黑龍江地區(qū)鯉春病毒血癥病毒的分離與基因型分析
發(fā)布時間：2019-04-27 20:48
【摘要】：對2015—2016年黑龍江不同地區(qū)的40個養(yǎng)殖場送檢的鯉(Cyprinus carpio)進(jìn)行鯉春病毒血癥病毒(spring viremia of carp virus,SVCV)的細(xì)胞培養(yǎng)分離、PCR鑒定、病毒滴度測定、病毒表面糖蛋白(glycoprotein,G)氨基酸序列聚類分析及基因分型研究。細(xì)胞培養(yǎng)結(jié)果顯示,來自4個不同養(yǎng)殖場的鯉組織樣本能夠感染鯉上皮細(xì)胞(epithelioma papulosum cyprini,EPC)產(chǎn)生典型細(xì)胞病變(cytopathic effect,CPE),收集病毒懸液分別稱為Shlj1~Shlj4。PCR鑒定結(jié)果表明,該4株病毒均為SVCV。病毒滴度實驗測算出SVCV Shlj 1~Shlj 4的滴度分別為10~(6.28)、10~(6.88)、10~(7.57)和106.38 TCID50/mL。Shlj的糖蛋白基因核苷酸序列的聚類分析和遺傳進(jìn)化分析結(jié)果顯示,Shlj 1~Shlj 4與Gen Bank收錄的中國參考株A2、BJ0505-2和美國參考株USA、212364聚為一簇,同源性為98.4%~99.8%;Shlj 1~Shlj 4毒株之間的糖蛋白核苷酸序列相似性在98.6%~99.8%,其中Shlj 3與美國SCVC毒株USA、212364具有最高的核苷酸相似性(99.8%),Shlj 2與英國參考毒株880163具有最低的相似性(88.0%)。糖蛋白氨基酸序列比對結(jié)果顯示,Shlj4中氨基酸突變最多,與另3個毒株差異較大。基因型分析結(jié)果顯示,Shlj 1~Shlj 4均為基因Ia型。本研究結(jié)果表明,黑龍江地區(qū)2015—2016年間的SVCV檢出率約為10%,并且來源于不同養(yǎng)殖場的病毒分離株的核酸序列呈現(xiàn)不同程度的差異,該結(jié)果進(jìn)一步證明SVCV毒株在中國不同的鯉養(yǎng)殖環(huán)境中正在不斷地進(jìn)化。
[Abstract]:Cell culture, PCR identification, viral titer determination and surface glycoprotein (glycoprotein,) of Cyprinus Carpio (Cyprinus carpio) from 40 farms in different areas of Heilongjiang Province from 2015 to 2016 were carried out by culture and isolation of Cyprinus Carpio viremia virus (spring viremia of carp virus,SVCV), PCR identification, viral titer determination and surface glycoprotein (glycoprotein,). G) amino acid sequence cluster analysis and genotyping. The results of cell culture showed that the common carp tissue samples from four different farms could infect common carp epithelial cells (epithelioma papulosum cyprini,EPC) to produce typical cytopathic effects (cytopathic effect,CPE). The results of Shlj1~Shlj4.PCR identification showed that the virus suspensions were collected respectively. The four strains were all SVCV.. The viral titers of SVCV Shlj 1~Shlj-4 were determined to be 106.28,106.88,107.57and 106.38 TCID50/mL.Shlj respectively by cluster analysis and genetic evolution analysis. Shlj 1~Shlj 4 was clustered with Chinese reference strain A2, BJ0505 / 2 and American reference strain USA,212364, and the homology was 98.4% and 99.8%, respectively, compared with that of Chinese reference strain A2, BJ0505 / 2 and American reference strain USA,212364. The nucleotide sequence similarity of glycoprotein among Shlj 1~Shlj 4 strains was 98.6% and 99.8% respectively. Among them, Shlj 3 had the highest nucleotide similarity with American SCVC strain USA,212364 (99.8%). The lowest similarity was found between Shlj 2 and British reference strain 880163 (88.0%). The results of amino acid sequence alignment showed that amino acid mutation was the most in Shlj4, which was significantly different from that of the other three strains. Genotype analysis showed that all Shlj 1~Shlj-4 were genotype Ia. The results showed that the detection rate of SVCV in Heilongjiang region from 2015 to 2016 was about 10%, and the nucleic acid sequences of virus isolates from different farms showed different degrees of difference. The results further demonstrate that the SVCV strain is evolving continuously in different culture environments of Cyprinus Carpio in China.
【作者單位】： 中國水產(chǎn)科學(xué)研究院黑龍江水產(chǎn)研究所;上海海洋大學(xué)水產(chǎn)與生命學(xué)院;
【基金】：黑龍江省應(yīng)用技術(shù)研究與開發(fā)計劃項目(GA13B401)
【分類號】：S943
本文編號：2467299
[Abstract]:Cell culture, PCR identification, viral titer determination and surface glycoprotein (glycoprotein,) of Cyprinus Carpio (Cyprinus carpio) from 40 farms in different areas of Heilongjiang Province from 2015 to 2016 were carried out by culture and isolation of Cyprinus Carpio viremia virus (spring viremia of carp virus,SVCV), PCR identification, viral titer determination and surface glycoprotein (glycoprotein,). G) amino acid sequence cluster analysis and genotyping. The results of cell culture showed that the common carp tissue samples from four different farms could infect common carp epithelial cells (epithelioma papulosum cyprini,EPC) to produce typical cytopathic effects (cytopathic effect,CPE). The results of Shlj1~Shlj4.PCR identification showed that the virus suspensions were collected respectively. The four strains were all SVCV.. The viral titers of SVCV Shlj 1~Shlj-4 were determined to be 106.28,106.88,107.57and 106.38 TCID50/mL.Shlj respectively by cluster analysis and genetic evolution analysis. Shlj 1~Shlj 4 was clustered with Chinese reference strain A2, BJ0505 / 2 and American reference strain USA,212364, and the homology was 98.4% and 99.8%, respectively, compared with that of Chinese reference strain A2, BJ0505 / 2 and American reference strain USA,212364. The nucleotide sequence similarity of glycoprotein among Shlj 1~Shlj 4 strains was 98.6% and 99.8% respectively. Among them, Shlj 3 had the highest nucleotide similarity with American SCVC strain USA,212364 (99.8%). The lowest similarity was found between Shlj 2 and British reference strain 880163 (88.0%). The results of amino acid sequence alignment showed that amino acid mutation was the most in Shlj4, which was significantly different from that of the other three strains. Genotype analysis showed that all Shlj 1~Shlj-4 were genotype Ia. The results showed that the detection rate of SVCV in Heilongjiang region from 2015 to 2016 was about 10%, and the nucleic acid sequences of virus isolates from different farms showed different degrees of difference. The results further demonstrate that the SVCV strain is evolving continuously in different culture environments of Cyprinus Carpio in China.
【作者單位】： 中國水產(chǎn)科學(xué)研究院黑龍江水產(chǎn)研究所;上海海洋大學(xué)水產(chǎn)與生命學(xué)院;
【基金】：黑龍江省應(yīng)用技術(shù)研究與開發(fā)計劃項目(GA13B401)
【分類號】：S943
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