RESUMO
OBJECTIVE: To develop and evaluate a simple method for Leptospira serovar identification, based upon single-strand conformation polymorphism (SSCP) analysis of sequences amplified using diagnostic primers. DESIGN AND METHODS: PCR products of the G1/G2 and B64-I/B64-II diagnostic primer pairs were subjected to SSCP analysis of 15 percent polyacrylamide gels. Silver staining revealed polymorphisms which were used to standardise the method using reference strains. The method was then applied to the identification of clinical isolates of Leptospira. RESULTS: Closely related serovars, such as copenhageni and icterohaemorrhagiae, gave distinct but similar patterns. All other serovars could be discriminated by their SSCP profiles. The PCR-SSCP method was then applied to DNA extracted from tissue samples, allowing the identification of Leptospira serovars present in the tissues, in the absence of positive cultures. CONCLUSIONS: The method developed permits rapid and simple identification of Leptospira serovars in culture or in clinical material. Identification of serovars causing disease in many Caribbean populations will be possible for the first time.(AU)
