The National Science Foundation's (NSF) Tokyo Office periodically receives and disseminates reports on research developments in Japan that are related to the Foundation's mission. NSF-sponsored researchers currently working in Japan prepare many of these reports. These reports present information for use by NSF program managers and policy makers; they are not statements of NSF policy. .
Ms. Erica Cline, a Ph.D. student in the College of Forest Resources at the University of Washington in Seattle, Washington, prepared the following report. Ms. Cline was a participant in the 1998 Summer Institute sponsored by NSF/NIH/USDA and the Science and Technology Agency of Japan. Dr. Kazuo Suzuki of the Laboratory of Forest Biology at the University of Tokyo, hosted Ms. Cline. Ms. Cline can be reached via email at: ecline@u.washington.edu
The pine mushroom or matsutake mushroom is highly prized in Japan and represents one of the uniquely Japanese culinary traditions. Tricholoma matsutake`s popularity as a Japanese delicacy makes it one of the most valuable edible mushrooms, with prices sometimes exceeding $500 per kilogram, depending on quality and demand. Japanese production peaked in 1941 but has decreased rapidly since then, partly due to forest decline, particularly of Japanese red pine (Pinus densiflora), matsutake's predominant host tree in Japan. As a result, the majority of Japanese demand is filled by importing T. matsutake and related species from other countries, including China, Korea, Canada, and the United States.
Concern about the status of matsutake in Japan has resulted in intense interest in understanding its ecology. T. matsutake is capable of forming ectomycorrhizae in a symbiotic association with various tree genera in the Pinaceae family, including Pinus, Tsuga, and Picea. Mushroom production occurs in a ring centered around the host tree, known as a "shiro". By marking the location of mushrooms each year, it is possible to trace the expansion of the mycelium through the soil. Locations of individual "shiros" are often closely guarded secrets, and harvesters generally return each year to each known "shiro".
A key area of research is how matsutake spreads from tree to tree, whether vegetatively through the fungal mycelium in the soil, or via airborne or animal-dispersed spores. Landowners sometimes attempt to inoculate forests by spraying spore suspensions or burying fragments of mushroom, but it is not clear whether these approaches are effective. In order to study dispersal of T. matsutake, it is helpful to first develop a means of identifying different strains, so that each strain's dispersal pattern can be traced.
My summer project investigated the effectiveness of molecular approaches in identifying individual strains of T. matsutake, as part of a broader study of its ecology and population dynamics. Our molecular work targeted the intergenic spacer (IGS) which is found between repeats of the ribosomal RNA genes. The IGS is a highly variable region of DNA flanked by the more conserved sequences of the ribosomal genes. Universal primers have been designed to amplify the IGS using the polymerase chain reaction (PCR). Species or strain-level differences in IGS sequences can be detected by using a suite of restriction enzymes to produce a characteristic pattern on an agarose gel. This restriction fragment length polymorphism (RFLP) analysis was used to identify polymorphisms of IGS sequences amplified from two mushrooms and seven cultivated strains of T. matsutake collected from various regions in Japan.
No polymorphisms were detected in strains and basidiomes collected from a single site, however two out of the four strains from unique locations were polymorphic with at least one enzyme. In contrast, the internal transcribed spacer (ITS) of the rRNA gene, a less variable region, showed no polymorphisms among the same samples. We conclude that the ITS is not variable enough to reveal strain-level differences, but the IGS has sufficient intraspecific variability to reveal differences in strains from geographically distant areas in Japan, and therefore could be useful in ecological population-level studies.