猪细小病毒灭活疫苗NJ株毒种分离及鉴定

为防控猪细小病毒病,进行了猪细小病毒灭活疫苗毒种研究。将南京某猪场初产母猪流产胎儿的脾、肺等组织病料处理后接种ST细胞培养,获得了1株猪细小病毒强毒,将分离毒株适当稀释接种ST细胞,经3轮蚀斑纯化获得1株纯净的猪细小病毒NJ株(PPV-NJ株),作为疫苗毒株。将NJ株接种ST细胞培养连续传15代,每代次病毒效价均不低于1 ml 107.25TCID50、血凝素效价不低于29。选择第5代病毒液,灭活后与矿物油佐剂混合乳化制成灭活疫苗,以不同剂量分别免疫豚鼠和后备母猪,用血凝抑制试验(HI)和ELISA检测抗体效价。分别用剂量0.125 ml、0.250 ml和0.500 ml疫苗免疫豚鼠28 d后,各免疫组HI抗体效价均高于28,0.5 ml组HI抗体效价高达211,ELISA抗体均显示阳性(OD630≥0.32),分别用剂量0.5 ml、1.0 ml和2.0 ml疫苗免疫后备母猪后,HI检测结果显示,3个剂量组免疫后1周HI抗体效价高于26,3周后达峰值,至免疫后4个月略有下降,2.00 ml剂量组HI效价最高(211.75)。ELISA检测结果显示,各剂量组在免疫后1周均转阳(OD630≥0.32),免疫后4～18周均维持较高水平。HI抗体效价和ELISA抗体水平与免疫剂量均呈正相关性。结果显示,利用NJ株病毒制备的灭活疫苗具有抗体产生快、效价高等特点,显示出良好的疫苗开发前景。

Isolation and identification of NJ strain of porcine parvovirus

In order to obtain a vaccine candidate for controlling porcine parvovirus(PPV),a PPV virus strain was isolated from the lung and spleen of aborted fetus which derived from a primiparity sow in a pig farm of Nanjing City.The isolate,named NJ strain,propagated in swine testicle(ST) cell line,and then was purified after 3-round plaque purification.NJ strain displayed high titer(≥107.25TCID50 per milliliter) and haemagglutinin titer(≥29 per milliliter) in every generation in serial passage to 15 generations in ST cell line.The fifth passage NJ virus was selected to produce inactive vaccine with mineral oil emulsion,and was then injected into guinea pigs or replacement gilts with various doses,respectively.The antibody against PPV was detected by both haemagglutinin inhibition(HI) test and ELISA.In guinea pigs,all HI titers ≥ 28,and ELISA antibodies were positive(OD630≥ 0.32) at day 28 post-inoculation(P.I) with doses of 0.125 ml,0.250 ml,or 0.500 ml,respectively.The highest HI titer was 211 in 0.5 ml inoculation treatment.In guinea pigs,all HI titers ≥ 26 at week 1 P.I,peaked at week 3 P.I,and slightly declined at mouth 4 P.I,at doses of 0.5 ml,1.0 ml or 2.0 ml,respectively.The highest HI titer was 211.75 in 2.0 ml inoculation treatment.ELISA antibodies were positive at week 1 P.I,(OD630≥ 0.32),and relative rich at 4-18 weeks P.I.HI antibody titers and ELISA level were both positively correlated with vaccination dose.The inactivated vaccine prepared with NJ isolate displayed fast antibody production and high titer,suggesting the prospective development of the vaccine.