Ma Chunli, Zhang Lili, Gao Mingyang, Ma Dexing
College of Food Science, Northeast Agricultural University, NO. 59 Mucai Street, Harbin 150030, China.
College of Veterinary Medicine, Northeast Agricultural University, NO. 59 Mucai Street, Harbin 150030, China; Heilongjiang Key Laboratory for Laboratory Animals and Comparative Medicine, NO. 59 Mucai Street, Harbin 150030, China.
Exp Parasitol. 2017 Jul;178:14-20. doi: 10.1016/j.exppara.2017.05.001. Epub 2017 May 17.
Two novel plasmids pTX8048-SP-Δ3-1E and pTX8048-SP-NAΔ3-1E-CWA were constructed. The plasmids were respectively electrotransformed into L. lactis NZ9000 to generate strain of L. lactis/pTX8048-SP-Δ3-1E in which 3-1E protein was expressed in secretion, and L. lactis/pTX8048-SP-NAΔ3-1E-CWA on which 3-1E protein was covalently anchored to the surface of bacteria cells. The expression of target proteins were examined by Western blot. The live lactococci expressing secreted 3-1E protein, anchored 3-1E protein, and cytoplasmic 3-1E protein was administered orally to chickens respectively, and the protective immunity and efficacy were compared by animal experiment. The results showed oral immunization to chickens with recombinant lactococci expressing anchored 3-1E protein elicited high 3-1E-specific serum IgG, increased high proportion of CD4 and CD8α cells in spleen, alleviated average lesion score in cecum, decreased the oocyst output per chicken compared to lactococci expressing cytoplasmic or secreted 3-1E protein. Taken together, these findings indicated the surface anchored Eimeria protein displayed by L. lacits can induce protective immunity and partial protection against homologous infection.
构建了两种新型质粒pTX8048-SP-Δ3-1E和pTX8048-SP-NAΔ3-1E-CWA。将这些质粒分别电转化到乳酸乳球菌NZ9000中，以产生分泌表达3-1E蛋白的乳酸乳球菌/pTX8048-SP-Δ3-1E菌株，以及3-1E蛋白共价锚定在细菌细胞表面的乳酸乳球菌/pTX8048-SP-NAΔ3-1E-CWA菌株。通过蛋白质免疫印迹法检测靶蛋白的表达。将表达分泌型3-1E蛋白、锚定型3-1E蛋白和细胞质3-1E蛋白的活乳酸乳球菌分别口服给鸡，并通过动物实验比较其保护性免疫和效果。结果表明，用表达锚定型3-1E蛋白重组乳酸乳球菌对鸡进行口服免疫，可诱导产生高水平的3-1E特异性血清IgG，增加脾脏中CD4和CD8α细胞的高比例，减轻盲肠平均病变评分，与表达细胞质或分泌型3-1E蛋白的乳酸乳球菌相比，降低每只鸡的卵囊产量。综上所述，这些发现表明乳酸乳球菌展示的表面锚定艾美耳球虫蛋白可诱导保护性免疫并对同源感染提供部分保护。
