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Highlights: [•] Fitness and stability of Cry1Ab resistance were assayed in two strains of Diatraea saccharalis. [•] Resistance in the two strains was not associated with fitness costs on non-toxic diet. [•] Resistance in both strains was stable in absence of selection pressures. [•] The Cry1Ab resistance in the two strains likely had a similar genetic basis. [Copyright &y& Elsevier]

BACKGROUND Larval movement of target pest populations among Bt and non- Bt plants is a major concern in the use of a seed mixture refuge strategy for Bt resistance management. In this study, occurrence and larval movement of the sugarcane borer, Diatraea saccharalis (F.), were evaluated in four planting patterns of non- Bt and Bt plants containing Genuity® SmartStaxTM traits in 2009-2011. The four planting patterns were: (1) a pure stand of 27 Bt plants; (2) one non- Bt plant in the center, surrounded by 26 Bt plants; (3) a pure stand of 27 non- Bt plants; (4) one Bt plant in the center, surrounded by 26 non- Bt plants. Studies were conducted under four conditions: (1) open field with natural infestation; (2) greenhouse with artificial infestations; open field with artificial infestations (3) on the center plants only and (4) on every plant. The major objective of this study was to determine whether refuge plants in a seed mixture strategy could provide a comparable refuge population of D. saccharalis to a 'structured refuge' planting. RESULTS Larvae of D. saccharalis showed the ability to move from infested plants to at least four plants away, as well as to adjacent rows, but the majority remained within the infested row. However, the number of larvae found on the non- Bt plants in the mixture plantings was not significantly reduced compared with the pure stand of non- Bt corn. CONCLUSION The results of this study show that refuge plants in a seed mixture may be able to provide a comparable refuge population of D. saccharalis to a structured refuge planting. © 2013 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Diatraea saccharalis is a major corn borer pest. Midgut serine proteinases are essential for insect growth and development. Alteration of midgut proteinases is responsible for Bt resistance development in some species. To clone midgut trypsin and chymotrypsin cDNAs and to test if the Cry1Ab resistance in D. saccharalis is associated with changes in midgut proteinases, total midgut tryptic and chymotryptic activities, cDNA sequences, and gene expressions of three trypsin and three chymotrypsin genes were comparatively examined between Cry1Ab-susceptible (Cry1Ab-SS) and Cry1Ab-resistant (Cry1Ab-RR) strains. Full-length cDNAs encoding three trypsin- and three chymotrypsin-like proteinases were sequenced from Cry1Ab-SS and Cry1Ab-RR larvae. These cDNAs code for active forms of midgut serine proteinases with all functional motifs, including signal peptide, conserved His-Asp-Ser for the catalytic triad, three pairs of cysteines for disulfide bridge configurations, and conserved substrate specificity determination residues. In general, cDNA and putative protein sequences are highly similar between Cry1Ab-SS and Cry1Ab-RR strains, except for a few nucleotide and predicted amino acid substitutions, whose function need to be further clarified. Total trypsin and chymotrypsin activities were also similar between Cry1Ab-SS and Cry1Ab-RR strains. Transcriptional levels of the trypsin and chymotrypsin genes had numerical difference between Cry1Ab-SS and Cry1Ab-RR strains, but the difference was not statistically significant. Data suggest that the development of Cry1Ab resistance in D. saccharalis was not significantly associated with these trypsins and chymotrypsins. Results clarified the role of six midgut proteinases and provided a foundation for continuing examination of potential involvement of other midgut proteinases in Bt resistance development and other important biochemical processes. [ABSTRACT FROM AUTHOR]

Abstract: Transgenic corn (Zea mays L.) products expressing multiple Bt proteins targeting a same group of insect pests have become commercially available in the U.S. The first commercialized pyramided Bt corn technologies for managing lepidopteran pests include Genuity® VT Triple Pro™ and Genuity® SmartStax™. In this study, larval survival and plant injury of Cry1Ab-susceptible (Cry1Ab-SS), -resistant (Cry1Ab-RR), and -heterozygous (Cry1Ab-RS) genotypes of the sugarcane borer, Diatraea saccharalis F., on five commercial corn hybrids were evaluated in a leaf tissue bioassay and two greenhouse trials during 2010–2011. The five hybrids included two non-Bt corn and three Bt corn hybrids representing three transgenic technologies: YieldGard® Corn Borer, Genuity® VT Triple Pro™, and Genuity® SmartStax™. YieldGard® Corn Borer corn contains a single Bt protein (Cry1Ab), while Genuity® VT Triple Pro™ and SmartStax™ contain two and three pyramided Cry genes targeting above-ground lepidopteran pests, respectively. An average of 75.5% of larvae survived after 6 d on non-Bt corn leaf tissue. The 6-d survivorship on Cry1Ab corn leaf tissue was 3% for Cry1Ab-SS, 19% for Cry1Ab-RS, and 35% for Cry1Ab-RR larvae, while none of the three insect genotypes survived for 6-d on the leaf tissue of the two pyramided Bt corn hybrids. After 21 d on whole plants in the greenhouse, 42.6–62.5% of larvae survived on non-Bt corn plants. Larval survivorship rates on YieldGard® Corn Borer plants after 21 d in the greenhouse were 4.7–5.6% for Cry1Ab-SS, 29.4–32.5 % for Cry1Ab-RS, and 36.6–45.6% for Cry1Ab-RR. Both pyramided Bt corn hybrids were very effective against D. saccharalis regardless of the insect genotype with 21-d survivorship of <2% for Cry1Ab-SS and Cry1Ab-RS, and <5% for Cry1Ab-RR. Cry1Ab-RS and –RR caused significant entry/exit holes and tunneling inside the plant stalks of non-Bt and YieldGard® corn plants, while they caused little injury on the two pyramided Bt corn hybrids. The data generated from both the leaf tissue and whole plant tests showed that the Cry1Ab-resistant D. saccharalis was susceptible to the pyramided Bt corn hybrids. The results suggest that the pyramided Bt corn containing Genuity® VT Triple Pro™ or SmartStax™ traits should offer a means for Cry1Ab resistance management in D. saccharalis. [Copyright &y& Elsevier]

Sugarcane borer, Diatraea saccharalis (F.), is a major target of transgenic maize expressing Bacillus thuringiensis (Bt) proteins in South America and the US mid-south region. Resistance development in target pest populations is a major threat to the sustainable use of Bt crops. In our field trials in 2009, a significant number of live borers and plant injury from D. saccharalis were observed in an experimental SmartStax™ maize line. The objective of this study was to assess the relative susceptibility of two field populations of D. saccharalis collected from non-Bt and Bt maize plants containing SmartStax™ traits to five individual Cry proteins. The five Bt proteins included two proteins (Cry1A.105 and Cry2Ab2) that were expressed in SmartStax™ maize plants and three other common Bt proteins (Cry1Aa, Cry1Ab and Cry1Ac) that were not produced in SmartStax™. Larval mortality and growth inhibition on Bt diet of the fourth generation after field collections were evaluated 7 days after release of neonates on the diet surface. The laboratory bioassays showed that 50% lethal concentration (LC50) values for Cry1A.105 and Cry2Ab2 for the population originated from Bt plants were 3.55- and 1.34-fold greater, respectively, than those of the population collected from non-Bt plants. In contrast, relative to the population from non-Bt plants, the LC50 of the population sampled from Bt plants were 3.85-, 2.5- and 1.64-fold more sensitive to Cry1Aa, Cry1Ab and Cry1Ac, respectively. The results did not provide clear evidence to conclude that the observed field survival of D. saccharalis on Bt plants was associated with increased levels of resistance. [ABSTRACT FROM AUTHOR]

The sugarcane borer, Diatraea saccharalis (F.), is a major target of transgenic maize expressing Bacillus thuringiensis (Bt) proteins in South America and the mid-Southern region of the United States. During 2007-2009, a total of 986 feral individuals of D. saccharalis were collected from maize fields in six locations of Louisiana and Mississippi and examined for resistance to Cry1Ab maize using F1/F2 screens. Major resistance alleles to Cry1Ab maize in the populations sampled from non-Bt maize plants during 2007 and 2008 in Louisiana and 2009 in Mississippi were rare. From a total of 487 individuals collected from three locations in Louisiana in 2007 and 2008, only one individual was identified with major resistance alleles. In addition, no major resistance alleles were detected in 242 individuals collected from three locations in Mississippi in 2009. The frequency of major resistance alleles was estimated to be 0.002 with a 95% CI of 0.00025- 0.0057 for the Louisiana populations and < 0.0061, with 95% probability, for the Mississippi populations. The resistance frequency estimated for the Louisiana populations in 2007 and 2008 was not significantly different from those reported previously for populations sampled in 2004-2006. However, among 200 individuals sampled from non-Bt maize plants in 2009 in Louisiana, six individuals were identified to possess major resistance alleles. The estimated major resistance allele frequency for the populations sampled from non-Bt maize plants in 2009 in Louisiana was 0.0176 with a 95% CI of 0.0072 to 0.0328, which was significantly greater than those estimated for the populations collected in 2004-2008. Similarly, the frequency of minor resistance alleles to Cry1Ab maize for the Louisiana populations collected in 2009 was also significantly greater than those estimated for the populations sampled before. In addition, two out of 57 feral individuals collected from Bt maize plants in Louisiana in 2009 were identified to carry major resistance alleles to Cry1Ab maize. Since 2010, transgenic maize expressing pyramided Bt genes has been planted in the US mid-Southern region and by 2011, pyramided Bt maize has replaced Cry1Ab maize as the dominant Bt maize for managing lepidopteran pests including D. saccharalis. The timely switching from single-gene Cry1Ab maize to the pyramided Bt maize should prevent further increases in Cry1Ab resistance allele frequency and thus ensure the continued success of Bt maize for managing D. saccharalis in the region. [ABSTRACT FROM AUTHOR]

The sugarcane borer, Diatraea saccharalis, is a major target pest of transgenic corn expressing Bacillus thuringiensis (Bt) proteins (i.e., Cry1Ab) in South America and the mid-southern region of the United States. Evolution of insecticide resistance in such target pests is a major threat to the durability of transgenic Bt crops. Understanding the pests' resistance mechanisms will facilitate development of effective strategies for delaying or countering resistance. Alterations in expression of cadherin- and alkaline phosphatase (ALP) have been associated with Bt resistance in several species of pest insects. In this study, neither the activity nor gene regulation of ALP was associated with Cry1Ab resistance in D. saccharalis. Total ALP enzymatic activity was similar between Cry1Ab-susceptible (Cry1Ab-SS) and -resistant (Cry1Ab-RR) strains of D. saccharalis. In addition, expression levels of three ALP genes were also similar between Cry1Ab-SS and -RR, and cDNA sequences did not differ between susceptible and resistant larvae. In contrast, altered expression of a midgut cadherin (DsCAD1) was associated with the Cry1Ab resistance. Whereas cDNA sequences of DsCAD1 were identical between the two strains, the transcript abundance of DsCAD1 was significantly lower in Cry1Ab-RR. To verify the involvement of DsCAD1 in susceptibility to Cry1Ab, RNA interference (RNAi) was employed to knock-down DsCAD1 expression in the susceptible larvae. Down-regulation of DsCAD1 expression by RNAi was functionally correlated with a decrease in Cry1Ab susceptibility. These results suggest that down-regulation of DsCAD1 is associated with resistance to Cry1Ab in D. saccharalis. [ABSTRACT FROM AUTHOR]

Using an F1 screen, 352 feral individuals of the sugarcane borer, Diatraea saccharalis (F.) (Lepidoptera: Crambidae), were examined for the presence of Bacillus thuringiensis (Bt)-resistance alleles. These insects represented four geographical populations collected in central and northeastern Louisiana, USA, and one field population from the Gulf Coast area of Texas, USA, during 2006. The F1 screen used various crosses between field-collected insects and a laboratory strain of Cry1Ab-resistant D. saccharalis, including both reciprocal crosses and group mating. F1 neonates of the crosses were screened for Bt resistance on Bt maize leaf tissue. One field-collected individual of D. saccharalis was shown to have a Bt-resistance allele. Based on Bayesian analysis procedures, the Bt-resistance allele frequency in the five populations of D. saccharalis was 0.0028 with a 95% confidence interval of 0.0003–0.0079. The successful identification of a resistance allele in a field collection of insects suggests that the F1 screening technique could be an effective tool for detecting and monitoring rare Bt-resistance alleles in field populations of D. saccharalis. [ABSTRACT FROM AUTHOR]

Transgenic Bacillus thuringiensis (Bt) corn, Zea mays L., has been widely used to manage a corn borer complex in the mid-southern region of the United States. The sugarcane borer, Diatraea saccharalis (F.) (Lepidoptera: Crambidae), has become a dominant cornstalk boring species in some areas of this region, especially in Louisiana. Therefore, management of sugarcane borer resistance to Bt corn is critical to ensure the long-term sustainability of Bt corn for the region. This study screened 280 two-parent family-lines of sugarcane borer from four geographical populations in Louisiana during 2005 to determine whether Bt resistance allele frequency in sugarcane borer is sufficiently low to meet the rare resistance assumption of the current "high dose/refuge" resistance management strategy for Bt corn. These sugarcane borer family-lines were examined for Bt resistance by using novel F2 screening procedures. No major Bt resistance alleles were detected in these four populations. The estimated frequency of major Bt resistance alleles was <0.0027, with a 95% probability and a detection power of 94%. The estimated minor resistance allele frequency was 0.0063, with a 95% CI of 0.0025-0.0117. During a previous study, a major Bt resistance allele was detected in one individual from 213 family-lines of another Louisiana population of sugarcane borer. Combining these data with the current screen, the frequency of major Bt resistance alleles across the five populations was 0.001, with a 95% credibility interval of 0.0001-0.0028 and a detection power of 95%. Major Bt resistance allele frequencies in Louisiana sugarcane borer populations seem to be low, and they should support the rare resistance allele requirement of the high dose/refuge strategy. [ABSTRACT FROM AUTHOR]

A Louisiana strain of the sugarcane borer, Diatraea saccharalis (F.) (Lepidoptera: Crambidae), was selected for resistance to the CrylAb protein of Bacillus thuringiensis (Bt) by using an F2 screening procedure. Survival of Bt-resistant, -susceptible, and -heterozygous genotypes of sugarcane borer was evaluated on vegetative and reproductive stages of five non-Bt and seven Bt field corn, Zea mays L., hybrids in a greenhouse study. Larval survival was recorded 21 d after infestation of neonates on potted plants. Larval survival across the three sugarcane borer genotypes and five non-Bt corn hybrids after 21 d ranged from 23.6 ± 5.2% (mean ± SEM) to 57.5 ± 5.2%. Mean survival of CrylAb-resistant larvae on vegetative and reproductive plant stages was 12 and 21%, respectively. During the vegetative stages, all seven Bt corn hybrids were highly efficacious against CrylAb- susceptible and -heterozygous genotypes of sugarcane borer, with a larval survival rate of <2% for the Bt-susceptible genotype and ⩽5% for the heterozygotes. However, 8-18% of the heterozygous genotype survived on reproductive stage plants for four of the seven Bt corn hybrids tested. The variation in performance of Bt corn cultivars at vegetative and reproductive growth stages against Cry lAb resistant sugarcane borer suggests differential seasonal expression that may hasten resistance in the field. Bt corn hybrids expressing a "high dose" for European corn borer, Ostrinia nubilalis (Hübner), may not produce a sufficient high dose for the sugarcane borer. [ABSTRACT FROM AUTHOR]

The sugarcane borer, Diatraea saccharalis (F.) (Lepidoptera: Crambidae), strain (F52-3-R) was developed from F3 survivors of a single-pair mating on commercial Cry1Ab Bacillus thuringiensis (Bt) corn plants in the greenhouse. The susceptibility of a Bt-susceptible and the F52-3-R strain of D. saccharalis to trypsin-activated Cry1Ab toxin was determined in a laboratory bioassay. Neonate-stage larvae were fed a meridic diet incorporating Cry1Ab toxin at a concentration range of 0.0625 to 32 µg g−1. Larval mortality, larval weight, and number of surviving larvae that did not gain significant weight (<0.1 mg per larva) were recorded on the 7th day after inoculation. The F52-3-R strain demonstrated a significant level of resistance to the activated Cry1Ab toxin. Larval mortality of the Bt-susceptible strain increased in response to higher concentrations of Cry1Ab toxin, exceeding 75% at 32 µg g−1, whereas mortality of the F52-3-R strain was below 8% across all Cry1Ab concentrations. Using a measure of practical mortality (larvae either died or gained no weight), the median lethal concentration (LC50) of the F52-3-R strain was 102-fold greater than that of the Bt-susceptible insects. Larval growth of both Bt-susceptible and F52-3-R strains was inhibited on Cry1Ab-treated diet, but the inhibition of the F52-3-R strain was significantly less than that of the Bt-susceptible insects. These results confirm that the survival of the F52-3-R strain on commercial Bt corn plants was related to Cry1Ab protein resistance and suggest that this strain may have considerable value in studying resistance management strategies for Bt corn. [ABSTRACT FROM AUTHOR]

A lepidopterous pest complex of southwestern corn borer, Diatraea grandiosella Dyar, and sugarcane borer, Diatraea saccharalis (F.), causes economic damage to field maize, Zea mays L., in the southern United States. Management of natural infestations was evaluated in irrigated and non-irrigated plots of Bacillus thuringiensis (Bt) transgenic maize hybrid expressing the insecticidal Cry1A(b) protein associated with the MON810 transformation event. Bt maize significantly (P ≤ 0.05) reduced the number of adult borer exit holes per plant, frequency of plants with adult exit holes in stalks, and frequency of plants with ear shank and cob tunneling compared to non-Bt maize regardless of irrigation regime. Ear length and grain yields were not significantly (P > 0.05) improved in the Bt hybrid compared to those in the non-Bt hybrid but were either similar or significantly (P ≤ 0.05) greater in irrigated than in non-irrigated maize. As stalk borer infestation persisted, a significant (P ≤ 0.05) increase was observed in the number of adult borer exit holes per stalk and in the number of plants affected with exit holes, ear shank tunneling, and cob tunneling in the irrigated non-Bt maize than in the non-irrigated, non-Bt maize. This suggests that irrigated non-Bt maize fields may offer a favorable environment for increased maize stalk borer infestation and survival. During the two-year study, sugarcane borer abundance increased compared to southwestern corn borer. Larvae removed from plants near crop maturity revealed a three to one predominance of sugarcane borer over southwestern corn borer. This might signal greater selection pressure on sugarcane borer in Bt maize in the southern United States. Current Insecticide Resistance Management (IRM) plans need to consider stalk borer species composition in an area and the differences in Bt susceptibility between sugarcane borer and other maize stalk-borer species to meet the high-dose IRM strategy. [ABSTRACT FROM AUTHOR]