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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01 Jan 2019
TL;DR: A novel approach to computationally identify new BHR plasmids was applied, in which the method searched for highly similar cognates found in multiple taxa within a comprehensive plasmid database.
Abstract: Describing the role of plasmids and their contribution to the exchange of genetic material among bacteria is essential for understanding the fields of plasmid epidemiology, microbial ecology, and commercial and synthetic microbiology. Broad-host-range (BHR) plasmids are those that are found not only in a single bacterial species, but in members of different taxonomic groups and are of significant interest to researchers in many fields. We applied a novel approach to computationally identify new BHR plasmids, in which we searched for highly similar cognate plasmids within a comprehensive plasmid database. After identifying 125 plasmid groups with highly similar cognates found in multiple taxa, we closely examined BHR plasmids found in multiple families. The majority of our identified BHR plasmids are found in members of the Enterobacteriaceae and closely related taxa, while three BHR plasmids of potential commercial significance were found in two species of Cyanobacteria. One plasmid with an exceptionally broad host range was found in both Gram-positive and Gram-negative bacterial species. This analysis demonstrates the utility of this method in identifying new BHR plasmids while highlighting unknown ranges of previously documented plasmids.
10 citations
Cites background or methods from "In Silico Detection and Typing of P..."
...Nordmann P, Naas T, Poirel L. Global spread of Carbapenemaseproducing Enterobacteriaceae....
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...Within the broad range of taxa represented in the comprehensive database, there was a heavy skew towards plasmids from medically relevant groupings, particularly the family Enterobacteriaceae, thus potentially limiting our ability to identify potential BHR plasmids....
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...Carattoli A. Resistance plasmid families in Enterobacteriaceae....
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...The pCAV plasmid, found in a number of human pathogens as well as the soil bacterium Citrobacter freundii, was found in additional human pathogens as well as several common plant pathogens, all from the family Enterobacteriaceae or closely related taxa....
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...Two antibiotic resistance encoding plasmids were found in members of the family Enterobacteriaceae and closely related families A beta-lactamase encoding plasmid was found in three entries of Enterobacter cloacae and one of Pantoea species [41] and when including additional BLAST results has an expanded range including a cognate found in the human pathogen Pseudomonas aeruginosa (File S2)....
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TL;DR: In this paper, the authors carried out a sentinel testing survey on the intestinal carriage of plasmid-mediated colistin-resistant Enterobacteriaceae in hospitalized patients.
Abstract: Background: In order to estimate the prevalence of plasmid borne colistin resistance and to characterize in detail the mcr-positive isolates, we carried out a sentinel testing survey on the intestinal carriage of plasmid-mediated colistin-resistant Enterobacteriaceae in hospitalized patients. Methods: Between June 2018 and September 2019, 1922 faecal samples from hospitalised patients were analysed by selective culture in presence of colistin (3.5 mg/L), and in parallel by direct detection of the mcr-1 to mcr-8 genes by qPCR. The mcr-positive isolates were characterised by whole-genome sequencing. Results: The prevalence of the mcr-1 gene was 0.21% (n = 4/1922); the mcr-2 to 8 genes were not detected. The mcr-1 gene was found to be localised in the IncX4 (n = 3) and IncHI2 (n = 1) plasmid type. One Escherichia coli isolate was susceptible to colistin due to the inactivation of the mcr-1 gene through the insertion of the IS2 element; however, the colistin resistance was inducible by culture in low concentrations of colistin. One human mcr-1 positive E. coli isolate was related genetically to the mcr-1 E. coli isolate derived from turkey meat of Czech origin. Conclusions:mcr-mediated colistin resistance currently poses little threat to patients hospitalised in Czech healthcare settings. The presence of the mcr-1 gene in the human population has a possible link to domestically produced, retail meat.
10 citations
TL;DR: Insight is provided into microbial profiling and antibiotic-resistant strains of chicken rinsate samples during poultry processing that feature resistance to the cephems class of antibiotics.
10 citations
TL;DR: In this paper , the authors reported increased antimicrobial resistance in E. coli O157:H7, which is an important food-borne and waterborne pathogen that causes hemorrhagic colitis and the hemolytic-uremic syndrome in humans and may cause serious morbidity and large outbreaks worldwide.
Abstract: Escherichia coli O157:H7 is an important food-borne and water-borne pathogen that causes hemorrhagic colitis and the hemolytic-uremic syndrome in humans and may cause serious morbidity and large outbreaks worldwide. People with bloody diarrhea have an increased risk of developing serious complications such as acute renal failure and neurological damage. The hemolytic-uremic syndrome (HUS) is a serious condition, and up to 50% of HUS patients can develop long-term renal dysfunction or blood pressure-related complications. Children aged two to six years have an increased risk of developing HUS. Clinical enteropathogenic Escherichia coli (EPEC) infections show fever, vomiting, and diarrhea. The EPEC reservoir is unknown but is suggested to be an asymptomatic or symptomatic child or an asymptomatic adult carrier. Spreading is often through the fecal-oral route. The prevalence of EPEC in infants is low, and EPEC is highly contagious in children. EPEC disease in children tends to be clinically more severe than other diarrheal infections. Some children experience persistent diarrhea that lasts for more than 14 days. Enterotoxigenic Escherichia coli (ETEC) strains are a compelling cause of the problem of diarrheal disease. ETEC strains are a global concern as the bacteria are the leading cause of acute watery diarrhea in children and the leading cause of traveler's diarrhea. It is contagious to children and can cause chronic diarrhea that can affect the development and well-being of children. Infections with diarrheagenic E. coli are more common in African countries. Antimicrobial agents should be avoided in the acute phase of the disease since studies showed that antimicrobial agents may increase the risk of HUS in children. The South African National Veterinary Surveillance and Monitoring Programme for Resistance to Antimicrobial Drugs has reported increased antimicrobial resistance in E. coli. Pathogenic bacterial strains have developed resistance to a variety of antimicrobial agents due to antimicrobial misuse. The induced heavy metal tolerance may also enhance antimicrobial resistance. The prevalence of antimicrobial resistance depends on the type of the antimicrobial agent, bacterial strain, dose, time, and mode of administration. Developing countries are severely affected by increased resistance to antimicrobial agents due to poverty, lack of proper hygiene, and clean water, which can lead to bacterial infections with limited treatment options due to resistance.
10 citations
TL;DR: A PCR method was developed to detect Salmonella Heidelberg in pure culture and chicken feces based on specific primers targeting genes conferring fosfomycin (fosA7) and third-generation cephalosporin (blaCMY-2) resistance as well as theSalmonella-specific invA gene and the universal 16S rRNA genes.
10 citations
Cites background from "In Silico Detection and Typing of P..."
...0 database (14) to determine their replicon types....
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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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