Insecticidal double-stranded RNAs (dsRNAs) silence expression of vital genes by activating the RNA interference (RNAi) mechanism in insect cells. Despite high commercial interest in insecticidal ...dsRNA, information on resistance to dsRNA is scarce, particularly for dsRNA products with non-transgenic delivery (ex. foliar/topical application) nearing regulatory review. We report the development of the CEAS 300 population of Colorado potato beetle (Leptinotarsa decemlineata Say) (Coleoptera: Chrysomelidae) with > 11,100-fold resistance to a dsRNA targeting the V-ATPase subunit A gene after nine episodes of selection using non-transgenic delivery by foliar coating. Resistance was associated with lack of target gene down-regulation in CEAS 300 larvae and cross-resistance to another dsRNA target (COPI β; Coatomer subunit beta). In contrast, CEAS 300 larvae showed very low (~ 4-fold) reduced susceptibility to the Cry3Aa insecticidal protein from Bacillus thuringiensis. Resistance to dsRNA in CEAS 300 is transmitted as an autosomal recessive trait and is polygenic. These data represent the first documented case of resistance in an insect pest with high pesticide resistance potential using dsRNA delivered through non-transgenic techniques. Information on the genetics of resistance and availability of dsRNA-resistant L. decemlineata guide the design of resistance management tools and allow research to identify resistance alleles and estimate resistance risks.
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•Phytophagy is linked to the diversification of gene families responsible for detoxification of xenobiotics in herbivorous arthropods.•Arthropod adaptations to chemical defenses of ...their host plants are similar to arthropod adaptations to synthetic pesticides.•Insecticides may change expression of particular subfamilies of detoxification genes that have evolved due to ancestral associations with host plants.•Treating insecticide resistance as an evolutionary process will increase long-term sustainability of chemical pest control.
Insecticide resistance is a serious economic problem that jeopardizes sustainability of chemical control of herbivorous insects and related arthropods. It can be viewed as a specific case of adaptation to toxic chemicals, which has been driven in large part, but not exclusively, by the necessity for insect pests to tolerate defensive compounds produced by their host plants. Synthetic insecticides may simply change expression of specific sets of detoxification genes that have evolved due to ancestral associations with host plants. Feeding on host plants with more abundant or novel secondary metabolites has even been shown to prime insect herbivores to tolerate pesticides. Clear understanding of basic evolutionary processes is important for achieving lasting success in managing herbivorous arthropods.
Colorado potato beetle (CPB,
Leptinotarsa decemlineata
) is a major pest of potato and other solanaceous vegetables in the Northern Hemisphere. The insect feeds on leaves and can completely defoliate ...crops. Because of the repeated use of single insecticide classes without rotating active ingredients, many chemicals are no longer effective in controlling CPB. Ledprona is a sprayable double-stranded RNA biopesticide with a new mode of action that triggers the RNA interference pathway. Laboratory assays with second instar larvae fed Ledprona showed a dose–response where 25×10
−6
g/L of dsPSMB5 caused 90% mortality after 6days of initial exposure. We also showed that exposure to Ledprona for 6h caused larval mortality and decreased target messenger RNA (mRNA) expression. Decrease in PSMB5 protein levels was observed after 48h of larval exposure to Ledprona. Both PSMB5 mRNA and protein levels did not recover over time. Ledprona efficacy was demonstrated in a whole plant greenhouse trial and performed similarly to spinosad. Ledprona, currently pending registration at EPA, represents a new biopesticide class integrated pest management and insecticide resistance management programs directed against CPB.
Larvae of black soldier flies, Hermetia illucens, are increasingly used for biological conversion of animal and plant wastes into ingredients of animal feeds on an industrial scale. The presence of ...pathogenic microorganisms in harvested larvae may be a serious problem for wide-scale adoption of this technology. Fortunately, black soldier fly larvae may have some antimicrobial properties. Methicillin-resistant Staphylococcus aureus (MRSA) is a bacterium associated with various environments that can be pathogenic to humans and farmed animals. We tested whether black soldier fly larvae suppress MRSA on potato substrate. Autoclaved potatoes containing black soldier fly larvae (P+BSFL), potatoes inoculated with MRSA and containing black soldier fly larvae (P+MRSA+BSFL), and potatoes inoculated with MRSA (P+MRSA) were incubated in glass jars. Substrate samples were taken after 3 and 7 days of incubation and plated on Trypticase soy agar (TSA) and Staphylococcus medium 110 agar (SA) to quantify total bacteria and MRSA, respectively. DNA was extracted from potato substrates on both days and sequenced to assess bacterial and fungal diversity using 515F/806R and internal transcribed spacer (ITS) 1/2 primers, respectively, and QIIME 2.0 software. Both total bacterial and MRSA-specific CFU were reduced in the presence of black soldier fly larvae, with a larger reduction for the latter. Twenty-five bacterial genera and 3 fungal genera were detected. Twenty bacterial genera were shared among the treatments and the days, but their relative abundances often varied. Among the most abundant genera, only
and
were universally present. Our findings confirm antimicrobial properties of black soldier fly larvae.
Larvae of black soldier flies, Hermetia illucens, may be used to provide an environmentally sustainable and economically viable method for biological conversion of animal and plant wastes into ingredients of animal feeds on an industrial scale. However, contamination of harvested larvae by pathogenic microorganisms inhabiting decaying substrates may be a serious problem for wide-scale adoption of this technology. Fortunately, black soldier fly larvae may have some antimicrobial properties, including suppression of several common pathogens. Our study showed that such a suppression applies to methicillin-resistant Staphylococcus aureus, which is a ubiquitous bacterium pathogenic to animals (including humans).
Arthropod Glutathione S-transferases (GSTs) constitute a large family of multifunctional enzymes that are mainly associated with xenobiotic or stress adaptation. GST-mediated xenobiotic adaptation ...takes place through direct metabolism or sequestration of xenobiotics, and/or indirectly by providing protection against oxidative stress induced by xenobiotic exposure. To date, the roles of GSTs in xenobiotic adaptation in the Colorado potato beetle (CPB), a notorious agricultural pest of plants within Solanaceae, have not been well studied. Here, we functionally expressed and characterized an unclassified-class GST, LdGSTu1. The three-dimensional structure of the LdGSTu1 was solved with a resolution up to 1.8 Å by X-ray crystallography. The signature motif VSDGPPSL was identified in the "G-site", and it contains the catalytically active residue Ser14. Recombinant LdGSTu1 was used to determine enzyme activity and kinetic parameters using 1-chloro-2, 4-dinitrobenzene (CDNB), GSH, p-nitrophenyl acetate (PNA) as substrates. The enzyme kinetic parameters and enzyme-substrate interaction studies demonstrated that LdGSTu1 could catalyze the conjugation of GSH to both CDNB and PNA, with a higher turnover number for CDNB than PNA. The LdGSTu1 enzyme inhibition assays demonstrated that the enzymatic conjugation of GSH to CDNB was inhibited by multiple pesticides, suggesting a potential function of LdGSTu1 in xenobiotic adaptation.
Agricultural systems are often simultaneously impacted by multiple stressors. In our study, we investigated the effects of two commonly occurring viruses of potato, Potato virus Y (PVY) and Potato ...leafroll virus (PLRV) on host plant selection and utilization across various spatial scales by the Colorado potato beetle. Adult beetles selected PLRV-infected foliage in the cage choice trails, and larvae that fed exclusively on PLRV-infected foliage developed into larger adults. PVY-infected foliage was avoided in the cage trials and had no effect on the weight of developing adults. In field trials, preference and utilization of potato plants was more dependent on plant size than on virus infection. These results demonstrate that while virus-infected plants can cause physiological changes that influence beetle preference, beetle abundance and/or distribution is driven by a more complex array of factors at the field level. Additionally, this study demonstrates the importance of investigating the applicability of laboratory findings at the field level.
Two-spotted spider mite, Tetranychus urticae Koch, is a ubiquitous and highly polyphagous plant pest with a worldwide distribution. Diapause is essential for its survival in the areas with ...unfavourable seasonal conditions. We investigated whether female mites overwintering close to the soil surface may be adapted to seasonal flooding. Among the tested mites diapausing on leaf surfaces, 76.3 ± 13.1% (mean ± SE) survived submergence in water for 10 days, 4.8 ± 3.0% - for 20-24 days, 3.0 ± 2.1% - for 30 days, and 8.7 ± 5.9% - for 42-46 days. Among the tested mites diapausing inside wheat straws, 33.3 ± 9.0% remained alive after being flooded for ten days, but none survived being submerged for 30 days. Seasonal floodwaters may be an important factor in spider mite dispersal, geographic isolation, and gene flow. Flooding may also be partially responsible for the formation of species assemblages along rivers.
is a common and ubiquitous bacterium that can cause foodborne illnesses in humans and other animals. Common methods of contact between foodborne pathogens and their victims include exposure through ...contaminated food or food containment products. Using larvae of black soldier flies,
, for biological conversion of wastes into components of animal feeds is a rapidly growing technology. However, contamination of larval biomass with pathogenic microorganisms may challenge its use on an industrial scale. We conducted laboratory experiments to test the effects of the black soldier fly larvae developing on simulated potato waste substrate on
abundance. We observed a general increase in the number of colony-forming units and concentration of
- gene when the larvae were present in the substrate, although the effect was modulated by larval densities and time since inoculation. It is possible that starch breakdown by black soldier fly larvae may provide a beneficial environment for
. Our results differ from the suppression in the presence by black soldier fly larvae reported for several other bacterial species and highlight the importance of taking proper food safety measures when using this technology.
Cannibalism, or intraspecific predation, can play a major role in changing individual fitness and population processes. In insects, cannibalism frequently occurs across life stages, with cannibals ...consuming a smaller or more vulnerable stage. Predation of adult insects on one another is considered to be uncommon. We investigated adult cannibalism in the Colorado potato beetle, Leptinotarsa decemlineata （Say）, which is an oligophagous herbivore specializing on plants in family Solanaceae, and an important agricultural pest. Under laboratory conditions, starvation and crowding encouraged teneral adults to feed upon each other, which reduced their weight loss during the period of starvation. However, pupae were attacked and consumed before adults. Injured beetles had a higher probability of being cannibalized than intact beetles. Males were more frequently attacked than females, but that appeared to be a function of their smaller size rather than other gender-specific traits. Cannibalizing eggs at a larval stage did not affect beetle propensity to cannibalize adults at an adult stage. When given a choice between conspecific adults and mealworrns, the beetles preferred to eat conspecifics. Cannibalistic behavior, including adult cannibalism, could be important for population persistence in this species.
The Colorado potato beetle, Leptinotarsa decemlineata (Say) (Coleoptera: Chrysomelidae) is an insect defoliator of the potato, Solanum tuberosum L. This species thrives in agricultural environments ...because of its flexible and complex life history, as well as its ability to evolve insecticide resistance. As a result, it has become a widely distributed agricultural pest. Ledprona (trade name Calantha) is a recently developed novel double-stranded RNA (dsRNA) insecticide that controls populations of Colorado potato beetle through RNA interference (RNAi). Previous studies have demonstrated the efficacy of ledprona through laboratory, greenhouse, and field studies. Colorado potato beetles from geographically distinct populations are known to vary in their response to insecticides, including experimental compounds based on RNAi. We tested the mortality and foliage consumption of beetles from different areas in the US treated with ledprona and found significant variation in both parameters. The beetles originating from New York were significantly less susceptible to ledprona in leaf disc assays compared to other populations. However, currently there is no evidence of reduced performance of ledprona against that population under field conditions, possibly because intoxicated beetles cannot withstand multiple stressors present in the field. The results of this study confirmed that the ledprona efficacy differs among geographically distinct populations, which may have implications for managing Colorado potato beetles.
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