The bacterial species used for microbial tests consisted of S. aureus (ATCC6538), P. aeruginosa (ATCC9027) and E. faecalis (ATCC29212), and C. albicans (ATCC10231) fungal species, which were provided by the Microbiology Laboratory of Tabriz Faculty of Medicine. Brain-heart broth (Merck, Darmstadt, Germany) and blood agar culture media were used to proliferate the bacterial species.
Brain heart broth
Manufactured by Merck Group
Sourced in Germany
Brain heart broth is a microbial culture medium used for the growth and isolation of a variety of microorganisms, including bacteria and fungi. It provides the necessary nutrients and growth factors for the cultivation of these organisms.
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Lab products found in correlation
Rose bengal, by Merck Group (1 mentions)
Sulfanilamide, by Merck Group (1 mentions)
Brain heart agar, by Merck Group (1 mentions)
Fetal calf serum (fcs), by Merck Group (1 mentions)
Hemin, by Merck Group (1 mentions)
L cysteine hydrochloride, by Merck Group (1 mentions)
Tryptone soy broth (tsb), by Thermo Fisher Scientific (1 mentions)
K2hpo4, by Carl Roth (1 mentions)
Na2co3, by Carl Roth (1 mentions)
Vitamin k1, by Carl Roth (1 mentions)
Myristic acid, by Merck Group (1 mentions)
Silver nitrate, by Merck Group (1 mentions)
Muller hinton, by Merck Group (1 mentions)
Mueller hinton agar (mha), by Kasvi (1 mentions)
Tris buffered saline (tbs), by Scharlab (1 mentions)
Purelink genomic dna kit, by Thermo Fisher Scientific (1 mentions)
Bigdye v3, by Thermo Fisher Scientific (1 mentions)
11 protocols using brain heart broth
1
Microbial Strains and Culture Media
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The bacterial species used for microbial tests consisted of S. aureus (ATCC6538), P. aeruginosa (ATCC9027) and E. faecalis (ATCC29212), and C. albicans (ATCC10231) fungal species, which were provided by the Microbiology Laboratory of Tabriz Faculty of Medicine. Brain-heart broth (Merck, Darmstadt, Germany) and blood agar culture media were used to proliferate the bacterial species.
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The bacterial species used for microbial tests consisted of S. aureus (ATCC6538), P. aeruginosa (ATCC9027) and E. faecalis (ATCC29212), and C. albicans (ATCC10231) fungal species, which were provided by the Microbiology Laboratory of Tabriz Faculty of Medicine. Brain-heart broth (Merck, Darmstadt, Germany) and blood agar culture media were used to proliferate the bacterial species.
2
Antibacterial Evaluation of Rose Bengal
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Rose Bengal, sulfanilamide, reagents for synthesis and solvents were purchased from Sigma-Aldrich Chemie GmbH (Schnelldorf, Germany). Methicillin was purchased from TOKU-E (Bellingham, WA, USA), growth media Brain Heart broth (BH) and Brain Heart agar (BHA) were purchased from Merck (Darmstadt, Germany) and Antibiotic medium 3 for minimal inhibitory concentration (MIC) studies was purchased from Becton Dickinson & Company (Le Pont de Claix, France). Antibiotic discs were purchased from Abtek Biologicals Ltd. (Liverpool, UK). Antibiotic sensitive and resistant strains of S. aureus and P. aeruginosa were hospital isolates from the Meir Medical Center (Kfar Saba, Israel). S. aureus strain ATCC 11541 and P. aeruginosa ATCC 25668 strains were purchased from ATCC (Manassas, VA, USA).
3
Anaerobic Growth of Mucispirillum schaedleri
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Unless otherwise stated, M. schaedleri ASF 457 MCS was cultured under anaerobic conditions under an N2 and 8% H2 atmosphere at 37°C without shaking using anaerobic Mucispirillum medium (AMM), which is based on Trypticase soy agar and contains (per liter) 18 g brain heart broth (Merck), 15 g tryptone soy broth (Oxoid), 5 g yeast extract (Bacto yeast extract), 2.5 g K2HPO4 (Carl Roth), 1 mg hemin (Sigma), 0.5 mg vitamin K1 (Carl Roth), 0.4 g Na2CO3 (Carl Roth), 3% fetal calf serum (Sigma), 0.5 mg l -cysteine hydrochloride (Sigma), and 0.5 mg alpha-(d +)-glucose monohydrate (Carl Roth) (87 ). M. schaedleri ASF 457 MCS was analyzed for growth in AMM with or without the presence of the following compounds: hydrogen (8%), formate (0.5, 2.5, 10, or 50 mM), nitrate (2 or 10 mM), and fumarate (10 or 50 mM). Growth was quantified by optical density measured at 600 nm (OD600) (M107 high-specification visible spectrophotometer; Spectronic Camspec Ltd., Leeds, United Kingdom). OD600 values were normalized by subtracting the background absorbance values of abiotic-medium controls.
4
Oral Swab Collection Procedure for Calves
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To collect the oral swabs, the calves were restrained by a feeding fence. A helper opened the calves’ mouth using a wooden mouth wedge with an opening in the middle through which the veterinarian took the sample with a sterile dry swab. Taking into account the defensive movements of the calves, the swab was taken from the buccal cavity or as far back in the throat as possible from the tonsils. Calves that had just eaten were sampled later to avoid contamination by feed residues. In between, the mouth wedge was washed in iodine soap containing povidone iod (Novasan Plus Jodseife, Ferd. Eimermacher GmbH and Co. KG, Nordwalde, Germany). After sampling, the swabs were transported in sterile test tubes containing 2 mL brain-heart-broth (Merck KGaA, Darmstadt, Germany) and shipped to the departmental laboratory in a cooling box for further analysis at the day of sampling.
5
Propagation of Campylobacter jejuni Strain
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The C. jejuni EMCC 1835 reference strain was brought from Microbial Resource Center (MIRCEN), Ain Shams University, Cairo, Egypt. The cell counts were accustomed to 106 CFU/mL as the infective dose is >105 CFU/g [20 (link)]. The strains were stored at a temperature of −80 °C in Brain Heart Broth (Merck, 1.10493.0500, Darmstadt, Germany), including both glycerol and lysed horse blood (LHB) (Oxoid, SR048C, Hampshire, UK), then they were sub-cultured at a temperature of 42 °C on Columbia agar base (CAB) (Oxoid, CM0331, Hampshire, UK) under certain conditions as follow: 5% O2, 85% N2, and 10% CO2.
6
Enzymatic Esterification of Fatty Acids
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Myristic acid, palmitic acid, glycerol, sulfuric acid, n-hexane, ethanol, acetone, sodium hydroxide, p-toluenesulfonic acid (pTSA), Amberlyst-15, sodium carbonate, potassium carbonate, ethyl acetate, chloroform, and anhydrous sodium sulfate were purchased from Merck (Darmstadt, Germany), while TLIM was obtained from Novozymes (Bagsværd, Denmark). All chemicals were used as received without any further purification. Thin layer chromatography (TLC) was carried out using an aluminium plate (Merck) coated with silica gel 60 F254 (20 × 20 cm). The brain-heart broth and Sabouraud agar powder which contains 4.00% of dextrose as the microbial nutrient was purchased from Merck.
7
Silver Nanoparticle Synthesis and Antimicrobial Testing
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Silver nitrate, ACS reagent, 99+% (AgNO3) for AgNP synthesis were obtained from Sigma-Aldrich, for appropriate microorganisms culturing Muller Hinton (MHB), Sabaroud Dextrose Broth (SDB) and Brain Heart Broth (BHB) were obtained from Merck. All microorganism strains were kept at −20˚C in the appropriate medium containing 10% glycerol and regenerated twice prior to the manipulations. For anticancer analyses all chemicals were purchased from Sigma Aldrich, Germany. Freshly prepared doubly distilled water was used throughout the experimental work.
8
Cultivation of Virulent Streptococcus pneumoniae
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We obtained the S. pneumoniae D39 serotype 2 from the Korea Centers for Disease Control and Prevention. D39 is the reference strain of S. pneumoniae, and is the most widely studied strain. It is highly encapsulated and results in a lethal outcome following infection. We cultured S. pneumoniae D39 on 5% sheep blood agar plates (Medexx) at 37℃/5% CO2 for 18 h before harvesting. Harvested S. pneumoniae D39 was rinsed and resuspended in sterilized phosphate-buffered saline (PBS) then transferred to brain heart broth (Merck) and cultured without shaking for 6 h at 37℃/5% CO2. The S. pneumoniae D39 cultures were centrifuged, washed and rinsed with sterile PBS. The infectious dose was determined by counting colony forming units (CFU) after plating serially diluted S. pneumoniae D39 suspensions on 5% sheep blood agar plates. Plates were cultured at 37℃/5% CO2 for 24 h.
9
Inactivation of Acinetobacter baumannii Cultures
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A. baumannii cultures were incubated overnight in brain heart broth (Merck, Darmstadt, Germany) under agitation at 120 rpm at 37 °C. Cell densities were at the optical density: 600 nm (OD600) = 0.8. Subsequently, they were inactivated using 3% formaldehyde (Alphatec®, São Paulo, Brazil) for 2 h at 30 °C, followed by 16 h at 60 °C, under agitation at 120 rpm. Inactivation was confirmed by adding 1 mL of the inactivated culture to Mueller–Hinton agar (Kasvi®, Paraná, Brazil) and incubating at 37 °C for 48 h [32 (link)].
10
Stainless Steel Biofilm Cultivation
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Clean and sterile stainless steel surfaces where monospecies and mixed biofilms are grown.
• Clean and sterile stainless steel surfaces, without microbial contamination, used as control • Clean and sterile stainless steel surfaces where planktonic cells are inoculated to show the presence of biofilms without biofilm formation.
Generic, non-selective culture media were employed for growing and forming biofilms: TBS (Typtic Soy Broth, Scharlab, Barcelona, Spain) and BHI (Brain Heart Broth, Merck, Madrid, Spain). Working temperature was 37 • C ± 1 • C, which is the optimal growth temperature for the microorganisms tested.
Biofilms were grown under static and dynamic conditions (11.6 mL/min), with and without replenishment of nutrition media, in order to optimize microbial adhesion to surfaces. Two biofilm growth periods, 3 and 10 days, were considered for testing TBF 300 against biofilms with different levels of strength.
• Clean and sterile stainless steel surfaces, without microbial contamination, used as control • Clean and sterile stainless steel surfaces where planktonic cells are inoculated to show the presence of biofilms without biofilm formation.
Generic, non-selective culture media were employed for growing and forming biofilms: TBS (Typtic Soy Broth, Scharlab, Barcelona, Spain) and BHI (Brain Heart Broth, Merck, Madrid, Spain). Working temperature was 37 • C ± 1 • C, which is the optimal growth temperature for the microorganisms tested.
Biofilms were grown under static and dynamic conditions (11.6 mL/min), with and without replenishment of nutrition media, in order to optimize microbial adhesion to surfaces. Two biofilm growth periods, 3 and 10 days, were considered for testing TBF 300 against biofilms with different levels of strength.
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