Viruses of the Fall Armyworm Spodoptera frugiperda: A Review with Prospects for Biological Control.

The fall armyworm (FAW), Spodoptera frugiperda, is a native pest species in the Western hemisphere. Since it was first reported in Africa in 2016, FAW has spread throughout the African continent and is now also present in several countries in Asia as well as Australia. The invasion of FAW in these areas has led to a high yield reduction in crops, leading to huge economic losses. FAW management options in the newly invaded areas are limited and mainly rely on the use of synthetic pesticides. Since there is a risk of resistance development against pesticides in addition to the negative environmental and human health impacts, other effective, sustainable, and cost-efficient control alternatives are desired. Insect pathogenic viruses fulfil these criteria as they are usually effective and highly host-specific with no significant harmful effect on beneficial insects and non-target organisms. In this review, we discuss all viruses known from FAW and their potential to be used for biological control. We specifically focus on baculoviruses and describe the recent advancements in the use of baculoviruses for biological control in the native geographic origin of FAW, and their potential use in the newly invaded areas. Finally, we identify current knowledge gaps and suggest new avenues for productive research on the use of viruses as a biopesticide against FAW.

Bacsnp: Using Single Nucleotide Polymorphism (SNP) Specificities and Frequencies to Identify Genotype Composition in Baculoviruses.

Natural isolates of baculoviruses (as well as other dsDNA viruses) generally consist of homogenous or heterogenous populations of genotypes. The number and positions of single nucleotide polymorphisms (SNPs) from sequencing data are often used as suitable markers to study their genotypic composition. Identifying and assigning the specificities and frequencies of SNPs from high-throughput genome sequencing data can be very challenging, especially when comparing between several sequenced isolates or samples. In this study, the new tool "bacsnp", written in R programming langue, was developed as a downstream process, enabling the detection of SNP specificities across several virus isolates. The basis of this analysis is the use of a common, closely related reference to which the sequencing reads of an isolate are mapped. Thereby, the specificities of SNPs are linked and their frequencies can be used to analyze the genetic composition across the sequenced isolate. Here, the downstream process and analysis of detected SNP positions is demonstrated on the example of three baculovirus isolates showing the fast and reliable detection of a mixed sequenced sample.

Entomopathogenic Viruses in the Neotropics: Current Status and Recently Discovered Species.

The market for biological control of insect pests in the world and in Brazil has grown in recent years due to the unwanted ecological and human health impacts of chemical insecticides. Therefore, research on biological control agents for pest management has also increased. For instance, insect viruses have been used to protect crops and forests around the world for decades. Among insect viruses, the baculoviruses are the most studied and used viral biocontrol agent. More than 700 species of insects have been found to be naturally infected by baculoviruses, with 90% isolated from lepidopteran insects. In this review, some basic aspects of baculovirus infection in vivo and in vitro infection, gene content, viral replication will be discussed. Furthermore, we provide examples of the use of insect viruses for biological pest control and recently characterized baculoviruses in Brazil.

A Nymphalid-Infecting Group I Alphabaculovirus Isolated from the Major Passion Fruit Caterpillar Pest Dione juno juno (Lepidoptera: Nymphalidae).

Baculoviruses are capable of infecting a wide diversity of insect pests. In the 1990s, the Dione juno nucleopolyhedrovirus (DijuNPV) was isolated from larvae of the major passionfruit defoliator pest Dione juno juno (Nymphalidae) and described at ultrastructural and pathological levels. In this study, the complete genome sequence of DijuNPV was determined and analyzed. The circular genome presents 122,075 bp with a G + C content of 50.9%. DijuNPV is the first alphabaculovirus completely sequenced that was isolated from a nymphalid host and may represent a divergent species. It appeared closely related to Orgyia pseudotsugata multiple nucleopolyhedrovirus (OpMNPV) and other Choristoneura-isolated group I alphabaculoviruses. We annotated 153 open reading frames (ORFs), including a set of 38 core genes, 26 ORFs identified as present in lepidopteran baculoviruses, 17 ORFs unique in baculovirus, and several auxiliary genes (e.g., bro, cathepsin, chitinase, iap-1, iap-2, and thymidylate kinase). The thymidylate kinase (tmk) gene was present fused to a dUTPase (dut) gene in other baculovirus genomes. DijuNPV likely lost the dut portion together with the iap-3 homolog. Overall, the genome sequencing of novel alphabaculoviruses enables a wide understanding of baculovirus evolution.

The complete genome of Rachiplusia nu nucleopolyhedrovirus (RanuNPV) and the identification of a baculoviral CPD-photolyase homolog.

We described a novel baculovirus isolated from the polyphagous insect pest Rachiplusia nu. The virus presented pyramidal-shaped occlusion bodies (OBs) with singly-embed nucleocapsids and a dose mortality response of 6.9 × 103 OBs/ml to third-instar larvae of R. nu. The virus genome is 128,587 bp long with a G + C content of 37.9% and 134 predicted ORFs. The virus is an alphabaculovirus closely related to Trichoplusia ni single nucleopolyhedrovirus, Chrysodeixis chalcites nucleopolyhedrovirus, and Chrysodeixis includens single nucleopolyhedrovirus and may constitute a new species. Surprisingly, we found co-evolution among the related viruses and their hosts at species level. Besides, auxiliary genes with homologs in other baculoviruses were found, e.g. a CPD-photolyase. The gene seemed to be result of a single event of horizontal transfer from lepidopterans to alphabaculovirus, followed by a transference from alpha to betabaculovirus. The predicted protein appears to be an active enzyme that ensures likely DNA protection from sunlight.

Genomic sequencing of Troides aeacus nucleopolyhedrovirus (TraeNPV) from golden birdwing larvae (Troides aeacus formosanus) to reveal defective Autographa californica NPV genomic features.

BACKGROUND: The golden birdwing butterfly (Troides aeacus formosanus) is a rarely observed species in Taiwan. Recently, a typical symptom of nuclear polyhedrosis was found in reared T. aeacus larvae. From the previous Kimura-2 parameter (K-2-P) analysis based on the nucleotide sequence of three genes in this isolate, polh, lef-8 and lef-9, the underlying virus did not belong to any known nucleopolyhedrovirus (NPV) species. Therefore, this NPV was provisionally named "TraeNPV". To understand this NPV, the nucleotide sequence of the whole TraeNPV genome was determined using next-generation sequencing (NGS) technology. RESULTS: The genome of TraeNPV is 125,477 bp in length with 144 putative open reading frames (ORFs) and its GC content is 40.45%. A phylogenetic analysis based on the 37 baculoviral core genes suggested that TraeNPV is a Group I NPV that is closely related to Autographa californica nucleopolyhedrovirus (AcMNPV). A genome-wide analysis showed that TraeNPV has some different features in its genome compared with other NPVs. Two novel ORFs (Ta75 and Ta139), three truncated ORFs (pcna, he65 and bro) and one duplicated ORF (38.7 K) were found in the TraeNPV genome; moreover, there are fewer homologous regions (hrs) than there are in AcMNPV, which shares eight hrs within the TraeNPV genome. TraeNPV shares similar genomic features with AcMNPV, including the gene content, gene arrangement and gene/genome identity, but TraeNPV lacks 15 homologous ORFs from AcMNPV in its genome, such as ctx, host cell-specific factor 1 (hcf-1), PNK/PNL, vp15, and apsup, which are involved in the auxiliary functions of alphabaculoviruses. CONCLUSIONS: Based on these data, TraeNPV would be clarified as a new NPV species with defective AcMNPV genomic features. The precise relationship between TraeNPV and other closely related NPV species were further investigated. This report could provide comprehensive information on TraeNPV for evolutionary insights into butterfly-infected NPV.

The complete genome sequence of a second alphabaculovirus from the true armyworm, Mythimna unipuncta: implications for baculovirus phylogeny and host specificity.

The Mythimna unipuncta nucleopolyhedrovirus isolate KY310 (MyunNPV-KY310) is an alphabaculovirus isolated from a true armyworm (Mythimna unipuncta) population in Kentucky, USA. Occlusion bodies of this virus were examined by electron microscopy and the genome sequence was determined by 454 pyrosequencing. MyunNPV-KY310 occlusion bodies consisted of irregular polyhedra measuring 0.8-1.8 µm in diameter and containing multiple virions, with one to six nucleocapsids per virion. The genome sequence was determined to be 156,647 bp with a nucleotide distribution of 43.9% G+C. 152 ORFs and six homologous repeat (hr) regions were annotated for the sequence, including the 38 core genes of family Baculoviridae and an additional group of 26 conserved alphabaculovirus genes. BLAST queries and phylogenetic inference confirmed that MyunNPV-KY310 is most closely related to the alphabaculovirus Leucania separata nucleopolyhedrovirus isolate AH1, which infects Mythimna separata. In contrast, MyunNPV-KY310 did not exhibit a close relationship with Mythimna unipuncta nucleopolyhedrovirus isolate #7, an alphabaculovirus from the same host species. MyunNPV-KY310 lacks the gp64 envelope glycoprotein, which is a characteristic of group II alphabaculoviruses. However, this virus and five other alphabaculoviruses lacking gp64 are placed outside the group I and group II clades in core gene phylogenies, further demonstrating that viruses of genus Alphabaculovirus do not occur in two monophyletic clades. Potential instances of MyunNPV-KY310 ORFs arising by horizontal transfer were detected. Although there are now genome sequences of four different baculoviruses from M. unipuncta, comparison of their genome sequences provides little insight into the genetic basis for their host specificity.

Genome-wide diversity in temporal and regional populations of the betabaculovirus Erinnyis ello granulovirus (ErelGV).

BACKGROUND: Erinnyis ello granulovirus (ErelGV) is a betabaculovirus infecting caterpillars of the sphingid moth E. ello ello (cassava hornworm), an important pest of cassava crops (Manihot esculenta). In this study, the genome of seven field isolates of the virus ErelGV were deep sequenced and their inter- and intrapopulational sequence diversity were analyzed. RESULTS: No events of gene gain/loss or translocations were observed, and indels were mainly found within highly repetitive regions (direct repeats, drs). A naturally occurring isolate from Northern Brazil (Acre State, an Amazonian region) has shown to be the most diverse population, with a unique pattern of polymorphisms. Overall, non-synonymous substitutions were found all over the seven genomes, with no specific gathering of mutations on hotspot regions. Independently of their sizes, some ORFs have shown higher levels of non-synonymous changes than others. Non-core genes of known functions and structural genes were among the most diverse ones; and as expected, core genes were the least variable genes. We observed remarkable differences on diversity of paralogous genes, as in multiple copies of p10, fgf, and pep. Another important contrast on sequence diversity was found on genes encoding complex subunits and/or involved in the same biological processes, as late expression factors (lefs) and per os infectivity factors (pifs). Interestingly, several polymorphisms in coding regions lie on sequences encoding specific protein domains. CONCLUSIONS: By comparing and integrating information about inter- and intrapopulational diversity of viral isolates, we provide a detailed description on how evolution operates on field isolates of a betabaculovirus. Our results revealed that 35-41% of the SNPs of ErelGV lead to amino acid changes (non-synonymous substitutions). Some genes, especially non-core genes of unknown functions, tend to accumulate more mutations, while core genes evolve slowly and are more conserved. Additional studies would be necessary to understand the actual effects of such gene variations on viral infection and fitness.

Baculovirus Kimura two-parameter species demarcation criterion is confirmed by the distances of 38 core gene nucleotide sequences.

Kimura two-parameter nucleotide distance comparisons based on polyhedrin/granulin (polh/gran), late expression factor 8 (lef-8) and late expression factor 9 (lef-9) are a widely applied method for species demarcation for lepidopteran-specific baculoviruses. Baculoviruses are considered to belong to the same species when a pairwise distance threshold of 0.015 is not exceeded and are considered as possibly belonging to the same species with a distance of up to 0.050. In the present work this method was revised and extended for 172 entirely sequenced lepidopteran, hymenopteran and dipteran baculovirus genomes by applying the nucleotide sequences of all 38 known baculovirus core genes for pairwise distance calculations. On the basis of this large dataset, the previously established standard thresholds for baculovirus species demarcation were adjusted for pairwise nucleotide distances estimated from the alignments of all 38 core genes. With the newly applied thresholds for the 38 core-gene dataset, a more sophisticated Kimura two-parameter method was established, avoiding the possible influence of the chimerical polh gene of the Autographa californica multiple nucleopolyhedrovirus. Based on the new dataset, the present classification of baculovirus species was confirmed. Thereby the Kimura two-parameter method for baculovirus demarcation was extended to include the information from all 38 Baculoviridae core genes, which represent the established standard information for baculovirus phylogeny to date.

ICTV Virus Taxonomy Profile: Baculoviridae.

The family Baculoviridae comprises large viruses with circular dsDNA genomes ranging from 80 to 180 kbp. The virions consist of enveloped, rod-shaped nucleocapsids and are embedded in distinctive occlusion bodies measuring 0.15-5 µm. The occlusion bodies consist of a matrix composed of a single viral protein expressed at high levels during infection. Members of this family infect exclusively larvae of the insect orders Lepidoptera, Hymenoptera and Diptera. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the taxonomy of the Baculoviridae, which is available at www.ictv.global/report/baculoviridae.

A Novel Betabaculovirus Isolated from the Monocot Pest Mocis latipes (Lepidoptera: Noctuidae) and the Evolution of Multiple-Copy Genes.

In this report, we described the genome of a novel baculovirus isolated from the monocot insect pest Mocis latipes, the striped grass looper. The genome has 134,272 bp in length with a G + C content of 38.3%. Based on the concatenated sequence of the 38 baculovirus core genes, we found that the virus is a betabaculovirus closely related to the noctuid-infecting betabaculoviruses including Pseudaletia unipuncta granulovirus (PsunGV), Trichoplusia ni granulovirus (TnGV), Helicoverpa armigera granulovirus (HearGV), and Xestia c-nigrum granulovirus (XecnGV). The virus may constitute a new Betabaculovirus species tentatively named Mocis latipes granulovirus (MolaGV). After gene content analysis, five open reading frames (ORFs) were found to be unique to MolaGV and several auxiliary genes were found including iap-3, iap-5, bro-a, bro-b, and three enhancins. The virus genome lacked both chitinase and cathepsin. We then looked at the evolutionary history of the enhancin gene and found that betabaculovirus acquired this gene from an alphabaculovirus followed by several duplication events. Gene duplication also happened to an endonuclease-like gene. Genomic and gene content analyses revealed both a strict collinearity and gene expansion into the genome of the MolaGV-related species. We also characterized the granulin gene using a recombinant Autographa californica multiple nucleopolyhedrovirus (AcMNPV) and found that occlusion bodies were produced into the nucleus of infected cells and presented a polyhedral shape and no occluded virions within. Overall, betabaculovirus genome sequencing is of importance to the field as few genomes are publicly accessible. Mocislatipes is a secondary pest of maize, rice, and wheat crops in Brazil. Certainly, both the discovery and description of novel baculoviruses may lead to development of greener and safer pesticides in order to counteract and effectively control crop damage-causing insect populations.

The complete genome sequence of the first hesperiid-infecting alphabaculovirus isolated from the leguminous pest Urbanus proteus (Lepidoptera: Hesperiidae).

Baculoviruses are insect viruses largely used as expression vectors and biopesticides. These viruses can efficiently infect the larval stage of several agricultural pests worldwide causing a lethal disease. In this work, we found a novel baculovirus isolated from the larval stage of Urbanus proteus (L.), the bean leafroller and characterized its complete genome. This is an important pest of several leguminous plants in Brazil and belongs to the butterfly family Hesperiidae, from where no baculovirus genome sequence has been described. This new virus was shown to have the smallest genome among all alphabaculoviruses sequenced to date, with 105,555 bp and 119 putative ORFs. We found ten unique genes, seven bro, and the 38 baculovirus core genes. UrprNPV was found to be related to the Adoxophyes-infecting baculoviruses AdorNPV and AdhoNPV with high genetic distance and a long branch length. Interestingly, few individual core gene-based phylogenies were found to support the relationship of UrprNPV to both AdorNPV and AdhoNPV. Importantly, the increase in number of completely sequenced baculovirus points to a very exciting way to understand baculovirus and its evolution and could potentially help the use of baculovirus as both biopesticides and expression vectors.

New Method for Differentiation of Granuloviruses (Betabaculoviruses) Based on Multitemperature Single Stranded Conformational Polymorphism.

Baculoviruses have been used as biopesticides for decades. Recently, due to the excessive use of chemical pesticides there is a need for finding new agents that may be useful in biological protection. Sometimes few isolates or species are discovered in one host. In the past few years, many new baculovirus species have been isolated from environmental samples, thoroughly characterized and thanks to next generation sequencing methods their genomes are being deposited in the GenBank database. Next generation sequencing (NGS) methodology is the most certain way of detection, but it has many disadvantages. During our studies, we have developed a method based on Polymerase chain reaction (PCR) followed by Multitemperature Single Stranded Conformational Polymorphism (MSSCP) which allows for distinguishing new granulovirus isolates in only a few hours and at low-cost. On the basis of phylogenetic analysis of betabaculoviruses, representative species have been chosen. The alignment of highly conserved genes-granulin and late expression factor-9, was performed and the degenerate primers were designed to amplify the most variable, short DNA fragments flanked with the most conserved sequences. Afterwards, products of PCR reaction were analysed by MSSCP technique. In our opinion, the proposed method may be used for screening of new isolates derived from environmental samples.

The Operophtera brumata Nucleopolyhedrovirus (OpbuNPV) Represents an Early, Divergent Lineage within Genus Alphabaculovirus.

Operophtera brumata nucleopolyhedrovirus (OpbuNPV) infects the larvae of the winter moth, Operophtera brumata. As part of an effort to explore the pesticidal potential of OpbuNPV, an isolate of this virus from Massachusetts (USA)-OpbuNPV-MA-was characterized by electron microscopy of OpbuNPV occlusion bodies (OBs) and by sequencing of the viral genome. The OBs of OpbuNPV-MA consisted of irregular polyhedra and contained virions consisting of a single rod-shaped nucleocapsid within each envelope. Presumptive cypovirus OBs were also detected in sections of the OB preparation. The OpbuNPV-MA genome assembly yielded a circular contig of 119,054 bp and was found to contain little genetic variation, with most polymorphisms occurring at a frequency of < 6%. A total of 130 open reading frames (ORFs) were annotated, including the 38 core genes of Baculoviridae, along with five homologous repeat (hr) regions. The results of BLASTp and phylogenetic analysis with selected ORFs indicated that OpbuNPV-MA is not closely related to other alphabaculoviruses. Phylogenies based on concatenated core gene amino acid sequence alignments placed OpbuNPV-MA on a basal branch lying outside other alphabaculovirus clades. These results indicate that OpbuNPV-MA represents a divergent baculovirus lineage that appeared early during the diversification of genus Alphabaculovirus.

Genomic analysis of a Trichoplusia ni Betabaculovirus (TnGV) with three different viral enhancing factors and two unique genes.

The complete genome of a Trichoplusia ni granulovirus (TnGV) is described and analyzed. The genome contains 175,360 bp (KU752557), becoming the third largest genome within the genus Betabaculovirus, smaller only than the Xestia c-nigrum GV (XecnGV) (178,733 pb) and the Pseudaletia unipuncta GV (PsunGV) (176,677 pb) genomes. The TnGV genome has a 39.81% C+G content and a total of 180 ORFs were identified, 96 of them in the granulin gene direction and 84 in the opposite direction. A total of 94.38% of the ORFs showed high identity with those of ClanGV, HaGV, and SlGV. Eight homologous regions (hrs) were identified as well as one apoptosis inhibitor (IAP-3). Interestingly, three viral enhancing factors (VEFs) were located in TnGV genome: VEF-1 (orf153), VEF-3 (orf155), and VEF-4 (orf164), additional to another metalloprotease (orf37). Two ORFs were unique to TnGV (orf100 and orf101) and another one was shared by only TnGV and AgseGV (orf2). Eleven of the deduced proteins showed high identity with proteins from nucleopolyhedroviruses, three with proteins from ascoviruses, and one with an entomopoxvirus protein. The largest deduced protein contains 1,213 amino acids (orf43) and the smallest deduced protein contains only 50 amino acids (orf143). Sequence identity and phylogenetic analyses showed that the closest related genomes to TnGV are, to date, those of PsunGV and XecnGV. This genome analysis may contribute to functional research on TnGV, and may form the bases for the utilization of this betabaculovirus as a pest control agent.

A betabaculovirus encoding a gp64 homolog.

BACKGROUND: A betabaculovirus (DisaGV) was isolated from Diatraea saccharalis (Lepidoptera: Crambidae), one of the most important insect pests of the sugarcane and other monocot cultures in Brazil. RESULTS: The complete genome sequence of DisaGV was determined using the 454-pyrosequencing method. The genome was 98,392 bp long, which makes it the smallest lepidopteran-infecting baculovirus sequenced to date. It had a G + C content of 29.7% encoding 125 putative open reading frames (ORF). All the 37 baculovirus core genes and a set of 19 betabaculovirus-specific genes were found. A group of 13 putative genes was not found in any other baculovirus genome sequenced so far. A phylogenetic analysis indicated that DisaGV is a member of Betabaculovirus genus and that it is a sister group to a cluster formed by ChocGV, ErelGV, PiraGV isolates, ClanGV, CaLGV, CpGV, CrleGV, AdorGV, PhopGV and EpapGV. Surprisingly, we found in the DisaGV genome a G protein-coupled receptor related to lepidopteran and other insect virus genes and a gp64 homolog, which is likely a product of horizontal gene transfer from Group 1 alphabaculoviruses. CONCLUSION: DisaGV represents a distinct lineage of the genus Betabaculovirus. It is closely related to the CpGV-related group and presents the smallest genome in size so far. Remarkably, we found a homolog of gp64, which was reported solely in group 1 alphabaculovirus genomes so far.

Genome of Cnaphalocrocis medinalis Granulovirus, the First Crambidae-Infecting Betabaculovirus Isolated from Rice Leaffolder to Sequenced.

Cnaphalocrocis medinalis is a major pest of rice in South and South-East Asia. Insecticides are the major means farmers use for management. A naturally occurring baculovirus, C. medinalis granulovirus (CnmeGV), has been isolated from the larvae and this has the potential for use as microbial agent. Here, we described the complete genome sequence of CnmeGV and compared it to other baculovirus genomes. The genome of CnmeGV is 112,060 base pairs in length, has a G+C content of 35.2%. It contains 133 putative open reading frames (ORFs) of at least 150 nucleotides. A hundred and one (101) of these ORFs are homologous to other baculovirus genes including 37 baculovirus core genes. Thirty-two (32) ORFs are unique to CnmeGV with no homologues detected in the GeneBank and 53 tandem repeats (TRs) with sequence length from 25 to 551 nt intersperse throughout the genome of CnmeGV. Six (6) homologous regions (hrs) were identified interspersed throughout the genome. Hr2 contains 11 imperfect palindromes and a high content of AT sequence (about 73%). The unique ORF28 contains a coiled-coil region and a zinc finger-like domain of 4-50 residues specialized by two C2C2 zinc finger motifs that putatively bound two atoms of zinc. ORF21 encoding a chit-1 protein suggesting a horizontal gene transfer from alphabaculovirus. The putative protein presents two carbohydrate-binding module family 14 (CBM_14) domains rather than other homologues detected from betabaculovirus that only contains one chit-binding region. Gene synteny maps showed the colinearity of sequenced betabaculovirus. Phylogenetic analysis indicated that CnmeGV grouped in the betabaculovirus, with a close relation to AdorGV. The cladogram obtained in this work grouped the 17 complete GV genomes in one monophyletic clade. CnmeGV represents a new crambidae host-isolated virus species from the genus Betabaculovirus and is most closely relative of AdorGV. The analyses and information derived from this study will provide a better understanding of the pathological symptoms caused by this virus and its potential use as a microbial pesticide.

Evolutionary analysis of the ubiquitin gene of baculovirus and insect hosts.

Baculovirus is the only virus that has been found to encode the ubiquitin protein. In this study, ubiquitin sequences from 16 insects and 49 viruses were collected and compared. The resulting sequences were aligned with virus genomes. Then MAGE 5.0, k-estimated software, as well as other software programs were used for systemic evolutionary, selection pressure, and evolutionary distance analysis. The results of the pairwise ratio of non-synonymous to synonymous substitution values and evolutionary distances showed that ubiquitin from baculovirus and insect hosts have been under purifying selection during evolution and are thus evolutionarily conserved. Moreover, genes from insect hosts were more conserved than those in baculovirus. Analysis of the non-synonymous to synonymous substitution rates at each site and entropy calculations revealed the evolutionary status of every site in the ubiquitin genes of baculovirus and their hosts. Genome locations and phylogenetic trees indicated that granuloviruses and non-photosynthetic vegetation evolved, and granulovirus evolution was more similar to that of insect hosts. Our results suggest that the ubiquitin gene in baculovirus may have been acquired through horizontal transfer from the host.

The genome and occlusion bodies of marine Penaeus monodon nudivirus (PmNV, also known as MBV and PemoNPV) suggest that it should be assigned to a new nudivirus genus that is distinct from the terrestrial nudiviruses.

BACKGROUND: Penaeus monodon nudivirus (PmNV) is the causative agent of spherical baculovirosis in shrimp (Penaeus monodon). This disease causes significant mortalities at the larval stage and early postlarval (PL) stage and may suppress growth and reduce survival and production in aquaculture. The nomenclature and classification status of PmNV has been changed several times due to morphological observation and phylogenetic analysis of its partial genome sequence. In this study, we therefore completed the genome sequence and constructed phylogenetic trees to clarify PmNV's taxonomic position. To better understand the characteristics of the occlusion bodies formed by this marine occluded virus, we also compared the chemical properties of the polyhedrin produced by PmNV and the baculovirus AcMNPV (Autographa californica nucleopolyhedrovirus). RESULTS: We used next generation sequencing and traditional PCR methods to obtain the complete PmNV genome sequence of 119,638 bp encoding 115 putative ORFs. Phylogenetic tree analysis showed that several PmNV genes and sequences clustered with the non-occluded nudiviruses and not with the baculoviruses. We also investigated the characteristics of PmNV polyhedrin, which is a functionally important protein and the major component of the viral OBs (occlusion bodies). We found that both recombinant PmNV polyhedrin and wild-type PmNV OBs were sensitive to acid conditions, but unlike the baculoviral OBs, they were not susceptible to alkali treatment. CONCLUSIONS: From the viral genome features and phylogenetic analysis we conclude that PmNV is not a baculovirus, and that it should be assigned to the proposed Nudiviridae family with the other nudiviruses, but into a distinct new genus (Gammanudivirus).

The isolation and genetic characterisation of a South African strain of Phthorimaea operculella granulovirus, PhopGV-SA.

The Phthorimaea operculella granulovirus (PhopGV) is considered a promising biopesticide that can be incorporated into integrated pest management programmes for sustainable control of the potato tuber moth, Phthorimaea operculella (Zeller) (Lepidoptera: Gelechiidae), a major pest of solanaceous crops in sub-tropical and tropical regions worldwide. Several PhopGV isolates recovered from geographically different insect populations have been genetically characterised, and the full genome of the Tunisian PhopGV-1346 isolate has been sequenced, providing a reference strain for comparison of novel isolates. Here we report the identification and genetic characterisation of a South African PhopGV isolate recovered from a P. operculella colony held under laboratory conditions. Transmission electron microscopy examination of purified occlusion bodies together with analysis of granulin and late expression factor-8 (lef-8) gene sequences confirmed the identity of the virus as PhopGV. The sequenced ecdysteroid UDP-glucosyltransferase (egt) gene was 1353nt in length, placing PhopGV-SA in egt group II. Finally, a phylogenetic analysis using a range of egt sequences grouped PhopGV-SA together with the Kenyan, Ecuadorian, Indonesian and Colombian isolates. The results are discussed with reference to the possible origin of PhopGV-SA, and provide a platform for future studies involving virulence evaluation against geographically different P. operculella populations with a view to biopesticide development.