Impact of cervicovaginal microbiome on the risk of cervical abnormalities development.

The vaginal microbiome has emerged as potentially influencing the natural history of Human Papillomavirus (HPV) infections and their clinical impact. We aimed to characterize the vaginal microbiome in samples from 807 high-risk HPVs (Hr-HPV) positive women with a mean age of 41.45 ± 10.79 years who participated in the Regional Cervical Cancer Screening Program from the Northern Region of Portugal. Microbiome analysis was performed with commercial kits for the detection of 21 microorganisms. The most frequent microorganisms were Ureaplasma parvum (52.5%), Gardnerella vaginalis (GV) (34.5%), Atopobium vaginae (AV) (32.6%), Lacto (30.7%), and Mycoplasma hominis (MH) (23.5%). The distribution according to age reveals that MH, Mega1, GV, BVab2, AV, and Mob were more prevalent in women older than 41 years of age (p < 0.050), while Lacto is significantly decreased in this group (23.5% vs. 39.4%, p < 0.001; RR = 0.47). The risk analysis showed that Hr-HPV-16/-18 and Hr-HPV-9val genotypes are associated with an increased risk of developing cervical abnormalities, while Lacto (p < 0.001; odd ratio [OR] = 0.33), GV (p = 0.0111; OR = 0.41), AV (p = 0.033; OR = 0.53) and Mob (p = 0.022; OR = 0.29) are associated with protection. Similar results were found for the risk of development atypical squamous cells cannot exclude HSIL/high-grade squamous intraepithelial lesion. Overall, the multivariate analysis confirmed that lactobacillus and bacteria associated with bacterial vaginosis (GV, AV, and Mob) are associated with protection against the development of cervical abnormalities. This study provides important data to be included in the future management of risk stratification for Hr-HPV-positive women.

Impact of high-risk Human Papillomavirus genotyping in cervical disease in the Northern region of Portugal: Real-world data from regional cervical cancer screening program.

Cervical cancer prevention is based on primary prevention with vaccines against Human Papillomavirus (HPV) and secondary prevention by screening with High-Risk-HPV (Hr-HPV) detection. Since 2017, cervical cancer screening in women aged 25-60 years has been performed in Portugal using Hr-HPV detection, followed by cytology in Hr-HPV-positive cases. Herein we report the prevalence of Hr-HPV genotypes and cytological abnormalities among 462 401 women (mean age: 43.73 ± 10.79; median age: 45; range: 24-66 years) that participated in the Regional Cervical Cancer Screening Program of the Northern Region of Portugal, performed between August 2016 and December 2021. Overall, we describe a prevalence rate of 12.50% for Hr-HPV varying from 20.76% at age 25% to 8.32% at age 64. The five most common Hr-HPV genotypes identified were HPV-68 (16.09%), HPV-31 (15.30%), HPV-51 (12.96%), HPV-16 (11.06%), and HPV-39 (11.01%). The prevalence of Hr-HPV included in the nonavalent vaccine (HPV-9valent) was 55.00% ranging from 47.78% to 59.18% across different age groups. Considering positive Hr-HPV cases, 65.68% had a Negative for Intraepithelial Lesion or Malignancy (NILM) cytology, 20.83% atypical squamous cells of undetermined significance (ASC-US), 8.85% Low-Grade Squamous Intraepithelial Lesion (LSIL), 1.65% High-Grade Squamous Intraepithelial Lesion (HSIL), 2.85% ASC-H, 0.09% Atypical Glandular Cells, 0.02% Adenocarcinomas, and 0.02% Squamous Cell Carcinoma (SCC). Our analysis revealed that HPV-9val genotypes were responsible for 52.13% NILM, 59.21% ASC-US, 55.06% LSIL, 90.14% HSIL, 83.50% ASC-H, and 100.00% SCC. Furthermore, multiple Hr-HPV infections (risk ratio [RR] = 1.46; 95% confidence interval [CI] 1.34-1.58), HPV-16/18 (RR = 5.16; 95% CI 4.75-5.93), or HPV-9val genotypes (RR = 5.23; 95% CI 4.68-5.85) were associated with a significant risk of developing > HSIL (p < 0.001). To date, this is the largest study on Hr-HPV genotyping in cervical cancer screening that includes data from a complete cycle of the screening program. Our findings suggest a high prevalence of HPV-9valent genotypes and a significant association with an increased risk of developing > HSIL. This constitutes important data for health authorities, which may help define the future of vaccination and cervical cancer screening strategies.

Epstein-Barr virus strains and variations: Geographic or disease-specific variants?

The Epstein-Barr Virus (EBV) is associated with the development of several diseases, including infectious mononucleosis (IM), Burkitt's Lymphoma (BL), Nasopharyngeal Carcinoma, and other neoplasias. The publication of EBV genome 1984 led to several studies regarding the identification of different viral strains. Currently, EBV is divided into EBV type 1 (B95-8 strain) and EBV type 2 (AG876 strain), also known as type A and type B, which have been distinguished based upon genetic differences in the Epstein-Barr nuclear antigens (EBNAs) sequence. Several other EBV strains have been described in the past 10 years considering variations on EBV genome, and many have attempted to clarify if these variations are ethnic or geographically correlated, or if they are disease related. Indeed, there is an increasing interest to describe possible specific disease associations, with emphasis on different malignancies. These studies aim to clarify if these variations are ethnic or geographically correlated, or if they are disease related, thus being important to characterize the epidemiologic genetic distribution of EBV strains on our population. Here, we review the current knowledge on the different EBV strains and variants and its association with different diseases. J. Med. Virol. 89:373-387, 2017. © 2016 Wiley Periodicals, Inc.

Mechanisms underlying the association between obesity and Hodgkin lymphoma.

A solid body of knowledge indicates that overweight and obese subjects are prone to develop cancer, aggressive disease, and death more than their lean counterparts. While obesity has been causally associated with various cancers, only a limited number of studies beheld the link with classical Hodgkin lymphoma (HL). Contemporary meta-analysis and prospective studies confirmed the association of body mass index with HL. Besides epidemiological evidence, excess adiposity is known to influence tumor behavior through adipokines, adipose-derived stem cell migration, and metabolism regulation, and by modulating immunoinflammatory response. Nevertheless, the obesity paradox has been described in few cancers. Considering that adipose tissue is an immunomodulatory organ, and that inflammation is the cornerstone of HL pathophysiology, the rationale for being causally related due to endocrine/paracrine interactions cannot be negligible. In this hypothesis-generating review, we explore the biologically plausible links between excess adiposity and HL in light of recent basic and clinical data, in order to create a basis for understanding the underlying mechanisms and foster applied research. The establishment of an association of excess adiposity with HL will determine public health preventive measures to fight obesity and eventually novel therapeutic approaches in HL patients.

Characterization of Epstein-Barr virus strains and LMP1-deletion variants in Portugal.

Variations in the genome sequence of Epstein-Barr Virus (EBV) are thought to lead to differential interaction with host cells, immune evasion, and transformation. The discussion regarding EBV strains as having a geographic or disease-association has been increasing and the majority of studies are performed in Asiatic populations. We developed a case-control study with 139 individuals, including 96 subjects with different malignancies and 43 healthy individuals, from the North region of Portugal. We have used PCR protocols for the characterization of EBV strains (type A or B) based on EBNA3C genome variation and for the LMP1 30bp-deletion variants (wt-LMP1 or del-LMP1). Our study showed that type A is the most prevalent in our population (100% of healthy controls, 96.9% of aHSCT patients, 90.8% of HNSCC patients, and 94.9% of NPC patients) and that type B was significantly associated with NPC (P = 0.019; RR = 8.90). Regarding the LMP1 30bp-deletion, we found a similar distribution of both wt- and del-LMP1 variants in controls and dispare results in cases: del-LMP1 was more frequent in aHSCT and HNSCC patients (64.7% and 63.2%, respectively) and wt-LMP1 in NPC patients (100%). In fact, the study reveals that wt-LPM1 was significantly associated with NPC (P < 0.001; RR = 18.4). Hence, our study showed that EBV type B and wt-LMP1 variant seem to be associated with NPC in our population, with a clear disease-association for wt-LMP1. These results contribute for the knowledge of EBV genetic diversity among Caucasian populations.

Characterization of cytomegalovirus and epstein-barr virus infection in cervical lesions in Portugal.

Infection by high-risk types of human papillomavirus (HPV) is considered necessary but not sufficient for the development of cervical cancer. Previous studies suggested that cytomegalovirus (CMV) and Epstein-barr virus (EBV) could be co-factors of HPV-associated carcinogenesis. The aim of this study was to characterize the prevalence of CMV and EBV and evaluate its association with the development cervical lesions in Portugal. The prevalence of CMV and EBV infections was determined by real-time PCR in 89 cervical samples from women with different histological lesions, who attended the Portuguese Institute of Oncology of Porto. This study revealed an overall prevalence of 4.5% for CMV and 10.1% for EBV. Age-stratified analysis revealed that CMV infection was present in individuals <30 and >60 years old, while EBV infection was present in all age groups. CMV was detected in 9.5% of low-grade lesions and in 22.2% of in situ/invasive carcinomas, while EBV infection was found in all different types of lesions. In addition, data revealed that CMV infection was associated with an increased risk of in situ/invasive carcinoma development (OR=1.28; P=0.035). The study reveals a low prevalence for both viruses; nevertheless, these results are important for knowledge on the shedding of EBV and CMV in cervical samples.

Association of Epstein­Barr virus infection with allogeneic hematopoietic stem cell transplantation in patients in Portugal.

The identification of patients at higher risk of developing Epstein­Barr virus (EBV) infection in hematopoietic stem cell transplants (HSCT) is useful for the prevention of EBV­associated diseases A prospective observational study was developed that included 40 patients (27 male and 13 females, with mean age of 32.2±1.5 years old) undergoing allogeneic­HSCT between January and December 2015. EBV was examined in whole blood samples collected during routine procedures at day (D)+30, D +60, +90, D+120, D+150 and D+180 post­transplant. EBV was detected, at least once during the follow­up period in 70.0% of our patients. Results indicated that patients with unrelated donors had increased risk of developing EBV infection at D+60 and D+150 (OR=3.9, P=0.058; OR=8.0, P=0.043; respectively). Moreover, myeloablative conditioning (OR=4.3, P=0.052), anti­thymocyte globulin use (OR=12.0, P=0.030) and graft­vs.­host disease (OR=6.7, P=0.032) were associated with EBV infection at D+60, D+150 and D+90, respectively. In our series, none of these patients developed post­transplant lymphoproliferative disease. To the best of our knowledge, the present study is the first study to report EBV infection in patients undergoing aHSCT from Portugal. The study revealed that EBV infection is associated with different factors. These findings provide evidence towards the identification of high­risk patients for EBV­infection and associated disease.

High-Risk human papillomavirus genotype distribution in the Northern region of Portugal: Data from regional cervical cancer screening program.

High-Risk Human papillomavirus (HR-HPV) full genotyping methods have been described as of great potential use in epidemiology and preventive strategies, including cervical cancer screening and HPV vaccination. We characterized the prevalence and distribution of HR-HPV genotypes in cervico-vaginal samples obtained from the Regional Cervical Cancer Screening Program from the Northern Region of Portugal. HR-HPV genotyping was performed using Anyplex™ II HPV-HR Detection kit in 105,458 women enrolled between August 2016 and December 2017. HR-HPVs were detected in 10,665 women (10.2%) with a prevalence ranging from 6.2 to 17.1% depending on age, and from 8.7 to 10.7% depending on geographical location. Multiple infections with two or more HR-HPVs were detected in 2736 (25.7%) of HR-HPV women ranging from 16.5 to 31.0% depending on age. Amongst HR-HPV positive women, HPV-16 (17.5%), HPV-39 (16.7%), HPV-31 (15.0%), HPV-68 (13.2%), HPV-52 (10.7%) and HPV-51 (10.6%) were the most common genotypes in our population, being HPV-16 more frequent in women aged from 30 to 45 years and HPV-39 in 50-65 years. Results also show that HPV16/18 are present in 22.1% and HPV16/18/31/33/45/52/58 in 47.6% of HR-HPV positive women. This is the largest study on HR-HPV genotyping for Cervical Cancer Screening in European populations and provides critical data for program management and vaccine policy.

Post­transplant lymphoproliferative disorder in hematopoietic stem cell transplant patients: A single center retrospective study between 2005 and 2012.

Post­transplant lymphoproliferative disorder (PTLD), despite its rarity, is an important mortality/morbidity event in transplant patients. The purpose of the present study was to retrospectively examine the clinical and pathologic characteristics, and outcomes of PTLD at the Portuguese Oncology Institute of Porto. A retrospective review of patient information was performed for patients that developed PTLD following allogeneic hematopoietic stem cell transplant (aHSCT) and were diagnosed between 2005 and 2012. The present study included a total of 15 patients, 8 females (53.3%) and 7 males (46.7%), with different clinicopathological characteristics. The most frequent clinical condition inducing aHSCT was acute lymphocytic leukemia (40.0%). Conditioning regimens consisted primarily in busulfan and cyclophosphamide, with anti­thymocyte globulin, and myeloablation was the preferential treatment. Epstein­Barr virus (EBV) was present in all patients with a median time of diagnosis following transplant of 75 days (range, 25­485 days) and a median viral load of 4.75 log10 copies/ml (range, 3.30­6.26 log10 copies/ml). PTLD diagnosis was mainly assessed by clinical findings, and histological confirmation was available for 5 patients: 3 monomorphic, 1 polymorphic and 1 with early lesions of PTLD. To the best of our knowledge, this is the first study to describe PTLD cases in HSCT patients in Portugal. The data reinforces the importance of performing EBV monitoring in high­risk patients, particularly those receiving a transplant from mismatch/unrelated donors, and those with myeloablative conditioning regimen including antithymocyte globulin. The results also suggested that EBV viral load may be significant for the prediction of PTLD development.

miR-34a and miR-125b Expression in HPV Infection and Cervical Cancer Development.

We aimed to characterize miR-125b and miR-34a expression in 114 women with different cervical lesions: normal epithelium with (n = 20) and without (n = 29) HPV infection; LSIL (n = 28); HSIL (n = 29); and ICC (n = 8). miRNA expression analysis was performed by comparing the distinct groups with the reference group (women with normal epithelium without HPV). For miR-125b, we observed a twofold (2(-ΔΔCt) = 2.11; P = 0.038) increased expression among women with normal epithelium with HPV infection and a trend of downregulation in different cervical lesions including an 80% reduction (2(-ΔΔCt) = 0.21; P = 0.004) in ICC. Similarly, miR-34a expression analysis revealed an increased expression (2(-ΔΔCt) = 1.69; P = 0.049) among women with normal cervix and HPV infection, and despite no significant correlation with cervical lesions, its expression was increased by twofold (2(-ΔΔCt) = 2.08; P = 0.042) in ICC. Moreover, miR-125b levels were able to predict invasive cancers with 88% sensitivity and 69% specificity. Results showed that while miR-34a expression seems to be correlated with invasive cervical cancer, miR-125b expression is significantly changed within the different cervical lesions and their levels should be further investigated as possible predictive/prognostic biomarkers using a noninvasive approach.

Characterization of human papillomavirus genotypes and HPV-16 physical status in cervical neoplasias of women from northern Portugal.

OBJECTIVE: To determine human papillomavirus (HPV) genotypes and the physical status of HPV-16 DNA among women from northern Portugal with cervical lesions. METHODS: The present retrospective study included samples of cervical exfoliated cells from 88 women (median age 42.0±13.1 years) who attended the Gynecology Service at the Portuguese Institute of Oncology in Porto during 2010. After DNA extraction, HPV genotyping was performed by polymerase chain reaction (PCR) followed by restriction fragment length polymorphism analysis using the MY09/MY11 primers. The physical status of HPV-16 was determined by real-time PCR. RESULTS: Overall, 69.3% of the samples tested positive. The prevalence of HPV infection was 38.5% in normal samples, 57.7% in cervicitis samples, and 87.2% in all cervical lesions including invasive cancers. Sixteen genotypes were detected, the most prevalent ones being HPV-16 (42.9%), HPV-31 (12.2%), and HPV-58 (10.2%); HPV-18 was rare. The overall prevalence of HPV-16 integration was 31.6%. The physical status of HPV-16 did not differ significantly by histology. CONCLUSION: The most frequent genotypes were HPV-16, -31, and -58. Integration of HPV-16 DNA seemed to be an early event in cervical carcinogenesis. Further studies are required to clarify the value of viral integration as a prognostic marker.

Cytomegalovirus infection and cervical cancer: from past doubts to present questions.

BACKGROUND: Since early 60's that Cytomegalovirus was studied for its possible role in cervical cancer development. Despite several decades of studies and the description of CMV DNA in cervical samples, it is still doubtful what is the prevalence of Cytomegalovirus in cervix and if CMV can act as a co-factor in cervical carcinogenesis. METHODS: In this Systematic Review we intend to summarize the frequency of Cytomegalovirus in cervical samples by revising all published studies between 1980 and 2011 regarding the detection of Cytomegalovirus in cervical samples and the development of lesions/ invasive cervical cancer. Crude and adjusted frequencies of Cytomegalovirus infection were calculated according to country and world region. RESULTS: This study revealed that the worldwide crude frequency of Cytomegalovirus infection in the cervix was 18.9% in all cervical samples and 36.5% in HPV positive women. Cytomegalovirus infection was present in all different types of lesions: 17.4% in normal/ cervicitis, 28.0% in LSIL, 19.7% in HSIL and 44.4% in CIS/ICC. The overall rate of Cytomegalovirus infection varied from 1.58% to 61.0% with an increased incidence in less developed countries. CONCLUSION: In this study we described a high frequency of positive Cytomegalovirus cases in all types of cervical samples, with increased incidence in both HPV-infected women and CIS/ICC cases. Hence, despite results showed that Cytomegalovirus shedding in cervical samples is frequent more studies should be performed to clarify if Cytomegalovirus infection is an opportunistic infection in HPV-infected cases, or if it contributes for cervical immunosuppression that will favor HPV-associated carcinogenesis.

Introdução: Desde os anos 60 que o Citomegalovírus tem sido estudado pelo potencial papel no desenvolvimento de cancro do colo do útero. Após várias décadas de estudos e relatos da presença de DNA viral em amostras cervico-vaginais, ainda permanecem dúvidas sobre qual a prevalência do Citomegalovírus no cólo do útero e se este pode ser um cofator da carcinogénese. Métodos: Nesta Revisão Sistemática pretendemos descrever a prevalência de Citomegalovírus em amostras cervicais, revendo todos os estudos publicados entre 1980 e 2011 que descrevem a infecção por Citomegalovírus em amostras cervicais e o desenvolvimento de lesões/carcinoma invasor. As frequências da infecção por Citomegalovírus foram calculadas de acordo com a região no mundo. Resultados: Este estudo revelou que a frequência bruta da infecção por Citomegalovírus no cólo do útero foi de 18,9% em todas as amostras e 36,5% em mulheres HPV positivas. A infecção por Citomegalovírus está presente em todas as diferentes lesões: 17,4% em normais/cervicite, 28,0% em LSIL, 19,7% em HSIL e 44,4% em CIS/ICC. A frequência global variou de 1,58% a 61,0%, com uma maior incidência em países menos desenvolvidos. Conclusão: Neste estudo, verificou-se uma elevada frequência de casos positivos para Citomegalovírus em todos os tipos de amostras cervico-vaginais, com maior incidência em mulheres infectadas por HPV e em casos de cancro. Assim, são necessários mais estudos para esclarecer se a infecção por Citomegalovírus é uma infecção oportunista ou se contribui para a imunossupressão favorecendo a carcinogénese associada ao HPV.