Functional characterisation of metabolism-related genes required for the survival of Mycoplasma bovis in association with host cells

Metabolic functions of mycoplasmas play an important role in their interactions with their host. Recent studies examining the capacity of mutants of Mycoplasma bovis to survive in co-culture with Madin-Darby bovine kidney (MDBK) cells have identified three genes, MBOVPG45_0728, MBOVPG45_0028 and MBOVPG45_0327, that are essential for the survival of M. bovis in association with host cells. In the study described here, the metabolic profiles of the mutant strains with transposons inserted into these genes (∆MBOVPG45_0728, ∆MBOVPG45_0028 and ∆MBOVPG45_0327) were compared with that of the parent strain, PG45, to investigate the metabolic functions of the proteins. Steady-state metabolomic analysis did not identify significant alterations in metabolites associated with glucose metabolism in the ∆MBOVPG45_0728 mutant, suggesting that the product of MBOVPG45_0728 does not have the role in phosphoglucomutase activity predicted by bioinformatic analysis, and may play a role in amino acid and lipid metabolism. Metabolomic footprinting analyses detected significant differences in changes in medium components in cultures of ∆MBOVPG45_0028 and ∆MBOVPG45_0327 compared to PG45. These results were consistent with bioinformatic predictions that these two genes encoded components of transporter systems, and suggested that the protein encoded by the MBOVPG45_0028 gene is involved in nucleoside uptake, and that encoded by the MBOVPG45_0327 gene is involved in the efflux of organic acids. Overall, comparative metabolomic profiling revealed metabolic functions of M. bovis that are critical in interactions with host cells, furthering our understanding of metabolic mechanisms required for successful infection with M. bovis.