Two of the Marsh Pride lions - stars of the BBC's long-running Big Cat Diary series - died after eating a poisoned cow carcass on Saturday night, while one more is missing and at least five are being treated by vets.
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Jonathan Scott, a zoologist and one of the show's original presenters who has been following the Marsh Pride for nearly 40 years, lamented the "end of an era".
Among the dead lions was 17-year old female Bibi, who had become something of a celebrity during the TV series' long and hugely popular run from 1996-2008.
A BBC wildlife crew at the scene said Bibi was found "lying on her side, foaming at the mouth, fitting and panting" before she died.
Conservationists first noticed something was wrong on Sunday morning when some of the lions, "were acting strangely collapsing and suffering from spasms," according to a statement from the David Sheldrick Wildlife Trust.
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Local cattle-herders who frequently bring their animals into the Masai Mara National Reserve to graze are suspected of the poisoning.
"Tens of thousands of cattle encroach into the reserve every night when visitors are safely out of sight," said Scott.
The grazing cattle are hunted by lions bringing the herders into conflict with the cats.
"There is nothing shocking anymore as to what is happening in the Masai Mara," he added, blaming years of "appalling management" by local authorities.
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In a statement Paula Kahumbu, chief executive of conservation group Wildlife Direct, condemned the poisoners as, "stupid, sick, cruel, and short-sighted people."
Two men were due to be charged with poisoning the lions on Tuesday, and face a maximum 20 million shilling ($196,000) fine or life imprisonment if found guilty.
"May they rot in jail," Kahumbu said.
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