Characterisation of complete high- and low-risk human papillomavirus genomes isolated from cervical specimens in southern Brazil.

The classification of human papillomavirus (HPV) intratypic lineages by complete genome sequencing is a determinant in understanding biological differences in association with this disease. In this work, we have characterised complete HPV genomes from southern Brazil. Fifteen cervicovaginal Pap smear negative samples previously categorised as HPV-positive were sequenced using ultradeep sequencing, and 18 complete genomes from 13 different HPV types were assembled. Phylogenetic and genetic distance analyses were performed to classify the HPV genomes into lineages and sublineages. This is the first report describing the distribution of HPV intratype lineages of high and low oncogenic risk in asymptomatic women from southern Brazil.

Human papillomavirus type distribution and HPV16 intratype diversity in southern Brazil in women with and without cervical lesions.

BACKGROUND: Increasing evidence suggests that human papillomavirus (HPV) intratype variants (specific lineages and sublineages) are associated with pathogenesis and progression from HPV infection to persistence and the development of cervical cancer. OBJECTIVES: This study aimed to verify the prevalence of HPV infection and distribution of HPV types and HPV16 variants in southern Brazil in women with normal cytology or intraepithelial lesions. METHODS: HPV typing was determined by L1 gene sequencing. To identify HPV16 variants, the LCR and E6 regions were sequenced, and characteristic single nucleotide variants were identified. FINDINGS: A total of 445 samples were studied, with 355 from cervical scrapes and 90 from cervical biopsies. HPV was detected in 24% and 91% of these samples, respectively. The most prevalent HPV types observed were 16 (cervical, 24%; biopsies, 57%) and 58 (cervical, 12%; biopsies, 12%). Seventy-five percent of the HPV16-positive samples were classified into lineages, with 88% defined as lineage A, 10% as lineage D, and 2% as lineage B. MAIN CONCLUSIONS: This study identified a high frequency of European and North American HPV16 lineages, consistent with the genetic background of the human population in southern Brazil.

High APOBEC3B mRNA Expression Is Associated with Human Papillomavirus Type 18 Infection in Cervical Cancer.

The APOBEC3 (A3) proteins are cytidine deaminases that exhibit the ability to insert mutations in DNA and/or RNA sequences. APOBEC3B (A3B) has been evidenced as a DNA mutagen with consistent high expression in several cancer types. Data concerning the A3B influence on HPV infection and cervical cancer are limited and controversial. We investigated the role of A3B expression levels in cervical cancer in affected women positive for infection by different HPV types. Tumor biopsies from cancerous uterine cervix were collected from 216 women registered at Hospital do Câncer II of Instituto Nacional de Câncer, and infecting HPV was typed. A3B expression levels were quantified from RNA samples extracted from cervical biopsies using real-time quantitative PCR. Median A3B expression levels were higher among HPV18+ samples when compared to HPV16+ counterparts and were also increased compared to samples positive for other HPV types. In squamous cell carcinoma, HPV18+ samples also showed increased median A3B expression when compared to HPV Alpha-9 species or only to HPV16+ samples. Our findings suggest that A3B expression is differentially upregulated in cervical cancer samples infected with HPV18. A3B could be potentially used as a biomarker for HPV infection and as a prognostic tool for clinical outcomes in the context of cervical cancer.

Human papillomavirus E6 triggers upregulation of the antiviral and cancer genomic DNA deaminase APOBEC3B.

UNLABELLED: Several recent studies have converged upon the innate immune DNA cytosine deaminase APOBEC3B (A3B) as a significant source of genomic uracil lesions and mutagenesis in multiple human cancers, including those of the breast, head/neck, cervix, bladder, lung, ovary, and other tissues. A3B is upregulated in these tumor types relative to normal tissues, but the mechanism is unclear. Because A3B also has antiviral activity in multiple systems and is a member of the broader innate immune response, we tested the hypothesis that human papillomavirus (HPV) infection causes A3B upregulation. We found that A3B mRNA expression and enzymatic activity were upregulated following transfection of a high-risk HPV genome and that this effect was abrogated by inactivation of E6. Transduction experiments showed that the E6 oncoprotein alone was sufficient to cause A3B upregulation, and a panel of high-risk E6 proteins triggered higher A3B levels than did a panel of low-risk or noncancer E6 proteins. Knockdown experiments in HPV-positive cell lines showed that endogenous E6 is required for A3B upregulation. Analyses of publicly available head/neck cancer data further support this relationship, as A3B levels are higher in HPV-positive cancers than in HPV-negative cancers. Taken together with the established role for high-risk E6 in functional inactivation of TP53 and published positive correlations in breast cancer between A3B upregulation and genetic inactivation of TP53, our studies suggest a model in which high-risk HPV E6, possibly through functional inactivation of TP53, causes derepression of A3B gene transcription. This would lead to a mutator phenotype that explains the observed cytosine mutation biases in HPV-positive head/neck and cervical cancers. IMPORTANCE: The innate immune DNA cytosine deaminase APOBEC3B (A3B) accounts for a large proportion of somatic mutations in cervical and head/neck cancers, but nothing is known about the mechanism responsible for its upregulation in these tumor types. Almost all cervical carcinomas and large proportions of head/neck tumors are caused by human papillomavirus (HPV) infection. Here, we establish a mechanistic link between HPV infection and A3B upregulation. The E6 oncoprotein of high-risk, but not low-risk, HPV types triggers A3B upregulation, supporting a model in which TP53 inactivation causes a derepression of A3B gene transcription and elevated A3B enzyme levels. This virus-induced mutator phenotype provides a mechanistic explanation for A3B signature mutations observed in HPV-positive head/neck and cervical carcinomas and may also help to account for the preferential cancer predisposition caused by high-risk HPV isolates.

The role of cytidine deaminases on innate immune responses against human viral infections.

The APOBEC family of proteins comprises deaminase enzymes that edit DNA and/or RNA sequences. The APOBEC3 subgroup plays an important role on the innate immune system, acting on host defense against exogenous viruses and endogenous retroelements. The role of APOBEC3 proteins in the inhibition of viral infection was firstly described for HIV-1. However, in the past few years many studies have also shown evidence of APOBEC3 action on other viruses associated with human diseases, including HTLV, HCV, HBV, HPV, HSV-1, and EBV. APOBEC3 inhibits these viruses through a series of editing-dependent and independent mechanisms. Many viruses have evolved mechanisms to counteract APOBEC effects, and strategies that enhance APOBEC3 activity constitute a new approach for antiviral drug development. On the other hand, novel evidence that editing by APOBEC3 constitutes a source for viral genetic diversification and evolution has emerged. Furthermore, a possible role in cancer development has been shown for these host enzymes. Therefore, understanding the role of deaminases on the immune response against infectious agents, as well as their role in human disease, has become pivotal. This review summarizes the state-of-the-art knowledge of the impact of APOBEC enzymes on human viruses of distinct families and harboring disparate replication strategies.

Human papillomavirus type distribution and HPV16 intratype diversity in southern Brazil in women with and without cervical lesions

BACKGROUND Increasing evidence suggests that human papillomavirus (HPV) intratype variants (specific lineages and sublineages) are associated with pathogenesis and progression from HPV infection to persistence and the development of cervical cancer. OBJECTIVES This study aimed to verify the prevalence of HPV infection and distribution of HPV types and HPV16 variants in southern Brazil in women with normal cytology or intraepithelial lesions. METHODS HPV typing was determined by L1 gene sequencing. To identify HPV16 variants, the LCR and E6 regions were sequenced, and characteristic single nucleotide variants were identified. FINDINGS A total of 445 samples were studied, with 355 from cervical scrapes and 90 from cervical biopsies. HPV was detected in 24% and 91% of these samples, respectively. The most prevalent HPV types observed were 16 (cervical, 24%; biopsies, 57%) and 58 (cervical, 12%; biopsies, 12%). Seventy-five percent of the HPV16-positive samples were classified into lineages, with 88% defined as lineage A, 10% as lineage D, and 2% as lineage B. MAIN CONCLUSIONS This study identified a high frequency of European and North American HPV16 lineages, consistent with the genetic background of the human population in southern Brazil.

Characterisation of complete high- and low-risk human papillomavirus genomes isolated from cervical specimens in southern Brazil

The classification of human papillomavirus (HPV) intratypic lineages by complete genome sequencing is a determinant in understanding biological differences in association with this disease. In this work, we have characterised complete HPV genomes from southern Brazil. Fifteen cervicovaginal Pap smear negative samples previously categorised as HPV-positive were sequenced using ultradeep sequencing, and 18 complete genomes from 13 different HPV types were assembled. Phylogenetic and genetic distance analyses were performed to classify the HPV genomes into lineages and sublineages. This is the first report describing the distribution of HPV intratype lineages of high and low oncogenic risk in asymptomatic women from southern Brazil.

Prevalência dos tipos de Papilomavírus Humano em mulheres atendidas em um Hospital Universitário no Sul do Brasil / Prevalence of Human Papillomavirus types in women attending at University hospital in southern Brazil

Determinar a prevalência e os genótipos do HPV em mulheres atendidas em um Hospital Universitário no Sul do Brasil. Metodologia: Foram coletadas amostras de secreções cérvico-vaginal de 200 mulheres. O HPV foi detectado pela Reação em Cadeia da Polimerase aninhada e os genótipos por sequenciamento. As variáveis foram analisadas pelo Teste Exato de Fisher e pelo Chi-quadrado de Pearson com o nível de significância < 5%. A força de associação foi calculada pela razão de prevalência e os seus intervalos de confiança a 95%. A análise Multivariada foi calculada pela Regressão Logística Binária para as variáveis com P <0,20. Resultados:O DNA do HPV foi detectado em 55 mulheres (27,5%). A prevalência do HPV foi associada a baixa renda(P =0,01), o início sexual precoce (P <0,001), a gestação (P = 0, 002), a infecção pelo HIV–1 (P = 0,001) e a coilocitose no exame citopatológico (P =0,006). Houve associação entre o status sorológico para o HIV–1 e os genótipos HPV–33 (P =0,001) e HPV–68 (P <0,001). Na análise multivariada, a prevalência do HPV foi associada ao início sexual precoce (P =0,001), a infecção pelo HIV–1 (P =0,01),a gestação (P =0,02) e a coilocitose no citopatológico (P =0,01). Sobre os genótipos, 90,4% eram de alto risco oncogênico (18 HPV–18, 14 HPV–16, quatro HPV–53, três HPV–31, dois HPV–58, dois HPV–59,dois HPV–68, um HPV–33 e um HPV–52) e 9,6% de baixo risco (dois HPV–11, dois HPV–16 e um HPV–70). Conclusões: Esse estudo teve a prevalência do HPV semelhante à prevalência descrita para esta região. Os genótipos do HPV de alto risco foram os mais prevalentes, sendo o HPV–18 o principal tipo viral encontrado...

Study design: cross-sectional. Objective: To determine the HPV prevalence and genotypes in women treated at University Hospital in southern Brazil. Methodology: Cervical cells samples from 200 women were collected. HPV was detected by nested polymerase chain reaction and genotypes were determined by sequencing. Variables were analyzed by the Fisher Exact Test and Chi-squared test of Pearson (X²)with a significance level of ≤ 5%. The strength of association was calculated by the prevalence ratio, with their confidence intervals at 95%. Multivariate analysis was calculated by Binary Logistic Regression for variables with P <0.20 Results: HPV DNA was detected in 55 women (27.5%). HPV prevalence was associate with income (P =0.01), early initiation of sexual life (P <0.001), pregnant (P = 0. 002), HIV-1 infection (P = 0. 001) and koilocytosis presence in cytological test (P =0.006). Were found an association between serological status for HIV-1 and the genotypes HPV–33 (P =0.001) and HPV–68 (P <0.001).Multivariate analysis showed that HPV prevalence was associated with patients who had early initiation of sexual life (P =0.001), was infected by HIV–1 (P = 0.01), was pregnant (P = 0.02), and women with koilocytosis in cytological test (P =0.01). Genotypes were 90.4% higher-risk oncogenic (18 HPV–18, 14HPV–16, four HPV–53, three HPV–31, two HPV–58, two HPV–59, two HPV–68, one HPV–33 and one HPV–52) and 9.6% low-risk (two HPV–11, two HPV–16 and one HPV–70). Conclusions: This study had the HPV prevalence similar to prevalence described in this region. The high-risk HPV genotypes were the most prevalent, being HPV–18 the main viral type found...