The influence of temperature on growth and verotoxin production by Escherichia coli strains was studied in brain heart infusion (BHI) broth both in shake cultures at various temperatures and in a temperature-gradient incubator. All strains of E. coli surveyed grew from at least 10 to 45°C, with some strains growing at 8° C. Verotoxin production (determined using the Vero cell–assay system) was a function of both temperature and time, with the highest titers produced at temperatures supporting the fastest growth (based on days to visible turbidity) and highest viable cell counts. However, for strains producing verotoxin, toxin production was detected at any temperature supporting growth. Three strains (of 16 tested) increased 1000-fold in viable count in 4 to 6 days at 10°C. The data presented here indicate that most E. coli strains surveyed can easily grow at ca. 10°C and thus suggest the potential for growth in temperature-abused refrigerated foods.
Two commercial serological kits (Oxoid BCET-RPLA and Tecra VIA) and a Chinese hamster ovary (CHO) cell cytotonicity assay for the detection of Bacillus cereus diarrhoeal enterotoxin were compared. Eleven B. cereus strains and one enterotoxigenic B. thuringiensis strain were evaluated. Both kits and the CHO cell assay yielded positive toxin responses for cell-free culture filtrates from eight out of 11 diarrhoeal enterotoxigenic strains. An emetic enterotoxin producing strain was negative with all three assays. Two B. cereus strains were negative using the BCET-RPLA kit, but positive with the Tecra VIA kit and CHO cell assay. The BCET-RPLA indicated significant levels of enterotoxin after samples were boiled, whereas the CHO cell and Tecra assays were negative. Overall, the cell culture assay was the most sensitive. However, the Tecra VIA kit provided similar results and was better suited for the rapid detection of B. cereus diarrhoeal enterotoxin.
Attachment of E. coli O157:H7 and E. coli K12 to beef tenderloin filet, chuck, and adipose tissues was studied. Most attachment occurred within 1 min of incubation; the number of attached organisms depended on the concentration of bacteria in the liquid inoculum. Similar levels of E. coli bound to the three types of beef tissues tested. E. coli O157:H7 was heavily piliated; however, there was no significant difference between levels of bound E. coli O157:H7 and E. coli K12, indicating that these surface structures apparently are not involved in attachment. Scanning electron photomicrographs of meat tissue and of purified collagen suggested that bacteria attached primarily to collagen fibers. Rinsing solutions consisting of 10% trisodium phosphate (TSP), 2% acetic acid (HAc), phosphate-buffered saline (PBS) and combinations of each were tested for effectiveness in reducing the number of attached E. coli. The level of bacteria removed from tenderloin tissue following TSP, HAc, or PBS rinses did not differ considerably. When beef tissues were stored at 4°C for 18 h after the various rinse combinations, TSP rinse treatments reduced the levels of E. coli K12 and O157:H7 attached to adipose tissue up to 3.4 and 2.7 log units, respectively, compared to PBS rinse treatments. Therefore, TSP may be effective for reducing populations of E. coli O157:H7 on beef carcass tissue.
The relationship between detection of heat-labile enterotoxin production by Bacillus cereus using the Oxoid BCET-RPLA kit and the production of a cytotonic response in HEp-2 and CHO cells was evaluated using 12 isolates. The strains were cultured in brain heart infusion + 0.1% glucose for 5 h at 37°C on a rotary shaker. Cell-free supernatants were prepared by filtration and tested using the two cell lines and the immunoassay. The primary biological response noted with the cell lines was cytotonicity, with the CHO cells being consistently more sensitive than the HEp-2 cells. While there was general agreement between the cell culture tests and the immunoassay, differences were noted for specific strains. Further, while activity in cell cultures was lost when cell-free supernatants were heated to 80°C for 5 min, the BCET-RPLA kits continued to detect significant levels of enterotoxin. The results suggest that confirmation of B. cereus enterotoxin using a suitable biological response may be warranted until additional research can clarify the specificity of the BCET-RPLA assay.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.