The latest 16 deaths were reported from the previously-unaffected districts of Unnao and Balrampur of Uttar Pradesh state, where hospitals were treating more than 1500 patients, said provincial health chief O.P. Singh.
The worst-hit district of Gorakhpur near Nepal's border has reported 392 deaths since late July, when encephalitis hit Uttar Pradesh, Singh told reporters in the state capital, Lucknow, adding that children were the main victims.
He said the latest casualties suggested the disease, which is transmitted through mosquito bites, had spread to 25 of the state's 70 districts.
The state's chief microbiologist, T.N. Dhole, who returned on Sunday after leading a scientists' team to Gorakhpur, painted a bleak picture of the situation in the remote area 250km southeast of Lucknow.
"It is catastrophic," he said. "The toll must be much, much higher because the government is only taking into account the deaths reported in state-run hospitals.
"Thousands of children must have died in the villages without medicines, and we also don't have any records of the deaths in private hospitals.
"When we visited the place, we interviewed villagers who had brought their sick children to hospital, and they told me that every village in and around Gorakhpur had 10 to 15 children suffering from the same symptoms.
"These poor people have no money to take them to city hospitals, leave alone pay for the medicines ... it's a disgrace," the government scientist added.
K.P. Kushwaha, who leads the paediatrics unit of Gorakhpur's state-run Medical College, sent an appeal on Sunday to the state administration.
"The toll must be much, much higher because the government is only taking into account the deaths reported in state-run hospitals"
Gorakhpur state chief microbiologist
"The toll will rise if steps are not taken. Kill the mosquitos, fast," he said by telephone from Gorakhpur, as Uttar Pradesh Chief Minister Mulayam Singh accused New Delhi of not dispatching enough vaccines.
Uttar Pradesh, India's most populous state, had sought federal supplies to vaccinate 7.5 million children below 15 years of age.
"But it has so far received only 200,000 vaccines," said health expert Rakesh Sehgal of India's Central Research Institute.
Japanese encephalitis, which first surfaced in Uttar Pradesh in 1978, causes inflammation of the brain and is followed by high fever and delirium.
Singh said efforts were on to eradicate mosquitos.
"We have intensified anti-vector fogging operations (spraying insecticide) and have given directions to smaller state-run district hospitals to open special wards so that children do not have to travel to Gorakhpur," Singh said.
He said a drive was under way to shut down pigpens in Lucknow and other towns. The virus is harboured by pigs and transmitted by mosquitoes to humans.