A Novel Reference for Bt-Resistance Mechanism in Plutella xylostella Based on Analysis of the Midgut Transcriptomes

The diamondback moth, Plutella xylostella, is a lepidopteran insect that mainly harms cruciferous vegetables, with strong resistance to a variety of agrochemicals, including Bacillus thuringiensis (Bt) toxins. This study intended to screen genes associated with Bt resistance in P. xylostella by comparing the midgut transcriptome of Cry1Ac-susceptible and -resistant strains together with two toxin-treated strains 24 h before sampling. A total of 12 samples were analyzed by BGISEQ-500, and each sample obtained an average of 6.35 Gb data. Additionally, 3284 differentially expressed genes (DEGs) were identified in susceptible and resistant strains. Among them, five DEGs for cadherin, 14 for aminopeptidase, zero for alkaline phosphatase, 14 for ATP binding cassette transport, and five heat shock proteins were potentially involved in resistance to Cry1Ac in P. xylostella. Furthermore, DEGs associated with “binding”, “catalytic activity”, “cellular process”, “metabolic process”, and “cellular anatomical entity” were more likely to be responsible for resistance to Bt toxin. Thus, together with other omics data, our results will offer prospective genes for the development of Bt resistance, thereby providing a brand new reference for revealing the resistance mechanism to Bt of P. xylostella.

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

Crops genetically engineered to produce insecticidal proteins from the bacterium Bacillus thuringiensis (Bt) have advanced pest management, but their benefits are diminished when pests evolve resistance. Elucidating the genetic basis of pest resistance to Bt toxins can improve resistance monitoring, resistance management, and design of new insecticides. Here, we investigated the genetic basis of resistance to Bt 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. Using a genome-wide association study, fine-scale mapping, and RNA-seq, we identified a 250kb QTL on chromosome 13 that was strongly associated with resistance in a strain of H. zea that had been selected for resistance in the field and lab. This QTL contains no genes with a previously reported role in resistance or susceptibility to Bt toxins. However, within this QTL, we discovered a premature stop codon in a kinesin gene. We hypothesize that this mutation contributes to resistance. The results indicate the mutation on chromosome 13 was necessary but not sufficient for resistance, and therefore conclude that mutations in more than one gene contributed to resistance. Moreover, 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 and extend the list of genes contributing to pest resistance to Bt toxins.

Early Warning of Resistance to Bt Toxin Vip3Aa in Helicoverpa zea

Evolution of resistance by pests can reduce the benefits of crops genetically engineered to produce insecticidal proteins from Bacillus thuringiensis (Bt). Because of the widespread resistance of Helicoverpa zea to crystalline (Cry) Bt toxins in the United States, the vegetative insecticidal protein Vip3Aa is the only Bt toxin produced by Bt corn and cotton that remains effective against some populations of this polyphagous lepidopteran pest. Here we evaluated H. zea resistance to Vip3Aa using diet bioassays to test 42,218 larvae from three lab strains and 71 strains derived from the field during 2016 to 2020 in Arkansas, Louisiana, Mississippi, Tennessee, and Texas. Relative to the least susceptible of the three lab strains tested (BZ), susceptibility to Vip3Aa of the field-derived strains decreased significantly from 2016 to 2020. Relative to another lab strain (TM), 7 of 16 strains derived from the field in 2019 were significantly resistant to Vip3Aa, with up to 13-fold resistance. Susceptibility to Vip3Aa was significantly lower for strains derived from Vip3Aa plants than non-Vip3Aa plants, providing direct evidence of resistance evolving in response to selection by Vip3Aa plants in the field. Together with previously reported data, the results here convey an early warning of field-evolved resistance to Vip3Aa in H. zea that supports calls for urgent action to preserve the efficacy of this toxin.

Study on using gamma radiation to inactivate Bacillus thuringiensis spores in biopesticide

Bacillus thuringiensis (Bt) produces different types of toxin that have potent and specific insecticidal activity. In recent years, Bt toxins have been used as the safe biological control agents to protect crops replacing for chemical insecticides. Bt-based biopesticides that have been commercialized as the alternative products to control pests and insects for sustainable agriculture, contain toxicity crystals and a significant number of spores that affects to the soil microflora. These uncontrollable changes may contaminate the cultivation soil, and eventually cause adverse effects to human and animal health. Therefore, the living cells and spores existing in the Bt-biopesticides should be controlled. This study evaluates the effects of gamma radiation on spore viability, germination and growth of the existing spores after spraying on the soil and the insecticidal effectiveness of a Bt-based biopesticide (VBT) against lepidoptera larvae. We attempted to identify the optimal dose that couldinactivate Bt spores but the toxicity of Bt still retain highly. The results revealed that the dose of 20 kGy is enough to control all living cells and spores in the product that consists of approximately 5.2 × 107 spores in the initial VBT. Though the growth of existing spores after spraying on the soil reduced by 85% or more by irradiation, their insecticidal activity against Heliothis armigera larvae reduced by 20-30% only as compared to that of the initial VBT. It suggested that gamma irradiation can be applied as useful way to control the living cells and spores existing in the commercial Bt-based bio-pesticides, and the radiation dose of 20 kGy is enough to kill all spores in VBT, but still kept its insecticidal effect for Heliothis armigera larvae

Detecting the Conspecific: Herbivory-Induced Olfactory Cues in the Fall Armyworm (Lepidoptera: Noctuidae)

The fall armyworm, Spodoptera frugiperda (Smith), is a polyphagous pest whose larval feeding threatens several economically important crops worldwide with especially severe damage to corn (Zea mays L.). Field-derived resistance to several conventional pesticides and Bt toxins have threatened the efficacy of current management strategies, necessitating the development of alternative pest management methods and technologies. One possible avenue is the use of volatile organic compounds (VOCs) and other secondary metabolites that are produced and sequestered by plants as a response to larval feeding. The effects of conspecific larval feeding on fall armyworm oviposition preferences and larval fitness were examined using two-choice oviposition experiments, larval feeding trials, targeted metabolomics, and VOC analyses. There was a significant preference for oviposition on corn plants that lacked larval feeding damage, and larvae fed tissue from damaged plants exhibited reduced weights and head capsule widths. All larval feeding promoted significantly increased metabolite and VOC concentrations compared to corn plants without any feeding. Metabolite differences were driven primarily by linoleic acid (which is directly toxic to fall armyworm) and tricarboxylic acids. Several VOCs with significantly increased concentrations in damaged corn plants were known oviposition deterrents that warrant further investigation in an integrated pest management context.

Aphicidal activities of Moroccan Bacillus thuringiensis strains against cotton aphid (Aphis gossypii)

The objective of this study was to evaluate the insecticidal effect of toxins from Moroccan Bacillus thuringiensis strains (Berliner) (Bt) on Aphis gossypii (Homoptera: Aphididae). Aphis gossypii is one of the most pests of Moroccan crops. Their management is based traditionally on using chemical products. Some of them are well known to be potentially toxic to the environment and human health. Therefore, alternative strategies for aphid management in crops have been developed in recent years, including a biological control using toxins of bacterial strains. In this study, the artificial diet bioassay was used to screen the aphicidal effect of 82 Bt toxins against first instar nymphs and third instar nymphs of A. gossypii. Among the examined Bt strains, eleven showed a high insecticide activity against A. gossypii stages. In addition, the assessment of the lethal concentration (LC50) of selected Bt revealed that the local BtA4, BtA1 and Bt21.6 exhibited higher insecticidal activity against first instar nymphs of A. gossypii (LC50 (BtA4)=0.15, LC50 (BtA1)=0.23 and LC50 (Bt21.6)=0.25 mg/ml) and the selected strains BtB6, BtA10 and Bt21.6 exhibited the relatively best activity third instar nymphs of A. gossypii (LC50 (BtB6)= 0.48, LC50 (BtA10)= 0.79 and LC50 (Bt21.6)= 1.14 mg/ml) of A. gossypii. Therefore, the results of this study indicate that the selected B. thuringiensis strains have great potential to be used in the integrated A. gossypii management.

Synergic Interaction Between Permethrin and Bt Toxins Discovered Using RNA-Seq

BACKGROUND: Acting against the development of resistance to antibiotics and insecticides, involving negatively correlated cross-resistance (NCR) is an alternative to use- and-discard approach. It is termed NCR that toxic chemicals interact with each other and resistance of target organisms to one chemical is sometimes associated with increased susceptibility to a second chemical when; an allele confers resistance to one toxic chemical and hyper-susceptibility to another, NCR occurs. However, only 11 toxin pairs have been revealed to cause NCR in insects. Finding novel NCRs is needed for integrated pest management. RESULTS: We analyzed permethrin, an insecticide, induced transcriptomes of cultured fat bodies of the silkworm Bombyx mori, a lepidopteran model insect. Differentially expressed gene analyses suggested Bacillus thuringiensis (Bt) toxin was an NCR toxin of permethrin. NCR to permethrin and Bt toxins in Thysanoplusia intermixta, the agricultural pest moth, was examined; the children of permethrin survivor T. intermixta had increased susceptibility to Bt toxin. CONCLUSIONS: A novel NCR toxin pair, permethrin and Bt toxin, was discovered. The screening and developmental method for negatively correlated cross-resistance toxins established in this study was effective, in vitro screening using model organisms and in vivo verification using agricultural pests.