The antimicrobial assay was conducted using the broth macrodilution method [17 (link)]. The A. ursinum extract was added to 1 mL of growth medium (Nutrient broth, Himedia, India) to give concentrations of 5, 10, 15, 20, 25 and 30 mg/mL. In order to obtain artificially contaminated samples at levels of 102, 103, 104, 105 and 106 cfu/mL, the appropriate volume of inoculum from the selected decimal dilution was aseptically transferred into 1 mL of growth media with a defined concentration of A. ursinum extract. Each sample was separately contaminated with individual test bacteria. Artificially contaminated samples of subcritical water extract of A. ursinum in Nutrient broth (Himedia, India) were incubated for 24 h at 37 ± 1 °C. Bacterial growth was determined by taking samples after incubation and plating on cultivation media (Plate count agar, Himedia, India) according to the standard method ISO 4833-1:2013. Artificially contaminated Nutrient broths (Himedia, India) without added plant extract were used as positive controls, while non-inoculated Nutrient broths (Himedia, India) containing extract of A. ursinum in selected concentrations were used as a negative controls. All experiments were conducted in triplicate.
Nutrient broth
Manufactured by HiMedia
Sourced in India, Pakistan, United States
Nutrient broth is a common laboratory medium used for the growth and cultivation of a wide range of microorganisms. It provides essential nutrients, including carbon, nitrogen, and other essential elements, to support the growth and proliferation of bacteria, fungi, and other microbes. The broth is typically clear, with a neutral pH, and can be used in a variety of microbiology applications, such as bacterial culture and isolation, antimicrobial susceptibility testing, and biochemical identification of microorganisms.
Automatically generated - may contain errors
Lab products found in correlation
Nutrient agar, by HiMedia (48 mentions)
Mueller hinton agar (mha), by HiMedia (13 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (9 mentions)
Agar, by HiMedia (6 mentions)
Ethanol, by Merck Group (6 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (6 mentions)
Potato dextrose broth (pdb), by HiMedia (5 mentions)
Muller hinton broth, by HiMedia (5 mentions)
Muller hinton agar, by HiMedia (5 mentions)
2 2 diphenyl 1 picrylhydrazyl (dpph), by Merck Group (5 mentions)
N hexane, by Merck Group (5 mentions)
Methanol, by Merck Group (5 mentions)
Luria bertani broth, by HiMedia (4 mentions)
Nutrient agar media, by HiMedia (4 mentions)
Mueller hinton broth (mhb), by HiMedia (4 mentions)
Glutaraldehyde, by Merck Group (4 mentions)
Methanol, by Loba Chemie (3 mentions)
Trolox, by Merck Group (3 mentions)
Sabouraud dextrose broth (sdb), by HiMedia (3 mentions)
Avance 2 400 nmr spectrometer, by Bruker (3 mentions)
199 protocols using nutrient broth
1
Antimicrobial Efficacy of Allium ursinum Extract
2
Cultivation of Bacterial and Fungal Species
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Escherichia was cultivated on agar and Muller–Hinton broth (Himedia, Mumbai, India) for 18–20 h at 37 °C. Staphylococcus was cultivated using Nutrientagar and Nutrientbroth (Himedia, India) and the GRM medium (Obolensk, Moscow, Russia) for 18–20 h at 37 °C. Nutrientagar and Nutrientbroth (Himedia, India) were used for Bacillus cultivation, and the culture was grown for 18–24 h. The cultivation of fungi of the genus Candida was conducted on Sabouraud’s medium at 30–37 °C for 48 h.
3
Protease Production from Cow Milk Isolate
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The CMSS isolate isolated from cow milk was used for the NK production. The 1% seed inoculums were inoculated in to three different medium containing 100 mL of nutrient broth, production medium (1% shrimp shell, 0.1% KH2PO4, 0.05% MgSO4, pH-7 at 37˚C) and nutrient broth (Hi-Media) with 0.1% shrimp shell and incubated at 37˚C for 24 hours in a shaker at 120 rpm. The medium was then centrifuged at 10,000 rpm for 10 minutes and the supernatant was used as the crude enzyme extract. The enzyme from the three different media was analyzed by radial caseinolytic plate assay (7 (link), 8 ).
4
Bacillus Spp. Antagonist Inoculum Preparation
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The inoculum preparation started by incubation of the potential producing Bacillus spp. strains on nutrient agar for 48 h at 28 °C. The second step included transferring the loopful biomass of antagonists into the liquid media (nutrient broth—Himedia, Laboratories, Mumbai, India) and incubating at 28 °C for 48 h on a rotary shaker with an agitation rate of 170 rpm. Inoculation of cultivation media was performed by adding 10% (v/v) of the prepared inocula (5 mL) to the Erlenmeyer flasks containing 50 mL of nutrient broth (Himedia Laboratories, Mumbai, India). The cultivation was carried out on a rotary shaker for 96 h, with a temperature of 28 °C, with an agitation rate of 170 rpm.
5
Isolation and Cultivation of Myroid gitamensis
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“Myroid gitamensis” BSH-3 a strain previously isolated from the soil sample collected from uncultivated land of the Mohali district of Punjab, India (30°53′ N 76°38′ E) (Kaur and Kaur, 2021 (link)) was used throughout the study. The pure culture was grown in Pikovskaya agar medium supplemented with 5% tricalcium phosphate (w/v) and 5% soya lecithin (w/v). The purified culture was preserved in glycerol stock at −20°C. The cells were sub-cultured in 5 mL nutrient broth (HiMedia) at 28 ± 2°C for 24 h. Then, the cultures were transferred into 500 mL of nutrient broth and incubated to grow in continuous shaking conditions at 28°C until the bacterial strain reached a concentration of 1012CFU/mL (Kumar et al., 2017 (link)). Cell pellets were harvested by centrifugation at 10,000g for 10 min at 4°C (Remi Centrifuge VCBS2252, Vasai, India). The pellets were washed three times with sterile 0.5 M phosphate buffer and suspended in 1 mL sterile distilled water to obtain concentrated “M. gitamensis” BSH-3 (~109 CFU/mL) (Kaur and Kaur, 2023 ).
6
Antimicrobial and Antioxidant Assays
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Ciprofloxacin and gentamicin (Microxpress, a division of Tulip Diagnostics Pvt. Ltd.) antibiotic discs were utilized as standard drugs for antibacterial assay. Nutrient broth (HiMedia Laboratories Pvt. Ltd., Mumbai), Mueller Hinton Agar (MHA) (HiMedia Laboratories Pvt. Ltd., Mumbai), 3 (4, 5 dimethylthiazol-2-yl)-2-5-diphenyl tetrazolium bromide (MTT) (Beyotime Biotechnology, China), DPPH (Thermo Fisher scientific, India Pvt. Ltd; Mumbai), Barium chloride (HiMedia Laboratories Pvt. Ltd., Mumbai), Folin-Ciocalteu reagent, quercetin, ascorbic acid dimethyl sulfoxide (Thermo Fisher Scientific, India Pvt. Ltd., Mumbai).
7
Trichoderma Strains from Coal Mines
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Strains used in this work were isolated from the underground of lignite mine Záhorie, Slovakia, except for Trichoderma sp. var. Modra, which was isolated from the rotten wood bark found near the town Modra, Slovakia. The isolate Trichoderma sp. var. Čáry, was isolated from the lignite layer in the drift wall opened to air for several years after cutting off the surface layer by a sterile chisel. The same is true for Trichoderma, var. Store which was isolated from a compact piece of coal found at the store of freshly excavated coal. Its ITS1 sequence showed a 98 % similarity to T. atroviride. Trichoderma isolates, designated as Čelba 1 and Čelba 2 also characterised by their ITS sequences. The obtained sequences were submitted to Gen-Bank and are available under the accession num-bers AY220263-AY220269. Their analyses showed that the isolated species from coal are closely related to Trichoderma viride and/or Trichoderma atroviride whereas Trichoderma sp. var. Modra is related to Trichoderma pseudokoningii. Fungal isolates were propagated on Czapek-Dox agar supplemented with 0.5 % (w/v) yeast extract at 26 o C in dark. For experiments, conidia were scrapped from the agar surface and washed once with water and counted. Isolated bacteria were cultivated on Nutrient Broth (HiMedia) plates at 37 o C.
8
Bacterial Challenge in Aquaculture Fish
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After the growth experiment, fish were challenged with S. aureus for 15 days. The S. aureus culture was prepared in 10 ml volume of nutrient broth (HiMedia Ltd., Lahore, Pakistan). Subsequently, the culture was vortexed, and incubated in shaker incubator for 24 hours at 37°C. The culture was centrifuged (Micro Prime Centrifuge, Pocklington, UK) at 5000 rpm for 15min at 4°C to get the hard pellet at the bottom. The obtained pellet underwent several washings, employing sterile phosphate buffer saline (PBS). Following thorough washing, the pellet was re-suspended in PBS (pH 7.4). To ascertain the optical density of bacterial suspension, a UV spectrophotometer was utilized to obtain concentration of 5×105 CFU/ml. The control group was split into two distinct subgroups: positive control (+ve T0) and negative control (-ve T0). Fish in -ve T0 was given bath with PBS only, whereas the other groups (+ve T0, T1, T2, T3, T4, T5, T6, and T7) (n = 15 for each treatment) were exposed to S. aureus (5×105 CFU/ml). Fish were bathed for 2 hours and the bath was repeated after seven days. Throughout the challenge period, all fish in all treatments were fed with their relevant diets.
9
Coumarin-Based Compound Evaluation
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Coumarin was obtained from
Koch Light Laboratories Limited, Saudi Arabia. Sodium phosphate dibasic
dehydrate, 14,000 Da, and poloxamer were obtained from (Sigma-Aldrich,
India). MTT and DMSO were obtained from Merck Chemicals, Germany.
Mueller–Hinton agar (MHA), Mueller–Hinton broth (MHB),
and nutrient broth were purchased from HiMedia Laboratories. Double-distilled
water (DD) was used throughout the experiment. Because the other chemicals
and solvents were of analytical purity, they did not require additional
purification prior to use.
Koch Light Laboratories Limited, Saudi Arabia. Sodium phosphate dibasic
dehydrate, 14,000 Da, and poloxamer were obtained from (Sigma-Aldrich,
India). MTT and DMSO were obtained from Merck Chemicals, Germany.
Mueller–Hinton agar (MHA), Mueller–Hinton broth (MHB),
and nutrient broth were purchased from HiMedia Laboratories. Double-distilled
water (DD) was used throughout the experiment. Because the other chemicals
and solvents were of analytical purity, they did not require additional
purification prior to use.
10
Isolation of Migratory Bird Gut Bacteria
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Only freshly dropped wet fecal samples were collected from the ground and tree leaves where the migratory birds were available. The sterile conditions were maintained during sample collection. A total of 66 fecal materials of migratory birds were collected from Baojani Baor, Mohammadpur Upazila (23.4056° N, 89.5686° E) in Magura district of Bangladesh during November 2019 and November 2020. The samples were collected by following the previously described procedure (Akter et al., 2020 (link)). In brief, each sample, collected by whirling a sterilized cotton bud into migratory bird’s dropping, was shifted to a separate sterilized zip lock bag with a particular tag number and directly transferred to the laboratory maintaining a cool chain. Each transferred sample was divided into two parts: one for seeding into 5 ml nutrient broth (HiMedia, India) and the other for taking into 5 ml Brain heart infusion (BHI) broth (Becton, Dickinson and Company, USA) containing 2% NaCl. All the broth containing sterile test tubes were then incubated aerobically for 18–24 h at 37 °C for the enrichment of the bacteria.
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