文献类型：期刊文献

中文题名：甲型副伤寒杆菌ompA基因分布及重组表达产物的免疫学鉴定

英文题名：Distribution of Salmonella paratyphi A ompA gene and immunological identification of the recombinant expressed product

作者：蒋锦琴[1,2];阮萍[3];丁威[4];孙爱华[2];毛亚飞[1];严杰[1]

机构：[1]浙江大学医学院病原生物学系,杭州310058;[2]浙江医学高等专科学校,杭州310053;[3]浙江绍兴文理学院医学院医学检验系,312000;[4]浙江省血液中心

年份：2010

期号：1

起止页码：1

中文期刊名：中华微生物学和免疫学杂志

外文期刊名：Chinese Journal of Microbiology and Immunology

收录：CSTPCD、、Scopus、CSCD2011_2012、北大核心2008、北大核心、CSCD

基金：浙江省自然科学基金（Y2090395）;浙江省医药卫生科学研究基金（2006A116）;浙江医学高等专科学校自然科学研究基金（2005xZ04）

语种：中文

中文关键词：甲型副伤寒杆菌;ompA基因;重组表达;免疫原性;免疫保护性

外文关键词：Salmonella paratyphi A ; ompA gene ; Recombinant expression ; Immunogenicity ; Immunoprotection

中文摘要：目的构建甲型副伤寒杆菌ompA基因原核表达系统，确定其重组表达产物rOmpA免疫原性和保护作用，检测甲型副伤寒杆菌临床菌株ompA基因携带率。方法采用PCR从甲型副伤寒杆菌临床株JH01中扩增ompA基因，T-A克隆后测序并构建ompA基因原核表达系统。采用SDS-PAGE和Bio—Rad凝胶图像分析系统检测rOmpA表达情况及其产量，采用免疫扩散法、微量肥达试验和Western blot鉴定其抗原性和免疫反应性。采用PCR检测98株甲型副伤寒杆菌临床菌株ompA基因携带率。采用小鼠感染模型，了解rOmpA对甲型副伤寒杆菌50001株感染的免疫保护作用。结果与报道的相关序列比较，所克隆的ompA基因核苷酸和氨基酸序列相似性均为100％。rOmpA表达量约为细菌总蛋白的65％。rOmpA能与其兔抗血清和甲型副伤寒杆菌全菌抗血清产生免疫反应（Western blot），并可在家兔中诱导产生抗体。94．9％（93／98）甲型副伤寒杆菌临床菌株含有ompA基因。100μg和200μg rOmpA对感染小鼠的免疫保护率分别为41．7％（5／12）和58．3％（7／12）。rOmpA免疫小鼠或保护试验存活小鼠血清对各副伤寒杆菌H抗原的凝集效价为1：5～1：40。结论ompA基因在甲型副伤寒杆菌临床菌株中分布广泛。rOmpA有良好的免疫原性和一定的免疫保护作用，可考虑为甲型副伤寒杆菌基因工程疫苗候选抗原。

外文摘要：Objective To construct a prokaryotic expression vector of Salmonella paratyphi A ompA gene, and to determine immunogenicity and immonuprotection of the recombinant expressed product rOmpA and carrying frequency of ompA gene in S. paratyphi A isolates. Methods ompA gene was amplified from a clinical S. paratyphi A strain JH01 by PCR, cloned into T-A cloning vector, and then the prokaryotic expression vector was constructed. The rOmpA expression was determined by SDS-PAGE. Antigenicity and immunoreactivity of rOmpA were determined by immunodiffusion test, Micro-Widal＇s test and Western blot assay, ompA gene exiting in 98 S. paratyphi A isolates was detected by PCR. By using immunoprotective effect of rOmpA against the challenge of S. paratyphi A strain 50001 in mice were determinded. Results Both the nucleotide and putative amino acid sequence identities of the cloned ompA gene were 100% com- pared to the reported corresponding sequences. The expression yield of rOmpA was approximately 65% of the total bacterial proteins, rOmpA was able to induce specific antibody in rabbits, and reacted efficiently with rabbit antiseruu or the antiserum against whole cell of S. paratyphi A detected by Western blot. 94.9% （93/98） of the S. paratyphi A isolates had ompA gene. 41. 7% （5/12） and 58.3% （7/12） of the mice immunized with 100 and 200 μg rOmpA were survival after lethal challenge with S. paratyphi A strain 50001. The titer of antibody to the H antigens of S. paratyphi spp in the sera from rOmpA immunized mice and survival mice in the S. paratyphi A challenge detected by Micro-Widal＇s test was 1：5-1 ： 40. Conclusion ompA gene has an extensive distribution in clinical isolates of S. paratyphi A. rOmpA possesses an immunogenicity and a certain immunoprotective effect, and can be used as the candidate antigen in genetic engineering vaccine.