Effects of Funneliformis mosseae on the utilization of organic phosphorus in Camellia oleifera Abel.

Arbuscular mycorrhizal (AM) fungi play an important role in the acquisition of phosphorus (P) by plants. The external hyphae of AM fungi function as an extension of plant roots and may downregulate related functions in the roots. It is not clear whether the ability of AM fungi to mineralize organic P affects root phosphatase activities. A pot experiment was conducted to investigate the effect of Funneliformis mosseae on soil organic P mineralization under phytate application and to explore root phosphatase activities, P uptake, and growth in Camellia oleifera Abel. The plants and their growth substrates were harvested 4 and 8 months after planting. The results showed that organic P application had no effect on the total dry mass of nonmycorrhizal plants, but differences in dry mass under P application were observed in mycorrhizal plants in both harvests. Inoculation with F. mosseae increased soil acid phosphatase, phytase, and alkaline phosphatase activities and reduced the soil organic P content. Mycorrhizal plants had higher root activity, shoot and root P contents and root acid phosphatase and phytase activities than nonmycorrhizal plants irrespective of organic P application. In conclusion, AM fungi enhanced the mineralization of soil organic P and positively affect root phosphatase activities.

Palmitvaccenic Acid (Δ11-cis-hexadecenoic acid) Is Synthesized by an OLE1-like Desaturase in the Arbuscular Mycorrhiza Fungus Rhizophagus irregularis.

Arbuscular mycorrhiza (AM) fungi deliver mineral nutrients to the plant host in exchange for reduced carbon in the form of sugars and lipids. Colonization with AM fungi upregulates a specific host lipid synthesis pathway resulting in the production of fatty acids. Predominantly palmitic acid (16:0) and the unusual palmitvaccenic acid (16:1Δ11cis) accumulate in the fungus Rhizophagus irregularis. Here, we present the isolation and characterization of RiOLE1-LIKE, the desaturase involved in palmitvaccenic acid synthesis, by heterologous expression in yeast and plants. Results are in line with the scenario in which RiOLE1-LIKE encodes an acyl-CoA desaturase with substrate specificity for C15-C18 acyl groups, in particular C16. Phylogenetic analysis of RiOLE1-LIKE-related sequences revealed that this gene is conserved in AM fungi from the Glomales and Diversisporales but is absent from nonsymbiotic Mortierellaceae and Mucoromycotina fungi, suggesting that 16:1Δ11cis provides a specific function during AM colonization.

DES2 is a fatty acid Δ11 desaturase capable of synthesizing palmitvaccenic acid in the arbuscular mycorrhizal fungus Rhizophagus irregularis.

Arbuscular mycorrhizal (AM) fungi are oleaginous organisms, and the most abundant fatty acyl moiety usually found in their lipids is palmitvaccenic acid (16:1Δ11cis ). However, it is not known how this uncommon fatty acid species is made. Here, we have cloned two homologues of lepidopteran fatty acyl-coenzyme A Δ11 desaturases from the AM fungus Rhizophagus irregularis. Both enzymes, DES1 and DES2, are expressed in intraradical mycelium and can complement the unsaturated fatty acid-requiring auxotrophic growth phenotype of the Saccharomyces cerevisiae ole1Δ mutant. DES1 expression leads almost exclusively to oleic acid (18:1Δ9cis ) production, whereas DES2 expression results in the production of 16:1Δ11cis and vaccenic acid (18:1Δ11cis ). DES2 therefore encodes a Δ11 desaturase that is likely to be responsible for the synthesis of 16:1Δ11cis in R. irregularis.

A mycorrhiza-specific H+ -ATPase is essential for arbuscule development and symbiotic phosphate and nitrogen uptake.

Most land plants can form symbiosis with arbuscular mycorrhizal (AM) fungi to enhance uptake of mineral nutrients, particularly phosphate (Pi) and nitrogen (N), from the soil. It is established that transport of Pi from interfacial apoplast into plant cells depends on the H+ gradient generated by the H+ -ATPase located on the periarbuscular membrane (PAM); however, little evidence regarding the potential link between mycorrhizal N transport and H+ -ATPase activity is available to date. Here, we report that a PAM-localized tomato H+ -ATPase, SlHA8, is indispensable for arbuscule development and mycorrhizal P and N uptake. Knockout of SlHA8 resulted in truncated arbuscule morphology, reduced shoot P and N accumulation, and decreased H+ -ATPase activity and acidification of apoplastic spaces in arbusculated cells. Overexpression of SlHA8 in tomato promoted both P and N uptake, and increased total colonization level, but did not affect arbuscule morphology. Heterogeneous expression of SlHA8 in the rice osha1 mutant could fully complement its defects in arbuscule development and mycorrhizal P and N uptake. Our results propose a pivotal role of the SlHA8 in energizing both the symbiotic P and N transport, and highlight the evolutionary conservation of the AM-specific H+ -ATPase orthologs in maintaining AM symbiosis across different mycorrhizal plant species.

Purification and characterization of Terfezia claveryi TcCAT-1, a desert truffle catalase upregulated in mycorrhizal symbiosis.

Terfezia claveryi Chatin is a mycorrhizal fungus that forms ectendomycorrhizal associations with plants of Helianthemum genus. Its appreciated edibility and drought resistance make this fungus a potential alternative crop in arid and semiarid areas of the Mediterranean region. In order to increase the knowledge about the biology of this fungus in terms of mycorrhiza formation and response to drought stress, a catalase from T. claveryi (TcCAT-1) has been purified to apparent homogeneity and biochemically characterized; in addition, the expression pattern of this enzyme during different stages of T. claveryi biological cycle and under drought stress conditions are reported. The results obtained, together with the phylogenetic analysis and homology modeling, indicate that TcCAT-1 is a homotetramer large subunit size monofunctional-heme catalase belonging to Clade 2. The highest expression of this enzyme occurs in mature mycorrhiza, revealing a possible role in mycorrhiza colonization, but it is not upregulated under drought stress. However, the H2O2 content of mycorrhizal plants submitted to drought stress is lower than in well watered treatments, suggesting that mycorrhization improves the plant's oxidative stress response, although not via TcCAT-1 upregulation.

The ability of a host plant to associate with different symbiotic partners affects ectomycorrhizal functioning.

Some plants that associate with ectomycorrhizal (ECM) fungi are also able to simultaneously establish symbiosis with other types of partners. The presence of alternative partners that may provide similar benefits may affect ECM functioning. Here we compared potential leucine-aminopeptidase (LA) and acid phosphatase (AP) enzyme activity (involved in N and P cycling, respectively) in ECM fungi of three hosts planted under the same conditions but differing in the type of partners: Pinus (ECM fungi only), Eucalyptus (ECM and arbuscular mycorrhizal -AM- fungi) and Acacia (ECM, AM fungi and rhizobial bacteria). We found that the ECM community on Acacia and Eucalyptus had higher potential AP activity than the Pinus community. The ECM community in Acacia also showed increased potential LA activity compared to Pinus. Morphotypes present in more than one host showed higher potential AP and LA activity when colonizing Acacia than when colonizing another host. Our results suggest that competition with AM fungi and rhizobial bacteria could promote increased ECM activity in Eucalyptus and Acacia. Alternatively, other host-related differences such as ECM community composition could also play a role. We found evidence for ECM physiological plasticity when colonizing different hosts, which might be key for adaptation to future climate scenarios.

Cadmium and arsenic responses in the ectomycorrhizal fungus Laccaria bicolor: glutathione metabolism and its role in metal(loid) homeostasis.

Ectomycorrhizal fungi play an important role in protecting their host plant from metal(loid) stresses by synthesizing various thiol rich compounds like metallothioneins and glutathione. We investigated the effect of cadmium (Cd) and arsenic (As) stress with a specific interest on glutathione (GSH) in the ectomycorrhizal fungus Laccaria bicolor. The total GSH levels inside the cell were significantly increased with increase in external metal(loid) stress. An analysis of the transcript levels of genes responsible for GSH synthesis, γ-glutamylcysteine synthetase (Lbγ-GCS) and glutathione synthetase (LbGS), using qPCR revealed that expression of both genes increased as a function of external metal(loid) concentration. The enzyme activity of both Lbγ-GCS and LbGS were increased with increase in external Cd and As concentration. Further, the functional role of Lbγ-GCS and LbGS genes in response to Cd and As stress was studied using their respective yeast mutant strains gsh1 Δ and gsh2 Δ . The mutant strains successfully expressed the two genes resulting in wild-type phenotype restoration of Cd and As tolerance. From these results, it was concluded that GSH act as a core component in the mycorrhizal defence system under Cd and As stress for metal(loid) homeostasis and detoxification.

Exposure to ionizing radiation affects the growth of ectomycorrhizal fungi and induces increased melanin production and increased capacities of reactive oxygen species scavenging enzymes.

Ectomycorrhizal (EM) fungi form symbioses with dominant tree families in boreal, temperate and tropical ecosystems and are important drivers of ecosystem function. EM fungal hyphae extend over a large area making them susceptible to enhanced radiation levels from naturally occurring or anthropogenically originating radioisotopes in the rhizosphere. In this study, the in-vitro effects of ionizing radiation on the growth and biomass of EM fungi Suillus luteus, S. bovinus and Rhizopogon luteolus were investigated. EM fungal cultures were exposed to gamma radiation from a 137Cs source for 137 h in darkness at 21 °C at dose rates of 404, 108.5 and 54.9 mGy h-1 resulting in total absorbed doses of 55.21, 14.82 and 7.50 Gy respectively. Cultures grown in the dark at 21 °C but not exposed to the 137Cs source served as the control. Our results show that EM fungi vary in their sensitivity to ionizing radiation. EM fungi used in this study produced melanin and reactive oxygen species scavenging enzymes such as catalase and superoxide dismutase as a response to ionizing radiation.

A methyltransferase gene from arbuscular mycorrhizal fungi involved in arsenic methylation and volatilization.

Arbuscular mycorrhizal fungi (AMF), ubiquitous symbiotic fungi associated with the majority of terrestrial plants, were demonstrated to play important roles in arsenic (As) translocation and transformation in the plant-soil continuum, and substantially influence plant As tolerance. However, the direct involvement of AMF in As methylation and volatilization and their molecular mechanisms remain unsolved. Here, an arsenite methyltransferase gene RiMT-11 was identified and characterized from AM fungus Rhizophagus irregularis. Heterologous expression of RiMT-11 enhanced arsenite resistance of E. coli (Δars) through methylating As into monomethylarsonic acid (MMA), dimethylarsinic acid (DMA) and ultimately volatile trimethyl arsine (TMAs). In a two-compartment in vitro monoxenic cultivation system, methylated and volatile As were also detected from AM symbioses with arsenate addition, accompanied by strong up-regulation of RiMT-11 expression in extraradical hyphae. The present study provided direct evidence and illustrated an underlying mechanism of As methylation and volatilization by AMF, leading to a deeper insight into the role of AMF in As biogeochemical cycling.

N-Acetylglucosaminidase activity, a functional trait of chitin degradation, is regulated differentially within two orders of ectomycorrhizal fungi: Boletales and Agaricales.

Chitin is one of the most abundant nitrogen-containing polymers in forest soil. Ability of ectomycorrhizal (EM) fungi to utilize chitin may play a key role in the EM symbiosis nutrition and soil carbon cycle. In forest, EM fungi exhibit high diversity, which could be based on function partitioning and trait complementarity. Although it has long been recognized that closely related species share functional characteristics, the phylogenetic conservatism of functional traits within microorganisms remains unclear. Because extracellular N-acetylglucosaminidase activity has been proposed as functional trait of chitin degradation, we screened this activity on 35 EM fungi species with or without chitin in the growth medium to (i) describe the functional diversity of EM fungi and (ii) identify potential links between this functional trait and EM fungal phylogeny. We observed large variations of the extracellular N-acetylglucosaminidase activities among the fungal strains. Furthermore, our results revealed two regulation patterns of extracellular N-acetylglucosaminidase activities. Indeed, these chitinolytic activities were stimulated or repressed in the presence of chitin, in comparison to the control treatment. These profiles of extracellular N-acetylglucosaminidase stimulation/repression might be conserved at a high phylogenetic level in the Basidiomycota phylum, as illustrated by the opposite patterns of regulation between Boletales and Agaricales. Finally, the downregulation of this activity by chitin, for some EM fungal groups, might suggest another chitin degradation pathway.

The ectomycorrhizal basidiomycete Laccaria bicolor releases a secreted ß-1,4 endoglucanase that plays a key role in symbiosis development.

In ectomycorrhiza, root ingress and colonization of the apoplast by colonizing hyphae is thought to rely mainly on the mechanical force that results from hyphal tip growth, but this could be enhanced by secretion of cell-wall-degrading enzymes, which have not yet been identified. The sole cellulose-binding module (CBM1) encoded in the genome of the ectomycorrhizal Laccaria bicolor is linked to a glycoside hydrolase family 5 (GH5) endoglucanase, LbGH5-CBM1. Here, we characterize LbGH5-CBM1 gene expression and the biochemical properties of its protein product. We also immunolocalized LbGH5-CBM1 by immunofluorescence confocal microscopy in poplar ectomycorrhiza. We show that LbGH5-CBM1 expression is substantially induced in ectomycorrhiza, and RNAi mutants with a decreased LbGH5-CBM1 expression have a lower ability to form ectomycorrhiza, suggesting a key role in symbiosis. Recombinant LbGH5-CBM1 displays its highest activity towards cellulose and galactomannans, but no activity toward L. bicolor cell walls. In situ localization of LbGH5-CBM1 in ectomycorrhiza reveals that the endoglucanase accumulates at the periphery of hyphae forming the Hartig net and the mantle. Our data suggest that the symbiosis-induced endoglucanase LbGH5-CBM1 is an enzymatic effector involved in cell wall remodeling during formation of the Hartig net and is an important determinant for successful symbiotic colonization.

A family of archaea-like carboxylesterases preferentially expressed in the symbiotic phase of the mychorrizal fungus Tuber melanosporum.

An increasing number of esterases is being revealed by (meta) genomic sequencing projects, but few of them are functionally/structurally characterized, especially enzymes of fungal origin. Starting from a three-member gene family of secreted putative "lipases/esterases" preferentially expressed in the symbiotic phase of the mycorrhizal fungus Tuber melanosporum ("black truffle"), we show here that these enzymes (TmelEST1-3) are dimeric, heat-resistant carboxylesterases capable of hydrolyzing various short/medium chain p-nitrophenyl esters. TmelEST2 was the most active (kcat = 2302 s-1 for p-nitrophenyl-butyrate) and thermally stable (T50 = 68.3 °C), while TmelEST3 was the only one displaying some activity on tertiary alcohol esters. X-ray diffraction analysis of TmelEST2 revealed a classical α/ß hydrolase-fold structure, with a network of dimer-stabilizing intermolecular interactions typical of archaea esterases. The predicted structures of TmelEST1 and 3 are overall quite similar to that of TmelEST2 but with some important differences. Most notably, the much smaller volume of the substrate-binding pocket and the more acidic electrostatic surface profile of TmelEST1. This was also the only TmelEST capable of hydrolyzing feruloyl-esters, suggestinng a possible role in root cell-wall deconstruction during symbiosis establishment. In addition to their potential biotechnological interest, TmelESTs raise important questions regarding the evolutionary recruitment of archaea-like enzymes into mesophilic subterranean fungi such as truffles.

Early-successional ectomycorrhizal fungi effectively support extracellular enzyme activities and seedling nitrogen accumulation in mature forests.

After stand-replacing disturbance, regenerating conifer seedlings become colonized by different ectomycorrhizal fungi (EMF) than the locally adapted EMF communities present on seedlings in mature forests. We studied whether EMF species that colonized subalpine fir (Abies lasiocarpa) seedlings in clearcuts differed from those that colonized seedlings in adjacent mature forests with respect to mycorrhizoplane extracellular enzyme activities (EEAs) and N status of the seedlings. We tested two alternate hypotheses: (1) that EEAs would differ between the two EMF communities, with higher activities associated with forest-origin communities, and (2) that acclimation to soil environment was considerable enough that EEAs would be determined primarily by the soil type in which the ectomycorrhizas were growing. Naturally colonized fir seedlings were reciprocally transplanted between clearcuts and forests, carrying different EMF communities with them. EEAs were influenced more by destination environment than by EMF community. EEAs were as high in early-successional as in late-successional communities in both destination environments. Buds of clearcut-origin seedlings had the same or higher N contents as forest seedlings after a growing season in either environment. These results indicate that (i) symbiotic EMF and/or their associated microbial communities demonstrate substantial ability to acclimate to new field environments; (ii) the ability to produce organic matter-degrading enzymes is not a trait that necessarily distinguishes early- and late-successional EMF communities in symbiosis; (iii) early-successional EMF are as capable of supporting seedling N accumulation in forest soils as late-successional EMF; and (iv) disturbed ecosystems where early-successional EMF are present should have high resilience for organic matter degradation.

Structural features of the aromatic/arginine constriction in the aquaglyceroporin GintAQPF2 are responsible for glycerol impermeability in arbuscular mycorrhizal symbiosis.

Carbon transport in arbuscular mycorrhizal (AM) symbiosis is of fundamental importance. However, the role of glycerol transport in AM symbiosis has not yet been resolved. Glycerol transport across the cell membrane is mediated by aquaglyceroporins (AQGPs), whereas our previous study revealed that it was disfavoured by GintAQPF2, an AQGP from AM fungi (AMF). Here, we analysed the function of two amino acid residues in the aromatic/arginine (ar/R) constriction known as the major selectivity filter in AQGPs. Replacement of phenylalanine-94 (Phe-94) by alanine (Ala) enlarged the diameter of the ar/R constriction and resulted in an increased intracellular glycerol accumulation and thus survival rate of yeast cells at high glycerol levels, while individual or joint replacement of Phe-94 and Ala-234 by tryptophan and glycine induced a closed state of GintAQPF2, suggesting that the potential double gates (Phe94-Phe243 and arginine-249) of the ar/R constriction also likely determined solute permeability. To figure out whether GintAQPF2 functions were relevant to the establishment of AM symbiosis, genomic analyses of four representative fungi with different lifestyles were performed. We found that glycerol facilitators existed in the facultative fungi (the ectomycorrhizal fungus Laccaria bicolor and hemibiotrophic pathogen Magnaporthe oryzae), but not in the obligatory fungi (the AMF Rhizophagus irregularis and necrotrophic pathogen Fusarium verticillioides), revealing a conserved pattern of glycerol transport in symbionts and pathogens. Our results suggested that glycerol blocks due to the special structural features of the ar/R constriction in the only AMF AQGP could potentially play a role in the establishment of AM symbiosis.

Characterization and purification of a bacterial chlorogenic acid esterase detected during the extraction of chlorogenic acid from arbuscular mycorrhizal tomato roots.

A Gram-negative bacterium able to grow using chlorogenic acid (5-caffeoylquinic acid) as sole carbon source has been isolated from the roots of tomato plants inoculated with the arbuscular mycorrhizal fungus Rhizophagus irregularis. An intracellular esterase exhibiting very high affinity (Km = 2 µM) for chlorogenic acid has been extracted and purified by FPLC from the chlorogenate-grown cultures of this bacterium. The molecular mass of the purified esterase determined by SDS-PAGE was 61 kDa and its isoelectric point determined by chromatofocusing was 7.75. The esterase hydrolysed chlorogenic acid analogues (caffeoylshikimate, and the 4- and 3-caffeoylquinic acid isomers), feruloyl esterases substrates (methyl caffeate and methyl ferulate), and even caffeoyl-CoA in vitro but all of them were less active than chlorogenic acid, demonstrating that the esterase is a genuine chlorogenic acid esterase. It was also induced when the bacterial strain was cultured in the presence of hydroxycinnamic acids (caffeic, p-coumaric or ferulic acid) as sole carbon source, but not in the presence of simple phenolics such as catechol or protocatechuic acid, nor in the presence of organic acids such as succinic or quinic acids. The purified esterase was remarkably stable in the presence of methanol, rapid formation of methyl caffeate occurring when its activity was measured in aqueous solutions containing 10-60% methanol. Our results therefore show that this bacterial chlorogenase can catalyse the transesterification reaction previously detected during the methanolic extraction of chlorogenic acid from arbuscular mycorrhizal tomato roots. Data are presented suggesting that colonisation by Rhizophagus irregularis could increase chlorogenic acid exudation from tomato roots, especially in nutrient-deprived plants, and thus favour the growth of chlorogenate-metabolizing bacteria on the root surface or in the mycorhizosphere.

Biochemical and ecophysiological responses to manganese stress by ectomycorrhizal fungus Pisolithus tinctorius and in association with Eucalyptus grandis.

At relatively low concentrations, the element manganese (Mn) is essential for plant metabolism, especially for photosynthesis and as an enzyme antioxidant cofactor. However, industrial and agricultural activities have greatly increased Mn concentrations, and thereby contamination, in soils. We tested whether and how growth of Pisolithus tinctorius is influenced by Mn and glucose and compare the activities of oxidative stress enzymes as biochemical markers of Mn stress. We also compared nutrient accumulation, ecophysiology, and biochemical responses in Eucalyptus grandis which had been colonized by the ectomycorrhizal Pisolithus tinctorius with those which had not, when both were exposed to increasing Mn concentrations. In vitro experiments comprised six concentrations of Mn in three concentrations of glucose. In vivo experiments used plants colonized by Pisolithus tinctorius, or not colonized, grown with three concentrations of Mn (0, 200, and 1000 µM). We found that fungal growth and glucose concentration were correlated, but these were not influenced by Mn levels in the medium. The anti-oxidative enzymes catalase and glutathione S-transferase were both activated when the fungus was exposed to Mn. Also, mycorrhizal plants grew more and faster than non-mycorrhizal plants, whatever Mn exposure. Photosynthesis rate, intrinsic water use efficiency, and carboxylation efficiency were all inversely correlated with Mn concentration. Thus, we originally show that the ectomycorrhizal fungus provides protection for its host plants against varying and potentially toxic concentrations of Mn.

Ectomycorrhizal Fungal Protein Degradation Ability Predicted by Soil Organic Nitrogen Availability.

In temperate and boreal forest ecosystems, nitrogen (N) limitation of tree metabolism is alleviated by ectomycorrhizal (ECM) fungi. As forest soils age, the primary source of N in soil switches from inorganic (NH4 (+) and NO3 (-)) to organic (mostly proteins). It has been hypothesized that ECM fungi adapt to the most common N source in their environment, which implies that fungi growing in older forests would have greater protein degradation abilities. Moreover, recent results for a model ECM fungal species suggest that organic N uptake requires a glucose supply. To test the generality of these hypotheses, we screened 55 strains of 13 Suillus species with different ecological preferences for their in vitro protein degradation abilities. Suillus species preferentially occurring in mature forests, where soil contains more organic matter, had significantly higher protease activity than those from young forests with low-organic-matter soils or species indifferent to forest age. Within species, the protease activities of ecotypes from soils with high or low soil organic N content did not differ significantly, suggesting resource partitioning between mineral and organic soil layers. The secreted protease mixtures were strongly dominated by aspartic peptidases. Glucose addition had variable effects on secreted protease activity; in some species, it triggered activity, but in others, activity was repressed at high concentrations. Collectively, our results indicate that protease activity, a key ectomycorrhizal functional trait, is positively related to environmental N source availability but is also influenced by additional factors, such as carbon availability.

Effect of Inoculation with Glomus versiforme on Cadmium Accumulation, Antioxidant Activities and Phytochelatins of Solanum photeinocarpum.

The plant growth, phosphate acquisition, Cd translocation, phytochelatins (PCs) production and antioxidant parameters [superoxide dismutase (SOD), catalase (CAT), guaiacol peroxidase (POD), ascorbate peroxidase (APX), glutathione reductase (GR), glutathione (GSH), ascorbate (ASA) and malonaldehyde (MDA)] were investigated in Cd-hyperaccumulator Solanum photeinocarpum inoculated with Glomus versiforme BGC GD01C (Gv) in Cd-added soils (0, 5, 10, 20, 40 mg Cd kg-1 soil). Mycorrhizal colonization rates were generally high (from 77% to 94%), and hardly affected by Cd. Gv colonization significantly enhanced P acquisition, growth and total Cd uptakes in both shoots and roots of S. photeinocarpum at all Cd levels. Meanwhile, Gv symbiosis significantly increased Cd concentration in the roots, and decreased Cd concentration in the shoots at all Cd levels, which indicates that Gv could promote phytostabilization by enhancing Cd accumulation in the roots to inhibit its translocation to shoots and the "dilution effects" linked to an increase in plant dry matter yield and a reduced Cd partitioning to shoots. Moreover, the improvement of CAT, POD and APX activities in the leaves of mycorrhizal plants infers that Gv symbiosis helped S. photeinocarpum to relieve oxidative damage to biomolecules in Cd-contaminated soil. The evident decline of MDA content in the leaves of mycorrhizal plants indicates that Gv symbiosis evidently improved antioxidant activities, and the enhancement of PCs production in the leaves of mycorrhizal plants suggests that Gv-inoculated plant may be more efficient to relieve Cd phytotoxicity. Therefore, the possible mechanisms of Cd phytotoxicity alleviation by Gv can be concluded as the decline of Cd concentration in the shoots and the improvement of P acquisition, PCs production and activities of CAT, POD, APX in mycorrhizal plants.

Arbuscular mycorrhizal fungal diversity, root colonization, and soil alkaline phosphatase activity in response to maize-wheat rotation and no-tillage in North China.

Monitoring the effects of no-tillage (NT) in comparison with conventional tillage (CT) on soil microbes could improve our understanding of soil biochemical processes and thus help us to develop sound management strategies. The objective of this study was to compare the species composition and ecological function of soil arbuscular mycorrhizal (AM) fungi during the growth and rotation of crops under NT and CT. From late June 2009 to early June 2010, 32 topsoil (0-15 cm) samples from four individual plots per treatment (CT and NT) were collected at both the jointing and maturation stages of maize (Zea mays L.) and wheat (Triticum aestivum L.) from a long-term experimental field that was established in an Aquic Inceptisol in North China in June 2006. The AM fungal spores were isolated and identified and then used to calculate species diversity indices, including the Shannon- Wiener index (H'), Evenness (E), and Simpson's index (D). The root mycorrhizal colonization and soil alkaline phosphatase activity were also determined. A total of 34 species of AM fungi within nine genera were recorded. Compared with NT, CT negatively affected the soil AM fungal community at the maize sowing stage, leading to decreases in the average diversity indices (from 2.12, 0.79, and 0.82 to 1.79, 0.72, and 0.74 for H', E, and D, respectively), root mycorrhizal colonization (from 28% to 20%), soil alkaline phosphatase activity (from 0.24 to 0.19 mg/g/24 h) and available phosphorus concentration (from 17.4 to 10.5 mg/kg) at the maize jointing stage. However, reductions in diversity indices of H', E, and D were restored to 2.20, 0.81, and 0.84, respectively, at the maize maturation stage. CT should affect the community again at the wheat sowing stage; however, a similar restoration in the species diversity of AM fungi was completed before the wheat jointing stage, and the highest Jaccard index (0.800) for similarity in the species composition of soil AM fungi between CT and NT was recorded at the wheat maturation stage. Our results also demonstrated that NT resulted in the positive protection of the community structure of AM fungi and played an important role in maintaining their functionality especially for maize seedlings.

Acid protease production in fungal root endophytes.

Fungal endophytes are ubiquitous in healthy root tissue, but little is known about their ecosystem functions, including their ability to utilize organic nutrient sources such as proteins. Root-associated fungi may secrete proteases to access the carbon and mineral nutrients within proteins in the soil or in the cells of their plant host. We compared the protein utilization patterns of multiple isolates of the root endophytes Phialocephala fortinii s.l., Meliniomyces variabilis and Umbelopsis isabellina with those of two ectomycorrhizal (ECM) fungi, Hebeloma incarnatulum and Laccaria bicolor, and the wood-decay fungus Irpex lacteus at pH values of 2-9 on liquid BSA media. We also assessed protease activity using a fluorescently labeled casein assay and gelatin zymography and characterized proteases using specific protease inhibitors. I. lacteus and U. isabellina utilized protein efficiently, while the ECM fungi exhibited poor protein utilization. ECM fungi secreted metallo-proteases and had pH optima above 4, while other fungi produced aspartic proteases with lower pH optima. The ascomycetous root endophytes M. variabilis and P. fortinii exhibited intermediate levels of protein utilization and M. variabilis exhibited a very low pH optimum. Comparing proteolytic profiles between fungal root endophytes and fungi with well defined ecological roles provides insight into the ecology of these cryptic root associates.