2022—2024年中国部分地区牛病毒性腹泻病毒分子检测及基因分型

doi:10.11843/j.issn.0366-6964.2025.10.050

Abstract

Abstract:

Bovine viral diarrhea virus (BVDV) is one of the major pathogens responsible for significant economic losses in cattle herds, particularly affecting the health and development of calves. This study aimed to establish a universal SYBR Green I-based real-time quantitative PCR (qPCR) assay for BVDV detection and to investigate the epidemiology, genotypes, and subtypes of BVDV in samples collected from free-range cattle herds in selected regions of China. A universal BVDV SYBR Green I qPCR assay was developed based on the 5′ untranslated region (5′UTR) sequences of various BVDV genotypes obtained from GenBank. Using this method, a total of 712 serum samples from free-range cattle, as well as tissue samples from calves exhibiting symptoms such as coughing, fever, and diarrhea collected between June 2022 and December 2024, were tested and subjected to phylogenetic analysis. Additionally, virus isolation, indirect immunofluorescence assay (IFA), and hematoxylin-eosin (HE) staining were performed on calf samples. The established BVDV SYBR Green I qPCR assay demonstrated high sensitivity, strong specificity, good linearity, and excellent reproducibility. The positive detection rate in serum samples was 19.10%, and the BVDV-24GS strain was successfully isolated. Phylogenetic analysis of the 5′UTR gene sequences revealed that the BVDV strains belonged to subtypes 1c, 1m, 1o, 1i, 1q, 1b, and 2. These findings indicate the successful development and application of a universal BVDV SYBR Green I qPCR assay, and the presence of varying degrees of BVDV infection among free-range cattle in parts of China. The identification of viral genotypes enriches the molecular epidemiological data of BVDV and provides a scientific basis for effective disease control, vaccine development, and optimization of immunization strategies.

Key words: bovine viral diarrhea virus, SYBR Green I real-time quantitative PCR, molecular detection, genotype identification

CLC Number:

S852.653

AN Lele, REN Xiaoting, LAN Qiuju, MO Yongli, CAO Xiaoan, DING Yulin, MA Zhongren, MA Xiaoxia. Molecular Detection and Genotyping of Bovine Viral Diarrhea Virus in Some Areas of China from 2022 to 2024[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(10): 5328-5334.

Figures/Tables 4

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References 21

1	HOUEH.Epidemiological features and economical importance of bovine virus diarrhoea virus (BVDV) infections[J].Vet Microbiol,1999,64(2-3):89-107. doi: 10.1016/S0378-1135(98)00262-4
2	阮文强,陈新诺,任玉鹏,等.不同来源的3株牛病毒性腹泻病毒对小鼠的致病性分析[J].畜牧兽医学报,2018,49(10):2232-2239. doi: 10.11843/j.issn.0366-6964.2018.10.019
	RUANW Q,CHENX N,RENY P,et al.Pathogenicity analysis of three bovine viral diarrhea viruses from different sources in mice[J].Acta Veterinaria et Zootechnica Sinica,2018,49(10):2232-2239. doi: 10.11843/j.issn.0366-6964.2018.10.019
3	COLLETTM S,LARSONR,BELZERS K,et al.Proteins encoded by bovine viral diarrhea virus: the genomic organization of a pestivirus[J].Virology,1988,165(1):200-208. doi: 10.1016/0042-6822(88)90673-3
4	ULLAHS,NOTSUK,SAITOA,et al.Direct TaqMan assay for the detection and genotyping of bovine viral diarrhea virus types 1 and 2[J].Arch Virol,2025,170(1):15. doi: 10.1007/s00705-024-06207-z
5	PASSLERT,DITCHKOFFS S,GIVENSM D,et al.Transmission of bovine viral diarrhea virus among white-tailed deer (Odocoileus virginianus)[J].Vet Res,2010,41(2):20. doi: 10.1051/vetres/2009068
6	GOMEZ-ROMERON,RIDPATHJ F,BASURTO-ALCANTARAF J,et al.Bovine viral diarrhea virus in cattle from Mexico: Current status[J].Front Vet Sci,2021,8,673577. doi: 10.3389/fvets.2021.673577
7	DENGM L,CHENN,GUIDARINIC,et al.Prevalence and genetic diversity of bovine viral diarrhea virus in dairy herds of China[J].Vet Microbiol,2020,242,108565. doi: 10.1016/j.vetmic.2019.108565
8	DE OLIVEIRAP S B,SILVA JÚNIORJ V J,WEIBLENR,et al.A new (old) bovine viral diarrhea virus 2 subtype: BVDV-2e[J].Arch Virol,2022,167(12):2545-2553. doi: 10.1007/s00705-022-05565-w
9	于江玮,程慧敏,林健,等.鸭瘟病毒TaqMan荧光定量PCR检测方法的建立及应用[J].畜牧兽医学报,2025,56(2):765-773. doi: 10.11843/j.issn.0366-6964.2025.02.026
	YUJ W,CHENGH M,LINJ,et al.Establishment and application of TaqMan fluorescent quantitative PCR detection method for duck plague virus[J].Acta Veterinaria et Zootechnica Sinica,2025,56(2):765-773. doi: 10.11843/j.issn.0366-6964.2025.02.026
10	王梦心. 黑龙江省BVDV分离鉴定及遗传进化分析[D]. 大庆: 黑龙江八一农垦大学, 2019.
	WANG M X. Isolation, identification and genetic evolution analysis of BVDV 1n Heilongjiang Province[D]. Daqing: Heilongjiang Bayi Agricultural University, 2019. (in Chinese)
11	谌婕,左之才,才冬杰,等.牛病毒性腹泻病毒全球基因型与亚型流行情况[J].浙江农业学报,2022,34(12):2622-2628.
	CHENJ,ZUOZ C,CAID J,et al.Global genotype and subgenotype prevalence of bovine viral diarrhea virus[J].Acta Agriculturae Zhejiangensis,2022,34(12):2622-2628.
12	舒鑫,董心仪,毛立,等.2株牛病毒性腹泻病毒基因2型的分离鉴定[J].畜牧与兽医,2018,50(10):105-109.
	SHUX,DONGX Y,MAOL,et al.Isolation and identification of two strains of bovine viral diarrhea virus 2[J].Animal Husbandry & Veterinary Medicine,2018,50(10):105-109.
13	林燕清. BVDV-2分离株XJ-04全基因组测序和抗BVDV单克隆抗体制备及初步应用[D]. 扬州: 扬州大学, 2011.
	LIN Y Q. Whole genome sequencing of BVDV-2 isolate XJ-04 and preparation and preliminary application of anti-BVDV monoclonal antibody[D]. Yangzhou: Yangzhou University, 2011. (in Chinese)
14	DENGM,JIS,FEIW,et al.Prevalence study and genetic typing of bovine viral diarrhea virus (BVDV) in four bovine species in China[J].PLoS One,2015,10(4):e0121718. doi: 10.1371/journal.pone.0121718
15	陈新诺,肖敏,阮文强,等.川藏地区牦牛牛病毒性腹泻病毒分子流行病学调查及分离鉴定[J].畜牧兽医学报,2018,49(3):606-613. doi: 10.11843/j.issn.0366-6964.2018.03.018
	CHENX N,XIAOM,RUANW Q,et al.Molecular epidemiological investigation and isolation of bovine viral diarrhea virus in yak in Sichuan-Tibet Plateau region[J].Acta Veterinaria et Zootechnica Sinica,2018,49(3):606-613. doi: 10.11843/j.issn.0366-6964.2018.03.018
16	ZHUJ,WANGC,ZHANGL,et al.Isolation of BVDV-1a, 1m, and 1v strains from diarrheal calf in china and identification of its genome sequence and cattle virulence[J].Front Vet Sci,2022,9,1008107. doi: 10.3389/fvets.2022.1008107
17	和彦良,李真光,苏玮玮,等.2018-2021年中国部分省份牛病毒性腹泻病毒的流行病学调查[J].中国兽医杂志,2023,59(10):9-15.
	HEY L,LIZ G,SUW W,et al.Epidemiological investigation of bovine viral diarrhea virus in some provinces of China from 2018 to 2021[J].Chinese Journal of Veterinary Medicine,2023,59(10):9-15.
18	夏九鲜,郑旺华,王磊,等.云南省部分地区猪感染BVDV流行病学调查[J].安徽农业科学,2023,51(19):92-94.
	XIAJ X,ZHENGW H,WANGL,et al.Epidemiological Investigation on BVDV 1nfection in pigs in some areas of Yunnan Province[J].Journal of Anhui Agricultural Sciences,2023,51(19):92-94.
19	张丁中,滕焯,程玉婷,等.青藏高原地区犊牦牛腹泻病原检测与分析[J].中国动物检疫,2024,41(4):1-5.
	ZHANGD Z,TENGZ,CHENGY T,et al.Detection and analysis of diarrhea pathogens in yak calves in Qinghai-Tibetan plateau region[J].China Animal Health Inspection,2024,41(4):1-5.
20	LESCOATC,PERROTTED,BARRYS,et al.Spatio-temporal distribution and international context of bovine viral diarrhoea virus genetic diversity in France[J].Vet Res,2024,55(1):129. doi: 10.1186/s13567-024-01377-9
21	ZHANGY Y,CHENGJ,GUOY,et al.Highly pathogenic bovine viral diarrhea virus BJ-11 unveils genetic evolution related to virulence in calves[J].Front Microbiol,2025,15,1540358. doi: 10.3389/fmicb.2024.1540358

质粒拷贝数/ (Copies·μL^-1) Copy No.	组内重复性试验Intra-group reproducibility tests		组间重复性试验Inter-group reproducibility tests
质粒拷贝数/ (Copies·μL^-1) Copy No.	$\bar x \pm s$	交异系数/% CV	$\bar x \pm s$	交异系数/% CV
1.80×10⁸	13.330±0.074	0.555	13.060±0.308	2.358
1.80×10⁷	17.120±0.036	0.210	16.860±0.298	1.767
1.80×10⁶	21.290±0.033	0.155	21.440±0.461	2.150
1.80×10⁵	25.470±0.095	0.371	25.470±0.214	0.840

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Molecular Detection and Genotyping of Bovine Viral Diarrhea Virus in Some Areas of China from 2022 to 2024

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