2020-2022年华中地区部分猪场猪圆环病毒3型的分子流行特征与遗传变异分析

doi:10.3864/j.issn.0578-1752.2024.03.014

中国农业科学 ›› 2024, Vol. 57 ›› Issue (3): 613-626.doi: 10.3864/j.issn.0578-1752.2024.03.014

• 畜牧·兽医 • 上一篇

2020-2022年华中地区部分猪场猪圆环病毒3型的分子流行特征与遗传变异分析

李博天¹(), 李春琪¹(), 刘国平¹(), 谢军²(), 曾攀¹, 赵润泽¹, 李桐¹, 裴洁², 郭利伟¹, 伍锐³, 谭磊¹

¹ 长江大学动物科学技术学院，湖北荆州 434020
² 湖北省动物疫病预防控制中心，武汉 430000
³ 四川农业大学动物医学院，成都 610000

收稿日期:2023-10-27 接受日期:2023-12-25 出版日期:2024-02-01 发布日期:2024-02-05
通信作者:
刘国平，E-mail：hhaaiieerr@163.com
谢军，E-mail：393396890@qq.com
联系方式: 李博天，E-mail：lbtt15972737737@163.com。李春琪，E-mail：lcq1447324386@163.com。李博天与李春琪为同等贡献作者。
基金资助:
湖北省重点研发项目(2023BBB045); 湖北省重点研发项目(2023BBB141); 石首市先行县“先进技术集成示范基地建设与定向攻关”揭榜项目(SS202311)

Molecular Epidemic Characteristics and Genetic Variation Analysis of Porcine Circovirus 3 in Some Pig Farms in Central China from 2020 to 2022

LI BOTIAN¹(), LI CHUNQI¹(), LIU GUOPING¹(), XIE JUN²(), ZENG PAN¹, ZHAO RUNZE¹, LI TONG¹, PEI JIE², GUO LIWEI¹, WU RUI³, TAN LEI¹

¹ College of Animal Science and Technology, Yangtze University, Jingzhou 434020 Hubei
² Hubei Provincial Center for Animal Disease Control and Prevention, Wuhan 430000
³ College of Veterinary Medcine, Sichuan Agricultural University, Chengdu 610000

Received:2023-10-27 Accepted:2023-12-25 Published:2024-02-01 Online:2024-02-05

摘要/Abstract

摘要：

【目的】通过系统性试验方案摸清我国华中部分地区猪圆环病毒3型（porcine circovirus 3）流行病学及遗传变异特点，为PCV3疫苗研究提供数据基础。【方法】对华中地区（湖北、湖南和河南）15个规模化养猪场的3 500份临床采集样品进行real-time PCR检测，分析不同猪群、不同组织器官、不同排毒量和对应流行病学症状特点关系。对部分阳性样本PCV3全基因组进行扩增、测序和分析。【结果】50.86%（1 780/3 500）被检样本为PCV3核酸阳性，不同发育阶段猪群PCV3均易感，保育猪、哺乳仔猪、生长育肥猪PCV3阳性率较高，分别为70.44%（498/707）、67.19%（596/887）、41.75%（177/424）。在不同组织器官和样品中，淋巴结、肺脏、产后胎盘样本PCV3阳性率为67.05%（59/88）、63.79%（74/116）、49.11%（55/112），血液、初乳样本PCV3阳性率分别为56.09%（502/895）、44.20%（278/629），且鼻拭子和唾液中均检出PCV3。出现猪繁殖障碍症状、厌食症状和呼吸障碍症状的猪群PCV3阳性率高，分别为44.43%（399/898）、36.43%（431/1183）、28.04%(233/831)。24条PCV3全基因组序列长度均为2000 nt，其全基因核苷酸序列同源性为98.4%—100%，与参考株PCV3全基因组的同源性为97.4%—99.5%。遗传进化分析结果显示，23株PCV3属于PCV3b亚型，1株属于PCV3a亚型。Rep氨基酸序列比对结果发现，PCV3-L2、L23和L14株分别在（N¹²⁴I)、（A¹⁸³E）和(V²⁴⁴I)出现独特变异位点；Cap蛋白氨基酸序列比对结果发现，PCV3-L15、L21、L3和L19株分别在(T⁴⁵P)、（R²K)、（R¹⁴K)和（F⁷L）出现独特变异位点。【结论】PCV3可感染不同发育阶段猪群，且分布于不同组织器官中；PCV3可通过垂直传播（如初乳和精液）和水平传播（如口鼻分泌物和唾液）感染猪群；PCV3感染可能与生猪繁殖障碍、呼吸障碍和多器官炎症和关节炎症密切相关。此外，被调查地区规模化猪场同时流行PCV3a和PCV3b两种亚型，其中PCV3b亚型为优势毒株。研究通过全基因组氨基酸序列分析发现部分独特变异位点位于Cap蛋白，这可能导致Cap蛋白赋予的免疫原性发生变化。对PCV3全基因序列的遗传进化做出详细阐述，为未来的PCV3疫苗研究奠定基础。

关键词: 猪圆环病毒3型, 病毒载量, 全基因组扩增, 遗传进化分析

Abstract:

【Objective】 The epidemiological and genetic variation characteristics of porcine circovirus 3 in some parts of central China were investigated through a systematic experimental protocol, which provided the data of basis for the research of PCV3 vaccine。【Methods】 A total of 3 500 clinical samples collected from 15 large-scale pig farms in central China (Hubei, Hunan and Henan) were detected by real-time PCR, and the relationship between different pig herds, different tissues and organs, different amount of detoxification and the characteristics of corresponding epidemiological symptoms was analyzed. The whole genome of PCV3 for some positive samples was amplified, sequenced and analyzed.【Result】 50.86% (1 780/3 500) of the tested samples were positive for PCV3 nucleic acid, pigs at different ages were susceptible to PCV3, and the positive rates of PCV3 were higher in nursing pigs, suckling piglets and growing finishing, which were 70.44% (498/707), 67.19% (596/887) and 41.75% (177/424), respectively. The positive rates of PCV3 in lymph nodes, lung, and postpartum placenta samples were 67.05% (59/88), 63.79% (74/116), and 49.11% (55/112), while the positive rates of PCV3 in blood and colostrum samples were 56.09% (502/895) and 44.20% (278/629), respectively, and PCV3 was detected in nasal swabs and saliva. The positive rates of PCV3 was high in the pig with symptoms of reproductive disorder, which were 44.43% (399/898), 36.43% (431/1183), and 28.04% (233/831), respectively. The length of the whole genome sequence of the 24 PCV3 was 2 000 nt, and the homology of the whole genome nucleotide sequence among the 24 sample strains was 98.4%-100%, and the homology of the whole genome with the reference strain PCV3 at home and abroad was 97.4%-99.5%. Genetic evolution analysis showed that 23 strains of PCV3 belonged to the PCV3b subtype and 1 strain belonged to the PCV3a subtype. The results of Rep amino acid sequence alignment showed that PCV3-L2, L23 and L14 strains had unique variation sites in (N¹²⁴I), (A¹⁸³E) and (V²⁴⁴I), respectively. The results of Cap protein amino acid sequence alignment showed that PCV3-L15, L21, L3 and L19 strains had unique variation sites in (T⁴⁵P), (R²K), (R¹⁴K) and (F⁷L), respectively.【Conclusion】 PCV3 can infect pigs at different ages and is distributed in different tissues and organs. PCV3 can infect pigs through vertical transmission (such as colostrum and semen) and horizontal transmission (such as oral and nasal secretions and saliva). PCV3 infection may be closely related to reproductive disorders, respiratory disorders, multiple organ inflammation and arthritis in pigs. In addition, both PCV3a and PCV3b subtypes were prevalent in large-scale pig farms in the surveyed area, and the PCV3b subtype was the dominant strain. In this study, genome-wide amino acid sequence analysis revealed that some unique variation sites were located in Cap protein, which may lead to changes in the immunogenicity conferred by Cap protein. In this study, the genetic evolution of the whole gene sequence of PCV3 is elaborated, aming to lay a foundation for future research on PCV3 vaccines.

Key words: porcine circovirus 3, viral load, whole genome amplification, genetic evolution analysis

李博天, 李春琪, 刘国平, 谢军, 曾攀, 赵润泽, 李桐, 裴洁, 郭利伟, 伍锐, 谭磊. 2020-2022年华中地区部分猪场猪圆环病毒3型的分子流行特征与遗传变异分析[J]. 中国农业科学, 2024, 57(3): 613-626.

LI BOTIAN, LI CHUNQI, LIU GUOPING, XIE JUN, ZENG PAN, ZHAO RUNZE, LI TONG, PEI JIE, GUO LIWEI, WU RUI, TAN LEI. Molecular Epidemic Characteristics and Genetic Variation Analysis of Porcine Circovirus 3 in Some Pig Farms in Central China from 2020 to 2022[J]. Scientia Agricultura Sinica, 2024, 57(3): 613-626.

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引物名称 Primer name	引物序列（5'-3'）Sequence (5'-3')	PCR 产物大小 Size of PCR product (bp)
PCV3-1IF	TAGTATTACCCGGCACCTCGG	816
PCV3-1IR	GCACCCTCCCCATTATAACCA	816
PCV3-2IF	CGGCTGCGCGGGAATTGAAGCTGT	491
PCV3-2IR	GGTAAACGCCCTCGCATGTGGGGA	491
PCV3-3IF	TGTCGCCCCCACCGAAGCATG	836
PCV3-3IR	GCCCGGCACCAAAATGAGACA	836
PCV3F	CACCCAGGACAAAGCCTCTT	89
PCV3R	AAGCAGTGCTCCCCATTGAA	89

序号 Number	毒株名称 Strain name	登录号 Accession number	地区 Location	基因型 Gene type	长度 Length (bp)
1	PCV3/KU-1605	KY996341.1	韩国South Korea	b	2000
2	PCV3-BR/RS/8	MF079254.1	巴西Brazil	b	2000
3	PCV3-Shandong-01	MH107161.1	中国China	b	2000
4	PCV3-China/JL17-42	MG870095.1	中国China	b	2000
5	PCV3/CN/Chongqing-148	KY075991.1	中国China	b	2000
6	PCV3/CN/Chongqing-147	KY075990.1	中国China	b	2000
7	PCV3/CN/Qinghai/2016	MG372486.1	中国China	b	2000
8	PCV3/CH/HB/HD/2017	MH101645.1	中国China	b	2000
9	PCV3 strain 2164	KX458235.1	美国America	b	2000
10	PCV3-AH-201706	MG868946.1	中国China	b	2000
11	PCV3-US/MO2015	KX778720.1	美国America	a	2000
12	CN/Liaoning-2017	MH177453.1	中国China	a	2000
13	PCV3/KU-1601	KY996337.1	韩国South Korea	a	2000
14	PCV3-BR/RS/6	MF079253.1	巴西Brazil	a	2000
15	PCV3-BJ-1	MF318453.1	中国China	a	2000
16	Hainan/2017	MG253684.1	中国China	a	2000
17	CN/FuJian-1215-2016	KY924474.1	中国China	a	2000
18	CN/FuJian-820-2016	KY924473.1	中国China	a	2000
19	CN/FuJian-318-2017	KY924475.1	中国China	a	2000
20	PCV3-Chian/GX2016-2	MF155642.1	中国China	a	2000
21	CN/Fujian-12/2016	KY075987.1	中国China	a	2000

不同猪场 Different pig farm		PCV3阳性数(份) Number of positives	样本数(份) Number of samples	PCV3阳性率 Positive rate
湖北 Hubei	武汉Wuhan	169	330	51.21%
	襄阳Xiangyang	107	228	46.93%
	荆州Jingzhou	97	199	48.74%
	恩施Enshi	148	265	55.85%
	鄂州Ezhou	107	212	50.47%
	随州Suizhou	252	398	63.32%
湖南 Hunan	常德Changde	203	320	63.44%
	湘潭Xiangtan	116	286	40.56%
	益阳 Yiyang	50	104	48.08%
	湘潭Xiangtan	91	157	57.96%
	永州Yongzhou	121	241	50.21%
河南 Henan	信阳Xinyang	112	193	58.03%
	兰考县Lankao	96	192	50.00%
	开封Kaifeng	101	185	54.59%
	新乡Xinxiang	106	190	55.79%
总计 Total		1876	3500	53.60%

不同阶段 Different stages	PCV3阳性数(份) Number of positives	样本数(份) Number of samples	PCV3阳性率 Positive rate
头胎母猪 Firstborn sow	114	366	31.15%
经产母猪Multiparous sows	243	675	36.00%
哺乳仔猪 Sucking pig	596	887	67.19%
保育猪 Nursery pig	498	707	70.44%
生长育肥猪 Growing-finishing pig	177	424	41.75%
种公猪 Boar	103	441	23.36%
总计 Total	1780	3500	50.86%

不同器官 Uneven organs	样品数(份) Number of samples	PCV3阳性数(份) Number of positives	PCV3阳性率 Positive rate
心脏 Heart	130	29	22.31%
肝脏 Liver	124	53	42.74%
脾脏 Spleen	84	20	23.81%
肺脏 Lungs	116	74	63.79%
肾脏 Kidney	195	77	39.49%
淋巴结 Lymph node	88	59	67.05%
胃 Stomach	100	36	36.00%
产后胎盘 Placenta	112	55	49.11%
总计 Total	949	403	22.31%

2020-2022年华中地区部分猪场猪圆环病毒3型的分子流行特征与遗传变异分析

Molecular Epidemic Characteristics and Genetic Variation Analysis of Porcine Circovirus 3 in Some Pig Farms in Central China from 2020 to 2022

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样品类型 Sample type	样品数(份) Number of samples	PCV3阳性数(份) Number of positives	PCV3阳性率 Positive rate
血液 Blood	895	502	56.09%
精液 Semen	658	206	31.31%
初乳 Colostrum	629	278	44.20%
外阴拭子 Vulvar swab	287	108	37.63%
鼻拭子 Nasal swab	431	112	25.99%
唾液 Saliva	400	93	23.25%
总计 Total	3300	1299	39.36%

症状 Symptoms	样品数(份) Number of samples	PCV3阳性数(份) Number of positives	PCV3阳性率 Positive rate
不吃 Skip	400	119	29.75%
少吃 Eat less	327	170	51.99%
未吃完 Not finished	456	142	31.14%
咳 Cough	831	233	28.04%
跛腿 Limp	102	25	24.51%
流产 Abortion	477	198	41.51%
早产 Premature birth	272	126	46.32%
产木乃伊Mummification	149	75	50.34%
腹泻 Diarrhea	83	4	4.82%
小猪发红 Sow flush	68	22	32.35%
死亡 Death	42	7	16.67%
总计 Total	3207	1121	34.95%

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