Insect Science

The tropical house cricket Gryllodes sigillatus is a major species used in the edible insect farming industry. Despite the rapid expansion of this sector, diagnostic tools for detecting infections in these species remain limited. The lack of validated reference genes compromises the reliability of RT-qPCR-based gene expression analyses, which are essential for the development of molecular tools for disease diagnosis and health monitoring in insect production systems. To address this gap, we evaluated the expression stability of six candidate reference genes (ACTB, EF1, GAPDH, HisH3, RPL5, and 18SrRNA) across four body parts (abdomen, head, legs, and whole body) using a combination of complementary statistical approaches, including geNorm, NormFinder, BestKeeper, the {Delta}Ct method, the R statistical environment, and the integrated RefFinder tool. Candidate genes were identified and annotated using the recently published G. sigillatus genome, through sequence comparisons with closely related insect species using BLAST and reciprocal BLAST analyses, multiple sequence alignments. All procedures complied with MIQE 2.0 guidelines to ensure methodological rigor and transparency. The results showed that ACTB, EF1, RPL5, and 18SrRNA exhibited stable and consistent expression across all analyzed tissues, whereas GAPDH and HisH3 displayed high variability and were generally unsuitable for normalization, except in head tissue where GAPDH remained stable. This study provides the first validated set of reference genes for G. sigillatus, establishing a robust foundation for accurate, reproducible, and comparable gene expression analyses. Furthermore, these findings support the development of RT-qPCR-based diagnostic tools, contributing to improved health monitoring and biosafety in insect production systems.

Iron is an essential nutrient for all types of organisms, including insects and the microbes that infect them. We predicted that insects fed an iron-supplemented diet would accumulate more iron in their hemolymph, and, because infectious microbes acquire iron from their hosts, that this extra iron would increase the severity of bacterial infections. To test this hypothesis, we studied the effects of dietary iron supplementation on infection outcomes in Manduca sexta (tobacco hornworm). Larvae were fed an artificial diet, with or without antibiotics, or the same diets supplemented with 10 mM iron. Control and iron-treated larvae were inoculated with non-pathogenic Escherichia coli or the entomopathogenic Enterococcus faecalis, and bacterial load and larval survival were measured. We found that dietary iron supplementation increased the iron content of hemolymph by approximately 20 fold; however, contrary to our prediction, this increase in iron did not result in an increase in the bacterial load of either E. coli or E. faecalis. The effect of iron supplementation on survival was more complicated. As expected, for larvae inoculated with nonpathogenic E. coli, iron supplementation had no effect. For larvae inoculated with E. faecalis, the effect of iron supplementation depended on whether antibiotics were present in the diet. Without antibiotics, iron supplementation prolonged larval survival; with antibiotics, iron supplementation decreased larval survival. The results of this study do not support the hypothesis that dietary iron supplementation increases infection severity in M. sexta. Instead, the results support the viewpoint that the relationship between dietary iron and infection outcome is complex.

BACKGROUNDThe two-spotted spider mite, Tetranychus urticae Koch, is a major agricultural pest with a rapid propensity for developing acaricide resistance. Bifenazate targets mitochondrial cytochrome b (CYTB). While the G126S mutation is frequently associated with resistance, its independent role remains unclear as it often occurs with other substitutions. This study explores the molecular basis of bifenazate resistance in a Russian laboratory strain derived from a St. Petersburg greenhouse population. RESULTSDisruptive selection with increasing bifenazate concentrations generated resistant and susceptible isofemale lines. AlphaFold2 structural modeling of CYTB indicated that G126S causes a steric clash, leading to conformational destabilization, whereas other reported mutations primarily affect the ligand-binding pocket. Oxford Nanopore sequencing revealed a very low initial frequency of the G126S allele (<1%; 226/35,895 reads) in the unselected population. After one year of stepwise selection (0.00005-0.031% a.i.), the mutant allele frequency surged to 90% (7,272/8,056 reads). No other known resistance-associated mutations were found in the analyzed cytb fragment. CONCLUSIONWe report the first identification of the G126S mutation in a Russian T. urticae population and demonstrate its rapid fixation under bifenazate selection. Within this genetic background, G126S alone appears sufficient to confer high-level resistance, emphasizing the population-specific nature of resistance evolution and the critical need for local monitoring.

O_LIPhenological shifts are a major ecological consequence of climate change, yet studies often focus on single life stages meaning that the potential for carryover effects between life stages remains poorly understood. Failing to account for these effects may lead to inaccurate estimates of phenological shifts, with consequences for predicted synchrony among interacting species. This is especially relevant for temperate systems where climate warming is occurring unevenly across the year. C_LIO_LIHere, we investigated how temperature experienced the previous autumn and winter (during the pupal and egg stage) influences spring phenology in the winter moth (Operophtera brumata), a herbivorous insect with distinct life stages. Using 50 years of local climate data to create five experimental temperature regimes, we first quantified phenotypic plasticity in the duration and temporal variability of pupal and egg development. We then examined how timing of adult moth emergence affects timing of offspring hatching. C_LIO_LIWe found divergent effects of temperature on different life stages; pupal development time was shortest at intermediate temperatures while egg development time decreased linearly with increasing temperature. Furthermore, phenological shifts due to the conditions experienced by the mother were carried over to influence the phenology of her offspring. While this carryover effect was partially compensated during subsequent stages, compensation decreased under warming conditions. C_LIO_LIThese results refine our understanding of the sensitivity of the annual cycle of winter moth phenology to variation in temperature with potential implications for population dynamics and interspecific interactions. Overall, our findings highlight the need to consider the impacts of warming across multiple life stages so that carryover effects can be properly accounted for. Doing so will improve predictions of phenological shifts under future climates. C_LI

Heat Shock Protein 90 (HSP90) functions as an evolutionary capacitor, allowing populations to store cryptic genetic variation that can be released under stress. While former studies have described the release of morphological variation, its behavioural consequences remain unexplored. In the red flour beetle, Tribolium castaneum, HSP90 inhibition released a phenotype with much smaller, less defined eyes that confers fitness benefits in continuous light and was subsequently assimilated. We hypothesized that altered eye morphology affects light perception and thereby changes light-dependent behaviours. To test whether phenotypes released via evolutionary capacitance can beneficially alter behaviour, we examined locomotor activity rhythm entrainment to light-dark cycles as well as individual and group light choice behaviour. Males of the reduced-eye phenotype exhibited a diminished startle response to sudden light exposure in locomotor activity assays. We also found reduced negative phototaxis in groups of beetles with reduced eyes. This modified behaviour, indicating reduced light sensitivity, may stem from impaired light perception caused by altered eye morphology. Lower light sensitivity could be beneficial under stressful environmental conditions by promoting the exploration of alternative niches. Therefore, this study provides the first evidence for potentially beneficial behavioural changes in a HSP90-released phenotype, reinforcing HSP90s role as an evolutionary capacitor.

MicroRNAs (miRNAs) play essential roles in the posttranscriptional regulation of gene expression in organisms. In the process of synthesizing mature miRNAs from miRNA precursors, the miRNA precursors are cleaved via Dicer at their loop structure, after which the miRNA precursors become mature and regulate transcription. However, the consequences of altering the loop sequence are not fully understood. The silkworm Bombyx mori is a lepidopteran insect with many genetic strains. We identified a mutant of the miRNA miR-3260 whose the part of the loop structure was lacking in a silkworm strain with translucent larval skin. Here, we aimed to analyze the role of wild-type miR-3260 and the influence of the mutation of the loop structure in B. mori. First, we identified the genomic region responsible for the translucent larval skin phenotype and determined that the mutated miR-3260 nucleotide sequences. Then, we predicted the binding partners of wild-type miR-3260 using the RNA hybrid tool and found two juvenile hormone (JH)-related genes as targets of wild-type miR-3260. Next, we assessed the relationships between miR-3260 and JH and found that miR-3260 was highly expressed in the Corpora allata and its expression responded to JH treatment. Meanwhile, miR-3260 mimic and inhibitor did not induce the typical phenotypes associated with JH in B. mori. Then, we compared the dicing products from wild-type and mutant miR-3260 precursors and observed that neither form underwent Dicer-mediated cleavage when the loop structure was altered. These results suggest that loop mutations in the miR-3260 precursor may not influence dicing activity, consistent with the lack of observable phenotypic effects.

Astyanax Baird & Girard, 1854 is a widely distributed and species-rich genus of Acestrorhamphidae, whose abundant populations in Neotropical basins play a crucial ecological role at the trophic level. Taxonomic uncertainties persist within the genus, as seen in Astyanax sp. (formerly designated as A. fasciatus) from the Magdalena basin in Colombia. Concerns about its genetic status are heightened due to ecological threats posed by hydroelectric dams, from habitat loss to river connectivity. We isolated and characterized 17 microsatellite loci to assess the population genetics of this species in a broad sample from the middle and lower sections of the Cauca River, now interrupted by the Ituango dam. Furthermore, a multidisciplinary approach integrating phylogenetic analyses of mitochondrial (COI) and nuclear (rag2) markers with geometric morphometric analyses was employed to evaluate potential cryptic diversity within Astyanax sp. Microsatellites revealed two genetic groups in the studied area, strongly supported as distinct lineages by phylogenetic analyses. Unexpectedly, one of these lineages of Astyanax sp. was recovered in an unresolved clade with samples of A. microlepis and allopatric samples of A. viejita from the Maracaibo Lake basin. Each genetic group showed high genetic diversity, but also evidence of recent bottleneck events and significant-high values of inbreeding. Morphometric analyses provided evidence of significant phenotypic differentiation among A. microlepis, Astyanax sp. 1 (Asp1), and Astyanax sp. 2 (Asp2). Morphological patterns ranged from the robust profile of A. microlepis to the streamlined shape of Astyanax sp. 2 (Asp2), with Astyanax sp. 1 (Asp1) displaying intermediate traits and localized differences in head length and fin placement. Statistical support from permutation tests and a high overall classification accuracy (95.65%) underscore the existence of distinct morphospecies, suggesting that phenotypic differentiation is well-established, despite the complex evolutionary history of the group. This study suggests the presence of cryptic diversity within Astyanax sp. and provides valuable genetic information for the conservation and management of their populations in the Magdalena basin.

Sentinel behaviour occurs when individuals use raised positions to scan for predators while the rest of the group forages. Here, we investigated whether a colonial cooperatively breeding species that forages in large groups, the sociable weaver, Philetairus socius, displays sentinel behaviour. This behaviour has been reported in species with similar ecology, behaviour and foraging habits, (e.g. ground foraging in open habitats where aerial predators are common) and, hence, we expected that it could occur in sociable weavers. On the other hand, sentinel behaviour appears to be less common in species that live in very large groups. We used an experimental set-up consisting of an artificial feeding station and perches to assess occurrence of sentinel related behaviours: (i) perching events > 30s on an elevated position, (ii) head-movements and (iii) alarm calling. Birds were seldom observed perching while others fed, and those that did, perched for periods that were too short to be considered as sentinel behaviour (less than 5s on average). Our results suggest that this behaviour is uncommon or even absent in sociable weavers. We discuss whether other factors such as foraging in very large groups, or interspecific foraging associations might make sentinel behaviour less important in this species.

Judgment bias tasks are increasingly used to assess affective states in animals, yet the extent to which they might reflect transient states or stable traits remains unclear. Here, we tested bumblebees (Bombus terrestris) in an active choice task across three repeated sessions to assess individual consistency in the absence of any manipulation. Bees were trained to associate each of two colours with either a high or a low reward, presented in separate chambers. During testing, they were presented with ambiguous colours. Bees were more likely to choose the high reward chamber and to choose more quickly in response to colours closer to the positive colour. The latency to choose the cues showed significant and moderate repeatability across sessions, suggesting a stable, trait-like underlying component. In contrast, the repeatability of the chamber choices was negligible, indicating that such responses might be largely state-dependent and influenced by situational factors. These findings suggest that judgment biases, particularly as assessed through an active choice task reflect states affected by external factors. Active choice tasks may help disentangle stable behavioural traits from transient affective states in invertebrates.

Understanding the dietary patterns of introduced predators is essential for assessing their impacts on freshwater ecosystems. Here, we investigated the feeding ecology of the invasive Korean perch (Coreoperca herzi) introduced to the Oyodo River system, Japan, by integrating gut content DNA metabarcoding and environmental DNA (eDNA) metabarcoding. Fifty specimens were collected, and prey taxa were identified using metabarcoding targeting fish, aquatic insects, and crustaceans. In parallel, eDNA metabarcoding of habitat water samples was used to assess prey availability and selectivity. The results revealed that the Korean perch prey extensively on aquatic insects and fish. Aquatic insect prey were dominated by epilithic clinger taxa inhabiting stone surfaces, particularly mayflies, suggesting visual-mediated prey selection. Fish predation was frequently detected even in small individuals (<100 mm SL), in contrast to previous studies based on conventional methods, indicating that piscivory begins early and ontogenetic dietary shifts are not pronounced. Furthermore, quantitative fish eDNA analysis showed a positive relationship between eDNA concentrations of prey species and predation frequency, indicating opportunistic feeding on abundant, size-accessible prey. By applying two metabarcoding approaches, this study provides an integrated assessment of prey utilisation and environmental context, highlighting ecological risks posed by the Korean perch to freshwater communities in Japan.

BackgroundThe cotton bollworm Helicoverpa armigera is a major global pest controlled by genetically engineered crops expressing Bacillus thuringiensis (Bt) toxins, including Vip3Aa. While Vip3Aa is widely deployed, the genetic basis of resistance remains poorly understood. Previous work identified disruption of a thyroglobulin-like gene (HaVipR1) as one mechanism of resistance, suggesting additional loci may be involved. ResultsUsing linkage analysis, transcriptomics, long-read sequencing, and CRISPR-Cas9 gene editing, we identify a second thyroglobulin-like gene, HaVipR2, as a novel mediator of Vip3Aa resistance. Resistance in a field-derived H. armigera line was shown to be monogenic, recessive, and autosomal, mapping to chromosome 29. Long-read sequencing revealed a [~]16 kb transposable element insertion disrupting HaVipR2, which was undetectable using standard short-read approaches. CRISPR-Cas9 knockout of HaVipR2 conferred >900-fold resistance, confirming its causal role. Comparative analyses show that HaVipR1 and HaVipR2 share conserved domain architecture, indicating that thyroglobulin-domain proteins represent a recurrent target of resistance evolution. ConclusionsOur findings establish thyroglobulin-domain proteins as a new class of Bt resistance genes in Lepidoptera and demonstrate that transposable element insertions can drive adaptive resistance while evading detection by conventional methods. These results highlight the importance of long-read sequencing and accurate genome annotation for resistance monitoring and provide new insights into the molecular basis and evolution of Vip3Aa resistance.

The coleoid cephalopods (octopus, cuttlefish, and squid) are emerging model organisms for neuroscience, development, and evolutionary biology. Determining their sex early in life is critical for population management and controlled experiments. Here, we present a protocol to non-invasively determine the sex of multiple cephalopod species as young as 3 hours post-hatching using a skin swab and quantitative PCR (qPCR). We describe steps for designing qPCR primers, swabbing live animals, extracting DNA, running the qPCR, and analyzing the results. For complete details on the use and execution of this protocol, please refer to Rubino et al.1 HighlightsO_LISwab live cephalopods as early as 3 hours post-hatching C_LIO_LIExtract DNA from cephalopod skin swabs C_LIO_LIPerform qPCR-based sex determination C_LIO_LIDesign and validate qPCR primers for new species C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=190 SRC="FIGDIR/small/715692v1_ufig1.gif" ALT="Figure 1"> View larger version (43K): org.highwire.dtl.DTLVardef@3aa68dorg.highwire.dtl.DTLVardef@8c7e61org.highwire.dtl.DTLVardef@1bd45d9org.highwire.dtl.DTLVardef@134cc4d_HPS_FORMAT_FIGEXP M_FIG C_FIG

The western honey bee (Apis mellifera) is an essential pollinator facing unprecedented threats from pesticide exposure. While pesticide resistance evolution is well documented in agricultural pests, our understanding of genetic variation in honey bee detoxification systems remains limited. This represents a missed opportunity, as harnessing naturally occurring detoxification diversity could provide new avenues for pollinator protection. Cytochrome P450 monooxygenases (CYPs), which are central to xenobiotic metabolism, offer a promising starting point. Here, we present the first comprehensive analysis of CYP genetic diversity in A. mellifera. We analysed the CYPome of 1,467 individuals representing 18 A. mellifera subspecies from 25 countries and identified 5,756 single-nucleotide polymorphisms (SNPs) in 46 CYP genes. Imputed McDonald-Kreitman testing revealed that 56% of non-synonymous CYP substitutions were driven by positive selection. Of the 1,302 haplotypes identified, 84% resided in CYP3, concentrated in the CYP9 and CYP6AS subfamilies implicated in xenobiotic detoxification. Population-level analysis of nucleotide diversity, Tajimas D selection signatures, FST-based differentiation, and McDonald-Kreitman testing pointed to CYP3 clan genes as the primary locus of adaptive variation. This work provides the first step toward building a comprehensive pharmacogenomic resource for honey bees, enabling the prediction of population-specific pesticide vulnerabilities and leveraging naturally occurring detoxification variants to enhance pollinator resilience - a critical step toward sustainable pollinator management.

Venom is an important functional trait that helps predatory animals capture prey. Centipede predatory venoms are complex cocktails of multiple proteins, such as neurotoxins (scoloptoxins), cytotoxins, {beta}-pore-forming toxins, and enzymes. We examined venom phenotypes in two closely related and co-occurring centipede species, Scolopendra morsitans (n=28) and S. hardwickei (n=11), in peninsular India to determine whether their venoms are similar or dissimilar. An integrated proteo-transcriptomic approach was used to characterise the venom phenotypes of the two species across multiple individuals in peninsular India. We used species occurrence records and species distribution models to assess the distributional overlap among these species within the peninsular Indian region. The species showed significant overlap in their current and projected geographical ranges, corresponding with their co-occurrence. We characterised the venom profiles of both species and found that the venoms were cocktails of enzymes, {beta}-pore-forming toxins, and neurotoxins comprising 110 and 84 proteins in S. morsitans and S. hardwickei, respectively. However, the venom composition of both species differed significantly in toxin abundance and species-specific protein repertoires. This indicates trait divergence in venom phenotypes, suggesting that distinct venom compositions may facilitate coexistence among ecologically similar predatory centipedes. The observed variation in venom phenotypes among co-distributed species opens up important avenues for future research into their ecological roles and functional significance. In this study, we provided a detailed account of venom composition across multiple individuals from the species geographic range and highlighted the importance of investigating the role of venom as a trait that could influence species interactions and shape communities in these diverse tropical forests.

Animals sense and integrate complex external cues to make developmental decisions that help them better survive and adapt to their natural habitats. Under environmental adversity, nematodes can enter an alternative developmental pathway to form a diapautic and stress-resistant stage, termed the dauer larvae. While dauer formation has been well characterized in Caenorhabditis elegans, how environmental factors influence analogous stages in other nematode species remains largely unexplored. This study examines how symbiotic bacteria, temperature, and pheromones affect the formation of the infective juvenile (IJ), a dauer-like stage, of the insect-parasitic nematode Steinernema hermaphroditum. In contrast to C. elegans, where dauer entry is promoted by heat, IJ development in S. hermaphroditum development is enhanced by reduced temperature. Moreover, the presence and absence of live symbiotic bacterium Xenorhabdus griffiniae functions as an ON-and-OFF switch that regulates the host IJ formation. Crude pheromone extracts from S. hermaphroditum liquid culture do not robustly induce IJ formation in a dose-responsive manner, unlike the potent pheromone-driven dauer entry observed in C. elegans. Nutrient-rich liver-kidney media that mimics host insect environment showed IJ entry induction in a pheromone-dependent manner. These data suggest that external cues, such as temperature, microbial diet, and pheromone, are perceived differently by S. hermaphroditum in comparison to that of C. elegans, reflecting species-specific adaptations to distinct ecological niches and life history strategies.

Schistosomiasis is a neglected tropical disease caused by parasitic flatworms of the genus Schistosoma, impacting hundreds of millions of people and animals globally. Disease pathology primarily originates from host immune responses to parasite eggs, which are produced only when female schistosomes are continuously paired with males. Past research focused on pairing-dependent female sexual maturation, while scarce data exist for the males reproductive biology. In this study, we characterized the G protein-coupled receptor Smgpcr9 (Smp_244240), an orphan Class A (Rhodopsin-like) GPCR with a testis-preferential and pairing-influenced expression profile in S. mansoni males. Previous bulk RNA-seq analyses of adult worms and their isolated gonads revealed that Smgpcr9 belongs to a subgroup of GPCR genes with abundant testis-preferential and pairing-influenced transcript levels in males but low and extremely low expression in unpaired and paired females, respectively. This male-/unpaired female-biased expression pattern mirrors that of neuropeptide (npp) genes of S. mansoni such as Smnpp26 and Smnpp41. In a deorphanization approach using yeast-two-hybrid analyses, GPCR internalization experiments, bioluminescence resonance energy transfer assays, and by modeling and docking analyses, we provide first evidence that both NPPs can interact with SmGPCR9. Furthermore, we optimized a GPCR RNAi approach and achieved efficient transcript knockdown (> 90%) enabling robust functional characterization of Smgpcr9. Following RNAi, physiological and morphological analyses revealed that SmGPCR9 regulates key aspects of male reproductive biology like testis morphology and spermatogenesis. Remarkably, ovary structure and egg production were also affected in paired females post RNAi. We observed similar phenotypes plus motility constraints and reduced stem-cell proliferation in both sexes upon RNAi of Smnpp26 and Smnpp41. In all cases, RNAi downstream analyses by RT-qPCR of marker genes substantiated the observed phenotypic effects. These results strongly indicate the importance of SmGPCR9, SmNPP26, and SmNPP41 for spermatogenesis and further physiological processes in male and female S. mansoni. Author SummaryResearch of the reproductive biology of schistosomes focused mainly on females so far, which upon pairing sexually mature to produce eggs that are important for the life cycle maintenance but also for the pathogenesis of schistosomiasis, the infectious disease caused by these parasites. We investigated a yet unknown G protein-coupled receptor, Smgpcr9, which showed a testis-preferential and pairing-influenced expression profile in Schistosoma mansoni males. To this end, we optimized an RNA interference (RNAi) approach for knockdown analysis, identified neuropeptides (NPPs) as potential ligands by different biochemical approaches and modeling and docking analyses, and we investigated the roles of SmGPCR9 and two interacting NPPs, SmNPP26 and SmNPP41, by physiological, microscopical, and molecular techniques. Our results strongly suggest that SmGPCR9 and both NPPs regulate spermatogenesis. Furthermore, we detected effects on ovary morphology, egg production, and stem-cell proliferation of paired females post RNAi. Taken together, we deorphanized SmGPCR9 and showed for the first time the essential role of a so far uncharacterized GPCR and two interacting neuropeptides for spermatogenesis. Our results shed first light on spermatogenesis regulatory processes controlled by GPCRs and neuropeptides in male S. mansoni and thus expand our understanding of the roles of GPCR-NPP signaling for schistosome reproductive biology.

Methods for estimating and modeling the long-term and short-term adult flight dynamics of the conifer silk moth Dendrolimus superans (Lepidoptera: Lasiocampidae) are examined. The analysis uses light trap adult catch data collected over 21 years, from 2005 to 2025. Three models of adult flight are considered: a flight-initiation model driven by weather factors, an autoregressive model of long-term catch dynamics, and a binary model of seasonal catch. For the flight-initiation model, we propose estimating the accumulated temperature sum ST from the date when the first derivative of the remote sensing vegetation index NDVI becomes positive until the date of the first adult capture of the season. ST is shown to be sufficiently stable across all years of observation, with flight each year beginning after this temperature sum is reached. The second model demonstrates that the long-term light trap catch time series is well described by a second-order autoregressive model AR(2), in which the catch of the current year depends on catches from the two preceding years. This long-term series is compared with a previously studied larval population density series of the Siberian silk moth; both are shown to be AR(2) series with similar coefficient values, which suggesting that adult catch data may serve as a proxy for absolute larval population density. In the third model, we describe the transition from absolute-scale seasonal catch dynamics (number of adults per day) to a binary scale (0, 1), where 0 denotes days on which no adults were attracted to the trap, and 1 denotes days on which at least one individual was captured. The seasonal absolute catch series is thereby transformed into a binary series of zeros and ones, and relationships between adjacent values in such a binary series are examined. A linear relationship between the absolute and binary seasonal dynamics series is demonstrated, making it possible to estimate absolute catches from binary catch values and to analyze seasonal flight in sparse pest populations. This potentially opens new avenues for understanding how outbreak populations function at chronically low density. Author summaryForest pests can cause catastrophic damage, yet predicting their outbreaks remains challenging. During periods of low population density, standard monitoring methods become labor-intensive and uninformative, while the transition to an outbreak often occurs unexpectedly. Using a 21-year dataset of adult Siberian silk moth (Dendrolimus superans) captures from light traps, we developed an approach combining three complementary models. First, we showed that moth flight begins upon reaching a specific temperature sum, with the starting point determined by NDVI vegetation index dynamics rather than a calendar date--making the forecast more ecologically relevant. Second, long-term adult population dynamics follow a second-order autoregressive model AR(2), matching the dynamics previously observed for larval populations. This establishes light trap data as a reliable proxy for absolute population density when ground surveys are impractical. Third, we introduced a method to analyze seasonal flight using binary data (presence/absence of moths per day), which we showed is linearly related to absolute abundance. This enables studying population dynamics during periods of extremely low density, when traditional methods fail. Our approach opens new possibilities for early warning systems to detect when a population risks transitioning from a latent state to an outbreak phase.

The common pine sawfly, Diprion pini, is a widespread defoliator of pine forests across Europe and Asia, with outbreaks causing substantial ecological and economic damages. However, genomic resources for this species have been limited, hindering advances in molecular ecology or pest management. Here, we present a near chromosome-level reference genome for D.pini, generated using PacBio HiFi reads, Oxford Nanopore MionION long reads, and 10x Genomics linked reads. The final assembly is organized into mostly chromosome-sized scaffolds. It spans a length of 268 Mb, comprises 81 scaffolds, and has a scaffold N50 of 18.7 Mb. BUSCO analysis (hymenoptera_odb10) indicates a high genome completeness of 97.2%. With 22,7 kb the mitochondrial genome is unusually large due to an extended non-coding control region (6,874 bp). Gene prediction identified 26,335 protein-coding genes, of which 12,769 were functionally annotated. Comparative analyses with other sawflies and Apocrita identified 2,472 proteins unique to D. pini, some of which are putatively associated with the processing of plant secondary metabolites. Notably, our genome assembly highlights that, when a closely related, high-quality reference genome is available, chromosome-scale assemblies can be generated without the need of Hi-C sequencing. The genome provides a valuable foundation for the development of improved monitoring and management strategies for D. pini outbreaks and contributes to advancing fundamental research on Hymenoptera evolution.

Capturing and handling wild animals is essential for ecological and evolutionary research, yet their effects on physiology, behaviour, and reproductive success remain poorly understood. We investigated short- and longer-term consequences of a capture-handling-restraint protocol in wild great tits (Parus major) over three breeding seasons. To assess short-term responses, we measured circulating corticosterone, a metabolic hormone that responds to unpredictable challenges, and automatically recorded provisioning behaviour. We also explored whether environmental and individual traits were related to provisioning latency (i.e., time to resume provisioning after capture). To evaluate longer-term effects, we monitored provisioning in the days following capture and related it to reproductive success (fledgling number and body condition). We predicted that longer handling would increase stress-induced corticosterone and provisioning latency, that these variables would be positively correlated, and that higher corticosterone and longer latencies would be associated with lower reproductive success. After capture, great tits showed elevated corticosterone and delayed provisioning. Contrary to our predictions, handling duration was negatively associated with stress-induced corticosterone in males (but not females) and did not affect provisioning latency. Provisioning latency was unrelated to corticosterone, environmental, or individual variables. Following capture, parents resumed provisioning, and short-term responses had little influence on reproductive success. We show that parental behaviour and physiology are affected by capture restraint protocols on the short term, but offspring condition and survival are not. However, these results should be interpreted cautiously, as our study lacks an uncaptured control group. Our findings highlight that evaluating welfare impacts requires rigorous study design incorporating both immediate and longer-term behavioural and fitness effects.

Stingless bees are key pollinators in tropical ecosystems, yet their ecological dynamics remain poorly understood in highly seasonal environments such as the seasonally dry tropical forests of Ecuador. These ecosystems experience pronounced climatic seasonality, with sharp transitions between dry and wet periods that strongly affect floral resource availability. Understanding interspecific competition and niche partitioning in such systems is critical, particularly given the global decline of pollinators. We investigated resource use and niche dynamics in two native stingless bees, Melipona mimetica and Scaptotrigona sp., by quantifying pollen, nectar, and resin collection across seasons. Log-linear models were used to test the effects of species, season, and their interaction on resource use, while non-metric multidimensional scaling (NMDS) assessed niche overlap. Contrary to the expectation that niche overlap increases under resource scarcity, we found greater overlap during the wet season, when resources are more abundant. This suggests that both species converge on high-quality floral resources during peak availability, reflecting an adaptive response to strong environmental seasonality. Pollen use remained stable across seasons, consistent with generalist foraging behavior. In contrast, nectar collection increased significantly during the wet season, while resin exhibited a shared seasonal peak, likely associated with synchronized nest construction or maintenance. These findings reveal context-dependent competition dynamics and highlight the role of environmental seasonality in shaping pollinator interactions. Our study provides new insights into the ecology of threatened stingless bees and contributes to their conservation in tropical dry forest ecosystems.