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Guohua Huang - One of the best experts on this subject based on the ideXlab platform.
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novel strategies for the biocontrol of noctuid pests lepidoptera based on improving Ascovirus infectivity using bacillus thuringiensis
Insect Science, 2021Co-Authors: Changjin Yang, Ying Zhao, Zhuangmei Chen, Michael J Adang, Guohua HuangAbstract:Identifying novel biocontrol agents and developing new strategies are urgent goals in insect pest biocontrol. Ascoviruses are potential competent insect viruses that may be developed into bioinsecticides, but this aim is impeded by their poor oral infectivity. To improve the per os infectivity of Ascovirus, Bacillus thuringiensis kurstaki (Btk) was employed as a helper to damage the midgut of lepidopteran larvae (Helicoverpa armigera, Mythimna separata, Spodoptera frugiperda, and S. litura) in formulations with Heliothis virescens Ascovirus isolates (HvAV-3h and HvAV-3j). Btk and Ascovirus mixtures (Btk/HvAV-3h and Btk/HvAV-3j) were fed to insect larvae (3rd instar). With the exception of S. frugiperda larvae, which exhibited low mortality after ingesting Btk, the larvae of the other tested species showed three types of response to feeding on the formulas: type I, the tested larvae (H. armigera) were killed by Btk infection so quickly that insufficient time and resources remained for ascoviral invasion; type II, both Btk and the Ascovirus were depleted by their competition, such that neither was successfully released or colonized the tissue; type III, Btk was eliminated by the Ascovirus, and the Ascovirus achieved systemic infection in the tested larvae. The feeding of Btk/Ascovirus formulas led to a great reduction in larval diet consumption and resulted in a significant decrease in the emergence rate of H. armigera, M. separata, and S. litura larvae, which suggested that the formulas exerted marked oral control effects on both the contemporary individuals and the next generation of these tested pest species.
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3h 31 a non structural protein of heliothis virescens Ascovirus 3h inhibits the host larval cathepsin and chitinase activities
Virologica Sinica, 2021Co-Authors: Yiyi Ouyang, Changjin Yang, Madoka Nakai, Guohua HuangAbstract:3h-31 of Heliothis virescens Ascovirus 3h (HvAV-3h) is a highly conserved gene of Ascoviruses. As an early gene of HvAV-3h, 3h-31 codes for a non-structural protein (3H-31) of HvAV-3h. In the study, 3h-31 was initially transcribed and expressed at 3 h post-infection (hpi) in the infected Spodoptera exigua fat body cells (SeFB). 3h-31 was further inserted into the bacmid of Autographa californica nucleopolyhedrovirus (AcMNPV) to generate an infectious baculovirus (AcMNPV-31). In vivo experiments showed that budded virus production and viral DNA replication decreased with the expression of 3H-31, and lucent tubular structures were found around the virogenic stroma in the AcMNPV-31-infected SeFB cells. In vivo, both LD50 and LD90 values of AcMNPV-31 were significantly higher than those of the wild-type AcMNPV (AcMNPV-wt) in third instar S. exigua larvae. An interesting finding was that the liquefaction of the larvae killed by the infection of AcMNPV-31 was delayed. Chitinase and cathepsin activities of AcMNPV-31-infected larvae were significantly lower than those of AcMNPV-wt-infected larvae. The possible regulatory function of the chitinase and cathepsin for 3H-31 was further confirmed by RNAi, which showed that larval cathepsin activity was significantly upregulated, but chitinase activity was not significantly changed due to the RNAi of 3h-31. Based on the obtained results, we assumed that the function of 3H-31 was associated with the inhibition of host larval chitinase and cathepsin activities, so as to restrain the hosts in their larval stages.
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melanization induced by heliothis virescens Ascovirus 3h promotes viral replication
Insect Science, 2021Co-Authors: Xiaohui Song, Min Wang, Shu Wang, Guohua HuangAbstract:Melanization is an important innate immune defense mechanism of insects, which can kill invading pathogens. Most pathogens, for their survival and reproduction, inhibit the melanization of the host. Interestingly, our results suggested that after infection with Heliothis virescens Ascovirus 3h (HvAV-3h), the speed of melanization in infected Spodoptera exigua larval hemolymph was accelerated and that the phenoloxidase (PO) activity of hemolymph in larvae infected with HvAV-3h increased significantly (1.20-fold at 96 hpi, 1.52-fold at 120 hpi, 1.23-fold at 144 hpi, 1.12-fold at 168 hpi). The transcription level of the gene encoding S. exigua prophenoloxidase-1 (SePPO-1 gene) was upregulated dramatically in the fat body during the middle stage of infection. In addition, when melanization was inhibited or promoted, the replication of HvAV-3h was inhibited or promoted, respectively. In conclusion, infection with HvAV-3h can markedly induce melanization in the middle stage of infection, and melanization is helpful for HvAV-3h viral replication.
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a spodoptera exigua serpin is involved in the immunity of hemolymph induced by heliothis virescens Ascovirus 3h
Journal of Asia-pacific Entomology, 2021Co-Authors: Ying Zhao, Guohua HuangAbstract:Abstract Serine protease inhibitors (serpins) are a broadly distributed protease superfamily in various organisms, which participate in the negative regulation of melanization in insects. Our pervious study demonstrated that induced melanization by Heliothis virescens Ascovirus 3h is helpful for HvAV-3h viral replication. In this study, the serpin-2.1 like gene from Spodoptera exigua (Seserpin-2.1 like) which is a specific serpin after HvAV-3h infection was identified. Seserpin-2.1 like has the typical characteristics of serpin proteins family. Seserpin-2.1 like widely distributed in various tissues (including hemolymph, fat body, midgut, cuticle and head), and expressed at a higher level in fat body, hemolymph and head. At different developmental stages, it tends to express at eggs and 5th instar larvae. Seserpin-2.1 like shows special phenomenon for HvAV-3h infection. On the one hand, the transcription level of Seserpin-2.1 like in hemolymph was significantly upregulated after HvAV-3h compared to other immune challenges Gram-positive bacteria (Bacillus thuringiensis), Gram-negative bacteria (Escherichia coli), and Baculovirus (Autographa californica nucleopolyhedrovirus). On the other hand, its expression disappeared during 72 to 120 h post infection. In summary, our results suggest that Seserpin-2.1 like may be involved in innate immunity of S. exigua after infection with HvAV-3h. And HvAV-3h may inhibit the expression of Seserpin-2.1 like to promote the melanization of host larvae for viral replication.
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characterization of heliothis virescens Ascovirus 3h orf21 that encodes a virion protein
Journal of Applied Virology, 2020Co-Authors: Ying Zhao, Guohua HuangAbstract:Homologues of Heliothis virescens Ascovirus 3h (HvAV-3h) orf21 are found in 9 completely sequenced members of the Ascoviruses, but so far their functions are unknown. Here, orf21 ( 3h-21 ) was cloned in-frame into a pET-28a bacterial expression vector. The fusion protein produced by this construct was used for the preparation of a polyclonal antiserum. RT-PCR analysis showed a single transcript of 3h-21 of approximately 0.7kb was transcribed beginning at 24h post-infection in infected Helicoverpa armigera larvae. Western blot analysis of extracts from HvAv-3h-infected Helicoverpa armigera larvae detected a 25.6 kDa protein late in infection. This antiserum also reacted with a 25.6 kDa protein in purified virions of HvAv-3h. The protein was not extensively modified post-translation. Immunoelectron microscopy confirmed that the 3H-21 is associated with the structure of HvAV-3h virions.
Xiaowen Cheng - One of the best experts on this subject based on the ideXlab platform.
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proteomic analysis of the heliothis virescens Ascovirus 3i hvav 3i virion
Journal of General Virology, 2019Co-Authors: Zishu Chen, Xiaowen Cheng, Xing Wang, Dianhai Hou, Guohua HuangAbstract:Ascoviruses are enveloped, circular, double-stranded DNA viruses that can effectively control the appetite of lepidopteran larvae, thereby reducing the consequent damage and economic losses to crops. In this study, the virion of a sequenced Heliothis virescens Ascovirus 3i (HvAV-3i) strain was used to perform proteomic analysis using both in-gel and in-solution digestion. A total of 81 viral proteins, of which 67 were associated with the virions, were identified in the proteome of HvAV-3i virions. Among these proteins, 23 with annotated functions were associated with DNA/RNA metabolism/transcription, virion assembly, sugar and lipid metabolism, signalling, cellular homoeostasis and cell lysis. Twenty-one viral membrane proteins were also identified. Some of the minor ‘virion’ proteins identified may be non-virion contaminants of viral proteins synthesized during replication, identified by more recent and highly sensitive methods. The extensive identification of the ascoviral proteome will establish a foundation for further investigation of ascoviral replication and infection.
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genomic analysis of a novel isolate heliothis virescens Ascovirus 3i hvav 3i and identification of ascoviral repeat orfs aros
Archives of Virology, 2018Co-Authors: Zishu Chen, Xiaowen Cheng, Xing Wang, Guohua HuangAbstract:Ascoviruses are circular double-stranded DNA viruses that infect insects. Herein we sequenced and analyzed the genome of the previously unrecorded Ascovirus isolate Heliothis virescens Ascovirus 3i (HvAV-3i). The genome size is 185,650 bp with 181 hypothetical open reading frames (ORFs). Additionally, definition based on Ascovirus repeated ORFs (aros) is proposed; whereby the 29 aros from all sequenced Ascoviridae genomes are divided into six distinct groups. The topological relationship among the isolates of Heliothis virescens Ascovirus 3a is (HvAV-3f, {HvAV-3h, [HvAV-3e, (HvAV-3g, HvAV-3i)]}) with every clade well supported by a Bayesian posterior probability of 1.00 and a Bootstrap value of 100%.
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complete genome sequence of a renamed isolate trichoplusia ni Ascovirus 6b from the united states
Genome Announcements, 2018Co-Authors: Yuanyuan Liu, Xiaowen Cheng, Wenfei Xian, Jin Xue, Yonglu Wei, Xing WangAbstract:ABSTRACT The complete genome of Trichoplusia ni Ascovirus 6b (TnAV-6b) was sequenced for the first time. The TnAV-6b isolate, which has its closest phylogenetic relationship with the TnAV-6a isolate, has a circular genome of 185,664 bp, with a G+C content of 46.0% and 178 predicted open reading frames.
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genome analysis of heliothis virescens Ascovirus 3h isolated from china
Virologica Sinica, 2017Co-Authors: Xiaowen Cheng, Guohua Huang, Manli Wang, Zhihong HuAbstract:No Ascovirus isolated from China has been sequenced so far. Therefore, in this study, we aimed to sequence the genome of Heliothis virescens Ascovirus 3h (HvAV-3h) using the 454 pyrosequencing technology. The genome was found to be 190,519-bp long with a G+C content of 45.5%. We also found that it encodes 185 hypothetical open reading frames (ORFs) along with at least 50 amino acids, including 181 ORFs found in other Ascoviruses and 4 unique ORFs. Gene-parity plots and phylogenetic analysis revealed a close relationship between HvAV-3h and three other HvAV-3a strains and a distant relationship with Spodoptera frugiperda Ascovirus 1a (SfAV-1a), Trichoplusia ni Ascovirus 6a (TnAV-6a), and Diadromus pulchellus Ascovirus 4a (DpAV-4a). Among the 185 potential genes encoded by the genome, 44 core genes were found in all the sequenced Ascoviruses. In addition, 25 genes were found to be conserved in all Ascoviruses except DpAV-4a. In the HvAV-3h genome, 24 baculovirus repeat ORFs (bros) were present, and the typical homologous repeat regions (hrs) were absent. This study supplies information important for understanding the conservation and functions of Ascovirus genes as well as the variety of ascoviral genomes.
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ICTV virus taxonomy profile: Ascoviridae
Journal of General Virology, 2017Co-Authors: Sassan Asgari, Yves Le Bigot, Dennis K Bideshi, Brian A. Federici, Xiaowen ChengAbstract:The family Ascoviridae includes viruses with circular dsDNA genomes of 100–200 kbp characterized by oblong enveloped virions of 200–400 nm in length. Ascoviruses mainly infect lepidopteran larvae and are mechanically transmitted by parasitoid wasps in which they may also replicate. Most known members belong to the genus Ascovirus, except one virus, that of the genus Toursvirus, which replicates in both its lepidopteran and parasitoid vector hosts. Ascoviruses cause high mortality among economically important insect pests, thereby controlling insect populations. This is a summary of the current International Committee on Taxonomy of Viruses (ICTV) Report on the taxonomy of the Ascoviridae, which is available at www.ictv.global/report/ascoviridae.
Sassan Asgari - One of the best experts on this subject based on the ideXlab platform.
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ICTV virus taxonomy profile: Ascoviridae
Journal of General Virology, 2017Co-Authors: Sassan Asgari, Yves Le Bigot, Dennis K Bideshi, Brian A. Federici, Xiaowen ChengAbstract:The family Ascoviridae includes viruses with circular dsDNA genomes of 100–200 kbp characterized by oblong enveloped virions of 200–400 nm in length. Ascoviruses mainly infect lepidopteran larvae and are mechanically transmitted by parasitoid wasps in which they may also replicate. Most known members belong to the genus Ascovirus, except one virus, that of the genus Toursvirus, which replicates in both its lepidopteran and parasitoid vector hosts. Ascoviruses cause high mortality among economically important insect pests, thereby controlling insect populations. This is a summary of the current International Committee on Taxonomy of Viruses (ICTV) Report on the taxonomy of the Ascoviridae, which is available at www.ictv.global/report/ascoviridae.
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effects of an Ascovirus hvav 3e on diamondback moth plutella xylostella and evidence for virus transmission by a larval parasitoid
Journal of Invertebrate Pathology, 2010Co-Authors: Michael J Furlong, Sassan AsgariAbstract:Ascoviruses (AVs) are pathogenic to lepidopteran larvae, and most commonly attack species in the Noctuidae. The unique pathology includes cleavage of host cells into virion-containing vesicles which leads to the milky white colouration of the hemolymph as opposed to the clear hemolymph of healthy larvae. Recently, we showed that a Heliothis virescens AV (HvAV-3e) isolate is able to induce disease in Crocidolomia pavonana F. (Lepidoptera: Crambidae), affecting feeding, growth and survival of infected larvae. In this study, we investigated the effect of different variants of HvAV-3e on diamondback moth, Plutella xylostella (Lepidoptera: Plutellidae) larvae, another non-noctuid host. In hemolymph inoculation bioassays fourth instar larvae showed a significant dose response to each of the HvAV-3e variants and significant differences between the virulence of the three variants were detected. Both second and fourth instars were readily infected with the virus and infected individuals demonstrated significant reductions in food consumption and growth. The majority of infected individuals died at the larval or pupal stage and individuals which developed into adults were usually deformed, less fecund than non-infected controls and died shortly after emergence. In transmission studies, Diadegma semiclausum (Hymenoptera: Ichneumonidae), a key parasitoid of diamondback moth, infected healthy host larvae during oviposition following previous attack of HvAV-3e infected hosts. In choice tests D. semiclausum did not discriminate between infected individuals but host infection had no detectable impact on the development of immature D. semiclausum or on subsequent adults.
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An Ascovirus-Encoded RNase III Autoregulates Its Expression and Suppresses RNA Interference-Mediated Gene Silencing
Journal of virology, 2010Co-Authors: Mazhar Hussain, Alexander M. Abraham, Sassan AsgariAbstract:RNase III proteins play vital roles in processing of several types of RNA molecules and gene silencing. Recently, it has been discovered that some plant and animal viruses encode RNase III-like proteins as well. Genome sequencing of four virus species belonging to the Ascoviridae family has revealed sequence conservation of an RNase III open reading frame among the viruses. These have not been explored in Ascoviruses, and therefore their role in host-virus interaction is unknown. Here, we confirmed expression of Heliothis virescens Ascovirus (HvAV-3e) open reading frame 27 (orf27) that encodes an RNase III-like protein after infection and demonstrated dsRNA specific endoribonuclease activity of the encoded protein. Analysis of the expression patterns of orf27 in virus-infected insect cells and a bacterial expression system revealed autoregulation of this protein over time. Moreover, HvAV-3e RNase III was found essential for virus DNA replication and infection using RNA interference (RNAi)-mediated gene silencing. In addition, using green fluorescent protein gene as a marker, we provide evidence that RNase III is involved in the suppression of gene silencing. To our knowledge, this is the first insect virus-encoded RNase III described and shown to suppress host cell RNAi defense mechanism.
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An Ascovirus-encoded RNase III autoregulates its expression and suppresses RNA interference-mediated gene silencing
2010Co-Authors: Mazhar Hussain, Er M. Abraham, Sassan AsgariAbstract:RNase III proteins play vital roles in processing of several types of RNA molecules and gene silencing. Recently, it has been discovered that some plant and animal viruses encode RNase III-like proteins as well. Genome sequencing of four virus species belonging to the Ascoviridae family has revealed sequence conserva-tion of an RNase III open reading frame among the viruses. These have not been explored in Ascoviruses, and therefore their role in host-virus interaction is unknown. Here, we confirmed expression of Heliothis virescens Ascovirus (HvAV-3e) open reading frame 27 (orf27) that encodes an RNase III-like protein after infection and demonstrated dsRNA specific endoribonuclease activity of the encoded protein. Analysis of the expression patterns of orf27 in virus-infected insect cells and a bacterial expression system revealed autoregulation of this protein over time. Moreover, HvAV-3e RNase III was found essential for virus DNA replication and infection using RNA interference (RNAi)-mediated gene silencing. In addition, using green fluorescent protein gene as a marker, we provide evidence that RNase III is involved in the suppression of gene silencing. To our knowledge, this is the first insect virus-encoded RNase III described and shown to suppress host cell RNAi defense mechanism. RNases III are double-stranded RNA (dsRNA)-specific processing enzymes present in prokaryotes, eukaryotes, an
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functional analysis of a cellular microrna in insect host Ascovirus interaction
Journal of Virology, 2010Co-Authors: Mazhar Hussain, Sassan AsgariAbstract:MicroRNAs (miRNAs) have emerged as key regulators in many biological processes, from development to defense, at almost all organismal levels. Recently, their role has been highlighted in pathogen-host interactions. Emerging evidence from a variety of virus-host systems indicates that cellular as well as virally encoded miRNAs influence viral replication. Here, we report changes in expression levels of host miRNAs upon Ascovirus infection in an insect cell line and investigated the role of a host miRNA, Hz-miR24, in the host-virus system. We found that Hz-miR24 is differentially expressed following virus infection, with an increase in its expression levels late in infection. Experimental evidence demonstrated that Hz-miR24 downregulates ascoviral DNA-dependent RNA polymerase and its β subunit transcript levels late in infection. The specific miRNA-target interactions were investigated and confirmed using the ectopic expression of Hz-miR24 and a green fluorescent protein-based reporter system. Further, the expression of the target gene was substantially enhanced in cells transfected with a synthesized inhibitor of Hz-miR24. These findings suggest that Ascoviruses manipulate host miRNAs that in turn regulate the expression of their genes at specific time points after infection. To our knowledge, this is the first cellular miRNA reported to interact with an insect virus.
Yves Le Bigot - One of the best experts on this subject based on the ideXlab platform.
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Ascovirus P64 Homologs: A Novel Family of Large Cationic Proteins That Condense Viral Genomic DNA for Encapsidation
Biology, 2018Co-Authors: Dennis K Bideshi, Yves Le Bigot, Yeping Tan, Tatsinda Spears, Heba Zaghloul, Brian FedericiAbstract:Eukaryotic dsDNA viruses use small basic protamine-like proteins or histones, typically
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Ascovirus P64 Homologs: A Novel Family of Large Cationic Proteins That Condense Viral Genomic DNA for Encapsidation
MDPI AG, 2018Co-Authors: Dennis K Bideshi, Yves Le Bigot, Yeping Tan, Tatsinda Spears, Heba A. H. Zaghloul, Brian A. FedericiAbstract:Eukaryotic dsDNA viruses use small basic protamine-like proteins or histones, typically <15 kDa, to condense and encapsidate their genomic (g)DNAs during virogenesis. Ascoviruses are large dsDNA (~100–200 kbp) viruses that are pathogenic to lepidopteran larvae. Little is known about the molecular basis for condensation and encapsidation of their gDNAs. Previous proteomic analysis showed that Spodoptera frugiperda Ascovirus (SfAV-1a) virions contain a large unique DNA-binding protein (P64; 64 kDa, pI = 12.2) with a novel architecture proposed to condense its gDNA. Here we used physical, biochemical, and transmission electron microscopy techniques to demonstrate that P64’s basic C-terminal domain condenses SfAV-1a gDNA. Moreover, we demonstrate that only P64 homologs in other Ascovirus virions are unique in stably binding DNA. As similar protein families or subfamilies were not identified in extensive database searches, our collective data suggest that Ascovirus P64 homologs comprise a novel family of atypical large viral gDNA condensing proteins
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ICTV virus taxonomy profile: Ascoviridae
Journal of General Virology, 2017Co-Authors: Sassan Asgari, Yves Le Bigot, Dennis K Bideshi, Brian A. Federici, Xiaowen ChengAbstract:The family Ascoviridae includes viruses with circular dsDNA genomes of 100–200 kbp characterized by oblong enveloped virions of 200–400 nm in length. Ascoviruses mainly infect lepidopteran larvae and are mechanically transmitted by parasitoid wasps in which they may also replicate. Most known members belong to the genus Ascovirus, except one virus, that of the genus Toursvirus, which replicates in both its lepidopteran and parasitoid vector hosts. Ascoviruses cause high mortality among economically important insect pests, thereby controlling insect populations. This is a summary of the current International Committee on Taxonomy of Viruses (ICTV) Report on the taxonomy of the Ascoviridae, which is available at www.ictv.global/report/ascoviridae.
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p64 a novel major virion dna binding protein potentially involved in condensing the spodoptera frugiperda Ascovirus 1a genome
Journal of Virology, 2009Co-Authors: Yeping Tan, Yves Le Bigot, Dennis K Bideshi, Jeffrey J. Johnson, Robert H. Hice, Tatsinda Spears, Brian A. FedericiAbstract:We recently identified 21 structural proteins in the virion of Spodoptera frugiperda Ascovirus 1a (SfAV1a), a virus with a large, double-stranded DNA genome of 157 kbp, which attacks species of the lepidopteran family Noctuidae. The two most abundant virion proteins were the major capsid protein and a novel protein (P64) of 64 kDa that contained two distinct domains not known previously to occur together. The amino-terminal half of P64 (residues 1 to 263) contained four repeats (a recently recognized motif with an unknown function) of a virus-specific two-cysteine adaptor. Adjoined to this, the carboxy-terminal half of P64 (residues 279 to 455) contained 14 copies of a highly basic, tandemly repeated motif rich in arginine and serine, having an 11- to 13-amino-acid consensus sequence, SPSQRRSTS(V/K)(A/S)RR, yielding a predicted isoelectric point of 12.2 for this protein. In the present study, we demonstrate by Southwestern analysis that SfAV1a P64 was the only virion structural protein that bound DNA. Additional electrophoretic mobility shift assays showed that P64 bound SfAV1a as well as non-SfAV1a DNA. Furthermore, we show through immunogold labeling of ultrathin sections that P64 is a component of virogenic stroma and appears to be progressively incorporated into the SfAV1a DNA core during virion assembly. As no other virion structural protein bound DNA and no basic DNA-binding proteins of lower mass are encoded by the SfAV1a genome or were identified by proteomic analysis, our results suggest that P64's function is to condense the large genome of this virus and assist in packaging this genome into its virion.
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proteomic analysis of the spodoptera frugiperda Ascovirus 1a virion reveals 21 proteins
Journal of General Virology, 2009Co-Authors: Yeping Tan, Yves Le Bigot, Dennis K Bideshi, Jeffrey J. Johnson, Brian A. FedericiAbstract:The Spodoptera frugiperda Ascovirus 1a (SfAV-1a) is a double-stranded DNA virus that attacks lepidopteran larvae, in which it produces enveloped virions with complex symmetry which have an average diameter of 130 nm and length of 400 nm. Here, we report identification of 21 SfAV-1a-encoded proteins that occur in the virion, as determined by nano-liquid chromatography/tandem mass spectrometry. These included a helicase (ORF009), nuclease (ORF075), ATPase (ORF047), serine/threonine-like protein kinase (ORF064), inhibitor of apoptosis-like protein (ORF015), thiol oxidoreductase-like protein (ORF061), CTD phosphatase (ORF109), major capsid protein (ORF041) and a highly basic protein, P64 (ORF048). The latter two were the most abundant. Apart from Ascoviruses, the closest orthologues were found in iridoviruses, providing further evidence that Ascoviruses evolved from invertebrate iridoviruses. These results establish a foundation for investigating how Ascovirus virion proteins interact to form their complex asymmetrical structure, as well as for elucidating the mechanisms involved in SfAV-1a virion morphogenesis.
Brian A. Federici - One of the best experts on this subject based on the ideXlab platform.
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Mitochondrial and Innate Immunity Transcriptomes from Spodoptera frugiperda Larvae Infected with the Spodoptera frugiperda Ascovirus.
Journal of virology, 2020Co-Authors: Heba Ahmed Hamed Mohamed Zaghloul, Dennis K Bideshi, Robert H. Hice, Peter Arensburger, Brian A. FedericiAbstract:Ascoviruses are large, enveloped DNA viruses that induce remarkable changes in cellular architecture during which the cell is partitioned into numerous vesicles for viral replication. Previous studies have shown that these vesicles arise from a process resembling apoptosis yet which differs after nuclear lysis in that mitochondria are not degraded but are modified by the virus, changing in size, shape, and motility. Moreover, infection does not provoke an obvious innate immune response. Thus, we used in vivo RNA sequencing to determine whether infection by the Spodoptera frugiperda Ascovirus 1a (SfAV-1a) modified expression of host mitochondrial, cytoskeletal, and innate immunity genes. We show that transcripts from many mitochondrial genes were similar to those from uninfected controls, whereas others increased slightly during vesicle formation, including those for ATP6, ATP8 synthase, and NADH dehydrogenase subunits, supporting electron microscopy (EM) data that these organelles were conserved for virus replication. Transcripts from 58 of 106 cytoskeletal genes studied increased or decreased more than 2-fold postinfection. More than half coded for mitochondrial motor proteins. Similar increases occurred for innate immunity transcripts and their negative regulators, including those for Toll, melanization, and phagocytosis pathways. However, those for many antimicrobial peptides, such as moricin, increased more than 20-fold. In addition, transcripts for gloverin-3, spod_x_tox, Hdd23, and lebocin, also antimicrobial, increased more than 20-fold. Interestingly, a phenoloxidase inhibitor transcript increased 12-fold, apparently to interfere with melanization. SfAV-1a destroys most fat body cells by 7 days postinfection, so innate immunity gene transcripts apparently occur in remaining cells in this tissue and possibly other major tissues, namely, epidermis and tracheal matrix.IMPORTANCE Ascoviruses are large DNA viruses that infect insects, inducing a cellular pathology that resembles apoptosis but which differs by causing enormous cellular hypertrophy followed by cleavage of the cell into numerous viral vesicles for replication. Previous EM studies suggest that mitochondria are important for vesicle formation. Transcriptome analyses of Spodoptera frugiperda larvae infected with SfAV-1a showed that mitochondrial transcripts were similar to those from uninfected controls or increased slightly during vesicle formation, especially for ATP6, ATP8 synthase, and NADH dehydrogenase subunits. This pattern resembles that for chronic disease-inducing viruses, which conserve mitochondria, differing markedly from viruses causing short-term viral diseases, which degrade mitochondrial DNA. Though mitochondrial transcript increases were low, our results demonstrate that SfAV-1a alters host mitochondrial expression more than any other virus. Regarding innate immunity, although SfAV-1a destroys most fat body cells, certain immunity genes were highly upregulated (greater than 20-fold), suggesting that these transcripts may originate from other tissues.
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Ascovirus P64 Homologs: A Novel Family of Large Cationic Proteins That Condense Viral Genomic DNA for Encapsidation
MDPI AG, 2018Co-Authors: Dennis K Bideshi, Yves Le Bigot, Yeping Tan, Tatsinda Spears, Heba A. H. Zaghloul, Brian A. FedericiAbstract:Eukaryotic dsDNA viruses use small basic protamine-like proteins or histones, typically <15 kDa, to condense and encapsidate their genomic (g)DNAs during virogenesis. Ascoviruses are large dsDNA (~100–200 kbp) viruses that are pathogenic to lepidopteran larvae. Little is known about the molecular basis for condensation and encapsidation of their gDNAs. Previous proteomic analysis showed that Spodoptera frugiperda Ascovirus (SfAV-1a) virions contain a large unique DNA-binding protein (P64; 64 kDa, pI = 12.2) with a novel architecture proposed to condense its gDNA. Here we used physical, biochemical, and transmission electron microscopy techniques to demonstrate that P64’s basic C-terminal domain condenses SfAV-1a gDNA. Moreover, we demonstrate that only P64 homologs in other Ascovirus virions are unique in stably binding DNA. As similar protein families or subfamilies were not identified in extensive database searches, our collective data suggest that Ascovirus P64 homologs comprise a novel family of atypical large viral gDNA condensing proteins
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transcriptome analysis of the spodoptera frugiperda Ascovirus in vivo provides insights into how its apoptosis inhibitors and caspase promote increased synthesis of viral vesicles and virion progeny
Journal of Virology, 2017Co-Authors: Robert H. Hice, Peter Arensburger, Heba A. H. Zaghloul, Brian A. FedericiAbstract:Ascoviruses are double-stranded DNA (dsDNA) viruses that attack caterpillars and differ from all other viruses by inducing nuclear lysis followed by cleavage of host cells into numerous anucleate vesicles in which virus replication continues as these grow in the blood. Ascoviruses are also unusual in that most encode a caspase or caspase-like proteins. A robust cell line to study the novel molecular biology of Ascovirus replication in vitro is lacking. Therefore, we used strand-specific transcriptome sequencing (RNA-Seq) to study transcription in vivo in third instars of Spodoptera frugiperda infected with the type species, Spodoptera frugiperda Ascovirus 1a (SfAV-1a), sampling transcripts at different time points after infection. We targeted transcription of two types of SfAV-1a genes; first, 44 core genes that occur in several Ascovirus species, and second, 26 genes predicted in silico to have metabolic functions likely involved in synthesizing viral vesicle membranes. Gene cluster analysis showed differences in temporal expression of SfAV-1a genes, enabling their assignment to three temporal classes: early, late, and very late. Inhibitors of apoptosis (IAP-like proteins; ORF016, ORF025, and ORF074) were expressed early, whereas its caspase (ORF073) was expressed very late, which correlated with apoptotic events leading to viral vesicle formation. Expression analysis revealed that a Diedel gene homolog (ORF121), the only known "virokine," was highly expressed, implying that this Ascovirus protein helps evade innate host immunity. Lastly, single-nucleotide resolution of RNA-Seq data revealed 15 bicistronic and tricistronic messages along the genome, an unusual occurrence for large dsDNA viruses.IMPORTANCE Unlike all other DNA viruses, Ascoviruses code for an executioner caspase, apparently involved in a novel cytopathology in which viral replication induces nuclear lysis followed by cell cleavage, yielding numerous large anucleate viral vesicles that continue to produce virions. Our transcriptome analysis of genome expression in vivo by the Spodoptera frugiperda Ascovirus shows that inhibitors of apoptosis are expressed first, enabling viral replication to proceed, after which the SfAV-1a caspase is synthesized, leading to viral vesicle synthesis and subsequent extensive production of progeny virions. Moreover, we detected numerous bicistronic and tricistronic mRNA messages in the Ascovirus transcriptome, implying that Ascoviruses use other noncanonical translational mechanisms, such as internal ribosome entry sites (IRESs). These results provide the first insights into the molecular biology of a unique coordinated gene expression pattern in which cell architecture is markedly modified, more than in any other known eukaryotic virus, to promote viral reproduction and transmission.
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ICTV virus taxonomy profile: Ascoviridae
Journal of General Virology, 2017Co-Authors: Sassan Asgari, Yves Le Bigot, Dennis K Bideshi, Brian A. Federici, Xiaowen ChengAbstract:The family Ascoviridae includes viruses with circular dsDNA genomes of 100–200 kbp characterized by oblong enveloped virions of 200–400 nm in length. Ascoviruses mainly infect lepidopteran larvae and are mechanically transmitted by parasitoid wasps in which they may also replicate. Most known members belong to the genus Ascovirus, except one virus, that of the genus Toursvirus, which replicates in both its lepidopteran and parasitoid vector hosts. Ascoviruses cause high mortality among economically important insect pests, thereby controlling insect populations. This is a summary of the current International Committee on Taxonomy of Viruses (ICTV) Report on the taxonomy of the Ascoviridae, which is available at www.ictv.global/report/ascoviridae.
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p64 a novel major virion dna binding protein potentially involved in condensing the spodoptera frugiperda Ascovirus 1a genome
Journal of Virology, 2009Co-Authors: Yeping Tan, Yves Le Bigot, Dennis K Bideshi, Jeffrey J. Johnson, Robert H. Hice, Tatsinda Spears, Brian A. FedericiAbstract:We recently identified 21 structural proteins in the virion of Spodoptera frugiperda Ascovirus 1a (SfAV1a), a virus with a large, double-stranded DNA genome of 157 kbp, which attacks species of the lepidopteran family Noctuidae. The two most abundant virion proteins were the major capsid protein and a novel protein (P64) of 64 kDa that contained two distinct domains not known previously to occur together. The amino-terminal half of P64 (residues 1 to 263) contained four repeats (a recently recognized motif with an unknown function) of a virus-specific two-cysteine adaptor. Adjoined to this, the carboxy-terminal half of P64 (residues 279 to 455) contained 14 copies of a highly basic, tandemly repeated motif rich in arginine and serine, having an 11- to 13-amino-acid consensus sequence, SPSQRRSTS(V/K)(A/S)RR, yielding a predicted isoelectric point of 12.2 for this protein. In the present study, we demonstrate by Southwestern analysis that SfAV1a P64 was the only virion structural protein that bound DNA. Additional electrophoretic mobility shift assays showed that P64 bound SfAV1a as well as non-SfAV1a DNA. Furthermore, we show through immunogold labeling of ultrathin sections that P64 is a component of virogenic stroma and appears to be progressively incorporated into the SfAV1a DNA core during virion assembly. As no other virion structural protein bound DNA and no basic DNA-binding proteins of lower mass are encoded by the SfAV1a genome or were identified by proteomic analysis, our results suggest that P64's function is to condense the large genome of this virus and assist in packaging this genome into its virion.
