牛结节性皮肤病病毒检测及蠓、蚊携带病毒调查

Detection of Lumpy Skin Disease Virus in Cattle and Investigation of Virus Carried by Midges and Mosquitoes

【目的】了解牛结节性皮肤病病毒(Lumpy skin disease virus,LSDV)在自然感染状况下病牛的带毒状况及其潜在的传播媒介种类,为牛结节性皮肤病防控提供参考依据。【方法】于2021年7月在云南省文山市采集疑似发病牛不同组织类型标本,2020年8月和2021年7月~2021年8月分别在云南省昆明市、文山市和双江县采集疑似LSDV感染的牛圈舍蠓、蚊标本,并进行形态学鉴定。采用实时荧光定量PCR方法检测发病牛不同组织及库蠓、库蚊标本中LSDV核酸;采用LSDV127基因特异引物对皮肤结节标本进行PCR扩增及测序;利用DNAStar和Mega 6.0等软件进行核苷酸序列相似性比对和系统进化分析。【结果】3头发病牛全血、血清、鼻拭子和皮肤结节标本中均检测到LSDV核酸,且皮肤结节标本中LSDV核酸扩增曲线信号最强;LSDV127基因序列分析结果显示,3份皮肤结节标本(17、18和19)的病毒核苷酸序列相似性为100%,与俄罗斯、泰国、越南和南非等LSDV分离株聚为一支,亲缘关系较近,核苷酸序列相似性在99.7%~100%之间。共采集鉴定条带库蠓、连斑库蠓和尖喙库蠓3种,库蠓标本4822只,库蚊标本150只。所有采集自发病牛圈舍的库蠓和库蚊标本LSDV核酸检测结果均为阴性。【结论】在自然感染LSDV的牛中,血液、鼻分泌物和皮肤结节等不同部位都可能为节肢动物媒介获得并传播LSDV提供很好的感染源;蠓、蚊等节肢动物媒介是否参与了当地LSDV的传播有必要进一步调查。

【Objective】This study was aimed to understand the virus-carrying status of infected cattle with Lumpy skin disease virus(LSDV)under natural infection and the potential vectors of the virus,so as to provide reference for the prevention and control of lumpy skin disease.【Method】The different tissue types of specimens were collected from cattle suspected infected with LSDV in Wenshan city,Yunnan province during July 2021.The midges and mosquitoes were collected from cattle feedlots which suspected infected with LSDV in Kunming,Wenshan city and Shuangjiang county,Yunnan province in August 2020 and July 2021 to August 2021,respectively,and the morphological identification was performed.The nucleic acid of LSDV in different tissue types of specimens of infected cattle and midges and mosquitoes were detected by Real-time quantitative PCR.PCR amplification and sequencing of the skin lesions specimens were carried out with specific primer pair of LSDV127 gene.The nucleotide homology and phylogenetic analysis were performed by DNAStar and Mega 6.0 softwares.【Result】The test of LSDV in whole blood,serum,nasal swab and skin lesion specimens of three infected cattle were positive,and the signals of LSDV amplification curve were the strongest in skin lesion specimens.In addition,the result of LSDV127 gene sequence analysis showed that three positive of nucleic acid of LSDV in skin lesion specimens were 100%homogeneity,and had the closer genetic relationship and clustered together with other LSDV which isolated from Russia,Thailand,Vietnam and South Africa,and showed 99.7%-100%homogeneity.A total of 4822 midges specimens were trapped and identified constituting 3 different species including C.tainanus,C.jacobsoni and C.oxystoma,as well as 150 specimens of mosquitoes.No specimens of midges and mosquitoes tested were positive for nucleic acid of LSDV.【Conclusion】These results suggested that the different parts such as blood,nasal secretions and skin lesions of cattle which naturally infected with LSDV might provide a plentiful source of pathogens for arthropod vectors to acquire and spread LSDV,and it was necessary to further investigate whether arthropod vectors such as midges and mosquitoes were involved in the transmission of local LSDV.