(P462) Extensive phase variation in Gardnerella: what could this mean for BV treatment and pathogenesis?

Background: The Gardnerella genus is associated with the polymicrobial disorder bacterial vaginosis (BV). However, the mechanistic details of Gardnerella pathogenesis remain poorly characterized, due in part to the substantial genetic and phenotypic heterogeneity within the genus. This heterogeneity is being parsed apart at the community and species levels, but clonal heterogeneity has yet to be explored in any great detail, though it could have important implications for BV treatment and pathogenesis. We recently discovered phase variable colony morphotypes characterized by size (large and small) and opacity (opaque and translucent) that are conserved among all tested Gardnerella strains, representing at least 10 different species. With the hypothesis that variants could play different roles in BV pathogenesis, we sought to characterize their differences.



Methods: We used in vitro assays, proteomics, and whole genome sequencing to assess phenotypic and genotypic differences between colony variants in three different Gardnerella species.



Results: Large colony variants (Lg) showed reduced vaginolysin secretion, faster growth rate, and enhanced biofilm production relative to small colony variants (Sm). The ability to inhibit growth of Neisseria gonorrhoeae and commensal lactobacillus species varied by strain and in some instances differed between variants. Proteomics analyses showed differential expression of 127-173 proteins, demonstrating that variants have distinct metabolic profiles. Proteins increased in Lg variants were predominantly involved in amino acid and protein synthesis, while those increased in Sm variants were related to nucleotide synthesis and DNA recombination. Whole genome sequencing analyses revealed an abundance of genes associated with variable homopolymeric tracts, implicating slipped strand mispairing in Gardnerella phase variation. The median number of poly-G/C tracts across Gardnerella genomes (n= 88) was 20. In our strains of interest, ~30% of tracts were intragenic, and the remainder were found within putative promoter regions. Genomic mutations detected in multiple strains between colony variants are currently being investigated for their contribution to colony phenotype.



Conclusion: Our results suggest that colony variants may be better suited to survive under different environmental conditions and illuminate previously unrecognized variability within clonal populations of multiple Gardnerella spp.