Application of whole genome sequencing to Australia-wide collections of extraintestinal pathogenic Escherichia coli from companion animals and commensal E. coli from pigs

Abstract

Escherichia coli is one of the most important bacterial pathogens associated with extraintestinal infections in both humans and animals. In companion animals, it is associated with urinary, respiratory, gastrointestinal tract, joint, wound, skin and soft tissue infections. These infections vary significantly between cats and dogs: for example, extraintestinal infection in cats is infrequent, and they rarely have simple, uncomplicated bacterial UTIs compared to dogs. E. coli strains causing UTIs in both cats and dogs share pathotypic similarities and are phylogenetically related to ExPEC isolates from humans. It is, therefore, possible for humans to share pathogenic ExPEC strains with their companion animals, which suggests the possibility of both zoonotic (animal-to-human) and anthropozoonotic (human-to-animal) transmission. Among phylogenetic group B2 FQ-susceptible cat clinical isolates from an Australia-wide collection (n = 323), it was found that ST73 was the dominant B2-associated ST, based on both WGS (23/53, 43%) and ST-specific PCR (45/221, 20%). Less dominant STs identified by WGS included ST127, ST12, and ST372 (4/53, 8% each). By contrast, among group B2 FQ-susceptible dog clinical isolates from an Australia-wide collection (n = 449), ST372, an infrequent human pathogen, was found to be the predominant B2-associated ST according to both WGS (24/77, 31%) and ST-specific PCR (53/240, 22%). The other primary STs identified by WGS included ST73 (13/77, 17%), ST12 and ST80 (5/77, 7% each). To further compare the relatedness of ST73 isolates from dogs, cats, and humans, WGS-based phylogenetic comparison was performed. This comparison showed that there was considerable overall phylogenetic diversity among the isolates. Although most clusters were species-specific, some contained closely related human and animal (dog>cat) isolates. These results confirmed that while there is clonal commonality among ExPEC isolated from cats, dogs and humans, there are dominant host-specific clonal groups – ST372 and ST73 in dogs and ST73 in cats – and potential for bi-directional (i.e., companion animal-human and human-companion animal) transmission of ST73 strains between host species. A collection of FQR E. coli isolates from the same Australia-wide collection of isolates from extraintestinal infections in Australian cats and dogs (n = 59) was also investigated to determine the prevalence of the globally disseminated MDR pandemic clonal groups ST131 and ST1193. The main STs identified in this study were ST224 (10/59, 17%), ST744 (8/59, 14%), and ST38 (8/59, 14%). The overall prevalence of ST131 was (6/59, 10%), however, no ST1193 isolates were identified in contrast to a 5-year-earlier report which identified two dog-source ST1193 isolates closely related to human source isolates. Furthermore, the findings showed that the prevalence of ST131 as a cause of FQR infections in Australian companion animals was relatively constant between this study and the 5-year-earlier study of Platell et al. (2010) (9/125 isolates, 7.2%). To investigate the clonal commonality of human and companion animal ST131, 20 Australian cat- and dog-source ST131 genomes were compared to 173 reference human-source ST131 genomes. This analysis revealed that the animal-sourced isolates were widely distributed throughout the ST131 phylogeny. There was also some minor sub-clustering of dog- and cat-source isolates, though most were closely related to human-source ST131 isolates. The absence of a separate clade containing only companion animal ST131 isolates combined with the higher prevalence of ST131 and ST1193 in humans reported in the literature suggests that companion animals act as spillover hosts rather than primary reservoirs for these lineages. The high degree of clonal commonality among these FQR clinical isolates from humans and companion animals further suggests the possibility of bi-directional between-species transmission. In addition to being both a commensal and pathogen in companion animals, E. coli is also the main facultative anaerobic bacteria colonising the pig gastrointestinal tract. While being a beneficial gut commensal, some E. coli pathotypes cause both intestinal and extraintestinal infections in pigs with AMR of increasing concern in both commensals and pathogens. On this basis the frequency of antimicrobial non-susceptibility among E. coli (n = 201) and Salmonella spp. (n = 69) isolated from rectal contents of healthy Australian finisher pigs at slaughter was determined. Only low levels of non-susceptibility to CIAs (ESCs and FQs) were identified, with non-susceptibility to both cefoxitin and ciprofloxacin observed in only 1% (2/201) of E. coli isolates. In addition, all E. coli isolates were fully susceptible to ceftiofur. None of the Salmonella spp. isolates showed any non-susceptibility to these CIAs. As the use of fluoroquinolones in food-producing animals in Australia is banned, the identification of fluoroquinolone resistance in E. coli from Australian pigs is significant as it demonstrates that even in the absence of local antimicrobial selection pressure resistance can occur. FQR E. coli can enter livestock production facilities despite strict biosecurity possibly via human carriers or wild birds. Other potential sources include feed, water, rodents, and insects. The results from these studies highlight the need for further surveillance of AMR and identification of prominent resistant clonal lineages in both companion and food-producing animals in Australia.