Monitoring Insect Transposable Elements in Large Double-Stranded DNA Viruses Reveals Host-to-Virus and Virus-to-Virus Transposition.

The mechanisms by which transposable elements (TEs) can be horizontally transferred between animals are unknown, but viruses are possible candidate vectors. Here, we surveyed the presence of host-derived TEs in viral genomes in 35 deep sequencing data sets produced from 11 host-virus systems, encompassing nine arthropod host species (five lepidopterans, two dipterans, and two crustaceans) and six different double-stranded (ds) DNA viruses (four baculoviruses and two iridoviruses). We found evidence of viral-borne TEs in 14 data sets, with frequencies of viral genomes carrying a TE ranging from 0.01% to 26.33% for baculoviruses and from 0.45% to 7.36% for iridoviruses. The analysis of viral populations separated by a single replication cycle revealed that viral-borne TEs originating from an initial host species can be retrieved after viral replication in another host species, sometimes at higher frequencies. Furthermore, we detected a strong increase in the number of integrations in a viral population for a TE absent from the hosts' genomes, indicating that this TE has undergone intense transposition within the viral population. Finally, we provide evidence that many TEs found integrated in viral genomes (15/41) have been horizontally transferred in insects. Altogether, our results indicate that multiple large dsDNA viruses have the capacity to shuttle TEs in insects and they underline the potential of viruses to act as vectors of horizontal transfer of TEs. Furthermore, the finding that TEs can transpose between viral genomes of a viral species sets viruses as possible new niches in which TEs can persist and evolve.

Elucidating the genetic diversity of Phthorimaea operculella granulovirus (PhopGV).

Twelve complete genome sequences of Phthorimaea operculella granulovirus (PhopGV) isolates from four different continents (Africa, South America, Asia and Europe) were analysed after Illumina next-generation sequencing (NGS). The isolates have a circular double-stranded DNA genome that is 118 355 to 119 177 bp in length and all of them encode 130 open reading frames (ORFs). Analysis of single-nucleotide polymorphisms (SNPs) revealed a unique set of SNP positions for every tested isolate. The genome sequences of the investigated PhopGV isolates were classified into a new system of four (1-4) groups according to the presence of group-specific SNPs as well as insertions and deletions. These genome groups correlated with phylogenetic lineages inferred from minimum-evolution trees of the whole-genome consensus nucleotide sequences. All members of group 3 originated from the Mediterranean area, whereas the geographical origin and the group assignment did not correlate for isolates belonging to genome groups 1, 2 or 4. The high degree of coverage facilitated the determination of variant nucleotide frequencies. We conclude that the geographical isolates of PhopGV are genetically highly similar. On the other hand, they were rarely genetically homogenous and in most cases appeared to be mixtures of multiple genotypes.

Agrotis segetum nucleopolyhedrovirus but not Agrotis segetum granulovirus replicate in AiE1611T cell line of Agrotisipsilon.

Both Agrotis segetum nucleopolyhedrovirus B (AgseNPV-B) and Agrotis segetum granulovirus (AgseGV) belong to a cluster of four baculoviruses that are infective for different Agrotis species. Belonging further to different baculovirus genera, namely Alphabaculovirus and Betabaculovirus, respectively, AgseNPV-B and AgseGV are candidates to investigate virus interactions in co-infections. However, for the investigation of virus interactions on a cellular level, permissive insect cell-lines are needed. The cell line AiE1611T deriving from Agrotisipsilon eggs has been shown to be permissive for several Alphabaculovirus isolates. In this study, virus replication was followed based on microscopic analysis of infected and transfected cells, as well as on a molecular level by PCR of DNA and cDNA of selected baculovirus transcripts. While the permissivity was not verified for AgseGV, AgseNPV-B produced occlusion bodies in both infection with hemolymph of infected larvae and Lipofectamin transfection with AgseNPV-B genomic DNA. In addition to the possibility to investigate virus interaction of AgseNPV-B with other alphabaculoviruses, the permissivity of AiE1611T for AgseNPV-B further offers the possibility a biological selection to separate AgseNPV-B from AgseGV.

Deciphering Single Nucleotide Polymorphisms and Evolutionary Trends in Isolates of the Cydia pomonella granulovirus.

Six complete genome sequences of Cydia pomonella granulovirus (CpGV) isolates from Mexico (CpGV-M and CpGV-M1), England (CpGV-E2), Iran (CpGV-I07 and CpGV-I12), and Canada (CpGV-S) were aligned and analyzed for genetic diversity and evolutionary processes. The selected CpGV isolates represented recently identified phylogenetic lineages of CpGV, namely, the genome groups A to E. The genomes ranged from 120,816 bp to 124,269 bp. Several common differences between CpGV-M, -E2, -I07, -I12 and -S to CpGV-M1, the first sequenced and published CpGV isolate, were highlighted. Phylogenetic analysis based on the aligned genome sequences grouped CpGV-M and CpGV-I12 as the most derived lineages, followed by CpGV-E2, CpGV-S and CpGV-I07, which represent the most basal lineages. All of the genomes shared a high degree of co-linearity, with a common setup of 137 (CpGV-I07) to 142 (CpGV-M and -I12) open reading frames with no translocations. An overall trend of increasing genome size and a decrease in GC content was observed, from the most basal lineage (CpGV-I07) to the most derived (CpGV-I12). A total number of 788 positions of single nucleotide polymorphisms (SNPs) were determined and used to create a genome-wide SNP map of CpGV. Of the total amount of SNPs, 534 positions were specific for exactly one of either isolate CpGV-M, -E2, -I07, -I12 or -S, which allowed the SNP-based detection and identification of all known CpGV isolates.

The genome sequence of Agrotis segetum granulovirus, isolate AgseGV-DA, reveals a new Betabaculovirus species of a slow killing granulovirus.

The European isolate Agrotis segetum granulovirus DA (AgseGV-DA) is a slow killing, type I granulovirus due to low dose-mortality responses within seven days post infection and a tissue tropism of infection restricted solely to the fat body of infected Agrotis segetum host larvae. The genome of AgseGV-DA was completely sequenced and compared to the whole genome sequences of the Chinese isolates AgseGV-XJ and AgseGV-L1. All three isolates share highly conserved genomes. The AgseGV-DA genome is 131,557bp in length and encodes for 149 putative open reading frames, including 37 baculovirus core genes and the per os infectivity factor ac110. Comprehensive investigations of repeat regions identified one putative non-hr like origin of replication in AgseGV-DA. Phylogenetic analysis based on concatenated amino acid alignments of 37 baculovirus core genes as well as pairwise distances based on the nucleotide alignments of partial granulin, lef-8 and lef-9 sequences with deposited betabaculoviruses confirmed AgseGV-DA, AgseGV-XJ and AgseGV-L1 as representative isolates of the same Betabaculovirus species. AgseGV encodes for a distinct putative enhancin, distantly related to enhancins from other granuloviruses.

Mortality of cutworm larvae is not enhanced by Agrotis segetum granulovirus and Agrotis segetum nucleopolyhedrovirus B coinfection relative to single infection by either virus.

Mixed infections of insect larvae with different baculoviruses are occasionally found. They are of interest from an evolutionary as well as from a practical point of view when baculoviruses are applied as biocontrol agents. Here, we report mixed-infection studies of neonate larvae of the common cutworm, Agrotis segetum, with two baculoviruses, Agrotis segetum nucleopolyhedrovirus B (AgseNPV-B) and Agrotis segetum granulovirus (AgseGV). By applying quantitative PCR (qPCR) analysis, coinfections of individual larvae were demonstrated, and occlusion body (OB) production within singly infected and coinfected larvae was determined in individual larvae. Mixtures of viruses did not lead to changes in mortality rates compared with rates of single-virus treatments, indicating an independent action within host larvae under our experimental conditions. AgseNPV-B-infected larvae showed an increase in OB production during 2 weeks of infection, whereas the number of AgseGV OBs did not change from the first week to the second week. Fewer OBs of both viruses were produced in coinfections than in singly infected larvae, suggesting a competition of the two viruses for larval resources. Hence, no functional or economic advantage could be inferred from larval mortality and OB production from mixed infections of A. segetum larvae with AgseNPV-B and AgseGV.

The genome sequence of Agrotis segetum nucleopolyhedrovirus B (AgseNPV-B) reveals a new baculovirus species within the Agrotis baculovirus complex.

The genome of Agrotis segetum nucleopolyhedrovirus B (AgseNPV-B) was completely sequenced and compared with whole genome sequences of the Agrotis segetum nucleopolyhedrovirus A (AgseNPV-A) and Agrotis ipsilon nucleopolyhedrovirus (AgipNPV). The AgseNPV-B genome is 148,981 bp in length and encodes 150 putative open reading frames. AgseNPV-B contains two copies of the gene viral enhancing factor (vef), making the Agrotis nucleopolyhedroviruses and A. segetum granulovirus (AgseGV) very rich in vef in comparison to other baculoviruses. Genome alignments of AgseNPV-B, AgseNPV-A and AgipNPV showed a very high genome co-linearity interspersed with variable regions, which are considered as putative sites of genomic recombination. Phylogenetic analyses revealed that all three viruses are distinct. However, AgseNPV-B is more closely related to AgipNPV suggesting that both viruses are at an early stage of phylogenetic divergence. It is proposed that AgseNPV-B belongs to a third Alphabaculovirus species of the Agrotis baculovirus complex. The Agrotis exclamationis nucleopolyhedrovirus (AgexNPV) shared high nucleotide sequence identities with AgseNPV-B, suggesting it is actually an AgseNPV-B isolate.