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Waprin, a WAP (Whey acidic protein) domain‐containing extracellular secretory protein, is widely known for its antibacterial properties. In this study, a waprin homologue (Tc_wapF) expressing in a female‐specific manner was identified in Tribolium castaneum, through the analysis of sex‐specific transcriptomes. Developmental‐ and tissue‐specific profiling revealed the widespread expression of Tc_wapF in adult female tissues, particularly in the ovary, gut and fatbody. This female‐specific expression of Tc_wapF is not regulated by the classical sex‐determination cascade of T. castaneum, as we fail to get any attenuation in Tc_wapF transcript levels in Tcdsx and Tctra (key players of sex determination cascade of T. castaneum) knockdown females. RNA interference‐mediated knockdown of Tc_wapF in females led to the non‐hatching of eggs laid by these females, suggesting the crucial role of Tc_wapF in the embryonic development in T. castaneum. This is the first report on the identification of a sex‐specific waprin homologue in an insect and its involvement in embryonic development. Future investigations on the functional conservation of insect waprins and their mechanistic role in embryonic development can be exploited for improving pest management strategies. [ABSTRACT FROM AUTHOR]

Plant extracts offer an alternative approach to safeguarding stored food products. Our research is focused on assessing the insecticidal properties of aqueous and ethanolic extracts containing bioactive compounds obtained from two species of Azolla; A. pinnata and A. Microphylla against pests that affect stored food items. Two drying processes were conducted in order to compare the yield of bioactive compounds (freeze-drying and oven-drying). We employed two extraction methods (maceration and decoction), using ethanol and water as solvents. Phytochemical screening of both extracts was carried out by CG-MS analysis. The insecticidal properties of the obtained extracts were assessed using the spraying method on larvae and adults of Tribolium castaneum. The results indicated that the freeze-dried method was more efficient comparing to oven-drying, recording the highest value of 26.07%, with the ethanolic extract by decoction. GC–MS analysis for both species confirmed the existence of primarily fatty acid, terpenoid, steroid, coumarin, and flavonoid derivatives compounds. Overall, the various extracts exhibited high toxicity against both larvae and adults. The mortality rate was increased according to the increasing concentration of extracts. The lowest LC50 of A. pinnata and A. microphylla extracts were found to be 872.42 and 894.65 µg/mL, respectively. The aqueous extract caused higher toxicity, reaching 96%. The findings of this study suggest that the dry matter from both azolla species exhibited significant toxicity, evidenced by the mortality of both the larvae and adults. Extracts from A. pinnata were found to be more toxic to T. castaneum in comparison with A. microphylla extracts. [ABSTRACT FROM AUTHOR]

As the catalytic subunit of the Elongator complex, Elongator protein 3 (Elp3) plays a crucial role in multiple physiological processes, including growth, development and immune responses. Previous studies on Elp3 have focused on Caenorhabditis elegans, Drosophila melanogaster, Homo sapiens (human) or Mus musculus (mouse), whereas there are few reports on Elp3 in agricultural pests. Here, the role of TcElp3 in reproduction in the red flour beetle, Tribolium castaneum, was investigated, and the underlying mechanisms were explored. The results showed that RNA interference (RNAi)‐mediated knockdown of TcElp3 in female pupae led to decreased fecundity in female adults. Consistently, the injection of dsTcElp3 into female pupae decreased the mRNA levels of the vitellogenin (Vg)‐encoding genes TcVg1 and TcVg2 in female adults. Notably, knockdown of TcElp3 upregulated the expression of forkhead box protein O (FOXO) at both the mRNA and the protein level in T. castaneum, and promoted the nuclear translocation of TcFOXO. Additionally, TcElp3 directly interacts with TcFOXO and the silencing of TcElp3 significantly decreased the acetylation level of TcFOXO. Overall, our studies reveal that Elp3 regulates beetle reproduction by interacting with FOXO and modulating its acetylation status. [ABSTRACT FROM AUTHOR]

Eucalyptol is one of the major insecticidal active ingredients in a variety of plant essential oils, and has good killing and avoidance effects on Tribolium castaneum. The presence of detoxifying enzymes glutathione S-transferase (GST) in T. castaneum makes it resistant to a variety of insecticides. However, whether GST is involved in regulating the sensitivity of eucalyptol by T. castaneum is not well understood. In our previous study, a glutathione S-transferase, TcGSTu1, was significantly up-regulated in RNA sequencing data when T. castaneum was exposed to eucalyptol. Therefore, in this study, the role of TcGSTu1 in the regulating the sensitivity of T. castaneum to eucalyptol was studied. The enzyme activities of GST and the transcription levels of TcGSTu1 were significantly increased following stimulation with eucalyptol. When using RNA interference technology knockdown TcGSTu1 heightens the sensitivity of T. castaneum to eucalyptol, demonstrating a link between TcGSTu1 and eucalyptol detoxification metabolism. Furthermore, TcGSTu1 is expressed in all developmental stages of T. castaneum, with higher expression levels observed particularly in the late egg stage. There was significant expression of TcGSTu1 in various tissues of different organisms, including larval head, fat body, and adult head. This observation indicated a possible connection between high TcGSTu1 expression and eucalyptol detoxification. The present findings suggest that TcGSTu1 may be involved in regulating the sensitivity and response of T. castaneum to treatment with eucalyptol, providing new research insight into pest control. [ABSTRACT FROM AUTHOR]

C-type lectins (CTLs), as a large family of pattern recognition receptors (PRRs), have been reported to be involved in bacterial infection, but the role of CTLs in development has been poorly understood in insects. The orthologues of Tribolium castaneum CTL9 (TcCTL9) have been identified among insects, but its functions were currently unclear. Therefore, we performed functional analysis of TcCTL9 in this study. Our results indicated that TcCTL9 could bind to bacteria through lipopolysaccharide and peptidoglycan, and agglutinate Gram-positive and Gram-positive bacteria in a Ca2+-dependent manner. Silencing TcCTL9 reduced the immune resistance to Staphylococcus aureus and Escherichia coli , decreased the expression of antimicrobial peptides and prophenoloxidase, and inhibited the phenoloxidase activity. These data suggested that TcCTL9 functioned in the immune response via the Toll and IMD pathways and prophenoloxidase system. During development, TcCTL9 had high expression in the periods of egg to larva and pupa to adult, and knockdown of TcCTL9 suppressed the metamorphosis, egg production and hatchability, and ovary development through ecdysone and juvenile hormone pathways in T. castaneum. This study comprehensively clarified the functions of TcCTL9 orthologues in insects and provided the theoretical basis for developing novel targets of pesticides. [Display omitted] • TcCTL9 could bind to bacteria via LPS and PGN. • TcCTL9 agglutinated the bacteria in a Ca2+ dependent manner. • Silencing TcCTL9 reduced the immune response via the Toll, IMD and proPO system. • TcCTL9 RNAi blocked the metamorphosis and reproduction by ecdysone and JH pathways. [ABSTRACT FROM AUTHOR]

The highly conserved Elongator complex plays important roles in histone acetylation and tRNA modification. Currently, Elongator complex has been shown to be essential for a range of biological processes, but its function in insect hormone signaling is poorly understood. In this study, the cDNA encoding TcElp3, the catalytic subunit of the Elongator complex in Tribolium castaneum, was cloned and functionally characterized. Analysis of temporal and spatial expression patterns revealed that TcElp3 is expressed at the highest level in the 20-day-old larvae and Malpighian tube of 7-day-old females, respectively. RNA interference of TcElp3 delayed the pupation of T. castaneum larvae by two days and led to significantly decreased pupation rate. Notably, knockdown of TcElp3 caused downregulation of ecdysteroid biosynthesis and ecdysone response genes as well as a decrease in ecdysone content in T. castaneum larvae. Further functional characterization of TcElp1, TcElp2, TcElp4, TcElp5 and TcElp6 revealed that knockdown of any of these five subunits of Elongator complex led to similar phenotypes observed in dsTcElp3-injected beetles. These results suggest a possible role of Elongator complex in the epigenetic regulation of T. castaneum ecdysteroid signaling, and provide further evidence in insects that the complete integrity of the Elongator complex is important for its function., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2025. Published by Elsevier B.V.)

Background: An increasing human population, the emergence of resistances against pesticides and their potential impact on the environment call for the development of new eco-friendly pest control strategies. RNA interference (RNAi)-based pesticides have emerged as a new option with the first products entering the market. Essentially, double-stranded RNAs targeting essential genes of pests are either expressed in the plants or sprayed on their surface. Upon feeding, pests mount an RNAi response and die. However, it has remained unclear whether RNAi-based insecticides should target the same pathways as classic pesticides or whether the different mode-of-action would favor other processes. Moreover, there is no consensus on the best genes to be targeted., Results: We performed a genome-wide screen in the red flour beetle to identify 905 RNAi target genes. Based on a validation screen and clustering, we identified the 192 most effective target genes in that species. The transfer to oral application in other beetle pests revealed a list of 34 superior target genes, which are an excellent starting point for application in other pests. Gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) analyses of our genome-wide dataset revealed that genes with high efficacy belonged mainly to basic cellular processes such as gene expression and protein homeostasis - processes not targeted by classic insecticides., Conclusion: Our work revealed the best target genes and target processes for RNAi-based pest control and we propose a procedure to transfer our short list of superior target genes to other pests. © 2024 The Author(s). Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry., (© 2024 The Author(s). Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.)

In this research, the essential oils (EOs) from different parts of Ocimum gratissimum var. suave were chemically characterized and evaluated for insecticidal activity, especially against two common storage pests of Chinese herbal medicines: Tribolium castaneum and Liposcelis bostrychophila. Ocimum gratissimum is a plant with several medicinal values in traditional Chinese medicine. In the study, EOs were successfully extracted from inflorescences (OGI) and stem-leaf (OGLS) parts of O. gratissimum by steam distillation and 16 compounds were identified by gas chromatography-mass spectrometry (GC-MS), of which eugenol was the major constituent in both extracts. In fumigation toxicity tests against both pests, the EOs showed limited toxicity against T. castaneum but showed better toxicity against L. bostrychophila. Contact toxicity tests showed that OGLS had better insecticidal potential than OGI, while the insecticidal effect of eugenol sometimes exceeded that of EOs. In addition, repellency experiments showed that O. gratissimum EOs repelled the pests to varying degrees, with the effect being influenced by concentration and exposure time. The results suggest that O. gratissimum EOs could be a promising alternative to synthetic insecticides for sustainable utilization., (© 2024 Wiley-VHCA AG, Zurich, Switzerland.)