Survival of Escherichia coli O157 in abattoir waste products.

AIMS: This study monitored survival and growth of Escherichia coli O157 in ovine and bovine abattoir waste. METHODS: Blood and gut contents were inoculated separately with cocktails of E. coli O157. Samples were stored aerophilically and microaerophilically at 5 degrees C, 15 degrees C and 30 degrees C to represent storage at different container depths and at extremes of UK ambient temperature. CONCLUSIONS: Results showed survival of E. coli O157 was irrespective of oxygen content with no significant differences observed between aerophilic and microaerophilic environments. Numbers of E. coli O157 in ovine and bovine gut contents showed no change when stored at 5 degrees C and increased 1-2 log(10) at 15 degrees C and 30 degrees C in 28 h. In ovine and bovine blood, irrespective of storage temperature, there was a 0.5-2 log(10) reduction or no change in numbers except in ovine blood stored at 30 degrees C where the fall in numbers was followed by a 3 log(10) increase. In aged (stored at 4 degrees C for 18 h before spiking) bovine blood there was no significant change in numbers at 5 degrees C while at 15 degrees C there was 2 log(10) rise after 48 h. At 30 degrees C there was an initial 1 log(10) decrease in numbers followed by a 1 log(10) rise over the following 40 h. SIGNIFICANCE AND IMPACT OF STUDY: Abattoir wastes may become contaminated from animals infected with Verocytotoxigenic E. coli O157 and in certain storage conditions these pathogens could significantly increase in numbers. There is need for care in abattoir waste disposal, not only for personnel subject to direct contact, but also in the prevention of cross contamination to adjacent land and water courses which could indirectly infect humans.

Optimizing enrichment conditions for the isolation of Escherichia coli O157 in soils by immunomagnetic separation.

AIMS: To compare a range of enrichment broths and enrichment temperatures for the isolation of Escherichia coli O157 by immunomagnetic separation (IMS) from sandy, loam and clay soils. METHODS AND RESULTS: Soils were spiked with cocktails of four atoxigenic strains of E. coli O157 and four strains of commensal E. coli. The organisms were stressed by subjecting soils to cycles of freeze/thawing, followed by drying at 20 degrees C for up to 4 days. Nine enrichment broths were trialled based on buffered peptone water, tryptone soya broths and EC broths supplemented with a range of selective additions. Enrichments were incubated for 6 h and assessed by target recovery after IMS on cefixime tellurite sorbitol MacConkey agar (CTSMAC) incubated at 37 degrees C for 24 h. A comparison of enrichment temperatures (37 and 42 degrees C) was also performed. Buffered peptone water (with or without vancomycin) and tryptone soya broth (with or without novobiocin) gave significant increases in recovery of E. coli O157 compared to others tested. In addition, broths incubated at 42 degrees C were superior to those at 37 degrees C for the recovery of E. coli O157. SIGNIFICANCE AND IMPACT OF THE STUDY: This study showed that sub-lethally damaged E. coli O157 surviving in soil can be sensitive to antimicrobial additions. The choice and concentration of these additions is vitally important to optimize target recovery. Some IMS protocols, established for the isolation of E. coli O157, may be unsuitable for the examination of soil samples.

Long-term survival of Escherichia coli O157 on pasture following an outbreak associated with sheep at a scout camp.

AIMS: To monitor the decay of E. coli O157 in soil (loamy sand) on a scout campsite following an outbreak in humans. METHODS AND RESULTS: Samples of soil and sheep faeces were collected from the campsite and tested for the presence of E. coli O157 by immunomagnetic separation (IMS) after enrichment in buffered peptone water + vancomycin at 42 degrees C for 6 h. Enumeration of target was carried out by direct plating onto sorbitol MacConkey agar plates supplemented with cefixime and tellurite (CTSMAC) incubated at 37 degrees C for 24 h. Low numbers (< 100 g(-1)) were estimated by the most probable number (3-tube MPN) technique. CONCLUSIONS: Survival was observed for 15 weeks. SIGNIFICANCE AND IMPACT OF THE STUDY: A number of laboratory studies have followed the decay of E. coli O157 in soil, animal faeces and water. This study follows (for the first time) the decay of the organism in soil after an outbreak associated with sheep. It demonstrates the long-term persistence of the organism in the environment and the results will be potentially important in performing risk assessments for both human and animal infection.

The optimization of isolation media used in immunomagnetic separation methods for the detection of Escherichia coli O157 in foods.

AIMS: To compare media used in immunomagnetic separation (IMS) techniques for the isolation of Escherichia coli O157 from food. METHODS AND RESULTS: Foods, both naturally contaminated and spiked, with low numbers (< 1 g(-1)) of stressed E. coli O157 were enriched in media based on buffered peptone water (BPW), tryptone soya and EC broths incubated at 30, 37, 40 and 42 degrees C. Following immunomagnetic separation, beads were plated on a range of selective agars. CONCLUSION: BPW supplemented with vancomycin (8 mg l(-1)) incubated at 42 degrees C, followed by IMS and subsequent plating of immunobeads onto cefixime tellurite sorbitol MacConkey agar plus either Rainbow or CHROMagar agars, proved optimum for the recovery of spiked, stressed E. coli O157 in minced beef, cheese, apple juice and pepperoni. The same protocol was optimum for recovery from naturally-contaminated minced beef and cheese. SIGNIFICANCE AND IMPACT OF THE STUDY: The optimum protocol would increase isolation rates of E. coli O157 from foods.

Improved isolation of Escherichia coli O157 using large enrichment volumes for immunomagnetic separation.

The recovery of low numbers of Escherichia coli O157 in foods by immunomagnetic separation (IMS) was improved, on average, ninefold by increasing the enrichment volume tested from 1 to 10 ml while maintaining the volume of immunobeads constant at 0.02 ml. Although 50 ml volumes also gave improved recoveries (approximately threefold), the 50 ml volume cannot be recommended until a suitable magnetic separation apparatus has been developed. By testing 10 ml volumes, the improved sensitivity of the IMS procedure will reduce false-negative E. coli O157 tests and help improve epidemiological studies of this pathogen.