The Microbiome of Endophytic, Wood Colonizing Bacteria from Pine Trees as Affected by Pine Wilt Disease

Abstract

Pine wilt disease (PWD) is a devastating forest disease present worldwide. In this study we analyzed the effects of the invasion of the pinewood nematode Bursaphelenchus xylophilus, the major pathogen causing PWD, on the endophytic microbiome of adult P. pinaster trees. Wood samples from trees with different degrees of PWD disease were collected at two sites (A and M) in Portugal. Endophytic bacteria were characterized based on directly extracted DNA by fingerprinting and barcoding using the 16S rRNA gene as marker. Furthermore, cultivation-based approaches were used to obtain isolates of the major taxa to study their ecophysiology. The endophytic microbiome from P. pinaster trees differed significantly between the two sampling sites. Main bacterial OTUs belonged to the Proteobacteria (39% (site M) - 97% (site A)), and Firmicutes (0.70% (site A) - 44% (site M)). However, consequences of the invasion with the pathogen were comparable. Interestingly diversity of wood endophytic bacteria increased with the severity of the diseases, with highest diversity levels observed in in the most affected trees. Our results suggest that in the first stages of the disease, the defence mechanisms of plants are repressed by the pathogen, resulting in a colonization of the wood interior by soil microorganisms.