Shouke Zhang

Hyphantria cunea (Drury) causes extensive ecological damage primarily during its larval stages, characterized by voracious feeding and rapid dispersal. Given that conventional dsRNA-mediated RNA interference (RNAi) is generally recalcitrant in Lepidoptera, endogenous microRNAs (miRNAs) may represent an additional class of regulatory molecules worthy of systematic investigation. In this study, we utilized high-throughput sequencing to construct nine comprehensive miRNA libraries across three critical developmental milestones (three biological replicates per instar): the 1st, 4th, and 7th instars (L1, L4, and L7). A total of 1667 miRNA entries were catalogued, including 1080 known and 587 bioinformatically predicted, as yet unvalidated novel miRNA candidates. Comparative transcriptomic analysis revealed 52 differentially expressed miRNAs with significant stage-dependent profiles, with the most pronounced divergence observed between the L1 and L7 groups. Bioinformatic prediction identified 16,784 non-redundant target genes. GO and KEGG enrichment analyses indicated that the predicted target genes of these differentially expressed miRNAs were enriched in developmental and metabolic categories, including cellular development, protein digestion, and nutrient absorption, suggesting that these miRNAs may be associated with tissue remodeling and larval developmental transitions. Collectively, our findings expand the currently available miRNA resource for H. cunea and define stage-associated miRNA expression patterns during larval development. Rather than establishing direct functional roles, this work provides a framework and candidate molecules for future design of RNAi-based biopesticides.

Mountain gradients act as natural experiments, and elevational mitochondrial DNA clines in insects are often interpreted as signatures of local metabolic adaptation. However, mitochondrial DNA is maternally co-inherited with heritable endosymbionts that can promote cytoplasmic hitchhiking and, when abundant, dominate marker-gene microbiome profiles, complicating inference about environmental forcing. Here, we evaluate a symbiont-aware cytoplasmic-axis framework in the tea green leafhopper Empoasca onukii using a densely replicated elevational survey across tea agroecosystems. Across 790 adults from 79 sites spanning 11 to 2750 meters above sea level, we quantified Wolbachia infection prevalence, within-host burden, and strain composition, and related these measures to mitochondrial haplotypes, a conservative nuclear reference marker, and whole-insect bacterial community profiles. Wolbachia prevalence, burden, and strain composition varied along elevation, with pronounced strain turnover. Mitochondrial diversity declined and haplotypes homogenized at high elevations, whereas the nuclear reference marker showed weak spatial structure, yielding mitochondrial–nuclear discordance consistent with cytoplasmic hitchhiking and sweep-like mtDNA homogenization. Bacterial community separation was strongest when Wolbachia features were retained but persisted after Wolbachia removal and renormalization, indicating both compositional dominance by Wolbachia and residual restructuring among non-Wolbachia taxa. In a balanced subset, mitochondrial coding variation and host energetic readouts provided observational functional context for the Wolbachia-centered cytoplasmic-axis pattern. Together, these results place Wolbachia at the center of a testable cytoplasmic framework linking elevational mitochondrial turnover with microbiome restructuring, and they highlight the broader importance of dominant heritable symbionts.

The extent to which herbivorous insects depend on gut bacteria and the molecular mechanisms by which these microbes help overcome host plant chemical defenses remain controversial. This study explored how the gut symbiont Acinetobacter sp. AS23 of a Camellia weevil (Curculio chinensis) (CW) enhances the weevil’s tolerance to toxic tea saponins in host plants. We first conducted toxicity assays in which third-instar CW larvae were exposed to fermentation filtrates containing tea saponin degradation products from the weevil’s gut bacterium, the AS23 strain. A combination of metabolomic and transcriptomic analyses was used to investigate the degradation pathway and key genes used by the AS23 strain in tea saponin metabolism. We then re-inoculated axenic larvae with bacterial mutants generated through CRISPR-Cas9 and verified gene functions in tea saponin degradation. Toxicity assays demonstrated that the AS23 strain exhibited time-dependent tea saponin degradation capabilities. The benzoate degradation pathway emerged as a core metabolic pathway enriched during tea saponin degradation, with the involvement of four key enzyme genes confirmed through qPCR and functional studies. Knockout strains exhibited a significantly reduced detoxification capacity and increased larval mortality when reintroduced into CWs’ gut. Our findings elucidated the key role of the AS23 strain in mediating CW larvae tolerance to tea saponins through the benzoate degradation pathway. This study highlights the potential of leveraging microbial saponin degradation pathways for developing environmentally friendly pest control strategies.

Root rot in Camellia oleifera complicates the development of targeted control measures owing to its complex aetiology. Although breeding resistant varieties of C. oleifera presents a promising solution, research into cultivation strategies and potential resistance mechanisms against root rot remains limited. In this study, we investigated six cultivars of C. oleifera that exhibit varying levels of resistance to root rot. We conducted transcriptome analysis, measurements of soil physicochemical properties and an analysis of the fungal microbiome to explore the relationship between Fusarium‐induced root rot and flavonoid compounds in the rhizosphere. The resistant cultivar CL18 demonstrated superior performance concerning root rot incidence, root health status and the expression levels of genes associated with flavonoid biosynthesis in this study. Significant differences were observed in the composition and diversity of rhizosphere fungal communities among the various cultivars of C. oleifera. The abundance of Fusarium in the rhizosphere soil of CL18 was low, and a negative correlation was identified between the flavonoid content in the soil and the abundance of Fusarium. Our study uncovers the role of flavonoids in the resistance of C. oleifera to root rot, thereby offering new strategies for disease management and the breeding of resistant cultivars.

Bamboo snout moths (Lepidoptera, Crambidae) comprise the four species: Eumorphobotys obscuralis , Circobotys aurealis , Demobotys pervulgalis , and Crypsiptya coclesalis . These economically important insect pests of bamboo are widely distributed in tropical and subtropical regions. The lack of precise mitochondrial genetic data has impeded the development of effective identification techniques, accurate classification strategies, and targeted prevention and treatment strategies. In this study, we obtained the complete mitochondrial genome sequences of four bamboo snout moth species using high‐throughput sequencing. The mitogenomes were 15,103–15,349 bp in length and contained 13 protein‐coding genes, 22 transfer RNA genes (tRNAs), two ribosomal RNA genes (rRNAs), and a noncoding region (A + T rich element), consistent with previously studied Crambidae mitogenomes. We reconstructed the phylogenetic relationships among the four species using Bayesian inference and maximum likelihood methods. The moths that fed on bamboo were well clustered in a single clade. Crypsiptya coclesalis was most closely related to D. pervulgalis , while E. obscuralis was most closely related to C. aurealis . The divergence among the main lineages of 97 Lepidoptera species was reconstructed using an uncorrelated relaxed molecular clock. Analyses of the phylogenetic relationships and divergence times showed that the evolution of lepidopteran species has been closely related to that of their hosts. The data support the development of molecular identification techniques for the four species of bamboo snout moth, and our results provide a basis for targeted control strategies.

Anthracnose, a fungal disease harming fruit trees and crops, poses a threat to agriculture. Traditional chemical pesticides face issues like environmental pollution and resistance. A strategy combining low-toxicity chemicals with biopesticides is proposed to enhance disease control while reducing chemical use. Our study found that mixing validamycin A (VMA) and Bacillus velezensis TCS001 effectively controlled anthracnose in Camellia oleifera. The combination increased antifungal efficacy by 65.62% over VMA alone and 18.83% over TCS001 alone. It caused pathogen deformities and loss of pathogenicity. Transcriptomic analysis revealed that the mix affected the pathogen’s metabolism and redox processes, particularly impacting cellular membrane functions and inducing apoptosis via glycolysis/gluconeogenesis. In vivo tests showed the treatment activated C. oleifera’s disease resistance, with a 161.72% increase in polyphenol oxidase concentration in treated plants. This research offers insights into VMA and TCS001’s mechanisms against anthracnose, supporting sustainable forestry and national edible oil security.