Dai Hirose

The roles of microorganisms in litter decomposition and soil formation

Much has been learned about the microbial decomposition of plant litter, but relatively little is known about microbial contributions to litter and soil chemistry. We conducted a 3-year litterbag experiment and measured hydrolyzable amino acids (AA) and amino sugars (AS) to gain insights about microbial contributions to the chemical characteristics of decomposing litter and soil. Microscopic observations of hyphae were used to estimate fungal contributions to litter. The carbon (C)-normalized yields of AA and AS increased during decomposition along with nitrogen (N), indicating a shift in chemical characteristics from C-rich plant-derived biopolymers to N-rich, microbially-derived biochemicals. The contributions of fungal biomass to C and N were minor, but necromass of fungi as melanized and clamp-bearing hyphae increased during litter decomposition. Yields of glucosamine and galactosamine in litter approached those in microorganisms, particularly bacteria, suggesting major contributions of bacterial residues to litter during decomposition. The microbial contributions to decomposing litter were consistent with those observed in organic and mineral soils. Microorganisms play important roles in the organization and stabilization of soil organic matter as well as N immobilization and organic C preservation.

Fungal colonization and decomposition of Castanopsis sieboldii leaves in a subtropical forest

Fungi play a crucial role in the decomposition of lignin in fallen leaves but few studies have examined the functional roles of ligninolytic fungi associated with the decomposition of fallen leaves on tropical forest soils. This study examined fungal populations responsible for lignin decomposition in Castanopsis sieboldii leaves in a subtropical evergreen broad-leaved forest in southern Japan. Fallen leaves of C. sieboldii are characterized by the occurrence of bleached portions attributable to fungal colonization of leaf tissues and decomposition of lignin. The bleached area accounted for 29.7%, on average, of the total area of C. sieboldii fallen leaves in the study site. Leaf mass per unit area (LMA) and lignin content were lower in the bleached area than in the surrounding nonbleached area of the same leaves, indicating that removal of lignin enhanced mass loss from leaf tissues and created small-scale heterogeneity of decomposition within single leaves. An unidentified species of Lachnocladiaceae (Basidiomycetes) was isolated frequently from the bleached area and caused selective decomposition of lignin in leaves under pure culture conditions, indicating that this fungus was responsible for the bleaching. The greater hyphal length of basidiomycetes in the bleached area than in the nonbleached area supported the finding that this Lachnocladiaceae sp. was associated with the bleaching. The relatively rapid decomposition of C. sieboldii leaves on the subtropical forest soil is partly attributable to colonization of the litter by this Lachnocladiaceae sp.

Contribution of symbiotic mycangial fungi to larval nutrition of a leaf-rolling weevil

Some phytophagous insects have been known to inoculate certain fungi on plant substrates. In many cases of such insect–fungi relationships it has been considered that fungi contribute to insects by decomposing lignin or polysaccharides, and that the insects feed on the decomposition products or fungi themselves. Females of the leaf-rolling weevil in the genus Euops (Attelabidae) store spores of symbiotic fungi in the mycangia and inoculate them on leaf rolls. To determine the effect of mycangial fungi on larval nutrition in E. lespedezae, the nutritional value was compared between leaves with and without mycangial fungi. Two Penicillium species were isolated from the mycangia. These mycangial fungi showed little effect on the decomposition of lignin and polysaccharides, and showed little effect on enhancement of soluble sugars within leaves. Thus, the mutualism between Euops and its mycangial fungi contrasts with the mainly nutritional mutualisms between wood-infesting insects (termites, bark/ambrosia beetles, and wood wasps) and lignin/polysaccharide-decomposing fungi.

Development and seasonal variations of Lophodermium populations on Pinus thunbergii needle litter

A quantitative method was developed to describe Lophodermium (Rhytismataceae, Ascomycetes) populations on Pinus thunbergii needle litter, and seasonal patterns of population variation were examined based on this method. We focused on the formation of black zone lines across needles and the production of Lophodermium ascocarps on needle portions delimitated by zone lines. The study was carried out at a soil erosion control site on a mountain slope and at a coastal sand dune site. Most Lophodermium spp. on needle litter were identified as L. pinastri according to morphological characteristics of ascocarps and ascospores and DNA analysis. Zone lines were produced on needles when isolates of L. pinastri were in contact with other isolates of L. pinastri or with isolates of other species in dual culture tests. This observation provided a rationale to consider that individual colonies with ascocarps and delimitated by zone lines were occupied by a single Lophodermium isolate. Frequency of occurrence of Lophodermium colonies, total colony length, and mean colony number per needle were higher at the coastal sand dune site than at the soil erosion control site. Total colony length and mean colony number also varied with season. Mean colony length and mean ascocarp number per colony were not different between sites or seasons.

Acremomannolipin A, the potential calcium signal modulator with a characteristic glycolipid structure from the filamentous fungus Acremonium strictum

By the newly developed assay method, the glycolipid Acremomannolipin A (1) was isolated from a filamentous fungus Acremonium strictum as a potential calcium signal modulator. The structure of 1 elucidated on the basis of intensive spectroscopic analyses as well as its degradation studies is quite unique: the d-mannopyranose is connected to d-mannitol through a β-glycoside linkage; all the hydroxyls in the mannose are highly masked as peresters with aliphatic acids, and this moiety is made hydrophobic, whereas the mannitol part exhibits a highly hydrophilic property. The compound (1) showed the characteristic bioactivity property, enabling calcineurin deletion cells to grow in the presence of Cl(-), which would be caused by calcium signal modulating. The activity was so potent as to exert the effect at a concentration of 200 nM.

Taxonomic study of the Japanese Dacrymycetes

The class Dacrymycetes is a rather small group of brown-rot wood-decaying jelly fungi characterised by forked basidia and an orange to yellow gelatinous to cartilaginous fruit body. In Japan, dacrymycetous fungi had not been investigated for a long time, justifying a taxonomic re-examination. In the present study we attempted an investigation of the dacrymycetous fungal flora of Japan, and recognised 28 taxonomic entities, including five new taxa, i.e. Dacrymyces ancyleus, D. aureosporus, D. pinacearum, D. subarcticus and Dacryopinax sphenocarpa, and nine new records. Due to the present survey, the total number of dacrymycetous species recorded from Japan increased from 28 to 42. Of the newly described species, Dacrymyces ancyleus is characterised by recurved, cylindrical basidiocarps and hyphae with clamp connections. Dacrymyces aureosporus resembles D. chrysospermus, but differs in wall thickness of its marginal hyphae. Dacrymyces pinacearum and D. subarcticus represent new coelomycetous anamorphic species. Dacryopinax sphenocarpa has sharp, spathulate basidiocarps, and hyphae with clamp connections. Descriptions, illustrations and photographs of fruit bodies are presented with some taxonomic notes. Molecular phylogenetic analyses were conducted to verify the species identification, and the remaining problems in Dacrymycetes taxonomy are discussed based on these data.

Does farm fungicide use induce azole resistance in Aspergillus fumigatus

Azole resistance of Aspergillus fumigatus isolates has been reported worldwide and it would appear to be mainly due to a point mutation in the 14α-sterol demethylase (CYP51A) gene, which is the target enzyme for azoles. The mutation has been confirmed in isolates from patients who received long-term itraconazole (ITZ) therapy and from agricultural fields where high levels of azole fungicides were employed. However, the relationship between farm environments and azole-resistant A. fumigatus has not been fully studied. In this investigation, 50 isolates of A. fumigatus were obtained from a farm where tetraconazole has been sprayed twice a year for more than 15 years. The mean minimum inhibitory concentration (MIC) of isolates was 0.74 (0.19-1.5) mg/L against ITZ, which was below the medical resistance level of ITZ. The sequence of CYP51A from isolates indicated no gene mutations in isolates from the farm. Antifungal susceptibility of isolates to tetraconazole showed that spraying with tetraconazole did not induce resistance to tetraconazole or ITZ in A. fumigatus.

Assessment of the fungal diversity and succession of ligninolytic endophytes in Camellia japonica leaves using clone library analysis

Fungal assemblages in live, newly shed and partly decomposed leaves of Camellia japonica were investigated with a clone library analysis to assess the fungal diversity and succession in a subtropical forest in southern Japan. Partly decomposed leaves were divided into bleached and adjacent nonbleached portions to estimate the fungi functionally associated with lignin decomposition in the bleached portions, with an emphasis on Coccomyces sinensis (Rhytismataceae, Ascomycota). From 144 cloned 28S ribosomal DNA (rDNA) sequences, 48 operational taxonomic units (OTUs) were defined based on a sequence similarity threshold of 98%. Forty-one (85%) of the 48 OTUs belonged to the Ascomycota and seven OTUs (15%) to the Basidiomycota. Twenty-six OTUs (54%) were detected only once (singletons). The number of OTUs and the diversity indices of the fungal assemblages in the different leaves were in this order: live leaves > newly shed leaves > bleached portions > nonbleached portions of partly decomposed leaves. The fungal assemblages were similar in newly shed leaves and the bleached portions of partly decomposed leaves. Ligninolytic fungi of the genera Coccomyces, Lophodermium and Xylaria were frequently detected in the bleached portions. OTU3, identified as Coccomyces sinensis, was detected in live and newly shed leaves and the bleached portions of partly decomposed leaves, suggesting that this fungus latently infects live leaves, persists after leaf fall and takes part in lignin decomposition.

Morphology and phylogeny of four Endogone species and Sphaerocreas pubescens collected in Japan

The order Endogonales within Mucoromycotina includes several ectomycorrhizal species and endophytes of lower land plants. As such, this order is a key taxon for investigation of the evolutionary process of plant–fungal symbioses. However, no taxonomic study of Endogonales based on morphological and phylogenetic data has been performed, and the geographic distribution of this order in Asia remains largely unknown. We collected E. incrassata, E. pisiformis, E. flammicorona, and E. lactiflua from forests in Japan, thus representing their first description in Asia. Collected sporocarps were examined microscopically to observe details of hyphal structures, particularly in zygosporangia and gametangia. In addition, we collected Japanese Sphaerocreas pubescens, recently reported to be related to Endogone-like mycobionts of liverworts, and described its morphology. Species were also analyzed phylogenetically based on large-subunit nuclear ribosomal RNA gene sequences. The phylogenetic relationships of the four Endogone species were supported by their zygosporangial morphogenesis: group A, comprising E. incrassata and E. pisiformis, formed homogametic zygosporangia with pale-colored sporangiothecia, whereas species in group B, E. flammicorona and E. lactiflua, formed heterogametic zygosporangia with orange-colored sporangiothecia. Sphaerocreas pubescens was phylogenetically close to Densospora solicarpa, which is an uncertain zygomycotan ectomycorrhizal fungus; however, the relationship between the “Sphaerocreas/Densospora” clade and Endogone was unclear.

Combined molecular and morphological data for improving phylogenetic hypothesis in Dacrymycetes

We analyzed the DNA sequences of four gene regions, 28S and 18S rDNA, the ITS region and rpb2, to obtain a high resolution phylogenetic tree of Dacrymycetes. In addition, we comparatively studied micro- and macromorphological characteristics of representative species. The traditional generic classification based on morphological characteristics was not reflected by our molecular phylogenies. Ancestral state reconstructions indicated that the morphology of basidia and clamp connections are evolutionarily stable. In contrast, basidiocarps and basidiospore septation patterns appear variable. Dacrymyces unisporus shares the dolipores with non-perforate parenthesomes typical of other dacrymycetous taxa but is a unique species having predominantly non-bifurcate basidia and subglobose to ovoid basidiospores with transverse and longitudinal septa. In molecular phylogenies this species is a member of Dacrymycetes but always occupies a sister position in relation to the rest of the Dacrymycetes. Based on our results we propose a new genus, Unilacryma, for D. unisporus. For proper accommodation of this taxon, we introduce the family Unilacrymaceae and the order Unilacrymales.