Genome sequencing and phylogenetic analysis of Banna virus(genus Seadornavirus,family Reoviridae)isolated from Culicoides

In an investigation of blood-sucking insects and arboviruses, a virus(YN12243) was isolated from Culicoides samples collected in the Sino-Burmese border region of Yunnan Province, China. The virus caused cytopathic effect(CPE) in C6/36 cells and passaged stably. Polyacrylamide gel analysis showed that the genome of YN12243 was composed of 12 segments of double-stranded RNA(dsRNA), with a distribution pattern of 6-6. The nucleotide and amino acid sequences of the coding region(1.12 segments) were17,803 bp and 5,925 amino acids in length, respectively. The phylogenetic analysis of VP1 protein(RdRp) revealed that YN12243 belonged to genus Seadornavirus of family Reoviridae, and further analysis indicated that YN12243 belongs to the Banna virus(BAV) genotype A2. Additionally, YN12243 was located in the same evolutionary cluster as BAV strains isolated from different mosquito species, suggesting that the BAV isolated from Culicoides does not have species barriers. These results indicate that Culicoides can also be a vector for BAV. In view of the hematophagous habits of Culicoides on cattle, horses, deer, and other large animals, as well as the possibility of spreading and causing a variety of animal arboviral diseases, it is important to improve infection detection and monitor the BAV in large livestock.

Longitudinal Surveillance of Betacoronaviruses in Fruit Bats in Yunnan Province,China During 2009–2016

Previous studies indicated that fruit bats carry two betacoronaviruses,BatCoV HKU9 and BatCoV GCCDC1.To investigate the epidemiology and genetic diversity of these coronaviruses,we conducted a longitudinal surveillance in fruit bats in Yunnan province,China during 2009–2016.A total of 59(10.63%)bat samples were positive for the two betacorona-viruses,46(8.29%)for HKU9 and 13(2.34%)for GCCDC1,or closely related viruses.We identified a novel HKU9 strain,tentatively designated as BatCoV HKU9-2202,by sequencing the full-length genome.The BatCoV HKU9-2202 shared 83%nucleotide identity with other BatCoV HKU9 stains based on whole genome sequences.The most divergent region is in the spike protein,which only shares 68%amino acid identity with BatCoV HKU9.Quantitative PCR revealed that the intestine was the primary infection organ of BatCoV HKU9 and GCCDC1,but some HKU9 was also detected in the heart,kidney,and lung tissues of bats.This study highlights the importance of virus surveillance in natural reservoirs and emphasizes the need for preparedness against the potential spill-over of these viruses to local residents living near bat caves.

Detection and characterization of diverse alpha-and betacoronaviruses from bats in China

Bats have been implicated as important reservoir hosts of alpha- and betacoronaviruses. In this study, diverse coronaviruses(CoVs) were detected in 50 of 951(positive rate 5.3%) intestinal specimens of eight bat species collected in four provinces and the Tibet Autonomous Region of China by pan-coronavirus RT-PCR screening. Based on 400-nt RNA-dependent RNA polymerase(Rd RP) sequence analysis, eight belonged to genus Alphacoronavirus and 42 to Betacoronavirus.Among the 50 positive specimens, thirteen gave rise to CoV full-length Rd RP gene amplification for further sequence comparison, of which three divergent sequences(two from a unreported province) were subjected to full genome sequencing. Two complete genomes of betacoronaviruses(JTMC15 and JPDB144) and one nearly-complete genome of alphacoronavirus(JTAC2) were sequenced and their genomic organization predicted. The present study has identified additional numbers of genetically diverse bat-borne coronaviruses with a wide distribution in China. Two new species of bat CoV, identified through sequence comparison and phylogenetic analysis, are proposed.

Coexistence of multiple coronaviruses in several bat colonies in an abandoned mineshaft

Since the 2002–2003 severe acute respiratory syndrome(SARS) outbreak prompted a search for the natural reservoir of the SARS coronavirus, numerous alpha- and betacoronaviruses have been discovered in bats around the world. Bats are likely the natural reservoir of alpha- and betacoronaviruses, and due to the rich diversity and global distribution of bats, the number of bat coronaviruses will likely increase. We conducted a surveillance of coronaviruses in bats in an abandoned mineshaft in Mojiang County, Yunnan Province, China, from 2012–2013. Six bat species were frequently detected in the cave: Rhinolophus sinicus, Rhinolophus affinis, Hipposideros pomona, Miniopterus schreibersii, Miniopterus fuliginosus, and Miniopterus fuscus. By sequencing PCR products of the coronavirus RNA-dependent RNA polymerase gene(Rd Rp), we found a high frequency of infection by a diverse group of coronaviruses in different bat species in the mineshaft. Sequenced partial Rd Rp fragments had 80%–99% nucleic acid sequence identity with well-characterized Alphacoronavirus species, including Bt CoV HKU2, Bt CoV HKU8, and Bt CoV1,and unassigned species Bt CoV HKU7 and Bt CoV HKU10. Additionally, the surveillance identified two unclassified betacoronaviruses, one new strain of SARS-like coronavirus, and one potentially new betacoronavirus species. Furthermore, coronavirus co-infection was detected in all six bat species, a phenomenon that fosters recombination and promotes the emergence of novel virus strains. Our findings highlight the importance of bats as natural reservoirs of coronaviruses and the potentially zoonotic source of viral pathogens.

Serological Evidence of Bat SARS-Related Coronavirus Infection in Humans,China

Dear Editor,Severe acute respiratory syndrome coronavirus(SARSCo V)is the causative agent of the 2002–2003 SARS pandemic,which resulted in more than 8000 human infections worldwide and an approximately 10%fatality rate(Ksiazek et al.2003;Peiris et al.2004).The virus infects both upper airway and alveolar epithelial cells,resulting in mild to severe lung injury in

Role of ecological approaches to eliminating schistosomiasis in Eryuan County evaluated by system modelling

Background:Schistosomiasis was severely prevalent in Yunnan Province,and it is difficult to achieve its elimination by convention approaches due to complexity of the nature.We explored the comprehensive model to eliminate schistosomiasis in Eryuan County,Yunnan Province,the People’s Republic of China,through integration with the ecological protection programme in Erhai Lake,in order to promote an efficient elimination strategy.We expected that this model is able to be tailored to other local settings,which help achieve the goal of precisely eliminating the disease in Yunnan Province.Methods:Eryuan County of Yunnan Province was chosen as the study area,where the data on environmental protection activities in Erhai Lake and on the schistosomiasis control programme were collected through different departments of Erhai County government since 2015.System modelling was performed using system dynamics software to establish a simulation model in order to evaluate the effectiveness of intervention activities.Results:Ecological approaches to control schistosomiasis in Eryuan County consist of three major components:(i)implementing precise interventions to stop schistosomiasis transmission by means of controlling the source of infection,blocking the biological transmission chains and cutting off the route of disease transmission;(ii)employing ecological approaches to improve the co-effectiveness of environmental protection and schistosomiasis prevention in the study area;and(iii)strengthening the professional skills of personnel involving in the schistosomiasis control programme.Simulation results showed that this strategy could speed up the progress of schistosomiasis control programme moving from the control stage to the elimination stage.Conclusions:Ecological approaches implemented in schistosomiasis endemic areas of the Eryuan region are able to improve the co-effectiveness of environmental protection and schistosomiasis control,providing a new avenue for eliminating schistosomiasis thanks to the application of precise interventions.

Chevrier＇s Field Mouse(Apodemus chevrieri) and Père David＇s Vole(Eothenomys melanogaster) in China Carry Orthohepeviruses that form Two Putative Novel Genotypes Within the Species Orthohepevirus C

Hepatitis E virus(HEV)is the prototype of the family Hepeviridae and the causative agent of common acute viral hepatitis.Genetically diverse HEV-related viruses have been detected in a variety of mammals and some of them may have zoonotic potential.In this study,we tested 278 specimens collected from seven wild small mammal species in Yunnan province,China,for the presence and prevalence of orthohepevirus by broad-spectrum reverse transcription(RT)-PCR.HEV-related sequences were detected in two rodent species,including Chevrier’s field mouse(Apodemus chevrieri,family Muridae)and Père David’s vole(Eothenomys melanogaster,family Cricetidae),with the infection rates of 29.20%(59/202)and 7.27%(4/55),respectively.Further four representative full-length genomes were generated:two each from Chevrier’s field mouse(named Rd HEVAc14 and Rd HEVAc86)and Père David’s vole(Rd HEVEm40 and Rd HEVEm67).Phylogenetic analyses and pairwise distance comparisons of whole genome sequences and amino acid sequences of the gene coding regions showed that orthohepeviruses identified in Chinese Chevrier’s field mouse and Père David’s vole belonged to the species Orthohepevirus C but were highly divergent from the two assigned genotypes:HEV-C1 derived from rat and shrew,and HEV-C2 derived from ferret and possibly mink.Quantitative real-time RT-PCR demonstrated that these newly discovered orthohepeviruses had hepatic tropism.In summary,our work discovered two putative novel genotypes orthohepeviruses preliminarily named HEVC3 and HEV-C4 within the species Orthohepevirus C,which expands our understanding of orthohepevirus infection in the order Rodentia and gives new insights into the origin,evolution,and host range of orthohepevirus.

Human rabies in China:evidence-based suggestions for improved case detection and data gathering

Background:China still suffers heavily from rabies,although reported human cases continue to decrease year over year.There are far fewer laboratory-confirmed human cases than clinically diagnosed cases,which is a big problem that needs to be addressed.In this report,we summarize analyses of all specimens from human cases tested in our laboratory over the past 15 years,in order to promote laboratory diagnosis of rabies.Methods:From 2005 to 2019,a total of 271 samples from 164 suspected rabies cases were collected from local hospitals by the local Centers for Disease Control and Prevention(CDCs)in China.Saliva,cerebrospinal fluid(CSF),serum(blood)and urine were collected for ante-mortem diagnosis,and brain tissue,neck skin tissue and cornea were collected for post-mortem diagnosis.All of the specimens were tested by reverse transcription-polymerase chain reaction(RT-PCR),and brain tissues were also tested using fluorescent antibody test(FAT).The number of positive test results obtained using different fluids or tissues,and at different stages of the disease,were compared using a chi-square test and a more effective sampling program is recommended.Results:As the national reference laboratory for rabies surveillance in China,our laboratory has tested 271 samples from 164 suspected rabies cases collected by local CDCs since 2005.We found that saliva gave the highest number of positive test results(32%),compared with CSF and other fluids.We also found that serum or blood specimens collected in the last 3 days of life can test positive by RT-PCR.Conclusions:Serum or blood samples collected in the last 3 days of a patient’s life can be used to measure viral RNA,which means that serum samples,as well as saliva and CSF,can be used to detect viral RNA for anti-mortem diagnosis of rabies.Because of our findings,we have modified our“National Surveillance Project for Human Rabies”,by adding the collection and testing of serum samples from the end of the survival period.This will improve our national surveillance and laboratory diagnosis of human rabies.

Longitudinal surveillance of SARS-like coronaviruses in bats by quantitative real-time PCR

Dear Editor,The 2002–2003 severe acute respiratory syndrome coronavirus（SARS-CoV）（Drosten et al.,2003）caused human pandemics that began in China and spread globally.Subsequently,

Schistosomiasis Surveillance--China,2015−2018

Background The prevalence of schistosomiasis in China is at the lowest level in history,and sentinel surveillance sites of schistosomiasis have fully covered all epidemic areas of China.This article has analyzed the surveillance data for the last four years and can help guide the next stage of surveillance work at the national level,including the scope of surveillance,surveillance content,and surveillance methods,etc.Methods Data from the National Schistosomiasis Surveillance System were collected.The infection rate of schistosomiasis among the human population,livestock,snails,and the change of the breeding area of snails in sentinel surveillance sites for four consecutive years were analyzed,and the trends in schistosomiasis prevalence in surveillance sites were determined.Results The prevalence of schistosomiasis in all sentinel surveillance sites of China showed downward trends from 2015 to 2018 with infection rates decreasing in this time period for local human population(0.05%to 0.00%),the floating population(0.020% to 0.003%),and cattle(0.037% to 0.000%).No infected snails were found during the period.From 2015 to 2018,the total area of newly found habitats of snails(Oncomelania hupensis,O.hupensis)were 34,730 m^(2),1,367,694 m^(2),18,944 m^(2),and 50,420 m^(2),and the total area of re-emergent habitats of snails(O.hupensis)were 822,194 m^(2),1,391,779 m^(2),1,516,292 m^(2),and 1,750,558 m^(2),respectively.Conclusions and Implications for Public Health Practice The prevalence of schistosomiasis in human and livestock is going down and is sustained at a very low level,but new and re-emerging habitats of O.hupensis are new challenges regardless of the achievement of schistosomiasis control.Therefore,two actions are necessary to effectively further reduce the transmission risk of schistosomiasis:1)taking effective measures to control the source of infection of schistosomiasis;and 2)reducing the breeding areas of O.hupensis.