This study investigated the efficacy of Rotstop^®, a native Latvian Phlebiopsis gigantea strain and 35% urea solution in combination with a stump cover treatment to control against natural spore infection by Heterobasidion spp. upon precommercial thinning of Norway spruce in three stands growing on former agricultural lands. The major findings were that (i) infection rates of Heterobasidion spp. on stumps treated with the native P. gigantea strain, Rotstop^® or urea are similar when stumps are uncovered, and (ii) stump cover promotes stump colonization by the Latvian P. gigantea strain and Rotstop^®, leading to a significantly smaller relative area colonized by Heterobasidion spp., as well greater efficiency against Heterobasidion in comparison with urea. Covering of stumps appears beneficial for controlling Heterobasidion stump colonization and may be valuable to forest owners if used in small-scale operations, but it is impractical in automatized thinnings, where managers should consider using regular Rotstop^® without covering the stumps.

There are several highly damaging Phytophthora species pathogenic to forest trees, many of which have been spread beyond their native range by the international trade of live plants and infested materials. Such introductions can be reduced through the development of better tools capable of the early, rapid, and high-throughput detection of contaminated plants. This study utilized a volatilomics approach (solid-phase microextraction coupled to gas chromatography–mass spectrometry) to differentiate between several Phytophthora species in culture and discriminate between healthy and Phytophthora-inoculated European beech and pedunculate oak trees. We tentatively identified 14 compounds that could differentiate eight Phytophthora species from each other in vitro. All of the Phytophthora species examined, except Phytophthora cambivora, uniquely produced at least one compound not observed in the other species; however, most detected compounds were shared between multiple species. Phytophthora polonica had the most unique compounds and was the least similar of all the species examined. The inoculated seedlings had qualitatively different volatile profiles and could be distinguished from the healthy controls by the presence of isokaurene, anisole, and a mix of three unknown compounds. This study supports the notion that volatiles are suitable for screening plant material, detecting tree pathogens, and differentiating between healthy and diseased material.

Hymenoscyphus fraxineus is an invasive pathogen native to East Asia, responsible for the widespread mortality of European ash ( Fraxinus excelsior ) throughout Europe. Asian ash species, which co-evolved with H. fraxineus , are considered more tolerant than European ash. However, within European ash populations, a small proportion of genotypes show low susceptibility to the pathogen. This study sought to characterize the underlying defence mechanisms to H. fraxineus by performing untargeted constitutive metabolomics profiling of phloem and leaf tissue of from thirteen F. excelsior genotypes (nine tolerant and four susceptible) and five genotypes representing three Asian ash species ( F. mandshurica , F. platypoda , and F. chinensis ). Here we report 57 and 36 compounds associated with lower or higher disease susceptibility, from phloem and leaf tissue, respectively. Flavonoids and coumarins were the main classes of detected compounds. In particular, quercitrin and fraxetin exhibited greater variation among the groups. In phloem tissue, quercitrin and fraxetin were more abundant in tolerant than in susceptible European ash and, lowest in Asian ash species. In leaves, however, quercitrin was highest in Asian ash, followed by tolerant and then susceptible European ash. Other flavonoids, coumarins, and iridoid glycosides also showed variation among groups, with stronger differences in phloem than in leaf tissue. Overall, this study advances our understanding of metabolite composition in Fraxinus species with different co-evolutionary histories and susceptibility to H. fraxineus and demonstrates the potential of untargeted metabolomics for investigating defence-related mechanisms in plant-pathogen interactions.

Many forest pathogens have become increasingly damaging as climate-driven stress intensifies disease outbreaks. A prominent example is Diplodia sapinea, which is causing more frequent and destructive shoot blight and dieback, particularly across northern Europe. Developing resistant planting stock is a priority, but screening currently depends on destructive inoculation assays and multi-year field trials, making the process slow and difficult to scale. This study evaluated whether a handheld Fourier-transform infrared (FT-IR) spectrometer that provides rapid and non-destructive sampling can identify constitutive chemical signatures associated with relative resistance prior to infection. FT-IR spectra were collected from needles, shoots, and stem phloem from Swedish pine families across two experimental years, followed by artificial inoculations to phenotype lesion development. A combined sparse partial least squares discriminant analysis and support vector machine workflow achieved 65–81% cross-validated accuracy in distinguishing resistant from susceptible trees, with shoot spectra consistently producing the strongest models. Principal component analysis indicated clear chemical differences between years, yet resistance-associated patterns were stable across tissues. The most frequently selected wavenumbers grouped into four biochemical domains: cell-wall polysaccharides, cellulose/hemicellulose bonding, phenolics and proteins, and aliphatic lipids/cutin, corresponding to known conifer defense pathways. These findings demonstrate that portable FT-IR spectroscopy can capture biologically meaningful variation in constitutive defense chemistry and offers a rapid, scalable approach for resistance phenotyping in Scots pine breeding programs.

Background: Tree seeds harbor diverse fungal communities, including both pathogens and mutualists, that can influence plant health. These communities comprise living, metabolically active organisms as well as dormant or dead cells. Because only active fungi interact with their hosts, distinguishing active from inactive taxa is crucial, especially for environmental and phytosanitary monitoring. Traditional culturing methods capture living fungi but account for only a small fraction of the total fungal diversity. Currently, these methods are increasingly replaced by high-throughput DNA metabarcoding, which detects a broader range of taxa. However, DNA persists after cell death and occurs in dormant cells, preventing distinction between active and inactive fungi. In contrast, RNA metabarcoding may better reflect living fungal communities than the other two methods, though its use in assessing plant-associated fungi remains underexplored. We used culturing, DNA-, and RNA-based metabarcoding to compare fungal communities associated with seeds of three key European tree species ( Fagus sylvatica , Abies alba , Pinus sylvestris ). Results: Dominant fungal communities in seeds were strongly shaped by host species identity and were largely shared across DNA and RNA metabarcoding datasets, with roughly half of the most abundant genera detected by both methods. Differences between DNA- and RNA-derived communities were predominantly associated with rare taxa in the RNA dataset, although distinguishing true biological signals from noise introduced by different methodological workflows remains challenging. Several cultured genera, likely both abundant and metabolically active, were consistently detected by both approaches. Conclusions: These results highlight the complementary nature of the three methods for characterising seed-associated fungi. Combining culturing, DNA- and RNA-based metabarcoding may provide the most comprehensive assessment of fungal diversity, while RNA metabarcoding alone offers a promising opportunity to identify the active members of fungal communities for improved environmental and phytosanitary monitoring.