Twenty-five grams of each samples was homogenized with 225 mL of sterilized peptone water. 1 mL aliquots of the homogenate was inoculated into 15 mL of Violet Red Bile Lactose Agar (Biokar Diagnostics, France) then incubated for 24 h at 37°C. The presumptive colonies were streaked into eosin methylene blue agar (Biokar Diagnostics, France). E. coli identification was based on morphological, microscopic characteristics and biochemical assays, including gram straining, oxidase, indole, fermentation, citrate degradation, then confirmed by API 20E kit (BioMerieux, France).
Eosin methylene blue agar
Manufactured by Solabia
Sourced in France
Eosin methylene blue agar is a selective and differential culture medium used for the isolation and identification of Gram-negative enteric bacteria, particularly Escherichia coli and other coliforms. It contains eosin and methylene blue as dyes, which inhibit the growth of Gram-positive bacteria and allow the differentiation of lactose-fermenting and non-lactose-fermenting Gram-negative bacteria based on colony color.
Automatically generated - may contain errors
2 protocols using eosin methylene blue agar
1
Quantitative Enumeration of E. coli in Samples
2
Isolation of E. coli from Faecal Samples
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One gram of homogenised faecal sample was added to 10 mL of sterile buffered peptone water (Biokar diagnostics) and plated onto SuperPolymyxin medium [19 (link)], an eosin methylene blue agar (Biokar diagnostics) supplemented with 3.5 μg/ml colistin (Sigma-Aldrich), 10 μg/ml daptomycin (Glentham Life Sciences), and 5 μg/ml amphotericin B (Glentham Life Sciences), and incubated for 24 h at 37°C. For each faecal sample, up to five colonies with a E. coli phenotype were isolated (i.e. metallic green sheen in SuperPolymyxin medium).
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