| Background Arboviruses are viruses that reproduce in blood-sucking insects such as mosquitoes and sandflies and transmit the virus to humans and animals through bites.There were 535 arboviruses registered in the International Arbovirus Center in 1992,of which 128 could cause disease to humans and animals,and in severe cases could cause death,and the disease burden was severe.At present,serious arbovirus diseases have been caused around the world,such as dengue fever and Zika virus disease.Therefore,arboviruses and related diseases are a public health issue of international concern.Genus Phlebovirus belongs to the order Bunyavirales,family Phenuiviridae.Many of which are sandfly-borne viruses,including Rift Valley fever virus,sandfly fever Naples virus(SFNV).At present,sandfly-borne Phleboviruses and their associated diseases are found primarily in countries along the Mediterranean coast,such as Italy,Turkey,and Cyprus,with no reports of sandfly-borne Phlebovirus in China or East Asia.Since the 1980 s,29 viruses belonging to 13 genera and seven families have been isolated from more than 1 million blood-sucking insect specimens collected in China.Although viral isolation and/or gene detection of sandfly-borne viruses were carried out on the collected sandfly specimens,no positive results were previously obtained.In 2011,severe fever with thrombocytopenia syndrome virus(SFTSV),a tick-borne virus known to cause fever and hemorrhaging,was first reported in China,and Guertu virus(GTV)was isolated from Dermacentor nuttalli ticks collected in Xinjiang,China.But these two viruses are tick-borne Phlebovirues.Shanxi province(N: 34°34′~40°44′,E: 110°14′~114°33′)is located in Central China,with an average elevation of 1000 m.Shanxi province is located to the Middle and lower reaches of the Yellow River.Shanxi province has a warm climate,dominated by mountains with an arid climate and less rain,which has contributed to the diversity of local vector insect species.In addition to mosquitoes,sandfly is an important blood-sucking insect vector in Shanxi Province.This is the first report of sandfly-borne Phlebovirus isolated in China,suggesting that sandfly-borne Phlebovirus not only is present in the Mediterranean Basin but is also emerging in China.Objective In order to understand the diversity and geographical distribution of arboviruses and arbovirus diseases,collecting various blood-sucking insect vectors to isolate viruses in Shanxi Province,which is not only beneficial to the prevention and control of arboviruses and related diseases in China,but also to public health in Asia and other countries is of great significance.Methods Blood-sucking insects were collected from feeding pens,such as chicken pens,and sheep pens,in Wuxiang county and Yangquan city,Shanxi Province in 2018.The specimens were classified according to their morphology under ice bath conditions.The specimens were stored in liquid nitrogen and transferred to the laboratory.BHK-21 cells(Mesocricetus auratus kidney cells)and C6/36 cells(Aedes albopictus cells)used in this study were isolated the viruses.The cells were cultured in incubators for three generations.The CPE of each well was observed each 12 h.The virus suspension was harvested when CPE occurred and stored at –80°C until further identification.Molecular biology methods was used to identification of viruses.The nucleotide and amino acid sequences were analyzed using software.Studies on the relationship between newly isolated viruses and humans and animals through animal experiments and serological experiments.Results In June 2018,4069 blood-sucking insects were collected in Wuxiang County,Shanxi Province.There were 3819 sandflies,including 3792 females and 27 males.In addition,250 mosquitoes were collected from the same breeding places,including Anopheles,and Armigeres.A new virus which is Wuxiang virus(17 strains)were isolated from sandfly specimens collected in the nature of Wuxiang County,Shanxi Province,in Central China.The virus can cause pathological changes of mammalian cells and morbidity and death of neonatal suckling mice.The titer of the virus is 108.09pfu/m L.The results of nucleotide and amino acid sequence analyses and genetic evolution analysis showed that the newly isolated virus belonged to a new member of the genus Phlebovirus,related to Toros virus(TORV)and Corfou virus(CFUV).In addition,the results of serum epidemiological studies showed that serum samples from local healthy residents were positive for Wuxiang virus-neutralizing antibodies.In June 2018,4070 blood-sucking insects were collected in Wuxiang County,Shanxi Province.There were 3996 sandflies,including 3904 females and 92 males.In addition,74 mosquitoes were collected from the same breeding places.10 strains of virus were isolated again from sandfly specimens collected in the natue of Yangquan city,Shanxi Province.The virus can cause pathological changes of mammalian cells and morbidity and death of neonatal suckling mice.The results of nucleotide and amino acid sequence analyses and genetic evolution analysis showed that the newly isolated viruses belonged to Wuxiang virus,related to Toros virus(TORV)and Corfou virus(CFUV).In addition,the results of serum epidemiological studies showed that human serum samples from local healthy residents and chicken serum samples were positive for Wuxiang virus-Ig M antibodies.Conclusions In this study,the genus Phlebovirus was isolated from the sandfly specimens collected in the nature of Shanxi Province,China,suggesting that:(1)Wuxiang virus isolated from the sandfly specimens is a new virus of the genus Phlebovirus,which provided new viral resources for viral biological phenotype and molecular genetic study;(2)This is the first report of Phlebovirus from natural sandflies in China;(3)Sandfly-borne Wuxiang viruses are isolated in Wuxiang and Yangquan,Shanxi Province and this reminds us that Wuxiang virus has a wide distribution in Shanxi Province and areas outside Shanxi Province;(4)It is worth investigating whether there is also sandfly-borne virus in areas outside Shanxi Province;(5)This explain provides new etiology basis of Fever of unknown causes at summer and autumn in Shanxi Province.This study suggests that the sandfly-borne virus is emerging in China,and detection of antibodies to the virus in human and animal serum samples suggests infection with the virus.Therefore,strengthening the detection and surveillance of diseases caused by sandfly-borne virus in humans and animals in China is of great public health significance for the prevention and control of diseases caused by sandfly-borne virus in China and even in East Asia.Limitations of this study(1)Collecting blood-sucking insect specimens only once,and investigating whether the virus persists for a long time requires continuous investigation;(2)The research on the relationship between sandfly-borne virus and local fever or encephalitis has not been performed;(3)The sandflies species of positive pools is Phlebotomus chinensis by the molecular biology identification,but others has not been performed;(4)Investigation and research on host animal species of sandfly-borne virus. |
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