Fungal Mats Research Articles

Characterization of the Neurospora crassa Galactosaminogalactan Biosynthetic Pathway.

The Neurospora crassa genome has a gene cluster for the synthesis of galactosaminogalactan (GAG). The gene cluster includes the following: (1) UDP-glucose-4-epimerase to convert UDP-glucose and UDP-N-acetylglucosamine to UDP-galactose and UDP-N-acetylgalactosamine (NCU05133), (2) GAG synthase for the synthesis of an acetylated GAG (NCU05132), (3) GAG deacetylase (/NCW-1/NCU05137), (4) GH135-1, a GAG hydrolase with specificity for N-acetylgalactosamine-containing GAG (NCU05135), and (5) GH114-1, a galactosaminidase with specificity for galactosamine-containing GAG (NCU05136). The deacetylase was previously shown to be a major cell wall glycoprotein and given the name of NCW-1 (non-GPI anchored cell wall protein-1). Characterization of the polysaccharides found in the growth medium from the wild type and the GAG synthase mutant demonstrates that there is a major reduction in the levels of polysaccharides containing galactosamine and N-acetylgalactosamine in the mutant growth medium, providing evidence that the synthase is responsible for the production of a GAG. The analysis also indicates that there are other galactose-containing polysaccharides produced by the fungus. Phenotypic characterization of wild-type and mutant isolates showed that deacetylated GAG from the wild type can function as an adhesin to a glass surface and provides the fungal mat with tensile strength, demonstrating that the deacetylated GAG functions as an intercellular adhesive. The acetylated GAG produced by the deacetylase mutant was found to function as an adhesive for chitin, alumina, celite (diatomaceous earth), activated charcoal, and wheat leaf particulates.

Fitness effects of a demography-dispersal trade-off in expandingSaccharomyces cerevisiaemats.

Fungi expand in space and time to form complex multicellular communities. The mechanisms by which they do so can vary dramatically and determine the life-history and dispersal traits of expanding populations. These traits influence deterministic and stochastic components of evolution, resulting in complex eco-evolutionary dynamics during colony expansion. We perform experiments on budding yeast strains genetically engineered to display rough-surface and smooth-surface phenotypes in colony-like structures called 'mats'. Previously, it was shown that the rough-surface strain has a competitive advantage over the smooth-surface strain when grown on semi-solid media. We experimentally observe the emergence and expansion of segments with a distinct smooth-surface phenotype during rough-surface mat development. We propose a trade-off between dispersal and local carrying capacity to explain the relative fitness of these two phenotypes. Using a modified stepping-stone model, we demonstrate that this trade-off gives the high-dispersing, rough-surface phenotype a competitive advantage from standing variation, but that it inhibits this phenotype's ability to invade a resident smooth-surface population via mutation. However, the trade-off improves the ability of the smooth-surface phenotype to invade in rough-surface mats, replicating the frequent emergence of smooth-surface segments in experiments. Together, these computational and experimental findings advance our understanding of the complex eco-evolutionary dynamics of fungal mat expansion.

Genome-wide identification of Sclerotinia sclerotiorum small RNAs and their endogenous targets

BackgroundSeveral phytopathogens produce small non-coding RNAs of approximately 18–30 nucleotides (nt) which post-transcriptionally regulate gene expression. Commonly called small RNAs (sRNAs), these small molecules were also reported to be present in the necrotrophic pathogen Sclerotinia sclerotiorum. S. sclerotiorum causes diseases in more than 400 plant species, including the important oilseed crop Brassica napus. sRNAs can further be classified as microRNAs (miRNAs) and short interfering RNAs (siRNAs). Certain miRNAs can activate loci that produce further sRNAs; these secondary sRNA-producing loci are called ‘phased siRNA’ (PHAS) loci and have only been described in plants. To date, very few studies have characterized sRNAs and their endogenous targets in S. sclerotiorum.ResultsWe used Illumina sequencing to characterize sRNAs from fungal mycelial mats of S. sclerotiorum spread over B. napus leaves. In total, eight sRNA libraries were prepared from in vitro, 12 h post-inoculation (HPI), and 24 HPI mycelial mat samples. Cluster analysis identified 354 abundant sRNA clusters with reads of more than 100 Reads Per Million (RPM). Differential expression analysis revealed upregulation of 34 and 57 loci at 12 and 24 HPI, respectively, in comparison to in vitro samples. Among these, 25 loci were commonly upregulated. Altogether, 343 endogenous targets were identified from the major RNAs of 25 loci. Almost 88% of these targets were annotated as repeat element genes, while the remaining targets were non-repeat element genes. Fungal degradome reads confirmed cleavage of two transposable elements by one upregulated sRNA. Altogether, 24 milRNA loci were predicted with both mature and milRNA* (star) sequences; these are both criteria associated previously with experimentally verified miRNAs. Degradome sequencing data confirmed the cleavage of 14 targets. These targets were related to repeat element genes, phosphate acetyltransferases, RNA-binding factor, and exchange factor. A PHAS gene prediction tool identified 26 possible phased interfering loci with 147 phasiRNAs from the S. sclerotiorum genome, suggesting this pathogen might produce sRNAs that function similarly to miRNAs in higher eukaryotes.ConclusionsOur results provide new insights into sRNA populations and add a new resource for the study of sRNAs in S. sclerotiorum.

Fungal Hyphosphere Microbiomes Are Distinct from Surrounding Substrates and Show Consistent Association Patterns.

Mat-forming fungi are common in forest and grassland soils across the world, where their activity contributes to important soil ecological processes. These fungi maintain dominance through aggressive and abundant hyphae that modify their internal physical and chemical environments and through these modifications select for what appears to be a suite of mycophilic bacteria. Here, the bacteria associated with the fungal mats of Leucopaxillus gentianeus and Leucopaxillus albissimus from western North America are compared to adjacent nonmat substrates. Within the mats, the bacterial richness and diversity were significantly reduced, and the community composition was significantly different. The bacterial community structure between the two fungal hosts was marginally significant and indicated a shared set of bacterial associates. The genera Burkholderia, Streptomyces, Bacillus, Paenibacillus, and Mycobacterium were significantly abundant within the fungal mats and represent core members of these hypha-rich environments. Comparison with the literature from fungal mat studies worldwide showed that these genera are common and often significantly found within fungal mats, further reinforcing the concept of a mycophilic bacterial guild. These genera are incorporated into a synthesis discussion in the context of our current understanding of the nature of fungal-bacterial interactions and the potential outcomes of these interactions in soil nutrient cycling, plant productivity, and human health. IMPORTANCE Fungi and bacteria are the most abundant and diverse organisms in soils (perhaps more so than any other habitat on earth), and together these microorganisms contribute to broad soil ecosystem processes. There is a suite of bacteria that appears consistently within the physical space called the hyphosphere, the area of influence surrounding fungal hyphae. How these bacteria are selected for, how they are maintained, and what broader ecological functions they perform are subjects of interest in this relatively new field-the cross-kingdom interactions between fungi and bacteria. Understanding their cooccurrence and their interactions can open new realms of understanding in soil ecological processes with global consequences.

Isolation, Molecular Characterization and Mycochemical Analysis of Aspergillus quadrilineatus

Secondary metabolites of microbial origin, either as pure compounds or crude extracts, provide opportunities for new drug development because of their chemical diversity. The aim of the present study is to isolate Molecular characterization and extract the bioactive compounds from the fungus of A. quadrilineatus (OL979151) in the rhizospheric soil of sunflower and mustard crop fields in Davanagere region. Dilution plate technique was used to isolate the rhizosphere fungi. The total genomic DNA was extracted from isolate of Aspergillus quadrilineatus based on Cetrimide Tetradecyl Trimethyl Ammonium Bromide (CTAB). Mass production of the fungal mat of A. quadrilineatus was established on potato dextrose broth medium. The dried methanol extract of fungus was subjected to HR-LCMS analysis; Methanol extract of A. quadrilineatus fungus shows that the compounds Amobarbital, Pirbuterol, Bambuterol, Cilazaprilat, Oxotremorine, Aminopyrine and Oryzanol as the major constituents. The compounds exhibit analgesic, anti-inflammatory, and antipyretic properties and various therapeutic uses.

A Preconditioning Paradox: Contrasting Effects of Initial Phyllosphere and Early Leaf Decomposer Microfungi on Subsequent Colonization by Leaf Decomposing Non-Unit-Restricted Basidiomycetes.

Fungal interactions during leaf decomposition can facilitate or inhibit other fungi. This experiment focused on whether preconditioning of leaf litter by microfungi that were confined to one leaf (Unit-Restricted) made leaf litter less likely to be colonized and decomposed by basidiomycetes that bind litter into mats (Non-Unit-Restricted) than non-preconditioned litter. Leaves of Manilkara bidentata in litterbags were preconditioned by incubating them for 0, 1, 2 or 3 months in flat litter/seed rain baskets 10 cm above the forest floor to avoid colonization by basidiomycete fungi. Preconditioned and non-preconditioned leaves were transferred to 5 replicate basidiomycete fungal mats of Gymnopus johnstonii for 6 weeks. Both attachment by basidiomycete fungi and percent mass loss after 6 weeks decreased significantly with increasing preconditioning time. In non-preconditioned leaves, gamma irradiation did not affect mass loss or percent white-rot despite having significantly increased numbers of basidiomycete fungal connections as compared to non-irradiated leaves. In non-preconditioned leaves, more basidiomycetes attachmented to non-irradiated than irradiated leaves suggest facilitation by phyllosphere microfungi. While basidiomycete colonization was initially facilitated by phyllosphere fungi, we inferred that degradation of resource quality led to fewer fungal attachments and less mass loss after 1–3 months of preconditioning by microfungi. The date suggest there is a 1-month time window for basidiomycete fungi to incorporate fallen leaves into their litter mats.

Disturbance reduces fungal white‐rot litter mat cover in a wet subtropical forest

AbstractFungi that bind leaf litter into mats and produce white‐rot via degradation of lignin and other aromatic compounds influence forest nutrient cycling and soil fertility. Extent of white‐rot litter mats formed by basidiomycete fungi in Puerto Rico decreased in response to disturbances—a simulated hurricane treatment executed by canopy trimming and debris addition in 2014, a drought in 2015, a treefall, and two hurricanes 10 days apart in September 2017. Percent fungal litter mat cover ranged from 0.4% after Hurricanes Irma and Maria to a high of 53% in forest with undisturbed canopy prior to the 2017 hurricanes, with means mostly between 10% and 45% of fungal litter mat cover in undisturbed forest. Drought decreased litter mat cover in both treatments, except in one control plot dominated by a drought‐resistant fungus, Marasmius crinis‐equi. Percent fungal litter mat cover sharply declined after hurricanes, a treefall, and a simulated hurricane treatment. Solar radiation was significantly inversely correlated with relative humidity (RH) and percent litter mat cover within each of the four climatic seasons. Solar radiation was also directly correlated with prior month litterfall, while RH was moderately correlated with throughfall, rain, and litter wetness. However, rainfall was inversely correlated with litter mat cover, possibly due to erosion or saturation during high rainfall events. Canopy opening reduced leaf fall and litter mat cover but these variables were not correlated except in winter. The main factor inhibiting basidiomycete fungi that bind leaf litter into mats was likely lower litter moisture associated with drought and increased solar radiation from canopy opening but secondary compounds in green litterfall may have contributed. Although higher litterfall likely increases fungal mat cover under closed canopy, changes in environmental factors apparently had a stronger inhibitory effect following canopy disturbances. Drought tolerance of some basidiomycete fungal litter mat species provided some resilience to drought.

Assessing the Fungal Simultaneous Removal Efficiency of Carbamazepine, Diclofenac and Ibuprofen in Aquatic Environment.

Unused pharmaceutical compounds (PhCs) discharged into the aquatic environment have been regarded as emerging pollutants due to potential harmful effects on humans and the environment. Microbial bioremediation is considered as a viable option for their removal from wastewater. The aim of this study was to assess the simultaneous removal of carbamazepine (CBZ), diclofenac (DCF) and ibuprofen (IBP) by previously isolated fungi (Aspergillus niger, Mucor circinelloides, Trichoderma longibrachiatum, Trametes polyzona, and Rhizopus microsporus). The tolerance to PhCs was conducted by tracking the fungal mycelium mat diameters in solid media and its dry biomass in liquid media, at the drug concentration range of 0.1 to 15 mg/L. The fungal enzymatic activities were determined for lignin peroxidase (LiP), manganese peroxidase (MnP) and laccase (Lac), respectively. The PhC removal efficiency of the fungi was assessed in aerated batch flasks and the drug concentrations and intermediate compounds formation were determined by using SPE-UPLC/MS. A tolerance over 70% was recorded for all the fungi at drug concentration of 0.1 mg/L. Manganese peroxidase was produced by all the fungi with very low amount of LiP, while all the enzymes were produced by T. polyzona. The pH of 4.3, temperature 37 ± 1.5°C and incubation time of 6 days were the optimum parameters for the fungal enzymatic activities. The best removal of CBZ (87%) was achieved by R. microsporus after 10 days. Between 78 and 100% removal of DCF was observed by all the fungi after 24 h, while 98% of IBP was removed after 2 days by M. circinelloides. Only a few intermediate compounds were identified after 3 days and disappeared after 10 days of incubation. This study demonstrated that apart from the basidiomycetes, the ascomycetes and zygomycetes are also producers of ligninolytic enzymes and have the ability to biodegrade emerging pollutants such as PhCs.

Fungal pigments on paper: Raman and quantum chemistry studies of Alternaria Sp

To advance our understanding of the molecular biochemistry of fungi which impact cultural heritage in libraries, museums and archives we investigated the diagnostic capacity of Raman spectroscopy to identify the composition of colored chromophores of fungi on paper. In this study we explored the diagnostic capacity of resonant Raman to distinguish chromophores in fungal filaments stimulated to grow on paper under high humidity with a focus on characterizing chromophores of Alternaria group species. To facilitate molecular analysis, we conducted quantum chemistry calculations of representative metabolites having optical absorption in the ultraviolet–visible spectral range. Comparing theory and experiment we show that fonsecin, erythroglaucin and aurasperone type chromophores occur in mature hyphal filaments with β-carotene dominant in yeast depositions on paper surfaces. Resonant Raman of mature filaments suggests a further contribution of carotenes longer than β-carotene to the spectral signature. Using microscopic resolution, we distinguish rich sets of Raman signatures that we assign to lignin, flavoglaucin, riboflavin, cycloleucomelon(e) and asperyellone molecular components in the spatial regions where filaments initiate from yeast depositions. In such regions, where filament microstructures stimulate development of a mature three-dimensional scaffold, the diversity of Raman resonances confirms a rich biochemistry of the developing structures. The library of computed optical and spectroscopic responses of characteristic fungal chromophores and metabolites presented here is essential for understanding the effect of fungi on a wide range of objects made from paper including books, prints, drawings, watercolors, engravings and even sculptures as well as designing next generation materials based on fungal hyphal mats.

Decolorisation of Azo Dye Congo Red (CR) by Termitomyces sp. Biomass

Aims: A study was conducted to evaluate decoloration of azo dye, Congo Red (CR) using fungal hyphal mat of beneficial bacidiomycete Termitomyces sp. TMS7 (MW694830) as bio sorbent material.
 Study design: Completely randomized block design (CRD).
 Place and duration of study: Department of Agricultural Microbiology, Tamil Nadu Agricultural University, Coimbatore, Tamil Nadu, India, between September 2019 and January 2020.
 Methodology: Isolation of white rot fungus from basidiocarb was done and screened based on their ligninolytic enzyme activity and Isolate TMS 7 was selected as best isolate and identified through ITS 1 and ITS 4 primers. Efficiency of fungal biomass to decolorize Congo red was assessed and per cent decoloration and kinetics were calculated.
 Results: Twelve fungal isolates were obtained and Isolate TMS 7 was selected as best isolate based on enzymatic activity. TMS 7 was identified as Termitomyces sp. using ITS 1 and ITS 4 primer. Ligninolytic enzymes i.e. cellulase (9.97 µ mol of glucose released/min/mg protein), and xylanase (9.55 µ mol of xylose released/min/mg protein) were quantified from the crude fungal extract of TMS 7, which was higher than standard (Termitomyces albuminosus -MTCC 1366). Decolorisation efficiency of termitomyces fungal biomass (1 g/100 ml) against different concentration of congo red dye (50-250 mg/L) was assessed. About 100 % (99.9) degradation was recorded in the minimum dye concentration of 50 mg/L within 3 days and 8 % decoloration was achieved at the highest dye concentration (250 mg/L) within 5 days.
 Conclusion: Possible mechanism of degradation is the presence of lignolytic enzyme especially cellulase, xylanase in the culture filtrate and bio sorption of degraded product by the fungal cell wall components viz., chitin, glucan other complex polymers.

Fungal mycelial mats used as textile by indigenous people of North America

ABSTRACT The indigenous people of the United States and Canada long have used forest fungi for food, tinder, medicine, paint, and many other cultural uses. New information about historical uses of fungi continues to be discovered from museums as accessions of fungi and objects made from fungi collected over the last 150+ years are examined and identified. Two textiles thought to be made from fungal mats are located in the Hood Museum of Art, Dartmouth College, and the Oakland Museum of California. Scanning electron microscopy and DNA sequencing were used to attempt to identify the fungus that produced the mats. Although DNA sequencing failed to yield a taxonomic identification, microscopy and characteristics of the mycelial mats suggest that the mats were produced by Laricifomes officinalis. This first report of fungal mats used for textile by indigenous people of North America will help to alert museum curators and conservators as well as mycological researchers to their existence and hopefully lead to more items being discovered that have been made from fungal fabric.

Fed-batch growth of Rhizopus oryzae: Eliminating ethanol formation by controlling glucose addition

Rhizopus oryzae is a prominent strain for producing fumarate, where biomass growth precedes fumarate production. The natural biofilm growth of R. oryzae as fungal mat was investigated using different glucose addition strategies in a novel fed-batch fermenter. Batch growth was compared through three fed-batch runs, each with a different glucose addition strategy. The fed-batch runs involved a constant glucose feed (CGF) of 0.075 g h−1 and controlled glucose feeds in order to keep the respiration quotient (RQ) at either 1.3 mol CO2 mol−1 O2 (RQ1.3) or 1.1 mol CO2 mol−1 O2 (RQ1.1). Ethanol overflow via the established Crabtree mechanism was completely negated for the CGF and RQ1.1 runs, while the batch and RQ1.3 runs exhibited significant ethanol formation. Biomass yield on glucose was found to be 0.476 g g−1 (RQ1.1), 0.194 g g−1 (RQ1.3), 0.125 g g−1 (CGF) and 0.144 g g−1 (batch). The results indicate a three-fold improvement in biomass yield when comparing the batch run with the RQ1.1 run. In addition, the RQ1.1 run resulted in zero detectable byproducts, unlike the batch scenario where pyruvate and fumarate were associated with ethanol formation. Clear evidence is provided that glucose overflow can be fully eliminated during R. oryzae growth, significantly affecting the biomass yield on glucose.

Diagnosis of Aeromonas sobria and Saprolegnia sp. co-infection in rainbow trout fry (Oncorhynchus mykiss)

The aim of the study was to determine a considerable level of mortalities occurred in rainbow trout fry (Oncorhynchus mykiss) cultured in a research hatchery located in Marmara Region/Turkey. Totally 18 individuals (1-8 g) were investigated by using bacteriological and mycological methods. The affected fish showed skin darkening, prolapses and fungal mats at the base of the fins. According to morphological and biochemical characteristics of the isolates, all of them were identified as Aeromonas sobria. Saprolegnia sp. was also observed in squash preparations of the skin. All isolates were determined to be sensitive against florfenicol, enrofloxacin and ciprofloxacin. Histopathologically, vacuolar degeneration, haemorrhage and hyperaemia in the liver, tubular necrosis, periglomerular edema, and multifocal melanomacrophage deposits in the kidney and depletion of white pulp in the spleen were determined. Distal type epithelial cells hyperplasia was also observed in the gills. In diseased fish, motile Aeromonad septicaemia was described and Saprolegnia sp. was also detected in fins and skin lesions but it was considered to be a secondary infection.

Sap Beetles (Coleoptera: Nitidulidae) in Oak Forests of Two Northeastern States: A Comparison of Trapping Methods and Monitoring for Phoretic Fungi.

Oak wilt is slowly expanding in the northeastern United States. Several nitidulid beetle species are known vectors of the fungus [Bretziella fagacearum (Bretz) Z. W. De Beer, Marinc., T. A. Duong, and M. J. Wingf (Microascales: Ceratocystidaceae)] that causes this disease, acquiring spores from fungal mats on infected trees and transmitting them to uninfected trees. Survey and fungal isolation from captured nitidulid beetles could be an important tool for detecting the presence of this disease in a geographic area not previously known to have oak wilt. In preparation for monitoring activities in such areas, two trapping studies were conducted in the northeastern United States: 1) trap test comparing the efficacy of wind-oriented pipe, multiple-funnel, and modified pitfall traps for nitidulids and 2) wet and dry collection cup comparison. Lures were a combination of nitidulid pheromones and fermenting liquid. Results support the use of multiple-funnel traps over the other two trap types, for both targeted species-specific surveys and community sampling. More total nitidulids, Colopterus truncatus (Randall), and Glischrochilus fasciatus (Olivier) were captured in wet collection cups compared with dry cups. Twenty-seven fungal species were isolated, none of which were B. fagacearum. Many fungi isolated from beetles were plant pathogens, indicating that in addition to the oak wilt fungus, sap beetles may contribute to the spread of other plant diseases.

Novel biomimetic chitin-glucan polysaccharide nano/microfibrous fungal-scaffolds for tissue engineering applications

Naturally occurring many biological structures have provided sources of inspiration for the fabrication of many novel nanostructures for various applications. Electrospun nano/microfibrous structures have great potential as scaffolds for cell attachment and proliferation in the field of tissue engineering. Here, for the first time, we report on the preparation of three-dimensional (3D) fungal mycelial mats with chitin-glucan polysaccharide cell walls as nano/microfibrous scaffolds for tissue engineering applications. Treatment of fungal-scaffolds (F-scaffolds) with β-mercaptoethanol (BME) improved hemocompatibility, and conferred biocompatibility with respect to the adhesion and proliferation of human keratinocytes. Field-emission scanning electron microscopy (FE-SEM) of BME-treated F-scaffolds revealed a meshwork of nano- and micro-fibrous mycelial structures with an average diameter of 2.94 ± 0.96 μm (range 0.92–5.6 μm). Tensile testing showed F-scaffolds had a mean tensile strength of 0.192 ± 0.07 MPa and a mean elongation at break of 10.74 ± 2.53%, respectively. The degradation rate of the F-scaffolds showed ~19.2 ± 1.9% weight loss in 28 days. FE-SEM of BME-treated F-scaffolds seeded with keratinocytes showed deposition of extracellular matrix (ECM) components and the formation of cell sheets in 14 days. In addition, the in vitro cytocompatibility of BME-treated F-scaffolds with keratinocytes was analyzed using resazurin-based assay, which showed a time-dependent increase in metabolic activity up to culture day 21. Overall, this novel investigation shows that filamentous fungal mats with a nano/microfibrous mycelial architecture are potentially useful for tissue engineering applications.

Further evidence for fungivory in the Lower Devonian (Lochkovian) of the Welsh Borderland, UK

The recent demonstrations that widespread mid-Palaeozoic Prototaxites and other nematophytes had fungal affinities indicate that terrestrial fungi were important elements in carbon cycling in the Early Devonian. Here, we provide evidence for their participation in the recycling of nutrients by early terrestrial invertebrates. Evidence is in the form of coprolites, both those associated with nematophytes or containing their fragmentary remains. Cylindrical coprolites consistently associated with fungal mats are placed in a new ichnospecies, Bacillafaex myceliorum. Their contents are granular to amorphous, suggestive of complete digestion of the ingested hyphae, with the inference of possession of chitinases in the digestive tracts of the consumers. A further single example comprises a cluster of cylindrical bodies attached to the lower surface of a Nematothallus fragment. Here, homogenisation was less complete, with traces of hyphae remaining. Terrestrial animal fossils have not been found at the locality, but scorpions, pseudoscorpions, Opiliones, mites, centipedes (carnivores) and millipedes, and Collembola (detritivores) have been recorded from the slightly younger Rhynie cherts. Studies of fungivory in extant arthropods have concentrated on Collembola and, to a lesser extent, mites, but their faecal pellets are much smaller than the fossil examples. Millipedes, based on body size and faeces of extant forms, are considered more realistic producers, but little is known about fungal feeding in these animals. Regardless of the affinities of the producers, the diversity in morphology, sizes, aggregations, and composition of nematophyte-containing examples suggests that fungivory was an important component of carbon cycling in early terrestrial ecosystems.

Magnetic-silica nanoshell for extraction of fungal genomic DNA from Rhizopus oryzae

The main objective of the present article describes an established genomic DNA purification technique improved in vitro by using magnetic nanoparticles as a stable phase adsorbent. Stöber technique was used to synthesis Fe3O4@SiO3 nanoshell, and described using transmission electron microscopy (TEM) and dynamic light scattering. The quality and quantity of purified DNA were verified by using agarose gel electrophoresis and some PCR strategies such as universal-primed UP-PCR, RAPD patterns. Up to 50 µg of DNA was obtained from a slight quantity of one hundred mg of fungal mycelium. Fungal DNA produced with the aid of this technique was applied as a fungal DNA template for PCR method to amplify UP-PCR, RAPD provided replicable profiles. DNA purified from fungal mats was 100 % pure enough to generally be used at high concentrations for PCR technique. The end result confirmed that Fe3O4@SiO3 nanoparticles of ∼140 nm diameter with concentration of 0.4 mg/ml produced premium quality and amount of isolated DNA produced from fungal mats. The obtained DNA was free from any polluting proteins, polysaccharides and coloured dyes. Magnetic nanoparticles (MNP)-mediated genomic DNA extraction is pretty simple (loose from filtration and centrifugation), speedy (30 min), and ecofriendly (not contain any toxic chemical compounds).

ANTIBACTERIAL PROPERTY AND MOLECULAR DOCKING STUDIES OF METHANOLIC FUNGAL PHYTOCHEMICALS OF CLAVICEPS PURPUREA INFECTED TO BAJRA (PENNISETUM GLAUCUM) CROP

Objective: The objective of this study is to study the antibacterial activity, minimum inhibitory concentration, preliminary phytochemical studies, molecular docking studies, and high-resolution liquid chromatography mass spectrometer (HR-LCMS) analysis of the methanol fungal extract of Claviceps purpurea. 
 Methods: Mass production of the fungal mat of C. purpurea was established on potato dextrose broth medium. The dried methanol extract of fungus was subjected to HR-LCMS analysis; antibacterial screening of the extract was carried out against pathogenic bacteria. Molecular docking study of HR-LCMS identified compounds was performed by docking with bacterial enzyme DNA gyrase.
 Results: HR-LCMS analysis of methanol extract of C. purpurea fungus shows that the compounds arecoline, benperidol, felbamate, solanidine, and triparanol as the major constituents. The antibacterial screening of methanol extract of fungus against bacterial pathogens showed a significant bactericidal activity against the strains Pseudomonas aeruginosa (14.40±0.04 mm), Xanthomonas campestris (13.60±0.06 mm), Escherichia coli (13.40±0.06 mm), Salmonella Typhi (11.50±0.05 mm), Staphylococcus aureus (11.37±0.04 mm.), and as matched to the standard drug ciprofloxacin. The molecular docking of triparanol against the bacterial enzyme DNA gyrase showed good binding affinity of −6.2 kcal/mol, good drug likeness (7.3767), 2 hydrogen bonds, and hydrophobic interaction with 8 amino acid residues so that triparanol shows good inhibitor as compared to other 4 compounds.
 Conclusion: Methanol fungal extract of C. purpurea showed significant antibacterial activity against pathogenic bacteria. Triparanol is the major compound present in C. purpurea fungus which showed good inhibitory activity against bacterial enzyme DNA gyrase. The methanol extract of C. purpurea fungus displayed a good antibacterial drug.

Biosynthesis of Silver Nanoparticles from Endophytic Fungi, and its Cytotoxic Activity

Nanoparticle research is currently an area of intense scientific interest due to a wide variety of potential applications in biomedical, optical, and electronic fields. The biological synthesis of nanoparticles includes the synthesis of nanoparticles using microbes which have the capability to absorb the nanoparticles that exist in various forms. In the biosynthesis of metal nanoparticles by a fungus Botryodiplodia theobromae, the fungus mycelium is exposed to the metal salt solution. The change in color of the medium indicates the synthesis of silver nanoparticles. The synthesized AgNPs were structurally characterized by UV-visible spectroscopy, scanning electron microscope (SEM), and Fourier-transform infrared spectroscopy (FTIR) analysis. The size of the silver nanoparticles was ranging from 62.77 –103 nm in the fungal mat and 66.75–111.23 nm in the cell filtrate respectively with the existence of free OH and NH groups, aromatic CH stretch, C–C stretch, unsaturation in the molecule, and C–Cl stretch. The cytotoxic efficiency of the AgNPs on human MCF7 breast cancer cell lines and A549 human lung cancer cell lines was appraised by cell viability assay. The synthesized AgNPs inhibited the propagation of cells at an IC50 concentration of 100 μg/mL. Thus, this study reveals that the green synthesis is an eco-friendly method for production of AgNPs from endophytic fungi which provided a powerful anti-proliferative action on MCF-7 and A549 cell lines, suggesting them as a novel chemotherapeutic agent against human breast and lung cancers.

Ecofriendly nanomaterials for controlling gray mold of table grapes and maintaining postharvest quality

Biodegradable antifungal nanomaterials are a recent novel measure against plant pathogens. In the present investigation, the synthesis and characterization of some ecofriendly nanomaterials, including silica, chitosan, and copper nanoparticles (NPs) and their combination, were carried out. Their fungicidal activity was studied in vitro and in vivo against Botrytis cinerea, the causal agent of gray mold on table grapes. In addition, the effect of those nanomaterials on physical and chemical properties of grape (TSS, TA, TSS/TA ratio and berries colour) were evaluated. Scanning electron microscopy (SEM) and analysis of DNA-binding profile were used to better understand their mechanism of action. SEM showed that chitosan and silica NPs caused inhibition of hyphal growth and/or alteration of hyphal morphology such as cell wall disruption, withering, and excessive septation. NPs interacted with DNA isolated from fungal mats: the highest concentration of chitosan and silica NPs affected DNA integrity and led to a significant degradation. A single application of chitosan or silica NPs at veraison stage was able to reduce gray mold of table grapes. Although further large scale trials are needed, the promising results of this research suggest nanomaterials compounds, i.e. silica and chitosan NPs, as effective antifungal agents for the control of gray mold of table grapes.