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.     

Importance of the experimental setup in research on attractiveness of odours in moths: an example with Busseola fusca

Wind tunnel and Y‐tube olfactometer studies are useful for demonstrating the responsiveness of male moths to sex pheromones. However, in the cereal stem borer, Busseola fusca (Fuller) (Lepidoptera: Noctuidae), wind tunnel bioassays yielded poor results when the behavioural responses of females to plant odours were tested. We demonstrated that for B. fusca females, the Y‐tube olfactometer was better suited for measuring the attractiveness of plant odours, compared with the wind tunnel. In particular, we showed B. fusca preference for odours of a host over a non‐host plant species using this apparatus. Behavioural responses for B. fusca after host finding are also discussed.     

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.     

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.     

Processing of the maize Bt toxin in the gut of Mythimna unipuncta caterpillars

Mythimna unipuncta Haworth (Lepidoptera: Noctuidae) is a well‐known moth species whose larvae can cause devastating damage to some Poaceae crops, including maize (Zea mays L.). The low susceptibility to the Bacillus thuringiensis Berliner (Bt) toxin observed in L6 larvae of this species has been the object of several studies. This study aimed to clarify whether the toxin eliminated from the content of the peritrophic membrane is degraded or excreted and whether the effects of the Bt toxin depend on the doses ingested. To this end, L6 larvae were fed on diets with different amounts of lyophilized Bt or non‐Bt maize leaves. The effect of the Bt concentrations on larval development was measured and the fate of the toxin in the larval tissues was tracked. Results indicated that the larvae of M. unipuncta fed on the various diets showed few differences in weight gain, duration of development, or pupal weight between sublethal Bt concentrations. The larvae rapidly excreted a large part of the toxin ingested, whereas inside the peritrophic membrane the toxin was eliminated, degraded, or sequestered at a rate that increased with the dose and the duration of feeding. As a consequence, little toxin reached the midgut epithelium and therefore the binding sites of the toxin. Moreover, larvae fed on the Bt toxin recovered quickly when they were transferred to a non‐Bt diet.     

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.     

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.     

Mortality impact of MON863 transgenic maize roots on western corn rootworm larvae in the field

Mortality of western corn rootworm (Diabrotica virgifera virgifera LeConte) due to feeding on MON863 transgenic maize (Zea mays L.) expressing the Cry3Bb1 protein was evaluated at three Missouri sites in both 2003 and 2004 and at one site each in South Dakota, Nebraska and Iowa in 2004. To do this, survivorship relative to survivorship on isoline maize (i.e. the same genetic background, but without Cry3Bb1) was evaluated. Comparisons were made using low (1650–2500 eggs/m) and high (3300–3500 eggs/m) western corn rootworm egg densities. Significantly fewer beetles were recovered from MON863 than from isoline maize. Emergence from MON863 as a percentage of viable eggs ranged from 0.02% to 0.10%, whereas percentage emergence from isoline maize ranged from 1.09% to 7.14%. Survivorship on MON863 relative to survivorship on isoline averaged 1.51% when averaged across all environments and both years, so mortality because of the Cry3Bb1 protein averaged 98.49%. The average time delay to 50% cumulative beetle emergence from MON863 was 18.3 days later than from isoline maize. Females comprised 56% and 71% of total beetles recovered from MON863 in 2003 and 2004, respectively. Results are discussed in relation to insect resistance management (IRM) of western corn rootworm.     

Fitness costs of Cry1F resistance in fall armyworm,Spodoptera frugiperda

Transgenic corn, Zea mays L., expressing the Bacillus thuringiensis Berliner (Bt) protein Cry1F has been registered for Spodoptera frugiperda (J. E. Smith) control since 2003 in the USA. Unexpected damage to Cry1F corn was reported in 2006 in Puerto Rico, and Cry1F resistance in S. frugiperda from Puerto Rico was documented. The study of fitness costs associated with insect resistance to Bt insecticidal proteins is important for understanding resistance evolution and for evaluating resistance management practices used to mitigate resistance to transgenic corn. Currently, no studies have addressed the fitness costs associated with Cry1F resistance in S. frugiperda. In this study, susceptible and resistant strains with similar genetic background and their reciprocal crosses were used to estimate Cry1F resistance fitness costs. Comparisons between life‐history traits and population growth rates of homozygous susceptible, heterozygous and homozygous resistant S. frugiperda were used to determine whether the resistance is associated with fitness costs. Major fitness costs were not apparent in either heterozygotes or homozygous resistant insects. However, there was a slight indication of hybrid vigour in the heterozygotes. Additionally, two lines in which the frequency of the resistant alleles was fixed at 0.5 were followed for seven generations, after which the frequency of resistant alleles slightly decreased in both lines. The lack of strong fitness costs associated with Cry1F resistance in S. frugiperda indicates that initial allele frequencies may be higher than expected in field populations and will tend to remain stable in field populations in the absence of selection pressure (e.g. Puerto Rico).     

Resistance monitoring of field populations of the corn borers Sesamia nonagrioides and Ostrinia nubilalis after 5 years of Bt maize cultivation in Spain

Approximately 22 000 hectares (5% of the total maize growing area) of transgenic maize expressing the Cry1Ab toxin from Bacillus thuringiensis (Bt maize) have been planted annually in Spain since 1998. Changes in the susceptibility to Cry1Ab of Spanish populations of the Mediterranean corn borer (MCB), Sesamia nonagrioides (Lefebvre) (Lepidoptera: Noctuidae), and the European corn borer (ECB), Ostrinia nubilalis (Hübner) (Lepidoptera: Crambidae), were assessed by annual monitoring on Bt maize fields. No increase in resistance was detected in the MCB populations from Ebro, Albacete, and Badajoz, nor in the ECB populations from Ebro and Badajoz during the period 1999–2002. The susceptibility of the MCB population from Madrid fluctuated from year to year, but a gradual trend towards higher levels of tolerance was not observed. Laboratory selection assays for eight generations yielded selected strains of MCB and ECB that were 21‐ and 10‐fold significantly more tolerant to Cry1Ab than the corresponding unselected strains, respectively. Nevertheless, none of the field‐collected or laboratory‐selected larvae were able to survive on Bt maize. Considering these data, no consistent shifts in susceptibility were found for Spanish populations of MCB nor ECB after 5 years of Bt maize cultivation, but systematic field monitoring needs to be continued.     

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.     

Rapid selection and characterization of Cry1F resistance in a Brazilian strain of fall armyworm

Transgenic maize (Zea mays L., Poaceae) event TC1507, producing the Cry1F protein of Bacillus thuringiensis Berliner, has been used for management of the fall armyworm, Spodoptera frugiperda (JE Smith) (Lepidoptera: Noctuidae), in Brazil since 2009. A strain of S. frugiperda, obtained from field collections of larvae in TC1507 maize in Minas Gerais state in 2010, was selected in the laboratory for resistance to Cry1F using leaves of TC1507 maize in two selection regimes. Continuous exposure of larvae to Cry1F was more effective than exposure for 6, 8, and 10 days in the selection of resistant S. frugiperda individuals. With only four generations of laboratory selection, a strain with high levels of resistance to Cry1F was obtained, as indicated by the survival of insects reared on leaves of TC1507 maize plants and by the more than 300‐fold resistance level measured in bioassays with the purified Cry1F protein. Importantly, reciprocal crosses between control and the Cry1F‐selected strains revealed that the resistance is autosomal and incompletely recessive, and the response obtained in the backcross of the F₁ generation with the resistant strain was consistent with simple monogenic inheritance. Additionally, there were no apparent fitness costs associated with resistance either for survival or larval growth on non‐Bt maize leaves. Our findings provide experimental evidence for rapid evolution of Cry1F resistance in S. frugiperda in the laboratory and further reinforce the potential of this species to evolve field resistance to the TC1507 maize as previously reported. The resistant strain isolated in this study provides an opportunity to estimate the resistance allele frequency in the field and to determine the biochemical and molecular basis of the resistance, which should provide further information to assist in the resistance management of S. frugiperda on transgenic maize producing B. thuringiensis proteins.     

Evaluation of adult emergence and larval root injury for Cry3Bb1‐resistant populations of the western corn rootworm

The transgenic maize (Zea mays L.) event MON 88017 produces the Bacillus thuringiensis Berliner (Bt) toxin Cry3Bb1 to provide protection from western corn rootworm (Diabrotica virgifera virgifera LeConte) larval feeding. In response to reports of reduced performance of Cry3Bb1‐expressing maize at two locations in Illinois, we conducted a two‐year experiment at these sites to characterize suspected resistance, as well as to evaluate root injury and adult emergence. Single‐plant bioassays were performed on larvae from each population that was suspected to be resistant. Results indicate that these populations had reduced mortality on Cry3Bb1‐expressing maize relative to susceptible control populations. No evidence of cross‐resistance between Cry3Bb1 and Cry34/35Ab1 was documented for the Cry3Bb1‐resistant populations. Field studies were conducted that included treatments with commercially available rootworm Bt hybrids and their corresponding non‐Bt near‐isolines. When compared with their near‐isolines, larval root injury and adult emergence were typically reduced for hybrids expressing Cry34/35Ab1 either alone or in a pyramid. In many instances, larval root injury and adult emergence were not significantly different for hybrids expressing mCry3A or Cry3Bb1 alone when compared with their non‐Bt near‐isolines. These findings suggest that Cry34/35Ab1‐expressing Bt maize may represent a valuable option for maize growers where Cry3Bb1 resistance is either confirmed or suspected. Consistent trends in adult size (head capsule width and dry mass) for individuals recovered from emergence cages were not detected during either year of this experiment. Because of the global importance of transgenic crops for managing insect pests, these results suggest that improved decision‐making for insect resistance management is needed to ensure the durability of Bt maize.     

Effects of three endophytic entomopathogens on sweet sorghum and on the larvae of the stalk borer Sesamia nonagrioides

Various endophytic fungi of native plants and crops are important entomopathogens. The objective of this study was to investigate the entomopathogenic action of Beauveria bassiana (Balsamo) Vuillemin, Metarhizium robertsii (Metchnikoff) Sorokin, and Isaria fumosorosea (Wize) Brown & Smith (all Ascomycota: Hypocreales) against larvae of Sesamia nonagrioides (Lefebvre) (Lepidoptera: Noctuidae) artificially introduced into Sorghum bicolor L. (Moench) (Poaceae) plants under natural environmental conditions. Sorghum bicolor is an economically important crop cultivated for grain, fiber, forage, and lately for biofuel, and S. nonagrioides is its main pest in Mediterranean areas. Young sorghum plants were inoculated with the entomopathogens by spraying in the field. Plant water status, chlorophyll concentration, photosynthesis, and transpiration were not affected. Thirty days after endophyte establishment, plants were infested with fourth instars of S. nonagrioides. The endophytes prevented 50–70% of larvae from entering stalks. Larval mortality was 70–100% and tunnel lengths were reduced by 60–87%. Larval infestation resulted in reduced electron transport capacity and net photosynthetic rate, which was ameliorated in the presence of I. fumosorosea and reversed by B. bassiana and M. robertsii. The growth of sorghum was unaffected in all treatments during the experimental period. Beauveria bassiana and M. robertsii can protect sweet sorghum from damage induced by S. nonagrioides under natural environmental conditions without affecting plant physiology and growth.     

Transportability of non‐target arthropod field data for the use in environmental risk assessment of genetically modified maize in Northern Mexico

In country, non‐target arthropod (NTA) field evaluations are required to comply with the regulatory process for cultivation of genetically modified (GM) maize in Mexico. Two sets of field trials, Experimental Phase and Pilot Phase, were conducted to identify any potential harm of insect‐protected and glyphosate‐tolerant maize (MON‐89Ø34‐3 × MON‐88Ø17‐3 and MON‐89Ø34‐3 × MON‐ØØ6Ø3‐6) and glyphosate‐tolerant maize (MON‐ØØ6Ø3‐6) to local NTAs compared to conventional maize. NTA abundance data were collected at 32 sites, providing high geographic and environmental diversity within maize production areas from four ecological regions (ecoregions) in northern Mexico. The most abundant herbivorous taxa collected included field crickets, corn flea beetles, rootworm beetles, cornsilk flies, aphids, leafhoppers, plant bugs and thrips while the most abundant beneficial taxa captured were soil mites, spiders, predatory ground beetles, rove beetles, springtails (Collembola), predatory earwigs, ladybird beetles, syrphid flies, tachinid flies, minute pirate bugs, parasitic wasps and lacewings. Across the taxa analysed, no statistically significant differences in abundance were detected between GM maize and the conventional maize control for 69 of the 74 comparisons (93.2%) indicating that the single or stacked insect‐protected and herbicide‐tolerant GM traits generally exert no marked adverse effects on the arthropod populations compared with conventional maize. The distribution of taxa observed in this study provides evidence that irrespective of variations in overall biodiversity of a given ecoregion, important herbivore, predatory and parasitic arthropod taxa within the commercial maize agroecosystem are highly similar indicating that relevant data generated in one ecoregion can be transportable for the risk assessment of the same or similar GM crop in another ecoregion.     

Plant defense against fall armyworm in micro‐sympatric maize (Zea mays ssp. mays) and Balsas teosinte (Zea mays ssp. parviglumis)

Maize [Zea mays L. ssp. mays (Poaceae)] was domesticated from Balsas teosinte (Zea mays ssp. parviglumis Iltis & Doebley) in present‐day Mexico. Fall armyworm, Spodoptera frugiperda JE Smith (Lepidoptera: Noctuidae), is among the most important pests of maize in Mexico and Central America. We compared the strength of plant defenses against fall armyworm between micro‐sympatric landrace maize and Balsas teosinte in the field and laboratory. The field comparison, conducted in Mexico, consisted of comparing the frequency of fall armyworm infestation between young maize and Balsas teosinte plants in dryland agricultural fields in which Balsas teosinte grew as a weed. The laboratory comparison contrasted the performance of fall armyworm larvae provided a diet of leaf tissue excised from maize or Balsas teosinte plants that were intact or had been primed by larval feeding. In the field, maize plants were more frequently infested with fall armyworm than Balsas teosinte plants: over 3 years and three fields, maize was infested at a ca. 1.8‐fold greater rate than Balsas teosinte. In the laboratory, larval growth, but not survivorship, was differently affected by feeding on maize vs. Balsas teosinte, and on primed vs. intact plants. Specifically, survivorship was ca. 98%, and did not differ between maize and Balsas teosinte, nor between primed and intact plants. Larvae grew less on intact vs. primed maize, and similarly on intact vs. primed Balsas teosinte; overall, growth was 1.2‐fold greater on maize compared to Balsas teosinte, and on primed compared to intact plants. Parallel observations showed that the differences in growth could not be attributed to the amount of leaf tissue consumed by larvae. We discuss our results in relation to differences in the strength of plant defenses between crops and their ancestors, the relevance of unmanaged Balsas teosinte introgression in the context of fall armyworm defenses in maize, and whether greater growth of larvae on primed vs. intact plants signifies herbivore offense.     

Combining next‐generation sequencing and progeny testing for rapid identification of induced recessive and dominant mutations in maize M2 individuals

Molecular identification of mutant alleles responsible for certain phenotypic alterations is a central goal of genetic analyses. In this study we describe a rapid procedure suitable for the identification of induced recessive and dominant mutations applied to two Zea mays mutants expressing a dwarf and a pale green phenotype, respectively, which were obtained through pollen ethyl methanesulfonate (EMS) mutagenesis. First, without prior backcrossing, induced mutations (single nucleotide polymorphisms, SNPs) segregating in a (M₂) family derived from a heterozygous (M₁) parent were identified using whole‐genome shotgun (WGS) sequencing of a small number of (M₂) individuals with mutant and wild‐type phenotypes. Second, the state of zygosity of the mutation causing the phenotype was determined for each sequenced individual by phenotypic segregation analysis of the self‐pollinated (M₃) offspring. Finally, we filtered for segregating EMS‐induced SNPs whose state of zygosity matched the determined state of zygosity of the mutant locus in each sequenced (M₂) individuals. Through this procedure, combining sequencing of individuals and Mendelian inheritance, three and four SNPs in linkage passed our zygosity filter for the homozygous dwarf and heterozygous pale green mutation, respectively. The dwarf mutation was found to be allelic to the an1 locus and caused by an insertion in the largest exon of the AN1 gene. The pale green mutation affected the nuclear W2 gene and was caused by a non‐synonymous amino acid exchange in encoded chloroplast DNA polymerase with a predicted deleterious effect. This coincided with lower cpDNA levels in pale green plants.     

Comparison of pheromone trap design and lures for Spodoptera frugiperda in Togo and genetic characterization of moths caught

Fall armyworm, Spodoptera frugiperda (JE Smith) (Lepidoptera: Noctuidae), is a pest of grain and vegetable crops endemic to the Western Hemisphere that has recently become widespread in sub‐Saharan Africa and has appeared in India. An important tool for monitoring S. frugiperda in the USA is pheromone trapping, which would be of value for use with African populations. Field experiments were conducted in Togo (West Africa) to compare capture of male fall armyworm using three commercially available pheromone lures and three trap designs. The objectives were to identify optimum trap × lure combinations with respect to sensitivity, specificity, and cost. Almost 400 moths were captured during the experiment. Differences were found in the number of S. frugiperda moths captured in the various trap designs and with the three pheromone lures, and in the number of non‐target moths captured with each lure. The merits of each trap × lure combination are discussed with respect to use in Africa. A nearly equal number of COI‐CS (161) and COI‐RS (158) moths was captured with no differences found in COI marker proportions among traps or lures. However, the diagnostic rice strain marker Tpi was rarely found. Overall, the genetic characterization of the pheromone trap collections indicated a consistent distribution of genetic markers from 2016 to 2017, suggesting a population at or near equilibrium.