Three serogroups of Shiga toxin-producing E. coli were used in this study. E. coli O157:H7 H1730 was a human isolate from a lettuce outbreak and E. coli O157:H7 (ATCC 43895) was from a 1982 ground beef outbreak. Both isolates were obtained from the culture collection at the Center for Food Safety, University of Georgia. E. coli O121:H19 (strain TW08980) and E. coli O26:H11 (strain 3012-03) were from a 2016 Missouri flour outbreak and obtained from the Michigan State University STEC Center. Isolates were revived from frozen storage by transferring to Tryptic Soy Broth (TSB, Neogen, Lansing, MI, USA) and incubating at 37°C for 24 h. The bacterial strains were evaluated for antibiotic resistance to ampicillin and streptomycin by streaking on Tryptic Soy Agar (TSA; Neogen Lansing, MI, USA) amended with 100 μg/ml of ampicillin, 100 μg/ml of streptomycin, and a combination of 100 μg/ml streptomycin and ampicillin. Plates were placed at an incubation temperature of 37°C for 24 h and observed for colony formation. Cultures that did not yield colonies in the antibiotic amended plates were considered to be susceptible to the antibiotics (Table 1 and Figure 1 ).
Tryptic soy agar tsa
Manufactured by Neogen
Sourced in United States
Tryptic Soy Agar (TSA) is a general-purpose microbiological growth medium. It is composed of tryptone, soy peptone, sodium chloride, and agar. TSA supports the growth of a wide range of microorganisms, including bacteria and fungi.
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Lab products found in correlation
Tryptic soy broth tsb, by Neogen (3 mentions)
Ethanol, by Chempur (1 mentions)
Chloroform, by Chempur (1 mentions)
Isopropanol, by Chempur (1 mentions)
Dichloromethane, by Chempur (1 mentions)
Ciprofloxacin, by Merck Group (1 mentions)
Mueller hinton broth (mhb), by Lab M (1 mentions)
Tobramycin, by Tokyo Chemical Industry (1 mentions)
Ultrasonic cleaner 3800, by Emerson (1 mentions)
Ferric ammonium citrate, by Merck Group (1 mentions)
Sodium thiosulfate, by Thermo Fisher Scientific (1 mentions)
5 protocols using tryptic soy agar tsa
1
Antibiotic Resistance Profiling of Shiga Toxin-Producing E. coli
2
Synthesis and Characterization of PCL Polymers
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PCL pellets (Mn 80,000, cat no.: 440744-500G) were obtained from Sigma Aldrich (Gillingham, U.K.), GS, USP grade was obtained from Fluorochem Ltd. (Glossop, U.K.). Solvents used such as dichloromethane (DCM), chloroform, ethanol, and iso-propanol, and components to prepare the phthaldialdehyde reagent were obtained from Chempur (Piekary Śląskie, Poland). Tryptic Soy Broth (TSB) and tryptic soy agar (TSA) were obtained from Neogen® (Lansing, MI, USA).
3
Antimicrobial Testing of P. aeruginosa Isolates
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Ten P. aeruginosa isolates were included in this study: three CF reference isolates AA2 (LMG 27630), DK2 (LMG 27626) and LESB58 (LMG 27622) [21 (link)], a hyper-biofilm forming isolate (CF127) [22 (link)] and six isolates recovered from chronically colonized CF patients receiving care at the Ghent University Hospital (this study was approved by the Ethics Committee of Ghent University Hospital, registration number B670201836204). Pure cultures were obtained by growth on Tryptic Soy Agar (TSA, Neogen, Heywood, UK) for 24 h at 37 °C. Overnight cultures were grown in Mueller Hinton Broth (MHB, Lab M, Moss Hall, UK) for 16 h at 37 °C. The following antibiotics were used: tobramycin (TCI Europe, Zwijndrecht, Belgium), ciprofloxacin (Merck Life Science, Darmstadt, Germany) and colistin sulphate (TCI Europe). For tobramycin and colistin, stock solutions of 5 mg/mL were prepared and for ciprofloxacin a stock solution of 3.2 mg/mL with 70 μL 1 M HCl was prepared. All stock solutions were prepared in MilliQ water, filter sterilized (PES, 0.22 μm, VWR, Haasrode, Belgium) and stored at 4–7 °C for maximum one week prior to use. SCFM2 was prepared as described previously [18 (link)], with the modification that mucin was autoclaved first instead of sterilized by UV exposure.
4
Quantification of Bacterial Biofilms on Titanium
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Titanium coupons were washed with TBS to remove remaining planktonic cells and then they were transferred into Falcon tubes containing 1 mL of 0.5% (w/v) Tween® 20-TSB solution. After that, the tubes were sonicated in an Ultrasonic Cleaner 3800 water sonicator (Branson Ultrasonics, Danbury, CT, USA) at 25 °C, for 5 min at 35 kHz. The tubes were mixed vigorously for 20 s prior to and after the sonication step. Serial dilutions were performed from the resulting bacterial suspensions and plated on Tryptic Soy Agar (TSA, Neogen®, Lansing, MI, USA) plates.
5
Salmonella Strain Preparation and Verification
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The strains of Salmonella enterica subsp. enterica serovars used in this research were S. Typhimurium E2009005811, S. Tennessee E200700502 and S. Agona. The first two strains were provided by the Minnesota Department of Health; they were isolated from patients linked to the S. Typhimurium E2009005811 and S. Tennessee E200700502 peanut butter outbreaks of 2009 and 2007, respectively [8 ,10 ]. The S. Agona strain was originally isolated from an outbreak related to toasted oats cereal from 1998 [6 ]. The stock cultures of the three serovars were stored in a 1:1 ratio of glycerol and tryptic soy broth (TSB; Neogen, Inc., East Lansing, MI, USA) at −55 °C. The working cultures of each serovar were prepared from frozen stocks and inoculated into TSB, grown overnight at 37 °C and then stored at 4 °C. In order to test the working stocks, they were re-transferred once a week and streaked onto tryptic soy agar (TSA; Neogen, Inc.) containing 0.8 g/L ferric ammonium citrate (Sigma-Aldrich™, St. Louis, MO, USA) and 6.8 g/L sodium thiosulfate. (Acros Organics, Morris Plains, NJ, USA). This medium was formulated to provide a differential but non-selective agar. Periodically, the serovars were also streaked onto bismuth sulfate agar (Neogen, Inc.) and xylose lysine deoxycholate agar (Neogen, Inc.).
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