Masters Theses

Date of Award

5-2000

Degree Type

Thesis

Degree Name

Master of Science

Major

Food Science and Technology

Major Professor

David A. Golden

Committee Members

F. Ann Draughon, P. Michael Davidson

Abstract

Recovery of Escherichia coli 0157:H7 and Yersinia enterocolitica from farm samples (feed, bedding, fecal samples, and water) from various animal sources (cattle, swine, and chickens) was investigated at inoculum levels of 104CFU/g (high level) and 10 CFU/g (low level) of sample. High-inoculum levels of E. coli 0157:H7 were recovered using direct plating on sorbitol MacConkey agar (SMAC), SMAC supplemented with cefixime and tellurite (CT-SMAC), hemorrhagic colitis agar (HC), modified eosin methylene blue agar (MEMB), and Rainbow ® agar 0157. Low-inoculum levels of E. coli 0157:H7 were recovered using a 24-hr enrichment at 37°C in modified tryptic soy broth supplemented with 0.02 g/1 novobiocin (mTSB+N), EHEC enrichment broth (EEB), and modified E. coli broth supplemented with 0.02 g/1 novobiocin (mEC+N) followed by streaking on to SMAC, CT-SMAC, HC, MEMB, and Rainbow ® agar 0157. In addition, low-inoculum levels of E. coli 0157:H7 were detected using immunomagnetic separation (IMS), EHEC-Tek ®, and Reveal ® for E. coli 0157:H7 test systems. High-inoculum levels of Y. enterocolitica were recovered using MacConkey agar (MAC), cefsulodin-irgasan-novobiocin agar (CIN), and KV-202 agar. Low- inoculum levels of Y. enterocoliticawere recovered using cold enrichment in peptone sorbitol bile broth (PSBB) followed by streaking on to MAC, CIN, and KV-202.

Eight sample types were evaluated. For direct plating, three repetitions were performed, giving a total of 24 (8x3) inoculated samples tested. Duplicate sample types were examined for low inoculum evaluations, also performed in triplicate, for a total of48 (8x2x3) individual samples evaluated. Rainbow ® agar 0157 recovered high-inoculum levels of E. coli 0157:H7 by direct plating from the most sample types (7 of 8), but provided the poorest numerical recovery in 6 of 7 sample types from which the organism was Recovered (PHigh-inoculum levels of Y. enterocolitica were recovered from the most sample types with CIN (7 of 8), followed by KV-202 (6 of 8), and MAC (5 of 8). CIN agar provided superior numerical recovery over KV-202 in TMR, pig fecal, chicken fecal, and pig feed sample types (P<0.05), but test media did not differ significantly in their ability to recover the organism from the pig feed sample type (46 individual samples evaluated) (P<0.05). After enrichment, streaking to CIN provided the best recovery (19 of 46), followed by MAC (13 of 46), and KV-202 (11 of 46).

These results indicate that some direct plating methods used for isolation of E.coli 0157:H7 and Y. enterocolitica from foods may be satisfactory for recovery of high levels of these pathogens from farm animal environments. While enrichment alone did not allow for consistent recovery of low levels of E. coli 0157:H7 from such environments, EHEC-Tek ® and Reveal ® for E. coli 0157:H7 test systems provide good recovery (18 and 20/24 samples respectively). Cold enrichment in PSBB is an unreliable method for recovery of low inocula of Y. enterocoliticafrom farm animal environments.

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