Comparison of five different polymerase chain reaction methods for detection of human papillomavirus in cervical cell specimens.
The polymerase chain reaction (PCR) methods enable the detection of large number of human papillomavirus (HPV) genotypes that infect the anogenital tract. In this study, two groups of cervical scrapes with abnormal cytomorphology were analysed. The first group was tested with three sets of consensus primers located within the L1 region of HPV genome, MY09/MY11 (i.e. MY), L1C1/L1C2-1/L1C2-2 (i.e. LC) and pI-1/pI-2 (i.e. pI) primer sets, while the second group of samples, which were all negative with the MY primers, was tested further with the LC primers, as well as with the GP5/GP6 (i.e. GP) primers. The GP primers were used in the nested PCR following amplification with the MY primers (i.e. MY/GP nested PCR). Samples from both groups were also tested with type-specific primers for HPV types 6/11, 16, 18, 31 and 33. In the first study group (N=164) there were 76.2% positive results obtained with at least one set of consensus primers. There were 62.2, 39, 62.2 and 59.1% positive results obtained with the MY, the pI, the LC and the HPV type-specific primer sets, respectively. The best results were obtained when both the MY and the LC primer sets were used, because in combination they detected 75% positive samples compared to 62.2% when used alone. There were 2. 4% samples negative with all consensus primers, but positive with one of the HPV type-specific primers, which increased the overall positivity rate to 78.6%. In the second study group (N=250) there were 8.4, 38.8 and 4% samples positive with the LC primers, the nested MY/GP and the HPV type-specific primer sets, respectively. Thus, the use of the MY/GP nested PCR increased significantly the positivity rate of HPV DNA detection and should be used for samples with a low copy number of HPV DNA. In conclusion, the following diagnostic protocol would be appropriate for detection of cancer-related HPVs: preselection of samples with the MY and the LC primers, additional amplification of the MY- and the LC-negative samples with the MY/GP nested PCR and HPV typing of consensus PCR-positive samples with the HPV type-specific primers.
Husnjak
K
K
Division of Molecular Medicine, Ruder Bosković' Institute, Bijenicka54, 10000, Zagreb, Croatia. husnjak@rudjer.irb.hr
Grce
M
M
Magdić
L
L
Pavelić
K
K
D000293:Adolescent; D000328:Adult; D000368:Aged; D017931:DNA Primers; D004279:DNA, Viral / Q000032:analysis; D005260:Female; D006801:Humans; D008875:Middle Aged; D027383:Papillomaviridae / Q000302:isolation & purification*; D030361:Papillomavirus Infections / Q000175:diagnosis*; D016133:Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity; D014626:Vaginal Smears
D017931:DNA Primers; D004279:DNA, Viral