|BIERNBAUM, ERIKA - Oak Ridge Institute For Science And Education (ORISE)|
|CASEY, THOMAS - Retired ARS Employee|
Submitted to: PLOS ONE
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/13/2022
Publication Date: 5/26/2022
Citation: Kudva, I.T., Trachsel, J.M., Biernbaum, E.N., Casey, T. 2022. Novel reusable animal model for comparative evaluation of in vivo growth and protein-expression of Escherichia coli O157 strains in the bovine rumen. PLoS ONE. 17(5). Article e0268645. https://doi.org/10.1371/journal.pone.0268645.
Interpretive Summary: Cattle are carriers of the foodborne, human pathogen Escherichia coli O157:H7 (O157). Understanding how O157 survives within the rumen of cattle, which is the first compartment of the gastrointestinal tract in this animal, could help identify possible targets for O157 control. However, lack of a method that would allow direct recovery of bacteria being tested from the rumen has limited such studies. In fact, most studies focus either on evaluating bacteria in rumen fluid in the laboratory or introduce bacteria into the rumen and recover the same only after the animal sheds the bacteria in feces. In case of latter, when bacteria being tested and therefore shed by the animal are human pathogens the animal is usually euthanized at end of study. Hence, to permit reuse of animals and allow recovery of bacteria being tested directly from the rumen, we developed a novel, reusable, rumen-fistulated cattle model. This animal model allows simultaneous evaluation of multiple bacteria within the bovine rumen in a non-terminal manner. Using this animal model, we tested three different strains of O157 in rumens of cattle fed either a high fiber maintenance or high protein lactation diets. These diets are commonly provided to dairy cattle, and we sought to determine if the changes in the rumen fluid brought about by these diets would alter growth of O157. We observed that some O157 strains did not thrive well in the acidic rumen fluid resulting from the lactation diet. Closer analysis of the proteins being produced by the O157 strains indicated that the differences in protein expression were mainly influenced by the animal’s diet and growth conditions, irrespective of the O157 strain being tested. The identified proteins, as produced by the O157 strains in the bovine rumen, help the bacteria to adapt to the rumen and could be targeted to control O157 in cattle.
Technical Abstract: Previously, we had demonstrated that Escherichia coli O157:H7 (O157) strain 86-24 expresses proteins involved in survival rather than virulence in vitro in rumen fluid from dairy cattle on the maintenance diet. Here, we verified if this observation would be true for different O157 strains grown in vitro in rumen fluid from, and in vivo in the rumen of, animals on contrasting maintenance (high fiber) and lactation (high protein) diets usually fed to dairy cattle. For the in vivo studies, an economical, novel, reusable and non-terminal rumen-fistulated animal model permitting simultaneous evaluation of multiple bacterial strains in the bovine rumen was developed. All experiments were conducted in duplicate using different animals to account for host-related variations. The O157 strains included, 86-24, EDL933 and the super shed SS-17; E. coli NalR (#5735), derived from a bovine intestinal commensal E. coli, was included as a control. As expected, diet influenced ruminal pH and volatile fatty acid (VFA) composition. The pH ranged from 6.2 - 7.0 and total VFA concentrations from 109 - 141 µM/ml, in animals fed the maintenance diet. In comparison, animals fed the lactation diet had a ruminal pH ranging between 5.18 – 6.0, and total VFA of 125 - 219 µM/ml. Strain dependent differences in O157 growth was observed in the rumen fluid/rumen of cattle fed either diet, both in vitro and in vivo, with O157 strains 86-24 and EDL933 demonstrating similar growth patterns. Analysis of the O157 proteomes expressed in the rumen fluid/rumen verified previous observations of adaptive responses. Any difference in the adaptive response was mainly influenced by the animal’s diet and growth conditions (in vitro and in vivo) and not the O157 strain. These new insights into the O157 responses could help formulate modalities to control O157 across strains in cattle at all stages of husbandry. Importance of this study: This study demonstrates that O157, irrespective of strain, have similar differences in growth and protein expression patterns in rumen fluid with variations being influenced primarily by rumen fluid composition and the in vitro or in vivo environment. Hence, evaluating microbes within the host is critical when seeking to understand colonization dynamics and identifying targets for control strategies. Most importantly, the method presented here for conducting such in vivo studies in the rumen will permit reuse of the animal and allow for multiple evaluations.