Detection of pathogenic leptospires in animals by PCR based on <i style="">lipL21</i> and <i style="">lipL32 </i>genes

Abstract

Efficacy of primers capable of amplifying conserved outer membrane protein (OMP) genes i.e.,<i style=""> lipL21</i> and<i style=""> lipL32 </i>of<i style=""> Leptospira </i>strains was tested for rapid and early diagnosis of the leptospirosis using a polymerase chain reaction (PCR). These OMP genes were found to be conserved in various leptospiral serovars viz., Canicola, Pomona, Icterohaemorrhagiae, Pyrogenes, Sejroe<i style="">,</i> Grippotyphosa, Ballum and Tarassovi as PCR products of 561 bp and 756 bp were obtained by PCR employing <i style="">lipL21 </i>and <i style="">lipL32</i> based primers, respectively, in all these serovars. Absence of such amplicons in DNA extracted from<i style=""> Pasteurella, Campylobacter </i>and <i style="">Brucella </i>confirmed the specificity of the primers. Serum and tissue samples collected from cattle, buffaloes and experimentally infected guinea pigs and calves were subjected to PCR using above primers as well as conventionally used primers G1/G2. All the sera and tissue samples, whether field samples or collected from experimentally infected animals, found positive for G1/G2 specific PCR were also positive for <i style="">lipL21</i> and <i style="">lipL32</i> specific PCR. The present study indicated that <i style="">lipL21 </i>and <i style="">lipL32</i> based primers could be used for PCR based diagnosis of leptospirosis. Since G1/G2 primers are known not to amplify the DNA of Grippotyphosa, the use of primers employed in the present study could have an additional advantage in detection of cases of the disease.