Potato dextrose agar pda

Manufactured by Merck Group

Sourced in Germany, United Kingdom

Potato Dextrose Agar (PDA) is a culture medium used for the growth and enumeration of yeasts and molds. It provides a nutritious environment for these microorganisms to thrive. PDA contains extracts from potatoes and dextrose, which serve as the primary sources of carbohydrates and nutrients.

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40 protocols using potato dextrose agar pda

Potato Fungal Pathogen Cultivation and Analysis

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Five fungal potato phytopathogens, Fusarium sambucinum DSM 62397, Alternaria tenuissima DSM 63360, Colletotrichum coccodes DSM 62126, Phoma exigua DSM 62040, and Rhizoctonia solani DSM 22843, were used in the experiments. The strains were purchased from the German Collection of Microorganisms and Cell Cultures GmbH (DSMZ, Braunschweig, Germany). All strains were activated on Potato Dextrose Agar (PDA; Merck, Darmstadt, Germany) and stored at 4 °C. Pathogen suspensions were prepared from the pure cultures on the PDA agar plates and adjusted to a final concentration of 10⁶ CFU/mL in saline solution (0.85% NaCl).
For fungal metabolite analysis (Section 2.3), samples of fungal cultures on Malt Extract Agar (MEA; Merck, Darmstadt, Germany) plates and potatoes infected with phytopathogens were prepared.

Steglińska A., Sulyok M., Janas R., Grzesik M., Liszkowska W., Kręgiel D, & Gutarowska B. (2023). Metabolite Formation by Fungal Pathogens of Potatoes (Solanum tuberosum L.) in the Presence of Bioprotective Agents. International Journal of Environmental Research and Public Health, 20(6), 5221.

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Fungal Secondary Metabolite Production

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The fungal isolates were revived by culturing them on Potato Dextrose agar (PDA, Merck, Darmstadt, Germany) and incubated at 25 °C for 10 days. A plug of active mycelia was inoculated into a 250 mL Erlenmeyer flask containing 50 mL of malt extract broth (MEB; Merck, Darmstadt, Germany). The numbers of spores were counted with a hemocytometer (Merck, Johannesburg, South Africa) and adjusted to 1 × 10⁶ conidia/mL. The secondary metabolites were produced by fermentation as described by Premjanu and Jaynthy [11 ] and each fermentation performed in triplicate. Briefly, fungal cell mass was removed by filtration through a 0.45 µm syringe filter and the resulting filtrate stored in sterile conical flasks at 4 °C, until further use.

Manganyi M.C., Tchatchouang C.D., Regnier T., Bezuidenhout C.C, & Ateba C.N. (2019). Bioactive Compound Produced by Endophytic Fungi Isolated From Pelargonium sidoides Against Selected Bacteria of Clinical Importance. Mycobiology, 47(3), 335-339.

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Antimicrobial Potato Dextrose Agar

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Potato dextrose agar (PDA), glucose, potato peptone, yeast extract, silver nitrate (AgNO₃), and sodium hydroxide (NaOH) were purchased from Merck (Darmstadt, Germany). Chitosan, sodium tripolyphosphate, Muller–Hinton broth, RPMI 1640 media, ceftazidime, and amphotericin B were obtained from Sigma-Aldrich (Darmstadt, Germany).

Hermosilla E., Díaz M., Vera J., Contreras M.J., Leal K., Salazar R., Barrientos L., Tortella G, & Rubilar O. (2023). Synthesis of Antimicrobial Chitosan-Silver Nanoparticles Mediated by Reusable Chitosan Fungal Beads. International Journal of Molecular Sciences, 24(3), 2318.

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Quantitative Microbiological Analysis of Cheese

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Representative duplicate 10 g portions were retrieved from each cheese sample during various time intervals and blended with 90 mL of sterilized 2% w/v tri-sodium citrate solution. The suspension was then submitted to 10 decimal serial dilutions with ¼ strength Ringer’s solution. Viable counts of total aerobic bacterial counts, Lactococci, Lactobacilli, yeasts and fungi, and coliforms were determined in triplicate by pour plating of appropriate dilutions (0.1 mL or 1 mL) on the selective media for each species according to instructions of the manufacturer [26 (link)]. Specifically, viable counts of total aerobic bacterial counts were enumerated on plate count agar (PCA) (Merck) by suspended 0.1 mL of the sample after incubation at 30 °C for 72 h. Lactococci were enumerated on M-17 agar (Merck) by suspended 1 mL of the sample after incubation 30 °C for 48–72 h. Lactobacilli were enumerated on MRS agar (Merck) by suspended 1 mL of the sample after incubation at 37 °C for 48 h. Yeasts and fungi were determined by suspended 0.1 mL of the sample after incubation on Potato Dextrose agar (PDA) (Merck) at 30 °C for 72 h. Finally, coliforms were enumerated on Violet Red Bile agar (LabM, Heywood, U.K.) by suspended 1 mL of the sample after anaerobic incubation at 30 °C for 24 h. All cell counts were expressed as log of mean colony forming units (CFU).

Mantzourani I., Terpou A., Alexopoulos A., Chondrou P., Galanis A., Bekatorou A., Bezirtzoglou E., Koutinas A.A, & Plessas S. (2018). Application of A Novel Potential Probiotic Lactobacillus paracasei Strain Isolated from Kefir Grains in the Production of Feta-Type Cheese. Microorganisms, 6(4), 121.

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Isolation of Endophytic Fungi from Medicinal Plants

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Endophytes were isolated from fresh leaves and stems of anticancer plants; C. citratus, M. koenigii, O. diffusa and P. bleo. These plants were sampled randomly from a housing garden at Bandar Tun Hussein Onn, Selangor, Malaysia. The plant tissues were first left under running tap water for 15 min. The leaf tissues were then cut into 2 cm × 2 cm segments whereas the stem tissues were cut to a length of 2 cm each. The tissues were subjected to triple sterilization technique, beginning with 40% household bleach for 5 min, then in 50%, 70%, 90% and 100% ethanol for 2 min each immersion, and finally a quick rinse in sterilized distilled water. This procedure was performed and repeated twice on the same batch of plant tissues [16] . Surface-sterilized tissue segments were then injured and seeded onto Potato Dextrose Agar (PDA, Merck) supplemented with Rose bengal (0.033 g L⁻¹) and chloramphenicol (0.05 g L⁻¹), to select for fungal endophytes [8] . Controls were prepared by seeding non-injured surface-sterilized tissue segments onto PDA plates. The absence of mycelial growth indicated effective surface sterilization while mycelial growth from injured tissues was identified as endophytes. All PDA plates were incubated at 28 ± 2 °C for 7–14 days. The fungal colonies were subsequently transferred to fresh PDA plates to establish pure cultures.

Chow Y, & Ting A.S. (2014). Endophytic l-asparaginase-producing fungi from plants associated with anticancer properties. Journal of Advanced Research, 6(6), 869-876.

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Culturing Fungal and Bacterial Pathogens

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The fungal pathogens Aspergillus flavus (FCBP-PTF-1265) and Aspergillus fumigatus (FCBP-MF-923) were obtained from the First Fungal Culture Bank of Pakistan (FCBP), University of the Punjab, Pakistan. Rhizopus oryzeae (ATCC 11886 (AY 803930)) was obtained from the Department of Microbiology, Quaid-i-Azam University, Islamabad.
The bacterial isolates were first sub-cultured in a nutrient broth (Sigma) and incubated at 37 °C for 18 h. The fungal isolates were sub-cultured on potato dextrose agar (PDA) (Merck) for 7 d at 25 °C.

Naz R., Ayub H., Nawaz S., Islam Z.U., Yasmin T., Bano A., Wakeel A., Zia S, & Roberts T.H. (2017). Antimicrobial activity, toxicity and anti-inflammatory potential of methanolic extracts of four ethnomedicinal plant species from Punjab, Pakistan. BMC Complementary and Alternative Medicine, 17, 302.

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Isolation of Phytopythium spp. from Kiwifruit

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Phytopythium spp. isolates were collected from 18 A. chinensis orchards during the period August–October of 2016, 2018 and 2019, in Piedmont, north-western Italy (Table 1 and Table S1). The strains were isolated from kiwifruit plants showing typical KVDS symptoms, i.e., reduction in plant vigour, leaf curling, or complete decline, focusing on microorganisms associated with infected tissues. Isolation was carried out from symptomatic rotten roots at the margin between infected and healthy tissue to fulfil the first postulate of Koch, as previously described by Prencipe et al. [10 (link)]. Briefly, roots were first surface-disinfected with 1% sodium hypochlorite, washed in sterile deionised water and air-dried. Five fragments of each root were cut at the symptom edges and plated onto Potato Dextrose Agar (PDA, Merck, Germany) and semi-selective oomycete PARP medium (17 g corn meal agar, 0.01 g Pimaricin, 0.01 g Ampicillin 0.01 g, Rifampicin and 0.07 g Pentachloronitrobenzene,) Petri dishes. After 4 days of incubation at 25 ± 1 °C, 71 representative isolates were selected (Table 1), based on colony morphology, and they were maintained in tubes of PARP medium.

Prencipe S., Schiavon G., Rosati M., Nari L., Schena L, & Spadaro D. (2023). Characterization of Phytopythium Species Involved in the Establishment and Development of Kiwifruit Vine Decline Syndrome. Microorganisms, 11(1), 216.

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Airborne Fungal Sampling by Settle Plate

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A total of 1092 air samples were prepared from 93 sampling sites during July 2012 using settle plate method according to Hoekstra et al.[30 ]. Sampling sites were chosen according to the extents of studied areas covering overall parts of each location. Plastic plates (9 mm Dia.) contained Malt extract agar (MEA; E. Merck, Darmstadt, Germany) and Potato dextrose agar (PDA; E. Merck, Darmstadt, Germany) with lids open were placed in sampling sites at the height of about one meter above the floor, for 30 min [31 ]. After that, the plates were transferred to the laboratory for fungal isolation and identification.

Shams-Ghahfarokhi M., Aghaei-Gharehbolagh S., Aslani N, & Razzaghi-Abyaneh M. (2014). Investigation on distribution of airborne fungi in outdoor environment in Tehran, Iran. Journal of Environmental Health Science and Engineering, 12, 54.

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Microbial Profiling of Vinegar Varieties

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Serial dilutions of vinegar samples (TTV, PTV and UTV) were performed in peptone water solution for microbial count. Acetic acid bacteria (AAB) counts were determined in Glucose Yeast Extract Calcium Carbonate Agar (GYC, pH 6.8, HiMedia, Mumbai, India). The surface plate method was used and incubated for 5–10 days at 30 °C in plates [27 (link)]. PCA (Plate Count Agar—Merck, Darmstadt, Germany) was used for TAPC (total aerobic plate count). Samples were incubated at 28 °C for 48 h. For yeast and mold count, Potato Dextrose Agar (PDA, pH 5.6, Merck, Germany) acidified with 10% tartaric acid (Merck, Darmstadt, Germany) was used. The Petri dishes were incubated at 25 °C for 3–5 days. Total Enterobacteriaceae count was determined in VRBG (Violet Red Bile Glucose Agar-Merck, Darmstadt, Germany) incubated at 37 °C for 24 h. The results were expressed as log colony forming units (CFU) per milliliter of PTV, TTV and UTV [28 (link)].

Yıkmış S., Aksu F., Altunatmaz S.S, & Çöl B.G. (2021). Ultrasound Processing of Vinegar: Modelling the Impact on Bioactives and Other Quality Factors. Foods, 10(8), 1703.

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Transformation of Ustilago maydis strain SG200

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All U. maydis strains from this study are derived from the solopathogenic strain SG200 (a1, mfa2, bE1/bW2; (Bölker et al., 1995 (link)). Protoplast preparation and transformation of U. maydis strain SG200 was carried out as previously described (Bösch et al., 2016 (link)). Briefly, for each construct, around 5μg of the plasmid DNA was linearized by digestion with restriction enzyme SspI, and U. maydis protoplasts were transformed with linearized DNA for the integration into the ip locus by homologous recombination using p123 derivatives as described previously (Loubradou et al., 2001 (link); Navarrete et al., 2022 (link)). The individual colonies were grown on Potato Dextrose Agar (PDA) (Merck, Kenilworth, NJ, USA) media with respective antibiotics (Carboxin, CBX 2μg/ml). The transformants were confirmed for the presence of the desired insert by PCR. All the U. maydis strains were grown at 28°C in YEPS liquid medium (0.4% yeast extract, 0.4% peptone, and 2% glucose) with overnight shaking at 180 rpm.

Ingole K.D., Nagarajan N., Uhse S., Giannini C, & Djamei A. (2022). Tetracycline-controlled (TetON) gene expression system for the smut fungus Ustilago maydis. Frontiers in Fungal Biology, 3, 1029114.

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