Stephen Cherne

Early Natural History of Incident, Type-Specific Human Papillomavirus Infections in Newly Sexually Active Young Women

Background: Characterizing short-term detection patterns of young womens incident α-genus human papillomavirus (HPV) infections may further our understanding of HPV transmission. Methods: Between 2000 and 2007, we followed 18- to 22-year-old female university students with triannual HPV DNA and Papanicolaou testing. Using Kaplan–Meier methods, we estimated duration of detectable, type-specific incident infections; time to redetection (among infections that became undetectable); and time to cervical lesion development after incident infection. We evaluated risk factors for short-term persistent versus transient infection with logistic regression. Results: Three hundred three incident, type-specific infections were detected in 85 sexually active women. Median time to first negative test after incident infection was 9.4 (95% CI: 7.8–11.2) months; 90.6% of infections became undetectable within 2 years. About 19.4% of infections that became undetectable were redetected within 1 year. Cervical lesions were common and 60% were positive for multiple HPV types in concurrent cervical swabs. Incident HPV detection in the cervix only (vs. the vulva/vagina only or both sites) was associated with short-term transience. Conclusions: Although most incident infections became undetectable within 2 years, redetection was common. Cervical lesions were a common early manifestation of HPV infection. Impact: It remains unclear whether potentially modifiable risk factors can be identified to reduce infection duration (and transmission likelihood). Cancer Epidemiol Biomarkers Prev; 20(4); 699–707. ©2011 AACR.

Emergence of Multiclass Drug–Resistance in HIV-2 in Antiretroviral-Treated Individuals in Senegal: Implications for HIV-2 Treatment in Resouce-Limited West Africa

BACKGROUND The efficacy of various antiretroviral (ARV) therapy regimens for human immunodeficiency virus type 2 (HIV-2) infection remains unclear. HIV-2 is intrinsically resistant to the nonnucleoside reverse-transcriptase inhibitors and to enfuvirtide and may also be less susceptible than HIV-1 to some protease inhibitors (PIs). However, the mutations in HIV-2 that confer ARV resistance are not well characterized. METHODS Twenty-three patients were studied as part of an ongoing prospective longitudinal cohort study of ARV therapy for HIV-2 infection in Senegal. Patients were treated with nucleoside reverse-transcriptase inhibitor (NRTI)- and PI (indinavir)-based regimens. HIV-2 pol genes from these patients were genotyped, and the mutations predictive of resistance in HIV-2 were assessed. Correlates of ARV resistance were analyzed. RESULTS Multiclass drug-resistance mutations (NRTI and PI) were detected in strains in 30% of patients; 52% had evidence of resistance to at least 1 ARV class. The reverse-transcriptase mutations M184V and K65R, which confer high-level resistance to lamivudine and emtricitabine in HIV-2, were found in strains from 43% and 9% of patients, respectively. The Q151M mutation, which confers multinucleoside resistance in HIV-2, emerged in strains from 9% of patients. HIV-1-associated thymidine analogue mutations (M41L, D67N, K70R, L210W, and T215Y/F) were not observed, with the exception of K70R, which was present together with K65R and Q151M in a strain from 1 patient. Eight patients had HIV-2 with PI mutations associated with indinavir resistance, including K7R, I54M, V62A, I82F, L90M, L99F; 4 patients had strains with multiple PI resistance-associated mutations. The duration of ARV therapy was positively associated with the development of drug resistance (P = .02). Nine (82%) of 11 patients with HIV-2 with no [corrected] detectable ARV resistance had undetectable plasma HIV-2 RNA loads (<1.4 log(10) copies/mL), compared with 3 (25%) of 12 patients with HIV-2 with detectable ARV resistance (P = .009). Patients with ARV-resistant virus had higher plasma HIV-2 RNA loads, compared with those with non-ARV-resistant virus (median, 1.7 log(10) copies/mL [range, <1.4 to 2.6 log(10) copies/mL] vs. <1.4 log(10) copies/mL [range, <1.4 to 1.6 log(10) copies/mL]; P = .003). CONCLUSIONS HIV-2-infected individuals treated with ARV therapy in Senegal commonly have HIV-2 mutations consistent with multiclass drug resistance. Additional clinical studies are required to improve the efficacy of primary and salvage treatment regimens for treating HIV-2 infection.

Detection of Genital HPV Types in Fingertip Samples from Newly Sexually Active Female University Students

Background: Little is known about detection of genital human papilloma virus (HPV) types in womens fingertips. The study objectives were to determine the presence of genital HPV types in fingertip samples and the agreement between fingertip and genital samples for detecting HPV. Methods: At triannual visits, genital and fingertip samples were collected from female university students and tested for 37 HPV genotypes by PCR-based assay. Type-specific concordance between paired fingertip and genital samples was evaluated using κ statistics for percent positive agreement (κ+). Paired samples with type-specific concordant fingertip and genital results were selected for variant characterization. Results: A total of 357 fingertip samples were collected from 128 women. HPV prevalence in fingertip samples was 14.3%. Although percent positive agreement between fingertips and genitals for detecting type-specific HPV was low (17.8%; κ+ = 0.17; 95% confidence interval, 0.10-0.25), 60.4% of type-specific HPV detected in the fingertips was detected in a concurrent genital sample. All but one of 28 paired concordant samples were positive for the same type-specific variant in the fingertip and genital sample. Redetection of HPV types at the subsequent visit was more common in genital samples (73.3%) than in fingertip samples (14.5%; P < 0.001). Conclusions: Detection of genital HPV types in the fingertips was not uncommon. Although impossible to distinguish between deposition of DNA from the genitals to the fingertips and true fingertip infection, the rarity of repeat detection in the fingertips suggests that deposition is more common. Impact: Finger-genital transmission is plausible but unlikely to be a significant source of genital HPV infection. Cancer Epidemiol Biomarkers Prev; 19(7); 1682–5. ©2010 AACR.

Validation for Clinical Use of a Novel HIV-2 Plasma RNA Viral Load Assay Using the Abbott m2000 Platform

BACKGROUND Optimal care of persons infected with human immunodeficiency virus type 2 (HIV-2) requires an accurate assessment of HIV-2 plasma viral load (VL), but no clinically approved quantitative HIV-2 RNA VL assay exists. OBJECTIVES To validate a novel quantitative HIV-2 RNA assay for clinical and research use. STUDY DESIGN The Abbott m2000sp/rt platform was adapted for quantification of HIV-2 RNA in plasma. Amplification targeted a region of the long terminal repeat conserved in Group A and B HIV-2. Electron microscopy-counted-HIV-2 standards, the WHO/NIBSC HIV-2 International Standard and clinical specimens (N=162) were used to determine the precision, sensitivity, specificity, linear range, accuracy, and clinical performance of the assay. RESULTS The quantitative linear range of the HIV-2 RNA assay was 10-1,000,000 copies/mL (R(2)>0.99), with a limit of detection of 8 copies/mL (95% CI, 5-18 copies/mL). The assay did not cross-react with HIV-1, and quantification of HIV-2 RNA was not affected by the presence of >5 log(10)HIV-1 RNA copies/mL. The total standard deviation (SD) and intra- and inter-run SD were 0.095, 0.093 and 0.162, respectively, at nominal inputs of 3.7, 1.7 and 1.0 log(10)HIV-2 RNA copies/mL. The HIV-2 WHO/NIBSC International Standard (1000 IU) was shown to contain 152 RNA copies/mL (95% CI 141-163). Overall, HIV-2 RNA was quantified at ≥10 copies/mL from 86 (53%) clinical specimens (median, 2.24 log(10) copies/mL; range 10-16,870), and nine specimens (6%) had HIV-2 RNA detected at <10 copies/mL. CONCLUSIONS We developed and validated a highly sensitive HIV-2 VL assay that is suitable for clinical and research use.

HIV-2 Integrase Variation in Integrase Inhibitor-Naïve Adults in Senegal, West Africa

Background Antiretroviral therapy for HIV-2 infection is hampered by intrinsic resistance to many of the drugs used to treat HIV-1. Limited studies suggest that the integrase inhibitors (INIs) raltegravir and elvitegravir have potent activity against HIV-2 in culture and in infected patients. There is a paucity of data on genotypic variation in HIV-2 integrase that might confer intrinsic or transmitted INI resistance. Methods We PCR amplified and analyzed 122 HIV-2 integrase consensus sequences from 39 HIV-2–infected, INI-naive adults in Senegal, West Africa. We assessed genetic variation and canonical mutations known to confer INI-resistance in HIV-1. Results No amino acid-altering mutations were detected at sites known to be pivotal for INI resistance in HIV-1 (integrase positions 143, 148 and 155). Polymorphisms at several other HIV-1 INI resistance-associated sites were detected at positions 72, 95, 125, 154, 165, 201, 203, and 263 of the HIV-2 integrase protein. Conclusion Emerging genotypic and phenotypic data suggest that HIV-2 is susceptible to the new class of HIV integrase inhibitors. We hypothesize that intrinsic HIV-2 integrase variation at “secondary” HIV-1 INI-resistance sites may affect the genetic barrier to HIV-2 INI resistance. Further studies will be needed to assess INI efficacy as part of combination antiretroviral therapy in HIV-2–infected patients.

Evaluation of Transported Dry and Wet Cervical Exfoliated Samples for Detection of Human Papillomavirus Infection

ABSTRACT We determined the feasibility of human papillomavirus (HPV) detection in cervical exfoliated cells collected as dry swab samples. Both dry cervical swab and specimen transport medium (STM) cervical swab samples were collected from 135 patients attending either colposcopy or womens clinics in Guayaquil, Ecuador, who had a cytology diagnosis within 6 months. HPV was detected by dot blot hybridization and genotyped by the liquid bead microarray assay (LBMA). Overall, 23.1% of dry samples were positive for any high-risk HPV types, and 24.6% of STM samples were positive for any high-risk HPV types. Of 125 paired samples, the type-specific high-risk HPV proportion positive agreement was 60.7% (kappa, 0.69; 95% confidence interval [CI], 0.53 to 0.82). Of six women with cytological evidence of invasive cervical cancer, high-risk HPV DNA was detected in three of their STM samples and in five of their dry samples. Dry samples were more likely to be insufficient for HPV testing than STM samples. Consistent with this observation, the amount of genomic DNA quantitated with the β-actin gene was almost 20 times lower in dry samples than in STM samples when detected by the real-time TaqMan assay; however, HPV DNA viral loads in dry samples were only 1.6 times lower than those in matched STM samples. We concluded that exfoliated cervical cells could be collected as dry swab samples for HPV detection.

Human Papillomavirus Types 16 and 18 DNA Load in Relation to Coexistence of Other Types, Particularly Those in the Same Species

Background: Infection with multiple human papillomavirus (HPV) types is common. However, it is unknown whether viral DNA load is related to the coexistence of other types. Methods: Study subjects were 802 and 303 women who were positive for HPV16 and HPV18, respectively, at enrollment into the Atypical Squamous Cells of Undetermined Significance and Low-Grade Squamous Intraepithelial Lesion Triage Study. HPV16 and HPV18 E7 copies per nanogram of cellular DNA in cervical swab samples were measured by real-time PCR in triplicate. Results: Concurrent coinfection was common in this population of women with minor cervical lesions; multiple HPV types were detected in 573 (71.4%) of 802 HPV16-positive women and 227 (74.9%) of 303 HPV18-positive women. The adjusted odds ratio associating coinfection with per 1 log unit increase in HPV16 DNA load was 0.78 (95% confidence interval, 0.68-0.89); it was 0.64 (95% confidence interval, 0.52-0.79) for a similar analysis of HPV18 DNA load. Women with, compared with without, coinfection of A9 species types possessed a significantly lower HPV16 DNA load (P < 0.001), whereas women with, compared with without, coinfection of A7 species types possessed a significantly lower HPV18 DNA load (P = 0.001). A trend of decrease in HPV16 DNA load with increasing number of the coexisting non-HPV16 A9 species types was statistically significant (Ptrend = 0.001). Conclusion: Coinfection with other types was associated with lower HPV16 and HPV18 DNA load. The extent of reduction was correlated to phylogenetic distance of the coexisting types to HPV16 and HPV18, respectively. (Cancer Epidemiol Biomarkers Prev 2009;18(9):2507–12)

Protocol for the Detection and Genotyping of Human Papillomaviruses Using a Liquid Bead Microarray Assay

More than 100 human papillomavirus (HPV) types have been identified, and over 40 of them infect the anogenital epithelium. Because each HPV type is associated with different risks for the development of cervical cancer, detecting and genotyping HPVs has increasingly become an integral part of cervical cancer control. Here, we describe a Luminex assay-based liquid bead microarray assay for genotyping 37 HPV types, which is objective, scalable, amenable to a high-throughput configuration, and has the potential to be automated.

Human papillomavirus type 16 viral load in relation to HIV infection, cervical neoplasia and cancer in Senegal.

BACKGROUND The importance of human papillomavirus (HPV) viral load in the pathogenesis of cervical cancer among HIV-infected and HIV-uninfected women has not yet been established. METHODS In this cross-sectional study, HPV-16 viral loads were measured using previously-collected and frozen cervical swab samples from 498 HPV-16 positive Senegalese women (368 HIV-seronegative, 126 HIV-1 and/or HIV-2 seropositive). The real-time polymerase chain reaction assay was used to quantify HPV-16 E7 copy number normalized by human cellular DNA (β-actin), and viral loads were log10 transformed. Associations between HPV-16 viral load, degree of cervical abnormality, and HIV status were assessed using multinomial and linear regression methods. RESULTS Compared to women with normal cytology, the likelihood of CIN1 (ORa: 1.21, 95% CI 0.93-1.57), CIN2-3 (ORa: 2.38, 95% CI 1.72-3.29) and cancer (ORa: 2.12, 95% CI 1.52-2.96) was found to increase for each 1-unit log10 increase in HPV-16 viral load. Compared to HIV-negative women, HIV-positive women had higher average HPV-16 viral load values (βa: 0.39, 95% CI 0.03-0.75), even after accounting for degree of cervical abnormality. CONCLUSION In our study of women including those with cancer, HPV-16 viral load was associated with a higher likelihood of cervical abnormalities. However, substantial overlaps across categories of disease severity existed. Higher viral load among HIV-infected individuals may indicate that HIV infection influences HPV viral replication factors.

Human Papillomavirus Prevalence Among American Indian Women of the Great Plains

BACKGROUND High-risk human papillomavirus (hrHPV) causes cervical cancer. In the United States, approximately 40% of women aged 14-59 years from all racial and ethnic groups are infected with HPV, and prevalence typically declines with age. However, American Indian (AI) women are insufficiently sampled to permit a population-specific estimate of hrHPV prevalence. METHODS Vaginal swabs were self-collected by 698 AI women aged 21-65 years from a tribal community in the Great Plains. We estimated the population prevalence of hrHPV and identified predominant genotypes. RESULTS The combined prevalence of hrHPV genotypes 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, and 68 was 34.8%. HPV-51 (7.6%), HPV-58 (5.3%), HPV-52 (4.3%), HPV-18 (4.3%), and HPV-16 (3.9%) were most prevalent. hrHPV prevalence declined with age, from 42.2% in women aged 21-24 years to 27.9% in women aged 50-65 years. CONCLUSIONS HPV-51 was the single most prevalent oncogenic genotype. The combined prevalence of hrHPV among AI women in our sample was high, particularly among women aged 50-65 years, for whom hrHPV prevalence was approximately triple that of other races. Cervical cancer screening efforts should be increased, particularly among women from the community aged 30 years and older.