Poster Presentation Australian Society for Microbiology Annual Scientific Meeting 2024

Deciphering the pan-methylome of Group A Streptococcus (#60)

August Mikucki 1 2 , Tim C Barnett 1 2 , Janessa Pickering 2 3
  1. The Marshall Centre for Infectious Diseases Research and Training, The University of Western Australia, Perth, WA, Australia
  2. Telethonk Kids Institute, Nedlands, WA, Australia
  3. School of Biomedical Sciences, University of Western Australia, Perth, WA, Australia

Group A Streptococcus (GAS) is a globally significant cause of superficial skin and throat infections and severe life-threatening illnesses. Research in other pathogenic bacteria has demonstrated that epigenetic control of gene expression through DNA methylation is an important regulator of growth, immune evasion, antibiotic resistance, and virulence. GAS has one major methylation system (HsdRSM) whose recognition motif differs by strain. However, very little is known about the role of DNA methylation in GAS. We sought to comprehensively define the putative methylome of GAS at a species-wide level. A GAS pangenome was constructed using 276 publicly available genomes using the Pangenome module of the Bactopia analysis pipeline. The methylation motif of the HsdRSM system was determined by BLAST comparison to enzymes in the REBASE database with previously confirmed motifs. For each genome, the corresponding motif was annotated to give the putative methylome of the isolate. Functional analysis was performed using the ClusterProfiler R package on gene clusters annotated with clusters of orthologous group (COG) functions using EggNOG mapper. The hsdRSM locus was intact in 97.8% (270/276) of isolates, with twelve unique methylation motifs identified. The mean number of methylation motifs per genome was 388.7 (range: 179-574). Principle component analysis of the methylated genes in each sample revealed four distinct clusters. Three were single-motif clusters but contained multiple unrelated genomic lineages, implying conserved methylation patterns. Despite all methylation motifs being present in more than one lineage, between 161 and 327 conserved methylation sites were present in all strains sharing each motif. The core methylome of three motifs was significantly enriched for genes encoding carbohydrate metabolism, nucleotide metabolism, energy production, and inorganic ion transport (COG categories G, F, C, and P adjusted p value <0.05). Despite diversity in the patterns of methylation sites between GAS strains, we have demonstrated conservation of methylation sites across genetically distant lineages. This work will form the basis of a larger study using both in silico and in vitro methods to establish the biological role of DNA methylation in GAS and determine the impact of DNA methylation on virulence, antibiotic resistance, and survival in this important pathogen.