Pine wilt disease (PWD), a destructive disease for pine trees, is caused by the pine wood nematode (PWN), Bursaphelenchus xylophilus and additional bacteria. In this study, extracts of Zostera marina showed a high nematicidal activity against PWN and some of the bacteria that it carries. Light yellow crystals were obtained from extracts of Z. marina through solvent extraction, followed by chromatography on AB-8 resin and crystallization. The NMR and HPLC analysis showed that the isolated compound was rosmarinic acid (RosA). RosA showed effective nematicidal activity, of which the LC₅₀ (50% lethal concentration) to PWN at 24 h, 48 h and 72 h was 1.18 mg/g, 1.05 mg/g and 0.95 mg/g, respectively. To get a high yield rate of RosA from Z. marina, single factor experiments and an L₉ (3⁴) orthogonal experiment were performed. This extraction process involved 70% ethanol for 3 h at 40 °C. The extraction dosage was 1:50 (w/v). The highest yield of RosA from Zostera was 3.13 mg/g DW (dried weight). The crude extracts of Zostera marina (10 mg/mL) and RosA (1 mg/mL) also showed inhibitory effects to some bacterial strains carried by PWN: Klebsiella sp., Stenotrophomonas maltophilia, Streptomyces sp. and Pantoea agglomerans. The results of these studies provide clues for preparing pesticide to control PWD from Z. marina.

Polylactic acid (PLA) is a biodegradable polymer made from natural sources, and its electrospinning (e-spinning) nanofiber membrane doped with antibacterial ingredients is widely used in the field of medical dressings. In this research, 9 wt% of rosmarinic acid (RosA) and 0.04 wt% of graphite oxide (GO) with synergistic antibacterial activity were introduced into the e-spinning PLA precursor solution, and the obtained PLA nanofiber membrane showed good antibacterial properties and wound healing effects. At the same time, a nonionic amphiphilic polymer, polyethylene glycol (PEG), was also introduced into this system to improve the hydrophilicity of the e-spinning membrane for wound healing application. The morphological characterization showed the RosA/GO and PEG did not affect the e-spinning of PLA. The tests of mechanical performance and wettability demonstrated that PEG and RosA/GO incorporated in PLA have migrated easily to the surface of the fiber. The e-spun PLA/PEG/RosA/GO membrane showed good antibacterial activity and promoted initial wound healing quickly, which would be a promising application in wound dressing.

A rich source of bioactive secondary metabolites from microorgannisms are widely used to control plant diseases in an eco-friendly way. To explore ideal candidates for prevention of pine wilt disease (PWD), a bacterial strain from rhizosphere of Pinus thunbergii, Lysinimonas M4, with nematicidal activity against pine wood nematode (PWN), Bursaphelenchus xylophilus, was isolated. Two nematicidal compounds were obtained from the culture of Lysinimonas M4 by silica gel chromatography based on bioactivity-guided fractionation and were subsequently identified as 2-coumaranone and cyclo-(Phe-Pro) by nuclear magnetic resonance (NMR) and mass spectrometry (MS). The 2-coumaranone and cyclo-(Phe-Pro) showed significant nematicidal activity against PWN, with LC₅₀ values at 24 h of 0.196 mM and 0.425 mM, respectively. Both compounds had significant inhibitory effects on egg hatching, feeding, and reproduction. The study on nematicidal mechanisms revealed that 2-coumaranone and cyclo-(Phe-Pro) caused the accumulation of reactive oxygen species (ROS) in nematodes, along with a notable decrease in CAT and POS activity and an increase in SOD activity in nematodes, which might contribute to the death of pine wood nematodes. Bioassay tests demonstrated that the two compounds could reduce the incidence of wilting in Japanese black pine seedlings. This research offers a new bacterial strain and two metabolites for biocontrol against PWN.

Pine wood nematode (PWN), Bursaphelenchus xylophilus, is a major pathogen of pine wilt disease (PWD), which is a devastating disease affecting pine trees. Eco-friendly plant-derived nematicides against PWN have been considered as promising alternatives to control PWD. In this study, the ethyl acetate extracts of Cnidium monnieri fruits and Angelica dahurica roots were confirmed to have significant nematicidal activity against PWN. Through bioassay-guided fractionations, eight nematicidal coumarins against PWN were separately isolated from the ethyl acetate extracts of C. monnieri fruits and A. dahurica roots, and they were identified to be osthol (Compound 1), xanthotoxin (Compound 2), cindimine (Compound 3), isopimpinellin (Compound 4), marmesin (Compound 5), isoimperatorin (Compound 6), imperatorin (Compound 7), and bergapten (Compound 8) by mass and nuclear magnetic resonance (NMR) spectral data analysis. Coumarins 1–8 were all determined to have inhibitory effects on the egg hatching, feeding ability, and reproduction of PWN. Moreover, all eight nematicidal coumarins could inhibit the acetylcholinesterase (AChE) and Ca²⁺ ATPase of PWN. Cindimine 3 from C. monnieri fruits showed the strongest nematicidal activity against PWN, with an LC₅₀ value of 64 μM at 72 h, and the highest inhibitory effect on PWN vitality. In addition, bioassays on PWN pathogenicity demonstrated that the eight nematicidal coumarins could effectively relieve the wilt symptoms of black pine seedlings infected by PWN. The research identified several potent botanical nematicidal coumarins for use against PWN, which could contribute to the development of greener nematicides for PWD control.