Isolation, identification and genome-wide analysis of Spring viremia of carp virus in Sichuan Province of China

doi:10.3969/j.issn.1004-1524.20240655

Acta Agriculturae Zhejiangensis ›› 2025, Vol. 37 ›› Issue (7): 1430-1440.DOI: 10.3969/j.issn.1004-1524.20240655

• Animal Science • Previous Articles Next Articles

Isolation, identification and genome-wide analysis of Spring viremia of carp virus in Sichuan Province of China

QI Yingji^a(), LIU Shuya^a, LAN Hanyun^a, WANG Xuemei^a, SUN Tiantian^a, LI Shuhan^a, LI Qiunan^a, HUANG Xiaoli^b, GENG Yi^a, CHEN Defang^b, OUYANG Ping^a^,^*()

College of Veterinary Medicine; b. College of Animal Science and Technology, Sichuan Agricultural University, Chengdu 611130, China

Received:2024-07-19 Online:2025-07-25 Published:2025-08-20

Abstract

Abstract:

In March 2022, large scale death of carp from a farm in Leshan City, Sichuan Province, suddenly developed acute mortality with a cumulative mortality rate of 80%. To study the etiology and epidemiological pattern of this disease, the diseased material was subjected to autopsy, pathological examination, bacteriological examination, PCR identification, virus isolation and identification and phylogenetic analysis. The results showed that histopathological lesions were mainly in the gills and intestines, intestinal congestion and hemorrhage, with necrosis and shedding of epithelial cells; and hyperplasia, necrosis and shedding of epithelial cells in gill lamellae. Based on pathological diagnosis, it was preliminarily determined that the disease was caused by Spring viremia of carp virus(SVCV) infection. RT-PCR detection results showed that the G protein gene sequence of SVCV was amplified in the visceral tissues and cell culture virus solution of naturally infected carp and artificially infected carp. Electron microscopic observation of ultrathin sections of cell cultures virus solution showed that the viral particles were 80-150 nm long and 40-60 nm wide, and were bullet-shaped. Artificially infected healthy carp showed clinical symptoms similar to those of naturally infected carp, with a cumulative mortality rate of 65% in the infected group. A virus strain was isolated from epithelioma papulosum cyprini cell(EPC), and whole genome sequencing and sequence analysis were performed on the isolate, which was named SVCV-2022 after sequencing conformation of the highest homology with SVCV. Combining the epidemiological and pathological characteristics of the disease, the isolation and identification of the virus, the results of the artificial infection test and transmission electron microscopy, the pathogen of the epidemic was identified as SVCV.

Key words: Spring viremia of carp virus, carp, cumulative mortality rate, phylogenetic analysis

CLC Number:

S941.416

QI Yingji, LIU Shuya, LAN Hanyun, WANG Xuemei, SUN Tiantian, LI Shuhan, LI Qiunan, HUANG Xiaoli, GENG Yi, CHEN Defang, OUYANG Ping. Isolation, identification and genome-wide analysis of Spring viremia of carp virus in Sichuan Province of China[J]. Acta Agriculturae Zhejiangensis, 2025, 37(7): 1430-1440.

Figures/Tables 11

Fig.1 Clinical symptoms and necropsy lesions of infected carp A-1, Superficial hemorrhage; A-2,Gill filaments pale with punctate hemorrhage; A-3, Hemoid ascites, mesenteric punctate hemorrhage; A-4, Redness and swelling in the anus and punctate hemorrhage in the caudal fin; A-5, Intestinal congestion, redness and swelling; B-1, Exophthalmos; B-2, Gill filaments pale; B-3, Spleen enlargement, ascites; B-4, Abdominal distension, redness and swelling in the anus; B-5, Punctate hemorrhage in swim bladder.

Fig.2 Histopathological damage of naturally infected SVCV carp(HE staining) A, Degeneration and necrosis of renal tubular epithelial cells( ), homogeneous, red-stained, translucent drops appeared in the lumen( ); B, Hepatic hemorrhage, vacuolar degeneration of hepatocytes; C, Nucleolysis, nuclear solidification and nuclear fragmentation of hepatocytes( ); D,Splenic hemorrhage, distribution of large quantities of hemosiderin, reticuloendothelial cell proliferation; E, Intestinal congestion and hemorrhage, necrosis and shedding of intestinal epithelial cells; F, Hyperplasia and necrosis of epithelial cells in gill lamellae( ),epithelial cell ballooning( ).

Fig.3 RT-PCR amplification of the SVCV G protein gene A, The first round PCR product(714 bp); B, The second round PCR product (606 bp); M, DL 2 000 DNA marker; “+”, Positive control; “-”, Negative control; 1, 2, 3 and 4, Carp sent for inspection.

Fig.4 Epithelioma papulosum cyprini cells inoculated with tissue homogenate of diseased carp A, Cell morphology of control group; B, Cell morphology of CPE appeared 48 h later.

Fig.5 RT-PCR identification of cell cultures A, The first round PCR product(714 bp); B, The second round PCR product (606 bp); M, DL2 000 DNA marker; 1, Negative control; 2, Positive control; 3, Control group; 4, Cell culture virus.

Fig.6 Electron microscopic observation results A, Apoptosis and a large number of virions around the cells (white arrow points); B, Virions aggregated in the cytoplasm.

Fig.7 Cumulative survival rate of carp after infection with Spring viremia of carp virus

Fig.8 Schematic representation of the genome structure of SVCV-2022(0P495631.1) isolate

Table 1 Gene information of virus strain for phylogenetic tree

编号 Number	GenBank登录号 GenBank accession number	分离株 Isolate	来源地 Source location	宿主名称 Host name	长度 Length/bp
1	DQ097384.2	A1	中国China	Cyprinus carpio	11 100
2	DQ491000.1	A2	中国China	—	10 990
3	EU177782.1	BJ0505-2	中国北京Beijing,China	—	11 047
4	MT675953.1	shlj4	中国China	Cyprinus carpio	11 029
5	KR012465.1	SH140501	中国上海Shanghai,China	金鱼Goldfish	11 029
6	KR012466.1	SH140502	中国上海Shanghai,China	金鱼Goldfish	11 029
7	KR012467.1	SH140503	中国上海Shanghai,China	金鱼Goldfish	11 029
8	KR012468.1	SH140522	中国上海Shanghai,China	鲤鱼Common carp	11 029
9	KT321307.1	SH150514	中国上海Shanghai,China	金鱼Goldfish	11 029
10	KY475636.1	SH160901	中国China	草鱼Grass carp	11 029
11	KJ513477.1	SVCV-265	中国China	锦鲤Koi	11 029
12	KU230365.1	202238	中国香港Hong Kong,China	Cynops orientalis	10 991
13	MG663514.1	ADC-SVC2016- 1	韩国Republic of Korea	Cyprinus carpio	11 034
14	MG663513.1	ADC-SVC2016-3	韩国Republic of Korea	Micropterus salmoides	11 034
15	MG663512.1	ADC-SVC2016-5	韩国Republic of Korea	Cyprinus carpionudus	11 029
16	NC_002803.1	ATCC VR-1390	南斯拉夫Yugoslavia	Cyprinus carpio	11 019
17	AJ318079.1	Fijan reference strain	南斯拉夫Yugoslavia	鲤Carp	11 019

Fig.9 Phylogenetic tree construction based on whole genome series of SVCV-2022

Fig.10 Similarity analysis of nucleotide sequences between isolate and reference strains of SVCV-2022(OP495631)

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Isolation, identification and genome-wide analysis of Spring viremia of carp virus in Sichuan Province of China

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