【摘要】 为了解辽宁沈阳地区猫泛白细胞减少症病毒(Feline panleukopenia virus, FPV)VP2和NS1基因的遗传变异情况，试验从辽宁省沈阳市3个动物医院和1个猫繁育场收集疑似FPV感染的猫粪便拭子样本13份(编号为FPV-8～FPV-20),利用胶体金试纸条进行FPV检测，提取阳性样本中的病毒DNA后进行VP2和NS1基因的PCR扩增、测序、同源性分析、编码蛋白的氨基酸突变位点分析及遗传进化树构建。结果表明：13份样本均呈FPV阳性，经PCR扩增均得到大小为1 755 bp的VP2基因和大小为2 007 bp的NS1基因。13份样本中的FPV VP2基因之间的核苷酸相似性为98.7%～100%,与GenBank中FPV参考毒株的核苷酸相似性为98.6%～100%,其中FPV-LN15、FPV-LN18、FPV-LN19与BJ540(MT270553.1)、FPV ZJFPV15(MW495840.1)和FPV-SH2003(MW811187.1)的核苷酸相似性为100%;13份样本中的FPV NS1基因之间的核苷酸相似性为99.0～100%,与GenBank中FPV参考毒株的核苷酸相似性为99.1%～99.9%,其中FPV-LN8、FPV-LN10和PPV-LN13的核苷酸相似性为100%,FPV-LN9、FPV-LN18和FPV-LN19的核苷酸相似性为100%,PPV-LN15和FPV-LN17的核苷酸相似性为100%。13份样本中的FPV VP2蛋白共存在5个氨基酸突变位点(第91,301,302,305,307位),FPV NS1蛋白共存在6个氨基酸突变位点(第17,23,60,247,443,596位)。基于VP2基因构建的遗传进化树中共分为2个大分支，FPV和水貂肠炎病毒(Mink enteritis virus, MEV)毒株聚为一支，犬细小病毒(Canine parvovirus, CPV)毒株处于一个独立分支。样本FPV-LN11、FPV-LN13、FPV-LN8、FPV-LN16、FPV-LN10、FPV-LN20处于一个较小分支，样本FPV-LN14处于独立小分支，且与FPV HNZZ2株(MZ005633.1,郑州)、FPV Barut株(MZ391096.1,土耳其)、FPV 17D01株(OP153925.1,韩国)、FPV DLC02株(MN418998.1,大连)处于不同小分支；样本FPV-LN9、FPV-LN12、FPV-LN15、FPV-LN18、FPV-LN19与FPV JSNJ-21G5株(OP796709.1,扬州)、FPV BJ540株(MT270553.1,北京)、FPV-SH2003株(MW811187.1,上海)、FPV ZJFPV15株(MW495840.1,扬州)处于一个较小分支，样本FPV-LN17处于独立小分支，且与FPV＿ARG08(FJ440714.1)株(阿根廷)处于不同小分支。基于NS1基因构建的遗传进化树共分为两个大分支，FPV毒株和CPV毒株分别为两个独立的分支，FPV-LN9、FPV-LN12、FPV-LN15、FPV-LN17、FPV-LN14、FPV-LN18、FPV-LN19与FPV-SH2003株(MW811187.1,上海)和FPV-SH2001株(MW650831.1,上海)处于一个较小分支，FPV-LN8、FPV-LN11、FPV-LN13、FPV-LN10、FPV-LN16、FPV-LN20株与FPV HNZZ2株(MZ005633.1,郑州)处于一个较小分支，均与FPV＿IZSSI＿42807＿15株(KX434462.1,意大利)、FPV株(X55115.1,澳大利亚)处于不同小分支。说明各样本两种基因的亲缘关系基本一致(除样本FPV-LN14外),FPV毒株在同一地理范围内具有较高的遗传相似性。更多还原

【Abstract】 In order to understand the genetic variation of VP2 and NS1 genes of Feline panleukopenia virus(FPV) in Shenyang City, Liaoning Province, in the test, thirteen fecal swab samples(numbered FPV-8 to FPV-20) of cats with suspected FPV infection were collected from three animal hospitals and one cat breeding farm in Shenyang City, Liaoning Province. FPV was detected by using colloidal gold test strips; PCR amplification of VP2 and NS1 genes, sequencing, homology analysis, mutation analysis of the amino acid sites of the encoded proteins, and the construction of genetic evolution tree were carried out after extracting the viral DNA of the positive samples. The results showed that all 13 samples were positive for FPV; the VP2 gene with a size of 1,755 bp and the NS1 gene with a size of 2,007 bp were amplified by PCR. The nucleotide similarity between VP2 genes of 13 samples was 98.7%-100%, and the nucleotide similarity with the FPV reference strains in GenBank was 98.6%-100%; among them, the nucleotide similarity between FPV-LN15, FPV-LN18, FPV-LN19 and BJ540(MT270553.1), FPV ZJFPV15(MW495840.1), FPV-SH2003(MW811187.1) was 100%. The nucleotide similarity between NS1 genes of 13 samples was 99.0-100%; the nucleotide similarity with the FPV reference strains in GenBank was 99.1%-99.9%; among them, the nucleotide similarity between FPV-LN8 and FPV-LN13 was 100%, the nucleotide similarity between FPV-LN9, FPV-LN18 and FPV-LN19 was 100%, and the nucletide similarity between FPV-LN15 and FPV-LN17 was 100%. A total of 5 amino acid mutation sites(positions 91, 301, 302, 305, 307) were present in the FPV VP2 protein in 13 samples; 6 amino acid mutation sites(positions 17, 23, 60, 247, 443, 596) were present in the FPV NS1 protein. In the genetic evolution tree constructed based on the VP2 gene, there were 2 large branches; the strains of FPV and Mink enteritis virus(MEV)were clustered into one branch, and Canine parvovirus(CPV) was in an independent branch. Samples FPV-LN11, FPV-LN13, FPV-LN8, FPV-LN16, FPV-LN10 and FPV-LN20 were in a smaller branch; sample FPV-LN14 was in a separate sub-branch and was in a different sub-branch from strains FPV HNZZ2(MZ005633.1)(Zhengzhou),FPV Barut(MZ391096.1)(Turkey), FPV 17D01(OP153925.1) strain(Korea), and FPV DLC02(MN418998.1) strain(Dalian) were in different sub-branches. The samples FPV-LN9, FPV-LN12, FPV-LN15, FPV-LN18, and FPV-LN19 were in a smaller branch with the FPV JSNJ-21G5(OP796709.1) strain(Yangzhou), the FPV BJ540(MT270553.1) strain(Beijing), the FPV-SH2003(MW811187.1) strain(Shanghai), the FPV ZJFPV15(MW495840.1) strain(Yangzhou) in a smaller branch; the sample FPV-LN17 was in a separate sub-branch and in a different sub-branch from the FPV＿ARG08(FJ440714.1) strain(Argentina). In the genetic evolution tree constructed based on the NS1 gene, there were two major branches; the strains of FPV and CPV formed two independent branches, respectively. FPV-LN9, FPV-LN12, FPV-LN15, FPV-LN17, FPV-LN14, FPV-LN18 and FPV-LN19 were in a smaller branch with strains FPV-SH2003(MW811187.1)(Shanghai) and FPV-SH2001(MW650831.1)(Shanghai). Strains FPV-LN8, FPV-LN11, FPV-LN13, FPV-LN10, FPV-LN16, and FPV-LN20 were in a smaller branch with strain HNZZ2(MZ005633.1)(Zhengzhou), and all were in different minor branches from FPV＿IZSSI＿42807＿15(KX434462.1)(Italy), FPV(X55115.1)(Australia). These results suggested that the relationship between the two genes was basically the same in all samples(except FPV-LN14), and that the FPV strains had a high degree of genetic similarity within the same geographic range.更多还原