This is the report from a BSPP Undergraduate ‘Vacation’ Bursary.
Click here to read more/apply for one yourself.
Lower crown dieback and blackened cankers have become increasingly prevalent on Scots pine across Scotland. A key agent implicated in these symptoms is the ascomycete fungus Curreya pithyophila, which colonises the outer bark layer, particularly at branch junctions. This fungus harbours colonies of Pineus pini, a woolly adelgid that feeds on the trees, creating wound entry points for secondary pathogens such as Crumenulopsis sororia, likely responsible for the blackened cankers. What was previously classified as C. pithyophila is now understood to comprise two distinct species with different spore types: phragmospores and dictyospores. This project aimed to investigate the pathogenic potential and genetic makeup of these species, focusing on their mating loci to better understand their reproductive strategies and ecological roles in Scots pine dieback. The project was conducted at Forest Research’s Northern Research Station near Edinburgh, supervised by Dr Sarah Green.
Pathogenicity tests involved inoculating 100 Scots pine saplings with mycelial plugs of both morphologically distinct forms of C. pithyophila at three sites (main stem, branch junction, and shoot tip). Each site was subjected to varying degrees of damage: light pricking to mimic P. pini feeding, moderate wounding with a thin layer of bark removed, and deeper wounding. Control saplings were inoculated with uncolonised agar plugs. Symptoms were minimal after eight weeks, with no significant differences between fungal types or wounding treatments. These findings align with our hypothesis that C. pithyophila is not a primary pathogen but instead colonises the bark, separating the bark layers facilitating feeding by Pineus pini, which is the primary agent of harm to the trees. We also hypothesise that the fungus benefits from the adelgids, potentially feeding off honeydew secretions, although further evidence is needed to confirm this. Notably, the mechanism by which C. pithyophila initiates P. pini infestations on Scots pine remains unknown and warrants further investigation.
A significant portion of my work focused on characterising the MAT loci of C. pithyophila. Using reference genomes assembled from PacBio sequences, I analysed several whole genomes of isolates representing both the phragmospore and dictyospore types from Illumina sequence data, after thoroughly cleaning and refining the reference genomes. To investigate the mating system, I performed a comprehensive search for MAT loci, drawing on annotated genomes of closely related species. This involved searching for both MAT1-1 and MAT1-2 genes, HMG domain-containing genes, common associated flanking genes, such as SLA2 and APN2, as well as key components of the pheromone response pathway, including STE12 and STE20, which are known to play roles in regulating mating and signal transduction. I identified the MAT1-1-1 gene in both phragmospore and dictyospore types, but no MAT1-2 gene was found. This suggests that C. pithyophila may be heterothallic, with each strain carrying only one mating idiomorph, requiring a complementary strain for reproduction. While MAT1-2 was absent in the current dataset, its existence cannot be ruled out, and further investigation across additional strains is necessary to fully resolve the mating system. To ensure the observed similarity in MAT1-1-1 idiomorphs between strains was not an artifact of assembling the Illumina data against the same reference genome, I also performed a de novo assembly using SPAdes. This allowed me to confirm that the high similarity between strains was not solely due to reference bias.
In parallel with the molecular analyses, I also assisted with field surveys of two native Caledonian pine populations and worked on dating cankers caused by C. pithyophila to understand the timeline of infection and disease development in Scots pine at these sites. This involved analysing cross-sections of cankers and counting the annual growth rings from the edge of the canker to the last full ring, which allowed us to estimate the year the cambium was killed. These analyses are still in progress and the results cannot be presented here.
This project provided me with a wide range of valuable skills in plant pathology, from molecular biology techniques such as DNA extraction, PCR, and bioinformatics analysis, to fieldwork and pathogenicity testing. The experience was incredibly rewarding, and I am truly grateful to the BSPP for funding the opportunity. The findings have contributed to the understanding of a new health threat to Scots pine. I would like to extend my sincere thanks to my supervisors, Dr Sarah Green and Maria Stanisz-Migal, for their guidance, and to Dr Heather Dun and Dr Debbie Frederickson-Matika for their assistance with sample collection and lab work. I am also deeply grateful to Dr Joanne Taylor, whose expertise and insights greatly contributed to the progress of this project. This project has deepened my passion for plant pathology, and I look forward to furthering my research in this field.
Finnley Tierney-Kitchener
This is the report from a BSPP Undergraduate ‘Vacation’ Bursary.
Click here to read more/apply for one yourself.
