蜂胶佐剂对异育银鲫嗜水气单胞菌灭活疫苗的免疫增强试验

本文就蜂胶佐剂对鱼类细菌性疫苗的免疫增强作用进行了首次评价。试验以异育银鲫嗜水气单胞菌灭活疫苗为对象，比较了蜂胶佐剂、铝胶佐剂和无佐剂三种疫苗对银鲫的免疫效果。经抗体监测表明，免疫接种后第2周，蜂胶佐剂组银鲫已检测到血清抗体，随后抗体效价迅速上升，第4周达到峰值2^5：铝胶佐剂组和无佐剂组第3周检测到抗体．铝胶佐剂组第5周达到峰值2^5，无佐剂组第4周达到峰值2^4。银鲫免疫后第5周进行攻毒保护试验，发现蜂胶佐剂组、铝胶佐剂组和无佐剂组的免疫保护率分别为77．3％．65．0％和56．3％。试验结果表明，蜂胶佐剂能有效增强异育银鲫细菌性灭活疫苗的免疫保护力。     

细菌性腹泻的疫苗

（1）常规疫苗可分为多价现地株灭活苗、多价菌毛亚单位苗两大类；按佐剂类型还可分为油乳剂灭活苗、蜂胶佐剂灭活苗、铝胶佐剂灭活苗和蜂胶中药佐荆灭活苗等多种剂型。以蜂胶和蜂胶中药佐剂灭活苗效果最佳。     

灭活菌苗免疫的中华倒刺鲃外周血免疫指标的变化

以温和气单胞菌灭活菌苗为免疫原,平均体重(100±25)g的健康中华倒刺鲃为实验对象,免疫组腹腔注射0.2mL浓度为1.0×108CFU/mL的免疫原,对照组注射等量灭菌生理盐水,分别在单次注射0、1、2、4、7、14、21、28、35d后随机从两组各取6尾实验鱼,尾静脉采血,测定外周血的血细胞数量、白细胞分类计数、吞噬活性、抗体效价和蛋白质含量等免疫指标的变化,第35天活菌攻毒。结果表明:温和气单胞菌灭活菌苗(F-AS)可诱导中华倒刺鲃红细胞和白细胞数量增加,并引起各种白细胞分类百分比变化,提高吞噬活性和抗体效价,血清中总蛋白及球蛋白含量增加,红细胞也具有免疫功能,灭活茵苗的相对免疫保护力达65.21%。免疫早期(第1周)主要是红细胞、单核细胞和中性粒细胞数量明显增加,吞噬细胞的吞噬活性迅速提高,吞噬百分比和吞噬指数第4天达峰值;随后则是淋巴细胞大量增殖,第21天淋巴细胞、抗体效价及球蛋白达峰值。可见灭活菌苗通过促进中华倒刺鲃血细胞增殖、提高吞噬细胞的吞噬活性、产生特异性抗体等方式提高免疫保护力;免疫早期非特异性细胞免疫起重要作用,之后特异性免疫起主要作用。     

海豚链球菌灭活疫苗对罗非鱼免疫效果的初步研究

将海豚链球菌(Streptococcus iniae)淡水分离株TBY-1菌株灭活后制备成无佐剂和添加白油佐剂的疫苗,利用浸泡和腹腔注射复合免疫方式对罗非鱼(Oreochromis niloticus)进行免疫,比较其相对保护率及血清中抗体凝集效价,以评价佐剂的存在与否及免疫次数对海豚链球菌灭活疫苗的免疫效果的影响,同...     

中华鳖对T3菌苗的回忆应答

研究证明用Ｔ３菌苗免疫的中华鳖具有回忆应答反应，它的回忆应答介于鱼类和哺乳类之间，具有以下特点：（１）回忆应答与初次应答的ＩＡＴ峰值差异明显，比率可达２左右；（２）抗体生成速度较快，第１６天左右ＩＡＴ可达峰值水平，ＩＡＴ峰值下降速度明显低于生成速度；（３）与初次应答相比，回忆应答抗体形成的速度较快，而消失的速度较慢。     

注射菌苗的生殖期黄颡鱼亲鱼的外周血细胞和体液免疫变化

阐明生殖期亲鱼在注射灭活菌苗后的血细胞和体液免疫变化规律，是预防亲鱼产后虚弱患病和仔鱼染病死亡的理论基础。本研究在试验组和对照组黄颡鱼胸鳍基部分别注射0.2 mL浓度为1.0×10⁸ CFU/mL福尔马林灭活温和气单胞菌苗（F-AS）和0.65%灭菌生理盐水，在免疫后第1、2、4、7、14、21、28、35天，从尾静脉采血，测定外周血液血细胞数量及组成比例，血细胞吞噬活性和抗体效价。结果显示：注射菌苗后白细胞数量和淋巴细胞的白细胞分类计数逐渐增加，显著高于对照；中性粒细胞、单核细胞的白细胞分类计数以及吞噬百分比（PP）、吞噬指数（PI）在4～7 d显著高于对照；抗体效价逐渐升高且在第14天达到最高，随后逐渐下降；而红细胞数和血栓细胞的白细胞分类计数逐渐下降，显著低于对照组。结果表明：单次注射菌苗后亲鱼可以产生良好的免疫应答，1～7 d以增强单核细胞和中性粒细胞等非特异细胞免疫为主；7～35 d淋巴细胞增值，并大量释放抗体且在第21天淋巴细胞出现极显著升高，此后虽逐渐下降但仍显著高于对照，抗体维持在较高水平上的时间为1周左右，建议在黄颡鱼催产前10～14 d进行单次菌苗注射以增强亲鱼免疫力。     

大口黑鲈弹状病毒的分离培养及其卵黄抗体的制备

本研究从爆发性死亡的大口黑鲈(Micropterus salmoides)鱼苗病样中分离培养1株大口黑鲈弹状病毒(M.salmoides rhabdovirus,MSRV)毒株,采用甲醛灭活大口黑鲈弹状病毒,制备佐剂灭活疫苗,免疫产蛋鸡,收集高免蛋,制备MSRV卵黄抗体,测定其效价、中和效果及对病毒复制和宿主细胞内凝集素基因(intelectin)表达的影响。结果显示,成功利用草鱼(Ctenopharyngodon idellus)卵巢细胞株CO细胞分离培养大口黑鲈鱼弹状病毒,并用于佐剂灭活疫苗的制备;MSRV佐剂灭活疫苗免疫蛋鸡,成功获得MRSV的卵黄抗体,效价为1∶256,稀释度为1∶64时的中和病毒的作用最为明显,中和作用率达38.29%以上,病毒核酸拷贝数明显降低。同时,还可上调细胞内凝集素的表达。研究表明,特异性卵黄抗体对大口黑鲈弹状病毒具有明显的中和效果,为后续卵黄抗体作为免疫制剂的应用奠定了基础。     

不同佐剂对大鲵嗜水气单胞菌灭活疫苗的免疫效果

为了探究添加蜂胶佐剂、弗氏佐剂和铝胶佐剂后对大鲵（Andrias davidianus）嗜水气单胞菌（Aeromonas hydrophila）灭活疫苗的免疫效果,取已制备好的大鲵嗜水气单胞菌灭活疫苗,分别加入3种佐剂,充分混匀后制备成含菌量为1×108 CFU/mL的佐剂疫苗,通过腹腔注射免疫健康大鲵(平均体长40 cm、平均体质量85 g）,每尾1 mL,并设不加佐剂组和对照组;分别于免疫后第1、4、7、14、21、28、35天尾静脉采血,利用凝集反应检测血清抗体效价,于免疫后第35天进行攻毒感染试验。结果表明,与对照组相比,蜂胶佐剂组抗体效价于免疫后第1～14天呈上升趋势,第21天达最高,为1: 469.33,弗氏佐剂组于免疫后第1～28天呈上升趋势,第35天达到最大值,为1: 448.00,随后呈下降趋势;铝胶佐剂组抗体效价于免疫后第1～21天呈上升趋势,第28天达到最大值,为1: 362.67,不加佐剂组于免疫后第21天达最高,为1: 384.00,随后呈下降趋势,而对照组基本稳定在1:4.00。攻毒感染试验表明,蜂胶佐剂组、弗氏佐剂组、铝胶佐剂组和不加佐剂组的死亡率为10%、20%、25%和35%,相对免疫保护率分别为88.9%、77.8%、72.2%和61.1%,对照组大鲵死亡率为90%;3种佐剂疫苗均有较好的免疫保护效果,其中以蜂胶佐剂疫苗效果最佳。     

禽流感灭活疫苗和重组禽流感灭活疫苗免疫鸡抗体消长规律的比较研究

<正>禽流感灭活疫苗(H5N2,N28株)和重组禽流感灭活疫苗(H5N1,Re-1株)各3批,以重组禽流感灭活疫苗(H5N1,Re-1株)推荐的免疫程序和剂量分组免疫5周龄SPF鸡和5周龄SPF鸭。结果为:(1)免疫后1周,各免疫组均未测到HI抗体;(2)鸡免疫后4⁵周,HI抗体效价达到峰值1∶26.05周,HI抗体效价达到峰值1∶26.0¹∶27.0,随后2周下降相对较快,平均每周下降0.31∶27.0,随后2周下降相对较快,平均每周下降0.3⁰.7个滴度,之后HI抗体效价缓慢下降,直到试验结束,平均每周下降0.040.7个滴度,之后HI抗体效价缓慢下降,直到试验结束,平均每周下降0.04⁰.17个滴度;(3)免疫     

中华鳖腐皮病的免疫预防研究

选择对中华鳖毒力最强的Ｐ－１菌株,经扩大培养制成灭活福尔马林菌苗（Ｆ－ＡＨ）及Ｓｐａｎ白油佐剂菌苗（Ｚ－ＡＨ）,对中华鳖进行了Ｆ－ＡＨ菌苗的１次注射、２次注射及Ｚ－ＡＨ菌苗的注射免疫试验。结果表明,受免疫组鳖血液白细胞吞噬活性、凝集抗体效价及脾脏吞噬活性均显著高于对照组鳖,免疫保护力均达到８８％以上。     

抗鲤疱疹Ⅱ型病毒卵黄抗体制备及其功能鉴定

为制备抗鲤疱疹Ⅱ型病毒(CyHV-2)的卵黄抗体,探索防治异育银鲫(Carassius auratus gibelio)鳃出血病的新方法和途径,本研究利用原核表达系统产生具免疫原性的重组CyHV-2-ORF72衣壳蛋白,纯化后免疫蛋鸡;二次免疫后采用间接ELISA法抽检免疫蛋的特异性卵黄抗体(IgY)含量,收集抗体效价达到1∶8000的免疫蛋。将免疫蛋分成全蛋组与蛋黄组,采用微包膜和冷冻干燥技术,分别制成全蛋粉或蛋黄粉,利用Western blot和ELISA技术检测全蛋粉和蛋黄粉中的IgY特异性及效价。将蛋黄粉稀释液与CyHV-2混合后孵育鲤鱼上皮瘤细胞,进行抗体中和试验。以0.15%和0.30%的添加量分别将抗CyHV-2-ORF72的免疫全蛋粉拌入商品饲料制成功能性饲料,投喂已发病的两个池塘的异育银鲫进行生产性治疗试验。结果显示,全蛋粉和蛋黄粉中的IgY均能特异性地结合CyHV-2-ORF72衣壳蛋白,效价达到1∶8000~1∶16000;抗CyHV-2-ORF72的IgY与CyHV-2具有较好的亲和力,中和活性高于鼠抗ORF72血清;两个池塘异育银鲫成活率分别达到74.3%和87.6%,远高于对照池的28.3%和31.5%。综上所述,本研究制备的抗CyHV-2-ORF72卵黄抗体可特异性地中和CyHV-2病毒,具较显著的免疫保护作用。     

精制禽流感高免卵黄抗体的研究

用禽流感油乳剂灭活苗按一定的免疫程序免疫SPF蛋鸡,制备高免卵黄抗体,用改良的水稀释法对IgY进行纯化,经过抗体效价检测、无菌检验、安全试验、动物试验,该卵黄抗体可以用于田间试验。保存期试验证明:精制禽流感高免卵黄抗体室温条件下可保存10个月,4℃条件下保存15个月,-20℃条件下保存18个月。     

Neutralizing antibody levels for protection against betanodavirus infection in sevenband grouper, Epinephelus septemfasciatus (Thunberg), immunized with an inactivated virus vaccine

An inactivated betanodavirus, red‐spotted grouper nervous necrosis virus (RGNNV), is a vaccine candidate for viral nervous necrosis (VNN). The present study was conducted to examine inoculation doses of the vaccine and neutralizing antibody titre levels to protect fish against VNN. Young sevenband grouper, Epinephelus septemfasciatus, averaging 25.4 g, were immunized at 25 °C water temperature by a single intraperitoneal injection of formalin‐inactivated RGNNV. Fish immunized at vaccine doses of 10^8.5, 10^8.0, 10^7.5, 10^7.0 and 10^6.5 TCID₅₀ per fish produced antibodies at mean titres of 1:907, 1:511, 1:259, 1:197 and 1:96, respectively, at 20 days post‐immunization (p.i.). Neutralizing antibodies were not detected in any control fish (titre <1:80). When fish were challenged with RGNNV (10^5.0 and 10^4.0 TCID₅₀/fish) at 20 days p.i., cumulative mortalities of the fish groups immunized with 10^8.5, 10^8.0, 10^7.5 and 10^7.0 TCID₅₀ per fish were significantly lower than those of the control group, and the relative percent survival values were higher than 60% in fish groups immunized with 10^7.5 TCID₅₀ per fish or higher doses. However, no significant differences were found in mortality between the group immunized with 10^6.5 TCID₅₀ per fish and the control group. From these results, it was deduced that the minimum effective inoculation dose of the vaccine is 10^7.0 TCID₅₀ per fish and the minimum mean neutralizing antibody titre giving significant protection is approximately 1:200. This antibody titre level is a possible measure of vaccine efficacy against VNN in sevenband grouper, instead of a virus challenge test.     

野生水禽高致病性禽流感卵黄抗体监测

为了解野生水禽对高致病性禽流感病毒(AIV)自然被动免疫状况,于2010—2011年,在大庆龙凤湿地分别采集绿头鸭、须浮鸥、黑翅长脚鹬三种野生水禽巢卵150、184和128枚,采用血凝抑制试验检测H5和H7两种AIV的卵黄抗体。结果表明,绿头鸭H5和H7亚型禽流感卵黄抗体的阳性率分别为53.33%和14.67%,平均卵黄抗体滴度分别为4.55和3.45(log2);须浮鸥H5和H7亚型禽流感卵黄抗体的阳性率分别为34.78%和11.96%,平均卵黄抗体滴度分别为3.78和3.18(log2);黑翅长脚鹬H5和H7亚型禽流感卵黄抗体的阳性率分别为31.25%和9.38%,平均卵黄抗体滴度分别为3.85和3.67(log2)。由此可见,野生水禽种群对于高致病性禽流感的接触情况是较为频繁的,提示需要进一步加强对其种群的病毒携带情况进行监测。     

特异性鸡蛋抗体防治中华鳖红脖子病的初步研究

选用引起中华鳖红脖子病的嗜水气单胞菌菌株，以灭活菌株作为抗原，对母鸡进行免疫，从鸡蛋中得到抗该病原的特异性鸡蛋抗体，适用浸泡及口服免疫法，并将该特异性鸡蛋抗体应用于中华鳖红脖子病的免疫防治试验。结果表明：特异性鸡蛋抗体对中华鳖红脖子病有较强的免疫保护率，浸泡浓度大于0．6ppm时，免疫保护率可达64．3％；口服免疫法每千克饲料必须添加0．2克鸡蛋抗体时，才具有较好的免疫作用。     

Effects of temperature on hatchability, development and growth of eggs and yolksac fry of Oreochromis mossambicus (Peters) under artificial incubation

Abstract. The eggs and yolk sac fry of the mouth brooding tilapia. Oreochromis mossambicus (Peters), were artificially reared in an experimental hatchery system at different temperatures. The hatchability, embryonic development of eggs and growth of yolk sac fry were studied under controlled conditions. It was shown that the upper lethal temperature for O. mossambicus eggs was above 40°C and the lower lethal temperature between 11°C and 17°C. For yolk sac fry these values lay between 34 and 40°C and 17 and 20°C respectively. Fry survival between 24·3 and 34·0°C was near to 100% while al 20°C it was less than 60%. The somatic growth rates of fry at temperature between 20 and 34·5°C up to 10 days post hatch are presented and were found to differ significantly. At 34·6°C a negative SGR was recorded during the period 6–9 days post hatching, but, during the first 6 days, the SGR at this temperature was more than four times higher than at 20°C. At the elevated temperatures, the utilization of yolk is faster and the loss of weight observed was due to starvation. This should be taken into account when incubating under artificial rearing condition so that initiation of exogenous feeding is timed most appropriately.     

Humoral response and susceptibility of five full-sib families of Atlantic salmon, Salmo salar L., to the haemoflagellate, Cryptobia salmositica

Susceptibility and antibody production against pathogenic and vaccine strains of the haemoflagellate, Cryptobia salmositica were investigated in five full‐sib families (A–E) of Atlantic salmon, Salmo salar. Humoral response and susceptibility of families were compared within three treatments: infection, vaccination and vaccination followed by challenge. Parasitaemias caused by the vaccine strain of C. salmositica were considerably lower than those caused by the pathogenic strain. All vaccinated families were protected when challenged with the pathogenic strain. Family B had significantly lower parasitaemias (with both strains) than the other families. When naïve fish were infected with the pathogenic strain, this family had a significantly lower and earlier peak parasitaemia (4.3 ±1.3 × 10⁶ parasites mL^?1 blood at 3 weeks post‐infection; w.p.i.) than the other families. Family C had the highest peak (11.1 ± 1.2 × 10⁶ parasites mL^?1 blood), which occurred at 4 w.p.i. Antibodies against C. salmositica were detected earlier in Family B (3 w.p.i.) than in Family C (5 w.p.i.). This demonstrates an association of increased susceptibility with a delayed antibody response. Western immunoblot identified antibodies against 112, 181 and 200 kDa antigens earlier in more resistant fish (Family B). Antigenic stimulation leading to a stronger antibody response was shown with the vaccine strain and in the later stages of infection.     

WSSV口服卵黄抗体的制备及对螯虾的毒性实验

将提纯的WSS(V白斑综合症病毒)接种于蛋鸡后获得了高效价的Ig(Y卵黄抗体),与一定比例的保护剂混合后,瞬间喷雾干燥制成干粉其效价基本维持不变。在以螯虾为实验模型的攻毒及治疗试验结果表明:饲料中0.1%的IgY含量能在一定程度上延缓螯虾死亡时间,而0.2%的IgY含量则能够使螯虾的最终死亡率从100%降至30%左右。这表明WSSV特异性IgY制剂是安全有效的抗WSSV感染的的新型生物制剂。