Head‐to‐head comparison of genotyping of human papillomavirus by real‐time multiplex PCR assay using type‐specific primers and SPF10‐PCR‐based line probe assay

Abstract The SPF10‐polymerase chain reaction (PCR)‐based line probe assay (LiPA‐25) with high analytical sensitivity and specificity for human papillomavirus (HPV) genotyping in clinical samples has been widely used in vaccine and epidemiologic studies. A real‐time multiplex PCR assay using type‐specific primers (Hybribio‐23) with low workload and cost has been developed recently. The study aimed to compare the performance of LiPA‐25 and Hybribio‐23 in selected 1731 cervical swab and 117 tissue samples, with a focus on 20 common HPV types (14 high‐risk: 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, and 68/73; 6 low‐risk: 6, 11, 42, 43, 44, and 53). The level of agreement of two assays was determined using Cohen's Kappa ( κ ) statistics. A total of 1296 (74.9%) swab samples were identified as HPV‐positive by Hybribio‐23 or LiPA‐25, of which 814 (62.8%) samples exhibited concordant, 358 (27.6%) showed additional or fewer types (compatible), and 124 (9.6%) were discordant. In addition, the two assays showed a perfect agreement for 20 HPV‐combined detection ( κ = 0.838) and 17 individual HPV types (all κ > 0.800), a good agreement for HPV31 ( κ = 0.792) and 43 ( κ = 0.696), and a moderate agreement for HPV42 ( κ = 0.504). Hybribio‐23 was significantly more sensitive for HPV58, 59, 68/73, 42, 43, and 44, and less sensitive for HPV35 and 66 than LiPA‐25 (McNemar's test: all p < 0.05). For 117 HPV‐positive tissue specimens, the identification of genotypes was 85.2% identical, 12.2% compatible, and only 2.6% discordant. The agreement for HPV31 ( κ = 0.786), 68/73 ( κ = 0.742), and HPV53 ( κ = 0.742) was good, while for other types (all κ > 0.853) and 20 HPV‐combined detection ( κ = 0.936) was perfect (all p > 0.05). In conclusion, Hybribio‐23 and LiPA‐25 are comparable. Hybribio‐23 could be used for the detection and genotyping of HPV in cervical samples for epidemiological and vaccine studies worldwide.