Novel genetic basis of resistance to Bt toxin Cry1Ac in Helicoverpa zea.

Crops genetically engineered to produce insecticidal proteins from the bacterium Bacillus thuringiensis have advanced pest management, but their benefits are diminished when pests evolve resistance. Elucidating the genetic basis of pest resistance to Bacillus thuringiensis toxins can improve resistance monitoring, resistance management, and the design of new insecticides. Here, we investigated the genetic basis of resistance to Bacillus thuringiensis toxin Cry1Ac in the lepidopteran Helicoverpa zea, one of the most damaging crop pests in the United States. To facilitate this research, we built the first chromosome-level genome assembly for this species, which has 31 chromosomes containing 375 Mb and 15,482 predicted proteins. Using a genome-wide association study, fine-scale mapping, and RNA-seq, we identified a 250-kb quantitative trait locus on chromosome 13 that was strongly associated with resistance in a strain of Helicoverpa zea that had been selected for resistance in the field and lab. The mutation in this quantitative trait locus contributed to but was not sufficient for resistance, which implies alleles in more than one gene contributed to resistance. This quantitative trait locus contains no genes with a previously reported role in resistance or susceptibility to Bacillus thuringiensis toxins. However, in resistant insects, this quantitative trait locus has a premature stop codon in a kinesin gene, which is a primary candidate as a mutation contributing to resistance. We found no changes in gene sequence or expression consistently associated with resistance for 11 genes previously implicated in lepidopteran resistance to Cry1Ac. Thus, the results reveal a novel and polygenic basis of resistance.

Balancing Bt Toxin Avoidance and Nutrient Intake by Helicoverpa zea (Lepidoptera: Noctuidae) Larvae.

To evaluate how the Cry1Ac Bacillus thuringiensis (Bt) toxin and the ratio of dietary protein to carbohydrate (P:C) independently and jointly affect Helicoverpa zea (Boddie) larval feeding performance and behavior, we conducted no-choice and binary choice experiments using chemically defined diets. We tested two related strains of this polyphagous pest: a strain selected for resistance to Cry1Ac in the laboratory (GA-R), and its field-derived parent strain (GA). In no-choice tests, feeding performance in GA and GA-R was superior on a non-Cry1Ac 80P:20C diet compared to a non-Cry1Ac 35P:65C diet or an 80P:20C diet containing Cry1Ac. Based on consumption and position measurements in binary choice tests, larvae preferred the non-Cry1Ac 80P:20C diet over the other two diets mentioned above. However, the association between preference and performance was weaker when comparing the Cry1Ac 80P:20C diet versus a Cry1Ac 35P:65C diet, and when comparing the Cry1Ac 80P:20C diet versus the non-Cry1Ac 35P:65C diet. In all choice situations, consumption preference occurred mainly by the percentage of larvae that fed almost entirely from one or the other diet, rather than from variation in the extent of diet mixing by individuals. Resistance to Cry1Ac affected the balance between toxin avoidance and nutrient intake: larvae from the more resistant GA-R strain consumed significantly more of the Cry1Ac 80P:20C diet when paired with non-Cry1Ac 35P:65C diet, while GA larvae consumed more of the latter, though not significantly. The results show that dietary P:C ratio, Cry1Ac, and resistance to Cry1Ac affected feeding behavior and performance of H. zea larvae.

Baseline susceptibilities of B- and Q-biotype Bemisia tabaci to anthranilic diamides in Arizona.

BACKGROUND: Development of pyriproxyfen and neonicotinoid resistance in the B-biotype whitefly and recent introduction of the Q biotype have the potential to threaten current whitefly management programs in Arizona. The possibility of integrating the novel anthranilic diamides chlorantraniliprole and cyantraniliprole into the current program to tackle these threats largely depends on whether these compounds have cross-resistance with pyriproxyfen and neonicotinoids in whiteflies. To address this question, the authors bioassayed a susceptible B-biotype strain, a pyriproxyfen-resistant B-biotype strain, four multiply resistant Q-biotype strains and 16 B-biotype field populations from Arizona with a systemic uptake bioassay developed in the present study. RESULTS: The magnitude of variations in LC(50) and LC(99) among the B-biotype populations or the Q-biotype strains was less than fivefold and tenfold, respectively, for both chlorantraniliprole and cyantraniliprole. The Q-biotype strains were relatively more tolerant than the B-biotype populations. No correlations were observed between the LC(50) (or LC(99)) values of the two diamides against the B- and Q-biotype populations tested and their survival rates at a discriminating dose of pyriproxyfen or imidacloprid. CONCLUSION: These results indicate the absence of cross-resistance between the two anthranilic diamides and the currently used neonicotinoids and pyriproxyfen. Future variation in susceptibility of field populations to chlorantraniliprole and cyantraniliprole could be documented according to the baseline susceptibility range of the populations tested in this study.

Extraordinary resistance to insecticides reveals exotic Q biotype of Bemisia tabaci in the New World.

A strain of the whitefly Bemisia tabaci (Gennadius) possessing unusually high levels of resistance to a wide range of insecticides was discovered in 2004 in the course of routine resistance monitoring in Arizona. The multiply resistant insects, collected from poinsettia (Euphorbia pulcherrima Willd. ex Klotzsch) plants purchased at a retail store in Tucson, were subjected to biotype analysis in three laboratories. Polyacrylamide gel electrophoresis of naphthyl esterases and sequencing of the mitochondrial cytochrome oxidase I gene (780 bp) confirmed the first detection of the Q biotype of B. tabaci in the New World. This U.S. Q biotype strain, referred to as Poinsettia'04, was highly resistant to two selective insect growth regulators, pyriproxyfen and buprofezin, and to mixtures of fenpropathrin and acephate. It was also unusually low in susceptibility to the neonicotinoid insecticides imidacloprid, acetamiprid, and thiamethoxam, relative to B biotype whiteflies. In 100 collections of whiteflies made in Arizona cotton (Gossypium spp.), vegetable, and melon (Cucumis melo L.) fields from 2001 to 2005, no Q biotypes were detected. Regions of the United States that were severely impacted by the introduction of the B biotype of B. tabaci in the 1980s would be well advised to promote measures that limit movement of the Q biotype from controlled environments into field systems and to formulate alternatives for managing this multiply-resistant biotype, in the event that it becomes more widely distributed.

Inheritance of resistance to pyriproxyfen in Bemisia tabaci (Hemiptera: Aleyrodidae) males and females (B biotype).

We evaluated effects of the insect growth regulator pyriproxyfen on Bemisia tabaci (Gennadius) (B biotype) (Hemiptera: Aleyrodidae) males and females in laboratory bioassays. Insects were treated with pyriproxyfen as either eggs or nymphs. In all tests, the LC50 for a laboratory-selected resistant strain was at least 620 times greater than for an unselected susceptible strain. When insects were treated as eggs, survival did not differ between males and females of either strain. When insects were treated as nymphs, survival did not differ between susceptible males and susceptible females, but resistant males had higher mortality than resistant females. The dominance of resistance decreased as pyriproxyfen concentration increased. Resistance was partially or completely dominant at the lowest concentration tested and completely recessive at the highest concentration tested. Hybrid female progeny from reciprocal crosses between the susceptible and resistant strains responded alike in bioassays; thus, maternal effects were not evident. Rapid evolution of resistance to pyriproxyfen could occur if individuals in field populations had resistance with traits similar to those of the laboratory-selected strain examined here.