饲料中添加益生菌对凡纳滨对虾抗感染和5种免疫基因表达的影响

为研究饲料中添加益生菌对凡纳滨对虾抗副溶血弧菌感染能力和非特异性免疫基因表达水平的影响,以初体质量为(6.95±1.20)g的凡纳滨对虾为研究对象,在室内养殖箱进行3周的养殖实验和2周的副溶血弧菌人工感染实验;其中,对照组每日投喂普通商品饲料,实验组每日投喂在普通商品饲料中添加地衣芽孢杆菌、枯草芽孢杆菌、地衣芽孢杆菌/枯草芽孢杆菌(1:1)配制成的3组实验饲料,实验饲料中益生菌的终浓度为10⁷ cfu/g。并采用实时荧光定量RT-PCR方法,对保护率最高的地衣芽孢杆菌/枯草芽孢杆菌实验组进行凡纳滨对虾相关免疫基因表达水平的分析。实验结果表明,在饲料中添加单一益生菌或复合益生菌均可显著提高对虾抗副溶血弧菌感染的能力(P<0.05),且复合益生菌的保护效果更佳,其相对免疫保护率为31.11%。感染副溶血弧菌后,地衣芽孢杆菌+枯草芽孢杆菌实验组凡纳滨对虾血淋巴中的先天免疫缺陷基因(innate immune deficiency gene,IMD)、对虾素3a分子(penaiedin 3a)、酚氧化酶原(prophenoloxidase,proPO)、溶菌酶(Lysozyme,LZM)和甲壳素Crustin的mRNA的相对表达量均显著上调,且分别在18～24 h达到最大值。实验结果提示:饲料中添加芽孢杆菌可有效提高凡纳滨对虾抗副溶血弧菌感染的能力,这种能力的提高可能是通过增加抗病相关基因的表达量实现的。

Effects of adding probiotics to the feed on anti-infection and five kinds of immune gene expression of Litopenaeus vannamei

A 3-week feeding experiment and a 2-week Vibrio parahaemolytics infection experiment were conducted to study the effects of adding probiotics to the feed on non-specific immune gene expression and resistance to V.parahaemolytics of L.vannameiinitial body weight(6.95±1.20)g].Litopenaeus vannamei was fed four different diets:Control group(commercial fodder)and experimental group(feed supplemented with Bacillus licheniformis;or with Bacillus subtilis and with mixed diet comprising Bacillus licheniformis/Bacillus subtilis at the ratio of 1:1),the final concentration of probiotics was 10⁷ cfu/g.And real-time PCR method was used to study the non-specific immune gene expression of L.vannamei in Bacillus licheniformis/Bacillus subtilis experimental group,which is the highest protection group after being infected with V.parahaemolytics.The results showed that all probiotics groups can effectively improve the ability of shrimp to resist the V.parahaemolytics's infection,and compound probiotics can better resist the V.parahaemolytics infections,its relative protection ratio is 31.11%.After infection with the V.parahaemolytics,the relative expressions of IMD,penaiedin 3a,proPO,LZM and Crustin were obviously up-regulated in hemolymph of shrimp fed diets supplemented with the compound probiotics(Bacillus licheniformis/Bacillus subtilis)compared to the control during the immunization.In hemolymph,the relative expression of immune deficiency gene,penaiedin 3a,prophenoloxidase,Lysozyme and Crustin were increased to the maximum levels at 18-24 h.The experimental result indicates:adding probiotics to the feed can effectively improve the ability of L.vannamei to resist the V.parahaemolytics infection,and the improvement of this ability may be made through enhancing the disease resistance related gene expression.