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Genomic characterization of an Escherichia coli O157:H7 strain linked to leafy greens-associated outbreaks dates its emergence to late 2015. One clade has notable accessory genomic content and a previously described mutation putatively associated with increased arsenic tolerance. This strain is a reoccurring, emerging, or persistent strain causing illness over an extended period.
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Escherichia coli O157 , Escherichia coli O157/genética , Surtos de Doenças , Genômica , MutaçãoRESUMO
BACKGROUND: Plasmids are mobile genetic elements, key in the dissemination of antibiotic resistance, virulence determinants and other adaptive traits in bacteria. Obtaining a robust method for plasmid classification is necessary to better understand the genetics and epidemiology of many pathogens. Until now, plasmid classification systems focused on specific traits, which limited their precision and universality. The definition of plasmid taxonomic units (PTUs), based on average nucleotide identity metrics, allows the generation of a universal plasmid classification scheme, applicable to all bacterial taxa. Here we present COPLA, a software able to assign plasmids to known and novel PTUs, based on their genomic sequence. RESULTS: We implemented an automated pipeline able to assign a given plasmid DNA sequence to its cognate PTU, and assessed its performance using a sample of 1000 unclassified plasmids. Overall, 41% of the samples could be assigned to a previously defined PTU, a number that reached 63% in well-known taxa such as the Enterobacterales order. The remaining plasmids represent novel PTUs, indicating that a large fraction of plasmid backbones is still uncharacterized. CONCLUSIONS: COPLA is a bioinformatic tool for universal, species-independent, plasmid classification. Offered both as an automatable pipeline and an open web service, COPLA will help bacterial geneticists and clinical microbiologists to quickly classify plasmids.
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Transferência Genética Horizontal , Genômica , Resistência Microbiana a Medicamentos , Plasmídeos/genética , Fatores de VirulênciaRESUMO
Enterobacteriaceae, quinolone resistance is largely attributed to mutations in the quinolone resistance-determining regions (QRDR) of gyrA, gyrB, parC, and parE, and plasmid-italiciated quinolone resistance (PMQR) genes (e.g., qnr genes, aac(6')-Ib-cr, or qepA). .
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Escherichia coli O26 has been identified as the most common non-O157 Shiga toxin-producing E. coli (STEC) serogroup to cause human illnesses in the United States and has been implicated in outbreaks around the world. E. coli has high genomic plasticity, which facilitates the loss or acquisition of virulence genes. Attaching and effacing E. coli (AEEC) O26 strains have frequently been isolated from bovine feces, and there is a need to better characterize the relatedness of these strains to defined molecular pathotypes and to describe the extent of their genetic diversity. High-throughput real-time PCR was used to screen 178 E. coli O26 isolates from a single U.S. cattle feedlot, collected from May to July 2011, for the presence or absence of 25 O26 serogroup-specific and virulence-associated markers. The selected markers were capable of distinguishing these strains into molecularly defined groups (yielding 18 unique marker combinations). Analysis of the clustered regularly interspaced short palindromic repeat 1 (CRISPR1) and CRISPR2a loci further discriminated isolates into 24 CRISPR types. The combination of molecular markers and CRISPR typing provided 20.8% diversity. The recent CRISPR PCR target SP_O26-E, which was previously identified only in stx2-positive O26:H11 human clinical strains, was identified in 96.4% (161/167 [95% confidence interval, 99.2 to 93.6%]) of the stx-negative AEEC O26:H11 bovine fecal strains. This supports that these stx-negative strains may have previously contained a prophage carrying stx or could acquire this prophage, thus possibly giving them the potential to become pathogenic to humans. These results show that investigation of specific genetic markers may further elucidate our understanding of the genetic diversity of AEEC O26 strains in bovine feces.
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Doenças dos Bovinos/epidemiologia , Doenças dos Bovinos/microbiologia , Infecções por Escherichia coli/veterinária , Fezes/microbiologia , Variação Genética , Escherichia coli Shiga Toxigênica/classificação , Escherichia coli Shiga Toxigênica/isolamento & purificação , Animais , Bovinos , Infecções por Escherichia coli/epidemiologia , Infecções por Escherichia coli/microbiologia , Genótipo , Ensaios de Triagem em Larga Escala , Tipagem Molecular , Reação em Cadeia da Polimerase em Tempo Real , Sorogrupo , Escherichia coli Shiga Toxigênica/genética , Estados Unidos , Fatores de Virulência/genéticaRESUMO
Bacterial relatedness measured using select chromosomal loci forms the basis of public health genomic surveillance. While approximating vertical evolution through this approach has proven exceptionally valuable for understanding pathogen dynamics, it excludes a fundamental dimension of bacterial evolution-horizontal gene transfer. Incorporating the accessory genome is the logical remediation and has recently shown promise in expanding epidemiological resolution for enteric pathogens. Employing k-mer-based Jaccard index analysis, and a novel genome length distance metric, we computed pangenome (i.e., core and accessory) relatedness for the globally important pathogen Salmonella enterica serotype Typhi (Typhi), and graphically express both vertical (homology-by-descent) and horizontal (homology-by-admixture) evolutionary relationships in a reticulate network of over 2,200 U.S. Typhi genomes. This analysis revealed non-random structure in the Typhi pangenome that is driven predominantly by the gain and loss of mobile genetic elements, confirming and expanding upon known epidemiological patterns, revealing novel plasmid dynamics, and identifying avenues for further genomic epidemiological exploration. With an eye to public health application, this work adds important biological context to the rapidly improving ways of analyzing bacterial genetic data and demonstrates the value of the accessory genome to infer pathogen epidemiology and evolution.IMPORTANCEGiven bacterial evolution occurs in both vertical and horizontal dimensions, inclusion of both core and accessory genetic material (i.e., the pangenome) is a logical step toward a more thorough understanding of pathogen dynamics. With an eye to public, and indeed, global health relevance, we couple contemporary tools for genomic analysis with decades of research on mobile genetic elements to demonstrate the value of the pangenome, known and unknown, annotated, and hypothetical, for stratification of Salmonella enterica serovar Typhi (Typhi) populations. We confirm and expand upon what is known about Typhi epidemiology, plasmids, and antimicrobial resistance dynamics, and offer new avenues of exploration to further deduce Typhi ecology and evolution, and ultimately to reduce the incidence of human disease.
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Colistin is a last-resort antibiotic used to treat infections caused by multidrug-resistant Gram-negative bacteria. People with a history of travel to the Dominican Republic have become sick with pathogenic bacteria carrying the mobile colistin resistance gene, mcr-1, during and after traveling. This investigation aimed to identify mcr genes in Enterobacteriaceae isolated from food animal sources in the Dominican Republic. Three hundred and eleven samples were tested, from which 1354 bacterial isolates were obtained. Real-time PCR tests showed that 70.7% (220 out of 311) of the samples and 3.2% (44 out of 1354) of the isolates tested positive for the mcr gene. All RT-PCR presumptive mcr-positive isolates (n = 44) and a subset (n = 133) of RT-PCR presumptive mcr-negative isolates were subjected to whole-genome sequencing. WGS analysis showed that 39 isolates carried the mcr gene, with 37 confirmed as positive through RT-PCR and two as negative. Further, all of the mcr-positive genomes were identified as Escherichia coli and all contained a IncX4 plasmid replicon. Resistant determinants for other antibiotics important for human health were found in almost all isolates carrying mcr genes.
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Enterobacteriaceae , Proteínas de Escherichia coli , Animais , Humanos , Colistina/farmacologia , República Dominicana/epidemiologia , Farmacorresistência Bacteriana/genética , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Escherichia coli , Plasmídeos , Proteínas de Escherichia coli/genética , Testes de Sensibilidade MicrobianaRESUMO
Background: The Global Typhoid Genomics Consortium was established to bring together the typhoid research community to aggregate and analyse Salmonella enterica serovar Typhi (Typhi) genomic data to inform public health action. This analysis, which marks 22 years since the publication of the first Typhi genome, represents the largest Typhi genome sequence collection to date (n=13,000). Methods: This is a meta-analysis of global genotype and antimicrobial resistance (AMR) determinants extracted from previously sequenced genome data and analysed using consistent methods implemented in open analysis platforms GenoTyphi and Pathogenwatch. Results: Compared with previous global snapshots, the data highlight that genotype 4.3.1 (H58) has not spread beyond Asia and Eastern/Southern Africa; in other regions, distinct genotypes dominate and have independently evolved AMR. Data gaps remain in many parts of the world, and we show the potential of travel-associated sequences to provide informal 'sentinel' surveillance for such locations. The data indicate that ciprofloxacin non-susceptibility (>1 resistance determinant) is widespread across geographies and genotypes, with high-level ciprofloxacin resistance (≥3 determinants) reaching 20% prevalence in South Asia. Extensively drug-resistant (XDR) typhoid has become dominant in Pakistan (70% in 2020) but has not yet become established elsewhere. Ceftriaxone resistance has emerged in eight non-XDR genotypes, including a ciprofloxacin-resistant lineage (4.3.1.2.1) in India. Azithromycin resistance mutations were detected at low prevalence in South Asia, including in two common ciprofloxacin-resistant genotypes. Conclusions: The consortium's aim is to encourage continued data sharing and collaboration to monitor the emergence and global spread of AMR Typhi, and to inform decision-making around the introduction of typhoid conjugate vaccines (TCVs) and other prevention and control strategies. Funding: No specific funding was awarded for this meta-analysis. Coordinators were supported by fellowships from the European Union (ZAD received funding from the European Union's Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement No 845681), the Wellcome Trust (SB, Wellcome Trust Senior Fellowship), and the National Health and Medical Research Council (DJI is supported by an NHMRC Investigator Grant [GNT1195210]).
Salmonella Typhi (Typhi) is a type of bacteria that causes typhoid fever. More than 110,000 people die from this disease each year, predominantly in areas of sub-Saharan Africa and South Asia with limited access to safe water and sanitation. Clinicians use antibiotics to treat typhoid fever, but scientists worry that the spread of antimicrobial-resistant Typhi could render the drugs ineffective, leading to increased typhoid fever mortality. The World Health Organization has prequalified two vaccines that are highly effective in preventing typhoid fever and may also help limit the emergence and spread of resistant Typhi. In low resource settings, public health officials must make difficult trade-off decisions about which new vaccines to introduce into already crowded immunization schedules. Understanding the local burden of antimicrobial-resistant Typhi and how it is spreading could help inform their actions. The Global Typhoid Genomics Consortium analyzed 13,000 Typhi genomes from 110 countries to provide a global overview of genetic diversity and antimicrobial-resistant patterns. The analysis showed great genetic diversity of the different strains between countries and regions. For example, the H58 Typhi variant, which is often drug-resistant, has spread rapidly through Asia and Eastern and Southern Africa, but is less common in other regions. However, distinct strains of other drug-resistant Typhi have emerged in other parts of the world. Resistance to the antibiotic ciprofloxacin was widespread and accounted for over 85% of cases in South Africa. Around 70% of Typhi from Pakistan were extensively drug-resistant in 2020, but these hard-to-treat variants have not yet become established elsewhere. Variants that are resistant to both ciprofloxacin and ceftriaxone have been identified, and azithromycin resistance has also appeared in several different variants across South Asia. The Consortium's analyses provide valuable insights into the global distribution and transmission patterns of drug-resistant Typhi. Limited genetic data were available fromseveral regions, but data from travel-associated cases helped fill some regional gaps. These findings may help serve as a starting point for collective sharing and analyses of genetic data to inform local public health action. Funders need to provide ongoing supportto help fill global surveillance data gaps.
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Salmonella typhi , Febre Tifoide , Humanos , Salmonella typhi/genética , Febre Tifoide/epidemiologia , Antibacterianos/farmacologia , Viagem , Farmacorresistência Bacteriana/genética , CiprofloxacinaRESUMO
The objective of this study was to characterize the epidemiology of Salmonella carried by dairy cows culled from herds in the Texas High Plains. Feces were collected from a convenience sample of 706 animals culled from nine dairy farms. In addition, individually paired fecal and hide samples were collected from 70 healthy milking cows on three of the dairies. Samples were cultured for Salmonella using routine methods; isolates were serotyped and subjected to a panel of antimicrobial drugs to determine susceptibility. Salmonella was recovered from 32.6% of culled cows. Whole-herd use of a vaccine containing siderophore receptors and porin proteins was associated (p=0.05) with reduced Salmonella prevalence in that the prevalence among herds that practiced whole-herd vaccination was 8.0% compared to 36.8% among herds that did not use this vaccine. The majority (88.6%) of isolates were pansusceptible or resistant to one drug. Of the 3.1% of isolates resistant to more than four drugs, all were Salmonella Newport and were recovered from one dairy. Various serotypes were recovered from individual fecal and hide samples. Salmonella Montevideo was recovered more frequently (p<0.01) from hide samples, whereas Salmonella Cerro was recovered more frequently (p<0.01) from feces. Salmonella was recovered from at least one cow on all dairies. While our study was not a priori designed to address herd-level factors, we found evidence that the whole-herd use of a siderophore receptor and porin protein-containing vaccine might be a useful aid in the control of Salmonella in groups of cattle. As this is a nonrandomized evaluation of an intervention, other herd-level factors that may be correlated with vaccine use, such as biosecurity, might have been responsible for the observed association.
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Vacinas Bacterianas/imunologia , Bovinos/microbiologia , Indústria de Laticínios , Farmacorresistência Bacteriana , Salmonella/classificação , Salmonella/isolamento & purificação , Animais , Antibacterianos/farmacologia , Carga Bacteriana/veterinária , Proteínas da Membrana Bacteriana Externa/antagonistas & inibidores , Proteínas de Bactérias/antagonistas & inibidores , Bovinos/imunologia , Farmacorresistência Bacteriana Múltipla , Fezes/microbiologia , Imunidade Coletiva , Testes de Sensibilidade Microbiana/veterinária , Porinas/antagonistas & inibidores , Receptores de Superfície Celular/antagonistas & inibidores , Salmonella/efeitos dos fármacos , Sorotipagem/veterinária , Pele/microbiologia , TexasRESUMO
In the United States, reports of Salmonella enterica carrying mcr-1 remain rare in humans, but when observed, the infection is often associated with travel. Here, we report 14 mcr-1-positive Salmonella enterica isolates from patients in the United States that reported travel to the Dominican Republic within the 12 months before illness.
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Despite being linked to a number of recent poultry-associated outbreaks in the United States, few reference genomes are available for Salmonella enterica serotype Hadar. Here, we address this need by reporting 18 Salmonella Hadar genomes from samples collected from patients in the United States between 2014 and 2020.
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Identification of genes encoding ß-lactamases (BLs) from short-read sequences remains challenging due to the high frequency of shared amino acid functional domains and motifs in proteins encoded by BL genes and related non-BL gene sequences. Divergent BL homologs can be frequently missed during similarity searches, which has important practical consequences for monitoring antibiotic resistance. To address this limitation, we built ROCker models that targeted broad classes (e.g., class A, B, C, and D) and individual families (e.g., TEM) of BLs and challenged them with mock 150-bp- and 250-bp-read data sets of known composition. ROCker identifies most-discriminant bit score thresholds in sliding windows along the sequence of the target protein sequence and hence can account for nondiscriminative domains shared by unrelated proteins. BL ROCker models showed a 0% false-positive rate (FPR), a 0% to 4% false-negative rate (FNR), and an up-to-50-fold-higher F1 score [2 × precision × recall/(precision + recall)] compared to alternative methods, such as similarity searches using BLASTx with various e-value thresholds and BL hidden Markov models, or tools like DeepARG, ShortBRED, and AMRFinder. The ROCker models and the underlying protein sequence reference data sets and phylogenetic trees for read placement are freely available through http://enve-omics.ce.gatech.edu/data/rocker-bla. Application of these BL ROCker models to metagenomics, metatranscriptomics, and high-throughput PCR gene amplicon data should facilitate the reliable detection and quantification of BL variants encoded by environmental or clinical isolates and microbiomes and more accurate assessment of the associated public health risk, compared to the current practice. IMPORTANCE Resistance genes encoding ß-lactamases (BLs) confer resistance to the widely prescribed antibiotic class ß-lactams. Therefore, it is important to assess the prevalence of BL genes in clinical or environmental samples for monitoring the spreading of these genes into pathogens and estimating public health risk. However, detecting BLs in short-read sequence data is technically challenging. Our ROCker model-based bioinformatics approach showcases the reliable detection and typing of BLs in complex data sets and thus contributes toward solving an important problem in antibiotic resistance surveillance. The ROCker models developed substantially expand the toolbox for monitoring antibiotic resistance in clinical or environmental settings.
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Antibacterianos , beta-Lactamases , Humanos , beta-Lactamases/genética , Filogenia , Antibacterianos/farmacologia , beta-Lactamas , Resistência Microbiana a MedicamentosRESUMO
Between 2018 and 2019, Salmonella enterica serotype Reading caused a large, multistate outbreak linked to contact with raw turkey products in the United States. Here, we provide five Salmonella Reading reference genomes collected from US patients between 2016 and 2018.
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From a global view of antimicrobial resistance over different sectors, seafood and the marine environment are often considered as potential reservoirs of antimicrobial resistance genes (ARGs) and mobile genetic elements (MGEs); however, there are few studies and sparse results on this sector. This study aims to provide new data and insights regarding the content of resistance markers in various seafood samples and sources, and therefore the potential exposure to humans in a global One Health approach. An innovative high throughput qPCR screening was developed and validated in order to simultaneously investigate the presence of 41 ARGs and 33 MGEs including plasmid replicons, integrons, and insertion sequences in Gram-negative bacteria. Analysis of 268 seafood isolates from the bacterial microflora of cod (n = 24), shellfish (n = 66), flat fishes (n = 53), shrimp (n = 10), and horse mackerel (n = 115) show the occurrence of sul-1, ant(3â³)-Ia, aph(3')-Ia, strA, strB, dfrA1, qnrA, and blaCTX-M-9 genes in Pseudomonas spp., Providencia spp., Klebsiella spp., Proteus spp., and Shewanella spp. isolates and the presence of MGEs in all bacterial species investigated. We found that the occurrence of MGE may be associated with the seafood type and the environmental, farming, and harvest conditions. Moreover, even if MGE were detected in half of the seafood isolates investigated, association with ARG was only identified for twelve isolates. The results corroborate the hypothesis that the incidence of antimicrobial-resistant bacteria (ARB) and ARG decreases with increasing distance from potential sources of fecal contamination. This unique and original high throughput micro-array designed for the screening of ARG and MGE in Gram-negative bacteria could be easily implementable for monitoring antimicrobial resistance gene markers in diverse contexts.
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Salmonella enterica serovar Fresno is an infrequently isolated serovar whose ecology and genomic characteristics have not yet been described. To further understand the genomic characteristics of this serovar, we sequenced the complete genome of a single isolate recovered from a bovine lymph node at harvest.
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Salmonella enterica subsp. enterica serovar Dublin is a host-adapted pathogen for cattle that can cause invasive disease in humans. To facilitate genomic comparisons characterizing virulence determinants of this pathogen, we present the complete genome sequences of three S. Dublin strains isolated from bovine sources at harvest.
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Shiga toxin-producing Escherichia coli of serotype O26:H11/H- constitute a diverse group of strains and several clones with distinct genetic characteristics have been identified and characterized. Whole genome sequencing was performed using Illumina and PacBio technologies on eight stx2-positive O26:H11 strains circulating in France. Comparative analyses of the whole genome of the stx2-positive O26:H11 strains indicate that several clones of EHEC O26:H11 are co-circulating in France. Phylogenetic analysis of the French strains together with stx2-positive and stx-negative E. coli O26:H11 genomes obtained from Genbank indicates the existence of four clonal complexes (SNP-CCs) separated in two distinct lineages, one of which comprises the "new French clone" (SNP-CC1) that appears genetically closely related to stx-negative attaching and effacing E. coli (AEEC) strains. Interestingly, the whole genome SNP (wgSNP) phylogeny is summarized in the cas gene phylogeny, and a simple qPCR assay targeting the CRISPR array specific to SNP-CC1 (SP_O26-E) can distinguish between the two main lineages. The PacBio sequencing allowed a detailed analysis of the mobile genetic elements (MGEs) of the strains. Numerous MGEs were identified in each strain, including a large number of prophages and up to four large plasmids, representing overall 8.7-19.8% of the total genome size. Analysis of the prophage pool of the strains shows a considerable diversity with a complex history of recombination. Each clonal complex (SNP-CC) is characterized by a unique set of plasmids and phages, including stx-prophages, suggesting evolution through separate acquisition events. Overall, the MGEs appear to play a major role in O26:H11 intra-serotype clonal diversification.
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Use, overuse, and misuse of antimicrobials contributes to selection and dissemination of bacterial resistance determinants that may be transferred to humans and constitute a global public health concern. Because of the continued emergence and expansion of antimicrobial resistance, combined with the lack of novel antimicrobial agents, efforts are underway to preserve the efficacy of current available life-saving antimicrobials in humans. As a result, uses of medically important antimicrobials in food animal production have generated debate and led to calls to reduce both antimicrobial use and the need for use. This manuscript, commissioned by the World Health Organization (WHO) to help inform the development of the WHO guidelines on the use of medically important antimicrobials in food animals, includes three illustrations of antimicrobial use in food animal production that has contributed to the selection-and subsequent transfer-of resistance determinants from food animals to humans. Herein, antimicrobial use and the epidemiology of bacterial resistance are described for streptothricins, glycopeptides, and colistin. Taken together, these historical and current narratives reinforce the need for actions that will preserve the efficacy of antimicrobials.
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To more fully characterize the burden of Salmonella enterica in bovine peripheral lymph nodes (PLN), PLN (n = 5,450) were collected from healthy cattle at slaughter in 12 commercial abattoirs that slaughtered feedlot-fattened (FF) cattle exclusively (n = 7), cattle removed (or culled) from breeding herds (n = 3), or both FF and cull cattle (n = 2). Qualitative and quantitative methods were used to estimate prevalence and concentration of Salmonella in PLN. Isolates were subjected to a variety of phenotypic, serological, and molecular assays. Overall, Salmonella prevalence in PLN from FF and cull cattle was 7.1 and 1.8%. However, burden varied by season in that observed prevalence in PLN collected in cooler or warmer seasons was 2.4 and 8.2%, respectively. Prevalence in PLN from cull cattle in the southwest region of the US was 2.1 and 1.1% for cool and warm seasons, respectively; however, prevalence in FF PLN was far greater in that it was 6.5 and 31.1%, respectively. Salmonella was recovered from 289 (5.6%) PLN and 2.9% (n = 160) of all PLN tested had quantifiable concentrations that varied from 1.6 to 4.9 log10 colony forming units/PLN. The most common serotypes isolated from PLN were Montevideo (26.9%), Lille (14.9%), Cerro (13.0%), Anatum (12.8%), and Dublin (6.9%). In all, 376 unique isolates were collected from the 289 Salmonella-positive PLN. Antimicrobial susceptibility testing revealed the majority (80.6%) of these isolates were pansusceptible; however, 10.7% of isolates were found to be resistant to two or more antimicrobial classes. We were able to document an observed increased in prevalence of Salmonella in PLN during the warmer season, particularly in FF cattle from the southwest region of the US. The mechanisms underlying the observed association between season, region, and production source have yet to be elucidated. Nevertheless, these findings increase our understanding of the sources of contamination of beef products and shed light on transmission dynamics that may be useful in targeting these sources.
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Current methods for screening Enterohemorrhagic Escherichia coli (EHEC) O157 and non-O157 in beef enrichments typically rely on the molecular detection of stx, eae, and serogroup-specific wzx or wzy gene fragments. As these genetic markers can also be found in some non-EHEC strains, a number of "false positive" results are obtained. Here, we explore the suitability of five novel molecular markers, espK, espV, ureD, Z2098, and CRISPRO26:H11 as candidates for a more accurate screening of EHEC strains of greater clinical significance in industrialized countries. Of the 1739 beef enrichments tested, 180 were positive for both stx and eae genes. Ninety (50%) of these tested negative for espK, espV, ureD, and Z2098, but 12 out of these negative samples were positive for the CRISPRO26:H11 gene marker specific for a newly emerging virulent EHEC O26:H11 French clone. We show that screening for stx, eae, espK, and espV, in association with the CRISPRO26:H11 marker is a better approach to narrow down the EHEC screening step in beef enrichments. The number of potentially positive samples was reduced by 48.88% by means of this alternative strategy compared to the European and American reference methods, thus substantially improving the discriminatory power of EHEC screening systems. This approach is in line with the EFSA (European Food Safety Authority) opinion on pathogenic STEC published in 2013.