R. Dewey Lee

EVALUATION OF YIELDGARD TRANSGENIC RESISTANCE FOR CONTROL OF FALL ARMYWORM AND CORN EARWORM (LEPIDOPTERA: NOCTUIDAE) ON CORN

Fall armyworm, Spodoptera frugiperda (J. E. Smith), and corn earworm, Helicoverpa zea Boddie, perennially cause leaf and ear damage to corn in the southeastern USA. Development of transgenic hybrids expressing insecticidal endotoxin from Bacillus thuringiensis (Bt) offers a new approach to managing these insects in field corn. Transgenic Bt hybrids with either the Bt11 or MON810 event, collectively known as YieldGard Technology, were evaluated for control fall armyworm and corn earworm in southern Georgia during 1998, which coincided with a severe outbreak of fall armyworm. YieldGard Bt resistance consistently reduced whorl infestation and damage to low levels and also reduced ear infestations and larval numbers per ear. However, larval establishment did occur on many ears of resistant plants, but once established in ears, larvae of both species developed more slowly and caused much less kernel damage on resistant than susceptible plants. We found no relationship between YieldGard Bt resistance and corn grain aflatoxin concentrations. Yield responses were variable with the prevention of yield loss being proportional to the severity of insect damage. These results indicate that YieldGard resistance is effective in preventing significant losses to field corn by fall armyworm and corn earworm. Further, evaluation under a variety of growing conditions and insect infestation levels is needed to clearly assess the value of YieldGard technology to corn growers in the Southeast.

Monitoring the Expression of Maize Genes in Developing Kernels under Drought Stress using Oligo‐microarray

Preharvest aflatoxin contamination of grain grown on the US southeastern Coast Plain is provoked and aggravated by abiotic stress. The primary abiotic stress is drought along with high temperatures. The objectives of the present study were to monitor gene expression in developing kernels in response to drought stress and to identify drought-responsive genes for possible use in germplasm assessment. The maize breeding line Tex6 was used, and gene expression profiles were analyzed in developing kernels under drought stress verses well-watered conditions at the stages of 25, 30, 35, 40, 45 d after pollination (DAP) using the 70 mer maize oligo-arrays. A total of 9 573 positive array spots were detected with unique gene IDs, and 7 988 were common in both stressed and well-watered samples. Expression patterns of some genes in several stress response-associated pathways, including abscisic acid, jasmonic acid and phenylalanine ammonia-lyase, were examined, and these specific genes were responsive to drought stress positively. Real-time quantitative polymerase chain reaction validated microarray expression data. The comparison between Tex6 and B73 revealed that there were significant differences in specific gene expression, patterns and levels. Several defense-related genes had been downregulated, even though some defense-related or drought responsive genes were upregulated at the later stages.

Plant-Incorporated Bacillus thuringiensis Resistance for Control of Fall Armyworm and Corn Earworm (Lepidoptera: Noctuidae) in Corn

Abstract Fall armyworm, Spodoptera frugiperda (J.E. Smith), and corn earworm, Helicoverpa zea (Boddie), perennially cause leaf and ear damage to corn, Zea mays L., in the southeastern United States. Transgenic Bacillus thuringiensis (Bt) hybrids with the Bt11, MON810, or 176 events expressing the Cry1Ab insecticidal endotoxin from were evaluated for control fall armyworm and corn earworm at seven locations in Georgia during 1999 and 2000. Corn was planted at the recommended time for each location and 1 and 2 mo later in the southern locations. All Bt events consistently reduced whorl infestation and damage, although event 176 did not prevent whorl damage in the later plantings in the southern locations in both years. All events also reduced seedling damage by the lesser cornstalk borer, Elasmopalpus lignosellus (Zeller), in one trial and stalk infestations and tunnel length by southwestern corn borers, Diatraea grandiosella Dyar, in another trial. Hybrids containing Bt11 and MON810 events reduced ear infestations in all trials, although reductions were small in later plantings. Nevertheless, both events reduced grain damage from earworms and armyworms by an average ± SE of 52.5 ± 5.1% in all trials. The hybrid containing event 176 did not reduce ear infestations and damage. Total grain aflatoxin concentrations were not significantly affected by Bt resistance in any trial (N = 17). Yield responses were variable with the prevention of yield loss being proportional to the severity of insect damage. Although plantings made after the recommended time did not consistently benefit from Bt resistance, Bt11 and MON810 events were effective in reducing damage to field corn when large infestations occurred. The Bt11 and MON810 events mitigated the risk of severe lepidopteran damage to corn, thereby making later plantings of corn feasible in double-cropping systems.

Impact of Brown Stink Bug (Heteroptera: Pentatomidae) Feeding on Corn Grain Yield Components and Quality

ABSTRACT Brown stink bug, Euschistus servus (Say) (Heteroptera: Pentatomidae), damage on developing corn, Zea mays L., ears was examined in 2005 and 2006 by using eight parameters related to its yield and kernel quality. Stink bug infestations were initiated when the corn plants were at tasseling (VT), mid-silking (R1), and blister (R2) stages by using zero, three, and six in 2005 or zero, one, two, and four bugs per ear in 2006, and maintained for 9 d. The percentage of discolored kernels was affected by stink bug number in both years, but not always affected by plant growth stage. The growth stage effect on the percentage of discolored kernels was significant in 2006, but not in 2005. The percentage of aborted kernels was affected by both stink bug number and plant growth stage in 2005 but not in 2006. Kernel weight was significantly reduced when three E. servus adults were confined on a corn ear at stage VT or R1 for 9 d in 2005, whereas one or two adults per ear resulted in no kernel weight loss, but four E. servus adults did cause significant kernel weight loss at stage VT in 2006. Stink bug feeding injury at stage R2 did not affect kernel damage, ear weight or grain weight in either year. The infestation duration (9 or 18 d) was positively correlated to the percentage of discolored kernels but did not affect kernel or ear weight. Based on the regression equations between the kernel weight and stink bug number, the gain threshold or economic injury level should be 0.5 bugs per ear for 9 d at stage VT and less for stage R1. This information will be useful in developing management guidelines for stink bugs in field corn during ear formation and early grain filling stages.

Root Morphology and Gene Expression Analysis in Response to Drought Stress in Maize (Zea mays)

Water-deficit stress tolerance is a complex trait, and water deficit results in various physiological and chemical changes in maize (Zea mays L.) and exacerbates pre-harvest aflatoxin contamination. The objective of this study was to characterize the variations in morphology, physiology, and gene expression in two contrasting inbred lines, Lo964 and Lo1016, in order to understand the differences in response to water-deficit stress. The results revealed that Lo964 was less sensitive to water-deficit stress, and had a strong lateral root system and a higher root/shoot ratio in comparison to Lo1016. In response to water-deficit stress by comparing stressed versus well-watered conditions, abscisic acid syntheses were increased in leaves, roots, and kernels of both Lo964 and Lo1016, but by different magnitudes. Indole-3-acetic acid (IAA) was undetectable in the leaves and roots of either genotype regardless of treatments, but increases of 58% and 8% in IAA concentration were observed in 20 DAP kernels, in response to water-deficit stress, respectively. The expression of the MIPS was up-regulated 7-fold in leaf tissues of Lo964 compared to Lo1016 at watered conditions, but decreased significantly to similar levels in both genotypes at water-deficit conditions. ZmPR10 and ZmFer1 expressions tended to up-regulate although ZmPR10 was expressed higher in root tissue while ZmFer1 was expressed higher in leaf tissue. Further study is needed to confirm if Lo964 has reduced aflatoxin contamination associated with the drought tolerance in the field in order to utilize the resistant trait in breeding.

Spatial patterns of aflatoxin levels in relation to ear-feeding insect damage in pre-harvest corn.

Key impediments to increased corn yield and quality in the southeastern US coastal plain region are damage by ear-feeding insects and aflatoxin contamination caused by infection of Aspergillus flavus. Key ear-feeding insects are corn earworm, Helicoverpa zea, fall armyworm, Spodoptera frugiperda, maize weevil, Sitophilus zeamais, and brown stink bug, Euschistus servus. In 2006 and 2007, aflatoxin contamination and insect damage were sampled before harvest in three 0.4-hectare corn fields using a grid sampling method. The feeding damage by each of ear/kernel-feeding insects (i.e., corn earworm/fall armyworm damage on the silk/cob, and discoloration of corn kernels by stink bugs), and maize weevil population were assessed at each grid point with five ears. The spatial distribution pattern of aflatoxin contamination was also assessed using the corn samples collected at each sampling point. Aflatoxin level was correlated to the number of maize weevils and stink bug-discolored kernels, but not closely correlated to either husk coverage or corn earworm damage. Contour maps of the maize weevil populations, stink bug-damaged kernels, and aflatoxin levels exhibited an aggregated distribution pattern with a strong edge effect on all three parameters. The separation of silk- and cob-feeding insects from kernel-feeding insects, as well as chewing (i.e., the corn earworm and maize weevil) and piercing-sucking insects (i.e., the stink bugs) and their damage in relation to aflatoxin accumulation is economically important. Both theoretic and applied ramifications of this study were discussed by proposing a hypothesis on the underlying mechanisms of the aggregated distribution patterns and strong edge effect of insect damage and aflatoxin contamination, and by discussing possible management tactics for aflatoxin reduction by proper management of kernel-feeding insects. Future directions on basic and applied research related to aflatoxin contamination are also discussed.

Field Screening of Experimental Corn Hybrids and Inbred Lines for Multiple Ear-Feeding Insect Resistance

Abstract Identifying and using native insect resistance genes is the core of integrated pest management. In this study, 10 experimental corn, Zea mays L., hybrids and 10 inbred lines were screened for resistance to major ear-feeding insects in the southeastern Coastal Plain region of the United States during 2004 and 2005. Ear-feeding insect damage was assessed at harvest by visual damage rating for the corn earworm, Helicoverpa zea (Boddie), and by the percentage of kernels damaged by the maize weevil, Sitophilus zeamais Motschulsky, and stink bugs [combination of Euschistus servus (Say) and southern green stink bug, Nezara viridula (L.)]. Among the eight inbred lines and two control populations examined, C3S1B73-5b was resistant to corn earworm, maize weevil, and stink bugs. In contrast, C3S1B73-4 was resistant to corn earworm and stink bugs, but not to maize weevil. In a similar manner, the corn hybrid S1W*CML343 was resistant to all three ear-feeding insects, whereas hybrid C3S1B73-3*Tx205 was resistant to corn earworm and maize weevil in both growing seasons, but susceptible to stink bugs in 2005. The silk-feeding bioassay showed that corn earworm developed better on corn silk than did fall armyworm. Among all phenotypic traits examined (i.e., corn ear size, husk extension, and husk tightness), only corn ear size was negatively correlated to corn earworm damage in the inbred lines examined, whereas only husk extension (i.e., coverage) was negatively correlated to both corn earworm and maize weevil damage on the experimental hybrids examined. Such information could be used to establish a baseline for developing agronomically elite corn germplasm that confers multiple ear-feeding insect resistance.

Oxidative stress and carbon metabolism influence Aspergillus flavus transcriptome composition and secondary metabolite production

Contamination of crops with aflatoxin is a serious global threat to food safety. Aflatoxin production by Aspergillus flavus is exacerbated by drought stress in the field and by oxidative stress in vitro. We examined transcriptomes of three toxigenic and three atoxigenic isolates of A. flavus in aflatoxin conducive and non-conducive media with varying levels of H2O2 to investigate the relationship of secondary metabolite production, carbon source, and oxidative stress. We found that toxigenic and atoxigenic isolates employ distinct mechanisms to remediate oxidative damage, and that carbon source affected the isolates’ expression profiles. Iron metabolism, monooxygenases, and secondary metabolism appeared to participate in isolate oxidative responses. The results suggest that aflatoxin and aflatrem biosynthesis may remediate oxidative stress by consuming excess oxygen and that kojic acid production may limit iron-mediated, non-enzymatic generation of reactive oxygen species. Together, secondary metabolite production may enhance A. flavus stress tolerance, and may be reduced by enhancing host plant tissue antioxidant capacity though genetic improvement by breeding selection.

Evaluation of spatial and temporal patterns of insect damage and aflatoxin level in the pre-harvest corn fields to improve management tactics

Spatial and temporal patterns of insect damage in relation to aflatoxin contamination in a corn field with plants of uniform genetic background are not well understood. After previous examination of spatial patterns of insect damage and aflatoxin in pre‐harvest corn fields, we further examined both spatial and temporal patterns of cob‐ and kernel‐feeding insect damage, and aflatoxin level with two samplings at pre‐harvest in 2008 and 2009. The feeding damage by each of the ear/kernel‐feeding insects (i.e., corn earworm/fall armyworm damage on the silk/cob, and discoloration of corn kernels by stink bugs) and maize weevil population were assessed at each grid point with five ears. Sampling data showed a field edge effect in both insect damage and aflatoxin contamination in both years. Maize weevils tended toward an aggregated distribution more frequently than either corn earworm or stink bug damage in both years. The frequency of detecting aggregated distribution for aflatoxin level was less than any of the insect damage assessments. Stink bug damage and maize weevil number were more closely associated with aflatoxin level than was corn earworm damage. In addition, the indices of spatial–temporal association (χ) demonstrated that the number of maize weevils was associated between the first (4 weeks pre‐harvest) and second (1 week pre‐harvest) samplings in both years on all fields. In contrast, corn earworm damage between the first and second samplings from the field on the Belflower Farm, and aflatoxin level and corn earworm damage from the field on the Lang Farm were dissociated in 2009.

Insect-attracting and antimicrobial properties of antifreeze for monitoring insect pests and natural enemies in stored corn.

Abstract Insect infestations in stored grain cause extensive damage worldwide. Storage insect pests, including the Indianmeal moth, Plodia interpunctella (Hübner) (Lepidoptera: Pyralidae); Sitophilus spp. (Coleoptera: Curculionidae); and their natural enemies [e.g., Cephalonomia tarsalis (Ashmead) (Hymenoptera: Bethylidae), and Anisopteromalus calandrae (Howard) (Hymenoptera: Pteromalidae)] inhabit a temporary, but stable ecosystem with constant environmental conditions. The objective of the present experiment was to assess the efficacy of using ethylene glycol antifreeze in combination with nutrient solutions to monitor storage insect pest and natural enemy populations in three bins of corn, Zea mays L. The treatments were deionized water, a diluted (1:5 antifreeze:water) antifreeze solution, 10% honey, 10% honey in the diluted antifreeze solution, 10% beer in the diluted antifreeze solution, 10% sucrose in the diluted antifreeze solution, and a commercial pheromone trap suspended in a 3.8-liter container filled with 300-ml of diluted antifreeze solution. The seven treatments captured storage insect pests and their natural enemies in the bins at 33–36°C and 51–55% RH. The pheromone trap in the container with the diluted antifreeze captured significantly more P. interpunctella than the other treatments, but a lower percentage (7.6%) of these captures were females compared with the rest of the treatments (>40% females). All trapping solutions also captured Sitophilus spp. and other beetle species, but the captures of the coleopteran pests were not significantly different among the seven treatments (P > 0.05). Two parasitoid wasps also were captured in the study. The number of A. calandrae was different among the seven treatments (P < 0.05), whereas the number of C. tarsalis was not different among the treatments (P > 0.05). Most A. calandrae adults were captured by the 10% honey in the diluted antifreeze, whereas the fewest were captured in the deionized water. Microbial growth was observed in the 10% honey solution, but no microbial growth occurred in the rest of the treatments, including 10% honey in the diluted antifreeze solution. The results of insect captures and microbial growth demonstrated that antifreeze could be used as a part of storage insect monitoring and/or control programs.