Diagnostic sensitivity of polymerase chain reaction and Southern blot hybridization for the detection of human papillomavirus DNA in biopsy specimens from cervical lesions.
Human papillomaviruses (HPVs) are associated with benign and malignant neoplasms of the cervix. One of the criteria for their etiologic role requires an assessment of whether virtually all or only a small fraction of lesions contain viral genomes. DNA preparations from colposcopically directed punch biopsies of cervical lesions were analyzed by Southern blot hybridization and the polymerase chain reaction (PCR) for the presence of HPV DNA. The biopsy specimens represented different pathologic entities (koilocytosis, condyloma, cervical intraepithelial neoplasia, and invasive carcinoma). In Southern blot hybridization with radioactive probes for HPV 11, 16, 18, 31, and 33, HPV DNA was detected in 74% of the biopsy specimens (42 of 57 cases), with the predominant types being HPV 16 and HPV 18. In contrast, after PCR amplification with primers yielding fragments of characteristic size for HPV 11, 16, and 18, the analysis of the same 57 biopsy specimens revealed that all samples were positive for at least one HPV type. To exclude false-positive PCR results, controls without HPV DNA were interspersed at regular intervals, and results were evaluated only if these controls remained HPV negative. To exclude false-negative results due to failure of the reaction, a target sequence within the c-Ha-ras-1 gene was used as an internal control. All HPV typing results obtained by Southern blot hybridization were in agreement with HPV typing by PCR. The higher number of positive samples in the latter analysis stems from the increased sensitivity of PCR, which was which was effective in identifying as few as 10-100 HPV DNA molecules; in contrast, the sensitivity of Southern blot hybridization was 1 pg, or approximately 10(5) molecules of HPV DNA. The authors conclude that, with sufficiently sensitive diagnostic methods, HPV DNA can be detected in most, if not all, neoplastic cervical lesions.
Tham
K M
KM
Department of Obstetrics and Gynecology, National University of Singapore, Kent Ridge.
Chow
V T
VT
Singh
P
P
Tock
E P
EP
Ching
K C
KC
Lim-Tan
S K
SK
Sng
I T
IT
Bernard
H U
HU
D001483:Base Sequence; D001706:Biopsy; D015139:Blotting, Southern; D002584:Cervix Uteri / Q000737:chemistry / Q000382:microbiology / Q000473:pathology; D004279:DNA, Viral / Q000032:analysis / Q000235:genetics*; D005260:Female; D006801:Humans; D008969:Molecular Sequence Data; D027383:Papillomaviridae / Q000235:genetics* / Q000302:isolation & purification; D016133:Polymerase Chain Reaction; D012846:Singapore / Q000453:epidemiology; D002577:Uterine Cervical Diseases / Q000453:epidemiology / Q000235:genetics* / Q000473:pathology; D002583:Uterine Cervical Neoplasms / Q000453:epidemiology / Q000235:genetics* / Q000473:pathology
D004279:DNA, Viral