Mid Sweden University

Essential oils from plants are often used in growth inhibition assays against bacteria, fungi, insects and viral infections. Turpentine from Norway spruce (Picea abies) is an essential oil produced in large quantities as a waste product in paper production. It is a complex mixture of secondary metabolites that is rich in terpenes and terpenoids. These compounds protect the tree from fungal and bacterial infection; turpentine should therefore be a good source for biologically active substances for inhibiting fungal growth. In this study, a combination of a reductive and a holistic approach was employed to find the most inhibiting compound; an approach that we have called an intermistic approach. The most inhibiting compounds was determined in three steps against growth of the saproxylic fungus Coniophora puteana: rejecting half of the turpentine search space in a reductive manner, finding the optimal concentration and finally combining fractions of the most active half of turpentine in a quadratic D-optimal design to investigate it holistically. Multivariate data analysis of gas chromatography coupled with mass spectrometry (GC-MS) chromatograms was used to zoom in on the most active compounds responsible for antifungal activity. The growth rate of C. puteana in mm/day was used as y-variable and the relative area percentage of 36 GC-MS peaks from the mixtures was used as x-variables to construct a partial least squares (PLS) regression model. The final model was reduced to 12 peaks and used to find the most promising antifungal compound within the mixture. In addition, an external validation was performed with preparatory GC on the most active fraction. From the model, one putative sesquiterpene was identified along with at least two other sesquiterpenes and one diterpene warrant further investigation. Herein, a general method for finding biologically active compounds against wood-rotting fungi, and potentially other organisms from a complex mixture, is proposed.

Heterobasidion parviporum and Gloeophyllum sepiarium are fungal species representing two different ecological niches. H. parviporum is a parasitic white-rot fungus and early colonizer of felled wood, while the brown-rot fungus G. sepiarium is an early-middle colonizer in the natural fungi succession order. In this study, we quantitatively examined the utilization of amino acids in the μM range, pH and final dry weight from common liquid glucose-malt extract medium when the two fungi were interacting at the near-contact level. An increase in glutamine concentration was observed, and lysine was utilized in a greater extent when fungi were interacting compared to when they grew on their own. Our results also show fungal interaction is a process with high variability both in pH, growth and amino acid utilization, depending on the type of interaction. A connection between the growth, according to dry weight, and pH is suggested in the interaction between H. parviporum and G. sepiarium.