-Glycosylation in Piroplasmids: Diversity within Simplicity.

-glycosylation has remained mostly unexplored in Piroplasmida, an order of tick-transmitted pathogens of veterinary and medical relevance. Analysis of 11 piroplasmid genomes revealed three distinct scenarios regarding -glycosylation: sensu stricto (s.s.) species add one or two -acetylglucosamine (NAcGlc) molecules to proteins; and add (NAcGlc)-mannose, while and s.s. synthesize dolichol-P-P-NAcGlc and dolichol-P-P-(NAcGlc) without subsequent transfer to proteins. All piroplasmids possess the gene complement needed for the synthesis of the -glycosylation substrates, dolichol-P and sugar nucleotides. The oligosaccharyl transferase of species, and , is predicted to be composed of only two subunits, STT3 and Ost1. Occurrence of short -glycans in merozoites was experimentally demonstrated by fluorescence microscopy using a NAcGlc-specific lectin. In vitro growth of was significantly impaired by tunicamycin, an inhibitor of -glycosylation, indicating a relevant role for -glycosylation in this pathogen. Finally, genes coding for -glycosylation enzymes and substrate biosynthesis are transcribed in blood and tick stages, suggesting that this pathway is biologically relevant throughout the parasite life cycle. Elucidation of the role/s exerted by -glycans will increase our understanding of these successful parasites, for which improved control measures are needed.