In our study of sampling and transport methods, oral rinse samples were significantly more sensitive than self-collected swabs and absorbing the oral rinse into a tampon for postage resulted in a significantly lower rate of oral HPV detection. HPV detection was more likely in those who had recently brushed their teeth and in current smokers. A higher number of lifetime sexual partners for tongue kissing and oral-penile sex, also predicted oral HPV detection. However only a minority of men with high numbers of oral sex partners were HPV positive, and prevalence in the mouth was much lower than prevalence in the anus, which was high in this and other studies of MSM. 
The same genotype of HPV was detected in 83% of the 12 men retested six months later. Together these data suggest that oral HPV infection may be difficult to acquire, but once present may persist many years.
The finding that recent tooth-brushing increases HPV detection also suggests that current sampling techniques may be improved by prior epithelial abrasion, similar to that used for anogenital HPV detection in men. 
The likelihood of detecting oral HPV fell in a linear fashion by about 14% with each additional hour after brushing teeth, suggesting that abrasion of oral mucosa improves collection of infected cells in an oral rinse. D’Souza and coworkers have reported an association between oropharyngeal cancer and infrequent toothbrushing, but this has not been reported for HPV detection and may be related to a different causal pathway for oropharyngeal cancer. 
These investigators have combined oropharyngeal brushings with oral rinses 
and have shown higher detection in oral rinses than in brushings 
but there are no reports comparing HPV detection in oral rinse samples with and without prior abrasion. Oral HPV detection was also associated with current smoking and this also has been reported by D’Souza. 
Smoking causes oral epithelial thickening and periodontal disease 
and given our observation that epithelial abrasion increases HPV detection, it may be that the epithelial effects of smoking contribute to increased detection in smokers.
Reports differ on whether oral HPV is significantly associated with recent or lifetime numbers of sexual partners 
. There are three potential reasons for these differences. Firstly, if only young adults within a few years of onset of sexual activity are studied, recent partners may approximate lifetime partners. This explanation is suggested by the observation that oral HPV is significantly associated with recent partners in studies that involve only young adults (age<24 
). Secondly, most studies are of women and heterosexual men with fewer partners than the MSM population assessed in the present study. Finally, we tested the association of oral HPV with partner numbers in non-overlapping time periods, whereas other investigators have not separated them.
We found that HPV detection rose with age, consistent with most other studies 
and importantly no studies have shown significantly declining prevalence with age. An increasing prevalence of HPV with age is consistent with the finding that lifetime, but not recent partner numbers, are most strongly associated with oral HPV detection suggesting that once acquired, oral HPV infection persists a long time. However if this is true the relatively low prevalence of HPV infection, despite a high numerical sexual exposure can only be explained if oral HPV infection is difficult to acquire. These findings contrast with cervical HPV infection which is acquired rapidly after commencing sexual activity, is related to recent sexual partners and its prevalence falls with age. 
Our study had a number of limitations. Firstly, it was cross-sectional and the associations are subject to the limitations of this design, such as unmeasured or incomplete adjustment for confounding. Secondly, the participants were from one sexual health service and therefore it may not be reasonable to generalise these findings to populations with a lower sexual risk. Finally, the reported number of sexual partners may have been affected by recall or social desirability bias.
Future studies could examine the effect of brushing prior to obtaining oral rinse samples to enhance HPV detection. We found a high level of HPV persistence in the small number of men tested after six months. Larger longitudinal studies are required to confirm this and to establish the age of acquisition of oral HPV to inform future vaccination policies.