Discussion

We found that alphapapillomavirus and betapapillomavirus types were detected together significantly less often than expected under independence of HPV types. Some of this was due to intra-genus clustering: both alphapapillomavirus and betapapillomavirus types are more likely to be detected with other types from the same genus. This clustering is likely due to shared common transmission routes with closely related types. There were still fewer than expected co-detections of alphapapillomavirus and betapapillomavirus types than expected after adjusting for intra-genus clustering, though confidence intervals could not exclude independence.
A substantial amount of previous research has shown that alphapapillomaviruses tend to cluster together; individuals who are positive for one alphapapillomavirus type are at a significantly higher risk of becoming positive for other alphapapillomavirus types.13-16 Presumably a large part of this association reflects shared risk factors across all alphapapillomavirus types, the most important being a common sexual transmission route as well as host-level susceptibility to infection. Therefore, it is unsurprising that we also found betapapillomaviruses tend to cluster together. This presumably also reflects a shared common transmission route across all betapapillomavirus types. However, what is much more surprising is the definite lack of clustering between alphapapillomaviruses and betapapillomaviruses. While the CIs of many of the measures of association include the null value expected under independence (1.0), all previous evidence13-16supports that independence between HPV types is not the expectation and that the expected clustering of HPVs of the alphapapillomavirus genus is of the order of OR/HRs of at least 1.5-3.0. Therefore, while the CIs include ORs and HRs of 1.0, we consider that this is a highly unusual result. We examine below two potential hypotheses for why alphapapillomaviruses and betapapillomaviruses may be less likely to be found together than expected.
The first hypothesis relates to potential biological interactions between genera. This could arise, for example, if infection with a first genus induces cross-immunity to subsequent infection with the other genus, or if both genera occupy the same ecological niche and infection with one genus leads to competitive exclusion of the other genus.17 We believe this hypothesis to be implausible for several reasons. HPV antibodies induced by natural infection tend to be type-specific,25 so it is unlikely that infection with types from one HPV genera would generate strong neutralizing cross-protection against infection with types from another HPV genera. Alphapapillomaviruses and betapapillomaviruses are also believed to have distinct tissue tropisms with preferred anatomical sites of infection.26 While alphapapillomaviruses have evolved to infect the mucosal epithelium, betapapillomaviruses have evolved to infect the cutaneous epithelium.2 Nonetheless, both genera are able to infect epithelial sites outside of their preferred trophic niche,6,8,27 so it is not impossible they could have overlapping niches leading to competitive exclusion. If this were the case, we would expect alphapapillomavirus types to be more likely to exclude cervicovaginal infections with betapapillomavirus types than the reverse due to their mucosal tropism. However, we did not find that alphapapillomavirus positivity reduced incident betapapillomavirus more than the reverse, so the hypothesis of competitive exclusion does not seem to be supported.
The second hypothesis is that the negative associations between genital alphapapillomaviruses and betapapillomaviruses could reflect independent or inversely correlated transmission mechanisms. Alphapapillomaviruses are sexually transmitted infections, with major risk factors including lifetime number of sex partners and new sex partners.28 Betapapillomaviruses are believed to be transmitted through skin-to-skin contacts,2 which include but are not restricted to sexual contacts. Interestingly, we found that lifetime and new sex partners were not risk factors for betapapillomavirus incidence, but having any sex during the previous interval was a risk factor. Most of the women who reported having sex in our study were having sex with a regular partner rather than a new one, and women who were married or living with their partner reported higher frequencies of sexual contacts than single or separated women. It could be that, in this study, having sex is an indicator for being in an ongoing intimate relationship with more skin-to-skin contacts, which include but may not be restricted to sex. It is also possible that betapapillomavirus detections may be depositions from recent sex rather than true infections, as the penile epithelium has been found to have a high betapapillomavirus prevalence.9 However, a previous study did not find that vaginal sex within the last 24 hours increased genital betapapillomavirus concordance between heterosexual partners.8 Nevertheless, that study and others have found that concordance was higher within than between couples, suggesting some form of transmission through intimate contact.8,29 Fingernails could be a potential reservoir for transmission and autoinoculation in the case of oral betapapillomavirus infections.11 We believe it is plausible that the negative association observed between betapapillomavirus and alphapapillomavirus types could partly reflect confounding due to inversely correlated transmission mechanisms, as women in our study who were having more regular sex and intimate skin-to-skin contact were women with fewer lifetime sexual partners or new sex partners. However, our adjustment for these sexual behaviors in multivariable models did not completely attenuate the negative association between alphapapillomaviruses and betapapillomaviruses. There could therefore remain some unmeasured confounding transmission risk factors explaining the negative association between betapapillomaviruses and alphapapillomaviruses.
The most detected betapapillomavirus types in cervicovaginal samples were HPV38, HPV5, HPV21, HPV22, and HPV8, respectively. Moscickiet al . also found that HPV38 was the most commonly detected betapapillomavirus type in cervicovaginal samples.8HPV38 is also one of the most commonly detected types in women’s oral samples,11 and among the most common betapapillomavirus types in men’s genital samples.9Genital HPV21 and HPV22 positivity were also common in male genital samples.9 HPV5 and HPV8 were conversely not among the most commonly reported detected types in male genital samples.9 HPV5 and HPV8 are the betapapillomavirus types which have been isolated from patients withepidermodysplasia verruciformis (EV) and are deemed to be possibly carcinogenic for individuals with EV by the International Agency for Research on Cancer.1,30 Oral HPV5 has also been associated with the risk of some head and neck cancers.3
By increasing the numbers of samples tested for betapapillomavirus, we were able to expand on our previous results which only included two observations per woman.19 The current study confirms our initial surprising findings that lifetime number of sex partners was inversely correlated with betapapillomavirus prevalence, and we propose above a potential explanation for this finding based on a multivariable analysis of sexual risk factors. However, this finding might reflect confounding from sexual behaviors that is specific to our study population, as studies of oral and penile betapapillomavirus infections have not found that number of lifetime sex partners was associated with betapapillomavirus prevalence at these sites.9,11,12,31 We are unaware of any other studies that have looked at risk factors for cervicovaginal betapapillomavirus prevalence. Due to the larger sample size, we were able to calculate incidence rates of betapapillomavirus positivity, and also detect lower than expected prevalent and incident co-detections of alphapapillomavirus and betapapillomavirus types which were not previously evident with fewer observations. However, despite the larger sample size, there were still large CIs for some associations; we therefore cannot exclude that our results could be due to chance. Our results would need to be confirmed in other studies of alphapapillomavirus and betapapillomavirus co-detection to verify whether they represent a more general trend across populations.
In conclusion, we found fewer prevalent and incident co-detections of alphapapillomaviruses and betapapillomaviruses than expected if HPV types belonging to these genera were transmitted independently. It is not clear whether these highly unusual findings reflect potential biological interactions between HPV genera, or are the result of inversely correlated transmission mechanisms, or simply represent a chance result. These findings need to be confirmed in other populations. This would help further elucidate the biology of betapapillomaviruses, a genus for which there is growing evidence of a role in carcinogenesis.