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: A clinical strain of Enterobacter cloacae, PIMB10EC27, isolated in Vietnam in 2010 that was resistant to 21 of 26 tested antibiotics, including carbapenems and colistin, is reported.
Abstract: We report a clinical strain of Enterobacter cloacae, PIMB10EC27, isolated in Vietnam in 2010 that was resistant to 21 of 26 tested antibiotics, including carbapenems (MICs >64 µg/mL) and colistin (MIC >128 µg/mL). The complete genome of strain PIMB10EC27 was sequenced by PacBio RSII and the Illumina Miseq system. Whole-genome analysis revealed that PIMB10EC27 contains a chromosome of the ST513 group (PIMBEC27, length 5,272,177 bp) and two plasmids, pEC27-1 of the IncX3 group (length 62,470 bp) and pEC27-2 of the IncHI1 group (length 84,602 bp). It also revealed that strain PIMB10EC27 carries 15 genes that confer resistance to at least 10 antibiotic groups. Particularly, the insertion of ISKpn19 and Tn6901 into the genomic context of blaNDM-1 was first identified and described. In another context, amino acid mutations G273D in PmrB and F515S in PmrC were first identified on the chromosome of PIMB10EC27, which may confer resistance to colistin in this strain.
11 citations
TL;DR: The emergence of mobile mcr genes mediating resistance to colistin is a critical public health issue that has hindered the treatment of serious infections caused by multidrug-resistant pathogens in humans and other animals as discussed by the authors.
Abstract: The emergence of mobile mcr genes mediating resistance to colistin is a critical public health issue that has hindered the treatment of serious infections caused by multidrug-resistant pathogens in humans and other animals. We report the emergence of the mcr-9.1 gene in a polymyxin-resistant extended-spectrum β-lactamase (ESBL)-producing Enterobacter kobei infecting a free-living franciscana dolphin (Pontoporia blainvillei), threatened with extinction in South America. Genomic analysis confirmed the presence of genes conferring resistance to clinically relevant β-lactam [blaCTX-M-15 , blaACT-9 , blaOXA-1 and blaTEM-1B ], aminoglycoside [aac(3)-IIa, aadA1, aph(3'')-Ib and aph(6)-Id], trimethoprim [dfrA14], tetracycline [tetA], quinolone [aac(6')-Ib-cr and qnrB1], fosfomycin [fosA], sulphonamide [sul2] and phenicol [catA1 and catB3] antibiotics. The identification of mcr-9.1 in a CTX-M-15-producing pathogen infecting a critically endangered animal is of serious concern, which should be interpreted as a sign of further spread of critical priority pathogens and their resistance genes in threatened ecosystems.
11 citations
01 Jan 2021
TL;DR: It is found that individual ST131 had small differences in AMR gene content relative to a larger shared resistome, and comparisons with a range of plasmids common in ST131 showed that AMRGene composition was driven by conjugation, recombination and mobile genetic elements.
Abstract: The human gut microbiome includes beneficial, commensal and pathogenic bacteria that possess antimicrobial resistance (AMR) genes and exchange these predominantly through conjugative plasmids. Escherichia coli is a significant component of the gastrointestinal microbiome and is typically non-pathogenic in this niche. In contrast, extra-intestinal pathogenic E. coli (ExPEC) including ST131 may occupy other environments like the urinary tract or bloodstream where they express genes enabling AMR and host cell adhesion like type 1 fimbriae. The extent to which commensal E. coli and uropathogenic ExPEC ST131 share AMR genes remains understudied at a genomic level, and we examined this here using a preterm infant resistome. We found that individual ST131 had small differences in AMR gene content relative to a larger shared resistome. Comparisons with a range of plasmids common in ST131 showed that AMR gene composition was driven by conjugation, recombination and mobile genetic elements. Plasmid pEK499 had extended regions in most ST131 Clade C isolates, and it had evidence of a co-evolutionary signal based on protein-level interactions with chromosomal gene products, as did pEK204 that had a type IV fimbrial pil operon. ST131 possessed extensive diversity of selective type 1, type IV, P and F17-like fimbriae genes that was highest in subclade C2. The structure and composition of AMR genes, plasmids and fimbriae vary widely in ST131 Clade C and this may mediate pathogenicity and infection outcomes.
11 citations
Cites result from "In Silico Detection and Typing of P..."
...Replicon typing used PlasmidFinder [56] and each plasmid was compared to the CARD....
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...Extensive plasmid rearrangements in closely related ST131 Clade C genomes To evaluate the origin and relationships of the AMR genes in these seven Clade C (Table 1), we initially found with PlasmidFinder [86] that all had IncF1A and some C2 had lost IncF1B, suggesting potential plasmid changes (Table S3)....
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...This dataset includes the PlasmidFinder results....
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...To evaluate the origin and relationships of the AMR genes in these seven Clade C (Table 1), we initially found with PlasmidFinder [86] that all had IncF1A and some C2 had lost IncF1B, suggesting potential plasmid changes (Table S3)....
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01 Jan 2021
TL;DR: Although the ST155 lineage is common in poultry from Africa and South America, the Gambian ST155 isolates belong to a unique cgMLST cluster comprising closely related (38–39 alleles differences) isolates from poultry and livestock from sub-Saharan Africa – suggesting that strains can be exchanged betweenoultry and livestock in this setting.
Abstract: Chickens and guinea fowl are commonly reared in Gambian homes as affordable sources of protein. Using standard microbiological techniques, we obtained 68 caecal isolates of Escherichia coli from 10 chickens and 9 guinea fowl in rural Gambia. After Illumina whole-genome sequencing, 28 sequence types were detected in the isolates (4 of them novel), of which ST155 was the most common (22/68, 32 %). These strains span four of the eight main phylogroups of E. coli, with phylogroups B1 and A being most prevalent. Nearly a third of the isolates harboured at least one antimicrobial resistance gene, while most of the ST155 isolates (14/22, 64 %) encoded resistance to ≥3 classes of clinically relevant antibiotics, as well as putative virulence factors, suggesting pathogenic potential in humans. Furthermore, hierarchical clustering revealed that several Gambian poultry strains were closely related to isolates from humans. Although the ST155 lineage is common in poultry from Africa and South America, the Gambian ST155 isolates belong to a unique cgMLST cluster comprising closely related (38-39 alleles differences) isolates from poultry and livestock from sub-Saharan Africa - suggesting that strains can be exchanged between poultry and livestock in this setting. Continued surveillance of E. coli and other potential pathogens in rural backyard poultry from sub-Saharan Africa is warranted.
11 citations
TL;DR: Wang et al. as mentioned in this paper reported the prevalence and molecular epidemiology of tet(X)-positive bacteria (TPB) from diverse sources, investigate the host-specificity of TPB and the transferability of Tet(X).
11 citations
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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