The molecular mechanism of efficient transmission of plant viruses in variable virus-vector-plant interactions

Plant viruses are mainly transmitted by insect vectors in the non-persistent,semi-persistent,or persistent modes.In the non-persistent mode,plant viruses are retained in the stylets of their insect vectors.In the semi-persistent mode,plant viruses are carried to vector foreguts or salivary glands,but they cannot spread to salivary glands.In the persistent mode,plant viruses are retained in vector guts and can spread to salivary glands.In the non-persistent and semi-persistent modes,plant viruses are retained for a short time and cannot enter the hemolymph of insect vectors,whereas in the persistent mode,plant viruses are retained for a relatively long time and can be found in the hemolymph.Here,we reviewed recent studies that uncovered molecular mechanisms of how plant viruses manipulate host traits for efficient transmission by insect vectors.Normally,plants that are infected with viruses,regardless of the transmission mode,tend to release more attractive volatiles to vectors.However,plant defensive systems are regulated differently by viruses in these three modes.In the non-persistent mode,virus infections significantly induce plant defense responses,which probably trigger vectors(e.g.,winged aphids)to disperse and transmit viruses in a short time.In the semi-persistent mode,virus infections frequently suppress plant defense responses,resulting in an increase of vector population and facilitating viral transmissions during vector outbreaks.In the persistent mode,virus infections reduce plant defense responses and manipulate plant traits to become suitable feeding sites in a relatively long period of time.Understanding the underlying mechanisms of virus–vector–plant interactions will lay a foundation for preventing virus transmission.

Impact of a novel Rickettsia symbiont on the life history and virus transmission capacity of its host whitefly(Bemisia tabaci)

Rickettsia consists of some of the most prevalent symbionts of insects and often plays a significant role in the biology of its hosts.Recently,a maternally inherited Torix group Rickettsia,provisionally named as RiTBt,was recorded in a species of notorious pest whitefly,tentatively named as Asia II 1,from the Bemisia tabaci complex.The role of this Rickettsia in the biology of its host is unknown.Here we investigated the impact of RiTBt on the performance and virus transmission capacity of Asia II 1.RiTBt did not significantly affect the life history parameters of the whitefly when the host insect was reared on tobacco,tomato,and cotton,three host plants with relatively low,medium and high suitability to the whitefly.Intriguingly,RiTBt slightly enhanced whitefly transmission of cotton leaf curl Multan virus(CLCuMuV),a virus that is transmitted by the whitefly in the field and has caused extensive damage to cotton production.Specifically,compared with whiteflies without RiTBt,following a 48 h virus acquisition whiteflies with RiTBt had higher titer of virus and showed higher efficiency of virus transmission.A rickettsial secretory protein BtR242 was identified as a putative virus-binding protein,and was observed to interact with the coat protein of CLCuMuV in vitro.Viral infection of the whitefly downregulated gene transcript levels of the BtR242 gene.These observations indicate that RiTBt has limited impact on the biology of the Asia II 1 whitefly,and whether this symbiont has functions in the biology of other host whiteflies warrants future investigation.

Possible Transmission of Irkut Virus from Dogs to Humans

Lyssaviruses, including Rabies virus, Duvenhage virus, European bat lyssavirus 1, European bat lyssavirus 2, Australian bat lyssavirus, and Irkut virus （IRKV）, have caused human fatalities, but infection of IRKV in dogs has not been previously reported. In China, a dead dog that previously bit a human was determined to be infected with IRKV. Pathogenicity tests revealed that IRKVs can cause rabies-like disease in dogs and cats after laboratory infection. The close relationship between humans and pets, such as dogs and cats, may generate a new spillover-spreading route for IRKV infection. Therefore, additional attention should be paid to trans-species infection of IRKV between bats and dogs or dogs and humans through investigation of the prevalence and circulation patterns of IRKV in China.

Insect-specific RNA virus affects the stylet penetration activity of brown citrus aphid(Aphis citricidus)to facilitate its transmission

Sap-sucking insects often transmit plant viruses but also carry insect viruses,which infect insects but not plants.The impact of such insect viruses on insect host biology and ecology is largely unknown.Here,we identified a novel insect-specific virus carried by brown citrus aphid(Aphis citricidus),which we tentatively named Aphis citricidus picornavirus(AcPV).Phylogenetic analysis discovered a monophyletic cluster with AcPV and other unassigned viruses,suggesting that these viruses represent a new family in order Picornavirales.Systemic infection with AcPV triggered aphid antiviral immunity mediated by RNA interference,resulting in asymptomatic tolerance.Importantly,we found that AcPV was transmitted horizontally by secretion of the salivary gland into the feeding sites of plants.AcPV influenced aphid stylet behavior during feeding and increased the time required for intercellular penetration,thus promoting its transmission among aphids with plants as an intermediate site.The gene expression results suggested that this mechanism was linked with transcription of salivary protein genes and plant defense hormone signaling.Together,our results show that the horizontal transmission of AcPV in brown citrus aphids evolved in a manner similar to that of the circulative transmission of plant viruses by insect vectors,thus providing a new ecological perspective on the activity of insect-specific viruses found in aphids and improving the understanding of insect virus ecology.

Identification,Isolation and Characterization of GaCyPI Gene in Gossypium arboreum under Cotton Leaf Curl Virus Disease Stress

Pakistan is facing the threat of Cotton Leaf Curl Virus(CLCuV)which is transmitted through whitefly to cotton crop.Molecular mechanism of leaf epicuticular wax protects the plants from different pathogens including insect attack and disease transmission.Objective of current study is the isolation and characterization of a wax related gene GaCyPI from Gossypium arboreum under CLCuV infection.A fragment of 475 bp was isolated from the total RNA and 3’and 5’RACE-PCR products were arranged by overlapping the extended sequences at both the ends.Deduced protein sequence of GaCyPI showed homology with Cyclophilin cis-trans isomerase gene of Gossypium ramondii and Gossypium barbadanse.Multiple sequence alignment also revealed homology among the coding sequences of same gene.GaCyPI protein comprised of 173 amino acids and ORF finder revealed the 69 bases upstream at 5’while 350 bp at 3’UTR.InterProScan revealed that it belongs to Cyclophilin-type peptidyl prolyl cis/trans isomerase(PPIase)family.Active sites are visible at specific amino acid positions and 3D structure was stable in Ramachandran plot.Prosa server showed protein residues have average 3D-1D score>=0.2 and Z-Score was−6.74.Phylogenetic analysis revealed that G.raimondii is the closest species that shares the same sequence.Hence,GaCyPI has strong role in plants’epicuticular wax and its genetic transformation may protect the cotton from whitefly which transmits CLCuV.

Cloning and Identification of S Gene from Chinese Isolate TH-98 of Transmissible Gastroenteritis Virus

Chinese isolate of transmissible gastroenteritis virus(TGEV)was propagated and harvested in swine testicle(ST)cells.Two pairs of primers were designed according to the published sequence with Oligo 4.1 and DNasis softwares.The products of RT-PCR were named Sa and Sb,of 2.3kb and 2.1kb respectively.Sa was inserted in EcoR I and Kpn I sites after Sb was cloned in Kpn I and Pst I sites of the same pUC18 plasmid.The recombinant designated pUC-S was verified and analyzed by corresponding restriction endonuclease(RE)and nested PCR on the basis of genetic sites of S gene and physical map of pUC18 plasmid,which was identified as S gene from Chinese isolate of TGEV.

Cloning and Homology Comparison of S Gene for Isolate TH-98 of Porcine Transmissible Gastroenteritis Virus

TH-98 isolate of transmissible gastroenteritis virus (TGEV) was propagated and harvested on swine testicle (ST) monolayer cell. Two pairs of primers were designed to amplify S gene by RT-PCR according to the published sequence of TGEV'S gene cDNA with Oligo version 4.1 and DNasis software. The products of PCR were named Sa and Sb, of 2.3 kb and 2.1 kb respectively. Sa was inserted in EcoR I and Kpn I sites after Sb was cloned in Kpn I and Pst I multiple cloning sites of the same pUC18 plasmid. The recombinant pUC-S plasmid was identified and analyzed by corresponding restriction endonuclease and nested PCR on the basis of the genetic sites of S gene and pUC18 plasmid, which was identified as S gene of TGEV. Recombinant pUC-S was sequenced and analyzed in comparison with the other strains. Gene sequence comparison indicated that TH-98 shared 99, 97, 98, 97 and 94% identities with Purdue-115(US), Miller(US), TO14(Japan), FS772(British), 96-1933(British), respectively, their deduced amino acid homology was 99, 97, 97, 96 and 93% correspondingly. In addition, the analysis report verified that pUC-S owned a complete open reading frame (ORF) including initiation codon, signal sequences, remaining sequences and termination codon as well. Therefore, the results affirmed that S gene of TGEV TH-98 was extremely conservative.

Effects of Different Alkaloids in Coptis chinensis on Inhibiting TGEV Proliferation

[Objectives]To study the effects of different alkaloids in Coptis chinensis on inhibiting the proliferation of Transmissible gastroenteritis virus(TGEV).[Methods]The components and content of the main alkaloids in the extract of C.chinensis were analyzed.The main alkaloids were selected as drugs to inhibit the proliferation of TGEV.The maximum non-toxic concentration of Columbamine,Jatrorrhizine,Epiberberine,Coptisine,Palmatine,and Berberine was screened.The protective rate of each drug on TGEV-infected ST cells was determined,and the transcriptional inhibitory effect of the drug on TGEV N gene was detected by fluorescent quantitative PCR.[Results]The extract of C.chinensis mainly contains 6 alkaloids:Columbamine,Jatrorrhizine,Epiberberine,Coptisine,Palmatine,and Berberine,accounting for 2.03%,8.88%,9.21%,15.07%,14.63%,and 50.18%,respectively.In the range of the safe concentration,Jatrorrhizine,Palmatine,and Coptisine had better protective effects on ST cells infected with TGEV;compared with the Columbamine group,the cell protection rate was significantly different(P<0.05);compared with the Berberine group,the difference was extremely significant(P<0.01).The Coptisine and Palmatine groups had significant inhibitory effects on the transcription of TGEV N gene,and the difference was extremely significant compared with the virus group(P<0.05).[Conclusions]Jatrorrhizine and Palmatine in C.chinensis are the main components to inhibit the proliferation of TGEV.

All-trans retinoic acid alleviates transmissible gastroenteritis virus-induced intestinal inflammation and barrier dysfunction in weaned piglets

Background Transmissible gastroenteritis virus(TGEV)is one of the main pathogens causing severe diarrhea of pig-lets.The pathogenesis of TGEV is closely related to intestinal inflammation.All-trans retinoic acid(ATRA)is the main active metabolite of vitamin A,which has immunomodulatory and anti-inflammatory properties.However,it is unclear whether ATRA can alleviate TGEV-induced intestinal inflammation and barrier dysfunction in piglets.This study aimed to investigate the effects of ATRA on growth performance,diarrhea,intestinal inflammation and intesti-nal barrier integrity of TGEV-challenged piglets.Methods In a 19-d study,32 weaned piglets were randomly divided into 4 treatments:Control group(basal diet),TGEV group(basal diet+TGEV challenge),TGEV+ATRA5 group(basal diet+5 mg/d ATRA+TGEV challenge)and TGEV+ATRA15 group(basal diet+15 mg/d ATRA+TGEV challenge).On d 14,piglets were orally administered TGEV or the sterile medium.Results Feeding piglets with 5 and 15 mg/d ATRA alleviated the growth inhibition and diarrhea induced by TGEV(P<0.05).Feeding piglets with 5 and 15 mg/d ATRA also inhibited the increase of serum diamine oxidase(DAO)activ-ity and the decrease of occludin and claudin-1 protein levels in jejunal mucosa induced by TGEV,and maintained intestinal barrier integrity(P<0.05).Meanwhile,5 mg/d ATRA feeding increased the sucrase activity and the expres-sions of nutrient transporter related genes(GLUT2 and SLC7A1)in jejunal mucosa of TGEV-challenged piglets(P<0.05).Furthermore,5 mg/d ATRA feeding attenuated TGEV-induced intestinal inflammatory response by inhibit-ing the release of interleukin(IL)-1β,IL-8 and tumor necrosis factor-α(TNF-α),and promoting the secretion of IL-10 and secretory immunoglobulin A(sIgA)(P<0.05).Feeding 5 mg/d ATRA also down-regulated the expressions of Toll-like receptors and RIG-I like receptors signaling pathway related genes(TLR3,TLR4,RIG-I,MyD88,TRIF and MAVS)and the phosphorylation level of nuclear factor-κB-p65(NF-κB p65),and up-regulated the inhibitor kappa B alpha(IκBα)protein level in jejunal mucosa of TGEV-challenged piglets(P<0.05).Conclusions ATRA alleviated TGEV-induced intestinal barrier damage by inhibiting inflammatory response,thus improving the growth performance and inhibiting diarrhea of piglets.The mechanism was associated with the inhibi-tion of NF-κB signaling pathway mediated by TLR3,TLR4 and RIG-I.

LONG-TERM IMMUNOGENICITY AND EFFICACY OF RECOMBINANT YEAST DERIVED HEPATITIS B VACCINE FOR INTERRUPTION OF MOTHER-INFANT TRANSMISSION OF HEPATITIS B VIRUS

Recombinant DNA Yeast-Derived Hepatitis B Vaccine (RYHB vaccine) is comparable to and can replace Plasma-Derived Hepatitis B Vaccine (PHB vaccine) for the prevention of mother-nfant transmission of hepatitis B virus (HBV), but the duration of immune efficacy of RYHB vaccine is not clear. This study indicates the long-term efficacy for the prevention of mother-infant transmission of HBV. One hundred and six neonates born to HBsAg-arrier mothers with HBeAg positive were randomly divided into two groups, one receiving 20 μg per dose of RYHB vaccine and the another receiving 20 μg per dose of PHB vaccine on the day of birth, at 1 month and at 6 months (three times). Physical examination and blood tests were performed for all infants at 6, 12, 24, 36, 48 and 60 months of age. The results showed that the protective efficacies at 6, 12, 24, 36, 48 and 60 months were 67%, 75%, 63%, 62%, 57% and 56%, respectively for the RYHB vaccine group and 58%, 76%, 51%, 41%, 24% and 18%, respectively for the PHB vaccine group. The protective efficacy was notably significant in the last two years. The study indicates that the duration of protective efficacy is over 5 years with RYHB vaccine, being longer than that of PHB vaccine. These recipients of RYHB vaccine showed no side effects, and the vaccine is regarded as safe and effective.

A global invasion by the thrip,Frankliniella occidentalis:Current virus vector status and its management

Western flower thrip,Frankliniella occidentalis(Pergande),is among the most economically important agricultural pests globally,attacking a wide range of vegetable and horticultural crops.In addition to causing extensive crop damage,the species is notorious for vectoring destructive plant viruses,mainly belonging to the genera Orthotospovirus,Ilarvirus.AIphacarmovirus and Machlomovirus.Once infected by orthotospoviruses,thrips can remain virulent throughout their lifespan and continue transmitting viruses to host plants when and wherever they feed.These irruptive viral outbreaks in crops will permanently disrupt functional integrated pest management systems,and typically require a remedial treatment involving insecticides,contributing to further development of insecticide resistance.To mitigate against this continuing cycle,the most effective management is early and comprehensive surveillance of the pest species and recognition of plant viruses in the field.This review provides information on the pest status of F.occidentalis,discusses the current global status of the viruses vectored by this thrip species,examines the mechanisms involved in transmitting virus-induced diseases by thrips,and reviews different management strategies,highlighting the potential management tactics developed for various cropping systems.The early surveillance and the utilization of potential methods for control of both F occidentalis and viruses are proposed.

Epidemiologic study on human immunodeficiency virus infection among children in a former paid plasma donating community in China

Background Illegal plasma collecting activities in mid 1990s caused a large number of human immunodeficiency virus (HIV) infections in rural areas of central-eastern China. Although most of these activities have been stopped, there were few reports on secondary transmission from infected former plasma donors to their spouses and from infected mothers to their children. This study was to determine the extent of HIV infections among young children in a rural community with a large proportion of plasma donors.Methods A survey was conducted among children aged under 7 years in a former plasma donating community in September 2000: finger blood was collected for HIV antibody testing. Another survey was repeated among children aged under 8 years and their families in the same community in April 2001: urine samples were collected for HIV testing. HIV positive children and samples of HIV negative children, whose mothers were positive based on 2001 survey, were followed up until September 2002 to investigate HIV seroconversion, disease progression and HIV strain analysis. Questionnaires were administered to collect information on children’s delivery, breast feeding, medical history and their parents’ commercial blood donation history and HIV status.Results Among 169 children surveyed in 2000, 10 (5.9%) were HIV positive. Of 224 children, 11 were positive in 2001. The overall prevalence rate in the two surveys was 5.0% (17/337) when counting 56 repeated children only once. Of children born to HIV positive mothers, 28.9% were infected. A seroconversion rate of 2.5 per 100 child-years was observed by following up 28 HIV negative children. No statistically significant associations were found between children’s HIV infection and their histories of blood transfusion, surgery, immunization injection or medical injections. All infections were HIV-1 subtype B’ strain, the average dispersion rate is 7.4%. DNA sequence analysis showed a close relationship between the seroconverted children and their infected mothers.Conclusions HIV vertical transmissions in the rural former plasma donating community was significant. Intervention measures should be taken to prevent further transmission. It was estimated that the HIV spread in this community occurred in 1994 or even earlier. Many infected people are developing AIDS now: treatment and care are urgently needed for these sick people.

宿主细胞蛋白HSPA2与TGEV Nsp2的相互作用及其对病毒复制的影响

为了探究猪传染性胃肠炎病毒(Transmissible gastroenteritis virus,TGEV)非结构蛋白2(Nsp2)与宿主细胞热休克蛋白70-2(HSPA2)的相互作用,以及HSPA2表达对TGEV复制的影响,试验利用RT-PCR方法获得猪源HSPA2基因并构建其真核表达载体pCMV-Myc-HSPA2,通过Western-blot和间接免疫荧光试验检测真核表达载体pCMV-Myc-HSPA2在IPEC-J2细胞中的表达情况。利用免疫共沉淀(Co-IP)和共定位试验进一步验证TGEV Nsp2和宿主细胞蛋白HSPA2之间是否存在相互作用,利用基因过表达技术在IPEC-J2细胞中过表达HSPA2后接种TGEV,分别测定接种后36,48小时时病毒TCID50。结果表明:真核表达载体pCMV-Myc-HSPA2能够在IPEC-J2细胞中成功表达,HSPA2与Nsp2之间存在相互作用,并且HSPA2表达量升高后TGEV的TCID50略有升高。说明宿主细胞蛋白HSPA2与TGEV Nsp2之间存在相互作用,且HSPA2的表达对TGEV复制具有一定促进作用。