Leptospiral phosphatidylinositol-specific phospholipase C: a novel conserved antigen for the definitive diagnosis of Leptospirosis.

Identification and characterisation of novel antigenic proteins of Leptospira are necessary to unravel the secrets of the Leptospiral disease biology. Besides, the current scenario of Leptospirosis demands the development of advanced diagnostics with the potential of an early and specific diagnosis. This work reports the diagnostic efficacy of a PI-PLC (LA_1375) of Leptospira interrogans serovar Lai strain 56601. Phospholipases are enzymes that catalyse the degradation of membrane phospholipids, with the after effects of pathophysiological consequences through the impairment of cellular signalling mechanisms. The immunoblotting analysis allows to identify the Leptospirosis positive serum samples demonstrated the expression and exposure of the protein to the immune system and confirmed the antigenic nature of the protein. Furthermore, evaluation of diagnostic efficacy of an ELISA to detect antibodies against recombinant PI-PLC has tested in indirect IgM ELISA using the purified 50 kDa recombinant protein as the coating antigen. A cut-off value of 0.218 at mean + 2SD and a sensitivity value of 100% and a specificity value of 95.65% explored for the rPI-PLC IgM ELISA. The PPV and NPV of ELISA for the Leptospirosis confirmed acute phase samples were 95.87% and 100% respectively. Comparison of the ELISA assay with MAT for the detection of Leptospiral antibodies showed a positive correlation (r value 0.345, p < 0.05), and signified the diagnostic efficacy of rPI-PLC for the definitive diagnosis of the Leptospirosis. In addition, the revealed conservation nature of PI-PLC among different Leptospira species highlights the diagnostic and vaccine candidate potential. Thus, the study emphasises the Leptospiral PI-PLC as a potential antigen still the importance as a novel biomarker candidate for the serodiagnosis and pathogenesis of Leptospirosis have to be investigated.

Phosphatidylinositol-specific phospholipase C contributes to survival of Staphylococcus aureus USA300 in human blood and neutrophils.

Staphylococcus aureus is an important human pathogen that employs a large repertoire of secreted virulence factors to promote disease pathogenesis. Many strains of S. aureus possess a plc gene that encodes a phosphatidylinositol (PI)-specific phospholipase C (PI-PLC) capable of hydrolyzing PI and cleaving glycosyl-PI (GPI)-linked proteins from cell surfaces. Despite being secreted by virulent staphylococci, the contribution of PI-PLC to the capacity of S. aureus to cause disease remains undefined. Our goal in these studies was to understand PI-PLC in the context of S. aureus biology. Among a collection of genetically diverse clinical isolates of S. aureus, community-associated methicillin-resistant S. aureus (CA-MRSA) USA300 secreted the most PI-PLC. Screening a collection of two-component system (TCS) mutants of S. aureus, we identified both the agr quorum-sensing system and the SrrAB TCS to be positive regulators of plc gene expression. Real-time PCR and PI-PLC enzyme assays of the TCS mutants, coupled with SrrA promoter binding studies, demonstrated that SrrAB was the predominant transcriptional activator of plc. Furthermore, plc regulation was linked to oxidative stress both in vitro and in vivo in a SrrAB-dependent manner. A Δplc mutant in a CA-MRSA USA300 background exhibited a survival defect in human whole blood and in isolated neutrophils. However, the same mutant strain displayed no survival defect in murine models of infection or murine whole blood. Overall, these data identify potential links between bacterial responses to the host innate immune system and to oxidative stress and suggest how PI-PLC could contribute to the pathogenesis of S. aureus infections.