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1.
Nat Commun ; 11(1): 4918, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-33004800

RESUMO

In order to control and eradicate epidemic cholera, we need to understand how epidemics begin, how they spread, and how they decline and eventually end. This requires extensive sampling of epidemic disease over time, alongside the background of endemic disease that may exist concurrently with the epidemic. The unique circumstances surrounding the Argentinian cholera epidemic of 1992-1998 presented an opportunity to do this. Here, we use 490 Argentinian V. cholerae genome sequences to characterise the variation within, and between, epidemic and endemic V. cholerae. We show that, during the 1992-1998 cholera epidemic, the invariant epidemic clone co-existed alongside highly diverse members of the Vibrio cholerae species in Argentina, and we contrast the clonality of epidemic V. cholerae with the background diversity of local endemic bacteria. Our findings refine and add nuance to our genomic definitions of epidemic and endemic cholera, and are of direct relevance to controlling current and future cholera epidemics.


Assuntos
Cólera/microbiologia , Doenças Endêmicas/prevenção & controle , Genoma Bacteriano/genética , Pandemias/prevenção & controle , Vibrio cholerae/genética , Argentina/epidemiologia , Cólera/epidemiologia , Cólera/prevenção & controle , DNA Bacteriano/genética , DNA Bacteriano/isolamento & purificação , História do Século XIX , História do Século XX , Humanos , Anotação de Sequência Molecular , Pandemias/história , Filogenia , Polimorfismo de Nucleotídeo Único , Análise de Sequência de DNA , Vibrio cholerae/isolamento & purificação , Vibrio cholerae/patogenicidade
2.
Nat Commun ; 11(1): 4379, 2020 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-32873785

RESUMO

The gut microbiome harbors a 'silent reservoir' of antibiotic resistance (AR) genes that is thought to contribute to the emergence of multidrug-resistant pathogens through horizontal gene transfer (HGT). To counteract the spread of AR, it is paramount to know which organisms harbor mobile AR genes and which organisms engage in HGT. Despite methods that characterize the overall abundance of AR genes in the gut, technological limitations of short-read sequencing have precluded linking bacterial taxa to specific mobile genetic elements (MGEs) encoding AR genes. Here, we apply Hi-C, a high-throughput, culture-independent method, to surveil the bacterial carriage of MGEs. We compare two healthy individuals with seven neutropenic patients undergoing hematopoietic stem cell transplantation, who receive multiple courses of antibiotics, and are acutely vulnerable to the threat of multidrug-resistant infections. We find distinct networks of HGT across individuals, though AR and mobile genes are associated with more diverse taxa within the neutropenic patients than the healthy subjects. Our data further suggest that HGT occurs frequently over a several-week period in both cohorts. Whereas most efforts to understand the spread of AR genes have focused on pathogenic species, our findings shed light on the role of the human gut microbiome in this process.


Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana/genética , Microbioma Gastrointestinal/genética , Transferência Genética Horizontal , Genes Bacterianos/efeitos dos fármacos , Adulto , Idoso , Antibacterianos/uso terapêutico , DNA Bacteriano/genética , DNA Bacteriano/isolamento & purificação , Microbioma Gastrointestinal/efeitos dos fármacos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Sequências Repetitivas Dispersas/efeitos dos fármacos , Pessoa de Meia-Idade
3.
Nat Commun ; 11(1): 4870, 2020 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-32978384

RESUMO

Little is known about the physiology of latent Mycobacterium tuberculosis infection. We studied the mutational rates of 24 index tuberculosis (TB) cases and their latently infected household contacts who developed active TB up to 5.25 years later, as an indication of bacterial physiological state and possible generation times during latent TB infection in humans. Here we report that the rate of new mutations in the M. tuberculosis genome decline dramatically after two years of latent infection (two-sided p < 0.001, assuming an 18 h generation time equal to log phase M. tuberculosis, with latency period modeled as a continuous variable). Alternatively, assuming a fixed mutation rate, the generation time increases over the latency duration. Mutations indicative of oxidative stress do not increase with increasing latency duration suggesting a lack of host or bacterial derived mutational stress. These results suggest that M. tuberculosis enters a quiescent state during latency, decreasing the risk for mutational drug resistance and increasing generation time, but potentially increasing bacterial tolerance to drugs that target actively growing bacteria.


Assuntos
Tuberculose Latente/microbiologia , Taxa de Mutação , Mycobacterium tuberculosis/genética , Tuberculose/microbiologia , Adulto , Brasil , DNA Bacteriano/isolamento & purificação , Feminino , Genoma Bacteriano , Humanos , Masculino , Mutação , Mycobacterium tuberculosis/patogenicidade , Estresse Oxidativo , Filogenia , Polimorfismo de Nucleotídeo Único , Fatores de Tempo , Adulto Jovem
4.
PLoS One ; 15(8): e0237748, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32866195

RESUMO

Soil microbiota are considered a source of undiscovered bioactive compounds, yet cultivation of most bacteria within a sample remains generally unsuccessful. Two main reasons behind the unculturability of bacteria are the presence of cells in a viable but not culturable state (such as dormant cells) and the failure to provide the necessary growth requirements in vitro (leading to the classification of some bacterial taxa as yet-to-be-cultured). The present work focuses on the development of a single procedure that helps distinguish between both phenomena of unculturability based on viability staining coupled with flow cytometry and fluorescence-activated cell sorting. In the selected soil sample, the success rate of cultured bacteria was doubled by selecting viable and metabolically active bacteria. It was determined that most of the uncultured fraction was not dormant or dead but likely required different growth conditions. It was also determined that the staining process introduced changes in the taxonomic composition of the outgrown bacterial biomass, which should be considered for further developments. This research shows the potential of flow cytometry and fluorescence-activated cell sorting applied to soil samples to improve the success rate of bacterial cultivation by estimating the proportion of dormant and yet-to-be-cultured bacteria and by directly excluding dormant cells from being inoculated into growth media.


Assuntos
Bactérias/crescimento & desenvolvimento , Técnicas Microbiológicas/métodos , Microbiota/fisiologia , Microbiologia do Solo , Bactérias/química , Bactérias/genética , Biomassa , Separação Celular/métodos , Meios de Cultura , DNA Bacteriano/isolamento & purificação , Estudos de Viabilidade , Citometria de Fluxo/métodos , Corantes Fluorescentes/química , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Coloração e Rotulagem/métodos
5.
Mem Inst Oswaldo Cruz ; 115: e200184, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32785422

RESUMO

BACKGROUND Carrion's disease (CD) is a neglected biphasic illness caused by Bartonella bacilliformis, a Gram-negative bacteria found in the Andean valleys. The spread of resistant strains underlines the need for novel antimicrobials against B. bacilliformis and related bacterial pathogens. OBJECTIVE The main aim of this study was to integrate genomic-scale data to shortlist a set of proteins that could serve as attractive targets for new antimicrobial discovery to combat B. bacilliformis. METHODS We performed a multidimensional genomic scale analysis of potential and relevant targets which includes structural druggability, metabolic analysis and essentiality criteria to select proteins with attractive features for drug discovery. FINDINGS We shortlisted seventeen relevant proteins to develop new drugs against the causative agent of Carrion's disease. Particularly, the protein products of fabI, folA, aroA, trmFO, uppP and murE genes, meet an important number of desirable features that make them attractive targets for new drug development. This data compendium is freely available as a web server (http://target.sbg.qb.fcen.uba.ar/). MAIN CONCLUSION This work represents an effort to reduce the costs in the first phases of B. bacilliformis drug discovery.


Assuntos
Antibacterianos/uso terapêutico , Infecções por Bartonella/tratamento farmacológico , Bartonella bacilliformis/efeitos dos fármacos , Bartonella bacilliformis/genética , Bartonella bacilliformis/isolamento & purificação , DNA Bacteriano/genética , DNA Bacteriano/isolamento & purificação , Genômica , Humanos , Reação em Cadeia da Polimerase
6.
PLoS One ; 15(8): e0237655, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32810167

RESUMO

BACKGROUND: Several tests are available for plague confirmation but bacteriological culture with Yersinia pestis strain isolation remains the gold standard according to the World Health Organization. However, this is a time consuming procedure; requiring specific devices and well-qualified staff. In addition, strain isolation is challenging if antibiotics have been administered prior to sampling. Here, we developed a loop-mediated isothermal amplification (LAMP) technique, a rapid, simple, sensitive and specific technique that would be able to detect Y. pestis in human biological samples. METHODS: LAMP primers were designed to target the caf1 gene which is specific to Y. pestis. The detection limit was determined by testing 10-fold serial dilution of Y. pestis DNA. Cross-reactivity was tested using DNA extracts from 14 pathogens and 47 residual samples from patients suffering from non-plague diseases. Specificity and sensitivity of the LAMP caf1 were assessed on DNA extracts of 160 human biological samples. Then, the performance of the LAMP caf1 assay was compared to conventional PCR and bacteriological culture. RESULTS: The detection limit of the developed Y. pestis LAMP assay was 3.79 pg/µl, similar to conventional PCR. The result could be read out within 45 min and as early as 35 minutes in presence of loop primer, using a simple water bath at 63°C. This is superior to culture with respect to time (requires up to 10 days) and simplicity of equipment compared to PCR. Furthermore, no cross-reactivity was found when tested on DNA extracts from other pathogens and human biological samples from patients with non-plague diseases. Compared to the gold standard, LAMP sensitivity and specificity were 97.9% (95% CI: 89.1%-99.9%) and 94.6% (95% CI: 88.6%-97.9%), respectively. CONCLUSION: LAMP detected Y. pestis effectively with high sensitivity and specificity in human plague biological samples. It can potentially be used in the field during outbreaks in resource limited countries such as Madagascar.


Assuntos
Técnicas Bacteriológicas/métodos , DNA Bacteriano/isolamento & purificação , Técnicas de Amplificação de Ácido Nucleico/métodos , Peste/diagnóstico , Yersinia pestis/isolamento & purificação , Técnicas Bacteriológicas/economia , Estudos de Viabilidade , Humanos , Limite de Detecção , Madagáscar , Técnicas de Amplificação de Ácido Nucleico/economia , Peste/microbiologia , Fatores de Tempo , Yersinia pestis/genética
7.
PLoS One ; 15(8): e0237704, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32804963

RESUMO

Since plastics degrade very slowly, they remain in the environment on much longer timescales than most natural organic substrates and provide a novel habitat for colonization by bacterial communities. The spectrum of relationships between plastics and bacteria, however, is little understood. The first objective of this study was to examine plastics as substrates for communities of Bacteria in estuarine surface waters. We used next-generation sequencing of the 16S rRNA gene to characterize communities from plastics collected in the field, and over the course of two colonization experiments, from biofilms that developed on plastic (low-density polyethylene, high-density polyethylene, polypropylene, polycarbonate, polystyrene) and glass substrates placed in the environment. Both field sampling and colonization experiments were conducted in estuarine tributaries of the lower Chesapeake Bay. As a second objective, we concomitantly analyzed biofilms on plastic substrates to ascertain the presence and abundance of Vibrio spp. bacteria, then isolated three human pathogens, V. cholerae, V. parahaemolyticus, and V. vulnificus, and determined their antibiotic-resistant profiles. In both components of this study, we compared our results with analyses conducted on paired samples of estuarine water. This research adds to a nascent literature that suggests environmental factors govern the development of bacterial communities on plastics, more so than the characteristics of the plastic substrates themselves. In addition, this study is the first to culture three pathogenic vibrios from plastics in estuaries, reinforcing and expanding upon earlier reports of plastic pollution as a habitat for Vibrio species. The antibiotic resistance detected among the isolates, coupled with the longevity of plastics in the aqueous environment, suggests biofilms on plastics have potential to persist and serve as focal points of potential pathogens and horizontal gene transfer.


Assuntos
Biofilmes/efeitos dos fármacos , Estuários , Plásticos , Vibrio/isolamento & purificação , Poluentes da Água , Antibacterianos/farmacologia , Oceano Atlântico , DNA Bacteriano/isolamento & purificação , Farmacorresistência Bacteriana/genética , Transferência Genética Horizontal , RNA Ribossômico 16S/genética , Água do Mar/microbiologia , Vibrio/efeitos dos fármacos , Vibrio/genética
8.
PLoS One ; 15(8): e0237775, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32813739

RESUMO

Nile tilapia, Oreochromis niloticus is the third most commonly farmed finfish species in the world, accounting for nearly 5% of global aquaculture production. In the past few decades much of the success of this species has been attributed to the development and distribution of Genetically Improved Farmed Tilapia (GIFT). Despite the increasing availability of GIFT, the productivity of small-scale farming remains highly variable, particularly in developing nations. Commercial fish-feed pellets can increase fish farm productivity; however, many small-scale farmers rely on other means of feeding fish due to the high cost and limited availability of commercial fish feed pellets. Therefore, understanding how locally-sourced feeds affect the production of GIFT is an important step towards improving feeding practices, particularly for farmers with low financial capital. This study used stable isotope analysis (SIA) and 16S rRNA gene sequencing to compare the effects of a locally-sourced vegetable-based diet and commercial pellet-based diets on the relative condition, nutrient assimilation patterns and gastrointestinal microbiota of GIFT. GIFT fed a locally-sourced diet were smaller, and in a significantly poorer condition than those fed with commercial fish feeds. SIA showed no differences in dietary carbon between the two diets; however, δ13C, poor fish condition and the abundance of specific bacterial taxa (of such as Fusobacteria) were correlated. SIA revealed that GIFT fed locally-sourced diets that predominantly consisted of vegetables were significantly enriched in δ15N despite a perceived lack of dietary protein. This enrichment suggests that GIFT fed a locally-sourced diet may be supplementing their diet via cannibalism, a behaviour representative of poor farming practice. Overall this study highlights the need to increase the availability of suitable GIFT feeds in developing nations. The development a low-cost feed alternative could improve the success of small-scale GIFT farmers in PNG, increasing both food and income security within the region.


Assuntos
Ração Animal , Animais Geneticamente Modificados/metabolismo , Aquicultura/métodos , Ciclídeos/metabolismo , Microbioma Gastrointestinal/fisiologia , Animais , Animais Geneticamente Modificados/genética , Animais Geneticamente Modificados/microbiologia , Aquicultura/economia , Aquicultura/organização & administração , Canibalismo , Ciclídeos/genética , Ciclídeos/microbiologia , DNA Bacteriano/isolamento & purificação , Suplementos Nutricionais/economia , Eficiência Organizacional/economia , Fazendas/economia , Fazendas/organização & administração , New South Wales , Nutrientes/metabolismo , RNA Ribossômico 16S/genética
9.
PLoS One ; 15(8): e0237779, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32834004

RESUMO

Microbiome data consists of operational taxonomic unit (OTU) counts characterized by zero-inflation, over-dispersion, and grouping structure among samples. Currently, statistical testing methods are commonly performed to identify OTUs that are associated with a phenotype. The limitations of statistical testing methods include that the validity of p-values/q-values depend sensitively on the correctness of models and that the statistical significance does not necessarily imply predictivity. Predictive analysis using methods such as LASSO is an alternative approach for identifying associated OTUs and for measuring the predictability of the phenotype variable with OTUs and other covariate variables. We investigate three strategies of performing predictive analysis: (1) LASSO: fitting a LASSO multinomial logistic regression model to all OTU counts with specific transformation; (2) screening+GLM: screening OTUs with q-values returned by fitting a GLMM to each OTU, then fitting a GLM model using a subset of selected OTUs; (3) screening+LASSO: fitting a LASSO to a subset of OTUs selected with GLMM. We have conducted empirical studies using three simulation datasets generated using Dirichlet-multinomial models and a real gut microbiome data related to Parkinson's disease to investigate the performance of the three strategies for predictive analysis. Our simulation studies show that the predictive performance of LASSO with appropriate variable transformation works remarkably well on zero-inflated data. Our results of real data analysis show that Parkinson's disease can be predicted based on selected OTUs after the binary transformation, age, and sex with high accuracy (Error Rate = 0.199, AUC = 0.872, AUPRC = 0.912). These results provide strong evidences of the relationship between Parkinson's disease and the gut microbiome.


Assuntos
Bactérias/classificação , Interpretação Estatística de Dados , Microbioma Gastrointestinal/genética , Modelos Biológicos , Doença de Parkinson/diagnóstico , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Bactérias/genética , Bactérias/isolamento & purificação , Estudos de Coortes , Simulação por Computador , DNA Bacteriano/isolamento & purificação , Conjuntos de Dados como Assunto , Feminino , Humanos , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Doença de Parkinson/microbiologia , Valor Preditivo dos Testes , Prognóstico , RNA Ribossômico 16S/genética , Fatores Sexuais
10.
PLoS One ; 15(8): e0237730, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32857799

RESUMO

Freshwater ecosystems play a key role in shaping the global carbon cycle and maintaining the ecological balance that sustains biodiversity worldwide. Surficial water bodies are often interconnected with groundwater, forming a physical continuum, and their interaction has been reported as a crucial driver for organic matter (OM) inputs in groundwater systems. However, despite the growing concerns related to increasing anthropogenic pressure and effects of global change to groundwater environments, our understanding of the dynamics regulating subterranean carbon flows is still sparse. We traced carbon composition and transformations in an arid zone calcrete aquifer using a novel multidisciplinary approach that combined isotopic analyses of dissolved organic carbon (DOC) and inorganic carbon (DIC) (δ13CDOC, δ13CDIC, 14CDOC and 14CDIC) with fluorescence spectroscopy (Chromophoric Dissolved OM (CDOM) characterisation) and metabarcoding analyses (taxonomic and functional genomics on bacterial 16S rRNA). To compare dynamics linked to potential aquifer recharge processes, water samples were collected from two boreholes under contrasting rainfall: low rainfall ((LR), dry season) and high rainfall ((HR), wet season). Our isotopic results indicate limited changes and dominance of modern terrestrial carbon in the upper part (northeast) of the bore field, but correlation between HR and increased old and 13C-enriched DOC in the lower area (southwest). CDOM results show a shift from terrestrially to microbially derived compounds after rainfall in the same lower field bore, which was also sampled for microbial genetics. Functional genomic results showed increased genes coding for degradative pathways-dominated by those related to aromatic compound metabolisms-during HR. Our results indicate that rainfall leads to different responses in different parts of the bore field, with an increase in old carbon sources and microbial processing in the lower part of the field. We hypothesise that this may be due to increasing salinity, either due to mobilisation of Cl- from the soil, or infiltration from the downstream salt lake during HR. This study is the first to use a multi-technique assessment using stable and radioactive isotopes together with functional genomics to probe the principal organic biogeochemical pathways regulating an arid zone calcrete system. Further investigations involving extensive sampling from diverse groundwater ecosystems will allow better understanding of the microbiological pathways sustaining the ecological functioning of subterranean biota.


Assuntos
Ciclo do Carbono , Isótopos de Carbono/análise , Monitoramento Ambiental/métodos , Água Subterrânea/química , Microbiota/fisiologia , Solo/química , Austrália , Isótopos de Carbono/metabolismo , Código de Barras de DNA Taxonômico , DNA Bacteriano/genética , DNA Bacteriano/isolamento & purificação , Água Subterrânea/microbiologia , RNA Ribossômico 16S/genética , Chuva , Salinidade , Microbiologia do Solo , Espectrometria de Fluorescência
11.
PLoS One ; 15(8): e0236054, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32750053

RESUMO

INTRODUCTION: Multi-drug resistance is a major challenge in the control of tuberculosis. Despite newer modalities for diagnosis and treatment, people are still suffering from this disease. Understanding the common gene mutations conferring rifampicin and isoniazid resistance is crucial for the implementation of effective molecular tools at local and national levels. Hence, this study aimed to evaluate the molecular detection of rifampicin and isoniazid-resistant gene mutations in M.tuberculosis isolates in Addis Ababa, Ethiopia. METHOD: Health Center-based cross-sectional study was conducted between January and September 2017 in Addis Ababa, Ethiopia. The collected sputum samples were processed for mycobacterial isolation and Region of difference 9 based polymerase chain reaction for species identification. To characterize the rifampicin and isoniazid-resistant M. tuberculosis isolates, a molecular genetic assay (GenoType MTBDRplus) was used; the assay is based on DNA-STRIP technology. RESULT: Culture positivity was confirmed in 82.6% (190/230) of smear-positive newly diagnosed pulmonary tuberculosis cases enrolled in the study. From 190 isolates 93.2% were sensitive for both rifampicin and isoniazid, and 6.8% of the isolates were resistant to at least one of the tested anti-TB drugs. Gene mutations were observed in all studied multidrug resistance-associated gene loci (rpoB, katG, and inhA). Two isolates exhibited heteroresistance, a mutated, as well as wild type sequences, were detected in the respective strains. MDR-TB case was observed in 1.1% (2/190) of the cases. All the MDR-TB cases were positive for HIV and found to have a history of prior hospital admission. CONCLUSION: In our finding a relatively high prevalence of any drug resistance was observed and the overall prevalence of multidrug-resistant tuberculosis was 1.1%.The majority of drug-resistant isolates demonstrated common mutations. Heteroresistant strains were detected, signaling the existence of an M.tuberculosis population with variable responses to anti-tuberculosis drugs or of mixed infections.


Assuntos
Antituberculosos/farmacologia , Farmacorresistência Bacteriana Múltipla/genética , Mycobacterium tuberculosis/genética , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Adolescente , Adulto , Antituberculosos/uso terapêutico , Estudos Transversais , Análise Mutacional de DNA , DNA Bacteriano/genética , DNA Bacteriano/isolamento & purificação , Etiópia , Feminino , Genes Bacterianos/genética , Loci Gênicos/genética , Humanos , Isoniazida/farmacologia , Isoniazida/uso terapêutico , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Mutação , Mycobacterium tuberculosis/isolamento & purificação , Reação em Cadeia da Polimerase , Rifampina/farmacologia , Rifampina/uso terapêutico , Escarro/microbiologia , Tuberculose Resistente a Múltiplos Medicamentos/microbiologia , Adulto Jovem
12.
PLoS One ; 15(8): e0237345, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32813724

RESUMO

BACKGROUND: Mixed/polyclonal infections due to different genotypes are reported in Tuberculosis. The current study was designed to understand the fate of mixed infections during the course of treatment and follow-up and its role in disease pathogenesis. METHODS: Sputum samples were collected on 0,1,2,3,6,12 and 24 months from 157 treatment-naïve patients, cultures subjected to Drug-Susceptibility-testing (MGIT 960), spoligotyping, MIRU-VNTR and SNP genotyping. All isolated colonies on thin layer agar (7H11) were subjected to spoligotyping. FINDINGS: One thirty three baseline cultures were positive (133/157, 84.7%), 43(32.3%) had mixture of genotypes. Twenty-four of these patients (55.8%) showed change in genotype while six showed different drug-susceptibility patterns while on treatment. Twenty-three (53.5%) patients with polyclonal infections showed resistance to at least one drug compared to 10/90 (11.1%) monoclonal infections (P<0.0001). Eight patients had recurrent TB, two with a new genotype and two with altered phenotypic DST. CONCLUSIONS: The coexistence of different genotypes and change of genotypes during the same disease episode, while on treatment, confirms constancy of polyclonal infections. The composition of the mixture of genotypes and the relative predominance may be missed by culture due to its limit of detection. Polyclonal infections in TB could be a rule rather than exception and challenges the age-old dogma of reactivation/reinfection.


Assuntos
Antituberculosos/farmacologia , Coinfecção/tratamento farmacológico , Farmacorresistência Bacteriana Múltipla/genética , Mycobacterium tuberculosis/genética , Tuberculose Pulmonar/microbiologia , Adolescente , Adulto , Antituberculosos/uso terapêutico , Técnicas de Tipagem Bacteriana , Evolução Clonal , Coinfecção/epidemiologia , Coinfecção/microbiologia , DNA Bacteriano/genética , DNA Bacteriano/isolamento & purificação , Feminino , Seguimentos , Técnicas de Genotipagem , Humanos , Limite de Detecção , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Técnicas de Diagnóstico Molecular , Mycobacterium tuberculosis/isolamento & purificação , Mycobacterium tuberculosis/patogenicidade , Filogenia , Polimorfismo de Nucleotídeo Único , Prevalência , Recidiva , Tuberculose Pulmonar/tratamento farmacológico , Tuberculose Pulmonar/epidemiologia , Adulto Jovem
13.
J Med Microbiol ; 69(8): 1079-1088, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32729813

RESUMO

Introduction. Linezolid-resistant (LZR) Staphylococcus capitis has recently emerged in our hospital, and its potential resistance mechanisms are still not clear.Aim. This study aimed to investigate the epidemiology, clinical and genetic characteristics, resistance mechanisms and biofilm formation capacity of LZR S. capitis isolated from patients at Huashan Hospital, Shanghai, PR China between 2012 and 2018.Methodology. Strains were subjected to antimicrobial susceptibility testing (AST) with antibiotics using the broth microdilution method according to the Clinical and Laboratory Standards Institute (CLSI) guidelines. The presence of cfr, optrA and poxtA, as well as mutations in the 23S ribosomal (r)RNA and ribosomal proteins, was investigated using PCR and sequencing techniques. The genetic relationship between isolates was analysed using pulsed-field gel electrophoresis (PFGE) and whole-genome sequencing (WGS). Biofilm biomasses were detected by using crystal violet staining.Results. Twenty-one LZR S. capitis strains displayed MICs of 32-512 µg ml-1. All LZR strains showed G2576T and C2104T mutations in the 23S rRNA V region. Besides G2576T and C2104T, no base mutations were detected in the V region. The cfr was detected in 12 strains, while optrA and poxtA were not amplified in 21 S. capitis strains. PFGE showed that the LZR S. capitis strains belonged to a single clone. The phylogenetic tree showed that 20 LZR S. capitis strains were highly similar to LNZR-1, isolated from Harbin (located in the north of China) in 2013, which showed resistance to linezolid.Conclusions. In this research, cfr-negative strains displayed linezolid MICs of 32 µg ml-1. In comparison, cfr-positive strains exhibited linezolid MICs of 128-512 µg ml-1, indicating that high levels of linezolid resistance appear to be related to the presence of cfr. The outbreak of LZR S. capitis in our hospital needs to be monitored closely.


Assuntos
Antibacterianos/farmacologia , Linezolida/farmacologia , Infecções Estafilocócicas/epidemiologia , Staphylococcus capitis/efeitos dos fármacos , Adulto , Idoso , Idoso de 80 Anos ou mais , Biofilmes/crescimento & desenvolvimento , China/epidemiologia , DNA Bacteriano/isolamento & purificação , Farmacorresistência Bacteriana/genética , Eletroforese em Gel de Campo Pulsado , Feminino , Humanos , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Mutação , Oxazolidinonas/farmacologia , RNA Ribossômico 23S/genética , Infecções Estafilocócicas/microbiologia , Staphylococcus capitis/genética , Staphylococcus capitis/fisiologia , Sequenciamento Completo do Genoma , Adulto Jovem
14.
PLoS Negl Trop Dis ; 14(7): e0008449, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32667914

RESUMO

BACKGROUND: The presence of Chlamydia trachomatis (Ct) DNA at non-ocular sites suggests that these sites may represent plausible routes of Ct transmission in trachoma. However, qPCR cannot discriminate between DNA from viable and non-viable bacteria. Here we use a propodium monoazide based viability PCR to investigate how long Ct remains viable at non-ocular sites under laboratory-controlled conditions. METHODS: Cultured Ct stocks (strain A2497) were diluted to final concentrations of 1000, 100, 10 and 1 omcB copies/µL and applied to plastic, woven mat, cotton cloth and pig skin. Swabs were then systemically collected from each surface and tested for the presence Ct DNA using qPCR. If Ct DNA was recovered, Ct viability was assessed over time by spiking multiple areas of the same surface type with the same final concentrations. Swabs were collected from each surface at 0, 2, 4, 6, 8 and 24 hours after spiking. Viability PCR was used to determine Ct viability at each timepoint. RESULTS: We were able to detect Ct DNA on all surfaces except the woven mat. Total Ct DNA remained detectable and stable over 24 hours for all concentrations applied to plastic, pig skin and cotton cloth. The amount of viable Ct decreased over time. For plastic and skin surfaces, only those where concentrations of 100 or 1000 omcB copies/µL were applied still had viable loads detectable after 24 hours. Cotton cloth showed a more rapid decrease and only those where concentrations of 1000 omcB copies/µL were applied still had viable DNA detectable after 24 hours. CONCLUSION: Plastic, cotton cloth and skin may contribute to transmission of the Ct strains that cause trachoma, by acting as sites where reservoirs of bacteria are deposited and later collected and transferred mechanically into previously uninfected eyes.


Assuntos
Chlamydia trachomatis/genética , DNA Bacteriano/isolamento & purificação , Fômites/microbiologia , Reação em Cadeia da Polimerase/métodos , Tracoma/microbiologia , Tracoma/transmissão , Humanos
15.
Nat Commun ; 11(1): 3772, 2020 07 29.
Artigo em Inglês | MEDLINE | ID: mdl-32728114

RESUMO

Selective and neutral forces shape human microbiota assembly in early life. The Tsimane are an indigenous Bolivian population with infant care-associated behaviors predicted to increase mother-infant microbial dispersal. Here, we characterize microbial community assembly in 47 infant-mother pairs from six Tsimane villages, using 16S rRNA gene amplicon sequencing of longitudinal stool and tongue swab samples. We find that infant consumption of dairy products, vegetables, and chicha (a fermented drink inoculated with oral microbes) is associated with stool microbiota composition. In stool and tongue samples, microbes shared between mothers and infants are more abundant than non-shared microbes. Using a neutral model of community assembly, we find that neutral processes alone explain the prevalence of 79% of infant-colonizing microbes, but explain microbial prevalence less well in adults from river villages with more regular access to markets. Our results underscore the importance of neutral forces during microbiota assembly. Changing lifestyle factors may alter traditional modes of microbiota assembly by decreasing the role of neutral processes.


Assuntos
Horticultura , Povos Indígenas , Microbiota , Adolescente , Adulto , Bolívia , Criança , Pré-Escolar , DNA Bacteriano/isolamento & purificação , Fezes/microbiologia , Feminino , Humanos , Lactente , Recém-Nascido , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Filogenia , RNA Ribossômico 16S/genética , Língua/microbiologia , Adulto Jovem
16.
PLoS One ; 15(7): e0236165, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32697804

RESUMO

In order to define the diversity and composition of the microbial communities colonizing of the soil microbiome of the Jinsha earthen relic, we used high-throughput sequencing technology to identify and characterize the microbiota in 22 samples collected from the Jinsha earthen relic in China during 2017 and 2018. We compared the taxonomy of the microbial communities from samples taken at different times and different sites. Our results showed that the identity of the dominant bacterial phyla differed among the samples. Proteobacteria (23-86.2%) were the predominant bacterial phylum in all samples taken from site A in both 2017 and 2018. However, Actinobacteria (21-92.3%) were the most popular bacterial phylum in samples from sites B and C in 2017 and 2018. Ascomycota were identified as the only fungal phyla in samples in 2017. However, the group varied drastically in relative abundance between 2017 and 2018. Functional analysis of the soil bacterial community suggested that abundant members of the microbiota may be associated with metabolism and the specific environment. This report was the first high-throughput sequencing study of the soil of the Jinsha earthen relic microbiome. Since soil microbiota can damage soil and archeological structures, comprehensive analyses of the microbiomes at archeological sites may contribute to the understand of the influence of microorganisms on the degradation of soil, as well as to the identification of potentially beneficial or undesirable members of these microbial communities in archeological sites. The study will be helpful to provide effective data and guidance for the prevention and control of microbial corrosion of the Jinsha earthen relic.


Assuntos
Arqueologia , Microbiota/genética , Microbiologia do Solo , Actinobacteria/genética , Actinobacteria/isolamento & purificação , Ascomicetos/genética , Ascomicetos/isolamento & purificação , China , DNA Bacteriano/isolamento & purificação , Sequenciamento de Nucleotídeos em Larga Escala , Proteobactérias/genética , Proteobactérias/isolamento & purificação , RNA Ribossômico 16S/genética
17.
Science ; 368(6495): 1135-1140, 2020 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-32499444

RESUMO

Determining where an object has been is a fundamental challenge for human health, commerce, and food safety. Location-specific microbes in principle offer a cheap and sensitive way to determine object provenance. We created a synthetic, scalable microbial spore system that identifies object provenance in under 1 hour at meter-scale resolution and near single-spore sensitivity and can be safely introduced into and recovered from the environment. This system solves the key challenges in object provenance: persistence in the environment, scalability, rapid and facile decoding, and biocontainment. Our system is compatible with SHERLOCK, a Cas13a RNA-guided nucleic acid detection assay, facilitating its implementation in a wide range of applications.


Assuntos
Código de Barras de DNA Taxonômico/métodos , DNA Bacteriano/isolamento & purificação , DNA Fúngico/isolamento & purificação , Microbiologia Ambiental , Microbiota/genética , Esporos/genética , Sistemas CRISPR-Cas , DNA Bacteriano/genética , DNA Fúngico/genética , RNA Guia
18.
J Med Microbiol ; 69(8): 1105-1113, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32597748

RESUMO

Introduction. Burkholderia cepacia complex (Bcc) bacteria, currently consisting of 23 closely related species, and Burkholderia gladioli, can cause serious and difficult-to-treat infections in people with cystic fibrosis. Identifying Burkholderia bacteria to the species level is considered important for understanding epidemiology and infection control, and predicting clinical outcomes. Matrix-assisted laser desorption/ionization time-of-flight MS (MALDI-TOF) is a rapid method recently introduced in clinical laboratories for bacterial species-level identification. However, reports on the ability of MALDI-TOF to accurately identify Bcc to the species level are mixed.Aim. The aim of this project was to evaluate the accuracy of MALDI-TOF using the Biotyper and VITEK MS systems in identifying isolates from 22 different Bcc species and B. gladioli compared to recA gene sequencing, which is considered the current gold standard for Bcc.Methodology. To capture maximum intra-species variation, phylogenetic trees were constructed from concatenated multi-locus sequence typing alleles and clustered with a novel k-medoids approach. One hundred isolates representing 22 Bcc species, plus B. gladioli, were assessed for bacterial identifications using the two MALDI-TOF systems.Results. At the genus level, 100 and 97.0 % of isolates were confidently identified as Burkholderia by the Biotyper and VITEK MS systems, respectively; moreover, 26.0 and 67.0 % of the isolates were correctly identified to the species level, respectively. In many, but not all, cases of species misidentification or failed identification, a representative library for that species was lacking.Conclusion. Currently available MALDI-TOF systems frequently do not accurately identify Bcc bacteria to the species level.


Assuntos
Burkholderia cepacia/isolamento & purificação , Burkholderia gladioli/isolamento & purificação , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Animais , Técnicas de Tipagem Bacteriana/métodos , Burkholderia cepacia/classificação , Burkholderia gladioli/classificação , Análise por Conglomerados , DNA Bacteriano/química , DNA Bacteriano/isolamento & purificação , Análise de Fourier , Humanos , Tipagem de Sequências Multilocus , Filogenia , Recombinases Rec A/genética , Alinhamento de Sequência
19.
PLoS One ; 15(6): e0233853, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32502212

RESUMO

Rapid detection of bacterial pathogens is a critical unmet need for both food and environmental samples such as irrigation water. As a part of the Food safety Modernization Act (FSMA), The Produce Safety rule has established several requirements for testing for the presence of generic Escherichia coli in water, but the current method available for testing (EPA M1603) demands specified multiple colony verification and highly trained personnel to perform these tests. The purpose of the study was to assess a phage induced bacterial lysis using quantitative image analysis to achieve rapid detection of E. coli at low concentrations within 8 hours. This study aimed to develop a simple yet highly sensitive and specific approach to detect target bacteria in complex matrices. In the study, E. coli cells were first enriched in tryptic soy broth (TSB), followed by T7 phage induced lysis, concentration, staining and fluorescent imaging. Image analysis was conducted including image pre-processing, image segmentation and quantitatively analysis of cellular morphological features (area, eccentricity and full width at half maximum). Challenge experiments using realistic matrices, including simulated fresh produce wash water, coconut water and spinach wash water, demonstrated the method can be applied for use in situations that occur in food processing facilities. The results indicated E. coli cells that are lysed by T7 phages demonstrated significantly (P < 0.05) higher extracellular DNA release, altered cellular shape (from rod to circular) and diffused fluorescent signal intensity. Using this biosensing strategy, a sensitivity to detect Escherichia coli at 10 CFU/ml within 8 hours was achieved, both in laboratory medium and in complex matrices. The proposed phage based biosensing strategy enables rapid detection of bacteria and is applicable to analysis of food systems. Furthermore, the steps involved in this assay can be automated to enable detection of target bacteria in food facilities without extensive resources.


Assuntos
Bacteriófago T7 , Técnicas Biossensoriais/métodos , DNA Ambiental/isolamento & purificação , Escherichia coli/isolamento & purificação , Processamento de Imagem Assistida por Computador , DNA Bacteriano/isolamento & purificação , Escherichia coli/genética , Escherichia coli/virologia , Microbiologia de Alimentos/normas , Inocuidade dos Alimentos , Microbiologia da Água/normas
20.
Int J Food Microbiol ; 331: 108716, 2020 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-32521375

RESUMO

Q fever is a common zoonotic disease with worldwide distribution. The causative agent of Q fever is Coxiella burnetii, a gram-negative and polymorphic rod bacterium. Sheep and goats are the primary reservoirs of this disease, although a variety of animal species can be infected. The main route of Q fever transmission from animals to humans is the inhalation of contaminated aerosols with C. burnetii. The bacterium is excreted in milk of infected animals and therefore; the consumption of unpasteurized milk and dairy products might be a route of coxiella burnetii transmission from animals to humans. The present study was conducted to determine the prevalence of C. burnetii in milk samples collected from sheep and goats in west Azerbaijan province, Iran. During 2018, a total number of 420 milk samples were collected from sheep (n = 210) and goats (n = 210) of different regions of the province. All milk samples were subjected to DNA extraction and examined by a highly and specific nested-PCR method. The results showed that 51 (12.1%) (95% CI: 9.3%-15.6%) examined samples [sheep; n = 16 (7.6%) and goat; n = 35 (16.6%)] were positive for C. burnetii. The prevalence of C. burnetii in goat milk samples was significantly higher than sheep milk samples (P < 0.05). The shedding of C. burnetii in milk was significantly higher in summer (25%) (P < 0.05, 95% CI: 17.7%-34%) than the other seasons. It was concluded that sheep and goat populations in west Azerbaijan play an important role in the epidemiology of Q fever.


Assuntos
Coxiella burnetii/genética , Doenças das Cabras/microbiologia , Leite/microbiologia , Febre Q/microbiologia , Doenças dos Ovinos/microbiologia , Zoonoses/microbiologia , Animais , Coxiella burnetii/isolamento & purificação , DNA Bacteriano/genética , DNA Bacteriano/isolamento & purificação , Doenças das Cabras/transmissão , Cabras , Humanos , Irã (Geográfico)/epidemiologia , Reação em Cadeia da Polimerase , Prevalência , Febre Q/epidemiologia , Ovinos , Doenças dos Ovinos/transmissão , Zoonoses/epidemiologia
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