EPIDEMIOLOGIA MOLECOLARE DELLE INFEZIONI DA HUMAN PAPILLOMAVIRUS (HPV) E CHLAMYDIA TRACHOMATIS NELLA POPOLAZIONE GENERALE FEMMINILE

Introduction Sexually transmitted infections (STIs) are a major public health concern: according to WHO estimates, 448 million new cases of curable STIs (syphilis, gonorrhoea, chlamydia and trichomoniasis) occur annually throughout the world in adults aged 15-49 years. There are more than 30 different sexually transmissible bacteria, viruses, mycetes and protozoa. Human Papillomavirus (HPV) and Chlamydia trachomatis are the causes of two of the most common STIs. HPVs are double-stranded DNA viruses, grouped into cutaneous and mucosal types according to their infection site, and further subdivided into high-risk (HR) and low-risk (LR) genotypes, depending on their association with disease malignancy. More than 40 HPV genotypes can infect the genital area: HR-HPVs (e.g. HPV types 16 and 18) can cause cervical cancers and LR-HPVs (e.g. HPV types 6 and 11) can cause genital warts. It has been estimated that more than 50% of sexually active persons become infected with HPV at least once in their lifetime. Most of these infections are asymptomatic or subclinical and, fortunately, usually self-limiting. However, persistence of HPV infection can occur in 30% of cases and when these infections are sustained by HR types the risk to develop cancer is increased. Chlamydia trachomatis is an obligate intracellular bacterium. Currently, 18 different serovars of C. trachomatis have been identified, 10 of these infect the genital area. Silent and asymptomatic infection is common in both women and men. In women, acute infection with C. trachomatis can result in pelvic inflammatory disease, whose long-term consequences include chronic pain, ectopic pregnancy and infertility. C. trachomatis infection has also been demonstrated to be a risk factor for the acquisition of HPV infection. This work aimed at studying the molecular epidemiology of HPV and C. trachomatis infections; carrying out a microbiological surveillance of these infections by molecular and phylogenetic methods; evaluating the HPV/C. trachomatis co-infection in sexually active young women aged 13-24 years; evaluating the epidemiological aspects in relation to novel preventive strategies (for example the opportunity to set C. trachomatis screening programme). Materials and methods 1,557 cervical brush samples were collected from 1,557 women between January 2009 and December 2011. In particular: - 688 were collected from women aged 25-64 years (median age 36 years); - 563 from sexually active females aged 13-24 years (median age 19 years); - 306 from sexually active young women (median age 19 years, range: 19-21 years) vaccinated with a tetravalent HPV vaccine. These samples were collected one year after the end of the vaccination schedule. All samples were analyzed to identify the presence of HPV infection and genotypes using molecular methods. HPV-16 and HPV-18 positive samples were analyzed through phylogenetic methods in their L1 gene coding for the major capsid protein to characterize the potential vaccine escape mutants and in the LCR to investigate the circulation of geographical variants. Samples collected from adolescents/young women aged 13-24 years were analyzed to determine the presence of C. trachomatis by molecular methods. All C. trachomatis positive samples were phylogenetically analyzed in the ompA gene (coding for the major membrane protein MOMP) to evaluate the circulating serovars. Results The prevalence of HPV infection in sexually active women aged 13-64 years was 16.5%. In details, among adolescent/young women aged 13-24 years the prevalence was 22.7%, whereas in women aged 25-64 was 11.3%. According to several epidemiological studies, our data show a decrease in prevalence with increasing age of women (26% in 19-24-year old, 18.5% in 25-34-year old, 6.9% in the 35-44 years old and 4.9% in the 45-64 years old). Over 86% (among women aged 25-64 years) and 64%(among women aged 13-24 years) of the detected infections were attributable to HR genotypes. The type-specific prevalence showed HPV-16 as the predominant type, being 22.7% among HPV-positive women aged 25-64 years and 20% among those aged 13-24 years. The prevalence of HPV infections observed in young women vaccinated with tetravalent HPV vaccine was 17.3%, significantly lower (p<0.05) than that (26%) observed in age-matched unvaccinated population. It is noteworthy that, though the occurrence of infections decreased in vaccinated women, the number of HPV-genotypes involved in such infections remained constant; most of these infections were sustained by HR-clade genotypes. Virological surveillance is necessary to capture potential changes in circulating viruses and the potential emergence of escape mutants, which may affect the vaccine efficacy. Thus, the phylogenetic analysis of the two prevalent oncogenic HPV (HPV-16 and HPV-18) identified the spread of European variants in analyzed samples. Only two HPV-16 sequences belonged to Asian-American lineage. To understand the circulation of potential escape mutants, characterization of the L1 protein showed 11 amino acid mutations in HPV-16 and 7 in HPV-18. Of these mutations 27.3% and 42.9%, respectively, fell into L1 epitopes. Nevertheless, analysis of selective pressures showed that these sequences were under strong purifying selection. The prevalence of C. trachomatis infections in adolescent/young women aged 13-24 years was 5.7%. The phylogenetic analysis of C. trachomatis positive samples demonstrated 7 different serovars (D, E, F, G, H, J, and K). The prevalent serovars were the E and F - 38.1% and 23.8%, respectively. It has been demonstrated that serovars E and F have a biological advantage over the other serovars thanks to both their ability to escape the host immune response and the presence of specific virulence factors, which can facilitate the transmission and infectious processes. Three percent of adolescent/young women had C. trachomatis/HPV co-infection. The prevalence of C. trachomatis infection was higher among HPV-DNA positive women than among HPV-negative ones (13.2% vs. 3.4%, p<0.001), thus suggesting that persons with an ongoing STI are more likely to acquire other STIs. Conclusion The results of this research project provide new information on the spread of HPV and Chlamydia trachomatis infections in women aged 13-64. In the HPV vaccination era, epidemiological studies and virological surveillance of HPV infections in the general population can improve the understanding of the natural history and dynamics of the HPV infection, can shed lights on how the vaccines affect the incidence of HPV-infection and can point out whether current vaccines can confer cross-protection against other viral types. The HPV/Chlamydia trachomatis co-infections observed in adolescent/young women emphasize the greater vulnerability of this cohort with risky sexual behaviors. These results suggest that the introduction of a screening program for Chlamydia trachomatis in Italy should be considered to better understand the spread of this infection and to early treat the subjects.