Use of Whole-Genus Genome Sequence Data To Develop a Multilocus Sequence Typing Tool That Accurately Identifies Yersinia Isolates to the Species and Subspecies Levels

Abstract

hThe genus Yersinia is a large and diverse bacterial genus consisting of human-pathogenic species, a fish-pathogenic species, and a large number of environmental species. Recently, the phylogenetic and population structure of the entire genus was elucidated through the genome sequence data of 241 strains encompassing every known species in the genus. Here we report the mining of this enormous data set to create a multilocus sequence typing-based scheme that can identify Yersinia strains to the species level to a level of resolution equal to that for whole-genome sequencing. Our assay is designed to be able to accurately subtype the important human-pathogenic species Yersinia enterocolitica to whole-genome resolution levels. We also report the validation of the scheme on 386 strains from reference laboratory collections across Europe. We propose that the scheme is an important molecular typing system to allow accurate and reproducible identification of Yersinia isolates to the species level, a process often inconsistent in nonspecialist laboratories. Additionally, our assay is the most phylogenetically informative typing scheme available for Y. enterocolitica. The Gram-negative Yersinia is one of the most important and well-studied bacterial genera, consisting of three human pathogens. Y. pestis is the causative agent of bubonic and pneumonic plague and is a recently diverged clone of Yersinia pseudotuberculosis (1), which alongside Y. enterocolitica is a zoonotic gastrointestinal pathogen (2). The remaining species are not associated with human disease and are considered to be environmental organisms, with the exception of the common fish pathogen Y. ruckeri (2) and the insecticidal species Y. entomophaga. Of the human-pathogenic species, Y. enterocolitica is the most common etiological agent of human disease, and in Germany and Scandinavia, the numbers of cases of human intestinal yersiniosis caused by Y. enterocolitica rival those caused by Salmonella (3). Y. enterocolitica is in itself a very diverse species that is classically subdivided into nonpathogenic, low-pathogenic, and high-pathogenic biotypes based on virulence in a mouse infection model (4). Biotype 1A isolates are considered nonpathogenic, which is concordant with a lack of the major Y. enterocolitica virulence factors such as pYV, invasin, YadA, and Ail (5), although there are numerous reports of biotype 1A human carriage (6, 7). Biotype 1B isolates are high pathogenic, which is concordant with carriage of the high-pathogenicity island, but isolation from human disease cases is very rare with the exception of notable outbreaks such as the recent emergence in Poland (8). Biotype 2 to 4 isolates are low pathogenic and are globally the most common causes of human gastrointestinal yersiniosis (4). Biotype 5 isolates are also considered low pathogenic but have only been isolated from wild hare populations and are very rare in nature (5).From a clinical perspective, the isolation and subsequent identification of Yersinia and in particular Y. enterocoli...

Multilocus sequence analysis for the taxonomic updating and identification of the genus Proteus and reclassification of Proteus genospecies 5 O’Hara et al. 2000, Proteus cibarius Hyun et al. 2016 as later heterotypic synonyms of Proteus terrae Behrendt et al. 2015

“…MLSA has been used for classi cation at the species level in numerous Enterobacteriaceae [14,15,[16][17][18][19][20][21]. MLSA has the advantage of being more convenient and more conducive to popularization in the primary research institution than the whole genome sequencing method.…”

“…Multilocus sequence analysis (MLSA) based on several housekeeping genes (HKGs) has previously been successfully employed to delineate boundaries between closely related bacterial species, subspecies and component strains [11][12][13]. Partial sequences of protein-encoding genes have proven useful for species identi cation and as phylogenetic markers in the family Enterobacteriaceae [14,15].…”

Yersinia canariaesp. nov., isolated from a human yersiniosis case

“…Whole genome sequencing data characterised this isolate as ST333, utilising the multi-locus sequence type (MLST) scheme developed by Hall et al . (8,10). A phylogenetic tree previously revealed that the isolate did not cluster with Y. enterocolitica instead being located on a distinct branch (8).…”

Prevalence and genomic characteristics of zoonotic gastro-intestinal pathogens and ESBL/pAmpC producing Enterobacteriaceae among Swedish corvid birds

“…Sequence data were uploaded to the European Nucleotide Archive (ebi.ac.uk/ena) and are publicly available under project accession number PRJEB34530. MLST for Yersinia [19] was performed using tools at EnteroBase [20], whereas Campylobacter and E. coli MLST was performed using the Center for genomic epidemiology (CGE) MLST tool [21]. New loci and sequence types found were submitted to the databases.…”