Displacement of Escherichia coli O157:H7 from Rumen Medium Containing Prebiotic Sugars

Abstract

A fed-batch, anaerobic culture system was developed to assess the behavior of Escherichia coli O157:H7 in a rumen-like environment. Fermentation medium consisted of either 50% (vol/vol) raw or sterile rumen fluid and 50% phosphate buffer. Additional rumen fluid was added twice per day, and samples were removed three times per day to simulate the exiting of digesta and microbes from the rumen environment under typical feeding regimens. With both types of medium, anaerobic and enteric bacteria reached 10 10 and 10 4 cells/ml, respectively, and were maintained at these levels for at least 5 days. When a rifampin-resistant strain of E. coli O157:H7 was inoculated into medium containing raw rumen fluid, growth did not occur. In contrast, when this strain was added to sterile rumen fluid medium, cell densities increased from 10 6 to 10 9 CFU/ml within 24 h. Most strains of E. coli O157:H7 are unable to ferment sorbitol; therefore, we assessed whether the addition of sorbitol as the only added carbohydrate could be used to competitively exclude E. coli O157:H7 from the culture system. When inoculated into raw rumen broth containing 3 g of sorbitol per liter, E. coli O157:H7 was displaced within 72 h. The addition of other competitive sugars, such as L-arabinose, trehalose, and rhamnose, to rumen medium gave similar results. However, whenever E. coli O157:H7 was grown in sterile rumen broth containing sorbitol, sorbitol-positive mutants appeared. These results suggest that a robust population of commensal ruminal microflora is required to invoke competitive exclusion of E. coli O157:H7 by the addition of "nonfermentable" sugars and that this approach may be effective as a preharvest strategy for reducing carriage of E. coli O157:H7 in the rumen.The emergence of Escherichia coli O157:H7 as a human pathogen has led to renewed interest in the ecology of this organism in the bovine rumen and gastrointestinal tract (19). The presence of E. coli O157:H7 and other Shiga toxin-producing bacterial strains in the bovine rumen has not been well studied, but recent studies indicate low prevalence rates (22) of these organisms as well as low cell densities in the rumen (28). The low pH and high concentration of volatile fatty acids (VFA) ordinarily seen with the grain-rich rations used by the cattle industry act to limit growth of enteric bacteria in the bovine rumen (3,25,27), and as a result, these bacteria are considered to represent a minor and rather transient population (11,14,18).However, despite the adverse growth conditions, the rumen is still widely considered to be an important reservoir of E. coli, and E. coli O157:H7 in particular, providing a continuous source of these bacteria into the intestinal tract of cattle (2,11,17). For these reasons, many of the strategies for decreasing the incidence of food-borne disease caused by E. coli O157:H7 are now focused on reducing carriage and shedding of the bacterium by cattle. Recently, changing the animals' diet from rolled corn (high starch) to alfalfa hay (low starch) was sho...

Biotic and Abiotic Factors Influencing In Vitro Growth of Escherichia coli O157:H7 in Ruminant Digestive Contents

“…We show that in raw rumen fluid, growth of E. coli O157:H7 is limited or even suppressed due to the presence of the resident microflora, which exerts a strong barrier effect. These data agree with previous observations made with continuous cultures in 50% diluted rumen fluid (12). Taken together, our results indicate that this barrier effect is due mostly to the observed increase in acetate concentration (acetate represents 65 to 75% of the total VFA mixture) and decrease in pH, resulting from the fermentative activity of the microflora during incubation, but also to other direct or indirect effects not yet identified.…”

“…Yet it is important to understand better how gut environmental conditions are likely to influence growth or survival of these potentially pathogenic bacteria. Most previous studies were done in sterile laboratory media or in diluted sterile rumen fluid supplemented with purified sugars or Trypticase (12,43,47,56). In this report, we describe original in vitro models for growth of EHEC strains in digestive contents directly harvested from ruminants.…”

Characterization of Haemorrhagic Enteritis in Dairy Goats and the Effectiveness of Probiotic and Prebiotic Applications in Alleviating Morbidity and Production Losses

“…In support of this role, the application of the prebiotic: probiotic mixture stopped symptoms in 50% of the symptomatic goats. The prebiotic, Celmanax ™ , contains significant mannose that is known to block FimH-mediated non-O157 STEC adherence to target cells [37] and in vitro studies indicated that it caused a higher magnitude of reduction in colonization for the non-O157 STEC compared with the O157 STEC [3]. The probiotic also produced a decrease in O157 and non-O157 STEC colonization, but the mechanism is not likely related to the presence of mannose.…”

Measurements of Fitness and Competition in CommensalEscherichia coliandE. coliO157:H7 Strains

“…Another six strains were obtained from the Karch laboratory in Germany. The commensal strains included 31 E. coli reference strains (37) and 8 cattle strains from the culture collections at the University of Wisconsin and University of Nebraska (17). The E. coli O157:H7 portion of the set was designed to represent the genetic and geographic diversity of this serotype in the United States, while the commensal E. coli portion of the set was designed to represent the entire genetic diversity of the species (54).…”