All materials were used without further purification unless otherwise noted. Tannic acid, vancomycin HCl (hereafter referred to as vancomycin unless otherwise noted), and poly-l -lysine (PLL, 30–70 kDa) were obtained from Sigma-Aldrich (St. Louis, MO). Poly(β-l -malic acid) (PMLA, 40 kDa) was cultured and purified as previously described from Physarum polycephalum.13 Bovine thrombin (high purity) was obtained from Biopharm Laboratories (Bluffdale, UT). Cation-adjusted Mueller Hinton broth (CaMHB) was obtained from Becton Dickenson (Franklin Lakes, NJ). All other chemicals unless otherwise noted, were obtained from Sigma-Aldrich (St. Louis, MO).
Cation adjusted mueller hinton broth
Manufactured by BD
Sourced in United States, France
Cation-adjusted Mueller Hinton broth is a standard culture medium used for antimicrobial susceptibility testing. It provides a consistent and reproducible growth environment for a variety of microorganisms, enabling accurate assessment of their antibiotic sensitivity profiles.
Automatically generated - may contain errors
Lab products found in correlation
Vitek 2 identification system, by bioMérieux (2 mentions)
Poly l lysine (pll), by Merck Group (1 mentions)
Tannic acid, by Merck Group (1 mentions)
Vancomycin hcl, by Merck Group (1 mentions)
Sepharose cl 4b, by GE Healthcare (1 mentions)
Mycophenolic acid, by Merck Group (1 mentions)
Adult bovine serum, by Sartorius (1 mentions)
Gc medium base, by BD (1 mentions)
Tryptic soy agar plates, by Liofilchem (1 mentions)
Sodium chloride (nacl), by Thermo Fisher Scientific (1 mentions)
Ethylenediaminetetraacetic acid, by Thermo Fisher Scientific (1 mentions)
Magnesium chloride, by Merck Group (1 mentions)
3 n morpholino propanesulfonic acid mops, by Merck Group (1 mentions)
Caco 2, by American Type Culture Collection (1 mentions)
Rpmi 1640, by Thermo Fisher Scientific (1 mentions)
Yeast extract, by Merck Group (1 mentions)
Phosphate buffered saline (pbs), by Thermo Fisher Scientific (1 mentions)
Brain heart infusion broth, by BD (1 mentions)
Hemin, by Merck Group (1 mentions)
L cysteine, by Merck Group (1 mentions)
11 protocols using cation adjusted mueller hinton broth
1
Tannic Acid-Vancomycin Antimicrobial Formulation
2
Mupirocin Nanoformulation Development and Evaluation
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Mupirocin was received from Teva (Debrecen, Hungary); hydroxy-propyl β-cyclodextrin (HPCD) from Roquette Frères (Lestrem, France); hydrogenated soy phosphatidylcholine (HSPC), 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-2000 Da] (mPEG DSPE) and cholesterol from Lipoid GmbH (Ludwigshafen, Germany); Sepharose CL-4B from GE Healthcare (Little Chalfont, UK); mycophenolic acid from Sigma-Aldrich (Saint Louis, MO, USA); adult bovine serum from Biological Industries (Beit Haemek, Israel); LC/MS-grade acetonitrile (ACN), methanol (MeOH) and water from Biolab Ltd. (Jerusalem, Israel); formic acid (FA) from J.T. Baker (Phillipsburg, NJ, USA). Analytical solvents were HPLC grade; other chemicals were commercial reagent grade.
Tryptic Soy Agar (TSA) plates containing 5% sheep blood and Chocolate Agar plates were from Liofilchem (Roseto degli Abruzzi, Italy). Cation-adjusted Mueller-Hinton broth (CA-MHB) and GC medium base were from Becton Dickinson (Franklin Lakes, NJ, USA). CA-MHB with TES (2-[[1,3-dihydroxy-2-(hydroxymethyl)propan-2-yl]amino]ethanesulfonic acid) was from TREK Diagnostic Systems (East Grinstead, UK).
Tryptic Soy Agar (TSA) plates containing 5% sheep blood and Chocolate Agar plates were from Liofilchem (Roseto degli Abruzzi, Italy). Cation-adjusted Mueller-Hinton broth (CA-MHB) and GC medium base were from Becton Dickinson (Franklin Lakes, NJ, USA). CA-MHB with TES (2-[[1,3-dihydroxy-2-(hydroxymethyl)propan-2-yl]amino]ethanesulfonic acid) was from TREK Diagnostic Systems (East Grinstead, UK).
3
Comparative Analysis of S. pneumoniae Susceptibility
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For S. pneumoniae, a comparative analysis was conducted between results observed for routine, conventional methods with data from the broth microdilution (BMD) method. Only blood isolates were included in this study. This work was performed with the collaboration of the SIREVA/GIVEBPVac (Grupo Interinstitutional para la Vigilancia de Enfermedades Bacterianas Prevenibles por Vacunación, in Spanish) network. Susceptibility testing was performed by BMD following the guidelines of the CLSI (2020) . S. pneumoniae ATCC 49619 was used as a control strain. Cation-adjusted Mueller Hinton broth (Becton, Dickinson, MD, USA), which had lysed horse blood added to a final concentration of 5%, was used as a medium.
4
Antibacterial Susceptibility Testing Protocol
5
Vancomycin-Resistant Enterococci Antibiotic Susceptibility
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Blood cultures were processed by the clinical microbiology laboratory. VRE was identified using the VITEK 2 identification system (bioMérieux Inc., La Balme les Grottes, France). Vancomycin resistance was defined as a MIC of at least 32 mg/L. VRE isolates collected at the two hospitals were stored at −80 °C until use. MIC tests of all available isolates were performed at the NTUH. The DAP MICs of enterococci were determined using the broth microdilution method and interpreted according to the Clinical and Laboratory Standards Institute20 . Cation-adjusted Mueller-Hinton broth (Becton Dickinson, Le Pont-de-Claix, France), with 50 μg/mL of supplemented calcium, was used for these tests.
6
Bacterial and Fungal Strain Characterization
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The bacterial and fungal strains (S2 Table in S1 File ) used in this study were clinical isolates obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA), the Centers for Disease Control and Prevention (CDC) and Biodefense and Emerging Infections Research Resources Repository (BEI Resources) (Manassas, VA, USA). RPMI 1640 (Thermo Fisher Scientific, Waltham, MA), YPD broth, YPD agar, cation adjusted Mueller-Hinton broth, brain heart infusion broth, and lactobacilli MRS broth (Becton, Dickinson and Company, Franklin Lakes, NJ) were purchased from commercial vendors. Phosphate buffered saline was purchased from Fisher Scientific (Waltham, MA). Yeast extract, L-cysteine, vitamin K, 3-(N-Morpholino)propanesulfonic acid (MOPS) and hemin were obtained from Sigma-Aldrich (St. Louis, MO). Human colorectal adenocarcinoma epithelial cells (Caco-2) (ATCC HTB-37), and monkey kidney epithelial cells (Vero) (ATCC CCL-81-VHG) were obtained from the American Type Culture Collection (ATCC) (Manassas, VA, USA).
7
Identification and Antibiotic Susceptibility of Enterococcus
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The blood cultures were processed at the clinical microbiology laboratory. Before 2017, Enterococcus spp. were identified using the VITEK 2 identification system (bioMérieux, Marcy l'Etoile, France). Subsequently, Enterococcus spp. were identified using matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (Bruker Biotyper system, MicroFlex LT; Bruker Daltonik GmbH, Bremen, Germany). Vancomycin-resistant enterococci were defined as Enterococcus isolates with a vancomycin MIC ≥ 32 mg/l. The MIC of daptomycin was determined by the broth microdilution method. Cation-Adjusted Mueller-Hinton Broth (Becton Dickinson, Le Pont-de-Claix, France) supplemented with 50 mg/l calcium was used. The fosfomycin MIC was determined using the agar dilution method in a medium supplemented with 25 mg/l glucose-6-phosphate. MIC for fosfomycin was interpreted based on the criteria for Enterococcus faecalis urinary tract isolates [24 ].
8
Bacterial Strains and Cell Line Protocol
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Neisseria gonorrhoeae clinical isolates (Table S1 ) used in this study were obtained from the CDC. S. aureus, MRSA, E. faecalis, E. faecium, and L. monocytogenes strains were obtained from the American Type Culture Collection (ATCC). E. coli BW25113 and JW25113 were obtained from the Coli Genetic Stock Center (CGSC), Yale University, USA. Media and reagents were purchased from commercial vendors: Brucella broth, chocolate II agar, cation-adjusted Mueller Hinton broth, tryptic soy broth (TSB), and tryptic soy agar (TSA) (Becton, Dickinson and Company, Cockeysville, MD, USA); yeast extract and dextrose (Fisher Bioreagents, Fairlawn, NJ, USA), proteose-peptone, nicotinamide adenine dinucleotide (NAD), agarose and tetracycline (Sigma-Aldrich, St. Louis, MO, USA); hematin, Tween 80, pyridoxal, linezolid and gentamicin sulfate (Chem-Impex International, Wood Dale, IL, USA); Dulbecco’s Modified Eagle Medium (DMEM), fetal bovine serum (FBS) and phosphate-buffered saline (PBS) (Corning, Manassas, VA, USA); and azithromycin (TCI America, Portland, OR, USA). Human colorectal adenocarcinoma epithelial cells (Caco-2) (ATCC HTB-37) was obtained from the American Type Culture Collection (ATCC) (Manassas, VA, USA). Compounds were synthesized from commercial sources in our laboratory.
9
Bacterial Genome Sequencing Workflow
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All bacterial isolates were grown on cation-adjusted Mueller-Hinton II agar (Becton-Dickinson) for 16 h at 37 °C, and sub-cultured into cation-adjusted Mueller-Hinton broth (Becton-Dickinson) for a further 16 h at 37 °C. Bacterial genomic DNA was extracted from liquid culture using the GenFind V3 Reagent Kit (Beckman Coulter) as per manufacturer’s instructions. Libraries for short read sequencing were prepared using the Nextera Flex DNA Library Prep Kit (Illumina), and 150 bp paired-end sequencing was performed on the NovaSeq 6000 system (Illumina). Libraries for long-read sequencing were prepared using the Ligation Sequencing Kit with Native Barcoding Expansion (Oxford Nanopore Technologies) and sequenced on the MinION instrument with an R9.4.1 flow cell (Oxford Nanopore Technologies) for 48 h. Basecalling was performed with Guppy v.4.0.14 using the ‘high accuracy’ basecalling model.
10
Tebipenem and Ertapenem Susceptibility Assay
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Tebipenem was provided by Spero Therapeutics (Cambridge, MA, USA). Ertapenem was purchased from Henry Schein (Melville, NY, USA). The challenge isolates utilized in these studies were either purchased from JMI Laboratories (North Liberty, IA) or provided by the National Collection of Type Cultures (NCTC). The challenge panel included five E. coli isolates that were selected based upon their multilocus sequence type (e.g., ST131), resistance mechanisms (e.g., ESBL production), and MIC values for tebipenem and ertapenem and confirmed resistance to other antibiotics (e.g., fluoroquinolone and TMP-SMX). Susceptibility studies were performed in accordance with the guidelines of the Clinical and Laboratory Standards Institute (21 ) and completed in triplicate over a 2-day period. The final MIC values were reported as modal values. Cation-adjusted Mueller-Hinton broth (BD Laboratories, Franklin Lakes, NJ, USA) was utilized in all susceptibility studies.
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