A police file on the shooting of two of Britain’s rarest birds, about which Prince Harry has been questioned, is to be sent to the Crown Prosecution Service.

The Prince, who was shooting on the Royal Family’s Sandringham estate when a conservation warden saw two hen harriers blasted out of the sky, has been interviewed by police but denied any knowledge of the crime. A conviction for killing a protected bird carries a maximum six-month jail sentence and a £5,000 fine.

Prince Harry and a friend were asked about the shooting of the hen harriers, whose prey include game birds, last week. Three people, including a Natural England warden, had reported the killing. Neither the Prince nor his friend were questioned under caution.

The linking of Prince Harry with the incident has strained relations between Buckingham Palace and the RSPB. The Queen is the longstanding patron of the organisation but her role was being questioned last night. Mark Avery, its director of conservation, insisted that the organisation was “very proud” of the royal patronage, but there were concerns.

The RSPB has campaigned strongly against the persecution of birds of prey, particularly hen harriers, of which there are thought to be fewer than 20 pairs left in England. Most of the British population of just over 500 pairs is to be found in Scotland.

Natural England refused to name the member of staff who witnessed the shooting because police were still investigating.

Ash Murray, the site manager for Dersingham Bog, where the birds were shot, said that it was the first incident of hen harriers in the area being persecuted since he took up his post in 1998. He said: “The hen harriers don’t breed here but they fly down from Scotland or Scandinavia and stay here from October through to March because it’s a bit warmer. I have seen as many as four of them, two pairs in one night.

“The reason they’ve been persecuted is because they do feed on grouse, but it is an illegal act and there are deserved severe penalties.”

Conservationists believe that if it were not for persecution there would be at least 200 pairs across Britain. Gamekeepers and landowners are believed to be the birds’ biggest enemy because of the conflict of interest over pheasants, grouse and other game birds. A hen harrier is one of forty bird species that is “red-listed” in terms of conservation importance in the UK, meaning that it is globally threatened and has rapidly declined in number in Britain in the past 25 years.

Norfolk police are conducting the inquiry. The witnesses saw the birds shot but were unable to see who killed them because trees were obstructing the view. Investigators have been hampered by the absence of any hen harrier carcasses.

Unless there is substantial evidence, it is possible to escape conviction by maintaining that the bird was mistaken for legitimate quarry.

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