In the case of M. pneumoniae, it is the STK, but not STP (PrpC), mutant which failed to adhere with culture flasks [20, 42]. Consistent with this negative adherence to culture flasks, this STK Erismodegib manufacturer mutant strain (MPN248 mutant) exhibits reduced levels of adherence related proteins, including P1, in SDS-PAGE. However, recent studies have demonstrated that deletion of
STP in strains of S. pyogenes (M1SF370) [22] and S. pneumoniae (D39)[25] leads to reduced adherence to pharyngeal cells. It appears, therefore, that disruption of both STK and STP can lead to adherence negative phenotype but it varies from species to species. However, the mechanism behind partial adherence of TIM207 to cultures flask remains elusive and it requires further study. TIM207 strain is less cytotoxic to HeLa cells Further to understand whether the lack of MG207 has any effect on other pathogenic mechanisms of M. genitalium, we examined the ability of TIM207 strain to cause cytotoxicity. Therefore, we infected HeLa cells with TIM207 and other control strains. Figure 5 shows the confocal microscopy observation of HeLa cells infected with M. genitalium strains. As can be seen, M. genitalium wild type strain G37 and a control strain TIM262, which hasTn4001 insertion in MG_262 encoding 3´-5´ exonuclease, had severe cytotoxic effects on HeLa cells, while TIM207 had no such effect and behaved similar to that of heat killed G37 (HKG37) strain. Since cytotoxicity of mycoplasmas is due partly to
the release of hydrogen peroxide by these CP-690550 cost species, we speculated that differences in cytotoxicity between the wild type and the mutant strains might be due to differences in the production of H2O2 by these strains. To rule out this possibility, we determined the H2O2 levels in these strains by FOX assay. The results
showed significantly reduced levels of H2O2 in TIM207 strain as compared to G37 strain (Figure 6). This indicated that deletion of MG_207 had some direct or indirect effect on the synthesis of H2O2 by M. genitalium. Mycoplasmas produce H2O2 by oxidizing the glycerophosphate of the glycolytic pathway by glycerophosphate oxidase [53]. It is likely that phosphorylation or dephosphorylation of some of the enzymes associated with this pathway leads to reduced production of H2O2 in TIM207 strain. Besides, in M. pneumoniae reduced cytotoxicity and H2O2 production is linked to reduced ability to utilize Reverse transcriptase glycerol [20]. To understand if the reduced H2O2 production by TIM207 has any correlation with glycerol utilization, we determined the growth of the TIM207 strain in SP-4 medium containing glycerol instead of dextrose. AZD1390 chemical structure Results presented in Additional file 3: Figure S2 reveal that this strain has a defect in the utilization of glycerol as compared to the wild type strain. These results, taken together, reiterate that reduced cytotoxicity of TIM207 is due partly to generation of relatively lower amount of H2O2 by this strain. Figure 5 Microscopic observation of cytotoxic effect by M.