文章摘要
单壁碳纳米管载嗜水气单胞菌外膜蛋白亚单位疫苗的研制及免疫效果评价
Preparation of single-walled carbon nanotubes carrying Aeromonas hydrophila outer membrane protein subunit vaccine and evaluation of immune effect
投稿时间:2019-04-20  修订日期:2019-06-27
DOI:
中文关键词: 嗜水气单胞菌  外膜蛋白  碳纳米管  注射免疫  浸泡免疫
英文关键词: Aeromonas hydrophila  outer membrane protein  carbon nanotubes  injection immunization  immersion immunizatio
基金项目:外膜蛋白LamB介导的细菌负调控耐药机制研究
作者单位E-mail
姜倩雯 福建农林大学生命科学学院 1274940988@qq.com 
张丽珊 福建农林大学生命科学学院  
李小艳 福建农林大学生命科学学院  
武瑶 福建农林大学生命科学学院  
赵怡扬 福建农林大学生命科学学院  
徐慧铭 福建农林大学生命科学学院  
林向民 福建农林大学生命科学学院 xiangmin@fafu.edu.cn 
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中文摘要:
      嗜水气单胞菌(Aeromonas hydrophilia)是一种常见的造成水产养殖业损失的主要病原菌。疫苗免疫被认为是抑制该菌感染最有效的途径之一。近年来,单壁碳纳米管(Single-walled carbon nanotube,SWCNTs)在生物医学领域得到较好的发展,并在水产养殖疫苗中得到初步的应用。课题组前期的研究发现在缺铁环境下高表达的嗜水气单胞菌外膜蛋白可能作为细菌的疫苗候选蛋白,本研究挑选了2个嗜水气单胞菌外膜蛋白(A0KLQ6、A0KH89)并以A0KHR9(OmpAII)为对照,比较外膜蛋白与碳纳米管载外膜蛋白,分别在腹腔注射与浸泡免疫模式对斑马鱼的免疫反应与免疫保护效果。结果显示:腹腔注射组和浸泡组,与普通蛋白组对比,碳纳米管载蛋白后能够显著提高斑马鱼免疫相关基因表达量。腹腔注射组在5μg剂量下免疫A0KHR9、A0KLQ6、A0KH89蛋白分别获得61.6 %、65.39 %、84.96 %相对免疫保护率(Relative immune protection rate,RPS),在5 μg剂量下免疫碳纳米管载蛋白SWCNTs-A0KHR9、SWCNTs-A0KLQ6、SWCNTs-A0KH89分别获得68.10 %、77.50 %、90.43 %的RPSs。浸泡组在40 mg/L剂量下免疫A0KHR9、A0KLQ6、A0KH89蛋白分别获得30.80 %、25.85 %、37.80 % RPSs,在40 mg/L最高剂量下免疫碳纳米管载蛋白疫苗SWCNTs-A0KHR9、SWCNTs-A0KLQ6、SWCNTs-A0KH89分别获得63.60 %、73.74 %、68.49 % RPSs。上述结果表明,铁离子限制下嗜水气单胞菌外膜蛋白A0KLQ6和A0KH89等蛋白能够作为抗嗜水气单胞菌的亚单位疫苗候选蛋白,而单壁碳纳米管载蛋白通过注射免疫与浸泡免疫均能显著提升免疫疗效,是一种疫苗的适宜纳米载体。以上研究为寻找高效的嗜水气单胞菌亚单位疫苗候选蛋白及佐剂提供了理论依据。
英文摘要:
      Aeromonas hydrophilia is a common human-animal-fish symbiotic aquatic pathogen and is one of the main pathogens causing bacterial disease in aquaculture. Vaccine immunization is considered to be one of the most effective ways to inhibit infection. In recent years, single-walled carbon nanotubes (SWCNTs) have been well developed in the biomedical field and have been initially applied in aquaculture vaccines. According to our reviously research, the iron related outer membrane proteins of A. hydrophila may be used as vaccine candidates. To investigate the immune response of zebrafish to iron-related proteins and SWCNT-encapsulated proteins, we overexpressed and purified two iron-related recombinant proteins (A0KLQ6 and A0KH89) from A. hydrophila. We then vaccinated zebrafish with these proteins and their SWCNT-encapsulated counterparts via both intraperitoneal injection and bath immunization, and found they evoked a strong immune response in zebrafish. When challenged with virulent A. hydrophila, zebrafish administered 5 μg intraperitoneal injections of A0KHR9, A0KLQ6, A0KH89, SWCNTs-A0KHR9, SWCNTs-A0KLQ6 and SWCNTs-A0KH89 obtained high relative percent survivals (RPSs) (51.6%, 60.59%, 84.96%, 68.10%, 77.50%, and 90.43%, respectively). Meanwhile, the zebrafish administered 40 mg/L bath immunizations of aboved samples had 30.80 %, 25.85 %, 37.80 %, 63.60%, 73.74% and 68.49% RPSs, respectively. These results indicate that the outer membrane proteins A0KLQ6 and A0KH89 can be used as potenial subunit vaccine candidates against A. hydrophila infection, and the SWCNTs -encapsulated counterparts can significantly improve the immune effect in both immunization ways. Our studies provide a theoretical basis for the development of vaccines and adjuvants candidates.
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