The Molecular Epidemiology Of HCV Genotype Among High Risk Populations In China

BackgroundThe global prevalence of HCV infection is about2percent, and it is around0.43percent in China. Chronic HCV infections can lead to chronic inflammation, necrosis of liver and liver fibrosis,2%～4%HCV patients will progress into liver cirrhosis and hepatocellular carcinoma(HCC) eventually. This fact attracts increasing attention from both society and public health. Some researches have shown:the risk of progressing into liver cirrhosis and HCC declines, when patients get SVR after the combine treatment of interferon and ribavirin; however, the sensitivity and prognostic of treatment are different among HCV genotypes. Therefore, information about infections with specific HCV genotype is necessary for choosing the optimal treatment and predicting prognostic, In China, DUS is the most high-risk population, the prevalence of HCV infections among people with transfusion history, patients with dialysis treatment, STD and FSW is lower than DUS. Therefore, it is important to know about the distribution of HCV genotypes among high-risk populations, and estimate public health investment and burden of diseased caused by HCV infection.AimsTo attain information about the distribution of HCV genotypes among HCV infections of DUS and sentinel surveillance populations, and HCV patients from hospital; Study on influence factors of the distribution of HCV genotypes; analysis on risk factors of intravenous drug use; discuss the relationship between HCV antibody positive results and NAT positive results; debate on the relationship between baseline viral load and HCV genotype.Methods1. Design proper primer for the molecular epidemiology study of HCV genotypes in China according to the mainly genotypes in HCV database; 2. Screen samples with domestic HCV ELISA reagents, if S/CO of ELISA result is over0.8, then recheck with imported HCV ELISA reagents, in order to find HCV antibody positive samples;3. Extract RNA by MagNA Pure LC-an automatic nucleic acids extraction machine produced by Roche, then RT-PCR of5’-UTR and gel electrophoresis of products for NAT testing;4. RT-PCR NS5B of RNA extractions with NAT positive results, and send positive products of NS5B to the company providing sequencing service;5. Clean, assemble and cluster of multiple sequences with the use of Chromas Pro1.5, BioEdit and Mega4.0;6. Contrast the amplification rate among designed one, primers recommended by HCV Database and "Guide of HCV testing technology";7. Compare the agreement of detecting HCV genotypes by In-house and Abbott HCV Ⅱ Genotype;8. Statistic methods adopted in this research include t test, chisq test, logistic regression and Anova analysis with the help of SAS9.1.Result1. The total number of HCV antibody positive samples was1664from HCV infected DUS, HCV patients in hospital and sentinel surveillance.1416of1664HCV antibody positive samples were NAT(+),1302of1416NAT(+) samples can be classified into subtype level. The most frequent subtypes of HCV infected DUS were3a,3b and6a; Among HCV patients in hospital and sentinel surveillance, the most frequent ones were lb and2a;2. Gender, age, district and population were influence factors of HCV genotype’s distribution, but the influence factors were different in population and districts;3. Male, young age and marry were risk factors of intravenous drug use;4. There was no statistical difference of the distribution of baseline viral load in different HCV genotypes infections;5. Designed primer was suitable for the molecular epidemiology of HCV genotype in China, where the main genotypes included la,lb,2a,3a,3b,6a,and6n; primer from HCV Database can be used to amplify HCV1,2and3genotypes; and primer from "Guide of HCV testing technology" is suit for the amplification of HCV1and2genotypes;6. The results of18sample (1,2,3and6genotype) from In-house and Abbott assays were coincidence. However, one1a sample by In-house was classified into1a/2co-infection, three6n samples could not be determined by Abbott way;7. There was no statistical difference of the distribution of ELISA S/CO between NAT(-) and NAT(+) samples.Conclusion1. The main genotypes of HCV infections of DUS were3and6, but the ones of HCV infections of Hebei sentinel surveillance population and HCV patients of hospital were1and2;2. Gender, age, district and population were influence factors of HCV genotype’s distribution, but the influence factors were different in population and districts;3. Male, young age and marry were risk factors of intravenous drug use;4. There was no statistical difference of the distribution of baseline viral load in different HCV genotypes infections;5. Designed primer could effectively amplify the main genotypes(1a,1b,2a,3a,3b,6a,6n) in China;6. The agreement percentage of In-house and Abbott Real time HCV Genotype II to detect HCV genotypes was satisfy;7. S/CO of ELISA cannot predict the results of NAT.