Cohen, Z. P., K. Brevik, Y. H. Chen, D. J. Hawthorne, B. D. Weibel, and S. D. Schoville. Molecular Ecology

Contextualizing evolutionary history and identifying genomic features of an insect that might contribute to its pest status is important in developing early detection and control tactics. In order to understand the evolution of pestiferousness, which we define as the accumulation of traits that contribute to an insect population's success in an agroecosystem, we tested the importance of known genomic properties associated with rapid adaptation in the Colorado potato beetle (CPB), Leptinotarsa decemlineata Say. Within the leaf beetle genus Leptinotarsa, only CPB, and a few populations therein, has risen to pest status on cultivated nightshades, Solanum. Using whole genomes from ten closely related Leptinotarsa species native to the United States, we reconstructed a high‐quality species tree and used this phylogenetic framework to assess evolutionary patterns in four genomic features of rapid adaptation: standing genetic variation, gene family expansion and contraction, transposable element abundance and location, and positive selection at protein‐coding genes. Throughout approximately 20 million years of history, Leptinotarsa species show little evidence of gene family turnover and transposable element variation. However, there is a clear pattern of CPB experiencing higher rates of positive selection on protein‐coding genes. We determine that these rates are associated with greater standing genetic variation due to larger effective population size, which supports the theory that the demographic history contributes to rates of protein evolution. Furthermore, we identify a suite of coding genes under positive selection that are putatively associated with pestiferousness in the Colorado potato beetle lineage. They are involved in the biological processes of xenobiotic detoxification, chemosensation and hormone function.

Doğan, C., S. Hänniger, D. G. Heckel, C. Coutu, D. D. Hegedus, L. Crubaugh, R. L. Groves, Ş. Bayram, U. Toprak. Archives of Insect Biochemistry and Physiology,

Molecular chaperones are crucial for the correct folding of newly synthesized polypeptides, in particular, under stress conditions. Various studies have revealed the involvement of molecular chaperones, such as heat shock proteins, in diapause maintenance and starvation; however, the role of other chaperones in diapause and starvation relatively is unknown. In the current study, we identified two lectin‐type chaperones with calcium affinity, a calreticulin (LdCrT) and a calnexin (LdCnX), that were present in the fat body of the Colorado potato beetle, Leptinotarsa decemlineata (Coleoptera: Chrysomelidae) during diapause. Both proteins possessed an N‐globular domain, a P‐arm domain, and a highly charged C‐terminal domain, while an additional transmembrane domain was present in LdCnX. Phylogenetic analysis revealed distinction at the order level. Both genes were expressed in multiple tissues in larval and adult stages, and constitutively throughout development, though a starvation response was detected only for LdCrT. In females, diapause‐related expression analysis in the whole body revealed an upregulation of both genes by post‐diapause, but a downregulation by diapause only for LdCrT. By contrast, males revealed no alteration in their diapause‐related expression pattern in the entire body for both genes. Fat body‐specific expression analysis of both genes in relation to diapause revealed the same expression pattern with no alteration in females and downregulation in males by post‐diapause. This study suggests that calcium‐binding chaperones play similar and possibly gender‐specific roles during diapause.

Voigt, D., M.Varenberg, J. Schuppert,, S.N. Gorb. Journal of Insect Physiology,

Green dock beetles Gastrophysa viridula and Colorado potato beetles Leptinotarsa decemlineata having distinctly different body mass and gait habits were compared with respect to their tarsal morphology and attachment ability. The focus laid on shapes and dimensions of tenent setae related to the peeling line, i.e., the sum of widths of all thin-film elements participating in contact. High-speed rotation of the two leaf beetle species attached to the horizontal and vertical sides of a Plexiglass drum resulted in higher attachment forces of the heavier beetle species that has a larger number of tarsal setae and a larger peeling line length. However, normalizing the measured forces with the corresponding peeling line lengths led to a reversed relationship. This allowed us to assume that the design of adhesive setae in different leaf beetle species matches the requirements imposed by their habitats. In accordance with the theory of thin film peeling, tangential forces were found to be higher than normal forces. The attachment system of females was found to exhibit stronger functional efficiency, which can be correlated to the morphology of their setae.

Brevik, K., E. M. Bueno, S. McKay, S. D. Schoville, Y. H. Chen.

Insecticide use is pervasive as a selective force in modern agroecosystems. Insect herbivores exposed to these insecticides have been able to rapidly evolve resistance to them, but how they are able to do so is poorly understood. One possible but largely unexplored explanation is that exposure to sublethal doses of insecticides may alter epigenetic patterns that are heritable.

For instance, epigenetic mechanisms, such as DNA methylation that modify gene expression without changing the underlying genetic code, may facilitate the emergence of resistant phenotypes in complex ways. We assessed the effects of sublethal insecticide exposure, with the neonicotinoid imidacloprid, on DNA methylation in the Colorado potato beetle, Leptinotarsa decemlineata, examining both global changes in DNA methylation and specific changes found within genes and transposable elements. We found that exposure to insecticide led to decreases in global DNA methylation for parent and F2 generations, and that many of the sites of changes in methylation are found within genes associated with insecticide resistance, such as cytochrome P450s, or within transposable elements. Exposure to sublethal doses of insecticide caused heritable changes in DNA methylation in an agricultural insect herbivore. Therefore, epigenetics may play a role in insecticide resistance, highlighting a fundamental mechanism of evolution while informing how we might better coexist with insect species in agroecosystems.

Middleton, E.G. and I. V. MacRae. 2021. Biological Control 152: 104463,

Wildflower plantings in agroecosystems can support arthropod predators, and may have the potential to increase conservation biological control of pest species in nearby crops. Colorado potato beetle (CPB) is a significant defoliator of potato that is resistant to many forms of management. Promoting natural enemies of CPB by establishing perennial wildflower plantings in field margins may provide a measure of control for this pest. We examined the impacts of floral plantings on the abundance of known CPB predators, predation of CPB egg masses, and CPB populations in a commercial agroecosystem. Floral plantings increased the abundance of CPB predators, but did not significantly increase the rate of predation of sentinel CPB egg masses within field margins. Within nearby potato fields, predator abundance and predation rates on CPB eggs were unaffected by the presence of flowers. Colorado potato beetle abundance in potato fields was also not impacted by floral plantings. However, floral margins may provide improved overwintering opportunities for CPB, and further investigation is needed. Perennial wildflower plantings show potential for attracting predators that prey on CPB, but these benefits do not extend into nearby potato crops.

Dively GP, Crossley MS, Schoville SD, Steinhauer N, Hawthorne DJ. 2020. Pest Management Science. 2020 Jul. DOI: 10.1002/ps.5992.

BACKGROUND: Agricultural insect pests frequently exhibit geographic variation in levels of insecticide resistance, which are often presumed to be due to the intensity of insecticide use for pest management. However, regional differences in the evolution of resistance to novel insecticides suggests that other factors are influencing rates of adaptation. We examined median lethal concentration (LC50) bioassay data spanning 15 years and six insecticides (abamectin, imidacloprid, spinosad, cyantraniliprole, chlorantraniliprole, and metaflumizone) for evidence of regional differences in Leptinotarsa decemlineata baseline sensitivity to insecticides as they became commercially available.

RESULTS: We consistently found that larvae from Colorado potato beetle populations from the northwestern USA had the highest baseline sensitivity to novel insecticides, while populations from the eastern USA had the lowest. Comparisons of gene expression between populations from these regions revealed constitutively elevated expression of an array of detoxification genes in the East, but no evidence of additional induction when exposed to imidacloprid.

CONCLUSIONS: Our results suggest a mechanism for geographic variation in rates of adaptation to insecticides, whereby baseline levels of gene expression determine a population's response to novel insecticides. These findings have implications for the regional development of insecticide resistance management strategies and for the fundamental question of what determines the rate of adaptation to insecticides.

Yoon JS, Kim K, Palli SR. 2020. Journal of Asia-Pacific Entomology. 23(4):1160–1164.

Despite extensive research during the past decade elucidating the mechanism of RNA interference (RNAi) in insects, it is not clear how ingested or injected double-stranded RNA (dsRNA) triggers RNAi response in the whole body or even its progeny, which is referred to as systemic RNAi. In the present study, we aim to understand how the dsRNA delivered into cells causes systemic RNAi using Colorado potato beetle cells (Lepd-SL1). We first tested if dsRNA treatment induces systemic RNAi in Lepd-SL1 cells. Exposure of a new batch of Lepd-SL1 cells to the conditioned medium where Lepd-SL1 cells treated with dsRNA targeting inhibitor of apoptosis were grown for 6 h induced apoptosis in these new batch of cells. We hypothesized the exosomes in the conditioned medium are responsible for RNAi-inducing effect. To test this hypothesis, we isolated exosomes from the conditioned medium from Lepd-SL1 cells that had been treated with dsGFP (dsRNA targeting gene coding for green fluorescent protein) or dsLuc (dsRNA targeting gene coding for the luciferase) were grown. RNA present in the purified exosomes was analyzed to check if long dsRNA or siRNA is accumulated in them. The results from the electrophoretic mobility shift assay clearly showed that the long dsRNAs are present in the exosomes. By knockdown of candidate genes involved in endosome recycling and generation pathways, we found that Rab4 and Rab35 are involved in exosome production and transport.

Wu JJ, Mu LL, Chen ZC, Fu KY, Guo WC, Li C, Li GQ. 2019. Journal of Asia-Pacific Entomology. 22(2):443–452.

Chitin deacetylases (CDAs) catalyze N-deacetylation of chitin, a crucial process for chitin modification. In the present paper, LdCDA1 was identified in Leptinotarsa decemlineata. It was copiously expressed in larval foregut, hindgut and epidermis. Just before the molt in the first, second and third larval instars, the mRNA levels of LdCDA1 were high. In the fourth (final)-instar larvae, a peak occurred 4 days after ecdysis. In vivo results revealed that LdCDA1 transcriptionally responded, positively and negatively respectively, to 20-hydroxyecdysone and juvenile hormone titers. Moreover, knockdown of LdCDA1 significantly reduced foliage consumption, lengthened developing period and prevented growth in the final instar larvae. Three distinct lethal phenotypes were noted in the LdCDA1 RNAi larvae. About 30% of the RNAi larvae became moribund and finally died; approximately 50% of deformed pupae died as pharate adults; and around 20% of LdCDA1 depleted pupae finally emerged as abnormal adults and eventually died within 1 week after emergence. Furthermore, chitin content was low and the mRNA levels of five chitin biosynthesis transcripts (LdUAP1, LdUAP2, LdChSAa, LdChSAb and LdChSB) were significantly declined in the LdCDA1 RNAi larvae. In addition, glucose, trehalose and glycogen contents were increased in the LdCDA1 depleted hypomorphs, along with highly expressed genes coding for trehalose and glycogen synthesis enzymes. The findings provide a compelling piece of evidence that CDA1 is critical for chitin deposition in L. decemlineata. Moreover, LdCDA1 may be a potential target for control of the larvae.

Rainio MJ, Margus A, Virtanen V, Lindström L, Salminen JP, Saikkonen K, Helander M. 2020. Chemosphere. 258:127254.

Glyphosate is the most used herbicide worldwide, targeting physiological pathways in plants. Recent studies have shown that glyphosate can also cause toxic effects in animals. We investigated the glyphosate-based herbicide (GBH)-induced changes in potato (Solanum tuberosum) plant chemistry and the effects of a GBH on the survival rate and oxidative status of the Colorado potato beetle (Leptinotarsa decemlineata). The beetles were reared on potato plants grown in pots containing soil treated with a GBH (Roundup Gold, 450 g/l) or untreated soil (water control). The 2nd instar larvae were introduced to the potato plants and then collected in 2 phases: as 4th instar larvae and as adults. The main glycoalkaloids of the potato plants, α-solanine and α-chaconine, were measured twice during the experiment. The α-solanine was reduced in potato plants grown in GBH-treated soil, which can be detrimental to plant defenses against herbivores. GBH treatment had no effect on the survival rate or body mass of the larvae or the adult beetles. In the larvae, total glutathione (tGSH) concentration and the enzyme activity of catalase (CAT), superoxide dismutase, and glutathione-S-transferase were increased in the GBH treatment group. In the adult beetles, CAT activity and tGSH levels were affected by the interactive effect of GBH treatment and the body mass. To conclude, environmentally relevant concentrations of a GBH can affect the potato plant's glycoalkaloid concentrations, but are not likely to directly affect the survival rate of the Colorado potato beetle, but instead, modify the antioxidant defense of the beetles via diet.

Miller T, Crossley MS, Fu Z, Meier AR, Crowder DW, Snyder WE. 2020. Biological Control. 151:104403.

Prey commonly must compete with conspecifics for resources while also defending themselves against predators. Both competition and defense can reduce feeding opportunities, or otherwise strain prey energy reserves, even when the prey is not killed. This suggests that stress from competition and anti-predator defense might yield non-lethal harm that differs quantitatively rather than qualitatively. We examined this possibility for an herbivorous prey species, the Colorado potato beetle (Leptinotarsa decemlineata), feeding among differing numbers of intraspecific competitors while also avoiding predation by Nabis alternatus and Hippodamia convergens. In a field cage experiment, predators and intraspecific competitors similarly reduced the herbivores’ larval survival. Yet, only exposure to predators heightened the risk of beetles later being killed by entomopathogenic nematodes and fungi, as the beetles pupated in the soil. This suggests that the threat of predation was exerting physiological harm to the prey, although it was also possible that a tradeoff existed where those beetles most resistant to predator attack were most susceptible to pathogen infection. In a second experiment, we found that regular touching of larvae by predatory Nabis or Hippodamia, in the absence of actual predation, consistently induced predator-avoidance behaviors by the larvae (e.g., rearing, vomiting, wiggling, etc.) that might be energetically costly. Indeed, only being touched by predators, but not by a wooden dowel as a disturbance control, led to greater susceptibility to entomopathogens as pupae. Altogether, our findings suggest that non-lethal interactions with predators might carry a particular cost to beetles in heightened pathogen susceptibility, not seen when facing competitors or simple disturbance. Additional work is needed to determine the specific physiological mechanism underlying this apparent synergism in the effects of predators and entomopathogens, which spans prey life stages.