Changes in mycorrhizal colonization and overall fungal communities across upland and wetland <Emphasis Type="Italic">Acer</Emphasis> forests of the Virginia Coastal Plain

Arbuscular mycorrhiza (AM) are common in the roots of most plants and provide benefits such as enhanced uptake of soil nutrients and water when the environment limits these resources. Two primary factors proposed in the literature as limiting the extent of mycorrhizal colonization of wetland plants are: (1) the low oxygen environment; and (2) the level of available phosphorus. The present study evaluated AM colonization of maple (Acer) roots and the fungal community structure in paired upland/wetland locations in the Virginia Coastal Plain to determine if wetland conditions affected the root fungal community. The range of observed AM colonization, based on 60 samples collected on 11 dates covering 3 years, was from 4 to 55%. Overall, soil redox potential and available phosphorus concentration were not significant in explaining the difference in AM colonization of Acer roots. In fact, the site with the greatest percent colonization in this study was a wetland site. A comparison of fungal diversity between the pooled wetland site communities and the pooled upland site communities was conducted and the difference was significant (p = 0.003), but the upland sites exhibited the lower diversity. Communities from all sites were dissimilar. Geographic location exerted a greater effect over community structure than did upland versus wetland status. This study concludes that the forested wetland environments studied here neither limited AM colonization nor reduced the overall fungal community and in fact may present a more favorable environment for fungal diversity.     

Land-use change impact on mycorrhizal symbiosis in female and male plants of wild <Emphasis Type="Italic">Carica papaya</Emphasis> (Caricaceae)

It has been acknowledged that land-use change has negative effects on genetic diversity and sex ratio in dioecious species, but less attention has been paid on the influence that land-use change has on the biotic interactions, especially between dioecious species and arbuscular mycorrhizal (AM) fungi. AM mutualism involves reciprocal transfer of carbohydrates and mineral nutrients between the host plant’s roots and these fungi. Here, we report spatial and temporal variation in AM colonization in dioecious wild Carica papaya plants growing in sites with different land use intensity. We tagged, recorded the basal stem circumference and collected roots of reproductive female and male Carica papaya plants in three wild sites during dry and rainy seasons of western Mexico. We also collected soil samples in each site to conduct soil chemical analyses. The sexes of C. papaya did not show significant differences in the frequency (percentage of root colonized by intraradical fungal structures) and abundance (length of intraradical hyphae) of AM fungi but the higher AM colonization was observed during the dry season, and in the site with the lowest disturbance. There was no relationship between soil chemistry and AM colonization. Overall, our findings suggest that land-use intensity has a negative effect on AM colonization and we discuss the consequences of habitat loss for the reproductive female and male plants, the implications of decreasing AM colonization for wild Carica papaya plants an important species that provides a source of genetic variation for the C. papaya varieties.     

Soil fungal community variation by large-scale reclamation in Sanjiang plain,China

Large-scale marshland reclamation can cause substantial changes to the soil fungal community by disturbances associated with the growth of crop plants and by the addition of fertilizers and pesticides. In this study, high-throughput sequencing of the fungal-specific internal transcribed spacer (ITS) gene region was used to identify fungal taxa. We analyzed the variation in soil fungi diversity and community composition in marshland, paddy, and farmland corn soils, and investigated the relationship between soil fungal community composition and soil physicochemical characteristics to quantify the effect of large-scale reclamation on marshland soil environment in the Sanjiang Plain, northeast China. Marshland soil contained most of the 1997 operational taxonomic units (OTUs) found across all sites (1241), while paddy soil had only 614 OTUs and farmland corn soil 817 OTUs. All reclaimed lands presented a decline in richness and diversity of soil fungi at the OTU level, and soil fungal richness was significantly different between marshland and reclaimed sites (P < 0.05), although it did not differ significantly between marshland and farmland corn sites. Additionally, soil fungal community composition showed different trends and structure after the reclamation. One-way analysis of variance showed Basidiomycota, Zygomycota, Glomeromycota, and Chytridiomycota composition differed significantly between marshland and reclaimed sites (P < 0.05). Nine dominant genera (relative abundance >1.5% in at least one site) and many unclassified genera showed significant variation between marshland and reclaimed sites, including Blumeria, Tomentella, Peziza, Hypholoma, Zopfiella, Mrakia, and Fusarium. Soil fungal community composition and diversity were affected by soil moisture, pH, total carbon (C), available nitrogen (N), soil organic carbon, soil dissolved organic carbon, and C/N (the ratio of total carbon to total nitrogen). The present results contribute to understanding the fungal community in marshland ecosystems, and the role of environmental variability as a predictor of fungal community composition.     

Biodiversity of arbuscular mycorrhizal fungi in the drawdown zone of the Three Gorges Reservoir under different fertilization histories

Impoundment of the Three Gorges Reservoir (TGR) has dramatically influenced the riparian environment and shaped a new drawdown zone, which has experienced long-term winter conditions and short periods of summer flooding. The community structure and diversity of arbuscular mycorrhizal (AM) fungi (AMF) were investigated in three areas with different fertilization histories [Area A (5 years of fertilization), Area B (3 years of fertilization) and Area C (no fertilization)] in the drawdown zone of the TGR. Altogether, 50 AMF species were identified; the genera Acaulospora, Funneliformis and Glomus were predominant. The AM fungal community differed among areas A, B and C. A higher isolation frequency and relative abundance of Acaulospora, Ambispora, Entrophospora and Paraglomus were observed in areas A and B; however, Claroideoglomus, Diversispora, Sclerocystis and Septoglomus were more abundant in Area C. The highest spore density occurred in Area C, and was slightly lower in Area A and lowest in Area B. Conversely, species richness and diversity indices (Shannon–Wiener and evenness indices) were the highest in Area A, followed by areas C and B. Based on nonmetric multidimensional scaling analyses, the distribution of AMF was influenced by plant host, fertilization practice and environmental factors. Among them, the soil physicochemical properties were the main drivers affecting AMF, in which three edaphic attributes (carbon/nitrogen ratio, available phosphorus and potassium content) were significantly correlated (P < 0.001) with the AM fungal community composition in the three areas of the drawdown zone of the TGR.     

AM fungal communities inhabiting the roots of submerged aquatic plant <Emphasis Type="Italic">Lobelia dortmanna</Emphasis> are diverse and include a high proportion of novel taxa

While the arbuscular mycorrhizal (AM) symbiosis is known to be widespread in terrestrial ecosystems, there is growing evidence that aquatic plants also form the symbiosis. It has been suggested that symbiosis with AM fungi may represent an important adaptation for isoëtid plants growing on nutrient-poor sediments in oligotrophic lakes. In this study, we address AM fungal root colonization intensity, richness and community composition (based on small subunit (SSU) ribosomal RNA (rRNA) gene sequencing) in five populations of the isoëtid plant species Lobelia dortmanna inhabiting oligotrophic lakes in Southern Sweden. We found that the roots of L. dortmanna hosted rich AM fungal communities and about 15 % of the detected molecular taxa were previously unrecorded. AM fungal root colonization intensity and taxon richness varied along an environmental gradient, being higher in oligotrophic and lower in mesotrophic lakes. The overall phylogenetic structure of this aquatic fungal community differed from that described in terrestrial systems: The roots of L. dortmanna hosted more Archaeosporaceae and fewer Glomeraceae taxa than would be expected based on global data from terrestrial AM fungal communities.     

Differences in the composition of arbuscular mycorrhizal fungal communities promoted by different propagule forms from a Mediterranean shrubland

As it is well known, arbuscular mycorrhizal (AM) colonization can be initiated from the following three types of fungal propagules: spores, extraradical mycelium (ERM), and mycorrhizal root fragments harboring intraradical fungal structures. It has been shown that biomass allocation of AM fungi (AMF) among these three propagule types varies between fungal taxa, as also differs the ability of the different AMF propagule fractions to initiate new colonizations. In this study, the composition of the AMF community in the roots of rosemary (Rosmarinus officinalis L., a characteristic Mediterranean shrub), inoculated with the three different propagule types, was analyzed. Accordingly, cuttings from this species were inoculated with either AMF spores, ERM, or colonized roots extracted from a natural soil. The AMF diversity within the rosemary roots was characterized using terminal restriction fragment length polymorphism (T-RFLP) of the small subunit (SSU) rDNA region. The AMF community established in the rosemary plants was significantly different according to the type of propagule used as inoculum. AMF taxa differed in their ability to initiate new colonizations from each propagule type. Results suggest different colonization strategies for the different AMF families involved, Glomeraceae and Claroideoglomeraceae colonizing mainly from colonized roots whereas Pacisporaceae and Diversisporaceae from spores and ERM. This supports that AMF taxa show contrasting life-history strategies in terms of their ability to initiate new colonizations from the different propagule types. Further research to fully understand the colonization and dispersal abilities of AMF is essential for their rational use in ecosystem restoration programs.     

Co-existing grass species have distinctive arbuscular mycorrhizal communities

Arbuscular mycorrhizal (AM) fungi are biotrophic symbionts colonizing the majority of land plants, and are of major importance in plant nutrient supply. Their diversity is suggested to be an important determinant of plant community structure, but the influence of host-plant and environmental factors on AM fungal community in plant roots is poorly documented. Using the terminal restriction fragment length polymorphism (T-RFLP) strategy, the diversity of AM fungi was assessed in 89 roots of three grass species (Agrostis capillaris, Festuca rubra, Poa pratensis) that co-occurred in the same plots of a field experiment. The impact of different soil amendments (nitrogen, lime, nitrogen and lime) and insecticide application on AM fungal community was also studied. The level of diversity found in AM fungal communities using the T-RFLP strategy was consistent with previous studies based on clone libraries. Our results clearly confirm that an AM fungal host-plant preference exists, even between different grass species. AM communities colonizing A. capillaris were statistically different from the others (P < 0.05). Although grass species evenness changed in amended soils, AM fungal community composition in roots of a given grass species remained stable. Conversely, in plots where insecticide was applied, we found higher AM fungal diversity and, in F. rubra roots, a statistically different AM fungal community.     

Incidence and diversity of arbuscular mycorrhizal fungi and successor herbaceous plants in an agro-system irrigated with produced water

The purpose of this study was to evaluate the diversity of herbaceous plants and arbuscular mycorrhizal fungi following the cultivation of sunflower (Helianthus annuus L., cv. BRS 321) irrigated with produced water. The sunflower plants were irrigated during three successive cycles with different types of water: produced water obtained through simple filtration (PWSF), and secondly, produced water treated by reverse osmosis (PWRO), and the control with groundwater from the aquifer Açu (WCA). In June 2014, five months after the final harvest, the treatments were evaluated in terms of the diversity of successor plants and their roots colonized by arbuscular mycorrhiza (AM); and samples of soil, in which the following were measured: the spore abundance of AM fungi, the levels of glomalin in easily extracted glomalin and total glomalin. Of a total of eighteen species of herbaceous plants which were identified in the experimental field, Dactyloctenium aegyptium was related with the use of PWSF, Panicum sp. and Diodella apiculata with the use of PWRO, and Trianthema portulacastrum and Eragrostis tenella with the control WCA. The diversity of AM fungi was affected by irrigation with PWSF, in which two species of Acaulospora, one species of Gigaspora and species of Paraglomus were absent, compared to the treatment with PWRO. Acaulospora sp.1 was related with the WCA control as an indicator species. The use of produced water which had undergone reverse osmosis had a short-term effect on the content of glomalin which is easily extractable from the soil but did not change the mycorrhization rates of plants. These results enable us to infer that irrigation with produced water leads to a reduction in the diversity of herbaceous plants and of arbuscular mycorrhizal fungi in the soil, confirming the importance of monitoring agro-systems irrigated with residual water.     

Illumina-based analysis of the rhizosphere microbial communities associated with healthy and wilted Lanzhou lily (<Emphasis Type="Italic">Lilium davidii</Emphasis> var. <Emphasis Type="Italic">unicolor</Emphasis>) plants grown in the field

Lanzhou lily (Liliumdavidii var. unicolor) is the best edible lily as well as a traditional medicinal plant in China. The microbes associated with plant roots play crucial roles in plant growth and health. However, little is known about the differences of rhizosphere microbes between healthy and wilted Lanzhou lily (Lilium davidii var. unicolor) plants. The objective of this study was to compare the rhizosphere microbial community and functional diversity of healthy and wilted plants, and to identify potential biocontrol agents with significant effect. Paired end Illumina Mi-Seq sequencing of 16S rRNA and ITS gene amplicons was employed to study the bacterial and fungal communities in the rhizosphere soil of Lanzhou lily plants. BIOLOG technology was adopted to investigate the microbial functional diversity. Our results indicated that there were major differences in the rhizosphere microbial composition and functional diversity of wilted samples compared with healthy samples. Healthy Lanzhou lily plants exhibited lower rhizosphere-associated bacterial diversity than diseased plants, whereas fungi exhibited the opposite trend. The dominant phyla in both the healthy and wilted samples were Proteobacteria and Ascomycota, i.e., 34.45 and 64.01 %, respectively. The microbial functional diversity was suppressed in wilted soil samples. Besides Fusarium, the higher relative abundances of Rhizoctonia, Verticillium, Penicillium, and Ilyonectria (Neonectria) in the wilted samples suggest they may pathogenetic root rot fungi. The high relative abundances of Bacillus in Firmicutes in healthy samples may have significant roles as biological control agents against soilborne pathogens. This is the first study to find evidence of major differences between the microbial communities in the rhizospheric soil of healthy and wilted Lanzhou lily, which may be linked to the health status of plants.     

Effects of using different host plants and long-term fertilization systems on population sizes of infective arbuscular mycorrhizal fungi

The effect of cultivation of mycorrhizal and non-mycorrhizal plants and mineral fertilization on the arbuscular mycorrhizal fungal (AMF) community structure of maize (Zea mays L.) plants was studied. Soil samples were collected from two field experiments treated for 5 years with three fertilization systems (Control – no fertilization; Mineral – NPK fertilization; and Organic – Farmyard manure fertilization). Soil samples containing soil and root fragments of rapeseed (Brassica napus L., non-mycorrhizal plant) and wheat (Triticum aestivum L., mycorrhizal plant) collected from the field plots were used as native microbial inoculum sources to maize plants. Maize plants were sown in pots containing these inoculum sources for four months under glasshouse conditions. Colonization of wheat roots by AMF, AMF community structure, AMF diversity (Shannon’s index), AMF dominance (Simpson’s index) and growth of maize were investigated. Sixteen AMF species were identified from rhizosphere soil samples as different species of genera Acaulospora, Claroideoglomus, Dentiscutata, Funneliformis, Gigaspora, Quatunica, Racocetra, and Rhizoglomus. Maize plants grown in manure-fertilized soils had a distinct AMF community structure from plants either fertilized with mineral NPK-fertilizer or non-fertilized. The results also showed that inoculum from non-mycorrhizal plants combined with mineral fertilization decreased AMF diversity (Shannon’s index), AMF dominance (Simpson’s index) and growth of maize. Our findings suggest that non-mycorrhizal plants, such as B. napus, can negatively affect the presence and the effects of soil inoculation on maize growth. Also, our results highlight the importance of considering the long-term effect of rapeseed cultivation system on the reduction of population sizes of infective AMF, and its effect on succeeding annual crops.     

Arbuscular mycorrhizal fungi can shift plant-soil feedback of grass-endophyte symbiosis from negative to positive

Background and aims

Variations in root-associated fungal communities contribute to the so-called ‘crop rotation benefit’ on soil productivity. We assessed the effects of chickpea, lentil, and pea in wheat-based rotations, as compared to wheat monoculture, on the structure of root-associated fungal communities, and described the legacy of pulses on a following wheat crop.

Methods

The internal transcribed spacer (ITS) and 18S rRNA gene markers, and 454 amplicon pyrosequencing were used to describe the fungal communities of crop roots and rhizosphere soil in a field experiment and agronomic data were collected.

Results

Pulses influenced only the structure of the non-mycorrhizal fungal community of roots. Fusarium tricinctum, Clonostachys rosea, Fusarium redolens, and Cryptococcus sp. were specific to certain crops. Despite the absence of selective effects of pulses on their associated arbuscular mycorrhizal (AM) fungal community, pea had a legacy effect on the structure of the AM fungal community associated with the roots of the following wheat crop, in one of the two year/sites examined. Species of Mortierella, Cryptococcus, and Paraglomus in wheat rhizosphere soil may benefit yield, whereas species of Fusarium, Davidiella, Lachnum, Sistotrema and Podospora may reduce yield.

Conclusion

The effect of pulse crops on root fungal communities varied with rotation crop species. Pulses had various effects on the physiology of the following wheat crop, including increased productivity.

     

Community Structure of Rhizomicrobiomes in Four Medicinal Herbs and Its Implication on Growth Management

Medicinal plants are the basic materials of traditional Chinese medicine. Soil characteristics and microbial contribution play important roles in the growth and product quality of medicinal plants, but the link between them in the rhizosphere of medicinal plants has been overlooked. Accordingly, Mentha haplocalyx, Perilla frutescens, Glycyrrhiza uralensis, and Astragalus membranaceus, four plants used in traditional Chinese medicines, were investigated in this study in order to elucidate bacterial and arbuscular mycorrhizal fungal (AMF) diversity in the rhizosphere and its possible association with soil quality. DGGE-based 16S rRNA and 18S rRNA gene sequencing results indicated that the diversity of both bacteria and AMF in Glycyrrhiza uralensis and Astragalus membranaceus was significantly higher than those in Mentha haplocalyx and Perilla frutescens, suggesting that medicinal plants have different preferences even under the same conditions. In addition, enzymatic activities and nutrition were enhanced in the rhizospheric soil of Mentha haplocalyx and Perilla frutescens, and the correlation among AMF diversity, soil enzymatic activities and nutrition was confirmed using RDA analysis. These results suggest the potential to grow medicinal plants with a reasonable rotation or intercrop in order to maintain long-term continuous soil development.     

Increased Diversity of Arbuscular Mycorrhizal Fungi in a Long-Term Field Experiment via Application of Organic Amendments to a Semiarid Degraded Soil

In this study, we tested whether communities of arbuscular mycorrhizal (AM) fungi associated with roots of plant species forming vegetative cover as well as some soil parameters (amounts of phosphatase and glomalin-related soil protein, microbial biomass C and N concentrations, amount of P available, and aggregate stability) were affected by different amounts (control, 6.5 kg m⁻², 13.0 kg m⁻², 19.5 kg m⁻², and 26.0 kg m⁻²) of an urban refuse (UR) 19 years after its application to a highly eroded, semiarid soil. The AM fungal small-subunit (SSU) rRNA genes were subjected to PCR, cloning, single-stranded conformation polymorphism analysis, sequencing, and phylogenetic analyses. One hundred sixteen SSU rRNA sequences were analyzed, and nine AM fungal types belonging to Glomus groups A and B were identified: three of them were present in all the plots that had received UR, and six appeared to be specific to certain amendment doses. The community of AM fungi was more diverse after the application of the different amounts of UR. The values of all the soil parameters analyzed increased proportionally with the dose of amendment applied. In conclusion, the application of organic wastes enhanced soil microbial activities and aggregation, and the AM fungal diversity increased, particularly when a moderate dose of UR (13.0 kg m⁻²) was applied.The semiarid Mediterranean areas of Southeastern Spain are affected by environmental degradation and erosive processes due to the fact that they are characterized by a set of climatic conditions that includes irregular and scarce rainfall and long, dry, and hot summers. Under these conditions, the soil organic matter content decreases, and the availability of nutrients and water for plants is reduced. Consequently, soil productivity decreases, levels of below-ground microbially diverse populations decline, and the water deficit limits plant growth so that the vegetation cover of natural soils cannot be sustained. Therefore, the development of revegetation techniques to reduce erosion, to remediate the effects of degradation, and, thus, to allow the restoration of biodiversity is needed. It was previously demonstrated that the application of organic amendments, such as urban refuse (UR), to soil increases the organic matter content of soil and improves the quality and productivity of degraded soils (17, 44, 57). Also, it was previously shown that the organic residues yield an improvement in levels of microbially diverse populations in the soil (43).A substantial part of the soil microbial communities belongs to the arbuscular mycorrhizal (AM) fungi, an ancient group of fungi belonging to the phylum Glomeromycota (49), which form mutualistic associations with the roots of the majority of land plants. These fungi have a variety of beneficial effects on their host plants, such as increasing the uptake of mineral nutrients, particularly phosphorus and nitrogen (41, 52); reduction of pathogen infections (7); improvement of water relations (12) and soil stability (58); and the limitation of heavy metal uptake (34). It is evident that AM fungi are an important factor contributing to the maintenance of terrestrial ecosystem functioning. Studies have shown that the diversity of AM fungal populations in the soil can affect plant diversity and productivity and ecosystem stability (62). Therefore, information on the species composition of the AM fungal community in roots is important for an understanding of mycorrhizal function as well as for the effective management and preservation of the diversity of AM fungal populations in ecological field studies.Thanks to advances in molecular techniques in recent years, it is possible to apply PCR-based molecular methods in order to analyze the diversity of AM fungi colonizing the roots of an individual plant at any given time. Traditional identification based on spore morphology is often problematic, and the abundance of spores in the soil may not accurately reflect AM fungal community composition and dynamics (8). The single-stranded conformation polymorphism (SSCP) approach is a very sensitive and reproducible technique for analyzing the sequence diversity of AM fungi within roots (30). This method is based on nucleotide differences between homologous sequence strands, which are detected by electrophoresis of single-stranded DNA under nondenaturing conditions (38).It is known that the application of organic amendments can have a positive effect on the proliferation of natural AM fungi in crop systems (20, 26). The stimulatory effects of the addition of organic matter on the development of AM fungi could be related to an improvement in the extensive network of AM fungal mycelium in the soil. In this way, the colonized plants are able to effectively exploit nutrients and water from soil (52). Moreover, AM fungi are able to exploit nutrients released by the mineralization of organic matter due to the activities of mineralizing microorganisms (28). However, there are many previous reports that showed a strong negative impact on the presence of AM fungal populations and mycorrhizal colonization when composted urban waste was added to the soil (19, 46). Also, research using trap cultures of host plants showed a decrease in the level of diversity of AM fungal species in soils amended with sewage sludge (25, 61).In a previous study carried out in 1992 at the site that is also the subject of the current work, Roldán and Albaladejo (43) found that the application of UR decreased levels of AM fungal populations in the first year after amendment; however, they observed an increase in levels of these populations 3 years after the addition. We hypothesized that after a long period of time, the application of UR could alter the diversity of AM fungal populations in a highly eroded, semiarid soil and that this effect could be influenced by the refuse application rate. In order to verify this hypothesis, we studied the diversity of the AM fungi associated with the roots of plant species forming the vegetative cover of five plots that received different amounts of UR 19 years after the amendment. Also, we determined whether there was an improvement in soil quality parameters related to soil microbial activity.     

Nutrient limitation drives response of <Emphasis Type="Italic">Calamagrostis epigejos</Emphasis> to arbuscular mycorrhiza in primary succession

Little is known about the functioning of arbuscular mycorrhizal (AM) symbiosis over the course of primary succession, where soil, host plants, and AM fungal communities all undergo significant changes. Over the course of succession at the studied post-mining site, plant cover changes from an herbaceous community to the closed canopy of a deciduous forest. Calamagrostis epigejos (Poaceae) is a common denominator at all stages, and it dominates among AM host species. Its growth response to AM fungi was studied at four distinctive stages of natural succession: 12, 20, 30, and 50 years of age, each represented by three spatially separated sites. Soils obtained from all 12 studied sites were γ-sterilized and used in a greenhouse experiment in which C. epigejos plants were (1) inoculated with a respective community of native AM fungi, (2) inoculated with reference AM fungal isolates from laboratory collection, or (3) cultivated without AM fungi. AM fungi strongly boosted plant growth during the first two stages but not during the latter two, where the effect was neutral or even negative. While plant phosphorus (P) uptake was generally increased by AM fungi, no contribution of mycorrhizae to nitrogen (N) uptake was recorded. Based on N:P in plant biomass, we related the turn from a positive to a neutral/negative effect of AM fungi on plant growth, observed along the chronosequence, to a shift in relative P and N availability. No functional differences were found between native and reference inocula, yet root colonization by the native AM fungi decreased relative to the reference inoculum in the later succession stages, thereby indicating shifts in the composition of AM fungal communities reflected in different functional characteristics of their members.     

Soil drying procedure affects the DNA quantification of <Emphasis Type="Italic">Lactarius vinosus</Emphasis> but does not change the fungal community composition

Drying soil samples before DNA extraction is commonly used for specific fungal DNA quantification and metabarcoding studies, but the impact of different drying procedures on both the specific fungal DNA quantity and the fungal community composition has not been analyzed. We tested three different drying procedures (freeze-drying, oven-drying, and room temperature) on 12 different soil samples to determine (a) the soil mycelium biomass of the ectomycorrhizal species Lactarius vinosus using qPCR with a specifically designed TaqMan® probe and (b) the fungal community composition and diversity using the PacBio® RS II sequencing platform. Mycelium biomass of L. vinosus was significantly greater in the freeze-dried soil samples than in samples dried at oven and room temperature. However, drying procedures had no effect on fungal community composition or on fungal diversity. In addition, there were no significant differences in the proportions of fungi according to their functional roles (moulds vs. mycorrhizal species) in response to drying procedures. Only six out of 1139 operational taxonomic units (OTUs) had increased their relative proportions after soil drying at room temperature, with five of these OTUs classified as mould or yeast species. However, the magnitude of these changes was small, with an overall increase in relative abundance of these OTUs of approximately 2 %. These results suggest that DNA degradation may occur especially after drying soil samples at room temperature, but affecting equally nearly all fungi and therefore causing no significant differences in diversity and community composition. Despite the minimal effects caused by the drying procedures at the fungal community composition, freeze-drying resulted in higher concentrations of L. vinosus DNA and prevented potential colonization from opportunistic species.     

Contribution of different arbuscular mycorrhizal fungal inoculum to <Emphasis Type="Italic">Elymus nutans</Emphasis> under nitrogen addition

Arbuscular mycorrhizal (AM) fungi are known to promote plant growth and nutrient uptake, but their role in nitrogen (N) uptake still remains unclear. Therefore, a pot experiment was set up to evaluate the impacts of N addition and AM inoculation (Diversispora eburnea, Claroideoglomus etunicatum, Paraglomus occultum, and their mixture) on AM root colonization, plant biomass, N and P nutrition in Elymus nutans. Our results showed that AM root colonization was unaffected by N addition but was significantly affected by different AM fungal species. D. eburnea and C. etunicatum showed significant higher root colonization than P. occultum. The E. nutans exhibited the highest biomass when inoculated with D. eburnea and significantly higher than non-mycorrhizal (the control) regardless of N addition. Under N addition treatment, D. eburnea significantly enhanced P content of roots, N content of shoots and roots, while AM mixture significantly enhanced shoot P content compared with non-mycorrhizal. However, N and P content in shoots and roots did not significantly vary among treatments when no N was added. In addition, inoculation with C. etunicatum and P. occultum showed no significant effect on plant biomass, N and P content regardless of N addition. In conclusion, this study revealed that the plant response to N addition depends on AM fungal species and also confirmed that significant functional diversity exists among AM fungal species.     

<Emphasis Type="Italic">Pyrola japonica</Emphasis>, a partially mycoheterotrophic Ericaceae,has mycorrhizal preference for russulacean fungi in central Japan

Mycorrhizal symbiosis often displays low specificity, except for mycoheterotrophic plants that obtain carbon from their mycorrhizal fungi and often have higher specificity to certain fungal taxa. Partially mycoheterotrophic (or mixotrophic, MX) plant species tend to have a larger diversity of fungal partners, e.g., in the genus Pyrola (Monotropoideae, Ericaceae). Preliminary evidence however showed that the Japanese Pyrola japonica has preference for russulacean fungi based on direct sequencing of the fungal internal transcribed spacer (ITS) region from a single site. The present study challenges this conclusion using (1) sampling of P. japonica in different Japanese regions and forest types and (2) fungal identification by ITS cloning. Plants were sampled from eight sites in three regions, in one of which the fungal community on tree ectomycorrhizal (ECM) tips surrounding P. japonica was also analyzed. In all, 1512 clone sequences were obtained successfully from 35 P. japonica plants and 137 sequences from ECM communities. These sequences were collectively divided into 74 molecular operational taxonomic units (MOTUs) (51 and 33 MOTUs, respectively). MOTUs from P. japonica involved 36 ECM taxa (96 % of all clones), and 17 of these were Russula spp. (76.2 % of all clones), which colonized 33 of the 35 sampled plants. The MOTU composition significantly differed between P. japonica and ECM tips, although shared species represented 26.3 % of the ECM tips community in abundance. This suggests that P. japonica has a preference for russulacean fungi.     

Insights into fungal communities colonizing the acarosphere in a forest soil habitat

Knowledge on the diversity and ecology of microfungi associated with soil-dwelling mites is rather limited. To get insights into associations between the two highly diverse groups, we studied composition and potential function of mite-associated fungal communities occurring in soil. Two different mite species living in temperate region pine forest soil were screened for associated fungi. The fungal community was assessed by restriction fragment length polymorphism (RFLP) analyses in a predatory (Leptogamasus obesus) and a predominantly saprobic (Oppiella subpectinata) mite species as well as in the organic soil layer. Key fungi were identified by sequencing, and community composition was exemplarily compared between the RFLP and a 454 metabarcoding approach. Composition of the fungal communities differed between mite species and between mites and organic soil layer. The mites were predominantly associated with Zygomycota, less frequently with Ascomycota, and rarely with Basidiomycota. The bulk soil was colonized by roughly equal proportions of the three phyla. Fungal taxa being known to exhibit chitinolytic activity were predominantly restricted to mites. Compositional and functional differences between the communities suggest that mites represent a particular microhabitat for fungi, the “acarosphere.” This mobile habitat may contribute to nutrient cycling by combining fungal and animal decomposition activities and serve as vector for soil-inhabiting fungi.     

The rhizosphere microbial community response to a bio-organic fertilizer: finding the mechanisms behind the suppression of watermelon <Emphasis Type="Italic">Fusarium</Emphasis> wilt disease

We aimed to evaluate the capability of bio-organic fertilizer suppressing watermelon Fusarium wilt disease, compare the variations of the rhizosphere bacterial and fungal community compositions after treatment with different fertilizers, and explore mechanisms causing disease suppression in rhizosphere microbial community. A rhizobacterium (Bacillus amyloliquefaciens JDF35) was identified to control watermelon Fusarium wilt disease. Bio-organic fertilizer JDF35 (BOF) was generated by inoculating JDF35 into the organic fertilizer (OF) composed of cow and chicken manure compost (1:50 v/w). A three successive growing season pot experiment was designed to evaluate the effects of BOF compared with OF and chemical fertilizer (CF). Next-generation sequencing using the Illumina MiSeq platform was used to investigate the variations in rhizosphere microbial community composition. The growth of the watermelon plants, soil pH, and available N, P and K concentrations were the highest in the BOF treatment. The Fusarium wilt incidence in the BOF treatment was lower than that in the CF and OF treatment, and the differences for disease incidence were significant (P < 0.001). The diversity of the rhizosphere bacterial community was higher, and that of the fungal was lower in the BOF treatment. Most importantly, the BOF treatment had lowest abundances of Fusarium. The application of the BOF altered the composition of rhizosphere microbial community, suppressing Fusarium wilt disease and promoting plant growth.