CAJ | 학술논문

以枯草芽孢杆菌(Bacillus subtilis)为活载体口服递呈对虾白斑综合征病毒(WSSV)囊膜蛋白 VP28,评价其抗病毒感染能力、对南美白对虾免疫相关基因表达以及血淋巴细胞对病毒特异性吞噬的影响。经口服免疫枯草重组菌株B. subtilis-VP28攻毒后,对虾的相对存活率达83.3%。为探讨重组菌株的抗病机理,比较研究了免疫相关基因-proPO(酚氧化酶原)、Peroxinectin(PE)和脂多糖-β-1,3-葡聚糖结合蛋白(LGBP)基因的表达差异,并进一步分析了血淋巴细胞吞噬活性和特异性。结果表明, B. subtilis-VP28菌液能显著提高(P<0.05)对虾proPO、PE和LGBP mRNA的表达水平和血细胞对WSSV的吞噬活性, B. subtilis组对免疫相关基因也有一定的激活作用,而B. subtilis-VP28发酵上清液则能增加血细胞吞噬活性；此外, B. subtilis-VP28菌液组血细胞对WSSV具有特异性吞噬作用。研究为枯草重组菌株B. subtilis-VP28抗WSSV感染作用及其作为特殊功能水产微生态制剂的应用提供了一定的科学依据。
이고초아포간균(Bacillus subtilis)위활재체구복체정대하백반종합정병독(WSSV)낭막단백 VP28,평개기항병독감염능력、대남미백대하면역상관기인표체이급혈림파세포대병독특이성탄서적영향。경구복면역고초중조균주B. subtilis-VP28공독후,대하적상대존활솔체83.3%。위탐토중조균주적항병궤리,비교연구료면역상관기인-proPO(분양화매원)、Peroxinectin(PE)화지다당-β-1,3-포취당결합단백(LGBP)기인적표체차이,병진일보분석료혈림파세포탄서활성화특이성。결과표명, B. subtilis-VP28균액능현저제고(P<0.05)대하proPO、PE화LGBP mRNA적표체수평화혈세포대WSSV적탄서활성, B. subtilis조대면역상관기인야유일정적격활작용,이B. subtilis-VP28발효상청액칙능증가혈세포탄서활성；차외, B. subtilis-VP28균액조혈세포대WSSV구유특이성탄서작용。연구위고초중조균주B. subtilis-VP28항WSSV감염작용급기작위특수공능수산미생태제제적응용제공료일정적과학의거。
The regulation of immune-related genes expression and phagocytosis of White Spot Syndrome Virus (WSSV) were evaluated by oral delivery of VP28 using Bacillus subtilis as vehicles in Litopenaeus vannamei. In our initial ex-periment, by oral delivery of B. subtilis spores harboring VP28 (B. subtilis-VP28) to L. vannamei, the extremely high survival (Relative Percent Survival:83.3%) upon challenge with WSSV can be observed. The differences of genes ex-pression levels of proPO, Peroxinectin (PE) and lipopolysaccharide-and beta-1, 3-glucan-binding protein (LGBP) were demonstrated among experimental groups of B. subtilis-VP28 bacterial spores, B. subtilis-VP28 supernatants, B. subtilis and control. The result showed that immune-related genes (proPO, PE and LGBP) were significantly (P<0.05) upregu-lated in both B. subtilis-VP28 bacterial spores and B. subtilis feeding groups compared to B. subtilis-VP28 supernatants and the control groups. The phagocytosis in groups of B. subtilis-VP28 bacterial spores and B. subtilis-VP28 super-natants were enhanced significantly (P<0.05). Besides, the percentage of haemocytes phagocytosing WSSV (46.8±3.9) in shrimp previously fed with B. subtilis-VP28 was significantly higher (P<0.001) than that of phagocytosing TSV (an unrelated virus of shrimp) (14.9±2.2). This suggests that the heightened phagocytic activity, and thus the high-level survival of shrimp after B. subtilis-VP28‘vaccination’ are selective or specific towards WSSV. These results may raise the application prospective of this recombinant B. subtilis spores against WSSV infection in shrimp farms.