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

Human Papillomavirus (HPV) infection and genotype frequency in the oral mucosa of newborns in Milan, Italy

Human papillomavirus (HPV) causes cutaneous and mucosal infections in both adults and children. In order to evaluate HPV prevalence and the spectrum of genotypes in the oral cavity of paediatric subjects, a retrospective study was carried out on oral-pharyngeal swabs collected from 177 newborns aged 0-6months. HPV-DNA was detected by a nested-PCR; the viral typing was made through DNA sequencing. HPV infection was identified in 25 subjects (14.1%) and the sequence analysis showed eight distinct genotypes. These data confirm HPV detection in newborn oral mucosa. Further investigations are needed to clarify the methods of HPV acquisition

HPV genotypes detected in the oropharyngeal mucosa of HIV-infected men who have sex with men in Northern Italy

The aim of this study was to investigate the epidemiological profile of HPV oropharyngeal infections in HIV-infected men who have sex with men. A total of 135 subjects were enrolled at the L. Sacco University Hospital (Milan, Italy) to evaluate their HPV oropharyngeal infection status at baseline and at a follow-up visit at least 12 months later. HPV DNA was detected from oropharyngeal swabs using an in-house nested PCR that amplifies a segment of the L1 gene. The PCR products were then sequenced and genotyped. A greater percentage of high-risk genotypes was identified compared to low-risk genotypes (13\ub77% vs. 6\ub79%, P &lt; 0\ub705), and two uncommon alpha-HPV genotypes were detected, i.e. HPV-102 and HPV-114. HPV infection prevalence was 24\ub74% and the cumulative incidence was 24\ub71%. During the follow-up period, one case of HPV infection (HPV-33) persisted, while the overall rate of infection clearance was 58\ub73%. HPV oropharyngeal infection was widespread in the cohort examined, and most of the infections were transient and cleared within 12 months. These results may help to clarify the role of HPV in the oropharynx and may also improve our understanding of the need to implement preventive strategies in at-risk populations

Molecular characterization and phylogenetic analysis of human influenza A viruses in three consecutive seasons with different epidemiological profiles

INTRODUCTION: Influenza activity and influenza virus circulation were observed in Lombardy (northern Italy) during three consecutive seasons and the molecular characteristics of circulating viruses analysed to control for introduction of new variants. METHODS: The molecular characterization of 38 isolates, namely 20 A/H3N2 and 18 A/H1N1 influenza strains from the 2005/06, 2006/07 and 2007/08 seasons, was performed by sequence analysis of the globular head region of the HA protein (HA1 subunit), specific for influenza virus A/H3 and A/H1. RESULTS AND DISCUSSION: The last three influenza seasons in the study region were characterized by medium-low activity. A typical co-circulation of several variants was shown for A/H3 viruses for approximately two years and were subsequently almost entirely substituted by new emerging variants. Vice versa, A/H1 viruses had a more homogeneous circulation with a single lineage clearly dominating each season. The HA sequences of the A/H3 and the A/H1 viruses isolated in the last three seasons fell into 4 and 3 principal phylogenetic groups, respectively. No evidence of positive or negative selection in the sequence alignments was observed. CONCLUSIONS: Molecular characterization of the influenza viruses in three consecutive seasons highlighted considerable heterogeneity in their HA sequences. A careful surveillance of genetic changes in the HA1 domain during seasonal influenza epidemics may reveal immune escape and provide early information on newly emerging strains with epidemiologic inference

Chlamydia trachomatis prevalence and chlamydial/ HPV co-infection among HPV-unvaccinated young italian females with normal cytology

Infections caused by Chlamydia trachomatis (Ct) and human papillomavirus (HPV) are the two main sexually transmitted infections; however, epidemiological data on Ct prevalence and Ct/HPV co-infection in Italy are scant. This study aimed at estimating the prevalence of Ct infection and Ct/HPV co-infection in young HPV-unvaccinated females with normal cytology, and placed particular attention on the possible association between Ct-DNA positivity and different HPV infecting genotypes. Five hundred 66 healthy females aged 16-26 years without cervical lesions, previously assessed for HPV infection (HPV-DNA prevalence: 18.2%), were tested for Ct-DNA. The overall prevalence of Ct was 5.8% (95% CI: 4.2-8.1), while Ct/HPV co-infection was recorded in 2.7% (95% CI: 1.6-4.3) of subjects. Compared with HPV-DNA-negative females, HPV-DNA positive subjects had significantly (P &lt; 0.001) higher odds of being infected with Ct (odds ratio of 4.20, 95% CI: 2.01-8.71). Both Ct and Ct/HPV infections were much more prevalent in under 18-year-olds than in older women. Subjects positive for single high-risk HPV genotypes and various multiple HPV infections had higher odds of being Ct-DNA positive. Our findings confirm that HPV and Ct infections are very common among asymptomatic young Italian females. This underlines the urgent need for nationwide Ct screening programs and reinforcement of sexual health education, which would be the most important public health strategies, since no Ct vaccines are currently available

Human papillomavirus detection in paraffin-embedded colorectal cancer tissues

Human papillomavirus (HPV) has a well-recognized aetiological role in the development of cervical cancer and other anogenital tumours. Recently, an association between colorectal cancer and HPV infection has been suggested, although this is still controversial. This study aimed at detecting and characterizing HPV infection in 57 paired biopsies from colorectal cancers and adjacent intact tissues using a degenerate PCR approach. All amplified fragments were genotyped by means of sequencing. Overall, HPV prevalence was 12.3 %. In particular, 15.8% of tumour tissues and 8.8% of non-cancerous tissue samples were HPV DNA-positive. Of these samples, 85.7% were genotyped successfully, with 41.7% of sequences identifying four genotypes of the HR (high oncogenic risk) clade Group 1; the remaining 58.3% of HPVgenotyped specimens had an unclassified \u3b2-HPV. Examining additional cases and analysing whole genomes will help to outline the significance of these findings

Measles surveillance activities in the Metropolitan Area of Milan during 2017-2018

Introduction In Italy, the transmission of measles is still endemic, and 7,919 cases were reported to the National Surveillance System between January 2017 and December 2018. Aim of this study is to report the results of the measles surveillance activities in the Metropolitan City of Milan from March 2017 to December 2018, and to evaluate the surveillance performance WHO indicators. Methods The Local Health Units (LHUs) carried out case investigations and collected specimens to send to the EpiSoMI Lab (Subnational Reference Laboratory, SRL) of the University of Milan for cases/outbreaks confirmation and genotyping performed according to the WHO Guidelines. Results Overall, 610 suspected measles cases were reported by the surveillance system of the Metropolitan City of Milan. A total of 439 out of 540 cases with adequate specimens were laboratory-confirmed by molecular and/or serological assays. Two-hundred and thirty-six cases were notified as sporadic and 203 as related to 94 outbreaks. The most confirmed cases were aged 15\u201339 years, almost all not vaccinated. Overall, 282 cases were genotype D8 and 118 genotype B3. The evaluation of a set of indicators to monitor the quality of surveillance activities demonstrated the proficiency of the EpiSoMI Lab. Conclusions A well-done investigation of cases and outbreaks by the surveillance local system, in a timely manner, in order to notify and investigate suspected cases and to laboratory confirm or discard cases is fundamental to reduce morbidity, to prevent further virus transmission and to achieve measles elimination

Ongoing large measles outbreak with nosocomial transmission in Milan, northern Italy, March-August 2017

A large measles outbreak has been ongoing in Milan and surrounding areas. From 1 March to 30 June 2017, 203 measles cases were laboratory-confirmed (108 sporadic cases and 95 related to 47 clusters). Phylogenetic analysis revealed the co-circulation of two different genotypes, D8 and B3. Both genotypes caused nosocomial clusters in two hospitals. The rapid analysis of epidemiological and phylogenetic data allowed effective surveillance and tracking of transmission pathways

Modifiable and non-modifiable factors related to HPV infection and cervical abnormalities in women at high risk: a cross-sectional analysis from the Valhidate Study

Abstract: Migrant women, and women infected with HIV, are at enhanced risk of cervical HPV infection and HPV-related cancers. We investigated factors that can reduce these risks through public health preventive and screening interventions. We compared prevalence and risk factors for cervical HPV infection/lesions in women with HIV-infection (HIW) and migrant women (RMW) with a control group of resident women (SPW) who were enrolled in the study for the eVALuation and monitoring of HPV Infections and relATEd cervical diseases in high-risk women (VALHIDATE). Among 3093 evaluable women, age-standardized HPV prevalence was 36.3% (95%CI: 28.1\u201344.4) in HIW, 21.6% (95%CI: 15.7\u201327.5) in RMW, and 14.3% (95%CI: 12.5\u201316.1) in SPW. Adjusted prevalence of HPV infection was 2.07 times higher among HIW (95%CI: 1.75\u20132.45), and 1.45 times higher among RMW (95%CI: 1.17\u20131.80) than in SPW. Prevalence-ratios of SIL and HG-SIL were 2.67 (95%CI: 2.06\u20133.45) and 2.82 (95%CI: 1.28\u20136.20), respectively, in HIW compared to controls. A multivariate log-binomial regression model showed modifiable risk factors associated with HPV infection/lesion to have different patterns among groups. Specific public-health intervention, including health and sexual-health education, safe-sex procedures, and improvements to screening programmes, could favorably affect these highly vulnerable women

Genetic characterisation of Measles virus variants identified during a large epidemic in Milan, Italy, March&#8211;December 2017

In 2017, Italy experienced a large measles epidemic with 5408 cases and four deaths. As Subnational Reference Laboratory of the Measles and Rubella surveillance NETwork (MoRoNET), the EpiSoMI (Epidemiology and Molecular Surveillance of Infections) Laboratory (University of Milan) set up rapid and active surveillance for the complete characterisation of the Measles virus (Mv) responsible for the large measles outbreak in Milan and surrounding areas (Lombardy, Northern Italy). The aims of this study were to describe the genetic profile of circulating viruses and to track the pathway of measles transmission. Molecular analysis was performed by sequencing the highly variable 450 nucleotides region of the N gene (N-450) of Mv genome. Two-hundred and ninety-nine strains of Mv were analysed. The phylogenetic analysis showed five different variants, two not previously described in the studied area, belonging to D8 and B3 genotypes. Three events of continuous transmission of autochthonous variants (D8-Osaka, D8-London and B3-Milan variants) and two events of continuous transmission of imported variants (B3-Dublin and D8-Hulu Langat) tracked five different transmission pathways. These pathways outlined two epidemic peaks: the first in April and the second in July 2017. The correlation between Mv variant and the epidemiological data may enable us to identify the sources of virus importation and recognise long-lasting virus transmission pathways