Molecular tools in leptospirosis diagnosis and characterization of isolates.

ABSTRACT

The incidence of leptospirosis in human beings has been increasing in recent years. Early diagnosis and treatment can prevent complications and reduce mortality. The conventional laboratory methods for diagnosis rely on the demonstration of leptospires in clinical specimens, recovering the organisms in culture or the demonstration of antibodies to leptospires. Demonstration techniques have low sensitivity and specificity. Leptospires grow slowly and the positivity rate in culture is very low. Although microscopic agglutination test has been the cornerstone of serological diagnosis, the procedure is complex. New tests, like ELISA, dipstick test, lateral flow, etc, are relatively simple and rapid, but sensitivity is low during the early stages of the disease. The cross agglutination absorption test (CAAT) and typing with monoclonal antibodies (MCA) are the techniques used for serological characterization. These techniques are complicated and might not help in the case of certain serogroups. An alternate method for early diagnosis and characterization focuses on DNA-based techniques. Polymerase chain reaction (PCR), in situ hybridization etc are some of the methods used for early diagnosis, whereas restriction endonuclease analysis (REA), random amplified polymorphic DNA (RAPD) fingerprinting, arbitrarily primed PCR (AP-PCR), pulsed field gel electrophoresis (PFGE), ribotyping and DNA sequencing are useful for characterization. PCR is the most popular and quickest method for diagnosis. It can detect even if only a small number of organisms are present in a clinical sample. Fingerprinting tools such as RAPD, REA, RFLP, PFGE etc translate the complex genetic code into easily recognizable patterns, which facilitates characterization of the isolates up to sub-serovar level.