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The use of CpGV strains as the basis for bioinsecticides is an effective and safe way to control Cydia pomonella. The research is aimed at the identification and study of new CpGV strains. Objects of identification and bioinformatic analysis: 18 CpGV strains. Sequencing was carried out on a NextSeq550. Genome assembly and annotation were carried out using Spades, Samtools 1.9, MinYS, Pilon, Gfinisher, Quast, and Prokka. Comparative genomic analysis was carried out in relation to the reference genome present in the «Madex Tween» strain-producer (biological standard) according to the average nucleotide identity (ANI) criterion. The presence/absence of IAP, cathepsin, MMP, and chitinase in the genetic sequences of the strains was determined using simply phylogeny. Entomopathogenic activity was assessed against C. pomonella according to the criterion of biological efficacy. Thus, molecular genetic identification revealed that 18 CpGV strains belong to a genus of Betabaculovirus. For all the strains under study ANI values of 99% or more were obtained, and the presence of the cathepsin, chitinase, IAP, and MMP genes was noted. The strains BZR GV 1, BZR GV 3, BZR GV 7, BZR GV 10, and BZR GV L-8 showed the maximum biological efficacy: 100% on the 15th day of observation. Strains BZR GV 4, BZR GV 8, and BZR GV 13 showed efficacy at the level of the «Madex Tween» preparation: 89.5% on the 15th day of observation. The strains with the highest mortality rate of the host insect were identified: BZR GV 9, BZR GV 10, BZR GV L-6, and BZR GV L-8. [ABSTRACT FROM AUTHOR]

El virus de la poliedrosis nuclear es un entomopatógeno que se ha usado para el manejo de insecto noctuidae, teniendo un alto porcentaje de efectividad. El objetivo de este estudio fue evaluar la efectividad del virus crudo de la poliedrosis nuclear (líquido) en mezclas de dos aislados virales, virus de la poliedrosis nuclear Spodoptera exigua (SeVPN), virus de la poliedrosis nuclear Spodoptera sunia (SsVPN) en sus propios huéspedes en condiciones de campo y laboratorio. Se utilizó un Diseño de Bloques Completos al Azar con cuatro tratamientos y cuatro repeticiones; siendo las unidades experimentales las larvas sobre alimentadas con hojas infestadas con virus de la poliedrosis nuclear. Los tratamientos evaluados fueron: 1) virus de SeVPN, 2) virus SsVPN, 3) Mezcla de virus de SeVPN+SsVPN (100 %) y 4) Mezcla de Virus de SeVPN+SsVPN (50 %). La aplicación de estos tratamientos se realizó utilizando el virus crudo (liquido) en cada una de las parcelas de cultivo de ajonjolí en horas de la tarde. Después de 24 horas de la aplicación, se colectaron las hojas del cultivo del estrato medio de las plantas para ser trasladadas al laboratorio de virus del Centro de Investigación y Reproducción de Controladores Biológico de la Universidad Nacional Autónoma de Nicaragua (UNAN, León), donde las larvas terminaron su desarrollo. La variable en estudio fue mortalidad. Se realizó un análisis de prueba de normalidad para la variable y se realizó un post hot según SNK en SPSS v. 2.1. No se determinaron diferencias estadísticas entre los tratamientos, pero si valores bajos de mortalidad, siendo de 38 % para Se+SsVPN100%, 39 % para Se+SsVPN50%, SeVPN con 37 % y en el caso de SsVPN de 21 %. La baja mortalidad posiblemente se debió a que en el estudio influyeron factores bióticos como contaminación por bacterias y hongos o por la influencia de la radiación solar en el campo.

The use of CpGV strains as the basis for bioinsecticides is an effective and safe way to control Cydia pomonella. The research is aimed at the identification and study of new CpGV strains. Objects of identification and bioinformatic analysis: 18 CpGV strains. Sequencing was carried out on a NextSeq550. Genome assembly and annotation were carried out using Spades, Samtools 1.9, MinYS, Pilon, Gfinisher, Quast, and Prokka. Comparative genomic analysis was carried out in relation to the reference genome present in the «Madex Tween» strain-producer (biological standard) according to the average nucleotide identity (ANI) criterion. The presence/absence of IAP, cathepsin, MMP, and chitinase in the genetic sequences of the strains was determined using simply phylogeny. Entomopathogenic activity was assessed against C. pomonella according to the criterion of biological efficacy. Thus, molecular genetic identification revealed that 18 CpGV strains belong to a genus of Betabaculovirus. For all the strains under study ANI values of 99% or more were obtained, and the presence of the cathepsin, chitinase, IAP, and MMP genes was noted. The strains BZR GV 1, BZR GV 3, BZR GV 7, BZR GV 10, and BZR GV L-8 showed the maximum biological efficacy: 100% on the 15th day of observation. Strains BZR GV 4, BZR GV 8, and BZR GV 13 showed efficacy at the level of the «Madex Tween» preparation: 89.5% on the 15th day of observation. The strains with the highest mortality rate of the host insect were identified: BZR GV 9, BZR GV 10, BZR GV L-6, and BZR GV L-8.

Grasserie is a major infectious disease of silkworms caused by the Bombyx mori nuclear polyhedrosis virus. The virus belongs to the family Baculoviridae and is composed of a polyhedron with a single molecule of circular supercoiled double-stranded deoxyribose nucleic acid. The disease causes massive mortality (more than 60% of the total crop loss caused by different diseases), causing significant great crop loss to the sericulturists by reducing the cocoon crop and silk produce. There has been no effective solution for preventing and treating the disease. In the present review, the authors explore the subject in detail and inspire the researchers to search for molecular mechanisms of gene action and promote disease-resistant silkworm breeding. [ABSTRACT FROM AUTHOR]

Antecedentes: Actualmente los productores usan insecticidas para las plagas, sin embargo una alternativa viable es el uso del Virus de la Poliedrosis Nuclear. objetivo del trabajo fue reactivar la cepa de VPNHz1991, para el manejo de Helicoverpa zea. Metodología, Se usaron seis tratamientos y seis repeticiones. Los tratamientos fueron: Concentración viral de 93 LE con tres instar larvales y concentración viral de 96 LE con tres instar larval. Se realizó un bioensayo con larvas de instar L1, L2 y L3, con 25 larvas por tratamiento, inoculando los tratamientos en la dieta artificial para que sea ingerida por las larvas. Se obtuvo una mortalidad del 100% sobre los instar L1 y L2 y 97% para instar L3. El tiempo letal que se obtuvo fue de 192 h cumpliendo con lo esperado del actuar del virus. Para determinar el conteo de cuerpos de inclusión poliedral CIP se usó la cámara de Neubauer, donde se usaron dos lotes cosechados de las concentraciones usadas. Resultados: El resultado del conteo para el lote cosechado (116 LE) de la concentración de 93 LE fue de 9.2X10. y para el lote cosechado (119) de la concentración de 96 LE fue de 1.34X1010 con 99% de mortalidad. Conclusiones: Se logró la reactivación de la cepa VPNHz1991 presentando una excelente actividad biológica en larvas de Helicoverpa zea pese a los años de inactividad, demostrando que la cepa viral VPNHz1991 usada en el estudio es apta para su uso como agente de control biológico para el manejo del gusano elotero.

Outbreaks of Anticarsia gemmatalis (Hübner, 1818) (Lepidoptera: Erebidae), a major pest of soybean, can be controlled below economic thresholds with methods that do not involve the application of synthetic insecticides. Formulations based on natural isolates of the Anticarsia gemmatalis multiple nucleopolyhedrovirus (AgMNPV) (Baculoviridae: Alphabaculovirus) played a significant role in integrated pest management programs in the early 2000s, but a new generation of chemical insecticides and transgenic soybean have displaced AgMNPV-based products over the past decade. However, the marked genotypic variability present among and within alphabaculovirus isolates suggests that highly insecticidal genotypic variants can be isolated and used to reduce virus production costs or overcome isolate-dependent host resistance. This study aimed to select novel variants of AgMNPV with suitable insecticidal traits that could complement the existing AgMNPV active ingredients. Three distinct AgMNPV isolates were compared using their restriction endonuclease profile and in terms of their occlusion body (OB) pathogenicity. One isolate was selected (AgABB51) from which eighteen genotypic variants were plaque purified and characterized in terms of their insecticidal properties. The five most pathogenic variants varied in OB pathogenicity, although none of them was faster-killing or had higher OB production characteristics than the wild-type isolate. We conclude that the AgABB51 wild-type isolates appear to be genotypically structured for fast speed of kill and high OB production, both of which would favor horizontal transmission. Interactions among the component variants are likely to influence this insecticidal phenotype. [ABSTRACT FROM AUTHOR]

Autophagy degrades cytoplasmic cargo by its delivery to lysosomes within double membrane autophagosomes. Synthesis of the phosphoinositide PI(3)P by the autophagic class III phosphatidylinositol-3 kinase complex I (PI3KC3-C1) and conjugation of ATG8/LC3 proteins to phagophore membranes by the ATG12-ATG5-ATG16L1 (E3) complex are two critical steps in autophagosome biogenesis, connected by WIPI2. Here, we present a complete reconstitution of these events. On giant unilamellar vesicles (GUVs), LC3 lipidation is strictly dependent on the recruitment of WIPI2 that in turn depends on PI(3)P. Ectopically targeting E3 to membranes in the absence of WIPI2 is insufficient to support LC3 lipidation, demonstrating that WIPI2 allosterically activates the E3 complex. PI3KC3-C1 and WIPI2 mutually promote the recruitment of each other in a positive feedback loop. When both PI 3-kinase and LC3 lipidation reactions were performed simultaneously, positive feedback between PI3KC3-C1 and WIPI2 led to rapid LC3 lipidation with kinetics similar to that seen in cellular autophagosome formation.

Simple Summary: Nucleopolyhedroviruses of caterpillars (Lepidoptera) produce occlusion bodies (OBs) that protect the virions within a protein matrix. This allows the virus to persist in the environment until ingested by a susceptible insect. The soil is an important reservoir of OBs that can be transported to plants for transmission to the host caterpillar. The Spodoptera frugiperda multiple nucleopolyhedrovirus is a lethal pathogen of the fall armyworm, a major pest of maize, rice, and sorghum. Analysis of 186 soil samples collected from maize fields in southern Mexico, Belize, and Guatemala revealed that almost 19% of samples had OBs present at low concentrations. Genetic analysis revealed the presence of genetic diversity in the soil OB populations. The presence of OBs was higher in maize fields with living crops and in specific types of soils. These findings suggest that the soil could be a valuable source of genetic diversity for the design of virus-based insecticides to control this important pest. The occlusion bodies (OBs) of lepidopteran nucleopolyhedroviruses can persist in soil for extended periods before being transported back on to the foliage for transmission to the host insect. A sensitive insect bioassay technique was used to detect OBs of Spodoptera frugiperda multiple nucleopolyhedrovirus (SfMNPV) in 186 soil samples collected from maize fields in the southern Mexican states of Chiapas, Tabasco, Campeche, Yucatán, and Quintana Roo, as well Belize and Guatemala. Overall, 35 (18.8%) samples proved positive for SfMNPV OBs. The frequency of OB-positive samples varied significantly among Mexican states and countries (p < 0.05). Between 1.7 and 4.4% of S. frugiperda larvae that consumed OB-positive samples died from polyhedrosis disease. Restriction endonuclease analysis using PstI and HindIII confirmed that the soil-derived isolates were strains of SfMNPV and that genetic diversity was evident among the isolates. The prevalence of OB-positive soil samples did not differ with altitude or extension (area) of the maize field, but it was significantly higher in fields with the presence of living maize plants compared to those containing dead plants or crop residues (p < 0.05). Georeferenced soil samples were used to identify soil types on digitized soil maps. Lithosol and Luvisol soils had a higher than average prevalence of OB-positive samples (42–45% positive) (p = 0.006), as did Andosol, Gleysol, and Vertisol soils (33–60% OB-positive), although the sample sizes were small (<5 samples) for the latter three soils. In contrast, Cambisol soils had a lower than average prevalence of OB-positive samples (5% positive). Bioassays on Acrisol, Fluvisol, Phaeozem, and Rendzina soils resulted in intermediate levels of OB-positive samples. We conclude that certain soil types may favor OB persistence and virus-mediated biological pest control. The soil is also likely to provide a valuable source of genetic diversity for the design of virus-based insecticides against this pest. [ABSTRACT FROM AUTHOR]

Abstract Background The baculovirus Spodoptera littoralis nucleopolyhedrovirus (SpliNPV) is an entomopathogenic virus utilized as a biological control agent of the Egyptian cotton leaf worm, Spodoptera littoralis. Several studies have focused on the identification of different SpliNPV isolates from a biological and molecular point of view, but few of them conducted in-depth analyses of the genomic composition of these isolates. Results Identification of a novel isolate of SpliNPV, termed Tun2, which was purified from infected S. littoralis larvae from Tunisia was reported. This isolate was propagated in vivo and its median lethal concentration (LC50) was determined to be 1.5 × 104 occlusion bodies (OBs)/ml for third instar S. littoralis larvae at 7 days of post-infection. OB production in late fourth instar larvae was estimated to be at least 2.7 × 109 OBs/g larval weight. The completely sequenced genome of SpliNPV-Tun2 was 137,099 bp in length and contained 132 open reading frames (ORF). It showed a 98.2% nucleotide identity to the Egyptian isolate SpliMNPV-AN1956, with some striking differences; between both genomes, insertion and deletion mutations were noticed in 9 baculovirus core genes, and also in the highly conserved polyhedrin gene. The homologs of ORF 106 and ORF 107 of SpliNPV-AN1956 appeared to be fused to a single ORF 106 in SpliNPV-Tun2, similar to the homologous ORF 110 in SpltNPV-G2. Conclusion SpliNPV-Tun2 is proposed as a new variant of SpliNPV and a potential candidate for further evaluation as a biocontrol agent for S. littoralis and probably other Spodoptera species.

Recruitment of poliovirus (PV) RNA to the human ribosome requires the coordinated interaction of the viral internal ribosome entry site (IRES) and several host cellular initiation factors and IRES trans-acting factors (ITAFs). Attenuated PV Sabin strains contain point mutations in the PV IRES domain V (dV) that inhibit viral translation. Remarkably, attenuation is most apparent in cells of the central nervous system, but the molecular basis to explain this is poorly understood. The dV contains binding sites for eukaryotic initiation factor 4G (eIF4G) and polypyrimidine tract-binding protein (PTB). Impaired binding of these proteins to the mutant IRESs has been observed, but these effects have not been quantitated. We used a fluorescence anisotropy assay to reveal that the Sabin mutants reduce the equilibrium dissociation constants of eIF4G and PTB to the PV IRES by up to 6-fold. Using the most inhibitory Sabin 3 mutant, we used a real-time fluorescence helicase assay to show that the apparent affinity of an active eIF4G/4A/4B helicase complex for the IRES is reduced by 2.5-fold. The Sabin 3 mutant did not alter the maximum rate of eIF4A-dependent helicase activity, suggesting that this mutant primarily reduces the affinity, rather than activity, of the unwinding complex. To confirm this affinity model of attenuation, we show that eIF4G overexpression in HeLa cells overcomes the attenuation of a Sabin 3 mutant PV-luciferase replicon. Our study provides a quantitative framework for understanding the mechanism of PV Sabin attenuation and provides an explanation for the previously observed cell type-specific translational attenuation.

Prior crystal structures of the vault have provided clues of its structural variability but are non-conclusive due to crystal packing. Here, we obtained vaults by engineering at the N terminus of rat major vault protein (MVP) an HIV-1 Gag protein segment and determined their near-atomic resolution (∼4.8 Å) structures in a solution/non-crystalline environment. The barrel-shaped vaults in solution adopt two conformations, 1 and 2, both with D39 symmetry. From the N to C termini, each MVP monomer has three regions: body, shoulder, and cap. While conformation 1 is identical to one of the crystal structures, the shoulder in conformation 2 is translocated longitudinally up to 10 Å, resulting in an outward-projected cap. Our structures clarify the structural discrepancies in the body region in the prior crystallography models. The vault's drug-delivery potential is highlighted by the internal disposition and structural flexibility of its Gag-loaded N-terminal extension at the barrel waist of the engineered vault.

Silk is an economically and culturally important product of Thailand that is threatened by the outbreaks of the Bombyx mori nucleopolyhedrovirus (BmNPV) in silkworm rearings. BmNPV infects the larval stages of the silkworm Bombyx mori, causing fatal viral infection and preventing the larvae from reaching the pupal stage. Numerous BmNPV isolates have been described from silk-producing countries from all over the world. In this study, the geographic distribution of BmNPV in Thailand was analyzed by collecting 18 samples from different regions. The BmNPV samples were analyzed by PCR and whole genome sequencing was performed for eight specimens. Their genome size ranged from 125,888 bp to 126,783 bp, comprising 138 open reading frames. Although the sequenced BmNPV genomes could be phylogenetically differentiated, no correlation between geographic and genetic distance was observed, indicating a close relationship between the BmNPV from Thailand., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

Outbreaks of Anticarsia gemmatalis (Hübner, 1818) (Lepidoptera: Erebidae), a major pest of soybean, can be controlled below economic thresholds with methods that do not involve the application of synthetic insecticides. Formulations based on natural isolates of the Anticarsia gemmatalis multiple nucleopolyhedrovirus (AgMNPV) (Baculoviridae: Alphabaculovirus) played a significant role in integrated pest management programs in the early 2000s, but a new generation of chemical insecticides and transgenic soybean have displaced AgMNPV-based products over the past decade. However, the marked genotypic variability present among and within alphabaculovirus isolates suggests that highly insecticidal genotypic variants can be isolated and used to reduce virus production costs or overcome isolate-dependent host resistance. This study aimed to select novel variants of AgMNPV with suitable insecticidal traits that could complement the existing AgMNPV active ingredients. Three distinct AgMNPV isolates were compared using their restriction endonuclease profile and in terms of their occlusion body (OB) pathogenicity. One isolate was selected (AgABB51) from which eighteen genotypic variants were plaque purified and characterized in terms of their insecticidal properties. The five most pathogenic variants varied in OB pathogenicity, although none of them was faster-killing or had higher OB production characteristics than the wild-type isolate. We conclude that the AgABB51 wild-type isolates appear to be genotypically structured for fast speed of kill and high OB production, both of which would favor horizontal transmission. Interactions among the component variants are likely to influence this insecticidal phenotype.

In rheumatoid arthritis (RA), the proliferation of fibroblast-like synoviocytes (FLS) is the cause of chronic inflammation in joints and of joint damage. Delivery of the pro-apoptotic gene PUMA to FLS via human adenovirus type 5 (HAdV5) vectors has been tested as a therapeutic approach, but efficiency is hampered by low transduction, as FLS do not express HAdV5 receptors on the cell surface. Here we show that efficient transduction of PUMA in FLS can be achieved by conjugating HAdV5 to a baculovirus, which binds to the cell surface via the envelope glycoprotein Gp64. Intra-articular injection in an adjuvant-induced rat model of RA induces apoptosis of FLS, leading to significant decrease in joint inflammation, joint damage, and bone loss with improvement in joint function and mobility. Our results demonstrate the therapeutic potential of PUMA gene therapy as a local treatment in various forms of arthritis in which abnormal FLS proliferation is implicated.Proliferation of synoviocytes contributes to joint damage in rheumatoid arthritis. Here the authors show that targeting of these cells by a vector, consisting of a baculovirus conjugated to an adenovirus carrying the pro-apoptotic gene PUMA, has therapeutic efficacy in a rat arthritis model.

We developed a novel antiviral strategy by combining transposon-based transgenesis and the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) system for the direct cleavage of Bombyx mori nucleopolyhedrovirus (BmNPV) genome DNA to promote virus clearance in silkworms. We demonstrate that transgenic silkworms constitutively expressing Cas9 and guide RNAs targeting the BmNPV immediate early-1 (ie-1) and me53 genes effectively induce target-specific cleavage and subsequent mutagenesis, especially large (∼7-kbp) segment deletions in BmNPV genomes, and thus exhibit robust suppression of BmNPV proliferation. Transgenic animals exhibited higher and inheritable resistance to BmNPV infection than wild-type animals. Our approach will not only contribute to modern sericulture but also shed light on future antiviral therapy.IMPORTANCE Pathogen genome targeting has shown its potential in antiviral research. However, transgenic CRISPR/Cas9 system-mediated viral genome targeting has not been reported as an antiviral strategy in a natural animal host of a virus. Our data provide an effective approach against BmNPV infection in a real-world biological system and demonstrate the potential of transgenic CRISPR/Cas9 systems in antiviral research in other species.

Background: The baculovirus Spodoptera littoralis nucleopolyhedrovirus (SpliNPV) is an entomopathogenic virus utilized as a biological control agent of the Egyptian cotton leaf worm, Spodoptera littoralis. Several studies have focused on the identification of different SpliNPV isolates from a biological and molecular point of view, but few of them conducted in-depth analyses of the genomic composition of these isolates. Results: Identification of a novel isolate of SpliNPV, termed Tun2, which was purified from infected S. littoralis larvae from Tunisia was reported. This isolate was propagated in vivo and its median lethal concentration (LC50) was determined to be 1.5 × 104 occlusion bodies (OBs)/ml for third instar S. littoralis larvae at 7 days of post-infection. OB production in late fourth instar larvae was estimated to be at least 2.7 × 109 OBs/g larval weight. The completely sequenced genome of SpliNPV-Tun2 was 137,099 bp in length and contained 132 open reading frames (ORF). It showed a 98.2% nucleotide identity to the Egyptian isolate SpliMNPV-AN1956, with some striking differences; between both genomes, insertion and deletion mutations were noticed in 9 baculovirus core genes, and also in the highly conserved polyhedrin gene. The homologs of ORF 106 and ORF 107 of SpliNPV-AN1956 appeared to be fused to a single ORF 106 in SpliNPV-Tun2, similar to the homologous ORF 110 in SpltNPV-G2. Conclusion: SpliNPV-Tun2 is proposed as a new variant of SpliNPV and a potential candidate for further evaluation as a biocontrol agent for S. littoralis and probably other Spodoptera species. [ABSTRACT FROM AUTHOR]

Helicoverpa armigera single nucleopolyhedrovirus (HearSNPV) is a virulent pathogen of lepidopterans in the genera Heliothis and Helicoverpa, whereas Helicoverpa armigera multiple nucleopolyhedrovirus (HearSNPV) is a different virus species with a broader host range. This study aimed to examine the consequences of coocclusion of HearSNPV and HearMNPV on the pathogenicity, stability and host range of mixed-virus occlusion bodies (OBs). HearSNPV OBs were approximately 6-fold more pathogenic than HearMNPV OBs, showed faster killing by approximately 13 h, and were approximately 45% more productive in terms of OB production per larva. For coocclusion, H. armigera larvae were first inoculated with HearMNPV OBs and subsequently inoculated with HearSNPV OBs at intervals of 0–72 h after the initial inoculation. When the interval between inoculations was 12–24 h, OBs collected from virus-killed insects were found to comprise 41–57% of HearSNPV genomes, but the prevalence of HearSNPV genomes was greatly reduced (3–4%) at later time points. Quantitative PCR (qPCR) analysis revealed the presence of HearSNPV genomes in a small fraction of multinucleocapsid ODVs representing 0.47–0.88% of the genomes quantified in ODV samples, indicating that both viruses had replicated in coinfected host cells. End-point dilution assays on ODVs from cooccluded mixed-virus OBs confirmed the presence of both viruses in 41.9–55.6% of wells that were predicted to have been infected by a single ODV. A control experiment indicated that this result was unlikely to be due to the adhesion of HearSNPV ODVs to HearMNPV ODVs or accidental contamination during ODV band extraction. Therefore, the disparity between the qPCR and end-point dilution estimates of the prevalence of mixed-virus ODVs likely reflected virus-specific differences in replication efficiency in cell culture and the higher infectivity of pseudotyped ODVs that were produced in coinfected parental cells. Bioassays on H. armigera, Spodoptera frugiperda and Mamestra brassicae larvae revealed that mixed-virus OBs were capable of infecting heterologous hosts, but relative potency values largely reflected the proportion of HearMNPV present in each mixed-virus preparation. The cooccluded mixtures were unstable in serial passage; HearSNPV rapidly dominated during passage in H. armigera whereas HearMNPV rapidly dominated during passage in the heterologous hosts. We conclude that mixed-virus coocclusion technology may be useful for producing precise mixtures of viruses with host range properties suitable for the control of complexes of lepidopteran pests in particular crops, although this requires validation by field testing. [ABSTRACT FROM AUTHOR]

The 'pseudokinase' SgK196 is a protein O-mannose kinase (POMK) that catalyzes an essential phosphorylation step during biosynthesis of the laminin-binding glycan on α-dystroglycan. However, the catalytic mechanism underlying this activity remains elusive. Here we present the crystal structure of Danio rerio POMK in complex with Mg2+ ions, ADP, aluminum fluoride, and the GalNAc-β3-GlcNAc-β4-Man trisaccharide substrate, thereby providing a snapshot of the catalytic transition state of this unusual kinase. The active site of POMK is established by residues located in non-canonical positions and is stabilized by a disulfide bridge. GalNAc-β3-GlcNAc-β4-Man is recognized by a surface groove, and the GalNAc-β3-GlcNAc moiety mediates the majority of interactions with POMK. Expression of various POMK mutants in POMK knockout cells further validated the functional requirements of critical residues. Our results provide important insights into the ability of POMK to function specifically as a glycan kinase, and highlight the structural diversity of the human kinome.

We investigate the relationship between the inherent secondary structure and aggregation propensity of peptides containing chameleon sequences (i.e., sequences that can adopt either α or β structure depending on context) using a combination of replica exchange molecular dynamics simulations, ion-mobility mass spectrometry, circular dichroism, and transmission electron microscopy. We focus on an eight-residue long chameleon sequence that can adopt an α-helical structure in the context of the iron-binding protein from Bacillus anthracis (PDB id 1JIG ) and a β-strand in the context of the baculovirus P35 protein (PDB id 1P35 ). We show that the isolated chameleon sequence is intrinsically disordered, interconverting between α-helical and β-rich conformations. The inherent conformational plasticity of the sequence can be constrained by addition of flanking residues with a given secondary structure propensity. Intriguingly, we show that the chameleon sequence with helical flanking residues aggregates rapidly into fibrils, whereas the chameleon sequence with flanking residues that favor β-conformations has weak aggregation propensity. This work sheds new insights into the possible role of α-helical intermediates in fibril formation.

The occlusion bodies (OBs) of lepidopteran nucleopolyhedroviruses can persist in soil for extended periods before being transported back on to the foliage for transmission to the host insect. A sensitive insect bioassay technique was used to detect OBs of Spodoptera frugiperda multiple nucleopolyhedrovirus (SfMNPV) in 186 soil samples collected from maize fields in the southern Mexican states of Chiapas, Tabasco, Campeche, Yucatán, and Quintana Roo, as well Belize and Guatemala. Overall, 35 (18.8%) samples proved positive for SfMNPV OBs. The frequency of OB-positive samples varied significantly among Mexican states and countries (p < 0.05). Between 1.7 and 4.4% of S. frugiperda larvae that consumed OB-positive samples died from polyhedrosis disease. Restriction endonuclease analysis using PstI and HindIII confirmed that the soil-derived isolates were strains of SfMNPV and that genetic diversity was evident among the isolates. The prevalence of OB-positive soil samples did not differ with altitude or extension (area) of the maize field, but it was significantly higher in fields with the presence of living maize plants compared to those containing dead plants or crop residues (p < 0.05). Georeferenced soil samples were used to identify soil types on digitized soil maps. Lithosol and Luvisol soils had a higher than average prevalence of OB-positive samples (42–45% positive) (p = 0.006), as did Andosol, Gleysol, and Vertisol soils (33–60% OB-positive), although the sample sizes were small (

The design of prophylactic and diagnostic tools specific to animal papillomaviruses is hampered by the difficulties of viral in vitro manipulation and by the scarce availability of dedicated biotechnological tools. This paper reports the production of Ovine Papillomavirus 3 (OaPV3)-based virus-like particles (OaPV3-VLPs) in the baculovirus system and their use to investigate host humoral immune response through the establishment of an indirect ELISA test., Polyclonal sera and monoclonal antibodies were generated against OaPV3-VLPs, and their isotype and reactivity were determined. Additionally, antibodies allowed OaPV3 detection in ovine squamous cell carcinoma (SCC) samples by immunohistochemistry. Results encourage the standardization of OaPV3-specific prophylactic and serological diagnostic tools, and open new perspectives for the study of host-viral interaction and SCC development., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Negatively-supercoiled-ds DNA molecules, including the genomes of baculoviruses, spontaneously wrap around cores of histones to form nucleosomes when present within eukaryotic nuclei. Hence, nucleosome management should be essential for baculovirus genome replication and temporal regulation of transcription, but this has not been documented. Nucleosome mobilization is the dominion of ATP-dependent chromatin-remodeling complexes. SWI/SNF and INO80, two of the best-studied complexes, as well as chromatin modifier TIP60, all contain actin as a subunit. Retrospective analysis of results of AcMNPV time course experiments wherein actin polymerization was blocked by cytochalasin D drug treatment implicate actin-containing chromatin modifying complexes in decatenating baculovirus genomes, shutting down host transcription, and regulating late and very late phases of viral transcription. Moreover, virus-mediated nuclear localization of actin early during infection may contribute to nucleosome management.

More than 30% of acute myeloid leukemia (AML) patients possess activating mutations in the receptor tyrosine kinase FMS-like tyrosine kinase 3 or FLT3. A small-molecule inhibitor of FLT3 (known as quizartinib or AC220) that is currently in clinical trials appears promising for the treatment of AML. Here, we report the co-crystal structure of the kinase domain of FLT3 in complex with quizartinib. FLT3 with quizartinib bound adopts an "Abl-like" inactive conformation with the activation loop stabilized in the "DFG-out" orientation and folded back onto the kinase domain. This conformation is similar to that observed for the uncomplexed intracellular domain of FLT3 as well as for related receptor tyrosine kinases, except for a localized induced fit in the activation loop. The co-crystal structure reveals the interactions between quizartinib and the active site of FLT3 that are key for achieving its high potency against both wild-type FLT3 as well as a FLT3 variant observed in many AML patients. This co-complex further provides a structural rationale for quizartinib-resistance mutations.

The in vivo multiplication of baculovirus Spodoptera frugiperda multiple nucleopolyhedrovirus (SfMNPV) in the specific host Spodoptera frugiperda features the issue of cannibalism of the pest species, with increase in costs and reduction of the final production. Current paper aims at improving the in vivo production method of SfMNPV based on the density of caterpillars and in the period of virus multiplication. Four-day old caterpillars of S. frugiperda were inoculated by virus suspension of isolate 6 (1 x 108 OB.mL-1) of SfMNPV in artificial diet. Caterpillars were kept at 31ºC in two densities (50 and 100 caterpillars/tray) and five periods for virus multiplication (48, 72, 96, 120 and 144 hours). After each period, the caterpillars of the two densities were frozen, weighed and ground to quantify occlusion bodies and the calculation of production parameters. Based on evaluated parameters, densities 50 and 100 caterpillars/tray and 96-h period are adequate for the improvement of in vivo production of isolate 6 of SfMNPV in caterpillars of S. frugiperda. [ABSTRACT FROM AUTHOR]

The enzyme Dicer generates 21-25 nucleotide RNAs that target specific mRNAs for silencing during RNA interference and related pathways. Although their active sites and RNA binding regions are functionally conserved, the helicase domains have distinct activities in the context of different Dicer enzymes. To examine the evolutionary origins of Dicer helicase functions, we investigated two related Dicer enzymes from the thermophilic fungus Sporotrichum thermophile. RNA cleavage assays showed that S. thermophile Dicer-1 (StDicer-1) can process hairpin precursor microRNAs, whereas StDicer-2 can only cleave linear double-stranded RNAs. Furthermore, only StDicer-2 possesses robust ATP hydrolytic activity in the presence of double-stranded RNA. Deletion of the StDicer-2 helicase domain increases both StDicer-2 cleavage activity and affinity for hairpin RNA. Notably, both StDicer-1 and StDicer-2 could complement the distantly related yeast Schizosaccharomyces pombe lacking its endogenous Dicer gene but only in their full-length forms, underscoring the importance of the helicase domain. These results suggest an in vivo regulatory function for the helicase domain that may be conserved from fungi to humans.

Sâu xanh da láng, tên khoa học là Spodoptera exigua (Lepidoptera: Noctuidae) là đối tượng gây hại chủ yếu trên nhiều loại rau màu tại ĐBSCL. Sử dụng thuốc hóa học hầu như không hiệu quả vì sâu đã phát triển tính kháng thuốc. Tác nhân gây bệnh côn trùng, Spodoptera exigua nucleopolyhedrovirus (SeNPV) đã được phát triển như một loại thuốc trừ sâu sinh học thương mãi để kiểm soát đối tượng này. Nghiên cứu được thực hiện nhằm xác định giá trị LC50 và LT50 của virus SeNPV chống lại ấu trùng sâu từ tuổi 1 đến tuổi 5 với phương pháp nhỏ giọt thức ăn. Kết quả cho thấy giá trị LC50 (nồng độ gây chết 50% cá thể) của virus SeNPV đối với sâu tuổi 1, 2, 3, 4 và 5 lần lượt là (2,0-8,6) x 101 OBs/ml; (0,22-20) x 102 OBs/ml; (17-0,31) x 103 OBs/ml; (0,14-7,8) x 105 OBs/ml và (1,5-0,32) x 107 OBs/ml. Thời gian gây chết 50% cá thể (LT50) của sâu từ tuổi 1 đến tuổi 5 dao động lần lượt từ 62,26 tới 69,00 giờ sau khi lây nhiễm (h.p.i); 102,00 tới 113,26 h.p.i; 123,24 tới 127,99 h.p.i; 139,51 tới 152,50 h.p.i và 163,51 tới 180,50 h.p.i.

The Cydia pomonella granulovirus (CpGV) has been used as a biological control agent of codling moth (Cydia pomonella), a severe global pest on pome fruit. Despite the economic importance, our knowledge of its molecular biology is still limited and a detailed picture of its gene expression is still missing. Here, we sequenced the transcriptome of codling moth larvae infected with the Mexican isolate CpGV-M and analyzed the expression of viral genes at 12, 48, and 96 h post infection (hpi). The results showed that two genes (p6.9 and pp31/39K) related to DNA binding of virus production, were highly expressed at 48 and 96 hpi. From 48 to 96 hpi, the expression of genes associated with virus replication and dissemination decreased, whereas the expression of genes related to infectious virion production and per os infectivity increased. This study provides a comprehensive view of CpGV gene expression patterns in host larvae. • The first temporal transcriptome of CpGV in host larvae was assessed using RNAseq. • Gene expression pattern of CpGV in host larvae was similar at 48 and 96 hpi. • p6.9 and pp31/39K were top two highly expressed genes and related to replication. • The expression of genes related to replication/dissemination decreased over time. • The expression of genes related to infectious virion production increased over time. [ABSTRACT FROM AUTHOR]

The fall armyworm, Spodoptera frugiperda (JE Smith, 1797) (Lepidoptera: Noctuidae) is considered the main pest of corn (Zea mays L.) in Brazil and feeds on several other plant species, including cassava (Manihot esculenta Crantz). The food substrate influences the control efficiency of baculovirus and there aren't studies on the effects of the application of baculovirus Spodoptera frugiperda multiple nucleopolyhedrovirus (SfMNPV) on cassava leaves in the mortality of S. frugiperda larvae. The main objectives of this study were to evaluate the efficiency of SfMNPV isolate 6 on S. frugiperda larvae fed on corn and cassava leaves. The food substrates were treated with three baculovirus concentrations (2 x 105, 2 x 106 and 2 x 107 OB mL-1) of a semipurified suspension, and of a commercial formulation (CartuchoVit®), both containing SfMNPV isolate 6, and then were offer to S. frugiperda second instar larvae. The semipurified virus was more virulent than the commercial formulation, regardless of the food substrate provided to insects, when concentrations of 2 x 105 and 2 x 106 OB mL-1 were used. However, there was no difference between treatments when baculovirus suspension of 2 x 107 OB mL-1 was use and the mortality rates were higher than 91%. The mortality was higher when the larvae fed on cassava leaves treated with SfMNPV. Therefore, the food substrate increased the efficiency of SfMNPV, which promises the use of this virus in the management of S. frugiperda in cassava crops. [ABSTRACT FROM AUTHOR]

Naturally occurring isolates of baculoviruses, such as the Bombyx mori nucleopolyhedrovirus (BmNPV), usually consist of numerous genetically different haplotypes. Deciphering the different haplotypes of such isolates is hampered by the large size of the dsDNA genome, as well as the short read length of next generation sequencing (NGS) techniques that are widely applied for baculovirus isolate characterization. In this study, we addressed this challenge by combining the accuracy of NGS to determine single nucleotide variants (SNVs) as genetic markers with the long read length of Nanopore sequencing technique. This hybrid approach allowed the comprehensive analysis of genetically homogeneous and heterogeneous isolates of BmNPV. Specifically, this allowed the identification of two putative major haplotypes in the heterogeneous isolate BmNPV-Ja by SNV position linkage. SNV positions, which were determined based on NGS data, were linked by the long Nanopore reads in a Position Weight Matrix. Using a modified Expectation-Maximization algorithm, the Nanopore reads were assigned according to the occurrence of variable SNV positions by machine learning. The cohorts of reads were de novo assembled, which led to the identification of BmNPV haplotypes. The method demonstrated the strength of the combined approach of short- and long-read sequencing techniques to decipher the genetic diversity of baculovirus isolates.

Polycomb Repressive Complex 2 (PRC2) is essential for gene silencing, establishing transcriptional repression of specific genes by tri-methylating Lysine 27 of histone H3, a process mediated by cofactors such as AEBP2. In spite of its biological importance, little is known about PRC2 architecture and subunit organization. Here, we present the first three-dimensional electron microscopy structure of the human PRC2 complex bound to its cofactor AEBP2. Using a novel internal protein tagging-method, in combination with isotopic chemical cross-linking and mass spectrometry, we have localized all the PRC2 subunits and their functional domains and generated a detailed map of interactions. The position and stabilization effect of AEBP2 suggests an allosteric role of this cofactor in regulating gene silencing. Regions in PRC2 that interact with modified histone tails are localized near the methyltransferase site, suggesting a molecular mechanism for the chromatin-based regulation of PRC2 activity.DOI:http://dx.doi.org/10.7554/eLife.00005.001.

Helicoverpa armigera single nucleopolyhedrovirus (HearSNPV) is a virulent pathogen of lepidopterans in the genera Heliothis and Helicoverpa, whereas Helicoverpa armigera multiple nucleopolyhedrovirus (HearSNPV) is a different virus species with a broader host range. This study aimed to examine the consequences of coocclusion of HearSNPV and HearMNPV on the pathogenicity, stability and host range of mixed-virus occlusion bodies (OBs). HearSNPV OBs were approximately 6-fold more pathogenic than HearMNPV OBs, showed faster killing by approximately 13 h, and were approximately 45% more productive in terms of OB production per larva. For coocclusion, H. armigera larvae were first inoculated with HearMNPV OBs and subsequently inoculated with HearSNPV OBs at intervals of 0–72 h after the initial inoculation. When the interval between inoculations was 12–24 h, OBs collected from virus-killed insects were found to comprise 41–57% of HearSNPV genomes, but the prevalence of HearSNPV genomes was greatly reduced (3–4%) at later time points. Quantitative PCR (qPCR) analysis revealed the presence of HearSNPV genomes in a small fraction of multinucleocapsid ODVs representing 0.47–0.88% of the genomes quantified in ODV samples, indicating that both viruses had replicated in coinfected host cells. End-point dilution assays on ODVs from cooccluded mixed-virus OBs confirmed the presence of both viruses in 41.9–55.6% of wells that were predicted to have been infected by a single ODV. A control experiment indicated that this result was unlikely to be due to the adhesion of HearSNPV ODVs to HearMNPV ODVs or accidental contamination during ODV band extraction. Therefore, the disparity between the qPCR and end-point dilution estimates of the prevalence of mixed-virus ODVs likely reflected virus-specific differences in replication efficiency in cell culture and the higher infectivity of pseudotyped ODVs that were produced in coinfected parental cells. Bioassays on H. armigera, Spodoptera frugiperda and Mamestra brassicae larvae revealed that mixed-virus OBs were capable of infecting heterologous hosts, but relative potency values largely reflected the proportion of HearMNPV present in each mixed-virus preparation. The cooccluded mixtures were unstable in serial passage; HearSNPV rapidly dominated during passage in H. armigera whereas HearMNPV rapidly dominated during passage in the heterologous hosts. We conclude that mixed-virus coocclusion technology may be useful for producing precise mixtures of viruses with host range properties suitable for the control of complexes of lepidopteran pests in particular crops, although this requires validation by field testing.

A biopesticide that combines Metarhizium rileyi and nucleopolyhedrovirus (NPV) in an oil dispersion (OD) was developed. Its efficacy against Anticarsia gemmatalis and Chrysodeixis includens was evaluated under laboratory and field conditions. First, each of the two selected fungal strains, combined or not in OD preparations with AgMNPV or ChinNPV, was tested in the laboratory against A. gemmatalis and C. includens, respectively. AgMNPV–fungus combinations had similar effect on overall mortality of A. gemmatalis compared to each pathogen used alone. C. includens was less susceptible to infection by M. rileyi strains, and ChinNPV became the major cause of larval death. The OD formulations with M. rileyi and/or NPV were applied in infested soybean fields. ChinNPV + M. rileyi caused 50.8% of overall mortality six days after spraying in a soybean crop infested by both pests, whereas for treatments with the fungus or the virus alone the percentages were 41.2% and 8.8%, respectively. In another field infested solely by A. gemmatalis, insect mortality was similar for the AgMNPV–fungus combination and the virus used alone. Larval parasitism by wasps and dipterans was not affected following biopesticide spraying. Our results suggest that the performance of a dual‐action biopesticide is related to the NPV and fungal strains that are combined and the proportion of host populations simultaneously infesting the crop. [ABSTRACT FROM AUTHOR]

The co-evolution between baculoviruses and their insect hosts results in selection of virus populations. To explore this phenomenon at the molecular level, seven natural isolates of Cydia pomonella granulovirus (CpGV) collected from orchards in northwest China were studied using Illumina next generation sequencing (NGS). A total of 540 genome positions with single nucleotide polymorphisms (SNPs) were detected in comparison with known CpGV isolates. New members of previously defined phylogenetic genome groups A, D and E of CpGV, as well as two novel phylogenetic lines, termed genome group F and G, were identified. Combining SNP frequency distribution with the prevalence of genome group-specific SNPs, revealed that six isolates of CpGV were mixtures of different ratios of at least two genotypes, whereas only one isolate, CpGV-WW, was genetically highly homogeneous. This study significantly extends our current understanding of the genetic diversity of CpGV and opens new lines of application of this virus. Image 1 • Genome analyses of seven natural Cydia pomonella granulovirus isolates from China. • Isolates are genotype mixtures belonging to known and new phylogenetic lines. • Analyses of single nucleotide polymorphisms (SNPs) reveal genotype mixtures. • Gene pe38 variability correlates with type I resistance-breaking activity. [ABSTRACT FROM AUTHOR]

Cydia pomonella granulovirus (CpGV) is successfully used worldwide as a biocontrol agent of the codling moth (CM) (Cydia pomonella). The occurrence of CM populations with different modes of resistance against commercial CpGV preparations in Europe, as well as the invasiveness of CM in China, threatening major apple production areas there, requires the development of new control options. Utilizing the naturally occurring genetic diversity of CpGV can improve such control strategies. Here, we report the identification of seven new CpGV isolates that were collected from infected CM larvae in northwest China. Resistance testing using a discriminating CpGV concentration and the determination of the median lethal concentration (LC50) were performed to characterize their levels of virulence against susceptible and resistant CM larvae. The isolates were further screened for the presence of the 2x12-bp-repeat insertion in CpGV gene pe38 (open reading frame 24 [ORF24]), which was shown to be the target of type I resistance. It was found that three isolates, CpGV-JQ, -KS1, and -ZY2, could break type I resistance, although delayed mortality was observed in the infection process. All isolates followed the pe38 model of breaking type I resistance, except for CpGV-WW, which harbored the genetic factor but failed to overcome type I resistance. However, CpGV-WW was able to overcome type II and type III resistance. The bioassay results and sequencing data of pe38 support previous findings that pe38 is the major target for type I resistance. The new isolates show some distinct virulence characteristics when infection of different CM strains is considered. [ABSTRACT FROM AUTHOR]

Baculoviruses are highly host specific, and their host range is usually restricted to a single or a few closely related insect species, except for few virus species, e.g. Alphabaculovirus aucalifonicae and Alphabaculovirus mabrassicae . In this study, two new alphabaculovirus isolates were isolated from the larvae of Mamestra brassicae and Mythimna separata , which were named as Mamestra brassicae multiple nucleopolyhedrovirus isolate QD (MbMNPV-QD) and Mythimna separata multiple nucleopolyhedrovirus isolate Hb (MyseMNPV-Hb), respectively. The Kimura two-parameter values based on the concatenated 38 core genes of baculovirus revealed that MbMNPV (isolates QD/CHb1/K1/CTa), MyseMNPV-Hb, Helicoverpa armigera multiple nucleopolyhedrovirus (HearMNPV) and Mamestra configurata nucleopolyhedrovirus B (MacoNPV-B) were different isolates of a same virus species. A phylogenetic tree of baculoviruses and nudiviruses constructed from their 20 homologous gene sequences, and that of their isolated hosts constructed from 13 protein-coding genes of the insect mitochondrial genomes, were used to analyse the coevolution of baculoviruses with their isolated hosts. The results showed that M. brassicae was the most likely ancestral host of these virus isolates, included MbMNPV isolates, MyseMNPV-Hb, HearMNPV, and MacoNPV-B. Therefore, we concluded that these virus isolates belong to the existing virus species - Alphabaculovirus mabrassicae with M. brassicae as their ancestral host.

Background: Spodoptera litura is a harmful pest that feeds on more than 80 species of plants, and can be infected and killed by Spodoptera litura nucleopolyhedrovirus (SpltNPV). SpltNPV-C3 is a type C SpltNPV clone, that was observed and collected in Japan. Compared with type A or type B SpltNPVs, SpltNPV-C3 can cause the rapid mortality of S. litura larvae., Methods: In this study, occlusion bodies (OBs) and occlusion-derived viruses (ODVs) of SpltNPV-C3 were purified, and OBs were observed by scanning electron microscopy (SEM). ODVs were observed under a transmission electron microscope (TEM)., Results: Both OBs and ODVs exhibit morphological characteristics typical of nucleopolyhedroviruses (NPVs).The genome of SpltNPV-C3 was sequenced and analyzed; the total length was 148,634 bp (GenBank accession 780,426,which was submitted as SpltNPV-II), with a G + C content of 45%. A total of 149 predicted ORFs were found. A phylogenetic tree of 90 baculoviruses was constructed based on core baculovirus genes. LC‒MS/MS was used to analyze the proteins of SpltNPV-C3; 34 proteins were found in the purified ODVs, 15 of which were core proteins. The structure of the complexes formed by per os infectivity factors 1, 2, 3 and 4 (PIF-1, PIF-2, PIF-3 and PIF-4) was predicted with the help of the AlphaFold multimer tool and predicted conserved sequences in PIF-3. SpltNPV-C3 is a valuable species because of its virulence, and the analysis of its genome and proteins in this research will be beneficial for pest control efforts., (© 2024. The Author(s).)

Background: Equine influenza (EI) is a transmissible viral respiratory sickness of the Equidae family. Two viruses, H7N7 and H3N8 caused EI; however, H7N7 has not been detected for decades. H3N8 has circulated and bifurcated into Eurasian and American lineages. The latter subsequently diversified into Kentucky, South America, and Florida sub-lineages. Florida clade 1 (FC1) and Florida clade 2 (FC2) strains are the only circulating EI viruses (EIVs) in the meantime. Immunization is considered the major means for the prevention and control of EI infection. Using disparate technologies and platforms, several vaccines have been developed and commercialized. According to the recommendations of the World Organization for Animal Health (WOAH), all commercial vaccines shall comprise representatives of both FC1 and FC2 strains. Unfortunately, most of the commercially available vaccines were not updated to incorporate a representative of FC2 strains., Aim: The purpose of this research was to develop a new EI vaccine candidate that incorporates the hemagglutinin (HA) antigen from the currently circulating FC2., Methods: In this study, we report the expression of the full-length recombinant HA gene of FC2 in the baculovirus expression system., Results: The HA recombinant protein has been proven to maintain its biological characteristics by hemadsorption (HAD) and hemagglutination tests. Moreover, using a reference-specific serum, the specificity of the HA has been confirmed through the implementation of immunoperoxidase and western immunoblotting assays., Conclusion: In conclusion, we report the expression of specific biologically active recombinant HA of FC2, which would act as a foundation for the generation of an updated EI subunit or virus vector vaccine candidates., Competing Interests: The authors affirm that they have no conflict of interest to disclose.

The mulberry silkworm, Bombyx mori (L.), is a model organism of lepidopteran insects with high economic importance. The viral diseases of the silkworm caused by Bombyx mori nucleopolyhedrovirus (BmNPV) and Bombyx mori bidensovirus (BmBDV) inflict huge economic losses and significantly impact the sericulture industry of India and other countries. To understand the distribution of Indian isolates of the BmNPV and to investigate their genetic composition, an in-depth population structure analysis was conducted using comprehensive and newly developed genomic analysis methods. The seven new Indian BmNPV isolates from Anantapur, Dehradun, Ghumarwin, Jammu, Kashmir, Mysore and Salem grouped in the BmNPV clade, and are most closely related to Autographa californica multiple nucleopolyhedrovirus and Rachiplusia ou multiple nucleopolyhedrovirus on the basis of gene sequencing and phylogenetic analyses of the partial polh, lef-8 and lef-9 gene fragments. The whole genome sequencing of three Indian BmNPV isolates from Mysore (-My), Jammu (-Ja) and Dehradun (-De) was conducted, and intra-isolate genetic variability was analyzed on the basis of variable SNP positions and the frequencies of alternative nucleotides. The results revealed that the BmNPV-De and BmNPV-Ja isolates are highly similar in their genotypic composition, whereas the population structure of BmNPV-My appeared rather pure and homogenous, with almost no or few genetic variations. The BmNPV-De and BmNPV-Ja samples further contained a significant amount of BmBDV belonging to the Bidnaviridae family. We elucidated the genotype composition within Indian BmNPV and BmBDV isolates, and the results presented have broad implications for our understanding of the genetic diversity and evolution of BmNPV and co-occurring BmBDV isolates.

Graphical abstract Highlights • NPV + Metarhizium rileyi co-infected Anticarsia gemmatalis and Spodoptera frugiperda. • Advantages in sequential applications of NPV and Metarhizium rileyi were not observed. • Simultaneous applications of NPV + Metarhizium rileyi showed an additive effect. • Mortalities of A. gemmatalis using mixed pathogens were higher than for M. rileyi alone. • Mortalities of S. frugiperda were similar for mixed pathogens and each pathogen alone. Abstract Members of the family Baculoviridae have been quite successfully used as biocontrol agents against some lepidopterans. Likewise, a number of fungi are important natural enemies of these pests. An interesting approach to increase control efficacy could be the combination of a given nucleopolyhedrovirus (NPV) and a fungus, since they possess distinct modes of action. As a first step towards this goal, we assessed the interaction between NPV (either AgMNPV-79 or SfMNPV-6nd) and the entomopathogenic fungus Metarhizium rileyi (either CG1153 or CG381), using Anticarsia gemmatalis and Spodoptera frugiperda as hosts. In sequential applications of these pathogens, per os inoculation of an NPV (leaf discs with 2.5 × 104 occlusion bodies) either two days before or two days post-spraying of its counterpart fungal strain (5 × 103 conidia.cm−2 sprays) usually resulted in an antagonistic effect. When both pathogens were simultaneously applied at different combined dosages, usually an additive effect was seen. Interestingly, a number of dead larvae showing signs of co-infections (partially with soft integument and partially mummified) were recorded. However, mixes with lower dosages of both pathogens did not cause significantly higher insect mortalities compared to low dosages of the fungus applied alone. The advantages and disadvantages of the simultaneous applications of NPV and M. rileyi aiming at the management of either A. gemmatalis or S. frugiperda were discussed. [ABSTRACT FROM AUTHOR]