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猪伪狂犬病是危害世界养猪业的重要疫病之一。近年来我国免疫猪群出现暴发、流行,经证实其抗原性和毒力发生了变异。为了有效区分变异毒株和经典毒株,根据国内外伪狂犬病病毒(PRV)变异毒株和经典毒株基因序列比对结果,针对UL44和UL36区域的基因序列设计了2对引物,建立两步法PCR。第一步PCR针对UL44区域,区分出经典毒株(疫苗株HB98除外)感染与变异毒株感染。第二步PCR针对UL36区域,区分出HB98株与伪狂犬病病毒变异毒株。两步法PCR的敏感度分别达到2~20TCID50和50TCID50病毒量。对猪繁殖与呼吸综合征病毒、猪圆环病毒2型、猪瘟病毒、日本脑炎病毒、脑心肌炎病毒和猪流行性腹泻病毒的核酸应用此方法进行扩增,结果均为阴性。该方法与猪伪狂犬病病毒国家标准PCR检测方法的符合率达到91%,可用于实验室快速诊断。
Pseudorabies is one of the important diseases that endanger the world pig industry. In recent years, the outbreaks of immunized pigs in our country, epidemic, confirmed its antigenicity and virulence have been mutated. In order to effectively distinguish between mutants and classical strains, two pairs of primers were designed according to the gene sequences of PRV and classical strains at home and abroad. Two pairs of primers were designed according to the sequence of UL44 and UL36 regions, Method PCR. The first step PCR targets the UL44 region, which distinguishes the classic strain (except vaccine strain HB98) from the infected and mutant strains. The second step of the PCR directed against the UL36 region identified the HB98 strain and the Pseudorabies virus variant strain. The sensitivity of the two-step PCR reached 2 ~ 20TCID50 and 50TCID50 respectively. This method was used to amplify the DNA of porcine reproductive and respiratory syndrome virus, porcine circovirus type 2, classical swine fever virus, Japanese encephalitis virus, encephalomyocarditis virus and porcine epidemic diarrhea virus. The results were negative. The coincidence rate between this method and the PRV PCR detection method reaches 91%, which can be used for rapid laboratory diagnosis.