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1.
Proc Natl Acad Sci U S A ; 116(48): 24084-24092, 2019 Nov 26.
Artigo em Inglês | MEDLINE | ID: mdl-31712444

RESUMO

An earlier report described a human case of necrotizing fasciitis (NF) caused by mixed infection with 4 Aeromonas hydrophila strains (NF1-NF4). While the NF2, NF3, and NF4 strains were clonal and possessed exotoxin A (ExoA), the NF1 strain was determined to be phylogenetically distinct, harboring a unique type 6 secretion system (T6SS) effector (TseC). During NF1 and NF2 mixed infection, only NF1 disseminated, while NF2 was rapidly killed by a contact-dependent mechanism and macrophage phagocytosis, as was demonstrated by using in vitro models. To confirm these findings, we developed 2 NF1 mutants (NF1ΔtseC and NF1ΔvasK); vasK encodes an essential T6SS structural component. NF1 VasK and TseC were proven to be involved in contact-dependent killing of NF2 in vitro, as well as in its elimination at the intramuscular injection site in vivo during mixed infection, with overall reduced mouse mortality. ExoA was shown to have an important role in NF by both NF1-exoA (with cis exoA) and NF2 during monomicrobial infection. However, the contribution of ExoA was more important for NF2 than NF1 in the murine peritonitis model. The NF2∆exoA mutant did not significantly alter animal mortality or NF1 dissemination during mixed infection in the NF model, suggesting that the ExoA activity was significant at the injection site. Immunization of mice to ExoA protected animals from NF2 monomicrobial challenge, but not from polymicrobial infection because of NF2 clearance. This study clarified the roles of T6SS and ExoA in pathogenesis caused by A. hydrophila NF strains in both mouse peritonitis and NF models in monomicrobial and polymicrobial infections.

2.
J Med Microbiol ; 68(10): 1507-1516, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31460858

RESUMO

Introduction. Prosthetic joint infections (PJIs) are challenging to treat therapeutically because the infectious agents often are resistant to antibiotics and capable of abundant growth in surface-attached biofilms. Though infection rates are low, ca. 1-2 %, the overall increase in the sheer number of joint replacement surgeries results in an increase in patients at risk.Aims. This study investigates the consensus of microbial species comprising PJI ecology, which is currently lacking.Methodology. In this study, PJI populations from seven patients were analysed using combined culturing and whole-genome shotgun sequencing (WGSS) to establish population profiles and compare WGSS and culture methods for detection and identification of the PJI microbiome.Results. WGSS detected strains when culture did not, notably dormant, culture-resistant and rare microbes. The CosmosID algorithm was used to predict micro-organisms present in the PJI and discriminate contaminants. However, culturing indicated the presence of microbes falling below the WGSS algorithm threshold. In these instances, microbes cultured are believed to be minor species. The two strategies were combined to build a population profile.Conclusions. Variability between and among PJIs showed that most infections were distinct and unique. Comparative analysis of populations revealed PJIs to form clusters that were related to, but separate from, vaginal, skin and gut microbiomes. Fungi and protists were detected by WGSS, but the role of fungi is just beginning to be understood and for protists it is unknown. These micro-organisms and their novel and strain-specific microbial interactions remain to be determined in current clinical tests.


Assuntos
Bactérias/genética , Bactérias/isolamento & purificação , Fungos/isolamento & purificação , Artropatias/microbiologia , Microbiota , Infecções Relacionadas à Prótese/microbiologia , Bactérias/classificação , Feminino , Fungos/classificação , Fungos/genética , Humanos , Articulações/microbiologia , Articulações/cirurgia , Masculino , Próteses e Implantes/efeitos adversos , Próteses e Implantes/microbiologia , Infecções Relacionadas à Prótese/etiologia , Estudos Retrospectivos , Sequenciamento Completo do Genoma
5.
Science ; 364(6435): 5, 2019 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-30948528
6.
Trends Microbiol ; 27(8): 670-677, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31031092

RESUMO

Now is an opportune time to foster collaborations across sectors and geographical boundaries to enable development of best practices for drinking water (DW) microbiome research, focusing on accuracy and reproducibility of meta-omic techniques (while learning from past microbiome projects). A large-scale coordinated effort that builds on this foundation will enable the urgently needed comprehensive spatiotemporal understanding and control of DW microbiomes by engineering interventions to protect public health. This opinion paper highlights the need to initiate and conduct a large-scale coordinated DW microbiome project by addressing key knowledge gaps and recommends a roadmap for this effort.

7.
Artigo em Inglês | MEDLINE | ID: mdl-30934749

RESUMO

The Little Bighorn River is the primary source of water for water treatment plants serving the local Crow Agency population, and has special significance in the spiritual and ceremonial life of the Crow tribe. Unfortunately, the watershed suffers from impaired water quality, with high counts of fecal coliform bacteria routinely measured during run-off events. A metagenomic analysis was carried out to identify potential pathogens in the river water. The Oxford Nanopore MinION platform was used to sequence DNA in near real time to identify both uncultured and a coliform-enriched culture of microbes collected from a popular summer swimming area of the Little Bighorn River. Sequences were analyzed using CosmosID bioinformatics and, in agreement with previous studies, enterohemorrhagic and enteropathogenic Escherichia coli and other E. coli pathotypes were identified. Noteworthy was detection and identification of enteroaggregative E. coli O104:H4 and Vibrio cholerae serotype O1 El Tor, however, cholera toxin genes were not identified. Other pathogenic microbes, as well as virulence genes and antimicrobial resistance markers, were also identified and characterized by metagenomic analyses. It is concluded that metagenomics provides a useful and potentially routine tool for identifying in an in-depth manner microbial contamination of waterways and, thereby, protecting public health.


Assuntos
Enterobacteriaceae/genética , Metagenômica/métodos , Rios/microbiologia , Escherichia coli/genética , Humanos , Montana/epidemiologia , Sorogrupo , Virulência , Qualidade da Água
9.
Mar Pollut Bull ; 136: 79-83, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30509844

RESUMO

Ballast water is used to safely stabilize and operate shipping vessels worldwide, in a multitude of aquatic settings, including inland, coastal and open oceans. However, ballast water may pose ecological, public health, and/or economic problems as it may serve as vehicles of transmission of microorganisms. Current ballast water regulations include limits of Escherichia coli, Enterococcus spp. and toxigenic Vibrio cholerae. Several United States Environmental Protection Agency approved standard operating protocols (SOPs) exist for detection of E. coli and Enterococci, yet none exists for V. cholerae. Current V. cholerae detection methods include colony blot hybridization, direct fluorescent antibody test (DFA), and/or polymerase chain reaction (PCR), which can be time consuming and difficult to perform. This study utilizes Cholera SMART II to determine its potential use in detection of V. cholerae. Validation of this method would help provide quick and accurate analysis for V. cholerae in ballast discharge waters in the field.

10.
Front Microbiol ; 9: 2435, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30416489

RESUMO

Conventional water resources are not sufficient in many regions to meet the needs of growing populations. Due to cyclical weather cycles, drought, and climate change, water stress has increased worldwide including in Southern California, which serves as a model for regions that integrate reuse of wastewater for both potable and non-potable use. The Orange County Water District (OCWD) Advanced Water Purification Facility (AWPF) is a highly engineered system designed to treat and produce up to 100 million gallons per day (MGD) of purified water from a municipal wastewater source for potable reuse. Routine facility microbial water quality analysis is limited to standard indicators at this and similar facilities. Given recent advances in high throughput DNA sequencing techniques, complete microbial profiling of communities in water samples is now possible. By using 16S/18S rRNA gene sequencing, metagenomic and metatranscriptomic sequencing coupled to a highly accurate identification method along with 16S rRNA gene qPCR, we describe a detailed view of the total microbial community throughout the facility. The total bacterial load of the water at stages of the treatment train ranged from 3.02 × 106 copies in source, unchlorinated wastewater feed to 5.49 × 101 copies of 16S rRNA gene/mL after treatment (consisting of microfiltration, reverse osmosis, and ultraviolet/advanced oxidation). Microbial diversity and load decreased by several orders of magnitude after microfiltration and reverse osmosis treatment, falling to almost non-detectable levels that more closely resembled controls of molecular grade laboratory water than the biomass detected in the source water. The presence of antibiotic resistance genes and viruses was also greatly reduced. Overall, system design performance was achieved, and comprehensive microbial community analysis was found to enable a more complete characterization of the water/wastewater microbial signature.

11.
Artigo em Inglês | MEDLINE | ID: mdl-30309013

RESUMO

The cholera epidemic that occurred in Haiti post-earthquake in 2010 has resulted in over 9000 deaths during the past eight years. Currently, morbidity and mortality rates for cholera have declined, but cholera cases still occur on a daily basis. One continuing issue is an inability to accurately predict and identify when cholera outbreaks might occur. To explore this surveillance gap, a metagenomic approach employing environmental samples was taken. In this study, surface water samples were collected at two time points from several sites near the original epicenter of the cholera outbreak in the Central Plateau of Haiti. These samples underwent whole genome sequencing and subsequent metagenomic analysis to characterize the microbial community of bacteria, fungi, protists, and viruses, and to identify antibiotic resistance and virulence associated genes. Replicates from sites were analyzed by principle components analysis, and distinct genomic profiles were obtained for each site. Cholera toxin converting phage was detected at one site, and Shiga toxin converting phages at several sites. Members of the Acinetobacter family were frequently detected in samples, including members implicated in waterborne diseases. These results indicate a metagenomic approach to evaluating water samples can be useful for source tracking and the surveillance of pathogens such as Vibrio cholerae over time, as well as for monitoring virulence factors such as cholera toxin.

12.
Environ Monit Assess ; 190(10): 565, 2018 Sep 03.
Artigo em Inglês | MEDLINE | ID: mdl-30178153

RESUMO

The objective of this study was to determine environmental parameters driving Vibrio populations in the estuarine zone of the Bengal delta. Spatio-temporal data were collected at river estuary, mangrove, beach, pond, and canal sites. Effects of salinity, tidal amplitude, and a cyclone and tsunami were included in the study. Vibrio population shifts were found to be correlated with tide-driven salinity and suspended particulate matter (SPM). Increased abundance of Vibrio spp. in surface water was observed after a cyclone, attributed to re-suspension of benthic particulate organic carbon (POC), and increased availability of chitin and dissolved organic carbon (DOC). Approximately a two log10 increase in the (p < 0.05) number of Vibrio spp. was observed in < 20 µm particulates, compared with microphytoplankton (20-60 µm) and zooplankton > 60 µm fractions. Benthic and suspended sediment comprised a major reservoir of Vibrio spp. Results of microcosm experiments showed enhanced growth of vibrios was related to concentration of organic matter in SPM. It is concluded that SPM, POC, chitin, and salinity significantly influence abundance and distribution of vibrios in the Bengal delta estuarine zone.


Assuntos
Clima , Processos Climáticos , Estuários , Rios/química , Vibrio/crescimento & desenvolvimento , Água/química , Áreas Alagadas , Animais , Ásia , Carbono , Quitina , Tempestades Ciclônicas , Monitoramento Ambiental , Sedimentos Geológicos , Material Particulado , Plâncton , Dinâmica Populacional , Salinidade , Cloreto de Sódio , Tsunamis , Zooplâncton
13.
Front Microbiol ; 9: 1361, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29988506

RESUMO

Background: There is a growing move to provide care for premature infants in a single family, private room neonatal intensive care unit (NICU) in place of the traditional shared space, open bay NICU. The resultant effect on the developing neonatal microbiota is unknown. Study Design: Stool and groin skin swabs were collected from infants in a shared-space NICU (old NICU) and a single-family room NICU (new NICU) on the same hospital campus. Metagenomic sequencing was performed and data analyzed by CosmosID bioinformatics software package. Results: There were no significant differences between the cohorts in gestational age, length of stay, and delivery mode; infants in the old NICU received significantly more antibiotics (p = 0.03). Differentially abundant antimicrobial resistance genes and virulence associated genes were found between the cohorts in stool and skin, with more differentially abundant antimicrobial resistance genes in the new NICU. The entire bacterial microbiota analyzed to the genus level significantly differed between cohorts in skin (p = 0.0001) but not in stool samples. There was no difference in alpha diversity between the two cohorts. DNA viruses and fungi were detected but did not differ between cohorts. Conclusion: Differences were seen in the resistome and virulome between the two cohorts with more differentially abundant antimicrobial resistance genes in the new NICU. This highlights the influence that different NICU environments can have on the neonatal microbiota. Whether the differences were due to the new NICU being a single-family NICU or located in a newly constructed building warrants exploration. Long term health outcomes from the differences observed must be followed longitudinally.

14.
Biol Rev Camb Philos Soc ; 93(4): 1813-1831, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-29732670

RESUMO

Environmentally transmitted diseases are comparatively poorly understood and managed, and their ecology is particularly understudied. Here we identify challenges of studying environmental transmission and persistence with a six-sided interdisciplinary review of the biology of anthrax (Bacillus anthracis). Anthrax is a zoonotic disease capable of maintaining infectious spore banks in soil for decades (or even potentially centuries), and the mechanisms of its environmental persistence have been the topic of significant research and controversy. Where anthrax is endemic, it plays an important ecological role, shaping the dynamics of entire herbivore communities. The complex eco-epidemiology of anthrax, and the mysterious biology of Bacillus anthracis during its environmental stage, have necessitated an interdisciplinary approach to pathogen research. Here, we illustrate different disciplinary perspectives through key advances made by researchers working in Etosha National Park, a long-term ecological research site in Namibia that has exemplified the complexities of the enzootic process of anthrax over decades of surveillance. In Etosha, the role of scavengers and alternative routes (waterborne transmission and flies) has proved unimportant relative to the long-term persistence of anthrax spores in soil and their infection of herbivore hosts. Carcass deposition facilitates green-ups of vegetation to attract herbivores, potentially facilitated by the role of anthrax spores in the rhizosphere. The underlying seasonal pattern of vegetation, and herbivores' immune and behavioural responses to anthrax risk, interact to produce regular 'anthrax seasons' that appear to be a stable feature of the Etosha ecosystem. Through the lens of microbiologists, geneticists, immunologists, ecologists, epidemiologists, and clinicians, we discuss how anthrax dynamics are shaped at the smallest scale by population genetics and interactions within the bacterial communities up to the broadest scales of ecosystem structure. We illustrate the benefits and challenges of this interdisciplinary approach to disease ecology, and suggest ways anthrax might offer insights into the biology of other important pathogens. Bacillus anthracis, and the more recently emerged Bacillus cereus biovar anthracis, share key features with other environmentally transmitted pathogens, including several zoonoses and panzootics of special interest for global health and conservation efforts. Understanding the dynamics of anthrax, and developing interdisciplinary research programs that explore environmental persistence, is a critical step forward for understanding these emerging threats.

15.
Anaerobe ; 2018 Apr 22.
Artigo em Inglês | MEDLINE | ID: mdl-29689301

RESUMO

The gut microbiome influences many, if not all, aspects of human health. Antibiotics, while lifesaving, have the unintended consequence of killing commensal microbiota inhabiting the gastrointestinal (GI) tract, which can lead to overgrowth of opportunistic pathogens such as Clostridium difficile and emergence of antibiotic-resistant organisms. Here, porcine models were developed to evaluate changes to the gut microbiome caused by two distinct types of beta-lactam antibiotics delivered via common administration routes, oral amoxicillin and intravenous ertapenem. Amoxicillin is one of the most often used broad-spectrum antibiotics, frequently prescribed to young children. Ertapenem, a carbapenem considered a last resort antibiotic, is used sparingly in humans and prohibited for use in animals. Cohorts of normal pigs (n = 5) were treated with amoxicillin (20 mg/kg, PO, BID) or ertapenem (30 mg/kg, IV, SID) for seven days. Microbiomes were evaluated using whole genome shotgun metagenomics analyses of fecal DNA collected prior to, during, and after antibiotic treatment. Each antibiotic resulted in significant and distinct changes in the microbiome, causing elimination of key commensal bacterial species and overgrowth of other, potentially pathogenic taxa. In addition, amoxicillin promoted propagation of a broad range of antibiotic resistance genes, many encoding efflux pump components and beta-lactamases, while ertapenem triggered emergence of genes encoding vancomycin resistance, and beta-lactamases, including the carbapenemase, IMP-27. Notably, microbiota alterations and antibiotic resistance gene propagation displayed unique patterns following exposure to amoxicillin or ertapenem. These data underscore the importance of understanding consequences of individual antibiotic use to predict and potentially mitigate adverse outcomes. The porcine models developed here can facilitate evaluation of therapeutic interventions to prevent antibiotic-mediated microbiome disruption.

16.
MBio ; 9(2)2018 04 17.
Artigo em Inglês | MEDLINE | ID: mdl-29666284

RESUMO

Vibrio cholerae, an estuarine bacterium, is the causative agent of cholera, a severe diarrheal disease that demonstrates seasonal incidence in Bangladesh. In an extensive study of V. cholerae occurrence in a natural aquatic environment, water and plankton samples were collected biweekly between December 2005 and November 2006 from Mathbaria, an estuarine village of Bangladesh near the mangrove forests of the Sundarbans. Toxigenic V. cholerae exhibited two seasonal growth peaks, one in spring (March to May) and another in autumn (September to November), corresponding to the two annual seasonal outbreaks of cholera in this region. The total numbers of bacteria determined by heterotrophic plate count (HPC), representing culturable bacteria, accounted for 1% to 2.7% of the total numbers obtained using acridine orange direct counting (AODC). The highest bacterial culture counts, including toxigenic V. cholerae, were recorded in the spring. The direct fluorescent antibody (DFA) assay was used to detect V. cholerae O1 cells throughout the year, as free-living cells, within clusters, or in association with plankton. V. cholerae O1 varied significantly in morphology, appearing as distinctly rod-shaped cells in the spring months, while small coccoid cells within thick clusters of biofilm were observed during interepidemic periods of the year, notably during the winter months. Toxigenic V. cholerae O1 was culturable in natural water during the spring when the temperature rose sharply. The results of this study confirmed biofilms to be a means of persistence for bacteria and an integral component of the annual life cycle of toxigenic V. cholerae in the estuarine environment of Bangladesh.IMPORTANCEVibrio cholerae, the causative agent of cholera, is autochthonous in the estuarine aquatic environment. This study describes morphological changes in naturally occurring V. cholerae O1 in the estuarine environment of Mathbaria, where the bacterium is culturable when the water temperature rises and is observable predominantly as distinct rods and dividing cells. In the spring and fall, these morphological changes coincide with the two seasonal peaks of endemic cholera in Bangladesh. V. cholerae O1 cells are predominantly coccoid within biofilms but are rod shaped as free-living cells and when attached to plankton or to particulate matter in interepidemic periods of the year. It is concluded that biofilms represent a stage of the annual life cycle of V. cholerae O1, the causative agent of cholera in Bangladesh.

17.
Genome Announc ; 6(10)2018 Mar 08.
Artigo em Inglês | MEDLINE | ID: mdl-29519847

RESUMO

We report here the first draft genome sequence of the non-O1/non-O139 Vibrio cholerae strain VcN1, isolated from Dhaka, Bangladesh. The data submitted to GenBank for this strain will contribute to advancing our understanding of this environmentally disseminated bacterium, including its virulence and its evolution as an important pathogen.

18.
Res Microbiol ; 169(1): 1-10, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-28888938

RESUMO

Africa is currently an important region in which cholera epidemics occur. Little is known about the presence of Vibrio cholerae in freshwater bodies in Africa. There are ca. 1700 lakes and reservoirs in Burkina Faso, most of which have been built within recent decades to secure water resources. The purpose of this study was to investigate the presence of V. cholerae in the water of reservoirs, using the most-probable-number polymerase chain reaction. Results showed that V. cholerae could be detected in water samples collected from 14 of 39 sampled reservoirs. The concentrations varied from 0 MPN/l to more than 1100 MPN/l. Fifty strains of V. cholerae isolated on CHROMagar™ vibrio were identified as V. cholerae non-O1/non-O139, none of which carried the ctxA gene. A significant positive correlation was found between the presence of V. cholerae in the reservoirs and both alkaline pH and phytoplankton biomass. V. cholerae was present in significantly higher numbers in reservoirs of urban areas than in rural areas. Since V. cholerae non-O1/non-O139 has been shown to be a causative agent of endemic diarrheal outbreaks, their presence in Burkina Faso reservoirs suggests they may play a role in gastroenteritis in that country.


Assuntos
Água Doce/microbiologia , Vibrio cholerae/isolamento & purificação , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Burkina Faso , Cólera/microbiologia , Humanos , Vibrio cholerae/classificação , Vibrio cholerae/genética , Vibrio cholerae/metabolismo , Poluição da Água , Recursos Hídricos
19.
Front Microbiol ; 8: 2283, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29218035

RESUMO

We report the complete sequence of two novel plasmids, pSDH-1 and pSDH-2, isolated from clinical Vibrio cholerae non-O1/non-O139 during the early phase of the 2010 Haitian cholera epidemic. Plasmids were revealed by employing single-cell genomics and their genome content suggests self-mobilization and, for pSDH-2, a toxin-antitoxin (TA) system for plasmid stabilization was identified. The putative origin of replication of pSDH-2 was mapped suggesting it replicates following the ColE1 model of plasmid replication. pSDH-1 and pSDH-2 were widespread among environmental V. cholerae non-O1/non-O139 with variable prevalence in four Haitian Departments. pSDH-2 was the most common element, either alone or with pSDH-1. The two plasmids detection adds to the composite scenario of mobile genetic elements (MGEs) observed in V. cholerae in Haiti. The role these small cryptic plasmids circulating in Vibrio spp. play in bacterial fitness or pathogenicity merits further investigation.

20.
Sci Rep ; 7(1): 16324, 2017 Nov 24.
Artigo em Inglês | MEDLINE | ID: mdl-29176730

RESUMO

Blowflies and houseflies are mechanical vectors inhabiting synanthropic environments around the world. They feed and breed in fecal and decaying organic matter, but the microbiome they harbour and transport is largely uncharacterized. We sampled 116 individual houseflies and blowflies from varying habitats on three continents and subjected them to high-coverage, whole-genome shotgun sequencing. This allowed for genomic and metagenomic analyses of the host-associated microbiome at the species level. Both fly host species segregate based on principal coordinate analysis of their microbial communities, but they also show an overlapping core microbiome. Legs and wings displayed the largest microbial diversity and were shown to be an important route for microbial dispersion. The environmental sequencing approach presented here detected a stochastic distribution of human pathogens, such as Helicobacter pylori, thereby demonstrating the potential of flies as proxies for environmental and public health surveillance.

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