Molecular Signatures of High-Grade Cervical Lesions.

Cervical cancer is the fourth most common neoplasia in women and the infection with human papilloma virus (HPV) is its necessary cause. Screening methods, currently based on cytology and HPV DNA tests, display low specificity/sensitivity, reducing the efficacy of cervical cancer screening programs. Herein, molecular signatures of cervical cytologic specimens revealed by liquid chromatography-mass spectrometry (LC-MS), were tested in their ability to provide a metabolomic screening for cervical cancer. These molecules were tested whether they could clinically differentiate insignificant HPV infections from precancerous lesions. For that, high-grade squamous intraepithelial lesions (HSIL)-related metabolites were compared to those of no cervical lesions in women with and without HPV infection. Samples were collected from women diagnosed with normal cervix (N = 40) and from those detected with HSIL from cytology and colposcopy (N = 40). Liquid-based cytology diagnosis, DNA HPV-detection test, and LC-MS analysis were carried out for all the samples. The same sample, in a customized collection medium, could be used for all the diagnostic techniques employed here. The metabolomic profile of cervical cancer provided by LC-MS was found to indicate unique molecular signatures for HSIL, being two ceramides and a sphingosine metabolite. These molecules occurred independently of women's HPV status and could be related to the pre-neoplastic phenotype. Statistical models based on such findings could correctly discriminate and classify HSIL and no cervical lesion women. The results showcase the potential of LC-MS as an emerging technology for clinical use in cervical cancer screening, although further validation with a larger sample set is still necessary.

Preoperative measurement of serum C-reactive protein: is it useful in the differential diagnosis of adnexal masses?

BACKGROUND: Serum biomarkers may help to discriminate malignant from benign adnexal masses with equivocal features on imaging. Adequate discrimination of such tumors is crucial for referring patients to either a specialized cancer center or a nonspecialized gynecology service. AIM: We aimed to investigate whether the preoperative level of serum C-reactive protein (CRP), alone or combined with CA125 and menopausal status in the Ovarian Score (OVS), is useful in the prediction of malignancy in women with ovarian tumors. METHODS: This cross-sectional study included 293 patients who underwent surgery in a tertiary cancer center. Receiver operating characteristic (ROC) areas under the curves (AUC) for CRP, CA125 and OVS were calculated in different scenarios, as well as their sensitivity and specificity, using standard cutoff points (for CRP, 10 mg/L; for CA125, 35 U/mL). RESULTS: CA125 and the OVS performed significantly better than CRP alone in the differentiation of benign disease from epithelial ovarian cancer (EOC) (AUC = 0.86 for CA125, 0.79 for OVS, and 0.73 for CRP). OVS and CRP alone were superior to CA125 only in the differentiation of borderline ovarian tumors from advanced stages of EOC and non-EOC. Sensitivity and specificity were 52.5% and 83%, respectively, for CRP, 77.9% and 66.7% for CA125, and 71.3% and 67.8% for OVS. CONCLUSIONS: OVS is as good as CA125 in the differentiation of benign tumors from ovarian cancer. The addition of CA125 and menopausal status to CRP enhanced the relatively low discriminatory power of isolated CRP.

Comparing the Copenhagen Index (CPH-I) and Risk of Ovarian Malignancy Algorithm (ROMA): Two equivalent ways to differentiate malignant from benign ovarian tumors before surgery?

AIM: To evaluate the prediction of malignancy in women with pelvic masses using the Copenhagen Index (CPH-I) and Risk of Ovarian Malignancy Algorithm (ROMA). PATIENTS AND METHODS: Three hundred eighty four women operated due to an ovarian mass were enrolled between January 2010 and June 2015. All patients had histopathological diagnosis, HE4 and CA125 measurement. CPH-I and ROMA were calculated and their performances compared in two distinct scenarios: 1) for the discrimination of benign ovarian disease from epithelial ovarian cancer (EOC), non-epithelial ovarian cancer, borderline ovarian tumors (BOT) and ovarian metastases, and 2) for the discrimination of benign disease from EOC. Receiver Operator Characteristics' Areas Under the Curves (AUC) were calculated for CPH-I and ROMA and compared. RESULTS: Of the 384 women, 224 presented a benign ovarian tumor, 32 BOT, 87 EOC, 26 non-epithelial ovarian cancer, and 15 had ovarian metastases. The best AUCs were obtained for the discrimination of EOC from benign tumors. CPH-I performed slightly better than ROMA, and both approached 89% sensitivity and 85% specificity. When all malignant tumors (EOC, BOT, ovarian metastases and non-epithelial ovarian cancer - entire cohort) were included, the performance of CPH-I and ROMA declined to nearly 72%, although the specificity remained close to 85%. CONCLUSION: CPH-I and ROMA performed similarly well for the discrimination of EOC from benign ovarian tumors. However, caution is necessary since, in practical situations, where all the histological possibilities for malignant ovarian tumors must be considered, the sensitivity of CPH-I and ROMA may not surpass 70%.

Bacterial vaginosis and inflammatory response showed association with severity of cervical neoplasia in HPV-positive women.

Vaginal infections may affect susceptibility to and clearance of human papillomavirus (HPV) infection and chronic inflammation has been linked to carcinogenesis. This study aimed to evaluate the association between bacterial vaginosis (BV) and inflammatory response (IR) with the severity of cervical neoplasia in HPV-infected women. HPV DNA was amplified using PGMY09/11 primers and genotyping was performed using a reverse line blot hybridization assay in 211 cervical samples from women submitted to excision of the transformation zone. The bacterial flora was assessed in Papanicolaou stained smears, and positivity for BV was defined as ≥ 20% of clue cells. Present inflammatory response was defined as ≥ 30 neutrophils per field at 1000× magnification. Age higher than 29 years (OR:1.91 95% CI 1.06-3.45), infections by the types 16 and/or 18 (OR:1.92 95% CI 1.06-3.47), single or multiple infections associated with types 16 and/or 18 (OR: 1.92 CI 95% 1.06-3.47), BV (OR: 3.54 95% CI 1.62-7.73) and IR (OR: 6.33 95% CI 3.06-13.07) were associated with severity of cervical neoplasia (CIN 2 or worse diagnoses), while not smoking showed a protective effect (OR: 0.51 95% CI 0.26-0.98). After controlling for confounding factors, BV(OR: 3.90 95% CI 1.64-9.29) and IR (OR: 6.43 95% CI 2.92-14.15) maintained their association with the severity of cervical neoplasia. Bacterial vaginosis and inflammatory response were independently associated with severity of cervical neoplasia in HPV-positive women, which seems to suggest that the microenvironment would relate to the natural history of cervical neoplasia.

Performance of the Risk of Malignancy Index for Discriminating Malignant Tumors in Women With Adnexal Masses.

OBJECTIVES: We examined the performance of 4 risk of malignancy index (RMI) variants in a medium-resource gynecologic cancer center. METHODS: A total of 158 women referred for adnexal masses were evaluated before surgery by the 4 RMI variants. Physicians with varied experience in ultrasound assessment of adnexal masses performed ultrasound examinations. We compared the performance of the 4 RMI variants using receiver operating characteristic curve analyses followed by calculation of sensitivity, specificity, and positive and negative likelihood ratios using the pathologic diagnosis of the masses as the reference standard. RESULTS: Among the 158 women with adnexal masses included in this study, 51 (32%) had malignant tumors; 26 (51%) of them were stage I. All RMI variants performed similarly (accuracy range, 74%-83%), regardless of menopausal status. Considering all women included, the positive likelihood ratios of the 4 RMI variants ranged from 3.52 to 4.41. In subset analyses, all RMI variants had decreased sensitivity for stage I malignant tumors and for those of nonepithelial histologic types. CONCLUSIONS: The 4 RMI variants performed acceptably in a medium-resource setting where ultrasound examiners were physicians with varied experience. This finding indicates a good tradeoff between performance and feasibility, since ultrasound RMI protocols are of low complexity.

DNA extraction from filter-paper spots of vaginal samples collected after sexual violence.

OBJECTIVE: To detect the presence of male DNA in vaginal samples collected from survivors of sexual violence and stored on filter paper. METHODS: A pilot study was conducted to evaluate 10 vaginal samples spotted on sterile filter paper: 6 collected at random in April 2009 and 4 in October 2010. Time between sexual assault and sample collection was 4-48hours. After drying at room temperature, the samples were placed in a sterile envelope and stored for 2-3years until processing. DNA extraction was confirmed by polymerase chain reaction for human ß-globin, and the presence of prostate-specific antigen (PSA) was quantified. The presence of the Y chromosome was detected using primers for sequences in the TSPY (Y7/Y8 and DYS14) and SRY genes. RESULTS: ß-Globin was detected in all 10 samples, while 2 samples were positive for PSA. Half of the samples amplified the Y7/Y8 and DYS14 sequences of the TSPY gene and 30% amplified the SRY gene sequence of the Y chromosome. Four male samples and 1 female sample served as controls. CONCLUSION: Filter-paper spots stored for periods of up to 3years proved adequate for preserving genetic material from vaginal samples collected following sexual violence.

Retrieval of HPV oncogenes E6 and E7 mRNA from cervical specimens using a manual open technology protocol.

BACKGROUND: HPV oncogenes mRNA detection gains momentum as an adjuvant for HPV-related cervical abnormalities diagnosis, but is based on costly detection assays not allowing viral type targeting. OBJECTIVE: To assess detection rate of HPV oncogenes E6/E7 mRNA from cervical specimens using a manual, open technology, fully customizable protocol and determine whether HPV-related epidemiological features influence mRNA retrieval. We reviewed literature and compared our retrieval rate with automated technologies. METHODS: We used 60 samples positive for HPV DNA types 16, 18, 31 and/or 45. We extracted mRNA with a TRizol-based protocol, and tested mRNA purity and concentration using light absorbance. We reverse-transcribed mRNA into cDNA for E6/7 detection. RESULTS: HPV oncogenes E6/E7 mRNA was retrieved from 36 (60%) out of 60 specimens. No HPV load-related clinical or epidemiological feature was significantly associated with mRNA retrieval. Presence of HPV-DNA 16/18 was associated with mRNA retrieval (OR = 9.08; 95% CI 1.26 to 65.32 for HPV 16; and 18.2; IC95% 1.86 to 391.44 for HPV 18). CONCLUSIONS: The open-technology protocol yielded an mRNA detection rate similar to that of automated technologies. Advantages are lower costs and target HPV type customization.

Symptoms, CA125 and HE4 for the preoperative prediction of ovarian malignancy in Brazilian women with ovarian masses.

BACKGROUND: This manuscript evaluates whether specific symptoms, a symptom index (SI), CA125 and HE4 can help identify women with malignant tumors in the group of women with adnexal masses previously diagnosed with ultrasound. METHODS: This was a cross-sectional study with data collection between January 2010 and January 2012. We invited 176 women with adnexal masses of suspected ovarian origin, attending the hospital of the Department of Obstetrics and Gynecology of the Unicamp School of Medicine. A control group of 150 healthy women was also enrolled. Symptoms were assessed with a questionnaire tested previously. Women with adnexal masses were interviewed before surgery to avoid recall bias. The Ward Agglomerative Method was used to define symptom clusters. Serum measurements of CA125 and HE4 were made. The Risk of Ovarian Malignancy Algorithm (ROMA) was calculated using standard formulae. RESULTS: Sixty women had ovarian cancer and 116 benign ovarian tumors. Six symptom clusters were formed and three specific symptoms (back pain, leg swelling and able to feel abdominal mass) did not agglomerate. A symptom index (SI) using clusters abdomen, pain and eating was formed. The sensitivity of the SI in discriminating women with malignant from those with benign ovarian tumors was 78.3%, with a specificity of 60.3%. Positive SI was more frequent in women with malignant than in women with benign tumors (OR 5.5; 95% CI 2.7 to 11.3). Elevated CA125 (OR 11.8; 95% CI 5.6 to 24.6) or HE4 (OR 7.6; 95% CI 3.7 to 15.6) or positive ROMA (OR 9.5; 95% CI 4.4 to 20.3) were found in women with malignant tumors compared with women with benign tumors. The AUC-ROC for CA125 was not different from that for HE4 or ROMA. The best specificity and negative predictive values were obtained using CA125 in women with negative SI. CONCLUSION: Women diagnosed with an adnexal mass could benefit from a short enquiry about presence, frequency and onset of six symptoms, and CA125 measurements. Primary care physicians can be thereby assisted in deciding as to whether or not reference the woman to often busy, congested specialized oncology centers.

HE4 can help discriminate women with malignant ovarian tumors only if CA125 levels are elevated.

Objective: To evaluate the performance of mesothelin, CA125, HE4 and ROMA index in the detection of malignant ovarian tumors. 
Study Design: This is a cross-sectional study including 199 women with adnexal masses (67 with malignant tumors and 132 with benign tumors) and 150 healthy women (controls). We used a recursive multivariate partitioning algorithm to assess the contribution of each of the serum markers and the ROMA index to the discrimination of women with malignant tumors. Also, the areas under the ROC curves (AUC) for each of the serum markers and for the ROMA index were compared using the Venkatraman's projection-permutation test. 
Results: In the recursive model, CA125 had the best performance in discriminating women with malignant tumors. In women with normal-range CA125 levels, none of the other markers contributed to the discrimination of women with malignant tumors. In women with elevated CA125 levels, elevated HE4 levels were associated with an increased risk of harboring a malignant tumor. The AUC for mesothelin was smaller than that for all the other curves, and both the ROMA index and CA125 had better AUC than HE4. 
 Conclusion: In women with normal CA125 levels, neither mesothelin nor HE4 contributed to discriminate women with malignant ovarian tumors; however, for women with elevated CA125 levels HE4 may help discriminating those who have a malignant ovarian tumor.

[Prevalence of Chlamydia trachomatis infection among women candidates for in vitro fertilization at a public institution of the State of São Paulo, Brazil]. / Prevalência de infecção por Chlamydia trachomatis em mulheres candidatas à fertilização in vitro em serviço público de referência do Estado de São Paulo.

PURPOSE: To evaluate the prevalence of Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) among women candidates to in vitro fertilization (IVF) in a reference public service in southeastern Brazil. METHODS: Women who were referred for IVF from April 1st, 2008 to December 31st, 2009 were enrolled sequentially in the study. A gynecological-obstetrical background questionnaire was applied and endocervical swab samples were obtained to search for CT and NG using hybrid capture and PCR. The variables studied were: age, color, education, duration of infertility, number of pregnancies and living children, history of miscarriage, ectopic pregnancy, number of sex partners, pelvic inflammatory disease (PID), pelvic surgery, manipulation of the uterine cavity, smoking, and illicit drug use. The women were distributed according to the presence/absence of confirmed chlamydia infection and descriptive analysis was employed. RESULTS: Among 176 women tested the prevalence of CT infection was 1.1% and there was no NG infection. Two thirds of the women were ≥30 years old, with schooling ≥8 years and ≤5 years of infertility, and 56.2% had no children. The main background data were pelvic surgery (77.8%), manipulation of the uterine cavity (62.5%) and PID (27.8%). The tubal factor was the most prevalent, 73.3% of women (from 129), 37.5% had been sterilized, 35.8% had not been sterilized, and other factors had a prevalence <30%. CONCLUSIONS: CT and NG infections had a low prevalence in this sample. Studies at other centers in the country are needed to confirm the prevalence of infection in this particular group of infertile women.

Inclusão dos sintomas na discriminação entre tumores anexiais benignos e malignos / Inclusion of symptoms in the discrimination between benign and malignant adnexal masses

OBJETIVO: Avaliar a associação entre sintomas clínicos e malignidade em mulheres com tumores anexiais, submetidas à cirurgia. MÉTODOS: Estudo de corte transversal com coleta prospectiva, no qual foram incluídas 105 mulheres, atendidas em um hospital de ensino do Estado de São Paulo de novembro de 2009 a março de 2011, devido ao tumor anexial e à indicação de laparotomia/laparoscopia. Todas foram submetidas a uma entrevista estruturada sobre a ocorrência de 18 sintomas associados ao câncer de ovário. A entrevista incluiu gravidade, frequência e duração dos sintomas nos 12 meses prévios à primeira consulta. Também foram avaliados os níveis de CA125 e a classificação ultrassonográfica. Foi calculada para cada sintoma a razão de prevalência com intervalo de confiança de 95%. O padrão-ouro foi o resultado do exame anatomopatológico das peças cirúrgicas. RESULTADOS: Das 105 mulheres incluídas, 75 (71,4%) apresentaram tumores benignos e 30 (28,6%), malignos. Em mulheres com tumores malignos, os sintomas foram mais frequentes, dentre eles: inchaço abdominal (70%), aumento do volume abdominal (67%), dor pélvica (60%), irregularidade menstrual (60%), empachamento (53%), dor abdominal (50%), dor nas costas (50%) e saciedade precoce (50%). As mulheres com tumores benignos apresentaram essencialmente dor pélvica (61%), irregularidade menstrual (61%) e inchaço abdominal (47%). Os sintomas significativamente associados com malignidade foram: sensação de inchaço abdominal (RP=2,0; IC95% 1,01 - 3,94), aumento objetivo do volume abdominal (RP=2,16; IC95% 1,12 - 4,16), dor nas costas (RP=1,97; IC95% 1,09 - 3,55), empachamento (RP=2,25; IC95% 1,25 - 4,07), saciedade precoce (RP=2,06; IC95% 1,14 - 3,70), massa abdominal (RP=1,83; IC95% 1,01 - 3,30), dificuldade para deglutir (RP=1,98; IC95% 1,10 - 3,56) e sangramento pós-menopausa (RP=2,91; IC95% 1,55 - 5,44). A presença de dor pélvica, constipação, dispareunia, fadiga, dor abdominal, náusea e/ou vômito, irregularidade menstrual, perda de peso, diarreia e sinusorragia foram semelhantes nos dois grupos. CONCLUSÕES: Em mulheres com tumores anexiais com indicação cirúrgica, a avaliação pré-operatória dos sintomas pode auxiliar na predição da malignidade.

PURPOSE: To assess the association between clinical symptoms and the diagnosis of malignancy in women with adnexal tumors who underwent surgery. METHODS: Cross-sectional study, in which 105 women with adnexal tumors and indication for laparotomy/laparoscopy were included. All women were treated at a teaching hospital in the state of São Paulo between November 2009 and March 2011. All patients underwent a structured interview about the occurrence of 18 symptoms associated with ovarian cancer. The interview included the severity, frequency, and duration of these symptoms in the 12 months prior to the first medical consultation. The CA125 levels and the ultrasound classification of the tumors were also evaluated. We calculated for each symptom the prevalence ratio with 95% confidence intervals. The golden-standard was the result of the pathological examination of the surgical specimens. RESULTS: Of the 105 women included, 75 (71.4%) had benign tumors and 30 (28.6%) had malignant ones. In women with malignant tumors, the most frequent symptoms were: abdominal bloating (70%), increased abdominal size (67%), pelvic pain (60%), menstrual irregularity (60%), swelling (53%), abdominal pain (50%), backache (50%), and early repletion (50%). Women with benign tumors showed essentially pelvic pain (61%), menstrual irregularities (61%), and abdominal swelling (47%). Symptoms significantly associated with malignancy were: bloating (PR=2.0; 95%CI 1.01 - 3.94), increased abdominal size (PR=2.16; 95%CI 1.12 - 4.16), backache (RP=1.97; 95%CI 1.09 - 3.55), swelling (PR=2.25; 95%CI 1.25 - 4.07), early repletion (RP=2.06; 95%CI 1.14 - 3.70), abdominal mass (PR=1.83; 95%CI 1.01 - 3.30), eating difficulties (PR=1.98; 95%CI 1.10 - 3.56), and postmenopausal bleeding (PR=2.91; 95%CI 1.55 - 5.44). The presence of pelvic pain, constipation, dyspareunia, fatigue, abdominal pain, nausea or vomiting, menstrual irregularity, weight loss, diarrhea, and bleeding after intercourse was similar in both groups. CONCLUSIONS: In women with adnexal tumors including indication of surgical treatment, the preoperative evaluation of symptoms may help predicting malignancy.

Prevalência de infecção por Chlamydia trachomatis em mulheres candidatas à fertilização in vitro em serviço público de referência do Estado de São Paulo / Prevalence of Chlamydia trachomatis infection among women candidates for in vitro fertilization at a public institution of the State of São Paulo, Brazil

OBJETIVO: Avaliar a prevalência de infecção por Chlamydia trachomatis (CT) e Neisseria gonorrhoeae (NG) em mulheres candidatas ao tratamento tópico e de fertilização in vitro (FIV) em serviço público de referência da Região Sudeste do Brasil. MÉTODOS: Mulheres que tiveram indicação de FIV, no período de 1º de abril de 2008 a 31 de outubro de 2009, foram admitidas sequencialmente no estudo. Foi aplicado um questionário sobre antecedentes ginecológicos e obstétricos e coletada amostra de swab endocervical para pesquisa de CT e NG através de captura híbrida e PCR. As variáveis estudadas foram: faixa etária, cor, escolaridade, tempo de infertilidade, número de gestações e filhos vivos, antecedentes de aborto, gestação ectópica, número de parceiros, Doença Inflamatória Pélvica (DIP), cirurgia pélvica, manipulação de cavidade uterina, tabagismo e uso de drogas ilícitas. As mulheres foram distribuídas segundo presença ou não de infecção por clamídia e a análise foi descritiva. RESULTADOS: Entre as 176 mulheres estudadas a prevalência de infecção por CT foi de 1,1%, não houve infecção por NG. Dois terços das mulheres tinham idade >30 anos, escolaridade >8 anos, <5 anos de infertilidade e 56,2% não tinham filhos. Os principais antecedentes foram cirurgia pélvica (77,8%), manipulação de cavidade uterina (62,5%) e DIP (27,8%). O fator tubário foi o mais prevalente, em 129 mulheres (73,3%), 37,5% com e 35,8% sem laqueadura, os demais fatores tiveram prevalência <30%. CONCLUSÕES: As infecções por CT e NG tiveram baixa prevalência na amostra estudada e são necessários estudos em outros centros do país para confirmar a prevalência de infecções nesse grupo particular de mulheres inférteis.

PURPOSE: To evaluate the prevalence of Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) among women candidates to in vitro fertilization (IVF) in a reference public service in southeastern Brazil. METHODS: Women who were referred for IVF from April 1st, 2008 to December 31st, 2009 were enrolled sequentially in the study. A ginecological-obstetrical background questionnaire was applied and endocervical swab samples were obtained to search for CT and NG using hybrid capture and PCR. The variables studied were: age, color, education, duration of infertility, number of pregnancies and living children, history of miscarriage, ectopic pregnancy, number of sex partners, pelvic inflammatory disease (PID), pelvic surgery, manipulation of the uterine cavity, smoking, and illicit drug use. The women were distributed according to the presence/absence of confirmed chlamydia infection and descriptive analysis was employed. RESULTS: Among 176 women tested the prevalence of CT infection was 1.1% and there was no NG infection. Two thirds of the women were >30 years old, with schooling >8 years and <5 years of infertility, and 56.2% had no children. The main background data were pelvic surgery (77.8%), manipulation of the uterine cavity (62.5%) and PID (27.8%). The tubal factor was the most prevalent, 73.3% of women (from 129), 37.5% had been sterilized, 35.8% had not been sterilized, and other factors had a prevalence <30%. CONCLUSIONS: CT and NG infections had a low prevalence in this sample. Studies at other centers in the country are needed to confirm the prevalence of infection in this particular group of infertile women.

[Inclusion of symptoms in the discrimination between benign and malignant adnexal masses]. / Inclusão dos sintomas na discriminação entre tumores anexiais benignos e malignos.

PURPOSE: To assess the association between clinical symptoms and the diagnosis of malignancy in women with adnexal tumors who underwent surgery. METHODS: Cross-sectional study, in which 105 women with adnexal tumors and indication for laparotomy/laparoscopy were included. All women were treated at a teaching hospital in the state of São Paulo between November 2009 and March 2011. All patients underwent a structured interview about the occurrence of 18 symptoms associated with ovarian cancer. The interview included the severity, frequency, and duration of these symptoms in the 12 months prior to the first medical consultation. The CA125 levels and the ultrasound classification of the tumors were also evaluated. We calculated for each symptom the prevalence ratio with 95% confidence intervals. The golden-standard was the result of the pathological examination of the surgical specimens. RESULTS: Of the 105 women included, 75 (71.4%) had benign tumors and 30 (28.6%) had malignant ones. In women with malignant tumors, the most frequent symptoms were: abdominal bloating (70%), increased abdominal size (67%), pelvic pain (60%), menstrual irregularity (60%), swelling (53%), abdominal pain (50%), backache (50%), and early repletion (50%). Women with benign tumors showed essentially pelvic pain (61%), menstrual irregularities (61%), and abdominal swelling (47%). Symptoms significantly associated with malignancy were: bloating (PR=2.0; 95%CI 1.01 - 3.94), increased abdominal size (PR=2.16; 95%CI 1.12 - 4.16), backache (RP=1.97; 95%CI 1.09 - 3.55), swelling (PR=2.25; 95%CI 1.25 - 4.07), early repletion (RP=2.06; 95%CI 1.14 - 3.70), abdominal mass (PR=1.83; 95%CI 1.01 - 3.30), eating difficulties (PR=1.98; 95%CI 1.10 - 3.56), and postmenopausal bleeding (PR=2.91; 95%CI 1.55 - 5.44). The presence of pelvic pain, constipation, dyspareunia, fatigue, abdominal pain, nausea or vomiting, menstrual irregularity, weight loss, diarrhea, and bleeding after intercourse was similar in both groups. CONCLUSIONS: In women with adnexal tumors including indication of surgical treatment, the preoperative evaluation of symptoms may help predicting malignancy.

[Prevalence of HPV 16, 18, 45 and 31 in women with cervical lesions]. / Prevalência dos HPV 16, 18, 45 e 31 em mulheres com lesão cervical.

PURPOSE: To determine the prevalence of HPV 16, 18, 31 and 45 in cervical screening samples of women with cellular changes and/or colposcopy suggestive of persistent high grade or low grade lesion who were submitted to conization. METHODS: A total of 120 women were included in the study. Histological analysis of the cervical cones revealed 7 cases of cervicitis, 22 of CIN1, 31 of CIN2, 54 of CIN3, and 6 invasive carcinomas. The cervical screening samples were analyzed before conization for the presence of HPV-DNA by PCR using the consensus primers PGMY09/11. HPV-DNA-positive samples were tested for the presence of HPV16, 18, 31 and 45 using type-specific primers for these HPV. RESULTS: HPV-DNA was detected in 67.5% of the studied women. HPV 16 (40%) was the most prevalent type in most ilesions, followed by HPV 31 (13.3%), 45 (13.3%), and 18 (4.1%). Multiple infections occurred in 15% of the cases and infections with other HPV types were detected in 14% of the sample. CONCLUSIONS: HPV 16 and 18 infections do not always occur as a single infection, and may be associated with other HPV types on different occasions.

Prevalência dos HPV 16, 18, 45 e 31 em mulheres com lesão cervical / Prevalence of HPV 16, 18, 45 and 31 in women with cervical lesions

OBJETIVO: avaliar a prevalência dos HPV 16, 18, 31 e 45 em amostras de raspado cervical de mulheres com alterações celulares e/ou colposcopia sugestiva de lesão de alto grau ou lesão de baixo grau persistente submetidas à conização. MÉTODOS: Foram incluídas 120 mulheres. A análise histológica dos cones cervicais revelou 7 casos de cervicite, 22 de NIC1, 31 de NIC2, 54 de NIC3 e 6 carcinomas invasores. Foram analisadas as amostras de raspado cervical coletadas antes da conização para a presença do DNA-HPV por PCR com os primers de consenso, PGMY09/11. As amostras positivas para DNA de HPV foram testadas para presença do HPV16, 18, 31 e 45 utilizando-se primers tipo específico para esses HPV. RESULTADOS: O DNA-HPV foi detectado em 67,5 por cento das mulheres. O HPV 16 (40 por cento) foi o tipo mais prevalente na maioria das lesões, seguido dos HPV 31 (13,3 por cento), 45 (13,3 por cento) e 18 (4,1 por cento). Infecções múltiplas ocorreram em 15 por cento dos casos e as infecções por outros tipos de HPV foram detectadas em 14 por cento da amostra. CONCLUSÕES: as infecções pelos HPV 16 e 18 nem sempre ocorrem de maneira solitária (infecção única), estando associadas a outros tipos de HPV em diversas ocasiões.

PURPOSE: to determine the prevalence of HPV 16, 18, 31 and 45 in cervical screening samples of women with cellular changes and/or colposcopy suggestive of persistent high grade or low grade lesion who were submitted to conization. METHODS: a total of 120 women were included in the study. Histological analysis of the cervical cones revealed 7 cases of cervicitis, 22 of CIN1, 31 of CIN2, 54 of CIN3, and 6 invasive carcinomas. The cervical screening samples were analyzed before conization for the presence of HPV-DNA by PCR using the consensus primers PGMY09/11. HPV-DNA-positive samples were tested for the presence of HPV16, 18, 31 and 45 using type-specific primers for these HPV. RESULTS: HPV-DNA was detected in 67.5 percent of the studied women. HPV 16 (40 percent) was the most prevalent type in most ilesions, followed by HPV 31 (13.3 percent), 45 (13.3 percent), and 18 (4.1 percent). Multiple infections occurred in 15 percent of the cases and infections with other HPV types were detected in 14 percent of the sample. CONCLUSIONS: HPV 16 and 18 infections do not always occur as a single infection, and may be associated with other HPV types on different occasions.

Phylogenetic classification of human papillomavirus genotypes in high-grade cervical intraepithelial neoplasia in women from a densely populated Brazilian urban region.

CONTEXT AND OBJECTIVE: Differences in human papillomavirus (HPV) types may correlate with the biological potential and invasion risk of high-grade cervical intraepithelial neoplasia (CIN 2 and CIN 3). The objective of this study was to determine the relationship between different combinations of HPV types and CIN severity. DESIGN AND SETTING: Cross-sectional study, at Universidade Estadual de Campinas (Unicamp). METHODS: Cervical samples from 106 women treated due to CIN 2 (18) or CIN 3 (88) were examined for specific HPV genotypes using Roche Linear Array (LA-HPV). The proportions of CIN 2 and CIN 3 in groups of women infected with the HPV phylogenetic groups A7 and A9 were compared. Three groups were formed: women with single infections; multiple infections; and the whole sample. RESULTS: Multiple infections were detected in 68 samples (64.7%). The most frequent high-risk genotypes detected (single/multiple) were HPV 16 (57.1%), HPV 58 (24.7%), HPV 33 (15.2%), HPV 52 (13.3%), HPV 31 (10.4%), HPV 51 (7.6%) and HPV 18 (6.6%). Women without infection with HPV species Alpha 9 were less likely to have CIN 3 than were their Alpha 9 HPV-infected counterparts. HPV 16 and/or HPV 18, with or without associations with other viral types, were more frequently found in women with CIN 3 than in those with CIN 2. CONCLUSIONS: The severity of high-grade CIN may be aggravated by the presence of HPV types included in the Alpha 9 phylogenetic classification and by infections including HPV 16 and 18, singly or in combination with other HPV genotypes.

Phylogenetic classification of human papillomavirus genotypes in high-grade cervical intraepithelial neoplasia in women from a densely populated Brazilian urban region / Classificação filogenética dos genótipos de papilomavírus humano em neoplasia intraepitelial cervical de alto grau em mulheres de uma região urbana brasileira densamente povoada

CONTEXT AND OBJECTIVE: Differences in human papillomavirus (HPV) types may correlate with the biological potential and invasion risk of high-grade cervical intraepithelial neoplasia (CIN 2 and CIN 3). The objective of this study was to determine the relationship between different combinations of HPV types and CIN severity. DESIGN AND SETTING: Cross-sectional study, at Universidade Estadual de Campinas (Unicamp). METHODS: Cervical samples from 106 women treated due to CIN 2 (18) or CIN 3 (88) were examined for specific HPV genotypes using Roche Linear Array® (LA-HPV). The proportions of CIN 2 and CIN 3 in groups of women infected with the HPV phylogenetic groups A7 and A9 were compared. Three groups were formed: women with single infections; multiple infections; and the whole sample. RESULTS: Multiple infections were detected in 68 samples (64.7 percent). The most frequent high-risk genotypes detected (single/multiple) were HPV 16 (57.1 percent), HPV 58 (24.7 percent), HPV 33 (15.2 percent), HPV 52 (13.3 percent), HPV 31 (10.4 percent), HPV 51 (7.6 percent) and HPV 18 (6.6 percent). Women without infection with HPV species Alpha 9 were less likely to have CIN 3 than were their Alpha 9 HPV-infected counterparts. HPV 16 and/or HPV 18, with or without associations with other viral types, were more frequently found in women with CIN 3 than in those with CIN 2. CONCLUSIONS: The severity of high-grade CIN may be aggravated by the presence of HPV types included in the Alpha 9 phylogenetic classification and by infections including HPV 16 and 18, singly or in combination with other HPV genotypes.

CONTEXTO E OBJETIVO: Diferentes tipos de papilomavírus humano (human papillomavirus, HPV) podem ser correlacionados com a capacidade biológica e risco de invasão das neoplasias intra-epitelial de alto grau cervical (NIC 2 e NIC 3). O objetivo deste estudo foi determinar a relação de diferentes tipos de HPV com a gravidade da NIC. TIPO DE ESTUDO E LOCAL: Estudo transversal na Universidade Estadual de Campinas (Unicamp). MÉTODOS: Foram avaliados os genótipos específicos de HPV da amostra cervical de 106 mulheres com NIC 2 (18) ou NIC 3 (88), utilizando Roche Linear Array® (LA) HPV genotyping assay. Foram comparadas as proporções de NIC 2 e NIC 3 em grupos de mulheres infectadas com tipos de HPV dos grupos filogenéticos A7 e A9. Três grupos foram formados: mulheres com infecção simples; infecção múltipla; e infecção simples e múltipla. RESULTADOS: Infecções múltiplas foram detectadas em 68 (64,7 por cento) das amostras. Os genótipos de alto risco mais frequentemente detectados em infecção simples ou múltipla foram HPV 16 (57,1 por cento), HPV 58 (24,7 por cento), HPV 33 (15,2 por cento), HPV 52 (13,3 por cento), HPV 31 (10,4 por cento), HPV 51 (7,6 por cento) e HPV 18 (6,6 por cento). A probabilidade de mulheres com NIC 3 serem infectadas com HPV que não da espécie Alfa 9 era menor do que com os tipos de HPV da espécie Alfa 9. HPV 16 e ou 18, associado ou não com outros tipos virais eram mais frequentemente encontrados nas mulheres com NIC 3 do que naquelas com NIC 2. CONCLUSÃO: A gravidade da NIC de alto grau pode ser aumentada pela presença de tipos de HPV incluídos na classificação filogenética Alfa 9 e por infecções que incluem HPV 16 e 18 combinados ou não com outros genótipos de HPV.

Recovery of DNA for the detection and genotyping of human papillomavirus from clinical cervical specimens stored for up to 2 years in a universal collection medium with denaturing reagent.

The recovery and stability of DNA for the detection and genotyping of HPV in UCM-containing specimens, after exposure to denaturing reagents and stored for up to 2 years were evaluated. Samples were collected from 60 women who had cervical cytology specimens harboring cervical intraepithelial neoplasia (CIN) 2 or 3. All samples were stored in UCM and had been frozen at -20 degrees C following the addition of the denaturing reagent (sodium hydroxide) and the removal of the aliquot required for Hybrid Capture 2 testing for the identification of HPV DNA. The samples had been stored for 6, 12 and 24 months (20 samples for each storage time). HPV DNA extraction was performed according to a protocol designed specifically and the presence and quality of DNA was confirmed by human beta-globin detection using the consensus primers G73 and G74. HPV DNA was amplified using the consensus primers PGMY09 and PGMY11, and reverse line-blot hybridization was used to detect type-specific amplicons for 37 HPV types. The DNA extracted from the denatured specimen was recovered in 57/60 (95%) of the samples. HPV DNA was detected in 56/57 (98%) of the recovered samples. Twenty-six of the 56 samples recovered (48%) were genotyped successfully.

Factors associated with HPV persistence after treatment for high-grade cervical intra-epithelial neoplasia with large loop excision of the transformation zone (LLETZ).

BACKGROUND AND OBJECTIVE: Human Papillomavirus (HPV) persistence after high-grade cervical intra-epithelial neoplasia (CIN) removal may be associated with residual lesions or risk of disease recurrence. Knowledge regarding the factors associated with HPV persistence following CIN treatment is still limited. The main purpose of this longitudinal study was to assess the association between characteristics of the patients and their cervical lesions with high-risk HPV-type persistence, detected by commercially available Hybrid Capture II (HC II), after CIN 2 and 3 treatment with large loop excision of the transformation zone (LLETZ). STUDY DESIGN: For this cohort study, a total of 94 women submitted to LLETZ between March 2001 and September 2002 were included. Only women with at least one follow-up visit at 6 or 12 months and confirmed CIN 2 or 3 in the cone specimen were considered. In each visit women answered to a questionnaire and undertook Pap smear and HC II specimens collection. McNemar's, chi-square and Fisher tests were used for univariate analysis. Generalized Estimating Equations (GEE) were used for multivariate analysis. All calculations were performed within 95% confidence intervals (95% CI). RESULTS: Histological evaluation showed 12 (13%) women with CIN, 2 and 82 (87%) with CIN 3 and conization margins were compromised in 27 (29%) cases. Eighty-seven (92%) women showed positive HC II tests prior to LLETZ. Of women initially HPV negative, none had a positive HC II during follow-up. The proportion of positive HPV tests was reduced from 92% to 20%(P < 0.01) at the first visit and to 22% (P < 0.01) at the second visit after LLETZ. Multivariate analysis showed that smoking and age above 35 years (irrespective of margin status) were strongly associated with positive HPV during follow-up. CONCLUSION: HPV persistence following LLETZ was associated with smoking and with the interaction between age and conization margins.