Molecular analysis and physiological roles of subtilases in sapstaining fungi.

摘要

The forest products industry is the largest industry in British Columbia and one of the main industries in Canada. Sapstain fungi cause cosmetic defects in wood and are a major problem to the industry. Millions of dollars are spent each year to prevent the growth of these fungi in sawn wood; however, current control methods are unsatisfactory. Modern chemical treatments have raised environmental concerns and do not have the duration of effectiveness desired by the industry. Kiln drying is expensive and not always practical and the rewetting of wood during transport can allow subsequent fungal growth and stain production. Current biological control agents are also unsatisfactory as they are unreliable and do not grow well in many environments. In order to develop new methods for stain control it is vital to fully understand the relationship between the staining fungi and the host wood. It is likely that the best stain control strategies will integrate these new methods with existing techniques, incorporating the strengths of each.; The nutrient content in wood affects the growth and melanin production of sapstaining fungi, and wood extractives are a major source of nutrients for these fungi. Levels of easily assimilated nitrogen sources in wood are extremely low and organic sources of nitrogen, such as proteins, are vital for the growth of sapstaining fungi. Fungi produce extracellular proteases in order to utilise these nitrogen sources, and these enzymes are likely necessary for fungal growth on wood.; The objective of this thesis was to understand the role of subtilases in the physiology of sapstaining fungi. To achieve this objective several steps were taken. First, a comprehensive study of the distribution and sequence variation of subtilases in sapstaining fungi was carried out. Using the obtained sequences three groups of fungal subtilases were delineated. This is the first report of these groups. Seven full-length sequences were also obtained providing a basis for future work on these genes. Second, the regulation of three representative subtilase genes was determined, extending what is known about the regulation of fungal subtilase genes and providing important information on the potential roles of these genes. Finally, the targeted disruption of opil1 is described. This is the first targeted disruption in any sapstaining species of Ophiostoma. The analysis of the obtained disruptants clearly showed that Opil1 is involved in nutrient acquisition for exogenous protein sources during growth in wood. This suggests that Opil1 could be a useful target for the development of new control strategies. This work also demonstrates the importance of nitrogen in fungal growth on wood, which could lead to better decisions in tree harvesting and be used to assist in the improved application of existing control strategies.