Andrés MF, Rossa GE, Cassel E, et al. Food and Chemical Toxicology. 2017;109:1086-1092.

In this study we evaluated the effect of a pressure gradient (1-2 atm) in the extraction and composition of the essential oil (EO) of Piper hispidinervum by steam distillation. We also evaluated the insect antifeedant effects (Spodoptera littoralis, Leptinotarsa decemlineata, Myzus persicae and Rhopalosiphum padi) and nematicidal activity (Meloidogyne javanica) of the oils, their major components and their synergistic interactions. Safrole was the major component (78-81%) followed by terpinolene (5-9%). The EOs tested were effective insect antifeedants. Safrole, explained most of the insect antifeedant action of P. hispidinervum EOs. When safrole and terpinolene were tested in binary combinations, low ratios of safrole improved the antifeedant effects of terpinolene. P. hispidinervum EOs caused higher mortality of M. javanica juveniles than their major components. In binary combinations, low ratios of terpinolene increased the nematicidal effects of safrole. The EO treatment strongly suppressed nematode egg hatching and juvenile infectivity. P. hispidinervum EOs affected the germination of S. lycopersicum and L. sativa mostly at 24 h of treatment, being L. sativa the most sensitive. Safrole moderately affected germination and root growth of L. sativa, S. lycopersicum and L. perenne. Terpinolene only affected S. lycopersicum root growth.

QingWei M, QingYu X, Pan D, KaiYun F, WenChao G, GuoQing L. Insect Biochem Mol Biol. 2018;97:1-9. doi: 10.1016/j.ibmb.2018.04.003.

In the tobacco hornworm Manduca sexta, juvenile hormone (JH) is critical for the control of species-specific size. However, whether the basic helix-loop-helix/Per-Arnt-Sim domain receptor methoprene-tolerant (Met) is involved remains unconfirmed. In the present paper, we found that RNA interference (RNAi)-aided knockdown of Met gene (LdMet) lowered the larval and pupal fresh weights and shortened the larval development period in the Colorado potato beetle Leptinotarsa decemlineata. Dietary introduction of JH into the LdMet RNAi larvae rescued neither the decreased weights nor the reduced development phase, even though JH ingestion by control larvae extended developmental time and caused large pupae. Moreover, the transcript levels of five genes involved in prothoracicotropic hormone and cap 'n' collar isoform C/Kelch-like ECH associated protein 1 pathways were upregulated in the LdMet silenced larvae. Ecdysteroidogenesis was thereby activated; 20-hydroxyecdysone (20E) titer was increased; and 20E signaling pathway was elicited in the LdMet RNAi larvae. Therefore, JH, acting through its receptor Met, inhibits PTTH production and release before the attainment of critical weight. Once the critical weight is reached, JH production and release are averted; and the hemolymph JH is removed. The elimination of JH allows the brain to release PTTH. PTTH subsequently stimulates ecdysteroid biosynthesis and release to start larval-pupal transition in L. decemlineata.

Morin MD, Lyons PJ, Crapoulet N, Boquel S, Morin PJ. International Journal of Molecular Sciences. 2017;18(12):2728. doi: 10.3390/ijms18122728.

The Colorado potato beetle (Leptinotarsa decemlineata (Say)) is a significant pest of potato plants that has been controlled for more than two decades by neonicotinoid imidacloprid. L. decemlineata can develop resistance to this agent even though the molecular mechanisms underlying this resistance are not well characterized. MicroRNAs (miRNAs) are short ribonucleic acids that have been linked to response to various insecticides in several insect models. Unfortunately, the information is lacking regarding differentially expressed miRNAs following imidacloprid treatment in L. decemlineata. In this study, next-generation sequencing and quantitative real-time polymerase chain reaction (qRT-PCR) were used to identify modulated miRNAs in imidacloprid-treated versus untreated L. decemlineata. This approach identified 33 differentially expressed miRNAs between the two experimental conditions. Of interest, miR-282 and miR-989, miRNAs previously shown to be modulated by imidacloprid in other insects, and miR-100, a miRNA associated with regulation of cytochrome P450 expression, were significantly modulated in imidacloprid-treated beetles. Overall, this work presents the first report of a miRNA signature associated with imidacloprid exposure in L. decemlineata using a high-throughput approach. It also reveals interesting miRNA candidates that potentially underly imidacloprid response in this insect pest.

YanWei W, YuZhe L, GuoQing L, PinJun W, Chao L. J Econ Entomol. 2019;112(2):912-923. doi: 10.1093/jee/toy396.

Structural cuticular proteins (CPs) are the primary components of insect cuticle, linings of salivary gland, foregut, hindgut and tracheae, and midgut peritrophic membrane. Variation of CPs in insect cuticle can cause penetration resistance to insecticides. Moreover, depletion of specific CP by RNA interference may be a suitable way for the development of potential pest control traits. Leptinotarsa decemlineata (Say) CPs are poorly characterized at present, and therefore, we mined the genome and transcriptome data to better annotate and classify L. decemlineata CPs in this study, by comparison with the annotated CPs of Tribolium castaneum Browse (Coleoptera: Tenebrionidae). We identified 175 CP genes. Except one miscellaneous CP with an 18-amino acid motif, these CPs were classified into 7 families based on motifs and phylogenetic analyses (CPs with a Rebers and Riddiford motif, CPR; CPs analogous to peritrophins, CPAP3 and CPAP1; CPs with a tweedle motif, TWDL; CPs with a 44-amino acid motif, CPF; CPs that are CPF-like, CPFL; and CPs with two to three copies of C-X5-C motif, CPCFC). Leptinotarsa decemlineata CPRs could be categorized into three subfamilies: RR-1 (50), RR-2 (85), and RR-3 (2). The RR-1 proteins had an additional motif with a conserved YTADENGF sequence. The RR-2 members possessed a conserved RDGDVVKG region and three copes of G-x(3)-VV. Few genes were found in TWDL (9), CPAP1 (9), CPAP3 (8), CPF (5), CPFL (4), and CPCFC (2) families. The findings provide valuable information to explore molecular modes of penetration resistance to insecticides and to develop dsRNA-based control method in L. decemlineata.

Tigreros N, Wang EH, Thaler JS. Funct Ecol. 2018;32(4):982-989. doi: 10.1111/1365-2435.13046.

Prey species can respond to the risk of predation with a range of antipredator behaviours and physiological changes. While these responses increase chances of survival, they often involve feeding reductions and greater energy expenditure (e.g. increases in metabolic rate). As a consequence, a prey response is constrained by its own nutritional condition. While a number of studies indeed demonstrate that prey in better nutritional condition have stronger antipredator behaviours, we do not yet understand how condition impacts the physiological component of the prey's response. Previous research revealed that Leptinotarsa decemlineata beetles experiencing predation risk improve their offspring's nutritional condition by promoting intraclutch egg cannibalism. Importantly, egg cannibalism decreased offspring vulnerability by increasing larval behavioural responses to chronic predation risk. In this study we test if egg cannibalism similarly impacts larval physiological responses by comparing how risk of predation in cannibals and non-cannibals affects their behaviour (e.g. feeding reductions), metabolic rate and energy stores. We found that non-cannibals did not exhibit antipredator behaviours but responded physiologically, by increasing metabolic rates. In contrast, cannibals responded behaviourally, suppressing feeding, but without altering metabolism. While cannibals and non-cannibals coupled food intake and energy expenditure differently, both reached similar growth rates and had similar energy stores when facing chronic predation risk. These results indicate that increases in predator avoidance behaviours are not merely mirrored by a stronger physiological response. Instead, changes in metabolism appear to ameliorate, within our experimental conditions, the costs associated with the behavioural response. Prey in poorer nutritional state are not less responsive to predators but appear to rely more heavily on physiological responses, demonstrating that how prey integrates behaviour and physiology depends on their own nutritional state.

Wu JJ, Chen ZC, Wang YW, Fu KY, Guo WC, Li GQ. Insect Mol Biol. 2019;28(1):52-64.

Insect chitin deacetylases (CDAs) are carbohydrate esterases that catalyze N-deacetylation of chitin to generate chitosan, a process essential for chitin organization and compactness during the formation of extracellular chitinous structure. Here we identified two CDA2 splice variants (LdCDA2a and LdCDA2b) in Leptinotarsa decemlineata. Both splices were abundantly expressed in larval foregut, rectum, and epidermis; their levels peaked immediately before ecdysis within each instar. In vivo results revealed that the two isoforms transcriptionally responded, positively and negatively respectively, to 20-hydroxyecdysone and juvenile hormone signaling pathways. RNA interference (RNAi)-aided knockdown of the two LdCDA2 variants (hereafter LdCDA2) or LdCDA2b, rather than LdCDA2a, resulted in three negative effects. First, foliage consumption was significantly reduced, larval developing period was lengthened, and larval growth was retarded. Second, chitin contents were reduced, whereas glucose, trehalose, and glycogen contents were increased in the LdCDA2 and LdCDA2b RNAi larvae. Third, approximately 20% of LdCDA2 and LdCDA2b RNAi larvae were trapped within the exuviae and finally died. About 60% of the abnormal pupae died as pharate adults. Around 20% of the RNAi pupae emerged as deformed adults, with small size and wrinkled wings. These adults eventually died within 1 week after molting. Our results reveal that knockdown of CDA2 affects chitin accumulation. Consequently, LdCDA2 may be a potential target for control of L. decemlineata larvae.

Yang L, KaiYun F, Tursun, Jiang H, WenChao G. Journal of Environmental Entomology. 2018;40(3):633-644.

As a microsatellite molecular marker method, SSR is widely used in genetic diversity research, and primers' efficiency is fundamental to the experiment's results. By the use of bioinformatics methods, SSR loci in Colorado potato beetle's genomes were analyzed, SSR primers were designed and the efficiency were verified. As results, 81 937 SSR loci were detected in the genomes, most of which were single nucleotide, dinucleotide and trinucleotide repeats, and the average distribution distance of SSR loci were 2.21 kb, 8.39 kb and 36.21 kb, respectively. Of the CPB genome SSR loci, advantage motifs account for a total of 68 388, up to 83.46%, among which the single nucleotide type as A/T repeats motif dominates, the dinucleotide type AG/CT repeats motif ranks secondly, and trinucleotide types AAT/ATT repeats ranks thirdly. Among the Class I length type of SSR loci, the largest number were dinucleotide and trinucleotide repeats and the proportion is 82.32%. 19 primers were designed to test efficiency, among which 14 primers were amplified successfully, 11 primers were confirmed as polymorphism primers with averaged 6.27 bands and the PIC value range between 0.375 to 0.794, the average was 0.600, of which larger than 0.600 was 6 pairs, accounting for 54.5%. The results showed that, it was more efficient for developing SSR primers by bioinformatics method. This result will help to study Colorado potato beetle's genetic diversity and lay foundation of developing SSR primers in insects.

Chertkova EA, Grizanova EV, Dubovskiy IM. J Invertebr Pathol. 2018;153:203-206. doi: 10.1016/j.jip.2018.02.020.

Dopamine (DA) is known as a hormone neurotransmitter molecule involved in several stress reactions in both vertebrates and invertebrates. Following infections with the fungi Metarhizium robertsii or Beauveria bassiana and the bacterium Bacillus thuringiensis, dopamine the concentration was measured at different time points in the haemolymph of the Colorado potato beetle, Leptinotarsa decemlineata and the larvae of the greater wax moth Galleria mellonella. The infection with M. robertsii increased (4 to 12-fold) DA concentrations in the haemolymph of the potato beetle larvae and the oral infection by B. thuringiensis also lead to a 30 and 45-fold increase. During infection of the greater wax moth larvae with Beauveria bassiana and B. thuringiensis DA increased 4 to 20-fold and about 2 to 2,5-fold respectively, compared to non-infected insects. The relative DA concentrations varied between the two insects and depended on the pathogens and post infection time.

Kaplanoglu E, Chapman P, Scott IM, Donly C. Scientific Reports. 2017;7(5):1762. doi: 10.1038/s41598-017-01961-4.

Current control of insect pests relies on chemical insecticides, however, insecticide resistance development by pests is a growing concern in pest management. The main mechanisms for insecticide resistance typically involve elevated activity of detoxifying enzymes and xenobiotic transporters that break-down and excrete insecticide molecules. In this study, we investigated the molecular mechanisms of imidacloprid resistance in the Colorado potato beetle, Leptinotarsa decemlineata (Say) (Coleoptera: Chrysomelidae), an insect pest notorious for its capacity to develop insecticide resistance rapidly. We compared the transcriptome profiles of imidacloprid-resistant and sensitive beetle strains and identified 102 differentially expressed transcripts encoding detoxifying enzymes and xenobiotic transporters. Of these, 74 were up-regulated and 28 were down-regulated in the resistant strain. We then used RNA interference to knock down the transcript levels of seven up-regulated genes in the resistant beetles. Ingestion of double-stranded RNA successfully knocked down the expression of the genes for three cytochrome P450s (CYP6BQ15, CYP4Q3 and CYP4Q7), one ATP binding cassette (ABC) transporter (ABC-G), one esterase (EST1), and two UDP-glycosyltransferases (UGT1 and UGT2). Further, we demonstrated that silencing of CYP4Q3 and UGT2 significantly increased susceptibility of resistant beetles to imidacloprid, indicating that overexpression of these two genes contributes to imidacloprid resistance in this resistant strain.

SeungHo C, Scully ED, Peiffer M, et al. Scientific Reports. 2017;7(1):39690. doi: 10.1038/srep39690.

Herbivore associated bacteria are vital mediators of plant and insect interactions. Host plants play an important role in shaping the gut bacterial community of insects. Colorado potato beetles (CPB; Leptinotarsa decemlineata) use several Solanum plants as hosts in their natural environment. We previously showed that symbiotic gut bacteria from CPB larvae suppressed jasmonate (JA)-induced defenses in tomato. However, little is known about how changes in the bacterial community may be involved in the manipulation of induced defenses in wild and cultivated Solanum plants of CPB. Here, we examined suppression of JA-mediated defense in wild and cultivated hosts of CPB by chemical elicitors and their symbiotic bacteria. Furthermore, we investigated associations between the gut bacterial community and suppression of plant defenses using 16 S rRNA amplicon sequencing. Symbiotic bacteria decreased plant defenses in all Solanum hosts and there were different gut bacterial communities in CPB fed on different host plants. When larvae were reared on different hosts, defense suppression differed among host plants. These results demonstrate that host plants influence herbivore gut bacterial communities and consequently affect the herbivore's ability to manipulate JA-mediated plant defenses. Thus, the presence of symbiotic bacteria that suppress plant defenses might help CPB adapt to host plants.