Esteya spp. reduce Bursaphelenchus xylophilus reproduction in Pinus pinaster seedlings without consistent improvements in disease outcomes.

BACKGROUND: Pine wilt disease, caused by Bursaphelenchus xylophilus, threatens pine forests worldwide, yet sustainable biological control strategies remain limited. We evaluated the effects of pre-emptive inoculation with Esteya vermicola and E. floridanum on nematode reproduction and disease outcomes in Pinus pinaster seedlings under greenhouse conditions. RESULTS: Seedlings inoculated with E. vermicola or E. floridanum remained free of visible symptoms during the 5-week pre-challenge period. Fungi were subsequently re-isolated from stem tissues at the experimental endpoint, and both treatments significantly reduced nematode reproduction relative to the inoculated control. However, reductions in nematode reproduction were not consistently accompanied by improvements across disease assessment endpoints, including symptom severity, disease progression, and survival. Symptom expression in nematode-free controls indicated that environmental stress contributed to disease assessment endpoints, limiting interpretation of symptom-based measures. Exploratory correlation and permutation analyses suggested treatment-specific differences in the association between nematode reproduction and symptom severity; however, the endpoint design and restricted range of reproduction values following fungal treatment do not distinguish this interpretation from alternative explanations. CONCLUSION: Esteya spp. significantly reduced pinewood nematode reproduction in P. pinaster seedlings, but improvements in disease outcomes were inconsistent under the conditions tested. These findings highlight that pathogen reproduction and disease assessment endpoints should both be evaluated when assessing biological control strategies for pine wilt disease. © 2026 Society of Chemical Industry.