A hotel in the town was visited by Gardai at the weekend and two women were removed from the premises.
It follows on from two incidents in the town a week previously when two women were removed from two different pubs.
Both women are believed to have been offering sexual services to customers.
Donegaldaily.com understands that at least 12 different prostitutes are operating in the Cathedral Town at any one time.
The majority of the women stay in the town for a number of days before they are moved around.
Increasing pressure is coming on Gardai to act on the prostitution problem in the town.
A source said prostitutes are becoming much more obvious.
“There are prostitutes in other towns in Donegal but Letterkenny has the greatest number because of its population.
“There are a number of long-term let apartments which are now being used for the trade.
“But people must remember that it is not the women who are making the majority of the money but the people who are running them,” he said.