重组乳酸乳球菌展示维氏气单胞菌 Aha1 与 LTB 佐剂在鲤鱼中的免疫效果。

Immunization effect of recombinant Lactobacillus casei displaying Aeromonas veronii Aha1 with an LTB adjuvant in carp.

机构信息

College of Animal Science and Technology, Jilin Agricultural University, Changchun, China.

College of Animal Science and Technology, Jilin University, Changchun, China.

出版信息

Fish Shellfish Immunol. 2023 Apr;135:108660. doi: 10.1016/j.fsi.2023.108660. Epub 2023 Mar 20.

DOI:10.1016/j.fsi.2023.108660

PMID:36940784

Abstract

Aeromonas veronii is an important aquatic zoonotic, which elicits a range of diseases, such as haemorrhagic septicemia. To develop an effective oral vaccine against Aeromonas veronii infection in carp, the Aeromonas veronii adhesion (Aha1) gene was used as a target molecule to attach to intestinal epithelial cells. Two anchored recombinant. Lactic acid bacteria strains (LC-pPG-Aha1 1038 bp and LC-pPG-Aha1-LTB 1383 bp) were constructed by fusing them with the E. coli intolerant enterotoxin B subunit (LTB) gene and using Lactobacillus casei as antigen delivery vector to evaluate immune effects of these in carp. Western blotting and immunofluorescence were used to confirm that protein expression was successful. Additionally, levels of specific IgM in serum and the activities of ACP, AKP, SOD, LYS, C3, C4, and lectin enzymes-were assessed. Cytokines IL-10, IL-1β, TNF-α, IgZ1, and IgZ2 were measured in the liver, spleen, kidney, intestines, and gills tissue by qRT-PCR, which showed an increasing trend compared with the control group (P < 0.05). A colonization assay showed that the two L. casei recombinants colonized the middle and hind intestines of immunized fish. When immunized carp were experimentally challenged with Aeromonas veronii the relative percentage protection of LC-pPG-Aha1 was 53.57%, and LC-pPG-Aha1-LTB was 60.71%. In conclusion, these results demonstrate that Aha1 is a promising candidate antigen when it is displayed on lactic acid bacteria (Lc-pPG-Aha1 and Lc-pPG-Aha1-LTB) seems promising for a mucosal therapeutic approach. We plan to investigate the molecular mechanism of the L. casei recombinant in regulating the intestinal tissue of carp in future studies.

摘要

维氏气单胞菌是一种重要的水生动物源性致病菌，可引起多种疾病，如出血性败血症。为了开发一种针对鲤鱼维氏气单胞菌感染的有效口服疫苗，将维氏气单胞菌黏附（Aha1）基因作为靶分子，使其附着在肠道上皮细胞上。将该基因与大肠杆菌不耐受肠毒素 B 亚基（LTB）基因融合，构建了两种锚定重组乳酸杆菌菌株（LC-pPG-Aha1 为 1038bp，LC-pPG-Aha1-LTB 为 1383bp），并将其用作乳杆菌抗原传递载体，以评估这些在鲤鱼中的免疫效果。通过 Western blot 和免疫荧光技术证实了蛋白表达的成功。此外，还评估了血清中特异性 IgM 的水平以及 ACP、AKP、SOD、LYS、C3、C4 和凝集素酶的活性。通过 qRT-PCR 测量了肝、脾、肾、肠和鳃组织中的细胞因子 IL-10、IL-1β、TNF-α、IgZ1 和 IgZ2，与对照组相比呈上升趋势（P<0.05）。定植试验表明，两种 L. casei 重组菌定植于免疫鱼的中后肠。当免疫鲤鱼受到维氏气单胞菌攻毒时，LC-pPG-Aha1 的相对保护率为 53.57%，LC-pPG-Aha1-LTB 为 60.71%。综上所述，这些结果表明 Aha1 是一种很有前途的候选抗原，当它在乳酸杆菌（LC-pPG-Aha1 和 LC-pPG-Aha1-LTB）上展示时，似乎对黏膜治疗方法有希望。我们计划在未来的研究中研究 L. casei 重组菌调节鲤鱼肠道组织的分子机制。