Sex workers who perform internal vaginal washing are three times more likely to get HIV than those who do not, a 10-year study in Kenya suggests.
And those who used detergents were at four times the risk, the University of Washington team reported.
The report, published in the Aids journal, suggested vigorous internal washing may cause inflammation, making infection more likely.
It is thought women may wrongly believe washing could get rid of infections.
However, the most effective way to protect against infections from HIV and other sexually transmitted diseases is to practise safe sex by using condoms.
The team from Washington University's International Aids Research and Training Programme spoke to more than 1,000 women in their study of risk factors for HIV-acquisition over a decade.
They found that internal vaginal washing was highly prevalent and was reported in a third of women from various clinical settings and regions in sub-Saharan Africa.
Study author Dr Scott McClelland said: "This is the first prospective study to demonstrate a significant association between vaginal washing and HIV acquisition.
"A causal association between vaginal washing and HIV acquisition seems biologically plausible."
He also argued that a thorough understanding of the beliefs surrounding the use of "intravaginal practices" was vital so that it could be tackled in a "culturally appropriate" manner.
The research team also said there was an urgent need for new initiatives to prevent HIV spreading to women, who account for a third of new infections.
Rod Watson, deputy head of health promotion at HIV/Aids charity Terrence Higgins Trust, said a link between HIV transmission and vaginal washing was not surprising.
"The vaginal lining is sensitive and easily damaged and the body's natural protection to some infections may be washed away.
"This means there's a greater chance that HIV or other sexually transmitted infections may be passed on.
"In an ideal world women would be educated about these risks and have access to condoms. Sadly, that kind of education is unavailable in many parts of Africa."
Chief Executive of HIV charity Crusaid Robin Brady said this kind of work was key to winning the fight against HIV/Aids in Africa.
He said: "Vaginal washing is commonplace in Africa and we need to understand how all intravaginal practices affect HIV transmission.
"The behavioural practices and cultural beliefs that lead to vaginal washing need to be better understood before any work to include the results of this study into prevention programmes on the ground can occur."
The research is published in Aids, the official journal of the International Aids Society.