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BACKGROUND: The Colorado potato beetle (CPB), Leptinotarsa decemlineata, is a major potato (Solanum tuberosum) pest, infesting over 16 million km2 and causing substantial economic losses. The insect cuticle forms an apical extracellular matrix (ECM) envelope covering exposed organs to direct morphogenesis and confer structural protection. While select chitinase (Cht) genes have proven essential for larval development, their potential activities directing ECM remodeling underlying adult wing maturation remain undefined. RESULTS: We investigated the expression patterns and performed an oral RNA interference (RNAi) screen targeting 19 LdChts in late‐instar L. decemlineata larvae. Subsequently, we assessed their effects on adult eclosion and wing characteristics. Knockdown of LdCht5, LdCht7, LdCht10, LdIDGF2, and LdIDGF4, as well as others from Group IV (LdCht15, LdCht12, LdCht17, and LdCht13) and Groups VII–X (LdCht2, LdCht11, LdCht1, and LdCht3), resulting in shrunken, misshapen elytra with reduced areal density, as well as transverse wrinkling and impaired wing‐tip folding in hindwings. Scanning electron micrographs revealed eroded elytral ridges alongside thinned, ruptured hindwing veins, indicative of mechanical fragility post‐LdCht suppression. Spectroscopic analysis uncovered biomolecular alterations underlying the elytral anomalies, including decreases in peaks representing chitin, proteins, and lipids. This loss of essential ECM components provides evidence for the fragility, wrinkling, and shrinkage observed in the RNAi groups. CONCLUSION: Our findings elucidate the crucial role of chitinases in the turnover of chitinous cuticles on beetle wings, offering insights into RNAi‐based control strategies against this invasive pest. © 2024 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Simple Summary: The Colorado potato beetle, Leptinotarsa decemlineata, poses a significant threat to potato crops worldwide, causing substantial agricultural damage. To better understand how these pests develop and potentially find ways to control them, we studied a group of genes called Knickkopf (Knk) genes in the larvae of the beetle. These genes are crucial for forming and maintaining the insect's outer shell, known as the cuticle, which protects them from the environment. We identified four LdKnk-family genes in the beetle and analyzed when and where these genes are expressed during the insect's growth stages. By using RNA interference (RNAi) to silence these genes, we observed that larvae became weaker, with damaged cuticles and higher death rates. These findings indicate that LdKnk-family genes are vital for the beetle's survival and development. Targeting LdKnk-family genes could provide a new strategy for pest control, potentially weakening the beetles' defenses and making them more susceptible to environmental factors and insecticides, ultimately helping to protect potato crops. The Colorado potato beetle (Leptinotarsa decemlineata) is a major pest of potato crops. While Knickkopf (Knk) genes are essential for insect cuticle formation, their roles in pests like L. decemlineata remain unclear. This study aims to identify and characterize Knk genes in L. decemlineata and explore their functions in larval development and cuticle integrity. We used genomic and transcriptomic databases to identify LdKnk-family genes, validated through RT-PCR and RACE. Gene expression was analyzed at various developmental stages and tissues using qRT-PCR. RNA interference (RNAi) and Transmission electron microscopy (TEM) were applied to determine the functional roles of these genes. Four LdKnk-family genes were identified. Spatio-temporal expression analysis indicated significant gene expression during larval molting and pupal stages, especially in the epidermis. RNAi experiments showed that silencing LdKnk and LdKnk3-5′ led to reduced larval weight, cuticle thinning, and increased mortality, while LdKnk3-FL knockdown caused abnormal cuticle thickening and molting disruptions. LdKnk2 knockdown increased epicuticle and endocuticle thickness without visible phenotypic changes. The study highlights the essential roles of LdKnk-family genes in maintaining cuticle structure and integrity, suggesting their potential as targets for RNAi-based pest control. [ABSTRACT FROM AUTHOR]

During choriogenesis in insects, chorion (eggshell) is formed by surrounding follicular epithelial cells in ovarioles. However, the regulatory endocrine factor(s) activating choriogenesis and the effect of chemical components on eggshell deserve further exploration. In two representative coleopterans, a coccinellid Henosepilachna vigintioctopunctata and a chrysomelid Leptinotarsa decemlineata, genes encoding the 20-hydroxyecdysone (20E) receptor heterodimer, ecdysone receptor (EcR) and ultraspiracle (USP), and two chitin biosynthesis enzymes UDP-N-acetylglucosamine pyrophosphorylase (UAP) and chitin synthase (ChS1), were highly expressed in ovaries of the young females. RNA interference (RNAi)-aided knockdown of either HvEcR or Hvusp in H. vigintioctopunctata inhibited oviposition, suppressed the expression of HvChS1, and lessened the positive signal of Calcofluor staining on the chorions, which suggests the reduction of a chitin-like substance (CLS) deposited on eggshells. Similarly, RNAi of LdEcR or Ldusp in L. decemlineata constrained oviposition, decreased the expression of LdUAP1 and LdChS1, and reduced CLS contents in the resultant ovaries. Knockdown of LdUAP1 or LdChS1 caused similar defective phenotypes, i.e., reduced oviposition and CLS contents in the L. decemlineata ovaries. These results, for the first time, indicate that 20E signaling activates choriogenesis in two coleopteran species. Moreover, our findings suggest the deposition of a CLS on the chorions. [ABSTRACT FROM AUTHOR]

BACKGROUND: Insect chitinases play crucial roles in degrading chitin in the extracellular matrix, affecting insect development and molting. However, our understanding of the specific functions of various chitinases in Leptinotarsa decemlineata is limited, hindering the deployment of novel gene‐targeting technologies as pest management strategies. RESULTS: We identified and characterized 19 full‐length complementary DNA (cDNA) sequences of chitinase genes (LdChts) in Leptinotarsa decemlineata. Despite having varying domain architectures, all these chitinases contained at least one chitinase catalytic domain. Phylogenetic analysis classified the chitinase proteins into ten distinct clusters (groups I–X). Expression profiles showed the highest expression in chitin‐rich tissues or during specific developmental stages from the larva‐to‐pupa transition. Gene‐specific RNA interference (RNAi) experiments provided valuable insight into chitinase gene function. Silencing of group II LdCht10 prevented larval–larval molting, larval–prepupal, and prepupal–pupal processes. Moreover, our study revealed that LdCht5, LdCht2, LdCht11, LdCht1, and LdCht3 from groups I and VII–X were specifically essential for the transition from prepupal to pupal stage, whereas LdIDGF2 from group V was necessary for the larval–prepupal metamorphic process. The chitinase gene LdCht7 from group III and LdIDGF4 from group V were involved in both the larva‐to‐prepupa and the prepupa‐to‐pupa shift. Additionally, our findings also shed light on the exclusive expression of nine chitinase genes within group IV in the digestive system, suggesting their potential role in regulating larval body weight and larva‐to‐pupa transition. CONCLUSION: Our results provide a comprehensive understanding of the functional specialization of chitinase genes during the molting process of various stages and identify potential targets for RNAi‐based management of Leptinotarsa decemlineata. © 2023 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

[Objectives] To determine the role of nitrogen in the population explosion of the Asian corn borer, Ostrinia furnacalis (Guenee), we compared the effects of artificial diets with different nitrogen contents on food utilization by, and nutritional content of, this species. [Methods] Five artificial diets (N1, N2, N3, N4 and N5) with nitrogen contents of 3.94%, 7.33%, 10.69%, 18.84% and 30.31% were fed to 3rd, 4th and 5th-instar larvae, and differences in food utilization and larval nutritional indices between treatment groups were compared and analyzed. [Results] The highest food utilization rate (16.28% ± 0.89%) and food conversion rate (41.05% ± 4.07%) was recorded in 3rd instar larvae in the N3 treatment group, significantly higher than the corresponding values in the N4 and N5 treatment groups. The lowest food utilization and food conversion rate, 3.32% ± 0.57% and 2.36% ± 0.69%, respectively, were recorded in the N2 treatment group. The highest food utilization rate in 4th instar larvae, 22.13% ± 1.51%, was recorded in the N5 treatment group. 5th instar larvae in the N5 and N4 treatment groups had the highest food utilization and approximate digestibility; 24.29% ± 7.01% and 40.93% ± 2.94%, respectively, but dietary nitrogen had no significant effect on food conversion and relative growth rates (P > 0.05). The nitrogen use efficiency of 3rd, 4th and 5th-instar larvae all increased with dietary nitrogen content; the highest nitrogen use efficiencies for each instar (72.33%, 86.09% and 90.91%, respectively) were all recorded in the N5 treatment group. [Conclusion] Dietary nitrogen has a direct impact on food utilization by, and the nutritional indices of, Asian corn borer larvae, and dietary nitrogen significantly increased food use efficiency, relative feeding rate and nitrogen use efficiency of 4th-instar and 5th-instar larvae. [ABSTRACT FROM AUTHOR]

BACKGROUND RNA interference (RNAi) is a breakthrough technology in pest control. It is highly efficient to Coleopteran pests such as the Colorado potato beetle Leptinotarsa decemlineata, a serious pest defoliator mainly attacking potatoes worldwide. The first step for effective pest control by RNAi is the development of effective and reliable target genes. RESULTS: Our results revealed that continuous ingestion of dsLdRan for 3 days successfully silenced the target gene, inhibited larval growth and killed 100% L. decemlineata larvae. When the bioassay began at the second‐, third/fourth‐instar larval stages, the larval lethality mainly occurred at the fourth larval instar and prepupal stages, respectively. Importantly, consumption of dsLdRan for 3 days by the newly‐emerged males and females effectively knocked down the target transcript, reduced fresh weights and caused 100% of lethality within a week. The LdRan females possessed underdeveloped ovaries. CONCLUSION: Considering that the larvae, adults and eggs are simultaneously sited on the potato plants, bacterially‐expressed dsLdRan is a potential RNAi‐based strategy for managing L. decemlineata in the potato field. © 2022 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

A heterodimer of two nuclear receptors, ecdysone receptor (EcR) and ultraspiracle, mediates 20‐hydroxyecdysone (20E) signaling to modulate many aspects in insect life, such as molting and metamorphosis, reproduction, diapause and innate immunity. In the present paper, we intended to determine the isoform‐specific roles of EcR during larval–pupal–adult transition in the Colorado potato beetle. Double‐stranded RNAs (dsRNAs) were prepared using the common (dsEcR) or isoform‐specific (dsEcRA, dsEcRB1) regions of EcR as templates. Ingestion of either dsEcR or dsEcRA, rather than dsEcRB1, by the penultimate (3rd) and final (4th) instar larvae caused failure of larval–pupal and pupal–adult ecdysis. The RNA interference (RNAi) larvae remained as prepupae, or became deformed pupae and adults. Determination of messenger RNA (mRNA) levels of EcR isoforms found that LdEcRA regulates the expression of LdEcRB1. Moreover, silencing the two EcR transcripts, LdEcRA or LdEcRB1 reduced the mRNA levels of Ldspo and Ldsad, and lowered 20E titer. In contrast, the expression levels of HR3, HR4, E74 and E75 were significantly decreased in the LdEcR or LdEcRA RNAi larvae, but not in LdEcRB1 depleted specimens. Dietary supplement with 20E did not restore the expression of five 20E signaling genes (USP, HR3, HR4, E74 and E75), and only partially alleviated the pupation defects in dsEcR‐ or dsEcRA‐fed beetles. These data suggest that EcR plays isoform‐specific roles in the regulation of ecdysteroidogenesis and the transduction of 20E signal in L. decemlineata. [ABSTRACT FROM AUTHOR]