Polymerase chain reaction (PCR)
The PCR is an in vitro method for the enzymatic synthesis of specific DNA sequences, using two oligonucleotide primers that hybridise to opposite strands and flank the region of interest in the target DNA (Erlich, 1989). It has been used to diagnose infectious diseases caused by fastidious bacteria (Bej et al., 1991; Merien et al., 1992). It is a rapid, reliable and sensitive test for the diagnosis of leptospirosis (Van Eys et al., 1989; Woodward et al., 1991). The application of the PCR for the diagnosis of leptospirosis has been reported by different workers.
In 1989, Van Eys et al. developed a PCR assay that could detect less than 10 L. hardjobovis added into the urine samples. Woodward et al. (1991) also developed a PCR which was specific for L. hardjobovis. Gerritsen et al. (1991) described a protocol to prepare a bovine urine sample for PCR assay, which improved the sensitivity of the assay.
Merien et al. (1992) described a PCR assay which could detect all pathogenic and non-pathogenic leptospires in clinical samples. Gravekamp et al. (1993) also developed a PCR assay using two sets of primers derived from genomic DNA libraries of L. icterohaemorrhagiae and bim which could detect all pathogenic leptospires in serum samples.
In 1994, Woodward and Redstone developed a PCR to amplify the flaB gene of 23 serovars of the genus Leptospira. The PCR products were subjected to REA and the profiles correlated well with phylogenetic relationships between these serovars. These data suggest that the PCR combined with REA could be a useful tool for rapid detection and preliminary differentiation of leptospires.