Effects of phyto-phenolic compounds on ammonia production by select amino acid fermenting bacteria

Authors: LAKES, JOURDAN - Oak Ridge Institute For Science And Education (ORISE); RAMOS, LEAH - Texas A&M University; CARDENAS, MAEDEAN - Texas A&M University; MAST, NATASHA - Texas A&M University; Flythe, Michael

Submitted to: FEMS Microbiology Letters
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/30/2025
Publication Date: 2/6/2025
Citation: Lakes, J.E., Ramos, L.I., Cardenas, M.L., Mast, N.L., Flythe, M.D. 2025. Effects of phyto-phenolic compounds on ammonia production by select amino acid fermenting bacteria. FEMS Microbiology Letters. 372:Article fnaf018. https://doi.org/10.1093/femsle/fnaf018.
DOI: https://doi.org/10.1093/femsle/fnaf018

Interpretive Summary: Certain bacteria within cattle destroy amino acids (protein) from the animal's diet by a process called amino acid fermentation, which results in ammonia that is excreted from the animal in the urine. This process negatively impacts the ability of beef cattle to gain weight and often results in the need to supplement feed with additional protein. The waste product can also contaminate groundwater. Therefore, the presence of the amino acid fermenting bacteria is wasteful and their control beneficial. Additionally, a similar process occurs in fermented forages (i.e., silage) where the fermentation of plant protein by amino acid fermenting bacteria also produces ammonia. This process can be ruinous because ammonia is basic and raises the pH of the silage. Successful fermentation of forages depends on a low pH (acidic). Plant-derived natural antimicrobials could be supplemented into the diets of cattle and into ensiled forages. The current study sought to investigate the antimicrobial potential of several plant compounds for the control of amino acid fermenting bacteria from ruminants (such as beef cattle) and stored forages (silage). Pure cultures of five amino acid fermenting bacteria were treated with five concentrations of each plant compound individually to assess antimicrobial activity. From this data, the lowest fully inhibitory concentration of each active compound was used on fully grown (stationary phase) cultures of each inhibited species to determine the effect on amino acid fermentation via ammonia production. The data demonstrated substantial and consistent reductions in ammonia when the five cultures were treated with 1 mM carvacrol and 10 mM eugenol. Thymol (1mM) greatly reduced the ammonia produced by four of the five species, and trans/hydro-cinnamic acid yielded modest reductions in ammonia produced by one species each. Thus, carvacrol, eugenol, and thymol appear to be candidate compounds of interest for the control of amino acid fermenting bacteria in ruminants and stored forages.

Technical Abstract: Bacteria that ferment amino acids to ammonia can be categorized as generalists or specialist hyper-ammonia producing bacteria. In the rumens of ruminant animals, most of the ammonia produced is eventually excreted as urea in urine. This study investigated the antimicrobial and metabolic suppressive potential of six phenolic compounds against five amino acid fermenting bacteria: Clostridium sporogenes MD1 and C. aminophilum F, Acetoanaerobium sticklandii SR, Peptostreptococcus sp. BG1, and Prevotella bryantii B14. Carvacrol (1 mM) inhibited the growth of all five species and thymol (1 mM) fully inhibited growth of four of the species with the exception of BG1. Eugenol inhibited the growth of all bacteria at the highest concentration used (10 mM). The cinnamic acids (trans- and hydro-) inhibited either fully or partially all organisms except F and B14. Carvacrol and eugenol produced the greatest reduction of ammonia by all organisms except B14 who produced no ammonia under the growth conditions. Thymol greatly reduced ammonia production of four organisms except for F. Hydro-cinnamic acid decreased ammonia in SR and trans-cinnamic acid reduced ammonia in MD1, only. Additionally, the maximum specific rates of ammonia production by MD1, F, and BG1 were altered by all treatments. Conversely, the maximum rate of ammonia produced by SR within the first hour of sampling was generally unaffected. These data demonstrate that eugenol, carvacrol, and thymol may be worthy antimicrobial candidates for the control of ammonia producing organisms.