Next‐generation transgenic cotton: pyramiding RNAi and Bt counters insect resistance

Transgenic crops producing insecticidal proteins from the bacterium Bacillus thuringiensis (Bt) are extensively cultivated worldwide. To counter rapidly increasing pest resistance to crops that produce single Bt toxins, transgenic plant ‘pyramids’ producing two or more Bt toxins that kill the same pest have been widely adopted. However, cross‐resistance and antagonism between Bt toxins limit the sustainability of this approach. Here we describe development and testing of the first pyramids of cotton combining protection from a Bt toxin and RNA interference (RNAi). We developed two types of transgenic cotton plants producing double‐stranded RNA (dsRNA) from the global lepidopteran pest Helicoverpa armigera designed to interfere with its metabolism of juvenile hormone (JH). We focused on suppression of JH acid methyltransferase (JHAMT), which is crucial for JH synthesis, and JH‐binding protein (JHBP), which transports JH to organs. In 2015 and 2016, we tested larvae from a Bt‐resistant strain and a related susceptible strain of H. armigera on seven types of cotton: two controls, Bt cotton, two types of RNAi cotton (targeting JHAMT or JHBP) and two pyramids (Bt cotton plus each type of RNAi). Both types of RNAi cotton were effective against Bt‐resistant insects. Bt cotton and RNAi acted independently against the susceptible strain. In computer simulations of conditions in northern China, where millions of farmers grow Bt cotton as well as abundant non‐transgenic host plants of H. armigera, pyramided cotton combining a Bt toxin and RNAi substantially delayed resistance relative to using Bt cotton alone.     

Stay or move: how Bt‐susceptible Helicoverpa armigera neonates behave on Bt cotton plants

Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) larvae occasionally have been reported to survive at management threshold levels in fields of Bollgard II^® cotton, Gossypium hirsutum L. (Malvaceae). The pattern and degree of larval survival is not easily predicted but depends on the ability of first instars to establish on host plants. Experiments were conducted with Bacillus thuringiensis Berliner (Bt)‐susceptible and Bt‐resistant larvae of H. armigera to understand how physiologically Bt‐susceptible H. armigera survive on Bt cotton plants, and examine how their first meal influences survival rates. In assays using cotton plant parts, both strains of larvae displayed similar tendencies to drop‐off specific plant parts of Bt and non‐Bt cotton. However, significantly more Bt‐susceptible larvae dropped off young leaves, mature leaves, and squares of Bt cotton compared to non‐Bt cotton plants. Egg cannibalism significantly improved the survival of Bt‐susceptible H. armigera larvae on Bt cotton plants. Larvae were more likely to eat live aged eggs, than newly laid or dead eggs. Survival significantly improved when larvae cannibalized eggs before feeding on Bt leaves. The behavior of Bt‐susceptible larvae with respect to drop‐off and egg cannibalism may help enhance their survival on Bt cotton plants.     

Fitness of Bt‐resistant cabbage loopers on Bt cotton plants

Development of resistance to the insecticidal toxins from Bacillus thuringiensis (Bt) in insects is the major threat to the continued success of transgenic Bt crops in agriculture. The fitness of Bt‐resistant insects on Bt and non‐Bt plants is a key parameter that determines the development of Bt resistance in insect populations. In this study, a comprehensive analysis of the fitness of Bt‐resistant Trichoplusia ni strains on Bt cotton leaves was conducted. The Bt‐resistant T. ni strains carried two genetically independent mechanisms of resistance to Bt toxins Cry1Ac and Cry2Ab. The effects of the two resistance mechanisms, individually and in combination, on the fitness of the T. ni strains on conventional non‐Bt cotton and on transgenic Bt cotton leaves expressing a single‐toxin Cry1Ac (Bollgard I) or two Bt toxins Cry1Ac and Cry2Ab (Bollgard II) were examined. The presence of Bt toxins in plants reduced the fitness of resistant insects, indicated by decreased net reproductive rate (R₀) and intrinsic rate of increase (r). The reduction in fitness in resistant T. ni on Bollgard II leaves was greater than that on Bollgard I leaves. A 12.4‐day asynchrony of adult emergence between the susceptible T. ni grown on non‐Bt cotton leaves and the dual‐toxin‐resistant T. ni on Bollgard II leaves was observed. Therefore, multitoxin Bt plants not only reduce the probability for T. ni to develop resistance but also strongly reduce the fitness of resistant insects feeding on the plants.     

The effect of Bt maize on Sesamia calamistis in South Africa

Bt maize, Zea mays L. (Poaceae) expressing Cry 1Ab insecticidal proteins was introduced for control of Busseola fusca (Fuller) (Lepidoptera: Noctuidae) and Chilo partellus (Swinhoe) (Lepidoptera: Crambidae) in South Africa after its development for control of crambid borers in North America. In the light of the reportedly lower toxicity of Bt maize to certain Noctuidae borers, the effect of Bt maize was evaluated on Sesamia calamistis (Hampson) (Lepidoptera: Noctuidae). The characteristic larval behaviour of S. calamistis may result in reduced exposure to Bt toxin and subsequent high levels of survival. Larvae do not feed on plant whorls like other borer species but penetrate stems directly from behind leaf sheaths where eggs are laid. Greenhouse and laboratory bioassays were done with three Bt maize hybrids and their iso‐hybrids. ‘Whole plant methods’ were used and potted plants artificially infested with eggs or larvae and survival recorded over time. Larval survival was also determined on different plant parts (whorls, stems, tillers, and ears) over time. Bt maize was shown to be highly toxic to S. calamistis. No larvae survived longer than 12–18 days on Bt maize plants in any of the experiments. Adults did not differentiate between Bt and non‐Bt plants in oviposition choice experiments. Sesamia calamistis is polyphagous and occurs in mixed populations with other borer species with which it shares many parasitoid species in Africa. The ecological impact of local extinction of S. calamistis caused by this highly effective transgenic event is therefore not expected to be great.     

Effect of Cry1F maize on the behavior of susceptible and resistant Spodoptera frugiperda and Ostrinia nubilalis

Understanding the behavior of pests targeted with Bacillus thuringiensis Berliner (Bt) crops is important to define resistance management strategies. Particularly the study of larval movement between plants is important to determine the feasibility of refuge configurations. Exposure to Bt maize, Zea mays L. (Poaceae), has been suggested to increase larval movement in lepidopteran species but few studies have examined the potential for resistance to interact with behavioral responses to Bt toxins. Choice and no‐choice experiments were conducted with Spodoptera frugiperda (JE Smith) (Lepidoptera: Noctuidae) and Ostrinia nubilalis (Hübner) (Lepidoptera: Crambidae) to determine whether Cry1F resistance influences neonate movement. Leaf discs of Cry1F maize and the corresponding isoline were used to characterize behavioral responses. In both experiments, the location (on or off of plant tissues) and mortality of susceptible and Cry1F resistant neonates was recorded for 5 days, but the analysis of larvae location was performed until 7 h. Our results indicated no strong difference between resistant and susceptible phenotypes in S. frugiperda and O. nubilalis, although a small percentage of susceptible neonates in both species abandoned maize tissue expressing Cry1F. However, significant behavioral differences were observed between species. Ostrinia nubilalis exhibited increased movement between leaf discs, whereas S. frugiperda selected plant tissue within the first 30 min and remained on the chosen plant regardless of the presence of Cry1F. Spodoptera frugiperda reduced larval movement may have implications to refuge configuration. This study represents the first step toward understanding the effects of Cry1F resistance on Lepidoptera larval behavior. Information regarding behavioral differences between species could aid in developing better and more flexible resistance management strategies.     

Fitness costs associated with Cry1F resistance in the European corn borer

Crops producing insecticidal toxins derived from the bacterium Bacillus thuringiensis (Bt) are widely planted to manage insect pests. Bt crops can provide an effective tool for pest management; however, the evolution of Bt resistance can diminish this benefit. The European corn borer, Ostrinia nubilalis Hübner, is a significant pest of maize and is widely managed with Bt maize in the Midwest of the United States. When Bt crops are grown in conjunction with non‐Bt refuges, fitness costs of Bt resistance can delay the evolution of resistance. Importantly, fitness costs often vary with ecological factors, including host‐plant genotype and diapause. In this study, we examined fitness costs associated with Cry1F resistance in O. nubilalis when insects were reared on three maize lines. Fitness costs were tested in two experiments. One experiment assessed the fitness costs when Cry1F‐resistant and Cry1F‐susceptible insects were reared on plants as larvae and experienced diapause. The second experiment tested resistant, susceptible and F1 heterozygotes that were reared on plants but did not experience diapause. Despite some evidence of greater adult longevity for Cry1F‐resistant insects, these insects produced fewer fertile eggs than Cry1F‐susceptible insects, and this occurred independent of diapause. Reduced fecundity was not detected among heterozygous individuals, which indicated that this fitness cost was recessive. Additionally, maize lines did not affect the magnitude of this fitness cost. The lower fitness of Cry1F‐resistant O. nubilalis may contribute to the maintenance of Cry1F susceptibility in field populations more than a decade after Cry1F maize was commercialized.     

Diamondback moth oviposition: effects of host plant and herbivory

The oviposition behaviour of Plutella xylostella L. (Lepidoptera: Plutellidae) on Chinese cabbage (Brassica rapa L. Pekinensis, cv. Wombok), canola (Brassica napus L. cv. Thunder TT), and cabbage (Brassica oleracea L. Capitata, cv. sugarloaf) (Brassicaceae) was studied in the laboratory. In no‐choice experiments moths laid most eggs on the stems and lower three leaves of cabbage plants, the lower three leaves of canola plants, but on the upper three leaves of Chinese cabbage plants. The effects of conspecific herbivore damage to foliage could be replicated by mechanical damage. When foliage was damaged, injured cabbage and canola plants were preferred for oviposition over intact conspecifics, whereas injured Chinese cabbage plants were less preferred than intact conspecifics. However, when root tissue was damaged, intact cabbage and canola plants were preferred over injured conspecifics, whereas moths did not discriminate between root‐damaged and intact Chinese cabbage plants. Injury to upper leaves significantly affected the intra‐plant distribution of eggs. In cabbage and canola plants, injury to leaf 6 significantly increased the number of eggs laid on this leaf, resulting in a significant decrease in the number of eggs laid on the lower foliage/stem of plants, whereas in Chinese cabbage plants it significantly decreased the number of eggs laid on leaf 6. Following oviposition on intact plants, neonate larvae established the vast majority of feeding sites on leaves 5–8 in all three host plants, indicating that larvae moved a considerable distance from preferred oviposition sites in cabbage and canola plants. The growth rate of neonates fed on leaf‐6 tissue was significantly greater than that of those fed on leaf‐1 tissue; >90% of larvae completed development when fed exclusively on leaf‐6 tissue but no larvae completed development when fed exclusively on leaf‐1 tissue. The study demonstrates the complex and unpredictable interactions between P. xylostella and its host plants and provides a basis from which we can begin to understand observed distributions of the pest in Brassica crops.     

Survival of Helicoverpa armigera larvae on and Bt toxin expression in various parts of transgenic Bt cotton (Bollgard II) plants

There is no conclusive evidence that Helicoverpa spp. (Lepidoptera: Noctuidae) in Australia have evolved significant levels of resistance to Bollgard II^® cotton (which expresses two Bt toxin genes, cry1Ac and cry2Ab). However, there is evidence of surviving larvae on Bollgard II cotton in the field. The distribution and survival of early‐instar Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) larvae were examined on whole Bollgard II and non‐Bt cotton plants in greenhouse bioassays. The expression of Cry toxins in various parts of Bollgard II plants was compared to the survival of larvae in those locations. Only 1% of larvae survived after 6 days on greenhouse‐grown Bollgard II plants compared to 31% on non‐Bt cotton plants. Overall, and across all time intervals, more larvae survived on reproductive parts (squares, flowers, and bolls) than on vegetative parts (leaves, stems, and petioles) on Bollgard II plants. The concentration of Cry1Ac toxin did not differ between plant structures, whereas Cry2Ab toxin differed significantly, but there was no relationship between the level of expression and the location of larvae. This study provides no evidence that lower expression of Cry toxins in the reproductive parts of plants explains the survival of H. armigera larvae on Bollgard II cotton.     

Sublethal effects of Bt toxin and chlorpyrifos on various Spodoptera exigua populations

Bt cotton (Cry1Ac) has been commercially grown in China since 1997, saving China's cotton production from attack by Bt‐target pests and also tremendously reducing pesticide usage. In recent years, however, Bt cotton, with 4.2 million ha of cultivation, has suffered from a secondary target pest, Spodoptera exigua (Hübner) (Lepidoptera: Noctuidae). In China, growers have even had to re‐adopt conventional pesticides to control the pest, and this practice has already caused serious pesticide residue. In order to clarify the sublethal effects of chemical pesticide, the responses of a Bt‐susceptible and a Bt‐tolerant (Bt10) S. exigua strain to three treatment combinations were examined, including Bt toxin, sublethal chlorpyrifos, and Bt + sublethal chlorpyrifos. The susceptible and the Bt10 strain responded differently to dual pressure. Bt toxin + sublethal chlorpyrifos treatment lowered larval mortality and stimulated population increase of the susceptible S. exigua, whereas it delayed growth and development of the Bt10 strain. Under dual pressure, although larvae of the Bt10 strain developed faster than larvae of the susceptible strain, the Bt10 population experienced higher larval mortality, prolonged pupal duration, decreased pupal weight, decreased emergence rate, and shortened adult longevity. Compared with the susceptible strain, the Bt10 strain was deleteriously affected by sublethal chlorpyrifos. The Bt‐tolerant/resistant S. exigua population was more vulnerable to chemical pesticides like chlorpyrifos regardless of whether it was exposed to Bt toxin or not. Our study provides a reference for increasing the efficacy of control of S. exigua in Bt‐cotton planting areas.     

Effects of adult emergence timing on susceptibility and fitness of Cry3Bb1‐resistant western corn rootworms

Field‐evolved resistance by the western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte to the Cry3Bb1 trait expressed in maize, has been documented in areas of Nebraska USA. Currently, only limited information is available on life‐history traits of Cry3Bb1‐resistant field populations. Therefore, the Gassmann on‐plant bioassay was used to investigate the potential variability among four Cry3Bb1‐resistant WCR field collections made in 2011–2012 by focusing on the key parameters: larval survival, developmental stage and weight with specific emphasis on the impact of adult emergence timing on these parameters in subsequent progeny. Key results: In three of four collections, the susceptibility of larval progeny from adults that emerged early or late within a generation from Cry3Bb1 plants was similar. Each of the three collections exhibited complete resistance; that is, survival on Cry3Bb1 plants was greater or equal to survival on non‐Bt isoline plants. Bioassays from an additional field collection from one site 2 years (2013) after the original collection (2011) (both from Cry3Bb1 maize) indicated that resistance to Cry3Bb1 was maintained over time at the site despite Bt trait rotation in 2012. In general, comparative WCR life‐history parameter data from Cry3Bb1 and isoline maize indicate that fitness of field collections exhibiting complete resistance was similar on each hybrid. The mean proportion of larvae in third instar and mean weight of larvae recovered in bioassays from progeny of early‐ and late‐emerged adults was not significantly affected by emergence period. This suggests that delays in development and associated mean adult emergence commonly observed in populations that are susceptible to Cry3Bb1 may become smaller as populations become resistant to Cry3Bb1. Results from this article will inform Cry3Bb1 resistance mitigation efforts and contribute to the development of sustainable WCR management programmes.     

Oviposition site preference and developmental performance of a gall‐inducing leafhopper on galled and non‐galled host plants

Oviposition preferences of herbivorous insects affect offspring performance. Both positive and negative links between oviposition preference and offspring performance have been reported for many species. A gall‐inducing leafhopper, Cicadulina bipunctata Melichar (Hemiptera: Cicadellidae), feeds on various Poaceae plants and induces galls of enhanced nutritional value for their offspring. Although gall induction by C. bipunctata improves nymphal performance, the oviposition preference of females between galled and non‐galled host plants is still unclear. In this paper, the nymphal performance and oviposition and feeding‐site preference of C. bipunctata were investigated using galled wheat, Triticum aestivum L., and non‐galled barley, Hordeum vulgare L., as host plants. The survival rate of C. bipunctata on wheat was significantly higher than on barley. In the choice test, significantly more eggs were laid into barley, whereas the number of eggs deposited on both hosts was not significantly different in the no‐choice test. The number of settling individuals per leaf area was not significantly different between wheat and barley, suggesting no clear preference for oviposition between these plants. Experience as a nymph with a growing host did not affect oviposition preference as adult female. The inconsistent correspondence between offspring performance and oviposition preference of C. bipunctata may reflect the high mobility of nymphs and/or differences in leaf area between host plants. The results indicate that the previous finding that oviposition preference and offspring performance are not always positively correlated in herbivorous insects is applicable to gall‐inducing insects.     

Quantifying rates of random mating in western corn rootworm emerging from Cry3Bb1‐expressing and refuge maize in field cages

The western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), is the most significant pest of field maize, Zea mays L. (Poaceae), in the USA. Maize plants expressing Bt toxins targeting the corn rootworm complex have been widely adopted and are the primary insecticidal control measure for this pest in North America. Insect resistance management tactics using various refuge structures have been adopted to ensure Bt products will retain durability. An assumption of the refuge strategy is that males and females emerging from Bt and refuge plantings mate randomly; this has not been tested in the field. We conducted cage studies using field populations of WCR in Indiana, USA, to generate empirical field data on mating rates between beetles emerging from Cry3Bb1‐expressing Bt and refuge maize plants. Two refuge configurations were tested; all refuge plants were labeled using the stable isotope ¹⁵N. This mark persists in adult beetles after eclosion, allowing for collection and analysis of isotopic ratios of all beetles. Additional data collected included adult emergence rates, timing and sex ratios for each of the treatments, and head capsule size and dry weights of beetles collected. Treatment had a significant effect on dry weight; mean dry weight decreased in Bt‐only treatments. Fisher's exact test of proportions of mating pairs of refuge and Bt insects indicated that mating was not random in 20% strip refuges and 5% seed blend treatments. We found high percentages of beetles that fed on Bt‐expressing plants as larvae, suggesting that mating between resistant beetles may not be rare even if random mating did occur.     

Antixenosis and larval and adult dispersal in the Mediterranean corn borer,Sesamia nonagrioides,in relation to Bt maize

Sesamia nonagrioides Lefèbvre (Lepidoptera: Noctuidae) is a key pest of maize [Zea mays L. (Poaceae)] and a main target of Bt maize in the Mediterranean area. To choose the most suitable non‐Bt refuge strategy for preventing or delaying resistance development in this maize borer, we examined its biology and behaviour. No antixenotic effects were found on numbers of eggs and egg batches per plant in choice (Bt vs. non‐Bt plants) and no‐choice assays. However, a greater ratio of young larvae dispersed from Bt than from non‐Bt plants. In addition, larvae that hatched on Bt plants tended to disperse more than those that hatched on non‐Bt plants, particularly during young growth stages. Many adults, especially females, could fly at least up to 400 m, as was found in a dispersal study with rubidium‐marked adults. The stimulation of larval dispersal by the Bt trait and the dispersal capacity of adults might compromise the efficacy of seed mixtures as an insecticide resistance management strategy.     

Influence of cabbage resistance and colour upon the diamondback moth and its parasitoid Oomyzus sokolowskii

Host plant resistance and biological control are vital integrated pest management tools against the diamondback moth (DBM), Plutella xylostella (L.) (Lepidoptera: Plutellidae), but to date no study has investigated this system including the DBM parasitoid Oomyzus sokolowskii (Kurdjumov) (Hymenoptera: Eulophidae). We examined oviposition and development of P. xylostella exposed to two commercial cabbage cultivars (green ‘Chato de quintal’ and red ‘Roxo’) and possible effects upon O. sokolowskii. Under free‐choice tests, DBM females laid significantly more eggs on plants of the green cabbage, even though several population growth parameters showed that DBM developed better on the red cabbage. Furthermore, a laboratory free‐choice test with artificially green‐ and red‐painted kale leaf discs demonstrated a similar oviposition preference pattern, with green colour being preferred over red colour. The preference was apparently visually mediated; olfactometer tests showed similar attraction of moths to both green and red cultivars in choice and non‐choice tests. Host plant cultivar had no statistically significant effect on female parasitoid behaviour towards DBM larvae, nor on parasitoid numbers or longevity. Moreover, wasps parasitizing DBM larvae reared on the green cultivar developed more quickly and in larger numbers per parasitized larva. Thus, feeding on green cabbage rather than red does not hinder, and potentially even enhances, control of DBM by O. sokolowskii. On a practical level, these results suggest that intercalating green cabbage cultivars as a trap crop might help protect more profitable red cultivars in growing fields.     

Effects of Bt maize expressing Cry1Ab and Bt spray on Spodoptera littoralis

Various studies have been conducted to assess the damage caused by secondary lepidopteran pests to transgenic Bt maize expressing Cry1Ab. However, to date little is known on the effects of transgenic maize on Spodoptera littoralis (Boisduval) (Lepidoptera: Noctuidae), a polyphagous herbivore which is considered a pest in Mediterranean maize growing areas. Here we present results on the effects of Bt maize (Bt‐11) and Bt spray (Dipel) on the various life stage parameters of this herbivore. We further assess the expression of Cry1Ab in different leaves and leaf parts in maize at a given plant growth stage, and determine whether the feeding damage of 3rd instar S. littoralis is influenced by Bt toxin expression. Contrary to previous literature reporting that S. littoralis is not sensitive to Bt Cry1Ab toxin, our results show that insects fed on either transgenic or Bt sprayed plants were negatively affected. Young S. littoralis larvae (1st and 2nd instars) were found to be the most sensitive to the Bt toxin. This was represented by a higher mortality and a slower developmental time of larvae maintained on transgenic or sprayed plants when compared to insects maintained on control plants. Moreover, Bt maize had a stronger and prolonged detrimental effect on insects when compared to Bt spray in maize. This was revealed by the fact that insects maintained on transgenic plants from 3rd instar to pupation took longer to reach adult emergence compared to insects that were maintained on sprayed plants. This was likely due to the continuous exposure of insects to the toxin when kept on transgenic maize. ELISA results showed a variation in the amount of Bt toxin among different leaf sections in transgenic maize at a given plant growth stage. These differences in Bt toxin were primarily found in the youngest leaf of growing plants. Although the lowest amounts of Bt toxin were detected in the growing leaf section of young leaves, this difference did not appear to influence the feeding behavior of 3rd instar S. littoralis.     

Oviposition acceptance and larval development of Chilo partellus stemborers in drought‐stressed wild and cultivated grasses of East Africa

Maternal host choices during oviposition by herbivorous insects determine the fitness of their offspring and may be influenced by environmental changes that can alter host‐plant quality. This is of particular relevance to ‘push‐pull’ cropping systems where host preferences are exploited to manage insect pest populations. We tested how drought stress in maize and companion plants that are used in these systems affect oviposition preference, larval feeding, and development of the spotted stemborer, Chilo partellus Swinhoe (Lepidoptera: Crambidae). Five host species were tested (all Poaceae): maize (Zea mays L.), Napier grass (Pennisetum purpureum Schumach), signal grass [Brachiaria brizantha (A. Rich) Stapf], Brachiaria cv. ‘Mulato’, and molasses grass [Melinis minutiflora (Beauv.)]. Under drought stress, maize experienced as much oviposition as control unstressed maize in choice and no‐choice experiments. Similarly, larval leaf damage was not significantly different in drought‐stressed vs. unstressed maize. In contrast, oviposition occurred less on drought‐stressed than on unstressed Napier and signal grass. Oviposition acceptance and leaf damage remained low in both drought‐stressed and unstressed molasses grass and Mulato. Larval survival and development remained high in drought‐stressed maize, but not in Napier, signal, and molasses grass and Mulato, where survival and development were low in both drought‐stressed and unstressed plants. Our results indicate that herbivore responses to drought‐stressed plants depend on the plant species and that drought stress can change host preference and acceptance rankings. In particular, trap‐crops such as Napier grass may not divert oviposition from the main maize crop under drought stress conditions.     

Physical leaf defenses – altered by Zea life‐history evolution,domestication, and breeding – mediate oviposition preference of a specialist leafhopper

Plant anti‐herbivore defenses are known to be affected by life‐history evolution, as well as by domestication and breeding in the case of crop species. A suite of plants from the maize genus Zea (Poaceae) and the specialist herbivore Dalbulus maidis (DeLong & Wolcott) (Hemiptera: Cicadellidae) were used to test the hypothesis that anti‐herbivore defenses are affected by plant life‐history evolution and human intervention through domestication and breeding for high yield. The suite of plants included a maize (Zea mays ssp. mays L.) commercial hybrid, a maize landrace, two populations of the annual Balsas teosinte (Z. mays ssp. parviglumis Iltis & Doebley), and perennial teosinte (Z. diploperennis Iltis, Doebley & Guzman). Leaf toughness, pubescence, and oviposition preference were compared among the suite of host plants looking for effects of transitions in life history (i.e., from perennial to annual life cycle), domestication (i.e., from wild annual to domesticated annual), and breeding (i.e., from landrace to hybrid maize) on defense against D. maidis. Results on leaf toughness suggested that the life‐history and domestication transitions weakened the plant's resistance to penetration by the mouthparts and ovipositor of D. maidis, whereas results on pubescence suggested that this putative defense was strengthened with the breeding transition, contrary to expectations. Results on oviposition preference of D. maidis coincided with the expectation that life‐history and domestication transitions would lead to preference for Balsas teosinte over perennial teosinte, and of landrace maize over Balsas teosinte. Also, a negative correlation suggested that oviposition preference is significantly influenced by leaf toughness. Overall, the results suggested that Zea defenses against the specialist herbivore D. maidis were variably affected by plant life‐history evolution, domestication, and breeding, and that chemical defense may play a role in Zea defense against D. maidis because leaf toughness and pubescence only partially explained its host preferences.     

Larval mortality and development for rotation‐resistant and rotation‐susceptible populations of western corn rootworm on Bt corn

The western corn rootworm, Diabrotica virgifera virgifera LeConte, is one of the most economically important insect pests threatening the production of corn, Zea mays (L.), in the United States. Throughout its history, this insect has displayed considerable adaptability by overcoming a variety of pest management tactics, including the cultural practice of annual crop rotation. Since first reported in Illinois in the late 1980s, populations of the rotation‐resistant western corn rootworm have spread over a wide area of the eastern Corn Belt. Currently, little information is available concerning the interaction of rotation resistance with the use of genetically modified corn expressing insecticidal toxins from Bacillus thuringiensis Berliner (Bt), a popular tactic for preventing larval injury and its associated yield loss. The goal of this greenhouse experiment was to determine whether rotation‐resistant and rotation‐susceptible western corn rootworm larvae differ with respect to survival or development when exposed to single‐ or dual‐toxin (pyramided) Bt corn. Individual corn plants were infested with 225 near‐hatch eggs at the V5 (five leaf collar) growth stage. Larvae developed undisturbed on the root systems for 17 days, after which they were recovered using Berlese–Tullgren funnels. Surviving larvae were counted to estimate mortality, and head capsule widths were measured to assess development. Rotation‐resistant and rotation‐susceptible larvae had statistically similar mean levels of mortality and head capsule widths when exposed to both single‐toxin (Cry3Bb1 or Cry34/35Ab1) and pyramided (Cry3Bb1+ Cry34/35Ab1) Bt corn, suggesting that these two populations do not differ with respect to survival or development when exposed to Bt corn. Additionally, the statistically similar mean levels of mortality for larvae exposed to single‐toxin and pyramided Bt corn suggest that pyramided Bt hybrids containing the Cry3Bb1 and Cry34/35Ab1 toxins do not result in additive mortality for western corn rootworm larvae. Implications for management of this economically important pest are discussed.     

Evaluation of Napier grass (Pennisetum purpureum) varieties for use as trap plants for the management of African stemborer (Busseola fusca) in a push–pull strategy

We evaluated eight Napier grass [Pennisetum purpureum Schumach (Poaceae)] varieties, used in various parts of eastern Africa as fodder, for their potential role as trap plants in the management of the African stemborer, Busseola fusca Füller (Lepidoptera: Noctuidae) through a push–pull strategy. Oviposition preference, larval orientation, settling, arrest and dispersal, feeding, mortality and survival, and development were determined for each of these varieties under laboratory and screen house conditions. Two‐choice tests showed that only two of the varieties tested (cv. Bana and cv. Uganda Hairless) were preferentially chosen by gravid female moths for oviposition over a susceptible maize variety, cv. Western Hybrid 502. Larval preference was, however, highly variable. Larval feeding by first instars on the maize leaves was more intense and significantly more than on leaves of all the Napier grass varieties evaluated. Food consumed and amounts assimilated by the third instars over a 24‐h period were not different among larvae fed on stems of maize and those fed on stems of the various Napier grass varieties. Larval survival was significantly lower on all the Napier grass varieties (below 3%) than on maize (about 44%). Similarly, larval development was about 2–3 weeks longer on majority of the Napier grass varieties. It was concluded that cv. Bana had potential for use as a trap plant in the management of B. fusca because it was more preferred by the moths for oviposition, equally preferred as maize by the larvae for orientation, settling, and arrest, and allowed minimal survival of the larvae. It can thus be used with such ‘push’ plants as Desmodium spp. (Fabaceae) in a ‘push–pull’ strategy, but the effectiveness of such a strategy would strictly depend on proper establishment and management of these companion plants.     

Impacts of nitrogen fertilizer on major insect pests and their predators in transgenic Bt rice lines T2A‐1 and T1C‐19

Nitrogen is a critical factor for plant development and nitrogen input is one of the important tactics to enhance the development and yield of crops. Nevertheless, nitrogen input could influence the occurrence of insects positively or negatively. Nitrogen is also one of the main elements composing the insecticidal crystal (Cry) protein. Cry protein production could affect nitrogen partitioning in Bt plants and as such nitrogen input may influence insect pest management in transgenic Bt rice, Oryza sativa L. (Poaceae). To test this possibility, we evaluated the impacts of nitrogen regimes on the main insect pests and their predators on two Bt rice lines, T2A‐1 and T1C‐19, expressing Cry2A and Cry1C, respectively, and their non‐transgenic parental counterpart MH63. The results showed that Cry proteins with different nitrogen regimes have enough insecticidal activity on rice leaffolder, Cnaphalocrocis medinalis Guenée (Lepidoptera: Crambidae), in both laboratory and field experiments. Laboratory studies indicated that relevant parameters of ecological fitness in brown planthopper, Nilaparvata lugens (Stål) (Hemiptera: Delphacidae), a non‐target insect pest, were significantly affected by nitrogen input both on Bt and MH63 rice lines. Nymphal survival, female adult longevity, and egg hatchability in N. lugens differed significantly among rice varieties. The experiments conducted in rice fields also demonstrated that nitrogen was positively correlated with the abundance of N. lugens on Bt rice, similar to that on MH63 rice. The abundances of two predators – the wolf spider Pirata subpiraticus (Boesenberg & Strand) (Araneae: Lycosidae) and the bug Cyrthorhinus lividipennis Reuter (Hemiptera: Miridae) – were significantly affected by rice growth stages but not by nitrogen input and rice varieties. In conclusion, the above results indicate that high nitrogen regimes for Bt rice (T2A‐1 and T1C‐19) and non‐Bt rice (MH63) cannot facilitate the management of insect pests.