The role of HPV infections in the development of cervical cancer has acquired fundamental importance, and in recent years, HPV has become a relevant diagnostic and prognostic tool. Therefore, knowing the HPV genotypes that predominate in each country region is crucial.
In addition to HPV infection, different risk factors have been associated with the development of cervical cancer, such as early age at first sexual intercourse, number of sexual partners, high parity, education level, smoking habit, use of hormonal contraceptives and certain dietary deficiencies [16,17,18,19,20,21]. In our study, as depicted in Table 1, the number of pregnancies in women with CC was significantly higher than that in patients with CIN 1; different reports have shown that parity ≥3 is correlated with the risk of CC development. This result agrees with results from some [19, 22] but not all previous studies [23,24,25]. Consistent with results from previously published reports [26, 27], our study highlighted the use of hormonal contraceptives in a high percentage of women with CC (28.1%), while their use was lower in women with CIN 1 (14.1%). However, other studies did not find an association between the use of hormonal contraceptives and the risk of CC development [19, 24, 25]. In relation to tobacco consumption, smoking is one of the most studied cofactors for cancer development . In our analysis, 34.4% of women diagnosed with CC claimed to be smokers compared to 8.1% of women with CIN 1. Although this cofactor has been associated with a two-fold increase in the risk of CC development , some authors suggest that tobacco intake is only associated with the risk of developing squamous cell carcinoma and not adenocarcinoma .
Focusing on HPV infections in the open-population among different regions of Mexico, 12.1% (364/3000) of women attending routine gynaecological examinations were found to be HPV-positive, with an overall prevalence of 2.6% for HPV16 (n = 77/3000), 0.5% for HPV18 (n = 16/3000) and 9.0% for other HPVs excluding 16 and 18 (n = 271/3000) (Table 2). Similar results, like those of the FRIDA study, have been previously reported in Mexico; this study found global HPV positivity of 11.0% in an open-population as determined by COBAS 4800, of which 8.8% corresponds to high-risk HPVs (excluding HPV16 and HPV18) [30, 31]. Different rates of infection have been recorded in diverse parts of the world. Studies in other regions have reported an overall HPV prevalence of 12.2% in Canada , 34.5% in Peru, 10.3% in Iran, and 9.3% in Australia [14, 33, 34]. On the other hand, Aoyama-Kikawa et al. reported the presence of HPV infection in only 4.6% of Japanese women . These results support the findings of Becker et al., who showed regional and ethnic differences in HPV prevalence .
On the other hand, the most commonly detected HPV genotypes that we found by linear array in the open-population group were 16, 59, 66, 52, 51, 31, 39 and 56 (Fig. 1a). Although these genotypes have been detected in the open-population, we observed that some of them are also frequently present in the CC group, such as HPV 16, 59, 52, 51 and 66 (Fig. 3a). These genotypes have been classified by the IARC as carcinogenic to humans (except for HPV66, which is classified as “possibly” carcinogenic) , and there is sufficient evidence of their presence in different types of cancer, such as anus, vulva, vagina, penis and some head and neck cancers . Nevertheless, the heterogeneity of the HPV genotypic distribution among states in Mexico is evident in this work (Fig. S1-Additional file 1). Expanding on the findings of each common genotype identified in this study, HPV16 was the most prevalent genotype found in all diagnostic groups, as previously reported in Mexico [13, 38,39,40] and worldwide [41, 42] by different authors. Interestingly, HPV59 ranks second in the open-population group, fourth in CIN 1 and third in CC, and it has been identified in previous studies among different populations of Mexico including Cozumel (an island in eastern Mexico), Monterrey (northern Mexico) and Michoacan (western Mexico) [43,44,45]. It is important to highlight that Salcedo et al. detected HPV59 in only 1% of CC samples from Mexican women . Moreover, in a previous study by our research group using only the linear array HPV genotyping test, we also found a very low rate of HPV59 in cervical cancer (0.8%) . In both previously mentioned studies, samples were obtained from Mexican women before 2014. In the present study, in which the samples were collected from 2015 to 2019, a dramatic increase in the prevalence of HPV59 was observed, reaching 11.5% when all CC samples were analysed (n = 11/96) (Fig. 3a). This HPV genotype has also been observed in the top five HPV genotypes detected in different regions of the world, as reported, for example, in Ghana , China [48,49,50], and Switzerland .
HPV66 is the third most common genotype that we detected in the open-population (16.3%); it was the second most common in CIN 1 (23.7%) and the ninth in CC (2.7%). HPV66 was recently reported to also have a high prevalence in low-grade cervical lesions of women from Mexico City (45.7%); the prevalence was lower in CC samples (3.6%), although they used a different detection method (INNO-LiPA) . HPV66 has also been frequently found in Chile , Africa [54, 55] and China , among others.
HPV52 and 51 were also frequently found in this study; however, they were mostly detected as coinfections. Both genotypes have been commonly reported as single infections and also as coinfections; the presence of HPV51-52 and coinfections of three genotypes, such as HPV16-51-52, are frequently detected in coinfections [57, 58].
Otherwise, even though a positivity rate of HPV above 99% has been reported in cervical cancer [6, 59], we found only 77%, which was unexpected. However, in Belgium, the annual screening reports since 2017 showed that 15% of CC cases were HPV negative . Additionally, using PCR with GP6/GP5 HPV universal primers in 113 cervical cancer biopsies from Iranian women, a similar HPV-positive percentage rate was reported (78%) . We plan to evaluate the negative CC samples using next-generation sequencing to determine whether they are infected by other genotypes of HPV not detected by the LA.
It is important to mention that HPVs 16 and 18 are the most commonly detected high-risk genotypes worldwide, accounting for approximately 75% of all cervical cancer cases . In our study, the prevalence of these genotypes in all CC samples analysed (n = 96) was only 53.1% (38.5% for HPV16 and 14.6% for HPV18). A study performed in central Mexico reported a prevalence in CC samples of 34% for HPV16 and 5% for HPV18 . Therefore, the currently available vaccines may not be as effective in Mexico, and special attention should be paid to different genotypes that are not covered by the vaccine. Concerning HPV coinfections, as visualized in Fig. 1b, Figs. 2 and 3 b, almost all HPV genotypes preferentially occurred as multiple infections in the open-population and CIN 1 groups, while more than 50% occurred as single infections in the CC group, especially when HPVs 16, 18, 45, 59 and 58 were detected. Similar results have been reported previously in a Mexican population, where the most prevalent genotype detected in a single infection was HPV59, followed by HPV51 and HPV45 . Worldwide, genotypes 16, 18 and 45 have been detected at a higher rate as single infections .
Considering these results, regional data on the prevalence and genotypic distribution of HPVs are essential for estimating the impact of vaccines on cervical cancer and screening programmes. Vaccination programmes with a quadrivalent vaccine (HPV6, 11, 16 and 18) and bivalent vaccine (HPV16 and 18), which have been approved by the FDA, have been implemented in over 40 countries [64, 65]. However, in 2012, Serrano et al. described the potential impact of a nine-valent (9-HPV) vaccine against HPVs 6, 11, 16, 18, 31, 33, 45, 52 and 58, reporting that this vaccine could prevent almost 90% of CC cases worldwide . Nevertheless, this vaccine confers no protection against other HPV genotypes frequently found in women in the present study (HPV 59, 66, 51, 39 and 56). Therefore, it is necessary to continue analysing the geographical distribution of HPV genotypes in Mexico and worldwide to design effective HPV screening systems and develop new HPV vaccines. This is increasingly important as new technologies are rapidly detecting new HPV genotypes that cannot be detected by commercial detection tests [67,68,69,70].
It is important to mention that a limitation of this study was the lack of histological or cytological diagnosis of the samples from the women in the open-population group.