A type-specific nested PCR assay established and applied for investigation of HB

StephenW

A type-specific nested PCR assay established and applied for investigation of HBV genotype and subgenotype in Chinese patients with chronic HBV infection
Jing-Jing Nie1 Email: [email protected]
Kui-Xia Sun1 Email: [email protected]
Jie Li1* * Corresponding author Email: [email protected]
Jie Wang1 Email: [email protected]
Hui Jin1 Email: [email protected]
Ling Wang1 Email: [email protected]
Feng-Min Lu1 Email: [email protected]
Tong Li1 Email: [email protected]
Ling Yan1 Email: [email protected]
Jing-Xian Yang1 Email: [email protected]
Mi-Shu Sun1 Email: [email protected]
Hui Zhuang1* * Corresponding author Email: [email protected]
1 Department of Microbiology, School of Basic Medical Sciences, Peking
University Health Science Center, Beijing 100191, China
Abstract
Background
Many studies have suggested that hepatitis B virus (HBV) genotypes show not only
geographical distribution and race specificity, but also are associated with disease progression and response to interferon treatment. The objective of this study was to develop a nested polymerase chain reaction (nPCR) assay for genotypes A-D and subgenotypes B1, B2, C1 and C2 of hepatitis B virus (HBV) and to investigate the distribution characteristics of HBV genotypes/subgenotype in China.
Methods
After redesigning the primers and optimizing the reaction conditions using common Taq
polymerase, the sensitivity, specificity and reproducibility of the method were evaluated
using plasmids and serum samples. In total, 642 serum samples from patients with chronic
HBV infection were applied to investigate the distribution of HBV genotype and subgenotype
in China.
Results
The genotype and subgenotype could be identified when the HBV DNA load of a sample was ≥10**2.3 IU/mL. For the 639 successfully genotyped samples, the sequencing results of 130 randomly selected samples (20.3%, 130/639) were consistent with those of the nPCR method.
The present study showed that HBV genotype B (11.2%, 72/642), C (68.2%, 438/642) and D (7.2%, 46/642) were circulating in China, while genotype C was the dominant strain except for western region where genotype D was the prevalent strain. The main subgenotypes of genotypes B and C were B2 (87.5%, 63/72) and C2 (92.9%, 407/438), respectively.
Conclusions
The low-cost nPCR method would be a useful tool for clinical and epidemiological
investigation in the regions where genotypes A-D are predominant.

StephenW

背景
许多研究表明，乙型肝炎病毒（HBV）基因型不仅显示与疾病进展的地理分布和种族特异性，但也有关联 和干扰素治疗的反应。本研究的目的是建立一个嵌套聚合酶链反应（nPCR）检测基因型AD和亚型B1，B2，C1的和C2 B型肝炎病毒（HBV）和调查乙肝病毒的分布特征
在中国的基因型/亚型。
方法
经过重新设计的引物和优化使用普通Taq酶的反应条件聚合酶，其敏感性，特异性和重复性的方法进行评估使用质粒和血清标本。总共642慢性乙型肝炎患者的血清标本被应用于乙型肝炎病毒感染乙型肝炎病毒基因型和亚型的分布调查在中国。
结果
基因型和亚型HBV DNA载量的样品时，可确定≥10**2.3 IU/毫升。 639样本的成功基因，测序结果130随机抽取的样品（20.3％，130/639）人与NPCR方法一致。
目前的研究表明，HBV B基因型（11.2％，72/642），C（68.2％，438/642）和D
（7.2％，46/642）在中国的循环，而C基因型是优势菌种，除了为西部地区，其中D型流行株。的主要亚型基因型B和C分别为B2（占87.5％，63/72）和C2（92.9％，407/438），分别。
结论
低成本nPCR的方法将是一个有用的工具，临床和流行病学调查基因型公元为主的地区。

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