豫北新发猪圆环病毒3型的鉴定及全基因序列分析
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摘要
为了对疑似新发猪圆环病毒3型(PCV-3)感染病例进行确诊,试验采用PCR技术鉴定1例疑似感染PCV-3的猪,并将鉴定得到的PCV-3毒株的全基因组分成2个片段进行PCR扩增,分别构建到pMD18-T载体上测序,通过MegAlign软件对全基因序列进行同源性和遗传进化分析,利用在线生物信息学软件预测Cap和Rep蛋白的信号肽、核定位信号和B细胞表位。结果表明:成功鉴定出1株豫北新发PCV-3毒株,将此毒株命名为PCV-3/CN/HNAY-201806;对2个基因片段测序后拼接发现其基因组长度为2 000 bp(登录号为MH683051);同源性和遗传进化分析显示,PCV-3/CN/HNAY-201806与吉林长春毒株PCV-3/CHN/JLCC2016(登录号为KY421348.1)核苷酸同源性(99.6%)较高,与广西毒株PCV-3/GXFC2017-7(登录号为MG250186.1)同源性(98.8%)较低,不同地区间PCV-3基因组有差异但较小;对该毒株Cap和Rep蛋白序列分析显示,2个蛋白都没有信号肽序列,但都有核定位信号,Cap蛋白有9个B细胞表位,Rep蛋白有14个B细胞表位。
In this study, it was identified by PCR that a pig suspected to be infected with Porcine circovirus type 3(PCV-3). The genome of the identified PCV-3 strain was amplified as two parts by PCR. Two parts were cloned into pMD18-T vector to be sequenced. Homology and polygenetic tree analysis were performed for the whole genome sequence by MegAlign. Signal peptides, nuclear localizations and B cell epitopes were predicted by online bioinformatics softwares. The results showed that a strain of PCV-3 from the North of Henan Province was successfully identified and named as PCV-3/CN/HNAY-201806. The length of its genome was 2 000 bp(GenBank No. MH683051) after two gene fragments sequencecl and spliced. Homology and polygenetic tree analysis suggested that PCV-3/CN/HNAY-201806(GenBank No. KY421348.1) had the higher homology(99.6%) with the strain PCV-3/CHN/JLC2016(GenBank No. KY421348.1) from Changchun City in Jilin Province, and a lower homology(98.8%)with the strain PCV-3/GXFC2017-7(GenBank No. MG250186.1) from Guangxi Province, indicating that there was different but little different for PCV-3 genome sequence in various regions. It showed that there were no signal peptide while there were own nuclear localizations for Cap and Rep proteins of this strain. The Cap protein had nine B cell epitopes, and Rep protein had fourteen B cell epitopes.
In this study, it was identified by PCR that a pig suspected to be infected with Porcine circovirus type 3(PCV-3). The genome of the identified PCV-3 strain was amplified as two parts by PCR. Two parts were cloned into pMD18-T vector to be sequenced. Homology and polygenetic tree analysis were performed for the whole genome sequence by MegAlign. Signal peptides, nuclear localizations and B cell epitopes were predicted by online bioinformatics softwares. The results showed that a strain of PCV-3 from the North of Henan Province was successfully identified and named as PCV-3/CN/HNAY-201806. The length of its genome was 2 000 bp(GenBank No. MH683051) after two gene fragments sequencecl and spliced. Homology and polygenetic tree analysis suggested that PCV-3/CN/HNAY-201806(GenBank No. KY421348.1) had the higher homology(99.6%) with the strain PCV-3/CHN/JLC2016(GenBank No. KY421348.1) from Changchun City in Jilin Province, and a lower homology(98.8%)with the strain PCV-3/GXFC2017-7(GenBank No. MG250186.1) from Guangxi Province, indicating that there was different but little different for PCV-3 genome sequence in various regions. It showed that there were no signal peptide while there were own nuclear localizations for Cap and Rep proteins of this strain. The Cap protein had nine B cell epitopes, and Rep protein had fourteen B cell epitopes.
引文
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[12] 冯选榕. 广西某市屠宰猪淋巴结PCV2和PCV3的病原检测及遗传进化分析[D]. 南宁:广西大学, 2017.
[13] SHEN H, LIU X, ZHANG P, et al. Genomecharacterization of a Porcine circovirus type 3 in South China [J]. Transb Emerg Dis, 2018, 65(1): 264-266.
[14] DENG J,LI X,ZHENG D,et al. Establishment andapplication of an indirect ELISA for Porcine circovirus 3[J].Arch Virol, 2018, 163(2): 479-482.
[2] TISCHER I, MIELDS W, WOLFF D, et al. Studies on epidemiology and pathogenicity of Porcine circovirus[J].Arch Virol, 1986, 91(3/4):271-276.
[3] XIAO C, HALBUR P, OPRIESSNIG T, Global molecular genetic analysis of Porcine circovirus type 2 (PCV2) sequences confirms the presence of four main PCV2 genotypes and reveals a rapid increase of PCV2d[J]. J Gen Virol,2015, 96(Pt7):1830-1841.
[4] PALINSKI R, PINEYRO P, SHANG P, et al. A novel Porcine circovirus distantly related to known circoviruses is associated with porcine dermatitis and nephropathy syndrome and reproductive failure[J]. J Virol, 2017, 91(1):e01879-16.
[5] KU X, CHEN F, LI P, et al. Identification and genetic characterization of Porcine circovirus type 3 in China[J]. Transb Emerg Dis, 2017, 64(3):703-708.
[6] ZHENG S, WU X, ZHANG L, et al. The occurrence of Porcine circovirus 3 without clinical infection signs in Shandong Province[J]. Transb Emerg Dis, 2017, 64(5):1337-1341.
[7] STADEJEK T, WOZNIAK A, MILEK D, et al. First detection of Porcine circovirus type 3 on commercial pig farms in Poland[J]. Transb Emerg Dis, 2017, 64(5):1350-1353.
[8] KWON T, YOO S, PARK C, et al. Prevalence of novel Porcine circovirus 3 in Korean pig populations[J].Vet Microbiol, 2017, 207:178-180.
[9] YE X, BERG M, FOSSUM C, et al. Detection and genetic characterisation of Porcine circovirus 3 from pigs in Sweden[J]. Virus Genes, 2018, 54 (3): 466-469.
[10] 宋婉如,王秋霞,余燕,等. 断奶仔猪圆环病毒2型和葡萄球菌混合感染的诊断[J]. 黑龙江畜牧兽医,2016(10下):138-140, 299.
[11] 湛洋,王东亮,王乃东,等.猪圆环病毒3型检测及其Cap结构序列和抗原性预测分析[J].畜牧兽医学报,2017,48(6):1076-1084.
[12] 冯选榕. 广西某市屠宰猪淋巴结PCV2和PCV3的病原检测及遗传进化分析[D]. 南宁:广西大学, 2017.
[13] SHEN H, LIU X, ZHANG P, et al. Genomecharacterization of a Porcine circovirus type 3 in South China [J]. Transb Emerg Dis, 2018, 65(1): 264-266.
[14] DENG J,LI X,ZHENG D,et al. Establishment andapplication of an indirect ELISA for Porcine circovirus 3[J].Arch Virol, 2018, 163(2): 479-482.