We report a unique primary cervical neoplasm in a 44-yr-old woman which we believe, based on the morphology and immunophenotype, represents an extremely unusual small cell variant of paraganglioma. This represents the first report of a primary cervical paraganglioma. Following chemoradiation treatment, the tumor underwent malignant transformation into an S100 and SOX10 positive sarcoma, morphologically and immunohistochemically resembling a malignant peripheral nerve sheath tumor, which we believe represents a sarcoma derived from the sustentacular cells of the paraganglioma. Mutational analysis detected a nonsense mutation of NF1 gene in the sarcoma. This further supports the diagnosis as both somatic and germline NF1 mutations have been associated with paragangliomas and malignant peripheral nerve sheath tumors. Targeted RNA sequencing (ARCHER, expanded sarcoma panel) covering many known genes implicated in sarcoma development, did not reveal any other molecular alteration (fusion or internal tandem duplication).

El cáncer cervical es la malignidad más común entre las mujeres del tercer mundo y en Guatemala, es responsable del 40% de todos los cánceres en ambos sexos y del 60% de todos los cánceres femeninos, La presencia de Virus del Papiloma Humano (VPH) se considera entre otras, como el factor de riesgo más importante para el desarrollo del cáncer cervical. El presente estudio fue llevado a cabo con el propósito de determinar la prevalencia de los tipos de VPH en un grupo de pacientes de Guatemala con cáncer cervical así como para evaluar los factores de riesgo asociados al desarrollo de la enfermedad. Con ese propósito se llevó a cabo un estudio de casos y controles en 112 pacientes con cáncer cervical invasivo (casos) y 102 mujeres sanas (controles). Se obtuvieron biopsias cervicales y se investigó la presencia de, VPH usando dos métodos moleculares, captura de híbridos 2 (HC-2™. Digene Co.) y la reacción en cadena de la polimerasa (PCR). Se utilizó un cuestionario epidemiológico para evaluar otros factores de riesgo agrupados en factores de comportamiento sexual, factores reproductivos y factores sociodemográficos. La información fue recolectada durante una entrevista personal con los sujetos de estudio. Un total de 89% de las pacientes y tin 18% de los controles fueron positivos para ADN del Virus del Papiloma Humano y los tipos detectados con mayor frecuencia fueron el VPH 16, 18 y 45. La evaluación de factores de riesgo demostró valores de p de tendencia significativos (mayor riesgo de desarrollar cáncer cervical) para factores como: edad temprana de inicio de relaciones sexuales, número elevado de compañeros sexuales durante toda la vida y compañeros estables, elevado número de embarazos y partos vaginales, baja escolaridad y un origen étnico categorizado como indígena, al contrario de otros como el control prenatal durante el último embarazo, el cual es un factor que reduce el riesgo.

BK virus (BKV), a common human polyomavirus infection latent in the kidneys, can reactivate with immunosuppression to cause renal disease. Some have suggested that BKV may contribute to the development of bladder cancer, and BKV sequences have been reported from bladder tumors. To further examine the role of BKV in human bladder cancer, a series of bladder tumors was investigated for BKV genomic sequences. Fresh‐frozen specimens from 76 transitional cell carcinoma tissues and 46 paired adjacent normal urothelial tissues archived at the H. Lee Moffitt Cancer Center were studied. All tissues were histopathologically reviewed. DNA extracted from the tissues was tested by quantitative real‐time polymerase chain reaction (QPCR) assays to detect BKV DNA sequences in the VP1 coding region. Amplification of ERV‐3 was conducted separately to quantify cell copy number. Conventional PCR targeting the BKV T‐antigen (T‐Ag) coding region and immunohistochemistry for BKV T‐Ag were also conducted on all tissues that tested positive for BKV by QPCR. Seventy‐three bladder tumors yielded ≥3,000 copies of ERV‐3, 4 (5.5%) of which tested positive for BKV with average copy numbers of 7.9, 15.8, 0.4 and 0.3 per 1,000 cells. Paired normal tissue was available for 2 of these BKV‐positive tumors, 1 of which was BKV‐positive (14.6 copies/1,000 cells). No other normal tissues were BKV‐positive by QPCR. The 6 BKV‐positive tissues by QPCR were also positive by conventional PCR, but all stained negative for BKV T‐Ag by immunohistochemistry. BKV is unlikely to be involved in the etiology of most bladder tumors. © 2006 Wiley‐Liss, Inc.

BACKGROUND:p53 mutations are relatively uncommon in medulloblastoma, but abnormalities in this cell cycle pathway have been associated with anaplasia and worse clinical outcomes. We correlated p53 protein expression with pathological subtype and clinical outcome in 75 embryonal brain tumors. The presence of JC virus, which results in p53 protein accumulation, was also examined.METHODS:p53 protein levels were evaluated semi-quantitatively in 64 medulloblastomas, 3 atypical teratoid rhabdoid tumors (ATRT), and 8 supratentorial primitive neuroectodermal tumors (sPNET) using immunohistochemistry. JC viral sequences were analyzed in DNA extracted from 33 frozen medulloblastoma and PNET samples using quantitative polymerase chain reaction.RESULTS:p53 expression was detected in 18% of non-anaplastic medulloblastomas, 45% of anaplastic medulloblastomas, 67% of ATRT, and 88% of sPNET. The increased p53 immunoreactivity in anaplastic medulloblastoma, ATRT, and sPNET was statistically significant. Log rank analysis of clinical outcome revealed significantly shorter survival in patients with p53 immunopositive embryonal tumors. No JC virus was identified in the embryonal brain tumor samples, while an endogenous human retrovirus (ERV-3) was readily detected.CONCLUSION:Immunoreactivity for p53 protein is more common in anaplastic medulloblastomas, ATRT and sPNET than in non-anaplastic tumors, and is associated with worse clinical outcomes. However, JC virus infection is not responsible for increased levels of p53 protein.

Esophageal squamous cell cancer is highly prevalent in south-western Kenya. The role of human papillomavirus (HPV) in esophageal cancers from this region was evaluated. Biopsies of 29 esophageal squamous cell cancers were assayed for HPV DNA sequences by reverse line blot polymerase chain reaction, using 27 HPV type-specific probes. Viral sequences were found in none of the specimens. These results suggest the HPV is unlikely to be an etiologic factor for esophageal squamous cell cancers in this region.

Purpose: Persistent infection with oncogenic human papillomaviruses (HPV) plays a central etiologic role in the development of squamous carcinomas of the cervix and their precursor lesions, cervical intraepithelial neoplasias (CIN). We carried out a prospective observational cohort study evaluating known, quantifiable prognostic variables of clinical behavior in women with high-grade cervical lesions.Experimental Design: Our study cohort included healthy women with high-grade cervical lesions (CIN2/3) with residual visible lesions after colposcopically directed biopsy. We prospectively followed 100 women over 15 weeks before standard resection. HPV typing was done using PCR and a reverse line blot detection method.Results: The rate of spontaneous histologic regression, defined as (CIN1 or less at resection) was 28%. The overall rate of HPV infection was 100%. HPV16 was identified in 68% of the lesions. Women with HPV16 only were significantly less likely to regress, compared with women with HPV types other than HPV16 (odds ratio, 0.342; 95% confidence interval, 0.117-0.997; P = 0.049). In the cohort with HPV16 only, patients who had an HLA*A201 allele had similar outcomes to those who did not carryA201. However, among patients with HPV types other than HPV16, the HLA*A201 allele interaction was significant; patients with HLA*A201 were the least likely to resolve.Conclusions: CIN2/3 lesions associated with HPV16 alone are significantly less likely to resolve spontaneously than those caused by other types. Interactions among HPV type, HLA type, and regression rate support a role for HLA-restricted HPV-specific immune responses in determining disease outcome.

Before 1963, poliovirus vaccine produced in the United States was contaminated with simian virus 40 (SV40), which causes cancer in animals. To examine whether early-life SV40 infection can cause human cancer, the authors studied 54,796 children enrolled in the US-based Collaborative Perinatal Project (CPP) in 1959-1966, 52 of whom developed cancer by their eighth birthday. Those children whose mothers had received pre-1963 poliovirus vaccine during pregnancy (22.5% of the children) had an increased incidence of neural tumors (hazard ratio = 2.6, 95% confidence interval: 1.0, 6.7; 18 cases) and hematologic malignancies (hazard ratio = 2.8, 95% confidence interval: 1.2, 6.4; 22 cases). For 50 CPP children with cancer and 200 CPP control children, the authors tested paired maternal serum samples from pregnancy for SV40 antibodies using a virus-like particle enzyme immunoassay and a plaque neutralization assay. Overall, mothers exhibited infrequent, low-level SV40 antibody reactivity, and only six case mothers seroconverted by either assay. Using the two SV40 assays, maternal SV40 seroconversion during pregnancy was not consistently related to children's case/control status or mothers' receipt of pre-1963 vaccine. The authors conclude that an increased cancer risk in CPP children whose mothers received pre-1963 poliovirus vaccine was unlikely to have been due to SV40 infection transmitted from mothers to their children.

The prevalence and risk factors for oral human papillomavirus (HPV) infection are unknown, despite evidence for an etiological role for HPV in oral cancers. Oral samples from human immunodeficiency virus (HIV)-seronegative (n=396) and HIV-seropositive (n=190) adults were tested for HPV DNA. High-risk HPV infections were present in 2.1% of tonsil and 6.3% of oral-rinse specimens. The prevalence of oral high-risk HPV infection was greater in HIV-seropositive individuals (13.7% vs. 4.5%; P<.001). In multiple logistic regression, odds of oral HPV infection increased with age, male sex, and herpes simplex virus (HSV)-2 seropositivity in HIV-seronegative individuals and with CD4 cell count <200 cells/mL, HSV-2 seropositivity, oral mucosal abnormalities, and >1 oral sex partner during the previous year (odds ratio, 12.8; 95% confidence interval, 3.1-52.7) among HIV-seropositive individuals. HPV type 16, which is present in most HPV-associated tonsillar cancers, was the most prevalent high-risk oral HPV infection.

Genomic sequences of the human polyomaviruses, JC virus (JCV) and BK virus (BKV), and simian virus 40 (SV40) have been reported from several types of human brain tumors, but there have been no population-based seroepidemiologic studies to evaluate the association between polyomavirus infection and brain tumors. We conducted a case-control study, nested within a prospective cohort, to investigate the association between antibodies to JCV, BKV, and SV40, as measured in serum collected 1-22 years before diagnosis and incident primary malignant brain tumors. Brain tumor cases (n = 44) and age-, gender-, and race-matched controls (n = 88) were identified from participants of two specimen banks in Washington County, Maryland. IgG antibodies to the capsid proteins of JCV and BKV were assessed using ELISAs. SV40-neutralizing antibodies were measured using plaque neutralization assays. Similar to the general population, the prevalence of JCV and BKV infection was high in our study population (77 and 85%, respectively). Antibodies to SV40 were less prevalent (11%). The odds ratio for subsequent brain tumor development was 1.46 [95% confidence interval (CI), 0.61-3.5] for JCV, 0.66 for BKV (95% CI, 0.22-1.95), and 1.00 for SV40 (95% CI, 0.30-3.32). Given the high prevalence of JCV and BKV infections and the millions who were potentially exposed to SV40 through contaminated polio vaccines, future studies should attempt to replicate these findings.

Carbone et al. advance a series of misstatements in criticizing our recently published study.1 Contrary to their assertions, our study provides evidence that simian virus 40 (SV40) is unlikely to be a cause of brain tumors in northern India. We respond to their numbered comments in order. 1. Carbone et al. state that the low SV40 copy number amplified from the positive controls (formalin-fixed mouse tumors) indicates either that we added very little DNA to our PCR reaction mixture or that our assay was insensitive. However, the possibility that SV40 was inefficiently amplified due to the effects of formalin fixation was not considered by the correspondents. As we write in our article, “The early phase of PCR amplification kinetics is susceptible to the DNA-degrading effects of formalin fixation, so the copy number derived from the growth curves for formalin-fixed tissues may not correspond to that for a similar amount of SV40 DNA present in the titration series.”1 Because of the effects of formalin fixation, one cannot directly extrapolate from an SV40 DNA titration series to estimate the number of SV40 copies present in a tumor sample. To address this problem, we simultaneously used quantitative PCRs for SV40 and a human gene, each standardized against an appropriate dilution series, to measure the amount of SV40 in tissue relative to cell content. Thus, as we write, we consider these results “semiquantitative.”1 In this regard, we chose the mouse tumors as positive controls to mimic the conditions of archived human tissues, i.e., small amounts of formalin-fixed tissue. Under these conditions, filter hybridizations from all 7 SV40-positive mouse tumors were positive (Fig. 2 of our article). The quantitative PCR also detected SV40 DNA in 6 of these tumors. The low absolute level that we measured (equivalent to < 10 copies of SV40 per reaction) likely arose from a combination of low tissue input and the effects of formalin fixation. Rather than indicate an insensitivity of our assay, as Carbone et al. claim, the published results demonstrate that, even in such challenging circumstances, our quantitative PCR is highly sensitive in detecting SV40. 2. In their second criticism, the logic of Carbone et al. is less clear. They apparently acknowledge that formalin fixation of tissues decreases the quality of DNA and leads to a lower quantitative PCR estimate of SV40 copy, as we indicate above. The correspondents then suggest that the DNA-degrading effects of fixation might require more than 50 cycles of PCR to detect SV40 in positive tumors. However, using only 50 PCR cycles, we consistently identified SV40-positive mouse tumors using both quantitative PCR and filter hybridization. Also, as we discuss next, the simultaneous PCR-based quantification of a human gene indicated that, despite the technical difficulties created by the formalin fixation, adequate amplifiable DNA was present in our experiment. 3. As mentioned, we standardized the measured SV40 copy number to the number of detected copies of a human cellular gene, glyceraldehyde-3 phosphate dehydrogenase (GAPDH). In our article, we mistakenly refer to this enzyme as glyceraldehyde-6-phospate dehydrogenase, and we thank the correspondents for correcting our typographical error. More substantively, Carbone et al. suggest that the presence of GAPDH pseudogenes could have caused us to overestimate the number of cell-equivalents of amplifiable DNA present in our reaction mixtures. We admit that we had not adequately considered this possibility. Based on a 1984 article, Carbone et al. point to the possible existence of 25 copies of GAPDH-“homologous” sequences in the human genome.2 However, by BLAST search, we identified only 4 known sequences in the human genome that matched our primers and that would result in amplification of the observed product size. The uncertainty raised by the limited data on this question prompted us to compare results obtained using our GAPDH quantitative PCR with those from a similar quantitative PCR assay for ERV-3,3 an endogenous retrovirus present in 1 copy per haploid genome. In duplicate, we prepared 60 aliquots of DNA extracted from Ramos cells, corresponding to approximately 50 to 4,000 cells per aliquot. These aliquots were assayed independently in separate laboratories for GAPDH and ERV-3 copy number. The GAPDH copy number was consistently greater than the ERV-3 copy number (median 9,300, range 700–99,000, vs. 1,200, range 120–8,300). The median GAPDH/ERV-3 ratio was 7.0, generally consistent with the results of the BLAST search. These observations lead us to recalculate 2 results from our study.1 First, in the single ependymoma that provided an unconfirmed positive result, we now estimate that SV40 was present in at most 1 copy per 50 cells (i.e., < 10 copies/[7,009 GAPDH copies/14 GAPDH copies per cell]). Second, based on the observed GAPDH copy numbers, 50% of our evaluated tumor specimens had at least 105 cell-equivalents of amplifiable DNA (i.e., median GAPDH copies 1,476/14 GAPDH copies per cell). None of the tumors had detectable SV40. Thus, the experiment ruled out SV40 at a level of more than 1 copy per 10 cells (i.e., 10 copies/105 cells). These 2 negative results, coupled with the negative filter hybridization,1 remain convincing arguments against the presence of SV40 in these tumors. Two additional comments by the correspondents deserve a brief response. They point out that K562 cells are triploid. However, we used K562 cells only as a ready source of human DNA for PCR experiments. Specifically, the GAPDH copy number in DNA extracted from these cells was standardized to a known concentration of a plasmid containing a single copy of GAPDH. Therefore, the ploidy of the source of DNA used in the standard is not relevant. Carbone et al. also argue that only a portion of our specimens consisted of tumor. However, our pathologic review of tissue sections adjacent to those evaluated by PCR indicated that all of the specimens were comprised predominantly of tumor cells. 4. Carbone et al. mistakenly write that we suggest that our assay could have detected 10 copies of SV40 in fixed tissue specimens. As stated already, we acknowledge that formalin fixation reduces the sensitivity of PCR. Thus, we do not believe that a measured amount of SV40 DNA amplified from formalin-fixed tissue would correspond to the amount that would have been amplifiable in the same tissue when fresh. However, because formalin fixation likely affects SV40 and GAPDH amplification equally, the absence of quantifiable SV40 DNA, when readily measurable GAPDH was present, is strong negative evidence. In coupling the SV40 and GAPDH PCR results, we rule out SV40 at a level of 10 or more copies per 105 cells. 5. In a final criticism, Carbone et al. claim that our study was not truly masked, based on an examination of our Figure 2, which shows the titration series and water controls placed consecutively. However, this assertion reflects another misunderstanding, because it was only the human tissues (brain tumor cases and other brain tissues that served as negative controls) that were masked and sorted in a random order. For example, in Figure 2 of our study,1 tissues in row A (columns 3–12) correspond to: tumor, normal, tumor, tumor, tumor, tumor, normal, tumor, normal, tumor. Because these tissues were randomly sorted and masked before extraction and PCR testing, our experiment is an appropriate test of whether SV40 is preferentially present in human tumors over tissues from other, presumably SV40-unrelated, brain conditions. Carbone et al argue that the sensitivity of our methods was limited. While acknowledging the challenges in working with archival tumor tissue, we have presented evidence that our experiment had adequate sensitivity to effectively rule out the presence of SV40 in Indian brain tumors. It is appropriate, then, to briefly highlight our concerns regarding published studies that have reported the detection of SV40 in brain tumors (see Table 1 of our article).1 Again we point out that an important weakness in these studies was that many lacked a control series of human tissues in which SV40 was expected, a priori, not to be present. Also, none of the studies evaluated tissues in a manner masked to tissue status (tumor vs. control). These limitations call into question the specificity of SV40 detection, which has been reported variably for diverse tumors (including but not limited to various brain tumors, sarcomas, mesothelioma, lymphoma) and normal tissues. This concern could be addressed rather simply, by including negative control tissues, evaluated in a masked manner concurrently with tumors of interest, analogous to our experiment. Importantly, in a masked multiinstitutional study evaluating mesothelioma specimens, the frequency of detection of SV40 sequences was very low and identical to that in masked controls.4 As we indicate, the quantification of SV40 DNA in evaluated tissue specimens, while rarely undertaken, can provide further relevant information regarding biologic plausibility.1 In their letter, Carbone et al. suggest that Southern blots can provide an approximate quantification of the amount of SV40 DNA present in cell lines. If this approach is feasible, it would be important to examine similar data for mesothelioma or brain tumor specimens obtained from humans. Of interest, Gorden et al. recently used a quantitative PCR assay similar to ours to detect and quantify SV40 DNA in U.S. mesothelioma specimens.5 Notably, SV40 DNA was present in only 2 specimens (6%), at a level of less than 1 copy per 134 cells. Overall, the suggested presence of SV40 DNA in a wide range of tumors and normal tissues, detected variably across laboratories and in at most extremely low levels, calls reported associations into question. We recognize the heated controversy surrounding the question of SV40 in human tumors. Therefore, it was inappropriate for the correspondents to suggest that there is resolution or consensus on this matter within the scientific community. Carbone et al. distort the record when they imply that the Institute of Medicine report concluded that there is compelling evidence in support of an association between SV40 and cancer. That report noted that “The conflicting results in the detection of SV40 have also led to questions about technical aspects of the detection of the virus. It is unclear whether positive findings are the result of overly sensitive but nonspecific tests that are detecting other viruses (i.e., BK or JC) or SV40 from laboratory contamination …”6 Carbone et al. also distort the record in their summary of a published commentary on a recent National Cancer Institute workshop.7 In that commentary, Wong et al. wrote “Many epidemiologists and some laboratory investigators remain unconvinced of the validity or consistency of detection of SV40 in human tumors or the significance of the various reported detections.”7 Given widespread doubt among scientists, it then seems remarkable that the “international consensus” meeting, organized by Dr. Carbone and colleagues in 2001, could have reached the conclusion that “there is now overwhelming evidence proving that SV40 is capable of infecting humans and that SV40 is present in some mesotheliomas …”8 Absent consensus, we agree with Wong et al. that “More research is needed to resolve the controversial issues … of whether SV40 is involved in human cancers.”7 We urge that future laboratory studies of SV40 in human cancers be conducted using rigorous designs that incorporate appropriate negative controls and masking. The authors thank Wendell Miley (Viral Epidemiology Section, AIDS Vaccine Program, National Cancer Institute-Frederick, Frederick, MD) for conducting the ERV-3 assays. Yours sincerely, Eric A. Engels*, Patti E. Gravitt* , Richard W. Daniel , Martha Quezado , Keerti V. Shah , * Division of Cancer Epidemiology and Genetics, National Cancer Institute, Rockville, MD, USA, Johns Hopkins Bloomberg School of Public Health, Baltimore, MD, USA, Department of Pathology, National Cancer Institute, Rockville, MD, USA.

Forty-two specimens from oropharyngeal (tonsil and base of tongue) squamous cell carcinoma patients (SCC) were studied for presence of HPV 16 by in situ hybridization and by immunohistochemistry for p53 and Cyclin D1 protein overexpression. Thirty-one per cent of cases were HPV-16 positive, which correlates with the prevalence reported worldwide. 74% of cases showed p53 protein overexpression and 79% showed Cyclin D1 overexpression. There was no correlation between HPV status and either p53 or Cyclin D1 overexpression (P>0.05). These three variables also did not correlate with factors such as grade of the tumour, stage of the disease or lymph nodal metastasis at presentation.

ABSTRACT Enzyme immunoassays (EIAs) for detection of serum antibodies to simian virus 40 (SV40), BK virus (BKV), and JC virus (JCV) were developed by using virus-like-particles (VLPs) produced in insect cells from recombinant baculoviruses expressing the VP1 protein of the respective virus. Rhesus macaque sera with neutralizing antibodies to SV40 showed a high level of reactivity in the SV40 VLP-based EIA, and these sera also showed lower levels of reactivity in the BKV and JCV VLP-based EIAs. Rhesus macaque sera negative for neutralizing antibodies to SV40 were negative in all three EIAs. Competitive binding assays showed that SV40 VLPs inhibited BKV reactivity. In rhesus macaque sera, high optical density (OD) values for antibodies to SV40 VLPs were correlated with high OD values for antibodies to BKV but not with high OD values for antibodies to JCV VLPs. Human sera with neutralizing antibodies to SV40 were more reactive to SV40 VLPs than human sera without neutralizing antibodies to SV40. The greater SV40 reactivities of human sera were correlated with greater reactivities to BKV VLPs but not JCV VLPs. These data suggest that cross-reactivity with BKV antibodies may account for part of the low-level SV40 reactivity seen in human sera. With their greater versatility and their suitability for large-scale testing, the VLP-based EIAs for SV40, BKV, and JCV are likely to contribute to a better understanding of the biology of these viruses.

Simian virus 40 (SV40), a monkey polyomavirus, was a contaminant of early poliovirus vaccines administered to millions of individuals in the 1950s and early 1960s. SV40 causes brain tumors in laboratory animals, and SV40 DNA sequences have been variably identified in human choroid plexus tumors and ependymomas. We studied the possible association between SV40 and human brain tumors in northern India, where humans have frequent contact with SV40-infected rhesus macaques. DNA from pathologic specimens from 33 ependymomas, 14 choroid plexus tumors and 18 control brain tissues (contused brain, brain metastases) was extracted and analyzed under masked conditions. We used real-time PCR to detect and quantify SV40 (T antigen) and human (GAPDH) DNA sequences. The SV40 PCR assay detected as few as 10 copies of SV40 DNA and had a linear range from 1 x 10(2) to 1 x 10(6) copies. SV40 DNA was detected in 1 specimen (an ependymoma). However, few SV40 DNA copies were detected in this sample (<10 copies, equivalent to <1 copy/350 cells, based on simultaneous GAPDH quantification), and SV40 was not detected when this sample was retested. Our findings do not support a role for SV40 in choroid plexus tumors or ependymomas from northern India.

Human immunodeficiency virus (HIV) infection and related immunosuppression are associated with excess risk for cervical neoplasia and human papillomavirus (HPV) persistence. Type-specific HPV infection was assessed at 6-month intervals for HIV-positive and HIV-negative women (median follow-up, 2.5 and 2.9 years, respectively). The type-specific incidence of HPV infection was determined, and risk factors for HPV persistence were investigated by statistical methods that accounted for repeated measurements. HIV-positive women were 1.8, 2.1, and 2.7 times more likely to have high-, intermediate-, and low-risk HPV infections, respectively, compared with HIV-negative women. In multivariate analysis, high viral signal, but not viral risk category, was independently associated with persistence among HIV-positive subjects (odds ratio [OR], 2.5; 95% confidence interval [CI], 2.1-2.9). Furthermore, persistence was 1.9 (95% CI, 1.5-2.3) times greater if the subject had a CD4 cell count <200 cells/<mu>L (vs. >500 cells/muL). Thus, HIV infection and immunosuppression play an important role in modulating the natural history of HPV infection.

This nine-laboratory multicenter investigation was designed to assess the sensitivity, specificity, and reproducibility of previously described assays for detection of SV40 DNA with three goals, i.e., (a) to compare methods for testing human tissues, (b) to examine the ability of these methods to detect SV40 in human mesotheliomas, and (c) to uncover assay differences that could explain conflicting findings in some past investigations. Each laboratory received, in a masked fashion, paired replicate DNA samples extracted from 25 fresh frozen mesotheliomas (50 samples) and one from each of 25 normal human lungs. Interspersed were masked positive (titrations of the SV40 genome) and negative control samples. Preliminary studies confirmed the adequacy of the samples for testing high molecular weight double-stranded linear DNA targets. All 15 PCR-based assays detected 5,000 copies or less of the SV40 genome spiked into 2 microg of WI-38 DNA. A high level of specificity and reproducibility was found among the PCR assays performed in most laboratories. However, none of the selected normal human lung tissue or the 25 mesothelioma tumor specimens obtained from archival samples at a single center was reproducibly positive for the presence of SV40 DNA. Further studies are needed to reconcile these results with previous reports of detection of SV40 DNA in tumor specimens.

OBJECTIVE:Human papillomavirus (HPV) assays are likely to be used with increasing frequency in clinical management of women with abnormal Papanicolaou smears and in cervical cancer screening. Our objective was to simplify the method of collection of female genital tract specimens. The utility of vaginal dry swabs for HPV diagnosis was evaluated.METHODS:Specimens for cytology and for HPV identification were collected by a clinician from 189 female soldiers attending a military clinic. Three methods of specimen collection for HPV identification were compared: a vaginal dry swab (v-DRY), and vaginal and cervical swabs placed into specimen transport medium (v-STM and c-STM). Swabs were shipped to a STD laboratory for processing. Specific HPV types were identified by a consensus primer based PCR based method. Results from 165 women were evaluable.RESULTS:HPV prevalence by the three methods was similar and ranged from 44.8% to 50.9%. 53 (32.1%) women were HPV positive and 60 (36.4%) women were HPV negative by all three collection methods. With respect to the risk categories of specific HPV types, there was greater agreement between the results from the two vaginal (v-DRY and v-STM) samples (kappa values of 0.69-0.81) than between the cervical (c-STM) and either of the vaginal samples (kappa values of 0.37-0.55). The HPV yield from c-STM was somewhat greater than that from the vaginal specimens but the correlation between cytological abnormalities and HPV was high for all three methods.CONCLUSION:A dry vaginal swab may be an acceptable method of specimen collection for HPV diagnosis.

Background: polymerase chain reaction (PCR)-based assays for human papillomavirus (HPV) sequences are in wide use in clinical and epidemiological studies. The reproducibility of these assays is not extensively studied. Objectives: to estimate the intra-laboratory reproducibility of generic and type-specific HPV diagnoses by the MY09/MY11/HMB01 consensus L1 primer-based PCR assay. Study design: systematically collected specimens (n=207) were masked and retested. Results: when specimens negative in both initial and repeat assays were excluded from analysis, the diagnostic reproducibility was 98.6% for β-globin, 90.7% for generic HPV (any HPV type), and 76.9% for type-specific HPVs. The reproducibility of type-specific diagnosis increased with increase in signal strength in the hybridization reaction of the initial assay. When a specimen contained five or more HPV types in the initial assay, it was rare to identify all of the HPV types in the repeat assay. Conclusions: the degree of reproducibility of the PCR diagnosis should be taken into account in the interpretation of HPV data in clinical and epidemiological studies.