In Vitro Infectious Risk Assessment of Heliothis virescens ascovirus 3j(HvAV-3j) toward Non-target Vertebrate Cells
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摘要
As specific pathogens of noctuid pests, including Spodoptera exigua, S. litura, Helicoverpa armigera, and Mythimna separata, ascoviruses are suitable for the development of bioinsecticides. In this study, the infectivity of Heliothis virescens ascovirus 3j(HvAV-3j) on insect and mammalian cells was evaluated. HvAV-3j infection induced drastic morphological changes in Sf9, HzAM1, SeFB, and HaFB cells, including swelling and detachment. Notably, the latter phenomena did not occur in HvAV-3j-inoculated mammalian cells(HEK293, 7402, HePG2, PK15, ST, and TM3). MTT assays indicated that HvAV-3j inhibited the growth of host insect cells from the 6 th hpi, but no effects were detected in the HvAV-3j-inoculated mammalian cells. Furthermore, viral DNA replication, gene transcription, and protein expression were investigated, and the results consistently suggested that HvAV-3j viruses were not able to replicate their genomic DNA, transcribe, or express their proteins in the non-target vertebrate cells. The HvAV-3j genes were only transcribed and expressed in the four insect cell lines. These results indicated that HvAV-3j was infectious to cells derived from S. frugiperda, S. exigua, H.armigera, and H. zea but not to cells derived from human, pig, and mouse, suggesting that ascoviruses are safe to nontarget vertebrate cells.
As specific pathogens of noctuid pests, including Spodoptera exigua, S. litura, Helicoverpa armigera, and Mythimna separata, ascoviruses are suitable for the development of bioinsecticides. In this study, the infectivity of Heliothis virescens ascovirus 3j(HvAV-3j) on insect and mammalian cells was evaluated. HvAV-3j infection induced drastic morphological changes in Sf9, HzAM1, SeFB, and HaFB cells, including swelling and detachment. Notably, the latter phenomena did not occur in HvAV-3j-inoculated mammalian cells(HEK293, 7402, HePG2, PK15, ST, and TM3). MTT assays indicated that HvAV-3j inhibited the growth of host insect cells from the 6 th hpi, but no effects were detected in the HvAV-3j-inoculated mammalian cells. Furthermore, viral DNA replication, gene transcription, and protein expression were investigated, and the results consistently suggested that HvAV-3j viruses were not able to replicate their genomic DNA, transcribe, or express their proteins in the non-target vertebrate cells. The HvAV-3j genes were only transcribed and expressed in the four insect cell lines. These results indicated that HvAV-3j was infectious to cells derived from S. frugiperda, S. exigua, H.armigera, and H. zea but not to cells derived from human, pig, and mouse, suggesting that ascoviruses are safe to nontarget vertebrate cells.
As specific pathogens of noctuid pests, including Spodoptera exigua, S. litura, Helicoverpa armigera, and Mythimna separata, ascoviruses are suitable for the development of bioinsecticides. In this study, the infectivity of Heliothis virescens ascovirus 3j(HvAV-3j) on insect and mammalian cells was evaluated. HvAV-3j infection induced drastic morphological changes in Sf9, HzAM1, SeFB, and HaFB cells, including swelling and detachment. Notably, the latter phenomena did not occur in HvAV-3j-inoculated mammalian cells(HEK293, 7402, HePG2, PK15, ST, and TM3). MTT assays indicated that HvAV-3j inhibited the growth of host insect cells from the 6 th hpi, but no effects were detected in the HvAV-3j-inoculated mammalian cells. Furthermore, viral DNA replication, gene transcription, and protein expression were investigated, and the results consistently suggested that HvAV-3j viruses were not able to replicate their genomic DNA, transcribe, or express their proteins in the non-target vertebrate cells. The HvAV-3j genes were only transcribed and expressed in the four insect cell lines. These results indicated that HvAV-3j was infectious to cells derived from S. frugiperda, S. exigua, H.armigera, and H. zea but not to cells derived from human, pig, and mouse, suggesting that ascoviruses are safe to nontarget vertebrate cells.
引文
Arai E,Ishii K,Ishii H,Sagawa S,Makiyama N,Mizutani T,Omatsu T,Katayama Y,Kunimi Y,Inoue MN,Nakai M(2018)An ascovirus isolated from Spodoptera litura(Noctuidae:Lepidoptera)transmitted by the generalist endoparasitoid Meteorus pulchrcornis(Braconidae:Hymenoptera).J Gen Virol99:574-584
Arif BM(2005)A brief journey with insect viruses with emphasis on baculoviruses.J Invertebr Pathol 89:39-45
Asgari S(2006)Replication of Heliothis virescens ascovirus in insect cell lines.Arch Virol 151:1689-1699
Asgari S,Davis J,Wood D,Wilson P,McGrath A(2007)Sequence and organization of the Heliothis virescens ascovirus genome.J Gen Virol 88:1120-1132
Asgari S,Bideshi DK,Bigot Y,Federici BA,Cheng X(2017)ICTVreport consortium,ICTV virus taxonomy profile:Ascoviridae.J Gen Virol 98:4-5
Bideshi DK,Tan KP,Bigot Y,Federici BA(2005)A viral caspase contributes to modified apoptosis for virus transmission.Gene Dev 19:1416-1421
Bigot Y,Rabouille A,Sizaret PY,Hamelin MH,Periquet G(1997)Particle and genomic characteristics of a new member of the Ascoviridae:Diadromus pulchellus ascovirus.J Gen Virol78:1139-1147
Carbonell LF,Klowden MJ,Miller LK(1985)Baculovirus mediated expression of bacterial genes in dipteran and mammalian cells.J Virol 56:153-160
Carner GR,Hudson JS(1983)Histopathology of virus-like particles in Heliothis spp.J Invertebr Pathol 41:238-249
Chen ZS,Hou DH,Cheng XW,Wang X,Huang GH(2018)Genomic analysis of a novel isolate Heliothis virescens ascovirus 3i(HvAV-3i)and identification of ascoviral repeat ORFs(aros).Arch Virol 163:2849-2853
Chen ZS,Cheng XW,Wang X,Hou DH,Huang GH(2019)Proteomic analysis of the Heliothis virescens ascovirus 3i(HvAV-3i)virion.J Gen Virol 100:301-307
Cheng XW,Carner GR,Brown TM(1999)Circular configuration of the genome of ascoviruses.J Gen Virol 80:1537-1540
Cheng XW,Carner GR,Arif BM(2000)A new ascovirus from Spodoptera exigua and its relatedness to the isolate from Spodoptera frugiperda.J Gen Virol 81:3083-3092
Cheng XW,Wang LH,Carner GR,Arif BM(2005)Characterization of three ascovirus isolates from cotton insects.J Invertebr Pathol89:193-202
Federici BA(1982)A new type of insect pathogen in larvae of the clover cutworm,Scotogramma trifolii.J Invertebr Pathol40:41-54
Federici BA(1983)Enveloped double-stranded DNA insect virus with novel structure and cytopathology.Proc Natl Acad Sci USA80:7664-7668
Federici BA,Govindarajan R(1990)Comparative histopathology larval of three ascovirus noctuids isolates in larval noctuids.J Invertebr Pathol 56:300-311
Federici BA,Vlak JM,Hamm JJ(1990)Comparative study of virion structure,protein composition and genomic DNA of three ascovirus isolates.J Gen Virol 71:1661-1668
Federici BA,Bideshi DK,Tan Y,Spears T,Bigot Y(2009)Ascoviruses:superb manipulators of apoptosis for viral replication and transmission.Curr Top Microbiol 328:171-196
Govindarajan R,Federici BA(1990)Ascovirus infectivity and effects of infection on the growth and development of noctuid larvae.J Invertebr Pathol 56:291-299
Groener A,Granados RR,Burand JP(1984)Interaction of Autographa californica nuclear polyhedrosis virus with two nonpermissive cell lines.Intervirology 21:203-209
Hamm JJ,Pair SD,Marti OG(1986)Incidence and host range of a new ascovirus isolated from fall armyworm,Spodoptera frugiperda(Lepidoptera:Noctuidae).Fla Entomol 69:524-531
Huang GH,Garretson TA,Cheng XH,Holztrager MS,Li SJ,Wang X,Cheng XW(2012a)Phylogenetic position and replication kinetics of Heliothis virescens ascovirus 3 h(HvAV-3h)isolated from Spodoptera exigua.PLoS ONE 7:e40225
Huang GH,Wang YS,Wang X,Garretson TA,Dai LY,Zhang CX,Cheng XW(2012b)Genomic sequence of Heliothis virescens ascovirus 3 g isolated from Spodoptera exigua.J Virol86:12467-12468
Huang GH,Hou DH,Wang M,Chen XW,Hu Z(2017)Genome analysis of Helithis virescens ascovirus 3h isolated from China.Virol Sin 32:147-154
Kim E,Jeon IS,Kim JW,Kim J,Jung HS,Lee SJ(2007)An MTT-based method for quantification of periodontal ligament cell viability.Oral Dis 13:495-499
Li SJ,Wang X,Zhou ZS,Zhu J,Hu J,Zhao YP,Zhou GW,Huang GH(2013)A comparison of growth and development of three major agricultural insect pests infected with Heliothis virescens ascovirus 3 h(HvAV-3h).PLoS ONE 8:e85704
Li SJ,Hopkins RJ,Zhao YP,Zhang YX,Hu J,Chen XY,Xu Z,Huang GH(2016)Imperfection works:survival,transmission and persistence in the system of Helithis virescens ascovirus 3h(HvAV-3h),Microplitis similis and Spodoptera exigua.Sci Rep16:21296
Liu YY,Xian WF,Xue J,Wei YL,Cheng XW,Wang X(2018)Complete Genome Sequence of a Renamed Isolate,Trichoplusia ni ascovirus 6b,from the United States.Genome Announc6:e00148-18
Tao XY,Choi JY,Kim WJ,Lee JH,Liu Q,Kim SE,An SB,Lee SH,Woo SD,Jin BR,Je YH(2013)The Autographa californica multiple nucleopolyhedrovirus ORF78 is essential for budded virus production and general occlusion body formation.J Virol87:8441-8450
Thiem SM(1997)Prospects for altering host range for baculovirus bioinsecticides.Curr Opin Biotechnol 8:317-322
Tjia ST,Altenschildesche GM,Doerfler W(1983)Autographa californica nuclear polyhedrosis virus(AcNPV)DNA does not persist in mass cultures of mammalian cells.Virology125:107-117
Volkman LE,Goldsmith PA(1983)In vitro survey of Autographa californica nuclear polyhedrosis virus interaction with nontarget vertebrate host cells.Appl Environ Microbiol 45:1085-1093
Wei YL,Hu J,Li SJ,Chen ZS,Cheng XW,Huang GH(2014)Genome sequence and organization analysis of Helithis virecens ascovirus 3f isolated form a Helicoverpa zea larva.J Invertebr Pathol 122:40-43
Wu WB,Lin TH,Pan LJ,Yu M,Li ZF,Pang Y,Yang K(2006)Autographa californica multiple nucleopolyhedrovirus nucleocapsid assembly is interrupted upon deletion of the 38 K gene.J Virol 80:11475-11485
Yu H,Xu J,Liu Q,Liu TX,Wang D(2015)Ha83,a chitin binding domain encoding gene,is important to Helicoverpa armigera nucleopolyhedrovirus budded virus production and occlusion body assembling.Sci Rep 5:11088
Yu H,He L,Ou-Yang YY,Huang GH(2018a)Preparation and molecular characterization of a polyclonal antibody as an efficient cutworm reference protein.J Asia Pac Entomol21:786-792
Yu H,Li ZQ,He L,Ou-Yang YY,Ni L,Huang GH(2018b)Response analysis of host Spodoptera exigua larvae to infection by Heliothis virescens ascovirus 3 h(HvAV-3h)via transcriptome.Sci Rep 8:5367
Zhang H,Zhang YA,Qin QL,Li X,Miao L,Wang YZ,Qu LJ,Zhang AJ,Yang Q(2009)A cell strain cloned from Spodoptera exigua cell line(IOZCAS-Spex-II)highly susceptible to S.exigua nucleopolyhedrovirus infection.In Vitro Cell Dev Biol Anim 45:201-204
Zhu SM,Wang W,Wang Y,Yuan MJ,Yang K(2013)The baculovirus core gene ac83 is required for nucleocapsid assembly and per os infectivity of Autographa californica nucleopolyhedrovirus.J Virol 87:10573-10586
Arif BM(2005)A brief journey with insect viruses with emphasis on baculoviruses.J Invertebr Pathol 89:39-45
Asgari S(2006)Replication of Heliothis virescens ascovirus in insect cell lines.Arch Virol 151:1689-1699
Asgari S,Davis J,Wood D,Wilson P,McGrath A(2007)Sequence and organization of the Heliothis virescens ascovirus genome.J Gen Virol 88:1120-1132
Asgari S,Bideshi DK,Bigot Y,Federici BA,Cheng X(2017)ICTVreport consortium,ICTV virus taxonomy profile:Ascoviridae.J Gen Virol 98:4-5
Bideshi DK,Tan KP,Bigot Y,Federici BA(2005)A viral caspase contributes to modified apoptosis for virus transmission.Gene Dev 19:1416-1421
Bigot Y,Rabouille A,Sizaret PY,Hamelin MH,Periquet G(1997)Particle and genomic characteristics of a new member of the Ascoviridae:Diadromus pulchellus ascovirus.J Gen Virol78:1139-1147
Carbonell LF,Klowden MJ,Miller LK(1985)Baculovirus mediated expression of bacterial genes in dipteran and mammalian cells.J Virol 56:153-160
Carner GR,Hudson JS(1983)Histopathology of virus-like particles in Heliothis spp.J Invertebr Pathol 41:238-249
Chen ZS,Hou DH,Cheng XW,Wang X,Huang GH(2018)Genomic analysis of a novel isolate Heliothis virescens ascovirus 3i(HvAV-3i)and identification of ascoviral repeat ORFs(aros).Arch Virol 163:2849-2853
Chen ZS,Cheng XW,Wang X,Hou DH,Huang GH(2019)Proteomic analysis of the Heliothis virescens ascovirus 3i(HvAV-3i)virion.J Gen Virol 100:301-307
Cheng XW,Carner GR,Brown TM(1999)Circular configuration of the genome of ascoviruses.J Gen Virol 80:1537-1540
Cheng XW,Carner GR,Arif BM(2000)A new ascovirus from Spodoptera exigua and its relatedness to the isolate from Spodoptera frugiperda.J Gen Virol 81:3083-3092
Cheng XW,Wang LH,Carner GR,Arif BM(2005)Characterization of three ascovirus isolates from cotton insects.J Invertebr Pathol89:193-202
Federici BA(1982)A new type of insect pathogen in larvae of the clover cutworm,Scotogramma trifolii.J Invertebr Pathol40:41-54
Federici BA(1983)Enveloped double-stranded DNA insect virus with novel structure and cytopathology.Proc Natl Acad Sci USA80:7664-7668
Federici BA,Govindarajan R(1990)Comparative histopathology larval of three ascovirus noctuids isolates in larval noctuids.J Invertebr Pathol 56:300-311
Federici BA,Vlak JM,Hamm JJ(1990)Comparative study of virion structure,protein composition and genomic DNA of three ascovirus isolates.J Gen Virol 71:1661-1668
Federici BA,Bideshi DK,Tan Y,Spears T,Bigot Y(2009)Ascoviruses:superb manipulators of apoptosis for viral replication and transmission.Curr Top Microbiol 328:171-196
Govindarajan R,Federici BA(1990)Ascovirus infectivity and effects of infection on the growth and development of noctuid larvae.J Invertebr Pathol 56:291-299
Groener A,Granados RR,Burand JP(1984)Interaction of Autographa californica nuclear polyhedrosis virus with two nonpermissive cell lines.Intervirology 21:203-209
Hamm JJ,Pair SD,Marti OG(1986)Incidence and host range of a new ascovirus isolated from fall armyworm,Spodoptera frugiperda(Lepidoptera:Noctuidae).Fla Entomol 69:524-531
Huang GH,Garretson TA,Cheng XH,Holztrager MS,Li SJ,Wang X,Cheng XW(2012a)Phylogenetic position and replication kinetics of Heliothis virescens ascovirus 3 h(HvAV-3h)isolated from Spodoptera exigua.PLoS ONE 7:e40225
Huang GH,Wang YS,Wang X,Garretson TA,Dai LY,Zhang CX,Cheng XW(2012b)Genomic sequence of Heliothis virescens ascovirus 3 g isolated from Spodoptera exigua.J Virol86:12467-12468
Huang GH,Hou DH,Wang M,Chen XW,Hu Z(2017)Genome analysis of Helithis virescens ascovirus 3h isolated from China.Virol Sin 32:147-154
Kim E,Jeon IS,Kim JW,Kim J,Jung HS,Lee SJ(2007)An MTT-based method for quantification of periodontal ligament cell viability.Oral Dis 13:495-499
Li SJ,Wang X,Zhou ZS,Zhu J,Hu J,Zhao YP,Zhou GW,Huang GH(2013)A comparison of growth and development of three major agricultural insect pests infected with Heliothis virescens ascovirus 3 h(HvAV-3h).PLoS ONE 8:e85704
Li SJ,Hopkins RJ,Zhao YP,Zhang YX,Hu J,Chen XY,Xu Z,Huang GH(2016)Imperfection works:survival,transmission and persistence in the system of Helithis virescens ascovirus 3h(HvAV-3h),Microplitis similis and Spodoptera exigua.Sci Rep16:21296
Liu YY,Xian WF,Xue J,Wei YL,Cheng XW,Wang X(2018)Complete Genome Sequence of a Renamed Isolate,Trichoplusia ni ascovirus 6b,from the United States.Genome Announc6:e00148-18
Tao XY,Choi JY,Kim WJ,Lee JH,Liu Q,Kim SE,An SB,Lee SH,Woo SD,Jin BR,Je YH(2013)The Autographa californica multiple nucleopolyhedrovirus ORF78 is essential for budded virus production and general occlusion body formation.J Virol87:8441-8450
Thiem SM(1997)Prospects for altering host range for baculovirus bioinsecticides.Curr Opin Biotechnol 8:317-322
Tjia ST,Altenschildesche GM,Doerfler W(1983)Autographa californica nuclear polyhedrosis virus(AcNPV)DNA does not persist in mass cultures of mammalian cells.Virology125:107-117
Volkman LE,Goldsmith PA(1983)In vitro survey of Autographa californica nuclear polyhedrosis virus interaction with nontarget vertebrate host cells.Appl Environ Microbiol 45:1085-1093
Wei YL,Hu J,Li SJ,Chen ZS,Cheng XW,Huang GH(2014)Genome sequence and organization analysis of Helithis virecens ascovirus 3f isolated form a Helicoverpa zea larva.J Invertebr Pathol 122:40-43
Wu WB,Lin TH,Pan LJ,Yu M,Li ZF,Pang Y,Yang K(2006)Autographa californica multiple nucleopolyhedrovirus nucleocapsid assembly is interrupted upon deletion of the 38 K gene.J Virol 80:11475-11485
Yu H,Xu J,Liu Q,Liu TX,Wang D(2015)Ha83,a chitin binding domain encoding gene,is important to Helicoverpa armigera nucleopolyhedrovirus budded virus production and occlusion body assembling.Sci Rep 5:11088
Yu H,He L,Ou-Yang YY,Huang GH(2018a)Preparation and molecular characterization of a polyclonal antibody as an efficient cutworm reference protein.J Asia Pac Entomol21:786-792
Yu H,Li ZQ,He L,Ou-Yang YY,Ni L,Huang GH(2018b)Response analysis of host Spodoptera exigua larvae to infection by Heliothis virescens ascovirus 3 h(HvAV-3h)via transcriptome.Sci Rep 8:5367
Zhang H,Zhang YA,Qin QL,Li X,Miao L,Wang YZ,Qu LJ,Zhang AJ,Yang Q(2009)A cell strain cloned from Spodoptera exigua cell line(IOZCAS-Spex-II)highly susceptible to S.exigua nucleopolyhedrovirus infection.In Vitro Cell Dev Biol Anim 45:201-204
Zhu SM,Wang W,Wang Y,Yuan MJ,Yang K(2013)The baculovirus core gene ac83 is required for nucleocapsid assembly and per os infectivity of Autographa californica nucleopolyhedrovirus.J Virol 87:10573-10586