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The rare achlorophyllous orchid Cyrtosia septentrionalis (Rchb.f.) Garay is thought to switch mycobiont from a Physisporinus species (‘TK-10’) to an Armillaria species as it develops. We conducted morphological, chemical, and molecular analyses of the root system of three adult plants growing sympatrically in Japan to assess reliance on different mycorrhizae. The root systems were found to comprise several long primary roots that radiate outwards from the emergent stem and many shorter lateral ones. Long roots branched most vigorously where they encountered Armillaria gallica in the soil, and the resulting lateral roots contained the highest density of intra-radical rhizomorphs. One plant was associated with ‘TK-10’ and the other two with A. gallica, but only the apical portion of long roots and a few lateral roots were colonized. Mycobionts isolated from the base of long roots proved incapable of germinating C. septentrionalis seeds. Whereas glucose was the dominant monosaccharide in non-mycorrhizal roots, galactose was in mycorrhizal ones. Lateral roots are specialized in mycorrhization and their velamen, and mucilage may play a key role in preventing desiccation. We hypothesize that C. septentrionalis can associate with either fungus from germination to adulthood, but that detection of A. gallica in the soil can prompt replacement of ‘TK-10’.

Brown root rot disease (BRRD), caused by

Systemic sclerosis (SSc) is a complex disease, in which an interaction of genetic and environmental factors plays an important role in its development and pathogenesis. A number of genetic studies, including candidate gene analysis and genome-wide association study, have found that the associated genetic variants are mainly localized in noncoding regions in the expression quantitative trait locus and influence corresponding gene expression. The gene variants identified as a risk for SSc susceptibility include those associated with innate immunity, adaptive immune response, and cell death, while there are only few SSc-associated genes involved in the fibrotic process or vascular homeostasis. Human leukocyte antigen class II genes are associated with SSc-related autoantibodies rather than SSc itself. Since the pathways between the associated genotype and phenotype are still poorly understood, further investigations using multi-omics technologies are necessary to characterize the complex molecular architecture of SSc, identify biomarkers useful to predict future outcomes and treatment responses, and discover effective drug targets.

Tuber himalayense is a species of edible Asiatic black truffle which is sister to Tuber indicum. Local populations contribute significantly to the reproduction of truffles, but fundamental data of fine-scale genetic structure is lacking in Asiatic truffle species. In this study, we provide the first report on the genotypic diversity and spatial fruiting pattern of an Asiatic truffle T. himalayense, in two spontaneous productive truffle grounds 140 m apart from one another in Yamanashi, Japan. Ascocarps were collected from 2004 to 2009 and in 2011. The spatial distribution of samples was recorded and genotypic diversity of the fungal individuals was examined using 15 newly developed and 4 existing simple sequence repeat (SSR) markers. The spatial distribution analyses suggested non-random, year-specific aggregation patterns of ascocarps in both plots. However, no consistent tendency was observed across the entire sampling period, with fruiting positions showing neither an obvious expansion nor a steady displacement. Although only a single SSR genotype, representing one gleba-forming maternal individual, was detected, the genotype was further divided into two different mating types, suggesting multiple, closely related individuals that were indistinguishable using the SSR analysis. The results indicated strong founding effects along with high inbreeding within populations, likely due to the limited dispersal opportunities into these sites. Despite low genetic diversity in the truffle grounds, the plots remained productive over the sampling years, indicating that low genetic diversity does not necessarily have an adverse effect on truffle production, at least not during a period of several years.

Phellinus noxius, the cause of brown root-rot disease, is an invasive fungal pathogen that causes a white rot among woody plants in Asia, Oceania, and Africa. Because the origin and diversity of this pathogen are unknown, it is difficult to predict its behavior and invasive capacity, especially under future climate-change scenarios. We characterized genetic relationships and ecological differences among P. noxius lineages across eastern Asia and Oceania to better understand evolutionary responses of the pathogen to environmental changes. Sequences of four loci (nuclear large subunit, internal transcribed spacers, partial RNA polymerase II, and partial translation elongation factor – 1 alpha) from 95 P. noxius isolates were used for genetic analyses. Our analyses revealed three genetically distinct lineages of P. noxius: 1) eastern Asia (Japan, Taiwan, Hong Kong, and Malaysia); 2) western Oceania/Japan/Taiwan (including Australia, Palau, Guam, Saipan, Yap, Pohnpei, and Kosrae) with some isolates from Japan and Taiwan; and 3) a distinct group from American Samoa. Population genetic analyses highlighted admixture and migration among the three lineages. Climate-based, species distribution models were used to predict ecological patterns of P. noxius for two genetic lineages: eastern Asia and western Oceania/Japan/Taiwan. Contemporary bioclimatic models depicted potential areas at high risk for P. noxius invasion, and predicted that suitable climate space (potential distribution) is lineage specific. Trade of important economic crops worldwide coupled with changing climates could exacerbate the spread of P. noxius into new geographic areas with suitable habitats for brown root-rot disease.

Phoretic nematodes associated with two mass-occurring populations of the millipede Parafontaria laminata were examined, focusing on Pristionchus spp. The nematodes that propagated on dissected millipedes were genotyped using the D2-D3 expansion segments of the 28S ribosomal RNA gene. Four Pristionchus spp. were detected: P. degawai, P. laevicollis, P. fukushimae, and P. entomophagus. Of the four, P. degawai dominated and it was isolated from more than 90% of the millipedes examined. The haplotypes of partial sequences of mitochondrial cytochrome oxidase subunit I examined for Pristionchus spp. and P. degawai showed high haplotype diversity.

Tricholoma bakamatsutake, which forms its fruit body in oak forests, is similar to T. matsutake in shape and odor; therefore, it is expected to have a high economic value similar to T. matsutake. Recently, the fruit bodies of T. bakamatsutake were produced after planting of a Quercus phillyraeoides sapling with a pure culture of the fungal mycelia. This fungus forms pigmented thick-walled spores (chlamydospores [CSs]) terminally on the hyphae in culture and in the field settings. Abundant CS production in vitro seems to be advantageous for cultivating mycorrhizal trees inoculated with T. bakamatsutake CSs. We investigated the effects of adding 16 different nitrogen (N) compounds (three inorganic and 13 organic) to a culture medium on vegetative hyphal growth and CS formation. The growth of T. bakamatsutake was reduced by many of the N sources used. Conversely, the rate of CS formation was improved by the addition of valine or glutamine.

Phoretic nematodes associated with two mass-occurring populations of the millipede Parafontaria laminata were examined, focusing on Pristionchus spp. The nematodes that propagated on dissected millipedes were genotyped using the D2-D3 expansion segments of the 28S ribosomal RNA gene. Four Pristionchus spp. were detected: P. degawai, P. laevicollis, P. fukushimae, and P. entomophagus. Of the four, P. degawai dominated and it was isolated from more than 90% of the millipedes examined. The haplotypes of partial sequences of mitochondrial cytochrome oxidase subunit I examined for Pristionchus spp. and P. degawai showed high haplotype diversity.

The activity concentrations of radiocesium (

The order Hymenochaetales of white rot fungi contain some of the most aggressive wood decayers causing tree deaths around the world. Despite their ecological importance and the impact of diseases they cause, little is known about the evolution and transmission patterns of these pathogens. Here, we sequenced and undertook comparative genomics analyses of Hymenochaetales genomes using brown root rot fungus Phellinus noxius, wood-decomposing fungus Phellinus lamaensis, laminated root rot fungus Phellinus sulphurascens, and trunk pathogen Porodaedalea pini. Many gene families of lignin-degrading enzymes were identified from these fungi, reflecting their ability as white rot fungi. Comparing against distant fungi highlighted the expansion of 1,3-beta-glucan synthases in P. noxius, which may account for its fast-growing attribute. We identified 13 linkage groups conserved within Agaricomycetes, suggesting the evolution of stable karyotypes. We determined that P. noxius has a bipolar heterothallic mating system, with unusual highly expanded ~60 kb A locus as a result of accumulating gene transposition. We investigated the population genomics of 60 P. noxius isolates across multiple islands of the Asia Pacific region. Whole-genome sequencing showed this multinucleate species contains abundant poly-allelic single-nucleotide-polymorphisms (SNPs) with atypical allele frequencies. Different patterns of intra-isolate polymorphism reflect mono-/heterokaryotic states which are both prevalent in nature. We have shown two genetically separated lineages with one spanning across many islands despite the geographical barriers. Both populations possess extraordinary genetic diversity and show contrasting evolutionary scenarios. These results provide a framework to further investigate the genetic basis underlying the fitness and virulence of white rot fungi.

After the accident at the Fukushima Dai-ichi Nuclear Power Plant in 2011, high activities of radiocaesium have been reported in wild mushrooms in Japan. Fungi play an important role in the dynamics of radiocaesium in forest ecosystems. We examined the contents of caesium (Cs), rubidium (Rb), and potassium (K) in the mycelium of 15 isolates of ectomycorrhizal (EM) fungi and nine isolates of saprotrophic (SA) fungi in a synthetic medium with either ammonium chloride (NH4Cl) or sodium nitrate (NaNO3), supplemented with 1 ppm caesium chloride and rubidium chloride. The mycelia were harvested after 8 weeks of incubation, and the contents of Cs, Rb, and K were measured by inductively coupled plasma mass spectrometry. The dry weight of the mycelium in the medium with NH4 was significantly higher than that with NO3, although some EM species, Hebeloma, Astraeus, Scleroderma, and Pisolithus, grew well in the medium with NO3. Among SA species, Crucibulum and Cyathus grew in the medium with NO3. The uptakes of Cs, Rb, and K by Suillus, Pisolithus, and Rhizopogon were higher than that in other EM and SA species when they grew on the medium with NH4, while the uptakes of these elements by Astraeus and Scleroderma were higher than those by other species grown on the medium with NO3. The content of Rb was positively correlated with Cs (r = 0.85, p

Systemic sclerosis (SSc) is a multi-organ fibrotic disease that affects the skin and various internal organs. Therapeutic strategies for tissue fibrosis have not been established; however, aberrantly activated fibroblasts in affected lesions are key targets for modulating fibrosis. Recently, increased intracellular cyclic GMP (cGMP) levels were demonstrated to improve fibrosis levels in various diseases. The purpose of this study was to assess the anti-fibrotic properties of cGMP in cultured fibroblasts from patients with SSc. The phosphodiesterase (PDE) 5 inhibitor sildenafil increased the intracellular cGMP levels in skin fibroblasts in a dose-dependent manner. Sildenafil treatment also significantly decreased the expression of several pro-fibrotic factors that were upregulated by TGF-β1 treatment in SSc skin fibroblasts. These inhibitory effects occurred via non-canonical TGF-β signaling. Our findings revealed that sildenafil might be a novel strategy to treat tissue fibrosis and vasculopathy in SSc.

The activity concentrations of radiocesium (137Cs) in wild mushrooms are reported to vary according to species, genus or ecological types. In addition, the concentration fluctuates among the same species collected within the same area. Therefore, we investigated whether the 137Cs concentration of wild mushrooms would be (1) spatially biased, or (2) influenced by the 137Cs or exchangeable potassium concentrations in the soils below. We set two survey plots 300 m apart in a Himalayan cedar forest in Tsukuba, Japan, where ca. 30 kBq/m2 of 137Cs was deposited after the Fukushima Nuclear Power Plant accident. From these plots, we collected fruit-bodies (fungal structures for spore production) of co-occurring Boletus hiratsukae, a mycorrhizal species, as well as from the soil below. The mean 137Cs concentrations in the fruit-bodies were significantly different between the two plots, but no difference was observed in the soil 137Cs concentration between the plots. Significant spatial autocorrelation was observed in the 137Cs concentration in the organic layer for both sites, but no significant spatial autocorrelation was observed in the 137Cs of fruit-bodies. Therefore, the variation in the 137Cs concentrations of co-occurred B. hiratsukae was not explained by spatial bias or radioactivity in the below soil. In contrast, the exchangeable potassium concentration in the soil was negatively correlated with the 137Cs in the fruit-bodies. Our results suggest that the 137Cs absorption of wild mushrooms would be suppressed by the competitive effect of exchangeable potassium in the surrounding soils.

Trunk rot, caused by Fomitiporia torreyae, is one of the most economically important sap rot diseases on Cryptomeria japonica, especially on a cutting cultivar ‘Sanbu-sugi’. This disease had been reported only from Chiba and Ibaraki prefectures in Japan; however, a similar trunk rot on Japanese cedars was found recently in Kyoto prefecture. We identified the causal agent of the trunk rot on Japanese cedars in Kyoto prefecture, clarified the genotype of ‘Sanbu-sugi’ (sensu stricto) using microsatellite markers, and then investigated the relatedness of these infected cedars with ‘Sanbu-sugi’. The pathogen was identified as F. torreyae by its DNA sequences and morphological characteristics of the fruiting body. The 25 ramets of ‘Sanbu-sugi’ had only one genotype based on eight microsatellite loci. Infected Japanese cedars in Kyoto were comprised of different genotypes and showed no close relatedness to ‘Sanbu-sugi’. This is the first report of trunk rot caused by F. torreyae on cultivars of Japanese ...

Armillaria possesses several intriguing characteristics that have inspired wide interest in understanding phylogenetic relationships within and among species of this genus. Nuclear ribosomal DNA sequence-based analyses of Armillaria provide only limited information for phylogenetic studies among widely divergent taxa. More recent studies have shown that translation elongation factor 1-α (tef1) sequences are highly informative for phylogenetic analysis of Armillaria species within diverse global regions. This study used Neighbor-net and coalescence-based Bayesian analyses to examine phylogenetic relationships of newly determined and existing tef1 sequences derived from diverse Armillaria species from across the Northern Hemisphere, with Southern Hemisphere Armillaria species included for reference. Based on the Bayesian analysis of tef1 sequences, Armillaria species from the Northern Hemisphere are generally contained within the following four superclades, which are named according to the specific epithet of the most frequently cited species within the superclade: (i) Socialis/Tabescens (exannulate) superclade including Eurasian A. ectypa, North American A. socialis (A. tabescens), and Eurasian A. socialis (A. tabescens) clades; (ii) Mellea superclade including undescribed annulate North American Armillaria sp. (Mexico) and four separate clades of A. mellea (Europe and Iran, eastern Asia, and two groups from North America); (iii) Gallica superclade including Armillaria Nag E (Japan), multiple clades of A. gallica (Asia and Europe), A. calvescens (eastern North America), A. cepistipes (North America), A. altimontana (western USA), A. nabsnona (North America and Japan), and at least two A. gallica clades (North America); and (iv) Solidipes/Ostoyae superclade including two A. solidipes/ostoyae clades (North America), A. gemina (eastern USA), A. solidipes/ostoyae (Eurasia), A. cepistipes (Europe and Japan), A. sinapina (North America and Japan), and A. borealis (Eurasia) clade 2. Of note is that A. borealis (Eurasia) clade 1 appears basal to the Solidipes/Ostoyae and Gallica superclades. The Neighbor-net analysis showed similar phylogenetic relationships. This study further demonstrates the utility of tef1 for global phylogenetic studies of Armillaria species and provides critical insights into multiple taxonomic issues that warrant further study.

Direct observation of hydrogel contact with a solid surface in water is indispensable for understanding the friction, lubrication, and adhesion of hydrogels under water. However, this is a difficult task since the refractive index of hydrogels is very close to that of water. In this paper, we present a novel method to in situ observe the macroscopic contact of hydrogels with a solid surface based on the principle of critical refraction. This method was applied to investigate the sliding friction of a polyacrylamide (PAAm) hydrogel with glass by using a strain-controlled parallel-plate rheometer. The study revealed that when the compressive pressure is not very high, the hydrogel forms a heterogeneous contact with the glass, and a macro-scale water drop is trapped at the soft interface. The pre-trapped water spreads over the interface to decrease the contact area with the increase in sliding velocity, which dramatically reduces the friction of the hydrogel. The study also revealed that this heterogeneous contact is the reason for the poor reproducibility of hydrogel friction that has been often observed in previous studies. Under the condition of homogeneous full contact, the molecular origin of hydrogel friction in water is discussed. This study highlights the importance of direct interfacial observation to reveal the friction mechanism of hydrogels.

The distribution of Phellinus noxius is currently limited to tropical and subtropical regions of Southeast and East Asia, Oceania, Central America, and Africa. The ability of this pathogen to produce disease in temperate climates is unknown. We examined the pathogenicity of Phellinus noxius on four important conifer plantation species grown under temperate conditions in Japan. The inoculation experiment with two isolates of P. noxius from Japan demonstrated clear fungal pathogenicity on the four conifer species, 3 months post-inoculation. Typical disease symptoms ranged from wilting, leaf chlorosis and necrosis, and defoliation resulting in the eventual death of most saplings. Mortality rate varied depending on tree species and the fungal isolate used. The results suggest that the possibility of serious damage if P. noxius invades the temperate zone of Japan (e.g., Kyushu and/or Honshu) from subtropical areas of Japan such as the Ryukyu Islands or foreign countries as global warming progresses.

Objective PM and DM are often complicated by interstitial lung disease (ILD). In this study we aimed to evaluate various serum cytokines in patients with PM/DM with ILD so as to clarify the differences in pathophysiology between anti-melanoma differentiation-associated gene 5 antibody-associated ILD (anti-MDA5-ILD) and anti-aminoacyl tRNA synthetase antibody-associated ILD (anti-ARS-ILD). Methods We evaluated the serum cytokine profiles of 38 patients with PM/DM and compared the cytokine profiles of the non-ILD and ILD subsets as well as the anti-MDA5-ILD and anti-ARS-ILD subsets. Results The myositis intention-to-treat activity index score, which indicates whole disease activity, significantly correlated with serum IL-6, IL-8, TNF-α and IP-10. These cytokine levels were significantly higher in the ILD subset than the non-ILD subset and were lower in the ILD subset following treatment. By multivariate analysis, TNF-α was the most significant cytokine [P = 0.0006, odds ratio (OR) 1.4, CI 1.1, 2.2] associated with PM/DM with ILD. IL-8 levels were significantly higher in anti-MDA5-ILD than in anti-ARS-ILD, although IL-6, TNF-α and IP-10 levels were high in both subsets. IL-8 was the most significant cytokine (P = 0.0006, OR 1.5, CI 1.1, 3.0) associated with anti-MDA5-ILD by multivariate analysis. Moreover, the ratio of IL-4 to IFN-γ was lower in anti-MDA5-ILD than in anti-ARS-ILD. Conclusion IL-6, IL-8, TNF-α and IP-10 are associated with global disease activity in PM/DM. These cytokine levels were high, especially in the ILD subset. Serum IL-8 levels and the balance between IL-4 and IFN-γ may contribute to the differences in pathophysiology between anti-ARS-ILD and anti-MDA5-ILD.

Objective: Adult-onset Still's disease (AOSD) is a clinical entity with heterogeneous etiology. We have encountered patients with AOSD who had severe polyarthritis and who fulfilled the classification criteria for rheumatoid arthritis (RA); however, most patients with AOSD typically exhibit mild arthritis. In this study, we proposed two clinical subsets of AOSD and investigated the clinically significant characteristics of the two subtypes. Methods: We retrospectively analyzed 71 consecutive patients with AOSD. We reviewed the medical records of all patients who were followed up for more than 2 years. We classified all the patients with AOSD into the following 2 subsets: an RA subtype for patients who met the criteria for RA according to the American College of Rheumatology and a non-RA subtype for patients who did not meet the criteria for RA. Results: Our results indicated that the non-RA subtype was accompanied by severe inflammatory complications, including pleuritis and hemophagocytic syndrome. In addition, the serum ferritin and serum IL-18 levels were significantly higher in patients with the non-RA subtype than in those with the RA subtype. Interestingly, only 1 patient with the RA subtype had anti-CCP antibodies, and 1 non-RA subtype patient had rheumatoid factor. These findings distinguish these patients from patients with true RA. Conclusions: There were two subsets of patients with AOSD in the examined population. Patients with high levels of IL-18 or ferritin presented with severe systemic inflammatory disorders (the non-RA subtype), and patients with low levels of IL-18 or ferritin developed severe arthritis (RA subtype).

Many γδ T cells typically bind antigen in a manner similar to that of immunoglobulins, recognizing and directly binding small molecules and intact proteins without presentation by classical MHC:Ag complexes. In this way, γδ T cells combine an innate-like immune response with an adaptive recognition strategy, providing both an immediate response to pathogen invasion and an ongoing, adaptive response to inflammation. However, γδ T cells often recombine tissue-specific, restricted sets of genes that have limited junctional diversity. Thus, somatic hypermutation (SHM) presents a useful solution for diversifying these antibody-like T cell receptors (TCR) by fine-tuning ligand recognition or permitting changes within the loci that allow receptors to evolve more rapidly to changing ligand environments. In fact, recent studies in sandbar shark have confirmed definitively that the γ chain locus of γδ T cells employs SHM as a mechanism to generate more diverse γδ TCR, and follow-up studies in dromedary camel indicated that loci of both γ and δ chains use SHM to diversify γδ TCR. In nurse sharks, we found evidence that SHM acts in the thymus, altering the α chain locus of αβ T cells to broaden the primary αβ T cell repertoire. Further, early results suggest that SHM modifies both γ and δ chain loci (but not β chain locus of αβ T cells) and this alteration may occur in the thymus, indicating that this receptor fine-tuning may precede ligand recognition.

Tricholoma dulciolens is an ectomycorrhizal basidiomycete that produces “matsutake”-like mushrooms in association with Picea abies in Fennoscandia. The phylogenetic position of T. dulciolens relative to matsutake species is, however, unknown. In the present study, we demonstrated that T. dulciolens is phylogenetically independent from T. matsutake and its allied species. In addition, herbarium specimens identified as T. caligatum from Abies alba forests in Spain were phylogenetically related to but distinct from T. dulciolens. Based on ribosomal internal transcribed spacer DNA sequence data, T. dulciolens appears to exist in North America, as well. This is the first phylogeny of T. dulciolens in Fennoscandia and its related species in southern Europe.

The decay risk of airborne wood-decay fungi in the same volume of air was investigated by using an air sampler over the course of a year at three different sampling sites. Japanese cedar disks measuring 7.8 cm in diameter and about 3 mm in thickness, and with a moisture content of about 100 % were placed in a “BIOSAMP” air sampler and then exposed to 1000 l of air in the northern, central, and southwest parts of Japan. The exposed disks were incubated for 20 weeks in a damp container maintained at 26 ± 2 °C and degraded by fungi trapped on the disks. The decay risk was calculated from the mass loss during incubation, and the factors affecting the said risk were explored. The results showed that sampling sites apparently do not affect decay risk, even though the Scheffer’s climate indexes of the sites were quite different. The relation between the sampling month and decay risk reveals that decay risk remains virtually the same year-round. Relative humidity on a sampling day is one of the key factors affecting decay risk in sampling conducted at the central or southwest site. In contrast, no weather factors influenced decay risk at the northern sampling site.

Objective Systemic sclerosis (SSc) is an autoimmune disease for which multiple susceptibility genes have been reported. Genome-wide association studies have shown that large numbers of susceptibility genes are shared among autoimmune diseases. Recently, our group identified 9 novel susceptibility genes associated with rheumatoid arthritis (RA) in a Japanese population. The aim of this study was to elucidate whether the 18 genes that displayed associations or suggestive associations for RA in our previous study are associated with SSc in Japanese. Methods We performed an association study that included 415 patients with SSc and 16,891 control subjects, followed by a replication study that included 315 patients and 21,054 control subjects. The 18 markers reported to display association with RA were analyzed for their associations with SSc in the first study, and 5 markers were further analyzed in the replication study. The inverse variance method was used to evaluate the associations of these markers with SSc in a combined study. Results In the phospholipase D4 gene (PLD4), rs2841277 displayed a significant association with SSc in Japanese patients (P = 0.00017). We observed that rs2841280 in exon 2 of PLD4 was in strong linkage disequilibrium with rs2841277 and introduced an amino acid alteration. We also observed associations between SSc and rs6932056 in TNFAIP3 and rs2280381 in IRF8 (P = 0.0000095 and P = 0.0030, respectively), both of which displayed associations with SSc in a European population. Conclusion We determined that PLD4 is a novel susceptibility gene for SSc in Japanese, thus confirming the involvement of PLD4 in autoimmunity. Associations between SSc and TNFAIP3 or IRF8 were also detected in our Japanese population. SSc and RA appear to share relatively large proportions of their genetic backgrounds.

Agrobacterium tumefaciens-mediated transformation (AMT) was successfully applied to mycelia of the 3 economically important mushrooms Hypsizigus marmoreus, Flammulina velutipes, and Grifola frondosa. We used the hygromycin B resistance gene (hph) under the control of the Cryptococcus neoformans actin promoter. Eighty-six resistant strains of H. marmoreus, 4 of F. velutipes, and 2 of G. frondosa were obtained. All transformants were highly resistant to hygromycin B, suggesting that the C. neoformans actin promoter has a potential universal promoter activity in basidiomycetes. Southern analysis revealed random but single integration of the hph gene.

Tricholoma matsutake (S. Ito & S. Imai) Singer and its allied species are referred to as matsutake worldwide and are the most economically important edible mushrooms in Japan. They are widely distributed in the northern hemisphere and established an ectomycorrhizal relationship with conifer and broadleaf trees. To clarify relationships among T. matsutake and its allies, and to delimit phylogenetic species, we analyzed multilocus datasets (ITS, megB1, tef, gpd) with samples that were correctly identified based on morphological characteristics. Phylogenetic analyses clearly identified four major groups: matsutake, T. bakamatsutake, T. fulvocasta- neum and T. caligatum; the latter three species were outside the matsutake group. The haplotype analyses and median-joining haplotype network analyses showed that the matsutake group included four closely related but clearly distinct taxa (T. matsutake, T. anatolicum, Tricholoma sp. from Mexico and T. magnivelare) from different geographical regions; these were considered to be distinct phylogenetic species.

Objective. The aim of this study was to investigate the precise clinical characteristics and to analyse the association between the anti-MDA5 antibody (anti-MDA5ab) titre and disease status in patients with anti-MDA5ab-positive DM. Methods. Twenty-seven patients who presented with DM and were positive for the anti-MDA5ab were enrolled. The association between the clinical manifestations and the clinical parameters, including the anti-MDA5ab, was analysed. Results. The complication of rapidly progressive interstitial lung disease (RP-ILD) occurred in 20 (74%) patients. The frequencies of fatal outcome, relapse and malignancy were 33, 4 and 4%, respectively. Remarkably, a fatal outcome occurred within the first 6 months. Compared with six non-RP-ILD patients, elderly age at onset, severely involved pulmonary function and high levels of serum ferritin were present in 20 RP-ILD patients with anti-MDA5ab. Alveolararterial oxygen difference (AaDO2) 532 mmHg and ferritin 5828 ng/ml at admission were poor prognostic factors in RP-ILD patients with anti-MDA5ab-positive DM. The median value of the anti-MDA5ab titre on admission was higher in patients who later died than in those who survived. The efficacy of treatment was indicated by the anti-MDA5ab, ferritin and IL-18 concentrations. The decline index of the anti-MDA5ab titre after treatment was lower in the subset of patients who died than in the subset of patients who lived. Sustained high levels of anti-MDA5ab, ferritin and IL-18 were present in the patients who died. Conclusion. Anti-MDA5ab titre and ferritin and IL-18 concentrations are useful for the evaluation of the response to treatment and the status of ILD in patients with anti-MAD5ab-positive DM.

Summary The fungus Phellinus noxius has a broad host range and causes brown root rot in a variety of tree species of various ages, irrespective of their original health. The fungus is widely distributed in tropical countries of Southeast Asia, the Pacific Islands and Australia, Central America and Caribbean, and Africa. Since 1988, when brown root rot was first found on Ishigaki Island, Okinawa Prefecture, the disease has been reported on several islands in Okinawa and Kagoshima Prefectures, and it has been causing serious problems to shade, windbreak, ornamental and landscape trees in the Ryukyu Islands, located in the subtropical region of southern Japan. Here, we report on the current status of P. noxius-caused brown root rot in the Ryukyu Islands, Japan, including symptoms, occurrence, dispersal, distribution and host tree species of the disease, pathogenicity and/or virulence of the pathogen, and disease management based on our surveys and previously published reports from Japan. Brown root rot has been confirmed in 53 tree species from 32 plant families at different sites on 10 of the 12 islands surveyed. Among the 53 tree species, 34 were first recorded in Japan as host plants of P. noxius. The disease occurs mainly in shade, ornamental and windbreak trees at sightseeing places, parks, roadsides, agricultural land such as sugar cane fields, and around residences or other places associated with human activities.

We have optimized a simple and rapid preparation procedure for mushroom DNA extraction from colonies on media or from fruiting bodies for PCR amplification. The protocol combines microwaving twice for 1 min, cooling for 10 min, and centrifuging for 5 min. By using this procedure, more than 100 samples of mushroom DNA can be prepared within 1 h. The DNA obtained can be used for (1) identifying mushroom species by PCR and subsequent sequencing, (2) amplifying low copy number genes (at least 2,000 bp), and (3) screening genetic transformants. This technique will contribute to the mycology of mushroom species.

The relationship between the taxa of airborne fungi and the decay risk was investigated. Airborne fungi in 1,000 l of air were trapped on Japanese cedar disks, and incubated in a damp container kept at 26oC. After 16-week incubation, filamentous fungi grown on the disks were isolated and DNA extracted from each isolate was amplified with the primers ITS4/ITS5. The DNA sequences of the amplified products were determined and compared to the sequence data of GenBank to determine the species or genus according to a BLAST search. This search revealed that the isolate consisted of 5 major taxa, namely Bjerkandera sp., Phanerochaete sp. (A), Phanerochaete sp. (B), Polyporales sp. Polyporus arcularius, and 6 minor ones. Statistical analysis revealed that the major taxa were trapped on the disks in similar weather conditions except for Bjerkandera sp., which was trapped at a cooler temperature. The analysis also proved the disks to which Phanerochaete spp. or Polyporales sp. were attached showed higher mass loss. It is concluded that, under these experimental conditions, related species of Phanerochaete sordida play an important role in increasing the decay risk caused by airborne wood-decay fungi.

A taxonomic study based on morphological and phylogenetic characters revealed that the specimens referred to as ‘Polyporus pseudobetulinus’ in Japan actually represent two distinct species, namely, P. pseudobetulinus and P. subvarius. These cam be distinguished by characteristics of the pileus surface, the presence/absence of clamp-connections on generative hyphae, and the shape of the basidiospores. P. subvarius is newly reported from Japan following the original description from China. The phylogenetic positions, detailed descriptions, and illustrations are provided for both species.

Dermatomyositis (DM) is occasionally complicated by interstitial lung disease. Acute/subacute interstitial pneumonia (A/SIP) with DM is intractable and life threatening. Clinically amyopathic dermatomyositis (C-ADM) is also reported to be complicated with A/SIP, especially in those patients with anti-melanoma differentiation-associated gene 5 (MDA5) antibody. In the present cases, we indicate that serum ferritin level correlated with activity of A/SIP with DM. Two patients, a 65-year-old woman and a 30-year-old woman, were diagnosed with anti-MDA5 antibody-associated A/SIP with DM. Serum ferritin was high, 1600 and 770 mg/dl, respectively, on admission. Immunosuppressive therapy ameliorated A/SIP in both cases. Similarly, serum ferritin was also decreasing. However, A/SIP was recurrent and progressive, and serum ferritin was also increasing again in one case. In conclusion, serum ferritin correlates with disease activity of anti-MDA5 antibody-associated A/SIP with DM. Intensity of treatment may be decided according to serum ferritin level.