Validation of a Low-Cost Human Papillomavirus Genotyping Assay Based on PGMY PCR and Reverse Blotting Hybridization with Reusable Membranes.

Détails

ID Serval
serval:BIB_B427B34E3E9E
Type
Article: article d'un périodique ou d'un magazine.
Collection
Publications
Titre
Validation of a Low-Cost Human Papillomavirus Genotyping Assay Based on PGMY PCR and Reverse Blotting Hybridization with Reusable Membranes.
Périodique
Journal of Clinical Microbiology
Auteur(s)
Estrade C., Menoud P.A., Nardelli-Haefliger D., Sahli R.
ISSN
1098-660X (Electronic)
ISSN-L
0095-1137
Statut éditorial
Publié
Date de publication
2011
Volume
49
Numéro
10
Pages
3474-3481
Langue
anglais
Résumé
The genotyping of human papillomaviruses (HPV) is essential for the surveillance of HPV vaccines. We describe and validate a low-cost PGMY-based PCR assay (PGMY-CHUV) for the genotyping of 31 HPV by reverse blotting hybridization (RBH). Genotype-specific detection limits were 50 to 500 genome equivalents per reaction. RBH was 100% specific and 98.61% sensitive using DNA sequencing as the gold standard (n = 1,024 samples). PGMY-CHUV was compared to the validated and commercially available linear array (Roche) on 200 samples. Both assays identified the same positive (n = 182) and negative samples (n = 18). Seventy-six percent of the positives were fully concordant after restricting the comparison to the 28 genotypes shared by both assays. At the genotypic level, agreement was 83% (285/344 genotype-sample combinations; κ of 0.987 for single infections and 0.853 for multiple infections). Fifty-seven of the 59 discordant cases were associated with multiple infections and with the weakest genotypes within each sample (P < 0.0001). PGMY-CHUV was significantly more sensitive for HPV56 (P = 0.0026) and could unambiguously identify HPV52 in mixed infections. PGMY-CHUV was reproducible on repeat testing (n = 275 samples; 392 genotype-sample combinations; κ of 0.933) involving different reagents lots and different technicians. Discordant results (n = 47) were significantly associated with the weakest genotypes in samples with multiple infections (P < 0.0001). Successful participation in proficiency testing also supported the robustness of this assay. The PGMY-CHUV reagent costs were estimated at $2.40 per sample using the least expensive yet proficient genotyping algorithm that also included quality control. This assay may be used in low-resource laboratories that have sufficient manpower and PCR expertise.
Pubmed
Web of science
Open Access
Oui
Création de la notice
03/11/2011 10:21
Dernière modification de la notice
09/05/2019 0:02
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