Algorithmic Approach to High-Throughput Molecular Screening for Alpha Interferon-Resistant Genotypes in Hepatitis C Patients
| Autor: | Srinand Sreevatsan, Lance Fors, Monika de Arruda, Raj R. Barathur, Raveendran Pottathil, Mridula R. Pottathil, David J. Marshall, Jack Bain Bookout, Fidel M. Ringpis, Christopher Murvine |
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| Rok vydání: | 1998 |
| Předmět: |
Microbiology (medical)
Genotype Hepatitis C virus Alpha interferon Hepacivirus Biology medicine.disease_cause Sensitivity and Specificity Substrate Specificity law.invention law Virology medicine Humans Genotyping Polymerase chain reaction Genetics Polymorphism Genetic Interferon-alpha Drug Resistance Microbial Sequence Analysis DNA Amplicon Endonucleases DNA Fingerprinting Hepatitis C Genotype frequency Genetic Techniques DNA profiling Costs and Cost Analysis Algorithms |
| Zdroj: | Journal of Clinical Microbiology. 36:1895-1901 |
| ISSN: | 1098-660X 0095-1137 |
| DOI: | 10.1128/jcm.36.7.1895-1901.1998 |
| Popis: | This study was designed to analyze the feasibility and validity of using Cleavase Fragment Length Polymorphism (CFLP) analysis as an alternative to DNA sequencing for high-throughput screening of hepatitis C virus (HCV) genotypes in a high-volume molecular pathology laboratory setting. By using a 244-bp amplicon from the 5′ untranslated region of the HCV genome, 61 clinical samples received for HCV reverse transcription-PCR (RT-PCR) were genotyped by this method. The genotype frequencies assigned by the CFLP method were 44.3% for type 1a, 26.2% for 1b, 13.1% for type 2b, and 5% type 3a. The results obtained by nucleotide sequence analysis provided 100% concordance with those obtained by CFLP analysis at the major genotype level, with resolvable differences as to subtype designations for five samples. CFLP analysis-derived HCV genotype frequencies also concurred with the national estimates (N. N. Zein et al., Ann. Intern. Med. 125:634–639, 1996). Reanalysis of 42 of these samples in parallel in a different research laboratory reproduced the CFLP fingerprints for 100% of the samples. Similarly, the major subtype designations for 19 samples subjected to different incubation temperature-time conditions were also 100% reproducible. Comparative cost analysis for genotyping of HCV by line probe assay, CFLP analysis, and automated DNA sequencing indicated that the average cost per amplicon was lowest for CFLP analysis, at $20 (direct costs). On the basis of these findings we propose that CFLP analysis is a robust, sensitive, specific, and an economical method for large-scale screening of HCV-infected patients for alpha interferon-resistant HCV genotypes. The paper describes an algorithm that uses as a reflex test the RT-PCR-based qualitative screening of samples for HCV detection and also addresses genotypes that are ambiguous. |
| Databáze: | OpenAIRE |
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