Last data update: Jun 17, 2024. (Total: 47034 publications since 2009)
Records 1-7 (of 7 Records) |
Query Trace: Swan DC [original query] |
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Patterns of cellular and HPV 16 methylation as biomarkers for cervical neoplasia
Patel DA , Rozek LS , Colacino JA , Van Zomeren-Dohm A , Ruffin MT , Unger ER , Dolinoy DC , Swan DC , Onyekwuluje J , Degraffinreid CR , Paskett ED . J Virol Methods 2012 184 84-92 ![]() Aberrant promoter methylation of biologically relevant genes in cervical cancer and uneven CpG distribution within the human papillomavirus 16 (HPV 16) enhancer region have been reported. Cervical samples and questionnaires from 151 women screened for cervical cancer in Appalachian Ohio were analyzed. Methylation was measured by bisulfite sequencing in candidate gene sites in ESR1, DCC, p16, and LINE1 elements. Among 89 HPV 16-positive women, CpG sites in the E6 promoter and enhancer regions and the L1 region of the HPV 16 genome were measured. Methylation levels were compared by cervical cytology and HPV 16 status. HPV methylation was low regardless of cytology status, however E6 methylation was significantly higher in women with normal cytology. ESR1 and DCC methylation were significantly higher in HPV 16-positive women. Increased methylation at sites in the E6 promoter region was associated with lower odds of abnormal cytology. Increased methylation in candidate genes was associated with higher odds of abnormal cytology, particularly DCC region 2.4, DCC region 2.6, ESR1 region 3.2, and LINE1 site 1.2. HPV 16 genome CpG methylation was low except for the L1 region. In general, lower HPV 16 methylation and higher candidate gene methylation levels were associated with higher odds of abnormal cytology. |
A real-time PCR assay for HPV52 detection and viral load quantification.
Onyekwuluje JM , Steinau M , Swan DC , Unger ER . Clin Lab 2012 58 61-6 ![]() BACKGROUND: Human papillomavirus (HPV) 52 is one of the most frequent high risk HPV types found in cervical and anal infections. Reliable and well characterized methods for HPV 52 detection are therefore of great importance. Unfortunately one of the most widely used commercially available HPV typing assays, the Roche Linear Array (LA), detects type 52 only through its XR probe which cross-reacts with types 33, 35 and 58 and fails to give unambiguous detection of HPV 52. METHODS: To address this problem a real-time TaqMan PCR assay for HPV 52 targeting the E6/E7 region was developed and validated, which can be applied as robust duplex assay simultaneously detecting beta-globin as genomic control and reference or as highly sensitive single target detection assay. RESULTS: Optimized primer and probe concentrations produced linear PCR amplifications over seven logs of targets (10(1) - 10(7)). The detection limit for HPV 52 was reproducibly at 10 copies per reaction for the duplex assay format and 5 copies for the single-plex format. The assay was very type-specific and no amplification signal was observed with 10(7) copies of the related HPV33, 35, and 58 DNA. Of 89 samples that tested unambiguously positive for HPV 52 in the LA, 75 were confirmed in the duplex format and 88 in the single-plex format. An additional 100 samples negative for HPV 52 in LA were all negative in both HPV 52 real-time PCR assay formats. CONCLUSIONS: These results indicate 92.6% and 99.5% accuracy relative to LA for the duplex and single-plex formats, respectively. In ongoing testing of 18,484 from various studies, 10.8% required the HPV52 TaqMan assay to unequivocally determine the status. Including the HPV 52 duplex assay provides the ability to monitor variations in the cell yield in various methods of sample collection and processing. This additional information is useful in QC monitoring of epidemiologic studies. |
Anogenital human papillomavirus in sexually abused and nonabused children: a multicenter study
Unger ER , Fajman NN , Maloney EM , Onyekwuluje J , Swan DC , Howard L , Beck-Sague CM , Sawyer MK , Girardet RG , Sautter RL , Hammerschlag MR , Black CM . Pediatrics 2011 128 (3) e658-65 OBJECTIVES: To characterize the epidemiology of genital human papillomavirus (HPV) infection in children without previous consensual sexual activity, comparing HPV prevalence by certainty of child sexual abuse (CSA). PATIENTS AND METHODS: Patients presenting for evaluation of CSA in 8 sites in Atlanta, Houston, Harrisburg, and New York City were recruited along with patients presenting for unrelated health visits. CSA certainty was classified as definite, probable, possible, or no evidence following published guidelines and the results of history, physical examination, and laboratory tests. Urine and swabs of external genitalia were tested for HPV using L1 consensus polymerase chain reaction. RESULTS: The study included 576 participants (89.9% female) aged 6 months to 13 years (mean: 7.9); 534 of whom were evaluated for CSA and 42 for unrelated reasons. Of those evaluated for CSA, 14 had genital warts. One or more HPV types were detected in 11.8% (61 of 517) of participants with adequate samples. HPV detection was more likely among abused participants (definite, probable, or possible) than among participants without evidence of CSA (13.7% and 1.3%, respectively; P < .0001) and increased with certainty of abuse (8.4%, 15.6%, and 14.5% in participants with possible, probable, and definite CSA, respectively; P < .0001). Participants aged 10 years or older had a higher prevalence of HPV (20.6%) than others (5.6%) (P < .0001). CSA, anogenital warts, and age were independently associated with HPV detection. CONCLUSIONS: HPV detection was associated with CSA and increased with CSA certainty. In this population, genital HPV seemed to behave as a sexually transmitted infection. |
Impact of HPV assay on observed population prevalence.
Unger ER , Steinau MS , Lin JM , Patel SS , Swan DC . Diagn Mol Pathol 2011 20 (2) 101-4 ![]() Type-specific surveillance of human papillomavirus (HPV) has been proposed as an early indicator of vaccine impact. Longitudinal comparison of HPV typing results requires stable assays with high type-specific reproducibility. Assays are evolving and the impact of even minor changes in the assay format may be difficult to anticipate. We initiated a population-based study of HPV with the prototype line blot (PLB) assay. These reagents were replaced by the research use only Linear Array (LA) HPV Genotyping kit. The assays are similar in principle and earlier comparisons found increased sensitivity and detection of more types per sample with LA; however, in samples from women with cervical abnormalities, the overall concordance was good. Slight changes in sensitivity may be more significant in samples from a general population with lower viral loads in the samples. Residual extracts from 3001 self-collected vaginal swabs from women in the general US population originally tested with PLB were retested with LA. With LA, all the samples were hybridized. PLB hybridization was restricted to samples with probable amplicon in gel electrophoresis. For HPV detection, the agreement between the 2 assays was 78.6% (kappa=0.55) with a positive concordance of 52.8%. However, this masks the observation that repeat testing with LA led to the detection of HPV in nearly twice as many samples. Agreement improves if comparison was restricted to the samples hybridized. These results emphasize that assay comparisons should consider the clinical-epidemiologic context of sample collection. Studies designed to examine temporal trends in type-specific prevalence should archive residual material to permit retesting if assays change. |
Human papillomavirus infection and cytologic abnormalities of the anus and cervix among HIV-infected women in the Study to Understand the Natural History of HIV/AIDS in the Era of Effective Therapy (The SUN Study)
Kojic EM , Cu-Uvin S , Conley L , Bush T , Onyekwuluje J , Swan DC , Unger ER , Henry K , Hammer JH , Overton ET , Darragh TM , Palefsky JM , Vellozzi C , Patel P , Brooks JT . Sex Transm Dis 2010 38 (4) 253-9 BACKGROUND: Human papillomavirus (HPV) infection of the cervix and related abnormal cervical cytology in HIV-infected women has been well described. Little is known about anal HPV infection in HIV-infected women. METHODS: The SUN Study is a prospective cohort study of 700 HIV-infected patients including 167 women. At baseline, patients completed a behavioral questionnaire and provided, among other samples, cervical and anal swabs for HPV detection and genotyping and for cytologic examination. Here, we present the available baseline data on the 167 women in the SUN study. RESULTS: Baseline results were available for 120 women (median age: 38 years, 57% non-Hispanic black, median CD4 cell count 444.5 cells/mm), of whom, 77% were taking antiretroviral therapy. The prevalences in the anus and cervix of any HPV were 90% and 83%, respectively (P = 0.039), and of high-risk (HR) types 85% and 70%, respectively, (P = 0.001). There was no significant difference in the prevalences of abnormal cytology between the anus and cervix: 38% and 33%, respectively (P = 0.217). Although the presence of abnormal cervical cytology was associated with the presence of abnormal anal cytology (relative risk: 1.7, P = 0.024), its sensitivity (52.5%) and positive predictive value positive (45.6%) for identifying women with abnormal anal cytology were poor. A history of anal sex was not associated with anal HPV infection or abnormal anal cytology. CONCLUSIONS: In this cohort of HIV-infected women, anal HPV infection was more prevalent and diverse than cervical HPV infection. Anal cytologic abnormalities were as prevalent as cervical cytologic abnormalities, and although abnormal cervical cytology was predictive of abnormal anal cytology, results were not highly concordant. These data support the need for studies of anal cytologic screening of HIV-infected women. |
Differences and changes of HPV16 variant status in HIV-positive adults are not uncommon
Steinau M , Swan DC , Onyekwuluje JM , Brooks JT , Vellozzi C , Unger ER , Sun Study Investigators . J Gen Virol 2010 91 2068-2072 HPV16 genotype variants have been the subject of several investigation, but study participants were rarely sampled more than ones. Among a cohort of Human immunodeficiency virus (HIV)-infected adults, we investigated HPV16 variants in samples collected concurrently from anus and cervix, as well as in serial samples collected from the same anatomical site at twelve-month intervals. We determined HPV16 variants in stored extracts of cervical and anal samples from subjects with multiple visits and at least one sample positive for HPV16. Seven polymorphic nucleotide positions within the E6 region were analyzed by pyrosequencing to determine genotype variants. Of 364 samples examined, 176 anal and 39 cervical swabs from 84 different subjects yielded unequivocal sequences of eight major HPV16 variants. Eight samples contained probable novel HPV16 variants and in 1 sample two variants were detected. In eight of 29 (27.6%) anal-cervical sample pairs positive for HPV 16, discordant variants were found. From 57 anal and 9 cervical sample series of HPV 16 positive samples a change in HPV16 variant status over time was seen in nine (15.8%) instances (7 anal, 2 cervical) from eight different participants. Changes of HPV 16 variants in HIV-infected adults was most frequently seen when different anatomic sites were sampled, but was also observed over time. |
Establishment of the 1st World Health Organization international standards for human papillomavirus type 16 DNA and type 18 DNA
Wilkinson DE , Baylis SA , Padley D , Heath AB , Ferguson M , Pagliusi SR , Quint WG , Wheeler CM , Collaborative Study Group , Unger E , Swan DC . Int J Cancer 2009 126 (12) 2969-83 ![]() A World Health Organization collaborative study was conducted to evaluate candidate International Standards for Human Papillomavirus (HPV) Type 16 DNA (NIBSC code 06/202) and HPV Type 18 DNA (NIBSC code 06/206) for use in the amplification and detection steps of nucleic acid-based assays. The freeze-dried candidate International Standards were prepared from bulk preparations of cloned plasmid containing full-length HPV-16 or HPV-18 genomic DNA. Nineteen laboratories from 13 countries participated in the study using a variety of commercial and in-house quantitative and qualitative assays. The data presented here indicate that, upon freeze-drying, there is no significant loss in potency for the candidate HPV-18 DNA and a slight loss in potency for the candidate HPV-16 DNA; although this is likely not scientifically relevant when assay precision is considered. In general, the individual laboratory mean estimates for each study sample were grouped +/- approximately 2 log(10) around the theoretical HPV DNA concentration of the reconstituted ampoule (1 x 10(7) HPV genome equivalents/mL). The agreement between laboratories is improved when potencies are made relative to the candidate International Standards, demonstrating their utility in harmonizing amplification and detection steps of HPV-16 and -18 DNA assays. Degradation studies indicate that the candidate International Standards are extremely stable and suitable for long-term use. Based on these findings, the candidate standards were established as the 1(st) WHO International Standards for HPV-16 DNA and HPV-18 DNA, each with a potency of 5 x 10(6) International Units (IU) per ampoule or 1 x 10(7) IU/mL when reconstituted as directed. (c) 2009 UICC. |
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