Upon collection of the ascocarps, several analyses were performed. First, material for genetic identification was collected directly upon sterilization of the tissue surface. Second, in those individuals with an intact fruiting body (16 individuals), fungal isolation was attempted either by germinating spores or by direct culturing pieces of hymenia. Isolation was performed on potato dextrose agar (PDA, Potato infusion powder, Sigma-Aldrich, 4 g/L + D(+) - glucose monohydrate, Roth, 20 g/L + Agar-Agar, Merck, 15 g/L). Pure isolates were obtained by successive plating in the same medium. In addition, two Chinese cultivars (NEU142 and NEU143) and one morel specimen from our collection (M84) were grown in PDA to be included in the analysis. Sclerotia were obtained by culturing on malt agar (Malt extract, Fluka, 12 g/L + Agar-Agar, Merck, 15 g/L; MA). All media were autoclaved at 121°C before pouring into Petri dishes.
Agar agar
Manufactured by Merck Group
Sourced in Germany, United States
Agar-agar is a natural, gelatinous substance extracted from red algae. It is commonly used as a gelling agent, stabilizer, and thickener in various laboratory applications. Agar-agar is heat-resistant, non-toxic, and capable of forming a reversible gel, making it a versatile and widely used product in the field of laboratory equipment and supplies.
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Lab products found in correlation
Coolpix 4500, by Nikon (5 mentions)
Ds u2, by Nikon (5 mentions)
Sodium chloride (nacl), by Merck Group (5 mentions)
Axiocam 506, by Zeiss (4 mentions)
Smz1500, by Nikon (4 mentions)
D glucose monohydrate, by Merck Group (4 mentions)
Axio imager a1, by Zeiss (4 mentions)
Potato dextrose agar (pda), by Merck Group (4 mentions)
Malt extract, by Merck Group (3 mentions)
Silver nitrate, by Merck Group (3 mentions)
Tryptic soy broth (tsb), by Merck Group (3 mentions)
Sz 60, by Nikon (2 mentions)
Coolpix 995, by Nikon (2 mentions)
Zen blue edition software, by Zeiss (2 mentions)
Bacto peptone, by Merck Group (2 mentions)
Titanium dioxide aeroxide tio2 p25, by Evonik (2 mentions)
Methanol, by Merck Group (2 mentions)
Hydrochloric acid (hcl), by Merck Group (2 mentions)
Potassium persulphate, by Merck Group (2 mentions)
Disodium phosphate, by Merck Group (2 mentions)
42 protocols using agar agar
1
Isolation and Cultivation of Morel Fungi
2
Fungal Growth on Ionic Liquids
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Malt extract media (MEM) agar was prepared as 3% malt extract (Roth) and 0.5% peptone from casein (Sigma) in 1.5% agar-agar (Sigma). Minimal medium (MM) agar for fungal spore generation was prepared as 1x Vogel’s salts [28 ] and 2% sucrose in 1.5% agar-agar (Sigma). MM agar with 2% carboxymethyl-cellulose (CMC low viscosity, Sigma Aldrich) instead of sucrose was used for ionic liquid dependent growth experiments for all fungal strains and was supplemented with varying amounts of different ILs provided by Nitrochemie Aschau GmbH (1-butyl-3-methylimidazolium acetate, 1-butyl-3-methylimidazolium chloride, 1-allyl-3-methylimidazolium-chloride, 1-hexyl-3-methylimidazolium-chloride) for some experiments as stated in the main text. Paper samples treated in the aforementioned paper reinforcement process [19 ] with 1-butyl-3-methylimidazolium chloride or 1-butyl-3-methylimidazolium acetate as cellulose solving ionic liquids were provided by Nitrochemie Aschau GmbH. 1-Butyl-3-Methylimidazolium chloride was applied as a solution in DMSO as it was solid at room temperature.
3
Culturing and Preserving Thyronectria Fungi
4
Fungal Cultivation and Microscopic Analysis
5
Swarming Assay for B. pseudomallei
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Swarming agar plates were prepared fresh on the day of the assay using the modified protocol of Tunpiboonsak et al. [55 (link)]. Briefly, plates were prepared using 0.5 % w/v agar-agar (Merck), 8 g/L nutrient broth No.2 (Oxoid) and 5 g/L D(+)- glucose (dextrose) (May and Baker) and dried carefully to give constant moisture content. Overnight cultures of B. pseudomallei parent strain K96243 and mutant strains were subcultured 1/50 into fresh medium with appropriate antibiotics (no selection for parent strain, 25 μg/ml tetracycline for all mutants and complemented mutants) and grown at 37 °C with shaking (200 rpm) to late exponential phase (OD600 of ~2.5). A 5 μl volume of each culture was spotted onto the centre of a swarming agar plate which was then incubated at 37 °C for 18 h in the dark. The diameter of the swarming population was then measured. Statistical analysis of swarming distance was determined by Student’s t-test with a P-value of <0.05 considered significant.
6
Phages Targeting Thermophilic Dairy Strains
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Thirty-five whey samples sourced from 27 distinct dairy fermentation facilities spanning ten countries on four continents were analysed for the presence of phages with lytic activity against 52 S. thermophilus strains. This was achieved using the standard double agar spot assay method [18 (link)], for which LM17 broth was supplemented with 0.25% glycine (Oxoid), 10 mM CaCl2, and 10 g/L (solid) or 4 g/L (semi-solid) agar-agar (Merck, Kenilworth, NJ, USA). Lytic phage activity was confirmed through the development of a clear zone of lysis within the bacterial lawn following overnight incubation at 42 °C. Phage positive samples were subsequently enumerated by plaque assays using the relevant whey–strain combination. Following overnight incubation, plaque morphology was assessed and isolates were deemed distinct based on size (small-medium-large) and/or halo formation. Individual plaque isolates were propagated on the relevant host strain in 10 mL LM17 broth supplemented with 10 mM CaCl2 at 42 °C, filtered through a 0.45 µm filter, and stored at 4 °C.
7
Phylogenetic Specimen Preparation and Analyses
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Collection data, hosts, herbarium, culture and GenBank accession numbers of the specimens used for phylogenetic analyses are provided in Table 1 . Single spore isolates were prepared and grown on 2 % malt extract agar (MEA; 2 % w/v malt extract, 2 % w/v agar-agar; Merck, Darmstadt, Germany). Details of the specimens used for morphological investigations are listed in the Taxonomy section after the respective descriptions.
8
Drosophila Larvae Acephate Toxicity
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Larvae of Drosophila melanogaster were used as experimental organism to assess the toxicological impact of Acephate on hemocyte abundance. Larvae were reared in Standard Drosophila medium (SDM) following the established laboratory rearing techniques (after Dutta et al., 2014 ; and Podder et al., 2012 ). SDM ingredients included corn meal, sucrose (SRL India), agar agar (Merck, India) and yeast extract powder (Merck, India). Nepagin (Supelco, USA) and Propionic acid (Himedia, India) were added for their antifungal properties.
9
Antioxidant Potential of NQNO Compound
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4-Nitroquinoline N-oxide (NQNO), DMSO, l -histidine monochloride, yeast extract, levallorphan, NADP+, glucose-6-phosphate sodium salt, glucose-6-phosphate dehydrogenase, potassium hydroxide, DPPH, gallic acid, and ascorbic acid were purchased from Sigma-Aldrich (Seelze, Germany); Nutrient Broth No. 2, from Argenta (Poznań, Poland); glycerol and neomycin sulfate, from Pharma Cosmetic (Kraków, Poland); sodium sulfate, sodium chloride, d -glucose, and dipotassium hydrogen phosphate from Chempur (Piekary Śląskie, Poland); dichloromethane from Stanlab (Lublin, Poland); and agar-agar, bacto-peptone, beef extract, potato dextrose agar, and 0.2-mm silica-coated aluminum TLC plates from Merck (Darmstadt, Germany).
10
Fungal Isolation Protocol for Ticks
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Ticks' homogenate was diluted with Phosphate Buffer Saline (PBS) to give a final volume of 100μl and diluted solutions these were spread on to Sabouraud Dextrose Agar (Peptone 1%, Glucose 2%, Agar-agar 1.5%; Merck, Germany) and Potato Dextrose Agar (Potato infusion 20%, Dextrose 2%, Agar 2%) plates and incubated for 2 weeks at 25 °C. The plates were periodically checked for fungal colonies. Identification of fungal isolates was performed according to a combination of macro and microscopic morphology.
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