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The prevalence of bovine tropical theileriosis (BTT) caused by Theileria annulata was studied in large ruminants of sub-Himalyan region of northern India using thin blood smear examination (TBE), polymerase chain reaction (PCR) and loop mediated isothermal amplification (LAMP) assays. About 11.04% blood samples were found positive for BTT. Maximum seasonal prevalence of BTT was observed in summer followed by rainy season and winter. Month-wise maximum prevalence was recorded in May and minimum in January. Laboratory examination with Giemsa staining for Koch’s blue bodies (KBB) and piroplasms revealed that 3.75% samples were positive for theileriosis. LAMP method gave the highest level of sensitivity followed by AS-PCR and TBE. Hence, it is suggested that LAMP can be a better molecular diagnostic tool for large scale epidemiological studies of bovine tropical theileriosis. Our results provide better understanding of this haemoprotozoan parasite and hence may help in devising timely control measures.