Many bacterial pathogens contain randomly-switching methyltransferases that control phase-variable regulons – phasevarions. All current examples control expression of genes involved in pathogenesis and host-adaptation, and many regulate putative and current vaccine candidates. Effective vaccines require stably expressed targets; individual phase-variable can be identified in silico as they contain easily recognised features, but genes controlled by phasevarions do not, complicating the rational design of vaccines. We have identified and begun to study phasevarions controlled by the switching of both Type I and Type III methyltransferases in several important bacterial veterinary pathogens: Streptococcus suis and Actinobacillus pleuropneumoniae are major swine pathogens, with S. suis also a major cause of bacterial meningitis in humans, particularly in S.E. Asia; Mannheimia haemolytica is a major bovine pathogen. All three organisms contain randomly switching methyltransferases, with all three pathogens containing both Type I and Type III methyltransferases that are able to phasevary – a phenomenon never before observed. Pacific Biosciences SMRT sequencing and methylome analysis of the methyltransferases from these organisms has deciphered their specificity, and demonstrated that different genes/alleles methylate different target sequences, and therefore control different phasevarions. Analysis of the protein profiles of strains containing phase-variable methyltransferases shows protein expression differences correlating with methyltransferase switching.

Our analysis shows that phasevarions are present in diverse veterinary pathogens, and need to be characterised in order to identify the stably expressed protein repertoire of these organisms. This work will provide a framework for the rational design of vaccines and treatments against these bacteria.