Pangenomic and biochemical analyses of Helcococcus ovis reveal widespread tetracycline resistance and a novel bacterial species, Helcococcus bovis

<i>Helcococcus ovis</i> (<i>H. ovis</i>) is an opportunistic bacterial pathogen of a wide range of animal hosts including domestic ruminants, swine, avians, and humans. In this study, we sequenced the genomes of 35 <i>Helcococcus</i> sp. clinical isolates from the uterus of dairy cows and explored their antimicrobial resistance and biochemical phenotypes <i>in vitro</i>. Phylogenetic and average nucleotide identity analyses classified four <i>Helcococcus</i> isolates within a cryptic clade representing an undescribed species, for which we propose the name <i>Helcococcus bovis</i> sp. nov. By establishing this new species clade, we also resolve the longstanding question of the classification of the Tongji strain responsible for a confirmed human conjunctival infection. This strain did not neatly fit into <i>H. ovis</i> and is instead a member of <i>H. bovis</i>. We applied whole genome comparative analyses to explore the pangenome, resistome, virulome, and taxonomic diversity of the remaining 31 <i>H. ovis</i> isolates. An overwhelming 97% of <i>H. ovis</i> strains (30 out of 31) harbor mobile tetracycline resistance genes and displayed significantly increased minimum inhibitory concentrations of tetracyclines <i>in vitro</i>. The high prevalence of mobile tetracycline resistance genes makes <i>H. ovis</i> a significant antimicrobial resistance gene reservoir in our food chain. Finally, the phylogenetic distribution of co-occurring high-virulence determinant genes of <i>H. ovis</i> across unlinked and distant loci highlights an instance of convergent gene loss in the species. In summary, this study showed that mobile genetic element-mediated tetracycline resistance is widespread in <i>H. ovis</i>, and that there is evidence of co-occurring virulence factors across clades suggesting convergent gene loss in the species. Finally, we introduced a novel <i>Helcococcus</i> species closely related to <i>H. ovis</i>, called <i>H. bovis</i> sp. nov., which has been reported to cause infection in humans.