Durable resistance to pine wilt disease in Pinus pinaster: evidence of nematode elimination in asymptomatic genotypes

Aim of study: To determine if specific Pinus pinaster trees exhibit resistance or tolerance to pine wilt disease, and to clarify whether artificially inoculated nematodes survive or are eliminated in asymptomatic plants. Area of study: Experiments were conducted in the greenhouse facilities of the Lourizán Forest Research Center (42° 24′ 33″ N, 8° 39′ 46″ W, Pontevedra, Spain). Material and methods: This study involved two sets of assays. In the first set, seedlings from 32 families of the Galician P. pinaster breeding program were artificially inoculated with Bursaphelenchus xylophilus three times over a year. Only 23 plants remained asymptomatic by the assay´s end, and four of these continued to be asymptomatic fifteen months after the final inoculation. At that point, these four ‘mother plants’ were clonally propagated via cuttings. Both the mother plants and their cuttings were subsequently checked for nematode presence at various time points. The second set of experiments comprised three inoculation experiments, evaluating clones derived from nematode-resistant P. pinaster parents of families. At the conclusion of these assays (109-141 days after inoculation), each plant was assigned a symptom stage, and nematode density was assessed. From each assay, 15 asymptomatic plants were sampled for nematode counting at 8 and 18 months post-inoculation. Main results: In the nematode counting performed in mother plants fifteen months following the last inoculation, nematodes were extracted solely from a section below the inoculation point of a single mother plant. Years later, none of the mother plants or their cuttings contained nematodes. In the inoculations experiments with resistant clones, only 16.5% of the plants that remained asymptomatic at the end of the assays contained nematodes. Research highlights: The possibility of producing pine wood nematode-free resistant plants by grafting or cutting is crucial to control pine wilt disease progression.