(P27-045-24) An Investigation of Honey’s Effects on Small Intestine Microbial Composition and Virulence Expression of the Foodborne Pathogen Enterotoxigenic E. coli

Objectives: Previously, our lab has shown that the foodborne pathogen enterotoxigenic E. coli (ETEC) displayed a significant decrease in survival in an in vitro gastric digestion with honey, as compared to a simple sugar control. However, this effect did not persist or prove to be protective in an in vitro small intestine fermentation in the presence of a mock community of small intestine microbial organisms (SIMO). Here, we build on this existing work to examine changes in microbial community composition, as well as abundance of the virulence factor sta2, in response to an in vitro small intestine fermentation with gastric-digested honey.

Methods: An in vitro gastric digestion of both filtered honey and a simple sugar control (42% fructose, 35% glucose, 23% water) was performed in a simulated gastric electrolyte solution containing pepsin and hydrochloric acid. Duplicate digests were spiked with 8x10^5 CFU/mL ETEC. Resulting digesta was added at 1% w/v to mock community SIMO fermentations, containing 1.0x10^5 cells each of 8 common small intestine commensal bacteria and fermented anaerobically for 6 hours at 37℃ and an initial pH of 5.8.

Bacterial suspensions were pelleted at 2 h intervals and half were preserved in RNALater and stored at -80℃ until DNA was extracted. Abundance of the virulence factor sta2 was measured via qPCR against the housekeeping gene arcA.

16S rRNA amplicon sequencing was performed on bacterial DNA collected from remaining bacterial pellets. Data analysis was performed in Qiime2, with SILVA 138 taxonomy classifiers.

Results: Analysis of virulence gene abundance found no significant differences in filtered clover honey as compared to a matched sugar control or media alone. In the small intestine fermentations, proportions of E. coli decreased and Bacteroides increased in honey relative to the sugar control, but these trends were not significant after correction for multiple comparisons.

Conclusions: Our results indicate that while honey consumption may have a dramatic impact on E. coli survival in the stomach environment, it likely does not significantly impact microbial community composition, nor reduce ETEC virulence expression within the context of an in vitro small intestine environment.

Funding Sources: The National Honey Board, USDA, and the Dean’s Distinguished Graduate Fellowship, UC Davis