Cyclin E expression and early cervical neoplasia in ThinPrep specimens. A feasibility study

OBJECTIVE: To investigate cyclin E expression as a possible marker for early cervical neoplasia using ThinPrep gynecologic specimens from premenopausal women. STUDY DESIGN: Archived ThinPrep liquid-based cervical/endocervical specimens (Cytyc Corporation, Boxborough, Massachusetts, U.S.A.) diagnosed as human papillomavirus infection (HPV) (20), atypical squamous cells of undetermined significance (ASCUS) (48) and within normal limits (WNL)/benign cellular changes (BCC) (21) were resampled in duplicate, fixed in 95% ethanol, subjected to immunocytochemical staining with the cyclin E antibody (clone 13A3, Novocastra Laboratories Ltd., Newcastle upon Tyne, U.K.) and HPV antibody (clone K1H8, Dako Corporation, Carpinteria, California, U.S.A.) and the expression scored by two pathologists and correlated with the cytologic diagnosis. A case was scored as positive if it contained > 10 abnormal squamous cells with nuclear immunocytochemical staining. RESULTS: The cylin E antibody assay was positive in 20 (100%) cases cytologically diagnosed as HPV. These cases were also anti-HPV antibody positive. Four cases (19%) cytologically diagnosed as WNL/BCC were cyclin E positive. Of these, two were anti-HPV antibody positive. Thirty-four (73%) cases cytologically diagnosed as ASCUS were positive for the cyclin E assay and for anti-HPV antibody staining. CONCLUSION: Cyclin E expression correlates strongly with morphologic features of HPV in ThinPrep specimens and may serve as a surrogate marker for HPV infection and early cervical preneoplastic lesions.     

A thin-layer,liquid-based pap test for mass screening in an area of China with a high incidence of cervical carcinoma. A cross-sectional,comparative study

OBJECTIVE: To confirm the accuracy of the ThinPrep Pap Test (Cytyc Corp., Boxborough, Massachusetts, U.S.A.) on the basis of histologic diagnosis by biopsy and the detection of human papillomavirus (HPV) DNA in mass screening. STUDY DESIGN: A total of 1,997 women residing in Xiangyuan County, Shanxi Province, P.R.C., an area with a high incidence of cervical carcinoma, were enrolled in this study. We collected exfoliative cervical samples from all subjects into a liquid buffer (Preserv-Cyt [Cytyc]) and utilized for both cytologic screening using the ThinPrep Pap Test and HPV DNA testing. Subsequent colposcopic biopsies were taken on all subjects. All the tests were performed in an independent and blinded fashion. We compared the ThinPrep Pap test with colposcopic biopsy and HPV test. RESULTS: High grade squamous intraepithelial lesions (HSIL) (CIN 2/3) were identified in 74 (3.7%) of 1,993 women adequately assessed, and there were 12 cases of squamous cell carcinoma (SCC). The false negative rate of ThinPrep cytology was 3.2% for biopsy-confirmed CIN 3 and 9.3% for CIN 2. Twenty-seven (87%) of the 31 women with biopsy-confirmed CIN 3 and 12 (100%) of 12 with biopsy-confirmed SCC had a diagnosis of either HSIL or greater abnormalities on ThinPrep cytology. In addition, the HPV DNA detection rates offered a good correlation between cytology and biopsy. CONCLUSION: The ThinPrep Pap performed extremely well in this primary screening trial. We found a good correlation between ThinPrep cytology and colposcopic biopsy on detection of HSIL and SCC; cervical specimens collected in ThinPrep liquid buffer serve as a direct test for HPV as well.     

Prevalence of human papillomavirus infection and genotype distribution determined by the cyclic-catcher melting temperature analysis in Korean medical checkup population

Although cytology screening has reduced the incidence and mortality rate of cervical cancer significantly, its usefulness is limited to samples from the site of the lesion, resulting in its low sensitivity and unsuitability for use in medical checkups. The purpose of the present study was to evaluate the prevalence of HPV infection using genotype distribution and to analyze the correlation of the HPV DNA test results with cytological results. We also evaluated the benefits of quantitative information obtained from cyclic-catcher melting temperature analysis (CMTA) in screening for cervical cancer. We performed cyclic-CMTA using Anyplex? II HPV28 Detection in combination with cervical cytology for 2,181 subjects. The following HPV positivity types were detected using cyclic-CMTA and HPV positivity was found to increase together with the severity of the cytology results: (1) For 419 HPV positive specimens, HPV DNA was detected in 18.1% of normal specimens, 78.3% of ASCUS, and all of LSIL and HSIL; (2) high-risk HPV DNAs were detected in 63.3% of normal (N=547), 65.9% of ASCUS (N=41), 76.9% of LSIL (N=13), and 88.9% of HSIL (N=9) among total detected HPV DNA regardless multiple detection; (3) multiple HPV genotypes were detected in 4.8% of normal specimens (N=2,146), 52.2% of ASCUS (N=23), 57.1% of LSIL (N=7), and 40.0% of HSIL (N=5). In addition, a high level of viral DNA was observed using cyclic-CMTA in all specimens beyond the LSIL stage according to cytology, while only 6% of specimens with normal cytology showed a correlation with viral quantitation by cyclic-CMTA. The combination of HPV genotyping with a quantitative assay and cytology will allow for a more accurate diagnosis of cervical cancer.     

Experience with a thin-layer,liquid-based cervical cytologic screening method

OBJECTIVE: To assess the utility of a thin-layer cytology system for cervicovaginal screening in clinical practice. STUDY DESIGN: Twenty-five hundred cervicovaginal split samples were processed with the traditional direct smearing method and with the ThinPrep Pap Test (Cytyc Corp., Boxborough, Massachusetts, U.S.A.) method and evaluated according to the Bethesda system, focusing mainly on the cytomorphologic features. RESULTS: The ThinPrep Pap Test yielded improved specimen adequacy and an increased detection rate of squamous abnormalities, which resulted in a decrease in the ASCUS/SIL ratio. Moreover, the thin-layer system provided adequate material for concomitant HPV testing, mainly in the LSIL and the ASCUS favor SIL cases, with satisfactory results, as well as for cell block preparations in a few selected cases that presented diagnostic difficulties. Furthermore, the morphologic features of the LSIL cases were virtually identical on both preparations, while in the HSIL cases, distinct features were noted on ThinPrep. CONCLUSION: The ThinPrep Pap Test is significantly more effective than the conventional smear in clinical practice. However, training and experience are necessary to take full advantage of this promising new technology.     

Outcomes of women with atypical squamous cells of undetermined significance and high-risk human papillomavirus DNA

OBJECTIVE: To study the risk of high-grade squamous intraepithelial lesion (HSIL) and cervical intraepithelial neoplasia (CIN) 2 or 3 in women with human papillomavirus (HPV)-positive atypical squamous cells of undetermined significance (ASCUS) cytology over a 2-year period using the 2001 Bethesda System and ThinPrep Paps. STUDY DESIGN: In 2002, 846 patients with ThinPrep cervical cytology having an ASCUS interpretation and positive for high-risk HPV DNA were identified. A cohort of 514 (60.8%) patients with follow-up by repeat cytology, cervical biopsy or both was included in the study. Patient age was 12-81 years, with a median of 25 years. RESULTS: There were 291 women (56.6%) with negative status by cytology, HPV testing or biopsy with a median interval of 8.5 months, and an additional 174 patients (33.9%) had persistent ASCUS, positive HPV DNA or low-grade SIL/CIN 1. Finally, 49 patients (9.5%) had CIN 2 or 3, with a median interval of 8.5 months. CONCLUSION: Our study suggests that HSIL or CIN 2 or 3 will be detected in 1 in 10 women with HPV-positive index ASCUS cervical cytology at initial colposcopy or within a 2-year follow-up period.     

Procedure for immunocytochemical detection of P16INK4A antigen in thin-layer, liquid-based specimens

OBJECTIVE: To develop a procedure for the immunocytochemical detection of P16INK4A in ThinPrep specimens. STUDY DESIGN: Archived ThinPrep, liquid-based cervical/endocervical cytology specimens (Cytyc Corp., Boxborough, Massachusetts, U.S.A.) diagnosed as LSIL, HSIL and WNL were resampled and fixed in 95% ethanol for at least three days. Rehydration and endogenous peroxidase blocking of both ThinPreps and formalin-fixed, paraffin-embedded tissues were accomplished on a Leica Autostainer (Leica, Deerfield, Illinois, U.S.A.). Microwave antigen retrieval with CitraPlus (Biogenex, San Ramon, California, U.S.A.) was performed using a Panasonic microwave oven (Matsushita Cooking Appliances, Franklin Park, Illinois, U.S.A.) on the high setting twice for five minutes each. After cooling for 20 minutes and undergoing a buffer rinse, the slides were placed in a Dako autostainer (Dako-USA, Carpinteria, California, U.S.A.). The P16INK4A primary antibody, clone E6H4 (MTM Laboratories, Heidelberg, Germany) was diluted 1:200 in antibody diluent buffer. Detection was accomplished with a mouse non-avidin-biotin EnVision+ polymer (Dako). The expression of P16INK4A in ThinPreps and corresponding biopsies were scored by two pathologists. A ThinPrep case was scored as positive if it contained > 10 abnormal cells with nuclear and cytoplasmic immunocytochemical staining. Corresponding biopsies were scored as exhibiting negative, sporadic, focal or diffuse staining, as described by Klaes et al, Overexpression of P16INK4A as specific marker for dysplastic and neoplastic epithelial cells of the cervix uteri (Int J Cancer 2001;92:276-284). RESULTS: The P16INK4A antibody assay was positive in 14 of 19 (73.68%) LSIL ThinPrep cases and in 25 of 26 (96.15%) HSIL ThinPrep cases. Thirty-eight of the 39 (97.44%) biopsies corresponding to the positively stained ThinPreps also were positive, with a staining score of at least focal positivity in the dysplastic regions. The P16INK4A antibody assay was negative in 5 of 19 (26.32%) LSIL ThinPrep cases and negative in 1 of 26 (3.85%) HSIL ThinPrep cases. The six biopsies corresponding to the negative ThinPreps were similarly negative. The two cytologic specimens diagnosed as WNL were negative for P16INK4A, as were two tissue control cases with benign diagnoses. Nondysplastic squamous epithelium, identified in 17 biopsy cases, did not stain, nor did nondysplastic squamous cells identified in ThinPrep cases. Sporadic staining of bacteria, inflammatory cells and occasional endocervical glandular cells was identified. CONCLUSION: P16INK4A expression in ThinPrep specimens correlates with tissue expression of P16INK4A, as implemented in the above protocol. P16INK4A may thus serve as a surrogate marker in gynecologic cytology for high-risk HPV infection and for the development of cervical neoplasia.     

Detection of HPV DNA in cervical specimens collected in cytologic solution by ligation-dependent PCR

OBJECTIVE: To determine the feasibility and sensitivity of detecting human papillomavirus (HPV) in specimens collected in Cytyc PreservCyt fluid (Boxborough, Massachusetts, U.S.A.) using ligation-dependent polymerase chain reaction (LD-PCR) and to demonstrate the diagnostic value of HPV DNA testing as an adjunct to cytology in the detection of cervical squamous intraepithelial lesions (SIL), especially in cases of atypical squamous cells of undetermined significance (ASCUS). STUDY DESIGN: LD-PCR is a recently invented DNA amplification technology that utilizes a capture probe for target isolation and 2 hemiprobes for target detection. The hemiprobes are designed in such a way that when they hybridize to their target, the 5' end of one probe and the 3' end of the other probe are brought together. Two hemiprobes can then be ligated into a full probe that can serve as a template for PCR amplification. A total of 94 cervical specimens were collected in cytologic fluid and tested with LD-PCR. The results were compared with those of the Digene Hybrid Capture II assay (HC II) (Beltville, Maryland, U.S.A.) and consensus PCR. RESULTS: The overall sensitivity for detecting HPV was 41.5% (39/94) by LD-PCR, 50% (47/94) by consensus PCR and 37.2% (35/94) by HC II. The prevalence of HPV by HC II, consensus PCR and LD-PCR were 87.5%, 100% and 87.5% in the high grade SIL group; 100%, 90.9% and 90.9% in the low grade SIL group; 30%, 52.5% and 40% in the ASCUS group; and 14.2%, 22.8% and 17.1% in women with normal cytology. These results indicate that all 3 methods have similar sensitivity in patients with SIL. However, there is greater variation in detection rates in the ASCUS and normal cytology groups. CONCLUSION: LD-PCR is a useful method of detecting HPV in liquid-based gynecologic cytologic preservatives, and HPV testing as a method adjunct to the liquid-based Pap test could be useful in detecting SILs, especially for the management of patients with ASCUS.     

Prevalence and typing of HPV DNA in atypical squamous cells in pregnant women

OBJECTIVE: To determine the prevalence and typing of HPV DNA in pregnant women with a diagnosis of atypical squamous cells (ASC) and to assess whether pregnancy-related changes contribute to the diagnosis of ASC. STUDY DESIGN: HPV testing was performed on residual specimens from the ThinPrep Pap test (Cytyc Corp., Boxborough, Massachusetts, U.S.A.) in pregnant women diagnosed as ASC (study group, n = 105), low and high grade squamous intraepithelial lesion (LSIL and HSIL) (positive control, n = 33) and negative for epithelial cell abnormality (negative control, n = 20). All cases were reviewed by 2 cytopathologists to obtain consensus diagnoses using the Bethesda System 2001 criteria. The study group cases were further subcategorized into ASC of undetermined significance (ASCUS, n = 99) and ASC cannot exclude HSIL (ASC-H, n = 6). HPV testing was also performed on an ASC control group consisting of 68 consecutive ASC cases in nonpregnant women, matched by age. RESULTS: Mean patient age was 23.7 years for the study group and 25.6 years for the ASC control group. HPV DNA was detected in 88.6% of cases in the study group, including 87.9% of ASC-US and 100% of ASC-H cases. Of the HPV positive cases, 79.6%, 4.3%, 5.4% and 10.8% had high-risk, mixed high- and low-risk, low-risk and unknown HPV types, respectively. The most frequent HPV types detected were: types 52 (31.2%), 16 (15.1%), 39 (11.8%), 53 (10.8%), and 18 and 58 (9.7% each). Multiple viral types were detected in 43.0% of cases. The prevalence of HPV DNA in the positive and negative controls in pregnant women was 100% and 55%, respectively. HPV DNA was detected in 83.8% of the ASC control group. CONCLUSION: Regardless of pregnancy-related changes, the prevalence of HPV DNA in pregnant women (88.6%) was similar to that found in ASC in nonpregnant women of the same reproductive-age group (83.8%), and the high-risk types accounted for the vast majority of cases (83.9%). These findings demonstrate that pregnancy-related changes do not contribute to the diagnosis of ASC in this subset of women. Furthermore, the high HPV DNA prevalence in reproductive-age women (< 40 years) suggests that HPV testing may have limited utility in effective management of these patients.     

Comparative study of the ThinPrep Pap test and conventional cytology results in a Canadian cohort

OBJECTIVE: To compare the frequency of Pap test results in a prospective series of direct to vial ThinPrep tests to a cohort of conventionally prepared tests. To follow-up all test results for a minimum of 2 years and assess performance based on this outcome. METHODS: All women presenting for either routine screening or colposcopic examination in 2001 were enrolled in the ThinPrep cohort. A similar, population of conventionally prepared tests was extracted from the year 2000 laboratory data. Information on all concurrent and follow-up cervical specimens over the ensuing 2 years was retrieved. RESULTS: The ThinPrep cohort comprised 2288 Pap tests and the conventional, 2211. The frequency of normal [within normal limits (WNL) and benign cellular changes (BCC)] results in the ThinPrep cohort was 6% lower and the frequency of abnormal [> or =atypical squamous cells of undetermined significance (ASCUS)] results was 6.8% higher. Respective ThinPrep and conventional cohort results were 1156 (51%) and 1291 (58%) WNL, 625 (27%) and 561 (25%) BCC, 101 (4%) and 65 (3%) ASCUS, 21 (1%) and 2 (0.1%) atypical glandular cells of undetermined significance, 301 (13%) and 224 (10%) low-grade squamous intraepithelial lesion (LSIL), and 74 (3%) and 40 (2%) high-grade SIL (HSIL) (P < 0.0001). Follow-up was available for nearly 80% of each cohort. LSIL or higher was confirmed in 57.5% (n = 266) of the abnormal ThinPrep and 60.9% (n = 190) of the abnormal conventional tests. The ThinPrep yield of confirmed tests however was almost 50% higher than the conventional test. CONCLUSION: In this population, ThinPrep was superior to the conventional Pap test.     

P16INK4A as an adjunct test in liquid-based cytology

OBJECTIVE: To assess the utility of P16INK4A as an adjunct test in liquid-based cytology in cases with equivocal morphologic changes of high grade squamous intraepithelial lesion (HSIL). STUDY DESIGN: P16INK4A immunoreactivity was investigated in residual ThinPrep material (Cytyc Corp., Boxborough, Massachusetts, U.S.A.) from 30 cases with equivocal diagnoses of HSIL that had corresponding follow-up biopsies. Two control ThinPrep cases were included: 1 HSIL with biopsy-confirmed cervical intraepithelial neoplasia (CIN) 3 and a negative specimen with a corresponding biopsy of squamous metaplasia. The expression of P16INK4A in ThinPrep specimens and corresponding biopsies was scored as previously described. A ThinPrep case was scored positive if it contained > 10 abnormal cells with nuclear and cytoplasmic immunocytochemical staining. Corresponding biopsies were scored as having negative, sporadic, focal or diffuse staining. RESULTS: The P16INK4A antibody assay was positive in 19 of 30 ThinPrep cases (63.3%). Seventeen of the 19 (89.4%) biopsies corresponding to the positively stained ThinPreps also were positive, with a score of at least focal positivity in the dysplastic regions (2 CIN 1, 4 CIN 2, 11 CIN 3; 2 lesions lost in the tissue recut). The assay was negative in 11 ThinPreps (36.6%) and 10 biopsies (33.3%) with tissue confirmation of chronic cervicitis (5), squamous metaplasia (2), CIN 1 (3) and 1 lesion lost in the tissue recut. Seventeen of 18 (94.4%) ThinPreps confirmed as high grade lesions upon biopsy showed P16INK4A positivity. The control HSIL case with a CIN 3 biopsy was diffusely positive for P16INK4A, and the control negative case with biopsy diagnosis of squamous metaplasia was negative. Nondysplastic squamous and metaplastic epithelium in 7 biopsies and nondysplastic squamous or metaplastic cells in ThinPrep cases were negative. Sporadic staining of bacteria, inflammatory cells and endocervical cells was noted. CONCLUSION: ThinPrep cases in the equivocal cytologic category with the corresponding tissue biopsy assayed for P16INK4A expression showed that there was utility for this type of testing. A larger series comparing corresponding ThinPrep and tissue biopsies will be undertaken. The role of HPV infection in these cases will also be explored.     

A human papillomavirus testing system in women with abnormal Pap results: a comparison study with follow-up biopsies

OBJECTIVE: To evaluate the efficacy of INFORM HPV using the SurePath collection method in women whose Pap tests indicated abnormal results. STUDY DESIGN: Ninety-two women from the gynecology clinics at The University of Texas M.D. Anderson Cancer Center who had Pap tests and underwent follow-up biopsies were selected for the study. This included 51 women with atypical squamous cells of undetermined significance (ASCUS), 23 women with low-grade squamous intraepithelial lesion (LSIL), 15 women with high-grade squamous intraepithelial lesion (HSIL) and 3 women with negative Pap results. The INFORM HPV, an in situ hybridization assay, testing for oncogenic types of HPV was performed, and the results were compared with follow-up biopsies. RESULTS: The positive rate of the INFORM HPV increased with higher grades of cytology diagnoses. The sensitivity of the INFORM HPV testing for predicting high-grade cervical intraepithelial neoplasia (CIN 2/3) also increased with higher grades of cytology diagnoses. A negative predictive value (NPV) of 94.9% and a specificity of 80.4% for predicting CIN 2/3 were observed in the ASCUS group. CONCLUSION: Using SurePath Pap specimens, the INFORM HPV lacks sufficient sensitivity and NPV for predicting CIN 2/3 in women with ASCUS. Therefore, use of the test as a triage tool is limited.     

Rapid detection of HSV from cytologic specimens collected into ThinPrep fixative

OBJECTIVE: Herpes simplex virus (HSV) infection is associated with substantial morbidity and mortality in neonates. A diagnosis of HSV on cervical cytologic studies could lead to a cesarean section, with an increase in the risk of maternal morbidity. The identification of viral lesions in sexually active women has medical and social implications. There have been reports of false positive diagnoses of HSV in patients with altered endocervical cells and with cervical intraepithelial neoplasia 3. We evaluated a polymerase chain reaction (PCR)-based assay to detect HSV-1 and HSV-2 in routinely collected cervical cytology specimens in ThinPrep fixative (Cytyc Corp., Marlborough, Massachussets, U.S.A.). STUDY DESIGN: DNA was extracted from five cases that demonstrated cytologic changes suggestive of an HSV infection. PCR amplification with subsequent gel electrophoresis was performed to detect the presence of HSV. RESULTS: HSV DNA was detected in three of five cases that were cytologically diagnosed as suspicious or strongly suspicious for HSV infection. CONCLUSION: The combination of the ThinPrep liquid-based method for cervical cytology with PCR allows prompt confirmation of the diagnosis of HSV without sacrificing the diagnostic morphology on the slide.     

Significance of high-risk human papillomavirus DNA-positive atypical squamous cells of undetermined significance pap smears in perimenopausal and postmenopausal women

OBJECTIVE: To determine the risk of high-grade squamous intraepithelial lesion (HSIL)/cervical intraepithelial neoplasia (CIN) 2 or 3 in perimenopausal and postmenopausal women with high-risk human papillomavirus (HPV) DNA-positive atypical squamous cells of undetermined significance (ASCUS) on Pap tests. STUDY DESIGN: A total of 752 perimenopausal and postmenopausal women (patients 45 years) with an ASCUS interpretation on ThinPrep Pap tests were identified in 2002 and 2003. High risk HPVDNA was detected in 191 (25.4%) women. The follow-up results in these 136 women were compared with those of women <45 years. RESULTS: Of the 136 perimenopausal and postmenopausal women, 64.7% became negative on cytology or biopsy and 30.9% revealed persistent low-grade SIL (LSIL)/CIN 1. Furthermore, 5 women (3.68%) were diagnosed with HSIL/CIN 2 or 3, and 1 patient had squamous cell carcinoma (0.74%). The rate of HSIL/CIN 2 or 3 or worse detected in perimenopausal and postmenopausal women was significant lower than 10.6% in women <45 years (p = 0.028). CONCLUSION: Our study showed that perimenopausal and postmenopausal women with high-risk HPV positivity and an ASCUS interpretation have a significantly lower risk of HSIL/CIN 2 or 3 detected. Nonetheless, the risk of a significant finding on workup, including invasive carcinoma, indicates the need for colposcopic evaluation of these women.     

ASCUS-LSIL Triage Study. Design, methods and characteristics of trial participants

OBJECTIVE: To describe the design and methods of the ASCUS-LSIL Triage Study (ALTS), a multicenter, randomized clinical trial designed to evaluate three alternative methods of managing low grade (LSIL) and equivocal (ASCUS) cervical cytologic diagnoses. STUDY DESIGN: Nonpregnant women, 18+ years old, with ASCUS or LSIL, no prior hysterectomy or ablative therapy to the cervix, were referred to one of four clinical centers around the United States. Eligible and consenting participants were administered a risk-factor questionnaire and underwent a pelvic examination, collection of cervical specimens for liquid-based cytology and human papillomavirus (HPV) testing and Cervicography (National Testing Laboratories, Fenton, Missouri, U.S.A.). Patients were randomized to one of three arms: (1) immediate referral for colposcopy at enrollment, (2) follow-up with cytology only, and (3) use of HPV DNA testing to triage to colposcopy. All women are followed every six months for two years with pelvic examinations, cytologic and masked HPV testing, and masked Cervicography. Digital cervical images and cytology and histology slides are externally reviewed to maximize patient safety. RESULTS: We enrolled and randomized 3,488 eligible women with ASCUS and 1,572 women with LSIL. CONCLUSION: The successful enrollment, randomization and high rates of follow-up are encouraging. The study will help clarify the optimal strategies for managing low grade cervical abnormalities.     

Triage of women with equivocal or low-grade cervical cytology results: a meta-analysis of the HPV test positivity rate

Consistent evidence underlines the utility of human papillomavirus (HPV) DNA testing in the management of women with equivocal cervical cytological abnormalities, but not in case of low-grade lesions. We performed a meta-analysis including studies where the high-risk probe of the Hybrid Capture-II is used to triage these two cytological categories. The triage test-positivity rate reflects the colposcopy referral workload.Data were pooled on the HPV test positivity rate in women with atypical squamous cells of undetermined significance (ASCUS/ASC-US) or low-grade squamous intraepithelial lesions (LSIL), derived from different cytological classification systems. The meta-analysis was restricted to studies, published between 1991 and 2007. A random-effect model was applied for meta-analytical pooling and the influence of covariates on the HPV positivity rate was analyzed by meta-regression. The variation by age was assessed within individual studies since age strata were not defined uniformly. On an average, 43% (95% CI: 40–46%) of women with ASCUS/ASC-US were high-risk HPV positive (range 23–74%). In women with LSIL, the pooled positivity rate was 76% (95% CI: 71–81%; range 55–89%). In spite of considerable inter-study heterogeneity, the difference in HPV positivity between the two triage groups was large and highly significant: 32% (95% CI: 27–38%). HPV rates dropped tremendously as age and cutoffs of test positivity increased. Other factors (cytological classification system, country, continent, collection method and year of publication) had no statistically significant impact, except in LSIL triage where HPV positivity was significantly lower in European compared to American studies. Women with LSIL, especially younger women, have high HPV positivity rates suggesting limited utility of reflex HPV triaging these cases. Research is needed to identify more specific methods to triage women with low-grade squamous cervical lesions.     

Utility of HPV DNA detection in thin-layer,liquid-based tests with atypical squamous metaplasia

OBJECTIVE: To determine if testing for HPV is useful in the management of patients with atypical squamous metaplasia (ASM) and to evaluate a small group of patients with atypical squamous cells in the setting of an atrophic cellular profile (estrogen test [ESTT]). STUDY DESIGN: Presence of HPV DNA was determined on 104 ASM and ESTT cases on residual ThinPrep specimens using Hybrid Capture II. Results of the HPV DNA test were correlated with subsequent biopsy or repeat Pap test results. RESULTS: Of 63 patients with ASM on ThinPrep Pap tests, 52% were associated with histologically proven SIL when HPV DNA was detected by Hybrid Capture II. None of the 17 patients who were negative for HPV DNA had subsequent evidence of HPV infection by biopsy or repeat Pap test. Among 14 patients with ESTT, none of the 12 who were negative for HPV DNA had subsequent evidence of HPV infection. CONCLUSION: ASM and ESTT with a negative HPV DNA test can be followed routinely, and a colposcopic examination is not warranted.     

Performance of monolayered cervical smears in a gynecology outpatient setting in Kuwait

OBJECTIVE: To compare ThinPrep (TP) Papanicolaou smears (Cytyc Corp., Box-borough, Massachusetts, U.S.A.) with matching conventional Papanicolaou (CP) smears for specimen adequacy, cytologic quality, diagnostic accuracy and screening time. STUDY DESIGN: In this prospective study of 1,024 women a split-sample, matched-pair design in favor of CP slides based on single-blind criteria was followed with a smear on a glass slide for CP and the remaining material collected in Preserv-Cyt solution (Cytyc) for a TP smear. A Cytobrush (Medscand, Hollywood, Florida, U.S.A.) was used for smear preparation for CP. TP smears were processed in ThinPrep 2000 (Cytyc). Smears were stained with Papanicolaou stain and were interpreted according to the Bethesda system. RESULTS: The number of satisfactory but limited (SBL) cases with TP were 77 (7.5%) as compared to 127 (12.4%) with the CP method. This reduction in SBL smears with the TP method and consequent increase in satisfactory smears were highly significant (P < .001) by McNemar's test. As regards unsatisfactory smears in discordant pairs, although the number of unsatisfactory smears was higher with TP (41 cases) as against CP (27 cases), the difference was not statistically significant (P < .05). The split-sample method showed a high correlation between the CP and TP diagnoses. TP smears had a significant advantage over CP smears in the reduction in the number of ASCUS and AGUS cases (14 vs. 29) (P < .05) and increased the pickup rate of LSIL, 6 vs. 1. Time taken to screen the TP smears was half that of CP smears. No cases of LSIL or HSIL were missed on TP smears. CONCLUSION: The liquid-based processor significantly improved the adequacy and quality of smears, resulting in fewer recall cases for SBL smears, leading to more definitive diagnoses in atypical cases, increasing the pickup rate of LSILs and reducing the screening time. A machine handling multiple specimens automatically would decrease cost and be an asset to a cytopathology laboratory.     

Prevalence of human papillomavirus among Croatian women attending regular gynecological visit

Human papillomavirus (HPV) infection has been identified as major risk factor for cervical intraepithelial neoplasia (CIN) and invasive cervical cancer. About 40 HPV viral types are commonly found in the genital tract. Most HPV infections resolve spontaneously, while persistent infection with oncogenic types, namely HPV 16 and 18 is necessary for CIN to occur and progress to cancer. Cervical screening is presently based on the Pap smear that is designed to diagnose precancerous lesions and cervical cancer The aim of this study was to investigate the prevalence of HPV DNA and to determine HPV types distribution among 361 women attending regular gynecological visit. There were 205 women (29+/-8 years old) without determined abnormal cervical lesions and 156 women (34+/-15 years old) with abnormal Pap smear; low grade squamous intraepitehelial lesions (LSIL, n=69), high grade squamous intraepithelial lesions (HSIL, n=72) and atypical squamous cells of undetermined significance (ASCUS, n=15). HPV DNA detection and genotyping was performed by Hybrid Capture 2 assay and additionally by consensus and type-specific primers directed PCR. The overall prevalence of high-risk HPV (hrHPV) in women with abnormal Pap smears was 67.9% (106/156), of which in ASCUS 33.4% (5/15), LSIL 62.3% (43/69) and HSIL 80.6% (58/72). In HPV positive specimens, HPV 16 was found as predominant type in 60.4% cases, followed by HPV 31 (8.5%), HPV 33 (6.6%) and HPV 18 (3.7%). In the group of women without obvious cervical changes the overall hrHPV prevalence was 35.6% with HPV 16 found in 43.8% cases, followed by HPV 31 (17.8%), HPV33 (9.5%) and HPV18 (6.8%). In both study groups, women with and without cervical lesions, the prevalence of HPVof indeterminate type was 14.2% and 13.7%, respectively. Our results indicate that cervical intraepithelial lesions are largely associated with HPV type 16, followed by HPV types 31, 33, 18 and HPV of indeterminate type. Although there is a significant difference in hrHPV DNA prevalence among two groups, no significant differences between particular hrHPV types distribution were observed.     

Newly developed liquid-based cytology. TACAS™: cytological appearance and HPV testing using liquid-based sample

Cell profiles determined by the thin-layer advanced cytology assay system (TACAS?), a liquid-based cytology technique newly developed in Japan, were analyzed in this study. Hybrid capture 2 (HC-2) was also performed using the liquid-based samples prepared by TACAS to ascertain its ability to detect human papillomavirus (HPV). Cell collection samples from uterine cervix were obtained from 359 patients and examined cytologically. A HC-2 assay for HPV was carried out in the cell specimens. All specimens were found to show background factors such as leukocytes. After excluding the 5 unsatisfactory cases from the total 354 cases, 82 cases (23.2%) were positive and 272 cases (76.8%) were negative for HPV. Cell specimens from 30 HPV-positive cases and 166 HPV-negative cases were subjected to 4 weeks of preservation at room temperature. Then, when subsequently re-assayed, 28 cases (93.3%) in the former group were found to be HPV positive and 164 cases (98.8%) in the latter group were found to be HPV negative. These results supported the excellent reproducibility of TACAS for HPV testing. A reasonable inference from the foregoing analysis is that TACAS may be distinguished from other liquid-based cytological approaches, such as ThinPrep and SurePath, in that it can retain the cell backgrounds. Furthermore, this study raises the possibility that cell specimens prepared using TACAS could be preserved for at least 4 weeks prior to carrying out a HC-2 assay for HPV.     

Can we detect cervical human papillomavirus (HPV) infection by cytomorphology alone? Diagnostic value of non‐classic cytological signs of HPV effect in minimally abnormal Pap tests

OBJECTIVE: Our aim was to assess the validity of non-classical cytological signs in minimally abnormal cervical smears for the prediction of HPV infection. METHODS: 164 ThinPrep monolayers were re-screened for mild nuclear changes, disorders of keratinisation, abortive koilocytes and 'measles cells', as well as degenerative changes. HPV DNA was detected by GP5+/6+ and MY09/MY11 consensus primer PCR assays. RESULTS: Seventy six of 164 cases (46.3%) had HPV positivity by PCR. All cytomorphological features studied were significantly associated with the presence of HPV. Mild nuclear changes had 100% sensitivity and 100% negative predictive value for HPV infection. CONCLUSIONS: Our results indicate that non-classic cytomorphological signs can improve the sensitivity of cytology for detecting HPV. Minimally abnormal Pap smears lacking mild nuclear changes (16%) in the present study--do not require further molecular HPV testing.