Hepatitis-hydropericardium syndrome (HPS) causes severe economic losses in the global poultry industry. The present study aims to explore oral immunization of recombinant Lactococcus lactis and Enterococcus faecalis expressing Hexon protein of fowl adenovirus 4 (FAdV-4). The bacteria L. lactis NZ9000 and E. faecalis MDXEF-1 were, respectively, modified as host strain to deliver truncated Hexon protein (ΔHexon) or ΔHexon protein fusing with dendritic cell (DC) targeting peptide (DC-ΔHexon) on the surface of bacteria. The expression of target protein in L. lactis NZ9000 and E. faecalis MDXEF-1 were detected by western blot. To evaluate the immune responses and protective efficacies provided by the live recombinant bacteria, chickens were immunized with the constructed ΔHexon-expressing bacteria three times at 2-week intervals, then experimentally challenged with hypervirulent FAdV-4/GX01. The results showed that oral immunizations with the four ΔHexon-expressing bacteria (NZ9000/ΔHexon-CWA, NZ9000/DC-ΔHexon-CWA, MDXEF-1/ΔHexon-CWA, and MDXEF-1/DC-ΔHexon-CWA), especially the two bacteria carrying DC-targeting peptide, stimulated higher levels of ΔHexon-specific sera IgG and secretory IgA (sIgA) in jejunal lavage fluid, higher proliferation of peripheral blood lymphocytes (PBLs) and higher levels of Th1/Th2-type cytokines, along with significantly decreased virus loads in liver and more offered protective efficacies against FAdV infection compared with PBS and empty vector control groups (p < 0.01). For chickens in the group MDXEF-1/DC-ΔHexon-CWA, the levels of aspartate transaminase (AST), alanine transaminase (ALT) and lactate dehydrogenase (LDH) in sera, and the virus loads in livers were significantly decreased vs. the other three ΔHexon-expressing bacteria (p < 0.01). The pathological changes in the hearts, livers, spleens and kidneys of chickens in MDXEF-1/DC-ΔHexon-CWA group were relatively slight compared to infection control group and other three ΔHexon-expressing bacteria groups. The rate of protection in MDXEF-1/DC-ΔHexon-CWA group was 90%. The present work demonstrated that cell surface-displayed target protein and immune enhancers in L. lactis and E. faecalis might be a promising approach to enhance immunity and immune efficacy against pathogen FAdV-4 infection.

肝炎-水囊病综合症（HPS）在全球家禽业中造成严重的经济损失。本研究旨在探讨重组体的口服免疫乳酸乳球菌 和 粪肠球菌表达禽腺病毒4（FAdV-4）的六邻体蛋白。细菌乳酸乳球菌 NZ9000和 粪肠球菌将MDXEF-1分别修饰为宿主菌株，以将截短的Hexon蛋白（ΔHexon）或ΔHexon蛋白与细菌表面上的树突状细胞（DC）靶向肽（DC-ΔHexon）融合。靶蛋白在组织中的表达乳酸乳球菌 NZ9000和 粪肠球菌通过蛋白质印迹检测MDXEF-1。为了评估由活重组细菌提供的免疫反应和保护效力，将鸡用构建的表达ΔHexon的细菌每2周间隔免疫3次，然后用高毒性FAdV-4 / GX01进行实验攻击。结果表明，使用四种表达ΔHexon的细菌（NZ9000 /ΔHexon-CWA，NZ9000 /DC-ΔHexon-CWA，MDXEF-1 /ΔHexon-CWA和MDXEF-1 /DC-ΔHexon-CWA）进行口服免疫。两种携带DC靶向肽的细菌在空肠灌洗液中刺激了更高水平的ΔHexon特异性血清IgG和分泌性IgA（sIgA），更高的外周血淋巴细胞（PBL）增殖和更高的Th1 / Th2型细胞因子水平，p<0.01）。对于MDXEF-1 /DC-ΔHexon-CWA组中的鸡，血清中的天冬氨酸转氨酶（AST），丙氨酸转氨酶（ALT）和乳酸脱氢酶（LDH）的水平与肝脏相比明显降低。其他三种表达ΔHexon的细菌（p<0.01）。与感染对照组和其他三个表达ΔHexon的细菌组相比，MDXEF-1 /DC-ΔHexon-CWA组的鸡的心脏，肝脏，脾脏和肾脏的病理变化相对较小。MDXEF-1 /DC-ΔHexon-CWA组的保护率为90％。目前的工作表明细胞表面展示的靶蛋白和免疫增强剂乳酸乳球菌 和 粪肠球菌 可能是增强针对病原体FAdV-4感染的免疫力和免疫效力的有前途的方法。