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Abstract Arhopalus unicolor is a carrier of the pine wood nematode (PWN), which causes pine wilt disease, killing pine trees and causing considerable economic and environmental losses. While the A. unicolor mitochondrial genome has been published, a high-quality genome assembly and annotation of A. unicolor is not yet available. To address this, we assembled a chromosome-level reference genome assembly of A. unicolor with a combination of Illumina, PacBio, and Hi-C sequencing technologies. The final genome size was determined to be 1268.11 Mb, with a GC% of 32.44%, and the scaffold N50 value was 19.30 Mb. A total of 98.77% of the assembled sequences mapped to 10 pseudochromosomes, and BUSCO analysis revealed high completeness, with 97.15% gene coverage. Furthermore, the genome contains 71.74% repeat elements and encompasses 16,450 predicted protein-coding genes. This genome sequence of A. unicolor will be a valuable resource for understanding the genetics and evolutionary history of this species and for developing effective management strategies for this PWN carrier.

Morphological attributes and chemical composition of host plants shape growth and development of phytophagous insects via influences on their behavior and physiological processes. This research delves into the relationship between Eriogyna pyretorum and various host plants through studuying how feeding on different host tree species affect growth, development, and physiological enzyme activities. We examined E. pyretorum response to three distinct host plants: Camphora officinarum, Liquidambar formosana and Pterocarya stenoptera. Notably, larvae feeding on C. officinarum and L. formosana displayed accelerated development, increased pupal length, and higher survival rates compared to those on P. stenoptera. This underlines the pivotal role of host plant selection in shaping the E. pyretorum’s life cycle. The activities of a-amylase, lipase and protective enzymes were the highest in larvae fed on the most suitable host L. formosana which indicated that the increase of these enzyme activities was closely related to growth and development. Furthermore, our investigation revealed a relationship between enzymatic activities and host plants. Digestive enzymes, protective enzymes, and detoxifying enzymes exhibited substantial variations contingent upon the ingested host plant. Moreover, the total phenolics content in the host plant leaves manifested a noteworthy positive correlation with catalase and lipase activities. In contrast, a marked negative correlation emerged with glutathione S-transferase and α-amylase activities. The total developmental duration of larvae exhibited a significant positive correlation with the activities of GST and CarE. The survival rate of larvae showed a significant positive correlation with CYP450. These observations underscore the insect’s remarkable adaptability in orchestrating metabolic processes in accordance with available nutritional resources. This study highlights the interplay between E. pyretorum and its host plants, offering novel insights into how different vegetation types influence growth, development, and physiological responses. These findings contribute to a deeper comprehension of insect-plant interactions, with potential applications in pest management and ecological conservation.

In this paper, the subgenus Thailandia Bílý, 1990 of the genus Anthaxia Eschscholtz, 1829 from China is reported, including two species: A. (T.) svatoplukbilyi Qi & Song, sp. nov. from Guangxi and A. (T.) rondoni Baudon, 1962 from Yunnan. The description and illustrations of the new species are provided, the illustrations and information of A. (T.) rondoni from Yunnan are given for the first time, and the diagnostic characters are provided to distinguish the new species from other related species.

IntroductionPine wilt disease (PWD) is responsible for extensive economic and ecological damage to Pinus spp. forests and plantations worldwide. PWD is caused by the pine wood nematode (PWN, Bursaphelenchus xylophilus) and transmitted into pine trees by a vector insect, the Japanese pine sawyer (JPS, Monochamus alternatus). Host infection by PWN will attract JPS to spawn, which leads to the co-existence of PWN and JPS within the host tree, an essential precondition for PWD outbreaks. Through the action of their metabolites, microbes can manipulate the co-existence of PWN and JPS, but our understanding on how key microorganisms engage in this process remains limited, which severely hinders the exploration and utilization of promising microbial resources in the prevention and control of PWD.MethodsIn this study we investigated how the PWN-associated fungus Aspergillus promotes the co-existence of PWN and JPS in the host trees (Pinus massoniana) via its secondary metabolite, sterigmatocystin (ST), by taking a multi-omics approach (phenomics, transcriptomics, microbiome, and metabolomics).ResultsWe found that Aspergillus was able to promote PWN invasion and pathogenicity by increasing ST biosynthesis in the host plant, mainly by suppressing the accumulation of ROS (reactive oxygen species) in plant tissues that could counter PWN. Further, ST accumulation triggered the biosynthesis of VOC (volatile organic compounds) that attracts JPS and drives the coexistence of PWN and JPS in the host plant, thereby encouraging the local transmission of PWD. Meanwhile, we show that application of an Aspergillus inhibitor (chiricanine A treatment) results in the absence of Aspergillus and decreases the in vivo ST amount, thereby sharply restricting the PWN development in host. This further proved that Aspergillus is vital and sufficient for promoting PWD transmission.DiscussionAltogether, these results document, for the first time, how the function of Aspergillus and its metabolite ST is involved in the entire PWD transmission chain, in addition to providing a novel and long-term effective nematicide for better PWD control in the field.

IntroductionTea is one of the most widely consumed beverages around the world. Larvae of the moth, Ectropis obliqua Prout (Geometridae, Lepidoptera), are one of the most destructive insect pests of tea in China. E. obliqua is a polyphagus insect that is of increasing concern due to the development of populations resistant to certain chemical insecticides. Microbial biological control agents offer an environmentally friendly and effective means for insect control that can be compatible with “green” and organic farming practices.MethodsTo identify novel E. obliqua biological control agents, soil and inset cadaver samples were collected from tea growing regions in the Fujian province, China. Isolates were analyzed morphologically and via molecular characterization to identity them at the species level. Laboratory and greenhouse insect bioassays were used to determine the effectiveness of the isolates for E. obliqua control.ResultsEleven isolates corresponding to ten different species of Metarhizium were identified according to morphological and molecular analyses from soil and/or insect cadavers found on tea plants and/or in the surrounding soil sampled from eight different regions within the Fujian province, China. Four species of Metarhizium including M. clavatum, M. indigoticum, M. pemphigi, and M. phasmatodeae were documented for the first time in China, and the other species were identified as M. anisopliae, M. brunneum, M. lepidiotae, M. majus, M. pinghaense, and M. robertsii. Insect bioassays of the eleven isolates of Metarhizium revealed significant variation in the efficacy of each isolate to infect and kill E. obliqua. Metarhizium pingshaense (MaFZ-13) showed the highest virulence reaching a host target mortality rate of 93% in laboratory bioassays. The median lethal concentration (LC50) and median lethal time (LT50) values of M. pingshaense MaFZ-13 were 9.6 × 104 conidia/mL and 4.8 days, respectively. Greenhouse experiments and a time-dose-mortality (TDM) models were used to further evaluate and confirm the fungal pathogenic potential of M. pingshaense MaFZ-13 against E. obliqua larvae.DiscussionIsolation of indigenous microbial biological control agents targeting specific pests is an effective approach for collecting resources that can be exploited for pest control with lowered obstacles to approval and commercialization. Our data show the presence of four different previously unreported Metarhizium species in China. Bioassays of the eleven different Metarhizium strains isolated revealed that each could infect and kill E. obliqua to different degrees with the newly isolated M. pingshaense MaFZ-13 strain representing a particularly highly virulent isolate potentially applicable for the control of E. obliqua larvae.

IntroductionBeauveria spp. and Dastarcus helophoroides Fairmaire adults were simultaneously released to attack elder larvae or pupae of Monochamus alternatus in pine forests in China. However, little is known about the pathogenicity virulence and biosafety of Beauveria spp. on beneficial adults of D. helophoroides, and specific Beauveria bassiana (Bb) strains should be selected for synthetic release together with D. helophoroides.MethodsA total of 17 strains of Beauveria spp. were collected, isolated, and purified, and then their mortality, cadaver rate, LT50, spore production, spore germination rate, and growth rate of D. helophoroide adults were calculated based on 0–20 days data after spore suspension and powder contact.Results and discussionThe lethality rate of BbMQ, BbFD, and BbMH-03 strains to D. helophoroides exceeded 50%, and the cadaver rate reached 70.6%, among which the mortality rate (82.22%), cadaver rate (47.78%), spore production (1.32 × 109 spores/ml), spore germination rate (94.71%), colony dimension (49.15 mm2), and LT50 (10.62 d) of the BbMQ strain were significantly higher than those of other strains (P < 0.01), and the mortality of D. helophoroides adults increased significantly with increased spore suspension concentration, with the highest mortality reaching 92.22%. This strain was identified as Beauveria bassiana by morphological and molecular methods, while the BbWYS strain had a minimum lethality of only 5.56%, which was safer compared to other strains of adult D. helophoroide. Consequently, the biological characteristics and pathogenicity of different Beauveria bassiana strains varied significantly in their effects on D. helophoroide adults, and the safety of different strains should be assessed when they are released or sprayed to control multiple pests in the forest. The BbMQ strain should not be simultaneously sprayed with releasing D. helophoroide adults in the same forest, while the BbWYS strain can be used in concert with D. helophoroide to synergize their effect.

Pinewood nematode (PWN), the causal agent of pine wilt disease (PWD), causes massive global losses of Pinus species each year. Bacteria and fungi existing in symbiosis with PWN are closely linked with the pathogenesis of PWD, but the relationship between PWN pathogenicity and the associated microbiota is still ambiguous. This study explored the relationship between microbes and the pathogenicity of PWN by establishing a PWN-associated microbe library, and used this library to generate five artificial PWN–microbe symbiont (APMS) assemblies with gnotobiotic PWNs. The fungal and bacterial communities of different APMSs (the microbiome) were explored by next-generation sequencing. Furthermore, different APMSs were used to inoculate the same Masson pine (Pinus massoniana) cultivar, and multi-omics (metabolome, phenomics, and transcriptome) data were obtained to represent the pathogenicity of different APMSs at 14 days post-inoculation (dpi). Significant positive correlations were observed between microbiome and transcriptome or metabolome data, but microbiome data were negatively correlated with the reactive oxygen species (ROS) level in the host. Five response genes, four fungal genera, four bacterial genera, and nineteen induced metabolites were positively correlated with the ROS level, while seven induced metabolites were negatively correlated. To further explore the function of PWN-associated microbes, single genera of functional microbes (Mb1–Mb8) were reloaded onto gnotobiotic PWNs and used to inoculate pine tree seedlings. Three of the genera (Cladophialophora, Ochroconis, and Flavobacterium) decreased the ROS level of the host pine trees, while only one genus (Penicillium) significantly increased the ROS level of the host pine tree seedlings. These results demonstrate a clear relationship between associated microbes and the pathogenicity of PWN, and expand the knowledge on the interaction between PWD-induced forest decline and the PWN-associated microbiome.

Plant viruses and entomopathogenic fungi (EPF) can both elicit immune responses in insects. This study was designed to clarify whether plant viruses could affect the efficacy of EPF and explore the immune responses of brown planthopper (BPH), Nilaparvata lugens, in response to different pathogen infections. In this study, a strain of Metarhizium anisopliae YTTR with high pathogenicity against BPH was selected and explored whether rice ragged stunt virus (RRSV) could affect its lethality against BPH. RNA-seq was used to detect the inner responses of BPH in response to RRSV and M. anisopliae YTTR infection. Results showed that M. anisopliae YTTR has strong lethality against BPH (RRSV-carrying and RRSV-free). RRSV invasion did not affect the susceptibility of BPH against M. anisopliae YTTR at all concentrations. At 1 × 108 spores/mL, M. anisopliae YTTR caused a cumulative mortality of 80% to BPH at 7 days post-treatment. The largest numbers of differentially expressed genes (DEGs) was obtained in BPH treated with the two pathogens than in other single pathogen treatment. In addition, KEGG enrichment analysis showed that the DEGs were mostly enriched in immune and physiological mechanisms-related pathways. Both RRSV and M. anisopliae YTTR could induce the expression changes of immune-related genes. However, most of the immune genes had varying expression patterns in different treatment. Our findings demonstrated that RRSV invasion did not have any significant effect on the pathogenicity of M. anisopliae YTTR, while the co-infection of M. anisopliae YTTR and RRSV induced more immune and physiological mechanisms -related genes’ responses. In addition, the presence of RRSV could render the interplay between BPH and M. anisopliae YTTR more intricate. These findings laid a basis for further elucidating the immune response mechanisms of RRSV-mediated BPH to M. anisopliae infection.

Prodigiosin is a promising secondary metabolite produced mainly by Serratia strains. To study the global regulatory mechanism of prodigiosin biosynthesis, a mutagenesis library containing 23,000 mutant clones was constructed with the EZ-Tn5 transposon, and 114 clones in the library showed altered prodigiosin production ability. For 37 of the 114 clones, transposon insertion occurred on the prodigiosin biosynthetic cluster genes; transposon inserted genes of the 77 clones belonged to 33 different outside prodigiosin biosynthetic cluster genes. These 33 genes can be divided into transcription-regulating genes, membrane protein-encoding genes, and metabolism enzyme-encoding genes. Most of the genes were newly reported to be involved in prodigiosin production. Transcriptional levels of the pigA gene were significantly downregulated in 22 mutants with different inserted genes, which was in accordance with the phenotype of decreased prodigiosin production. Functional confirmation of the mutant genes involved in the pyrimidine nucleotide biosynthesis pathway was carried out by adding orotate and uridylate (UMP) into the medium. Gene complementation confirmed the regulatory function of the EnvZ/OmpR two-component regulatory system genes envZ and ompR in prodigiosin production.