Cervical Brush Research Articles

High detection rate of human papillomavirus in anal brushings from women attending a proctology clinic

To study human papillomavirus (HPV) anal infection in anal brushings from women attending a proctology clinic, and compare results with those obtained from paired cervical brushings. Women attending a university hospital proctology clinic for anal conditions or as part of a screening campaign, were enrolled consecutively, excluding those reporting previous HPV-related pathologies. HPV genotypes in anal and cervical brushings were determined by sequencing and, in most cases, type-specific viral loads were measured. HPV DNA was detected in 28.3% of anal brushings, with 47.4% of HPV genotypes being high risk. Cervical HPV detection was at almost the same rate but HPV status was discordant in about half those women with at least one positive specimen. Abnormal cytological findings were more common in anal than in cervical samples, in particular in the proctology outpatients. Viral load measurements excluded the existence of a multiple infection with genotypes detected in discordant anal- and cervical-paired samples and showed a significant correlation between anal and cervical paired concordant samples. The high rate of HPV detection in anal brushings that is not usually related to HPV positivity in cervical brushings could provide support for offering HPV DNA tests to women attending proctology clinics.

Prevalence and genotyping of HPV, by cervical brushing, in Irpinia area of Campania region

Cervical cancer is due to persistent genital infection with Human Papillomavirus (HPV).The purpose of this study was to evaluate prevalence of HPV in Irpinia (Campania region, Italy), distribution of different viral genotypes, correlating cytological results and virological investigations. In the period 2006-2011, were made 1080 cervical samples of women aged 18-65 years for HPV identification and genotyping. Detection of the virus was performed by Multiplex-PCR System (Seegene,Arrow) and typing with INNO-LiPA HPV Genotyping Extra test (Innogenetics). Out of the 1080 tested samples, 330 (30.6%) samples were positive for HPV DNA. The most frequently occurring High Risk (HR)-HPV genotype in single infections was HPV16 (16.6%), followed by HPV51 (10.7%), in multiple infections HPV16 (15.7%) and 31 (14.6%). The prevalence of infection, correlated with age of patients studied, is greater in the group aged 26-30 years (42.5%). HR-HPV were detected in different percent in patients with Pap test scores: 22.5% in normal Pap smear (20% HPV16), 14.5% ASCUS (47.6% HPV16), 24% LSIL (20% HPV16), 79.3% HSIL (72.7% HPV16; 9.1% HPV18 detected only in this type of cellular alteration). The high prevalence of HR-HPV in patients with ASCUS or normal Pap test, suggesting the real advantage of HPV screening test, more sensitive in selecting the actual population at risk. Based on the findings of our epidemiological study, HR-HPV screening and HPV genotyping test should be strongly advised also to the vaccinated population for the high incidence of genotypes which are not included in vaccines (67%).

Management of missing strings in users of intrauterine contraceptives

BackgroundA common question among health care professionals is how to manage nonvisible strings in users of intrauterine contraceptives (IUCs) at repeat follow-up visits. This study assessed the position of the IUCs in women who consulted repeatedly with nonvisible IUC strings. Study DesignThe medical records of the clinic were reviewed to identify new acceptors and switchers who had an IUC inserted between 1990 and 2009. All women were identified whose IUC string could not be visualized at the external os of the cervix by the health care professional at any given follow-up visit, even after attempting a standard maneuver of sweeping the strings from the cervical canal using a cervical brush or trying to visualize the strings in the cervical canal using colposcopy. Data were also retrieved on the use of ultrasonography and/or pelvic X-ray to assess IUC position, as well as data from any subsequent visits at which the IUC strings were nonvisible. ResultsThe medical charts of 14,935 patients using an IUC were reviewed, and 750 women (5.0%) presenting for the first time with missing IUC strings at any follow-up visit were identified. Ultrasound scans showed the IUC to be in situ in 735 cases (98.0%), while 9 women (1.2%) had expelled the device and, in 5 cases (0.7%), the device was found in the pelvis following uterine perforation. IUC strings were missing on a second occasion in 297 cases. The device was found to be in situ in 295 cases (99.3%) and had been expelled in 2 (0.7%). At subsequent consultations, (between 1 and 18 years after the first consultation), strings were missing in 113, 55, 19 and 5 cases. In 111 (98.2%), 54 (98.2%), 18 (94.7%) and 5 (100%) of these cases, respectively, the IUC was found to be in situ, while in the remaining cases, the device had been expelled. ConclusionsMissing IUC strings are an uncommon finding, and ultrasonography confirmed that the device was in situ in the majority of these cases. For women with persistent missing IUC strings after one ultrasound scan that has verified appropriate intrauterine position, given the 2.4% likelihood that expulsion may have occurred at the time of subsequent visits, repeating the ultrasound (if available) should be considered for at least one (and possibly two) additional women's visit.

Chlamydia trachomatis as a probable cofactor in human papillomavirus infection in aboriginal women from northeastern Argentina

Objectives: High-risk types of human papillomavirus (HPV) are strongly associated with cervical cancer (CC), and Chlamydia trachomatis (CT), the most frequent sexually transmitted bacterial infection (STBI) worldwide, seems to be a risk factor for HPV infection and for CC. It is also known that both agents are more prevalent in vulnerable communities where lack of adequate primary health care is a cause for concern. The aim of this work was to determine the impact of CT and HPV infections in women belonging to an isolated aboriginal population (Pilaga community) from a poor region in Northern Argentina (province of Formosa). For this purpose, a cross-sectional study was performed in all sexually active Pilaga women, who attended a local community-based gynecological health screening project. The polymerase chain reaction (PCR) method on a cervical brush specimen was used to detect both agents. Results: A total of 227 women (20% of the total female population of the Pilaga community) were studied and the overall prevalence was 26.4% for CT, 46.7% for HPV and 16.3% for concurrent infection. CT infection was higher in HPV DNA positive (34.2%) than in HPV DNA negative women (19%; OR: 2.22/95% CI = 1.16-4.28 / p = 0.009) and the most prevalent HPV types were HPV-16 (19.4%), 6 and 18 (5.3%), 58 (3.5%) and 33 (3.1%). Conclusions: The prevalence of CT and HPV observed in Pilaga women are among the worst registered in Latin America. Also, data collected suggest that chlamydial infection may play an important role in the natural history of HPV infection. On this respect, we propose that the association between these two agents seems to be more related to a mutual potentiation than to the fact that they share a common route of transmission.

Chlamydia trachomatis as a probable cofactor in human papillomavirus infection in aboriginal women from northeastern Argentina

ObjectivesHigh-risk types of human papillomavirus (HPV) are strongly associated with cervical cancer (CC), and Chlamydia trachomatis (CT), the most frequent sexually transmitted bacterial infection (STBI) worldwide, seems to be a risk factor for HPV infection and for CC. It is also known that both agents are more prevalent in vulnerable communities where lack of adequate primary health care is a cause for concern. The aim of this work was to determine the impact of CT and HPV infections in women belonging to an isolated aboriginal population (Pilaga community) from a poor region in Northern Argentina (province of Formosa). For this purpose, a cross-sectional study was performed in all sexually active Pilaga women, who attended a local community-based gynecological health screening project. The polymerase chain reaction (PCR) method on a cervical brush specimen was used to detect both agents. ResultsA total of 227 women (20% of the total female population of the Pilaga community) were studied and the overall prevalence was 26.4% for CT, 46.7% for HPV and 16.3% for concurrent infection. CT infection was higher in HPV DNA positive (34.2%) than in HPV DNA negative women (19%; OR: 2.22/95% CI=1.16-4.28/p=0.009) and the most prevalent HPV types were HPV-16 (19.4%), 6 and 18 (5.3%), 58 (3.5%) and 33 (3.1%). ConclusionsThe prevalence of CT and HPV observed in Pilaga women are among the worst registered in Latin America. Also, data collected suggest that chlamydial infection may play an important role in the natural history of HPV infection. On this respect, we propose that the association between these two agents seems to be more related to a mutual potentiation than to the fact that they share a common route of transmission.

Non-Invasive Cytology Brush PCR Diagnostic Testing in Mucosal Leishmaniasis: Superior Performance to Conventional Biopsy with Histopathology

BackgroundTraditional methods of diagnosing mucosal leishmaniasis (ML), such as biopsy with histopathology, are insensitive and require collection of an invasive diagnostic specimen.MethodsWe compared standard invasive procedures including biopsy histopathology, biopsy PCR, and leishmanin skin test (LST) to a novel, non-invasive, cytology-brush based PCR for the diagnosis of ML in Lima, Peru. Consensus reference standard was 2/4 tests positive, and outcome measures were sensitivity and specificity. Leishmania species identification was performed by PCR-based assays of positive specimens.ResultsTwenty-eight patients were enrolled, 23 of whom fulfilled criteria for a diagnosis of ML. Sensitivity and specificity of biopsy with histopathology were 21.7% [95% CI 4.9–38.5%] and 100%; 69.6% [95% CI 50.8–88.4%] and 100% for LST; 95.7% [95% CI 87.4–100%] and 100% for biopsy PCR; and 95.7% [95% CI 87.4–100%] and 90% [95% CI 71.4–100%] for cytology brush PCR using both Cervisoft® and Histobrush® cervical cytology brushes. Represented species identified by PCR-RFLP included: L. (V). braziliensis (n = 4), and L. (V). peruviana (n = 3).ConclusionsUse of commercial grade cytology brush PCR for diagnosis of ML is sensitive, rapid, well tolerated, and carries none of the risks of invasive diagnostic procedures such as biopsy. Further optimization is required for adequate species identification. Further evaluation of this method in field and other settings is warranted.

PCR‐based identification of eight lactobacillus species and 18 hr‐HPV genotypes in fixed cervical samples of south african women at risk of HIV and BV

Vaginal lactobacilli assessed by PCR-based microarray and PCR-based genotyping of HPV in South African women at risk for HIV and BV. Vaginal lactobacilli can be defined by microarray techniques in fixed cervical samples of South African women. Cervical brush samples suspended in the coagulant fixative BoonFix of one hundred women attending a health centre for HIV testing in South Africa were available for this study. In the Ndlovu Medical Centre in Elandsdoorn, South Africa, identification of 18 hr-HPV genotypes was done using the INNO-LiPA method. An inventory of lactobacilli organisms was performed using microarray technology. On the basis of the Lactobacillus and Lactobacillus biofilm scoring, the cases were identified as Leiden bacterial vaginosis (BV) negative (BV-; n = 41), Leiden BV intermediate (BV±; n = 25), and Leiden BV positive (BV+; n = 34). Fifty-one women were HIV positive and 49 HIV negative. Out of the 51 HIV positive women, 35 were HPV infected. These 51 HIV positive women were frequently infected with HPV16 and HPV18. In addition, HPV35, HPV52, HPV33, and HPV66 were often detected in these samples. Lactobacillus salivarius and Lactobacillus iners were the most prevalent lactobacilli as established by the microarray technique. In women with HPV infection, the prevalence of Lactobacillus crispatus was significantly reduced. In both HIV and HPV infection, a similar (but not identical) shift in the composition of the lactobacillus flora was observed. We conclude that there is a shift in the composition of vaginal lactobacilli in HIV-infected women. Because of the prominence of HPV35, HPV52, HPV33, and HPV66, vaccination for exclusively HPV16 and HPV18 might be insufficient in South African HIV+ women.

Application of the Hybrid Capture 2 assay to squamous cell carcinomas of the head and neck

A growing proportion of head and neck squamous cell carcinoma (HNSCC) is caused by the human papillomavirus (HPV). In light of the unique natural history and prognosis of HPV-related HNSCCs, routine HPV testing is being incorporated into diagnostic protocols. Accordingly, there is an escalating demand for an optimal detection strategy that is sensitive and specific, transferrable to the diagnostic laboratory, standardized across laboratories, cost-effective, and amenable to broad application across specimen types including cytologic preparations. Cytologic preparations (fine-needle aspirates [FNAs] and brushes) were obtained from surgically resected HNSCCs and evaluated for the presence of high-risk HPV using the Hybrid Capture 2 assay. HPV analysis was also performed on the corresponding tissue sections using HPV in situ hybridization and p16 immunohistochemistry. In cases in which the immunohistochemical and in situ hybridization results were discordant, HPV status was determined by real-time polymerase chain reaction detection of E7 expression. HPV status in the tissues and corresponding cytologic samples was compared. Based on benchmark HPV testing of the tissue sections, 14 HNSCCs were classified as HPV positive and 10 as HPV negative. All corresponding cytologic preparations were correctly classified using the Hybrid Capture 2 assay. The Hybrid Capture 2 strategy, already widely used for the detection of high-risk HPV in cervical brushes, is readily transferrable to HNSCCs. Consistent accuracy in cytologic preparation suggests its potential application in FNAs from patients who present with lymph node metastases, and may eliminate the need to obtain tissue solely for the purpose of HPV testing.

Descriptive epidemiology of Pap test results from women with gynecologic symptoms in Georgia

Descriptive epidemiology of Pap test results from women with gynecologic symptoms in Georgia

Optimising human papillomavirus self-testing for high risk women

ObjectivesHuman papillomavirus self-tests that can be used at home and returned by mail may increase cervical cancer screening rates. Growing evidence suggests that self-test methods could increase screening for high-risk...

Testing for human papillomavirus and measurement of viral load of HPV 16 and 18 in self‐collected vaginal swabs of women who do not undergo cervical cytological screening in Southern France

Self-sampling using vaginal swabs could be a valuable alternative to screen for cervical cancer for women who do not attend regular cytological screening. The aim of this study was to determine the prevalence of high and low-risk HPV types and of HPV type 16 and 18 DNA load in self-collected vaginal swabs from 35- to 69-year-old Southern French women of low socioeconomic level or migrant populations who do not attend regular cervical screening. A good concordance (93.1%) was found between cervical brush and vaginal swabs in 29 samples. Self-collected vaginal swabs were examined from 120 women. HPV infection was found in 28 women (23.3%; median age 48 years), 17 (14.1%) of whom harbored high-risk HPV types. HPV type 16 was the high risk type found most frequently, followed by types 53, 31, 18, 58, and 66. The low-risk type detected most frequently was HPV type 6, followed by types 61, 70, and 81. The mean HPV 16 and 18 load was 6.3 log(10) copies/10(6) cells and 2.4 log(10) copies/10(6) cells, respectively. These results suggest that vaginal self-swabs can be a reliable tool for cervical cancer screening in non-attending and inadequately screened elderly women.

Prenatal diagnosis of trisomy 21 through detection of trophoblasts in cervical smears

Background Fetal cells exfoliate in the uterine cavity during early pregnancy and are a potential source of material for NIPD. Aims This study was designed to test the hypothesis that fetal cells obtained from the uterine cervix during the first trimester of pregnancy could be utilized for prenatal diagnosis of chromosomal aneuploidy. Study design Fetal cells retrieved from the distal endocervical canal during the first trimester of pregnancy were hybridized with chromosome 21 specific FISH probes and analyzed with an automated fluorescence microscope. Subjects and outcome measures Cells with 3 copies of chromosome 21 were detected in 5 out of 5 trisomy 21 pregnancies. Results The number of trisomic cells detected ranged from 1 to 27 with a median value of 5. Conclusions FISH-based scanning can identify trisomy 21 pregnancies by analysis of routine cervical brushings. The approach offers the potential for non-invasive prenatal diagnosis as early as 5 weeks gestation.

Exploiting the Residual of Cervical Thin Layer Brush Samples Through Cytohistology in Cases with Invasive Carcinoma with Application of Antibodies

To exploit cervical thin layer brush samples through cytohistology in cases with invasive carcinoma with application of antibodies.Fourteen cases from women with carcinoma diagnosed in 2006 were selected out of 29 invasive carcinomas. From these 14 cases liquid-based cervical cytology material was available to prepare cytohistology. Eight women had squamous cell carcinoma, 4 endocervical adenocarcinoma, 1 endometrial adenocarcinoma and 1 ovarian adenocarcinoma. The residual material from the thin layer sample, collected by brushes by general practitioners, was used to prepare paraffin sections. These were stained with the Papanicolaou method and for the biomarkers Ki-67 and p16 and, if desired, for differentiation markers, including carcinoembryonic antigen, vimentin, cytokeratin 7 and cytokeratin 20 to establish the immunoprofile of the carcinoma.The morphologic details in the cancer nuclei in the paraffin sections were excellent, while in all cases the thin layer cytology slide contained thick epithelial fragments with blurred nuclei. In 5 of the 6 adenocarcinomas, the glandular architecture diagnostic of adenocarcinoma was visible in the cytohistology, which was highlighted in the biomarker stainings, particularly so in the Ki-67 sections. With the exception of endometrial adenocarcinoma, all p16(INK4a) stainings were positive, as they were in the ovarian adenocarcinoma case.Cytohistology is an adjunct to routine cervical cytologic examination of thin layer samples, allowing an unequivocal and refined diagnosis.

Sensitive HPV detection in oropharyngeal cancers

BackgroundHuman papillomaviruses (HPV) are the aetiological agents of certain benign and malignant tumours of skin and mucosae; the most important of which is cervical cancer. Also, the incidence of ano-genital warts, HPV-anal cancer and oropharyngeal cancers are rising. To help ascertain a useful PCR detection protocol for oropharyngeal cancers, we directly compared three commonly used primer sets in detection of HPV from different clinical samples.MethodsWe compared PGMY09/11, MY09/11 and GP5+/6+ primers sets in PCRs of 34 clinically diagnosed samples of genital warts, cervical brushings (with associated histological diagnosis) and vulval biopsies. All negative samples were subsequently tested using the previously reported PGMY/GP PCR method and amplicons directly sequenced for confirmation and typing. An optimised PCR protocol was then compared to a line blot assay for detection of HPV in 15 oropharyngeal cancer samples.ResultsPGMY09/11 primers detected HPV presence in more cervical brushing (100%) and genital wart (92.9%) samples compared to MY09/11 (90% and 64.3%) and GP5+/6+ (80% and 64.3%) primer sets, respectively. From vulval biopsies, HPV detection rates were: MY09/11 (63.6%), GP5+/6+ (54.5%) and PGMY09/11 (54.5%). PGMY/GP nested PCR demonstrated that HPV was present, and direct sequencing confirmed genotypes. This nested PCR protocol showed detection of HPV in 10/15 (66.7%) of oropharyngeal cancer samples.ConclusionsPGMY09/11 primers are the preferred primer set among these three for primary PCR screening with different clinical samples. MY09/11 and GP5+/6+ may be used (particularly for cervical samples) but demonstrate lower detection rates. A nested PCR approach (i.e. a PGMY-GP system) may be required to confirm negativity or to detect low levels of HPV, undetectable using current primary PCR methods, as demonstrated using oropharyngeal cancer samples.

Clinical Performance of Human Papillomavirus E6 and E7 mRNA Testing for High-Grade Lesions of the Cervix

Infection with high-risk (HR) human papillomavirus (HPV) is the major cause of cervical cancer. However, relatively few infections progress to malignant disease. Progression to malignancy requires the overexpression of the E6 and E7 genes in the integrated HPV genome. It follows that the E6 and E7 transcripts could be useful markers of disease progression. The study presented here tests this possibility, using data from colposcopy and from cytological and histological tests to compare RNA assays for the E6 and E7 genes with DNA testing. A total of 180 women underwent colposcopy, cytology, and biopsy of suspected lesions (143 cases). Cervical brush specimens were analyzed for HPV DNA and for E6 and E7 mRNA. DNA from HR HPV was found in 57.8% of the specimens; E6 and E7 transcripts were found in 45%. The rates of detection of HPV DNA and of E6 and E7 transcripts were 33.3% and 25%, respectively, for specimens with normal findings; 51.4% and 31.9%, respectively, for specimens with cervical intraepithelial neoplasia grade 1 (CIN1); and 61.1% and 44.2% for specimens with CIN2, respectively. All specimens with CIN3 and 95.5% of specimens from patients with squamous cell carcinoma were positive by both assays. Thirty-seven patients with normal colposcopy findings did not undergo biopsy. HPV DNA and mRNA transcripts were found in 32.4% and 18.9% of these cases, respectively. Comparisons with cytological tests produced similar results. Overall, the mRNA tests showed a higher specificity than the DNA tests for high-grade lesions (72.7% and 56.2%, respectively) and a higher positive predictive value (59.3% and 49.0%, respectively). These findings suggest that mRNA assays could be more powerful than DNA testing for predicting the risk of progression and offer a strong potential as a tool for triage and patient follow-up.

Biomarker p16INK4a on Paraffin Sections Prepared from Cervical Brush Samples Highlights Nuclear Changes Resulting in Unquestionable Cytohistodiagnosis

To optimize the morphology of pl6-positive atypical and (pre)neoplastic cells in paraffin cytoblock sections with the aim to minimize equivocal diagnoses on cytology samples.Patients with negative cytology results or results within normal limits (WNL) (n=38), atypical squamous cells of undetermined significance (ASCUS) (n=25) and high grade squamous intraepithelial lesion (HSIL) (n=16) were selected. The residual material of the cervical brush samples was processed to cytoblock paraffin sections and stained for pl1. The cytohistodiagnosis of the pl1-stained paraffin sections was based on the cytologic and histologic morphology.Of the 38 cytologically negative cases, only 4 contained afe w faintly positive pl61cells. Of the 25AS CUS cases, I 1as cytohistologically upgraded to low grade squamous intraepithelial lesion (LSIL). All 16 HSIL cases contained cells with outspoken diffuse positive immunostaining, highlighting chromatin clumping in the (pre)malignant cells. In the paraffin sections the tissue fragments showed architecture consistent with that of HSIL. The nuclear to cytoplasmic ratio in the HSIL was severely disturbed.Cervical brush samples allow an unequivocal cytohistodiagnosis based on the (pre)malignant nuclear changes highlighted by the p16 staining of the paraffin sections.

Human Papillomaviruses and genital co-infections in gynaecological outpatients

BackgroundHigh grade HPV infections and persistence are the strongest risk factors for cervical cancer. Nevertheless other genital microorganisms may be involved in the progression of HPV associated lesions.MethodsCervical samples were collected to search for human Papillomavirus (HPV), bacteria and yeast infections in gynaecologic outpatients. HPV typing was carried out by PCR and sequencing on cervical brush specimens. Chlamydia trachomatis was identified by strand displacement amplification (SDA) and the other microorganisms were detected by conventional methods.ResultsIn this cross-sectional study on 857 enrolled outpatients, statistical analyses revealed a significant association of HPV with C. trachomatis and Ureaplasma urealyticum (at high density) detection, whereas no correlation was found between HPV infection and bacterial vaginosis, Streptococcus agalactiae, yeasts, Trichomonas vaginalis and U. urealyticum. Mycoplasma hominis was isolated only in a few cases both in HPV positive and negative women and no patient was infected with Neisseria gonorrhoeae.ConclusionAlthough bacterial vaginosis was not significantly associated with HPV, it was more common among the HPV positive women. A significant association between HPV and C. trachomatis was found and interestingly also with U. urealyticum but only at a high colonization rate. These data suggest that it may be important to screen for the simultaneous presence of different microorganisms which may have synergistic pathological effects.

Clinical follow-up of women infected with human papillomavirus-16, either alone or with other human papillomavirus types: identification of different risk groups

Evaluation of the clinical impact of multiple infections of the cervix by human papillomavirus, including human papillomavirus-16, compared with single human papillomavirus-16 infection. One hundred sixty-nine women were classified in 3 categories depending on their human papillomavirus profile: human papillomavirus-16 only, human papillomavirus-16 and low-risk type(s), and human papillomavirus-16 and other high-risk type(s). Cervical brush samples were analyzed for human papillomavirus DNA by polymerase chain reaction and reverse line blot hybridization. All women were evaluated with colposcopy during 24 months or more. Management was according to the Bethesda recommendations. Women infected with human papillomavirus-16 and other high-risk human papillomavirus type(s) presented more progression or no change in the grade of dysplasia, compared with women of the other groups (relative risk [RR], 1.39; 95% confidence interval [CI], 1.07-1.82; P = .02 at 6 months; RR, 2.10; 95% CI, 1.46-3.02; P < .001 at 12 months; RR, 1.82; 95% CI, 1.21-2.72; P = .004 at 24 months). Coinfection of women with human papillomavirus-16 and other high-risk human papillomavirus type(s) increases the risk of unfavorable evolution.

Genotoxic effects of X-rays on keratinized mucosa cells during panoramic dental radiography

The aim of this study was to evaluate the genotoxic effects of X-rays on epithelial gingival cells during panoramic dental radiography using a differentiated protocol for the micronucleus test. 40 healthy individuals who underwent this procedure for diagnostic purposes on request from their dentists agreed to participate in this study. All of them answered a questionnaire before the examination. Epithelial gingival cells were obtained from the keratinized mucosa of the upper dental arcade by gentle scraping with a cervical brush immediately before exposure and 10 days later. Cytological preparations were stained according to the Feulgen-Rossenbeck reaction, counterstained with fast green 1% for 1 min and analysed under a light microscope. Micronuclei, nuclear projections (broken eggs) and degenerative nuclear alterations (pyknosis, karyolysis, karyorrhexis and condensed chromatin) were scored. The frequency of micronuclei was significantly higher after exposure (P < 0.05), as were the frequencies of nuclear alterations indicative of apoptosis (P < 0.001). These results indicate that X-ray radiation emitted during panoramic dental radiography induces a genotoxic effect on epithelial gingival cells that increases the frequency of chromosomal damage and nuclear alterations indicative of apoptosis.

Y‐STR Profiling in Extended Interval (≥3 days) Postcoital Cervicovaginal Samples*

Depending upon specific situations, some victims of sexual assault provide vaginal samples more than 36-48 h after the incident. We have tested the ability of commercial and in-house Y-STR systems to provide DNA profiles from extended interval (> or =3 days) postcoital samples. The commercial Y-STR systems tested included the AmpFlSTR Yfiler (Applied Biosystems), PowerPlex Y (Promega) and Y-PLEX 12 (Reliagene) products whereas the in-house systems comprised Multiplex I (MPI) and Multiplex B (MPB). Three donor couples were recruited for the study. Postcoital cervicovaginal swabs (x2) were recovered by each of the three females at specified intervals after sexual intercourse (3-7 days). Each time point sample was collected after a separate act of sexual intercourse and was preceded by a 7-day abstention period. As a negative control, a precoital swab was also recovered prior to coitus for each sampling and only data from postcoital samples that demonstrated a lack of male DNA in the associated precoital sample was used. A number of DNA profile enhancement strategies were employed including sampling by cervical brushing, nondifferential DNA extraction methodology, and post-PCR purification. Full Y-STR profiles from cervicovaginal samples recovered 3-4 days after intercourse were routinely obtained. Profiles were also obtainable 5-6 days postcoitus although by this stage partial profiles rather than full profiles were a more likely outcome. The DNA profiles from the sperm fraction of a differential lysis were superior to that obtained when a nondifferential method was employed in that the allelic signal intensities were generally higher and more balanced and exhibited less baseline noise. The incorporation of a simple post-PCR purification process significantly increased the ability to obtain Y-STR profiles, particularly from 5- to 6-day postcoital samples. Remarkably an 8 locus Y-STR profile was obtained from a 7-day postcoital sample, which is approaching the reported time limit for sperm detection in the cervix.