College of Animal Science and Technology, Jilin Agricultural University, Changchun, 130118, China; Jilin Provincial Key Laboratory of Animal Nutrition and Feed Science, Jilin Agricultural University, Changchun, 130118, China; Joint Laboratory of Modern Agricultural Technology International Cooperation, Ministry of Education, Jilin Agricultural University, Changchun, Jilin, 130118, China; Ministry of Education Laboratory of Animal Production and Quality Security, Jilin Agricultural University, Changchun, Jilin, 130118, China.
College of Animal Science and Technology, Jilin Agricultural University, Changchun, 130118, China.
Fish Shellfish Immunol. 2020 Apr;99:73-85. doi: 10.1016/j.fsi.2020.02.008. Epub 2020 Feb 4.
In the present study, we constructed two recombinant Lactobacillus casei (L. casei) Lc-pPG-1-AcrV (surface-displayed) and Lc-pPG-2-AcrV (secretory) constitutively expressing AcrV protein of Aeromonas veronii (A. veronii). Expression of recombinant AcrV protein was verified by western blot and immunofluorescence technique. Compared with PBS group, the final weight (FW), weight gain (WG) and specific growth rate (SGR) of fish fed Lc-pPG-1-AcrV, Lc-pPG-2-AcrV and Lc-pPG diets after 56 days observed significantly increase (p < 0.05), while the feed conversion ratio (FCR) showed a significantly decrease (p < 0.05). The recombinant L. casei strains were orally administrated to crucian carp, and significant increased (p < 0.05) the immunoglobulin M (IgM), elevated the acid phosphatase (ACP), alkaline phosphatase (AKP), lysozyme (LZM) and superoxide dismutase (SOD) activity in serum. Moreover, leukocytes phagocytosis percentage and index of the recombinant L. casei were both enhanced. The results demonstrated that the recombinant L. casei could elicit systemic immune responses and increase the serum immunological index. The Interleukin-10 (IL-10), Interleukin-1β (IL-1β), interferon-γ (IFN-γ) and Tumor Necrosis Factor-α (TNF-α) levels in liver, spleen, kidney and intestine have up regulated significantly in tissues (p < 0.05), suggesting that the recombinant L. casei has the ability to induce expression of cytokines and enhance the innate immune response. Higher survival rates were exhibited that crucian carp immunized with Lc-pPG-1-AcrV (67.5%) and Lc-pPG-2-AcrV (52.5%) after challenge with A. veronii. In conclusion, these two recombinant L. casei vaccine were effective in improving crucian carp growth, immunity response and disease resistance. The recombinant L. casei strains may be a promising candidate for the development of an oral vaccine against A. veronii.
在本研究中，我们构建了两种重组干酪乳杆菌（L. casei）Lc-pPG-1-AcrV（表面展示）和 Lc-pPG-2-AcrV（分泌型），它们持续表达维氏气单胞菌（A. veronii）的 AcrV 蛋白。通过 Western blot 和免疫荧光技术验证了重组 AcrV 蛋白的表达。与 PBS 组相比，经过 56 天喂养后，Lc-pPG-1-AcrV、Lc-pPG-2-AcrV 和 Lc-pPG 饮食组的终重（FW）、增重（WG）和特定生长率（SGR）显著增加（p<0.05），而饲料转化率（FCR）显著降低（p<0.05）。将重组 L. casei 菌株口服给药给鲫鱼，显著增加（p<0.05）血清中的免疫球蛋白 M（IgM），提高酸性磷酸酶（ACP）、碱性磷酸酶（AKP）、溶菌酶（LZM）和超氧化物歧化酶（SOD）的活性。此外，白细胞吞噬率和重组 L. casei 的指数均得到增强。结果表明，重组 L. casei 可以引起全身免疫反应并提高血清免疫学指标。肝、脾、肾和肠组织中的白细胞介素 10（IL-10）、白细胞介素 1β（IL-1β）、干扰素-γ（IFN-γ）和肿瘤坏死因子-α（TNF-α）水平均显著上调（p<0.05），表明重组 L. casei 具有诱导细胞因子表达和增强固有免疫反应的能力。经 Lc-pPG-1-AcrV（67.5%）和 Lc-pPG-2-AcrV（52.5%）免疫的鲫鱼在受到维氏气单胞菌攻击后，存活率更高。综上所述，这两种重组 L. casei 疫苗可有效提高鲫鱼的生长、免疫反应和抗病能力。重组 L. casei 菌株可能是开发针对维氏气单胞菌的口服疫苗的有前途的候选者。
