: 44 U/L and AST: 45 U/L. HBV-DNA level was detected as 700 IU/ml by real-time PCR (Artus HBV QS RGQ Qiagen, Germany). HBV-DNA isolated from the serum sample of the patient was amplified by PCR and polymerase gene segment of HBV was directly sequenced. UPGMA method was used for phylogenetic analysis and Inno-LIPA HBV genotyping method (Innogenetics, Belgium) was performed to determine multiple HBV genotype infection. On the basis of those methods the genotype of the virus was identified as genotype E. The partial sequences of the HBV polymerase gene were loaded to the international DNA data bank (GenBank) for contribution to the global HBV surveillance. This report emphasized that besides genotype D the other HBV genotypes could be found in Turkey. Since the patient was an inactive HBsAg carrier before his residence in Turkey, this

  10. Genotypic and chemotypic diversity of Neotyphodium endophytes in tall fescue from Greece.

    Science.gov (United States)

    Takach, Johanna E; Mittal, Shipra; Swoboda, Ginger A; Bright, Sherrita K; Trammell, Michael A; Hopkins, Andrew A; Young, Carolyn A

    2012-08-01

    Epichloid endophytes provide protection from a variety of biotic and abiotic stresses for cool-season grasses, including tall fescue. A collection of 85 tall fescue lines from 15 locations in Greece, including both Continental and Mediterranean germplasm, was screened for the presence of native endophytes. A total of 37 endophyte-infected lines from 10 locations were identified, and the endophytes were classified into five distinct groups (G1 to G5) based on physical characteristics such as colony morphology, growth rate, and conidial morphology. These classifications were supported by phylogenetic analyses of housekeeping genes tefA and tubB, and the endophytes were further categorized as Neotyphodium coenophialum isolates (G1, G4, and G5) or Neotyphodium sp. FaTG-2 (Festuca arundinacea taxonomic group 2 isolates (G2 and G3). Analyses of the tall fescue matK chloroplast genes indicated a population-wide, host-specific association between N. coenophialum and Continental tall fescue and between FaTG-2 and Mediterranean tall fescue that was also reflected by differences in colonization of host tillers by the native endophytes. Genotypic analyses of alkaloid gene loci combined with chemotypic (chemical phenotype) profiles provided insight into the genetic basis of chemotype diversity. Variation in alkaloid gene content, specifically the presence and absence of genes, and copy number of gene clusters explained the alkaloid diversity observed in the endophyte-infected tall fescue, with one exception. The results from this study provide insight into endophyte germplasm diversity present in living tall fescue populations.

  11. Genetic diversity of avian paramyxovirus type 1: Proposal for a unified nomenclature and classification system of Newcastle disease virus genotypes

    Science.gov (United States)

    Genetically diverse Newcastle disease virus (NDV) isolates circulate and cause disease in different geographic locations of the world. The differences found on the genome of distinct NDV isolates have been used to classify different isolates into genetic groups called genotypes or lineages. Both l...

  12. Assessment of the genotypic diversity of antibiotic-producing Pseudomonas species in the rhizosphere by denaturing gradient gel electrophoresis

    NARCIS (Netherlands)

    Bergsma-Vlami, M.; Prins, M.E.; Staats, M.; Raaijmakers, J.M.

    2005-01-01

    The genotypic diversity of antibiotic-producing Pseudomonas spp. provides an enormous resource for identifying strains that are highly rhizosphere competent and superior for biological control of plant diseases. In this study, a simple and rapid method was developed to determine the presence and gen

  13. Comparison of genotypes I and III in Japanese encephalitis virus reveals distinct differences in their genetic and host diversity.

    Science.gov (United States)

    Han, Na; Adams, James; Chen, Ping; Guo, Zhen-yang; Zhong, Xiang-fu; Fang, Wei; Li, Na; Wen, Lei; Tao, Xiao-yan; Yuan, Zhi-ming; Rayner, Simon

    2014-10-01

    Japanese encephalitis (JE) is an arthropod-borne disease associated with the majority of viral encephalitis cases in the Asia-Pacific region. The causative agent, Japanese encephalitis virus (JEV), has been phylogenetically divided into five genotypes. Recent surveillance data indicate that genotype I (GI) is gradually replacing genotype III (GIII) as the dominant genotype. To investigate the mechanism behind the genotype shift and the potential consequences in terms of vaccine efficacy, human cases, and virus dissemination, we collected (i) all full-length and partial JEV molecular sequences and (ii) associated genotype and host information comprising a data set of 873 sequences. We then examined differences between the two genotypes at the genetic and epidemiological level by investigating amino acid mutations, positive selection, and host range. We found that although GI is dominant, it has fewer sites predicted to be under positive selection, a narrower host range, and significantly fewer human isolates. For the E protein, the sites under positive selection define a haplotype set for each genotype that shows striking differences in their composition and diversity, with GIII showing significantly more variety than GI. Our results suggest that GI has displaced GIII by achieving a replication cycle that is more efficient but is also more restricted in its host range. Japanese encephalitis is an arthropod-borne disease associated with the majority of viral encephalitis cases in the Asia-Pacific region. The causative agent, Japanese encephalitis virus (JEV), has been divided into five genotypes based on sequence similarity. Recent data indicate that genotype I (GI) is gradually replacing genotype III (GIII) as the dominant genotype. Understanding the reasons behind this shift and the potential consequences in terms of vaccine efficacy, human cases, and virus dissemination is important for controlling the spread of the virus and reducing human fatalities. We

  14. Annual research review: Rare genotypes and childhood psychopathology--uncovering diverse developmental mechanisms of ADHD risk.

    Science.gov (United States)

    Scerif, Gaia; Baker, Kate

    2015-03-01

    Through the increased availability and sophistication of genetic testing, it is now possible to identify causal diagnoses in a growing proportion of children with neurodevelopmental disorders. In addition to developmental delay and intellectual disability, many genetic disorders are associated with high risks of psychopathology, which curtail the wellbeing of affected individuals and their families. Beyond the identification of significant clinical needs, understanding the diverse pathways from rare genetic mutations to cognitive dysfunction and emotional-behavioural disturbance has theoretical and practical utility. We overview (based on a strategic search of the literature) the state-of-the-art on causal mechanisms leading to one of the most common childhood behavioural diagnoses - attention deficit hyperactivity disorder (ADHD) - in the context of specific genetic disorders. We focus on new insights emerging from the mapping of causal pathways from identified genetic differences to neuronal biology, brain abnormalities, cognitive processing differences and ultimately behavioural symptoms of ADHD. First, ADHD research in the context of rare genotypes highlights the complexity of multilevel mechanisms contributing to psychopathology risk. Second, comparisons between genetic disorders associated with similar psychopathology risks can elucidate convergent or distinct mechanisms at each level of analysis, which may inform therapeutic interventions and prognosis. Third, genetic disorders provide an unparalleled opportunity to observe dynamic developmental interactions between neurocognitive risk and behavioural symptoms. Fourth, variation in expression of psychopathology risk within each genetic disorder points to putative moderating and protective factors within the genome and the environment. A common imperative emerging within psychopathology research is the need to investigate mechanistically how developmental trajectories converge or diverge between and within

  15. Genetic diversity of "Pimenta Longa" genotypes (Piper spp., Piperaceae of the Embrapa Acre germplasm collection

    Directory of Open Access Journals (Sweden)

    Lúcia Helena de Oliveira Wadt

    2004-01-01

    Full Text Available The commonly known Pimenta longa is a commercially valuable natural resource found wild in Acre, Brazil. Specifically, three Piperaceae species with contested taxonomic status were studied, Piper hispidinervum, Piper aduncum, and Piper hispidum, to assesses the inter- and intra-specific genetic relationship of 49 Piper genotypes kept in the Pimenta longa germplasm collection at Embrapa Acre, using sixty six Random Amplified Polymorphic DNA (RAPD markers. The DNA polymorphism level detected was high (96.97%, but the marker frequencies for each species showed polymorphism levels of 79.4% for Piper hispidinervum and 5.3% for P. aduncum. The genetic similarity clustering analysis resulted in three distinct groups corresponding to Piper hispidinervum, Piper aduncum, and Piper hispidum. Four and nine characteristic RAPD markers were identified for P. hispidinervum and P. aduncum, respectively, supporting the existence of two separate species. However, six genotypes collected in Tarauacá county formed a distinct subgroup within the P. hispidinervum group and may be considered as an ecotype of this species or an intermediate between the P. hispidinervum and P. aduncum groups. More extensive sampling of both P. hispidinervum and P. aduncum populations throughout the region are needed to further establish their relation and its implication for breeding efforts.

  16. Pathogenic potential and genotypic diversity of Campylobacter jejuni: a neglected food-borne pathogen in Brazil.

    Science.gov (United States)

    Frazão, Miliane Rodrigues; Medeiros, Marta Inês Cazentini; Duque, Sheila da Silva; Falcão, Juliana Pfrimer

    2017-03-01

    Purpose and methodology.Campylobacter jejuni is a major zoonotic pathogen that causes food-borne gastroenteritis worldwide. However, there are only a few studies available that have molecularly characterized C. jejuni strains isolated in Brazil. The aim of this study was to genotype 111 C. jejuni strains isolated from sick humans (43), monkey faeces (19), chicken faeces (14), chicken meat (33) and sewage (2) between 1996 and 2016 in Brazil using flaA-SVR (short variable region) sequencing and PFGE. Furthermore, the presence of 16 virulence genes was analysed by PCR. Using PFGE and flaA-SVR sequencing, the 111 C. jejuni strains studied were grouped into three and two clusters, respectively, and some strains of different origin presented a similarity of ≥80 %. In total, 35 flaA-SVR alleles were detected. Alleles gt45, gt49 and gt57 were the most prevalent, in contrast with those frequently described in the PubMLST database. All 111 C. jejuni strains contained the genes flaA, flhA, cadF, docA, cdtA, cdtB, cdtC, iamA, ciaB, sodB, dnaJ, pldA, racR and csrA. The wlaN gene was detected in 11 strains (9.9 %), and the virB11 in just one strain (0.9 %). In conclusion, the pathogenic potential of the C. jejuni strains studied was highlighted by the high frequency of the majority of the virulence genes searched. The flaA-SVR sequencing and PFGE results showed that some of the strains studied presented a high genotypic similarity, suggesting potential for transmission between animal sources and humans in this country. Altogether, the results characterize further C. jejuni isolates from Brazil, an important producer and exporter of chicken meat.

  17. Capillary electrophoresis with laser-induced fluorescence detection for fast and reliable apolipoprotein E genotyping

    NARCIS (Netherlands)

    Somsen, GW; Welten, HTME; Mulder, FP; Swart, CW; Kema, IP; de Jong, GJ

    2002-01-01

    The use of capillary electrophoresis (CE) with laser-induced fluorescence (LIF) detection for the rapid determination of apolipoprotein E (apoE) genotypes was studied. High resolution and sensitive detection of the concerned DNA restriction fragments was achieved using CE buffers with hydroxypropylm

  18. Genetic diversity of Bacillus anthracis in Europe: genotyping methods in forensic and epidemiologic investigations.

    Science.gov (United States)

    Derzelle, Sylviane; Thierry, Simon

    2013-09-01

    Bacillus anthracis, the etiological agent of anthrax, a zoonosis relatively common throughout the world, can be used as an agent of bioterrorism. In naturally occurring outbreaks and in criminal release of this pathogen, a fast and accurate diagnosis is crucial to an effective response. Microbiological forensics and epidemiologic investigations increasingly rely on molecular markers, such as polymorphisms in DNA sequence, to obtain reliable information regarding the identification or source of a suspicious strain. Over the past decade, significant research efforts have been undertaken to develop genotyping methods with increased power to differentiate B. anthracis strains. A growing number of DNA signatures have been identified and used to survey B. anthracis diversity in nature, leading to rapid advances in our understanding of the global population of this pathogen. This article provides an overview of the different phylogenetic subgroups distributed across the world, with a particular focus on Europe. Updated information on the anthrax situation in Europe is reported. A brief description of some of the work in progress in the work package 5.1 of the AniBioThreat project is also presented, including (1) the development of a robust typing tool based on a suspension array technology and multiplexed single nucleotide polymorphisms scoring and (2) the typing of a collection of DNA from European isolates exchanged between the partners of the project. The know-how acquired will contribute to improving the EU's ability to react rapidly when the identity and real origin of a strain need to be established.

  19. Genetic diversity for grain Zn concentration in finger millet genotypes: Potential for improving human Zn nutrition

    Directory of Open Access Journals (Sweden)

    Ramegowda Yamunarani

    2016-06-01

    Full Text Available Nearly half of the world population suffers from micronutrient malnutrition, particularly Zn deficiency. It is important to understand genetic variation for uptake and translocation behaviors of Zn in relevant crop species to increase Zn concentration in edible parts. In the present study, genetic variation in grain Zn concentration of 319 finger millet genotypes was assessed. Large genetic variation was found among the genotypes, with concentrations ranging from 10 to 86 μg g− 1 grain. Uptake and translocation studies with Zn/65Zn application in 12 selected low-Zn genotypes showed wide variation in root uptake and shoot translocation, with genotypes GEC331 and GEC164 showing greater uptake and translocation. Genotypes GEC164 and GEC543 showed increased grain Zn concentration. Genotypes GEC331 and GEC164 also showed improved yield under Zn treatment. Appreciable variation in grain Zn concentration among finger millet genotypes found in this study offers opportunities to improve Zn nutrition through breeding.

  20. Genotype MTBDR plus assay for molecular detection of rifampicin and isoniazid resistance in Mycobacterium tuberculosis

    Directory of Open Access Journals (Sweden)

    Soniya Sharma

    2014-01-01

    Full Text Available Aim: This study was performed for the rapid identification of Mycobacterium tuberculosis complex and its resistance to rifampicin and isoniazid, directly from the sputum samples of pulmonary tuberculosis patients. Materials and Methods: A commercially available genotype MTBDR plus assay was used for the identification and detection of mutations in Mycobacterial isolates. A total of 100 sputum samples of pulmonary tuberculosis patients were analyzed by using the genotype MTBDR plus assay. The MTBDR plus assay is designed to detect the mutations in the hotspot region of rpoB gene, katG and regulatory region of inhA gene. Results: The genotype MTBDR plus assay detected 22% multidrug resistant (MDR, 2% rifampicin (RMP monoresistant and 1% isoniazid (INH monoresistant isolates. In 22 MDR isolates, the codons most frequently involved in RMP-associated mutations were codon 531 (54.55%, 516 (31.82% and 526 (13.63%, and 90.90% of MDR isolates showed KatG S315T mutations and 9.1% showed inhA C-15T mutations associated with INH resistance. Conclusion: The new genotype MTBDR plus assay represents a rapid, reliable tool for the detection of MDR-TB, wherein results are obtained in 5 h allowing early and appropriate treatment, which is essential to cut the transmission path and reduce the spread of MDR-TB. The genotype MTBDR plus assay can readily be included in a routine laboratory work for the early diagnosis and control of MDR-TB.

  1. Detection of Helicobacter pylori and the genotypes of resistance to clarithromycin and the heterogeneous genotype to this antibiotic in biopsies obtained from symptomatic children.

    Science.gov (United States)

    Aguilera-Correa, John Jairo; Urruzuno, Pedro; Barrio, Josefa; Martinez, María José; Agudo, Sonia; Somodevilla, Angela; Llorca, Laura; Alarcón, Teresa

    2017-02-01

    The aim of this study was to use a commercially available kit (GenoType® HelicoDR; Hain Life Science, Germany) to detect Helicobacter pylori infection and clarithromycin resistance genotype in biopsies obtained from symptomatic children.

  2. Genotypic and functional diversity of phenotypically defined primitive hematopoietic cells in patients with chronic myeloid leukemia.

    Science.gov (United States)

    Sloma, Ivan; Beer, Philip A; Saw, Kyi Min; Chan, Matthew; Leung, Donna; Raghuram, Kamini; Brimacombe, Cedric; Johnston, Bobby; Lambie, Karen; Forrest, Donna; Jiang, Xiaoyan; Eaves, Connie J

    2013-10-01

    Much progress has been made in the management of chronic-phase chronic myeloid leukemia (CP-CML), but there is a continuing imperative to develop curative treatments, predict patient responses to specific modalities, and anticipate disease relapse or progression. These needs underlie continuing interest in methods to detect and quantify the relevant leukemic cells in clinical samples with improved reliability and specificity. We report the results of comparing three methods to enumerate primitive CP-CML cells in the same samples: genotyping CD34(+)38(-) cells directly by fluorescence in situ hybridization, and measuring BCR-ABL1 transcript-genotyped colony-forming cell outputs in either 5-week long-term cultures (LTCs) containing non-engineered mouse fibroblasts or in 6-week LTCs containing mouse fibroblasts engineered to produce human Steel factor, granulocyte colony-stimulating factor, and IL-3. The results demonstrate that the first two methods significantly overestimate the prevalence of primitive CP-CML cells by comparison to the third. In additional studies, we found that CML-CD34(+) cells can repopulate the marrow and spleen of serially transplanted adult NOD/SCID-IL-2Rγ chain-null mice for more than 1 year with an almost exclusive myeloid differentiation in primary and secondary recipients and without evidence of disease progression. These findings underscore the importance of long-term functional in vitro and in vivo endpoints to identify and characterize CP-CML stem cells. Crown Copyright © 2013. Published by Elsevier Inc. All rights reserved.

  3. Detection of Rotavirus Genotypes in Korea 5 Years after the Introduction of Rotavirus Vaccines.

    Science.gov (United States)

    Chung, Ju-Young; Kim, Min-Sung; Jung, Tae Woong; Kim, Seong Joon; Kang, Jin-Han; Han, Seung Beom; Kim, Sang Yong; Rhim, Jung Woo; Kim, Hwang-Min; Park, Jae Hong; Jo, Dae Sun; Ma, Sang Hyuk; Jeong, Hye-Sook; Cheon, Doo-Sung; Kim, Jong-Hyun

    2015-10-01

    Rotavirus (RV) is one of the most important viral etiologic agents of acute gastroenteritis (AGE) in children. Although effective RV vaccines (RVVs) are now used worldwide, novel genotypes and outbreaks resulting from rare genotype combinations have emerged. This study documented RV genotypes in a Korean population of children with AGE 5 yr after the introduction of RVV and assessed potential genotype differences based on vaccination status or vaccine type. Children less than 5-yr-old diagnosed with AGE between October 2012 and September 2013 admitted to 9 medical institutions from 8 provinces in Korea were prospectively enrolled. Stool samples were tested for RV by enzyme immunoassay and genotyped by multiplex reverse-transcription polymerase chain reaction. In 346 patients, 114 (32.9%) were RV-positive. Among them, 87 (76.3%) patients were infected with RV alone. Eighty-six of 114 RV-positive stool samples were successfully genotyped, and their combinations of genotypes were G1P[8] (36, 41.9%), G2P[4] (12, 14.0%), and G3P[8] (6, 7.0%). RV was detected in 27.8% of patients in the vaccinated group and 39.8% in the unvaccinated group (P=0.035). Vaccination history was available for 67 of 86 cases with successfully genotyped RV-positive stool samples; RotaTeq (20, 29.9%), Rotarix (7, 10.4%), unvaccinated (40, 59.7%). The incidence of RV AGE is lower in the RV-vaccinated group compared to the unvaccinated group with no evidence of substitution with unusual genotype combinations.

  4. Detection of multiple sapovirus genotypes and genogroups in oyster-associated outbreaks.

    Science.gov (United States)

    Nakagawa-Okamoto, Reiko; Arita-Nishida, Tomoko; Toda, Shoichi; Kato, Hirotomo; Iwata, Hiroyuki; Akiyama, Miho; Nishio, Osamu; Kimura, Hirokazu; Noda, Mamoru; Takeda, Naokazu; Oka, Tomoichiro

    2009-01-01

    This report describes multiple viruses in stool specimens from oyster-associated gastroenteritis. Eleven outbreaks of oyster-associated gastroenteritis were examined for enteric viruses between January 2002 and March 2006 in Japan. Multiple norovirus genotypes were detected in all outbreaks; moreover, kobuvirus, sapovirus, and astrovirus were also detected in 6, 3, and 1 of the 11 outbreaks, respectively. Notably, multiple sapovirus genogroups were detected in the stool specimens from subjects in two oyster-associated gastroenteritis outbreaks.

  5. Genotypic and phenotypic detection of efflux pump in Rhodococcus equi.

    Science.gov (United States)

    Gressler, Letícia Trevisan; de Vargas, Agueda Castagna; da Costa, Mateus Matiuzzi; Pötter, Luciana; da Silveira, Bibiana Petri; Sangioni, Luis Antônio; de Avila Botton, Sônia

    2014-01-01

    The req_39680 gene, associated to a putative efflux system, was detected in 60% (54/90) of R. equi isolates by PCR. The phenotypic expression of efflux mechanism was verified in 20% of the isolates using ethidium bromide. For the first time, the expression of efflux mechanism was demonstrated in R. equi.

  6. Genotypic and phenotypic detection of efflux pump in Rhodococcus equi

    Directory of Open Access Journals (Sweden)

    Letícia Trevisan Gressler

    2014-06-01

    Full Text Available The req_39680 gene, associated to a putative efflux system, was detected in 60% (54/90 of R. equi isolates by PCR. The phenotypic expression of efflux mechanism was verified in 20% of the isolates using ethidium bromide. For the first time, the expression of efflux mechanism was demonstrated in R. equi.

  7. Phylogenetic diversity and genotypical complexity of H9N2 influenza A viruses revealed by genomic sequence analysis.

    Directory of Open Access Journals (Sweden)

    Guoying Dong

    Full Text Available H9N2 influenza A viruses have become established worldwide in terrestrial poultry and wild birds, and are occasionally transmitted to mammals including humans and pigs. To comprehensively elucidate the genetic and evolutionary characteristics of H9N2 influenza viruses, we performed a large-scale sequence analysis of 571 viral genomes from the NCBI Influenza Virus Resource Database, representing the spectrum of H9N2 influenza viruses isolated from 1966 to 2009. Our study provides a panoramic framework for better understanding the genesis and evolution of H9N2 influenza viruses, and for describing the history of H9N2 viruses circulating in diverse hosts. Panorama phylogenetic analysis of the eight viral gene segments revealed the complexity and diversity of H9N2 influenza viruses. The 571 H9N2 viral genomes were classified into 74 separate lineages, which had marked host and geographical differences in phylogeny. Panorama genotypical analysis also revealed that H9N2 viruses include at least 98 genotypes, which were further divided according to their HA lineages into seven series (A-G. Phylogenetic analysis of the internal genes showed that H9N2 viruses are closely related to H3, H4, H5, H7, H10, and H14 subtype influenza viruses. Our results indicate that H9N2 viruses have undergone extensive reassortments to generate multiple reassortants and genotypes, suggesting that the continued circulation of multiple genotypical H9N2 viruses throughout the world in diverse hosts has the potential to cause future influenza outbreaks in poultry and epidemics in humans. We propose a nomenclature system for identifying and unifying all lineages and genotypes of H9N2 influenza viruses in order to facilitate international communication on the evolution, ecology and epidemiology of H9N2 influenza viruses.

  8. Diversity in the carotenoid profiles and the expression of genes related to carotenoid accumulation among citrus genotypes

    OpenAIRE

    Ikoma, Yoshinori; Matsumoto, Hikaru; Kato, Masaya

    2016-01-01

    Carotenoids are not only important to the plants themselves but also are beneficial to human health. Since citrus fruit is a good source of carotenoids for the human diet, it is important to study carotenoid profiles and the accumulation mechanism in citrus fruit. Thus, in the present paper, we describe the diversity in the carotenoid profiles of fruit among citrus genotypes. In regard to carotenoids, such as β-cryptoxanthin, violaxanthin, lycopene, and β-citraurin, the relationship between t...

  9. Cerebral and non-cerebral coenurosis: on the genotypic and phenotypic diversity of Taenia multiceps.

    Science.gov (United States)

    Christodoulopoulos, Georgios; Dinkel, Anke; Romig, Thomas; Ebi, Dennis; Mackenstedt, Ute; Loos-Frank, Brigitte

    2016-12-01

    We characterised the causative agents of cerebral and non-cerebral coenurosis in livestock by determining the mitochondrial genotypes and morphological phenotypes of 52 Taenia multiceps isolates from a wide geographical range in Europe, Africa, and western Asia. Three studies were conducted: (1) a morphological comparison of the rostellar hooks of cerebral and non-cerebral cysts of sheep and goats, (2) a morphological comparison of adult worms experimentally produced in dogs, and (3) a molecular analysis of three partial mitochondrial genes (nad1, cox1, and 12S rRNA) of the same isolates. No significant morphological or genetic differences were associated with the species of the intermediate host. Adult parasites originating from cerebral and non-cerebral cysts differed morphologically, e.g. the shape of the small hooks and the distribution of the testes in the mature proglottids. The phylogenetic analysis of the mitochondrial haplotypes produced three distinct clusters: one cluster including both cerebral isolates from Greece and non-cerebral isolates from tropical and subtropical countries, and two clusters including cerebral isolates from Greece. The majority of the non-cerebral specimens clustered together but did not form a monophyletic group. No monophyletic groups were observed based on geography, although specimens from the same region tended to cluster. The clustering indicates high intraspecific diversity. The phylogenetic analysis suggests that all variants of T. multiceps can cause cerebral coenurosis in sheep (which may be the ancestral phenotype), and some variants, predominantly from one genetic cluster, acquired the additional capacity to produce non-cerebral forms in goats and more rarely in sheep.

  10. Novel Phl-producing genotypes of finger millet rhizosphere associated pseudomonads and assessment of their functional and genetic diversity.

    Science.gov (United States)

    Sekar, Jegan; Prabavathy, Vaiyapuri Ramalingam

    2014-07-01

    Genetic diversity of phlD gene, an essential gene in the biosynthesis of 2,4-diacetylphloroglucinol, was studied by restriction fragment length polymorphism (RFLP) in 20 Phl-producing pseudomonads isolated from finger millet rhizosphere. RFLP analysis of phlD gene displayed three patterns with HaeIII and TaqI enzymes. phlD gene sequence closely correlated with RFLP results and revealed the existence of three new genotypes G, H and I. Further, the phylogenetic and concatenated sequence analysis of the 16S rRNA, rpoB, gyrB, rpoD genes supported the hypothesis that these genotypes G, H and I were different from reported genotypes A-F. In all phylogenetic studies, the genotype G formed a distant clade from the groups of Pseudomonas putida and P. aeruginosa (sensu strictu), but the groups H and I were closely related to P. aeruginosa/P. stutzeri group. The Phl-producing pseudomonads exhibited antagonistic activity against Pyricularia grisea (TN508), Gaeumannomyces graminis (DSM1463), Fusarium oxysporum (DSM62297), Xanthomonas campestris (DSM3586) and Erwinia persicina (HMGU155). In addition, these strains exhibited various plant growth-promoting traits. In conclusion, this study displays the existence of novel Phl-producing pseudomonads genotypes G, H and I from finger millet rhizosphere, which formed taxonomically outward phylogenetic lineage from the groups of P. putida and P. aeruginosa (sensu strictu).

  11. Genetic diversity of group A rotaviruses associated with repeated outbreaks of diarrhea in a farrow-to-finish farm: identification of a porcine rotavirus strain bearing a novel VP7 genotype, G26

    Science.gov (United States)

    2011-01-01

    Group A rotaviruses (GARs) are one of the most common causes of diarrhea in suckling pigs. Although a number of G and P genotypes have been identified in porcine GARs, few attempts have been made to study the molecular epidemiology of these viruses associated with diarrhea outbreaks within a farm over an extended period of time. Here, we investigated the molecular characteristics of GARs that caused four outbreaks of diarrhea among suckling pigs in a farrow-to-finish farm over the course of a year. G and P genotyping of GARs detected at each outbreak demonstrated genetic diversity in this farm as follows: G9P[23] was detected at the first outbreak, G9P[13]/[22] and G9P[23] at the second, G3P[7] at the third, and G9P[23], G5P[13]/[22], and P[7] combined with an untypeable G genotype at the fourth. Sequence analysis of the detected GARs revealed that such genetic diversity could have resulted not only from the introduction of new GAR strains, but also from gene reassortment between GAR strains within the farm. Further, the GAR strain carrying the untypeable G genotype was shown to be a novel porcine GAR bearing a new G26 genotype, as confirmed by the Rotavirus Classification Working Group. PMID:22067072

  12. [Detection and genotype analysis of sapovirus associated with sporadic diarrhea in Shenzhen in 2009].

    Science.gov (United States)

    Wang, Yan-ge; He, Ya-qing; Xiang, Wen-long; Liao, Yu-xue; Chen, Qing; Yu, Shou-yi; Hu, Gui-fang

    2011-06-01

    To conduct an epidemiological and genotype analysis of sapovirus (SaV) associated with sporadic diarrhea in Shenzhen in the year 2009. A total of 852 fecal samples were collected from sporadic cases of diarrhea in Shenzhen in 2009 and detected by reverse-transcription polymerase chain reaction (RT-PCR) using the primers of SLV5317/5749. The PCR products were analyzed with 1.5% agarose gel electrophoresis and sequenced to construct the phylogenetic tree. Sixteen samples were found positive for SaV, with a positivity rate of 1.88%. Sequence analysis identified 8 isolates as SaV GI genotype (including 3 SaV GI.1 and 5 SaV GI.2), 7 as SaV GIV genotype, and 1 as GII genotype. SaV infection is present in Shenzhen with GI as the predominant genotype. This is the first report of SaV GIV strains in China, which differs from the strains of Anhui-A141 and Beijing-CHN99/BJ360, suggesting the genotypic variety of SaV infection in China.

  13. Variation in the vernalization response of a geographically diverse collection of timothy genotypes

    DEFF Research Database (Denmark)

    Fiil, Alice; Jensen, Louise Bach; Fjellheim, Siri;

    2011-01-01

    . Genotypes of non-Nordic origin with a strong vernalization response were either di- or tetraploid, whereas respective Nordic genotypes were hexaploid. The ploidy level in relation to vernalization response is discussed. This study clearly demonstrates the presence of considerable genetic variation...

  14. Assessment of Genetic Diversity and Structure of Large Garlic (Allium sativum) Germplasm Bank, by Diversity Arrays Technology "Genotyping-by-Sequencing" Platform (DArTseq).

    Science.gov (United States)

    Egea, Leticia A; Mérida-García, Rosa; Kilian, Andrzej; Hernandez, Pilar; Dorado, Gabriel

    2017-01-01

    Garlic (Allium sativum) is used worldwide in cooking and industry, including pharmacology/medicine and cosmetics, for its interesting properties. Identifying redundancies in germplasm blanks to generate core collections is a major concern, mostly in large stocks, in order to reduce space and maintenance costs. Yet, similar appearance and phenotypic plasticity of garlic varieties hinder their morphological classification. Molecular studies are challenging, due to the large and expected complex genome of this species, with asexual reproduction. Classical molecular markers, like isozymes, RAPD, SSR, or AFLP, are not convenient to generate germplasm core-collections for this species. The recent emergence of high-throughput genotyping-by-sequencing (GBS) approaches, like DArTseq, allow to overcome such limitations to characterize and protect genetic diversity. Therefore, such technology was used in this work to: (i) assess genetic diversity and structure of a large garlic-germplasm bank (417 accessions); (ii) create a core collection; (iii) relate genotype to agronomical features; and (iv) describe a cost-effective method to manage genetic diversity in garlic-germplasm banks. Hierarchical-cluster analysis, principal-coordinates analysis and STRUCTURE showed general consistency, generating three main garlic-groups, mostly determined by variety and geographical origin. In addition, high-resolution genotyping identified 286 unique and 131 redundant accessions, used to select a reduced size germplasm-bank core collection. This demonstrates that DArTseq is a cost-effective method to analyze species with large and expected complex genomes, like garlic. To the best of our knowledge, this is the first report of high-throughput genotyping of a large garlic germplasm. This is particularly interesting for garlic adaptation and improvement, to fight biotic and abiotic stresses, in the current context of climate change and global warming.

  15. Assessment of Genetic Diversity and Structure of Large Garlic (Allium sativum) Germplasm Bank, by Diversity Arrays Technology “Genotyping-by-Sequencing” Platform (DArTseq)

    Science.gov (United States)

    Egea, Leticia A.; Mérida-García, Rosa; Kilian, Andrzej; Hernandez, Pilar; Dorado, Gabriel

    2017-01-01

    Garlic (Allium sativum) is used worldwide in cooking and industry, including pharmacology/medicine and cosmetics, for its interesting properties. Identifying redundancies in germplasm blanks to generate core collections is a major concern, mostly in large stocks, in order to reduce space and maintenance costs. Yet, similar appearance and phenotypic plasticity of garlic varieties hinder their morphological classification. Molecular studies are challenging, due to the large and expected complex genome of this species, with asexual reproduction. Classical molecular markers, like isozymes, RAPD, SSR, or AFLP, are not convenient to generate germplasm core-collections for this species. The recent emergence of high-throughput genotyping-by-sequencing (GBS) approaches, like DArTseq, allow to overcome such limitations to characterize and protect genetic diversity. Therefore, such technology was used in this work to: (i) assess genetic diversity and structure of a large garlic-germplasm bank (417 accessions); (ii) create a core collection; (iii) relate genotype to agronomical features; and (iv) describe a cost-effective method to manage genetic diversity in garlic-germplasm banks. Hierarchical-cluster analysis, principal-coordinates analysis and STRUCTURE showed general consistency, generating three main garlic-groups, mostly determined by variety and geographical origin. In addition, high-resolution genotyping identified 286 unique and 131 redundant accessions, used to select a reduced size germplasm-bank core collection. This demonstrates that DArTseq is a cost-effective method to analyze species with large and expected complex genomes, like garlic. To the best of our knowledge, this is the first report of high-throughput genotyping of a large garlic germplasm. This is particularly interesting for garlic adaptation and improvement, to fight biotic and abiotic stresses, in the current context of climate change and global warming. PMID:28775737

  16. Assessment of Genetic Diversity and Structure of Large Garlic (Allium sativum Germplasm Bank, by Diversity Arrays Technology “Genotyping-by-Sequencing” Platform (DArTseq

    Directory of Open Access Journals (Sweden)

    Leticia A. Egea

    2017-07-01

    Full Text Available Garlic (Allium sativum is used worldwide in cooking and industry, including pharmacology/medicine and cosmetics, for its interesting properties. Identifying redundancies in germplasm blanks to generate core collections is a major concern, mostly in large stocks, in order to reduce space and maintenance costs. Yet, similar appearance and phenotypic plasticity of garlic varieties hinder their morphological classification. Molecular studies are challenging, due to the large and expected complex genome of this species, with asexual reproduction. Classical molecular markers, like isozymes, RAPD, SSR, or AFLP, are not convenient to generate germplasm core-collections for this species. The recent emergence of high-throughput genotyping-by-sequencing (GBS approaches, like DArTseq, allow to overcome such limitations to characterize and protect genetic diversity. Therefore, such technology was used in this work to: (i assess genetic diversity and structure of a large garlic-germplasm bank (417 accessions; (ii create a core collection; (iii relate genotype to agronomical features; and (iv describe a cost-effective method to manage genetic diversity in garlic-germplasm banks. Hierarchical-cluster analysis, principal-coordinates analysis and STRUCTURE showed general consistency, generating three main garlic-groups, mostly determined by variety and geographical origin. In addition, high-resolution genotyping identified 286 unique and 131 redundant accessions, used to select a reduced size germplasm-bank core collection. This demonstrates that DArTseq is a cost-effective method to analyze species with large and expected complex genomes, like garlic. To the best of our knowledge, this is the first report of high-throughput genotyping of a large garlic germplasm. This is particularly interesting for garlic adaptation and improvement, to fight biotic and abiotic stresses, in the current context of climate change and global warming.

  17. Detection of Common, Emerging and Uncommon VP4, and VP7 Human Group A Rotavirus Genotypes from Urban Sewage Samples in Uruguay.

    Science.gov (United States)

    Tort, Luis Fernando Lopez; Victoria, Matías; Lizasoain, Andrés; García, Mariana; Berois, Mabel; Cristina, Juan; Leite, José Paulo Gagliardi; Gómez, Mariela Martínez; Miagostovich, Marize Pereira; Colina, Rodney

    2015-12-01

    Environmental approach has proven to be a useful tool for epidemiological studies demonstrating through environmental studies the diversity of viruses circulating in a given population. The aim of this study was to perform a phylogenetic characterization of the group A rotavirus (RVA) glycoprotein (G)- and protease-sensitive (P)-genotypes obtained from sewage samples (n = 116) collected in six cities of Uruguay during March 2011 to April 2013. A worldwide standardized semi-nested multiplex RT-PCR (SNM RT-PCR) protocol directed against VP4 and VP7 genes were conducted for RVA detection and consensual DNA fragments were submitted to nucleotide sequencing. P and/or G genotype was successfully determined by phylogenetic analysis in 61% (n = 37) of the positive samples obtained by SNM RT-PCR (n = 61). The RVA genotypes were as follow: G1 (n = 2), G2 (n = 14), G3 (n = 5), G12 (n = 2), P[4] (n = 4), P[8] (n = 16), and P[3] (n = 2). Interestingly, through phylogenetic analysis, emerging, and uncommon human genotypes could be detected. Results obtained from the comparison of RVA genotypes detected in the current study and Uruguayan RVA strains previously described for contemporary clinical pediatric cases showed that monitoring sewage may be a good screening option for a rapid and economical overview of the circulating genotypes in the surrounding human population and a useful approximation to study RVA epidemiology in a future vaccine monitoring program. The present study represents the first report in Uruguay that describes the phylogenetic diversity of RVA from urban sewage samples.

  18. Using Neighbor Diversity to Detect Fraudsters in Online Auctions

    Directory of Open Access Journals (Sweden)

    Jun-Lin Lin

    2014-05-01

    Full Text Available Online auctions attract not only legitimate businesses trying to sell their products but also fraudsters wishing to commit fraudulent transactions. Consequently, fraudster detection is crucial to ensure the continued success of online auctions. This paper proposes an approach to detect fraudsters based on the concept of neighbor diversity. The neighbor diversity of an auction account quantifies the diversity of all traders that have transactions with this account. Based on four different features of each trader (i.e., the number of received ratings, the number of cancelled transactions, k-core, and the joined date, four measurements of neighbor diversity are proposed to discern fraudsters from legitimate traders. An experiment is conducted using data gathered from a real world auction website. The results show that, although the use of neighbor diversity on k-core or on the joined date shows little or no improvement in detecting fraudsters, both the neighbor diversity on the number of received ratings and the neighbor diversity on the number of cancelled transactions improve classification accuracy, compared to the state-of-the-art methods that use k-core and center weight.

  19. Identification of genome-wide copy number variations among diverse pig breeds using SNP genotyping arrays.

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    Jiying Wang

    Full Text Available Copy number variations (CNVs are important forms of genetic variation complementary to SNPs, and can be considered as promising markers for some phenotypic and economically important traits or diseases susceptibility in domestic animals. In the present study, we performed a genome-wide CNV identification in 14 individuals selected from diverse populations, including six types of Chinese indigenous breeds, one Asian wild boar population, as well as three modern commercial foreign breeds. We identified 63 CNVRs in total, which covered 9.98 Mb of polymorphic sequence and corresponded to 0.36% of the genome sequence. The length of these CNVRs ranged from 3.20 to 827.21 kb, with an average of 158.37 kb and a median of 97.85 kb. Functional annotation revealed these identified CNVR have important molecular function, and may play an important role in exploring the genetic basis of phenotypic variability and disease susceptibility among pigs. Additionally, to confirm these potential CNVRs, we performed qPCR for 12 randomly selected CNVRs and 8 of them (66.67% were confirmed successfully. CNVs detected in diverse populations herein are essential complementary to the CNV map in the pig genome, which provide an important resource for studies of genomic variation and the association between various economically important traits and CNVs.

  20. Diversity of microcystin-producing genotypes in Brazilian strains of Microcystis (Cyanobacteria

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    MC Bittencourt-Oliveira

    Full Text Available Microcystis Kützing ex Lemmermann is among the genera of cyanobacteria often associated to toxic blooms with the release of microcystins. A gene cluster codes for microcystin synthetases, which are involved in the biosynthesis of this toxin. The aim of the present study was to investigate the genetic diversity of the mcyB gene, specifically the B1 module, in Brazilian strains of Microcystis spp. and its microcystin variants. Broad genetic diversity was revealed in this region. From the phylogenetic analysis, three clusters were obtained that were not related to the geographic origin or morphospecies of the strains, nor with the variant of the microcystin produced. A group of strains that did not produce microcystins was found, despite the presence of the mcyB1 fragment. Eight microcystin isoforms were detected: MC-LR, [D-Asp³]-MC-LR, [Asp³]-MC-LR, MC-RR, [Dha7]-MC-LR, MC-LF, MC-LW and [D-Asp³, EtAdda5]-MC-LH, the latter of which is described for the first time in Brazil. Moreover, five other variants were not identified and indicate being new.

  1. Development of T m -shift genotyping method for detection of cat-derived Giardia lamblia.

    Science.gov (United States)

    Pan, Weida; Fu, Yeqi; Abdullahi, Auwalu Yusuf; Wang, Mingwei; Shi, Xianli; Yang, Fang; Yu, Xingang; Yan, Xinxin; Zhang, Pan; Hang, Jianxiong; Li, Guoqing

    2017-04-01

    To develop T m -shift genotyping method for detection of cat-derived Giardia lamblia, two sets of primers with two GC-rich tails of unequal length attached to their 5'-end were designed according to two SNPs (BG434 and BG170) of β-giardin (bg) gene, and specific PCR products were identified by inspection of a melting curve on real-time PCR thermocycler. A series of experiments on the stability, sensitivity, and accuracy of T m -shift method was tested, and clinical samples were also detected. The results showed that two sets of primers based on SNP could distinguish accurately between assemblages A and F. Coefficient of variation of T m values of assemblage A and F was 0.14 and 0.07% in BG434 and 0.10 and 0.11% in BG170, respectively. The lowest detection concentration was 4.52 × 10(-5) and 4.88 × 10(-5) ng/μL samples of assemblage A and F standard plasmids. The T m -shift genotyping results of ten DNA samples from the cat-derived G. lamblia were consistent with their known genotypes. The detection rate of clinical samples by T m -shift was higher than that by microscopy, and their genotyping results were in complete accordance with sequencing results. It is concluded that the T m -shift genotyping method is rapid, specific, and sensitive and may provide a new technological mean for molecular detection and epidemiological investigation of the cat-derived G. lamblia.

  2. Detection and differentiation of Japanese encephalitis virus genotype I and genotype III by reverse transcription loop-mediated isothermal amplification combined with restriction fragment length polymorphism.

    Science.gov (United States)

    Zhang, Liang; Cao, Sanjie; Wu, Rui; Zhu, Shuquan; Liu, Hanyang; Yuan, Lei; Shi, Shuangyan; Zhang, Dan; Huang, Xiaobo; Wen, Xintian; Wen, Yiping; Yan, Qigui; Huang, Yong; Ma, Xiaoping

    2015-04-01

    Japanese encephalitis (JE), which is a mosquito-borne arboviral infection, is the leading cause of viral encephalitis in Asian countries. The causative agent of JE is Japanese encephalitis virus (JEV), in which the predominant genotype has changed from genotype III (G III) to genotype I (G I). However, a method for the rapid differentiation between JEV G I and G III remains unavailable. This study aimed to establish a rapid JEV genotyping method using reverse transcription loop-mediated isothermal amplification (RT-LAMP). An Spe I site, which was located in the target sequence (C gene) of JEV G III strains but not in JEV G I strains, was selected as the RT-LAMP target. After testing 64 specimens, results showed that RT-LAMP can detect and differentiate JEV G I and G III specifically. Thus, a novel RT-LAMP system for the rapid detection and differentiation of JEV G I and G III was developed successfully.

  3. Genotypic and phenotypic diversity of Ralstonia pickettii and Ralstonia insidiosa isolates from clinical and environmental sources including High-purity Water. Diversity in Ralstonia pickettii

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    Adley Catherine C

    2011-08-01

    Full Text Available Abstract Background Ralstonia pickettii is a nosocomial infectious agent and a significant industrial contaminant. It has been found in many different environments including clinical situations, soil and industrial High Purity Water. This study compares the phenotypic and genotypic diversity of a selection of strains of Ralstonia collected from a variety of sources. Results Ralstonia isolates (fifty-nine from clinical, industrial and environmental origins were compared genotypically using i Species-specific-PCR, ii PCR and sequencing of the 16S-23S rRNA Interspatial region (ISR iii the fliC gene genes, iv RAPD and BOX-PCR and v phenotypically using biochemical testing. The species specific-PCR identified fifteen out of fifty-nine designated R. pickettii isolates as actually being the closely related species R. insidiosa. PCR-ribotyping of the 16S-23S rRNA ISR indicated few major differences between the isolates. Analysis of all isolates demonstrated different banding patterns for both the RAPD and BOX primers however these were found not to vary significantly. Conclusions R. pickettii species isolated from wide geographic and environmental sources appear to be reasonably homogenous based on genotypic and phenotypic characteristics. R. insidiosa can at present only be distinguished from R. pickettii using species specific PCR. R. pickettii and R. insidiosa isolates do not differ significantly phenotypically or genotypically based on environmental or geographical origin.

  4. Easy and fast detection and genotyping of high-risk human papillomavirus by dedicated DNA microarrays.

    Science.gov (United States)

    Albrecht, Valérie; Chevallier, Anne; Magnone, Virginie; Barbry, Pascal; Vandenbos, Fanny; Bongain, André; Lefebvre, Jean-Claude; Giordanengo, Valérie

    2006-11-01

    Persistent cervical high-risk human papillomavirus (HPV) infection is correlated with an increased risk of developing a high-grade cervical intraepithelial lesion. A two-step method was developed for detection and genotyping of high-risk HPV. DNA was firstly amplified by asymmetrical PCR in the presence of Cy3-labelled primers and dUTP. Labelled DNA was then genotyped using DNA microarray hybridization. The current study evaluated the technical efficacy of laboratory-designed HPV DNA microarrays for high-risk HPV genotyping on 57 malignant and non-malignant cervical smears. The approach was evaluated for a broad range of cytological samples: high-grade squamous intraepithelial lesions (HSIL), low-grade squamous intraepithelial lesions (LSIL) and atypical squamous cells of high-grade (ASC-H). High-risk HPV was also detected in six atypical squamous cells of undetermined significance (ASC-US) samples; among them only one cervical specimen was found uninfected, associated with no histological lesion. The HPV oligonucleotide DNA microarray genotyping detected 36 infections with a single high-risk HPV type and 5 multiple infections with several high-risk types. Taken together, these results demonstrate the sensitivity and specificity of the HPV DNA microarray approach. This approach could improve clinical management of patients with cervical cytological abnormalities.

  5. APPLICATION OF RYE SSR MARKERS FOR DETECTION OF GENETIC DIVERSITY IN TRITICALE

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    Želmíra Balážová

    2016-06-01

    Full Text Available Present study aims to testify usefulness of particular rye SSR markers for the detection of genetic diversity degree in the set of 20 triticale cultivars coming from different European countries. For this purpose, a set of six rye SSR markers were used. The set of six polymorphic markers provided 22 alleles with an average frequency of 3.67 alleles per locus. The number of alleles ranged between 2 (SCM43 and 5 (SCM28, SCM86. Resulting from the number and frequency of alleles diversity index (DI, polymorphic information content (PIC and probability of identity (PI were calculated. An average value of PIC for 6 SSR markers was 0.505, the highest value was calculated for rye SSR marker SCM86 (0.706. Based on UPGMA algorithm, a dendrogram was constructed. In dendrogram cultivars were divided into two main clusters. The first cluster contained two cultivars, Russian cultivar Greneder and Slovak cultivar Largus, and second included 18 cultivars. Genetically the closest were two Greek cultivars (Niobi and Thisbi and were close to other Greek cultivar Vrodi. It was possible to separate triticale cultivars of spring and winter form in dendrogram. Results showed the utility of rye microsatellite markers for estimation of genetic diversity of European triticale genotypes leading to genotype identification.

  6. Study of Mycobacterium tuberculosis complex genotypic diversity in Malaysia reveals a predominance of ancestral East-African-Indian lineage with a Malaysia-specific signature

    National Research Council Canada - National Science Library

    Ismail, Fazli; Couvin, David; Farakhin, Izzah; Abdul Rahman, Zaidah; Rastogi, Nalin; Suraiya, Siti

    2014-01-01

    .... This study intended a first assessment of spoligotyping-based MTBC genotypic diversity in Malaysia followed by a comparison of strains with those prevailing in neighboring countries by comparison...

  7. High Diversity of vacA and cagA Helicobacter pylori Genotypes in Patients with and without Gastric Cancer

    Science.gov (United States)

    López-Vidal, Yolanda; Ponce-de-León, Sergio; Castillo-Rojas, Gonzalo; Barreto-Zúñiga, Rafael; Torre-Delgadillo, Aldo

    2008-01-01

    Background Helicobacter pylori is associated with chronic gastritis, peptic ulcers, and gastric cancer. The aim of this study was to assess the topographical distribution of H. pylori in the stomach as well as the vacA and cagA genotypes in patients with and without gastric cancer. Methodology/Principal Findings Three gastric biopsies, from predetermined regions, were evaluated in 16 patients with gastric cancer and 14 patients with dyspeptic symptoms. From cancer patients, additional biopsy specimens were obtained from tumor centers and margins; among these samples, the presence of H. pylori vacA and cagA genotypes was evaluated. Positive H. pylori was 38% and 26% in biopsies obtained from the gastric cancer and non-cancer groups, respectively (p = 0.008), and 36% in tumor sites. In cancer patients, we found a preferential distribution of H. pylori in the fundus and corpus, whereas, in the non-cancer group, the distribution was uniform (p = 0.003). A majority of the biopsies were simultaneously cagA gene-positive and -negative. The fundus and corpus demonstrated a higher positivity rate for the cagA gene in the non-cancer group (p = 0.036). A mixture of cagA gene sizes was also significantly more frequent in this group (p = 0.003). Ninety-two percent of all the subjects showed more than one vacA gene genotype; s1b and m1 vacA genotypes were predominantly found in the gastric cancer group. The highest vacA-genotype signal-sequence diversity was found in the corpus and 5 cm from tumor margins. Conclusion/Significance High H. pylori colonization diversity, along with the cagA gene, was found predominantly in the fundus and corpus of patients with gastric cancer. The genotype diversity observed across systematic whole-organ and tumor sampling was remarkable. We find that there is insufficient evidence to support the association of one isolate with a specific disease, due to the multistrain nature of H. pylori infection shown in this work. PMID:19050763

  8. High diversity of vacA and cagA Helicobacter pylori genotypes in patients with and without gastric cancer.

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    Yolanda López-Vidal

    Full Text Available BACKGROUND: Helicobacter pylori is associated with chronic gastritis, peptic ulcers, and gastric cancer. The aim of this study was to assess the topographical distribution of H. pylori in the stomach as well as the vacA and cagA genotypes in patients with and without gastric cancer. METHODOLOGY/PRINCIPAL FINDINGS: Three gastric biopsies, from predetermined regions, were evaluated in 16 patients with gastric cancer and 14 patients with dyspeptic symptoms. From cancer patients, additional biopsy specimens were obtained from tumor centers and margins; among these samples, the presence of H. pylori vacA and cagA genotypes was evaluated. Positive H. pylori was 38% and 26% in biopsies obtained from the gastric cancer and non-cancer groups, respectively (p = 0.008, and 36% in tumor sites. In cancer patients, we found a preferential distribution of H. pylori in the fundus and corpus, whereas, in the non-cancer group, the distribution was uniform (p = 0.003. A majority of the biopsies were simultaneously cagA gene-positive and -negative. The fundus and corpus demonstrated a higher positivity rate for the cagA gene in the non-cancer group (p = 0.036. A mixture of cagA gene sizes was also significantly more frequent in this group (p = 0.003. Ninety-two percent of all the subjects showed more than one vacA gene genotype; s1b and m1 vacA genotypes were predominantly found in the gastric cancer group. The highest vacA-genotype signal-sequence diversity was found in the corpus and 5 cm from tumor margins. CONCLUSION/SIGNIFICANCE: High H. pylori colonization diversity, along with the cagA gene, was found predominantly in the fundus and corpus of patients with gastric cancer. The genotype diversity observed across systematic whole-organ and tumor sampling was remarkable. We find that there is insufficient evidence to support the association of one isolate with a specific disease, due to the multistrain nature of H. pylori infection shown in this work.

  9. Genetic diversity of Toxoplasma gondii isolates in Egyptian feral cats reveals new genotypes.

    Science.gov (United States)

    Al-Kappany, Y M; Rajendran, C; Abu-Elwafa, S A; Hilali, M; Su, C; Dubey, J P

    2010-12-01

    Cats are important in the epidemiology of Toxoplasma gondii because they are the only hosts that excrete environmentally resistant oocysts in feces. In the present study, 115 viable T. gondii isolates from tissues of cats from Egypt were genotyped using 10 PCR-restriction fragment length polymorphism markers (SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico) and DNA from tachyzoites. Seven genotypes were recognized including the clonal Type II, Type III (2 genotypes), and 4 atypical genotypes. Ninety percent (103 of 115) of isolates were clonal, i.e., Type II (n  =  61) and Type III (n  =  42) strains. Of the 61 Type II strains, all had the Type II alleles at all loci, except for 2 strains that had allele I at Apico. Eight isolates were divided into 4 atypical genotypes. One of these genotypes (with 4 isolates) was previously reported in dogs from Sri Lanka and in sand cats from the United Arab Emirates. Four isolates had mixed infections. These results revealed a strong clonal population structure with the dominance of clonal Type II and III lineages of T. gondii in feral cats from Egypt.

  10. Genotypic Diversity of Mycobacterium tuberculosis Clinical Isolates in the Multiethnic Area of the Xinjiang Uygur Autonomous Region in China

    Science.gov (United States)

    Liu, Jie; Li, Junlian; Liu, Jiao; Lian, Lulu; Lu, Bing; Yu, Qin; Zhang, Jingrui; Qi, Yingcheng

    2017-01-01

    Objectives. We studied the genetic diversity of clinical isolates from patients with tuberculosis in the multiethnic area of Xinjiang autonomous region in China. A total of 311 clinical M. tuberculosis isolates were collected in 2006 and 2011 and genotyped by two genotyping methods. All isolates were grouped into 68 distinct spoligotypes using the spoligotyping method. The Beijing family was dominant, followed by T1 and CAS. MIRU-VNTR results showed that a total of 195 different VNTR types were identified. Ten of the 15 loci were highly or moderately discriminant according to their HGDI scores, and 13 loci had good discriminatory power in non-Beijing family strains, whereas only two loci had good discriminatory power in Beijing family strains. Chi-square tests demonstrated that there were no correlations between four characteristics (sex, age, type of case, and treatment history) and the Beijing family. In summary, Beijing family strains were predominant in Xinjiang, and the VNTR-15China locus-set was suitable for genotyping all Xinjiang strains, but not for the Beijing family strains. Thus, these data suggested that different genotype distributions may exist in different regions; MLVA locus-sets should be adjusted accordingly, with newly added loci to increase resolution if necessary.

  11. HPV detection methods and genotyping techniques in screening for cervical cancer.

    Science.gov (United States)

    Eide, Maj Liv; Debaque, Hervé

    2012-12-01

    Human papillomaviruses (HPV), double-stranded DNA viruses, are causing many mucocutaneous diseases, benign or malignant, ranging from common warts to malignancies involving the upper aerodigestive tract and the anogenital sphere. The diagnosis of HPV infection is based primarily on the viral genome detection by molecular biological methods, given the difficulty in routine cultivation of these viruses. The current trend in screening against cervical cancer is to improve the sensitivity of screening with new methods and to propose new algorithms for diagnostic and therapeutic decisions. The development of liquid-based cytology facilitates the cytologic diagnosis and molecular assays from the same sample. There are two main types of HPV detection methods used on uterine cervical samples: signal amplification methods (hybridization techniques in liquid phase) and target amplification methods (the techniques of gene amplification or Polymerase Chain Reaction [PCR]). Genotyping techniques are also developed: they are based on an amplification technique followed by hybridization with probe specific types. In addition to the detection, genotyping techniques allow quantitative detection of viral DNA of HPV genotype and so monitor changes in viral load over time. Another approach relies on the detection of messenger RNA (mRNA) of HPV proteins E6 and E7 oncogenes, which would appear to be a relevant marker to identify and monitor women at risk of progression to a precancerous lesion or cervical cancer.

  12. Hepatitis C Virus Genotype Diversity among Intravenous Drug Users in Yunnan Province, Southwestern China

    Science.gov (United States)

    Wu, Wenlong; Feng, Ruilin; Wu, Zhongxiang; Cun, Wei; Dong, Shaozhong

    2013-01-01

    Background Recently, high proportions (15.6%–98.7%) of intravenous drug users (IDUs) in China were found to be positive for hepatitis C virus (HCV). Yunnan Province is located in southwestern China and borders one of the world's most important opium-producing regions, thus it is an important drug trafficking route to other regions of China. Methodology/Principal Findings Here, we assessed 100 HCV-positive plasma samples from IDUs who were enrolled through the Kunming Center for Disease Control and Prevention in 2012. HCV C/E1 fragments were PCR-amplified and sequenced. We identified eight HCV subtypes (1a, 1b, 3a, 3b, 6a, 6n, 6u and 6v), of which genotype 6 was most predominant (frequency, 47%) followed by genotypes 3 (41%) and 1 (12%). HCV subtypes 6n (30%) and 3b (29%) were most common and were identified in 59% of the IDUs. We compared HCV genotypes among IDUs in Yunnan Province with those from other regions and found that the distribution patterns of HCV genotypes in Yunnan Province were similar to those in southern China, but different from those in eastern China. However, the distribution patterns of HCV subtypes varied among Yunnan Province and southern China, despite the shared similar genotypes. A comparison of the current data with those previously reported showed that the frequency of HCV genotype 6 increased from 25% to 47% within 5 years, especially subtypes 6a (5% to 15%) and 6n (11.2% to 30%). In contrast, the frequencies of subtypes 3b and 1b decreased by almost 50% within 5 years. Conclusion/Significance Our results provided further information to support the assertion that drug trafficking routes influence HCV transmission patterns among IDUs in Yunnan Province. The frequency of HCV genotypes and subtypes changed rapidly among IDUs in Yunnan Province and subtypes 6a and 6n may have originated in Vietnam and Myanmar, respectively. PMID:24358211

  13. Hepatitis C virus genotype diversity among intravenous drug users in Yunnan Province, Southwestern China.

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    Zhihui Zhang

    Full Text Available BACKGROUND: Recently, high proportions (15.6%-98.7% of intravenous drug users (IDUs in China were found to be positive for hepatitis C virus (HCV. Yunnan Province is located in southwestern China and borders one of the world's most important opium-producing regions, thus it is an important drug trafficking route to other regions of China. METHODOLOGY/PRINCIPAL FINDINGS: Here, we assessed 100 HCV-positive plasma samples from IDUs who were enrolled through the Kunming Center for Disease Control and Prevention in 2012. HCV C/E1 fragments were PCR-amplified and sequenced. We identified eight HCV subtypes (1a, 1b, 3a, 3b, 6a, 6n, 6u and 6v, of which genotype 6 was most predominant (frequency, 47% followed by genotypes 3 (41% and 1 (12%. HCV subtypes 6n (30% and 3b (29% were most common and were identified in 59% of the IDUs. We compared HCV genotypes among IDUs in Yunnan Province with those from other regions and found that the distribution patterns of HCV genotypes in Yunnan Province were similar to those in southern China, but different from those in eastern China. However, the distribution patterns of HCV subtypes varied among Yunnan Province and southern China, despite the shared similar genotypes. A comparison of the current data with those previously reported showed that the frequency of HCV genotype 6 increased from 25% to 47% within 5 years, especially subtypes 6a (5% to 15% and 6n (11.2% to 30%. In contrast, the frequencies of subtypes 3b and 1b decreased by almost 50% within 5 years. CONCLUSION/SIGNIFICANCE: Our results provided further information to support the assertion that drug trafficking routes influence HCV transmission patterns among IDUs in Yunnan Province. The frequency of HCV genotypes and subtypes changed rapidly among IDUs in Yunnan Province and subtypes 6a and 6n may have originated in Vietnam and Myanmar, respectively.

  14. Genetic History of Hepatitis C Virus in Venezuela: High Diversity and Long Time of Evolution of HCV Genotype 2

    Science.gov (United States)

    Sulbarán, Maria Z.; Di Lello, Federico A.; Sulbarán, Yoneira; Cosson, Clarisa; Loureiro, Carmen L.; Rangel, Héctor R.; Cantaloube, Jean F.; Campos, Rodolfo H.; Moratorio, Gonzalo; Cristina, Juan; Pujol, Flor H.

    2010-01-01

    Background The subtype diversity of the hepatitis C virus (HCV) genotypes is unknown in Venezuela. Methodology/Principal Findings Partial sequencing of the NS5B region was performed in 310 isolates circulating in patients from 1995 to 2007. In the samples collected between 2005 and 2007, HCV genotype 1 (G1) was the most common genotype (63%), composed as expected of mainly G1a and G1b. G2 was the second most common genotype (33%), being G2a almost absent and G2j the most frequent subtype. Sequence analysis of the core region confirmed the subtype assignment performed within the NS5b region in 63 isolates. The complete genome sequence of G2j was obtained. G2j has been described in France, Canada and Burkina Fasso, but it was not found in Martinique, where several subtypes of G2 circulate in the general population. Bayesian coalescence analysis indicated a most recent common ancestor (MRCA) of G2j around 1785, before the introduction of G1b (1869) and G1a (1922). While HCV G1a and G1b experienced a growth reduction since 1990, coincident with the time when blood testing was implemented in Venezuela, HCV G2j did not seem to reach growth equilibrium during this period. Conclusions/Significance Assuming the introduction of G2j from Africa during the slave trade, the high frequency of G2j found in Venezuela could suggest: 1- the introduction of African ethnic groups different from the ones introduced to Martinique or 2- the occurrence of a founder effect. This study represents an in-depth analysis of the subtype diversity of HCV in Venezuela, which is still unexplored in the Americas and deserves further studies. PMID:21179440

  15. Colorimetric Detection of 23 Human Papillomavirus Genotypes by Loop-Mediated Isothermal Amplification.

    Science.gov (United States)

    Lin, Junxiao; Ma, Biao; Fang, Jiehong; Wang, Ye; He, Haizhen; Lin, Wei; Su, Wei; Zhang, Mingzhou

    2017-03-01

    Human papillomavirus (HPV) infection is linked to cervical cancer. With the technological development of molecular biology and epidemiology, detection and treatment of HPV has become an important mean to prevent cervical cancer. A simple, rapid, and sensitive colorimetric loop-mediated isothermal amplification (LAMP) method was established herein to detect 23 HPV genotypes. The sequences of the primers for the LAMP reaction were located in the L1 gene of the HPV genome. As it is a fluorescent dye, calcein was added before the reaction. The reaction was run under isothermal conditions at 65°C for 40 minutes. A positive reaction was indicated by a color change from yellow to fluorescent green. The fluorescence curve diagram represents the monitoring of real time quantitative instrument. 450 cervical swab samples from patients with single infections of 23 different HPV genotypes were examined to evaluate the specificity. The results revealed no cross-reaction with other HPV genotypes. A serial dilution of a cloned plasmid containing 23 HPV L1 gene sequences was employed to evaluate the sensitivity. Different HPV subtypes have different detection capability. The sensitivity of different HPV subtypes tested by LAMP assay was in the range from 1.0 x10 to 4.0 x 103 copies per reaction. The LAMP assay and the RDB (reverse dot blot) were compared for detecting and genotyping HPV among the 450 clinical samples. There were 385 (85.6%) and 375 (83.3%) HPV positive specimens detected by LAMP and RDB, respectively, as well as 306 (68.0%) and 296 (65.8%) for HR-HPV positive specimens. The agreement between the LAMP and RDB assays was 93.3% (κ = 0.75) for HPV positivity and 94.7% (κ = 0.88) for HR-HPV positivity. It was concluded that this colorimetric LAMP assay had potential application for the rapid screening of the HPV infection in resource-limited hospitals or rural clinics.

  16. Highly specific and sensitive electrochemical genotyping via gap ligation reaction and surface hybridization detection.

    Science.gov (United States)

    Huang, Yong; Zhang, Yan-Li; Xu, Xiangmin; Jiang, Jian-Hui; Shen, Guo-Li; Yu, Ru-Qin

    2009-02-25

    This paper developed a novel electrochemical genotyping strategy based on gap ligation reaction with surface hybridization detection. This strategy utilized homogeneous enzymatic reactions to generate molecular beacon-structured allele-specific products that could be cooperatively annealed to capture probes stably immobilized on the surface via disulfide anchors, thus allowing ultrasensitive surface hybridization detection of the allele-specific products through redox tags in close proximity to the electrode. Such a unique biphasic architecture provided a universal methodology for incorporating enzymatic discrimination reactions in electrochemical genotyping with desirable reproducibility, high efficiency and no interferences from interficial steric hindrance. The developed technique was demonstrated to show intrinsic high sensitivity for direct genomic analysis, and excellent specificity with discriminativity of single nucleotide variations.

  17. Mycobacterium tuberculosis Complex Genotype Diversity and Drug Resistance Profiles in a Pediatric Population in Mexico

    Directory of Open Access Journals (Sweden)

    Mercedes Macías Parra

    2011-01-01

    Full Text Available The aim of this study was to determine the frequency of drug resistance and the clonality of genotype patterns in M. tuberculosis clinical isolates from pediatric patients in Mexico (n=90 patients from 19 states; time period—January 2002 to December 2003. Pulmonary disease was the most frequent clinical manifestation (71%. Children with systemic tuberculosis (TB were significantly younger compared to patients with localized TB infections (mean 7.7±6.2 years versus 15±3.4 years P=0.001. Resistance to any anti-TB drug was detected in 24/90 (26.7% of the isolates; 21/90 (23.3% and 10/90 (11.1% were resistant to Isoniazid and Rifampicin, respectively, and 10/90 (11.1% strains were multidrug-resistant (MDR. Spoligotyping produced a total of 55 different patterns; 12/55 corresponded to clustered isolates (n=47, clustering rate of 52.2%, and 43/55 to unclustered isolates (19 patterns were designated as orphan by the SITVIT2 database. Database comparison led to designation of 36 shared types (SITs; 32 SITs (n=65 isolates matched a preexisting shared type in SITVIT2, whereas 4 SITs (n=6 isolates were newly created. Lineage classification based on principal genetic groups (PGG revealed that 10% of the strains belonged to PGG1 (Bovis and Manu lineages. Among PGG2/3 group, the most predominant clade was the Latin-American and Mediterranean (LAM in 27.8% of isolates, followed by Haarlem and T lineages. The number of single drug-resistant (DR and multidrug-resistant (MDR-TB isolates in this study was similar to previously reported in studies from adult population with risk factors. No association between the spoligotype, age, region, or resistance pattern was observed. However, contrary to a study on M. tuberculosis spoligotyping in Acapulco city that characterized a single cluster of SIT19 corresponding to the EAI2-Manila lineage in 70 (26% of patients, not a single SIT19 isolate was found in our pediatric patient population. Neither did we find any

  18. Mycobacterium tuberculosis Complex Genotype Diversity and Drug Resistance Profiles in a Pediatric Population in Mexico

    Science.gov (United States)

    Macías Parra, Mercedes; Kumate Rodríguez, Jesús; Arredondo García, José Luís; López-Vidal, Yolanda; Castañón-Arreola, Mauricio; Balandrano, Susana; Rastogi, Nalin; Gutiérrez Castrellón, Pedro

    2011-01-01

    The aim of this study was to determine the frequency of drug resistance and the clonality of genotype patterns in M. tuberculosis clinical isolates from pediatric patients in Mexico (n = 90 patients from 19 states; time period—January 2002 to December 2003). Pulmonary disease was the most frequent clinical manifestation (71%). Children with systemic tuberculosis (TB) were significantly younger compared to patients with localized TB infections (mean 7.7 ± 6.2 years versus 15 ± 3.4 years P = 0.001). Resistance to any anti-TB drug was detected in 24/90 (26.7%) of the isolates; 21/90 (23.3%) and 10/90 (11.1%) were resistant to Isoniazid and Rifampicin, respectively, and 10/90 (11.1%) strains were multidrug-resistant (MDR). Spoligotyping produced a total of 55 different patterns; 12/55 corresponded to clustered isolates (n = 47, clustering rate of 52.2%), and 43/55 to unclustered isolates (19 patterns were designated as orphan by the SITVIT2 database). Database comparison led to designation of 36 shared types (SITs); 32 SITs (n = 65 isolates) matched a preexisting shared type in SITVIT2, whereas 4 SITs (n = 6 isolates) were newly created. Lineage classification based on principal genetic groups (PGG) revealed that 10% of the strains belonged to PGG1 (Bovis and Manu lineages). Among PGG2/3 group, the most predominant clade was the Latin-American and Mediterranean (LAM) in 27.8% of isolates, followed by Haarlem and T lineages. The number of single drug-resistant (DR) and multidrug-resistant (MDR-TB) isolates in this study was similar to previously reported in studies from adult population with risk factors. No association between the spoligotype, age, region, or resistance pattern was observed. However, contrary to a study on M. tuberculosis spoligotyping in Acapulco city that characterized a single cluster of SIT19 corresponding to the EAI2-Manila lineage in 70 (26%) of patients, not a single SIT19 isolate was found in our pediatric patient population. Neither did we

  19. The Genetic Diversity and Phylogenetic Characteritics of Rotavirus VP4(P Genotypes in Children With Acute Diarrhea

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    Haghshenas Z

    2011-11-01

    Full Text Available Background: Acute gastroenteritis is a major cause of morbidity and mortality among children in developing countries. Rotaviruses are recognized as the most common etiologic factors of gastroenteritis. In this study, we determined the epidemiologic features, clinical symptoms and molecular structure of rotavirus VP4(P genotypes in children with acute diarrhea in Bahrami Hospital in Tehran Iran, during 2009 for justifying the routine use of rotavirus vaccines in children. Methods: One hundred fifty fecal samples from 150 children with acute diarrhea in Bahrami Pediatric Hospital in Tehran, Iran were collected from January to December 2009. The patients’ mean age was 20.90+18.19 years (ranging from 1 month to 14 years. Fecal samples were transported on ice to the laboratory of virology department of Pasture Institute of Iran. The demographic and clinical data for each case were entered in an author-devised questionnaire. Group A rotavirus was detected by dsRNA-PAGE. Subsequently, rotavirus genotyping (VP4 was performed by semi-nested multiple RT-PCR and the phylogenetic tree of the Rotavirus nucleotides was constructed. The data were analyzed by statistical tests including Wilcoxon signed and Mann-Whitney U. Results: Rotavirus was isolated in 19.3% of the samples, more than 90% of which had long RNA patterns. The predominant genotype (VP4 was P[8] (86% and other genotypes respectively were P[6] (6.9% and P[4] (6.9%. Conclusion: A high prevalence of the P[8] genotype was found to be the cause of acute diarrhea. The analysis of P[8] genotype sequence showed a high level of similarity of the virus in this study with those of other Asian countries.

  20. Comparative transcriptional analysis of human macrophages exposed to animal and human isolates of Mycobacterium avium subspecies paratuberculosis with diverse genotypes.

    Science.gov (United States)

    Motiwala, Alifiya S; Janagama, Harish K; Paustian, Michael L; Zhu, Xiaochun; Bannantine, John P; Kapur, Vivek; Sreevatsan, Srinand

    2006-11-01

    Mycobacterium avium subsp. paratuberculosis is the causative agent of Johne's disease in animals and has been hypothesized to be associated with Crohn's disease in humans. Recently, M. avium subsp. paratuberculosis isolates recovered from Crohn's disease patients were shown to have limited diversity, implying the existence of human disease-associated genotypes and strain sharing with animals (A. H. Ghadiali et al., J. Clin. Microbiol. 42:5345-5348, 2004). To explore whether these genotypic differences or similarities among human and animal isolates translated to functionally significant attributes such as variance in host preference and/or difference in magnitude of infections, we performed a global scale analysis of M. avium subsp. paratuberculosis isolates that were representative of different genotypes and host species using DNA microarrays. Genome-wide characterization of the transcriptional changes was carried out using a human monocytic cell line (THP-1 cells) in response to different genotypes of M. avium subsp. paratuberculosis isolates recovered from various hosts. We identified several differentially expressed genes during early intracellular infection, including those involved in common canonical pathways such as NF-kappaB, interleukin-6 (IL-6), mitogen-activated protein kinase/extracellular signal-regulated kinase, and Jun N-terminal protein kinase signaling, as well as genes involved in T helper type 1 (Th1) responses (such as CCL5 ligand) and those that encode several proinflammatory cytokines and chemokine receptors. The cattle and human isolates of M. avium subsp. paratuberculosis, regardless of their short sequence repeat (SSR) genotype, induced similar global gene expression patterns in THP-1 cells. They differentially regulated genes necessary for cell survival without causing major alterations in proinflammatory genes. In contrast, the sheep isolates representing diverse SSR genotypes closely resembled the global gene expression pattern of an M

  1. Genotypic Variation in Grain P Loading across Diverse Rice Growing Environments and Implications for Field P Balances

    Science.gov (United States)

    Vandamme, Elke; Wissuwa, Matthias; Rose, Terry; Dieng, Ibnou; Drame, Khady N.; Fofana, Mamadou; Senthilkumar, Kalimuthu; Venuprasad, Ramaiah; Jallow, Demba; Segda, Zacharie; Suriyagoda, Lalith; Sirisena, Dinarathna; Kato, Yoichiro; Saito, Kazuki

    2016-01-01

    More than 60% of phosphorus (P) taken up by rice (Oryza spp.) is accumulated in the grains at harvest and hence exported from fields, leading to a continuous removal of P. If P removed from fields is not replaced by P inputs then soil P stocks decline, with consequences for subsequent crops. Breeding rice genotypes with a low concentration of P in the grains could be a strategy to reduce maintenance fertilizer needs and slow soil P depletion in low input systems. This study aimed to assess variation in grain P concentrations among rice genotypes across diverse environments and evaluate the implications for field P balances at various grain yield levels. Multi-location screening experiments were conducted at different sites across Africa and Asia and yield components and grain P concentrations were determined at harvest. Genotypic variation in grain P concentration was evaluated while considering differences in P supply and grain yield using cluster analysis to group environments and boundary line analysis to determine minimum grain P concentrations at various yield levels. Average grain P concentrations across genotypes varied almost 3-fold among environments, from 1.4 to 3.9 mg g−1. Minimum grain P concentrations associated with grain yields of 150, 300, and 500 g m−2 varied between 1.2 and 1.7, 1.3 and 1.8, and 1.7 and 2.2 mg g−1 among genotypes respectively. Two genotypes, Santhi Sufaid and DJ123, were identified as potential donors for breeding for low grain P concentration. Improvements in P balances that could be achieved by exploiting this genotypic variation are in the range of less than 0.10 g P m−2 (1 kg P ha−1) in low yielding systems, and 0.15–0.50 g P m−2 (1.5–5.0 kg P ha−1) in higher yielding systems. Improved crop management and alternative breeding approaches may be required to achieve larger reductions in grain P concentrations in rice. PMID:27729916

  2. Geographical patterns of Toxoplasma gondii genetic diversity revealed by multilocus PCR-RFLP genotyping

    Science.gov (United States)

    In recent years, an extensive collection of Toxoplasma gondii samples have been typed by the multilocus PCR-RFLP method using a standardized set of 10 genetic markers. Here we summarize the data reported until the end of 2012. A total of 1457 samples were typed into 189 genotypes. Overall, only a fe...

  3. Effects of environment and genotype on phenolic acids in wheat in the HEALTHGRAIN diversity screen.

    Science.gov (United States)

    Fernandez-Orozco, Rebeca; Li, Li; Harflett, Claudia; Shewry, Peter R; Ward, Jane L

    2010-09-08

    Phenolic acid content and composition have been determined in 26 wheat genotypes grown in Hungary over three consecutive years and at three additional locations (France, United Kingdom, and Poland) during the third year. Fractions comprising free, soluble conjugated, and bound phenolic acids were analyzed using HPLC with measurements being made for individual phenolic acids in each fraction. Statistically significant differences in phenolic acid content occurred across the different growing locations with the average total phenolic acid content being highest in the genotypes grown in Hungary. The growth year in Hungary also had a large impact, especially on the free and conjugated phenolic acid contents. Certain genotypes were more resistant to environmental impacts than others. Of the genotypes with high levels of total phenolic acids, Lynx, Riband, Tommi, and Cadenza were most stable with respect to their total contents, whereas Valoris, Herzog, and Malacca, also high in phenolic acid content, were least stable. Of the three fractions analyzed, the free and conjugated phenolic acids were most variable and were also susceptible to the effect of environment, whereas bound phenolic acids, which comprised the greatest proportion of the total phenolic acids, were the most stable.

  4. Clonality and micro-diversity of a nationwide spreading genotype of Mycobacterium tuberculosis in Japan.

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    Takayuki Wada

    Full Text Available Mycobacterium tuberculosis transmission routes can be estimated from genotypic analysis of clinical isolates from patients. In Japan, still a middle-incidence country of TB, a unique genotype strain designated as 'M-strain' has been isolated nationwide recently. To ascertain the history of the wide spread of the strain, 10 clinical isolates from different areas were subjected to genome-wide analysis based on deep sequencers. Results show that all isolates possessed common mutations to those of referential strains. The greatest number of accumulated single nucleotide variants (SNVs from the oldest coalescence was 13 nucleotides, indicating high clonality of these isolates. When an SNV common to the isolates was used as a surrogate marker of the clone, authentic clonal isolates with variation in a reliable subset of variable number of tandem repeat (VNTR genotyping method can be selected successfully from clinical isolates populations of M. tuberculosis. When the authentic clones can also be assigned to sub-clonal groups by SNVs derived from the genomic comparison, they are classifiable into three sub-clonal groups with a bias of geographical origins. Feedback from genomic analysis of clinical isolates of M. tuberculosis to genotypic markers will be an efficient strategy for the big data in various settings for public health actions against TB.

  5. Clonality and micro-diversity of a nationwide spreading genotype of Mycobacterium tuberculosis in Japan.

    Science.gov (United States)

    Wada, Takayuki; Iwamoto, Tomotada; Tamaru, Aki; Seto, Junji; Ahiko, Tadayuki; Yamamoto, Kaori; Hase, Atushi; Maeda, Shinji; Yamamoto, Taro

    2015-01-01

    Mycobacterium tuberculosis transmission routes can be estimated from genotypic analysis of clinical isolates from patients. In Japan, still a middle-incidence country of TB, a unique genotype strain designated as 'M-strain' has been isolated nationwide recently. To ascertain the history of the wide spread of the strain, 10 clinical isolates from different areas were subjected to genome-wide analysis based on deep sequencers. Results show that all isolates possessed common mutations to those of referential strains. The greatest number of accumulated single nucleotide variants (SNVs) from the oldest coalescence was 13 nucleotides, indicating high clonality of these isolates. When an SNV common to the isolates was used as a surrogate marker of the clone, authentic clonal isolates with variation in a reliable subset of variable number of tandem repeat (VNTR) genotyping method can be selected successfully from clinical isolates populations of M. tuberculosis. When the authentic clones can also be assigned to sub-clonal groups by SNVs derived from the genomic comparison, they are classifiable into three sub-clonal groups with a bias of geographical origins. Feedback from genomic analysis of clinical isolates of M. tuberculosis to genotypic markers will be an efficient strategy for the big data in various settings for public health actions against TB.

  6. Diverse Genotypes of Yersinia pestis Caused Plague in Madagascar in 2007.

    Science.gov (United States)

    Riehm, Julia M; Projahn, Michaela; Vogler, Amy J; Rajerison, Minoaerisoa; Andersen, Genevieve; Hall, Carina M; Zimmermann, Thomas; Soanandrasana, Rahelinirina; Andrianaivoarimanana, Voahangy; Straubinger, Reinhard K; Nottingham, Roxanne; Keim, Paul; Wagner, David M; Scholz, Holger C

    2015-06-01

    Yersinia pestis is the causative agent of human plague and is endemic in various African, Asian and American countries. In Madagascar, the disease represents a significant public health problem with hundreds of human cases a year. Unfortunately, poor infrastructure makes outbreak investigations challenging. DNA was extracted directly from 93 clinical samples from patients with a clinical diagnosis of plague in Madagascar in 2007. The extracted DNAs were then genotyped using three molecular genotyping methods, including, single nucleotide polymorphism (SNP) typing, multi-locus variable-number tandem repeat analysis (MLVA), and Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) analysis. These methods provided increasing resolution, respectively. The results of these analyses revealed that, in 2007, ten molecular groups, two newly described here and eight previously identified, were responsible for causing human plague in geographically distinct areas of Madagascar. Plague in Madagascar is caused by numerous distinct types of Y. pestis. Genotyping method choice should be based upon the discriminatory power needed, expense, and available data for any desired comparisons. We conclude that genotyping should be a standard tool used in epidemiological investigations of plague outbreaks.

  7. Phenotypic diversity of basic characteristics of genotypes from the Serbia onion collection

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    Gvozdanović-Varga Jelica

    2013-01-01

    Full Text Available The onion is a widely distributed vegetable crop, which takes an important place in the vegetable production in Serbia. The traditional planting method is the one from sets. Old cultivars and populations and, in recent years, foreign cultivars are grown. The large variety of genotypes, including both domestic populations and cultivars, comprises the significant gene pool of this region. The onion collection of the Institute of Field and Vegetable Crops in Novi Sad is based on the populations and cultivars from the territory of the former Yugoslavia. This paper reviews 30 onion genotypes on the basic IPGRI descriptors (ANNEX I. Variability of the reviewed characteristics was determined by PC analysis. High variability values have been established for bulb skin color, bulb flesh color, bulb hearting and bulb skin thickness. The genotypes varied in bulb skin color as well as in bulb flesh color from white to dark violet. These two characteristics had the largest impact on clustering, with a single genotype being heterogeneous exactly for these two characteristics. [Projekat Ministarstva nauke Republike Srbije, br. TR 31030

  8. Exploiting genotypic diversity of 2,4-Diacetylphloroglucinol-producing Pseudomonas spp.: characterization of superior root-colonizing P. fluorescens strain Q8r1-96

    NARCIS (Netherlands)

    Raaijmakers, J.M.; Weller, D.M.

    2001-01-01

    The genotypic diversity that occurs in natural populations of antagonistic microorganisms provides an enormous resource for improving biological control of plant diseases. In this study, we determined the diversity of indigenous 2,4-diacetylphloroglucinol (DAPG)-producing Pseudomonas spp. occurring

  9. Molecular characterization and genetic diversity in an avocado collection of cultivars and local Spanish genotypes using SSRs.

    Science.gov (United States)

    Alcaraz, M L; Hormaza, J I

    2007-12-01

    In this work, 75 avocado accessions maintained in an ex situ germplasm collection at the E.E. la Mayora in Málaga (Spain) were characterized with 16 microsatellites previously developed in this species. This avocado collection includes both local Spanish genotypes obtained through prospection and genotypes obtained by exchange with different countries. A total of 156 different amplification fragments were detected ranging from 4 to 16 per locus with an average of 9.75 alleles per locus. All the microsatellites were highly informative with an expected heterozygosity higher than 0.5 and a probability of identity below 0.36. The total probability of identity was 2.85x10(-14). Fifteen of the 16 loci studied showed a positive Wright's fixation index (F) indicating a deficit of heterozygotes with an average over all the SSRs of 0.18. A dendrogram was generated using UPGMA (unweighted pair group method with arithmetic averages) based on the Nei and Li similarity index. This dendrogram classified most of the genotypes analyzed into three major groups which mainly differed in racial origin although with low bootstrap support probably due to the presence of many interracial hybrids in the collection. All the genotypes studied could be unequivocally distinguished with the combination of SSRs used except some putative mutations of 'Hass' and an additional group of two cultivars. The results obtained indicate that the set of SSRs used is highly informative and are discussed in terms of their implications for avocado germplasm characterization and management.

  10. Subgenotype diversity of hepatitis B virus American genotype F in Amerindians from Venezuela and the general population of Colombia.

    Science.gov (United States)

    Devesa, M; Loureiro, C L; Rivas, Y; Monsalve, F; Cardona, N; Duarte, M C; Poblete, F; Gutierrez, M F; Botto, C; Pujol, F H

    2008-01-01

    The objective of this study was the evaluation of the genetic diversity found in HBV circulating among Venezuelan Amerindians and the general population in Colombia. Phylogenetic analysis of the S region in 194 isolates showed that genotype F is highly predominant in Colombia and Venezuela. This might be related to the genetic background of the population. F3 is the main subgenotype which circulates in both countries. Phylogenetic analysis of 61 complete genome sequences of HBV American genotypes confirms the presence of two genotypes F and H, and 4 F subgenotypes. In Venezuela, subgenotypes F1, F2, and F3 circulate in East and West Amerindians, while only F3 was found among South Amerindians. Japreira community derived from Yucpa Amerindians around 150 years ago. However, several Japreira HBV sequences were forming a clade that can be classified as subgenotype 2b, differing from Yucpa sequences that belong mainly to subgenotype F3. The apparent absence of correlation between the phylogenetic groupings of HBV isolates with the ethnical origin in aboriginal populations might be suggesting a recent origin of HBV American subgenotypes, or a genetic drift effect.

  11. Potential pathogens, antimicrobial patterns and genotypic diversity of Escherichia coli isolates in constructed wetlands treating swine wastewater.

    Science.gov (United States)

    Ibekwe, A M; Murinda, Shelton E; DebRoy, Chitrita; Reddy, Gudigopura B

    2016-02-01

    Escherichia coli populations originating from swine houses through constructed wetlands were analyzed for potential pathogens, antimicrobial susceptibility patterns, and genotypic diversity. Escherichia coli isolates (n = 493) were screened for the presence of the following virulence genes: stx1, stx2 and eae (Shiga toxin-producing E. coli [STEC]), heat-labile enterotoxin (LT) genes and heat stable toxin STa and STb (enterotoxigenic E. coli (ETEC), cytotoxin necrotizing factors 1 and 2 (cnf1 and cnf2 [necrotoxigenic E. coli- NTEC]), as well as O and H antigens, and the presence of the antibiotic resistance genes blaTEM, blaSHV, blaCMY-2, tet A, tet B, tet C, mph(A), aadA, StrA/B, sul1, sul2 and sul3. The commensal strains were further screened for 16 antimicrobials and characterized by BOX AIR-1 PCR for unique genotypes. The highest antibiotic resistance prevalence was for tetracycline, followed by erythromycin, ampicillin, streptomycin, sulfisoxazole and kanamycin. Our data showed that most of the isolates had high distribution of single or multidrug-resistant (MDR) genotypes. Therefore, the occurrence of MDR E. coli in the wetland is a matter of great concern due to possible transfer of resistance genes from nonpathogenic to pathogenic strains or vice versa in the environment.

  12. Antimicrobial Resistance and Genotypic Diversity of Campylobacter Isolated from Pig, Dairy and Beef Cattle in Tanzania

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    Isaac eKashoma

    2015-11-01

    Full Text Available Foodborne Campylobacter infections pose a serious threat to public health worldwide. However, the occurrence and characteristics of Campylobacter in food animals and products remain largely unknown in Tanzania. The objective of this study was to determine the prevalence, antibiotic resistance, and genetic profiles (sequence types, STs of Campylobacter isolated from feces of pigs and dairy and beef cattle in Tanzania. Overall, 259 (~ 30% of 864 samples were positive for Campylobacter spp, which were detected in 32.5%, 35.4%, and 19.6% of the pig, dairy, and beef cattle samples, respectively. Multiplex PCR analysis identified 64.5% and 29.3% of the Campylobacter isolates as C. coli and C. jejuni, respectively. The majority (91.9% of the isolates from pig samples were identified as C. coli, while C. jejuni accounted for 65.5% of the isolates from cattle. Antimicrobial susceptibility testing using the disk diffusion assay and the broth microdilution method revealed resistance to: ampicillin (70% and 76%, gentamicin (1.8% and 12.6%, respectively, streptomycin (65.8% and 74.8%, erythromycin (41.4% and 48.7%, tetracycline (18.9% and 23.4%, and ciprofloxacin (14.4% and 7.2%. Resistance to nalidixic acid (39.6%, azithromycin (13.5%, and chloramphenicol (4.5% was determined using the disk diffusion assay only, while resistance to tylosin (38.7% was quantified using the broth microdilution method. Multilocus sequence typing of 111 Campylobacter isolates resulted in the identification of 48 STs (26 C. jejuni and 22 C. coli of which 7 were novel (6 C. jejuni and 1 C. coli. Taken together, this study revealed the high prevalence, genetic diversity and antimicrobial resistance of Campylobacter in important food animals in Tanzania, which highlights the urgent need for the surveillance and control of Campylobacter in this country.

  13. Genotypic Diversity of Staphylococcus aureus α-Hemolysin Gene (hla and Its Association with Clonal Background: Implications for Vaccine Development.

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    Meng Xiao

    Full Text Available The α-hemolysin, encoded by the hla gene, is a major virulence factor in S. aureus infections. Changes in key amino acid residues of α-hemolysin can result in reduction, or even loss, of toxicity. The aim of this study was to investigate the diversity of the hla gene sequence and the relationship of hla variants to the clonal background of S. aureus isolates. A total of 47 clinical isolates from China were used in this study, supplemented with in silico analysis of 318 well-characterized whole genome sequences from globally distributed isolates. A total of 28 hla genotypes were found, including three unique to isolates from China, 20 found only in the global genomes and five found in both. The hla genotype generally correlated with the clonal background, particularly the multilocus sequence type, but was not related to geographic origin, host source or methicillin-resistance phenotype. In addition, the hla gene showed greater diversity than the seven loci utilized in the MLST scheme for S. aureus. Our investigation has provided genetic data which may be useful for future studies of toxicity, immunogenicity and vaccine development.

  14. Phenotypic and genotypic diversity of Pseudomonas aeruginosa strains isolated from hospitals in siedlce (Poland

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    Katarzyna Wolska

    2012-03-01

    Full Text Available A total of 62 Pseudomonas aeruginosa strains isolated from two hospitals in Siedlce (Poland were studied by repetitive element based PCR (rep-PCR using BOX primer. BOX-PCR results revealed the presence of 7 numerous genotypes and 31 unique patterns among isolates. Generally, the strains of P. aeruginosa were characterized by resistance to many antibiotics tested and by differences in serogroups and types of growth on cetrimide agar medium. However, the P. aeruginosa strains isolated from faeces showed much lower phenotypic and genotypic variations in comparison with strains obtained from other clinical specimens. It was observed that genetic techniques supported by phenotypic tests have enabled to conduct a detailed characterization of P. aeruginosa strains isolated from a particular environment at a particular time.

  15. Detection and genotyping of Chlamydia species responsible for reproductive disorders in Algerian small ruminants.

    Science.gov (United States)

    Merdja, Salah-Eddine; Khaled, Hamza; Aaziz, Rachid; Vorimore, Fabien; Bertin, Claire; Dahmani, Ali; Bouyoucef, Abdallah; Laroucau, Karine

    2015-02-01

    Chlamydiosis in small ruminants is a zoonotic disease mainly related to Chlamydia abortus. This bacterium is responsible for abortions and reproductive disorders in sheep and goats. Stillbirth and infertility, leading to important economic losses, are also associated with this pathology. In Algeria, abortion cases are frequently reported by veterinarians but, except for brucellosis which is a notifiable disease in this country, abortive diseases are in general poorly studied. In order to detect and genotype Chlamydia species in small ruminants in different areas of Algeria, a study was conducted on samples collected from females (164 blood samples and 199 vaginal swabs) between October 2011 and March 2013. Serum samples were tested with a C. abortus-specific indirect ELISA test. Fourteen samples (8.5 %), from six farms (6/20, 30 %) were tested positive. Vaginal swabs were analysed with a real-time PCR targeting all Chlamydiaceae spp. Thirty samples (15 %) were diagnosed positive in 16 farms (16/25, 64 %). Positive samples were all re-tested with a C. abortus- and a C. pecorum-specific real-time PCR. Finally, 13/30 (43.3 %) and 6/30 (20 %) were identified as C. abortus and C. pecorum, respectively. Enough concentrated C. abortus samples were genotyped by multi-loci variable number of tandem repeat (VNTR) analysis (MLVA), and all were related to the genotype [2] group which mainly includes French C. abortus isolates. C. pecorum-positive samples were genotyped by multi-locus sequence typing (MLST). Interestingly, two of them were successfully genotyped and showed identical MLST sequences to VB2, AB10, E58 and SBE, a group which includes C. pecorum isolates considered as highly pathogenic. These findings suggest a possible role of C. abortus and C. pecorum strains in the aetiology of abortion in Algerian small ruminants.

  16. Inference of antibiotic resistance and virulence among diverse group A Streptococcus strains using emm sequencing and multilocus genotyping methods.

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    David Metzgar

    Full Text Available BACKGROUND: Group A Streptococcus pyogenes (GAS exhibits a high degree of clinically relevant phenotypic diversity. Strains vary widely in terms of antibiotic resistance (AbR, clinical severity, and transmission rate. Currently, strain identification is achieved by emm typing (direct sequencing of the genomic segment coding for the antigenic portion of the M protein or by multilocus genotyping methods. Phenotype analysis, including critical AbR typing, is generally achieved by much slower and more laborious direct culture-based methods. METHODOLOGY/PRINCIPAL FINDINGS: We compare genotype identification (by emm typing and PCR/ESI-MS with directly measured phenotypes (AbR and outbreak associations for 802 clinical isolates of GAS collected from symptomatic patients over a period of 6 years at 10 military facilities in the United States. All independent strain characterization methods are highly correlated. This shows that recombination, horizontal transfer, and other forms of reassortment are rare in GAS insofar as housekeeping genes, primary virulence and antibiotic resistance determinants, and the emm gene are concerned. Therefore, genotyping methods offer an efficient way to predict emm type and the associated AbR and virulence phenotypes. CONCLUSIONS/SIGNIFICANCE: The data presented here, combined with much historical data, suggest that emm typing assays and faster molecular methods that infer emm type from genomic signatures could be used to efficiently infer critical phenotypic characteristics based on robust genotype: phenotype correlations. This, in turn, would enable faster and better-targeted responses during identified outbreaks of constitutively resistant or particularly virulent emm types.

  17. Genetic Diversity Assessment and Identification of New Sour Cherry Genotypes Using Intersimple Sequence Repeat Markers

    OpenAIRE

    Roghayeh Najafzadeh; Kazem Arzani; Naser Bouzari; Ali Saei

    2014-01-01

    Iran is one of the chief origins of subgenus Cerasus germplasm. In this study, the genetic variation of new Iranian sour cherries (which had such superior growth characteristics and fruit quality as to be considered for the introduction of new cultivars) was investigated and identified using 23 intersimple sequence repeat (ISSR) markers. Results indicated a high level of polymorphism of the genotypes based on these markers. According to these results, primers tested in this study specially IS...

  18. Diversity of cacao trees in Waslala, Nicaragua: associations between genotype spectra, product quality and yield potential.

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    Bodo Trognitz

    Full Text Available The sensory quality and the contents of quality-determining chemical compounds in unfermented and fermented cocoa from 100 cacao trees (individual genotypes representing groups of nine genotype spectra (GG, grown at smallholder plantings in the municipality of Waslala, Nicaragua, were evaluated for two successive harvest periods. Cocoa samples were fermented using a technique mimicking recommended on-farm practices. The sensory cocoa quality was assessed by experienced tasters, and seven major chemical taste compounds were quantified by near infrared spectrometry (NIRS. The association of the nine, partially admixed, genotype spectra with the analytical and sensory quality parameters was tested. The individual parameters were analyzed as a function of the factors GG and harvest (including the date of fermentation, individual trees within a single GG were used as replications. In fermented cocoa, significant GG-specific differences were observed for methylxanthines, theobromine-to-caffeine (T/C ratio, total fat, procyanidin B5 and epicatechin, as well as the sensory attributes global score, astringency, and dry fruit aroma, but differences related to harvest were also apparent. The potential cocoa yield was also highly determined by the individual GG, although there was significant tree-to-tree variation within every single GG. Non-fermented samples showed large harvest-to-harvest variation of their chemical composition, while differences between GG were insignificant. These results suggest that selection by the genetic background, represented here by groups of partially admixed genotype spectra, would be a useful strategy toward enhancing quality and yield of cocoa in Nicaragua. Selection by the GG within the local, genetically segregating populations of seed-propagated cacao, followed by clonal propagation of best-performing individuals of the selected GG could be a viable alternative to traditional propagation of cacao by seed from open

  19. Diversity of cacao trees in Waslala, Nicaragua: associations between genotype spectra, product quality and yield potential.

    Science.gov (United States)

    Trognitz, Bodo; Cros, Emile; Assemat, Sophie; Davrieux, Fabrice; Forestier-Chiron, Nelly; Ayestas, Eusebio; Kuant, Aldo; Scheldeman, Xavier; Hermann, Michael

    2013-01-01

    The sensory quality and the contents of quality-determining chemical compounds in unfermented and fermented cocoa from 100 cacao trees (individual genotypes) representing groups of nine genotype spectra (GG), grown at smallholder plantings in the municipality of Waslala, Nicaragua, were evaluated for two successive harvest periods. Cocoa samples were fermented using a technique mimicking recommended on-farm practices. The sensory cocoa quality was assessed by experienced tasters, and seven major chemical taste compounds were quantified by near infrared spectrometry (NIRS). The association of the nine, partially admixed, genotype spectra with the analytical and sensory quality parameters was tested. The individual parameters were analyzed as a function of the factors GG and harvest (including the date of fermentation), individual trees within a single GG were used as replications. In fermented cocoa, significant GG-specific differences were observed for methylxanthines, theobromine-to-caffeine (T/C) ratio, total fat, procyanidin B5 and epicatechin, as well as the sensory attributes global score, astringency, and dry fruit aroma, but differences related to harvest were also apparent. The potential cocoa yield was also highly determined by the individual GG, although there was significant tree-to-tree variation within every single GG. Non-fermented samples showed large harvest-to-harvest variation of their chemical composition, while differences between GG were insignificant. These results suggest that selection by the genetic background, represented here by groups of partially admixed genotype spectra, would be a useful strategy toward enhancing quality and yield of cocoa in Nicaragua. Selection by the GG within the local, genetically segregating populations of seed-propagated cacao, followed by clonal propagation of best-performing individuals of the selected GG could be a viable alternative to traditional propagation of cacao by seed from open pollination. Fast and

  20. Development of a rapid HRM genotyping method for detection of dog-derived Giardia lamblia.

    Science.gov (United States)

    Tan, Liping; Yu, Xingang; Abdullahi, Auwalu Yusuf; Wu, Sheng; Zheng, Guochao; Hu, Wei; Song, Meiran; Wang, Zhen; Jiang, Biao; Li, Guoqing

    2015-11-01

    Giardia lamblia is a zoonotic flagellate protozoan in the intestine of human and many mammals including dogs. To assess a threat of dog-derived G. lamblia to humans, the common dog-derived G. lamblia assemblages A, C, and D were genotyped by high-resolution melting (HRM) technology. According to β-giardin gene sequence, the qPCR-HRM primers BG5 and BG7 were designed. A series of experiments on the stability, sensitivity, and accuracy of the HRM method were also tested. Results showed that the primers BG5 and BG7 could distinguish among three assemblages A, C, and D, which Tm value differences were about 1 °C to each other. The melting curves of intra-assay reproducibility were almost coincided, and those of inter-assay reproducibility were much the same shape. The lowest detection concentration was about 5 × 10(-6)-ng/μL sample. The genotyping results from 21 G. lamblia samples by the HRM method were in complete accordance with sequencing results. It is concluded that the HRM genotyping method is rapid, stable, specific, highly sensitive, and suitable for clinical detection and molecular epidemiological survey of dog-derived G. lamblia.

  1. A comparison of growth and development patterns in diverse genotypes of broilers. 1. Male broiler growth.

    Science.gov (United States)

    Reddish, J M; Lilburn, M S

    2004-07-01

    Selection for breast muscle yield and BW in commercial broilers has resulted in genotypes far different from broilers processed in the past. When comparative studies with commercial lines are conducted, it is often difficult to differentiate between carcass effects resulting from direct genetic selection vs. correlated effects that partially reflect genetic changes in BW. The objective of the present experiment was to compare growth and development characteristics of male broilers from commercial lines exhibiting similar rates of BW gain based on a percentage of 8-wk BW but exhibiting different carcass traits. Male broilers from 2 commercial genotypes exhibiting increased breast muscle yield (A and B) were compared with broilers from a commercial line that was "unimproved" with respect to conformation (C). All birds were fed a commercial-type broiler starter diet throughout the study and were processed at 8 wk of age for carcass comparisons. No differences were observed among lines for breast-free BW or weight of the abdominal fat pad. The absolute and relative weights of pectoralis major and pectoralis minor breast muscles were heavier in lines A and B than C (P A > B; P C >A; P C; P A = B, P A = B, P development between commercial genotypes with similar growth patterns.

  2. Detecting pin diversion from pressurized water reactors spent fuel assemblies

    Energy Technology Data Exchange (ETDEWEB)

    Ham, Young S.; Sitaraman, Shivakumar

    2017-01-10

    Detecting diversion of spent fuel from Pressurized Water Reactors (PWR) by determining possible diversion including the steps of providing a detector cluster containing gamma ray and neutron detectors, inserting the detector cluster containing the gamma ray and neutron detectors into the spent fuel assembly through the guide tube holes in the spent fuel assembly, measuring gamma ray and neutron radiation responses of the gamma ray and neutron detectors in the guide tube holes, processing the gamma ray and neutron radiation responses at the guide tube locations by normalizing them to the maximum value among each set of responses and taking the ratio of the gamma ray and neutron responses at the guide tube locations and normalizing the ratios to the maximum value among them and producing three signatures, gamma, neutron, and gamma-neutron ratio, based on these normalized values, and producing an output that consists of these signatures that can indicate possible diversion of the pins from the spent fuel assembly.

  3. [Molecular test Genotype® MTBDRplus, an alternative to rapid detection of multidrug resistance tuberculosis].

    Science.gov (United States)

    Asencios, Luis; Galarza, Marco; Quispe, Neyda; Vásquez, Lucy; Leo, Elena; Valencia, Eddy; Ramírez, Juan; Acurio, Margoth; Salazar, Rosario; Mendoza-Ticona, Alberto; Cáceres, Omar

    2012-03-01

    The Genotype®MTBDRplus molecular test is a method that allows identification of the most frequent mutations associated with resistance to major first-line antituberculosis drugs, Isoniazid (INH) and Rifampicin (RFP). The aim of this study was to evaluate the performance of the molecular test with culture and smear- positive sputum samples. We evaluated 95 cultures and 100 sputum samples with resistance profiles previously determined by the reference method "Agar Plate Proportions" (APP). The molecular test from cultures showed a sensitivity of 100 %, 97,5 % and 96,97 % for RIF, INH and MDR respectively while from sputums the sensitivity was 95,65 %, 96,77 % and 95,24 % for RIF, INH and MDR respectively. We conclude that the molecular test Genotype®MTBDRplus is a very useful tool to detect resistance to isoniazid and rifampicin simultaneously (MDR-TB) in up to 72 hours from sputum samples or cultures.

  4. Genotypic and symbiotic diversity of Rhizobium populations associated with cultivated lentil and pea in sub-humid and semi-arid regions of Eastern Algeria.

    Science.gov (United States)

    Riah, Nassira; Béna, Gilles; Djekoun, Abdelhamid; Heulin, Karine; de Lajudie, Philippe; Laguerre, Gisèle

    2014-07-01

    The genetic structure of rhizobia nodulating pea and lentil in Algeria, Northern Africa was determined. A total of 237 isolates were obtained from root nodules collected on lentil (Lens culinaris), proteaginous and forage pea (Pisum sativum) growing in two eco-climatic zones, sub-humid and semi-arid, in Eastern Algeria. They were characterised by PCR-restriction fragment length polymorphism (RFLP) of the 16S-23S rRNA intergenic region (IGS), and the nodD-F symbiotic region. The combination of these haplotypes allowed the isolates to be clustered into 26 distinct genotypes, and all isolates were classified as Rhizobium leguminosarum. Symbiotic marker variation (nodD-F) was low but with the predominance of one nod haplotype (g), which had been recovered previously at a high frequency in Europe. Sequence analysis of the IGS further confirmed its high variability in the studied strains. An AMOVA analysis showed highly significant differentiation in the IGS haplotype distribution between populations from both eco-climatic zones. This differentiation was reflected by differences in dominant genotype frequencies. Conversely, no host plant effect was detected. The nodD gene sequence-based phylogeny suggested that symbiotic gene diversity in pea and lentil nodulating rhizobial populations in Algeria was low compared to that reported elsewhere in the world.

  5. A single-nucleotide-polymorphism-based genotyping assay for simultaneous detection of different carbendazim-resistant genotypes in the Fusarium graminearum species complex

    Science.gov (United States)

    Zhang, Hao; Brankovics, Balázs; van der Lee, Theo A.J.; Waalwijk, Cees; van Diepeningen, Anne A.D.; Xu, Jin; Xu, Jingsheng

    2016-01-01

    The occurrence resistance to methyl benzimidazole carbamates (MBC)-fungicides in the Fusarium graminearum species complex (FGSC) is becoming a serious problem in the control of Fusarium head blight in China. The resistance is caused by point mutations in the β2-tubulingene. So far, five resistant genotypes (F167Y, E198Q, E198L, E198K and F200Y) have been reported in the field. To establish a high-throughput method for rapid detection of all the five mutations simultaneously, an efficient single-nucleotide-polymorphism-based genotyping method was developed based on the Luminex xMAP system. One pair of amplification primers and five allele specific primer extension probes were designed and optimized to specially distinguish the different genotypes within one single reaction. This method has good extensibility and can be combined with previous reported probes to form a highly integrated tool for species, trichothecene chemotype and MBC resistance detection. Using this method, carbendazim resistant FGSC isolates from Jiangsu, Anhui and Sichuan Province in China were identified. High and moderate frequencies of resistance were observed in Jiangsu and Anhui Province, respectively. Carbendazim resistance in F. asiaticum is only observed in the 3ADON genotype. Overall, our method proved to be useful for early detection of MBC resistance in the field and the result aids in the choice of fungicide type. PMID:27812414

  6. Genetic Drift, Purifying Selection and Vector Genotype Shape Dengue Virus Intra-host Genetic Diversity in Mosquitoes

    Science.gov (United States)

    Fontaine, Albin; Ar Gouilh, Meriadeg; Moltini-Conclois, Isabelle

    2016-01-01

    Due to their error-prone replication, RNA viruses typically exist as a diverse population of closely related genomes, which is considered critical for their fitness and adaptive potential. Intra-host demographic fluctuations that stochastically reduce the effective size of viral populations are a challenge to maintaining genetic diversity during systemic host infection. Arthropod-borne viruses (arboviruses) traverse several anatomical barriers during infection of their arthropod vectors that are believed to impose population bottlenecks. These anatomical barriers have been associated with both maintenance of arboviral genetic diversity and alteration of the variant repertoire. Whether these patterns result from stochastic sampling (genetic drift) rather than natural selection, and/or from the influence of vector genetic heterogeneity has not been elucidated. Here, we used deep sequencing of full-length viral genomes to monitor the intra-host evolution of a wild-type dengue virus isolate during infection of several mosquito genetic backgrounds. We estimated a bottleneck size ranging from 5 to 42 founding viral genomes at initial midgut infection, irrespective of mosquito genotype, resulting in stochastic reshuffling of the variant repertoire. The observed level of genetic diversity increased following initial midgut infection but significantly differed between mosquito genetic backgrounds despite a similar initial bottleneck size. Natural selection was predominantly negative (purifying) during viral population expansion. Taken together, our results indicate that dengue virus intra-host genetic diversity in the mosquito vector is shaped by genetic drift and purifying selection, and point to a novel role for vector genetic factors in the genetic breadth of virus populations during infection. Identifying the evolutionary forces acting on arboviral populations within their arthropod vector provides novel insights into arbovirus evolution. PMID:27304978

  7. Genetic Drift, Purifying Selection and Vector Genotype Shape Dengue Virus Intra-host Genetic Diversity in Mosquitoes.

    Science.gov (United States)

    Lequime, Sebastian; Fontaine, Albin; Ar Gouilh, Meriadeg; Moltini-Conclois, Isabelle; Lambrechts, Louis

    2016-06-01

    Due to their error-prone replication, RNA viruses typically exist as a diverse population of closely related genomes, which is considered critical for their fitness and adaptive potential. Intra-host demographic fluctuations that stochastically reduce the effective size of viral populations are a challenge to maintaining genetic diversity during systemic host infection. Arthropod-borne viruses (arboviruses) traverse several anatomical barriers during infection of their arthropod vectors that are believed to impose population bottlenecks. These anatomical barriers have been associated with both maintenance of arboviral genetic diversity and alteration of the variant repertoire. Whether these patterns result from stochastic sampling (genetic drift) rather than natural selection, and/or from the influence of vector genetic heterogeneity has not been elucidated. Here, we used deep sequencing of full-length viral genomes to monitor the intra-host evolution of a wild-type dengue virus isolate during infection of several mosquito genetic backgrounds. We estimated a bottleneck size ranging from 5 to 42 founding viral genomes at initial midgut infection, irrespective of mosquito genotype, resulting in stochastic reshuffling of the variant repertoire. The observed level of genetic diversity increased following initial midgut infection but significantly differed between mosquito genetic backgrounds despite a similar initial bottleneck size. Natural selection was predominantly negative (purifying) during viral population expansion. Taken together, our results indicate that dengue virus intra-host genetic diversity in the mosquito vector is shaped by genetic drift and purifying selection, and point to a novel role for vector genetic factors in the genetic breadth of virus populations during infection. Identifying the evolutionary forces acting on arboviral populations within their arthropod vector provides novel insights into arbovirus evolution.

  8. [HPV genotypes prevalence in México and worldwide detected by Linear Array].

    Science.gov (United States)

    Flores-Miramontes, María G; Torres-Reyes, Luis A; Aguilar-Lemarroy, Adriana; Vallejo-Ruíz, Verónica; Piña-Sánchez, Patricia; Cortés-Gutiérrez, Elva; Reyes-Leyva, Julio; Jave-Suárez, Luis Felipe

    2015-01-01

    Infection with human papillomavirus (HPV) is the main factor associated with the development of cervical cancer (CC). Knowing about the prevalence of HPVs at different stages in the development of CC is important for determining the HPV oncogenic risk, the development of screening strategies, the evaluation of prevention programs, and also for vaccine designing. This paper is a meta-analysis of HPV prevalence worldwide and in Mexico from studies using the Linear Array® HPV Genotyping Test as a diagnostic test (it is the commercial test that, up to date, identifies the largest number of HPV genotypes in a single sample) in DNA of cervical samples from women with normal cytology, with low grade squamous intraepithelial lesions (LGSIL), with high grade squamous intraepithelial lesions (HGSIL) and with CC. The most prevalent genotypes after HPV-16 and -18 in women with CC varies depending on geographic region, which supports the need to develop detection and prevention strategies according to the characteristics of the population.

  9. Effects of salinity stress on carotenoids, anthocyanins, and color of diverse tomato genotypes.

    Science.gov (United States)

    Borghesi, Eva; González-Miret, M Lourdes; Escudero-Gilete, M Luisa; Malorgio, Fernando; Heredia, Francisco J; Meléndez-Martínez, Antonio J

    2011-11-09

    One nonanthocyanin-accumulating (Ailsa Craig) and three anthocyanin-accumulating tomato genotypes (Anthocyanin fruit type, Atroviolaceum, and Sun Black) were analyzed to assess differences in their carotenoid and anthocyanin levels and color and to evaluate the effects of nutrient solutions with different salt concentrations on these parameters. The carotenoid content of control Atroviolaceum tomatoes was ca. 2-2.5-fold higher relative to the other two types, and the color of its puree could be visually distinguished from those of other genotypes. Salinity stress led in some cases to a 2-3-fold increase in the lycopene content. Saline treatment increased the accumulation of total anthocyanins in fruits of Sun Black (2-fold increase), while it reduced it in fruits of Anthocyanin (10-fold decrease). In general, the treatment increased the differences in color of different purees. These results indicate that salinity stress can lead to similar or higher increases in tomato carotenoids than those achieved by genetic engineering. In addition, these changes were accompanied by visually discernible color differences in tomato products. Our findings show the considerable potential of exploiting saline soils to obtain tomatoes with higher levels of secondary metabolites like carotenoids and anthocyanins.

  10. Genotypically Different Clones of Staphylococcus aureus Are Diverse in the Antimicrobial Susceptibility Patterns and Biofilm Formations

    Directory of Open Access Journals (Sweden)

    Salman Sahab Atshan

    2013-01-01

    Full Text Available This study evaluated whether genotypically different clinical isolates of S. aureus have similar susceptibilities to individual antibiotics. It further aims to check the impact of biofilm on the in vitro activity of vancomycin, daptomycin, linezolid, and tigecycline against S. aureus clones. The study used a total of 60 different clinical MSSA and MRSA isolates. Susceptibilities were performed in planktonic cultures by macrobroth dilution and epsilon-test (E test system. Biofilm production was determined using an adherent plate assay. The efficacy of antimicrobial activities against biofilms formation was checked using confocal laser scanning microscopy (CLSM. The study found that similar and different spa, MLST, and SCCmec types displayed high variation in their susceptibilities to antibiotics with tigecycline and daptomycin being the most effective. The biofilms were found resistant to high concentrations of most antibiotics tested with daptomycin being the most effective drug used in adhesive biofilms. A considerable difference exists among similar and various clone types against antibiotics tested. This variation could have contributed to the degree of virulence even within the same clonal genotype and enhanced heterogeneity in the infection potential. Thus, the development of a rapid and precise identification profile for each clone in human infections is important.

  11. Genotypic Diversity of Coagulase-Negative Staphylococci Causing Endocarditis: a Global Perspective▿

    Science.gov (United States)

    Petti, Cathy A.; Simmon, Keith E.; Miro, Jose M.; Hoen, Bruno; Marco, Francesc; Chu, Vivian H.; Athan, Eugene; Bukovski, Suzana; Bouza, Emilio; Bradley, Suzanne; Fowler, Vance G.; Giannitsioti, Efthymia; Gordon, David; Reinbott, Porl; Korman, Tony; Lang, Selwyn; Garcia-de-la-Maria, Cristina; Raglio, Annibale; Morris, Arthur J.; Plesiat, Patrick; Ryan, Suzanne; Doco-Lecompte, Thanh; Tripodi, Francesca; Utili, Riccardo; Wray, Dannah; Federspiel, J. Jeffrey; Boisson, K.; Reller, L. Barth; Murdoch, David R.; Woods, Christopher W.

    2008-01-01

    Coagulase-negative staphylococci (CNS) are important causes of infective endocarditis (IE), but their microbiological profiles are poorly described. We performed DNA target sequencing and susceptibility testing for 91 patients with definite CNS IE who were identified from the International Collaboration on Endocarditis—Microbiology, a large, multicenter, multinational consortium. A hierarchy of gene sequences demonstrated great genetic diversity within CNS from patients with definite endocarditis that represented diverse geographic regions. In particular, rpoB sequence data demonstrated unique genetic signatures with the potential to serve as an important tool for global surveillance. PMID:18367572

  12. Genotypic diversity of coagulase-negative staphylococci causing endocarditis: a global perspective.

    Science.gov (United States)

    Petti, Cathy A; Simmon, Keith E; Miro, Jose M; Hoen, Bruno; Marco, Francesc; Chu, Vivian H; Athan, Eugene; Bukovski, Suzana; Bouza, Emilio; Bradley, Suzanne; Fowler, Vance G; Giannitsioti, Efthymia; Gordon, David; Reinbott, Porl; Korman, Tony; Lang, Selwyn; Garcia-de-la-Maria, Cristina; Raglio, Annibale; Morris, Arthur J; Plesiat, Patrick; Ryan, Suzanne; Doco-Lecompte, Thanh; Tripodi, Francesca; Utili, Riccardo; Wray, Dannah; Federspiel, J Jeffrey; Boisson, K; Reller, L Barth; Murdoch, David R; Woods, Christopher W

    2008-05-01

    Coagulase-negative staphylococci (CNS) are important causes of infective endocarditis (IE), but their microbiological profiles are poorly described. We performed DNA target sequencing and susceptibility testing for 91 patients with definite CNS IE who were identified from the International Collaboration on Endocarditis-Microbiology, a large, multicenter, multinational consortium. A hierarchy of gene sequences demonstrated great genetic diversity within CNS from patients with definite endocarditis that represented diverse geographic regions. In particular, rpoB sequence data demonstrated unique genetic signatures with the potential to serve as an important tool for global surveillance.

  13. Genotypic detection of rifampicin and isoniazid resistant Mycobacterium tuberculosis strains by DNA sequencing: a randomized trial

    Directory of Open Access Journals (Sweden)

    El mashad Noha

    2009-01-01

    Full Text Available Abstract Background Tuberculosis is a growing international health concern. It is the biggest killer among the infectious diseases in the world today. Early detection of drug resistance allows starting of an appropriate treatment. Resistance to drugs is due to particular genomic mutations in specific genes of Mycobacterium tuberculosis(MTB. The aim of this study was to identify the presence of Isoniazid (INH and Rifampicin(RIF drug resistance in new and previously treated tuberculosis (TB cases using DNA sequencing. Methods This study was carried out on 153 tuberculous patients with positive Bactec 460 culture for acid fast bacilli. Results Of the 153 patients, 105 (68.6% were new cases and 48 (31.4% were previously treated cases. Drug susceptibility testing on Bactec revealed 50 resistant cases for one or more of the first line antituberculous. Genotypic analysis was done only for rifampicin resistant specimens (23 cases and INH resistant specimens (26 cases to detect mutations responsible for drug resistance by PCR amplification of rpoB gene for rifampicin resistant cases and KatG gene for isoniazid resistant cases. Finally, DNA sequencing was done for detection of mutation within rpoB and KatG genes. Genotypic analysis of RIF resistant cases revealed that 20/23 cases (86.9% of RIF resistance were having rpoB gene mutation versus 3 cases (13.1% having no mutation with a high statistical significant difference between them (P Conclusion We can conclude that rifampicin resistance could be used as a useful surrogate marker for estimation of multidrug resistance. In addition, Genotypic method was superior to that of the traditional phenotypic method which is time-consuming taking several weeks or longer.

  14. Satellite detection of wastewater diversion plumes in Southern California

    Science.gov (United States)

    Gierach, Michelle M.; Holt, Benjamin; Trinh, Rebecca; Jack Pan, B.; Rains, Christine

    2017-02-01

    Multi-sensor satellite observations proved useful in detecting surfacing wastewater plumes during the 2006 Hyperion Treatment Plant (HTP) and 2012 Orange County Sanitation District (OCSD) wastewater diversion events in Southern California. Satellite sensors were capable of detecting biophysical signatures associated with the wastewater, compared to ambient ocean waters, enabling monitoring of environmental impacts over a greater spatial extent than in situ sampling alone. Thermal satellite sensors measured decreased sea surface temperatures (SSTs) associated with the surfacing plumes. Ocean color satellite sensors did not measure a distinguishable biological response in terms of chlorophyll-a (chl-a) concentrations during the short lived, three-day long, 2006 HTP diversion. A period of decreased chl-a concentration was observed during the three-week long 2012 OCSD diversion, likely in association with enhanced chlorination of the discharged wastewater that suppressed the phytoplankton response and/or significant uptake by heterotrophic bacteria. Synthetic aperture radar (SAR) satellite data were able to identify and track the 2006 HTP wastewater plume through changes in surface roughness related to the oily components of the treated surfacing wastewater. Overall, it was found that chl-a and SST values must have differences of at least 1 mg m-3 and 0.5 °C, respectively, in comparison with adjacent waters for wastewater plumes and their biophysical impact to be detectable from satellite. For a wastewater plume to be identifiable in SAR imagery, wind speeds must range between ∼3 and 8 m s-1. The findings of this study illustrate the benefit of utilizing multiple satellite sensors to monitor the rapidly changing environmental response to surfacing wastewater plumes, and can help inform future wastewater diversions in coastal areas.

  15. Molecular Detection and Genotyping of Chlamydia psittaci in Captive Psittacines from Costa Rica

    Directory of Open Access Journals (Sweden)

    Jessica Sheleby-Elías

    2013-01-01

    Full Text Available Oropharyngeal and cloacal swabs from 117 captive psittacine birds presented at veterinary clinics (88 and from shelters/rescue centers of wildlife (29 were collected to determine the prevalence of C. psittaci in captive birds in Costa Rica. Samples were collected during 2009 from a total of 19 different species of parrots, with Ara macao (33, Amazona autumnalis (24, Amazona ochrocephala (21, and Ara ararauna (8 being the most representative species sampled. C. psittaci was detected in four (3.4% birds using molecular detection (PCR. The positive samples belonged to birds presented at veterinary clinics; three of them were Ara macao and one Amazona ochrocephala. Three birds were adults; all positive birds showed no symptoms of illness and lived in homes with other birds, two in San José and two in Heredia. Sequencing was used to confirm the PCR positive results, showing that two samples of C. psittaci belonged to genotype A, representing the first report of the presence of this genotype in Costa Rica. The detection of this bacterium in captive psittacine birds shows that there is a potential risk for people living or having contact with them and that there is a possibility of infecting other birds.

  16. Detection of hepatitis B virus genotypes using oligonucleotide chip among hepatitis B virus carriers in Eastern China

    Institute of Scientific and Technical Information of China (English)

    Xiang-Rong Tang; Ji-Shen Zhang; Hui Zhao; Yu-Hua Gong; Yong-Zhong Wang; Jian-Long Zhao

    2007-01-01

    AIM: To determine the genotype distribution of hepatitis B virus (HBV) with a newly oligonucleotide chip assay among the HBV carriers in Eastern China.METHODS: An assay using oligonucleotide chip was developed for detection of HBV genotypes in serum samples from HBV DNA-positive patients in Eastern China. This method is based on the principle of reverse hybridization with Cy5-labeled amplicons hybridizing to type-specific oligonucleotide probes that are immobilized on slides. The results of 80 randomly chosen sera were confirmed by direct sequencing.RESULTS: HBV genotype B, C and mixed genotype were detected in 400 serum samples, accounting for8.3% (n = 33), 83.2% (n = 333), and 8.5% (n = 34),respectively. The evaluation of the oligonucleotide assay showed 100% concordance with the amplicon phylogenetic analysis except 9 mixed genotype infections undetected by sequencing.CONCLUSION: The study indicates that HBV genotype C and B prevail in the Eastern China. It is suggested that the oligonucleotide chip is a reliable and convenient tool for the detection of HBV genotyping.

  17. Yellow fever virus: genetic and phenotypic diversity and implications for detection, prevention and therapy.

    Science.gov (United States)

    Beasley, David W C; McAuley, Alexander J; Bente, Dennis A

    2015-03-01

    Yellow fever virus (YFV) is the prototypical hemorrhagic fever virus, yet our understanding of its phenotypic diversity and any molecular basis for observed differences in disease severity and epidemiology is lacking, when compared to other arthropod-borne and haemorrhagic fever viruses. This is, in part, due to the availability of safe and effective vaccines resulting in basic YFV research taking a back seat to those viruses for which no effective vaccine occurs. However, regular outbreaks occur in endemic areas, and the spread of the virus to new, previously unaffected, areas is possible. Analysis of isolates from endemic areas reveals a strong geographic association for major genotypes, and recent epidemics have demonstrated the emergence of novel sequence variants. This review aims to outline the current understanding of YFV genetic and phenotypic diversity and its sources, as well as the available animal models for characterizing these differences in vivo. The consequences of genetic diversity for detection and diagnosis of yellow fever and development of new vaccines and therapeutics are discussed.

  18. Genotype diversity in structure of amylopectin of waxy rice and its influence on gelatinization properties.

    Science.gov (United States)

    Lin, Jheng-Hua; Singh, Harinder; Ciao, Jhih-Ying; Kao, Wen-Tzu; Huang, Wei-Hsiang; Chang, Yung-Ho

    2013-02-15

    A set of 13 waxy rice genotypes prepared by chemically-induced mutation of non-waxy rice variety TNG67 and 7 waxy rice varieties widely planted in Taiwan were screened for various structural and gelatinization properties of starches. Wide variation on physicochemical properties and molecular structure of amylopectin for the 20 waxy rice starches were obtained and relationship between gelatinization properties and molecular structure of starch were discussed. More attributes on swelling and gelatinization thermal properties, comparing to pasting attributes, showed significant correlation with molecular structure parameters. The swelling and gelatinization thermal properties of waxy rice starch did not show significant correlation with molecular size of amylopectin, while significant correlations were found between the swelling or gelatinization thermal properties and chain length of amylopectin. Results suggest that the swelling and pasting of waxy starch is essentially dominated by granule architecture and is dependent on the interactions among amylopectin chains. Copyright © 2012 Elsevier Ltd. All rights reserved.

  19. Diversity in the carotenoid profiles and the expression of genes related to carotenoid accumulation among citrus genotypes.

    Science.gov (United States)

    Ikoma, Yoshinori; Matsumoto, Hikaru; Kato, Masaya

    2016-01-01

    Carotenoids are not only important to the plants themselves but also are beneficial to human health. Since citrus fruit is a good source of carotenoids for the human diet, it is important to study carotenoid profiles and the accumulation mechanism in citrus fruit. Thus, in the present paper, we describe the diversity in the carotenoid profiles of fruit among citrus genotypes. In regard to carotenoids, such as β-cryptoxanthin, violaxanthin, lycopene, and β-citraurin, the relationship between the carotenoid profile and the expression of carotenoid-biosynthetic genes is discussed. Finally, recent results of quantitative trait locus (QTL) analyses of carotenoid contents and expression levels of carotenoid-biosynthetic genes in citrus fruit are shown.

  20. Genotyping of Leptospira directly in urine samples of cattle demonstrates a diversity of species and strains in Brazil.

    Science.gov (United States)

    Hamond, C; Pestana, C P; Medeiros, M A; Lilenbaum, W

    2016-01-01

    The aim of this study was to identify Leptospira in urine samples of cattle by direct sequencing of the secY gene. The validity of this approach was assessed using ten Leptospira strains obtained from cattle in Brazil and 77 DNA samples previously extracted from cattle urine, that were positive by PCR for the genus-specific lipL32 gene of Leptospira. Direct sequencing identified 24 (31·1%) interpretable secY sequences and these were identical to those obtained from direct DNA sequencing of the urine samples from which they were recovered. Phylogenetic analyses identified four species: L. interrogans, L. borgpetersenii, L. noguchii, and L. santarosai with the most prevalent genotypes being associated with L. borgpetersenii. While direct sequencing cannot, as yet, replace culturing of leptospires, it is a valid additional tool for epidemiological studies. An unexpected finding from this study was the genetic diversity of Leptospira infecting Brazilian cattle.

  1. Molecular and evolutionary bases of within-patient genotypic and phenotypic diversity in Escherichia coli extraintestinal infections.

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    Maxime Levert

    Full Text Available Although polymicrobial infections, caused by combinations of viruses, bacteria, fungi and parasites, are being recognised with increasing frequency, little is known about the occurrence of within-species diversity in bacterial infections and the molecular and evolutionary bases of this diversity. We used multiple approaches to study the genomic and phenotypic diversity among 226 Escherichia coli isolates from deep and closed visceral infections occurring in 19 patients. We observed genomic variability among isolates from the same site within 11 patients. This diversity was of two types, as patients were infected either by several distinct E. coli clones (4 patients or by members of a single clone that exhibit micro-heterogeneity (11 patients; both types of diversity were present in 4 patients. A surprisingly wide continuum of antibiotic resistance, outer membrane permeability, growth rate, stress resistance, red dry and rough morphotype characteristics and virulence properties were present within the isolates of single clones in 8 of the 11 patients showing genomic micro-heterogeneity. Many of the observed phenotypic differences within clones affected the trade-off between self-preservation and nutritional competence (SPANC. We showed in 3 patients that this phenotypic variability was associated with distinct levels of RpoS in co-existing isolates. Genome mutational analysis and global proteomic comparisons in isolates from a patient revealed a star-like relationship of changes amongst clonally diverging isolates. A mathematical model demonstrated that multiple genotypes with distinct RpoS levels can co-exist as a result of the SPANC trade-off. In the cases involving infection by a single clone, we present several lines of evidence to suggest diversification during the infectious process rather than an infection by multiple isolates exhibiting a micro-heterogeneity. Our results suggest that bacteria are subject to trade-offs during an infectious

  2. Molecular and evolutionary bases of within-patient genotypic and phenotypic diversity in Escherichia coli extraintestinal infections.

    Science.gov (United States)

    Levert, Maxime; Zamfir, Oana; Clermont, Olivier; Bouvet, Odile; Lespinats, Sylvain; Hipeaux, Marie Claire; Branger, Catherine; Picard, Bertrand; Saint-Ruf, Claude; Norel, Françoise; Balliau, Thierry; Zivy, Michel; Le Nagard, Hervé; Cruveiller, Stéphane; Cruvellier, Stéphane; Chane-Woon-Ming, Béatrice; Nilsson, Susanna; Gudelj, Ivana; Phan, Katherine; Ferenci, Thomas; Tenaillon, Olivier; Denamur, Erick

    2010-09-30

    Although polymicrobial infections, caused by combinations of viruses, bacteria, fungi and parasites, are being recognised with increasing frequency, little is known about the occurrence of within-species diversity in bacterial infections and the molecular and evolutionary bases of this diversity. We used multiple approaches to study the genomic and phenotypic diversity among 226 Escherichia coli isolates from deep and closed visceral infections occurring in 19 patients. We observed genomic variability among isolates from the same site within 11 patients. This diversity was of two types, as patients were infected either by several distinct E. coli clones (4 patients) or by members of a single clone that exhibit micro-heterogeneity (11 patients); both types of diversity were present in 4 patients. A surprisingly wide continuum of antibiotic resistance, outer membrane permeability, growth rate, stress resistance, red dry and rough morphotype characteristics and virulence properties were present within the isolates of single clones in 8 of the 11 patients showing genomic micro-heterogeneity. Many of the observed phenotypic differences within clones affected the trade-off between self-preservation and nutritional competence (SPANC). We showed in 3 patients that this phenotypic variability was associated with distinct levels of RpoS in co-existing isolates. Genome mutational analysis and global proteomic comparisons in isolates from a patient revealed a star-like relationship of changes amongst clonally diverging isolates. A mathematical model demonstrated that multiple genotypes with distinct RpoS levels can co-exist as a result of the SPANC trade-off. In the cases involving infection by a single clone, we present several lines of evidence to suggest diversification during the infectious process rather than an infection by multiple isolates exhibiting a micro-heterogeneity. Our results suggest that bacteria are subject to trade-offs during an infectious process and that

  3. The genetic diversity of triticale genotypes involved in Polish breeding programs.

    Science.gov (United States)

    Niedziela, Agnieszka; Orłowska, Renata; Machczyńska, Joanna; Bednarek, Piotr T

    2016-01-01

    Genetic diversity analysis of triticale populations is useful for breeding programs, as it helps to select appropriate genetic material for classifying the parental lines, heterotic groups and predicting hybrid performance. In our study 232 breeding forms were analyzed using diversity arrays technology markers. Principal coordinate analysis followed by model-based Bayesian analysis of population structure revealed the presence of weak data structuring with three groups of data. In the first group, 17 spring and 17 winter forms were clustered. The second and the third groups were represented by 101 and 26 winter forms, respectively. Polymorphic information content values, as well as Shannon's Information Index, were higher for the first (0.319) and second (0.309) than for third (0.234) group. AMOVA analysis demonstrated a higher level of within variation (86 %) than among populations (14 %). This study provides the basic information on the presence of structure within a genetic pool of triticale breeding forms.

  4. Genotypic diversity among rhizospheric bacteria of three legumes assessed by cultivation-dependent and cultivation-independent techniques.

    Science.gov (United States)

    Pongsilp, Neelawan; Nimnoi, Pongrawee; Lumyong, Saisamorn

    2012-02-01

    The genotypic diversity of rhizospheric bacteria of 3 legumes including Vigna radiata, Arachis hypogaea and Acacia mangium was compared by using cultivation-dependent and cultivation-independent methods. For cultivation-dependent method, Random amplified polymorphic DNA (RAPD) profiles revealed that the bacterial genetic diversity of V. radiata and A. mangium rhizospheres was higher than that of A. hypogaea rhizosphere. For cultivation-independent method, Denaturing gradient gel electrophoresis (DGGE) profiles of PCR-amplified 16S rRNA genes revealed the difference in bacterial community and diversity of rhizospheres collected from 3 legumes. The ribotype richness which indicates species diversity, was highest in V. radiata rhizosphere, followed by A. hypogaea and A. mangium rhizospheres, respectively. Three kinds of media were used to cultivate different target groups of bacteria. The result indicates that the communities of cultivable bacteria in 3 rhizospheres recovered from nutrient agar (NA) medium were mostly different from each other, while Bradyrhizobium selective medium (BJSM) and nitrogen-free medium shaped the communities of cultivable bacteria. Nine isolates grown on BJSM were identified by 16S rRNA gene sequence analysis. These isolates were very closely related (with 96% to 99% identities) to either one of the three groups including Cupriavidus-Ralstonia group, Bacillus group and Bradyrhizobium-Bosea-Afipia group. The rhizospheres were also examined for their enzymatic patterns. Of 19 enzymes tested, 3 rhizospheres were distinguishable by the presence or the absence of leucine acrylamidase and acid phosphatase. The selected cultivable bacteria recovered from NA varied in their abilities to produce indole-acetic acid and ammnonia. The resistance to 10 antibiotics was indistinguishable among bacteria isolated from different rhizospheres.

  5. Five species, many genotypes, broad phenotypic diversity: When agronomy meets functional ecology.

    Science.gov (United States)

    Prieto, Ivan; Litrico, Isabelle; Violle, Cyrille; Barre, Philippe

    2017-01-01

    Current ecological theory can provide insight into the causes and impacts of plant domestication. However, just how domestication has impacted intraspecific genetic variability (ITV) is unknown. We used 50 ecotypes and 35 cultivars from five grassland species to explore how selection drives functional trait coordination and genetic differentiation. We quantified the extent of genetic diversity among different sets of functional traits and determined how much genetic diversity has been generated within populations of natural ecotypes and selected cultivars. In general, the cultivars were larger (e.g., greater height, faster growth rates) and had larger and thinner leaves (greater SLA). We found large (average 63%) and trait-dependent (ranging from 14% for LNC to 95.8% for growth rate) genetic variability. The relative extent of genetic variability was greater for whole-plant than for organ-level traits. This pattern was consistent within ecotypes and within cultivars. However, ecotypes presented greater ITV variability. The results indicated that genetic diversity is large in domesticated species with contrasting levels of heritability among functional traits and that selection for high yield has led to indirect selection of some associated leaf traits. These findings open the way to define which target traits should be the focus in selection programs, especially in the context of community-level selection. © 2017 Botanical Society of America.

  6. Novel genotype of Ehrlichia canis detected in samples of human blood bank donors in Costa Rica.

    Science.gov (United States)

    Bouza-Mora, Laura; Dolz, Gaby; Solórzano-Morales, Antony; Romero-Zuñiga, Juan José; Salazar-Sánchez, Lizbeth; Labruna, Marcelo B; Aguiar, Daniel M

    2017-01-01

    This study focuses on the detection and identification of DNA and antibodies to Ehrlichia spp. in samples of blood bank donors in Costa Rica using molecular and serological techniques. Presence of Ehrlichia canis was determined in 10 (3.6%) out of 280 blood samples using polymerase chain reaction (PCR) targeting the ehrlichial dsb conserved gene. Analysis of the ehrlichial trp36 polymorphic gene in these 10 samples revealed substantial polymorphism among the E. canis genotypes, including divergent tandem repeat sequences. Nucleotide sequences of dsb and trp36 amplicons revealed a novel genotype of E. canis in blood bank donors from Costa Rica. Indirect immunofluorescence assay (IFA) detected antibodies in 35 (35%) of 100 serum samples evaluated. Thirty samples showed low endpoint titers (64-256) to E. canis, whereas five sera yielded high endpoint titers (1024-8192); these five samples were also E. canis-PCR positive. These findings represent the first report of the presence of E. canis in humans in Central America.

  7. Multi-locus genotypes of Enterocytozoon bieneusi in captive Asiatic black bears in southwestern China: High genetic diversity, broad host range, and zoonotic potential

    Science.gov (United States)

    Cao, Xuefeng; Song, Yuan; Wang, Wuyou; Huang, Xiangming; Liu, Xuehan; Hu, Yanchun; Fu, Hualin; He, Min; Wang, Ya; Zhang, Yue; Wu, Kongju; Peng, Guangneng

    2017-01-01

    Enterocytozoon bieneusi is an obligate eukaryotic intracellular parasite that infects a wide variety of vertebrate and invertebrate hosts. Although considerable research has been conducted on this organism, relatively little information is available on the occurrence of E. bieneusi in captive Asiatic black bears. The present study was performed to determine the prevalence, genetic diversity, and zoonotic potential of E. bieneusi in captive Asiatic black bears in zoos in southwestern China. Fecal specimens from Asiatic black bears in four zoos, located in four different cities, were collected and analyzed for the prevalence of E. bieneusi. The average prevalence of E. bieneusi was 27.4% (29/106), with the highest prevalence in Guiyang Zoo (36.4%, 16/44). Altogether, five genotypes of E. bieneusi were identified among the 29 E. bieneusi-positive samples, including three known genotypes (CHB1, SC02, and horse2) and two novel genotypes named ABB1 and ABB2. Multi-locus sequence typing using three microsatellites (MS1, MS3, and MS7) and one minisatellite (MS4) revealed V, III, V, and IV genotypes at these four loci, respectively. Phylogenetic analysis showed that the genotypes SC02 and ABB2 were clustered into group 1 of zoonotic potential, the genotypes CHB1 and ABB1 were clustered into a new group, and the genotype horse2 was clustered into group 6 of unclear zoonotic potential. In conclusion, this study identified two novel E. bieneusi genotypes in captive Asiatic black bears, and used microsatellite and minisatellite markers to reveal E. bieneusi genetic diversity. Moreover, our findings show that genotypes SC02 (identified in humans) and ABB2 belong to group 1 with zoonotic potential, suggesting the risk of transmission of E. bieneusi from Asiatic black bears to humans and other animals. PMID:28182656

  8. Genotyping of Norovirus strains detected in outbreaks between April 2002 and March 2003 in Osaka City, Japan.

    Science.gov (United States)

    Seto, Yoshiyuki; Iritani, Nobuhiro; Kubo, Hideyuki; Kaida, Atsushi; Murakami, Tsukasa; Haruki, Kosuke; Nishio, Osamu; Ayata, Minoru; Ogura, Hisashi

    2005-01-01

    Noroviruses (NVs) are the major cause of food- and waterborne nonbacterial gastroenteritis in Japan. Between April 2002 and March 2003, a total of 111 fecal specimens from 40 outbreaks of acute nonbacterial gastroenteritis in Osaka City, Japan were subject to NV detection. Seventy-two samples (64.9%) from 31 outbreaks (77.5%) were NV positive by a real time reverse transcription (RT)-PCR assay. To further determine the genotype of individual NV strains, we sequenced the capsid N-terminal/shell (N/S) domain of some representative strains from each outbreak. The 51 NV strains detected in this study were segregated into 15 genotypes (6 in genogroup I and 9 in genogroup II), and GII/5 genotype NV was a dominant outbreak genotype.

  9. Detection of a new subgenotype of hepatitis B virus genotype A in Cameroon but not in neighbouring Nigeria.

    Science.gov (United States)

    Hübschen, J M; Mbah, P O; Forbi, J C; Otegbayo, J A; Olinger, C M; Charpentier, E; Muller, C P

    2011-01-01

    To further investigate the genetic diversity of hepatitis B virus (HBV) genotype A in Africa, we analysed 263 HBV strains from Nigeria (n=163) and Cameroon (n=100). Phylogenetic analysis of S fragment sequences attributed 175 strains (66.5%) to genotype E and 88 (33.5%) to genotype A. In Cameroon, genotype A strains were the most prevalent (79/100, 79.0%), whereas, in Nigeria, genotype E was highly dominant (154/163, 94.5%). The genotype A strains grouped with reference strains of subgenotype A3 (n=8), the provisional subgenotype A5 (n=43), a recently reported new variant from Rwanda (n=35), or as outliers (n=2). Ten complete genome sequences obtained from strains that clustered with the new variant from Rwanda formed a separate group supported by a bootstrap value of 96. The between-group distance to other potential or recognized subgenotypes of genotype A was at least 3.81%. Thus, the new group of strains could be considered as a new subgenotype of HBV genotype A, tentatively named 'A7'. Interestingly, the 'A7' strains from Rwanda and Cameroon showed an interspersed clustering, but essentially no other (sub)genotypes were shared between the two countries, suggesting that 'A7' may have evolved in a yet unknown place and may have only relatively recently spread to Rwanda and Western Cameroon. Strains attributed to provisional subgenotype A5 were found for the first time in Cameroon (n=36) and Central Nigeria (n=2), indicating that A5 is more widespread than previously thought.

  10. Lentil (Lens culinaris Medik.) nodulates with genotypically and phenotypically diverse rhizobia in Ethiopian soils.

    Science.gov (United States)

    Tena, Wondwosen; Wolde-Meskel, Endalkachew; Degefu, Tulu; Walley, Fran

    2017-01-01

    Forty-eight lentil-nodulating rhizobia were isolated from soil samples collected from diverse agro-ecological locations in Ethiopia, and characterized based on 76 phenotypic traits. Furthermore, 26 representative strains were selected and characterized using multilocus sequence analyses (MLSA) of core (16S rRNA, recA, atpD, glnII and gyrB) and symbiotic (nodA and nifH) genes. Numerical analysis of phenotypic characteristics showed that the 48 test strains fell into three major distinct clusters. The phylogenetic trees based on 16S rRNA genes showed that they belong to the Rhizobium genus. Our phylogenetic reconstruction based on combined gene trees (recA, atpD and glnII) supported three distinct sub-lineages (Clades I-III). While genospecies I and II could be classified with Rhizobium etli and Rhizobium leguminosarum, respectively, genospecies III, might be an unnamed genospecies within the genus Rhizobium. Phylogenetic reconstruction based on the symbiosis-related genes supported a single cluster, indicating differences in the evolutionary histories between chromosomal and symbiotic genes. Overall, these results confirmed the presence of a great diversity of lentil-nodulating Rhizobium species in Ethiopia, inviting further exploration. Moreover, the differences in symbiotic effectiveness of the test strains indicated the potential for selecting and using them as inoculants to improve the productivity of lentil in the country.

  11. Detection of Francisella tularensis in ticks and identification of their genotypes using multiple-locus variable-number tandem repeat analysis

    Directory of Open Access Journals (Sweden)

    Yang Hong

    2008-09-01

    Full Text Available Abstract Background Tularemia was reported in China over 50 years ago, however, many epidemical characteristics remain unclear. In the present study, the prevalence of Francisella tularensis in ticks was investigated during an epidemiological surveillance in China and then we measured their genetic diversity by conducting multiple-locus variable- number tandem repeat analysis (MLVA. Results 1670 ticks from 2 endemic areas (Inner Mongolia Autonomous Region and Heilongjiang Province and 2 non-endemic areas (Jilin and Fujian Provinces were collected and tested for evidence of tularemia by nested PCR. The prevalence of Francisella tularensis in ticks averaged 1.98%. The positive rates were significantly different among tick species, with Dermacentor silvarum and Ixodes persulatus responsible for all positive numbers. All F. tularensis that were detected in ticks belonged to F. tularensis subsp. holarctica and MLVA disclosed genetic diversity. One subtype was identified in 17 of 33 positive tick samples in three different study areas. Another subtype belonging to F. tularensis subsp. holarctica genotype was described for the first time in the current study. Conclusion The study showed two tick species, D. silvarum and I. persulatus harboring the pathogen of tularemia in natural environment, indicating these two tick species might have a role in tularemia existence in China. MLVA results disclosed the genetic diversity F. tularensis and identified one genotype as the most prevalent among the investigated ticks in China.

  12. Genetic Diversity and Selective Pressure in Hepatitis C Virus Genotypes 1–6: Significance for Direct-Acting Antiviral Treatment and Drug Resistance

    Science.gov (United States)

    Cuypers, Lize; Li, Guangdi; Libin, Pieter; Piampongsant, Supinya; Vandamme, Anne-Mieke; Theys, Kristof

    2015-01-01

    Treatment with pan-genotypic direct-acting antivirals, targeting different viral proteins, is the best option for clearing hepatitis C virus (HCV) infection in chronically infected patients. However, the diversity of the HCV genome is a major obstacle for the development of antiviral drugs, vaccines, and genotyping assays. In this large-scale analysis, genome-wide diversity and selective pressure was mapped, focusing on positions important for treatment, drug resistance, and resistance testing. A dataset of 1415 full-genome sequences, including genotypes 1–6 from the Los Alamos database, was analyzed. In 44% of all full-genome positions, the consensus amino acid was different for at least one genotype. Focusing on positions sharing the same consensus amino acid in all genotypes revealed that only 15% was defined as pan-genotypic highly conserved (≥99% amino acid identity) and an additional 24% as pan-genotypic conserved (≥95%). Despite its large genetic diversity, across all genotypes, codon positions were rarely identified to be positively selected (0.23%–0.46%) and predominantly found to be under negative selective pressure, suggesting mainly neutral evolution. For NS3, NS5A, and NS5B, respectively, 40% (6/15), 33% (3/9), and 14% (2/14) of the resistance-related positions harbored as consensus the amino acid variant related to resistance, potentially impeding treatment. For example, the NS3 variant 80K, conferring resistance to simeprevir used for treatment of HCV1 infected patients, was present in 39.3% of the HCV1a strains and 0.25% of HCV1b strains. Both NS5A variants 28M and 30S, known to be associated with resistance to the pan-genotypic drug daclatasvir, were found in a significant proportion of HCV4 strains (10.7%). NS5B variant 556G, known to confer resistance to non-nucleoside inhibitor dasabuvir, was observed in 8.4% of the HCV1b strains. Given the large HCV genetic diversity, sequencing efforts for resistance testing purposes may need to be

  13. Genetic Diversity and Selective Pressure in Hepatitis C Virus Genotypes 1–6: Significance for Direct-Acting Antiviral Treatment and Drug Resistance

    Directory of Open Access Journals (Sweden)

    Lize Cuypers

    2015-09-01

    Full Text Available Treatment with pan-genotypic direct-acting antivirals, targeting different viral proteins, is the best option for clearing hepatitis C virus (HCV infection in chronically infected patients. However, the diversity of the HCV genome is a major obstacle for the development of antiviral drugs, vaccines, and genotyping assays. In this large-scale analysis, genome-wide diversity and selective pressure was mapped, focusing on positions important for treatment, drug resistance, and resistance testing. A dataset of 1415 full-genome sequences, including genotypes 1–6 from the Los Alamos database, was analyzed. In 44% of all full-genome positions, the consensus amino acid was different for at least one genotype. Focusing on positions sharing the same consensus amino acid in all genotypes revealed that only 15% was defined as pan-genotypic highly conserved (≥99% amino acid identity and an additional 24% as pan-genotypic conserved (≥95%. Despite its large genetic diversity, across all genotypes, codon positions were rarely identified to be positively selected (0.23%–0.46% and predominantly found to be under negative selective pressure, suggesting mainly neutral evolution. For NS3, NS5A, and NS5B, respectively, 40% (6/15, 33% (3/9, and 14% (2/14 of the resistance-related positions harbored as consensus the amino acid variant related to resistance, potentially impeding treatment. For example, the NS3 variant 80K, conferring resistance to simeprevir used for treatment of HCV1 infected patients, was present in 39.3% of the HCV1a strains and 0.25% of HCV1b strains. Both NS5A variants 28M and 30S, known to be associated with resistance to the pan-genotypic drug daclatasvir, were found in a significant proportion of HCV4 strains (10.7%. NS5B variant 556G, known to confer resistance to non-nucleoside inhibitor dasabuvir, was observed in 8.4% of the HCV1b strains. Given the large HCV genetic diversity, sequencing efforts for resistance testing purposes may

  14. SNiPlay: a web-based tool for detection, management and analysis of SNPs. Application to grapevine diversity projects.

    Science.gov (United States)

    Dereeper, Alexis; Nicolas, Stéphane; Le Cunff, Loïc; Bacilieri, Roberto; Doligez, Agnès; Peros, Jean-Pierre; Ruiz, Manuel; This, Patrice

    2011-05-05

    High-throughput re-sequencing, new genotyping technologies and the availability of reference genomes allow the extensive characterization of Single Nucleotide Polymorphisms (SNPs) and insertion/deletion events (indels) in many plant species. The rapidly increasing amount of re-sequencing and genotyping data generated by large-scale genetic diversity projects requires the development of integrated bioinformatics tools able to efficiently manage, analyze, and combine these genetic data with genome structure and external data. In this context, we developed SNiPlay, a flexible, user-friendly and integrative web-based tool dedicated to polymorphism discovery and analysis. It integrates:1) a pipeline, freely accessible through the internet, combining existing softwares with new tools to detect SNPs and to compute different types of statistical indices and graphical layouts for SNP data. From standard sequence alignments, genotyping data or Sanger sequencing traces given as input, SNiPlay detects SNPs and indels events and outputs submission files for the design of Illumina's SNP chips. Subsequently, it sends sequences and genotyping data into a series of modules in charge of various processes: physical mapping to a reference genome, annotation (genomic position, intron/exon location, synonymous/non-synonymous substitutions), SNP frequency determination in user-defined groups, haplotype reconstruction and network, linkage disequilibrium evaluation, and diversity analysis (Pi, Watterson's Theta, Tajima's D).Furthermore, the pipeline allows the use of external data (such as phenotype, geographic origin, taxa, stratification) to define groups and compare statistical indices.2) a database storing polymorphisms, genotyping data and grapevine sequences released by public and private projects. It allows the user to retrieve SNPs using various filters (such as genomic position, missing data, polymorphism type, allele frequency), to compare SNP patterns between populations, and to

  15. A comparison of growth and development patterns in diverse genotypes of broilers. 2. Pullet growth.

    Science.gov (United States)

    Reddish, J M; Lilburn, M S

    2004-07-01

    Genetic selection within commercial broiler lines continues to generate improvements in BW, feed conversion, and breast meat yield. The objective of the current experiment was to compare carcass and reproductive characteristics of broiler breeder pullets from 2 dam lines that produce heavy broilers with increased breast yield (A and B) with pullets from a commercial line that does not have the extremes in breast yield (C). Restricted-fed BW were similar at all ages of the experiment in the 3 genotypes. All pullets were photostimulated at 23 wk of age, and carcass and reproductive tract measurements were made at 27 wk of age. There were no differences in BW among the lines at 27 wk of age, but the weights of the pectoralis major and minor breast muscles were heavier in lines A and B compared with line C (P abdominal fad pad, however, was heavier in line C (P < 0.04). There was no difference in total drum weight or total thigh weight among lines A, B, or C. Tibia length and tibia width were similar in lines A and C, and the measurements were larger in these lines than in line B (P < 0.02). Femur length was longer (P < 0.001) in line C than in lines A and B, whereas femur width was greater in line C than in line A (P < 0.001) but similar to line B (P < 0.001). Oviduct weight was greater in line C compared with lines A and B (P < 0.004), but there were no differences in total ovarian weight, follicle number, or follicle weight. The information gathered in the present experiment suggested that line C may prove to be useful for reproductive comparisons with commercial lines exhibiting significant differences in carcass traits.

  16. Distribution of AdeABC efflux system genes in genotypically diverse strains of clinical Acinetobacter baumannii.

    Science.gov (United States)

    Wieczorek, Piotr; Sacha, Paweł; Czaban, Sławomir; Hauschild, Tomasz; Ojdana, Dominika; Kowalczuk, Oksana; Milewski, Robert; Poniatowski, Bogusław; Nikliński, Jacek; Tryniszewska, Elżbieta

    2013-10-01

    Acinetobacter baumannii has emerged as a highly problematic hospital-associated pathogen. Different mechanisms contribute to the formation of multidrug resistance in A. baumannii, including the AdeABC efflux system. Distribution of the structural and regulatory genes encoding the AdeABC efflux system among genetically diverse clinical A. baumannii strains was achieved by using PCR and pulsed-field gel electrophoresis techniques. The distribution of adeABRS genes is extremely high among our A. baumannii strains, except the adeC gene. We have observed a large proportion of strains presenting multidrug-resistance phenotype for several years. The efflux pump could be an important mechanism in these strains in resistance to antibiotics.

  17. [Effectiveness of human papillomavirus genotyping for detection of high-grade anal intraepithelial neoplasia compared to anal cytology].

    Science.gov (United States)

    Padilla-España, Laura; Repiso-Jiménez, Juan Bosco; Fernández-Sánchez, Fernando; Pereda, Teresa; Rivas-Ruiz, Francisco; Fernández-Morano, Teresa; de la Torre-Lima, Javier; Palma, Fermín; Redondo, Maximino; de Troya-Martín, Magdalena

    2016-01-01

    The incidence of high-grade anal intraepithelial neoplasia (HGAIN) -with an aetiological based on high-risk types of human papillomavirus- is increasing in some high-risk groups. Screening for HGAIN includes routine anal cytology and, more recently, HPV genotyping. The main objective of this study was to determine the sensitivity and specificity of anal cytology and HPV genotyping for the detection of HGAIN. This is a study to determine the correlation of cytological and microbiological findings with anal biopsy findings in a cohort of patients at high risk of developing AIN referred to the department of sexually transmitted infections of the Hospital Costa del Sol, Spain, between January 2008 and December 2014. Of the 151 patients subjected to screening, a total of 92 patients, all of them with the result of three screening test (anal cytology, genotyping and biopsy) were included in the study. Just under two-thirds (62%) of them were HIV-positive. The sensitivity and specificity of anal cytology to detect HGAIN were 52.8 and 85.7%, respectively (k: 0.328), and 78 and 62.8% to detect two or more HPV oncogenic genotypes (k: 0.417). The detection of oncogenic HPV genotypes allowed the identification of 23 new cases of HGAIN that had been underdiagnosed with anal cytology, with 14 cases containing at least three high-risk genotypes. Anal cytology did not show enough sensitivity in HGAIN screening. HPV genotyping has shown to be a useful tool to detect HGAIN cases, although it could lead to an over-diagnosis as a solitary screening procedure. Copyright © 2016 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

  18. High Prevalence and Genotype Diversity of Anal HPV Infection among MSM in Northern Thailand.

    Directory of Open Access Journals (Sweden)

    Taweewat Supindham

    Full Text Available HPV infection is common and may cause cancer among men who have sex with men (MSM. Anal HPV infection (HPV+ was found in 85% of HIV-positive (HIV+ and 59% of HIV-negative (HIV- MSM in Bangkok, central Thailand. As little is known about HPV in this group in northern Thailand, we studied MSM subgroups comprised of gay men (GM, bisexual men (BM, and transgender women (TGW.From July 2012 through January 2013, 85 (42.5% of 200 GM, 30 (15% BM, and 85 (42.5% TGW who practiced receptive anal intercourse were recruited after informed consent, followed by self-assisted computer interview, HIV testing, and anal swabs for HPV genotyping.Of 197 adequate specimens, the overall prevalence of any HPV was 157 (80%. Prevalence was 89% (76/85 in GM, 48% (14/29 in BM, and 81% (67/83 in TGW. The most common high-risk types were HPV16 (27% of 197, HPV58 (23%, and HPV51 (18%. Prevalence of high-risk types was 74% in 85 GM, 35% in 29 BM, and 71% in 83 TGW. Prevalence of any HPV type, or high-risk type, was 100% and 94%, respectively, among 48 HIV+ MSM, 70% and 54% among 120 HIV- MSM. Of the 197 specimens, 36% (70 had HPV types 16 and/or 18 in the bivalent vaccine, compared to 48% (95 with ≥1 of types 16/18/06/11 in the quadrivalent, 56% (111 for 16/18/31/33/45/52/58 in the 7-valent, and 64% (126 for 16/18/31/33/45/52/58/06/11 in the 9-valent. HIV+, GM, and TGW were independently associated with HPV infection.We found higher rates of both any HPV and high-risk types than previous studies. Among the heretofore unstudied TGW, their equivalent HPV rates were comparable to GM. Current and investigational HPV vaccines could substantially protect GM, BM, and TGW from the serious consequences of HPV infection especially among HIV + MSM.

  19. Simultaneous detection and differentiation of three genotypes of Brassica yellows virus by multiplex reverse transcription-polymerase chain reaction.

    Science.gov (United States)

    Zhang, Xiaoyan; Peng, Yanmei; Wang, Ying; Zhang, Zongying; Li, Dawei; Yu, Jialin; Han, Chenggui

    2016-11-22

    Brassica yellows virus (BrYV), proposed to be a new polerovirus species, three distinct genotypes (BrYV-A, BrYV-B and BrYV-C) have been described. This study was to develop a simple, rapid, sensitive, cost-effective method for simultaneous detection and differentiation of three genotypes of BrYV. In this study, a multiplex reverse transcription-polymerase chain reaction (mRT-PCR) was developed for simultaneous detection and differentiation of the three genotypes of BrYV. The three genotypes of BrYV and Tunip yellows virus (TuYV) could be differentiated simultaneously using six optimized specific oligonucleotide primers, including one universal primer for detecting BrYV, three BrYV genotype-specific primers, and a pair of primers for specific detection of TuYV. Primers were designed from conserved regions of each virus and their specificity was confirmed by sequencing PCR products. The mRT-PCR products were 278 bp for BrYV-A, 674 bp for BrYV-B, 505 bp for BrYV-C, and 205 bp for TuYV. Amplification of three target genotypes was optimized by increasing the PCR annealing temperatures to 62 °C. One to three fragments specific for the virus genotypes were simultaneously amplified from infected samples and identified by their specific molecular sizes in agarose gel electrophoresis. No specific products could be amplified from cDNAs of other viruses which could infect crucifer crops. Detection limits of the plasmids for multiplex PCR were 100 fg for BrYV-A and BrYV-B, 10 pg for BrYV-C, and 1 pg for TuYV, respectively. The mRT-PCR was applied successfully for detection of three BrYV genotypes from field samples collected in China. The simple, rapid, sensitive, and cost-effective mRT-PCR was developed successfully for detection and differentiation of the three genotypes of BrYV.

  20. Simultaneous detection of hepatitis B virus genotypes and mutations associated with resistance to lamivudine, adefovir, and telbivudine by the polymerase chain reaction-ligase detection reaction

    Directory of Open Access Journals (Sweden)

    Yong-Zhong Wang

    2011-12-01

    Full Text Available OBJECTIVES: Detection of mutations associated to nucleos(tide analogs and hepatitis B virus (HBV genotyping are essential for monitoring treatment of HBV infection. We developed a multiplex polymerase chain reaction-ligase detection reaction (PCR-LDR assay for the rapid detection of HBV genotypes and mutations associated with lamivudine, adefovir, and telbivudine resistance in HBV-infected patients. METHODS: HBV templates were amplified by PCR, followed by LDR and electrophoresis on a sequencer. The assay was evaluated using plasmids that contained wild-type or mutant HBV sequences and 216 clinical samples. RESULTS: The PCR-LDR assay and sequencing gave comparable results for 158 of the 216 samples (73.1% with respect to mutation detection and genotyping. Complete agreement between the two methods was observed for all the samples (100% at codon 180 and codon 204. Concordant results were observed for 99.4% of the 158 samples at codon 181 and 98.7% at codon 236. The genotyping results were completely concordant between the PCR-LDR assay and sequencing. The PCR-LDR assay could detect a proportion of 1% mutant plasmid in a background of wild-type plasmid. CONCLUSION: The PCR-LDR assay is sensitive and specific for detection of HBV genotypes and drug resistance mutations, and could be helpful for decision making in the treatment of HBV infection.

  1. Phenotypic and genotypic diversity of rhizobia nodulating Pterocarpus erinaceus and P. lucens in Senegal.

    Science.gov (United States)

    Sylla, Samba Ndao; Samba, Ramatoulaye Thiaba; Neyra, Marc; Ndoye, Ibrahima; Giraud, Eric; Willems, Anne; de Lajudie, Philippe; Dreyfus, Bernard

    2002-12-01

    A total of fifty root nodules isolates of fast-growing and slow growing rhizobia from Pterocarpus ennaceus and Pterocarpus lucens respectively native of sudanean and sahelian regions of Senegal were characterized. These isolates were compared to representative strains of known rhizobial species. Twenty-two new isolates were slow growers and twenty-eight were fast growers. A polyphasic approach was performed including comparative total protein sodium dodecyl sulphate polyacrylamide gel (SDS-PAGE) profile analysis; 16S rDNA and 16S-23S rDNA intergenic spacer (IGS) sequence analysis. By SDS-PAGE the slow growing isolates grouped in one major cluster containing reference strains of Bradyrhizobium sp. including strains isolated in Africa, in Brazil and in New Zealand. Most of the fast-growing rhizobia grouped in four different clusters or were separate strains related to Rhizobium and Mesorhizobium strains. The 16S rDNA and 16S-23S rDNA IGS sequences analysis showed accurately the differentiation of fast growing rhizobia among the Rhizobium and Mesorbizobium genospecies. The representative strains of slow growing rhizobia were identified as closely related to Bradyrbizobium elkanii and Bradyrhizobium japonicum. Based on 16S rDNA sequence analysis, one slow growing strain (ORS199) was phylogenetically related to Bradyrbizobium sp. (Lupinus) and Blastobacter denitrificans. This position of ORS 199 was not confirmed by IGS sequence divergence. We found no clear relation between the diversity of strains, the host plants and the ecogeographical origins.

  2. Evaluation of DNA Extraction Methods Suitable for PCR-based Detection and Genotyping of Clostridium botulinum

    DEFF Research Database (Denmark)

    Auricchio, Bruna; Anniballi, Fabrizio; Fiore, Alfonsina

    2013-01-01

    Sufficient quality and quantity of extracted DNA is critical to detecting and performing genotyping of Clostridium botulinum by means of PCR-based methods. An ideal extraction method has to optimize DNA yield, minimize DNA degradation, allow multiple samples to be extracted, and be efficient...... in terms of cost, time, labor, and supplies. Eleven botulinum toxin–producing clostridia strains and 25 samples (10 food, 13 clinical, and 2 environmental samples) naturally contaminated with botulinum toxin–producing clostridia were used to compare 4 DNA extraction procedures: Chelex® 100 matrix, Phenol......-Cloroform-Isoamyl alcohol, NucliSENS® magnetic extraction kit, and DNeasy® Blood & Tissue kit. Integrity, purity, and amount of amplifiable DNA were evaluated. The results show that the DNeasy® Blood & Tissue kit is the best extraction method evaluated because it provided the most pure, intact, and amplifiable DNA. However...

  3. Selective genotyping to detect QTL for multiple traits in outbred populations

    NARCIS (Netherlands)

    Bovenhuis, H.; Spelman, R.J.

    2000-01-01

    Selective genotyping (i.e., genotyping of individuals that are extreme for a quantitative trait) has been undertaken in many studies but has usually been limited to one trait of interest. This paper outlines the implications of selective genotyping when applied to a daughter design. Formulas are pre

  4. Helicobacter Genotyping and Detection in Peroperative Lavage Fluid in Patients with Perforated Peptic Ulcer

    NARCIS (Netherlands)

    N.A.P. Komen (Niels); M.J.O.E. Bertleff (Marietta); J.F. Lange (Johan); P.W. de Graaf

    2008-01-01

    textabstractIntroduction and Objectives Certain Helicobacter pylori genotypes are associated with peptic ulcer disease; however, little is known about associations between the H. pylori genotype and perforated peptic ulcer (PPU). The primary aim of this study was to evaluate which genotypes are pre

  5. Genotypic and phenotypic diversity of rhizobia isolated from Lathyrus japonicus indigenous to Japan.

    Science.gov (United States)

    Aoki, Seishiro; Kondo, Tetsuya; Prévost, Danielle; Nakata, Sayuri; Kajita, Tadashi; Ito, Motomi

    2010-11-01

    Sixty-one rhizobial strains from Lathyrus japonicus nodules growing on the seashore in Japan were characterized and compared to two strains from Canada. The PCR-based method was used to identify test strains with novel taxonomic markers that were designed to discriminate between all known Lathyrus rhizobia. Three genomic groups (I, II, and III) were finally identified using RAPD, RFLP, and phylogenetic analyses. Strains in genomic group I (related to Rhizobium leguminosarum) were divided into two subgroups (Ia and Ib) and subgroup Ia was related to biovar viciae. Strains in subgroup Ib, which were all isolated from Japanese sea pea, belonged to a distinct group from other rhizobial groups in the recA phylogeny and PCR-based grouping, and were more tolerant to salt than the isolate from an inland legume. Test strains in genomic groups II and III belonged to a single clade with the reference strains of R. pisi, R. etli, and R. phaseoli in the 16S rRNA phylogeny. The PCR-based method and phylogenetic analysis of recA revealed that genomic group II was related to R. pisi. The analyses also showed that genomic group III harbored a mixed chromosomal sequence of different genomic groups, suggesting a recent horizontal gene transfer between diverse rhizobia. Although two Canadian strains belonged to subgroup Ia, molecular and physiological analyses showed the divergence between Canadian and Japanese strains. Phylogenetic analysis of nod genes divided the rhizobial strains into several groups that reflected the host range of rhizobia. Symbiosis between dispersing legumes and rhizobia at seashore is discussed.

  6. Phased genotyping-by-sequencing enhances analysis of genetic diversity and reveals divergent copy number variants in maize

    Science.gov (United States)

    High-throughput sequencing of reduced representation genomic libraries has ushered in an era of genotyping-by-sequencing (GBS), where genome-wide genotype data can be obtained for nearly any species. However, there remains a need for imputation-free GBS methods for genotyping large samples taken fr...

  7. Evaluation of the Genotype MTBDR Assay for Rapid Detection of Rifampin and Isoniazid Resistance in Mycobacterium tuberculosis Isolates

    OpenAIRE

    2006-01-01

    A novel PCR-based reverse hybridization method Genotype MTBDR assay (Hain Lifescience GmbH, Nehren, Germany) was evaluated for rapid detection of rifampin (RIF) and isoniazid (INH) resistance in Turkish Mycobacterium tuberculosis isolates. The Genotype MTBDR assay is designed to detect mutations within the 81-bp hotspot region of rpoB and mutations at katG codon 315. A total of 41 RIF-resistant M. tuberculosis isolates with rpoB mutations that were previously tested by the INNO-LiPA Rif.TB ki...

  8. A first insight into the genotypic diversity of Mycobacterium tuberculosis from Rwanda

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    Gafirita James

    2012-11-01

    Full Text Available Abstract Background Mycobacterium tuberculosis complex (MTC is the causative agent of tuberculosis (TB. Globally, increasing evidence shows that in M. tuberculosis, transmission varies from strain to strain and that different strains exhibit a range of geographical and host specificities, pathogenicity, and drug susceptibility. Therefore rapid and accurate differentiation of the members of MTC is critical in guiding treatment and public health decisions. We carried out a study at different health units and the National Reference Laboratory in Rwanda identify Mycobacterium tuberculosis complex species prevalent in TB patients in Rwanda. We further characterized the isolates using spoligotyping in order to gain an insight into the strain diversity of drug resistant and susceptible isolates of M. tuberculosis in this setting. Methods A total of 151 isolates from culture positive sputum samples were harvested, heat killed at 80°C for two hours, and then shipped to Makerere University College of Health Sciences, Uganda, for speciation and typing. Species identification was achieved by regions of difference (RD analysis, while Spoligotyping was done to identify strain types. Results Region of difference analysis identified all the 151 isolates as M. tuberculosis. Spoligotyping revealed predominance of the T2 family (58.3%, 88/151, with SIT 52 being the most prevalent strain (31.8%, 48/151. Among the 151 isolates, 64 (42.4% were multidrug resistant (MDR with 3 cases on mono-resistance. Of 94 retreatment cases, 48 (51.1% were MDR and of 46 newly presenting cases 14 (30.4% were MDR. There was a significant difference (p=0.01 in anti-TB drug resistance between new and retreatment cases in the sample. However, there was no significant relationship between HIV serostatus and the two major strain types SIT 52 (p =0.15and SIT 152 (p = 0.41. Conclusion Mycobacterium tuberculosis is the most prevalent species of Mycobacterium tuberculosis complex in Rwanda

  9. Does fingerprinting truly represent the diversity of wine yeasts? A case study with interdelta genotyping of Saccharomyces cerevisiae strains.

    Science.gov (United States)

    Pfliegler, W P; Sipiczki, M

    2016-12-01

    Simple and efficient genotyping methods are widely used to assess the diversity of a large number of microbial strains, e.g. wine yeasts isolated from a specific geographical area or a vintage. Such methods are often also the first to be applied, to decrease the number of strains deemed interesting for a more time-consuming physiological characterization. Here, we aimed to use a physiologically characterized strain collection of 69 Saccharomyces cerevisiae strains from Hungarian wine regions to determine whether geographical origin or physiological similarity can be recovered by clustering the strains with one or two simultaneously used variations of interdelta genotyping. Our results indicate that although a detailed clustering with high resolution can be achieved with this method, the clustering of strains is largely contrasting when different primer sets are used and it does not recover geographical or physiological groups. Genotyping is routinely used for assessing the diversity of a large number of isolates/strains of a single species, e.g. a collection of wine yeasts. We tested the efficiency of interdelta genotyping on a collection of Saccharomyces wine yeasts from four wine regions of Hungary that was previously characterized physiologically. Interdelta fingerprinting recovered neither physiological nor geographical similarities, and in addition, the two different primer pairs widely used for this method showed conflicting and barely comparable results. Thus, this method does not necessarily represent the true diversity of a strain collection, but detailed clustering may be achieved by the combined use of primer sets. © 2016 The Society for Applied Microbiology.

  10. Plasmodium falciparum genotypes diversity in symptomatic malaria of children living in an urban and a rural setting in Burkina Faso

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    Konaté Amadou T

    2009-06-01

    Full Text Available Abstract Background The clinical presentation of malaria, considered as the result of a complex interaction between parasite and human genetics, is described to be different between rural and urban areas. The analysis of the Plasmodium falciparum genetic diversity in children with uncomplicated malaria, living in these two different areas, may help to understand the effect of urbanization on the distribution of P. falciparum genotypes. Methods Isolates collected from 75 and 89 children with uncomplicated malaria infection living in a rural and an urban area of Burkina Faso, respectively, were analysed by a nested PCR amplification of msp1 and msp2 genes to compare P. falciparum diversity. Results The K1 allelic family was widespread in children living in the two sites, compared to other msp1 allelic families (frequency >90%. The MAD 20 allelic family of msp1 was more prevalent (p = 0.0001 in the urban (85.3% than the rural area (63.2%. In the urban area, the 3D7 alleles of msp2 were more prevalent compared to FC27 alleles, with a high frequency for the 3D7 300bp allele (>30%. The multiplicity of infection was in the range of one to six in the urban area and of one to seven in the rural area. There was no difference in the frequency of multiple infections (p = 0.6: 96.0% (95% C.I: 91.6–100 in urban versus 93.1% (95%C.I: 87.6–98.6 in rural areas. The complexity of infection increased with age [p = 0.04 (rural area, p = 0.06 (urban area]. Conclusion Urban-rural area differences were observed in some allelic families (MAD20, FC27, 3D7, suggesting a probable impact of urbanization on genetic variability of P. falciparum. This should be taken into account in the implementation of malaria control measures.

  11. Diversity of Enterococcus faecalis Genotypes from Multiple Oral Sites Associated with Endodontic Failure Using Repetitive Sequence-based Polymerase Chain Reaction and Arbitrarily Primed Polymerase Chain Reaction.

    Science.gov (United States)

    Delboni, Maraísa G; Gomes, Brenda P F A; Francisco, Priscila A; Teixeira, Fabrício B; Drake, David

    2017-03-01

    The aim of this study was to evaluate the diversity and similarity of Enterococcus faecalis genotype isolates from multiple oral sites using repetitive sequence-based polymerase chain reaction and arbitrarily primed polymerase chain reaction (AP-PCR). Forty-two endodontically treated teeth with apical periodontitis were selected. A total of 126 microbial samples were collected from 3 different sites (saliva, pulp chamber, and root canals, all n = 42) during the nonsurgical retreatment procedures. After growth on m-Enterococcus agar, the colonies were isolated, characterized as gram-positive catalase negative cocci, and identified using an API 20 Strep kit (bioMérieux, Marcy-l'Etoile, France). Seventy-four colonies from 10 patients were confirmed as E. faecalis by polymerase chain reaction (16S ribosomal RNA). Repetitive sequence-based polymerase chain reactions using ERIC and AP-PCR using RW3A primers were performed in all 74 colonies. Fingerprints were analyzed and separated into genotypic groups based on the Dice coefficient percentage of similarity (82% or greater) as determined by ERIC reproducibility assays involving E. faecalis controls. Seven different E. faecalis genotypes (GTs) (GT1 = 27%, GT2 = 17.6%, GT3 = 1.3%, GT4 = 18.9%, GT5 = 9.5%, GT6 = 14.9%, and GT7 = 10.8%) were observed in different subjects and oral sites associated with endodontic failure. Remarkably, in 4 of 5 patients, the same GTs present in the infected root canals were also isolated from either the pulp chamber or the saliva samples. In particular, GT6 was detected in all 3 oral sites of patient 37. E. faecalis GTs isolated from saliva, the pulp chamber, and the root canal were similar using the Rep-PCR and AP-PCR methods. These findings suggest that coronal microleakage is a conceivable cause of endodontic failure. Copyright © 2016 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  12. Detecting small-scale genotype-environment interactions in apomictic dandelion (Taraxacum officinale) populations.

    Science.gov (United States)

    McLeod, K A; Scascitelli, M; Vellend, M

    2012-08-01

    Studies of genotype × environment interactions (G × E) and local adaptation provide critical tests of natural selection's ability to counter opposing forces such as gene flow. Such studies may be greatly facilitated in asexual species, given the possibility for experimental replication at the level of true genotypes (rather than populations) and the possibility of using molecular markers to assess genotype-environment associations in the field (neither of which is possible for most sexual species). Here, we tested for G × E in asexual dandelions (Taraxacum officinale) by subjecting six genotypes to experimental drought, mown and benign (control) conditions and subsequently using microsatellites to assess genotype-environment associations in the field. We found strong G × E, with genotypes that performed poorly under benign conditions showing the highest performance under stressful conditions (drought or mown). Our six focal genotypes comprise > 80% of plants in local populations. The most common genotype in the field showed its highest relative performance under mown conditions (the most common habitat in our study area), and almost all plants of this genotype in the field were found growing in mowed lawns. Genotypes performing best under benign experimental conditions were found most frequently in unmown conditions in the field. These results are strongly indicative of local adaptation at a very small scale, with unmown microsites of only a few square metres typically embedded within larger mown lawns. By studying an asexual species, we were able to map genotypes with known ecological characteristics to environments with high spatial precision.

  13. Detection of parasitizing coccidia and determination of host crane species, sex and genotype by faecal DNA analysis.

    Science.gov (United States)

    Honma, H; Suyama, Y; Nakai, Y

    2011-11-01

    In Japan, the three main crane species are the endangered red-crowned crane (Grus japonensis) inhabiting Hokkaido, the northernmost island of Japan; the vulnerable hooded crane (Grus monacha); and the vulnerable white-naped crane (Grus vipio). Both the hooded and white-naped cranes migrate in winter to Izumi in Kyushu, the southern island of Japan. In this study, we investigated the cranes and their coccidian parasites, through a targeted molecular approach using faecal DNA to develop a noninvasive method for infectious disease research. To determine the origin of noninvasively collected faecal samples, host species were identified by sequencing a region of approximately 470 bp of the mitochondrial 16S ribosomal RNA gene in the faecal DNA. Furthermore, to avoid sample redundancy, individual determination was performed by fragment analysis using microsatellite and sex-linked markers. For microsatellite genotyping, previously reported markers and markers isolated in this study were examined, and seven loci for red-crowned cranes, eight for hooded cranes and six for white-naped cranes displayed polymorphisms. A low error rate was demonstrated by comparing microsatellite data generated from faecal DNA samples with that generated from feather DNA samples, indicating a high reliability. Polymerase chain reaction-based capillary electrophoresis (PCR-CE), employing genetic markers in the second internal transcribed spacer (ITS2) of nuclear ribosomal DNA, was employed to detect crane coccidia. The sensitivity of detection of PCR-CE using faecal DNA was inferior to that with traditional microscopy; however, our results suggest that PCR-CE can depict crane coccidia diversity with higher resolution and it is a useful tool to characterize community composition of coccidia in detail.

  14. Development of a novel genus-specific real-time PCR assay for detection and differentiation of Bartonella species and genotypes.

    Science.gov (United States)

    Diaz, Maureen H; Bai, Ying; Malania, Lile; Winchell, Jonas M; Kosoy, Michael Y

    2012-05-01

    The genus Bartonella includes numerous species with varied host associations, including several that infect humans. Development of a molecular diagnostic method capable of detecting the diverse repertoire of Bartonella species while maintaining genus specificity has been a challenge. We developed a novel real-time PCR assay targeting a 301-bp region of the ssrA gene of Bartonella and demonstrated specific amplification in over 30 Bartonella species, subspecies, and strains. Subsequent analysis of ssrA sequences was sufficient to discriminate Bartonella species and provided phylogenetic data consistent with that of gltA, a commonly used gene for differentiating Bartonella genotypes. Using this assay, we identified Bartonella DNA in 29% and 47% of blood specimens from elk in Wyoming and cattle in the Republic of Georgia, respectively. Sequence analysis of a subset of genotypes from elk specimens revealed a cluster most closely related to Bartonella capreoli, and genotypes from cattle were identified as Bartonella bovis, both Bartonella species commonly found in wild and domestic ruminants. Considering the widespread geographic distribution and infectivity potential to a variety of hosts, this assay may be an effective diagnostic method for identification of Bartonella infections in humans and have utility in Bartonella surveillance studies.

  15. Genotypic diversity and virulence markers of Yersinia enterocolitica biotype 1A strains isolated from clinical and non-clinical origins.

    Science.gov (United States)

    Campioni, Fábio; Falcão, Juliana P

    2014-03-01

    Yersinia enterocolitica biotype 1A (B1A) strains are considered as non-pathogenic; however, some reports have identified some strains as the causal agents of infection. In South America, few studies molecularly characterized the strains of this biotype. This work typed 51 B1A strains isolated from clinical and non-clinical sources from Brazil and Chile by Enterobacterial Repetitive Intergenic Consensus-PCR (ERIC-PCR) to elucidate their genotypic diversity, and verify the distribution of 11 virulence markers by PCR. The strains were divided into two groups, ERIC-A and ERIC-B, clustered independently of their clinical or non-clinical origin. No differences were observed in the frequencies of the virulence markers between clinical and non-clinical strains. However, the genes ystB, hreP and myfA occurred exclusively in the strains of the group ERIC-A. Some clinical and non-clinical strains were clustered in the same genetic group and presented the same number of virulence markers, which might suggest the role of the environment and food as a potential source of infection for humans and animals. The results corroborate with the hypothesis that B1A strains are divided into two main clusters that differ in the frequency of some virulence markers, a fact observed for the first time in South American strains.

  16. Expression of Hepatitis B virus surface antigen (HBsAg from genotypes A, D and F and influence of amino acid variations related or not to genotypes on HBsAg detection

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    Natalia M. Araujo

    Full Text Available The impact of hepatitis B virus (HBV genotypes on the sensitivity of surface antigen (HBsAg detection assays has been poorly investigated. Here, plasmids carrying consensus or variant coding sequences for HBV surface proteins from genotypes A, D and F, were constructed. HBsAg levels were evaluated in medium and extracts of transfected CHO cells by a commercial polyclonal-based assay. We show that HBsAg detection values of consensus forms from genotypes D and F were, respectively, 37% and 30% lower than those obtained by genotype A. However, the presence of two single variations, T143M in genotype A, and T125M in genotype D, produced a decrease of 44% and an increase of 34%, respectively, on HBsAg mean values in comparison with their consensus forms. In conclusion, HBsAg detection levels varied among HBV genotypes. However, unique amino acid substitutions not linked to genotypes, such as T125M and T143M described here, should have more implications in HBV immunological diagnostics than the set of variations characteristic of each HBV genotype.

  17. Expression of Hepatitis B virus surface antigen (HBsAg from genotypes A, D and F and influence of amino acid variations related or not to genotypes on HBsAg detection

    Directory of Open Access Journals (Sweden)

    Natalia M. Araujo

    2009-08-01

    Full Text Available The impact of hepatitis B virus (HBV genotypes on the sensitivity of surface antigen (HBsAg detection assays has been poorly investigated. Here, plasmids carrying consensus or variant coding sequences for HBV surface proteins from genotypes A, D and F, were constructed. HBsAg levels were evaluated in medium and extracts of transfected CHO cells by a commercial polyclonal-based assay. We show that HBsAg detection values of consensus forms from genotypes D and F were, respectively, 37% and 30% lower than those obtained by genotype A. However, the presence of two single variations, T143M in genotype A, and T125M in genotype D, produced a decrease of 44% and an increase of 34%, respectively, on HBsAg mean values in comparison with their consensus forms. In conclusion, HBsAg detection levels varied among HBV genotypes. However, unique amino acid substitutions not linked to genotypes, such as T125M and T143M described here, should have more implications in HBV immunological diagnostics than the set of variations characteristic of each HBV genotype.

  18. Genotyping hepatitis C viruses from Southeast Asia by a novel line probe assay that simultaneously detects core and 5′ untranslated regions

    NARCIS (Netherlands)

    S. Noppornpanth (Suwanna); B.L. Haagmans (Bart); E. Sablon (Erwin); K. de Nys (Kathy); T.X. Lien (Truong Xuang); J. Brouwer (Jan); M. van Brussel (Marianne); S.L. Smits (Saskia); Y. Poovorawan (Yong); A.D.M.E. Osterhaus (Albert)

    2006-01-01

    textabstractHepatitis C viruses (HCVs) display a high level of sequence diversity and are currently divided into six genotypes. A line probe assay (LiPA), which targets the 5′ untranslated region (5′UTR) of the HCV genome, is widely used for genotyping. However, this assay cannot distinguish many ge

  19. Porphyrin-based sensor nanoarchitectonics in diverse physical detection modes.

    Science.gov (United States)

    Ishihara, Shinsuke; Labuta, Jan; Van Rossom, Wim; Ishikawa, Daisuke; Minami, Kosuke; Hill, Jonathan P; Ariga, Katsuhiko

    2014-06-07

    Porphyrins and related families of molecules are important organic modules as has been reflected in the award of the Nobel Prizes in Chemistry in 1915, 1930, 1961, 1962, 1965, and 1988 for work on porphyrin-related biological functionalities. The porphyrin core can be synthetically modified by introduction of various functional groups and other elements, allowing creation of numerous types of porphyrin derivatives. This feature makes porphyrins extremely useful molecules especially in combination with their other interesting photonic, electronic and magnetic properties, which in turn is reflected in their diverse signal input-output functionalities based on interactions with other molecules and external stimuli. Therefore, porphyrins and related macrocycles play a preeminent role in sensing applications involving chromophores. In this review, we discuss recent developments in porphyrin-based sensing applications in conjunction with the new advanced concept of nanoarchitectonics, which creates functional nanostructures based on a profound understanding of mutual interactions between the individual nanostructures and their arbitrary arrangements. Following a brief explanation of the basics of porphyrin chemistry and physics, recent examples in the corresponding fields are discussed according to a classification based on physical modes of detection including optical detection (absorption/photoluminescence spectroscopy and energy and electron transfer processes), other spectral modes (circular dichroism, plasmon and nuclear magnetic resonance), electronic and electrochemical modes, and other sensing modes.

  20. Antibody Detection, Isolation, Genotyping, and Virulence of Toxoplasma gondii in Captive Felids from China

    Directory of Open Access Journals (Sweden)

    Yu-Rong Yang

    2017-07-01

    Full Text Available The felids are the only definitive hosts of Toxoplasma gondii, which could excrete oocysts into the environment and provide an infection source for toxoplasmosis in various warm-blooded animal species, particularly the captive felids that live close to human communities. The infection rate of the captive felids is a perfect standard in detecting the presence of Toxoplasma gondii oocysts in the environment. In this study, sera or tissue samples from zoo (1 young tiger, 2 adult tigers, 6 young lions, farm (10 masked palm civets, and pet hospital (28 cats from Henan Province (China were collected. The sera (n = 47 were tested for immunoglobulin G (IgG antibodies against T. gondii by using modified agglutination test (MAT, whereas the hearts tissue (n = 40 were bioassayed in mice to isolate T. gondii strains. The genotype was distinguished by using PCR-RFLP of 10 loci (SAG1, SAG2, SAG3, GRA6, BTUB, L358, c22-8, PK1, c29-2, and Apico. The detection rate for the T. gondii antibody in captive felids was 21.3% (10/47. One viable T. gondii strain (TgCatCHn4 was obtained from a cat heart tissue, and its genotype was ToxoDB#9. The oocysts of ToxoDB#9 were collected from a T. gondii-free cat. The virulence of TgCatCHn4 was low and no cysts were detected in the brain of mice at 60 days post-inoculation. The finding of the present study suggested a widespread exposure of T. gondii for felids in Henan Province of central China, particularly those from the zoological gardens and homes. ToxoDB#9 was the predominant strain in China. Preventive measures against T. gondii oocyst contamination of various components of the environment should thus be implemented, including providing pre-frozen meat, well-cooked cat food, cleaned fruits and vegetables, monitoring birds and rodents, inactive T. gondii oocysts in felids feces, and proper hygiene.

  1. Genotypic and phenotypic detection of macrolide and lincosamide resistance in Streptococcus uberis.

    Science.gov (United States)

    Schmitt-Van de Leemput, E; Zadoks, R N

    2007-11-01

    Streptococcus uberis isolates (n = 55) were obtained from milk samples of cases of mild clinical mastitis in 55 dairy cows from 35 herds serviced by one veterinary practice in Mayenne, France. Isolates were tested for macrolide and lincosamide resistance by using phenotypic and genotypic methods. Erythromycin resistance was detected in 12 of the 55 (22%) isolates based on agar disc diffusion testing and MIC measurements, and was encoded by ermB. This gene also conferred phenotypic resistance to pirlimycin based on MIC measurements, but the D-test was needed for detection of the resistance phenotype in the agar disc diffusion test. Isolates with ermB were also highly resistant to the macrolide antibiotic spiramycin. Seventeen of the 55 isolates (31%) were classified as resistant to spiramcyin only and as having intermediate susceptibility to spiramycin based on agar disc diffusion testing and MIC measurements, respectively. The genetic mechanism behind this phenotype and its clinical relevance are unknown. The efflux pump gene mefA was not detected in any of the 55 isolates in this study. Pirlimycin resistance without macrolide resistance was encoded by the lincosamide resistance gene linB in 4 isolates. Based on current guidelines, some linB-positive isolates would be classified as susceptible by using phenotypic tests, and alternative values for the interpretation of the agar disc diffusion test are suggested. We conclude that the agar disc diffusion test is a useful indicator for macrolide and lincosamide resistance of Strep. uberis in veterinary practice, provided that the D-test is used for detection of pirlimycin resistance.

  2. H. pylori clinical isolates have diverse babAB genotype distributions over different topographic sites of stomach with correlation to clinical disease outcomes

    Directory of Open Access Journals (Sweden)

    Sheu Shew-Meei

    2012-05-01

    Full Text Available Abstract Background Intragenomic recombination between babA and babB mediates antigenic variations and may help H. pylori colonization. This study determined whether variable genotypes of babA and babB correlate to different clinical disease outcomes, and can distribute over the different gastric niches. Results This study enrolled 92 clinical strains (45 from peptic ulcer, 27 from gastritis, and 20 from gastric cancer to detect whether the babA and babB are at locus A or B by PCR reactions using the primers designed from the upstream and variable region of the babA and babB genes. Four genotypes of babA and babB (A B, AB B, A AB, AB AB were found. The distribution of the 4 genotypes in 92 clinical strains was significantly different among patients with different gastric diseases (p vs. 9.7%, p p p > 0.05. Besides, the study enrolled 19 patients to verify whether variable genotypes of babAB existed in the different gastric niches. Among the patients infected with more than one babAB genotypes over antrum and corpus, there were higher rate of genotypes as A B or AB AB in isolates from antrum than in those from corpus (75.0 % vs. 16.7%, p  Conclusions The H. pylori isolate with the AB AB genotype correlates with an increased gastric cancer risk, and colonize in an antrum predominant manner.

  3. Phenotypic and genotypic characterization of vancomycin-resistant Enterococcus faecium clinical isolates from two hospitals in Mexico: First detection of VanB phenotype-vanA genotype.

    Science.gov (United States)

    Bocanegra-Ibarias, Paola; Flores-Treviño, Samantha; Camacho-Ortiz, Adrián; Morfin-Otero, Rayo; Villarreal-Treviño, Licet; Llaca-Díaz, Jorge; Martínez-Landeros, Erik Alan; Rodríguez-Noriega, Eduardo; Calzada-Güereca, Andrés; Maldonado-Garza, Héctor Jesús; Garza-González, Elvira

    2016-01-01

    Enterococcus faecium has emerged as a multidrug-resistant nosocomial pathogen involved in outbreaks worldwide. Our aim was to determine the antimicrobial susceptibility, biofilm production, and clonal relatedness of vancomycin-resistant E. faecium (VREF) clinical isolates from two hospitals in Mexico. Consecutive clinical isolates (n=56) were collected in two tertiary care hospitals in Mexico from 2011 to 2014. VREF isolates were characterized by phenotypic and molecular methods including pulsed-field gel electrophoresis (PFGE). VREF isolates were highly resistant to vancomycin, erythromycin, norfloxacin, high-level streptomycin, and teicoplanin, and showed lower resistance to tetracycline, nitrofurantoin and quinupristin-dalfopristin. None of the isolates were resistant to linezolid. The vanA gene was detected in all isolates. Two VanB phenotype-vanA genotype isolates, highly resistant to vancomycin and susceptible to teicoplanin, were detected. Furthermore, 17.9% of the isolates were classified as biofilm producers, and the espfm gene was found in 98.2% of the isolates. A total of 37 distinct PFGE patterns and 6 clones (25% of the isolates as clone A, 5.4% as clone B, and 3.6% each as clone C, D, E, and F) were detected. Clone A was detected in 5 different wards of the same hospital during 14 months of surveillance. The high resistance to most antimicrobial agents and the moderate cross-transmission of VREF detected accentuates the need for continuous surveillance of E. faecium in the hospital setting. This is also the first reported incidence of the E. faecium VanB phenotype-vanA genotype in the Americas. Copyright © 2015 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

  4. Postnatal and non-invasive prenatal detection of β-thalassemia mutations based on Taqman genotyping assays

    Science.gov (United States)

    Breveglieri, Giulia; Travan, Anna; D’Aversa, Elisabetta; Cosenza, Lucia Carmela; Pellegatti, Patrizia; Guerra, Giovanni; Gambari, Roberto

    2017-01-01

    The β-thalassemias are genetic disorder caused by more than 200 mutations in the β-globin gene, resulting in a total (β0) or partial (β+) deficit of the globin chain synthesis. The most frequent Mediterranean mutations for β-thalassemia are: β039, β+IVSI-110, β+IVSI-6 and β0IVSI-1. Several molecular techniques for the detection of point mutations have been developed based on the amplification of the DNA target by polymerase chain reaction (PCR), but they could be labor-intensive and technically demanding. On the contrary, TaqMan® genotyping assays are a simple, sensitive and versatile method suitable for the single nucleotide polymorphism (SNP) genotyping affecting the human β-globin gene. Four TaqMan® genotyping assays for the most common β-thalassemia mutations present in the Mediterranean area were designed and validated for the genotype characterization of genomic DNA extracted from 94 subjects comprising 25 healthy donors, 33 healthy carriers and 36 β-thalassemia patients. In addition, 15 specimens at late gestation (21–39 gestational weeks) and 11 at early gestation (5–18 gestational weeks) were collected from pregnant women, and circulating cell-free fetal DNAs were extracted and analyzed with these four genotyping assays. We developed four simple, inexpensive and versatile genotyping assays for the postnatal and prenatal identification of the thalassemia mutations β039, β+IVSI-110, β+IVSI-6, β0IVSI-1. These genotyping assays are able to detect paternally inherited point mutations in the fetus and could be efficiently employed for non-invasive prenatal diagnosis of β-globin gene mutations, starting from the 9th gestational week. PMID:28235086

  5. A novel strategy for human papillomavirus detection and genotyping with SybrGreen and molecular beacon polymerase chain reaction

    NARCIS (Netherlands)

    Szuhai, K; Sandhaus, E; Kolkman-Uljee, SM; Lemaitre, M; Truffert, JC; Dirks, RW; Tanke, HJ; Fleuren, GJ; Schuuring, E; Raap, AK

    2001-01-01

    Human papillomaviruses (HPVs) play an important role in the pathogenesis of cervical cancer. For identification of the large number of different HPV types found in (pre)malignant lesions, a robust methodology is needed that combines general HPV detection with HPV genotyping. We have developed for

  6. A novel strategy for human papillomavirus detection and genotyping with SybrGreen and molecular beacon polymerase chain reaction

    NARCIS (Netherlands)

    Szuhai, K; Sandhaus, E; Kolkman-Uljee, SM; Lemaitre, M; Truffert, JC; Dirks, RW; Tanke, HJ; Fleuren, GJ; Schuuring, E; Raap, AK

    2001-01-01

    Human papillomaviruses (HPVs) play an important role in the pathogenesis of cervical cancer. For identification of the large number of different HPV types found in (pre)malignant lesions, a robust methodology is needed that combines general HPV detection with HPV genotyping. We have developed for fo

  7. Birth of correctly genotyped calves after multiplex marker detection from bovine embryo microblade biopsies.

    Science.gov (United States)

    Peippo, Jaana; Viitala, Sirja; Virta, Jouni; Räty, Mervi; Tammiranta, Niina; Lamminen, Terttu; Aro, Johanna; Myllymäki, Hannu; Vilkki, Johanna

    2007-11-01

    We report a method for multiplex genotyping of bovine embryo microblade biopsies. We have tested the reliability of the method and the viability of the embryos in vitro and in vivo. Two polymorphic gene markers (GHR F279Y and PRLR S18N) associated with milk production traits and one marker for sex diagnosis (ZFX/ZFY) were genotyped simultaneously with a method that combines nested PCR and allelic discrimination. To test the accuracy of genotyping, in the first experiment the genotypes of 134 biopsies from in vitro produced embryos were compared to genotypes determined from the corresponding embryos after biopsy. The method proved to be highly accurate as only in three cases (two for PRLR S18N and one for GHR F279Y) out of 395 genotypes the genotype was in disagreement between the two samples. The viability of similarly biopsied embryos was tested in parallel: after 24-hr culture 94.6% of embryos recovered in vitro. In the second experiment, a total of 150 in vivo-produced embryos were biopsied on Day 7 and genotyped. After the genotyping results were obtained on Day 8, female embryos were selected for transfer. From a total of 57 selected embryos 43 were transferred individually and 14 as pairs. After single embryo transfers, 19 recipients became pregnant and after embryo transfers in pairs one became pregnant. The success of genotyping was tested with the genotypes of donors and bulls and also from the hair samples of born calves. All calves were females and of the same genotypes determined from the biopsy.

  8. Molecular Detection and Genotypic Characterization of Toxoplasma gondii in Paraffin-Embedded Fetoplacental Tissues of Women with Recurrent Spontaneous Abortion

    Science.gov (United States)

    Abdoli, Amir; Dalimi, Abdolhossein; Soltanghoraee, Haleh; Ghaffarifar, Fatemeh

    2017-01-01

    Background Congenital toxoplasmosis is an important cause of spontaneous abortion worldwide. However, there is limited information on detection and genotypic characterization of Toxoplasma gondii (T. gondii) in women with recurrent spontaneous abortion (RSA). The aim of this study is the molecular detection and genotypic characterization of T. gondii in formalin-fixed, paraffin-embedded fetoplacental tissues (FFPTs) of women with RSA that have referred to the Avicenna Research Institute in Tehran, Iran. Materials and Methods This experimental research was undertaken on 210 FFPTs of women with RSA. The information of the patients was collected from the archives of Avicenna Research Institute in Tehran, Iran. After DNA extraction, the presence of T. gondii was examined by nested polymerase chain reaction targeting the GRA6 gene. Genotyping was performed on positive samples using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) that targeted the GRA6 and SAG3 genes. Sequencing was conducted on two GRA6 positive samples. Results T. gondii DNA was detected in 3.8% (8/210) of the samples. Genotyping showed that all positive samples belonged to type III of the T. gondii genotype. Sequencing two genomic DNAs of the GRA6 gene revealed 99% similarity with each other and 99-100% similarity with T. gondii sequences deposited in GenBank. There were six patients with histories of more than three abortions; one patient had a healthy girl and another patient had two previous abortions. Abortions occurred in the first trimester of pregnancy in seven patients and in the second trimester of pregnancy in one patient. Conclusion The results of this study have indicated that genotype III is the predominant type of T. gondii in women with RSA in Tehran, Iran. Also, our findings suggest that toxoplasmosis may play a role in the pathogenesis of RSA. However, further studies are needed to elucidate a clear relationship between T. gondii infection and RSA. PMID

  9. Molecular Detection and Genotypic Characterization of Toxoplasma gondii in Paraffin-Embedded Fetoplacental Tissues of Women with Recurrent Spontaneous Abortion

    Directory of Open Access Journals (Sweden)

    Amir Abdoli

    2016-11-01

    Full Text Available Background: Congenital toxoplasmosis is an important cause of spontaneous abortion worldwide. However, there is limited information on detection and genotypic characterization of Toxoplasma gondii (T. gondii in women with recurrent spontaneous abortion (RSA. The aim of this study is the molecular detection and genotypic characterization of T. gondii in formalin-fixed, paraffin-embedded fetoplacental tissues (FFPTs of women with RSA that have referred to the Avicenna Research Institute in Tehran, Iran. Materials and Methods: This experimental research was undertaken on 210 FFPTs of women with RSA. The information of the patients was collected from the archives of Avicenna Research Institute in Tehran, Iran. After DNA extraction, the presence of T. gondii was examined by nested polymerase chain reaction targeting the GRA6 gene. Genotyping was performed on positive samples using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP that targeted the GRA6 and SAG3 genes. Sequencing was conducted on two GRA6 positive samples. Results: T. gondii DNA was detected in 3.8% (8/210 of the samples. Genotyping showed that all positive samples belonged to type III of the T. gondii genotype. Sequencing two genomic DNAs of the GRA6 gene revealed 99% similarity with each other and 99-100% similarity with T. gondii sequences deposited in GenBank. There were six patients with histories of more than three abortions; one patient had a healthy girl and another patient had two previous abortions. Abortions occurred in the first trimester of pregnancy in seven patients and in the second trimester of pregnancy in one patient. Conclusion: The results of this study have indicated that genotype III is the predominant type of T. gondii in women with RSA in Tehran, Iran. Also, our findings suggest that toxoplasmosis may play a role in the pathogenesis of RSA. However, further studies are needed to elucidate a clear relationship between T. gondii

  10. Clonal diversity and population genetic structure of arbuscular mycorrhizal fungi (Glomus spp.) studied by multilocus genotyping of single spores

    DEFF Research Database (Denmark)

    Holtgrewe-Stukenbrock, Eva; Rosendahl, Søren

    2005-01-01

    characterized by SSCP (single stranded conformation polymorphism) and sequencing.   All spore genotypes were unique suggesting that no recombination was taking place in the populations. There were no overall differences in the distribution of genotypes in the two fields and identical genotypes could be sampled......A nested multiplex PCR (polymerase chain reaction) approach was used for multilocus genotyping of arbuscular mycorrhizal fungal populations. This method allowed us to amplify multiple loci from Glomus single spores in a single PCR amplification. Variable introns in the two protein coding genes Gm...

  11. Phenotypic and genotypic characterization of Stenotrophomonas maltophilia isolates from patients with cystic fibrosis: genome diversity, biofilm formation, and virulence.

    Science.gov (United States)

    Pompilio, Arianna; Pomponio, Stefano; Crocetta, Valentina; Gherardi, Giovanni; Verginelli, Fabio; Fiscarelli, Ersilia; Dicuonzo, Giordano; Savini, Vincenzo; D'Antonio, Domenico; Di Bonaventura, Giovanni

    2011-07-05

    Stenotrophomonas maltophilia is emerging as one of the most frequently found bacteria in cystic fibrosis (CF) patients. In the present study, phenotypic and genotypic traits of a set of 98 isolates of S. maltophilia obtained from clinical (CF and non-CF patients) and environmental sources were comparatively evaluated. S. maltophilia exhibited a high level of genomic diversity in both CF and non-CF group, thus possibly allowing this bacterium to expand its pathogenic potentials. Strains sharing the same pulsotype infected different patients, thus likely indicating the occurrence of clonal spread or acquisition by a common source. CF isolates differed greatly in some phenotypic traits among each other and also when compared with non-CF isolates, demonstrating increased mean generation time and susceptibility to oxidative stress, but reduced ability in forming biofilm. Furthermore, in CF isolates flagella- and type IV pili-based motilities were critical for biofilm development, although not required for its initiation. Sequential isogenic strains isolated from the same CF patient displayed heterogeneity in biofilm and other phenotypic traits during the course of chronic infection. CF and non-CF isolates showed comparable virulence in a mouse model of lung infection. Overall, the phenotypic differences observed between CF and non-CF isolates may imply different selective conditions and persistence (adaptation) mechanisms in a hostile and heterogeneous environment such as CF lung. Molecular elucidation of these mechanisms will be essential to better understand the selective adaptation in CF airways in order to design improved strategies useful to counteract and eradicate S. maltophilia infection.

  12. A novel method for detecting uniparental disomy from trio genotypes identifies a significant excess in children with developmental disorders.

    Science.gov (United States)

    King, Daniel A; Fitzgerald, Tomas W; Miller, Ray; Canham, Natalie; Clayton-Smith, Jill; Johnson, Diana; Mansour, Sahar; Stewart, Fiona; Vasudevan, Pradeep; Hurles, Matthew E

    2014-04-01

    Exome sequencing of parent-offspring trios is a popular strategy for identifying causative genetic variants in children with rare diseases. This method owes its strength to the leveraging of inheritance information, which facilitates de novo variant calling, inference of compound heterozygosity, and the identification of inheritance anomalies. Uniparental disomy describes the inheritance of a homologous chromosome pair from only one parent. This aberration is important to detect in genetic disease studies because it can result in imprinting disorders and recessive diseases. We have developed a software tool to detect uniparental disomy from child-mother-father genotype data that uses a binomial test to identify chromosomes with a significant burden of uniparentally inherited genotypes. This tool is the first to read VCF-formatted genotypes, to perform integrated copy number filtering, and to use a statistical test inherently robust for use in platforms of varying genotyping density and noise characteristics. Simulations demonstrated superior accuracy compared with previously developed approaches. We implemented the method on 1057 trios from the Deciphering Developmental Disorders project, a trio-based rare disease study, and detected six validated events, a significant enrichment compared with the population prevalence of UPD (1 in 3500), suggesting that most of these events are pathogenic. One of these events represents a known imprinting disorder, and exome analyses have identified rare homozygous candidate variants, mainly in the isodisomic regions of UPD chromosomes, which, among other variants, provide targets for further genetic and functional evaluation.

  13. Molecular diversity in irregular or refugee immigrant patients with HBV-genotype-E infection living in the metropolitan area of Naples.

    Science.gov (United States)

    Sagnelli, Caterina; Ciccozzi, Massimo; Coppola, Nicola; Minichini, Carmine; Lo Presti, Alessandra; Starace, Mario; Alessio, Loredana; Macera, Margherita; Cella, Eleonora; Gualdieri, Luciano; Caprio, Nunzio; Pasquale, Giuseppe; Sagnelli, Evangelista

    2016-11-02

    In a recent testing in the metropolitan area of Naples, Italy, on 945 irregular immigrants or refugees, 87 HBsAg chronic carriers were identified, 53 of whom were infected by HBV-genotype E. The aim of the present study was to identify the genetic diversity of HBV-genotype E in these 53 immigrants. The 53 immigrant patients with HBV-genotype-E infection were born in Africa, central or eastern Asia, eastern Europe or Latin America. These patients had been seen for a clinical consultation at one of the four first-level units from January 2012 to 2013. The first dataset contained 53 HBV-S gene isolates plus 128 genotype/subgenotype specific reference sequences downloaded from the National Center for Biotechnology Information. The second dataset, comprising the 53 HBV-S gene isolates, previously classified as HBV-genotype E, was used to perform the time-scaled phylogeny reconstruction using a Bayesian approach. Phylogenetic analysis showed that all 53 HBV-S isolates belonged to HBV-genotype E. Bayes factor analysis showed that the relaxed clock exponential growth model fitted the data significantly better than the other models. The time-scaled Bayesian phylogenetic tree of the second dataset showed that the root of the tree dated back to the year 1990 (95% HPD:1984-2000). Four statistically supported clusters were identified. Cluster A dated back to 2012 (95% HPD:1997-2012); cluster B dated back to 2008 (95% HPD:2001-2015); cluster C to 2006 (95% HPD:1999-2013); cluster D to 2004 (95% HPD:1998-2011). This study disclosed the genetic evolution and phylogenesis in a group of HBV-genotype-E-infected immigrants. J. Med. Virol. © 2016 Wiley Periodicals, Inc.

  14. Diversion of drugs within health care facilities, a multiple-victim crime: patterns of diversion, scope, consequences, detection, and prevention.

    Science.gov (United States)

    Berge, Keith H; Dillon, Kevin R; Sikkink, Karen M; Taylor, Timothy K; Lanier, William L

    2012-07-01

    Mayo Clinic has been involved in an ongoing effort to prevent the diversion of controlled substances from the workplace and to rapidly identify and respond when such diversion is detected. These efforts have found that diversion of controlled substances is not uncommon and can result in substantial risk not only to the individual who is diverting the drugs but also to patients, co-workers, and employers. We believe that all health care facilities should have systems in place to deter controlled substance diversion and to promptly identify diversion and intervene when it is occurring. Such systems are multifaceted and require close cooperation between multiple stakeholders including, but not limited to, departments of pharmacy, safety and security, anesthesiology, nursing, legal counsel, and human resources. Ideally, there should be a broad-based appreciation of the dangers that diversion creates not only for patients but also for all employees of health care facilities, because diversion can occur at any point along a long supply chain. All health care workers must be vigilant for signs of possible diversion and must be aware of how to engage a preexisting group with expertise in investigating possible diversions. In addition, clear policies and procedures should be in place for dealing with such investigations and for managing the many possible outcomes of a confirmed diversion. This article provides an overview of the multiple types of risk that result from drug diversion from health care facilities. Further, we describe a system developed at Mayo Clinic for evaluating episodes of potential drug diversion and for taking action once diversion is confirmed.

  15. HPV detection and genotyping in males from the city of Córdoba, Argentina.

    Science.gov (United States)

    Venezuela, F; Kremer, L E; Kiguen, X; Cuffini, C

    2010-01-01

    A wide range of human papillomavirus (HPV) types can infect the anogenital region of males. Although there is a vast knowledge on HPV infections in women as well as on their association with cervical cancer, the study of HPV infections in males is scarce and controversial. The aim of the present work was to detect and typify HPV infections of the anogenital region in males and analyze the associated risk factors in the population studied. Anogenital samples from 37 patients (30 of whom were HIV carriers) attending the Infectology Service at the Hospital Nacional de Clinicas in C6rdoba, Argentina, were studied. Nine of these patients tested HPV-positive and five out of these nine were found to have mixed infections, being 18 and 61 the most frequent genotypes. There was a significant correlation between the HPV-positive patients and those having an HPV-compatible lesion or AIDS. The present work is the first study in the city of Cordoba which contributes relevant results to the knowledge of HPV infection and to the possible implementation of measures for its prevention.

  16. A new sieving matrix for DNA sequencing, genotyping and mutation detection and high-throughput genotyping with a 96-capillary array system

    Energy Technology Data Exchange (ETDEWEB)

    Gao, David [Iowa State Univ., Ames, IA (United States)

    1999-11-08

    Capillary electrophoresis has been widely accepted as a fast separation technique in DNA analysis. In this dissertation, a new sieving matrix is described for DNA analysis, especially DNA sequencing, genetic typing and mutation detection. A high-throughput 96 capillary array electrophoresis system was also demonstrated for simultaneous multiple genotyping. The authors first evaluated the influence of different capillary coatings on the performance of DNA sequencing. A bare capillary was compared with a DB-wax, an FC-coated and a polyvinylpyrrolidone dynamically coated capillary with PEO as sieving matrix. It was found that covalently-coated capillaries had no better performance than bare capillaries while PVP coating provided excellent and reproducible results. The authors also developed a new sieving Matrix for DNA separation based on commercially available poly(vinylpyrrolidone) (PVP). This sieving matrix has a very low viscosity and an excellent self-coating effect. Successful separations were achieved in uncoated capillaries. Sequencing of M13mp18 showed good resolution up to 500 bases in treated PVP solution. Temperature gradient capillary electrophoresis and PVP solution was applied to mutation detection. A heteroduplex sample and a homoduplex reference were injected during a pair of continuous runs. A temperature gradient of 10 C with a ramp of 0.7 C/min was swept throughout the capillary. Detection was accomplished by laser induced fluorescence detection. Mutation detection was performed by comparing the pattern changes between the homoduplex and the heteroduplex samples. High throughput, high detection rate and easy operation were achieved in this system. They further demonstrated fast and reliable genotyping based on CTTv STR system by multiple-capillary array electrophoresis. The PCR products from individuals were mixed with pooled allelic ladder as an absolute standard and coinjected with a 96-vial tray. Simultaneous one-color laser-induced fluorescence

  17. Seroprevalence, Detection of DNA in Blood and Milk, and Genotyping of Toxoplasma gondii in a Goat Population in Italy

    Directory of Open Access Journals (Sweden)

    Francesca Mancianti

    2013-01-01

    Full Text Available Toxoplasma gondii is the causative agent of a major zoonosis with cosmopolitan distribution and is known to be transmitted mainly by the ingestion of undercooked or raw animal products. Drinking unpasteurized goat’s milk is a risk factor associated with human toxoplasmosis. However, very little is known about the excretion of DNA in goat milk. Aim of the present study was to determine the seroprevalence of T. gondii infection using a modified agglutination test (MAT, to detect T. gondii DNA by nested-PCR (n-PCR in samples of blood and milk from seropositive goats, and to genotype DNA isolates using 11 molecular markers in 127 adult lactating goats from 6 farms in Italy. Positive MAT results were found in 60.6% of goats while 13% of blood and milk samples from seropositive goats were positive to n-PCR. A kappa coefficient of 1 indicated a perfect agreement between blood and milk n-PCR. Genetic characterization of isolates revealed the occurrence of genotype III (, genotype I (, and atypical genotypes with hints for genotype I (. Our results suggest that the risk of excretion of Toxoplasma tachyzoites might frequently occur in milk of seropositive goats testing positive to n-PCR on blood.

  18. Genetic Variation, Heritability, and Diversity Analysis of Upland Rice (Oryza sativa L. Genotypes Based on Quantitative Traits

    Directory of Open Access Journals (Sweden)

    Mst. Tuhina-Khatun

    2015-01-01

    Full Text Available Upland rice is important for sustainable crop production to meet future food demands. The expansion in area of irrigated rice faces limitations due to water scarcity resulting from climate change. Therefore, this research aimed to identify potential genotypes and suitable traits of upland rice germplasm for breeding programmes. Forty-three genotypes were evaluated in a randomised complete block design with three replications. All genotypes exhibited a wide and significant variation for 22 traits. The highest phenotypic and genotypic coefficient of variation was recorded for the number of filled grains/panicle and yields/plant (g. The highest heritability was found for photosynthetic rate, transpiration rate, stomatal conductance, intercellular CO2, and number of filled grains/panicle and yields/plant (g. Cluster analysis based on 22 traits grouped the 43 rice genotypes into five clusters. Cluster II was the largest and consisted of 20 genotypes mostly originating from the Philippines. The first four principle components of 22 traits accounted for about 72% of the total variation and indicated a wide variation among the genotypes. The selected best trait of the number of filled grains/panicle and yields/plant (g, which showed high heritability and high genetic advance, could be used as a selection criterion for hybridisation programmes in the future.

  19. Genetic Variation, Heritability, and Diversity Analysis of Upland Rice (Oryza sativa L.) Genotypes Based on Quantitative Traits.

    Science.gov (United States)

    Tuhina-Khatun, Mst; Hanafi, Mohamed M; Rafii Yusop, Mohd; Wong, M Y; Salleh, Faezah M; Ferdous, Jannatul

    2015-01-01

    Upland rice is important for sustainable crop production to meet future food demands. The expansion in area of irrigated rice faces limitations due to water scarcity resulting from climate change. Therefore, this research aimed to identify potential genotypes and suitable traits of upland rice germplasm for breeding programmes. Forty-three genotypes were evaluated in a randomised complete block design with three replications. All genotypes exhibited a wide and significant variation for 22 traits. The highest phenotypic and genotypic coefficient of variation was recorded for the number of filled grains/panicle and yields/plant (g). The highest heritability was found for photosynthetic rate, transpiration rate, stomatal conductance, intercellular CO₂, and number of filled grains/panicle and yields/plant (g). Cluster analysis based on 22 traits grouped the 43 rice genotypes into five clusters. Cluster II was the largest and consisted of 20 genotypes mostly originating from the Philippines. The first four principle components of 22 traits accounted for about 72% of the total variation and indicated a wide variation among the genotypes. The selected best trait of the number of filled grains/panicle and yields/plant (g), which showed high heritability and high genetic advance, could be used as a selection criterion for hybridisation programmes in the future.

  20. Genetic diversity and comparison of physicochemical and nutritional characteristics of six quinoa (Chenopodium quinoa willd. genotypes cultivated in Chile

    Directory of Open Access Journals (Sweden)

    Margarita Miranda

    2012-12-01

    Full Text Available The present study was focused on the analysis of agronomical, nutritional, physicochemical, and antioxidant properties of six genetically different quinoa (Chenopodium quinoa Willd genotypes cultivated in three distinctive geographical zones of Chile. Ancovinto and Cancosa genotypes from the northern Altiplano (19 ºS, Cáhuil and Faro from the central region (34 ºS, and Regalona and Villarica from the southern region (39 ºS are representative of high genetic differentiation among the pooled samples, in particular between Altiplano and the central-southern groups. A Common-Garden Assay at 30 ºS showed significant differences among seed origins in all morphometric parameters and also in yields. Altiplano genotypes had larger panicule length but no seed production. A significant influence of the different quinoa genotypes on chemical composition and functional properties was also observed. Protein concentration ranged from 11.13 to 16.18 g.100 g-1 d.m., while total dietary fiber content ranged from 8.07-12.08 g.100 g-1 d.m., and both were the highest in Villarrica ecotype. An adequate balance of essential amino acids was also observed. Sucrose was the predominant sugar in all genotypes. Antioxidant activity was high in all genotypes, and it was highest in Faro genotype (79.58% inhibition.

  1. Genetic diversity for grain Zn concentration in finger millet genotypes:Potential for improving human Zn nutrition

    Institute of Scientific and Technical Information of China (English)

    Ramegowda Yamunarani; Geetha Govind; Venkategowda Ramegowda; Harshavardhan Vokkaliga Thammegowda; Shankar Ambarahalli Guligowda

    2016-01-01

    Nearly half of the world population suffers from micronutrient malnutrition,particularly Zn deficiency.It is important to understand genetic variation for uptake and translocation behaviors of Zn in relevant crop species to increase Zn concentration in edible parts.In the present study,genetic variation in grain Zn concentration of 319 finger millet genotypes was assessed.Large genetic variation was found among the genotypes,with concentrations ranging from 10 to 86 μg g-1grain.Uptake and translocation studies with Zn/65 Zn application in 12 selected low-Zn genotypes showed wide variation in root uptake and shoot translocation,with genotypes GEC331 and GEC164 showing greater uptake and translocation.Genotypes GEC164 and GEC543 showed increased grain Zn concentration.Genotypes GEC331 and GEC164 also showed improved yield under Zn treatment.Appreciable variation in grain Zn concentration among finger millet genotypes found in this study offers opportunities to improve Zn nutrition through breeding.

  2. Development and Clinical Evaluation of a Highly Sensitive DNA Microarray for Detection and Genotyping of Human Papillomaviruses

    Science.gov (United States)

    Oh, TaeJeong; Kim, ChangJin; Woo, SukKyung; Kim, TaeSeung; Jeong, DongJun; Kim, MyungSoon; Lee, Sunwoo; Cho, HyunSill; An, Sungwhan

    2004-01-01

    Human papillomavirus (HPV) has been found in cervical cancer, tonsillar cancer, and certain types of head and neck cancers. We report on a DNA microarray-based method for the simultaneous detection and typing of HPVs. The genotype spectrum discriminated by this HPV DNA microarray includes 15 high-risk HPV genotypes and 12 low-risk HPV genotypes. The HPV DNA microarray showed high degrees of specificity and reproducibility. We evaluated the performance of the HPV DNA microarray by application to three HPV-positive cell lines (HeLa, Caski, and SiHa cells) and two HPV-negative cell lines (C33A and A549 cells). The HPV DNA microarray successfully identified the known types of HPV present in the cell lines. The detection limit of the HPV DNA microarray was at least 100-fold higher than that of PCR. To assess the clinical applicability of the HPV DNA microarray, we performed the HPV genotyping assay with 73 nonmalignant and malignant samples from 39 tonsillar cancer patients. Twenty-five of the 39 (64.1%) malignant samples were positive for HPV, whereas 3 of 34 (8.8%) nonmalignant samples were positive for HPV. This result shows a preferential association of HPV with tonsillar carcinomas. The correlations of the presence of HPV with the grade of differentiation and risk factors were not significant. Our data show that the HPV DNA microarray may be useful for the diagnosis and typing of HPV in large-scale epidemiological studies. PMID:15243092

  3. Early detection of resistance to Mycosphaerella fijiensis in local genotypes of Musa in Ecuador

    OpenAIRE

    García, Galo Cedeño; Capello, Carmen Suarez; Coello, Danilo Vera; Fadda, Carlo; Jarvis, Devra; De Santis, Paola

    2017-01-01

    The aim of this research was to determine resistance reaction of 14 Musa genotypes against Mycosphaerella fijensis, causal agent of Black Sigatoka disease. Two experiments were conducted in laboratory and shadehause at Pichilingue research Station from the National Institute of Agricultural Research (INIAP), Ecuador. Genotypes considered were: Orito (AA), Guineo de jardín, Williams, Filipino, Gross Mitchel, Mulato and Morado, (AAA); Limeño, Maqueño, Dominico, Dominico-Hartón, Dominico gigante...

  4. Allele mining across DREB1A and DREB1B in diverse rice genotypes suggest a highly conserved pathway inducible by low temperature

    Indian Academy of Sciences (India)

    Clarissa Challam; Tapu Ghosh; Mayank Rai; Wricha Tyagi

    2015-06-01

    Low temperature stress is one of the major limiting factors affecting rice productivity in higher altitudes. DREB1A and DREB1B, are two transcription factors that have been reported to play key regulatory role in low temperature tolerance. In order to understand whether natural genetic variation in these two loci leads to cold tolerance or susceptibility, OsDREB1A and OsDREB1B were targeted across several rice genotypes showing differential response to low temperature. Expression data suggests induction of gene expression in shoots in response to low temperature in both tolerant and susceptible genotypes. Upon sequence analysis of 20 rice genotypes, eight nucleotide changes were identified including two in the coding region and six in the $5'$UTR. None of the discovered novel variations lie in the conserved region of the genes under study, thereby causing little or no changes in putative function of the corresponding proteins. In silico analysis using a diverse set of 400 O. sativa revealed much lower nucleotide diversity estimates across two DREB loci and one other gene (MYB2) involved in DREB pathway than those observed for other rice genes. None of the changes showed association with seedling stage cold tolerance, suggesting that nucleotide changes in DREB loci are unlikely to contribute to low temperature tolerance. So far, data concerning the physiological role and regulation of DREB1 in different genetic background are very limited; it is to be expected that they will be studied extensively in the near future.

  5. Genetic Diversity of the Hepatitis B Virus Strains in Cuba: Absence of West-African Genotypes despite the Transatlantic Slave Trade.

    Science.gov (United States)

    Rodríguez Lay, Licel A; Corredor, Marité B; Villalba, Maria C; Frómeta, Susel S; Wong, Meilin S; Valdes, Lidunka; Samada, Marcia; Sausy, Aurélie; Hübschen, Judith M; Muller, Claude P

    2015-01-01

    Cuba is an HBsAg low-prevalence country with a high coverage of anti-hepatitis B vaccine. Its population is essentially the result of the population mix of Spanish descendants and former African slaves. Information about genetic characteristics of hepatitis B virus (HBV) strains circulating in the country is scarce. The HBV genotypes/subgenotypes, serotypes, mixed infections, and S gene mutations of 172 Cuban HBsAg and HBV-DNA positive patients were determined by direct sequencing and phylogenetic analysis. Phylogenetic analysis of HBV S gene sequences showed a predominance of genotype A (92.4%), subgenotype A2 (84.9%) and A1 (7.6%). Genotype D (7.0%) and subgenotype C1 (0.6%) were also detected but typical (sub)genotypes of contemporary West-Africa (E, A3) were conspicuously absent. All genotype A, D, and C strains exhibited sequence characteristics of the adw2, ayw2, and adrq serotypes, respectively. Thirty-three (19.1%) patients showed single, double, or multiple point mutations inside the Major Hydrophilic domain associated with vaccine escape; eighteen (10.5%) patients had mutations in the T-cell epitope (amino acids 28-51), and there were another 111 point mutations downstream of the S gene. One patient had an HBV A1/A2 mixed infection. This first genetic study of Cuban HBV viruses revealed only strains that were interspersed with strains from particularly Europe, America, and Asia. The absence of genotype E supports previous hypotheses about an only recent introduction of this genotype into the general population in Africa. The presence of well-known vaccine escape (3.5%) and viral resistance mutants (2.9%) warrants strain surveillance to guide vaccination and treatment strategies.

  6. Genetic Diversity of the Hepatitis B Virus Strains in Cuba: Absence of West-African Genotypes despite the Transatlantic Slave Trade

    Science.gov (United States)

    Rodríguez Lay, Licel A.; Corredor, Marité B.; Villalba, Maria C.; Frómeta, Susel S.; Wong, Meilin S.; Valdes, Lidunka; Samada, Marcia; Sausy, Aurélie; Hübschen, Judith M.; Muller, Claude P.

    2015-01-01

    Cuba is an HBsAg low-prevalence country with a high coverage of anti-hepatitis B vaccine. Its population is essentially the result of the population mix of Spanish descendants and former African slaves. Information about genetic characteristics of hepatitis B virus (HBV) strains circulating in the country is scarce. The HBV genotypes/subgenotypes, serotypes, mixed infections, and S gene mutations of 172 Cuban HBsAg and HBV-DNA positive patients were determined by direct sequencing and phylogenetic analysis. Phylogenetic analysis of HBV S gene sequences showed a predominance of genotype A (92.4%), subgenotype A2 (84.9%) and A1 (7.6%). Genotype D (7.0%) and subgenotype C1 (0.6%) were also detected but typical (sub)genotypes of contemporary West-Africa (E, A3) were conspicuously absent. All genotype A, D, and C strains exhibited sequence characteristics of the adw2, ayw2, and adrq serotypes, respectively. Thirty-three (19.1%) patients showed single, double, or multiple point mutations inside the Major Hydrophilic domain associated with vaccine escape; eighteen (10.5%) patients had mutations in the T-cell epitope (amino acids 28-51), and there were another 111 point mutations downstream of the S gene. One patient had an HBV A1/A2 mixed infection. This first genetic study of Cuban HBV viruses revealed only strains that were interspersed with strains from particularly Europe, America, and Asia. The absence of genotype E supports previous hypotheses about an only recent introduction of this genotype into the general population in Africa. The presence of well-known vaccine escape (3.5%) and viral resistance mutants (2.9%) warrants strain surveillance to guide vaccination and treatment strategies. PMID:25978398

  7. Genotypic and phenotypic diversity of Ralstonia pickettii and Ralstonia insidiosa isolates from clinical and environmental sources including High-purity Water.

    LENUS (Irish Health Repository)

    Ryan, Michael P

    2011-08-30

    Abstract Background Ralstonia pickettii is a nosocomial infectious agent and a significant industrial contaminant. It has been found in many different environments including clinical situations, soil and industrial High Purity Water. This study compares the phenotypic and genotypic diversity of a selection of strains of Ralstonia collected from a variety of sources. Results Ralstonia isolates (fifty-nine) from clinical, industrial and environmental origins were compared genotypically using i) Species-specific-PCR, ii) PCR and sequencing of the 16S-23S rRNA Interspatial region (ISR) iii) the fliC gene genes, iv) RAPD and BOX-PCR and v) phenotypically using biochemical testing. The species specific-PCR identified fifteen out of fifty-nine designated R. pickettii isolates as actually being the closely related species R. insidiosa. PCR-ribotyping of the 16S-23S rRNA ISR indicated few major differences between the isolates. Analysis of all isolates demonstrated different banding patterns for both the RAPD and BOX primers however these were found not to vary significantly. Conclusions R. pickettii species isolated from wide geographic and environmental sources appear to be reasonably homogenous based on genotypic and phenotypic characteristics. R. insidiosa can at present only be distinguished from R. pickettii using species specific PCR. R. pickettii and R. insidiosa isolates do not differ significantly phenotypically or genotypically based on environmental or geographical origin.

  8. Genotype, soil type, and locale effects on reciprocal transplant vigor, endophyte growth, and microbial functional diversity of a narrow sagebrush hybrid zone in Salt Creek Canyon, Utah

    Science.gov (United States)

    Miglia, K.J.; McArthur, E.D.; Redman, R.S.; Rodriguez, R.J.; Zak, J.C.; Freeman, D.C.

    2007-01-01

    When addressing the nature of ecological adaptation and environmental factors limiting population ranges and contributing to speciation, it is important to consider not only the plant's genotype and its response to the environment, but also any close interactions that it has with other organisms, specifically, symbiotic microorganisms. To investigate this, soils and seedlings were reciprocally transplanted into common gardens of the big sagebrush hybrid zone in Salt Creek Canyon, Utah, to determine location and edaphic effects on the fitness of parental and hybrid plants. Endophytic symbionts and functional microbial diversity of indigenous and transplanted soils and sagebrush plants were also examined. Strong selection occurred against the parental genotypes in the middle hybrid zone garden in middle hybrid zone soil; F1 hybrids had the highest fitness under these conditions. Neither of the parental genotypes had superior fitness in their indigenous soils and habitats; rather F1 hybrids with the nonindigenous maternal parent were superiorly fit. Significant garden-by-soil type interactions indicate adaptation of both plant and soil microorganisms to their indigenous soils and habitats, most notably in the middle hybrid zone garden in middle hybrid zone soil. Contrasting performances of F1 hybrids suggest asymmetrical gene flow with mountain, rather than basin, big sagebrush acting as the maternal parent. We showed that the microbial community impacted the performance of parental and hybrid plants in different soils, likely limiting the ranges of the different genotypes.

  9. DETECTION OF MENDELIAN AND GENOTYPE FREQUENCY OF GROWTH HORMONE GENE IN ONGOLE CROSSBRED CATTLE MATED BY THE ARTIFICIAL INSEMINATION TECHNIQUE

    Directory of Open Access Journals (Sweden)

    U. Paputungan

    2012-06-01

    Full Text Available The objectives of this study were to detect the Mendelian mode inheritance of growth hormone (GH and to establish genotype frequency of GH gene in Ongole-crossbred cattle mated by the artificial insemination (AI technique. Total of 76 blood samples were collected from Ongole-crossbred cows and bulls (G0, and their progenies (G1 at the Tumaratas AI service center in North Sulawesi province, Indonesia. All blood samples were screened for the presence of GH locus using a PCR-RFLP method involving restricted enzyme Msp1 on 1.2 % of agarose gel. Data were analyzed using statistical program function in Excel XP. The results showed that GH locus using alleles of Msp1+ and Msp1- enzyme restriction in Ongole-crossbred cows and bulls was inherited to their Ongole-crossbred progenies following the Mendelian mode inheritance. This Mendelian inheritance generated by AI technique was not under genetic equilibrium for the Msp1 genotype frequencies in groups of G0 and G1. The breeding program using genotypes of bulls and cows (G0 for generating the genotype of GH Msp1 enzyme restriction by AI technique should be maintained to increase these various allele dispersion rates for breeding under genetic equilibrium of the Ongole-crossbred cattle population.

  10. Rapid Detection of Rare Deleterious Variants by Next Generation Sequencing with Optional Microarray SNP Genotype Data.

    Science.gov (United States)

    Watson, Christopher M; Crinnion, Laura A; Gurgel-Gianetti, Juliana; Harrison, Sally M; Daly, Catherine; Antanavicuite, Agne; Lascelles, Carolina; Markham, Alexander F; Pena, Sergio D J; Bonthron, David T; Carr, Ian M

    2015-09-01

    Autozygosity mapping is a powerful technique for the identification of rare, autosomal recessive, disease-causing genes. The ease with which this category of disease gene can be identified has greatly increased through the availability of genome-wide SNP genotyping microarrays and subsequently of exome sequencing. Although these methods have simplified the generation of experimental data, its analysis, particularly when disparate data types must be integrated, remains time consuming. Moreover, the huge volume of sequence variant data generated from next generation sequencing experiments opens up the possibility of using these data instead of microarray genotype data to identify disease loci. To allow these two types of data to be used in an integrated fashion, we have developed AgileVCFMapper, a program that performs both the mapping of disease loci by SNP genotyping and the analysis of potentially deleterious variants using exome sequence variant data, in a single step. This method does not require microarray SNP genotype data, although analysis with a combination of microarray and exome genotype data enables more precise delineation of disease loci, due to superior marker density and distribution.

  11. Comparing Human Metapneumovirus and Respiratory Syncytial Virus: Viral Co-Detections, Genotypes and Risk Factors for Severe Disease

    Science.gov (United States)

    Moe, Nina; Krokstad, Sidsel; Stenseng, Inger Heimdal; Christensen, Andreas; Skanke, Lars Høsøien; Risnes, Kari Ravndal; Nordbø, Svein Arne; Døllner, Henrik

    2017-01-01

    Background It is unclarified as to whether viral co-detection and human metapneumovirus (HMPV) genotypes relate to clinical manifestations in children with HMPV and lower respiratory tract infection (LRTI), and if the clinical course and risk factors for severe LRTI differ between HMPV and respiratory syncytial virus (RSV). Methods We prospectively enrolled hospitalized children aged <16 years with LRTI from 2006 to 2015. Children were clinically examined, and nasopharyngeal aspirates were analyzed using semi-quantitative, real-time polymerase chain reaction tests for HMPV, RSV and 17 other pathogens. HMPV-positive samples were genotyped. Results A total of 171 children had HMPV infection. HMPV-infected children with single virus (n = 106) and co-detections (n = 65) had similar clinical manifestations. No clinical differences were found between HMPV genotypes A (n = 67) and B (n = 80). The HMPV-infected children were older (median 17.2 months) than RSV-infected children (median 7.3 months, n = 859). Among single virus-infected children, no differences in age-adjusted LRTI diagnoses were found between HMPV and RSV. Age was an important factor for disease severity among single virus-infected children, where children <6 months old with HMPV had a milder disease than those with RSV, while in children 12–23 months old, the pattern was the opposite. In multivariable logistic regression analysis for each virus type, age ≥12 months (HMPV), and age <6 months (RSV), prematurity, ≥1 chronic disease and high viral loads of RSV, but not high HMPV viral loads, were risk factors for severe disease. Conclusions Among hospitalized children with LRTI, HMPV manifests independently of viral co-detections and HMPV genotypes. Disease severity in HMPV- and RSV-infected children varies in relation to age. A history of prematurity and chronic disease increases the risk of severe LRTI among HMPV- and RSV-infected children. PMID:28095451

  12. A multiplex PCR for the simultaneous detection and genotyping of the Echinococcus granulosus complex.

    Directory of Open Access Journals (Sweden)

    Ghalia Boubaker

    Full Text Available Echinococcus granulosus is characterized by high intra-specific variability (genotypes G1-G10 and according to the new molecular phylogeny of the genus Echinococcus, the E. granulosus complex has been divided into E. granulosus sensu stricto (G1-G3, E. equinus (G4, E. ortleppi (G5, and E. canadensis (G6-G10. The molecular characterization of E. granulosus isolates is fundamental to understand the spatio-temporal epidemiology of this complex in many endemic areas with the simultaneous occurrence of different Echinococcus species and genotypes. To simplify the genotyping of the E. granulosus complex we developed a single-tube multiplex PCR (mPCR allowing three levels of discrimination: (i Echinococcus genus, (ii E. granulosus complex in common, and (iii the specific genotype within the E. granulosus complex. The methodology was established with known DNA samples of the different strains/genotypes, confirmed on 42 already genotyped samples (Spain: 22 and Bulgaria: 20 and then successfully applied on 153 unknown samples (Tunisia: 114, Algeria: 26 and Argentina: 13. The sensitivity threshold of the mPCR was found to be 5 ng Echinoccoccus DNA in a mixture of up to 1 µg of foreign DNA and the specificity was 100% when template DNA from closely related members of the genus Taenia was used. Additionally to DNA samples, the mPCR can be carried out directly on boiled hydatid fluid or on alkaline-lysed frozen or fixed protoscoleces, thus avoiding classical DNA extractions. However, when using Echinococcus eggs obtained from fecal samples of infected dogs, the sensitivity of the mPCR was low (<40%. Thus, except for copro analysis, the mPCR described here has a high potential for a worldwide application in large-scale molecular epidemiological studies on the Echinococcus genus.

  13. A multiplex PCR for the simultaneous detection and genotyping of the Echinococcus granulosus complex.

    Science.gov (United States)

    Boubaker, Ghalia; Macchiaroli, Natalia; Prada, Laura; Cucher, Marcela A; Rosenzvit, Mara C; Ziadinov, Iskender; Deplazes, Peter; Saarma, Urmas; Babba, Hamouda; Gottstein, Bruno; Spiliotis, Markus

    2013-01-01

    Echinococcus granulosus is characterized by high intra-specific variability (genotypes G1-G10) and according to the new molecular phylogeny of the genus Echinococcus, the E. granulosus complex has been divided into E. granulosus sensu stricto (G1-G3), E. equinus (G4), E. ortleppi (G5), and E. canadensis (G6-G10). The molecular characterization of E. granulosus isolates is fundamental to understand the spatio-temporal epidemiology of this complex in many endemic areas with the simultaneous occurrence of different Echinococcus species and genotypes. To simplify the genotyping of the E. granulosus complex we developed a single-tube multiplex PCR (mPCR) allowing three levels of discrimination: (i) Echinococcus genus, (ii) E. granulosus complex in common, and (iii) the specific genotype within the E. granulosus complex. The methodology was established with known DNA samples of the different strains/genotypes, confirmed on 42 already genotyped samples (Spain: 22 and Bulgaria: 20) and then successfully applied on 153 unknown samples (Tunisia: 114, Algeria: 26 and Argentina: 13). The sensitivity threshold of the mPCR was found to be 5 ng Echinoccoccus DNA in a mixture of up to 1 µg of foreign DNA and the specificity was 100% when template DNA from closely related members of the genus Taenia was used. Additionally to DNA samples, the mPCR can be carried out directly on boiled hydatid fluid or on alkaline-lysed frozen or fixed protoscoleces, thus avoiding classical DNA extractions. However, when using Echinococcus eggs obtained from fecal samples of infected dogs, the sensitivity of the mPCR was low (Echinococcus genus.

  14. Detection of mumps virus genotype H in two previously vaccinated patients from Mexico City.

    Science.gov (United States)

    Del Valle, Alberto; García, Alí A; Barrón, Blanca L

    2016-06-01

    Infections caused by mumps virus (MuV) have been successfully prevented through vaccination; however, in recent years, an increasing number of mumps outbreaks have been reported within vaccinated populations. In this study, MuV was genotyped for the first time in Mexico. Saliva samples were obtained from two previously vaccinated patients in Mexico City who had developed parotitis. Viral isolation was carried out in Vero cells, and the SH and HN genes were amplified by RT-PCR. Amplicons were sequenced and compared to a set of reference sequences to identify the MuV genotype.

  15. MIRU-VNTR genotype diversity and indications of homoplasy in M. avium strains isolated from humans and slaughter pigs in Latvia.

    Science.gov (United States)

    Kalvisa, Adrija; Tsirogiannis, Constantinos; Silamikelis, Ivars; Skenders, Girts; Broka, Lonija; Zirnitis, Agris; Jansone, Inta; Ranka, Renate

    2016-09-01

    Diseases which are caused by non-tuberculous mycobacteria (NTM) are an increasing problem in the developed countries. In Latvia, one of the most clinically important members of NTM is Mycobacterium avium (M. avium), an opportunistic pathogen which has been isolated from several lung disease patients and tissue samples of slaughter pigs. This study was designed to characterize the genetic diversity of the M. avium isolates in Latvia and to compare the distribution of genotypic patterns among humans and pigs. Eleven (Hall and Salipante, 2010) clinical M. avium samples, isolated from patients of Center of Tuberculosis and Lung Diseases (years 2003-2010), and 32 isolates from pig necrotic mesenterial lymph nodes in different regions (years 2003-2007) were analyzed. The majority (42 of 43) of samples were identified as M. avium subsp. hominissuis; one porcine isolate belonged to M. avium subsp. avium. MIRU-VNTR genotyping revealed 13 distinct genotypes, among which nine genotype patterns, including M. avium subsp. avium isolate, were newly identified. IS1245 RFLP fingerprinting of 25 M. avium subsp. hominissuis samples yielded 17 different IS1245 RFLP patterns, allowing an efficient discrimination of isolates. Clusters of identical RFLP profiles were observed within host species, geographical locations and time frame of several years. Additional in silico analysis on simulated MIRU-VNTR genotype population datasets showed that the MIRU-VNTR pattern similarity could partly arise due to probabilistic increase of acquiring homoplasy among subpopulations, thus the similar MIRU-VNTR profiles of M. avium strains even in close geographical proximity should be interpreted with caution.

  16. DNA detection and genotypic identification of potentially human-pathogenic microsporidia from asymptomatic pet parrots in South Korea as a risk factor for zoonotic emergence.

    Science.gov (United States)

    Lee, So-Young; Lee, Sung-Seok; Lyoo, Young S; Park, Hee-Myung

    2011-12-01

    We detected and identified genotypes of human-pathogenic microsporidia in fecal samples from 51 asymptomatic captive-bred pet parrots in South Korea. Microsporidia were identified in 8 samples (15.7%); 7 parrots tested positive for Encephalitozoon hellem, and 1 parrot tested positive for both E. hellem and Encephalitozoon cuniculi. In genotypic identifications, E. hellem was present in genotypes 1A and 2B and E. cuniculi was present in genotype II. Pet parrots might be a source of human microsporidian infection.

  17. Detection and genotyping of cutaneous leishmaniasis species in the southeast of Iran: restriction enzyme analysis (RFLP

    Directory of Open Access Journals (Sweden)

    Khalili M

    2009-06-01

    Full Text Available "n Normal 0 false false false EN-GB X-NONE AR-SA MicrosoftInternetExplorer4 /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-qformat:yes; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri","sans-serif"; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-fareast-font-family:"Times New Roman"; mso-fareast-theme-font:minor-fareast; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-bidi-font-family:Arial; mso-bidi-theme-font:minor-bidi;} Background: Leishmaniasis is a parasitic infectious disease which causes skin sores. There is no effective laboratory screening tests for leishmaniasis. Some diagnostic techniques exist that allow parasite detection and species identification by special culture and microscopy, biochemical (Isoenzymes, immunologic (immunoassays, and molecular (PCR approaches. Specific major objectives of this study was to genotyping of Leishmania species in Bam and Shiraz city."n"n Methods: A total of 83 samples of Leishmania were collected from patients clinically suspected of cutaneous leishmaniasis. The geographic distributions of the samples were 55 samples from Bam and 28 from Shiraz city. For this propose samples of skin and bloods were blotted on filter paper. Genomic DNA extracted with a Genomic DNA extraction kit (AccuPrep, BIONEER. Aliquots of extracted DNA were kept at -20°C. region of ITS1 amplified with the published Leishmania-specific primers. 15-20mL of these amplicons, containing the amplified ITS1 region, was digested for 2h with HaeIII."n"n Results: All 55 samples from Bam were considered as L. tropica and the positive samples from Shiraz considered as L. tropica and just one sample was L. major which was belonged to

  18. Comparative analysis of genotypic diversity in Entamoeba nuttalli isolates from Tibetan macaques and rhesus macaques in China.

    Science.gov (United States)

    Guan, Yue; Feng, Meng; Cai, Junlong; Min, Xiangyang; Zhou, Xingyu; Xu, Qing; Tan, Ning; Cheng, Xunjia; Tachibana, Hiroshi

    2016-03-01

    We have recently demonstrated the potentially virulent species Entamoeba nuttalli as one of the highly prevalent parasites in rhesus macaques (Macaca mulatta) in Mount Long-hu and Gui-yang in China. Tibetan macaque (Macaca thibetana) is a unique species living in China. To evaluate the prevalence of Entamoeba species in wild Tibetan macaques, we obtained 89 stool samples in Mount E-mei of Si-chuan Province in China. PCR analysis detected E. nuttalli, Entamoeba coli, and Entamoeba polecki ST2 in 17%, 42%, and 66% of the samples, respectively, whereas Entamoeba histolytica and Entamoeba dispar were undetected. This study is the first to report on the detection of E. nuttalli from Tibetan macaques. Six E. nuttalli isolates were obtained, 18S rRNA gene and six tRNA-linked short tandem repeat (STR) loci of the isolates were sequenced. The Mantel test results gave an r value of 0.97 of relationships between geographical distance and genetic diversity of Chinese E. nuttalli populations, indicating a significant isolation-by-distance effect in Chinese E. nuttalli according to the tRNA-STR loci sequences. Structural analysis of E. nuttalli isolates based on tRNA-linked STR loci demonstrated three Chinese E. nuttalli populations with their respective features, but the Gui-yang population was located in the middle. In the distance-based NJ tree, E. nuttalli isolates were divided into five different branches, and E-mei isolates were attributed to an independent branch to distinguish them from Gui-yang and Long-hu isolates. Genetic analysis in this study provided clues of the genetic differences between E. nuttalli isolates from Tibetan macaques and rhesus macaques in China.

  19. Multi-allele genotyping platform for the simultaneous detection of mutations in the Wilson disease related ATP7B gene.

    Science.gov (United States)

    Amvrosiadou, Maria; Petropoulou, Margarita; Poulou, Myrto; Tzetis, Maria; Kanavakis, Emmanuel; Christopoulos, Theodore K; Ioannou, Penelope C

    2015-12-01

    Wilson's disease is an inherited disorder of copper transport in the hepatocytes with a wide range of genotype and phenotype characteristics. Mutations in the ATP7B gene are responsible for the disease. Approximately, over 500 mutations in the ATP7B gene have been described to date. We report a method for the simultaneous detection of the ten most common ATP7B gene mutations in Greek patients. The method comprises 3 simple steps: (i) multiplex PCR amplification of fragments in the ATP7B gene flanking the mutations (ii) multiplex primer extension reaction of the unpurified amplification products using allele-specific primers and (iii) visual detection of the primer extension reaction products within minutes by means of dry-reagent multi-allele dipstick assay using anti-biotin conjugated gold nanoparticles. Optimization studies on the efficiency and specificity of the PEXT reaction were performed. The method was evaluated by genotyping 46 DNA samples of known genotype and 34 blind samples. The results were fully concordant with those obtained by reference methods. The method is simple, rapid, cost-effective and it does not require specialized instrumentation or highly qualified personnel.

  20. Genetic Diversity of the Hepatitis B Virus Strains in Cuba: Absence of West-African Genotypes despite the Transatlantic Slave Trade.

    Directory of Open Access Journals (Sweden)

    Licel A Rodríguez Lay

    Full Text Available Cuba is an HBsAg low-prevalence country with a high coverage of anti-hepatitis B vaccine. Its population is essentially the result of the population mix of Spanish descendants and former African slaves. Information about genetic characteristics of hepatitis B virus (HBV strains circulating in the country is scarce. The HBV genotypes/subgenotypes, serotypes, mixed infections, and S gene mutations of 172 Cuban HBsAg and HBV-DNA positive patients were determined by direct sequencing and phylogenetic analysis. Phylogenetic analysis of HBV S gene sequences showed a predominance of genotype A (92.4%, subgenotype A2 (84.9% and A1 (7.6%. Genotype D (7.0% and subgenotype C1 (0.6% were also detected but typical (subgenotypes of contemporary West-Africa (E, A3 were conspicuously absent. All genotype A, D, and C strains exhibited sequence characteristics of the adw2, ayw2, and adrq serotypes, respectively. Thirty-three (19.1% patients showed single, double, or multiple point mutations inside the Major Hydrophilic domain associated with vaccine escape; eighteen (10.5% patients had mutations in the T-cell epitope (amino acids 28-51, and there were another 111 point mutations downstream of the S gene. One patient had an HBV A1/A2 mixed infection. This first genetic study of Cuban HBV viruses revealed only strains that were interspersed with strains from particularly Europe, America, and Asia. The absence of genotype E supports previous hypotheses about an only recent introduction of this genotype into the general population in Africa. The presence of well-known vaccine escape (3.5% and viral resistance mutants (2.9% warrants strain surveillance to guide vaccination and treatment strategies.

  1. Cultural diversity and work-group performance : Detecting the rules

    NARCIS (Netherlands)

    Girndt, T.

    2000-01-01

    With greater levels of international cooperation, work-groups are increasingly composed of members from different cultures. These groups often suffer from communication problems; however, research suggests that they also benefit from their members cultural diversity and generate higher ranges of pro

  2. Cultural diversity and work-group performance : Detecting the rules

    NARCIS (Netherlands)

    Girndt, T.

    2000-01-01

    With greater levels of international cooperation, work-groups are increasingly composed of members from different cultures. These groups often suffer from communication problems; however, research suggests that they also benefit from their members cultural diversity and generate higher ranges of

  3. Cultural diversity and work-group performance : Detecting the rules

    NARCIS (Netherlands)

    Girndt, T.

    2000-01-01

    With greater levels of international cooperation, work-groups are increasingly composed of members from different cultures. These groups often suffer from communication problems; however, research suggests that they also benefit from their members cultural diversity and generate higher ranges of pro

  4. The microbiology of Bandji, palm wine of Borassus akeassii from Burkina Faso: identification and genotypic diversity of yeasts, lactic acid and acetic acid bacteria.

    Science.gov (United States)

    Ouoba, L I I; Kando, C; Parkouda, C; Sawadogo-Lingani, H; Diawara, B; Sutherland, J P

    2012-12-01

    To investigate physicochemical characteristics and especially genotypic diversity of the main culturable micro-organisms involved in fermentation of sap from Borassus akeassii, a newly identified palm tree from West Africa. Physicochemical characterization was performed using conventional methods. Identification of micro-organisms included phenotyping and sequencing of: 26S rRNA gene for yeasts, 16S rRNA and gyrB genes for lactic acid bacteria (LAB) and acetic acid bacteria (AAB). Interspecies and intraspecies genotypic diversities of the micro-organisms were screened respectively by amplification of the ITS1-5.8S rDNA-ITS2/16S-23S rDNA ITS regions and repetitive sequence-based PCR (rep-PCR). The physicochemical characteristics of samples were: pH: 3.48-4.12, titratable acidity: 1.67-3.50 mg KOH g(-1), acetic acid: 0.16-0.37%, alcohol content: 0.30-2.73%, sugars (degrees Brix): 2.70-8.50. Yeast included mainly Saccharomyces cerevisiae and species of the genera Arthroascus, Issatchenkia, Candida, Trichosporon, Hanseniaspora, Kodamaea, Schizosaccharomyces, Trigonopsis and Galactomyces. Lactobacillus plantarum was the predominant LAB species. Three other species of Lactobacillus were also identified as well as isolates of Leuconostoc mesenteroides, Fructobacillus durionis and Streptococcus mitis. Acetic acid bacteria included nine species of the genus Acetobacter with Acetobacter indonesiensis as predominant species. In addition, isolates of Gluconobacter oxydans and Gluconacetobacter saccharivorans were also identified. Intraspecies diversity was observed for some species of micro-organisms including four genotypes for Acet. indonesiensis, three for Candida tropicalis and Lactobacillus fermentum and two each for S. cerevisiae, Trichosporon asahii, Candida pararugosa and Acetobacter tropicalis. fermentation of palm sap from B. akeassii involved multi-yeast-LAB-AAB cultures at genus, species and intraspecies level. First study describing microbiological and

  5. Detection and genotyping of human papillomavirus in urine samples from unvaccinated male and female adolescents in Italy.

    Science.gov (United States)

    Bianchi, Silvia; Frati, Elena Rosanna; Panatto, Donatella; Martinelli, Marianna; Amicizia, Daniela; Zotti, Carla Maria; Martinese, Morena; Bonanni, Paolo; Boccalini, Sara; Coppola, Rosa Cristina; Masia, Giuseppina; Meloni, Angelo; Castiglia, Paolo; Piana, Andrea; Gasparini, Roberto; Tanzi, Elisabetta

    2013-01-01

    The introduction of vaccination against Human Papillomavirus (HPV) in adolescent girls in 2006 has focused virological surveillance on this age group. As few studies have evaluated HPV infections in young populations, further data are needed in order to improve and extend prophylactic policy and to monitor epidemiological changes. The present study aimed at evaluating overall and type-specific HPV prevalence in both female and male adolescents in Italy. HPV DNA detection and genotyping was performed on urine samples collected from 870 unvaccinated adolescents (369 females, 501 males, 11-18 years of age) in five cities in Italy. Following DNA extraction by means of a commercial kit (NucliSENS(®)-miniMAG(®), bioMérieux), the L1 gene fragment was PCR amplified and genotyped by restriction fragment length polymorphism analysis. HPV DNA was detected in 1.5% of all samples, and in 3% and 0.4% of samples from females and males, respectively. In approximately 70% of HPV DNA positive adolescents, the infection was due to a single genotype, with 88.9% of genotypes belonging to the HR-clade. The only two HPV-positive boys (14 and 18 years old) had HPV-70 genotype. Only one of the 11 HPV-infected girls was in the 11-14 age-group. HPV prevalence was 4.2% in girls aged 15-18 years and 60% of infections were due to vaccine types HPV-16 or HPV-6/-11. This is one of the few studies, the first conducted in Italy, on HPV infection in adolescents. Urine testing is the easier way of detecting HPV infection in younger populations. Our data revealed a very low HPV prevalence, and no infections were observed in the 12-year-old vaccine target population. The majority of infections were seen in females aged 15-18 years. Overall, more than 50% and 30% of the potentially persistent HPV infections detected in this group could have been prevented by the quadrivalent and the bivalent vaccines, respectively.

  6. Detection and genotyping of human papillomavirus in urine samples from unvaccinated male and female adolescents in Italy.

    Directory of Open Access Journals (Sweden)

    Silvia Bianchi

    Full Text Available The introduction of vaccination against Human Papillomavirus (HPV in adolescent girls in 2006 has focused virological surveillance on this age group. As few studies have evaluated HPV infections in young populations, further data are needed in order to improve and extend prophylactic policy and to monitor epidemiological changes. The present study aimed at evaluating overall and type-specific HPV prevalence in both female and male adolescents in Italy. HPV DNA detection and genotyping was performed on urine samples collected from 870 unvaccinated adolescents (369 females, 501 males, 11-18 years of age in five cities in Italy. Following DNA extraction by means of a commercial kit (NucliSENS(®-miniMAG(®, bioMérieux, the L1 gene fragment was PCR amplified and genotyped by restriction fragment length polymorphism analysis. HPV DNA was detected in 1.5% of all samples, and in 3% and 0.4% of samples from females and males, respectively. In approximately 70% of HPV DNA positive adolescents, the infection was due to a single genotype, with 88.9% of genotypes belonging to the HR-clade. The only two HPV-positive boys (14 and 18 years old had HPV-70 genotype. Only one of the 11 HPV-infected girls was in the 11-14 age-group. HPV prevalence was 4.2% in girls aged 15-18 years and 60% of infections were due to vaccine types HPV-16 or HPV-6/-11. This is one of the few studies, the first conducted in Italy, on HPV infection in adolescents. Urine testing is the easier way of detecting HPV infection in younger populations. Our data revealed a very low HPV prevalence, and no infections were observed in the 12-year-old vaccine target population. The majority of infections were seen in females aged 15-18 years. Overall, more than 50% and 30% of the potentially persistent HPV infections detected in this group could have been prevented by the quadrivalent and the bivalent vaccines, respectively.

  7. Detection of east/central/south African genotype of chikungunya virus in Myanmar, 2010.

    Science.gov (United States)

    Tun, Mya Myat Ngwe; Thant, Kyaw Zin; Inoue, Shingo; Nabeshima, Takeshi; Aoki, Kotaro; Kyaw, Aung Kyaw; Myint, Tin; Tar, Thi; Maung, Kay Thwe Thwe; Hayasaka, Daisuke; Morita, Kouichi

    2014-08-01

    In 2010, chikungunya virus of the East Central South African genotype was isolated from 4 children in Myanmyar who had dengue-like symptoms. Phylogenetic analysis of the E1 gene revealed that the isolates were closely related to isolates from China, Thailand, and Malaysia that harbor the A226V mutation in this gene.

  8. Microsporidia detection and genotyping study of human pathogenic E. bieneusi in animals from Spain.

    Directory of Open Access Journals (Sweden)

    Ana Luz Galván-Díaz

    Full Text Available Microsporidia are ubiquitous parasites infecting all animal phyla and we present evidence that supports their zoonotic potential. Fecal samples taken from domestic (cats and dogs, farm (pigs, rabbits and ostriches and wild animals (foxes from different provinces of Spain were evaluated for microsporidia infection by light microscopy and PCR. After Microsporidia species identification, E. bieneusi genotypes were additionally studied by sequence analysis of the ITS region. Eighty-five samples out of 159 exhibited structures that were compatible with microsporidia spores by Webeŕs stain with 37 of them being confirmed by PCR. Microsporidia species identified included E. bieneusi, E. intestinalis and A. algerae. We report the first diagnosis of E. intestinalis and E. bieneusi in ostriches and A. algerae in pigs. We also provide new information on the molecular characterization of E. bieneusi isolates both in rabbits and ostriches. All of the E. bieneusi genotypes identified belonged to the zoonotic group of genotypes (Group I including genotypes A (dogs, I (pigs, D (rabbits and foxes and type IV (ostriches. Our results demonstrate that microsporidia are present in domestic, farm and wild animals in Spain, corroborating their potential role as a source of human infection and environmental contamination.

  9. Detecting specific genotype by environment interaction using marginal maximum likelihood estimation in the classical twin design

    NARCIS (Netherlands)

    D. Molenaar; S. van der Sluis; D.I. Boomsma; C.V. Dolan

    2012-01-01

    Considerable effort has been devoted to the analysis of genotype by environment (G × E) interactions in various phenotypic domains, such as cognitive abilities and personality. In many studies, environmental variables were observed (measured) variables. In case of an unmeasured environment, van der

  10. Detecting High-Order Epistasis in Nonlinear Genotype-Phenotype Maps.

    Science.gov (United States)

    Sailer, Zachary R; Harms, Michael J

    2017-03-01

    High-order epistasis has been observed in many genotype-phenotype maps. These multi-way interactions between mutations may be useful for dissecting complex traits and could have profound implications for evolution. Alternatively, they could be a statistical artifact. High-order epistasis models assume the effects of mutations should add, when they could in fact multiply or combine in some other nonlinear way. A mismatch in the "scale" of the epistasis model and the scale of the underlying map would lead to spurious epistasis. In this article, we develop an approach to estimate the nonlinear scales of arbitrary genotype-phenotype maps. We can then linearize these maps and extract high-order epistasis. We investigated seven experimental genotype-phenotype maps for which high-order epistasis had been reported previously. We find that five of the seven maps exhibited nonlinear scales. Interestingly, even after accounting for nonlinearity, we found statistically significant high-order epistasis in all seven maps. The contributions of high-order epistasis to the total variation ranged from 2.2 to 31.0%, with an average across maps of 12.7%. Our results provide strong evidence for extensive high-order epistasis, even after nonlinear scale is taken into account. Further, we describe a simple method to estimate and account for nonlinearity in genotype-phenotype maps.

  11. Novel biosensor-based microarray assay for detecting rs8099917 and rs12979860 genotypes

    Institute of Scientific and Technical Information of China (English)

    Pei-Yuan Li; Xiao-Jun Zhou; Lan Yao; Xin-Hua Fang; Jiang-Nan Ren; Jia-Wu Song

    2012-01-01

    AIM:To evaluate a novel biosensor-based microarray (BBM) assay for detecting rs12979860 and rs8099917genotypes.METHODS:Four probes specific for rs8099917C/T or rs12979860G/T detection and three sets of quality control probes were designed,constructed and arrayed on an optical biosensor to develop a microarray assay.Two sets of primers were used in a one tube polymerase chain reaction (PCR) system to amplify two target fragments simultaneously.The biosensor microarray contained probes that had been sequenced to confirm that they included the rsS099917C/T or rs12979860G/T alleles of interest and could serve as the specific assay standards.In addition to rehybridization of four probes of known sequence,a total of 40 clinical samples collected from hepatitis C seropositive patients were also tested.The target fragments of all 40 samples were amplified in a 50 μL PCR system.Ten μL of each amplicon was tested by BBM assay,and another 40 μL was used for sequencing.The agreement of the results obtained by the two methods was tested statistically using the kappa coefficient.The sensitivity of the BBM assay was evaluated using serial dilutions of ten clinical blood samples containing 103-104 white cells/lμL.RESULTS:As shown by polyacrylamide gel electrophoresis,two target segments of the interleukin 28B-associated polymorphisms (SNPs) were successfully amplified in the one-tube PCR system.The lengths of the two amplified fragments were consistent with the known length of the target sequences,137 and 159bps.After hybridization of the PCR amplicons with the probes located on the BBM array,the signals of each allele of both the rs8099917 SNPs and rs12979860 SNPs were observed simultaneously and were clearly visible by the unaided eye.The signals were distinct from each other,could be interpreted visually,and accurately recorded using an ordinary digital camera.To evaluate the specificity of the assay,both the plasmids and clinical samples were applied to the microarray

  12. Molecular sequence typing reveals genotypic diversity among Escherichia coli isolates recovered from a cantaloupe packinghouse in Northwestern Mexico

    Science.gov (United States)

    The increase in the consumption of fresh produce in the United States has correlated with a rise in the number of reported foodborne illnesses. To identify potential risk factors associated with post-harvest practices, the present study employed multilocus sequence typing (MLST) for the genotypic c...

  13. Extended Genetic Diversity of Bovine Viral Diarrhea Virus and Frequency of Genotypes and Subtypes in Cattle in Italy between 1995 and 2013

    OpenAIRE

    Camilla Luzzago; Stefania Lauzi; Erika Ebranati; Monica Giammarioli; Ana Moreno; Vincenza Cannella; Loretta Masoero; Elena Canelli; Annalisa Guercio; Claudio Caruso; Massimo Ciccozzi; Gian Mario De Mia; Pier Luigi Acutis; Gianguglielmo Zehender; Simone Peletto

    2014-01-01

    Genetic typing of bovine viral diarrhea virus (BVDV) has distinguished BVDV-1 and BVDV-2 species and an emerging putative third species (HoBi-like virus), recently detected in southern Italy, signaling the occurrence of natural infection in Europe. Recognizing the need to update the data on BVDV genetic variability in Italy for mounting local and European alerts, a wide collection of 5′ UTR sequences (n = 371) was selected to identify the frequency of genotypes and subtypes at the herd level....

  14. Systematic Inference of Copy-Number Genotypes from Personal Genome Sequencing Data Reveals Extensive Olfactory Receptor Gene Content Diversity

    Science.gov (United States)

    Waszak, Sebastian M.; Hasin, Yehudit; Zichner, Thomas; Olender, Tsviya; Keydar, Ifat; Khen, Miriam; Stütz, Adrian M.; Schlattl, Andreas; Lancet, Doron; Korbel, Jan O.

    2010-01-01

    Copy-number variations (CNVs) are widespread in the human genome, but comprehensive assignments of integer locus copy-numbers (i.e., copy-number genotypes) that, for example, enable discrimination of homozygous from heterozygous CNVs, have remained challenging. Here we present CopySeq, a novel computational approach with an underlying statistical framework that analyzes the depth-of-coverage of high-throughput DNA sequencing reads, and can incorporate paired-end and breakpoint junction analysis based CNV-analysis approaches, to infer locus copy-number genotypes. We benchmarked CopySeq by genotyping 500 chromosome 1 CNV regions in 150 personal genomes sequenced at low-coverage. The assessed copy-number genotypes were highly concordant with our performed qPCR experiments (Pearson correlation coefficient 0.94), and with the published results of two microarray platforms (95–99% concordance). We further demonstrated the utility of CopySeq for analyzing gene regions enriched for segmental duplications by comprehensively inferring copy-number genotypes in the CNV-enriched >800 olfactory receptor (OR) human gene and pseudogene loci. CopySeq revealed that OR loci display an extensive range of locus copy-numbers across individuals, with zero to two copies in some OR loci, and two to nine copies in others. Among genetic variants affecting OR loci we identified deleterious variants including CNVs and SNPs affecting ∼15% and ∼20% of the human OR gene repertoire, respectively, implying that genetic variants with a possible impact on smell perception are widespread. Finally, we found that for several OR loci the reference genome appears to represent a minor-frequency variant, implying a necessary revision of the OR repertoire for future functional studies. CopySeq can ascertain genomic structural variation in specific gene families as well as at a genome-wide scale, where it may enable the quantitative evaluation of CNVs in genome-wide association studies involving high

  15. Development of a single nucleotide polymorphism DNA microarray for the detection and genotyping of the SARS coronavirus.

    Science.gov (United States)

    Guo, Xi; Geng, Peng; Wang, Quan; Cao, Boyang; Liu, Bin

    2014-10-01

    Severe acute respiratory syndrome (SARS), a disease that spread widely in the world during late 2002 to 2004, severely threatened public health. Although there have been no reported infections since 2004, the extremely pathogenic SARS coronavirus (SARS-CoV), as the causative agent of SARS, has recently been identified in animals, showing the potential for the re-emergence of this disease. Previous studies showed that 27 single nucleotide polymorphism (SNP) mutations among the spike (S) gene of this virus are correlated closely with the SARS pathogenicity and epidemicity. We have developed a SNP DNA microarray in order to detect and genotype these SNPs, and to obtain related information on the pathogenicity and epidemicity of a given strain. The microarray was hybridized with PCR products amplified from cDNAs obtained from different SARS-CoV strains. We were able to detect 24 SNPs and determine the type of a given strain. The hybridization profile showed that 19 samples were detected and genotyped correctly by using our microarray, with 100% accuracy. Our microarray provides a novel method for the detection and epidemiological surveillance of SARS-CoV.

  16. Detection of Cryptosporidium hominis and novel Cryptosporidium bat genotypes in wild and captive Pteropus hosts in Australia.

    Science.gov (United States)

    Schiller, Sabine Eva; Webster, Koa Narelle; Power, Michelle

    2016-10-01

    Spillover of zoonotic pathogens from wildlife to humans has been identified as a primary threat to global health. In contrast, the process of reverse pathogen transmission (zooanthroponosis), whereby pathogens move from humans into wildlife species is still largely unexplored. Globally, increasing urbanisation and habitat loss are driving many wildlife species into urban and regional centres. In Australia, large numbers of flying foxes now live in close proximity to humans, increasing the risk of zooanthroponosis. The protozoan parasite Cryptosporidium is an emerging zoonotic parasite that infects a wide range of vertebrates yet there are limited studies on transmission potential of Cryptosporidium between humans and urban wildlife. To explore the presence of zooanthroponosis in flying foxes in Australia the occurrence and diversity of Cryptosporidium was investigated in urbanised wild and captive flying foxes. PCR screening of faecal samples (n=281) from seven wild sites and two captive facilities identified the presence of Cryptosporidium in 3.2% (95% CI 1.5% to 6.0%) of faecal samples. In faecal samples from wild sites Cryptosporidium occurrence was 1.7% (95% CI 0.3% to 4.8%) versus 5.9% (95% CI 2.2% to 12.4%) in samples from captive individuals, with no significant difference between captive and wild sites (p=0.077). Multilocus sequencing using three loci, 18s rDNA, actin and gp60 was used to identify Cryptosporidium in flying fox species. The host specific Cryptosporidium hominis was identified in faecal samples from two captive flying foxes, and one of these samples was confirmed as C. hominis at both actin and gp60. Sequencing of the 18s rDNA also revealed four novel Cryptosporidium genotypes in wild and captive individuals, actin and gp60 amplification and sequencing were unreliable for all four novel genotypes. These novel genotypes have been designated Cryptosporidium bat genotypes VIII-XI. This first report of Cryptosporidium spp. in Australian flying

  17. Genotype diversity of H9N2 viruses isolated from wild birds and chickens in Hunan Province, China.

    Science.gov (United States)

    Wang, Ba; Liu, Zhihua; Chen, Quanjiao; Gao, Zhimin; Fang, Fang; Chang, Haiyan; Chen, Jianjun; Xu, Bing; Chen, Ze

    2014-01-01

    Three H9N2 avian influenza viruses were isolated from the Dongting Lake wetland, among which one was from fresh egret feces, the other two were from chicken cloacal swabs in poultry markets. Phylogenetic analyses suggested that eight genes of the egret-derived H9N2 virus might come from Korean-like or American-like lineages. The two poultry-derived H9N2 viruses were reassortants between the CK/BJ/94-like and G1-like viruses. Except the PB1 genes (90.6%), the nucleotide sequence of other internal genes of the two viruses exhibited high homology (>95%). In addition, they also exhibited high homology (96-98.3%) with some genes of the H7N9 virus that caused an epidemic in China in 2013. Nucleotide sequence of the poultry-derived and egret-derived H9N2 viruses shared low homology. Infection studies showed that the egret-derived H9N2 virus was non-pathogenic to both mice and chickens, and the virus was unable to infect chickens even through 8 passages continuously in the lung. On the other hand, the chickens infected by poultry-derived viruses showed obvious clinical symptoms and even died; the infected mice showed no noticeable clinical symptoms and weight loss, but viruses could be detected in their lungs. In conclusion, for the egret-derived H9N2 virus, it would take a long adaptation process to achieve cross-species transmission in poultry and mammals. H9N2 viruses isolated at different times from the same host species in the same geographical region presented different evolutionary status, and virus isolated from different hosts in the same geographical region exhibited genetic diversity. Therefore, it is important to continue the H9N2 virus surveillance for understanding their evolutionary trends so as to provide guidance for disease control and prevention.

  18. Molecular detection and genotyping of Acanthamoeba spp. among stray dogs using conjunctival swab sampling.

    Science.gov (United States)

    Karakuş, Mehmet; Aykur, Mehmet; Özbel, Yusuf; Töz, Seray; Dağcı, Hande

    2016-12-01

    Acanthamoeba is one of the most common free-living amoebas (FLA) that present in environment. In humans, Acanthamoeba can cause an infection of the eye termed Acanthamoeba keratitis, which mostly occurs in contact lens wearers. In the present study, we aimed to screen the presence of Acanthamoeba DNA in stray dogs using previously collected conjunctival swab samples in a hyper-endemic area for canine leishmaniasis. Totally, 184 dogs were included in the study and 27 of them (14.6%) were found positive for Acanthamoeba according to the 18s rRNA gene sequencing. Two different genotypes (T4 and T5) were identified and T5 was firstly reported in Turkey in the present study. Statistical analysis was performed and no correlation was found between Leishmania and Acanthamoeba positivity (PAcanthamoeba among stray dogs. Further studies are necessary to reveal the infection status and genotypes among dogs and its possible correlation with leishmaniasis.

  19. Detection and molecular characterisation of Giardia lamblia genotypes in Sharjah, United Arab Emirates.

    Science.gov (United States)

    ElBakri, Ali; Samie, Amidou; Bessong, Pascal; Potgieter, Natasha; Odeh, Ra'ed Abu

    2014-08-01

    No data are available on Giardia lamblia genotypes in the United Arab Emirates (UAE). This study aimed to identify G. lamblia from DNA extracted from human stool samples to gain information on its prevalence and to perform molecular analysis on isolates collected from expatriates from different localities residing in Sharjah, UAE. In total, 111 healthy expatriates residing in Sharjah were screened for G. lamblia using nested PCR amplification of the small subunit ribosomal RNA (ssu-rRNA) gene. Positive samples were genotyped using a nested PCR amplifying the triosephosphate isomerase (tpi) gene to differentiate between the two human assemblages (A and B). A subset of the PCR products (n=23) were sequenced and their phylogenetic relationships were determined. Of the 111 participants, 67 (60.4%) were identified as positive for the ssu-rRNA gene. When genotyped for the tpi gene, 18.9% (21/111) were of assemblage A, 17.1% (19/111) belonged to assemblage B and 5.4% (6/111) showed patterns compatible with mixed infections. A strong correlation between the presence of diarrhoea and assemblage B was observed (χ(2)=10.553; p=0.001). Moreover, an association was also observed between mixed infections (A+B) and diarrhoea (χ(2)=8.899; p=0.003). No correlation between age, gender and geographic origin of the infected individual was noted. Phylogenetic analysis showed three clusters for the tpi gene. No relationship between the clusters and the origin of samples was noted. This study is the first to determine the infection rate and genotypic composition of Giardia in Sharjah, UAE. © The Author 2014. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  20. Study of Mycobacterium tuberculosis complex genotypic diversity in Malaysia reveals a predominance of ancestral East-African-Indian lineage with a Malaysia-specific signature.

    Science.gov (United States)

    Ismail, Fazli; Couvin, David; Farakhin, Izzah; Abdul Rahman, Zaidah; Rastogi, Nalin; Suraiya, Siti

    2014-01-01

    Tuberculosis (TB) still constitutes a major public health problem in Malaysia. The identification and genotyping based characterization of Mycobacterium tuberculosis complex (MTBC) isolates causing the disease is important to determine the effectiveness of the control and surveillance programs. This study intended a first assessment of spoligotyping-based MTBC genotypic diversity in Malaysia followed by a comparison of strains with those prevailing in neighboring countries by comparison with an international MTBC genotyping database. Spoligotyping was performed on a total of 220 M. tuberculosis clinical isolates collected in Kelantan and Kuala Lumpur. The results were compared with the SITVIT2 international database of the Pasteur Institute of Guadeloupe. Spoligotyping revealed 77 different patterns: 22 corresponded to orphan patterns while 55 patterns containing 198 isolates were assigned a Spoligo International Type (SIT) designation in the database (the latter included 6 newly created SITs). The eight most common SITs grouped 141 isolates (5 to 56 strains per cluster) as follows: SIT1/Beijing, n = 56, 25.5%; SIT745/EAI1-SOM, n = 33, 15.0%; SIT591/EAI6-BGD1, n = 13, 5.9%; SIT256/EAI5, n = 12, 5.5%; SIT236/EAI5, n = 10, 4.6%; SIT19/EAI2-Manila, n = 9, 4.1%; SIT89/EAI2-Nonthaburi, n = 5, 2.3%; and SIT50/H3, n = 3, 1.4%. The association between city of isolation and lineages was statistically significant; Haarlem and T lineages being higher in Kuala Lumpur (pMalaysia, and its probable ongoing evolution with locally evolved strains sharing a specific signature characterized by absence of spacers 37, 38, and 40. Pending complementary genotyping confirmation, we propose that SIT745/EAI-SOM is tentatively reclassified as SIT745/EAI-MYS.

  1. High Diversity of vacA and cagA Helicobacter pylori Genotypes in Patients with and without Gastric Cancer

    OpenAIRE

    Yolanda López-Vidal; Sergio Ponce-de-León; Gonzalo Castillo-Rojas; Rafael Barreto-Zúñiga; Aldo Torre-Delgadillo

    2008-01-01

    BACKGROUND: Helicobacter pylori is associated with chronic gastritis, peptic ulcers, and gastric cancer. The aim of this study was to assess the topographical distribution of H. pylori in the stomach as well as the vacA and cagA genotypes in patients with and without gastric cancer. METHODOLOGY/PRINCIPAL FINDINGS: Three gastric biopsies, from predetermined regions, were evaluated in 16 patients with gastric cancer and 14 patients with dyspeptic symptoms. From cancer patients, additional biops...

  2. Detection of Genotype 4 Swine Hepatitis E Virus in Systemic Tissues in Cross-Species Infected Rabbits

    Science.gov (United States)

    Wu, Qiaoxing; An, Junqing; She, Ruiping; Shi, Ruihan; Hao, Wenzhuo; Soomro, MajidHussain; Yuan, Xuerui; Yang, Jinling; Wang, Jingyuan

    2017-01-01

    Increasing evidence demonstrates that hepatitis E virus (HEV) can be transmitted across species. According to previous reports, swine HEV has two genotypes, genotype 3 and 4, and both can infect humans by the fecal-oral route. Thus, it is crucial for the control of HEV zoonotic transmission to evaluate the dynamics of viral shedding and distribution in different tissues during cross-species infection by HEV. In this study, rabbits were infected with genotype 4 swine HEV by the intraperitoneal route. The results showed that HEV RNA not only shed in the feces but also in the saliva of some rabbits during infection with swine HEV. Viremia appeared late after infection, and anti-HEV IgG was not obvious until the appearance of high viremia levels. After the rabbits were euthanized, a histopathological examination showed that the livers developed overt hepatitis accompanied by an elevation of alanine aminotransferase (ALT) and aspartate transaminase (AST). Furthermore, HEV RNA was detected in various tissues, especially in the salivary glands and tonsils. Subsequently, negative-stranded HEV RNA was practiced in tissues with positive HEV RNA, which demonstrated that HEV replicated in the tissues. Next, we harvested additional tissues from the liver, salivary gland, tonsil, spleen, thymus gland, lymph node and intestine, which are known as replication sites of swine HEV. Additionally, we also observed the HEV antigen distributed in the organs above through immunohistochemical staining. These results demonstrate that rabbits could be used as an animal model for researching cross-species infection of genotype 4 HEV. It is also noteworthy that HEV can shed in the saliva and presents the risk of droplet transmission. These new data provide valuable information for understanding cross-species infection by HEV. PMID:28129390

  3. Simultaneous Detection and Genotype Determination of HSV 1 and 2 by Real-time PCR Using Melting Curve Analysis and a Unique Pair of Primers.

    Science.gov (United States)

    Paryan, Mahdi; Mohammadi-Yeganeh, Samira; Rezvan, Houri; Kia, Vahid; Mansouri, Ardalan; Mirab Samiee, Siamak

    2017-02-01

    Herpes simplex virus (HSV) is a human pathogen that causes different pathologic manifestations. Rapid and feasible detection and discrimination methods for HSV genotyping is a challenge in clinical laboratories, especially in children suffering from herpetic encephalitis. A quantitative real-time polymerase chain reaction (PCR)-based genotyping assay using SYBR Green I was established. We designed only 1 pair of primer for HSV 1 and 2, targeting thymidine kinase gene conserved region. HSV genotypes were determined by PCR using melting curve analysis with LightCycler. Different HSV genotypes were successfully detected in all clinical samples. The melting temperature for HSV 1 and 2 was 85.5±0.78°C and 89±0.53°C, respectively. These 2 genotypes were completely distinguished by means of the accurate melting assay. Importantly, detection was reliably performed within only 1 hour. The assay had no cross-reactivity across species, an excellent dynamic range from 10 to 10 copies per reaction, a good intra-assay and interassay reproducibility, and a detection limit of a single copy per reaction. Our homebrew designed and validated quantitative real-time PCR followed by a melting curve analysis provided a rapid and convenient screening test for differential identification of HSV genotypes 1 and 2. We recommend the large-scale application of this method for HSV 1 and 2 detection.

  4. Genotyping-By-Sequencing (GBS) Detects Genetic Structure and Confirms Behavioral QTL in Tame and Aggressive Foxes (Vulpes vulpes).

    Science.gov (United States)

    Johnson, Jennifer L; Wittgenstein, Helena; Mitchell, Sharon E; Hyma, Katie E; Temnykh, Svetlana V; Kharlamova, Anastasiya V; Gulevich, Rimma G; Vladimirova, Anastasiya V; Fong, Hiu Wa Flora; Acland, Gregory M; Trut, Lyudmila N; Kukekova, Anna V

    2015-01-01

    The silver fox (Vulpes vulpes) offers a novel model for studying the genetics of social behavior and animal domestication. Selection of foxes, separately, for tame and for aggressive behavior has yielded two strains with markedly different, genetically determined, behavioral phenotypes. Tame strain foxes are eager to establish human contact while foxes from the aggressive strain are aggressive and difficult to handle. These strains have been maintained as separate outbred lines for over 40 generations but their genetic structure has not been previously investigated. We applied a genotyping-by-sequencing (GBS) approach to provide insights into the genetic composition of these fox populations. Sequence analysis of EcoT22I genomic libraries of tame and aggressive foxes identified 48,294 high quality SNPs. Population structure analysis revealed genetic divergence between the two strains and more diversity in the aggressive strain than in the tame one. Significant differences in allele frequency between the strains were identified for 68 SNPs. Three of these SNPs were located on fox chromosome 14 within an interval of a previously identified behavioral QTL, further supporting the importance of this region for behavior. The GBS SNP data confirmed that significant genetic diversity has been preserved in both fox populations despite many years of selective breeding. Analysis of SNP allele frequencies in the two populations identified several regions of genetic divergence between the tame and aggressive foxes, some of which may represent targets of selection for behavior. The GBS protocol used in this study significantly expanded genomic resources for the fox, and can be adapted for SNP discovery and genotyping in other canid species.

  5. Genotyping-By-Sequencing (GBS Detects Genetic Structure and Confirms Behavioral QTL in Tame and Aggressive Foxes (Vulpes vulpes.

    Directory of Open Access Journals (Sweden)

    Jennifer L Johnson

    Full Text Available The silver fox (Vulpes vulpes offers a novel model for studying the genetics of social behavior and animal domestication. Selection of foxes, separately, for tame and for aggressive behavior has yielded two strains with markedly different, genetically determined, behavioral phenotypes. Tame strain foxes are eager to establish human contact while foxes from the aggressive strain are aggressive and difficult to handle. These strains have been maintained as separate outbred lines for over 40 generations but their genetic structure has not been previously investigated. We applied a genotyping-by-sequencing (GBS approach to provide insights into the genetic composition of these fox populations. Sequence analysis of EcoT22I genomic libraries of tame and aggressive foxes identified 48,294 high quality SNPs. Population structure analysis revealed genetic divergence between the two strains and more diversity in the aggressive strain than in the tame one. Significant differences in allele frequency between the strains were identified for 68 SNPs. Three of these SNPs were located on fox chromosome 14 within an interval of a previously identified behavioral QTL, further supporting the importance of this region for behavior. The GBS SNP data confirmed that significant genetic diversity has been preserved in both fox populations despite many years of selective breeding. Analysis of SNP allele frequencies in the two populations identified several regions of genetic divergence between the tame and aggressive foxes, some of which may represent targets of selection for behavior. The GBS protocol used in this study significantly expanded genomic resources for the fox, and can be adapted for SNP discovery and genotyping in other canid species.

  6. Detection and diversity of various Arcobacter species in Danish poultry

    DEFF Research Database (Denmark)

    Atabay, H.I.; Wainø, Michael; Madsen, Mogens

    2006-01-01

    The prevalence and diversity of different Arcobacter spp. in various poultry species in Denmark were investigated using cultural and multiplex PCR methods. A pool of three fresh droppings obtained at the production site from 70 broiler chicken flocks aged 4-5 weeks was examined. In addition, pools...... from ducks at the abattoir were analyzed individually. In total, 85 Arcobacter isolates were obtained. Of these 45, 20 and 7 were identified as Arcobacter butzleri, Arcobacter cryaerophilus and Arcobacter skirrowii, respectively, using a multiplex PCR. Interestingly, some chicken isolates of A...... positive for A. cryaerophilus (7), A. butzleri (2) or A. skirrowii (2). Four (11%) of the 37 turkey flocks analyzed harboured either A. butzleri or A. cryaerophilus. The carriage rate of Arcobacter was higher in live ducks than those of live broiler chickens and turkeys in the present study. In addition...

  7. Genotyping and genetic diversity of Arcobacter butzleri and Arcobacter cryaerophilus isolated from different sources by using ERIC-PCR from India.

    Science.gov (United States)

    Ramees, Thadiyam Puram; Rathore, Ramswaroop Singh; Bagalkot, Prashanth Suresh; Sailo, Blessa; Mohan, Hosakote Venkatappa; Kumar, Ashok; Dhama, Kuldeep; Singh, Raj Kumar

    2014-01-01

    Arcobacters are important zoonotic pathogens and are transmitted through food and water. They are implicated in causing enteritis in animals and humans. Among the Arcobacter species, a wide genetic diversity has been documented, which reflects continuous evolving nature of these pathogens. To genotype and to know the genetic diversity of Arcobacter spp. (Arcobacter butzleri and Arcobacter cryaerophilus) isolated from different sources in India. Enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) was performed using genomic DNA of 49 Arcobacter isolates (27 A. butzleri and 22 A. cryaerophilus), recovered from a total of 506 samples of chicken meat, poultry skin, dairy cow milk and human stool as template and employing published primers. ERIC sequence was found to be present in all the 27 A. butzleri isolates which were grouped into 18 subtypes, while it was present in 20 out of 22 A. cryaerophilus isolates which were grouped into 14 subtypes. Less variation was observed within sequences of both the Arcobacter species as revealed in dendrogram analysis. The genotyping of A. butzleri isolates showed the presence of 2-8 distinct bands (∼150 to ∼1600 bp), while A. cryaerophilus showed 1-10 distinct bands (∼120 to ∼2900 bp). This study is the first report regarding genetic diversity of Indian Arcobacter isolates using ERIC-PCR. Close clustering between arcobacters of human and animal origin are indicative of probable zoonotic significance. So for these purposes, further explorative studies are suggested which would also help revealing the possibility of epidemiological relationships of different Arcobacter spp. as well as their public health concerns.

  8. Detection Performance of Polarization and Spatial Diversities for Indoor GNSS Applications

    Directory of Open Access Journals (Sweden)

    Mohammadreza Zaheri

    2012-01-01

    Full Text Available Multipath fading in the form of signal power fluctuation poses a formidable challenge to GNSS signal detection in harsh multipath environments such as indoors. Antenna diversity techniques such as polarization and spatial diversities can be used to combat multipath fading in wireless propagation channels. This paper studies and compares GPS signal detection performance enhancements arising from the spatial and polarization diversity techniques. Performance enhancements are quantified from a theoretical perspective and later verified based on several test measurements in various indoor environments. Enhancement is quantified based on measuring the correlation coefficient values between diversity branches, SNR levels, and computing the level crossing rate and average fade duration. In addition, the processing gain is quantified and the performance of each individual diversity system is evaluated. Experimental results show that, for a given target detection performance in terms of the probability of false alarm and the probability of detection, the required input SNR level to meet the target detection performance can be significantly reduced utilizing the diversity system.

  9. Detection, diversity, and population dynamics of waterborne Phytophthora ramorum populations

    Science.gov (United States)

    Catherine Eyre; Matteo Garbelotto

    2015-01-01

    Sudden oak death, the tree disease caused by Phytophthora ramorum, has significant environmental and economic impacts on natural forests on the U.S. west coast, plantations in the United Kingdom, and in the worldwide nursery trade. Stream baiting is vital for monitoring and early detection of the pathogen in high-risk areas and is performed...

  10. Diversity among other worlds: characterization of exoplanets by direct detection

    CERN Document Server

    Schneider, J; Aylward, A; Baudoz, P; Beuzit, J L; Brown, R; Cho, J; Dohlen, K; Ferrari, M; Galicher, R; Grasset, O; Grenfell, L; Guyon, O; Hough, J; Kasper, M; Keller, Ch; Longmore, A; López, B; Martin, E; Mawet, D; Ménard, F; Merin, B; Palle, E; Perrin, G; Pinfield, D; Sein, E; Shore, P; Sotin, Ch; Sozzetti, A; Stam, D; Surdej, J; Tamburini, F; Tinetti, G; Udry, S; Verinaud, C; Walker, D

    2008-01-01

    The physical characterization of exoplanets will require to take spectra at several orbital positions. For that purpose, a direct imaging capability is necessary. Direct imaging requires an efficient stellar suppression mechanism, associated with an ultrasmooth telescope. We show that before future large space missions (interferometer, 4-8 m class coronograph, external occulter or Fresnel imager), direct imaging of giant planets and close-by super-Earth are at the cross-road of a high scientific interest and a reasonable feasibility. The scientific interest lies in the fact that super-Earths share common geophysical attributes with Earths. They already begin to be detected by radial velocity (RV) and, together with giant planets, they have a larger area than Earths, making them detectable with a 1.5-2 m class telescope in reflected light. We propose such a (space) telescope be a first step before large direct imaging missions.

  11. Genotypic detection of acyclovir-resistant HSV-1: characterization of 67 ACV-sensitive and 14 ACV-resistant viruses.

    Science.gov (United States)

    Frobert, Emilie; Cortay, Jean-Claude; Ooka, Tadamasa; Najioullah, Fatiha; Thouvenot, Danielle; Lina, Bruno; Morfin, Florence

    2008-07-01

    Infections due to herpes simplex virus (HSV) resistant to acyclovir (ACV) represent an important clinical concern in immunocompromised patients. In order to switch promptly to an appropriate treatment, rapid viral susceptibility assays are required. We developed herein a genotyping analysis focusing on thymidine kinase gene (TK) mutations in order to detect acyclovir-resistant HSV in clinical specimens. A total of 85 HSV-1 positive specimens collected from 69 patients were analyzed. TK gene could be sequenced directly for 81 clinical specimens (95%) and 68 HSV-1 specimens could be characterized as sensitive or resistant by genotyping (84%). Genetic characterization of 67 susceptible HSV-1 specimens revealed 10 polymorphisms never previously described. Genetic characterization of 14 resistant HSV-1 revealed 12 HSV-1 with either TK gene additions/deletions (8 strains) or substitutions (4 strains) and 2 HSV-1 with no mutation in the TK gene. DNA polymerase gene was afterwards explored. With this rapid PCR-based assay, ACV-resistant HSV could be detected directly in clinical specimens within 24 h.

  12. Evaluation of a Probe-Based PCR-ELISA System for Simultaneous Semi Quantitative Detection and Genotyping of Human Cytomegalovirus (HCMV) Infection in Clinical Specimens.

    Science.gov (United States)

    Talkhabifard, Majid; Javid, Naeme; Moradi, Abdolvahab; Ghaemi, Amir; Tabarraei, Alijan

    2017-01-01

    Human cytomegalovirus (HCMV) is a common opportunistic pathogen that causes serious complications in immunosuppressed patients and infected newborns. In this study, PCR-ELISA was optimized for semi-quantitative detection of infection in clinical specimens and simultaneous genotyping of glycoprotein B for 4 major genotypes, due to its significance. During DIG-labeling PCR, a pair of primers amplifies a fragment of variable region of the glycoprotein B encoding sequence. Under optimized conditions, labeled Target amplicons hybridize to biotinated specific probes and are detected in an ELISA system. PCR-ELISA system showed specific performance with detection limit of approximately 100 copies of CMV DNA. The linear correlation was observed between the PCR-ELISA results (OD) and logarithmic scale of CMV (r=0.979). Repeatability of PCR-ELISA detection system for intra-assay and inter-assay was evaluated for negative and positive samples. In optimized conditions of hybridization, differentiation between genotypes of glycoprotein B was feasible using genotype-specific probes in PCR-ELISA genotyping system. In comparison with sequencing method, genotyping system was confirmed with kappa index of 1. PCR-ELISA is proposed as an applicable and reliable technique for semi-quantitative diagnosis and typing of the infection. This technique is flexible to apply in a variety of molecular fields.

  13. Diversity and Adaptation of Human Respiratory Syncytial Virus Genotypes Circulating in Two Distinct Communities: Public Hospital and Day Care Center

    Directory of Open Access Journals (Sweden)

    Gustavo Rocha Garcia

    2012-10-01

    Full Text Available HRSV is one of the most important pathogens causing acute respiratory tract diseases as bronchiolitis and pneumonia among infants. HRSV was isolated from two distinct communities, a public day care center and a public hospital in São José do Rio Preto – SP, Brazil. We obtained partial sequences from G gene that were used on phylogenetic and selection pressure analysis. HRSV accounted for 29% of respiratory infections in hospitalized children and 7.7% in day care center children. On phylogenetic analysis of 60 HRSV strains, 48 (80% clustered within or adjacent to the GA1 genotype; GA5, NA1, NA2, BA-IV and SAB1 were also observed. SJRP GA1 strains presented variations among deduced amino acids composition and lost the potential O-glycosilation site at amino acid position 295, nevertheless this resulted in an insertion of two potential O-glycosilation sites at positions 296 and 297. Furthermore, a potential O-glycosilation site insertion, at position 293, was only observed for hospital strains. Using SLAC and MEME methods, only amino acid 274 was identified to be under positive selection. This is the first report on HRSV circulation and genotypes classification derived from a day care center community in Brazil.

  14. Genetic diversity of NS5A protein from hepatitis C virus genotype 3a and its relationship to therapy response

    Directory of Open Access Journals (Sweden)

    Rahal Paula

    2010-02-01

    Full Text Available Abstract Background The quasispecies nature of HCV may have important implications for viral persistence, pathogenicity and resistance to antiviral agents. The variability of one of the viral proteins, NS5A, is believed to be related to the response to IFN therapy, the standard treatment for infection. In this study we analyzed the quasispecies composition of NS5A protein in patients infected with HCV genotype 3a, before IFN therapy. Methods Viral RNA was isolated from samples of 12 patients: four sustained virological responders (SVR, four non-responders (NR, and four end-of-treatment responders (ETR. cDNA was synthesized, the NS5A region was amplified and the fragments obtained were cloned. Fifteen clones from each patient were sequenced with eight primers, generating 179 contigs. Results Higher values for substitution (either synonymous or non-synonymous and for distance were found in the SVR group. However, the NR group showed relatively more non-synonymous mutations than the other groups, owing to the higher values of dN/dS in complete NS5A and most specific regions. Overall, NS5A protein is undergoing purifying selection, since all dN/dS ratios values are below 0.5. Conclusions Our study provides an overview of the genetic variability of complete NS5A protein in HCV genotype 3a.

  15. Diversity and Detection of Nitrate Assimilation Genes in Marine Bacteria

    OpenAIRE

    Allen, Andrew E.; Booth, Melissa G.; Frischer, Marc E.; Verity, Peter G.; Jonathan P Zehr; Zani, Sabino

    2001-01-01

    A PCR approach was used to construct a database of nasA genes (called narB genes in cyanobacteria) and to detect the genetic potential for heterotrophic bacterial nitrate utilization in marine environments. A nasA-specific PCR primer set that could be used to selectively amplify the nasA gene from heterotrophic bacteria was designed. Using seawater DNA extracts obtained from microbial communities in the South Atlantic Bight, the Barents Sea, and the North Pacific Gyre, we PCR amplified and se...

  16. Increasing the power to detect causal associations by combining genotypic and expression data in segregating populations.

    Directory of Open Access Journals (Sweden)

    Jun Zhu

    2007-04-01

    Full Text Available To dissect common human diseases such as obesity and diabetes, a systematic approach is needed to study how genes interact with one another, and with genetic and environmental factors, to determine clinical end points or disease phenotypes. Bayesian networks provide a convenient framework for extracting relationships from noisy data and are frequently applied to large-scale data to derive causal relationships among variables of interest. Given the complexity of molecular networks underlying common human disease traits, and the fact that biological networks can change depending on environmental conditions and genetic factors, large datasets, generally involving multiple perturbations (experiments, are required to reconstruct and reliably extract information from these networks. With limited resources, the balance of coverage of multiple perturbations and multiple subjects in a single perturbation needs to be considered in the experimental design. Increasing the number of experiments, or the number of subjects in an experiment, is an expensive and time-consuming way to improve network reconstruction. Integrating multiple types of data from existing subjects might be more efficient. For example, it has recently been demonstrated that combining genotypic and gene expression data in a segregating population leads to improved network reconstruction, which in turn may lead to better predictions of the effects of experimental perturbations on any given gene. Here we simulate data based on networks reconstructed from biological data collected in a segregating mouse population and quantify the improvement in network reconstruction achieved using genotypic and gene expression data, compared with reconstruction using gene expression data alone. We demonstrate that networks reconstructed using the combined genotypic and gene expression data achieve a level of reconstruction accuracy that exceeds networks reconstructed from expression data alone, and that

  17. One size fits all? : optimization of rainbow trout breeding program under diverse preferences and genotype-by-environment interaction

    NARCIS (Netherlands)

    Sae-Lim, P.

    2013-01-01

    Global fish breeders distribute improved animal material to several continents to be farmed under diverse environments, and for very different market conditions. When establishing a global breeding program, there is a need to assess whether or not a single breeding objective satisfies the markets ac

  18. One size fits all? : optimization of rainbow trout breeding program under diverse preferences and genotype-by-environment interaction

    NARCIS (Netherlands)

    Sae-Lim, P.

    2013-01-01

    Global fish breeders distribute improved animal material to several continents to be farmed under diverse environments, and for very different market conditions. When establishing a global breeding program, there is a need to assess whether or not a single breeding objective satisfies the markets

  19. Detection and genotyping of vancomycin-resistant Enterococcus spp. by multiplex polymerase chain reaction in Korean aquatic environmental samples.

    Science.gov (United States)

    Nam, Sehee; Kim, Min-jeong; Park, Chulmin; Park, Jong-Geun; Maeng, Pil Jae; Lee, Gyu-Cheol

    2013-07-01

    The distribution characteristics of Enterococcus spp., which are indicators of fecal pollution, were investigated at 33 sites within the 3 major water systems of Korea. Enterococci were detected at concentrations ranging from 1 to 37 CFU/100mL in 41 of 132 samples (31.1%) from the 3 major water systems. The overall average detected concentration was 1.2 CFU/100mL, while the average concentration for all detection sites was 5.3 CFU/100mL. After optimized multiplex polymerase chain reaction (PCR) was performed with newly developed VanA, VanB, VanC-1, and VanC-2/3 primers, concentrations of vancomycin-resistant Enterococcus spp. (VRE) ranging from 1 to 23 CFU/100mL were detected in 17 of 132 samples (12.9%). Of 216 individual enterococcal colonies, 64 (29.6%) displayed the VanC genotype. The results of a susceptibility test to vancomycin showed that the range of the minimal inhibitory concentration (MIC), an indicator of bacterial resistance, was 4 to 24μg/mL, with the average MIC at 9.2±4.5μg/mL. Of the bacterial isolates, 1 colony with the VanC-1 genotype was identified as E. gallinarum by 16S rDNA sequencing, whereas the other 63 colonies had the VanC-2/3 genotype and were identified as E. casseliflavus. Although these results imply that the major head bays of Korea are not contaminated with the highly vancomycin-resistant VanA- or VanB-type VREs, the misuse of antibiotics should be prohibited to minimize the presence of VREs and to maintain a safe water supply for protecting the health of the general population. Based on the study results, we also recommend the implementation of a continuous, broad-spectrum inspection program for Enterococcus spp. and VRE contamination in the major head bays. Furthermore, the multiplex PCR method described in this study can be used effectively for this purpose.

  20. Genetic diversity and phylogeography of highly zoonotic Echinococcus granulosus genotype G1 in the Americas (Argentina, Brazil, Chile and Mexico) based on 8279bp of mtDNA.

    Science.gov (United States)

    Laurimäe, Teivi; Kinkar, Liina; Andresiuk, Vanessa; Haag, Karen Luisa; Ponce-Gordo, Francisco; Acosta-Jamett, Gerardo; Garate, Teresa; Gonzàlez, Luis Miguel; Saarma, Urmas

    2016-11-01

    Echinococcus granulosus is a taeniid cestode and the etiological agent of an infectious zoonotic disease known as cystic echinococcosis (CE) or hydatid disease. CE is a serious public health concern in many parts of the world, including the Americas, where it is highly endemic in many regions. Echinococcus granulosus displays high intraspecific genetic variability and is divided into multiple genotypes (G1-G8, G10) with differences in their biology and etiology. Of these, genotype G1 is responsible for the majority of human and livestock infections and has the broadest host spectrum. However, despite the high significance to the public and livestock health, the data on genetic variability and regional genetic differences of genotype G1 in America are scarce. The aim of this study was to evaluate the genetic variability and phylogeography of G1 in several countries in America by sequencing a large portion of the mitochondrial genome. We analysed 8279bp of mtDNA for 52 E. granulosus G1 samples from sheep, cattle and pigs collected in Argentina, Brazil, Chile and Mexico, covering majority of countries in the Americas where G1 has been reported. The phylogenetic network revealed 29 haplotypes and a high haplotype diversity (Hd=0.903). The absence of phylogeographic segregation between different regions in America suggests the importance of animal transportation in shaping the genetic structure of E. granulosus G1. In addition, our study revealed many highly divergent haplotypes, indicating a long and complex evolutionary history of E. granulosus G1 in the Americas.

  1. Microbial diversity in lake sediments detected by PCR-DGGE

    Institute of Scientific and Technical Information of China (English)

    Xinqing ZHAO; Liuyan YANG; Can CHEN; Lin XIAO; Lijuan JIANG; Zhe MA; Haowei ZHU; Zhenyang YU; Daqiang YIN

    2008-01-01

    In this study,PCR-denaturing gradient gel electrophoresis (DGGE) was applied to analyze the microbial communities in lake sediments from Lake Xuanwu,Lake Mochou in Nanjing and Lake Taihu in Wuxi.Sediment samples from seven locations in three lakes were collected and their genomic DNAs were extracted.The DNA yields of the sediments of Lake Xuanwu and Lake Mochou were high (10 μg/g),while that of sediments in Lake Taihu was relatively low.After DNA purification,the 16S rDNA genes (V3 to V5 region) were amplified and the amplified DNA fragments were separated by parallel DGGE.The DGGE profiles showed that there were five common bands in all the lake sediment samples indicating that there were similarities among the populations of microorganisms in all the lake sediments.The DGGE profiles of Lake Xuanwu and Lake Mochou were similar and about 20 types of micro-organisms were identified in the sediment samples of both lakes.These results suggest that the sediment samples of these two city lakes (Xuanwu,Mochou) have similar microbial communities.However,the DGGE profiles of sediment samples in Lake Taihu were significantly differ-ent from these two lakes.Furthermore,the DGGE pro-files of sediment samples in different locations in Lake Taihu were also different,suggesting that the microbial communities in Lake Taihu are more diversified than those in Lake Xuanwu and Lake Mochou.The differences in microbial diversity may be caused by the different environmental conditions,such as redox potential,pH,and the concentrations of organic matters.Seven major bands of 16S rDNA genes fragments from the DGGE profiles of sediment samples were further re-amplified and sequenced.The results of sequencing analysis indicate that five sequences shared 99%-100% homology with known sequences (Bacillus and Brevibacillus,uncultured bacteria),while the other two sequences shared 93%-96% homology with known sequences (Acinetobacter,and Bacillus).The study shows that the PCR-DGGE tech

  2. The geographical patterns of symbiont diversity in the invasive legume Mimosa pudica can be explained by the competitiveness of its symbionts and by the host genotype.

    Science.gov (United States)

    Melkonian, Rémy; Moulin, Lionel; Béna, Gilles; Tisseyre, Pierre; Chaintreuil, Clémence; Heulin, Karine; Rezkallah, Naïma; Klonowska, Agnieszka; Gonzalez, Sophie; Simon, Marcelo; Chen, Wen-Ming; James, Euan K; Laguerre, Gisèle

    2014-07-01

    Variations in the patterns of diversity of symbionts have been described worldwide on Mimosa pudica, a pan-tropical invasive species that interacts with both α and β-rhizobia. In this study, we investigated if symbiont competitiveness can explain these variations and the apparent prevalence of β- over α-rhizobia. We developed an indirect method to measure the proportion of nodulation against a GFP reference strain and tested its reproducibility and efficiency. We estimated the competitiveness of 54 strains belonging to four species of β-rhizobia and four of α-rhizobia, and the influence of the host genotype on their competitiveness. Our results were compared with biogeographical patterns of symbionts and host varieties. We found: (i) a strong strain effect on competitiveness largely explained by the rhizobial species, with Burkholderia phymatum being the most competitive species, followed by B. tuberum, whereas all other species shared similar and reduced levels of competitiveness; (ii) plant genotype can increase the competitiveness of Cupriavidus taiwanensis. The latter data support the likelihood of the strong adaptation of C. taiwanensis with the M. pudica var. unijuga and help explain its prevalence as a symbiont of this variety over Burkholderia species in some environments, most notably in Taiwan. © 2013 Society for Applied Microbiology and John Wiley & Sons Ltd.

  3. Rapid detection of apolipoprotein E genotypes in Alzheimer's disease using polymerase chain reaction-single strand conformation polymorphism.

    Science.gov (United States)

    Kamruecha, Worawan; Chansirikarnjana, Sirintorn; Nimkulrat, Ekapot; Udommongkol, Chesda; Wongmek, Wanna; Thangnipon, Wipawan

    2006-07-01

    Apolipoprotein E (APOE) gene on chromosome 19q13.2 is encoded by three common alleles designated as epsilon2, epsilon3 and epsilon4. In Alzheimer's disease (AD) the epsilon4 allele is over-represented and is considered to be a major genetic risk factor. Several methods have been developed to determine APOE genotypes. Among them, polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) appears to be highly reliable. In this study, we improved the nonisotopic PCR-SSCP method for determining APOE genotypes in 42 cases of AD patients, 40 cases of non-AD dementia patients, and 49 cases of age-matched controls. DNA from the target sequence on APOE was amplified by PCR from peripheral blood genomic DNA. PCR products were electrophoresed in a non-denaturing polyacrylamide gel and visualized by silver staining. We found that the epsilon4 allele had a significantly high frequency of occurrence in AD patients (33.3%) compared with age-matched controls (13.3%) (chi(2) = 10.43, p = 0.001) and non-AD dementia (10%) (chi(2) = 13.02, p<0.001) whereas the epsilon3 allele was of high frequency in non-AD dementia (90%) compared with age-matched controls (85.7%) and AD patients (66.7%). APOE epsilon4 homozygotes were found only in AD groups. On the other hand, the epsilon2 allele was found only in an age-matched control. This study confirmed that the APOE psilon4 allele is a risk factor in Thai AD subjects and that the PCR-SSCP method is a rapid and useful means of detecting the APOE genotype in AD.

  4. Genotypic and phenotypic diversity of Douglas fir mycorrhizae grown under elevated atmospheric CO{sub 2} and temperature

    Energy Technology Data Exchange (ETDEWEB)

    Martin, K.J. [Mantech Environmental Services, Inc., Corvallis, OR (United States); Tuininga, A.R. [Oregon State Univ., Corvallis, OR (United States); Rygiewicz, P.T. [Environmental Research Laboratory, Corvallis, OR (United States)

    1995-09-01

    Our knowledge of the combined effects of elevated CO{sub 2} and global climate change (GCC) on below ground processes is limited. Given the interaction between ectomycorrhyzae (ECM) and plant carbon and nutrient physiology, studies of changes in ECM may be critical to expanding this knowledge. We are examining the diversity of Douglas fir ECM at the TERA GCC facility (US EPA). Morphotypes are tracked using gross morphological traits, and genetic diversity is assessed using PCR based restriction fragment length polymorphism (RFLP). Numbers of morphotypes increased during the experiment while some types decreased in abundance. Prevalence of each morphotype varies by GCC treatment and soil horizon. We have found multiple RFLP patterns associated with a single morphotype. Multiple morphotypes are also expect to share RFLP patterns (phenotypic variance within a genetic type).

  5. Genetic Diversity Strategy for the Management and Use of Rubber Genetic Resources: More than 1,000 Wild and Cultivated Accessions in a 100-Genotype Core Collection.

    Directory of Open Access Journals (Sweden)

    Livia Moura de Souza

    Full Text Available The rubber tree [Hevea brasiliensis (Willd. ex Adr. de Juss. Muell. Arg.] is the only plant species worldwide that is cultivated for the commercial production of natural rubber. This study describes the genetic diversity of the Hevea spp. complex that is available in the main ex situ collections of South America, including Amazonian populations that have never been previously described. Genetic data were analyzed to determine the genetic structure of the wild populations, quantify the allelic diversity and suggest the composition of a core collection to capture the maximum genetic diversity within a minimal sample size. A total of 1,117 accessions were genotyped with 13 microsatellite markers. We identified a total of 408 alleles, 319 of which were shared between groups and 89 that were private in different groups of accessions. In a population structure and principal component analysis, the level of clustering reflected a primary division into the following two subgroups: cluster 1, which consisted of varieties from the advanced breeding germplasm that originated from the Wickham and Mato Grosso accessions; and cluster 2, which consisted of the wild germplasm from the Acre, Amazonas, Pará and Rondônia populations and Hevea spp. The analyses revealed a high frequency of gene flow between the groups, with the genetic differentiation coefficient (GST estimated to be 0.018. Additionally, no distinct separation among the H. brasiliensis accessions and the other species from Amazonas was observed. A core collection of 99 accessions was identified that captured the maximum genetic diversity. Rubber tree breeders can effectively utilize this core collection for cultivar improvement. Furthermore, such a core collection could provide resources for forming an association panel to evaluate traits with agronomic and commercial importance. Our study generated a molecular database that should facilitate the management of the Hevea germplasm and its use for

  6. Genetic Diversity Strategy for the Management and Use of Rubber Genetic Resources: More than 1,000 Wild and Cultivated Accessions in a 100-Genotype Core Collection

    Science.gov (United States)

    Cerqueira-Silva, Carlos Bernardo Moreno; Silva, Carla Cristina; Mantello, Camila Campos; Conson, Andre Ricardo Oliveira; Vianna, João Paulo Gomes; Zucchi, Maria Imaculada; Scaloppi Junior, Erivaldo José; Fialho, Josefino de Freitas; de Moraes, Mario Luis Teixeira; Gonçalves, Paulo de Souza; de Souza, Anete Pereira

    2015-01-01

    The rubber tree [Hevea brasiliensis (Willd. ex Adr. de Juss.) Muell. Arg.] is the only plant species worldwide that is cultivated for the commercial production of natural rubber. This study describes the genetic diversity of the Hevea spp. complex that is available in the main ex situ collections of South America, including Amazonian populations that have never been previously described. Genetic data were analyzed to determine the genetic structure of the wild populations, quantify the allelic diversity and suggest the composition of a core collection to capture the maximum genetic diversity within a minimal sample size. A total of 1,117 accessions were genotyped with 13 microsatellite markers. We identified a total of 408 alleles, 319 of which were shared between groups and 89 that were private in different groups of accessions. In a population structure and principal component analysis, the level of clustering reflected a primary division into the following two subgroups: cluster 1, which consisted of varieties from the advanced breeding germplasm that originated from the Wickham and Mato Grosso accessions; and cluster 2, which consisted of the wild germplasm from the Acre, Amazonas, Pará and Rondônia populations and Hevea spp. The analyses revealed a high frequency of gene flow between the groups, with the genetic differentiation coefficient (GST) estimated to be 0.018. Additionally, no distinct separation among the H. brasiliensis accessions and the other species from Amazonas was observed. A core collection of 99 accessions was identified that captured the maximum genetic diversity. Rubber tree breeders can effectively utilize this core collection for cultivar improvement. Furthermore, such a core collection could provide resources for forming an association panel to evaluate traits with agronomic and commercial importance. Our study generated a molecular database that should facilitate the management of the Hevea germplasm and its use for subsequent genetic

  7. Genotypic and phenotypic diversity of PGPR fluorescent pseudomonads isolated from the rhizosphere of sugarcane (Saccharum officinarum L.).

    Science.gov (United States)

    Rameshkumar, Neelamegam; Ayyadurai, Niraikulam; Kayalvizhi, Nagarajan; Gunasekaran, Paramasamy

    2012-01-01

    The genetic diversity of plant growth-promoting rhizobacterial (PGPR) fluorescent pseudomonads associated with the sugarcane (Saccharum officinarum L.) rhizosphere was analyzed. Selected isolates were screened for plant growthpromoting properties including production of indole acetic acid, phosphate solubilization, denitrification ability, and production of antifungal metabolites. Furthermore, 16S rDNA sequence analysis was performed to identify and differentiate these isolates. Based on 16S rDNA sequence similarity, the isolates were designated as Pseudomonas plecoglossicida, P. fluorescens, P. libaniensis, and P. aeruginosa. Differentiation of isolates belonging to the same group was achieved through different genomic DNA fingerprinting techniques, including randomly amplified polymorphic DNA (RAPD), amplified ribosomal DNA restriction analysis (ARDRA), repetitive extragenic palindromic (REP), enterobacterial repetitive intergenic consensus (ERIC), and bacterial repetitive BOX elements (BOX) analyses. The genetic diversity observed among the isolates and rep-PCR-generated fingerprinting patterns revealed that PGPR fluorescent pseudomonads are associated with the rhizosphere of sugarcane and that P. plecoglossicida is a dominant species. The knowledge obtained herein regarding the genetic and functional diversity of fluorescent pseudomonads associated with the sugarcane rhizosphere is useful for understanding their ecological role and potential utilization in sustainable agriculture.

  8. Within Host Evolution Selects for a Dominant Genotype of Mycobacterium tuberculosis while T Cells Increase Pathogen Genetic Diversity

    Science.gov (United States)

    Copin, Richard; Louie, Eddie; Escuyer, Vincent; Gagneux, Sebastien; Palmer, Guy H.

    2016-01-01

    Molecular epidemiological assessments, drug treatment optimization, and development of immunological interventions all depend on understanding pathogen adaptation and genetic variation, which differ for specific pathogens. Mycobacterium tuberculosis is an exceptionally successful human pathogen, yet beyond knowledge that this bacterium has low overall genomic variation but acquires drug resistance mutations, little is known of the factors that drive its population genomic characteristics. Here, we compared the genetic diversity of the bacteria that established infection to the bacterial populations obtained from infected tissues during murine M. tuberculosis pulmonary infection and human disseminated M. bovis BCG infection. We found that new mutations accumulate during in vitro culture, but that in vivo, purifying selection against new mutations dominates, indicating that M. tuberculosis follows a dominant lineage model of evolution. Comparing bacterial populations passaged in T cell-deficient and immunocompetent mice, we found that the presence of T cells is associated with an increase in the diversity of the M. tuberculosis genome. Together, our findings put M. tuberculosis genetic evolution in a new perspective and clarify the impact of T cells on sequence diversity of M. tuberculosis. PMID:27973588

  9. Mycobacterial species diversity at a general hospital on the island of Crete: first detection of Mycobacterium lentiflavum in Greece.

    Science.gov (United States)

    Neonakis, Ioannis K; Gitti, Zoe; Kourbeti, Irene S; Michelaki, Helen; Baritaki, Maria; Alevraki, Georgia; Papadomanolaki, Evangelia; Tsafaraki, Ekaterini; Tsouri, Anna; Baritaki, Stavroula; Krambovitis, Elias; Spandidos, Demetrios A

    2007-01-01

    The objective of the present study was to investigate the diversity of mycobacterial isolates in a general hospital in Crete, Greece. 48 positive Lowenstein-Jensen cultures over a 3-y period were analysed by means of AccuProbe and GenoType assays. Non-tuberculous mycobacteria (NTM) comprised the majority of the isolates (56.3%, 27/48) vs 33.3% (16/48) of M. tuberculosis; 10.4% of the isolates could not be classified. Among NTM, M. lentiflavum was the predominant species isolated (9/27) followed by M. kansasii, M. gordonae and M. peregrinum, whereas no M. avium complex isolates were detected. This is the first detection of M. lentiflavum in Greece. The susceptibilities of the M. lentiflavum isolates to an extended panel of antibiotics were determined by the proportions method and the medical files of the 9 patients were reviewed. Three isolates were from urine, which is an unusual site. All strains exhibited multidrug resistance. The patients were adults with immunosuppression or predisposing conditions for NTM infection. Diagnosis of true infection was either not pursued or the patients died shortly after isolation.

  10. Detecting shifts in diversity limits from molecular phylogenies: what can we know?

    Science.gov (United States)

    McInnes, Lynsey; Orme, C David L; Purvis, A

    2011-11-01

    Large complete species-level molecular phylogenies can provide the most direct information about the macroevolutionary history of clades having poor fossil records. However, extinction will ultimately erode evidence of pulses of rapid speciation in the deep past. Assessment of how well, and for how long, phylogenies retain the signature of such pulses has hitherto been based on a--probably untenable--model of ongoing diversity-independent diversification. Here, we develop two new tests for changes in diversification 'rules' and evaluate their power to detect sudden increases in equilibrium diversity in clades simulated with diversity-dependent speciation and extinction rates. Pulses of diversification are only detected easily if they occurred recently and if the rate of species turnover at equilibrium is low; rates reported for fossil mammals suggest that the power to detect a doubling of species diversity falls to 50 per cent after less than 50 Myr even with a perfect phylogeny of extant species. Extinction does eventually draw a veil over past dynamics, suggesting that some questions are beyond the limits of inference, but sudden clade-wide pulses of speciation can be detected after many millions of years, even when overall diversity is constrained. Applying our methods to existing phylogenies of mammals and angiosperms identifies intervals of elevated diversification in each.

  11. Detection and molecular diversity of Giardia duodenalis and Cryptosporidium spp. in sheltered dogs and cats in Northern Spain.

    Science.gov (United States)

    Gil, Horacio; Cano, Lourdes; de Lucio, Aida; Bailo, Begoña; de Mingo, Marta Hernández; Cardona, Guillermo A; Fernández-Basterra, José A; Aramburu-Aguirre, Juan; López-Molina, Nuria; Carmena, David

    2017-06-01

    Domestic dogs and cats may act as natural reservoirs of a large number of zoonotic pathogens, including the enteric parasites Giardia duodenalis and Cryptosporidium spp., the most relevant protozoan species causing gastrointestinal disease worldwide. A cross-sectional epidemiological study aiming to assess the prevalence and molecular diversity of G. duodenalis and Cryptosporidium spp. was conducted in an animal rescue centre in the province of Álava (Northern Spain). A total of 194 and 65 faecal dropping samples from individual dogs and cats, respectively, were collected between November 2013 and June 2016. G. duodenalis cysts and Cryptosporidium spp. oocysts were detected by direct fluorescence microscopy and PCR-based methods targeting the small subunit ribosomal RNA gene of these parasites. Overall, G. duodenalis and Cryptosporidium spp. were detected in 33% (63/194) and 4.1% (8/194) of dogs, and 9.2% (6/65) and 4.6% (3/65) of cats, respectively. G. duodenalis and Cryptosporidium co-infections were observed in 1.5% (3/194) of dogs, but not in cats. No significant differences in infection rates could be demonstrated among dogs or cats according to their sex, age group, status, or geographical origin. Multi-locus sequence-based genotyping of the glutamate dehydrogenase and β-giardin genes of G. duodenalis allowed the characterization of 19 canine isolates that were unambiguously assigned to sub-assemblages AII (n=7), BIII (n=1), and BIV (n=7), and assemblages C (n=3) and D (n=1). Two feline isolates were genotyped as assemblages A and F, respectively. No mixed assemblage or sub-assemblage infections were identified. C. canis (n=5) and C. hominis (n=1) were the Cryptosporidium species found in dogs, whereas C. felis (n=1) was identified in cats. The finding of G. duodenalis sub-assemblages AII, BIII, and BIV circulating in dogs (but not cats) may have zoonotic potential, although most of the AII and BIV isolates sub-genotyped corresponded to genetic variants not

  12. Study of Mycobacterium tuberculosis complex genotypic diversity in Malaysia reveals a predominance of ancestral East-African-Indian lineage with a Malaysia-specific signature.

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    Fazli Ismail

    Full Text Available Tuberculosis (TB still constitutes a major public health problem in Malaysia. The identification and genotyping based characterization of Mycobacterium tuberculosis complex (MTBC isolates causing the disease is important to determine the effectiveness of the control and surveillance programs.This study intended a first assessment of spoligotyping-based MTBC genotypic diversity in Malaysia followed by a comparison of strains with those prevailing in neighboring countries by comparison with an international MTBC genotyping database.Spoligotyping was performed on a total of 220 M. tuberculosis clinical isolates collected in Kelantan and Kuala Lumpur. The results were compared with the SITVIT2 international database of the Pasteur Institute of Guadeloupe.Spoligotyping revealed 77 different patterns: 22 corresponded to orphan patterns while 55 patterns containing 198 isolates were assigned a Spoligo International Type (SIT designation in the database (the latter included 6 newly created SITs. The eight most common SITs grouped 141 isolates (5 to 56 strains per cluster as follows: SIT1/Beijing, n = 56, 25.5%; SIT745/EAI1-SOM, n = 33, 15.0%; SIT591/EAI6-BGD1, n = 13, 5.9%; SIT256/EAI5, n = 12, 5.5%; SIT236/EAI5, n = 10, 4.6%; SIT19/EAI2-Manila, n = 9, 4.1%; SIT89/EAI2-Nonthaburi, n = 5, 2.3%; and SIT50/H3, n = 3, 1.4%. The association between city of isolation and lineages was statistically significant; Haarlem and T lineages being higher in Kuala Lumpur (p<0.01. However, no statistically significant differences were noted when comparing drug resistance vs. major lineages, nor between gender and clades.The ancestral East-African-Indian (EAI lineage was most predominant followed by the Beijing lineage. A comparison of strains with those prevailing in neighboring countries in South Asia, East Asia and South East Asia underlined the phylogeographical specificity of SIT745 for Malaysia, and its probable ongoing evolution

  13. Detection of Self Incompatibility Genotypes in Prunus africana: Characterization, Evolution and Spatial Analysis.

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    Judith Ssali Nantongo

    Full Text Available In flowering plants, self-incompatibility is an effective genetic mechanism that prevents self-fertilization. Most Prunus tree species exhibit a homomorphic gametophytic self-incompatibility (GSI system, in which the pollen phenotype is encoded by its own haploid genome. To date, no identification of S-alleles had been done in Prunus africana, the only member of the genus in Africa. To identify S-RNase alleles and hence determine S-genotypes in African cherry (Prunus africana from Mabira Forest Reserve, Uganda, primers flanking the first and second intron were designed and these amplified two bands in most individuals. PCR bands on agarose indicated 26 and 8 different S-alleles for second and first intron respectively. Partial or full sequences were obtained for all these fragments. Comparison with published S-RNase data indicated that the amplified products were S-RNase alleles with very high interspecies homology despite the high intraspecific variation. Against expectations for a locus under balancing selection, frequency and spatial distribution of the alleles in a study plot was not random. Implications of the results to breeding efforts in the species are discussed, and mating experiments are strongly suggested to finally prove the functionality of SI in P. africana.

  14. Detection of Self Incompatibility Genotypes in Prunus africana: Characterization, Evolution and Spatial Analysis.

    Science.gov (United States)

    Nantongo, Judith Ssali; Eilu, Gerald; Geburek, Thomas; Schueler, Silvio; Konrad, Heino

    2016-01-01

    In flowering plants, self-incompatibility is an effective genetic mechanism that prevents self-fertilization. Most Prunus tree species exhibit a homomorphic gametophytic self-incompatibility (GSI) system, in which the pollen phenotype is encoded by its own haploid genome. To date, no identification of S-alleles had been done in Prunus africana, the only member of the genus in Africa. To identify S-RNase alleles and hence determine S-genotypes in African cherry (Prunus africana) from Mabira Forest Reserve, Uganda, primers flanking the first and second intron were designed and these amplified two bands in most individuals. PCR bands on agarose indicated 26 and 8 different S-alleles for second and first intron respectively. Partial or full sequences were obtained for all these fragments. Comparison with published S-RNase data indicated that the amplified products were S-RNase alleles with very high interspecies homology despite the high intraspecific variation. Against expectations for a locus under balancing selection, frequency and spatial distribution of the alleles in a study plot was not random. Implications of the results to breeding efforts in the species are discussed, and mating experiments are strongly suggested to finally prove the functionality of SI in P. africana.

  15. Detection of mutually exclusive mosaicism in a girl with genotype-phenotype discrepancies.

    Science.gov (United States)

    Luo, Minjie; Mulchandani, Surabhi; Dubbs, Holly A; Swarr, Daniel; Pyle, Louise; Zackai, Elaine H; Spinner, Nancy B; Conlin, Laura K

    2015-12-01

    Discordance between clinical phenotype and genotype has multiple causes, including mosaicism. Phenotypes can be modified due to tissue distribution, or the presence of multiple abnormal cell lines with different genomic contributions. We have studied a 20-month-old female whose main phenotypes were failure to thrive, developmental delay, and patchy skin pigmentation. Initial chromosome and SNP microarray analysis of her blood revealed a non-mosaic ∼24 Mb duplication of 15q25.1q26.3 resulting from the unbalanced translocation of terminal 15q to the short arm of chromosome 15. The most common feature associated with distal trisomy 15q is prenatal and postnatal overgrowth, which was not consistent with this patient's phenotype. The phenotypic discordance, in combination with the patchy skin pigmentation, suggested the presence of mosaicism. Further analysis of skin biopsies from both hyper- and hypopigmented regions confirmed the presence of an additional cell line with the short arm of chromosome X deleted and replaced by the entire long arm of chromosome 15. The Xp deletion, consistent with a variant Turner Syndrome diagnosis, better explained the patient's phenotype. Parental studies revealed that the alterations in both cell lines were de novo and the duplicated distal 15q and the deleted Xp were from different parental origins, suggesting a mitotic event. The possible mechanism for the occurrence of two mutually exclusive structural rearrangements with both involving the long arm of chromosome 15 is discussed.

  16. Detection of Leptospira spp. in wildlife reservoir hosts in Ontario through comparison of immunohistochemical and polymerase chain reaction genotyping methods.

    Science.gov (United States)

    Shearer, Karen E; Harte, Michael J; Ojkic, Davor; Delay, Josepha; Campbell, Douglas

    2014-03-01

    A total of 460 kidney samples from wildlife (beavers, coyotes, deer, foxes, opossums, otters, raccoons, skunks) were obtained from road-kill and hunter/trapper donations in Ontario between January 2010 and November 2012. The objectives of the study were to detect Leptospira spp. by immunohistochemistry and polymerase chain reaction (PCR), to map presence of leptospires in wildlife relative to livestock and human populations, and to characterize positive samples by sequencing and comparison to leptospires known to affect domestic animals and humans. The proportion of samples that tested positive ranged from 0% to 42%, with the highest rates in skunks and raccoons. Leptospira spp. were present in kidneys of wildlife across Ontario, particularly in areas of high human density, and areas in which livestock populations are abundant. The PCR was too weak in most samples to permit genotyping and examination of the relationship between the leptospires found in this study and those affecting domestic animals and humans.

  17. Advances in alloimmune thrombocytopenia: perspectives on current concepts of human platelet antigens, antibody detection strategies, and genotyping.

    Science.gov (United States)

    Hayashi, Tomoya; Hirayama, Fumiya

    2015-07-01

    Alloimmunisation to platelets leads to the production of antibodies against platelet antigens and consequently to thrombocytopenia. Numerous molecules located on the platelet surface are antigenic and induce immune-mediated platelet destruction with symptoms that can be serious. Human platelet antigens (HPA) cause thrombocytopenias, such as neonatal alloimmune thrombocytopenia, post-transfusion purpura, and platelet transfusion refractoriness. Thirty-four HPA are classified into 28 systems. Assays to identify HPA and anti-HPA antibodies are critically important for preventing and treating thrombocytopenia caused by anti-HPA antibodies. Significant progress in furthering our understanding of HPA has been made in the last decade: new HPA have been discovered, antibody-detection methods have improved, and new genotyping methods have been developed. We review these advances and discuss issues that remain to be resolved as well as future prospects for preventing and treating immune thrombocytopenia.

  18. Prevalence and Distribution of High-Risk Genotypes of HPV in Women with Severe Cervical Lesions in Madrid, Spain: Importance of Detecting Genotype 16 and Other High-Risk Genotypes.

    Science.gov (United States)

    Mateos Lindemann, Maria Luisa; Sánchez Calvo, Juan Manuel; Chacón de Antonio, Jesús; Sanz, Itziar; Diaz, Esperanza; Rubio, Maria Dolores; de la Morena, Maria Luisa

    2011-01-01

    Background. Persistent infection with high-risk human papillomavirus (HR-HPV) has been demonstrated to be the necessary causal factor for developing cervical cancer. To know the most prevalent HR-HPV in different geographical areas is important to design diagnostic tests and implementation of vaccines. Objectives. The goal of this study is to evaluate the prevalence of HR-HPV in a total of 1001 patients, 198 with normal cytology results, 498 with low-grade squamous intraepithelial lesion (LSIL), and 205 with high-grade squamous intraepithelial lesion (HSIL) who attended our gynaecology department for opportunistic screening of HPV infection. Study design. Cervical samples were taken in a PreservCyt vial (Cytyc Corporation, Boxborough, MA). Hybrid capture assay was carried out following the manufacturer's instructions (Digene Corp., Gaithersburg, MD). All samples were further studied with polymerase chain reaction (PCR) (Linear Array HPV Genotyping Test, Roche Diagnostics, Mannheim, Germany). Results. Genotype 16 was the most prevalent HR-HPV in the three groups, 17.8% in the patients with normal cytology results, 22.3% in the LSIL group, and 60% in the HSIL group. Genotype 18 had a very low prevalence in all groups. Other HR-HPV genotypes such as genotype 31, genotype 58 and genotype 52 were found in significant numbers in HSIL patients. Discussion. Our data show that genotypes 16, 31, 58, and 52 are the most prevalent HR-HPV in cervical samples with severe intraepithelial lesion in Spain. There may be some geographical variation in prevalence of carcinogenic types, and it must be considered for designing diagnostic tests and vaccine.

  19. Mode of endosperm and wholemeal arabinoxylans solubilisation during rye breadmaking: genotypic diversity in the level, substitution degree and macromolecular characteristics.

    Science.gov (United States)

    Cyran, Malgorzata R; Dynkowska, Wioletta M

    2014-02-15

    Insight into solubilisation mechanisms of rye arabinoxylans during breadmaking is important for understanding the biochemical processes that affect bread attributes. Purified ethanol precipitated water-extractable arabinoxylans (WE-AX) and residual unextractable counterparts (WU-AX) were isolated from rye flours and resulting breads. While the endosperm flours had lower endoxylanase activities and higher arabinose-to-xylose ratios of WU-AX than those of corresponding wholemeals, there were not any significant differences between them in the mean amounts of WU-AX hydrolysed during breadmaking. Nevertheless, they were highly affected by rye cultivar used for breadmaking. On average, 42% and 36% of WU-AX were recovered in bread WE-AX fraction, causing its 11% and 8% increase, respectively for endosperm and wholemeal breads. Bread WE-AX, however, had lower molecular weights than those of starting flours, implying chains depolymerisation. Degree of AX solubilisation depends mainly on rye genotype used, determining combined effect of enzymatic and acid hydrolyses, associations and fine structure of AX.

  20. Stenotrophomonas maltophilia Phenotypic and Genotypic Diversity during a 10-year Colonization in the Lungs of a Cystic Fibrosis Patient.

    Science.gov (United States)

    Pompilio, Arianna; Crocetta, Valentina; Ghosh, Dipankar; Chakrabarti, Malabika; Gherardi, Giovanni; Vitali, Luca Agostino; Fiscarelli, Ersilia; Di Bonaventura, Giovanni

    2016-01-01

    The present study was carried out to understand the adaptive strategies developed by Stenotrophomonas maltophilia for chronic colonization of the cystic fibrosis (CF) lung. For this purpose, 13 temporally isolated strains from a single CF patient chronically infected over a 10-year period were systematically characterized for growth rate, biofilm formation, motility, mutation frequencies, antibiotic resistance, and pathogenicity. Pulsed-field gel electrophoresis (PFGE) showed over time the presence of two distinct groups, each consisting of two different pulsotypes. The pattern of evolution followed by S. maltophilia was dependent on pulsotype considered, with strains belonging to pulsotype 1.1 resulting to be the most adapted, being significantly changed in all traits considered. Generally, S. maltophilia adaptation to CF lung leads to increased growth rate and antibiotic resistance, whereas both in vivo and in vitro pathogenicity as well as biofilm formation were decreased. Overall, our results show for the first time that S. maltophilia can successfully adapt to a highly stressful environment such as CF lung by paying a "biological cost," as suggested by the presence of relevant genotypic and phenotypic heterogeneity within bacterial population. S. maltophilia populations are, therefore, significantly complex and dynamic being able to fluctuate rapidly under changing selective pressures.

  1. Stenotrophomonas maltophilia phenotypic and genotypic diversity during a 10-year colonization in the lungs of a cystic fibrosis patient

    Directory of Open Access Journals (Sweden)

    Arianna Pompilio

    2016-09-01

    Full Text Available The present study was carried out to understand the adaptive strategies developed by S. maltophilia for chronic colonization of the cystic fibrosis (CF lung. For this purpose, 12 temporally isolated strains from a single CF patient chronically infected over a 10-year period were systematically characterized for growth rate, biofilm formation, motility, mutation frequencies, antibiotic resistance, and pathogenicity. Pulsed-field gel electrophoresis showed over time the presence of 2 distinct groups, each consisting of 2 different pulsotypes. The pattern of evolution followed by S. maltophilia was dependent on pulsotype considered, with strains belonging to pulsotype 1.1 resulting to be the most adapted, being significantly changed in all traits considered. Generally, S. maltophilia adaptation to CF lung leads to increased growth rate and antibiotic resistance, whereas both in vivo and in vitro pathogenicity as well as biofilm formation were decreased. Our results show for the first time that S. maltophilia can successfully adapt to a highly stressful environment such as CF lung by paying a biological cost, as suggested by the presence of relevant genotypic and phenotypic heterogeneity within bacterial population. This indicates that S. maltophilia populations are significantly more complex and dynamic than can be described by the analysis of any single isolate and can fluctuate rapidly to changing selective pressures.

  2. Stenotrophomonas maltophilia Phenotypic and Genotypic Diversity during a 10-year Colonization in the Lungs of a Cystic Fibrosis Patient

    Science.gov (United States)

    Pompilio, Arianna; Crocetta, Valentina; Ghosh, Dipankar; Chakrabarti, Malabika; Gherardi, Giovanni; Vitali, Luca Agostino; Fiscarelli, Ersilia; Di Bonaventura, Giovanni

    2016-01-01

    The present study was carried out to understand the adaptive strategies developed by Stenotrophomonas maltophilia for chronic colonization of the cystic fibrosis (CF) lung. For this purpose, 13 temporally isolated strains from a single CF patient chronically infected over a 10-year period were systematically characterized for growth rate, biofilm formation, motility, mutation frequencies, antibiotic resistance, and pathogenicity. Pulsed-field gel electrophoresis (PFGE) showed over time the presence of two distinct groups, each consisting of two different pulsotypes. The pattern of evolution followed by S. maltophilia was dependent on pulsotype considered, with strains belonging to pulsotype 1.1 resulting to be the most adapted, being significantly changed in all traits considered. Generally, S. maltophilia adaptation to CF lung leads to increased growth rate and antibiotic resistance, whereas both in vivo and in vitro pathogenicity as well as biofilm formation were decreased. Overall, our results show for the first time that S. maltophilia can successfully adapt to a highly stressful environment such as CF lung by paying a “biological cost,” as suggested by the presence of relevant genotypic and phenotypic heterogeneity within bacterial population. S. maltophilia populations are, therefore, significantly complex and dynamic being able to fluctuate rapidly under changing selective pressures. PMID:27746770

  3. Applicability of anatid and galliform microsatellite markers to the genetic diversity studies of domestic geese (Anser anser domesticus through the genotyping of the endangered zatorska breed

    Directory of Open Access Journals (Sweden)

    Kapkowska Ewa

    2011-03-01

    Full Text Available Abstract Background The lack of a sufficient number of molecular markers seriously limits the cognition of genetic relationships within and between populations of many species. Likewise, the genetic diversity of domestic goose (Anser anser domesticus, with a great number of breeds throughout the world, remains poorly understood at the molecular level. Findings Thirty-five goose, seventeen duck and eight chicken microsatellite primer pairs were screened for their utility in the cross-species amplification on DNA from 96 individuals of Zatorska breed of domestic geese. Twenty-seven of 42 amplifying primer pairs revealed length-polymorphic products, but three of them were difficult to score. Fifteen primer pairs amplifying the same length product across all individuals. One polymorphic microsatellite locus was assigned by genotyping of known sex individuals to the Z-chromosome. Conclusions We present a set of 24 polymorphic microsatellite markers useful for population genetic studies of the domestic goose. Another 15 markers were classified as monomorphic, but they might also be suitable for the assessment of genetic diversity in geese.

  4. Epidemiological analysis of the dynamic and diversity of Salmonella spp. in five German pig production clusters using pheno- and genotyping methods: an exploratory study.

    Science.gov (United States)

    Niemann, J; Tietze, E; Ruddat, I; Fruth, A; Prager, R; Rabsch, W; Blaha, T; Münchhausen, C; Merle, R; Kreienbrock, L

    2015-03-23

    An exploratory study in five conventional pig production clusters was carried out to investigate the dynamic and diversity of Salmonella spp. within different production stages and sample site categories (pooled feces, direct and non-direct environment). Observing two production cycles per production cluster, a total of 1276 samples were collected along the pig production chain. Following a microbiological examination via culture, 2246 subcultures were generated out of 285 Salmonella positive samples and analysed by pheno- and genotyping methods. Based on a combination of serotyping, MLVA (multiple-locus variable-number tandem repeat (VNTR) analysis), PFGE (pulse-field gel electrophoresis) and MLST (multilocus sequence typing), an amount of 22.3% Salmonella positive samples were characterized in clonal lineages and its variants. Within each production cluster, one main clonal lineage could be identified and persisted over both production cycles with a large diversity of variants and a wide distribution in sample site categories and production stages. Results underline the importance of biosecurity with emphasis on the environment to prevent persistence and circulation of Salmonella within herds. Furthermore, the combined implementation of MLVA, PFGE and MLST with conventional culture techniques for isolate classification could be successfully applied as an effective and valuable tool for identifying similar pattern of Salmonella occurrence within pig production clusters.

  5. Molecular epidemiology of group A rotavirus among children admitted to hospital in Salto, Uruguay, 2011-2012: first detection of the emerging genotype G12.

    Science.gov (United States)

    Tort, Luis Fernando López; Victoria, Matías; Lizasoain A, Andrés; Castells, Matías; Maya, Leticia; Gómez, Mariela Martínez; Arreseigor, Edit; López, Patricia; Cristina, Juan; Leite, Jose Paulo Gagliardi; Colina, Rodney

    2015-05-01

    Group A rotavirus (RVA) is the most important etiologic agent of infant acute gastroenteritis (AGE) worldwide. Detection and molecular characterization of RVA in Salto department, Northwestern region of Uruguay, was conducted on 175 clinical samples, being 153 stool and 22 vomit samples, collected from hospitalized children with AGE, between 0-15 years old, from two hospitals of Salto city during 2011 and 2012. RVA was detected and genotyped by seminested multiplex RT-PCR in order to determine G- and P-genotypes. Positive samples were sequenced and phylogenetic analyses were carried out in order to determine lineages and sub-lineages. RVA were detected in 64 (37%) of the samples and the G and P genotypes observed were: 6% G1P[8], 23% G2P[4]/G2P[X]/GXP[4], 23% G3P[8]/G3P[X], 14% G12P[8]/G12P[X], 16% GXP[8], 1,5% G12P[9], 3% G2P[4]/[8], and 16% non-typeable. VP7 and VP4 genotypes related to DS-1 like gene constellation were prevalent during 2011 and those VP7 and VP4 genotypes related to Wa-like constellation were prevalent during 2012 (mainly represented by G3P[8]). Interestingly, RVA was detected in vomit samples in a high prevalence (41%). RVA was observed mainly in the age group between 1 and 5 years old (75% of the cases), and seasonality with a high detection rate in winter season was observed for the two consecutive years of surveillance. To our knowledge, this study represents the first detection and molecular characterization of RVA in Salto department, Northwestern region of Uruguay; and the first identification of the emerging genotype G12 in the country.

  6. Genotyping of Coxiella burnetii from domestic ruminants in northern Spain

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    Astobiza Ianire

    2012-12-01

    Full Text Available Abstract Background Information on the genotypic diversity of Coxiella burnetii isolates from infected domestic ruminants in Spain is limited. The aim of this study was to identify the C. burnetii genotypes infecting livestock in Northern Spain and compare them to other European genotypes. A commercial real-time PCR targeting the IS1111a insertion element was used to detect the presence of C. burnetii DNA in domestic ruminants from Spain. Genotypes were determined by a 6-loci Multiple Locus Variable number tandem repeat analysis (MLVA panel and Multispacer Sequence Typing (MST. Results A total of 45 samples from 4 goat herds (placentas, N = 4, 12 dairy cattle herds (vaginal mucus, individual milk, bulk tank milk, aerosols, N = 20 and 5 sheep flocks (placenta, vaginal swabs, faeces, air samples, dust, N = 21 were included in the study. Samples from goats and sheep were obtained from herds which had suffered abortions suspected to be caused by C. burnetii, whereas cattle samples were obtained from animals with reproductive problems compatible with C. burnetii infection, or consisted of bulk tank milk (BTM samples from a Q fever surveillance programme. C. burnetii genotypes identified in ruminants from Spain were compared to those detected in other countries. Three MLVA genotypes were found in 4 goat farms, 7 MLVA genotypes were identified in 12 cattle herds and 4 MLVA genotypes were identified in 5 sheep flocks. Clustering of the MLVA genotypes using the minimum spanning tree method showed a high degree of genetic similarity between most MLVA genotypes. Overall 11 different MLVA genotypes were obtained corresponding to 4 different MST genotypes: MST genotype 13, identified in goat, sheep and cattle from Spain; MST genotype 18, only identified in goats; and, MST genotypes 8 and 20, identified in small ruminants and cattle, respectively. All these genotypes had been previously identified in animal and human clinical samples from several

  7. Improved resolution of reef-coral endosymbiont (Symbiodinium species diversity, ecology, and evolution through psbA non-coding region genotyping.

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    Todd C LaJeunesse

    Full Text Available Ribosomal DNA sequence data abounds from numerous studies on the dinoflagellate endosymbionts of corals, and yet the multi-copy nature and intragenomic variability of rRNA genes and spacers confound interpretations of symbiont diversity and ecology. Making consistent sense of extensive sequence variation in a meaningful ecological and evolutionary context would benefit from the application of additional genetic markers. Sequences of the non-coding region of the plastid psbA minicircle (psbA(ncr were used to independently examine symbiont genotypic and species diversity found within and between colonies of Hawaiian reef corals in the genus Montipora. A single psbA(ncr haplotype was recovered in most samples through direct sequencing (~80-90% and members of the same internal transcribed spacer region 2 (ITS2 type were phylogenetically differentiated from other ITS2 types by substantial psbA(ncr sequence divergence. The repeated sequencing of bacterially-cloned fragments of psbA(ncr from samples and clonal cultures often recovered a single numerically common haplotype accompanied by rare, highly-similar, sequence variants. When sequence artifacts of cloning and intragenomic variation are factored out, these data indicate that most colonies harbored one dominant Symbiodinium genotype. The cloning and sequencing of ITS2 DNA amplified from these same samples recovered numerically abundant variants (that are diagnostic of distinct Symbiodinium lineages, but also generated a large amount of sequences comprising PCR/cloning artifacts combined with ancestral and/or rare variants that, if incorporated into phylogenetic reconstructions, confound how small sequence differences are interpreted. Finally, psbA(ncr sequence data from a broad sampling of Symbiodinium diversity obtained from various corals throughout the Indo-Pacific were concordant with ITS lineage membership (defined by denaturing gradient gel electrophoresis screening, yet exhibited

  8. High recombination frequency creates genotypic diversity in colonies of the leaf-cutting ant Acromyrmex echinatior

    DEFF Research Database (Denmark)

    Sirviö, A.; Gadau, J.; Rueppell, O.

    2006-01-01

    Honeybees are known to have genetically diverse colonies because queens mate with many males and the recombination rate is extremely high. Genetic diversity among social insect workers has been hypothesized to improve general performance of large and complex colonies, but this idea has not been...... tested in other social insects. Here, we present a linkage map and an estimate of the recombination rate for Acromyrmex echinatior, a leaf-cutting ant that resembles the honeybee in having multiple mating of queens and colonies of approximately the same size. A map of 145 AFLP markers in 22 linkage...... groups yielded a total recombinational size of 2076 cM and an inferred recombination rate of 161 kb cM-1 (or 6.2 cM Mb-1). This estimate is lower than in the honeybee but, as far as the mapping criteria can be compared, higher than in any other insect mapped so far. Earlier studies on A. echinatior have...

  9. Detection of four beta-thalassemia point mutations in Iranians using a PCR-ELISA genotyping system.

    Science.gov (United States)

    Gill, Pooria; Forouzandeh, Mehdi; Eshraghi, Naser; Ghalami, Mostafa; Safa, Majid; Noori-Daloii, Mohammad-Reza

    2008-04-01

    Development of molecular techniques with analytical capability of mutation detection can realize the medical diagnosis of diseases and improve people's health. beta-Thalassemia is one of the most prevalent genetic disorders in Iran and using a simple and rapid test in laboratories for the mass screening and prenatal diagnosis is essential. Here, we described a simple method for rapid detection of four common beta-thalassemia point mutations in Iranians (IVS-II-1 (G-->A), IVS-I-5 (G-->C), FSC 8/9 (+G), IVS-I-110 (G-->A)) using a PCR-ELISA genotyping system. After DNA isolation from whole blood, a segment of beta-globin gene was amplified by DIG-labeling PCR. The DIG-labeled PCR amplicons were denatured and added to biotinylated normal probe (for normal gene allele) and mutant probe (for mutant gene allele). The hybrids were detected by colorimetric ELISA method. The optical densities obtained using normal and mutant probes with heterozygous PCR products were very similar. The optical densities obtained using mutant probes were higher than normal probes with homozygous PCR products. In vice versa, the optical densities obtained using normal probes were higher than mutant probes with normal PCR products. All the results demonstrated that the PCR-ELISA has similar specificity in comparison to the amplification refractory mutation system.

  10. 重度龋儿童远缘链球菌基因多态性研究%Genotypic diversity of Streptococcus sobrinus in 3 to 4-year-old children suffering with severe early childhood caries

    Institute of Scientific and Technical Information of China (English)

    秦秀荣; 周琼; 秦满

    2009-01-01

    目的 采用随机引物聚合酶链反应(arbitrarily primed-polymerase chain reaction,AP-PCR)法初步探讨远缘链球菌在重度低龄儿童龋(severe early childhood caries,S-ECC)儿童与无龋儿童口腔中基因型分布的情况,分析其与婴幼儿龋发生之间的关系.方法 选取北京市海淀区和西城区14所幼儿园178名42~54个月龄儿童,S-ECC(患龋牙数≥5)组87例,无龋组91人.嚼蜡法采集刺激性唾液进行分离培养,典型菌落行革兰染色、生化鉴定并保种,提取基因组DNA,PCR鉴定变形链球菌和远缘链球菌,AP-PCR法对远缘链球菌临床分离株行基因型分析.结果 S-ECC组远缘链球菌检出率18%(16/87),显著高于无龋组的3%(3/91),差异有统计学意义(P<0.01).53株远缘链球菌临床分离株共检出22种基因型,S-ECC组个体基因型为1~3种,无龋组均为1种;此外,S-ECC组3名个体间存在相同基因型的菌株.S-ECC组基因型数与龋失补牙数间存在相关性(r=0.50,P<0.05).结论 S-ECC儿童远缘链球菌检出率明显高于无龋儿童,且菌株间存在基因多态性,个体携带基因型的种类数与其致龋性间存在相关性;远缘链球菌无关个体间存在相同基因型的菌株.%Objective To evaluate the genotypic diversity of Streptococcus sobrinus( Ss) between children suffering with severe early childhood caries (S-ECC) and caries-free children by arbitrarily primed-polymerase chain reaction(AP-PCR). Methods A total of 178 children aged from 42 to 54 months were recruited from 14 urban kindergartens. The S-ECC group contained 87 children with more than 5 decayed teeth, and the control group was composed of 91 caries-free children. Stimulated whole saliva was collected by chewing paraffin. All mutans streptococcus isolates were subcultured, biochemically characterised and identified by PCR as Streptococcus mutans(Sm)and Ss.Then the Ss isolates were genotyped by AP-PCILResults The frequency of Ss detection was 18% in S

  11. Root transcriptomes of two acidic soil adapted Indica rice genotypes suggest diverse and complex mechanism of low phosphorus tolerance.

    Science.gov (United States)

    Tyagi, Wricha; Rai, Mayank

    2017-03-01

    Low phosphorus (P) tolerance in rice is a biologically and agronomically important character. Low P tolerant Indica-type rice genotypes, Sahbhagi Dhan (SD) and Chakhao Poreiton (CP), are adapted to acidic soils and show variable response to low P levels. Using RNAseq approach, transcriptome data was generated from roots of SD and CP after 15 days of low P treatment to understand differences and similarities at molecular level. In response to low P, number of genes up-regulated (1318) was more when compared with down-regulated genes (761). Eight hundred twenty-one genes found to be significantly regulated between SD and CP in response to low P. De novo assembly using plant database led to further identification of 1535 novel transcripts. Functional annotation of significantly expressed genes suggests two distinct methods of low P tolerance. While root system architecture in SD works through serine-threonine kinase PSTOL1, suberin-mediated cell wall modification seems to be key in CP. The transcription data indicated that CP relies more on releasing its internally bound Pi and coping with low P levels by transcriptional and translational modifications and using dehydration response-based signals. Role of P transporters seems to be vital in response to low P in CP while sugar- and auxin-mediated pathway seems to be preferred in SD. At least six small RNA clusters overlap with transcripts highly expressed under low P, suggesting role of RNA super clusters in nutrient response in plants. These results help us to understand and thereby devise better strategy to enhance low P tolerance in Indica-type rice.

  12. M. tuberculosis genotypic diversity and drug susceptibility pattern in HIV- infected and non-HIV-infected patients in northern Tanzania

    Directory of Open Access Journals (Sweden)

    van Soolingen Dick

    2007-05-01

    Full Text Available Abstract Background Tuberculosis (TB is a major health problem and HIV is the major cause of the increase in TB. Sub-Saharan Africa is endemic for both TB and HIV infection. Determination of the prevalence of M. tuberculosis strains and their drug susceptibility is important for TB control. TB positive culture, BAL fluid or sputum samples from 130 patients were collected and genotyped. The spoligotypes were correlated with anti-tuberculous drug susceptibility in HIV-infected and non-HIV patients from Tanzania. Results One-third of patients were TB/HIV co-infected. Forty-seven spoligotypes were identified. Fourteen isolates (10.8% had new and unique spoligotypes while 116 isolates (89.2% belonged to 33 known spoligotypes. The major spoligotypes contained nine clusters: CAS1-Kili 30.0%, LAM11- ZWE 14.6%, ND 9.2%, EAI 6.2%, Beijing 5.4%, T-undefined 4.6%, CAS1-Delhi 3.8%, T1 3.8% and LAM9 3.8%. Twelve (10.8% of the 111 phenotypically tested strains were resistant to anti-TB drugs. Eight (7.2% were monoresistant strains: 7 to isoniazid (INH and one to streptomycin. Four strains (3.5% were resistant to multiple drugs: one (0.9% was resistant to INH and streptomycin and the other three (2.7% were MDR strains: one was resistant to INH, rifampicin and ethambutol and two were resistant to all four anti-TB drugs. Mutation in the katG gene codon 315 and the rpoB hotspot region showed a low and high sensitivity, respectively, as predictor of phenotypic drug resistance. Conclusion CAS1-Kili and LAM11-ZWE were the most common families. Strains of the Beijing family and CAS1-Kili were not or least often associated with resistance, respectively. HIV status was not associated with spoligotypes, resistance or previous TB treatment.

  13. Detection of cytotoxin genotypes of Helicobacter pylori in stomach, saliva and dental plaque

    NARCIS (Netherlands)

    Silva, D.G.; Stevens, R.H.; Macedo, J.M.B.; Albano, R.M.; Falabella, M.E.V.; Veerman, E.C.I.; Tinoco, E.M.B.

    2009-01-01

    The aim of this study was to detect the presence of Helicobacter pylori and its virulent cagA genes in the oral cavity of individuals with upper gastric diseases. Sixty-two individuals (42 ± 2.3 years) with dispepsy symptoms, referred for gastroscopy and who were H. pylori positive in the gastric

  14. Detection of cytotoxin genotypes of Helicobacter pylori in stomach, saliva and dental plaque

    NARCIS (Netherlands)

    Silva, D.G.; Stevens, R.H.; Macedo, J.M.B.; Albano, R.M.; Falabella, M.E.V.; Veerman, E.C.I.; Tinoco, E.M.B.

    2009-01-01

    The aim of this study was to detect the presence of Helicobacter pylori and its virulent cagA genes in the oral cavity of individuals with upper gastric diseases. Sixty-two individuals (42 ± 2.3 years) with dispepsy symptoms, referred for gastroscopy and who were H. pylori positive in the gastric bi

  15. Helicobacter pylori vacA and cagA genotype diversity and interferon gamma expression in patients with chronic gastritis and patients with gastric cancer.

    Science.gov (United States)

    Martínez-Carrillo, D N; Atrisco-Morales, J; Hernández-Pando, R; Reyes-Navarrete, S; Betancourt-Linares, R; Cruz-del Carmen, I; Illades Aguiar, B; Román-Román, A; Fernández-Tilapa, G

    2014-01-01

    Helicobacter pylori (H. pylori) is the main risk factor for the development of chronic gastritis, gastric ulcer, and gastric cancer. In H. pylori-infected individuals, the clinical result is dependent on various factors, among which are bacterial components, the immune response, and environmental influence. To compare IFN-γ expression with the H. pylori vacA and cagA genotypes in patients with chronic gastritis and patients with gastric cancer. Ninety-five patients diagnosed with chronic gastritis and 20 with gastric cancer were included in the study. Three gastric biopsies were taken; one was used for the molecular detection and genotyping of H. pylori; another was fixed in absolute alcohol and histologic sections were made for determining IFN-γ expression through immunohistochemistry. No differences were found in the cells that expressed IFN-γ between the patients with chronic gastritis (median percentage of positive cells: 82.6% in patients without H. pylori and 82% in infected persons) and those with gastric cancer (70.5% in H. pylori-negative patients and 78.5% in infected persons). IFN-γ expression was 69% in chronic gastritis patients infected with H. pylori vacAs2m2/cagA⁻ it was 86.5% in patients infected with H. pylori vacAs1m2/cagA⁻, 86.5% in vacAs1m1/cagA⁻, and 82% in vacAs1m1/cagA⁺. Similar data were found in the patients with gastric cancer. IFN-γ expression varied depending on the H. pylori vacA and cagA genotype, but not in accordance with the presence of chronic gastritis or gastric cancer.

  16. A survey of sesamin and composition of tocopherol variability from seeds of eleven diverse sesame (Sesamum indicum L.) genotypes using HPLC-PAD-ECD.

    Science.gov (United States)

    Williamson, Kelly S; Morris, J Brad; Pye, Quentin N; Kamat, Chandrashekhar D; Hensley, Kenneth

    2008-01-01

    The objective of this study was to determine the composition and content of sesamin and desmethyl tocopherols such as alpha-tocopherol (alphaT), delta-tocopherol (deltaT) and gamma-tocopherol (gammaT) in seeds of sesame (Sesamum indicum L.) for 11 genotypes conserved in the United States Department of Agriculture (USDA), Agricultural Research Service (ARS) and Plant Genetic Resources Conservation Unit (PGRCU) in Griffin, Georgia, USA. Seed accessions studied were collections from eight countries worldwide, including one landrace from Thailand and two cultivars from Texas, USA. Novel methodologies and analytical techniques described herein consisted of reverse-phase high-performance liquid chromatography (HPLC) connected in series with two detection systems specific for each analyte class. Photodiode array detection was employed for sesamin analysis and electrochemical array detection was used in the determination of tocopherols. A preliminary study was conducted to assess sesamin levels in 2003 and tocopherol levels in 2004 from sesame seed samples conserved at the USDA, ARS and PGRCU. In 2005, sesame seed samples were grown, harvested and evaluated for sesamin as well as tocopherol levels. The overall results (n = 3) showed that sesamin, alphaT, deltaT and gammaT levels were 0.67-6.35 mg/g, 0.034-0.175 microg/g, 0.44-3.05 microg/g and 56.9-99.3 microg/g respectively, indicating that the sesame seed accessions contained higher levels of sesamin and gammaT compared with alphaT and deltaT. Statistical analysis was conducted and significant differences were observed among the 11 different sesame genotypes. This suggests that genetic, environmental and geographical factors influence sesamin and desmethyl tocopherol content.

  17. Integrase inhibitor (INI) genotypic resistance in treatment-naive and raltegravir-experienced patients infected with diverse HIV-1 clades

    Science.gov (United States)

    Doyle, Tomas; Dunn, David T.; Ceccherini-Silberstein, Francesca; De Mendoza, Carmen; Garcia, Frederico; Smit, Erasmus; Fearnhill, Esther; Marcelin, Anne-Genevieve; Martinez-Picado, Javier; Kaiser, Rolf; Geretti, Anna Maria

    2015-01-01

    Objectives The aim of this study was to characterize the prevalence and patterns of genotypic integrase inhibitor (INI) resistance in relation to HIV-1 clade. Methods The cohort comprised 533 INI-naive subjects and 255 raltegravir recipients with viraemia who underwent integrase sequencing in routine care across Europe, including 134/533 (25.1%) and 46/255 (18.0%), respectively, with non-B clades (A, C, D, F, G, CRF01, CRF02, other CRFs, complex). Results No major INI resistance-associated mutations (RAMs) occurred in INI-naive subjects. Among raltegravir recipients with viraemia (median 3523 HIV-1 RNA copies/mL), 113/255 (44.3%) had one or more major INI RAMs, most commonly N155H (45/255, 17.6%), Q148H/R/K + G140S/A (35/255, 13.7%) and Y143R/C/H (12/255, 4.7%). In addition, four (1.6%) raltegravir recipients showed novel mutations at recognized resistance sites (E92A, S147I, N155D, N155Q) and novel mutations at other integrase positions that were statistically associated with raltegravir exposure (K159Q/R, I161L/M/T/V, E170A/G). Comparing subtype B with non-B clades, Q148H/R/K occurred in 42/209 (20.1%) versus 2/46 (4.3%) subjects (P = 0.009) and G140S/A occurred in 36/209 (17.2%) versus 1/46 (2.2%) subjects (P = 0.005). Intermediate- to high-level cross-resistance to twice-daily dolutegravir was predicted in 40/255 (15.7%) subjects, more commonly in subtype B versus non-B clades (39/209, 18.7% versus 1/46, 2.2%; P = 0.003). A glycine (G) to serine (S) substitution at integrase position 140 required one nucleotide change in subtype B and two nucleotide changes in all non-B clades. Conclusions No major INI resistance mutations occurred in INI-naive subjects. Reduced occurrence of Q148H/R/K + G140S/A was seen in non-B clades versus subtype B, and was explained by the higher genetic barrier to the G140S mutation observed in all non-B clades analysed. PMID:26311843

  18. Multiplex Detection and Genotyping of Point Mutations Involved in Charcot-Marie-Tooth Disease Using a Hairpin Microarray-Based Assay

    Directory of Open Access Journals (Sweden)

    Yasser Baaj

    2009-01-01

    Full Text Available We previously developed a highly specific method for detecting SNPs with a microarray-based system using stem-loop probes. In this paper we demonstrate that coupling a multiplexing procedure with our microarray method is possible for the simultaneous detection and genotyping of four point mutations, in three different genes, involved in Charcot-Marie-Tooth disease. DNA from healthy individuals and patients was amplified, labeled with Cy3 by multiplex PCR; and hybridized to microarrays. Spot signal intensities were 18 to 74 times greater for perfect matches than for mismatched target sequences differing by a single nucleotide (discrimination ratio for “homozygous” DNA from healthy individuals. “Heterozygous” mutant DNA samples gave signal intensity ratios close to 1 at the positions of the mutations as expected. Genotyping by this method was therefore reliable. This system now combines the principle of highly specific genotyping based on stem-loop structure probes with the advantages of multiplex analysis.

  19. A high-density Diversity Arrays Technology (DArT microarray for genome-wide genotyping in Eucalyptus

    Directory of Open Access Journals (Sweden)

    Myburg Alexander A

    2010-06-01

    Full Text Available Abstract Background A number of molecular marker technologies have allowed important advances in the understanding of the genetics and evolution of Eucalyptus, a genus that includes over 700 species, some of which are used worldwide in plantation forestry. Nevertheless, the average marker density achieved with current technologies remains at the level of a few hundred markers per population. Furthermore, the transferability of markers produced with most existing technology across species and pedigrees is usually very limited. High throughput, combined with wide genome coverage and high transferability are necessary to increase the resolution, speed and utility of molecular marker technology in eucalypts. We report the development of a high-density DArT genome profiling resource and demonstrate its potential for genome-wide diversity analysis and linkage mapping in several species of Eucalyptus. Findings After testing several genome complexity reduction methods we identified the PstI/TaqI method as the most effective for Eucalyptus and developed 18 genomic libraries from PstI/TaqI representations of 64 different Eucalyptus species. A total of 23,808 cloned DNA fragments were screened and 13,300 (56% were found to be polymorphic among 284 individuals. After a redundancy analysis, 6,528 markers were selected for the operational array and these were supplemented with 1,152 additional clones taken from a library made from the E. grandis tree whose genome has been sequenced. Performance validation for diversity studies revealed 4,752 polymorphic markers among 174 individuals. Additionally, 5,013 markers showed segregation when screened using six inter-specific mapping pedigrees, with an average of 2,211 polymorphic markers per pedigree and a minimum of 859 polymorphic markers that were shared between any two pedigrees. Conclusions This operational DArT array will deliver 1,000-2,000 polymorphic markers for linkage mapping in most eucalypt pedigrees

  20. Molecular detection and genotyping of Japanese Encephalitis Virus in mosquitoes during a 2010 outbreak in the Republic of Korea

    Science.gov (United States)

    Seo, Hyun-Ji; Kim, Heung Chul; Klein, Terry A.; Ramey, Andrew M.; Lee, Ji-Hyee; Kyung, Soon-Goo; Park, Jee-Yong; Cho, In-Soo; Yeh, Jung-Yong

    2013-01-01

    Japanese encephalitis virus (JEV), a mosquito-borne zoonotic pathogen, is one of the major causes of viral encephalitis. To reduce the impact of Japanese encephalitis among children in the Republic of Korea (ROK), the government established a mandatory vaccination program in 1967. Through the efforts of this program only 0-7 (mean 2.1) cases of Japanese encephalitis were reported annually in the ROK during the period of 1984-2009. However, in 2010 there was an outbreak of 26 confirmed cases of Japanese encephalitis, including 7 deaths. This represented a >12-fold increase in the number of confirmed cases of Japanese encephalitis in the ROK as compared to the mean number reported over the last 26 years and a 3.7-fold increase over the highest annual number of cases during this same period (7 cases). Surveillance of adult mosquitoes was conducted during the 2010 outbreak of Japanese encephalitis in the ROK. A total of 6,328 culicine mosquitoes belonging to 12 species from 5 genera were collected at 6 survey sites from June through October 2010 and assayed by reverse-transcription polymerase chain reaction (RT-PCR) for the presence of JEV. A total of 34/371 pooled samples tested positive for JEV (29/121 Culex tritaeniorhynchus, 4/64 Cx. pipiens, and 1/26 Cx. bitaeniorhynchus) as confirmed by sequencing of the pre-membrane and envelope protein coding genes. The maximum likelihood estimates of JEV positive individuals per 1,000 culicine vectors for Cx. tritaeniorhynchus, Cx. pipiens, and Cx. bitaeniorhynchus were 11.8, 5.6, and 2.8, respectively. Sequences of the JEV pre-membrane and envelope protein coding genes amplified from the culicine mosquitoes by RT-PCR were compared with those of JEV genotypes I-V. Phylogenetic analyses support the detection of a single genotype (I) among samples collected from the ROK in 2010.

  1. Genetic diversity and frequency of bovine viral diarrhea virus (BVDV) detected in cattle in Turkey.

    Science.gov (United States)

    Yilmaz, Huseyin; Altan, Eda; Ridpath, Julia; Turan, Nuri

    2012-09-01

    The aim of this study was to investigate the frequency and diversity of bovine viral diarrhea viruses (BVDV) infecting cattle in Turkey. A total of 1124 bovine blood samples from 19 farms in 4 different Turkish regions were tested by antigen capture ELISA (ACE). BVDV antigen was found in 26 samples from 13 farms. Only 20 of the 26 initial test positive cattle were available for retesting. Of these, 6 of 20 tested positive for BVDV, by ACE and real-time RT-PCR, one month after initial testing. Phylogenetic analysis, based on comparison of the E2 or the 5'UTR coding regions, from 19 of the 26 initial positive samples, indicated that 17 belonged to the BVDV-1 genotype and 2 to the BVDV-2 genotype. Comparison of 5'UTR sequences segregated 8 BVDV-1 strains (strains 5, 6, 10, 11, 12, 13, 17, and 19) to the BVDV1f, 1 strain (strain 8) to the BVDV1i and 1 strain (strain 14) to the BVDV1d subgenotypes. One strain (strain 4) did not group with other subgenotypes but was closer to the BVDV1f. The remaining 6 BVDV-1 strains (strains 1, 2, 3, 7, 9, and 18) segregated to a novel subgenotype. The E2 sequence comparison results were similar, with the exception that strain 5 grouped with the novel subgenotype rather than BVDV1f subgenotype. It appears that among the diverse BVDV strains in circulation there may be a subgenotype that is unique to Turkey. This should be considered in the design of diagnostics and vaccines to be used in Turkey.

  2. Detection, quantification and genotyping of noroviruses in oysters implicated in disease outbreaks

    DEFF Research Database (Denmark)

    Haefeli, Deborah; Gantenbein-Demarchi, Corinne; Böttiger, Blenda

    2012-01-01

    Noroviruses (NoVs) are a major cause of foodborne outbreaks of acute gastroenteritis in humans. Transmission of NoV is commonly linked to the consumption of oysters as they accumulate viruses through filter feeding in faecal-contaminated water. The NoV genogroups (G)I, GII and GIV infect humans....... While GI and GII have often been verified as causative agents of oyster-transmitted illness, GIV is rarely detected and has so far not been confirmed in outbreaks related to oysters. The aim of this study was to determine whether NoVs from oysters implicated in a disease outbreak were linked to the GI...

  3. Optical coherent detection Brillouin distributed optical fiber sensor based on orthogonal polarization diversity reception

    Institute of Scientific and Technical Information of China (English)

    Muping Song; Bin Zhao; Xianmin Zhang

    2005-01-01

    In Brillouin distributed optical fiber sensor, using optical coherent detection to detect Brillouin scattering optical signal is a good method, but there exists the polarization correlated detection problem. A novel detecting scheme is presented and demonstrated experimentally, which adopts orthogonal polarization diversity reception to resolve the polarization correlated detection problem. A laser is used as pump and reference light sources, a microwave electric-optical modulator (EOM) is adopted to produce frequency shift reference light, a polarization controller is used to control the polarization of the reference light which is changed into two orthogonal polarization for two adjacent acquisition periods. The Brillouin scattering light is coherently detected with the reference light, and the Brillouin scattering optical signal is taken out based on Brillouin frequency shift. After electronic processing, better Brillouin distributed sensing signal is obtained. A 25-km Brillouin distributed optical fiber sensor is achieved.

  4. Rapid and sensitive detection of fluoroquinolone-resistant Escherichia coli from urine samples using a genotyping DNA microarray.

    Science.gov (United States)

    Yu, Xiaolei; Susa, Milorad; Weile, Jan; Knabbe, Cornelius; Schmid, Rolf D; Bachmann, Till T

    2007-10-01

    Urinary tract infections (UTI) are among the most common bacterial infections in humans, with Escherichia coli being the major cause of infection. Fluoroquinolone resistance of uropathogenic E. coli has increased significantly over the last decade. In this study a microarray-based assay was developed and applied, which provides a rapid, sensitive and specific detection of fluoroquinolone-resistant E. coli in urine. The capture probes were designed against previously identified and described hotspots for quinolone resistance (codons 83 and 87 of gyrA). The key goals of this development were to reduce assay time while increasing the sensitivity and specificity as compared with a pilot version of a gyrA genotyping DNA microarray. The performance of the assay was demonstrated with pure cultures of 30 E. coli isolates as well as with urine samples spiked with 6 E. coli isolates. The microarray results were confirmed by standard DNA sequencing and were in full agreement with the phenotypic antimicrobial susceptibility testing using standard methods. The DNA microarray test displayed an assay time of 3.5h, a sensitivity of 100CFU/ml, and the ability to detect fluoroquinolone-resistant E. coli in the presence of a 10-fold excess of fluoroquinolone-susceptible E. coli cells. As a consequence, we believe that this microarray-based determination of antibiotics resistance has a true potential for the application in clinical routine laboratories in the future.

  5. Phenotypic and genotypic detection of methicillin-resistant Staphylococcus aureus in hunting dogs in Maiduguri metropolitan, Borno State, Nigeria

    Directory of Open Access Journals (Sweden)

    Muhammad Mustapha

    2016-05-01

    Full Text Available Aim: To determine the presence of MRSA in hunting dogs in Maiduguri metropolitan. Materials and Methods: Phenotypic methods used includes microscopic technique, colony morphology study, catalase-coagulase tests, and the use of mannitol salt agar test, oxacillin resistance screening agar base, and antibiotic susceptibility testing methods. Genotypic approach was used for deoxyribonucleic acid extraction, and the presence of nuc and mecA gene was detected using polymerase chain reaction (PCR techniques. Results: Examination of 416 swab samples from nasal and perineal region of dogs revealed a total of 79.5% of S. aureus, where 62.5% of the isolates were MRSA. Molecular analysis revealed that 7nuc genes specific for S. aureus from 20 presumptive MRSA assay were all mecA PCR negative. The isolates were sensitive to gentamicin and ciprofloxacin but proved resistant to cefoxitin and oxacillin. Conclusion: High isolation rate of MRSA was found in hunting dogs. Significant level (p<0.05 of MRSA was isolated in the nasal cavity of hunting dogs than its perineum. Only nuc genes were detected from the MRSA isolates.

  6. Linking microbial oxidation of arsenic with detection and phylogenetic analysis of arsenite oxidase genes in diverse geothermal environments.

    Science.gov (United States)

    Hamamura, N; Macur, R E; Korf, S; Ackerman, G; Taylor, W P; Kozubal, M; Reysenbach, A-L; Inskeep, W P

    2009-02-01

    The identification and characterization of genes involved in the microbial oxidation of arsenite will contribute to our understanding of factors controlling As cycling in natural systems. Towards this goal, we recently characterized the widespread occurrence of aerobic arsenite oxidase genes (aroA-like) from pure-culture bacterial isolates, soils, sediments and geothermal mats, but were unable to detect these genes in all geothermal systems where we have observed microbial arsenite oxidation. Consequently, the objectives of the current study were to measure arsenite-oxidation rates in geochemically diverse thermal habitats in Yellowstone National Park (YNP) ranging in pH from 2.6 to 8, and to identify corresponding 16S rRNA and aroA genotypes associated with these arsenite-oxidizing environments. Geochemical analyses, including measurement of arsenite-oxidation rates within geothermal outflow channels, were combined with 16S rRNA gene and aroA functional gene analysis using newly designed primers to capture previously undescribed aroA-like arsenite oxidase gene diversity. The majority of bacterial 16S rRNA gene sequences found in acidic (pH 2.6-3.6) Fe-oxyhydroxide microbial mats were closely related to Hydrogenobaculum spp. (members of the bacterial order Aquificales), while the predominant sequences from near-neutral (pH 6.2-8) springs were affiliated with other Aquificales including Sulfurihydrogenibium spp., Thermocrinis spp. and Hydrogenobacter spp., as well as members of the Deinococci, Thermodesulfobacteria and beta-Proteobacteria. Modified primers designed around previously characterized and newly identified aroA-like genes successfully amplified new lineages of aroA-like genes associated with members of the Aquificales across all geothermal systems examined. The expression of Aquificales aroA-like genes was also confirmed in situ, and the resultant cDNA sequences were consistent with aroA genotypes identified in the same environments. The aroA sequences

  7. High-resolution melting analysis for detection of a single-nucleotide polymorphism and the genotype of the myostatin gene in warmblood horses.

    Science.gov (United States)

    Serpa, Priscila B S; Garbade, Petra; Natalini, Cláudio C; Pires, Ananda R; Tisotti, Tainor M

    2017-01-01

    OBJECTIVE To develop a high-resolution melting (HRM) assay to detect the g.66493737C>T polymorphism in the myostatin gene (MSTN) and determine the frequency of 3 previously defined g.66493737 genotypes (T/T, T/C, and C/C) in warmblood horses. SAMPLES Blood samples from 23 horses. PROCEDURES From each blood sample, DNA was extracted and analyzed by standard PCR methods and an HRM assay to determine the MSTN genotype. Three protocols (standard protocol, protocol in which a high-salt solution was added to the reaction mixture before the first melting cycle, and protocol in which an unlabeled probe was added to the reaction mixture before analysis) for the HRM assay were designed and compared. Genotype results determined by the HRM protocol that generated the most consistent melting curves were compared with those determined by sequencing. RESULTS The HRM protocol in which an unlabeled probe was added to the reaction mixture generated the most consistent melting curves. The genotypes of the g.66493737C>T polymorphism were determined for 22 horses (16 by HRM analysis and 20 by sequencing); 14, 7, and 1 had the T/T, T/C, and C/C genotypes, respectively. The genotype determined by HRM analysis agreed with that determined by sequencing for 14 of 16 horses. The frequency of alleles T and C was 79.5% and 20.5%, respectively. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that HRM analysis may be a faster and more economical alternative than PCR methods for genotyping. Genotyping results might be useful as predictors of athletic performance for horses.

  8. Development of a real-time RT-PCR and Reverse Line probe Hybridisation assay for the routine detection and genotyping of Noroviruses in Ireland.

    LENUS (Irish Health Repository)

    Menton, John F

    2007-01-01

    BACKGROUND: Noroviruses are the most common cause of non-bacterial gastroenteritis. Improved detection methods have seen a large increase in the number of human NoV genotypes in the last ten years. The objective of this study was to develop a fast method to detect, quantify and genotype positive NoV samples from Irish hospitals. RESULTS: A real-time RT-PCR assay and a Reverse Line Blot Hybridisation assay were developed based on the ORF1-ORF2 region. The sensitivity and reactivity of the two assays used was validated using a reference stool panel containing 14 NoV genotypes. The assays were then used to investigate two outbreaks of gastroenteritis in two Irish hospitals. 56 samples were screened for NoV using a real-time RT-PCR assay and 26 samples were found to be positive. Genotyping of these positive samples found that all positives belonged to the GII\\/4 variant of NoV. CONCLUSION: The combination of the Real-time assay and the reverse line blot hybridisation assay provided a fast and accurate method to investigate a NoV associated outbreak. It was concluded that the predominant genotype circulating in these Irish hospitals was GII\\/4 which has been associated with the majority of NoV outbreaks worldwide. The assays developed in this study are useful tools for investigating NoV infection.

  9. Detection and genotyping of human papillomavirus in self-obtained cervicovaginal samples by using the FTA cartridge: new possibilities for cervical cancer screening.

    NARCIS (Netherlands)

    Lenselink, C.H.; Bie, R.P. de; Hamont, D. van; Bakkers, J.M.J.E.; Quint, W.G.V.; Massuger, L.F.A.G.; Bekkers, R.L.M.; Melchers, W.J.G.

    2009-01-01

    This study assesses human papillomavirus (HPV) detection and genotyping in self-sampled genital smears applied to an indicating FTA elute cartridge (FTA cartridge). The study group consisted of 96 women, divided into two sample sets. All samples were analyzed by the HPV SPF(10)-Line Blot 25. Set 1

  10. Differential detection of Human Papillomavirus genotypes and cervical intraepithelial neoplasia by four commercial assays

    DEFF Research Database (Denmark)

    Rebolj, Matejka; Bonde, Jesper; Preisler, Sarah;

    2016-01-01

    Laboratories can nowadays choose from >100 Human Papillomavirus (HPV) assays for cervical screening. Our previous analysis based on the data from the Danish Horizon study, however, showed that four widely used assays, Hybrid Capture 2 (HC2), cobas, CLART and APTIMA, frequently do not detect...... the same HPV infections. Here, we determined the characteristics of the concordant (all four assays returning a positive HPV test result) and discordant samples (all other HPV-positive samples) in primary cervical screening at 30-65 years (n=2859) and in a concurrent referral population from the same...... catchment area (n=885). HPV testing followed the manufacturers' protocols. Women with abnormal cytology were managed according to the routine recommendations. Cytology-normal/HPV-positive women were invited for repeated testing in 18 months. Screening history and histologically confirmed cervical...

  11. Evaluation of Genotypic and Phenotypic Methods to Detect Carbapenemase Production in Gram-Negative Bacilli.

    Science.gov (United States)

    McMullen, Allison R; Yarbrough, Melanie L; Wallace, Meghan A; Shupe, Angela; Burnham, Carey-Ann D

    2017-03-01

    Carbapenemase-producing gram-negative bacteria (CP-GNB) are an urgent and expanding public health threat. Rapid and accurate identification of these organisms facilitates infection prevention efforts in healthcare facilities. The objective of our study was to evaluate methods to detect and identify CP-GNB. We examined 189 carbapenem-resistant GNB(CR-GNB), including Enterobacteriaceae, Pseudomonas aeruginosa, and Acinetobacter baumannii complex, using 3 different methods: 2 methods to screen isolates of GNB for carbapenemase production [the carbapenem inactivation method (CIM) and 2 chromogenic agars] and a molecular method (Cepheid GeneXpert Carba-R) to identify the mechanism of carbapenem resistance and the associated resistance genes (blaKPC, blaNDM, blaIMP, blaOXA-48-like, and blaVIM). The CIM was a simple and inexpensive phenotypic screen to differentiate between CR-GNB and CP-GNB, with improved analytical performance characteristics and inter-reader correlation compared to the modified Hodge test. Both chromogenic agars evaluated (HardyCHROM CRE and chromID CARBA) were able to support growth of most of the organisms tested, including isolates possessing the blaOXA-48-like gene. However, these media had a low analytical specificity for carbapenemase production, with breakthrough of CR-GNB that did not produce a carbapenemase. The Xpert Carba-R assay was rapid and easy to perform, and demonstrated 100% positive and negative agreement for characterization of genetic determinants of carbapenem resistance. Screening by CIM followed by the Xpert Carba-R PCR is an accurate method for detecting and characterizing CP-GNB, including Enterobacteriaceae, P. aeruginosa, and A. baumannii complex. © 2016 American Association for Clinical Chemistry.

  12. Hepatitis C Virus Genotypes

    Directory of Open Access Journals (Sweden)

    Kayhan Azadmanesh

    2005-09-01

    the ultimate source of the virus's genetic diversity. HCV circulates as a heterogeneous population of genetically different but closely related genomes known as the quasispecies(15.As only 30-35% of nucleotides actually differ, there is obviously considerable heterogeneity in evolutionary rates among nucleotide sites in the genome. This heterogeneity is the result of variable evolutionary constraints. The 5'-UTR contains extensive secondary RNA structure and is correspondingly the slowest evolving genomic region(16. The next slowest region is the C (Core gene, which evolves three times faster than the 5'- UTR. The envelope genes E1 and E2 constitute the most diverse genome region and evolve about nine times faster than the 5'-UTR(16, probably as a result of their presumed role in evading the host immune response. Genomic Heterogeneity and ClassificationSystemsShortly after its discovery in 1989, it became clear that HCV had substantial nucleotide sequence diversity, with only 66 to 80% overall sequencesimilarity among strains belonging to different genotypes or subtypes(17. HCV isolates show four levels of genomic variations: types, subtypes, isolates, andquasispecies. The overall sequence similarities over complete genomic sequences are at least 91% within quasispecies, approximately 79% (range, 77 to 80% between subtypes, and about 68% (range, 66 to 69% between different types. This quasispecies is composed of a group of heterogeneous RNA sequences centered around a dominant nucleotide sequence that changes, throughout the course of the infection, under the selective pressure of the host immune system(18. More than one genotype can be found in the circulations of some HCV-infected patients, particularly in individuals who have received multiple transfusions and intravenous drug users. These are referred to as mixed-genotype infections(19, 20.The lack of a routinely available cell culture system and an easily available animal model has rendered classification of HCV

  13. Extended Genetic Diversity of Bovine Viral Diarrhea Virus and Frequency of Genotypes and Subtypes in Cattle in Italy between 1995 and 2013

    Directory of Open Access Journals (Sweden)

    Camilla Luzzago

    2014-01-01

    Full Text Available Genetic typing of bovine viral diarrhea virus (BVDV has distinguished BVDV-1 and BVDV-2 species and an emerging putative third species (HoBi-like virus, recently detected in southern Italy, signaling the occurrence of natural infection in Europe. Recognizing the need to update the data on BVDV genetic variability in Italy for mounting local and European alerts, a wide collection of 5′ UTR sequences (n = 371 was selected to identify the frequency of genotypes and subtypes at the herd level. BVDV-1 had the highest frequency, followed by sporadic BVDV-2. No novel HoBi-like viruses were identified. Four distribution patterns of BVDV-1 subtypes were observed: highly prevalent subtypes with a wide temporal-spatial distribution (1b and 1e, low prevalent subtypes with a widespread geographic distribution (1a, 1d, 1g, 1h, and 1k or a restricted geographic distribution (1f, and sporadic subtypes detected only in single herds (1c, 1j, and 1l. BVDV-1c, k, and l are reported for the first time in Italy. A unique genetic variant was detected in the majority of herds, but cocirculation of genetic variants was also observed. Northern Italy ranked first for BVDV introduction, prevalence, and dispersion. Nevertheless, the presence of sporadic variants in other restricted areas suggests the risk of different routes of BVDV introduction.

  14. Extended genetic diversity of bovine viral diarrhea virus and frequency of genotypes and subtypes in cattle in Italy between 1995 and 2013.

    Science.gov (United States)

    Luzzago, Camilla; Lauzi, Stefania; Ebranati, Erika; Giammarioli, Monica; Moreno, Ana; Cannella, Vincenza; Masoero, Loretta; Canelli, Elena; Guercio, Annalisa; Caruso, Claudio; Ciccozzi, Massimo; De Mia, Gian Mario; Acutis, Pier Luigi; Zehender, Gianguglielmo; Peletto, Simone

    2014-01-01

    Genetic typing of bovine viral diarrhea virus (BVDV) has distinguished BVDV-1 and BVDV-2 species and an emerging putative third species (HoBi-like virus), recently detected in southern Italy, signaling the occurrence of natural infection in Europe. Recognizing the need to update the data on BVDV genetic variability in Italy for mounting local and European alerts, a wide collection of 5' UTR sequences (n = 371) was selected to identify the frequency of genotypes and subtypes at the herd level. BVDV-1 had the highest frequency, followed by sporadic BVDV-2. No novel HoBi-like viruses were identified. Four distribution patterns of BVDV-1 subtypes were observed: highly prevalent subtypes with a wide temporal-spatial distribution (1b and 1e), low prevalent subtypes with a widespread geographic distribution (1a, 1d, 1g, 1h, and 1k) or a restricted geographic distribution (1f), and sporadic subtypes detected only in single herds (1c, 1j, and 1l). BVDV-1c, k, and l are reported for the first time in Italy. A unique genetic variant was detected in the majority of herds, but cocirculation of genetic variants was also observed. Northern Italy ranked first for BVDV introduction, prevalence, and dispersion. Nevertheless, the presence of sporadic variants in other restricted areas suggests the risk of different routes of BVDV introduction.

  15. Performance analysis of fiber-based free-space optical communications with coherent detection spatial diversity.

    Science.gov (United States)

    Li, Kangning; Ma, Jing; Tan, Liying; Yu, Siyuan; Zhai, Chao

    2016-06-10

    The performances of fiber-based free-space optical (FSO) communications over gamma-gamma distributed turbulence are studied for multiple aperture receiver systems. The equal gain combining (EGC) technique is considered as a practical scheme to mitigate the atmospheric turbulence. Bit error rate (BER) performances for binary-phase-shift-keying-modulated coherent detection fiber-based free-space optical communications are derived and analyzed for EGC diversity receptions through an approximation method. To show the net diversity gain of a multiple aperture receiver system, BER performances of EGC are compared with a single monolithic aperture receiver system with the same total aperture area (same average total incident optical power on the aperture surface) for fiber-based free-space optical communications. The analytical results are verified by Monte Carlo simulations. System performances are also compared for EGC diversity coherent FSO communications with or without considering fiber-coupling efficiencies.

  16. Development and Evaluation of a Seminested PCR for Detection and Differentiation of Babesia gibsoni (Asian Genotype) and B. canis DNA in Canine Blood Samples

    OpenAIRE

    Birkenheuer, Adam J.; Levy, Michael G.; Breitschwerdt, Edward B.

    2003-01-01

    Canine babesiosis has recently been recognized as an emerging infectious disease of dogs in North America. We sought to develop a seminested PCR to detect and differentiate Babesia gibsoni (Asian genotype), B. canis subsp. vogeli, B. canis subsp. canis, and B. canis subsp. rossi DNA in canine blood samples. An outer primer pair was designed to amplify an ∼340-bp fragment of the 18S rRNA genes from B. gibsoni (Asian genotype), B. canis subsp. vogeli, B. canis subsp. rossi, and B. canis subsp. ...

  17. Phenotypic and Genotypic Detection of Metallo-beta-lactamases among Imipenem-Resistant Gram Negative Isolates

    Directory of Open Access Journals (Sweden)

    Mohammad Mohammadzadeh

    2016-08-01

    Full Text Available Background:   Imipenem-resistant gram negative bacteria, resulting from metallo-beta-lactamase (MBLs-producing strains have been reported to be among the important causes of nosocomial infections and of serious therapeutic problem worldwide. Because of their broad range, potent carbapenemase activity and resistance to inhibitors, these enzymes can confer resistance to almost all beta-lactams. The prevalence of metallo-beta-lactamase among imipenem-resistant Acinetobacter spp., Pseudomonas spp. and Enerobacteriaceae isolates is determined.Methods:   In this descriptive study 864 clinical isolates of Acinetobacter spp., Pseudomonas spp. and Enterobacteriaceae, were initially tested for imipenem susceptibility. The metallo-beta-lactamase production was detected using combined disk diffusion, double disk synergy test, and Hodge test. Then all imipenem resistant isolates were tested by PCR for imp, vim and ndm genes. Results:   Among 864 isolates, 62 (7.17 % were imipenem-resistant. Positive phonetypic test for metallo-beta-lactamase was 40 (64.5%, of which 24 (17.1% and 16 (9.2% isolates were Acinetobacter spp. and Pseudomonas spp., respectively. By PCR method 30 (48.4% of imipenem resistant Acinetobacter, and Pseudomonas isolates were positive for MBL-producing genes. None of the Enterobacteriaceae isolates were positive for metallo-beta-lactamase activity. Conclusion:   The results of this study are indicative of the growing number of nosocomial infections associated with multidrug-resistant gram negative bacteria in this region leading to difficulties in antibiotic therapy. Thereby, using of phenotypic methods can be helpful for management of this problem.

  18. Familial Case of Pelizaeus-Merzbacher Disorder Detected by Oligoarray Comparative Genomic Hybridization: Genotype-to-Phenotype Diagnosis

    Directory of Open Access Journals (Sweden)

    Kimia Najafi

    2017-01-01

    Full Text Available Introduction. Pelizaeus-Merzbacher disease (PMD is an X-linked recessive hypomyelinating leukodystrophy characterized by nystagmus, spastic quadriplegia, ataxia, and developmental delay. It is caused by mutation in the PLP1 gene. Case Description. We report a 9-year-old boy referred for oligoarray comparative genomic hybridization (OA-CGH because of intellectual delay, seizures, microcephaly, nystagmus, and spastic paraplegia. Similar clinical findings were reported in his older brother and maternal uncle. Both parents had normal phenotypes. OA-CGH was performed and a 436 Kb duplication was detected and the diagnosis of PMD was made. The mother was carrier of this 436 Kb duplication. Conclusion. Clinical presentation has been accepted as being the mainstay of diagnosis for most conditions. However, recent developments in genetic diagnosis have shown that, in many congenital and sporadic disorders lacking specific phenotypic manifestations, a genotype-to-phenotype approach can be conclusive. In this case, a diagnosis was reached by universal genomic testing, namely, whole genomic array.

  19. Genetic diversity in Australian Cedar genotypes selected by mixed models Diversidade genética em genótipos de Cedro Australiano selecionados via modelos mistos

    Directory of Open Access Journals (Sweden)

    Rulfe Tavares

    2012-04-01

    Full Text Available The increasing demand for raw material for multiple uses of forest products and by-products has attracted the interest for fast growing species, such as the Australian Cedar (Toona ciliata, which presents high productive and economic potential. The present work aimed at estimating genetic diversity by DNA markers and morphological traits supported for the mixed models. The following traits were measured and genotypes were sampled randomly in different areas: diameter at breast height, height, cylindrical volume, diameter, distance between nodes and crown diameter. Twelve RAPD primers were used and generated a total of 91 marks, 82 of which were polymorphic. The high percentage of polymorphic markers, 90.10%, demonstrated that discrimination in this species is efficient, but it yet little studied, for this case we can find the extent of the genetic basis for the application of technical improvement. The assessment of genetic diversity by the UPGMA method using the binary and morphological data provided the expression of genetic dissimilarities among the accessions evaluated, optimizing the perception of this divergence. The use of mixed models was efficient to assess combined genetic diversity to optimize the selection of genotypes with divergent genetic values for diameter at breast height.A crescente demanda por matéria-prima para múltiplos usos dos produtos florestais e subprodutos tem despertado o interesse para espécies de crescimento rápido, como o cedro australiano (Toona ciliata, que apresenta potencial produtivo e econômico. Neste trabalho, objetivou-se estimar a diversidade genética por marcadores de DNA e caracteres morfológicos com o uso dos modelos mistos. Os seguintes caracteres foram medidos e os genótipos foram amostrados aleatoriamente em diferentes áreas de plantio: diâmetro à altura do peito, altura, volume cilíndrico, distância entre nós e diâmetro de copa. Doze primers RAPD foram utilizados e geraram um total

  20. Comparison Between AFLP and RFLP Markers in Detecting the Diversity of Rice (Oryza sativa L.)

    Institute of Scientific and Technical Information of China (English)

    CHEN Liang; LIANG Chun-yang; SUN Chuan-qing; JIN De-min; JIANG Ting-bo; WANG Bin; WANG Xiang-kun

    2002-01-01

    AFLP and RFLP were used to study the diversity of 20 rice cultivars. 15 primer combinations were used in the AFLP analysis and 47 - 118 bands were amplified in each lane. A total of 107 polymorphic bands were detected in the RFLP analysis using 49 RFLP probes. The cluster analysis based on RFLP data showed that 20 rice cultivars could be divided into an indica group and a japonica group, as did the AFLP data; however AFLP is more suitable in detecting the difference of pedigree and ecotype among the 20 cultivars.The genetic distance based on AFLP and RFLP data showed the same tendency, but AFLP markers increased the measure of genetic distance among intra-subspecific cultivars and decreased the measure of genetic distance among inter-subspecific cultivars relative to RFLP markers. That indicated that AFLP is more suitable than RFLP in the diversity study and RFLP is more suitable to study the indica-japonica differentiation of cultivars.

  1. Detection of neonatal unit clusters of Candida parapsilosis fungaemia by microsatellite genotyping: Results from laboratory-based sentinel surveillance, South Africa, 2009-2010.

    Science.gov (United States)

    Magobo, Rindidzani E; Naicker, Serisha D; Wadula, Jeannette; Nchabeleng, Maphoshane; Coovadia, Yacoob; Hoosen, Anwar; Lockhart, Shawn R; Govender, Nelesh P

    2017-05-01

    Neonatal candidaemia is a common, deadly and costly hospital-associated disease. To determine the genetic diversity of Candida parapsilosis causing fungaemia in South African neonatal intensive care units (NICUs). From February 2009 through to August 2010, cases of candidaemia were reported through laboratory-based surveillance. C. parapsilosis isolates from neonatal cases were submitted for identification by internal transcribed spacer (ITS) region sequencing, antifungal susceptibility testing and microsatellite genotyping. Cluster analysis was performed using Unweighted Pair Group Method with Arithmetic Mean (UPGMA). Of 1671 cases with a viable Candida isolate, 393 (24%) occurred among neonates. Isolates from 143 neonatal cases were confirmed as C. parapsilosis sensu stricto. Many isolates were resistant to fluconazole (77/143; 54%) and voriconazole (20/143; 14%). Of 79 closely-related genotypes, 18 were represented by ≥2 isolates; 61 genotypes had a single isolate each. Seven clusters, comprised of 82 isolates, were identified at five hospitals in three provinces. Isolates belonging to certain clusters were significantly more likely to be fluconazole resistant: all cluster 7 isolates and the majority of cluster 4 (78%), 5 (89%) and 6 (67%) isolates (PCandida parapsilosis-associated candidaemia in public-sector NICUs was caused by closely related genotypes and there was molecular evidence of undetected outbreaks as well as intra-hospital transmission. © 2017 Blackwell Verlag GmbH.

  2. Development and evaluation of a seminested PCR for detection and differentiation of Babesia gibsoni (Asian genotype) and B. canis DNA in canine blood samples.

    Science.gov (United States)

    Birkenheuer, Adam J; Levy, Michael G; Breitschwerdt, Edward B

    2003-09-01

    Canine babesiosis has recently been recognized as an emerging infectious disease of dogs in North America. We sought to develop a seminested PCR to detect and differentiate Babesia gibsoni (Asian genotype), B. canis subsp. vogeli, B. canis subsp. canis, and B. canis subsp. rossi DNA in canine blood samples. An outer primer pair was designed to amplify an approximately 340-bp fragment of the 18S rRNA genes from B. gibsoni (Asian genotype), B. canis subsp. vogeli, B. canis subsp. rossi, and B. canis subsp. canis but not mammalian DNA. Forward primers were designed that would specifically amplify a smaller fragment from each organism in a seminested PCR. The practical limit of detection was 50 organisms/ml of mock-infected EDTA anticoagulated whole blood. The primer pair also amplified an approximately 370-bp fragment of the B. gibsoni (USA/California genotype) 18S rRNA gene from the blood of an experimentally infected dog with a high percentage of parasitemia. Amplicons were not detected when DNA extracted from the blood of a dog that was naturally infected with Theileria annae at a low percentage of parasitemia was amplified. Due to limited sensitivity, this test is not recommended for the routine diagnosis of B. gibsoni (USA/California genotype) or T. annae. The PCR test did not amplify Toxoplasma gondii, Neospora caninum, Leishmania infantum, Cryptosporidium parvum, or canine DNA under any of the conditions tested. The seminested PCR test was able to detect and discriminate B. gibsoni (Asian genotype), B. canis subsp. vogeli, B. canis subsp. canis, and B. canis subsp. rossi DNA in blood samples from infected dogs.

  3. COLD-PCR and innovative microarray substrates for detecting and genotyping MPL exon 10 W515 substitutions.

    Science.gov (United States)

    Brisci, Angela; Damin, Francesco; Pietra, Daniela; Galbiati, Silvia; Boggi, Sabrina; Casetti, Ilaria; Rumi, Elisa; Chiari, Marcella; Cazzola, Mario; Ferrari, Maurizio; Cremonesi, Laura

    2012-12-01

    Myeloproliferative neoplasms (MPNs) include polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF). Somatic mutations in exon 10 of the MPL (myeloproliferative leukemia virus oncogene) gene, mainly substitutions encoding W515 variants, have recently been described in a minority of patients with ET or PMF. We optimized analytically sensitive methods for detecting and genotyping MPL variants. We used DNA previously isolated from circulating granulocytes of 60 patients with MPN that had previously been analyzed by high-resolution melting (HRM), direct sequencing, and the TaqMan allelic-discrimination assay. We developed conditions for enriching tumor mutant alleles with COLD-PCR (coamplification at lower denaturation temperature PCR) and coupled it with direct sequencing. Assays were designed for identifying MPL W515 substitutions with full COLD-PCR protocols. In parallel, we used innovative microarray substrates to develop assays for evaluating the mutant burden in granulocyte cells. Mutations that were present at very low levels in patients who had previously been scored as having an MPL variant by HRM and as wild type by direct sequencing were successfully identified in granulocyte DNA. Notably, the microarray approach displayed analytical sensitivities of 0.1% to 5% mutant allele, depending on the particular mutation. This analytical sensitivity is similar to that obtained with COLD-PCR. The assay requires no enrichment strategy and allows both the characterization of each variant allele and the evaluation of its proportion in every patient. These procedures, which are transferable to clinical diagnostic laboratories, can be used for detecting very low proportions of minority mutant alleles that cannot be identified by other, conventional methods. © 2012 American Association for Clinical Chemistry

  4. Highly multiplex and sensitive SNP genotyping method using a three-color fluorescence-labeled ligase detection reaction coupled with conformation-sensitive CE.

    Science.gov (United States)

    Choi, Woong; Jung, Gyoo Yeol

    2017-02-01

    For the development of clinically useful genotyping methods for SNPs, accuracy, simplicity, sensitivity, and cost-effectiveness are the most important criteria. Among the methods currently being developed for SNP genotyping technology, the ligation-dependent method is considered the simplest for clinical diagnosis. However, sensitivity is not guaranteed by the ligation reaction alone, and analysis of multiple targets is limited by the detection method. Although CE is an attractive alternative to error-prone hybridization-based detection, the multiplex assay process is complicated because of the size-based DNA separation principle. In this study, we employed the ligase detection reaction coupled with high-resolution CE-SSCP to develop an accurate, sensitive, and simple multiplex genotyping method. Ligase detection reaction could amplify ligated products through recurrence of denaturation and ligation reaction, and SSCP could separate these products according to each different structure conformation without size variation. Thus, simple and sensitive SNP analysis can be performed using this method involving the use of similar-sized probes, without complex probe design steps. We found that this method could not only accurately discriminate base mismatches but also quantitatively detect 37 SNPs of the tp53 gene, which are used as targets in multiplex analysis, using three-color fluorescence-labeled probes.

  5. Genome-wide high-throughput SNP discovery and genotyping for understanding natural (functional) allelic diversity and domestication patterns in wild chickpea

    Science.gov (United States)

    Bajaj, Deepak; Das, Shouvik; Badoni, Saurabh; Kumar, Vinod; Singh, Mohar; Bansal, Kailash C.; Tyagi, Akhilesh K.; Parida, Swarup K.

    2015-01-01

    We identified 82489 high-quality genome-wide SNPs from 93 wild and cultivated Cicer accessions through integrated reference genome- and de novo-based GBS assays. High intra- and inter-specific polymorphic potential (66–85%) and broader natural allelic diversity (6–64%) detected by genome-wide SNPs among accessions signify their efficacy for monitoring introgression and transferring target trait-regulating genomic (gene) regions/allelic variants from wild to cultivated Cicer gene pools for genetic improvement. The population-specific assignment of wild Cicer accessions pertaining to the primary gene pool are more influenced by geographical origin/phenotypic characteristics than species/gene-pools of origination. The functional significance of allelic variants (non-synonymous and regulatory SNPs) scanned from transcription factors and stress-responsive genes in differentiating wild accessions (with potential known sources of yield-contributing and stress tolerance traits) from cultivated desi and kabuli accessions, fine-mapping/map-based cloning of QTLs and determination of LD patterns across wild and cultivated gene-pools are suitably elucidated. The correlation between phenotypic (agromorphological traits) and molecular diversity-based admixed domestication patterns within six structured populations of wild and cultivated accessions via genome-wide SNPs was apparent. This suggests utility of whole genome SNPs as a potential resource for identifying naturally selected trait-regulating genomic targets/functional allelic variants adaptive to diverse agroclimatic regions for genetic enhancement of cultivated gene-pools. PMID:26208313

  6. Complete Genome Sequencing of Influenza A Viruses within Swine Farrow-to-Wean Farms Reveals the Emergence, Persistence, and Subsidence of Diverse Viral Genotypes.

    Science.gov (United States)

    Diaz, Andres; Marthaler, Douglas; Culhane, Marie; Sreevatsan, Srinand; Alkhamis, Moh; Torremorell, Montserrat

    2017-09-15

    Influenza A viruses (IAVs) are endemic in swine and represent a public health risk. However, there is limited information on the genetic diversity of swine IAVs within farrow-to-wean farms, which is where most pigs are born. In this longitudinal study, we sampled 5 farrow-to-wean farms for a year and collected 4,190 individual nasal swabs from three distinct pig subpopulations. Of these, 207 (4.9%) samples tested PCR positive for IAV, and 124 IAVs were isolated. We sequenced the complete genomes of 123 IAV isolates and found 31 H1N1, 26 H1N2, 63 H3N2, and 3 mixed IAVs. Based on the IAV hemagglutinin, seven different influenza A viral groups (VGs) were identified. Most of the remaining IAV gene segments allowed us to differentiate the same VGs, although an additional viral group was identified for gene segment 3 (PA). Moreover, the codetection of more than one IAV VG was documented at different levels (farm, subpopulation, and individual pigs), highlighting the environment for potential IAV reassortment. Additionally, 3 out of 5 farms contained IAV isolates (n = 5) with gene segments from more than one VG, and 79% of all the IAVs sequenced contained a signature mutation (S31N) in the matrix gene that has been associated with resistance to the antiviral amantadine. Within farms, some IAVs were detected only once, while others were detected for 283 days. Our results illustrate the maintenance and subsidence of different IAVs within swine farrow-to-wean farms over time, demonstrating that pig subpopulation dynamics are important to better understand the diversity and epidemiology of swine IAVs.IMPORTANCE On a global scale, swine are one of the main reservoir species for influenza A viruses (IAVs) and play a key role in the transmission of IAVs between species. Additionally, the 2009 IAV pandemics highlighted the role of pigs in the emergence of IAVs with pandemic potential. However, limited information is available regarding the diversity and distribution of swine IAVs

  7. [Evaluation of rapid genotype assay for the identification of gram-positive cocci from blood cultures and detection of mecA and van genes].

    Science.gov (United States)

    Gülhan, Barış; Atmaca, Selahattin; Ozekinci, Tuncer; Suay, Adnan

    2011-10-01

    Rapid and accurate identification of bacterial pathogens grown in blood cultures of patients with sepsis is crucial for prompt initiation of appropriate therapy in order to decrease related morbidity and mortality rates. Although current automated blood culture systems led to a significant improvement in bacterial detection time, more rapid identification systems are still needed to optimise the establishment of treatment. Novel genotype technology which is developed for the rapid diagnosis of sepsis, is a molecular genetic assay based on DNA multiplex amplification with biotinylated primers followed by hybridization to membrane bound probes. The aim of this study was to evaluate the performance of "Genotype® BC gram-positive” test for the identification of gram-positive cocci grown in blood cultures and rapid detection of mecA and van genes. This test uses DNA.STRIP® technology which includes a panel of probes for identification of 17 gram-positive bacterial species and is able to determinate the methicillin and vancomycin resistance mediating genes (mecA and vanA, vanB, vanC1, vanC2/C3) simultaneously, in a single test run. A total of 55 positive blood cultures from BACTECTM Plus/F (Becton Dickinson, USA) aerobic and pediatric blood culture vials were included in the study. The isolates which exhibit gram-positive coccus morphology by Gram staining were identified by Genotype ® BC gram-positive test (Hain Life Science, Germany). All of the samples were also identified with the use of Phoenix PMIC/ID Panel (Becton Dickinson, USA) and antibiotic susceptibilities were determined. Of the 55 blood culture isolates, 17 were identified as Staphylococcus epidermidis [all were methicillin-resistant (MR)], 9 were S.aureus (one was MR), 18 were S.hominis (10 were MR), 4 were E.faecalis, 3 were E. faecium (one was vanconycin-resistant), 2 were S.saprophyticus (one was MR), 1 was S.warneri and 1 was S.haemolyticus, by Phoenix automated system. Genotype® BC gram

  8. Prevalence of Hepatitis C Virus Genotypes in Asymptomatic Blood Donors in NCT of Delhi

    OpenAIRE

    2015-01-01

    Hepatitis C virus shows high genomic diversity which poses a challenge for drug therapy and vaccine development. Here, we present a study on prevalence of HCV genotypes and subtypes in blood donors of Delhi. Forty two HCV samples were taken from blood bags found positive at various blood banks located in Delhi. From these samples 5’UTR were amplified, sequenced and genotyped. Genotype 1, 3 and 4 were detected but genotype 1 of HCV was found prevalent. Based on sequence analysis the HCV isolat...

  9. Diversity detection in non-Gaussian noise employing the generalized approach to signal processing in noise with fading diversity channels

    Science.gov (United States)

    Tuzlukov, Vyacheslav

    2011-06-01

    In this paper, we consider the problem of M-ary signal detection based on the generalized approach to signal processing (GASP) in noise over a single-input multiple-output (SIMO) channel affected by frequency-dispersive Rayleigh distributed fading and corrupted by additive non-Gaussian noise modeled as spherically invariant random process. We derive both the optimum generalized detector (GD) structure based on GASP and a suboptimal reduced-complexity GD applying the low energy coherence approach jointly with the GASP in noise. Both GD structures are independent of the actual noise statistics. We also carry out a performance analysis of both GDs and compare with the conventional receivers. The performance analysis is carried out with reference to the case that the channel is affected by a frequency-selective fading and for a binary frequency-shift keying (BFSK) signaling format. The results obtained through both a Chernoff-bounding technique and Monte Carlo simulations reveal that the adoption of diversity also represents a suitable means to restore performance in the presence of dispersive fading and impulsive non-Gaussian noise. It is also shown that the suboptimal GD incurs a limited loss with respect to the optimum GD and this loss is less in comparison with the conventional receiver.

  10. Characterization of Enterococcus faecium with macrolide resistance and reduced susceptibility to quinupristin/dalfopristin in a Japanese hospital: detection of extensive diversity in erm(B)-regulator regions.

    Science.gov (United States)

    Isogai, Nayuta; Urushibara, Noriko; Kawaguchiya, Mitsuyo; Ghosh, Souvik; Suzaki, Keisuke; Watanabe, Naoki; Quiñones, Dianelys; Kobayashi, Nobumichi

    2013-08-01

    Cross-resistance to macrolide, lincosamide, and streptogramin B (MLSB) antibiotics is mainly mediated by the erm (erythromycin ribosome methylation) genes that encode 23S rRNA methylases in enterococi, and various mechanisms are involved in the streptogramin B resistance. Prevalence of MLSB resistance and its genetic mechanisms were analyzed for a total of 159 strains of Enterococcus faecium isolated from clinical specimens in a university hospital in Japan from 1997 to 2006. Resistance to erythromycin (EM) and clindamycin was detected in 88.1% and 89.9% of all the strains examined, respectively, and expression of resistance was totally constitutive. Although none of the strain was resistant to quinupristin/dalfopristin (Q/D), 28 strains (17.6%) showed intermediate resistance to Q/D (MIC: 2 μg/ml). The erm(B) gene was detected in 139 strains (87.4%), and msrC was found in all the strains examined, whereas no other known MLSB resistance genes were identified. The erm(B) regulator region (RR) containing a coding region of the leader peptide was classified into 13 genetic variations (L1-L3, M, S1-S7, D, and R genotypes) in 56 strains. However, no relatedness was identified between the erm(B) RR genotype and EM resistance, or reduced susceptibility to Q/D, although most of Q/D-intermediate strains were assigned to the L1, L2, and S1 genotypes. Q/D-intermediate strains were classified into five multiple-locus variable-number tandem-repeat analysis (MLVA) types, including four types of clonal complex (CC)-C1, five sequence types (STs), including four STs of CC-17, and several resistance gene/virulence factor profiles. The present study revealed the occurrence of Q/D-intermediate E. faecium, which are composed of heterogeneous strains in Japan, and more genetic diversity in the erm(B) RRs than those reported previously.

  11. Development of New InDel Marker to Detect Genotypes of Rf-1a Conferring Fertility Restoration of BT-Type Cytoplasmic Male Sterility in Rice

    Institute of Scientific and Technical Information of China (English)

    CHEN Tao; ZHANG Ya-dong; ZHU Zhen; ZHAO Ling; ZHAO Qing-yong; ZHOU Li-hui; YAO Shu; YU Xin; WANG Cai-lin

    2014-01-01

    Restorer line breeding is an important approach to enhance the heterosis and improve the yields of japonica hybrid rice. To improve the selection efficiency of restorer lines for BT-type cytoplasmic male sterility (CMS) in japonica rice, a functional marker InDel-Rf-1a based on the difference of nucleotide sequence in Rf-1a locus between BT-type CMS lines and restorer lines was developed to detect the genotypes of different rice materials. Conventional indica rice varieties, restorer and maintainer lines without 574 bp deletion could restore the fertility for BT-type CMS in japonica rice. By contrast, most conventional japonica rice varieties except Aichi 106 and Yijing 12, with genotype of rf-1arf-1a showed the 574 bp deletion maintained sterility for BT-type CMS lines. To further verify the effect of genotyping detection in Rf-1a locus, this marker was also used to amplify the genomic DNA in different japonica rice restorer lines, CMS lines, hybrids and F2 segregation population, and three genotypes in Rf-1a locus could be distinguished distinctly. Therefore, the marker InDel-Rf-1a could be widely used for genetic identification and marker-assisted selection (MAS) in breeding japonica restorer lines.

  12. Comparing triage algorithms using HPV DNA genotyping, HPV E7 mRNA detection and cytology in high-risk HPV DNA-positive women.

    Science.gov (United States)

    Luttmer, Roosmarijn; Berkhof, Johannes; Dijkstra, Maaike G; van Kemenade, Folkert J; Snijders, Peter J F; Heideman, Daniëlle A M; Meijer, Chris J L M

    2015-06-01

    High-risk human papillomavirus (hrHPV) DNA positive women require triage testing to identify those with high-grade cervical intraepithelial neoplasia or cancer (≥CIN2). Comparing three triage algorithms (1) E7 mRNA testing following HPV16/18/31/33/45/52/58 genotyping (E7 mRNA test), (2) HPV16/18 DNA genotyping and (3) cytology, for ≥CIN2 detection in hrHPV DNA-positive women. hrHPV DNA-positive women aged 18-63 years visiting gynecology outpatient clinics were included in a prospective observational cohort study. From these women a cervical scrape and colposcopy-directed biopsies were obtained. Cervical scrapes were evaluated by cytology, HPV DNA genotyping by bead-based multiplex genotyping of GP5+6+-PCR-products, and presence of HPV16/18/31/33/45/52/58 E7 mRNA using nucleic acid sequence-based amplification (NASBA) in DNA positive women for respective HPV types. Sensitivities and specificities for ≥CIN2 were compared between E7 mRNA test and HPV16/18 DNA genotyping in the total group (n=348), and E7 mRNA test and cytology in a subgroup of women referred for non-cervix-related gynecological complaints (n=133). Sensitivity for ≥CIN2 of the E7 mRNA test was slightly higher than that of HPV16/18 DNA genotyping (66.9% versus 60.9%; ratio 1.10, 95% CI: 1.0002-1.21), at similar specificity (54.8% versus 52.3%; ratio 1.05, 95% CI: 0.93-1.18). Neither sensitivity nor specificity of the E7 mRNA test differed significantly from that of cytology (sensitivity: 68.8% versus 75.0%; ratio 0.92, 95% CI: 0.72-1.17; specificity: 59.4% versus 65.3%; ratio 0.91, 95% CI: 0.75-1.10). For detection of ≥CIN2 in hrHPV DNA-positive women, an algorithm including E7 mRNA testing following HPV16/18/31/33/45/52/58 DNA genotyping performs similar to HPV16/18 DNA genotyping or cytology. Copyright © 2015 Elsevier B.V. All rights reserved.

  13. Development and evaluation of a real-time RT-qPCR for detection of Crimean-Congo hemorrhagic fever virus representing different genotypes.

    Science.gov (United States)

    Jääskeläinen, Anne J; Kallio-Kokko, Hannimari; Ozkul, Aykut; Bodur, Hurrem; Korukruoglu, Gulay; Mousavi, Mehrdad; Pranav, Patel; Vaheri, Antti; Mirazimi, Ali; Vapalahti, Olli

    2014-12-01

    Crimean-Congo hemorrhagic fever (CCHF) is a zoonotic disease caused by a nairovirus belonging to family Bunyaviridae. The CCHF virus (CCHFV) can be transmitted to humans by Hyalomma ticks as well as by direct contact with infected body fluids or tissues from viremic livestock or humans. Our aim was to set up a fast RT-qPCR for detection of the different CCHFV genotypes in clinical samples, including an inactivation step to make the sample handling possible in lower biosafety levels (BSL) than BSL-4. This method was evaluated against commercial reference assays and international External Quality Assessment (EQA) samples. The analytical limit of detection for the developed CCHFV-S RT-qPCR was 11 CCHFV genomes per reaction. After exclusion of four dubious samples, we studied 38 CCHFV-positive samples (using reference tests) of which 38 were found positive by CCHFV-S RT-qPCR, suggesting a sensitivity of 100%. CCHFV-S RT q-PCR detected all eight different CCHFV strains representing five different CCHFV genotypes. In conclusion, the CCHFV-S RT-qPCR described in this study was evaluated using various sources of CCHFV samples and shown to be an accurate tool to detect human CCHFV infection caused by different genotypes of the virus.

  14. Molecular characterization of Dobrava and Kurkino genotypes of Dobrava-Belgrade hantavirus detected in Hungary and Northern Croatia.

    Science.gov (United States)

    Németh, Viktória; Oldal, Miklós; Madai, Mónika; Horváth, Győző; Kemenesi, Gábor; Dallos, Bianka; Bányai, Krisztián; Jakab, Ferenc

    2013-12-01

    Among the Hantavirus genus, Saaremaa virus (SAAV) has been the subject of taxonomical debates. While the International Committee on Taxonomy of Viruses declares SAAV as a distinct species, several European hantavirus experts proposed that SAAV is in fact a genotype of Dobrava-Belgrade virus (DOBV). In the present study we performed S-segment-based phylogenetic analysis of eight DOBV strains identified in rodents in Hungary and Northern Croatia. These new sequences considerably increase the number of complete nucleoprotein gene sequences deposited in the NCBI database. Our phylogenetic analysis clearly support the taxonomical nomenclature recently proposed for DOBV, i.e., genotypes such as Dobrava, Saaremaa, Kurkino, and Sochi should indeed be classified within the DOBV hantavirus species. Moreover, we found that only the Dobrava and Kurkino genotypes of DOBV species are circulating in Hungary while currently there is no evidence for the presence of Saaremaa genotype.

  15. Genotypic diversity, pathogenic potential and the resistance profile of Salmonella Typhimurium strains isolated from humans and food from 1983 to 2013 in Brazil.

    Science.gov (United States)

    Almeida, Fernanda; Medeiros, Marta Inês Cazentini; Rodrigues, Dália dos Prazeres; Falcão, Juliana Pfrimer

    2015-11-01

    Salmonella enterica subsp. enterica serovar Typhimurium is one of the leading serovars that causes salmonellosis worldwide. However, few studies have molecularly characterized S. Typhimurium strains in Brazil. In this study, we genotyped 92 S. Typhimurium strains isolated from humans (43) and food (49) between 1983 and 2013 in Brazil using PFGE, multiple-locus variable number of tandem repeats analysis (MLVA) and enterobacterial repetitive intergenic consensus PCR (ERIC-PCR). Moreover, we assessed the frequency of 12 virulence markers by PCR and the resistance profile against 12 antimicrobials. More than 85.8% of the strains studied carried 11 of the virulence markers or more. Thirty-three strains (25%) were multidrug resistant (MDR). The 92 S. Typhimurium studied were grouped by PFGE as PFGE-A, PFGE-B1 and PFGE-B2; by MLVA as MLVA-A, MLVA-B1 and MLVA-B2; and, finally, by ERIC-PCR as ERIC-A and ERIC-B. The strains isolated from humans before the mid-1990s were allocated to all clusters. The strains isolated from humans after the mid-1990s were distributed in the PFGE-B1, MLVA-B1, MLVA-B2 and ERIC-A clusters. The strains isolated from food were distributed in all clusters, except in PFGE-B2. All typing results suggested that the S. Typhimurium strains of human clinical origin isolated before the mid-1990s were genetically more diverse, which might indicate the selection of a more adapted S. Typhimurium subtype after Salmonella Enteritidis became the most prevalent serovar in Brazil. Regarding strains isolated from food, the results suggest the current circulation of more than one subtype. Furthermore, the high frequency of virulence genes and the presence of MDR strains reinforces their potential hazard for humans and the risk of their presence in foods in Brazil.

  16. Genotyping of Endocervical Chlamydia trachomatis Strains and Detection of Serological Markers of Acute and Chronic Inflammation in Their Host

    Directory of Open Access Journals (Sweden)

    Behrouz Taheri Beni

    2012-01-01

    Full Text Available Background: Chlamydia trachomatis (C. trachomatis is the most prevalent cause ofbacterial sexually transmitted infections (STI recognized throughout the world. Theaim of this study is to determine different genotypes of genital C. trachomatis andthe association between the serological markers of inflammation and genotypes ofC. trachomatis in sexually active women (n=80 attending Shahid Beheshti Hospitalin Isfahan, Iran.Materials and Methods: In this descriptive study, endocervical swabs were collectedfrom 80 women. There were 17 endocervical samples that showed positivity for C. trachomatisby plasmid polymerase chain reaction (PCR using KL1 and KL2 primers. Theomp1 gene was directly amplified in 17 plasmid PCR positive samples and was usedto differentiate the clinical genotypes by omp1 gene PCR-restriction fragment lengthpolymorphism (PCR-RFLP. The levels of IgG and IgA specific to C. trachmatis andC-reactive protein (CRP were evaluated.Results: Based on restriction-digestion patterns, four genotypes were identified. GenotypesE (35.3% and F (35.3% were the most prevalent, followed by D/Da (23.5% and K(5.9%. There was no significant association between genotypes and the presence ofIgG and CRP. Patients infected with genotype E showed a serological marker of chronicinflammation, i.e. IgA seropositivity, significantly more than patients infected with othergenotypes (p=0.042.Conclusion: Nested PCR could increase the sensitivity of omp1 amplification. Based onthe presence of IgA, chronic C. trachomatis infections were observed more frequentlyamong genotype E-infected patients in our population.

  17. Detection of Genetic Diversity in Synthetic Hexaploid Wheats Using Microsatellite Markers

    Institute of Scientific and Technical Information of China (English)

    CHEN Guo-yue; LI Li-hui

    2007-01-01

    Ninety-five synthetic hexaploid wheats(2n=6x=42,AABBDD)were analyzed using 45 microsatellite markers to investigate the potential genetic diversity in wheat breeding programs.A total of 326 alleles were detected by these microsatellite primer pairs,with an average of 6.65 alleles per locus.The polymorphic information content(PIC),Simpson index(SI),and genetic similarity(GS)coefficient showed that the D genome is of the highest genetic diversity among the A,B,and D genomes in the synthetic hexaploid wheats.The results also indicated that the synthetic hexaploid wheat is an efficient way to enrich wheat genetic backgrounds,especially to use the genetic variations of the D genome from Aegilops squarrosa for wheat improvement.The UPGMA dendogram,based on a similarity matrix by a simple matching coefficient algorithm,delineated the above accessions into 5 major clusters and was in accordance with the available pedigree information.The results demonstrated the utility of microsatellite markers in detecting DNA polymorphism and estimating genetic diversity.

  18. Hepatitis C virus long-term persistence in peripheral blood mononuclear cells in patients with haemophilia. Detection of occult genotype 1.

    Science.gov (United States)

    Parodi, C; García, G; Monzani, M C; Culasso, A; Aloisi, N; Corti, M; Campos, R; de E de Bracco, M M; Baré, P

    2015-07-01

    Peripheral blood mononuclear cells (PBMC) from chronic hepatitis C virus-infected persons can harbour viral variants that are not detected in plasma samples. We explored the presence and persistence of HCV genotypes in plasma and PBMC cultures from 25 HCV-monoinfected and 25 HIV/HCV-coinfected patients with haemophilia. Cell cultures were performed at different time points between 1993 and 2010-2011, and the HCV genome was examined in culture supernatants. Sequential plasma samples were studied during the same time period. Analysing sequential plasma samples, 21% of patients had mixed-genotype infections, while 50% had mixed infections determined from PBMC culture supernatants. HIV coinfection was significantly associated with the presence of mixed infections (OR = 4.57, P = 0.02; 95% CI = 1.38-15.1). In our previous study, genotype 1 was found in 72% of 288 patients of this cohort. Similar results were obtained with the sequential plasma samples included in this study, 69% had genotype 1. However, when taking into account plasma samples and the results from PBMC supernatants, genotype 1 was identified in 94% of the population. The PBMC-associated variants persisted for 10 years in some subjects, emphasizing their role as long-term reservoirs. The presence of genotype 1 in PBMC may be associated with therapeutic failure and should not be disregarded when treating haemophilic patients who have been infected by contaminated factor concentrates. The clinical implications of persistent lymphotropic HCV variants deserve further examination among multiple exposed groups of HCV-infected patients.

  19. E2 GLYCOPROTEIN OF GENOTYPE Ⅲ CHINESE ISOLATES OF HEPATITIS C VIRUS EXPRESSED IN MAMMALIAN CELL AS ANTIGEN FOR ANTI-E2 ANTIBODY DETECTION

    Institute of Scientific and Technical Information of China (English)

    吴朝栋; 陶其敏

    1998-01-01

    Expression vector inserted with E2/NS1 gane derived from ganotype Ⅲ Chinese isolates of HCV was transfected into mammalian cells to express E2 glycoprotein. Expressed protein was used as antigen for anti-E2 antibody detection in 19 cases of hepatitis C patients by Western blot. It was first to express E2 glycoprotein of genotype Ⅲ Chinese hepatitis C virus isolates. For anti-E2 detection, 14 cases of patients were positive of antibodies against E2(73.7%). These results indicated that E2 glycoprotein expressed in mammalian cells had good immunoganicity and cross reactivity to serum infected with genotype Ⅱ Chinese hepatitis C virus isolates.

  20. Hepatitis B virus genotype E detected in Brazil in an African patient who is a frequent traveler

    Directory of Open Access Journals (Sweden)

    R Sitnik

    2007-12-01

    Full Text Available Genotype E of hepatitis B virus (HBV has not been described in Brazil and is found mainly in Africa. Genotype A is the most prevalent in Brazil, and genotypes B, C, D, and F have already been reported. We report here an HBV genotype E-infected patient and some characterization of surface (S protein, DNA polymerase (P and precore/core (preC/C coding regions based on the viral genome. The patient is a 31-year-old black man with chronic hepatitis B who was born and raised in Angola. He has been followed by a hepatologist in São Paulo, Brazil, since November 2003, and he is a frequent traveler to Latin America, Africa, and Europe. In 2003, he was diagnosed with HBV infection and started treatment with lamivudine with the later addition of adefovir dipivoxil. No known risk factor was identified. Serologically, he is HBsAg and anti-HBe positive, but HBeAg and anti-HBs negative. DNA sequence analysis of the S/P region confirmed that this patient is infected with genotype E, subtype ayw4. The preC/C region showed G1896A and G1899A mutations but no mutations in the basal core promoter. Nucleotide substitutions common in genotype E were also observed (C1772, T1858 and A1757. Although this is not an autochthonous case and there is no evidence of further spread, the description of this case in Brazil highlights the current risk of viral genotypes spreading with unprecedented speed due to constant travel around the world.

  1. Evaluation of genotype MTBDRplus VER 2.0 line probe assay for the detection of MDR-TB in smear positive and negative sputum samples.

    Science.gov (United States)

    Meaza, Abyot; Kebede, Abebaw; Yaregal, Zelalem; Dagne, Zekarias; Moga, Shewki; Yenew, Bazezew; Diriba, Getu; Molalign, Helina; Tadesse, Mengistu; Adisse, Desalegn; Getahun, Muluwork; Desta, Kassu

    2017-04-17

    Multi drug resistant tuberculosis (MDR-TB) poses formidable challenges to TB control due to its complex diagnostic and treatment challenges and often associated with a high rate of mortality. Accurate and rapid detection of MDR-TB is critical for timely initiation of treatment. Line Probe Assay (LPA) is a qualitative in vitro diagnostic test based on DNA-STRIP technology for the identification of the M. tuberculosis complex and its resistance to rifampicin (RMP) and/or isoniazid (INH). Hain Lifescience, GmbH, Germany has improved the sensitivity of Genotype MTBDRplus VER 2.0 LPA for the detection of MDR-TB; with the possibility of applying the tool in smear negative sputum samples. A cross sectional study was conducted on 274 presumptive MDR-TB patients referred to the National TB Reference Laboratory (NTRL), Ethiopian Public Health Institute (EPHI) who submitted sputum samples for laboratory diagnosis of drug resistant-TB testing. Seventy-two smear and culture positive samples processed in smear positive direct LPA category and 197 smear negative sputum samples were processed for direct LPA. Among the smear negative samples 145 (73.6%) were culture negative and 26 (13.2%) were culture positive. All specimens were processed using NALC-NaOH method and ZN smear microscopy done from sediments. Genotype MTBDRplus VER 2.0 done from processed sputum sediments and the result was compared against the reference, BACTEC MGIT 960 culture and DST. Sensitivity, specificity, PPV and NPV of Genotype MTBDRplus VER 2.0 assay was determined and P-value TB from direct smear positive sputum samples. The sensitivity, specificity, PPV and NPV of Genotype MTBDR plus VER 2.0 LPA were 77.8, 97.2, 82.4 and 97.2%, respectively, for the detection of M. tuberculosis from direct smear negative sputum samples. Fourteen (53.8%) samples had valid results with LPA among the 26 smear negative culture positive samples. The remaining 8 (30.8%) and 4 (15.4%) were invalid and negative with LPA

  2. Detection of rotavirus species A, B and C in domestic mammalian animals with diarrhoea and genotyping of bovine species A rotavirus strains.

    Science.gov (United States)

    Otto, Peter H; Rosenhain, Stefanie; Elschner, Mandy C; Hotzel, Helmut; Machnowska, Patrycja; Trojnar, Eva; Hoffmann, Kathrin; Johne, Reimar

    2015-09-30

    Rotaviruses (RVs) are a major cause of neonatal diarrhoea in humans and animals worldwide. In this study, 425 faecal samples were collected between 1999 and 2013 from diarrhoeic livestock and companion animals at different locations in Germany and tested for RVs. A previously published real-time RT-PCR assay was optimized for detection of a larger variety of RV species A (RVA) strains, and real-time RT-PCR assays for detection of RV species B (RVB) and C (RVC) were newly developed. The detection limits of the assays were 1.54×10(2), 3.95×10(2) and 3.60×10(3) genome copies for RVA, RVB and RVC, respectively. RVA was identified in 85.2% of bovine samples, 51.2% of porcine samples, 50.0% of feline samples, 43.2% of equine samples and 39.7% of canine samples. RVB was found in 3.0% of bovine samples, 2.7% of equine samples and 1.6% of porcine samples. RVC was detected in 31.0% of porcine samples, 21.7% of feline samples, 9.0% of canine samples and 6.0% of bovine samples. For genotyping, 101 RVA-positive bovine samples were further analysed by semi-nested RT-PCR. Genotype combination G6P[5] was most frequently detected (67.3% of samples), followed by G6P[11] (13.9%), G10P[5] (4.0%), G8P[11] (3.0%), G6P[1] (1.0%), and G10P[11] (1.0%). Mixed RVA infections were detected in 5.9% of samples; no or incomplete typing was possible in 4.0% of the samples. This first overview on RV species and RVA genotypes in diarrhoeic livestock and companion animals from Germany indicates a broad circulation of a large variety of RVs.

  3. (Brassica napus L.) genotypes

    African Journals Online (AJOL)

    STORAGESEVER

    2009-10-05

    Oct 5, 2009 ... The genetic diversity and relationships among rapeseed genotypes were ... dent of environment and plant growth stage, unlimited ..... interactions that lead to the expression of particular traits .... thesis, Faculty of Agriculture, University of Novi Sad. ... in the U.S. hard red winter wheat cultivars as reveled by.

  4. Fiber-Based Polarization Diversity Detection for Polarization-Sensitive Optical Coherence Tomography

    Directory of Open Access Journals (Sweden)

    Hamid Pahlevaninezhad

    2014-09-01

    Full Text Available We present a new fiber-based polarization diversity detection (PDD scheme for polarization sensitive optical coherence tomography (PSOCT. This implementation uses a new custom miniaturized polarization-maintaining fiber coupler with single mode (SM fiber inputs and polarization maintaining (PM fiber outputs. The SM fiber inputs obviate matching the optical lengths of the two orthogonal OCT polarization channels prior to interference while the PM fiber outputs ensure defined orthogonal axes after interference. Advantages of this detection scheme over those with bulk optics PDD include lower cost, easier miniaturization, and more relaxed alignment and handling issues. We incorporate this PDD scheme into a galvanometer-scanned OCT system to demonstrate system calibration and PSOCT imaging of an achromatic quarter-wave plate, fingernail in vivo, and chicken breast, salmon, cow leg, and basa fish muscle samples ex vivo.

  5. Fiber-optic polarization diversity detection for rotary probe optical coherence tomography.

    Science.gov (United States)

    Lee, Anthony M D; Pahlevaninezhad, Hamid; Yang, Victor X D; Lam, Stephen; MacAulay, Calum; Lane, Pierre

    2014-06-15

    We report a polarization diversity detection scheme for optical coherence tomography with a new, custom, miniaturized fiber coupler with single mode (SM) fiber inputs and polarization maintaining (PM) fiber outputs. The SM fiber inputs obviate matching the optical lengths of the X and Y OCT polarization channels prior to interference and the PM fiber outputs ensure defined X and Y axes after interference. Advantages for this scheme include easier alignment, lower cost, and easier miniaturization compared to designs with free-space bulk optical components. We demonstrate the utility of the detection system to mitigate the effects of rapidly changing polarization states when imaging with rotating fiber optic probes in Intralipid suspension and during in vivo imaging of human airways.

  6. Low prevalence of pneumococcal carriage and high serotype and genotype diversity among adults over 60 years of age living in Portugal.

    Directory of Open Access Journals (Sweden)

    Sónia T Almeida

    Full Text Available Pneumococcal disease is frequent at the extremes of age. While several studies have looked at colonization among young children, much less is known among the elderly. We aimed to evaluate pneumococcal carriage among elderly adults living in Portugal. Between April 2010 and December 2012, nasopharyngeal and oropharyngeal swabs of adults over 60 years of age, living in an urban area (n = 1,945 or in a rural area (n = 1,416, were obtained. Pneumococci were isolated by culture-based standard procedures, identified by optochin susceptibility, bile solubility and PCR screening for lytA and cpsA, and characterized by antibiotype, serotype, and MLST. Associations between pneumococcal carriage, socio-demographic and clinical characteristics were evaluated by univariate analysis and multiple logistic regression. The global prevalence of carriage was 2.3% (95% CI: 1.8-2.8. In the multiple logistic regression analysis, smoking, being at a retirement home, and living in a rural area increased the odds of being a pneumococcal carrier by 4.4-fold (95% CI: 1.9-9.2, 2.0-fold (95% CI: 1.1-3.6 and 2.0-fold (95% CI: 1.2-3.5, respectively. Among the 77 pneumococcal isolates, 26 serotypes and 40 STs were identified. The most prevalent serotypes were (in decreasing order 19A, 6C, 22F, 23A, 35F, 11A, and 23B, which accounted, in total, for 60.0% of the isolates. Most isolates (93.5% had STs previously described in the MLST database. Resistance to macrolides, non-susceptibility to penicillin and multidrug resistance were found in 19.5%, 11.7%, and 15.6% of the isolates, respectively. We conclude that the prevalence of pneumococcal carriage in the elderly, in Portugal, as determined by culture-based methods, is low. Serotype and genotype diversity is high. Living in a rural area, in a retirement home, and being a smoker increased the risk of pneumococcal carriage. This study contributes to the establishment of a baseline that may be used to monitor how novel

  7. Population samples and genotyping technology.

    Science.gov (United States)

    Mack, S J; Sanchez-Mazas, A; Single, R M; Meyer, D; Hill, J; Dron, H A; Jani, A J; Thomson, G; Erlich, H A

    2007-04-01

    The 14th International HLA (human leukocyte antigen) Immunogenetics Workshop (14th-IHIWS) Biostatistics and Anthropology/Human Genetic Diversity project continues the population sampling, genotype data generation, and biostatistic analyses of the 13th International Histocompatibility Workshop Anthropology/Human Genetic Diversity Component, with the overall goal of further characterizing global HLA allele and haplotype diversity and better describing the relationships between major histocompatibility complex diversity, geography, linguistics, and population history. Since the 13th Workshop, new investigators have and continue to be recruited to the project and new high-resolution class I and class II genotype data are being generated for 112 population samples from around the world.

  8. Association Mapping of Total Carotenoids in Diverse Soybean Genotypes Based on Leaf Extracts and High-Throughput Canopy Spectral Reflectance Measurements.

    Directory of Open Access Journals (Sweden)

    Arun Prabhu Dhanapal

    Full Text Available Carotenoids are organic pigments that are produced predominantly by photosynthetic organisms and provide antioxidant activity to a wide variety of plants, animals, bacteria, and fungi. The carotenoid biosynthetic pathway is highly conserved in plants and occurs mostly in chromoplasts and chloroplasts. Leaf carotenoids play important photoprotective roles and targeted selection for leaf carotenoids may offer avenues to improve abiotic stress tolerance. A collection of 332 soybean [Glycine max (L. Merr.] genotypes was grown in two years and total leaf carotenoid content was determined using three different methods. The first method was based on extraction and spectrophotometric determination of carotenoid content (eCaro in leaf tissue, whereas the other two methods were derived from high-throughput canopy spectral reflectance measurements using wavelet transformed reflectance spectra (tCaro and a spectral reflectance index (iCaro. An association mapping approach was employed using 31,253 single nucleotide polymorphisms (SNPs to identify SNPs associated with total carotenoid content using a mixed linear model based on data from two growing seasons. A total of 28 SNPs showed a significant association with total carotenoid content in at least one of the three approaches. These 28 SNPs likely tagged 14 putative loci for carotenoid content. Six putative loci were identified using eCaro, five loci with tCaro, and nine loci with iCaro. Three of these putative loci were detected by all three carotenoid determination methods. All but four putative loci were located near a known carotenoid-related gene. These results showed that carotenoid markers can be identified in soybean using extract-based as well as by high-throughput canopy spectral reflectance-based approaches, demonstrating the utility of field-based canopy spectral reflectance phenotypes for association mapping.

  9. Association Mapping of Total Carotenoids in Diverse Soybean Genotypes Based on Leaf Extracts and High-Throughput Canopy Spectral Reflectance Measurements

    Science.gov (United States)

    Dhanapal, Arun Prabhu; Ray, Jeffery D.; Singh, Shardendu K.; Hoyos-Villegas, Valerio; Smith, James R.; Purcell, Larry C.; King, C. Andy; Fritschi, Felix B.

    2015-01-01

    Carotenoids are organic pigments that are produced predominantly by photosynthetic organisms and provide antioxidant activity to a wide variety of plants, animals, bacteria, and fungi. The carotenoid biosynthetic pathway is highly conserved in plants and occurs mostly in chromoplasts and chloroplasts. Leaf carotenoids play important photoprotective roles and targeted selection for leaf carotenoids may offer avenues to improve abiotic stress tolerance. A collection of 332 soybean [Glycine max (L.) Merr.] genotypes was grown in two years and total leaf carotenoid content was determined using three different methods. The first method was based on extraction and spectrophotometric determination of carotenoid content (eCaro) in leaf tissue, whereas the other two methods were derived from high-throughput canopy spectral reflectance measurements using wavelet transformed reflectance spectra (tCaro) and a spectral reflectance index (iCaro). An association mapping approach was employed using 31,253 single nucleotide polymorphisms (SNPs) to identify SNPs associated with total carotenoid content using a mixed linear model based on data from two growing seasons. A total of 28 SNPs showed a significant association with total carotenoid content in at least one of the three approaches. These 28 SNPs likely tagged 14 putative loci for carotenoid content. Six putative loci were identified using eCaro, five loci with tCaro, and nine loci with iCaro. Three of these putative loci were detected by all three carotenoid determination methods. All but four putative loci were located near a known carotenoid-related gene. These results showed that carotenoid markers can be identified in soybean using extract-based as well as by high-throughput canopy spectral reflectance-based approaches, demonstrating the utility of field-based canopy spectral reflectance phenotypes for association mapping. PMID:26368323

  10. Association Mapping of Total Carotenoids in Diverse Soybean Genotypes Based on Leaf Extracts and High-Throughput Canopy Spectral Reflectance Measurements.

    Science.gov (United States)

    Dhanapal, Arun Prabhu; Ray, Jeffery D; Singh, Shardendu K; Hoyos-Villegas, Valerio; Smith, James R; Purcell, Larry C; King, C Andy; Fritschi, Felix B

    2015-01-01

    Carotenoids are organic pigments that are produced predominantly by photosynthetic organisms and provide antioxidant activity to a wide variety of plants, animals, bacteria, and fungi. The carotenoid biosynthetic pathway is highly conserved in plants and occurs mostly in chromoplasts and chloroplasts. Leaf carotenoids play important photoprotective roles and targeted selection for leaf carotenoids may offer avenues to improve abiotic stress tolerance. A collection of 332 soybean [Glycine max (L.) Merr.] genotypes was grown in two years and total leaf carotenoid content was determined using three different methods. The first method was based on extraction and spectrophotometric determination of carotenoid content (eCaro) in leaf tissue, whereas the other two methods were derived from high-throughput canopy spectral reflectance measurements using wavelet transformed reflectance spectra (tCaro) and a spectral reflectance index (iCaro). An association mapping approach was employed using 31,253 single nucleotide polymorphisms (SNPs) to identify SNPs associated with total carotenoid content using a mixed linear model based on data from two growing seasons. A total of 28 SNPs showed a significant association with total carotenoid content in at least one of the three approaches. These 28 SNPs likely tagged 14 putative loci for carotenoid content. Six putative loci were identified using eCaro, five loci with tCaro, and nine loci with iCaro. Three of these putative loci were detected by all three carotenoid determination methods. All but four putative loci were located near a known carotenoid-related gene. These results showed that carotenoid markers can be identified in soybean using extract-based as well as by high-throughput canopy spectral reflectance-based approaches, demonstrating the utility of field-based canopy spectral reflectance phenotypes for association mapping.

  11. Using RNA-Seq for gene identification, polymorphism detection and transcript profiling in two alfalfa genotypes with divergent cell wall composition in stems

    Science.gov (United States)

    2011-01-01

    Background Alfalfa, [Medicago sativa (L.) sativa], a widely-grown perennial forage has potential for development as a cellulosic ethanol feedstock. However, the genomics of alfalfa, a non-model species, is still in its infancy. The recent advent of RNA-Seq, a massively parallel sequencing method for transcriptome analysis, provides an opportunity to expand the identification of alfalfa genes and polymorphisms, and conduct in-depth transcript profiling. Results Cell walls in stems of alfalfa genotype 708 have higher cellulose and lower lignin concentrations compared to cell walls in stems of genotype 773. Using the Illumina GA-II platform, a total of 198,861,304 expression sequence tags (ESTs, 76 bp in length) were generated from cDNA libraries derived from elongating stem (ES) and post-elongation stem (PES) internodes of 708 and 773. In addition, 341,984 ESTs were generated from ES and PES internodes of genotype 773 using the GS FLX Titanium platform. The first alfalfa (Medicago sativa) gene index (MSGI 1.0) was assembled using the Sanger ESTs available from GenBank, the GS FLX Titanium EST sequences, and the de novo assembled Illumina sequences. MSGI 1.0 contains 124,025 unique sequences including 22,729 tentative consensus sequences (TCs), 22,315 singletons and 78,981 pseudo-singletons. We identified a total of 1,294 simple sequence repeats (SSR) among the sequences in MSGI 1.0. In addition, a total of 10,826 single nucleotide polymorphisms (SNPs) were predicted between the two genotypes. Out of 55 SNPs randomly selected for experimental validation, 47 (85%) were polymorphic between the two genotypes. We also identified numerous allelic variations within each genotype. Digital gene expression analysis identified numerous candidate genes that may play a role in stem development as well as candidate genes that may contribute to the differences in cell wall composition in stems of the two genotypes. Conclusions Our results demonstrate that RNA-Seq can be

  12. Baseline predictors of sputum culture conversion in pulmonary tuberculosis: importance of cavities, smoking, time to detection and W-Beijing genotype.

    Directory of Open Access Journals (Sweden)

    Marianne E Visser

    Full Text Available BACKGROUND: Time to detection (TTD on automated liquid mycobacterial cultures is an emerging biomarker of tuberculosis outcomes. The M. tuberculosis W-Beijing genotype is spreading globally, indicating a selective advantage. There is a paucity of data on the association between baseline TTD and W-Beijing genotype and tuberculosis outcomes. AIM: To assess baseline predictors of failure of sputum culture conversion, within the first 2 months of antitubercular therapy, in participants with pulmonary tuberculosis. DESIGN: Between May 2005 and August 2008 we conducted a prospective cohort study of time to sputum culture conversion in ambulatory participants with first episodes of smear and culture positive pulmonary tuberculosis attending two primary care clinics in Cape Town, South Africa. Rifampicin resistance (diagnosed on phenotypic susceptibility testing was an exclusion criterion. Sputum was collected weekly for 8 weeks for mycobacterial culture on liquid media (BACTEC MGIT 960. Due to missing data, multiple imputation was performed. Time to sputum culture conversion was analysed using a Cox-proportional hazards model. Bayesian model averaging determined the posterior effect probability for each variable. RESULTS: 113 participants were enrolled (30.1% female, 10.5% HIV-infected, 44.2% W-Beijing genotype, and 89% cavities. On Kaplan Meier analysis 50.4% of participants underwent sputum culture conversion by 8 weeks. The following baseline factors were associated with slower sputum culture conversion: TTD (adjusted hazard ratio (aHR = 1.11, 95% CI 1.02; 1.2, lung cavities (aHR = 0.13, 95% CI 0.02; 0.95, ever smoking (aHR = 0.32, 95% CI 0.1; 1.02 and the W-Beijing genotype (aHR = 0.51, 95% CI 0.25; 1.07. On Bayesian model averaging, posterior probability effects were strong for TTD, lung cavitation and smoking and moderate for W-Beijing genotype. CONCLUSION: We found that baseline TTD, smoking, cavities and W-Beijing genotype were

  13. Using RNA-Seq for gene identification, polymorphism detection and transcript profiling in two alfalfa genotypes with divergent cell wall composition in stems

    Directory of Open Access Journals (Sweden)

    Lamb JoAnn FS

    2011-04-01

    Full Text Available Abstract Background Alfalfa, [Medicago sativa (L. sativa], a widely-grown perennial forage has potential for development as a cellulosic ethanol feedstock. However, the genomics of alfalfa, a non-model species, is still in its infancy. The recent advent of RNA-Seq, a massively parallel sequencing method for transcriptome analysis, provides an opportunity to expand the identification of alfalfa genes and polymorphisms, and conduct in-depth transcript profiling. Results Cell walls in stems of alfalfa genotype 708 have higher cellulose and lower lignin concentrations compared to cell walls in stems of genotype 773. Using the Illumina GA-II platform, a total of 198,861,304 expression sequence tags (ESTs, 76 bp in length were generated from cDNA libraries derived from elongating stem (ES and post-elongation stem (PES internodes of 708 and 773. In addition, 341,984 ESTs were generated from ES and PES internodes of genotype 773 using the GS FLX Titanium platform. The first alfalfa (Medicago sativa gene index (MSGI 1.0 was assembled using the Sanger ESTs available from GenBank, the GS FLX Titanium EST sequences, and the de novo assembled Illumina sequences. MSGI 1.0 contains 124,025 unique sequences including 22,729 tentative consensus sequences (TCs, 22,315 singletons and 78,981 pseudo-singletons. We identified a total of 1,294 simple sequence repeats (SSR among the sequences in MSGI 1.0. In addition, a total of 10,826 single nucleotide polymorphisms (SNPs were predicted between the two genotypes. Out of 55 SNPs randomly selected for experimental validation, 47 (85% were polymorphic between the two genotypes. We also identified numerous allelic variations within each genotype. Digital gene expression analysis identified numerous candidate genes that may play a role in stem development as well as candidate genes that may contribute to the differences in cell wall composition in stems of the two genotypes. Conclusions Our results demonstrate that RNA

  14. Validation study for using lab-on-chip technology for Coxiella burnetii multi-locus-VNTR-analysis (MLVA) typing: application for studying genotypic diversity of strains from domestic ruminants in France.

    Science.gov (United States)

    Prigent, Myriam; Rousset, Elodie; Yang, Elise; Thiéry, Richard; Sidi-Boumedine, Karim

    2015-01-01

    Coxiella burnetii, the etiologic bacterium of Q fever zoonosis, is still difficult to control. Ruminants are often carriers and involved in human epidemics. MLVA is a promising genotyping method for molecular epidemiology. Different techniques are used to resolve the MLVA band profiles such as electrophoresis on agarose gels, capillary electrophoresis or using the microfluidic Lab-on-Chip system. In this study, system based on microfluidics electrophoresis with Lab-on-Chip technology was assessed and applied on DNA field samples to investigate the genotypic diversity of C. burnetii strains circulating in France. The Lab-on-Chip technology was first compared to agarose gel electrophoresis. Subsequently, the set-up Lab-on-Chip technology was applied on 97 samples collected from ruminants in France using the 17 markers previously described. A discordance rate of 27% was observed between Lab-on-Chip and agarose gel electrophoresis. These discrepancies were checked and resolved by sequencing. The cluster analysis revealed classification based on host species and/or geographic origin criteria. Moreover, the circulation of different genotypic strains within the same farm was also observed. In this study, MLVA with Lab-on-Chip technology was shown to be more accurate, reproducible, user friendly and safer than gel electrophoresis. It also provides an extended data set from French ruminant C. burnetii circulating strains useful for epidemiological investigations. Finally, it raises some questions regarding the standardization and harmonization of C. burnetii MLVA genotyping.

  15. A novel species within the Fusarium graminearum complex from Ethiopia detected by a multilocus genotyping assay and molecular phylogenetics

    Science.gov (United States)

    Twenty isolates resembling members of the Fusarium graminearum species complex (Fg complex; O’Donnell et al., Fungal Genet. Biol. 41:600-623, 2004) were isolated from ground wheat samples collected in two different geographic areas in Ethiopia. Results of a multilocus genotyping (MLGT) assay (Ward ...

  16. Genotypic and phenotypic detection of capsular polysaccharides in Staphylococcus aureus isolated from bovine intramammary infections in Argentina

    Directory of Open Access Journals (Sweden)

    C. Camussone

    2012-09-01

    Full Text Available Staphylococcus aureus (n=157 isolated from intramammary infections in Argentine dairy areas were evaluated for presence of cap5 and cap8 loci. Isolates carrying cap5 and cap8 were serotyped using specific antisera. Sixty four percent of the isolates were genotyped as cap5 or cap8 and 50% of them expressed CP5 or 8.

  17. Genotyping of Giardia lamblia and methods of its detection%蓝氏贾第鞭毛虫基因型及其检测方法概述

    Institute of Scientific and Technical Information of China (English)

    郑佳

    2011-01-01

    蓝氏贾第鞭毛虫致病性与其基因型密切相关.最近发展的一项分子技术real-time PCR以其实时定量、灵敏度高、特异性强等优势,被广泛应用于病毒、细菌和寄生虫的临床检测.本文就蓝氏贾第鞭毛虫基因型及检测方法的研究进行了综述.%The pathogenicity of giardia is closely related to its genotype. Recent advances in a molecular technique, realtime PCR, have provided real-time quantification, a high degree of sensitivity, a high degree of specificity, and other advantages. This technique is widely used to clinically test for viruses, bacteria, and parasites. This paper summarizes the genotyping and detection of Giardia lamblia.

  18. Detection of the emerging rotavirus G12P[8] genotype at high frequency in brazil in 2014: Successive replacement of predominant strains after vaccine introduction.

    Science.gov (United States)

    Luchs, Adriana; Cilli, Audrey; Morillo, Simone Guadagnucci; Gregório, Debora de Souza; de Souza, Karen Aparecida Farias; Vieira, Heloísa Rosa; Fernandes, Adeline de Mira; Carmona, Rita de Cássia Compagnoli; Timenetsky, Maria do Carmo Sampaio Tavares

    2016-04-01

    The continuum characterization of rotavirus (RVA) genotypes is essential to understand how vaccine introduction could impact virus epidemiology. In the present study, an unexpected rapid changing pattern of RVA genotypes distribution in Brazilian population during three followed seasons is described. From January/2012 to December/2014, a total of 3441 fecal specimens were collected from collaborating centers across Southern, Southeastern and Midwest of Brazil. All specimens were screened for RVA using ELISA, and genotyped by RT-PCR. Differences in proportions were tested using Chi-Squares. A p-value of less than 0.05 was considered statistically significant. RVA was detected in 19.7% (677/3441). Among RVA positive cases (n=677), a total of 652 (96.3%) samples were successfully amplified by RT-PCR. G3P[8] remained prevalent in 2012 (37.6%, 69/185) and 2013 (40.1%, 74/186) (χ(2)=0.107, p=0.743), but declined markedly in 2014 (3.5%, 10/281) (χ(2)=71.770, p=0.000). G12P[8] was second highest strain in 2012 (22.7%, 42/185), decrease rapidly in 2013 (2.7%, 5/186) (χ(2)=26.224, p=0.000) and re-emerged as the predominant genotype in 2014 (86.6%, 243/281) (χ(2)=118.299, p=0.000). From July/2014, G12P[8] was the single genotype detected in all regions studied. The sudden emergence, spread and predominance of G12P[8] strain in Brazil, raised the hypothesis of a possible G12 outbreak being in progress. Nationally, the long term decline in gastroenteritis hospitalization observed in the country after RVA vaccine introduction was confirmed. Nevertheless, the sharp increase in diarrhea hospitalization prevalence from 2013 to 2014 observed in Southern and Southeastern regions is consistent with what appears to be an outbreak of G12P[8]. Continued surveillance is needed to verify the effectiveness of the RotarixTM vaccine in Brazil together with potential emergence of unusual genotypes.

  19. Prueba molecular Genotype® MTBDRplus, una alternativa para la detección rápida de tuberculosis multidrogorresistente Molecular test Genotype® MTBDRplus, an alternative to rapid detection of multidrug resistance tuberculosis

    Directory of Open Access Journals (Sweden)

    Luis Asencios

    2012-03-01

    Full Text Available La prueba molecular Genotype®MTBDRplus, es un método que permite identificar las mutaciones más frecuentes asociadas con la resistencia a las drogas antituberculosas de primera línea: isoniacida (INH y rifampicina (RIF. El objetivo de este estudio fue evaluar el desempeño de la prueba molecular con cultivos y muestras de esputo con baciloscopía positiva. Se evaluó 95 cultivos y 100 esputos con perfiles de resistencia previamente determinados por el método de referencia "proporciones agar en placa" (APP. La prueba molecular a partir de cultivos mostró una sensibilidad de 100%; 97,5% y 96,9% para RIF, INH y multidrogorresistente (MDR respectivamente; mientras que para esputo la sensibilidad fue de 95,7%; 96,8% y 95,2% para RIF, INH y MDR respectivamente. Se concluye que Genotype®MTBDRplus es una herramienta muy útil para la detección rápida de la resistencia a INH y RIF simultáneamente (MDR en un máximo de 72 h a partir de esputo o de cultivo.The Genotype®MTBDRplus molecular test is a method that allows identification of the most frequent mutations associated with resistance to major first-line antituberculosis drugs, Isoniazid (INH and Rifampicin (RFP. The aim of this study was to evaluate the performance of the molecular test with culture and smear- positive sputum samples. We evaluated 95 cultures and 100 sputum samples with resistance profiles previously determined by the reference method "Agar Plate Proportions" (APP. The molecular test from cultures showed a sensitivity of 100 %, 97,5 % and 96,97 % for RIF, INH and MDR respectively while from sputums the sensitivity was 95,65 %, 96,77 % and 95,24 % for RIF, INH and MDR respectively. We conclude that the molecular test Genotype®MTBDRplus is a very useful tool to detect resistance to isoniazid and rifampicin simultaneously (MDR-TB in up to 72 hours from sputum samples or cultures.

  20. Toward fully automated genotyping: Allele assignment, pedigree construction, phase determination, and recombination detection in Duchenne muscular dystrophy

    Energy Technology Data Exchange (ETDEWEB)

    Perlin, M.W.; Burks, M.B. [Carnegie Mellon Univ., Pittsburgh, PA (United States); Hoop, R.C.; Hoffman, E.P. [Univ. of Pittsburgh School of Medicine, PA (United States)

    1994-10-01

    Human genetic maps have made quantum leaps in the past few years, because of the characterization of >2,000 CA dinucleotide repeat loci: these PCR-based markers offer extraordinarily high PIC, and within the next year the