CN109198268A - 鳜鱼抗病中草药与卵黄抗体复合添加剂及投喂方法 - Google Patents
鳜鱼抗病中草药与卵黄抗体复合添加剂及投喂方法 Download PDFInfo
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Abstract
本发明是一种鳜鱼抗病中草药与卵黄抗体复合添加剂,该添加剂由抗鳜鱼细菌性败血症卵黄抗体、3味中草药浸出液及β‑环糊精;其中抗鳜鱼细菌性败血症卵黄抗体,主要是使鳜鱼降低或免受强致病性的嗜水气单胞菌的感染;3味中草药浸出液中的大黄可以抑制或杀灭病原嗜水气单胞菌,具有使鳜鱼降低或免受强致病性的嗜水气单胞菌的感染的作用,女贞子具有增强免疫力的作用,干草具有解毒作用;β‑环糊精主要是使卵黄抗体和中草药浸出液混合物起到粘合及缓释作用。本发明能预防与治疗鳜鱼养殖期间由病原嗜水气单胞菌引起的败血症,增加鳜鱼养殖产量,显著地提高鳜鱼养殖的经济效益。解决目前鳜鱼养殖生产过程中易发生细菌性败血症的问题。
Description
技术领域
本发明涉及一种水产养殖用饲料添加剂,特别涉及一种用于抗鳜鱼细菌性败血症的饲料添加剂。
背景技术
翘嘴鳜(Siniperca chuatsi)是淡水凶猛肉食性鱼类,生长速度快,因其肉质细嫩、营养含量高而深受欢迎,目前鳜养殖已成为我国水产养殖业一大特色。随着养殖集约化程度的增高,养殖密度的增大以及养殖水环境的恶化,鳜鱼的病害问题日益突出,目前已成为威胁我国鳜鱼养殖业健康发展的主要因素。这些病害中,嗜水气单胞菌引起的细菌性败血症等疾病危害较为严重,给养殖户带来了巨大的经济损失,严重制约了鳜鱼养殖业的健康稳定发展。因此,鳜鱼细菌性败血症防控技术研究,是产业发展的重大技术亟需。
目前鳜鱼养殖生产中很难控制由嗜水气单胞菌引起的疾病的发生与流行,尤其是鳜鱼以活鱼(异育银鲫及赤眼鳟等)为饵料,药物及免疫制剂等不能通过配合饲料投喂。因此,开发研制鳜鱼抗细菌性败血症饲料添加剂,并通过投喂饵料鱼将药物及免疫制剂带入鳜鱼体内,对于有效控制鳜鱼养殖生产中败血症的发生与流行具有重要的应用前景。
发明内容
本发明所要解决的技术问题是针对现有技术的不足,提供一种提高鳜鱼抗病力、降低鳜鱼养殖生产中由病原嗜水气单胞菌所造成的危害、以满足鳜鱼养殖生产上的抗病需求的饲料添加剂。
本发明所要解决的另一个技术问题是提供了前述复合添加剂的制备方法。
本发明所要解决的再一个技术问题是提供了前述复合添加剂的投喂方法。
本发明所要解决的又一个技术问题是提供了前述复合添加剂在制备鳜鱼抗病养殖饲料(特别是鳜鱼抗细菌性败血症饲料)中的用途。
本发明所要解决的技术问题是通过以下的技术方案来实现的。本发明是一种一种鳜鱼抗病中草药与卵黄抗体复合添加剂,其特点是,所述的复合添加剂由以下重量配比的三种成分组成,
(1)抗鳜鱼细菌性败血症卵黄抗体液 1;
(2)3味中草药浸出液 1-3;
(3)β-环糊精 适量;
所述的抗鳜鱼细菌性败血症卵黄抗体液的制备方法如下:
①将引起鳜鱼细菌性败血症的强致病性的病原嗜水气单胞菌 (Aeromonas hydrophila ) G3菌株分别接种在普通营养肉汤培养基上,28-30℃摇床培养24h,加入体积终浓度0.3%的甲醛28℃灭活48h,离心得灭活菌体;
②取黄芪用蒸馏水浸泡30-60min后煎煮,过滤,浓缩至终浓度为原药1g/mL,得黄芪浸出液;
③将灭活菌体与黄芪浸出液混合,制得水剂型疫苗;疫苗液中病原嗜水气单胞菌G3免疫原达到109cells/mL,黄芪浸出液为1g/mL;
④用上述制得的病原嗜水气单胞菌水剂型灭活疫苗,采用口服及滴鼻免疫的方法免疫成年产蛋鸡,第1次免疫剂量为:口服免疫是将疫苗液加入到饮水中,加入量为5 mL;滴鼻免疫是将疫苗液滴入蛋鸡鼻腔,滴入量为1 mL;一周后按前述方法加强免疫,连续加强免疫5次,共免疫6次,免疫结束后第6周,卵黄抗体效价达212-214;收集被免疫鸡所产鸡蛋,分离出卵黄,即得抗鳜鱼细菌性败血症卵黄抗体液;
所述的3味中草药浸出液的制备方法如下:将重量配比为2-4:0.8-1.2:1的大黄、女贞子和甘草加入蒸馏水,置水浴煮沸20-40min,取上清液;再于中草药中加入蒸馏水,二次水浴煮沸20-40min后取上清液;两次的上清液混合、并补足蒸馏水,3味中草药终浓度为0.2g/mL,制得3味中草药浸出液。
以上所述的本发明鳜鱼抗病中草药与卵黄抗体复合添加剂中,进一步优选的技术方案是:所述的复合添加剂中,抗鳜鱼细菌性败血症卵黄抗体液与3味中草药浸出液的重量比为1:2;所述的β-环糊精占复合添加剂总重量的5~8%。
以上所述的本发明鳜鱼抗病中草药与卵黄抗体复合添加剂中,进一步优选的技术方案是:所述的3味中草药浸出液的具体制备方法中:原料大黄、女贞子和甘草将的重量配比为3:1:1。
本发明还提供了一种如以上技术方案所述的鳜鱼抗病中草药与卵黄抗体复合添加剂的制备方法,其特点是,其步骤如下:按所述重量配比取抗鳜鱼细菌性败血症卵黄抗体液,加入3味中草药浸出液稀释,加入β-环糊精,喷雾干燥制粉,喷雾干燥时喷口温度设置为140℃,制得中草药与卵黄抗体喷雾微胶囊干粉。
本发明还提供了一种鳜鱼抗病中草药与卵黄抗体复合添加剂投喂方法,其特点是,应用以上技术方案所述的复合添加剂;其使用步骤如下:先将鳜鱼抗病中草药与卵黄抗体复合添加剂以5%-10%(进一步优选8%)的添加量添加至异育银鲫及赤眼鳟配合饲料中,再用前述配合饲料投喂异育银鲫及赤眼鳟;待异育银鲫及赤眼鳟食用配合饲料2h后,将其投喂给鳜鱼作饵料鱼。
本发明所述的鳜鱼强致病性的病原嗜水气单胞菌G3菌株已经在公开文献《翘嘴鳜病原嗜水气单胞菌分子特征及LAMP检测方法的建立》中所公开(菌株G3),上述公开文献发表于《水生生物学报》,2017,41(6):1225-1231。菌株G3在Genebank 中16S rRNA基因登录号为KX822740,gyrB基因登录号为KX822741。
菌株G3为公知公用材料,现保存在扬州大学动物科学与技术学院水产动物病害实验室,在本专利申请日起二十年内,公众如果需要,扬州大学动物科学与技术学院水产动物病害实验室可对外发放。
本发明鳜鱼抗病中草药与卵黄抗体复合添加剂,不仅具有良好的抗病作用,而且使用方便,同时又具有较高的营养价值。主要是使鳜鱼降低或免受强致病性的嗜水气单胞菌的感染;3味中草药浸出液中的大黄可以抑制或杀灭病原嗜水气单胞菌,具有使鳜鱼降低或免受强致病性的嗜水气单胞菌的感染的作用,女贞子具有增强免疫力的作用,干草具有解毒作用;β-环糊精主要是使卵黄抗体和中草药浸出液混合物起到粘合及缓释作用,各原料协同增效,大大地增强了鳜鱼对嗜水气单胞菌的抵抗力,提高鳜鱼的成活率,增加鳜鱼养殖生产的经济效益,以解决目前鳜鱼养殖生产过程中易发生由嗜水气单胞菌引起的细菌性败血症的问题。
具体实施方式
以下对本发明的具体实施方式进行详细描述,以便于本领域的技术人员进一步地理解本发明,而不构成对本发明权利的限制。
实施例1,一种鳜鱼抗病中草药与卵黄抗体复合添加剂,所述的复合添加剂由以下重量配比的三种成分组成,
(1)抗鳜鱼细菌性败血症卵黄抗体液 1;
(2)3味中草药浸出液 1;
(3)β-环糊精占复合添加剂总重量的5%;
所述的抗鳜鱼细菌性败血症卵黄抗体液的制备方法如下:
①将引起鳜鱼细菌性败血症的强致病性的病原嗜水气单胞菌 (Aeromonas hydrophila ) G3菌株分别接种在普通营养肉汤培养基上,28℃摇床培养24h,加入体积终浓度0.3%的甲醛28℃灭活48h,离心得灭活菌体;
②取黄芪用蒸馏水浸泡30min后煎煮,过滤,浓缩至终浓度为原药1g/mL,得黄芪浸出液;
③将灭活菌体与黄芪浸出液混合,制得水剂型疫苗;疫苗液中病原嗜水气单胞菌G3免疫原达到109cells/mL,黄芪浸出液为1g/mL;
④用上述制得的病原嗜水气单胞菌水剂型灭活疫苗,采用口服及滴鼻免疫的方法免疫成年产蛋鸡,第1次免疫剂量为:口服免疫是将疫苗液加入到饮水中,加入量为5 mL;滴鼻免疫是将疫苗液滴入蛋鸡鼻腔,滴入量为1 mL;一周后按前述方法加强免疫,连续加强免疫5次,共免疫6次,免疫结束后第6周,卵黄抗体效价可达212-214;收集被免疫鸡所产鸡蛋,分离出卵黄,即得抗鳜鱼细菌性败血症卵黄抗体液;
所述的3味中草药浸出液的制备方法如下:将重量配比为2:0.8:1的大黄、女贞子和甘草加入蒸馏水,置水浴煮沸20min,取上清液;再于中草药中加入蒸馏水,二次水浴煮沸20min后取上清液;两次的上清液混合、并补足蒸馏水,3味中草药终浓度为0.2g/mL,制得3味中草药浸出液。
实施例2,一种鳜鱼抗病中草药与卵黄抗体复合添加剂,所述的复合添加剂由以下重量配比的三种成分组成,
(1)抗鳜鱼细菌性败血症卵黄抗体液 1;
(2)3味中草药浸出液 3;
(3)β-环糊精占复合添加剂总重量的8%;
所述的抗鳜鱼细菌性败血症卵黄抗体液的制备方法如下:
①将引起鳜鱼细菌性败血症的强致病性的病原嗜水气单胞菌 (Aeromonas hydrophila ) G3菌株分别接种在普通营养肉汤培养基上, 30℃摇床培养24h,加入体积终浓度0.3%的甲醛28℃灭活48h,离心得灭活菌体;
②取黄芪用蒸馏水浸泡60min后煎煮,过滤,浓缩至终浓度为原药1g/mL,得黄芪浸出液;
③将灭活菌体与黄芪浸出液混合,制得水剂型疫苗;疫苗液中病原嗜水气单胞菌G3免疫原达到109cells/mL,黄芪浸出液为1g/mL;
④用上述制得的病原嗜水气单胞菌水剂型灭活疫苗,采用口服及滴鼻免疫的方法免疫成年产蛋鸡,第1次免疫剂量为:口服免疫是将疫苗液加入到饮水中,加入量为5 mL;滴鼻免疫是将疫苗液滴入蛋鸡鼻腔,滴入量为1 mL;一周后按前述方法加强免疫,连续加强免疫5次,共免疫6次,免疫结束后第6周,卵黄抗体效价可达212-214;收集被免疫鸡所产鸡蛋,分离出卵黄,即得抗鳜鱼细菌性败血症卵黄抗体液;
所述的3味中草药浸出液的制备方法如下:将重量配比为4: 1.2:1的大黄、女贞子和甘草加入蒸馏水,置水浴煮沸40min,取上清液;再于中草药中加入蒸馏水,二次水浴煮沸40min后取上清液;两次的上清液混合、并补足蒸馏水,3味中草药终浓度为0.2g/mL,制得3味中草药浸出液。
实施例3,一种鳜鱼抗病中草药与卵黄抗体复合添加剂,所述的复合添加剂由以下重量配比的三种成分组成,
(1)抗鳜鱼细菌性败血症卵黄抗体液 1;
(2)3味中草药浸出液 2;
(3)β-环糊精占复合添加剂总重量的6%;
所述的抗鳜鱼细菌性败血症卵黄抗体液的制备方法如下:
①将引起鳜鱼细菌性败血症的强致病性的病原嗜水气单胞菌 (Aeromonas hydrophila ) G3菌株分别接种在普通营养肉汤培养基上,29℃摇床培养24h,加入体积终浓度0.3%的甲醛28℃灭活48h,离心得灭活菌体;
②取黄芪用蒸馏水浸泡45min后煎煮,过滤,浓缩至终浓度为原药1g/mL,得黄芪浸出液;
③将灭活菌体与黄芪浸出液混合,制得水剂型疫苗;疫苗液中病原嗜水气单胞菌G3免疫原达到109cells/mL,黄芪浸出液为1g/mL;
④用上述制得的病原嗜水气单胞菌水剂型灭活疫苗,采用口服及滴鼻免疫的方法免疫成年产蛋鸡,第1次免疫剂量为:口服免疫是将疫苗液加入到饮水中,加入量为5 mL;滴鼻免疫是将疫苗液滴入蛋鸡鼻腔,滴入量为1 mL;一周后按前述方法加强免疫,连续加强免疫5次,共免疫6次,免疫结束后第6周,卵黄抗体效价可达212-214;收集被免疫鸡所产鸡蛋,分离出卵黄,即得抗鳜鱼细菌性败血症卵黄抗体液;
所述的3味中草药浸出液的制备方法如下:将重量配比为3:3:1的大黄、女贞子和甘草加入蒸馏水,置水浴煮沸30min,取上清液;再于中草药中加入蒸馏水,二次水浴煮沸30min后取上清液;两次的上清液混合、并补足蒸馏水,3味中草药终浓度为0.2g/mL,制得3味中草药浸出液。
其制备方法步骤如下:按所述重量配比取抗鳜鱼细菌性败血症卵黄抗体液,加入3味中草药浸出液稀释,加入β-环糊精,喷雾干燥制粉,喷雾干燥时喷口温度设置为140℃,制得中草药与卵黄抗体喷雾微胶囊干粉。
其投喂方法步骤如下:先将鳜鱼抗病中草药与卵黄抗体复合添加剂以8%的添加量添加至异育银鲫及赤眼鳟配合饲料中,再用前述配合饲料投喂异育银鲫及赤眼鳟;待异育银鲫及赤眼鳟食用配合饲料2h后,将其投喂给鳜鱼作饵料鱼。
实施例4,实施例3制得的鳜鱼抗病中草药与卵黄抗体复合添加剂的应用实验及其结果。
实验时间:2017年度8月。
实验地点:江苏省扬州大学水产动物病害实验室与养殖温室。
实验动物:赤眼鳟与翘嘴鳜。
(1)使用实施例3制得的鳜鱼抗病中草药及卵黄抗体复合添加剂微胶囊干粉。
(2)实验组赤眼鳟(100尾)投喂本发明鳜鱼抗病中草药与卵黄抗体复合添加剂,按8%的比例添加在赤眼鳟配合饲料,对照组赤眼鳟(100尾)投喂市售赤眼鳟配合饲料。
(3)实验组和对照组赤眼鳟投喂2h后,实验组和对照组赤眼鳟分别投喂翘嘴鳜(40尾)。
(4)投喂24h后,对投喂实验组赤眼鳟和对照组赤眼鳟的翘嘴鳜进行保护力检验,分别取翘嘴鳜20尾,用嗜水气单胞菌G3进行浸浴感染,水体中菌体终浓度均达到106CFU/mL。试验期间继续按上述(2)和(3)进行投喂,试验期为2周。
(5)实验结果:实验组赤眼鳟投喂实施例3所述的鳜鱼抗病中草药与卵黄抗体复合添加剂(8%)后,再投喂给鳜鱼,可使鳜鱼获得较高的抗病能力。实验翘嘴鳜用嗜水气单胞菌G3菌液进行浸浴感染,投喂实施例3所述的鳜鱼抗病中草药与卵黄抗体复合添加剂(8%)及市售赤眼鳟配合饲料(92%)的实验组,翘嘴鳜感染2周后存活率达76%,投喂对照组赤眼鳟的翘嘴鳜存活率仅为28%,投喂实验组赤眼鳟(鳜鱼抗病中草药与卵黄抗体复合添加剂)的翘嘴鳜成活率比对照组高48%。实验结果表明鳜鱼抗病中草药与卵黄抗体复合添加剂通过投喂给饵料鱼可翘嘴鳜获得较高的免疫保护力。
Claims (7)
1.一种鳜鱼抗病中草药与卵黄抗体复合添加剂,其特征在于,所述的复合添加剂由以下重量配比的三种组分组成:
(1)抗鳜鱼细菌性败血症卵黄抗体液 1;
(2)3味中草药浸出液 1-3;
(3)β-环糊精 适量;
所述的抗鳜鱼细菌性败血症卵黄抗体液的制备方法如下:
①将引起鳜鱼细菌性败血症的强致病性的病原嗜水气单胞菌 (Aeromonas hydrophila ) G3菌株分别接种在普通营养肉汤培养基上,28-30℃摇床培养24h,加入体积终浓度0.3%的甲醛28℃灭活48h,离心得灭活菌体;
②取黄芪用蒸馏水浸泡30-60min后煎煮,过滤,浓缩至终浓度为原药1g/mL,得黄芪浸出液;
③将灭活菌体与黄芪浸出液混合,制得水剂型疫苗;疫苗液中病原嗜水气单胞菌G3免疫原达到109cells/mL,黄芪浸出液为1g/mL;
④用上述制得的病原嗜水气单胞菌水剂型灭活疫苗,采用口服及滴鼻免疫的方法免疫成年产蛋鸡,第1次免疫剂量为:口服免疫是将疫苗液加入到饮水中,加入量为5 mL;滴鼻免疫是将疫苗液滴入蛋鸡鼻腔,滴入量为1 mL;一周后按前述方法加强免疫,连续加强免疫5次,共免疫6次,免疫结束后第6周,卵黄抗体效价达212-214;收集被免疫鸡所产鸡蛋,分离出卵黄,即得抗鳜鱼细菌性败血症卵黄抗体液;
所述的3味中草药浸出液的制备方法如下:将重量配比为2-4:0.8-1.2:1的大黄、女贞子和甘草加入蒸馏水,置水浴煮沸20-40min,取上清液;再于中草药中加入蒸馏水,二次水浴煮沸20-40min后取上清液;两次的上清液混合、并补足蒸馏水,3味中草药终浓度为0.2g/mL,制得3味中草药浸出液。
2.根据权利要求1所述的鳜鱼抗病中草药与卵黄抗体复合添加剂,其特征在于:所述的复合添加剂中,抗鳜鱼细菌性败血症卵黄抗体液与3味中草药浸出液的重量比为1:2;所述的β-环糊精占复合添加剂总重量的5~8%。
3.根据权利要求1所述的鳜鱼抗病中草药与卵黄抗体复合添加剂,其特征在于:所述的3味中草药浸出液的具体制备方法中:原料大黄、女贞子和甘草将的重量配比为3:1:1。
4.一种如权利要求1或2或3所述的鳜鱼抗病中草药与卵黄抗体复合添加剂的制备方法,其特征在于,其步骤如下:按所述重量配比取抗鳜鱼细菌性败血症卵黄抗体液,加入3味中草药浸出液稀释,加入β-环糊精,喷雾干燥制粉,喷雾干燥时喷口温度设置为140℃,制得中草药与卵黄抗体喷雾微胶囊干粉。
5.一种鳜鱼抗病中草药与卵黄抗体复合添加剂投喂方法,其特征在于,应用权利要求1或2或3所述的复合添加剂;其使用步骤如下:先将鳜鱼抗病中草药与卵黄抗体复合添加剂以5%-10%的添加量添加至异育银鲫及赤眼鳟配合饲料中,再用前述配合饲料投喂异育银鲫及赤眼鳟;待异育银鲫及赤眼鳟食用配合饲料2h后,将其投喂给鳜鱼作饵料鱼。
6.根据权利要求5所述的鳜鱼抗病中草药与卵黄抗体复合添加剂投喂方法,其特征在于,复合添加剂以8%的添加量添加至异育银鲫及赤眼鳟配合饲料中。
7.一种如权利要求1或2或3所述的复合添加剂的用途,其特征在于,所述的用途为复合添加剂在制备鳜鱼抗病养殖饲料中的用途。
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