Greenhouse Evaluation of Dose– and Time–Mortality Relationships of Two Nucleopolyhedroviruses for the Control of Beet Armyworm, Spodoptera exigua, on Chrysanthemum

Dose– and time–mortality relationships of baculoviruses in pest insects are important for the determination of effective spraying regimes. A series of experiments with Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV) and Spodoptera exigua MNPV (SeMNPV) against synchronized populations of S. exigua larvae in greenhouse chrysanthemum was conducted. Dose– and time–mortality relationships of different virus concentrations and S. exigua target stages were determined and the area foliage consumption was measured. Crop injury was greatly reduced when S. exigua were controlled as second or third instar larvae, whereas virus applications against fourth instar larvae could not prevent considerable crop injury, even at high concentrations. SeMNPV was approximately 10 times as infectious as AcMNPV when applied on greenhouse chrysanthemum. The relative virulence of AcMNPV and SeMNPV corresponded reasonably well with previously published laboratory bioassay data. SeMNPV killed second and fourth instar S. exigua larvae approximately 12 h faster than did AcMNPV in chrysanthemum, but no difference in speed of action was found for third instar larvae. The relative speed of action of AcMNPV and SeMNPV determined in chrysanthemum and in laboratory bioassays did not correspond for third instar S. exigua larvae; laboratory bioassay data can therefore not simply be extrapolated to the crop level.     

The influence of greenhouse chrysanthemum on the interaction between the beet armyworm, Spodoptera exigua, and the baculovirus SeMNPV: parameter quantification for a process-based simulation model

During the building of a process-based simulation model for the epidemiology of the multicapsid nucleopolyhedrovirus of S. exigua (SeMNPV) in populations of Spodoptera exigua (Hübner) in greenhouse chrysanthemum, it was found that the effect of host plants had been under-rated. 'Missing links' included (i) the 'natural' background mortality of larvae of S. exigua in practical cropping conditions; (ii) the developmental rate of larvae of S. exigua on plant substrate in a glasshouse as compared to artificial medium in the laboratory; (iii) the validity of the results of dose-mortality and time-mortality bioassays conducted on artificial medium as compared to natural plant substrate; (iv) the distribution of inoculum released from deceased caterpillars over chrysanthemum leaves; and (v) the leaf visit rate of healthy caterpillars (as it affects horizontal transmission). Experiments were carried out to quantify these processes. Developmental rates of S. exigua larvae on greenhouse chrysanthemum were 36% lower than on an artificial diet. The fraction survival during the first, second, third and fourth instar S. exigua larvae in greenhouse chrysanthemum was 0.60, 0.80, 0.88 and 0.95, respectively. Forty percent of the first instar larvae reached the fifth larval stage. Second instar S. exigua larvae reared on chrysanthemum were significantly more susceptible to SeMNPV than larvae reared on an artificial diet. The food source had no effect on the time to kill S. exigua larvae. Cadavers of second, third and fourth instar S. exigua larvae contaminated on average 1.4, 2.5 and 3.3 chrysanthemum leaves. Second to fourth instar S. exigua larvae visited 2–3 leaves per day and spent 15–55% of the time on the underside of leaves. The above information is of critical importance for a trustworthy simulation of the epidemiology of SeMNPV in chrysanthemum.     

Selection of nuclear polyhedrosis viruses as biological control agents ofSpodoptera exigua [Lep.: Noctuidae]

The virulence of 5 nuclear polyhedrosis viruses infectious for larvae of beet armyworm,Spodoptera exigua, was studied and their potential as biological control agents of this accidentally introduced pest in Dutch greenhouse crops is discussed. Three of the virus isolates were collected from deceased beet armyworm larvae found in Dutch greenhouses. Based on restriction endonuclease patterns of their DNA they appeared to be closely related toMamestra brassicae nuclear polyhedrosis virus (MbMNPV) and therefore were named MbMNPV-NL80, MbMNPV-NL82 and MbMNPV-NL83. These isolates were not related toAutographa californica MNPV (AcMNPV) or toSpodoptera exigua MNPV (SeMNPV), both originating from the USA. Comparison of the oiological activity of these 5 isolates showed that the SeMNPV was more virulent against beet armyworm than the other isolates. There was no significant difference in virulence between MbMNPV-NL80, NL82, NL83 and AcMNPV forS. exigua. The LD-50 values of the 5 isolates for 2nd instar larvae were 3, 26, 14, 17 and 18 polyhedra, respectively. Despite compensating qualities of the other MNPVs, such as a broader host range and potential production in alternate hosts or cell-lines, SeMNPV is considered to be the most suitable candidate as biological control agent of beet armyworm.      

Baculovirus infection triggers a positive phototactic response in caterpillars to induce ‘tree-top’ disease

Many parasites manipulate host behaviour to enhance parasite transmission and survival. A fascinating example is baculoviruses, which often induce death in caterpillar hosts at elevated positions (‘tree-top’ disease). To date, little is known about the underlying processes leading to this adaptive host manipulation. Here, we show that the baculovirus Spodoptera exigua multiple nucleopolyhedrovirus (SeMNPV) triggers a positive phototactic response in S. exigua larvae prior to death and causes the caterpillars to die at elevated positions. This light-dependent climbing behaviour is specific for infected larvae, as movement of uninfected caterpillars during larval development was light-independent. We hypothesize that upon infection, SeMNPV captures a host pathway involved in phototaxis and/or light perception to induce this remarkable behavioural change.     

Chemical and biological stress factors on the activation of nucleopolyhedrovirus infections in covertly infected Spodoptera exigua

Following the consumption of baculovirus occlusion bodies (OBs), insects may succumb to lethal disease, but the survivors can harbour sublethal covert infections and may develop, reproduce and transmit the infection to their offspring. The use of different chemical and biological stressors was examined to determine whether they could be used to activate covert infections in populations of Spodoptera exigua larvae infected by the homologous nucleopolyhedrovirus (SeMNPV). Treatment of covertly infected S. exigua second instars with Tinopal UNPA‐GX, hydroxylamine, paraquat, Bacillus thuringiensis var. kurstaki crystals, spores or mixtures of crystals + spores, or a heterologous nucleopolyhedrovirus (Chrysodeixis chalcites SNPV) did not result in the activation of SeMNPV covert infections. Similarly, virus treatments involving permissive NPVs did not result in greater mortality in covertly infected insects compared with the virus‐free controls. In contrast, 0.1% copper sulphate, 1% iron (II) sulphate and 1 mg/l sodium selenite treatments resulted in 12–41% lethal polyhedrosis disease in covertly infected larvae. A greenhouse trial using copper sulphate and sodium selenite as activation factors applied to covertly infected S. exigua larvae on sweet pepper plants resulted in very low levels of SeMNPV activation (<3%). These results highlight the important roles of copper, iron and selenium in insect immunity and baculovirus‐induced disease. However, these substances seem unlikely to prove useful for the activation of covert SeMNPV infections in S. exigua larvae under greenhouse conditions.     

Competition between wild-type and a marked recombinant baculovirus (Spodoptera exigua nucleopolyhedrovirus) with enhanced speed of action in insect larvae

Competition between virus genotypes in insect hosts is a key element of virus fitness, affecting their long-term persistence in agro-ecosystems. Little information is available on virus competition in insect hosts or during serial passages from one cohort of hosts to the next. Here we report on the competition between two genotypes of Spodoptera exigua nucleopolyhedrovirus (SeMNPV), when serially passaged as mixtures in cohorts of 4th instar S. exigua larvae. One of the genotypes was a SeMNPV wild-type isolate, SeUS1, while the other was a SeMNPV recombinant (SeMNPV-XD1) having a greater speed of kill than SeUS1. SeXD1 lacks a suite of genes, including the ecdysteroid UDP-glucosyl transferase (egt) gene. SeXD1 expresses the green fluorescent protein (GFP) gene, enabling the identification of SeXD1 in cell culture and in insects. The relative proportion of SeUS1 and SeXD1 in successive passages of mixed infections in various ratios was determined by plaque assays of budded virus from infected larvae and by polymerase chain reactions and restriction enzyme analyses. The SeUS1 genotype outcompeted recombinant SeXD1 over successive passages. Depending on the initial virus genotype ratio, the recombinant SeXD1 was no longer detected after 6-12 passages. A mathematical model was developed to characterize the competition dynamics. Overall, the ratio SeUS1/XD1 increased by a factor 1.9 per passage. The findings suggest that under the experimental conditions recombinant SeXD1 is displaced by the wild-type strain SeUS1, but further studies are needed to ascertain that this is also the case when the same baculoviruses would be used in agro-ecosystems.     

Insecticidal activity of recombinant baculovirus co-expressing Bacillus thuringiensis crystal protein and Kunitz-type toxin isolated from the venom of bumblebee Bombus ignitus

To improve the insecticidal activity of Autographa californica nucleopolyhedrovirus (AcMNPV), using co-expression of Bacillus thuringiensis crystal protein and a Kunitz-type toxin isolated from bumblebee Bombus ignitus venom, a recombinant AcMNPV, ApPolh5-3006BiKTI, expressing Bi-KTI under the control of early promoter from Cotesia plutellae bracovirus (CpBV) was constructed. In this recombinant virus, B. thuringiensis cry1-5 crystal protein gene was introduced into the genome by the fusion of polyhedrin-cry1-5 under the control of polyhedrin gene promoter. RT-PCR analysis indicated that both Bi-KTI and polyhedrin-cry1-5 fusion protein were successfully expressed from the infected cells. In addition, SDS-PAGE revealed that polyhedrin-cry1-5 fusion protein expressed by recombinant viruses was occluded into the polyhedra. ApPolh5-3006BiKTI showed an improved insecticidal activity against larvae of Plutella xylostella and Spodoptera exigua. At low dosage rates, it was more effective against S. exigua than on P. xylostella, but more rapid insecticidal activity was shown in P. xylostella. These results strongly suggest that co-expression of Bt toxin and Kunitz-type toxins could be successfully applied to improve the insecticidal activity of baculoviruses.     

Activity and Persistence of Steinernema carpocapsae and Spodoptera exigua Nuclear Polyhedrosis Virus against S. exigua Larvae on Soybean

Experiments were conducted to assess the effects of the entomopathogenic nematode Steinernema carpocapsae and Spodoptera exigua multinucleocapsid nuclear polyhedrosis virus (SeMNPV), alone and in combinations, on mortality of the beet armyworm, S. exigua, larvae on soybean. In 1991 tests, field-grown soybean plants were treated with S. carpocapsae at 0.3 and 0.6 nematodes/cm² of leaflet, SeMNPV at 20 and 40 polyhedral inclusion bodies (PIB)/cm², and all possible combinations. Treated leaflets were collected from plants and bioassayed with 5-day-old larvae. The combination of S. carpocapsae at 0.6 nematodes/cm² + SeMNPV at 40 PIB/cm² produced significantly higher larval mortality (61.7%) compared with either S. carpocapsae (24.8-35.1%) or SeMNPV (26.5-33.7%) alone. In 1992, similar tests were repeated using S. carpocapsae at 0.2 and 0.5 nematodes/cm², and SeMNPV at 14 and 35 PIB/cm². The combination of 0.5 nematodes/cm² + 35 PIB/cm² resulted in significantly higher larval mortality (64.0%) than either pathogen alone (41.5-49.0%). Steinernema carpocapsae and SeMNPV produced additive effects on beet arlnyworm mortality. Persistence of S. carpocapsae was 12-24 hours and SeMNPV was 96-120 hours on soybean.     

Interactions between an ectoparasitoid and a nucleopolyhedrovirus when simultaneously attacking Spodoptera exigua (Lepidoptera: Noctuidae)

Insect pathogenic viruses and parasitoids represent distinct biological entities that exploit a shared host resource and have similar effects in suppressing host populations. This study explores the interactions between the ectoparasitoid Euplectrus plathypenae (Hymenoptera: Eulophidae) and the Spodoptera exigua multiple nucleopolyhedrovirus (SeMNPV) in larvae of S. exigua (Lepidoptera: Noctuidae). Parasitoid progeny failed to complete development in hosts that had been infected prior to parasitism. However, infection of S. exigua fourth instars at 48 h post‐parasitism had no significant effects on the survival of parasitoid progeny. Larval and pupal development times of E. plathypenae that survived on virus‐infected S. exigua did not differ significantly from that of parasitoids on healthy hosts. Virus‐induced mortality and the production of occlusion bodies were very similar in parasitized and non‐parasitized S. exigua. The virus was genetically stable over three passages in parasitized and unparasitized hosts. These results suggest that applications of SeMNPV‐based insecticides are unlikely to disrupt pest control exerted by the parasitoid E. plathypenae in biological pest control programs as long as virus applications are timed not to coincide with parasitoid releases.     

Biological activity of SeMNPV, AcMNPV, and three AcMNPV deletion mutants against Spodoptera exigua larvae (Lepidoptera: noctuidae)

Virulence and speed of action, as related to dose, are important effectiveness-determining properties of insect-pathogenic biocontrol agents. We used the droplet-feeding bioassay to compare dose responses between two wild-type baculoviruses, Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV) and Spodoptera exigua MNPV (SeMNPV), and three deletion mutants of AcMNPV in S. exigua larvae. In each mutant one gene was deleted by genetic engineering: pp34, coding for the polyhedral membrane; egt, coding for ecdysteroid UDP-glucosyltransferase; or p10, coding for fibrillar structures in infected insect cells. SeMNPV had the lowest median lethal dose (LD(50)) as well as the highest speed of action (LT(50)) of all viruses investigated. In our comparative bioassays the only significant effect of gene deletions in AcMNPV was a slightly lower speed of action for the p10 deletion mutant. Otherwise, wild-type and recombinant AcMNPVs had similar biological activities. Our results suggest, in contrast to what is generally assumed, that gene deletions in AcMNPV for improved insecticidal activity should be critically assessed in each host system prior to further implementation as a control agent. Insertion of foreign genes coding for entomotoxins is less questionable and more promising in this respect.     

Efficacy of Spodoptera exigua multiple nucleopolyhedrovirus as a biological insecticide for beet armyworm control in greenhouses of southern Spain

Chemical control measures targeted at Spodoptera exigua in greenhouse sweet pepper crops in Spain have resulted in pest resistance to virtually all commercially available insecticidal products. A multicapsid nucleopolyhedrovirus (SeMNPV), isolated from diseased S. exigua in Spain, was produced in laboratory reared larvae, tested for insecticidal activity in a laboratory bioassay, and was then applied in eleven commercial greenhouses planted with sweet pepper. Virus occlusion bodies (OBs) were applied on two occasions, at an interval of ～7 days, at a rate of 5×10⁸ OBs/L of spray in a volume of ～600 L/ha, depending on crop phenology and greenhouse area. The percentage of plants showing recent (<48 h old) feeding damage fell dramatically in greenhouses with high infestations of S. exigua; the same pattern was observed, although less dramatically, in greenhouses with low infestations. Average mortality of larvae collected from treated plants at 4 days after each application, and reared in the laboratory until death, was high (70–89%) and was not significantly affected by the degree of crop infestation. In a separate trial, the rate of acquisition of infection was examined in larvae that fed on plants treated with 1×10⁸ or 5×10⁸ OBs/L of spray. Of the 27 and 60% of larvae, respectively, that acquired infection in the 48 h period after spraying, about half became infected in the first 6 h post-application, irrespective of application rate. Acquisition of infection proceeded more slowly during the night-time compared to the daytime period, underlining the advantages of early morning applications of the virus. We conclude that the Spanish SeMNPV isolate merits registration as a biological insecticide for use in greenhouse crops in this region.     

Baculovirus‐induced tree‐top disease: how extended is the role of egt as a gene for the extended phenotype?

Many parasites alter host behaviour to enhance their chance of transmission. Recently, the ecdysteroid UDP‐glucosyl transferase (egt) gene from the baculovirus Lymantria dispar multiple nucleopolyhedrovirus (LdMNPV) was identified to induce tree‐top disease in L. dispar larvae. Infected gypsy moth larvae died at elevated positions (hence the term tree‐top disease), which is thought to promote dissemination of the virus to lower foliage. It is, however, unknown whether egt has a conserved role among baculoviruses in inducing tree‐top disease. Here, we studied tree‐top disease induced by the baculovirus Autographa californica multiple nucleopolyhedrovirus (AcMNPV) in two different host insects, Trichoplusia ni and Spodoptera exigua, and we investigated the role of the viral egt gene therein. AcMNPV induced tree‐top disease in both T. ni and S. exigua larvae, although in S. exigua a moulting‐dependent effect was seen. Those S. exigua larvae undergoing a larval moult during the infection process died at elevated positions, while larvae that did not moult after infection died at low positions. For both T. ni and S. exigua, infection with a mutant AcMNPV lacking egt did not change the position where the larvae died. We conclude that egt has no highly conserved role in inducing tree‐top disease in lepidopteran larvae. The conclusion that egt is a ‘gene for an extended phenotype’ is therefore not generally applicable for all baculovirus–host interactions. We hypothesize that in some baculovirus–host systems (including LdMNPV in L. dispar), an effect of egt on tree‐top disease can be observed through indirect effects of egt on moulting‐related climbing behaviour.     

NeuroBactrus,a Novel,Highly Effective,and Environmentally Friendly Recombinant Baculovirus Insecticide

A novel recombinant baculovirus, NeuroBactrus, was constructed to develop an improved baculovirus insecticide with additional beneficial properties, such as a higher insecticidal activity and improved recovery, compared to wild-type baculovirus. For the construction of NeuroBactrus, the Bacillus thuringiensis crystal protein gene (here termed cry1-5) was introduced into the Autographa californica nucleopolyhedrovirus (AcMNPV) genome by fusion of the polyhedrin–cry1-5–polyhedrin genes under the control of the polyhedrin promoter. In the opposite direction, an insect-specific neurotoxin gene, AaIT, from Androctonus australis was introduced under the control of an early promoter from Cotesia plutellae bracovirus by fusion of a partial fragment of orf603. The polyhedrin–Cry1-5–polyhedrin fusion protein expressed by the NeuroBactrus was not only occluded into the polyhedra, but it was also activated by treatment with trypsin, resulting in an ∼65-kDa active toxin. In addition, quantitative PCR revealed that the neurotoxin was expressed from the early phase of infection. NeuroBactrus showed a high level of insecticidal activity against Plutella xylostella larvae and a significant reduction in the median lethal time against Spodoptera exigua larvae compared to those of wild-type AcMNPV. Rerecombinant mutants derived from NeuroBactrus in which AaIT and/or cry1-5 were deleted were generated by serial passages in vitro. Expression of the foreign proteins (B. thuringiensis toxin and AaIT) was continuously reduced during the serial passage of the NeuroBactrus. Moreover, polyhedra collected from S. exigua larvae infected with the serially passaged NeuroBactrus showed insecticidal activity similar to that of wild-type AcMNPV. These results suggested that NeuroBactrus could be recovered to wild-type AcMNPV through serial passaging.     

Relative activity of baculoviruses of the diamondback moth, <Emphasis Type="Italic">Plutella xylostella</Emphasis> (L.) (Lepidoptera: Plutellidae)

The relative activities of the granulovirus (PxGV) and the nucleopolyhedrovirus (PxMNPV) of the diamondback moth, Plutella xylostella (L.), against this insect were evaluated in the laboratory. Similar numbers of occlusion bodies (OBs) of PxGV and PxMNPV were required to kill P. xylostella. However, cadavers of insects infected with PxGV contained more OBs per unit weight than did cadavers infected with PxMNPV. Less cadaver material was thus required to kill insects with PxGV. Larvae that were killed by PxGV survived approximately 2 d longer than did those killed by PxMNPV. When P. xylostella larvae were fed both viruses together, mortality was higher than that caused by either virus alone. The virulence of PxMNPV produced in P.␣xylostella did not differ from that of PxMNPV produced in the beet armyworm, Spodoptera exigua (Hübner). S. exigua was less susceptible to PxMNPV than to its homologous nucleopolyhedrovirus, SeMNPV. Use of trade names does not imply endorsements by USDA-ARS of products named nor criticism of similar ones not mentioned.     

Protein Tyrosine Phosphatase-Induced Hyperactivity Is a Conserved Strategy of a Subset of BaculoViruses to Manipulate Lepidopteran Host Behavior

Many parasites manipulate host behavior to increase the probability of transmission. To date, direct evidence for parasitic genes underlying such behavioral manipulations is scarce. Here we show that the baculovirus Autographa californica nuclear polyhedrovirus (AcMNPV) induces hyperactive behavior in Spodoptera exigua larvae at three days after infection. Furthermore, we identify the viral protein tyrosine phosphatase (ptp) gene as a key player in the induction of hyperactivity in larvae, and show that mutating the catalytic site of the encoded phosphatase enzyme prevents this induced behavior. Phylogenetic inference points at a lepidopteran origin of the ptp gene and shows that this gene is well-conserved in a group of related baculoviruses. Our study suggests that ptp-induced behavioral manipulation is an evolutionarily conserved strategy of this group of baculoviruses to enhance virus transmission, and represents an example of the extended phenotype concept. Overall, these data provide a firm base for a deeper understanding of the mechanisms behind baculovirus-induced insect behavior.     

Adaptation of the Se301 insect cell line to suspension culture. Effect of turbulence on growth and on production of nucleopolyhedrovius (SeMNPV)

As chemical pesticides are being banned as control agents for agricultural pests, the use of the highly specific, safe to non-target organisms baculoviruses has been proposed. These viruses can be produced either in vivo or in vitro. In vitro production requires appropriated host insect cell lines with the ability for growing as freely-suspended cells. In this work, the Spodoptera exigua Se301 cell line was used to produce the commercially available S. exigua nucleopolyhedrovirus (SeMNPV) in suspension. Se301 cells showed to be very sensitive to the hydrodynamic shear rates developed in bioreactors. A process of progressive adaptation to freely-suspended cultures using protective additives against shear stress and disaggregant was proposed. The best combinations were polyvinyl alcohol (PVA) or polyvinyl pyrrolidone (PVP) with the disaggregant dextran sulfate (DS). Both static and freely-suspended Se301 cell cultures were successfully infected with the SeMNPV baculovirus. Production of occluded baculovirus (OB) increased with the multiplicity of infection (MOI > 0.1).     

Juvenile hormone analogs greatly increase the production of a nucleopolyhedrovirus

The commercial production of baculovirus insecticides is limited by the need to produce the virus in living insects. The influence of juvenile hormone analogs (JHA) on the growth and survival of Spodoptera exigua larvae placed on treated diet in the fifth instar was examined. Weight increases observed in methoprene- and fenoxycarb-treated larvae were over three-fold greater than that of control insects, whereas other compounds resulted in lower weight gains (pyriproxyfen) or highly variable responses (hydroprene). Approximately 90% and 70% of fenoxycarb and methoprene-treated larvae, respectively, molted to a supernumerary sixth instar and attained a final weight at 8–10 days post-treatment that was approximately double the maximum weight observed in control larvae. Inoculation of fenoxycarb and methoprene-treated sixth instars with a nucleopolyhedrovirus (SeMNPV) resulted in 2.4- or 2.9-fold increases in final weights, compared to control larvae inoculated in the fifth instar. The total yield of SeMNPV occlusion bodies (OBs) per larva was 2.7- and 2.9-fold greater in fenoxycarb- and methoprene-treated larvae, respectively, compared to fifth instar controls. A significant but small increase in the yield of OBs/mg larval weight was observed in fenoxycarb-treated insects but not in the methoprene treatment. The LC₅₀ value of OBs harvested from fenoxycarb-treated insects was slightly higher than that of OBs from control insects, whereas no such difference was observed in OBs from methoprene-treated insects. We conclude that appropriate use of JHA technology is likely to provide considerable benefits for the mass production of baculoviruses.     

ANALYSIS OF HELICOVERPA ARMIGERA SINGLE NUCLEOPOLYHEDROVIRUS IMMEDIATE EARLY 1 (IE‐1) GENE*

Abstract A 6.12 kb Xbal‐H fragment of the Helicoverpa armigem single nucleopolyhedrovirus (HaSNPV) gemone was cloned and the complete sequence of this fragment was sequenced by random sequencing method. Sequence comparison and analysis revealed an ORF13 which was homologous to ie‐1 of Auiographa California nucleopolyhedrovirus (AcMNPV). The homologous encoding gene is ie‐1. The total length of the encoding region of HaSNPV gene was 1986 bp and was predicted to encode 661 amino acid protein(IE‐1) with molecular weight of 76.5 kD. The alingment of putative HaSNPV IE‐1 amino acid sequence with those of other 9 reported baculoviruses IE‐Is showed that the HaSNPV IE‐1 was most closely related to Helicoverpa zea nucleopolyhedrovirus (HzNPV) IE‐1, with 97% amino acid identidy. But it showed a low degree of sequence similarity to those of AcMNPV, Bombyx mori nucleopolyhedrovirus (BmNPV), Choristoneura fumiferana nucleopolyhedrovirus (CfMNPV), Lymantria dispar nucleopolyhedrovirus (LdMNPV), Orgyia pseudotsugata nucleopolyhedrovirus (OpMNPV), Spodoptera exigua nucleopolyhedrovirus (SeMNPV), Plutella xylostella granulovirus(PxGV) and Xestia c‐nigrum granulovirus (XcGV), with 23%, 23%, 23%, 25%, 23%, 14%, 27% and 7% amino acid identity, respectively. A phylogenetic tree of ten baculoviruses IE‐1 was also given.     

Insecticidal activity of the chitinase from the Spodoptera litura nucleopolyhedrovirus

Baculovirus chitinase gene (chiA) is a late gene essential for liquefying the host insect at a late stage of infection for its hydrolyzing chitin function. In a previous report, baculovirus ChiA has been shown to offer many interesting new opportunities for pest control. Recently, a putative chiA gene was identified in the Korean isolate of the Spodoptera litura nucleopolyhedorvirus (SpliMNPV‐K1) genome. The open reading frame (ORF) contains 1692 nucelotides and encodes a protein of 563 amino acids with a predicted molecular weight of about 62.6 kDa. To study the insecticidal activity of ChiA from SpliMNPV‐K1, we constructed a recombinant AcMNPV, Ap‐SlChiA, which is designed to express the ChiA under the control of a polyhedrin promoter. Western blot analysis indicated that ChiA was successfully expressed by this recombinant virus. Chitinase assay revealed that the chitobiosidase and endochitinase activity of the recombinant virus was 2.5‐ and 3.9‐flods higher than those of wild‐type AcMNPV, respectively. In addition, the recombinant virus showed higher evident insecticidal activity against 3rd instar larvae of Spodotera exigua than that of the AcMNPV. These results suggest that the chiA gene from SpliMNPV‐K1 could be successfully applied to improve pathogenicity of baculoviruses.