BBC Countryfile and Jeremy Clarkson are facing investigation by police after the show "disturbed" protected barn owls during filming for the show on his estate in the Cotswolds.
Last Sunday's edition of the countryside magazine show featured host Chris Packham photographing the protected animals where Clarkson lives in Chipping Norton, Gloucestershire.
After approaching one of the animals' nests, two of the birds flew off, prompting complaints from concerned viewers.
It is an offence to disturb the habitat of protected animals such as barn owls while they are nesting, and the stunt could see the BBC levied fines of near £5,000, according to the Daily Telegraph.
The two were taking photographs of the birds for a competition, which in itself requires a licence from the Department for Environment, Food and Rural Affairs (Defra).
It is also unclear whether the pair had obtained a license to snap the birds.
A spokesman for the Barn Owl Trust said: "We are slightly concerned that the programme gave the impression that flushing pairs of birds from potential nest sites is a common occurrence.
"This item was a great opportunity to mention the great care that is taken when potential nest sites are visited under licence.
"This is, after all, a Schedule 1-protected species.
"If the producers knew that the birds were not breeding and therefore that flushing them out was not an issue, it would have been good to have made this clear at the outset."
Thames Valley Police confirmed that an investigation is underway into the incident.
Wildlife expert Packham contests that the animals were disturbed. In a statement released by the BBC, he said: "I've been doing this kind of work for years and I'm confident there was no disturbance of the barn owls."
The BBC added that: "No-one from the team entered the barn while the birds were there or approached the nest directly."
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