本试验旨在研究枯草芽孢杆菌(Bacillus subtilis)对免疫和未免疫黄河鲤免疫功能和抗病力的影响。选择初始平均体重为(35.36±0.46) g的黄河鲤1 920尾,随机分为8组(Ⅰ~Ⅷ组),每组3个重复,每个重复80尾。Ⅰ~Ⅳ组为注射抗原的免疫组,Ⅴ~Ⅷ组为注射无菌生理盐水的对照组。试验饲料中枯草芽孢杆菌的添加量分别为0(Ⅰ、Ⅴ组)、1.5×1011(Ⅱ、Ⅵ组)、3.0×1011(Ⅲ、Ⅶ组)、4.5×1011 CFU/kg(Ⅳ、Ⅷ组)。各组试验鱼在投喂试验饲料28 d后,改喂基础饲料28 d。于免疫后第7、14、21、28、35、42天测定各组试验鱼的白细胞吞噬活性、血清溶菌酶活性和血清抗体效价。于免疫后第42天采用嗜水气单孢菌对各组试验鱼进行攻毒,14 d后统计各组的死亡率,计算免疫保护率。结果表明:饲料中添加枯草芽孢杆菌能显著提高免疫和未免疫黄河鲤的白细胞吞噬百分比和吞噬指数以及血清溶菌酶活性(P<0.05)。Ⅲ组黄河鲤的血清抗体效价最高,其几何平均值为486.7;其次是Ⅳ组,为372.6;再次是Ⅱ组,为234.6;而Ⅰ组最低,为192.8。免疫组黄河鲤均获得了一定的免疫保护率,以枯草芽孢杆菌添加量为4.5×1011 CFU/kg时的免疫保护率最高。由此得出,在本试验条件下,饲料中添加枯草芽孢杆菌可提高免疫和未免疫黄河鲤的免疫功能和抗病力,建议添加量为3.0×1011~4.5×1011 CFU/kg。
This experiment was conducted to study the effects of Bacillus subtilis on immune function and disease resistance of immunized and unimmunized Huanghe carp (Cyprinus carpio Huanghe var.). A total of 1 920 Huanghe carp with an average initial body weight of (35.36±0.46) g were randomly divided into 8 groups (groups Ⅰ to Ⅷ) with 3 replicates per group and 80 fish per replicate. The groups Ⅰ to Ⅳ were immunized groups which were injected antigen, and the groups Ⅴ to Ⅷ were control groups which were injected stroke-physiological saline solution. The supplemental levels of Bacillus subtilis in the experimental diets were 0 (groups Ⅰ and Ⅴ), 1.5×1011 (groups Ⅱ and Ⅵ), 3.0×1011 (groups Ⅲ and Ⅶ) and 4.5×1011 CFU/kg (groups Ⅳ and Ⅷ), respectively. The experimental fish were fed the experimental diets for 28 days, and subsequently fed the basal diet for 28 days. The leukocytes phagocytic activity, serum lysozyme activity and serum antibody titer were determined on the 7th, 14th, 21st, 28th, 35th and 42nd day after immunization. The experimental fish were challenged by Aeromonas hydrophila on the 42nd day after immunization, and the mortality of each group was counted for the calculation of immune protection rate. The results showed as follows: Bacillus subtilis supplementation could significantly improve the leukocytes phagocytic percentage and phagocytic index, and serum lysozyme activity of immunized and unimmunized Huanghe carp (P<0.05). The serum antibody titer of Huanghe carp in group Ⅲ was the highest, which was 486.7; the second was group Ⅳ, which was 372.6; the third was group Ⅱ, which was 234.6; the last one was group Ⅰ, which was 192.8. The Huanghe carp in immunized groups all had a certain immune protection rate, and the supplementation of 4.5×1011 CFU/kg Bacillus subtils induced the highest immune protection rate of immunized Huanghe carp. It is concluded that the supplementation of Bacillus subtilis can improve the immune function and disease resistance of immunized and unimmunized Huanghe carp, and the supplemental level of Bacillus subtilis in diets is suggested to be 3.0×1011 to 4.5×1011 CFU/kg.
[1] SHEN W Y,FU L L,LI W F,et al.Effect of dietary supplementation with Bacillus subtilis on the growth,performance,immune response and antioxidant activities of the shrimp (Litopenaeus vannamei)[J].Aquaculture Research,2010,41:1691-1698.
[2] ZIAEI-NEJAD S,REZAEI M H,TAKAMI G A,et al.The effect of Bacillus sp. bacteria used as probiotics on digestive enzyme activity,survival and growth in the Indian white shrimp Fenneropenaeus indicu[J].Aquaculture,2006,252:516-524.
[3] NAYAK S K,SWAIN P,MUKHERJEE S C.Effect of dietary supplementation of probiotic and vitamin C on the immune response of Indian major carp,Labeo rohita[J].Fish & Shellfish Immunology,2007,23(4):892-896.
[4] 郭玉娟,陈学年.肽聚糖对鲫鱼嗜水气单胞菌灭活疫苗免疫增强效果的研究[J].中国兽医科学,2006,36(1):33-36.
[5] 孟小亮,陈昌福,吴志新,等.嗜水气单胞菌3种疫苗对斑点叉尾 免疫原性比较研究[J].淡水渔业,2009,39(14):49-53.
[6] AMAR E C,KIRON V,SATOH S,et al.Enhancement of innate immunity in rainbow trout (Oncorhynchus mykiss Walbaum) associated with dietary intake of carotenoids from natural products[J].Fish & Shellfish Immunology,2004,16:527-537.
[7] ZHOU X,TIAN Z,WANG Y,et al.Effect of treatment with probiotics as water additives on tilapia (Oreochromis niloticus) growth performance and immune response[J].Fish Physiology and Biochemistry,2010,36:501-509.
[8] 大野尚仁.βグルカンの生体防御系修饰作用[J].日本细菌学杂志,2000,55(3):527-537.
[9] ALY S M,ABDEL-GALIL A Y,ABDEL-AZIZ G A,et al.Studies on Bacillus subtilis and Lactobacillus acidophilus,as potential probiotics,on the immune response and resistance of Tilapia nilotica (Oreochromis niloticus) to challenge infections[J].Fish & Shellfish Immunology,2008,25:128-136.
[10] YANG H L,SUN Y Z,MA R L,et al.PCR-DGGE analysis of the autochthonous gut microbiota of grouper Epinephelus coioides following probiotic Bacillus clausii administration[J].Aquaculture Research,2011.doi:10.1111/j.1365-2109.2011.02852.X.
[11] NEWAJ-FYZUL A,ADESIYUNA A,MUTANI A,et al.Bacillus subtilis AB1 controls Aeromonas infection in rainbow trout (Oncorhynchus mykiss Walbaum)[J].Journal of Applied Microbiology,2007,103(5):1669-1706.
[12] MARJA M,ANTTI S.Changes in plasma lysozyme and blood leucocyte levels of hatchery-reared Atlantic salmon and sea trout (Salmo tuotta L) during parr-smolt transformation[J].Aquaculture,1992,106:75-87.
[13] 潘康成,杨汉博.饲用芽孢菌作用机理的研究进展[J].饲料工业,1997,18(9):32-34.
[14] TEJADA-SIMON M V,PESTKA J J.Proinflammatory cytokine and nitric oxide induction in murine macrophages by cell wall and cytoplasmic extracts of lactic acid bacteria[J].Journal of Food Protection,l999,62(12):1435-1444.
[15] ANDERSON D P.Immunostimulants,adjuvants,and vaccine in fish:applications to aquaculture[J].Annual Review of Fish Diseases,1992,2:281-307.
[16] HIMANEN J P,SARVAS M,HELANDER I M.Assessment of non-protein impurities in potential vaccine proteins produced by Bacillus subtilis[J].Vaccine,1993,11(9):970-973.
[17] 安利国,冯程强,邢维贤,等.灭活疫苗对鲤鱼血清溶菌酶和腹腔吞噬细胞活性的作用[J].山东师范大学学报:自然科学版,1999,l4(2):175-179.
[18] RAJESH K,MUKHERJEE S C,RITESH R,et al.Enhanced innate immune parameters in Labeo rohita (Ham.) following oral administration of Bacillus subtilis[J].Fish & Shellfish Immunology,2008,24(2):168-172.
[19] MERRIFIELD D L,DIMITROGLOU A,BRADLEY G,et al.Probiotic applications for rainbow trout (Oncorhynchus mykiss Walbaum):Ⅱ.Effects on growth performance,feed utilization,intestinal microbiota and related health criteria postantibiotic treatment[J].Aquaculture Nutrition,2010,16(5):496-503.
[20] NIKL L,ALBRIGHT L J,EVELYN T P T.Influence of seven immunostimulants on the immune responses of coho salmon to Aeromonas salmonicida[J].Diseases Aquactic Organisms,1991,12:7-12.
[21] 孙建和,严亚贤,陈怀青.嗜水气单胞菌亚单位疫苗的研制[J].中国兽医学报,1996,16(1):11-15.
[22] MICHEL C B,FAIVTE B.Occurrence and significance of agglutinating antibodies in experimental forunculosis of rainbow trout,Salmo gairdneri[J].Journal of Fish Diseases,1982,5:429-432.
[23] POST G.Response of rainbow trout to antigens of Aeromonas hydrophila[J].Journal of the Fisheries Research Board of Canada,1966,23:1487-1494.
[24] SCHACTE J H.Studies on the immunization of the channel catfish against two bacterial pathogens.Ph.D.Thesis.Auburn,A.L.:Auburn University,1976:81-89.
[25] SCHROERS V,VAN DER MAREL M,NEUHAUS H,et al.Changes of intestinal mucus glycoproteins after peroral application of Aeromonas hydrophila to common carp (Cyprinus carpio)[J].Aquaculture,2009,288(3/4):184-189.
[26] ARDÓ L,JENEY Z,ADAM A,et al.Immune responses of resistant and sensitive common carp families following experimental challenge with Aeromonas hydrophila[J].Fish & Shellfish Immunology,2010,29(1):111-116.
京公网安备 11010802041926号