Tobacco Budworm Heliothis Virescens Research Articles

Identification and Functional Characterization of Toxoneuron nigriceps Ovarian Proteins Involved in the Early Suppression of Host Immune Response.

Simple SummaryThe endoparasitoid of the tobacco budworm Heliothis virescens, Toxoneuron nigriceps, has several strategies to survive, including venom and calyx fluid. This latter contains a Polydnavirus and Ovarian Proteins (OPs). They are injected into the host body together with the egg. Although much research has focused on venom protein components, little is known about OPs. OPs can disrupt the cellular immune response of the host, acting on host hemocytes, the immune cells. In this study we investigated the action of HPLC fractions derived from OPs. Two fractions caused multiple and significant changes in hemocytes, including cellular oxidative stress and actin cytoskeleton disruption, which might explain the high incidence of hemocyte death and loss of function. Furthermore, using a transcriptome and proteomic approach, we identify the proteins of the two fractions that may be involved in the observed host hemocyte alterations. Our results will help to better understand the OP components and their involvement in parasitization strategies.The endophagous parasitoid Toxoneuron nigriceps (Viereck) (Hymenoptera, Braconidae) of the larval stages of the tobacco budworm Heliothis virescens (Fabricius) (Lepidoptera, Noctuidae) injects the egg, the venom, the calyx fluid, which includes a Polydnavirus (T. nigriceps BracoVirus: TnBV) and the Ovarian Proteins (OPs) into the host body during oviposition. The host metabolism and immune system are disrupted prematurely shortly after parasitization by the combined action of the TnBV, venom, and OPs. OPs are involved in the early suppression of host immune response, before TnBV infects and expresses its genes in the host tissues. In this work, we evaluated the effect of HPLC fractions deriving from in toto OPs. Two fractions caused a reduction in hemocyte viability and were subsequently tested to detect changes in hemocyte morphology and functionality. The two fractions provoked severe oxidative stress and actin cytoskeleton disruption, which might explain the high rate of hemocyte mortality, loss of hemocyte functioning, and hence the host’s reduced hemocyte encapsulation ability. Moreover, through a transcriptome and proteomic approach we identify the proteins of the two fractions: eight proteins were identified that might be involved in the observed host hemocyte changes. Our findings will contribute to a better understanding of the secreted ovarian components and their role in parasitoid wasp strategy for evading host immune responses.

Role of Ovarian Proteins Secreted by Toxoneuron nigriceps (Viereck) (Hymenoptera, Braconidae) in the Early Suppression of Host Immune Response.

Simple SummaryToxoneuron nigriceps is an endoparasitoid of the tobacco budworm Heliothis virescens. Parasitoid strategies to survive involve different regulating factors that are injected into the host body together with the egg: the venom and the calyx fluid, containing a Polydnavirus (PDV) and Ovarian Proteins (OPs). The combination of these factors increases the success of parasitism. Although many studies have been reported on venom protein components and the knowledge on PDVs is increasing, little is known on OPs. These secretions are able to interfere early with the host cellular immune response, acting specifically on host haemocytes, cells involved in immune response. Our results show that OPs induce several alterations on haemocytes, including cellular oxidative stress condition and modifications of actin cytoskeleton, so inducing both a loss of haemocyte functionality and cell death. Overall, in synergy with PDV and venom, OPs positively contribute to the evasion of the host immune response by T. nigriceps.Toxoneuron nigriceps (Viereck) (Hymenoptera, Braconidae) is an endophagous parasitoid of the larval stages of the tobacco budworm, Heliothis virescens (Fabricius) (Lepidoptera, Noctuidae). During oviposition, T. nigriceps injects into the host body, along with the egg, the venom, the calyx fluid, which contains a Polydnavirus (T. nigriceps BracoVirus: TnBV), and the Ovarian Proteins (OPs). Although viral gene expression in the host reaches detectable levels after a few hours, a precocious disruption of the host metabolism and immune system is observed right after parasitization. This alteration appears to be induced by female secretions including TnBV venom and OPs. OPs, originating from the ovarian calyx cells, are involved in the induction of precocious symptoms in the host immune system alteration. It is known that OPs in braconid and ichneumonid wasps can interfere with the cellular immune response before Polydnavirus infects and expresses its genes in the host tissues. Here we show that T. nigriceps OPs induce several alterations on host haemocytes that trigger cell death. The OP injection induces an extensive oxidative stress and a disorganization of actin cytoskeleton and these alterations can explain the high-level of haemocyte mortality, the loss of haemocyte functionality, and so the reduction in encapsulation ability by the host.

Elucidating Efficacy of Ingested Positively Charged Zein Nanoparticles Against Noctuidae.

A meridic diet overlay bioassay using empty, positively charged zein nanoparticles ((+)ZNP) was performed on soybean looper (Chrysodeixis includens (Walker)), tobacco budworm (Heliothis virescens (F.)), and velvetbean caterpillar (Anticarsia gemmatalis Hübner) (Lepidoptera: Noctuidae). Assessment of effects on mortality and development weights 7 d after ingestion of (+)ZNP were evaluated on larvae of each species. Treatments involved different concentrations, with H. virescens and A. gemmatalis offered 0 and 3,800 ppm (+)ZNP, whereas C. includens colonies were offered 0, 630, 1,260, and 2,520 ppm (+)ZNP. Mortality of A. gemmatalis and C. includens increased after ingestion of the highest (+)ZNP concentrations, while H. virescens neonate mortality was unaffected. Neonate and third-instar weights of A. gemmatalis and C. includens, and neonate H. virescens, decreased with high (+)ZNP concentrations. Following mortality results from A. gemmatalis neonates, a concentration response test was performed using a range of (+)ZNP concentrations. The LC50 for A. gemmatalis was 1,478 ppm. The potential of (+)ZNP as a pest management tactic is discussed.

Response of Bt and Non-Bt Cottons to High Infestations of Bollworm (Helicoverpa zea Boddie) and Tobacco Budworm (Heliothis virescens (F.)) under Sprayed and Unsprayed Conditions

Early-maturing and full-season Bt and non-Bt cottons were exposed to high densities of tobacco budworm (Heliothis virescens (F.)) and bollworm (Helicoverpa zea Boddie) in 0.04 ha field cages during the summers of 2011 and 2012 to measure the possible need for supplemental use of insecticides on Bt cotton. Fruit survival within-season and at-harvest was carefully mapped on individual plants within comparative plots of all cottons untreated and sprayed with lambda-cyhalothin (0.0448 kg a.i./ha) or chlorantraniliprole (0.1009 kg a.i./ha) following insect infestations. Differences in lint yields among cotton maturity groups were not always detected, but early-maturing Bt cottons were among the higher yielding experimental plots for both years. Depending on the insecticide treatment, average harvested fruit ranged from 0.3 to 7.1 open bolls per plant for non-Bt cotton plots, while Bt cotton plots ranged from 1.8 to 7.5 open bolls per plant during the two-year study. Bt cottons generally protected fruit from insect damage and resulted in final yields comparable to those of insecticide sprayed Bt and non-Bt cottons. Unsprayed non-Bt cottons were significantly damaged by insects in these high-infestation environments.

The expression patterns of SNMP1 and SNMP2 underline distinct functions of two CD36-related proteins in the olfactory system of the tobacco budworm Heliothis virescens.

In insects, male and female pheromone signals are detected by olfactory sensory neurons (OSNs) expressing the "sensory neuron membrane protein type 1". SNMP1 is supposed to function as a co-receptor involved in the transfer of pheromones to adjacent pheromone receptors. In the moth Heliothis virescens, we previously found OSNs that project their dendrites into pheromone-responsive trichoid sensilla and are associated with cells containing transcripts for the HvirSNMP1-related protein HvirSNMP2. Like HvirSNMP1, HvirSNMP2 belongs to the CD36-family of two-transmembrane domain receptors and transporters for lipophilic compounds, but its role in the olfactory system is unknown. Here, we generated polyclonal anti-peptide antibodies against HvirSNMP2 as well as HvirSNMP1 and conducted an in-depth immunohistochemical analysis of their subcellular localization in the antenna of both sexes. In line with a function in pheromone detection, HvirSNMP1 was immunodetected in the somata and the dendrites of distinct OSNs in subsets of trichoid sensilla. These trichoid sensilla contained only one α-SNMP1-positive OSN in males and clusters of 2-3 labeled cells in females. In contrast, experiments with α-SNMP2-antibodies revealed a broad labeling of non-neuronal support cells (SCs) that are associated with OSNs in likely all trichoid and basiconic sensilla of the antenna with no differences between sexes. Detailed confocal microscope examinations of olfactory sensilla revealed SNMP2-like immunoreactivity close to the apical membrane of SCs and interestingly inside the sensillum. Together, these findings indicate a potential function of SNMP2 in pheromone- as well as general odorant-responsive sensilla and a role fundamentally different from SNMP1.

Ecdysteroidogenesis in Heliothis virescens (Lepidoptera: Noctuidae): Recombinant Prothoracicotropic Hormone and Brain Extract Show Comparable Effects.

Prothoracicotropic hormone (PTTH) is a neuropeptide that triggers a cascade of events within the prothoracic gland (PG) cells, leading to the activation of all the crucial enzymes involved in ecdysone biosynthesis, the main insect steroid hormone. Studies concerning ecdysteroidogenesis predicted PTTH action using brain extract (BE), consisting in a complex mixture in which some components positively or negatively interfere with PTTH-stimulated ecdysteroidogenesis. Consequently, the integration of these opposing factors in steroidogenic tissues leads to a complex secretory pattern. A recombinant form of prothoracicotropic hormone (rPTTH) from the tobacco budworm Heliothis virescens (F.) (Lepidoptera: Noctuidae) was expressed and purified to perform in vitro tests in a standard and repeatable manner. A characterization of rPTTH primary and secondary structures was performed. The ability of rPTTH and H. virescens BE to stimulate ecdysteroidogenesis was investigated on the third day of fifth larval stage. rPTTH activity was compared with the BE mixture by enzyme immunoassay and western blot, revealing that they equally stimulate the production of significant amount of ecdysone, through a transduction cascade that includes the TOR pathway, by the phosphorylation of 4E binding protein (4E-BP) and S6 kinase (S6K), the main targets of TOR protein. The results of these experiments suggest the importance of obtaining a functional pure hormone to perform further studies, not depending on the crude brain extract, composed by different elements and susceptible to different uncontrollable variables.

HT-SuperSAGE of the gut tissue of a Vip3Aa-resistant Heliothis virescens (Lepidoptera: Noctuidae) strain provides insights into the basis of resistance.

Multitoxin Bt‐crops expressing insecticidal toxins with different modes of action, for example, Cry and Vip, are expected to improve resistance management in target pests. While Cry1A resistance has been relatively well characterized in some insect species, this is not the case for Vip3A, for which no mechanism of resistance has yet been identified. Here we applied HT‐SuperSAGE to analyze the transcriptome of the gut tissue of tobacco budworm Heliothis virescens (F.) laboratory‐selected for Vip3Aa resistance. From a total of 1 324 252 sequence reads, 5 895 126‐bp tags were obtained representing 17 751 nonsingleton unique transcripts (UniTags) from genetically similar Vip3Aa‐resistant (Vip–Sel) and susceptible control (Vip–Unsel) strains. Differential expression was significant (≥2.5 fold or ≤0.4; P < 0.05) for 1989 sequences (11.2% of total UniTags), where 420 represented overexpressed (OE) and 1569 underexpressed (UE) genes in Vip–Sel. BLASTN searches mapped 419 UniTags to H. virescens sequence contigs, of which, 416 (106 OE and 310 UE) were unambiguously annotated to proteins in NCBI nonredundant protein databases. Gene Ontology distributed 345 of annotated UniTags in 14 functional categories with metabolism (including serine‐type hydrolases) and translation/ribosome biogenesis being the most prevalent. A UniTag homologous to a particular member of the REsponse to PAThogen (REPAT) family was found among most overexpressed, while UniTags related to the putative Vip3Aa‐binding ribosomal protein S2 (RpS2) were underexpressed. qRT‐PCR of a subset of UniTags validated the HT‐SuperSAGE data. This study is the first providing lepidopteran gut transcriptome associated with Vip3Aa resistance and a foundation for future attempts to elucidate the resistance mechanism.

Distribution of tachykinin-related peptides in the brain of the tobacco budworm Heliothis virescens.

Invertebrate tachykinin-related peptides (TKRPs) comprise a group of signaling molecules having sequence similarities to mammalian tachykinins. A growing body of evidence has demonstrated the presence of TKRPs in the central nervous system of insects. In this investigation, we used an antiserum against locustatachykinin-II to reveal the distribution pattern of these peptides in the brain of the moth Heliothis virescens. Immunolabeling was found throughout the brain of the heliothine moth. Most of the roughly 500 locustatachykinin-II immunoreactive cell bodies, that is, ca. 400, were located in the protocerebrum, whereas the rest was distributed in the deutocerebrum, tritocerebrum, and the gnathal ganglion. Abundant immunoreactive processes were located in the same regions. Labeled processes in the protocerebrum were especially localized in optic lobe, central body, lateral accessory lobe, superior protocerebrum, and lateral protocerebrum, while those in the deutocerebrum were present exclusively in the antennal lobe. In addition to brain interneurons, four pairs of median neurosecretory cells in the pars intercerebralis with terminal processes in the corpora cardiaca and aorta wall were immunostained. No sexual dimorphism in immunoreactivity was found. Comparing the data obtained here with findings from other insect species reveals considerable differences, suggesting species-specific roles of tachykinin-related peptides in insects.

Dynamics of transcriptomic response to infection by the nematode Heterorhabditis bacteriophora and its bacterial symbiont Photorhabdus temperata in Heliothis virescens larvae.

Entomopathogenic nematodes in the Heterorhabditis genus and their symbiotic Photorhabdus bacteria are important biocontrol agents of insect pests and models for the study of microbe-host interactions. In this work, we used larvae of the tobacco budworm (Heliothis virescens) as a model to study its defensive mechanisms against Heterorhabditis bacteriophora nematodes carrying symbiotic Photorhabdus temperata. We first determined time points of initial nematode entry and release of bacteria into the haemolymph to perform transcriptional analysis of insect gene expression during these steps in the infective process. RNA-Sequencing analyses were then performed to profile differential gene expression in the insect during nematode invasion, bacterial release and final steps of infection, relative to the untreated controls. Our results support the theory that insect immune response genes are induced upon nematode invasion, but themajority of these genes are suppressed upon the release of bacteria by the nematodes into the haemolymph. Overall, these findings provide information onthe dynamics of the insect's response to a progressing infection by this entomopathogenic nematode-bacteria complex and facilitate development of Hel.virescens as a pest model for future functional studies of the key insect defence factors.

Registration of ‘Acala 1517‐16 B2RF’, a New Acala Cotton Cultivar with Insect and Herbicide Resistance

Insect pests and weeds cause significant yield losses in cotton (Gossypium hirsutum L.) production. The New Mexico Agricultural Experiment Station announces the release of an insect‐resistant and glyphosate herbicide‐tolerant Acala cotton cultivar, ‘Acala 1517‐16 B2RF’ (Reg. No. CV‐133, PI 678370), possessing Bollguard II (BGII or B2) and Roundup‐Ready Flex (RF) traits. Cotton with the B2 trait has an excellent season‐long in‐plant protection against pink bollworm [Pectinophora gossypiella (Saund.)], tobacco budworm (Heliothis virescens F.), American bollworm [Helicoverpa armigera (Hubner)], and glyphosate and an improved resistance to beet armyworm [Spodoptera exigua (Hübner)], soybean looper (Pseudoplusia includens Walker), and bollworm (H. zea Boddie). The new cultivar was selected as a progeny row derived from a backcross BC3F3 between a Monsanto B2RF donor and NM 970123 as the recurrent parent. Acala 1517‐16 B2RF was tested in 11 replicated field trials in New Mexico from 2007 to 2009, one field test in the High Plains of Texas, and nine locations outside of the Southwest region in 2009. Acala 1517‐16 B2RF had similar lint yield to ‘Acala 1517‐99W’ containing two Bt genes and two conventional cultivars ‘Acala 1517‐08’ and ‘PHY 72 Acala’. The most improved trait was its greater boll weight. In general, Acala 1517‐16 B2RF had similar fiber quality to the three checks in most of the tests, but its fibers were coarser than Acala 1517‐99W and finer than Acala 1517‐08 and PHY 72 Acala. This insect‐resistant and glyphosate‐tolerant Acala cotton cultivar is best adapted to the U.S. Southwest arid region of the Cotton Belt, where conventional Acala 1517 cultivars are grown.

Optimal Cotton Insecticide Application Termination Timing: A Meta-Analysis.

The concept of insecticide termination timing is generally accepted among cotton (Gossypium hirsutum) researchers; however, exact timings are often disputed. Specifically, there is uncertainty regarding the last economic insecticide application to control fruit-feeding pests including tarnished plant bug (Lygus lineolaris (Palisot de Beauvois)), boll weevil (Anthonomus grandis), bollworm (Helicoverpa zea), tobacco budworm (Heliothis virescens), and cotton fleahopper (Pseudatomoscelis seriatus). A systematic review of prior studies was conducted within a meta-analytic framework. Nine publicly available articles were amalgamated to develop an optimal timing principle. These prior studies reported 53 independent multiple means comparison field experiments for a total of 247 trial observations. Stochastic plateau theory integrated with econometric meta-analysis methodology was applied to the meta-database to determine the shape of the functional form of both the agronomic optimal insecticide termination timing and corresponding yield potential. Results indicated that current university insecticide termination timing recommendations are later than overall estimated timing suggested. The estimated 159 heat units (HU) after the fifth position above white flower (NAWF5) was found to be statistically different than the 194 HU termination used as the status quo recommended termination timing. Insecticides applied after 159 HU may have been applied in excess, resulting in unnecessary economic and environmental costs. Empirical results also suggested that extending the insecticide termination time by one unit resulted in a cotton lint yield increase of 0.27 kilograms per hectare up to the timing where the plateau began. Based on economic analyses, profit-maximizing producers may cease application as soon as 124 HU after NAWF5. These results provided insights useful to improve production systems by applying inputs only when benefits were expected to be in excess of the respective costs.

Potential detoxification of gossypol by UDP-glycosyltransferases in the two Heliothine moth species Helicoverpa armigera and Heliothis virescens

The cotton bollworm Helicoverpa armigera and the tobacco budworm Heliothis virescens are closely related generalist insect herbivores and serious pest species on a number of economically important crop plants including cotton. Even though cotton is well defended by its major defensive compound gossypol, a toxic sesquiterpene dimer, larvae of both species are capable of developing on cotton plants. In spite of severe damage larvae cause on cotton plants, little is known about gossypol detoxification mechanisms in cotton-feeding insects. Here, we detected three monoglycosylated and up to five diglycosylated gossypol isomers in the feces of H. armigera and H. virescens larvae fed on gossypol-supplemented diet. Candidate UDP-glycosyltransferase (UGT) genes of H. armigera were selected by microarray studies and in silico analyses and were functionally expressed in insect cells. In enzymatic assays, we show that UGT41B3 and UGT40D1 are capable of glycosylating gossypol mainly to the diglycosylated gossypol isomer 5 that is characteristic for H. armigera and is absent in H. virescens feces. In conclusion, our results demonstrate that gossypol is partially metabolized by UGTs via glycosylation, which might be a crucial step in gossypol detoxification in generalist herbivores utilizing cotton as host plant.

Moth pheromone receptors: gene sequences, function, and evolution

The detection of female-released species-specific sex pheromones in moths is mediated by the pheromone receptors that are expressed in the sensory neurons in the olfactory sensilla of conspecific male antennae. Since the pioneering studies on the tobacco budworm Heliothis virescens and the silkworm Bombyx mori a decade ago, genes encoding pheromone receptors have been identified from a number of moth species. Pheromone receptor genes constitute a specialized olfactory receptor subfamily that shares sequence homology. In most cases the pheromone receptor genes are more abundantly expressed in male antennae, and the expression is confined to the neurons in the long sensilla trichodea, which are responsible for pheromone sensing. Both highly specific and more broadly tuned pheromone receptors have been described in various moth species. We review the advances in moth pheromone receptor studies over the past decade, including the methods used in receptor gene isolation and functional characterization, the different ligand profiles of the identified receptors, and the evolution of this multigene family.

Functional analysis of an immune gene of Spodoptera littoralis by RNAi

Insect immune defences rely on cellular and humoral responses targeting both microbial pathogens and metazoan parasites. Accumulating evidence indicates functional cross-talk between these two branches of insect immunity, but the underlying molecular mechanisms are still largely unknown. We recently described, in the tobacco budworm Heliothis virescens, the presence of amyloid fibers associated with melanogenesis in immune capsules formed by hemocytes, and identified a protein (P102) involved in their assembly. Non-self objects coated by antibodies directed against this protein escaped hemocyte encapsulation, suggesting that P102 might coordinate humoral and cellular defence responses at the surface of foreign invaders. Here we report the identification of a cDNA coding for a protein highly similar to P102 in a related Lepidoptera species, Spodoptera littoralis. Its transcript was abundant in the hemocytes and the protein accumulated in large cytoplasmic compartments, closely resembling the localization pattern of P102 in H. virescens. RNAi-mediated gene silencing provided direct evidence for the role played by this protein in the immune response. Oral delivery of dsRNA molecules directed against the gene strongly suppressed the encapsulation and melanization response, while hemocoelic injections did not result in evident phenotypic alterations. Shortly after their administration, dsRNA molecules were found in midgut cells, en route to the hemocytes where the target gene was significantly down-regulated. Taken together, our data demonstrate that P102 is a functionally conserved protein with a key role in insect immunity. Moreover, the ability to target this gene by dsRNA oral delivery may be exploited to develop novel technologies of pest control, based on immunosuppression as a strategy for enhancing the impact of natural antagonists.

Evaluation of biweekly pesticide applications of new insecticides for tobacco budworm (Heliothis virescens (Fabricius)) management in tobacco (Nicotiana tabacum L.) seed production

Field studies were conducted in 2010 and 2011 in North Carolina tobacco grown to produce seed to assess the longevity and efficacy of the insecticide active ingredients chlorantraniliprole and flubendiamide on Heliothis virescens (Fabricius), the most significant pest of tobacco grown for seed. Results suggest that incorporating these materials into an integrated pest management program for lepidopterous pests in seed tobacco could potentially decrease the number of insecticide applications with limited impact on natural enemies. Soil applications of chlorantraniliprole may potentially delay H. virescens infestations, further decreasing foliar insecticide applications, but activity of soil applications was not consistent between sites and years.

Evaluating host plant resistance in cotton (&amp;lt;i&amp;gt;Gossypium hirsutum&amp;lt;/i&amp;gt; L.) with varying gland densities to tobacco budworm (&amp;lt;i&amp;gt;Heliothis virescens&amp;lt;/i&amp;gt; F.) and bollworm (&amp;lt;i&amp;gt;Helicoverpa zea&amp;lt;/i&amp;gt; Boddie) in the field and laboratory

Cotton (Gossypium hirsutum L.) produces a number of toxic terpenoid aldehyde (TA) compounds contained in epidermal glands that help protect the plant from pests and diseases. In the seed, one of these toxic compounds, gossypol, limits the use of the seed to ruminants such as dairy cows. There are breeding techniques and germplasm available to decrease gossypol in the seed, but the breeding process also needs to include methods to evaluate the plant’s ability to resist insect pests. Three approaches were used to assess resistance of cotton to herbivory from bollworm (Helicoverpa zea Boddie) and tobacco budworm (Heliothis virescens F.) including field counts, controlled field antibiosis assays and laboratory feeding tests of young field grown leaves. Results indicated that both field and laboratory evaluation could provide an assessment of the cotton host’s resistance. Measurements of terpenoid aldehydes (TAs) in the seed and the leaves, confirmed that the levels and types of TAs in the seed were not always good estimators of leaf TAs and that other TAs such as hemigossypolone and heliocides contribute to host plant resistance.

Pinellia ternata agglutinin expression in chloroplasts confers broad spectrum resistance against aphid, whitefly, Lepidopteran insects, bacterial and viral pathogens

Broad spectrum protection against different insects and pathogens requires multigene engineering. However, such broad spectrum protection against biotic stress is provided by a single protein in some medicinal plants. Therefore, tobacco chloroplasts were transformed with the agglutinin gene from Pinellia ternata (pta), a widely cultivated Chinese medicinal herb. Pinellia ternata agglutinin (PTA) was expressed up to 9.2% of total soluble protein in mature leaves. Purified PTA showed similar hemagglutination activity as snowdrop lectin. Artificial diet with purified PTA from transplastomic plants showed marked and broad insecticidal activity. In planta bioassays conducted with T0 or T1 generation PTA lines showed that the growth of aphid Myzus persicae (Sulzer) was reduced by 89%-92% when compared with untransformed (UT) plants. Similarly, the larval survival and total population of whitefly (Bemisia tabaci) on transplastomic lines were reduced by 91%-93% when compared with UT plants. This is indeed the first report of lectin controlling whitefly infestation. When transplastomic PTA leaves were fed to corn earworm (Helicoverpa zea), tobacco budworm (Heliothis virescens) or the beet armyworm (spodoptera exigua), 100% mortality was observed against all these three insects. In planta bioassays revealed Erwinia population to be 10,000-fold higher in control than in PTA lines. Similar results were observed with tobacco mosaic virus (TMV) challenge. Therefore, broad spectrum resistance to homopteran (sap-sucking), Lepidopteran insects as well as anti-bacterial or anti-viral activity observed in PTA lines provides a new option to engineer protection against biotic stress by hyper-expression of an unique protein that is naturally present in a medicinal plant.

Hvmar1, a mariner‐like element from the tobacco budworm Heliothis virescens, can transpose in Drosophila melanogaster

AbstractTransposable elements of the mariner family are widespread and have been found in the genome of plants, animals and insects. However, most of these elements contain multiple inactivating mutations and so far, only three naturally occurring mariner elements are known to be functional. In a previous study, a mariner‐like element called Hvmar1 was discovered in the genome of the tobacco budworm Heliothis virescens. Further analysis of the Hvmar1 nucleotide sequence revealed the presence of 30‐bp imperfect inverted terminal repeats and an intact open reading frame, which is considered to encode a functional transposase. In the present study, we show that the Hvmar1 element is active using interplasmid transposition assays in Drosophila melanogaster embryos. When injected into Drosophila embryos, the helper plasmid produced a transposase that was able to mediate transposition of the Hvmar1 element from a donor to a target plasmid. The transposition efficiency of Hvmar1 in D. melanogaster is approximately 11‐fold lower than that of the well‐known Mos1 mariner transposon. However, this efficiency is comparable to those observed previously with Mos1 in non‐Drosophila insects. We identified 10 independent interplasmid transposition events, albeit the recovery of these events was rare. In each case the Hvmar1 element transposed in a precise manner, with the characteristic TA dinucleotides being duplicated on insertion. Furthermore, two of the target sites identified have been used previously by Mos1 for insertion. The active transposition of Hvmar1 in D. melanogaster provides a basis for examining the mobility of this element in its natural host as well as a starting point for comparative studies with Mos1 and other functional mariner transposons.

Long-term Benefits of GM crops: Potential for Diabrotica Suppression in Europe using Bt Maize

AbstractTransgenic crops producing insecticidal proteins from Bacillus thuringiensis (Bt) have been widely adopted since 1996 in the United States of America to combat important pests of maize and cotton. There is growing evidence that several target pest populations have been dramatically reduced in areas where the Bt crops have been most intensively adopted over multiple years. The evidence is most dramatic for nonmigratory monophagous and oligophagous species that show high mortality on Bt crops, such as European corn borer (Ostrinia nubilalis), and tobacco budworm (Heliothis virescens). Bt cotton is currently being used in the southwestern USA as part of an area-wide eradication program for pink bollworm (Pectinophora gossypiella). Single, stacked, and pyramided Bt maize lines producing coleopteran-active insecticidal proteins have been shown in some cases to cause &gt;99% mortality of western corn rootworm (Diabrotica virgifera virgifera) larvae every year. Simulation models suggest that long-term area-wide cultivation of these Bt corn lines can lead to dramatic population reduction, and even local extinction, of corn rootworms. An eradication program for this economically important pest, based around Bt maize and incorporating a combination of other tools, would benefit European agriculture and the environment. Post-commercial monitoring programs should be designed to investigate long-term benefits to agricultural production systems that can be reasonably expected from the widespread cultivation of Bt crops in Europe.

Soybean Flour and Wheat Germ Proportions in Artificial Diet and Their Effect on the Growth Rates of the Tobacco Budworm,Heliothis virescens

Soybean flour and wheat germ are the two most important protein components of wheat germ-based insect artificial diets. The effect of modifying the proportion of these two ingredients in a Noctuidae-specific diet was investigated utilizing the tobacco budworm Heliothis virescens (F.) (Lepidoptera: Noctuidae), with the goal of developing a suboptimal diet that, without drastically affecting this insect's growth and reproductive rates, could manifest subtle negative effects in this insect. The original diet formula contained 2.51% protein. When the proportions of soybean flour and wheat germ were changed to 2.15% protein the net reproductive rate of the first generation was significantly lower. In the second generation, the net reproductive rate, development time, percent female survivorship, fertility, intrinsic rate of increase, finite rate of increase and female longevity were significantly lower in both the 2.15% and 2.26% protein diets. The survival rate of immatures to the adult stage was 1% in the 2.05% protein diet in the first generation. Interestingly, females exposed to these suboptimal diets produced a significantly higher number of eggs but the survival of their larvae was significantly reduced. It is evident from these results that modifications to the protein content and the nutrient composition profile of the original wheat germ-based insect artificial formula can be used to produce subtle negative effects on the growth of tobacco budworm.