2016—2020年浙江地区猪圆环病毒2型分子流行病学分析

doi:10.3785/j.issn.1008-9209.2021.11.051

浙江大学学报（农业与生命科学版）

2022, Vol. 48

Issue (5): 644-652 DOI: 10.3785/j.issn.1008-9209.2021.11.051

动物科学与动物医学

2016—2020年浙江地区猪圆环病毒2型分子流行病学分析

徐丽华¹(

),苏菲¹,李军星¹,余斌¹,叶十一¹,杨富文¹,邓利荣²,毛慧敏²,袁秀芳¹(

)

^1.浙江省农业科学院畜牧兽医研究所，杭州 310021
^2.江山康农农业发展有限公司，浙江衢州 324100

Molecular epidemiological analysis of porcine circovirus type 2 in Zhejiang Province from 2016 to 2020

Lihua XU¹(

),Fei SU¹,Junxing LI¹,Bin YU¹,Shiyi YE¹,Fuwen YANG¹,Lirong DENG²,Huimin MAO²,Xiufang YUAN¹(

)

^1.Institute of Animal Husbandry and Veterinary Science, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China
^2.Jiangshan Kangnong Agricultural Development Co. , Ltd. , Quzhou 324100, Zhejiang, China

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摘要：

为了解猪圆环病毒2型（porcine circovirus type 2, PCV2）在浙江地区的流行病学特点和遗传变异规律，本研究采用聚合酶链反应（polymerase chain reaction, PCR）对2016—2020年采自浙江省不同地区猪场的1 725份疑似猪圆环病毒2型感染的临床病料进行PCV2的病原学检测，同时对部分阳性样品进行全基因组扩增、克隆和测序，并与GenBank中收录的16株参考株序列进行比对和遗传进化分析。结果表明：PCV2检测阳性的样品有359份，平均阳性率为20.8%，2016—2020年PCV2阳性率分别为38.1%、23.2%、24.1%、12.5%和10.7%；测序所得PCV2 36个全基因序列的核苷酸同源性为94.0%～99.9%，与国产疫苗株核苷酸同源性为94.7%～98.5%，与参考株核苷酸同源性为92.9%～99.8%；遗传进化分析显示PCV2 36个全基因序列可分为3个基因型，其中11株为PCV2a型，8株为PCV2b型，17株为PCV2d型，PCV2d为优势基因型；ORF2基因全长分别为705 bp（16株）和702 bp（20株），相互间核苷酸同源性为88.7%～100.0%，与国产疫苗株核苷酸同源性为90.2%～99.6%，与参考株核苷酸同源性为85.0%～100.0%。Cap蛋白的氨基酸序列分析表明，36株序列除了唯一糖基化位点呈高度保守外，存在较多的点突变，且不同基因型具有其代表性突变位点，优势基因型PCV2d的特异性突变位点主要集中在I⁵³、N⁶⁸、L⁸⁹、T¹²¹、R¹⁶⁸、I²¹⁵。本研究为浙江地区PCV2的免疫预防提供了参考，为新型疫苗的研制积累了有效材料。

关键词： 猪圆环病毒2型; 全基因组; Cap蛋白; 分子流行病学; 遗传变异

Abstract:

In order to investigate the molecular epidemiology and genetic variation of porcine circovirus type 2 (PCV2) in Zhejiang Province, a total of 1 725 clinical samples suspected of PCV2 infection were collected from different regions of Zhejiang Province between 2016 and 2020. All these samples were subjected to pathogenic detection by polymerase chain reaction (PCR). The whole genomes of some positive samples were amplified, cloned and sequenced, and compared with sequences of 16 reference strains in the GenBank for genetic evolution analysis. The results showed that 359 samples were tested positive, with an average positive rate of 20.8%, and the positive rates from 2016 to 2020 were 38.1%, 23.2%, 24.1%, 12.5%, and 10.7%, respectively. The whole genomes of 36 isolates were sequenced. Multiple sequence alignments showed that the nucleotide sequence homologies among 36 PCV2 strains were 94.0%-99.9%, while the homologies were up to 94.7%-98.5% compared with the domestic vaccine strains, and 92.9%-99.8% compared with the reference strains. The phylogenetic analysis showed that these 36 isolates belong to three genotypes, including 11 strains of PCV2a genotype, eight strains of PCV2b genotype and 17 strains of PCV2d genotype. Among these, PCV2d was the predominant genotype. The length of PCV2 ORF2 gene was 705 bp in 16 isolates and 702 bp in 20 isolates. The nucleotide sequence homologies of ORF2 gene among these 36 isolates were 88.7%-100.0%, while the homologies were 90.2%-99.6% compared with the domestic vaccine strains, and 85.0%-100.0% compared with the reference strains. The amino acid sequence analysis of Cap protein indicated a large number of point mutations in addition to the only highly conserved glycosylation site. Moreover, different genotypes had characteristic mutation sites, and the specific mutation sites of dominant genotype PCV2d were mainly concentrated in I⁵³, N⁶⁸, L⁸⁹, T¹²¹, R¹⁶⁸, and I²¹⁵. This study provides a reference for the immune prevention of PCV2 in Zhejiang Province and accumulates effective materials for the development of new vaccines.

Key words: porcine circovirus type 2 whole genome Cap protein molecular epidemiology genetic variation

收稿日期: 2021-11-05 出版日期: 2022-11-02

CLC:

S 855.3

基金资助: 浙江省重点研发计划项目(2019C02052-3)

通讯作者: 袁秀芳 E-mail: xulihua08@yeah.net;xiufangyuan@126.com

作者简介: 徐丽华（https://orcid.org/0000-0001-6601-6599），E-mail：xulihua08@yeah.net

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引用本文:

徐丽华,苏菲,李军星,余斌,叶十一,杨富文,邓利荣,毛慧敏,袁秀芳. 2016—2020年浙江地区猪圆环病毒2型分子流行病学分析[J]. 浙江大学学报（农业与生命科学版）, 2022, 48(5): 644-652.

Lihua XU,Fei SU,Junxing LI,Bin YU,Shiyi YE,Fuwen YANG,Lirong DENG,Huimin MAO,Xiufang YUAN. Molecular epidemiological analysis of porcine circovirus type 2 in Zhejiang Province from 2016 to 2020. Journal of Zhejiang University (Agriculture and Life Sciences), 2022, 48(5): 644-652.

链接本文:

https://www.zjujournals.com/agr/CN/10.3785/j.issn.1008-9209.2021.11.051 或 https://www.zjujournals.com/agr/CN/Y2022/V48/I5/644

表1 用于PCR的引物

表2 PCV2参考株信息

图1 猪圆环病毒2型PCR扩增结果1：阴性对照；2：阳性对照；3～20：临床样品；M： DL1000 DNA分子标志物。

表3 不同年份（2016—2020年）PCV2的阳性率

图2 基于PCV2全基因组的系统进化树★代表疫苗株；▲代表参考株。

图3 36株序列 ORF2 基因核苷酸序列同源性比较

图4 PCV2 Cap蛋白的氨基酸序列多重比对分析不同颜色框架区域代表不同亚型的主要突变位点；蓝色框架区域代表唯一糖基化位点。

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