Reverse transcriptase polymerase chain reaction (RT‐PCR) detection of HLP gene expression in Candida glabrata and its possible role in in vitro haemolysin production

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Abstract
Although haemolysins are known to be putative virulence factors contributing to pathogenicity in Candida species, the haemolytic activity of Candida glabrata and its genetic expression is ill understood at present. Thus, we studied a total of 34 Candida glabrata isolates for their in vitro haemolytic activity using a previously described plate assay system. The mRNA expression of HLP, a putative haemolysin gene, of these isolates was also evaluated using a semi-quantitative, non-competitive RT-PCR assay. All 34 C. glabrata isolates exhibited both partial (alpha) and complete (beta) haemolytic activity to varying degrees. In parallel with the haemolytic activity, all isolates were also positive for HLP mRNA expression. The expression levels of HLP mRNA (as relative units) ranged from 1.01 to 1.82, with a mean value of 1.32. On regression analysis of latter values and the haemolytic activity (in terms of the dimension of the haemolytic zone in the plate assay) of the C. glabrata isolates a highly significant positive correlation was noted (r=0.759, p<0.0001). Taken together, our data illustrate not only the phenotypic characteristics of haemolysin(s) and HLP expression of a battery of C. glabrata clinical isolates, but also, for the first time, evidence for a role of HLP in haemolysis.