Nutrient utilization by caterpillars of the generalist beet armyworm, Spodoptera exigua

Abstract.  Beet armyworm, Spodoptera exigua (Hübner) (Lepidoptera: Noctuidae), caterpillars are destructive crop pests responsible for considerable annual economic losses. These generalist herbivores are challenged with a diversity of dietary choices that can impact their survival, development and fecundity. In the present study, dietary choices of S. exigua caterpillars, based on the protein to digestible carbohydrate (P : C) ratio of the food, and the impact of nutritionally unbalanced foods on caterpillar performance are assessed. In choice experiments, individual third-instar caterpillars are offered simultaneously a P-biased and a C-biased food until pupation. Caterpillars feed nonrandomly and select a slightly P-biased diet (22P : 20C). In no-choice experiments, second instar caterpillars are reared until pupation on diets ranging in P : C ratio from extremely P- to extremely C-biased. High mortality and delayed development are observed on the C-rich, P-poor diets, highlighting the potential deleterious effects of excess carbohydrates and the importance of protein for growth and development. Diet-dependent differences in pupal weight or pupal lipid reserves are not observed. This contrasts with closely-related Spodoptera species where pupal mass and lipid stores increase on C-rich, P-poor diets. On the extremely P-biased diet, performance is similar to that of individuals reared on the self-selected diet, suggesting that these caterpillars may efficiently be deaminating excess amino acids to generate carbon skeletons, which are shunted into lipid biosynthesis. Spodoptera exigua caterpillars exhibit flexible and efficient pre-ingestive nutrient intake regulation and post-ingestive utilization, allowing these generalist feeders to cope with the heterogeneous diets they may encounter.     

Diet factors responsible for the change of the glucose oxidase activity in labial salivary glands of Helicoverpa armigera

We investigated the change of the glucose oxidase (GOX) activity in labial salivary glands of Helicoverpa armigera larvae fed with the artificial diet or host plant tobacco and the major factors responsible for such a change. Throughout larval development, the labial salivary GOX activities in caterpillars reared on the artificial diet were remarkably higher than those fed with the plant. After fifth-instar plant-fed caterpillars were transferred to the artificial diet, their labial salivary GOX activity increased quickly, which was closely correlated with the time spent feeding on the artificial diet. The total sugar content of the artificial diet was 68 times higher than that of the tobacco leaves. We hypothesized that sugars and secondary metabolites are the possible causes of induction of GOX activity. When fifth-instar caterpillars were fed with tobacco leaves coated with glucose or sucrose, their labial salivary GOX activity was significantly higher than those fed with leaves without sugar coating. Following native PAGE, 1 single band of the labial salivary GOX was observed in all the caterpillars fed with different diets, implying that only the activity of the isoenzyme was changed in response to different diets. Furthermore, the labial salivary GOX activity was determined after caterpillars were fed with artificial diets containing chlorogenic acid, rutin, and quercetin. The results showed that all these phenolic compounds had no effect on the GOX activity. We conclude that sugar in diets was a major factor influencing the labial salivary GOX activity of the larvae. Arch. Insect Biochem. Physiol. 2008.     

Evidence that caterpillar labial saliva suppresses infectivity of potential bacterial pathogens

Salivary enzyme, glucose oxidase (GOX) from the caterpillar Helicoverpa zea, catalyzes the conversion of glucose to gluconic acid and hydrogen peroxide. Because hydrogen peroxide has well-known antimicrobial properties, we examined whether caterpillar labial saliva could reduce the infectivity of bacterial pathogens. We examined the effects of caterpillar saliva on the growth of two bacteria species Serratia marcescens and Pseudomonas aeruginosa. Wells formed in LB agar contained a solution of salivary gland extract (Sx) and glucose, GOX and glucose, Sx only, GOX only, or glucose only. After 18 h of incubation, the diameter of cleared bacteria was measured. Wells treated with only GOX, Sx, or glucose showed no measurable area of clearing, while wells treated with GOX with glucose or Sx with glucose had considerable clearing. To determine if saliva could provide protection to caterpillars in vivo, a surgery was performed on caterpillars that prevented the secretion of labial saliva. Caterpillars were fed a diet containing either no added bacteria or treated with high levels of S. marcescens or P. aeruginosa. Caterpillars that could not secrete saliva had significantly higher levels of mortality when feeding on diet treated with either bacterium than caterpillars that could secrete saliva when feeding on equal levels of bacteria-treated diet. Our evidence demonstrates for the first time that insect saliva in situ can provide protection against bacterial pathogens and that the salivary enzyme GOX appears to provide the antimicrobial properties.     

Sex-specific differences in nutrient regulation in a capital breeding caterpillar, Spodoptera litura (Fabricius)

Nutrient requirements by male and female insects are likely to differ, but relatively little is known regarding how sexes differ in their regulation of macronutrient acquisition. The present study reports the results from a laboratory experiment in which behavioural and physiological components of nutrient regulation were compared between male and female caterpillars of Spodoptera litura (Fabricius). When provided with choices between two nutritionally complementary foods (one is a protein-biased diet and the other a carbohydrate-biased diet), both males and females adjusted their food selection to defend an intake target. However, the composition of diet preferred by the two differed, with females selecting significantly more protein than males with no difference in carbohydrate intake between the two. When confined to single diets with varying mixtures of protein and carbohydrate [P:C ratios, expressed as the percentage of diet by dry mass: protein 42%:carbohydrate 0% (p42:c0), p35:c7, p28:c14, p21:c21, p14:c28, p7:c35], females consumed more macronutrients than did males across on all P:C diets except the extremely carbohydrate-biased diet (p7:c35). Under both choice and no-choice feeding condition, such sex differences in nutrient intake were not expressed until late in the feeding stage of the final stadium. Sexes also differed in post-ingestive utilization of ingested nutrients. Females utilized ingested protein for body growth with greater efficiency compared to males, presumably reflecting provisioning their adult needs for protein to develop eggs, whereas males were more efficient at depositing lipids from carbohydrate intake than females.     

A geometric analysis of nutrient regulation in the generalist caterpillar Spodoptera littoralis (Boisduval)

Nutritional regulatory responses of the generalist caterpillar Spodoptera littoralis (Boisduval) were explored, in choice and no-choice experiments, using the Geometric Framework. In the choice experiment, newly moulted final instar larvae were provided with one of three protein-biased foods (PB-food: p35:c7, p28:c5.6 or p21:c4.2) and one of three equal protein-carbohydrate ratio foods (ER-food: p21:c21, p16.8:c16.8 or p12.6:c12.6). On five of the nine treatments, caterpillars independently regulated protein and carbohydrate intake to a mixture of 57 and 43%, respectively. However, when the concentration of the ER-food decreased and that of the paired PB-food increased, caterpillars progressively abandoned regulation and ate more of the PB-food. Despite these regulatory differences, performance (survivorship, growth and development) was similar across all nine treatments. In the no-choice experiment, caterpillars were given one of five foods (p35:c7, p28:c14, p21:c21, p14:c28 and p7:c35). Results indicated that caterpillars moved to a point in protein-carbohydrate space that was consistent with the Equal Distance Rule of compromise, a pattern previously predicted for generalist feeders. The insects on the two extreme foods, p35:c7 and p7:c35, showed reduced pupal mass and longer development, respectively. There was also strong evidence for post-ingestive regulation of nutrient utilisation, notably for protein.     

Three hundred and fifty generations of extreme food specialisation: testing predictions of nutritional ecology

We used a strain of diamondback moth, Plutella xylostella L. (Lepidoptera: Plutellidae), that had been reared for approximately 350 generations in a precisely characterised environment to test hypotheses regarding the influence of nutritional heterogeneity on the evolution of nutrient regulatory responses. Caterpillars were maintained with ad libitum access to a diet that emulated that of an extreme nutritional specialist, comprising a homogeneous food of fixed nutrient composition. We measured performance (survival, development rate, and pupal mass), as well as the protein and carbohydrate intake of individual caterpillars confined to one of a range of single foods differing in their protein, carbohydrate, and water content. In a separate experiment, we measured the amount and balance of protein and carbohydrate self-selected by caterpillars presented with nutritionally complementary foods. Results showed a close fit with three of four predictions about the nutritional responses of 'nutrient specialist' feeders: (1) survival, development rate, and pupal mass were highest for animals given diets with the protein:carbohydrate composition of the ancestral culture diet, and dropped off sharply with higher and lower protein:carbohydrate balance, (2) caterpillars coped poorly with dietary dilution by water, irrespective of the macronutrient balance, and (3) the self-selected intake point corresponded with the macronutrient balance that gave peak performance (i.e., that of the ancestral culture diet). The fourth prediction, that caterpillars would be disinclined to over-ingest nutrients on imbalanced diets, was at best weakly met. We hypothesise that the evolution and maintenance of the specialist strategy might, paradoxically, require some degree of environmental heterogeneity.     

Do food protein and carbohydrate content influence the pattern of feeding and the tendency to explore of forest tent caterpillars?

This study examines whether the ratio of protein to carbohydrate affects the timing of meals and the propensity to explore of forest tent caterpillars (Malacosoma disstria). The behavior of fourth instar caterpillars was observed on three semi-defined artificial diets varying in protein (p)-carbohydrate (c) ratio. These diets were (a) p14:c28, (b) p28:c14, and (c) p35:c7. The probability of initiating feeding at first contact with the food and the duration of the first feeding event did not vary across diets, suggesting not much difference in phagostimulatory power. There was also no difference in the total time spent eating, at rest and in motion between diets. However, the timing and duration of meals varied significantly; more short meals were observed on the carbohydrate-biased diet. The duration of pauses between meals also increased with food protein content. Furthermore, caterpillars on the carbohydrate-biased diet were more likely to leave the trail leading to the known food source and to discover a second food source, suggesting that protein deprivation promotes exploration. These findings shed insight into the physiological responses to protein and carbohydrate ingestion and demonstrate how post-ingestive effects can favor consumption of foods containing protein without invoking an explicit mechanism of independent nutrient regulation, but simply by influencing the pattern of feeding and the propensity to explore.     

Dietary self‐selection and rules of compromise by fifth‐instar Vanessa cardui

We examined dietary self‐selection and rules of compromise for protein (P) and digestible carbohydrate (C) intake by fifth‐instar Vanessa cardui L. (Lepidoptera: Nymphalidae: Nymphalini). We presented six fat‐free diet pairs to larvae in a choice trial to determine the ‘intake target’. In addition, we fed larvae seven fat‐free single diets differing in dietary nutrient ratio in no‐choice trials to determine the rules of compromise they exhibit when constrained to a singular, sub‐optimal dietary source. In choice trials, caterpillars regulated nutrient intake to a ratio of 1 protein to 1.09 carbohydrate (1P:1.09C), exhibiting tighter regulation of protein than of carbohydrate. Furthermore, larvae from different diet pair treatments did not differ in pupal mass or stadium duration. In no‐choice experiments, larvae reduced consumption on increasingly protein‐biased diets and increased consumption on increasingly carbohydrate‐biased diets, relative to a 1P:1C ratio diet. Differences in carbohydrate consumption were much greater between no‐choice treatments than differences in protein consumption. Dietary nutrient ratio affected pupal mass when accounting for initial larval mass. Pupal mass decreased as nutrient ratio was shifted off of 1P:1C, but to a greater extent when the ratio was skewed toward carbohydrate. Stadium duration increased as nutrient ratio diverged from 1P:1C, being more pronounced when shifted toward carbohydrate than toward protein. Regulation to near 1P:1C is consistent with results found for other Lepidoptera, and the rule of compromise exhibited by V. cardui is consistent with that expected for a generalist herbivore.     

Revisiting macronutrient regulation in the polyphagous herbivore Helicoverpa zea (Lepidoptera: Noctuidae): New insights via nutritional geometry

Insect herbivores that ingest protein and carbohydrates in physiologically-optimal proportions and concentrations show superior performance and fitness. The first-ever study of protein–carbohydrate regulation in an insect herbivore was performed using the polyphagous agricultural pest Helicoverpa zea. In that study, experimental final instar caterpillars were presented two diets – one containing protein but no carbohydrates, the other containing carbohydrates but no protein – and allowed to self-select their protein–carbohydrate intake. The results showed that H. zea selected a diet with a protein-to-carbohydrate (p:c) ratio of 4:1. At about this same time, the geometric framework (GF) for the study of nutrition was introduced. The GF is now established as the most rigorous means to study nutrient regulation (in any animal). It has been used to study protein–carbohydrate regulation in several lepidopteran species, which exhibit a range of self-selected p:c ratios between 0.8 and 1.5. Given the economic importance of H. zea, and it is extremely protein-biased p:c ratio of 4:1 relative to those reported for other lepidopterans, we decided to revisit its protein–carbohydrate regulation. Our results, using the experimental approach of the GF, show that H. zea larvae self-select a p:c ratio of 1.6:1. This p:c ratio strongly matches that of its close relative, Heliothis virescens, and is more consistent with self-selected p:c ratios reported for other lepidopterans. Having accurate protein and carbohydrate regulation information for an insect herbivore pest such as H. zea is valuable for two reasons. First, it can be used to better understand feeding patterns in the field, which might lead to enhanced management. Second, it will allow researchers to develop rearing diets that more accurately reflect larval nutritional needs, which has important implications for resistance bioassays and other measures of physiological stress.     

Competition for dietary carbohydrates between streptococci in dental plaque

Abstract The predominant microorganisms in dental plaque, i.e., streptococci and actinomycetes, are carbohydrate fermenters. In the natural environment these organisms experience carbohydrate limitation interrupted by periods of substrate excess following food intake by the host. Recently, we have studied the competition between pairs of oral Streptococcus species in the chemostat under continuous glucose or sucrose limitation and under glucose pulsed conditions.
In the present study we have investigated the competition for dietary carbohydrates in dental plaque of gnotobiotic rats infected with the same combinations of streptococci. The rats were exposed to similar regimes of carbohydrate administration as in the chemostat by incorporation of low molecular carbohydrates in the diet and drinking water. Addition of glucose to the diet favoured the organism with the highest q _max for glucose. This result parallels the outcome of earlier competition experiments in glucose-pulsed chemostats, which showed that the organism with the highest q _max glucose became dominant in the cultures. Previous observations that bacteriocin production and extracellular glucan synthesis are major ecological factors for oral streptococci were also confirmed in this experiment.     

Temperature‐by‐nutrient interactions affecting growth rate in an insect ectotherm

Temperature and nutrition are two prominent environmental variables influencing juvenile growth rate in ectotherms. These two factors interact in complex ways. Here, we present a comprehensive analysis of the interactive effects of temperature and nutrition on various components of fitness (growth rate, survival), food intake, and level of energy storage in an insect herbivore, caterpillars of Spodoptera exigua Hübner (Lepidoptera: Noctuidae). In a factorial experimental design, final‐instar caterpillars (i.e., fifth instars) were individually reared at one of three constant temperatures (18, 26, and 34 °C), in which they received one of six diets differing in their ratio of protein and digestible carbohydrate [P:C mixture, expressed as the percentage of diet by dry mass: protein 42%:carbohydrate 0% (42:0), 35:7, 28:14, 21:21, 14:28, and 7:35]. Within the range of test temperatures, larval growth rate increased with rising temperature and was strongly affected by P:C mixture, reaching a maximum on moderate P:C diets at each temperature and falling at very high and low P:C mixtures. There was a significant temperature*diet interaction, such that the difference in growth rates between temperatures was greatest on moderate P:C diets and least on the most extreme diets (42:0 and 7:35). Food intake rate patterns followed a similar trend to growth rate. Rapidly growing animals at high ambient temperature suffered high mortality across all dietary P:C mixtures, but to a greater extent on the extremely unbalanced diets. This suggests that there are developmental and physiological costs associated with fast growth at high temperature, as indicated by high rate of pupation failure and reduced lipid storage efficiency. Our study shows how temperature and nutrition interplay to mediate phenotypic variations in growth rates and energy utilization in an insect ectotherm.     

Influence of dietary composition on growth and energy reserves in tench (Tinca tinca)

The effects of different protein, lipid and carbohydrate diets on growth and energy storage in tench, Tinca tinca L., were studied. Over a 2-month period fish were fed four different diets: control, protein-enriched, carbohydrate-enriched and lipid-enriched. The best growth rates were obtained with the control and protein-enriched diets; the carbohydrate diet produced the worst results (lowest specific growth rate, weight gain, nutritional index and hepatosomatic index). These results suggest that it is not advisable to reduce dietary fish protein below 35%, and that it is not possible to obtain a protein-sparing effect of either lipids or carbohydrates, at least in our experimental conditions. The high-protein diet resulted in the storage of energy excess as muscle proteins and hepatic glycogen. Tench fed the high-carbohydrate diet stored carbohydrates as muscle glycogen and reduced plasma triglycerides. Finally, both liver and muscle lipid content were in positive correlation to dietary lipid.     

A comparison of nutrient regulation between solitarious and gregarious phases of the specialist caterpillar, Spodoptera exempta (Walker)

Nutritional regulatory responses were compared between solitarious and gregarious phases of the African armyworm, Spodoptera exempta. When allowed to mix between two nutritionally imbalanced but complementary foods, final-instar caterpillars in both phases selected a diet comprising more carbohydrate than protein. This contrasts with other larval lepidopterans studied to date. Only minor differences were found in the position of the intake target for the two phases, despite their different energetic requirements for migration as adults. When restricted to nutritionally imbalanced diets, caterpillars of both phases were less disposed to overeat protein on high-protein diets than carbohydrate on high-carbohydrate diets, relative to the self-composed intake target. However, in both cases gregarious larvae overingested the excess nutrient to a greater degree than did solitarious larvae. Furthermore, gregarious larvae showed higher nitrogen conversion efficiency on an extreme protein-limiting diet, and accumulated more lipid per amount of carbohydrate consumed on carbohydrate-deficient diets. These phase-associated nutritional differences are consistent with the life-history strategies of the two phases.     

Salivary signals of European corn borer induce indirect defenses in tomato

Plants can recognize the insect elicitors and activate its defense mechanisms. European Corn Borer (ECB; Ostrinia nubilalis) saliva, produced from the labial salivary glands and released through the spinneret, is responsible for inducing direct defenses in host plants. Glucose oxidase (GOX) present in the ECB saliva induced direct defenses in tomato. By contrast, GOX activity in ECB saliva was insufficient to trigger defenses in maize, suggesting that host-specific salivary elicitors are responsible for inducing direct defenses in host plants. Our current study further examined whether ECB saliva can trigger indirect defenses in tomato. Relative expression levels of TERPENE SYNTHASE5 (TPS5) and HYDROPEROXIDE LYASE (HPL), marker for indirect defenses in host plants, were monitored. Quantitative real-time PCR analysis revealed that ECB saliva can induce the expression of TPS5 and HPL, suggesting that salivary signals can induce indirect defenses in addition to the direct defenses. Further experiments are required to identify different ECB elicitors that are responsible for inducing direct and indirect defenses in host plants.     

Dietary carbohydrates induce changes in glucosensing capacity and food intake of rainbow trout

We hypothesize that variations in dietary carbohydrate levels produce changes in glucosensor parameters in previously characterized glucosensing areas (hypothalamus and hindbrain) related with the regulation of food intake of a carnivorous fish species like rainbow trout. Therefore, we fed trout with standard, carbohydrate-free (CF) or high-carbohydrate (HC) diets for 10 days to assess changes in glucosensing system and food intake. Fish fed CF diet displayed hypoglycemia and increased food intake. Fish fed a HC diet displayed hyperglycemia and decreased food intake. Changes in food intake due to dietary carbohydrates were accompanied in hypothalamus and hindbrain of fish fed with HC diet by changes in parameters involved in glucosensing, such as increased glucose, glucose 6-phosphate, and glycogen levels and increased glucokinase (GK), glycogen synthase, and pyruvate kinase activities as well as increased GK and GLUT2 expression. All those results address for the first time in fish, despite the relative intolerance to glucose of carnivorous species, that dietary carbohydrates are important regulators of the glucosensing system in carnivorous fish, suggesting that the information generated by this system can be associated with the changes observed in food intake.     

Autolysis and ageing of Penicillium chrysogenum under carbon starvation: respiration and glucose oxidase production

During the exponential growth phase of Penicillium chrysogenum NCAIM 00237 the effective conversion of glucose and O2 to gluconate and H2O2 by glucose oxidase (GOX) was the most likely source of intracellular ROS measured. In glucose-supplemented autolysing cultures, the increased of intracellular ROS concentration was attributed to respiration in the absence of any significant GOX activity. The induction of GOX and catalase by glucose and H2O2 was clearly age-dependent in P. chrysogenum. In ageing cryptic growth phase cultures, superoxide dismutase and cyanide-resistant respiration were the major elements of antioxidative defence but these activities were insufficient to prevent the progressive accumulation of ROS and the concomitant decrease in cell vitality.     

Determinants of polyphagy by a woolly bear caterpillar: a test of the physiological efficiency hypothesis

The physiological efficiency hypothesis argues that the physiological efficiency of food utilization determines feeding habits of herbivorous insects. Although relatively unsuccessful at explaining dietary specificity, it may explain the food-mixing habit of individually polyphagous herbivores because they may opportunistically increase physiological efficiency by optimizing nutrient balance or diluting toxins in the course of feeding on multiple host-plant species. This study tests two predictions of the physiological efficiency hypothesis with the woolly bear caterpillar, Grammia geneura (Strecker) (Lepidoptera: Arctiidae). Namely, both herbivore performance (survival, developmental rate, pupal mass) and growth efficiency should be better on mixed-plant diets than on single-plant diets. In a series of three laboratory experiments, I found that larval survival and developmental rate on mixed-plant diets were superior to performance on single-plant diets in only one of four cases. In all other cases, mixed-plant diets were either inferior (female pupal mass) to single-plant diets or not detectably different from them. Larval growth efficiency on mixed-plant diets was never superior to that on single-plant diets. In mixed-plant treatments, caterpillars often selected a diet that included plant species of relatively low suitability alone, thereby suffering reduced performance and growth efficiency. These results contradict predictions of the physiological efficiency hypothesis, indicating the limitations of the conventional focus on the physiological constraints on food utilization as an explanation for both individual polyphagy and dietary specificity in herbivorous insects.