Hepatitis C virus (HCV) genotyping by annealing reverse transcription-PCR products with genotype-specific capture probes

Abstract

The genotype of the hepatitis C virus (HCV) strain infecting a given patient is an important predictive factor for the clinical outcome of chronic liver disease and its response to anti-viral therapeutic agents. We herein sought to develop a new easy, sensitive and accurate HCV genotyping method using annealing genotype-specific capture probes (AGSCP) in an automation-friendly 96-well plate format. The validation of our new AGSCP was performed using the Standard HCV Genotype Panel. We then used both our AGSCP and the commercially available INNO-LiPA assay to analyze the HCV genotypes from 111 Korean patients. Discordant results were analyzed by direct sequencing. AGSCP successfully genotyped the standard panel. The genotypes of 111 patient samples were also obtained successfully by AGSCP and INNO-LiPA. We observed a high concordance rate (93 matched samples, 83.8%) between the two assays. Sequencing analysis of the 18 discordant results revealed that the AGSCP had correctly identified 12 samples, whereas the INNO-LiPA had correctly identified only 6. These results collectively indicate that AGSCP assay is a convenient and sensitive method for large-scale genotyping, and it may be a promising tool for the determination of HCV and other genotypes in clinical settings.

A method for rapid detection and genotype identification of hepatitis C virus 1–6 by one-step reverse transcription loop-mediated isothermal amplification

“…While 31 these tests are highly sensitive, they remain expensive and 32 laborious and require well-trained personnel, as well as sophisti-33 cated laboratory facilities. [14][15][16][17] Furthermore, the application of 34 these tests may be limited regarding their ability to detect and 35 simultaneously identify the specific HCV genotypes. 18 Besides, 36 several (reverse transcription) loop-mediated isothermal amplifi-37 cation (RT)-LAMP assays have been designed for the detection of 38 various pathogens including HCV, but they detect a limited 39 number of genotypes and rarely demonstrate genotype identifica-40 tion of the pathogens targeted for detection.…”

Determination of hepatitis C virus genotype distribution in Adıyaman province

“…Bu hatalar sonucu, kronik hepatit C'li hastalarda HCV' nin farklı genetik sekansları ile heterojen bir topluluk oluşmaktadır. Günümüze kadar yapılan çalışmalarda HCV'nin en az 6 genotipi ve 80' den fazla alt tipi vardır [6,7]. Bunlardan genotip 1a, 1b,2a, 2b, 3a tüm dünyada yaygın olarak görülürken, genotip 4, 5, 6 sadece belli bölgelerde bulunmaktadır [8].…”

Determination of Hepatitis C Virus Genotype Distribution in Mersin Province, Turkey

“…HCV enfeksiyonunun seroprevalansı Kuzey Amerika'da yaklaşık %1, Akdeniz ve Asya ülkelerinde %3-4, Orta Afrika ve Mısır'da %10-20 olarak bildirilmiş-tir 2 . Akut hepatit C enfeksiyonlarının %70-80 oranında kronikleşmesi ve çoğunlukla asemptomatik seyirli olması en önemli problemdir 1. Flaviviridae ailesinin bir üyesi olan HCV, pozitif polariteli, tek zincirli 9.6 kilobaz (kb) büyüklüğünde RNA genomuna sahiptir 3 . Viral genom, yüksek mutasyon oranı ve yüksek derecede genomik heterojenite göstermekte olup, bu da tedavi etkinliği için büyük öne-me sahiptir 4 .…”