The aim of this study was to provide a model for development of healthy box lunches for the middle-aged and elderly. Three types of box lunch were proposed: a box lunch plenty of mushrooms, a box lunch high in calcium and a makunouchi box lunch. We conducted taste testing of the box lunches and a convenience store makunouchi box lunch (CVS box lunch) and surveyed university faculty members and students. The energy density of the three box lunches developed was lower and the salt content was 30-50% less than that of the CVS box lunch, making them healthier. The three box lunches developed scored higher for seasoning and tastiness than the CVS box lunch and the box lunch plenty of mushrooms and the makunouchi box lunch scored especially higher. The energy density of the box lunch plenty of mushrooms was the lowest and it demonstrated an energy density that was low in comparison to its observed volume, though and it provided a feeling of satiety. Mushrooms are suggested to be a suitable food ingredient for the middle-aged and elderly.

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In field trials of in vitro-generated pine ectomycorrhizal seedlings associated with Tricholoma matsutake, we examined the survivability of the ectomycorrhizas on Pinus densiflora seedlings one to three years after outplanting. In total, 55 seedlings (1 to 2 years old) were transplanted to two different P. densiflora experimental forests in Ibaraki Prefecture, Japan, from 2004 to 2008. One to three years after transplantation, the root systems of the seedlings were examined microscopically and their ectomycorrhizal tips were tested for the presence of fungus. T. matsutake was isolated from five seedlings one year after transplantation, and the identity of the fungal species was confirmed by PCR-RFLP analysis of the cultured mycelia. The same analysis showed that T. matsutake survived on one seedling two years after transplantation. These results suggest that T. matsutake ectomycorrhizas synthesized in vitro can survive at least two years after transplantation into a natural pine forest.

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High-resolution melting (HRM) analysis is a closed-tube method for detecting nucleotide polymorphisms, enabling rapid genotyping. In this study, we have developed a rapid strain-typing method for Kinko 115, a premium shiitake cultivar, by HRM analysis. When HRM analyses of tyrosinase (tyr) genes of 64 shiitake cultivars were carried out, 3 cultivars including Kinko 115 could be distinguished from another 61 cultivars. After that, HRM analyses of the ribosomal intergenic spacer (IGS) region in these 3 cultivars revealed that Kinko 115 was distinguished from the other 2 cultivars. Consequently, Kinko 115 could be distinguished from other cultivars by HRM analyses of the tyr gene and the IGS region. Furthermore, Kinko 115 could be distinguished from the others by HRM analyses using not only purified DNA, but also crude DNA extracts from mycelia or gills of dried fruiting bodies. Thus, the HRM analyses developed in this study would be useful for rapidly and easily identifying Kinko 115.

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We evaluated effects of addition of lactic acid bacteria on the quality of fermented apple pomace (FAP), which can be preserved for a long period as a mushroom medium. Sixteen strains belonging to nine species (five Lactobacillus, one Leuconostoc, two Pediococcus and one Lactococcus), were obtained from an institute for microbial strain distribution and evaluated. Results of laboratory-scale fermentation tests using an anaerobic pouch system indicated that the addition of lactic acid bacteria contributes to apple pomace fermentation. Subsequently we evaluated whether FAP can be included in media for bunashimeji (Hypsizygus marmoreus) cultivation. Neither the growth (cultivation days or yield of the fruit body) nor free amino acid composition was affected by a low or moderate level of inclusion of FAP fermented by a Lactobacillus plantarum strain, while mycelia growth was inhibited by excess inclusion of FAP. Our results suggest that FAP can be used in a medium for mushroom cultivation if it is included at up to 9% of the dry matter.

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We utilized autoradiography to visualize the localization of radioactive cesium in the fruiting body and spore print of shiitake mushrooms (Lentinula edodes), grown by the open culture method using wood stored in a restricted habitation area after the Fukushima Dai-ichi nuclear power plant accident. Autoradiography of the cap edge showed higher density compared to that of the stipe. Meanwhile, autoradiography of the spore print revealed its shape. The γ-ray spectrum of the fruiting body showed peaks of 134Cs and 137Cs, while that of the spore print showed peaks of 134Cs, 137Cs and 40K. This indicates that the fruiting body had been contaminated with 134Cs and 137Cs, and greater contamination was distributed at the cap edge compared to the stipe. Moreover, the specific activity was observed to increase toward the cap edge. In addition, it was suggested that the 134Cs and 137Cs localized in the fruiting body were incorporated into the spores.

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