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  1. Environmental mycobacteria – Future directions

    Directory of Open Access Journals (Sweden)

    D E Schraufnagel

    2015-01-01

    An interferon gamma release assay for infection with nontuberculous mycobacteria is desperately needed. Success with tuberculosis was attained by finding bacterial antigens associated with virulence and invasive disease. This may be difficult because the damage caused by environmental mycobacteria tends to be epithelial and subepithelial. The cavities may be bronchiectatic in origin and deeper invasion may result from host reactions. Serologic testing has not yielded important clinical help for diseases caused by either M. tuberculosis or the nontuberculous mycobacteria, possibly because good bacterial or host response proteins have not been identified. This could change with a metabolomics approach. Genomic studies that give incremental gains may set the stage for major breakthroughs, but require coordination with good phenotyping. Tools are in place to open an exciting new era for understanding and control of mycobacteria.

  2. Role of genotype® mycobacterium common mycobacteria/additional species assay for rapid differentiation between Mycobacterium tuberculosis complex and different species of non-tuberculous mycobacteria

    Directory of Open Access Journals (Sweden)

    Amresh Kumar Singh

    2013-01-01

    Full Text Available Background: Mycobacterium tuberculosis complex (MTBC and non-tuberculous mycobacteria (NTM may or may not have same clinical presentations, but the treatment regimens are always different. Laboratory differentiation between MTBC and NTM by routine methods are time consuming and cumbersome to perform. We have evaluated the role of GenoType® Mycobacterium common mycobacteria/additional species (CM/AS assay for differentiation between MTBC and different species of NTM in clinical isolates from tuberculosis (TB cases. Materials and Methods: A total of 1080 clinical specimens were collected from January 2010 to June 2012. Diagnosis was performed by Ziehl-Neelsen staining followed by culture in BacT/ALERT 3D system (bioMerieux, France. A total of 219 culture positive clinical isolates (BacT/ALERT® MP cultures were selected for differentiation by p-nitrobenzoic acid (PNB sensitivity test as and BIO-LINE SD Ag MPT64 TB test considering as the gold standard test. Final identification and differentiation between MTBC and different species of NTM were further confirmed by GenoType® Mycobacterium CM/AS assay (Hain Lifescience, Nehren, Germany. Results: Out of 219 BacT/ALERT® MP culture positive isolates tested by PNB as 153 MTBC (69.9% and by GenoType® Mycobacterium CM/AS assay as 159 (72.6% MTBC and remaining 60 (27.4% were considered as NTM species. The GenoType® Mycobacterium CM/AS assay was proved 99.3% sensitive and 98.3% specific for rapid differentiation of MTBC and NTM. The most common NTM species were; Mycobacterium fortuitum 20 (33.3% among rapid growing mycobacteria and Mycobacterium intracellulare 11 (18.3% among slow growing mycobacteria. Conclusion: The GenoType® Mycobacterium assay makes rapid and accurate identification of NTM species as compared with different phenotypic and molecular diagnostic tool and helps in management of infections caused by different mycobacteria.

  3. Host-directed strategies using lipid nanoparticles to reduce mycobacteria survival

    Energy Technology Data Exchange (ETDEWEB)

    Pereira, L. [Faculty of Sciences and Technology of the New University of Lisbon (Portugal); Diogo, J.; Mateus, R.; Pimentel, M.; Videira, M., E-mail: mvideira@ff.ul.pt [iMed.UL - Research Institute for Medicines and Pharmaceutical Sciences, Faculty of Pharmacy of the University of Lisbon, Intracellular Trafficking Modulation for Advanced Drug Delivery InTraCell-ADD Research Group (Portugal)

    2015-02-15

    Antibiotic-resistant infections and the stagnations in the development of new drugs have increased the demand for new therapeutic approaches against Mycobacterium tuberculosis. Innovative systems that are able to target and eradicate the bacteria in the infected host cells may represent a therapeutic breakthrough while avoiding latency. The development of nanosystems aiming a controlled and targeted intracellular drug release, have proved to increase cytosolic therapeutic concentration while reducing undesired side effects. This work’s main goal was to develop a host-directed strategy against mycobacterial infection through the design of a biocompatible nanocarrier for phage-derived protein delivery, using M. smegmatis as model. Since mycobacterial pathogenicity is strongly supported by the presence of lipids in the cell wall, their degradation induces bacterial destruction through cell wall hydrolysis. Phage-based lipolytic enzymes such as, LysB a mycolylarabinogalactan esterase, represent an appealing therapeutic approach. The herein proposed Ms6 LysB-containing lipid nanocarrier (SLN-LysB) explores the known advantages of nanomedicine-based systems for phagocytic cells selectively targeting thus allowing LysB intracellular accumulation and a more pronounced mycobacterial infection eradication. Adsorption efficiency value indicates the potential of this system as a protein nanocarrier. Moreover, promising outcomes were obtained in host-infected macrophages treated with SLN-LysB. The results show that the herein proposed strategy was more effective in inhibiting the growth of M. smegmatis than free LysB, which might be related to the nanocarrier internalization. Acting as effective protein nanocarriers, the protein-guided delivery in the infected phagocytic cells allows it to exert its hydrolytic action on the lipid layer of the Mycobacterium.

  4. Host-directed strategies using lipid nanoparticles to reduce mycobacteria survival

    Science.gov (United States)

    Pereira, L.; Diogo, J.; Mateus, R.; Pimentel, M.; Videira, M.

    2015-02-01

    Antibiotic-resistant infections and the stagnations in the development of new drugs have increased the demand for new therapeutic approaches against Mycobacterium tuberculosis. Innovative systems that are able to target and eradicate the bacteria in the infected host cells may represent a therapeutic breakthrough while avoiding latency. The development of nanosystems aiming a controlled and targeted intracellular drug release, have proved to increase cytosolic therapeutic concentration while reducing undesired side effects. This work's main goal was to develop a host-directed strategy against mycobacterial infection through the design of a biocompatible nanocarrier for phage-derived protein delivery, using M. smegmatis as model. Since mycobacterial pathogenicity is strongly supported by the presence of lipids in the cell wall, their degradation induces bacterial destruction through cell wall hydrolysis. Phage-based lipolytic enzymes such as, LysB a mycolylarabinogalactan esterase, represent an appealing therapeutic approach. The herein proposed Ms6 LysB-containing lipid nanocarrier (SLN_LysB) explores the known advantages of nanomedicine-based systems for phagocytic cells selectively targeting thus allowing LysB intracellular accumulation and a more pronounced mycobacterial infection eradication. Adsorption efficiency value indicates the potential of this system as a protein nanocarrier. Moreover, promising outcomes were obtained in host-infected macrophages treated with SLN_LysB. The results show that the herein proposed strategy was more effective in inhibiting the growth of M. smegmatis than free LysB, which might be related to the nanocarrier internalization. Acting as effective protein nanocarriers, the protein-guided delivery in the infected phagocytic cells allows it to exert its hydrolytic action on the lipid layer of the Mycobacterium.

  5. Host-directed strategies using lipid nanoparticles to reduce mycobacteria survival

    International Nuclear Information System (INIS)

    Pereira, L.; Diogo, J.; Mateus, R.; Pimentel, M.; Videira, M.

    2015-01-01

    Antibiotic-resistant infections and the stagnations in the development of new drugs have increased the demand for new therapeutic approaches against Mycobacterium tuberculosis. Innovative systems that are able to target and eradicate the bacteria in the infected host cells may represent a therapeutic breakthrough while avoiding latency. The development of nanosystems aiming a controlled and targeted intracellular drug release, have proved to increase cytosolic therapeutic concentration while reducing undesired side effects. This work’s main goal was to develop a host-directed strategy against mycobacterial infection through the design of a biocompatible nanocarrier for phage-derived protein delivery, using M. smegmatis as model. Since mycobacterial pathogenicity is strongly supported by the presence of lipids in the cell wall, their degradation induces bacterial destruction through cell wall hydrolysis. Phage-based lipolytic enzymes such as, LysB a mycolylarabinogalactan esterase, represent an appealing therapeutic approach. The herein proposed Ms6 LysB-containing lipid nanocarrier (SLN-LysB) explores the known advantages of nanomedicine-based systems for phagocytic cells selectively targeting thus allowing LysB intracellular accumulation and a more pronounced mycobacterial infection eradication. Adsorption efficiency value indicates the potential of this system as a protein nanocarrier. Moreover, promising outcomes were obtained in host-infected macrophages treated with SLN-LysB. The results show that the herein proposed strategy was more effective in inhibiting the growth of M. smegmatis than free LysB, which might be related to the nanocarrier internalization. Acting as effective protein nanocarriers, the protein-guided delivery in the infected phagocytic cells allows it to exert its hydrolytic action on the lipid layer of the Mycobacterium

  6. Direct molecular mass determination of trehalose monomycolate from 11 species of mycobacteria by MALDI-TOF mass spectrometry.

    Science.gov (United States)

    Fujita, Yukiko; Naka, Takashi; Doi, Takeshi; Yano, Ikuya

    2005-05-01

    Direct estimation of the molecular mass of single molecular species of trehalose 6-monomycolate (TMM), a ubiquitous cell-wall component of mycobacteria, was performed by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. When less than 1 microg TMM was analysed by MALDI-TOF mass spectrometry, quasimolecular ions [M+Na]+ of each molecular species were demonstrated and the numbers of carbons and double bonds (or cyclopropane rings) were determined. Since the introduction of oxygen atoms such as carbonyl, methoxy and ester groups yielded the appropriate shift of mass ions, the major subclasses of mycolic acid (alpha, methoxy, keto and wax ester) were identified without resorting to hydrolytic procedures. The results showed a marked difference in the molecular species composition of TMM among mycobacterial species. Unexpectedly, differing from other mycoloyl glycolipids, TMM from Mycobacterium tuberculosis showed a distinctive mass pattern, with abundant odd-carbon-numbered monocyclopropanoic (or monoenoic) alpha-mycolates besides dicyclopropanoic mycolate, ranging from C75 to C85, odd- and even-carbon-numbered methoxymycolates ranging from C83 to C94 and even- and odd-carbon-numbered ketomycolates ranging from C83 to C90. In contrast, TMM from Mycobacterium bovis (wild strain and BCG substrains) possessed even-carbon-numbered dicyclopropanoic alpha-mycolates. BCG Connaught strain lacked methoxymycolates almost completely. These results were confirmed by MALDI-TOF mass analysis of mycolic acid methyl esters liberated by alkaline hydrolysis and methylation of the original TMM. Wax ester-mycoloyl TMM molecular species were demonstrated for the first time as an intact form in the Mycobacterium avium-intracellulare group, M. phlei and M. flavescens. The M. avium-intracellulare group possessed predominantly C85 and C87 wax ester-mycoloyl TMM, while M. phlei and the rapid growers tested contained C80, C81, C82 and C83 wax ester

  7. Direct maximum parsimony phylogeny reconstruction from genotype data.

    Science.gov (United States)

    Sridhar, Srinath; Lam, Fumei; Blelloch, Guy E; Ravi, R; Schwartz, Russell

    2007-12-05

    Maximum parsimony phylogenetic tree reconstruction from genetic variation data is a fundamental problem in computational genetics with many practical applications in population genetics, whole genome analysis, and the search for genetic predictors of disease. Efficient methods are available for reconstruction of maximum parsimony trees from haplotype data, but such data are difficult to determine directly for autosomal DNA. Data more commonly is available in the form of genotypes, which consist of conflated combinations of pairs of haplotypes from homologous chromosomes. Currently, there are no general algorithms for the direct reconstruction of maximum parsimony phylogenies from genotype data. Hence phylogenetic applications for autosomal data must therefore rely on other methods for first computationally inferring haplotypes from genotypes. In this work, we develop the first practical method for computing maximum parsimony phylogenies directly from genotype data. We show that the standard practice of first inferring haplotypes from genotypes and then reconstructing a phylogeny on the haplotypes often substantially overestimates phylogeny size. As an immediate application, our method can be used to determine the minimum number of mutations required to explain a given set of observed genotypes. Phylogeny reconstruction directly from unphased data is computationally feasible for moderate-sized problem instances and can lead to substantially more accurate tree size inferences than the standard practice of treating phasing and phylogeny construction as two separate analysis stages. The difference between the approaches is particularly important for downstream applications that require a lower-bound on the number of mutations that the genetic region has undergone.

  8. Direct maximum parsimony phylogeny reconstruction from genotype data

    Directory of Open Access Journals (Sweden)

    Ravi R

    2007-12-01

    Full Text Available Abstract Background Maximum parsimony phylogenetic tree reconstruction from genetic variation data is a fundamental problem in computational genetics with many practical applications in population genetics, whole genome analysis, and the search for genetic predictors of disease. Efficient methods are available for reconstruction of maximum parsimony trees from haplotype data, but such data are difficult to determine directly for autosomal DNA. Data more commonly is available in the form of genotypes, which consist of conflated combinations of pairs of haplotypes from homologous chromosomes. Currently, there are no general algorithms for the direct reconstruction of maximum parsimony phylogenies from genotype data. Hence phylogenetic applications for autosomal data must therefore rely on other methods for first computationally inferring haplotypes from genotypes. Results In this work, we develop the first practical method for computing maximum parsimony phylogenies directly from genotype data. We show that the standard practice of first inferring haplotypes from genotypes and then reconstructing a phylogeny on the haplotypes often substantially overestimates phylogeny size. As an immediate application, our method can be used to determine the minimum number of mutations required to explain a given set of observed genotypes. Conclusion Phylogeny reconstruction directly from unphased data is computationally feasible for moderate-sized problem instances and can lead to substantially more accurate tree size inferences than the standard practice of treating phasing and phylogeny construction as two separate analysis stages. The difference between the approaches is particularly important for downstream applications that require a lower-bound on the number of mutations that the genetic region has undergone.

  9. Direct and inverted repeats elicit genetic instability by both exploiting and eluding DNA double-strand break repair systems in mycobacteria.

    Directory of Open Access Journals (Sweden)

    Ewelina A Wojcik

    Full Text Available Repetitive DNA sequences with the potential to form alternative DNA conformations, such as slipped structures and cruciforms, can induce genetic instability by promoting replication errors and by serving as a substrate for DNA repair proteins, which may lead to DNA double-strand breaks (DSBs. However, the contribution of each of the DSB repair pathways, homologous recombination (HR, non-homologous end-joining (NHEJ and single-strand annealing (SSA, to this sort of genetic instability is not fully understood. Herein, we assessed the genome-wide distribution of repetitive DNA sequences in the Mycobacterium smegmatis, Mycobacterium tuberculosis and Escherichia coli genomes, and determined the types and frequencies of genetic instability induced by direct and inverted repeats, both in the presence and in the absence of HR, NHEJ, and SSA. All three genomes are strongly enriched in direct repeats and modestly enriched in inverted repeats. When using chromosomally integrated constructs in M. smegmatis, direct repeats induced the perfect deletion of their intervening sequences ~1,000-fold above background. Absence of HR further enhanced these perfect deletions, whereas absence of NHEJ or SSA had no influence, suggesting compromised replication fidelity. In contrast, inverted repeats induced perfect deletions only in the absence of SSA. Both direct and inverted repeats stimulated excision of the constructs from the attB integration sites independently of HR, NHEJ, or SSA. With episomal constructs, direct and inverted repeats triggered DNA instability by activating nucleolytic activity, and absence of the DSB repair pathways (in the order NHEJ>HR>SSA exacerbated this instability. Thus, direct and inverted repeats may elicit genetic instability in mycobacteria by 1 directly interfering with replication fidelity, 2 stimulating the three main DSB repair pathways, and 3 enticing L5 site-specific recombination.

  10. Direct and inverted repeats elicit genetic instability by both exploiting and eluding DNA double-strand break repair systems in mycobacteria.

    Science.gov (United States)

    Wojcik, Ewelina A; Brzostek, Anna; Bacolla, Albino; Mackiewicz, Pawel; Vasquez, Karen M; Korycka-Machala, Malgorzata; Jaworski, Adam; Dziadek, Jaroslaw

    2012-01-01

    Repetitive DNA sequences with the potential to form alternative DNA conformations, such as slipped structures and cruciforms, can induce genetic instability by promoting replication errors and by serving as a substrate for DNA repair proteins, which may lead to DNA double-strand breaks (DSBs). However, the contribution of each of the DSB repair pathways, homologous recombination (HR), non-homologous end-joining (NHEJ) and single-strand annealing (SSA), to this sort of genetic instability is not fully understood. Herein, we assessed the genome-wide distribution of repetitive DNA sequences in the Mycobacterium smegmatis, Mycobacterium tuberculosis and Escherichia coli genomes, and determined the types and frequencies of genetic instability induced by direct and inverted repeats, both in the presence and in the absence of HR, NHEJ, and SSA. All three genomes are strongly enriched in direct repeats and modestly enriched in inverted repeats. When using chromosomally integrated constructs in M. smegmatis, direct repeats induced the perfect deletion of their intervening sequences ~1,000-fold above background. Absence of HR further enhanced these perfect deletions, whereas absence of NHEJ or SSA had no influence, suggesting compromised replication fidelity. In contrast, inverted repeats induced perfect deletions only in the absence of SSA. Both direct and inverted repeats stimulated excision of the constructs from the attB integration sites independently of HR, NHEJ, or SSA. With episomal constructs, direct and inverted repeats triggered DNA instability by activating nucleolytic activity, and absence of the DSB repair pathways (in the order NHEJ>HR>SSA) exacerbated this instability. Thus, direct and inverted repeats may elicit genetic instability in mycobacteria by 1) directly interfering with replication fidelity, 2) stimulating the three main DSB repair pathways, and 3) enticing L5 site-specific recombination.

  11. Mycobacteria of clinical interest

    International Nuclear Information System (INIS)

    Casal, M.

    1986-01-01

    This book is based upon a symposium on mycobacteria of clinical interest. Due to the multidisciplinary participation of, among others, microbiologists, clinicians, immunologists and epidemiologists, a very wide and thorough presentation of the present state of clinical research in this field is ensured. Topics of particular interest included in this volume were the new antimicrobial agents active against mycobacteria; new therapeutic possibilities; a system of rapid diagnosis of tuberculosis and mycobacteriosis; mycobacteriosis in AIDS; progress in immunopathology of tuberculosis and leprosy; progress in bacteriology and vaccination in leprosy; progress in immunological diagnosis and new epidemiological biovars of M. tuberculosis. (Auth.)

  12. Macrolide Resistance in Mycobacteria

    Czech Academy of Sciences Publication Activity Database

    Doucet-Populaire, F.; Buriánková, Karolína; Weiser, Jaroslav; Pernodet, J.-L.

    2005-01-01

    Roč. 2, - (2005), s. 511-523 ISSN 0198-6325 R&D Projects: GA ČR GA310/03/0292 Institutional research plan: CEZ:AV0Z50200510 Keywords : mycobacteria * mycobacterium tuberculosis * ribosome Subject RIV: EE - Microbiology, Virology Impact factor: 7.964, year: 2005

  13. HIV and mycobacteria.

    Science.gov (United States)

    Procop, Gary W

    2017-07-01

    The importance of mycobacteria as opportunistic pathogens, particularly members of the M. avium complex (MAC), in patients with progressive HIV infection was recognized early in the AIDS epidemic. It took longer to appreciate the global impact and devastation that would result from the deadly synergy that exists between HIV and M. tuberculosis. This HIV/M. tuberculosis co-pandemic is ongoing and claiming millions of lives every year. In addition to MAC, a number of other non-tuberculous mycobacteria have been recognized as opportunistic pathogens in HIV-infected individuals; some of these are more commonly encountered (e.g., M. kansasii) than others (M. haemophilum and M. genevense). Finally, there are challenges to concomitantly treating the HIV and the infecting Mycobacterium species, because of antimicrobial resistance, therapeutic side-effects and the complex pharmacologic interactions of the antiretroviral and antimycobacterial multidrug therapy. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Direct maximum parsimony phylogeny reconstruction from genotype data

    OpenAIRE

    Sridhar, Srinath; Lam, Fumei; Blelloch, Guy E; Ravi, R; Schwartz, Russell

    2007-01-01

    Abstract Background Maximum parsimony phylogenetic tree reconstruction from genetic variation data is a fundamental problem in computational genetics with many practical applications in population genetics, whole genome analysis, and the search for genetic predictors of disease. Efficient methods are available for reconstruction of maximum parsimony trees from haplotype data, but such data are difficult to determine directly for autosomal DNA. Data more commonly is available in the form of ge...

  15. Nontuberculous mycobacteria in the environment.

    Science.gov (United States)

    Falkinham, Joseph O

    2002-09-01

    It is likely that the incidence of infection by environmental opportunistic mycobacteria will continue to rise. Part of the rise will be caused by the increased awareness of these microbes as human pathogens and improvements in methods of detection and culture. Clinicians and microbiologists will continue to be challenged by the introduction of new species to the already long list of mycobacterial opportunists (see Table 3). The incidence of infection will also rise because an increasing proportion of the population is aging or subject to some type of immunosuppression. A second reason for an increase in the incidence of environmental mycobacterial infection is that these microbes are everywhere. They are present in water, biofilms, soil, and aerosols. They are natural inhabitants of the human environment, especially drinking water distribution systems. Thus, it is likely that everyone is exposed on a daily basis. It is likely that certain human activities can lead to selection of mycobacteria. Important lessons have been taught by study of cases of hypersensitivity pneumonitis associated with exposure to metalworking fluid. First, the implicated metalworking fluids contained water, the likely source of the mycobacteria. Second, the metalworking fluids contain hydrocarbons (e.g., pine oils) and biocides (e.g., morpholine) both of which are substrates for the growth of mycobacteria [53,193]. Third, outbreak of disease followed disinfection of the metalworking fluid [136,137]. Although the metalworking fluid was contaminated with microorganisms, it was only after disinfection that symptoms developed in the workers. Because mycobacteria are resistant to disinfectants, it is likely that the recovery of the mycobacteria from the metalworking fluid [137] was caused by their selection. Disinfection may also contribute, in part, to the persistence of M avium and M intracellulare in drinking water distribution systems [33,89,240]. M avium and M intracellulare are many times

  16. Life-threatening mycobacteria infection

    NARCIS (Netherlands)

    Faber, William R.

    2005-01-01

    Most mycobacteria cause localized and often harmless infections of the skin, Leprosy. which dates back to approximately 60 BC in India,(1) was supposed to be eliminated as a Public health problem by the year 2000.(2) With a new case detection rate between 600,000 and 700,000 yearly, however.(3)

  17. Response of rice genotypes to weed competition in dry direct-seeded rice in India.

    Science.gov (United States)

    Mahajan, Gulshan; Ramesha, Mugalodi S; Chauhan, Bhagirath S

    2014-01-01

    The differential weed-competitive abilities of eight rice genotypes and the traits that may confer such attributes were investigated under partial weedy and weed-free conditions in naturally occurring weed flora in dry direct-seeded rice during the rainy seasons of 2011 and 2012 at Ludhiana, Punjab, India. The results showed genotypic differences in competitiveness against weeds. In weed-free plots, grain yield varied from 6.6 to 8.9 t ha(-1) across different genotypes; it was lowest for PR-115 and highest for the hybrid H-97158. In partial weedy plots, grain yield and weed biomass at flowering varied from 3.6 to 6.7 t ha(-1) and from 174 to 419 g m(-2), respectively. In partial weedy plots, grain yield was lowest for PR-115 and highest for PR-120. Average yield loss due to weed competition ranged from 21 to 46% in different rice genotypes. The study showed that early canopy closure, high leaf area index at early stage, and high root biomass and volume correlated positively with competitiveness. This study suggests that some traits (root biomass, leaf area index, and shoot biomass at the early stage) could play an important role in conferring weed competitiveness and these traits can be explored for dry-seeded rice.

  18. Mycobacteria in nail salon whirlpool footbaths, California.

    Science.gov (United States)

    Vugia, Duc J; Jang, Yvonne; Zizek, Candi; Ely, Janet; Winthrop, Kevin L; Desmond, Edward

    2005-04-01

    In 2000, an outbreak of Mycobacterium fortuitum furunculosis affected customers using whirlpool footbaths at a nail salon. We swabbed 30 footbaths in 18 nail salons from 5 California counties and found mycobacteria in 29 (97%); M. fortuitum was the most common. Mycobacteria may pose an infectious risk for pedicure customers.

  19. Prediction of direct and indirect genetic gains and genotypic correlations in rubber tree progenies

    Directory of Open Access Journals (Sweden)

    Cecília Khusala Verardi

    2011-09-01

    Full Text Available The objective of this work was to estimate the genetic parameters, genotypic and phenotypic correlations, and direct and indirect genetic gains among and within rubber tree (Hevea brasiliensis progenies. The experiment was set up at the Municipality of Jaú, SP, Brazil. A randomized complete block design was used, with 22 treatments (progenies, 6 replicates, and 10 plants per plot at a spacing of 3x3 m. Three‑year‑old progenies were assessed for girth, rubber yield, and bark thickness by direct and indirect gains and genotypic correlations. The number of latex vessel rings showed the best correlations, correlating positively and significantly with girth and bark thickness. Selection gains among progenies were greater than within progeny for all the variables analyzed. Total gains obtained were high, especially for girth increase and rubber yield, which were 93.38 and 105.95%, respectively. Young progeny selection can maximize the expected genetic gains, reducing the rubber tree selection cycle.

  20. Natural Disasters and Nontuberculous Mycobacteria

    Science.gov (United States)

    Bernhard, Jon N.; Chan, Edward D.

    2015-01-01

    Infectious diseases acquired by survivors of large-scale natural disasters complicate the recovery process. During events such as tsunamis, hurricanes, earthquakes, and tornados and well into the recovery period, victims often are exposed to water-soil mixtures that have relocated with indigenous microbes. Because nontuberculous mycobacteria (NTM) are ubiquitous in water and soil, there is potential for increased exposure to these organisms during natural disasters. In this hypothesis-driven commentary, we discuss the rise in NTM lung disease and natural disasters and examine the geographic overlap of NTM infections and disaster frequencies in the United States. Moreover, we show an increased number of positive NTM cultures from Louisiana residents in the years following three of the relatively recent epic hurricanes and posit that such natural disasters may help to drive the increased number of NTM infections. Finally, we advocate for increased environmental studies and surveillance of NTM infections before and after natural disasters. PMID:25644904

  1. Sinusitis from Nontuberculous Mycobacteria in Tap Water

    Centers for Disease Control (CDC) Podcasts

    Dr. Wellington S. Tichenor. Associate Clinical Professor of Medicine at New York Medical College and in private practice in Manhattan, New York, discusses his investigation of sinusitis from nontuberculous mycobacteria in tap water.

  2. The effects of stabilizing and directional selection on phenotypic and genotypic variation in a population of RNA enzymes.

    Science.gov (United States)

    Hayden, Eric J; Bratulic, Sinisa; Koenig, Iwo; Ferrada, Evandro; Wagner, Andreas

    2014-02-01

    The distribution of variation in a quantitative trait and its underlying distribution of genotypic diversity can both be shaped by stabilizing and directional selection. Understanding either distribution is important, because it determines a population's response to natural selection. Unfortunately, existing theory makes conflicting predictions about how selection shapes these distributions, and very little pertinent experimental evidence exists. Here we study a simple genetic system, an evolving RNA enzyme (ribozyme) in which a combination of high throughput genotyping and measurement of a biochemical phenotype allow us to address this question. We show that directional selection, compared to stabilizing selection, increases the genotypic diversity of an evolving ribozyme population. In contrast, it leaves the variance in the phenotypic trait unchanged.

  3. Mycobacteria in Finnish cooling tower waters.

    Science.gov (United States)

    Torvinen, Eila; Suomalainen, Sini; Paulin, Lars; Kusnetsov, Jaana

    2014-04-01

    Evaporative cooling towers are water systems used in, e.g., industry and telecommunication to remove excess heat by evaporation of water. Temperatures of cooling waters are usually optimal for mesophilic microbial growth and cooling towers may liberate massive amounts of bacterial aerosols. Outbreaks of legionellosis associated with cooling towers have been known since the 1980's, but occurrences of other potentially pathogenic bacteria in cooling waters are mostly unknown. We examined the occurrence of mycobacteria, which are common bacteria in different water systems and may cause pulmonary and other soft tissue infections, in cooling waters containing different numbers of legionellae. Mycobacteria were isolated from all twelve cooling systems and from 92% of the 24 samples studied. Their numbers in the positive samples varied from 10 to 7.3 × 10(4) cfu/L. The isolated species included M. chelonae/abscessus, M. fortuitum, M. mucogenicum, M. peregrinum, M. intracellulare, M. lentiflavum, M. avium/nebraskense/scrofulaceum and many non-pathogenic species. The numbers of mycobacteria correlated negatively with the numbers of legionellae and the concentration of copper. The results show that cooling towers are suitable environments for potentially pathogenic mycobacteria. Further transmission of mycobacteria from the towers to the environment needs examination. © 2013 APMIS. Published by John Wiley & Sons Ltd.

  4. Molecular epidemiology of Bordetella pertussis in Cambodia determined by direct genotyping of clinical specimens.

    Science.gov (United States)

    Moriuchi, Takumi; Vichit, Ork; Vutthikol, Yong; Hossain, Md Shafiqul; Samnang, Chham; Toda, Kohei; Grabovac, Varja; Hiramatsu, Yukihiro; Otsuka, Nao; Shibayama, Keigo; Kamachi, Kazunari

    2017-09-01

    This study sought to determine the genotypes of circulating Bordetella pertussis, the causative agent of pertussis, in Cambodia by direct molecular typing of clinical specimens. DNA extracts from nasopharyngeal swabs obtained from 82 pertussis patients in 2008-2016 were analyzed by multilocus variable-number tandem repeat analysis (MLVA). B. pertussis virulence-associated allelic genes (ptxA, prn, and fim3) and the pertussis toxin promoter ptxP were also investigated by DNA sequence-based typing. Forty-four DNA extracts (54%) yielded a complete MLVA profile, and these were sorted into 8 MLVA types (MT18, MT26, MT27, MT29, MT43, MT72, MT95, and MT200). MT27 and MT29, which are common in developed countries, were the predominant strain types (total 73%). The predominant profile of virulence-associated allelic genes was the combination of ptxP3/ptxA1/prn2/fim3A (48%). MT27 strains were detected during the entire study period, whereas MT29 strains were only found in 2014-2016. The B. pertussis population in Cambodia, where a whole-cell pertussis vaccine (WCV) has been continuously used, resembled those observed previously in developed countries where acellular pertussis vaccines are used. Circulating B. pertussis strains in Cambodia were distinct from those in other countries using WCVs. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

  5. Impact of hepatitis C virus genotype-4 eradication following direct acting antivirals on liver stiffness measurement

    Directory of Open Access Journals (Sweden)

    Tag-Adeen M

    2017-10-01

    Full Text Available Mohammed Tag-Adeen,1,2 Ahlam Mohamed Sabra,1 Yuko Akazawa,2 Ken Ohnita,2 Kazuhiko Nakao2 1Department of Internal Medicine, Qena School of Medicine, South Valley University, Qena, Egypt; 2Department of Gastroenterology and Hepatology, Nagasaki School of Biomedical Sciences, Nagasaki University, Nagasaki, Japan Background: Liver fibrosis is the most important prognostic factor in chronic hepatitis C virus (HCV patients, and Egypt shows the highest worldwide HCV prevalence with genotype-4 predominance. The aim of this study was to investigate the degree of liver stiffness measurement (LSM improvement after successful HCV eradication.Patients and methods: The study included 84 chronic HCV Egyptian patients, and was conducted at Qena University Hospital from November 1, 2015 till October 31, 2016. LSM was obtained by FibroScan® before starting direct acting antiviral (DAA treatment and after achieving sustained virologic response-24 (SVR-24. Based on baseline LSM, patients were stratified into F0–F1, F2, F3 and F4 groups (METAVIR. LSM and laboratory data after achieving SVR-24 was compared with that before starting therapy in each fibrosis group (F0-F4, p-value <0.05 was statistically significant.Results: Following DAA treatment, 80 patients achieved SVR-24; of these, 50 were males (62.5%, mean age: 54.2±7.6 years, and mean body mass index: 28.6±2.2 kg/m2. Mean baseline LSM dropped from 15.6±10.8 to 12.1±8.7 kPa post-SVR; the maximum change of −5.8 occurred in F4 versus −2.79, −1.28 and +0.08 in F3, F2 and F0–F1 respectively (p<0.0001. At baseline, 41 patients were in the F4 group; only 16 (39% regressed to non-cirrhotic range (<12.5 kPa, while 25 (61% were still cirrhotic despite achieving SVR-24 (p<0.0001. Patients who achieved LSM improvement (n=64 have had significantly higher baseline aspartate transferase (AST and alanine transaminase (ALT. Also, those patients showed significant improvement in AST, AST/platelets ratio index

  6. Sinusitis from Nontuberculous Mycobacteria in Tap Water

    Centers for Disease Control (CDC) Podcasts

    2012-12-21

    Dr. Wellington S. Tichenor. Associate Clinical Professor of Medicine at New York Medical College and in private practice in Manhattan, New York, discusses his investigation of sinusitis from nontuberculous mycobacteria in tap water.  Created: 12/21/2012 by National Center for Emerging and Zoonotic Infectious Diseases (NCEZID).   Date Released: 12/31/2012.

  7. Tuberculosis and non-tuberculous mycobacteria among HIV-infected individuals in Ghana

    DEFF Research Database (Denmark)

    Bjerrrum, Stephanie; Oliver-Commey, Joseph; Kenu, Ernest

    2016-01-01

    OBJECTIVES: To assess the prevalence and clinical importance of previously unrecognised tuberculosis (TB) and isolation of non-tuberculous mycobacteria (NTM) among HIV-infected individuals in a teaching hospital in Ghana. METHODS: Intensified mycobacterial case finding was conducted among HIV...... for mycobacteria with smear microscopy, culture and Xpert MTB/RIF assay. NTM species were identified with the GenoType Mycobacterium CM/AS or sequence analysis of 16S rRNA gene. RESULTS: Of 473 participants, 60 (12.7%) had confirmed pulmonary TB, and 38 (8.0%) had positive cultures for NTM. Mycobacterium avium...... cell count, BMI, prolonged fever and ART initiation. CONCLUSIONS: Intensified mycobacterial screening of HIV-infected individuals revealed a high burden of unrecognised pulmonary TB before ART initiation, which increased risk of death within six months. NTM were frequently isolated and associated...

  8. Highly productive mutant genotypes in barley - direct use in practice and in successive recombination

    International Nuclear Information System (INIS)

    Gustafsson, Aa.; Lundqvist, U.

    1984-01-01

    Three special cases of induced mutations in barley are discussed in this paper. They are denoted here as the Gunilla, the Pallas and the Mari cases, after the three named varieties to which the original mutants gave rise. The original mutants described represent just a small sample of the induced mutants, many of which have been tested in practice and have been further studied in basic genetics and evolutionary research. The three approved varieties have given rise to further recombination families, which also to some extent have been fused. Two of the mutant cases - Pallas and Mari - were directly useful in practice and officially approved. The third case involved a mutant of special appearance - a ''bushy type'' with an intense blue wax coating and with a supreme lodging resistance. The mutant was used in developing the Gunilla variety, which arose by recombination breeding. This variety has been highly satisfactory in further gene recombination work. A similar situation has prevailed with regard to the Pallas and Mari families arising after gene recombination, too. Up to now, the Gunilla, Pallas and Mari families include a long series of released and officially approved varieties. Several of them represent valuable agricultural contributions with wide areas of cultivation. These three mutants - with their recombination families - led to greatly increased straw stiffness and high grain production. Their phenotypic expression often corresponds to a dwarf or semidwarf description. One of the mutants - the Mari genotype - represents a group of genes and alleles which give rise to profound changes in the photoperiod (and partially also in the thermoperiod) behaviour. In fact, often even such small changes have a fundamental influence on adaptation and distribution. Data are presented analysing the property of lodging resistance with the background of plant, tiller and internode structure. A method of partial back-mutation was worked out in separating traits generally

  9. COMT Val(158)Met genotype determines the direction of cognitive effects produced by catechol-O-methyltransferase inhibition.

    Science.gov (United States)

    Farrell, Sarah M; Tunbridge, Elizabeth M; Braeutigam, Sven; Harrison, Paul J

    2012-03-15

    Catechol-O-methyltransferase (COMT) metabolizes dopamine. The COMT Val(158)Met polymorphism influences its activity, and multiple neural correlates of this genotype on dopaminergic phenotypes, especially working memory, have been reported. COMT activity can also be regulated pharmacologically by COMT inhibitors. The inverted-U relationship between cortical dopamine signaling and working memory predicts that the effects of COMT inhibition will differ according to COMT genotype. Thirty-four COMT Met(158)Met (Met-COMT) and 33 COMT Val(158)Val (Val-COMT) men were given a single 200-mg dose of the brain-penetrant COMT inhibitor tolcapone or placebo in a randomized, double-blind, between-subjects design. They completed the N-back task of working memory and a gambling task. In the placebo group, Met-COMT subjects outperformed Val-COMT subjects on the 2- back, and they were more risk averse. Tolcapone had opposite effects in the two genotype groups: it worsened N-back performance in Met-COMT subjects but enhanced it in Val-COMT subjects. Tolcapone made Met-COMT subjects less risk averse but Val-COMT subjects more so. In both tasks, tolcapone reversed the baseline genotype differences. Depending on genotype, COMT inhibition can enhance or impair working memory and increase or decrease risky decision making. To our knowledge, the data are the clearest demonstration to date that the direction of effect of a drug can be influenced by a polymorphism in its target gene. The results support the inverted-U model of dopamine function. The findings are of translational relevance, because COMT inhibitors are used in the adjunctive treatment of Parkinson's disease and are under evaluation in schizophrenia and other disorders. Copyright © 2012 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

  10. 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.

  11. Enhanced capillary electrophoretic screening of Alzheimer based on direct apolipoprotein E genotyping and one-step multiplex PCR.

    Science.gov (United States)

    Woo, Nain; Kim, Su-Kang; Sun, Yucheng; Kang, Seong Ho

    2018-01-01

    Human apolipoprotein E (ApoE) is associated with high cholesterol levels, coronary artery disease, and especially Alzheimer's disease. In this study, we developed an ApoE genotyping and one-step multiplex polymerase chain reaction (PCR) based-capillary electrophoresis (CE) method for the enhanced diagnosis of Alzheimer's. The primer mixture of ApoE genes enabled the performance of direct one-step multiplex PCR from whole blood without DNA purification. The combination of direct ApoE genotyping and one-step multiplex PCR minimized the risk of DNA loss or contamination due to the process of DNA purification. All amplified PCR products with different DNA lengths (112-, 253-, 308-, 444-, and 514-bp DNA) of the ApoE genes were analyzed within 2min by an extended voltage programming (VP)-based CE under the optimal conditions. The extended VP-based CE method was at least 120-180 times faster than conventional slab gel electrophoresis methods In particular, all amplified DNA fragments were detected in less than 10 PCR cycles using a laser-induced fluorescence detector. The detection limits of the ApoE genes were 6.4-62.0pM, which were approximately 100-100,000 times more sensitive than previous Alzheimer's diagnosis methods In addition, the combined one-step multiplex PCR and extended VP-based CE method was also successfully applied to the analysis of ApoE genotypes in Alzheimer's patients and normal samples and confirmed the distribution probability of allele frequencies. This combination of direct one-step multiplex PCR and an extended VP-based CE method should increase the diagnostic reliability of Alzheimer's with high sensitivity and short analysis time even with direct use of whole blood. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Mycobacteria inactivation using Engineered Water Nanostructures (EWNS).

    Science.gov (United States)

    Pyrgiotakis, Georgios; McDevitt, James; Gao, Ya; Branco, Alan; Eleftheriadou, Mary; Lemos, Bernardo; Nardell, Edward; Demokritou, Philip

    2014-08-01

    Airborne transmitted pathogens such as Mycobacterium tuberculosis (Mtb) cause serious, often fatal infectious disease with enormous global health implications. Due to their unique cell wall and slow growth, mycobacteria are among the most resilient microbial forms. Herein we evaluate the ability of an emerging, chemical-free, nanotechnology-based method to inactivate M. parafortuitum (Mtb surrogate). This method is based on the transformation of atmospheric water vapor into engineered water nano-structures (EWNS) via electrospray. We demonstrate that the EWNS can interact with and inactivate airborne mycobacteria, reducing their concentration levels significantly. Additionally, EWNS can inactivate M. parafortuitum on surfaces eight times faster than the control. The mechanism of mycobacteria inactivation was also investigated in this study. It was demonstrated that the EWNS effectively deliver the reactive oxygen species, encapsulated during the electrospray process, to the bacteria oxidizing their cell membrane resulting into inactivation. Overall, this is a method with the potential to become an effective intervention technology in the battle against airborne infections. This study demonstrates the feasibility of mycobacterium inactivation in airborne form or on contact surfaces using electrospray activated water nano-structures. Given that the method is free of toxic chemicals, this might become an important tool in the prevention of mycobacterial infections, which are notoriously hard to treat. Copyright © 2014 Elsevier Inc. All rights reserved.

  13. Nontuberculous mycobacteria in captive and pet reptiles

    Directory of Open Access Journals (Sweden)

    Irena Reil

    2017-01-01

    Full Text Available The aim of this study was to highlight the importance of nontuberculous mycobacteria species in the pathology of various reptilian pet species as well as their epidemiological significance of infection transmission to humans. Faeces samples from six living reptiles and organs from ten carcasses were submitted to bacteriological testing during the years 2003–2015. Positive colonies from one faeces sample and two organs showed the presence of a gene coding 65kDa antigen common for all mycobacteria. Further identification to the species level revealed that the isolates belong to Mycobacterium fortuitum and Mycobacterium avium subsp. hominissuis, later subjected to drug susceptibility testing which confirmed high resistance levels in both isolates. In conclusion, there is a great significance of the occurrence of nontuberculous mycobacteria in captive and pet reptiles, presenting reptiles as possible hosts representing a serious threat of transmission of high resistance mycobacterial isolates to humans. To our knowledge, this is the first report of M. avium subsp. hominissuis occurrence in reptiles.

  14. 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

  15. High prevalence of Human Parechovirus (HPeV) genotypes in the Amsterdam region and the identification of specific HPeV variants by direct genotyping of stool samples

    NARCIS (Netherlands)

    Benschop, K.; Thomas, X.; Serpenti, C.; Molenkamp, R.; Wolthers, K.

    2008-01-01

    Human Parechoviruses (HPeV) are widespread pathogens belonging to the Picornavirus family. Six genotypes are known which have predominantly been isolated from children. Data on prevalence of HPeV genotypes are generally based on cell culture, which may underestimate the prevalence of certain HPeV

  16. An update on mycobacteria and the development of allergic diseases

    NARCIS (Netherlands)

    Folkerts, G.

    2015-01-01

    Mycobacteria can diminish allergic and asthmatic manifestations. This means that mycobacteria could offer therapeutical opportunities as an 'anti-allergic' vaccine.In humans, the genetic background and the environment probably contribute to the development of allergies. Over the last 20. years, a

  17. Mycobacterium bovis BCG mycobacteria--new application.

    Science.gov (United States)

    Kowalewicz-Kulbat, Magdalena; Pestel, Joël; Biet, Franck; Locht, Camille; Tonnel, André-Bernard; Druszczyńska, Magdalena; Rudnicka, Wiesława

    2006-01-01

    The polarized response of T helper-2 (Th2) lymphocytes to an allergen is considered to be the main cause of the pathogenesis of asthma. In this study, we asked a question whether M. bovis BCG mycobacteria which are known for the preferential stimulation of T helper-1 (Th1) immunity, diminish the effector functions of Th2 cells from allergic patients upon stimulation with a common house dust mite Der p-1 allergen. Our results allow a positive answer to this question. We demonstrate that BCG modulates the dendritic cell-dependent allergen presentation process and switches naive T lymphocytes towards an anti-allergic Th1 profile.

  18. Short communication: Genotyping of cows to speed up availability of genomic estimated breeding values for direct health traits in Austrian Fleckvieh (Simmental) cattle--genetic and economic aspects.

    Science.gov (United States)

    Egger-Danner, C; Schwarzenbacher, H; Willam, A

    2014-07-01

    The aim of this study was to quantify the impact of genotyping cows with reliable phenotypes for direct health traits on annual monetary genetic gain (AMGG) and discounted profit. The calculations were based on a deterministic approach using ZPLAN software (University of Hohenheim, Stuttgart, Germany). It was assumed that increases in reliability of the total merit index (TMI) of 5, 15, and 25 percentage points were achieved through genotyping 5,000, 25,000, and 50,000 cows, respectively. Costs for phenotyping, genotyping, and genomic estimated breeding values vary between €150 and €20 per cow. The gain in genotyping cows for traits with medium to high heritability is more than for direct health traits with low heritability. The AMGG is increased by 1.5% if the reliability of TMI is 5 percentage points higher (i.e., 5,000 cows genotyped) and 6.53% higher AMGG can be expected when the reliability of TMI is increased by 25 percentage points (i.e., 50,000 cows genotyped). The discounted profit depends not only on the costs of genotyping but also on the population size. This study indicates that genotyping cows with reliable phenotypes is feasible to speed up the availability of genomic estimated breeding values for direct health traits. But, because of the huge amount of valid phenotypes and genotypes needed to establish an efficient genomic evaluation, it is likely that financial constraints will be the main limiting factor for implementation into breeding program such as Fleckvieh Austria. Copyright © 2014 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  19. Learning epigenetic regulation from mycobacteria

    Directory of Open Access Journals (Sweden)

    Sanjeev Khosla

    2016-01-01

    Full Text Available In a eukaryotic cell, the transcriptional fate of a gene is determined by the profile of the epigenetic modifications it is associated with and the conformation it adopts within the chromatin. Therefore, the function that a cell performs is dictated by the sum total of the chromatin organization and the associated epigenetic modifications of each individual gene in the genome (epigenome. As the function of a cell during development and differentiation is determined by its microenvironment, any factor that can alter this microenvironment should be able to alter the epigenome of a cell. In the study published in Nature Communications (Yaseen [2015] Nature Communications 6:8922 doi: 10.1038/ncomms9922, we show that pathogenic Mycobacterium tuberculosis has evolved strategies to exploit this pliability of the host epigenome for its own survival. We describe the identification of a methyltransferase from M. tuberculosis that functions to modulate the host epigenome by methylating a novel, non-canonical arginine, H3R42 in histone H3. In another study, we showed that the mycobacterial protein Rv2966c methylates cytosines present in non-CpG context within host genomic DNA upon infection. Proteins with ability to directly methylate host histones H3 at a novel lysine residue (H3K14 has also been identified from Legionella pnemophilia (RomA. All these studies indicate the use of non-canonical epigenetic mechanisms by pathogenic bacteria to hijack the host transcriptional machinery.

  20. 'Emerging' mycobacteria in South Africa : review article

    Directory of Open Access Journals (Sweden)

    P.D. Van Helden

    2009-05-01

    Full Text Available Disease can be caused by various species of the genus Mycobacterium. A number of reports, both published and unpublished, of rarely reported mycobacteria have surfaced in South Africa in the last few years. Some unusual hosts have also been involved, causing concern in some quarters.These include reports on Mycobacterium goodii in a spotted hyaena (Crocuta crocuta, M. xenopi in a ruffed lemur (Varecia variegata, M. intracellulare in wild-caught chacma baboons (Papio ursinus, the 'dassie bacillus' in free ranging rock hyrax (dassies; Procavia capensis the 'oryx bacillus' from free-ranging buffalo (Syncerus caffer and M. tuberculosis in suricates (Suricata suricatta, a domestic dog and in baboons. In this article it has been attempted to put these in context and show how improved surveillance and technologies have allowed mycobacteria to be identified to species level more easily. Most of the unusual mycobacterial species have most likely been present in the region for many years and have probably caused disease episodes before, but have been misdiagnosed. Each case must be evaluated carefully with respect to the animal species involved, the environment in which the host is found and the mycobacterial species, and operational decisions made accordingly.

  1. AAA+ Machines of Protein Destruction in Mycobacteria.

    Science.gov (United States)

    Alhuwaider, Adnan Ali H; Dougan, David A

    2017-01-01

    The bacterial cytosol is a complex mixture of macromolecules (proteins, DNA, and RNA), which collectively are responsible for an enormous array of cellular tasks. Proteins are central to most, if not all, of these tasks and as such their maintenance (commonly referred to as protein homeostasis or proteostasis) is vital for cell survival during normal and stressful conditions. The two key aspects of protein homeostasis are, (i) the correct folding and assembly of proteins (coupled with their delivery to the correct cellular location) and (ii) the timely removal of unwanted or damaged proteins from the cell, which are performed by molecular chaperones and proteases, respectively. A major class of proteins that contribute to both of these tasks are the AAA+ (ATPases associated with a variety of cellular activities) protein superfamily. Although much is known about the structure of these machines and how they function in the model Gram-negative bacterium Escherichia coli , we are only just beginning to discover the molecular details of these machines and how they function in mycobacteria. Here we review the different AAA+ machines, that contribute to proteostasis in mycobacteria. Primarily we will focus on the recent advances in the structure and function of AAA+ proteases, the substrates they recognize and the cellular pathways they control. Finally, we will discuss the recent developments related to these machines as novel drug targets.

  2. Prevalence of Nontuberculous Mycobacteria among Extrapulmonary Tuberculosis Cases in Tertiary Care Centers in Northern India

    Directory of Open Access Journals (Sweden)

    A. K. Maurya

    2015-01-01

    Full Text Available The reports of nontuberculous mycobacteria (NTM associated with extrapulmonary diseases are increasing in tertiary care hospitals. Despite a significant increase in knowledge about NTM infections, they still represent a diagnostic and therapeutic challenge. The aim of this study is to know the prevalence of NTN among extrapulmonary tuberculosis cases in tertiary care centers in Northern India. A total of 227 culture positive isolates from 756 cases were tested for niacin production and catalase assay. BIO-LINE SD Ag MPT64 TB test and final identification and differentiation between MTBC and different species of NTM were further confirmed by GenoType Mycobacterium CM/AS assay. 71 cases (9.3% were positive for AFB by ZN staining and 227 cases (30.1% were positive for mycobacteria by culture. Niacin production and catalase activity were negative in 62/227 (27.4% strains and after using a panel of different biochemicals and final confirmation by GenoType Mycobacterium CM assay. Out of 227 cultures tested, 165 (72.6% strains were confirmed as M. tuberculosis complex, and 62 (27.4% were confirmed as NTM. The most common NTM species identified were M. fortuitum 17 (27.5% and M. intracellulare 13 (20.9%. The rapid identification of NTM species may help in targeted therapy and management of the diseases.

  3. Multiplicative Genotype-Environment Interaction as a Cause of Reversed Response to Directional Selection

    OpenAIRE

    Gimelfarb, A.

    1986-01-01

    In experiments with directional selection on a quantitative character a "reversed response" to selection is occasionally observed, when selection of individuals for a higher (lower) value of the character results in a lower (higher) value of the character among their offspring. A sudden change in environments or random drift is often assumed to be responsible for this. It is demonstrated in this paper that these two causes cannot account for the reversed response at least in some of the exper...

  4. UGT1A1 (TA)n genotyping in sickle-cell disease: high resolution melting (HRM) curve analysis or direct sequencing, what is the best way?

    Science.gov (United States)

    Thomas, Vincent; Mazard, Blandine; Garcia, Caroline; Lacan, Philippe; Gagnieu, Marie-Claude; Joly, Philippe

    2013-09-23

    Minucci et al. have proposed in 2010 a rapid, simple and cost-effective HRM method on the LightCycler 480® apparatus (Roche) for the determination of the 6/6, 6/7 and 7/7 genotypes of the (TA)n UGT1A1 promoter polymorphism. However, they have not studied the n=5 and n=8 alleles which can be quite frequent in sickle-cell disease patients. The aim of our study was to test this HRM protocol to all the 10 possible (TA)n UGT1A1 genotypes (i.e. 5/5, 5/6, 5/7, 5/8, 6/6, 6/7, 6/8, 7/7, 7/8 and 8/8) by using our SCD cohort of patients. All genotypes could be unambiguously identified except 6/7 and 6/8 which give a similar HRM profile. For those two genotypes, the differentiation necessitates either a direct Sanger sequencing or a second PCR protocol followed by a 3% agarose gel migration. For the (TA)n UGT1A1 promoter genotyping of African patients, each lab has to wonder what is the best way between (i) direct Sanger sequencing of all patients and (ii) HRM protocol for all patients followed by a complementary analysis to differentiate the 6/7 and 6/8 genotypes. © 2013. Published by Elsevier B.V. All rights reserved.

  5. High-resolution melt PCR analysis for genotyping of Ureaplasma parvum isolates directly from clinical samples.

    Science.gov (United States)

    Payne, Matthew S; Tabone, Tania; Kemp, Matthew W; Keelan, Jeffrey A; Spiller, O Brad; Newnham, John P

    2014-02-01

    Ureaplasma sp. infection in neonates and adults underlies a variety of disease pathologies. Of the two human Ureaplasma spp., Ureaplasma parvum is clinically the most common. We have developed a high-resolution melt (HRM) PCR assay for the differentiation of the four serovars of U. parvum in a single step. Currently U. parvum strains are separated into four serovars by sequencing the promoter and coding region of the multiple-banded antigen (MBA) gene. We designed primers to conserved sequences within this region for PCR amplification and HRM analysis to generate reproducible and distinct melt profiles that distinguish clonal representatives of serovars 1, 3, 6, and 14. Furthermore, our HRM PCR assay could classify DNA extracted from 74 known (MBA-sequenced) test strains with 100% accuracy. Importantly, HRM PCR was also able to identify U. parvum serovars directly from 16 clinical swabs. HRM PCR performed with DNA consisting of mixtures of combined known serovars yielded profiles that were easily distinguished from those for single-serovar controls. These profiles mirrored clinical samples that contained mixed serovars. Unfortunately, melt curve analysis software is not yet robust enough to identify the composition of mixed serovar samples, only that more than one serovar is present. HRM PCR provides a single-step, rapid, cost-effective means to differentiate the four serovars of U. parvum that did not amplify any of the known 10 serovars of Ureaplasma urealyticum tested in parallel. Choice of reaction reagents was found to be crucial to allow sufficient sensitivity to differentiate U. parvum serovars directly from clinical swabs rather than requiring cell enrichment using microbial culture techniques.

  6. Consideration of Viral Resistance for Optimization of Direct Antiviral Therapy of Hepatitis C Virus Genotype 1-Infected Patients.

    Science.gov (United States)

    Dietz, Julia; Susser, Simone; Berkowski, Caterina; Perner, Dany; Zeuzem, Stefan; Sarrazin, Christoph

    2015-01-01

    Different highly effective interferon-free treatment options for chronic hepatitis C virus (HCV) infection are currently available. Pre-existence of resistance associated variants (RAVs) to direct antiviral agents (DAAs) reduces sustained virologic response (SVR) rates by 3-53% in hepatitis C virus (HCV) genotype 1 infected patients depending on different predictors and the DAA regimen used. Frequencies of single and combined resistance to NS3, NS5A and NS5B inhibitors and consequences for the applicability of different treatment regimens are unknown. Parallel population based sequencing of HCV NS3, NS5A and NS5B genes in 312 treatment-naïve Caucasian HCV genotype 1 infected patients showed the presence of major resistant variants in 20.5% (NS3), 11.9% (NS5A), and 22.1% (NS5B) with important differences for HCV subtypes. In NS3, Q80K was observed in 34.7% and 2.1% of subtype 1a and 1b patients, respectively while other RAVs to second generation protease inhibitors were detected rarely (1.4%). Within NS5A RAVs were observed in 7.1% of subtype 1a and 17.6% in subtype 1b infected patients. RAVs to non-nucleoside NS5B inhibitors were observed in 3.5% and 44.4% of subtype 1a and 1b patients, respectively. Considering all three DAA targets all subtype 1a and 98.6% of subtype 1b infected patients were wildtype for at least one interferon free DAA regimen currently available. In conclusion, baseline resistance testing allows the selection of at least one RAVs-free treatment option for nearly all patients enabling a potentially cost- and efficacy-optimized treatment of chronic hepatitis C.

  7. Consideration of Viral Resistance for Optimization of Direct Antiviral Therapy of Hepatitis C Virus Genotype 1-Infected Patients.

    Directory of Open Access Journals (Sweden)

    Julia Dietz

    Full Text Available Different highly effective interferon-free treatment options for chronic hepatitis C virus (HCV infection are currently available. Pre-existence of resistance associated variants (RAVs to direct antiviral agents (DAAs reduces sustained virologic response (SVR rates by 3-53% in hepatitis C virus (HCV genotype 1 infected patients depending on different predictors and the DAA regimen used. Frequencies of single and combined resistance to NS3, NS5A and NS5B inhibitors and consequences for the applicability of different treatment regimens are unknown. Parallel population based sequencing of HCV NS3, NS5A and NS5B genes in 312 treatment-naïve Caucasian HCV genotype 1 infected patients showed the presence of major resistant variants in 20.5% (NS3, 11.9% (NS5A, and 22.1% (NS5B with important differences for HCV subtypes. In NS3, Q80K was observed in 34.7% and 2.1% of subtype 1a and 1b patients, respectively while other RAVs to second generation protease inhibitors were detected rarely (1.4%. Within NS5A RAVs were observed in 7.1% of subtype 1a and 17.6% in subtype 1b infected patients. RAVs to non-nucleoside NS5B inhibitors were observed in 3.5% and 44.4% of subtype 1a and 1b patients, respectively. Considering all three DAA targets all subtype 1a and 98.6% of subtype 1b infected patients were wildtype for at least one interferon free DAA regimen currently available. In conclusion, baseline resistance testing allows the selection of at least one RAVs-free treatment option for nearly all patients enabling a potentially cost- and efficacy-optimized treatment of chronic hepatitis C.

  8. Genotypic Differences in Growth and Physiological Responses to Transplanting and Direct Seeding Cultivation in Rice

    Directory of Open Access Journals (Sweden)

    Song Chen

    2009-06-01

    Full Text Available The field experiments were conducted to investigate the growth and physiological responses of six super hybrid rice combinations to two planting methods, transplanting (TP and direct seeding (DS during 2006–2007 and 2007–2008. The 1000-grain weight and number of tillers per plant at the early growth stage, the maximum quantum yield of PSII (Fv/Fm and transpiration rate (Tr were higher in DS plants than in TP ones, whereas the grain yield, number of panicles per square meter, seed setting rate, net photosynthetic rate (Pn and stomatal conductance were lower in DS plants. However, little difference was detected in number of grains per panicle, stem (shoot and leaf weight between the combinations in the two planting methods. The responses of plant growth and physiological traits to planting method differed greatly among the six combinations. In both planting methods, Chouyou 58 and Yongyou 6 had the highest and lowest panicle biomass and Pn, respectively. The higher yield of Chunyou 58 was associated with more numbers of panicles per square meter and grains per panicle in both planting methods. The results indicate that lower grain yield in DS relative to TP is attributed to more excessive tillers at the early stage, lower leaf biomass and photosynthetic rate at the late stage.

  9. RNA-seq analysis of mycobacteria stress response to microgravity

    Data.gov (United States)

    National Aeronautics and Space Administration — The aim of this work is to determine whether mycobacteria have enhanced virulence during space travel and what mechanisms they use to adapt to microgravity. M....

  10. Nisin depletes ATP and proton motive force in mycobacteria.

    Science.gov (United States)

    Chung, H J; Montville, T J; Chikindas, M L

    2000-12-01

    This study examined the inhibitory effect of nisin and its mode of action against Mycobacterium smegmatis, a non-pathogenic species of mycobacteria, and M. bovis-Bacill Carmette Guerin (BCG), a vaccine strain of pathogenic M. bovis. In agar diffusion assays, 2.5 mg ml(-1) nisin was required to inhibit M. bovis-BCG. Nisin caused a slow, gradual, time- and concentration-dependent decrease in internal ATP levels in M. bovis-BCG, but no ATP efflux was detected. In mycobacteria, nisin decreased both components of proton motive force (membrane potential, Delta Psi and Delta pH) in a time- and concentration-dependent manner. However, mycobacteria maintained their intracellular ATP levels during the initial time period of Delta Psi and Delta pH dissipation. These data suggest that the mechanism of nisin in mycobacteria is similar to that in food-borne pathogens.

  11. Characterization and Strains of Mycobacteria Isolated from Cattle ...

    African Journals Online (AJOL)

    Characterization and Strains of Mycobacteria Isolated from Cattle Carcasses in Refrigerated Slaughterhouse Bamako Goal. Y S Kone, A Cisse, S S Sidibe, Z Tarnagda, S Diarra, D Yassa, P Coulibaly, B Traore ...

  12. Performance Assessment of the CapitalBio Mycobacterium Identification Array System for Identification of Mycobacteria

    Science.gov (United States)

    Liu, Jingbo; Yan, Zihe; Han, Min; Han, Zhijun; Jin, Lingjie; Zhao, Yanlin

    2012-01-01

    The CapitalBio Mycobacterium identification microarray system is a rapid system for the detection of Mycobacterium tuberculosis. The performance of this system was assessed with 24 reference strains, 486 Mycobacterium tuberculosis clinical isolates, and 40 clinical samples and then compared to the “gold standard” of DNA sequencing. The CapitalBio Mycobacterium identification microarray system showed highly concordant identification results of 100% and 98.4% for Mycobacterium tuberculosis complex (MTC) and nontuberculous mycobacteria (NTM), respectively. The sensitivity and specificity of the CapitalBio Mycobacterium identification array for identification of Mycobacterium tuberculosis isolates were 99.6% and 100%, respectively, for direct detection and identification of clinical samples, and the overall sensitivity was 52.5%. It was 100% for sputum, 16.7% for pleural fluid, and 10% for bronchoalveolar lavage fluid, respectively. The total assay was completed in 6 h, including DNA extraction, PCR, and hybridization. The results of this study confirm the utility of this system for the rapid identification of mycobacteria and suggest that the CapitalBio Mycobacterium identification array is a molecular diagnostic technique with high sensitivity and specificity that has the capacity to quickly identify most mycobacteria. PMID:22090408

  13. Surgical site infections due to rapidly growing mycobacteria in puducherry, India.

    Science.gov (United States)

    Kannaiyan, Kavitha; Ragunathan, Latha; Sakthivel, Sulochana; Sasidar, A R; Muralidaran; Venkatachalam, G K

    2015-03-01

    Rapidly growing Mycobacteria are increasingly recognized, nowadays as an important pathogen that can cause wide range of clinical syndromes in humans. We herein describe unrelated cases of surgical site infection caused by Rapidly growing Mycobacteria (RGM), seen during a period of 12 months. Nineteen patients underwent operations by different surgical teams located in diverse sections of Tamil Nadu, Pondicherry, Karnataka, India. All patients presented with painful, draining subcutaneous nodules at the infection sites. Purulent material specimens were sent to the microbiology laboratory. Gram stain and Ziehl-Neelsen staining methods were used for direct examination. Culture media included blood agar, chocolate agar, MacConkey agar, Sabourauds agar and Lowenstein-Jensen medium for Mycobacteria. Isolated microorganisms were identified and further tested for antimicrobial susceptibility by standard microbiologic procedures. Mycobacterium fortuitum and M.chelonae were isolated from the purulent drainage obtained from wounds by routine microbiological techniques from all the specimens. All isolates analyzed for antimicrobial susceptibility pattern were sensitive to clarithromycin, linezolid and amikacin but were variable to ciprofloxacin, rifampicin and tobramycin. Our case series highlights that a high level of clinical suspicion should be maintained for patients presenting with protracted soft tissue lesions with a history of trauma or surgery as these infections not only cause physical but also emotional distress that affects both the patients and the surgeon.

  14. 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...

  15. Retreatment of patients with treatment failure of direct-acting antivirals: Focus on hepatitis C virus genotype 1b.

    Science.gov (United States)

    Kanda, Tatsuo; Nirei, Kazushige; Matsumoto, Naoki; Higuchi, Teruhisa; Nakamura, Hitomi; Yamagami, Hiroaki; Matsuoka, Shunichi; Moriyama, Mitsuhiko

    2017-12-14

    The recent development of direct-acting antiviral agents (DAAs) against hepatitis C virus (HCV) infection could lead to higher sustained virological response (SVR) rates, with shorter treatment durations and fewer adverse events compared with regimens that include interferon. However, a relatively small proportion of patients cannot achieve SVR in the first treatment, including DAAs with or without peginterferon and/or ribavirin. Although retreatment with a combination of DAAs should be conducted for these patients, it is more difficult to achieve SVR when retreating these patients because of resistance-associated substitutions (RASs) or treatment-emergent substitutions. In Japan, HCV genotype 1b (GT1b) is founded in 70% of HCV-infected individuals. In this minireview, we summarize the retreatment regimens and their SVR rates for HCV GT1b. It is important to avoid drugs that target the regions targeted by initial drugs, but next-generation combinations of DAAs, such as sofosbuvir/velpatasvir/voxilaprevir for 12 wk or glecaprevir/pibrentasvir for 12 wk, are proposed to be potential solution for the HCV GT1b-infected patients with treatment failure, mainly on a basis of targeting distinctive regions. Clinicians should follow the new information and resources for DAAs and select the proper combination of DAAs for the retreatment of HCV GT1b-infected patients with treatment failure.

  16. Direct costs of first-generation protease inhibitors for the treatment of genotype 1 chronic hepatitis C viral infection.

    Science.gov (United States)

    Sethi, N; Tapper, E B; Vong, A; Sethi, S; Rourke, M; Afdhal, N H

    2015-12-01

    To evaluate the cost-effectiveness of Hepatitis C therapy, robust real-world data are needed to understand the costs and benefits of treatment alternatives. The objective of this study was to evaluate the true direct cost of treatment in an unselected sequential population of patients treated at a tertiary care centre for hepatitis C virus genotype 1. A total of 200 consecutive patients were treated with interferon, ribavirin and a first-generation direct-acting antiviral agent (DAA) between 2011 and 2013. A total of 41% had cirrhosis, 31% were prior relapsers, and 41% were prior partial or null responders. Costs used were wholesale acquisition cost prices for medications, average hospital costs per day for each diagnosis code based on US inpatient hospital charges. All costs were adjusted to 2013 dollars. Sustained virologic response (SVR) was achieved in 97 patients (48.5%). A total of 14% experienced relapse, 19% breakthrough or nonresponse, and 18.5% discontinued secondary to side effects. Twenty per cent of patients had at least one hospitalization attributable to a complication of therapy. Thirty-seven per cent of patients required erythropoietin-stimulating agents, 16% received filgastrim, and 15% needed a red blood cell transfusion. The mean overall cost of treatment was $83,851 per patient. The cost per SVR was $172,889; $266,670 for patients with cirrhosis. The costs per SVR after treatment with first-generation DAAs are dependent on the stage of disease and therapy side effects. These real-world costs significantly exceed those described in prior cost-effectiveness assessments and should be used instead for future studies. © 2015 John Wiley & Sons Ltd.

  17. Evaluation of amplified rDNA restriction analysis (ARDRA for the identification of cultured mycobacteria in a diagnostic laboratory

    Directory of Open Access Journals (Sweden)

    Rottiers Sylvianne

    2002-03-01

    Full Text Available Abstract Background The development of DNA amplification for the direct detection of M. tuberculosis from clinical samples has been a major goal of clinical microbiology during the last ten years. However, the limited sensitivity of most DNA amplification techniques restricts their use to smear positive samples. On the other hand, the development of automated liquid culture has increased the speed and sensitivity of cultivation of mycobacteria. We have opted to combine automated culture with rapid genotypic identification (ARDRA: amplified rDNA restriction analysis for the detection resp. identification of all mycobacterial species at once, instead of attempting direct PCR based detection from clinical samples of M. tuberculosis only. Results During 1998–2000 a total of approx. 3500 clinical samples was screened for the presence of M. tuberculosis. Of the 151 culture positive samples, 61 were M. tuberculosis culture positive. Of the 30 smear positive samples, 26 were M. tuberculosis positive. All but three of these 151 mycobacterial isolates could be identified with ARDRA within on average 36 hours. The three isolates that could not be identified belonged to rare species not yet included in our ARDRA fingerprint library or were isolates with an aberrant pattern. Conclusions In our hands, automated culture in combination with ARDRA provides with accurate, practically applicable, wide range identification of mycobacterial species. The existing identification library covers most species, and can be easily updated when new species are studied or described. The drawback is that ARDRA is culture-dependent, since automated culture of M. tuberculosis takes on average 16.7 days (range 6 to 29 days. However, culture is needed after all to assess the antibiotic susceptibility of the strains.

  18. 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

  19. Epidemiology of nontuberculous mycobacteria isolations among central North Carolina residents, 2006-2010

    Science.gov (United States)

    BACKGROUND: Nontuberculous mycobacteria (NTM) are environmental mycobacteria associated with a range of infections. Reports of NTM epidemiology have primarily focused on pulmonary infections and isolations, however extrapulmonary infections of the skin, soft tissues and sterile s...

  20. Isolation of rapid growing mycobacteria from soil and water in Iran

    African Journals Online (AJOL)

    USER

    2010-06-14

    Jun 14, 2010 ... an abundant occurrence of mycobacteria in low pH (P value = 0001). We also ..... between large numbers of mycobacterial and high soil acidity as ... (2002). Chlorine disinfection of atypical mycobacteria isolated from a.

  1. IFN-γ fails to overcome inhibition of selected macrophage activation events in response to pathogenic mycobacteria.

    Directory of Open Access Journals (Sweden)

    Shyamala Thirunavukkarasu

    Full Text Available According to most models of mycobacterial infection, inhibition of the pro-inflammatory macrophage immune responses contributes to the persistence of bacteria. Mycobacterium avium subsp. paratuberculosis (MAP is a highly successful pathogen in cattle and sheep and is also implicated as the causative agent of Crohn's disease in humans. Pathogenic mycobacteria such as MAP have developed multiple strategies to evade host defence mechanisms including interfering with the macrophages' capacity to respond to IFN-γ, a feature which might be lacking in non-pathogenic mycobacteria such as M. smegmatis. We hypothesized that pre-sensitisation of macrophages with the pro-inflammatory cytokine IFN-γ would help in overcoming the inhibitory effect of MAP or its antigens on macrophage inflammatory responses. Herein we have compared a series of macrophage activation parameters in response to MAP and M. smegmatis as well as mycobacterial antigens. While IFN-γ did overcome the inhibition in immune suppressive mechanisms in response to MAP antigen as well as M. smegmatis, we could not find a clear role for IFN-γ in overcoming the inhibition of macrophage inflammatory responses to the pathogenic mycobacterium, MAP. We demonstrate that suppression of macrophage defence mechanisms by pathogenic mycobacteria is unlikely to be overcome by prior sensitization with IFN-γ alone. This indicates that IFN-γ signaling pathway-independent mechanisms may exist for overcoming inhibition of macrophage effector functions in response to pathogenic mycobacteria. These findings have important implications in understanding the survival mechanisms of pathogenic mycobacteria directed towards finding better therapeutics and vaccination strategies.

  2. Mycobacteria in Terrestrial Small Mammals on Cattle Farms in Tanzania

    DEFF Research Database (Denmark)

    Durnez, Lies; Katakweba, Abdul; Sadiki, Harrison

    2011-01-01

    The control of bovine tuberculosis and atypical mycobacterioses in cattle in developing countries is important but difficult because of the existence of wildlife reservoirs. In cattle farms in Tanzania, mycobacteria were detected in 7.3% of 645 small mammals and in cow's milk. The cattle farms we....... However, because of the high prevalence of mycobacteria in some small mammal species, these infected animals can pose a risk to humans, especially in areas with a high HIV-prevalence as is the case in Tanzania.......The control of bovine tuberculosis and atypical mycobacterioses in cattle in developing countries is important but difficult because of the existence of wildlife reservoirs. In cattle farms in Tanzania, mycobacteria were detected in 7.3% of 645 small mammals and in cow's milk. The cattle farms were...... and PCR in the small mammals were atypical mycobacteria. Analysis of the presence of mycobacteria in relation to the reactor status of the cattle farms does not exclude transmission between small mammals and cattle but indicates that transmission to cattle from another source of infection is more likely...

  3. Macrophage sphingolipids are essential for the entry of mycobacteria.

    Science.gov (United States)

    Viswanathan, Gopinath; Jafurulla, Md; Kumar, G Aditya; Raghunand, Tirumalai R; Chattopadhyay, Amitabha

    2018-07-01

    Mycobacteria are intracellular pathogens that can invade and survive within host macrophages. Mycobacterial infections remain a major cause of mortality and morbidity worldwide, with serious concerns of emergence of multi and extensively drug-resistant tuberculosis. While significant advances have been made in identifying mycobacterial virulence determinants, the detailed molecular mechanism of internalization of mycobacteria into host cells remains poorly understood. Although several studies have highlighted the crucial role of sphingolipids in mycobacterial growth, persistence and establishment of infection, the role of sphingolipids in the entry of mycobacteria into host cells is not known. In this work, we explored the role of host membrane sphingolipids in the entry of Mycobacterium smegmatis into J774A.1 macrophages. Our results show that metabolic depletion of sphingolipids in host macrophages results in a significant reduction in the entry of M. smegmatis. Importantly, the entry of Escherichia coli into host macrophages under similar conditions remained invariant, implying the specificity of the requirement of sphingolipids in mycobacterial entry. To the best of our knowledge, our results constitute the first report demonstrating the role of host macrophage sphingolipids in the entry of mycobacteria. Our results could help in the development of novel therapeutic strategies targeting sphingolipid-mediated entry of mycobacteria into host cells. Copyright © 2018 Elsevier B.V. All rights reserved.

  4. Nontuberculous mycobacteria in non-AIDS patients

    Directory of Open Access Journals (Sweden)

    A. Marinho

    2008-05-01

    Full Text Available Nontuberculous mycobacteria (NTM play an increasingly significant pathogenic role in HIV-positive patients, in patients with chronic lung disease, in other chronic conditions and in the elderly. Aims: Evaluate the importance of NTM isolation in respiratory samples in patients without HIV-infection. Methods: Retrospective evaluation of our hospital patients with no known AIDS, with at least one NTM positive respiratory sample, from 1997-2004. Results: We found 102 patients, with a median age of 63 years; 67% male. Sixty-three (62% had underlying lung disease, mainly tuberculosis sequelae (n = 19. The majority (47% of the isolations were Mycobacterium avium complex (MAC.A diagnosis of Mycobacterium pulmonary disease was made in 16 patients (15.7%, 14 of which met the American Thoracic Society diagnostic criteria. Ten male and 6 female; median age 65 years. Twelve had underlying lung disease. All of them had respiratory infection complaints. Chest X-rays showed mainly pulmonary infiltrates, linear opacities and cavitation. MAC was the cause of mycobacterium respiratory disease in 12 patients (75%. Conclusion: NTM isolation did not equal pulmonary NTM disease in the majority of cases, even in patients with underlying lung disease. MAC was the most commonly isolated agent and its relative importance was higher in the presence of NTM disease. Resumo: As micobactérias não tuberculosas (MNT têm um papel patogénico de importância crescente em doentes com serologia positiva para o vírus da imunodeficiência humana (VIH, em doentes com doença pulmonar crónica, em outras doencas crónicas, e ainda nos idosos. Objectivo: Avaliar a importância do isolamento de MNT em amostras respiratórias em doentes adultos sem infecção VIH. Material e métodos: Estudo retrospectivo dos doentes do Hospital de São João sem infecção VIH conhecida, com pelo menos uma amostra respiratória positiva para MNT, entre 1997 e 2004. Resultados: Foram encontrados 102

  5. Nontuberculous mycobacteria in Middle East: Current situation and future challenges

    Directory of Open Access Journals (Sweden)

    Ali Akbar Velayati

    2015-01-01

    Mycobacterium fortuitum was the most common rapid growing mycobacteria (RGM isolated from both clinical (269 out of 447 RGM; 60.1% and environmental (135 out of 289 RGM; 46.7% samples. Mycobacterium avium complex (MAC was the most common slow growing mycobacteria (SGM isolated from clinical samples (140 out of 637 SGM; 21.9%. An increasing trend in NTM isolation from the Middle East was noted over the last 5 years. This review demonstrates the increasing concern regarding NTM disease in the Middle East, emphasizing the need for regional collaboration and coordination in order to respond appropriately.

  6. First detection of mycobacteria in African rodents and insectivores, using stratified pool screening

    DEFF Research Database (Denmark)

    Durnez, L.; Eddyani, M.; Mgode, G. F.

    2007-01-01

    With the rising number of patients with human immunodeficiency virus (HIV)/AIDS in developing countries, the control of mycobacteria is of growing importance. Previous studies have shown that rodents and insectivores are carriers of mycobacteria. However, it is not clear how widespread mycobacteria...... spp.) were isolated from C. gambianus, Mastomys natalensis, and C. hirta. This study is the first to report findings of mycobacteria in African rodents and insectivores and the first in mycobacterial ecology to estimate the prevalence of mycobacteria after stratified pool screening. The fact...... that small mammals in urban areas carry more mycobacteria than those in the fields and that potentially pathogenic mycobacteria were isolated identifies a risk for other animals and humans, especially HIV/AIDS patients, that have a weakened immune system. ...

  7. Ionizing radiation in the disinfection of water contaminated with potentially pathogenic mycobacteria

    International Nuclear Information System (INIS)

    Kubin, M.; Sedlackova, J.; Vacek, K.

    1982-01-01

    Sterile drinking water samples were artificially colonized with M. kansasii, M. gardonae and M. fortuitum suspensions (the numbers of viable units in 1 ml were 1.2x10 3 , 48.5 and 3.2x10 3 , respectively) prepared from mycobacterial strains replicated in Tween 80-free liquid Dubos medium STO. The contaminated water samples were irradiated from a rotary cobalt 60 source (gamma radiation, E=1.17 and 1.33 MeV, dose rate 1 kJ/kg.h at room temperature) with doses 0.7, 1.5, 2.2, 3, 9, 16 and 27 kJ/kg. The disinfecting effectiveness was assessed by direct cultivation tests (0.5 ml volumes of water inoculated on egg medium) and by cultivation on membrane filtres after filtering the whole amount of the water examined (about 500 ml). Total disinfection was recorded for M. kansasii and M. fortuitum irradiated with 9 kJ/kg and for M. gordonae after irradiation with 1.5 kJ/kg. The calculated value of D 10 =0.4 kJ/kg (i.e., the radiation dose that reduces the number of viable mycobacteria by an order of magnitude) is suggestive of a strong disinfecting effect of ionizing radiation on the tested strains of potentially pathogenic mycobacteria. The results indicate that ionizing radiation could be applxcable in disinfecting supply and potable water contaminated with mycobacteria difficult to remove by other methods which, as a rule, cannot ensure permanent disinfection. (author)

  8. Ionizing radiation in the disinfection of water contaminated with potentially pathogenic mycobacteria

    Energy Technology Data Exchange (ETDEWEB)

    Kubin, M [Institut Hygieny a Epidemiologie, Prague (Czechoslovakia); Sedlackova, J; Vacek, K [Ustav Jaderneho Vyzkumu CSKAE, Rez (Czechoslovakia)

    1982-01-01

    Sterile drinking water samples were artificially colonized with M. kansasii, M. gardonae and M. fortuitum suspensions (the numbers of viable units in 1 ml were 1.2x10/sup 3/, 48.5 and 3.2x10/sup 3/, respectively) prepared from mycobacterial strains replicated in Tween 80-free liquid Dubos medium STO. The contaminated water samples were irradiated from a rotary cobalt 60 source (gamma radiation, E=1.17 and 1.33 MeV, dose rate 1 kJ/kg.h at room temperature) with doses 0.7, 1.5, 2.2, 3, 9, 16 and 27 kJ/kg. The disinfecting effectiveness was assessed by direct cultivation tests (0.5 ml volumes of water inoculated on egg medium) and by cultivation on membrane filtres after filtering the whole amount of the water examined (about 500 ml). Total disinfection was recorded for M. kansasii and M. fortuitum irradiated with 9 kJ/kg and for M. gordonae after irradiation with 1.5 kJ/kg. The calculated value of D/sub 10/=0.4 kJ/kg (i.e., the radiation dose that reduces the number of viable mycobacteria by an order of magnitude) is suggestive of a strong disinfecting effect of ionizing radiation on the tested strains of potentially pathogenic mycobacteria. The results indicate that ionizing radiation could be applicable in disinfecting supply and potable water contaminated with mycobacteria difficult to remove by other methods which, as a rule, cannot ensure permanent disinfection.

  9. Subcellular trafficking of mycobacteria : Implications for virulence and immunogenicity

    NARCIS (Netherlands)

    Houben, D.

    2011-01-01

    The aim of this thesis is to determine the properties of the compartment where mycobacteria end up after phagocytosis and which mycobacterial genes play a role in this process. In most cases, bacterial pathogens are taken up by the cell, processed in the endocytic pathway and eventually bacterial

  10. Resistance mechanisms and drug susceptibility testing of nontuberculous mycobacteria.

    NARCIS (Netherlands)

    Ingen, J. van; Boeree, M.J.; Soolingen, D. van; Mouton, J.W.

    2012-01-01

    Nontuberculous mycobacteria (NTM) are increasingly recognized as causative agents of opportunistic infections in humans. For most NTM infections the therapy of choice is drug treatment, but treatment regimens differ by species, in particular between slow (e.g. Mycobacterium avium complex,

  11. Thiacetazone, an antitubercular drug that inhibits cyclopropanation of cell wall mycolic acids in mycobacteria.

    Directory of Open Access Journals (Sweden)

    Anuradha Alahari

    2007-12-01

    Full Text Available Mycolic acids are a complex mixture of branched, long-chain fatty acids, representing key components of the highly hydrophobic mycobacterial cell wall. Pathogenic mycobacteria carry mycolic acid sub-types that contain cyclopropane rings. Double bonds at specific sites on mycolic acid precursors are modified by the action of cyclopropane mycolic acid synthases (CMASs. The latter belong to a family of S-adenosyl-methionine-dependent methyl transferases, of which several have been well studied in Mycobacterium tuberculosis, namely, MmaA1 through A4, PcaA and CmaA2. Cyclopropanated mycolic acids are key factors participating in cell envelope permeability, host immunomodulation and persistence of M. tuberculosis. While several antitubercular agents inhibit mycolic acid synthesis, to date, the CMASs have not been shown to be drug targets.We have employed various complementary approaches to show that the antitubercular drug, thiacetazone (TAC, and its chemical analogues, inhibit mycolic acid cyclopropanation. Dramatic changes in the content and ratio of mycolic acids in the vaccine strain Mycobacterium bovis BCG, as well as in the related pathogenic species Mycobacterium marinum were observed after treatment with the drugs. Combination of thin layer chromatography, mass spectrometry and Nuclear Magnetic Resonance (NMR analyses of mycolic acids purified from drug-treated mycobacteria showed a significant loss of cyclopropanation in both the alpha- and oxygenated mycolate sub-types. Additionally, High-Resolution Magic Angle Spinning (HR-MAS NMR analyses on whole cells was used to detect cell wall-associated mycolates and to quantify the cyclopropanation status of the cell envelope. Further, overexpression of cmaA2, mmaA2 or pcaA in mycobacteria partially reversed the effects of TAC and its analogue on mycolic acid cyclopropanation, suggesting that the drugs act directly on CMASs.This is a first report on the mechanism of action of TAC, demonstrating the

  12. ISOLATION AND ANTIBIOTIC SUSCEPTIBILITY TESTING OF RAPIDLY-GROWING MYCOBACTERIA FROM GRASSLAND SOILS

    Directory of Open Access Journals (Sweden)

    Martina Kyselková

    2013-08-01

    Full Text Available Rapidly growing mycobacteria (RGM are common soil saprophytes, but certain strains cause infections in human and animals. The infections due to RGM have been increasing in past decades and are often difficult to treat. The susceptibility to antibiotics is regularly evaluated in clinical isolates of RGM, but the data on soil RGM are missing. The objectives of this study was to isolate RGM from four grassland soils with different impact of manuring, and assess their resistance to antibiotics and the ability to grow at 37°C and 42°C. Since isolation of RGM from soil is a challenge, a conventional decontamination method (NaOH/malachite green/cycloheximide and a recent method based on olive oil/SDS demulsification were compared. The olive oil/SDS method was less efficient, mainly because of the emulsion instability and plate overgrowing with other bacteria. Altogether, 44 isolates were obtained and 23 representatives of different RGM genotypes were screened. The number of isolates per soil decreased with increasing soil pH, consistently with previous findings that mycobacteria were more abundant in low pH soils. Most of the isolates belonged to the Mycobacterium fortuitum group. The majority of isolates was resistant to 2-4 antibiotics. Multiresistant strains occurred also in a control soil that has a long history without the exposure to antibiotic-containing manure. Seven isolates grew at 37°C, including the species M. septicum and M. fortuitum known for infections in humans. This study shows that multiresistant RGM close to known human pathogens occur in grassland soils regardless the soil history of manuring.

  13. Verification of Frequency in Species of Nontuberculous Mycobacteria in Kermanshah Drinking Water Supplies Using the PCR-Sequencing Method.

    Science.gov (United States)

    Mohajeri, Parviz; Yazdani, Laya; Shahraki, Abdolrazagh Hashemi; Alvandi, Amirhoshang; Atashi, Sara; Farahani, Abbas; Almasi, Ali; Rezaei, Mansour

    2017-04-01

    Nontuberculous mycobacteria are habitants of environment, especially in aquatic systems. Some of them cause problems in immunodeficient patients. Over the last decade, 16S rRNA gene sequencing was established in 45 novel species of nontuberculous mycobacteria. Experiences revealed that this method underestimates the diversity, but does not distinguish between some of mycobacterium subsp. To recognize emerging rapidly growing mycobacteria and identify their subsp, rpoB gene sequencing has been developed. To better understand the transmission of nontuberculous mycobacterial species from drinking water and preventing the spread of illness with these bacteria, the aim of this study was to detect the presence of bacteria by PCR-sequencing techniques. Drinking water samples were collected from different areas of Kermanshah city in west of IRAN. After decontamination with cetylpyridinium chloride, samples were filtered with 0.45-micron filters, the filter transferred directly on growth medium waiting to appear in colonies, then DNA extraction and PCR were performed, and products were sent to sequencing. We found 35/110 (32%) nontuberculous mycobacterial species in drinking water samples, isolates included Mycobacterium goodii, Mycobacterium aurum, and Mycobacterium gastri with the most abundance (11.5%), followed by Mycobacterium smegmatis, Mycobacterium porcinum, Mycobacterium peregrinum, Mycobacterium mucogenicum, and Mycobacterium chelonae (8%). In this study, we recognized the evidence of contamination by nontuberculous mycobacteria in corroded water pipes. As a result of the high prevalence of these bacteria in drinking water in Kermanshah, this is important evidence of transmission through drinking water. This finding can also help public health policy makers control these isolates in drinking water supplies in Kermanshah.

  14. Urease testing of mycobacteria with BACTEC radiometric instrumentation

    International Nuclear Information System (INIS)

    Damato, J.J.; Collins, M.T.; McClatchy, J.K.

    1982-01-01

    A total of 140 mycobacterial isolates from patients treated at Fitzsimons Army Medical Center or the National Jewish Hospital and Research Center and from animal specimens submitted to the National Veterinary Services Laboratory were tested by using a urease procedure modified for use with a BACTEC model 301. Mycobacterial suspensions were prepared by using Middlebrook 7H10 Tween broth. Of the 98 mycobacteria isolates which were urease positive utilizing standard methodology, all were positive using the radiometric procedures. Similarly, all 42 urease-negative isolates were also negative employing the new methodology. Although maximum radiometric readings were observed at 48 h, all positive strains were readily identified 24 h after inoculation without sacrificing either test sensitivity or specificity. Thus, urease testing of mycobacteria, using the modified BACTEC radiometric methodology, was as sensitive, as specific, and more rapid than conventional methods

  15. Malachite green interferes with postantibiotic recovery of mycobacteria.

    Science.gov (United States)

    Gelman, Ekaterina; McKinney, John D; Dhar, Neeraj

    2012-07-01

    The genus Mycobacterium comprises slow-growing species with generation times ranging from hours to weeks. The protracted incubation time before colonies appear on solid culture medium can result in overgrowth by faster-growing microorganisms. To prevent contamination, the solid media used in laboratories and clinics for cultivation of mycobacteria contain the arylmethane compound malachite green, which has broad-spectrum antimicrobial activity. Malachite green has no impact on the plating efficiency of mycobacteria when cells are grown under normal conditions. However, we found that malachite green interfered with colony formation when bacteria were preexposed to antibiotics targeting cell wall biogenesis (isoniazid, ethionamide, ethambutol). This inhibitory effect of malachite green was not observed when bacteria were preexposed to antibiotics targeting cellular processes other than cell wall biogenesis (rifampin, moxifloxacin, streptomycin). Sputum specimens from tuberculosis patients are routinely evaluated on solid culture medium containing high concentrations of malachite green. This practice could lead to underestimation of bacterial loads and overestimation of chemotherapeutic efficacy.

  16. Surveillance of Tuberculosis in Taipei: The Influence of Nontuberculous Mycobacteria

    OpenAIRE

    Chiang, Chen-Yuan; Yu, Ming-Chih; Yang, Shiang-Lin; Yen, Muh-Yong; Bai, Kuan-Jen

    2015-01-01

    Background Notification of tuberculosis (TB) but not nontuberculous mycobacteria (NTM) is mandatory in Taiwan. Partly due to the strict regulation on TB notification, several patients infected with NTM were notified as TB cases. Notification of patients infected with NTM as TB cases can trigger public health actions and impose additional burdens on the public health system. We conducted a study to assess the influence of NTM infection on surveillance of TB in Taipei. Methodology/Principal Fin...

  17. Nontuberculous mycobacteria in Middle East: Current situation and future challenges

    OpenAIRE

    Velayati, Ali Akbar; Rahideh, Sanaz; Derakhshani Nezhad, Zahra; Farnia, Parissa; Mirsaeidi, Mehdi

    2015-01-01

    Nontuberculous mycobacteria (NTM) are a diverse group of bacterial species that are distributed in the environment. Many of these environmental bacteria can cause disease in humans. The identification of NTM in environmental sources is important for both clinical and epidemiological purposes. In this study, the distribution of NTM species from environmental and clinical samples in the Middle East was reviewed. In order to provide an overview of NTM, as well as recent epidemiological trends, a...

  18. Detection and recovery of mycobacteria by a radiometric procedure

    International Nuclear Information System (INIS)

    Takahashi, H.; Foster, V.

    1983-01-01

    During a 6-month period, 5,375 clinical specimens were cultured on Middlebrook-Cohn 7H10 medium, on Lowenstein-Jensen medium, and in Middlebrook 7H12 medium containing [ 14 C]palmitic acid. More mycobacteria were recovered when all three media were used than when either the conventional method with 7H10 agar and Lowenstein-Jensen slants or the radiometric method with 7H12 broth was used alone

  19. Methodology of mycobacteria tuberculosis bacteria detection by Raman spectroscopy

    Science.gov (United States)

    Zyubin, A.; Lavrova, A.; Manicheva, O.; Dogonadze, M.; Tsibulnikova, A.; Samusev, I.

    2018-01-01

    We have developed a methodology for the study of deactivated strains of Mycobacterium tuberculosis. Strains of the Beijing species obtained from pulmonary patient secrete (XDR strain) and reference strain (H37Rv) were investigated by Raman spectrometry with He-Ne (632,8 nm) laser excitation source. As a result of the research, the optimal experimental parameters have been obtained to get spectra of mycolic acids, which are part of the cell wall of mycobacteria.

  20. Nontuberculous mycobacteria in Middle East: Current situation and future challenges.

    Science.gov (United States)

    Velayati, Ali Akbar; Rahideh, Sanaz; Nezhad, Zahra Derakhshani; Farnia, Parissa; Mirsaeidi, Mehdi

    2015-03-01

    Nontuberculous mycobacteria (NTM) are a diverse group of bacterial species that are distributed in the environment. Many of these environmental bacteria can cause disease in humans. The identification of NTM in environmental sources is important for both clinical and epidemiological purposes. In this study, the distribution of NTM species from environmental and clinical samples in the Middle East was reviewed. In order to provide an overview of NTM, as well as recent epidemiological trends, all studies addressing NTM in the Middle East from 1984 to 2014 were reviewed. A total of 96 articles were found, in which 1751 NTM strains were isolated and 1084 of which were obtained from clinical samples, 619 from environmental samples and 48 were cited by case reports. Mycobacterium fortuitum was the most common rapid growing mycobacteria (RGM) isolated from both clinical (269 out of 447 RGM; 60.1%) and environmental (135 out of 289 RGM; 46.7%) samples. Mycobacterium avium complex (MAC) was the most common slow growing mycobacteria (SGM) isolated from clinical samples (140 out of 637 SGM; 21.9%). An increasing trend in NTM isolation from the Middle East was noted over the last 5years. This review demonstrates the increasing concern regarding NTM disease in the Middle East, emphasizing the need for regional collaboration and coordination in order to respond appropriately. Copyright © 2015 Asian African Society for Mycobacteriology. Published by Elsevier Ltd. All rights reserved.

  1. ANALYSIS OF MUTATIONS OF TUBERCULOUS MYCOBACTERIA DEFINING DRUG RESISTANCE IN HIV POSITIVE AND HIV NEGATIVE TUBERCULOSIS PATIENTS WITHOUT PRIOR HISTORY OF TREATMENT IN SVERDLOVSK REGION

    Directory of Open Access Journals (Sweden)

    G. V. Panov

    2017-01-01

    Full Text Available Goal of the study: to identify profile of mutations of tuberculous mycobacteria responsible for resistance to anti-tuberculosis drugs in HIV positive and HIV negative tuberculosis patients without prior history of treatment.Materials and methods. 165 strains of tuberculous mycobacteria from HIV positive patients and 166 strains of tuberculous mycobacteria from HIV negative patients were studied in Sverdlovsk Region (TB Dispensary, Yekaterinburg. Mutations in genes were identified using microchips of TB-BIOCHIP® and TB-BIOCHIP®-2 in compliance with the manufacturer's guidelines (OOO Biochip-IMB, Moscow.Results. It was observed that 85/165 (51.52% strains isolated from HIV positive tuberculosis patients and 58/166 (34.94% strains isolated from tuberculosis patients not associated with HIV possessed MDR genotype (p < 0.01. The majority of MDR strains had mutations in the 531th codon of rpoB (Ser→Leu and 315th codon of katG (Ser→Thr (64/85, 75.29% and 38/58, 65.52% respective the groups, resulting in the high level of resistance to rifampicin and isoniazid. Each group also had approximately equal ratio (11/165, 6.67% and 12/166, 7.23% respective the groups of strains with genomic mutations defining the resistance to isoniazid, rifampicin and fluoruquinolones. No confident difference was found in mutation patterns of genome of tuberculous mycobacteria isolated from HIV positive and HIV negative tuberculosis patients. 

  2. Hepatitis C virus eradication by direct antiviral agents improves glucose tolerance and reduces post-load insulin resistance in nondiabetic patients with genotype 1.

    Science.gov (United States)

    Salomone, Federico; Catania, Maurizio; Montineri, Arturo; Bertino, Gaetano; Godos, Justyna; Rizzo, Leonardo; Magrì, Giovanni; Li Volti, Giovanni

    2017-12-19

    Genotype 1 chronic hepatitis C is associated with an impairment of glucose homoeostasis, especially in the advanced stages of the disease. Glucose tolerance is an independent predictor of liver-related mortality in patients with cirrhosis because of chronic hepatitis C. However, no study has demonstrated so far weather hepatitis C virus clearance affects glucose tolerance. To this aim, we performed a prospective study assessing the effects of direct antiviral agents treatment in nondiabetic cirrhotic patients with genotypes 1a/1b and impaired glucose tolerance based on a 75-g oral glucose tolerance test. Impaired glucose tolerance was diagnosed by a 2-hour plasma glucose between 140 and 199 mg/dL. Insulin resistance was estimated by the oral glucose insulin sensitivity index, an oral glucose tolerance test-derived measure. After meeting the inclusion criteria, the study population included 32 outpatients (26/6 genotypes 1b/1a; age 62 ± 7.4 years; 18 males) with compensated Child-A cirrhosis. All patients achieved a sustained virological response following direct antiviral agents treatment. After viral eradication, we did not observe change in fasting plasma glucose (103.5 ± 7.1 vs 102.8 ± 7.2 mg/dL, P = .15) but 2-hour plasma glucose was reduced (165.2 ± 22.7 vs 138.5 ± 21.3 mg/dL, P Hepatitis C virus eradication led also to a significant reduction in HbA1c (6.1 ± 0.2% vs 5.7 ± 0.3%, P resistance as assessed by the oral glucose insulin sensitivity index (6.92 ± 1.56 vs 9.52 ± 1.39 mg/kg/min, P  .5). Our results indicate that hepatitis C virus eradication may early improve glucose tolerance in patients with hepatitis C virus-related cirrhosis. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  3. Essential Role of the ESX-5 Secretion System in Outer Membrane Permeability of Pathogenic Mycobacteria

    KAUST Repository

    Ates, Louis S.

    2015-05-04

    Mycobacteria possess different type VII secretion (T7S) systems to secrete proteins across their unusual cell envelope. One of these systems, ESX-5, is only present in slow-growing mycobacteria and responsible for the secretion of multiple substrates. However, the role of ESX-5 substrates in growth and/or virulence is largely unknown. In this study, we show that esx-5 is essential for growth of both Mycobacterium marinum and Mycobacterium bovis. Remarkably, this essentiality can be rescued by increasing the permeability of the outer membrane, either by altering its lipid composition or by the introduction of the heterologous porin MspA. Mutagenesis of the first nucleotide-binding domain of the membrane ATPase EccC5 prevented both ESX-5-dependent secretion and bacterial growth, but did not affect ESX-5 complex assembly. This suggests that the rescuing effect is not due to pores formed by the ESX-5 membrane complex, but caused by ESX-5 activity. Subsequent proteomic analysis to identify crucial ESX-5 substrates confirmed that all detectable PE and PPE proteins in the cell surface and cell envelope fractions were routed through ESX-5. Additionally, saturated transposon-directed insertion-site sequencing (TraDIS) was applied to both wild-type M. marinum cells and cells expressing mspA to identify genes that are not essential anymore in the presence of MspA. This analysis confirmed the importance of esx-5, but we could not identify essential ESX-5 substrates, indicating that multiple of these substrates are together responsible for the essentiality. Finally, examination of phenotypes on defined carbon sources revealed that an esx-5 mutant is strongly impaired in the uptake and utilization of hydrophobic carbon sources. Based on these data, we propose a model in which the ESX-5 system is responsible for the transport of cell envelope proteins that are required for nutrient uptake. These proteins might in this way compensate for the lack of MspA-like porins in slow

  4. Essential Role of the ESX-5 Secretion System in Outer Membrane Permeability of Pathogenic Mycobacteria.

    Directory of Open Access Journals (Sweden)

    Louis S Ates

    2015-05-01

    Full Text Available Mycobacteria possess different type VII secretion (T7S systems to secrete proteins across their unusual cell envelope. One of these systems, ESX-5, is only present in slow-growing mycobacteria and responsible for the secretion of multiple substrates. However, the role of ESX-5 substrates in growth and/or virulence is largely unknown. In this study, we show that esx-5 is essential for growth of both Mycobacterium marinum and Mycobacterium bovis. Remarkably, this essentiality can be rescued by increasing the permeability of the outer membrane, either by altering its lipid composition or by the introduction of the heterologous porin MspA. Mutagenesis of the first nucleotide-binding domain of the membrane ATPase EccC5 prevented both ESX-5-dependent secretion and bacterial growth, but did not affect ESX-5 complex assembly. This suggests that the rescuing effect is not due to pores formed by the ESX-5 membrane complex, but caused by ESX-5 activity. Subsequent proteomic analysis to identify crucial ESX-5 substrates confirmed that all detectable PE and PPE proteins in the cell surface and cell envelope fractions were routed through ESX-5. Additionally, saturated transposon-directed insertion-site sequencing (TraDIS was applied to both wild-type M. marinum cells and cells expressing mspA to identify genes that are not essential anymore in the presence of MspA. This analysis confirmed the importance of esx-5, but we could not identify essential ESX-5 substrates, indicating that multiple of these substrates are together responsible for the essentiality. Finally, examination of phenotypes on defined carbon sources revealed that an esx-5 mutant is strongly impaired in the uptake and utilization of hydrophobic carbon sources. Based on these data, we propose a model in which the ESX-5 system is responsible for the transport of cell envelope proteins that are required for nutrient uptake. These proteins might in this way compensate for the lack of Msp

  5. Essential Role of the ESX-5 Secretion System in Outer Membrane Permeability of Pathogenic Mycobacteria

    KAUST Repository

    Ates, Louis S.; Ummels, Roy; Commandeur, Susanna; van der Weerd, Robert; Sparrius, Marion; Weerdenburg, Eveline; Alber, Marina; Kalscheuer, Rainer; Piersma, Sander R.; Abdallah, Abdallah; Abd El Ghany, Moataz; Abdel-Haleem, Alyaa M.; Pain, Arnab; Jimé nez, Connie R.; Bitter, Wilbert; Houben, Edith N.G.

    2015-01-01

    Mycobacteria possess different type VII secretion (T7S) systems to secrete proteins across their unusual cell envelope. One of these systems, ESX-5, is only present in slow-growing mycobacteria and responsible for the secretion of multiple substrates. However, the role of ESX-5 substrates in growth and/or virulence is largely unknown. In this study, we show that esx-5 is essential for growth of both Mycobacterium marinum and Mycobacterium bovis. Remarkably, this essentiality can be rescued by increasing the permeability of the outer membrane, either by altering its lipid composition or by the introduction of the heterologous porin MspA. Mutagenesis of the first nucleotide-binding domain of the membrane ATPase EccC5 prevented both ESX-5-dependent secretion and bacterial growth, but did not affect ESX-5 complex assembly. This suggests that the rescuing effect is not due to pores formed by the ESX-5 membrane complex, but caused by ESX-5 activity. Subsequent proteomic analysis to identify crucial ESX-5 substrates confirmed that all detectable PE and PPE proteins in the cell surface and cell envelope fractions were routed through ESX-5. Additionally, saturated transposon-directed insertion-site sequencing (TraDIS) was applied to both wild-type M. marinum cells and cells expressing mspA to identify genes that are not essential anymore in the presence of MspA. This analysis confirmed the importance of esx-5, but we could not identify essential ESX-5 substrates, indicating that multiple of these substrates are together responsible for the essentiality. Finally, examination of phenotypes on defined carbon sources revealed that an esx-5 mutant is strongly impaired in the uptake and utilization of hydrophobic carbon sources. Based on these data, we propose a model in which the ESX-5 system is responsible for the transport of cell envelope proteins that are required for nutrient uptake. These proteins might in this way compensate for the lack of MspA-like porins in slow

  6. Characterization of mycobacteria in HIV/AIDS patients of Nepal.

    Science.gov (United States)

    Dhungana, G P; Ghimire, P; Sharma, S; Rijal, B P

    2008-01-01

    Besides Mycobacterium tuberculosis, a number of other Mycobacterium species are also occasional human pathogens. Tuberculosis due to Mycobacterium avium complex (MAC) and Mycobacterium kansasii is particularly prevalent in AIDS patients as compared to the normal population. A cross-sectional study was carried out during January 2004 to August 2005 in 100 HIV-infected persons visiting Tribhuvan University, Teaching Hospital, and about a dozen of HIV/AIDS care centers of Kathmandu with the objectives to characterize the different mycobacterial species in HIV/AIDS patients. Three sputum specimens from each person were used to investigate tuberculosis by Ziehl-Neelsen staining, culture and identification tests. Among the 100 HIV-infected cases, 66 (66%) were males and 34 (34%) were females. Sixty percent of the cases were in the age group of 21-30 years. Mycobacteria were detected in 23 (23%) HIV cases of which 15 (65.2%) were in the age group of 21-30 years ; 17(74%) were males and 6 (26 %) were females. Among 23 co-infected cases, 22 were culture positive for mycobacteria. Among these, the predominant one was Mycobacterium avium complex (MAC), 9 (41%), followed by M. tuberculosis, 6 (27%), M .kansasii, 4 (18%), M. fortuitum, 2 (10%) and M. chelonae 1 (4%). Significant relationship was established between smoking/alcoholism and the subsequent development of tuberculosis (chi(2)=7.24, p<0.05 for smoking habit and chi(2)=4.39, p<0.05 for alcoholism). Fourteen (61%) co-infected cases presented with weight loss and cough whereas diarrhea was presented only by those patients with atypical mycobacterial co-infection, which was as high as 5 (56%) in patients with MAC co-infection. This study demonstrated the predominance of atypical mycobacteria, mainly MAC, in HIV/AIDS cases and most of them were from sputum smear-negative cases.

  7. [Breeding and management of mycobacteria-free guinea pigs (author's transl)].

    Science.gov (United States)

    Kazda, J

    1976-08-01

    A number of mycobacterial species are detectable under conventional holding condition of guinea pigs. These mycobacteria originating in drinking water and litter caused cross reactions in the Jones-Mote hypersensitivity test. Using suitable precautions it was possible to breed and hold the animals mycobacteria-free. The precautions depend mainly in alteration of the wire mesh floor in cages to avoide the contact of the animals with the litter, in cleaning and desinfection of water bottles, in using of heated water and food and in the prevention of mycobacterial contamination from the staff. The control examination on mycobacteria without treating is given in details. Cases are refered in which a oral rece ption of mycobacteria can alter the immune response. The modification of guinea pigs management to the mycobacteria-free ones is possible in a short time and with minimal cost.

  8. Subcellular trafficking of mycobacteria: Implications for virulence and immunogenicity

    OpenAIRE

    Houben, D.

    2011-01-01

    The aim of this thesis is to determine the properties of the compartment where mycobacteria end up after phagocytosis and which mycobacterial genes play a role in this process. In most cases, bacterial pathogens are taken up by the cell, processed in the endocytic pathway and eventually bacterial derived peptides are presented on MHC class II molecules to CD4+ T-cells. Proteins from viral pathogens in contrast, are degraded in the cytosol and transported into the ER for presentation on MHC cl...

  9. Destruction of Various Kinds of Mycobacteria in Milk by Pasteurization

    Science.gov (United States)

    Harrington, Rube; Karlson, Alfred G.

    1965-01-01

    Various strains of unclassified mycobacteria, Mycobacterium tuberculosis (including H37Rv strains), M. bovis, M. avium, M. fortuitum, and bacille Calmette-Guerin, were exposed to the temperature and time of pasteurization in skim milk in test tubes. Of the 195 strains tested, there were a few surviving colonies among 6 of 33 skotochromogens, 1 of 26 photochromogens, 10 of 79 nonchromogens, and 1 of 9 rapid growers. Subcultures of the surviving colonies failed to resist the pasteurization tests on subsequent trials. PMID:14325295

  10. Mycobacteria in water and loose deposits of drinking water distribution systems in Finland.

    Science.gov (United States)

    Torvinen, Eila; Suomalainen, Sini; Lehtola, Markku J; Miettinen, Ilkka T; Zacheus, Outi; Paulin, Lars; Katila, Marja-Leena; Martikainen, Pertti J

    2004-04-01

    Drinking water distribution systems were analyzed for viable counts of mycobacteria by sampling water from waterworks and in different parts of the systems. In addition, loose deposits collected during mechanical cleaning of the main pipelines were similarly analyzed. The study covered 16 systems at eight localities in Finland. In an experimental study, mycobacterial colonization of biofilms on polyvinyl chloride tubes in a system was studied. The isolation frequency of mycobacteria increased from 35% at the waterworks to 80% in the system, and the number of mycobacteria in the positive samples increased from 15 to 140 CFU/liter, respectively. Mycobacteria were isolated from all 11 deposits with an accumulation time of tens of years and from all 4 deposits which had accumulated during a 1-year follow-up time. The numbers of mycobacteria were high in both old and young deposits (medians, 1.8 x 10(5) and 3.9 x 10(5) CFU/g [dry weight], respectively). Both water and deposit samples yielded the highest numbers of mycobacteria in the systems using surface water and applying ozonation as an intermediate treatment or posttreatment. The number and growth of mycobacteria in system waters correlated strongly with the concentration of assimilable organic carbon in the water leaving the waterworks. The densities of mycobacteria in the developing biofilms were highest at the distal sites of the systems. Over 90% of the mycobacteria isolated from water and deposits belonged to Mycobacterium lentiflavum, M. tusciae, M. gordonae, and a previously unclassified group of mycobacteria. Our results indicate that drinking water systems may be a source for recently discovered new mycobacterial species.

  11. Double-Strand DNA Break Repair in Mycobacteria.

    Science.gov (United States)

    Glickman, Michael S

    2014-10-01

    Discontinuity of both strands of the chromosome is a lethal event in all living organisms because it compromises chromosome replication. As such, a diversity of DNA repair systems has evolved to repair double-strand DNA breaks (DSBs). In part, this diversity of DSB repair systems has evolved to repair breaks that arise in diverse physiologic circumstances or sequence contexts, including cellular states of nonreplication or breaks that arise between repeats. Mycobacteria elaborate a set of three genetically distinct DNA repair pathways: homologous recombination, nonhomologous end joining, and single-strand annealing. As such, mycobacterial DSB repair diverges substantially from the standard model of prokaryotic DSB repair and represents an attractive new model system. In addition, the presence in mycobacteria of a DSB repair system that can repair DSBs in nonreplicating cells (nonhomologous end joining) or when DSBs arise between repeats (single-strand annealing) has clear potential relevance to Mycobacterium tuberculosis pathogenesis, although the exact role of these systems in M. tuberculosis pathogenesis is still being elucidated. In this article we will review the genetics of mycobacterial DSB repair systems, focusing on recent insights.

  12. Specific Proteins in Nontuberculous Mycobacteria: New Potential Tools

    Directory of Open Access Journals (Sweden)

    Patricia Orduña

    2015-01-01

    Full Text Available Nontuberculous mycobacteria (NTM have been isolated from water, soil, air, food, protozoa, plants, animals, and humans. Although most NTM are saprophytes, approximately one-third of NTM have been associated with human diseases. In this study, we did a comparative proteomic analysis among five NTM strains isolated from several sources. There were different numbers of protein spots from M. gordonae (1,264, M. nonchromogenicum type I (894, M. nonchromogenicum type II (935, M. peregrinum (806, and M. scrofulaceum/Mycobacterium mantenii (1,486 strains, respectively. We identified 141 proteins common to all strains and specific proteins to each NTM strain. A total of 23 proteins were selected for its identification. Two of the common proteins identified (short-chain dehydrogenase/reductase SDR and diguanylate cyclase did not align with M. tuberculosis complex protein sequences, which suggest that these proteins are found only in the NTM strains. Some of the proteins identified as common to all strains can be used as markers of NTM exposure and for the development of new diagnostic tools. Additionally, the specific proteins to NTM strains identified may represent potential candidates for the diagnosis of diseases caused by these mycobacteria.

  13. Changes of Functional and Directed Resting-State Connectivity Are Associated with Neuronal Oscillations, ApoE Genotype and Amyloid Deposition in Mild Cognitive Impairment

    Directory of Open Access Journals (Sweden)

    Lars Michels

    2017-09-01

    Full Text Available The assessment of effects associated with cognitive impairment using electroencephalography (EEG power mapping allows the visualization of frequency-band specific local changes in oscillatory activity. In contrast, measures of coherence and dynamic source synchronization allow for the study of functional and effective connectivity, respectively. Yet, these measures have rarely been assessed in parallel in the context of mild cognitive impairment (MCI and furthermore it has not been examined if they are related to risk factors of Alzheimer’s disease (AD such as amyloid deposition and apolipoprotein ε4 (ApoE allele occurrence. Here, we investigated functional and directed connectivities with Renormalized Partial Directed Coherence (RPDC in 17 healthy controls (HC and 17 participants with MCI. Participants underwent ApoE-genotyping and Pittsburgh compound B positron emission tomography (PiB-PET to assess amyloid deposition. We observed lower spectral source power in MCI in the alpha and beta bands. Coherence was stronger in HC than MCI across different neuronal sources in the delta, theta, alpha, beta and gamma bands. The directed coherence analysis indicated lower information flow between fronto-temporal (including the hippocampus sources and unidirectional connectivity in MCI. In MCI, alpha and beta RPDC showed an inverse correlation to age and gender; global amyloid deposition was inversely correlated to alpha coherence, RPDC and beta and gamma coherence. Furthermore, the ApoE status was negatively correlated to alpha coherence and RPDC, beta RPDC and gamma coherence. A classification analysis of cognitive state revealed the highest accuracy using EEG power, coherence and RPDC as input. For this small but statistically robust (Bayesian power analyses sample, our results suggest that resting EEG related functional and directed connectivities are sensitive to the cognitive state and are linked to ApoE and amyloid burden.

  14. High-Throughput, Multiplex Genotyping Directly from Blood or Dried Blood Spot without DNA Extraction for the Screening of Multiple G6PD Gene Variants at Risk for Drug-Induced Hemolysis.

    Science.gov (United States)

    Tian, Xiaoyi; Zhou, Jun; Zhao, Ye; Cheng, Zhibin; Song, Wenqi; Sun, Yu; Sun, Xiaodong; Zheng, Zhi

    2017-09-01

    Clinical or epidemiologic screening of single-nucleotide polymorphism markers requires large-scale multiplexed genotyping. Available genotyping tools require DNA extraction and multiplex PCR, which may limit throughput and suffer amplification bias. Herein, a novel genotyping approach has been developed, multiplex extension and ligation-based probe amplification (MELPA), which eliminates DNA extraction and achieves uniform PCR amplification. MELPA lyses blood or dried blood spot and directly captures specific target DNA to 96-well plates using tailed probes. Subsequent enzymatic extension and ligation form target single-nucleotide polymorphism-spanning single-stranded templates, which are PCR-amplified using universal primers. Multiplexed genotyping by single-base primer extension is analyzed by mass spectrometry, with a call rate >97%. MELPA was compared with a commercial assay (iPLEX) for detecting 24 G6PD variants known to be at risk for primaquine-induced hemolysis. MELPA provided results that were more reliable than iPLEX, with higher throughput and lower cost. Genotyping archival blood from 106 malaria patients taking primaquine found 10 G6PD-deficient variants, including 1 patient with a hemizygous Mahidol mutation who had hemolysis. Preemptive G6PD genotyping of 438 dried blood spots from a malaria-endemic area identified three variants. MELPA also enabled pooled genotyping without diluting rare alleles, in which undesired common-allele background increased by sample pooling can be repressed by adding specific common allele blockers. Thus, MELPA represents a high-throughput, cost-effective approach to targeted genotyping at the population level. Copyright © 2017 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

  15. Prevalence of Nontuberculous Mycobacteria (NTM in Iranian Clinical Specimens: Systematic Review and Meta-Analysis

    Directory of Open Access Journals (Sweden)

    Azad Khaledi

    2017-01-01

    Full Text Available Background:    Although, nontuberculous mycobacteria can cause disease in different organisms, they usually are not reported in most countries because scientists in general consider them as non-pathogens. But, increasing nontuberculous mycobacteria diseases occurrence has changed this belief. Nevertheless, there is no meta-analysis review about prevalence of nontuberculous mycobacteria in Iran. Methods:   Any data about prevalence of nontuberculous mycobacteria in clinical specimens in Iran were retrieved by searching data bases such as Pub Med, MEDLINE, and Iranian data bases. Then the meta-analysis was performed by comprehensive meta-analysis software (CMA. Results:    The meta-analysis showed that the prevalence of nontuberculous mycobacteria in the clinical specimens in Iran was 1.3%. In the studies that had sample size less than 300, and in studies conducted after 2004, the prevalence was higher. Also, the prevalence of nontuberculous mycobacteria was higher in the West of Iran. In this study, the most prevalent rapid-growing mycobacterium was Mycobacterium. fortuitum and  most prevalent slow-growing mycobacterium was M. simiae with the prevalence 44.2% and 14.3%, respectively.Conclusion:   M. simiae is the most prevalent nontuberculous mycobacteria in the clinical specimens in Iran. As this species of nontuberculous mycobacteria has similar clinical and radiological manifestations with tuberculosis, it is often treated as tuberculosis. Unfortunately, M. simiae is resistant against first-line anti-TB drugs resulting in treatment failure after using routine anti-TB medication. Therefore, there is an urgent need for application of new diagnostic strategy for identification of nontuberculous mycobacteria species.

  16. In vitro activity of flomoxef against rapidly growing mycobacteria.

    Science.gov (United States)

    Tsai, Moan-Shane; Tang, Ya-Fen; Eng, Hock-Liew

    2008-06-01

    The aim of this study was to determine the in vitro sensitivity of rapidly growing mycobacteria (RGM) to flomoxef in respiratory secretions collected from 61 consecutive inpatients and outpatients at Chang Gung Memorial Hospital-Kaohsiung medical center between July and December, 2005. Minimal inhibitory concentrations (MIC) of flomoxef were determined by the broth dilution method for the 61 clinical isolates of RGMs. The MICs of flomoxef at which 90% of clinical isolates were inhibited was >128 microg/mL in 26 isolates of Mycobacterium abscessus and 4 microg/mL in 31 isolates of M. fortuitum. Three out of 4 clinical M. peregrinum isolates were inhibited by flomoxef at concentrations of 4 microg/mL or less. Although the numbers of the clinical isolates of RGMs were small, these preliminary in vitro results demonstrate the potential activity of flomoxef in the management of infections due to M. fortuitum, and probably M. peregrinum in humans.

  17. Natural disasters and nontuberculous mycobacteria: a recipe for increased disease?

    Science.gov (United States)

    Honda, Jennifer R; Bernhard, Jon N; Chan, Edward D

    2015-02-01

    Infectious diseases acquired by survivors of large-scale natural disasters complicate the recovery process. During events such as tsunamis, hurricanes, earthquakes, and tornados and well into the recovery period, victims often are exposed to water-soil mixtures that have relocated with indigenous microbes. Because nontuberculous mycobacteria (NTM) are ubiquitous in water and soil, there is potential for increased exposure to these organisms during natural disasters. In this hypothesis-driven commentary, we discuss the rise in NTM lung disease and natural disasters and examine the geographic overlap of NTM infections and disaster frequencies in the United States. Moreover, we show an increased number of positive NTM cultures from Louisiana residents in the years following three of the relatively recent epic hurricanes and posit that such natural disasters may help to drive the increased number of NTM infections. Finally, we advocate for increased environmental studies and surveillance of NTM infections before and after natural disasters.

  18. Nontuberculous mycobacteria infection after mesotherapy: preliminary report of 15 cases.

    Science.gov (United States)

    Sañudo, Alejandra; Vallejo, Fernando; Sierra, Martha; Hoyos, Juan G; Yepes, Sandra; Wolff, Juan Carlos; Correa, Luis A; Montealegre, Carlos; Navarro, Pilar; Bedoya, Elina; Sanclemente, Gloria

    2007-06-01

    Mesotherapy is an increasingly used technique which is currently causing several mycobacterial infections owing to contaminated substances being injected, and also to poor aseptic measures being held by nonprofessional practitioners. We collected 15 cases of nontuberculous mycobacteria (NTM) infection after mesotherapy in a 6-month period. All patients were female with ages ranging from 19 to 52 years; the main substances injected were procaine and lecithin, and the time between mesotherapy and the appearance of the lesions varied between 1 and 12 weeks. Clinical lesions were mostly nodules and abscesses, which were localized in the abdomen and buttocks in the majority of cases. The main patient complaint was local pain but some presented with systemic symptoms such as fever and malaise. Biopsies reported granulomatous chronic inflammation in the majority of cases. Skin cultures were positive for NTM and Mycobacterium chelonae. Mesotherapy not performed with quality controlled substances can be a predisposing factor for NTM infection.

  19. Recent advances on nontuberculous mycobacteria diseases in Asia

    Directory of Open Access Journals (Sweden)

    Po-Ren Hsueh

    2015-01-01

    Full Text Available The isolation rate of nontuberculous mycobacteria (NTM species and the prevalence of NTM-associated diseases are on the rise worldwide; however, the species distribution of NTM isolates and the types of diseases caused by NTM species vary from region to region. Treatment of a NTM disease is complicated, and there is no comprehensive guideline regarding the in vitro susceptibility of each antimicrobial agent against NTM. Therefore, appropriate anti-NTM treatment can only be recommended based on individual NTM species and local surveillance studies of anti-NTM resistance. Previous studies on the in vitro susceptibility of Mycobacterium avium complex (MAC to clarithromycin in some Asian countries have revealed a low rate of resistance to that antimicrobial agent. Thus, a clarithromycin-based anti-MAC regimen should be effective for MAC infections. However, clarithromycin resistance due to the mutation of the 23S rRNA gene in MAC strains has been detected in many countries. Therefore, physicians should avoid monotherapy with clarithromycin and consider the possibility of clarithromycin resistance in patients who do not respond to clarithromycin-based regimens. Rifampicin is the critical component of successful management of Mycobacterium kansasii diseases. Although most M. kansasii isolates are susceptible to rifampicin in Western countries and in Japan, this agent may not work well in Taiwan. Rapidly growing mycobacteria (RGM is a prevalent NTM group worldwide, particularly in Asia; however, each NTM species in this group may have its own distinct antibiotic susceptibility pattern, and close monitoring of the antibiotic-resistance patterns of RGM is necessary. Most important of all, the in vitro susceptibility may not represent the in vivo activity until the confirmation of the clinical study. Therefore, further investigation of the clinical effectiveness of the anti-NTM agents is warranted.

  20. Utility of rpoB Gene Sequencing for Identification of Nontuberculous Mycobacteria in the Netherlands

    NARCIS (Netherlands)

    Zwaan, R. de; Ingen, J. van; Soolingen, D. van

    2014-01-01

    In the Netherlands, clinical isolation of nontuberculous mycobacteria (NTM) has increased over the past decade. Proper identification of isolates is important, as NTM species differ strongly in clinical relevance. Most of the currently applied identification methods cannot distinguish between all

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

    Science.gov (United States)

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

  2. A low prevalence of mycobacteria in freshwater fish from water reservoirs, ponds and farms

    Czech Academy of Sciences Publication Activity Database

    Mrlík, V.; Slaný, M.; Kubečka, Jan; Seďa, Jaromír; Nečas, A.; Babák, V.; Slaná, I.; Kříž, P.; Pavlík, I.

    2012-01-01

    Roč. 35, č. 7 (2012), s. 497-504 ISSN 0140-7775 Institutional support: RVO:60077344 Keywords : food safety * mycobacteriosis * potentially pathogenic mycobacteria * zoonoses Subject RIV: DA - Hydrology ; Limnology Impact factor: 1.591, year: 2012

  3. Differential Macrophage Response to Slow- and Fast-Growing Pathogenic Mycobacteria

    Directory of Open Access Journals (Sweden)

    A. Cecilia Helguera-Repetto

    2014-01-01

    Full Text Available Nontuberculous mycobacteria (NTM have recently been recognized as important species that cause disease even in immunocompetent individuals. The mechanisms that these species use to infect and persist inside macrophages are not well characterised. To gain insight concerning this process we used THP-1 macrophages infected with M. abscessus, M. fortuitum, M. celatum, and M. tuberculosis. Our results showed that slow-growing mycobacteria gained entrance into these cells with more efficiency than fast-growing mycobacteria. We have also demonstrated that viable slow-growing M. celatum persisted inside macrophages without causing cell damage and without inducing reactive oxygen species (ROS, as M. tuberculosis caused. In contrast, fast-growing mycobacteria destroyed the cells and induced high levels of ROS. Additionally, the macrophage cytokine pattern induced by M. celatum was different from the one induced by either M. tuberculosis or fast-growing mycobacteria. Our results also suggest that, in some cases, the intracellular survival of mycobacteria and the immune response that they induce in macrophages could be related to their growth rate. In addition, the modulation of macrophage cytokine production, caused by M. celatum, might be a novel immune-evasion strategy used to survive inside macrophages that is different from the one reported for M. tuberculosis.

  4. 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.

  5. Naturally Occurring Resistance-Associated Variants to Hepatitis C Virus Direct-Acting Antiviral Agents in Treatment-Naive HCV Genotype 6a-Infected Patients

    Directory of Open Access Journals (Sweden)

    Zhanyi Li

    2017-01-01

    Full Text Available Background and Objective. The direct-acting antiviral agents (DAAs antiviral therapy has drastically improved the prognosis of hepatitis C virus (HCV patients. However, the viral drug resistance-associated variants (RAVs can limit the efficacy of DAAs. For the HCV-6a is not the predominant prevalent genotype; the data on the prevalence of naturally occurring RAVs in it is scarce. Our study aims to assess the prevalence of RAVs in treatment-naive HCV-6a patients. Methods. Nested PCR assays were performed on 95 HCV-6a patients to amplify HCV viral regions of NS3, NS5A, and NS5B. Results. In NS3/4A region, we detected Q80K in 95.5% isolates (84/88 and D168E in 2.3% isolates (2/88. In NS5A region, we detected Q30R in 93.2% isolates (82/88, L31M in 4.6% isolates (4/88, and H58P in 6.8% isolates (6/88. In NS5B region, we detected A15G in 2.3% isolates (2/88, S96T in 1.1% isolates (1/88, and S282T in 20.7% isolates (17/88 and we detected I482L in 100% isolates (4/4, V494A in 50% isolates (2/4, and V499A in 100% isolates (4/4. Conclusions. RAVs to DAAs preexist in treatment-naive HCV-6a patients. Further studies should address the issue of the impact of RAVs in response to DAA therapies for HCV-6a patients.

  6. Pegylated-interferon plus ribavirin treatment does not alter the prevalence of resistance-associated substitutions to direct-acting antivirals in HCV genotype 1a patients

    Directory of Open Access Journals (Sweden)

    Chen Z

    2017-08-01

    Full Text Available Zhi-wei Chen,* Xi-chen Pang,* Zhao Li, Hong Ren, Peng Hu Department of Infectious Diseases, Institute for Viral Hepatitis, The Key Laboratory of Molecular Biology for Infectious Diseases, Chinese Ministry of Education, The Second Affiliated Hospital of Chongqing Medical University, Chongqing, China *These authors contributed equally to this work Background: Direct-acting antiviral (DAA resistance-associated substitutions (RASs can jeopardize the effectiveness of DAAs in patients with hepatitis C virus (HCV. The selection pressure by pegylated-interferon (Peg-IFN plus ribavirin (P/R treatment may enhance HCV genome variation. However, whether P/R treatment alters the rate of change of RASs is still unclear. Materials and methods: We retrieved the genomic sequences of HCV genotype (GT 1a patients from GenBank, which included patients naïve to P/R (pre-IFN group and those previously treated with P/R (post-IFN group. The sequences were aligned and analyzed by using MEGA 6.0 software. Clinically relevant RASs were summarized from the current medical literature. Results: In the cross-sectional study, the total prevalence of clinically relevant RASs was high, independent of the treatment group (pre-IFN: 219/403 [54.34%] vs post-IFN: 67/131 [51.15%]. The high prevalence was mainly detected in the NS3 region RAS at Q80 (40.69% vs 36.64%. The RASs in the NS5A region, such as M28, Q30, L31 and Y93, were uncommon (0%–5%. Similarly, all RASs showed no difference between the two groups. One exception was the RAS at I170 in the NS3 region, which was significantly higher in the post-IFN group than in the pre-IFN group. In the longitudinal study, similar results were observed. However, no difference in RAS at I170 was observed between the two groups. Finally, no clinically relevant RASs were detected in response to the DAA regimens approved for GT 1a patients treated with P/R. Conclusion: Our results suggest that previous P/R treatment failure was not

  7. Comprehensive Screening for Naturally Occurring Hepatitis C Virus Resistance to Direct-Acting Antivirals in the NS3, NS5A, and NS5B Genes in Worldwide Isolates of Viral Genotypes 1 to 6.

    Science.gov (United States)

    Patiño-Galindo, Juan Ángel; Salvatierra, Karina; González-Candelas, Fernando; López-Labrador, F Xavier

    2016-04-01

    There is no comprehensive study available on the natural hepatitis C virus (HCV) polymorphism in sites associated with resistance including all viral genotypes which may present variable susceptibilities to particular direct-acting antivirals (DAAs). This study aimed to analyze the frequencies, genetic barriers, and evolutionary histories of naturally occurring resistance-associated variants (RAVs) in the six main HCV genotypes. A comprehensive analysis of up to 103 RAVs was performed in 2,901, 2,216, and 1,344 HCV isolates for the NS3, NS5A, and NS5B genes, respectively. We report significant intergenotypic differences in the frequencies of natural RAVs for these three HCV genes. In addition, we found a low genetic barrier for the generation of new RAVs, irrespective of the viral genotype. Furthermore, in 1,126 HCV genomes, including sequences spanning the three genes, haplotype analysis revealed a remarkably high frequency of viruses carrying more than one natural RAV to DAAs (53% of HCV-1a, 28.5% of HCV-1b, 67.1% of HCV-6, and 100% of genotype 2, 3, 4, and 5 haplotypes). With the exception of HCV-1a, the most prevalent haplotypes showed RAVs in at least two different viral genes. Finally, evolutionary analyses revealed that, while most natural RAVs appeared recently, others have been efficiently transmitted over time and cluster in well-supported clades. In summary, and despite the observed high efficacy of DAA-based regimens, we show that naturally occurring RAVs are common in all HCV genotypes and that there is an overall low genetic barrier for the selection of resistance mutations. There is a need for natural DAA resistance profiling specific for each HCV genotype. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  8. Pathogenic mycobacteria achieve cellular persistence by inhibiting the Niemann-Pick Type C disease cellular pathway.

    Science.gov (United States)

    Fineran, Paul; Lloyd-Evans, Emyr; Lack, Nathan A; Platt, Nick; Davis, Lianne C; Morgan, Anthony J; Höglinger, Doris; Tatituri, Raju Venkata V; Clark, Simon; Williams, Ian M; Tynan, Patricia; Al Eisa, Nada; Nazarova, Evgeniya; Williams, Ann; Galione, Antony; Ory, Daniel S; Besra, Gurdyal S; Russell, David G; Brenner, Michael B; Sim, Edith; Platt, Frances M

    2016-11-18

    Tuberculosis remains a major global health concern. The ability to prevent phagosome-lysosome fusion is a key mechanism by which intracellular mycobacteria, including Mycobacterium tuberculosis , achieve long-term persistence within host cells. The mechanisms underpinning this key intracellular pro-survival strategy remain incompletely understood. Host macrophages infected with persistent mycobacteria share phenotypic similarities with cells taken from patients suffering from Niemann-Pick Disease Type C (NPC), a rare lysosomal storage disease in which endocytic trafficking defects and lipid accumulation within the lysosome lead to cell dysfunction and cell death. We investigated whether these shared phenotypes reflected an underlying mechanistic connection between mycobacterial intracellular persistence and the host cell pathway dysfunctional in NPC. The induction of NPC phenotypes in macrophages from wild-type mice or obtained from healthy human donors was assessed via infection with mycobacteria and subsequent measurement of lipid levels and intracellular calcium homeostasis. The effect of NPC therapeutics on intracellular mycobacterial load was also assessed. Macrophages infected with persistent intracellular mycobacteria phenocopied NPC cells, exhibiting accumulation of multiple lipid types, reduced lysosomal Ca 2+ levels, and defects in intracellular trafficking. These NPC phenotypes could also be induced using only lipids/glycomycolates from the mycobacterial cell wall. These data suggest that persistent intracellular mycobacteria inhibit the NPC pathway, likely via inhibition of the NPC1 protein, and subsequently induce altered acidic store Ca 2+ homeostasis. Reduced lysosomal calcium levels may provide a mechanistic explanation for the reduced levels of phagosome-lysosome fusion in mycobacterial infection. Treatments capable of correcting defects in NPC mutant cells via modulation of host cell calcium were of benefit in promoting clearance of mycobacteria

  9. Flux through trehalose synthase flows from trehalose to the alpha anomer of maltose in mycobacteria.

    Science.gov (United States)

    Miah, Farzana; Koliwer-Brandl, Hendrik; Rejzek, Martin; Field, Robert A; Kalscheuer, Rainer; Bornemann, Stephen

    2013-04-18

    Trehalose synthase (TreS) was thought to catalyze flux from maltose to trehalose, a precursor of essential trehalose mycolates in mycobacterial cell walls. However, we now show, using a genetic approach, that TreS is not required for trehalose biosynthesis in Mycobacterium smegmatis, whereas two alternative trehalose-biosynthetic pathways (OtsAB and TreYZ) are crucial. Consistent with this direction of flux, trehalose levels in Mycobacterium tuberculosis decreased when TreS was overexpressed. In addition, TreS was shown to interconvert the α anomer of maltose and trehalose using (1)H and (19)F-nuclear magnetic resonance spectroscopies using its normal substrates and deoxyfluoromaltose analogs, with the nonenzymatic mutarotation of α/β-maltose being slow. Therefore, flux through TreS in mycobacteria flows from trehalose to α-maltose, which is the appropriate anomer for maltose kinase of the GlgE α-glucan pathway, which in turn contributes to intracellular and/or capsular polysaccharide biosynthesis. Copyright © 2013 Elsevier Ltd. All rights reserved.

  10. Comparison of the fibronectin-binding ability and antitumor efficacy of various mycobacteria.

    Science.gov (United States)

    Hudson, M A; Ritchey, J K; Catalona, W J; Brown, E J; Ratliff, T L

    1990-07-01

    Although the mechanism by which Bacillus Calmette-Guerin (BCG) exerts an antitumor effect on superficial bladder tumors is not fully understood, recent evidence has implicated binding of BCG organisms to fibronectin (FN) as requisite for this antitumor efficacy. Various substrains of BCG and other mycobacteria were tested in vitro for their relative capacities to bind both matrix and soluble FN. A substrain of Mycobacterium kansasii, designated the "high-binding strain," was found to bind FN more readily (P less than 0.05) in in vitro studies, when compared to commercially available substrains of BCG (Tice, Connaught, and Armand Frappier). The binding by the three commercial strains of BCG to FN in vitro appeared to be equivalent. The high-binding strain was further demonstrated to attach more readily in vivo to the acutely injured murine bladder (P less than 0.005) than the Armand Frappier substrain. Finally, using the MB49 murine bladder tumor model, an enhanced antitumor effect (P less than 0.05) was noted in mice treated with intravesical high-binding strain, in comparison to the Armand Frappier substrain, during five weekly treatments. It appears not only that the commercial substrains of BCG bind FN in an equivalent manner but also that the relative binding capacities of the substrains correlate directly with antitumor activity. A substrain of M. kansasii appears to have been identified which may prove more clinically effective than the currently available strains of BCG.

  11. Detection of molecular markers by comparative sequence analysis of enzymes from mycobacteria species

    International Nuclear Information System (INIS)

    Asad, S.; Hussain, M.; Siddiqua, A.; Ain, Q.U.

    2014-01-01

    Mycobacterial species are one of the most important pathogens and among these members of non-tuberculous mycobacteria (NTM) and mycobacterial tuberculousis complex (MTC) are the causative agent of a relatively milder form of Tuberculosis. Traditional methods for identification of these groups of pathogens are time consuming, lack specificity and sensitivity and furthermore lead to the misidentification due to high similarity index. Therefore, more rapid, specific and cost-effective methods are required for the accurate identification of Mycobacterium species in routine diagnostics. In our study, we identified molecular markers in order to differentiate closely related cousin species of genus Mycobacterium including M. bovis, M. avium, M. leprae and M. tuberculosis. The nucleotide sequences of selected unique markers, i.e., enzymes (used previously in various biochemical tests for the identification of M. species) were selected and their ORFs were retrieved and selected functional proteins of respective biosynthetic pathways were compared in-silico. Result suggested that the variations in nucleotide sequences of the selected enzymes can be directly used for M. species discrimination in one step PCR test. We believe that the in-silico identification and storage of these distinctive characteristics of individual M. species will help in more precise recognition of pathogenic strains and hence specie specific targeted therapy. (author)

  12. Rapidly growing mycobacteria in Singapore, 2006-2011.

    Science.gov (United States)

    Tang, S S; Lye, D C; Jureen, R; Sng, L-H; Hsu, L Y

    2015-03-01

    Nontuberculous mycobacteria infection is a growing global concern, but data from Asia are limited. This study aimed to describe the distribution and antibiotic susceptibility profiles of rapidly growing mycobacterium (RGM) isolates in Singapore. Clinical RGM isolates with antibiotic susceptibility tests performed between 2006 and 2011 were identified using microbiology laboratory databases and minimum inhibitory concentrations of amikacin, cefoxitin, clarithromycin, ciprofloxacin, doxycycline, imipenem, linezolid, moxifloxacin, sulfamethoxazole or trimethoprim-sulfamethoxazole, tigecycline and tobramycin were recorded. Regression analysis was performed to detect changes in antibiotic susceptibility patterns over time. A total of 427 isolates were included. Of these, 277 (65%) were from respiratory specimens, 42 (10%) were related to skin and soft tissue infections and 36 (8%) were recovered from blood specimens. The two most common species identified were Mycobacterium abscessus (73%) and Mycobacterium fortuitum group (22%), with amikacin and clarithromycin being most active against the former, and quinolones and trimethoprim-sulfamethoxazole against the latter. Decreases in susceptibility of M. abscessus to linezolid by 8.8% per year (p 0.001), M. fortuitum group to imipenem by 9.5% per year (p 0.023) and clarithromycin by 4.7% per year (p 0.033) were observed. M. abscessus in respiratory specimens is the most common RGM identified in Singapore. Antibiotic options for treatment of RGM infections are increasingly limited. Copyright © 2014 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

  13. Iron Deprivation Affects Drug Susceptibilities of Mycobacteria Targeting Membrane Integrity

    Directory of Open Access Journals (Sweden)

    Rahul Pal

    2015-01-01

    Full Text Available Multidrug resistance (MDR acquired by Mycobacterium tuberculosis (MTB through continuous deployment of antitubercular drugs warrants immediate search for novel targets and mechanisms. The ability of MTB to sense and become accustomed to changes in the host is essential for survival and confers the basis of infection. A crucial condition that MTB must surmount is iron limitation, during the establishment of infection, since iron is required by both bacteria and humans. This study focuses on how iron deprivation affects drug susceptibilities of known anti-TB drugs in Mycobacterium smegmatis, a “surrogate of MTB.” We showed that iron deprivation leads to enhanced potency of most commonly used first line anti-TB drugs that could be reverted upon iron supplementation. We explored that membrane homeostasis is disrupted upon iron deprivation as revealed by enhanced membrane permeability and hypersensitivity to membrane perturbing agent leading to increased passive diffusion of drug and TEM images showing detectable differences in cell envelope thickness. Furthermore, iron seems to be indispensable to sustain genotoxic stress suggesting its possible role in DNA repair machinery. Taken together, we for the first time established a link between cellular iron and drug susceptibility of mycobacteria suggesting iron as novel determinant to combat MDR.

  14. Nontuberculous Mycobacteria Infections at a Provincial Reference Hospital, Cambodia.

    Science.gov (United States)

    Bonnet, Maryline; San, Kim Chamroeun; Pho, Yati; Sok, Chandara; Dousset, Jean-Philippe; Brant, William; Hurtado, Northan; Eam, Khun Kim; Ardizzoni, Elisa; Heng, Seiha; Godreuil, Sylvain; Yew, Wing-Wai; Hewison, Cathy

    2017-07-01

    Prevalence of nontuberculous mycobacteria (NTM) disease is poorly documented in countries with high prevalence of tuberculosis (TB). We describe prevalence, risk factors, and TB program implications for NTM isolates and disease in Cambodia. A prospective cohort of 1,183 patients with presumptive TB underwent epidemiologic, clinical, radiologic, and microbiologic evaluation, including >12-months of follow-up for patients with NTM isolates. Prevalence of NTM isolates was 10.8% and of disease was 0.9%; 217 (18.3%) patients had TB. Of 197 smear-positive patients, 171 (86.8%) had TB confirmed (167 by culture and 4 by Xpert MTB/RIF assay only) and 11 (5.6%) had NTM isolates. HIV infection and past TB were independently associated with having NTM isolates. Improved detection of NTM isolates in Cambodia might require more systematic use of mycobacterial culture and the use of Xpert MTB/RIF to confirm smear-positive TB cases, especially in patients with HIV infection or a history of TB.

  15. Nontuberculous mycobacteria: incidence in Southwest Ireland from 1987 to 2000.

    LENUS (Irish Health Repository)

    Kennedy, M P

    2012-02-03

    SETTING: The Southwest of Ireland (Counties Cork and Kerry) 1987-2000, average population 549,500. OBJECTIVE: Nontuberculous mycobacteria (NTM) cause significant morbidity worldwide and the study of epidemiology and characteristics helps in their prevention and treatment. This study was performed to determine the incidence of NTM disease in comparison to Mycobacterium tuberculosis (M. tuberculosis) and Mycobacterium bovis (M. bovis) in Southwest Ireland, over the above time period. DESIGN: A retrospective study was carried out in all human isolates of NTM, M. tuberculosis and M. bovis between 1987 and 2000, in the Southwest Region of Ireland. RESULTS: The mean incidence of NTM (0.4\\/100,000 population) has risen since 1995, principally of pulmonary Mycobacterium avium intracellulare complex (MAC). The annual incidence of M. tuberculosis in humans over 14 years in the same region was 971\\/100,000 population with a significant reduction since 1994 and M. bovis remained constant at 0.5\\/100,000 population. CONCLUSION: The increasing incidence of disease causing NTM noted in Southwest Ireland reflects global data and is surmised to be due to an ageing population, increased incidence related to chronic fibrotic lung disease, and environmental mycobacterial factors.

  16. Specific and straightforward molecular investigation of β-thalassemia mutations in the Malaysian Malays and Chinese using direct TaqMan genotyping assays.

    Science.gov (United States)

    Kho, S L; Chua, K H; George, E; Tan, J A M A

    2013-07-15

    Beta-thalassemia is a life-threatening inherited blood disorder. Rapid characterization of β-globin gene mutations is necessary because of the high frequency of Malaysian β-thalassemia carriers. A combination real-time polymerase chain reaction genotyping assay using TaqMan probes was developed to confirm β-globin gene mutations. In this study, primers and probes were designed to specifically identify 8 common β-thalassemia mutations in the Malaysian Malay and Chinese ethnic groups using the Primer Express software. "Blind tests" using DNA samples from healthy individuals and β-thalassemia patients with different genotypes were performed to determine the specificity and sensitivity of this newly designed assay. Our results showed 100% sensitivity and specificity for this novel assay. In conclusion, the TaqMan genotyping assay is a straightforward assay that allows detection of β-globin gene mutations in less than 40 min. The simplicity and reproducibility of the TaqMan genotyping assay permit its use in laboratories as a rapid and cost-effective diagnostic tool for confirmation of common β-thalassemia mutations in Malaysia.

  17. Stable Regulation of Cell Cycle Events in Mycobacteria: Insights From Inherently Heterogeneous Bacterial Populations.

    Science.gov (United States)

    Logsdon, Michelle M; Aldridge, Bree B

    2018-01-01

    Model bacteria, such as E. coli and B. subtilis , tightly regulate cell cycle progression to achieve consistent cell size distributions and replication dynamics. Many of the hallmark features of these model bacteria, including lateral cell wall elongation and symmetric growth and division, do not occur in mycobacteria. Instead, mycobacterial growth is characterized by asymmetric polar growth and division. This innate asymmetry creates unequal birth sizes and growth rates for daughter cells with each division, generating a phenotypically heterogeneous population. Although the asymmetric growth patterns of mycobacteria lead to a larger variation in birth size than typically seen in model bacterial populations, the cell size distribution is stable over time. Here, we review the cellular mechanisms of growth, division, and cell cycle progression in mycobacteria in the face of asymmetry and inherent heterogeneity. These processes coalesce to control cell size. Although Mycobacterium smegmatis and Mycobacterium bovis Bacillus Calmette-Guérin (BCG) utilize a novel model of cell size control, they are similar to previously studied bacteria in that initiation of DNA replication is a key checkpoint for cell division. We compare the regulation of DNA replication initiation and strategies used for cell size homeostasis in mycobacteria and model bacteria. Finally, we review the importance of cellular organization and chromosome segregation relating to the physiology of mycobacteria and consider how new frameworks could be applied across the wide spectrum of bacterial diversity.

  18. Stable Regulation of Cell Cycle Events in Mycobacteria: Insights From Inherently Heterogeneous Bacterial Populations

    Directory of Open Access Journals (Sweden)

    Michelle M. Logsdon

    2018-03-01

    Full Text Available Model bacteria, such as E. coli and B. subtilis, tightly regulate cell cycle progression to achieve consistent cell size distributions and replication dynamics. Many of the hallmark features of these model bacteria, including lateral cell wall elongation and symmetric growth and division, do not occur in mycobacteria. Instead, mycobacterial growth is characterized by asymmetric polar growth and division. This innate asymmetry creates unequal birth sizes and growth rates for daughter cells with each division, generating a phenotypically heterogeneous population. Although the asymmetric growth patterns of mycobacteria lead to a larger variation in birth size than typically seen in model bacterial populations, the cell size distribution is stable over time. Here, we review the cellular mechanisms of growth, division, and cell cycle progression in mycobacteria in the face of asymmetry and inherent heterogeneity. These processes coalesce to control cell size. Although Mycobacterium smegmatis and Mycobacterium bovis Bacillus Calmette-Guérin (BCG utilize a novel model of cell size control, they are similar to previously studied bacteria in that initiation of DNA replication is a key checkpoint for cell division. We compare the regulation of DNA replication initiation and strategies used for cell size homeostasis in mycobacteria and model bacteria. Finally, we review the importance of cellular organization and chromosome segregation relating to the physiology of mycobacteria and consider how new frameworks could be applied across the wide spectrum of bacterial diversity.

  19. Mycobacteria exploit nitric oxide-induced transformation of macrophages into permissive giant cells.

    Science.gov (United States)

    Gharun, Kourosh; Senges, Julia; Seidl, Maximilian; Lösslein, Anne; Kolter, Julia; Lohrmann, Florens; Fliegauf, Manfred; Elgizouli, Magdeldin; Vavra, Martina; Schachtrup, Kristina; Illert, Anna L; Gilleron, Martine; Kirschning, Carsten J; Triantafyllopoulou, Antigoni; Henneke, Philipp

    2017-12-01

    Immunity to mycobacteria involves the formation of granulomas, characterized by a unique macrophage (MΦ) species, so-called multinucleated giant cells (MGC). It remains unresolved whether MGC are beneficial to the host, that is, by prevention of bacterial spread, or whether they promote mycobacterial persistence. Here, we show that the prototypical antimycobacterial molecule nitric oxide (NO), which is produced by MGC in excessive amounts, is a double-edged sword. Next to its antibacterial capacity, NO propagates the transformation of MΦ into MGC, which are relatively permissive for mycobacterial persistence. The mechanism underlying MGC formation involves NO-induced DNA damage and impairment of p53 function. Moreover, MGC have an unsurpassed potential to engulf mycobacteria-infected apoptotic cells, which adds a further burden to their antimycobacterial capacity. Accordingly, mycobacteria take paradoxical advantage of antimicrobial cellular efforts by driving effector MΦ into a permissive MGC state. © 2017 The Authors.

  20. MALDI-TOF MS Andromas strategy for the routine identification of bacteria, mycobacteria, yeasts, Aspergillus spp. and positive blood cultures.

    Science.gov (United States)

    Bille, E; Dauphin, B; Leto, J; Bougnoux, M-E; Beretti, J-L; Lotz, A; Suarez, S; Meyer, J; Join-Lambert, O; Descamps, P; Grall, N; Mory, F; Dubreuil, L; Berche, P; Nassif, X; Ferroni, A

    2012-11-01

    All organisms usually isolated in our laboratory are now routinely identified by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) using the Andromas software. The aim of this study was to describe the use of this strategy in a routine clinical microbiology laboratory. The microorganisms identified included bacteria, mycobacteria, yeasts and Aspergillus spp. isolated on solid media or extracted directly from blood cultures. MALDI-TOF MS was performed on 2665 bacteria isolated on solid media, corresponding to all bacteria isolated during this period except Escherichia coli grown on chromogenic media. All acquisitions were performed without extraction. After a single acquisition, 93.1% of bacteria grown on solid media were correctly identified. When the first acquisition was not contributory, a second acquisition was performed either the same day or the next day. After two acquisitions, the rate of bacteria identified increased to 99.2%. The failures reported on 21 strains were due to an unknown profile attributed to new species (9) or an insufficient quality of the spectrum (12). MALDI-TOF MS has been applied to 162 positive blood cultures. The identification rate was 91.4%. All mycobacteria isolated during this period (22) were correctly identified by MALDI-TOF MS without any extraction. For 96.3% and 92.2% of yeasts and Aspergillus spp., respectively, the identification was obtained with a single acquisition. After a second acquisition, the overall identification rate was 98.8% for yeasts (160/162) and 98.4% (63/64) for Aspergillus spp. In conclusion, the MALDI-TOF MS strategy used in this work allows a rapid and efficient identification of all microorganisms isolated routinely. © 2011 The Authors. Clinical Microbiology and Infection © 2011 European Society of Clinical Microbiology and Infectious Diseases.

  1. Prevalence of Toxoplasma gondii infection in HIV-infected patients and food animals and direct genotyping of T. gondii isolates, Southern Ghana.

    Science.gov (United States)

    Pappoe, Faustina; Cheng, Weisheng; Wang, Lin; Li, Yuanling; Obiri-Yeboah, Dorcas; Nuvor, Samuel Victor; Ambachew, Henock; Hu, Xiaodong; Luo, Qingli; Chu, Deyong; Xu, Yuanhong; Shen, Jilong

    2017-06-01

    Toxoplasma gondii is of public health and veterinary importance causing severe diseases in immunocompromised individuals including HIV/AIDS patients and in congenital cases and animals. There is limited information on the epidemiology of T. gondii infection in humans, particularly HIV patients and food animals and the parasite genotypes in Ghana. A total of 394 HIV-infected patients from three hospitals were screened for T. gondii anti-IgG and IgM using ELISA. DNAs from blood samples of seropositve participants and 95 brain tissues of food animals were PCR assayed to detect Toxoplasma gra6. DNA positive samples were genotyped using multilocus nested polymerase chain reaction restriction fragment length polymorphism at 10 loci: sag1, alt.sag2, sag3, btub, gra6, l358, c22-8, c29-2, pk1, and apico. The overall seroprevalence was 74.37% (293/394). Toxoplasma DNAs were detected in 3.07% of the seropositive participants and 9.47% of the animals. Six of the human DNA positive samples were partly typed at sag3: 33.33, 50, and 16.67% isolates had type I, II, and III alleles, respectively. All nine isolates from food animals typed at nine loci except apico were atypical: six isolates were identical to ToxoDB #41 and #145, and one was identical to TgCkBrRj2 all identified in Brazil. The genotype of two isolates has not been reported previously and was named as TgCtGh1. T. gondii seroprevalence is high among the HIV-infected individuals with T. gondii circulating in Ghana being genetically diverse.

  2. The Genotype MTBDRplus ver. 2.0 test as a quick indicator of resistance to rifampicin and isoniazid in Mycobacterium tuberculosis strains

    Directory of Open Access Journals (Sweden)

    Salvatore Nisticò

    2013-08-01

    Full Text Available Tuberculosis is still a global emergency and a major public health problem, in some cases related to the appearance of strains of multi drug resistance (MDR and extensive drug resistance (XDR Mycobacterium tuberculosis complex.The correct determination of antibiotic sensitivity profiles is therefore crucial to carry out appropriate treatment aimed to decrease the infectivity of each patient and to reduce mortality. The poor adherence to treatment by the patient or the use of therapies based on a single drug, as a result of incorrect requirements, promote the development of drug-resistance. Have some time on the market of molecular diagnostic tests that allow, quickly and directly from biological sample to search for resistance genes some key drugs of anti-TB therapy (Rifampicin and Isoniazid. One of the tests in question is the Genotype MTBDRplus ver 2.0 which can reveal the presence of genes for resistance to Isoniazid (INH and Rifampin (RMP.The loci analyzed are those corresponding to the rpoB gene for rifampicin, katG and inhA for isoniazid. Our study is based on the analysis of 83 strains of tubercular Mycobacteria identified and isolated from patients with tuberculosis disease and subjected to the tests sensitivity, searching for mutations and phenotypic susceptibility testing for Rifampicin and Isoniazid.The comparison of the results has shown that the results obtained using the Genotype MTBDRplus ver 2.0 test, were similar to the results obtained by the traditional susceptibility testing.

  3. Surveillance of Tuberculosis in Taipei: The Influence of Nontuberculous Mycobacteria.

    Directory of Open Access Journals (Sweden)

    Chen-Yuan Chiang

    Full Text Available Notification of tuberculosis (TB but not nontuberculous mycobacteria (NTM is mandatory in Taiwan. Partly due to the strict regulation on TB notification, several patients infected with NTM were notified as TB cases. Notification of patients infected with NTM as TB cases can trigger public health actions and impose additional burdens on the public health system. We conducted a study to assess the influence of NTM infection on surveillance of TB in Taipei.The study population included all individuals with a positive culture for Mycobacterium who were citizens of Taipei City and notified as TB cases in the calendar years 2007-2010. Of the 4216 notified culture-positive tuberculosis (TB cases, 894 (21.2% were infected with NTM. The average annual reported case rate of infection with NTM was 8.6 (95% confidence interval 7.7-9.4 per 100,000 people. The reported case rate of NTM increased with age in both males and females. The proportion of reported TB cases infected with NTM was significantly higher in females than in males (27.6% vs 17.8%, adjusted OR (adjOR 1.93, 95% confidence interval (CI 1.63-2.28; in smear-positive than in smear-negative (23.1% vs 19.2%, adjOR 1.26, 95% CI 1.08-1.47; and in previously treated cases than in new cases (35.7% vs 19.1%, adjOR 2.30, 95% CI 1.88-2.82. The most frequent species was M. avium complex (32.4%, followed by M. chelonae complex (17.6%, M. fortuitum complex (17.0% and M. kansasii (9.8%. Of the 890 notified NTM cases assessed, 703 (79.0% were treated with anti-TB drugs, and 730 (82.0% were de-notified.The influence of NTM on surveillance of TB in Taipei was substantial. Health authorities should take action to ensure that nucleic acid amplification tests are performed in all smear-positive cases in a timely manner to reduce the misdiagnosis of patients infected with NTM as TB cases.

  4. Surveillance of Tuberculosis in Taipei: The Influence of Nontuberculous Mycobacteria.

    Science.gov (United States)

    Chiang, Chen-Yuan; Yu, Ming-Chih; Yang, Shiang-Lin; Yen, Muh-Yong; Bai, Kuan-Jen

    2015-01-01

    Notification of tuberculosis (TB) but not nontuberculous mycobacteria (NTM) is mandatory in Taiwan. Partly due to the strict regulation on TB notification, several patients infected with NTM were notified as TB cases. Notification of patients infected with NTM as TB cases can trigger public health actions and impose additional burdens on the public health system. We conducted a study to assess the influence of NTM infection on surveillance of TB in Taipei. The study population included all individuals with a positive culture for Mycobacterium who were citizens of Taipei City and notified as TB cases in the calendar years 2007-2010. Of the 4216 notified culture-positive tuberculosis (TB) cases, 894 (21.2%) were infected with NTM. The average annual reported case rate of infection with NTM was 8.6 (95% confidence interval 7.7-9.4) per 100,000 people. The reported case rate of NTM increased with age in both males and females. The proportion of reported TB cases infected with NTM was significantly higher in females than in males (27.6% vs 17.8%, adjusted OR (adjOR) 1.93, 95% confidence interval (CI) 1.63-2.28); in smear-positive than in smear-negative (23.1% vs 19.2%, adjOR 1.26, 95% CI 1.08-1.47); and in previously treated cases than in new cases (35.7% vs 19.1%, adjOR 2.30, 95% CI 1.88-2.82). The most frequent species was M. avium complex (32.4%), followed by M. chelonae complex (17.6%), M. fortuitum complex (17.0%) and M. kansasii (9.8%). Of the 890 notified NTM cases assessed, 703 (79.0%) were treated with anti-TB drugs, and 730 (82.0%) were de-notified. The influence of NTM on surveillance of TB in Taipei was substantial. Health authorities should take action to ensure that nucleic acid amplification tests are performed in all smear-positive cases in a timely manner to reduce the misdiagnosis of patients infected with NTM as TB cases.

  5. Characterization of non-tuberculosis mycobacteria by neutron radiography

    International Nuclear Information System (INIS)

    Silva, Jaqueline M. da; Crispim, Verginia Reis; Gomes da Silva, Marlei; Furtado, Vanessa Rodrigues; Duarte, Rafael Da Silva

    2013-01-01

    The genus Mycobacterium shares many characteristics with Corynebacterium and Actinomyces genera, among which the genomic guanine plus cytosine content and the production of long branched-chain fatty acids, known as mycolic acids are enhanced. Growth rate and optimal temperature of mycobacteria are variable. The genus comprises more than 140 known species; however Mycobacterium fortuitum, a fast growing nontuberculous mycobacterium, is clinically significant, because it has been associated to several lesions following surgery procedures such as liposuction, silicone breast and pacemaker implants, exposure to prosthetic materials besides sporadic lesions in the skin, soft tissues and rarely lungs. The objective of the present study is to reduce the time necessary for M. fortuitum characterization based on its morphology and the use of the neutron radiography technique substituting the classical biochemical assays. We also aim to confirm the utility of dendrimers as boron carriers. The samples were sterilized through conventional protocols using 10% formaldehyde. In the incubation process, two solutions with different molar ratios (10:1 and 20:1) of sodium borate and PAMAM G4 dendrimer and also pure sodium borate were used. After doping and sterilization procedures, the samples were deposited on CR-39 sheets, irradiated with a 4.6×10 5 n/cm 2 s thermal neutron flux for 30 min, from the J-9 irradiation channel of the Argonauta IEN/CNEN reactor. The images registered in the CR-39 were visualized in a Nikon E400 optical transmission microscope and captured by a Nikon Coolpix 995 digital camera. Developing the nuclear tracks registered in the CR-39 allowed a 1000× enlargement of mycobacterium images, facilitating their characterization, the use of more sophisticated equipment not being necessary. The use of neutron radiography technique reduced the time necessary for characterization. Doping with PAMAM dendrimer improved the visualization of NTM in neutron radiography

  6. The geographic diversity of nontuberculous mycobacteria isolated from pulmonary samples: an NTM-NET collaborative study

    NARCIS (Netherlands)

    Hoefsloot, W.; Ingen, J. van; Andrejak, C.; Angeby, K.; Bauriaud, R.; Bemer, P.; Beylis, N.; Boeree, M.J.; Cacho, J.; Chihota, V.; Chimara, E.; Churchyard, G.; Cias, R.; Daza, R.; Daley, C.L.; Dekhuijzen, P.N.R.; Domingo, D.; Drobniewski, F.; Esteban, J. De; Fauville-Dufaux, M.; Folkvardsen, D.B.; Gibbons, N.; Gomez-Mampaso, E.; Gonzalez, R.; Hoffmann, H.; Hsueh, P.R.; Indra, A.; Jagielski, T.; Jamieson, F.; Jankovic, M.; Jong, E. de; Keane, J.; Koh, W.J.; Lange, B. de; Leao, S.; Macedo, R.; Mannsaker, T.; Marras, T.K.; Maugein, J.; Milburn, H.J.; Mlinko, T.; Morcillo, N.; Morimoto, K.; Papaventsis, D.; Palenque, E.; Paez-Pena, M.; Piersimoni, C.; Polanova, M.; Rastogi, N.; Richter, E.; Ruiz-Serrano, M.J.; Silva, A.; Silva, M.P. da; Simsek, H.; Soolingen, D. van; Szabo, N.; Thomson, R.; Fernandez, T. Tortola; Tortoli, E.; Totten, S.E.; Tyrrell, G.; Vasankari, T.; Villar, M.; Walkiewicz, R.; Winthrop, K.L.; Wagner, D.; Trials, G. Nontuberculous

    2013-01-01

    A significant knowledge gap exists concerning the geographical distribution of nontuberculous mycobacteria (NTM) isolation worldwide. To provide a snapshot of NTM species distribution, global partners in the NTM-Network European Trials Group (NET) framework (www.ntm-net.org), a branch of the

  7. Analysis of recombinant mycobacteria as T helper type 1 vanccines in an allergy challange model

    NARCIS (Netherlands)

    Janssen, R.; Kruisselbrink, A.; Hoogteyling, L.; Lamb, J.R.; Young, D.B.; Thole, J.E.R.

    2001-01-01

    The potential for development of mycobacteria as T helper type 1 (Th1) vaccines capable of induction of Th1 responses to recombinant antigens was explored in a model system based on an immunodominant peptide from house dust mite. Different recombinant mycobacterial preparations were compared for

  8. Epidemiology of nontuberculous mycobacteria among patients with cystic fibrosis in Scandinavia

    DEFF Research Database (Denmark)

    Qvist, Tavs; Gilljam, Marita; Jönsson, Bodil

    2015-01-01

    BACKGROUND: Nontuberculous mycobacteria (NTM) are an emerging threat to cystic fibrosis (CF) patients but their epidemiology is not well described. METHODS: In this retrospective observational study we identified all Scandinavian CF patients with a positive NTM culture from airway secretions from...

  9. Decolorization of Malachite Green and Crystal Violet by Waterborne Pathogenic Mycobacteria

    OpenAIRE

    Jones, Jefferson J.; Falkinham III, Joseph O.

    2003-01-01

    Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium scrofulaceum, Mycobacterium marinum, and Mycobacterium chelonae tolerate high concentrations of the dyes malachite green and crystal violet. Cells of strains of those species decolorized (reduced) both malachite green and crystal violet. Because decolorized malachite green lacked antimicrobial activity, the resistance of these mycobacteria could be due, in part, to their ability to decolorize the dyes. Small amounts of malachite...

  10. Production of monoclonal antibodies against Mycobacterium leprae and armadillo-derived mycobacteria

    NARCIS (Netherlands)

    Kolk, A. H.; Ho, M. L.; Klatser, P. R.; Eggelte, T. A.; Portaels, F.

    1985-01-01

    Six monoclonal antibodies to Mycobacterium leprae and armadillo-derived mycobacteria were produced. The monoclonal antibodies were characterized by an immunofluorescence assay using 22 mycobacterial strains. One monoclonal antibody, F47-21-3, reacted only with M. leprae; two, F45-9 and F45-15,

  11. Structural insights into the mycobacteria transcription initiation complex from analysis of X-ray crystal structures

    Energy Technology Data Exchange (ETDEWEB)

    Hubin, Elizabeth A.; Lilic, Mirjana; Darst, Seth A.; Campbell, Elizabeth A.

    2017-07-13

    The mycobacteria RNA polymerase (RNAP) is a target for antimicrobials against tuberculosis, motivating structure/function studies. Here we report a 3.2 Å-resolution crystal structure of a Mycobacterium smegmatis (Msm) open promoter complex (RPo), along with structural analysis of the Msm RPo and a previously reported 2.76 Å-resolution crystal structure of an Msm transcription initiation complex with a promoter DNA fragment. We observe the interaction of the Msm RNAP α-subunit C-terminal domain (αCTD) with DNA, and we provide evidence that the αCTD may play a role in Mtb transcription regulation. Our results reveal the structure of an Actinobacteria-unique insert of the RNAP β' subunit. Finally, our analysis reveals the disposition of the N-terminal segment of Msm σA, which may comprise an intrinsically disordered protein domain unique to mycobacteria. The clade-specific features of the mycobacteria RNAP provide clues to the profound instability of mycobacteria RPo compared with E. coli.

  12. In vitro Anti-mycobacteria Sensitivity and Kill-kinetics of Allium ...

    African Journals Online (AJOL)

    Allium ascalonicum L. (Shallot) was one of the herbs repeatedly identified from the result of our ethnobotanical survey for the treatment of tuberculosis and nontuberculous mycobacteria diseases. It has been reported to show inhibitory potentials against several pathogens. This plant is also known to form part of the diet of ...

  13. Structural analysis of biofilm formation by rapidly and slowly growing nontuberculous mycobacteria

    Science.gov (United States)

    Mycobacterium avium complex (MAC) and rapidly growing mycobacteria (RGM) such as M. abscessus, M. mucogenicum, M. chelonae and M. fortuitum, implicated in healthcare-associated infections, are often isolated from potable water supplies as part of the microbial flora. To understa...

  14. The tracing of mycobacteria in drinking water supply systems by culture, conventional, and real time PCRs

    Czech Academy of Sciences Publication Activity Database

    Klanicová, B.; Seďa, Jaromír; Slaná, I.; Slaný, M.; Pavlík, I.

    2013-01-01

    Roč. 67, č. 6 (2013), s. 725-731 ISSN 0343-8651 Institutional support: RVO:60077344 Keywords : avium subsp paratuberculosis * nontuberculous mycobacteria * potable water * survival * intracellulare Subject RIV: EE - Microbiology, Virology Impact factor: 1.359, year: 2013

  15. Mycobacteria-specific cytokine responses as correlates of treatment response in active and latent tuberculosis.

    Science.gov (United States)

    Clifford, Vanessa; Tebruegge, Marc; Zufferey, Christel; Germano, Susie; Forbes, Ben; Cosentino, Lucy; McBryde, Emma; Eisen, Damon; Robins-Browne, Roy; Street, Alan; Denholm, Justin; Curtis, Nigel

    2017-08-01

    A biomarker indicating successful tuberculosis (TB) therapy would assist in determining appropriate length of treatment. This study aimed to determine changes in mycobacteria-specific antigen-induced cytokine biomarkers in patients receiving therapy for latent or active TB, to identify biomarkers potentially correlating with treatment success. A total of 33 adults with active TB and 36 with latent TB were followed longitudinally over therapy. Whole blood stimulation assays using mycobacteria-specific antigens (CFP-10, ESAT-6, PPD) were done on samples obtained at 0, 1, 3, 6 and 9 months. Cytokine responses (IFN-γ, IL-1ra, IL-2, IL-10, IL-13, IP-10, MIP-1β, and TNF-α) in supernatants were measured by Luminex xMAP immunoassay. In active TB cases, median IL-1ra (with CFP-10 and with PPD stimulation), IP-10 (CFP-10, ESAT-6), MIP-1β (ESAT-6, PPD), and TNF-α (ESAT-6) responses declined significantly over the course of therapy. In latent TB cases, median IL-1ra (CFP-10, ESAT-6, PPD), IL-2 (CFP-10, ESAT-6), and IP-10 (CFP-10, ESAT-6) responses declined significantly. Mycobacteria-specific cytokine responses change significantly over the course of therapy, and their kinetics in active TB differ from those observed in latent TB. In particular, mycobacteria-specific IL-1ra responses are potential correlates of successful therapy in both active and latent TB. Copyright © 2017 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

  16. Isolation of mycobacteria other than Mycobacterium avium from porcine lymph nodes

    NARCIS (Netherlands)

    Ingen, van J.; Wisselink, H.J.; Solt-Smits, van C.B.; Boeree, M.J.; Soolingen, D.

    2010-01-01

    Mycobacterium avium causes lymphadenitis in pigs. This presents an economical burden, as these pigs meat is considered inappropriate for consumption. In humans, lymphadenitis due to nontuberculous mycobacteria (NTM) primarily affects children and is caused by a variety of NTM, though M. avium

  17. Isolation of mycobacteria other than Mycobacterium avium from porcine lymph nodes.

    NARCIS (Netherlands)

    Ingen, J. van; Wisselink, H.J.; Solt-Smits, C.B. van; Boeree, M.J.; Soolingen, D. van

    2010-01-01

    Mycobacterium avium causes lymphadenitis in pigs. This presents an economical burden, as these pigs meat is considered inappropriate for consumption. In humans, lymphadenitis due to nontuberculous mycobacteria (NTM) primarily affects children and is caused by a variety of NTM, though M. avium

  18. Pulmonary infection with hypervirulent Mycobacteria reveals a crucial role for the P2X7 receptor in aggressive forms of tuberculosis.

    Directory of Open Access Journals (Sweden)

    Eduardo P Amaral

    2014-07-01

    Full Text Available The purinergic P2X7 receptor (P2X7R is a sensor of extracellular ATP, a damage-associated molecule that is released from necrotic cells and that induces pro-inflammatory cytokine production and cell death. To investigate whether the innate immune response to damage signals could contribute to the development of pulmonary necrotic lesions in severe forms of tuberculosis, disease progression was examined in C57BL/6 and P2X7R-/- mice that were intratracheally infected with highly virulent mycobacterial strains (Mycobacterium tuberculosis strain 1471 of the Beijing genotype family and Mycobacterium bovis strain MP287/03. The low-dose infection of C57BL/6 mice with bacteria of these strains caused the rapid development of extensive granulomatous pneumonia with necrotic areas, intense bacillus dissemination and anticipated animal death. In contrast, in P2X7R-/- mice, the lung pathology presented with moderate infiltrates of mononuclear leukocytes without visible signs of necrosis; the disease attenuation was accompanied by a delay in mortality. In vitro, the hypervirulent mycobacteria grew rapidly inside macrophages and induced death by a P2X7R-dependent mechanism that facilitated the release of bacilli. Furthermore, these bacteria were resistant to the protective mechanisms elicited in macrophages following extracellular ATP stimulation. Based on this study, we propose that the rapid intracellular growth of hypervirulent mycobacteria results in massive macrophage damage. The ATP released by damaged cells engages P2X7R and accelerates the necrotic death of infected macrophages and the release of bacilli. This vicious cycle exacerbates pneumonia and lung necrosis by promoting widespread cell destruction and bacillus dissemination. These findings suggest the use of drugs that have been designed to inhibit the P2X7R as a new therapeutic approach to treat the aggressive forms of tuberculosis.

  19. Elephant grass genotypes for bioenergy production by direct biomass combustion Genótipos de capim-elefante para produção de bioenergia por combustão direta da biomassa

    Directory of Open Access Journals (Sweden)

    Rafael Fiusa de Morais

    2009-02-01

    Full Text Available The objective of this work was to evaluate elephant grass (Pennisetum purpureum Schum. genotypes for bioenergy production by direct biomass combustion. Five elephant grass genotypes grown in two different soil types, both of low fertility, were evaluated. The experiment was carried out at Embrapa Agrobiologia field station in Seropédica, RJ, Brazil. The design was in randomized complete blocks, with split plots and four replicates. The genotypes studied were Cameroon, Bag 02, Gramafante, Roxo and CNPGL F06-3. Evaluations were made for biomass production, total biomass nitrogen, biomass nitrogen from biological fixation, carbon/nitrogen and stem/leaf ratios, and contents of fiber, lignin, cellulose and ash. The dry matter yields ranged from 45 to 67 Mg ha-1. Genotype Roxo had the lowest yield and genotypes Bag 02 and Cameroon had the highest ones. The biomass nitrogen accumulation varied from 240 to 343 kg ha-1. The plant nitrogen from biological fixation was 51% in average. The carbon/nitrogen and stem/leaf ratios and the contents of fiber, lignin, cellulose and ash did not vary among the genotypes. The five genotypes are suitable for energy production through combustion.O objetivo deste trabalho foi avaliar genótipos de capim-elefante (Pennisetum purpureum Schum. quanto ao potencial para a produção de bioenergia por combustão direta da biomassa. Avaliaram-se cinco genótipos de capim-elefante, em dois solos com baixa fertilidade. Os experimentos foram conduzidos na estação experimental da Embrapa Agrobiologia, em Seropédica, RJ. O delineamento experimental foi o de blocos ao acaso, em parcelas subdivididas, com quatro repetições. Os genótipos estudados foram Cameroon, Bag 02, Gramafante, Roxo e CNPGL F06-3. Determinaram-se a produção de biomassa, o acúmulo de nitrogênio na biomassa, o nitrogênio da biomassa proveniente da fixação biológica, as relações carbono/nitrogênio e talo/folha, e os teores de fibra, lignina

  20. Comparison among three methods for mycobacteria identification Comparación entre tres métodos para identificar micobacterias

    OpenAIRE

    Misael Mondragón-Barreto; Carlos A. Vázquez-Chacón; Candelaria Barrón-Rivero; Patricia Acosta-Blanco; Kenneth C. Jost Jr; Susana Balandrano; Hiram Olivera-Díaz

    2000-01-01

    OBJECTIVE: To compare three methods: Biochemical tests, high-performance liquid chromatography (HPLC) and polymerase chain reaction-restriction fragments length polymorphism (PCR-RFLP), for the identification of mycobacteria, and to perform a cost-benefit analysis to define an optimum identification algorithm. MATERIAL AND METHODS: One-hundred-and-seven mycobacteria isolates were identified by the three methods at Instituto de Diagnóstico y Referencia Epidemiológicos, between February of 1999...

  1. A pilot evaluation of external quality assessment of GenoType MTBDRplus versions 1 and 2 using dried culture spot material.

    Science.gov (United States)

    Gous, N; Isherwood, L E; David, A; Stevens, W; Scott, L E

    2015-04-01

    Dried culture spots (DCS) of inactivated Mycobacteria strains designed as part of an external quality assessment (EQA) program for the GeneXpert system has applications to other molecular tuberculosis (TB) diagnostic platforms. DCS tested on the GenoType MTBDRplus and Mycobacterium CM assays performed well with MTBDRplus version 2 but require increased bacterial concentration for use with version 1. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  2. Mycobacteria emulsified in olive oil-in-water trigger a robust immune response in bladder cancer treatment

    Science.gov (United States)

    Noguera-Ortega, Estela; Blanco-Cabra, Núria; Rabanal, Rosa Maria; Sánchez-Chardi, Alejandro; Roldán, Mónica; Guallar-Garrido, Sandra; Torrents, Eduard; Luquin, Marina; Julián, Esther

    2016-01-01

    The hydrophobic composition of mycobacterial cell walls leads to the formation of clumps when attempting to resuspend mycobacteria in aqueous solutions. Such aggregation may interfere in the mycobacteria-host cells interaction and, consequently, influence their antitumor effect. To improve the immunotherapeutic activity of Mycobacterium brumae, we designed different emulsions and demonstrated their efficacy. The best formulation was initially selected based on homogeneity and stability. Both olive oil (OO)- and mineral oil-in-water emulsions better preserved the mycobacteria viability and provided higher disaggregation rates compared to the others. But, among both emulsions, the OO emulsion increased the mycobacteria capacity to induce cytokines’ production in bladder tumor cell cultures. The OO-mycobacteria emulsion properties: less hydrophobic, lower pH, more neutralized zeta potential, and increased affinity to fibronectin than non-emulsified mycobacteria, indicated favorable conditions for reaching the bladder epithelium in vivo. Finally, intravesical OO-M. brumae-treated mice showed a significantly higher systemic immune response, together with a trend toward increased tumor-bearing mouse survival rates compared to the rest of the treated mice. The physicochemical characteristics and the induction of a robust immune response in vitro and in vivo highlight the potential of the OO emulsion as a good delivery vehicle for the mycobacterial treatment of bladder cancer. PMID:27265565

  3. Cultivable mycobacteria in sphagnum vegetation of moors in South Sweden and coastal Norway.

    Science.gov (United States)

    Kazda, J; Müller, K; Irgens, L M

    1979-04-01

    Intact sphagnum vegetation from moors in south Sweden and coastal areas of west Norway contained cultivable mycobacteria in 32% and 30% of the specimens, respectively. This frequency of specimens is lower than the 50% previously found in the partly altered moors of northwestern Germany, but the Scandinavian moors contained a larger variety of species. On both intact and altered moors M. chelonei and M. sphagni sp. nov. were found, the latter a homologous group of 151 strains. In south Sweden the highest frequency was found in S. balticum, S. recurvum. S. tenellum and S. compactum & molle. (40-65%). In coastal Norway the highest frequency was found in S. rubellum (48%) which offers favourable conditions for the accumulation of solar energy due to the red brown colour in the upper parts. Combined with a high humidity in coastal Norway in summer, this may contribute to the growth of mesophilic mycobacteria. A significant affinity of M. chelonei to S. tenellum was stated.

  4. Using riboswitches to regulate gene expression and define gene function in mycobacteria.

    Science.gov (United States)

    Van Vlack, Erik R; Seeliger, Jessica C

    2015-01-01

    Mycobacteria include both environmental species and many pathogenic species such as Mycobacterium tuberculosis, an intracellular pathogen that is the causative agent of tuberculosis in humans. Inducible gene expression is a powerful tool for examining gene function and essentiality, both in in vitro culture and in host cell infections. The theophylline-inducible artificial riboswitch has recently emerged as an alternative to protein repressor-based systems. The riboswitch is translationally regulated and is combined with a mycobacterial promoter that provides transcriptional control. We here provide methods used by our laboratory to characterize the riboswitch response to theophylline in reporter strains, recombinant organisms containing riboswitch-regulated endogenous genes, and in host cell infections. These protocols should facilitate the application of both existing and novel artificial riboswitches to the exploration of gene function in mycobacteria. © 2015 Elsevier Inc. All rights reserved.

  5. Distribution of nontuberculous mycobacteria in treated patients with pulmonary disease in Greece - relation to microbiological data.

    Science.gov (United States)

    Manika, Katerina; Tsikrika, Stamatoula; Tsaroucha, Emilia; Karabela, Simona; Karachaliou, Iris; Bosmi, Ioulia; Kioumis, Ioannis; Papavasileiou, Apostolos

    2015-01-01

    The aim was to assess the distribution of nontuberculous mycobacteria (NTM) in treated patients with pulmonary disease (PD) in Greece. Patients treated for NTM PD at the two largest chest diseases hospitals in Greece, in the period 1990-2013 were investigated. For the years 2005-2013 data on NTM isolation frequency were recorded. M. avium complex (MAC) was the predominant cause of NTM PD disease followed by M. kansasii and rapid growing mycobacteria (RGM). The pathogenicity of RGM was significantly lower than this of MAC and M. kansasii. An increase was observed in the percentage of isolated NTM species that were considered clinically significant over the study period. The increasing number of NTM PD in Greece is a consequence of their isolation being more frequently considered as clinically relevant.

  6. Non-tuberculous mycobacteria and microbial populations in drinking water distribution systems

    Directory of Open Access Journals (Sweden)

    Rossella Briancesco

    2010-01-01

    Full Text Available Data on the occurrence of non-tuberculous mycobacteria (NTM, in parallel with those obtained for bacterial indicators and amoebae, are presented with the aim to collect information on the spread of NTM in drinking water distribution systems in Italy. Samples were collected from taps of hospitals and households in Central and Southern Italy. The concentration values obtained for the more traditional microbial parameters complied with the mandatory requirements for drinking water. Conversely, moderate-to-high microbial loads (till 300 CFU/L were observed for the NTM. Positive samples were obtained from 62% of the investigated water samples. Analogous results were observed for amoebae showing a higher percentage of positive samples (76%. In terms of public health, the presence of mycobacteria in water distribution systems may represent a potential risk especially for vulnerable people such as children, the elderly or immunocompromised individuals.

  7. Nontuberculous mycobacteria in Denmark, incidence and clinical importance during the last quarter-century

    DEFF Research Database (Denmark)

    Hermansen, Thomas S; Ravn, Pernille; Svensson, Erik

    2017-01-01

    and trends in annual incidence rates. 524,119 clinical specimens were cultured for mycobacteria from 1991 through 2015 at the International Reference Laboratory of Mycobacteriology in Denmark. Among these, 8,227 NTM strains were identified from 3,462 patients and distributed according to microbiological...... disease criteria. We observed no increase in NTM disease incidence or proportion of patients with positive NTM cultures during the study period (Quasi-Poisson regression, p = 0.275 and 0.352 respectively). Annual incidence rates were 1.20/105 for definite NTM disease, 0.49/105 for possible NTM disease...... and 0.88/105 for NTM colonization. The incidence rate of NTM disease was highest in children aged 0-4 years (5.36/105/year), predominantly with cervical Mycobacterium avium complex (MAC) adenitis. Surprisingly, based on more than half a million clinical specimens cultured for mycobacteria in Denmark...

  8. Nontuberculous mycobacteria in Denmark, incidence and clinical importance during the last quarter-century

    DEFF Research Database (Denmark)

    Hermansen, Thomas S; Ravn, Pernille; Svensson, Erik

    2017-01-01

    and trends in annual incidence rates. 524,119 clinical specimens were cultured for mycobacteria from 1991 through 2015 at the International Reference Laboratory of Mycobacteriology in Denmark. Among these, 8,227 NTM strains were identified from 3,462 patients and distributed according to microbiological...... disease criteria. We observed no increase in NTM disease incidence or proportion of patients with positive NTM cultures during the study period (Quasi-Poisson regression, p = 0.275 and 0.352 respectively). Annual incidence rates were 1.20/10(5) for definite NTM disease, 0.49/10(5) for possible NTM disease...... and 0.88/10(5) for NTM colonization. The incidence rate of NTM disease was highest in children aged 0-4 years (5.36/10(5)/year), predominantly with cervical Mycobacterium avium complex (MAC) adenitis. Surprisingly, based on more than half a million clinical specimens cultured for mycobacteria in Denmark...

  9. Clinical and Taxonomic Status of Pathogenic Nonpigmented or Late-Pigmenting Rapidly Growing Mycobacteria

    OpenAIRE

    Brown-Elliott, Barbara A.; Wallace, Richard J.

    2002-01-01

    The history, taxonomy, geographic distribution, clinical disease, and therapy of the pathogenic nonpigmented or late-pigmenting rapidly growing mycobacteria (RGM) are reviewed. Community-acquired disease and health care-associated disease are highlighted for each species. The latter grouping includes health care-associated outbreaks and pseudo-outbreaks as well as sporadic disease cases. Treatment recommendations for each species and type of disease are also described. Special emphasis is on ...

  10. Understanding the relative contributions of direct environmental effects and passive genotype-environment correlations in the association between familial risk factors and child disruptive behavior disorders.

    Science.gov (United States)

    Bornovalova, M A; Cummings, J R; Hunt, E; Blazei, R; Malone, S; Iacono, W G

    2014-03-01

    Previous work reports an association between familial risk factors stemming from parental characteristics and offspring disruptive behavior disorders (DBDs). This association may reflect (a) the direct effects of familial environment and (b) a passive gene-environment correlation (r(GE)), wherein the parents provide both the genes and the environment. The current study examined the contributions of direct environmental influences and passive r(GE) by comparing the effects of familial risk factors on child DBDs in genetically related (biological) and non-related (adoptive) families. Participants were 402 adoptive and 204 biological families. Familial environment was defined as maternal and paternal maladaptive parenting and antisociality, marital conflict and divorce; offspring DBDs included attention deficit hyperactivity disorder (ADHD), conduct disorder (CD) and oppositional defiant disorder (ODD). Mixed-level regressions estimated the main effects of familial environment, adoption status and the familial environment by adoption status interaction term, which tested for the presence of passive r(GE). There was a main effect of maternal and paternal maladaptive parenting and marital discord on child DBDs, indicating a direct environmental effect. There was no direct environmental effect of maternal or paternal antisociality, but maternal and paternal antisociality had stronger associations with child DBDs in biological families than adoptive families, indicating the presence of a passive r(GE). Many familial risk factors affected children equally across genetically related and non-related families, providing evidence for direct environmental effects. The relationship of parental antisociality and offspring DBDs was best explained by a passive r(GE), where a general vulnerability toward externalizing psychopathology is passed down by the parents to the children.

  11. ESX-1-mediated translocation to the cytosol controls virulence of mycobacteria

    KAUST Repository

    Houben, Diane; Demangel, Caroline; Van Ingen, Jakko; Perez, Jorge; Baldeó n, Lucy R.; Abdallah, Abdallah; Caleechurn, Laxmee; Bottai, Daria; Van Zon, Maaike; De Punder, Karin; Van Der Laan, Tridia; Kant, Arie; Bossers-De Vries, Ruth; Willemsen, Peter Th J; Bitter, Wilbert M.; Van Soolingen, Dick; Brosch, Roland; Van Der Wel, Nicole N.; Peters, Peter J.

    2012-01-01

    Mycobacterium species, including Mycobacterium tuberculosis and Mycobacterium leprae, are among the most potent human bacterial pathogens. The discovery of cytosolic mycobacteria challenged the paradigm that these pathogens exclusively localize within the phagosome of host cells. As yet the biological relevance of mycobacterial translocation to the cytosol remained unclear. In this current study we used electron microscopy techniques to establish a clear link between translocation and mycobacterial virulence. Pathogenic, patient-derived mycobacteria species were found to translocate to the cytosol, while non-pathogenic species did not. We were further able to link cytosolic translocation with pathogenicity by introducing the ESX-1 (type VII) secretion system into the non-virulent, exclusively phagolysosomal Mycobacterium bovis BCG. Furthermore, we show that translocation is dependent on the C-terminus of the early-secreted antigen ESAT-6. The C-terminal truncation of ESAT-6 was shown to result in attenuation in mice, again linking translocation to virulence. Together, these data demonstrate the molecular mechanism facilitating translocation of mycobacteria. The ability to translocate from the phagolysosome to the cytosol is with this study proven to be biologically significant as it determines mycobacterial virulence. © 2012 Blackwell Publishing Ltd.

  12. Antimicrobial susceptibility testing of rapidly growing mycobacteria by microdilution - Experience of a tertiary care centre

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    Set R

    2010-01-01

    Full Text Available Purpose: The objective of the study was to perform antimicrobial susceptibility testing of rapidly growing mycobacteria (RGM isolated from various clinically suspected cases of extrapulmonary tuberculosis, from January 2007 to April 2008, at a tertiary care centre in Mumbai. Materials and Methods: The specimens were processed for microscopy and culture using the standard procedures. Minimum inhibitory concentrations (MIC were determined by broth microdilution, using Sensititre CA MHBT. Susceptibility testing was also carried out on Mueller Hinton agar by the Kirby Bauer disc diffusion method. Results: Of the 1062 specimens received for mycobacterial cultures, 104 (9.79% grew mycobacteria. Of the mycobacterial isolates, six (5.76% were rapid growers. M. abscessus and M. chelonae appeared to be resistant organisms, with M. chelonae showing intermediate resistance to amikacin and minocycline. However, all the six isolates showed sensitivity to vancomycin and gentamicin by the disc diffusion test. Also all three isolates of M. abscessus were sensitive to piperacillin and erythromycin. Further studies are required to test their sensitivity to these four antimicrobials by using the microbroth dilution test, before they can be prescribed to patients. Conclusions: We wish to emphasize that reporting of rapidly growing mycobacteria from clinical settings, along with their sensitivity patterns, is an absolute need of the hour.

  13. Nontuberculous Mycobacteria Isolation from Clinical and Environmental Samples in Iran: Twenty Years of Surveillance

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    Ali Akbar Velayati

    2015-01-01

    Full Text Available Nontuberculous mycobacteria (NTM are opportunistic pathogens that are widely distributed in the environment. There is a lack of data on species distribution of these organisms from Iran. This study consists of a review of NTM articles published in Iran between the years 1992 and 2014. In this review, 20 articles and 14 case reports were identified. Among the 20 articles, 13 (65% studies focused on NTM isolates from clinical specimens, 6 (30% studies examined NTM isolates from environmental samples, and one (5% article included both clinical and environmental isolates. M. fortuitum (229/997; 23% was recorded as the most prevalent and rapid growing mycobacteria (RGM species in both clinical (28% and environmental (19% isolated samples (P < 0.05. Among slow growing mycobacteria (SGM, M. simiae (103/494; 21% demonstrated a higher frequency in clinical samples whereas in environmental samples it was M. flavescens (44/503; 9%. These data represent information from 14 provinces out of 31 provinces of Iran. No information is available in current published data on clinical or environmental NTM from the remaining 17 provinces in Iran. These results emphasize the potential importance of NTM as well as the underestimation of NTM frequency in Iran. NTM is an important clinical problem associated with significant morbidity and mortality in Iran. Continued research is needed from both clinical and environmental sources to help clinicians and researchers better understand and address NTM treatment and prevention.

  14. Nontuberculous Mycobacteria Isolation from Clinical and Environmental Samples in Iran: Twenty Years of Surveillance.

    Science.gov (United States)

    Velayati, Ali Akbar; Farnia, Parissa; Mozafari, Mohadese; Mirsaeidi, Mehdi

    2015-01-01

    Nontuberculous mycobacteria (NTM) are opportunistic pathogens that are widely distributed in the environment. There is a lack of data on species distribution of these organisms from Iran. This study consists of a review of NTM articles published in Iran between the years 1992 and 2014. In this review, 20 articles and 14 case reports were identified. Among the 20 articles, 13 (65%) studies focused on NTM isolates from clinical specimens, 6 (30%) studies examined NTM isolates from environmental samples, and one (5%) article included both clinical and environmental isolates. M. fortuitum (229/997; 23%) was recorded as the most prevalent and rapid growing mycobacteria (RGM) species in both clinical (28%) and environmental (19%) isolated samples (P < 0.05). Among slow growing mycobacteria (SGM), M. simiae (103/494; 21%) demonstrated a higher frequency in clinical samples whereas in environmental samples it was M. flavescens (44/503; 9%). These data represent information from 14 provinces out of 31 provinces of Iran. No information is available in current published data on clinical or environmental NTM from the remaining 17 provinces in Iran. These results emphasize the potential importance of NTM as well as the underestimation of NTM frequency in Iran. NTM is an important clinical problem associated with significant morbidity and mortality in Iran. Continued research is needed from both clinical and environmental sources to help clinicians and researchers better understand and address NTM treatment and prevention.

  15. MicroRNA-155 promotes autophagy to eliminate intracellular mycobacteria by targeting Rheb.

    Science.gov (United States)

    Wang, Jinli; Yang, Kun; Zhou, Lin; Minhaowu; Wu, Yongjian; Zhu, Min; Lai, Xiaomin; Chen, Tao; Feng, Lianqiang; Li, Meiyu; Huang, Chunyu; Zhong, Qiu; Huang, Xi

    2013-01-01

    Mycobacterium tuberculosis is a hard-to-eradicate intracellular pathogen that infects one-third of the global population. It can live within macrophages owning to its ability to arrest phagolysosome biogenesis. Autophagy has recently been identified as an effective way to control the intracellular mycobacteria by enhancing phagosome maturation. In the present study, we demonstrate a novel role of miR-155 in regulating the autophagy-mediated anti-mycobacterial response. Both in vivo and in vitro studies showed that miR-155 expression was significantly enhanced after mycobacterial infection. Forced expression of miR-155 accelerated the autophagic response in macrophages, thus promoting the maturation of mycobacterial phagosomes and decreasing the survival rate of intracellular mycobacteria, while transfection with miR-155 inhibitor increased mycobacterial survival. However, macrophage-mediated mycobacterial phagocytosis was not affected after miR-155 overexpression or inhibition. Furthermore, blocking autophagy with specific inhibitor 3-methyladenine or silencing of autophagy related gene 7 (Atg7) reduced the ability of miR-155 to promote autophagy and mycobacterial elimination. More importantly, our study demonstrated that miR-155 bound to the 3'-untranslated region of Ras homologue enriched in brain (Rheb), a negative regulator of autophagy, accelerated the process of autophagy and sequential killing of intracellular mycobacteria by suppressing Rheb expression. Our results reveal a novel role of miR-155 in regulating autophagy-mediated mycobacterial elimination by targeting Rheb, and provide potential targets for clinical treatment.

  16. MicroRNA-155 promotes autophagy to eliminate intracellular mycobacteria by targeting Rheb.

    Directory of Open Access Journals (Sweden)

    Jinli Wang

    Full Text Available Mycobacterium tuberculosis is a hard-to-eradicate intracellular pathogen that infects one-third of the global population. It can live within macrophages owning to its ability to arrest phagolysosome biogenesis. Autophagy has recently been identified as an effective way to control the intracellular mycobacteria by enhancing phagosome maturation. In the present study, we demonstrate a novel role of miR-155 in regulating the autophagy-mediated anti-mycobacterial response. Both in vivo and in vitro studies showed that miR-155 expression was significantly enhanced after mycobacterial infection. Forced expression of miR-155 accelerated the autophagic response in macrophages, thus promoting the maturation of mycobacterial phagosomes and decreasing the survival rate of intracellular mycobacteria, while transfection with miR-155 inhibitor increased mycobacterial survival. However, macrophage-mediated mycobacterial phagocytosis was not affected after miR-155 overexpression or inhibition. Furthermore, blocking autophagy with specific inhibitor 3-methyladenine or silencing of autophagy related gene 7 (Atg7 reduced the ability of miR-155 to promote autophagy and mycobacterial elimination. More importantly, our study demonstrated that miR-155 bound to the 3'-untranslated region of Ras homologue enriched in brain (Rheb, a negative regulator of autophagy, accelerated the process of autophagy and sequential killing of intracellular mycobacteria by suppressing Rheb expression. Our results reveal a novel role of miR-155 in regulating autophagy-mediated mycobacterial elimination by targeting Rheb, and provide potential targets for clinical treatment.

  17. ESX-1-mediated translocation to the cytosol controls virulence of mycobacteria

    KAUST Repository

    Houben, Diane

    2012-05-08

    Mycobacterium species, including Mycobacterium tuberculosis and Mycobacterium leprae, are among the most potent human bacterial pathogens. The discovery of cytosolic mycobacteria challenged the paradigm that these pathogens exclusively localize within the phagosome of host cells. As yet the biological relevance of mycobacterial translocation to the cytosol remained unclear. In this current study we used electron microscopy techniques to establish a clear link between translocation and mycobacterial virulence. Pathogenic, patient-derived mycobacteria species were found to translocate to the cytosol, while non-pathogenic species did not. We were further able to link cytosolic translocation with pathogenicity by introducing the ESX-1 (type VII) secretion system into the non-virulent, exclusively phagolysosomal Mycobacterium bovis BCG. Furthermore, we show that translocation is dependent on the C-terminus of the early-secreted antigen ESAT-6. The C-terminal truncation of ESAT-6 was shown to result in attenuation in mice, again linking translocation to virulence. Together, these data demonstrate the molecular mechanism facilitating translocation of mycobacteria. The ability to translocate from the phagolysosome to the cytosol is with this study proven to be biologically significant as it determines mycobacterial virulence. © 2012 Blackwell Publishing Ltd.

  18. EVALUATION OF THE EFFECT OF SPUTUM STORAGE CONDITIONS ON THE VITAL PROPERTIES OF TUBERCULOUS MYCOBACTERIA

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    Yu. D. Rodionova

    2017-01-01

    Full Text Available Goal of the study: to enhance efficiency of laboratory diagnostics of tuberculous infection through investigating the effect of various storage conditions of samples on the growth properties of Tuberculosis complex mycobacteria.Materials and Methods. 2058 samples of sputum collected by coughing were examined. All sputum samples were aliquoted into 5 parts stored under various conditions: in the fridge under +7°С for 2 hours; frozen for 7 days; by room indoor temperature for 48-72 hours; with the use preservative 10% solution of triple-substituted natrium phosphate for 48 hours; in the fridge under +7°С for 2 hours with consequent treatment by 1% solution of N-acetyl-L-cysteine.Results. The most optimal sputum storage conditions are freezing by -20°С, providing maximum safety of mycobacteria and minimum contamination of the samples with foreign bacteria. Sputum storage by indoor room temperature for 2-3 days reduces the number of positive results of cultures on nutritive media. Using 10% solution of triple-substituted natrium phosphate provides high positive results of bacterioscopic and cultural testing techniques. Time prolongation for biomaterials storage in the preservative solution for more than 72 hours results in the death tuberculous mycobacteria

  19. Hepatitis C virus genotypes in Myanmar.

    Science.gov (United States)

    Win, Nan Nwe; Kanda, Tatsuo; Nakamoto, Shingo; Yokosuka, Osamu; Shirasawa, Hiroshi

    2016-07-21

    Myanmar is adjacent to India, Bangladesh, Thailand, Laos and China. In Myanmar, the prevalence of hepatitis C virus (HCV) infection is 2%, and HCV infection accounts for 25% of hepatocellular carcinoma. In this study, we reviewed the prevalence of HCV genotypes in Myanmar. HCV genotypes 1, 3 and 6 were observed in volunteer blood donors in and around the Myanmar city of Yangon. Although there are several reports of HCV genotype 6 and its variants in Myanmar, the distribution of the HCV genotypes has not been well documented in areas other than Yangon. Previous studies showed that treatment with peginterferon and a weight-based dose of ribavirin for 24 or 48 wk could lead to an 80%-100% sustained virological response (SVR) rates in Myanmar. Current interferon-free treatments could lead to higher SVR rates (90%-95%) in patients infected with almost all HCV genotypes other than HCV genotype 3. In an era of heavy reliance on direct-acting antivirals against HCV, there is an increasing need to measure HCV genotypes, and this need will also increase specifically in Myanmar. Current available information of HCV genotypes were mostly from Yangon and other countries than Myanmar. The prevalence of HCV genotypes in Myanmar should be determined.

  20. Validation of biomarkers for distinguishing Mycobacterium tuberculosis from non-tuberculous mycobacteria using gas chromatography-mass spectrometry and chemometrics.

    Directory of Open Access Journals (Sweden)

    Ngoc A Dang

    Full Text Available Tuberculosis (TB remains a major international health problem. Rapid differentiation of Mycobacterium tuberculosis complex (MTB from non-tuberculous mycobacteria (NTM is critical for decisions regarding patient management and choice of therapeutic regimen. Recently we developed a 20-compound model to distinguish between MTB and NTM. It is based on thermally assisted hydrolysis and methylation gas chromatography-mass spectrometry and partial least square discriminant analysis. Here we report the validation of this model with two independent sample sets, one consisting of 39 MTB and 17 NTM isolates from the Netherlands, the other comprising 103 isolates (91 MTB and 12 NTM from Stellenbosch, Cape Town, South Africa. All the MTB strains in the 56 Dutch samples were correctly identified and the model had a sensitivity of 100% and a specificity of 94%. For the South African samples the model had a sensitivity of 88% and specificity of 100%. Based on our model, we have developed a new decision-tree that allows the differentiation of MTB from NTM with 100% accuracy. Encouraged by these findings we will proceed with the development of a simple, rapid, affordable, high-throughput test to identify MTB directly in sputum.

  1. [Soft-tissue infections due to non-tuberculous mycobacteria following mesotherapy. What is the price of beauty].

    Science.gov (United States)

    Rivera-Olivero, Ismar Alejandra; Guevara, Armando; Escalona, Arnelly; Oliver, Margarita; Pérez-Alfonzo, Ricardo; Piquero, Jaime; Zerpa, Olga; de Waard, Jacobus H

    2006-05-01

    Mesotherapy is widely used In Latin America for cosmetic purposes, particularly in obese individuals. We describe the clinical and epidemiological characteristics, microbiological diagnosis, treatment and follow-up of patients from Caracas (Venezuela) with soft tissue infection caused by non-tuberculous mycobacteria following mesotherapy. Between March 2002 and December 2003, we evaluated 49 cases of skin and soft tissue infection following mesotherapy. Specimens obtained from the lesions and 15 products used in the mesotherapy procedure were cultured for the presence of non-tuberculous mycobacteria. Isolated mycobacteria were identified by PCR restriction fragment length polymorphism analysis of the hsp65 gene. Infection by non-tuberculous mycobacteria was confirmed in 81.6% of the 49 cases. Mycobacterium abscessus and M. fortuitum were the most common species, but M. chelonae, M. peregrinum, M. simiae and a new species that was designated "M. cosmeticum" were also isolated. Patients were treated with species-specific antibiotic agents for 3 to 18 months. Investigation into the source of the infection revealed that 21 patients were clustered within 3 different outbreaks and two products were found to be contaminated with M. fortuitum and M. abscessus, respectively. Physicians should be alerted to the possibility of infection by non-tuberculous mycobacteria in patients with a history of mesotherapy who develop late-onset skin and soft tissue infection, particularly if they do not respond to conventional antibiotic treatment.

  2. Azurophil granule proteins constitute the major mycobactericidal proteins in human neutrophils and enhance the killing of mycobacteria in macrophages.

    Directory of Open Access Journals (Sweden)

    Prajna Jena

    Full Text Available Pathogenic mycobacteria reside in, and are in turn controlled by, macrophages. However, emerging data suggest that neutrophils also play a critical role in innate immunity to tuberculosis, presumably by their different antibacterial granule proteins. In this study, we purified neutrophil azurophil and specific granules and systematically analyzed the antimycobacterial activity of some purified azurophil and specific granule proteins against M. smegmatis, M. bovis-BCG and M. tuberculosis H37Rv. Using gel overlay and colony forming unit assays we showed that the defensin-depleted azurophil granule proteins (AZP were more active against mycobacteria compared to other granule proteins and cytosolic proteins. The proteins showing antimycobacterial activity were identified by MALDI-TOF mass spectrometry. Electron microscopic studies demonstrate that the AZP disintegrate bacterial cell membrane resulting in killing of mycobacteria. Exogenous addition of AZP to murine macrophage RAW 264.7, THP-1 and peripheral blood monocyte-derived macrophages significantly reduced the intracellular survival of mycobacteria without exhibiting cytotoxic activity on macrophages. Immunofluorescence studies showed that macrophages actively endocytose neutrophil granular proteins. Treatment with AZP resulted in increase in co-localization of BCG containing phagosomes with lysosomes but not in increase of autophagy. These data demonstrate that neutrophil azurophil proteins may play an important role in controlling intracellular survival of mycobacteria in macrophages.

  3. The Comparison of Biochemical and Sequencing 16S rDNA Gene Methods to Identify Nontuberculous Mycobacteria

    Directory of Open Access Journals (Sweden)

    Shafipour1, M.

    2014-11-01

    Full Text Available The identification of Mycobacteria in the species level has great medical importance. Biochemical tests are laborious and time-consuming, so new techniques could be used to identify the species. This research aimed to the comparison of biochemical and sequencing 16S rDNA gene methods to identify nontuberculous Mycobacteria in patients suspected to tuberculosis in Golestan province which is the most prevalent region of tuberculosis in Iran. Among 3336 patients suspected to tuberculosis referred to hospitals and health care centres in Golestan province during 2010-2011, 319 (9.56% culture positive cases were collected. Identification of species by using biochemical tests was done. On the samples recognized as nontuberculous Mycobacteria, after DNA extraction by boiling, 16S rDNA PCR was done and their sequencing were identified by NCBI BLAST. Of the 319 positive samples in Golestan Province, 300 cases were M.tuberculosis and 19 cases (5.01% were identified as nontuberculous Mycobacteria by biochemical tests. 15 out of 19 nontuberculous Mycobacteria were identified by PCR and sequencing method as similar by biochemical methods (similarity rate: 78.9%. But after PCR, 1 case known as M.simiae by biochemical test was identified as M. lentiflavum and 3 other cases were identified as Nocardia. Biochemical methods corresponded to the 16S rDNA PCR and sequencing in 78.9% of cases. However, in identification of M. lentiflavum and Nocaria sp. the molecular method is better than biochemical methods.

  4. Techniques of DNA hybridization detect small numbers of mycobacteria with no cross-hybridization with non-mycobacterial respiratory organisms

    International Nuclear Information System (INIS)

    Shoemaker, S.A.; Fisher, J.H.; Scoggin, C.H.

    1985-01-01

    The traditional methods used in identifying mycobacteria, such as acid-fast bacillus stains and culture, are often time-consuming, insensitive, and nonspecific. As part of an ongoing program to improve diagnosis and characterization of mycobacteria, the authors have found that deoxyribonucleic acid (DNA) hybridization techniques using isotopically labeled, single-stranded, total DNA can be used to detect as little as 10(-4) micrograms of Mycobacterium tuberculosis (MTb) DNA. This amount of DNA represents approximately 2 X 10(4) genomes. They have also shown the MTb DNA is sufficiently different from the DNA of non-mycobacterial microorganisms such that cross-hybridization with MTb DNA does not occur under the hybridization conditions employed. The authors speculate that DNA hybridization techniques may allow the rapid, sensitive, and specific identification of mycobacteria

  5. Molecular epidemiology of nontuberculous mycobacteria isolates from clinical and environmental sources of a metropolitan city.

    Science.gov (United States)

    Velayati, Ali Akbar; Farnia, Parissa; Mozafari, Mohadese; Malekshahian, Donya; Seif, Shima; Rahideh, Snaz; Mirsaeidi, Mehdi

    2014-01-01

    While NTM infection is mainly acquired from environmental exposure, monitoring of environmental niches for NTM is not a routine practice. This study aimed to find the prevalence of environmental NTM in soil and water in four highly populated suburbs of Tehran, Iran. A total of 4014 samples from soil and water resources were collected and studied. Sediments of each treated sample were cultured in Lowenstein-Jensen medium and observed twice per week for growth rate, colony morphology, and pigmentation. Colonies were studied with phenotypic tests. Molecular analysis was performed on single colonies derived from subculture of original isolates. Environmental samples were compared with 34 NTM isolates from patients who were residents of the study locations. Out of 4014 samples, mycobacteria were isolated from 862 (21.4%) specimens; 536 (62.1%) belonged to slow growing mycobacteria (SGM) and 326 (37.8%) were rapid growing mycobacteria (RGM). The five most frequent NTM were M. farcinogens (105/862; 12.1%), M. fortuitum (72/862; 8.3%), M. senegalense (58/862; 6.7%), M. kansasii (54/862; 6.2%), and M. simiae (46/862; 5.3%). In total, 62.5% (539/862) of mycobacterial positive samples were isolated from water and only 37.4% (323/862) of them were isolated from soil samples (Pdistribution pattern of environmental NTM isolates with clinical isolates suggests a possible transmission link, but this does not apply to all environmental NTM species. Our study confirms an increasing trend of NTM isolation from clinical samples that needs further investigation.

  6. Th1-skewed tissue responses to a mycolyl glycolipid in mycobacteria-infected rhesus macaques

    Energy Technology Data Exchange (ETDEWEB)

    Morita, Daisuke; Miyamoto, Ayumi; Hattori, Yuki; Komori, Takaya [Laboratory of Cell Regulation, Institute for Virus Research, Kyoto University, 53 Kawahara-cho, Shogoin, Sakyo-ku, Kyoto 606-8507 (Japan); Nakamura, Takashi [Faculty of Pharmaceutical Sciences, Hokkaido University, Kita 12 Nishi 6, Kita-ku, Sapporo, Hokkaido 060-0812 (Japan); Igarashi, Tatsuhiko [Laboratory of Primate Model, Institute for Virus Research, Kyoto University, 53 Kawahara-cho, Shogoin, Sakyo-ku, Kyoto 606-8507 (Japan); Harashima, Hideyoshi [Faculty of Pharmaceutical Sciences, Hokkaido University, Kita 12 Nishi 6, Kita-ku, Sapporo, Hokkaido 060-0812 (Japan); Sugita, Masahiko [Laboratory of Cell Regulation, Institute for Virus Research, Kyoto University, 53 Kawahara-cho, Shogoin, Sakyo-ku, Kyoto 606-8507 (Japan)

    2013-11-08

    Highlights: •Glucose monomycolate (GMM) is a marker glycolipid for active tuberculosis. •Tissue responses to GMM involved up-regulation of Th1-attracting chemokines. •Th1-skewed local responses were mounted at the GMM-injected tissue. -- Abstract: Trehalose 6,6′-dimycolate (TDM) is a major glycolipid of the cell wall of mycobacteria with remarkable adjuvant functions. To avoid detection by the host innate immune system, invading mycobacteria down-regulate the expression of TDM by utilizing host-derived glucose as a competitive substrate for their mycolyltransferases; however, this enzymatic reaction results in the concomitant biosynthesis of glucose monomycolate (GMM) which is recognized by the acquired immune system. GMM-specific, CD1-restricted T cell responses have been detected in the peripheral blood of infected human subjects and monkeys as well as in secondary lymphoid organs of small animals, such as guinea pigs and human CD1-transgenic mice. Nevertheless, it remains to be determined how tissues respond at the site where GMM is produced. Here we found that rhesus macaques vaccinated with Mycobacterium bovis bacillus Calmette–Guerin mounted a chemokine response in GMM-challenged skin that was favorable for recruiting T helper (Th)1 T cells. Indeed, the expression of interferon-γ, but not Th2 or Th17 cytokines, was prominent in the GMM-injected tissue. The GMM-elicited tissue response was also associated with the expression of monocyte/macrophage-attracting CC chemokines, such as CCL2, CCL4 and CCL8. Furthermore, the skin response to GMM involved the up-regulated expression of granulysin and perforin. Given that GMM is produced primarily by pathogenic mycobacteria proliferating within the host, the Th1-skewed tissue response to GMM may function efficiently at the site of infection.

  7. Th1-skewed tissue responses to a mycolyl glycolipid in mycobacteria-infected rhesus macaques

    International Nuclear Information System (INIS)

    Morita, Daisuke; Miyamoto, Ayumi; Hattori, Yuki; Komori, Takaya; Nakamura, Takashi; Igarashi, Tatsuhiko; Harashima, Hideyoshi; Sugita, Masahiko

    2013-01-01

    Highlights: •Glucose monomycolate (GMM) is a marker glycolipid for active tuberculosis. •Tissue responses to GMM involved up-regulation of Th1-attracting chemokines. •Th1-skewed local responses were mounted at the GMM-injected tissue. -- Abstract: Trehalose 6,6′-dimycolate (TDM) is a major glycolipid of the cell wall of mycobacteria with remarkable adjuvant functions. To avoid detection by the host innate immune system, invading mycobacteria down-regulate the expression of TDM by utilizing host-derived glucose as a competitive substrate for their mycolyltransferases; however, this enzymatic reaction results in the concomitant biosynthesis of glucose monomycolate (GMM) which is recognized by the acquired immune system. GMM-specific, CD1-restricted T cell responses have been detected in the peripheral blood of infected human subjects and monkeys as well as in secondary lymphoid organs of small animals, such as guinea pigs and human CD1-transgenic mice. Nevertheless, it remains to be determined how tissues respond at the site where GMM is produced. Here we found that rhesus macaques vaccinated with Mycobacterium bovis bacillus Calmette–Guerin mounted a chemokine response in GMM-challenged skin that was favorable for recruiting T helper (Th)1 T cells. Indeed, the expression of interferon-γ, but not Th2 or Th17 cytokines, was prominent in the GMM-injected tissue. The GMM-elicited tissue response was also associated with the expression of monocyte/macrophage-attracting CC chemokines, such as CCL2, CCL4 and CCL8. Furthermore, the skin response to GMM involved the up-regulated expression of granulysin and perforin. Given that GMM is produced primarily by pathogenic mycobacteria proliferating within the host, the Th1-skewed tissue response to GMM may function efficiently at the site of infection

  8. Molecular epidemiology of nontuberculous mycobacteria isolates from clinical and environmental sources of a metropolitan city.

    Directory of Open Access Journals (Sweden)

    Ali Akbar Velayati

    Full Text Available While NTM infection is mainly acquired from environmental exposure, monitoring of environmental niches for NTM is not a routine practice. This study aimed to find the prevalence of environmental NTM in soil and water in four highly populated suburbs of Tehran, Iran.A total of 4014 samples from soil and water resources were collected and studied. Sediments of each treated sample were cultured in Lowenstein-Jensen medium and observed twice per week for growth rate, colony morphology, and pigmentation. Colonies were studied with phenotypic tests. Molecular analysis was performed on single colonies derived from subculture of original isolates. Environmental samples were compared with 34 NTM isolates from patients who were residents of the study locations.Out of 4014 samples, mycobacteria were isolated from 862 (21.4% specimens; 536 (62.1% belonged to slow growing mycobacteria (SGM and 326 (37.8% were rapid growing mycobacteria (RGM. The five most frequent NTM were M. farcinogens (105/862; 12.1%, M. fortuitum (72/862; 8.3%, M. senegalense (58/862; 6.7%, M. kansasii (54/862; 6.2%, and M. simiae (46/862; 5.3%. In total, 62.5% (539/862 of mycobacterial positive samples were isolated from water and only 37.4% (323/862 of them were isolated from soil samples (P<0.05. Out of 5314 positive clinical samples for mycobacteria, 175 (3.2% isolates were NTM. The trend of NTM isolates increased from 1.2% (13 out of 1078 in 2004 to 3.8% (39 out of 1005 in 2014 (P = 0.0001. The major clinical isolates were M. simiae (51; 29.1%, M. kansasii (26; 14.8%, M. chelonae (28; 16%, and M. fortuitum (13; 7.4%.Comparing the distribution pattern of environmental NTM isolates with clinical isolates suggests a possible transmission link, but this does not apply to all environmental NTM species. Our study confirms an increasing trend of NTM isolation from clinical samples that needs further investigation.

  9. MicroRNAs play big roles in modulating macrophages response toward mycobacteria infection.

    Science.gov (United States)

    Abdalla, Abualgasim Elgaili; Duan, Xiangke; Deng, Wanyan; Zeng, Jie; Xie, Jianping

    2016-11-01

    Macrophages are crucial player in the defense against multiple intracellular pathogens. Mycobacterium tuberculosis, the causative agent of tuberculosis which inflicted around one third of global population, can replicate and persist within macrophages. MicroRNAs, endogenous, small noncoding RNA, can regulate the expression of macrophages genes required for appropriate signaling. Mycobacteria can manipulate the expression of macrophages microRNAs to subvert cell response for its survival and persistence. This review summarized the progress of microRNAs in mycobacterial pathogenesis. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. [Identification of mycobacteria to the species level by molecular methods in the Public Health Laboratory of Bogotá, Colombia].

    Science.gov (United States)

    Hernández-Toloza, Johana Esther; Rincón-Serrano, María de Pilar; Celis-Bustos, Yamile Adriana; Aguillón, Claudia Inés

    2016-01-01

    Global epidemiology of non-tuberculous mycobacteria (NTM) is unknown due to the fact that notification is not required in many countries, however the number of infection reports and outbreaks caused by NTM suggest a significant increase in the last years. Traditionally, mycobacteria identification is made through biochemical profiles which allow to differentiate M. tuberculosis from NTM, and in some cases the mycobacteria species. Nevertheless, these methods are technically cumbersome and time consuming. On the other hand, the introduction of methods based on molecular biology has improved the laboratory diagnosis of NTM. To establish the NTM frequency in positive cultures for acid-fast bacilli (AAFB) which were sent to Laboratorio de Salud Pública de Bogotá over a 12 month period. A total of 100 positive cultures for acid-fast bacilli from public and private hospitals from Bogotá were identified by both biochemical methods and the molecular methods PRA (PCR-restriction enzyme analysis) and multiplex-PCR. Furthermore, low prevalence mycobacteria species and non-interpretable results were confirmed by 16SrDNA sequentiation analysis. Identification using the PRA method showed NMT occurrence in 11% of cultures. In addition, this molecular methodology allowed to detect the occurrence of more than one mycobacteria in 4% of the cultures. Interestingly, a new M. kubicae pattern of PCR-restriction analysis is reported in our study. Using a mycobacteria identification algorithm, which includes the molecular method PRA, improves the diagnostic power of conventional methods and could help to advance both NTM epidemiology knowledge and mycobacteriosis control. Copyright © 2015 Elsevier España, S.L.U. y Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

  11. Identification of a Novel Conjugative Plasmid in Mycobacteria That Requires Both Type IV and Type VII Secretion

    KAUST Repository

    Ummels, R.

    2014-09-23

    Conjugative plasmids have been identified in a wide variety of different bacteria, ranging from proteobacteria to firmicutes, and conjugation is one of the most efficient routes for horizontal gene transfer. The most widespread mechanism of plasmid conjugation relies on different variants of the type IV secretion pathway. Here, we describe the identification of a novel type of conjugative plasmid that seems to be unique for mycobacteria. Interestingly, while this plasmid is efficiently exchanged between different species of slow-growing mycobacteria, including Mycobacterium tuberculosis, it could not be transferred to any of the fast-growing mycobacteria tested. Genetic analysis of the conjugative plasmid showed the presence of a locus containing homologues of three type IV secretion system components and a relaxase. In addition, a new type VII secretion locus was present. Using transposon insertion mutagenesis, we show that in fact both these secretion systems are essential for conjugation, indicating that this plasmid represents a new class of conjugative plasmids requiring two secretion machineries. This plasmid could form a useful new tool to exchange or introduce DNA in slow-growing mycobacteria. IMPORTANCE: Conjugative plasmids play an important role in horizontal gene transfer between different bacteria and, as such, in their adaptation and evolution. This effect is most obvious in the spread of antibiotic resistance genes. Thus far, conjugation of natural plasmids has been described only rarely for mycobacterial species. In fact, it is generally accepted that M. tuberculosis does not show any recent sign of horizontal gene transfer. In this study, we describe the identification of a new widespread conjugative plasmid that can also be efficiently transferred to M. tuberculosis. This plasmid therefore poses both a threat and an opportunity. The threat is that, through the acquisition of antibiotic resistance markers, this plasmid could start a rapid spread of

  12. Direct stamp of technology or origin on the genotypic and phenotypic variation of indigenous Saccharomyces cerevisiae population in a natural model of boiled grape juice fermentation into traditional Msalais wine in China.

    Science.gov (United States)

    Zhu, Li-Xia; Wang, Guan-Qiong; Xue, Ju-Lan; Gou, Dong-Qi; Duan, Chang-Qing

    2017-08-01

    Saccharomyces cerevisiae strains worldwide show genetic and phenotypic diversity and their population substructures are greatly affected by their technological application or geographical origins. Msalais is a traditional wine obtained via a unique method of spontaneous fermentation of local boiled grape juice in Southern Xinjiang. We analyzed 436 indigenous S. cerevisiae strains associated with Msalais fermentation. These strains were highly diverse with respect to the interdelta region and 24 phenotypic traits, with apparent differentiation according to strain origins and technologies used to produce Msalais. The genetic and phenotypic diversity of strains from traditional workshops was higher than in strains from modern plants. These local strains had different origin- or technology-specific fermentative characteristics. Strains growing in large-scale fermentation tanks tolerated high temperature, whereas strains from traditional workshops tolerated high alcohol content (16%) and low temperature (13°C). Almost all the strains were characterized by the highest fermenting vigor, with weak H2S production and no histamine, cadaverine, phenethylamine and tryptamine production. Majority of strains had pronounced autolytic activity with high β-glucosidase and polygalacturonase activity and alcohol production. Our study reveals a direct stamp of technology or origin on genotypic and phenotypic variation of an indigenous S. cerevisiae population. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  13. Utility and diagnostic performance of Mycobacterium tuberculosis complex by two immunochromatographic assays as compared with the molecular Genotype assay in Nigeria

    Directory of Open Access Journals (Sweden)

    Benjamin Thumamo Pokam

    2013-01-01

    Full Text Available Among the disadvantages of smear microscopy for detection of tuberculosis cases is its inability to differentiate between Mycobacterium tuberculosis (MTB and non-tuberculous mycobacteria (NTM. This study evaluated two, new immunochromatographic assays – Capilia TB-Neo and SD Bioline – on unheated and heated cultures at 80 °C for 30 min respectively for their ability to discriminate between MTB complex and NTM as compared with the molecular Genotype assay. Mycobacteria used in the study were obtained from smear-positive specimens collected from patients at four major hospitals in Cross River State, Nigeria. Capilia TB-Neo and SD Bioline showed sensitivities of 98.8% and 93.8% respectively and 100% specificity for both assays. Heating the isolates did not significantly impact the test performance. Both tests are recommended for use in rapid differentiation of strains isolated in Nigeria.

  14. Engineering more stable, selectable marker-free autoluminescent mycobacteria by one step.

    Science.gov (United States)

    Yang, Feng; Njire, Moses M; Liu, Jia; Wu, Tian; Wang, Bangxing; Liu, Tianzhou; Cao, Yuanyuan; Liu, Zhiyong; Wan, Junting; Tu, Zhengchao; Tan, Yaoju; Tan, Shouyong; Zhang, Tianyu

    2015-01-01

    In our previous study, we demonstrated that the use of the autoluminescent Mycobacterium tuberculosis as a reporter strain had the potential to drastically reduce the time, effort, animals and costs consumed in evaluation of the activities of drugs and vaccines in live mice. However, the strains were relatively unstable and lost reporter with time without selection. The kanamycin selection marker used wasn't the best choice as it provides resistance to amino glycosides which are an important class of second line drugs used in tuberculosis treatment. In addition, the marker could limit utility of the strains for screening of new potential drugs or evaluating drug combinations for tuberculosis treatment. Limited selection marker genes for mycobacterial genetic manipulation is a major drawback for such a marker-containing strain in many research fields. Therefore, selectable marker-free, more stable autoluminescent mycobacteria are highly needed. After trying several strategies, we created such mycobacterial strains successfully by using an integrative vector and removing both the resistance maker and integrase genes by Xer site-specific recombination in one step. The corresponding plasmid vectors developed in this study could be very convenient in constructing other selectable marker-free, more stable reporter mycobacteria with diverse applications.

  15. Engineering more stable, selectable marker-free autoluminescent mycobacteria by one step.

    Directory of Open Access Journals (Sweden)

    Feng Yang

    Full Text Available In our previous study, we demonstrated that the use of the autoluminescent Mycobacterium tuberculosis as a reporter strain had the potential to drastically reduce the time, effort, animals and costs consumed in evaluation of the activities of drugs and vaccines in live mice. However, the strains were relatively unstable and lost reporter with time without selection. The kanamycin selection marker used wasn't the best choice as it provides resistance to amino glycosides which are an important class of second line drugs used in tuberculosis treatment. In addition, the marker could limit utility of the strains for screening of new potential drugs or evaluating drug combinations for tuberculosis treatment. Limited selection marker genes for mycobacterial genetic manipulation is a major drawback for such a marker-containing strain in many research fields. Therefore, selectable marker-free, more stable autoluminescent mycobacteria are highly needed. After trying several strategies, we created such mycobacterial strains successfully by using an integrative vector and removing both the resistance maker and integrase genes by Xer site-specific recombination in one step. The corresponding plasmid vectors developed in this study could be very convenient in constructing other selectable marker-free, more stable reporter mycobacteria with diverse applications.

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

    Science.gov (United States)

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

    2017-06-20

    Complexities associated with drinking water plumbing systems can result in undesirable interactions among plumbing components that undermine engineering controls for opportunistic pathogens (OPs). In this study, we examine the effects of plumbing system materials and two commonly applied disinfectants, copper and chloramines, on water chemistry and the growth of Legionella and mycobacteria across a transect of bench- and pilot-scale hot water experiments carried out with the same municipal water supply. We discovered that copper released from corrosion of plumbing materials can initiate evolution of >1100 times more hydrogen (H 2 ) from water heater sacrificial anode rods than does presence of copper dosed as soluble cupric ions. H 2 is a favorable electron donor for autotrophs and causes fixation of organic carbon that could serve as a nutrient for OPs. Dosed cupric ions acted as a disinfectant in stratified stagnant pipes, inhibiting culturable Legionella and biofilm formation, but promoted Legionella growth in pipes subject to convective mixing. This difference was presumably due to continuous delivery of nutrients to biofilm on the pipes under convective mixing conditions. Chloramines eliminated culturable Legionella and prevented L. pneumophila from recolonizing biofilms, but M. avium gene numbers increased by 0.14-0.76 logs in the bulk water and were unaffected in the biofilm. This study provides practical confirmation of past discrepancies in the literature regarding the variable effects of copper on Legionella growth, and confirms prior reports of trade-offs between Legionella and mycobacteria if chloramines are applied as secondary disinfectant residual.

  17. Blending genomes: distributive conjugal transfer in mycobacteria, a sexier form of HGT.

    Science.gov (United States)

    Gray, Todd A; Derbyshire, Keith M

    2018-04-18

    This review discusses a novel form of horizontal gene transfer (HGT) found in mycobacteria called Distributive Conjugal Transfer (DCT). While satisfying the criteria for conjugation, DCT occurs by a mechanism so distinct from oriT-mediated conjugation that it could be considered a fourth category of HGT. DCT involves the transfer of chromosomal DNA between mycobacteria and, most significantly, generates transconjugants with mosaic genomes of the parental strains. Multiple segments of donor chromosomal DNA can be co-transferred regardless of their location or the genetic selection and, as a result, the transconjugant genome contains many donor-derived segments; hence the name DCT. This distinguishing feature of DCT separates it from the other known mechanisms of HGT, which generally result in the introduction of a single, defined segment of DNA into the recipient chromosome (Fig. ). Moreover, these mosaic progeny are generated from a single conjugal event, which provides enormous capacity for rapid adaptation and evolution, again distinguishing it from the three classical modes of HGT. Unsurprisingly, the unusual mosaic products of DCT are generated by a conjugal mechanism that is also unusual. Here, we will describe the unique features of DCT and contrast those to other mechanisms of HGT, both from a mechanistic and an evolutionary perspective. Our focus will be on transfer of chromosomal DNA, as opposed to plasmid mobilization, because DCT mediates transfer of chromosomal DNA and is a chromosomally encoded process. © 2018 John Wiley & Sons Ltd.

  18. Rapidly growing non-tuberculous mycobacteria infection of prosthetic knee joints: A report of two cases.

    Science.gov (United States)

    Kim, Manyoung; Ha, Chul-Won; Jang, Jae Won; Park, Yong-Beom

    2017-08-01

    Non-tuberculous mycobacteria (NTM) cause prosthetic knee joint infections in rare cases. Infections with rapidly growing non-tuberculous mycobacteria (RGNTM) are difficult to treat due to their aggressive clinical behavior and resistance to antibiotics. Infections of a prosthetic knee joint by RGNTM have rarely been reported. A standard of treatment has not yet been established because of the rarity of the condition. In previous reports, diagnoses of RGNTM infections in prosthetic knee joints took a long time to reach because the condition was not suspected, due to its rarity. In addition, it is difficult to identify RGNTM in the lab because special identification tests are needed. In previous reports, after treatment for RGNTM prosthetic infections, knee prostheses could not be re-implanted in all cases but one, resulting in arthrodesis or resection arthroplasty; this was most likely due to the aggressiveness of these organisms. In the present report, two cases of prosthetic knee joint infection caused by RGNTM (Mycobacterium abscessus) are described that were successfully treated, and in which prosthetic joints were finally reimplanted in two-stage revision surgery. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Optimal DNA Isolation Method for Detection of Nontuberculous Mycobacteria by Polymerase Chain Reaction.

    Science.gov (United States)

    Mohammadi, Samira; Esfahani, Bahram Nasr; Moghim, Sharareh; Mirhendi, Hossein; Zaniani, Fatemeh Riyahi; Safaei, Hajieh Ghasemian; Fazeli, Hossein; Salehi, Mahshid

    2017-01-01

    Nontuberculous mycobacteria (NTM) are a group of opportunistic pathogens and these are widely dispersed in water and soil resources. Identification of mycobacteria isolates by conventional methods including biochemical tests, growth rates, colony pigmentation, and presence of acid-fast bacilli is widely used, but these methods are time-consuming, labor-intensive, and may sometimes remain inconclusive. The DNA was extracted from NTM cultures using CTAB, Chelex, Chelex + Nonidet P-40, FTA ® Elute card, and boiling The quantity and quality of the DNA extracted via these methods were determined using UV-photometer at 260 and 280 nm, and polymerase chain reaction (PCR) amplification of the heat-shock protein 65 gene with serially diluted DNA samples. The CTAB method showed more positive results at 1:10-1:100,000 at which the DNA amount was substantial. With the Chelex method of DNA extraction, PCR amplification was detected at 1:10 and 1:1000 dilutions. According to the electrophoresis results, the CTAB and Chelex DNA extraction methods were more successful in comparison with the others as regard producing suitable concentrations of DNA with the minimum use of PCR inhibitor.

  20. Optimal DNA Isolation Method for Detection of Nontuberculous Mycobacteria by Polymerase Chain Reaction

    Directory of Open Access Journals (Sweden)

    Samira Mohammadi

    2017-01-01

    Full Text Available Background: Nontuberculous mycobacteria (NTM are a group of opportunistic pathogens and these are widely dispersed in water and soil resources. Identification of mycobacteria isolates by conventional methods including biochemical tests, growth rates, colony pigmentation, and presence of acid-fast bacilli is widely used, but these methods are time-consuming, labor-intensive, and may sometimes remain inconclusive. Materials and Methods: The DNA was extracted from NTM cultures using CTAB, Chelex, Chelex + Nonidet P-40, FTA® Elute card, and boiling The quantity and quality of the DNA extracted via these methods were determined using UV-photometer at 260 and 280 nm, and polymerase chain reaction (PCR amplification of the heat-shock protein 65 gene with serially diluted DNA samples. Results: The CTAB method showed more positive results at 1:10–1:100,000 at which the DNA amount was substantial. With the Chelex method of DNA extraction, PCR amplification was detected at 1:10 and 1:1000 dilutions. Conclusions: According to the electrophoresis results, the CTAB and Chelex DNA extraction methods were more successful in comparison with the others as regard producing suitable concentrations of DNA with the minimum use of PCR inhibitor.

  1. Synthesis of avibactam derivatives and activity on β-lactamases and peptidoglycan biosynthesis enzymes of mycobacteria.

    Science.gov (United States)

    Edoo, Zainab; Iannazzo, Laura; Compain, Fabrice; Li de la Sierra Gallay, Inès; van Tilbeurgh, Herman; Fonvielle, Matthieu; Bouchet, Flavie; Le Run, Eva; Mainardi, Jean-Luc; Arthur, Michel; Ethève-Quelquejeu, Mélanie; Hugonnet, Jean-Emmanuel

    2018-03-30

    There is a renewed interest for β-lactams for treating infections due to Mycobacterium tuberculosis and M. abscessus since their β-lactamases are inhibited by classical (clavulanate) or new generation (avibactam) inhibitors, respectively. Here, we report access to an azido derivative of the diazabicyclooctane (DBO) scaffold of avibactam for functionalization by the Huisgen-Sharpless cycloaddition reaction. The amoxicillin-DBO combinations were active indicating that the triazole ring is compatible with drug penetration (minimal inhibitory concentration of 16 µg/ml for both species). Mechanistically, β-lactamase inhibition was not sufficient to account for the potentiation of amoxicillin by DBOs. Thus, we investigated the latter compounds as inhibitors of L,D-transpeptidases (LDTs), which are the main peptidoglycan polymerases in mycobacteria. The DBOs acted as slow-binding inhibitors of LDTs by S-carbamoylation indicating that optimization of DBOs for LDT inhibition is an attractive strategy to obtain drugs selectively active on mycobacteria. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  2. Antibiotic management of lung infections in cystic fibrosis. II. Nontuberculous mycobacteria, anaerobic bacteria, and fungi.

    Science.gov (United States)

    Chmiel, James F; Aksamit, Timothy R; Chotirmall, Sanjay H; Dasenbrook, Elliott C; Elborn, J Stuart; LiPuma, John J; Ranganathan, Sarath C; Waters, Valerie J; Ratjen, Felix A

    2014-10-01

    Airway infections are a key component of cystic fibrosis (CF) lung disease. Whereas the approach to common pathogens such as Pseudomonas aeruginosa is guided by a significant body of evidence, other infections often pose a considerable challenge to treating physicians. In Part I of this series on the antibiotic management of difficult lung infections, we discussed bacterial organisms including methicillin-resistant Staphylococcus aureus, gram-negative bacterial infections, and treatment of multiple bacterial pathogens. Here, we summarize the approach to infections with nontuberculous mycobacteria, anaerobic bacteria, and fungi. Nontuberculous mycobacteria can significantly impact the course of lung disease in patients with CF, but differentiation between colonization and infection is difficult clinically as coinfection with other micro-organisms is common. Treatment consists of different classes of antibiotics, varies in intensity, and is best guided by a team of specialized clinicians and microbiologists. The ability of anaerobic bacteria to contribute to CF lung disease is less clear, even though clinical relevance has been reported in individual patients. Anaerobes detected in CF sputum are often resistant to multiple drugs, and treatment has not yet been shown to positively affect patient outcome. Fungi have gained significant interest as potential CF pathogens. Although the role of Candida is largely unclear, there is mounting evidence that Scedosporium species and Aspergillus fumigatus, beyond the classical presentation of allergic bronchopulmonary aspergillosis, can be relevant in patients with CF and treatment should be considered. At present, however there remains limited information on how best to select patients who could benefit from antifungal therapy.

  3. Hypersensitivity of hypoxia grown Mycobacterium smegmatis to DNA damaging agents: implications of the DNA repair deficiencies in attenuation of mycobacteria.

    Science.gov (United States)

    Rex, Kervin; Kurthkoti, Krishna; Varshney, Umesh

    2013-10-01

    Mycobacteria are an important group of pathogenic bacteria. We generated a series of DNA repair deficient strains of Mycobacterium smegmatis, a model organism, to understand the importance of various DNA repair proteins (UvrB, Ung, UdgB, MutY and Fpg) in survival of the pathogenic strains. Here, we compared tolerance of the M. smegmatis strains to genotoxic stress (ROS and RNI) under aerobic, hypoxic and recovery conditions of growth by monitoring their survival. We show an increased susceptibility of mycobacteria to genotoxic stress under hypoxia. UvrB deficiency led to high susceptibility of M. smegmatis to the DNA damaging agents. Ung was second in importance in strains with single deficiencies. Interestingly, we observed that while deficiency of UdgB had only a minor impact on the strain's susceptibility, its combination with Ung deficiency resulted in severe consequences on the strain's survival under genotoxic stress suggesting a strong interdependence of different DNA repair pathways in safeguarding genomic integrity. Our observations reinforce the possibility of targeting DNA repair processes in mycobacteria for therapeutic intervention during active growth and latency phase of the pathogen. High susceptibility of the UvrB, or the Ung/UdgB deficient strains to genotoxic stress may be exploited in generation of attenuated strains of mycobacteria. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  4. Isolation of nontuberculous mycobacteria from soil using Middlebrook 7H10 agar with increased malachite green concentration.

    Science.gov (United States)

    Hu, Yuli; Yu, Xinglong; Zhao, Dun; Li, Runcheng; Liu, Yang; Ge, Meng; Hu, Huican

    2017-12-01

    Environmental exposure is considered to be responsible for nontuberculous mycobacterial infections in humans. To facilitate the isolation of mycobacteria from soil, Middlebrook 7H10 agar was optimized as an enhanced selective medium by increasing the concentration of malachite green. A series of modified Middlebrook 7H10 agar media with malachite green concentrations ranging from 2.5 to 2500 mg/L was evaluated using 20 soil samples decontaminated with 3% sodium dodecyl sulfate plus 2% NaOH for 30 min. Among these modified Middlebrook 7H10 media, the medium with malachite green at a concentration of 250 mg/L, i.e., at the same concentration as in Löwenstein-Jensen medium, was the most effective in terms of the number of plates with mycobacterial growth. This medium was further evaluated with 116 soil samples. The results showed that 87.1% (101/116) of the samples produced mycobacterial growth, and 15 samples (12.9%) produced no mycobacterial growth. Of the plates inoculated with the soil samples, each in duplicate, 5.2% (12/232) showed late contamination. In total, 19 mycobacterial species were isolated, including seven (36.8%) rapidly growing mycobacteria and 12 (63.2%) slowly growing mycobacteria. Our results demonstrate that the modified Middlebrook 7H10 agar with 250 mg/L malachite green is useful for the primary isolation of nontuberculous mycobacteria from soil.

  5. Comparing the harmful effects of nontuberculous mycobacteria and Gram negative bacteria on lung function in patients with cystic fibrosis

    DEFF Research Database (Denmark)

    Qvist, Tavs; Taylor-Robinson, David; Waldmann, Elisabeth

    2015-01-01

    BACKGROUND: To better understand the relative effects of infection with nontuberculous mycobacteria and Gram negative bacteria on lung function decline in cystic fibrosis, we assessed the impact of each infection in a Danish setting. METHODS: Longitudinal registry study of 432 patients with cystic...

  6. Genomic evaluations with many more genotypes

    Directory of Open Access Journals (Sweden)

    Wiggans George R

    2011-03-01

    Full Text Available Abstract Background Genomic evaluations in Holstein dairy cattle have quickly become more reliable over the last two years in many countries as more animals have been genotyped for 50,000 markers. Evaluations can also include animals genotyped with more or fewer markers using new tools such as the 777,000 or 2,900 marker chips recently introduced for cattle. Gains from more markers can be predicted using simulation, whereas strategies to use fewer markers have been compared using subsets of actual genotypes. The overall cost of selection is reduced by genotyping most animals at less than the highest density and imputing their missing genotypes using haplotypes. Algorithms to combine different densities need to be efficient because numbers of genotyped animals and markers may continue to grow quickly. Methods Genotypes for 500,000 markers were simulated for the 33,414 Holsteins that had 50,000 marker genotypes in the North American database. Another 86,465 non-genotyped ancestors were included in the pedigree file, and linkage disequilibrium was generated directly in the base population. Mixed density datasets were created by keeping 50,000 (every tenth of the markers for most animals. Missing genotypes were imputed using a combination of population haplotyping and pedigree haplotyping. Reliabilities of genomic evaluations using linear and nonlinear methods were compared. Results Differing marker sets for a large population were combined with just a few hours of computation. About 95% of paternal alleles were determined correctly, and > 95% of missing genotypes were called correctly. Reliability of breeding values was already high (84.4% with 50,000 simulated markers. The gain in reliability from increasing the number of markers to 500,000 was only 1.6%, but more than half of that gain resulted from genotyping just 1,406 young bulls at higher density. Linear genomic evaluations had reliabilities 1.5% lower than the nonlinear evaluations with 50

  7. Prevalence of hepatitis C virus-resistant association substitutions to direct-acting antiviral agents in treatment-naïve hepatitis C genotype 1b-infected patients in western China

    Directory of Open Access Journals (Sweden)

    Li Z

    2017-10-01

    Full Text Available Zhao Li,* Zhi-wei Chen,* Hu Li, Hong Ren, Peng Hu Department of Infectious Diseases, Institute for Viral Hepatitis, Key Laboratory of Molecular Biology for Infectious Diseases, Chinese Ministry of Education, Second Affiliated Hospital of Chongqing Medical University, Chongqing, China *These authors contributed equally to this work. Background: Direct-acting antivirals (DAAs against hepatitis C virus (HCV are potent and highly efficacious. However, resistance-associated substitutions (RASs relevant to DAAs can impair treatment effectiveness even at baseline. Moreover, the prevalence of baseline RASs in HCV genotype 1b-infected patients in western China is still unclear. Materials and methods: Direct sequencing of the HCV NS3, NS5A, and NS5B regions was performed in baseline serum samples of 70 DAAs treatment-naïve HCV 1b-infected patients in western China. The sequences were analyzed with MEGA version 5.05 software. Evolutionary patterns of RASs and amino-acid covariance patterns in the NS3, NS5A, and NS5B genes were analyzed by MEGA and Cytoscape (version 3.2.1, respectively.Results: The presence of at least one RAS in the NS3 region (C16S, T54S, Q80R/L, A87T, R117H, S122G, V132I, V170I was observed in 85.48% (53 of 62 of patients, RASs in the NS5A region (L28M, R30Q, Q54H, P58S/T, Q62H/R, Y93H were observed in 42.42% (28 of 66 of patients, and RASs in the NS5B region (N142S, A300T, C316N, A338V, S365A, L392I, M414L, I424V, A442T, V499A, S556G were observed in 100% (44 of 44 of patients. Evolutionary patterns of RASs and amino-acid covariance patterns for the NS3, NS5A, and NS5B genes are reported.Conclusion: The prevalence of RASs relevant to DAAs detected in the NS3, NS5A, and NS5B regions of HCV 1b from DAA treatment-naïve patients is high. Therefore, more attention should be paid to RASs associated with DAAs in the upcoming DAA-treatment era in China. Keywords: hepatitis C virus, unstructured proteins, resistance

  8. Hepatitis C Virus: Viral Quasispecies and Genotypes

    Directory of Open Access Journals (Sweden)

    Kyoko Tsukiyama-Kohara

    2017-12-01

    Full Text Available Hepatitis C virus (HCV mainly replicates in the cytoplasm, where it easily establishes persistent infection, resulting in chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. Due to its high rate of mutation, HCV forms viral quasispecies, categorized based on the highly variable regions in the envelope protein and nonstructural 5A protein. HCV possesses seven major genotypes, among which genotype 1 is the most prevalent globally. The distribution of HCV genotypes varies based on geography, and each genotype has a different sensitivity to interferon treatment. Recently-developed direct-acting antivirals (DAAs, which target viral proteases or polymerases, mediate drastically better antiviral effects than previous therapeutics. Although treatment with DAAs has led to the development of drug-resistant HCV mutants, the most recently approved DAAs show improved pan-genomic activity, with a higher barrier to viral resistance.

  9. Hepatitis C Virus: Viral Quasispecies and Genotypes.

    Science.gov (United States)

    Tsukiyama-Kohara, Kyoko; Kohara, Michinori

    2017-12-22

    Hepatitis C virus (HCV) mainly replicates in the cytoplasm, where it easily establishes persistent infection, resulting in chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. Due to its high rate of mutation, HCV forms viral quasispecies, categorized based on the highly variable regions in the envelope protein and nonstructural 5A protein. HCV possesses seven major genotypes, among which genotype 1 is the most prevalent globally. The distribution of HCV genotypes varies based on geography, and each genotype has a different sensitivity to interferon treatment. Recently-developed direct-acting antivirals (DAAs), which target viral proteases or polymerases, mediate drastically better antiviral effects than previous therapeutics. Although treatment with DAAs has led to the development of drug-resistant HCV mutants, the most recently approved DAAs show improved pan-genomic activity, with a higher barrier to viral resistance.

  10. Frequency of Natural Resistance within NS5a Replication Complex Domain in Hepatitis C Genotypes 1a, 1b: Possible Implication of Subtype-Specific Resistance Selection in Multiple Direct Acting Antivirals Drugs Combination Treatment

    Directory of Open Access Journals (Sweden)

    Sabrina Bagaglio

    2016-03-01

    Full Text Available Different HCV subtypes may naturally harbor different resistance selection to anti-NS5a inhibitors. 2761 sequences retrieved from the Los Alamos HCV database were analyzed in the NS5a domain 1, the target of NS5a inhibitors. The NS5a resistance-associated polymorphisms (RAPs were more frequently detected in HCV G1b compared to G1a. The prevalence of polymorphisms associated with cross-resistance to compounds in clinical use (daclatasvir, DCV, ledipasvir, LDV, ombitasvir, and OMV or scheduled to come into clinical use in the near future (IDX719, elbasvir, and ELV was higher in G1b compared to G1a (37/1552 (2.4% in 1b sequences and 15/1209 (1.2% in 1a isolates, p = 0.040. Interestingly, on the basis of the genotype-specific resistance pattern, 95 (6.1% G1b sequences had L31M RAP to DCV/IDX719, while 6 sequences of G1a (0.5% harbored L31M RAP, conferring resistance to DCV/LDV/IDX719/ELV (p < 0.0001. Finally, 28 (2.3% G1a and none of G1b isolates harbored M28V RAP to OMV (p < 0.0001. In conclusion, the pattern of subtype-specific resistance selection in the naturally occurring strains may guide the treatment option in association with direct acting antivirals (DAAs targeting different regions, particularly in patients that are difficult to cure, such as those with advanced liver disease or individuals who have failed previous DAAs.

  11. APOE Genotyping, Cardiovascular Disease

    Science.gov (United States)

    ... Resources For Health Professionals Subscribe Search APOE Genotyping, Cardiovascular Disease Send Us Your Feedback Choose Topic At a ... help understand the role of genetic factors in cardiovascular disease . However, the testing is sometimes used in clinical ...

  12. Radiosensitivity of fingermillet genotypes

    Energy Technology Data Exchange (ETDEWEB)

    Raveendran, T S; Nagarajan, C; Appadurai, R; Prasad, M N; Sundaresan, N [Tamil Nadu Agricultural Univ., Coimbatore (India)

    1984-07-01

    Varietal differences in radiosensitivity were observed in a study involving 4 genotypes of fingermillet (Eleusine coracana (Linn.) Gaertn.) subjected to gamma-irradiation. Harder seeds were found to tolerate a higher dose of the mutagen.

  13. Baby bottle steam sterilizers for disinfecting home nebulizers inoculated with non-tuberculous mycobacteria.

    Science.gov (United States)

    Towle, D; Callan, D A; Lamprea, C; Murray, T S

    2016-03-01

    Non-tuberculous mycobacteria (NTMb), present in environmental water sources, can contribute to respiratory infection in patients with chronic pulmonary disease. Contaminated nebulizers are a potential source of respiratory infection. Treatment with baby bottle steam sterilizers disinfects home nebulizers inoculated with bacterial pathogens but whether this method works for disinfection of NTMb is unclear. Baby bottle steam sterilization was compared with vigorous water washing for disinfecting home nebulizers inoculated with NTMb mixed with cystic fibrosis sputum. No NTMb was recovered from any nebulizers after steam treatment whereas viable NTMb grew after water washing, demonstrating that steam sterilization effectively disinfects NTMb-inoculated nebulizers. Copyright © 2015 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.

  14. COMPARATIVE ANALYSIS OF METHODS FOR IDENTIFICATION OF NONTUBERCULOUS MYCOBACTERIA ISOLATED FROM CLINICAL MATERIAL

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    A. V. Lyamin

    2017-01-01

    -ToF spectrometry, DNA hybridization were not determined to species. 17 (21.8% of microbial strains which have been identified using the method of DNA hybridization, identified by spectrometry, including slow-growing microorganisms, non-mycobacteria strains seven (9.0%: Gordonia rubriperticta, Nocardia forcinica, Tsukumurella spp., Rhodotorula mucilaginosa. Accurate species identification NTMB is fundamental to determine the tactics of treatment of patients with mycobacteriosis. Due to this rather limited possibility of identification of non-tuberculous mycobacteria, using a DNA-hybridization method is inadequate to date. The introduction of new techniques, such as MALDI-ToF spectrometry, can identify a greater number of species of nontuberculous mycobacteria, as well as other types of slow-growing microorganisms having similarities with mycobacteria on cultural and morphological properties, which significantly increases the diagnostic capabilities of laboratories.

  15. Rapid detection and identification of pathogenic mycobacteria by combining radiometric and nucleic acid probe methods

    International Nuclear Information System (INIS)

    Ellner, P.D.; Kiehn, T.E.; Cammarata, R.; Hosmer, M.

    1988-01-01

    The combination of radiometric methodology (BACTEC 12B) and probe technology for recovery and identification of mycobacteria was studied in two large hospital laboratories. The sediment from vials with positive growth indices was tested with DNA probes specific for Mycobacterium tuberculosis, Mycobacterium avium, and Mycobacterium intracellulare. The sensitivity of the radiometric method and the specificity of the probes resulted in a marked reduction in the time to the final report. Biochemical testing could be eliminated on isolates giving a positive reaction with one of the probes. Some 176 isolates of M. tuberculosis, 110 of M. avium, and 5 of M. intracellulare were recovered. Two-thirds of these isolates were detected and identified within 2 weeks of inoculation and the remainder was detected by 4 weeks, a reduction of 5 to 7 weeks to the final report

  16. Are we overlooking infections owing to non-tuberculous mycobacteria during routine conventional laboratory investigations?

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    Kushal Garima

    2012-01-01

    Full Text Available A large number of potentially pathogenic non-tuberculous mycobacteria (NTM encountered in the clinical laboratory makes it necessary to identify their species to ensure appropriate treatment. However, labor-intensive conventional methods of speciation are not used in every laboratory, and hence NTM infections are often ignored. Polymerase chain reaction (PCR restriction analysis (PRA was applied in this study for early identification and speciation of mycobacterial species on 306 cultures of acid-fast bacilli isolated from patients suspected of suffering from tuberculosis. Mycobacterium tuberculosis was identified in 85.6% of the isolates. The NTM isolated most commonly was Mycobacterium kansasii/gastri group (3.5%, followed by Mycobacterium fortuitum (3.2%. Four of the M. fortuitum were grown from cultures obtained on the same day, but from samples from different patients and were probably laboratory contaminants. Mycobacterium intracellulare and Mycobacterium avium were identified in 2.94% and 2.28% of the isolates, respectively. Three isolates of M. avium and two isolates of M. intracellulare were obtained in repeated cultures from sputum samples of the same patients and were thus pathogenic. A single isolate of Mycobacterium abscessus was obtained from a breast abscess. A rare pathogen Mycobacterium phocaicum was isolated from one patient with epididymitis. However, whether it was the causative agent of epididymitis in this patient remains doubtful. The results of this study highlight the importance of speciation of mycobacteria for appropriate diagnosis and the importance of including molecular assays to augment conventional methods of diagnosis of mycobacterial diseases for rapid identification of NTM so that these potential pathogens are not overlooked in routine diagnostic procedures.

  17. Sulfate Reducing Bacteria and Mycobacteria Dominate the Biofilm Communities in a Chloraminated Drinking Water Distribution System.

    Science.gov (United States)

    Gomez-Smith, C Kimloi; LaPara, Timothy M; Hozalski, Raymond M

    2015-07-21

    The quantity and composition of bacterial biofilms growing on 10 water mains from a full-scale chloraminated water distribution system were analyzed using real-time PCR targeting the 16S rRNA gene and next-generation, high-throughput Illumina sequencing. Water mains with corrosion tubercles supported the greatest amount of bacterial biomass (n = 25; geometric mean = 2.5 × 10(7) copies cm(-2)), which was significantly higher (P = 0.04) than cement-lined cast-iron mains (n = 6; geometric mean = 2.0 × 10(6) copies cm(-2)). Despite spatial variation of community composition and bacterial abundance in water main biofilms, the communities on the interior main surfaces were surprisingly similar, containing a core group of operational taxonomic units (OTUs) assigned to only 17 different genera. Bacteria from the genus Mycobacterium dominated all communities at the main wall-bulk water interface (25-78% of the community), regardless of main age, estimated water age, main material, and the presence of corrosion products. Further sequencing of the mycobacterial heat shock protein gene (hsp65) provided species-level taxonomic resolution of mycobacteria. The two dominant Mycobacteria present, M. frederiksbergense (arithmetic mean = 85.7% of hsp65 sequences) and M. aurum (arithmetic mean = 6.5% of hsp65 sequences), are generally considered to be nonpathogenic. Two opportunistic pathogens, however, were detected at low numbers: M. hemophilum (arithmetic mean = 1.5% of hsp65 sequences) and M. abscessus (arithmetic mean = 0.006% of hsp65 sequences). Sulfate-reducing bacteria from the genus Desulfovibrio, which have been implicated in microbially influenced corrosion, dominated all communities located underneath corrosion tubercules (arithmetic mean = 67.5% of the community). This research provides novel insights into the quantity and composition of biofilms in full-scale drinking water distribution systems, which is critical for assessing the risks to public health and to the

  18. Identification of a Novel Conjugative Plasmid in Mycobacteria That Requires Both Type IV and Type VII Secretion

    KAUST Repository

    Ummels, R.; Abdallah, A. M.; Kuiper, V.; Aajoud, A.; Sparrius, M.; Naeem, R.; Spaink, H. P.; van Soolingen, D.; Pain, Arnab; Bitter, W.

    2014-01-01

    Conjugative plasmids play an important role in horizontal gene transfer between different bacteria and, as such, in their adaptation and evolution. This effect is most obvious in the spread of antibiotic resistance genes. Thus far, conjugation of natural plasmids has been described only rarely for mycobacterial species. In fact, it is generally accepted that M. tuberculosis does not show any recent sign of horizontal gene transfer. In this study, we describe the identification of a new widespread conjugative plasmid that can also be efficiently transferred to M. tuberculosis. This plasmid therefore poses both a threat and an opportunity. The threat is that, through the acquisition of antibiotic resistance markers, this plasmid could start a rapid spread of antibiotic resistance genes between pathogenic mycobacteria. The opportunity is that we could use this plasmid to generate new tools for the efficient introduction of foreign DNA in slow-growing mycobacteria.

  19. Azurophil granule proteins constitute the major mycobactericidal proteins in human neutrophils and enhance the killing of mycobacteria in macrophages

    DEFF Research Database (Denmark)

    Jena, Prajna; Mohanty, Soumitra; Mohanty, Tirthankar

    2012-01-01

    Pathogenic mycobacteria reside in, and are in turn controlled by, macrophages. However, emerging data suggest that neutrophils also play a critical role in innate immunity to tuberculosis, presumably by their different antibacterial granule proteins. In this study, we purified neutrophil azurophil...... and specific granules and systematically analyzed the antimycobacterial activity of some purified azurophil and specific granule proteins against M. smegmatis, M. bovis-BCG and M. tuberculosis H37Rv. Using gel overlay and colony forming unit assays we showed that the defensin-depleted azurophil granule...... proteins (AZP) were more active against mycobacteria compared to other granule proteins and cytosolic proteins. The proteins showing antimycobacterial activity were identified by MALDI-TOF mass spectrometry. Electron microscopic studies demonstrate that the AZP disintegrate bacterial cell membrane...

  20. Pretreatment of clinical specimens with sodium dodecyl (lauryl) sulfate is not suitable for the mycobacteria growth indicator tube cultivation method.

    OpenAIRE

    Pfyffer, G E; Welscher, H M; Kissling, P

    1997-01-01

    When using the Mycobacteria Growth Indicator Tube (MGIT), pretreatment of clinical specimens with N-acetyl-L-cysteine-NaOH is recommended by the manufacturer. Processing of clinical specimens (n = 1,000) with sodium dodecyl (lauryl) sulfate-NaOH resulted in both poor recovery and delayed mean time to detection of acid-fast bacilli. Values were comparable to those obtained on solid media.

  1. Developing whole mycobacteria cell vaccines for tuberculosis: Workshop proceedings, Max Planck Institute for Infection Biology, Berlin, Germany, July 9, 2014.

    Science.gov (United States)

    2015-06-12

    On July 9, 2014, Aeras and the Max Planck Institute for Infection Biology convened a workshop entitled "Whole Mycobacteria Cell Vaccines for Tuberculosis" at the Max Planck Institute for Infection Biology on the grounds of the Charité Hospital in Berlin, Germany, close to the laboratory where, in 1882, Robert Koch first identified Mycobacterium tuberculosis (Mtb) as the pathogen responsible for tuberculosis (TB). The purpose of the meeting was to discuss progress in the development of TB vaccines based on whole mycobacteria cells. Live whole cell TB vaccines discussed at this meeting were derived from Mtb itself, from Bacille Calmette-Guérin (BCG), the only licensed vaccine against TB, which was genetically modified to reduce pathogenicity and increase immunogenicity, or from commensal non-tuberculous mycobacteria. Inactivated whole cell TB and non-tuberculous mycobacterial vaccines, intended as immunotherapy or as safer immunization alternatives for HIV+ individuals, also were discussed. Workshop participants agreed that TB vaccine development is significantly hampered by imperfect animal models, unknown immune correlates of protection and the absence of a human challenge model. Although a more effective TB vaccine is needed to replace or enhance the limited effectiveness of BCG in all age groups, members of the workshop concurred that an effective vaccine would have the greatest impact on TB control when administered to adolescents and adults, and that use of whole mycobacteria cells as TB vaccine candidates merits greater support, particularly given the limited understanding of the specific Mtb antigens necessary to generate an immune response capable of preventing Mtb infection and/or disease. Copyright © 2015. Published by Elsevier Ltd.. All rights reserved.

  2. High recurrence rate of lymphadenitis due to nontuberculous mycobacteria and its association with concurrent Salmonella infection in Taiwan.

    Science.gov (United States)

    Hsiao, Cheng-Hsiang; Lai, Chih-Cheng; Hsueh, Po-Ren

    2014-06-01

    The objective of this study is to investigate the clinical characteristics of lymphadenitis due to nontuberculous mycobacteria (NTM) in Taiwan. We retrospectively reviewed the medical records of all patients who presented to the National Taiwan University Hospital with culture-positive NTM lymphadenitis during the period 2000-2010. Patients with concurrent extranodal involvement were excluded. From 2000 to 2010, 15 patients with lymphadenitis caused by nontuberculous mycobacteria were identified. Most patients (80%, n = 12) were infected with rapidly growing mycobacteria. Mycobacterium abscessus was the most common infective species (n = 8). Recurrence of infection involving multiple organs occurred 2-7 years after the completion of treatment in 11 (73%) patients. Five (33.3%) patients had concurrent Salmonella infections (4 patients with bacteremia and 1 patient with empyema thoracis) during the course of the disease. In Taiwanese patients, we found a high recurrence rate of NTM lymphadenitis that was closely associated with Salmonella infections. We also noted that the clinical and epidemiological manifestations of NTM lymphadenitis in Taiwan differed from their manifestations in western countries. Copyright © 2012. Published by Elsevier B.V.

  3. Comparison of a radiometric method (BACTEC) and conventional culture media for recovery of mycobacteria from smear-negative specimens

    International Nuclear Information System (INIS)

    Morgan, M.A.; Horstmeier, C.D.; DeYoung, D.R.; Roberts, G.D.

    1983-01-01

    The BACTEC system and three conventional media (Middlebrook 7H10, selective Middlebrook 7H11 [S7H11], and Lowenstein-Jensen [LJ] were compared for their mean recovery times and recovery rates of mycobacteria from acid-fast, smear-negative clinical specimens. Of the 71 smear-negative, culture-positive specimens recovered from 2,165 submitted smear-negative cultures, the BACTEC system detected 71.8%, compared with 88.7% for the conventional three-medium system. When media were individually compared, BACTEC medium (Middlebrook 7H12) was more successful in recovering mycobacteria (71.8%) than was LJ (62%), Middlebrook medium 7H10 (55.9%), or Middlebrook S7H11 medium (52.1%). Middlebrook 7H11 medium containing sodium selenate was also evaluated and did not increase the recovery rate or decrease the recovery time of mycobacterial species when compared with LJ, Middlebrook 7H10, S7H11, and 7H12 media. The mean detection time for the BACTEC system was less than that by conventional methods for the seven species of mycobacteria recovered. Detection times for Mycobacterium tuberculosis on the BACTEC system and conventional cultural systems were 13.7 and 26.3 days, respectively

  4. Increase in nontuberculous mycobacteria isolated in Shanghai, China: results from a population-based study.

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    Jie Wu

    Full Text Available BACKGROUND: In China, the prevalence of nontuberculous mycobacteria (NTM in isolates from mycobacterial culture-positive patients with pulmonary tuberculosis (TB is largely unknown. METHODS: We used conventional biochemical and 16S rRNA gene sequencing to identify species of mycobacteria in specimens from patients suspected of having TB. Drug-susceptibility testing was performed on NTM isolates using the proportion method. We also determined the independent risk factors associated with infection with NTM compared with infection with Mycobacterium tuberculosis. RESULTS: The overall rate of NTM isolated from mycobacterial culture-positive patients was 5.9% in this population, with a significantly increasing trend from 3.0% in 2008 to 8.5% in 2012 (P for trend <0.001. The organism most frequently identified was M. kansasii (45.0%, followed by M. intracellulare (20.8% and M. chelonae/abscessus (14.9%. The overall proportion of isolates resistant to the four first-line anti-TB agents were 64.6% for isoniazid, 77.6% for streptomycin, 63.3% for rifampicin and 75.1% for ethambutol. The risk factors most often associated with NTM infection were older age (P for trend <0.001, being a resident of Shanghai (adjusted odds ratio [aOR], 1.48; 95% CI, 1.10-2.00, having been treated for tuberculosis (aOR, 1.64; 95% CI, 1.18-2.29, having a cavity on chest X-ray (aOR, 1.51; 95% CI, 1.16-1.96, and being sputum smear-negative (aOR, 1.59; 95% CI, 1.16-2.18. CONCLUSIONS: The prevalence of NTM isolated in Shanghai increased between 2008 and 2012, thus clinicians should consider NTM as a possible cause of TB-like disease. Accurate species identification is imperative so that proper treatment can be administered for diseases caused by the diversity of NTM species.

  5. Differential effects of Radix Paeoniae Rubra (Chishao on cytokine and chemokine expression inducible by mycobacteria

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    Li James

    2011-03-01

    Full Text Available Abstract Background Upon initial infection with mycobacteria, macrophages secrete multiple cytokines and chemokines, including interleukin-6 (IL-6, IL-8 and tumor necrosis factor-α (TNF-α, to mediate host immune responses against the pathogen. Mycobacteria also induce the production of IL-10 via PKR activation in primary human monocytes and macrophages. As an anti-inflammatory cytokine, over-expression of IL-10 may contribute to mycobacterial evasion of the host immunity. Radix Paeoniae Rubra (RPR, Chishao, a Chinese medicinal herb with potentials of anti-inflammatory, hepatoprotective and neuroprotective effects, is used to treat tuberculosis. This study investigates the immunoregulatory effects of RPR on primary human blood macrophages (PBMac during mycobacterial infection. Methods The interaction of Bacillus Calmette-Guerin (BCG with PBMac was used as an experimental model. A series of procedures involving solvent extraction and fractionation were used to isolate bioactive constituents in RPR. RPR-EA-S1, a fraction with potent immunoregulatory effects was obtained with a bioactivity guided fractionation scheme. PBMac were treated with crude RPR extracts or RPR-EA-S1 before BCG stimulation. The expression levels of IL-6, IL-8, IL-10 and TNF-α were measured by qPCR and ELISA. Western blotting was used to determine the effects of RPR-EA-S1 on signaling kinases and transcriptional factors in the BCG-activated PBMac. Results In BCG-stimulated macrophages, crude RPR extracts and fraction RPR-EA-S1 specifically inhibited IL-10 production while enhanced IL-8 expression at both mRNA and protein levels without affecting the expressions of IL-6 and TNF-α. Inhibition of BCG-induced IL-10 expression by RPR-EA-S1 occurred in a dose- and time-dependent manner. RPR-EA-S1 did not affect the phosphorylation of cellular protein kinases including MAPK, Akt and GSK3β. Instead, it suppressed the degradation of IκBα in the cytoplasm and inhibited the

  6. Capacity of lung stroma to educate dendritic cells inhibiting mycobacteria-specific T-cell response depends upon genetic susceptibility to tuberculosis.

    Science.gov (United States)

    Kapina, Marina A; Rubakova, Elvira I; Majorov, Konstantin B; Logunova, Nadezhda N; Apt, Alexander S

    2013-01-01

    The balance between activation and inhibition of local immune responses in affected tissues during prolonged chronic infections is important for host protection. There is ample evidence that regulatory, tolerogenic dendritic cells (DC) are developed and present in tissues and inhibit overwhelming inflammatory reactions. Also, it was firmly established that stromal microenvironment of many organs is able to induce development of immature regulatory DC (DCreg), an essential element of a general immune regulatory network. However, direct experimental data demonstrating inhibition of immune responses by stroma-instructed immature DCreg in infectious models are scarce, and virtually nothing is known about functioning of this axis of immunity during tuberculosis (TB) infection. In this study, we demonstrate that lung stromal cells are capable of supporting the development in culture of immature CD11b(+)CD11c(low)CD103(-) DCreg from lineage-negative (lin(-)) bone marrow precursors. DCreg developed on lung stroma isolated from mice of genetically TB-hyper-susceptible I/St and relatively resistant B6 inbred strains inhibited proliferative response of mycobacteria-specific CD4(+) T-cell lines a dose-dependent manner. Importantly, the inhibitory activity of B6 DCreg was substantially higher than that of I/St Dcreg. Moreover, when the donors of stromal cells were chronically infected with virulent mycobacteria, the capacity to instruct inhibitory DCreg was retained in B6, but further diminished in I/St stromal cells. DCreg-provided suppression was mediated by a few soluble mediators, including PGE2, NO and IL-10. The content of CD4(+)Foxp3(+) Treg cells in the mediastinal, lung-draining lymph nodes at the advanced stages of chronic infection did not change in I/St, but increased 2-fold in B6 mice, and lung pathology was much more pronounced in the former mice. Taken together, these data provide genetic evidence that the capacity to maintain populations of regulatory cells

  7. Capacity of lung stroma to educate dendritic cells inhibiting mycobacteria-specific T-cell response depends upon genetic susceptibility to tuberculosis.

    Directory of Open Access Journals (Sweden)

    Marina A Kapina

    Full Text Available The balance between activation and inhibition of local immune responses in affected tissues during prolonged chronic infections is important for host protection. There is ample evidence that regulatory, tolerogenic dendritic cells (DC are developed and present in tissues and inhibit overwhelming inflammatory reactions. Also, it was firmly established that stromal microenvironment of many organs is able to induce development of immature regulatory DC (DCreg, an essential element of a general immune regulatory network. However, direct experimental data demonstrating inhibition of immune responses by stroma-instructed immature DCreg in infectious models are scarce, and virtually nothing is known about functioning of this axis of immunity during tuberculosis (TB infection. In this study, we demonstrate that lung stromal cells are capable of supporting the development in culture of immature CD11b(+CD11c(lowCD103(- DCreg from lineage-negative (lin(- bone marrow precursors. DCreg developed on lung stroma isolated from mice of genetically TB-hyper-susceptible I/St and relatively resistant B6 inbred strains inhibited proliferative response of mycobacteria-specific CD4(+ T-cell lines a dose-dependent manner. Importantly, the inhibitory activity of B6 DCreg was substantially higher than that of I/St Dcreg. Moreover, when the donors of stromal cells were chronically infected with virulent mycobacteria, the capacity to instruct inhibitory DCreg was retained in B6, but further diminished in I/St stromal cells. DCreg-provided suppression was mediated by a few soluble mediators, including PGE2, NO and IL-10. The content of CD4(+Foxp3(+ Treg cells in the mediastinal, lung-draining lymph nodes at the advanced stages of chronic infection did not change in I/St, but increased 2-fold in B6 mice, and lung pathology was much more pronounced in the former mice. Taken together, these data provide genetic evidence that the capacity to maintain populations of regulatory

  8. Helicobacter pylori from Peptic Ulcer Patients in Uganda Is Highly Resistant to Clarithromycin and Fluoroquinolones: Results of the GenoType HelicoDR Test Directly Applied on Stool

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    Denish Calmax Angol

    2017-01-01

    Full Text Available Background. Around 70–90% of peptic ulcer disease (PUD is due to Helicobacter pylori and requires treatment with antimicrobials to which these bacteria are susceptible. Common H. pylori diagnostic tests do not provide drug susceptibility data. Using the GenoType HelicoDR PCR test designed for gastric biopsies for simultaneous detection of H. pylori and its resistance to clarithromycin (CLA/fluoroquinolones (FLQ, we present evidence for stool as an optional test specimen and also provide data on prevalence of H. pylori resistance to CLA and FLQ in Uganda. Methods. Stool from 142 symptomatic PUD patients at three hospitals in Kampala was screened for H. pylori using a rapid antigen test. The GenoType HelicoDR test was run on all H. pylori antigen positives to determine PCR positivity and resistance to CLA/FLQ. Results. Thirty-one samples (22% were H. pylori antigen positive, and 21 (68% of these were H. pylori PCR positive. Six of the 21 (29% were resistant to CLA and eight to FLQ (42%, while two gave invalid FLQ resistance results. Conclusion. Stool is a possible specimen for the GenoType HelicoDR test for rapid detection of H. pylori and drug resistance. In Uganda, Helicobacter pylori is highly resistant to CLA and FLQ.

  9. Pathogenic mycobacteria achieve cellular persistence by inhibiting the Niemann-Pick Type C disease cellular pathway [version 2; referees: 2 approved, 2 approved with reservations

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    Paul Fineran

    2017-06-01

    Full Text Available Background. Tuberculosis remains a major global health concern. The ability to prevent phagosome-lysosome fusion is a key mechanism by which intracellular mycobacteria, including Mycobacterium tuberculosis, achieve long-term persistence within host cells. The mechanisms underpinning this key intracellular pro-survival strategy remain incompletely understood. Host macrophages infected with intracellular mycobacteria share phenotypic similarities with cells taken from patients suffering from Niemann-Pick Disease Type C (NPC, a rare lysosomal storage disease in which endocytic trafficking defects and lipid accumulation within the lysosome lead to cell dysfunction and cell death. We investigated whether these shared phenotypes reflected an underlying mechanistic connection between mycobacterial intracellular persistence and the host cell pathway dysfunctional in NPC.  Methods. The induction of NPC phenotypes in macrophages from wild-type mice or obtained from healthy human donors was assessed via infection with mycobacteria and subsequent measurement of lipid levels and intracellular calcium homeostasis. The effect of NPC therapeutics on intracellular mycobacterial load was also assessed.  Results. Macrophages infected with intracellular mycobacteria phenocopied NPC cells, exhibiting accumulation of multiple lipid types, reduced lysosomal Ca 2+ levels, and defects in intracellular trafficking. These NPC phenotypes could also be induced using only lipids/glycomycolates from the mycobacterial cell wall. These data suggest that intracellular mycobacteria inhibit the NPC pathway, likely via inhibition of the NPC1 protein, and subsequently induce altered acidic store Ca 2+ homeostasis. Reduced lysosomal calcium levels may provide a mechanistic explanation for the reduced levels of phagosome-lysosome fusion in mycobacterial infection. Treatments capable of correcting defects in NPC mutant cells via modulation of host cell calcium were of benefit in

  10. Pathogenic mycobacteria achieve cellular persistence by inhibiting the Niemann-Pick Type C disease cellular pathway [version 1; referees: 2 approved, 1 approved with reservations

    Directory of Open Access Journals (Sweden)

    Paul Fineran

    2016-11-01

    Full Text Available Background. Tuberculosis remains a major global health concern. The ability to prevent phagosome-lysosome fusion is a key mechanism by which intracellular mycobacteria, including Mycobacterium tuberculosis, achieve long-term persistence within host cells. The mechanisms underpinning this key intracellular pro-survival strategy remain incompletely understood. Host macrophages infected with persistent mycobacteria share phenotypic similarities with cells taken from patients suffering from Niemann-Pick Disease Type C (NPC, a rare lysosomal storage disease in which endocytic trafficking defects and lipid accumulation within the lysosome lead to cell dysfunction and cell death. We investigated whether these shared phenotypes reflected an underlying mechanistic connection between mycobacterial intracellular persistence and the host cell pathway dysfunctional in NPC. Methods. The induction of NPC phenotypes in macrophages from wild-type mice or obtained from healthy human donors was assessed via infection with mycobacteria and subsequent measurement of lipid levels and intracellular calcium homeostasis. The effect of NPC therapeutics on intracellular mycobacterial load was also assessed. Results. Macrophages infected with persistent intracellular mycobacteria phenocopied NPC cells, exhibiting accumulation of multiple lipid types, reduced lysosomal Ca2+ levels, and defects in intracellular trafficking. These NPC phenotypes could also be induced using only lipids/glycomycolates from the mycobacterial cell wall. These data suggest that persistent intracellular mycobacteria inhibit the NPC pathway, likely via inhibition of the NPC1 protein, and subsequently induce altered acidic store Ca2+ homeostasis. Reduced lysosomal calcium levels may provide a mechanistic explanation for the reduced levels of phagosome-lysosome fusion in mycobacterial infection. Treatments capable of correcting defects in NPC mutant cells via modulation of host cell calcium were

  11. Distinct mechanisms of DNA repair in mycobacteria and their implications in attenuation of the pathogen growth.

    Science.gov (United States)

    Kurthkoti, Krishna; Varshney, Umesh

    2012-04-01

    About a third of the human population is estimated to be infected with Mycobacterium tuberculosis. Emergence of drug resistant strains and the protracted treatment strategies have compelled the scientific community to identify newer drug targets, and to develop newer vaccines. In the host macrophages, the bacterium survives within an environment rich in reactive nitrogen and oxygen species capable of damaging its genome. Therefore, for its successful persistence in the host, the pathogen must need robust DNA repair mechanisms. Analysis of M. tuberculosis genome sequence revealed that it lacks mismatch repair pathway suggesting a greater role for other DNA repair pathways such as the nucleotide excision repair, and base excision repair pathways. In this article, we summarize the outcome of research involving these two repair pathways in mycobacteria focusing primarily on our own efforts. Our findings, using Mycobacterium smegmatis model, suggest that deficiency of various DNA repair functions in single or in combinations severely compromises their DNA repair capacity and attenuates their growth under conditions typically encountered in macrophages. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  12. Non-tuberculous mycobacteria: baseline data from three sites in Papua New Guinea, 2010-2012.

    Science.gov (United States)

    Ley, Serej; Carter, Robyn; Millan, Korai; Phuanukoonnon, Suparat; Pandey, Sushil; Coulter, Christopher; Siba, Peter; Beck, Hans-Peter

    2015-01-01

    To determine the proportion of non-tuberculous mycobacteria (NTM) in samples of pulmonary tuberculosis (TB) cases from Papua New Guinea who were diagnosed using acid-fast microscopy. As part of a case detection study for TB, conducted in three provincial hospitals in Papua New Guinea, sputum samples of suspected tuberculous cases aged 15 years or older were collected from November 2010 to July 2012. Mycobacterial species isolated from sputum and grown in culture were examined to distinguish between NTM and the Mycobacterium tuberculosis complex (MTBC). NTM were detected in 4% (9/225) of sputum samples grown in culture. Five (2.2%) of them were identified as NTM only and four (1.8%) were identified as mixed cultures containing both MTBC and NTM. Four different NTM species were identified; M. fortuitum, M. intracellulare, M. terrae and M. avium. This is the first report from Papua New Guinea identifying NTM in three different locations. As NTM cannot be distinguished from M. tuberculosis through smear microscopy, the presence of NTM can lead to a false-positive diagnosis of tuberculosis. The prevalence of NTM should be determined and a diagnostic algorithm developed to confirm acid-fast bacilli in a smear as M. tuberculosis.

  13. Non-tuberculous mycobacteria: baseline data from three sites in Papua New Guinea, 2010–2012

    Science.gov (United States)

    Ley, Serej; Carter, Robyn; Millan, Korai; Phuanukoonnon, Suparat; Pandey, Sushil; Coulter, Christopher; Siba, Peter

    2015-01-01

    Objective To determine the proportion of non-tuberculous mycobacteria (NTM) in samples of pulmonary tuberculosis (TB) cases from Papua New Guinea who were diagnosed using acid-fast microscopy. Methods As part of a case detection study for TB, conducted in three provincial hospitals in Papua New Guinea, sputum samples of suspected tuberculous cases aged 15 years or older were collected from November 2010 to July 2012. Mycobacterial species isolated from sputum and grown in culture were examined to distinguish between NTM and the Mycobacterium tuberculosis complex (MTBC). Results NTM were detected in 4% (9/225) of sputum samples grown in culture. Five (2.2%) of them were identified as NTM only and four (1.8%) were identified as mixed cultures containing both MTBC and NTM. Four different NTM species were identified; M. fortuitum, M. intracellulare, M. terrae and M. avium. Discussion This is the first report from Papua New Guinea identifying NTM in three different locations. As NTM cannot be distinguished from M. tuberculosis through smear microscopy, the presence of NTM can lead to a false-positive diagnosis of tuberculosis. The prevalence of NTM should be determined and a diagnostic algorithm developed to confirm acid-fast bacilli in a smear as M. tuberculosis. PMID:26798558

  14. Drug Susceptibility Testing of 31 Antimicrobial Agents on Rapidly Growing Mycobacteria Isolates from China.

    Science.gov (United States)

    Pang, Hui; Li, Guilian; Zhao, Xiuqin; Liu, Haican; Wan, Kanglin; Yu, Ping

    2015-01-01

    Several species of rapidly growing mycobacteria (RGM) are now recognized as human pathogens. However, limited data on effective drug treatments against these organisms exists. Here, we describe the species distribution and drug susceptibility profiles of RGM clinical isolates collected from four southern Chinese provinces from January 2005 to December 2012. Clinical isolates (73) were subjected to in vitro testing with 31 antimicrobial agents using the cation-adjusted Mueller-Hinton broth microdilution method. The isolates included 55 M. abscessus, 11 M. fortuitum, 3 M. chelonae, 2 M. neoaurum, and 2 M. septicum isolates. M. abscessus (75.34%) and M. fortuitum (15.07%), the most common species, exhibited greater antibiotic resistance than the other three species. The isolates had low resistance to amikacin, linezolid, and tigecycline, and high resistance to first-line antituberculous agents, amoxicillin-clavulanic acid, rifapentine, dapsone, thioacetazone, and pasiniazid. M. abscessus and M. fortuitum were highly resistant to ofloxacin and rifabutin, respectively. The isolates showed moderate resistance to the other antimicrobial agents. Our results suggest that tigecycline, linezolid, clofazimine, and cefmetazole are appropriate choices for M. abscessus infections. Capreomycin, sulfamethoxazole, tigecycline, clofazimine, and cefmetazole are potentially good choices for M. fortuitum infections. Our drug susceptibility data should be useful to clinicians.

  15. Clinical and taxonomic status of pathogenic nonpigmented or late-pigmenting rapidly growing mycobacteria.

    Science.gov (United States)

    Brown-Elliott, Barbara A; Wallace, Richard J

    2002-10-01

    The history, taxonomy, geographic distribution, clinical disease, and therapy of the pathogenic nonpigmented or late-pigmenting rapidly growing mycobacteria (RGM) are reviewed. Community-acquired disease and health care-associated disease are highlighted for each species. The latter grouping includes health care-associated outbreaks and pseudo-outbreaks as well as sporadic disease cases. Treatment recommendations for each species and type of disease are also described. Special emphasis is on the Mycobacterium fortuitum group, including M. fortuitum, M. peregrinum, and the unnamed third biovariant complex with its recent taxonomic changes and newly recognized species (including M. septicum, M. mageritense, and proposed species M. houstonense and M. bonickei). The clinical and taxonomic status of M. chelonae, M. abscessus, and M. mucogenicum is also detailed, along with that of the closely related new species, M. immunogenum. Additionally, newly recognized species, M. wolinskyi and M. goodii, as well as M. smegmatis sensu stricto, are included in a discussion of the M. smegmatis group. Laboratory diagnosis of RGM using phenotypic methods such as biochemical testing and high-performance liquid chromatography and molecular methods of diagnosis are also discussed. The latter includes PCR-restriction fragment length polymorphism analysis, hybridization, ribotyping, and sequence analysis. Susceptibility testing and antibiotic susceptibility patterns of the RGM are also annotated, along with the current recommendations from the National Committee for Clinical Laboratory Standards (NCCLS) for mycobacterial susceptibility testing.

  16. Drug Susceptibility Testing of 31 Antimicrobial Agents on Rapidly Growing Mycobacteria Isolates from China

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    Hui Pang

    2015-01-01

    Full Text Available Objectives. Several species of rapidly growing mycobacteria (RGM are now recognized as human pathogens. However, limited data on effective drug treatments against these organisms exists. Here, we describe the species distribution and drug susceptibility profiles of RGM clinical isolates collected from four southern Chinese provinces from January 2005 to December 2012. Methods. Clinical isolates (73 were subjected to in vitro testing with 31 antimicrobial agents using the cation-adjusted Mueller-Hinton broth microdilution method. The isolates included 55 M. abscessus, 11 M. fortuitum, 3 M. chelonae, 2 M. neoaurum, and 2 M. septicum isolates. Results. M. abscessus (75.34% and M. fortuitum (15.07%, the most common species, exhibited greater antibiotic resistance than the other three species. The isolates had low resistance to amikacin, linezolid, and tigecycline, and high resistance to first-line antituberculous agents, amoxicillin-clavulanic acid, rifapentine, dapsone, thioacetazone, and pasiniazid. M. abscessus and M. fortuitum were highly resistant to ofloxacin and rifabutin, respectively. The isolates showed moderate resistance to the other antimicrobial agents. Conclusions. Our results suggest that tigecycline, linezolid, clofazimine, and cefmetazole are appropriate choices for M. abscessus infections. Capreomycin, sulfamethoxazole, tigecycline, clofazimine, and cefmetazole are potentially good choices for M. fortuitum infections. Our drug susceptibility data should be useful to clinicians.

  17. Dehalogenation of Haloalkanes by Mycobacterium tuberculosis H37Rv and Other Mycobacteria

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    Jesenská, Andrea; Sedlác̆ek, Ivo; Damborský, Jir̆í

    2000-01-01

    Haloalkane dehalogenases convert haloalkanes to their corresponding alcohols by a hydrolytic mechanism. To date, various haloalkane dehalogenases have been isolated from bacteria colonizing environments that are contaminated with halogenated compounds. A search of current databases with the sequences of these known haloalkane dehalogenases revealed the presence of three different genes encoding putative haloalkane dehalogenases in the genome of the human parasite Mycobacterium tuberculosis H37Rv. The ability of M. tuberculosis and several other mycobacterial strains to dehalogenate haloaliphatic compounds was therefore studied. Intact cells of M. tuberculosis H37Rv were found to dehalogenate 1-chlorobutane, 1-chlorodecane, 1-bromobutane, and 1,2-dibromoethane. Nine isolates of mycobacteria from clinical material and four strains from a collection of microorganisms were found to be capable of dehalogenating 1,2-dibromoethane. Crude extracts prepared from two of these strains, Mycobacterium avium MU1 and Mycobacterium smegmatis CCM 4622, showed broad substrate specificity toward a number of halogenated substrates. Dehalogenase activity in the absence of oxygen and the identification of primary alcohols as the products of the reaction suggest a hydrolytic dehalogenation mechanism. The presence of dehalogenases in bacterial isolates from clinical material, including the species colonizing both animal tissues and free environment, indicates a possible role of parasitic microorganisms in the distribution of degradation genes in the environment. PMID:10618227

  18. Nontuberculous mycobacteria in respiratory samples from patients with pulmonary tuberculosis in the state of Rondonia, Brazil

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    Cleoni Alves Mendes de Lima

    2013-06-01

    Full Text Available The main cause of pulmonary tuberculosis (TB is infection with Mycobacterium tuberculosis (MTB. We aimed to evaluate the contribution of nontuberculous mycobacteria (NTM to pulmonary disease in patients from the state of Rondônia using respiratory samples and epidemiological data from TB cases. Mycobacterium isolates were identified using a combination of conventional tests, polymerase chain reaction-based restriction enzyme analysis of hsp65 gene and hsp65 gene sequencing. Among the 1,812 cases suspected of having pulmonary TB, 444 yielded bacterial cultures, including 369 cases positive for MTB and 75 cases positive for NTM. Within the latter group, 14 species were identified as Mycobacterium abscessus, Mycobacterium avium, Mycobacterium fortuitum, Mycobacterium intracellulare, Mycobacterium gilvum, Mycobacterium gordonae, Mycobacterium asiaticum, Mycobacterium tusciae, Mycobacterium porcinum, Mycobacterium novocastrense, Mycobacterium simiae, Mycobacterium szulgai, Mycobacterium phlei and Mycobacterium holsaticum and 13 isolates could not be identified at the species level. The majority of NTM cases were observed in Porto Velho and the relative frequency of NTM compared with MTB was highest in Ji-Paraná. In approximately half of the TB subjects with NTM, a second sample containing NTM was obtained, confirming this as the disease-causing agent. The most frequently observed NTM species were M. abscessus and M. avium and because the former species is resistant to many antibiotics and displays unsatisfactory cure rates, the implementation of rapid identification of mycobacterium species is of considerable importance.

  19. Decolorization of Malachite Green and Crystal Violet by Waterborne Pathogenic Mycobacteria

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    Jones, Jefferson J.; Falkinham III, Joseph O.

    2003-01-01

    Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium scrofulaceum, Mycobacterium marinum, and Mycobacterium chelonae tolerate high concentrations of the dyes malachite green and crystal violet. Cells of strains of those species decolorized (reduced) both malachite green and crystal violet. Because decolorized malachite green lacked antimicrobial activity, the resistance of these mycobacteria could be due, in part, to their ability to decolorize the dyes. Small amounts of malachite green and its reduced, decolorized product were detected in the lipid fraction of M. avium strain A5 cells grown in the presence of malachite green, suggesting that a minor component of resistance could be due to sequestering the dyes in the extensive mycobacterial cell surface lipid. The membrane fraction of M. avium strain A5 had at least a fivefold-higher specific decolorization rate than did the crude extract, suggesting that the decolorization activity is membrane associated. The malachite green-decolorizing activity of the membrane fraction of M. avium strain A5 was abolished by either boiling or proteinase exposure, suggesting that the decolorizing activity was due to a protein. Decolorization activity of membrane fractions was stimulated by ferrous ion and inhibited by dinitrophenol and metyrapone. PMID:12821489

  20. Mutation of environmental mycobacteria to resist silver nanoparticles also confers resistance to a common antibiotic.

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    Larimer, Curtis; Islam, Mohammad Shyful; Ojha, Anil; Nettleship, Ian

    2014-08-01

    Non-tuberculous mycobacteria are a threat to human health, gaining entry to the body through contaminated water systems, where they form persistent biofilms despite extensive attempts at disinfection. Silver is a natural antibacterial agent and in nanoparticle form activity is increased by a high surface area. Silver nanoparticles (AgNPs) have been used as alternative disinfectants in circulating water systems, washing machines and even clothing. However, nanoparticles, like any other antibiotic that has a pervasive durable presence, carry the risk of creating a resistant population. In this study Mycobacterium smegmatis strain mc(2)155 was cultured in AgNP enriched agar such that only a small population survived. Surviving cultures were isolated and re-exposed to AgNPs and AgNO3 and resistance to silver was compared to a negative control. After only a single exposure, mutant M. smegmatis populations were resistant to AgNPs and AgNO3. Further, the silver resistant mutants were exposed to antibiotics to determine if general resistance had been conferred. The minimum inhibitory concentration of isoniazid was four times higher for silver resistant mutants than for strain mc(2)155. However, core resistance was not conferred to other toxic metal ions. The mutants had lower resistance to CuSO4 and ZnSO4 than the mc(2)155 strain.

  1. Current Methods in the Molecular Typing of Mycobacterium tuberculosis and Other Mycobacteria

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    van Ingen, Jakko; Dziadek, Jarosław; Mazur, Paweł K.; Bielecki, Jacek

    2014-01-01

    In the epidemiology of tuberculosis (TB) and nontuberculous mycobacterial (NTM) diseases, as in all infectious diseases, the key issue is to define the source of infection and to disclose its routes of transmission and dissemination in the environment. For this to be accomplished, the ability of discerning and tracking individual Mycobacterium strains is of critical importance. Molecular typing methods have greatly improved our understanding of the biology of mycobacteria and provide powerful tools to combat the diseases caused by these pathogens. The utility of various typing methods depends on the Mycobacterium species under investigation as well as on the research question. For tuberculosis, different methods have different roles in phylogenetic analyses and person-to-person transmission studies. In NTM diseases, most investigations involve the search for environmental sources or phylogenetic relationships. Here, too, the type of setting determines which methodology is most suitable. Within this review, we summarize currently available molecular methods for strain typing of M. tuberculosis and some NTM species, most commonly associated with human disease. For the various methods, technical practicalities as well as discriminatory power and accomplishments are reviewed. PMID:24527454

  2. Control of biotin biosynthesis in mycobacteria by a pyruvate carboxylase dependent metabolic signal.

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    Lazar, Nathaniel; Fay, Allison; Nandakumar, Madhumitha; Boyle, Kerry E; Xavier, Joao; Rhee, Kyu; Glickman, Michael S

    2017-12-01

    Biotin is an essential cofactor utilized by all domains of life, but only synthesized by bacteria, fungi and plants, making biotin biosynthesis a target for antimicrobial development. To understand biotin biosynthesis in mycobacteria, we executed a genetic screen in Mycobacterium smegmatis for biotin auxotrophs and identified pyruvate carboxylase (Pyc) as required for biotin biosynthesis. The biotin auxotrophy of the pyc::tn strain is due to failure to transcriptionally induce late stage biotin biosynthetic genes in low biotin conditions. Loss of bioQ, the repressor of biotin biosynthesis, in the pyc::tn strain reverted biotin auxotrophy, as did reconstituting the last step of the pathway through heterologous expression of BioB and provision of its substrate DTB. The role of Pyc in biotin regulation required its catalytic activities and could be supported by M. tuberculosis Pyc. Quantitation of the kinetics of depletion of biotinylated proteins after biotin withdrawal revealed that Pyc is the most rapidly depleted biotinylated protein and metabolomics revealed a broad metabolic shift in wild type cells upon biotin withdrawal which was blunted in cell lacking Pyc. Our data indicate that mycobacterial cells monitor biotin sufficiency through a metabolic signal generated by dysfunction of a biotinylated protein of central metabolism. © 2017 John Wiley & Sons Ltd.

  3. Technical note: Avoiding the direct inversion of the numerator relationship matrix for genotyped animals in single-step genomic best linear unbiased prediction solved with the preconditioned conjugate gradient.

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    Masuda, Y; Misztal, I; Legarra, A; Tsuruta, S; Lourenco, D A L; Fragomeni, B O; Aguilar, I

    2017-01-01

    This paper evaluates an efficient implementation to multiply the inverse of a numerator relationship matrix for genotyped animals () by a vector (). The computation is required for solving mixed model equations in single-step genomic BLUP (ssGBLUP) with the preconditioned conjugate gradient (PCG). The inverse can be decomposed into sparse matrices that are blocks of the sparse inverse of a numerator relationship matrix () including genotyped animals and their ancestors. The elements of were rapidly calculated with the Henderson's rule and stored as sparse matrices in memory. Implementation of was by a series of sparse matrix-vector multiplications. Diagonal elements of , which were required as preconditioners in PCG, were approximated with a Monte Carlo method using 1,000 samples. The efficient implementation of was compared with explicit inversion of with 3 data sets including about 15,000, 81,000, and 570,000 genotyped animals selected from populations with 213,000, 8.2 million, and 10.7 million pedigree animals, respectively. The explicit inversion required 1.8 GB, 49 GB, and 2,415 GB (estimated) of memory, respectively, and 42 s, 56 min, and 13.5 d (estimated), respectively, for the computations. The efficient implementation required <1 MB, 2.9 GB, and 2.3 GB of memory, respectively, and <1 sec, 3 min, and 5 min, respectively, for setting up. Only <1 sec was required for the multiplication in each PCG iteration for any data sets. When the equations in ssGBLUP are solved with the PCG algorithm, is no longer a limiting factor in the computations.

  4. Mycobacteria exploit three genetically distinct DNA double-strand break repair pathways.

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    Gupta, Richa; Barkan, Daniel; Redelman-Sidi, Gil; Shuman, Stewart; Glickman, Michael S

    2011-01-01

    Bacterial pathogens rely on their DNA repair pathways to resist genomic damage inflicted by the host. DNA double-strand breaks (DSBs) are especially threatening to bacterial viability. DSB repair by homologous recombination (HR) requires nucleases that resect DSB ends and a strand exchange protein that facilitates homology search. RecBCD and RecA perform these functions in Escherichia coli and constitute the major pathway of error-free DSB repair. Mycobacteria, including the human pathogen M. tuberculosis, elaborate an additional error-prone pathway of DSB repair via non-homologous end-joining (NHEJ) catalysed by Ku and DNA ligase D (LigD). Little is known about the relative contributions of HR and NHEJ to mycobacterial chromosome repair, the factors that dictate pathway choice, or the existence of additional DSB repair pathways. Here we demonstrate that Mycobacterium smegmatis has three DSB repair pathway options: HR, NHEJ and a novel mechanism of single-strand annealing (SSA). Inactivation of NHEJ or SSA is compensated by elevated HR. We find that mycobacterial RecBCD does not participate in HR or confer resistance to ionizing radiation (IR), but is required for the RecA-independent SSA pathway. In contrast, the mycobacterial helicase-nuclease AdnAB participates in the RecA-dependent HR pathway, and is a major determinant of resistance to IR and oxidative DNA damage. These findings reveal distinctive features of mycobacterial DSB repair, most notably the dedication of the RecBCD and AdnAB helicase-nuclease machines to distinct repair pathways. © 2010 Blackwell Publishing Ltd.

  5. Oral Tolerance to Environmental Mycobacteria Interferes with Intradermal, but Not Pulmonary, Immunization against Tuberculosis.

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    Dominique N Price

    2016-05-01

    Full Text Available Bacille Calmette-Guérin (BCG is currently the only approved vaccine against tuberculosis (TB and is administered in over 150 countries worldwide. Despite its widespread use, the vaccine has a variable protective efficacy of 0-80%, with the lowest efficacy rates in tropical regions where TB is most prevalent. This variability is partially due to ubiquitous environmental mycobacteria (EM found in soil and water sources, with high EM prevalence coinciding with areas of poor vaccine efficacy. In an effort to elucidate the mechanisms underlying EM interference with BCG vaccine efficacy, we exposed mice chronically to Mycobacterium avium (M. avium, a specific EM, by two different routes, the oral and intradermal route, to mimic human exposure. After intradermal BCG immunization in mice exposed to oral M. avium, we saw a significant decrease in the pro-inflammatory cytokine IFN-γ, and an increase in T regulatory cells and the immunosuppressive cytokine IL-10 compared to naïve BCG-vaccinated animals. To circumvent the immunosuppressive effect of oral M. avium exposure, we vaccinated mice by the pulmonary route with BCG. Inhaled BCG immunization rescued IFN-γ levels and increased CD4 and CD8 T cell recruitment into airways in M. avium-presensitized mice. In contrast, intradermal BCG vaccination was ineffective at T cell recruitment into the airway. Pulmonary BCG vaccination proved protective against Mtb infection regardless of previous oral M. avium exposure, compared to intradermal BCG immunization. In conclusion, our data indicate that vaccination against TB by the pulmonary route increases BCG vaccine efficacy by avoiding the immunosuppressive interference generated by chronic oral exposure to EM. This has implications in TB-burdened countries where drug resistance is on the rise and health care options are limited due to economic considerations. A successful vaccine against TB is necessary in these areas as it is both effective and economical.

  6. In-vitro antimycobacterial drug susceptibility testing of non-tubercular mycobacteria by tetrazolium microplate assay.

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    Sankar, Manimuthu Mani; Gopinath, Krishnamoorthy; Singla, Roopak; Singh, Sarman

    2008-07-11

    Non-tubercular mycobacteria (NTM) has not been given due attention till the recent epidemic of HIV. This has led to increasing interest of health care workers in their biology, epidemiology and drug resistance. However, timely detection and drug susceptibility profiling of NTM isolates are always difficult in resource poor settings like India. Furthermore, no standardized methodology or guidelines are available to reproduce the results with clinical concordance. To find an alternative and rapid method for performing the drug susceptibility assay in a resource limited settings like India, we intended to evaluate the utility of Tetrazolium microplate assay (TEMA) in comparison with proportion method for reporting the drug resistance in clinical isolates of NTM. A total of fifty-five NTM isolates were tested for susceptibility against Streptomycin, Rifampicin, Ethambutol, Ciprofloxacin, Ofloxacin, Azithromycin, and Clarithromycin by TEMA and the results were compared with agar proportion method (APM). Of the 55 isolates, 23 (41.8%) were sensitive to all the drugs and the remaining 32 (58.2%) were resistant to at least one drug. TEMA had very good concordance with APM except with minor discrepancies. Susceptibility results were obtained in the median of 5 to 9 days by TEMA. The NTM isolates were highly sensitive against Ofloxacin (98.18% sensitive) and Ciprofloxacin (90.09% sensitive). M. mucogenicum was sensitive only to Clarithromycin and resistant to all the other drugs tested. The concordance between TEMA and APM ranged between 96.4 - 100%. Tetrazolium Microplate Assay is a rapid and highly reproducible method. However, it must be performed only in tertiary level Mycobacteriology laboratories with proper bio-safety conditions.

  7. Microculture in biphasic medium with silicone-coated slides for isolation of mycobacteria.

    Science.gov (United States)

    Shibayama, H; Galván, F J; Contreras, C

    1996-09-01

    The study reported here, seeking to develop a simple, practical, sensitive, and inexpensive technique for microbial diagnosis of tuberculosis, used a combination of biphasic media and microculture techniques to augment the sensitivity of traditional culture methods. A total of 540 sputum samples (5 mL each) were obtained from 180 patients with suspected tuberculosis in Mexico City. These samples were treated with Hanks reagent, neutralized with 25% HCl, and centrifuged. In each case the resulting residue was combined with liquid media (Sula medium or a phosphate-buffered control solution) and was inoculated into a bottle containing a solid medium (Löwenstein-Jensen-Holm or Middlebrook). A silicone-coated slide appropriate for culture of hydrophobic mycobacteria was inserted in each bottle, and the cultures (examined weekly) were incubated at 37 degrees C until the first macroscopic bacterial growth was detected or for up to eight weeks if none was detected. When such growth was detected, or at the end of eight weeks, each slide was withdrawn from the bottle, sterilized, stained by Kinyoun's method, and examined microscopically. Following 2-4 weeks of incubation, macroscopic bacterial growth was detected in 71 bottles and was confirmed by microscopic examination of the corresponding slides. No macroscopic bacterial growth was found in any of the remaining 469 bottles, but microscopic growth was observed on 77 of the slides examined after eight weeks. The authors conclude that this method represents a noteworthy improvement over standard culture methods in terms of bacterial isolation and suggest that its case, economy, and practicality make it suitable for application in developing countries.

  8. Unique flexibility in energy metabolism allows mycobacteria to combat starvation and hypoxia.

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    Michael Berney

    Full Text Available Mycobacteria are a group of obligate aerobes that require oxygen for growth, but paradoxically have the ability to survive and metabolize under hypoxia. The mechanisms responsible for this metabolic plasticity are unknown. Here, we report on the adaptation of Mycobacterium smegmatis to slow growth rate and hypoxia using carbon-limited continuous culture. When M. smegmatis is switched from a 4.6 h to a 69 h doubling time at a constant oxygen saturation of 50%, the cells respond through the down regulation of respiratory chain components and the F1Fo-ATP synthase, consistent with the cells lower demand for energy at a reduced growth rate. This was paralleled by an up regulation of molecular machinery that allowed more efficient energy generation (i.e. Complex I and the use of alternative electron donors (e.g. hydrogenases and primary dehydrogenases to maintain the flow of reducing equivalents to the electron transport chain during conditions of severe energy limitation. A hydrogenase mutant showed a 40% reduction in growth yield highlighting the importance of this enzyme in adaptation to low energy supply. Slow growing cells at 50% oxygen saturation subjected to hypoxia (0.6% oxygen saturation responded by switching on oxygen scavenging cytochrome bd, proton-translocating cytochrome bc1-aa3 supercomplex, another putative hydrogenase, and by substituting NAD+-dependent enzymes with ferredoxin-dependent enzymes thus highlighting a new pattern of mycobacterial adaptation to hypoxia. The expression of ferredoxins and a hydrogenase provides a potential conduit for disposing of and transferring electrons in the absence of exogenous electron acceptors. The use of ferredoxin-dependent enzymes would allow the cell to maintain a high carbon flux through its central carbon metabolism independent of the NAD+/NADH ratio. These data demonstrate the remarkable metabolic plasticity of the mycobacterial cell and provide a new framework for understanding their

  9. Mycobacteria attenuate nociceptive responses by formyl peptide receptor triggered opioid peptide release from neutrophils.

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    Heike L Rittner

    2009-04-01

    Full Text Available In inflammation, pain is regulated by a balance of pro- and analgesic mediators. Analgesic mediators include opioid peptides which are secreted by neutrophils at the site of inflammation, leading to activation of opioid receptors on peripheral sensory neurons. In humans, local opioids and opioid peptides significantly downregulate postoperative as well as arthritic pain. In rats, inflammatory pain is induced by intraplantar injection of heat inactivated Mycobacterium butyricum, a component of complete Freund's adjuvant. We hypothesized that mycobacterially derived formyl peptide receptor (FPR and/or toll like receptor (TLR agonists could activate neutrophils, leading to opioid peptide release and inhibition of inflammatory pain. In complete Freund's adjuvant-induced inflammation, thermal and mechanical nociceptive thresholds of the paw were quantified (Hargreaves and Randall-Selitto methods, respectively. Withdrawal time to heat was decreased following systemic neutrophil depletion as well as local injection of opioid receptor antagonists or anti-opioid peptide (i.e. Met-enkephalin, beta-endorphin antibodies indicating an increase in pain. In vitro, opioid peptide release from human and rat neutrophils was measured by radioimmunoassay. Met-enkephalin release was triggered by Mycobacterium butyricum and formyl peptides but not by TLR-2 or TLR-4 agonists. Mycobacterium butyricum induced a rise in intracellular calcium as determined by FURA loading and calcium imaging. Opioid peptide release was blocked by intracellular calcium chelation as well as phosphoinositol-3-kinase inhibition. The FPR antagonists Boc-FLFLF and cyclosporine H reduced opioid peptide release in vitro and increased inflammatory pain in vivo while TLR 2/4 did not appear to be involved. In summary, mycobacteria activate FPR on neutrophils, resulting in tonic secretion of opioid peptides from neutrophils and in a decrease in inflammatory pain. Future therapeutic strategies may aim

  10. Rapid-Growing Mycobacteria Infections in Medical Tourists: Our Experience and Literature Review.

    Science.gov (United States)

    Singh, Mansher; Dugdale, Caitlin M; Solomon, Isaac H; Huang, Anne; Montgomery, Mary W; Pomahac, Bohdan; Yawetz, Sigal; Maguire, James H; Talbot, Simon G

    2016-09-01

    "Medical tourism" has gained popularity over the past few decades. This is particularly common with patients seeking elective cosmetic surgery in the developing world. However, the risk of severe and unusual infectious complications appears to be higher than for patients undergoing similar procedures in the United States. The authors describe their experience with atypical mycobacterial infections in cosmetic surgical patients returning to the United States postoperatively. A review of patient medical records presenting with infectious complications after cosmetic surgery between January 2010 and July 2015 was performed. Patients presenting with mycobacterial infections following cosmetic surgery were reviewed in detail. An extensive literature review was performed for rapid-growing mycobacteria (RGM) related to cosmetic procedures. Between January 2010 and July 2015, three patients presented to our institution with culture-proven Mycobacterium abscessus at the sites of recent cosmetic surgery. All had surgery performed in the developing world. The mean age of these patients was 36 years (range, 29-44 years). There was a delay of up to 16 weeks between the initial presentation and correct diagnosis. All patients were treated with surgical drainage and combination antibiotics with complete resolution. We present series of patients with mycobacterial infections after cosmetic surgery in the developing world. This may be related to the endemic nature of these bacteria and/or inadequate sterilization or sterile technique. Due to low domestic incidence of these infections, diagnosis may be difficult and/or delayed. Consulting physicians should have a low threshold to consider atypical etiologies in such scenarios. 5 Therapeutic. © 2016 The American Society for Aesthetic Plastic Surgery, Inc. Reprints and permission: journals.permissions@oup.com.

  11. Effects of mycobacteria major secretion protein, Ag85B, on allergic inflammation in the lung.

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    Yusuke Tsujimura

    Full Text Available Many epidemiological studies have suggested that the recent increase in prevalence and severity of allergic diseases such as asthma is inversely correlated with Mycobacterium bovis bacillus Calmette Guerin (BCG vaccination. However, the underlying mechanisms by which mycobacterial components suppress allergic diseases are not yet fully understood. Here we showed the inhibitory mechanisms for development of allergic airway inflammation by using highly purified recombinant Ag85B (rAg85B, which is one of the major protein antigens secreted from M. tuberculosis. Ag85B is thought to be a single immunogenic protein that can elicit a strong Th1-type immune response in hosts infected with mycobacteria, including individuals vaccinated with BCG. Administration of rAg85B showed a strong inhibitory effect on the development of allergic airway inflammation with induction of Th1-response and IL-17and IL-22 production. Both cytokines induced by rAg85B were involved in the induction of Th17-related cytokine-production innate immune cells in the lung. Administration of neutralizing antibodies to IL-17 or IL-22 in rAg85B-treated mice revealed that IL-17 induced the infiltration of neutrophils in BAL fluid and that allergen-induced bronchial eosinophilia was inhibited by IL-22. Furthermore, enhancement of the expression of genes associated with tissue homeostasis and wound healing was observed in bronchial tissues after rAg85B administration in a Th17-related cytokine dependent manner. The results of this study provide evidence for the potential usefulness of rAg85B as a novel approach for anti-allergic effect and tissue repair other than the role as a conventional TB vaccine.

  12. Two novel species of rapidly growing mycobacteria: Mycobacterium lehmannii sp. nov. and Mycobacterium neumannii sp. nov.

    Science.gov (United States)

    Nouioui, Imen; Sangal, Vartul; Carro, Lorena; Teramoto, Kanae; Jando, Marlen; Montero-Calasanz, Maria Del Carmen; Igual, José Mariano; Sutcliffe, Iain; Goodfellow, Michael; Klenk, Hans-Peter

    2017-12-01

    Two rapidly growing mycobacteria with identical 16S rRNA gene sequences were the subject of a polyphasic taxonomic study. The strains formed a well-supported subclade in the mycobacterial 16S rRNA gene tree and were most closely associated with the type strain of Mycobacterium novocastrense. Single and multilocus sequence analyses based on hsp65, rpoB and 16S rRNA gene sequences showed that strains SN 1900 T and SN 1904 T are phylogenetically distinct but share several chemotaxonomic and phenotypic features that are are consistent with their classification in the genus Mycobacterium. The two strains were distinguished by their different fatty acid and mycolic acid profiles, and by a combination of phenotypic features. The digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) values for strains SN 1900 T and SN 1904 T were 61.0 % and 94.7 %, respectively; in turn, the corresponding dDDH and ANI values with M. novocastrense DSM 44203 T were 41.4 % and 42.8 % and 89.3 % and 89.5 %, respectively. These results show that strains SN1900 T and SN 1904 T form new centres of taxonomic variation within the genus Mycobacterium. Consequently, strains SN 1900 T (40 T =CECT 8763 T =DSM 43219 T ) and SN 1904 T (2409 T =CECT 8766 T =DSM 43532 T ) are considered to represent novel species, for which the names Mycobacteriumlehmannii sp. nov. and Mycobacteriumneumannii sp. nov. are proposed. A strain designated as 'Mycobacteriumacapulsensis' was shown to be a bona fide member of the putative novel species, M. lehmannii.

  13. High Rates of Non-Tuberculous Mycobacteria Isolation in Mozambican Children with Presumptive Tuberculosis.

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    Elisa López-Varela

    Full Text Available Non-tuberculous mycobacteria (NTM can cause disease which can be clinically and radiologically undistinguishable from tuberculosis (TB, posing a diagnostic and therapeutic challenge in high TB settings. We aim to describe the prevalence of NTM isolation and its clinical characteristics in children from rural Mozambique.This study was part of a community TB incidence study in children <3 years of age. Gastric aspirate and induced sputum sampling were performed in all presumptive TB cases and processed for smear testing using fluorochrome staining and LED Microscopy, liquid and solid culture, and molecular identification by GenoType® Mycobacterium CM/AS assays.NTM were isolated in 26.3% (204/775 of children. The most prevalent NTM species was M. intracellulare (N = 128, followed by M. scrofulaceum (N = 35 and M. fortuitum (N = 9. Children with NTM were significantly less symptomatic and less likely to present with an abnormal chest radiograph than those with M. tuberculosis. NTM were present in 21.6% of follow-up samples and 25 children had the same species isolated from ≥2 separate samples. All were considered clinically insignificant and none received specific treatment. Children with NTM isolates had equal all cause mortality and likelihood of TB treatment as those with negative culture although they were less likely to have TB ruled out.NTM isolation is frequent in presumptive TB cases but was not clinically significant in this patient cohort. However, it can contribute to TB misdiagnosis. Further studies are needed to understand the epidemiology and the clinical significance of NTM in children.

  14. Identification of non-tuberculous mycobacteria isolated from clinical specimens at a tertiary care hospital: a cross-sectional study.

    Science.gov (United States)

    Ahmed, Imran; Jabeen, Kauser; Hasan, Rumina

    2013-10-22

    Non-tuberculous mycobacteria (NTM) are opportunistic pathogens in immuno-compromised patients. They are also increasingly recognized as pathogens in immuno-competent individuals. Globally, an increase in NTM isolation is being reported with a varied geographic prevalence of different species around the world. There is lack of data on species distribution of these organisms from Pakistan. Treatment options differ according to the species isolated and its susceptibility profile. Knowledge of local species variation would help targeted therapy. This study was performed to determine frequencies of different NTM species isolated from various clinical specimens submitted at a tertiary care hospital laboratory. NTM isolated from 25955 clinical specimens over a period of two years (2010 to 2011) were included. All NTM were identified using conventional tests. Drug susceptibility testing (DST) was performed by broth microdilution and interpreted according to Clinical and Laboratory Standards Institute's document M24-A2. A total of 104 NTM were included in the study. Of these, 76% (54/71) rapidly growing mycobacteria (RGM) and 57.6% (19/33) slow growing mycobacteria (SGM) could be further identified. Mycobacterium fortuitum (21/54) was the commonest NTM identified among RGM followed by M. mucogenicum (12/54) and M. smegmatis (11/54). Among SGM, M. avium complex (MAC) was the most frequent (14/19). Clinical significance could be assessed in a limited number (52/104) of NTM isolates and MAC appeared to be the commonest significant NTM. Three extra-pulmonary cases were found to be healthcare associated infections. DST results for RGM showed susceptibility to amikacin (100%), clarithromycin (100%, except M. fortuitum where it is not reportable), linezolid (90%) and moxifloxacin (75%). Whereas SGM were susceptible to clarithromycin (100%), linezolid (58.8%) and moxifloxacin (64.7%). This is the first study reporting NTM species and their clinical significance isolated from

  15. Non-tuberculous mycobacteria I: one year clinical isolates identification in Tertiary Hospital Aids Reference Center, Rio de Janeiro, Brazil, in pre highly active antiretroviral therapy era

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    Ferreira Rosa Maria Carvalho

    2002-01-01

    Full Text Available The aim of this study was to determine the prevalence of non-tuberculous mycobacteria (NTM isolates at University Hospital, Reference Center for Aids in Rio de Janeiro, Brazil, during one year. We used standard biochemical tests for species identification and IS1245 PCR amplification was applied as a Mycobacterium avium specific identification marker. Four hundred and four specimens from 233 patients yielded acid-fast bacilli growth. M. tuberculosis was identified in 85% of the patients and NTM in 15%. NTM disseminated infection was a common event correlated with human immunodeficiency virus (HIV infected patients and only in HIV negative patients the source of NTM was non sterile site. M. avium complex (MAC was biochemically identified in 57.8% (49/83 of NTM isolates, most of them from sterile sites (75.5%, and in 94% (46/49 the IS 1245 marker specific for M. avium was present. Twenty NTM strains showed a MAC biochemical pattern with the exception of a urease-positive (99% of MAC are urease-negative, however IS1245 was detected in 96% of the strains leading to their identification as M. avium. In this group differences in NTM source was not significant. The second most frequently isolated NTM was identified as M. scrofulaceum (7.2%, followed by M. terrae (3.6%, M. gordonae (2.4%, M. chelonae (1.2%, M. fortuitum (1.2% and one strain which could not be identified. All were IS1245 negative except for one strain identified as M. scrofulaceum. It is interesting to note that non-sterile sites were the major source of these isolates (92.8%. Our finding indicated that M. avium is still the major atypical species among in the MAC isolates recovered from Brazilian Aids patients without highty active antiretroviral therapy schema. Some discrepancies were seen between the identification methods and further investigations must be done to better characterize NTM isolates using other phenotypic and genotypic methods.

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

    Science.gov (United States)

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

    2014-09-01

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

  17. Common genotypes of hepatitis B virus

    International Nuclear Information System (INIS)

    Idrees, M.; Khan, S.; Riazuddin, S.

    2004-01-01

    Objective: To find out the frequency of common genotypes of hepatitis-B virus (HBV). Subjects and Methods: HBV genotypes were determined in 112 HBV DNA positive sera by a simple and precise molecular genotyping system base on PCR using type-specific primers for the determination of genotypes of HBV A through H. Results: Four genotypes (A,B,C and D) out of total eight reported genotypes so far were identified. Genotypes A, B and C were predominant. HBV genotype C was the most predominant in this collection, appearing in 46 samples (41.7%). However, the genotypes of a total of 5 (4.46%) samples could not be determined with the present genotyping system. Mixed genotypes were seen in 8(7.14% HBV) isolates. Five of these were infected with genotypes A/D whereas two were with genotypes C/D. One patient was infected with 4 genotypes (A/B/C/D). Genotype A (68%) was predominant in Sindh genotype C was most predominant in North West Frontier Province (NWFP) (68.96) whereas genotype C and B were dominant in Punjab (39.65% and 25.86% respectively). Conclusion: All the four common genotypes of HBV found worldwide (A,B,C and D) were isolated. Genotype C is the predominant Genotypes B and C are predominant in Punjab and N.W.F.P. whereas genotype A is predominant in Sindh. (author)

  18. Amoebae as potential environmental hosts for Mycobacterium ulcerans and other mycobacteria, but doubtful actors in Buruli ulcer epidemiology.

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    Sophie Gryseels

    Full Text Available The reservoir and mode of transmission of Mycobacterium ulcerans, the causative agent of Buruli ulcer, remain unknown. Ecological, genetic and epidemiological information nonetheless suggests that M. ulcerans may reside in aquatic protozoa.We experimentally infected Acanthamoeba polyphaga with M. ulcerans and found that the bacilli were phagocytised, not digested and remained viable for the duration of the experiment. Furthermore, we collected 13 water, 90 biofilm and 45 detritus samples in both Buruli ulcer endemic and non-endemic communities in Ghana, from which we cultivated amoeboid protozoa and mycobacteria. M. ulcerans was not isolated, but other mycobacteria were as frequently isolated from intracellular as from extracellular sources, suggesting that they commonly infect amoebae in nature. We screened the samples as well as the amoeba cultures for the M. ulcerans markers IS2404, IS2606 and KR-B. IS2404 was detected in 2% of the environmental samples and in 4% of the amoeba cultures. The IS2404 positive amoeba cultures included up to 5 different protozoan species, and originated both from Buruli ulcer endemic and non-endemic communities.This is the first report of experimental infection of amoebae with M. ulcerans and of the detection of the marker IS2404 in amoeba cultures isolated from the environment. We conclude that amoeba are potential natural hosts for M. ulcerans, yet remain sceptical about their implication in the transmission of M. ulcerans to humans and their importance in the epidemiology of Buruli ulcer.

  19. Mycobacteria in water used for personal hygiene in heavy industry and collieries: a potential risk for employees.

    Science.gov (United States)

    Ulmann, Vit; Kracalikova, Anna; Dziedzinska, Radka

    2015-03-04

    Environmental mycobacteria (EM) constitute a health risk, particularly for immunocompromised people. Workers in heavy industry and in collieries represent an at-risk group of people as their immunity is often weakened by long-term employment in dusty environments, frequent smoking and an increased occurrence of pulmonary diseases. This study was concerned with the presence of EM in non-drinking water used for the hygiene of employees in six large industrial companies and collieries. Over a period of ten years, 1096 samples of surface water treated for hygiene purposes (treated surface water) and treated surface water diluted with mining water were examined. EM were detected in 63.4 and 41.5% samples of treated surface water and treated surface water diluted with mining water, respectively. Mycobacterium gordonae, M. avium-intracellulare and M. kansasii were the most frequently detected species. Adoption of suitable precautions should be enforced to reduce the incidence of mycobacteria in shower water and to decrease the infectious pressure on employees belonging to an at-risk group of people.

  20. Mycobacteria in Water Used for Personal Hygiene in Heavy Industry and Collieries: A Potential Risk for Employees

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    Vit Ulmann

    2015-03-01

    Full Text Available Environmental mycobacteria (EM constitute a health risk, particularly for immunocompromised people. Workers in heavy industry and in collieries represent an at-risk group of people as their immunity is often weakened by long-term employment in dusty environments, frequent smoking and an increased occurrence of pulmonary diseases. This study was concerned with the presence of EM in non-drinking water used for the hygiene of employees in six large industrial companies and collieries. Over a period of ten years, 1096 samples of surface water treated for hygiene purposes (treated surface water and treated surface water diluted with mining water were examined. EM were detected in 63.4 and 41.5% samples of treated surface water and treated surface water diluted with mining water, respectively. Mycobacterium gordonae, M. avium-intracellulare and M. kansasii were the most frequently detected species. Adoption of suitable precautions should be enforced to reduce the incidence of mycobacteria in shower water and to decrease the infectious pressure on employees belonging to an at-risk group of people.

  1. HBV genotypic variability in Cuba.

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    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.

  2. 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

  3. Mycobacteria contain two groEL genes: the second Mycobacterium leprae groEL gene is arranged in an operon with groES

    NARCIS (Netherlands)

    Rinke de Wit, T. F.; Bekelie, S.; Osland, A.; Miko, T. L.; Hermans, P. W.; van Soolingen, D.; Drijfhout, J. W.; Schöningh, R.; Janson, A. A.; Thole, J. E.

    1992-01-01

    In contrast to other bacterial species, mycobacteria were thus far considered to contain groEL and groES genes that are present on separate loci on their chromosomes, Here, by screening a Mycobacterium leprae lambda gt11 expression library with serum from an Ethiopian lepromatous leprosy patient,

  4. [Identification of mycobacteria by matrix-assisted laser desorption-ionization time-of-flight mass spectrometry--using reference strains and clinical isolates of Mycobacterium].

    Science.gov (United States)

    Niitsuma, Katsunao; Saito, Miwako; Koshiba, Shizuko; Kaneko, Michiyo

    2014-05-01

    Matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF MS) method is being played an important role for the inspection of clinical microorganism as a rapid and the price reduction. Mass spectra obtained by measuring become points of identification whether the peak pattern match any species mass spectral pattern. We currently use MALDI-TOF MS for rapid and accurate diagnosis of inactivated reference and clinical isolates of Mycobacterium because of the improved pretreatment techniques compared with former inspection methods that pose a higher risk of infection to the operator. The identification matching rate of score value (SV) peak pattern spectra was compared with that of conventional methods such as strain diffusion/amplification. Also, cultures were examined after a fixed number of days. Compared with the initial inspection technique, the pretreatment stage of current MALDI-TOF MS inspection techniques can improve the analysis of inactivated acid-fast bacteria that are often used as inspection criteria strains of clinical isolates. Next, we compared the concordance rate for identification between MALDI-TOF MS and conventional methods such as diffusion/amplification by comparison of peak pattern spectra and evaluated SV spectra to identify differences in the culture media after the retention period. In examination of 158 strains of clinical isolated Mycobacterium tuberculosis complex (MTC), the identification coincidence rate in the genus level in a matching pattern was 99.4%, when the species level was included 94.9%. About 37 strains of nontuberculous mycobacteria (NTM), the identification coincidence rate in the genus level was 94.6%. M. bovis BCG (Tokyo strain) in the reference strain was judged by the matching pattern to be MTC, and it suggested that they are M. tuberculosis and affinity species with high DNA homology. Nontuberculous mycobacterial M. gordonae strain JATA 33-01 shared peak pattern spectra, excluding the

  5. Echinococcus granulosus genotypes in Iran

    Science.gov (United States)

    Sharafi, Seyedeh Maryam; Rostami-Nejad, Mohammad; Moazeni, Mohammad; Yousefi, Morteza; Saneie, Behnam; Hosseini-Safa, Ahmad

    2014-01-01

    Hydatidosis, caused by Echinococcus granulosus is one of the most important zoonotic diseases, throughout most parts of the world. Hydatidosis is endemic in Iran and responsible for approximately 1% of admission to surgical wards. There are extensive genetic variations within E. granulosus and 10 different genotypes (G1–G10) within this parasite have been reported. Identification of strains is important for improvement of control and prevention of the disease. No new review article presented the situation of Echinococcus granulosus genotypes in Iran in the recent years; therefore in this paper we reviewed the different studies regarding Echinococcus granulosus genotypes in Iran. PMID:24834298

  6. Single systemic administration of Ag85B of mycobacteria DNA inhibits allergic airway inflammation in a mouse model of asthma

    Directory of Open Access Journals (Sweden)

    Karamatsu K

    2012-12-01

    Full Text Available Katsuo Karamatsu,1,2 Kazuhiro Matsuo,3 Hiroyasu Inada,4 Yusuke Tsujimura,1 Yumiko Shiogama,1,2 Akihiro Matsubara,1,2 Mitsuo Kawano,5 Yasuhiro Yasutomi1,21Laboratory of Immunoregulation and Vaccine Research, Tsukuba Primate Research Center, National Institute of Biomedical Innovation, Tsukuba, 2Division of Immunoregulation, Department of Molecular and Experimental Medicine, Mie University Graduate School of Medicine, Tsu, 3Department of Research and Development, Japan BCG Laboratory, Tokyo, 4Department of Pathology, Suzuka University of Medical Science, Suzuka, 5Department of Microbiology and Molecular Genetics, Mie University Graduate School of Medicine, Tsu, JapanAbstract: The immune responses of T-helper (Th and T-regulatory cells are thought to play a crucial role in the pathogenesis of allergic airway inflammation observed in asthma. The correction of immune response by these cells should be considered in the prevention and treatment of asthma. Native antigen 85B (Ag85B of mycobacteria, which cross-reacts among mycobacteria species, may play an important biological role in host–pathogen interaction since it elicits various immune responses by activation of Th cells. The current study investigated the antiallergic inflammatory effects of DNA administration of Ag85B from Mycobacterium kansasii in a mouse model of asthma. Immunization of BALB/c mice with alum-adsorbed ovalbumin followed by aspiration with aerosolized ovalbumin resulted in the development of allergic airway inflammation. Administration of Ag85B DNA before the aerosolized ovalbumin challenge protected the mice from subsequent induction of allergic airway inflammation. Serum and bronchoalveolar lavage immunoglobulin E levels, extent of eosinophil infiltration, and levels of Th2-type cytokines in Ag85B DNA-administered mice were significantly lower than those in control plasmid-immunized mice, and levels of Th1- and T-regulatory-type cytokines were enhanced by Ag85B

  7. Genotypic character relationship and phenotypic path coefficient analysis in chili pepper genotypes grown under tropical condition.

    Science.gov (United States)

    Usman, Magaji G; Rafii, Mohd Y; Martini, Mohammad Y; Oladosu, Yusuff; Kashiani, Pedram

    2017-03-01

    Studies on genotypic and phenotypic correlations among characters of crop plants are useful in planning, evaluating and setting selection criteria for the desired characters in a breeding program. The present study aimed to estimate the phenotypic correlation coefficients among yield and yield attributed characters and to work out the direct and indirect effects of yield-related characters on yield per plant using path coefficient analysis. Twenty-six genotypes of chili pepper were laid out in a randomized complete block design with three replications. Yield per plant showed positive and highly significant (P ≤ 0.01) correlations with most of the characters studied at both the phenotypic and genotypic levels. By contrast, disease incidence and days to flowering showed a significant negative association with yield. Fruit weight and number of fruits exerted positive direct effect on yield and also had a positive and significant (P ≤ 0.01) correlation with yield per plant. However, fruit length showed a low negative direct effect with a strong and positive indirect effect through fruit weight on yield and had a positive and significant association with yield. Longer fruits, heavy fruits and a high number of fruits are variables that are related to higher yields of chili pepper under tropical conditions and hence could be used as a reliable indicator in indirect selection for yield. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

  8. Regulation of macrophage accessory cell activity by mycobacteria. I. Ia expression in normal and irradiated mice infected with Mycobacterium mycroti

    International Nuclear Information System (INIS)

    Kaye, P.M.; Feldmann, M.

    1986-01-01

    CBA/Ca mice were infected by either the intravenous or intraperitoneal route with Mycobacterium microti and the subsequent changes in local macrophage populations examined. Following infection, the number of macrophages increased and they showed greater expression of both MHC Class II molecules. This response was not dependent on viability of the mycobacteria, in contrast to reports with other microorganisms such as Listeria. Studies in sublethally irradiated mice indicated that persistent antigen could give rise to a response after a period of host recovery which was radiation dose dependent. This procedure also highlighted differences in the regulation of different murine class II antigens in vivo, as seen by delayed re-expression of I-E antigens. Macrophage accessory cell function, as assessed by an in vitro T cell proliferation assay, correlated with Ia expression after fixation, but not after indomethacin treatment; this highlights the diverse nature of regulatory molecules produced by these cells. (author)

  9. Comparison among three methods for mycobacteria identification Comparación entre tres métodos para identificar micobacterias

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    Misael Mondragón-Barreto

    2000-11-01

    Full Text Available OBJECTIVE: To compare three methods: Biochemical tests, high-performance liquid chromatography (HPLC and polymerase chain reaction-restriction fragments length polymorphism (PCR-RFLP, for the identification of mycobacteria, and to perform a cost-benefit analysis to define an optimum identification algorithm. MATERIAL AND METHODS: One-hundred-and-seven mycobacteria isolates were identified by the three methods at Instituto de Diagnóstico y Referencia Epidemiológicos, between February of 1999 and January of 2000 and the results were compared with those of a reference laboratory using the Q-Cochran statistical test. RESULTS: PCR-RFLP was the most rapid and specific procedure but also the most expensive; biochemical tests excelled for identification of Mycobacterium tuberculosis, but were lengthy and expensive for other mycobacteria; HPLC ranked in the middle for price, speed and specificity. CONCLUSIONS: Considering the expected proportion of M. tuberculosis, the following algorithm was proposed: Initially, biochemical tests should be performed; if the results indicate a non-tuberculous mycobacteria, the isolate should be analyzed with HPLC; if results are unclear, the isolate should be analyzed using PCR-RFLP. Isolates showing a previously undescribed PCR-RFLP pattern should be characterized by DNA sequencing.OBJETIVO: Comparar tres métodos: pruebas bioquímicas, cromatografía líquida de alta resolución (HPLC, por sus siglas en inglés y reacción en cadena de la polimerasa-polimorfismo del tamaño de fragmentos de restricción (PCR-RFLP para identificar micobacterias a nivel especie, analizando costo-beneficio y proponiendo un algoritmo de identificación. MATERIAL Y MÉTODOS: Entre febrero de 1999 y enero de 2000, en los laboratorios del Instituto de Diagnóstico y Referencia Epidemiológicos se tipificaron 107 aislados de micobacterias y los resultados se compararon con los obtenidos en un laboratorio de referencia utilizando la prueba

  10. Identification of polymorphic inversions from genotypes

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    Cáceres Alejandro

    2012-02-01

    Full Text Available Abstract Background Polymorphic inversions are a source of genetic variability with a direct impact on recombination frequencies. Given the difficulty of their experimental study, computational methods have been developed to infer their existence in a large number of individuals using genome-wide data of nucleotide variation. Methods based on haplotype tagging of known inversions attempt to classify individuals as having a normal or inverted allele. Other methods that measure differences between linkage disequilibrium attempt to identify regions with inversions but unable to classify subjects accurately, an essential requirement for association studies. Results We present a novel method to both identify polymorphic inversions from genome-wide genotype data and classify individuals as containing a normal or inverted allele. Our method, a generalization of a published method for haplotype data 1, utilizes linkage between groups of SNPs to partition a set of individuals into normal and inverted subpopulations. We employ a sliding window scan to identify regions likely to have an inversion, and accumulation of evidence from neighboring SNPs is used to accurately determine the inversion status of each subject. Further, our approach detects inversions directly from genotype data, thus increasing its usability to current genome-wide association studies (GWAS. Conclusions We demonstrate the accuracy of our method to detect inversions and classify individuals on principled-simulated genotypes, produced by the evolution of an inversion event within a coalescent model 2. We applied our method to real genotype data from HapMap Phase III to characterize the inversion status of two known inversions within the regions 17q21 and 8p23 across 1184 individuals. Finally, we scan the full genomes of the European Origin (CEU and Yoruba (YRI HapMap samples. We find population-based evidence for 9 out of 15 well-established autosomic inversions, and for 52 regions

  11. Macropinocytosis is responsible for the uptake of pathogenic and non-pathogenic mycobacteria by B lymphocytes (Raji cells

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    García-Pérez Blanca Estela

    2012-10-01

    Full Text Available Abstract Background The classical roles of B cells include the production of antibodies and cytokines and the generation of immunological memory, these being key factors in the adaptive immune response. However, their role in innate immunity is currently being recognised. Traditionally, B cells have been considered non-phagocytic cells; therefore, the uptake of bacteria by B cells is not extensively documented. In this study, we analysed some of the features of non-specific bacterial uptake by B lymphocytes from the Raji cell line. In our model, B cells were infected with Mycobacterium tuberculosis (MTB, Mycobacterium smegmatis (MSM, and Salmonella typhimurium (ST. Results Our observations revealed that the Raji B cells were readily infected by the three bacteria that were studied. All of the infections induced changes in the cellular membrane during bacterial internalisation. M. smegmatis and S. typhimurium were able to induce important membrane changes that were characterised by abundant filopodia and lamellipodia formation. These membrane changes were driven by actin cytoskeletal rearrangements. The intracellular growth of these bacteria was also controlled by B cells. M. tuberculosis infection also induced actin rearrangement-driven membrane changes; however, the B cells were not able to control this infection. The phorbol 12-myristate 13-acetate (PMA treatment of B cells induced filopodia and lamellipodia formation, the production of spacious vacuoles (macropinosomes, and the fluid-phase uptake that is characteristic of macropinocytosis. S. typhimurium infection induced the highest fluid-phase uptake, although both mycobacteria also induced fluid uptake. A macropinocytosis inhibitor such as amiloride was used and abolished the bacterial uptake and the fluid-phase uptake that is triggered during the bacterial infection. Conclusions Raji B cells can internalise S. typhimurium and mycobacteria through an active process, such as

  12. Transcriptome Analysis of Mycobacteria-Specific CD4+ T Cells Identified by Activation-Induced Expression of CD154.

    Science.gov (United States)

    Kunnath-Velayudhan, Shajo; Goldberg, Michael F; Saini, Neeraj K; Johndrow, Christopher T; Ng, Tony W; Johnson, Alison J; Xu, Jiayong; Chan, John; Jacobs, William R; Porcelli, Steven A

    2017-10-01

    Analysis of Ag-specific CD4 + T cells in mycobacterial infections at the transcriptome level is informative but technically challenging. Although several methods exist for identifying Ag-specific T cells, including intracellular cytokine staining, cell surface cytokine-capture assays, and staining with peptide:MHC class II multimers, all of these have significant technical constraints that limit their usefulness. Measurement of activation-induced expression of CD154 has been reported to detect live Ag-specific CD4 + T cells, but this approach remains underexplored and, to our knowledge, has not previously been applied in mycobacteria-infected animals. In this article, we show that CD154 expression identifies adoptively transferred or endogenous Ag-specific CD4 + T cells induced by Mycobacterium bovis bacillus Calmette-Guérin vaccination. We confirmed that Ag-specific cytokine production was positively correlated with CD154 expression by CD4 + T cells from bacillus Calmette-Guérin-vaccinated mice and show that high-quality microarrays can be performed from RNA isolated from CD154 + cells purified by cell sorting. Analysis of microarray data demonstrated that the transcriptome of CD4 + CD154 + cells was distinct from that of CD154 - cells and showed major enrichment of transcripts encoding multiple cytokines and pathways of cellular activation. One notable finding was the identification of a previously unrecognized subset of mycobacteria-specific CD4 + T cells that is characterized by the production of IL-3. Our results support the use of CD154 expression as a practical and reliable method to isolate live Ag-specific CD4 + T cells for transcriptomic analysis and potentially for a range of other studies in infected or previously immunized hosts. Copyright © 2017 by The American Association of Immunologists, Inc.

  13. Translation of Genotype to Phenotype by a Hierarchy of Cell Subsystems

    OpenAIRE

    Yu, Michael Ku; Kramer, Michael; Dutkowski, Janusz; Srivas, Rohith; Licon, Katherine; Kreisberg, Jason F.; Ng, Cherie T.; Krogan, Nevan; Sharan, Roded; Ideker, Trey

    2016-01-01

    Summary Accurately translating genotype to phenotype requires accounting for the functional impact of genetic variation at many biological scales. Here we present a strategy for genotype-phenotype reasoning based on existing knowledge of cellular subsystems. These subsystems and their hierarchical organization are defined by the Gene Ontology or a complementary ontology inferred directly from previously published datasets. Guided by the ontology?s hierarchical structure, we organize genotype ...

  14. Transforming microbial genotyping: a robotic pipeline for genotyping bacterial strains.

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    Brian O'Farrell

    Full Text Available Microbial genotyping increasingly deals with large numbers of samples, and data are commonly evaluated by unstructured approaches, such as spread-sheets. The efficiency, reliability and throughput of genotyping would benefit from the automation of manual manipulations within the context of sophisticated data storage. We developed a medium- throughput genotyping pipeline for MultiLocus Sequence Typing (MLST of bacterial pathogens. This pipeline was implemented through a combination of four automated liquid handling systems, a Laboratory Information Management System (LIMS consisting of a variety of dedicated commercial operating systems and programs, including a Sample Management System, plus numerous Python scripts. All tubes and microwell racks were bar-coded and their locations and status were recorded in the LIMS. We also created a hierarchical set of items that could be used to represent bacterial species, their products and experiments. The LIMS allowed reliable, semi-automated, traceable bacterial genotyping from initial single colony isolation and sub-cultivation through DNA extraction and normalization to PCRs, sequencing and MLST sequence trace evaluation. We also describe robotic sequencing to facilitate cherrypicking of sequence dropouts. This pipeline is user-friendly, with a throughput of 96 strains within 10 working days at a total cost of 200,000 items were processed by two to three people. Our sophisticated automated pipeline can be implemented by a small microbiology group without extensive external support, and provides a general framework for semi-automated bacterial genotyping of large numbers of samples at low cost.

  15. High prevalence of Hepatitis C virus genotype 6 in Vietnam.

    Science.gov (United States)

    Pham, Duc Anh; Leuangwutiwong, Pornsawan; Jittmittraphap, Akanitt; Luplertlop, Nattanej; Bach, Hoa Khanh; Akkarathamrongsin, Srunthron; Theamboonlers, Apiradee; Poovorawan, Yong

    2009-01-01

    This study aimed to update the prevalence of the various Hepatitis C virus genotypes in Vietnamese blood donors. One hundred and three HCV antibody-positive plasma samples were collected from blood donors at the National Institute of Hematology and Blood Transfusion, Hanoi, Vietnam. All specimens were subjected to RT-PCR of the 5' untranslated region (UTR) to confirm the presence of HCV RNA. The core and NS5B regions of thh positive samples were subsequently amplified by RT-PCR followed by direct sequencing and phylogenetic analysis. Seventy out of 103 samples (68.0%) were RNA positive. Core and NS5B were successfully amplified and sequences were obtained for 70 and 65 samples, respectively. Phylogenetic analysis revealed that genotype 6a was the most predominant among Vietnamese blood donors with a prevalence of 37.1% (26/70), followed by genotype 1a at 30.0% (21/70) and genotype 1b at 17.1% (12/70). The prevalence of two other genotype 6 variants, 6e and 61 was 8.6% and 1.4%, respectively. Further analysis of recent studies showed that the geographic distribution of genotype 6 covered mainly southern China and the mainland of Southeast Asia including Vietnam, Laos, Thailand, and Myanmar. The GenBank accession numbers for the sequences reported in this study are FJ768772-FJ768906.

  16. Interleukin-1 or tumor necrosis factor-alpha augmented the cytotoxic effect of mycobacteria on human fibroblasts: application to evaluation of pathogenesis of clinical isolates of Mycobacterium tuberculosis and M. avium complex.

    Science.gov (United States)

    Takii, T; Abe, C; Tamura, A; Ramayah, S; Belisle, J T; Brennan, P J; Onozaki, K

    2001-03-01

    Mycobacteria-induced in vitro events reflecting human tuberculosis can contribute to the evaluation of the pathogenesis of Mycobacterium tuberculosis (MTB). In this study, we propose such an in vitro method based on live mycobacteria-induced cytotoxicity to human cell lines. When human lung-derived normal fibroblast cell line MRC-5 was infected with various strains of mycobacteria (M. tuberculosis H(37)Rv and H(37) Ra, Mycobacterium avium 427S and 2151SmO, and Mycobacterium bovis BCG Pasteur and Tokyo), the fibroblasts were killed by mycobacteria according to the degree of virulence. Other human originated macrophage (U-937, THP-1), myeloid (HL-60), and epithelial carcinoma (A549) cell lines exhibited a similar cytotoxic response to virulent mycobacteria. MRC-5 was most susceptible to virulent mycobacteria among various human cell lines examined. The cytotoxicity was enhanced by the proinflammatory cytokines, interleukin-1 (IL-1) and tumor necrosis factor-a (TNF-alpha), which in the absence of mycobacteria stimulate the growth of normal human fibroblasts. This in vitro evaluation system was applied to clinical isolates of drug-sensitive MTB (DS-MTB), drug-resistant MTB (DR-MTB) including multidrug-resistant (MDR-MTB), and M. avium complex (MAC). MTB strains (n = 24) exhibited strong cytotoxic activity, but MAC strains (n = 5) had only weak activity. Furthermore, there was no significant difference in cytotoxicity between DS-MTB (n = 11) and DR-MTB (n = 13). Collectively, these results suggest that this new in vitro system is useful for evaluating the pathogenesis of mycobacteria and that there was no difference in the pathogenesis between drug-susceptible and drug-resistant clinical isolates.

  17. Identification of zoonotic genotypes of Giardia duodenalis.

    Directory of Open Access Journals (Sweden)

    Hein Sprong

    Full Text Available Giardia duodenalis, originally regarded as a commensal organism, is the etiologic agent of giardiasis, a gastrointestinal disease of humans and animals. Giardiasis causes major public and veterinary health concerns worldwide. Transmission is either direct, through the faecal-oral route, or indirect, through ingestion of contaminated water or food. Genetic characterization of G. duodenalis isolates has revealed the existence of seven groups (assemblages A to G which differ in their host distribution. Assemblages A and B are found in humans and in many other mammals, but the role of animals in the epidemiology of human infection is still unclear, despite the fact that the zoonotic potential of Giardia was recognised by the WHO some 30 years ago. Here, we performed an extensive genetic characterization of 978 human and 1440 animal isolates, which together comprise 3886 sequences from 4 genetic loci. The data were assembled into a molecular epidemiological database developed by a European network of public and veterinary health Institutions. Genotyping was performed at different levels of resolution (single and multiple loci on the same dataset. The zoonotic potential of both assemblages A and B is evident when studied at the level of assemblages, sub-assemblages, and even at each single locus. However, when genotypes are defined using a multi-locus sequence typing scheme, only 2 multi-locus genotypes (MLG of assemblage A and none of assemblage B appear to have a zoonotic potential. Surprisingly, mixtures of genotypes in individual isolates were repeatedly observed. Possible explanations are the uptake of genetically different Giardia cysts by a host, or subsequent infection of an already infected host, likely without overt symptoms, with a different Giardia species, which may cause disease. Other explanations for mixed genotypes, particularly for assemblage B, are substantial allelic sequence heterogeneity and/or genetic recombination. Although the

  18. Duplex detection of the Mycobacterium tuberculosis complex and medically important non-tuberculosis mycobacteria by real-time PCR based on the rnpB gene.

    Science.gov (United States)

    Abdeldaim, Guma; Svensson, Erik; Blomberg, Jonas; Herrmann, Björn

    2016-11-01

    A duplex real-time PCR based on the rnpB gene was developed for Mycobacterium spp. The assay was specific for the Mycobacterium tuberculosis complex (MTB) and also detected all 19 tested species of non-tuberculous mycobacteria (NTM). The assay was evaluated on 404 clinical samples: 290 respiratory samples and 114 from tissue and other non-respiratory body sites. M. tuberculosis was detected by culture in 40 samples and in 30 samples by the assay. The MTB assay showed a sensitivity similar to Roche Cobas Amplicor MTB-PCR (Roche Molecular Systems, Pleasanton, CA, USA). There were only nine samples with non-tuberculous mycobacteria detected by culture. Six of them were detected by the PCR assay. © 2016 APMIS. Published by John Wiley & Sons Ltd.

  19. Genotyping of African swine fever virus (ASFV) isolates associated ...

    African Journals Online (AJOL)

    Four of these viruses were isolated directly from serum samples. All the viruses were classified within the domesticpig cycle-associated p72 and p54 genotype IX which also includes viruses responsible for ASF outbreaks in Kenya in 2006 and 2007 and Uganda in 2003. To define virus relationships at higher resolution, ...

  20. Increasing Recovery of Nontuberculous Mycobacteria from Respiratory Specimens over a 10-Year Period in a Tertiary Referral Hospital in South Korea

    OpenAIRE

    Koh, Won-Jung; Chang, Boksoon; Jeong, Byeong-Ho; Jeon, Kyeongman; Kim, Su-Young; Lee, Nam Yong; Ki, Chang-Seok; Kwon, O Jung

    2013-01-01

    Background The number of patients with pulmonary disease caused by nontuberculous mycobacteria (NTM) has been increasing worldwide. The aim of this study was to evaluate long-term trends in the NTM recovery rate from respiratory specimens over a 10-year period in a tertiary referral hospital in South Korea. Methods We retrospectively reviewed the records of mycobacterial cultures of respiratory specimens at Samsung Medical Center from January 2001 to December 2011. Results During the study pe...

  1. Lipoproteins of slow-growing Mycobacteria carry three fatty acids and are N-acylated by apolipoprotein N-acyltransferase BCG_2070c.

    Science.gov (United States)

    Brülle, Juliane K; Tschumi, Andreas; Sander, Peter

    2013-10-05

    Lipoproteins are virulence factors of Mycobacterium tuberculosis. Bacterial lipoproteins are modified by the consecutive action of preprolipoprotein diacylglyceryl transferase (Lgt), prolipoprotein signal peptidase (LspA) and apolipoprotein N- acyltransferase (Lnt) leading to the formation of mature triacylated lipoproteins. Lnt homologues are found in Gram-negative and high GC-rich Gram-positive, but not in low GC-rich Gram-positive bacteria, although N-acylation is observed. In fast-growing Mycobacterium smegmatis, the molecular structure of the lipid modification of lipoproteins was resolved recently as a diacylglyceryl residue carrying ester-bound palmitic acid and ester-bound tuberculostearic acid and an additional amide-bound palmitic acid. We exploit the vaccine strain Mycobacterium bovis BCG as model organism to investigate lipoprotein modifications in slow-growing mycobacteria. Using Escherichia coli Lnt as a query in BLASTp search, we identified BCG_2070c and BCG_2279c as putative lnt genes in M. bovis BCG. Lipoproteins LprF, LpqH, LpqL and LppX were expressed in M. bovis BCG and BCG_2070c lnt knock-out mutant and lipid modifications were analyzed at molecular level by matrix-assisted laser desorption ionization time-of-flight/time-of-flight analysis. Lipoprotein N-acylation was observed in wildtype but not in BCG_2070c mutants. Lipoprotein N- acylation with palmitoyl and tuberculostearyl residues was observed. Lipoproteins are triacylated in slow-growing mycobacteria. BCG_2070c encodes a functional Lnt in M. bovis BCG. We identified mycobacteria-specific tuberculostearic acid as further substrate for N-acylation in slow-growing mycobacteria.

  2. Diversity, Community Composition, and Dynamics of Nonpigmented and Late-Pigmenting Rapidly Growing Mycobacteria in an Urban Tap Water Production and Distribution System

    OpenAIRE

    Dubrou, S.; Konjek, J.; Macheras, E.; Welté, B.; Guidicelli, L.; Chignon, E.; Joyeux, M.; Gaillard, J. L.; Heym, B.; Tully, T.; Sapriel, G.

    2013-01-01

    Nonpigmented and late-pigmenting rapidly growing mycobacteria (RGM) have been reported to commonly colonize water production and distribution systems. However, there is little information about the nature and distribution of RGM species within the different parts of such complex networks or about their clustering into specific RGM species communities. We conducted a large-scale survey between 2007 and 2009 in the Parisian urban tap water production and distribution system. We analyzed 1,418 w...

  3. Discrepancy between Hepatitis C Virus Genotypes and NS4-Based Serotypes: Association with Their Subgenomic Sequences

    Directory of Open Access Journals (Sweden)

    Nan Nwe Win

    2017-01-01

    Full Text Available Determination of hepatitis C virus (HCV genotypes plays an important role in the direct-acting agent era. Discrepancies between HCV genotyping and serotyping assays are occasionally observed. Eighteen samples with discrepant results between genotyping and serotyping methods were analyzed. HCV serotyping and genotyping were based on the HCV nonstructural 4 (NS4 region and 5′-untranslated region (5′-UTR, respectively. HCV core and NS4 regions were chosen to be sequenced and were compared with the genotyping and serotyping results. Deep sequencing was also performed for the corresponding HCV NS4 regions. Seventeen out of 18 discrepant samples could be sequenced by the Sanger method. Both HCV core and NS4 sequences were concordant with that of genotyping in the 5′-UTR in all 17 samples. In cloning analysis of the HCV NS4 region, there were several amino acid variations, but each sequence was much closer to the peptide with the same genotype. Deep sequencing revealed that minor clones with different subgenotypes existed in two of the 17 samples. Genotyping by genome amplification showed high consistency, while several false reactions were detected by serotyping. The deep sequencing method also provides accurate genotyping results and may be useful for analyzing discrepant cases. HCV genotyping should be correctly determined before antiviral treatment.

  4. Acid-fast Smear and Histopathology Results Provide Guidance for the Appropriate Use of Broad-Range Polymerase Chain Reaction and Sequencing for Mycobacteria.

    Science.gov (United States)

    Miller, Kennon; Harrington, Susan M; Procop, Gary W

    2015-08-01

    New molecular diagnostic tests are attractive because of the potential they hold for improving diagnostics in microbiology. The value of these tests, which is often assumed, should be investigated to determine the best use of these potentially powerful tools. To investigate the usefulness of broad-range polymerase chain reaction (PCR), followed by sequencing, in mycobacterial infections. We reviewed the test performance of acid-fast bacilli (AFB) PCR and traditional diagnostic methods (histopathology, AFB smear, and culture). We assessed the diagnostic effect and cost of the unrestricted ordering of broad-range PCR for the detection and identification of mycobacteria in clinical specimens. The AFB PCR was less sensitive than culture and histopathology and was less specific than culture, AFB smear, and histopathology. During 18 months, $93 063 was spent on 183 patient specimens for broad-range PCR and DNA sequencing for mycobacteria to confirm one culture-proven Mycobacterium tuberculosis infection that was also known to be positive by AFB smear and histopathology. In this cohort, there was a false-negative AFB PCR for M tuberculosis and a false-positive AFB PCR for Mycobacterium lentiflavum . Testing of AFB smear-negative specimens from patients without an inflammatory response supportive of a mycobacterial infection is costly and has not been proven to improve patient care. Traditional diagnostics (histopathology, AFB smear, and culture) should remain the primary methods for the detection of mycobacteria in clinical specimens.

  5. The Phenotype/Genotype Correlation of Lactase Persistence among Omani Adults

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    Abdulrahim Al-Abri

    2013-09-01

    Full Text Available Objective: To examine the correlation of lactase persistence phenotype with genotype in Omani adults.Methods: Lactase persistence phenotype was tested by hydrogen breath test in 52 Omani Adults using the Micro H2 analyzer. Results were checked against genotyping using direct DNA sequencing.Results: Forty one individuals with C/C-13910 and T/T-13915 genotypes had positive breath tests (≥20 ppm; while eight of nine individuals with T/C-13910 or T/G-13915 genotypes had negative breath tests (<20 ppm and two subjects were non-hydrogen producers. The agreement between phenotype and genotype using Kappa value was very good (0.93.Conclusion: Genotyping both T/C-13910 and T/G-13915 alleles can be used to assist diagnosis and predict lactose intolerance in the Omani population.

  6. Measurements of experimental precision for trials with cowpea (Vigna unguiculata L. Walp.) genotypes.

    Science.gov (United States)

    Teodoro, P E; Torres, F E; Santos, A D; Corrêa, A M; Nascimento, M; Barroso, L M A; Ceccon, G

    2016-05-09

    The aim of this study was to evaluate the suitability of statistics as experimental precision degree measures for trials with cowpea (Vigna unguiculata L. Walp.) genotypes. Cowpea genotype yields were evaluated in 29 trials conducted in Brazil between 2005 and 2012. The genotypes were evaluated with a randomized block design with four replications. Ten statistics that were estimated for each trial were compared using descriptive statistics, Pearson correlations, and path analysis. According to the class limits established, selective accuracy and F-test values for genotype, heritability, and the coefficient of determination adequately estimated the degree of experimental precision. Using these statistics, 86.21% of the trials had adequate experimental precision. Selective accuracy and the F-test values for genotype, heritability, and the coefficient of determination were directly related to each other, and were more suitable than the coefficient of variation and the least significant difference (by the Tukey test) to evaluate experimental precision in trials with cowpea genotypes.

  7. Disseminated mycobacteria chelonae infection in a kidney-pancreas transplant recipient: A case report and review of the literature

    Directory of Open Access Journals (Sweden)

    Shafi Malik

    2016-01-01

    Full Text Available A 40-year-old male with a long-standing history of type 1 diabetes with end-stage renal failure underwent combined kidney-pancreas (KP transplant from a standard criteria donor. Post-operative course was uncomplicated with good primary function of both transplant grafts. Induction was with thymoglobulin and maintenance immunosuppression was with tacrolimus, mycophenolate mofetil and prednisolone. Nine weeks post-transplant, the patient developed dysfunction of both grafts. Panel reactive antibody testing revealed that the patient had developed a de novo donor-specific antibody and considering an antibody-mediated rejection the patient was treated with intravenous pulse methyl prednisone 500 mg ×3 doses, IV immunoglobulin 2 mg/kg in two divided doses, and ATG 7 mg/kg (total dose of 700 mg. In addition, his baseline immunosuppression was increased. Cr decreased to baseline levels, and blood sugars were in the range of 7-8 mmol/L, serum amylase normalized to 63 U/L, and the patient was discharged home. Nine days post-discharge, the patient presented to the hospital with a five-day history of fever, pain, and swelling in the left knee along with subcutaneous, erythematous, tender, nodular lesions in both legs and both arms. Skin biopsy showed Ziehl-Neelsen stain positive rods and biopsy culture and blood culture grew Mycobacteria chelonae. Antimicrobials were switched to azithromycin 500 mg OD, moxifloxacin 400 mg OD, and linezolid 600 mg BID and baseline immunosuppression was reduced to tacrolimus trough target 8-10 ng/mL and MMF to 250 mg BID. The patient gradually improved and was discharged after 28 days in the hospital. Six weeks following the diagnosis of nontuberculous mycobacteria infection, the patient′s pancreas graft failed, presumably due to reduction in immuno-suppression and he is now back on insulin treatment. His renal graft continued to function well. Although rapidly growing mycobacterial infections are rare among transplant

  8. Counting mycobacteria in infected human cells and mouse tissue: a comparison between qPCR and CFU.

    Directory of Open Access Journals (Sweden)

    Sharad Pathak

    Full Text Available Due to the slow growth rate and pathogenicity of mycobacteria, enumeration by traditional reference methods like colony counting is notoriously time-consuming, inconvenient and biohazardous. Thus, novel methods that rapidly and reliably quantify mycobacteria are warranted in experimental models to facilitate basic research, development of vaccines and anti-mycobacterial drugs. In this study we have developed quantitative polymerase chain reaction (qPCR assays for simultaneous quantification of mycobacterial and host DNA in infected human macrophage cultures and in mouse tissues. The qPCR method cannot discriminate live from dead bacteria and found a 10- to 100-fold excess of mycobacterial genomes, relative to colony formation. However, good linear correlations were observed between viable colony counts and qPCR results from infected macrophage cultures (Pearson correlation coefficient [r] for M. tuberculosis = 0.82; M. a. avium = 0.95; M. a. paratuberculosis = 0.91. Regression models that predict colony counts from qPCR data in infected macrophages were validated empirically and showed a high degree of agreement with observed counts. Similar correlation results were also obtained in liver and spleen homogenates of M. a. avium infected mice, although the correlations were distinct for the early phase (< day 9 post-infection and later phase (≥ day 20 post-infection liver r = 0.94 and r = 0.91; spleen r = 0.91 and r = 0.87, respectively. Interestingly, in the mouse model the number of live bacteria as determined by colony counts constituted a much higher proportion of the total genomic qPCR count in the early phase (geometric mean ratio of 0.37 and 0.34 in spleen and liver, respectively, as compared to later phase of infection (geometric mean ratio of 0.01 in both spleen and liver. Overall, qPCR methods offer advantages in biosafety, time-saving, assay range and reproducibility compared to colony counting. Additionally, the duplex format allows

  9. FTO genotype and weight loss

    DEFF Research Database (Denmark)

    Livingstone, Katherine M; Celis-Morales, Carlos; Papandonatos, George D

    2016-01-01

    : Ovid Medline, Scopus, Embase, and Cochrane from inception to November 2015. ELIGIBILITY CRITERIA FOR STUDY SELECTION: Randomised controlled trials in overweight or obese adults reporting reduction in body mass index, body weight, or waist circumference by FTO genotype (rs9939609 or a proxy) after...

  10. FTO genotype and weight loss

    DEFF Research Database (Denmark)

    Livingstone, Katherine M; Celis-Morales, Carlos; Papandonatos, George D

    2016-01-01

    OBJECTIVE: To assess the effect of the FTO genotype on weight loss after dietary, physical activity, or drug based interventions in randomised controlled trials. DESIGN: Systematic review and random effects meta-analysis of individual participant data from randomised controlled trials. DATA SOURC...

  11. Distribution of Human papilloma virus genotypes in cervical cancer tissues

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    Stamenković M.

    2014-01-01

    Full Text Available Cervical cancer incidence and mortality rates in Serbia are among the highest in Europe and data on Human papilloma virus (HPV type distribution are scarce. The aim of this study was to determine the prevalence of HPV types in archival specimens of cervical cancer tissues of women in the Serbian population. A total of 45 paraffin-embedded tissue samples of cervical carcinoma were used in this study. The procedure included deparaffinization of tissue samples, DNA extraction, PCR, gel electrophoresis and HPV genotyping by direct sequencing. HPV was detected in 32 samples (71%. Genotyping revealed the presence of 6 high-risk HPV types 16, 18, 33, 45, 53 and 58, where HPV type 16 was the most prevalent type (73.7%. The results of this study and further studies will provide more detailed information about HPV genotype distribution and may contribute to the formulation of national guidelines for the prevention of cervical cancer. [175073

  12. Methods for MHC genotyping in non-model vertebrates.

    Science.gov (United States)

    Babik, W

    2010-03-01

    Genes of the major histocompatibility complex (MHC) are considered a paradigm of adaptive evolution at the molecular level and as such are frequently investigated by evolutionary biologists and ecologists. Accurate genotyping is essential for understanding of the role that MHC variation plays in natural populations, but may be extremely challenging. Here, I discuss the DNA-based methods currently used for genotyping MHC in non-model vertebrates, as well as techniques likely to find widespread use in the future. I also highlight the aspects of MHC structure that are relevant for genotyping, and detail the challenges posed by the complex genomic organization and high sequence variation of MHC loci. Special emphasis is placed on designing appropriate PCR primers, accounting for artefacts and the problem of genotyping alleles from multiple, co-amplifying loci, a strategy which is frequently necessary due to the structure of the MHC. The suitability of typing techniques is compared in various research situations, strategies for efficient genotyping are discussed and areas of likely progress in future are identified. This review addresses the well established typing methods such as the Single Strand Conformation Polymorphism (SSCP), Denaturing Gradient Gel Electrophoresis (DGGE), Reference Strand Conformational Analysis (RSCA) and cloning of PCR products. In addition, it includes the intriguing possibility of direct amplicon sequencing followed by the computational inference of alleles and also next generation sequencing (NGS) technologies; the latter technique may, in the future, find widespread use in typing complex multilocus MHC systems. © 2009 Blackwell Publishing Ltd.

  13. Hepatitis C Virus: Virology and Genotypes

    KAUST Repository

    Abdelaziz, Ahmed

    2017-01-01

    Hepatitis C virus (HCV) is a major causative agent of chronic liver disease worldwide. HCV is characterized by genetic heterogeneity, with at least six genotypes identified. The geographic distribution of genotypes has shown variations in different

  14. Acidic methanolysis v. alkaline saponification in gas chromatographic characterization of mycobacteria: differentiation between Mycobacterium avium-intracellulare and Mycobacterium gastri.

    Science.gov (United States)

    Larsson, L

    1983-08-01

    Mycobacterium avium-intracellulare and M.gastri were analyzed with capillary gas chromatography after each strain had been subjected to acidic methanolysis or to alkaline saponification followed by methylation. Prominent peaks of myristic, palmitoleic, palmitic, oleic, stearic and tuberculostearic acids were found in the chromatograms of both species, whereas 2-octadecanol and 2-eicosanol were detected only in M. avium-intracellulare. In initial runs, both of the derivatization principles yielded virtually identical chromatograms for a given strain. After repeated injections of extracts from alkaline saponification, however, the alcohol peaks showed pronounced tailing and finally almost disappeared from the chromatograms. This disadvantage, which was not observed when only acid methanolysis was used, could be overcome with trifluoroacetylation. Restored peak shape of the underivatized alcohols could be achieved by washing the cross-linked stationary phase in the capillary tubing with organic solvents. The study demonstrated the importance of conditions which enable separation of 2-octadecanol and 2-eicosanol when gas chromatography is used for species identification of mycobacteria.

  15. Comparison of manual mycobacteria growth indicator tube and epsilometer test with agar proportion method for susceptibility testing of Mycobacterium tuberculosis

    Directory of Open Access Journals (Sweden)

    N Karabulut

    2014-01-01

    Full Text Available Background and Objectives: Antimycobacterial susceptibility tests take weeks, and delayed therapy can lead to spread of Mycobacterium tuberculosis. Therefore, rapid, accurate and cost-effective methods are required for proper therapy selection. In this study, the Mycobacteria growth indicator tube (MGIT and epsilometer test (Etest methods were compared to the agar proportion method for susceptibility testing of Mycobacterium tuberculosis. Materials and Methods: The susceptibility tests against isoniazid (INH, rifampin (RIF, streptomycin (STM and ethambutol (ETM of 51 M. tuberculosis complex isolates were analyzed by the MGIT, Etest and agar proportion methods. Results: The concordance between MGIT/Etest and agar proportion methods was 98% for INH and 100% for RIF, STM, ETM. There were not statistically significant differences in results of the susceptibility tests between MGIT/Etest and the reference agar proportion method. Conclusion: The results have shown that MGIT and Etest methods can be used instead of the agar proportion method, because these two methods are more rapid and easier than the agar proportion method.

  16. The epidemiology of pulmonary nontuberculous mycobacteria: data from a general hospital in Athens, Greece, 2007-2013.

    Science.gov (United States)

    Panagiotou, Marios; Papaioannou, Andriana I; Kostikas, Konstantinos; Paraskeua, Maria; Velentza, Ekaterini; Kanellopoulou, Maria; Filaditaki, Vasiliki; Karagiannidis, Napoleon

    2014-01-01

    The epidemiology of pulmonary nontuberculous mycobacteria (NTM) in Greece is largely unknown. To determine the incidence and the demographic, microbiological, and clinical characteristics of patients with pulmonary NTM infection and pulmonary NTM disease. A retrospective review of the demographic, microbiological, and clinical characteristics of patients with NTM culture-positive respiratory specimens from January 2007 to May 2013. A total of 120 patients were identified with at least one respiratory NTM isolate and 56 patients (46%) fulfilled the microbiological ATS/IDSA criteria for NTM disease. Of patients with adequate data, 16% fulfilled the complete ATS/IDSA criteria for NTM disease. The incidence of pulmonary NTM infection and disease was 18.9 and 8.8 per 100.000 inpatients and outpatients, respectively. The spectrum of NTM species was high (13 species) and predominated by M. avium-intracellulare complex (M. avium (13%), M. intracellulare (10%)), M. gordonae (14%), and M. fortuitum (12%). The ratio of isolation of NTM to M. tuberculosis in all hospitalized patients was 0.59. The first data on the epidemiology of pulmonary NTM in Athens, Greece, are presented. NTM infection is common in patients with chronic respiratory disease. However, only a significantly smaller proportion of patients fulfill the criteria for NTM disease.

  17. The Epidemiology of Pulmonary Nontuberculous Mycobacteria: Data from a General Hospital in Athens, Greece, 2007–2013

    Science.gov (United States)

    Papaioannou, Andriana I.; Paraskeua, Maria; Velentza, Ekaterini; Kanellopoulou, Maria; Filaditaki, Vasiliki; Karagiannidis, Napoleon

    2014-01-01

    Background. The epidemiology of pulmonary nontuberculous mycobacteria (NTM) in Greece is largely unknown. Objectives. To determine the incidence and the demographic, microbiological, and clinical characteristics of patients with pulmonary NTM infection and pulmonary NTM disease. Methods. A retrospective review of the demographic, microbiological, and clinical characteristics of patients with NTM culture-positive respiratory specimens from January 2007 to May 2013. Results. A total of 120 patients were identified with at least one respiratory NTM isolate and 56 patients (46%) fulfilled the microbiological ATS/IDSA criteria for NTM disease. Of patients with adequate data, 16% fulfilled the complete ATS/IDSA criteria for NTM disease. The incidence of pulmonary NTM infection and disease was 18.9 and 8.8 per 100.000 inpatients and outpatients, respectively. The spectrum of NTM species was high (13 species) and predominated by M. avium-intracellulare complex (M. avium (13%), M. intracellulare (10%)), M. gordonae (14%), and M. fortuitum (12%). The ratio of isolation of NTM to M. tuberculosis in all hospitalized patients was 0.59. Conclusions. The first data on the epidemiology of pulmonary NTM in Athens, Greece, are presented. NTM infection is common in patients with chronic respiratory disease. However, only a significantly smaller proportion of patients fulfill the criteria for NTM disease. PMID:25132991

  18. Clinical Relevance of Nontuberculous Mycobacteria Isolated from Sputum in a Gold Mining Workforce in South Africa: An Observational, Clinical Study

    Directory of Open Access Journals (Sweden)

    Clare L. van Halsema

    2015-01-01

    Full Text Available Background. The clinical relevance of nontuberculous mycobacteria (NTM, detected by liquid more than solid culture in sputum specimens from a South African mining workforce, is uncertain. We aimed to describe the current spectrum and relevance of NTM in this population. Methods. An observational study including individuals with sputum NTM isolates, recruited at workforce tuberculosis screening and routine clinics. Symptom questionnaires were administered at the time of sputum collection and clinical records and chest radiographs reviewed retrospectively. Results. Of 232 individuals included (228 (98% male, median age 44 years, M. gordonae (60 individuals, M. kansasii (50, and M. avium complex (MAC: 38 were the commonest species. Of 38 MAC isolates, only 2 (5.3% were from smear-positive sputum specimens and 30/38 grew in liquid but not solid culture. MAC was especially prevalent among symptomatic, HIV-positive individuals. HIV prevalence was high: 57/74 (77% among those tested. No differences were found in probability of death or medical separation by NTM species. Conclusions. M. gordonae, M. kansasii, and MAC were the commonest NTM among miners with suspected tuberculosis, with most MAC from smear-negative specimens in liquid culture only. HIV testing and identification of key pathogenic NTM in this setting are essential to ensure optimal treatment.

  19. THE CELLS WITH MYCOBACTERIA IN GRANULOMATOUS AGGREGATES FROM MICE WITH LATENT TUBERCULOUS INFECTION IN EX VIVO CULTURE

    Directory of Open Access Journals (Sweden)

    E. G. Ufimtseva

    2013-01-01

    Full Text Available Abstract. The aim of this study was to obtain ex vivo monolayer culture cells migrated from individual granulomas isolated from the spleens of the Balb/c line mice through 1–2 months after BCG vaccine infection. The second goal was to evaluate influence of different types of cells in the development of granulomatic inflammation and analysis of BCG bacteria content in these cells in the latent stage of tuberculosis. Granulomas were presented by macrophages in general. The number of granulomas was varied as in one mouse as between mice. Granulomas contained also dendritic cells (in average 10% from macrophages of granulomas and lymphocytes. In some granulomas fibroblasts, neutrophils, eosiniphils, multinuclear cells of Pirogov–Langhans, megacariocytes and platelets were observed in all stages of infection. The number of these cells was also varied between granulomas. The acid staining BCG bacteria were only detected in macrophages, dendritic cells and Pirogov–Langhans cells of mice granulomas. Mice were different as by number of cells with BCG bacteria in granulomas as by number of granulomas with BCG-containing cells. The proposed model of granuloma cells of mice in ex vivo culture can be used to study interaction between host cells and mycobacteria to find new ways and methods of influence to intracellular pathogens in latent stage of tuberculosis. 

  20. The study of correlation between HBV genotype and the response to transcatheter arterial chemoembolization therapy in hepatocellular carcinoma patients

    International Nuclear Information System (INIS)

    Huang Keyao; Yang Weizhu; Jiang Na; Zheng Qubing

    2004-01-01

    Objective: To evaluate the influence of hepatitis B virus(HBV) genotype on response to transcatheter arterial embolization therapy in patients with HBV-related HCC. Methods: Transcatheter arterial chemoem-bolization therapy was conducted in patients with HBV-related HCC and response to embolization therapy were observed according to the tumor necrosis rate, the HCC recurrence rate, the cumulative incidence of survival rate and the change of AFP. The HBV genotype was determined by sequencing directly the polymerase chain reaction products of the HBV S gene. The response of HCC to embolization therapy was compared between patients who were infected with different genotypic HBV. Results: The tumor necrosis rate of genotype C patients was similar to that of genotype B patients (P=0.099). The HCC recurrence rate of genotype B was lower than that of genotype C patients (P=0.036). The cumulative incidence of survival rates of 2 and 3 years were significantly higher in the genotype B patients (P=0.036 and P=0.013). There was no difference between the two genotypes, patients in the change of AFP (P>0.05). Conclusions: HBV genotype B patients seem to have a better response to embolization therapy as compared to genotype C patients. Determination of HBV genotype may be useful in predicting the outcomes of TACE therapy in HBV-related HCC. (authors)

  1. Genotype x environment interaction and optimum resource ...

    African Journals Online (AJOL)

    ... x E) interaction and to determine the optimum resource allocation for cassava yield trials. The effects of environment, genotype and G x E interaction were highly significant for all yield traits. Variations due to G x E interaction were greater than those due to genotypic differences for all yield traits. Genotype x location x year ...

  2. Identification of Zoonotic Genotypes of Giardia duodenalis

    DEFF Research Database (Denmark)

    Sprong, H.; Cacciò, S.M.; van der Giessen, J.W.B

    2009-01-01

    Giardia duodenalis, originally regarded as a commensal organism, is the etiologic agent of giardiasis, a gastrointestinal disease of humans and animals. Giardiasis causes major public and veterinary health concerns worldwide. Transmission is either direct, through the faecal-oral route, or indirect......, through ingestion of contaminated water or food. Genetic characterization of G. duodenalis isolates has revealed the existence of seven groups (assemblages A to G) which differ in their host distribution. Assemblages A and B are found in humans and in many other mammals, but the role of animals......, mixtures of genotypes in individual isolates were repeatedly observed. Possible explanations are the uptake of genetically different Giardia cysts by a host, or subsequent infection of an already infected host, likely without overt symptoms, with a different Giardia species, which may cause disease. Other...

  3. Genetic Divergence in Sugarcane Genotypes

    OpenAIRE

    Tahir, Mohammad; Rahman, Hidayatur; Gul, Rahmani; Ali, Amjad; Khalid, Muhammad

    2012-01-01

    To assess genetic divergence of sugarcane germplasm, an experiment comprising 25 sugarcane genotypes was conducted at Sugar Crops Research Institute (SCRI), Mardan, Khyber Pakhtunkhwa, Pakistan, in quadruple lattice design during 2008-09. Among the 14 parameters evaluated, majority exhibited significant differences while some showed nonsignificant mean squares. The initial correlation matrix revealed medium to high correlations. Principal Component Analysis (PCA) showed that there were two pr...

  4. Most of the benefits from genomic selection can be realised by genotyping a proportion of selection candidates

    DEFF Research Database (Denmark)

    Henryon, Mark; Berg, Peer; Sørensen, Anders Christian

    2012-01-01

    allocated to male and female candidates at ratios of 100:0, 75:25, 50:50, 25:75, and 0:100. For genotyped candidates, a direct-genomic value (DGV) was sampled with reliabilities 0.10, 0.50, and 0.90. Ten sires and 300 dams with the highest breeding values after genotyping were selected at each generation......We reasoned that there are diminishing marginal returns from genomic selection as the proportion of genotyped selection candidates is increased and breeding values based on a priori information are used to choose the candidates that are genotyped. We tested this premise by stochastic simulation...... of breeding schemes that resembled those used for pigs. We estimated rates of genetic gain and inbreeding realized by genomic selection in breeding schemes where candidates were phenotyped before genotyping and 0-100% of the candidates were genotyped based on predicted breeding values. Genotypings were...

  5. Disease caused by non-tuberculous mycobacteria: diagnostic procedures and treatment evaluation in the North of Buenos Aires Province

    Directory of Open Access Journals (Sweden)

    Belén Imperiale

    2012-03-01

    Full Text Available Non-tuberculous mycobacteria (NTM have emerged as pathogens frequently associated to HIV co-infection. The aims of this study were to describe the clinical importance of NTM in patients from the North of Buenos Aires Province and the drug-susceptibility patterns in relation with the therapy used. A total of 23,624 clinical specimens were investigated during the period 2004-2010. Ziehl-Neelsen stain and cultures were used for diagnosis. Molecular and biochemical tests were performed to identify the mycobacteria. TB and mycobacterioses cases were 2 118 and 108 respectively. Sixteen NTM species were found: Mycobacterium avium and Mycobacterium intracellulare as the main causative agents. Infections produced by more than one species at the same time were confirmed (4 cases. Macrolides and fluoroquinolones were the most active in vitro drugs. Treatment evaluation showed that 68.0 % of the cases completed the therapy, 20 % died; and 12 % were relapses. The cases in which the treatment outcome was evaluated received an individual tailor-made therapeutic scheme including those drugs showing in vitro activity and presumed in vivo usefulness. More than a quarter of the patients had HIV co-infection and the majority of the deaths were associated with this co-infection.Enfermedad causada por micobacterias no tuberculosas: diagnóstico y evaluación del tratamiento en el norte del Gran Buenos Aires. Las micobacterias no tuberculosas (MNT emergieron como patógenos frecuentemente asociados a la co-infección con el HIV. EL objetivo del estudio fue describir la importancia clínica de las MNT en pacientes de la región norte de la provincia de Buenos Aires y los patrones de drogo-sensibilidad en relación con la terapia empleada. Se investigó un total de 23.624 especímenes clínicos durante, el período 2004-2010. La tinción de Ziehl-Neelsen y los cultivos se utilizaron para diagnóstico. Las micobacterias fueron identificadas mediante pruebas bioqu

  6. [Nicotinamidase and the so-called pyrazinamidase in mycobacteria; the simultaneous occurrence of both activites (author's transl)].

    Science.gov (United States)

    Tarnok, I; Pechmann, H; Krallmann-Wenzel, U; Röhrscheidt, E; Tarnok, Z

    1979-07-01

    Nicotin- and the so-called pyrazinamidase (in the following: "pyrazinamidase") have been found in strains of four mycobacteria species, M. fortuitum, M. gastri, M. bovis and M. microti. These findings are in contradiction to those summarized in Bergey's Manual of Determinative Bacteriology (1974). The reason for the discrepancies is that the original method (Bönicke, 1961) for amidase determination has not taken the following aspects into consideration: a) The inducibility of the nicotin- and "pyrazinamidase" (example: M. fortuitum); b) The temperature sensitivity of these enzymes (M. gastri); c) The light sensitivity of nicotinamidase (in photochromogenic M. gastri strains); d) The optimal substrate concentration which must be at least 4 mM instead of 0,8 mm. The following consequences can be drawn for the taxonomy and biochemistry of the tested organisms: e) The species status of M. gastri should be annuled. The main difference between M. gastri and M. kansasii consists only of the non-agglutinability of M. gastri by anti-M. kansasii serum. "Pyrazinamidase" and also nitrate reductase (Tarnok et al., in press) are positive in strains of both species; f) M. bovis possesses nicotin- and "pyrazinamidase" as M. tuberculosis too. Thus, these two species are more closely related than suggested earlier; g) Till now, no Mycobacterium has been found showing nicotinamidase without "pyrazinamidase" activity (or vice versa). It seems to be very probable that nicotinamidase, an enzyme of low substrate specificity, is able to hydrolyze several compounds with a nicotinamide-like structure such as pyrazinamide. Thus, we suggest the annulment of the term pyrazinamidase or the employment of quotation marks ("pyrazinamidase") to show the fictitious value of this designation.

  7. Prevalence and species spectrum of both pulmonary and extrapulmonary nontuberculous mycobacteria isolates at a tertiary care center

    Directory of Open Access Journals (Sweden)

    Jyoti Umrao

    2016-01-01

    Full Text Available Objective/background: Nontuberculous mycobacteria (NTM infection associated with pulmonary and extrapulmonary disease has been increasing globally. Despite an increase in incidence rate of NTM infection, its prevalence, species diversity, and circulation pattern in India is largely unknown. This study sought to investigate the overall burden and diversity of NTM among both pulmonary and extrapulmonary clinical isolates from a Northern Indian population. Methods: The study was conducted in the Department of Microbiology, from January 2013 to December 2015. A total of 4620 clinical samples were collected from patients suspected to have pulmonary and extrapulmonary tuberculosis. Preliminary diagnosis was performed using Ziehl–Neelsen staining followed by liquid culture in BacT/ALERT three-dimensional system. A total of 906 positive cultures obtained were differentiated as either NTM or Mycobacterium tuberculosis complex using a biochemical and MPT64 antigen test. Further identification of NTM species was confirmed with a line probe assay. Results: Out of 906 cultures isolates, 263 (29.0% were confirmed as NTM and 643 (71.0% were identified as Mycobacterium tuberculosis complex. A total of 79.4% of the NTM were recovered from pulmonary and 18.2% from extrapulmonary specimens. The diversity of NTM species was high (13 species and predominated by Mycobacterium abscessus (31.3% followed by Mycobacterium fortuitum (22%, Mycobacterium intracellulare (13.6%, Mycobacterium chelonae (9.1%, however, M. abscessus and M. fortuitum were the predominant species in both types of clinical isolates. Men (60.4% and older patients aged greater than 55 years were the predominated risk group for NTM infection. Conclusion: The high prevalence and species diversity of NTM suggests the need for immediate and accurate characterization of NTM for proper treatment and management of patients.

  8. Comparison of clinico-radiological features of patients with positive cultures of nontuberculous mycobacteria and patients with tuberculosis

    International Nuclear Information System (INIS)

    Ba-Hammam, Ahmed; Sharif, Yasir; Masood, Mohammad; Isnani, Arthur; Youssef, Ismael; Kambal, Abdelmageed; Shaikh, Shaffi

    2005-01-01

    To identify the clinico-radiological features of patients with positive cultures for nontuberculous mycobacteria (NTM) and compare those to a sample of patients with tuberculosis (MTB). A laboratory database was used to retrieve all specimens submitted to King Khalid University Hospital, Riyadh, mycobacteriology laboratory for mycobacterial smears and cultures during the period from October 1999-April 2002. Using this database, the original records of the mycobacteriology laboratory and a review of the patient's health records, a standard proforma was completed that included demographic, clinical, radiological and laboratory information on patients included in this study. The patients were divided into 2 groups; the NTM group, which included patients with positive cultures for NTM and the MTB group, which included a sample of patients with documented tuberculosis. During the study period, 286 patients had positive mycobacterial cultures. Seventy patients (24.5%) grew NTM and 216 (75.5%) grew MTB. For patients with MTB, 54 patients were included as per the selection protocol of the study. There was no difference between the 2 groups in all measured demographic variables. The presence of weight loss and fever was significantly more in the MTB group. Radiologically, the presence of hilar adenopathy was more significant among patients with MTB than those with NTM (17% versus 4%, p=0.02). However, bronchiectatic changes were seen significantly more among NTM patients compared to patients with MTB (26% versus 11%, p=0.03). The isolation of NTM in the mycobacteriology laboratory is high. The clinico-radiological features were not sufficiently specific to differentiate patients with NTM from patients with MTB. Local studies are needed to explore NTM disease in various developing countries and identify the NTM species causing infections in non-immunosuppressed patients in each locality. (author)

  9. 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.

  10. Insulin resistance and liver steatosis in chronic hepatitis C infection genotype 3.

    Science.gov (United States)

    Abenavoli, Ludovico; Masarone, Mario; Peta, Valentina; Milic, Natasa; Kobyliak, Nazarii; Rouabhia, Samir; Persico, Marcello

    2014-11-07

    Hepatitis C virus (HCV) infection is a common chronic liver disease worldwide. Non-alcoholic fatty liver disease and insulin resistance (IR) are the major determinants of fibrosis progression and response to antiviral therapy. The pathogenetic link between IR and chronic HCV infection is complex, and is associated with HCV genotype. Liver steatosis is the most common in the patients infected with genotype 3 virus, possibly due to direct effects of genotype 3 viral proteins. To the contrary, hepatic steatosis in the patients infected with other genotypes is thought to be mostly due to the changes in host metabolism, involving IR. In HCV genotype 3, liver steatosis correlates with viral load, reverts after reaching the sustained virologic response and reoccurs in the relapsers. A therapeutic strategy to improve IR and liver steatosis and subsequently the response to antiviral treatment in these patients is warranted.

  11. 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.

  12. Norovirus Genotype Profiles Associated with Foodborne Transmission, 1999??"2012

    OpenAIRE

    Verhoef, Linda; Hewitt, Joanne; Barclay, Leslie; Ahmed, Sharia; Lake, Rob; Hall, Aron J.; Lopman, Ben; Kroneman, Annelies; Vennema, Harry; VinjA(c), Jan; Koopmans, Marion

    2015-01-01

    textabstractWorldwide, noroviruses are a leading cause of gastroenteritis. They can be transmitted from person to person directly or indirectly through contaminated food, water, or environments. To estimate the proportion of foodborne infections caused by noroviruses on a global scale, we used norovirus transmission and genotyping information from multiple international outbreak surveillance systems (Noronet, CaliciNet, EpiSurv) and from a systematic review of peer-reviewed literature. The pr...

  13. [Analytic methods for seed models with genotype x environment interactions].

    Science.gov (United States)

    Zhu, J

    1996-01-01

    Genetic models with genotype effect (G) and genotype x environment interaction effect (GE) are proposed for analyzing generation means of seed quantitative traits in crops. The total genetic effect (G) is partitioned into seed direct genetic effect (G0), cytoplasm genetic of effect (C), and maternal plant genetic effect (Gm). Seed direct genetic effect (G0) can be further partitioned into direct additive (A) and direct dominance (D) genetic components. Maternal genetic effect (Gm) can also be partitioned into maternal additive (Am) and maternal dominance (Dm) genetic components. The total genotype x environment interaction effect (GE) can also be partitioned into direct genetic by environment interaction effect (G0E), cytoplasm genetic by environment interaction effect (CE), and maternal genetic by environment interaction effect (GmE). G0E can be partitioned into direct additive by environment interaction (AE) and direct dominance by environment interaction (DE) genetic components. GmE can also be partitioned into maternal additive by environment interaction (AmE) and maternal dominance by environment interaction (DmE) genetic components. Partitions of genetic components are listed for parent, F1, F2 and backcrosses. A set of parents, their reciprocal F1 and F2 seeds is applicable for efficient analysis of seed quantitative traits. MINQUE(0/1) method can be used for estimating variance and covariance components. Unbiased estimation for covariance components between two traits can also be obtained by the MINQUE(0/1) method. Random genetic effects in seed models are predictable by the Adjusted Unbiased Prediction (AUP) approach with MINQUE(0/1) method. The jackknife procedure is suggested for estimation of sampling variances of estimated variance and covariance components and of predicted genetic effects, which can be further used in a t-test for parameter. Unbiasedness and efficiency for estimating variance components and predicting genetic effects are tested by

  14. Applications of blood group genotyping

    Directory of Open Access Journals (Sweden)

    Mariza A. Mota

    2006-03-01

    Full Text Available Introduction: The determination of blood group polymorphism atthe genomic level facilitates the resolution of clinical problemsthat cannot be addressed by hemagglutination. They are useful to(a determine antigen types for which currently available antibodiesare weakly reactive; (b type patients who have been recentlytransfused; (c identify fetuses at risk for hemolytic disease of thenewborn; and (d to increase the reliability of repositories of antigennegative RBCs for transfusion. Objectives: This review assessedthe current applications of blood group genotyping in transfusionmedicine and hemolytic disease of the newborn. Search strategy:Blood group genotyping studies and reviews were searched ingeneral database (MEDLINE and references were reviewed.Selection criteria: All published data and reviews were eligible forinclusion provided they reported results for molecular basis ofblood group antigens, DNA analysis for blood group polymorphisms,determination of fetal group status and applications of blood groupgenotyping in blood transfusion. Data collection: All data werecollected based on studies and reviews of blood grouppolymorphisms and their clinical applications.

  15. Grain yield stability of early maize genotypes

    Directory of Open Access Journals (Sweden)

    Chitra Bahadur Kunwar

    2016-12-01

    Full Text Available The objective of this study was to estimate grain yield stability of early maize genotypes. Five early maize genotypes namely Pool-17, Arun1EV, Arun-4, Arun-2 and Farmer’s variety were evaluated using Randomized Complete Block Design along with three replications at four different locations namely Rampur, Rajahar, Pakhribas and Kabre districts of Nepal during summer seasons of three consecutive years from 2010 to 2012 under farmer’s fields. Genotype and genotype × environment (GGE biplot was used to identify superior genotype for grain yield and stability pattern. The genotypes Arun-1 EV and Arun-4 were better adapted for Kabre and Pakhribas where as pool-17 for Rajahar environments. The overall findings showed that Arun-1EV was more stable followed by Arun-2 therefore these two varieties can be recommended to farmers for cultivation in both environments.

  16. 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.

  17. Hepatitis C Virus: Virology and Genotypes

    KAUST Repository

    Abdelaziz, Ahmed

    2017-12-01

    Hepatitis C virus (HCV) is a major causative agent of chronic liver disease worldwide. HCV is characterized by genetic heterogeneity, with at least six genotypes identified. The geographic distribution of genotypes has shown variations in different parts of the world over the past decade because of variations in population structure, immigration, and routes of transmission. Genotype differences are of epidemiologic interest and help the study of viral transmission dynamics to trace the source of HCV infection in a given population. HCV genotypes are also of considerable clinical importance because they affect response to antiviral therapy and represent a challenging obstacle for vaccine development.

  18. Robust HCV Genotype 3a Infectious Cell Culture System Permits Identification of Escape Variants With Resistance to Sofosbuvir

    DEFF Research Database (Denmark)

    Ramirez Almeida, Santseharay; Mikkelsen, Lotte S.; Gottwein, Judith M.

    2016-01-01

    Background & Aims Direct-acting antivirals (DAAs) effectively eradicate chronic hepatitis C virus (HCV) infection, although HCV genotype 3a is less responsive to these drugs. We aimed to develop genotype 3a infectious cultures and study the effects of inhibitors of NS5A and NS5B and resistance to...

  19. Increasing Recovery of Nontuberculous Mycobacteria from Respiratory Specimens over a 10-Year Period in a Tertiary Referral Hospital in South Korea.

    Science.gov (United States)

    Koh, Won-Jung; Chang, Boksoon; Jeong, Byeong-Ho; Jeon, Kyeongman; Kim, Su-Young; Lee, Nam Yong; Ki, Chang-Seok; Kwon, O Jung

    2013-11-01

    The number of patients with pulmonary disease caused by nontuberculous mycobacteria (NTM) has been increasing worldwide. The aim of this study was to evaluate long-term trends in the NTM recovery rate from respiratory specimens over a 10-year period in a tertiary referral hospital in South Korea. We retrospectively reviewed the records of mycobacterial cultures of respiratory specimens at Samsung Medical Center from January 2001 to December 2011. During the study period, 32,841 respiratory specimens from 10,563 patients were found to be culture-positive for mycobacteria. These included 12,619 (38%) Mycobacterium tuberculosis and 20,222 (62%) NTM isolates. The proportion of NTM among all positive mycobacterial cultures increased from 43% (548/1,283) in 2001 to 70% (3,341/4,800) in 2011 (ptrend). The recovery rate of NTM isolates from acid-fast bacilli smear-positive specimens increased from 9% (38/417) in 2001 to 64% (1,284/1,997) in 2011 (ptrend). The proportion of positive liquid cultures was higher for NTM than for M. tuberculosis (p<0.001). The most frequently isolated NTM were Mycobacterium avium-intracellulare complex (53%) and Mycobacterium abscessus-massiliense complex (25%). The recovery rate of NTM from respiratory specimens in South Korea has increased steadily.

  20. Occurrence of Mycobacterium bovis and non-tuberculous mycobacteria (NTM) in raw and pasteurized milk in the northwestern region of Paraná, Brazil.

    Science.gov (United States)

    Sgarioni, Sônia Aparecida; Hirata, Rosario Dominguez Crespo; Hirata, Mario Hiroyuki; Leite, Clarice Queico Fujimura; de Prince, Karina Andrade; de Andrade Leite, Sergio Roberto; Filho, Dirceu Vedovello; Siqueira, Vera Lucia Dias; Caleffi-Ferracioli, Katiany Rizzieri; Cardoso, Rosilene Fressatti

    2014-01-01

    Milk is widely consumed in Brazil and can be the vehicle of agent transmission. In this study, was evaluated the occurrence of Mycobacterium bovis and non-tuberculous mycobacteria (NTM) in raw and pasteurized milk consumed in the northwestern region of Paraná, Brazil. Fifty-two milk samples (20 pasteurized and 32 raw) from dairy farms near the municipality of Maringa, Parana State, Brazil were collected. Milk samples were decontaminated using 5% oxalic acid method and cultured on Lowenstein-Jensen and Stonebrink media at 35 °C and 30 °C, with and without 5-10% CO2. Mycobacteria isolates were identified by morphological features, PCR-Restriction Fragment Length Polymorphism Analysis (PCR-PRA) and Mycolic acids analysis. Thirteen (25%) raw and 2 (4%) pasteurized milk samples were positive for acid fast bacilli growth. Nine different species of NTM were isolated (M. nonchromogenicum, M. peregrinum, M. smegmatis, M. neoaurum, M. fortuitum, M. chelonae, M. flavescens, M. kansasii and M. scrofulaceum). M. bovis was not detected. Raw and pasteurized milk may be considered one source for NTM human infection. The paper reinforces the need for intensification of measures in order to avoid the milk contamination and consequently prevent diseases in the south of Brazil.

  1. Occurrence of Mycobacterium bovis and non-tuberculous mycobacteria (NTM in raw and pasteurized milk in the northwestern region of Paraná, Brazil

    Directory of Open Access Journals (Sweden)

    Sônia Aparecida Sgarioni

    2014-06-01

    Full Text Available Milk is widely consumed in Brazil and can be the vehicle of agent transmission. In this study, was evaluated the occurrence of Mycobacterium bovis and non-tuberculous mycobacteria (NTM in raw and pasteurized milk consumed in the northwestern region of Paraná, Brazil. Fifty-two milk samples (20 pasteurized and 32 raw from dairy farms near the municipality of Maringa, Parana State, Brazil were collected. Milk samples were decontaminated using 5% oxalic acid method and cultured on Lowenstein-Jensen and Stonebrink media at 35 °C and 30 °C, with and without 5-10% CO2. Mycobacteria isolates were identified by morphological features, PCR-Restriction Fragment Length Polymorphism Analysis (PCR-PRA and Mycolic acids analysis. Thirteen (25% raw and 2 (4% pasteurized milk samples were positive for acid fast bacilli growth. Nine different species of NTM were isolated (M. nonchromogenicum, M. peregrinum, M. smegmatis, M. neoaurum, M. fortuitum, M. chelonae, M. flavescens, M. kansasii and M. scrofulaceum. M. bovis was not detected. Raw and pasteurized milk may be considered one source for NTM human infection. The paper reinforces the need for intensification of measures in order to avoid the milk contamination and consequently prevent diseases in the south of Brazil.

  2. Evaluation of the Abbott Real Time HCV genotype II assay for Hepatitis C virus genotyping.

    Science.gov (United States)

    Sariguzel, Fatma Mutlu; Berk, Elife; Gokahmetoglu, Selma; Ercal, Baris Derya; Celik, Ilhami

    2015-01-01

    The determination of HCV genotypes and subtypes is very important for the selection of antiviral therapy and epidemiological studies. The aim of this study was to evaluate the performance of Abbott Real Time HCV Genotype II assay in HCV genotyping of HCV infected patients in Kayseri, Turkey. One hundred patients with chronic hepatitis C admitted to our hospital were evaluated between June 2012 and December 2012, HCV RNA levels were determined by the COBAS® AmpliPrep/COBAS® TaqMan® 48 HCV test. HCV genotyping was investigated by the Abbott Real Time HCV Genotype II assay. With the exception of genotype 1, subtypes of HCV genotypes could not be determined by Abbott assay. Sequencing analysis was used as the reference method. Genotypes 1, 2, 3 and 4 were observed in 70, 4, 2 and 24 of the 100 patients, respectively, by two methods. The concordance between the two systems to determine HCV major genotypes was 100%. Of 70 patients with genotype 1, 66 showed infection with subtype 1b and 4 with subtype 1a by Abbott Real Time HCV Genotype II assay. Using sequence analysis, 61 showed infection with subtype 1b and 9 with subtype 1a. In determining of HCV genotype 1 subtypes, the difference between the two methods was not statistically significant (P>0.05). HCV genotype 4 and 3 samples were found to be subtype 4d and 3a, respectively, by sequence analysis. There were four patients with genotype 2. Sequence analysis revealed that two of these patients had type 2a and the other two had type 2b. The Abbott Real Time HCV Genotype II assay yielded results consistent with sequence analysis. However, further optimization of the Abbott Real Time HCV Genotype II assay for subtype identification of HCV is required.

  3. Hepatitis C virus genotyping of organ donor samples to aid in transplantation of HCV-positive organs.

    Science.gov (United States)

    Gentile, Caren; Van Deerlin, Vivianna M; Goldberg, David S; Reese, Peter P; Hasz, Richard D; Abt, Peter; Blumberg, Emily; Farooqi, Midhat S

    2018-02-01

    Given the availability of new highly efficacious anti-HCV therapies, some clinicians have advocated for wider use of kidneys from hepatitis C virus-positive (HCV+) donors, including transplanting them into HCV-negative recipients. As treatment regimens for HCV are commonly guided by genotype, pretransplant HCV genotyping of tissue donors would be beneficial. To our knowledge, donor HCV genotyping has never been reported. We retrieved archived frozen plasma samples for 17 previous organ donors through a local organ procurement organization. We performed HCV genotyping using the eSensor HCVg Direct Test (GenMark Diagnostics) and also by Sanger sequencing, for confirmation (Retrogen). In addition, viral loads were measured using the COBAS AmpliPrep/TaqMan system (Roche Diagnostics). We found that most of the samples (n = 14) were HCV Genotype 1a with the remainder being Genotype 2b (n = 1) or Genotype 3 (n = 2). All genotyping results were concordant with Sanger sequencing. The average HCV viral load in the sample group was ~ 1.6 million IU/mL (range: ~16 000 IU/mL to 7 million IU/mL). We demonstrate that viral RNA from organ donor plasma can be successfully genotyped for HCV. This ability suggests that transplantation of HCV+ kidneys into HCV-negative recipients, followed by genotype-guided antiviral therapy, could be feasible. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  4. Race moderates the association of Catechol-O-methyltransferase genotype and posttraumatic stress disorder in preschool children.

    Science.gov (United States)

    Humphreys, Kathryn L; Scheeringa, Michael S; Drury, Stacy S

    2014-10-01

    The present study sought to replicate previous findings of an association between the Catechol-O-methyltransferase (COMT) val158met polymorphism with posttraumatic stress disorder (PTSD) and symptomatology in a novel age group, preschool children. COMT genotype was determined in a sample of 171 3-6-year-old trauma-exposed children. PTSD was assessed with a semistructured interview. Accounting for sex, trauma type, and age, genotype was examined in relation to categorical and continuous measures of PTSD both controlling for race and within the two largest racial categories (African American [AA] and European American [EA]). Race significantly moderated the association between genotype and PTSD. Specifically, the genotype associated with increased PTSD symptoms in one racial group had the opposite association in the other racial group. For AA children the met/met genotype was associated with more PTSD symptoms. However, for EA children, val allele carriers had more PTSD symptoms. Whereas every AA child with the met/met genotype met criteria for PTSD, none of the EA children with the met/met genotype did. This genetic association with COMT genotype, in both races but in opposite directions, was most associated with increased arousal symptoms. These findings replicate previous findings in participants of African descent, highlight the moderating effect of race on the association between COMT genotype and PTSD, and provide direct evidence that consideration of population stratification within gene-by-environment studies is valuable to prevent false negative findings.

  5. Testing GxG interactions between coinfecting microbial parasite genotypes within hosts

    Directory of Open Access Journals (Sweden)

    Rebecca D Schulte

    2014-05-01

    Full Text Available Host-parasite interactions represent one of the strongest selection pressures in nature. They are often governed by genotype-specific (GxG interactions resulting in host genotypes that differ in resistance and parasite genotypes that differ in virulence depending on the antagonist’s genotype. Another type of GxG interactions, which is often neglected but which certainly influences host-parasite interactions, are those between coinfecting parasite genotypes. Mechanistically, within-host parasite interactions may range from competition for limited host resources to cooperation for more efficient host exploitation. The exact type of interaction, i.e. whether competitive or cooperative, is known to affect life-history traits such as virulence. However, the latter has been shown for chosen genotype combinations only, not considering whether the specific genotype combination per se may influence the interaction (i.e. GxG interactions. Here, we want to test for the presence of GxG interactions between coinfections of the bacterium Bacillus thuringiensis infecting the nematode Caenorhabditis elegans by combining two non-pathogenic and five pathogenic strains in all possible ways. Furthermore, we evaluate whether the type of interaction, reflected by the direction of virulence change of multiple compared to single infections, is genotype-specific. Generally, we found no indication for GxG interactions between non-pathogenic and pathogenic bacterial strains, indicating that virulence of pathogenic strains is equally affected by both non-pathogenic strains. Specific genotype combinations, however, differ in the strength of virulence change, indicating that the interaction type between coinfecting parasite strains and thus the virulence mechanism is specific for different genotype combinations. Such interactions are expected to influence host-parasite interactions and to have strong implications for coevolution.

  6. Temperature Switch PCR (TSP: Robust assay design for reliable amplification and genotyping of SNPs

    Directory of Open Access Journals (Sweden)

    Mather Diane E

    2009-12-01

    Full Text Available Abstract Background Many research and diagnostic applications rely upon the assay of individual single nucleotide polymorphisms (SNPs. Thus, methods to improve the speed and efficiency for single-marker SNP genotyping are highly desirable. Here, we describe the method of temperature-switch PCR (TSP, a biphasic four-primer PCR system with a universal primer design that permits amplification of the target locus in the first phase of thermal cycling before switching to the detection of the alleles. TSP can simplify assay design for a range of commonly used single-marker SNP genotyping methods, and reduce the requirement for individual assay optimization and operator expertise in the deployment of SNP assays. Results We demonstrate the utility of TSP for the rapid construction of robust and convenient endpoint SNP genotyping assays based on allele-specific PCR and high resolution melt analysis by generating a total of 11,232 data points. The TSP assays were performed under standardised reaction conditions, requiring minimal optimization of individual assays. High genotyping accuracy was verified by 100% concordance of TSP genotypes in a blinded study with an independent genotyping method. Conclusion Theoretically, TSP can be directly incorporated into the design of assays for most current single-marker SNP genotyping methods. TSP provides several technological advances for single-marker SNP genotyping including simplified assay design and development, increased assay specificity and genotyping accuracy, and opportunities for assay automation. By reducing the requirement for operator expertise, TSP provides opportunities to deploy a wider range of single-marker SNP genotyping methods in the laboratory. TSP has broad applications and can be deployed in any animal and plant species.

  7. Hepatitis delta genotypes in chronic delta infection in the northeast of Spain (Catalonia).

    Science.gov (United States)

    Cotrina, M; Buti, M; Jardi, R; Quer, J; Rodriguez, F; Pascual, C; Esteban, R; Guardia, J

    1998-06-01

    Based on genetic analysis of variants obtained around the world, three genotypes of the hepatitis delta virus have been defined. Hepatitis delta virus variants have been associated with different disease patterns and geographic distributions. To determine the prevalence of hepatitis delta virus genotypes in the northeast of Spain (Catalonia) and the correlation with transmission routes and clinical disease, we studied the nucleotide divergence of the consensus sequence of HDV RNA obtained from 33 patients with chronic delta hepatitis (24 were intravenous drug users and nine had no risk factors), and four patients with acute self-limited delta infection. Serum HDV RNA was amplified by the polymerase chain reaction technique and a fragment of 350 nucleotides (nt 910 to 1259) was directly sequenced. Genetic analysis of the nucleotide consensus sequence obtained showed a high degree of conservation among sequences (93% of mean). Comparison of these sequences with those derived from different geographic areas and pertaining to genotypes I, II and III, showed a mean sequence identity of 92% with genotype I, 73% with genotype II and 61% with genotype III. At the amino acid level (aa 115 to 214), the mean identity was 87% with genotype I, 63% with genotype II and 56% with genotype III. Conserved regions included the RNA editing domain, the carboxyl terminal 19 amino acids of the hepatitis delta antigen and the polyadenylation signal of the viral mRNA. Hepatitis delta virus isolates in the northeast of Spain are exclusively genotype I, independently of the transmission route and the type of infection. No hepatitis delta virus subgenotypes were found, suggesting that the origin of hepatitis delta virus infection in our geographical area is homogeneous.

  8. Genotyping isolates of the entomopathogenic fungus Beauveria ...

    African Journals Online (AJOL)

    Multi-locus denaturing gradient gel electrophoresis (DGGE) analysis was developed to investigate the genotypes of Beauveria bassiana sensu lato. ... These results demonstrated that multi-locus DGGE is a potentially useful molecular marker for genotyping, identifying and tracking the fates of experimentally released ...

  9. 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.

  10. Toward fully automated genotyping: Genotyping microsatellite markers by deconvolution

    Energy Technology Data Exchange (ETDEWEB)

    Perlin, M.W.; Lancia, G.; See-Kiong, Ng [Carnegie Mellon Univ., Pittsburgh, PA (United States)

    1995-11-01

    Dense genetic linkage maps have been constructed for the human and mouse genomes, with average densities of 2.9 cM and 0.35 cM, respectively. These genetic maps are crucial for mapping both Mendelian and complex traits and are useful in clinical genetic diagnosis. Current maps are largely comprised of abundant, easily assayed, and highly polymorphic PCR-based microsatellite markers, primarily dinucleotide (CA){sub n} repeats. One key limitation of these length polymorphisms is the PCR stutter (or slippage) artifact that introduces additional stutter bands. With two (or more) closely spaced alleles, the stutter bands overlap, and it is difficult to accurately determine the correct alleles; this stutter phenomenon has all but precluded full automation, since a human must visually inspect the allele data. We describe here novel deconvolution methods for accurate genotyping that mathematically remove PCR stutter artifact from microsatellite markers. These methods overcome the manual interpretation bottleneck and thereby enable full automation of genetic map construction and use. New functionalities, including the pooling of DNAs and the pooling of markers, are described that may greatly reduce the associated experimentation requirements. 32 refs., 5 figs., 3 tabs.

  11. Comparison of Direct Sequencing, Real-Time PCR-High Resolution Melt (PCR-HRM) and PCR-Restriction Fragment Length Polymorphism (PCR-RFLP) Analysis for Genotyping of Common Thiopurine Intolerant Variant Alleles NUDT15 c.415C>T and TPMT c.719A>G (TPMT*3C).

    Science.gov (United States)

    Fong, Wai-Ying; Ho, Chi-Chun; Poon, Wing-Tat

    2017-05-12

    Thiopurine intolerance and treatment-related toxicity, such as fatal myelosuppression, is related to non-function genetic variants encoding thiopurine S-methyltransferase (TPMT) and Nudix hydrolase 15 (NUDT15). Genetic testing of the common variants NUDT15:NM_018283.2:c.415C>T (Arg139Cys, dbSNP rs116855232 T allele) and TPMT: NM_000367.4:c.719A>G (TPMT*3C, dbSNP rs1142345 G allele) in East Asians including Chinese can potentially prevent treatment-related complications. Two complementary genotyping approaches, real-time PCR-high resolution melt (PCR-HRM) and PCR-restriction fragment length morphism (PCR-RFLP) analysis were evaluated using conventional PCR and Sanger sequencing genotyping as the gold standard. Sixty patient samples were tested, revealing seven patients (11.7%) heterozygous for NUDT15 c.415C>T, one patient homozygous for the variant and one patient heterozygous for the TPMT*3C non-function allele. No patient was found to harbor both variants. In total, nine out of 60 (15%) patients tested had genotypic evidence of thiopurine intolerance, which may require dosage adjustment or alternative medication should they be started on azathioprine, mercaptopurine or thioguanine. The two newly developed assays were more efficient and showed complete concordance (60/60, 100%) compared to the Sanger sequencing results. Accurate and cost-effective genotyping assays by real-time PCR-HRM and PCR-RFLP for NUDT15 c.415C>T and TPMT*3C were successfully developed. Further studies may establish their roles in genotype-informed clinical decision-making in the prevention of morbidity and mortality due to thiopurine intolerance.

  12. Plant genotype shapes ant-aphid interactions: implications for community structure and indirect plant defense.

    Science.gov (United States)

    Mooney, Kailen A; Agrawal, Anurag A

    2008-06-01

    Little is known about the mechanisms by which plant genotype shapes arthropod community structure. In a field experiment, we measured the effects of milkweed (Asclepias syriaca) genotype and ants on milkweed arthropods. Populations of the ant-tended aphid Aphis asclepiadis and the untended aphid Myzocallis asclepiadis varied eight- to 18-fold among milkweed genotypes, depending on aphid species and whether ants were present. There was no milkweed effect on predatory arthropods. Ants increased Aphis abundance 59%, decreased Myzocallis abundance 52%, and decreased predator abundance 56%. Milkweed genotype indirectly influenced ants via direct effects on Aphis and Myzocallis abundance. Milkweed genotype also modified ant-aphid interactions, influencing the number of ants attracted per Aphis and Myzocallis. While ant effects on Myzocallis were consistently negative, effects on Aphis ranged from antagonistic to mutualistic among milkweed genotypes. As a consequence of milkweed effects on ant-aphid interactions, ant abundance varied 13-fold among milkweed genotypes, and monarch caterpillar survival was negatively correlated with genetic variation in ant abundance. We speculate that heritable variation in milkweed phloem sap drives these effects on aphids, ants, and caterpillars. In summary, milkweed exerts genetic control over the interactions between aphids and an ant that provides defense against foliage-feeding caterpillars.

  13. Identifying the Genotypes of Hepatitis B Virus (HBV) with DNA Origami Label.

    Science.gov (United States)

    Liu, Ke; Pan, Dun; Wen, Yanqin; Zhang, Honglu; Chao, Jie; Wang, Lihua; Song, Shiping; Fan, Chunhai; Shi, Yongyong

    2018-02-01

    The hepatitis B virus (HBV) genotyping may profoundly affect the accurate diagnosis and antiviral treatment of viral hepatitis. Existing genotyping methods such as serological, immunological, or molecular testing are still suffered from substandard specificity and low sensitivity in laboratory or clinical application. In a previous study, a set of high-efficiency hybridizable DNA origami-based shape ID probes to target the templates through which genetic variation could be determined in an ultrahigh resolution of atomic force microscopy (AFM) nanomechanical imaging are established. Here, as a further confirmatory research to explore the sensitivity and applicability of this assay, differentially predesigned DNA origami shape ID probes are also developed for precisely HBV genotyping. Through the specific identification of visualized DNA origami nanostructure with clinical HBV DNA samples, the genetic variation information of genotypes can be directly identified under AFM. As a proof-of-concept, five genotype B and six genotype C are detected in 11 HBV-infected patients' blood DNA samples of Han Chinese population in the single-blinded test. The AFM image-based DNA origami shape ID genotyping approach shows high specificity and sensitivity, which could be promising for virus infection diagnosis and precision medicine in the future. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Histoimmunogenetics Markup Language 1.0: Reporting next generation sequencing-based HLA and KIR genotyping.

    Science.gov (United States)

    Milius, Robert P; Heuer, Michael; Valiga, Daniel; Doroschak, Kathryn J; Kennedy, Caleb J; Bolon, Yung-Tsi; Schneider, Joel; Pollack, Jane; Kim, Hwa Ran; Cereb, Nezih; Hollenbach, Jill A; Mack, Steven J; Maiers, Martin

    2015-12-01

    We present an electronic format for exchanging data for HLA and KIR genotyping with extensions for next-generation sequencing (NGS). This format addresses NGS data exchange by refining the Histoimmunogenetics Markup Language (HML) to conform to the proposed Minimum Information for Reporting Immunogenomic NGS Genotyping (MIRING) reporting guidelines (miring.immunogenomics.org). Our refinements of HML include two major additions. First, NGS is supported by new XML structures to capture additional NGS data and metadata required to produce a genotyping result, including analysis-dependent (dynamic) and method-dependent (static) components. A full genotype, consensus sequence, and the surrounding metadata are included directly, while the raw sequence reads and platform documentation are externally referenced. Second, genotype ambiguity is fully represented by integrating Genotype List Strings, which use a hierarchical set of delimiters to represent allele and genotype ambiguity in a complete and accurate fashion. HML also continues to enable the transmission of legacy methods (e.g. site-specific oligonucleotide, sequence-specific priming, and Sequence Based Typing (SBT)), adding features such as allowing multiple group-specific sequencing primers, and fully leveraging techniques that combine multiple methods to obtain a single result, such as SBT integrated with NGS. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  15. Mycobacteria and TB

    National Research Council Canada - National Science Library

    Kaufmann, S. H. E. (Stephan H. E.); Hahn, Helmut

    2003-01-01

    .... Scientists investigating the epidemiology, immunology and molecular biology of TB or engaged in vaccine and drug development as well as physicians and social workers treating TB patients will benefit...

  16. Mycobacteria and innate cells

    Indian Academy of Sciences (India)

    PRAKASH KUMAR G

    Effective adaptive immune responses to pathogenic and ... Protective immunity against mycobacterial infections such as Mycobacterium tuberculosis is mediated by interactions ..... 4. γδ T cells as special guests in the antimycobacterial.

  17. Nontuberculous Mycobacteria (NTM)

    Science.gov (United States)

    ... NTM) Infections Program NTM Center of Excellence Our Specialists Shannon H. Kasperbauer, MD + × Shannon H. Kasperbauer, MD ... URL based on the top item shown if (Math.abs(scroll_pos - last_scroll) > $(window).height() * 0. ...

  18. Of Microsoft and Mycobacteria

    Directory of Open Access Journals (Sweden)

    Hans Pasterkamp

    2001-01-01

    Full Text Available Bill Gates should have run for President of the United States. Not only would he have been a candidate with some legitimate claim to the Internet, but his call for a major increase in funding for new defence systems would have been supported by the general public, both at home and abroad. The founder of Microsoft himself spent US$1.44 billion last year to support the fight against global health threats from infectious diseases such as AIDS, malaria and tuberculosis (TB. This contribution from the Bill and Melinda Gates Foundation was more than one-quarter of the total amount that all industrialized nations together raised for this cause.

  19. 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.

  20. Genotype X/C recombinant (putative genotype I) of hepatitis B virus is rare in Hanoi, Vietnam--genotypes B4 and C1 predominate.

    Science.gov (United States)

    Phung, Thi Bich Thuy; Alestig, Erik; Nguyen, Thanh Liem; Hannoun, Charles; Lindh, Magnus

    2010-08-01

    There are eight known genotypes of hepatitis B virus, A-H, and several subgenotypes, with rather well-defined geographic distributions. HBV genotypes were evaluated in 153 serum samples from Hanoi, Vietnam. Of the 87 samples that could be genotyped, genotype B was found in 67 (77%) and genotype C in 19 (22%). All genotype C strains were of subgenotype C1, and the majority of genotype B strains were B4, while a few were B2. The genotype X/C recombinant strain, identified previously in Swedish patients of indigenous Vietnamese origin, was found in one sample. This variant, proposed to be classified as genotype I, has been found recently also by others in Vietnam and Laos. The current study indicates that the genotype X/C recombinant may represent approximately 1% of the HBV strains circulating in Vietnam. (c) 2010 Wiley-Liss, Inc.

  1. Translation of Genotype to Phenotype by a Hierarchy of Cell Subsystems.

    Science.gov (United States)

    Yu, Michael Ku; Kramer, Michael; Dutkowski, Janusz; Srivas, Rohith; Licon, Katherine; Kreisberg, Jason; Ng, Cherie T; Krogan, Nevan; Sharan, Roded; Ideker, Trey

    2016-02-24

    Accurately translating genotype to phenotype requires accounting for the functional impact of genetic variation at many biological scales. Here we present a strategy for genotype-phenotype reasoning based on existing knowledge of cellular subsystems. These subsystems and their hierarchical organization are defined by the Gene Ontology or a complementary ontology inferred directly from previously published datasets. Guided by the ontology's hierarchical structure, we organize genotype data into an "ontotype," that is, a hierarchy of perturbations representing the effects of genetic variation at multiple cellular scales. The ontotype is then interpreted using logical rules generated by machine learning to predict phenotype. This approach substantially outperforms previous, non-hierarchical methods for translating yeast genotype to cell growth phenotype, and it accurately predicts the growth outcomes of two new screens of 2,503 double gene knockouts impacting DNA repair or nuclear lumen. Ontotypes also generalize to larger knockout combinations, setting the stage for interpreting the complex genetics of disease.

  2. Molecular identification of nontuberculous mycobacteria isolated from pyogenic bovine tissues in South Darfur State and Alsabalouga slaughterhouse at Omdurman area, Sudan

    Directory of Open Access Journals (Sweden)

    A.E. El Tigani-Asil

    2014-02-01

    Full Text Available This study identified nontuberculous mycobacteria (NTM recovered from bovine pyogenic affections obtained at necropsy using the molecular target 16S-23S rDNA internal transcribed spacer region. Postmortem inspection of cattle was conducted at South Darfur State abattoirs and Alsabalouga Slaughterhouse at Omdurman area during 2007-2009. Specimens were examined for the presence of acid fast bacteria (AFB using microscopic and standard culturing techniques. AFB were identified phenotypically and confirmed by 16S-23S rDNA ITS. Fifty nine NTM were recovered and confirmed as acid fast filaments out of 165 positive AFB specimens, of which 52 isolates were identified as bovine farcy causative agents, while 7 cultures were excluded due to drying. 16S-23S rDNA ITS of NTM revealed three different amplicons 500 bp. (32 isolates, 550 bp. (2 isolates and 600 bp. (14 isolates. Four isolates were contaminated.

  3. Molecular identification of nontuberculous mycobacteria isolated from pyogenic bovine tissues in South Darfur State and Alsabalouga slaughterhouse at Omdurman area, Sudan.

    Science.gov (United States)

    Tigani-Asil, A E El; Sanousi, S M El; Aljameel, M A; Beir, H El; Adam, A; Abdallatif, M M; Hamid, M E

    2014-01-01

    This study identified nontuberculous mycobacteria (NTM) recovered from bovine pyogenic affections obtained at necropsy using the molecular target 16S-23S rDNA internal transcribed spacer region. Postmortem inspection of cattle was conducted at South Darfur State abattoirs and Alsabalouga Slaughterhouse at Omdurman area during 2007-2009. Specimens were examined for the presence of acid fast bacteria (AFB) using microscopic and standard culturing techniques. AFB were identified phenotypically and confirmed by 16S-23S rDNA ITS. Fifty nine NTM were recovered and confirmed as acid fast filaments out of 165 positive AFB specimens, of which 52 isolates were identified as bovine farcy causative agents, while 7 cultures were excluded due to drying. 16S-23S rDNA ITS of NTM revealed three different amplicons 500 bp. (32) isolates, 550 bp. (2) isolates and 600 bp. (14) isolates. Four isolates were contaminated.

  4. Molecular genotyping of Echinococcus granulosus in the North of Iraq.

    Science.gov (United States)

    Hammad, Salam Jumaah; Cavallero, Serena; Milardi, Giovanni Luigi; Gabrielli, Simona; D Amelio, Stefano; Al-Nasiri, Fatima Shihab

    2018-01-15

    Cystic echinococcosis/hydatidosis is an important cosmopolitan zoonotic disease that causes large economic losses and human suffering. The larval stages of Echinococcus granulosus are the etiological agents of cystic echinococcosis that showed different genotypes in different regions in the world. The present study was aimed at the detection of E. granulosus strains circulating in two cities from north of Iraq (Kirkuk and Sulaimania). A total of 47 specimens of hydatid cysts were collected from patients and from different domestic intermediate hosts including cattle, sheep, goat and buffalo from slaughterhouses. Molecular characterization was performed by direct sequencing of the mitochondrial DNA (mtDNA) genes coding for the cytochrome c oxidase I (cox1) and the small subunit ribosomal RNA (rrnS). The results showed a high prevalence for the sheep strain (G1), an isolated finding of the buffalo strain (G3) and the presence of seven and three different microvariants for cox1 and rrnS, respectively. This is the first contribution on molecular genotyping of E. granulosus in Iraq with the observation of genotypes other than G1. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. Esters of pyrazinoic acid are active against pyrazinamide-resistant strains of Mycobacterium tuberculosis and other naturally resistant mycobacteria in vitro and ex vivo within macrophages.

    KAUST Repository

    Pires, David

    2015-10-05

    Pyrazinamide (PZA) is active against major Mycobacterium tuberculosis species (M. tuberculosis, M. africanum, and M. microti), but not against M. bovis and M. avium. The latter two are mycobacteria species involved in human and cattle tuberculosis and in HIV co-infections, respectively. PZA is a first-line agent for the treatment of human tuberculosis and requires activation by a mycobacterial pyrazinamidase to form the active metabolite pyrazinoic acid (POA). As a result of this mechanism, resistance to PZA as often found in tuberculosis patients is caused by point mutations in pyrazinamidase. In previous work, we have shown that POA esters and amides synthesized in our laboratory were stable in plasma. Although the amides did not present significant activity, the esters were active against sensitive mycobacteria at concentrations 5-to-10 fold lower than those of PZA. Here, we report that these POA derivatives possess antibacterial efficacy in vitro and ex vivo against several species and strains of Mycobacterium with natural or acquired resistance to PZA, including M. bovis and M. avium. Our results indicate that the resistance was probably overcome by cleavage of the prodrugs into POA and a long-chain alcohol. Although it is not possible to rule out that the esters may have intrinsic activity per se, we bring evidence here that long-chain fatty alcohols possess a significant anti-mycobacterial effect against PZA-resistant species and strains and are not mere inactive promoieties. These findings may lead to candidate dual-drugs having enhanced activity against both PZA-susceptible and PZA-resistant isolates and being suitable for clinical development.

  6. Comparative Analyses of Nonpathogenic, Opportunistic, and Totally Pathogenic Mycobacteria Reveal Genomic and Biochemical Variabilities and Highlight the Survival Attributes of Mycobacterium tuberculosis

    Science.gov (United States)

    Singh, Yadvir; Kohli, Sakshi; Ahmad, Javeed; Ehtesham, Nasreen Z.; Tyagi, Anil K.

    2014-01-01

    ABSTRACT Mycobacterial evolution involves various processes, such as genome reduction, gene cooption, and critical gene acquisition. Our comparative genome size analysis of 44 mycobacterial genomes revealed that the nonpathogenic (NP) genomes were bigger than those of opportunistic (OP) or totally pathogenic (TP) mycobacteria, with the TP genomes being smaller yet variable in size—their genomic plasticity reflected their ability to evolve and survive under various environmental conditions. From the 44 mycobacterial species, 13 species, representing TP, OP, and NP, were selected for genomic-relatedness analyses. Analysis of homologous protein-coding genes shared between Mycobacterium indicus pranii (NP), Mycobacterium intracellulare ATCC 13950 (OP), and Mycobacterium tuberculosis H37Rv (TP) revealed that 4,995 (i.e., ~95%) M. indicaus pranii proteins have homology with M. intracellulare, whereas the homologies among M. indicus pranii, M. intracellulare ATCC 13950, and M. tuberculosis H37Rv were significantly lower. A total of 4,153 (~79%) M. indicus pranii proteins and 4,093 (~79%) M. intracellulare ATCC 13950 proteins exhibited homology with the M. tuberculosis H37Rv proteome, while 3,301 (~82%) and 3,295 (~82%) M. tuberculosis H37Rv proteins showed homology with M. indicus pranii and M. intracellulare ATCC 13950 proteomes, respectively. Comparative metabolic pathway analyses of TP/OP/NP mycobacteria showed enzymatic plasticity between M. indicus pranii (NP) and M. intracellulare ATCC 13950 (OP), Mycobacterium avium 104 (OP), and M. tuberculosis H37Rv (TP). Mycobacterium tuberculosis seems to have acquired novel alternate pathways with possible roles in metabolism, host-pathogen interactions, virulence, and intracellular survival, and by implication some of these could be potential drug targets. PMID:25370496

  7. Hepatitis B virus genotypes circulating in Brazil: molecular characterization of genotype F isolates

    Directory of Open Access Journals (Sweden)

    Virgolino Helaine A

    2007-11-01

    Full Text Available Abstract Background Hepatitis B virus (HBV isolates have been classified in eight genotypes, A to H, which exhibit distinct geographical distributions. Genotypes A, D and F are predominant in Brazil, a country formed by a miscegenated population, where the proportion of individuals from Caucasian, Amerindian and African origins varies by region. Genotype F, which is the most divergent, is considered indigenous to the Americas. A systematic molecular characterization of HBV isolates from different parts of the world would be invaluable in establishing HBV evolutionary origins and dispersion patterns. A large-scale study is needed to map the region-by-region distribution of the HBV genotypes in Brazil. Results Genotyping by PCR-RFLP of 303 HBV isolates from HBsAg-positive blood donors showed that at least two of the three genotypes, A, D, and F, co-circulate in each of the five geographic regions of Brazil. No other genotypes were identified. Overall, genotype A was most prevalent (48.5%, and most of these isolates were classified as subgenotype A1 (138/153; 90.2%. Genotype D was the most common genotype in the South (84.2% and Central (47.6% regions. The prevalence of genotype F was low (13% countrywide. Nucleotide sequencing of the S gene and a phylogenetic analysis of 32 HBV genotype F isolates showed that a great majority (28/32; 87.5% belonged to subgenotype F2, cluster II. The deduced serotype of 31 of 32 F isolates was adw4. The remaining isolate showed a leucine-to-isoleucine substitution at position 127. Conclusion The presence of genotypes A, D and F, and the absence of other genotypes in a large cohort of HBV infected individuals may reflect the ethnic origins of the Brazilian population. The high prevalence of isolates from subgenotype A1 (of African origin indicates that the African influx during the colonial slavery period had a major impact on the circulation of HBV genotype A currently found in Brazil. Although most genotype F

  8. Forensic SNP genotyping with SNaPshot

    DEFF Research Database (Denmark)

    Fondevila, M; Børsting, C; Phillips, C

    2017-01-01

    to routine STR profiling, use of SNaPshot is an important part of the development of SNP sets for a wide range of forensic applications with these markers, from genotyping highly degraded DNA with very short amplicons to the introduction of SNPs to ascertain the ancestry and physical characteristics......This review explores the key factors that influence the optimization, routine use, and profile interpretation of the SNaPshot single-base extension (SBE) system applied to forensic single-nucleotide polymorphism (SNP) genotyping. Despite being a mainly complimentary DNA genotyping technique...... of an unidentified contact trace donor. However, this technology, as resourceful as it is, displays several features that depart from the usual STR genotyping far enough to demand a certain degree of expertise from the forensic analyst before tackling the complex casework on which SNaPshot application provides...

  9. HMSRP Hawaiian Monk Seal Microsatellite Genotypes

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Currently ~2,400 Hawaiian monk seal specimens have been analyzed genetically, providing genotypes at 18 microsatellite loci. These data are organized by individual,...

  10. Global distribution of novel rhinovirus genotype

    DEFF Research Database (Denmark)

    Briese, Thomas; Renwick, Neil; Venter, Marietjie

    2008-01-01

    Global surveillance for a novel rhinovirus genotype indicated its association with community outbreaks and pediatric respiratory disease in Africa, Asia, Australia, Europe, and North America. Molecular dating indicates that these viruses have been circulating for at least 250 years Udgivelsesdato...

  11. Assessment of antibiotic susceptibilities, genotypic characteristics ...

    African Journals Online (AJOL)

    Jane

    2011-09-28

    Sep 28, 2011 ... Staphylococcus aureus and Salmonella Typhimurium ... This study was designed to evaluate the antibiotic susceptibilities, genotypic characteristics and ..... Distribution of reference and virulence genes among antibiotic-sensitive S. aureus (SAS), .... environmental factors such as temperature, water activity,.

  12. Early seedling development of Medicago truncatula genotypes ...

    African Journals Online (AJOL)

    adel

    2014-01-08

    Jan 8, 2014 ... heat shock proteins; ABA, abscisic acid. Page 2. Amar et al. 323. Figure 1. Seed vigor of M. truncatula genotypes under different salt stress conditions. Results are means ..... (HSPs) that accumulate during seed late maturation.

  13. Hearing impairment in genotyped Wolfram syndrome patients.

    NARCIS (Netherlands)

    Plantinga, R.F.; Pennings, R.J.E.; Huygen, P.L.M.; Bruno, R.; Eller, P.; Barrett, T.G.; Vialettes, B.; Paquis-Fluklinger, V.; Lombardo, F.; Cremers, C.W.R.J.

    2008-01-01

    OBJECTIVES: Wolfram syndrome is a progressive neurodegenerative syndrome characterized by the features "DIDMOAD" (diabetes insipidus, diabetes mellitus, optic atrophy, and deafness). We sought to study the audiometric data of genotyped Wolfram syndrome patients with sensorineural hearing impairment.

  14. Phylogeography of Japanese encephalitis virus: genotype is associated with climate.

    Directory of Open Access Journals (Sweden)

    Amy J Schuh

    Full Text Available The circulation of vector-borne zoonotic viruses is largely determined by the overlap in the geographical distributions of virus-competent vectors and reservoir hosts. What is less clear are the factors influencing the distribution of virus-specific lineages. Japanese encephalitis virus (JEV is the most important etiologic agent of epidemic encephalitis worldwide, and is primarily maintained between vertebrate reservoir hosts (avian and swine and culicine mosquitoes. There are five genotypes of JEV: GI-V. In recent years, GI has displaced GIII as the dominant JEV genotype and GV has re-emerged after almost 60 years of undetected virus circulation. JEV is found throughout most of Asia, extending from maritime Siberia in the north to Australia in the south, and as far as Pakistan to the west and Saipan to the east. Transmission of JEV in temperate zones is epidemic with the majority of cases occurring in summer months, while transmission in tropical zones is endemic and occurs year-round at lower rates. To test the hypothesis that viruses circulating in these two geographical zones are genetically distinct, we applied Bayesian phylogeographic, categorical data analysis and phylogeny-trait association test techniques to the largest JEV dataset compiled to date, representing the envelope (E gene of 487 isolates collected from 12 countries over 75 years. We demonstrated that GIII and the recently emerged GI-b are temperate genotypes likely maintained year-round in northern latitudes, while GI-a and GII are tropical genotypes likely maintained primarily through mosquito-avian and mosquito-swine transmission cycles. This study represents a new paradigm directly linking viral molecular evolution and climate.

  15. Existence of various human parvovirus B19 genotypes in Chinese plasma pools: genotype 1, genotype 3, putative intergenotypic recombinant variants and new genotypes.

    Science.gov (United States)

    Jia, Junting; Ma, Yuyuan; Zhao, Xiong; Huangfu, Chaoji; Zhong, Yadi; Fang, Chi; Fan, Rui; Lv, Maomin; Zhang, Jingang

    2016-09-17

    Human parvovirus B19 (B19V) is a frequent contaminant of blood and plasma-derived medicinal products. Three distinct genotypes of B19V have been identified. The distribution of the three B19V genotypes has been investigated in various regions or countries. However, in China, data on the existence of different B19V genotypes are limited. One hundred and eighteen B19V-DNA positive source plasma pool samples collected from three Chinese blood products manufacturers were analyzed. The subgenomic NS1/VP1u region junction of B19V was amplified by nested PCR. These amplified products were then cloned and subsequently sequenced. For genotyping, their phylogenetic inferences were constructed based on the NS1/VP1-unique region. Then putative recombination events were analyzed and identified. Phylogenetic analysis of 118 B19V sequences attributed 61.86 % to genotype 1a, 10.17 % to genotype 1b, and 17.80 % to genotype 3b. All the genotype 3b sequences obtained in this study grouped as a specific, closely related cluster with B19V strain D91.1. Four 1a/3b recombinants and 5 new atypical B19V variants with no recombination events were identified. There were at least 3 subtypes (1a, 1b and 3b) of B19V circulating in China. Furthermore, putative B19V 1a/3b recombinants and unclassified strains were identified as well. Such recombinant and unclassified strains may contribute to the genetic diversity of B19V and consequently complicate the B19V infection diagnosis and NAT screening. Further studies will be required to elucidate the biological significance of the recombinant and unclassified strains.

  16. Popcorn genotypes resistance to fall armyworm

    Directory of Open Access Journals (Sweden)

    Nádia Cristina de Oliveira

    2018-02-01

    Full Text Available ABSTRACT: The aim of this study was to evaluate popcorn genotypes for resistance to the fall armyworm, Spodoptera frugiperda. The experiment used a completely randomized design with 30 replicates. The popcorn genotypes Aelton, Arzm 05 083, Beija-Flor, Colombiana, Composto Chico, Composto Gaúcha, Márcia, Mateus, Ufvm Barão Viçosa, Vanin, and Viviane were evaluated,along with the common maize variety Zapalote Chico. Newly hatched fall armyworm larvae were individually assessed with regard to biological development and consumption of food. The data were subjected to multivariate analyses of variance and genetic divergence among genotypes was evaluated through the clustering methods of Tocher based on generalized Mahalanobis distances and canonical variable analyses. Seven popcorn genotypes, namely, Aelton, Arzm 05 083, Composto Chico, Composto Gaúcha, Márcia, Mateus, and Viviane,were shown to form a cluster (cluster I that had antibiosis as the mechanism of resistance to the pest. Cluster I genotypes and the Zapalote Chico genotype could be used for stacking genes for antibiosis and non-preference resistance.

  17. Comparison of Three Different Commercial Kits for the Human Papilloma Virus Genotyping.

    Science.gov (United States)

    Lim, Yong Kwan; Choi, Jee-Hye; Park, Serah; Kweon, Oh Joo; Park, Ae Ja

    2016-11-01

    High-risk type human papilloma virus (HPV) is the most important cause of cervical cancer. Recently, real-time polymerase chain reaction and reverse blot hybridization assay-based HPV DNA genotyping kits are developed. So, we compared the performances of different three HPV genotyping kits using different analytical principles and methods. Two hundred positive and 100 negative cervical swab specimens were used. DNA was extracted and all samples were tested by the MolecuTech REBA HPV-ID, Anyplex II HPV28 Detection, and HPVDNAChip. Direct sequencing was performed as a reference method for confirming high-risk HPV genotypes 16, 18, 45, 52, and 58. Although high-level agreement results were observed in negative samples, three kits showed decreased interassay agreement as screening setting in positive samples. Comparing the genotyping results, three assays showed acceptable sensitivity and specificity for the detection of HPV 16 and 18. Otherwise, various sensitivities showed in the detection of HPV 45, 52, and 58. The three assays had dissimilar performance of HPV screening capacity and exhibited moderate level of concordance in HPV genotyping. These discrepant results were unavoidable due to difference in type-specific analytical sensitivity and lack of standardization; therefore, we suggested that the efforts to standardization of HPV genotyping kits and adjusting analytical sensitivity would be important for the best clinical performance. © 2016 Wiley Periodicals, Inc.

  18. Serotonin transporter genotype linked to adolescent substance use treatment outcome through externalizing behavior

    Directory of Open Access Journals (Sweden)

    Tammy eChung

    2014-07-01

    Full Text Available Meta-analyses suggest that the serotonin transporter linked polymorphic region (5-HTTLPR short (S allele, relative to the long (L allele, is associated with risk for alcohol dependence, particularly among individuals with early onset antisocial alcoholism. Youth in substance use treatment tend to show antisocial or externalizing behaviors, such as conduct problems, which predict worse treatment outcome. This study examined a pathway in which 5-HTTLPR genotype is associated with externalizing behavior, and the intermediate phenotype of externalizing behavior serves as a link between 5-HTTLPR genotype and substance use treatment outcome in youth. Adolescents (n=142 who were recruited from addictions treatment were genotyped for 5-HTTLPR polymorphisms (S and LG carriers vs. LALA, assessed for externalizing and internalizing behaviors shortly after starting treatment, and followed over 6-months. 5-HTTLPR genotype was not associated with internalizing behaviors, and was not directly associated with 6-month substance use outcomes. However, 5-HTTLPR genotype was associated with externalizing behaviors (S and LG > LALA, and externalizing behaviors predicted alcohol and marijuana problem severity at 6-month follow-up. Results indicated an indirect (p<.05 and non-specific (i.e., both alcohol and marijuana severity effect of 5-HTTLPR genotype on youth substance use treatment outcomes, with externalizing behaviors as an important linking factor. Adolescents in substance use treatment with low expressing (S and LG 5-HTTLPR alleles and externalizing behavior might benefit from intervention that addresses serotonergic functioning, externalizing behaviors, and substance use to improve outcomes.

  19. Physical and verbal aggressive behavior and COMT genotype: Sensitivity to the environment.

    Science.gov (United States)

    Tuvblad, Catherine; Narusyte, Jurgita; Comasco, Erika; Andershed, Henrik; Andershed, Anna-Karin; Colins, Olivier F; Fanti, Kostas A; Nilsson, Kent W

    2016-07-01

    Catechol-O-methyltransferase (COMT) genotype has been implicated as a vulnerability factor for several psychiatric diseases as well as aggressive behavior, either directly, or in interaction with an adverse environment. The present study aimed at investigating the susceptibility properties of COMT genotype to adverse and favorable environment in relation to physical and verbal aggressive behavior. The COMT Val158Met polymorphism was genotyped in a Swedish population-based cohort including 1,783 individuals, ages 20-24 years (47% males). A significant three-way interaction was found, after correction for multiple testing, between COMT genotype, exposure to violence, and parent-child relationship in association with physical but not verbal aggressive behavior. Homozygous for the Val allele reported lower levels of physical aggressive behavior when they were exposed to violence and at the same time experienced a positive parent-child relationship compared to Met carriers. Thus, susceptibility properties of COMT genotype were observed in relation to physical aggressive behavior supporting the hypothesis that COMT genotypes are modifying the sensitivity to environment that confers either risk or protection for aggressive behavior. As these are novel findings, they warrant further investigation and replication in independent samples. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  20. Comparison of leaf proteomes of cassava (Manihot esculenta Crantz cultivar NZ199 diploid and autotetraploid genotypes.

    Directory of Open Access Journals (Sweden)

    Feifei An

    Full Text Available Cassava polyploid breeding has drastically improved our knowledge on increasing root yield and its significant tolerance to stresses. In polyploid cassava plants, increases in DNA content highly affect cell volumes and anatomical structures. However, the mechanism of this effect is poorly understood. The purpose of the present study was to compare and validate the changes between cassava cultivar NZ199 diploid and autotetraploid at proteomic levels. The results showed that leaf proteome of cassava cultivar NZ199 diploid was clearly differentiated from its autotetraploid genotype using 2-DE combined MS technique. Sixty-five differential protein spots were seen in 2-DE image of autotetraploid genotype in comparison with that of diploid. Fifty-two proteins were identified by MALDI-TOF-MS/MS, of which 47 were up-regulated and 5 were down-regulated in autotetraploid genotype compared with diploid genotype. The classified functions of 32 up-regulated proteins were associated with photosynthesis, defense system, hydrocyanic acid (HCN metabolism, protein biosynthesis, chaperones, amino acid metabolism and signal transduction. The remarkable variation in photosynthetic activity, HCN content and resistance to salt stress between diploid and autotetraploid genotypes is closely linked with expression levels of proteomic profiles. The analysis of protein interaction networks indicated there are direct interactions between the 15 up-regulation proteins involved in the pathways described above. This work provides an insight into understanding the protein regulation mechanism of cassava polyploid genotype, and gives a clue to improve cassava polyploidy breeding in increasing photosynthesis and resistance efficiencies.

  1. Cotton genotypes selection through artificial neural networks.

    Science.gov (United States)

    Júnior, E G Silva; Cardoso, D B O; Reis, M C; Nascimento, A F O; Bortolin, D I; Martins, M R; Sousa, L B

    2017-09-27

    Breeding programs currently use statistical analysis to assist in the identification of superior genotypes at various stages of a cultivar's development. Differently from these analyses, the computational intelligence approach has been little explored in genetic improvement of cotton. Thus, this study was carried out with the objective of presenting the use of artificial neural networks as auxiliary tools in the improvement of the cotton to improve fiber quality. To demonstrate the applicability of this approach, this research was carried out using the evaluation data of 40 genotypes. In order to classify the genotypes for fiber quality, the artificial neural networks were trained with replicate data of 20 genotypes of cotton evaluated in the harvests of 2013/14 and 2014/15, regarding fiber length, uniformity of length, fiber strength, micronaire index, elongation, short fiber index, maturity index, reflectance degree, and fiber quality index. This quality index was estimated by means of a weighted average on the determined score (1 to 5) of each characteristic of the HVI evaluated, according to its industry standards. The artificial neural networks presented a high capacity of correct classification of the 20 selected genotypes based on the fiber quality index, so that when using fiber length associated with the short fiber index, fiber maturation, and micronaire index, the artificial neural networks presented better results than using only fiber length and previous associations. It was also observed that to submit data of means of new genotypes to the neural networks trained with data of repetition, provides better results of classification of the genotypes. When observing the results obtained in the present study, it was verified that the artificial neural networks present great potential to be used in the different stages of a genetic improvement program of the cotton, aiming at the improvement of the fiber quality of the future cultivars.

  2. Prevalence and Genotype Distribution of Pneumocystis jirovecii in Cuban Infants and Toddlers with Whooping Cough

    Science.gov (United States)

    Monroy-Vaca, Ernesto X.; de Armas, Yaxsier; Illnait-Zaragozí, María T.; Toraño, Gilda; Diaz, Raúl; Vega, Dania; Alvarez-Lam, Ileana; Calderón, Enrique J.

    2014-01-01

    This study describes the prevalence and genotype distribution of Pneumocystis jirovecii obtained from nasopharyngeal (NP) swabs from immunocompetent Cuban infants and toddlers with whooping cough (WC). A total of 163 NP swabs from 163 young Cuban children with WC who were admitted to the respiratory care units at two pediatric centers were studied. The prevalence of the organism was determined by a quantitative PCR (qPCR) assay targeting the P. jirovecii mitochondrial large subunit (mtLSU) rRNA gene. Genotypes were identified by direct sequencing of mtLSU ribosomal DNA (rDNA) and restriction fragment length polymorphism (RFLP) analysis of the dihydropteroate synthase (DHPS) gene amplicons. qPCR detected P. jirovecii DNA in 48/163 (29.4%) samples. mtLSU rDNA sequence analysis revealed the presence of three different genotypes in the population. Genotype 2 was most common (48%), followed in prevalence by genotypes 1 (23%) and 3 (19%); mixed-genotype infections were seen in 10% of the cases. RFLP analysis of DHPS PCR products revealed four genotypes, 18% of which were associated with resistance to sulfa drugs. Only contact with coughers (prevalence ratio [PR], 3.51 [95% confidence interval {CI}, 1.79 to 6.87]; P = 0.000) and exposure to tobacco smoke (PR, 1.82 [95% CI, 1.14 to 2.92]; P = 0.009) were statistically associated with being colonized by P. jirovecii. The prevalence of P. jirovecii in infants and toddlers with WC and the genotyping results provide evidence that this population represents a potential reservoir and transmission source of P. jirovecii. PMID:24131683

  3. Hepatitis C virus genotypes in Bahawalpur

    International Nuclear Information System (INIS)

    Qazi, M.A.; Fayyaz, M.; Chaudhry, G.M.D.; Jamil, A.

    2006-01-01

    This study was conducted at Medical Unit-II Bahawal Victoria Hospital / Quaid-e-Azam Medical College Bahawalpur from May 1st , 2005 to December 31st 2005. The objective of this study was to determine hepatitis C virus (HCV) genotypes in Bahawalpur, Pakistan. In consecutive 105 anti-HCV (ELISA-3) positive patients, complete history and physical examination was performed. Liver function tests, complete blood counts and platelet count, blood sugar fasting and 2 hours after breakfast, prothrombin time, serum albumin, serum globulin and abdominal ultrasound were carried out in all the patients. Tru cut biopsy was performed on 17 patients. We studied HCV RNA in all these patients by Nested PCR method. HCV RNA was detected in 98 patients and geno typing assay was done by genotype specific PCR. Among total of 105 anti-HCV positive patients, HCV-RNA was detected in 98 patients. Out of these 98 patients there were 57 (58.2%) males and 41 (42.8%) females. Their age range was 18-75 years. The age 18-29 years 26 (26.5%), 30-39 years 35 (35.7%) and 40-75 37 (37.8%), while 10 (10.2%) patients were diabetics and 34 (34.7%) patients were obese. Liver cirrhosis was present in 10 (10.2%) patients. Forty two (43.9%) patients were symptomatic while 56 (57.1%) were asymptomatic. Out of 98 patients 11 (11.2%) were un type-able and 87 (88.8%) were type able. 70/98 (71.4%) were genotype 3; 10/98 (10.2%) were genotype 1; 03/98 (3.1%) were genotype 2; 03/98 (3.1%) were mixed genotype 2 and 3; 01/98 (1%) were mixed genotype 3a and 3b. Genotype 3 is the most common HCV virus in our area which shows that both virological and biochemical response will be better. Because HCV genotype 3 is more frequent among the drug users which points towards unsafe injection practices in our area. (author)

  4. The genotypic structure of a multi-host bumblebee parasite suggests a role for ecological niche overlap.

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    Rahel M Salathé

    Full Text Available The genotypic structure of parasite populations is an important determinant of ecological and evolutionary dynamics of host-parasite interactions with consequences for pest management and disease control. Genotypic structure is especially interesting where multiple hosts co-exist and share parasites. We here analyze the natural genotypic distribution of Crithidia bombi, a trypanosomatid parasite of bumblebees (Bombus spp., in two ecologically different habitats over a time period of three years. Using an algorithm to reconstruct genotypes in cases of multiple infections, and combining these with directly identified genotypes from single infections, we find a striking diversity of infection for both data sets, with almost all multi-locus genotypes being unique, and are inferring that around half of the total infections are resulting from multiple strains. Our analyses further suggest a mixture of clonality and sexuality in natural populations of this parasite species. Finally, we ask whether parasite genotypes are associated with host species (the phylogenetic hypothesis or whether ecological factors (niche overlap in flower choice shape the distribution of parasite genotypes (the ecological hypothesis. Redundancy analysis demonstrates that in the region with relatively high parasite prevalence, both host species identity and niche overlap are equally important factors shaping the distribution of parasite strains, whereas in the region with lower parasite prevalence, niche overlap more strongly contributes to the distribution observed. Overall, our study underlines the importance of ecological factors in shaping the natural dynamics of host-parasite systems.

  5. Phenotypic and genotypic variation in Iranian Pistachios

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    Somayeh Tayefeh Aliakbarkhani

    2015-12-01

    Full Text Available As Iran is one of the richest pistachio germplasms a few studies have been conducted on different sexes of pistachio trees, in areas where this crop emerged. To this end, 40 male and female Iranian pistachio genotypes from Feizabad region, Khorasan, Iran; were evaluated using morphological characters and randomly amplified polymorphic DNA (RAPD markers. For morphological assessments, 54 variables were considered to investigate similarities between and among the studied genotypes. Morphological data indicated relative superiority in some female genotypes (such as Sefid 1, Sefid Sabuni 2, Garmesiah, and Ghermezdorosht Z regarding characters such as halfcrackedness, the percentages of protein and fat content. 115 polymorphic bands were recorded with 92.83% average polymorphism among all primers. The total resolving power (Rp of the primers was 74.54. The range of genetic similarity varied from about 0.31 to about 0.70. Genotypes were segregated into eight groups at the similarity limit of 0.41. Results of present investigation could be helpful for strategic decisions for maintaining Iranian pistachio genotypes.

  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. Application of high-resolution DNA melting for genotyping in lepidopteran non-model species: Ostrinia furnacalis (Crambidae.

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    FengBo Li

    Full Text Available Development of an ideal marker system facilitates a better understanding of the genetic diversity in lepidopteran non-model organisms, which have abundant species, but relatively limited genomic resources. Single nucleotide polymorphisms (SNPs discovered within single-copy genes have proved to be desired markers, but SNP genotyping by current techniques remain laborious and expensive. High resolution melting (HRM curve analysis represents a simple, rapid and inexpensive genotyping method that is primarily confined to clinical and diagnostic studies. In this study, we evaluated the potential of HRM analysis for SNP genotyping in the lepidopteran non-model species Ostrinia furnacalis (Crambidae. Small amplicon and unlabeled probe assays were developed for the SNPs, which were identified in 30 females of O. furnacalis from 3 different populations by our direct sequencing. Both assays were then applied to genotype 90 unknown female DNA by prior mixing with known wild-type DNA. The genotyping results were compared with those that were obtained using bi-directional sequencing analysis. Our results demonstrated the efficiency and reliability of the HRM assays. HRM has the potential to provide simple, cost-effective genotyping assays and facilitates genotyping studies in any non-model lepidopteran species of interest.

  8. Saponin profile of green asparagus genotypes.

    Science.gov (United States)

    Vázquez-Castilla, Sara; Jaramillo-Carmona, Sara; Fuentes-Alventosa, Jose María; Jiménez-Araujo, Ana; Rodríguez-Arcos, Rocío; Cermeño-Sacristán, Pedro; Espejo-Calvo, Juan Antonio; Guillén-Bejarano, Rafael

    2013-11-20

    The main goal of this study was to determine the saponin profiles of different "triguero" asparagus genotypes and to compare them to green asparagus commercial hybrids. The samples consisted of 31 commercial hybrids and 58 genotypes from the Huétor-Tájar (HT) population variety ("triguero"). The saponin analysis by high-performance liquid chromatography-mass spectrometry allowed for the determination of 12 saponins derived from a furostan-type steroidal genin, 4 of which had never been described in the edible part of asparagus. The saponin profile of "triguero" asparagus was a combination of these new saponins and protodioscin. Although protodioscin was the major saponin found in commercial hybrids, some of these 12 saponins were detected as major components in some of the commercial hybrids. The total contents of saponins described in some of these HT genotypes reach values as high as 10-100 times higher than those found in commercial hybrids.

  9. Carcass traits of four rabbit genotypes

    Directory of Open Access Journals (Sweden)

    Ajda Kermauner

    2010-01-01

    Full Text Available Seventy-three rabbits of four genotypes (A - SIKA maternal line; C - SIKA sire line; AxC - hybrids between line A and C; AxCal - crossbreds between line A and the Californian breed were used to evaluate the effect of genotype on carcass traits. Rabbits were weaned at 35 days and slaughtered at 93 days of age. Rabbits were fed standard feed mixture ad libitum. The highest live weight at slaughter and dressing percentage was achieved by line C, and the lowest in line A. Hybrids between line A and C exhibited slightly worse carcass traits than rabbits in line C, but the differences were not statistically significant. The Californian breed gave worse results than crossbreeding with line C, though in most cases the differences between AxC and AxCal were not significant. The differences between genotypes in hind leg tissue composition, pH and meat colour were not statistically significant.

  10. Differentiation of Mycobacterium tuberculosis complex from non-tubercular mycobacteria by nested multiplex PCR targeting IS6110, MTP40 and 32kD alpha antigen encoding gene fragments.

    Science.gov (United States)

    Sinha, Pallavi; Gupta, Anamika; Prakash, Pradyot; Anupurba, Shampa; Tripathi, Rajneesh; Srivastava, G N

    2016-03-12

    Control of the global burden of tuberculosis is obstructed due to lack of simple, rapid and cost effective diagnostic techniques that can be used in resource poor-settings. To facilitate the early diagnosis of TB directly from clinical specimens, we have standardized and validated the use of nested multiplex PCR, targeting gene fragments IS6110, MTP40 and 32kD α-antigen encoding genes specific for Mycobacterium tuberculosis complex and non-tubercular mycobacteria (NTM), in comparison to smear microscopy, solid culture and single step multiplex PCR. The results were evaluated in comparison to a composite reference standard (CRS) comprising of microbiological results (smear and culture), clinical, radiological and cytopathological findings, clinical treatment and response to anti-tubercular therapy. The nested multiplex PCR (nMPCR) assay was evaluated to test its utility in 600 (535 pulmonary and 65 extra-pulmonary specimens) clinically suspected TB cases. All specimens were processed for smear, culture, single step multiplex PCR and nested multiplex PCR testing. Out of 535 screened pulmonary and 65 extra-pulmonary specimens, 329 (61.5%) and 19 (29.2%) cases were culture positive for M. tuberculosis. Based on CRS, 450 patients had "clinical TB" (definitive-TB, probable-TB and possible-TB). Remaining 150 were confirmed "non-TB" cases. For culture, the sensitivity was low, 79.3% for pulmonary and 54.3% for extra-pulmonary cases. The sensitivity and specificity results for nMPCR test were evaluated taken composite reference standard as a gold standard. The sensitivity of the nMPCR assay was 97.1% for pulmonary and 91.4% for extra-pulmonary TB cases with specificity of 100% and 93.3% respectively. Nested multiplex PCR using three gene primers is a rapid, reliable and highly sensitive and specific diagnostic technique for the detection and differentiation of M. tuberculosis complex from NTM genome and will be useful in diagnosing paucibacillary samples. Nested multiplex

  11. Genotype x environment interaction for grain yield of wheat genotypes tested under water stress conditions

    International Nuclear Information System (INIS)

    Sail, M.A.; Dahot, M.U.; Mangrio, S.M.; Memon, S.

    2007-01-01

    Effect of water stress on grain yield in different wheat genotypes was studied under field conditions at various locations. Grain yield is a complex polygenic trait influenced by genotype, environment and genotype x environment (GxE) interaction. To understand the stability among genotypes for grain yield, twenty-one wheat genotypes developed Through hybridization and radiation-induced mutations at Nuclear Institute of Agriculture (NIA) TandoJam were evaluated with four local check varieties (Sarsabz, Thori, Margalla-99 and Chakwal-86) in multi-environmental trails (MET/sub s/). The experiments were conducted over 5 different water stress environments in Sindh. Data on grain yield were recorded from each site and statistically analyzed. Combined analysis of variance for all the environments indicated that the genotype, environment and genotype x environment (GxE) interaction were highly significant (P greater then 0.01) for grain yield. Genotypes differed in their response to various locations. The overall highest site mean yield (4031 kg/ha) recorded at Moro and the lowest (2326 kg/ha) at Thatta. Six genotypes produced significantly (P=0.01) the highest grain yield overall the environments. Stability analysis was applied to estimate stability parameters viz., regression coefficient (b), standard error of regression coefficient and variance due to deviation from regression (S/sub 2/d) genotypes 10/8, BWS-78 produced the highest mean yield over all the environments with low regression coefficient (b=0.68, 0.67 and 0.63 respectively and higher S/sup 2/ d value, showing specific adaptation to poor (un favorable) environments. Genotype 8/7 produced overall higher grain yield (3647 kg/ha) and ranked as third high yielding genotype had regression value close to unity (b=0.9) and low S/sup d/ value, indicating more stability and wide adaptation over the all environments. The knowledge of the presence and magnitude of genotype x environment (GE) interaction is important to

  12. A rapid genotyping method for an obligate fungal pathogen, Puccinia striiformis f.sp. tritici, based on DNA extraction from infected leaf and Multiplex PCR genotyping

    Directory of Open Access Journals (Sweden)

    Enjalbert Jérôme

    2011-07-01

    Full Text Available Abstract Background Puccinia striiformis f.sp. tritici (PST, an obligate fungal pathogen causing wheat yellow/stripe rust, a serious disease, has been used to understand the evolution of crop pathogen using molecular markers. However, numerous questions regarding its evolutionary history and recent migration routes still remains to be addressed, which need the genotyping of a large number of isolates, a process that is limited by both DNA extraction and genotyping methods. To address the two issues, we developed here a method for direct DNA extraction from infected leaves combined with optimized SSR multiplexing. Findings We report here an efficient protocol for direct fungal DNA extraction from infected leaves, avoiding the costly and time consuming step of spore multiplication. The genotyping strategy we propose, amplified a total of 20 SSRs in three Multiplex PCR reactions, which were highly polymorphic and were able to differentiate different PST populations with high efficiency and accuracy. Conclusion These two developments enabled a genotyping strategy that could contribute to the development of molecular epidemiology of yellow rust disease, both at a regional or worldwide scale.

  13. Oilseed rape genotypes response to boron toxicity

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    Savić Jasna

    2013-01-01

    Full Text Available Response of 16 oilseed rape genotypes to B (boron toxicity was analyzed by comparing the results of two experiments conducted in a glasshouse. In Experiment 1 plants were grown in standard nutrient solutions with 10 µMB (control and 1000 µM B. Relative root and shoot growth varied from 20-120% and 31-117%, respectively. Variation in B concentration in shoots was also wide (206.5-441.7 µg B g-1 DW as well as total B uptake by plant (62.3-281.2 µg B g1. Four selected genotypes were grown in Experiment 2 in pots filled with high B soil (8 kg ha-1 B; B8. Shoot growth was not affected by B8 treatment, while root and shoot B concentration was significantly increased compared to control. Genotypes Panther and Pronto which performed low relative root and shoot growth and high B accumulation in plants in Experiment 1, had good growth in B8 treatment. In Experiment 2 genotype NS-L-7 had significantly lower B concentration in shots under treatment B8, but also very high B accumulation in Experiment 1. In addition, cluster analyses classified genotypes in three groups according to traits contrasting in their significance for analyzing response to B toxicity. The first group included four varieties based on their shared characteristics that have small value for the relative growth of roots and shoots and large values of B concentration in shoot. In the second largest group were connected ten genotypes that are heterogeneous in traits and do not stand out on any characteristic. Genotypes NS-L-7 and Navajo were separated in the third group because they had big relative growth of root and shoot, but also a high concentration of B in the shoot, and high total B uptake. Results showed that none of tested genotypes could not be recommended for breeding process to tolerance for B toxicity. [Projekat Ministarstva nauke Republike Srbije, br. OI 173028

  14. [Individual Identification of Cartilage by Direct Amplification in Mass Disasters].

    Science.gov (United States)

    Wang, C H; Xu, C; Li, X Q; Wu, Y; Du, Z

    2017-06-01

    To explore the effectiveness of direct amplification for the STR analysis of cartilage, and to accelerate the effectiveness of disaster victim identification. Eighty-eight cartilage samples were directly amplified by PowerPle® 21 kit, and the results of genotyping were compared with that obtained by the magnetic beads method. In 88 cartilage samples, the STR genotypes were successfully detected from 84 samples by direct amplification and magnetic beads method, and both the results of genotyping by two method were consistent. Direct amplification with PowerPlex® 21 kit can be used for STR genotyping of cartilages. This method is operated easily and promptly, which has a potential application in the individual identification of mass disasters. Copyright© by the Editorial Department of Journal of Forensic Medicine

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

    OpenAIRE

    Kocjan, Boštjan; Poljak, Mario; Oštrbenk, Anja

    2015-01-01

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

  16. Genotyping microarray (gene chip) for the ABCR (ABCA4) gene.

    Science.gov (United States)

    Jaakson, K; Zernant, J; Külm, M; Hutchinson, A; Tonisson, N; Glavac, D; Ravnik-Glavac, M; Hawlina, M; Meltzer, M R; Caruso, R C; Testa, F; Maugeri, A; Hoyng, C B; Gouras, P; Simonelli, F; Lewis, R A; Lupski, J R; Cremers, F P M; Allikmets, R

    2003-11-01

    Genetic variation in the ABCR (ABCA4) gene has been associated with five distinct retinal phenotypes, including Stargardt disease/fundus flavimaculatus (STGD/FFM), cone-rod dystrophy (CRD), and age-related macular degeneration (AMD). Comparative genetic analyses of ABCR variation and diagnostics have been complicated by substantial allelic heterogeneity and by differences in screening methods. To overcome these limitations, we designed a genotyping microarray (gene chip) for ABCR that includes all approximately 400 disease-associated and other variants currently described, enabling simultaneous detection of all known ABCR variants. The ABCR genotyping microarray (the ABCR400 chip) was constructed by the arrayed primer extension (APEX) technology. Each sequence change in ABCR was included on the chip by synthesis and application of sequence-specific oligonucleotides. We validated the chip by screening 136 confirmed STGD patients and 96 healthy controls, each of whom we had analyzed previously by single strand conformation polymorphism (SSCP) technology and/or heteroduplex analysis. The microarray was >98% effective in determining the existing genetic variation and was comparable to direct sequencing in that it yielded many sequence changes undetected by SSCP. In STGD patient cohorts, the efficiency of the array to detect disease-associated alleles was between 54% and 78%, depending on the ethnic composition and degree of clinical and molecular characterization of a cohort. In addition, chip analysis suggested a high carrier frequency (up to 1:10) of ABCR variants in the general population. The ABCR genotyping microarray is a robust, cost-effective, and comprehensive screening tool for variation in one gene in which mutations are responsible for a substantial fraction of retinal disease. The ABCR chip is a prototype for the next generation of screening and diagnostic tools in ophthalmic genetics, bridging clinical and scientific research. Copyright 2003 Wiley

  17. Scanning fluorescence detector for high-throughput DNA genotyping

    Science.gov (United States)

    Rusch, Terry L.; Petsinger, Jeremy; Christensen, Carl; Vaske, David A.; Brumley, Robert L., Jr.; Luckey, John A.; Weber, James L.

    1996-04-01

    A new scanning fluorescence detector (SCAFUD) was developed for high-throughput genotyping of short tandem repeat polymorphisms (STRPs). Fluorescent dyes are incorporated into relatively short DNA fragments via polymerase chain reaction (PCR) and are separated by electrophoresis in short, wide polyacrylamide gels (144 lanes with well to read distances of 14 cm). Excitation light from an argon laser with primary lines at 488 and 514 nm is introduced into the gel through a fiber optic cable, dichroic mirror, and 40X microscope objective. Emitted fluorescent light is collected confocally through a second fiber. The confocal head is translated across the bottom of the gel at 0.5 Hz. The detection unit utilizes dichroic mirrors and band pass filters to direct light with 10 - 20 nm bandwidths to four photomultiplier tubes (PMTs). PMT signals are independently amplified with variable gain and then sampled at a rate of 2500 points per scan using a computer based A/D board. LabView software (National Instruments) is used for instrument operation. Currently, three fluorescent dyes (Fam, Hex and Rox) are simultaneously detected with peak detection wavelengths of 543, 567, and 613 nm, respectively. The detection limit for fluorescein-labeled primers is about 100 attomoles. Planned SCAFUD upgrades include rearrangement of laser head geometry, use of additional excitation lasers for simultaneous detection of more dyes, and the use of detector arrays instead of individual PMTs. Extensive software has been written for automatic analysis of SCAFUD images. The software enables background subtraction, band identification, multiple- dye signal resolution, lane finding, band sizing and allele calling. Whole genome screens are currently underway to search for loci influencing such complex diseases as diabetes, asthma, and hypertension. Seven production SCAFUDs are currently in operation. Genotyping output for the coming year is projected to be about one million total genotypes (DNA

  18. An Affymetrix Microarray Design for Microbial Genotyping

    Science.gov (United States)

    2009-10-01

    les échantillons qui ne se prêtent pas aux méthodes culturales de la microbiologie classique. La puce à ADN est une technologie qui permet la... area of microbial genotyping there are multiple platforms that can identify one or a few microbial targets in a single assay iteration. For most

  19. Human papillomavirus genotyping by multiplex pyrosequencing in ...

    Indian Academy of Sciences (India)

    PRAKASH KUMAR G

    malignant cervical samples ... low- and high-risk HPV genotypes without identifying ... Since these samples were not from “healthy .... major capsid protein, any variation in its coding sequence is .... worldwide: a meta-analysis; Br. J. Cancer 88 63–73.

  20. Physicochemical and sensorial quality of banana genotypes

    Directory of Open Access Journals (Sweden)

    Ronielli Cardoso Reis

    2016-03-01

    Full Text Available Despite the diversity of banana varieties in Brazil, only a few cultivars have the proper agronomic traits and fruit quality for commercial exploitation. This study aimed at evaluating the physicochemical traits and sensorial acceptance of banana genotypes, in order to identify those with potential for commercial growing. Six improved banana genotypes were assessed (BRS Maravilha, PC 0101, FHIA 18, TM 2803, YB 4203 and BRS Caipira, as well as three commercial cultivars (Grand Naine, Pacovan and Prata Anã. Analyses of peel and pulp color, peel thickness, pulp yield, moisture, pH, soluble solids, titratable acidity, total carotenoids and sensorial acceptance were performed. The BRS Maravilha, FHIA 18, YB 4203 and BRS Caipira genotypes presented physicochemical traits similar to the Grand Naine, Pacovan and Prata Anã commercial cultivars. The BRS Maravilha and TM 2803 genotypes had sensorial acceptance similar to the Prata Anã and Grand Naine cultivars, and are therefore promising for commercial growing, with the advantage of being resistant to the black Sigatoka and Panama disease.

  1. Genotyping of human pappilomavirus in cervical precancerous ...

    African Journals Online (AJOL)

    Background: Cervical cancer caused by human papilloma virus (HPV), is the second most common cancer for women. This cancer is distributed worldwide, with ~80% of cases are found in the developing countries. In Indonesia, data of HPV genotypes are still limited and do not represent all regions of the country. Thus ...

  2. Cryptosporidium Pig Genotype II in Immunocompetent Man

    Czech Academy of Sciences Publication Activity Database

    Kváč, Martin; Květoňová, Dana; Sak, Bohumil; Ditrich, Oleg

    2009-01-01

    Roč. 15, č. 6 (2009), s. 982-983 ISSN 1080-6040 R&D Projects: GA ČR GP523/07/P117 Institutional research plan: CEZ:AV0Z60220518 Keywords : immunocompetent patients * cryptosporidiosis * Cryptosporidium pig genotype II Subject RIV: GJ - Animal Vermins ; Diseases , Veterinary Medicine Impact factor: 6.794, year: 2009

  3. Morphometric characteristics of Lotus corniculatus L. genotypes ...

    African Journals Online (AJOL)

    The aim of this study was to examine the degree of variability in morphological and agronomic characteristics of 20 Lotus corniculatus L. local genotypes, and also to set aside germplasm that will be used as a source of genetic basis for improvement of the studied properties. In poor quality soils, L. corniculatus L. plays an ...

  4. Genotype X Fertility Interactions in Seedling Sweetgum

    Science.gov (United States)

    Scott X. Chang; Daniel J. Robison

    2002-01-01

    Genotype x fertility interactions may affect the suitability of sweetgum (Liquidambar styraciflua L.) for specific sites or the efficiency of nutrient use. To gain a better understanding of these interactions, 2-year-old sweetgum seedlings from two half-sib families were tested for growth response to N (0 and 100 kg/ha equivalent) and P (0 and 50 kg...

  5. (AMMI) and genotype by environment interaction

    African Journals Online (AJOL)

    SARAH

    2014-04-30

    Apr 30, 2014 ... Background and justification: Lack of stable high yielding cultivars is one ... of advanced finger millet genotypes evaluated in multiple environments, and (ii) identify stable high yielding .... for interaction principal component axis (IPCA) n, γgn ..... Table 2: Analysis of variance for grain yield using AMMI model.

  6. Impact of inter-genotypic recombination and probe cross-reactivity on the performance of the Abbott RealTime HCV Genotype II assay for hepatitis C genotyping.

    Science.gov (United States)

    Sridhar, Siddharth; Yip, Cyril C Y; Chan, Jasper F W; To, Kelvin K W; Cheng, Vincent C C; Yuen, Kwok-Yung

    2018-05-01

    The Abbott RealTime HCV Genotype II assay (Abbott-RT-HCV assay) is a real-time PCR based genotyping method for hepatitis C virus (HCV). This study measured the impact of inter-genotypic recombination and probe cross-reactivity on the performance of the Abbott-RT-HCV assay. 517 samples were genotyped using the Abbott-RT-HCV assay over a one-year period, 34 (6.6%) were identified as HCV genotype 1 without further subtype designation raising the possibility of inaccurate genotyping. These samples were subjected to confirmatory sequencing. 27 of these 34 (79%) samples were genotype 1b while five (15%) were genotype 6. One HCV isolate was an inter-genotypic 1a/4o recombinant. This is a novel natural HCV recombinant that has never been reported. Inter-genotypic recombination and probe cross-reactivity can affect the accuracy of the Abbott-RT-HCV assay, both of which have significant implications on antiviral regimen choice. Confirmatory sequencing of ambiguous results is crucial for accurate genotyping. Copyright © 2018 Elsevier Inc. All rights reserved.

  7. Personal genome testing in medical education: student experiences with genotyping in the classroom.

    Science.gov (United States)

    Vernez, Simone Lucia; Salari, Keyan; Ormond, Kelly E; Lee, Sandra Soo-Jin

    2013-01-01

    Direct-to-consumer (DTC) personal genotyping services are beginning to be adopted by educational institutions as pedagogical tools for learning about human genetics. However, there is little known about student reactions to such testing. This study investigated student experiences and attitudes towards DTC personal genome testing. Individual interviews were conducted with students who chose to undergo personal genotyping in the context of an elective genetics course. Ten medical and graduate students were interviewed before genotyping occurred, and at 2 weeks and 6 months after receiving their genotype results. Qualitative analysis of interview transcripts assessed the expectations and experiences of students who underwent personal genotyping, how they interpreted and applied their results; how the testing affected the quality of their learning during the course, and what were their perceived needs for support. Students stated that personal genotyping enhanced their engagement with the course content. Although students expressed skepticism over the clinical utility of some test results, they expressed significant enthusiasm immediately after receiving their personal genetic analysis, and were particularly interested in results such as drug response and carrier testing. However, few reported making behavioral changes or following up on specific results through a healthcare provider. Students did not report utilizing genetic counseling, despite feeling strongly that the 'general public' would need these services. In follow-up interviews, students exhibited poor recall on details of the consent and biobanking agreements, but expressed little regret over their decision to undergo genotyping. Students reported mining their raw genetic data, and conveyed a need for further consultation support in their exploration of genetic variants. Personal genotyping may improve students' self-reported motivation and engagement with course material. However, consultative support that

  8. 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

  9. Genotyping of Mycobacterium avium subsp. avium isolates from naturally infected lofts of domestic pigeons by IS901 RFLP

    Directory of Open Access Journals (Sweden)

    K Parvandar Asadollahi

    2015-01-01

    In conclusion: It is suggested that more DNA fingerprinting tests for non-tuberculous Mycobacteria, particularly M. avium complex isolated from infected birds and humans, be conducted to find the source of their infections.

  10. In Vitro Conservation of Sweet Potato Genotypes

    Directory of Open Access Journals (Sweden)

    Maria de Fátima Arrigoni-Blank

    2014-01-01

    Full Text Available The aim of this study was to develop a protocol for the in vitro conservation of sweet potato genotypes using the slow growth technique. The first experiment was conducted in a 4×5×2 factorial scheme, testing four genotypes (IPB-007, IPB-052, IPB-072, and IPB-137, five concentrations of abscisic acid (ABA (0.0, 1.0, 2.0, 4.0, and 8.0 mg·L−1, and two temperatures (18 and 25°C. The second experiment was conducted in a 4×3×3 factorial scheme at 18°C, testing four genotypes (IPB-007, IPB-052, IPB-072, and IPB-137, three variations of MS salts (50, 75, and 100%, and three concentrations of sucrose (10, 20, and 30 g·L−1. Every three months, we evaluated the survival (%, shoot height, and shoot viability. In vitro conservation of the sweet potato genotypes IPB-052 and IPB-007 was obtained over three and six months, respectively, using MS medium plus 2.0 mg·L−1 of ABA at either 18 or 25°C. Genotypes IPB-072 and IPB-137 can be kept for three and six months, respectively, in MS medium without ABA at 18°C. It is possible to store IPB-052 and IPB-072 for six months and IPB-007 and IPB-137 for nine months using 30 g·L−1 of sucrose and 50% MS salts.

  11. Esters of pyrazinoic acid are active against pyrazinamide-resistant strains of Mycobacterium tuberculosis and other naturally resistant mycobacteria in vitro and ex vivo within macrophages.

    KAUST Repository

    Pires, David; Valente, Emí lia; Simoes, Marta; Carmo, Nuno; Testa, Bernard; Constantino, Luí s; Anes, Elsa

    2015-01-01

    In previous work, we have shown that POA esters and amides synthesized in our laboratory were stable in plasma. Although the amides did not present significant activity, the esters were active against sensitive mycobacteria at concentrations 5-to-10 fold lower than those of PZA. Here, we report that these POA derivatives possess antibacterial efficacy in vitro and ex vivo against several species and strains of Mycobacterium with natural or acquired resistance to PZA, including M. bovis and M. avium. Our results indicate that the resistance was probably overcome by cleavage of the prodrugs into POA and a long-chain alcohol. Although it is not possible to rule out that the esters may have intrinsic activity per se, we bring evidence here that long-chain fatty alcohols possess a significant anti-mycobacterial effect against PZA-resistant species and strains and are not mere inactive promoieties. These findings may lead to candidate dual-drugs having enhanced activity against both PZA-susceptible and PZA-resistant isolates and being suitable for clinical development.

  12. In Vitro Comparison of Ertapenem, Meropenem, and Imipenem against Isolates of Rapidly Growing Mycobacteria and Nocardia by Use of Broth Microdilution and Etest.

    Science.gov (United States)

    Brown-Elliott, Barbara A; Killingley, Jessica; Vasireddy, Sruthi; Bridge, Linda; Wallace, Richard J

    2016-06-01

    We compared the activities of the carbapenems ertapenem, meropenem, and imipenem against 180 isolates of rapidly growing mycobacteria (RGM) and 170 isolates of Nocardia using the Clinical and Laboratory Standards Institute (CLSI) guidelines. A subset of isolates was tested using the Etest. The rate of susceptibility to ertapenem and meropenem was limited and less than that to imipenem for the RGM. Analysis of major and minor discrepancies revealed that >90% of the isolates of Nocardia had higher MICs by the broth microdilution method than by Etest, in contrast to the lower broth microdilution MICs seen for >80% of the RGM. Imipenem remains the most active carbapenem against RGM, including Mycobacterium abscessus subsp. abscessus For Nocardia, imipenem was significantly more active only against Nocardia farcinica Although there may be utility in testing the activities of the newer carbapenems against Nocardia, their activities against the RGM should not be routinely tested. Testing by Etest is not recommended by the CLSI. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  13. Utility of gastric aspirates for diagnosing tuberculosis in children in a low prevalence area: predictors of positive cultures and significance of non-tuberculous mycobacteria.

    Science.gov (United States)

    Kordy, Faisal; Richardson, Susan E; Stephens, Derek; Lam, Ray; Jamieson, Frances; Kitai, Ian

    2015-01-01

    In countries with low rates of tuberculosis (TB), yields of gastric aspirates (GAs) for Mycobacterium tuberculosis (MTB) culture are low. The significance of non-tuberculous mycobacteria (NTM) isolated from GA is uncertain. We reviewed clinical, microbiologic and radiologic data for children who underwent GA between 1999 and 2011 at Sick Kids, Toronto. Radiologic features of cases were compared with those of age matched controls. 785 GAs were obtained from 285 patients of whom 20 (7%) had positive MTB cultures: in 15 patients the GA was the only positive culture for MTB. Of 15 culture-positive patients who underwent exactly 3 GAs, MTB was isolated from the first lavage in 10 (67%), only from the second in 3 (20%) and only from the third in 2 (13%). On univariate analysis, miliary disease and intrathoracic lymphadenopathy were associated with a positive GA MTB culture. On multiple conditional logistic regression analysis, adenopathy remained significant (OR 10.2 [95% CI 2.0-51.4] p =0.005). Twelve patients had NTM isolated, most commonly M. avium complex: none had evidence of invasive NTM disease during a median duration of 12 months of follow-up. Causal pathogens different from the GA NTM culture were isolated from biopsies or bronchoalveolar lavage in 3. GAs continue to be important for TB diagnosis in children. Three GAs have a yield better than 1. Those with miliary or disseminated TB and intrathoracic lymphadenopathy have highest yields. NTM isolates from GA are likely unimportant and can be clinically misleading.

  14. 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...

  15. Genotypic Variation of Early Maturing Soybean Genotypes for Phosphorus Utilization Efficiency under Field Grown Conditions

    Energy Technology Data Exchange (ETDEWEB)

    Abaidoo, R. C. [Kwame Nkrumah University of Technology, Kumasi (Ghana); International Institute of Tropical Agriculture, Ibadan (Nigeria); Opoku, A.; Boahen, S. [Kwame Nkrumah University of Technology, Kumasi (Ghana); Dare, M. O. [Federal University of Agriculture, Abeokuta (Nigeria)

    2013-11-15

    Variability in the utilization of phosphorus (P) by 64 early-maturing soybean (Glycine max L. Merr.) genotypes under low-P soil conditions were evaluated in 2009 and 2010 at Shika, Nigeria. Fifteen phenotypic variables; number of nodules, nodule dry weight, grain yield, plant biomass, total biomass, biomass N and P content, Phosphorus Utilization Index (PUI), shoot P Utilization efficiency (PUIS), grain P Utilization efficiency (PUIG), Harvest Index (HI), Biological N fixed (BNF), total N fixed and N and P uptake were measured. The four clusters revealed by cluster analysis were basically divided along (1) plant biomass and uptake, (2) nutrient acquisition and utilization and (3) nodulation components. Three early maturing genotypes, TGx1842-14E, TGx1912-11F and TGx1913-5F, were identified as having high P utilization index and low P uptake. These genotypes could be a potential source for breeding for P use efficiency in early maturing soybean genotypes. (author)

  16. Rapid ABO genotyping by high-speed droplet allele-specific PCR using crude samples.

    Science.gov (United States)

    Taira, Chiaki; Matsuda, Kazuyuki; Takeichi, Naoya; Furukawa, Satomi; Sugano, Mitsutoshi; Uehara, Takeshi; Okumura, Nobuo; Honda, Takayuki

    2018-01-01

    ABO genotyping has common tools for personal identification of forensic and transplantation field. We developed a new method based on a droplet allele-specific PCR (droplet-AS-PCR) that enabled rapid PCR amplification. We attempted rapid ABO genotyping using crude DNA isolated from dried blood and buccal cells. We designed allele-specific primers for three SNPs (at nucleotides 261, 526, and 803) in exons 6 and 7 of the ABO gene. We pretreated dried blood and buccal cells with proteinase K, and obtained crude DNAs without DNA purification. Droplet-AS-PCR allowed specific amplification of the SNPs at the three loci using crude DNA, with results similar to those for DNA extracted from fresh peripheral blood. The sensitivity of the methods was 5%-10%. The genotyping of extracted DNA and crude DNA were completed within 8 and 9 minutes, respectively. The genotypes determined by the droplet-AS-PCR method were always consistent with those obtained by direct sequencing. The droplet-AS-PCR method enabled rapid and specific amplification of three SNPs of the ABO gene from crude DNA treated with proteinase K. ABO genotyping by the droplet-AS-PCR has the potential to be applied to various fields including a forensic medicine and transplantation medical care. © 2017 Wiley Periodicals, Inc.

  17. Use of a New High Resolution Melting Method for Genotyping Pathogenic Leptospira spp.

    Directory of Open Access Journals (Sweden)

    Florence Naze

    Full Text Available Leptospirosis is a worldwide zoonosis that is endemic in tropical areas, such as Reunion Island. The species Leptospira interrogans is the primary agent in human infections, but other pathogenic species, such as L. kirschner and L. borgpetersenii, are also associated with human leptospirosis.In this study, a melting curve analysis of the products that were amplified with the primer pairs lfb1 F/R and G1/G2 facilitated an accurate species classification of Leptospira reference strains. Next, we combined an unsupervised high resolution melting (HRM method with a new statistical approach using primers to amplify a two variable-number tandem-repeat (VNTR for typing at the subspecies level. The HRM analysis, which was performed with ScreenClust Software, enabled the identification of genotypes at the serovar level with high resolution power (Hunter-Gaston index 0.984. This method was also applied to Leptospira DNA from blood samples that were obtained from Reunion Island after 1998. We were able to identify a unique genotype that is identical to that of the L. interrogans serovars Copenhageni and Icterohaemorrhagiae, suggesting that this genotype is the major cause of leptospirosis on Reunion Island.Our simple, rapid, and robust genotyping method enables the identification of Leptospira strains at the species and subspecies levels and supports the direct genotyping of Leptospira in biological samples without requiring cultures.

  18. 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.

  19. High-throughput genotyping of single nucleotide polymorphisms with rolling circle amplification

    Directory of Open Access Journals (Sweden)

    Sun Zhenyu

    2001-08-01

    Full Text Available Abstract Background Single nucleotide polymorphisms (SNPs are the foundation of powerful complex trait and pharmacogenomic analyses. The availability of large SNP databases, however, has emphasized a need for inexpensive SNP genotyping methods of commensurate simplicity, robustness, and scalability. We describe a solution-based, microtiter plate method for SNP genotyping of human genomic DNA. The method is based upon allele discrimination by ligation of open circle probes followed by rolling circle amplification of the signal using fluorescent primers. Only the probe with a 3' base complementary to the SNP is circularized by ligation. Results SNP scoring by ligation was optimized to a 100,000 fold discrimination against probe mismatched to the SNP. The assay was used to genotype 10 SNPs from a set of 192 genomic DNA samples in a high-throughput format. Assay directly from genomic DNA eliminates the need to preamplify the target as done for many other genotyping methods. The sensitivity of the assay was demonstrated by genotyping from 1 ng of genomic DNA. We demonstrate that the assay can detect a single molecule of the circularized probe. Conclusions Compatibility with homogeneous formats and the ability to assay small amounts of genomic DNA meets the exacting requirements of automated, high-throughput SNP scoring.

  20. Antioxidant activity of seedling growth in selected soybean genotypes (Glycine max (L.) Merrill) responses of submergence

    Science.gov (United States)

    Damanik, R. I.; Marbun, P.; Sihombing, L.

    2016-08-01

    In order to better understand the physiological and biochemical responses relating to direct seeding establishment in soybeans, the plant growth rate and antioxidative defense responses of seedlings in seven Indonesian soybean genotypes (Anjasmoro, Detam-1, Detam-2, Dieng, Grobogan, Tanggamus, and Willis) at different submergence periods (4, and 8 days) were examined. Twelve-day old seedlings were hydroponically grown in limited oxygen conditions. The results showed that the chlorophyll content in soybean seedlings was reduced beginning as early as 4 d under submerged condition, except for Detam-1, Detam-2, and Grobogan genotypes. The dry weight and protein concentration of seedlings were significantly higher at control condition (0 d) than those in submerged condition. The activities of superoxide dismutase (SOD) increased linearly until 8 d submerged for all genotypes. On the other hand, our results showed that catalase (CAT) and ascorbate peroxidase (APX) activities did not work together, meaning that CAT is activated and APX deactivated, or vice versa, in response to submergence conditions, except for Grobogan and Tanggamus genotypes which had an effect on both CAT and APX activities. Submergence stress led to a significant increase in glutathione reductase (GR) together with APX activity for Detam-2 and Dieng genotypes at 8 d submerged.

  1. 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.

  2. Differential growth responses of Brachypodium distachyon genotypes to inoculation with plant growth promoting rhizobacteria.

    Science.gov (United States)

    do Amaral, Fernanda P; Pankievicz, Vânia C S; Arisi, Ana Carolina M; de Souza, Emanuel M; Pedrosa, Fabio; Stacey, Gary

    2016-04-01

    Plant growth promoting rhizobacteria (PGPR) can associate and enhance the growth of important crop grasses. However, in most cases, the molecular mechanisms responsible for growth promotion are not known. Such research could benefit by the adoption of a grass model species that showed a positive response to bacterial inoculation and was amenable to genetic and molecular research methods. In this work we inoculated different genotypes of the model grass Brachypodium distachyon with two, well-characterized PGPR bacteria, Azospirillum brasilense and Herbaspirillum seropedicae, and evaluated the growth response. Plants were grown in soil under no nitrogen or with low nitrogen (i.e., 0.5 mM KNO3). A variety of growth parameters (e.g., shoot height, root length, number of lateral roots, fresh and dry weight) were measured 35 days after inoculation. The data indicate that plant genotype plays a very important role in determining the plant response to PGPR inoculation. A positive growth response was observed with only four genotypes grown under no nitrogen and three genotypes tested under low nitrogen. However, in contrast, relatively good root colonization was seen with most genotypes, as measured by drop plate counting and direct, microscopic examination of roots. In particular, the endophytic bacteria H. seropedicae showed strong epiphytic and endophytic colonization of roots.

  3. A new HCV genotype 6 subtype designated 6v was confirmed with three complete genome sequences.

    Science.gov (United States)

    Wang, Yizhong; Xia, Xueshan; Li, Chunhua; Maneekarn, Niwat; Xia, Wenjie; Zhao, Wenhua; Feng, Yue; Kung, Hsiang Fu; Fu, Yongshui; Lu, Ling

    2009-03-01

    Although hepatitis C virus (HCV) genotype 6 is classified into 21 subtypes, 6a-6u, new variants continue to be identified. To characterize the full-length genomes of three novel HCV genotype 6 variants: KMN02, KM046 and KM181. From sera of patients with HCV infection, the entire HCV genome was amplified by RT-PCR followed by direct DNA sequencing and phylogenetic analysis. The sera contained HCV genomes of 9461, 9429, and 9461nt in length, and each harboured a single ORF of 9051nt. The genomes showed 95.3-98.1% nucleotide similarity to each other and 72.2-75.4% similarity to 23 genotype 6 reference sequences, which represent subtypes 6a-6u and unassigned variants km41 and gz52557. Phylogenetic analyses demonstrated that they were genotype 6, but were subtypically distinct. Based on the current criteria of HCV classification, they were designed to represent a new subtype, 6v. Analysis of E1 and NS5B region partial sequences revealed two additional related variants, CMBD-14 and CMBD-86 that had been previously reported in northern Thailand and sequences dropped into Genbank. Three novel HCV genotype 6 variants were entirely sequenced and designated subtype 6v.

  4. Distribution of genotype network sizes in sequence-to-structure genotype-phenotype maps.

    Science.gov (United States)

    Manrubia, Susanna; Cuesta, José A

    2017-04-01

    An essential quantity to ensure evolvability of populations is the navigability of the genotype space. Navigability, understood as the ease with which alternative phenotypes are reached, relies on the existence of sufficiently large and mutually attainable genotype networks. The size of genotype networks (e.g. the number of RNA sequences folding into a particular secondary structure or the number of DNA sequences coding for the same protein structure) is astronomically large in all functional molecules investigated: an exhaustive experimental or computational study of all RNA folds or all protein structures becomes impossible even for moderately long sequences. Here, we analytically derive the distribution of genotype network sizes for a hierarchy of models which successively incorporate features of increasingly realistic sequence-to-structure genotype-phenotype maps. The main feature of these models relies on the characterization of each phenotype through a prototypical sequence whose sites admit a variable fraction of letters of the alphabet. Our models interpolate between two limit distributions: a power-law distribution, when the ordering of sites in the prototypical sequence is strongly constrained, and a lognormal distribution, as suggested for RNA, when different orderings of the same set of sites yield different phenotypes. Our main result is the qualitative and quantitative identification of those features of sequence-to-structure maps that lead to different distributions of genotype network sizes. © 2017 The Author(s).

  5. 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.

  6. Behavior of durum wheat genotypes under normal irrigation and ...

    African Journals Online (AJOL)

    Behavior of durum wheat genotypes under normal irrigation and drought stress conditions in the greenhouse. ... African Journal of Biotechnology ... Genotypes were grouped in cluster analysis (using Ward's method) based on Yp, Ys and ...

  7. Genetic variability in cowpea (Vigna unguiculata (L.) Walp.) genotypes

    African Journals Online (AJOL)

    sive protein in human diets with grains containing about. 23–25% protein ... Keywords: heritability, phenotype, principal component analysis, variance. Introduction .... be due to genotype, environment, and the interaction of genotype and ...

  8. magnitude of genotype x environment interaction for bacterial leaf

    African Journals Online (AJOL)

    ACSS

    African Crop Science Journal, Vol. ... effects of treatments into genotype, environment, and genotype x environment (G x E) interactions. Results .... method is economically effective (Niño-Liu et al., ..... This phenomenon indicated differences in.

  9. 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.

  10. HPV genotypes in invasive cervical cancer in Danish women

    DEFF Research Database (Denmark)

    Kirschner, Benny; Junge, Jette; Holl, Katsiaryna

    2013-01-01

    Human papillomavirus (HPV) genotype distribution in invasive cervical cancers may differ by geographic region. The primary objective of this study was to estimate HPV-genotype distribution in Danish women with a diagnosis of invasive cervical cancer.......Human papillomavirus (HPV) genotype distribution in invasive cervical cancers may differ by geographic region. The primary objective of this study was to estimate HPV-genotype distribution in Danish women with a diagnosis of invasive cervical cancer....

  11. Evaluation of promising sweetpotato genotypes for high altitude ...

    African Journals Online (AJOL)

    The trials were set up to identify sweetpotato genotypes with adaptation to highland agroecologies with special reference to resistance to Ahemaria blight ... growth and at harvest, four genotypes and the local check, Magabari, bad high levels of resistance toA/Jemaria blight. Eight genotypes had total storage root yield ...

  12. Hepatitis B virus Genotypes in West Azarbayjan Province, Northwest Iran

    Directory of Open Access Journals (Sweden)

    Mohammad Hasan Khadem Ansari

    2017-12-01

    CONCLUSIONS: The results reveal that D genotype is the main genotype of HBV in West Azarbayjan province. Presence of this genotype conformed with the low rate of acute liver diseases caused by hepatitis B chronic infection, cirrhosis of the liver and hepatocellular carcinoma.

  13. Genotype-based personalised nutrition for obesity prevention and ...

    African Journals Online (AJOL)

    Typically, genotype-based personalised nutrition involves genotyping for a number of susceptibility SNPs associated with the prevention, or management, of a particular disease. Dietary advice is then personalised to the individual's genotype to ensure optimal prevention or treatment outcomes. To ensure evidence-based ...

  14. Introduction to a special issue on genotype by environment interaction

    Science.gov (United States)

    Expression of a phenotype is a function of the genotype, the environment, and the differential sensitivity of certain genotypes to different environments, also known as genotype by environment (G × E) interaction. This special issue of Crop Science includes a collection of manuscripts that reviews t...

  15. Core Gene Expression and Association of Genotypes with Viral ...

    African Journals Online (AJOL)

    Purpose: To determine genotypic distribution, ribonucleic acid (RNA) RNA viral load and express core gene from Hepatitis C Virus (HCV) infected patients in Punjab, Pakistan. Methods: A total of 1690 HCV RNA positive patients were included in the study. HCV genotyping was tested by type-specific genotyping assay, viral ...

  16. Effect of Genotype and Age on Some Morphometric, Body Linear ...

    African Journals Online (AJOL)

    A population of 231 roosters of the Nigerian indigenous chickens of normal feathered frizzle feathered and naked neck genotypes was evaluated for the effect of genotype and age on some morphometric body linear measurements and semen characteristics of three Nigerian chicken genotypes. 20 roosters from each ...

  17. 21 CFR 862.3360 - Drug metabolizing enzyme genotyping system.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Drug metabolizing enzyme genotyping system. 862... Test Systems § 862.3360 Drug metabolizing enzyme genotyping system. (a) Identification. A drug metabolizing enzyme genotyping system is a device intended for use in testing deoxyribonucleic acid (DNA...

  18. Relationship of status of polymorphic rapd bands with genotypic ...

    African Journals Online (AJOL)

    Relationship of status of polymorphic rapd bands with genotypic adaptation in early finger millet genotypes. S Das, RC Misra, GR Rout, MC Pattanaik, S Aparajita. Abstract. Molecular characterisation of the 15 early duration finger millet (Eleusine coracana G) genotypes was done through RAPD markers. Twenty-five ...

  19. Genotype x Environment Interaction for Tuber Yield, Dry Matter ...

    African Journals Online (AJOL)

    A study was conducted to determine stability of tuber yield, dry matter content and specific gravity, and the nature and magnitude of genotype x environment (G x E) interaction in elite tetraploid potato genotypes. Eleven potato genotypes including two standard checks were evaluated in the eastern part of Ethiopia at ...

  20. [Evaluation of hepatitis B virus genotyping EIA kit].

    Science.gov (United States)

    Tanaka, Yasuhito; Sugauchi, Fuminaka; Matsuuraa, Kentaro; Naganuma, Hatsue; Tatematsu, Kanako; Takagi, Kazumi; Hiramatsu, Kumiko; Kani, Satomi; Gotoh, Takaaki; Wakimoto, Yukio; Mizokami, Masashi

    2009-01-01

    Clinical significance of Hepatitis B virus(HBV) genotyping is increasingly recognized. The aim of this study was to evaluate reproducibility, accuracy, and sensitivity of an enzyme immunoassay (EIA) based HBV genotyping kit, which designed to discriminate between genotypes to A, B, C, or D by detecting genotype-specific epitopes in PreS2 region. Using the four genotypes panels, the EIA demonstrated complete inter and intra-assay genotyping reproducibility. Serum specimens had stable results after 8 days at 4 degrees C, or 10 cycles of freezing-thawing. In 91 samples that have been genotyped by DNA sequencing, 87(95.6%) were in complete accordance with EIA genotyping. Of examined 344 HBsAg-positive serum specimens, genotypes A, B, C and D were determined in 26 (7.6%), 62 (18.0%), 228 (66.3%), and 9 (2.6%) cases, respectively. Of 19 (5.5%) specimens unclassified by the EIA, 13 were found to have low titer of HBsAg concentration (< 3 IU/ml), and the other 5 had amino acid mutations or deletions within targeted PreS2 epitopes. The EIA allowed genotyping even in HBV DNA negative samples (96.2%). In conclusion, HBV genotype EIA is reliable, sensitive and easy assay for HBV genotyping. The assay would be useful for clinical use.

  1. Genotype by environment interactions and yield stability of stem ...

    African Journals Online (AJOL)

    In a maize breeding program, potential genotypes are usually evaluated in different environments before desirable ones are selected. Genotype x environment (G x E) interaction is associated with the differential performance of genotypes tested at different locations and in different years, and influences selection and ...

  2. Genotype x environment interaction and stability analysis for yield ...

    African Journals Online (AJOL)

    Chickpea is the major pulse crop cultivated in Ethiopia. However, its production is constrained due to genotype instability and environmental variability. This research was carried out to examine the magnitude of environmental effect on yield of chickpea genotypes and to investigate the stability and adaptability of genotypes ...

  3. Reactions of some potato genotypes to late blight in Cameroon ...

    African Journals Online (AJOL)

    Reactions of some potato genotypes to late blight in Cameroon. D. K. Njualem, P. Demo, H. A. Mendoza, J. T. Koi, S. F. Nana. Abstract. Field experiments were conducted in Cameroon in 1995 and 1996 to evaluate reactions of different potato genotypes to late blight. There were significant differences among genotypes for ...

  4. Discovery of novel variants in genotyping arrays improves genotype retention and reduces ascertainment bias

    Directory of Open Access Journals (Sweden)

    Didion John P

    2012-01-01

    Full Text Available Abstract Background High-density genotyping arrays that measure hybridization of genomic DNA fragments to allele-specific oligonucleotide probes are widely used to genotype single nucleotide polymorphisms (SNPs in genetic studies, including human genome-wide association studies. Hybridization intensities are converted to genotype calls by clustering algorithms that assign each sample to a genotype class at each SNP. Data for SNP probes that do not conform to the expected pattern of clustering are often discarded, contributing to ascertainment bias and resulting in lost information - as much as 50% in a recent genome-wide association study in dogs. Results We identified atypical patterns of hybridization intensities that were highly reproducible and demonstrated that these patterns represent genetic variants that were not accounted for in the design of the array platform. We characterized variable intensity oligonucleotide (VINO probes that display such patterns and are found in all hybridization-based genotyping platforms, including those developed for human, dog, cattle, and mouse. When recognized and properly interpreted, VINOs recovered a substantial fraction of discarded probes and counteracted SNP ascertainment bias. We developed software (MouseDivGeno that identifies VINOs and improves the accuracy of genotype calling. MouseDivGeno produced highly concordant genotype calls when compared with other methods but it uniquely identified more than 786000 VINOs in 351 mouse samples. We used whole-genome sequence from 14 mouse strains to confirm the presence of novel variants explaining 28000 VINOs in those strains. We also identified VINOs in human HapMap 3 samples, many of which were specific to an African population. Incorporating VINOs in phylogenetic analyses substantially improved the accuracy of a Mus species tree and local haplotype assignment in laboratory mouse strains. Conclusion The problems of ascertainment bias and missing

  5. COMT genotype, gambling activity, and cognition

    DEFF Research Database (Denmark)

    Grant, Jon E; Leppink, Eric W; Redden, Sarah A

    2015-01-01

    adjustment and delay aversion) and the Spatial Working Memory task (total errors). This study adds to the growing literature on the role of COMT in impulsive behaviors by showing that the Val/Val genotype was associated with specific clinical and cognitive elements among young adults who gamble......Neuropsychological studies of adults with problem gambling indicate impairments across multiple cognitive domains. Catechol-O-methyltransferase (COMT) plays a unique role in the regulation of dopamine in the prefrontal cortex, and has been implicated in the cognitive dysfunction evident in problem...... gambling. This study examined adults with varying levels of gambling behavior to determine whether COMT genotype was associated with differences in gambling symptoms and cognitive functioning. 260 non-treatment-seeking adults aged 18-29 years with varying degrees of gambling behavior provided saliva...

  6. FRUIT QUALITY CHARACTERISTICS OF SOME BLUEBERRY GENOTYPES

    Directory of Open Access Journals (Sweden)

    Irina Ancu

    2013-12-01

    Full Text Available In Romania the blueberry breeding program started in 1982 and till now was conducted by dr. Paulina Mladin. For inducing the variability, different genetic resources of American blueberry cultivars (V. corymbosum, V. angustifolium were involved in a high number of crosses. For identify the genotype with the best fruit quality, some biometric quality indicators (average fruit weight, size index and basically chemical compounds of fruits including ascorbic acid, dry matter, ash, soluble solids, total sugar, titratable acidity, tanoid substances, pectic substances, protein crude, phosphorus and potassium were determined. Of the eleven chemical studied properties who reflected the fruits quality, for five of them were found no statistically significant differences. The purpose of this paper work was to evaluate fruit quality and to identify the valuable genotypes resulted from Romanian blueberry breeding program.

  7. BCL2 genotypes and prostate cancer survival

    Energy Technology Data Exchange (ETDEWEB)

    Renner, Wilfried [Medical University of Graz, Clinical Institute of Medical and Chemical Laboratory Diagnostics, Graz (Austria); Langsenlehner, Uwe [GKK Outpatient Department, Division of Internal Medicine, Graz (Austria); Krenn-Pilko, Sabine; Langsenlehner, Tanja [Medical University of Graz, Department of Therapeutic Radiology and Oncology, Graz (Austria); Eder, Petra [University Hospital Wuerzburg, Department of Internal Medicine I, Wuerzburg (Germany)

    2017-06-15

    The antiapoptotic B-cell lymphoma 2 (BCL2) gene is a key player in cancer development and progression. A functional single-nucleotide polymorphism (c.-938C>A, rs2279115) in the inhibitory P2 BCL2 gene promoter has been associated with clinical outcomes in various types of cancer. Aim of the present study was to analyze the role of BCL2-938C>A genotypes in prostate cancer mortality. The association between BCL2-938C>A (rs2279115) genotypes and prostate cancer outcome was studied within the prospective PROCAGENE study comprising 702 prostate cancer patients. During a median follow-up time of 92 months, 120 (17.1%) patients died. A univariate Cox regression model showed a significant association of the CC genotype with reduced cancer-specific survival (CSS; hazard ratio, HR, 2.13, 95% confidence interval, CI, 1.10-4.12; p = 0.024) and overall survival (OS; HR 2.34, 95% CI 1.58-3.47; p < 0.001). In a multivariate Cox regression model including age at diagnosis, risk group, and androgen deprivation therapy, the CC genotype remained a significant predictor of poor CSS (HR 2.05, 95% CI 1.05-3.99; p = 0.034) and OS (HR 2.25, 95% CI 1.51-3.36; p < 0.001). This study provides evidence that the homozygous BCL2-938 CC genotype is associated with OS and C in prostate cancer patients. (orig.) [German] Das antiapoptotische Gen B cell lymphoma 2 (BCL2) spielt eine Schluesselrolle in der Entstehung und Progression von Krebserkrankungen. Ein funktioneller Einzelnukleotid-Polymorphismus (c.-938C>A, rs2279115) im inhibitorischen P2-BCL2-Promotor wurde mit dem klinischen Outcome verschiedener Krebserkrankungen verknuepft. Ziel der vorliegenden Studie war die Untersuchung der Rolle von BCL2-938C>A-Genotypen fuer die Mortalitaet bei Patienten mit Prostatakarzinom. Der Zusammenhang zwischen BCL2-938C>A-Genotypen (rs2279115) und dem Outcome bei Prostatakrebs wurde in der prospektiven PROCAGENE-Studie, die 702 Patienten mit Prostatakarzinom umfasste, untersucht. Waehrend der medianen

  8. Emerging genotype (GGIIb) of norovirus in drinking water, Sweden.

    Science.gov (United States)

    Nygård, Karin; Torvén, Maria; Ancker, Camilla; Knauth, Siv Britt; Hedlund, Kjell-Olof; Giesecke, Johan; Andersson, Yvonne; Svensson, Lennart

    2003-12-01

    From May through June 2001, an outbreak of acute gastroenteritis that affected at least 200 persons occurred in a combined activity camp and conference center in Stockholm County. The source of illness was contaminated drinking water obtained from private wells. The outbreak appears to have started with sewage pipeline problems near the kitchen, which caused overflow of the sewage system and contaminated the environment. While no pathogenic bacteria were found in water or stools specimens, norovirus was detected in 8 of 11 stool specimens and 2 of 3 water samples by polymerase chain reaction. Nucleotide sequencing of amplicons from two patients and two water samples identified an emerging genotype designated GGIIb, which was circulating throughout several European countries during 2000 and 2001. This investigation documents the first waterborne outbreak of viral gastroenteritis in Sweden, where nucleotide sequencing showed a direct link between contaminated water and illness.

  9. Novel approach for CES1 genotyping

    DEFF Research Database (Denmark)

    Bjerre, Ditte; Berg Rasmussen, Henrik

    2018-01-01

    AIM: Development of a specific procedure for genotyping of CES1A1 (CES1) and CES1A2, a hybrid of CES1A1 and the pseudogene CES1P1. MATERIALS & METHODS: The number of CES1A1 and CES1A2 copies and that of CES1P1 were determined using real-time PCR. Long range PCRs followed by secondary PCRs allowed...

  10. Blood group genotyping: the power and limitations of the Hemo ID Panel and MassARRAY platform.

    Science.gov (United States)

    McBean, Rhiannon S; Hyland, Catherine A; Flower, Robert L

    2015-01-01

    Matrix-assisted laser desorption/ionization, time-of-flight mass spectrometry (MALDI-TOF MS), is a sensitive analytical method capable of resolving DNA fragments varying in mass by a single nucleotide. MALDI-TOF MS is applicable to blood group genotyping, as the majority of blood group antigens are encoded by single nucleotide polymorphisms. Blood group genotyping by MALDI-TOF MS can be performed using a panel (Hemo ID Blood Group Genotyping Panel, Agena Bioscience Inc., San Diego, CA) that is a set of genotyping assays that predict the phenotype for 101 antigens from 16 blood group systems. These assays involve three fundamental stages: multiplex target-specific polymerase chain reaction amplification, allele-specific single base primer extension, and MALDI-TOFMS analysis using the MassARRAY system. MALDI-TOF MS-based genotyping has many advantages over alternative methods including high throughput, high multiplex capability, flexibility and adaptability, and the high level of accuracy based on the direct detection method. Currently available platforms for MALDI-TOF MS-based genotyping are not without limitations, including high upfront instrumentation costs and the number of non-automated steps. The Hemo ID Blood Group Genotyping Panel, developed and optimized in a collaboration between the vendor and the Blood Transfusion Service of the Swiss Red Cross in Zurich, Switzerland, is not yet widely utilized, although several laboratories are currently evaluating the MassARRAY system for blood group genotyping. Based on the accuracy and other advantages offered by MALDITOF MS analysis, in the future, this method is likely to become widely adopted for blood group genotyping, in particular, for population screening.

  11. Screening cotton genotypes for seedling drought tolerance

    Directory of Open Access Journals (Sweden)

    Penna Julio C. Viglioni

    1998-01-01

    Full Text Available The objectives of this study were to adapt a screening method previously used to assess seedling drought tolerance in cereals for use in cotton (Gossypium hirsutum L. and to identify tolerant accessions among a wide range of genotypes. Ninety genotypes were screened in seven growth chamber experiments. Fifteen-day-old seedlings were subjected to four 4-day drought cycles, and plant survival was evaluated after each cycle. Three cycles are probably the minimum required in cotton work. Significant differences (at the 0.05 level or lower among entries were obtained in four of the seven experiments. A "confirmation test" with entries previously evaluated as "tolerant" (high survival and "susceptible" (low survival was run. A number of entries duplicated their earlier performance, but others did not, which indicates the need to reevaluate selections. Germplasms considered tolerant included: `IAC-13-1', `IAC-RM4-SM5', `Minas Sertaneja', `Acala 1517E-1' and `4521'. In general, the technique is simple, though time-consuming, with practical value for screening a large number of genotypes. Results from the screening tests generally agreed with field information. The screening procedure is suitable to select tolerant accessions from among a large number of entries in germplasm collections as a preliminary step in breeding for drought tolerance. This research also demonstrated the need to characterize the internal lack of uniformity in growth chambers to allow for adequate designs of experiments.

  12. Tunisian Toxoplasma gondii strains genotyping by the use of AK69 marker

    Directory of Open Access Journals (Sweden)

    Aoun Karim

    2011-08-01

    Full Text Available Abstract Background Clinical manifestation due to infection by Toxoplasma gondii is closely linked to the infecting strain of the parasite. Several genetic markers are available to determinate its genotype but few of them are able to discriminate between the three predominant lineages, namely types I, II and III. The number of markers decreases when atypical, recombinant/mixed genotypes need to be identified. Findings In our study, the contribution of sequence polymorphisms in the AK69 gene as typing markers for T. gondii was investigated for the first time in an epidemiological study. The coding region of the marker was amplified, sequenced and aligned for different Toxoplasma strains. The identified nucleotide polymorphism at 12 positions was able to highly discriminate between the different congenital toxoplasmosis Tunisian strains. Moreover the high detection sensitivity level of the marker enabled unambiguous identification of mixed/recombinant genotypes directly. Conclusion It can be, thus, very useful for direct typing in areas where such genotypes are frequently encountered, mainly in the African continent.

  13. Genotypic diversity of root and shoot characteristics of

    Directory of Open Access Journals (Sweden)

    ali ganjali

    2009-06-01

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

  14. 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)

  15. Influence of ESAT-6 secretion system 1 (RD1) of Mycobacterium tuberculosis on the interaction between mycobacteria and the host immune system.

    Science.gov (United States)

    Majlessi, Laleh; Brodin, Priscille; Brosch, Roland; Rojas, Marie-Jésus; Khun, Huot; Huerre, Michel; Cole, Stewart T; Leclerc, Claude

    2005-03-15

    The chromosomal locus encoding the early secreted antigenic target, 6 kDa (ESAT-6) secretion system 1 of Mycobacterium tuberculosis, also referred to as "region of difference 1 (RD1)," is absent from Mycobacterium bovis bacillus Calmette-Guerin (BCG). In this study, using low-dose aerosol infection in mice, we demonstrate that BCG complemented with RD1 (BCG::RD1) displays markedly increased virulence which albeit does not attain that of M. tuberculosis H37Rv. Nevertheless, phenotypic and functional analyses of immune cells at the site of infection show that the capacity of BCG::RD1 to initiate recruitment/activation of immune cells is comparable to that of fully virulent H37Rv. Indeed, in contrast to the parental BCG, BCG::RD1 mimics H37Rv and induces substantial influx of activated (CD44highCD45RB(-)CD62L(-)) or effector (CD45RB(-)CD27(-)) T cells and of activated CD11c(+)CD11bhigh cells to the lungs of aerosol-infected mice. For the first time, using in vivo analysis of transcriptome of inflammatory cytokines and chemokines of lung interstitial CD11c+ cells, we show that in a low-dose aerosol infection model, BCG::RD1 triggered an activation/inflammation program comparable to that induced by H37Rv while parental BCG, due to its overattenuation, did not initiate the activation program in lung interstitial CD11c+ cells. Thus, products encoded by the ESAT-6 secretion system 1 of M. tuberculosis profoundly modify the interaction between mycobacteria and the host innate and adaptive immune system. These modifications can explain the previously described improved protective capacity of BCG::RD1 vaccine candidate against M. tuberculosis challenge.

  16. Non-Tuberculous Mycobacteria multispecies biofilms in cystic fibrosis: development of an in vitro Mycobacterium abscessus and Pseudomonas aeruginosa dual species biofilm model.

    Science.gov (United States)

    Rodríguez-Sevilla, Graciela; García-Coca, Marta; Romera-García, David; Aguilera-Correa, John Jairo; Mahíllo-Fernández, Ignacio; Esteban, Jaime; Pérez-Jorge, Concepción

    2018-04-01

    Lung disease in cystic fibrosis (CF) is characterized by the progressive colonization of the respiratory tract by different bacteria, which develop polymicrobial biofilms. In the past decades, there has been an increase in the number of CF patients infected with Non-Tuberculous Mycobacteria (NTM). Although Mycobacterium abscessus is the main NTM isolated globally, little is known about M. abscessus multispecies biofilm formation. In the present study we developed an in vitro model to study the phenotypic characteristics of biofilms formed by M. abscessus and Pseudomonas aeruginosa, a major pathogen in CF. For that purpose, dual species biofilms were grown on polycarbonate membranes with a fixed concentration of P. aeruginosa and different inoculums of M. abscessus. The biofilms were sampled at 24, 48, and 72 h and bacteria were quantified in specific media. The results revealed that the increasing initial concentration of M. abscessus in dual species biofilms had an effect on its population only at 24 and 48 h, whereas P. aeruginosa was not affected by the different concentrations used of M. abscessus. Time elapsed increased biofilm formation of both species, specially between 24 and 48 h. According to the results, the conditions to produce a mature dual species biofilm in which the relative species distribution remained stable were 72 h growth of the mixed microbial culture at a 1:1 ratio. A significant decrease in mycobacterial population in dual compared to single species biofilms was found, suggesting that P. aeruginosa has a negative influence on M. abscessus. Finally, in a proof of concept experiment, young and mature dual species biofilms were exposed to clarithromycin. Copyright © 2018 Elsevier GmbH. All rights reserved.

  17. ¹³C metabolic flux analysis identifies an unusual route for pyruvate dissimilation in mycobacteria which requires isocitrate lyase and carbon dioxide fixation.

    Directory of Open Access Journals (Sweden)

    Dany J V Beste

    2011-07-01

    Full Text Available Mycobacterium tuberculosis requires the enzyme isocitrate lyase (ICL for growth and virulence in vivo. The demonstration that M. tuberculosis also requires ICL for survival during nutrient starvation and has a role during steady state growth in a glycerol limited chemostat indicates a function for this enzyme which extends beyond fat metabolism. As isocitrate lyase is a potential drug target elucidating the role of this enzyme is of importance; however, the role of isocitrate lyase has never been investigated at the level of in vivo fluxes. Here we show that deletion of one of the two icl genes impairs the replication of Mycobacterium bovis BCG at slow growth rate in a carbon limited chemostat. In order to further understand the role of isocitrate lyase in the central metabolism of mycobacteria the effect of growth rate on the in vivo fluxes was studied for the first time using ¹³C-metabolic flux analysis (MFA. Tracer experiments were performed with steady state chemostat cultures of BCG or M. tuberculosis supplied with ¹³C labeled glycerol or sodium bicarbonate. Through measurements of the ¹³C isotopomer labeling patterns in protein-derived amino acids and enzymatic activity assays we have identified the activity of a novel pathway for pyruvate dissimilation. We named this the GAS pathway because it utilizes the Glyoxylate shunt and Anapleurotic reactions for oxidation of pyruvate, and Succinyl CoA synthetase for the generation of succinyl CoA combined with a very low flux through the succinate--oxaloacetate segment of the tricarboxylic acid cycle. We confirm that M. tuberculosis can fix carbon from CO₂ into biomass. As the human host is abundant in CO₂ this finding requires further investigation in vivo as CO₂ fixation may provide a point of vulnerability that could be targeted with novel drugs. This study also provides a platform for further studies into the metabolism of M. tuberculosis using ¹³C-MFA.

  18. Diversity, community composition, and dynamics of nonpigmented and late-pigmenting rapidly growing mycobacteria in an urban tap water production and distribution system.

    Science.gov (United States)

    Dubrou, S; Konjek, J; Macheras, E; Welté, B; Guidicelli, L; Chignon, E; Joyeux, M; Gaillard, J L; Heym, B; Tully, T; Sapriel, G

    2013-09-01

    Nonpigmented and late-pigmenting rapidly growing mycobacteria (RGM) have been reported to commonly colonize water production and distribution systems. However, there is little information about the nature and distribution of RGM species within the different parts of such complex networks or about their clustering into specific RGM species communities. We conducted a large-scale survey between 2007 and 2009 in the Parisian urban tap water production and distribution system. We analyzed 1,418 water samples from 36 sites, covering all production units, water storage tanks, and distribution units; RGM isolates were identified by using rpoB gene sequencing. We detected 18 RGM species and putative new species, with most isolates being Mycobacterium chelonae and Mycobacterium llatzerense. Using hierarchical clustering and principal-component analysis, we found that RGM were organized into various communities correlating with water origin (groundwater or surface water) and location within the distribution network. Water treatment plants were more specifically associated with species of the Mycobacterium septicum group. On average, M. chelonae dominated network sites fed by surface water, and M. llatzerense dominated those fed by groundwater. Overall, the M. chelonae prevalence index increased along the distribution network and was associated with a correlative decrease in the prevalence index of M. llatzerense, suggesting competitive or niche exclusion between these two dominant species. Our data describe the great diversity and complexity of RGM species living in the interconnected environments that constitute the water production and distribution system of a large city and highlight the prevalence index of the potentially pathogenic species M. chelonae in the distribution network.

  19. US Cystic Fibrosis Foundation and European Cystic Fibrosis Society consensus recommendations for the management of non-tuberculous mycobacteria in individuals with cystic fibrosis.

    Science.gov (United States)

    Floto, R Andres; Olivier, Kenneth N; Saiman, Lisa; Daley, Charles L; Herrmann, Jean-Louis; Nick, Jerry A; Noone, Peadar G; Bilton, Diana; Corris, Paul; Gibson, Ronald L; Hempstead, Sarah E; Koetz, Karsten; Sabadosa, Kathryn A; Sermet-Gaudelus, Isabelle; Smyth, Alan R; van Ingen, Jakko; Wallace, Richard J; Winthrop, Kevin L; Marshall, Bruce C; Haworth, Charles S

    2016-01-01

    Non-tuberculous mycobacteria (NTM) are ubiquitous environmental organisms that can cause chronic pulmonary infection, particularly in individuals with pre-existing inflammatory lung disease such as cystic fibrosis (CF). Pulmonary disease caused by NTM has emerged as a major threat to the health of individuals with CF but remains difficult to diagnose and problematic to treat. In response to this challenge, the US Cystic Fibrosis Foundation (CFF) and the European Cystic Fibrosis Society (ECFS) convened an expert panel of specialists to develop consensus recommendations for the screening, investigation, diagnosis and management of NTM pulmonary disease in individuals with CF. Nineteen experts were invited to participate in the recommendation development process. Population, Intervention, Comparison, Outcome (PICO) methodology and systematic literature reviews were employed to inform draft recommendations. An anonymous voting process was used by the committee to reach consensus. All committee members were asked to rate each statement on a scale of: 0, completely disagree, to 9, completely agree; with 80% or more of scores between 7 and 9 being considered 'good' agreement. Additionally, the committee solicited feedback from the CF communities in the USA and Europe and considered the feedback in the development of the final recommendation statements. Three rounds of voting were conducted to achieve 80% consensus for each recommendation statement. Through this process, we have generated a series of pragmatic, evidence-based recommendations for the screening, investigation, diagnosis and treatment of NTM infection in individuals with CF as an initial step in optimising management for this challenging condition. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/

  20. Identification of the Regulator Gene Responsible for the Acetone-Responsive Expression of the Binuclear Iron Monooxygenase Gene Cluster in Mycobacteria

    Science.gov (United States)

    Furuya, Toshiki; Hirose, Satomi; Semba, Hisashi; Kino, Kuniki

    2011-01-01

    The mimABCD gene cluster encodes the binuclear iron monooxygenase that oxidizes propane and phenol in Mycobacterium smegmatis strain MC2 155 and Mycobacterium goodii strain 12523. Interestingly, expression of the mimABCD gene cluster is induced by acetone. In this study, we investigated the regulator gene responsible for this acetone-responsive expression. In the genome sequence of M. smegmatis strain MC2 155, the mimABCD gene cluster is preceded by a gene designated mimR, which is divergently transcribed. Sequence analysis revealed that MimR exhibits amino acid similarity with the NtrC family of transcriptional activators, including AcxR and AcoR, which are involved in acetone and acetoin metabolism, respectively. Unexpectedly, many homologs of the mimR gene were also found in the sequenced genomes of actinomycetes. A plasmid carrying a transcriptional fusion of the intergenic region between the mimR and mimA genes with a promoterless green fluorescent protein (GFP) gene was constructed and introduced into M. smegmatis strain MC2 155. Using a GFP reporter system, we confirmed by deletion and complementation analyses that the mimR gene product is the positive regulator of the mimABCD gene cluster expression that is responsive to acetone. M. goodii strain 12523 also utilized the same regulatory system as M. smegmatis strain MC2 155. Although transcriptional activators of the NtrC family generally control transcription using the σ54 factor, a gene encoding the σ54 factor was absent from the genome sequence of M. smegmatis strain MC2 155. These results suggest the presence of a novel regulatory system in actinomycetes, including mycobacteria. PMID:21856847

  1. 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

  2. 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.

  3. 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.

  4. AMMI model in the analysis of genotype by environment interaction of conventionally and organically grown onion

    Directory of Open Access Journals (Sweden)

    Brdar-Jokanović Milka

    2016-01-01

    Full Text Available This study was aimed to assess the stability of direct yield components (bulb weight and number plot-1 and other yield contributing characteristics (bulb diameter, height and index, neck diameter and length, plant height, emergence and vegetation period in five commercial onion cultivars grown in conventional and organic environments, by employing additive main effect and multiplicative interaction (AMMI statistical model in data analysis. The two-year field trial organized in complete randomized blocks included the plots maintained in four regimes: mineral fertilization (conventional, without fertilization, fertilization with farmyard manure and with bacterial fertilizer (organic. Each treatment by year combination was considered as an environment. Analysis of variance of AMMI model calculated for the investigated traits showed that all sources of variation (genotypes, environments, genotype by environment interaction were highly significant. The largest proportions of the total sum of squares were encompassed by environments, except for emergence and bulb index with the pronounced effect of genotypes (67.26 and 52.54%, respectively and neck length with the genotype by environment interaction amounting 44.59%. Generally, the effects of the interactions were in the common range. The AMMI model with two axes was concluded as the best model for the investigated traits. Onions grown in conventional system outperformed the organic ones. However, good performance of the genotypes was accompanied with low stability across the environments and vice versa. Therefore breeding programs intended to develop cultivars adapted to alternative production systems should rely on the experiments set in the corresponding environments that include various combinations of genotypes and agro-technical procedures based on the principles of organic agriculture. [Projekat Ministarstva nauke Republike Srbije, br. TR 31059

  5. Phenotype, Genotype, and Drug Resistance in Subtype C HIV-1 Infection.

    Science.gov (United States)

    Derache, Anne; Wallis, Carole L; Vardhanabhuti, Saran; Bartlett, John; Kumarasamy, Nagalingeswaran; Katzenstein, David

    2016-01-15

    Virologic failure in subtype C is characterized by high resistance to first-line antiretroviral (ARV) drugs, including efavirenz, nevirapine, and lamivudine, with nucleoside resistance including type 2 thymidine analog mutations, K65R, a T69del, and M184V. However, genotypic algorithms predicting resistance are mainly based on subtype B viruses and may under- or overestimate drug resistance in non-B subtypes. To explore potential treatment strategies after first-line failure, we compared genotypic and phenotypic susceptibility of subtype C human immunodeficiency virus 1 (HIV-1) following first-line ARV failure. AIDS Clinical Trials Group 5230 evaluated patients failing an initial nonnucleoside reverse-transcriptase inhibitor (NNRTI) regimen in Africa and Asia, comparing the genotypic drug resistance and phenotypic profile from the PhenoSense (Monogram). Site-directed mutagenesis studies of K65R and T69del assessed the phenotypic impact of these mutations. Genotypic algorithms overestimated resistance to etravirine and rilpivirine, misclassifying 28% and 32%, respectively. Despite K65R with the T69del in 9 samples, tenofovir retained activity in >60%. Reversion of the K65R increased susceptibility to tenofovir and other nucleosides, while reversion of the T69del showed increased resistance to zidovudine, with little impact on other NRTI. Although genotype and phenotype were largely concordant for first-line drugs, estimates of genotypic resistance to etravirine and rilpivirine may misclassify subtype C isolates compared to phenotype. © The Author 2015. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

  6. Laboratory Information Management Software for genotyping workflows: applications in high throughput crop genotyping

    Directory of Open Access Journals (Sweden)

    Prasanth VP

    2006-08-01

    Full Text Available Abstract Background With the advances in DNA sequencer-based technologies, it has become possible to automate several steps of the genotyping process leading to increased throughput. To efficiently handle the large amounts of genotypic data generated and help with quality control, there is a strong need for a software system that can help with the tracking of samples and capture and management of data at different steps of the process. Such systems, while serving to manage the workflow precisely, also encourage good laboratory practice by standardizing protocols, recording and annotating data from every step of the workflow. Results A laboratory information management system (LIMS has been designed and implemented at the International Crops Research Institute for the Semi-Arid Tropics (ICRISAT that meets the requirements of a moderately high throughput molecular genotyping facility. The application is designed as modules and is simple to learn and use. The application leads the user through each step of the process from starting an experiment to the storing of output data from the genotype detection step with auto-binning of alleles; thus ensuring that every DNA sample is handled in an identical manner and all the necessary data are captured. The application keeps track of DNA samples and generated data. Data entry into the system is through the use of forms for file uploads. The LIMS provides functions to trace back to the electrophoresis gel files or sample source for any genotypic data and for repeating experiments. The LIMS is being presently used for the capture of high throughput SSR (simple-sequence repeat genotyping data from the legume (chickpea, groundnut and pigeonpea and cereal (sorghum and millets crops of importance in the semi-arid tropics. Conclusion A laboratory information management system is available that has been found useful in the management of microsatellite genotype data in a moderately high throughput genotyping

  7. Genotyping of Coxiella burnetii from domestic ruminants in northern Spain

    Directory of Open Access Journals (Sweden)

    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

  8. Audioprofiles and antioxidant enzyme genotypes in presbycusis.

    Science.gov (United States)

    Angeli, Simon I; Bared, Anthony; Ouyang, Xiaomei; Du, Li Lin; Yan, Denise; Zhong Liu, Xue

    2012-11-01

    Audiometric patterns have been shown to indirectly provide information regarding the pathophysiology of presbycusis and be useful in the phenotyping of hereditary deafness. Hospital-based cohort study of adults with presbycusis, comparing the association of audiometric patterns and polymorphisms of antioxidant enzymes that have been linked to presbycusis: GSTT1, GSTM1 and NAT2. All subjects underwent a clinical evaluation and completed questionnaires regarding ototoxicity and noise exposure. Pure-tone threshold audiometry was obtained and subjects' audiograms were classified into specific patterns. DNA was extracted from blood and the polymorphisms of GSTT1, GSTM1, and the NAT2 variants (NAT2* 5A; NAT2* 6A,B) were analyzed by PCR. The audiometric patterns that were more prevalent in our cohort were "High-Frequency Steeply Sloping" or HFSS (33%), "High-Frequency Gently Sloping" or HFGS (31%), and "Flat" (27%), with other patterns being rare. We did not find a statistical significant effect of gender, age, hearing level, and ear side on the audiometric pattern. Subjects with mutant alleles for GSTT1 were more likely to have a HFSS audiogram than subjects with the wild type genotype. In this cohort, there was a similar prevalence for the three audiometric configurations HFSS, HFGS, and Flat, with other configurations being rare. Subjects with mutant alleles for GSTT1 were more likely to have a HFSS audiogram than subjects with the wild type genotype, suggesting that the basal turn of the cochlea is susceptible to GSTT1 regulated oxidative stress. However, further studies of audioprofiles with larger sample sizes may be needed to establish phenotype-genotype correlations in presbycusis. Copyright © 2012 The American Laryngological, Rhinological, and Otological Society, Inc.

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

    Science.gov (United States)

    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. Relationship of some upland rice genotype after gamma irradiation

    Science.gov (United States)

    Suliartini, N. W. S.; Wijayanto, T.; Madiki, A.; Boer, D.; Muhidin; Juniawan

    2018-02-01

    The objective of the research was to group local upland rice genotypes after being treated with gamma irradiation. The research materials were upland rice genotypes resulted from mutation of the second generation and two parents: Pae Loilo (K3D0) and Pae Pongasi (K2D0) Cultivars. The research was conducted at the Indonesian Sweetener and Fiber Crops Research Institute, Malang Regency, and used the augmented design method. Research data were analyzed with R Program. Eight hundred and seventy one genotypes were selected with the selection criteria were based on yields on the average parents added 1.5 standard deviation. Based on the selection, eighty genotypes were analyzed with cluster analyses. Nine observation variables were used to develop cluster dendrogram using average linked method. Genetic distance was measured by euclidean distance. The results of cluster dendrogram showed that tested genotypes were divided into eight groups. Group 1, 2, 7, and 8 each had one genotype, group 3 and 6 each had two genotypes, group 4 had 25 genotypes, and group 5 had 51 genotypes. Check genotypes formed a separate group. Group 6 had the highest yield per plant of 126.11 gram, followed by groups 5 and 4 of 97.63 and 94.08 gram, respectively.

  11. Cow genotyping strategies for genomic selection in a small dairy cattle population.

    Science.gov (United States)

    Jenko, J; Wiggans, G R; Cooper, T A; Eaglen, S A E; Luff, W G de L; Bichard, M; Pong-Wong, R; Woolliams, J A

    2017-01-01

    This study compares how different cow genotyping strategies increase the accuracy of genomic estimated breeding values (EBV) in dairy cattle breeds with low numbers. In these breeds, few sires have progeny records, and genotyping cows can improve the accuracy of genomic EBV. The Guernsey breed is a small dairy cattle breed with approximately 14,000 recorded individuals worldwide. Predictions of phenotypes of milk yield, fat yield, protein yield, and calving interval were made for Guernsey cows from England and Guernsey Island using genomic EBV, with training sets including 197 de-regressed proofs of genotyped bulls, with cows selected from among 1,440 genotyped cows using different genotyping strategies. Accuracies of predictions were tested using 10-fold cross-validation among the cows. Genomic EBV were predicted using 4 different methods: (1) pedigree BLUP, (2) genomic BLUP using only bulls, (3) univariate genomic BLUP using bulls and cows, and (4) bivariate genomic BLUP. Genotyping cows with phenotypes and using their data for the prediction of single nucleotide polymorphism effects increased the correlation between genomic EBV and phenotypes compared with using only bulls by 0.163±0.022 for milk yield, 0.111±0.021 for fat yield, and 0.113±0.018 for protein yield; a decrease of 0.014±0.010 for calving interval from a low base was the only exception. Genetic correlation between phenotypes from bulls and cows were approximately 0.6 for all yield traits and significantly different from 1. Only a very small change occurred in correlation between genomic EBV and phenotypes when using the bivariate model. It was always better to genotype all the cows, but when only half of the cows were genotyped, a divergent selection strategy was better compared with the random or directional selection approach. Divergent selection of 30% of the cows remained superior for the yield traits in 8 of 10 folds. Copyright © 2017 American Dairy Science Association. Published by

  12. Genotyping of Trichomonas vaginalis isolates from women in Shahrekord city (Southwestern Iran

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    Khalili Bahman

    2017-01-01

    Full Text Available Trichomonas vaginalis is a causative agent of vaginitis in female and urethritis in men. It is primarily transmitted by sexually route. It is known that each geographical area has its own set of Trichomonas vaginalis strain. Parasite strains in each region have its specific characterizations and different strains of the parasite are able to cause various diseases with the acuity and severity. The aim of this study was to determine the genotyping of Trichomonas vaginalis strains in the Shahrekord city (Chaharmahal Va Bakhtiari province, southwest Iran. A total of 1725 vaginal samples were taken from clinically suspected women for Trichomonas vaginalis infection and 21 specimens were diagnosed as positive by direct smear wet mount and culture repeated passage of the parasite in the modified TYI-S-33 medium. The genomic DNA was extracted from each sample and the nested polymerase chain reaction was applied using specific oligonucleotide primers for actin gene amplification. Finally, the restriction fragment length polymorphism using RsaI, MseI, and HindII restriction enzymes were done on PCR products for genotyping. PCR-RFLP analysis of 21 positive cases (1.22% was showing the most frequent genotype was H (8 cases, followed by G (4 cases, E (3 cases, and P (2 cases. N and I genotypes were detected in each 1 case. Also, there was 2 cases mix (E and H genotype. The findings of the present work were showed 7 different genetic strains in isolated Trichomonas vaginalis from symptomatic and asymptomatic women in Shahrekord city. In this study high level of H genotype in referred women in Shahrekord city was observed and H, G, E, and I genotypes were may be related to burning and itching as well as H, P, and mix genotypes were associated with malodorous discharge with pelvic pain in this region of Iran. For a suggestion, it would be better in further studies the accurate determination of genetic diversity of this parasite done in Chaharmahal Va Bakhtiari

  13. NS3 protease polymorphisms and genetic barrier to drug resistance of distinct hepatitis C virus genotypes from worldwide treatment-naïve subjects.

    Science.gov (United States)

    Vidal, L L; Soares, M A; Santos, A F

    2016-11-01

    Hepatitis C virus (HCV) NS3 protease inhibitors have been primarily designed against genotype 1, the one with the lowest response to dual therapy. However, less evidence of their efficacy on non-1 genotypes is available, and any such information is mostly concentrated on genotypes 2-4. This study evaluated HCV protease resistance profiles in the major six HCV genotypes and identified genetic barrier (GB) profiles to each available protease inhibitor across HCV strains from different locations worldwide. We obtained 15 099 HCV sequences from treatment-naïve subjects retrieved at the Los Alamos HCV Sequence Database. The wild-type codons of different HCV genotypes were used to analyse the smallest number of nucleotide substitution steps required for changing that codon to the closest one associated with drug resistance. The 36L and 175L RAVs were found as genetic signatures of genotypes 2-5, while the 80K RAV was found in all genotype 5 sequences. Genotypes 4 and 6 showed a higher GB to RAV mutations conferring resistance to telaprevir, while genotypes 2-5 presented baseline resistance to that drug, carrying the 36L mutation. Genotype 4 had a higher GB to simeprevir resistance, requiring three substitutions to acquire the 155K mutation. Subtype 1b showed a higher GB than subtype 1a to resistance for most PIs, with RAVs at codons 36 and 155. Geographic disparities were also found in frequencies of certain RAVs in genotypes 2 and 3. Under a scenario of unprecedented evolution of anti-HCV direct-acting agents, the genetic composition of the circulating HCV sequences should be evaluated worldwide to choose the most appropriate/feasible therapeutic schemes with the highest genetic barriers to resistance. © 2016 John Wiley & Sons Ltd.

  14. Direct Democracy

    DEFF Research Database (Denmark)

    Beramendi, Virginia; Ellis, Andrew; Kaufman, Bruno

    While many books on direct democracy have a regional or national approach, or simply focus on one of the many mechanisms associated with direct democracy, this Handbook delves into a global comparison of direct democracy mechanisms, including referendums, citizens' initiatives, agenda initiatives...... learned. In addition, the uniquely comprehensive world survey outlines direct democracy provisions in 214 countries and territories and indicates which, if any, of these provisions are used by each country or territory at both the national and sub-national levels. Furthermore, the world survey includes...

  15. Global surveillance of emerging Influenza virus genotypes by mass spectrometry.

    Directory of Open Access Journals (Sweden)

    Rangarajan Sampath

    2007-05-01

    Full Text Available Effective influenza surveillance requires new methods capable of rapid and inexpensive genomic analysis of evolving viral species for pandemic preparedness, to understand the evolution of circulating viral species, and for vaccine strain selection. We have developed one such approach based on previously described broad-range reverse transcription PCR/electrospray ionization mass spectrometry (RT-PCR/ESI-MS technology.Analysis of base compositions of RT-PCR amplicons from influenza core gene segments (PB1, PB2, PA, M, NS, NP are used to provide sub-species identification and infer influenza virus H and N subtypes. Using this approach, we detected and correctly identified 92 mammalian and avian influenza isolates, representing 30 different H and N types, including 29 avian H5N1 isolates. Further, direct analysis of 656 human clinical respiratory specimens collected over a seven-year period (1999-2006 showed correct identification of the viral species and subtypes with >97% sensitivity and specificity. Base composition derived clusters inferred from this analysis showed 100% concordance to previously established clades. Ongoing surveillance of samples from the recent influenza virus seasons (2005-2006 showed evidence for emergence and establishment of new genotypes of circulating H3N2 strains worldwide. Mixed viral quasispecies were found in approximately 1% of these recent samples providing a view into viral evolution.Thus, rapid RT-PCR/ESI-MS analysis can be used to simultaneously identify all species of influenza viruses with clade-level resolution, identify mixed viral populations and monitor global spread and emergence of novel viral genotypes. This high-throughput method promises to become an integral component of influenza surveillance.

  16. Genotyping and phylogenetic analysis of Pneumocystis jirovecii isolates from India.

    Science.gov (United States)

    Gupta, Rashmi; Mirdha, Bijay Ranjan; Guleria, Randeep; Agarwal, Sanjay Kumar; Samantaray, Jyotish Chandra; Kumar, Lalit; Kabra, Sushil Kumar; Luthra, Kalpana; Sreenivas, Vishnubhatla

    2010-08-01

    Pneumocystis jirovecii is the cause of Pneumocystis pneumonia (PCP) in immuno-compromised individuals. The aim of this study was to describe the genotypes/haplotypes of P. jirovecii in immuno-compromised individuals with positive polymerase chain reaction (PCR) result for PCP. The typing was based on sequence polymorphism at internal transcribed spacer (ITS) regions of rRNA operon. Phylogenetic relationship between Indian and global haplotypes was also studied. Between January 2005 to October 2008, 43 patients were found to be positive for Pneumocystis using PCR targeting mitochondrial large subunit rRNA (mt LSU rRNA) and ITS region. Genotyping of all the positive samples was performed at the ITS locus by direct sequencing. Nine ITS1 alleles (all previously known) and 11 ITS2 alleles (nine previously defined and two new) were observed. A total of 19 ITS haplotypes, including five novel haplotypes (DEL1r, Edel2, Hr, Adel3 and SYD1a), were observed. The most prevalent type was SYD1g (16.3%), followed by types Ea (11.6%), Ec (9.3%), Eg (6.9%), DEL1r (6.9%), Ne (6.9%) and Ai (6.9%). To detect mixed infection, 30% of the positive isolates were cloned and 4-5 clones were sequenced from each specimen. Cloning and sequencing identified two more haplotypes in addition to the 19 types. Mixed infection was identified in 3 of the 13 cloned samples (23.1%). Upon construction of a haplotype network of 21 haplotypes, type Eg was identified as the most probable ancestral type. The present study is the first study that describes the haplotypes of P. jirovecii based on the ITS gene from India. The study suggests a high diversity of P. jirovecii haplotypes in the population. Copyright 2010 Elsevier B.V. All rights reserved.

  17. ABO Blood Group Genotyping by Real-time PCR in Kazakh Population

    Directory of Open Access Journals (Sweden)

    Pavel Tarlykov

    2014-12-01

    Full Text Available Introduction. ABO blood group genotyping is a new technology in hematology that helps prevent adverse transfusion reactions in patients. Identification of antigens on the surface of red blood cells is based on serology; however, genotyping employs a different strategy and is aimed directly at genes that determine the surface proteins. ABO blood group genotyping by real-time PCR has several crucial advantages over other PCR-based techniques, such as high rapidity and reliability of analysis. The purpose of this study was to examine nucleotide substitutions differences by blood types using a PCR-based method on Kazakh blood donors.Methods. The study was approved by the Ethics Committee of the National Center for Biotechnology. Venous blood samples from 369 healthy Kazakh blood donors, whose blood types had been determined by serological methods, were collected after obtaining informed consent. The phenotypes of the samples included blood group A (n = 99, B (n = 93, O (n = 132, and AB (n = 45. Genomic DNA was extracted using a salting-out method. PCR products of ABO gene were sequenced on an ABI 3730xl DNA analyzer (Applied Biosystems. The resulting nucleotide sequences were compared and aligned against reference sequence NM_020469.2. Real-time PCR analysis was performed on CFX96 Touch™ Real-Time PCR Detection System (BioRad.Results. Direct sequencing of ABO gene in 369 samples revealed that the vast majority of nucleotide substitutions that change the ABO phenotype were limited to exons 6 and 7 of the ABO gene at positions 261, 467, 657, 796, 803, 930 and 1,060. However, genotyping of only three of them (261, 796 and 803 resulted in identification of major ABO genotypes in the Kazakh population. As a result, TaqMan probe based real-time PCR assay for the specific detection of genotypes 261, 796 and 803 was developed. The assay did not take into account several other mutations that may affect the determination of blood group, because they have a

  18. Linking genotypes database with locus-specific database and genotype-phenotype correlation in phenylketonuria.

    Science.gov (United States)

    Wettstein, Sarah; Underhaug, Jarl; Perez, Belen; Marsden, Brian D; Yue, Wyatt W; Martinez, Aurora; Blau, Nenad

    2015-03-01

    The wide range of metabolic phenotypes in phenylketonuria is due to a large number of variants causing variable impairment in phenylalanine hydroxylase function. A total of 834 phenylalanine hydroxylase gene variants from the locus-specific database PAHvdb and genotypes of 4181 phenylketonuria patients from the BIOPKU database were characterized using FoldX, SIFT Blink, Polyphen-2 and SNPs3D algorithms. Obtained data was correlated with residual enzyme activity, patients' phenotype and tetrahydrobiopterin responsiveness. A descriptive analysis of both databases was compiled and an interactive viewer in PAHvdb database was implemented for structure visualization of missense variants. We found a quantitative relationship between phenylalanine hydroxylase protein stability and enzyme activity (r(s) = 0.479), between protein stability and allelic phenotype (r(s) = -0.458), as well as between enzyme activity and allelic phenotype (r(s) = 0.799). Enzyme stability algorithms (FoldX and SNPs3D), allelic phenotype and enzyme activity were most powerful to predict patients' phenotype and tetrahydrobiopterin response. Phenotype prediction was most accurate in deleterious genotypes (≈ 100%), followed by homozygous (92.9%), hemizygous (94.8%), and compound heterozygous genotypes (77.9%), while tetrahydrobiopterin response was correctly predicted in 71.0% of all cases. To our knowledge this is the largest study using algorithms for the prediction of patients' phenotype and tetrahydrobiopterin responsiveness in phenylketonuria patients, using data from the locus-specific and genotypes database.

  19. Chemical compounds related to nutraceutical and industrial qualities of non-transgenic soybean genotypes.

    Science.gov (United States)

    Carrera, Constanza S; Dardanelli, Julio L; Soldini, Diego O

    2014-05-01

    Information about the chemical profile of soybean seed is valuable for breeding programs aimed at obtaining value-added products to meet the demands of niche markets. The objective of this study was to determine seed composition of non-transgenic soybean genotypes with specialty characters in different environments of Argentina. Protein and oil contents ranged from 396 to 424 g kg⁻¹ and from 210 to 226 g kg⁻¹, respectively. Oleic and linolenic acid ratio, the general indicator of oil quality, varied from 2.7 to 3.8. The oil contained high levels of total tocopherols (1429-1558 mg kg⁻¹) and the meal exhibited high levels of total isoflavones (2.91-4.62 mg g⁻¹). The biplot showed that oleic, linoleic and linolenic acids, γ-, δ- and total tocopherols, genistin, malonyl daidzin and genistin, acetyl daidzin and glycitin and total isoflavones allowed the greatest discrimination among the genotypes studied. Different chemical profiles of each non-transgenic genotype analyzed were established and, therefore, their identity was defined. These results are important for breeders who intend to obtain new genotypes with improved meal and oil quality, as well as for processors and exporters, who could use them directly as raw material for soyfood processing for nutraceutical purposes. © 2013 Society of Chemical Industry.

  20. 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.

  1. Hepatitis C Virus Resistance Testing in Genotype 1: The Changing Role in Clinical Utility.

    Science.gov (United States)

    Molino, Suzanne; Martin, Michelle T

    2017-09-01

    To review the role and utility of baseline resistance testing with currently available and pipeline genotype 1 hepatitis C virus (HCV) treatment. Authors reviewed liver meeting abstracts for data on currently-available and pipeline genotype 1 retreatment regimens from January 1, 2015, to March 23, 2017. Additional trials were identified from a review of clinicaltrials.gov using the pipeline medication names. Authors identified reports of current and pipeline genotype 1 retreatment regimens. Seven references were clinical study results presented at the meetings of the American Association for the Study of Liver Diseases and the European Association for the Study of the Liver, and 2 studies were from clinicaltrials.gov . Retreatment trial data of currently available salvage regimens indicate that baseline NS5A resistance-associated substitutions (RASs) may decrease sustained virological response (SVR) rates when retreating with ledipasvir/sofosbuvir but are not affected when using elbasvir/grazoprevir + sofosbuvir + ribavirin, paritaprevir/ritonavir/ombitasvir + dasabuvir + sofosbuvir, or sofosbuvir/velpatasvir + ribavirin. Pipeline data indicate that baseline NS5A RASs do not affect SVR rates when retreating with sofosbuvir/velpatasvir/voxilaprevir or glecaprevir/pibrentasvir. Baseline resistance testing was used for decisional support for 3 clinical scenarios in patients with HCV genotype 1 infection at the time of manuscript submission. Pending the approval of 2 new direct-acting antiviral regimens in the third quarter of 2017, the rapidly evolving HCV treatment guidelines will likely reflect a decreased clinical utility for resistance testing.

  2. Bound volatile precursors in genotypes in the pedigree of 'Marion' blackberry (Rubus sp.).

    Science.gov (United States)

    Du, Xiaofen; Finn, Chad E; Qian, Michael C

    2010-03-24

    Glycosidically bound volatiles and precursors in genotypes representing the pedigree for 'Marion' blackberry were investigated over two growing seasons. The volatile precursors were isolated using a C18 solid-phase extraction column. After enzymatic hydrolysis, the released volatiles were analyzed using stir bar sorptive extraction gas chromatography-mass spectrometry (GC-MS) and direct microvial insert thermal desorption GC-MS. The most abundant volatile precursors in the genotypes were alcohols, followed by shikimic acid derivatives. High amounts of furanone glycosides were also detected, while norisoprenoids only existed in a small amount in blackberries. The volatile precursor composition in the genotypes in the 'Marion' pedigree was very similar to their free volatile distribution. 'Logan' and 'Olallie' predominantly had bound norisoprenoids. Wild 'Himalaya' predominated with terpene alcohol and furaneol glycosides, whereas 'Santiam' and 'Chehalem' contained a high level of terpene alcohol glycosides. A similar inheritance pattern was also observed for some volatile precursors in the genotypes in the 'Marion' pedigree. A high content of linalool, hydroxylinalool, and alpha-ionol glycosides in 'Olallie' and a low content in 'Chehalem' resulted in a moderate level in their offspring 'Marion', while a low content of (E)-linalool oxide precursor in 'Olallie' and a high content in 'Chehalem' also resulted in a moderate level in 'Marion'. However, the concentration of furaneol glycosides in 'Marion' exceeded that of its two parents.

  3. Ecological effects of aphid abundance, genotypic variation, and contemporary evolution on plants.

    Science.gov (United States)

    Turley, Nash E; Johnson, Marc T J

    2015-07-01

    Genetic variation and contemporary evolution within populations can shape the strength and nature of species interactions, but the relative importance of these forces compared to other ecological factors is unclear. We conducted a field experiment testing the effects of genotypic variation, abundance, and presence/absence of green peach aphids (Myzus persicae) on the growth, leaf nitrogen, and carbon of two plant species (Brassica napus and Solanum nigrum). Aphid genotype affected B. napus but not S. nigrum biomass explaining 20 and 7% of the total variation, respectively. Averaging across both plant species, the presence/absence of aphids had a 1.6× larger effect size (Cohen's d) than aphid genotype, and aphid abundance had the strongest negative effects on plant biomass explaining 29% of the total variation. On B. napus, aphid genotypes had different effects on leaf nitrogen depending on their abundance. Aphids did not influence leaf nitrogen in S. nigrum nor leaf carbon in either species. We conducted a second experiment in the field to test whether contemporary evolution could affect plant performance. Aphid populations evolved in as little as five generations, but the rate and direction of this evolution did not consistently vary between plant species. On one host species (B. napus), faster evolving populations had greater negative effects on host plant biomass, with aphid evolutionary rate explaining 23% of the variation in host plant biomass. Together, these results show that genetic variation and evolution in an insect herbivore can play important roles in shaping host plant ecology.

  4. BDNF rs6265 methylation and genotype interact on risk for schizophrenia.

    Science.gov (United States)

    Ursini, Gianluca; Cavalleri, Tommaso; Fazio, Leonardo; Angrisano, Tiziana; Iacovelli, Luisa; Porcelli, Annamaria; Maddalena, Giancarlo; Punzi, Giovanna; Mancini, Marina; Gelao, Barbara; Romano, Raffaella; Masellis, Rita; Calabrese, Francesca; Rampino, Antonio; Taurisano, Paolo; Di Giorgio, Annabella; Keller, Simona; Tarantini, Letizia; Sinibaldi, Lorenzo; Quarto, Tiziana; Popolizio, Teresa; Caforio, Grazia; Blasi, Giuseppe; Riva, Marco A; De Blasi, Antonio; Chiariotti, Lorenzo; Bollati, Valentina; Bertolino, Alessandro

    2016-01-01

    Epigenetic mechanisms can mediate gene-environment interactions relevant for complex disorders. The BDNF gene is crucial for development and brain plasticity, is sensitive to environmental stressors, such as hypoxia, and harbors the functional SNP rs6265 (Val(66)Met), which creates or abolishes a CpG dinucleotide for DNA methylation. We found that methylation at the BDNF rs6265 Val allele in peripheral blood of healthy subjects is associated with hypoxia-related early life events (hOCs) and intermediate phenotypes for schizophrenia in a distinctive manner, depending on rs6265 genotype: in ValVal individuals increased methylation is associated with exposure to hOCs and impaired working memory (WM) accuracy, while the opposite is true for ValMet subjects. Also, rs6265 methylation and hOCs interact in modulating WM-related prefrontal activity, another intermediate phenotype for schizophrenia, with an analogous opposite direction in the 2 genotypes. Consistently, rs6265 methylation has a different association with schizophrenia risk in ValVals and ValMets. The relationships of methylation with BDNF levels and of genotype with BHLHB2 binding likely contribute to these opposite effects of methylation. We conclude that BDNF rs6265 methylation interacts with genotype to bridge early environmental exposures to adult phenotypes, relevant for schizophrenia. The study of epigenetic changes in regions containing genetic variation relevant for human diseases may have beneficial implications for the understanding of how genes are actually translated into phenotypes.

  5. Antioxidant capacity of anthocyanins from acerola genotypes

    Directory of Open Access Journals (Sweden)

    Vera Lúcia Arroxelas Galvão De Lima

    2011-03-01

    Full Text Available Anthocyanins from 12 acerola genotypes cultivated at the Active Germplasm Bank at Federal Rural University of Pernambuco were isolated for antioxidant potential evaluation. The antioxidant activity and radical scavenging capacity of the anthocyanin isolates were measured according to the β-carotene bleaching method and 1,1-diphenyl-2-picrylhydrazyl (DPPH free radical scavenging assay, respectively. The antioxidant activity varied from 25.58 to 47.04% at 0.2 mg.mL-1, and it was measured using the β-carotene bleaching method. The free radical scavenging capacity increased according to the increase in concentration and reaction time by the DPPH assay. At 16.7 μg.mL-1 concentration and after 5 minutes and 2 hours reaction time, the percentage of scavenged radicals varied from 36.97 to 63.92% and 73.27 to 94.54%, respectively. Therefore, the antioxidant capacity of acerola anthocyanins varied amongst acerola genotypes and methods used. The anthocyanins present in this fruit may supply substantial dietary source of antioxidant which may promote health and produce disease prevention effects.

  6. [Mexican phenotype and genotype Vibrio cholerae 01].

    Science.gov (United States)

    Giono, S; Gutiérrez Cogno, L; Rodríguez Angeles, G; del Rio Zolezzi, A; Valdespino González, J L; Sepúlveda Amor, J

    1995-01-01

    This paper presents the phenotypical and genotypical characterization of 26922 Vibrio cholerae 01 strains isolated in Mexico from 1991 to 1993. All strains isolated were El Tor biovar. Strains were sensitive to antibiotics excluding furazolidone, streptomycin and sulfisoxasole to which we found resistance in 97% and we are using this characteristic as epidemiological markers. We detected a marked change in frequency of Inaba serotype from 1991, when it was dominant, with 99.5%, until 1992 when Ogawa serotype turned to be dominant with 95% of isolates. All Vibrio cholerae 01 strains, except one Ogawa strain, were to igenic, and V. choleraeno 01 were not toxigenic by ELISA, PCR and cell culture tests. Dominant ribotype was 5, but we found some strains with 6a pattern and two with ribotype 12. We are searching for ribotype 2 among hemolytic strains in order to learn if there is any relation to Gulf Coast strains prevalent in the USA, but until now we have not found any V. cholerae ribotype 2 in our isolates. Even if rapid tests are recommended for immediate diagnosis of cholera, it is necessary to continue bacterial isolation in order to have strains for phenotyping and genotyping studies that may support epidemiological analysis.

  7. HFE genotype affects exosome phenotype in cancer.

    Science.gov (United States)

    Mrowczynski, Oliver D; Madhankumar, A B; Slagle-Webb, Becky; Lee, Sang Y; Zacharia, Brad E; Connor, James R

    2017-08-01

    Neuroblastoma is the third most common childhood cancer, and timely diagnosis and sensitive therapeutic monitoring remain major challenges. Tumor progression and recurrence is common with little understanding of mechanisms. A major recent focus in cancer biology is the impact of exosomes on metastatic behavior and the tumor microenvironment. Exosomes have been demonstrated to contribute to the oncogenic effect on the surrounding tumor environment and also mediate resistance to therapy. The effect of genotype on exosomal phenotype has not yet been explored. We interrogated exosomes from human neuroblastoma cells that express wild-type or mutant forms of the HFE gene. HFE, one of the most common autosomal recessive polymorphisms in the Caucasian population, originally associated with hemochromatosis, has also been associated with increased tumor burden, therapeutic resistance boost, and negative impact on patient survival. Herein, we demonstrate that changes in genotype cause major differences in the molecular and functional properties of exosomes; specifically, HFE mutant derived exosomes have increased expression of proteins relating to invasion, angiogenesis, and cancer therapeutic resistance. HFE mutant derived exosomes were also shown to transfer this cargo to recipient cells and cause an increased oncogenic functionality in those recipient cells. Copyright © 2017. Published by Elsevier B.V.

  8. Haplotype-Based Genotyping in Polyploids

    Directory of Open Access Journals (Sweden)

    Josh P. Clevenger

    2018-04-01

    Full Text Available Accurate identification of polymorphisms from sequence data is crucial to unlocking the potential of high throughput sequencing for genomics. Single nucleotide polymorphisms (SNPs are difficult to accurately identify in polyploid crops due to the duplicative nature of polyploid genomes leading to low confidence in the true alignment of short reads. Implementing a haplotype-based method in contrasting subgenome-specific sequences leads to higher accuracy of SNP identification in polyploids. To test this method, a large-scale 48K SNP array (Axiom Arachis2 was developed for Arachis hypogaea (peanut, an allotetraploid, in which 1,674 haplotype-based SNPs were included. Results of the array show that 74% of the haplotype-based SNP markers could be validated, which is considerably higher than previous methods used for peanut. The haplotype method has been implemented in a standalone program, HAPLOSWEEP, which takes as input bam files and a vcf file and identifies haplotype-based markers. Haplotype discovery can be made within single reads or span paired reads, and can leverage long read technology by targeting any length of haplotype. Haplotype-based genotyping is applicable in all allopolyploid genomes and provides confidence in marker identification and in silico-based genotyping for polyploid genomics.

  9. Hearing impairment in genotyped Wolfram syndrome patients.

    Science.gov (United States)

    Plantinga, Rutger F; Pennings, Ronald J E; Huygen, Patrick L M; Bruno, Rocco; Eller, Philipp; Barrett, Timothy G; Vialettes, Bernard; Paquis-Fluklinger, Veronique; Lombardo, Fortunato; Cremers, Cor W R J

    2008-07-01

    Wolfram syndrome is a progressive neurodegenerative syndrome characterized by the features "DIDMOAD" (diabetes insipidus, diabetes mellitus, optic atrophy, and deafness). We sought to study the audiometric data of genotyped Wolfram syndrome patients with sensorineural hearing impairment. Pure tone threshold data of 23 Wolfram syndrome patients were used for cross-sectional analysis in subgroups (age less than 16 years or between 19 and 25 years, gender, and origin). All subgroups, with 1 exception, showed a fairly similar type of hearing impairment with, on average, thresholds of about 25 dB (range, 0 to 65 dB) at 0.25 to 1 kHz, gently sloping downward to about 60 dB (range, 25 to 95 dB) at 8 kHz. The subgroup of Dutch women, which was excluded from the calculations of the average hearing thresholds, showed a higher degree of hearing impairment. Only the latter subgroup showed progression; however, contrary to the previous longitudinal analysis, progression was not significant in the present cross-sectional analysis, presumably because of the high degree of cross-subject variability. This unique collection of audiometric data from genotyped Wolfram syndrome patients shows no substantial progression in sensorineural hearing impairment with advancing age, no relation to the types of WFS1 mutations identified, and, with exclusion of the subgroup of Dutch female patients, no significant sex-related differences.

  10. Directing 101.

    Science.gov (United States)

    Pintoff, Ernest

    Providing an introduction to anyone considering directing as a field of study or career, this book takes a broad look at the process of directing and encourages students and professionals alike to look outside of the movie industry for inspiration. Chapters in the book discuss selecting and acquiring material; budgeting and financing; casting and…

  11. Performance comparison between the mycobacteria growth indicator tube system and Löwenstein-Jensen medium in the routine detection of Mycobacterium tuberculosis at public health care facilities in Rio de Janeiro, Brazil: preliminary results of a pragmatic clinical trial.

    Science.gov (United States)

    Moreira, Adriana da Silva Rezende; Huf, Gisele; Vieira, Maria Armanda; Fonseca, Leila; Ricks, Monica; Kritski, Afrânio Lineu

    2013-01-01

    In view of the fact that the World Health Organization has recommended the use of the mycobacteria growth indicator tube (MGIT) 960 system for the diagnosis of tuberculosis and that there is as yet no evidence regarding the clinical impact of its use in health care systems, we conducted a pragmatic clinical trial to evaluate the clinical performance and cost-effectiveness of the use of MGIT 960 at two health care facilities in the city of Rio de Janeiro, Brazil, where the incidence of tuberculosis is high. Here, we summarize the methodology and preliminary results of the trial. (ISRCTN.org Identifier: ISRCTN79888843 [http://isrctn.org/]) In view of the fact that the World Health Organization has recommended the use of the mycobacteria growth indicator tube (MGIT) 960 system for the diagnosis of tuberculosis and that there is as yet no evidence regarding the clinical impact of its use in health care systems, we conducted a pragmatic clinical trial to evaluate the clinical performance and cost-effectiveness of the use of MGIT 960 at two health care facilities in the city of Rio de Janeiro, Brazil, where the incidence of tuberculosis is high. Here, we summarize the methodology and preliminary results of the trial. (ISRCTN.org Identifier: ISRCTN79888843 [http://isrctn.org/]).

  12. Deep sequencing analysis of HBV genotype shift and correlation with antiviral efficiency during adefovir dipivoxil therapy.

    Directory of Open Access Journals (Sweden)

    Yuwei Wang

    Full Text Available Viral genotype shift in chronic hepatitis B (CHB patients during antiviral therapy has been reported, but the underlying mechanism remains elusive.38 CHB patients treated with ADV for one year were selected for studying genotype shift by both deep sequencing and Sanger sequencing method.Sanger sequencing method found that 7.9% patients showed mixed genotype before ADV therapy. In contrast, all 38 patients showed mixed genotype before ADV treatment by deep sequencing. 95.5% mixed genotype rate was also obtained from additional 200 treatment-naïve CHB patients. Of the 13 patients with genotype shift, the fraction of the minor genotype in 5 patients (38% increased gradually during the course of ADV treatment. Furthermore, responses to ADV and HBeAg seroconversion were associated with the high rate of genotype shift, suggesting drug and immune pressure may be key factors to induce genotype shift. Interestingly, patients with genotype C had a significantly higher rate of genotype shift than genotype B. In genotype shift group, ADV treatment induced a marked enhancement of genotype B ratio accompanied by a reduction of genotype C ratio, suggesting genotype C may be more sensitive to ADV than genotype B. Moreover, patients with dominant genotype C may have a better therapeutic effect. Finally, genotype shifts was correlated with clinical improvement in terms of ALT.Our findings provided a rational explanation for genotype shift among ADV-treated CHB patients. The genotype and genotype shift might be associated with antiviral efficiency.

  13. The genotype-phenotype map of an evolving digital organism

    OpenAIRE

    Fortuna, Miguel A.; Zaman, Luis; Ofria, Charles; Wagner, Andreas

    2017-01-01

    To understand how evolving systems bring forth novel and useful phenotypes, it is essential to understand the relationship between genotypic and phenotypic change. Artificial evolving systems can help us understand whether the genotype-phenotype maps of natural evolving systems are highly unusual, and it may help create evolvable artificial systems. Here we characterize the genotype-phenotype map of digital organisms in Avida, a platform for digital evolution. We consider digital organisms fr...

  14. Molecular genotyping of HCV infection in seropositive blood donor

    Science.gov (United States)

    Zarin, Siti Noraziah Abu; Ibrahim, Nazlina

    2013-11-01

    This study is to investigate the prevalence of hepatitis C virus infection in seropositive blood donor. RNA was extracted from 32 positive samples in National Blood Centre and Melaka Hospital. The core and NS5B sequences were obtained from 23 samples. Genotype 3a is most prevalent in this study followed by genotype 1a. Evidence of mixed-genotypes (3a and 1b) infections was found in 5 subjects.

  15. Genomic Variants Revealed by Invariably Missing Genotypes in Nelore Cattle.

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    Joaquim Manoel da Silva

    Full Text Available High density genotyping panels have been used in a wide range of applications. From population genetics to genome-wide association studies, this technology still offers the lowest cost and the most consistent solution for generating SNP data. However, in spite of the application, part of the generated data is always discarded from final datasets based on quality control criteria used to remove unreliable markers. Some discarded data consists of markers that failed to generate genotypes, labeled as missing genotypes. A subset of missing genotypes that occur in the whole population under study may be caused by technical issues but can also be explained by the presence of genomic variations that are in the vicinity of the assayed SNP and that prevent genotyping probes from annealing. The latter case may contain relevant information because these missing genotypes might be used to identify population-specific genomic variants. In order to assess which case is more prevalent, we used Illumina HD Bovine chip genotypes from 1,709 Nelore (Bos indicus samples. We found 3,200 missing genotypes among the whole population. NGS re-sequencing data from 8 sires were used to verify the presence of genomic variations within their flanking regions in 81.56% of these missing genotypes. Furthermore, we discovered 3,300 novel SNPs/Indels, 31% of which are located in genes that may affect traits of importance for the genetic improvement of cattle production.

  16. Phosphorus use efficiency in pima cotton (Gossypium barbadense L. genotypes

    Directory of Open Access Journals (Sweden)

    Elcio Santos

    2015-06-01

    Full Text Available In the Brazilian Cerrado, P deficiency restricts cotton production, which requires large amounts of phosphate fertilizer. To improve the yield of cotton crops, genotypes with high P use efficiency must be identified and used. The present study evaluated P uptake and use efficiency of different Gossypium barbadense L. genotypes grown in the Cerrado. The experiment was carried out in a greenhouse with a completely randomized design, 15 x 2 factorial treatment structure (15 genotypes x 2 P levels, and four replicates. The genotypes were MT 69, MT 70, MT 87, MT 91, MT 92, MT 94, MT 101, MT 102, MT 103, MT 105, MT 106, MT 110, MT 112, MT 124, and MT 125; P levels were sufficient (1000 mg pot-1, PS treatment or deficient (PD treatment. Dry matter (DM and P levels were determined in cotton plant parts and used to calculate plant P content and use efficiency. In general, DM and P content were higher in the PS than in the PD treatment, with the exception of root DM and total DM in some genotypes. Genotypes also differed in terms of P uptake and use capacity. In the PS treatment, genotypes MT 92 and MT 102 had the highest response to phosphate fertilization. Genotype MT 69 exhibited the most efficient P uptake in the PD treatment. Genotype MT 124 showed the best shoot physiological efficiency, apparent recovery efficiency, and utilization efficiency, whereas MT 110 exhibited the highest root physiological efficiency.

  17. Intact molecular characterization of cord factor (trehalose 6,6'-dimycolate) from nine species of mycobacteria by MALDI-TOF mass spectrometry.

    Science.gov (United States)

    Fujita, Yukiko; Naka, Takashi; McNeil, Michael R; Yano, Ikuya

    2005-10-01

    Cord factor (trehalose 6,6'-dimycolate, TDM) is an unique glycolipid with a trehalose and two molecules of mycolic acids in the mycobacterial cell envelope. Since TDM consists of two molecules of very long branched-chain 3-hydroxy fatty acids, the molecular mass ranges widely and in a complex manner. To characterize the molecular structure of TDM precisely and simply, an attempt was made to determine the mycolic acid subclasses of TDM and the molecular species composition of intact TDM by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry for the first time. The results showed that less than 1 microg mycolic acid methyl ester of TDM from nine representative species of mycobacteria and TDM from the same species was sufficient to obtain well-resolved mass spectra composed of pseudomolecular ions [M+Na]+. Although the mass ion distribution was extremely diverse, the molecular species of each TDM was identified clearly by constructing a molecular ion matrix consisting of the combination of two molecules of mycolic acids. The results showed a marked difference in the molecular structure of TDM among mycobacterial species and subspecies. TDM from Mycobacterium tuberculosis (H37Rv and Aoyama B) showed a distinctive mass pattern and consisted of over 60 molecular ions with alpha-, methoxy- and ketomycolate. TDM from Mycobacterium bovis BCG Tokyo 172 similarly showed over 35 molecular ions, but that from M. bovis BCG Connaught showed simpler molecular ion clusters consisting of less than 35 molecular species due to a complete lack of methoxymycolate. Mass ions due to TDM from M. bovis BCG Connaught and Mycobacterium kansasii showed a biphasic distribution, but the two major peaks of TDM from M. kansasii were shifted up two or three carbon units higher compared with M. bovis BCG Connaught. Within the rapid grower group, in TDM consisting of alpha-, keto- and wax ester mycolate from Mycobacterium phlei and Mycobacterium flavescens, the

  18. 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.

  19. Directing Creativity

    DEFF Research Database (Denmark)

    Darsø, Lotte; Ibbotson, Piers

    2008-01-01

    In this article we argue that leaders facing complex challenges can learn from the arts, specifically that leaders can learn by examining how theatre directors direct creativity through creative constraints. We suggest that perceiving creativity as a boundary phenomenon is helpful for directing it....... Like leaders, who are caught in paradoxical situations where they have to manage production and logistics simultaneously with making space for creativity and innovation, theatre directors need to find the delicate balance between on one hand renewal of perceptions, acting and interaction...... and on the other hand getting ready for the opening night. We conclude that the art of directing creativity is linked to developing competencies of conscious presence, attention and vigilance, whereas the craft of directing creativity concerns communication, framing and choice....

  20. Direct Heat

    Energy Technology Data Exchange (ETDEWEB)

    Lienau, P J

    1990-01-01

    Potential resources and applications of earth heat in the form of geothermal energy are large. United States direct uses amount to 2,100 MWt thermal and worldwide 8,850 MWt above a reference temperature of 35 degrees Celsius. Space and district heating are the major direct uses of geothermal energy. Equipment employed in direct use projects is of standard manufacture and includes downhole and circulation pumps, transmission and distribution pipelines, heat exchangers and convectors, heat pumps and chillers. Direct uses of earth heat discussed are space and district heating, greenhouse heating and fish farming, process and industrial applications. The economic feasibility of direct use projects is governed by site specific factors such as location of user and resource, resource quality, system load factor and load density, as well as financing. Examples are presented of district heating in Klamath Falls, and Elko. Further developments of direct uses of geothermal energy will depend on matching user needs to the resource, and improving load factors and load density.

  1. Plant genotype, microbial recruitment and nutritional security.

    Science.gov (United States)

    Patel, Jai S; Singh, Akanksha; Singh, Harikesh B; Sarma, Birinchi K

    2015-01-01

    Agricultural food products with high nutritional value should always be preferred over food products with low nutritional value. Efforts are being made to increase nutritional value of food by incorporating dietary supplements to the food products. The same is more desirous if the nutritional value of food is increased under natural environmental conditions especially in agricultural farms. Fragmented researches have demonstrated possibilities in achieving the same. The rhizosphere is vital in this regard for not only health and nutritional status of plants but also for the microorganisms colonizing the rhizosphere. Remarkably robust composition of plant microbiome with respect to other soil environments clearly suggests the role of a plant host in discriminating its colonizers (Zancarini et al., 2012). A large number of biotic and abiotic factors are believed to manipulate the microbial communities in the rhizosphere. However, plant genotype has proven to be the key in giving the final shape of the rhizosphere microbiome (Berendsen et al., 2012; Marques et al., 2014).

  2. Sources of Wilhelm Johannsen's genotype theory.

    Science.gov (United States)

    Roll-Hansen, Nils

    2009-01-01

    This paper describes the historical background and early formation of Wilhelm Johannsen's distinction between genotype and phenotype. It is argued that contrary to a widely accepted interpretation (For instance, W. Provine, 1971. The Origins of Theoretical Population Genetics. Chicago: The University of Chicago Press; Mayr, 1973; F. B. Churchill, 1974. Journal of the History of Biology 7: 5-30; E. Mayr, 1982. The Growth of Biological Thought, Cambridge: Harvard University Press; J. Sapp, 2003. Genesis. The Evolution of Biology. New York: Oxford University Press) his concepts referred primarily to properties of individual organisms and not to statistical averages. Johannsen's concept of genotype was derived from the idea of species in the tradition of biological systematics from Linnaeus to de Vries: An individual belonged to a group - species, subspecies, elementary species - by representing a certain underlying type (S. Müller-Wille and V. Orel, 2007. Annals of Science 64: 171-215). Johannsen sharpened this idea theoretically in the light of recent biological discoveries, not least those of cytology. He tested and confirmed it experimentally combining the methods of biometry, as developed by Francis Galton, with the individual selection method and pedigree analysis, as developed for instance by Louis Vilmorin. The term "genotype" was introduced in W. Johannsen's 1909 (Elemente der Exakten Erblichkeitslehre. Jena: Gustav Fischer) treatise, but the idea of a stable underlying biological "type" distinct from observable properties was the core idea of his classical bean selection experiment published 6 years earlier (W. Johannsen, 1903. Ueber Erblichkeit in Populationen und reinen Linien. Eine Beitrag zur Beleuchtung schwebender Selektionsfragen, Jena: Gustav Fischer, pp. 58-59). The individual ontological foundation of population analysis was a self-evident presupposition in Johannsen's studies of heredity in populations from their start in the early 1890s till his

  3. Enterovirus genotypes causing hand foot and mouth disease in Shanghai, China: a molecular epidemiological analysis

    Science.gov (United States)

    2013-01-01

    Background A rapid expansion of hand, foot, and mouth disease (HFMD) outbreaks has occurred and caused deaths in China in recent years, but little is known about the other etiologic agents except enterovirus 71 (EV71) and coxsackievirus A 16 (CA16). The objective of this study is to determine the genotype compositions of enterovirus causing HFMD in Shanghai and identify any associations between enterovirus types and clinical manifestations. Methods Stool specimens were collected from patients hospitalized for treatment of HFMD, from May 2010 to April 2011. Enterovirus was detected by reverse transcription PCR and directly genotyped by sequencing the PCR products. Phylogenetic analysis was based on the VP1 partial gene. Results Of 290 specimens, 277 (95.5%) tested positive for enterovirus. The major genotypes were EV71 (63.8%), CA10 (9.0%), CA6 (8.3%), CA16 (6.9%), CA12 (2.4%), and CA4 (1.4%). The EV71 strains belonged to the C4a subtype and CA16 belonged to the B subtype. CA6 was closely related to strains detected in Japan, Taiwan and China, and CA10, CA12 and CA4 were phylogenetically similar to other strains circulating in China. Mean hospital stays and the prevalence of complications in patients with EV71 infection were higher than those in patients in CA6, CA10 or CA16 infection (P enterovirus genotypes. It deserves our attention as early identification of enterovirus genotypes is important for diagnosis and treatment of HFMD patients. PMID:24148902

  4. Validation of Genotyping-By-Sequencing Analysis in Populations of Tetraploid Alfalfa by 454 Sequencing

    Science.gov (United States)

    Rocher, Solen; Jean, Martine; Castonguay, Yves; Belzile, François

    2015-01-01

    Genotyping-by-sequencing (GBS) is a relatively low-cost high throughput genotyping technology based on next generation sequencing and is applicable to orphan species with no reference genome. A combination of genome complexity reduction and multiplexing with DNA barcoding provides a simple and affordable way to resolve allelic variation between plant samples or populations. GBS was performed on ApeKI libraries using DNA from 48 genotypes each of two heterogeneous populations of tetraploid alfalfa (Medicago sativa spp. sativa): the synthetic cultivar Apica (ATF0) and a derived population (ATF5) obtained after five cycles of recurrent selection for superior tolerance to freezing (TF). Nearly 400 million reads were obtained from two lanes of an Illumina HiSeq 2000 sequencer and analyzed with the Universal Network-Enabled Analysis Kit (UNEAK) pipeline designed for species with no reference genome. Following the application of whole dataset-level filters, 11,694 single nucleotide polymorphism (SNP) loci were obtained. About 60% had a significant match on the Medicago truncatula syntenic genome. The accuracy of allelic ratios and genotype calls based on GBS data was directly assessed using 454 sequencing on a subset of SNP loci scored in eight plant samples. Sequencing depth in this study was not sufficient for accurate tetraploid allelic dosage, but reliable genotype calls based on diploid allelic dosage were obtained when using additional quality filtering. Principal Component Analysis of SNP loci in plant samples revealed that a small proportion (<5%) of the genetic variability assessed by GBS is able to differentiate ATF0 and ATF5. Our results confirm that analysis of GBS data using UNEAK is a reliable approach for genome-wide discovery of SNP loci in outcrossed polyploids. PMID:26115486

  5. Influence of Genotype on Warfarin Maintenance Dose Predictions Produced Using a Bayesian Dose Individualization Tool.

    Science.gov (United States)

    Saffian, Shamin M; Duffull, Stephen B; Roberts, Rebecca L; Tait, Robert C; Black, Leanne; Lund, Kirstin A; Thomson, Alison H; Wright, Daniel F B

    2016-12-01

    A previously established Bayesian dosing tool for warfarin was found to produce biased maintenance dose predictions. In this study, we aimed (1) to determine whether the biased warfarin dose predictions previously observed could be replicated in a new cohort of patients from 2 different clinical settings, (2) to explore the influence of CYP2C9 and VKORC1 genotype on predictive performance of the Bayesian dosing tool, and (3) to determine whether the previous population used to develop the kinetic-pharmacodynamic model underpinning the Bayesian dosing tool was sufficiently different from the test (posterior) population to account for the biased dose predictions. The warfarin maintenance doses for 140 patients were predicted using the dosing tool and compared with the observed maintenance dose. The impact of genotype was assessed by predicting maintenance doses with prior parameter values known to be altered by genetic variability (eg, EC50 for VKORC1 genotype). The prior population was evaluated by fitting the published kinetic-pharmacodynamic model, which underpins the Bayesian tool, to the observed data using NONMEM and comparing the model parameter estimates with published values. The Bayesian tool produced positively biased dose predictions in the new cohort of patients (mean prediction error [95% confidence interval]; 0.32 mg/d [0.14-0.5]). The bias was only observed in patients requiring ≥7 mg/d. The direction and magnitude of the observed bias was not influenced by genotype. The prior model provided a good fit to our data, which suggests that the bias was not caused by different prior and posterior populations. Maintenance doses for patients requiring ≥7 mg/d were overpredicted. The bias was not due to the influence of genotype nor was it related to differences between the prior and posterior populations. There is a need for a more mechanistic model that captures warfarin dose-response relationship at higher warfarin doses.

  6. Automated genotyping of dinucleotide repeat markers

    Energy Technology Data Exchange (ETDEWEB)

    Perlin, M.W.; Hoffman, E.P. [Carnegie Mellon Univ., Pittsburgh, PA (United States)]|[Univ. of Pittsburgh, PA (United States)

    1994-09-01

    The dinucleotide repeats (i.e., microsatellites) such as CA-repeats are a highly polymorphic, highly abundant class of PCR-amplifiable markers that have greatly streamlined genetic mapping experimentation. It is expected that over 30,000 such markers (including tri- and tetranucleotide repeats) will be characterized for routine use in the next few years. Since only size determination, and not sequencing, is required to determine alleles, in principle, dinucleotide repeat genotyping is easily performed on electrophoretic gels, and can be automated using DNA sequencers. Unfortunately, PCR stuttering with these markers generates not one band for each allele, but a pattern of bands. Since closely spaced alleles must be disambiguated by human scoring, this poses a key obstacle to full automation. We have developed methods that overcome this obstacle. Our model is that the observed data is generated by arithmetic superposition (i.e., convolution) of multiple allele patterns. By quantitatively measuring the size of each component band, and exploiting the unique stutter pattern associated with each marker, closely spaced alleles can be deconvolved; this unambiguously reconstructs the {open_quotes}true{close_quotes} allele bands, with stutter artifact removed. We used this approach in a system for automated diagnosis of (X-linked) Duchenne muscular dystrophy; four multiplexed CA-repeats within the dystrophin gene were assayed on a DNA sequencer. Our method accurately detected small variations in gel migration that shifted the allele size estimate. In 167 nonmutated alleles, 89% (149/167) showed no size variation, 9% (15/167) showed 1 bp variation, and 2% (3/167) showed 2 bp variation. We are currently developing a library of dinucleotide repeat patterns; together with our deconvolution methods, this library will enable fully automated genotyping of dinucleotide repeats from sizing data.

  7. Directed polymers versus directed percolation

    Science.gov (United States)

    Halpin-Healy, Timothy

    1998-10-01

    Universality plays a central role within the rubric of modern statistical mechanics, wherein an insightful continuum formulation rises above irrelevant microscopic details, capturing essential scaling behaviors. Nevertheless, occasions do arise where the lattice or another discrete aspect can constitute a formidable legacy. Directed polymers in random media, along with its close sibling, directed percolation, provide an intriguing case in point. Indeed, the deep blood relation between these two models may have sabotaged past efforts to fully characterize the Kardar-Parisi-Zhang universality class, to which the directed polymer belongs.

  8. Improved Ancestry Estimation for both Genotyping and Sequencing Data using Projection Procrustes Analysis and Genotype Imputation

    Science.gov (United States)

    Wang, Chaolong; Zhan, Xiaowei; Liang, Liming; Abecasis, Gonçalo R.; Lin, Xihong

    2015-01-01

    Accurate estimation of individual ancestry is important in genetic association studies, especially when a large number of samples are collected from multiple sources. However, existing approaches developed for genome-wide SNP data do not work well with modest amounts of genetic data, such as in targeted sequencing or exome chip genotyping experiments. We propose a statistical framework to estimate individual ancestry in a principal component ancestry map generated by a reference set of individuals. This framework extends and improves upon our previous method for estimating ancestry using low-coverage sequence reads (LASER 1.0) to analyze either genotyping or sequencing data. In particular, we introduce a projection Procrustes analysis approach that uses high-dimensional principal components to estimate ancestry in a low-dimensional reference space. Using extensive simulations and empirical data examples, we show that our new method (LASER 2.0), combined with genotype imputation on the reference individuals, can substantially outperform LASER 1.0 in estimating fine-scale genetic ancestry. Specifically, LASER 2.0 can accurately estimate fine-scale ancestry within Europe using either exome chip genotypes or targeted sequencing data with off-target coverage as low as 0.05×. Under the framework of LASER 2.0, we can estimate individual ancestry in a shared reference space for samples assayed at different loci or by different techniques. Therefore, our ancestry estimation method will accelerate discovery in disease association studies not only by helping model ancestry within individual studies but also by facilitating combined analysis of genetic data from multiple sources. PMID:26027497

  9. Genotypic and phenotypic characterization of Chikungunya virus of different genotypes from Malaysia.

    Directory of Open Access Journals (Sweden)

    I-Ching Sam

    Full Text Available BACKGROUND: Mosquito-borne Chikungunya virus (CHIKV has recently re-emerged globally. The epidemic East/Central/South African (ECSA strains have spread for the first time to Asia, which previously only had endemic Asian strains. In Malaysia, the ECSA strain caused an extensive nationwide outbreak in 2008, while the Asian strains only caused limited outbreaks prior to this. To gain insight into these observed epidemiological differences, we compared genotypic and phenotypic characteristics of CHIKV of Asian and ECSA genotypes isolated in Malaysia. METHODS AND FINDINGS: CHIKV of Asian and ECSA genotypes were isolated from patients during outbreaks in Bagan Panchor in 2006, and Johor in 2008. Sequencing of the CHIKV strains revealed 96.8% amino acid similarity, including an unusual 7 residue deletion in the nsP3 protein of the Asian strain. CHIKV replication in cells and Aedes mosquitoes was measured by virus titration. There were no differences in mammalian cell lines. The ECSA strain reached significantly higher titres in Ae. albopictus cells (C6/36. Both CHIKV strains infected Ae. albopictus mosquitoes at a higher rate than Ae. aegypti, but when compared to each other, the ECSA strain had much higher midgut infection and replication, and salivary gland dissemination, while the Asian strain infected Ae. aegypti at higher rates. CONCLUSIONS: The greater ability of the ECSA strain to replicate in Ae. albopictus may explain why it spread far more quickly and extensively in humans in Malaysia than the Asian strain ever did, particularly in rural areas where Ae. albopictus predominates. Intergenotypic genetic differences were found at E1, E2, and nsP3 sites previously reported to be determinants of host adaptability in alphaviruses. Transmission of CHIKV in humans is influenced by virus strain and vector species, which has implications for regions with more than one circulating CHIKV genotype and Aedes species.

  10. Methods for discovering and validating relationships among genotyped animals

    Science.gov (United States)

    Genomic selection based on single-nucleotide polymorphisms (SNPs) has led to the collection of genotypes for over 2.2 million animals by the Council on Dairy Cattle Breeding in the United States. To assure that a genotype is assigned to the correct animal and that the animal’s pedigree is correct, t...

  11. Variation of meat quality traits among five genotypes of chicken.

    Science.gov (United States)

    Tang, H; Gong, Y Z; Wu, C X; Jiang, J; Wang, Y; Li, K

    2009-10-01

    The main objective of this study was to examine the diversity of meat quality traits among 5 chicken genotypes. The genotypes included 2 Chinese native breeds (Wenchang,WCH, and Xianju), 1 commercial broiler line (Avian, AV), 1 commercial layer line (Hy-Line Brown, HLB), and 1 Chinese commercial broiler line (Lingnanhuang, LNH) synthesized by exotic and native breeds, which were slaughtered at their market ages: 16, 7, 16, and 8 wk, respectively. The effects of genotype, muscle type, and sex on meat quality traits were examined. Birds from slow-growing genotypes (WCH, Xianju, and HLB) exhibited higher shear value, inosine-5'-monophosphate concentration, lower cook loss, and more fat than those from fast-growing genotypes (AV and LNH). Chickens from WCH possessed the lowest expressible moisture, cook loss, and the highest lipid (%) among the 3 slow-growing genotypes. The HLB birds were intermediate in expressible moisture and cook loss and lowest in lipid among all genotypes. The LNH cross birds were similar to AV broilers in most meat quality parameters, although they had a lower shear force value and higher fat content than AV broilers. Breast muscle had higher expressible moisture, shear force, protein (%), inosine-5'-monophosphate content, lower cook loss, and lipid (%) than leg muscle. Muscles from male chickens had higher expressible moisture than those from the females. Variability of meat quality characteristics is mainly related to genotype and muscle type differences.

  12. Screening of Wheat Genotypes for Boron Efficiency in Bangladesh

    Science.gov (United States)

    A number of Bangladeshi wheat genotypes (varieties and advanced lines) have been tested for boron efficiency through sand culture experiments over two years (2007-08 & 2008-09) against two Thai check varieties ‘Fang 60’ (boron efficient) and ‘SW41’ (boron inefficient). Performances of the genotypes ...

  13. Crossbreeding of large white and Nsukka local pigs: Genotype and ...

    African Journals Online (AJOL)

    5-Blood samples from 80 pigs of 4 genotypes - the Nsukka local (Lo), the exotic Large White (LW), the one-way F1 (LW x Lo) and the F2 crosses belonging to 5 age groups, were analyzed, to determine the mean values of the haematological parameters in the genotypes and different age groups and to check if and how the ...

  14. Enhanced fodder yield of maize genotypes under saline irrigation is ...

    African Journals Online (AJOL)

    Poor quality irrigation water adversely affects the growth and yield of crops. This study was designed to evaluate the growth, fodder yield and ionic concentration of three promising maize (Zea mays L.) genotypes under the influence of varying quality irrigation water, with different salinity levels. The genotypes, such as ...

  15. Breeding of a Tomato Genotype Readily Accessible to Genetic Manipulation

    NARCIS (Netherlands)

    Koornneef, Maarten; Hanhart, Corrie; Jongsma, Maarten; Toma, Ingrid; Weide, Rob; Zabel, Pim; Hille, Jacques

    1986-01-01

    A tomato genotype, superior in regenerating plants from cell cultures, was obtained by transferring regeneration capacity from Lycopersicon peruvianum into L. esculentum by classical breeding. This genotype, MsK93, greatly facilitates genetic manipulation of tomato, as was demonstrated by successful

  16. Application of mixed models for the assessment genotype and ...

    African Journals Online (AJOL)

    Application of mixed models for the assessment genotype and environment interactions in cotton ( Gossypium hirsutum ) cultivars in Mozambique. ... The cultivars ISA 205, STAM 42 and REMU 40 showed superior productivity when they were selected by the Harmonic Mean of Genotypic Values (HMGV) criterion in relation ...

  17. The influence of host genotype X environment Interactions on the ...

    African Journals Online (AJOL)

    Mean squares for environments, genotypes and G x E interactions were highly significant (P<0.0001) for anthracnose infection. Significant G x E interactions, accounting for 19% of the treatment sums of squares, indicated that genotypes responded differentially to anthracnose infection across environments. The additive ...

  18. The genotype-phenotype map of an evolving digital organism.

    Directory of Open Access Journals (Sweden)

    Miguel A Fortuna

    2017-02-01

    Full Text Available To understand how evolving systems bring forth novel and useful phenotypes, it is essential to understand the relationship between genotypic and phenotypic change. Artificial evolving systems can help us understand whether the genotype-phenotype maps of natural evolving systems are highly unusual, and it may help create evolvable artificial systems. Here we characterize the genotype-phenotype map of digital organisms in Avida, a platform for digital evolution. We consider digital organisms from a vast space of 10141 genotypes (instruction sequences, which can form 512 different phenotypes. These phenotypes are distinguished by different Boolean logic functions they can compute, as well as by the complexity of these functions. We observe several properties with parallels in natural systems, such as connected genotype networks and asymmetric phenotypic transitions. The likely common cause is robustness to genotypic change. We describe an intriguing tension between phenotypic complexity and evolvability that may have implications for biological evolution. On the one hand, genotypic change is more likely to yield novel phenotypes in more complex organisms. On the other hand, the total number of novel phenotypes reachable through genotypic change is highest for organisms with simple phenotypes. Artificial evolving systems can help us study aspects of biological evolvability that are not accessible in vastly more complex natural systems. They can also help identify properties, such as robustness, that are required for both human-designed artificial systems and synthetic biological systems to be evolvable.

  19. The genotype-phenotype map of an evolving digital organism.

    Science.gov (United States)

    Fortuna, Miguel A; Zaman, Luis; Ofria, Charles; Wagner, Andreas

    2017-02-01

    To understand how evolving systems bring forth novel and useful phenotypes, it is essential to understand the relationship between genotypic and phenotypic change. Artificial evolving systems can help us understand whether the genotype-phenotype maps of natural evolving systems are highly unusual, and it may help create evolvable artificial systems. Here we characterize the genotype-phenotype map of digital organisms in Avida, a platform for digital evolution. We consider digital organisms from a vast space of 10141 genotypes (instruction sequences), which can form 512 different phenotypes. These phenotypes are distinguished by different Boolean logic functions they can compute, as well as by the complexity of these functions. We observe several properties with parallels in natural systems, such as connected genotype networks and asymmetric phenotypic transitions. The likely common cause is robustness to genotypic change. We describe an intriguing tension between phenotypic complexity and evolvability that may have implications for biological evolution. On the one hand, genotypic change is more likely to yield novel phenotypes in more complex organisms. On the other hand, the total number of novel phenotypes reachable through genotypic change is highest for organisms with simple phenotypes. Artificial evolving systems can help us study aspects of biological evolvability that are not accessible in vastly more complex natural systems. They can also help identify properties, such as robustness, that are required for both human-designed artificial systems and synthetic biological systems to be evolvable.

  20. The influence of temperature on photosynthesis of different tomato genotypes

    NARCIS (Netherlands)

    Gosiewski, W.; Nilwik, H.J.M.; Bierhuizen, J.F.

    1982-01-01

    Net photosynthesis and dark respiration from whole plants of various tomato genotypes were measured in a closed system. At low irradiance (27 W m−2) and low external CO2 concentration (550 mg m−3), net photosynthesis of 10 genotypes was found to vary between 0.122 and 0.209 mg CO2 m−2 s−1.

  1. characterisation of common bean genotypes based on storage

    African Journals Online (AJOL)

    ACSS

    of pliers and then ground to fine powder with a ... segregation of genotypes were Rm 23.75, 32.50,. 33.75, 22.50 ... Figure 1. Positions of Phaseolus vulgarisL. genotypes on the first and second correspondence scores based on storage protein.

  2. Integrated analysis for genotypic adaptation in rice | Das | African ...

    African Journals Online (AJOL)

    Integrated analysis for genotypic adaptation in rice. S Das, RC Misra, MC Pattnaik, SK Sinha. Abstract. Development of varieties with high yield potential coupled with wide adaptability is an important plant breeding objective. The presence of genotype by environment (GxE) interaction plays a crucial role in determining the ...

  3. 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

  4. Beijing/W genotype Mycobacterium tuberculosis and drug resistance.

    NARCIS (Netherlands)

    Glynn, Judith R; Kremer, Kristin; Borgdorff, Martien W; Rodriguez, Mar Pujades; Soolingen, Dick van

    2006-01-01

    Beijing/W genotype Mycobacterium tuberculosis is widespread, may be increasing, and may have a predilection for drug resistance. Individual-level data on >29,000 patients from 49 studies in 35 countries were combined to assess the Beijing genotype's prevalence worldwide, trends over time and with

  5. Apolipoprotein E genotype, cardiovascular biomarkers and risk of stroke

    DEFF Research Database (Denmark)

    Khan, Tauseef A; Shah, Tina; Prieto, David

    2013-01-01

    At the APOE gene, encoding apolipoprotein E, genotypes of the ε2/ε3/ε4 alleles associated with higher LDL-cholesterol (LDL-C) levels are also associated with higher coronary risk. However, the association of APOE genotype with other cardiovascular biomarkers and risk of ischaemic stroke is less c...

  6. Genotype dependent callus induction and shoot regeneration in ...

    African Journals Online (AJOL)

    This study aims to observe the effect of genotype, hormone and culture conditions on sunflower (Helianthus annuus L.) callus induction and indirect plant regeneration. Calli were obtained from hypocotyl and cotyledon explants of five different sunflower genotypes; Trakya 80, Trakya 129, Trakya 259, Trakya 2098 and ...

  7. Efficiency and response of conilon coffee genotypes to nitrogen supply

    African Journals Online (AJOL)

    The objective of the study was to differentiate genotypes with higher efficiency and responsiveness to nitrogen supply, to understand how the nitrogen supply can impact the dry matter allocation and the accumulation of this nutrient in the different plant compartments of genotypes of conilon coffee, cultivated under ...

  8. Haemoglobin genotype of children with severe malaria seen at the ...

    African Journals Online (AJOL)

    Abstract: Introduction: Types of haemoglobin (Hb) genotype have been found to be crucial to the rate of red blood cell parasite invasion, multiplication, and destruction as well as outcome of malaria disease. In a bid to provide more information on the relationship between Hb genotype and level of protection conferred by ...

  9. participatory selection of mungbean genotypes in uganda abstract

    African Journals Online (AJOL)

    ACSS

    2017-05-29

    May 29, 2017 ... that there were no significant differences in genotype choices based on gender ... However, there were significant genotype preference differences between eastern and northern regions (χ2 = ..... for southern Ethiopia. ... of improved cowpea cultivars in the Guinea and Sudan savanna zones of north east.

  10. Genetic variation of response to water deficit in parental genotypes ...

    African Journals Online (AJOL)

    dgomi

    In this study, we investigated morphological and photosynthetic responses to water deficit in parental genotypes of M. ... for adaptation to water deficit in legumes is a prerequisite for any research aiming to improve legume yields. ...... tolerant genotypes in rainfed lowland rice. Field Crop. Res. 99:48-58. Rouached A, Slama I, ...

  11. Clusters of incompatible genotypes evolve with limited dispersal

    Science.gov (United States)

    Erin L. Landguth; Norman A. Johnson; Samuel A. Cushman

    2015-01-01

    Theoretical and empirical studies have shown heterogeneous selection to be the primary driver for the evolution of reproductively isolated genotypes in the absence of geographic barriers. Here, we ask whether limited dispersal alone can lead to the evolution of reproductively isolated genotypes despite the absence of any geographic barriers or heterogeneous...

  12. Evaluation of sorghum genotypes under drought stress conditions ...

    African Journals Online (AJOL)

    Seven genotypes of sorghum (Sorghum bicolour (L.) Moench) were studied in both drought and normal conditions. In each condition, the genotypes were evaluated using a split plot based randomized complete block design with three replications. Drought tolerance indices including stability tolerance index (STI), mean ...

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

    Directory of Open Access Journals (Sweden)

    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.

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

    Science.gov (United States)

    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...

  15. Molecular genotyping of Echinococcus granulosus from dromedaries (Camelus dromedarius) in eastern Iran.

    Science.gov (United States)

    Moghaddas, E; Borji, H; Naghibi, A; Shayan, P; Razmi, G R

    2015-01-01

    With the aim of genotyping Echinococcus granulosus cysts found in Iranian dromedaries (Camelus dromedarius), 50 cysts of E. granulosus were collected from five geographical regions in Iran. Cysts were characterized using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis of the internal transcribed spacer 1 (ITS1) gene and sequencing fragments of the genes coding for mitochondrial cytochrome c oxidase subunit 1 (cox1). Morphological criteria using rostellar hook dimensions were also undertaken. The present results have shown that 27 out of 50 E. granulosus cysts (54%) were determined as the G1 strain, and the other (46%) were determined as the G6 strain. The molecular analysis of the ITS1 region of ribosomal DNA corresponded with the morphological findings. Because of its recognized infectivity in humans, the G1 genotype is a direct threat to human health and its presence in Iranian dromedaries is of urgent public health importance.

  16. SNPMClust: Bivariate Gaussian Genotype Clustering and Calling for Illumina Microarrays

    Directory of Open Access Journals (Sweden)

    Stephen W. Erickson

    2016-07-01

    Full Text Available SNPMClust is an R package for genotype clustering and calling with Illumina microarrays. It was originally developed for studies using the GoldenGate custom genotyping platform but can be used with other Illumina platforms, including Infinium BeadChip. The algorithm first rescales the fluorescent signal intensity data, adds empirically derived pseudo-data to minor allele genotype clusters, then uses the package mclust for bivariate Gaussian model fitting. We compared the accuracy and sensitivity of SNPMClust to that of GenCall, Illumina's proprietary algorithm, on a data set of 94 whole-genome amplified buccal (cheek swab DNA samples. These samples were genotyped on a custom panel which included 1064 SNPs for which the true genotype was known with high confidence. SNPMClust produced uniformly lower false call rates over a wide range of overall call rates.

  17. Micropropagation of six Paulownia genotypes through tissue culture

    Directory of Open Access Journals (Sweden)

    Lydia Shtereva

    2014-12-01

    Full Text Available We investigated the effect of genotype and culture medium on the in vitro germination and development of plantlets from seeds of 6 different Paulownia genotypes (P. tomentosa, hybrid lines P. tomentosa P. fortunei (Mega, Ganter and Caroline, P. elongata and hybrid line P. elongata P. fortunei. Nodal and shoot tip explants were used for micropropagation of Paulownia genotypes by manipulating plant growth regulators. The highest germination percentage for all genotypes was obtained for seeds inoculated on medium supplemented with 50 mg*L GA3 (MSG2. On Thidiazuron containing media, the explants of hybrid line P. elongata P. fortunei exhibited the highest frequency of axillary shoot proliferation following by P. tomentosa P. fortunei. The results are discussed with the perspective of applying an improved protocol for in vitro seed germination and plantlet formation in several economically valuable Paulownia genotypes.

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