Potential of Fungal Endophytes Isolated from Pasture Species in Spanish Dehesas to Produce Enzymes under Salt Conditions.

Endophytic fungi have been found to produce a wide range of extracellular enzymes, which are increasingly in demand for their industrial applications. Different by-products from the agrifood industry could be used as fungal growth substrates for the massive production of these enzymes, specifically as a way to revalorize them. However, such by-products often present unfavorable conditions for the microorganism's growth, such as high salt concentrations. Therefore, the objective of the present study was to evaluate the potential of eleven endophytic fungi-which were isolated from plants growing in a harsh environment, specifically, from the Spanish dehesas-for the purposes of the in vitro production of six enzymes (i.e., amylase, lipase, protease, cellulase, pectinase and laccase) under both standard and salt-amended conditions. Under standard conditions, the studied endophytes produced between two and four of the six enzymes evaluated. In most of the producer fungal species, this enzymatic activity was relatively maintained when NaCl was added to the medium. Among the isolates evaluated, Sarocladium terricola (E025), Acremonium implicatum (E178), Microdiplodia hawaiiensis (E198), and an unidentified species (E586) were the most suitable candidates for the massive production of enzymes by using growth substrates with saline properties (such as those found in the many by-products from the agrifood industry). This study should be considered an initial approach by which to further study the identification of these compounds as well as to develop the optimization of their production by directly using those residues.

Protective Effects of Filtrates and Extracts from Fungal Endophytes on Phytophthora cinnamomi in Lupinus luteus.

Fungal endophytes have been found to protect their hosts against multiple fungal pathogens. Frequently, the secondary metabolites produced by the endophyte are responsible for antifungal activity. To develop new bio-products that are more environmentally friendly than synthetic pesticides against Phytophthora cinnamomi, a serious pathogen of many plant species, the antifungal activity of filtrates or extracts from four endophytes was evaluated in different in vitro tests and in plants of Lupinus luteus. In the dual culture assays, the filtrate of one of the endophytes (Drechslera biseptata) completely inhibited the mycelial growth of the pathogen. Moreover, it showed a very low minimal inhibitory concentration (MIC). Epicoccum nigrum, an endophyte that also showed high inhibitory activity and a low MIC against P. cinnamomi in those two experiments, provided a clear growth promotion effect when the extracts were applied to L. luteus seedlings. The extract of Fusarium avenaceum also manifested such a promotion effect and was the most effective in reducing the disease severity caused by the pathogen in lupine plants (73% reduction). Results demonstrated the inhibitory activity of the filtrates or extracts of these endophytes against P. cinnamomi. A better insight into the mechanisms involved may be gained by isolating and identifying the metabolites conferring this inhibitory effect against this oomycete pathogen.

Metabolites Produced by Fungi against Fungal Phytopathogens: Review, Implementation and Perspectives.

Many fungi, especially endophytes, have been found to produce multiple benefits in their plant hosts, with many of these benefits associated with the protection of plants against fungal diseases. This fact could be used in the development of new bio-products that could gradually reduce the need for chemical fungicides, which have been associated with multiple health and environmental problems. However, the utilization of the living organism may present several issues, such as an inconsistency in the results obtained and more complicated management and application, as fungal species are highly influenced by environmental conditions, the type of relationship with the plant host and interaction with other microorganisms. These issues could be addressed by using the bioactive compounds produced by the fungus, in cases where they were responsible for positive effects, instead of the living organism. Multiple bioactive compounds produced by fungal species, especially endophytes, with antifungal properties have been previously reported in the literature. However, despite the large amount of these metabolites and their potential, extensive in-field application on a large scale has not yet been implemented. In the present review, the main aspects explaining this limited implementation are analyzed, and the present and future perspectives for its development are discussed.

Different dry hopping and fermentation methods: influence on beer nutritional quality.

BACKGROUND: Nowadays, the craft beer market is booming and the consumer trend for trying something new is increasing. Here, nine different treatments of a craft beer were realized in a pilot plant, studying fermentation and dry-hopping types. Quality parameters of the beer such as polyphenols, antioxidants, bitterness, colour and alcohol were analysed. In addition, an electronic nose was used to distinguish beer types. RESULTS: Results showed that dry hopping in maturation with warm temperature increased the bitterness from 33 to 40 IBUs. The treatment using two yeasts and two fermentation temperatures resulted in the highest antioxidant capacity of the beer (around 92%). Antioxidant activity was increased by late dry hopping using ale yeasts for fermenting. Principal component analysis performed with electronic nose data explained up to 97% of the total variability of the compounds in the study. CONCLUSIONS: Combined use of ale and lager yeasts seems to increase the antioxidant capacity and total polyphenol content of beer. Antioxidant activity is increased by late dry hopping. An electronic nose is a suitable device for discriminating the volatile profile complexity in beer. © 2020 Society of Chemical Industry.

Hyfraxins A and B, cytotoxic ergostane-type steroid and lanostane triterpenoid glycosides from the invasive ash dieback ascomycete Hymenoscyphus fraxineus.

A virulent culture of Hymenoscyphus fraxineus, the causal agent of ash dieback, was investigated for its production of secondary metabolites in a 70 L batch fermentation. Chemical analysis of the mycelial extract by means of flash chromatography and preparative HPLC led to the isolation of a new ergostane-type steroid (1) and a new related lanostane triterpenoid (2), both revealing the same glycosylation pattern. While their planar structures were elucidated by HR-ESIMS and NMR data, relative stereochemistry was assigned by ROESY correlations in conjunction with H,H and C,H coupling constants. Absolute configuration was determined based on ROESY correlations between the aglycons and the sugar moieties, which were identified in both cases as d-mannose by GC/MS analysis of the trimethylsilylated derivatives. The isolated compounds, for which we propose the trivial names hyfraxins A (1) and B (2), were found to be cytotoxic against the mouse fibroblast cell line L929 and exhibited moderate to weak activity against Gram-positive bacteria.

Effect of Fungal Endophytes on Biomass Yield, Nutritive Value and Accumulation of Minerals in Ornithopus compressus.

Yellow serradella (Ornithopus compressus), a valuable pasture species in Mediterranean areas, presents a high diversity of endophytic mycoflora. In the present work, the hypothesis of a significant effect of fungal endophytic species on the parameters of forage production, nutritive value and mineral status of herbage was tested. O. compressus plants were inoculated with each of seven endophytes (four in 2012/2013 and three in 2013/2014). After inoculation, two experiments (under greenhouse and field conditions) were established. Results evidenced a certain influence of several endophytes on herbage yield, nutritive value and mineral status of O. compressus forage. Byssochlamys spectabilis increased herbage biomass yield by around 42% in the field experiment. Stemphylium sp. improved the nutritive value of forage either by increasing crude protein, digestibility and the concentration of essential minerals (such as B, Mo, P or S) or by reducing the concentration of toxic elements such as Al or Pb. In conclusion, the results presented here provide evidence that plant inoculation with endophytes could be a suitable strategy to increase forage yield and its nutritive value or to deal with potential nutrient deficiencies or potential mineral toxicity in forage.

Selenium speciation in malt, wort, and beer made from selenium-biofortified two-rowed barley grain.

Selenium (Se) biofortification of barley is a suitable strategy to increase the Se concentration in grain. In the present paper, the suitability of this Se-biofortified grain for making Se-enriched beer is analyzed. The aim of the present study was to evaluate the effect of different Se fertilizer doses (0, 10, and 20 g of Se ha(-1)) and forms (sodium selenate or sodium selenite) on the Se loss during the malting and brewing processes and Se speciation in grain, malt, wort, and beer. Samples were analyzed using inductively coupled plasma-mass spectrometry (ICP-MS) and high-performance liquid chromatography (HPLC)-ICP-MS for total Se and speciation. Mashing-lautering was the process with the greatest Se loss (83.8%). After malting and brewing, only 7.3% of the initial Se was retained in beer, mainly in selenite form. Even so, the fertilizer application of sodium selenate at 20 g ha(-1) increased the total Se concentration almost 6-fold in the final beer in comparison to the use of grain derived from unfertilized barley. The present paper provides evidence that the use of Se-biofortified barley grain as a raw material to produce Se-enriched beer is possible, and the results are comparable to other methods in terms of efficiency.

Selenium accumulation and speciation in biofortified chickpea (Cicer arietinum L.) under Mediterranean conditions.

BACKGROUND: Millions of people have Se-deficient diets and Se-biofortified crops could prevent such deficiency. The aim of the present study was to evaluate the potential of chickpea for use in Se fertilization programs in order to increase available Se. Two foliar Se fertilizers (sodium selenate and sodium selenite) at four rates (0, 10, 20, 40 g ha(-1)) were tested in the 2010/2011 and 2011/2012 growing seasons in a field experiment conducted under semiarid Mediterranean conditions. RESULTS: Sodium selenate was much more effectively taken by plants than sodium selenite, and there was a strong and linear relationship between total Se content and Se rate for both. For each gram of Se fertilizer, applied either as sodium selenate or sodium selenite, the increases of total Se concentration in grain were 126 and 87, and 25 and 19 µg Se kg(-1) dry weight, in 2010/2011 and 2011/2012, respectively. Se was found to be incorporated into chickpea grains mainly (>70%) as selenomethionine. CONCLUSION: Se-enriched chickpeas would be a good candidate for inclusion in biofortification programs under semiarid Mediterranean conditions and for promotion as a 'functional food'.

Evaluation of the potential of peas (Pisum sativum L.) to be used in selenium biofortification programs under Mediterranean conditions.

Selenium (Se), which has antioxidant, anticancer, and antiviral properties, is an essential micronutrient for humans and animals. This micronutrient is found in high quantity in legumes. Peas have an ever-increasing importance in Spain, and to increase their nutritional value, two foliar Se fertilizers: sodium selenate and sodium selenite, at five different rates: 0, 10, 20, 40, 80 g ha(-1), were studied during the 2010/2011 crop season on semiarid Mediterranean conditions. Sodium selenate was much more effectively taken up by plants compared to sodium selenite. There was a strong linear relationship between the total Se content and Se rate in both sodium selenate and selenite. For each gram of Se fertilization as either sodium selenate or sodium selenite, the increase of total Se concentration in the grain was 148 and 19 µg Se kg(-1) dry weight, respectively. Ingestion of 100 g of peas previously fertilized with 10 g of sodium selenate per hectare would result in an intake of 179 µg of Se. This is almost 90 % of the daily recommended dose needed to reduce the chance of some cancers and about 179 % of the minimum concentration required to prevent Se deficiency diseases in animals. The pea has shown to have a strong ability to uptake and accumulate Se under Mediterranean conditions; therefore, this would make it a very strong candidate for inclusion in biofortification programs aiming to increase Se in the food chain.