Prevalence of high-grade cervical intraepithelial neoplasia in women with persistent high-risk HPV genotypes and negative cytology.

INTRODUCTION: Primary HPV screening will be implemented into the English Cervical Screening Programme by 2019. Its impact upon women referred to colposcopy, with negative cytology but persistently positive high-risk HPV (hrHPV), remains unreported from UK Sentinel sites. HPV primary screening was introduced in Sheffield, UK in April 2013; this paper reports its impact on the service. METHODS: A retrospective cohort study was performed from June 2014 to July 2016 at the Jessop Wing Colposcopy Unit, Sheffield. UK. Data were obtained from the pathology and colposcopy databases and cross-referenced with case-notes and pathology results for women referred with persistently positive hrHPV, cytology negative samples. Patient demographics, hrHPV genotype, biopsy rates, histological diagnoses, management, and outcomes were collected and baseline statistics performed. RESULTS: During the study 1076 women were seen. Most frequent hrHPV genotypes were: hrHPV other, 41%; and HPV16, 33%. The majority (72%) were found to have normal colposcopy; 28% had an abnormal colposcopic assessment (11% low-grade; 11% high-grade; 6% inadequate). The majority were discharged (83%) and only 5% underwent LLETZ. No cancers were detected. High-grade cervical intraepithelial neoplasia (CIN) was found in 7%; overall risk of CIN2 was 1/29; 1/30 for CIN3. Presence of HPV16 was associated with a significantly higher risk of high-grade CIN; 1/9. CONCLUSION: This is the first study to report results for women referred to colposcopy with cytology negative, persistently positive hrHPV. Disease prevalence is low, although women with HPV16 have a significantly higher likelihood of high-grade disease compared to other HPV subtypes.

Eight cases of malignant mixed Müllerian tumor (carcinosarcoma) of the uterus: findings in SurePath™ cervical cytology.

Malignant mixed Müllerian tumor (MMMT)/carcinosarcoma is a rare neoplasm of the female genital tract characterized by a mixture of epithelial and mesenchymal components. There are published reports of conventional cervical smear findings in uterine MMMT, but to the best of our knowledge the cytomorphology of MMMT in SurePath™ liquid-based cytology samples has not been described. We present a series of eight cases of uterine MMMT in SurePath™ cervical samples. The mean age of the women was 65.5 years (range, 60-82 years) and the most common presenting symptom was postmenopausal bleeding. Three out of the eight cases had both epithelial and sarcomatous elements in highly cellular cervical samples. In two cases, abnormal glandular cell clusters were closely associated with the mesenchymal component. The glandular component was usually represented by three-dimensional clusters of malignant glandular cells and a dispersed population of large, round to oval malignant epithelial cells with single prominent nucleoli. Necrosis or tumor diathesis was not seen in any cases. Six out of the eight cases were initially reported as adenocarcinoma, endometrial type, and two cases as normal. Five cases had an atrophic background and tiny atypical glandular clusters that could easily be overlooked. Although cervical sample is not the method of choice for primary uterine carcinosarcoma screening, the possibility of the same should be kept in mind, when reporting a cervical sample from a woman with postmenopausal vaginal bleeding.

Atypical squamous cells, cannot exclude high-grade squamous intraepithelial lesion: diagnostic features in surepath™ cervical samples.

This study was undertaken to identify the situations in which a diagnosis of "Atypical squamous cells, cannot exclude a high-grade squamous intraepithelial lesion (ASC-H)" is offered in SurePath™ cervical samples and to identify cytological criteria helpful in predicting high-grade disease. 2,335 (3.4%) SurePath samples reported as atypical squamous cells (ASC) over a period of 2 years, including 1,112 cases with known hrHPV status were retrieved. 105/1,112 cases were categorized into ASC-H, and slides were available for review in 88/105 cases. These 88 samples were divided into two categories based on follow-up histological outcome and hrHPV status-category A: cases with CIN2+ lesions on follow-up (n = 48) and category B: cases with ≤CIN1 lesions or hrHPV negative status (n = 40). 78% (82/105) cases of ASC-H tested positive for hrHPV. Overall CIN2+ lesions were found in 50.3% (53/105) cases. Of 88 cases reviewed, HCGs were noted in 56.3% (27/48) cases in category A and 75% (30/40) cases in category B. Dispersed metaplastic cells and scattered small atypical cells were seen in 37.5% (18/48) cases in category A and 12.5%(5/40) in category B. The majority of cases with dispersed atypical cells had <20 cells/sample and cases with HCGs had <10 HCGs per sample. The majority of the cases reported as ASC-H contained HCGs. Of these groups with nuclear crowding, disorganization and those with steep edges ("blocks") are likely to predict high-grade disease. The samples with only dispersed atypical cells had <20 cells/sample in majority of cases. In these, a disproportionate and especially high nuclear: cytoplasmic ratio and irregular chromatin were the most useful features in predicting high-grade disease.

Factors contributing to false-negative and potential false-negative cytology reports in SurePath™ liquid-based cervical cytology.

OBJECTIVES: The characteristics of false-negative conventional cervical cytology smears have been well documented, but there is limited literature available for liquid-based cytology (LBC), especially SurePath™ samples. We aimed to assess the characteristics of false-negative SurePath LBC samples. METHODS: Over a period of 5 years, an audit of false-negative reports in SurePath cervical cytology was undertaken. In a workload of 183, 112 samples, 481 (0.3%) false negatives were identified using two routes: those detected by routine laboratory internal quality control (rapid pre-screening) (n = 463) and those reported as normal (true false negatives) with concurrent high-grade cervical histology (n = 18). Ninety-five false-negative cases with a subsequent biopsy reported as at least cervical intraepithelial neoplasia grade 2 (CIN2+) were reviewed for a number of different cytomorphological features. RESULTS: Of 95 samples with subsequent CIN2+, 30.5% predominately contained microbiopsies/hyperchromatic crowded cell groups (HCGs), 27.3% sparse dyskarytotic cells, 4.2% pale cell dyskaryosis, 6.3% small dyskaryotic cells; 3.2% were misinterpreted cells, 8.4% contained other distracting cells, 7.4% were low contrast, 5.3% were unexplained and 7.4% were true negatives. The mean number of microbiopsies/HCGs in that category was 4.6. The mean number of abnormal cells in the sparse dyskaryotic cell category was 13.8. CONCLUSIONS: Microbiopsies/HCGs were the commonest reason for false negatives. They were usually present in sufficient numbers to be detected but interpretation could be problematic. Dispersed single abnormal cells were usually not identified because of their scarcity or the presence of distracters.

ABC3 Part I: a review of the guidelines for terminology, classification and management of cervical cytology in England.

The provision of guidance on cytology reporting and evaluation, first outlined in 1995 with the publication of Achievable Standards, Benchmarks for Reporting, and Criteria for Evaluating Cervical Cytopathology (ABC), and subsequently revised and expanded in a second edition in 2000, has been pivotal to the success of the National Health Service Cervical Screening Programme (NHSCSP), ensuring that standards are upheld, and that rigorous evaluation and quality assurance take place. In the last decade, major changes to the NHSCSP, notably the adoption of revised age ranges and screening intervals for all women in England, implementation of liquid-based cytology and, most recently, the decision to introduce high-risk human papillomavirus (HR-HPV) testing for triage of low-grade and borderline (equivalent to 'atypical') cytological abnormalities and test of cure after treatment of cervical intraepithelial neoplasia (CIN) determined that an updated version of ABC was required. The third edition of ABC recommends adoption, with minor modification, of the revised British Society for Clinical Cytology terminology and provides guidance on the management of abnormal cytology results linked to this terminology taking account of HR-HPV testing. To accommodate these changes, expanded result codes, which are electronic codes used to transfer management information to central computers for follow-up, call and recall of individual women, have been developed. Further guidance on specimen adequacy is also provided. Revised performance indicators are described and explained in a separate article by R. Blanks in this issue of Cytopathology. All the changes in ABC3 are designed to support the mission statement of the NHSCSP that 'the objective of cervical screening is to reduce cervical cancer incidence and mortality by screening with a high sensitivity for the detection of CIN2 or worse, whilst maintaining a high specificity'.

Outcome of SurePath™ cervical samples reported as borderline nuclear change by cytological subtype and high-risk HPV status.

BACKGROUND: The average borderline rate in cervical cytology samples for English laboratories was 3.8% with the range being 2.0-6.8% at the time of the present study, which was undertaken in order to determine the association between different subtypes of borderline nuclear change (BNC), high-grade cervical intraepithelial neoplasia (CIN) and high-risk human papillomavirus (hrHPV) status. MATERIALS AND METHODS: Of 68,551 SurePath(TM) cervical samples reported in one laboratory over a period of 2 years, 2335 (3.4%) were reported as BNC. hrHPV status was known in 1112 cases (47.6%). The outcome was known only for women with hrHPV-positive BNC, who were recommended for colposcopy under the National Health Service Cervical Screening Programme sentinel site protocol. Women with hrHPV-negative BNC were returned to 3-yearly recall. The cases were subdivided into BNC, high-grade dyskaryosis cannot be excluded (B-HG; 105 cases); BNC with koilocytosis (B-K; 421 cases); BNC with other features of HPV (B-HPV; 160 cases); and BNC, not otherwise specified (B-NOS; 426 cases) and were correlated with the histological outcome where available. RESULTS: The study population age ranged from 23 to 65 years. Cases that tested positive for hrHPV by Qiagen HCII assay comprised 78.1%, 81.0%, 73.1% and 67.8% of B-HG, B-K, B-HPV and B-NOS categories, respectively. CIN2 or worse (CIN2+) was found in 64.6%, 10.0%, 19.7% and 20.1% of hrHPV-positive cases of B-HG, B-K, B-HPV and B-NOS, respectively, which was significantly higher in the B-HG category (P < 0.001) and lower in the B-K category compared with B-NOS (p < 0.001) and B-HPV (p = 0.006) respectively. CIN3+ comprised 55.6%, 6.3%, 26.3% and 19.1% of biopsies in the same categories, respectively. CONCLUSIONS: Subtyping BNC is useful, especially B-K and B-HG, which, respectively, had the lowest and highest rates of detection of both CIN2+ and CIN3+, confirming that koilocytosis is likely to be associated with transient HPV infection. Women with B-HG should be referred to colposcopy in the absence of HPV triage.

Survey of medical training in cytopathology carried out by the journal Cytopathology.

This report of the Editorial Advisory Board of Cytopathology gives the results of a survey of medical practitioners in cytopathology, which aimed to find out their views on the current situation in undergraduate and postgraduate training in their institutions and countries. The results show that training in cytopathology and histopathology are largely carried out at postgraduate level and tend to be organized nationally rather than locally. Histopathology was regarded as essential for training in cytopathology by 89.5% of respondents and was mandatory according to 83.1%. Mandatory cytopathology sections of histopathology were reported by 67.3% and specific examinations in cytopathology by 55.4%. The main deficiencies in training were due to its variability; there were insufficient numbers of pathologists interested in cytology and a consequent lack of training to a high level of competence. Pathologists without specific training in cytopathology signed out cytology reports according to 54.7% of responses, more often in centres where training was 3-6 months or less duration. Although 92.2% of respondents thought that specialist cytology should not be reported by pathologists without experience in general cytopathology, that practice was reported by 30.9%, more often in centres with small workloads. The survey report recommends that 6-12 months should be dedicated to cytopathology during histopathology training, with optional additional training for those wanting to carry out independent practice in cytopathology. Formal accreditation should be mandatory for independent practice in cytopathology. When necessary, temporary placements to centres of good practice should be available for trainees intending to practise independently in cytopathology. There should be adequate numbers of pathologists trained in cytopathology to a high level of competence; some of their time could be released by training cytotechnologists and trainee pathologists to prescreen cytology slides and assess adequacy of fine-needle aspiration samples when immediate diagnosis was not required. The survey demonstrated a clear need for European and international guidelines for training in cytopathology.

The revised BSCC terminology for abnormal cervical cytology.

The BSCC terminology was originally published in 1986 and although highly successful, requires revision. Through a process of professional consensus and literature review this has been undertaken by the BSCC. The revision takes account of recent developments and improvements in understanding of morphology and disease process and is compatible with other terminologies in use elsewhere, whilst still maintaining a focus on practice in the UK cervical screening programmes.

Proposed Sheffield quantitative criteria in cervical cytology to assist the diagnosis and grading of squamous intra-epithelial lesions, as some Bethesda system definitions require amendment.

OBJECTIVE: This study assesses the accuracy of published quantitative and qualitative criteria in the Bethesda System (TBS) for squamous intra-epithelial lesions. METHODS: Quantitative image analysis was undertaken on illustrations from TBS publications and also from slides in Cytology Training Centre teaching sets. Comparisons were also made with the British Society for Clinical Cytology (BSCC) terminology in cervical cytology, using the illustrations in their terminology publication and amalgamating the results into their proposed new two-tier model. RESULTS: TBS quantitatively defines low-grade squamous intra-epithelial lesions (LSIL) in both conventional and liquid-based cytology (LBC) preparations as showing nuclear enlargement more than x3 the area of a normal intermediate squamous cell nucleus. This study found that the increase in mean nuclear area was limited to only x2 in conventional preparations. In LBC (SurePath preparations, there was only a statistically non-significant x1.2 increase. This study identified a progressive and statistically significant reduction in mean cytoplasmic area from normal intermediate cells to LSIL and then to high-grade squamous intra-epithelial lesions (HSIL) in both conventional and LBC preparations. Furthermore, the most consistent quantitative finding in both conventional and LBC preparations was a statistically significant increase in the mean area and diameter ratios from normal intermediate cells to LSIL and then to HSIL. In all instances this varied from x2 to just below x3. This is in agreement with TBS, which states that the cytoplasmic area in HSIL is decreased leading to a marked increase in nuclear to cytoplasmic (NC) ratio. With the exception of an increase in mean nuclear area in conventional preparations from normal intermediate cells to LSIL, the predominant cause for this increase in NC ratios was a reduction in mean cytoplasmic area. The numerical increase in NC ratio for LSIL identified in this study was greater than implied by the 'slightly increased' statement in TBS. TBS comments that some HSIL cells can have the same degree of nuclear enlargement as in LSIL and that other HSIL cells may have much smaller nuclei than in LSIL. Both of these qualitative comments were supported in this study. The mean diameter NC ratios of 33% and 50% could provide useful diagnostic assistance in the distinction of normal intermediate cells and LSIL and between LSIL and HSIL, respectively. Because of overlapping individual ranges, however, additional diagnostic features such as nuclear morphology must be used in the distinction of normal intermediate cells, LSIL and HSIL. No statistical difference was identified in the mean diameter NC ratios between ASC-US and LSIL in TBS publications. In addition, the proposed new BSCC low and high grades of squamous abnormality were not statistically different from ASC-US/LSIL and HSIL, respectively. This provides support that the proposed BSCC two-tier system of squamous abnormalities is comparable to TBS. This study shows that LBC has variable but major and significant effects on nuclear and cytoplasmic morphology and that quantitative definitions in conventional preparations cannot be automatically extrapolated to LBC methodology. CONCLUSIONS: The study shows that some TBS quantitative and qualitative criteria require amendment and that an alternative quantitative approach, such as diameter NC ratio has a more valid scientific evidence base. Furthermore, use of NC ratios avoids the problems associated with the variable changes in nuclear and cytoplasmic areas, occurring between conventional and different commercial LBC preparations. By contrast, classifications based on area comparisons must be tailored to the specific conventional or commercial LBC preparation.

Proposed Sheffield quantitative criteria in cervical cytology to assist the grading of squamous cell dyskaryosis, as the British Society for Clinical Cytology definitions require amendment.

OBJECTIVE: In 1986, the British Society for Clinical Cytology (BSCC) published quantitative criteria to assist diagnosis in a three-tier grading system of squamous cell dyskaryosis. In dyskaryotic cells, area nuclear to cytoplasmic (NC) ratios below 50%, between 50% and 66% and over 66% were defined as equating with mild, moderate and severe grades respectively. Following the Terminology Conference in 2002, however, the BSCC recommended on their website that the three-tier model should be replaced by a new two-tier system of low- and high-grade squamous abnormalities. The latter broadly equate with the two-grade Bethesda System (TBS) for reporting squamous intraepithelial lesions. The purpose of this study was to assess the accuracy and reproducibility of the BSCC three-tier quantitative definitions, to investigate if they were applicable to liquid-based cytology (LBC) and to see how they related to the proposed new two-tier BSCC system. METHODS: Quantitative image analysis was undertaken on illustrations from the 1986 BSCC terminology publication and on microscope slides from external quality assessment and Cytology Training Centre teaching sets. RESULTS: Analysis of mean NC ratios showed that mild, moderate and severe dyskaryosis exist as statistically different populations. Overlap of NC ratio ranges, however, limits their practical application in the three-tier model, although interestingly no overlap was noted between mild and severe dyskaryosis. No grade of dyskaryosis had a mean area NC ratio over 50%, indicating that the BSCC quantitative definitions are incorrect. The mean diameter NC ratios for mild, moderate and severe dyskaryosis were found to be 40%, 49% and 66% respectively. Accordingly it is possible that those reporting cervical cytology could be interpreting the BSCC NC ratios as meaning diameter rather than area. Amalgamation of the three-tier results into the proposed two-tier model shows that the resulting mean NC area and diameter ratios identify statistically different low- and high-grade populations. The reduced degree of overlap, however, of NC ratio ranges in the two-tier model implies that NC ratios could have a useful practical role in the separation of the low- and high-grade categories. The two categories were reasonably well separated by mean area and diameter NC ratios of 25% and 50% respectively. A two-tier model combining mild with moderate rather than severe dyskaryosis was found to be a statistically valid alternative but gave rise to NC ratios that would be difficult to use in practice. Except for moderate dyskaryosis, no significant differences were identified between the mean NC ratios of either conventional and LBC preparations or LBC preparations using two different commercial methodologies (SurePath and ThinPrep). Differences, however, were noted in area measurements between SurePath and ThinPrep and this has potential implications for classifications (such as TBS) using area comparisons as their basis. In addition, it was found that the increased NC ratio, associated with higher grades of dyskaryosis is more a consequence of progressive cytoplasmic area reduction rather than nuclear area increase. The similar NC ratios of borderline nuclear changes associated with human papilloma virus and mild dyskaryosis support the BSCC proposal that these can be combined to constitute a low-grade category. This study shows that the BSCC area NC ratio criteria of grading squamous cell dyskaryosis require amendment. In addition, this study supports the new BSCC recommendation of low- and high-grade squamous cell categories. CONCLUSIONS: The study proposes Sheffield quantitative criteria to assist the grading of squamous cell abnormalities. Quantitative diameter NC ratio measurements, however, must always be accompanied by detailed assessment of qualitative morphological features and in particular those relating to nuclear chromatin. This is equally relevant to both two- and three-tier models.

Bethesda 2001.

The Bethesda system (TBS) for reporting cervical/vaginal cytological diagnoses was originally developed in 1988 at a National Cancer Institute (NCI) workshop convened to consider the benefits of increased standardization in the diagnostic reports provided by cytology laboratories. It rapidly gained acceptance in laboratory practice in the USA and beyond and three years later the NCI sponsored another workshop to assess the use of TBS in practice and consider areas for improvement. Subsequently an illustrated guide was published.

Is microscopic assessment of macroscopically normal hysterectomy specimens necessary?

AIM: To determine whether microscopic examination of macroscopically normal hysterectomy specimens yields findings that could alter subsequent clinical management. METHODS: All pathology reports on hysterectomy specimens submitted to the department of histopathology at the Northern General Hospital from January 1997 to December 1998 were reviewed. Cases were included for further assessment if the hysterectomy specimen was regarded as macroscopically normal by a consultant pathologist and if the patient had no history of, or suspicion of, neoplastic disease. The subsequent microscopic findings from these cases were assessed to determine whether any lesions of clinical importance were identified. RESULTS: Eight hundred and fifty four specimens were reviewed, of which 139 were suitable for inclusion. Only one of the 139 cases harboured a microscopic abnormality that necessitated specific clinical follow up; this was a focus of cervical intraepithelial neoplasia 2 (CIN 2). On follow up of that patient, no further neoplastic disease was identified. CONCLUSION: Microscopic assessment of macroscopically normal hysterectomy specimens does not contribute to patient management and is unnecessary in an era of manpower shortage and cost containment.

Anal colposcopy and the diagnosis of anal intraepithelial neoplasia in high-risk gynecologic patients.

The objective of this study was to define the colposcopic features of the normal anal canal and of anal human papillomavirus (HPV)-associated lesions, including anal intraepithelial neoplasia (AIN), and to correlate the colposcopic impression with the final histopathologic diagnosis. A controled colposcopic screening study of women considered at risk for HPV-associated anal epithelial abnormalities was carried out. All colposcopic assessments included a biopsy with matching histopathologic diagnosis. The study group consisted of 213 women who were considered at risk of anal HPV infection and AIN. A further group of 50 women, who had no previous history of ano-genital HPV infection or AIN and whose recent cervical smear was negative were recruited as controls. Informed consent was obtained from all patients, and the study was approved by the local ethical committee. In the control group of 50 women no AIN was detected. Normal histology was obtained in 45/50 (90%) biopsies where normality had been predicted on colposcopy. Histologic diagnosis in the at-risk group was normal in 143 (67%), subclinical papillomarvirus infection (SPI) in 24 (11%), and AIN of all grades (including three cases of early invasive squamous cancer in a field change of AIN III) in 46 (22%) patients. Nineteen of 24 (79%) cases of SPI were incorrectly predicted as normal on colposcopy, and another one (4%) as AIN I-II. Only four (17%) cases of SPI were correctly predicted at colposcopy. Of the 46 cases of histologically proven AIN, 26 (56%) were AIN I-II, and 20 (44%) were AIN III. Some 50% of AIN I-II were incorrectly predicted as SPI on colposcopy. Of the 20 AIN III lesions, 15 (75%) were correctly predicted by colposcopy. Three (20%) of these lesions contained foci of early invasion, of which in only one case (33.3%) was invasive disease suspected at colposcopy. Some 25% (5/20) of AIN III lesions were incorrectly diagnosed as AIN I-II at colposcopy. As is the experience with colposcopic assessment of the cervix, anal colposcopy predictions correlated well with the final histologic diagnosis, at the normal and high-grade AIN ends of the spectrum. The colposcopic predictive distinction between SPI and low-grade AIN (I-II) was less accurate. It was difficult to distinguish early invasive lesions within a field change of AIN III, from pure AIN III. In these studies there were three cases of early anal squamous carcinoma arising in AIN III lesions, two of which were unsuspected clinically.