IS1533-BASED PCR ASSAY FOR IDENTIFICATION OF LEPTOSPIRA-INTERROGANS SENSU-LATO SEROVARS

A PCR-based assay was developed for typing L. interrogans sensu late serovars. The assay is designed to exploit the presence of many copies of the leptospiral insertion sequence IS1533 and IS1533-like sequences present in the genomes of most leptospiral serovars. The PCR primers were designed to amplify DNA of unknown sequence between closely placed IS1533 or IS1533-like sequences. Amplification reactions primed with IS1533-based primers generated products of different sizes. When few copies of IS1533 were present in the genome, amplification of a few products was still detected. These results suggest that IS1533 elements may be found close together. Analysis of DNA amplified from different serovars showed the presence of differently sized products, thus enabling the serovars to be identified. Genetic variation among isolates within the same serovar was also demonstrated with the IS1533-based primers. Amplification reactions using DNA extracted from the urine of infected animals generated specific products which were similar to the products generated from purified bacterial DNA. These results demonstrate that this assay is selective enough to be used for typing leptospiral serovars from clinical material and thus allows leptospiral typing without isolation of the bacteria in pure culture.