Reference microbial strains were obtained from the American Type Culture Collection (ATCC), including Staphylococcus aureus ATCC 25923, Enterococcus faecalis ATCC 29212, Bacillus subtilis ATCC 6635, Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27833 and Shigella flexneri ATCC 12022. The Listeria monocytogenes PCM 2191 and Bacillus cereus PCM 1948 strains were taken from the Polish Collection of Microorganisms (PCM). One bacterial strain Salmonella Enteritidis ZMF 279 was derived from the collection of the Department of Pharmaceutical Microbiology and Diagnostic Microbiology, Medical University of Lodz. Two fungal strains were also used: Candida albicans ATCC 10231 and Aspergillus brasiliensis ATCC 16404. All tested microorganisms were stored at –80 °C in 15% glycerol stocks. Before the investigation, the bacterial strains were transferred to Mueller-Hinton agar medium (Oxoid, Thermo Fisher Scientific, Waltham, MA, USA) and cultured overnight at 37 °C. Fungal strains were transferred on Sabouraud agar medium (Oxoid, Thermo Fisher Scientific, Waltham, MA, USA) and cultured for two days at 30 °C.
Sabouraud agar medium
Manufactured by Thermo Fisher Scientific
Sourced in United Kingdom
Sabouraud agar medium is a microbiological culture medium used for the isolation and identification of fungi, including yeasts and molds. It is formulated to support the growth of a wide range of fungal species while inhibiting the growth of bacteria. The medium contains nutrients, such as peptone and dextrose, that are specifically tailored to the nutritional requirements of fungi.
Automatically generated - may contain errors
Lab products found in correlation
Bacillus subtilis, by American Type Culture Collection (1 mentions)
Shigella flexneri, by American Type Culture Collection (1 mentions)
Mueller hinton agar medium, by Thermo Fisher Scientific (1 mentions)
Escherichia coli, by American Type Culture Collection (1 mentions)
Pseudomonas aeruginosa, by American Type Culture Collection (1 mentions)
Staphylococcus aureus, by American Type Culture Collection (1 mentions)
Enterococcus faecalis, by American Type Culture Collection (1 mentions)
3 protocols using sabouraud agar medium
1
Microbial Strain Collection and Culture Conditions
2
High-Pressure Homogenization of Microbial Cells
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A PANDA high-pressure homogenizer (Niro Soavi, Parma, Italy) was used for all homogenizing treatments. The initial cell load before HPH treatment was about 7.7 log CFU ml-1. The control treatment was performed at 0.1 MPa, and the mild treatment was performed at 80 MPa. The machine was supplied with a homogenizing PS-type valve with a flow rate of 10 l h-1. The valve assembly included a ball-type impact head made of ceramic, a stainless steel, large-inner-diameter impact ring and a tungsten carbide passage head. The inlet temperature of the cell suspension was 40°C, and the rate of temperature increase was approximately 2°C 10 MPa-1. During the treatment, the cells remained under pressure for a few milliseconds. Immediately after the treatment at 0.1 and 80 MPa, the cell loads were verified by plating the cell suspension on Sabouraud agar medium (Oxoid, Basigstone, UK) after sonication with a Liarre Starsonic 90 to separate cell clumps.
3
Quantifying Airborne and Drilling Microbes
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The bacteria and fungi collected on polycarbonate filters were extracted in 5.5 mL sterile solution (0.05% Tween 80 and 0.85% NaCl) by orbital shaking (500 rpm) for 15 min at room temperature. The dust suspensions were plated in different dilutions on three types of agar media: NA for the quantification and identification of bacteria; DG18 agar and Sabouraud agar medium (SA, Oxoid, Basingstoke, UK) for quantification of fungi. The plates were incubated at different temperatures with or without oxygen.
The drilling waste samples were after arrival at the laboratory shaken (500 rpm) for 15 min at room temperature. The samples were plated and incubated as described in thesupplementary file (Methods section). Part of each suspension was stored with 33% glycerol at −80°C for re-plating and biofilm assay.
Bacteria able to grow aerobically at 25°C are called bacteria, and bacteria grown anaerobically are called anaerobic bacteria. The number of fungi on DG18 and SA did not differ significantly, and counting numbers are presented for DG18 data. The data on airborne microorganisms are presented as TWA (CFU)/m3 air, and microorganisms in the drilling waste samples as CFU/mL.
The drilling waste samples were after arrival at the laboratory shaken (500 rpm) for 15 min at room temperature. The samples were plated and incubated as described in the
Bacteria able to grow aerobically at 25°C are called bacteria, and bacteria grown anaerobically are called anaerobic bacteria. The number of fungi on DG18 and SA did not differ significantly, and counting numbers are presented for DG18 data. The data on airborne microorganisms are presented as TWA (CFU)/m3 air, and microorganisms in the drilling waste samples as CFU/mL.
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