Cd exposure confers ß-cypermethrin tolerance in Lymantria dispar by activating the ROS/CnCC signaling pathway-mediated P450 detoxification.

Heavy metal pollutants are important abiotic environmental factors affecting pest habitats. In this study, Cd pre-exposure significantly increased the tolerance of Lymantria dispar larvae to ß-cypermethrin, but did not significantly alter their tolerance to λ-cyhalothrin and bifenthrin. The activation of P450 by Cd exposure is the key mechanism that induces insecticide cross-tolerance in L. dispar larvae. Both before and after ß-cypermethrin treatment, Cd exposure significantly increased the expression of CYP6AB224 and CYP6AB226 in L. dispar larvae. Silencing CYP6AB224 and CYP6AB226 reduced the tolerance of Cd-treated L. dispar larvae to ß-cypermethrin. Transgenic CYP6AB224 and CYP6AB226 genes significantly increased the tolerance of Drosophila and Sf9 cells to ß-cypermethrin, and the recombinant proteins of both genes could significantly metabolise ß-cypermethrin. Cd exposure significantly increased the expression of CnCC and Maf. CnCC was found to be a key transcription factor regulating CYP6AB224- and CYP6AB226-activated insecticide cross-tolerance in Cd-treated larvae. Decreasing reactive oxygen species (ROS) levels in the Cd-treated larvae or increasing ROS levels in the untreated larvae reduced or enhanced the expression of CnCC, CYP6AB224 and CYP6AB226 and ß-cypermethrin tolerance in L. dispar larvae, respectively. Collectively, Cd exposure confers ß-cypermethrin tolerance in L. dispar larvae through the ROS/CnCC signalling pathway-mediated P450 detoxification.

Genetics of flight in spongy moths (Lymantria dispar ssp.): functionally integrated profiling of a complex invasive trait.

BACKGROUND: Flight can drastically enhance dispersal capacity and is a key trait defining the potential of exotic insect species to spread and invade new habitats. The phytophagous European spongy moths (ESM, Lymantria dispar dispar) and Asian spongy moths (ASM; a multi-species group represented here by L. d. asiatica and L. d. japonica), are globally invasive species that vary in adult female flight capability-female ASM are typically flight capable, whereas female ESM are typically flightless. Genetic markers of flight capability would supply a powerful tool for flight profiling of these species at any intercepted life stage. To assess the functional complexity of spongy moth flight and to identify potential markers of flight capability, we used multiple genetic approaches aimed at capturing complementary signals of putative flight-relevant genetic divergence between ESM and ASM: reduced representation genome-wide association studies, whole genome sequence comparisons, and developmental transcriptomics. We then judged the candidacy of flight-associated genes through functional analyses aimed at addressing the proximate demands of flight and salient features of the ecological context of spongy moth flight evolution. RESULTS: Candidate gene sets were typically non-overlapping across different genetic approaches, with only nine gene annotations shared between any pair of approaches. We detected an array of flight-relevant functional themes across gene sets that collectively suggest divergence in flight capability between European and Asian spongy moth lineages has coincided with evolutionary differentiation in multiple aspects of flight development, execution, and surrounding life history. Overall, our results indicate that spongy moth flight evolution has shaped or been influenced by a large and functionally broad network of traits. CONCLUSIONS: Our study identified a suite of flight-associated genes in spongy moths suited to exploration of the genetic architecture and evolution of flight, or validation for flight profiling purposes. This work illustrates how complementary genetic approaches combined with phenotypically targeted functional analyses can help to characterize genetically complex traits.

Cd pollution in agroforestry ecosystems: An abiotic factor affecting the passive and active lethal efficiency of Beauveria bassiana to Lymantria dispar larvae.

This study evaluated the effects of cadmium (Cd) exposure on the passive and active lethal efficiency of Beauveria bassiana (Bb) to Lymantria dispar larvae and analyzed the corresponding mechanism from mycelial vegetative growth, fungal and host nutrient competition, and fungal spore performance. The results showed that the passive lethal efficiency of Bb to Cd-exposed L. dispar larvae was significantly higher than that of larvae not exposed to Cd. After Bb infection, the fungal biomass in living larvae and the mycelium encapsulation index of dead larvae were significantly decreased under Cd exposure. Cd exposure damaged the mycelial structure, as well as inhibited the mycelial growth and sporulation quantity. A total of 15 and 39 differentially accumulated mycotoxin metabolites were identified in Bb mycelia treated with low Cd and high Cd, respectively, and the contents of these differentially accumulated mycotoxins in the low Cd and high Cd treatment groups were overall lower than those in the control group. Nutrient content and energy metabolism-related gene expression were significantly decreased in Cd-exposed larvae, both before and after Bb infection. Trehalose supplementation alleviated the nutritional deficiency of larvae under the combined treatment of Cd and Bb and decreased the larval susceptibility to Bb. Compared with untreated Bb, the lethal efficiency of low Cd-exposed Bb to larvae increased significantly, while high Cd-exposed Bb was significantly less lethal to larvae. Cd exposure promoted at low concentrations but inhibited the hydrophobicity and adhesion of spores at higher concentrations. Spore germination rate and stress resistance of Bb decreased significantly under the treatment of both Cd concentrations. Taken together, heavy metals can be regarded as an abiotic environmental factor that directly affects the lethal efficiency of Bb to insect pests.

A new insight into Cd exposure-induced hemocyte reduction in Lymantria dispar larvae: Involvement of the ROS-ATF6-ER stress-apoptosis pathway.

Hemocytes are important targets for heavy metal-induced immunotoxicity in insects. This study aimed to investigate the mechanism by which cadmium (Cd) exposure affects the hemocyte count in Lymantria dispar larvae. The results showed that the number of larval hemocytes was significantly decreased under Cd exposure, accompanied by a significant increase in the apoptosis rate and the expression of Caspase-3. The endoplasmic reticulum (ER) of hemocytes in the Cd-treated group showed irregular swelling. Expression levels of ER stress indicator genes (CHOP, Bip1, Bip2, Bip3, and Bip4) were significantly higher in the Cd-treated group. Among the three pathways that potentially mediate ER stress, only the key genes in the ATF6 pathway (ATF6, S1P-1, S1P-2, and WFS1) exhibited differential responses to Cd exposure. Cd exposure significantly increased the levels of reactive oxygen species (ROS) and the expression of oxidative stress-related genes (CNCC, P38, and ATF2) in hemocytes. Studies using inhibitors confirmed that apoptosis mediated the decrease in hemocyte count, ER stress mediated apoptosis, ATF6 pathway mediated ER stress, and ROS or oxidative stress mediated ER stress through the activation of the ATF6 pathway. Taken together, the ROS-ATF6-ER stress-apoptosis pathway is responsible for the reduction in the hemocyte count of Cd-treated L. dispar larvae.

CncC-Keap1-P450s pathway is involved in the detoxification of emamectin benzoate in the spongy moth Lymantria dispar.

The detoxification of insecticides in insects is dependent on the expression and activity of multiple detoxification enzymes. As an important modulator of detoxification enzymes, the CncC-Keap1 pathway was involved in the detoxification of various pesticides. However, whether the CncC-Keap1 pathway is involved in the detoxification of emamectin benzoate (EMB) is unclear. In this study, we cloned the LdCncC and LdKeap1 from spongy moths (Lymantria dispar). Our results showed that EMB exposure induced oxidative stress, and activated the CncC-Keap1 pathway at mRNA and protein levels. Removing ROS by N-acetylcysteine remarkably decreased H2O2 levels and restored the expression of LdCncC and LdKeap1. The silencing LdCncC, not LdKeap1, by dsRNA significantly decreased the cytochrome P450 activities, and increased the sensitivity of larvae to EMB. Besides, the expression of CYP6B7v1, CYP321A7 and CYP4S4v1 were significantly decreased after silencing LdCncC. Notably, the knockdown of CYP6B7v1, CYP321A7 or CYP4S4v1 significantly increased the mortality induced by EMB exposure. Therefore, we proposed that activation of CncC-Keap1 pathway induced by ROS increased the detoxification of EMB in spongy moths by regulating the expression of CYP6B7v1, CYP321A7 and CYP4S4v1. Our study strengthened the understanding of the detoxification of EMB from the perspective of CncC-Keap1-P450s pathway.

Building a reference indicator model using co-kriging interpolation to determine the geographical origin of the flighted spongy moth complex in China.

Using stable isotopes to detect and analyze the geographical origin of insects represents an important traceability technology, which requires a rich isotope database. In this study, we representatively sampled the Chinese provinces where flighted spongy moth complex (FSMC) has been reported and, for the first time, used co-kriging interpolation to predict the distribution patterns of FSMC δ13C values in the main distribution areas. From 2020 to 2022, we set up 60 traps in 12 provinces and cities in China and collected 795 FSMCs. Then, 6 main climatic factors were obtained by multi-collinearity screening from 21 types of meteorological data collected at the sample plots, and a correlation analysis was carried out by combining longitude, latitude, and altitude data with the δ13C values of FSMC. Next, we performed a co-kriging interpolation using the 2 climatic factors with the highest correlation (isothermality and altitude) and the δ13C values of FSMC. A cross-validation was performed to systematically test 11 candidate models and select the best semi-variogram model ("Exponential"), which was then used to build a co-kriging interpolation model. The geographical distribution patterns of the FSMC δ13C values obtained from the 2 interpolation models (i.e., interpolated with isothermality and altitude, respectively) were almost the same. Moreover, the δ13C values varied significantly at the regional scale, showing regular changes in spatial distribution. Overall, the reference indicator map of the δ13C values generated from stable isotopes can be used to greatly improve the efficiency of discrimination analyses on the geographical origin of FSMC.

Host-specific growth responses of Larix kaempferi and Quercus acutissima to Asian gypsy moth defoliation in central Korea.

As the risk of gypsy moth outbreaks that have detrimental effects on forest ecosystem in the Northern Hemisphere increase due to climate change, a quantitative evaluation of the impact of gypsy moth defoliation is needed to support the adaptive forest management. To evaluate the host-specific impact of gypsy moth defoliation, radial growth and annual carbon accumulation were compared for one severely defoliated (Larix kaempferi (Lamb.) Carrière) and one moderate defoliated (Quercus acutissima Carruth.) host, in defoliated and non-defoliated site using tree-ring analysis. Finally, the resilience indices of radial growth variables were calculated to assess the ability of sampled trees to withstand defoliation. Gypsy moth defoliation mainly decreased latewood width and caused reduction in annual carbon absorption more than 40% for both tree species. However, L. kaempferi, showed the reduced growth until the year following defoliation, while Q. acutissima, showed no lagged growth depression and rapid growth recover. The findings show how each species reacts differently to gypsy moth defoliation and highlight the need of managing forests in a way that takes resilient tree species into account.

Effect of fluorescent brighteners on the insecticidal activity of Bacillus thuringiensis var. kurstaki and LdMNPV on Lymantria dispar asiatica in Korea.

Stilbene-based fluorescent brighteners (FBs) have been demonstrated to improve the insecticidal activities of entomopathogenic viruses; however, there is limited information regarding their effect on entomopathogenic bacteria. We conducted this study to investigate the effect of two FBs (FB 28 and FB 71) on the insecticidal activities of Bacillus thuringiensis var. kurstaki (Btk) and Lymantria dispar multiple nuclear polyhedrosis virus (LdMNPV) on Lymantria dispar asiatica. FB 28 and Btk combination at low concentration (1.6 × 102 IU/mL) increased the mortality, whereas FB 71 and Btk combination at intermediate and high concentrations (1.6 × 103 and 1.6 × 104 IU/mL) slightly reduced the mortality compared with that with Btk alone. The lethal time was also shorter with combinations of Btk and FB 28 than with FB 71. Both FB 28 and FB 71 increased the mortality in combination with LdMNPV at all concentrations (3 × 102 , 3 × 104 , and 3 × 106 polyhedral occlusion bodies/mL compared with that with LdMNPV alone. Our results suggest that FBs improve the insecticidal activities of Btk and LdMNPV, and their activities depend on their interactions with the midgut structures of the host insect species.

Chromosome-level genome assembly of the Asian spongy moths Lymantria dispar asiatica.

The Asian spongy moth, Lymantria dispar asiatica, is one of the most devastating forestry defoliators. The absence of a high-quality genome limited the understanding of its adaptive evolution. Here, we conducted the first chromosome-level genome assembly of L. dispar asiatica using PacBio HIFI long reads, Hi-C sequencing reads and transcriptomic data. The total assembly size is 997.59 Mb, containing 32 chromosomes with a GC content of 38.91% and a scaffold N50 length of 35.42 Mb. The BUSCO assessment indicated a completeness estimate of 99.4% for this assembly. A total of 19,532 protein-coding genes was predicted. Our study provides a valuable genomics resource for studying the mechanisms of adaptive evolution and facilitate an efficient control of L. dispar asiatica.