【作者】 陈果亮；

【导师】 余兴龙；

【作者基本信息】 湖南农业大学 ， 临床兽医学， 2013， 博士

【摘要】 目的：本研究以规模化种猪场为依托,以猪瘟阻断ELISA抗体检测为主要手段,研究猪瘟疫苗免疫中的困境,以期找到一个可广泛采用的猪瘟免疫程序。并通过与猪场的实际生产管理相结合,提出规模化猪场猪瘟群体免疫的建议。方法：1)选择4个厂家猪瘟疫苗免疫试验猪群,分离血清进行抗体检测；以105TCID5o和106TCID50两个不同剂量的疫苗免疫仔猪并攻毒,通过临床症状观察、病理剖检和组织病理学观察、抗体检测、特异性淋巴细胞增殖试验及病原检测等方法评价疫苗免疫保护效果。2)按季度对怀孕母猪、后备母猪、公猪及哺乳母猪群体进行采血监测,评估种猪群现有免疫的状况；选择抗体水平不同的6头产房母猪,跟踪其仔猪母源抗体消长；随机选25窝同批次哺乳仔猪及其母猪为对象,分析母猪及其仔猪5周龄母源抗体的相互关系；3)通过选择疫苗按照既定技术路线免疫并进行抗体检测,综合评定10套仔猪免疫程序；25窝仔猪执行35-70日龄免疫程序,分别于免疫后34d、55d、62d、69d、98d、105d采血进行抗体检测,分析母猪抗体水平不同对其仔猪执行相同程序的影响；选取34日龄阻断值在9个不同区段的断奶仔猪,不同时间点进行抗体检测：4)选择抗体水平不同的母猪分为四组,跟踪免疫后猪瘟抗体的持续时间；选择4个不同猪场对阻断值低于70%的母猪加强免疫,分析免疫前后抗体消长情况。结果：1)不同厂家的疫苗在临床使用中,抗体应答及抗体水平存在差异,从中筛选了广东某厂的猪瘟ST细胞苗。该疫苗两个不同剂量一次免疫后第9d即能100%抵抗致死剂量强毒(1×104TCID50)攻击。2)4个不同群体采样1902份,抗体阳性率以怀孕母猪最高(91.71%)；平均阻断率以怀孕母猪最高(74.67%±22.22),后备猪次之；阻断率在70%以上的个体数比例以后备母猪比例最高(66.96%),各群体免疫状况良好。抗体水平阻断值高于85%的哺乳母猪其所产仔猪在乳后1天的抗体水平略高于母猪本身,且到乳后28天抗体最低值仍高于66.25%。低于75%的哺乳母猪其所产仔猪在乳后1天的抗体平均水平明显高于其母猪本身,于7日龄后开始下降,但至28日龄时抗体水平仍有40.1%-53.07%。通过25头母猪与其5周龄仔猪抗体分析,母猪抗体阻断率在70%以上时,其对应仔猪抗体阻断率在40%以上；在40%～70%区间,其对应仔猪阻断率在30%以下；窝间存在抗体水平差异性,且窝内仔猪个体间也存在差异。3)10套免疫程序以35日龄首免、70日龄二免的效果最好。首免后35d检测,34日龄母源抗体低于30%的组都为阳性,在30%-39%之间的组抗体值为疑似阳性,在40%以上的组首免后35d抗体为阴性；二免后4-6周,除母源抗体高于80%的分组外,其他组都为阳性,而且其抗体滴度与母源抗体的滴度呈负相关。4)高于80%阻断值的两组母猪跟踪至免疫后300天,抗体下降不明显。70%区间组抗体水平下降明显,跟踪至240天时抗体水平接近阳性临界值,差异性显著；母猪抗体阻断值低于50%,加强免疫后增幅大,而高于70%的则增幅不明显。结论：1)即便使用相同免疫程序,不同厂家的猪瘟疫苗在临床使用中抗体应答及抗体水平也存在明显差异；猪瘟ST细胞苗对仔猪安全有效。2)虽然在基础群抗体水平调查中表明群体有较好的免疫效果,但抗体高低仍存在差异,会进一步造成仔猪免疫前后的差异性增大,给猪群带来潜在的风险：哺乳母猪抗体水平与仔猪母源抗体持续时间呈明显的正相关；3)35-70免疫程序具有较好的推广价值：高母源抗体对仔猪一免产生影响,推迟一免的时间可以减轻其影响;母源抗体水平整齐有助于设计仔猪免疫程序,34日龄母源抗体在40%-60%对首免的效果有明显影响,但仍有免疫应答,低于40%对首免效果影响不大。4)抗体水平高于80%阻断值的母猪一年可以只进行一次免疫;加强免疫试验中,低于50%阻断值的母猪抗体上升幅度较大,大于70%阻断值的分组提升幅度有限,据此推断,根据不同情况对猪群进行普查和抽查,低于70%阻断值的母猪加强免疫,淘汰不理想母猪,可提高母猪群体抗体水平的整齐度。更多还原

【Abstract】 The purpose of this study is to get a reasonable, scientific immunity procedure of CSF which is suitable for the large-scaled pig farm. A commercial blocking antibody ELIS A Kits was employed to detect the titer of antibody against classical swine fever virus (CSFV), by which the influence of the quantity of vaccine on the level of antibody, the relation between the level of piglets’maternal derived antibody(MDA) and pregnant sows, the dynamic change of piglets’MDA, the affect of MDA on antibody of piglets trigered by vaccine and the the dynamic change of antibody level in sow were investigated.Several commercial cell original CSF vaccines were chosen to immunize pre-tests herds for analyses of the antiboy titer in sera and effect of protection agaist CSFV infection, and the results revealed that the antibody (MDA) response and level of antibody titer for each vaccine which from various companies are different obviously. Finally, a vaccine from a Guangdong producer was set as a reference vaccine in this study for its best protection against CSFV infection. As the results showed, single vaccination with this vaccine in two different dose (106TCID50and105TCIDso) can provide the piglets a complete protection against challenge with strong virulent CSFV strain in a lethal dose (1x104TCID50)The antibody titers of different herds were obsevered to analysis the differences among pregnant sows, lactating sows, boars and replacement breeding pigs, and results indicated that pregnant sows got the highest score with91.71%in antibody positive rate and74.67%±22.22in blocking rate. The titer and duration of piglets’ MDA were directly related to the antiboy titer of sows, as results showed the higher level of antibody in sows, the higher antibody titer, the slower declease rate, and longer duration in piglets; otherwise contrast completely.Ten serials of immunity procedure were conducted to estimate the merits of the immunity procedure, and it was demonstrated that piglets vaccinated twice on35d and70d was the best procedure. Piglets with various level of MDA titer were divided into different groups and vaccinated to observe the influence of MDA on vaccination in piglets. It was found that the effect of inmmunity was seriously disturbed by MDA after the fisrt vaccination both in antibody titer and positive rate, resulting a contrary effect, same in the second vaccination.In order to evaluate the duration of antibody in sows, the change dynamic of antibody titer was obersevered among sows with different level antibody. It was found that the antibody titer was not decreased any more in sows with high level antibody (above80%in blocking rate), comparely it was obviousely in sows with70-80%. Inaddtionally, sows, which antiboy level under70%, were conducted a second inmmunity to investigate its effect to improve antiboy level. Intersetingly, the positive was obviously in sows which antibody level was under50%, while less in those above this level.According to the results above, we can get a conclusion:the quantity of CSFV vaccine is critial to the antibody level and effective protection; the level of MDA in piglets is depended on the antibody titer of sows, however it presented a negative infact on the effect of vaccine on piglets inmmunity; there is a direct relation between antibody duration and level in sows, and secondary vaccination can improve antibody titer of sows which present a low level. Thus, it is very important to investigate the antibody titer of sows in modern, large-scaled pig farm, which is dispensible for producing a reasonable, scientific immunity procedure to provent CSF.更多还原