猪繁殖与呼吸综合征（PRRS）病原分离鉴定及防控措施研究

发布时间：2018-06-08 13:42

本文选题：PRRSV + 遗传变异　；参考：《吉林大学》2016年博士论文

【摘要】：本研究采集河南省及周边不同地区疑似PRRS病猪的组织病料进行PRRSV分离与鉴定,并对分离毒株进行遗传变异分析,以河南省PRRS的流行状况调查结果为基础,研究PRRSV弱毒疫苗、三仁汤复方中药制剂和抗PRRSV免疫血清对PRRS的防治效果,以寻求简捷、有效、实用的PRRS防控措施,以提高养猪业的生产水平。1、收集河南省及湖北部分市区2006~2014年疑似PRRS发病猪的病料180余份,用Marc-145细胞培养进行病毒分离与鉴定,分离得到10株PRRSV,其中河南省共9株,湖北襄樊1株,以美国经典株ATCCVR-2332和中国经典株CH-1a为参考毒株,将分离株的NSP2和ORF5-7片段分别克隆到p MD-18T载体并进行测序。结果表明PRRSV分离株均为美洲型毒株,且Nsp2基因均已发生了缺失,GP5基因没有缺失,只有基因位点突变;其中He N1-06与ATCCVR-2332处于同一个进化分支,其它9株毒株(He N2-06,He N3-06,He N1-07,He N2-07,He N3-07,He N1-08,He N2-08,He N3-08,He N4-08)与CH-1a株处于同一进化分支。证实了河南省PRRSV一直在变异,且以高致病性毒株为主流。2、PRRS母源抗体消长规律试验结果表明:仔猪母源抗体水平高低和维持时间长短与母猪初乳抗体水平呈正相关,母源抗体的S/P在14日龄最高,随后开始下降,21日龄下降速度加快,38日龄降到最低,平均半衰期12天,由此确定免疫方案:仔猪21日龄实行首免,45日龄二免,剂量1头份/头弱毒疫苗肌肉注射,种猪每年普免3次,剂量2头份/次弱毒疫苗肌肉注射。3、通过三仁汤复方中药制剂作用于Marc145细胞后,观察Marc145细胞的病变率(CPE),研究三仁汤复方中药对PRRSV在Marc-145上的吸附阻断作用,增殖抑制作用以及直接灭活作用。结果表明:三仁汤复方中药制剂对PRRSV有较强的吸附阻断作用、增殖抑制作用和直接灭活作用,其最小质量浓度分别为125g/L、31.25g/L和15.625g/L。4、PRRS临床防控研究结果表明:按照上述免疫方案实施后,种猪配种分娩率由76.6%提高到90.5%,提高了13.9%;保育猪成活率由89.0%提高到98.0%,提高了9.0%;育肥猪成活率由87.0%提高到95.0%,提高了8.0%;三仁汤复方中药临床治疗1825头发生PRRS保育猪成活率达到99.28%,治疗8994头发生PRRS的育肥猪成活率达到95.73%;临床对照治疗实验结果表明:用三仁汤复方中药治疗1153头发生PRRS病猪成活率为97.66%,日增重390g/d,用抗生素控制继发感染治疗1040头发生PRRS病猪成活率为80.48%,日增重310g/d,成活率提高了17.18%,日增重提高了80g/d;用三仁汤治疗发生PRRS种猪群配种分娩率提高了39.8%,新生仔猪健仔率提高了53.2%。用抗PRRSV免疫血清在保育猪发生PRRS早期按1m L/kg体重剂量肌肉注射,成活率由86.56%提高到96.85%,提高了10.32%。综上所述,本研究证明,河南省PRRSV的感染与流行十分普遍,毒株已发生了不同程度的变异,运用PRRSV弱毒疫苗、三仁汤复方中药和抗PRRSV免疫血清联合防治PRRS的发生,有效减少了母猪流产和仔猪死亡,改善了猪群健康状况,提高猪场的生产水平。
[Abstract]:In this study, tissue samples of suspected PRRS pigs in Henan Province and its surrounding areas were collected for PRRSV isolation and identification, and genetic variation of the isolates was analyzed. Based on the results of the investigation on the prevalence of PRRS in Henan Province, PRRSv attenuated vaccine was studied. The prevention and control effect of Sanren decoction compound traditional Chinese medicine preparation and anti-PRRSv immune serum on PRRS, in order to seek simple, effective and practical preventive and control measures of PRRS. In order to improve the production level of pig industry, more than 180 samples of suspected PRRS infected pigs from 2006 to 2014 in Henan Province and some urban areas of Hubei Province were collected. The virus was isolated and identified by Marc-145 cell culture, and 10 strains of PRRSVS were isolated and identified, of which 9 strains were isolated from Henan Province. The NSP2 and ORF5-7 fragments of Xiangfan strain were cloned into pMD-18T vector and sequenced using the American classic strain ATCCVR-2332 and the Chinese classical strain CH-1a as reference strains. The results showed that PRRSv isolates were all American strains, and the deletion of GP5 gene was not found in all of the Nsp2 genes, but only the mutation of gene locus, in which he N1-06 was in the same evolutionary branch as ATCCVR-2332. The other 9 strains, he N2-06HN3-06H1-06H1-06HN1-07H1-07HN1-07H1-07HN1-08H1-08H3-08He N4-08) are in the same evolutionary branch as the CH-1a strain, and the other 9 strains are in the same phylogenetic branch as the CH-1a strain, and the other 9 strains are in the same phylogenetic branch as the CH-1a strain. It was confirmed that PRRSv had been mutating all the time in Henan Province, and the results showed that the maternal antibody level of piglets and the duration of maintenance were positively correlated with the antibody level of porcine colostrum. The S / P of maternal antibody was highest at 14 days of age, then began to decline, and then began to decline at 21 days of age. The decline rate was accelerated to the lowest at 38 days of age, with an average half-life of 12 days. Thus, the immunization scheme was determined: the first day of 21 days old piglets were given the second immunity at 45 days old. A dose of 1 head / head attenuated vaccine was injected intramuscularly, 3 times a year for pig immunizations, and 2 doses per second for intramuscular injection of attenuated vaccine. The cells were treated with Sanren decoction compound Chinese medicine preparation. To observe the cytopathic rate of Marc145 cells and to study the inhibition of PRRSv on Marc-145, the inhibition of proliferation and the direct inactivation of PRRSv by Sanren decoction. The results showed that Sanren decoction had strong adsorption blocking, proliferation inhibition and direct inactivation on PRRSV, and its minimum mass concentration was 125g / L 31.25g / L and 15.625g / L / L respectively. The birth rate of breeding pigs increased from 76.6% to 90.5%, which increased 13.9%; the survival rate of preserved pigs increased from 89.0% to 98.0, which increased 9.0%; the survival rate of fattening pigs increased from 87.0% to 95.0%, which increased 8.0%. The survival rate of the fattened pigs treated with 8994 hair PRRS was 95.730.The results of clinical controlled treatment showed that the survival rate of 1153 hair pigs treated with Sanren decoction was 97.66, the daily gain was 390g / d, and the secondary infection was controlled by antibiotics. The survival rate was 80.48g / d, the daily gain was 310g / d, the survival rate was 17.18g / d, the daily gain was increased 80g / d, the rate of breeding and delivery was increased by 39.8g / d, and the healthy rate of newborn piglets was increased 53.2d. After intramuscular injection of PRRS with anti-PRRSV immune serum, the survival rate was increased from 86.56% to 96.85 in the early stage of PRRS. In conclusion, this study proved that the infection and prevalence of PRRSv in Henan Province is very common, the virus strains have been mutated to different degrees, using PRRSv attenuated vaccine, Sanren decoction compound Chinese medicine and anti-PRRSV immunized serum to prevent and treat PRRS. It can effectively reduce abortion and piglet death, improve the health status of pig herd and improve the production level of pig farm.
【学位授予单位】：吉林大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：S858.28

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