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A total of 102 serum samples were collected randomly from buffaloes exposed with different clinical conditions (abortion, repeat breeding, fever, mastitis, anorexia) suspected for leptospirosis and apparently healthy. These serum samples were subjected to seroepidemiogical study using microscopic agglurination test (MAT) having different serovars of Leptospira spp. The seroprevalence of leptospirosis among buffaloes was noted to be 15.69% (16/102). All the three districts of South Gujarat showed the presence of leptospiral antibodies without any significant difference (P≤0.05) with the highest rate in Tapi (50.00%) followed by Navsari (14.89%) and Surat (13.72%). Jafrabadi breed showed 50.00% seropositivity followed by Surati (16.67%), Mehsana (15.00%) and Non-Descript (5.55%). In female buffaloes the seroprevalence positivity was noted in 16.49% cases. However, none of male exhibited seropositivity. In respect of age groups the highest rate of seropositivity (19.23%) was observed in age group of 1 to 4 years followed by above 4 years (15.71%) and below 1 year (00%) without significant difference (P≤0.05). In buffaloes out of 102 sera screened, 16 were positive with one or more serovars. The highest number of seropositivity was recorded against serovar Kaup (17.39%).