Strategies for recovering of planktonic and sessile cells of Escherichia coli O157:H7 from freshwater environment.

Abstract

The experiments were performed with Escherichia coli O157:H7 EDL 933 in freshwater microcosms at 12°C. At 35, 45, and 70days, samples were taken and filtered through 0.45 μm membrane filters. The following alternatives were tested to evaluate the recovery percentage of injured cells: (1) selective media CHROMagar(™)O157 and chromID(™)O157:H7 agar, at 37°C for 24h; (2) tryptic soy agar supplemented with yeast extract (TSAE), incubated at 25°C for 2 or 4h, then transferred to CHROMagar(™)O157 or chromID(™)O157:H7 agar at 37°C (TSAE2h-CHROM, TSAE4h-CHROM and TSAE2h-ID, TSAE4h-ID); (3) thin agar layer (TAL) method, TSAE was overlaid on CHROMagar(™)O157 or chromID(™)O157:H7 agar (TALCHROM and TALID, respectively) and incubated at 37°C for 24h; and (4) TALCHROM at 25°C for 4h, then continued up to complete 24h at 37°C (TALCHROM4h). Furthermore, the recovery of E. coli O157:H7 cells adhering to glass coverslips were evaluated to mimic biofilm conditions. The recovery percentages obtained from each alternative were calculated relative to TSAE counts. After 70days, TSAE4h-CHROM and TALCHROM4h showed the highest recovery percentage (>90%) from water microcosms. Despite the improved recovery of cell adhering to glass surfaces, the percentages obtained with TSAE4h-CHROM were low. Further studies for the recovery of biofilm-forming E. coli O157:H7 are required. Pre-incubation on TSAE at 25°C for 4h, combined with CHROMagar(™)O157, or by thin agar layer method (TALCHROM) enhanced significantly the recovery of viable cells of E. coli O157:H7 after prolonged stay in water microcosms.