逃脱免疫和耐药的乙肝表面抗原（HBsAg ）

StephenW

Decreased antigenicity profiles of immune-escaped and drug-resistant hepatitis B surface antigen (HBsAg) double mutants

Mingshun Zhang12†, Guohong Ge3†, Yonglin Yang4, Xubing Cai4, Qiang Fu4, Jie Cai4* and Zuhu Huang1*   

    * Corresponding authors: Jie Cai [email protected] - Zuhu Huang [email protected]
    † Equal contributors

Author Affiliations

1 Department of Infectious Disease, the First Affiliated Hospital of Nanjing Medical University, Nanjing, China

2 Department of Microbiology and Immunology, Nanjing Medical University, Nanjing, China

3 The Third Hospital of Zhenjiang City, Zhenjiang, China

4 Nanjing Red Cross Blood Center, Nanjing, China

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Virology Journal 2013, 10:292 doi:10.1186/1743-422X-10-292

The electronic version of this article is the complete one and can be found online at: http://www.virologyj.com/content/10/1/292

Received:    29 March 2013
Accepted:    17 September 2013
Published:    22 September 2013

© 2013 Zhang et al.; licensee BioMed Central Ltd.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Abstract
Background

Selective pressure from either the immune response or the use of nucleoside analogs in antiviral therapy could be driving the emergence of HBV mutants. Because of the overlap of the open reading frame (ORF) S for the HBsAg and ORF P for viral polymerase, rtM204I and rtM204V mutations in the polymerase would produce sI195M and sW196S in the HBsAg. The combined effects of immune-escaped mutations (sT118M, sG145K, sG145R) and drug-resistant mutations (rtM204I, rtM204V) on the antigenicity profiles of HBsAg has not been widely explored.
Methods

To determine the combined effects of immune-escaped and drug-resistant mutants on the antigenicity profiles of HBsAg, recombinant plasmids encoding HBsAg double mutants were constructed using site-directed mutagenesis. The supernatant from each plasmid transfection was analyzed for HBsAg in the western-blotting and five of the most commonly used commercial ELISA kits in China.
Results

Western-blotting assay showed the successful expression of each HBsAg mutant. All five ELISA kits manifested similar avidity, which were demonstrated by the slope of the curves, for the sT118M mutant, and sT118M-rtM204I (sT118M-sI195M) and sT118M-rtM204V (sT118M-sW196S) double mutants, suggesting that drug-resistant YMDD mutants caused negligible losses in the antigenicity of immune-escaped sT118M HBsAg. In contrast, the presence of the rtM204I (sI195M) mutation, but not rtM204V (sW196S) in combination with the sG145K mutation significantly reduced the avidity of sG145K HBsAg. The rtM204I (sI195M) mutation also decreased the antigenicity profiles for sG145R HBsAg.
Conclusions

Drug-resistant mutations rtM204I (sI195M) and rtM204V (sW196S) caused significant reduction in antigenicity for the immune-escaped HBsAg mutants sG145K and sG145R, which may hamper HBV diagnosis and disease control from HBV blood-transfusion transmissions in China. The development of ELISA kits with a greater sensitivity for drug-resistant and immune-escaped HBsAg warrants further consideration.

StephenW

明顺Zhang12 † ，国宏舸† ，永霖Yang4 ，王绪兵Cai4 ， FU4强，杰Cai4 * Zuhu Huang1 *

    *通讯作者：蔡杰[email protected]的 - 黄[email protected] Zuhu
    †平等贡献者

作者单位

1部传染病，南京中医药大学，南京，中国第一附属医院

2 ，南京医科大学微生物学和免疫学系，南京，中国

3镇江市第三人民医院，镇江，中国

4南京红十字血液中心，南京，中国


病毒学杂志2013 ， 10:292 DOI ： 10.1186/1743-422X-10-292

电子版本的这篇文章是完整的，并可以在网上找到： http://www.virologyj.com/content/10/1/292

收稿日期： 2013年3月29日
接受日期： 2013年9月17日
发布时间： 2013年9月22日

© 2013张等;持牌BioMed Central的有限公司

这是一个开放的分布的条款下的知识共享署名许可（ http://creativecommons.org/licenses/by/2.0 ） ，允许无限制地使用，分发和再现任何媒体的访问文章，提供了原来的工作是正确引用。
抽象
背景

从免疫响应，或者在使用核苷类似物抗病毒治疗的选择压力可以驾驶HBV突变体的出现。由于重叠的开放阅读框（ORF） ， HBsAg和ORF P S为病毒聚合酶， rtM204I和rtM204V突变的聚合酶产生的HBsAg sI195M sW196S 。免疫逃脱的突变（ sT118M ， sG145K sG145R ）和耐药突变（ rtM204I ， rtM204V ） HBsAg的抗原分布的综合影响还没有被广泛探讨。
方法

要确定的综合影响免疫逃脱，采用定点突变耐药突变对HBsAg的抗原型材，构建了重组质粒编码HBsAg的双突变体。从每个质粒转染上清液中HBsAg西部印迹和五个最常用的商业在中国的ELISA试剂盒分析。
结果

蛋白质印迹检测结果显示每一个乙肝表面抗原突变体的成功表达。所有五个ELISA试剂盒表现相似的亲合力，由该曲线的斜率，突变体sT118M ， ： rtM204I sT118M （ sT118M - sI195M ）和sT118M rtM204V （ sT118M - sW196S ）双突变体进行了论证，表明耐药性YMDD突变引起的免疫逃脱sT118M的的HBsAg抗原可以忽略不计的损失。与此相反，在存在下rtM204I （ sI195M ）的突变，但不rtM204V （ sW196S ）结合的sG145K突变可显着降低的亲和力sG145K HBsAg的。的sG145R乙肝表面抗原rtM204I （ sI195M ）的突变也降低了抗原型材。
结论

耐药突变rtM204I （ sI195M ）的rtM204V （ sW196S ）造成显着减少抗原的免疫逃脱乙肝表面抗原突变体sG145K sG145R ，这可能妨碍乙肝诊断和疾病控制在中国从HBV输血传输的。 ELISA试剂盒的发展具有更大的敏感性耐药和免疫逃脱HBsAg的认股权证进一步考虑。