ABSTRACT
BACKGROUND: Nosocomial infections pose a considerable risk to patients who are susceptible, and this is particularly acute in intensive care units when hospital-associated bacteria are endemic. During the first wave of the COVID-19 pandemic, the surge of patients presented a significant obstacle to the effectiveness of infection control measures. We aimed to assess the risks and extent of nosocomial pathogen transmission under a high patient burden by designing a novel bacterial pan-pathogen deep-sequencing approach that could be integrated with standard clinical surveillance and diagnostics workflows. METHODS: We did a prospective cohort study in a region of northern Italy that was severely affected by the first wave of the COVID-19 pandemic. Inpatients on both ordinary and intensive care unit (ICU) wards at the San Matteo hospital, Pavia were sampled on multiple occasions to identify bacterial pathogens from respiratory, nasal, and rectal samples. Diagnostic samples collected between April 7 and May 10, 2020 were cultured on six different selective media designed to enrich for Acinetobacter baumannii, Escherichia coli, Enterococcus faecium, Enterococcus faecalis, Klebsiella pneumoniae, Pseudomonas aeruginosa, Staphylococcus aureus, and Streptococcus pneumoniae, and DNA from each plate with positive growth was deep sequenced en masse. We used mSWEEP and mGEMS to bin sequencing reads by sequence cluster for each species, followed by mapping with snippy to generate high quality alignments. Antimicrobial resistance genes were detected by use of ARIBA and CARD. Estimates of hospital transmission were obtained from pairwise bacterial single nucleotide polymorphism distances, partitioned by within-patient and between-patient samples. Finally, we compared the accuracy of our binned Acinetobacter baumannii genomes with those obtained by single colony whole-genome sequencing of isolates from the same hospital. FINDINGS: We recruited patients from March 1 to May 7, 2020. The pathogen population among the patients was large and diverse, with 2148 species detections overall among the 2418 sequenced samples from the 256 patients. In total, 55 sequence clusters from key pathogen species were detected at least five times. The antimicrobial resistance gene prevalence was correspondingly high, with key carbapenemase and extended spectrum ß-lactamase genes detected in at least 50 (40%) of 125 patients in ICUs. Using high-resolution mapping to infer transmission, we established that hospital transmission was likely to be a significant mode of acquisition for each of the pathogen species. Finally, comparison with single colony Acinetobacter baumannii genomes showed that the resolution offered by deep sequencing was equivalent to single-colony sequencing, with the additional benefit of detection of co-colonisation of highly similar strains. INTERPRETATION: Our study shows that a culture-based deep-sequencing approach is a possible route towards improving future pathogen surveillance and infection control at hospitals. Future studies should be designed to directly compare the accuracy, cost, and feasibility of culture-based deep sequencing with single colony whole-genome sequencing on a range of bacterial species. FUNDING: Wellcome Trust, European Research Council, Academy of Finland Flagship program, Trond Mohn Foundation, and Research Council of Norway.
ABSTRACT
Since 2016, Yemen has been experiencing the largest cholera outbreak in modern history. Multidrug resistance (MDR) emerged among Vibrio cholerae isolates from cholera patients in 2018. Here, to characterize circulating genotypes, we analysed 260 isolates sampled in Yemen between 2018 and 2019. Eighty-four percent of V. cholerae isolates were serogroup O1 belonging to the seventh pandemic El Tor (7PET) lineage, sub-lineage T13, whereas 16% were non-toxigenic, from divergent non-7PET lineages. Treatment of severe cholera with macrolides between 2016 and 2019 coincided with the emergence and dominance of T13 subclones carrying an incompatibility type C (IncC) plasmid harbouring an MDR pseudo-compound transposon. MDR plasmid detection also in endemic non-7PET V. cholerae lineages suggested genetic exchange with 7PET epidemic strains. Stable co-occurrence of the IncC plasmid with the SXT family of integrative and conjugative element in the 7PET background has major implications for cholera control, highlighting the importance of genomic epidemiological surveillance to limit MDR spread.
Subject(s)
Cholera , Vibrio cholerae O1 , Humans , Cholera/epidemiology , Vibrio cholerae O1/genetics , Yemen/epidemiology , Plasmids/genetics , GenomicsABSTRACT
Healthcare-associated infections (HCAIs) affect the most vulnerable people in society and are increasingly difficult to treat in the face of mounting antimicrobial resistance (AMR). Routine surveillance represents an effective way of understanding the circulation and burden of bacterial resistance and transmission in hospital settings. Here, we used whole-genome sequencing (WGS) to retrospectively analyse carbapenemase-producing Gram-negative bacteria from a single hospital in the UK over 6 years (n=165). We found that the vast majority of isolates were either hospital-onset (HAI) or HCAI. Most carbapenemase-producing organisms were carriage isolates, with 71â% isolated from screening (rectal) swabs. Using WGS, we identified 15 species, the most common being Escherichia coli and Klebsiella pneumoniae. Only one significant clonal outbreak occurred during the study period and involved a sequence type (ST)78 K. pneumoniae carrying bla NDM-1 on an IncFIB/IncHI1B plasmid. Contextualization with public data revealed little evidence of this ST outside of the study hospital, warranting ongoing surveillance. Carbapenemase genes were found on plasmids in 86â% of isolates, the most common types being bla NDM- and bla OXA-type alleles. Using long-read sequencing, we determined that approximately 30â% of isolates with carbapenemase genes on plasmids had acquired them via horizontal transmission. Overall, a national framework to collate more contextual genomic data, particularly for plasmids and resistant bacteria in the community, is needed to better understand how carbapenemase genes are transmitted in the UK.
Subject(s)
Hospitals , Klebsiella pneumoniae , Humans , Retrospective Studies , Plasmids/genetics , Klebsiella pneumoniae/genetics , Escherichia coli/genetics , Genomics , United Kingdom/epidemiologyABSTRACT
Shigella sonnei causes shigellosis, a severe gastrointestinal illness that is sexually transmissible among men who have sex with men (MSM). Multidrug resistance in S. sonnei is common including against World Health Organisation recommended treatment options, azithromycin, and ciprofloxacin. Recently, an MSM-associated outbreak of extended-spectrum ß-lactamase producing, extensively drug resistant S. sonnei was reported in the United Kingdom. Here, we aimed to identify the genetic basis, evolutionary history, and international dissemination of the outbreak strain. Our genomic epidemiological analyses of 3,304 isolates from the United Kingdom, Australia, Belgium, France, and the United States of America revealed an internationally connected outbreak with a most recent common ancestor in 2018 carrying a low-fitness cost resistance plasmid, previously observed in travel associated sublineages of S. flexneri. Our results highlight the persistent threat of horizontally transmitted antimicrobial resistance and the value of continuing to work towards early and open international sharing of genomic surveillance data.
Subject(s)
Sexual and Gender Minorities , Shigella , Male , Humans , Shigella sonnei/genetics , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/therapeutic use , Homosexuality, Male , Travel , Drug Resistance, Bacterial/genetics , Microbial Sensitivity TestsABSTRACT
BACKGROUND: Patients with prolonged hospitalisation have a significant risk of carriage of and subsequent infection with extended spectrum ß-lactamase (ESBL)-producing and carbapenemase-producing Klebsiella pneumoniae. However, the distinctive roles of the community and hospital environments in the transmission of ESBL-producing or carbapenemase-producing K pneumoniae remain elusive. We aimed to investigate the prevalence and transmission of K pneumoniae within and between the two tertiary hospitals in Hanoi, Viet Nam, using whole-genome sequencing. METHODS: We did a prospective cohort study of 69 patients in intensive care units (ICUs) from two hospitals in Hanoi, Viet Nam. Patients were included if they were aged 18 years or older, admitted for longer than the mean length of stay in their ICU, and cultured K pneumoniae from their clinical samples. Longitudinally collected samples from patients (collected weekly) and the ICU environment (collected monthly) were cultured on selective media, and whole-genome sequences from K pneumoniae colonies analysed. We did phylogenetic analyses and correlated phenotypic antimicrobial susceptibility testing with genotypic features of K pneumoniae isolates. We constructed transmission networks of patient samples, relating ICU admission times and locations with genetic similarity of infecting K pneumoniae. FINDINGS: Between June 1, 2017, and Jan 31, 2018, 69 patients were in the ICUs and eligible for inclusion, and a total of 357 K pneumoniae isolates were cultured and successfully sequenced. 228 (64%) of K pneumoniae isolates carried between two and four different ESBL-encoding and carbapenemase-encoding genes, with 164 (46%) isolates carrying genes encoding both, with high minimum inhibitory concentrations. We found a novel co-occurrence of blaKPC-2 and blaNDM-1 in 46·6% of samples from the globally successful ST15 lineage. Despite being physically and clinically separated, the two hospitals shared closely related strains carrying the same array of antimicrobial resistance genes. INTERPRETATION: These results highlight the high prevalence of ESBL-positive carbapenem-resistant K pneumoniae in ICUs in Viet Nam. Through studying K pneumoniae ST15 in detail, we showed how important resistance genes are contained within these strains that are carried broadly by patients entering the two hospitals directly or through referral. FUNDING: Medical Research Council Newton Fund, Ministry of Science and Technology, Wellcome Trust, Academy of Medical Sciences, Health Foundation, and National Institute for Health and Care Research Cambridge Biomedical Research Centre.
Subject(s)
Klebsiella pneumoniae , Humans , Klebsiella pneumoniae/genetics , Vietnam/epidemiology , Prospective Studies , Phylogeny , Tertiary Care CentersABSTRACT
Here, this report presents two genomes of Vibrio cholerae O1 serotype Ogawa, recovered from cholera cases in Australia linked to travel to Pakistan in 2022. Their multidrug-resistant genotype represents the current activity of cholera within the seventh pandemic. One of the genome sequences was assembled using both short- and long-read sequences.
ABSTRACT
The plasmid pCD-METRO confers metronidazole resistance in Clostridioides difficile. We showed high sequence similarity among pCD-METRO plasmids from different isolates and identified pCD-METRO and associated metronidazole-resistant isolates in clinical and veterinary reservoirs in the Americas. We recommend using PCR or genomic assays to detect pCD-METRO in metronidazole-resistant C. difficile.
Subject(s)
Clostridioides difficile , Clostridium Infections , Humans , Metronidazole/pharmacology , Clostridioides difficile/genetics , Ribotyping , Clostridium Infections/veterinary , Clostridium Infections/drug therapy , Clostridioides , Drug Resistance, Bacterial/genetics , Microbial Sensitivity Tests , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/therapeutic useABSTRACT
Plasmids are mobile elements that can carry genes encoding traits of clinical concern, including antimicrobial resistance (AMR) and virulence. Population-level studies of Enterobacterales, including Escherichia coli, Shigella and Klebsiella, indicate that plasmids are important drivers of lineage expansions and dissemination of AMR genes. Salmonella Typhimurium is the second most common cause of salmonellosis in humans and livestock in the UK and Europe. The long-term dynamics of plasmids between S. Typhimurium were investigated using isolates collected through national surveillance of animals in England and Wales over a 25-year period. The population structure of S. Typhimurium and its virulence plasmid (where present) were inferred through phylogenetic analyses using whole-genome sequence data for 496 isolates. Antimicrobial resistance genes and plasmid markers were detected in silico. Phenotypic plasmid characterization, using the Kado and Liu method, was used to confirm the number and size of plasmids. The differences in AMR and plasmids between clades were striking, with livestock clades more likely to carry one or more AMR plasmid and be multi-drug-resistant compared to clades associated with wildlife and companion animals. Multiple small non-AMR plasmids were distributed across clades. However, all hybrid AMR-virulence plasmids and most AMR plasmids were highly clade-associated and persisted over decades, with minimal evidence of horizontal transfer between clades. This contrasts with the role of plasmids in the short-term dissemination of AMR between diverse strains in other Enterobacterales in high-antimicrobial-use settings, with implications for predicting plasmid dissemination amongst S. Typhimurium.
Subject(s)
Anti-Infective Agents , Salmonella typhimurium , Animals , Anti-Bacterial Agents/pharmacology , Drug Resistance, Bacterial/genetics , Escherichia coli/genetics , Humans , Phylogeny , Plasmids/genetics , Salmonella typhimurium/genetics , Virulence/geneticsABSTRACT
Dissemination of antimicrobial resistance (AMR) genes by horizontal gene transfer (HGT) mediated through plasmids is a major global concern. Genomic epidemiology studies have shown varying success of different AMR plasmids during outbreaks, but the underlying reasons for these differences are unclear. Here, we investigated two Shigella plasmids (pKSR100 and pAPR100) that circulated in the same transmission network but had starkly contrasting epidemiological outcomes to identify plasmid features that may have contributed to the differences. We used plasmid comparative genomics to reveal divergence between the two plasmids in genes encoding AMR, SOS response alleviation and conjugation. Experimental analyses revealed that these genomic differences corresponded with reduced conjugation efficiencies for the epidemiologically successful pKSR100, but more extensive AMR, reduced fitness costs, and a reduced SOS response in the presence of antimicrobials, compared with the less successful pAPR100. The discrepant phenotypes between the two plasmids are consistent with the hypothesis that plasmid-associated phenotypes contribute to determining the epidemiological outcome of AMR HGT and suggest that phenotypes relevant in responding to antimicrobial pressure and fitness impact may be more important than those around conjugation in this setting. Plasmid phenotypes could thus be valuable tools in conjunction with genomic epidemiology for predicting AMR dissemination.
Subject(s)
Drug Resistance, Bacterial , Shigella , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/therapeutic use , Drug Resistance, Bacterial/genetics , Gene Transfer, Horizontal , Phenotype , Plasmids , Shigella/geneticsABSTRACT
We examined 185 complete, publicly available FII-33 plasmid sequences, characterizing their backbone and various insertions. The variable characteristic insertions facilitated evolutionary reconstruction for this plasmid group, beginning with the acquisition of a primary resistance region (PRR) over 10 years ago. FII-33 plasmids have evolved by acquiring additional resistance genes in the PRR via translocatable elements and by forming cointegrates with plasmids of other types. In all cases, IS26 is suspected to have mediated cointegration. Plasmid cointegration has contributed to the accumulation of resistance genes and may have increased the transmissibility, stability, and host range of the original FII-33 lineage. A particularly important sublineage was formed by a replicative IS26 cointegration event that fused an FII-33 plasmid with a blaKPC-2-containing R-type plasmid, interrupting the FII-33 traI gene encoding the conjugative relaxase. The FII-33:R cointegrate arose in the Klebsiella pneumoniae ST11 clone and remains largely confined there due to the abolition of transfer ability by the FII-33:R cointegration event. However, in some cases FII-33:R cointegrates have fused with additional plasmids and acquired complete transfer regions or oriT sequences that might restore their ability to transfer horizontally. Cointegration events across FII-33 plasmid sublineages have involved plasmids of at least 15 different types. This suggests that plasmid cointegration occurs readily and is more common than previously appreciated, raising questions about the effects of cointegrate formation on plasmid host range, stability, and capacity for horizontal transfer. Resources are provided for detecting and characterizing FII-33 plasmid sublineages from complete or draft genome sequences. IMPORTANCE Effective genomic surveillance of antibiotic-resistant bacterial pathogens must consider plasmids, which are frequently implicated in the accumulation and transfer of resistance genes between bacterial strains or species. However, the evolution of plasmids is complex, and simple typing or comparison tools cannot accurately determine whether plasmids belong to the same sublineages. This precludes precise tracking of plasmid movement in bacterial populations. We have examined the FII-33 group, which has been associated with multidrug resistance and particularly carbapenem resistance in clinically significant members of the Enterobacterales in China. Our analysis has provided insight into the evolution of this important plasmid group, allowing us to develop resources for rapidly typing them to the sublineage level in complete or draft genome sequences. Our approach will improve detection and characterization of FII-33 plasmids and facilitate surveillance within and outside China. The approach can serve as a model for similar studies of other plasmid types.
Subject(s)
Bacterial Proteins , DNA Transposable Elements , Bacterial Proteins/genetics , Plasmids , Drug Resistance, Multiple , GenomicsABSTRACT
The open sharing of genomic data provides an incredibly rich resource for the study of bacterial evolution and function and even anthropogenic activities such as the widespread use of antimicrobials. However, these data consist of genomes assembled with different tools and levels of quality checking, and of large volumes of completely unprocessed raw sequence data. In both cases, considerable computational effort is required before biological questions can be addressed. Here, we assembled and characterised 661,405 bacterial genomes retrieved from the European Nucleotide Archive (ENA) in November of 2018 using a uniform standardised approach. Of these, 311,006 did not previously have an assembly. We produced a searchable COmpact Bit-sliced Signature (COBS) index, facilitating the easy interrogation of the entire dataset for a specific sequence (e.g., gene, mutation, or plasmid). Additional MinHash and pp-sketch indices support genome-wide comparisons and estimations of genomic distance. Combined, this resource will allow data to be easily subset and searched, phylogenetic relationships between genomes to be quickly elucidated, and hypotheses rapidly generated and tested. We believe that this combination of uniform processing and variety of search/filter functionalities will make this a resource of very wide utility. In terms of diversity within the data, a breakdown of the 639,981 high-quality genomes emphasised the uneven species composition of the ENA/public databases, with just 20 of the total 2,336 species making up 90% of the genomes. The overrepresented species tend to be acute/common human pathogens, aligning with research priorities at different levels from individual interests to funding bodies and national and global public health agencies.
Subject(s)
Bacteria/genetics , Biodiversity , DNA, Bacterial/genetics , Data Curation , Base Sequence , Drug Resistance, Bacterial/genetics , Species SpecificityABSTRACT
BACKGROUND: Escherichia coli is a leading cause of bloodstream infections. Developing interventions to reduce E coli infections requires an understanding of the frequency of nosocomial transmission, but the available evidence is scarce. We aimed to detect and characterise transmission of E coli and associated plasmids in a hospital setting. METHODS: In this prospective observational cohort study, patients were admitted to two adult haematology wards at the Cambridge University Hospitals NHS Foundation Trust in England. Patients aged 16 years and older who were treated for haematological malignancies were included. Stool samples were collected from study participants on admission, once per week, and at discharge. We sequenced multiple E coli isolates (both extended spectrum ß-lactamase [ESBL]-producing and non-ESBL-producing) from each stool sample. A genetic threshold to infer E coli transmission was defined by maximum within-host single nucleotide polymorphism (SNP) diversity and the probability of drawing observed pairs of between-patient isolates at different SNP thresholds. Putative transmission clusters were identified when sequences were less than the genetic threshold. Epidemiological links for each transmission event were investigated. We sequenced all E coli positive blood samples from the two adult haematology wards. FINDINGS: We recruited 174 (51%) of 338 adult patients admitted to the wards between May 13 and Nov 13, 2015. We obtained and cultured 376 stool samples from 149 patients, of which 152 samples from 97 (65%) patients grew E coli. Whole-genome sequencing was done on 970 isolates. We identified extensive diversity in the bacterial population (90 sequence types) and mixed E coli sequence type carriage. 24 (26%) patients carried two sequence types, 12 (13%) carried three, and six (6%) patients carried four or more sequence types. Using a 17 SNP cutoff we identified ten clusters in 20 patients. The largest cluster contained seven patients, whereas four patients were included in multiple clusters. Strong epidemiological links were found between patients in seven clusters. 17 (11%) of 149 patients had stool samples positive for ESBL-producing E coli, the most common of which was associated with bla CTX-M-15 (12 [71%] of 17). Long-read sequencing revealed that bla CTX-M-15 was often integrated into the chromosome, with little evidence for plasmid transmission. Seven patients developed E coli bloodstream infection, four with identical strains to those in their stool; two of these had documented nosocomial acquisition. INTERPRETATION: We provide evidence of bacterial transmission and endogenous infection during routine care by integrating genomic and epidemiological data and by determining a genetic cutoff informed by within-host diversity in the studied population. Our findings challenge single colony-based investigations, and the widely accepted notion of plasmid spread. FUNDING: UK Department of Health, Wellcome Trust, UK National Institute for Health Research.
Subject(s)
Cross Infection , Escherichia coli Infections , Adult , Anti-Bacterial Agents/pharmacology , Cross Infection/epidemiology , Drug Resistance, Bacterial , Escherichia coli/genetics , Escherichia coli Infections/epidemiology , Genomics , Humans , Prospective Studies , beta-Lactamases/geneticsABSTRACT
Members of the bacterial genus Vibrio utilize chitin both as a metabolic substrate and a signal to activate natural competence. Vibrio cholerae is a bacterial enteric pathogen, sub-lineages of which can cause pandemic cholera. However, the chitin metabolic pathway in V. cholerae has been dissected using only a limited number of laboratory strains of this species. Here, we survey the complement of key chitin metabolism genes amongst 195 diverse V. cholerae. We show that the gene encoding GbpA, known to be an important colonization and virulence factor in pandemic isolates, is not ubiquitous amongst V. cholerae. We also identify a putatively novel chitinase, and present experimental evidence in support of its functionality. Our data indicate that the chitin metabolic pathway within V. cholerae is more complex than previously thought, and emphasize the importance of considering genes and functions in the context of a species in its entirety, rather than simply relying on traditional reference strains.
Subject(s)
Bacterial Proteins/genetics , Vibrio cholerae/genetics , Chitin/metabolism , Chitinases/genetics , Cholera/microbiology , Cloning, Molecular , Fimbriae Proteins/genetics , Gene Expression Regulation, Bacterial , Sequence Alignment , Vibrio cholerae/physiology , Virulence Factors/geneticsABSTRACT
Enterotoxigenic Escherichia coli (ETEC) is an enteric pathogen responsible for the majority of diarrheal cases worldwide. ETEC infections are estimated to cause 80,000 deaths annually, with the highest rates of burden, ca 75 million cases per year, amongst children under 5 years of age in resource-poor countries. It is also the leading cause of diarrhoea in travellers. Previous large-scale sequencing studies have found seven major ETEC lineages currently in circulation worldwide. We used PacBio long-read sequencing combined with Illumina sequencing to create high-quality complete reference genomes for each of the major lineages with manually curated chromosomes and plasmids. We confirm that the major ETEC lineages all harbour conserved plasmids that have been associated with their respective background genomes for decades, suggesting that the plasmids and chromosomes of ETEC are both crucial for ETEC virulence and success as pathogens. The in-depth analysis of gene content, synteny and correct annotations of plasmids will elucidate other plasmids with and without virulence factors in related bacterial species. These reference genomes allow for fast and accurate comparison between different ETEC strains, and these data will form the foundation of ETEC genomics research for years to come.
Subject(s)
Enterotoxigenic Escherichia coli/genetics , Escherichia coli Infections/microbiology , Escherichia coli Proteins/metabolism , Virulence Factors/metabolism , Antineoplastic Agents/pharmacology , Diarrhea/microbiology , Drug Resistance, Bacterial , Enterotoxigenic Escherichia coli/drug effects , Enterotoxigenic Escherichia coli/isolation & purification , Escherichia coli Infections/drug therapy , Escherichia coli Infections/genetics , Escherichia coli Proteins/genetics , Genome, Bacterial , Genomics , Humans , Phylogeny , Reference Standards , Virulence , Virulence Factors/geneticsABSTRACT
OBJECTIVES: To understand the acquisition of resistance genes by a non-GC1, non-GC2 Acinetobacter baumannii strain responsible for a 4 year outbreak at a Sydney hospital. METHODS: Representative isolates were screened for resistance to antibiotics. Three were subjected to WGS using Illumina HiSeq. One genome was completed with MinION long reads. Resistance regions were compared with known sequences using bioinformatics. RESULTS: Isolates were resistant to third-generation cephalosporins, gentamicin and tobramycin, sulfamethoxazole and erythromycin. Sequenced isolates were ST49 (Institut Pasteur scheme) and ST128 (Oxford scheme) and carried KL11 at the capsule locus and OCL8 at the lipooligosaccharide outer core locus. The complete genome of isolate J9 revealed that the resistance genes were all in plasmids; pRAY* contained aadB, and a large plasmid, pJ9-3, contained sul2 and floR genes and a dif module containing the mph(E)-msr(E) macrolide resistance genes. Transposon Tn6168, consisting of a second copy of the chromosomal ampC gene region flanked by ISAba1s, confers resistance to third-generation cephalosporins. Tn6168 is located inside the mph(E)-msr(E) dif module. pJ9-3 includes a set of four dif modules and the orientation of the pdif sites, XerC-XerD or XerD-XerC, alternates. A large transposon, Tn6175, containing tniCABDE transposition genes and genes annotated as being involved in heavy metal metabolism, uptake or export was found in the comM gene. Other ST49:ST128:KL11:OCL8 genomes found in the GenBank WGS database carried Tn6175 but neither of the plasmids carrying the resistance genes. CONCLUSIONS: An early carbapenem-susceptible A. baumannii outbreak recorded in Australia was caused by an unusual clone that had acquired plasmids carrying antibiotic resistance genes.
Subject(s)
Acinetobacter Infections , Acinetobacter baumannii , Acinetobacter Infections/epidemiology , Acinetobacter baumannii/genetics , Anti-Bacterial Agents/pharmacology , Australia/epidemiology , Disease Outbreaks , Drug Resistance, Bacterial , Hospitals , Humans , Macrolides , Plasmids/genetics , Sequence Analysis, DNAABSTRACT
Escherichia coli is a highly diverse organism that includes a range of commensal and pathogenic variants found across a range of niches and worldwide. In addition to causing severe intestinal and extraintestinal disease, E. coli is considered a priority pathogen due to high levels of observed drug resistance. The diversity in the E. coli population is driven by high genome plasticity and a very large gene pool. All these have made E. coli one of the most well-studied organisms, as well as a commonly used laboratory strain. Today, there are thousands of sequenced E. coli genomes stored in public databases. While data is widely available, accessing the information in order to perform analyses can still be a challenge. Collecting relevant available data requires accessing different sources, where data may be stored in a range of formats, and often requires further manipulation and processing to apply various analyses and extract useful information. In this study, we collated and intensely curated a collection of over 10â000 E. coli and Shigella genomes to provide a single, uniform, high-quality dataset. Shigella were included as they are considered specialized pathovars of E. coli. We provide these data in a number of easily accessible formats that can be used as the foundation for future studies addressing the biological differences between E. coli lineages and the distribution and flow of genes in the E. coli population at a high resolution. The analysis we present emphasizes our lack of understanding of the true diversity of the E. coli species, and the biased nature of our current understanding of the genetic diversity of such a key pathogen.
Subject(s)
Databases, Genetic , Escherichia coli Proteins/genetics , Escherichia coli/genetics , Shigella/genetics , Access to Information , Computational Biology/methods , Data Curation , Escherichia coli/classification , Gene Flow , Genome, Bacterial , Shigella/classificationABSTRACT
Conjugative, broad host-range plasmids of the L/M complex have been associated with antibiotic resistance since the 1970s. They are found in Gram-negative bacterial genera that cause human infections and persist in hospital environments. It is crucial that these plasmids are typed accurately so that their clinical and global dissemination can be traced in epidemiological studies. The L/M complex has previously been divided into L, M1 and M2 subtypes. However, those types do not encompass all diversity seen in the group. Here, we have examined 148 complete L/M plasmid sequences in order to understand the diversity of the complex and trace the evolution of distinct lineages. The backbone sequence of each plasmid was determined by removing translocatable genetic elements and reversing their effects in silico. The sequence identities of replication regions and complete backbones were then considered for typing. This supported the distinction of L and M plasmids and revealed that there are five L and eight M types, where each type is comprised of further sub-lineages that are distinguished by variation in their backbone and translocatable element content. Regions containing antibiotic resistance genes in L and M sub-lineages have often formed by initial rare insertion events, followed by insertion of other translocatable elements within the inceptive element. As such, islands evolve in situ to contain genes conferring resistance to multiple antibiotics. In some cases, different plasmid sub-lineages have acquired the same or related resistance genes independently. This highlights the importance of these plasmids in acting as vehicles for the dissemination of emerging resistance genes. Materials are provided here for typing plasmids of the L/M complex from complete sequences or draft genomes. This should enable rapid identification of novel types and facilitate tracking the evolution of existing lineages.
Subject(s)
Anti-Bacterial Agents , Gram-Negative Bacteria , Anti-Bacterial Agents/pharmacology , Drug Resistance, Microbial , Gram-Negative Bacteria/genetics , Humans , Plasmids/geneticsABSTRACT
Antibiotic resistant strains of Acinetobacter baumannii are responsible for a large and increasing burden of nosocomial infections in Thailand and other countries of Southeast Asia. New approaches to their control and treatment are urgently needed and an attractive strategy is to remove the bacterial polysaccharide capsule, and thus the protection from the host's immune system. To examine phylogenetic relationships, distribution of capsule chemotypes, acquired antibiotic resistance determinants, susceptibility to complement and other traits associated with systemic infection, we sequenced 191 isolates from three tertiary referral hospitals in Thailand and used phenotypic assays to characterize key aspects of infectivity. Several distinct lineages were circulating in three hospitals and the majority belonged to global clonal group 2 (GC2). Very high levels of resistance to carbapenems and other front-line antibiotics were found, as were a number of widespread plasmid replicons. A high diversity of capsule genotypes was encountered, with only three of these (KL6, KL10, and KL47) showing more than 10% frequency. Almost 90% of GC2 isolates belonged to the most common capsule genotypes and were fully resistant to the bactericidal action of human serum complement, most likely protected by their polysaccharide capsule, which represents a key determinant of virulence for systemic infection. Our study further highlights the importance to develop therapeutic strategies to remove the polysaccharide capsule from extensively drug-resistant A. baumanii during the course of systemic infection.
ABSTRACT
Most Acinetobacter plasmids are genus specific but their properties have not been investigated. Small plasmids with Rep_3 family replication initiation proteins and iterons are common in Acinetobacter baumannii and often carry antibiotic resistance genes and toxin-antitoxin systems. A RepAci1 plasmid, carrying the carbapenem resistance gene oxa23 in Tn2006 and a RepAci2 plasmid carrying the amikacin (kanamycin and neomycin) resistance gene aphA6 in TnaphA6 were identified. These two plasmids have related rep regions; the consensus 22â¯bp iteron repeats differ only at three positions and the RepA proteins are 84% identical. However, they were shown to be compatible, whereas the RepAci1 plasmid displaced another RepAci1 plasmid demonstrating that they were incompatible. Despite encoding no mobilisation proteins, the RepAci1 plasmid was transferred to a new host at low frequency when a conjugatively proficient RepAci6 plasmid was present, whereas the RepAci2 plasmid carrying mobA and mobC mobilisation genes was not. Comparison of the sequences of the mobilised and mobilising plasmids revealed a short region of high similarity that is upstream of the predicted mobilisation genes in the RepAci6 plasmid, and has an organisation similar to that of F-type oriT transfer origins. The segment carrying the oriT-like region is present in many RepAci1 plasmids, including ones carrying the cabarpenem resistance genes oxa24 or oxa58 in dif modules, and in some RepAci2 or other Rep_3 plasmids of further types, including one carrying the tet39 tetracycline resistance determinant. These plasmids are also likely to be mobilised, spreading resistance.