嗜水气单胞菌ompA、flaA基因重组干酪乳杆菌对鲤生长及免疫效果分析
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国家现代农业产业技术体系建设专项 (CARS- 46); 国家自然科学基金(31372540)


Effects of recombinant Lactobacillus casei from Aeromonas hydrophila ompA and flaA genes on the growth and immunity in common carp (Cyprinus carpio)
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    摘要:

    为构建嗜水气单胞菌ompAflaA基因重组干酪乳杆菌并分别检测其表达产物对鲤生长及免疫效果的影响,实验将目的基因克隆至乳酸菌穿梭表达质粒pPG612中,并电转至干酪乳杆菌中,经诱导后包被饲料对鲤进行饲养56 d,称重并采集血清及组织,分析生长指标及免疫指标变化。在56 d饲养免疫结束后结果显示,生长指标显示重组干酪乳杆菌组与对照组无显著差异,表明重组干酪乳杆菌对生长无影响;免疫效果分析显示,血清中 IgM 表达水平显著上升。血清中AKP、LZM、SOD、CAT、C3和C4也均显著升高。荧光定量PCR检测发现,免疫后肝脏、脾脏、头肾及肠组织中的细胞免疫因子基因IL-1βIL-8、TNF-αNF-κBTLR5及MYD88的表达水平有不同程度的显著提升;其中TLR5及MYD88表达量在Lc-mcs-flaA组高于Lc-mcs-ompA组。攻毒后Lc-mcs-ompA与Lc-mcs-flaA组免疫保护率分别为59%与54%,显著高于对照组。研究表明,本实验构建的重组干酪乳杆菌Lc-mcs-ompA与Lc-mcs-flaA免疫鲤能刺激机体产生免疫应答反应,进而提高自身存活率,为预防鱼类嗜水气单胞菌感染的口服免疫制剂的研发奠定基础并提供科学理论依据。

    Abstract:

    To construct recombinant Lactobacillus casei with ompA and flaA of Aeromonas hydrophila and to detect the growth and immunogenicity of their expression products to common carp, Cyprinus carpio, the target genes were cloned into Lactobacillus Shuttle expression plasmid pPG612, and electric-transfered into L. casei. Fish were fed with dietary mixed with different recombinant L. casei for 56 days, and we weighed and collected serum and tissues, and analyzed the growth and related immune index changes; feeding at 56 days after the immunization, the growth index results showed that the recombinant L. casei had no significant difference compared with the control group, which showed that it had no effect on growth, immune indexes. Results showed that level of IgM in serum was significantly increased, alkaline phosphatase (AKP), lysozyme (LZM), superoxide dismutase (SOD), catalase (CAT), Complement 3 (C3) and Complement 4 (C4) were also significantly increased, qRT-PCR detection found that the expression levels of Interleukin 1β (IL-1β), Interleukin 8 (IL-8), Tumor Necrosis Factor α (TNF-α), Nuclear factor-kappa B (NF-κB), Toll-like receptor 5 (TLR) and myeloid differentiation factor 88 (MYD88) in liver, spleen, kidney and intestinal tissues after immunization increased significantly to varying degrees; the expression of TLR5 in the Lc-mcs-flaA group was higher than that in Lc-mcs-ompA groups; after challenge, the immune protection rates of Lc-mcs-ompA and Lc-mcs-flaA were 59% and 54%, respectively, and the survival rate was significantly higher than control group. The constructed recombinant L. casei Lc-mcs-ompA and Lc-mcs-flaA immuned carp could stimulate the body to activate immune response, thereby increasing its survival rate. It is an oral solution for preventing fish Aeromonas hydrophila infection. The research and development of immune preparations laid the foundation and provided scientific theoretical basis.

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赵林辉,田佳鑫,孔祎頔,彭思博,单晓枫,王桂芹.嗜水气单胞菌ompA、flaA基因重组干酪乳杆菌对鲤生长及免疫效果分析[J].水产学报,2022,46(1):73~84

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  • 收稿日期:2020-02-21
  • 最后修改日期:2021-04-11
  • 录用日期:2021-04-11
  • 在线发布日期: 2022-01-05
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