Molecular characterisation of theChlamydia pecorumplasmid from porcine, ovine, bovine, and koala strains indicates plasmid-strain co-evolution
Background.Highly stable, evolutionarily conserved, small, non-integrative plasmids are commonly found in members of theChlamydiaceaeand, in some species, these plasmids have been strongly linked to virulence. To date, evidence for such a plasmid inChlamydia pecorumhas been ambiguous. In a recent comparative genomic study of porcine, ovine, bovine, and koalaC. pecorumisolates, we identified plasmids (pCpec) in a pig and three koala strains, respectively. Screening of further porcine, ovine, bovine, and koalaC. pecorumisolates for pCpec showed that pCpecis common, but not ubiquitous inC. pecorumfrom all of the infected hosts.Methods.We used a combination of (i) bioinformatic mining of previously sequencedC. pecorumgenome data sets and (ii) pCpec PCR-amplicon sequencing to characterise a further 17 novel pCpecs inC. pecorumisolates obtained from livestock, including pigs, sheep, and cattle, as well as those from koala.Results and Discussion.This analysis revealed that pCpec is conserved with all eight coding domain sequences (CDSs) present in isolates from each of the hosts studied. Sequence alignments revealed that the 21 pCpecs show 99% nucleotide sequence identity, with 83 single nucleotide polymorphisms (SNPs) shown to differentiate all of the plasmids analysed in this study. SNPs were found to be mostly synonymous and were distributed evenly across all eight pCpecCDSs as well as in the intergenic regions. Although conserved, analyses of the 21 pCpecsequences resolved plasmids into 12 distinct genotypes, with five shared between pCpecs from different isolates, and the remaining seven genotypes being unique to a single pCpec. Phylogenetic analysis revealed congruency and co-evolution of pCpecs with their cognate chromosome, further supporting polyphyletic origin of the koalaC. pecorum. This study provides further understanding of the complex epidemiology of this pathogen in livestock and koala hosts and paves the way for studies to evaluate the function of this putativeC. pecorumvirulence factor.