Variation in the microbiome of the urogenital tract of Chlamydia-free female koalas (Phascolarctos cinereus) with and without 'wet bottom'.

Koalas (Phascolarctos cinereus) are iconic Australian marsupials currently threatened by several processes, including infectious diseases and ecological disruption. Infection with Chlamydia pecorum, is considered a key driver of population decline. The clinical sign of 'wet bottom', a staining of the rump associated with urinary incontinence, is often caused by chlamydial urinary tract infections. However, wet bottom has been recorded in koalas free of C. pecorum, suggesting other causative agents in those individuals. We used 16S rRNA diversity profiling to investigate the microbiome of the urogenital tract of ten female koalas in order to identify potential causative agents of wet bottom, other than C. pecorum. Five urogenital samples were processed from koalas presenting with wet bottom and five were clinically normal. All koalas were negative for C. pecorum infection. We detected thirteen phyla across the ten samples, with Firmicutes occurring at the highest relative abundance (77.6%). The order Lactobacillales, within the Firmicutes, comprised 70.3% of the reads from all samples. After normalising reads using DESeq2 and testing for significant differences (P < 0.05), there were 25 operational taxonomic units (OTUs) more commonly found in one group over the other. The families Aerococcaceae and Tissierellaceae both had four significantly differentially abundant OTUs. These four Tissierellaceae OTUs were all significantly more abundant in koalas with wet bottom. This study provides the foundation for future investigations of causes of koala wet bottom, other than C. pecorum infection. This is of clinical relevance as wet bottom is often assumed to be caused by C. pecorum and treated accordingly. Our research highlights that other organisms may be causing wet bottom, and these potential aetiological agents need to be further investigated to fully address the problems this species faces.