The present work helped to confirm the disease after feeding of the contaminated groundnut hay. The fungi isolated were Rhizopus oryzae, Fusarium sporotrichoides and Fusarium semitectum. Macroscopic, microscopic and molecular characterization of the fungi were done for confirmation. The fungi contaminated wheat materials were subjected to mycotoxins analysis using LC-MS/MS method. Fusarium sporotrichoides infected material had had apicidin 28.80 µg/kg and beauvericin 5126.67 µg/kg.Fusarium semitectum infected material was having apicidin 5375 µg/kg and beauvericin 2732.50 µg/kg of infected material. Sub chronic toxicity study of culture filtrates and fungi infected wheat materials were conducted in rats. The fungi were isolated from the contaminated groundnut hay which caused mycotoxicosis in cattle, exhibited the clinical signs of colic, tenesmus, ruminal atony, anorexia, bleeding from nostrils, rectum and fly bite site. Clinical signs observed in rats were diarrhoea, weakness, severe arching of back, swollen forehead and conjunctival haemorrhage.This study helped the farming community by identifying cause of the death of the cattle and reduced further mortality.