Differential responses of midgut soluble aminopeptidases of Helicoverpa armigera to feeding on various host and non-host plant diets

Differential responses of midgut soluble aminopeptidases were studied in Helicoverpa armigera larvae fed on various host (chickpea and pigeon pea) and non-host (bitter gourd and chili) plant diets. Larval growth was significantly reduced by non-host plant diets. Although the serine proteinase activities were inhibited, aminopeptidase activities were significantly increased in the larvae fed on non-host plant diets. Results were qualitatively and quantitatively confirmed with in vivo and in vitro analyses. It was noted that aminopeptidases had given more preference to ApNA than LpNA on non-host plant diets and vice versa on host plant diets. However, optimum pH for aminopeptidase activity (around pH 7.0–8.0) and susceptibility to inhibitors was similar in the larvae fed on host as well as non-host plant diets. These results suggest that H. armigera regulates digestive enzyme levels to obtain better nourishment from the diet and avoid toxicity due to nutritional imbalance. A detailed biochemical and molecular analysis of gut aminopeptidases upon exposure of the insect to a particular diet will highlight their specific roles.     

Compensatory proteolytic responses to dietary proteinase inhibitors from Albizia lebbeck seeds in the Helicoverpa armigera larvae

Plant proteinase inhibitors (PIs) have been shown to reduce the growth rates in larvae of numerous insect species. On the other hand, insects can also regulate their proteinases against plant PIs. In the present study, we report the compensatory activities of Helicoverpa armigera (Hubner) (Lepidoptera: Noctuidae) gut proteinases against the PIs of Albizia lebbeck seeds. Total of ten proteinase inhibitor bands were detected in the seed extract of A. lebbeck. Bioassays were conducted by feeding H. armigera larvae on diet containing partially purified PIs from A. lebbeck seeds. Results show that larval growth and survival was significantly reduced by A. lebbeck PIs. We found that higher activity H. armigera gut proteinase (HGP) isoforms observed in the midgut of control larvae were inhibited in the midgut of larvae fed on test diet. Some HGP isoforms were induced in the larvae fed on PI containing test diet; however, these isoforms showed lower activity in the larvae fed on control diet. Aminopeptidase activities were significantly increased in the midgut of larvae fed on test diet. A population of susceptible and resistant enzymes was observed in the midgut of H. armigera, when fed on diet containing PIs from A. lebbeck seeds. Our initial observations indicate that H. armigera can regulate its digestive proteinase activity against non-host plant PIs, too. It is important to study the exact biochemical and molecular mechanisms underlying this phenomenon in order to develop PI-based insect control strategies.     

Antifeedant and cytotoxic activity of gibberellic acid against Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae)

The present study investigates the effect of gibberellic acid (GA3), a terpenoid and phytohormone, on the digestive physiology and intermediary metabolism of the cotton bollworm Helicoverpa armigera Hübner (Lepidoptera, Noctuidae). Incorporation of GA3 (800 μg g^?1 diet) in an artificial diet results in significant reductions in the rates of diet consumption and the efficiency of conversion of food consumption by by H. armigera larvae. The relative growth rate decreases as the concentration increases. The relative α‐amylase activity in sixth‐instar larvae of H. armigera decreases significantly after ingestion of four concentrations of GA3. Histological studies of the midgut in GA3‐treated larvae (800 μg g^?1 diet) show degeneration of the epithelial cells. The alanine and aspartate aminotransferase activity decreases at the highest concentration. However, acid phosphatase, alkaline phosphatase, γ‐glutamyl transferase and lactate dehydrogenase activity increase significantly compared with the control. The results clearly demonstrate the adverse effects of GA3 on H. armigera via interruption of nutritional physiology and metabolism.     

Behavioral and morphological responses of an insect herbivore to low nutrient quality are inhibited by plant chemical defenses

Animals have several strategies to contend with nutritionally poor diets, including compensatory consumption and enhanced food utilization efficiencies. Plants produce a diversity of defense compounds that affect the ability of herbivores to utilize these strategies in response to variation in food nutritional quality. Little is known, however, about effects of allelochemicals on herbivores utilizing integrated behavioral and morphological responses to reduced food quality. Our objectives were to (1) examine how variation in diet nutritional quality influences compensatory responses of a generalist insect herbivore, and (2) determine how plant defenses affect these processes. Gypsy moth (Lymantria dispar) larvae were administered one of nine combinations of diet having low, moderate, or high nutritional quality and 0, 2, or 4 % purified aspen (Populus tremuloides) salicinoids. We quantified larval growth, consumption, frass production, and biomass allocation to midgut tissue over a 4-day bioassay. In the absence of salicinoids, larvae compensated for reduced nutritional quality and maintained similar growth across all diets through increased consumption, altered midgut biomass allocation, and improved processing efficiencies. Dietary salicinoids reduced larval consumption, midgut biomass allocation, digestive efficiencies, and growth at all nutritional levels, but the effect size was more pronounced when larvae were fed nutritionally suboptimal diets. Our findings demonstrate that integrated behavioral and morphological compensatory responses to reduced food quality are affected by plant defenses, ultimately limiting compensatory responses and reducing larval performance.     

Silencing of acetylcholinesterase gene of Helicoverpa armigera by siRNA affects larval growth and its life cycle

RNA interference is an effective means of regulation of gene expression both in vitro and in vivo. We studied the effect of siRNA on larval development by selective targeting of the acetylcholinesterase (AChE) gene of Helicoverpa armigera. Chemically synthesized siRNA molecules were directly fed to H. armigera larvae along with the artificial diet. The siRNA treatment resulted in specific gene silencing of AChE and consequently brought about mortality, growth inhibition of larvae, reduction in the pupal weight, malformation and drastically reduced fecundity as compared to control larvae. Our studies suggest some novel roles for AChE in growth and development of insect larvae and demonstrate that siRNA can be readily taken up by insect larvae with their diet.     

Effect of jasmonic acid and salicylic acid induced resistance in groundnut on Helicoverpa armigera

Induced resistance in plants affects insect growth and development as a result of the up‐regulation of defence‐related secondary metabolites or enzyme‐binding proteins. In the present study, the effects of jasmonic acid (JA) and salicylic acid (SA) induced resistance in groundnut on Helicoverpa armigera (Hübner) are examined. Larval survival, larval weights and the activities of digestive enzymes (total serine protease and trypsin) and of detoxifying enzymes [glutathione S‐transferase (GST) and esterase (EST)] are studied in insects fed on four groundnut genotypes with moderate levels of resistance to H. armigera (ICGV 86699, ICGV 86031, ICG 2271 and ICG 1697) and a susceptible genotype (JL 24). The plants are pre‐ and/or simultaneously treated with JA and SA, and then infested with H. armigera, which are allowed to feed for 6 days. Significantly lower serine protease and trypsin activities are observed in H. armigera fed on plants treated with JA. Greater GST activity is recorded in insects fed on JA and SA treated plants, whereas EST activity is low in H. armigera larvae fed on plants treated with JA and SA. Serine proteases, trypsin and GST activities and larval weights (r = 0.74–0.95) and larval survival (r = 0.77–0.93) are positively correlated, whereas EST activity and larval weight (r = ?0.55) and larval survival (r = ?0.65) are negatively correlated. The results suggest that midgut digestive and detoxifying enzymes can be used as indicators of the adverse effects of constitutive and/or induced resistance in crop plants on the insect pests and the role of JA and SA in insect pest management.     

A host-plant specialist, Helicoverpa assulta, is more tolerant to capsaicin from Capsicum annuum than other noctuid species

Plant secondary compounds not only play an important role in plant defense, but have been a driving force for host adaptation by herbivores. Capsaicin (8-methyl-N-vanillyl-6-nonenamide), an alkaloid found in the fruit of Capsicum spp. (Solanaceae), is responsible for the pungency of hot pepper fruits and is unique to the genus. The oriental tobacco budworm, Helicoverpa assulta (Lepidoptera: Noctuidae), is a specialist herbivore feeding on solanaceous plants including Capsicum annuum, and is one of a very few insect herbivores worldwide capable of feeding on hot pepper fruits. To determine whether this is due in part to an increased physiological tolerance of capsaicin, we compared H. assulta with another specialist on Solanaceae, Heliothis subflexa, and four generalist species, Spodoptera frugiperda, Heliothis virescens, Helicoverpa armigera, and Helicoverpa zea, all belonging to the family Noctuidae. When larvae were fed capsaicin-spiked artificial diet for the entire larval period, larval mortality increased in H. subflexa and H. zea but decreased in H. assulta. Larval growth decreased on the capsaicin-spiked diet in four of the species, was unaffected in H. armigera and increased in H. assulta. Food consumption and utilization experiments showed that capsaicin decreased relative consumption rate (RCR), relative growth rate (RGR) and approximate digestibility (AD) in H. zea, and increased AD and the efficiency of conversion of ingested food (ECI) in H. armigera; whereas it did not significantly change any of these nutritional indices in H. assulta. The acute toxicity of capsaicin measured by injection into early fifth instar larvae was less in H. assulta than in H. armigera and H. zea. Injection of high concentrations produced abdominal paralysis and self-cannibalism. Injection of sub-lethal doses of capsaicin resulted in reduced pupal weights in H. armigera and H. zea, but not in H. assulta. The results indicate that H. assulta is more tolerant to capsaicin than the other insects tested, suggesting that this has facilitated expansion of its host range within Solanaceae to Capsicum after introduction of the latter to the Old World about 500 years ago. The increased larval survival and growth due to chronic dietary exposure to capsaicin suggests further adaptation of H. assulta to that compound, the mechanisms of which remain to be investigated.     

Coupling diet quality and Bowman-Birk and Kunitz-type soybean proteinase inhibitor effectiveness to Diatraea saccharalis development and mortality

We used bioassays to investigate the effect of Bowman‐Birk and Kunitz‐type soybean proteinase inhibitors on two artificial diets (diets 1 and 2) which are commonly used to feed laboratory colonies of larvae of the moth Diatraea saccharalis, monitoring food intake and utilization, and larval development and mortality. Diet 1 was less nutritious, with a low protein content and reduced mineral and essential amino acid (e.g., cysteine, lysine, and methionine) content, while diet 2 was richer and more complete. When proteinase inhibitors were incorporated into the artificial diets, the effects on larval development were significantly greater for those larvae fed diet 1, with the chronic ingestion of proteinase inhibitors reducing the level of trypsin‐like activity in the midgut of larvae fed this diet. Larvae fed diet 2 also showed a reduced level of tryptic activity in the midgut, but this was less marked than for diet 1. These results indicate that despite their inhibitory effect on midgut enzymes, the effectiveness of proteinase inhibitors is directly dependent on the quality of the diet. The different effects seen on insect biology when proteinase inhibitors are added to rich or poor diets suggests that the role of anti‐nutritional proteins in the control of insects might not be adequately addressed by bioassays based on the incorporation of inhibitors into artificial diets.     

CARBOHYDRASES IN THE DIGESTIVE SYSTEM OF THE SPINED SOLDIER BUG,Podisus maculiventris (SAY) (HEMIPTERA: PENTATOMIDAE)

The spined soldier bug, Podisus maculiventris, is a generalist predator of insects and has been used in biological control. However, information on the digestion of food in this insect is lacking. Therefore, we have studied the digestive system in P. maculiventris, and further characterized carbohydrases in the digestive tract. The midgut of all developmental stages was composed of anterior, median, and posterior regions. The volumes of the anterior midgut decreased and the median midgut increased in older instars and adults, suggesting a more important role of the median midgut in food digestion. However, carbohydrase activities were predominant in the anterior midgut. In comparing the specific activity of carbohydrases, α‐amylase activity was more in the salivary glands (with two distinct activity bands in zymograms), and glucosidase and galactosidase activities were more in the midgut. Salivary α‐amylases were detected in the prey hemolymph, demonstrating the role of these enzymes in extra‐oral digestion. However, the catalytic efficiency of midgut α‐amylase activity was approximately twofold more than that of the salivary gland enzymes, and was more efficient in digesting soluble starch than glycogen. Midgut α‐amylases were developmentally regulated, as one isoform was found in first instar compared to three isoforms in fifth instar nymphs. Starvation significantly affected carbohydrase activities in the midgut, and acarbose inhibited α‐amylases from both the salivary glands and midgut in vitro and in vivo. The structural diversity and developmental regulation of carbohydrases in the digestive system of P. maculiventris demonstrate the importance of these enzymes in extra‐oral and intra‐tract digestion, and may explain the capability of the hemipteran to utilize diverse food sources.     

Penicillin-induced oxidative stress: effects on antioxidative response of midgut tissues in instars of Galleria mellonella

Penicillin and other antibiotics are routinely incorporated in insect culture media. Although culturing insects in the presence of antibiotics is a decades-old practice, antibiotics can exert deleterious influences on insects. In this article, we test the hypothesis that one of the effects of dietary penicillin is to increase oxidative stress on insects. The effects of penicillin on midgut concentrations of the oxidative stress indicator malondialdehyde (MDA) and on midgut antioxidant enzyme (superoxide dismutase [SOD], catalase [CAT], glutathione S-transferase [GST], and glutathione peroxidase [GPx]) and transaminases (alanine aminotransferase and aspartate aminotransferase) activities in greater wax moth, Galleria mellonella (L.), were investigated. The insects were reared from first instars on artificial diets containing 0.001, 0.01, 0.1, or 1.0 g penicillin per 100 g of diets. MDA content was significantly increased in the midgut tissues of each larval instar reared in the presence of high penicillin concentrations. Activities of antioxidant and transaminase enzymes did not show a consistent pattern with respect to penicillin concentrations in diet or age of larvae. Despite the increased penicillin-induced oxidative stress in gut tissue, antioxidant and transaminase enzymes did not correlate with oxidative stress level or between each other in larvae of other age stages except for the seventh instar. We found a significant negative correlation of MDA content with SOD and GST activities in seventh instars. SOD activity was also negatively correlated with CAT activity in seventh instars. These results suggest that exposure to dietary penicillin resulted in impaired enzymatic antioxidant defense capacity and metabolic functions in wax moth larval midgut tissues and that the resulting oxidative stress impacts midgut digestive physiology.     

The effect of GNA lectin on the α-amylase activity of the beet armyworm,Spodoptera exigua Hb. (Lepidoptera: Noctuidae)

Beet armyworm (Spodoptera exigua Hb.) (Lepidoptera: Noctuidae) is the major pest of sugar beet (Beta vulgaris). Pesticide applications are the main method of the insect control. So, alternative method/s is/are needed to control this insect species. So, in the current study, the effect of Galanthus nivalis agglutinin (GNA) (snowdrop lectin) on beet armyworm α-amylase was studied. Measurement of the amylase activity of the larval midgut fed on artificial diet and sugar beet leaves showed that the enzyme activity was higher when the larvae fed on artificial diet. However, in both cases, the fourth instar larvae had the greatest amylase activity. Thus, fourth instar larvae were offered artificial diet containing 1 and 2% GNA. Both treatments of the lectin significantly reduced the α-amylase activity of the insect. For example, amylase activity of the fourth instar larvae in the control (fed only on artificial diet) was 2.62 Uml^?1 whilst the activity of the enzyme in the two treatments including diet containing 2 and 1% lectin was 1.45 and 1.75 Uml^?1, respectively. The achieved data showed that lectin, in addition to have toxic effect on the larval growth and development, affects the α-amylase activity of the insect gut.     

Elevated ozone induces jasmonic acid defense of tomato plants and reduces midgut proteinase activity in Helicoverpa armigera

As a consequence of membrane lipid peroxidation, foliar defense compounds are changed by elevated ozone (O₃), which in turn affects the palatability and performance of insect herbivores. The induced defense of two tomato [Solanum esculentum L. (Solanaceae)] genotypes, namely jasmonic acid (JA) pathway‐deficient mutant spr2 and its wild‐type control, was studied in response to cotton bollworm, Helicoverpa armigera Hübner (Lepidoptera: Noctuidae), as well as the digestive adaptation of these insects under elevated O₃ in open‐top field chambers. Our data indicated that elevated O₃ increased foliar JA and salicylic acid (SA) levels simultaneously and up‐regulated proteinase inhibitors (PIs) and lipoxidase activities in wild‐type plants, regardless of H. armigera infestation. In contrast, only the O₃+H. armigera treatment increased free SA levels in spr2 plants, but did not affect JA level or PI activities. Additionally, the lower activity of midgut digestive enzymes, including active alkaline trypsin‐like enzyme and chymotrypsin‐like enzyme, was observed in the midgut of cotton bollworms after they consumed wild‐type plants treated for 2 h with elevated O₃. With temporary increases at 8 h, all four digestive enzymes of interest in the insect midgut dropped when they were fed with wild‐type plants under elevated O₃ treatment. Increases in atmospheric O₃ are thought to increase JA signaling and consequently reduce the activities of midgut digestive enzymes in H. armigera, therefore enhancing plant resistance against insect herbivores.     

Bioinsecticidal activity of Talisia esculenta reserve protein on growth and serine digestive enzymes during larval development of Anticarsia gemmatalis

Plants synthesize a variety of molecules to defend themselves against an attack by insects. Talisin is a reserve protein from Talisia esculenta seeds, the first to be characterized from the family Sapindaceae. In this study, the insecticidal activity of Talisin was tested by incorporating the reserve protein into an artificial diet fed to the velvetbean caterpillar Anticarsia gemmatalis, the major pest of soybean crops in Brazil. At 1.5% (w/w) of the dietary protein, Talisin affected larval growth, pupal weight, development and mortality, adult fertility and longevity, and produced malformations in pupae and adult insects. Talisin inhibited the trypsin-like activity of larval midgut homogenates. The trypsin activity in Talisin-fed larvae was sensitive to Talisin, indicating that no novel protease-resistant to Talisin was induced in Talisin-fed larvae. Affinity chromatography showed that Talisin bound to midgut proteinases of the insect A. gemmatalis, but was resistant to enzymatic digestion by these larval proteinases. The transformation of genes coding for this reserve protein could be useful for developing insect resistant crops.     

Pupal diapause of Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) mediated by larval host plants: pupal weight is important

Facultative diapause, a strategy that allows insects to initiate additional generations when conditions are favorable or to enter diapause when they are not, has a profound effect on the ecology and evolution of species. Most previous studies have concentrated on the role of photoperiod and temperature in inducing facultative diapause in insects. In contrast, here we studied pupal diapause mediated by larval host plants in the cotton bollworm Helicoverpa armigera, and confirmed that pupal weight is a critical factor. Two groups of third instar H. armigera larvae, kept at 25 °C with L:D = 8:16 and 20 °C with photoperiod of L:D = 8:16, respectively, were fed on six host plants and on artificial diet (as a control) to determine how larval host plants affect diapause incidence and related traits (such as pupal weight and developmental duration). The data showed larval host plants affected diapause incidence significantly and the effects could be masked by low temperature. Further analysis showed that pupal size, not the length of the sensitive stage, affected the decision to enter diapause. In a further experiment, third-instar to final-stage larvae deprived of artificial diet for 2 days demonstrated a direct relationship between pupal weight and diapause incidence. These results suggest that larval host plants, by affecting pupal size, may influence diapause occurrence in H. armigera. This has important adaptive significance for both over-wintering survival and the possibility for completing an additional generation.     

Insecticidal Potential of Defense Metabolites from Ocimum kilimandscharicum against Helicoverpa armigera

Genus Ocimum contains a reservoir of diverse secondary metabolites, which are known for their defense and medicinal value. However, the defense-related metabolites from this genus have not been studied in depth. To gain deeper insight into inducible defense metabolites, we examined the overall biochemical and metabolic changes in Ocimum kilimandscharicum that occurred in response to the feeding of Helicoverpa armigera larvae. Metabolic analysis revealed that the primary and secondary metabolism of local and systemic tissues in O. kilimandscharicum was severely affected following larval infestation. Moreover, levels of specific secondary metabolites like camphor, limonene and β-caryophyllene (known to be involved in defense) significantly increased in leaves upon insect attack. Choice assays conducted by exposing H. armigera larvae on O. kilimandscharicum and tomato leaves, demonstrated that O. kilimandscharicum significantly deters larval feeding. Further, when larvae were fed on O. kilimandscharicum leaves, average body weight decreased and mortality of the larvae increased. Larvae fed on artificial diet supplemented with O. kilimandscharicum leaf extract, camphor, limonene and β-caryophyllene showed growth retardation, increased mortality rates and pupal deformities. Digestive enzymes of H. armigera - namely, amylase, protease and lipase- showed variable patterns after feeding on O. kilimandscharicum, which implies striving of the larvae to attain required nutrition for growth, development and metamorphosis. Evidently, selected metabolites from O. kilimandscharicum possess significant insecticidal activity.     

Aqueous washings of host plant surface influence the growth inhibitory effects of δ‐endotoxins of Bacillus thuringiensis against Helicoverpa armigera

Biological activity of the bacterium Bacillus thuringiensis Berliner (Bt) against insect pests is influenced by the host plants. To understand the underlying mechanism of variation in biological activity of Bt on host plants, we studied the effect of chemicals from the surface of chickpea (Cicer arietinum L., Fabaceae) leaves (ICCC 37 and ICC 506EB), sorghum [Sorghum bicolor (L.) Moench, Poaceae] grain (ICSV 745 and IS 18698), pigeon pea [Cajanus cajan (L.) Millsp., Fabaceae] pods (ICPL 87 and ICPL 332WR), and cotton (Gossypium hirsutum L., Malvaceae) squares (RCH 2 and Bt RCH 2), on which Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) feeds under natural conditions. Surface chemicals extracted in water from host plant leaves were added to the standard artificial diet containing a commercial formulation of Bt or Cry1Ac. Data were recorded on larval and pupal weights, pupation, adult emergence, larval and pupal periods, adult longevity, and fecundity. Weights of H. armigera at 5 days after initiation of the experiment were significantly reduced on artificial diets containing Bt + pod washings of ICPL 87 and ICPL 332WR, grain washings of ICSV 745, or square washings of RCH 2, and Cry1Ac + leaf‐surface washings of ICC 506EB. Pupal weights were lower on diets containing leaf‐surface washings of ICCC 37 + Bt than on standard artificial diet. Larval periods were prolonged on diets containing Bt + leaf‐surface washings of ICCC 37, pod washings of ICPL 87, and square washings of RCH 2, and on standard artificial diet + Cry1Ac. Pupation was significantly higher on standard artificial diet + Cry1Ac than on diets with Bt + grain washings of ICSV 745 and Cry1Ac + square washings of RCH 2 and Bt RCH 2. Adult emergence was lowest on diets with square washings of RCH 2 + Bt and grain washings of ICSV 745 + Cry1Ac. The results suggested that leaf‐surface washings play an important role in biological activity of Bt/Cry1Ac against H. armigera.     

Ultrastructure and immunolocalization of digestive enzymes in the midgut of Podisus nigrispinus (Heteroptera: Pentatomidae)

The predatory stinkbug Podisus nigrispinus has been utilized in biological control programs. Its midgut is anatomically divided into anterior, middle and posterior regions, which play different roles in the digestive process. We describe the midgut ultrastructure and the secretion of digestive enzymes in the midgut of P. nigrispinus. Midguts were analyzed with transmission electron microscopy and the digestive enzymes amylase, cathepsin L, aminopeptidase and α-glucosidase were immunolocalized. The ultrastructural features of the digestive cells in the anterior, middle and posterior midgut regions suggest that they play a role in digestive enzyme synthesis, ion and nutrient absorption, storage and excretion. The digestive enzymes have different distribution along the midgut regions of the predator P. nigrispinus. Amylase, aminopeptidase and α-glucosidase occur in three midgut regions, whereas cathepsin L occurs in the middle and posterior midgut regions. The anterior midgut region of P. nigrispinus seems to play a role in water absorption, the middle midgut may be involved in nutrient absorption and the posterior midgut region is responsible for water transport to the midgut lumen.     

Composition of the Spruce Budworm (Choristoneura fumiferana) Midgut Microbiota as Affected by Rearing Conditions

The eastern spruce budworm (Choristoneura fumiferana) is one of the most destructive forest insect pests in Canada. Little is known about its intestinal microbiota, which could play a role in digestion, immune protection, communication and/or development. The present study was designed to provide a first characterization of the effects of rearing conditions on the taxonomic diversity and structure of the C. fumiferana midgut microbiota, using a culture-independent approach. Three diets and insect sources were examined: larvae from a laboratory colony reared on a synthetic diet and field-collected larvae reared on balsam fir or black spruce foliage. Bacterial DNA from the larval midguts was extracted to amplify and sequence the V6-V8 region of the 16S rRNA gene, using the Roche 454 GS-FLX technology. Our results showed a dominance of Proteobacteria, mainly Pseudomonas spp., in the spruce budworm midgut, irrespective of treatment group. Taxonomic diversity of the midgut microbiota was greater for larvae reared on synthetic diet than for those collected and reared on host plants, a difference that is likely accounted for by several factors. A greater proportion of bacteria from the phylum Bacteroidetes in insects fed artificial diet constituted the main difference between this group and those reared on foliage; within the phylum Proteobacteria, the presence of the genus Bradyrhizobium was also unique to insects reared on artificial diet. Strikingly, a Bray-Curtis analysis showed important differences in microbial diversity among the treatment groups, pointing to the importance of diet and environment in defining the spruce budworm midgut microbiota.     

Characterization of the Mamestra configurata (Lepidoptera: Noctuidae) larval midgut protease complement and adaptation to feeding on artificial diet,Brassica species,and protease inhibitor

The midgut protease profiles from 5th instar Mamestra configurata larvae fed various diets (standard artificial diet, low protein diet, low protein diet with soybean trypsin inhibitor [SBTI], or Brassica napus) were characterized by one‐dimensional enzymography in gelatin gels. The gut protease profile of larvae fed B. napus possessed protease activities of molecular masses of approximately 33 and 55 kDa, which were not present in the guts of larvae fed artificial diet. Similarly, larvae fed artificial diet had protease activities of molecular masses of approximately 21, 30, and 100 kDa that were absent in larvae fed B. napus. Protease profiles changed within 12 to 24 h after switching larvae from artificial diet to plant diet and vice versa. The gut protease profiles from larvae fed various other brassicaceous species and lines having different secondary metabolite profiles did not differ despite significant differences in larval growth rates on the different host plants. Genes encoding putative digestive proteolytic enzymes, including four carboxypeptidases, five aminopeptidases, and 48 serine proteases, were identified in cDNA libraries from 4th instar M. configurata midgut tissue. Many of the protease‐encoding genes were expressed at similar levels on all diets; however, three chymoptrypsin‐like genes (McSP23, McSP27, and McSP37) were expressed at much higher levels on standard artificial diet and diet containing SBTI as was the trypsin‐like gene McSP34. The expression of the trypsin‐like gene McSP50 was highest on B. napus. The adaptation of M. configurata digestive biochemistry to different diets is discussed in the context of the flexibility of polyphagous insects to changing diet sources. Published 2010 Wiley Periodicals, Inc.