In Silico Detection and Typing of Plasmids using PlasmidFinder and Plasmid Multilocus Sequence Typing

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.

[1]  A. Carattoli,et al.  Reversion to susceptibility of a carbapenem-resistant clinical isolate of Klebsiella pneumoniae producing KPC-3. , 2013, The Journal of antimicrobial chemotherapy.

[2]  R. Kaas,et al.  Genotyping using whole-genome sequencing is a realistic alternative to surveillance based on phenotypic antimicrobial susceptibility testing. , 2013, The Journal of antimicrobial chemotherapy.

[3]  H. Hasman,et al.  Characterization of IncN plasmids carrying bla CTX-M-1 and qnr genes in Escherichia coli and Salmonella from animals, the environment and humans. , 2013, The Journal of antimicrobial chemotherapy.

[4]  P. Nordmann,et al.  Complete Sequence of the IncT-Type Plasmid pT-OXA-181 Carrying the blaOXA-181 Carbapenemase Gene from Citrobacter freundii , 2013, Antimicrobial Agents and Chemotherapy.

[5]  F. de la Cruz,et al.  A Degenerate Primer MOB Typing (DPMT) Method to Classify Gamma-Proteobacterial Plasmids in Clinical and Environmental Settings , 2012, PloS one.

[6]  S. Rasmussen,et al.  Identification of acquired antimicrobial resistance genes , 2012, The Journal of antimicrobial chemotherapy.

[7]  P. Nordmann,et al.  Complete sequencing of an IncH plasmid carrying the blaNDM-1, blaCTX-M-15 and qnrB1 genes. , 2012, The Journal of antimicrobial chemotherapy.

[8]  H. Hasman,et al.  Expansion of the IncX plasmid family for improved identification and typing of novel plasmids in drug-resistant Enterobacteriaceae. , 2012, Plasmid.

[9]  P. Nordmann,et al.  Characterization of an IncFII Plasmid Encoding NDM-1 from Escherichia coli ST131 , 2012, PloS one.

[10]  A. Carattoli,et al.  Klebsiella pneumoniae ST258 Producing KPC-3 Identified in Italy Carries Novel Plasmids and OmpK36/OmpK35 Porin Variants , 2012, Antimicrobial Agents and Chemotherapy.

[11]  Ole Lund,et al.  Multilocus Sequence Typing of Total-Genome-Sequenced Bacteria , 2012, Journal of Clinical Microbiology.

[12]  P. Nordmann,et al.  Evolution of IncA/C blaCMY-2-Carrying Plasmids by Acquisition of the blaNDM-1 Carbapenemase Gene , 2011, Antimicrobial Agents and Chemotherapy.

[13]  P. Nordmann,et al.  Analysis of the Resistome of a Multidrug-Resistant NDM-1-Producing Escherichia coli Strain by High-Throughput Genome Sequencing , 2011, Antimicrobial Agents and Chemotherapy.

[14]  V. Miriagou,et al.  Multilocus sequence typing of IncN plasmids. , 2011, The Journal of antimicrobial chemotherapy.

[15]  N. Thomson,et al.  Complete Sequence and Molecular Epidemiology of IncK Epidemic Plasmid Encoding blaCTX-M-14 , 2011, Emerging infectious diseases.

[16]  Martin C. J. Maiden,et al.  BIGSdb: Scalable analysis of bacterial genome variation at the population level , 2010, BMC Bioinformatics.

[17]  Alessandra Carattoli,et al.  Replicon sequence typing of IncF plasmids carrying virulence and resistance determinants. , 2010, The Journal of antimicrobial chemotherapy.

[18]  Narmada Thanki,et al.  CDD: a Conserved Domain Database for the functional annotation of proteins , 2010, Nucleic Acids Res..

[19]  N. Woodford,et al.  Complete nucleotide sequence of the IncN plasmid pKOX105 encoding VIM-1, QnrS1 and SHV-12 proteins in Enterobacteriaceae from Bolzano, Italy compared with IncN plasmids encoding KPC enzymes in the USA. , 2010, The Journal of antimicrobial chemotherapy.

[20]  A. Carattoli,et al.  Plasmid double locus sequence typing for IncHI2 plasmids, a subtyping scheme for the characterization of IncHI2 plasmids carrying extended-spectrum beta-lactamase and quinolone resistance genes. , 2010, The Journal of antimicrobial chemotherapy.

[21]  N. Woodford,et al.  Complete Nucleotide Sequences of Plasmids pEK204, pEK499, and pEK516, Encoding CTX-M Enzymes in Three Major Escherichia coli Lineages from the United Kingdom, All Belonging to the International O25:H4-ST131 Clone , 2009, Antimicrobial Agents and Chemotherapy.

[22]  A. Carattoli,et al.  Characterization of plasmids harbouring qnrS1, qnrB2 and qnrB19 genes in Salmonella. , 2009, The Journal of antimicrobial chemotherapy.

[23]  S. Nair,et al.  Variation in Salmonella enterica Serovar Typhi IncHI1 Plasmids during the Global Spread of Resistant Typhoid Fever , 2008, Antimicrobial Agents and Chemotherapy.

[24]  A. Carattoli,et al.  Multilocus sequence typing of IncI1 plasmids carrying extended-spectrum beta-lactamases in Escherichia coli and Salmonella of human and animal origin. , 2008, The Journal of antimicrobial chemotherapy.

[25]  V. Miriagou,et al.  Antimicrobial resistance islands: resistance gene clusters in Salmonella chromosome and plasmids. , 2006, Microbes and infection.

[26]  A. Carattoli,et al.  Identification of plasmids by PCR-based replicon typing. , 2005, Journal of microbiological methods.

[27]  M. Gilmour,et al.  The complete nucleotide sequence of the resistance plasmid R478: defining the backbone components of incompatibility group H conjugative plasmids through comparative genomics. , 2004, Plasmid.

[28]  S. Sørensen,et al.  Plasmid-Encoded Multidrug Efflux Pump Conferring Resistance to Olaquindox in Escherichia coli , 2004, Antimicrobial Agents and Chemotherapy.

[29]  V. Burland,et al.  The complete DNA sequence and analysis of R27, a large IncHI plasmid from Salmonella typhi that is temperature sensitive for transfer. , 2000, Nucleic acids research.

[30]  M. Couturier,et al.  Identification and classification of bacterial plasmids. , 1988, Microbiological reviews.

[31]  R. Stephan,et al.  Epidemiology of Virulence-Associated Plasmids and Outer Membrane Protein Patterns Within Seven Common Salmonella Serotypes , 1986, Infection and immunity.

[32]  N. Datta,et al.  Compatibility Groups among fi− R Factors , 1971, Nature.

[33]  P. Nordmann,et al.  Complete sequencing of an IncHI1 plasmid encoding the carbapenemase NDM-1, the ArmA 16S RNA methylase and a resistance-nodulation-cell division/multidrug efflux pump. , 2013, The Journal of antimicrobial chemotherapy.

[34]  D. Bartosik,et al.  Mobilizable narrow host range plasmids as natural suicide vectors enabling horizontal gene transfer among distantly related bacterial species. , 2012, FEMS microbiology letters.