Molecular detection and characterisation of potentially zoonotic bacteria in bathyergids from the Western Cape Province

Abstract

Globally emphasis has been given to identify emerging and re-emerging pathogens. Rapid urban expansion creates a problem which is two-fold. Firstly, increasing slum living conditions due to inadequate rate of infrastructure development results in an increased reliance on natural resources, including the capture and consumption of surrounding wildlife to subsist, thereby facilitating the transfer of emergent zoonotic pathogens. Secondly, through activities such as pollution or alien species introductions, the rapid transformation of once pristine environments, alters natural systems, potentially exposing these environments to new bacterial pathogens. Therefore, the main aim of this study was to assess overlooked bacterial species harboured by four host species (Bathyergus suillus, Georychus capensis, Cryptomys hottentotus hottentotus and Fukomys damarensis) belonging to the subterranean rodent family Bathyergidae, which inhabit an environment well-suited for an array of bacterial species, and which varied in their exposure to human settlements. Bacterial prevalence and diversity was initially evaluated using broad-range PCR techniques in combination with nucleotide sequencing. This revealed high levels of bacterial prevalence (82.91%) and mixed infections (22.60%) in bathyergid species. Two bacterial groups, the Bacillus cereus complex (a group of soil-dwelling bacterial strains with pathogenic potential with an overall prevalence of 8.55%) and haemotropic Mycoplasma strains (vector-borne bacterial strains of zoonotic potential with an overall prevalence of 1.28%) were subsequently selected for further genetic analysis with genus and species-specific PCRs. Bacillus molecular screening and phylogenetic analyses was achieved by targeting four gene regions with seven published primer assays and two novel PCR assays. This enabled identification of two B. cereus complex strains in bathyergid lungs and revealed an overall B. cereus complex prevalence of 17.95% for the 234 bathyergid lung samples screened. Bacillus genome prevalence was significantly higher in B. suillus individuals (45.35%), sampled in a peri-urban environment, compared to the other bathyergid species sampled from pristine habitats (ranging from 0% - 4.44%). Anthropogenic activities in the area where B. suillus was sampled could, at least partially, attribute to the perceived difference between urban and naturally sampled bathyergid species, highlighting the role of B. suillus to act as both a reservoir of potentially zoonotic pathogens and as a sentinel for anthropogenic soiling. Mycoplasma molecular screening using three different PCR assays, all targeting the 16S rRNA gene region, confirmed an overall haemotropic Mycoplasma prevalence of 24.13% in the 286 bathyergid organs (lung, spleen and liver) screened. A significantly higher prevalence and diversity of haemotropic Mycoplasma strains was found in B. suillus lungs (41.86%) compared to its naturally occurring relatives (ranging from 0%-36%). Phylogenetic analyses identified six novel haemotropic Mycoplasma strains, all grouping within a discrete monophyletic cluster, sister to Mycoplasma coccoides, and comprising two well-supported sub-clusters. The human introduction of commensal rodents harbouring Mycoplasma strains transferred through cosmopolitan arthropod vectors to indigenous bathyergids, likely underlies the higher prevalence in urban areas, although other biotic and abiotic factors affecting ectoparasite load also merit consideration. The data generated by the current study indicate the need to identify largely overlooked and potentially zoonotic bacterial pathogens in subterranean mammals and emphasises the importance of monitoring anthropogenically-introduced, opportunistic pathogens and the threats they pose to vulnerable communities and co-occurring, free-living animal species