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Desert truffle crop is a pioneer in southeastern Spain, a region where native edible hypogeous fungi are adapted to the semiarid areas with low annual rainfall. Terfezia claveryi Chatin was the first species of desert truffle to be cultivated, and has been increasing in recent years as an alternative rainfed crop in the Iberian Peninsula. However, its behaviour in the field has yet not been investigated. For this purpose, specific primers were designed for the soil DNA quantification of both T. claveryi and Terfezia crassiverrucosa and a real-time qPCR protocol was developed, using the ITS rDNA region as a target. Moreover, a young desert truffle orchard was sampled for environmental validation. The results showed the highest efficiency for the TerclaF3/TerclaR1 primers pair, 89%, and the minimal fungal biomass that could be reliable detected was set at 4.23 µg mycelium/g soil. The spatial distribution of fungal biomass was heterogeneous, and there was not a direct relationship between the quantity of winter soil mycelium and the location/productivity of desert truffles. This protocol could be applied to tracking these species in soil and understand their mycelial dynamics in plantations and wild areas. [ABSTRACT FROM AUTHOR]

Desert truffle is becoming a new crop in semiarid areas. Climatic parameters and the presence of microorganisms influence the host plant physiology and alter desert truffle production. Desert truffle plants present a typical summer deciduous plant phenology divided into four stages: summer dormancy, autumn bud break, winter photosynthetic activity, and spring fruiting. We hypothesize that the bacterial community associated with desert truffle plants will show a seasonal trend linked to their plant growth–promoting rhizobacteria (PGPR) traits. This information will provide us with a better understanding about its potential role in this symbiosis and possible management implementations. Bacteria were isolated from root-adhering soil at the four described seasons. A total of 417 isolated bacteria were phenotypically and biochemically characterized and gathered by molecular analysis into 68 operational taxonomic units (OTUs). They were further characterized for PGPR traits such as indole acetic acid production, siderophore production, calcium phosphate solubilization, and ACCD (1-amino-cyclopropane-1-carboxilatedeaminase) activity. These PGPR traits were used to infer functional PGPR diversity and cultivable bacterial OTU composition at different phenological moments. The different seasons induced shifts in the OTU composition linked to their PGPR traits. Summer was the phenological stage with the lowest microbial diversity and PGPR functions, whereas spring was the most active one. Among the PGPR traits analyzed, P-solubilizing rhizobacteria were harbored in the mycorrhizosphere during desert truffle fruiting in spring. [ABSTRACT FROM AUTHOR]

Summary: The desert truffle Terfezia claveryi is one of the few mycorrhizal fungi currently in cultivation in semiarid and arid areas. Agroclimatic parameters seem to affect its annual yield, but there is no information on the influence of biotic factors. In this study, fungal diversity was analysed by high‐throughput sequencing of the ITS2 rDNA region from soil and root samples to compare productive and non‐productive mycorrhizal plants in a 4‐years old plantation (Murcia, Spain). The fungal metaprofile was dominated by Ascomycota phylum. Desert truffle productivity was driven by different patterns of fungal species composition in soil (species replacement) and root (species richness differences). Moreover, positive associations for ectomycorrhizal and negative for arbuscular mycorrhizal guilds were found in productive roots, and positive associations for fungal parasite‐plant pathogen guild in non‐productive ones. Soil samples were dominated by pathotroph and saprotroph trophic modes, showing positive associations for Aureobasidium pullulans and Alternaria sp. in productive areas, and positive associations for Fusarium sp. and Mortierella sp. were found in non‐productive soils. Finally, some significant OTUs were identified and associated to ascocarp producing patches, which could serve as predictive and location markers of desert truffle production. [ABSTRACT FROM AUTHOR]

Summary: Desert truffles are edible hypogeous fungi forming ectendomycorrhizal symbiosis with plants of Cistaceae family. Knowledge about the reproductive modes of these fungi and the molecular mechanisms driving the ectendomycorrhizal interaction is lacking.Genomes of the highly appreciated edible desert truffles Terfezia claveryi Chatin and Tirmania nivea Trappe have been sequenced and compared with other Pezizomycetes. Transcriptomes of T. claveryi × Helianthemum almeriense mycorrhiza from well‐watered and drought‐stressed plants, when intracellular colonizations is promoted, were investigated.We have identified the fungal genes related to sexual reproduction in desert truffles and desert‐truffles‐specific genomic and secretomic features with respect to other Pezizomycetes, such as the expansion of a large set of gene families with unknown Pfam domains and a number of species or desert‐truffle‐specific small secreted proteins differentially regulated in symbiosis. A core set of plant genes, including carbohydrate, lipid‐metabolism, and defence‐related genes, differentially expressed in mycorrhiza under both conditions was found.Our results highlight the singularities of desert truffles with respect to other mycorrhizal fungi while providing a first glimpse on plant and fungal determinants involved in ecto to endo symbiotic switch that occurs in desert truffle under dry conditions. [ABSTRACT FROM AUTHOR]

Predicted increases in atmospheric concentration of carbon dioxide (CO2) coupled with increased temperatures and drought are expected to strongly influence the development of most of the plant species in the world, especially in areas with high risk of desertification like the Mediterranean basin. Helianthemum almeriense is an ecologically important Mediterranean shrub with an added interest because it serves as the host for the Terfezia claveryi mycorrhizal fungus, which is a desert truffle with increasingly commercial interest. Although both plant and fungi are known to be well adapted to dry conditions, it is still uncertain how the increase in atmospheric CO2 will influence them. In this article we have addressed the physiological responses of H. almeriense × T. claveryi mycorrhizal plants to increases in atmospheric CO2 coupled with drought and high vapor pressure deficit. This work reports one of the few estimations of mesophyll conductance in a drought deciduous Mediterranean shrub and evaluates its role in photosynthesis limitation. High atmospheric CO2 concentrations help desert truffle mycorrhizal plants to cope with the adverse effects of progressive drought during Mediterranean springs by improving carbon net assimilation, intrinsic water use efficiency and dispersal of the species through increased flowering events. [ABSTRACT FROM AUTHOR]

The cultivation of desert truffle Terfezia claveryi using Helianthemum almeriense as a host plant has recently become a solid alternative crop in the Mediterranean region due to its adaptation to arid and semiarid ecosystems, which are expected to increase during the following years because of climate change. However, management models are still being developed in order to improve and stabilize the production, which varies greatly from one year to another. According to gatherers and farmers, one of the key factors for desert truffle production is the plant phenology in spring, which, in turn, depends on environmental conditions. In this manuscript, we have characterized the physiological, morphological, and molecular responses of the mycorrhizal plants in spring, coinciding with the fructification period of the plant and fungal species. Thanks to this characterization, a sigmoidal relationship between stomatal conductance and vapor pressure deficit (VPD) was found, which can be used as a marker of plant phenological switch. In order to confirm that this phenology status is related to desert truffle fructification, this marker has been successfully correlated to total truffle production. The results of this manuscript suppose a big step forward that will help to develop management models for the desert truffle crop. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Terfezia claveryi Chatin was the first desert truffle species to be cultivated, the mycorrhizal plants being successfully produced by using both desert truffle spores and mycelia. However, it is more advisable to use mycelium than spores whenever possible and profitable. Given the low yields of mycelia obtained using traditional culture methods of this truffle, the medium composition was modified in an attempt to determine its nutritional requirements. For this, an assay involving response surface methodology was performed using Box-Behnken design to find the optimal parameters for the high production of mycelial biomass. The best results were obtained with glucose as carbon source, buffering the pH at 5 during culture, adding a pool of vitamins, and adjusting the optimal concentrations of carbon and nitrogen sources of the MMN medium. Biomass production increased from 0.3 to 3 g L−1 dry weight and productivity increased from 10.7 to 95.8 mg L−1 day−1 dry weight. The produced mycelium was able to colonize Helianthemum roots efficiently, providing more than 50% ectomycorrhizal colonization. [ABSTRACT FROM AUTHOR]

Sixty-four native bacterial colonies were isolated from mycorrhizal roots of Helianthemum almeriense colonized by Terfezia claveryi, mycorrhizosphere soil, and peridium of T. claveryi to evaluate their effect on mycorrhizal plant production. Based on the phylogenetic analysis of the 16S rDNA partial sequence, 45 different strains from 17 genera were gathered. The largest genera were Pseudomonas (40.8 % of the isolated strains), Bacillus (12.2 % of isolated strains), and Varivorax (8.2 % of isolated strains). All the bacteria were characterized phenotypically and by their plant growth-promoting rhizobacteria (PGPR) traits (auxin and siderophore production, phosphate solubilization, and ACC deaminase activity). Only bacterial combinations with several PGPR traits or Pseudomonas sp. strain 5, which presents three different PGPR traits, had a positive effect on plant survival and growth. Particularly relevant were the bacterial treatments involving auxin release, which significantly increased the root-shoot ratio and mycorrhizal colonization. Moreover, Pseudomonas mandelii strain 29 was able to considerably increase mycorrhizal colonization but not plant growth, and could be considered as mycorrhiza-helper bacteria. Therefore, the mycorrhizal roots, mycorrhizosphere soil, and peridium of desert truffles are environments enriched in bacteria which may be used to increase the survival and mycorrhization in the desert truffle plant production system at a semi-industrial scale. [ABSTRACT FROM AUTHOR]

Sporocarp formation is part of the reproductive stage in the life cycle of many mycorrhizal macrofungi. Sporocarp formation is accompanied by a transcriptomic switch and profound changes in regulation of the gene families that play crucial roles in the sporocarp initiation and maturation. Since sporocarp growth requires efficient water delivery, in the present study, we investigated changes in transcript abundance of six fungal aquaporin genes that could be cloned from the ectomycorrhizal fungus Laccaria bicolor strain UAMH8232, during the initiation and development of its basidiocarp. Aquaporins are intrinsic membrane proteins facilitating the transmembrane transport of water and other small neutral molecules. In controlled-environment experiments, we induced basidiocarp formation in L. bicolor, which formed ectomycorrhizal associations with white spruce ( Picea glauca) seedlings. We profiled transcript abundance corresponding to six fungal aquaporin genes at six different developmental stages of basidiocarp growth and development. We also compared physiological parameters of non-inoculated to mycorrhizal seedlings with and without the presence of basidiocarps. Two L. bicolor aquaporins- JQ585592, a functional channel for CO, NO and HO, and JQ585595, a functional water channel-showed the greatest degree of upregulation during development of the basidiocarp. Our findings point to the importance of aquaporin-mediated transmembrane water and CO transport during distinct stages of basidiocarp development. [ABSTRACT FROM AUTHOR]

The development of ectomycorrhizal associations is crucial for growth of many forest trees. However, the signals that are exchanged between the fungus and the host plant during the colonization process are still poorly understood. In this study, we have identified the relationship between expression patterns of L accaria bicolor aquaporin LbAQP1 and the development of ectomycorrhizal structures in trembling aspen ( P opulus tremuloides) seedlings. The peak expression of LbAQP1 was 700-fold higher in the hyphae within the root than in the free-living mycelium after 24 h of direct interaction with the roots. Moreover, in LbAQP1 knock-down strains, a non-mycorrhizal phenotype was developed without the Hartig net and the expression of the mycorrhizal effector protein MiSSP7 quickly declined after an initial peak on day 5 of interaction of the fungal hyphae with the roots. The increase in the expression of LbAQP1 required a direct contact of the fungus with the root and it modulated the expression of MiSSP7. We have also determined that LbAQP1 facilitated NO, H2O2 and CO2 transport when heterologously expressed in yeast. The report demonstrates that the L . bicolor aquaporin LbAQP1 acts as a molecular signalling channel, which is fundamental for the development of Hartig net in root tips of P . tremuloides. [ABSTRACT FROM AUTHOR]

The influence of inorganic and organic phosphorus (P) and the absence of P in the culture medium on the type of mycorrhizal colonization formed (ecto-, ectendo-, or endomycorrhiza) during Helianthemum almeriense x Terfezia claveryi symbiosis in in vitro conditions was analyzed. This is the first time that the relative proportions of the different mycorrhizal types in mycorrhizal roots of H. almeriense have been quantified and statistically analyzed. The relative proportions of the mycorrhizal types depended on the P source in the medium, suggesting that it is the organic P form that induces the formation of intracellular colonization. The above association should be considered as a continuum between intra- and intercellular colonizations, the most appropriate term for defining it being ectendomycorrhiza. The influence of the endogenous concentration of P on plant growth was also analyzed. P translocation was observed from shoot to roots, especially in mycorrhizal plants because mycorrhizal roots showed higher growth than non-mycorrhizal roots and/or because of an extra P demand from mycelium inside the roots. Soluble and cell wall acid phosphatases activities from H. almeriense roots were kinetically characterized at optimum pH (5.0), using p-nitrophenyl phosphate as substrate, with K values of 3.4 and 1.8 mM, respectively. Moreover, the plant acid phosphatase and fungal alkaline phosphatases activities were histochemically localised in mycorrhizal H. almeriense roots by fluorescence with enzyme-labelled fluorescence substrate. [ABSTRACT FROM AUTHOR]

The ability of two species of desert truffle, Terfezia claveryi strain TcS2 and Picoa lefebvrei strain OL2, to tolerate water stress in pure culture has been investigated. Both T. claveryi and P. lefebvrei strains exhibited a mycelium growth pattern characteristic of drought tolerant species. However, they were only tolerant to moderate water stress, below −1.07 MPa, with the P. lefebvrei isolate being slightly more drought tolerant than the T. claveryi isolate. The increased alkaline phosphatase (ALP) activity observed in both fungi at moderate water stress with respect to the control indicated the functional adaptation of these mycelia to these drought conditions. ALP activity can be used as an indicator of the metabolic activity of these fungi. Slight water stress (−0.45 MPa) could improve mycelial inoculum production of these desert truffles. Moreover, P. lefebvrei could be a good candidate for further desert truffle mycorrhizal plant cultivation programmes in semiarid Mediterranean areas. [ABSTRACT FROM AUTHOR]

Physiological parameters of mycorrhizal symbiosis by Helianthemum almeriense and Terfezia claveryi in orchards were characterized under water deficit conditions. Our orchard included 40 mycorrhizal and 40 nonmycorrhizal plants. Only mycorrhizal plants survived at the beginning of the experimental period, indicating dependency on fungal symbionts in roots for survival. Drought stress significantly affected the mycorrhizal colonization percentage which was 70% in nonirrigated mycorrhizal and 48% in irrigated mycorrhizal plants. No significant differences in plant growth were observed between nonirrigated and irrigated mycorrhizal plants before and after drought stress. Stomatal conductance was more sensitive to water stress than shoot water potential. It decreased more than two-fold under drought-stress compared to control mycorrhizal plants under irrigation/light saturating conditions, indicating important stomatal closure with water deficit. Plants' water use efficiency improved with drought with stomatal conductance values below 0.3 mol m s. The ability to maintain open stomata and photosynthesis under drought increased carbon supply for growth, and ascocarp fruiting which requires current photosynthates. Basically, H. almeriense shows a conservative water use strategy based mainly on avoiding drought stress by reducing stomatal conductance as soil water potential decreases and atmospheric conditions dry. The results show that mycorrhizal H. almeriense plants maintain good physiological parameters with low soil matric potentials, thus making them an alternative agricultural crop in arid/semi-arid areas. [ABSTRACT FROM AUTHOR]

In the present paper, we confirmed that alkaline phosphatase (ALP) is the main phosphatase present in ascocarps of the edible mycorrhizal fungus Terfezia claveryi. The enzyme was partially purified by precipitation with polyethylene glycol. The purification achieved from a crude extract was fivefold, with 53% of the activity recovered, and acid phosphatase, most of the lipids and phenolic compounds were eliminated. Alkaline phosphatase was kinetically characterised at pH 10.0, the optimum for this enzyme, using p-nitrophenyl phosphate as substrate. The Vmax and Km values were 0.3 μmol·min−1·mg−1 protein and 9.0 mm, respectively. Orthovanadate was a competitive inhibitor of ALP, with a Ki of 42.5 μm. The enzyme was histochemically localised in the peridium, the hypothecium and in the ascogenic hyphae of the gleba using both colour and fluorescent reactions. The results presented suggest that the ascocarp of T. claveryi, at some stages of its development, may become nutritionally autonomous and independent of the host plant. [ABSTRACT FROM AUTHOR]

Desert truffles are edible hypogeous (forming fruit bodies below ground) fungi that grow in semi-arid and arid areas. They are highly valued for both their culinary and medicinal properties in the Mediterranean basin, the Middle East and the Gulf areas. Desert truffles form mycorrhizae mostly with plants belonging to the Cistaceae family, mainly with Helianthemum species. These truffles are still, usually, collected from the wild, but loss of habitats due to urbanization, desertification, intensive agriculture and global warming, along with an urgent need to develop new crops adapted to arid conditions, are currently hastening efforts towards their domestication. Here, we sum up the successful research leading to cultivation of this crop, based on plots that were established in sandy to silt soils with high pH values and low mineral contents. We report suitable methods for production of mycorrhized seedlings and preferred planting methods. We found that under natural conditions yields are affected by water availability, so irrigation regimes to ensure good yields were sought. Although good yields were indeed obtained in some years, fluctuations in yields over the years were significant; the reasons for this are not entirely clear and are currently under study. This crop is particularly well suited to relatively marginal conditions but prospects for establishment of desert truffles as a niche crop for arid and semi-arid areas depend on further improvements in yields. [ABSTRACT FROM AUTHOR]

Desert truffles have become an alternative agricultural crop in semiarid areas of the Iberian Peninsula due to their much appreciated edible value and their low water requirements for cultivation. Although most studies related to desert truffle production point to the sole importance of precipitation, this work is the first systematic study carried out to characterize whether other important agroclimatic parameters, for example reference evapotranspiration, soil water potential, relative air humidity %, aridity index or air vapour pressure deficit, may have an impact on a desert truffle production in an orchard with mycorrhizal plants of Helianthemum almeriense × Terfezia claveryi for 15 years from the plantation. The results show for the first time that T. claveryi production has two key periods, during its annual cycle: autumn (September to October) and spring (end of March). The aridity index and soil water potential seem to be the most manageable parameters in the field and can be easily controlled by applying irrigation during the abovementioned periods. Agroclimatic parameters can influence the final crop a long time before the desert truffle fruiting season contrary to what happens with other edible mycorrhizal mushrooms. Four different models to manage desert truffle plantations are proposed based on these agroclimatic parameters in order to optimize and stabilize carpophore fructifications over the years. [ABSTRACT FROM AUTHOR]

Aquaporins are membrane integral proteins responsible for the transmembrane transport of water and other small neutral molecules. Despite their well-acknowledged importance in water transport, their significance in gas transport processes remains unclear. Growing evidence points to the involvement of plant aquaporins in CO2 delivery for photosynthesis. The role of these channel proteins in the transport of O2 and other gases may also be more important than previously envisioned. In this study, we examined O2 permeability of various human, plant, and fungal aquaporins by co-expressing heterologous aquaporin and myoglobin in yeast. Two of the most promising O2-transporters (Homo sapiens AQP1 and Nicotiana tabacum PIP1;3) were confirmed to facilitate O2 transport in the spectrophotometric assay using yeast protoplasts. The over-expression of NtPIP1;3 in yeasts significantly increased their O2 uptake rates in suspension culture. In N. tabacum roots subjected to hypoxic hydroponic conditions, the transcript levels of the O2-transporting aquaporin NtPIP1;3 significantly increased after the seven-day hypoxia treatment, which was accompanied by the increase of ATP levels in the apical root segments. Our results suggest that the functional significance of aquaporin-mediated O2 transport and the possibility of controlling the rate of transmembrane O2 transport should be further explored. [ABSTRACT FROM AUTHOR]