Impact of native predators and parasites on Spodoptera exigua, an introduced pest of alfalfa hay in northern California

A long-term study of Spodoptera exigua (Hübner) (Lepidoptera: Noctuidae) revealed that a complex of native predators and parasites exploit this introduced pest in alfalfa-hay fields in northern California. In field trials, predators typically fed on >50% of sentinel egg masses during a given sample interval; mean predation rate ranged from 40 to 100% and percentage predation per individual egg mass was density-independent (spatial context) for 16 of 17 sample intervals. The egg-predator guild consisted primarily of adults and nymphs of Lygus hesperus Knight & L. elisus Van Duzee (Miridae), Nabis americoferus Carayon (Nabidae), and Orius tristicolor (White) (Anthocoridae); and adults of Collops vittatus (Say) (Melyridae) and Hippodamia convergens Guerin-Meneville (Coccinellidae). In laboratory feeding trials, these species readily fed on eggs, as well as neonate larvae of S. exigua. The parasite guild associated with small- to medium-sized larvae consisted of Hyposoter exiguae (Viereck) and Pristomerus spinator (F.) (Ichneumonidae), both larval endoparasites; and Chelonus insularis Cresson (Braconidae), an egg-larval endoparasite. Total parasitization by these species in field samples ranged from 5 to 75%, and was density-independent (spatial context) for each of four sample dates. A few larvae were infected with a nuclear polyhedrosis virus but no epizootics were observed. These results are consistent with the hypothesis that native natural enemies, especially generalist predators, can maintain populations of S. exigua at levels well below the treatment threshold for this exotic pest of alfalfa grown for hay.     

An evaluation of some natural enemies of >Nezara viridula in northern California

Natural enemies associated with eggs of >Nezara viridula (L.) (Heteroptera: Pentatomidae) wereevaluated by placing sentinel egg masses inweeds and cultivated tomato and bean crops innorthern California. Egg predation wasgenerally less than 10% and normally involvedpredators with chewing mouth parts. Predatorsseldom destroyed an entire egg mass, typicallyeating <40 eggs per exploited mass.Laboratory evaluation of >25 species ofpotential arthropod predators revealed that fewfed on >N. viridula eggs to any extent;however, numerous species fed on >N.viridula nymphs. Five species of eggparasites were recovered from sentinel eggmasses: >Trissolcus basalis (Wollaston),>Gryon obesum Masner and >Telenomuspodisi Ashmead (Scelionidae); and>Ooencyrtus californicus Girault and >O.johnsoni (Howard) (Encyrtidae). The major eggparasite was >T. basalis, the only exoticmember of the parasite guild; it typicallyparasitized 100% of the eggs in an exploitedegg mass. The results indicate thatparasitization of eggs and predation of smallnymphs can be important biotic mortalityfactors for >N. viridula populations innorthern California. It is suggested that acombination of factors – viz., eggparasitization, nymphal predation, regionalshortage of overwintering sites, and localshortages of suitable hosts – maintains thisexotic pest at relatively low levels in theregion.     

Effect of CryIC toxin from Bacillus thuringiensis on larval feeding behavior of Spodoptera exigua

The lack of data on the effect of Bacillus thuringiensis (B.t.) toxins on larval feeding behavior of the pest Spodoptera exigua (Hübner) (Noctuidae: Amphypyrini) prompted us to investigate the effect of three delivery systems of CryIC, a commercial formulation, inclusion bodies, and the activated CryIC toxin. The commercial formulation was the least and CryIC toxin the most lethal form to neonates of susceptible colonies. All but two of the treatments in choice tests with neonates and third instars showed significant avoidance of B.t. treated diet, with greater proportion of larvae from susceptible (UCR-S and AUBURN-S) and resistant (AUBURN-R) colonies on untreated diet than on diet treated with any of the CryIC forms and concentrations tested. Furthermore, third instars consumed significantly more control than treated diet for all CryIC forms, colonies and concentrations. The avoidance of CryIC toxin by neonates and third instars strongly suggests that CryIC, which also is present in the commercial formulation and in the inclusion bodies, is responsible for eliciting avoidance behavior by S. exigua larvae. Behavioral observations of third instars in a no-choice test on either treated or control diet indicated that questing behavior in susceptible larvae appears to be positively related with presence of CryIC toxin in the diet. Furthermore, resistant third instars were on the whole more active than susceptible thirds on both treated and control diet. Resistant thirds raised on CryIC treated diet (AUBURN-RC) spent more time eating treated diet than resistant larvae raised on control diet (AUBURN-R), suggesting that diet conditioning plays an important role on feeding behavior of S. exigua. The implications of these results are discussed.     

Nuclear polyhedrosis virus for control of Spodoptera exigua larvae on glasshouse crops

the efficacy of a nuclear polyhedrosis virus, SeNPV, for controlling larval populations of the beet armyworm, Spodoptera exigua (Noctuidae) was studied in chrysanthemum, gerbera, kalanchoë and tomato crops in glasshouses. Applications of 1×10⁸ PIBs/m² on the various crops resulted in 95–100% larval mortality. Virus applications caused comparable levels of mortality in populations of early and late instar larvae. Reduction in feeding damage to the crops, however, was more pronounced when the larvae were early instars at the time of virus application. Single applications with the virus resulted in higher levels of larval mortality as double applications with 0.1% methomyl and diflubenzuron.The virus appears to be a potential control agent for the beet armyworm in Dutch glasshouses.

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Résumé Les résultats d'applications d'un virus de la polyédrose nucléaire contre des populations de chenilles de Spodoptera exigua sont examinés dans des chrysanthèmes, gerberas, kalanchoës et tomates. Une dose de 1×10⁸ PIBs/m² résulte à 95–100% mortalité larvale dans tous les végétaux. L'effet sur chemilles de première et seconde stade larvale et sur chenilles de troisième et quatrième stade larvale était pareille. Malgré le niveau élevé du mortalité larvale les végéteaux étaient damagés sérieusement, particulièrement quand le virus était appliqué contre des chenilles de troisième et quatrième stade larvale.Comparée avec des pesticides chimiques, méthomyl et diflubenzuron, applications de virus résultent à un mortalité larvale plus haute. Le conclusion était que le virus est un candidat sérieus pour l'utilisation dans le lutte biologique contre les chenilles de S. exigua dans des serres en Pays Bas.

     

Phenotypic characteristics and relative proportions of three genotypic variants isolated from a nucleopolyhedrovirus of Spodoptera exigua

US2A, US2D, and US2F are three in vivocloned genotypic variants from the wild-type strain of a Spodoptera exiguanucleopolyhedrovirus (SeMNPV) isolated in Florida (USA) and is the active component of the commercial bioinsecticide Spod-X^®. These variants were compared in terms of pathogenicity (LD₅₀), speed of kill (expressed as mean time to death) and viral progeny productivity (OBs/larva). LD₅₀values were similar for the three cloned genotypes. The mean time to death value for US2D (113.7 h) was significantly higher than those of US2A (31.7 h) and US2F (27.8 h). Virus yield was determined for L₄larvae infected with the estimated LD₉₉. US2D infected larvae attained higher weight than those infected with US2A and US2F, and produced a higher OB yield than larvae infected with US2A or US2F. An outstanding feature of US2F, in contrast to US2A and US2D, was its inability to disrupt the teguments of NPV-killed larvae. To study the relative proportion of the three genotypic variants throughout successive passages, S. exigualarvae were originally infected with a viral inoculum containing a 1:1:1 mixture of the three genotypes. After three successive passages, US2D was no longer detected in either of the three replicate experiments performed, while US2A was the predominant genotype in all of them, and US2F remained at similar proportions throughout the three passages. The influence of the phenotypic characteristics of the three variants on their relative proportions in mixed infections is discussed.     

RNA interference of an antimicrobial peptide, gloverin, of the beet armyworm, Spodoptera exigua, enhances susceptibility to Bacillus thuringiensis

Gloverin is known to be an inducible antimicrobial peptide. This study reports a gloverin gene (Seglv) identified from the beet armyworm, Spodoptera exigua. Seglv encodes 175 amino acids with a signal peptide. Its amino acid sequence is highly homologous (>95%) to other known gloverins. Seglv was expressed from egg to adult stages even without immune challenge. Especially, in larval stage, it was expressed in all tested tissues, such as hemocyte, fat body, gut, and epidermis. However, the constitutive expression level was significantly elevated in response to bacterial challenge. Expression of a Toll gene was required for expression of Seglv. A recombinant Seglv protein was synthesized using a bacterial expression system and purified with an affinity chromatography. The recombinant protein showed a specific antibacterial activity against a Gram-positive bacterium, but no activity against a Gram-negative Escherichia coli. Injection of specific double stranded RNA (dsRNA) against Seglv could suppress its expression. Knockdown of Seglv expression induced a significant developmental retardation and resulted in hypotrophy pupae. The larvae treated with dsRNA were much more susceptible to Bacillus thuringiensis than the control larvae. These results suggest that Seglv acts as an antimicrobial peptide especially against Gram-positive bacteria including B. thuringiensis.     

Effects of plant chemical extracts and physical characteristics of Apium graveolens and Chenopodium murale on host choice by Spodoptera exigua larvae

Choice tests with whole plants and leaf discs indicated that fourth instar Spodoptera exigua (Hübner) (Noctuidae: Amphypyrini) were found more frequently and ate significantly more of the weed Chenopodium murale than the associated crop plant Apium graveolens. In order to explain the preference, plant extracts, plant volatiles, soluble protein concentrations, water contents, and leaf toughness of the two plants were investigated. Bioassays of aqueous methanol (90%) and hexane extracts of leaves on cellulose discs indicated that neither attractants in C. murale nor repellents in A. graveolens could account for the observed preference. No significant difference could be found between the effects of plant volatiles from C. murale, A. graveolens and a control on larval dispersal by S. exigua. Selective feeding for higher levels of proteins also was not a factor, because A. graveolens had nearly twice the soluble protein of C. murale. Water content was approximately 6% higher (by weight) in C. murale than A. graveolens but most polyphagous larvae do not typically show compensatory feeding for water alone. However, the potentially related characteristic of leaf toughness was significantly different, with A. graveolens exhibiting 1.53 times the toughness of C. murale. Studies comparing five types of larval behavior on both plant species showed that the time spent in swallowing behavior was significantly greater on the tougher A. graveolens leaves relative to C. murale. To test the hypothesis that leaf toughness was affecting larval host choice, both plants were finely ground and incorporated into agar blocks. No differences in feeding behavior were detected. The implications of leaf toughness for larval diet and host choice are discussed.     

Adaptation of Spodoptera exigua (Lepidoptera: Noctuidae) to barley trypsin inhibitor BTI-CMe expressed in transgenic tobacco

Nicotiana tabacum plants were transformed with the cDNA of barley trypsin inhibitor BTI-CMe under the control of the 35S CaMV promoter. Although the transgene was expressed and the protein was active in the homozygous lines selected, growth of Spodoptera exigua (Lepidoptera: Noctuidae) larvae reared on transgenic plants was not affected. The protease activity in larval midgut extracts after 2 days feeding on transformed tobacco leaves from the highest expressing plant showed a reduction of 25% in the trypsin-like activity compared to that from insects fed on non-transformed controls. The susceptibility of digestive serine-proteases to inhibition by BTI-CMe was confirmed by activity staining gels. This decrease was compensated with a significant induction of leucine aminopeptidase-like and carboxipeptidase A-like activities, whilechymotrypsin-, elastase-, and carboxipeptidase B-like proteases were not affected.     

Feeding and dispersion of Spodoptera exigua larvae and its relevance for control with a nuclear polyhedrosis virus

On chrysanthemum crops the larvae of the beet armyworm Spodoptera exigua (Hübner) (Lep.: Noctuidae) moved upwards to the top of plants after hatching and predominantly fed on the upper foliage layers. On tomato, however, the larvae did not move upwards and mainly fed on the lower leaves. On chrysanthemums up to the fourth instar most feeding occurred at the underside of foliage while the upper-epidermis remained intact. Larvae dispersing from a single egg batch of 35 eggs damaged about 90 small and 50 tall chrysanthemum plants during their development. The successive larval stages contributed respectively 0.1, 0.4, 4, 20 and 75 percent to the total foliage consumption.The results indicated that the virus preferably should be applied to the lower leaves of tomatoes and chrysanthemums when young instars are present, but to the upper-middle and top leaves of chrysanthemums when the larvae are older than second instars.

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Résumé La consommation et la dispersion des chenilles de Spodoptera exigua sont examinées sur chrysanthèmes et tomates de serre. Les adults déposent les oeufs généralement au feuilles près du sol. Apres l'éclosion les chenilles graduellement se mouvent en haute dans les chrysanthèmes et consomment principalement les feuilles les plus hautes. Dans tomate, cependant, les chenilles s'alimentent principalement au feuilles près du sol.Pendant leur dévelopment les chenilles originaires d'une seule pond de 35 oeufs peuvent endommager environ go petits ou so hautes crysanthè mes. Jusqu'à la quatrième stade larvale les chenilles s'alimentent principalement avec le surface inférieure des feuilles, sans consommer l'épiderme supérieure.Les cinq stades larvales contribuent respectivement 0,1, 0,4, 4, 20 et 75% a la consommation totale des feuilles de chrysanthème.Les resultats des expériments sur la conduite larvale suggestent que les viruses de la polyédrose nucléaire doivent être appliqués préférablement sur la face inférieure des feuilles bas de chrysanthème et de tomate avant que les chenilles se développent a la troisième stade larvale. Quand les larves sur chrysanthèmes sont déja dans le troisième stade larvale, application du virus sur les feuilles plus haute probablement donnait des resultats optimal.

     

斜纹夜蛾天敌作用的评价

通过组建斜纹夜蛾第4代和第8代自然种群生命表,运用排除作用控制指数分析了生物因子对斜纹夜蛾种群的自然控制作用。结果表明,低龄(1～3龄)幼虫的捕食性天敌是影响斜纹夜蛾种群数量动态的重要因子。对第4代和第8代种群的排除作用控制指数分别为13.904和12.946.如果没有捕食性天敌的作用,下代种群数量将分别增长到当代的15.1206和74.678倍。病原微生物是影响第4代斜纹夜蛾种群数量的另一重要因子,其排除作用控制指数为2.4726.     

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.     

斜纹夜蛾天敌作用的评价

通过组建斜纹夜蛾第４代和第８代自然种群生命表,运用排除作用控制指数分析了生物因子对斜纹夜蛾种群的自然控制作用．结果表明,低龄（１～３龄）幼虫的捕食性天敌是影响斜纹夜蛾种群数量动态的重要因子．对第４代和第８代种群的排除作用控制指数分别为１３．９０４和１２．９４６．如果没有捕食性天敌的作用,下代种群数量将分别增长到当代的１５．１２０６和７４．６７８倍．病原微生物是影响第４代斜纹夜蛾种群数量的另一重要因子,其排除作用控制指数为２．４７２６．     

Effects of individual Bacillus thuringiensis insecticidal crystal proteins on adult Heliothis virescens (F.) and Spodoptera exigua (Hubner) (Lepidoptera: Noctuidae)

Bacillus thuringiensis (Bt) is a grampositive, spore forming bacterium, which is principally distinguishedfrom other bacilli by the production of large, insecticidal,protein crystals (Insecticidal Crystal Proteins, or ICPs). Theseproteins are usually thought to act only on the actively feedinglarvae of susceptible species by a mechanism which involvesconsumption and proteolytic processing of the protein followed bybinding to, and lysis of, midgut epithelial cells. However, few authorshave reported Bt toxicity to adult insects. In the followingpaper, we expand on previous reports of toxicity to adult insects andpresent data which demonstrate that: (1) proteolytically activated ICPssignificantly reduce the lifespans of adult Heliothis virescensand Spodoptera exigua at concentrations of 500 g/ml, butnot 167 or 25 g/ml, (2) individual activated ICPs are differentiallytoxic to adult H. virescens and S. exigua, and (3)adult S. exigua are sensitive to Cry1C protoxin at aconcentration of 1 mg/ml.Deceased     

An entomopathogenic bacterium, Xenorhabdus nematophila, inhibits hemocyte phagocytosis of Spodoptera exigua by inhibiting phospholipase A(2)

Phagocytosis is a hemocytic behavior against bacterial infection. An entomopathogenic bacterium, Xenorhabdus nematophila, inhibits immune responses of target insects and causes hemolymph septicemia. This study analyzed how X. nematophila could inhibit phagocytosis to increase its pathogenicity. Granular cells and plasmatocytes were the main phagocytic hemocytes of Spodoptera exigua determined by observing fluorescence-labeled bacteria in the cytosol. X. nematophila significantly inhibited phagocytosis of both hemocytes, while heat-killed X. nematophila lost its inhibitory potency. However, co-injection of X. nematophila with arachidonic acid did not show any significant inhibition of hemocyte phagocytosis. In fact, hemocytes of S. exigua infected with X. nematophila showed significant reduction in phospholipase A(2) (PLA(2)) activity. Dexamethasone, a specific PLA(2) inhibitor, significantly inhibited phagocytosis of both cell types. However, the inhibitory effect of dexamethasone was recovered by addition of arachidonic acid. Incubation of hemocytes with benzylideneacetone, a metabolite of X. nematophila, inhibited phagocytosis in a dose-dependent manner. These results suggest that X. nematophila produces and secretes PLA(2) inhibitor(s), which in turn inhibit the phagocytic response of hemocytes.     

PGE(2) induces oenocytoid cell lysis via a G protein-coupled receptor in the beet armyworm, Spodoptera exigua

Eicosanoids mediate cellular and humoral immune responses in the beet armyworm, Spodoptera exigua, including activation of prophenoloxidase (PPO). PPO activation begins with release of its inactive zymogen, PPO, from oenocytoids in response to prostaglandins (PGs). Based on the biomedical literature, we hypothesized that PGs exert their actions via specific G protein-coupled receptor(s) in S. exigua. This study reports a G protein-coupled receptor (Se-hcPGGPCR1) gene, which is expressed in the hemocytes of S. exigua. The Se-hcPGGPCR1 consists of 420 amino acids and belongs to rhodopsin-type GPCRs. The high content of hydrophobic amino acid residues within the Se-hcPGGPCR1 protein is explained by prediction of seven-transmembrane domains that are characteristic of these GPCRs. Except for the eggs, Se-hcPGGPCR1 was expressed in all life stages. During the larval stage, it was expressed in hemocytes and gut, but not in fat body nor in epidermis. Real time quantitative RT-PCR showed that bacterial challenge induced more than 20-fold increases in its expression level. Fluorescence in situ hybridization showed that Se-hcPGGPCR1 was expressed in a specific hemocyte type, the oenocytoids. A specific eicosanoid, PGE₂, significantly induced oenocytoid lysis and increased PO activity in the plasma. In contrast, when Se-hcPGGPCR1 expression was suppressed by RNA interference (RNAi), the oenocytoid lysis and the PO activation in response to PGE₂ were not elevated above basal levels. A binding assay using intracellular calcium mobilization showed that the RNAi-treated hemocytes were significantly less responsive to PGE₂ than the control hemocytes. These results support our hypothesis with the specific finding that PGE₂ acts through Se-hcPGGPCR1 to activate PPO by lysing oenocytoids.     

Role of a small G protein Ras in cellular immune response of the beet armyworm, Spodoptera exigua

Insect cellular immune responses accompany cytoskeletal rearrangement of hemocytes to exhibit filopodial and pseudopodial extension of their cytoplasm. Small G proteins are postulated to be implicated in the hemocyte cellular processes to perform phagocytosis, nodulation, and encapsulation behaviors. A small G protein ras gene (Se-Ras) was cloned from cDNAs prepared from hemocytes of the beet armyworm, Spodoptera exigua. The open reading frame of Se-Ras encoded 179 amino acids with a predicted molecular weight of 20.0 kDa, in which 114 residues at amino terminus were predicted to be a GTP binding domain. It showed high sequence similarities (86.1-92.8%) with known ras genes in other insects. Se-Ras was constitutively expressed in all developmental stages from egg to adult without any significant change in expression levels in response to bacterial challenge. A specific double strand RNA (dsRNA) could knockdown its expression in the hemocytes after 48 h post-injection. While the RNA interference (RNAi) did not show any change in total or differential hemocyte counts, it impaired hemocyte behaviors. The RNAi of Se-Ras significantly suppressed hemocyte spreading, cytoskeleton extension, and nodulation behaviors in response to bacterial challenge. Release of prophenoloxidase from oenocytoids was significantly inhibited by the RNAi, which resulted in significant suppression in PO activation in response to an inducer, PGE₂. These results suggest that Se-Ras is implicated in mediating cellular processes of S. exigua hemocytes. This is the first report of Ras role in insect cellular immune response.     

Effect of protein type and quantity on growth and development of larval Heliothis zea and Spodoptera exigua and the endoparasitoid Hyposoter exiguae

Based on the use of casein, soy protein, and corn gluten in artificial diets, we show that larval growth of Heliothis zea (Boddie) and Spodoptera exigua (Hubner) (Lepidoptera: Noctuidae) is significantly reduced both by a deficit and surfeit of dietary protein. The optimal level of protein varies with protein type. Similar protein treatments were also used to examine the influence of host diet on growth and development of the endoparasitoid Hyposoter exiguae (Viereck) (Hymenoptera: Ichneumonidae). Dietary protein regimens reducing host larval growth to the greatest extent are most stressful to the parasitoid, irrespective of protein type. Parasitoid growth appeared to be regulated by the nutritional status of the host. The parasitoid required the host to reach ca 20 mg before it could successfully complete its larval development and pupate. This suggests that host hormones may also be involved in parasitoid development. The implications of these findings for theory of tritrophic interactions are discussed.

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Zusammenfassung Mit Hilfe des Gebrauchs von Kasein, Sojaprotein und Maisgluten in künstlicher Diät zeigen wir, dass das Wachstum der Larven von Heliothis zea (Boddie) und Spodoptera exigua (Hubner) (Lepidoptera: Noctuidae) signifikant reduziert ist durch Defizit oder Überschuss von Protein in der Diät. Der optimale Proteingehalt hängt von der Proteinart ab. Ähnliche Proteindiäten wurden auch benutzt, um den Einfluss von Wirtsdiät auf Wachstum und Entwicklung des Endoparasiten Hyposoter exiguae (Viereck) (Hymenoptera: Icheneumonidae) zu untersuchen. Diejenigen Proteingehaltsstufen, die das Larvenwachstum des Wirts zum grössten Ausmass reduzieren, produzieren den grössten Stress für den Parasiten, unabhängig von der Proteinart. Das Wachstum des Parasiten scheint durch den Ernährungszustand des Wirtes reguliert zu werden. Für die erfolgreiche Larvenentwicklung und Verpuppung des Parasiten war ein Gewicht des Wirtes von ca. 20 mg Voraussetzung. Das deutet auf die Möglichkeit hin, dass Wirtshormone für die Entwicklung des Parasiten eine Rolle spielen können. Schlussfolgerungen aus diesen Ergebnissen für die Theorie tritrophischer Wechselbeziehungen werden diskutiert.

     

In vivo development of the entomogeneous hyphomycetePaecilomyces farinosus in hostSpodoptera exigua (beet armyworm) larvae

The in vivo development of the entomogenous hyphomycetePaecilomyces farinosus inSpodoptera exigua (beet armyworm) larvae was examined using light and electron microscopic techniques. Blastospores injected into larval hemocoels (500 blastospores/larva) were immediately ingested by phagocytic hemocytes, and no fungal cells were detected in the hemolymph until 36 h post-injection. As indicated by immunocytochemical methods, the in vivo-produced blastosopres, in contrast to in vitro blastospores, lacked a galacto-mannan surface layer required for opsonization by aS. exigua humoral lectin. Therefore, these in vivo cells were not recognized by phagocytic granulocytes and were freely-circulating in the hemolymph. Hyphae differentiating from the blastospores were recognized by the hemocytes and induced formation of multicellular hemocytic nodules. By 72 h post-injection, mycelia were observed emerging from the nodules and by 96 h, larvae had become mummified due to extensive proliferation of the fungus throughout host tissues. Neither phagocytosis of the initially injected in vitro-produced blastospores nor nodule formation around hyphal cells later in the infection process was effective in stopping fungal growth. The in vivo development ofP. farinosus was similar to that of another hyphomycete,Beauveria bassiana except that in the latter case, extensive nodule formation was inhibited by the production of fungal metabolites.