Phylogenetic Position and Replication Kinetics of Heliothis virescens Ascovirus 3h (HvAV-3h) Isolated from Spodoptera exigua

Abstract

Insect-specific ascoviruses with a circular genome are distributed in the USA, France, Australia and Indonesia. Here, we report the first ascovirus isolation from Spodoptera exigua in Hunan, China. DNA-DNA hybridization to published ascoviruses demonstrated that the new China ascovirus isolate is a variant of Heliothis virescens ascovirus 3a (HvAV-3a), thus named HvAV-3h. We investigated the phylogenetic position, cell infection, vesicle production and viral DNA replication kinetics of HvAV-3h, as well as its host-ranges. The major capsid protein (MCP) gene and the delta DNA polymerase (DNA po1) gene of HvAV-3h were sequenced and compared with the available ascovirus isolates for phylogenetic analysis. This shows a close relationship with HvAV-3g, originally isolated from Indonesia, HvAV-3e from Australia and HvAV-3c from United States. HvAV-3h infection induced vesicle production in the SeE1 cells derived from S. exigua and Sf9 cells derived from S. frugiperda , resulting in more vesicles generated in Sf9 than SeE1. Viral DNA replication kinetics of HvAV-3h also demonstrated a difference between the two cell lines tested. HvAV-3h could readily infect three important insect pests Helicoverpa armigera (Hübner), Spodoptera exigua (Hübner) and Spodoptera litura (Fabricius) from two genera in different subfamilies with high mortalities.

Genomic Sequence of Heliothis virescens Ascovirus 3g Isolated from Spodoptera exigua

“…The peptidase also has Y-glutamyl hydrolase activity in humans (1), and GvpU is required to produce gas vesicles in B. megaterium (13). These two genes were not found in the other reported ascoviruses, including the closely related HvAV-3e (12). Five unique homologous regions (hrs) and 25 baculovirus repeated ORFs (bro), which are highly variable, were found in the genome of HvAV-3g.…”

“…A total of 300,000 PE reads were used to assemble a single contig by Edena (11). The assembled contig representing the entire HvAV-3g genome sequence was confirmed by comparing six predicted restriction fragment profiles from the genome, for HindIII, EcoRI, BamHI, XbaI, XhoI, and PstI, with the matching fragment profiles produced by actual restriction digestions of the HvAV-3g genome (7,8,12).…”

Genome sequence and organization analysis of Heliothis virescens ascovirus 3f isolated from a Helicoverpa zea larva

“…A total of 6,244,397 clean pair-end (PE) reads were obtained. Genome assembly was performed using the Edena assembly software package (Hernandez et al, 2008 assembled contig representing the entire HvAV-3f genome sequence was confirmed by six restriction enzyme digestion profiles (BamHI, EcoRI, HindIII, PstI, XbaI and XhoI) that produced the predicted fragments (Cheng et al, 2005;Xue and Cheng, 2011;Huang et al, 2012a). ORFs with 50 aa and longer considered were identified through the GeneMarkS program (Besemer and Borodovsky, 2005), ORF finder (NCBI) and Geneious program (Drummond et al, 2010).…”

“…Based on virion morphology, DNA sequence information, host range and tissue tropism, five ascovirus species have been recognized by the International Committee on Taxonomy of Viruses, which are consisting of Spodoptera frugiperda ascovirus 1a (SfAV-1a), Trichoplusia ni ascovirus 2a (TnAV-2a), Heliothis virescens ascovirus 3a (HvAV-3a), and Diadromus pulchellus ascovirus 4a (DpAV-4a) and Trichoplusia ni ascovirus 6a (ex-TnAV-2c) (Bigot et al, 2011). Eighteen isolates or strains belonging to four ascovirus species have been reported in the world (Bigot et al, 2011;Huang et al, 2012a), in which five ascovirus isolates genomes that are SfAV-1a, TnAV-6a (Trichoplusia ni ascovirus 6a, previously named at its discovery TnAV-2c (Wang et al, 2006)), HvAV-3e, HvAV-3g, and DpAV-4a have been sequenced (Wang et al, 2006;Bideshi et al, 2006;Asgari et al, 2007;Bigot et al, 2009;Huang et al, 2012b). All these reported genomes have a double-stranded circular DNA ranging from 100 to 199 kbp in size (Xue and Cheng, 2011;Huang et al, 2012b).…”

“…All these reported genomes have a double-stranded circular DNA ranging from 100 to 199 kbp in size (Xue and Cheng, 2011;Huang et al, 2012b). Based on the biological characteristics analysis, the species HvAV-3a is the most diverse and widely distributed from America to Asia and Australia (Hamm et al, 1998;Huang et al, 2012a), in which two isolates HvAV-3e and HvAV-3g genomic sequences were reported from Australia and Indonesia, respectively (Asgari et al, 2007;Huang et al, 2012b), but no HvAV-3a isolate from America is sequenced. To understand the diversity and, evolution and geographical differentiation of the HvAV-3a isolates, we report the complete genome of HvAV-3f which was a HvAV-3a isolate from USA, and make comparison for HvAV-3f with the other 5 previously published ascovirus genomes.…”