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Introduction: The pinewood nematode (PWN) is responsible for causing pine wilt disease (PWD), which has led to the significant decline of conifer species in Eurasian forests and has become a globally invasive quarantine pest. Manipulating plant-associated microbes to control nematodes is an important strategy for sustainable pest management. However, it has proven difficult to find pine-associated bacteria that possess both nematocidal activity and the ability to colonize pine tissues., Methods: The stress experiments with turpentine and pine tissue extract were carried out to screen for the desired target strain that could adapt to the internal environment of pine trees. This strain was used to construct an engineered nematocidal strain. Additionally, a fluorescent strain was constructed to determine its dispersal ability in Pinus massoniana seedlings through plate separation, PCR detection, and fluorescence microscopy observations. The engineered nematocidal strain was tested in the greenhouse experiment to assess its ability to effectively protect P. massoniana seedlings from nematode infection., Results: This study isolated a Bacillus toyonensis strain Bxy19 from the healthy pine stem, which showed exceptional tolerance in stress experiments. An engineered nematocidal strain Bxy19P3C6 was constructed, which expressed the Cry6Aa crystal protein and exhibited nematocidal activity. The fluorescent strain Bxy19GFP was also constructed and used to test its dispersal ability. It was observed to enter the needles of the seedlings through the stomata and colonize the vascular bundle after being sprayed on the seedlings. The strain was observed to colonize and spread in the tracheid after being injected into the stems. The strain could colonize the seedlings and persist for at least 50 days. Furthermore, the greenhouse experiments indicated that both spraying and injecting the engineered strain Bxy19P3C6 had considerable efficacy against nematode infection., Discussion: The evidence of the colonization ability and persistence of the strain in pine advances our understanding of the control and prediction of the colonization of exogenously delivered bacteria in pines. This study provides a promising approach for manipulating plant-associated bacteria and using Bt protein to control nematodes., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Li, Li, Wang, Zhang, Wen, Liu, Feng and Zhang.)

Salicylic acid (SA) is generally considered to be a critical signal transduction factor in plant defenses against pathogens. It could be converted to methyl salicylate (MeSA) for remote signals by salicylic acid methyltransferase (SAMT) and converted back to SA by SA-binding protein 2 (SABP2). In order to verify the function of SAMT in poplar plants, we isolated the full-length cDNA sequence of PagSAMT from 84K poplar and cultivated PagSAMT overexpression lines (OE-2 isolate) to test its role in SA-mediated defenses against the virulent fungal pathogen Botryosphaeria dothidea . Our results showed that after inoculation with B. dothidea , OE-2 significantly increased MeSA content and reduced SA content which is associated with increased expression of SAMT in both infected and uninfected leaves, when compared against the wild type (WT). Additionally, SAMT overexpression plant lines (OE-2) exhibited higher expression of pathogenesis-related genes PR-1 and PR -5, but were still susceptible to B. dothidea suggesting that in poplar SA might be responsible for resistance against this pathogen. This study expands the current understanding of joint regulation of SAMT and SABP2 and the balance between SA and MeSA in poplar responses to pathogen invasion., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Dong, Zhang, Li, Feng, Wang, Liu, Li, Wen, Ma and Zhang.)

The tree-of-heaven root weevil ( Eucryptorrhynchus scrobiculatus ) and the tree-of-heaven trunk weevil ( Eucryptorrhynchus brandti ) are closely related species that monophagously feed on the same host plant, the Ailanthus altissima (Mill) Swingle, at different locations. However, the mechanisms of how they select different parts of the host tree are unclear. As chemosensory systems play important roles in host location and oviposition, we screened candidate chemosensory protein genes from the transcriptomes of the two weevils at different developmental stages. In this study, we identified 12 candidate chemosensory proteins (CSPs) of E. scrobiculatus and E. brandti , three EscrCSPs, and one EbraCSPs, respectively, were newly identified. The qRT-PCR results showed that EscrCSP7/8a/9 and EbraCSP7/8/9 were significantly expressed in adult antennae, while EscrCSP8a and EbraCSP8 shared low sequence identity, suggesting that they may respond to different odorant molecule binding. Additionally, EbraCSP6 and EscrCSP6 were mainly expressed in antennae and proboscises and likely participate in the process of chemoreception. The binding simulation of nine volatile compounds of the host plant to EscrCSP8a and EbraCSP8 indicated that (1R)-(+)-alpha-pinene, (-)-beta-caryophyllene, and beta-elemen have higher binding affinities with EscrCSP8a and lower affinities with EbraCSP8. In addition, there were seven, two, and one EbraCSPs mainly expressed in pupae, larvae, and eggs, respectively, indicating possible developmental-related roles in E. brandti . We screened out several olfactory-related possible CSP genes in E. brandti and E. scrobiculatus and simulated the binding model of CSPs with different compounds, providing a basis for explaining the niche differentiation of the two weevils., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Wang, Wen, Lu and Wen.)

The key to the coexistence of two or more species on the same host is ecological niche separation. Adult Eucryptorrhynchus scrobiculatus and E. brandti both feed on the tree of heaven, Ailanthus altissima , but on different sections of the plant. Olfaction plays a vital role in foraging for food resources. Chemosensory genes on the antennae, the main organ for insect olfaction, might explain their feeding differentiation. In the present study, we identified 130 and 129 putative chemosensory genes in E. scrobiculatus and E. brandti , respectively, by antennal transcriptome sequencing, including 31 odorant-binding proteins (OBPs), 11 chemosensory proteins (CSPs), 49 odorant receptors (ORs), 17 ionotropic receptors (IRs), 19 gustatory receptors (GRs), and three sensory neuron membrane proteins (SNMPs) in E. scrobiculatus and 28 OBPs, 11 CSPs, 45 ORs, 25 IRs, 17 GRs, and three SNMPs in E. brandti . We inferred that EscrOBP8 ( EscrPBP1 ), EscrOBP24 ( EscrPBP2 ) and EbraOBP8 ( EbraPBP1 ), EbraOBP24 ( EbraPBP2 ) were putative PBPs by the phylogenetic analysis. We identified species-specific OR transcripts (10 EscrORs and 8 EbraORs) with potential roles in the recognition of specific volatiles of A. altissima . In addition to conserved "antennal IRs," we also found several "divergent IRs" orthologues in E. scrobiculatus and E. brandti , such as EscrIR16 , EbraIR19 , and EbraIR20 . Compared with other chemosensory genes, GRs between E. scrobiculatus and E. brandti shared lower amino acid identities, which could explain the different feeding habits of the species. We examined OBP expression patterns in various tissues and sexes. Although amino acid sequence similarities were high between EscrOBPs and EbraOBPs, the homologous OBPs showed different tissue expression pattern between two weevils. Our systematic comparison of chemosensory genes in E. scrobiculatus and E. brandti provides a foundation for studies of olfaction and olfactory differentiation in the two weevils as well as a theoretical basis for studying species differentiation.