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The company has defended the policy as in the interests of children.
Sydney fireman Johnny McGirr, 33, said he was flying home from Brisbane in April when he took his seat next to two boys he estimated to be between 8 and 10 years old.
He was assigned the window seat but sat in the aisle seat so the two boys could look out the window.
However, a flight attendant approached him just as passengers were asked to put on their seatbelts, asking him to move.
Mr McGirr said when he asked why, he was told, "Well you can't sit next to two unaccompanied minors."
"She said it was the policy and I said, 'Well, that's pretty sexist and discriminatory. You can't just say becauseI'm a man I can't sit there,' and she just apologised and said that was the policy.
"By this stage everyone around me had started looking."
Mr McGirr said the attendant then asked a fellow female passenger, "Can you please sit in this seat because he is not allowed to sit next to minors."
"After that I got really embarrassed because she didn't even explain. I just got up and shook my head a little, trying to get some dignity out of the situation," he said.
"And that was it, I pretty much sat through the flight getting angrier."
Mr McGirr pointed out that he works as a fireman in Newtown in Sydney and was trusted in his job to look out for the welfare of children.
"(The attitude of the airline) is 'we respect you but as soon as you board a Virgin airline you are a potential paedophile', and that strips away all the good that any male does regardless of his standing in society, his profession or his moral attitudes," he said.
A spokeswoman for Virgin Australia confirmed the policy and said while they didn't want to offend male passengers, their priority was the safety of children.
"In our experience, most guests thoroughly understand that the welfare of the child is our priority," she said.
The spokeswoman said staff usually tried to keep the seat empty but when that was not possible a woman was seated next to the child.
"Virgin Australia takes the safety of all guests very seriously and in the case of unaccompanied minors, we take additional steps to ensure their flight is safe and trouble free in every respect."
Mr McGirr, who wrote to Virgin to complain, said the policy was flawed.
"(It's) blatant discrimination that just because I'm a male I can't sit there," he said. "...they apologised that it happened on the flight and said it shouldn't have happened then but my issue is not with the mistakes made there, my issue is with the policy in general.
"The majority of sexual assaults are (also) committed by men, does that mean that we can't sit next to women? Should we just have a seat by ourselves and that way women and children will be protected?"
Mr McGirr said he understood the children were vulnerable when not with an adult but fears about crimes committed by a small minority of people should not rule society.
Mr McGirr said Virgin should either allocate a chaperone for children to sit with them for the entire flight, have staff do regular checks on the children to see if they were all right or ask parents to purchase the seat that is vacant so it is always left empty.
Among other Australian airlines, budget carriers Jetstar and Tiger Airways do not accept unaccompanied minors on their flights, though the two airlines have different definitions of what constitutes a minor. Qantas has been contacted for comment on its unaccompanied minor policy.