In Silico Detection and Typing of Plasmids using PlasmidFinder and Plasmid Multilocus Sequence Typing
01 Jul 2014-Antimicrobial Agents and Chemotherapy (American Society for Microbiology)-Vol. 58, Iss: 7, pp 3895-3903
TL;DR: Two easy-to-use Web tools for in silico detection and characterization of whole-genome sequence (WGS) and whole-plasmid sequence data from members of the family Enterobacteriaceae are designed and developed.
Abstract: In the work presented here, we designed and developed two easy-to-use Web tools for in silico detection and characterization of whole-genome sequence (WGS) and whole-plasmid sequence data from members of the family Enterobacteriaceae. These tools will facilitate bacterial typing based on draft genomes of multidrug-resistant Enterobacteriaceae species by the rapid detection of known plasmid types. Replicon sequences from 559 fully sequenced plasmids associated with the family Enterobacteriaceae in the NCBI nucleotide database were collected to build a consensus database for integration into a Web tool called PlasmidFinder that can be used for replicon sequence analysis of raw, contig group, or completely assembled and closed plasmid sequencing data. The PlasmidFinder database currently consists of 116 replicon sequences that match with at least at 80% nucleotide identity all replicon sequences identified in the 559 fully sequenced plasmids. For plasmid multilocus sequence typing (pMLST) analysis, a database that is updated weekly was generated from www.pubmlst.org and integrated into a Web tool called pMLST. Both databases were evaluated using draft genomes from a collection of Salmonella enterica serovar Typhimurium isolates. PlasmidFinder identified a total of 103 replicons and between zero and five different plasmid replicons within each of 49 S . Typhimurium draft genomes tested. The pMLST Web tool was able to subtype genomic sequencing data of plasmids, revealing both known plasmid sequence types (STs) and new alleles and ST variants. In conclusion, testing of the two Web tools using both fully assembled plasmid sequences and WGS-generated draft genomes showed them to be able to detect a broad variety of plasmids that are often associated with antimicrobial resistance in clinically relevant bacterial pathogens.
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TL;DR: It is observed that horizontal gene transfer is likely to have played an overwhelming role in species diversification, favouring a public-goods-like mechanism of gene ‘acquisition and resampling’ over a tree-of-life-like vertical inheritance model of evolution.
Abstract: Campylobacter hyointestinalis is a member of an emerging group of zoonotic Campylobacter spp. that are increasingly identified in both gastric and non-gastric disease in humans. Here, we discovered C. hyointestinalis in three separate classes of New Zealand ruminant livestock; cattle, sheep and deer. To investigate the relevance of these findings we performed a systematic literature review on global C. hyointestinalis epidemiology and used comparative genomics to better understand and classify members of the species. We found that C. hyointestinalis subspecies hyointestinalis has an open pangenome, with accessory gene contents involved in many essential processes such as metabolism, virulence and defence. We observed that horizontal gene transfer is likely to have played an overwhelming role in species diversification, favouring a public-goods-like mechanism of gene 'acquisition and resampling' over a tree-of-life-like vertical inheritance model of evolution. As a result, simplistic gene-based inferences of taxonomy by similarity are likely to be misleading. Such genomic plasticity will also mean that local evolutionary histories likely influence key species characteristics, such as host-association and virulence. This may help explain geographical differences in reported C. hyointestinalis epidemiology and limits what characteristics may be generalised, requiring further genomic studies of C. hyointestinalis in areas where it causes disease.
27 citations
01 Jul 2020
TL;DR: This study highlights that the transparency of global surveillance genomic data could accelerate understanding of virulence and antimicrobial resistance makeup of a previously unknown threat.
Abstract: Salmonella is composed of a wide variety of serovars, causing human self-limited gastrointestinal illnesses or invasive infections Invasive non-typhoidal Salmonella (iNTS) is well documented, with high mortality for children and immunocompromised adults in sub-Saharan Africa and has recently been reported in Southeast Asia However, iNTS in China remains unknown In May 2019, a case of invasive infection caused by Salmonella enterica serovar Uzaramo (S Uzaramo) was reported for the first time in China Phylogenomic analysis was performed by genomic sequencing the available contextualized isolates, which separated the two Chinese strains into different sublineages Both phenotypic and genomic characterization demonstrated that the S Uzaramo isolates showed in general low antimicrobial resistance potential, except one isolated from lake-water in China Additional comparative genomic analysis and Caenorhabditis elegans killing assays suggested a unique combination of virulence factors, including typhoid toxin and tcf fimbrial adhesin, which might play a role in the invasive infection This study highlights that the transparency of global surveillance genomic data could accelerate understanding of virulence and antimicrobial resistance makeup of a previously unknown threat
27 citations
Cites methods from "In Silico Detection and Typing of P..."
...The assembled contigs were analysed for plasmids and antimicrobial resistance genes using the CGE PlasmidFinder database (similarity 95 %) [28] and ResFinder database (similarity 90 %) through ABRicate v....
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TL;DR: The presence of Cronobacter spp.
27 citations
TL;DR: Pathogenwatch as mentioned in this paper is a web application for real-time analysis of Klebsiella genomes to aid surveillance at local, national, and global levels, which can identify hospital outbreaks, detect introductions from abroad, and uncover clonal expansions associated with resistance and virulence genes.
Abstract: Background Klebsiella species, including the notable pathogen K. pneumoniae, are increasingly associated with antimicrobial resistance (AMR). Genome-based surveillance can inform interventions aimed at controlling AMR. However, its widespread implementation requires tools to streamline bioinformatic analyses and public health reporting. Methods We developed the web application Pathogenwatch, which implements analytics tailored to Klebsiella species for integration and visualization of genomic and epidemiological data. We populated Pathogenwatch with 16 537 public Klebsiella genomes to enable contextualization of user genomes. We demonstrated its features with 1636 genomes from 4 low- and middle-income countries (LMICs) participating in the NIHR Global Health Research Unit (GHRU) on AMR. Results Using Pathogenwatch, we found that GHRU genomes were dominated by a small number of epidemic drug-resistant clones of K. pneumoniae. However, differences in their distribution were observed (eg, ST258/512 dominated in Colombia, ST231 in India, ST307 in Nigeria, ST147 in the Philippines). Phylogenetic analyses including public genomes for contextualization enabled retrospective monitoring of their spread. In particular, we identified hospital outbreaks, detected introductions from abroad, and uncovered clonal expansions associated with resistance and virulence genes. Assessment of loci encoding O-antigens and capsule in K. pneumoniae, which represent possible vaccine candidates, showed that 3 O-types (O1-O3) represented 88.9% of all genomes, whereas capsule types were much more diverse. Conclusions Pathogenwatch provides a free, accessible platform for real-time analysis of Klebsiella genomes to aid surveillance at local, national, and global levels. We have improved representation of genomes from GHRU participant countries, further facilitating ongoing surveillance.
27 citations
17 Jan 2020
TL;DR: Findings suggest that the acquisition of blaNDM-1-bearing plasmid structure (p18-43_01), horizontal transfer and clonal dissemination facilitate the spread of carbapenemases in South Africa, emphasizing the importance of targeted infection control measures to prevent dissemination.
Abstract: Whole-genome sequence (WGS) analyses were employed to investigate the genomic epidemiology of extensively drug-resistant Klebsiella pneumoniae strains, focusing on the carbapenem resistance-encoding determinants, mobile genetic support, clonal and epidemiological relationships. A total of ten isolates were obtained from patients admitted to the intensive care unit (ICU) in a public hospital in South Africa. Five isolates were from rectal swabs of colonized patients and five from blood cultures of patients with invasive carbapenem-resistant infections. Following microbial identification and antibiotic susceptibility tests, the isolates were subjected to WGS on the Illumina MiSeq platform. All the isolates showed genotypic resistance to tested β-lactams (NDM-1, OXA-1, CTX-M-15, TEM-1B, SHV-1) and other antibiotics. All but one isolate belonged to the ST152 with a novel sequence type, ST3136, differing by a single-locus variant. The isolates had the same plasmid multilocus sequence type (IncF[K12:A-:B36]) and capsular serotype (KL149), supporting the epidemiological linkage between the clones. Resistance to carbapenems in the 10 isolates was conferred by the blaNDM-1 mediated by the acquisition of multi-replicon [ColRNAI, IncFIB(pB171), Col440I, IncFII, IncFIB(K) and IncFII(Yp)] p18-43_01 plasmid. These findings suggest that the acquisition of blaNDM-1-bearing plasmid structure (p18-43_01), horizontal transfer and clonal dissemination facilitate the spread of carbapenemases in South Africa. This emphasizes the importance of targeted infection control measures to prevent dissemination.
27 citations
Additional excerpts
...WGS Identification of Mobile Genetic Elements (MGEs)/Genetic Support Plasmid replicons were predicted through PlasmidFinder [38]....
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...WGS analysis via the PlasmidFinder online platform revealed different plasmid replicon types (Inc FIB(K), Inc FII, Inc FIB (pB171), Inc FII(Yp), ColRNAI, and Col440I) grouped into two different combinations in the CRKP isolates (Table 3)....
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...Plasmid replicons were predicted through PlasmidFinder [38]....
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References
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TL;DR: A web server providing a convenient way of identifying acquired antimicrobial resistance genes in completely sequenced isolates was created, and the method was evaluated on WGS chromosomes and plasmids of 30 isolates.
Abstract: Objectives
Identification of antimicrobial resistance genes is important for understanding the underlying mechanisms and the epidemiology of antimicrobial resistance. As the costs of whole-genome sequencing (WGS) continue to decline, it becomes increasingly available in routine diagnostic laboratories and is anticipated to substitute traditional methods for resistance gene identification. Thus, the current challenge is to extract the relevant information from the large amount of generated data.
3,956 citations
"In Silico Detection and Typing of P..." refers methods in this paper
...To extract the relevant information from the large amount of data generated, a Web-based tool, ResFinder, for the identification of acquired or intrinsically present antimicrobial resistance genes in whole-genome data was recently developed (15)....
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TL;DR: NCBI’s Conserved Domain Database (CDD) is a resource for the annotation of protein sequences with the location of conserved domain footprints, and functional sites inferred from these footprints.
Abstract: NCBI's Conserved Domain Database (CDD) is a resource for the annotation of protein sequences with the location of conserved domain footprints, and functional sites inferred from these footprints. CDD includes manually curated domain models that make use of protein 3D structure to refine domain models and provide insights into sequence/structure/function relationships. Manually curated models are organized hierarchically if they describe domain families that are clearly related by common descent. As CDD also imports domain family models from a variety of external sources, it is a partially redundant collection. To simplify protein annotation, redundant models and models describing homologous families are clustered into superfamilies. By default, domain footprints are annotated with the corresponding superfamily designation, on top of which specific annotation may indicate high-confidence assignment of family membership. Pre-computed domain annotation is available for proteins in the Entrez/Protein dataset, and a novel interface, Batch CD-Search, allows the computation and download of annotation for large sets of protein queries. CDD can be accessed via http://www.ncbi.nlm.nih.gov/Structure/cdd/cdd.shtml.
2,934 citations
"In Silico Detection and Typing of P..." refers background in this paper
...In particular, the replicase proteins showing the pfam02387 or pfam01051 conserved domains were assigned to the FII and FIB groups, respectively (31)....
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TL;DR: Results indicated that the inc/rep PCR method demonstrates high specificity and sensitivity in detecting replicons on reference plasmids and also revealed the presence of recurrent and common plasmid in epidemiologically unrelated Salmonella isolates of different serotypes.
2,163 citations
"In Silico Detection and Typing of P..." refers methods in this paper
...A collection of 24 previously characterized and fully FIG 1 Numbers of fully sequenced plasmids (y axis) classified into incompatibility groups occurring in the different bacterial species of the Enterobacteriaceae family....
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...Since 2005, a PCR-based replicon typing (PBRT) scheme has been available that targets in multiplex PCRs the replicons of the major plasmid families occurring in members of the family Enterobacteriaceae (2)....
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...Here, we present two free, easy-to-use Web tools, PlasmidFinder and pMLST, to analyze and classify plasmids from bacterial species of the family Enterobacteriaceae....
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...Here, we describe the design of two new easy-to-use Web tools useful for the rapid identification of plasmids in Enterobacteriaceae species that are of interest for epidemiological and clinical microbiology investigations of the plasmid-associated spread of antimicrobial resistance....
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...This method was initially developed to detect the replicons of plasmids belonging to the 18 major incompatibility (Inc) groups of Enterobacteriaceae species (3)....
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TL;DR: The Bacterial Isolate Genome Sequence Database (BIGSDB) represents a freely available resource that will assist the broader community in the elucidation of the structure and function of bacteria by means of a population genomics approach.
Abstract: The opportunities for bacterial population genomics that are being realised by the application of parallel nucleotide sequencing require novel bioinformatics platforms These must be capable of the storage, retrieval, and analysis of linked phenotypic and genotypic information in an accessible, scalable and computationally efficient manner The Bacterial Isolate Genome Sequence Database (BIGSDB) is a scalable, open source, web-accessible database system that meets these needs, enabling phenotype and sequence data, which can range from a single sequence read to whole genome data, to be efficiently linked for a limitless number of bacterial specimens The system builds on the widely used mlstdbNet software, developed for the storage and distribution of multilocus sequence typing (MLST) data, and incorporates the capacity to define and identify any number of loci and genetic variants at those loci within the stored nucleotide sequences These loci can be further organised into 'schemes' for isolate characterisation or for evolutionary or functional analyses Isolates and loci can be indexed by multiple names and any number of alternative schemes can be accommodated, enabling cross-referencing of different studies and approaches LIMS functionality of the software enables linkage to and organisation of laboratory samples The data are easily linked to external databases and fine-grained authentication of access permits multiple users to participate in community annotation by setting up or contributing to different schemes within the database Some of the applications of BIGSDB are illustrated with the genera Neisseria and Streptococcus The BIGSDB source code and documentation are available at http://pubmlstorg/software/database/bigsdb/
Genomic data can be used to characterise bacterial isolates in many different ways but it can also be efficiently exploited for evolutionary or functional studies BIGSDB represents a freely available resource that will assist the broader community in the elucidation of the structure and function of bacteria by means of a population genomics approach
1,943 citations
TL;DR: A Web-based method for MLST of 66 bacterial species based on whole-genome sequencing data that enables investigators to determine the sequence types of their isolates on the basis of WGS data.
Abstract: Accurate strain identification is essential for anyone working with bacteria. For many species, multilocus sequence typing (MLST) is considered the “gold standard” of typing, but it is traditionally performed in an expensive and time-consuming manner. As the costs of whole-genome sequencing (WGS) continue to decline, it becomes increasingly available to scientists and routine diagnostic laboratories. Currently, the cost is below that of traditional MLST. The new challenges will be how to extract the relevant information from the large amount of data so as to allow for comparison over time and between laboratories. Ideally, this information should also allow for comparison to historical data. We developed a Web-based method for MLST of 66 bacterial species based on WGS data. As input, the method uses short sequence reads from four sequencing platforms or preassembled genomes. Updates from the MLST databases are downloaded monthly, and the best-matching MLST alleles of the specified MLST scheme are found using a BLAST-based ranking method. The sequence type is then determined by the combination of alleles identified. The method was tested on preassembled genomes from 336 isolates covering 56 MLST schemes, on short sequence reads from 387 isolates covering 10 schemes, and on a small test set of short sequence reads from 29 isolates for which the sequence type had been determined by traditional methods. The method presented here enables investigators to determine the sequence types of their isolates on the basis of WGS data. This method is publicly available at www.cbs.dtu.dk/services/MLST.
1,620 citations
"In Silico Detection and Typing of P..." refers methods in this paper
...If raw sequence reads are uploaded, they are first assembled (after the sequencing platform is given by the user) as described previously (16)....
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