Essential role of proline synthesis and the one-carbon metabolism pathways for systemic virulence of Streptococcus pneumoniae

Previous virulence screens have indicated potential roles during Streptococcus pneumoniae infection for the one-carbon metabolism pathway component Fhs and proline synthesis mediated by ProABC. To define how these metabolic pathways affect S. pneumoniae virulence we have investigated phenotypes and transcription profiles of {Delta}fhs and {Delta}proABC strain mutants. S. pneumoniae capsular serotype 6B BHN418 {Delta}fhs and {Delta}proABC mutant strains were markedly reduced virulence in mouse models of systemic infection and pneumonia, but were still able to colonise the nasopharynx. Although the {Delta}fhs and {Delta}proABC mutant strains grew normally in complete media, both mutant strains had markedly impaired growth in chemically defined medium, human serum and human CSF. The {Delta}proABC strain also had impaired growth under conditions of osmotic and oxidative stress. When transferred to the serotype 2 D39 S. pneumoniae strain background, the {Delta}fhs mutation replicated the virulence and growth in serum phenotype of the BHN418 mutation. In contrast, the D39 {Delta}proABC mutant could cause septicaemia and grow in human serum, indicating the role of this genetic locus during virulence is strain-specific. In human sera the {Delta}fhs and {Delta}proABC mutants both had major derangements in global gene transcription affecting multiple but different metabolic pathways, identifying the corresponding S. pneumoniae metabolic functions affected by these genes under infection-related conditions. Our data demonstrate an essential role for the S. pneumoniae one- carbon metabolism and a strain-conditional role for proline biosynthesis for growth in physiological fluids and therefore systemic infection, and further demonstrate the vital importance of bacterial metabolism for disease pathogenesis. ImportanceRapid adaptation to grow within the physiological conditions found in the host environment is an essential but poorly understood virulence requirement for systemic pathogens such as Streptococcus pneumoniae. We have now demonstrated an essential role for the one-carbon metabolism pathway and a conditional role depending on strain background for proline biosynthesis for S. pneumoniae growth in serum or CSF and therefore for systemic virulence. RNAseq data demonstrated that loss of one carbon metabolism or proline biosynthesis both have profound but differing effects on S. pneumoniae metabolism in human serum, identifying the metabolic processes dependent on each pathway during systemic infection. These data provide a more detailed understanding of the adaptations required by systemic bacterial pathogens in order to cause infection, and demonstrate that the requirement for some of these adaptations vary between strains from the same species and could therefore underpin strain variations in virulence potential.