A 25-year-old doctor died from the Crimean Congo Haemorrhagic Fever, also known as the "Congo fever", in the western city of Ahmedabad.
The dead include a woman who was infected with the virus, and the doctor and the nurse who treated her at a hospital in the city.
The virus has been detected in India for the first time, health officials say.
The doctor who died on Monday was an intern working in a government hospital.
It is not clear whether he came in contact with any of the previous victims of the disease.
Twenty teams of health officials fanned out within 5km of Kolat village in Gujarat state last month after the disease was detected, but no fresh cases were found.
Crimean Congo Haemorrhagic Fever is a viral haemorrhagic fever transmitted by ticks. It causes bleeding from the skin, mouth and nose.
The virus can also be transmitted from animals to humans.