Analysis of the Differences between Bethesda Groups according to Conventional Smear and Liquid-Based Cytology Methods in Cervicovaginal Cytology: A Single-Center Experience with 165,915 Cases.

INTRODUCTION: Liquid-based cytology (LBC) has replaced conventional smear (CS) in the world. In this study, through a series with a large number of cases, we aimed to make a comparison and general evaluation in all groups, primarily epithelial abnormalities, according to LBC and CS methods. This study was carried out in a private pathology laboratory located in a metropolitan city, where cytological materials sent from many clinics were examined. MATERIAL AND METHODS: There were 165,915 cases whose smears were examined between 2012 and 2020, most of them conventional (131,224 CS, 34,691 LBC). Cases were evaluated on the basis of the Bethesda 2014 classification and divided into sub-diagnostic categories after they were divided into two main groups as "with epithelial abnormalities" and "without." χ2 and Fischer's precision statistical tests were conducted using SPSS 23.0 package. In the CS process, cervical samples were obtained using an endocervical brush and a spatula. Cells were directly spread onto the slides and promptly fixed in 95% ethanol, followed by staining with the standard Papanicolaou stain. For LBC ThinPrep, cervical specimens were gathered using a cervix brush. The brush was washed in a vial and discarded. Finally, cells were isolated through vacuum filtration and transferred to the slide using air pressure. RESULTS: Squamous cell abnormalities (atypical squamous cells of undetermined significance [ASC-US], atypical squamous cells - cannot exclude high-grade squamous intraepithelial lesion [ASC-H], low-grade squamous intraepithelial lesion [LSIL], high-grade squamous intraepithelial lesion [HSIL], squamous cell carcinoma, atypical glandular cells of undetermined significance) were reported in 5,696 (3.43%) cases. ASC (ASC-US + ASC-H)/SIL ratio (1.36/2.04) was found to be 0.67 (recommended Bethesda ratio is <3). ASC-US (p < 0.001), ASC-H (p < 0.001), and HSIL(p < 0.001) detection rate of LBC was found to be significantly higher than CS. ASC-US (1.8/1.2), ASC-H (0.08/0.008), and HSIL (0.6/0.3) case ratios of LBC/CS were found to be significantly higher in LBC. LSIL (1.72/1.66) rate was similar. CONCLUSION: LBC is superior to CS in detecting epithelial lesions. In addition to being used as a screening method, it is clear that it makes a great contribution to reducing cervical carcinomas due to HPV typing. Definitive comments regarding comparison of methods on reactive changes and microorganism detection are challenging. Preanalytical factors might account for these situations.

Comparison of liquid-based cytology and cell blocks prepared from cell remnants for diagnosis of cervical pathology.

BACKGROUND: Cervical cancer is a global public health problem with high mortality. Advances in screening programs for cervical cancer are considered key to eliminate cervical cancer. We aimed to examine the contribution of cell block analysis to the detection of epithelial cell abnormalities in cervical smear samples. METHODS: A total of 559 patients with suspected cervical pathology were examined, and their samples were analyzed by both liquid-based cytology (LBC) and cell blocks. The biopsy results of 149 out of the 559 patients were obtained. RESULTS: Of the 50 patients who were identified as HSIL by biopsy, only 12 were diagnosed as HSIL by the LBC method, 22 as LSIL, 12 as ASCUS, and 4 as ASC-H (p < 0.001). With the cell block analysis, results for these patients were: 20 HSIL, 17 LSIL, 7 NILM, 4 'unsatisfactory', and 2 ASC cases (p < 0.001). LBC detected only 1 of the 10 patients with biopsy-diagnosed tumors, while 7 of these were defined as HSIL, 1 as ASCUS and 1 as AGC. The results of cell block analysis in patients with biopsy-diagnosed tumors were as follows: 7 HSIL, 1 tumor, 1 ASC and 1 LSIL. CONCLUSIONS: Cell block analysis might be superior to LBC in terms of diagnostic accuracy in cervical pathologies, particularly in the detection of HSIL. However, both methods were similarly poor in diagnosing tumors. Cell blocks may improve diagnostic accuracy and can be a complementary method to LBC, while having the advantage of revealing histological architecture.

Defining the Longitudinal Risk of CIN 3+ for

OBJECTIVE: To determine the baseline and cumulative risk of cervical intraepithelial neoplasia (CIN)3 and invasive cervical cancer in participants referred to colposcopy with high-grade cytology and

Added value of electrical impedance spectroscopy in adjunction of colposcopy: a prospective cohort study.

OBJECTIVE: To assess whether electrical impedance spectroscopy (EIS) as an adjunctive technology enhances the performance of colposcopy. DESIGN: Prospective cohort study. SETTING: University Hospital colposcopy clinic. PARTICIPANTS: Colposcopy with EIS for 647 women and conventional colposcopy for 962 women. INTERVENTIONS: Comparison of the performance of colposcopy by referral cervical cytology in two cohorts, with and without EIS as an adjunctive technology. OUTCOME MEASURES: Prevalence of cervical intraepithelial neoplasia grade 2 or worse (CIN2+), diagnostic testing accuracy to detect CIN2+ with and without EIS and their relative differences between cohorts. RESULTS: The prevalence of CIN2+ varied between the cohorts according to referral cytology: 17.0% after abnormal squamous cells of unknown significance referral cytology in EIS cohort and 9.1% in the reference cohort, 16.5% and 18.9% after low-grade squamous intraepithelial lesion (LSIL), 44.3% and 58.2% after atypical squamous cells, cannot exclude high-grade squamous intraepithelial lesion (HSIL) (atypical squamous cells that cannot exclude HSIL), and 81.9% and 77.0% after HSIL cytology, respectively. Sensitivity to detect CIN2+ was higher in the EIS cohort, varying from 1.79 (95% CI 1.30 to 2.45) after LSIL referral cytology to 1.16 (95% CI 1.09 to 1.23) after HSIL referral cytology, with correspondingly lower specificity after any referral cytology. CONCLUSIONS: Colposcopy with EIS had overall higher sensitivity but lower specificity to detect CIN2+ than conventional colposcopy. CIN2+ prevalence rates were, however, not consistently higher in the EIS cohort, suggesting innate differences between the cohorts or truly lower detection rates of CIN2+ for EIS, highlighting the need for randomised controlled trials on the effectiveness of EIS.

Are HPV-negative lesions concerned for the introduction of primary HPV testing for cervical cancer screening in Japan?

AIM: In Japan, primary human papilloma virus (HPV) testing has not been introduced for cervical cancer screening due to concerns that HPV-negative lesions may be missed and a lack of Japanese data. The purpose of this study was to evaluate the validity of primary HPV testing in Japan by analyzing cervical intraepithelial neoplasia 2 (CIN2) or more/high-risk HPV- (CIN2+/hrHPV-) cases in cervical cancer screening. METHODS: Data from 35 525 cervical cancer screenings with HPV testing and cervical cytology from 2011 to 2019 in Saga City, Japan, were reviewed. The cases with low-grade squamous intraepithelial lesion (LSIL+)/hrHPV- were analyzed in detail. RESULTS: The results of the 35 525 examinees were as follows: 31 123 were negative for intraepithelial lesion or malignancy (NILM)/hrHPV-, 2612 were NILM/hrHPV+, 262 were atypical squamous cells of undetermined significance (ASC-US)/hrHPV-, 213 were ASC-US/hrHPV+, 291 were LSIL+/hrHPV-, and 1024 were LSIL+/hrHPV+. Of the 256 LSIL+/hrHPV- examinees for whom histology was available, CIN2+ were CIN2 9.4% (24/256), CIN3 3.9% (10/256), cervical adenocarcinoma 0.4% (1/256), uterine corpus cancer 1.2% (3/256), and uterine sarcoma 0.4% (1/256). Overall, the rate of LSIL+/hrHPV- was 0.82% (291/35.525), 0.1% (36/35525) of which were cervical lesions with CIN2+. Only one cervical adenocarcinoma was detected, but gastric-type adenocarcinoma was not included. CONCLUSION: HPV-negative CIN2+ or cervical adenocarcinoma is not a concern for the introduction of primary HPV screening in Japan. Primary HPV testing in cervical cancer screening is considered a feasible method that can be used in Japan, although an algorithm suitable for Japan and a national-level management system need to be established.

High-risk human papillomavirus genotyping in women with atypical squamous cells of undetermined significance.

We conducted a prospective study to evaluate the prevalence of high-risk human papillomavirus (hr-HPV) positivity in women with atypical squamous cells of undetermined significance (ASC-US). Additionally, we assessed the association of hr-HPV positivity with the pathology of high-grade squamous intraepithelial lesions or worse (HSIL+) and the risk of subsequent detection of squamous intraepithelial lesions. A total of 376 women were included, with 242 (64.4%) exhibiting hr-HPV positivity. The predominant HPV genotypes were 16, 52 and 58. Factors associated with the immediate detection of HSIL+ pathology included a colposcopic impression of high-grade lesions, hr-HPV positivity, HPV 16 positivity, HPV 18 positivity, HPV 58 positivity, age less than 40 years, and biopsy of two or more pieces. However, only the first three factors were statistically significant in multivariate analysis. Among the 291 women who continued surveillance for 6 months or more, the median follow-up period was 41.8 months (interquartile range [IQR] 26.5-54.0). The prevalence of subsequent HSIL in women with hr-HPV positivity versus negativity was 3.6% versus 0.98%, respectively. The median time to the subsequent detection of SIL was 28.7 months (IQR 14.9-41.7). In conclusion, women with ASC-US in our study had a high proportion of hr-HPV positivity. Type-specific HPV testing could play a pivotal role in the development of specific management protocols for women with ASC-US.Clinical trial registration: https://thaiclinicaltrials.org , TCTR20161017002.

Evaluation of the clinical performance of p16/Ki-67 dual-staining cytology for cervical lesion detection in premenopausal and postmenopausal Chinese women.

BACKGROUND: Studies on the clinical performance of p16/Ki-67 dual-staining in detecting cervical lesions by menopausal status were limited. METHODS: 4364 eligible women were enrolled with valid p16/Ki-67, HR-HPV, and LBC test results, including 542 cancer and 217 CIN2/3 cases. The positivity rates of p16 and Ki-67 single staining and p16/ Ki-67 dual-staining were analyzed by different pathological grades and age groups. The sensitivity (SEN), specificity (SPE), positive predictive value (PPV), and negative predictive value (NPV) of each test in different subgroups were calculated and compared. RESULTS: P16/Ki-67 dual-staining positivity increased with histopathological severity in premenopausal and postmenopausal women (P < 0.05), while no increasing trends of individual expression of p16 single staining and Ki-67 single staining were observed in postmenopausal women. P16/Ki-67 showed higher SPE (88.09% vs. 81.91%, P < 0.001) and PPV (33.8% vs. 13.18%, P < 0.001) in detecting CIN2/3, and higher SEN (89.97% vs. 82.61%, P = 0.012) and SPE (83.22% vs. 79.89%, P = 0.011) in detecting cancer in premenopausal women than postmenopausal women. For triaging the HR-HPV+ population to identify CIN2/3, p16/Ki-67 performed comparably to LBC in the premenopausal women, and showed higher PPV (51.14% vs. 23.08%, P < 0.001) in premenopausal than postmenopausal women. For triaging ASC-US/LSIL population, p16/Ki-67 demonstrated higher SPE and lower colposcopy referral rate than HR-HPV in both premenopausal and postmenopausal women. CONCLUSIONS: Expressions of p16/Ki-67 dual-staining between premenopausal and postmenopausal women are varied. P16/Ki-67 performs better in detecting cervical lesions in premenopausal women. For triaging, p16/Ki-67 is suitable for HR-HPV+ women, especially premenopausal women, to identify CIN2/3 and women with ASC-US/LSIL.

13-Type HPV DNA Test versus 5-Type HPV mRNA Test in Triage of Women Aged 25-33 Years with Minor Cytological Abnormalities-6 Years of Follow-Up.

BACKGROUND: A specific, cost-effective triage test for minor cytological abnormalities is essential for cervical cancer screening among younger women to reduce overmanagement and unnecessary healthcare utilization. We compared the triage performance of one 13-type human papillomavirus (HPV) DNA test and one 5-type HPV mRNA test. METHODS: We included 4115 women aged 25-33 years with a screening result of atypical squamous cells of undetermined significance (ASC-US) or low-grade squamous intraepithelial lesions (LSIL) recorded in the Norwegian Cancer Registry during 2005-2010. According to Norwegian guidelines, these women went to triage (HPV testing and repeat cytology: 2556 were tested with the Hybrid Capture 2 HPV DNA test, which detects the HPV types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, and 68; and 1559 were tested with the PreTect HPV-Proofer HPV mRNA test, which detects HPV types 16, 18, 31, 33, and 45). Women were followed through December 2013. RESULTS: HPV positivity rates at triage were 52.8% and 23.3% among DNA- and mRNA-tested women (p < 0.001), respectively. Referral rates for colposcopy and biopsy and repeat testing (HPV + cytology) after triage were significantly higher among DNA-tested (24.9% and 27.9%) compared to mRNA-tested women (18.3% and 5.1%), as were cervical intraepithelial neoplasia grade 3 or worse (CIN3+) detection rates (13.1% vs. 8.3%; p < 0.001). Ten cancer cases were diagnosed during follow-up; eight were in DNA-tested women. CONCLUSION: We observed significantly higher referral rates and CIN3+ detection rates in young women with ASC-US/LSIL when the HPV DNA test was used at triage. The mRNA test was as functional in cancer prevention, with considerably less healthcare utilization.

Evaluation of 1,030,482 Cervical Smear Results in Brazilian Population.

BACKGROUND: The objective was to determine the prevalence of abnormal cytologic smears in a large population of a Brazilian city. METHODS: Retrospective study of cervical cytology results performed at a private laboratory in São Paulo - Brazil. A total of 1,030,482 cytology tests were performed between January 2010 and December 2015. RESULTS: Among the satisfactory cytologies, we observed abnormal results in 8.9% (91,371). Analyzing the proportion of exams with altered results over the years, there was no change in these rates. We observed that the proportion of abnormal exams decreased with increasing age. Atypical squamous cells of undetermined significance, possibly non-neoplastic (ASC-US) cytology was the most common abnormality found in the general population with 73.19%, followed by low-grade intraepithelial lesion (LSIL) with 20.5%; these 2 cytological abnormalities add up to 93.69 % of all abnormal results. There was an increase in ASC-US with advancing age, a decrease in prevalence of LSIL, especially after the age of 30 and high-grade intraepithelial lesion (HSIL) shows two peaks of elevation, respectively in the age groups of 30-39 years and over 70 years. CONCLUSION: Women under the age of 30 showed a higher prevalence of cytological abnormalities, however most of these changes are low-grade lesions. In older women, cytological abnormalities are largely high-grade lesions, requiring greater care to prevent progression to cancer.

Efficacy of optical coherence tomography in the triage of women with minor abnormal cervical cytology before colposcopy.

OBJECTIVES: To evaluate the efficacy of Optical Coherence Tomography (OCT) for detecting cervical lesions in women with minor abnormal cytology results (atypical squamous cells of undetermined significance (ASC-US) and low-grade squamous intraepithelial lesion (LSIL)). METHODS: A prospective study was conducted at gynecologic clinic from Mar 2021 to Sep 2021. The recruited women with cervical cytological findings of ASC-US or LSIL were inspected with OCT before colposcopy-directed cervical biopsy. The diagnostic performance of OCT, alone and in combination with high-risk human papillomavirus (hrHPV) testing were evaluated to detect cervical intraepithelial neoplasia of grade 2 or worse (CIN2+)/CIN3 or worse (CIN3+). The rate of colposcopy referral and the immediate risk of CIN3+ of OCT were calculated. RESULTS: A total of 349 women with minor abnormal cervical cytology results were enrolled. For detection of CIN2+/CIN3+, the sensitivity and NPV of OCT were lower than those of hrHPV testing (CIN2+: 71.3% vs. 95.4%, 89.0% vs. 91.1%, P < 0.001; CIN3+: 75% vs. 93.8%, 96.5% vs. 95.6%, P < 0.001), but the specificity, accuracy and PPV were higher than those of hrHPV testing (CIN2+: 77.5% vs. 15.6%, 75.9% vs. 35.5%, 51.2% vs. 27.3%, P < 0.001; CIN3+: 69.4% vs. 13.6%, 69.9% vs. 20.9%, 19.8% vs. 9.9%, P < 0.001). OCT combined with hrHPV testing (CIN2+: 80.9%; CIN3+: 72.6%) showed higher specificity than that of OCT alone (P < 0.001). The colposcopy referral rate base on OCT classification was lower than that based on hrHPV testing (34.7% vs. 87.1%, P < 0.001). Patients with hrHPV-positive ASC-US and hrHPV-negative LSIL cytology, the immediate CIN3+ risk in OCT negative cases was less than 4%. CONCLUSIONS: OCT alone or combination with hrHPV testing shows good performance for detecting CIN2+/CIN3+ in patients with ASC-US/LSIL cytology. OCT is an effective method for colposcopy triage in women with hrHPV-positive ASC-US and hrHPV-negative LSIL cytology.

Cervical cytology results among pregnant and non-pregnant women in Brazil.

OBJECTIVE: To compare the prevalence of abnormal cervical smear results among pregnant and non-pregnant women with and without representation of the transformation zone (TZ) in a mid-sized city in Brazil. STUDY DESIGN: This observational analytical and retrospective study analyzed cervical smear results from pregnant and non-pregnant women in a mid-sized city in Brazil. Private data were collected at private clinics over seven years and from public services over 16 years. All results were reported according to Bethesda System. Pregnant women were separated from non-pregnant women, and women were stratified into two age groups (25-29 years and 30-34 years) in general analyses. For TZ representation and cytologic-histologic correlation, there was no age stratification. RESULTS: Among public services cervical smears, the frequency of atypical squamous cells of undetermined significance (ASC-US) was higher for pregnant women in the entire group (odds ratio [OR] 1.91; confidence interval [CI] 1.07-3.39) and among women from 30 to 40 years old (OR 2.79; CI 1.38-5.66). The same occurred for low-grade squamous intraepithelial lesion (LSIL) in the total group (OR 3.50; CI 1.44-8.53) and among women 30 to 40 years old (OR 4.57; CI 1.45-14.42). The frequency of ASC-US was higher in pregnant women with TZ representation exams than those without (OR 4.62; CI 2.17-9.84). The same occurred for those with atypical squamous cells, which cannot exclude high-grade squamous intraepithelial lesion (OR 9.81; CI 1.63-59.15) and LSIL (OR 10.47; CI 3.39-32.36). Among pregnant women with abnormal cervical smear results, 85.74% were followed up through cytology, and an average of 26.12% of patients underwent biopsy, of which almost all were positive for dysplasia. CONCLUSION: Pregnant and non-pregnant women did not differ in cervical smears with actual precursor lesions. Nevertheless, considering the opportunistic nature of Brazilian cervical cancer screening, prenatal visits present an opportunity to perform cervical smears in pregnant women. TZ representation correlated with higher rates of abnormal cervical smears, reinforcing the clinical importance of endocervical/metaplastic cell representation, including among pregnant women.

Safety Run-in of Intramuscular pNGVL4a-Sig/E7(detox)/HSP70 DNA and TA-CIN Protein Vaccination as Treatment for HPV16+ ASC-US, ASC-H, or LSIL/CIN1.

Patients with human papillomavirus type 16 (HPV16) infection and low-grade cervical dysplasia [low-grade squamous intraepithelial lesion (LSIL)/CIN1] or atypical squamous cells [atypical squamous cells of undetermined significance (ASC-US)/atypical squamous cells- cannot exclude high-grade squamous intraepithelial lesion (ASC-H)] require active surveillance for disease progression. A safe and effective immunotherapy to clear HPV16 is an unmet medical need. The safety run-in cohort of a randomized double-blind, placebo-controlled phase II trial of PVX2 [vaccination twice with HPV16-targeting pNGVL4a-Sig/E7(detox)/HSP70 plasmid and once with the HPV16 L2E7E6 fusion protein "TA-CIN"] as immunotherapy for patients with HPV16+ ASC-US, ASC-H, or LSIL/CIN1 (NCT03911076) was recently completed. The primary objective of this cohort was to determine the safety and tolerability of PVX2 vaccination. Subjects were confirmed to have HPV16 infection and LSIL/CIN1, ASC-US, or ASC-H. Adverse events were evaluated using Common Terminology Criteria for Adverse Events v5.0. HPV typing by HPV16 18/45 Aptima Assay was done at baseline, month 6, and month 12, with simultaneous cytology analysis. Cervical biopsies and endocervical curettage were performed at baseline and month 6. In the safety run-in cohort 12 eligible patients were enrolled. Each received three monthly immunizations. One was lost to follow-up after week 12. There were no serious adverse events. A total of five adverse events were noted by 4 patients; 4 were considered not vaccine-related, and one 'unlikely related' by the investigator. At month 6, 45% (5/11) of participants converted to HPV16-negative and 2 others developed CIN2+ and received a loop electrosurgical excision procedure. At month 12, 64% (7/11) were HPV16-negative, including those HPV16-negative at month 6. In conclusion, PVX2 immunotherapy was well tolerated and associated with viral regression, supporting further testing. PREVENTION RELEVANCE: This safety run-in study cohort suggests that PVX2 immunotherapy is well tolerated in the target population and is sufficiently safe to warrant further clinical testing in a randomized study. The combined vaccines may facilitate higher-than-expected rate of human papillomavirus type 16 viral clearance 6 and 12 months after treatment, although this requires validation.

Cytology compared with Hybrid Capture 2 human papilloma virus cervical cancer screening in HIV positive and HIV negative South African women.

OBJECTIVES: Cervical cancer is preventable and caused by persistent infection with oncogenic human papilloma virus (HPV) types. HPV screening is more sensitive and is the preferred screening test. HPV screening data are mainly from developed settings, and the purpose of this study was to investigate the performance of HPV screening in previously unscreened HIV positive and negative women. METHODS: In this cross sectional multicenter study, liquid based cytology and HPV testing were performed on women attending different clinics. Patients with positive screening tests had colposcopy and biopsy or large loop excision of the transformation zone. Some women with normal screening had colposcopy and biopsy. Data of women with histology results, and data of HIV positive and negative women were analyzed for comparison. For women without histology results, data were imputed using a statistical model. RESULTS: In 903 women with known HIV status, 683 (75.6%) had negative cytology, 202 women (22.4%) had abnormal cytology, and in 18 patients (2.0%) the results were uncertain. Mean age was 41.4 years (range 25-65). HPV tests were negative in 621 women (68.8%). In HIV positive women, 54.5% tested negative compared with 79.7% HIV negative women (p<0.0001). HPV screening had higher sensitivity (60.9%), but lower specificity (82.4%), compared with cytology (48.6% and 86.7%) for detection of cervical intraepithelial neoplasia (CIN) 2+ in all women. For detection of CIN 3+, HPV screening had higher sensitivity (70.4%) compared with cytology (62.9%), and specificity (75.5%) was lower compared with cytology at a threshold of atypical squamous cells of undetermined significance (ASCUS+) (82.4%). CONCLUSION: HPV screening was more sensitive than cytology in HIV positive and HIV negative women, but specificity was lower. Although HPV screening should be the preferred screening test, cytology is a suitable screening test in HIV positive women in low resource settings. TRIAL REGISTRATION NUMBER: NCT02956031.

Feasibility of implementing cytology-based cervical cancer screening national program in Lebanon: A pilot study.

OBJECTIVE: To review our national cervical cancer screening program using existing Ministry of Public Health primary healthcare centers (PHCs) and report the impact of women's knowledge, attitude, behavior, and practices on screening uptake and outcome. METHODS: A cross-sectional study on cervical cancer screening offered to sexually active Lebanese women aged 21 years and above visiting PHCs. Exclusions were history of complete hysterectomy, gynecologic cancers, and current pregnancy. Data were collected through a questionnaire and conventional cervical smear performed by trained healthcare providers and sent to one centralized cytopathology laboratory. RESULTS: Of 12 273 eligible women, 1.7% had an abnormal cervical smear test including 161 atypical squamous cells (ASC) of undetermined significance, 6 atypical glandular cells of undetermined significance, 16 low-grade squamous intraepithelial lesion (SIL), 17 ASC-cannot rule out high-grade SIL, 11 high-grade SIL, and one invasive carcinoma. Knowledge and attitudes significantly affected participation in screening; women lacking awareness had rarely undergone a cervical smear. CONCLUSION: In Lebanon, cytology-based cervical cancer screening is feasible within the PHCs. Positive screen incidence was low. Despite previous campaigns, a low level of knowledge persists, and affects women's com with the screening guidelines. Advocacy and awareness activities by key healthcare providers may help to improve participation.

The interplay between HPV, other Sexually Transmissible Infections and genital microbiome on cervical microenvironment (MicroCervixHPV study).

Background: The importance of Cervicovaginal Microbiota in protecting against infections (such as HPV) is already well established, namely through Lactobacillus spp., as well as the mechanism through which HPV leads to Cervical Neoplasia. However, it is not possible to classify HPV as a complete carcinogen. Thus, the importance of exploring Cervicovaginal dysbiosis with the intention of deciphering this interaction with HPV, takes on greater relevance. The main objectives of this study were: 1) Comparison of the MCV composition of women with or without HPV and women with ASCUS or LSIL; 2) Characterization of cytokines present in the vaginal microenvironment; 3) Evaluation of the blood count ratios as prognostic systemic inflammatory biomarkers; 4) Correlation between MCV, HPV serotypes and cytokines. Methods: This was a retrospective, observational, multicenter, cross-sectional study. CVM analysis was performed by isolation RNA and sequencing on a NGS platform. Cytokine concentrations of CVM were obtained through Multiplex platform. Statistical analysis was performed in SPSS v 26.0. An α of 0.05 was considered statistically significant. Results: Highlighting the core of the study, CVM types of CST I and CST IV were found to influence the emergence of cervical lesions. Neutrophil-to-Lymphocyte ratio was found to impact the prognosis of ASCUS. Within CVM, Lactobacillus prevent the growth of other CST IV species, while the latter express symbiotic relationships with each other and show affinity for specific HPV serotypes. At last, RANTES chemokine is significantly elevated in cervicovaginal infections. Conclusion: The importance of using vaginal cytokine profiles and CVM is highlighted in the hypothesis of prevention of Cervical Neoplasia development, as well as in its use as a prognostic biomarker. Taken together, these insights are one step closer to personalized medicine.

Condyloma acuminatum of the urinary tract demonstrates atypical squamous cells in urine cytology.

Urine cytology of urinary tract condylomas has not been systematically studied. We analyzed cytologic features of urinary tract condylomas and evaluated potential diagnostic challenges and pitfalls. We retrospectively reviewed urine cytology of urinary tract condylomas from 2 academic institutions (2015-2022). Among 20 patients with urinary tract condylomas, 6 had urine cytology (2 samples in 1 patient), including 3 men and 3 women (mean age, 74.3 years; range, 65-86 years). Original interpretations were negative for high-grade urothelial carcinoma (NHGUC; n = 4), atypical urothelial cells (n = 1), reactive urothelial cells (n = 1), and negative for malignancy (n = 1). Squamous cells were noted in 3 cases, atypical squamous cells (ASC) consistent with low-grade squamous intraepithelial lesion (LSIL) were noted in 1 case, and in 3 cases, the presence of squamous cells was not mentioned. All urines were reclassified according to The Paris System as NHGUC. Specimens were composed of benign urothelial cells and groups or isolated ASC consistent with LSIL (n = 4), atypical keratinized squamous cells (n = 2), and ASC that did not meet LSIL criteria (n = 1). The LSIL cells showed nuclear enlargement (n = 4), hyperchromasia (n = 4), perinuclear halo (n = 3), nuclear membrane irregularity (n = 4), orangeophilic cytoplasm (n = 3), and binucleation (n = 4). The atypical keratinized squamous cells showed hyperchromasia (n = 2), nuclear membrane irregularity (n = 2), keratin pearls (n = 2), and binucleation (n = 1). The ASC that did not meet LSIL criteria showed nuclear enlargement and orangeophilic cytoplasm. Many urinary tract condylomas (57%) demonstrate classic LSIL features in urine cytology. Less frequent cases can mimic keratinizing squamous cell carcinoma (28%) or demonstrate ASC not diagnostic of LSIL (15%).

Applicability of On-Site Evaluation of Cervical Cytology Smears Stained with Toluidine Blue to Reduce Unsatisfactory Results.

INTRODUCTION: Carcinoma of the uterine cervix is a major health problem faced by Indian women. Screening techniques like visual inspection with acetic acid, Lugol's iodine, Papanicolaou smear, and human papillomavirus DNA testing have been suggested. Pap smear is a simple, safe, cost-effective, and reliable technique used for screening cervical lesions. Rapid on-site evaluation (ROSE) using the 1% aq. toluidine blue staining method has been less studied in cervical cytology. MATERIALS AND METHODS: Our study was a prospective study done over a period of 2 years. All the cervical cytology smears were reported as per the Bethesda system 2014. Rapid stain using aqueous toluidine blue (1%) and conventional Pap stain was done on the smears received. RESULTS: We evaluated a total of 1,300 cases, with 97.6% satisfactory samples. The spectrum of cases included 96.3% of negative for malignancy cases (including bacterial vaginosis, trichomonas, candida, and atrophic smears), atypical squamous cell of undetermined significance and atypical squamous cells cannot exclude high-grade squamous intraepithelial lesion in 0.5% cases, low-grade squamous intraepithelial lesion and high-grade intraepithelial lesion in 0.3% cases, squamous cell carcinoma in 0.3% cases, and atypical glandular cells/adenocarcinoma in 0.2% cases. Turnaround time was within 48 h in 77% cases. With rapid stain, our unsatisfactory rate was reduced from 12% to approx. 2.4%. CONCLUSION: ROSE has been attempted on routine FNA cytology samples with success. However, the use of ROSE in cervical cytology has not been attempted to date. Lower unsatisfactory rate is an important indicator for the successful implementation of cervical cancer screening technique.

P16/Ki-67 Dual Staining in Positive Human Papillomavirus DNA Testing for Predictive Diagnosis of Abnormal Cervical Lesions in Northeastern Thai Women.

OBJECTIVE: Cervical cancer screening can effectively reduce new cervical cancer cases, including in Thailand. The abnormal results are subsequently referred for colposcopy. To avoid unnecessary colposcopy, an efficient triage is still needed for validation. This study aimed to investigate the overall positivity of cytology-based screening, HPV detection, and p16/Ki-67 dual staining and evaluate different triage strategies for predictive diagnosis of abnormal cervical lesions in northeastern Thailand. METHODS: Cervical cells were collected from 191 women who came for cervical screening in the gynecological outpatient department during March 2019-February 2020. Pap smear samples were classified into 6 groups including 17 atypical glandular cells (AGC), 21 atypical squamous cells of undetermined significance (ASC-US), 7 atypical squamous cells - cannot exclude HSIL (ASC-H), 26 low-grade squamous intraepithelial lesions (LSILs), 19 high-grade SILs (HSILs) and 101 no squamous intraepithelial lesion (noSIL). Polymerase chain reaction (PCR) was performed for HPV DNA detection. HPV genotyping was determined by reverse line blot hybridization. P16/Ki-67 dual staining was performed by using CINtec PLUS Cytology kit. Biopsies from abnormal screening were collected for surgical pathology classification. RESULTS: High-risk HPV (HR-HPV) infection was 2.97%, 29.41%, 38.10%, 57.14%, 46.15% and 84.21% in noSIL, AGC, ASC-US, ASC-H, LSIL and HSIL cytology respectively. P16/ Ki-67 in noSIL, AGC, ASC-US, ASC-H, LSIL and HSIL was 0.99%, 5.88%, 9.52%, 42.86%, 26.92% and 63.16%, respectively (P-value < 0.001). Among p16/Ki-67 positive cases, 96.15% (25/26) were infected with HPV and 84.62% (22/26) were HR-HPV. The overall positivity of each and co-testing between cytology or HPV DNA testing or p16/Ki-67 dual staining was evaluated. In each cervical lesion, primary HPV DNA testing showed the highest sensitivity, but low specificity. The combined all HPV/HR-HPV with p16/Ki-67 detection increased the specificity of abnormal cervical lesions. CONCLUSION: P16/Ki-67 dual stain cytology in HPV-positive women performs well for diagnosis of abnormal cervical lesions and should be considered for management of HPV-positive women to avoid unnecessary colposcopy referrals.

LINE-1 and Alu Methylation in hrHPV-Associated Precancerous Cervical Samples.

OBJECTIVE: This study was conducted to determine global DNA methylation patterns in cervical cells cytologically identified as atypical squamous cells of unknown significance (ASCUS) with a normal, LSIL, or HSIL histopathological result. METHODS: Methylation patterns of long interspersed nuclear elements (LINE-1) and short interspersed element (Alu) sequences were assessed using the combined bisulfite restriction analysis (COBRA) method in cervical samples with cytology-diagnosed cervical lesions. RESULTS: In cervical precancerous lesions with hrHPV positive, the percentage of overall (mC) and mCmC LINE-1 methylation levels showed a stepwise increase from hrHPV positive normal to HSIL with significant differences (p<0.001). However, both methylation levels were significantly higher in hrHPV negative normal than in hrHPV positive normal (p<0.001). The overall (mC) Alu methylation in hrHPV positive LSIL and HSIL was lower than in hrHPV positive normal, with a significant difference (p<0.05). Remarkably, the percentage of uCmC and mCuC of LINE-1 and Alu in three different hrHPV positive cervical lesions showed a stepwise decrease from hrHPV positive normal, LSIL and HSIL, respectively. Furthermore, receiver operating characteristic (ROC) curve analyses revealed that the LINE-1 mC and mCmC patterns have high sensitivity and specificity for distinguishing HSIL from normal/LSIL in hrHPV positive cases at the appropriate cutoff levels. CONCLUSION: We have demonstrated the LINE-1 and Alu methylation data in normal and premalignant cervical epithelia. LINE-1 hypomethylation was found in hrHPV positive normal cells, with lower methylation levels associated with cancer features. In cytologically diagnosed Atypical Squamous Cells of Unknown Significance (ASCUS), the levels of mC and the mCmC pattern could be utilized in concert with hrHPV detection to classify the ASCUS sample prior to colposcopy.

Identification of Malignancy in PAP Smear Samples Using the CGB3 and NOP56 Genes as Methylation Markers.

BACKGROUND: Although various improvements have been made in the reporting of the Papanicolaou (PAP) test in recent years, there remain several challenges that have yet to be addressed in terms of determining a standardized methodology for categorizing atypical squamous cells of undetermined significance (ASC US). METHODS: The present study focuses on evaluating the performance of the methylation status of two genes (CGB3 and NOP56) using a total of 200 PAP samples, which were divided into the "determined" group, with 78 samples based on cytology, and the "undetermined" group (ASC US), with 122 samples. The promoter methylation status of the CGB3 and NOP56 genes was detected for the 200 PAP samples using methylation specific PCR (MSP). The diagnostic abilities of the CGB3 and NOP56 genes in PAP samples were measured, and receiver operating characteristic (ROC) curves were generated using Python programming language. RESULTS: Based on the validation of CGB3 and NOP56 methylation in the 200 PAP samples, both genes exhibited higher methylation percentages in abnormal samples compared with normal samples. In addition, on the basis of diagnostic performance analysis, the CGB3 gene exhibited the highest sensitivity and specificity in both histology based ASC US and cytology based 'determined' PAP samples, with significant diagnostic abilities [area under the curve (AUC) values of 0.83 and 0.74, respectively, where AUC ≥0.5 was determined to be significant] to distinguish between the "normal" and "abnormal" samples. CONCLUSION: The findings of the present study will contribute toward identifying a DNA methylation marker for the early detection of abnormal samples before they reach the initial stages of cervical cancer, and should prove to be helpful for clinicians in terms of diagnosing patients whose cells are ASC US.
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