Analysis of human papillomavirus types in exophytic condylomata acuminata by hybrid capture and Southern blot techniques.

Exophytic condylomata acuminata of the external genitalia of 40 patients were analyzed for human papillomavirus (HPV) DNA by the Southern blot and hybrid capture methods. All lesions were initially analyzed by the Southern blot method by using a mixture of HPV type 6, 11, 16, and 18 whole genomic probes. Southern blots demonstrated characteristic PstI restriction patterns of HPV type 6, 11, or 16 in all but one lesion. HPV 6 subtypes accounted for 28 of 39 HPV-positive lesions. Twenty-seven of these 28 lesions contained HPV type 6a, and 1 lesion contained HPV type 6c. Eight lesions contained HPV type 11 and three contained HPV type 16. Two of the three condylomata acuminata containing HPV type 16 were obtained from solid-organ transplant recipients receiving immunosuppressive medications. The third lesion containing HPV type 16 was a typical exophytic condyloma acuminatum from a woman with previously resected vulvar carcinoma. The hybrid capture assay detected HPV DNAs in all lesions except the Southern blot-negative lesion. Twenty-five lesions were positive for the A probe only (HPV types 6 and 11 and related types). All of these lesions were found to contain HPV type 6 or 11 sequences in the Southern blot assay. The remaining 14 lesions were positive for both the A probe and the B probe (HPV types 16 and 18 and related types). The strongest signal in these 14 lesions by the hybrid capture assay was consistent with the result of the Southern blot assay in all but one case. We conclude that (i) HPV type 6a is the most common type found in these lesions, (ii) HPV type 16 may be present more often in exophytic condylomata acuminata from immunosuppressed individuals, (iii) hybrid capture is a useful tool for documenting the presence of HPV sequences in DNAs from exophytic condylomata acuminata, and (iv) in samples containing multiple HPV types, hybrid capture allows detection of minority HPV types.     

Characterization of human papilloma virus types in condylomata acuminata in children by in situ hybridization

The presence of human papilloma virus (HPV) DNA sequences of types 6, 11, 16, and 18 was determined by in situ hybridization under stringent conditions on archival paraffin-embedded tissue sections in eight condylomata acuminata observed in children below the age of 12 years. Viral sequences were detected in seven of eight cases: all of them contained HPV 6, four contained also HPV 11, and one contained HPV 16 and 18. Papilloma virus common antigen was detected in only three of eight cases, all of them being positive also by in situ hybridization. We conclude that most condylomata acuminata in children are associated with the same types found in anogenital lesions in adults. Since little is known about the long-term significance of genital condylomas in children the identification of the papilloma virus type may prove to be important as a prognostic tool particularly in patients infected with HPV types 16 and 18, thought to have high oncogenic potential.     

Identification of human papillomavirus types in male urethral condylomata acuminata by in situ hybridization

An in situ hybridization technique was applied under stringent conditions to paraffin sections of urethral condylomata from male patients to determine the presence of DNA sequences of human papillomavirus (HPV) types 6, 11, 16, and 18. The material consisted of 15 classical condylomata acuminata, two flat condylomata, and five recurrent lesions. HPV DNA sequences could be identified in all 15 condylomata acuminata; in 13 lesions, two types of viral DNA were observed (types 6 and 11 in 12, types 6 and 18 in one). In the remaining two condylomata acuminata, only HPV type 11 was present. One of the two flat condylomata was negative with all the probes, and one was borderline-positive for HPV 6. Four of five recurrent lesions contained the same types of viral DNA as the primary lesions, albeit with slight differences in the intensity of viral expression. One lesion was negative with all probes. We conclude that urethral condylomata in males contain the same types of HPV as seen in other anogenital lesions of both sexes and that infection with two viral types is common. In situ hybridization with HPV DNA probes is applicable to archival material and therefore may prove to be of value in future epidemiologic studies comparing lesions in sexual partners. The determination of viral type may have therapeutic implications.     

Two newly identified human papillomavirus types (HPV 40 and 57) isolated from mucosal lesions

Two new papillomaviruses, HPV 40 and HPV 57, were isolated from a PIN lesion and an inverted papilloma of the maxillary sinus, respectively. HPV 40 showed a 13% homology to HPV 7 by reassociation kinetics and HPV 57 showed a 17% homology to HPV 2 and 25% homology to HPV 27. Hybridization of the DNA of these papillomaviruses to a wide variety of different tumor biopsies revealed that HPV 40 was present in a few genital condylomata acuminata as well as in bowenoid lesions. HPV 57 DNA was present in an oral wart, a genital condyloma acuminatum, and verrucae vulgares lesions from two immunosuppressed patients.     

Detection and typing of human papillomaviruses in mucosal and cutaneous biopsies from immunosuppressed and immunocompetent patients and patients with epidermodysplasia verruciformis: a unified diagnostic approach.

AIM: To develop a unified diagnostic approach for the detection of human papillomavirus (HPV) DNA in skin and mucosal biopsies from both immunocompetent and immunosuppressed individuals using a degenerate polymerase chain reaction (PCR) method. METHODS: The sensitivity and specificity of three published degenerate primer sets (HVP2/B5 and F14/B15; MY09/MY11; CP62/69 outer and CP65/68 nested primer pairs) were evaluated in PCR reactions with serial dilutions of 12 representative cloned HPV types. This combination of primers was then used to detect HPV DNA in 49 benign and malignant lesions of cutaneous and mucosal origin from immunosuppressed, immunocompetent, and epidermodysplasia verruciformis (EV) patients, and compared with detection rates using single primer sets alone. RESULTS: The observed sensitivity of MY09/MY11 and CP62/69 + CP65/68 was high for mucosal and EV HPV types, respectively. The sensitivity of all primer sets for cutaneous types was low, but nonetheless the use of this combination of primers allowed HPV DNA detection in all of the benign warts analysed. Several mixed infections were also identified. A high prevalence of HPV DNA was similarly detected in squamous cell carcinomas from immunocompromised patients; the HPV types found were exclusively EV related. CONCLUSIONS: The use of a combined degenerate primer PCR approach considerably improves HPV DNA detection over individual primer sets and allows detection of mixed infections. The findings may help explain the discrepancies in published reports relating to HPV DNA detection in benign and malignant skin lesions. Further modifications to this method are in progress which should significantly improve comprehensive HPV detection and typing for diagnostic purposes.     

Use of PGMY primers in L1 consensus PCR improves detection of human papillomavirus DNA in genital samples

The novel PGMY L1 consensus primer pair is more sensitive than the MY09 and MY11 primer mix for detection and typing with PCR of human papillomavirus (HPV) DNA in genital specimens. We assessed the diagnostic yield of PGMY primers for the detection and typing of HPV by comparing the results obtained with PGMY09/PGMY11 and MY09/MY11/HMB01 on 299 genital samples. Amplicons generated with PGMY primers were typed with the line blot assay (PGMY-line blot), while HPV amplicons obtained with the degenerate primer pool MY09/MY11/HMB01 were detected with type-specific radiolabeled probes in a dot blot assay (standard consensus PCR test). Cervicovaginal lavage samples (N = 272) and cervical scrape samples (N = 27) were tested in parallel with both PCR tests. The PGMY-line blot test detected the presence of HPV DNA more frequently than the standard consensus PCR assay. The concordance for HPV typing between the two assays was 84.3% (214 of 255 samples), for a good kappa value of 0.69. Of the 177 samples containing HPV DNA by at least one method, 40 samples contained at least one HPV type detected only with PGMY-line blot, whereas positivity exclusively with the standard consensus PCR test was found for only 7 samples (P < 0.001). HPV types 45 and 52 were especially more frequently detected with PGMY than MY primers. However, most HPV types were better amplified with PGMY primers, including HPV-16. Samples with discordant results between the two PCR assays more frequently contained multiple HPV types. Studies using PGMY instead of MY primers have the potential to report higher detection rates of HPV infection not only for newer HPV types but also for well-known genital types.     

HPV6，11型在女性生殖器尖锐湿疣和乳头瘤样增生中的分布

从３４２名上海地区患者的外阴、阴道和宫颈的病变部，采集４８７份活检标本。５种病理诊断类型，经斑点杂交法检测，ＨＰＶ６、１１ＤＮＡ总阳性率：尖锐湿疣８５％、乳头瘤样增生１０．１０％、慢性宫颈炎７．１４％、鳞状上皮增生５％、生殖器正常粘膜２．６５％。５１例尖锐湿疣和１０例乳头瘤样增生，ＨＰＶ６、１１和６＋１１感染阳性率分别为：１５．６９％、２５．４９％、５８．８２％和２０％、３０％、５０％。Ｓｏｕｔｈｅｒｎ印迹转移杂交与斑点杂交的符合率，在尖锐湿疣中，ＨＰＶ６、１１和６＋１１分别为７５％、９２．３％、８６．７％；在乳头瘤样增生中，分别为１００％、３３．３％、６０％。ＡＢＣ法检测阳性率仅为６１．６７％和０％。本调查结果证实，上海地区女性尖锐湿疣和乳头瘤样增生与ＨＰＶ６、１１型感染密切相关，且以ＨＰＶ两种型别（ＨＰＶ６＋１１）混合感染为主。     

Human papillomavirus DNA determination of anal condylomata, dysplasias, and squamous carcinomas with in situ hybridization

Infection with types 6, 11, 16, and 18 of the human papillomavirus (HPV) is associated with condylomatous, dysplastic, or carcinomatous changes in the genital tract. Emerging evidence suggests that a similar series of lesions develops in the anal canal after exposure to the same HPV types. In situ hybridization was performed with the use of biotinylated DNA probes to HPV 6, 11, 16, and 18, so as to determine the frequency of HPV DNA in 45 perianal and/or anal condylomata, 6 anal intraepithelial neoplasias, and 13 anal squamous cell carcinomas. Of the 33 perianal and/or anal condylomata in which HPV DNA was detected, 13 contained HPV 6 and 11, 12 HPV 6, 7 HPV 11, and 1 HPV 6, 11, and 18. Two of four severe anal dysplasias contained HPV 16, whereas one case each of mild and moderate anal dysplasia contained HPV 6. No HPV DNA was detected in the anal squamous cell carcinomas. The study demonstrated the presence of HPV DNA in 73% of condylomata and 67% of anal dysplasias. The observations suggest that the cloacogenically derived anal epithelium is susceptible to infection by the same HPV types as infect the similarly derived epithelium of the lower female genital tract and that these HPV types result in some similar lesions, i.e., condylomata and dysplasias in both sites. A role in the genesis of anal cancer was not found in this study.     

High-grade dysplasia in genital warts from two patients infected with the human immunodeficiency virus

Cancer-associated human papillomavirus (HPV) types are detected in genital warts removed from immunosuppressed individuals more commonly than from those occurring in otherwise healthy individuals. The prognosis of genital warts containing cancer-associated HPV types is not known. Because it is assumed that genital warts are benign lesions, they are usually treated by destructive therapies without prior knowledge of histopathology. The aim of the present study was to determine whether genital warts from individuals with or without human immunodeficiency virus (HIV) contain high-risk HPV types or areas of dysplasia. The study design was a nonrandomized analysis of genital warts removed by excision biopsy from 15 HIV-infected patients and 15 HIV-negative patients. The tissue was analyzed for HPV DNA by hybrid capture, and microscopic sections of each biopsy were examined for areas of dysplasia. Genital warts from HIV-infected patients contained cancer-associated (“high risk”) HPV types in 9 of 15 cases, including 1 that contained only a high-risk type. High-grade dysplastic abnormalities were present in 2 of the 15 lesions from this group, both of which contained high-risk HPV types. Four genital warts removed from HIV-negative patients contained high-risk HPV types, but none contained dysplastic abnormalities. It is concluded that genital warts from HIV-infected patients often contain high-risk HPV types. Such lesions may exhibit dysplastic changes. The frequency of dysplastic changes in genital warts from HIV-infected patients is not known. Biopsy of genital warts may be indicated prior to additional therapy in HIV-infected patients, and surgical removal should be considered as a preferred treatment option in these patients. J. Med. Virol. 54:69–73, 1998. © 1998 Wiley-Liss,Inc.     

Detection of human papillomavirus deoxyribonucleic acid in the female genital tract

A total of 336 biopsies, scrapes and exfoliated cells from the cervix and from the lower genital tract were screened for human papilloma (HP) viral sequences of types 6, 11, 16 and 18 by Southern blot, dot blot and filter in situ (FISH) hybridizations with cloned ³²P-radiolabeled HPV DNA probes. The specimens included cervical intraepithelial neoplasias (CIN I–III), carcinoma in situ and invasive carcinoma of the cervix and vagina, adenocarcinomas, vulvar and vaginal condylomata acuminata and healthy epithelial samples. The oncogenic HPV 16 was found in 46% of the cervical carcinomas. Most of the type 16 occurences (75%) represented the third stage of inooperable cases. Similarly, HPV 18 was also most frequently present in this stage as well as in carcinoma in situ and in CIN III (25%, 18%). At the same time, in condylomata acuminata, types 6 and 11 were detectable in 88.7% of cares. In all, 13.5% of the normal samples harboured HPV DNA.     

Analysis of benign and malignant urogenital tumors for human papillomavirus infection by labelling cellular DNA

A total of 268 biopsies from the genital region was screened for the presence of human papillomavirus DNA. The specimens included carcinoma of the vulva, vagina, cervix, corpus uteri, ovaries and penis, and Bowen's carcinomas, Bowenoid papuloses, Bowen's disease, cervical intraepithelial neoplasias (CIN I to III), Buschke-Löwenstein tumors, a cervical polyp, decidua, endometrium and histologically normal biopsies. Of 45 carcinomas, 18 contained either HPV 16 and/or 18 and 3 HPV 6-related sequences.In a few individual biopsies double or even triple infections were noted. Unusual was the presence of HPV 2-related DNA in one biopsy from Bowen's disease, whereas 2 condylomata acuminata contained HPV 3-related DNA and one contained HPV DNA related to a group of epidermal HPV's found in epidermodysplasia verruciformis lesions.     

Human papillomavirus detection in urine samples from male patients by the polymerase chain reaction.

Human papillomavirus (HPV) detection was performed using the polymerase chain reaction technique on urine samples from 17 male patients with condylomata acuminata in the meatus urethrae. Urine samples from 14 male laboratory volunteers were analyzed as controls. The DNA was extracted and purified from urine sediments, centrifuged at 1,800 and 100,000 x g, and subjected to 40 cycles of amplification with HPV 6 and HPV 11 type-specific anticontamination primers and the heat-stable Taq DNA polymerase. HPV was detected in the urine of 15 (88%) patients. In all positive patients the urine sediments of both the 1,800 and 100,000 x g centrifugation steps contained HPV DNA. Eight patients were found to be positive for HPV 6 DNA, six were positive for HPV 11 DNA, and one was positive for both HPV 6 and HPV 11 DNA. None of the males in the control group was positive for either HPV 6 or HPV 11 DNA. The results demonstrate that HPV can be transported by the urine, probably in exfoliated HPV-infected cells. A similar mechanism may occur during ejaculation, allowing sexual transmission of HPV viruses harbored in the cells of the male genital tract.     

Loop-mediated isothermal amplification method for detection of human papillomavirus type 6, 11, 16, and 18

A new method was developed for detection of human papillomavirus (HPV) by loop-mediated isothermal amplification (LAMP), which was compared with the polymerase chain reaction (PCR), and real-time PCR for specificity and sensitivity. All initial validation studies with the control DNA proved to be type-specific. In order to evaluate the reliability of HPV type-specific LAMP detecting HPV DNA from clinical samples, tissue specimens were obtained from 27 patients with external genital polypoid lesions. The histologic diagnoses included condyloma acuminatum (n = 21), bowenoid papulosis (n = 2), seborrheic keratosis (n = 2), epidermolytic acanthoma (n = 1), and hairy nymphae (n = 1). HPV-6 DNA and HPV-11 DNA were detected in 18 and 3 of 21 condylomata acuminata, respectively, and there was no simultaneous infection. HPV-16 DNA was detected in one of two bowenoid papuloses. HPV DNA was not detected in the seborrheic keratoses, epidermolytic acanthoma, and hairy nymphae. These results correlated perfectly with those from real-time PCR analysis. Most positive samples contained high copy numbers of HPV DNA. HPV-11 DNA was detected in one case that could not be detected by PCR. The average reaction time was about 59 min. There was a linear correlation between the genome quantity and reaction time to reach the threshold. The LAMP method has an additional advantage as a quantitative method, and is superior in terms of sensitivity, specificity, rapidity, and simplicity, and can potentially be a valuable tool for the detection of HPV DNA.     

Analyses of human papillomavirus genotypes and viral loads in anogenital warts

Condylomata acuminata (genital warts) are the most common sexually transmitted viral diseases. These lesions are caused by infection with mucosal human papillomaviruses (HPVs). However, there is limited information on HPV strain distribution involved in the molecular pathogenesis of these lesions. To address this, the strain prevalence and the frequency of multiple HPV infections were determined in wart tissue obtained from 31 patients attending a wart clinic. These lesions were bisected and subjected to parallel DNA and mRNA extractions. HPV-type prevalence and incidence of multiple infections were determined by the Roche Linear Array assay. qPCR compared HPV 6, 11, 16, and 18 viral loads and RT-qPCR measured HPV 6 and 11 E6 genomic expression levels. Seventy-one percent of these samples were infected with multiple HPVs. Only one sample was negative for HPV 6 or 11 DNA. Forty-eight percent of samples were positive for a high risk (oncogenic) HPV. The results show that multiple infections in tissue are frequent and the subsequent analysis of HPV 6 and 11 E6 DNA viral loads suggested that other HPVs could be causing lesions. Further analysis of HPV 6/11 E6 mRNA levels showed that there was no discernable relationship between HPV 6 E6 DNA viral load and relative HPV 6 or 11 E6 mRNA levels thereby questioning the relevance of viral load to lesion causality.     

Detection of human papillomavirus DNA in genital lesions by using a modified commercially available in situ hybridization assay.

A modified, commercially available DNA-DNA in situ hybridization test that uses biotinylated probes for the identification of human papillomavirus (HPV) DNA types 6/11, 16/18, and 31/33/35 was evaluated. HPV DNA was detected in 314 of 787 (40%) histologically abnormal genital biopsy specimens by using the ViraType in situ assay (Life Technologies, Gaithersburg, Md.), in which the hybridization time was increased from 2 to 16 h. Ninety percent of positive condyloma acuminata specimens contained HPV type 6/11 DNA. The prevalences of HPV DNA for cervical intraepithelial neoplasia I, II, and III lesions by this in situ hybridization test were 42, 54, and 55%, respectively. The combined prevalence of HPV type 16/18 and 31/33/35 DNAs increased with the severity of the lesion, while the prevalence of type 6/11 DNA decreased. HPV type 6/11 DNA was found only in 1 of 16 (6%) positive cervical intraepithelial neoplasia III specimens. HPV type 16/18 and 31/33/35 DNA was detected in 11 of 16 (69%) and 4 of 16 (25%) in situ hybridization-positive cervical intraepithelial neoplasia III specimens, respectively. Thus, the observation that certain "higher-risk" HPV genotypes are associated with upper-grade cervical precancer lesions was confirmed by this commercial hybridization system. In general, the assay was found to be well suited for use in the clinical laboratory. The ViraType in situ procedure modified for a longer hybridization time may be helpful in identifying lesions containing higher-risk HPV strains.     

High prevalence of papillomavirus-associated penile intraepithelial neoplasia in sexual partners of women with cervical intraepithelial neoplasia

To determine whether neoplastic cervical lesions in women are associated with papillomavirus infections in their sexual partners, we used a colposcope to examine male sexual partners of women with cervical flat condyloma (294 cases) or cervical intraepithelial neoplasia (186 cases), before and after 5 percent acetic acid was applied to the penis and the anogenital area. Condylomata acuminata, papules, and macules were observed in 309 of the 480 men (64.4 percent). In 204 of them (42.5 percent), macules or slightly elevated papules were detected only after application of acetic acid. Condylomata acuminata or lesions showing histologic features of condyloma were found in 121 partners (41.2 percent) of women with condyloma, but in only 10 partners (5.4 percent) of women with cervical intraepithelial neoplasia. Penile lesions showing histologic features of intraepithelial neoplasia were found in 61 partners (32.8 percent) of women with cervical intraepithelial neoplasia, but in only 4 partners (1.4 percent) of women with flat condyloma. Thirty-six (60 percent) of the 60 macules or papules tested contained papillomavirus DNA sequences. Human papillomavirus types 16 and 33 were almost exclusively found in penile intraepithelial neoplasia. Type 6, type 11, and the recently recognized type 42 were found in lesions showing features of condyloma or minimal histologic changes. As yet uncharacterized papillomaviruses were found in 15 percent of the specimens. These data support the concept that cervical carcinomas and precancerous lesions in women may be associated with genital papillomavirus infection in their male sexual partners.     

Comparison of peroxidase-labeled DNA probes with radioactive RNA probes for detection of human papillomaviruses by in situ hybridization in paraffin sections.

A study comparing in situ hybridization using nonradioactive DNA probes directly conjugated with horseradish peroxidase (HRP), and 35S-labeled antisense RNA probes for human papillomavirus (HPV) types 6/11, 16, and 18 was performed on formalin-fixed, paraffin-embedded tissue from 34 lesions of the cervix and vulva. These lesions included exophytic condylomas and intraepithelial and invasive neoplasms. HPV 6/11 was detected in two of four condylomata acuminata by both in situ techniques. HPV 16 was detected in 13 of 30 cases of intraepithelial and invasive neoplasms by both methods. Discordance between the two methods occurred in two instances. The radiolabeled probe but not the HRP probe detected HPV 16 in one case of cervical intraepithelial neoplasia (CIN 3), whereas the converse occurred in one case of vulvar intraepithelial neoplasia (VIN 3). HPV 18 was not detected in any of the specimens by either method. This study demonstrates that nonradioactive HRP-labeled probes for the detection of specific HPV types are as sensitive as the more laborious and potentially hazardous radioactive probes.     

Human papillomavirus segregation patterns in genital and nongenital warts in prepubertal children and adults.

This study compared the segregation patterns of human papillomavirus (HPV) in genital and nongenital warts in prepubertal children and adults. HPV 2 was detected in most nongenital warts in children and adults, whereas neither HPV 6 or 11 was detected at nongenital sites in either group with the use of in situ or Southern blot hybridization analyses. Of nine genital tract lesions in children. HPV 2 was detected in two and HPV 6 or 11 in six. More than 90% of cases of regional tract condylomata in adults contained HPV 6 or 11. HPV 2 was not detected in any of 99 genital tract lesions in adults. It is concluded that HPV 6/11 cannot proliferate at nongenital cutaneous sites and HPV 2 can proliferate in the genital tract of children but not adults. Thus, the detection of HPV 6 or 11 in a genital wart in a child implies, assuming cutaneous transmission, infection from a genital site, whereas the detection of HPV 2 presumes nongenital transmission.     

Human papillomavirus (HPV) type distribution and serological response to HPV type 6 virus-like particles in patients with genital warts.

Thirty-nine patients with condylomas (12 women and 27 men) attending a dermatology clinic were tested for genital human papillomavirus (HPV) DNA and for seroprevalence to HPV type 6 (HPV6) L1 virus-like particles. The L1 consensus PCR system (with primers MY09 and MY11) was used to determine the presence and types of HPV in sample specimens. All 37 (100%) patients with sufficient DNA specimens were positive for HPV DNA, and 35 (94%) had HPV6 DNA detected at the wart site. Three patients (8%) had HPV11 detected at the wart site, and one patient had both HPV6 and -11 detected at the wart site. Thirteen additional HPV types were detected among the patients; the most frequent were HPV54 (8%) and HPV58 (8%). Baculovirus-expressed HPV6 L1 virus-like particles were used in enzyme-linked immunosorbent assays to determine seroprevalence among the patients with warts. Seronegativity was defined by a control group of 21 women who were consistently PCR negative for HPV DNA. Seroprevalence was also determined for reference groups that included cytologically normal women who had detectable DNA from either HPV6 or HPV16 and women with HPV16-associated cervical intraepithelial neoplasia. Among the asymptomatic women with HPV6, only 2 of 9 (22%) were seropositive, compared with 12 of 12 (100%) female patients with warts. A similar trend in increased HPV6 seropositivity with increased grade of disease was found with the HPV16 DNA-positive women, whose seroprevalence increased from 1 in 11 (9%) in cytologically normal women to 6 in 15 (40%) among women with cervical intraepithelial neoplasia 1 or 3. However, only 4 of 25 (16%) male patients were seropositive.(ABSTRACT TRUNCATED AT 250 WORDS)     

Detection of human papillomavirus DNA in cervical lavage specimens by a nonisotopic consensus PCR assay.

A gene amplification method that combines PCR with an enzyme immunoassay (PCR-EIA) for quantitation of amplified DNA was developed for the detection of human papillomavirus (HPV). Samples were amplified with consensus primers MY09 and MY11. Amplified DNA products were reacted in solution with type-specific nested RNA probes labelled with digoxigenin-11-UTP. Hybrids were captured on a microtiter plate coated with an antidigoxigenin antibody. Bound DNA-RNA hybrids were quantitated by the addition of an alkaline phosphatase-labelled monoclonal antibody directed against DNA-RNA hybrids and a fluorogenic substrate. The detection limit of PCR-EIA was six copies of HPV type 18 DNA in the original specimen. The assay was used to assess HPV infection of the uterine cervixes of 65 women referred to a colposcopy clinic. In 66 cervicovaginal lavage specimens, all 23 HPV strains detected by a standard isotopic PCR assay were also detected by the PCR-EIA (sensitivity, 100%; 95% confidence interval, 85.2 to 100%). Forty-two of the 43 samples that did not contain HPV types 6/11, 16, 18, 31, 33, 35, and 45 were also negative by PCR-EIA, for a specificity of 97.7%. Low-level cross-reactivity was encountered between HPV types 18 and 45 as well as between types 33 and 58. PCR-EIA provides a convenient means of objectively measuring PCR-amplified HPV DNA from common genital HPV types.