Mehall, Lindsey Nicole (2015-12). Evaluation of the Appropriateness of Previously Developed Escherichia coli Biotype I Surrogates as Predictors of Non-O157:H7 Shiga Toxin-Producing E. coli in Beef Processing. Doctoral Dissertation.
Thesis
Non-O157 Shiga-toxin producing Escherichia coli (STECs) serovars O145:NM, O45:H2, O26:H11, O103:H11, O111 (organism was originally ordered from American Type Culture Collection (ATCC) under accession # BAA-2217, and has since then been reclassified as O147, see Appendix), O121:H19, and E. coli biotype I surrogates were individually cultured in tryptic soy broth (TSB) at 37?C for 18 hours. Rifampicin-resistant (rif-resistant) surrogates were compared to the parent strains listed above. Stationary phase and acid-adapted organisms were each transferred into phosphate buffer saline (PBS) acidified with L-lactic acid at pH 2.5, 3.0, and 3.5, and enumerated for survivors following a 2-hour exposure time. In order to construct a thermal destruction curve, organisms were transferred into a capillary tube, flame sealed, submerged in a water bath at 55, 60 and 65 +- 0.5?C, and enumerated for survivors. For freezing (-20 +- 0.5?C) and refrigerated (4 +- 0.5?C) storage, bacterial strains were enumerated on days 0, 7, 14, 21, 28, 60, and 90 to determine the response to freezing and refrigerated storage. Growth curves of E. coli biotype I surrogates (parent and rif-resistant) were similar to those of STECs throughout the evaluation. For acid resistance, acid-adapted organisms at pH 3.0 showed initial log reductions (CFU/ml) ranging from 1.7-2.5, where organism O26:H11 had the greatest log reduction (2.5). After 2 hours exposure time, reductions ranged from 5.1-7.4 log CFU/ml. D-values were calculated for each organism at 55, 60 and 65 +- 0.5?C. Acid-adapted organisms at 65?C had D-values ranging from 0.13-0.64 min, with a rif-resistant E. coli biotype I surrogate (BAA-1429 rif) having the highest D-value at this temperature. For the response to refrigeration and freezing temperatures, there were no notable trends or patterns observed, and no one single E. coli biotype I surrogate represented all of the non-O157 STECs. Organisms were analyzed individually and in sets (surrogates, rif-resistant surrogates and STECs) to represent a mean. Analyzing the organisms in sets eliminated certain individual strain-to-strain variation, and showed fewer differences (P < 0.05). Surrogates analyzed as a mean of the five strains indicate they may be best utilized in combination to represent all six of the non-O157 STECs.
