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Sample records for genotype diversity detected

  1. Low genetic diversity of bovine Mycobacterium avium subspecies paratuberculosis isolates detected by MIRU-VNTR genotyping.

    Science.gov (United States)

    de Kruijf, Marcel; Lesniak, Olga N; Yearsley, Dermot; Ramovic, Elvira; Coffey, Aidan; O'Mahony, Jim

    2017-05-01

    Mycobacterial interspersed repetitive unit and variable number tandem repeat (MIRU-VNTR) has been developed as a simple, rapid and cost efficient molecular typing method to differentiate Mycobacterium avium subspecies paratuberculosis (MAP) isolates. The aim of this study was to determine the genomic diversity of MAP across the Republic of Ireland by utilising the MIRU-VNTR typing method on a large collection of MAP isolates. A total of 114 MAP isolates originated from 53 herds across 19 counties in the Republic of Ireland were genotyped based on eight established MIRU-VNTR loci. Four INMV groups were observed during this study. INMV 1 was found in 67 MAP isolates (58.8%) and INMV 2 was observed in 45 isolates (39.4%). INMV 3 and INMV 116 recorded only one isolate each (0.9%). The unique INMV 116 group has never been reported among herds thus far and the molecular pattern of the MAP isolate classified in INMV 116 showed a difference at the MIRU-VNTR X3 locus compared to the other three INMV groups observed. INMV 1, INMV 2 and INMV 3 are observed frequently in Europe and comprised 99.1% of the total MAP isolates characterised in this study, indicating that MAP exhibited low level of genetic diversity across the Republic of Ireland using the MIRU-VNTR method. By the implementation of SNP analysis or MLSSR as an additional typing method, MAP genetic diversity would increase. INMV 3 is unique to Ireland and whereas INMV 116 has never been previously reported among herds by MIRU-VNTR typing. Copyright © 2017 Elsevier B.V. All rights reserved.

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

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    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. Whole genome analysis of porcine astroviruses detected in Japanese pigs reveals genetic diversity and possible intra-genotypic recombination.

    Science.gov (United States)

    Ito, Mika; Kuroda, Moegi; Masuda, Tsuneyuki; Akagami, Masataka; Haga, Kei; Tsuchiaka, Shinobu; Kishimoto, Mai; Naoi, Yuki; Sano, Kaori; Omatsu, Tsutomu; Katayama, Yukie; Oba, Mami; Aoki, Hiroshi; Ichimaru, Toru; Mukono, Itsuro; Ouchi, Yoshinao; Yamasato, Hiroshi; Shirai, Junsuke; Katayama, Kazuhiko; Mizutani, Tetsuya; Nagai, Makoto

    2017-06-01

    Porcine astroviruses (PoAstVs) are ubiquitous enteric virus of pigs that are distributed in several countries throughout the world. Since PoAstVs are detected in apparent healthy pigs, the clinical significance of infection is unknown. However, AstVs have recently been associated with a severe neurological disorder in animals, including humans, and zoonotic potential has been suggested. To date, little is known about the epidemiology of PoAstVs among the pig population in Japan. In this report, we present an analysis of nearly complete genomes of 36 PoAstVs detected by a metagenomics approach in the feces of Japanese pigs. Based on a phylogenetic analysis and pairwise sequence comparison, 10, 5, 15, and 6 sequences were classified as PoAstV2, PoAstV3, PoAstV4, and PoAstV5, respectively. Co-infection with two or three strains was found in individual fecal samples from eight pigs. The phylogenetic trees of ORF1a, ORF1b, and ORF2 of PoAstV2 and PoAstV4 showed differences in their topologies. The PoAstV3 and PoAstV5 strains shared high sequence identities within each genotype in all ORFs; however, one PoAstV3 strain and one PoAstV5 strain showed considerable sequence divergence from the other PoAstV3 and PoAstV5 strains, respectively, in ORF2. Recombination analysis using whole genomes revealed evidence of multiple possible intra-genotype recombination events in PoAstV2 and PoAstV4, suggesting that recombination might have contributed to the genetic diversity and played an important role in the evolution of Japanese PoAstVs. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Detection of diverse genotypes of Methicillin-resistant Staphylococcus aureus from hospital personnel and the environment in Armenia

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    Hermine V. Mkrtchyan

    2017-02-01

    Full Text Available Abstract Background Methicillin-resistant Staphylococcus aureus (MRSA is a public health concern internationally. Studies examining a range of cohorts have been reported from various regions of the world, but little is known about the molecular epidemiology of MRSA in Armenia. Methods Between May and September 2013, twenty isolates of methicillin-resistant Staphylococcus aureus (MRSA; mecA positive were recovered from hospital personnel (n = 10; 9 females, 1 male and environmental sites (n = 10 in the maternity ward of one of the teaching hospitals in Armenia. Results Multi-locus sequence type clonal complex (MLST-CC assignments inferred from spa typing data revealed the majority belonged to 3 pandemic lineages of MRSA including: t008-CC8-SCCmecV (n = 10; 7 from personnel; t021-CC30-SCCmecIV (n = 5; all environmental; and t1523-CC45 (n = 2; 1 from personnel, one harboured SCCmecV the other was SCCmec non-typable. The remainder identified as belonging to genotype t364-CC182, both of which harboured a novel SCCmec cassette with kdp, rif5, ccrB2 and ccrC detected by PCR (both from personnel; and t325-CC88-SCCmecIV (n = 1; environmental. All MRSA were negative for the Panton-Valentine Leukocidin (PVL locus and three CC8 strains were positive for the arginine catabolic element (ACME. Conclusions In this small study, we report for the first time of the occurrence of diverse MRSA genotypes belonging to both pandemic and more sporadic international clones in Armenia harbouring the smaller SCCmec types and/or ACME, both of which have been associated with strain fitness. Further surveillance is warranted to better understand the prevalence, clinical and molecular epidemiology of MRSA throughout Armenia.

  5. High degree of genetic diversity among genotypes of the forage grass Brachiaria ruziziensis (Poaceae) detected with ISSR markers.

    Science.gov (United States)

    Azevedo, A L S; Costa, P P; Machado, M A; de Paula, C M P; Sobrinho, F S

    2011-11-17

    The grasses of the genus Brachiaria account for 80% of the cultivated pastures in Brazil. Despite its importance for livestock production, little information is available for breeding purposes. Embrapa has a population of B. ruziziensis from different regions of Brazil, representing most of existing variability. This population was used to initiate an improvement program based on recurrent selection. In order to assist the genetic improvement program, we estimated the molecular variability among 93 genotypes of Embrapa's collection using ISSR (inter-simple sequence repeat) markers. DNA was extracted from the leaves. Twelve ISSR primers generated 89 polymorphic bands in the 93 genotypes. The number of bands identified by each primer ranged from two to 13, with a mean of 7.41. Cluster analysis revealed a clearly distinct group, containing most of the B. ruziziensis genotypes apart from the outgroup genotypes. Genetic similarity coefficients ranged from 0.0 to 0.95, with a mean of 0.50 and analysis of molecular variance indicated higher variation within (73.43%) than among species (26.57%). We conclude that there is a high genetic diversity among these B. ruziziensis genotypes, which could be explored by breeding programs.

  6. Precise genotyping and recombination detection of Enterovirus

    Science.gov (United States)

    2015-01-01

    Enteroviruses (EV) with different genotypes cause diverse infectious diseases in humans and mammals. A correct EV typing result is crucial for effective medical treatment and disease control; however, the emergence of novel viral strains has impaired the performance of available diagnostic tools. Here, we present a web-based tool, named EVIDENCE (EnteroVirus In DEep conception, http://symbiont.iis.sinica.edu.tw/evidence), for EV genotyping and recombination detection. We introduce the idea of using mixed-ranking scores to evaluate the fitness of prototypes based on relatedness and on the genome regions of interest. Using phylogenetic methods, the most possible genotype is determined based on the closest neighbor among the selected references. To detect possible recombination events, EVIDENCE calculates the sequence distance and phylogenetic relationship among sequences of all sliding windows scanning over the whole genome. Detected recombination events are plotted in an interactive figure for viewing of fine details. In addition, all EV sequences available in GenBank were collected and revised using the latest classification and nomenclature of EV in EVIDENCE. These sequences are built into the database and are retrieved in an indexed catalog, or can be searched for by keywords or by sequence similarity. EVIDENCE is the first web-based tool containing pipelines for genotyping and recombination detection, with updated, built-in, and complete reference sequences to improve sensitivity and specificity. The use of EVIDENCE can accelerate genotype identification, aiding clinical diagnosis and enhancing our understanding of EV evolution. PMID:26678286

  7. Pollen diversity, viability and floral structure of some Musa genotypes

    African Journals Online (AJOL)

    Pollen diversity, viability and floral structure of some Musa genotypes. ... This experiment was designed to study the floral structure, pollen morphology and the potential pollen viability of five Musa genotypes obtained ... HOW TO USE AJOL.

  8. Genotypic and phenotypic diversity of Alicyclobacillus acidocaldarius isolates.

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    Félix-Valenzuela, L; Guardiola-Avila, I; Burgara-Estrella, A; Ibarra-Zavala, M; Mata-Haro, V

    2015-10-01

    The fruit juice industry recognizes Alicyclobacillus as a major quality control target micro-organism. In this study, we analysed 19 bacterial isolates to identify Alicyclobacillus species by polymerase chain reaction (PCR) and sequencing analyses. Phenotypic and genomic diversity among isolates were investigated by API 50CHB system and ERIC-PCR (enterobacterial repetitive intergenic consensus-PCR) respectively. All bacterial isolates were identified as Alicyclobacillus acidocaldarius, and almost all showed identical DNA sequences according to their 16S rRNA (rDNA) gene partial sequences. Only few carbohydrates were fermented by A. acidocaldarius isolates, and there was little variability in the biochemical profile. Genotypic fingerprinting of the A. acidocaldarius isolates showed high diversity, and clusters by ERIC-PCR were distinct to those obtained from the 16S rRNA gene phylogenetic tree. There was no correlation between phenotypic and genotypic variability in the A. acidocaldarius isolates analysed in this study. Detection of Alicyclobacillus strains is imperative in fruit concentrates and juices due to the production of guaiacol. Identification of the genera originates rejection of the product by processing industry. However, not all the Alicyclobacillus species are deteriorative and hence the importance to differentiate among them. In this study, partial 16S ribosomal RNA sequence alignment allowed the differentiation of species. In addition, ERIC-PCR was introduced for the genotypic characterization of Alicyclobacillus, as an alternative for differentiation among isolates from the same species. © 2015 The Society for Applied Microbiology.

  9. Plant genotypic diversity reduces the rate of consumer resource utilization.

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    McArt, Scott H; Thaler, Jennifer S

    2013-07-07

    While plant species diversity can reduce herbivore densities and herbivory, little is known regarding how plant genotypic diversity alters resource utilization by herbivores. Here, we show that an invasive folivore--the Japanese beetle (Popillia japonica)--increases 28 per cent in abundance, but consumes 24 per cent less foliage in genotypic polycultures compared with monocultures of the common evening primrose (Oenothera biennis). We found strong complementarity for reduced herbivore damage among plant genotypes growing in polycultures and a weak dominance effect of particularly resistant genotypes. Sequential feeding by P. japonica on different genotypes from polycultures resulted in reduced consumption compared with feeding on different plants of the same genotype from monocultures. Thus, diet mixing among plant genotypes reduced herbivore consumption efficiency. Despite positive complementarity driving an increase in fruit production in polycultures, we observed a trade-off between complementarity for increased plant productivity and resistance to herbivory, suggesting costs in the complementary use of resources by plant genotypes may manifest across trophic levels. These results elucidate mechanisms for how plant genotypic diversity simultaneously alters resource utilization by both producers and consumers, and show that population genotypic diversity can increase the resistance of a native plant to an invasive herbivore.

  10. Pollen diversity, viability and floral structure of some Musa genotypes

    African Journals Online (AJOL)

    Prof. Ogunji

    Pollen diversity, viability and floral structure of some Musa genotypes ... at the Faculty of Agriculture & Natural Resources Management farm, Ebonyi State University,. Abakaliki. ..... Roots, tuber, plantains and bananas in human nutrition. Rome,.

  11. Genetic diversity of maize genotypes on the basis of morpho ...

    African Journals Online (AJOL)

    Genetic diversity of maize genotypes on the basis of morpho-physiological and simple sequence repeat (SSR) markers. Ashish Kumar, Arunita Rakshit, Naresh K Mangilipelli, Y Varalaxmi, T Vijayalakshmi, Jainender M Vanaja, SK Yadav, B Venkateswarlu, M Maheswari ...

  12. DNA landmarks for genetic diversity assessment in tea genotypes ...

    African Journals Online (AJOL)

    Tea (Camellia sinensis) is one of the most important non-alcoholic beverages of the world. Natural genetic diversity in tea has been reduced due to continue selection in favor of desirable traits. The present study was conducted to estimate genetic diversity in tea genotypes cultivated in Pakistan using 20 randomly amplified ...

  13. Genetic diversity of Pakistani maize genotypes using chromosome ...

    African Journals Online (AJOL)

    For improvement of maize crop presence of genetic diversity in the germplasm is very important. This study was conducted to determine genetic diversity among 17 Pakistani maize genotypes using 10 simple sequence repeat (SSR) primer sets. All the amplification products were in the range of <250-750 bp. To estimate the ...

  14. Protein landmarks for diversity assessment in wheat genotypes ...

    African Journals Online (AJOL)

    Grain proteins from 20 Indian wheat genotypes were evaluated for diversity assessment based seed storage protein profiling on sodium dodecylsulphate polyacrylamide gel electrophoresis (SDS-PAGE). Genetic diversity was evaluated using Nei's index, Shannon index and Unweighted pair group method with arithmetic ...

  15. 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...... 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...... 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...

  16. Genotyping-By-Sequencing for Plant Genetic Diversity Analysis: A Lab Guide for SNP Genotyping

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    Gregory W. Peterson

    2014-10-01

    Full Text Available Genotyping-by-sequencing (GBS has recently emerged as a promising genomic approach for exploring plant genetic diversity on a genome-wide scale. However, many uncertainties and challenges remain in the application of GBS, particularly in non-model species. Here, we present a GBS protocol we developed and use for plant genetic diversity analysis. It uses two restriction enzymes to reduce genome complexity, applies Illumina multiplexing indexes for barcoding and has a custom bioinformatics pipeline for genotyping. This genetic diversity-focused GBS (gd-GBS protocol can serve as an easy-to-follow lab guide to assist a researcher through every step of a GBS application with five main components: sample preparation, library assembly, sequencing, SNP calling and diversity analysis. Specifically, in this presentation, we provide a brief overview of the GBS approach, describe the gd-GBS procedures, illustrate it with an application to analyze genetic diversity in 20 flax (Linum usitatissimum L. accessions and discuss related issues in GBS application. Following these lab bench procedures and using the custom bioinformatics pipeline, one could generate genome-wide SNP genotype data for a conventional genetic diversity analysis of a non-model plant species.

  17. 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.

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

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    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 lack of dominant fecal sources in the watershed. High temporal variability of E. coli genotypes in soil was also observed, which suggests a dynamic E. coli population corresponding with the frequently observed high concentrations in tropical soils. When E. coli genotypes detected from the same sampling events were compared, limited sharing between the soil and water samples was observed in the majority of comparisons (73.5%). However, several comparisons reported up to 33.3% overlap of E. coli genotypes between soil and water, illustrating the potential for soil-water interactions under favorable environmental conditions. In addition, genotype accumulation curves for E. coli from water and soil indicated that the sampling efforts in the Manoa watershed could not exhaust the overall genotypic diversity. Comparisons of E. coli genotypes from other watersheds on Oahu, Hawaii, identified no apparent grouping according to sampling locations. The results of the present study demonstrate the complexity of using E. coli as a fecal indicator bacterium in tropical watersheds and highlight the need to differentiate environmental sources of E. coli from fecal sources in water quality monitoring. PMID:21515724

  19. 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.

  20. AFLP analysis of Genetic Diversity Among Different Jatropha curcas L. Genotypes from Africa and Ecuador

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    Konan, NO.

    2018-01-01

    Full Text Available Six populations amounting to a total number of seventy genotypes of Jatropha curcas L. originating from Africa (Senegal, Mali, Burkina Faso and Madagascar and Ecuador were investigated for genetic diversity using two AFLP primer combinations. The results revealed a high genetic diversity in the populations studied. The population with greatest genetic diversity was Madagascar (He = 0.2638 and I = 0.4066 and the least diverse was Senegal-Tamba (He = 0.1962 and I = 0.3079. AMOVA (analysis of molecular variance detected the highest proportion of variation within populations (81% of the total molecular variation. This may be attributed to the high level of allogamy observed in this species. The Nei's standard unbiased genetic distance (D between the populations ranged from 0.010 (Senegal-Tamba and Burkina Faso to 0.131 (Mali and Ecuador; the average was 0.063. Analysis of the genetic relationships among the 6 populations using both neighbor-joining cluster analysis and principal component analysis (PCoA showed five clusters with globally, groupings of i most of Burkina Faso and Senegal-Tamba genotypes, ii most of Mali and Senegal-Diobass genotypes , iii most of Madagascar and Ecuador genotypes, and iv some mixings of genotypes with different origins. Considering the distance existing between the different origins there are prospects to develop F1 hybrids. The greatest heterosis might be expected from crossing involving genotypes of cluster I and cluster V which group the more distant genotypes. Such crossing schemes might produce greater success in the production of genetic variability and might maximize the exploitation of heterosis and segregation.

  1. Genotypic diversity of european Phytophthora ramorum isolates based on SSR analysis

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    Kris Van Poucke; Annelies Vercauteren; Martine Maes; Sabine Werres; Kurt Heungens

    2013-01-01

    in Scotland were genotyped using seven microsatellite markers as described by Vercauteren et al. (2010). Thirty multilocus genotypes were identified within the Scottish population, with 51 percent of the isolates belonging to the main European genotype EU1MG1 and 13 unique detected genotypes. Ten of those genotypes were site specific, often represented by...

  2. 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.

  3. Yersinia enterocolitica of porcine origin: carriage of virulence genes and genotypic diversity.

    Science.gov (United States)

    Tadesse, Daniel A; Bahnson, Peter B; Funk, Julie A; Morrow, W E Morgan; Abley, Melanie J; Ponte, Valeria A; Thakur, Siddhartha; Wittum, Thomas; DeGraves, Fred J; Rajala-Schultz, Paivi J; Gebreyes, Wondwossen A

    2013-01-01

    Yersinia enterocolitica is an important foodborne pathogen, and pigs are recognized as a major reservoir and potential source of pathogenic strains to humans. A total of 172 Y. enterocolitica recovered from conventional and antimicrobial-free pig production systems from different geographic regions (North Carolina, Ohio, Michigan, Wisconsin, and Iowa) were investigated to determine their pathogenic significance to humans. Phenotypic and genotypic diversity of the isolates was assessed using antibiogram, serogrouping, and amplified fragment length polymorphism (AFLP). Carriage of chromosomal and plasmid-borne virulence genes were investigated using polymerase chain reaction. A total of 12 antimicrobial resistance patterns were identified. More than two-thirds (67.4%) of Y. enterocolitica were pan-susceptible, and 27.9% were resistant against β-lactams. The most predominant serogroup was O:3 (43%), followed by O:5 (25.6%) and O:9 (4.1%). Twenty-two of 172 (12.8%) isolates were found to carry Yersinia adhesion A (yadA), a virulence gene encoded on the Yersinia virulence plasmid. Sixty-nine (40.1%) isolates were found to carry ail gene. The ystA and ystB genes were detected in 77% and 26.2% of the strains, respectively. AFLP genotyping of isolates showed wide genotypic diversity and were grouped into nine clades with an overall genotypic similarity of 66.8-99.3%. AFLP analysis revealed that isolates from the same production system showed clonal relatedness, while more than one genotype of Y. enterocolitica circulates within a farm.

  4. Genotypic diversity and clonal structure of Erigeron annuus (Asteraceae in Lithuania

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    Tunaitienė, Virginija

    2014-02-01

    Full Text Available This study was conducted to assess the clonal structure and genetic diversity of alien herbaceous plant species Erigeron annuus. The global warming and changes in agriculture practice in the past few decades were favourable for the expansion of this species in Lithuania. We used RAPD and ISSR assays to assess genetic variation within and among 29 populations of E. annuus. A total of 278 molecular markers were revealed. Our study detected reduced level of genetic diversity of invasive populations of E. annuus. Significant differences in DNA polymorphism among populations of E. annuus were also found. Some populations of this species are composed of genetically identical plants, while others were polymorphic. Clonal diversity of study populations ranged from 0.083 to 0.4 for both DNA marker systems. The Simpsons diversity index values ranged from 0.0 to 0.636. The average number of genotypes per population established using both assays was about 1.7. Out of 328 E. annuus individuals only 16 showed unique RAPD and 14 unique ISSR banding patterns. The remaining plants were clones of different size. The most common genotype of E. annuus identified in our study was represented by predominate in nine populations.

  5. Genotypic diversity of root and shoot characteristics of

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    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

  6. Genotypic Diversity and Short-term Response to Shading Stress in a Threatened Seagrass: Does Low Diversity Mean Low Resilience?

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    Suzanna M. Evans

    2017-08-01

    Full Text Available Seagrasses that are predominantly clonal often have low levels of genetic variation within populations and predicting their response to changing conditions requires an understanding of whether genetic variation confers increased resistance to environmental stressors. A higher level of genetic diversity is assumed to benefit threatened species due to the increased likelihood of those populations having genotypes that can persist under environmental change. To test this idea, we conducted an in situ shading experiment with six geographically distinct meadows of the threatened seagrass Posidonia australis that vary in genetic diversity. Different genotypes within meadows varied widely in their physiological and growth responses to reduced light during a simulated short-term turbidity event. The majority of meadows were resistant to the sudden reduction in light availability, but a small subset of meadows with low genotypic diversity were particularly vulnerable to the early effects of shading, showing substantially reduced growth rates after only 3 weeks. Using the photosynthetic performance (maximum quantum yield of known genotypes, we simulated meadows of varying genetic diversity to show that higher diversity can increase meadow resilience to stress by ensuring a high probability of including a high-performing genotype. These results support the hypothesis that complementarity among genotypes enhances the adaptive capacity of a population, and have significant implications for the conservation of declining P. australis meadows close to the species range edge on the east coast of Australia, where the genotypic diversity is low.

  7. Genotypic Diversity and Short-term Response to Shading Stress in a Threatened Seagrass: Does Low Diversity Mean Low Resilience?

    Science.gov (United States)

    Evans, Suzanna M; Vergés, Adriana; Poore, Alistair G B

    2017-01-01

    Seagrasses that are predominantly clonal often have low levels of genetic variation within populations and predicting their response to changing conditions requires an understanding of whether genetic variation confers increased resistance to environmental stressors. A higher level of genetic diversity is assumed to benefit threatened species due to the increased likelihood of those populations having genotypes that can persist under environmental change. To test this idea, we conducted an in situ shading experiment with six geographically distinct meadows of the threatened seagrass Posidonia australis that vary in genetic diversity. Different genotypes within meadows varied widely in their physiological and growth responses to reduced light during a simulated short-term turbidity event. The majority of meadows were resistant to the sudden reduction in light availability, but a small subset of meadows with low genotypic diversity were particularly vulnerable to the early effects of shading, showing substantially reduced growth rates after only 3 weeks. Using the photosynthetic performance (maximum quantum yield) of known genotypes, we simulated meadows of varying genetic diversity to show that higher diversity can increase meadow resilience to stress by ensuring a high probability of including a high-performing genotype. These results support the hypothesis that complementarity among genotypes enhances the adaptive capacity of a population, and have significant implications for the conservation of declining P. australis meadows close to the species range edge on the east coast of Australia, where the genotypic diversity is low.

  8. Genotyping of ancient Mycobacterium tuberculosis strains reveals historic genetic diversity.

    Science.gov (United States)

    Müller, Romy; Roberts, Charlotte A; Brown, Terence A

    2014-04-22

    The evolutionary history of the Mycobacterium tuberculosis complex (MTBC) has previously been studied by analysis of sequence diversity in extant strains, but not addressed by direct examination of strain genotypes in archaeological remains. Here, we use ancient DNA sequencing to type 11 single nucleotide polymorphisms and two large sequence polymorphisms in the MTBC strains present in 10 archaeological samples from skeletons from Britain and Europe dating to the second-nineteenth centuries AD. The results enable us to assign the strains to groupings and lineages recognized in the extant MTBC. We show that at least during the eighteenth-nineteenth centuries AD, strains of M. tuberculosis belonging to different genetic groups were present in Britain at the same time, possibly even at a single location, and we present evidence for a mixed infection in at least one individual. Our study shows that ancient DNA typing applied to multiple samples can provide sufficiently detailed information to contribute to both archaeological and evolutionary knowledge of the history of tuberculosis.

  9. 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

  10. Genotypic diversity of merozoite surface antigen 1 of Babesia bovis within an endemic population.

    Science.gov (United States)

    Lau, Audrey O T; Cereceres, Karla; Palmer, Guy H; Fretwell, Debbie L; Pedroni, Monica J; Mosqueda, Juan; McElwain, Terry F

    2010-08-01

    Multiple genetically distinct strains of a pathogen circulate and compete for dominance within populations of animal reservoir hosts. Understanding the basis for genotypic strain structure is critical for predicting how pathogens respond to selective pressures and how shifts in pathogen population structure can lead to disease outbreaks. Evidence from related Apicomplexans such as Plasmodium, Toxoplasma, Cryptosporidium and Theileria suggests that various patterns of population dynamics exist, including but not limited to clonal, oligoclonal, panmictic and epidemic genotypic strain structures. In Babesia bovis, genetic diversity of variable merozoite surface antigen (VMSA) genes has been associated with disease outbreaks, including in previously vaccinated animals. However, the extent of VMSA diversity within a defined population in an endemic area has not been examined. We analyzed genotypic diversity and temporal change of MSA-1, a member of the VMSA family, in individual infected animals within a reservoir host population. Twenty-eight distinct MSA-1 genotypes were identified within the herd. All genotypically distinct MSA-1 sequences clustered into three groups based on sequence similarity. Two thirds of the animals tested changed their dominant MSA-1 genotypes during a 6-month period. Five animals within the population contained multiple genotypes. Interestingly, the predominant genotypes within those five animals also changed over the 6-month sampling period, suggesting ongoing transmission or emergence of variant MSA-1 genotypes within the herd. This study demonstrated an unexpected level of diversity for a single copy gene in a haploid genome, and illustrates the dynamic genotype structure of B. bovis within an individual animal in an endemic region. Co-infection with multiple diverse MSA-1 genotypes provides a basis for more extensive genotypic shifts that characterizes outbreak strains.

  11. 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

  12. 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.

  13. Analysis of the genetic diversity of four rabbit genotypes using ...

    African Journals Online (AJOL)

    Dr.Ola

    2013-05-15

    May 15, 2013 ... consumption and low cost, it has been widely utilized in genetics analysis in ... isozyme variation among the selected individuals within each rabbit genotype. ... with different embryo survival (Bolet and Theau-Clement, 1994).

  14. 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.

  15. The genetic diversity of hepatitis A genotype I in Bulgaria.

    Science.gov (United States)

    Cella, Eleonora; Golkocheva-Markova, Elitsa N; Trandeva-Bankova, Diljana; Gregori, Giulia; Bruni, Roberto; Taffon, Stefania; Equestre, Michele; Costantino, Angela; Spoto, Silvia; Curtis, Melissa; Ciccaglione, Anna Rita; Ciccozzi, Massimo; Angeletti, Silvia

    2018-01-01

    The purpose of this study was to analyze sequences of hepatitis A virus (HAV) Ia and Ib genotypes from Bulgarian patients to investigate the molecular epidemiology of HAV genotype I during the years 2012 to 2014. Around 105 serum samples were collected by the Department of Virology of the National Center of Infectious and Parasitic Diseases in Bulgaria. The sequenced region encompassed the VP1/2A region of HAV genome. The sequences obtained from the samples were 103. For the phylogenetic analyses, 5 datasets were built to investigate the viral gene in/out flow among distinct HAV subpopulations in different geographic areas and to build a Bayesian dated tree, Bayesian phylogenetic and migration pattern analyses were performed. HAV Ib Bulgarian sequences mostly grouped into a single clade. This indicates that the Bulgarian epidemic is partially compartmentalized. It originated from a limited number of viruses and then spread through fecal-oral local transmission. HAV Ia Bulgarian sequences were intermixed with European sequences, suggesting that an Ia epidemic is not restricted to Bulgaria but can affect other European countries. The time-scaled phylogeny reconstruction showed the root of the tree dating in 2008 for genotype Ib and in 1999 for genotype Ia with a second epidemic entrance in 2003. The Bayesian skyline plot for genotype Ib showed a slow but continuous growth, sustained by fecal-oral route transmission. For genotype Ia, there was an exponential growth followed by a plateau, which suggests better infection control. Bidirectional viral flow for Ib genotype, involving different Bulgarian areas, was observed, whereas a unidirectional flow from Sofia to Ihtiman for genotype Ia was highlighted, suggesting the fecal-oral transmission route for Ia. Copyright © 2017 The Authors. Published by Wolters Kluwer Health, Inc. All rights reserved.

  16. Molecular characterization and genetic diversity of different genotypes of Oryza sativa and Oryza glaberrima

    Directory of Open Access Journals (Sweden)

    Caijin Chen

    2017-11-01

    Conclusions: Genetic diversity studies revealed that 50 rice types were clustered into different subpopulations whereas three genotypes were admixtures. Molecular fingerprinting and 10 specific markers were obtained to identify the 53 rice genotypes. These results can facilitate the potential utilization of sibling species in rice breeding and molecular classification of O. sativa and O. glaberrima germplasms.

  17. Impact of enumeration method on diversity of Escherichia coli genotypes isolated from surface water.

    Science.gov (United States)

    Martin, E C; Gentry, T J

    2016-11-01

    There are numerous regulatory-approved Escherichia coli enumeration methods, but it is not known whether differences in media composition and incubation conditions impact the diversity of E. coli populations detected by these methods. A study was conducted to determine if three standard water quality assessments, Colilert ® , USEPA Method 1603, (modified mTEC) and USEPA Method 1604 (MI), detect different populations of E. coli. Samples were collected from six watersheds and analysed using the three enumeration approaches followed by E. coli isolation and genotyping. Results indicated that the three methods generally produced similar enumeration data across the sites, although there were some differences on a site-by-site basis. The Colilert ® method consistently generated the least diverse collection of E. coli genotypes as compared to modified mTEC and MI, with those two methods being roughly equal to each other. Although the three media assessed in this study were designed to enumerate E. coli, the differences in the media composition, incubation temperature, and growth platform appear to have a strong selective influence on the populations of E. coli isolated. This study suggests that standardized methods of enumeration and isolation may be warranted if researchers intend to obtain individual E. coli isolates for further characterization. This study characterized the impact of three USEPA-approved Escherichia coli enumeration methods on observed E. coli population diversity in surface water samples. Results indicated that these methods produced similar E. coli enumeration data but were more variable in the diversity of E. coli genotypes observed. Although the three methods enumerate the same species, differences in media composition, growth platform, and incubation temperature likely contribute to the selection of different cultivable populations of E. coli, and thus caution should be used when implementing these methods interchangeably for

  18. Lunasin detection in coloured wheat genotype

    Directory of Open Access Journals (Sweden)

    Milan Chňapek

    2016-01-01

    Full Text Available Lunasin is a biologically active protein, composed of 43 amino acid residues. There has been proven many health-promoting effects of lunasin peptide. The most important health benefits include: anti-hypertension, antioxidant activity, cancer prevention or therapy. It was also demonstrated anti-inflammation, hypocholesterolemic activity, anti-obesity and immunomodulation. The focus of our research is to summarize the discovery, characterization and biological activities of lunasin, which will provide a reference for the future development and utilization of lunasin, and a basis for exploring the underlying mechanisms of these health-beneficial functions. Lunasin was first isolated in 1987 at Niigata University School of Medicine in Japan, during the screening of protease inhibitors from soybean seeds. It was subsequently found in other beans, grains and herbal plants, including wheat, barley, rye, triticale. Concentration of lunasin is ranging from 0.013 to 70.5 mg protein lunasin/g of protein. Big step forward in the understanding of the lunasine operating mechanism in the fight against cancer has arisen after study on cloning of the soybean lunasin gene and subsequent transfection into mammalian cells which led to the discovery that the lunasin gene can disrupt mitosis and induce chromosome breakage, ultimately leading to cell apoptosis. The main goal of our work was to evaluate collection of wheat with unusual grain colour for presence of lunasin gene. DNA was extracted by commercial kit and lunasin gene was detected by PCR reaction. Our results showed presence of lunasin gene detected by 3 combinations of 2 sets of primer pair and indicated lunasin peptide presence in cereal grains. These findings are necessary to confirmed by proteome analysis. Normal 0 21 false false false CS JA X-NONE

  19. Genotypic diversity of an invasive plant species promotes litter decomposition and associated processes.

    Science.gov (United States)

    Wang, Xiao-Yan; Miao, Yuan; Yu, Shuo; Chen, Xiao-Yong; Schmid, Bernhard

    2014-03-01

    Following studies that showed negative effects of species loss on ecosystem functioning, newer studies have started to investigate if similar consequences could result from reductions of genetic diversity within species. We tested the influence of genotypic richness and dissimilarity (plots containing one, three, six or 12 genotypes) in stands of the invasive plant Solidago canadensis in China on the decomposition of its leaf litter and associated soil animals over five monthly time intervals. We found that the logarithm of genotypic richness was positively linearly related to mass loss of C, N and P from the litter and to richness and abundance of soil animals on the litter samples. The mixing proportion of litter from two sites, but not genotypic dissimilarity of mixtures, had additional effects on measured variables. The litter diversity effects on soil animals were particularly strong under the most stressful conditions of hot weather in July: at this time richness and abundance of soil animals were higher in 12-genotype litter mixtures than even in the highest corresponding one-genotype litter. The litter diversity effects on decomposition were in part mediated by soil animals: the abundance of Acarina, when used as covariate in the analysis, fully explained the litter diversity effects on mass loss of N and P. Overall, our study shows that high genotypic richness of S. canadensis leaf litter positively affects richness and abundance of soil animals, which in turn accelerate litter decomposition and P release from litter.

  20. Genetic Diversity and Population Structure of F3:6 Nebraska Winter Wheat Genotypes Using Genotyping-By-Sequencing.

    Science.gov (United States)

    Eltaher, Shamseldeen; Sallam, Ahmed; Belamkar, Vikas; Emara, Hamdy A; Nower, Ahmed A; Salem, Khaled F M; Poland, Jesse; Baenziger, Peter S

    2018-01-01

    The availability of information on the genetic diversity and population structure in wheat ( Triticum aestivum L.) breeding lines will help wheat breeders to better use their genetic resources and manage genetic variation in their breeding program. The recent advances in sequencing technology provide the opportunity to identify tens or hundreds of thousands of single nucleotide polymorphism (SNPs) in large genome species (e.g., wheat). These SNPs can be utilized for understanding genetic diversity and performing genome wide association studies (GWAS) for complex traits. In this study, the genetic diversity and population structure were investigated in a set of 230 genotypes (F 3:6 ) derived from various crosses as a prerequisite for GWAS and genomic selection. Genotyping-by-sequencing provided 25,566 high-quality SNPs. The polymorphism information content (PIC) across chromosomes ranged from 0.09 to 0.37 with an average of 0.23. The distribution of SNPs markers on the 21 chromosomes ranged from 319 on chromosome 3D to 2,370 on chromosome 3B. The analysis of population structure revealed three subpopulations (G1, G2, and G3). Analysis of molecular variance identified 8% variance among and 92% within subpopulations. Of the three subpopulations, G2 had the highest level of genetic diversity based on three genetic diversity indices: Shannon's information index ( I ) = 0.494, diversity index ( h ) = 0.328 and unbiased diversity index (uh) = 0.331, while G3 had lowest level of genetic diversity ( I = 0.348, h = 0.226 and uh = 0.236). This high genetic diversity identified among the subpopulations can be used to develop new wheat cultivars.

  1. Pollen diversity, viability and floral structure of some Musa genotypes

    African Journals Online (AJOL)

    Prof. Ogunji

    This is related to sterility due to triploidy, variation in genome size and structure ... Pollen grains are structures that house the male gametophytes generation of .... the mean value for all the genotypes was recorded for PITA 14, an 11TA hybrid.

  2. Genetic diversity among some productive genotypes of tomato ...

    African Journals Online (AJOL)

    HP

    2015-06-03

    Jun 3, 2015 ... weight, cluster VIII had highest number of locules per fruit, fruit yield per plant and yield per hectare and cluster XVII was .... Cluster means for yield and quality contributing characters of 60 genotypes of tomato. Cluster lycopene β- ..... Sharma JP, Singh AK, Satesh K, Sanjeev K. (2009). Identification of.

  3. Estimation of genetic diversity in rice ( Oryza sativa L. ) genotypes ...

    African Journals Online (AJOL)

    Thirty rice genotypes comprising land races, pure lines, somaclones, breeding lines and varieties specifically adapted to costal saline environments were characterized by SSR markers and morphological characters in this study. Out of 35 primers of SSR markers, 28 were found to be polymorphic. The PIC value ranged ...

  4. Genetic diversity of popcorn genotypes using molecular analysis.

    Science.gov (United States)

    Resh, F S; Scapim, C A; Mangolin, C A; Machado, M F P S; do Amaral, A T; Ramos, H C C; Vivas, M

    2015-08-19

    In this study, we analyzed dominant molecular markers to estimate the genetic divergence of 26 popcorn genotypes and evaluate whether using various dissimilarity coefficients with these dominant markers influences the results of cluster analysis. Fifteen random amplification of polymorphic DNA primers produced 157 amplified fragments, of which 65 were monomorphic and 92 were polymorphic. To calculate the genetic distances among the 26 genotypes, the complements of the Jaccard, Dice, and Rogers and Tanimoto similarity coefficients were used. A matrix of Dij values (dissimilarity matrix) was constructed, from which the genetic distances among genotypes were represented in a more simplified manner as a dendrogram generated using the unweighted pair-group method with arithmetic average. Clusters determined by molecular analysis generally did not group material from the same parental origin together. The largest genetic distance was between varieties 17 (UNB-2) and 18 (PA-091). In the identification of genotypes with the smallest genetic distance, the 3 coefficients showed no agreement. The 3 dissimilarity coefficients showed no major differences among their grouping patterns because agreement in determining the genotypes with large, medium, and small genetic distances was high. The largest genetic distances were observed for the Rogers and Tanimoto dissimilarity coefficient (0.74), followed by the Jaccard coefficient (0.65) and the Dice coefficient (0.48). The 3 coefficients showed similar estimations for the cophenetic correlation coefficient. Correlations among the matrices generated using the 3 coefficients were positive and had high magnitudes, reflecting strong agreement among the results obtained using the 3 evaluated dissimilarity coefficients.

  5. Protein landmarks for diversity assessment in wheat genotypes

    African Journals Online (AJOL)

    jai ganesha

    2013-07-17

    Jul 17, 2013 ... of genetic diversity in wheat has been on differences in morphological and ... glutenins, are the main components of gluten, which is the main contributor to the .... However, there was no within variety diversity observed as a ...

  6. 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

  7. Genotypic Diversity for Biomass Accumulation and Shoot-Root Allometry in the Grass Brachypodium distachyon

    Science.gov (United States)

    Jansson, C.; Handakumbura, P. P.; Fortin, D.; Stanfill, B.; Rivas-Ubach, A.

    2017-12-01

    Predicting carbon uptake, assimilation and allocation for current and future biogeographical environments, including climate, is critical for our ability to select and/or design plant genotypes to meet increasing demand for plant biomass going into food, feed and energy production, while at the same time maintain or increase soil organic matter (SOM for soil fertility and carbon storage, and reduce emission of greenhouse gasses. As has been demonstrated for several plant species allometric relationships may differ between plant genotypes. Exploring plant genotypic diversity for biomass accumulation and allometry will potentially enable selection of genotypes with high CO2 assimilation and favorable allocation of recent photosynthate into above-ground and below-ground biomass. We are investigating genotypic diversity for PFTs in natural accessions of the annual C3 grass Brachypodium distachyon under current and future climate scenarios and how genotypic diversity correlates with metabolite profiles in aboveground and below-ground biomass. In the current study, we compare effects from non-stressed and drought conditions on biomass accumulation and shoot-root allometry.

  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. Host-parasite genotypic interactions in the honey bee: the dynamics of diversity.

    Science.gov (United States)

    Evison, Sophie E F; Fazio, Geraldine; Chappell, Paula; Foley, Kirsten; Jensen, Annette B; Hughes, William O H

    2013-07-01

    Parasites are thought to be a major driving force shaping genetic variation in their host, and are suggested to be a significant reason for the maintenance of sexual reproduction. A leading hypothesis for the occurrence of multiple mating (polyandry) in social insects is that the genetic diversity generated within-colonies through this behavior promotes disease resistance. This benefit is likely to be particularly significant when colonies are exposed to multiple species and strains of parasites, but host-parasite genotypic interactions in social insects are little known. We investigated this using honey bees, which are naturally polyandrous and consequently produce genetically diverse colonies containing multiple genotypes (patrilines), and which are also known to host multiple strains of various parasite species. We found that host genotypes differed significantly in their resistance to different strains of the obligate fungal parasite that causes chalkbrood disease, while genotypic variation in resistance to the facultative fungal parasite that causes stonebrood disease was less pronounced. Our results show that genetic variation in disease resistance depends in part on the parasite genotype, as well as species, with the latter most likely relating to differences in parasite life history and host-parasite coevolution. Our results suggest that the selection pressure from genetically diverse parasites might be an important driving force in the evolution of polyandry, a mechanism that generates significant genetic diversity in social insects.

  10. Analysis of the genetic diversity of four rabbit genotypes using ...

    African Journals Online (AJOL)

    Dr.Ola

    2013-05-15

    May 15, 2013 ... diseases or market conditions. A considerable number of genetic diversity studies for several livestock species have been carried out during recent years by research. *Corresponding author. E-mail: ola.galal@agr.kfs.edu.eg or olagalal2002@yahoo.com , Tel/Fax: +2-0479102930. Abbreviations: APRI ...

  11. Microsatellite genotyping reveals high genetic diversity but low ...

    African Journals Online (AJOL)

    JMwacharos

    2016-03-16

    Mar 16, 2016 ... diversity and (2) Investigate population structure and extent of admixture .... to estimate and partition genetic variation within and ... K between 1 and 40 and inferred its most optimal value ... populations of 0.84 ± 0.021 with the lowest mean in ..... on population stratification and the distribution of genetic.

  12. Evaluation of Genetic Diversity of Iris Genotypes (Iris spp Using ISSR

    Directory of Open Access Journals (Sweden)

    seyedeh zeinab attari

    2017-02-01

    Full Text Available Introduction: Some of Iris species are growing in different parts of the Iran as wild species. Iris species have important medicinal and horticultural properties. Understanding of the genetic variation within and between populations is essential for the establishment of effective and efficient methods for conservation of the plants. Genetic variation studies are fundamental for the management and conservation of this species. The use of molecular markers is a powerful tool in the genetic study of populations. The use of DNA marker, such as AFLP, SSR, RAPD and ISSR represents an alternative method in detection of polymorphism. ISSRs are highly variable, require less investment in time, money and labor than other methods. ISSR can generate higher percentages of polymorphic loci than other PCR methods. These can serve as an efficient tool for phylogenetic studies. ISSRs had reported that used in studies of cultivated species to produce genetic linkage maps and to determine the relatedness of lines of agriculturally important species. ISSR analysis involves the PCR amplification of regions between adjacent, inversely oriented microsatellites, using a single simple sequence repeat (SSR motifs (dinucleotide, trinucleotide, tetranucleotide or penta nucleotides. Therefore, little is known about the genetic variability of the Iranian Iris ssp .The objectives of this study were to evaluate genetic diversity among genotypes using ISSR markers and the degree of polymorphism generated from ISSR technique as a pre-requisite for their applicability to population genetics studies in Iris ssp. Materials and Methods: To evaluate genetic variations in some wild Iris genotypes, Iris kopetdaghensis ،Iris songarica and Iris fosteriana were collected from some parts of Khorasan province. Genomic DNA was extracted from young leaves following the cetyltrimethylammonium bromide (CTAB procedure. Extracted DNA concentration was quantified by using the spectrophotometer

  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. Global phylogeography and genetic diversity of the zoonotic tapeworm Echinococcus granulosus sensu stricto genotype G1.

    Science.gov (United States)

    Kinkar, Liina; Laurimäe, Teivi; Acosta-Jamett, Gerardo; Andresiuk, Vanessa; Balkaya, Ibrahim; Casulli, Adriano; Gasser, Robin B; van der Giessen, Joke; González, Luis Miguel; Haag, Karen L; Zait, Houria; Irshadullah, Malik; Jabbar, Abdul; Jenkins, David J; Kia, Eshrat Beigom; Manfredi, Maria Teresa; Mirhendi, Hossein; M'rad, Selim; Rostami-Nejad, Mohammad; Oudni-M'rad, Myriam; Pierangeli, Nora Beatriz; Ponce-Gordo, Francisco; Rehbein, Steffen; Sharbatkhori, Mitra; Simsek, Sami; Soriano, Silvia Viviana; Sprong, Hein; Šnábel, Viliam; Umhang, Gérald; Varcasia, Antonio; Saarma, Urmas

    2018-05-19

    Echinococcus granulosus sensu stricto (s.s.) is the major cause of human cystic echinococcosis worldwide and is listed among the most severe parasitic diseases of humans. To date, numerous studies have investigated the genetic diversity and population structure of E. granulosus s.s. in various geographic regions. However, there has been no global study. Recently, using mitochondrial DNA, it was shown that E. granulosus s.s. G1 and G3 are distinct genotypes, but a larger dataset is required to confirm the distinction of these genotypes. The objectives of this study were to: (i) investigate the distinction of genotypes G1 and G3 using a large global dataset; and (ii) analyse the genetic diversity and phylogeography of genotype G1 on a global scale using near-complete mitogenome sequences. For this study, 222 globally distributed E. granulosus s.s. samples were used, of which 212 belonged to genotype G1 and 10 to G3. Using a total sequence length of 11,682 bp, we inferred phylogenetic networks for three datasets: E. granulosus s.s. (n = 222), G1 (n = 212) and human G1 samples (n = 41). In addition, the Bayesian phylogenetic and phylogeographic analyses were performed. The latter yielded several strongly supported diffusion routes of genotype G1 originating from Turkey, Tunisia and Argentina. We conclude that: (i) using a considerably larger dataset than employed previously, E. granulosus s.s. G1 and G3 are indeed distinct mitochondrial genotypes; (ii) the genetic diversity of E. granulosus s.s. G1 is high globally, with lower values in South America; and (iii) the complex phylogeographic patterns emerging from the phylogenetic and geographic analyses suggest that the current distribution of genotype G1 has been shaped by intensive animal trade. Copyright © 2018 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.

  15. Assessment of RAPD Markers to Analyse the Genetic Diversity among Sunflower (Helianthus annuus L. Genotypes

    Directory of Open Access Journals (Sweden)

    Ali Raza

    2018-02-01

    Full Text Available Genetic diversity estimation among different species is an important tool for genetic improvement to maximize the yield, desirable quality, wider adaptation, pest and insect resistance that ultimately boosting traditional plant breeding methods. The most efficient way of diversity estimation is application of molecular markers. In this study, twenty random amplified polymorphic DNA (RAPD primers were utilized to estimate the genetic diversity between ten sunflower genotypes. Overall 227 bands were amplified by 20 primers with an average of 11.35 bands per primer. RAPD data showed 86.34% polymorophic bands and 13.65% of monomorophic bands. Genetic similarity was ranged from 50.22% to 87.22%. The lowest similarity (50.22% was observed between FH-352 and FH-359 and the maximum similarity 87.22% was observed between A-23 and G-46. Polymorphic information content (PIC values were varying from 0.05 to 0.12 with a mean of 0.09. Cluster analysis based on RAPD results displayed two major distinct groups 1 and 2. Group-2 contains FH-352 which was the most diverse genotype, while group-1 consists of few sub groups with all other genotypes. Ample diversity was found in all the genotypes. Present study reveals novel information about sunflower genome which can be used in future studies for sunflower improvement.

  16. SSR based genetic diversity of pigmented and aromatic rice (Oryza sativa L.) genotypes of the western Himalayan region of India.

    Science.gov (United States)

    Ashraf, Humaira; Husaini, Amjad M; Ashraf Bhat, M; Parray, G A; Khan, Salim; Ganai, Nazir A

    2016-10-01

    A set of 24 of SSR markers were used to estimate the genetic diversity in 16 rice genotypes found in Western Himalayas of Kashmir and Himachal Pradesh, India. The level of polymorphism among the genotypes of rice was evaluated from the number of alleles and PIC value for each of the 24 SSR loci. A total of 68 alleles were detected across the 16 genotypes through the use of these 24 SSR markers The number of alleles per locus generated varied from 2 (RM 338, RM 452, RM 171) to 6 (RM 585, RM 249, RM 481, RM 162). The PIC values varied from 0.36 (RM 1) to 0.86 (RM 249) with an average of 0.62 per locus. Based on information generated, the genotypes got separated in six different clusters. Cluster 1 comprised of 4 genotypes viz; Zag 1, Zag 13, Pusa sugandh 3, and Zag 14, separated from each other at a similarity value of 0.40. Cluster second comprised of 3 landraces viz; Zag 2. Zag 4 and Zag10 separated from each other at a similarity value of 0.45. Cluster third comprised of 3 genotypes viz; Grey rice, Mushk budji and Kamad separated from each other at a similarity value of 0.46. Cluster fourth had 2 landraces viz; Kawa kreed and Loual anzul, and was not sub clustered. Fifth cluster had 3 genotypes viz; Zag 12, Purple rice and Jhelum separated from each other at a similarity value of 0.28. Cluster 6 comprised of a single popular variety i.e. Shalimar rice 1 with independent lineage.

  17. Genetic diversity and phylogenetic relationship in different genotypes of cotton for future breeding

    Directory of Open Access Journals (Sweden)

    Jehan

    2017-11-01

    Full Text Available Background: To make the plants well adapted and more resistant to diseases and other environmental stresses there is always a need to improve the quality of plant’s genome i.e. to increase its genetic diversity. Methods: In the present study six variety and six lines of cotton were investigated for their genetic diversity and phylogenetic relationship. For this purpose 35 different RAPD primers obtained from the Gene Link Technologies, USA were used. Results: Among 35 RAPD primers, 13 primers produced reproducible PCR bands while the rest failed to show any amplification product. Our results indicated that the total count of the reproducible bands was 670 and polymorphic loci were counted to be 442 which constitute 66% of total loci. Phylogenetic analysis revealed two major groups each consists of 7 and 5 genotypes respectively. Genotypes Lp1 and Tp4 were placed at maximum genetic distance and in separate groups and could be utilized for future cotton breeding. Conclusions: RAPD analysis is a cheaper and time saving technique for the determination of genetic diversity of different cotton genotypes. Cotton genotype Lp1 and Tp4 could be the best candidates for future breeding programs as both genotypes are genetically distant from each other.

  18. DNA landmarks for genetic relatedness and diversity assessment in Pakistani wheat genotypes using RAPD markers

    International Nuclear Information System (INIS)

    Siddiqui, M.F.; Iqbal, S.; Naz, N.; Khan, S.; Erum, S.

    2010-01-01

    DNA profiles from 10 Pakistani wheat genotypes were evaluated for diversity assessment based on RAPD markers. A total of 79 DNA fragments were generated by 10 RAPD primers, with an average of 7.9 bands primer-1. Of these, 64 fragments (81%) were polymorphic among 10 genotypes. Genetic diversity was evaluated via UPGMA cluster analysis by constructing dendrogram, which were used for the calculation of similarity coefficients between these genotypes. The greatest similarity (95%) was observed between PR-94 and PR-95, whereas PR-96 with PR-90 showed the lowest similarity (60%). Adoption of this technology would be useful to the plant protection regulatory systems, especially for plant variety identification and registration of new plant varieties, breeding programs and protection purposes. (author)

  19. DNA landmarks for genetic relatedness and diversity assessment in Pakistani wheat genotypes using RAPD markers

    Energy Technology Data Exchange (ETDEWEB)

    Siddiqui, M F; Iqbal, S; Naz, N; Khan, S [Federal Seed Certification and Registration Dept., Islamabad (Pakistan); Erum, S [National Agricultural Research Centre, Islamabad (Pakistan). Plant Genetic Resources Inst.

    2010-04-15

    DNA profiles from 10 Pakistani wheat genotypes were evaluated for diversity assessment based on RAPD markers. A total of 79 DNA fragments were generated by 10 RAPD primers, with an average of 7.9 bands primer-1. Of these, 64 fragments (81%) were polymorphic among 10 genotypes. Genetic diversity was evaluated via UPGMA cluster analysis by constructing dendrogram, which were used for the calculation of similarity coefficients between these genotypes. The greatest similarity (95%) was observed between PR-94 and PR-95, whereas PR-96 with PR-90 showed the lowest similarity (60%). Adoption of this technology would be useful to the plant protection regulatory systems, especially for plant variety identification and registration of new plant varieties, breeding programs and protection purposes. (author)

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

    Directory of Open Access Journals (Sweden)

    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.

  1. Genetic diversity in South African maize ( Zea mays L.) genotypes as ...

    African Journals Online (AJOL)

    One thousand and forty three (1043) maize genotypes including white and yellow maize inbred lines as well as hybrids from the public germplasm collection were characterized with 80 microsatellite markers distributed throughout the genome. A total of 1874 alleles were amplified and used in the genetic diversity analysis.

  2. 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...

  3. Genotypic and phenotypic diversity among Bacillus species isolated ...

    African Journals Online (AJOL)

    DIRECTOR

    2013-03-20

    Mar 20, 2013 ... 1Department of Food Engineering and Quality Control, University Institute of Technology,. P. O. Box 454 Ngaoundere, Cameroon. 2Department of ... potentially help in maintaining consumer health. (Mohamadou et ... used to this effect. Phenotypic ..... However, the detection of B. cereus, a food poisoning,.

  4. Microsatellite genotyping and genome-wide single nucleotide polymorphism-based indices of Plasmodium falciparum diversity within clinical infections.

    Science.gov (United States)

    Murray, Lee; Mobegi, Victor A; Duffy, Craig W; Assefa, Samuel A; Kwiatkowski, Dominic P; Laman, Eugene; Loua, Kovana M; Conway, David J

    2016-05-12

    In regions where malaria is endemic, individuals are often infected with multiple distinct parasite genotypes, a situation that may impact on evolution of parasite virulence and drug resistance. Most approaches to studying genotypic diversity have involved analysis of a modest number of polymorphic loci, although whole genome sequencing enables a broader characterisation of samples. PCR-based microsatellite typing of a panel of ten loci was performed on Plasmodium falciparum in 95 clinical isolates from a highly endemic area in the Republic of Guinea, to characterize within-isolate genetic diversity. Separately, single nucleotide polymorphism (SNP) data from genome-wide short-read sequences of the same samples were used to derive within-isolate fixation indices (F ws), an inverse measure of diversity within each isolate compared to overall local genetic diversity. The latter indices were compared with the microsatellite results, and also with indices derived by randomly sampling modest numbers of SNPs. As expected, the number of microsatellite loci with more than one allele in each isolate was highly significantly inversely correlated with the genome-wide F ws fixation index (r = -0.88, P 10 % had high correlation (r > 0.90) with the index derived using all SNPs. Different types of data give highly correlated indices of within-infection diversity, although PCR-based analysis detects low-level minority genotypes not apparent in bulk sequence analysis. When whole-genome data are not obtainable, quantitative assay of ten or more SNPs can yield a reasonably accurate estimate of the within-infection fixation index (F ws).

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

    Directory of Open Access Journals (Sweden)

    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.

  6. 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.

  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. Copyright © 2015 Elsevier B.V. All rights reserved.

  8. Toxoplasma gondii seroprevalence and genotype diversity in select wildlife species from the southeastern United States

    Directory of Open Access Journals (Sweden)

    Richard W. Gerhold

    2017-10-01

    Full Text Available Abstract Background Toxoplasma gondii is a widespread protozoan parasite that infects humans and other animals. Previous studies indicate some genotypes of T. gondii are more frequently isolated in wildlife than agricultural animals, suggesting a wild/feral animal diversity model. To determine seroprevalence and genetic diversity of T. gondii in southeastern US wildlife, we collected sera from 471 wild animals, including 453 mammals and 18 birds, between 2011 and 2014. These serum samples were assayed for T. gondii infection using the modified agglutination test (MAT. Heart or tongue tissues from 66 seropositive animals were bioassayed in mice and 19 isolates were obtained. The isolated parasites were genotyped by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP method employing 10 genetic markers. Results One hundred and ninety-six of 471 samples (41.6% had a titer ≥1:32 and were considered positive for T. gondii infection. Of 453 mammals, 195 (43% were seropositive, whereas only one (5.6% of 18 birds was seropositive. The seroprevalence in mammals was significantly higher than in the birds. Mammalian hosts with adequate samples size (≥ 20 comprised white-tailed deer (n = 241, feral hogs (n = 100, raccoons (n = 34 and coyotes (n = 22, with seroprevalences of 41.0%, 51.0%, 50.0% and 72.7%, respectively. Coyotes had significantly higher seroprevalence than the white-tailed deer. Genotyping revealed five distinct genotypes, including the ToxoDB PCR-RFLP genotype #5 (a.k.a type 12 for 15 isolates, genotype #3 (a.k.a. type II for 1 isolate, and genotypes #154, #167 and #216, each for 1 isolate. The results showed moderate to high infection rates of T. gondii in white-tailed deer, feral hogs, raccoons and coyotes. Genotyping results indicated limited genetic diversity and a dominance of genotype #5, which has been reported as a major type in wildlife in North America. Conclusions We conclude that T. gondii

  9. 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

  10. 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.

  11. Appraisal of genetic diversity of different peach cultivars and genotypes through rapd markers

    International Nuclear Information System (INIS)

    Bakht, J.; Jamal, N.; Shafi, M

    2012-01-01

    The present study was amid to investigate the genetic diversity of twenty peach cultivars and genotypes by RAPD primers at the Institute of Biotechnology and Genetic Engineering, KPK Agricultural University Peshawar. The result indicated that fifteen primers (GLCO9, GLC20, GLA20, GLA13, GLB10, GLB20, GLB06, GLB19, GLA19, GLB19, GLD16, GLB15, GLA15, GLB12, GLB11) gave genetic distance among the peach cultivars and genotypes under study by PCR amplification. Average genetic diversity (estimated as genetic distance) ranged between 12 and 58%. The molecular size of most of the bands were from 150 bp to 1000 bp. Based on dendrogram analysis, Khyber 1 and Khyber 2 was grouped in cluster A, and Tex-A6-69 and BY-8-135 in cluster B, Candan and 6A were most closely related cultivars and genotypes among the 20 peach cultivars and genotypes while Lering, Flam crest, Tex x-9, early grand and Floradaking were distinctly grouped when compared with the rest of population. (author)

  12. 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.

  13. Genotype Diversity and Distribution of Orientia tsutsugamushi Causing Scrub Typhus in Thailand

    Science.gov (United States)

    2011-07-01

    Society for Microbiology. All Rights Reserved. Genotype Diversity and Distribution of Orientia tsutsugamushi Causing Scrub Typhus in Thailand" Toon...2011 Scrub typhus , caused by antigenically disparate isolates of Orientia tsutsugamushi, is a widely distributed mite-borne human disease in the Asia...evaluation of scrub typhus - specific diagnostic assays and vaccines. Using indirect immunoHuorescence assays (IF A) and PCR assays, 0. tsutsugamushi

  14. The genotypic diversity and lipase production of some thermophilic bacilli from different genera

    OpenAIRE

    Koc, Melih; Cokmus, Cumhur; Cihan, Arzu Coleri

    2015-01-01

    Abstract Thermophilic 32 isolates and 20 reference bacilli were subjected to Rep-PCR and ITS-PCR fingerprinting for determination of their genotypic diversity, before screening lipase activities. By these methods, all the isolates and references could easily be differentiated up to subspecies level from each other. In screening assay, 11 isolates and 7 references were found to be lipase producing. Their extracellular lipase activities were measured quantitatively by incubating in both tributy...

  15. Genotypic and Phenotypic Diversity Does Not Affect Productivity and Drought Response in Competitive Stands of Trifolium repens.

    Science.gov (United States)

    Huber, Heidrun; During, Heinjo J; Bruine de Bruin, Fabienne; Vermeulen, Peter J; Anten, Niels P R

    2016-01-01

    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 affected by soil

  16. 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...

  17. Human population structure detection via multilocus genotype clustering

    Directory of Open Access Journals (Sweden)

    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.

  18. 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.

  19. Genotypic diversity of Coxiella burnetii in the 2007-2010 Q fever outbreak episodes in The Netherlands

    NARCIS (Netherlands)

    Tilburg, Jeroen J. H. C.; Rossen, John W. A.; van Hannen, Erik J.; Melchers, Willem J. G.; Hermans, Mirjam H. A.; van de Bovenkamp, Jeroen; Roest, Hendrik Jan I. J.; de Bruin, Arnout; Nabuurs-Franssen, Marrigje H.; Horrevorts, Alphons M.; Klaassen, Corne H. W.

    The genotypic diversity of Coxiella burnetii in clinical samples obtained from the Dutch Q fever outbreak episodes of 2007-2010 was determined by using a 6-locus variable-number tandem repeat analysis panel. The results are consistent with the introduction of one founder genotype that is gradually

  20. Evaluation of genetic diversity among soybean (Glycine max) genotypes using univariate and multivariate analysis.

    Science.gov (United States)

    Oliveira, M M; Sousa, L B; Reis, M C; Silva Junior, E G; Cardoso, D B O; Hamawaki, O T; Nogueira, A P O

    2017-05-31

    The genetic diversity study has paramount importance in breeding programs; hence, it allows selection and choice of the parental genetic divergence, which have the agronomic traits desired by the breeder. This study aimed to characterize the genetic divergence between 24 soybean genotypes through their agronomic traits, using multivariate clustering methods to select the potential genitors for the promising hybrid combinations. Six agronomic traits evaluated were number of days to flowering and maturity, plant height at flowering and maturity, insertion height of the first pod, and yield. The genetic divergence evaluated by multivariate analysis that esteemed first the Mahalanobis' generalized distance (D 2 ), then the clustering using Tocher's optimization methods, and then the unweighted pair group method with arithmetic average (UPGMA). Tocher's optimization method and the UPGMA agreed with the groups' constitution between each other, the formation of eight distinct groups according Tocher's method and seven distinct groups using UPGMA. The trait number of days for flowering (45.66%) was the most efficient to explain dissimilarity between genotypes, and must be one of the main traits considered by the breeder in the moment of genitors choice in soybean-breeding programs. The genetic variability allowed the identification of dissimilar genotypes and with superior performances. The hybridizations UFU 18 x UFUS CARAJÁS, UFU 15 x UFU 13, and UFU 13 x UFUS CARAJÁS are promising to obtain superior segregating populations, which enable the development of more productive genotypes.

  1. 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.

  2. Genetic diversity within and among two-spotted spider mite resistant and susceptible common bean genotypes

    Directory of Open Access Journals (Sweden)

    Zeinab YOUSEFI

    2017-12-01

    Full Text Available Two-spotted spider mite (Tetranychus urticae C. L. Koch, 1836, is one of the most destructive herbivores of common bean. Very little is known about the diversity among resistant sources in this crop. The present study was conducted to characterize 22 resistant and susceptible common bean genotypes by 8 Simple Sequence Repeats (SSRs and 8 Random Amplified Polymorphic DNA (RAPD markers. These SSR and RAPD primers produced 100 % and 81.8 % polymorphic bands. Based on RAPD fingerprints and SSR profiles, pairwise genetic similarity ranged from 0.0 to 0.857 and from 0.125 to 1, respectively. The resistant and susceptible common bean accessions were grouped together in the dendrograms generated from RAPD and SSR clustering analyses. The results indicate that RAPD and SSR analysis could be successfully used for the estimation of genetic diversity among genotypes. SSR markers could group genotypes according to their resistibility and susceptibility to the spotted spider mite but RAPD could not. Therefore, the SSR markers can facilitate the development of resistant common bean cultivars through breeding programs against T. urticae.

  3. Diversity among Modern Tomato Genotypes at Different Levels in Fresh-Market Breeding

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    Krishna Bhattarai

    2018-01-01

    Full Text Available Cultivated tomato has been in existence for about 400 years and breeding activities have been conducted for only eight decades. However, more than 10,000 tomato cultivars have already been developed. Ninety-one tomato genotypes were characterized for twenty-one morphological traits using developmental, vegetative, and fruit traits. Correlation, principal component, and cluster analysis between the traits were carried out. Higher correlations between fruit traits including fruit shape, fruit size, and fruit types were observed. These correlations indicate that specific fruit types require specific traits like branched inflorescence and a greater number of fruits per inflorescence are beneficial only for smaller fruit sizes like cherry and grape tomatoes. Contrastingly, traits like determinate growth habit and fruit maturity are preferred in all fruit types of tomato for better cultivation practices and longer production duration and hence showed lower correlations. Principal component analysis clustered tomato genotypes into three main clusters with multiple subgroups. Similar tomato genotypes were placed into one or more clusters confirming the results from correlation analysis. Involvement of private breeding programs in cultivar development has increased the competition on introgression of novel and desired traits across new cultivars. Understanding the diversity present in modern cultivars and potential traits identification in related wild species can enhance tomato diversity and improve quality and production.

  4. Genotypic and phenotypic diversity of Bacillus spp. isolated from steel plant waste

    Directory of Open Access Journals (Sweden)

    Chartone-Souza Edmar

    2008-10-01

    Full Text Available Abstract Background Molecular studies of Bacillus diversity in various environments have been reported. However, there have been few investigations concerning Bacillus in steel plant environments. In this study, genotypic and phenotypic diversity and phylogenetic relationships among 40 bacterial isolates recovered from steel plant waste were investigated using classical and molecular methods. Results 16S rDNA partial sequencing assigned all the isolates to the Bacillus genus, with close genetic relatedness to the Bacillus subtilis and Bacillus cereus groups, and to the species Bacillus sphaericus. tDNA-intergenic spacer length polymorphisms and the 16S–23S intergenic transcribed spacer region failed to identify the isolates at the species level. Genomic diversity was investigated by molecular typing with rep (repetitive sequence based PCR using the primer sets ERIC2 (enterobacterial repetitive intergenic consensus, (GTG5, and BOXAIR. Genotypic fingerprinting of the isolates reflected high intraspecies and interspecies diversity. Clustering of the isolates using ERIC-PCR fingerprinting was similar to that obtained from the 16S rRNA gene phylogenetic tree, indicating the potential of the former technique as a simple and useful tool for examining relationships among unknown Bacillus spp. Physiological, biochemical and heavy metal susceptibility profiles also indicated considerable phenotypic diversity. Among the heavy metal compounds tested Zn, Pb and Cu were least toxic to the bacterial isolates, whereas Ag inhibited all isolates at 0.001 mM. Conclusion Isolates with identical 16S rRNA gene sequences had different genomic fingerprints and differed considerably in their physiological capabilities, so the high levels of phenotypic diversity found in this study are likely to have ecological relevance.

  5. Identification of genetically diverse genotypes for photoperiod insensitivity in soybean using RAPD markers.

    Science.gov (United States)

    Singh, R K; Bhatia, V S; Yadav, Sanjeev; Athale, Rashmi; Lakshmi, N; Guruprasad, K N; Chauhan, G S

    2008-10-01

    Most of the Indian soybean varieties were found to be highly sensitive to photoperiod, which limits their cultivation in only localized area. Identification of genetically diverse source of photoperiod insensitive would help to broaden the genetic base for this trait. Present study was undertaken with RAPD markers for genetic diversity estimation in 44 accessions of soybean differing in response to photoperiod sensitivity. The selected twenty-five RAPD primers produced a total of 199 amplicons, which generated 89.9 % polymorphism. The number of amplification products ranged from 2 to 13 for different primers. The polymorphism information content ranged from 0.0 for monomorphic loci to 0.5 with an average of 0.289. Genetic diversity between pairs of genotypes was 37.7% with a range of 3.9 to 71.6%. UPGMA cluster analysis placed all the accessions of soybean into four major clusters. No discernable geographical patterns were observed in clustering however; the smaller groups corresponded well with pedigree. Mantel's test (r = 0.915) indicates very good fit for clustering pattern. Two genotypes, MACS 330 and 111/2/1939 made a very divergent group from other accessions of soybean and highly photoperiod insensitive that may be potential source for broadening the genetic base of soybean for this trait.

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

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    Mathew A Beale

    Full Text Available 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

  7. Genetic variation of european maize genotypes (zea mays l. Detected using ssr markers

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    Martin Vivodík

    2017-01-01

    Full Text Available The SSR molecular markers were used to assess genetic diversity in 40 old European maize genotypes. Ten SSR primers revealed a total of 65 alleles ranging from 4 (UMC1060 to 8 (UMC2002 and UMC1155 alleles per locus with a mean value of 6.50 alleles per locus. The PIC values ranged from 0.713 (UMC1060 to 0.842 (UMC2002 with an average value of 0.810 and the DI value ranged from 0.734 (UMC1060 to 0.848 (UMC2002 with an average value of 0.819. 100% of used SSR markers had PIC and DI values higher than 0.7 that means high polymorphism of chosen markers used for analysis. Probability of identity (PI was low ranged from 0.004 (UMC1072 to 0.022 (UMC1060 with an average of 0.008. A dendrogram was constructed from a genetic distance matrix based on profiles of the 10 maize SSR loci using the unweighted pair-group method with the arithmetic average (UPGMA. According to analysis, the collection of 40 diverse accessions of maize was clustered into four clusters. The first cluster contained nine genotypes of maize, while the second cluster contained the four genotypes of maize. The third cluster contained 5 maize genotypes. Cluster 4 contained five genotypes from Hungary (22.73%, two genotypes from Poland (9.10%, seven genotypes of maize from Union of Soviet Socialist Republics (31.81%, six genotypes from Czechoslovakia (27.27%, one genotype from Slovak Republic (4.55% and one genotype of maize is from Yugoslavia (4.55%. We could not distinguish 4 maize genotypes grouped in cluster 4, (Voroneskaja and Kocovska Skora and 2 Hungarian maize genotypes - Feheres Sarga Filleres and Mindszentpusztai Feher, which are genetically the closest.

  8. Protection of macaques with diverse MHC genotypes against a heterologous SIV by vaccination with a deglycosylated live-attenuated SIV.

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    Chie Sugimoto

    Full Text Available HIV vaccine development has been hampered by issues such as undefined correlates of protection and extensive diversity of HIV. We addressed these issues using a previously established SIV-macaque model in which SIV mutants with deletions of multiple gp120 N-glycans function as potent live attenuated vaccines to induce near-sterile immunity against the parental pathogenic SIVmac239. In this study, we investigated the protective efficacy of these mutants against a highly pathogenic heterologous SIVsmE543-3 delivered intravenously to rhesus macaques with diverse MHC genotypes. All 11 vaccinated macaques contained the acute-phase infection with blood viral loads below the level of detection between 4 and 10 weeks postchallenge (pc, following a transient but marginal peak of viral replication at 2 weeks in only half of the challenged animals. In the chronic phase, seven vaccinees contained viral replication for over 80 weeks pc, while four did not. Neutralizing antibodies against challenge virus were not detected. Although overall levels of SIV specific T cell responses did not correlate with containment of acute and chronic viral replication, a critical role of cellular responses in the containment of viral replication was suggested. Emergence of viruses with altered fitness due to recombination between the vaccine and challenge viruses and increased gp120 glycosylation was linked to the failure to control SIV. These results demonstrate the induction of effective protective immune responses in a significant number of animals against heterologous virus by infection with deglycosylated attenuated SIV mutants in macaques with highly diverse MHC background. These findings suggest that broad HIV cross clade protection is possible, even in hosts with diverse genetic backgrounds. In summary, results of this study indicate that deglycosylated live-attenuated vaccines may provide a platform for the elucidation of correlates of protection needed for a

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

    OpenAIRE

    Wadt, Lúcia Helena de Oliveira; Ehringhaus, Christiane; Kageyama, Paulo Yoshio

    2004-01-01

    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 w...

  10. 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.

  11. High Prevalence and Genotypic Diversity of the Human Papillomavirus in Amazonian Women, Brazil

    Directory of Open Access Journals (Sweden)

    Danielle Albuquerque Pires Rocha

    2013-01-01

    Full Text Available The aim of this study was to determine the prevalence of human papillomavirus (HPV in a women population living within the state of Amazonas, Brazil, and to determine the viral genotypes found. The study included 361 sexually active women over 18 years of age. We performed the Pap test and the molecular diagnosis for HPV DNA using polymerase chain reaction (PCR. The amplicons obtained were sequenced in automatic sequencer for genotyping. The presence of HPV DNA was found in 29.1% (105 of the women. Only 321 women presented satisfactory slides for cytological diagnosis, 97.9% (314 had normal cytology (negative for cancer, and 2.1% (7 had abnormal cytology (4 ASCUS, 1 LSIL, and 2 HSIL. The types more frequently found were HPV 16 (58.1% and HPV 58 (20.0%. Additionally, we found more 13 types of HPV. Compared with previous studies in Brazil, our data confirmed a high prevalence and genotypic diversity of HPV in Brazilian women.

  12. 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.

  13. Genetic diversity and distribution of Mycobacterium tuberculosis genotypes in Limpopo, South Africa.

    Science.gov (United States)

    Maguga-Phasha, N T C; Munyai, N S; Mashinya, F; Makgatho, M E; Mbajiorgu, E F

    2017-12-12

    Tuberculosis remains a major health problem and knowledge of the diversity of Mycobacterium tuberculosis strains in specific geographical regions can contribute to the control of the disease. This study describes the genetic profile of M. tuberculosis in five districts of Limpopo Province. A total 487 isolates were collected from the National Health Laboratory Services from all regions/districts of Limpopo Province. Only 215 isolates were confirmed to be M. tuberculosis by Bactec Mycobacterium Growth Indicator Tube 960® and Rhodamine-Auramine staining. Isolates were subcultured on Löwenstein-Jensen medium agar slants to validate purity. They were spoligotyped and data analysed using the international spoligotyping database 4 (SpolDB4). Of the 215 isolates, 134 (62.3%) were genotyped into 21 genotype families while 81 (37.7%) were orphans. The 81 orphans were further subjected to resolution employing SpolDB3/RIM. Overall, the study revealed a high diversity of strains of 32 predominantly the non-Beijing lineages: the LAM- LAM3 (9.8%), LAM9 (4.7%) and LAM11- ZWE (3.3%), the T-T1(15.0%), T2 (0.9%), T2-T3 (1.4%), the CAS-CAS1-Delhi 5 (1.9%) and CAS1-KILI (1.4%) the MANU2 (1.4%), U (0.5%), X-X1(1.4%), X3 (1.9%), S (9.8%), CAS (1.4%), LAM7(0.9%), T3(0.5%), LAM8(4.7%), T4(1.4%), X2(0.4%), AI5(1.9%), LAM1(0.5%), FAMILY33 (1.9%), EAI4(1.4%), M. microti (1.9%). The Beijing and Beijing-like families were (14.9%) and (0.9%), respectively. A total of 28(13%) clusters and 77(36%) unique cases were identified. Beijing strain (SIT 1) formed the biggest cluster constituting 14%, followed by LAM3 (SIT 33), T1 (SIT 53) and LAM4 (SIT 811) with 7%, 5.1% and 2.8%, respectively. The Beijing family was the only genotype found in all the five districts and was predominant in Mopani (18.8%), Sekhukhune (23.7%) and Vhembe (23.3%). Dominant genotypes in Capricorn and Waterberg were LAM3 (11.9%) and T1 (13.3%), respectively. A wide diversity of lineages was demonstrated at district level. A

  14. Genotypic and Phenotypic Diversity of Cryptococcus gattii VGII Clinical Isolates and Its Impact on Virulence

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    Vanessa A. Barcellos

    2018-02-01

    Full Text Available The Cryptococcus gattii species complex harbors the main etiological agents of cryptococcosis in immunocompetent patients. C. gattii molecular type VGII predominates in the north and northeastern regions of Brazil, leading to high morbidity and mortality rates. C. gattii VGII isolates have a strong clinical relevance and phenotypic variations. These phenotypic variations among C. gattii species complex isolates suggest that some strains are more virulent than others, but little information is available related to the pathogenic properties of those strains. In this study, we analyzed some virulence determinants of C. gattii VGII strains (CG01, CG02, and CG03 isolated from patients in the state of Piauí, Brazil. The C. gattii R265 VGIIa strain, which was isolated from the Vancouver outbreak, differed from C. gattii CG01, CG02 and CG03 isolates (also classified as VGII when analyzed the capsular dimensions, melanin production, urease activity, as well as the glucuronoxylomannan (GXM secretion. Those differences directly reflected in their virulence potential. In addition, CG02 displayed higher virulence compared to R265 (VGIIa strain in a cryptococcal murine model of infection. Lastly, we examined the genotypic diversity of these strains through Multilocus Sequence Type (MLST and one new subtype was described for the CG02 isolate. This study confirms the presence and the phenotypic and genotypic diversity of highly virulent strains in the Northeast region of Brazil.

  15. Genotyping-by-Sequencing Analysis for Determining Population Structure of Finger Millet Germplasm of Diverse Origins

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    Anil Kumar

    2016-07-01

    Full Text Available Finger millet [ (L. Gaertn.] is grown mainly by subsistence farmers in arid and semiarid regions of the world. To broaden its genetic base and to boost its production, it is of paramount importance to characterize and genotype the diverse gene pool of this important food and nutritional security crop. However, as a result of nonavailability of the genome sequence of finger millet, the progress could not be made in realizing the molecular basis of unique qualities of the crop. In the present investigation, attempts have been made to characterize the genetically diverse collection of 113 finger millet accessions through whole-genome genotyping-by-sequencing (GBS, which resulted in a genome-wide set of 23,000 single-nucleotide polymorphisms (SNPs segregating across the entire collection and several thousand SNPs segregating within every accession. A model-based population structure analysis reveals the presence of three subpopulations among the finger millet accessions, which are in parallel with the results of phylogenetic analysis. The observed population structure is consistent with the hypothesis that finger millet was domesticated first in Africa, and from there it was introduced to India some 3000 yr ago. A total of 1128 gene ontology (GO terms were assigned to SNP-carrying genes for three main categories: biological process, cellular component, and molecular function. Facilitated access to high-throughput genotyping and sequencing technologies are likely to improve the breeding process in developing countries, and as such, this data will be very useful to breeders who are working for the genetic improvement of finger millet.

  16. Genotyping-by-Sequencing Analysis for Determining Population Structure of Finger Millet Germplasm of Diverse Origins.

    Science.gov (United States)

    Kumar, Anil; Sharma, Divya; Tiwari, Apoorv; Jaiswal, J P; Singh, N K; Sood, Salej

    2016-07-01

    Finger millet [ (L.) Gaertn.] is grown mainly by subsistence farmers in arid and semiarid regions of the world. To broaden its genetic base and to boost its production, it is of paramount importance to characterize and genotype the diverse gene pool of this important food and nutritional security crop. However, as a result of nonavailability of the genome sequence of finger millet, the progress could not be made in realizing the molecular basis of unique qualities of the crop. In the present investigation, attempts have been made to characterize the genetically diverse collection of 113 finger millet accessions through whole-genome genotyping-by-sequencing (GBS), which resulted in a genome-wide set of 23,000 single-nucleotide polymorphisms (SNPs) segregating across the entire collection and several thousand SNPs segregating within every accession. A model-based population structure analysis reveals the presence of three subpopulations among the finger millet accessions, which are in parallel with the results of phylogenetic analysis. The observed population structure is consistent with the hypothesis that finger millet was domesticated first in Africa, and from there it was introduced to India some 3000 yr ago. A total of 1128 gene ontology (GO) terms were assigned to SNP-carrying genes for three main categories: biological process, cellular component, and molecular function. Facilitated access to high-throughput genotyping and sequencing technologies are likely to improve the breeding process in developing countries, and as such, this data will be very useful to breeders who are working for the genetic improvement of finger millet. Copyright © 2016 Crop Science Society of America.

  17. High-throughput genotyping for species identification and diversity assessment in germplasm collections.

    Science.gov (United States)

    Mason, Annaliese S; Zhang, Jing; Tollenaere, Reece; Vasquez Teuber, Paula; Dalton-Morgan, Jessica; Hu, Liyong; Yan, Guijun; Edwards, David; Redden, Robert; Batley, Jacqueline

    2015-09-01

    Germplasm collections provide an extremely valuable resource for breeders and researchers. However, misclassification of accessions by species often hinders the effective use of these collections. We propose that use of high-throughput genotyping tools can provide a fast, efficient and cost-effective way of confirming species in germplasm collections, as well as providing valuable genetic diversity data. We genotyped 180 Brassicaceae samples sourced from the Australian Grains Genebank across the recently released Illumina Infinium Brassica 60K SNP array. Of these, 76 were provided on the basis of suspected misclassification and another 104 were sourced independently from the germplasm collection. Presence of the A- and C-genomes combined with principle components analysis clearly separated Brassica rapa, B. oleracea, B. napus, B. carinata and B. juncea samples into distinct species groups. Several lines were further validated using chromosome counts. Overall, 18% of samples (32/180) were misclassified on the basis of species. Within these 180 samples, 23/76 (30%) supplied on the basis of suspected misclassification were misclassified, and 9/105 (9%) of the samples randomly sourced from the Australian Grains Genebank were misclassified. Surprisingly, several individuals were also found to be the product of interspecific hybridization events. The SNP (single nucleotide polymorphism) array proved effective at confirming species, and provided useful information related to genetic diversity. As similar genomic resources become available for different crops, high-throughput molecular genotyping will offer an efficient and cost-effective method to screen germplasm collections worldwide, facilitating more effective use of these valuable resources by breeders and researchers. © 2015 John Wiley & Sons Ltd.

  18. Genetic diversity in soybean genotypes using phenotypic characters and enzymatic markers.

    Science.gov (United States)

    Zambiazzi, E V; Bruzi, A T; Sales, A P; Borges, I M M; Guilherme, S R; Zuffo, A M; Lima, J G; Ribeiro, F O; Mendes, A E S; Godinho, S H M; Carvalho, M L M

    2017-09-21

    The objective of this study was to evaluate the genetic diversity of soybean cultivars by adopting phenotypic traits and enzymatic markers, the relative contribution of agronomic traits to diversity, as well as diversity between the level of technology used in soybean cultivars and genetic breeding programs in which cultivars were inserted. The experiments were conducted on the field at the Center for Scientific and Technological Development in crop-livestock production and the Electrophoresis Laboratory of Lavras Federal University. The agronomic traits adopted were grain yield, plant height, first legume insertion, plant lodging, the mass of one thousand seeds, and days for complete maturation, in which the Euclidean distance, grouped by Tocher and UPGMA criteria, was obtained. After electrophorese gels for enzymatic systems, dehydrogenase alcohol, esterase, superoxide dismutase, and peroxidase were performed. The genetic similarity estimative was also obtained between genotypes by the Jaccard coefficient with subsequent grouping by the UPGMA method. The formation of two groups was shown using phenotypic characters in the genetic diversity study and individually discriminating the cultivar 97R73 RR. The character with the greatest contribution to the genetic divergence was grain yield with contribution higher than 90.0%. To obtain six different groups, individually discriminating the cultivars CG 8166 RR, FPS Jupiter RR, and BRS MG 780 RR, enzymatic markers were used. Cultivars carrying the RR technology presented more divergence than conventional cultivars and IPRO cultivars.

  19. 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

  20. Genetic Diversity and Geographic Population Structure of Bovine Neospora caninum Determined by Microsatellite Genotyping Analysis

    Science.gov (United States)

    Regidor-Cerrillo, Javier; Díez-Fuertes, Francisco; García-Culebras, Alicia; Moore, Dadín P.; González-Warleta, Marta; Cuevas, Carmen; Schares, Gereon; Katzer, Frank; Pedraza-Díaz, Susana; Mezo, Mercedes; Ortega-Mora, Luis M.

    2013-01-01

    The cyst-forming protozoan parasite Neospora caninum is one of the main causes of bovine abortion worldwide and is of great economic importance in the cattle industry. Recent studies have revealed extensive genetic variation among N . caninum isolates based on microsatellite sequences (MSs). MSs may be suitable molecular markers for inferring the diversity of parasite populations, molecular epidemiology and the basis for phenotypic variations in N . caninum , which have been poorly defined. In this study, we evaluated nine MS markers using a panel of 11 N . caninum -derived reference isolates from around the world and 96 N . caninum bovine clinical samples and one ovine clinical sample collected from four countries on two continents, including Spain, Argentina, Germany and Scotland, over a 10-year period. These markers were used as molecular tools to investigate the genetic diversity, geographic distribution and population structure of N . caninum . Multilocus microsatellite genotyping based on 7 loci demonstrated high levels of genetic diversity in the samples from all of the different countries, with 96 microsatellite multilocus genotypes (MLGs) identified from 108 N . caninum samples. Geographic sub-structuring was present in the country populations according to pairwise F ST. Principal component analysis (PCA) and Neighbor Joining tree topologies also suggested MLG segregation partially associated with geographical origin. An analysis of the MLG relationships, using eBURST, confirmed that the close genetic relationship observed between the Spanish and Argentinean populations may be the result of parasite migration (i.e., the introduction of novel MLGs from Spain to South America) due to cattle movement. The eBURST relationships also revealed genetically different clusters associated with the abortion. The presence of linkage disequilibrium, the co-existence of specific MLGs to individual farms and eBURST MLG relationships suggest a predominant clonal

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

    Directory of Open Access Journals (Sweden)

    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.

  2. detection of the predominant microcystin-producing genotype of ...

    African Journals Online (AJOL)

    use

    2011-12-21

    Dec 21, 2011 ... genotypes of MC-producing cyanobacteria in Mozambique. Polymerase chain ...... 1(7): 359-366. Pan H, Song L, Liu Y, Börner T (2002). ... Miles CO (2009). A convenient and cost-effective method for monitoring marine algal ...

  3. RT-PCR for detection of all seven genotypes of Lyssavirus genus.

    Science.gov (United States)

    Vázquez-Morón, S; Avellón, A; Echevarría, J E

    2006-08-01

    The Lyssavirus genus includes seven species or genotypes named 1-7. Rabies genotypes correlate with geographical distribution and specific hosts. Co-circulation of different lyssaviruses, imported cases, and the presence of unknown viruses, such as Aravan, Khujand, Irkut and West Caucasian Bat Virus, make it necessary to use generic methods able to detect all lyssaviruses. Primer sequences were chosen from conserved regions in all genotypes in order to optimise a generic RT-PCR. Serial dilutions of 12 RNA extracts from all seven Lyssavirus genotypes were examined to compare the sensitivity of the RT-PCR standardised in this study with a published RT-PCR optimised for EBLV1 detection and capable of amplifying RNA from all seven lyssaviruses. All seven genotypes were detected by both RT-PCRs, however, the sensitivity was higher with the new version of the test. Twenty samples submitted for rabies diagnosis were tested by the new RT-PCR. Eight out of 20 samples from six dogs, one horse and one bat were found positive, in agreement with immunofluorescence results. Seven samples from terrestrial mammals were genotype 1 and one from a bat was genotype 5. In conclusion, this method can be used to complement immunofluorescence for the diagnosis of rabies, enabling the detection of unexpected lyssaviruses during rabies surveillance.

  4. Genotypic Diversity of Phytophthora cinnamomi and P. plurivora in Maryland's Nurseries and Mid-Atlantic Forests.

    Science.gov (United States)

    Beaulieu, Justine; Ford, Blaine; Balci, Yilmaz

    2017-06-01

    Genetic diversity of two Phytophthora spp.-P. cinnamomi (102 isolates), commonly encountered in Maryland nurseries and forests in the Mid-Atlantic United States, and P. plurivora (186 isolates), a species common in nurseries-was characterized using amplified fragment length polymorphism. Expected heterozygosity and other indices suggested a lower level of diversity among P. cinnamomi than P. plurivora isolates. Hierarchical clustering showed P. cinnamomi isolates separated into four clusters, and two of the largest clusters were closely related, containing 80% of the isolates. In contrast, P. plurivora isolates separated into six clusters, one of which included approximately 40% of the isolates. P. plurivora isolates recovered from the environment (e.g., soil and water) were genotypically more diverse than those found causing lesions. For both species, isolate origin (forest versus nursery or among nurseries) was a significant factor of heterozygosity. Clonal groups existed within P. cinnamomi and P. plurivora and included isolates from both forest and nurseries, suggesting that a pathway from nurseries to forests or vice versa exists.

  5. 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.

  6. DQ High genotypic diversity among methicillin-resistant Staphylococcus pseudintermedius isolated from canine infections in Denmark

    DEFF Research Database (Denmark)

    Damborg, Peter; Moodley, Arshnee; Aalbaek, Bent

    2016-01-01

    genotypic diversity and antimicrobial resistance of clinical MRSP isolates obtained from dogs, including dogs sampled on multiple occasions, in Denmark over a six-year period. For that purpose a total of 46 clinical MRSP isolates obtained from 36 dogs between 2009 and 2014 were subjected to antimicrobial...... susceptibility testing, multilocus-sequence typing (MLST) and SCCmec typing. Results: Twenty-three sequence types were identified with ST71, mostly associated with SCCmec II-III, as the most common occurring in 13 dogs. Among the remaining 33 isolates, 19 belonged to clonal complex (CC) 258 comprising ST258...... resistant and almost all CC258 isolates being susceptible. Sixteen of the 19 CC258 isolates had oxacillin MICs of 0.5 g/L, whereas MICs for CC71 isolates were consistently above 4 g/L. Four of five dogs representing multiple isolates had distinct STs on different sampling events. Conclusions: The overall...

  7. The genotypic diversity and lipase production of some thermophilic bacilli from different genera

    Directory of Open Access Journals (Sweden)

    Melih Koc

    2015-12-01

    Full Text Available Abstract Thermophilic 32 isolates and 20 reference bacilli were subjected to Rep-PCR and ITS-PCR fingerprinting for determination of their genotypic diversity, before screening lipase activities. By these methods, all the isolates and references could easily be differentiated up to subspecies level from each other. In screening assay, 11 isolates and 7 references were found to be lipase producing. Their extracellular lipase activities were measured quantitatively by incubating in both tributyrin and olive oil broths at 60 °C and pH 7.0. During the 24, 48 and 72-h period of incubation, the changes in the lipase activities, culture absorbance, wet weight of biomass and pH were all measured. The activity was determined by using pNPB in 50 mM phosphate buffer at pH 7.0 at 60 °C. The lipase production of the isolates in olive oil broths varied between 0.008 and 0.052, whereas these values were found to be 0.002-0.019 (U/mL in the case of tyributyrin. For comparison, an index was established by dividing the lipase activities to cell biomass (U/mg. The maximum thermostable lipase production was achieved by the isolates F84a, F84b, and G. thermodenitrificans DSM 465T (0.009, 0.008 and 0.008 U/mg within olive oil broth, whereas G. stearothermophilus A113 displayed the highest lipase activity than its type strain in tyributyrin. Therefore, as some of these isolates displayed higher activities in comparison to references, new lipase producing bacilli were determined by presenting their genotypic diversity with DNA fingerprinting techniques.

  8. Detection and genotyping of human papilloma virus in cervical cancer specimens from Saudi patients.

    Science.gov (United States)

    Al-Badawi, Ismail A; Al-Suwaine, Abdulrahman; Al-Aker, Murad; Asaad, Lina; Alaidan, Alwaleed; Tulbah, Asma; Fe Bohol, Marie; Munkarah, Adnan R

    2011-07-01

    To determine the rates and types of human papilloma virus (HPV) infection in cervical cancer specimens from Saudi patients. One hundred specimens were randomly selected and retrieved from the achieved samples stored in the pathology department accessioned under the diagnosis of cervical cancer and carcinoma in situ between the years 1997 and 2007. Human papilloma virus in the clinical samples was detected using polymerase chain reaction amplification methods. Two primer systems are commonly used: the MY09-MY11 primers and the GP5+-GP6+ that amplify a wide range of HPV genotypes. Human papilloma virus isolates were genotyped using DNA sequencing and reverse line blot hybridization assay to identify the high-risk HPV genotypes. Ninety cases fulfilled the diagnostic criteria and were analyzed. The rate of HPV genotype detection among cervical cancer samples was 95.5%. The most common HPV genotype detected by both methods was HPV-16 (63.4%), followed by HPV-18 (11.1%), HPV-45 (4.5%), HPV-33 (3.3%), and HPV-31, HPV-52, HPV-53, HPV-58, HPV-59, and HPV-66 with 2.2% prevalence rate each. Prevalence of HPV genotypes among patients with cervical cancer in Saudi Arabia is comparable to the international rates. The use of the reverse line blot hybridization assay genotyping method could be useful for classifying oncogenic HPV-positive women. It is relatively inexpensive and reliable and can be performed in routine practice or epidemiological study compared with the available standard commercial kits.

  9. Genotypic diversity of multidrug-, quinolone- and extensively drug-resistant Mycobacterium tuberculosis isolates in Thailand.

    Science.gov (United States)

    Disratthakit, Areeya; Meada, Shinji; Prammananan, Therdsak; Thaipisuttikul, Iyarit; Doi, Norio; Chaiprasert, Angkana

    2015-06-01

    Drug-resistant tuberculosis (TB), which includes multidrug-resistant (MDR-TB), quinolone-resistant (QR-TB) and extensively drug-resistant tuberculosis (XDR-TB), is a serious threat to TB control. We aimed to characterize the genotypic diversity of drug-resistant TB clinical isolates collected in Thailand to establish whether the emergence of drug-resistant TB is attributable to transmitted resistance or acquired resistance. We constructed the first molecular phylogeny of MDR-TB (n=95), QR-TB (n=69) and XDR-TB (n=28) in Thailand based on spoligotyping and proposed 24-locus multilocus variable-number of tandem repeat analysis (MLVA). Clustering analysis was performed using the unweighted pair group method with arithmetic mean. Spoligotyping identified the Beijing strain (SIT1) as the most predominant genotype (n=139; 72.4%). The discriminatory power of 0.9235 Hunter-Gaston Discriminatory Index (HGDI) with the 15-locus variable-number tandem repeats of mycobacterial interspersed repetitive units typing was improved to a 0.9574 HGDI with proposed 24-locus MLVA, thereby resulting in the subdivision of a large cluster of Beijing strains (SIT1) into 17 subclusters. We identified the spread of drug-resistant TB clones caused by three different MLVA types in the Beijing strain (SIT1) and a specific clone of XDR-TB caused by a rare genotype, the Manu-ancestor strain (SIT523). Overall, 49.5% of all isolates were clustered. These findings suggest that a remarkable transmission of drug-resistant TB occurred in Thailand. The remaining 50% of drug-resistant TB isolates were unique genotypes, which may have arisen from the individual acquisition of drug resistance. Our results suggest that transmitted and acquired resistance have played an equal role in the emergence of drug-resistant TB. Further characterization of whole genome sequences of clonal strains could help to elucidate the mycobacterial genetic factors relevant for drug resistance, transmissibility and virulence

  10. Detection and genotyping of group A rotaviruses isolated from sewage samples in Monastir, Tunisia between April 2007 and April 2010.

    Science.gov (United States)

    Hassine-Zaafrane, M; Kaplon, J; Ben Salem, I; Sdiri-Loulizi, K; Sakly, N; Pothier, P; Aouni, M; Ambert-Balay, K

    2015-11-01

    To ascertain the viral load, the distribution of G and P types of group A rotaviruses (RV-A) in sewage samples and to compare strains in clinical, animal and environmental samples. During our study from April 2007 to April 2010, 518 samples of raw and treated sewage were collected from two biological sewage treatment plants (STPs) located in the Monastir region, Tunisia. RV-A was detected by real-time RT-PCR in 375 (72·4%) sewage samples. According to the quantification results of RV-A, it appears that the viral load in raw and treated sewage of the two STPs was quite similar (P = 0·735). The genotyping of RV-A strains detected in sewage samples showed a great diversity with 10 G types and 8 P types. Most of them were described as common in humans, but we also detected genotypes commonly found in animals. All the genotypes detected in two previous studies performed in our laboratory on clinical and bovine samples were also found in environmental samples. However, some genotypes commonly found in animal were only found in sewage samples. The comparison of environmental, clinical and animal data suggests that STPs may convey not only human sewage but also animal wastes, both of them contaminated with numerous RV-A strains which are not efficiently eliminated by the sewage treatment process and may spread to surface waters. This work demonstrates the potential release of human and animal RV-A into water sources, representing a public health risk, by inducing gastroenteritis in population, but also by increasing the risk of zoonotic transmission and formation of reassortant viruses which could get a higher infectious potential. Our findings also suggest that monitoring of sewage may provide an additional tool to determine the epidemiology of RV-A circulating in a given community. © 2015 The Society for Applied Microbiology.

  11. Detection of HCV genotypes using molecular and radio-isotopic methods

    International Nuclear Information System (INIS)

    Ahmad, N.; Baig, S.M.; Shah, W.A.; Khattak, K.F.; Khan, B.; Qureshi, J.A.

    2004-01-01

    Hepatitis C virus (HCV) accounts for most cases of acute and chronic non-A and non-B liver diseases. Persistent HCV infection may lead to liver cirrhosis and hepatocellular carcinoma. Six major HCV genotypes have been recognized. Infection with different genotypes results in different clinical pictures and responses to antiviral therapy. In the area of Faisalabad (Punjab province of Pakistan), the prevalence and molecular epidemiology of Hepatitis C virus infection had never been investigated before. In this study, we have made an attempt to determine the prevalence, distribution and clinical significance of HCV infection in 1100 suspected patients of liver disease by nested reverse transcriptase polymerase chain reaction (RTPCR) over a period of four years. HCV genotypes of isolates were determined by dot-blot hybridization with genotype specific radiolabeled probes in 337 subjects. The proportion of patients with HCV genotypes 1,2,3 and 4 were 37.38%, 1.86%, 16.16% and 0.29% respectively. Mixed infection of HCV genotype was detected in 120 (35.6%) patients, whereas 31 (9.1%) samples remained unclassified. This study revealed changing epidemiology of hepatitis C virus genotype 1 and 3 in the patients. Multiple infection of HCV genotype in the same patient may be of great clinical and pathological importance and interest. (author)

  12. 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

  13. Invasion Dynamics and Genotypic Diversity of Cogongrass (Imperata cylindrica) at the Point of Introduction in the Southeastern United States

    Science.gov (United States)

    Ludovic J. A. Capo-chichi; Wilson H. Faircloth; A. G. Williamson; Michael G. Patterson; James H. Miller; Edzard van Santen

    2008-01-01

    Nine sites of cogongrass were included in a study of genotypic dimity and spread dynamics at the point of introduction and its adjacent areas in the southern United States. Clones evaluated with two primer pairs yielded a total of 137 amplified fragment length polymorphism (AFLP) hi of which 102 (74.4%) were polymorphic. Genetic diversity was measured as the percentage...

  14. 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.

  15. 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

  16. Single Nucleotide Polymorphisms in Common Bean: Their Discovery and Genotyping Using a Multiplex Detection System

    Directory of Open Access Journals (Sweden)

    E. Gaitán-Solís

    2008-11-01

    Full Text Available Single nucleotide polymorphism (SNP markers are by far the most common form of DNA polymorphism in a genome. The objectives of this study were to discover SNPs in common bean ( L. by comparing sequences from coding and noncoding regions obtained from the GenBank and genomic DNA and to compare sequencing results with those obtained using single base extension (SBE assays on the Luminex-100 system for use in high-throughput germplasm evaluation. We assessed the frequency of SNPs in 47 fragments of common bean DNA, using SBE as the evaluation methodology. We conducted a sequence analysis of 10 genotypes of cultivated and wild beans belonging to the Mesoamerican and Andean genetic pools of . For the 10 genotypes evaluated, a total of 20,964 bp of sequence were analyzed in each genotype and compared, resulting in the discovery of 239 SNPs and 133 InDels, giving an average SNP frequency of one per 88 bp and an InDel frequency of one per 157 bp. This is the equivalent of a nucleotide diversity (θ of 6.27 × 10. Comparisons with the SNP genotypes previously obtained by direct sequencing showed that the SBE assays on the Luminex-100 were accurate, with 2.5% being miscalled and 1% showing no signal. These results indicate that the Luminex-100 provides a high-throughput system that can be used to analyze SNPs in large samples of genotypes both for purposes of assessing diversity and also for mapping studies.

  17. Prevent cervical cancer by screening with reliable human papillomavirus detection and genotyping

    International Nuclear Information System (INIS)

    Ge, Shichao; Gong, Bo; Cai, Xushan; Yang, Xiaoer; Gan, Xiaowei; Tong, Xinghai; Li, Haichuan; Zhu, Meijuan; Yang, Fengyun; Zhou, Hongrong; Hong, Guofan

    2012-01-01

    The incidence of cervical cancer is expected to rise sharply in China. A reliable routine human papillomavirus (HPV) detection and genotyping test to be supplemented by the limited Papanicolaou cytology facilities is urgently needed to help identify the patients with cervical precancer for preventive interventions. To this end, we evaluated a nested polymerase chain reaction (PCR) protocol for detection of HPV L1 gene DNA in cervicovaginal cells. The PCR amplicons were genotyped by direct DNA sequencing. In parallel, split samples were subjected to a Digene HC2 HPV test which has been widely used for “cervical cancer risk” screen. Of the 1826 specimens, 1655 contained sufficient materials for analysis and 657 were truly negative. PCR/DNA sequencing showed 674 infected by a single high-risk HPV, 188 by a single low-risk HPV, and 136 by multiple HPV genotypes with up to five HPV genotypes in one specimen. In comparison, the HC2 test classified 713 specimens as infected by high-risk HPV, and 942 as negative for HPV infections. The high-risk HC2 test correctly detected 388 (57.6%) of the 674 high-risk HPV isolates in clinical specimens, mislabeled 88 (46.8%) of the 188 low-risk HPV isolates as high-risk genotypes, and classified 180 (27.4%) of the 657 “true-negative” samples as being infected by high-risk HPV. It was found to cross-react with 20 low-risk HPV genotypes. We conclude that nested PCR detection of HPV followed by short target DNA sequencing can be used for screening and genotyping to formulate a paradigm in clinical management of HPV-related disorders in a rapidly developing economy

  18. Multilocus genotypic data reveal high genetic diversity and low population genetic structure of Iranian indigenous sheep

    International Nuclear Information System (INIS)

    Vahidi, S.M.F.; Faruque, M.O.; Falahati Anbaran, M.; Afraz, F.; Mousavi, S.M.; Boettcher, P.; Joost, S.; Han, J.L.; Colli, L.; Periasamy, K.; Negrini, R.; Ajmone-Marsan, P.

    2016-01-01

    Full text: Iranian livestock diversity is still largely unexplored, in spite of the interest in the populations historically reared in this country located near the Fertile Crescent, a major livestock domestication centre. In this investigation, the genetic diversity and differentiation of 10 Iranian indigenous fat-tailed sheep breeds were investigated using 18 microsatellite markers. Iranian breeds were found to host a high level of diversity. This conclusion is substantiated by the large number of alleles observed across loci (average 13.83, range 7–22) and by the high within-breed expected heterozygosity (average 0.75, range 0.72–0.76). Iranian sheep have a low level of genetic differentiation, as indicated by the analysis of molecular variance, which allocated a very small proportion (1.67%) of total variation to the between-population component, and by the small fixation index (FST = 0.02). Both Bayesian clustering and principal coordinates analysis revealed the absence of a detectable genetic structure. Also, no isolation by distance was observed through comparison of genetic and geographical distances. In spite of high within-breed variation, signatures of inbreeding were detected by the FIS indices, which were positive in all and statistically significant in three breeds. Possible factors explaining the patterns observed, such as considerable gene flow and inbreeding probably due to anthropogenic activities in the light of population management and conservation programmes are discussed. (author)

  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. SV2: accurate structural variation genotyping and de novo mutation detection from whole genomes.

    Science.gov (United States)

    Antaki, Danny; Brandler, William M; Sebat, Jonathan

    2018-05-15

    Structural variation (SV) detection from short-read whole genome sequencing is error prone, presenting significant challenges for population or family-based studies of disease. Here, we describe SV2, a machine-learning algorithm for genotyping deletions and duplications from paired-end sequencing data. SV2 can rapidly integrate variant calls from multiple structural variant discovery algorithms into a unified call set with high genotyping accuracy and capability to detect de novo mutations. SV2 is freely available on GitHub (https://github.com/dantaki/SV2). jsebat@ucsd.edu. Supplementary data are available at Bioinformatics online.

  1. UPD detection using homozygosity profiling with a SNP genotyping microarray.

    Science.gov (United States)

    Papenhausen, Peter; Schwartz, Stuart; Risheg, Hiba; Keitges, Elisabeth; Gadi, Inder; Burnside, Rachel D; Jaswaney, Vikram; Pappas, John; Pasion, Romela; Friedman, Kenneth; Tepperberg, James

    2011-04-01

    Single nucleotide polymorphism (SNP) based chromosome microarrays provide both a high-density whole genome analysis of copy number and genotype. In the past 21 months we have analyzed over 13,000 samples primarily referred for developmental delay using the Affymetrix SNP/CN 6.0 version array platform. In addition to copy number, we have focused on the relative distribution of allele homozygosity (HZ) throughout the genome to confirm a strong association of uniparental disomy (UPD) with regions of isoallelism found in most confirmed cases of UPD. We sought to determine whether a long contiguous stretch of HZ (LCSH) greater than a threshold value found only in a single chromosome would correlate with UPD of that chromosome. Nine confirmed UPD cases were retrospectively analyzed with the array in the study, each showing the anticipated LCSH with the smallest 13.5 Mb in length. This length is well above the average longest run of HZ in a set of control patients and was then set as the prospective threshold for reporting possible UPD correlation. Ninety-two cases qualified at that threshold, 46 of those had molecular UPD testing and 29 were positive. Including retrospective cases, 16 showed complete HZ across the chromosome, consistent with total isoUPD. The average size LCSH in the 19 cases that were not completely HZ was 46.3 Mb with a range of 13.5-127.8 Mb. Three patients showed only segmental UPD. Both the size and location of the LCSH are relevant to correlation with UPD. Further studies will continue to delineate an optimal threshold for LCSH/UPD correlation. Copyright © 2011 Wiley-Liss, Inc.

  2. Diversity of dengue virus-3 genotype III in Jeddah, Saudi Arabia.

    Science.gov (United States)

    Hashem, Anwar M; Sohrab, Sayed S; El-Kafrawy, Sherif A; Abd-Alla, Adly M M; El-Ela, Saeid Abo; Abujamel, Turki S; Hassan, Ahmed M; Farraj, Suha A; Othman, Noura A; Charrel, Remi N; Azhar, Esam I

    2018-07-01

    Dengue is the most important arboviral disease in tropical and subtropical countries. Dispersal of the vector and an increase in migratory flow between countries have led to large epidemics and severe clinical outcomes. Over the past 20 years, dengue epidemics have become more wide-spread and frequent. Previous studies have shown that dengue is endemic in Jeddah, Makkah and Al-Madinah in western Saudi Arabia as well as in Jazan region in the southern part of the country. The four serotypes of dengue virus (DENV) have been reported from western Saudi Arabia. It has been suggested that pilgrims could play a significant and unique role in DENV-1 and DENV-2 introduction into Saudi Arabia, especially in the cities of Jeddah, Makkah and Al-Madinah during Hajj and Umrah seasons. However, only limited data on DENV-3 in Saudi Arabia are available. All available DENV-3 sequences published and unpublished from Saudi Arabia and other countries were retrieved from Genbank and gene sequence repository and phylogenetically analyzed to examine the diversity of DENV-3 into the city of Jeddah. Based on the analysis of the envelope gene and non-structural 1 (E/NS1) junction sequences, we show that there were at least four independent introductions of DENV-3, all from genotype III into Jeddah. The first introduction was most probably before 1997 as Saudi virus isolates from 1997 formed a cluster without any close relationship to other globally circulating isolates, suggesting their local circulation from previous introduction events. Two introductions were most probably in 2004 with isolates closely-related to isolates from Africa and India (Asia), in addition to another introduction in 2014 with isolates clustering with those from Singapore (Asia). Our data shows that only genotype III isolates of DENV-3 are circulating in Jeddah and highlights the potential role of pilgrims in DENV-3 importation into western Saudi Arabia and subsequent exportation to their home countries during Hajj

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

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    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.

  4. 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). © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  5. 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.

  6. A Multipoint Method for Detecting Genotyping Errors and Mutations in Sibling-Pair Linkage Data

    OpenAIRE

    Douglas, Julie A.; Boehnke, Michael; Lange, Kenneth

    2000-01-01

    The identification of genes contributing to complex diseases and quantitative traits requires genetic data of high fidelity, because undetected errors and mutations can profoundly affect linkage information. The recent emphasis on the use of the sibling-pair design eliminates or decreases the likelihood of detection of genotyping errors and marker mutations through apparent Mendelian incompatibilities or close double recombinants. In this article, we describe a hidden Markov method for detect...

  7. The Chado Natural Diversity module: a new generic database schema for large-scale phenotyping and genotyping data.

    Science.gov (United States)

    Jung, Sook; Menda, Naama; Redmond, Seth; Buels, Robert M; Friesen, Maren; Bendana, Yuri; Sanderson, Lacey-Anne; Lapp, Hilmar; Lee, Taein; MacCallum, Bob; Bett, Kirstin E; Cain, Scott; Clements, Dave; Mueller, Lukas A; Main, Dorrie

    2011-01-01

    Linking phenotypic with genotypic diversity has become a major requirement for basic and applied genome-centric biological research. To meet this need, a comprehensive database backend for efficiently storing, querying and analyzing large experimental data sets is necessary. Chado, a generic, modular, community-based database schema is widely used in the biological community to store information associated with genome sequence data. To meet the need to also accommodate large-scale phenotyping and genotyping projects, a new Chado module called Natural Diversity has been developed. The module strictly adheres to the Chado remit of being generic and ontology driven. The flexibility of the new module is demonstrated in its capacity to store any type of experiment that either uses or generates specimens or stock organisms. Experiments may be grouped or structured hierarchically, whereas any kind of biological entity can be stored as the observed unit, from a specimen to be used in genotyping or phenotyping experiments, to a group of species collected in the field that will undergo further lab analysis. We describe details of the Natural Diversity module, including the design approach, the relational schema and use cases implemented in several databases.

  8. 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.

  9. Human papillomavirus detection with genotyping by the cobas and Aptima assays: Significant differences in HPV 16 detection?

    Science.gov (United States)

    Chorny, Joseph A; Frye, Teresa C; Fisher, Beth L; Remmers, Carol L

    2018-03-23

    The primary high-risk human papillomavirus (hrHPV) assays in the United States are the cobas (Roche) and the Aptima (Hologic). The cobas assay detects hrHPV by DNA analysis while the Aptima detects messenger RNA (mRNA) oncogenic transcripts. As the Aptima assay identifies oncogenic expression, it should have a lower rate of hrHPV and genotype detection. The Kaiser Permanente Regional Reference Laboratory in Denver, Colorado changed its hrHPV assay from the cobas to the Aptima assay. The rates of hrHPV detection and genotyping were compared over successive six-month periods. The overall hrHPV detection rates by the two platforms were similar (9.5% versus 9.1%) and not statistically different. For genotyping, the HPV 16 rate by the cobas was 1.6% and by the Aptima it was 1.1%. These differences were statistically different with the Aptima detecting nearly one-third less HPV 16 infections. With the HPV 18 and HPV 18/45, there was a slightly higher detection rate of HPV 18/45 by the Aptima platform (0.5% versus 0.9%) and this was statistically significant. While HPV 16 represents a low percentage of hrHPV infections, it was detected significantly less by the Aptima assay compared to the cobas assay. This has been previously reported, although not highlighted. Given the test methodologies, one would expect the Aptima to detect less HPV 16. This difference appears to be mainly due to a significantly increased number of non-oncogenic HPV 16 infections detected by the cobas test as there were no differences in HPV 16 detection rates in the high-grade squamous intraepithelial lesions indicating that the two tests have similar sensitivities for oncogenic HPV 16. © 2018 Wiley Periodicals, Inc.

  10. Genotype diversity of Trypanosoma cruzi in small rodents and Triatoma sanguisuga from a rural area in New Orleans, Louisiana.

    Science.gov (United States)

    Herrera, Claudia P; Licon, Meredith H; Nation, Catherine S; Jameson, Samuel B; Wesson, Dawn M

    2015-02-24

    Chagas disease is an anthropozoonosis caused by the protozoan parasite Trypanosoma cruzi that represents a major public health problem in Latin America. Although the United States is defined as non-endemic for Chagas disease due to the rarity of human cases, the presence of T. cruzi has now been amply demonstrated as enzootic in different regions of the south of the country from Georgia to California. In southeastern Louisiana, a high T. cruzi infection rate has been demonstrated in Triatoma sanguisuga, the local vector in this area. However, little is known about the role of small mammals in the wild and peridomestic transmission cycles. This study focused on the molecular identification and genotyping of T. cruzi in both small rodents and T. sanguisuga from a rural area of New Orleans, Louisiana. DNA extractions were prepared from rodent heart, liver, spleen and skeletal muscle tissues and from cultures established from vector feces. T. cruzi infection was determined by standard PCR using primers specific for the minicircle variable region of the kinetoplastid DNA (kDNA) and the highly repetitive genomic satellite DNA (satDNA). Genotyping of discrete typing units (DTUs) was performed by amplification of mini-exon and 18S and 24Sα rRNA genes and subsequent sequence analysis. The DTUs TcI, TcIV and, for the first time, TcII, were identified in tissues of mice and rats naturally infected with T. cruzi captured in an area of New Orleans, close to the house where the first human case of Chagas disease was reported in Louisiana. The T. cruzi infection rate in 59 captured rodents was 76%. The frequencies of the detected DTUs in such mammals were TcI 82%, TcII 22% and TcIV 9%; 13% of all infections contained more than one DTU. Our results indicate a probable presence of a considerably greater diversity in T. cruzi DTUs circulating in the southeastern United States than previously reported. Understanding T. cruzi transmission dynamics in sylvatic and peridomestic cycles

  11. Preliminary concepts for detecting national diversion of LWR spent fuel

    International Nuclear Information System (INIS)

    Sonnier, C.S.; Cravens, M.N.

    1978-04-01

    Preliminary concepts for detecting national diversion of LWR spent fuel during storage, handling and transportation are presented. Principal emphasis is placed on means to achieve timely detection by an international authority. This work was sponsored by the Department of Energy/Office of Safeguards and Security (DOE/OSS) as part of the overall Sandia Fixed Facility Physical Protection Program

  12. Genetic diversity analysis of cyanogenic potential (CNp) of root among improved genotypes of cassava using simple sequence repeat markers.

    Science.gov (United States)

    Moyib, O K; Mkumbira, J; Odunola, O A; Dixon, A G

    2012-12-01

    Cyanogenic potential (CNp) of cassava constitutes a serious problem for over 500 million people who rely on the crop as their main source of calories. Genetic diversity is a key to successful crop improvement for breeding new improved variability for target traits. Forty-three improved genotypes of cassava developed by International Institute of Tropical Agriculture (ITA), Ibadan, were characterized for CNp trait using 35 Simple Sequence.Repeat (SSR) markers. Essential colorimetry picric test was used for evaluation of CNp on a color scale of 1 to 14. The CNp scores obtained ranged from 3 to 9, with a mean score of 5.48 (+/- 0.09) based on Statistical Analysis System (SAS) package. TMS M98/ 0068 (4.0 +/- 0.25) was identified as the best genotype with low CNp while TMS M98/0028 (7.75 +/- 0.25) was the worst. The 43 genotypes were assigned into 7 phenotypic groups based on rank-sum analysis in SAS. Dissimilarity analysis representatives for windows generated a phylogenetic tree with 5 clusters which represented hybridizing groups. Each of the clusters (except 4) contained low CNp genotypes that could be used for improving the high CNp genotypes in the same or near cluster. The scatter plot of the genotypes showed that there was little or no demarcation for phenotypic CNp groupings in the molecular groupings. The result of this study demonstrated that SSR markers are powerful tools for the assessment of genetic variability, and proper identification and selection of parents for genetic improvement of low CNp trait among the IITA cassava collection.

  13. 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.

  14. 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.

  15. Genotypic Diversity within a Single Pseudomonas aeruginosa Strain Commonly Shared by Australian Patients with Cystic Fibrosis.

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    Anna Sze Tai

    Full Text Available In cystic fibrosis (CF, Pseudomonas aeruginosa undergoes intra-strain genotypic and phenotypic diversification while establishing and maintaining chronic lung infections. As the clinical significance of these changes is uncertain, we investigated intra-strain diversity in commonly shared strains from CF patients to determine if specific gene mutations were associated with increased antibiotic resistance and worse clinical outcomes. Two-hundred-and-one P. aeruginosa isolates (163 represented a dominant Australian shared strain, AUST-02 from two Queensland CF centres over two distinct time-periods (2001-2002 and 2007-2009 underwent mexZ and lasR sequencing. Broth microdilution antibiotic susceptibility testing in a subset of isolates was also performed. We identified a novel AUST-02 subtype (M3L7 in adults attending a single Queensland CF centre. This M3L7 subtype was multi-drug resistant and had significantly higher antibiotic minimum inhibitory concentrations than other AUST-02 subtypes. Prospective molecular surveillance using polymerase chain reaction assays determined the prevalence of the 'M3L7' subtype at this centre during 2007-2009 (170 patients and 2011 (173 patients. Three-year clinical outcomes of patients harbouring different strains and subtypes were compared. MexZ and LasR sequences from AUST-02 isolates were more likely in 2007-2009 than 2001-2002 to exhibit mutations (mexZ: odds ratio (OR = 3.8; 95% confidence interval (CI: 1.1-13.5 and LasR: OR = 2.5; 95%CI: 1.3-5.0. Surveillance at the adult centre in 2007-2009 identified M3L7 in 28/509 (5.5% P. aeruginosa isolates from 13/170 (7.6% patients. A repeat survey in 2011 identified M3L7 in 21/519 (4.0% P. aeruginosa isolates from 11/173 (6.4% patients. The M3L7 subtype was associated with greater intravenous antibiotic and hospitalisation requirements, and a higher 3-year risk of death/lung transplantation, than other AUST-02 subtypes (adjusted hazard ratio [HR] = 9.4; 95%CI: 2

  16. 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.

  17. Symmetric Pin Diversion Detection using a Partial Defect Detector (PDET)

    International Nuclear Information System (INIS)

    Sitaraman, S.; Ham, Y.S.

    2009-01-01

    Since the signature from the Partial Defect Detector (PDET) is principally dependent on the geometric layout of the guide tube locations, the capability of the technique in detecting symmetric diversion of pins needs to be determined. The Monte Carlo simulation study consisted of cases where pins were removed in a symmetric manner and the resulting signatures were examined. In addition to the normalized gamma-to-neutron ratios, the neutron and gamma signatures normalized to their maximum values, were also examined. Examination of the shape of the three curves as well as of the peak-to-valley differences in excess of the maximum expected in intact assemblies, indicated pin diversion. A set of simulations with various symmetric patterns of diversion were examined. The results from these studies indicated that symmetric diversions as low as twelve percent could be detected by this methodology

  18. Genetic variation and relationships of old maize genotypes (Zea mays l. detected using SDS-page

    Directory of Open Access Journals (Sweden)

    Martin Vivodík

    2016-11-01

    Full Text Available The assessment of genetic diversity among the members of a species is of vital importance for successful breeding and adaptability. In the present study 40 old genotypes of maize from Hungary, Union of Soviet Socialist Republics, Poland, Czechoslovakia, Yugoslavia and Slovak Republic  were evaluated for the total seed storage proteins using sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE through vertical slab unit. The number of total scorable protein bands was twentythree as a result of SDS-PAGE technique but those that were not cosistent in reproducibility and showed occasional variation in sharpness and density were not considered. Out of twentythree polypeptide bands, 6 (31% were commonly present in all accessions and considered as monomorphic, while 17 (65% showed variations and considered as polymorphic. On the basis of banding profiles of proteins of different kDa, gel was divided into zones A, B and C. The major protein bands were lied in zones A and B, while minor bands were present in zones C. In zone A out of 10 protein bands, 1 were monomorphic and 9 were polymorphic. In zone B out of 8 protein bands, 3 was monomorphic and 5 was polymorphic and in zone C out of 5 protein bands, 2 were monomorphic whereas 3 polymorphic. The dendrogram tree demonstrated the relationship among the forty registered old maize genotypes according to the similarity index, using UPGMA cluster analysis. The dendrogram was divided into two main clusters. The first one contained eleven genotypes from maize, while the second cluster contained the twentynine genotypes of maize. Similarly the present study of genetic variability in the seed storage polypeptide determined by SDS-PAGE technique proved that it is fruitful to identify genetic diversity among accessions of maize. 

  19. Phenotypic and Genotypic Detection of Campylobacter jejuni at Local Chicken and Chicken Meat

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    A Rosyidi

    2010-05-01

    Full Text Available The Objective of this study was to identify the existence of Campylobacter jejuni based on phenotypic and genotypic characteristic in local chicken and chicken meats. Samples of local chicken intestine and meat were tested for the bacterial existence. Phenotypic examination was carried out by means of cultivation followed by gram staining and biochemical tests. Genotypic examination was conducted by polymerase chain reaction (PCR using genus specific16S rRNA gene at 816 bp and membrane-associated protein A (mapA gene at 589 bp as Campylobacter jejuni species-specific gene. The result of phenotypic detection revealed the existence of Campylobacter spp as gram negative, curved rod shape, oxidase positive, urease negative and motile. Genotypic examination also indicated the existence of bacteria using both primers. However, no Campylobacter jejuni detected from meat of the chickens. The results suggest that the method of PCR using a primer detecting species-specific gene of Campylobacter jejuni gives a rapid and accurate detection of the bacteria as compared to that using phenotypic and biochemical test. Identification of Campylobacter spp from chicken meats should be improved with enrichment method and sample collection. (Animal Production 12(2: 128-134 (2010Key Words: Campylobacter jejuni, mapA gene, local chicken

  20. Start codon targeted (scot polymorphism reveals genetic diversity in european old maize (zea mays l. Genotypes

    Directory of Open Access Journals (Sweden)

    Martin Vivodík

    2016-11-01

    Full Text Available Maize (Zea mays L. is one of the world's most important crop plants following wheat and rice, which provides staple food to large number of human population in the world. It is cultivated in a wider range of environments than wheat and rice because of its greater adaptability. Molecular characterization is frequently used by maize breeders as an alternative method for selecting more promising genotypes and reducing the cost and time needed to develop hybrid combinations. In the present investigation 40 genotypes of maize from Czechoslovakia, Hungary, Poland, Union of Soviet Socialist Republics, Slovakia and Yugoslavia were analysed using 20 Start codon targeted (SCoT markers. These primers produced total 114 fragments across 40 maize genotypes, of which 86 (76.43% were polymorphic with an average of 4.30 polymorphic fragments per primer and number of amplified fragments ranged from 2 (SCoT 45 to 8 (SCoT 28 and SCoT 63. The polymorphic information content (PIC value ranged from 0.374 (ScoT 45 to 0.846 (SCoT 28 with an average of 0.739. The dendrogram based on hierarchical cluster analysis using UPGMA algorithm was prepared. The hierarchical cluster analysis showed that the maize genotypes were divided into two main clusters. Unique maize genotype (cluster 1, Zuta Brzica, originating from Yugoslavia separated from others. Cluster 2 was divided into two main clusters (2a and 2b. Subcluster 2a contained one Yugoslavian genotype Juhoslavanska and subcluster 2b was divided in two subclusters 2ba and 2bb. The present study shows effectiveness of employing SCoT markers in analysis of maize, and would be useful for further studies in population genetics, conservation genetics and genotypes improvement.

  1. Genetic diversity among 16 genotypes of Coffea arabica in the Brazilian cerrado.

    Science.gov (United States)

    Machado, C M S; Pimentel, N S; Golynsk, A; Ferreira, A; Vieira, H D; Partelli, F L

    2017-09-21

    For the selection of coffee plants that have favorable characteristics, it is necessary to evaluate variables related to production. Knowledge of the genetic divergence of arabica coffee is of extreme importance, as this knowledge can be associated with plant breeding programs in order to combine genetic divergence with good productive performance. The objective of this study was to evaluate the genetic divergence among 16 genotypes of Coffea arabica with the purpose of identifying the most dissimilar genotypes for the establishment of breeding programs and adaptation to the Brazilian cerrado. The genetic divergence was evaluated using multivariate procedures, the analysis of the average grouping unweighted pair group method with arithmetic mean (UPGMA) and main components in 2013 and 2014. Eight characters were evaluated in an experiment conducted in Morrinhos, Goiás. The presence of genetic divergence among the 16 C. arabica genotypes under cerrado conditions was recorded. The formation of UPGMA groups for the evaluated characteristics was pertinent due to the number of genotypes. The first three major components accounted for 81.77% of the total variance. The genotype H-419-3-4-4-13(C-241) of low size was the most divergent, followed by Catucaí 2 SL and Catiguá MG2, according to the main components.

  2. 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

  3. Genetic history of hepatitis C virus in Venezuela: high diversity and long time of evolution of HCV genotype 2.

    Directory of Open Access Journals (Sweden)

    Maria Z Sulbarán

    Full Text Available 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.

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

    Science.gov (United States)

    Raaijmakers, J M; Weller, D M

    2001-06-01

    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 on roots of wheat grown in a soil naturally suppressive to take-all disease of wheat. Among 101 isolates, 16 different groups were identified by random amplified polymorphic DNA (RAPD) analysis. One RAPD group made up 50% of the total population of DAPG-producing Pseudomonas spp. Both short- and long-term studies indicated that this dominant genotype, exemplified by P. fluorescens Q8r1-96, is highly adapted to the wheat rhizosphere. Q8r1-96 requires a much lower dose (only 10 to 100 CFU seed(-1) or soil(-1)) to establish high rhizosphere population densities (10(7) CFU g of root(-1)) than Q2-87 and 1M1-96, two genotypically different, DAPG-producing P. fluorescens strains. Q8r1-96 maintained a rhizosphere population density of approximately 10(5) CFU g of root(-1) after eight successive growth cycles of wheat in three different, raw virgin soils, whereas populations of Q2-87 and 1M1-96 dropped relatively quickly after five cycles and were not detectable after seven cycles. In short-term studies, strains Q8r1-96, Q2-87, and 1M1-96 did not differ in their ability to suppress take-all. After eight successive growth cycles, however, Q8r1-96 still provided control of take-all to the same level as obtained in the take-all suppressive soil, whereas Q2-87 and 1M1-96 gave no control anymore. Biochemical analyses indicated that the superior rhizosphere competence of Q8r1-96 is not related to in situ DAPG production levels. We postulate that certain rhizobacterial genotypes have evolved a preference for colonization of specific crops. By exploiting diversity of antagonistic rhizobacteria that share a common trait, biological control can be improved significantly.

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

    Directory of Open Access Journals (Sweden)

    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.

  6. Evaluating late detection capability against diverse insider adversaries

    International Nuclear Information System (INIS)

    Sicherman, A.

    1987-01-01

    This paper describes a model for evaluating the late (after-the-fact) detection capability of material control and accountability (MCandA) systems against insider theft or diversion of special nuclear material. Potential insider cover-up strategies to defeat activities providing detection (e.g., inventories) are addressed by the model in a tractable manner. For each potential adversary and detection activity, two probabilities are assessed and used to fit the model. The model then computes the probability of detection for activities occurring periodically over time. The model provides insight into MCandA effectiveness and helps identify areas for safeguards improvement. 4 refs., 4 tabs

  7. 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

  8. Study of Genetic Diversity of Some Persian walnut Genotypes in Mashhad Commercial Orchards by using ISSR Marker

    Directory of Open Access Journals (Sweden)

    Shadi Attar

    2018-02-01

    Full Text Available Introduction: Persian walnut (Juglans regia L., belonging to the Juglandaceae family, has its natural origin in the mountainous regions of central Asia and especially northern forests of Iran. Most walnut genotypes are seedling and sexually reproduced. Conducting studies on the genetic structure of these genotypes to identify, select and maintain their genetic resources is important. Identifying and collecting local varieties of fruit trees is considered as the first step on the path of breeding programs and lack of information regarding plants genetic characteristics causes the breeding work to be done slowly. Various methods have been used for studying genetic diversity and determining the genetic relationship between European and Asian varieties of walnut and identifying commercial walnut varieties, among which we can mention: Morphologic indices, Alozyme, Isozym, RFLP, RAPD, AFLP and ISSR markers. ISSR molecular marker was used in order to investigate genetic diversity of some genotypes of Persian walnut (Juglans regia L. in Mashhad orchards. . Materials and methods: To begin with, about 56 walnut trees from 4 orchards in Mashhad (Esteghlal (1, Golestan (2, Alandasht (3 and Emam Reza (4 were selected and tagged from 2014 to 2016. In the spring of 2014 with the beginning of trees growth and opening of leaves, a number of leaves from each genotype were collected. After DNA extraction, the quality of samples by agarose gel (1 percentage and electrophoresis method and quantity of them via spectrophotometer device at 260 and 280 nm wavelengths were determined. First, 24 primers of ISSR marker were prepared and after initial evaluation on 5 random genotypes, 9 primers with high polymorphism and repeatability were selected for further investigation. For PCR reaction, Amplicon kit (code 180 301, made in Denmark was used. Gel electrophoresis images of primers that produced polymorphic bands with suitable resolution were analyzed manually. After

  9. Active neutron technique for detecting attempted special nuclear material diversion

    International Nuclear Information System (INIS)

    Smith, G.W.; Rice, L.G. III.

    1979-01-01

    The identification of special nuclear material (SNM) diversion is necessary if SNM inventory control is to be maintained at nuclear facilities. (Special nuclear materials are defined for this purpose as either 235 U of 239 Pu.) Direct SNM identification by the detection of natural decay or fission radiation is inadequate if the SNM is concealed by appropriate shielding. The active neutron interrogation technique described combines direct SNM identification by delayed fission neutron (DFN) detection with implied SNM detection by the identification of materials capable of shielding SNM from direct detection. This technique is being developed for application in an unattended material/equipment portal through which items such as electronic instruments, packages, tool boxes, etc., will pass. The volume of this portal will be 41-cm wide, 53-cm high and 76-cm deep. The objective of this technique is to identify an attempted diversion of at least 20 grams of SNM with a measurement time of 30 seconds

  10. 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.

  11. 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. PMID:25734518

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

    Directory of Open Access Journals (Sweden)

    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

  13. Genotypic diversity and mixed infection in newborn disease and hearing loss in congenital cytomegalovirus infection.

    Science.gov (United States)

    Pati, Sunil K; Pinninti, Swetha; Novak, Zdenek; Chowdhury, Nazma; Patro, Raj K; Fowler, Karen; Ross, Shannon; Boppana, Suresh

    2013-10-01

    Congenital cytomegalovirus (cCMV) is a common congenital infection and a leading nongenetic cause of sensorineural hearing loss (SNHL). CMV exhibits extensive genetic variability, and infection with multiple CMV strains (mixed infection) was shown to be common in congenital CMV. The role of mixed infections in disease and outcome remains to be defined. Genotyping of envelope glycoproteins, UL55 (gB), UL73 (gN) and UL75 (gH), was performed on saliva specimens of 79 infants from the ongoing CMV and Hearing Multicenter Screening (CHIMES) Study and on blood and urine specimens of 52 infants who participated in natural history studies at the University of Alabama at Birmingham. Genotyping of UL144 and US28 was also performed in the CHIMES cohort. The association of individual genotypes and mixed infection with clinical findings at birth and SNHL was examined. Thirty-seven of 131 infants (28%) were symptomatic at birth and 26 (20%) had SNHL at birth. All known genotypes of UL55, UL75, UL73 and US28 were represented, and no particular genotype was associated with symptomatic infection or SNHL. UL144 subtype C was more common in symptomatic infants but not associated with SNHL. Mixed infection was observed in 59 infants (45%) and not associated with symptoms (P = 0.43) or SNHL at birth (P = 0.82). In the cohort of 52 infants with long-term hearing outcome, mixed infection at birth was not predictive of SNHL. Mixed infection is common in infants with congenital CMV but is neither associated with symptomatic infection nor associated with SNHL.

  14. Genetic diversity of bread wheat genotypes in Iran for some nutritional value and baking quality traits.

    Science.gov (United States)

    Amiri, Reza; Sasani, Shahryar; Jalali-Honarmand, Saeid; Rasaei, Ali; Seifolahpour, Behnaz; Bahraminejad, Sohbat

    2018-02-01

    Genetic variation among 78 irrigated bread wheat genotypes was studied for their nutritional value and baking quality traits as well as some agronomic traits. The experiment was conducted in a randomized complete block design with three replicates under normal and terminal drought stress conditions in Kermanshah, Iran during 2012-2013 cropping season. The results of combined ANOVA indicated highly significant genotypic differences for all traits. All studied traits except grain yield, hectoliter weight and grain fiber content were significantly affected by genotype × environment interaction. Drought stress reduced grain yield, thousand kernel weight, gluten index, grain starch content and hectoliter weight and slightly promoted grain protein and fiber contents, falling number, total gluten and ratio of wet gluten to grain protein content. Grain yield by 31.66% and falling number by 9.20% attained the highest decrease and increase due to drought stress. There were negative and significant correlations among grain yield with grain protein and fiber contents under both conditions. Results of cluster analysis showed that newer genotypes had more grain yield and gluten index than older ones, but instead, they had the lower grain protein and fiber contents. It is thought that wheat breeders have bred cultivars with high grain yield, low protein content, and improved bread-making attributes during last seven decades. While older genotypes indicated significantly higher protein contents, and some of them had higher gluten index. We concluded from this study that it is imperative for breeders to pay more attention to improve qualitative traits coordinated to grain yield.

  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. 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.

  17. 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

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

    Directory of Open Access Journals (Sweden)

    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.

  19. 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 t...

  20. Association of Heavy Rainfall on Genotypic Diversity in V. cholerae Isolates from an Outbreak in India

    Directory of Open Access Journals (Sweden)

    A. K. Goel

    2011-01-01

    Full Text Available The outbreak of waterborne disease cholera has been associated with rainfall and flooding events by contamination of potable water with environmental Vibrio cholerae. The continuation of the epidemic in a region, however, is often due to secondary transmission of the initial outbreak strain through human waste. This paper reports, on the contrary, a rapid shift of genotype from one V. cholerae strain to another one in an epidemic region. V. cholerae isolated from patients during 2005 cholera epidemic in Chennai, India were characterized using PCR identification of toxin genes, antibiogram, and genomic fingerprinting analysis. The results showed that in spite of the similarity of toxin genes and antibiogram, the Vibrio isolates grouped into two different clusters based on the ERIC-PCR fingerprinting. Each cluster corresponded to a distinct peak of cholera outbreak, which occurred after separate heavy rainfall. The results suggest that the rainfall event can bring various genotypes of V. cholerae strains causing multiple outbreaks.

  1. 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

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

    Directory of Open Access Journals (Sweden)

    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

  3. 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

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

    Directory of Open Access Journals (Sweden)

    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.

  5. Construction and applications of DNA probes for detection of polychlorinated biphenyl-degrading genotypes in toxic organic-contaminated soil environments

    International Nuclear Information System (INIS)

    Walia, S.; Khan, A.; Rosenthal, N.

    1990-01-01

    Several DNA probes for polychlorinated biphenyl (PCB)-degrading genotypes were constructed from PCB-degrading bacteria. These laboratory-engineered DNA probes were used for the detection, enumeration, and isolation of specific bacteria degrading PCBs. Dot blot analysis of purified DNA from toxic organic chemical-contaminated soil bacterial communities showed positive DNA-DNA hybridization with a 32P-labeled DNA probe (pAW6194, cbpABCD). Less than 1% of bacterial colonies isolated from garden topsoil and greater than 80% of bacteria isolated from PCB-contaminated soils showed DNA homologies with 32P-labeled DNA probes. Some of the PCB-degrading bacterial isolates detected by the DNA probe method did not show biphenyl clearance. The DNA probe method was found to detect additional organisms with greater genetic potential to degrade PCBs than the biphenyl clearance method did. Results from this study demonstrate the usefulness of DNA probes in detecting specific PCB-degrading bacteria, abundance of PCB-degrading genotypes, and genotypic diversity among PCB-degrading bacteria in toxic chemical-polluted soil environments. We suggest that the DNA probe should be used with caution for accurate assessment of PCB-degradative capacity within soils and further recommend that a combination of DNA probe and biodegradation assay be used to determine the abundance of PCB-degrading bacteria in the soil bacterial community

  6. Detection and genotyping of Entamoeba histolytica, Entamoeba dispar, Giardia lamblia, and Cryptosporidium parvum by oligonucleotide microarray.

    Science.gov (United States)

    Wang, Zheng; Vora, Gary J; Stenger, David A

    2004-07-01

    Entamoeba histolytica, Giardia lamblia, and Cryptosporidium parvum are the most frequently identified protozoan parasites causing waterborne disease outbreaks. The morbidity and mortality associated with these intestinal parasitic infections warrant the development of rapid and accurate detection and genotyping methods to aid public health efforts aimed at preventing and controlling outbreaks. In this study, we describe the development of an oligonucleotide microarray capable of detecting and discriminating between E. histolytica, Entamoeba dispar, G. lamblia assemblages A and B, and C. parvum types 1 and 2 in a single assay. Unique hybridization patterns for each selected protozoan were generated by amplifying six to eight diagnostic sequences/organism by multiplex PCR; fluorescent labeling of the amplicons via primer extension; and subsequent hybridization to a set of genus-, species-, and subtype-specific covalently immobilized oligonucleotide probes. The profile-based specificity of this methodology not only permitted for the unequivocal identification of the six targeted species and subtypes, but also demonstrated its potential in identifying related species such as Cryptosporidium meleagridis and Cryptosporidium muris. In addition, sensitivity assays demonstrated lower detection limits of five trophozoites of G. lamblia. Taken together, the specificity and sensitivity of the microarray-based approach suggest that this methodology may provide a promising tool to detect and genotype protozoa from clinical and environmental samples.

  7. Recurrent high level parvovirus B19/genotype 2 viremia in a renal transplant recipient analyzed by real-time PCR for simultaneous detection of genotypes 1 to 3.

    Science.gov (United States)

    Liefeldt, Lutz; Plentz, Annelie; Klempa, Boris; Kershaw, Olivia; Endres, Anne-Sophie; Raab, Ulla; Neumayer, Hans-H; Meisel, Helga; Modrow, Susanne

    2005-01-01

    Organ transplant recipients infected with parvovirus B19 frequently develop persistent viremia associated with chronic anemia and pure red cell aplasia. In this study, a male renal transplant recipient who had been infected with parvovirus B19/genotype 2 after renal transplantation at the age of 34 years is described. The patient was repeatedly treated with high dose intravenous immunoglobulin (IVIG) that resulted in the resolvement of symptoms but not in virus eradication. During an observation period of 33 months after transplantation three phases associated with high parvovirus B19 viremia were observed. Both the first and the second viremic phases were combined with severe anemia. Parvovirus B19 specific IgM-antibodies were initially detected at the beginning of the second phase in continually rising concentrations. Initially eradication of the virus by immunoglobulin therapy was reported after the first viremic phase [Liefeldt et al. (2002): Nephrol Dial Transplant 17:1840-1842]. Retrospectively this statement has to be corrected. It was based on the use of a qualitative PCR assay specific for parvovirus B19 genotype 1 associated with reduced sensitivity for detection of genotype 2. After sequence analysis of the viral DNA and adjustment of a real-time PCR assay (TaqMan) for quantitative detection of all three B19 virus genotypes analysis of consecutive serum samples allowed the demonstration of long lasting phases with reduced viral loads following IVIG-treatment. These results demonstrate that IVIG treatment of parvovirus B19-triggered anemia in transplant recipients offers an opportunity to resolve symptoms, but does not guarantee eradication of the virus. Since reactivation of parvovirus B19 infection can result in high virus load associated with the recurrence of symptoms repeated screening for viral DNA is recommended using the TaqMan system established for quantitative detection of all three genotypes of parvovirus B19. Copyright 2005 Wiley-Liss, Inc.

  8. Evaluation of genetic diversity amongst Descurainia sophia L. genotypes by inter-simple sequence repeat (ISSR) marker.

    Science.gov (United States)

    Saki, Sahar; Bagheri, Hedayat; Deljou, Ali; Zeinalabedini, Mehrshad

    2016-01-01

    Descurainia sophia is a valuable medicinal plant in family of Brassicaceae. To determine the range of diversity amongst D. sophia in Iran, 32 naturally distributed plants belonging to six natural populations of the Iranian plateau were investigated by inter-simple sequence repeat (ISSR) markers. The average percentage of polymorphism produced by 12 ISSR primers was 86 %. The PIC values for primers ranged from 0.22 to 0.40 and Rp values ranged between 6.5 and 19.9. The relative genetic diversity of the populations was not high (Gst =0.32). However, the value of gene flow revealed by the ISSR marker was high (Nm = 1.03). UPGMA clustering method based on Jaccard similarity coefficient grouped the genotypes into two major clusters. Graph results from Neighbor-Net Network generated after a 1000 bootstrap test using Jaccard coefficient, and STRUCTURE analysis confirmed the UPGMA clustering. The first three PCAs represented 57.31 % of the total variation. The high levels of genetic diversity were observed within populations, which is useful in breeding and conservation programs. ISSR is found to be an eligible marker to study genetic diversity of D. sophia.

  9. 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.

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

    Science.gov (United States)

    Sheleby-Elías, Jessica; Solórzano-Morales, Antony; Romero-Zuñiga, Juan José; Dolz, Gaby

    2013-01-01

    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.

  11. Comparative ultrastructure of fruit plastids in three genetically diverse genotypes of apple (Malus × domestica Borkh.) during development

    Science.gov (United States)

    Schaeffer, Scott M.; Christian, Ryan; Castro-Velasquez, Nohely; Hyden, Brennan; Lynch-Holm, Valerie

    2017-01-01

    Plastids are the defining organelle for a plant cell and are critical for myriad metabolic functions. The role of leaf plastid, chloroplast, is extensively documented; however, fruit plastids—chromoplasts—are poorly understood, especially in the context of the diverse metabolic processes operating in these diverse plant organs. Recently, in a comparative study of the predicted plastid-targeted proteomes across seven plant species, we reported that each plant species is predicted to harbor a unique set of plastid-targeted proteins. However, the temporal and developmental context of these processes remains unknown. In this study, an ultrastructural analysis approach was used to characterize fruit plastids in the epidermal and collenchymal cell layers at 11 developmental timepoints in three genotypes of apple (Malus × domestica Borkh.): chlorophyll-predominant ‘Granny Smith’, carotenoid-predominant ‘Golden Delicious’, and anthocyanin-predominant ‘Top Red Delicious’. Plastids transitioned from a proplastid-like plastid to a chromoplast-like plastid in epidermis cells, while in the collenchyma cells, they transitioned from a chloroplast-like plastid to a chloro-chromo-amyloplast plastid. Plastids in the collenchyma cells of the three genotypes demonstrated a diverse array of structures and features. This study enabled the identification of discrete developmental stages during which specific functions are most likely being performed by the plastids as indicated by accumulation of plastoglobuli, starch granules, and other sub-organeller structures. Information regarding the metabolically active developmental stages is expected to facilitate biologically relevant omics studies to unravel the complex biochemistry of plastids in perennial non-model systems. PMID:28698906

  12. 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

    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......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...

  13. Genetic diversity of wild and cultivated genotypes of pigeonpea through RAPD and SSR markers.

    Science.gov (United States)

    Walunjkar, Babasaheb C; Parihar, Akarsh; Singh, Nirbhay Kumar; Parmar, L D

    2015-03-01

    Eight wild and four cultivated pigeonpea genotypes were subjected to RAPD and microsatellite analysis, with 40 primers each. Out of these, eight RAPD and five SSR primers were found polymorphic. RAPD primers showed 100% polymorphism and produced a total of 517 DNA fragments, whereas SSR primers produced 67 fragments and they too showed 100% polymorphism. The RAPD markers revealed highest similarity co-efficient of 0.93 (GT-100 and ICPL-87), whereas the highest similarity co-efficient obtained with SSR markers was 1.00 (GTH-1 and GT-100). Average PIC value obtained with RAPD and SSR were 0.90 and 0.18, respectively. The arithmetic mean heterozygosity and marker index were 0.90 and 22.47 respectively with RAPD marker, whereas the corresponding values for SSR markers were 0.18 and 33.66. Moreover; the four wild genotypes (Cajanus scarabaeoides, Rhyncosia rufescence, Cajanus cajanifolius and Rhyncosia canna) and the four cultivars (GTH-1, GT-100, ICPL-87 and GT-1) grouped distinctly in the same subgroups of the dendrograms obtained with both RAPD and SSR analysis. Therefore, the findings of SSR supplement and validate the results obtained with RAPD analysis.

  14. Evaluating late detection capability against diverse insider adversaries

    International Nuclear Information System (INIS)

    Sicherman, A.

    1987-01-01

    The threat of theft or diversion of special nuclear material (SNM) by insiders is a key concern for safeguards planners. Different types of employees having varying degrees of access to both SNM and safeguards systems pose a difficult challenge for theft detection. Safeguards planners rely on physical security, material control, and accountability to provide detection of a theft attempt. When detection occurs too late to prevent a theft, it is called a late detection or late alarm. Activities or events that many provide late detection usually belong to material control and accountability (MC ampersand A) activities. A model has been developed for evaluating the probability of late detection as a function of time elapsed since the theft. Late detection capability is beneficial if it is timely enough to improve the ability to determine the cause of an alarm, speed recovery of SNM, prevent an incorrect response to a threat demand, or promote assurance that no theft has occurred in the absence of an alarm. The model provides insight into the effectiveness of late detection safeguards components in place and helps to identify areas where the MC ampersand A can be most effectively improved

  15. First Insight into a Nationwide Genotypic Diversity of Mycobacterium tuberculosis among Previously Treated Pulmonary Tuberculosis Cases in Benin, West Africa.

    Science.gov (United States)

    Affolabi, Dissou; Sanoussi, N'Dira; Codo, Sergio; Sogbo, Fréderic; Wachinou, Prudence; Massou, Faridath; Kehinde, Aderemi; Anagonou, Séverin

    2017-01-01

    Molecular studies on tuberculosis (TB) are rare in low-resource countries like Benin, where data on molecular study on previously treated TB cases is unavailable. From January to December 2014, all smear- and culture-positive previously treated pulmonary TB patients from all TB clinics were systematically recruited. Drug susceptibility testing and spoligotyping were performed on all isolates. Of the 100 patients recruited, 71 (71.0%) were relapse cases and 24 (24.0%) were failure cases, while 5 (5.0%) were default cases. Resistance rate to any first-line drug was 40.0%, while 12.0% of strains were multidrug-resistant (MDR) and no strain was extensively drug-resistant (XDR). A total of 40 distinct spoligotypes were found to be corresponding to a genotypic diversity of 40.0%. ST61 was the most predominant spoligotype with prevalence of 33.0%. In all, 31 single spoligotypes and nine clusters were observed with 2 to 33 strains per cluster giving a clustering rate of 69.0%. Euro-American (Lineage 4) was the most prevalent lineage (74.0%) and Lineage 2 was associated with resistance to streptomycin. This first insight into genetic diversity of previously treated pulmonary TB patients in Benin showed a relatively high genetic diversity of Mycobacterium tuberculosis .

  16. First Insight into a Nationwide Genotypic Diversity of Mycobacterium tuberculosis among Previously Treated Pulmonary Tuberculosis Cases in Benin, West Africa

    Directory of Open Access Journals (Sweden)

    Dissou Affolabi

    2017-01-01

    Full Text Available Background. Molecular studies on tuberculosis (TB are rare in low-resource countries like Benin, where data on molecular study on previously treated TB cases is unavailable. Materials and Methods. From January to December 2014, all smear- and culture-positive previously treated pulmonary TB patients from all TB clinics were systematically recruited. Drug susceptibility testing and spoligotyping were performed on all isolates. Results. Of the 100 patients recruited, 71 (71.0% were relapse cases and 24 (24.0% were failure cases, while 5 (5.0% were default cases. Resistance rate to any first-line drug was 40.0%, while 12.0% of strains were multidrug-resistant (MDR and no strain was extensively drug-resistant (XDR. A total of 40 distinct spoligotypes were found to be corresponding to a genotypic diversity of 40.0%. ST61 was the most predominant spoligotype with prevalence of 33.0%. In all, 31 single spoligotypes and nine clusters were observed with 2 to 33 strains per cluster giving a clustering rate of 69.0%. Euro-American (Lineage 4 was the most prevalent lineage (74.0% and Lineage 2 was associated with resistance to streptomycin. Conclusion. This first insight into genetic diversity of previously treated pulmonary TB patients in Benin showed a relatively high genetic diversity of Mycobacterium tuberculosis.

  17. 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. Copyright © 2016 Elsevier GmbH. All rights reserved.

  18. GENETIC OF DIVERSITY AND RELATIONSHIP BAMBARA GROUNDNUT (Vigna Subteranea L. GENOTYPE LANDRACES OF WEST JAVA

    Directory of Open Access Journals (Sweden)

    Enceng Sobari

    2017-12-01

    Full Text Available Bambara groundnut (Vigna subteranea L. is one of underutliized crops in Indonesia. Bambara groundnut is potential to be developed and can be utilized as an alternative food source in Indonesia. Bambara groundnut greatly varies and has a very wide area of adaptation. The experiment was conducted at the experimental field station at Ciparanje in Padjadjaran University. Starting on September 2014 until March 2015 with Randomized Block Design (RBD and repeated two times. The research used 30 accessions originally from various locations in West Java (Bandung, Tasikmalaya, Garut, Sumedang, Bogor, Majalengka and East Java (Lamongan, Madura. Genetic variability of Bambara groundnut landrace  in some West Java showed broad criteria on the characters fresh pod weight, dry pod weight, weight of 100 seeds, and weight per plot. Genotypes which had many similarities in some characters based on euclidian distance coefficient had close relationship.

  19. Diversity of Physiological Traits In jerusalem Artichoke genotypes under Non-stress and Drought Stress

    International Nuclear Information System (INIS)

    Ruttanaprasert, R.; Banterng, P.; Vorasoot, N.; Kesmala, T.; Patanothai, A.; Jogloy, S.

    2016-01-01

    Physiological traits such as SPAD Chlorophyll Meter Reading (SCMR), specific leaf area (SLA) and harvest index (HI) play an important role in crop yield. The objectives of this work were to study the effect of drought stress on HI, SCMR and SLA and explore genetic variability for these physiological traits in Jerusalem artichoke (JA) (Helianthus tuberosus L.). Field experiments were conducted in the dry period of 2010/11 and 2011/12 in the Northeast of Thailand using a strip plot design with four replications. A horizontal factor was three different water regimes (W1:100 percent Crop water requirement (ETcrop), W2: 75 percent ETcrop and W3: 45 percent ETcrop) and a vertical factor was 40 JA genotypes. Measurements on HI, relative water content (RWC), SLA and SCMR were conducted at 40, 60 and 70 days after transplantation. Drought stress significantly reduced RWC and SLA but significantly increased SCMR. High variations in SCMR (32-59) and SLA (78-213 cm/sup 2/ g/sup -1/) were found among genotypes. The correlations between HI and SCMR (r = 0.56 to 0.78, p<=0.01) were positive and significant, whereas the respective ones between HI and SLA (r = -0.60 to -0.76, p<=0.01) were negative and significant as those between SCMR and SLA (r = -0.73 to -0.90, p<=0.01). These findings suggested that SCMR was linked with SLA and HI in JA. SCMR could be used as a physiological trait for indirect selection for HI and productivity under various water regimes in JA. (author)

  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. 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. Copyright © 2015 Elsevier B.V. All rights reserved.

  2. Assessment of genetic diversity, population structure and relationships in Indian and non-Indian genotypes of finger millet (Eleusine coracana (L.) Gaertn) using genomic SSR markers.

    Science.gov (United States)

    Ramakrishnan, M; Antony Ceasar, S; Duraipandiyan, V; Al-Dhabi, N A; Ignacimuthu, S

    2016-01-01

    We evaluated the genetic variation and population structure in Indian and non-Indian genotypes of finger millet using 87 genomic SSR primers. The 128 finger millet genotypes were collected and genomic DNA was isolated. Eighty-seven genomic SSR primers with 60-70 % GC contents were used for PCR analysis of 128 finger millet genotypes. The PCR products were separated and visualized on a 6 % polyacrylamide gel followed by silver staining. The data were used to estimate major allele frequency using Power Marker v3.0. Dendrograms were constructed based on the Jaccard's similarity coefficient. Statistical fitness and population structure analyses were performed to find the genetic diversity. The mean major allele frequency was 0.92; the means of polymorphic alleles were 2.13 per primer and 1.45 per genotype; the average polymorphism was 59.94 % per primer and average PIC value was 0.44 per primer. Indian genotypes produced an additional 0.21 allele than non-Indian genotypes. Gene diversity was in the range from 0.02 to 0.35. The average heterozygosity was 0.11, close to 100 % homozygosity. The highest inbreeding coefficient was observed with SSR marker UGEP67. The Jaccard's similarity coefficient value ranged from 0.011 to 0.836. The highest similarity value was 0.836 between genotypes DPI009-04 and GPU-45. Indian genotypes were placed in Eleusine coracana major cluster (EcMC) 1 along with 6 non-Indian genotypes. AMOVA showed that molecular variance in genotypes from various geographical regions was 4 %; among populations it was 3 % and within populations it was 93 %. PCA scatter plot analysis showed that GPU-28, GPU-45 and DPI009-04 were closely dispersed in first component axis. In structural analysis, the genotypes were divided into three subpopulations (SP1, SP2 and SP3). All the three subpopulations had an admixture of alleles and no pure line was observed. These analyses confirmed that all the genotypes were genetically diverse and had been grouped based on

  3. 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.

  4. Relationship between ureB Sequence Diversity, Urease Activity and Genotypic Variations of Different Helicobacter pylori Strains in Patients with Gastric Disorders.

    Science.gov (United States)

    Ghalehnoei, Hossein; Ahmadzadeh, Alireza; Farzi, Nastaran; Alebouyeh, Masoud; Aghdaei, Hamid Asadzadeh; Azimzadeh, Pendram; Molaei, Mahsa; Zali, Mohammad Reza

    2016-01-01

    Association of the severity of Helicobacter pylori induced diseases with virulence entity of the colonized strains was proven in some studies. Urease has been demonstrated as a potent virulence factor for H. pylori. The main aim of this study was investigation of the relationships of ureB sequence diversity, urease activity and virulence genotypes of different H. pylori strains with histopathological changes of gastric tissue in infected patients suffering from different gastric disorders. Analysis of the virulence genotypes in the isolated strains indicated significant associations between the presence of severe active gastritis and cagA+ (P = 0.039) or cagA/iceA1 genotypes (P = 0.026), and intestinal metaplasia and vacA m1 (P = 0.008) or vacA s1/m2 (P = 0.001) genotypes. Our results showed a 2.4-fold increased risk of peptic ulcer (95% CI: 0.483-11.93), compared with gastritis, in the infected patients who had dupA positive strains; however this association was not statistically significant. The results of urease activity showed a significant mean difference between the isolated strains from patients with PUD and NUD (P = 0.034). This activity was relatively higher among patients with intestinal metaplasia. Also a significant association was found between the lack of cagA and increased urease activity among the isolated strains (P = 0.036). While the greatest sequence variation of ureB was detected in a strain from a patient with intestinal metaplasia, the sole determined amino acid change in UreB sequence (Ala201Thr, 30%), showed no influence on urease activity. In conclusion, the supposed role of H. pylori urease to form peptic ulcer and advancing of intestinal metaplasia was postulated in this study. Higher urease activity in the colonizing H. pylori strains that present specific virulence factors was indicated as a risk factor for promotion of histopathological changes of gastric tissue that advance gastric malignancy.

  5. 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)

    Deng, Lei; Li, Wei; Zhong, Zhijun; Gong, Chao; 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.

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

    Directory of Open Access Journals (Sweden)

    Lei Deng

    Full Text Available 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.

  7. [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.

  8. Detection and genotyping of human papillomavirus in gynaecologic outpatients of Messina, eastern Sicily, Italy.

    Science.gov (United States)

    Giuffrè, Giuseppe; Simone, Angela; Todaro, Paolo; Le Donne, Maria; Caruso, Carmela; Pizzo, Alfonsa; Granese, Domenico

    2010-03-01

    In order to determine the prevalence of human papillomavirus (HPV) infection in sexually active female population in Messina, we tested cervical scrapes of women referred to university clinics for routine gynaecologic care. Between March and December 2008, a total of 680 cervical samples of 598 patients (573 Italian from province of Messina and 25 resident aliens) were examined consecutively from laboratory of molecular biology at the Department of Human Pathology. For each sample, cervical cells were collected by centrifugation and DNA was extracted (QIAamp DNA mini kit, Qiagen), followed by a PCR-based HPV DNA assay and reverse dot blot genotyping (HPV-HS Bio plus HPV-strip, AB Analytica or HPV-type, AB Analytica). The overall rate of HPV DNA detection in Italian patients (mean age 34 years; range 15-69) was 70.5% (404/573), with 163 cases of multiple infections (40.3%). In 335 patients (82.9%) a high-risk HPV infection was detected. In this group the coexistence of a low-risk HPV infection was documented in 97 cases while 65 patients exhibited only a low-risk HPV infection. HPV-16 was the most prevalent (33.4%), followed by HPV-6 (28.0%), HPV-31 (24.3%), HPV-58 (11.4%), HPV-66 (11.1%), HPV-53 (6.4%), HPV-18 (6.2%), HPV-56 (5.4%), HPV-33 (5.2%) while the other genotypes identified (HPV-11, -40, -42, -43, -44, -54, -61, -70, -81, -26, -35, -39, -45, -51, -52, -59, -68, -73, -82) were below 5%. HPV prevalence (any type) was 78.7% at age or =45 years. A significant association (chi2=12.718; P=0.006) between HPV DNA detection and the younger age was encountered. Since available data on the prevalence and distribution of HPV infection in Italy are somewhat discordant, this study represents a helpful contribution to the knowledge on the circulation of precise genotypes in east Sicily in order to improve new HPV vaccines.

  9. Detection of oncogenic human papillomavirus genotypes on spermatozoa from male partners of infertile couples.

    Science.gov (United States)

    Schillaci, Rosaria; Capra, Giuseppina; Bellavia, Carmela; Ruvolo, Giovanni; Scazzone, Concetta; Venezia, Renato; Perino, Antonio

    2013-11-01

    To evaluate the prevalence of human papillomavirus (HPV) sperm infection and its correlation with sperm parameters in patients who attended a fertility clinic. Cross-sectional clinical study. University-affiliated reproductive medicine clinic. A total of 308 male partners of couples undergoing in vitro fertilization techniques. Specimens of semen were collected from all patients. Sperm parameters were evaluated according to the World Health Organization manual. The presence of HPV DNA was researched by the combined use of two HPV assays and a highly sensitive nested polymerase chain reaction assay followed by HPV genotyping. To examine whether HPV was associated with the sperm, in situ hybridization (ISH) analysis was performed. Results of HPV investigation were compared with sperm parameters and ISH analysis. Twenty-four out of 308 semen samples (7.8%) were HPV DNA positive, but HPV infection did not seem to affect semen quality. Moreover, ISH revealed a clear HPV localization at the equatorial region of sperm head in infected samples. Oncogenic HPV genotypes were detected on spermatozoa from asymptomatic subjects, but a role of the infection in male infertility was not demonstrated. Copyright © 2013 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  10. 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

    2015-09-16

    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

  11. Phenotypic- and Genotypic-Resistance Detection for Adaptive Resistance Management in Tetranychus urticae Koch.

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    Deok Ho Kwon

    Full Text Available Rapid resistance detection is necessary for the adaptive management of acaricide-resistant populations of Tetranychus urticae. Detection of phenotypic and genotypic resistance was conducted by employing residual contact vial bioassay (RCV and quantitative sequencing (QS methods, respectively. RCV was useful for detecting the acaricide resistance levels of T. urticae, particularly for on-site resistance detection; however, it was only applicable for rapid-acting acaricides (12 out of 19 tested acaricides. QS was effective for determining the frequencies of resistance alleles on a population basis, which corresponded to 12 nonsynonymous point mutations associated with target-site resistance to five types of acaricides [organophosphates (monocrotophos, pirimiphos-methyl, dimethoate and chlorpyrifos, pyrethroids (fenpropathrin and bifenthrin, abamectin, bifenazate and etoxazole]. Most field-collected mites exhibited high levels of multiple resistance, as determined by RCV and QS data, suggesting the seriousness of their current acaricide resistance status in rose cultivation areas in Korea. The correlation analyses revealed moderate to high levels of positive relationships between the resistance allele frequencies and the actual resistance levels in only five of the acaricides evaluated, which limits the general application of allele frequency as a direct indicator for estimating actual resistance levels. Nevertheless, the resistance allele frequency data alone allowed for the evaluation of the genetic resistance potential and background of test mite populations. The combined use of RCV and QS provides basic information on resistance levels, which is essential for choosing appropriate acaricides for the management of resistant T. urticae.

  12. 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.

  13. 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

    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.

  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. Population genetic diversity and hybrid detection in captive zebras.

    Science.gov (United States)

    Ito, Hideyuki; Langenhorst, Tanya; Ogden, Rob; Inoue-Murayama, Miho

    2015-08-21

    Zebras are members of the horse family. There are three species of zebras: the plains zebra Equus quagga, the Grevy's zebra E. grevyi and the mountain zebra E. zebra. The Grevy's zebra and the mountain zebra are endangered, and hybridization between the Grevy's zebra and the plains zebra has been documented, leading to a requirement for conservation genetic management within and between the species. We characterized 28 microsatellite markers in Grevy's zebra and assessed cross-amplification in plains zebra and two of its subspecies, as well as mountain zebra. A range of standard indices were employed to examine population genetic diversity and hybrid populations between Grevy's and plains zebra were simulated to investigate subspecies and hybrid detection. Microsatellite marker polymorphism was conserved across species with sufficient variation to enable individual identification in all populations. Comparative diversity estimates indicated greater genetic variation in plains zebra and its subspecies than Grevy's zebra, despite potential ascertainment bias. Species and subspecies differentiation were clearly demonstrated and F1 and F2 hybrids were correctly identified. These findings provide insights into captive population genetic diversity in zebras and support the use of these markers for identifying hybrids, including the known hybrid issue in the endangered Grevy's zebra.

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

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    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.

  17. Genotypic and phenotypic diversity in tropical strains of Aspergillus spp. (section Circumdati) isolated from insects.

    Science.gov (United States)

    Moraes, Aurea; Holanda, Veronica; Zahner, Viviane

    2006-04-01

    The morphology, multilocus enzyme electrophoresis (MLEE), and RAPD-PCR profiles of a panel of 63 strains of Aspergilus section Circumdati, all isoloated from Brazilian insects, were examined. When compared to the descriptions reported in the literature, differences were observed in terms of colony diameter for the representatives studies. Numerical taxonomy based on data generated by MLEE identified two distinct subgroups among the A. ochraceus isolates. In addition, phosphoglucose isomerase (GPI-1) was detected only in A. sclerotiorum, while phosphofructokinase (FK-1) and acid phosphatase (ACP-2) were present only in strains of A. sulphureus, suggesting that these alleles (bands) could be used for species-specific detection. Using RAPD-PCR, species-specific molecular markers were identified for both A. petrakii and A. sulphureus. These results are important from the taxonomic viewpoint and may also be used in the design of screening programs for the isoloation of new strains.

  18. Preliminary concepts for detecting diversion of LWR spent fuel

    International Nuclear Information System (INIS)

    Sellers, T.A.

    Sandia Laboratories, under the sponsorship of the Department of Energy, Office of Safeguards and Security, has been developing conceptual designs of advanced systems to rapidly detect diversion of LWR spent fuel. Three detection options have been identified and compared on the basis of timeliness of detection and cost. Option 1 is based upon inspectors visiting each facility on a periodic basis to obtain and review data acquired by surveillance instruments and to verify the inventory. Option 2 is based upon continuous inspector presence, aided by surveillance instruments. Option 3 is based upon the collection of data from surveillance instruments with periodic readout either at the facility or at a remote central monitoring and display module and occasional inspection. Surveillance instruments are included in each option to assure a sufficiently high probability of detection. An analysis technique with an example logic tree that was used to identify performance requirements is described. A conceptual design has been developed for Option 3 and the essential hardware elements are not being developed. These elements include radiation, crane and pool acoustic sensors, a Data Collection Module, a Local Collection Module, a Local Display Module and a Central Monitoring and Display Module. A demonstration, in operating facilities, of the overall system concept is planned for the March to June 1979 time frame

  19. Diversity in boron toxicity tolerance of Australian barley (Hordeum vulgare L.) genotypes.

    Science.gov (United States)

    Hayes, Julie E; Pallotta, Margaret; Garcia, Melissa; Öz, Mehmet Tufan; Rongala, Jay; Sutton, Tim

    2015-09-26

    Boron (B) is an important micronutrient for plant growth, but is toxic when levels are too high. This commonly occurs in environments with alkaline soils and relatively low rainfall, including many of the cereal growing regions of southern Australia. Four major genetic loci controlling tolerance to high soil B have been identified in the landrace barley, Sahara 3771. Genes underlying two of the loci encode the B transporters HvBot1 and HvNIP2;1. We investigated sequence and expression level diversity in HvBot1 and HvNIP2;1 across barley germplasm, and identified five novel coding sequence alleles for HvBot1. Lines were identified containing either single or multiple copies of the Sahara HvBot1 allele. We established that only the tandemly duplicated Sahara allele conferred B tolerance, and this duplicated allele was found only in a set of nine lines accessioned in Australian collections as Sahara 3763-3771. HvNIP2;1 coding sequences were highly conserved across barley germplasm. We identified the likely causative SNP in the 5'UTR of Sahara HvNIP2;1, and propose that the creation of a small upstream open reading frame interferes with HvNIP2;1 translation in Sahara 3771. Similar to HvBot1, the tolerant HvNIP2;1 allele was unique to the Sahara barley accessions. We identified a new source of the 2H B tolerance allele controlling leaf symptom development, in the landrace Ethiopia 756. Ethiopia 756, as well as the cultivar Sloop Vic which carries both the 2H and HvBot1 B tolerance alleles derived from Sahara 3771, may be valuable as alternative parents in breeding programs targeted to high soil B environments. There is significant diversity in B toxicity tolerance among contemporary Australian barley varieties but this is not related to variation at any of the four known B tolerance loci, indicating that novel, as yet undiscovered, sources of tolerance exist.

  20. Diversity and Antimicrobial Resistance Genotypes in Non-Typhoidal Salmonella Isolates from Poultry Farms in Uganda

    Directory of Open Access Journals (Sweden)

    Terence Odoch

    2018-02-01

    Full Text Available Non-typhoidal Salmonella (NTS are foodborne pathogens of global public health significance. The aim of this study was to subtype a collection of 85 NTS originating from poultry farms in Uganda, and to evaluate a subgroup of phenotypically resistant isolates for common antimicrobial resistance genes and associated integrons. All isolates were subtyped by pulsed-field gel electrophoresis (PFGE. Phenotypically resistant isolates (n = 54 were screened by PCR for the most relevant AMR genes corresponding to their phenotypic resistance pattern, and all 54 isolates were screened by PCR for the presence of integron class 1 and 2 encoding genes. These genes are known to commonly encode resistance to ampicillin, tetracycline, ciprofloxacin, trimethoprim, sulfonamide and chloramphenicol. PFGE revealed 15 pulsotypes representing 11 serotypes from 75 isolates, as 10 were non-typable. Thirty one (57.4% of the 54 resistant isolates carried at least one of the seven genes (blaTEM-1, cmlA, tetA, qnrS, sul1, dhfrI, dhfrVII identified by PCR and six (11% carried class 1 integrons. This study has shown that a diversity of NTS-clones are present in Ugandan poultry farm settings, while at the same time similar NTS-clones occur in different farms and areas. The presence of resistance genes to important antimicrobials used in human and veterinary medicine has been demonstrated, hence the need to strengthen strategies to combat antimicrobial resistance at all levels.

  1. 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.

  2. 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.

  3. 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.

  4. 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.

  5. Staphylococcus aureus Isolates from Bovine Mastitis in Eight Countries: Genotypes, Detection of Genes Encoding Different Toxins and Other Virulence Genes

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    Valentina Monistero

    2018-06-01

    Full Text Available Staphylococcus aureus is recognized worldwide as one of the major agents of dairy cow intra-mammary infections. This microorganism can express a wide spectrum of pathogenic factors used to attach, colonize, invade and infect the host. The present study evaluated 120 isolates from eight different countries that were genotyped by RS-PCR and investigated for 26 different virulence factors to increase the knowledge on the circulating genetic lineages among the cow population with mastitis. New genotypes were observed for South African strains while for all the other countries new variants of existing genotypes were detected. For each country, a specific genotypic pattern was found. Among the virulence factors, fmtB, cna, clfA and leucocidins genes were the most frequent. The sea and sei genes were present in seven out of eight countries; seh showed high frequency in South American countries (Brazil, Colombia, Argentina, while sel was harboured especially in one Mediterranean country (Tunisia. The etb, seb and see genes were not detected in any of the isolates, while only two isolates were MRSA (Germany and Italy confirming the low diffusion of methicillin resistance microorganism among bovine mastitis isolates. This work demonstrated the wide variety of S. aureus genotypes found in dairy cattle worldwide. This condition suggests that considering the region of interest might help to formulate strategies for reducing the infection spreading.

  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

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

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    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.

  8. Evaluation of a New Test, GenoType HelicoDR, for Molecular Detection of Antibiotic Resistance in Helicobacter pylori▿

    Science.gov (United States)

    Cambau, Emmanuelle; Allerheiligen, Vera; Coulon, Céline; Corbel, Céline; Lascols, Christine; Deforges, Lionel; Soussy, Claude-James; Delchier, Jean-Charles; Megraud, Francis

    2009-01-01

    The eradication rate of Helicobacter pylori by standard therapy is decreasing due to antibiotic resistance, mainly to clarithromycin. Our aim was to provide a new molecular test to guide the treatment of new and relapsed cases. We first studied 126 H. pylori strains for phenotypic (MIC) and genotypic resistance to clarithromycin (rrl mutation) and levofloxacin (gyrA mutation) and then developed a DNA strip genotyping test on the basis of the correlation results and literature data. Clinical strains (n = 92) and gastric biopsy specimens containing H. pylori (n = 105) were tested blindly with the new molecular test GenoType HelicoDR. The presence of mutations or the absence of hybridization with wild-type sequences was predictive, in rrl for clarithromycin resistance in 91 cases (mostly the A2147G mutation) and in gyrA for levofloxacin resistance in 58 cases (mutations at codon 87 or 91). Genotyping revealed a mix of genotypes in 33% of the cases, reflecting a coinfection or selection for resistant mutants. The sensitivity and specificity of detecting resistance were 94% and 99% for clarithromycin and 87% and 98.5% for levofloxacin, respectively. The concordance scores were 0.96 for clarithromycin and 0.94 for levofloxacin. With global resistance rates of 46% for clarithromycin and 25% for levofloxacin, which were observed for consecutive positive biopsy specimens from 2007 and 2008, the positive and negative predictive values for detecting resistance were 99% and 94% for clarithromycin and 96% and 96% for fluoroquinolone. GenoType HelicoDR is efficient at detecting mutations predictive of antibiotic resistance in H. pylori when applied to strains or directly to gastric biopsy specimens. PMID:19759218

  9. Evaluation of a new test, genotype HelicoDR, for molecular detection of antibiotic resistance in Helicobacter pylori.

    Science.gov (United States)

    Cambau, Emmanuelle; Allerheiligen, Vera; Coulon, Céline; Corbel, Céline; Lascols, Christine; Deforges, Lionel; Soussy, Claude-James; Delchier, Jean-Charles; Megraud, Francis

    2009-11-01

    The eradication rate of Helicobacter pylori by standard therapy is decreasing due to antibiotic resistance, mainly to clarithromycin. Our aim was to provide a new molecular test to guide the treatment of new and relapsed cases. We first studied 126 H. pylori strains for phenotypic (MIC) and genotypic resistance to clarithromycin (rrl mutation) and levofloxacin (gyrA mutation) and then developed a DNA strip genotyping test on the basis of the correlation results and literature data. Clinical strains (n = 92) and gastric biopsy specimens containing H. pylori (n = 105) were tested blindly with the new molecular test GenoType HelicoDR. The presence of mutations or the absence of hybridization with wild-type sequences was predictive, in rrl for clarithromycin resistance in 91 cases (mostly the A2147G mutation) and in gyrA for levofloxacin resistance in 58 cases (mutations at codon 87 or 91). Genotyping revealed a mix of genotypes in 33% of the cases, reflecting a coinfection or selection for resistant mutants. The sensitivity and specificity of detecting resistance were 94% and 99% for clarithromycin and 87% and 98.5% for levofloxacin, respectively. The concordance scores were 0.96 for clarithromycin and 0.94 for levofloxacin. With global resistance rates of 46% for clarithromycin and 25% for levofloxacin, which were observed for consecutive positive biopsy specimens from 2007 and 2008, the positive and negative predictive values for detecting resistance were 99% and 94% for clarithromycin and 96% and 96% for fluoroquinolone. GenoType HelicoDR is efficient at detecting mutations predictive of antibiotic resistance in H. pylori when applied to strains or directly to gastric biopsy specimens.

  10. Detection, identification and genotyping of Borrellia spp. in rodents in Slovenia by PCR and culture.

    Science.gov (United States)

    Cerar, Tjaša; Korva, Miša; Avšič-Županc, Tatjana; Ružić-Sabljić, Eva

    2015-08-08

    Borrelia burgdorferi sensu lato, the agent of Lyme borreliosis, is mainly maintained in natural foci through the transmission cycles of competent tick vectors (Ixodes sp.) and a vertebrate reservoir. Specific rodents have been identified as the principal reservoir of Borrelia burgdorferi sensu lato in Europe. Borrelia miyamotoi is the only relapsing fever spirochete transmitted by the same tick. The aim of the present study was to perform an epidemiological survey to determine the presence of B. burgdorferi sensu lato in rodents occurring in Slovenia and to explore the presence of Borrelia miyamotoi. The study was performed in two parts, retrospective and prospective; a total of 297 rodents was analyzed. Detection and identification of borrelia was performed by molecular methods and additionally in the prospective study by isolation and genotyping (MluI-LRFP and MLST). During the prospective part of the study, borrelia was isolated from 2/46 (4.3 %) lung specimens and from 10/46 (21.7 %) heart specimens of rodents. All isolated strains were identified as B. afzelii subtype Mla1, and MLST analysis revealed 5 distinct sequence types. Borrelia DNA was successfully detected by one or other of the PCR methods in 18/46 (39.1 %) and 75/251 (29.9 %) samples in the prospective and retrospective studies, respectively. LightMix® was found to be more sensitive than the ''in-house" nested PCR (91/297 (30.6 %) vs 48/297 (16.1 %)). Borrelia miyamotoi DNA was detected in 1/251 (0.4 %) and in 1/46 (2.2 %) heart specimens, in the retrospective and prospective parts of the study, respectively. We determined the prevalence of B. afzelii in rodents and report for the first time the presence of B. miyamotoi in Slovenia.

  11. Cluster-distinguishing genotypic and phenotypic diversity of carbapenem-resistant Gram-negative bacteria in solid-organ transplantation patients: a comparative study.

    Science.gov (United States)

    Karampatakis, Theodoros; Geladari, Anastasia; Politi, Lida; Antachopoulos, Charalampos; Iosifidis, Elias; Tsiatsiou, Olga; Karyoti, Aggeliki; Papanikolaou, Vasileios; Tsakris, Athanassios; Roilides, Emmanuel

    2017-07-31

    Solid-organ transplant recipients may display high rates of colonization and/or infection by multidrug-resistant bacteria. We analysed and compared the phenotypic and genotypic diversity of carbapenem-resistant (CR) strains of Klebsiella pneumoniae, Pseudomonas aeruginosa and Acinetobacter baumannii isolated from patients in the Solid Organ Transplantation department of our hospital. Between March 2012 and August 2013, 56 CR strains from various biological fluids underwent antimicrobial susceptibility testing with VITEK 2, molecular analysis by PCR amplification and genotypic analysis with pulsed-field gel electrophoresis (PFGE). They were clustered according to antimicrobial drug susceptibility and genotypic profiles. Diversity analyses were performed by calculating Simpson's diversity index and applying computed rarefaction curves.Results/Key findings. Among K. pneumoniae, KP-producers predominated (57.1 %). VIM and OXA-23 carbapenemases prevailed among P. aeruginosa and A. baumannii (89.4 and 88.9 %, respectively). KPC-producing K. pneumoniae and OXA-23 A. baumannii were assigned in single PFGE pulsotypes. VIM-producing P. aeruginosa generated multiple pulsotypes. CR K. pneumoniae strains displayed phenotypic diversity in tigecycline, colistin (CS), amikacin (AMK), gentamicin (GEN) and co-trimoxazole (SXT) (16 clusters); P. aeruginosa displayed phenotypic diversity in cefepime (FEP), ceftazidime, aztreonam, piperacillin, piperacillin-tazobactam, AMK, GEN and CS (9 clusters); and A. baumannii displayed phenotypic diversity in AMK, GEN, SXT, FEP, tobramycin and rifampicin (8 clusters). The Simpson diversity indices for the interpretative phenotype and PFGE analysis were 0.89 and 0.6, respectively, for K. pneumoniae strains (P<0.001); 0.77 and 0.6 for P. aeruginosa (P=0.22); and 0.86 and 0.19 for A. baumannii (P=0.004). The presence of different antimicrobial susceptibility profiles does not preclude the possibility that two CR K. pneumoniae or A. baumannii

  12. 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

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

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    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.

  14. 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

    2017-06-01

    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. 89:1015-1024, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  15. 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. Copyright © 2012 Mayo Foundation for Medical Education and Research. Published by Elsevier Inc. All rights reserved.

  16. Detection and genotyping of Toxoplasma gondii DNA in the blood and milk of naturally infected donkeys (Equus asinus).

    Science.gov (United States)

    Mancianti, Francesca; Nardoni, Simona; Papini, Roberto; Mugnaini, Linda; Martini, Mina; Altomonte, Iolanda; Salari, Federica; D'Ascenzi, Carlo; Dubey, Jitender P

    2014-04-03

    Toxoplasma gondii is a worldwide zoonotic protozoan. Consumption of raw milk from infected animals is considered a risk factor for acquiring toxoplasmosis in humans. Recently, donkey milk has been indicated for therapeutic and nutritional purposes and T. gondii infection is common in donkeys. The purpose of the present paper was to detect the presence of parasite DNA in milk of T. gondii positive donkeys. Antibodies to T. gondii were found in 11 out of 44 healthy lactating donkeys by IFAT. T. gondii DNA was detected by PCR in blood of 6 and milk of 3 seropositive jennies. Results of limited RFLP-PCR genotyping indicated the presence of T. gondii genotype II or III, commonly found in Europe. The occurrence of T. gondii DNA in milk suggests that the consumption of raw milk from seropositive donkeys could be a potential source of human infection.

  17. Evaluation of genotype MTBDRplus assay for rapid detection of isoniazid and rifampicin resistance in Mycobacterium tuberculosis clinical isolates from Pakistan

    Directory of Open Access Journals (Sweden)

    Hasnain Javed

    2016-01-01

    Conclusions: As evidenced in this study, the major concern with the GenoType MTBDRplus assay were false negative results. In comparison to conventional drug susceptibility testing, the assay was unable to detect 30 (30/100; 30% strains resistant to INH and 23 (23/100; 23% strains resistant to RMP. The GenoType MTBDRplus failed to identify 38 MDR (38/100; 38% strains. Resistance in those strains probably originate from mutations in other codons and/or genes than those covered by the test. For detecting INH and RMP resistance in TB cases, especially in high TB incidence countries, such as Pakistan, molecular approaches should still be a complement rather than areplacement to conventional drug susceptibility testing.

  18. Assessment of Genetic Variation and Population Structure of Diverse Rice Genotypes Adapted to Lowland and Upland Ecologies in Africa Using SNPs

    Directory of Open Access Journals (Sweden)

    Marie Noelle Ndjiondjop

    2018-04-01

    Full Text Available Using interspecific crosses involving Oryza glaberrima Steud. as donor and O. sativa L. as recurrent parents, rice breeders at the Africa Rice Center developed several ‘New Rice for Africa (NERICA’ improved varieties. A smaller number of interspecific and intraspecific varieties have also been released as ‘Advanced Rice for Africa (ARICA’. The objective of the present study was to investigate the genetic variation, relatedness, and population structure of 330 widely used rice genotypes in Africa using DArTseq-based single nucleotide polymorphisms (SNPs. A sample of 11 ARICAs, 85 NERICAs, 62 O. sativa spp. japonica, and 172 O. sativa spp. indica genotypes were genotyped with 27,560 SNPs using diversity array technology (DArT-based sequencing (DArTseq platform. Nearly 66% of the SNPs were polymorphic, of which 15,020 SNPs were mapped to the 12 rice chromosomes. Genetic distance between pairs of genotypes that belong to indica, japonica, ARICA, and NERICA varied from 0.016 to 0.623, from 0.020 to 0.692, from 0.075 to 0.763, and from 0.014 to 0.644, respectively. The proportion of pairs of genotypes with genetic distance > 0.400 was the largest within NERICAs (35.1% of the pairs followed by ARICAs (18.2%, japonica (17.4%, and indica (5.6%. We found one pair of japonica, 11 pairs of indica, and 35 pairs of NERICA genotypes differing by <2% of the total scored alleles, which was due to 26 pairs of genotypes with identical pedigrees. Cluster analysis, principal component analysis, and the model-based population structure analysis all revealed two distinct groups corresponding to the lowland (primarily indica and lowland NERICAs and upland (japonica and upland NERICAs growing ecologies. Most of the interspecific lowland NERICAs formed a sub-group, likely caused by differences in the O. glaberrima genome as compared with the indica genotypes. Analysis of molecular variance revealed very great genetic differentiation (FST = 0.688 between the

  19. Assessment of Genetic Variation and Population Structure of Diverse Rice Genotypes Adapted to Lowland and Upland Ecologies in Africa Using SNPs.

    Science.gov (United States)

    Ndjiondjop, Marie Noelle; Semagn, Kassa; Sow, Mounirou; Manneh, Baboucarr; Gouda, Arnaud C; Kpeki, Sèdjro B; Pegalepo, Esther; Wambugu, Peterson; Sié, Moussa; Warburton, Marilyn L

    2018-01-01

    Using interspecific crosses involving Oryza glaberrima Steud. as donor and O. sativa L. as recurrent parents, rice breeders at the Africa Rice Center developed several 'New Rice for Africa (NERICA)' improved varieties. A smaller number of interspecific and intraspecific varieties have also been released as 'Advanced Rice for Africa (ARICA)'. The objective of the present study was to investigate the genetic variation, relatedness, and population structure of 330 widely used rice genotypes in Africa using DArTseq-based single nucleotide polymorphisms (SNPs). A sample of 11 ARICAs, 85 NERICAs, 62 O. sativa spp. japonica , and 172 O. sativa spp. indica genotypes were genotyped with 27,560 SNPs using diversity array technology (DArT)-based sequencing (DArTseq) platform. Nearly 66% of the SNPs were polymorphic, of which 15,020 SNPs were mapped to the 12 rice chromosomes. Genetic distance between pairs of genotypes that belong to indica, japonica, ARICA, and NERICA varied from 0.016 to 0.623, from 0.020 to 0.692, from 0.075 to 0.763, and from 0.014 to 0.644, respectively. The proportion of pairs of genotypes with genetic distance > 0.400 was the largest within NERICAs (35.1% of the pairs) followed by ARICAs (18.2%), japonica (17.4%), and indica (5.6%). We found one pair of japonica, 11 pairs of indica, and 35 pairs of NERICA genotypes differing by <2% of the total scored alleles, which was due to 26 pairs of genotypes with identical pedigrees. Cluster analysis, principal component analysis, and the model-based population structure analysis all revealed two distinct groups corresponding to the lowland (primarily indica and lowland NERICAs) and upland (japonica and upland NERICAs) growing ecologies. Most of the interspecific lowland NERICAs formed a sub-group, likely caused by differences in the O. glaberrima genome as compared with the indica genotypes. Analysis of molecular variance revealed very great genetic differentiation ( F ST = 0.688) between the lowland and upland

  20. Characterization of diverse soybean genotypes using 14 carbon for organic acid exudation under phosphorus stress and its relationship with phosphorus acquisition efficiency

    International Nuclear Information System (INIS)

    Vengavasi, Krishnapriya; Pandey, Renu

    2017-01-01

    To characterise the physiological aspects influencing organic acid exudation under low phosphorus (P) stress, experiment was conducted to screen a diverse soybean panel (comprising 116 genotypes) for total carbon exudation employing shoot labelling with 14 CO 2 technique. Among the traits measured at seedling stage, total carbon ( 14 C) exudation, P uptake and total dry weight contributed to the maximum genotypic variability in soybean. The proportion of organic acids (including oxalate, citrate, succinate and fumarate) was the highest among root-exuded compounds induced by low P stress in soybean. Improved root length, surface area and volume coupled with higher activity of enzymes in TCA cycle contributed to enhanced organic acid exudation under low P. Efficient soybean genotypes (EC-232019 and G-2344) exhibited superior growth and P acquisition efficiency under low soil P availability attributed to its higher root exudation potential aiding in mining fixed soil P. To understand the molecular mechanism differentially regulating root exudation potential in contrasting genotypes (EC-232019 and EC-113396), root proteome analysis at low P stress was carried out. Among the total proteins visualised by 2D-gel electrophoresis, 105 genotypes (32%) were differentially expressed between sufficient and low P levels. A total of 44 (14%) proteins were down-regulated by more than two-fold while 61 (15%) proteins were up-regulated by more than two-fold at low P. The differential proteins were involved in a myriad of functions including organic acid accumulation, carbohydrate, protein and lipid metabolism under low P stress. Characterisation of 17 proteins with unknown function indicated the role of novel genes under low P stress. The identified genotypes have potential to be used as donors in crop improvement programs to develop high-yielding P-efficient cultivars which may be an asset to low-input sustainable agriculture. (author)

  1. The IAEA concept of detection of diversion through nuclear material accountancy (2)

    International Nuclear Information System (INIS)

    Akiba, Mitsunori

    2005-01-01

    Diversion into D (falsification of accounting report) and diversion into MUF could be detected by the Inspectorate through nuclear material accountancy. The Inspectorate designs inspection activities to detect diversion into D in cost effective ways. As a result, detection of diversion into D is divided into two statistics, one is item difference statistics which could detect major defects and the other is material balance statistics which could detect remaining small defects. MUF statistics could detect Diversion into MUF. Item statistics has many useful characteristics from safeguards view points, so it is examined in details. Material balance statistics and MUF statistics stem from measurement error associated with equipment inevitably. The above-mentioned concept is called 'IAEA decision structure'. Hereafter, designing safeguards (inspection activities) approach will be based on the IAEA decision structure. (author)

  2. Recognition of Y Fragment Deletion by Genotyping Graphs after Amplified by PowerPlex® 21 Detection Kit.

    Science.gov (United States)

    Wang, S C; Ding, M M; Wei, X L; Zhang, T; Yao, F

    2016-06-01

    To recognize the possibility of Y fragment deletion of Amelogenin gene intuitively and simply according to the genotyping graphs. By calculating the ratio of total peak height of genotyping graphs, the statistics of equilibrium distribution between Amelogenin and D3S1358 loci, Amelogenin X-gene and Amelogenin Y-gene, and different alleles of D3S1358 loci from 1 968 individuals was analyzed after amplified by PowerPlex ® 21 detection kit. Sum of peak height of Amelogenin X allele was not less than 60% that of D3S1358 loci alleles in 90.8% female samples, and sum of peak height of Amelogenin X allele was not higher than 70% that of D3S1358 loci alleles in 94.9% male samples. The result of genotyping after amplified by PowerPlex ® 21 detection kit shows that the possibility of Y fragment deletion should be considered when only Amelogenin X-gene of Amelogenin is detected and the peak height of Amelogenin X-gene is not higher than 70% of the total peak height of D3S1358 loci. Copyright© by the Editorial Department of Journal of Forensic Medicine

  3. 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

  4. 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.

  5. Bacterial diversity on the surface of potato tubers in soil and the influence of the plant genotype.

    Science.gov (United States)

    Weinert, Nicole; Meincke, Remo; Gottwald, Christine; Heuer, Holger; Schloter, Michael; Berg, Gabriele; Smalla, Kornelia

    2010-10-01

    The surface of tubers might be a reservoir for bacteria that are disseminated with seed potatoes or that affect postharvest damage. The numbers of culturable bacteria and their antagonistic potential, as well as bacterial community fingerprints were analysed from tubers of seven field-grown potato genotypes, including two lines with tuber-accumulated zeaxanthin. The plant genotype significantly affected the number of culturable bacteria only at one field site. Zeaxanthin had no effect on the bacterial plate counts. In dual culture, 72 of 700 bacterial isolates inhibited at least one of the potato pathogens Rhizoctonia solani, Verticillium dahliae or Phytophthora infestans, 12 of them suppressing all three. Most of these antagonists were identified as Bacillus or Streptomyces. From tubers of two plant genotypes, including one zeaxanthin line, higher numbers of antagonists were isolated. Most antagonists showed glucanase, cellulase and protease activity, which could represent mechanisms for pathogen suppression. PCR-DGGE fingerprints of the 16S rRNA genes of bacterial communities from the tuber surfaces revealed that the potato genotype significantly affected the Pseudomonas community structure at one site. However, the genotypes showed nearly identical fingerprints for Bacteria, Actinobacteria, Alphaproteobacteria, Betaproteobacteria, Bacillus and Streptomycetaceae. In conclusion, tuber-associated bacteria were only weakly affected by the plant genotype. © 2010 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  6. Evaluation of diverse wheat genotypes for potential biomass production through physiological parameters at seedling stage under controlled environment

    International Nuclear Information System (INIS)

    Khattak, G.S.S.

    2014-01-01

    Thirty wheat genotypes from UK, CIMMYT and Pakistan were evaluated under controlled environment conditions for their potential biomass production by measuring stomatal conductance (gs; porometry), leaf photosynthesis (IRGA), carbon isotope discrimination and carbon content (isotope ratio and mass analysis) at Rothamsted Research, Harpenden, UK during 2011. Amongst the dwarf genotypes, Rht2 and Rht3 showed lower stomatal conductance than Seri 32B, Seri 87B and Bathoor-07. For these 5 genotypes and another genotype Inqalab photosynthetic performance was determined by means of IRGA measurements. Of these genotypes Inqalab had the highest photosynthetic activity (A), stomatal conductance (gs), transpiration (E) and leaf intercellular CO/sub 2/ but it also had the lowest water use efficiency (A/gs) and intrinsic water use efficiency (A/E). Seri-87B had the greatest water use efficiency (A/gs) and intrinsic water use efficiency (A/E). All the Pakistani genotypes had large stomatal conductances and high 13C delta and thus may be expected to produce high biomass under irrigations and optimum inputs. (author)

  7. Direct Molecular Detection and Genotyping of Borrelia burgdorferi from Whole Blood of Patients with Early Lyme Disease

    Science.gov (United States)

    Eshoo, Mark W.; Crowder, Christopher C.; Rebman, Alison W.; Rounds, Megan A.; Matthews, Heather E.; Picuri, John M.; Soloski, Mark J.; Ecker, David J.; Schutzer, Steven E.; Aucott, John N.

    2012-01-01

    Direct molecular tests in blood for early Lyme disease can be insensitive due to low amount of circulating Borrelia burgdorferi DNA. To address this challenge, we have developed a sensitive strategy to both detect and genotype B. burgdorferi directly from whole blood collected during the initial patient visit. This strategy improved sensitivity by employing 1.25 mL of whole blood, a novel pre-enrichment of the entire specimen extract for Borrelia DNA prior to a multi-locus PCR and electrospray ionization mass spectrometry detection assay. We evaluated the assay on blood collected at the initial presentation from 21 endemic area patients who had both physician-diagnosed erythema migrans (EM) and positive two-tiered serology either at the initial visit or at a follow-up visit after three weeks of antibiotic therapy. Results of this DNA analysis showed detection of B. burgdorferi in 13 of 21 patients (62%). In most cases the new assay also provided the B. burgdorferi genotype. The combined results of our direct detection assay with initial physician visit serology resulted in the detection of early Lyme disease in 19 of 21 (90%) of patients at the initial visit. In 5 of 21 cases we demonstrate the ability to detect B. burgdorferi in early Lyme disease directly from whole blood specimens prior to seroconversion. PMID:22590620

  8. 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.

  9. 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.

  10. 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. PMID:23350011

  11. 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.

  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. 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.

  14. 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.

  15. Detection and characterization of two chimpanzee polyomavirus genotypes from different subspecies

    NARCIS (Netherlands)

    I. Deuzing (Ilona); Z. Fagrouch (Zahra); M.J. Groenewoud (Marlous); H. Niphuis (Henk); I. Kondova (Ivanela); W. Bogers (Willy); E.J. Verschoor (Ernst)

    2010-01-01

    textabstractThe complete nucleotide sequences of three chimpanzee polyomavirus genetic variants were determined. Phylogenetic analysis indicated that the viruses form two different genotypes of ChPyV. Comparison with other primate polyomaviruses revealed a putative agnogene, and an unusually long

  16. 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. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. 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

    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......FOX2 and GmTOR2 were applied as codominant genetic markers together with the LSU rDNA.   Genetic structure of Glomus spp. populations from an organically and a conventionally cultured field were compared by hierarchical sampling of spores from four plots in each field. Multilocus genotypes were...

  18. Genotypes of Pestivirus RNA detected n anti influenza virus vaccines for human use

    Directory of Open Access Journals (Sweden)

    M. Giangaspero

    2004-02-01

    Full Text Available Nine polyvalent human influenza virus vaccines were tested by reverse transcriptase-polymerase chain reaction (RT-PCR for the presence of pestivirus RNA. Samples were selected from manufacturers in Europe and the USA. Three samples of the nine vaccines tested (33.3% gave positive results for pestivirus RNA. The 5´-untranslated genomic region sequence of the contaminant pestivirus RNA was analysed based on primary nucleotide sequence homology and on secondary sequence structures characteristic to genotypes. Two sequences belonged to Pestivirus type-1 (bovine viral diarrhoea virus [BVDV] species, genotypes BVDV-1b and BVDV-1e. These findings confirm previous reports, suggesting an improvement in preventive measures against contamination of biological products for human use.

  19. 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

  20. 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.

  1. Molecular epidemiology and genetic diversity of hepatitis B virus genotype E in an isolated Afro-Colombian community.

    Science.gov (United States)

    Alvarado Mora, Mónica Viviana; Romano, Camila Malta; Gomes-Gouvêa, Michele Soares; Gutierrez, Maria Fernanda; Carrilho, Flair José; Pinho, João Renato Rebello

    2010-02-01

    Hepatitis B virus (HBV) infection is a significant public health concern with 350 million chronic carriers worldwide. Eight HBV genotypes (A-H) have been described so far. Genotype E (HBV/E) is widely distributed in West Africa and has rarely been found in other continents, except for a few cases in individuals with an African background. In this study, we characterized HBV genotypes in Quibdó, Colombia, by partial S/P gene sequencing, and found, for the first time, HBV/E circulating in nine Afro-Colombian patients who had no recent contact with Africa. The presence of HBV/E in this community as a monophyletic group suggests that it was a result of a recent introduction by some Afro-descendent contact or, alternatively, that the virus came with slaves brought to Colombia. By using sequences with sampling dates, we estimated the substitution rate to be about 3.2 x 10(-4) substitutions per site per year, which resulted in a time to the most recent common ancestor (TMRCA) of 29 years. In parallel, we also estimated the TMRCA for HBV/E by using two previously estimated substitution rates (7.7 x 10(-4) and 1.5 x 10(-5) substitutions per site per year). The TMRCA was around 35 years under the higher rate and 1500 years under the slower rate. In sum, this work reports for the first time the presence of an exclusively African HBV genotype circulating in South America. We also discuss the time of the entry of this virus into America based on different substitution rates estimated for HBV.

  2. Genetic Diversity of Plasmodium falciparum Field Isolates in Central Sudan Inferred by PCR Genotyping of Merozoite Surface Protein 1 and 2.

    Science.gov (United States)

    Hamid, Muzamil M Abdel; Mohammed, Sara B; El Hassan, Ibrahim M

    2013-02-01

    Characterization of Plasmodium falciparum diversity is commonly achieved by amplification of the polymorphic regions of the merozoite surface proteins 1 (MSP1) and 2 (MSP2) genes. The present study aimed to determine the allelic variants distribution of MSP1 and MSP2 and multiplicity of infection in P. falciparum field isolates from Kosti, central Sudan, an area characterized by seasonal malaria transmission. Total 121 samples (N = 121) were collected during a cross-sectional survey between March and April 2003. DNA was extracted and MSP1 and MSP2 polymorphic loci were genotyped. The total number of alleles identified in MSP1 block 2 was 11, while 16 alleles were observed in MSP2 block 3. In MSP1, RO33 was found to be the predominant allelic type, carried alone or in combination with MAD20 and K1 types, whereas FC27 family was the most prevalent in MSP2. Sixty two percent of isolates had multiple genotypes and the overall mean multiplicity of infection was 1.93 (CI 95% 1.66-2.20). Age correlated with parasite density (P = 0.017). In addition, a positive correlation was observed between parasite densities and the number of alleles (P = 0.022). Genetic diversity in P. falciparum field isolates in central Sudan was high and consisted of multiple clones.

  3. Diversity

    Science.gov (United States)

    Portraits In Courage Vol. VIII Portraits In Courage Vol. IX Portraits In Courage Vol. X AF Sites Social -Wide Initiative to Promote Diversity and Inclusion in the Federal Workforce Executive Order 13548 : Virtual Diversity Conference Air Force Diversity & Inclusion Air Force Diversity Graphic There is no

  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. Variability in amino acid digestibility of triticale grain from diverse genotypes as studied in cecectomized laying hens.

    Science.gov (United States)

    Zuber, T; Maurer, H P; Möhring, J; Nautscher, N; Siegert, W; Rosenfelder, P; Rodehutscord, M

    2016-12-01

    Triticale, an anthropogenic hybrid grain, is increasing in importance as a feed grain for laying hens. However, our limited knowledge of its nutritional qualities and their impact on hen performance prevents optimization of its use. The present study investigated the digestibility of amino acids ( AA: ) in triticale grain in laying hens, and additionally examined relationships between AA digestibility and chemical and physical characteristics of the grain. Twenty genotypes of triticale were grown under standardized agronomic and environmental conditions and were characterized according to their physical properties (thousand-seed weight, test weight, falling number, extract viscoelasticity), chemical composition (proximate nutrients, non-starch polysaccharides, AA, minerals, inositol phosphates) and gross energy concentration. Additionally, the in vitro solubility of nitrogen was determined. The animal trial comprised 4 Latin Squares (6 × 6) distributed among 2 subsequent runs. Twelve cecectomized LSL-Classic hens were individually housed in metabolism cages and either fed a basal diet containing 500 g/kg cornstarch or one of 20 triticale diets, each replacing the cornstarch with one triticale genotype, for 8 d. During the last 4 d, feed intake was recorded and excreta were collected quantitatively. Amino acid digestibility of the triticale genotypes was calculated by linear regression. The digestibility of all AA differed significantly between the 20 genotypes, including Lys (digestibility range 68 to 80%), Met (77 to 86%), Thr (68 to 78%) and Trp (74 to 83%). However, AA digestibility only correlated with characteristics of the grain in few cases, without a consistent pattern among AA. Equations to predict AA digestibility based on the grain's physical and chemical characteristics were calculated by multiple linear regression. The explanatory power (adjusted R 2 ;) of these prediction equations was below 0.7 for most AA and thus not sufficiently precise to be

  6. High performance of a new PCR-based urine assay for HPV-DNA detection and genotyping.

    Science.gov (United States)

    Tanzi, Elisabetta; Bianchi, Silvia; Fasolo, Maria Michela; Frati, Elena R; Mazza, Francesca; Martinelli, Marianna; Colzani, Daniela; Beretta, Rosangela; Zappa, Alessandra; Orlando, Giovanna

    2013-01-01

    Human papillomavirus (HPV) testing has been proposed as a means of replacing or supporting conventional cervical screening (Pap test). However, both methods require the collection of cervical samples. Urine sample is easier and more acceptable to collect and could be helpful in facilitating cervical cancer screening. The aim of this study was to evaluate the sensitivity and specificity of urine testing compared to conventional cervical smear testing using a PCR-based method with a new, designed specifically primer set. Paired cervical and first voided urine samples collected from 107 women infected with HIV were subjected to HPV-DNA detection and genotyping using a PCR-based assay and a restriction fragment length polymorphism method. Sensitivity, specificity, Positive Predictive Value (PPV), and Negative Predictive Value (NPV) were calculated using the McNemar's test for differences. Concordance between tests was assessed using the Cohen's unweighted Kappa (k). HPV DNA was detected in 64.5% (95% CI: 55.1-73.1%) of both cytobrush and urine samples. High concordance rates of HPV-DNA detection (k = 0.96; 95% CI: 0.90-1.0) and of high risk-clade and low-risk genotyping in paired samples (k = 0.80; 95% CI: 0.67-0.92 and k = 0.74; 95% CI: 0.60-0.88, respectively) were observed. HPV-DNA detection in urine versus cervix testing revealed a sensitivity of 98.6% (95% CI: 93.1-99.9%) and a specificity of 97.4% (95% CI: 87.7-99.9%), with a very high NPV (97.4%; 95% CI: 87.7-99.9%). The PCR-based assay utilized in this study proved highly sensitive and specific for HPV-DNA detection and genotyping in urine samples. These data suggest that a urine-based assay would be a suitable and effective tool for epidemiological surveillance and, most of all, screening programs. Copyright © 2012 Wiley Periodicals, Inc.

  7. Concomitant genotyping revealed diverse spreading between methicillin-resistant Staphylococcus aureus and methicillin-susceptible Staphylococcus aureus in central Taiwan.

    Science.gov (United States)

    Ho, Cheng-Mao; Lin, Chien-Yu; Ho, Mao-Wang; Lin, Hsiao-Chuan; Peng, Ching-Tien; Lu, Jang-Jih

    2016-06-01

    Staphylococcus aureus is a versatile bacterium, which can lead to various infectious diseases. Various molecular typing methods are applied to the evolution and epidemiology surveys of S. aureus, mostly for methicillin-resistant S. aureus (MRSA). However, methicillin-susceptible S. aureus (MSSA) is still an important pathogen, but their molecular typing is evaluated infrequently. Pulsed-field gel electrophoresis (PFGE), spa typing, and detection of five virulent genes for 95 MRSA and 56 MSSA isolates (July-December 2008 and July 2008-December 2009, respectively) during an overlapping period were performed. More diversity was found in MSSA isolates (23 pulsotypes and 25 spa types, excluding 4 new-type and 1 nontypable isolates for spa typing) than in MRSA isolates (19 pulsotypes and 16 spa types, excluding 1 new-type and 1 nontypable isolates for spa typing). By spa typing, t002 (n = 30), t037 (n = 23), t437 (n = 21), t234 (n = 3), t1081 (n = 3), and t1094 (n = 3) were the six major MRSA clones. For MSSA isolates, t189 (n = 13), t437 (n = 4), t084 (n = 3), t213 (n = 3), t701 (n = 3), and t7200 (n = 3) were the six major types. Combining PFGE and spa typing, there were five combinations (pulsotype + spa type) that contained both MRSA and MSSA isolates (pulsotype 9-t437, pulsotype 15-t037, pulsotype 19-t002, pulsotype 21-t002, and pulsotype 28-t1081). For all 151 S. aureus or 95 MRSA isolates, the PFGE typing had more discrimination power, but spa typing had larger discrimination index for 56 MSSA isolates. In conclusion, there were different predominant MRSA and MSSA clones clinically. Continuing longitudinal tracking of molecular typing is necessary for elucidating the evolution of this important clinical pathogen. Copyright © 2014. Published by Elsevier B.V.

  8. Comparative ultrastructure of fruit plastids in three genetically diverse genotypes of apple (Malus × domestica Borkh.) during development.

    Science.gov (United States)

    Schaeffer, Scott M; Christian, Ryan; Castro-Velasquez, Nohely; Hyden, Brennan; Lynch-Holm, Valerie; Dhingra, Amit

    2017-10-01

    Comparative ultrastructural developmental time-course analysis has identified discrete stages at which the fruit plastids undergo structural and consequently functional transitions to facilitate subsequent development-guided understanding of the complex plastid biology. Plastids are the defining organelle for a plant cell and are critical for myriad metabolic functions. The role of leaf plastid, chloroplast, is extensively documented; however, fruit plastids-chromoplasts-are poorly understood, especially in the context of the diverse metabolic processes operating in these diverse plant organs. Recently, in a comparative study of the predicted plastid-targeted proteomes across seven plant species, we reported that each plant species is predicted to harbor a unique set of plastid-targeted proteins. However, the temporal and developmental context of these processes remains unknown. In this study, an ultrastructural analysis approach was used to characterize fruit plastids in the epidermal and collenchymal cell layers at 11 developmental timepoints in three genotypes of apple (Malus × domestica Borkh.): chlorophyll-predominant 'Granny Smith', carotenoid-predominant 'Golden Delicious', and anthocyanin-predominant 'Top Red Delicious'. Plastids transitioned from a proplastid-like plastid to a chromoplast-like plastid in epidermis cells, while in the collenchyma cells, they transitioned from a chloroplast-like plastid to a chloro-chromo-amyloplast plastid. Plastids in the collenchyma cells of the three genotypes demonstrated a diverse array of structures and features. This study enabled the identification of discrete developmental stages during which specific functions are most likely being performed by the plastids as indicated by accumulation of plastoglobuli, starch granules, and other sub-organeller structures. Information regarding the metabolically active developmental stages is expected to facilitate biologically relevant omics studies to unravel the complex

  9. Using AFLP-RGA markers to assess genetic diversity among pigeon pea (Cajanus cajan genotypes in relation to major diseases

    Directory of Open Access Journals (Sweden)

    Prakash G Pati

    2014-06-01

    Full Text Available Resistance gene analog (RGA-anchored amplified fragment length polymorphism (AFLP-RGA marker system was used in order to evaluate genetic relationships among 22 pigeon pea genotypes with varied responses to Fusarium wilt and sterility mosaic disease. Five AFLP-RGA primer combinations (E-CAG/wlrk-S, M-GTG/wlrk-S, M-GTG/wlrk-AS, E-CAT/S1-INV and E-CAG/wlrk-AS produced 173 scorable fragments, of which 157 (90.7% were polymorphic, with an average of 31.4 fragments per primer combination. The polymorphism rates obtained with the five primers were 83.3%, 92.0%, 92.3%, 93.0% and 93.1%, respectively. Mean polymorphic information content (PIC values ranged from 0.24 (with E-CAT/S1-INV to 0.30 (with E-CAG/wlrk-AS, whereas resolving power (RP values varied from 11.06 (with M-GTG/wlrk-S to 25.51 (with E-CAG/wlrk-AS and marker index (MI values ranged from 5.98 (with M-GTG/wlrk-S to 12.30 (with E-CAG/wlrk-AS. We identified a positive correlation between MI and RP (r²=0.98, p<0.05, stronger that that observed for the comparison between PIC and RP (r²=0.88, p<0.05. That implies that either MI or RP is the best parameter for selecting more informative AFLP-RGA primer combinations. The Jaccard coefficient ranged from 0.07 to 0.72, suggesting a broad genetic base in the genotypes studied. A neighbor-joining tree, based on the unweighted pair group method with arithmetic mean, distinguished cultivated species from wild species. The grouping of resistant genotypes in different clusters would help in the selection of suitable donors for resistance breeding in pigeon pea.

  10. Detection of species diversity of arbuscular mycorrhizal fungi (AMF ...

    African Journals Online (AJOL)

    Arbuscular-mycorhizal fungi (AMF) from melon plants grown in Van province, were studied by nested-PCR method to establish colonization ratio of related fungi in plants and to detect the fungi at species level. From 10 different locations, a total of 100 soil samples were taken from rhizosphere area of melon plants.

  11. 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

  12. Detection of Sickle Cell Hemoglobin in Haiti by Genotyping and Hemoglobin Solubility Tests

    Science.gov (United States)

    Carter, Tamar E.; von Fricken, Michael; Romain, Jean R.; Memnon, Gladys; St. Victor, Yves; Schick, Laura; Okech, Bernard A.; Mulligan, Connie J.

    2014-01-01

    Sickle cell disease is a growing global health concern because infants born with the disorder in developing countries are now surviving longer with little access to diagnostic and management options. In Haiti, the current state of sickle cell disease/trait in the population is unclear. To inform future screening efforts in Haiti, we assayed sickle hemoglobin mutations using traditional hemoglobin solubility tests (HST) and add-on techniques, which incorporated spectrophotometry and insoluble hemoglobin separation. We also generated genotype data as a metric for HST performance. We found 19 of 202 individuals screened with HST were positive for sickle hemoglobin, five of whom did not carry the HbS allele. We show that spectrophotometry and insoluble hemoglobin separation add-on techniques could resolve false positives associated with the traditional HST approach, with some limitations. We also discuss the incorporation of insoluble hemoglobin separation observation with HST in suboptimal screening settings like Haiti. PMID:24957539

  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. Understanding and Detecting Supporting Arguments of Diverse Types

    OpenAIRE

    Hua, Xinyu; Wang, Lu

    2017-01-01

    We investigate the problem of sentence-level supporting argument detection from relevant documents for user-specified claims. A dataset containing claims and associated citation articles is collected from online debate website idebate.org. We then manually label sentence-level supporting arguments from the documents along with their types as study, factual, opinion, or reasoning. We further characterize arguments of different types, and explore whether leveraging type information can facilita...

  15. Exploring origins, invasion history and genetic diversity of Imperata cylindrica (L.) P. Beauv. (Cogongrass) in the United States using genotyping by sequencing.

    Science.gov (United States)

    Burrell, A Millie; Pepper, Alan E; Hodnett, George; Goolsby, John A; Overholt, William A; Racelis, Alexis E; Diaz, Rodrigo; Klein, Patricia E

    2015-05-01

    Imperata cylindrica (Cogongrass, Speargrass) is a diploid C4 grass that is a noxious weed in 73 countries and constitutes a significant threat to global biodiversity and sustainable agriculture. We used a cost-effective genotyping-by-sequencing (GBS) approach to identify the reproductive system, genetic diversity and geographic origins of invasions in the south-eastern United States. In this work, we demonstrated the advantage of employing the closely related, fully sequenced crop species Sorghum bicolor (L.) Moench as a proxy reference genome to identify a set of 2320 informative single nucleotide and insertion-deletion polymorphisms. Genetic analyses identified four clonal lineages of cogongrass and one clonal lineage of Imperata brasiliensis Trin. in the United States. Each lineage was highly homogeneous, and we found no evidence of hybridization among the different lineages, despite geographical overlap. We found evidence that at least three of these lineages showed clonal reproduction prior to introduction to the United States. These results indicate that cogongrass has limited evolutionary potential to adapt to novel environments and further suggest that upon arrival to its invaded range, this species did not require local adaptation through hybridization/introgression or selection of favourable alleles from a broad genetic base. Thus, cogongrass presents a clear case of broad invasive success, across a diversity of environments, in a clonal organism with limited genetic diversity. © 2015 John Wiley & Sons Ltd.

  16. Diversity of Survival Patterns among Escherichia coli O157:H7 Genotypes Subjected to Food-Related Stress Conditions.

    Science.gov (United States)

    Elhadidy, Mohamed; Álvarez-Ordóñez, Avelino

    2016-01-01

    The purpose of this study was to evaluate the resistance patterns to food-related stresses of Shiga toxin producing Escherichia coli O157:H7 strains belonging to specific genotypes. A total of 33 E. coli O157:H7 strains were exposed to seven different stress conditions acting as potential selective pressures affecting the transmission of E. coli O157:H7 to humans through the food chain. These stress conditions included cold, oxidative, osmotic, acid, heat, freeze-thaw, and starvation stresses. The genotypes used for comparison included lineage-specific polymorphism, Shiga-toxin-encoding bacteriophage insertion sites, clade type, tir (A255T) polymorphism, Shiga toxin 2 subtype, and antiterminator Q gene allele. Bacterial resistance to different stressors was calculated by determining D-values (times required for inactivation of 90% of the bacterial population), which were then subjected to univariate and multivariate analyses. In addition, a relative stress resistance value, integrating resistance values to all tested stressors, was calculated for each bacterial strain and allowed for a ranking-type classification of E. coli O157:H7 strains according to their environmental robustness. Lineage I/II strains were found to be significantly more resistant to acid, cold, and starvation stress than lineage II strains. Similarly, tir (255T) and clade 8 encoding strains were significantly more resistant to acid, heat, cold, and starvation stress than tir (255A) and non-clade 8 strains. Principal component analysis, which allows grouping of strains with similar stress survival characteristics, separated strains of lineage I and I/II from strains of lineage II, which in general showed reduced survival abilities. Results obtained suggest that lineage I/II, tir (255T), and clade 8 strains, which have been previously reported to be more frequently associated with human disease cases, have greater multiple stress resistance than strains of other genotypes. The results from this

  17. 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. Copyright © 2016 Elsevier B.V. All rights reserved.

  18. Investigating Genetic Diversity Among Cotton Genotypes Available in the Iranian Gene Bank (Gossypium sp. Using ISSR Molecular Marker

    Directory of Open Access Journals (Sweden)

    F Shahriari Ahmadi

    2013-04-01

    Full Text Available Cotton is one of the most important world crops and is considered as a major cash crop in the North East of Iran. All selections in plant breeding are based on diversity and an increase in genetic diversity determines the range of selection. In the present study, 24 cultivars of cotton available at the research station for cotton in the East of Iran -Kashmar- were studied using the ISSR marker. A total number of 13 primers, with repeated simple sequences, were used for the amplification of genomic DNA. Overall, 128 bands were obtained, 109 of which showed polymorphism. To evaluate genetic similarity between cultivars, cluster analysis accompanied by the similarity coefficient developed by Jaccard and Nee (1972, were applied using the UPGMA method. Dendrogram analysis showed a high diversity in the cotton cultivars and two main groups with 70 percent genetic similarity dividing the cotton cultivars into two main groups; namely, tetraploid and diploid. The highest polymorphism percentage was related to 5' (CT8RC3' (100% and the lowest belonged to 5' (AG8YA3' and 5' (TC8G3' (25% primers. Based on the similarity matrix, the highest genetic similarity was found in Varamin and Khordad and the lowest in Avangard and Bakhtegan cultivars. Based on the obtained results, ISSR markers can be efficiently used for the investigation of genetic diversity among cotton cultivars.

  19. Tracking the distribution of Puccinia psidii genotypes that cause rust disease on diverse myrtaceous trees and shrubs

    Science.gov (United States)

    Amy L. Ross-Davis; Rodrigo N. Graca; Acelino C. Alfenas; Tobin L. Peever; Jack W. Hanna; Janice Y. Uchida; Rob D. Hauff; Chris Y. Kadooka; Mee-Sook Kim; Phil G. Cannon; Shigetou Namba; Nami Minato; Sofia Simeto; Carlos A. Perez; Min B. Rayamajhi; Mauricio Moran; D. Jean Lodge; Marcela Arguedas; Rosario Medel-Ortiz; M. Armando Lopez-Ramirez; Paula Tennant; Morag Glen; Ned B. Klopfenstein

    2014-01-01

    Puccinia psidii Winter (Basidiomycota, Uredinales) is a biotrophic rust fungus that was first reported in Brazil from guava in 1884 (Psidium guajava; Winter 1884) and later from eucalypt in 1912 (Joffily 1944). Considered to be of neotropical origin, the rust has also been reported to infect diverse myrtaceous hosts elsewhere in South America, Central America, the...

  20. 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

  1. 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.

  2. Genotypic diversity of oscillatoriacean strains belonging to the genera Geitlerinema and Spirulina determined by 16S rDNA restriction analysis.

    Science.gov (United States)

    Margheri, Maria C; Piccardi, Raffaella; Ventura, Stefano; Viti, Carlo; Giovannetti, Luciana

    2003-05-01

    Genotypic diversity of several cyanobacterial strains mostly isolated from marine or brackish waters, belonging to the genera Geitlerinema and Spirulina, was investigated by amplified 16S ribosomal DNA restriction analysis and compared with morphological features and response to salinity. Cluster analysis was performed on amplified 16S rDNA restriction profiles of these strains along with profiles obtained from sequence data of five Spirulina-like strains, including three representatives of the new genus Halospirulina. Our strains with tightly coiled trichomes from hypersaline waters could be assigned to the Halospirulina genus. Among the uncoiled strains, the two strains of hypersaline origin clustered together and were found to be distant from their counterparts of marine and freshwater habitat. Moreover, another cluster, formed by alkali-tolerant strains with tightly coiled trichomes, was well delineated.

  3. Development and evaluation of novel one-step TaqMan realtime RT-PCR assays for the detection and direct genotyping of genogroup I and II noroviruses

    DEFF Research Database (Denmark)

    Schultz, Anna Charlotte; Vega, Everado; Dalsgaard, Anders

    2011-01-01

    BackgroundCurrent detection and genotyping methods of genogroup (G) I and II noroviruses (NoVs) consist of a 2-step approach including detection of viral RNA by TaqMan realtime RT-PCR (RT-qPCR) followed by conventional RT-PCR and sequencing of partial regions of ORF1 or ORF2. ObjectiveTo develop ......Man RT-qPCR assays for the sensitive detection and direct genotyping of GI and GII NoVs from clinical and environmental matrices...... novel long-template one-step TaqMan assays (L-RT-qPCR) for the rapid detection and direct genotyping of GI and GII NoVs and to evaluate the sensitivity and specificity of the assays. Study designGI and GII-specific broadly reactive L-RT-qPCR assays were developed by combining existing NoV primers...... and probes targeting the open reading frame (ORF)1–ORF2 junction as well as region C at the 5′–ORF2. The assays were validated using GI and GII RNA transcripts and a coded panel of 75 stool samples containing NoV strains representing 9 GI genotypes and 12 GII genotypes, as well as sapoviruses, astroviruses...

  4. MLL2 mutation detection in 86 patients with Kabuki syndrome: a genotype-phenotype study.

    Science.gov (United States)

    Makrythanasis, P; van Bon, B W; Steehouwer, M; Rodríguez-Santiago, B; Simpson, M; Dias, P; Anderlid, B M; Arts, P; Bhat, M; Augello, B; Biamino, E; Bongers, E M H F; Del Campo, M; Cordeiro, I; Cueto-González, A M; Cuscó, I; Deshpande, C; Frysira, E; Izatt, L; Flores, R; Galán, E; Gener, B; Gilissen, C; Granneman, S M; Hoyer, J; Yntema, H G; Kets, C M; Koolen, D A; Marcelis, C l; Medeira, A; Micale, L; Mohammed, S; de Munnik, S A; Nordgren, A; Psoni, S; Reardon, W; Revencu, N; Roscioli, T; Ruiterkamp-Versteeg, M; Santos, H G; Schoumans, J; Schuurs-Hoeijmakers, J H M; Silengo, M C; Toledo, L; Vendrell, T; van der Burgt, I; van Lier, B; Zweier, C; Reymond, A; Trembath, R C; Perez-Jurado, L; Dupont, J; de Vries, B B A; Brunner, H G; Veltman, J A; Merla, G; Antonarakis, S E; Hoischen, A

    2013-12-01

    Recently, pathogenic variants in the MLL2 gene were identified as the most common cause of Kabuki (Niikawa-Kuroki) syndrome (MIM#147920). To further elucidate the genotype-phenotype correlation, we studied a large cohort of 86 clinically defined patients with Kabuki syndrome (KS) for mutations in MLL2. All patients were assessed using a standardized phenotype list and all were scored using a newly developed clinical score list for KS (MLL2-Kabuki score 0-10). Sequencing of the full coding region and intron-exon boundaries of MLL2 identified a total of 45 likely pathogenic mutations (52%): 31 nonsense, 10 missense and four splice-site mutations, 34 of which were novel. In five additional patients, novel, i.e. non-dbSNP132 variants of clinically unknown relevance, were identified. Patients with likely pathogenic nonsense or missense MLL2 mutations were usually more severely affected (median 'MLL2-Kabuki score' of 6) as compared to the patients without MLL2 mutations (median 'MLL2-Kabuki score' of 5), a significant difference (p < 0.0014). Several typical facial features such as large dysplastic ears, arched eyebrows with sparse lateral third, blue sclerae, a flat nasal tip with a broad nasal root, and a thin upper and a full lower lip were observed more often in mutation positive patients. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  5. 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

  6. Genetic diversity of Pinus halepensis Mill. populations detected by RAPD loci

    OpenAIRE

    Gómez , Aránzazu; Alía , Ricardo; Bueno , María

    2001-01-01

    International audience; Genetic diversity of Pinus halepensis Mill. was analysed in nine populations (six Spanish populations and one each from Tunisia, France and Greece). Twenty four RAPD loci were amplified with 60 megagametophyte DNA samples from each population. Populations' contribution to Nei gene diversity and to allelic richness were calculated. Results showed higher within population genetic variation but also a $G_{{\\rm ST}} = 13.6\\%$ higher than those detected in previous studies ...

  7. 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

  8. FIRST MOLECULAR DETECTION AND VP7 (G GENOTYPING OF GROUP A ROTAVIRUS BY SEMI-NESTED RT-PCR FROM SEWAGE IN NIGERIA

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    Babatunde Olanrewaju MOTAYO

    Full Text Available SUMMARY Rotavirus is the leading cause of viral gastroenteritis worldwide, and sewage is a major source of the virus dissemination in the environment. Our aim was to detect and genotype rotaviruses from sewages in Nigeria. One hundred and ninety sewage samples were collected between June 2014 and January 2015. The two phase concentration method using PEG 6000 and dextran was used to concentrate sewage samples following WHO protocols. Molecular detection was performed by RT-PCR, and VP7 genotyping by semi-nested multiplex PCR. A total of 14.2% (n = 27 samples tested positive. Monthly distribution showed that June to September had a lower rate (3.7% to 7.4%, while October to January recorded 11% to 26%. Genotype G1 predominated followed by G8, G9, G4 and lastly G2, 7.4% (n = 2 of isolates were nontypeable. This is the first report of rotavirus detection in sewages from Nigeria. Genotype G1 remains the most prevalent genotype. This observation calls for an effort by the governmental authorities to implement a molecular surveillance, both clinical and environmental, in order to provide vital information for the control and the vaccine efficacy not only in Nigeria, but globally.

  9. Detecting genotyping errors and describing black bear movement in northern Idaho

    Science.gov (United States)

    Michael K. Schwartz; Samuel A. Cushman; Kevin S. McKelvey; Jim Hayden; Cory Engkjer

    2006-01-01

    Non-invasive genetic sampling has become a favored tool to enumerate wildlife. Genetic errors, caused by poor quality samples, can lead to substantial biases in numerical estimates of individuals. We demonstrate how the computer program DROPOUT can detect amplification errors (false alleles and allelic dropout) in a black bear (Ursus americanus) dataset collected in...

  10. A new molecular diagnostic tool for quantitatively detecting and genotyping “Candidatus Liberibacter species”

    Science.gov (United States)

    A new molecular diagnostic method was developed for quantitative detection of “Candidatus Liberibacter” species associated with citrus Huanglongbing (“Ca. Liberibacter asiaticus”, “Ca. Liberibacter africanus” and “Ca. Liberibacter americanus”) and potato zebra chip disorder (“Ca. Liberibacter solana...

  11. Sensitive detection of multiple hepatitis A virus genotypes with a single polony-based assay

    Science.gov (United States)

    Hepatitis A virus (HAV) is one of the major causes of non-bacterial gastroenteritis in humans worldwide. HAV is mostly transmitted via direct person-to-person contact, or by consumption of contaminated foods and water. Since only a few viral particles may cause disease, detection of low levels of HA...

  12. Conceptual design of a system for detecting national diversion of LWR spent fuel

    International Nuclear Information System (INIS)

    Holmes, J.P.

    1978-09-01

    A conceptual design for detecting the national diversion of light water reactor spent fuel in water basin storage or in transit between facilities is described. This is the third in a series of reports dealing with this topic. The first report provides the spent fuel facilities and operations baseline description; the second report discusses cost and performance tradeoffs for three inspection and surveillance concepts for the detection of a national diversion of spent fuel. The conceptual design presented herein will provide a basis for future feasibility investigations and tradeoff analyses of hardware configurations and inspection options

  13. 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

  14. Searching for Celiac Disease Screening-detected celiac disease in an HLA-genotyped birth cohort

    OpenAIRE

    Björck, Sara

    2015-01-01

    Objectives: Celiac disease is a common immune mediated enteropathy strongly associated with HLA-DQB1*02 (DQ2), *0302 (DQ8), or both and the presence of tissue transglutaminase autoantibodies (tTGA). Prevalence studies have revealed that most affected individuals go undetected because of subclinical signs or being asymptomatic rendering screening a method for identification. However, less is known about subclinical manifestations of screening-detected celiac disease during childhood and if the...

  15. 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.

  16. M. tuberculosis genotypic diversity and drug susceptibility pattern in HIV- infected and non-HIV-infected patients in northern Tanzania

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    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.

  17. 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.

  18. Multiplex Detection and Genotyping of Point Mutations Involved in Charcot-Marie-Tooth Disease Using a Hairpin Microarray-Based Assay

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    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. 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.

  20. Detection of Aggregatibacter actinomycetemcomitans leukotoxin and fimbria-associated protein gene genotypes among periodontitis patients and healthy controls: A case–control study

    Science.gov (United States)

    Mahalakshmi, Krishnan; Krishnan, Padma; Chandrasekaran, S. C.

    2018-01-01

    Background: Aggregatibacter actinomycetemcomitans has been reported in higher proportions in subgingival microbiota of individuals with aggressive periodontitis (AgP) compared with those with chronic periodontitis (ChP) and healthy controls. The major virulence factors are the ones that help in colonization and evasion of host's defenses. Hence, this study was aimed to assess the prevalence of A. actinomycetemcomitans 16S rRNA and its virulent genotypes (leukotoxin [lktA] and fimbria-associated protein [fap]). Materials and Methods: In this case– control study We performed periodontal examination and measured probing depth and clinical attachment level (CAL). Subgingival plaque samples from 200 (ChP: n = 128 and AgP: n = 72) periodontitis patients and 200 healthy controls were screened for the presence of A. actinomycetemcomitans 16S rRNA, lktA, and fap genotypes by polymerase chain reaction. The prevalence of genotypes between periodontitis patients and healthy controls was compared with Pearson's Chi-square test. P periodontitis patients, while A. actinomycetemcomitans fap genotype showed 31.8% prevalence. The prevalence of these genotypes was insignificant in healthy controls. Conclusion: The high odds ratio for A. actinomycetemcomitans prevalence suggests its strong link to periodontitis. Detection of A. actinomycetemcomitans lktA + genotype may be a useful marker for AgP as its prevalence was found to be high in AgP. PMID:29922337

  1. Evaluation of the BACTEC MGIT 960 SL DST Kit and the GenoType MTBDRsl Test for Detecting Extensively Drug-resistant Tuberculosis Cases.

    Science.gov (United States)

    Tekin, Kemal; Albay, Ali; Simsek, Hulya; Sig, Ali Korhan; Guney, Mustafa

    2017-10-01

    The present study aimed to evaluate the performances of the BACTEC MGIT 960 SL DST kit and the GenoType MTBDRsl test for detecting second-line antituberculosis drug resistance in Multidrug-resistant TB (MDR-TB) cases. Forty-six MDR-TB strains were studied. Second-line antituberculosis drug resistances were detected using the BACTEC MGIT 960 SL DST kit and the GenoType MTBDRsl test. The Middlebrook 7H10 agar proportion method was used as the reference test. The sensitivity and specificity values for the BACTEC MGIT 960 SL DST kit were both 100% for amikacin, kanamycin, capreomycin (4 µg/mL), and ofloxacin; 100% and 95.3%, respectively, for capreomycin (10 µg/mL); and 85.7% and 100%, respectively, for moxifloxacin (0.5 µg/mL). The sensitivity and specificity values for the GenoType MTBDRsl test to detect fluoroquinolone and aminoglycoside/cyclic peptide resistance were 88.9% and 100%, respectively, for ofloxacin and 85.7% and 94.9%, respectively, for moxifloxacin (0.5 µg/mL). The accuracy of the GenoType MTBDRsl assay for kanamycin, capreomycin, ofloxacin, and moxifloxacin was lower than that of the BACTEC MGIT 960 SL DST. The BACTEC MGIT 960 SL DST kit and the GenoType MTBDRsl were successful in detecting second-line antituberculosis drug resistance. Preliminary results of the GenoType MTBDRsl are very valuable for early treatment decisions, but we still recommend additional BACTEC MGIT 960 SL DST kit usage in the routine evaluation of drug-resistant tuberculosis.

  2. Evaluation of the BACTEC MGIT 960 SL DST Kit and the GenoType MTBDRsl Test for Detecting Extensively Drug-resistant Tuberculosis Cases

    Science.gov (United States)

    Tekin, Kemal; Albay, Ali; Simsek, Hulya; Sig, Ali Korhan; Guney, Mustafa

    2017-01-01

    Objective: The present study aimed to evaluate the performances of the BACTEC MGIT 960 SL DST kit and the GenoType MTBDRsl test for detecting second-line antituberculosis drug resistance in Multidrug-resistant TB (MDR-TB) cases. Materials and Methods: Forty-six MDR-TB strains were studied. Second-line antituberculosis drug resistances were detected using the BACTEC MGIT 960 SL DST kit and the GenoType MTBDRsl test. The Middlebrook 7H10 agar proportion method was used as the reference test. Results: The sensitivity and specificity values for the BACTEC MGIT 960 SL DST kit were both 100% for amikacin, kanamycin, capreomycin (4 µg/mL), and ofloxacin; 100% and 95.3%, respectively, for capreomycin (10 µg/mL); and 85.7% and 100%, respectively, for moxifloxacin (0.5 µg/mL). The sensitivity and specificity values for the GenoType MTBDRsl test to detect fluoroquinolone and aminoglycoside/cyclic peptide resistance were 88.9% and 100%, respectively, for ofloxacin and 85.7% and 94.9%, respectively, for moxifloxacin (0.5 µg/mL). The accuracy of the GenoType MTBDRsl assay for kanamycin, capreomycin, ofloxacin, and moxifloxacin was lower than that of the BACTEC MGIT 960 SL DST. Conclusion: The BACTEC MGIT 960 SL DST kit and the GenoType MTBDRsl were successful in detecting second-line antituberculosis drug resistance. Preliminary results of the GenoType MTBDRsl are very valuable for early treatment decisions, but we still recommend additional BACTEC MGIT 960 SL DST kit usage in the routine evaluation of drug-resistant tuberculosis. PMID:29123441

  3. FTA liver impressions as DNA template for detecting and genotyping fowl adenovirus.

    Science.gov (United States)

    Moscoso, Hugo; Bruzual, Jose J; Sellers, Holly; Hofacre, Charles L

    2007-03-01

    The feasibility of using liver impressions on Flinders Technology Associates (FTA filter paper for the collection, inactivation, and molecular analysis of fowl adenovirus (FAV) was evaluated. FAV I European Union (EU) serotype 1 spotted on FTA was shown to be inactivated using specific-pathogen-free (SPF) primary chicken embryo liver cell culture as indicated by absence of cytopathic effect. Sensitivity of the polymerase chain reaction (PCR) test using tenfold dilutions of allantoic fluid from 100 to 10-4 for the detection of adenovirus serotype 1 on FTA cards was determined to be 0.0005 mean tissue culture infectious dose per FTA spot. The stability of the DNA from liver impressions on the FTA was found to be 198 days when stored at -20 degrees C. In a trial, inclusion body hepatitis (IBH) was experimentally reproduced in SPF chickens inoculated with FAV I EU serogroup 1, 4, 8, or 11, which presented weakness, pallor, depression, dehydration, and mortality within 6 days after inoculation. PCR performed on FTA liver impressions from the inoculated birds was able to detect all four viruses, and the nucleotide sequence analysis of the amplified PCR products (1219 bp of the hexone gene) revealed the expected serotypes. In addition to the trial, 55 clinical samples were analyzed from liver impressions on FTA cards, and FAV was detected in 11 of 55 (20%). Sequencing analysis showed that the viruses were EU serotypes 4, 5, 9, and 10. The results demonstrate that FTA filter paper inactivates the FAV I and maintains the DNA template for molecular analysis.

  4. Phenotypic and genotypic detection of methicillin-resistant Staphylococcus aureus in hunting dogs in Maiduguri metropolitan, Borno State, Nigeria.

    Science.gov (United States)

    Mustapha, Muhammad; Bukar-Kolo, Yachilla Maryam; Geidam, Yaqub Ahmed; Gulani, Isa Adamu

    2016-05-01

    To determine the presence of MRSA in hunting dogs in Maiduguri metropolitan. 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. 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. 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.

  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. Agreement for HPV genotyping detection between self-collected specimens on a FTA cartridge and clinician-collected specimens.

    Science.gov (United States)

    Guan, Yaoyao; Gravitt, Patti E; Howard, Roslyn; Eby, Yolanda J; Wang, Shaoming; Li, Belinda; Feng, Changyan; Qiao, You-Lin; Castle, Philip E

    2013-04-01

    The current method of transporting self-collected cervicovaginal specimen for HPV DNA testing relies on liquid based medium, which is challenging and expensive to transport. A novel, dry storage and transportation device, Whatman indicating FTA™ Elute Cartridge, avoids some of the pitfalls of liquid-based medium. This method has been shown to be comparable to liquid-based collection medium, but relative performance of self-collected (SC) and clinician-collected (CC) samples onto FTA cards has not been reported. The objective of this study is to compare the analytic performance of self- and clinician-collected samples onto FTA cartridges for the detection of carcinogenic HPV using Linear Array. There was a 91% agreement, 69% positive agreement, and kappa of 0.75 between the clinician-collected and self-collected specimens for detection of any carcinogenic HPV genotype. When the HPV results were categorized hierarchically according to cervical cancer risk, there was no difference in the distribution of the HPV results for the clinician- and self-collected specimens (p=0.7). This study concludes that FTA elute cartridge is a promising method of specimen transport for cervical cancer screening programs considering using self-collected specimen and HPV testing. Larger studies with clinical endpoints are now needed to assess the clinical performance. Copyright © 2012 Elsevier B.V. All rights reserved.

  7. Agreement for HPV genotyping detection between self-collected specimens on a FTA cartridge and clinician-collected specimens

    Science.gov (United States)

    Guan, YaoYao; Gravitt, Patti E.; Howard, Roslyn; Eby, Yolanda J.; Wang, Shaoming; Li, Belinda; Feng, Changyan; Qiao, You-Lin; Castle, Philip E.

    2016-01-01

    The current method of transporting self-collected cervicovaginal specimen for HPV DNA testing relies on liquid based medium, which is challenging and expensive to transport. A novel, dry storage and transportation device, Whatman indicating FTA™ Elute Cartridge, avoids some of the pitfalls of liquid-based medium. This method has been shown to be comparable to liquid-based collection medium, but relative performance of self-collected (SC) and clinician-collected (CC) samples onto FTA cards has not been reported. The objective of this study is to compare the analytic performance of self- and clinician-collected samples onto FTA cartridges for the detection of carcinogenic HPV using Linear Array. There was a 91% agreement, 69% positive agreement, and kappa of 0.75 between the clinician-collected and self-collected specimens for detection of any carcinogenic HPV genotype. When the HPV results were categorized hierarchically according to cervical cancer risk, there was no difference in the distribution of the HPV results for the clinician- and self-collected specimens (p = 0.7). This study concludes that FTA elute cartridge is a promising method of specimen transport for cervical cancer screening programs considering using self-collected specimen and HPV testing. Larger studies with clinical endpoints are now needed to assess the clinical performance. PMID:23370404

  8. 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

    . 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......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...

  9. 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.

  10. Application of DNA-DNA colony hybridization to the detection of catabolic genotypes in environmental samples

    International Nuclear Information System (INIS)

    Sayler, G.S.; Shields, M.S.; Tedford, E.T.; Breen, A.; Hooper, S.W.; Sirotkin, K.M.; Davis, J.W.

    1985-01-01

    The application of preexisting DNA hybridization techniques was investigated for potential in determining populations of specific gene sequences in environmental samples. Cross-hybridizations among two degradative plasmids, TOL and NAH, and two cloning vehicles, pLAFR1 and RSF1010, were determined. The detection limits for the TOL plasmid against a nonhomologous plasmid-bearing bacterial background was ascertained. The colony hybridization technique allowed detection of one colony containing TOL plasmid among 10(6) Escherichia coli colonies of nonhomologous DNA. Comparisons between population estimates derived from growth on selective substrates and from hybridizations were examined. Findings indicated that standard sole carbon source enumeration procedures for degradative populations lead to overestimations due to nonspecific growth of other bacteria on the microcontaminant carbon sources present in the media. Population estimates based on the selective growth of a microcosm population on two aromatic substrates (toluene and naphthalene) and estimates derived from DNA-DNA colony hybridizations, using the TOL or NAH plasmid as a probe, corresponded with estimates of substrate mineralization rates and past exposure to environmental contaminants. The applications of such techniques are hoped to eventually allow enumeration of any specific gene sequences in the environment, including both anabolic and catabolic genes. In addition, this procedure should prove useful in monitoring recombinant DNA clones released into environmental situations

  11. Role of static fluid MR urography in detecting post urinary diversion complications

    Directory of Open Access Journals (Sweden)

    Amr Farouk Ibrahim Moustafa

    2018-06-01

    Full Text Available Aim of work: The aim of the study was to assess the diagnostic performance of static MR urography in detection of post cystectomy complications & the ability of static fluid MR urography in visualization of urinary tract segments. Material & methods: We prospectively reviewed 21 MR urograms with urinary diversion. The most common surgical procedures included Ileal conduit & Ileocecal neobladder diversion.Magnetic resonance urography examinations were performed with 1.5-T MR scanners. T2 weighted (static fluid MR urography techniques were done, in addition to conventional T1- and T2-weighted axial and coronal sequences. Urinary tract was divided in different parts: pelvicalyceal systems, upper, mid and lower ureteric segments & the reservoir or conduit Imaging features of the urinary collecting systems were evaluated for their visualization and complications detection. Results: T2-weighted MR urography could demonstrate 95% of urinary tract segments & together with conventional MR sequences all urinary tract segments can be visualized. Urinary diversion related complications were encountered included in 15 patients (71.4% & no urological complications were seen in 6 patients (28.6%. Conclusion: Comprehensive T2-weighted MR urography is an effective imaging method for the visualization of the urinary system and detection of early and late postoperative complications in patients with urinary diversion. Keywords: MR urography, Urinary diversion, Cancer bladder

  12. 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.

  13. Label-free genotyping of cytochrome P450 2D6*10 using ligation-mediated strand displacement amplification with DNAzyme-based chemiluminescence detection.

    Science.gov (United States)

    Wang, Hong-Qi; Wu, Zhan; Zhang, Yan; Tang, Li-Juan; Yu, Ru-Qin; Jiang, Jian-Hui

    2012-01-13

    Genotyping of cytochrome P450 monooxygenase 2D6*10 (CYP2D6*10) plays an important role in pharmacogenomics, especially in clinical drug therapy of Asian populations. This work reported a novel label-free technique for genotyping of CYP2D6*10 based on ligation-mediated strand displacement amplification (SDA) with DNAzyme-based chemiluminescence detection. Discrimination of single-base mismatch is firstly accomplished using DNA ligase to generate a ligation product. The ligated product then initiates a SDA reaction to produce aptamer sequences against hemin, which can be probed by chemiluminescence detection. The proposed strategy is used for the assay of CYP2D6*10 target and the genomic DNA. The results reveal that the proposed technique displays chemiluminescence responses in linear correlation to the concentrations of DNA target within the range from 1 pM to 1 nM. A detection limit of 0.1 pM and a signal-to-background ratio of 57 are achieved. Besides such high sensitivity, the proposed CYP2D6*10 genotyping strategy also offers superb selectivity, great robustness, low cost and simplified operations due to its label-free, homogeneous, and chemiluminescence-based detection format. These advantages suggest this technique may hold considerable potential for clinical CYP2D6*10 genotyping and association studies. Copyright © 2011 Elsevier B.V. All rights reserved.

  14. Microfluidic PCR Amplification and MiSeq Amplicon Sequencing Techniques for High-Throughput Detection and Genotyping of Human Pathogenic RNA Viruses in Human Feces, Sewage, and Oysters

    Directory of Open Access Journals (Sweden)

    Mamoru Oshiki

    2018-04-01

    Full Text Available Detection and genotyping of pathogenic RNA viruses in human and environmental samples are useful for monitoring the circulation and prevalence of these pathogens, whereas a conventional PCR assay followed by Sanger sequencing is time-consuming and laborious. The present study aimed to develop a high-throughput detection-and-genotyping tool for 11 human RNA viruses [Aichi virus; astrovirus; enterovirus; norovirus genogroup I (GI, GII, and GIV; hepatitis A virus; hepatitis E virus; rotavirus; sapovirus; and human parechovirus] using a microfluidic device and next-generation sequencer. Microfluidic nested PCR was carried out on a 48.48 Access Array chip, and the amplicons were recovered and used for MiSeq sequencing (Illumina, Tokyo, Japan; genotyping was conducted by homology searching and phylogenetic analysis of the obtained sequence reads. The detection limit of the 11 tested viruses ranged from 100 to 103 copies/μL in cDNA sample, corresponding to 101–104 copies/mL-sewage, 105–108 copies/g-human feces, and 102–105 copies/g-digestive tissues of oyster. The developed assay was successfully applied for simultaneous detection and genotyping of RNA viruses to samples of human feces, sewage, and artificially contaminated oysters. Microfluidic nested PCR followed by MiSeq sequencing enables efficient tracking of the fate of multiple RNA viruses in various environments, which is essential for a better understanding of the circulation of human pathogenic RNA viruses in the human population.

  15. 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

  16. Detection of Genotype 4 Swine Hepatitis E Virus in Systemic Tissues in Cross-Species Infected Rabbits

    OpenAIRE

    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...

  17. 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

    intraepithelial neoplasia (CIN) in 2.5 years after the baseline testing were determined from the national pathology register. HPV-positive women undergoing primary screening having concordant samples were more likely to harbor high-risk infections and less likely to harbor only low-risk infections than women......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...

  18. 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

  19. Detection of enterotoxins and genotyping of Staphylococcus aureus strains isolated from Isfahan Educational Hospital, Iran

    Directory of Open Access Journals (Sweden)

    Seyed Asghar Havaei

    2017-10-01

    Full Text Available Background and aims: Staphylococcus aureus is known as one of the most important nosocomial pathogens, which may lead to several infections. The aim of this study was determining the enterotoxins A, C, and TSST-1 and molecular characterization of S. aureus strains with PFGE and MLST typing methods. Materials and methods: In the present study during the sixmonths sampling, fifty S. aureus strains were isolated from patients admitted to Al-Zahra university hospital. Antimicrobial susceptibility testing, Multiplex PCR for detection of enterotoxin A, C and TSST-1, pulse field gel electrophoresis (PFGE and multilocus sequence typing (MLST were used for molecular typing. Results: In antibiogram the highest and lowest percentage of resistance was belonged to tetracycline and rifampin respectively. Multiplex PCR indicated that 30% of the strains harbored sea and 34% harbored sec genes. However, only 4% of our collected isolates had tsst gene. In PFGE method analysis on all S. aureus strains, a total of 19 different patterns were identified. Nine various sequence types in 27 selected S. aureus isolates were identified by MLST. Conclusions: Present study indicates a possible higher variability among our S. aureus strains by two different molecular typing methods; nevertheless four main common types (CT1, CT7, CT9, and CT11 with at least one toxin genes were determined.

  20. One-step simultaneous detection of Ureaplasma parvum and genotypes SV1, SV3 and SV6 from clinical samples using PlexPCR technology.

    Science.gov (United States)

    Payne, M S; Furfaro, L L; Tucker, R; Tan, L Y; Mokany, E

    2017-08-01

    Ureaplasma spp. are associated with preterm birth. In recent times, it has become apparent that Ureaplasma parvum, but not Ureaplasma urealyticum, is of most relevance. We recently demonstrated this in Australian pregnant women and using high-resolution melt (HRM) PCR, further showed that U. parvum genotype SV6 was of particular significance. However, our assay was unable to identify multiple genotypes in the same sample, required a separate species-level qPCR for low titre samples and was not ideal for diagnostic laboratories due to the nature of HRM PCR result interpretation. Consequently, our current study developed a novel, one-step PlexPCR assay capable of detecting U. parvum and genotypes SV1, SV3 and SV6 in a single reaction directly from clinical samples. We then validated this using vaginal swab DNA from our Australian cohort of pregnant women. The PlexPCR was highly sensitive, detecting all targets to between 0.4 × 10 -5  ng DNA (SV3) and 0.4 × 10 -6  ng DNA (U. parvum, SV1 and SV6). Compared to our HRM PCR, the PlexPCR defined genotype distribution in all seven cases previously reported as 'mixed', and detected another eight cases where multiple genotypes (two) were present in samples previously reported as single genotypes using HRM PCR. Ureaplasma spp. have been associated with prematurity for decades, however, only a minority of studies have examined this beyond the genus level. In those that have, Ureaplasma parvum has been strongly associated with preterm birth. We recently demonstrated this in Australian women and further showed that U. parvum genotype SV6 was of particular significance. Our PlexPCR assay allows rapid detection and concurrent genotyping of U. parvum in clinical samples and may be of particular interest to obstetricians, particularly those caring for women at a high risk of preterm birth, and any other disease phenotypes where U. parvum is of interest. © 2017 The Authors. Letters in Applied Microbiology published by John

  1. 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.

  2. Detection of a diverse marine fish fauna using environmental DNA from seawater samples

    DEFF Research Database (Denmark)

    Thomsen, Philip Francis; Kielgast, Jos; Iversen, Lars Lønsmann

    2012-01-01

    eDNA from 15 different fish species, including both important consumption species, as well as species rarely or never recorded by conventional monitoring. We also detect eDNA from a rare vagrant species in the area; European pilchard (Sardina pilchardus). Additionally, we detect four bird species....... Records in national databases confirmed the occurrence of all detected species. To investigate the efficiency of the eDNA approach, we compared its performance with 9 methods conventionally used in marine fish surveys. Promisingly, eDNA covered the fish diversity better than or equal to any of the applied...

  3. 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.

  4. High genotypic diversity of the reef-building coral Porites lobata (Scleractinia: Poritidae in Isla del Coco National Park, Costa Rica

    Directory of Open Access Journals (Sweden)

    Jennifer N. Boulay

    2012-11-01

    Full Text Available The isolated Isla del Coco experiences periodic, extreme disturbances which devastate coral reefs surrounding the island. Scleractinian corals build the physical structure of the reef therefore ecosystem recovery relies on coral species recovery. Coral recruits can be of sexual or asexual origin, and the relative success of the two recruit types influences the speed and spread of recovery processes. Here we focus on the massive coral, Porites lobata, because it is the main reef-builder around Isla del Coco to describe the relative contribution of asexual and sexual recruits to population maintenance. P. lobata samples were collected using a spatially explicit random sampling design in three plots at Isla del Coco: Punta Ulloa (n=17, Bahía Weston (n=20 and Punta María (n=20 and samples were genotyped with 11 microsatellite markers. Additional sampling was conducted at three “coastal” sites near the Costa Rican mainland (Isla del Caño Biological Reserve: Caño1 (n=8, Caño2 (n=10, Caño5 (n=11 to compare the contributions of asexual and sexual recruits at Isla del Coco sites to coastal sites. Isla del Coco sites were characterized by small colony size (>60% of colonies <0.5m2 and high sexual reproduction. Sites were either mostly or entirely sexual,consisting of only unique genotypes (N G/N= 0.90-1.00; G O/G E=0.83-1.00; D=0.99-1.00. Although there were no significant differences in genetic diversity (number of alleles per locus, number of private alleles or colony size between Isla del Coco and the coastal sites, the coastal sites exhibited a greater range of genotypic diversity from moderately asexual (N G/N=0.5; G O/G E=0.36; D=0.8 to purely sexual (N G/N=1.0; G O/G E=1.0; D=1.0. The mode of asexual reproduction in P. lobata is likely fragmentation of adult colonies rather than asexual larval production because ramets of P. lobata occurred close together and asexually produced larvae have not been reported in gonochoric broadcast

  5. Detection and gB genotyping of CMV in Mexican preterm infants in the context of maternal seropositivity.

    Science.gov (United States)

    Arellano-Galindo, José; Villanueva-García, Dina; Cruz-Ramirez, José Luis; Yalaupari-Mejìa, Juan Pablo; Uribe-Gutiérrez, Gabriel; Velazquez-Guadarrama, Norma; Nava-Frias, Margarita; Munoz-Hernández, Onofre; Mejía-Arangure, Juan Manuel

    2014-06-11

    Congenital (CI) and perinatal cytomegalovirus (CMV) infections (PI) can be linked to maternal CMV seropositivity, with fatal consequences in preterm newborns. GB genotyping has been used to analyze genotypic similarity in mothers and infants. The frequency of CMV infection in the context of maternal seropositivity and the viral gB genotypes as well as the genotypic similarity in mothers and preterm infants were investigated. Saliva samples and dry blood spots (DBS) were taken weekly from preterm newborns  from birth until the first month of life, and breast milk samples were taken from their mothers weekly during the first month of lactation. CMV IgG seroprevalence of the mothers and CI or PI in the infants were established. The gB status and genotypic similarities were established retrospectively in DBS and in the breast milk samples. In total, 387 neonates and 375 mothers were enrolled. The maternal CMV-positive IgG serology was 97.3% (365/375). Neonatal CMV was found in 5.1% (20/387) of newborns, and one infant presented with CMV-compatible symptoms. CI was 2.5% and PI in the first month after birth was 11.8%. GB2 was the most prevalent genotype and was also the genotype preferentially transmitted to newborns by mothers with mixed infections. CMV PI and CI in preterm infants from highly seropositive mothers was high, but the rate of symptomatic infection was low. The prevalent genotype was gB2, and this genotype was preferentially transmitted to newborns by mothers with mixed infections.

  6. A high resolution melting (HRM) technology-based assay for cost-efficient clinical detection and genotyping of herpes simplex virus (HSV)-1 and HSV-2.

    Science.gov (United States)

    Lieveld, M; Carregosa, A; Benoy, I; Redzic, N; Berth, M; Vanden Broeck, D

    2017-10-01

    Genital herpes can be caused by two very similar viruses, herpes simplex virus (HSV)-1 or HSV-2. These two HSV types cannot be distinguished clinically, but genotyping is recommended in the first-episodes of genital herpes to guide counselling and management. Quantitative polymerase chain reaction (qPCR) is the preferred diagnostic method for HSV typing. However, commercial qPCR methods use expensive fluorescent labeled probes for detection. Furthermore, most low-cost methods are not able to differentiate between HSV-1 and -2. The aim of this study was to develop a high resolution melting (HRM) technology-based assay for sensitive HSV-1 and HSV-2 detection and genotyping. Using a panel of 46 clinical specimens, the performance of the HRM assay was compared to two commercial HSV tests: the HRM assay detected HSV in all 23 positive samples, with no false positive results (100% concordance with HSV I/II Real-TM assay). Additionally, the HRM assay correctly genotyped both HSV types in a subset of these clinical samples, as determined by the Realstar HSV PCR Kit. The HSV HRM assay provides a cost-effective alternative method to conventional more expensive assays and can be used in routine clinical specimens, in cases where it is particularly necessary to detect and distinguish HSV-1 from -2. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Diversity in the Enteric Viruses Detected in Outbreaks of Gastroenteritis from Mumbai, Western India

    Directory of Open Access Journals (Sweden)

    Renu Bhardwaj

    2012-03-01

    Full Text Available Faecal specimens collected from two outbreaks of acute gastroenteritis that occurred in southern Mumbai, India in March and October, 2006 were tested for seven different enteric viruses. Among the 218 specimens tested, 95 (43.6% were positive, 73 (76.8% for a single virus and 22 (23.2% for multiple viruses. Single viral infections in both, March and October showed predominance of enterovirus (EV, 33.3% and 40% and rotavirus A (RVA, 33.3% and 25%. The other viruses detected in these months were norovirus (NoV, 12.1% and 10%, rotavirus B (RVB, 12.1% and 10%, enteric adenovirus (AdV, 6.1% and 7.5%, Aichivirus (AiV, 3% and 7.5% and human astrovirus (HAstV, 3% and 0%. Mixed viral infections were largely represented by two viruses (84.6% and 88.9%, a small proportion showed presence of three (7.7% and 11% and four (7.7% and 0% viruses in the two outbreaks. Genotyping of the viruses revealed predominance of RVA G2P[4], RVB G2 (Indian Bangladeshi lineage, NoV GII.4, AdV-40, HAstV-8 and AiV B types. VP1/2A junction region based genotyping showed presence of 11 different serotypes of EVs. Although no virus was detected in the tested water samples, examination of both water and sewage pipelines in gastroenteritis affected localities indicated leakages and possibility of contamination of drinking water with sewage water. Coexistence of multiple enteric viruses during the two outbreaks of gastroenteritis emphasizes the need to expand such investigations to other parts of India.

  8. Diversity in the enteric viruses detected in outbreaks of gastroenteritis from Mumbai, Western India.

    Science.gov (United States)

    Chitambar, Shobha; Gopalkrishna, Varanasi; Chhabra, Preeti; Patil, Pooja; Verma, Harsha; Lahon, Anismrita; Arora, Ritu; Tatte, Vaishali; Ranshing, Sujata; Dhale, Ganesh; Kolhapure, Rajendra; Tikute, Sanjay; Kulkarni, Jagannath; Bhardwaj, Renu; Akarte, Sulbha; Pawar, Sashikant

    2012-03-01

    Faecal specimens collected from two outbreaks of acute gastroenteritis that occurred in southern Mumbai, India in March and October, 2006 were tested for seven different enteric viruses. Among the 218 specimens tested, 95 (43.6%) were positive, 73 (76.8%) for a single virus and 22 (23.2%) for multiple viruses. Single viral infections in both, March and October showed predominance of enterovirus (EV, 33.3% and 40%) and rotavirus A (RVA, 33.3% and 25%). The other viruses detected in these months were norovirus (NoV, 12.1% and 10%), rotavirus B (RVB, 12.1% and 10%), enteric adenovirus (AdV, 6.1% and 7.5%), Aichivirus (AiV, 3% and 7.5%) and human astrovirus (HAstV, 3% and 0%). Mixed viral infections were largely represented by two viruses (84.6% and 88.9%), a small proportion showed presence of three (7.7% and 11%) and four (7.7% and 0%) viruses in the two outbreaks. Genotyping of the viruses revealed predominance of RVA G2P[4], RVB G2 (Indian Bangladeshi lineage), NoV GII.4, AdV-40, HAstV-8 and AiV B types. VP1/2A junction region based genotyping showed presence of 11 different serotypes of EVs. Although no virus was detected in the tested water samples, examination of both water and sewage pipelines in gastroenteritis affected localities indicated leakages and possibility of contamination of drinking water with sewage water. Coexistence of multiple enteric viruses during the two outbreaks of gastroenteritis emphasizes the need to expand such investigations to other parts of India.

  9. Detection of Diverse Novel Bat Astrovirus Sequences in the Czech Republic.

    Science.gov (United States)

    Dufkova, Lucie; Straková, Petra; Širmarová, Jana; Salát, Jiří; Moutelíková, Romana; Chrudimský, Tomáš; Bartonička, Tomáš; Nowotny, Norbert; Růžek, Daniel

    2015-08-01

    Astroviruses are a major cause of gastroenteritis in humans and animals. Recently, novel groups of astroviruses were identified in apparently healthy insectivorous bats. We report the detection of diverse novel astrovirus sequences in nine different European bat species: Eptesicus serotinus, Hypsugo savii, Myotis emarginatus, M. mystacinus, Nyctalus noctula, Pipistrellus nathusii or P. pygmaeus, P. pipistrellus, Vespertilio murinus, and Rhinolophus hipposideros. In six bat species, astrovirus sequences were detected for the first time. One astrovirus strain detected in R. hipposideros clustered phylogenetically with Chinese astrovirus strains originating from bats of the families Rhinolophidae and Hipposideridae. All other Czech astrovirus sequences from vesper bats formed, together with one Hungarian sequence, a separate monophyletic lineage within the bat astrovirus group. These findings provide new insights into the molecular epidemiology, ecology, and prevalence of astroviruses in European bat populations.

  10. Comparison of Detection Rate and Mutational Pattern of Drug-Resistant Mutations Between a Large Cohort of Genotype B and Genotype C Hepatitis B Virus-Infected Patients in North China.

    Science.gov (United States)

    Li, Xiaodong; Liu, Yan; Xin, Shaojie; Ji, Dong; You, Shaoli; Hu, Jinhua; Zhao, Jun; Wu, Jingjing; Liao, Hao; Zhang, Xin-Xin; Xu, Dongping

    2017-06-01

    The study aimed to investigate the association of prevalent genotypes in China (HBV/C and HBV/B) with HBV drug-resistant mutations. A total of 13,847 nucleos(t)ide analogue (NA)-treated patients with chronic HBV infection from North China were enrolled. HBV genotypes and resistant mutations were determined by direct sequencing and confirmed by clonal sequencing if necessary. HBV/B, HBV/C, and HBV/D occupied 14.3%, 84.9%, and 0.8% across the study population, respectively. NA usage had no significant difference between HBV/B- and HBV/C-infected patients. Lamivudine-resistant mutations were more frequently detected in HBV/C-infected patients, compared with HBV/B-infected patients (31.67% vs. 25.26%, p M250 V/I/L substitution (0.67% vs. 1.46%, p < 0.01). Multidrug-resistant mutations (defined as coexistence of mutation to nucleoside and nucleotide analogues) were detected in 104 patients. HBV/C-infected patients had a higher detection rate of multidrug-resistant mutation than HBV/B-infected patients (0.83% vs. 0.35%, p < 0.05). The study for the first time clarified that HBV/C-infected patients had a higher risk to develop multidrug-resistant mutations, compared with HBV/B-infected patients; and HBV/C- and HBV/B-infected patients had different inclinations in the ETV-resistant mutational pattern.

  11. 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 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.To assess baseline predictors of failure of sputum culture conversion, within the first 2 months of antitubercular therapy, in participants with pulmonary tuberculosis.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.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.We found that baseline TTD, smoking, cavities and W-Beijing genotype were associated with delayed 2 month sputum culture

  12. 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

  13. Baseline predictors of sputum culture conversion in pulmonary tuberculosis: importance of cavities, smoking, time to detection and W-Beijing genotype.

    Science.gov (United States)

    Visser, Marianne E; Stead, Michael C; Walzl, Gerhard; Warren, Rob; Schomaker, Michael; Grewal, Harleen M S; Swart, Elizabeth C; Maartens, Gary

    2012-01-01

    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. To assess baseline predictors of failure of sputum culture conversion, within the first 2 months of antitubercular therapy, in participants with pulmonary tuberculosis. 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. 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. We found that baseline TTD, smoking, cavities and W-Beijing genotype were associated with delayed 2 month sputum culture. Larger

  14. 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.

  15. Lack of detection of host associated differences in Newcastle disease viruses of genotype VIId isolated from chickens and geese

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    Wang Yuyang

    2012-09-01

    Full Text Available Abstract Background The goose is usually considered to be resistant even to strains of Newcastle disease virus (NDV that are markedly virulent for chickens. However, ND outbreaks have been frequently reported in goose flocks in China since the late 1990s with the concurrent emergence of genotype VIId NDV in chickens. Although the NDVs isolated from both chickens and geese in the past 15 years have been predominantly VIId viruses, published data comparing goose- and chicken-originated ND viruses are scarce and controversial. Results In this paper, we compared genotype VIId NDVs originated from geese and chickens genetically and pathologically. Ten entire genomic sequences and 329 complete coding sequences of individual genes from genotype VIId NDVs of both goose- and chicken-origin were analyzed. We then randomly selected two goose-originated and two chicken-originated VIId NDVs and compared their pathobiology in both geese and chickens in vivo and in vitro with genotype IV virus Herts/33 as a reference. The results showed that all the VIId NDVs either from geese or from chickens shared high sequence homology and characteristic amino acid substitutions and clustered together in phylogenetic trees. In addition, geese and chickens infected by goose or chicken VIId viruses manifested very similar pathological features distinct from those of birds infected with Herts/33. Conclusions There is no genetic or phenotypic difference between genotype VIId NDVs originated from geese and chickens. Therefore, no species-preference exists for either goose or chicken viruses and more attention should be paid to the trans-species transmission of VIId NDVs between geese and chickens for the control and eradication of ND.

  16. 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 their density is expected to reach intervals of a few centimorgans per marker. These new genetic maps open new avenues for disease gene research, including large-scale genotyping for both simple and complex disease loci. However, the allele patterns of many dinucleotide repeat loci can be complex and difficult to interpret, with genotyping errors a recognized problem. Furthermore, the possibility of genotyping individuals at hundreds or thousands of polymorphic loci requires improvements in data handling and analysis. The automation of genotyping and analysis of computer-derived haplotypes would remove many of the barriers preventing optimal use of dense and informative dinucleotide genetic maps. Toward this end, we have automated the allele identification, genotyping, phase determinations, and inheritance consistency checks generated by four CA repeats within the 2.5-Mbp, 10-cM X-linked dystrophin gene, using fluorescein-labeled multiplexed PCR products analyzed on automated sequencers. The described algorithms can deconvolute and resolve closely spaced alleles, despite interfering stutter noise; set phase in females; propagate the phase through the family; and identify recombination events. We show the implementation of these algorithms for the completely automated interpretation of allele data and risk assessment for five Duchenne/Becker muscular dystrophy families. The described approach can be scaled up to perform genome-based analyses with hundreds or thousands of CA-repeat loci, using multiple fluorophors on automated sequencers. 16 refs., 5 figs., 1 tab.

  17. 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.

  18. Evaluation of Diversity Based on Morphological Variabilities and ISSR Molecular Markers in Iranian Cynodon dactylon (L.) Pers. Accessions to Select and Introduce Cold-Tolerant Genotypes.

    Science.gov (United States)

    Akbari, M; Salehi, H; Niazi, A

    2018-04-01

    The main goals of the present study were to screen Iranian common bermudagrasses to find cold-tolerant accessions and evaluate their genetic and morphological variabilities. In this study, 49 accessions were collected from 18 provinces of Iran. One foreign cultivar of common bermudagrass was used as control. Morphological variation was evaluated based on 14 morphological traits to give information about taxonomic position of Iranian common bermudagrass. Data from morphological traits were evaluated to categorize all accessions as either cold sensitive or tolerant using hierarchical clustering with Ward's method in SPSS software. Inter-Simple Sequence Repeat (ISSR) primers were employed to evaluate genetic variability of accessions. The results of our taxonomic investigation support the existence of two varieties of Cynodon dactylon in Iran: var. dactylon (hairless plant) and var. villosous (plant with hairs at leaf underside and/or upper side surfaces or exterior surfaces of sheath). All 15 primers amplified and gave clear and highly reproducible DNA fragments. In total, 152 fragments were produced, of which 144 (94.73%) being polymorphic. The polymorphic information content (PIC) values ranged from 0.700 to 0.928. The average PIC value obtained with 15 ISSR primers was 0.800, which shows that all primers were informative. Probability identity (PI) and discriminating power between all primers ranged from 0.029 to 0.185 and 0.815 to 0.971, respectively. Genetic data were converted into a binary data matrix. NTSYS software was used for data analysis. Clustering was done by the unweighted pair-group method with arithmetic averages and principle coordinate analysis, separated the accessions into six main clusters. According to both morphological and genetic diversity investigations of accessions, they can be clustered into three groups: cold sensitive, cold semi-tolerant, and cold tolerant. The most cold-tolerant accessions were: Taft, Malayear, Gorgan, Safashahr

  19. Sensitivity of the ViroSeq HIV-1 Genotyping System for Detection of the K103N Resistance Mutation in HIV-1 Subtypes A, C, and D

    Science.gov (United States)

    Church, Jessica D.; Jones, Dana; Flys, Tamara; Hoover, Donald; Marlowe, Natalia; Chen, Shu; Shi, Chanjuan; Eshleman, James R.; Guay, Laura A.; Jackson, J. Brooks; Kumwenda, Newton; Taha, Taha E.; Eshleman, Susan H.

    2006-01-01

    The US Food and Drug Administration-cleared ViroSeq HIV-1 Genotyping System (ViroSeq) and other population sequencing-based human immunodeficiency virus type 1 (HIV-1) genotyping methods detect antiretroviral drug resistance mutations present in the major viral population of a test sample. These assays also detect some mutations in viral variants that are present as mixtures. We compared detection of the K103N nevirapine resistance mutation using ViroSeq and a sensitive, quantitative point mutation assay, LigAmp. The LigAmp assay measured the percentage of K103N-containing variants in the viral population (percentage of K103N). We analyzed 305 samples with HIV-1 subtypes A, C, and D collected from African women after nevirapine administration. ViroSeq detected K103N in 100% of samples with >20% K103N, 77.8% of samples with 10 to 20% K103N, 71.4% of samples with 5 to 10% K103N, and 16.9% of samples with 1 to 5% K103N. The sensitivity of ViroSeq for detection of K103N was similar for subtypes A, C, and D. These data indicate that the ViroSeq system reliably detects the K103N mutation at levels above 20% and frequently detects the mutation at lower levels. Further studies are needed to compare the sensitivity of different assays for detection of HIV-1 drug resistance mutations and to determine the clinical relevance of HIV-1 minority variants. PMID:16931582

  20. 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.

  1. 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.

  2. [Genetic diversity of wild Cynodon dactylon germplasm detected by SRAP markers].

    Science.gov (United States)

    Yi, Yang-Jie; Zhang, Xin-Quan; Huang, Lin-Kai; Ling, Yao; Ma, Xiao; Liu, Wei

    2008-01-01

    Sequence-related amplified polymorphism (SRAP) molecular markers were used to detect the genetic diversity of 32 wild accessions of Cynodon dactylon collected from Sichuan, Chongqing, Guizhou and Tibet, China. The following results were obtained. (1) Fourteen primer pairs produced 132 polymorphic bands, averaged 9.4 bands per primer pair. The percentage of polymorphic bands in average was 79.8%. The Nei's genetic similarity coefficient of the tested accessions ranged from 0.591 to 0.957, and the average Nei's coefficient was 0.759. These results suggested that there was rich genetic diversity among the wild resources of Cynodon dactylon tested. (2) Thirty two wild accessions were clustered into four groups. Moreover, the accessions from the same origin frequently clustered into one group. The findings implied that a correlation among the wild resources, geographical and ecological environment. (3) Genetic differentiation between and within six eco-geographical groups of C. dactylon was estimated by Shannon's diversity index, which showed that 65.56% genetic variance existed within group, and 34.44% genetic variance was among groups. (4) Based on Nei's unbiased measures of genetic identity, UPGMA cluster analysis measures of six eco-geographical groups of Cynodon dactylon, indicated that there was a correlation between genetic differentiation and eco-geographical habits among the groups.

  3. Diversity arrays technology (DArT) markers in apple for genetic linkage maps

    NARCIS (Netherlands)

    Schouten, H.J.; Weg, van de W.E.; Carling, J.; Khan, S.A.; McKay, S.J.; Kaauwen, van M.P.W.

    2012-01-01

    Diversity Arrays Technology (DArT) provides a high-throughput whole-genome genotyping platform for the detection and scoring of hundreds of polymorphic loci without any need for prior sequence information. The work presented here details the development and performance of a DArT genotyping array for

  4. An analysis of genetic diversity and inbreeding in Wuchereria bancrofti: implications for the spread and detection of drug resistance.

    Directory of Open Access Journals (Sweden)

    Thomas S Churcher

    2008-04-01

    Full Text Available Estimates of genetic diversity in helminth infections of humans often have to rely on genotyping (immature parasite transmission stages instead of adult worms. Here we analyse the results of one such study investigating a single polymorphic locus (a change at position 200 of the beta-tubulin gene in microfilariae of the lymphatic filarial parasite Wuchereria bancrofti. The presence of this genetic change has been implicated in benzimidazole resistance in parasitic nematodes of farmed ruminants. Microfilariae were obtained from patients of three West African villages, two of which were sampled prior to the introduction of mass drug administration. An individual-based stochastic model was developed showing that a wide range of allele frequencies in the adult worm populations could have generated the observed microfilarial genetic diversity. This suggests that appropriate theoretical null models are required in order to interpret studies that genotype transmission stages. Wright's hierarchical F-statistic was used to investigate the population structure in W. bancrofti microfilariae and showed significant deficiency of heterozygotes compared to the Hardy-Weinberg equilibrium; this may be partially caused by a high degree of parasite genetic differentiation between hosts. Studies seeking to quantify accurately the genetic diversity of helminth populations by analysing transmission stages should increase their sample size to account for the variability in allele frequency between different parasite life-stages. Helminth genetic differentiation between hosts and non-random mating will also increase the number of hosts (and the number of samples per host that need to be genotyped, and could enhance the rate of spread of anthelmintic resistance.

  5. Characteristic cardiac phenotypes are detected by cardiovascular magnetic resonance in patients with different clinical phenotypes and genotypes of mitochondrial myopathy.

    Science.gov (United States)

    Florian, Anca; Ludwig, Anna; Stubbe-Dräger, Bianca; Boentert, Matthias; Young, Peter; Waltenberger, Johannes; Rösch, Sabine; Sechtem, Udo; Yilmaz, Ali

    2015-05-22

    Mitochondrial myopathies (MM) are a heterogeneous group of inherited conditions resulting from a primary defect in the mitochondrial respiratory chain with consecutively impaired cellular energy metabolism. Small sized studies using mainly electrocardiography (ECG) and echocardiography have revealed cardiac abnormalities ranging from conduction abnormalities and arrhythmias to hypertrophic or dilated cardiomyopathy in these patients. Recently, characteristic patterns of cardiac involvement were documented by cardiovascular magnetic resonance (CMR) in patients with chronic progressive external ophthalmoplegia (CPEO)/Kearns-Sayre syndrome (KSS) and with mitochondrial encephalopathy with lactic acidosis and stroke-like episodes (MELAS). The present study aimed to characterize the prevalence and pattern of cardiac abnormalities and to test the additional diagnostic value of CMR in this patient population. The hypothesis that different neuromuscular MM syndromes present with different cardiac disease phenotypes was evaluated. Sixty-four MM patients (50 ± 15 years, 44% male) and 25 matched controls (52 ± 14 years, 36% male) prospectively underwent cardiac evaluations including CMR (comprising cine- and late-gadolinium-enhancement (LGE) imaging). Based on the neuromuscular phenotype and genotype, the patients were grouped: (a) CPEO/KSS (N = 33); (b) MELAS/-like (N = 11); c) myoclonic epilepsy with ragged-red fibers (MERRF) (N = 3) and d) other non-specific MM forms (N = 17). Among the 64 MM patients, 34 (53%) had at least one abnormal CMR finding: 18 (28%) demonstrated an impaired left ventricular ejection-fraction (LV-EF patients showed significantly higher maximal wall thickness (10 ± 3 vs. 8 ± 2 mm, p = 0.005) and concentricity (LV mass to end-diastolic volume: 0.84 ± 0.27 vs. 0.67 ± 0.11, p patients showed the highest frequency of cardiac disease (in 10/11 (91%)), a mostly concentric LV hypertrophy (6/9; 67%) with or

  6. Detection and Genotyping of Human Papillomavirus in Self-Obtained Cervicovaginal Samples by Using the FTA Cartridge: New Possibilities for Cervical Cancer Screening ▿

    Science.gov (United States)

    Lenselink, Charlotte H.; de Bie, Roosmarie P.; van Hamont, Dennis; Bakkers, Judith M. J. E.; Quint, Wim G. V.; Massuger, Leon F. A. G.; Bekkers, Ruud L. M.; Melchers, Willem 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 SPF10-Line Blot 25. Set 1 consisted of 45 women attending the gynecologist; all obtained a self-sampled cervicovaginal smear, which was applied to an FTA cartridge. HPV results were compared to a cervical smear (liquid based) taken by a trained physician. Set 2 consisted of 51 women who obtained a self-sampled cervicovaginal smear at home, which was applied to an FTA cartridge and to a liquid-based medium. DNA was obtained from the FTA cartridges by simple elution as well as extraction. Of all self-obtained samples of set 1, 62.2% tested HPV positive. The overall agreement between self- and physician-obtained samples was 93.3%, in favor of the self-obtained samples. In sample set 2, 25.5% tested HPV positive. The overall agreement for high-risk HPV presence between the FTA cartridge and liquid-based medium and between DNA elution and extraction was 100%. This study shows that HPV detection and genotyping in self-obtained cervicovaginal samples applied to an FTA cartridge is highly reliable. It shows a high level of overall agreement with HPV detection and genotyping in physician-obtained cervical smears and liquid-based self-samples. DNA can be obtained by simple elution and is therefore easy, cheap, and fast. Furthermore, the FTA cartridge is a convenient medium for collection and safe transport at ambient temperatures. Therefore, this method may contribute to a new way of cervical cancer screening. PMID:19553570

  7. Detection and genotyping of human papillomavirus in self-obtained cervicovaginal samples by using the FTA cartridge: new possibilities for cervical cancer screening.

    Science.gov (United States)

    Lenselink, Charlotte H; de Bie, Roosmarie P; van Hamont, Dennis; Bakkers, Judith M J E; Quint, Wim G V; Massuger, Leon F A G; Bekkers, Ruud L M; Melchers, Willem J G

    2009-08-01

    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 consisted of 45 women attending the gynecologist; all obtained a self-sampled cervicovaginal smear, which was applied to an FTA cartridge. HPV results were compared to a cervical smear (liquid based) taken by a trained physician. Set 2 consisted of 51 women who obtained a self-sampled cervicovaginal smear at home, which was applied to an FTA cartridge and to a liquid-based medium. DNA was obtained from the FTA cartridges by simple elution as well as extraction. Of all self-obtained samples of set 1, 62.2% tested HPV positive. The overall agreement between self- and physician-obtained samples was 93.3%, in favor of the self-obtained samples. In sample set 2, 25.5% tested HPV positive. The overall agreement for high-risk HPV presence between the FTA cartridge and liquid-based medium and between DNA elution and extraction was 100%. This study shows that HPV detection and genotyping in self-obtained cervicovaginal samples applied to an FTA cartridge is highly reliable. It shows a high level of overall agreement with HPV detection and genotyping in physician-obtained cervical smears and liquid-based self-samples. DNA can be obtained by simple elution and is therefore easy, cheap, and fast. Furthermore, the FTA cartridge is a convenient medium for collection and safe transport at ambient temperatures. Therefore, this method may contribute to a new way of cervical cancer screening.

  8. Sequence-specific label-free nucleic acid biosensor for the detection of the hepatitis C virus genotype 1a using a disposable pencil graphite electrode.

    Science.gov (United States)

    Donmez, Soner; Arslan, Fatma; Arslan, Halit

    2016-05-01

    In this paper, we demonstrate a simple, sensitive, inexpensive, disposable and label-free electrochemical nucleic acid biosensor for the detection of the hepatitis C virus genotype 1a (HCV1a). The nucleic acid biosensor was designed with the amino-linked inosine-substituted 20-mer probes, which were immobilized onto a disposable pencil graphite electrode (PGE) by covalent linking. The proposed nucleic acid biosensor was linear in the range of 0.05 and 0.75 μM, exhibiting a limit of detection of 54.9 nM. The single-stranded synthetic PCR product analogs of HCV1a were also detected with satisfactory results under optimal conditions, showing the potential application of this biosensor.

  9. In Vitro vs In Silico Detected SNPs for the Development of a Genotyping Array: What Can We Learn from a Non-Model Species?

    Science.gov (United States)

    Lepoittevin, Camille; Frigerio, Jean-Marc; Garnier-Géré, Pauline; Salin, Franck; Cervera, María-Teresa; Vornam, Barbara; Harvengt, Luc; Plomion, Christophe

    2010-01-01

    Background There is considerable interest in the high-throughput discovery and genotyping of single nucleotide polymorphisms (SNPs) to accelerate genetic mapping and enable association studies. This study provides an assessment of EST-derived and resequencing-derived SNP quality in maritime pine (Pinus pinaster Ait.), a conifer characterized by a huge genome size (∼23.8 Gb/C). Methodology/Principal Findings A 384-SNPs GoldenGate genotyping array was built from i/ 184 SNPs originally detected in a set of 40 re-sequenced candidate genes (in vitro SNPs), chosen on the basis of functionality scores, presence of neighboring polymorphisms, minor allele frequencies and linkage disequilibrium and ii/ 200 SNPs screened from ESTs (in silico SNPs) selected based on the number of ESTs used for SNP detection, the SNP minor allele frequency and the quality of SNP flanking sequences. The global success rate of the assay was 66.9%, and a conversion rate (considering only polymorphic SNPs) of 51% was achieved. In vitro SNPs showed significantly higher genotyping-success and conversion rates than in silico SNPs (+11.5% and +18.5%, respectively). The reproducibility was 100%, and the genotyping error rate very low (0.54%, dropping down to 0.06% when removing four SNPs showing elevated error rates). Conclusions/Significance This study demonstrates that ESTs provide a resource for SNP identification in non-model species, which do not require any additional bench work and little bio-informatics analysis. However, the time and cost benefits of in silico SNPs are counterbalanced by a lower conversion rate than in vitro SNPs. This drawback is acceptable for population-based experiments, but could be dramatic in experiments involving samples from narrow genetic backgrounds. In addition, we showed that both the visual inspection of genotyping clusters and the estimation of a per SNP error rate should help identify markers that are not suitable to the GoldenGate technology in species

  10. In vitro vs in silico detected SNPs for the development of a genotyping array: what can we learn from a non-model species?

    Directory of Open Access Journals (Sweden)

    Camille Lepoittevin

    2010-06-01

    Full Text Available There is considerable interest in the high-throughput discovery and genotyping of single nucleotide polymorphisms (SNPs to accelerate genetic mapping and enable association studies. This study provides an assessment of EST-derived and resequencing-derived SNP quality in maritime pine (Pinus pinaster Ait., a conifer characterized by a huge genome size ( approximately 23.8 Gb/C.A 384-SNPs GoldenGate genotyping array was built from i/ 184 SNPs originally detected in a set of 40 re-sequenced candidate genes (in vitro SNPs, chosen on the basis of functionality scores, presence of neighboring polymorphisms, minor allele frequencies and linkage disequilibrium and ii/ 200 SNPs screened from ESTs (in silico SNPs selected based on the number of ESTs used for SNP detection, the SNP minor allele frequency and the quality of SNP flanking sequences. The global success rate of the assay was 66.9%, and a conversion rate (considering only polymorphic SNPs of 51% was achieved. In vitro SNPs showed significantly higher genotyping-success and conversion rates than in silico SNPs (+11.5% and +18.5%, respectively. The reproducibility was 100%, and the genotyping error rate very low (0.54%, dropping down to 0.06% when removing four SNPs showing elevated error rates.This study demonstrates that ESTs provide a resource for SNP identification in non-model species, which do not require any additional bench work and little bio-informatics analysis. However, the time and cost benefits of in silico SNPs are counterbalanced by a lower conversion rate than in vitro SNPs. This drawback is acceptable for population-based experiments, but could be dramatic in experiments involving samples from narrow genetic backgrounds. In addition, we showed that both the visual inspection of genotyping clusters and the estimation of a per SNP error rate should help identify markers that are not suitable to the GoldenGate technology in species characterized by a large and complex genome.

  11. Real-time PCR assays for detection of Brucella spp. and the identification of genotype ST27 in bottlenose dolphins (Tursiops truncatus).

    Science.gov (United States)

    Wu, Qingzhong; McFee, Wayne E; Goldstein, Tracey; Tiller, Rebekah V; Schwacke, Lori

    2014-05-01

    Rapid detection of Brucella spp. in marine mammals is challenging. Microbiologic culture is used for definitive diagnosis of brucellosis, but is time consuming, has low sensitivity and can be hazardous to laboratory personnel. Serological methods can aid in diagnosis, but may not differentiate prior exposure versus current active infection and may cross-react with unrelated Gram-negative bacteria. This study reports a real-time PCR assay for the detection of Brucella spp. and application to screen clinical samples from bottlenose dolphins stranded along the coast of South Carolina, USA. The assay was found to be 100% sensitive for the Brucella strains tested, and the limit of detection was 0.27fg of genomic DNA from Brucella ceti B1/94 per PCR volume. No amplification was detected for the non-Brucella pathogens tested. Brucella DNA was detected in 31% (55/178) of clinical samples tested. These studies indicate that the real-time PCR assay is highly sensitive and specific for the detection of Brucella spp. in bottlenose dolphins. We also developed a second real-time PCR assay for rapid identification of Brucella ST27, a genotype that is associated with human zoonotic infection. Positive results were obtained for Brucella strains which had been identified as ST27 by multilocus sequence typing. No amplification was found for other Brucella strains included in this study. ST27 was identified in 33% (18/54) of Brucella spp. DNA-positive clinical samples. To our knowledge, this is the first report on the use of a real-time PCR assay for identification of Brucella genotype ST27 in marine mammals. Copyright © 2014 Elsevier B.V. All rights reserved.

  12. Porphyromonas gingivalis Fim-A genotype distribution among Colombians

    Science.gov (United States)

    Jaramillo, Adriana; Parra, Beatriz; Botero, Javier Enrique; Contreras, Adolfo

    2015-01-01

    Introduction: Porphyromonas gingivalis is associated with periodontitis and exhibit a wide array of virulence factors, including fimbriae which is encoded by the FimA gene representing six known genotypes. Objetive: To identify FimA genotypes of P. gingivalis in subjects from Cali-Colombia, including the co-infection with Aggregatibacter actinomycetemcomitans, Treponema denticola, and Tannerella forsythia. Methods: Subgingival samples were collected from 151 people exhibiting diverse periodontal condition. The occurrence of P. gingivalis, FimA genotypes and other bacteria was determined by PCR. Results: P. gingivalis was positive in 85 patients. Genotype FimA II was more prevalent without reach significant differences among study groups (54.3%), FimA IV was also prevalent in gingivitis (13.0%). A high correlation (p= 0.000) was found among P. gingivalis, T. denticola, and T. forsythia co-infection. The FimA II genotype correlated with concomitant detection of T. denticola and T. forsythia. Conclusions: Porphyromonas gingivalis was high even in the healthy group at the study population. A trend toward a greater frequency of FimA II genotype in patients with moderate and severe periodontitis was determined. The FimA II genotype was also associated with increased pocket depth, greater loss of attachment level, and patients co-infected with T. denticola and T. forsythia. PMID:26600627

  13. Detection and genotyping of torque teno virus (TTV) in healthy blood donors and patients infected with HBV or HCV in Qatar.

    Science.gov (United States)

    AbuOdeh, Raed; Al-Mawlawi, Naema; Al-Qahtani, Ahmed A; Bohol, Marie Fe F; Al-Ahdal, Mohammed N; Hasan, Haydar A; AbuOdeh, Lamees; Nasrallah, Gheyath K

    2015-07-01

    Torque Teno virus (TTV) has been associated with non A-G hepatitis. The goal of this study was to estimate the infection rates and genotypic characteristics of TTV in the State of Qatar. A total of 644 blood samples representing different nationalities: (i) Qatari (118) and (ii) non-Qatari (526) nationals (mostly from Arab and South Eeast Asia countries) were tested for the presence of TTV DNA by nested PCR. The majority (573) of the blood samples belonged to healthy blood donors, whereas 54 and 53 of the blood samples belonged to patients infected with hepatitis B virus (HBV) and hepatitis C virus (HCV), respectively. The results obtained showed that the TTV infection rates in the healthy blood donors, and those infected with HBV or HCV patients were 81.4, 90.75 and 84.9%, respectively. Significant association between TTV viremia and age, or nationality was observed. Sequence analysis of PCR fragments amplified from the 5'-untranslated region (5'-UTR) of all (531) TTV positive samples showed that 65.5% (348/531) of the PCR fragment sequences were classified into main genogroup 3, followed by main genogroups 5 (24%), 2 (5.8%), and 1 (4.7%). Genogroup 4 was not detected among the our studied subjects. Phylogenetic and pairwise analyses using sequences from TTV viremic samples also showed an overall close similarity to the main genogroup 3. In conclusion, there was no significant difference in the rates of TTV detection among Qataris and non-Qataris and several genotypes, mainly genotype 3, were isolated. © 2015 Wiley Periodicals, Inc.

  14. DNA methylation changes detected by methylation-sensitive amplified polymorphism in two contrasting rice genotypes under salt stress.

    Science.gov (United States)

    Wang, Wensheng; Zhao, Xiuqin; Pan, Yajiao; Zhu, Linghua; Fu, Binying; Li, Zhikang

    2011-09-20

    DNA methylation, one of the most important epigenetic phenomena, plays a vital role in tuning gene expression during plant development as well as in response to environmental stimuli. In the present study, a methylation-sensitive amplified polymorphism (MSAP) analysis was performed to profile DNA methylation changes in two contrasting rice genotypes under salt stress. Consistent with visibly different phenotypes in response to salt stress, epigenetic markers classified as stable inter-cultivar DNA methylation differences were determined between salt-tolerant FL478 and salt-sensitive IR29. In addition, most tissue-specific DNA methylation loci were conserved, while many of the growth stage-dependent DNA methylation loci were dynamic between the two genotypes. Strikingly, salt stress induced a decrease in DNA methylation specifically in roots at the seedling stage that was more profound in IR29 than in the FL478. This result may indicate that demethylation of genes is an active epigenetic response to salt stress in roots at the seedling stage, and helps to further elucidate the implications of DNA methylation in crop growth and development. Copyright © 2011. Published by Elsevier Ltd.

  15. Detection of superior genotype of fatty acid synthase in Korean native cattle by an environment-adjusted statistical model

    Directory of Open Access Journals (Sweden)

    Jea-Young Lee

    2017-06-01

    Full Text Available Objective This study examines the genetic factors influencing the phenotypes (four economic traits:oleic acid [C18:1], monounsaturated fatty acids, carcass weight, and marbling score of Hanwoo. Methods To enhance the accuracy of the genetic analysis, the study proposes a new statistical model that excludes environmental factors. A statistically adjusted, analysis of covariance model of environmental and genetic factors was developed, and estimated environmental effects (covariate effects of age and effects of calving farms were excluded from the model. Results The accuracy was compared before and after adjustment. The accuracy of the best single nucleotide polymorphism (SNP in C18:1 increased from 60.16% to 74.26%, and that of the two-factor interaction increased from 58.69% to 87.19%. Also, superior SNPs and SNP interactions were identified using the multifactor dimensionality reduction method in Table 1 to 4. Finally, high- and low-risk genotypes were compared based on their mean scores for each trait. Conclusion The proposed method significantly improved the analysis accuracy and identified superior gene-gene interactions and genotypes for each of the four economic traits of Hanwoo.

  16. HBV genotypic variability in Cuba.

    Directory of Open Access Journals (Sweden)

    Carmen L Loureiro

    Full Text Available The genetic diversity of HBV in human population is often a reflection of its genetic admixture. The aim of this study was to explore the genotypic diversity of HBV in Cuba. The S genomic region of Cuban HBV isolates was sequenced and for selected isolates the complete genome or precore-core sequence was analyzed. The most frequent genotype was A (167/250, 67%, mainly A2 (149, 60% but also A1 and one A4. A total of 77 isolates were classified as genotype D (31%, with co-circulation of several subgenotypes (56 D4, 2 D1, 5 D2, 7 D3/6 and 7 D7. Three isolates belonged to genotype E, two to H and one to B3. Complete genome sequence analysis of selected isolates confirmed the phylogenetic analysis performed with the S region. Mutations or polymorphisms in precore region were more common among genotype D compared to genotype A isolates. The HBV genotypic distribution in this Caribbean island correlates with the Y lineage genetic background of the population, where a European and African origin prevails. HBV genotypes E, B3 and H isolates might represent more recent introductions.

  17. HBV Genotypic Variability in Cuba

    Science.gov (United States)

    Loureiro, Carmen L.; Aguilar, Julio C.; Aguiar, Jorge; Muzio, Verena; Pentón, Eduardo; Garcia, Daymir; Guillen, Gerardo; Pujol, Flor H.

    2015-01-01

    The genetic diversity of HBV in human population is often a reflection of its genetic admixture. The aim of this study was to explore the genotypic diversity of HBV in Cuba. The S genomic region of Cuban HBV isolates was sequenced and for selected isolates the complete genome or precore-core sequence was analyzed. The most frequent genotype was A (167/250, 67%), mainly A2 (149, 60%) but also A1 and one A4. A total of 77 isolates were classified as genotype D (31%), with co-circulation of several subgenotypes (56 D4, 2 D1, 5 D2, 7 D3/6 and 7 D7). Three isolates belonged to genotype E, two to H and one to B3. Complete genome sequence analysis of selected isolates confirmed the phylogenetic analysis performed with the S region. Mutations or polymorphisms in precore region were more common among genotype D compared to genotype A isolates. The HBV genotypic distribution in this Caribbean island correlates with the Y lineage genetic background of the population, where a European and African origin prevails. HBV genotypes E, B3 and H isolates might represent more recent introductions. PMID:25742179

  18. Use of (D, MUF) and maximum-likelihood methods for detecting falsification and diversion in data-verification problems

    International Nuclear Information System (INIS)

    Goldman, A.S.; Beedgen, R.

    1982-01-01

    The investigation of data falsification and/or diversion is of major concern in nuclear materials accounting procedures used in international safeguards. In this paper, two procedures, denoted by (D,MUF) and LR (Likelihood Ratio), are discussed and compared when testing the hypothesis that neither diversion nor falsification has taken place versus the one-sided alternative that at least one of these parameters is positive. Critical regions and detection probabilities are given for both tests. It is shown that the LR method outperforms (D,MUF) when diversion and falsification take place

  19. Detecting differential DNA methylation from sequencing of bisulfite converted DNA of diverse species.

    Science.gov (United States)

    Huh, Iksoo; Wu, Xin; Park, Taesung; Yi, Soojin V

    2017-07-21

    DNA methylation is one of the most extensively studied epigenetic modifications of genomic DNA. In recent years, sequencing of bisulfite-converted DNA, particularly via next-generation sequencing technologies, has become a widely popular method to study DNA methylation. This method can be readily applied to a variety of species, dramatically expanding the scope of DNA methylation studies beyond the traditionally studied human and mouse systems. In parallel to the increasing wealth of genomic methylation profiles, many statistical tools have been developed to detect differentially methylated loci (DMLs) or differentially methylated regions (DMRs) between biological conditions. We discuss and summarize several key properties of currently available tools to detect DMLs and DMRs from sequencing of bisulfite-converted DNA. However, the majority of the statistical tools developed for DML/DMR analyses have been validated using only mammalian data sets, and less priority has been placed on the analyses of invertebrate or plant DNA methylation data. We demonstrate that genomic methylation profiles of non-mammalian species are often highly distinct from those of mammalian species using examples of honey bees and humans. We then discuss how such differences in data properties may affect statistical analyses. Based on these differences, we provide three specific recommendations to improve the power and accuracy of DML and DMR analyses of invertebrate data when using currently available statistical tools. These considerations should facilitate systematic and robust analyses of DNA methylation from diverse species, thus advancing our understanding of DNA methylation. © The Author 2017. Published by Oxford University Press.

  20. Genetic diversity analysis of sugarcane germplasm based on fluorescence-labeled simple sequence repeat markers and a capillary electrophoresis-based genotyping platform

    Science.gov (United States)

    Genetic diversity analysis, which refers to the elaboration of total extent of genetic characteristics in the genetic makeup of a certain species, constitutes a classical strategy for the study of diversity, population genetic structure, and breeding practices. In this study, fluorescence-labeled se...

  1. Novel procedure for genotyping of the human serotonin transporter gene-linked polymorphic region (5-HTTLPR)--a region with a high level of allele diversity

    DEFF Research Database (Denmark)

    Rasmussen, Henrik B; Werge, Thomas M

    2007-01-01

    determination. After having developed a 5-HTTLPR genotyping assay, we examined all samples of DNA in two separate rounds of analyses and found complete agreement between the results from these two rounds. CONCLUSION: On the basis of simultaneous analysis of tandem repeat size variation and variation of single......BACKGROUND: The serotonin transporter, the target of a group of antidepressant drugs, is involved in the regulation of the availability and reuptake of serotonin. A variable number of tandem repeats in the promoter region of the serotonin transporter gene, designated 5-HTTLPR, affects...... for detailed genotyping of 5-HTTLPR based upon simultaneous analysis of tandem repeat size variation and single nucleotide variations. METHODS: We elaborated a list of all known 5-HTTLPR alleles to provide an overview of the allele repertoire at this polymorphic locus. Fragments of 5-HTTLPR were PCR...

  2. Quick survey for detection, identification and characterization of Acanthamoeba genotypes from some selected soil and water samples in Pakistan

    Directory of Open Access Journals (Sweden)

    Tania Tanveer

    2015-05-01

    Full Text Available Acanthamoeba is an opportunistic protozoan pathogen which is widely distributed in nature and plays a pivotal role in ecosystem. Acanthamoeba species may cause blinding keratitis and fatal granulomatous encephalitis involving central nervous system. In this study, we investigated the presence of Acanthamoeba in soil and water resources of Pakistan. Here, Acanthamoeba were recovered on non-nutrient agar plate lawn with E.coli and identified by morphological characteristics of the cyst. Furthermore PCR was performed with genus-specific primers followed by direct sequencing of the PCR product for molecular identification. Overall our PCR and sequencing results confirmed pathogenic genotypes including T4 and T15 from both soil and water samples. This is our first report of Acanthamoeba isolation from both soil and water resources of Pakistan which may serve as a potential treat to human health across the country.

  3. SNP design from 454 sequencing of Podosphaera plantaginis transcriptome reveals a genetically diverse pathogen metapopulation with high levels of mixed-genotype infection.

    Directory of Open Access Journals (Sweden)

    Charlotte Tollenaere

    Full Text Available Molecular tools may greatly improve our understanding of pathogen evolution and epidemiology but technical constraints have hindered the development of genetic resources for parasites compared to free-living organisms. This study aims at developing molecular tools for Podosphaera plantaginis, an obligate fungal pathogen of Plantago lanceolata. This interaction has been intensively studied in the Åland archipelago of Finland with epidemiological data collected from over 4,000 host populations annually since year 2001.A cDNA library of a pooled sample of fungal conidia was sequenced on the 454 GS-FLX platform. Over 549,411 reads were obtained and annotated into 45,245 contigs. Annotation data was acquired for 65.2% of the assembled sequences. The transcriptome assembly was screened for SNP loci, as well as for functionally important genes (mating-type genes and potential effector proteins. A genotyping assay of 27 SNP loci was designed and tested on 380 infected leaf samples from 80 populations within the Åland archipelago. With this panel we identified 85 multilocus genotypes (MLG with uneven frequencies across the pathogen metapopulation. Approximately half of the sampled populations contain polymorphism. Our genotyping protocol revealed mixed-genotype infection within a single host leaf to be common. Mixed infection has been proposed as one of the main drivers of pathogen evolution, and hence may be an important process in this pathosystem.The developed SNP panel offers exciting research perspectives for future studies in this well-characterized pathosystem. Also, the transcriptome provides an invaluable novel genomic resource for powdery mildews, which cause significant yield losses on commercially important crops annually. Furthermore, the features that render genetic studies in this system a challenge are shared with the majority of obligate parasitic species, and hence our results provide methodological insights from SNP calling to field

  4. Genomic Analysis of 15 Human Coronaviruses OC43 (HCoV-OC43s Circulating in France from 2001 to 2013 Reveals a High Intra-Specific Diversity with New Recombinant Genotypes

    Directory of Open Access Journals (Sweden)

    Nathalie Kin

    2015-05-01

    Full Text Available Human coronavirus OC43 (HCoV-OC43 is one of five currently circulating human coronaviruses responsible for respiratory infections. Like all coronaviruses, it is characterized by its genome’s high plasticity. The objectives of the current study were to detect genetically distinct genotypes and eventually recombinant genotypes in samples collected in Lower Normandy between 2001 and 2013. To this end, we sequenced complete nsp12, S, and N genes of 15 molecular isolates of HCoV-OC43 from clinical samples and compared them to available data from the USA, Belgium, and Hong-Kong. A new cluster E was invariably detected from nsp12, S, and N data while the analysis of nsp12 and N genes revealed the existence of new F and G clusters respectively. The association of these different clusters of genes in our specimens led to the description of thirteen genetically distinct genotypes, among which eight recombinant viruses were discovered. Identification of these recombinant viruses, together with temporal analysis and tMRCA estimation, provides important information for understanding the dynamics of the evolution of these epidemic coronaviruses.

  5. Detection, quantification and genotyping of Herpes Simplex Virus in cervicovaginal secretions by real-time PCR: a cross sectional survey

    Directory of Open Access Journals (Sweden)

    Natividad-Sancho Angels

    2005-08-01

    Full Text Available Abstract Background Herpes Simplex Virus (HSV Genital Ulcer Disease (GUD is an important public health problem, whose interaction with HIV results in mutually enhancing epidemics. Conventional methods for detecting HSV tend to be slow and insensitive. We designed a rapid PCR-based assay to quantify and type HSV in cervicovaginal lavage (CVL fluid of subjects attending a Genito-Urinary Medicine (GUM clinic. Vaginal swabs, CVL fluid and venous blood were collected. Quantitative detection of HSV was conducted using real time PCR with HSV specific primers and SYBR Green I. Fluorogenic TaqMan Minor Groove Binder (MGB probes designed around a single base mismatch in the HSV DNA polymerase I gene were used to type HSV in a separate reaction. The Kalon test was used to detect anti-HSV-2 IgG antibodies in serum. Testing for HIV, other Sexually Transmitted Infections (STI and related infections was based on standard clinical and laboratory methods. Results Seventy consecutive GUM clinic attendees were studied. Twenty-seven subjects (39% had detectable HSV DNA in CVL fluid; HSV-2 alone was detected in 19 (70% subjects, HSV-1 alone was detected in 4 (15% subjects and both HSV types were detected in 4 (15% subjects. Eleven out of 27 subjects (41% with anti-HSV-2 IgG had detectable HSV-2 DNA in CVL fluid. Seven subjects (10% were HIV-positive. Three of seven (43% HIV-infected subjects and two of five subjects with GUD (40% were secreting HSV-2. None of the subjects in whom HSV-1 was detected had GUD. Conclusion Quantitative real-time PCR and Taqman MGB probes specific for HSV-1 or -2 were used to develop an assay for quantification and typing of HSV. The majority of subjects in which HSV was detected had low levels of CVL fluid HSV, with no detectable HSV-2 antibodies and were asymptomatic.

  6. Assessing Symbiodinium diversity in scleractinian corals via next-generation sequencing-based genotyping of the ITS2 rDNA region

    KAUST Repository

    Arif, Chatchanit; Daniels, Camille; Bayer, Till; Banguera Hinestroza, Eulalia; Barbrook, Adrian; Howe, Christopher J.; LaJeunesse, Todd C.; Voolstra, Christian R.

    2014-01-01

    The persistence of coral reef ecosystems relies on the symbiotic relationship between scleractinian corals and intracellular, photosynthetic dinoflagellates in the genus Symbiodinium. Genetic evidence indicates that these symbionts are biologically diverse and exhibit discrete patterns of environmental and host distribution. This makes the assessment of Symbiodinium diversity critical to understanding the symbiosis ecology of corals. Here, we applied pyrosequencing to the elucidation of Symbiodinium diversity via analysis of the internal transcribed spacer 2 (ITS2) region, a multicopy genetic marker commonly used to analyse Symbiodinium diversity. Replicated data generated from isoclonal Symbiodinium cultures showed that all genomes contained numerous, yet mostly rare, ITS2 sequence variants. Pyrosequencing data were consistent with more traditional denaturing gradient gel electrophoresis (DGGE) approaches to the screening of ITS2 PCR amplifications, where the most common sequences appeared as the most intense bands. Further, we developed an operational taxonomic unit (OTU)-based pipeline for Symbiodinium ITS2 diversity typing to provisionally resolve ecologically discrete entities from intragenomic variation. A genetic distance cut-off of 0.03 collapsed intragenomic ITS2 variants of isoclonal cultures into single OTUs. When applied to the analysis of field-collected coral samples, our analyses confirm that much of the commonly observed Symbiodinium ITS2 diversity can be attributed to intragenomic variation. We conclude that by analysing Symbiodinium populations in an OTU-based framework, we can improve objectivity, comparability and simplicity when assessing ITS2 diversity in field-based studies.

  7. Assessing Symbiodinium diversity in scleractinian corals via next-generation sequencing-based genotyping of the ITS2 rDNA region

    KAUST Repository

    Arif, Chatchanit

    2014-09-01

    The persistence of coral reef ecosystems relies on the symbiotic relationship between scleractinian corals and intracellular, photosynthetic dinoflagellates in the genus Symbiodinium. Genetic evidence indicates that these symbionts are biologically diverse and exhibit discrete patterns of environmental and host distribution. This makes the assessment of Symbiodinium diversity critical to understanding the symbiosis ecology of corals. Here, we applied pyrosequencing to the elucidation of Symbiodinium diversity via analysis of the internal transcribed spacer 2 (ITS2) region, a multicopy genetic marker commonly used to analyse Symbiodinium diversity. Replicated data generated from isoclonal Symbiodinium cultures showed that all genomes contained numerous, yet mostly rare, ITS2 sequence variants. Pyrosequencing data were consistent with more traditional denaturing gradient gel electrophoresis (DGGE) approaches to the screening of ITS2 PCR amplifications, where the most common sequences appeared as the most intense bands. Further, we developed an operational taxonomic unit (OTU)-based pipeline for Symbiodinium ITS2 diversity typing to provisionally resolve ecologically discrete entities from intragenomic variation. A genetic distance cut-off of 0.03 collapsed intragenomic ITS2 variants of isoclonal cultures into single OTUs. When applied to the analysis of field-collected coral samples, our analyses confirm that much of the commonly observed Symbiodinium ITS2 diversity can be attributed to intragenomic variation. We conclude that by analysing Symbiodinium populations in an OTU-based framework, we can improve objectivity, comparability and simplicity when assessing ITS2 diversity in field-based studies.

  8. Clinical application of RT-nested PCR integrated with RFLP in Hantavirus detection and genotyping: a prospective study in Shandong Province, PR China.

    Science.gov (United States)

    Liu, Yun-Xi; Zhao, Zhong-Tang; Cao, Wu-Chun; Xu, Xiao-Qun; Suo, Ji-Jiang; Xing, Yu-Bin; Jia, Ning; Du, Ming-Mei; Liu, Bo-Wei; Yao, Yuan

    2013-01-01

    The aim of the present study was to evaluate the clinical usefulness of applying RT-nested PCR along with RFLP as a method for diagnosis and genotypic differentiation of Hantavirus in the acute-stage sera of HFRS patients as compared to the ELISA technique. A prospective study of patients with suspected HFRS patients was carried out. Sera were collected for serological evaluation by ELISA and RT-nested PCR testing. Primers were selected from the published sequence of the S segment of HTNV strain 76-118 and SEOV strain SR-11, which made it possible to obtain an amplicon of 403 bp by RT-nested PCR. The genotypic differentiations of the RT-nested PCR amplicons were carried out by RFLP. Sequence analyses of the amplicons were used to confirm the accuracy of the results obtained by RFLP. Of the 48 acute-stage sera from suspected HFRS patients, 35 were ELISA-positive while 41 were positive by RT-nested PCR. With Hind III and Hinf I, RFLP profiles of the RT-nested PCR amplicons of the 41 positive sera exhibited two patterns. 33 had RFLP profiles similar to the reference strain R22, and thus belonged to the SEOV type. The other 8 samples which were collected during October-December had RFLP profiles similar to the reference strain 76-118, and thus belonged to the HTNV type. Sequence phylogenetic analysis of RT-nested PCR amplicons revealed sdp1, sdp2 YXL-2008, and sdp3 as close relatives of HTNV strain 76-118, while sdp22 and sdp37 as close relatives of SEOV strain Z37 and strain R22 located in two separate clusters in the phylogenetic tree. These results were identical to those acquired by RFLP. RT-nested PCR integrated with RFLP was a rapid, simple, accurate method for detecting and differentiating the genotypes of Hantavirus in the acute-stage sera of suspected HFRS patients. In Shandong province, the main genotypes of Hantavirus belonged to the SEOV types, while the HTNV types were observed during the autumn-winter season.

  9. Molecular detection of genotype II grass carp reovirus based on nucleic acid sequence-based amplification combined with enzyme-linked immunosorbent assay (NASBA-ELISA).

    Science.gov (United States)

    Zeng, Weiwei; Yao, Wei; Wang, Yingying; Li, Yingying; Bermann, Sven M; Ren, Yan; Shi, Cunbin; Song, Xinjian; Huang, Qiwen; Zheng, Shuchen; Wang, Qing

    2017-05-01

    Grass carp reovirus (GCRV) is the causative agent of the grass carp hemorrhagic disease that has resulted in severe economic losses in the grass carp (Ctenopharyngodon idella) farming industry in China. Early diagnosis and vaccine administration are important priorities for GCRV control. In this study, a nucleic acid sequence-based amplification with enzyme-linked immunosorbent assay (NASBA-ELISA) was developed for to detect genotype II GCRV (GCRV- II). Primers specifically targeting viral RNA genome segment 6 were utilized for amplification in an isothermal digoxigenin-labeling NASBA process, resulting in DIG-labeled RNA amplicons. The amplicons were hybridized to specific biotinylated DNA probes and the products were detected colorimetrically using horseradish peroxidase and a microplate reader. The new method is able to detect GCRV at 14 copies/μL within 5h and had a diagnostic sensitivity and a specificity of 100% when GCRV-II and non-target virus were tested. This NASBA-ELISA was evaluated using a panel of clinical samples (n=103) to demonstrate that it is a rapid, effective and sensitive method for GCRV detection in grass carp aquaculture. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. THE USE OF MULTIPLE DISPLACEMENT AMPLIFICATION TO INCREASE THE DETECTION AND GENOTYPING OF TRYPANOSOMA SPECIES SAMPLES IMMOBILISED ON FTA FILTERS

    Science.gov (United States)

    MORRISON, LIAM J.; McCORMACK, GILLIAN; SWEENEY, LINDSAY; LIKEUFACK, ANNE C. L.; TRUC, PHILIPPE; TURNER, C. MICHAEL; TAIT, ANDY; MacLEOD, ANNETTE

    2007-01-01

    Whole genome amplification methods are a recently developed tool for amplifying DNA from limited template. We report its application in trypanosome infections, characterised by low parasitaemias. Multiple Displacement Amplification (MDA) amplifies DNA with a simple in vitro step, and was evaluated on mouse blood samples on FTA filter cards with known numbers of Trypanosoma brucei parasites. The data showed a twenty-fold increase in the number of PCRs possible per sample, using primers diagnostic for the multi-copy ribosomal ITS region or 177 bp repeats, and a twenty-fold increase in sensitivity over nested PCR against a single copy microsatellite. Using MDA for microsatellite genotyping caused allele dropout at low DNA concentrations, which was overcome by pooling multiple MDA reactions. The validity of using MDA was established with samples from Human African Trypanosomiasis patients. The use of MDA allows maximal use of finite DNA samples and may prove a valuable tool in studies where multiple reactions are necessary, such as population genetic analyses. PMID:17556624

  11. Detection of Fetomaternal Genotype Associations in Early-Onset Disorders: Evaluation of Different Methods and Their Application to Childhood Leukemia

    Directory of Open Access Journals (Sweden)

    Jasmine Healy

    2010-01-01

    Full Text Available Several designs and analytical approaches have been proposed to dissect offspring from maternal genetic contributions to early-onset diseases. However, lack of parental controls halts the direct verification of the assumption of mating symmetry (MS required to assess maternally-mediated effects. In this study, we used simulations to investigate the performance of existing methods under mating asymmetry (MA when parents of controls are missing. Our results show that the log-linear, likelihood-based framework using a case-triad/case-control hybrid design provides valid tests for maternal genetic effects even under MA. Using this approach, we examined fetomaternal associations between 29 SNPs in 12 cell-cycle genes and childhood pre-B acute lymphoblastic leukemia (ALL. We identified putative fetomaternal effects at loci CDKN2A rs36228834 (P=.017 and CDKN2B rs36229158 (P=.022 that modulate the risk of childhood ALL. These data further corroborate the importance of the mother's genotype on the susceptibility to early-onset diseases.

  12. PCR detection and genetic diversity of bovine hemoprotozoan parasites in Vietnam.

    Science.gov (United States)

    Sivakumar, Thillaiampalam; Lan, Dinh Thi Bich; Long, Phung Thang; Yoshinari, Takeshi; Tattiyapong, Muncharee; Guswanto, Azirwan; Okubo, Kazuhiro; Igarashi, Ikuo; Inoue, Noboru; Xuan, Xuenan; Yokoyama, Naoaki

    2013-11-01

    Hemoprotozoan infections often cause serious production losses in livestock. In the present study, we conducted a PCR-based survey of Babesia bovis, Babesia bigemina, Theileria annulata, Theileria orientalis, Trypanosoma evansi and Trypanosoma theileri, using 423 DNA samples extracted from blood samples of cattle (n=202), water buffaloes (n=43), sheep (n=51) and goats (n=127) bred in the Hue and Hanoi provinces of Vietnam. With the exception of T. annulata and T. evansi, all other parasite species (B. bovis, B. bigemina, T. orientalis and T. theileri) were detected in the cattle populations with B. bovis being the most common among them. Additionally, four water buffaloes and a single goat were infected with B. bovis and B. bigemina, respectively. The Hue province had more hemoprotozoan-positive animals than those from the Hanoi region. In the phylogenetic analyses, B. bovis-MSA-2b, B. bigemina-AMA-1 and T. theileri-CATL gene sequences were dispersed across four, one and three different clades in the respective phylograms. This is the first study in which the presence of Babesia, Theileria and Trypanosoma parasites was simultaneously investigated by PCR in Vietnam. The findings suggest that hemoprotozoan parasites, some of which are genetically diverse, continue to be a threat to the livestock industry in this country.

  13. The Use of TILLING Technique to Detect Mutations and genetic diversity in Potato

    International Nuclear Information System (INIS)

    Elias, R; Al-Safadi, B; Till, B

    2008-01-01

    TILLING technique has been used, for the first time, to detect genetic variation, at the molecular level, among potato mutants (induced by gamma irradiation) and genetic diversity among 3 potato cultivars. Three potato mutant lines (every mutant represents a cultivar) tolerant to salinity have been used along with their controls. Three primer pairs were designed with the help of Potato Genome Sequencing Consortium on the web and were evaluated using agarose gel then sequencing. Primer pairs passing these tests were fluorescently labeled. Li-Cor based TILLING was applied using 2 forward primers one of them is labeled and 2 reverse primers one of them is labeled. The results have shown the success of using this technique on potato (tetraploid species) where the average density of nucleotide polymorphisms per sample was 16 polymorphisms per 1 kb. The optimal concentration was also determined between 0.1 and 1 ng/ul for potato genomic DNAs to be used in Li-Cor based TILLING assays. (author)

  14. Coherent scattering noise reduction method with wavelength diversity detection for holographic data storage system

    Science.gov (United States)

    Nakamura, Yusuke; Hoshizawa, Taku; Takashima, Yuzuru

    2017-09-01

    A new method, wavelength diversity detection (WDD), for improving signal quality is proposed and its effectiveness is numerically confirmed. We consider that WDD is especially effective for high-capacity systems having low hologram diffraction efficiencies. In such systems, the signal quality is primarily limited by coherent scattering noise; thus, effective improvement of the signal quality under a scattering-limited system is of great interest. WDD utilizes a new degree of freedom, the spectrum width, and scattering by molecules to improve the signal quality of the system. We found that WDD improves the quality by counterbalancing the degradation of the quality due to Bragg mismatch. With WDD, a higher-scattering-coefficient medium can improve the quality. The result provides an interesting insight into the requirements for material characteristics, especially for a large-M/# material. In general, a larger-M/# material contains more molecules; thus, the system is subject to more scattering, which actually improves the quality with WDD. We propose a pathway for a future holographic data storage system (HDSS) using WDD, which can record a larger amount of data than a conventional HDSS.

  15. Comparison of the performance in detection of HPV infections between the high-risk HPV genotyping real time PCR and the PCR-reverse dot blot assays.

    Science.gov (United States)

    Zhang, Lahong; Dai, Yibei; Chen, Jiahuan; Hong, Liquan; Liu, Yuhua; Ke, Qiang; Chen, Yiwen; Cai, Chengsong; Liu, Xia; Chen, Zhaojun

    2018-01-01

    A new multiplex real-time PCR assay, the high-risk HPV genotyping real time PCR assay (HR HPV RT-PCR), has been developed to detect 15 high-risk HPV types with respective viral loads. In this report, a total of 684 cervical specimens from women diagnosed with vaginitis were assessed by the HR HPV RT-PCR and the PCR reaction and reverse dot blot (PCR-RDB) assays, using a PCR-sequencing method as a reference standard. A total coincidence of 97.7% between the HR HPV RT PCR and the PCR-RDB assays was determined with a Kappa value of 0.953. The HR HPV RT PCR assay had sensitivity, specificity, and concordance rates (accuracy) of 99.7%, 99.7%, and 99.7%, respectively, as confirmed by PCR-sequencing, while the PCR-RDB assay had respective rates of 98.8%, 97.1%, and 98.0%. The overall rate of HPV infection, determined by PCR-sequencing, in women diagnosed with vaginitis was 49.85%, including 36.26% of single infection and 13.6% of multiple infections. The most common infections among the 15 high-risk HPV types in women diagnosed with vaginitis were HPV-52, HPV-16, and HPV-58, with a total detection rate of 10.23%, 7.75%, and 5.85%, respectively. We conclude that the HR HPV RT PCR assay exhibits better clinical performance than the PCR-RDB assay, and is an ideal alternative method for HPV genotyping. In addition, the HR HPV RT PCR assay provides HPV DNA viral loads, and could serve as a quantitative marker in the diagnosis and treatment of single and multiple HPV infections. © 2017 Wiley Periodicals, Inc.

  16. Primary Screening for Cervical Cancer Based on High-Risk Human Papillomavirus (HPV) Detection and HPV 16 and HPV 18 Genotyping, in Comparison to Cytology

    Science.gov (United States)

    Constantinidis, Theocharis; Constantinidis, Theodoros C.

    2015-01-01

    Objectives The objective of the present study is to assess the performance of a high-risk human papillomavirus (HR-HPV) DNA test with individual HPV-16/HPV-18 genotyping as a method for primary cervical cancer screening compared with liquid-based cytology (LBC) in a population of Greek women taking part in routine cervical cancer screening. Methods The study, conducted by the “HEllenic Real life Multicentric cErvical Screening” (HERMES) study group, involved the recruitment of 4,009 women, aged 25–55, who took part in routine cervical screening at nine Gynecology Departments in Greece. At first visit cervical specimens were collected for LBC and HPV testing using the Roche Cobas 4800 system. Women found positive for either cytology or HPV were referred for colposcopy, whereas women negative for both tests will be retested after three years. The study is ongoing and the results of the first screening round are reported herein. Results Valid results for cytology and HPV testing were obtained for 3,993 women. The overall prevalence of HR-HPV was 12.7%, of HPV-16 2.7% and of HPV-18 1.4%. Of those referred for colposcopy, cervical intraepithelial neoplasia grade 2 or worse (CIN2+) was detected in 41 women (1.07%). At the threshold of CIN2+, cytology [atypical squamous cells of undetermined significance (ASC-US) or worse] and HPV testing showed a sensitivity of 53.7% and 100% respectively, without change between age groups. Cytology and HPV testing showed specificity of 96.8% and 90.3% respectively, which was increased in older women (≥30) in comparison to younger ones (25–29). Genotyping for HPV16/18 had similar accuracy to cytology for the detection of CIN2+ (sensitivity: 58.5%; specificity 97.5%) as well as for triage to colposcopy (sensitivity: 58.5% vs 53.7% for cytology). Conclusion HPV testing has much better sensitivity than cytology to identify high-grade cervical lesions with slightly lower specificity. HPV testing with individual HPV-16/HPV-18

  17. Desmanthus GENOTYPES

    Directory of Open Access Journals (Sweden)

    JOSÉ HENRIQUE DE ALBUQUERQUE RANGEL

    2015-01-01

    Full Text Available Desmanthus is a genus of forage legumes with potential to improve pastures and livestock produc-tion on clay soils of dry tropical and subtropical regions such as the existing in Brazil and Australia. Despite this patterns of natural or enforced after-ripening of Desmanthus seeds have not been well established. Four year old seed banks of nine Desmanthus genotypes at James Cook University were accessed for their patterns of seed softe-ning in response to a range of temperatures. Persistent seed banks were found to exist under all of the studied ge-notypes. The largest seeds banks were found in the genotypes CPI 78373 and CPI 78382 and the smallest in the genotypes CPI’s 37143, 67643, and 83563. An increase in the percentage of softened seeds was correlated with higher temperatures, in two patterns of response: in some accessions seeds were not significantly affected by tempe-ratures below 80º C; and in others, seeds become soft when temperature rose to as little as 60 ºC. At 80 °C the heat started to depress germination. High seed production of Desmanthus associated with dependence of seeds on eleva-ted temperatures to softening can be a very important strategy for plants to survive in dry tropical regions.

  18. Genetic diversity of five local Swedish chicken breeds detected by microsatellite markers.

    Directory of Open Access Journals (Sweden)

    Abiye Shenkut Abebe

    Full Text Available This study aimed at investigating the genetic diversity, relationship and population structure of 110 local Swedish chickens derived from five breeds (Gotlandshöna, Hedemorahöna, Öländsk dvärghöna, Skånsk blommehöna, and Bohuslän- Dals svarthöna, in the rest of the paper the shorter name Svarthöna is used using 24 microsatellite markers. In total, one hundred thirteen alleles were detected in all populations, with a mean of 4.7 alleles per locus. For the five chicken breeds, the observed and expected heterozygosity ranged from 0.225 to 0.408 and from 0.231 to 0.515, with the lowest scores for the Svarthöna and the highest scores for the Skånsk blommehöna breeds, respectively. Similarly, the average within breed molecular kinship varied from 0.496 to 0.745, showing high coancestry, with Skånsk blommehöna having the lowest and Svarthöna the highest coancestry. Furthermore, all breeds showed significant deviations from Hardy-Weinberg expectations. Across the five breeds, the global heterozygosity deficit (FIT was 0.545, population differentiation index (FST was 0.440, and the global inbreeding of individuals within breed (FIS was 0.187. The phylogenetic relationships of chickens were examined using neighbor-joining trees constructed at the level of breeds and individual samples. The neighbor-joining tree constructed at breed level revealed two main clusters, with Hedemorahöna and Öländsk dvärghöna breeds in one cluster, and Gotlandshöna and Svarthöna breeds in the second cluster leaving the Skånsk blommehöna in the middle. Based on the results of the STRUCTURE analysis, the most likely number of clustering of the five breeds was at K = 4, with Hedemorahöna, Gotlandshöna and Svarthöna breeds forming their own distinct clusters, while Öländsk dvärghöna and Skånsk blommehöna breeds clustered together. Losses in the overall genetic diversity of local Swedish chickens due to breeds extinction varied from -1.46% to -6

  19. Diversidade bacteriana da rizosfera de genótipos de milho contrastantes na eficiência de uso de fósforo Bacterial diversity in the rhizosphere of maize genotypes contrasting for phosphorus use efficiency

    Directory of Open Access Journals (Sweden)

    Christiane Abreu de Oliveira

    2009-11-01

    Full Text Available O objetivo deste trabalho foi avaliar a diversidade funcional e genética de bactérias associadas à rizosfera de genótipos de milho contrastantes quanto à eficiência de uso de fósforo, por meio do teste de fontes de carbono no sistema EcoPlate e da eletroforese em gel de gradiente desnaturante (DGGE dos fragmentos amplificados dos genes 16S ribossomais (rDNA das bactérias. Foram coletadas amostras de solo da rizosfera de linhagens e híbridos contrastantes quanto à eficiência de uso de fósforo, cultivados em Latossolo Vermelho-Escuro fase cerrado, com baixo e alto teor de P. Bactérias da rizosfera de híbridos e linhagens eficientes, sob estresse de P, analisadas pelo sistema EcoPlate, tenderam a se agrupar conforme a análise de componentes principais, o que indica que utilizaram fontes de carbono semelhantes. Não houve diferença na diversidade bacteriana, analisada pela DGGE, entre bactérias associadas a genótipos eficientes e ineficientes no uso de P. Com base no sequenciamento do 16S rDNA, foi verificado que a rizosfera de genótipos de milho sob estresse de P parece selecionar grupos específicos de bactérias. A estrutura populacional genética e metabólica de bactérias da rizosfera foi mais influenciada pelo teor de fósforo no solo do que pela eficiência das plantas em usar o fósforo.The objective of this work was to evaluate the functional and genetic diversity of bacteria associated to the rhizosphere of maize genotypes contrasting for phosphorus use efficiency by means of the EcoPlate carbon source test and denaturing gradient gel electrophoresis (DGGE of amplified 16S ribosomal DNA (rDNA fragments of bacteria. Rhizosphere soil samples of maize genotypes (hybrids and lineages contrasting for phosphorus use efficiency cultivated in an Oxisol with high and low P content were collected. Bacteria from the rhizosphere of P-efficient maize genotypes under P stress conditions analyzed by the EcoPlate system tended to group

  20. Genetic and morphological diversity of okra (Abelmoschus esculentus [L.] Moench.) genotypes and their possible relationships, with particular reference to Greek landraces

    NARCIS (Netherlands)

    Kyriakopoulou, O.G.; Arens, P.F.P.; Pelgrom, K.T.B.; Karapanos, I.; Bebeli, P.; Passam, H.C.

    2014-01-01

    Despite its high economic value in many countries (especially in developing regions of the tropics and sub-tropics), okra has received little attention with respect to its source of origin and genetic diversity, particularly at the molecular level. Phenotypic description (morphology, pod

  1. High levels of genetic and genotypic diversity in field populations of the barley pathogen Ramularia collo-cygni

    DEFF Research Database (Denmark)

    Hjortshøj, Rasmus Lund; Ravnshøj, A.R.; Nyman, M.

    2013-01-01

    The ascomycete pathogen Ramularia collo-cygni causes Ramularia leaf spot (RLS) on barley. Although R. collo-cygni is considerd an emerging disease of barley, little is known about genetic diversity or population genetic structure of this pathogen. We applied a set of polymorphic AFLP (Amplified F...

  2. Trait specific expression profiling of salt stress responsive genes in diverse rice genotypes as determined by modified Significance Analysis of Microarrays

    Directory of Open Access Journals (Sweden)

    Mohammad Rashed Hossain

    2016-05-01

    Full Text Available Stress responsive gene expression is commonly profiled in a comparative manner involving different stress conditions or genotypes with contrasting reputation of tolerance/resistance. In contrast, this research exploited a wide natural variation in terms of taxonomy, origin and salt sensitivity in eight genotypes of rice to identify the trait specific patterns of gene expression under salt stress. Genome wide transcptomic responses were interrogated by the weighted continuous morpho-physiological trait responses using modified Significance Analysis of Microarrays. More number of genes was found to be differentially expressed under salt stressed compared to that of under unstressed conditions. Higher numbers of genes were observed to be differentially expressed for the traits shoot Na+/K+, shoot Na+, root K+, biomass and shoot Cl-, respectively. The results identified around sixty genes to be involved in Na+, K+ and anion homeostasis, transport and transmembrane activity under stressed conditions. Gene Ontology (GO enrichment analysis identified 1.36% (578 genes of the entire transcriptome to be involved in the major molecular functions such as signal transduction (>150 genes, transcription factor (81 genes and translation factor activity (62 genes etc. under salt stress. Chromosomal mapping of the genes suggests that majority of the genes are located on chromosomes 1, 2, 3, 6 & 7. The gene network analysis showed that the transcription factors and translation initiation factors formed the major gene networks and are mostly active in nucleus, cytoplasm and mitochondria whereas the membrane and vesicle bound proteins formed a secondary network active in plasma membrane and vacuoles. The novel genes and the genes with unknown functions thus identified provide picture of a synergistic salinity response representing the potentially fundamental mechanisms that are active in the wide natural genetic background of rice and will be of greater use once

  3. Detecting the Diversity of Mycoplasma and Ureaplasma Endosymbionts Hosted by Trichomonas vaginalis Isolates

    Science.gov (United States)

    Ioannidis, Anastasios; Papaioannou, Panagiota; Magiorkinis, Emmanouil; Magana, Maria; Ioannidou, Vasiliki; Tzanetou, Konstantina; Burriel, Angeliki R.; Tsironi, Maria; Chatzipanagiotou, Stylianos

    2017-01-01

    Objectives: The symbiosis of Trichomonas vaginalis and Mycoplasma hominis is the first described association between two obligate human parasites. Trichomonas is the niche and the vector for the transmission of M. hominis infection. This clinically significant symbiosis may affect T. vaginalis virulence and susceptibility to treatment. The aims of this study were to investigate the intracellularly present Mycoplasma and Ureaplasma species in T. vaginalis strains isolated from the vaginal discharge of infected women as well as to trace the diversity pattern among the species detected in the isolated strains. Methods: Hundred pure T. vaginalis cultures were isolated from ~7,500 patient specimens presented with clinical purulent vaginitis. PCR and sequencing for Mycoplasma/Ureaplasma spp. were performed in DNA extracted from the pure cultures. In addition, vaginal discharge samples were cultured for the presence of M. hominis and U. urealyticum. Phylogenetic analysis assisted the identification of interspecies relationships between the Mycoplasma and Ureaplasma isolates. Results: Fifty four percentage of T. vaginalis isolates were harboring Mycoplasma spp. Phylogenetic analysis revealed three distinct clusters, two with already characterized M. hominis and Ureaplasma spp. (37% of total Mycoplasma spp.), whereas one group formed a distinct cluster matched with the newly identified species Candidatus Mycoplasma girerdii (59.3%) and one or more unknown Mycoplasma spp. (3.7%). Conclusions: T. vaginalis strains associated with vaginal infection might host intracellular mycoplasmas or ureaplasmas. Intracellular Mollicutes that remain undetected in the extracellular environment when conventional diagnostic methods are implemented may comprise either novel species, such as Candidatus M. giredii, or unknown species with yet unexplored clinical significance. PMID:28702014

  4. Detecting the Diversity of Mycoplasma and Ureaplasma Endosymbionts Hosted by Trichomonas vaginalis Isolates

    Directory of Open Access Journals (Sweden)

    Anastasios Ioannidis

    2017-06-01

    Full Text Available Objectives: The symbiosis of Trichomonas vaginalis and Mycoplasma hominis is the first described association between two obligate human parasites. Trichomonas is the niche and the vector for the transmission of M. hominis infection. This clinically significant symbiosis may affect T. vaginalis virulence and susceptibility to treatment. The aims of this study were to investigate the intracellularly present Mycoplasma and Ureaplasma species in T. vaginalis strains isolated from the vaginal discharge of infected women as well as to trace the diversity pattern among the species detected in the isolated strains.Methods: Hundred pure T. vaginalis cultures were isolated from ~7,500 patient specimens presented with clinical purulent vaginitis. PCR and sequencing for Mycoplasma/Ureaplasma spp. were performed in DNA extracted from the pure cultures. In addition, vaginal discharge samples were cultured for the presence of M. hominis and U. urealyticum. Phylogenetic analysis assisted the identification of interspecies relationships between the Mycoplasma and Ureaplasma isolates.Results: Fifty four percentage of T. vaginalis isolates were harboring Mycoplasma spp. Phylogenetic analysis revealed three distinct clusters, two with already characterized M. hominis and Ureaplasma spp. (37% of total Mycoplasma spp., whereas one group formed a distinct cluster matched with the newly identified species Candidatus Mycoplasma girerdii (59.3% and one or more unknown Mycoplasma spp. (3.7%.Conclusions:T. vaginalis strains associated with vaginal infection might host intracellular mycoplasmas or ureaplasmas. Intracellular Mollicutes that remain undetected in the extracellular environment when conventional diagnostic methods are implemented may comprise either novel species, such as Candidatus M. giredii, or unknown species with yet unexplored clinical significance.

  5. Genetic and DNA methylation changes in cotton (Gossypium genotypes and tissues.

    Directory of Open Access Journals (Sweden)

    Kenji Osabe

    Full Text Available In plants, epigenetic regulation is important in normal development and in modulating some agronomic traits. The potential contribution of DNA methylation mediated gene regulation to phenotypic diversity and development in cotton was investigated between cotton genotypes and various tissues. DNA methylation diversity, genetic diversity, and changes in methylation context were investigated using methylation-sensitive amplified polymorphism (MSAP assays including a methylation insensitive enzyme (BsiSI, and the total DNA methylation level was measured by high-performance liquid chromatography (HPLC. DNA methylation diversity was greater than the genetic diversity in the selected cotton genotypes and significantly different levels of DNA methylation were identified between tissues, including fibre. The higher DNA methylation diversity (CHG methylation being more diverse than CG methylation in cotton genotypes suggest epigenetic regulation may be important for cotton, and the change in DNA methylation between fibre and other tissues hints that some genes may be epigenetically regulated for fibre development. The novel approach using BsiSI allowed direct comparison between genetic and epigenetic diversity, and also measured CC methylation level that cannot be detected by conventional MSAP.

  6. Genetic and DNA methylation changes in cotton (Gossypium) genotypes and tissues.

    Science.gov (United States)

    Osabe, Kenji; Clement, Jenny D; Bedon, Frank; Pettolino, Filomena A; Ziolkowski, Lisa; Llewellyn, Danny J; Finnegan, E Jean; Wilson, Iain W

    2014-01-01

    In plants, epigenetic regulation is important in normal development and in modulating some agronomic traits. The potential contribution of DNA methylation mediated gene regulation to phenotypic diversity and development in cotton was investigated between cotton genotypes and various tissues. DNA methylation diversity, genetic diversity, and changes in methylation context were investigated using methylation-sensitive amplified polymorphism (MSAP) assays including a methylation insensitive enzyme (BsiSI), and the total DNA methylation level was measured by high-performance liquid chromatography (HPLC). DNA methylation diversity was greater than the genetic diversity in the selected cotton genotypes and significantly different levels of DNA methylation were identified between tissues, including fibre. The higher DNA methylation diversity (CHG methylation being more diverse than CG methylation) in cotton genotypes suggest epigenetic regulation may be important for cotton, and the change in DNA methylation between fibre and other tissues hints that some genes may be epigenetically regulated for fibre development. The novel approach using BsiSI allowed direct comparison between genetic and epigenetic diversity, and also measured CC methylation level that cannot be detected by conventional MSAP.

  7. Performance Evaluation of Manual and Automated (MagNA Pure Nucleic Acid Isolation in HPV Detection and Genotyping Using Roche Linear Array HPV Test

    Directory of Open Access Journals (Sweden)

    Aikaterini Chranioti

    2011-01-01

    Full Text Available Nucleic acids of human papillomavirus (HPV isolated by manual extraction method (AmpliLute and automated MagNA pure system were compared and evaluated with cytohistological findings in 253 women. The concordance level between AmpliLute and MagNA was very good 93.3% (=0.864, <.0001. Overall HPVpositivity detected by AmpliLute was 57.3% (30.4% as single and 27% as multiple infections in contrast to MagNA 54.5% (32% and 23%, resp.. Discrepant results observed in 25 cases: 11 MagNA(−/AmpliLute(+, 10 of which had positive histology; 5 MagNA(+/AmpliLute(− with negative histology; 8 MagNA(+/AmpliLute(+: in 7 of which AmpliLute detected extra HPV genotypes and 1 MagNA(invalid/AmpliLute(+ with positive histology. Both methods performed well when compared against cytological (area under curve (AUC of AmpliLute 0.712 versus 0.672 of MagNA and histological diagnoses (AUC of AmpliLute 0.935 versus 0.877 of MagNA, with AmpliLute showing a slightly predominance over MagNA. However, higher sensitivities, specificities, and positive/negative predictive values were obtained by AmpliLute.

  8. Improved detection of CXCR4-using HIV by V3 genotyping: application of population-based and "deep" sequencing to plasma RNA and proviral DNA.

    Science.gov (United States)

    Swenson, Luke C; Moores, Andrew; Low, Andrew J; Thielen, Alexander; Dong, Winnie; Woods, Conan; Jensen, Mark A; Wynhoven, Brian; Chan, Dennison; Glascock, Christopher; Harrigan, P Richard

    2010-08-01

    Tropism testing should rule out CXCR4-using HIV before treatment with CCR5 antagonists. Currently, the recombinant phenotypic Trofile assay (Monogram) is most widely utilized; however, genotypic tests may represent alternative methods. Independent triplicate amplifications of the HIV gp120 V3 region were made from either plasma HIV RNA or proviral DNA. These underwent standard, population-based sequencing with an ABI3730 (RNA n = 63; DNA n = 40), or "deep" sequencing with a Roche/454 Genome Sequencer-FLX (RNA n = 12; DNA n = 12). Position-specific scoring matrices (PSSMX4/R5) (-6.96 cutoff) and geno2pheno[coreceptor] (5% false-positive rate) inferred tropism from V3 sequence. These methods were then independently validated with a separate, blinded dataset (n = 278) of screening samples from the maraviroc MOTIVATE trials. Standard sequencing of HIV RNA with PSSM yielded 69% sensitivity and 91% specificity, relative to Trofile. The validation dataset gave 75% sensitivity and 83% specificity. Proviral DNA plus PSSM gave 77% sensitivity and 71% specificity. "Deep" sequencing of HIV RNA detected >2% inferred-CXCR4-using virus in 8/8 samples called non-R5 by Trofile, and <2% in 4/4 samples called R5. Triplicate analyses of V3 standard sequence data detect greater proportions of CXCR4-using samples than previously achieved. Sequencing proviral DNA and "deep" V3 sequencing may also be useful tools for assessing tropism.

  9. Genetic diversity in maize genotypes with and without a topdressing of nitrogen fertilizer = Divergência genética de genótipos de milho com e sem adubação nitrogenada em cobertura

    Directory of Open Access Journals (Sweden)

    Karen Cristina Leite Silva

    2015-06-01

    Full Text Available The use of genetic diversity as a basis for identifying combinations which are superior to the parents, with a greater heterozygosity, is important in view of the difficulty when selecting promising genotypes for a breeding program. Given the above, the aim of this work was to evaluate genetic diversity in maize genotypes with and without a topdressing of nitrogen fertiliser, using characteristics of the growth stage of the crop. Two field experiments were carried out in Gurupi, in the south of the state of Tocantins, Brazil (TO, one with and another without a topdressing of N fertilizer (1 - no N topdressing, 2 - 150 kg N ha-1. The treatments consisted of 12 genotypes (six open-pollinated populations, and six S5 strains. In applying the technique of clustering to the genotypes, the Generalised Mahalanobis Distance (D2 was adopted as dissimilarity measure. To establish similar groups, the agglomerative hierarchical method of optimisation proposed by Tocher was applied. In addition, Singh’s criterion was used to quantify the relative contribution to genetic divergence of the characteristics under evaluation. The characteristics, Chlorophyll-a and total chlorophyll, displayed the greatest contribution to genetic divergence, when there was no topdressing of nitrogen fertiliser and with the use of 150 kg N ha-1 respectively. A topdressing of nitrogen influenced both the vegetative development of the genotypes, and the expression of their genetic variability. = A utilização da divergência genética como base para a identificação de combinações superiores aos progenitores, apresentando maior heterozigose, faz-se importante diante da dificuldade de escolha de genótipos promissores em um programa de melhoramento. Com base no exposto, objetivou-se com este trabalho avaliar a divergência genética de genótipos de milho com e sem adubação nitrogenada em cobertura, utilizando características do estágio vegetativo da cultura, no sul do Estado

  10. The use of microsatellite markers for genetic diversity assessment of ...

    African Journals Online (AJOL)

    In this study, gene diversity and genetic relationships among 30 genotypes of genus Hordeum from Kerman province (Iran) were assessed using 10 simple sequence repeat (SSR) primers. Seven of these markers were highly polymorphic. A total of 96 alleles were detected. The number of alleles per microsatellite marker ...

  11. Examination of food chain-derived Listeria monocytogenes strains of different serotypes reveals considerable diversity in inlA genotypes, mutability, and adaptation to cold temperatures.

    Science.gov (United States)

    Kovacevic, Jovana; Arguedas-Villa, Carolina; Wozniak, Anna; Tasara, Taurai; Allen, Kevin J

    2013-03-01

    Listeria monocytogenes strains belonging to serotypes 1/2a and 4b are frequently linked to listeriosis. While inlA mutations leading to premature stop codons (PMSCs) and attenuated virulence are common in 1/2a, they are rare in serotype 4b. We observed PMSCs in 35% of L. monocytogenes isolates (n = 54) recovered from the British Columbia food supply, including serotypes 1/2a (30%), 1/2c (100%), and 3a (100%), and a 3-codon deletion (amino acid positions 738 to 740) seen in 57% of 4b isolates from fish-processing facilities. Caco-2 invasion assays showed that two isolates with the deletion were significantly more invasive than EGD-SmR (P cold temperature following a downshift from 37°C to 4°C. Overall, three distinct cold-adapting groups (CAG) were observed: 46% were fast (200 h) adaptors. Intermediate CAG strains (70%) more frequently possessed inlA PMSCs than did fast (20%) and slow (10%) CAGs; in contrast, 87% of fast adaptors lacked inlA PMSCs. In conclusion, we report food chain-derived 1/2a and 4b serotypes with a 3-codon deletion possessing invasive behavior and the novel association of inlA genotypes encoding a full-length InlA with fast cold-adaptation phenotypes.

  12. Role of static fluid MR urography in detecting post urinary diversion ...

    African Journals Online (AJOL)

    Amr Farouk Ibrahim Moustafa

    mon surgical procedures included Ileal conduit & Ileocecal neobladder diversion. Material & methods: ... tory urograms giving an overview of the urinary tract & quickly ..... A secondary fistula was seen in 1 patient where fistulous track occurred ...

  13. A newly developed real-time PCR assay for detection and quantification of Fusarium oxysporum and its use in compatible and incompatible interactions with grafted melon genotypes.

    Science.gov (United States)

    Haegi, Anita; Catalano, Valentina; Luongo, Laura; Vitale, Salvatore; Scotton, Michele; Ficcadenti, Nadia; Belisario, Alessandra

    2013-08-01

    A reliable and species-specific real-time quantitative polymerase chain reaction (qPCR) assay was developed for detection of the complex soilborne anamorphic fungus Fusarium oxysporum. The new primer pair, designed on the translation elongation factor 1-α gene with an amplicon of 142 bp, was highly specific to F. oxysporum without cross reactions with other Fusarium spp. The protocol was applied to grafted melon plants for the detection and quantification of F. oxysporum f. sp. melonis, a devastating pathogen of this cucurbit. Grafting technologies are widely used in melon to confer resistance against new virulent races of F. oxysporum f. sp. melonis, while maintaining the properties of valuable commercial varieties. However, the effects on the vascular pathogen colonization have not been fully investigated. Analyses were performed on 'Charentais-T' (susceptible) and 'Nad-1' (resistant) melon cultivars, both used either as rootstock and scion, and inoculated with F. oxysporum f. sp. melonis race 1 and race 1,2. Pathogen development was compared using qPCR and isolations from stem tissues. Early asymptomatic melon infections were detected with a quantification limit of 1 pg of fungal DNA. The qPCR protocol clearly showed that fungal development was highly affected by host-pathogen interaction (compatible or incompatible) and time (days postinoculation). The principal significant effect (P ≤ 0.01) on fungal development was due to the melon genotype used as rootstock, and this effect had a significant interaction with time and F. oxysporum f. sp. melonis race. In particular, the amount of race 1,2 DNA was significantly higher compared with that estimated for race 1 in the incompatible interaction at 18 days postinoculation. The two fungal races were always present in both the rootstock and scion of grafted plants in either the compatible or incompatible interaction.

  14. Prevalence of ESBL-producing Pseudomonas aeruginosa isolates in Warsaw, Poland, detected by various phenotypic and genotypic methods.

    Directory of Open Access Journals (Sweden)

    Agnieszka E Laudy

    Full Text Available Knowledge of the prevalence of ESBL enzymes among P. aeruginosa strains compared to the Enterobacteraiceae family is limited. The phenotypic tests recommended by EUCAST for the detection of ESBL-producing Enterobacteriaceae are not always suited for P. aeruginosa strains. This is mainly due to the presence of other families of ESBLs in P. aeruginosa isolates more often than in Enterobacteriaceae, production of natural AmpC cephalosporinase and its overexpression, and co-production of metallo-β-lactamases. The aim of this study was to determine the occurrence of ESBLs in P. aeruginosa isolated from patients from hospitals in Warsaw, to evaluate the ESBL production of these isolates using currently available phenotypic tests, their modifications, multiplex PCR and molecular typing of ESBL-positive isolates by PFGE. Clinical isolates of P. aeruginosa were collected in 2000-2014 from four Warsaw hospitals. Based on the data obtained in this study, we suggest using three DDST methods with inhibitors, such as clavulanic acid, sulbactam and imipenem, to detect ESBL-producing P. aeruginosa strains. Depending on the appearance of the plates, we suggest a reduction in the distance between discs with antibiotics to 15 mm and the addition of boronic acid at 0.4 mg per disc. The analysed isolates carried genes encoding ESBL from the families VEB (69 isolates with VEB-9, GES (6 with GES-1, 1 GES-5, 5 GES-13 and 2 with GES-15, OXA-2 (12 with OXA-15, 1 OXA-141, 1 OXA-210, 1 OXA-543 and 1 with OXA-544 and OXA-10 (5 isolates with OXA-74 and one with OXA-142. The most important result of this study was the discovery of three new genes, blaGES-15, blaOXA-141 and blaOXA-142; their nucleotide sequences have been submitted to the NCBI GenBank. It is also very important to note that this is the first report on the epidemiological problem of VEB-9-producing bacterial strains, not only in Poland but also worldwide.

  15. Diversity of Archaea and detection of crenarchaeotal amoA genes in the rivers Rhine and Têt

    OpenAIRE

    Herfort, L.; Kim, J.H.; Coolen, M.J.L.; Abbas, B.; Schouten, S.; Herndl, G.J.; Sinninghe Damste, J.S.

    2009-01-01

    Pelagic archaeal phylogenetic diversity and the potential for crenarchaeotal nitrification of Group 1.1a were determined in the rivers Rhine and Têt by 16S rRNA sequencing, catalyzed reported deposition-fluorescence in situ hybridization (CARD–FISH) and quantification of 16S rRNA and functional genes. Euryarchaeota were, for the first time, detected in temperate river water even though a net predominance of crenarchaeotal phylotypes was found. Differences in phylogenic distribution were obser...

  16. Genotyping of friesian horses to detect a hydrocephalus-associated c.1423C>T mutation in B3GALNT2 using PCR-RFLP and PCR-PIRA methods: Frequency in stallion horses in México.

    Science.gov (United States)

    Ayala-Valdovinos, Miguel Angel; Galindo-García, Jorge; Sánchez-Chiprés, David; Duifhuis-Rivera, Theodor

    2017-04-01

    Hydrocephalus in Friesian horses is an autosomal recessive hereditary disease that can result in an abortion, a stillbirth, or euthanization of a newborn foal. Here, the hydrocephalus-associated c.1423C > T mutation in B3GALNT2 gene was detected with PCR-RFLP and PCR-PIRA methods for horse genotyping. A preliminary genotyping survey was performed on 83 randomly selected Friesian stallion horses to determine the current allele frequency in Mexico. The frequency of the mutant T allele was 9.6%. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. Phylogenetic Diversity in Core Region of Hepatitis C Virus Genotype 1a as a Factor Associated with Fibrosis Severity in HIV-1-Coinfected Patients

    Directory of Open Access Journals (Sweden)

    Micaela Parra

    2017-01-01

    Full Text Available High hepatitis C virus (HCV genetic diversity impacts infectivity/pathogenicity, influencing chronic liver disease progression associated with fibrosis degrees and hepatocellular carcinoma. HCV core protein is crucial in cell-growth regulation and host-gene expression. Liver fibrosis is accelerated by unknown mechanisms in human immunodeficiency virus-1- (HIV-1- coinfected individuals. We aimed to study whether well-defined HCV-1a core polymorphisms and genetic heterogeneity are related to fibrosis in a highly homogeneous group of interferon-treated HIV-HCV-coinfected patients. Genetic heterogeneity was weighed by Faith’s phylogenetic diversity (PD, which has been little studied in HCV. Eighteen HCV/HIV-coinfected patients presenting different liver fibrosis stages before anti-HCV treatment-initiation were recruited. Sampling at baseline and during and after treatment was performed up to 72 weeks. At inter/intrahost level, HCV-1a populations were studied using molecular cloning and Sanger sequencing. Over 400 complete HCV-1a core sequences encompassing 573 positions of C were obtained. Amino acid substitutions found previously at positions 70 and 91 of HCV-1b core region were not observed. However, HCV genetic heterogeneity was higher in mild than in severe fibrosis cases. These results suggest a potential utility of PD as a virus-related factor associated with chronic hepatitis C progression. These observations should be reassessed in larger cohorts to corroborate our findings and assess other potential covariates.

  18. American foulbrood of the honey bee: occurrence and distribution of different genotypes of Paenibacillus larvae in the administrative district of Arnsberg (North Rhine-Westphalia).

    Science.gov (United States)

    Peters, M; Kilwinski, J; Beringhoff, A; Reckling, D; Genersch, E

    2006-03-01

    Between March 2003 and October 2004, Paenibacillus larvae, the aetiological agent of American foulbrood disease of the honey bee, was isolated from broodcombs and honey samples of 54 apiaries in the administrative district of Arnsberg (North Rhine-Westphalia, Germany). Genotyping of 176 P. larvae isolates with repetitive element polymerase chain reaction fingerprinting (rep-PCR) using BOX A1R and MBO REP1 primers revealed five different genotypes (AB, Ab, ab, ass, Acapital BE, Cyrillic). In samples of three apiaries, more than one genotype was detected. A combination of two genotypes was isolated from honey samples of the same hive two times (ab/ass and Ab/ab). The five genotypes were not randomly distributed in the district, but revealed a certain geographical clustering. Possible factors with impact on the genotype diversity and the distribution pattern are discussed.

  19. Detection of porcine circovirus genotypes 2a and 2b in aborted foetuses from infected swine herds in the State of São Paulo, Brazil

    Directory of Open Access Journals (Sweden)

    de Castro Alessandra MMG

    2012-05-01

    Full Text Available Abstract Background Porcine circovirus type 2 (PCV2 has been associated with several disease complexes, including reproductive failure. The aim of this study was to identify the subtypes of PCV2 that are associated with reproductive failure in pigs from the State of São Paulo, Brazil and to investigate co-infections with other infectious organisms. Findings Samples of 168 aborted foetuses or mummified foetuses from five farrow-to-finish swine farms known to be infected with PCV2 and located in the State of São Paulo were tested for PCV2 by polymerase chain reaction (PCR. Positive samples were additionally tested for porcine parvovirus (PPV, Leptospira spp. and Brucella spp. by PCR. PCV2 was detected in 18 of the samples (10.7%. PPV, Brucella spp. and Leptospira spp were found in 2, 10 and 0 cases, respectively. Eleven PCV2 strains were sequenced and determined to be either genotype 2a (n = 1 or 2b (n = 10. Conclusions The findings indicate that the frequency of PCV2 infections in aborted porcine foetuses from the State of São Paulo is rather low (10.7% and that co-infection with other pathogens is common and may be involved in PCV2 associated reproductive failure. No repeatable, characteristic amino acid motifs for regions of the PCV2 capsid protein seemed to be associated with abortion in sows.

  20. Mapping epistasis and environment × QTX interaction based on four -omics genotypes for the detected QTX loci controlling complex traits in tobacco

    Directory of Open Access Journals (Sweden)

    Liyuan Zhou

    2013-12-01

    Full Text Available Using newly developed methods and software, association mapping was conducted for chromium content and total sugar in tobacco leaf, based on four -omics datasets. Our objective was to collect data on genotype and phenotype for 60 leaf samples at four developmental stages, from three plant architectural positions and for three cultivars that were grown in two locations. Association mapping was conducted to detect genetic variants at quantitative trait SNP (QTS loci, quantitative trait transcript (QTT differences, quantitative trait protein (QTP variability, and quantitative trait metabolite (QTM changes, which can be summarized as QTX locus variation. The total heritabilities of the four -omics loci for both traits tested were 23.60% for epistasis and 15.26% for treatment interaction. Epistasis and environment × treatment interaction had important impacts on complex traits at all -omics levels. For decreasing chromium content and increasing total sugar in tobacco leaf, six methylated loci can be directly used for marker-assisted selection, and expression of ten QTTs, seven QTPs and six QTMs can be modified by selection or cultivation.

  1. Detection of uncommon G3P[3] rotavirus A (RVA) strain in rat possessing a human RVA-like VP6 and a novel NSP2 genotype.

    Science.gov (United States)

    Ianiro, Giovanni; Di Bartolo, Ilaria; De Sabato, Luca; Pampiglione, Guglielmo; Ruggeri, Franco M; Ostanello, Fabio

    2017-09-01

    Rotavirus is one of the leading causes of acute gastroenteritis in infants and young children. RVAs infect not only humans but also a wide range of mammals including rats, which represent a reservoir of several other zoonotic pathogens. Due to the segmented nature of the RVA genome, animal RVA strains can easily adapt to the human host by reassortment with co-infecting human viruses. This study aims to detect and characterize RVA in the intestinal content of Italian sinantropic rats (Rattus rattus). Out of 40 samples examined following molecular approach, one resulted positive for RVA. The molecular characterization of VP1-4, 6 and 7, and NSP1-5 genes by sequencing revealed the genomic constellation G3-P[3]-I1-R11-C11-M10-A22-N18-T14-E18-H13. This uncommon genomic combination includes: the VP1-4,VP7, the NSP1, 3, 4 and 5 gene segments, closely related to those of RVA from rodents, the N18 novel genotype established for the NSP2 gene segment and the human Wa-like VP6 gene, suggesting interspecies reassortment. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. Targeted genomic enrichment and sequencing of CyHV-3 from carp tissues confirms low nucleotide diversity and mixed genotype infections

    Directory of Open Access Journals (Sweden)

    Saliha Hammoumi

    2016-09-01

    Full Text Available Koi herpesvirus disease (KHVD is an emerging disease that causes mass mortality in koi and common carp, Cyprinus carpio L. Its causative agent is Cyprinid herpesvirus 3 (CyHV-3, also known as koi herpesvirus (KHV. Although data on the pathogenesis of this deadly virus is relatively abundant in the literature, still little is known about its genomic diversity and about the molecular mechanisms that lead to such a high virulence. In this context, we developed a new strategy for sequencing full-length CyHV-3 genomes directly from infected fish tissues. Total genomic DNA extracted from carp gill tissue was specifically enriched with CyHV-3 sequences through hybridization to a set of nearly 2 million overlapping probes designed to cover the entire genome length, using KHV-J sequence (GenBank accession number AP008984 as reference. Applied to 7 CyHV-3 specimens from Poland and Indonesia, this targeted genomic enrichment enabled recovery of the full genomes with >99.9% reference coverage. The enrichment rate was directly correlated to the estimated number of viral copies contained in the DNA extracts used for library preparation, which varied between ∼5000 and ∼2×107. The average sequencing depth was >200 for all samples, thus allowing the search for variants with high confidence. Sequence analyses highlighted a significant proportion of intra-specimen sequence heterogeneity, suggesting the presence of mixed infections in all investigated fish. They also showed that inter-specimen genetic diversity at the genome scale was very low (>99.95% of sequence identity. By enabling full genome comparisons directly from infected fish tissues, this new method will be valuable to trace outbreaks rapidly and at a reasonable cost, and in turn to understand the transmission routes of CyHV-3.

  3. Determining rejection regions and power that optimize the inspector's chances of detecting diversion and/or falsification

    International Nuclear Information System (INIS)

    Goldman, A.S.; Whitty, W.J.; Hafer, J.F.; Markin, J.T.; Shipley, J.P.

    1981-01-01

    Three specific models are investigated: (1) diversion and k distinct falsifications (k + 1 degrees of freedom); (2) diversion and the accumulation of all the falsifications (2 degrees of freedom); (3) diversion only (1 degree of freedom). A test statistic has been derived for models (1) and (2) by the likelihood ratio procedure under the hypothesis of zero falsification and zero diversion versus a one-sided alternative of positive diversion, positive falsification, or both. An analogous test has been developed for model (3) for diversion only. A detailed discussion of this development is given by Shipley. J.P. Shipley, Notes for the Seminar on Data Generation and Evaluation for Safeguards, Los Alamos National Laboratory Seminar, Los Alamos, New Mexico, July 7-9, 1980 (to be published). Utilizing his notation, the test variables called Inspector's Sufficient Statistics - ISS 0 , ISS 1 , and ISS 2 - are used for testing the above three models (1), (2), and (3) respectively. Note that ISS 2 corresponds to the well-known MUF-D statistic, which is currently in vogue. These two tests, for purposes of this discussion, are equivalent. The objectives of this paper are to review the ISS procedures, develop optimal critical regions for ISS tests, and compare detection probability (power) curves for models (1), (2), and (3). Optimal critical regions depend upon values obtained from one-sided chi-squared tests with more than one degree of freedom. Details of such a procedure are not to be found in the literature, and this strange neglect was one of the motivating factors in this study

  4. TEST KIT FOR THE DETECTION AND GENOTYPING OF HIGHLY PATHOGENIC INFLUENZA VIRUS A H5N1 BY REAL-TIME POLYMERASE CHAIN REACTION

    Directory of Open Access Journals (Sweden)

    S. V. Stepaniuk

    2014-06-01

    Full Text Available Results of the annual monitoring of epizooties indicate that highly pathogenic HPAI/H5N1 avian influenza widely circulated in Eurasian region. Over a period of 2010–2013 years more than 165 cases of outbreaks in 14 countries were found out. Ukraine became one of the first countries in Europe where in Autonomous Republic of Crimea in October 2005 outbreak of avian epizootic with HPAI/H5N1 was documented and until February 2008 more than 236,000 poultry were killed. Since then the question of monitoring of infected both migrating birds and poultry in places of cross contact in Ukraine remains of high priority. The test system is developed for identification and genotyping A H5N1 on three genes (M, H5 and N1 HPAI/H5N1 in real-time mode for polymerase chain reaction. Test kit capacity to detect HPAI/h5n1avian influenza virus and differentiate it from the other viral infection agents of birds and animals were studied by testing of HPAI/H5N1 virus isolated during mass infection outbreak in Crimea in 2005 and cultural specimens of other viral pathogens. It was established that the «DIA Real Avian Influenza» test kit was capable to detect RNA influenza A virus of high pathogenic H5N1 strains having high sensitivity (100% while RNA of the Crimean HPAI/H5N1 isolate studying and specificity (100% while RNA viruses of Newcastle birds disease, fowl powershift, syndrome of drop in egg production and horse influenza studying.

  5. Comprehensive detection of diverse exon 19 deletion mutations of EGFR in lung Cancer by a single probe set.

    Science.gov (United States)

    Bae, Jin Ho; Jo, Seong-Min; Kim, Hak-Sung

    2015-12-15

    Detection of exon 19 deletion mutation of EGFR, one of the most frequently occurring mutations in lung cancer, provides the crucial information for diagnosis and treatment guideline in non-small-cell lung cancer (NSCLC). Here, we demonstrate a simple and efficient method to detect various exon 19 deletion mutations of EGFR using a single probe set comprising of an oligo-quencher (oligo-Q) and a molecular beacon (MB). While the MB hybridizes to both the wild and mutant target DNA, the oligo-Q only binds to the wild target DNA, leading to a fluorescent signal in case of deletion mutation. This enables the comprehensive detection of the diverse exon 19 deletion mutations using a single probe set. We demonstrated the utility and efficiency of the approach by detecting the frequent exon 19 deletion mutations of EGFR through a real-time PCR and in situ fluorescence imaging. Our approach enabled the detection of genomic DNA as low as 0.02 ng, showing a detection limit of 2% in a heterogeneous DNA mixture, and could be used for detecting mutations in a single cell level. The present MB and oligo-Q dual probe system can be used for diagnosis and treatment guideline in NSCLC. Copyright © 2015 Elsevier B.V. All rights reserved.

  6. Distribution and factors associated with Salmonella enterica genotypes in a diverse population of humans and animals in Qatar using multi-locus sequence typing (MLST).

    Science.gov (United States)

    Chang, Yu C; Scaria, Joy; Ibraham, Mariamma; Doiphode, Sanjay; Chang, Yung-Fu; Sultan, Ali; Mohammed, Hussni O

    2016-01-01

    Salmonella enterica is one of the most commonly reported causes of bacterial foodborne illness around the world. Understanding the sources of this pathogen and the associated factors that exacerbate its risk to humans will help in developing risk mitigation strategies. The genetic relatedness among Salmonella isolates recovered from human gastroenteritis cases and food animals in Qatar were investigated in the hope of shedding light on these sources, their possible transmission routes, and any associated factors. A repeat cross-sectional study was conducted in which the samples and associated data were collected from both populations (gastroenteritis cases and animals). Salmonella isolates were initially analyzed using multi-locus sequence typing (MLST) to investigate the genetic diversity and clonality. The relatedness among the isolates was assessed using the minimum spanning tree (MST). Twenty-seven different sequence types (STs) were identified in this study; among them, seven were novel, including ST1695, ST1696, ST1697, ST1698, ST1699, ST1702, and ST1703. The pattern of overall ST distribution was diverse; in particular, it was revealed that ST11 and ST19 were the most common sequence types, presenting 29.5% and 11.5% within the whole population. In addition, 20 eBurst Groups (eBGs) were identified in our data, which indicates that ST11 and ST19 belonged to eBG4 and eBG1, respectively. In addition, the potential association between the putative risk factors and eBGs were evaluated. There was no significant clustering of these eBGs by season; however, a significant association was identified in terms of nationality in that Qataris were six times more likely to present with eBG1 compared to non-Qataris. In the MST analysis, four major clusters were presented, namely, ST11, ST19, ST16, and ST31. The linkages between the clusters alluded to a possible transmission route. The results of the study have provided insight into the ST distributions of S. enterica and

  7. Low-Complexity Full-Diversity Detection in Two-User MIMO X Channels

    KAUST Repository

    Ismail, Amr

    2014-01-26

    Several interference cancellation (IC) schemes have been recently proposed to suppress multi-user interference for various network configurations (e.g., multiple access and X channels). However, most of these schemes trade-off diversity for implementation complexity or vice-versa. In this paper, we propose a full-diversity interference cancellation scheme in a multiple-input multiple-output (MIMO) X channel with two sources and two destinations while maintaining low decoding complexity. We provide sufficient conditions for a wide range of space-time block codes (STBCs) to achieve full-diversity gain under the so-called partial interference cancellation group decoding (PICGD) in the configuration of interest. A systematic construction is then proposed to achieve full-diversity. The constructed scheme is compared to recently proposed IC scheme in terms of performance and decoding complexity. Our IC scheme outperforms the recently proposed scheme in the case it provides higher transmission rate, while it loses slightly in the case of equal rates. In terms of decoding complexity, both schemes are equivalent.

  8. On low-complexity full-diversity detection of multi-user space-time coding

    KAUST Repository

    Ismail, Amr

    2013-06-01

    The incorporation of multiple input multiple output (MIMO) schemes in recent wireless communication standards paved the way to exploit the newly introduced dimension (i.e. space) to efficiently cancel the interference without requiring additional resources. In this paper, we focus on multiple input multiple ouitput (MIMO) multiple access channel (MAC) case and we answer the question about whether it is possible to suppress the interference in a MIMO MAC channel for completely blind users while achieving full-diversity with a simplified decoder in the affirmative. In fact, this goal can be attained by employing space-time block codes (STBC)s that achive full-diversity under partial interference cancellation (PIC). We derive sufficient conditions for a wide range of STBCs to achieve full-diversity under PIC group decoding with or without successive interference cancellation (SIC). Based on the provided design criteria we derive an upper-bound on the achievable rate for a class of codes. A two-user MIMO MAC interference cancellation scheme is presented and proved to achieve full-diversity under PIC group decoding. We compare our scheme to existing beamforming schemes with full or limit feedback. © 2013 IEEE.

  9. On low-complexity full-diversity detection of multi-user space-time coding

    KAUST Repository

    Ismail, Amr; Alouini, Mohamed-Slim

    2013-01-01

    for a wide range of STBCs to achieve full-diversity under PIC group decoding with or without successive interference cancellation (SIC). Based on the provided design criteria we derive an upper-bound on the achievable rate for a class of codes. A two

  10. Low-Complexity Full-Diversity Detection in Two-User MIMO X Channels

    KAUST Repository

    Ismail, Amr; Abediseid, Walid; Alouini, Mohamed-Slim

    2014-01-01

    Several interference cancellation (IC) schemes have been recently proposed to suppress multi-user interference for various network configurations (e.g., multiple access and X channels). However, most of these schemes trade-off diversity for implementation complexity or vice-versa. In this paper, we propose a full-diversity interference cancellation scheme in a multiple-input multiple-output (MIMO) X channel with two sources and two destinations while maintaining low decoding complexity. We provide sufficient conditions for a wide range of space-time block codes (STBCs) to achieve full-diversity gain under the so-called partial interference cancellation group decoding (PICGD) in the configuration of interest. A systematic construction is then proposed to achieve full-diversity. The constructed scheme is compared to recently proposed IC scheme in terms of performance and decoding complexity. Our IC scheme outperforms the recently proposed scheme in the case it provides higher transmission rate, while it loses slightly in the case of equal rates. In terms of decoding complexity, both schemes are equivalent.

  11. Diversity of group A rotavirus genes detected in the Triângulo Mineiro region, Minas Gerais, Brazil

    Directory of Open Access Journals (Sweden)

    Ana Carolina Bernardes Dulgheroff

    Full Text Available ABSTRACT Group A rotaviruses are the main causative agent of infantile gastroenteritis. The segmented nature of the viral genome allows reassortment of genome segments, which can generate genetic variants. In this study, we characterized the diversity of the VP7, VP4 (VP8*, VP6, NSP4, and NSP5 genes of the rotaviruses that circulated from 2005 to 2011 in the Triângulo Mineiro (TM region of Brazil. Samples with genotypes G2 (sublineages IVa-1 and IVa-3, G1 (sublineage I-A, G9 (lineage III, G12 (lineages II and III, G8 (lineage II, G3 (lineage III, P[4] (sublineages IVa and IVb, P[8] (sublineages P[8]-3.6, P[8]-3.3, and P[8]-3.1, I2 (lineage VII, E2 (lineages VI, XII, and X, and H2 (lineage III were identified. The associations found in the samples were G1, G9, or G12 with P[8]-I1-E1-H1; G2 or G8 with P[4]-I2-E2-H2; G12 with I3-E3-H6; and G3 with P[4]-I2-E3-H3 (previously unreported combination. Reassortment events in G2P[4] strains and an apparent pattern of temporal segregation within the lineages were observed. Five TM samples contained genes that exhibited high nucleotide and amino acid identities with strains of animal origin. The present study includes a period of pre- and post-introduction of rotavirus vaccination in all Brazilian territories, thereby serving as a basis for monitoring changes in the genetic constitution of rotaviruses. The results also contribute to the understanding of the diversity and evolution of rotaviruses in a global context.

  12. High genotypic diversity of the reef-building coral Porites lobata (Scleractinia: Poritidae in Isla del Coco National Park, Costa Rica

    Directory of Open Access Journals (Sweden)

    Jennifer N. Boulay

    2012-11-01

    Full Text Available The isolated Isla del Coco experiences periodic, extreme disturbances which devastate coral reefs surrounding the island. Scleractinian corals build the physical structure of the reef therefore ecosystem recovery relies on coral species recovery. Coral recruits can be of sexual or asexual origin, and the relative success of the two recruit types influences the speed and spread of recovery processes. Here we focus on the massive coral, Porites lobata, because it is the main reef-builder around Isla del Coco to describe the relative contribution of asexual and sexual recruits to population maintenance. P. lobata samples were collected using a spatially explicit random sampling design in three plots at Isla del Coco: Punta Ulloa (n=17, Bahía Weston (n=20 and Punta María (n=20 and samples were genotyped with 11 microsatellite markers. Additional sampling was conducted at three “coastal” sites near the Costa Rican mainland (Isla del Caño Biological Reserve: Caño1 (n=8, Caño2 (n=10, Caño5 (n=11 to compare the contributions of asexual and sexual recruits at Isla del Coco sites to coastal sites. Isla del Coco sites were characterized by small colony size (>60% of colonies Los ambientes marinos del Parque Nacional Isla del Coco experimentan perturbaciones extremas periódicamente como por ejemplo El Niño-Oscilación del Sur (ENOS que han devastado las comunidades coralinas. La cobertura coralino se redujo drásticamente durante El Niño de 1982-83. Los corales escleractinios construyen la estructura física de los arrecifes así que la recuperación de estos ecosistemas depende de que los corales se recuperen. Los corales pueden reproducirse sexual y asexualmente, y el éxito relativo de cada forma de reproducción va a guiar el proceso de recuperación con implicaciones potenciales a la diversidad de las comunidades asociadas. En la mayoría de los arrecifes alrededor de la Isla del Coco, el coral masivo, Porites lobata, es la especie

  13. OGDD (Olive Genetic Diversity Database): a microsatellite markers' genotypes database of worldwide olive trees for cultivar identification and virgin olive oil traceability.

    Science.gov (United States)

    Ben Ayed, Rayda; Ben Hassen, Hanen; Ennouri, Karim; Ben Marzoug, Riadh; Rebai, Ahmed

    2016-01-01

    Olive (Olea europaea), whose importance is mainly due to nutritional and health features, is one of the most economically significant oil-producing trees in the Mediterranean region. Unfortunately, the increasing market demand towards virgin olive oil could often result in its adulteration with less expensive oils, which is a serious problem for the public and quality control evaluators of virgin olive oil. Therefore, to avoid frauds, olive cultivar identification and virgin olive oil authentication have become a major issue for the producers and consumers of quality control in the olive chain. Presently, genetic traceability using SSR is the cost effective and powerful marker technique that can be employed to resolve such problems. However, to identify an unknown monovarietal virgin olive oil cultivar, a reference system has become necessary. Thus, an Olive Genetic Diversity Database (OGDD) (http://www.bioinfo-cbs.org/ogdd/) is presented in this work. It is a genetic, morphologic and chemical database of worldwide olive tree and oil having a double function. In fact, besides being a reference system generated for the identification of unkown olive or virgin olive oil cultivars based on their microsatellite allele size(s), it provides users additional morphological and chemical information for each identified cultivar. Currently, OGDD is designed to enable users to easily retrieve and visualize biologically important information (SSR markers, and olive tree and oil characteristics of about 200 cultivars worldwide) using a set of efficient query interfaces and analysis tools. It can be accessed through a web service from any modern programming language using a simple hypertext transfer protocol call. The web site is implemented in java, JavaScript, PHP, HTML and Apache with all major browsers supported. Database URL: http://www.bioinfo-cbs.org/ogdd/. © The Author(s) 2016. Published by Oxford University Press.

  14. Genetic diversity of Echinococcus spp. in Russia.

    Science.gov (United States)

    Konyaev, Sergey V; Yanagida, Tetsuya; Nakao, Minoru; Ingovatova, Galina M; Shoykhet, Yakov N; Bondarev, Alexandr Y; Odnokurtsev, Valeriy A; Loskutova, Kyunnyay S; Lukmanova, Gulnur I; Dokuchaev, Nikolai E; Spiridonov, Sergey; Alshinecky, Mikhail V; Sivkova, Tatyana N; Andreyanov, Oleg N; Abramov, Sergey A; Krivopalov, Anton V; Karpenko, Sergey V; Lopatina, Natalia V; Dupal, Tamara A; Sako, Yasuhito; Ito, Akira

    2013-11-01

    In Russia, both alveolar and cystic echinococcoses are endemic. This study aimed to identify the aetiological agents of the diseases and to investigate the distribution of each Echinococcus species in Russia. A total of 75 Echinococcus specimens were collected from 14 host species from 2010 to 2012. Based on the mitochondrial DNA sequences, they were identified as Echinococcus granulosus sensu stricto (s.s.), E. canadensis and E. multilocularis. E. granulosus s.s. was confirmed in the European Russia and the Altai region. Three genotypes, G6, G8 and G10 of E. canadensis were detected in Yakutia. G6 was also found in the Altai region. Four genotypes of E. multilocularis were confirmed; the Asian genotype in the western Siberia and the European Russia, the Mongolian genotype in an island of Baikal Lake and the Altai Republic, the European genotype from a captive monkey in Moscow Zoo and the North American genotype in Yakutia. The present distributional record will become a basis of public health to control echinococcoses in Russia. The rich genetic diversity demonstrates the importance of Russia in investigating the evolutionary history of the genus Echinococcus.

  15. Selective inferior petrosal sinus sampling without venous outflow diversion in the detection of a pituitary adenoma in Cushing's syndrome

    International Nuclear Information System (INIS)

    Andereggen, Lukas; Schroth, Gerhard; Gralla, Jan; Ozdoba, Christoph; Seiler, Rolf; Mariani, Luigi; Beck, Juergen; Widmer, Hans-Rudolf; Andres, Robert H.; Christ, Emanuel

    2012-01-01

    Conventional MRI may still be an inaccurate method for the non-invasive detection of a microadenoma in adrenocorticotropin (ACTH)-dependent Cushing's syndrome (CS). Bilateral inferior petrosal sinus sampling (BIPSS) with ovine corticotropin-releasing hormone (oCRH) stimulation is an invasive, but accurate, intervention in the diagnostic armamentarium surrounding CS. Until now, there is a continuous controversial debate regarding lateralization data in detecting a microadenoma. Using BIPSS, we evaluated whether a highly selective placement of microcatheters without diversion of venous outflow might improve detection of pituitary microadenoma. We performed BIPSS in 23 patients that met clinical and biochemical criteria of CS and with equivocal MRI findings. For BIPSS, the femoral veins were catheterized bilaterally with a 6-F catheter and the inferior petrosal sinus bilaterally with a 2.7-F microcatheter. A third catheter was placed in the right femoral vein. Blood samples were collected from each catheter to determine ACTH blood concentration before and after oCRH stimulation. In 21 patients, a central-to-peripheral ACTH gradient was found and the affected side determined. In 18 of 20 patients where transsphenoidal partial hypophysectomy was performed based on BIPSS findings, microadenoma was histologically confirmed. BIPSS had a sensitivity of 94% and a specificity of 67% after oCRH stimulation in detecting a microadenoma. Correct localization of the adenoma was achieved in all Cushing's disease patients. BIPSS remains the gold standard in the detection of a microadenoma in CS. Our findings show that the selective placement of microcatheters without venous outflow diversion might further enhance better recognition to localize the pituitary tumor. (orig.)

  16. Detection of temporal behaviour patterns of free-ranging cattle by means of diversity spectra

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    de Miguel, J. M.

    1991-06-01

    Full Text Available The aim of this paper is to detect temporal patterns of cattle behaviour. The method, diversity spectra, provides, on the one hand, the number of parts into which a temporary transect should be divided in order to understand the maximum segregation of cattle activities and, on the other, the clarity with which each segregation is defined. In the case under study (a 'dehesa' pasture-land in central Spain the maximum segregation of fundamental activities in cattle behaviour is reached by considering the year as divided into two periods: spring-summer and autumn-winter. Cattle behaviour shows an annual "coarse grain" pattern, which is associated with management activities and with the meteorological seasonality of the Mediterranean climate. However, within each of the two annual periods, maximum segregation is reached considering separately the days of observation. This "fine grain" pattern indicates within each season, a certain capacity for response to a fluctuating environment and determines very different behaviour on close days. During autumn-winter period cattle show seasonal and daily activity segregations which are clearer than during spring-summer. In the former period, the lack of grass, more severe climatic conditions and management would seem to be determining factors of this temporal behaviour pattern.

    [es] El objetivo del trabajo es identificar patrones temporales de comportamiento del ganado. El procedimiento utilizado, espectros de diversidad, permite apreciar, por un lado, el número de partes en que debe dividirse un transecto temporal para detectar la máxima segregación de las actividades del ganado y, por otro, el grado de definición con que se manifiesta dicha segregación. En el caso estudiado (una dehesa del centro de España la máxima segregación de las actividades fundamentales de comportamiento del ganado se produce al considerar el año dividido en dos periodos: primavera-verano y otoño-invierno. El

  17. Heterogeneous recombination among Hepatitis B virus genotypes.

    Science.gov (United States)

    Castelhano, Nadine; Araujo, Natalia M; Arenas, Miguel

    2017-10-01

    The rapid evolution of Hepatitis B virus (HBV) through both evolutionary forces, mutation and recombination, allows this virus to generate a large variety of adapted variants at both intra and inter-host levels. It can, for instance, generate drug resistance or the diverse viral genotypes that currently exist in the HBV epidemics. Concerning the latter, it is known that recombination played a major role in the emergence and genetic diversification of novel genotypes. In this regard, the quantification of viral recombination in each genotype can provide relevant information to devise expectations about the evolutionary trends of the epidemic. Here we measured the amount of this evolutionary force by estimating global and local recombination rates in >4700 HBV complete genome sequences corresponding to nine (A to I) HBV genotypes. Counterintuitively, we found that genotype E presents extremely high levels of recombination, followed by genotypes B and C. On the other hand, genotype G presents the lowest level, where recombination is almost negligible. We discuss these findings in the light of known characteristics of these genotypes. Additionally, we present a phylogenetic network to depict the evolutionary history of the studied HBV genotypes. This network clearly classified all genotypes into specific groups and indicated that diverse pairs of genotypes are derived from a common ancestor (i.e., C-I, D-E and, F-H) although still the origin of this virus presented large uncertainty. Altogether we conclude that the amount of observed recombination is heterogeneous among HBV genotypes and that this heterogeneity can influence on the future expansion of the epidemic. Copyright © 2017 Elsevier B.V. All rights reserved.

  18. Decoding noises in HIV computational genotyping.

    Science.gov (United States)

    Jia, MingRui; Shaw, Timothy; Zhang, Xing; Liu, Dong; Shen, Ye; Ezeamama, Amara E; Yang, Chunfu; Zhang, Ming

    2017-11-01

    Lack of a consistent and reliable genotyping system can critically impede HIV genomic research on pathogenesis, fitness, virulence, drug resistance, and genomic-based healthcare and treatment. At present, mis-genotyping, i.e., background noises in molecular genotyping, and its impact on epidemic surveillance is unknown. For the first time, we present a comprehensive assessment of HIV genotyping quality. HIV sequence data were retrieved from worldwide published records, and subjected to a systematic genotyping assessment pipeline. Results showed that mis-genotyped cases occurred at 4.6% globally, with some regional and high-risk population heterogeneities. Results also revealed a consistent mis-genotyping pattern in gp120 in all studied populations except the group of men who have sex with men. Our study also suggests novel virus diversities in the mis-genotyped cases. Finally, this study reemphasizes the importance of implementing a standardized genotyping pipeline to avoid genotyping disparity and to advance our understanding of virus evolution in various epidemiological settings. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. Molecular detection and sequence characterization of diverse rhabdoviruses in bats, China.

    Science.gov (United States)

    Xu, Lin; Wu, Jianmin; Jiang, Tinglei; Qin, Shaomin; Xia, Lele; Li, Xingyu; He, Biao; Tu, Changchun

    2018-01-15

    The Rhabdoviridae is among the most diverse families of RNA viruses and currently classified into 18 genera with some rhabdoviruses lethal to humans and other animals. Herein, we describe genetic characterization of three novel rhabdoviruses from bats in China. Of these, two viruses (Jinghong bat virus and Benxi bat virus) found in Rhinolophus bats showed a phylogenetic relationship with vesiculoviruses, and sequence analyses indicate that they represent two new species within the genus Vesiculovirus. The remaining Yangjiang bat virus found in Hipposideros larvatus bats were only distantly related to currently known rhabdoviruses. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. Comparison of Modified Impedance Whole Blood Platelet Aggregation Method Detecting Platelet Function in ACS Patients with Different CYP2C19 Genotypes.

    Science.gov (United States)

    Cui, Chanjuan; Qiao, Rui; Zhang, Jie

    2016-01-01

    A reliable laboratory test to monitor onclopidogrel platelet reactivity (PR) is very necessary. In addition, genetic factors also play an important part in onclopidogrel PR. This study aimed to modify the original impedance whole blood platelet aggregation assay associated with the release assay to monitor onclopidogrel PR and assess their relationship with genotype. We adjusted the concentration of calcium in the in vitro reaction system of platelet aggregation to modify the original impedance whole blood platelet aggregation assay. Meanwhile, chronolume, which quantified the adenosine triphosphate (ATP) released from platelet dense granules, is added to this reaction system to reflect the platelet release function. In the modified assay, platelet magnified activation time (MAT) and the maximal platelet ATP release value (RV) were used to reflect platelet function parameters. In the original assay, the electrical resistance (omega) and RV were used to reflect platelet function parameters. Onclopidogrel PR was detected by the original impedance whole blood platelet aggregation assay, modified assay, and flow cytometric vasodilator stimulated phosphoprotein (VASP) assay in 168 patients with acute coronary syndromes (ACS). CYP2C19*2 and CYP2C19*3 polymorphisms were also detected in all of these patients. This modified method showed that when 12.5 microL CaCl2 (0.2 mmol/L) was added to the reaction system, MAT was appropriate (93 +/- 23 seconds). The CVs for the modified impedance assay and release assay were 9.31% and 6.13%, respectively. The mean VASP-PRI in the patient group treated with clopidogrel was significantly lower than that in the control group without antiplatelet therapy (54.88 +/- 16.81% vs. 79.86 +/- 10.24%, p 50% group were shorter than that in the PRI 50% group were higher than that in the PRI omega) and RV of the original method showed no differences between the two groups [0 (0-2) vs. 0 (0-1.25), 0.05 (0-0.25) vs. 0.08 (0-0.24); p > 0.05, p > 0

  1. Genetic diversity detection of the domestic horse (Equus caballus by genes associated with coat color

    Directory of Open Access Journals (Sweden)

    Luz Correa A

    2015-09-01

    Full Text Available Objective. To assess the population structure and genetic diversity in populations of domestic horse (Equus caballus in the municipality Cienaga de Oro-Córdoba (Colombia. Materials and methods. Random sampling were conducted between August and October 2013, in adult animals on farms seven districts, which was carried out phenotypic characterization of each animal, based on autosomal markers encoding morphological Extension (E , Agouti (A, Cream (C, White (W, Gray (G, Tobiano (TO, Overo (O and Roan (RN. Population genetic parameters: allele frequency, genetic diversity, gene flow, Hardy-Weinberg equilibrium and genetic distance were calculated through the program POPGENE 1.31; the genetic structure was assessed using the program FSTAT v. 2.9.3.2. Results. 341 individuals were analyzed in the seven populations studied, where the Extension gene Was the MOST faq frequently as the Overo and Tobiano genes showed the lowest values. Insignificant values of genetic variability and population recorded a global level, likewise, low genetic differentiation among populations, accompanied by a high gene flow was obtained; an excess of heterozygotes at population and global level was observed; to this is added the presence of Hardy-Weinberg equilibrium in all populations relative to the markers studied and low genetic distance values were reported. Conclusions. The populations are highly genetically related, a situation that may result from the existing geographical proximity between them, favoring genetic exchange and the establishment of a metapopulation.

  2. Widespread recombination, reassortment, and transmission of unbalanced compound viral genotypes in natural arenavirus infections.

    Directory of Open Access Journals (Sweden)

    Mark D Stenglein

    2015-05-01

    Full Text Available Arenaviruses are one of the largest families of human hemorrhagic fever viruses and are known to infect both mammals and snakes. Arenaviruses package a large (L and small (S genome segment in their virions. For segmented RNA viruses like these, novel genotypes can be generated through mutation, recombination, and reassortment. Although it is believed that an ancient recombination event led to the emergence of a new lineage of mammalian arenaviruses, neither recombination nor reassortment has been definitively documented in natural arenavirus infections. Here, we used metagenomic sequencing to survey the viral diversity present in captive arenavirus-infected snakes. From 48 infected animals, we determined the complete or near complete sequence of 210 genome segments that grouped into 23 L and 11 S genotypes. The majority of snakes were multiply infected, with up to 4 distinct S and 11 distinct L segment genotypes in individual animals. This S/L imbalance was typical: in all cases intrahost L segment genotypes outnumbered S genotypes, and a particular S segment genotype dominated in individual animals and at a population level. We corroborated sequencing results by qRT-PCR and virus isolation, and isolates replicated as ensembles in culture. Numerous instances of recombination and reassortment were detected, including recombinant segments with unusual organizations featuring 2 intergenic regions and superfluous content, which were capable of stable replication and transmission despite their atypical structures. Overall, this represents intrahost diversity of an extent and form that goes well beyond what has been observed for arenaviruses or for viruses in general. This diversity can be plausibly attributed to the captive intermingling of sub-clinically infected wild-caught snakes. Thus, beyond providing a unique opportunity to study arenavirus evolution and adaptation, these findings allow the investigation of unintended anthropogenic impacts on

  3. First detection of multiple knockdown resistance (kdr)-like mutations in voltage-gated sodium channel using three new genotyping methods in Anopheles sinensis from Guangxi Province, China.

    Science.gov (United States)

    Tan, Wei L; Li, Chun X; Wang, Zhong M; Liu, Mei D; Dong, Yan D; Feng, Xiang Y; Wu, Zhi M; Guo, Xiao X; Xing, Dan; Zhang, Ying M; Wang, Zhong C; Zhao, Tong Y

    2012-09-01

    To investigate knockdown resistance (kdr)-like mutations associated with pyrethroid resistance in Anopheles sinensis (Wiedemann, 1828), from Guangxi province, southwest China, a segment of a sodium channel gene was sequenced and genotyped using three new genotyping assays. Direct sequencing revealed the presence of TTG-to-TCG and TG-to-TTT mutations at allele position L1014, which led to L1014S and L1014F substitutions in a few individual and two novel substitutions of N1013S and L1014W in two DNA templates. A low frequency of the kdr allele mostly in the heterozygous state of L1014S and L1014F was observed in this mosquito population. In this study, the genotyping of An. sinensis using three polymerase chain reaction-based methods generated consistent results, which agreed with the results of DNA sequencing. In total, 52 mosquitoes were genotyped using a direct sequencing assay. The number of mosquitoes and their genotypes were as follows: L/L = 24, L/S = 19, L/F = 8, and F/W = 1. The allelic frequency of L1014, 1014S, and 1014F were 72, 18, and 9%, respectively.

  4. Genetic diversity of merozoite surface antigens in Babesia bovis detected from Sri Lankan cattle.

    Science.gov (United States)

    Sivakumar, Thillaiampalam; Okubo, Kazuhiro; Igarashi, Ikuo; de Silva, Weligodage Kumarawansa; Kothalawala, Hemal; Silva, Seekkuge Susil Priyantha; Vimalakumar, Singarayar Caniciyas; Meewewa, Asela Sanjeewa; Yokoyama, Naoaki

    2013-10-01

    Babesia bovis, the causative agent of severe bovine babesiosis, is endemic in Sri Lanka. The live attenuated vaccine (K-strain), which was introduced in the early 1990s, has been used to immunize cattle populations in endemic areas of the country. The present study was undertaken to determine the genetic diversity of merozoite surface antigens (MSAs) in B. bovis isolates from Sri Lankan cattle, and to compare the gene sequences obtained from such isolates against those of the K-strain. Forty-four bovine blood samples isolated from different geographical regions of Sri Lanka and judged to be B. bovis-positive by PCR screening were used to amplify MSAs (MSA-1, MSA-2c, MSA-2a1, MSA-2a2, and MSA-2b), AMA-1, and 12D3 genes from parasite DNA. Although the AMA-1 and 12D3 gene sequences were highly conserved among the Sri Lankan isolates, the MSA gene sequences from the same isolates were highly diverse. Sri Lankan MSA-1, MSA-2c, MSA-2a1, MSA-2a2, and MSA-2b sequences clustered within 5, 2, 4, 1, and 9 different clades in the gene phylograms, respectively, while the minimum similarity values among the deduced amino acid sequences of these genes were 36.8%, 68.7%, 80.3%, 100%, and 68.3%, respectively. In the phylograms, none of the Sri Lankan sequences fell within clades containing the respective K-strain sequences. Additionally, the similarity values for MSA-1 and MSA-2c were 40-61.8% and 90.9-93.2% between the Sri Lankan isolates and the K-strain, respectively, while the K-strain MSA-2a/b sequence shared 64.5-69.8%, 69.3%, and 70.5-80.3% similarities with the Sri Lankan MSA-2a1, MSA-2a2, and MSA-2b sequences, respectively. The present study has shown that genetic diversity among MSAs of Sri Lankan B. bovis isolates is very high, and that the sequences of field isolates diverged genetically from the K-strain. Copyright © 2013 Elsevier B.V. All rights reserved.

  5. Molecular detection and genetic diversity of Babesia gibsoni in dogs in Bangladesh.

    Science.gov (United States)

    Terao, Masashi; Akter, Shirin; Yasin, Md Golam; Nakao, Ryo; Kato, Hirotomo; Alam, Mohammad Zahangir; Katakura, Ken

    2015-04-01

    Babesia gibsoni is a tick-borne hemoprotozoan parasite of dogs that often causes fever and hemolytic illness. Detection of B. gibsoni has been predominantly reported in Asian countries, including Japan, Korea, Taiwan, Malaysia, Bangladesh and India. The present study shows the first molecular characterization of B. gibsoni detected from dogs in Bangladesh. Blood samples were collected on FTA® Elute cards from 50 stray dogs in Mymensingh District in Bangladesh. DNA eluted from the cards was subjected to nested PCR for the 18S rRNA gene of Babesia species. Approximately 800bp PCR products were detected in 15 of 50 dogs (30%). Based on restriction fragment length polymorphism (RFLP) and direct sequencing of the PCR products, all parasite isolates were identified as B. gibsoni. Furthermore, the BgTRAP (B. gibsoni thrombospondin-related adhesive protein) gene fragments were detected in 13 of 15 18S rRNA gene PCR positive blood samples. Phylogenetic analysis of the BgTRAP gene revealed that B. gibsoni parasites in Bangladesh formed a cluster, which was genetically different from other Asian B. gibsoni isolates. In addition, tandem repeat analysis of the BgTRAP gene clearly showed considerable genetic variation among Bangladeshi isolates. These results suggested that B. gibsoni parasites in a different genetic clade are endemic in dogs in Bangladesh. Further studies are required to elucidate the origin, distribution, vector and pathogenesis of B. gibsoni parasites circulating in dogs in Bangladesh. Copyright © 2015 Elsevier B.V. All rights reserved.

  6. Combined diversity and improved energy detection in cooperative spectrum sensing with faded reporting channels

    Directory of Open Access Journals (Sweden)

    Srinivas Nallagonda

    2016-04-01

    Full Text Available In this paper we evaluate the performance of cooperative spectrum sensing (CSS where each cognitive radio (CR employs an improved energy detector (IED with multiple antennas and uses selection combining (SC for detecting the primary user (PU in noisy and faded sensing (S channels. We derive an expression for the probability of false alarm and expressions for probability of missed detection in non-faded (AWGN and Rayleigh faded sensing environments in terms of cumulative distribution function (CDF. Each CR transmits its decision about PU via noisy and faded reporting (R channel to fusion center (FC. In this paper we assume that S-channels are noisy and Rayleigh faded while several cases of fading are considered for R-channels such as: (i Hoyt (or Nakagami-q, (ii Rayleigh, (iii Rician (or Nakagami-n, and (iv Weibull. A Binary Symmetric channel (BSC with a fixed error probability (r in the R-channel is also considered. The impact of fading in R-channel, S-channel and several network parameters such as IED parameter, normalized detection threshold, number of CRs, and number of antennas on missed detection and total error probability is assessed. The effects of Hoyt, Rician, and Weibull fading parameters on overall performance of IED-CSS are also highlighted.

  7. Distribution of Porphyromonas gingivalis fimA genotypes in primary endodontic infections.

    Science.gov (United States)

    Rôças, Isabela N; Siqueira, José F

    2010-03-01

    Long fimbriae (FimA) are important virulence factors of Porphyromonas gingivalis. Based on the diversity of the fimA gene, this species is classified into 6 genotypes. This study surveyed samples from primary endodontic infections for the presence of these P. gingivalis fimA variants. Genomic DNA isolated from samples taken from 25 root canals of teeth with chronic apical periodontitis and 25 aspirates from acute apical abscess was used as template in polymerase chain reaction (PCR) assays directed toward the detection of the different P. gingivalis fimA genotypes. Porphyromonas gingivalis was detected by a 16S rRNA gene-based PCR in 36% of the total number of cases sampled (44% of chronic apical periodontitis and 28% of abscess aspirates). In cases of chronic apical periodontitis, P. gingivalis variant type IV was the most prevalent (24%), followed by types I (20%), II (16%), and III (8%). In acute abscess samples, variant type II was the most prevalent (12%), followed by types III and IV (8% of each) and type I (4%). Combinations of up to 3 different genotypes were detected in a few cases. No single fimA genotype variant or combination thereof was significantly associated with symptoms. Overall, fimA types IV (16%), II (14%), and I (12%) were the most prevalent. Findings demonstrated that different P. gingivalis fimA genotypes can be present in primary endodontic infections. Copyright 2010 Mosby, Inc. All rights reserved.

  8. Genotyping of vacA alleles of Helicobacter pylori strains recovered ...

    African Journals Online (AJOL)

    commonly detected genotypes in the meat-based foods, viz, vegetable sandwich and ready to eat fish, were vacA ... Keywords: Helicobacter pylori, VacA genotypes, Genotyping, Food items ..... Microbiology and Quality Control, Islamic Azad.

  9. Diversity of virulence genes in Brucella melitensis and Brucella abortus detected from patients with rheumatoid arthritis.

    Science.gov (United States)

    Rahdar, Hossein Ali; Golmohammadi, Reza; Mirnejad, Reza; Ataee, Ramezan Ali; Alishiri, Gholam Hossein; Kazemian, Hossein

    2018-03-22

    The presence of Brucella melitensis and Brucella abortus genomes were investigated in the synovial fluid (SF) samples from 90 patients with rheumatoid arthritis (RA). DNA extraction and PCR assay were performed for simultaneous identification and discrimination of B. melitensis and B. abortus from the SF using three specific primers. After gel electrophoresis, the PCR products were confirmed by DNA sequencing. The cbg, omp31, manA, virB, and znuA virulence genes typing were performed by multiplex-PCR. Of the 90 samples, 14 were positive for B. melitensis (n = 9; 10%) and B. abortus (n = 5; 5.5%). The virulotyping of positive samples revealed the presence of all five virulence genes in B. melitensis. The virB, cbg, and om31 were detected in all five samples of B. abortus. In addition, zhuA and manA were detected in three (60%) and four (80%) samples, respectively, of the B. abortus-positive samples. Moreover, a total of 94.2% and 89.2% of the 14 positive samples were also found positive for manA and znuA, respectively. Our findings revealed that the Brucella spp. genomes can be detected in the SF of RA patients by the PCR-based method. We thus suggest that physicians should consider the Brucella spp. as indicators of potential RA for the timely diagnosis and treatment of RA. Copyright © 2018. Published by Elsevier Ltd.

  10. Molecular Diversity of Trypanosoma cruzi Detected in the Vector Triatoma protracta from California, USA.

    Directory of Open Access Journals (Sweden)

    Lisa A Shender

    2016-01-01

    Full Text Available Trypanosoma cruzi, causative agent of Chagas disease in humans and dogs, is a vector-borne zoonotic protozoan parasite that can cause fatal cardiac disease. While recognized as the most economically important parasitic infection in Latin America, the incidence of Chagas disease in the United States of America (US may be underreported and even increasing. The extensive genetic diversity of T. cruzi in Latin America is well-documented and likely influences disease progression, severity and treatment efficacy; however, little is known regarding T. cruzi strains endemic to the US. It is therefore important to expand our knowledge on US T. cruzi strains, to improve upon the recognition of and response to locally acquired infections.We conducted a study of T. cruzi molecular diversity in California, augmenting sparse genetic data from southern California and for the first time investigating genetic sequences from northern California. The vector Triatoma protracta was collected from southern (Escondido and Los Angeles and northern (Vallecito California regions. Samples were initially screened via sensitive nuclear repetitive DNA and kinetoplast minicircle DNA PCR assays, yielding an overall prevalence of approximately 28% and 55% for southern and northern California regions, respectively. Positive samples were further processed to identify discrete typing units (DTUs, revealing both TcI and TcIV lineages in southern California, but only TcI in northern California. Phylogenetic analyses (targeting COII-ND1, TR and RB19 genes were performed on a subset of positive samples to compare Californian T. cruzi samples to strains from other US regions and Latin America. Results indicated that within the TcI DTU, California sequences were similar to those from the southeastern US, as well as to several isolates from Latin America responsible for causing Chagas disease in humans.Triatoma protracta populations in California are frequently infected with T. cruzi

  11. Genetic relationship among Musa genotypes revealed by ...

    African Journals Online (AJOL)

    enoh

    2012-03-29

    Mar 29, 2012 ... A banana germplasm was established containing 44 Musa genotypes collected from various locations in Malaysia. To detect their genetic variation and to rule out duplicates among cultivar, microsatellite markers were used in their analysis. The microsatellite profiles of 44 Musa genotypes of various origins.

  12. Genome-wide detection of copy number variations among diverse horse breeds by array CGH.

    Directory of Open Access Journals (Sweden)

    Wei Wang

    Full Text Available Recent studies have found that copy number variations (CNVs are widespread in human and animal genomes. CNVs are a significant source of genetic variation, and have been shown to be associated with phenotypic diversity. However, the effect of CNVs on genetic variation in horses is not well understood. In the present study, CNVs in 6 different breeds of mare horses, Mongolia horse, Abaga horse, Hequ horse and Kazakh horse (all plateau breeds and Debao pony and Thoroughbred, were determined using aCGH. In total, seven hundred CNVs were identified ranging in size from 6.1 Kb to 0.57 Mb across all autosomes, with an average size of 43.08 Kb and a median size of 15.11 Kb. By merging overlapping CNVs, we found a total of three hundred and fifty-three CNV regions (CNVRs. The length of the CNVRs ranged from 6.1 Kb to 1.45 Mb with average and median sizes of 38.49 Kb and 13.1 Kb. Collectively, 13.59 Mb of copy number variation was identified among the horses investigated and accounted for approximately 0.61% of the horse genome sequence. Five hundred and eighteen annotated genes were affected by CNVs, which corresponded to about 2.26% of all horse genes. Through the gene ontology (GO, genetic pathway analysis and comparison of CNV genes among different breeds, we found evidence that CNVs involving 7 genes may be related to the adaptation to severe environment of these plateau horses. This study is the first report of copy number variations in Chinese horses, which indicates that CNVs are ubiquitous in the horse genome and influence many biological processes of the horse. These results will be helpful not only in mapping the horse whole-genome CNVs, but also to further research for the adaption to the high altitude severe environment for plateau horses.

  13. Phenotypic and genotypic detection of ESBL mediated cephalosporin resistance in Klebsiella pneumoniae: emergence of high resistance against cefepime, the fourth generation cephalosporin.

    Science.gov (United States)

    Grover, S S; Sharma, Meenakshi; Chattopadhya, D; Kapoor, Hema; Pasha, S T; Singh, Gajendra

    2006-10-01

    common resistance phenotypes probably belonged to a single strain as evident by MIC patterns, genotypic characterization and resistance profile to non-cephalosporin group of antimicrobials thereby pointing out the possibility of an outbreak. PCR may be regarded as a reliable method for detection of ESBL since in addition to the strains that could be identified as ESBL producers by DDST and E-test ESBL; PCR could demonstrate ESBL production among additional 32 strains (15 in phase I and 17 in phase II). Continued uses of cephalosporin group appear to be a potential risk factor for emergence of ESBL producing K. pneumoniae strains. In addition, as noted in the present study, the rise of resistance to cefepime that has been introduced recently in this country for therapeutic use could be of concern.

  14. Sequence diversity within the HA-1 gene as detected by melting temperature assay without oligonucleotide probes

    Directory of Open Access Journals (Sweden)

    Mattiuz Pier

    2005-10-01

    Full Text Available Abstract Background The minor histocompatibility antigens (mHags are self-peptides derived from common cellular proteins and presented by MHC class I and II molecules. Disparities in mHags are a potential risk for the development of graft-versus-host disease (GvHD in the recipients of bone marrow from HLA-identical donors. Two alleles have been identified in the mHag HA-1. The correlation between mismatches of the mHag HA-1 and GvHD has been suggested and methods to facilitate large-scale testing were afterwards developed. Methods We used sequence specific primer (SSP PCR and direct sequencing to detect HA-1 gene polymorphisms in a sample of 131 unrelated Italian subjects. We then set up a novel melting temperature (Tm assay that may help identification of HA-1 alleles without oligonucleotide probes. Results We report the frequencies of HA-1 alleles in the Italian population and the presence of an intronic 5 base-pair deletion associated with the immunogeneic allele HA-1H. We also detected novel variable sites with respect to the consensus sequence of HA-1 locus. Even though recombination/gene conversion events are documented, there is considerable linkage disequilibrium in the data. The gametic associations between HA-1R/H alleles and the intronic 5-bp ins/del polymorphism prompted us to try the Tm analysis with SYBR® Green I. We show that the addition of dimethylsulfoxide (DMSO during the assay yields distinct patterns when amplicons from HA-1H homozygotes, HA-1R homozygotes, and heterozygotes are analysed. Conclusion The possibility to use SYBR® Green I to detect Tm differences between allelic variants is attractive but requires great caution. We succeeded in allele discrimination of the HA-1 locus using a relatively short (101 bp amplicon, only in the presence of DMSO. We believe that, at least in certain assets, Tm assays may benefit by the addition of DMSO or other agents affecting DNA strand conformation and stability.

  15. Propionibacterium acnes: disease-causing agent or common contaminant? Detection in diverse patient samples by next generation sequencing

    DEFF Research Database (Denmark)

    Mollerup, Sarah; Friis-Nielsen, Jens; Vinner, Lasse

    2016-01-01

    Propionibacterium acnes is the most abundant bacterium on human skin, particularly in sebaceous areas. P. acnes is suggested to be an opportunistic pathogen involved in the development of diverse medical conditions, but is also a proven contaminant of human samples and surgical wounds. Its...... significance as a pathogen is consequently a matter of debate.In the present study we investigated the presence of P. acnes DNA in 250 next generation sequencing datasets generated from 180 samples of 20 different sample types, mostly of cancerous origin. The samples were either subjected to microbial...... enrichment, involving nuclease treatment to reduce the amount of host nucleic acids, or shotgun-sequenced.We detected high proportions of P. acnes in enriched samples, particularly skin derived and other tissue samples, with levels being higher in enriched compared to shotgun-sequenced samples. P. acnes...

  16. Detection of braking intention in diverse situations during simulated driving based on EEG feature combination.

    Science.gov (United States)

    Kim, Il-Hwa; Kim, Jeong-Woo; Haufe, Stefan; Lee, Seong-Whan

    2015-02-01

    We developed a simulated driving environment for studying neural correlates of emergency braking in diversified driving situations. We further investigated to what extent these neural correlates can be used to detect a participant's braking intention prior to the behavioral response. We measured electroencephalographic (EEG) and electromyographic signals during simulated driving. Fifteen participants drove a virtual vehicle and were exposed to several kinds of traffic situations in a simulator system, while EEG signals were measured. After that, we extracted characteristic features to categorize whether the driver intended to brake or not. Our system shows excellent detection performance in a broad range of possible emergency situations. In particular, we were able to distinguish three different kinds of emergency situations (sudden stop of a preceding vehicle, sudden cutting-in of a vehicle from the side and unexpected appearance of a pedestrian) from non-emergency (soft) braking situations, as well as from situations in which no braking was required, but the sensory stimulation was similar to stimulations inducing an emergency situation (e.g., the sudden stop of a vehicle on a neighboring lane). We proposed a novel feature combination comprising movement-related potentials such as the readiness potential, event-related desynchronization features besides the event-related potentials (ERP) features used in a previous study. The performance of predicting braking intention based on our proposed feature combination was superior compared to using only ERP features. Our study suggests that emergency situations are characterized by specific neural patterns of sensory perception and processing, as well as motor preparation and execution, which can be utilized by neurotechnology based braking assistance systems.

  17. Detection of braking intention in diverse situations during simulated driving based on EEG feature combination

    Science.gov (United States)

    Kim, Il-Hwa; Kim, Jeong-Woo; Haufe, Stefan; Lee, Seong-Whan

    2015-02-01

    Objective. We developed a simulated driving environment for studying neural correlates of emergency braking in diversified driving situations. We further investigated to what extent these neural correlates can be used to detect a participant's braking intention prior to the behavioral response. Approach. We measured electroencephalographic (EEG) and electromyographic signals during simulated driving. Fifteen participants drove a virtual vehicle and were exposed to several kinds of traffic situations in a simulator system, while EEG signals were measured. After that, we extracted characteristic features to categorize whether the driver intended to brake or not. Main results. Our system shows excellent detection performance in a broad range of possible emergency situations. In particular, we were able to distinguish three different kinds of emergency situations (sudden stop of a preceding vehicle, sudden cutting-in of a vehicle from the side and unexpected appearance of a pedestrian) from non-emergency (soft) braking situations, as well as from situations in which no braking was required, but the sensory stimulation was similar to stimulations inducing an emergency situation (e.g., the sudden stop of a vehicle on a neighboring lane). Significance. We proposed a novel feature combination comprising movement-related potentials such as the readiness potential, event-related desynchronization features besides the event-related potentials (ERP) features used in a previous study. The performance of predicting braking intention based on our proposed feature combination was superior compared to using only ERP features. Our study suggests that emergency situations are characterized by specific neural patterns of sensory perception and processing, as well as motor preparation and execution, which can be utilized by neurotechnology based braking assistance systems.

  18. Cytogenetics of Physalis peruviana L. and Physalis floridana Rydb. genotypes with differential response to Fusarium oxysporum

    Directory of Open Access Journals (Sweden)

    Sara A. Liberato G.

    2014-01-01

    Full Text Available Vascular wilt caused by the fungus Fusarium oxysporumis considered the main constraint of cape gooseberry, Physalis peruviana, production in Colombia. P. peruvianaand P. floridana genotypes with differential resistance responses against F. oxysporum have been identified previously. In the present study, the genotypes were evaluated in order to complement the knowledge of cytogenetics diversity in Physalisand to design hybridization strategies to support breeding of cape gooseberry crop. The chromosome number in mitotic dividing cells from root-tips of tissue culture plantlets was determined, from which the average mitotic hour was estimated at 12:00 hours for P. peruviana and 10:00 for P. floridana. Chromosomic complements of 2n = 4x = 48 and 2n = 2x = 24 were found for each one of the two species. Additionally, flow cytometry analyses detected variation within P. peruviana with a nuclear DNA content of 2.33 pg for the 2n = 24 genotype and variations ranged from 5.77 to 8.12 pg for 2n = 48 genotypes. In P. floridanaDNA content was 2.29 pg in the 2n = 24 genotype and 4.03 pg in the 2n = 48 genotype. There was a significant effect (α = 0.01 of the number of chromosomes on nuclear DNA content for the two species.

  19. Detecting exact breakpoints of deletions with diversity in hepatitis B viral genomic DNA from next-generation sequencing data.

    Science.gov (United States)

    Cheng, Ji-Hong; Liu, Wen-Chun; Chang, Ting-Tsung; Hsieh, Sun-Yuan; Tseng, Vincent S

    2017-10-01

    Many studies have suggested that deletions of Hepatitis B Viral (HBV) are associated with the development of progressive liver diseases, even ultimately resulting in hepatocellular carcinoma (HCC). Among the methods for detecting deletions from next-generation sequencing (NGS) data, few methods considered the characteristics of virus, such as high evolution rates and high divergence among the different HBV genomes. Sequencing high divergence HBV genome sequences using the NGS technology outputs millions of reads. Thus, detecting exact breakpoints of deletions from these big and complex data incurs very high computational cost. We proposed a novel analytical method named VirDelect (Virus Deletion Detect), which uses split read alignment base to detect exact breakpoint and diversity variable to consider high divergence in single-end reads data, such that the computational cost can be reduced without losing accuracy. We use four simulated reads datasets and two real pair-end reads datasets of HBV genome sequence to verify VirDelect accuracy by score functions. The experimental results show that VirDelect outperforms the state-of-the-art method Pindel in terms of accuracy score for all simulated datasets and VirDelect had only two base errors even in real datasets. VirDelect is also shown to deliver high accuracy in analyzing the single-end read data as well as pair-end data. VirDelect can serve as an effective and efficient bioinformatics tool for physiologists with high accuracy and efficient performance and applicable to further analysis with characteristics similar to HBV on genome length and high divergence. The software program of VirDelect can be downloaded at https://sourceforge.net/projects/virdelect/. Copyright © 2017. Published by Elsevier Inc.

  20. Exploiting orthologue diversity for systematic detection of gain-of-function phenotypes

    Directory of Open Access Journals (Sweden)

    Cantarella Daniela

    2008-05-01

    Full Text Available Abstract Background Systematic search for genes whose gain-of-function by exogenous expression confers an advantage in cell-based selective screenings is a powerful method for unbiased functional exploration of the genome, and has the potential to disclose new targets for cancer therapy. A major limit of this approach resides in the labor-intensive cloning of resistant cells, identification of the integrated genes and validation of their ability to confer a selective advantage. Moreover, the selection has to be drastic and genes conferring a limited advantage are typically missed. Results We developed a new functional screening strategy based on transduction of mammalian cells of a given species with an expression library from another species, followed by one-shot quantitative tracing with DNA microarrays of all library-derived transcripts before and after selection. In this way, exogenous transcripts enriched after selection, and therefore likely to confer resistance, are readily detected. We transduced a retroviral cDNA expression library from mouse testis into human and canine cells, and optimized the use of commercial murine gene expression arrays for species-specific detection of library-derived transcripts. We then conducted a functional screening by growing library-transduced canine MDCK cells in suspension, to enrich for cDNAs conferring anchorage independence. Notably, these cells show partial resistance to loss of anchorage, and the selection can be of limited stringency, compromising approaches based on clonal selection or anyway requiring high stringency. Microarray analysis revealed reproducible enrichment after three weeks of growth on polyhema for seven genes, among which the Hras proto-oncogene and Sox5. When individually transduced into MDCK cells, Sox5 specifically promoted anchorage-independent growth, thereby confirming the validity and specificity of the approach. Conclusion The procedure described here brings substantial

  1. Detection and characterization of diverse coccidian protozoa shed by California sea lions

    Science.gov (United States)

    Girard, Yvette A.; Johnson, Christine K.; Fritz, Heather M.; Shapiro, Karen; Packham, Andrea E.; Melli, Ann C.; Carlson-Bremer, Daphne; Gulland, Frances M.; Rejmanek, Daniel; Conrad, Patricia A.

    2015-01-01

    Tissue-cyst forming coccidia in the family Sarcocystidae are etiologic agents of protozoal encephalitis in marine mammals including the federally listed Southern sea otter (Enhydra lutris). California sea lions (Zalophus californianus), whose coastal habitat overlaps with sea otters, are definitive hosts for coccidian protozoa provisionally named Coccidia A, B and C. While Coccidia A and B have unknown clinical effects on aquatic wildlife hosts, Coccidia C is associated with severe protozoal disease in harbor seals (Phoca vitulina). In this study, we conducted surveillance for protozoal infection and fecal shedding in hospitalized and free-ranging California sea lions on the Pacific Coast and examined oocyst morphology and phenotypic characteristics of isolates via mouse bioassay and cell culture. Coccidia A and B were shed in similar frequency, particularly by yearlings. Oocysts shed by one free-ranging sea lion sampled at Año Nuevo State Park in California were previously unidentified in sea lions and were most similar to coccidia infecting Guadalupe fur seals (Arctocephalus townsendi) diagnosed with protozoal disease in Oregon (USA). Sporulated Coccidia A and B oocysts did not replicate in three strains of mice or in African green monkey kidney cells. However, cultivation experiments revealed that the inoculum of fecally-derived Coccidia A and B oocysts additionally contained organisms with genetic and antigenic similarity to Sarcocystis neurona; despite the absence of detectable free sporocysts in fecal samples by microscopic examination. In addition to the further characterization of Coccidia A and B in free-ranging and hospitalized sea lions, these results provide evidence of a new role for sea lions as putative mechanical vectors of S. neurona, or S. neurona-like species. Future work is needed to clarify the distribution, taxonomical status, and pathogenesis of these parasites in sea lions and other marine mammals that share their the near-shore marine

  2. Detection and characterization of diverse coccidian protozoa shed by California sea lions.

    Science.gov (United States)

    Girard, Yvette A; Johnson, Christine K; Fritz, Heather M; Shapiro, Karen; Packham, Andrea E; Melli, Ann C; Carlson-Bremer, Daphne; Gulland, Frances M; Rejmanek, Daniel; Conrad, Patricia A

    2016-04-01

    Tissue-cyst forming coccidia in the family Sarcocystidae are etiologic agents of protozoal encephalitis in marine mammals including the federally listed Southern sea otter (Enhydra lutris). California sea lions (Zalophus californianus), whose coastal habitat overlaps with sea otters, are definitive hosts for coccidian protozoa provisionally named Coccidia A, B and C. While Coccidia A and B have unknown clinical effects on aquatic wildlife hosts, Coccidia C is associated with severe protozoal disease in harbor seals (Phoca vitulina). In this study, we conducted surveillance for protozoal infection and fecal shedding in hospitalized and free-ranging California sea lions on the Pacific Coast and examined oocyst morphology and phenotypic characteristics of isolates via mouse bioassay and cell culture. Coccidia A and B were shed in similar frequency, particularly by yearlings. Oocysts shed by one free-ranging sea lion sampled at Año Nuevo State Park in California were previously unidentified in sea lions and were most similar to coccidia infecting Guadalupe fur seals (Arctocephalus townsendi) diagnosed with protozoal disease in Oregon (USA). Sporulated Coccidia A and B oocysts did not replicate in three strains of mice or in African green monkey kidney cells. However, cultivation experiments revealed that the inoculum of fecally-derived Coccidia A and B oocysts additionally contained organisms with genetic and antigenic similarity to Sarcocystis neurona; despite the absence of detectable free sporocysts in fecal samples by microscopic examination. In addition to the further characterization of Coccidia A and B in free-ranging and hospitalized sea lions, these results provide evidence of a new role for sea lions as putative mechanical vectors of S. neurona, or S. neurona-like species. Future work is needed to clarify the distribution, taxonomical status, and pathogenesis of these parasites in sea lions and other marine mammals that share their the near-shore marine

  3. Application of GC-MS for the detection of lipophilic compounds in diverse plant tissues

    Directory of Open Access Journals (Sweden)

    Hellmann Hanjo

    2009-04-01

    Full Text Available Abstract Background The concept of metabolite profiling has been around for decades and technical innovations are now enabling it to be carried out on a large scale with respect to the number of both metabolites measured and experiments carried out. However, studies are generally confined to polar compounds alone. Here we describe a simple method for lipophilic compounds analysis in various plant tissues. Results We choose the same preparative and instrumental platform for lipophilic profiling as that we routinely use for polar metabolites measurements. The method was validated in terms of linearity, carryover, reproducibility and recovery rates, as well as using various plant tissues. As a first case study we present metabolic profiling of Arabidopsis root and shoot tissue of wild type (C24 and mutant (rsr4-1 plants deficient on vitamin B6. We found significant alterations in lipid constituent contents, especially in the roots, which were characterised by dramatic increases in several fatty acids, thus providing further hint for the role of pyridoxine in oxidative stress and lipid peroxidation. The second example is the lipophilic profiling of red and green tomato fruit cuticles of wild type (Alisa Craig and the DFD (delayed fruit deterioration mutant, which we compared and contrasted with the more focused wax analysis of these plants reported before. Conclusion We can rapidly and reliably detect and quantify over 40 lipophilic metabolites including fatty acids, fatty alcohols, alkanes, sterols and tocopherols. The method presented here affords a simple and rapid, yet robust complement to previously validated methods of polar metabolite profiling by gas-chromatography mass-spectrometry.

  4. Detection and characterization of diverse coccidian protozoa shed by California sea lions

    Directory of Open Access Journals (Sweden)

    Yvette A. Girard

    2016-04-01

    Full Text Available Tissue-cyst forming coccidia in the family Sarcocystidae are etiologic agents of protozoal encephalitis in marine mammals including the federally listed Southern sea otter (Enhydra lutris. California sea lions (Zalophus californianus, whose coastal habitat overlaps with sea otters, are definitive hosts for coccidian protozoa provisionally named Coccidia A, B and C. While Coccidia A and B have unknown clinical effects on aquatic wildlife hosts, Coccidia C is associated with severe protozoal disease in harbor seals (Phoca vitulina. In this study, we conducted surveillance for protozoal infection and fecal shedding in hospitalized and free-ranging California sea lions on the Pacific Coast and examined oocyst morphology and phenotypic characteristics of isolates via mouse bioassay and cell culture. Coccidia A and B were shed in similar frequency, particularly by yearlings. Oocysts shed by one free-ranging sea lion sampled at Año Nuevo State Park in California were previously unidentified in sea lions and were most similar to coccidia infecting Guadalupe fur seals (Arctocephalus townsendi diagnosed with protozoal disease in Oregon (USA. Sporulated Coccidia A and B oocysts did not replicate in three strains of mice or in African green monkey kidney cells. However, cultivation experiments revealed that the inoculum of fecally-derived Coccidia A and B oocysts additionally contained organisms with genetic and antigenic similarity to Sarcocystis neurona; despite the absence of detectable free sporocysts in fecal samples by microscopic examination. In addition to the further characterization of Coccidia A and B in free-ranging and hospitalized sea lions, these results provide evidence of a new role for sea lions as putative mechanical vectors of S. neurona, or S. neurona-like species. Future work is needed to clarify the distribution, taxonomical status, and pathogenesis of these parasites in sea lions and other marine mammals that share their the near

  5. First detection and genotyping of Giardia intestinalis in stool samples collected from children in Ghazni Province, eastern Afghanistan and evaluation of the PCR assay in formalin-fixed specimens.

    Science.gov (United States)

    Lass, Anna; Karanis, Panagiotis; Korzeniewski, Krzysztof

    2017-08-01

    It is estimated that faecal-orally transmitted diseases are common in Afghanistan, as a consequence of poor hygienic standards of life and widespread contamination of water and food with both human and animal faeces. However, there is little information in the literature concerning infections caused by intestinal parasites in the Afghan population. In this study, we report the occurrence of Giardia intestinalis assemblages (A and B) in formalin-fixed stool samples collected from 245 Afghan schoolchildren living in Ghazni Province in eastern Afghanistan. Detection of the parasite's DNA and genotyping was performed using real-time PCR, specific to the β-giardin gene of G. intestinalis. Positive results were recorded in 52 (21.2%) samples. Genotyping was successful in 39 faecal samples and showed the predominance of assemblage B of G. intestinalis in this population (15 assemblage A and 24 assemblage B). Co-infection with both genotypes A and B was detected in four samples. Additionally, we evaluated the effect of 10% buffered formalin fixative on the detection of G. intestinalis DNA using real-time PCR and nested PCR characterised by different lengths of PCR products (74 and 479 bp, respectively). The human faeces containing the Giardia cysts were tested for 16 weeks. Amplification of G. intestinalis DNA with real-time PCR was possible up to 6 weeks of preservation of stool sample in formalin, compared to only 2 weeks with nested PCR. This suggests that real-time PCR is a more suitable tool in cases where stool samples have to be kept in formalin for longer periods of time.

  6. One-step multiplex real-time RT-PCR assay for detecting and genotyping wild-type group A rotavirus strains and vaccine strains (Rotarix® and RotaTeq®) in stool samples

    Science.gov (United States)

    Mijatovic-Rustempasic, Slavica; Esona, Mathew D.; Tam, Ka Ian; Quaye, Osbourne; Bowen, Michael D.

    2016-01-01

    Background. Group A rotavirus (RVA) infection is the major cause of acute gastroenteritis (AGE) in young children worldwide. Introduction of two live-attenuated rotavirus vaccines, RotaTeq® and Rotarix®, has dramatically reduced RVA associated AGE and mortality in developed as well as in many developing countries. High-throughput methods are needed to genotype rotavirus wild-type strains and to identify vaccine strains in stool samples. Quantitative RT-PCR assays (qRT-PCR) offer several advantages including increased sensitivity, higher throughput, and faster turnaround time. Methods. In this study, a one-step multiplex qRT-PCR assay was developed to detect and genotype wild-type strains and vaccine (Rotarix® and RotaTeq®) rotavirus strains along with an internal processing control (Xeno or MS2 RNA). Real-time RT-PCR assays were designed for VP7 (G1, G2, G3, G4, G9, G12) and VP4 (P[4], P[6] and P[8]) genotypes. The multiplex qRT-PCR assay also included previously published NSP3 qRT-PCR for rotavirus detection and Rotarix® NSP2 and RotaTeq® VP6 qRT-PCRs for detection of Rotarix® and RotaTeq® vaccine strains respectively. The multiplex qRT-PCR assay was validated using 853 sequence confirmed stool samples and 24 lab cultured strains of different rotavirus genotypes. By using thermostable rTth polymerase enzyme, dsRNA denaturation, reverse transcription (RT) and amplification (PCR) steps were performed in single tube by uninterrupted thermocycling profile to reduce chances of sample cross contamination and for rapid generation of results. For quantification, standard curves were generated using dsRNA transcripts derived from RVA gene segments. Results. The VP7 qRT-PCRs exhibited 98.8–100% sensitivity, 99.7–100% specificity, 85–95% efficiency and a limit of detection of 4–60 copies per singleplex reaction. The VP7 qRT-PCRs exhibited 81–92% efficiency and limit of detection of 150–600 copies in multiplex reactions. The VP4 qRT-PCRs exhibited 98.8

  7. Reflectance Spectra Diversity of Silica-Rich Materials: Sensitivity to Environment and Implications for Detections on Mars

    Science.gov (United States)

    Rice, M. S.; Cloutis, E. A.; Bell, J. F., III; Bish, D. L.; Horgan, B. H.; Mertzman, S. A.; Craig, M. A.; Renault, R. W.; Gautason, B.; Mountain, B.

    2013-01-01

    Hydrated silica-rich materials have recently been discovered on the surface of Mars by the Mars Exploration Rover (MER) Spirit, the Mars Reconnaissance Orbiter (MRO) Compact Reconnaissance Imaging Spectrometer for Mars (CRISM), and the Mars Express Observatoire pour la Mineralogie, l'Eau, les Glaces, et l'Activite'(OMEGA) in several locations. Having been interpreted as hydrothermal deposits and aqueous alteration products, these materials have important implications for the history of water on the martian surface. Spectral detections of these materials in visible to near infrared (Vis NIR) wavelengths have been based on a H2O absorption feature in the 934-1009 nm region seen with Spirit s Pancam instrument, and on SiOH absorption features in the 2.21-2.26 micron range seen with CRISM. Our work aims to determine how the spectral reflectance properties of silica-rich materials in Vis NIR wavelengths vary as a function of environmental conditions and formation. Here we present laboratory reflectance spectra of a diverse suite of silica-rich materials (chert, opal, quartz, natural sinters and synthetic silica) under a range of grain sizes and temperature, pressure, and humidity conditions. We find that the H2O content and form of H2O/OH present in silica-rich materials can have significant effects on their Vis NIR spectra. Our main findings are that the position of the approx.1.4 microns OH feature and the symmetry of the approx.1.9 microns feature can be used to discern between various forms of silica-rich materials, and that the ratio of the approx.2.2 microns (SiOH) and approx.1.9 microns (H2O) band depths can aid in distinguishing between silica phases (opal-A vs. opal-CT) and formation conditions (low vs. high temperature). In a case study of hydrated silica outcrops in Valles Marineris, we show that careful application of a modified version of these spectral parameters to orbital near-infrared spectra (e.g., from CRISM and OMEGA) can aid in characterizing the

  8. Multifunctional Pt(II) Reagents: Covalent Modifications of Pt Complexes Enable Diverse Structural Variation and In-Cell Detection.

    Science.gov (United States)

    White, Jonathan D; Haley, Michael M; DeRose, Victoria J

    2016-01-19

    To enhance the functionality of Pt-based reagents, several strategies have been developed that utilize Pt compounds modified with small, reactive handles. This Account encapsulates work done by us and other groups regarding the use of Pt(II) compounds with reactive handles for subsequent elaboration with fluorophores or other functional moieties. Described strategies include the incorporation of substituents for well-known condensation or nucleophilic displacement-type reactions and their use, for example, to tether spectroscopic handles to Pt reagents for in vivo investigation. Other chief uses of displacement-type reactions have included tethering various small molecules exhibiting pharmacological activity directly to Pt, thus adding synergistic effects. Click chemistry-based ligation techniques have also been applied, primarily with azide- and alkyne-appended Pt complexes. Orthogonally reactive click chemistry reactions have proven invaluable when more traditional nucleophilic displacement reactions induce side-reactivity with the Pt center or when systematic functionalization of a larger number of Pt complexes is desired. Additionally, a diverse assortment of Pt-fluorophore conjugates have been tethered via click chemistry conjugation. In addition to providing a convenient synthetic path for diversifying Pt compounds, the use of click-capable Pt complexes has proved a powerful strategy for postbinding covalent modification and detection with fluorescent probes. This strategy bypasses undesirable influences of the fluorophore camouflaged as reactivity due to Pt that may be present when detecting preattached Pt-fluorophore conjugates. Using postbinding strategies, Pt reagent distributions in HeLa and lung carcinoma (NCI-H460) cell cultures were observed with two different azide-modified Pt compounds, a monofunctional Pt(II)-acridine type and a difunctional Pt(II)-neutral complex. In addition, cellular distribution was observed with an alkyne-appended difunctional

  9. Detection of the Diversity of Cytoplasmic Male Sterility Sources in Broccoli (Brassica Oleracea var. Italica) Using Mitochondrial Markers.

    Science.gov (United States)

    Shu, Jinshuai; Liu, Yumei; Li, Zhansheng; Zhang, Lili; Fang, Zhiyuan; Yang, Limei; Zhuang, Mu; Zhang, Yangyong; Lv, Honghao

    2016-01-01

    Broccoli (Brassica oleracea var. italica) is an important commercial vegetable crop. As part of an efficient pollination system, cytoplasmic male sterility (CMS) has been widely used for broccoli hybrid production. Identifying the original sources of CMS in broccoli accessions has become an important part of broccoli breeding. In this study, the diversity of the CMS sources of 39 broccoli accessions, including 19 CMS lines and 20 hybrids, were analyzed using mitochondrial markers. All CMS accessions contained the ogu orf138-related DNA fragment and the key genes of nap CMS, pol CMS, and tour CMS were not detected. The 39 CMS accessions were divided into five groups using six orf138-related and two simple sequence repeat markers. We observed that ogu CMS R3 constituted 79.49% of the CMS sources. CMS6 and CMS26 were differentiated from the other accessions using a specific primer. CMS32 was distinguished from the other accessions based on a 78-nucleotide deletion at the same locus as the orf138-related sequence. When the coefficient was about 0.90, five CMS accessions (13CMS6, 13CMS23, 13CMS24, 13CMS37, and 13CMS39) exhibiting abnormal floral organs with poor seed setting were grouped together. The polymerase chain reaction amplification profiles for these five accessions differed from those of the other accessions. We identified eight useful molecular markers that can be used to detect CMS types during broccoli breeding. Our data also provide important information relevant to future studies on the possible origins and molecular mechanisms of CMS in broccoli.

  10. Next-Generation Sequencing Analysis of the Diversity of Human Noroviruses in Japanese Oysters.

    Science.gov (United States)

    Imamura, Saiki; Kanezashi, Hiromi; Goshima, Tomoko; Haruna, Mika; Okada, Tsukasa; Inagaki, Nobuya; Uema, Masashi; Noda, Mamoru; Akimoto, Keiko

    2017-08-01

    To obtain detailed information on the diversity of infectious norovirus in oysters (Crossostrea gigas), oysters obtained from fish producers at six different sites (sites A, B, C, D, E, and F) in Japan were analyzed once a month during the period spanning October 2015-February 2016. To avoid false-positive polymerase chain reaction (PCR) results derived from noninfectious virus particles, samples were pretreated with RNase before reverse transcription-PCR (RT-PCR). RT-PCR products were subjected to next-generation sequencing to identify norovirus genotypes in oysters. As a result, all GI genotypes were detected in the investigational period. The detection rate and proportion of norovirus GI genotypes differed depending on the sampling site and month. GII.3, GII.4, GII.13, GII.16, and GII.17 were detected in this study. Both the detection rate and proportion of norovirus GII genotypes differed depending on the sampling site and month. In total, the detection rate and proportion of GII.3 were highest from October to December among all detected genotypes. In January, the detection rates of GII.4 and GII.17 reached the same level as that of GII.3. The proportion of GII.17 was relatively lower from October to December, whereas it was the highest in January. To our knowledge, this is the first investigation on noroviruses in oysters in Japan, based on a method that can distinguish their infectivity.

  11. Food-Borne Outbreak Investigation and Molecular Typing: High Diversity of Staphylococcus aureus Strains and Importance of Toxin Detection

    Science.gov (United States)

    Denayer, Sarah; Nia, Yacine; Botteldoorn, Nadine

    2017-01-01

    Staphylococcus aureus is an important aetiological agent of food intoxications in the European Union as it can cause gastro-enteritis through the production of various staphylococcal enterotoxins (SEs) in foods. Reported enterotoxin dose levels causing food-borne illness are scarce and varying. Three food poisoning outbreaks due to enterotoxin-producing S. aureus strains which occurred in 2013 in Belgium are described. The outbreaks occurred in an elderly home, at a barbecue event and in a kindergarten and involved 28, 18, and six cases, respectively. Various food leftovers contained coagulase positive staphylococci (CPS). Low levels of staphylococcal enterotoxins ranging between 0.015 ng/g and 0.019 ng/g for enterotoxin A (SEA), and corresponding to 0.132 ng/g for SEC were quantified in the food leftovers for two of the reported outbreaks. Molecular typing of human and food isolates using pulsed-field gel electrophoresis (PFGE) and enterotoxin gene typing, confirmed the link between patients and the suspected foodstuffs. This also demonstrated the high diversity of CPS isolates both in the cases and in healthy persons carrying enterotoxin genes encoding emetic SEs for which no detection methods currently exist. For one outbreak, the investigation pointed out to the food handler who transmitted the outbreak strain to the food. Tools to improve staphylococcal food poisoning (SFP) investigations are presented. PMID:29261162

  12. Food-Borne Outbreak Investigation and Molecular Typing: High Diversity of Staphylococcus aureus Strains and Importance of Toxin Detection.

    Science.gov (United States)

    Denayer, Sarah; Delbrassinne, Laurence; Nia, Yacine; Botteldoorn, Nadine

    2017-12-20

    Staphylococcus aureus is an important aetiological agent of food intoxications in the European Union as it can cause gastro-enteritis through the production of various staphylococcal enterotoxins (SEs) in foods. Reported enterotoxin dose levels causing food-borne illness are scarce and varying. Three food poisoning outbreaks due to enterotoxin-producing S. aureus strains which occurred in 2013 in Belgium are described. The outbreaks occurred in an elderly home, at a barbecue event and in a kindergarten and involved 28, 18, and six cases, respectively. Various food leftovers contained coagulase positive staphylococci (CPS). Low levels of staphylococcal enterotoxins ranging between 0.015 ng/g and 0.019 ng/g for enterotoxin A (SEA), and corresponding to 0.132 ng/g for SEC were quantified in the food leftovers for two of the reported outbreaks. Molecular typing of human and food isolates using pulsed-field gel electrophoresis (PFGE) and enterotoxin gene typing, confirmed the link between patients and the suspected foodstuffs. This also demonstrated the high diversity of CPS isolates both in the cases and in healthy persons carrying enterotoxin genes encoding emetic SEs for which no detection methods currently exist. For one outbreak, the investigation pointed out to the food handler who transmitted the outbreak strain to the food. Tools to improve staphylococcal food poisoning (SFP) investigations are presented.

  13. Welcome to the neighbourhood: interspecific genotype by genotype interactions in Solidago influence above- and belowground biomass and associated communities.

    Science.gov (United States)

    Genung, Mark A; Bailey, Joseph K; Schweitzer, Jennifer A

    2012-01-01

    Intra- and interspecific plant-plant interactions are fundamental to patterns of community assembly and to the mixture effects observed in biodiversity studies. Although much research has been conducted at the species level, very little is understood about how genetic variation within and among interacting species may drive these processes. Using clones of both Solidago altissima and Solidago gigantea, we found that genotypic variation in a plant's neighbours affected both above- and belowground plant traits, and that genotype by genotype interactions between neighbouring plants impacted associated pollinator communities. The traits for which focal plant genotypic variation explained the most variation varied by plant species, whereas neighbour genotypic variation explained the most variation in coarse root biomass. Our results provide new insight into genotypic and species diversity effects in plant-neighbour interactions, the extended consequences of diversity effects, and the potential for evolution in response to competitive or to facilitative plant-neighbour interactions. © 2011 Blackwell Publishing Ltd/CNRS.

  14. Developmental plasticity: re-conceiving the genotype.

    Science.gov (United States)

    Sultan, Sonia E

    2017-10-06

    In recent decades, the phenotype of an organism (i.e. its traits and behaviour) has been studied as the outcome of a developmental 'programme' coded in its genotype. This deterministic view is implicit in the Modern Synthesis approach to adaptive evolution as a sorting process among genetic variants. Studies of developmental pathways have revealed that genotypes are in fact differently expressed depending on environmental conditions. Accordingly, the genotype can be understood as a repertoire of potential developmental outcomes or norm of reaction. Reconceiving the genotype as an environmental response repertoire rather than a fixed developmental programme leads to three critical evolutionary insights. First, plastic responses to specific conditions often comprise functionally appropriate trait adjustments, resulting in an individual-level, developmental mode of adaptive variation. Second, because genotypes are differently expressed depending on the environment, the genetic diversity available to natural selection is itself environmentally contingent. Finally, environmental influences on development can extend across multiple generations via cytoplasmic and epigenetic factors transmitted to progeny individuals, altering their responses to their own, immediate environmental conditions and, in some cases, leading to inherited but non-genetic adaptations. Together, these insights suggest a more nuanced understanding of the genotype and its evolutionary role, as well as a shift in research focus to investigating the complex developmental interactions among genotypes, environments and previous environments.

  15. Performance of chickpea genotypes under Swat valley conditions

    International Nuclear Information System (INIS)

    Khan, A.; Rahim, M.; Ahmad, F.; Ali, A.

    2004-01-01

    Twenty-two genetically diverse chickpeas genotypes were studied for their physiological efficiency to select the most desirable genotype/genotypes for breeding program on chickpea. Genotype 'CM7-1' was found physiologically efficient stain with maximum harvest index (37.33%) followed by genotype 'CM1571-1-A' with harvest index of 35.73%. Genotype '90206' produced maximum biological yield (7463 kg ha/sup -1/) followed by genotypes 'CM31-1' and 'E-2034' with biological yield of 7352 and 7167 kg ha/sup -1/, respectively. Harvest index and economic yield showed significant positive correlation value of (r=+0.595), while negative correlation value of (r = -0.435) was observed between harvest index and biological yield. (author)

  16. Characteristics of Streptococcus mutans genotypes and dental caries in children

    Science.gov (United States)

    Cheon, Kyounga; Moser, Stephen A.; Wiener, Howard W.; Whiddon, Jennifer; Momeni, Stephanie S.; Ruby, John D.; Cutter, Gary R.; Childers, Noel K.

    2013-01-01

    This longitudinal cohort study evaluated the diversity, commonality, and stability of Streptococcus mutans genotypes associated with dental caries history. Sixty-seven 5 and 6 yr-old children, considered being at high caries risk, had plaque collected from baseline through 36 months for S. mutans isolation and genotyping with repetitive extragenic palindromic-PCR (4,392 total isolates). Decayed, missing, filled surfaces (dmfs/DMFS) for each child were recorded at baseline. At baseline, 18 distinct genotypes were found among 911 S. mutans isolates from 67 children (diversity) and 13 genotypes were shared by at least 2 children (commonality). The number of genotypes per individual was positively associated with the proportion of decayed surfaces (p-ds) at baseline. Twenty-four of the 39 children who were available at follow-up visits maintained a predominant genotype for the follow-up periods (stability) and was negatively associated with p-ds. The observed diversity, commonality, and stability of S. mutans genotypes represent a pattern of dental caries epidemiology in this high caries risk community, which suggest fewer decayed surfaces are significantly associated with lower diversity and stability of S. mutans genotypes. PMID:23659236

  17. Molecular detection and genetic diversity of bovine Babesia spp., Theileria orientalis, and Anaplasma marginale in beef cattle in Thailand.

    Science.gov (United States)

    Jirapattharasate, Charoonluk; Adjou Moumouni, Paul Franck; Cao, Shinuo; Iguchi, Aiko; Liu, Mingming; Wang, Guanbo; Zhou, Mo; Vudriko, Patrick; Efstratiou, Artemis; Changbunjong, Tanasak; Sungpradit, Sivapong; Ratanakorn, Parntep; Moonarmart, Walasinee; Sedwisai, Poonyapat; Weluwanarak, Thekhawet; Wongsawang, Witsanu; Suzuki, Hiroshi; Xuan, Xuenan

    2017-02-01

    Babesia spp., Theileria orientalis, and Anaplasma marginale are significant tick-borne pathogens that affect the health and productivity of cattle in tropical and subtropical areas. In this study, we used PCR to detect the presence of Babesia bovis, Babesia bigemina, and T. orientalis in 279 beef cattle from Western Thailand and A. marginale in 608 beef cattle from the north, northeastern, and western regions. The PCRs were performed using species-specific primers based on the B. bovis spherical body protein 2 (BboSBP2), B. bigemina rhoptry-associated protein 1a (BbiRAP-1a), T. orientalis major piroplasm surface protein (ToMPSP), and A. marginale major surface protein 4 (AmMSP4) genes. To determine the genetic diversity of the above parasites, amplicons of B. bovis and B. bigemina ITS1-5.8s rRNA gene-ITS2 regions (B. bovis ITS, B. bigemina ITS), ToMPSP, and AmMSP4 genes were sequenced for phylogenetic analysis. PCR results revealed that the prevalence of B. bovis, B. bigemina, T. orientalis, and A. marginale in the Western region was 11.1, 12.5, 7.8, and 39.1 %, respectively. Coinfections of two or three parasites were observed in 17.9 % of the animals sampled. The study revealed that the prevalence of A. marginale in the western region was higher than in the north and northeastern regions (7 %). Sequence analysis showed the BboSBP2 gene to be more conserved than B. bovis ITS in the different isolates and, similarly, the BbiRAP-1a was more conserved than B. bigemina ITS. In the phylogenetic analysis, T. orientalis MPSP sequences were classified into types 3, 5, and 7 as previously reported. A. marginale MSP4 gene sequences shared high identity and similarity with each other and clustered with isolates from other countries. This study provides information on the prevalence and genetic diversity of tick-borne pathogens in beef cattle and highlights the need for effective strategies to control these pathogens in Thailand.

  18. Genotype 3 is the predominant hepatitis C genotype in a multi-ethnic Asian population in Malaysia.

    Science.gov (United States)

    Ho, Shiaw-Hooi; Ng, Kee-Peng; Kaur, Harvinder; Goh, Khean-Lee

    2015-06-01

    Genotypes of hepatitis C virus (HCV) are distributed differently across the world. There is a paucity of such data in a multi-ethnic Asian population like Malaysia. The objectives of this study were to determine the distribution of HCV genotypes between major ethnic groups and to ascertain their association with basic demographic variables like age and gender. This was a cross-sectional prospective study conducted from September 2007 to September 2013. Consecutive patients who were detected to have anti-HCV antibodies in the University of Malaya Medical Centre were included and tested for the presence of HCV RNA using Roche Cobas Amplicor Analyzer and HCV genotype using Roche single Linear Array HCV Genotyping strip. Five hundred and ninety-six subjects were found to have positive anti-HCV antibodies during this period of time. However, only 396 (66.4%) were HCV RNA positive and included in the final analysis. Our results showed that HCV genotype 3 was the predominant genotype with overall frequency of 61.9% followed by genotypes 1 (35.9%), 2 (1.8%) and 6 (0.5%). There was a slightly higher prevalence of HCV genotype 3 among the Malays when compared to the Chinese (P=0.043). No other statistical significant differences were observed in the distribution of HCV genotypes among the major ethnic groups. There was also no association between the predominant genotypes and basic demographic variables. In a multi-ethnic Asian society in Malaysia, genotype 3 is the predominant genotype among all the major ethnic groups with genotype 1 as the second commonest genotype. Both genotypes 2 and 6 are uncommon. Neither genotype 4 nor 5 was detected. There is no identification of HCV genotype according to ethnic origin, age and gender.

  19. Prevalence of hepatitis viruses in patients with acute hepatitis and characterization of the detected genotype 4 hepatitis E virus sequences in Mongolia.

    Science.gov (United States)

    Tsatsralt-Od, Bira; Baasanjav, Nachin; Nyamkhuu, Dulmaa; Ohnishi, Hiroshi; Takahashi, Masaharu; Okamoto, Hiroaki

    2016-02-01

    Hepatitis E is considered to be a worldwide public health problem. Although the prevalence of hepatitis E virus (HEV) antibodies in healthy individuals is noted to be 11%, no patients with acute hepatitis E have previously been identified in Mongolia. Three hundred two consecutive patients (183 males and 119 females; median age of 22.0 [Interquartile range: 18.3-25.0] years) who were clinically diagnosed with sporadic acute hepatitis during 2012-2013 in Ulaanbaatar, Mongolia, were studied. By serological and/or molecular approaches, 77 (25.5%), 93 (30.8%), 19 (6.3%), 48 (15.9%), and 12 (4.0%) of the patients were diagnosed with acute hepatitis of types A, B, C, D (superinfection of hepatitis delta virus on a background of chronic hepatitis B virus infection) and E, respectively, while the cause of hepatitis was unknown in the remaining 53 patients (17.5%). The 12 hepatitis E patients had no history of travel abroad in the 3 months before the onset of disease, and lived separately in fixed or movable houses with water supplied via pipe, tank or well, denying transmission from a common water supply. The 12 HEV isolates obtained from the patients showed high nucleotide identities of 99.7-100%, and a representative HEV isolate, MNE13-227, was closest to the Chinese isolates of genotype 4, with the highest identity of 97.3% in the 304-nt ORF2 sequence and 92.1% over the entire genome. The present study revealed the occurrence of autochthonous acute hepatitis E in Mongolia, caused by a monophyletic genotype 4 HEV strain. © 2015 Wiley Periodicals, Inc.

  20. Whole genome sequencing of genotype VI Newcastle disease viruses from formalin-fixed paraffinembedded tissues from wild pigeons reveals continuous evolution and previously unrecognized genetic diversity in the U.S.

    Science.gov (United States)

    Background: Newcastle disease viruses (NDV) are highly contagious and can cause disease in both wild birds and poultry. A pigeon-adapted variant of genotype VI NDV, termed pigeon paramyxovirus 1, is commonly isolated from Columbiform birds in the United States. Complete genomic characterization of t...

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

    Science.gov (United States)

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

    2018-01-01

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

  2. Genetic diversity in some Turkish pepper (Capsicum annuum L ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-09-15

    Sep 15, 2009 ... RFLP, RAPD, AFLP, and micro-satellites have been beneficial by being ... List of pepper genotypes used in the experiment. Genotype. Type. Source. Classification of ..... Artificial selection might decrease the genetic diversity ...

  3. Multilocus genotyping of Giardia duodenalis in captive non-human primates in Sichuan and Guizhou provinces, Southwestern China.

    Directory of Open Access Journals (Sweden)

    Zhijun Zhong

    Full Text Available Giardia duodenalis is a common human and animal pathogen. It has been increasingly reported in wild and captive non-human primates (NHPs in recent years. However, multilocus genotyping information for G. duodenalis infecting NHPs in southwestern China is limited. In the present study, the prevalence and multilocus genotypes (MLGs of G. duodenalis in captive NHPs in southwestern China were determined. We examined 207 fecal samples from NHPs in Sichuan and Guizhou provinces, and 16 specimens were positive for G. duodenalis. The overall infection rate was 7.7%, and only assemblage B was identified. G. duodenalis was detect positive in northern white-cheeked gibbon (14/36, 38.9%, crab-eating macaque (1/60, 1.7% and rhesus macaques (1/101, 0.9%. Multilocus sequence typing based on beta-giardin (bg, triose phosphate isomerase (tpi and glutamate dehydrogenase (gdh revealed nine different assemblage B MLGs (five known genotypes and four novel genotypes. Based on a phylogenetic analysis, one potentially zoonotic genotype of MLG SW7 was identified in a northern white-cheeked gibbon. A high degree of genetic diversity within assemblage B was observed in captive northern white-cheeked gibbons in Southwestern China, including a potentially zoonotic genotype, MLG SW7. To the best of our knowledge, this is the first report using a MLGs approach to identify G. duodenalis in captive NHPs in Southwestern China.

  4. An Evaluation of Quantitative PCR Assays (TaqMan® and SYBR Green for the Detection of Babesia bigemina and Babesia bovis, and a Novel Fluorescent-ITS1-PCR Capillary Electrophoresis Method for Genotyping B. bovis Isolates

    Directory of Open Access Journals (Sweden)

    Bing Zhang

    2016-09-01

    Full Text Available Babesia spp. are tick-transmitted haemoparasites causing tick fever in cattle. In Australia, economic losses to the cattle industry from tick fever are estimated at AUD$26 Million per annum. If animals recover from these infections, they become immune carriers. Here we describe a novel multiplex TaqMan qPCR targeting cytochrome b genes for the identification of Babesia spp. The assay shows high sensitivity, specificity and reproducibility, and allows quantification of parasite DNA from Babesia bovis and B. bigemina compared to standard PCR assays. A previously published cytochrome b SYBR Green qPCR was also tested in this study, showing slightly higher sensitivity than the Taqman qPCRs but requires melting curve analysis post-PCR to confirm specificity. The SYBR Green assays were further evaluated using both diagnostic submissions and vaccinated cattle (at 7, 9, 11 and 14 days post-inoculation showed that B. bigemina can be detected more frequently than B. bovis. Due to fewer circulating parasites, B. bovis detection in carrier animals requires higher DNA input. Preliminary data for a novel fluorescent PCR genotyping based on the Internal Transcribed Spacer 1 region to detect vaccine and field alleles of B. bovis are described. This assay is capable of detecting vaccine and novel field isolate alleles in a single sample.

  5. Construction of High Density Sweet Cherry (Prunus avium L. Linkage Maps Using Microsatellite Markers and SNPs Detected by Genotyping-by-Sequencing (GBS.

    Directory of Open Access Journals (Sweden)

    Verónica Guajardo

    Full Text Available Linkage maps are valuable tools in genetic and genomic studies. For sweet cherry, linkage maps have been constructed using mainly microsatellite markers (SSRs and, recently, using single nucleotide polymorphism markers (SNPs from a cherry 6K SNP array. Genotyping-by-sequencing (GBS, a new methodology based on high-throughput sequencing, holds great promise for identification of high number of SNPs and construction of high density linkage maps. In this study, GBS was used to identify SNPs from an intra-specific sweet cherry cross. A total of 8,476 high quality SNPs were selected for mapping. The physical position for each SNP was determined using the peach genome, Peach v1.0, as reference, and a homogeneous distribution of markers along the eight peach scaffolds was obtained. On average, 65.6% of the SNPs were present in genic regions and 49.8% were located in exonic regions. In addition to the SNPs, a group of SSRs was also used for construction of linkage maps. Parental and consensus high density maps were constructed by genotyping 166 siblings from a 'Rainier' x 'Rivedel' (Ra x Ri cross. Using Ra x Ri population, 462, 489 and 985 markers were mapped into eight linkage groups in 'Rainier', 'Rivedel' and the Ra x Ri map, respectively, with 80% of mapped SNPs located in genic regions. Obtained maps spanned 549.5, 582.6 and 731.3 cM for 'Rainier', 'Rivedel' and consensus maps, respectively, with an average distance of 1.2 cM between adjacent markers for both 'Rainier' and 'Rivedel' maps and of 0.7 cM for Ra x Ri map. High synteny and co-linearity was observed between obtained maps and with Peach v1.0. These new high density linkage maps provide valuable information on the sweet cherry genome, and serve as the basis for identification of QTLs and genes relevant for the breeding of the species.

  6. Coronavirus Infection and Diversity in Bats in the Australasian Region.

    Science.gov (United States)

    Smith, C S; de Jong, C E; Meers, J; Henning, J; Wang, L- F; Field, H E

    2016-03-01

    Following the SARS outbreak, extensive surveillance was undertaken globally to detect and identify coronavirus diversity in bats. This study sought to identify the diversity and prevalence of coronaviruses in bats in the Australasian region. We identified four different genotypes of coronavirus, three of which (an alphacoronavirus and two betacoronaviruses) are potentially new species, having less than 90% nucleotide sequence identity with the most closely related described viruses. We did not detect any SARS-like betacoronaviruses, despite targeting rhinolophid bats, the putative natural host taxa. Our fin