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BackgroundLeptospirosis is a re-emerging infectious disease that is under-recognized due to low-sensitivity and cumbersome serological tests. MAT is the gold standard test and it is the only serogroup specific test used till date. Rapid reliable alternative serogroup specific tests are needed for surveillance studies to identify locally circulating serogroups in the study area.Methods/Principal FindingsIn the present investigation the serological specificity of leptospiral lipopolysaccharides (LPS) was evaluated by enzyme linked immunosorbent assay (ELISA), dot blot assay and rapid immunochromatography based lateral flow assay (ICG-LFA). Sera samples from 120 MAT positive cases, 174 cases with febrile illness other than leptospirosis, and 121 seronegative healthy controls were evaluated for the diagnostic sensitivity and specificity of the developed assays. LPS was extracted from five locally predominant circulating serogroups including: Australis (27.5%), Autumnalis (11.7%), Ballum (25.8%), Grippotyphosa (12.5%), Pomona (10%) and were used as antigens in the diagnostics to detect IgM antibodies in patients’ sera. The sensitivity observed by IgM ELISA and dot blot assay using various leptospiral LPS was >90% for homologous sera. Except for Ballum LPS, no other LPS showed cross-reactivity to heterologous sera. An attempt was made to develop LPS based ICG-LFA for rapid and sensitive serogroup specific diagnostics of leptospirosis. The developed ICG-LFA showed sensitivity in the range between 93 and 100% for homologous sera. The Wilcoxon analysis showed LPS based ICG-LFA did not differ significantly from the gold standard MAT (P>0.05).ConclusionThe application of single array of LPS for serogroup specific diagnosis is first of its kind. The developed assay could potentially be evaluated and employed for as MAT alternative.

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Leptospirosis has been recognized as the disease of the dogs. The prevention of canine leptospirosis can block the transmission of the etiological agent to humans. Commercial vaccines prevent not only clinical leptospirosis but also the renal carrier state in dogs. Thus in this present investigation, the humoral immune response of the vaccinated dogs with the multivalent vaccine (Megavac-6) was analyzed. Enzyme linked immunosorbent assay (ELISA) with preparations of 2 whole cell lysate, 2 leptospiral LPS, 4 purified recombinant proteins were used to test sera from 30 dogs vaccinated with MEGAVAC-6, a commercial vaccine practiced by the animal husbandary Departments in Tamilnadu. All 30 sera were positive by ELISA with whole cell lysate of Canicola and Icterohaemorrhagiae, and ELISA with Canicola LPS, Icterohaemorrhagiae LPS; antibody titres ranged from 20 to ≥10,240. Although there was less frequent reactivity of sera with recombinant antigens, antibodies to LipL32 and LigA were detected in 25 (83.3%) serum samples. Less frequent reactivity was noted when recombinant GroEl, LK73.5 antigens were included separately in ELISAs. The highest MAT titre was observed against the serovar Canicola and Icterohaemorrhagiae. The more reactivity against LPS may be due to the dominance of serovar specific leptospiral LPS, which may be the dominant immunogenic antigen in inactivated bivalent leptospiral vaccines. Eventhough LPS is the dominant antigen. It is serovars specific and hence recommends the incorporation of the locally circulating serovars in vaccine for its efficient use. The efficiency at this point can be increased by the use of subunit vaccines rather than recombinant proteins as such. The present study has proposed certain epitopes with increased antigenic potency.