The root-associated arbuscular mycorrhizal fungal assemblages of exotic alien plants are simplified in invaded distribution ranges, but dominant species are retained: A trans-continental perspective.

Arbuscular mycorrhizal fungi (AMF) provide crucial support for the establishment of plants in novel environments. We hypothesized that the OTU/genus richness and diversity of soil- and root-associated AMF associated with alien plant species in their exotic ranges are lower than those in their native ranges. We examined the root-associated and soil-dwelling AMF of 11 invasive plant species in their native and exotic ranges in the United States and Europe by DNA sequencing of the ITS2 locus. Examined root-associated AMF assemblages were simplified, which manifested as the loss of several AMF genera in the exotic ranges of the plants. These fungal assemblages were also characterized by greater dominance and simplification of the fungal assemblages. The dominant fungal genera were present regardless of whether their host plants were in their native or exotic ranges. Interestingly, both the native and invaded soils hosted diverse local AMF assemblages. Therefore, alien plant invasions were not limited to soils with low AMF diversity. Some AMF taxa could be context-dependent passengers rather than drivers of alien plant invasions. Further studies should identify functions of AMF missing or less abundant in roots of plants growing in exotic ranges.

Study of the effects of mineral salts on the biofilm formation on polypropylene fibers using three quantification methods.

The microbial biofilms are ubiquitous in nature and represent important biological entities that affect various aspects of human life. As such, they attracted considerable attention during last decades, with the factors affecting the biofilm development being among the frequently studied topics. In our work, the biofilm was cultivated on the surface of polypropylene fibers in a nutrient medium inoculated by the suspension of two unsterile soils. The effects of ionic strength and valence of salt on the amount of the produced biofilm and on composition of biofilm microbial communities were investigated. The effect of valence was significant in some OTUs: Arthrobacter/Pseudarthrobacter/Paenarthrobacter and Bacillus with positive response to monovalent salt (KCl) and Streptomyces, Lysinibacillus, Pseudomonas, and Ensifer with positive response to divalent salt (MgSO4). The significant preference for a certain concentration of salts was observed in the case of OTUs Agrobacterium, Bacillus (both 100 mM), and Brevundimonas (30 mM). A new quantification method based on measuring of oxidizable organic carbon in biofilm biomass, based on dichromate oxidation, was used. We compared the results obtained using this method with results of crystal violet destaining and measuring of extracted DNA concentration as proxies of the biofilm biomass. The dichromate oxidation is simple, inexpensive, and fast, and our results show that it may be more sensitive than crystal violet destaining. The highest biomass values tended to associate with high concentrations of the divalent salt. This trend was not observed in treatments where the monovalent salt was added. Our data confirm the importance of inorganic ions for biofilm composition and biomass accumulation.

Biofilm and planktonic microbial communities in highly acidic soil (pH < 3) in the Soos National Nature Reserve, Czech Republic.

Biofilm formation is a typical life strategy used by microorganisms populating acidic water systems. The same strategy might be used by microbes in highly acidic soils that are, however, neglected in this regard. In the present study, the microbial community in such highly acidic soil in the Soos National Nature Reserve (Czech Republic) has been investigated using high-throughput DNA sequencing and the organisms associated with biofilm life mode and those preferring planktonic life were distinguished using the biofilm trap technique. Our data show the differences between biofilm and planktonic microbiota fraction, although the majority of the organisms were capable of using both life modes. The by far most abundant prokaryotic genus was Acidiphilium and fungi were identified among the most abundant eukaryotic elements in biofilm formations. On the other hand, small flagellates from diverse taxonomical groups predominated in plankton. The application of cellulose amendment as well as the depth of sampling significantly influenced the composition of the detected microbial community.

Dead Rhizophagus irregularis biomass mysteriously stimulates plant growth.

Arbuscular mycorrhizal (AM) fungi establish symbiotic associations with many plant species, transferring significant amounts of soil nutrients such as phosphorus to plants and receiving photosynthetically fixed carbon in return. Functioning of AM symbiosis is thus based on interaction between two living partners. The importance of dead AM fungal biomass (necromass) in ecosystem processes remains unclear. Here, we applied either living biomass or necromass (0.0004 potting substrate weight percent) of monoxenically produced AM fungus (Rhizophagus irregularis) into previously sterilized potting substrate planted with Andropogon gerardii. Plant biomass production significantly improved in both treatments as compared to non-amended controls. Living AM fungus, in contrast to the necromass, specifically improved plant acquisition of nutrients normally supplied to the plants by AM fungal networks, such as phosphorus and zinc. There was, however, no difference between the two amendment treatments with respect to plant uptake of other nutrients such as nitrogen and/or magnesium, indicating that the effect on plants of the AM fungal necromass was not primarily nutritional. Plant growth stimulation by the necromass could thus be either due to AM fungal metabolites directly affecting the plants, indirectly due to changes in soil/root microbiomes or due to physicochemical modifications of the potting substrate. In the necromass, we identified several potentially bioactive molecules. We also provide experimental evidence for significant differences in underground microbiomes depending on the amendment with living or dead AM fungal biomass. This research thus provides the first glimpse into possible mechanisms responsible for observed plant growth stimulation by the AM fungal necromass.

Geography and habitat predominate over climate influences on arbuscular mycorrhizal fungal communities of mid-European meadows.

Despite the crucial importance of arbuscular mycorrhizal fungi (AMF) for numerous processes within terrestrial ecosystems, knowledge of the determinants of AMF community structure still is limited, mainly because of the limited scope of the available individual case studies which often only include a few environmental variables. Here, we describe the AMF diversity of mid-European meadows (mown or regularly cut grasslands, or recently abandoned lands where grasslands established spontaneously) within a considerably heterogeneous landscape over a scale of several hundred kilometers with regard to macroclimatic, microclimatic, and soil parameters. We include data describing the habitat (including vegetation type), geography, and climate, and test their contribution to the structure of the AMF communities at a regional scale. We amplified and sequenced the ITS 2 region of the ribosomal DNA operon of the AMF from soil samples using nested PCR and Illumina pair-end amplicon sequencing. Habitat (especially soil pH) and geographical parameters (spatial distance, altitude, and longitude) were the main determinants of the structure of the AMF communities in the meadows at a regional scale, with the abundance of genera Septoglomus, Paraglomus, Archaeospora, Funneliformis, and Dominikia driving the main response. The effects of climate and vegetation type were not significant and were mainly encompassed within the geography and/or soil pH effects. This study illustrates how important it is to have a large set of environmental metadata to compare the importance of different factors influencing the AMF community structure at large spatial scales.

Disentangling the factors of contrasting silver and copper accumulation in sporocarps of the ectomycorrhizal fungus Amanita strobiliformis from two sites.

Amanita strobiliformis (European Pine Cone Lepidella) is an ectomycorrhizal fungus of the Amanitaceae family known to hyperaccumulate Ag in the sporocarps. Two populations (ecotypes) of A. strobiliformis collected from two urban forest plantations in Prague, Czech Republic, were investigated. The concentrations of Ag, Cu, Cd, and Zn were determined in the mushrooms. The metal mobility and fractionation in the soils was investigated by single extractions and sequential extraction. The soil distribution of A. strobiliformis mycelium was assessed by quantitative polymerase chain reaction (qPCR). The metal uptake from the soil into the mushroom sporocarps was traced by Pb isotopic fingerprinting. The findings suggested that A. strobiliformis (i) accumulates primarily Ag from the topsoil layer (circa 12cm deep) and (ii) accumulates Ag associated with the "reducible soil fraction". The concentrations of all metals, particularly Ag and Cu, were significantly higher in the A. strobiliformis sporocarps from one of the investigated sites (Klícov). The elevated concentrations of Ag in the sporocarps from Klícov can possibly be attributed to the higher Ag content in the topsoil layer found at this site. However, the simultaneously elevated concentrations of Cu in A. strobiliformis from Klícov cannot be explained by the differences in the geochemical background and should be attributed to biological factors.

Fungi, a neglected component of acidophilic biofilms: do they have a potential for biotechnology?

Fungi from extreme environments, including acidophilic ones, belong to biotechnologically most attractive organisms. They can serve as a source of enzymes and metabolites with potentially uncommon properties and may actively participate within bioremediation processes. In respect of their biotechnological potential, extremophilic fungi are mostly studied as individual species. Nevertheless, microorganisms rarely live separately and they form biofilms instead. Living in biofilms is the most successful life strategy on the Earth and the biofilm is the most abundant form of life in extreme environments including highly acidic ones. Compared to bacterial fraction, fungal part of acidophilic biofilms represents a largely unexplored source of organisms with possible use in biotechnology and especially data on biofilms of highly acidic soils are missing. The functioning of the biofilm results from interactions between organisms whose metabolic capabilities are efficiently combined. When we look on acidophilic fungi and their biotechnological potential we should take this fact into account as well. The practical problem to be resolved in connection with extensive studies of exploitable properties and abilities of acidophilic fungi is the methodology of isolation of strains from the nature. In this respect, novel isolation techniques should be developed.

Soil Matrix Determines the Outcome of Interaction Between Mycorrhizal Symbiosis and Biochar for Andropogon gerardii Growth and Nutrition.

Biochar has been heralded as a multipurpose soil amendment to sustainably increase soil fertility and crop yields, affect soil hydraulic properties, reduce nutrient losses, and sequester carbon. Some of the most spectacular results of biochar (and organic nutrient) inputs are the terra preta soils in the Amazon, dark anthropogenic soils with extremely high fertility sustained over centuries. Such soil improvements have been particularly difficult to achieve on a short run, leading to speculations that biochar may need to age (weather) in soil to show its best. Further, interaction of biochar with arbuscular mycorrhizal fungi (AMF), important root symbionts of a great majority of terrestrial plants including most agricultural crops, remains little explored. To study the effect of aged biochar on highly mycotrophic Andropogon gerardii plants and their associated AMF, we made use of softwood biochar, collected from a historic charcoal burning site. This biochar (either untreated or chemically activated, the latter serving as a proxy for freshly prepared biochar) was added into two agricultural soils (acid or alkaline), and compared to soils without biochar. These treatments were further crossed with inoculation with a synthetic AMF community to address possible interactions between biochar and the AMF. Biochar application was generally detrimental for growth and mineral nutrition of our experimental plants, but had no effect on the extent of their root colonized by the AMF, nor did it affect composition of their root-borne AMF communities. In contrast, biochar affected development of two out of five AMF (Claroideoglomus and Funneliformis) in the soil. Establishment of symbiosis with AMF largely mitigated biochar-induced suppression of plant growth and mineral nutrition, mainly by improving plant acquisition of phosphorus. Both mycorrhizal and non-mycorrhizal plants grew well in the acid soil without biochar application, whereas non-mycorrhizal plants remained stunted in the alkaline soils under all situations (with or without biochar). These different and strong effects indicate that response of plants to biochar application are largely dependent on soil matrix and also on microbes such as AMF, and call for further research to enable qualified predictions of the effects of different biochar applications on field-grown crops and soil processes.

Utilization of organic nitrogen by arbuscular mycorrhizal fungi-is there a specific role for protists and ammonia oxidizers?

Arbuscular mycorrhizal (AM) fungi can significantly contribute to plant nitrogen (N) uptake from complex organic sources, most likely in concert with activity of soil saprotrophs and other microbes releasing and transforming the N bound in organic forms. Here, we tested whether AM fungus (Rhizophagus irregularis) extraradical hyphal networks showed any preferences towards certain forms of organic N (chitin of fungal or crustacean origin, DNA, clover biomass, or albumin) administered in spatially discrete patches, and how the presence of AM fungal hyphae affected other microbes. By direct 15N labeling, we also quantified the flux of N to the plants (Andropogon gerardii) through the AM fungal hyphae from fungal chitin and from clover biomass. The AM fungal hyphae colonized patches supplemented with organic N sources significantly more than those receiving only mineral nutrients, organic carbon in form of cellulose, or nothing. Mycorrhizal plants grew 6.4-fold larger and accumulated, on average, 20.3-fold more 15N originating from the labeled organic sources than their nonmycorrhizal counterparts. Whereas the abundance of microbes (bacteria, fungi, or Acanthamoeba sp.) in the different patches was primarily driven by patch quality, we noted a consistent suppression of the microbial abundances by the presence of AM fungal hyphae. This suppression was particularly strong for ammonia oxidizing bacteria. Our results indicate that AM fungi successfully competed with the other microbes for free ammonium ions and suggest an important role for the notoriously understudied soil protists to play in recycling organic N from soil to plants via AM fungal hyphae.

Appropriate nonmycorrhizal controls in arbuscular mycorrhiza research: a microbiome perspective.

Establishment of nonmycorrhizal controls is a "classic and recurrent theme" in mycorrhizal research. For decades, authors reported mycorrhizal plant growth/nutrition as compared to various nonmycorrhizal controls. In such studies, uncertainties remain about which nonmycorrhizal controls are most appropriate and, in particular, what effects the control inoculations have on substrate and root microbiomes. Here, different types of control and mycorrhizal inoculations were compared with respect to plant growth and nutrition, as well as the structure of root and substrate microbiomes, assessed by next-generation sequencing. We compared uninoculated ("absolute") control to inoculation with blank pot culture lacking arbuscular mycorrhizal fungi, filtrate of that blank inoculum, and filtrate of complex pot-produced mycorrhizal inoculum. Those treatments were compared to a standard mycorrhizal treatment, where the previously sterilized substrate was inoculated with complex pot-produced inoculum containing Rhizophagus irregularis SYM5. Besides this, monoxenically produced inoculum of the same fungus was applied either alone or in combination with blank inoculum. The results indicate that the presence of mycorrhizal fungus always resulted in stimulation of Andropogon gerardii plant biomass as well as in elevated phosphorus content of the plants. The microbial (bacterial and fungal) communities developing in the differently inoculated treatments, however, differed substantially from each other and no control could be obtained comparable with the treatment inoculated with complex mycorrhizal inoculum. Soil microorganisms with significant biological competences that could potentially contribute to the effects of the various inoculants on the plants were detected in roots and in plant cultivation substrate in some of the treatments.

Utilization of organic nitrogen by arbuscular mycorrhizal fungi-is there a specific role for protists and ammonia oxidizers?

Arbuscular mycorrhizal (AM) fungi can significantly contribute to plant nitrogen (N) uptake from complex organic sources, most likely in concert with activity of soil saprotrophs and other microbes releasing and transforming the N bound in organic forms. Here, we tested whether AM fungus (Rhizophagus irregularis) extraradical hyphal networks showed any preferences towards certain forms of organic N (chitin of fungal or crustacean origin, DNA, clover biomass, or albumin) administered in spatially discrete patches, and how the presence of AM fungal hyphae affected other microbes. By direct 15N labeling, we also quantified the flux of N to the plants (Andropogon gerardii) through the AM fungal hyphae from fungal chitin and from clover biomass. The AM fungal hyphae colonized patches supplemented with organic N sources significantly more than those receiving only mineral nutrients, organic carbon in form of cellulose, or nothing. Mycorrhizal plants grew 6.4-fold larger and accumulated, on average, 20.3-fold more 15N originating from the labeled organic sources than their nonmycorrhizal counterparts. Whereas the abundance of microbes (bacteria, fungi, or Acanthamoeba sp.) in the different patches was primarily driven by patch quality, we noted a consistent suppression of the microbial abundances by the presence of AM fungal hyphae. This suppression was particularly strong for ammonia oxidizing bacteria. Our results indicate that AM fungi successfully competed with the other microbes for free ammonium ions and suggest an important role for the notoriously understudied soil protists to play in recycling organic N from soil to plants via AM fungal hyphae.

Soil-derived organic particles and their effects on the community of culturable microorganisms.

Soil microbial community interacts with a range of particulate material in the soil, consisting of both inorganic and organic compounds with different levels of water solubility. Though sparingly water-soluble and insoluble organic compounds in the soil may affect living organisms, they are difficult to introduce into microbiological media. Their biological activity (i.e., their effect on soil microorganisms) thus has been almost neglected in most of the cultivation assays. To fill this gap, we propose the use of fine organic particles prepared from soil organic matter that are introduced into a laboratory medium where microbial community is cultivated. To this purpose, submicrometer particles consisting of sparingly water-soluble or insoluble soil organic matter were obtained from humic horizons of two soils by precipitation of organics dissolved in tetrahydrofuran by addition of water. The particles could then be size fractionated by centrifugation, and coarse fraction obtained from humic horizon formed under spruce forest was tested for effects on complex microbial community developing under laboratory conditions. The results indicate that low concentration (20 mg/L) of the particles is efficient to affect the composition of the bacterial community revealed by terminal restriction fragment length polymorphism. The work contributes to understanding the factors that determine the composition of soil microbial community.

Truffle biogeography-A case study revealing ecological niche separation of different Tuber species.

Ecology of hypogeic mycorrhizal fungi, such as truffles, remains largely unknown, both in terms of their geographical distribution and their environmental niches. Occurrence of true truffles (Tuber spp.) was therefore screened using specific polymerase chain reaction (PCR) assays and subsequent PCR amplicon sequencing in tree roots collected at 322 field sites across the Czech Republic. These sites spanned a wide range of climatic and soil conditions. The sampling was a priori restricted to areas thought to be suitable for Tuber spp. inasmuch as they were characterized by weakly acidic to alkaline soils, warmer climate, and with tree species previously known to host true truffles. Eight operational taxonomic units (OTUs) corresponding to Tuber aestivum, T. borchii, T. foetidum, T. rufum, T. indicum, T. huidongense, T. dryophilum, and T. oligospermum were detected. Among these, T. borchii was the OTU encountered most frequently. It was detected at nearly 19% of the sites. Soil pH was the most important predictor of Tuber spp. distribution. Tuber borchii preferred weakly acidic soils, T. foetidum and T. rufum were most abundant in neutral soils, and T. huidongense was restricted to alkaline soils. Distribution of T. aestivum was mainly dictated by climate, with its range restricted to the warmest sites. Host preferences of the individual Tuber spp. were weak compared to soil and climatic predictors, with the notable exception that T. foetidum appeared to avoid oak trees. Our results open the way to better understanding truffle ecology and, through this new knowledge, also to better-informed trufficulture.

Resurrection of Cortinarius coalescens: taxonomy, chemistry, and ecology.

Cortinarius coalescens Kärcher & Seibt is a rare European species of the subgenus Phlegmacium, section Phlegmacioides, neglected in recent molecular studies. New primers (CortF and CortR) designed for species in the section Phlegmacioides allowed to obtain ITS rDNA sequence data from the holotype collection of C. coalescens; according to the results, this epithet has priority over C. crassorum Rob. Henry ex Rob. Henry, C. pardinus Reumaux, and C. parargutus Bidaud, Moënne-Locc. & Reumaux. Morphological and ecological observations on recent collections of C. coalescens from the Czech Republic in comparison with the co-occurring C. largus are discussed. Nomenclatural and taxonomic comments on C. tomentosus Rob. Henry, C. balteatotomentosus Rob. Henry, and C. subtomentosus Reumaux are also provided. So far, C. coalescens is known with certainty from Germany, France, and the Czech Republic, where it grows in deciduous forests on acid to neutral soils. Arsenic and its compounds were determined in C. coalescens and related species of the section Phlegmacioides: C. largus, C. pseudodaulnoyae, and C. variecolor. Total arsenic concentrations were in the range 3.6-30.2 mg kg-1 (dry matter) and arsenobetaine was the major arsenic compound.

Monitoring CO2 emissions to gain a dynamic view of carbon allocation to arbuscular mycorrhizal fungi.

Quantification of carbon (C) fluxes in mycorrhizal plants is one of the important yet little explored tasks of mycorrhizal physiology and ecology. 13CO2 pulse-chase labelling experiments are increasingly being used to track the fate of C in these plant-microbial symbioses. Nevertheless, continuous monitoring of both the below- and aboveground CO2 emissions remains a challenge, although it is necessary to establish the full C budget of mycorrhizal plants. Here, a novel CO2 collection system is presented which allows assessment of gaseous CO2 emissions (including isotopic composition of their C) from both belowground and shoot compartments. This system then is used to quantify the allocation of recently fixed C in mycorrhizal versus nonmycorrhizal Medicago truncatula plants with comparable biomass and mineral nutrition. Using this system, we confirmed substantially greater belowground C drain in mycorrhizal versus nonmycorrhizal plants, with the belowground CO2 emissions showing large variation because of fluctuating environmental conditions in the glasshouse. Based on the assembled 13C budget, the C allocation to the mycorrhizal fungus was between 2.3% (increased 13C allocation to mycorrhizal substrate) and 2.9% (reduction of 13C allocation to mycorrhizal shoots) of the plant gross photosynthetic production. Although the C allocation to shoot respiration (measured during one night only) did not differ between the mycorrhizal and nonmycorrhizal plants under our experimental conditions, it presented a substantial part (∼10%) of the plant C budget, comparable to the amount of CO2 released belowground. These results advocate quantification of both above- and belowground CO2 emissions in future studies.

Bioaccumulation of heavy metals, metalloids, and chlorine in ectomycorrhizae from smelter-polluted area.

Ectomycorrhizal (ECM) fungi contribute to the survival of host trees on metal-rich soils by reducing the transfer of toxic metals into roots. However, little is known about the ability of ECM fungi to accumulate elements in ectomycorrhizae (ECMs). Here we report Ag, As, Cd, Cl, Cu, Sb, V, and Zn contents in wild-grown Norway spruce ECMs collected in a smelter-polluted area at Lhota near Príbram, Czech Republic. The ECMs data were compared with the element concentrations determined in the corresponding non-mycorrhizal fine roots, soils, and soil extracts. Bioaccumulation factors were calculated to differentiate the element accumulation ability of ECMs inhabited by different mycobionts, which were identified by ITS rDNA sequencing. Among the target elements, the highest contents were observed for Ag, Cl, Cd, and Zn; Imleria badia ECMs showed the highest capability to accumulate these elements. ECMs of Amanita muscaria, but not of other species, accumulated V. The analysis of the proportions of I. badia and A. muscaria mycelia in ECMs by using species-specific quantitative real-time PCR revealed variable extent of the colonization of roots, with median values close to 5% (w/w). Calculated Ag, Cd, Zn and Cl concentrations in the mycelium of I. badia ECMs were 1 680, 1 510, 2 670, and 37,100 mg kg-1 dry weight, respectively, indicating substantial element accumulation capacity of hyphae of this species in ECMs. Our data strengthen the idea of an active role of ECM fungi in soil-fungal-plant interactions in polluted environments.

Organic Nitrogen-Driven Stimulation of Arbuscular Mycorrhizal Fungal Hyphae Correlates with Abundance of Ammonia Oxidizers.

Large fraction of mineral nutrients in natural soil environments is recycled from complex and heterogeneously distributed organic sources. These sources are explored by both roots and associated mycorrhizal fungi. However, the mechanisms behind the responses of arbuscular mycorrhizal (AM) hyphal networks to soil organic patches of different qualities remain little understood. Therefore, we conducted a multiple-choice experiment examining hyphal responses to different soil patches within the root-free zone by two AM fungal species (Rhizophagus irregularis and Claroideoglomus claroideum) associated with Medicago truncatula, a legume forming nitrogen-fixing root nodules. Hyphal colonization of the patches was assessed microscopically and by quantitative real-time PCR (qPCR) using AM taxon-specific markers, and the prokaryotic and fungal communities in the patches (pooled per organic amendment treatment) were profiled by 454-amplicon sequencing. Specific qPCR markers were then designed and used to quantify the abundance of prokaryotic taxa showing the strongest correlation with the pattern of AM hyphal proliferation in the organic patches as per the 454-sequencing. The hyphal density of both AM fungi increased due to nitrogen (N)-containing organic amendments (i.e., chitin, DNA, albumin, and clover biomass), while no responses as compared to the non-amended soil patch were recorded for cellulose, phytate, or inorganic phosphate amendments. Abundances of several prokaryotes, including Nitrosospira sp. (an ammonium oxidizer) and an unknown prokaryote with affiliation to Acanthamoeba endosymbiont, which were frequently recorded in the 454-sequencing profiles, correlated positively with the hyphal responses of R. irregularis to the soil amendments. Strong correlation between abundance of these two prokaryotes and the hyphal responses to organic soil amendments by both AM fungi was then confirmed by qPCR analyses using all individual replicate patch samples. Further research is warranted to ascertain the causality of these correlations and particularly which direct roles (if any) do these prokaryotes play in the observed AM hyphal responses to organic N amendment, organic N utilization by the AM fungus and its (N-unlimited) host plant. Further, possible trophic dependencies between the different players in the AM hyphosphere needs to be elucidated upon decomposing the organic N sources.

Genetic transformation of extremophilic fungi Acidea extrema and Acidothrix acidophila.

Intact, growing cells of strongly acidophilic fungi Acidea extrema and Acidothrix acidophila have been successfully transformed by introduction of heterologous DNA fragment (composed of the glyceraldehyde-phosphate-dehydrogenase gene promoter from Emericella nidulans, a metallothionein-coding gene AsMt1 from Amanita strobiliformis and glyceraldehyde-phosphate-dehydrogenase gene terminator from Colletotrichum gloeosporioides) with the length of 1690 bp. The transformation procedure was based on the DNA transfer mediated by Agrobacterium tumefaciens bearing disarmed helper plasmid pMP90 and binary vector pCambia1300 with inserted DNA fragment of interest. The transformants proved to be mitotically stable, and the introduced gene was expressed at least at the level of transcription. Our work confirms that metabolic adaptations of strongly acidophilic fungi do not represent an obstacle for genetic transformation using conventional methods and can be potentially used for production of heterologous proteins. A promising role of the fast growing A. acidophila as active biomass in biotechnological processes is suggested not only by the low susceptibility of the culture grown at low pH to contaminations but also by reduced risk of accidental leaks of genetically modified microorganisms into the environment because highly specialized extremophilic fungi can poorly compete with common microflora under moderate conditions.

Duration and intensity of shade differentially affects mycorrhizal growth- and phosphorus uptake responses of Medicago truncatula.

Plant and fungal partners in arbuscular mycorrhizal symbiosis trade mineral nutrients for carbon, with the outcome of this relationship for plant growth and nutrition being highly context-dependent and changing with the availability of resources as well as with the specific requirements of the different partners. Here we studied how the model legume Medicago truncatula, inoculated or not with a mycorrhizal fungus Rhizophagus irregularis, responded to a gradient of light intensities applied over different periods of time, in terms of growth, phosphorus nutrition and the levels of root colonization by the mycorrhizal fungus. Short-term (6 d) shading, depending on its intensity, resulted in a rapid decline of phosphorus uptake to the shoots of mycorrhizal plants and simultaneous accumulation of phosphorus in the roots (most likely in the fungal tissues), as compared to the non-mycorrhizal controls. There was, however, no significant change in the levels of mycorrhizal colonization of roots due to short-term shading. Long-term (38 d) shading, depending on its intensity, provoked a multitude of plant compensatory mechanisms, which were further boosted by the mycorrhizal symbiosis. Mycorrhizal growth- and phosphorus uptake benefits, however, vanished at 10% of the full light intensity applied over a long-term. Levels of root colonization by the mycorrhizal fungus were significantly reduced by long-term shading. Our results indicate that even short periods of shade could have important consequences for the functioning of mycorrhizal symbiosis in terms of phosphorus transfer between the fungus and the plants, without any apparent changes in root colonization parameters or mycorrhizal growth response, and call for more focused research on temporal dynamics of mycorrhizal functioning under changing environmental conditions.

On the possible role of macrofungi in the biogeochemical fate of uranium in polluted forest soils.

Interactions of macrofungi with U, Th, Pb and Ag were investigated in the former ore mining district of Príbram, Czech Republic. Samples of saprotrophic (34 samples, 24 species) and ectomycorrhizal (38 samples, 26 species) macrofungi were collected from a U-polluted Norway spruce plantation and tailings and analyzed for metal content. In contrast to Ag, which was highly accumulated in fruit-bodies, concentrations of U generally did not exceed 3mg/kg which indicates a very low uptake rate and efficient exclusion of U from macrofungi. In ectomycorrhizal tips (mostly determined to species level by DNA sequencing), U contents were practically identical with those of the non-mycorrhizal fine spruce roots. These findings suggest a very limited role of macrofungi in uptake and biotransformation of U in polluted forest soils. Furthermore, accumulation of U, Th, Pb and Ag in macrofungal fruit-bodies apparently does not depend on total content and chemical fractionation of these metals in soils (tested by the BCR sequential extraction in this study).