表达铜绿假单胞菌保护性抗原基因F1I2重组鼠伤寒沙门菌在水貂上的免疫原性研究

doi:10.16431/j.cnki.1671-7236.2020.04.021

中国畜牧兽医 ›› 2020, Vol. 47 ›› Issue (4): 1156-1162.doi: 10.16431/j.cnki.1671-7236.2020.04.021

表达铜绿假单胞菌保护性抗原基因F1I2重组鼠伤寒沙门菌在水貂上的免疫原性研究

张明亮^1,2,3, 孙长江², 顾敬敏², 崔子寅², 韩文瑜²

1. 安阳工学院生物与食品工程学院, 安阳 455000;
2. 吉林大学动物医学学院, 长春 130062;
3. 河南省动物疫病防控与营养免疫院士工作站, 安阳 455000;
4. 河南省兽用生物制品研发与应用国际联合实验室, 安阳 455000

收稿日期:2019-09-18 出版日期:2020-04-20 发布日期:2020-04-17
通讯作者: 韩文瑜 E-mail:hanwy@jlu.edu.cn
作者简介:张明亮(1985-),男,河南遂平人,博士,讲师,研究方向:病原微生物和分子免疫学,E-mail:mingliang90909@163.com;孙长江(1978-),男,吉林梅河口人,博士,讲师,研究方向:人畜共患传染病防治,E-mail:18021346@qq.com
基金资助:
国家高技术研究发展计划项目（2011AA10A210）；国家自然科学基金青年基金项目（31802170）；安阳工学院博士科研启动基金项目（BSJ2016013）

Immunogenicity of Recombinant Salmonella Typhimurium Expressing Protective Antigen Gene F1I2 of Pseudomonas aeruginosa in Mink

ZHANG Mingliang^1,2,3, SUN Changjiang², GU Jingmin², CUI Ziyin², HAN Wenyu²

1. College of Biological Science and Food Engineering, Anyang Institute of Technology, Anyang 455000, China;
2. College of Veterinary Medicine, Jilin University, Changchun 130062, China;
3. Academician Workstation of Animal Disease Control and Nutrition Immunity in Henan Province, Anyang 455000, China;
4. Henan Joint International Research Laboratory of Veterinary Biologics Research and Application, Anyang 455000, China

Received:2019-09-18 Online:2020-04-20 Published:2020-04-17

摘要/Abstract

摘要： 本试验旨在研究携带铜绿假单胞菌保护性抗原基因F_190-342（F1）和I_21-83（I2）的重组鼠伤寒沙门菌株ΔasdLH430（pYA-F1I2）在水貂上的免疫原性。将9只7月龄健康水貂随机分为3组，每组3只：对照组，皮下注射无菌生理盐水；LH430菌株免疫组，皮下注射菌株2.0×10⁸ CFU/只；ΔasdLH430（pYA-F1I2）菌株免疫组，皮下注射菌株2.0×10⁸ CFU/只。首免后第15天各组加强免疫一次。分别于第1次免疫后第15、30及45天断指采集水貂血液，测定血清中IgG水平。在第1次免疫后第45天分离外周血单核细胞，Eli-Spot检测F1I2特异性抗体分泌细胞数量和沙门菌特异性抗体分泌细胞数量。同时，取脾脏分离淋巴细胞，MTT法检测F1I2和沙门菌特异性的淋巴细胞增殖情况。结果表明，ΔasdLH430（pYA-F1I2）组血清中F1I2特异性IgG抗体和沙门菌特异性IgG抗体滴度在取样点逐渐升高，同时在第45天达到最大值；免疫后第45天，ΔasdLH430（pYA-F1I2）组F1I2特异性IgG分泌细胞的数量极显著高于对照组及LH430组（P < 0.01），沙门菌特异性抗体分泌细胞的数量极显著高于对照组（P < 0.01），与LH430组差异不显著（P > 0.05）。同时，F1I2抗原和沙门菌抗原刺激组极显著增强了水貂脾脏淋巴细胞的增殖（P < 0.01）。本研究可为开发防控水貂出血性肺炎和沙门菌感染的实用新型双价基因工程疫苗提供理论依据。

关键词: 铜绿假单胞菌; 水貂; 出血性肺炎; 免疫原性

Abstract: The purpose of this study was to investigate the immunogenicity of recombinant Salmonella Typhimurium strain Δasdlh430(pYA-F1I2) carrying Pseudomonas aeruginosa protective antigen genes F_190-342(F1) and I_21-83(I2) in mink.Nine 7-month-old healthy minks were randomly divided into three groups,three in each group:Control group,hypodermic injection of sterile normal saline;LH430 strain immunization group,hypodermic injection of 2.0×10⁸ CFU per mink;Δasdlh430(pYA-F1I2) strain immunization group,hypodermic injection of 2.0×10⁸ CFU per mink.Every group was immunized once on the 15th day after the first immunization.On the 15th,30th and 45th day after the first immunization,the blood of mink was collected and the level of IgG in serum was measured.Peripheral blood monocytes were isolated on the 45th day after the first immunization.The number of F1I2 specific antibody secreting cells and Salmonella specific antibody secreting cells were detected by Eli-Spot.At the same time,lymphocytes were isolated from the spleen,and the specific lymphocyte proliferation of F1I2 and Salmonella was detected by MTT method.The results showed that the titers of F1I2 specific IgG antibody and Salmonella specific IgG antibody in serum of Δasdlh430(pYA-F1I2) group increased gradually at the sampling point,and reached the maximum value at the 45th day.On the 45th day after immunization,the number of F1I2 specific IgG secretory cells in Δasdlh430(pYA-F1I2) group was extremely significantly higher than that in control and LH430 groups (P < 0.01),and the number of Salmonella specific antibody secretory cells was extremely significantly higher than that of control group (P < 0.01),but there was no significant difference with LH430 group (P > 0.05).At the same time,F1I2 antigen and Salmonella antigen stimulated the proliferation of mink spleen lymphocytes significantly (P < 0.01).This study could provide a theoretical basis for the development of a new bivalent genetic engineering vaccine against mink hemorrhagic pneumonia and Salmonella infection.

Key words: Pseudomonas aeruginosa; mink; hemorrhagic pneumonia; immunogenicity

中图分类号:

S852.61

张明亮, 孙长江, 顾敬敏, 崔子寅, 韩文瑜. 表达铜绿假单胞菌保护性抗原基因F1I2重组鼠伤寒沙门菌在水貂上的免疫原性研究[J]. 中国畜牧兽医, 2020, 47(4): 1156-1162.

ZHANG Mingliang, SUN Changjiang, GU Jingmin, CUI Ziyin, HAN Wenyu. Immunogenicity of Recombinant Salmonella Typhimurium Expressing Protective Antigen Gene F1I2 of Pseudomonas aeruginosa in Mink[J]. China Animal Husbandry and Veterinary Medicine, 2020, 47(4): 1156-1162.

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表达铜绿假单胞菌保护性抗原基因F1I2重组鼠伤寒沙门菌在水貂上的免疫原性研究

Immunogenicity of Recombinant Salmonella Typhimurium Expressing Protective Antigen Gene F1I2 of Pseudomonas aeruginosa in Mink

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