Submitted to: Applied and Environmental Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/26/2001
Publication Date: 3/15/2002
Citation: RYCHLIK, J.L., RUSSELL, J.B. BACTERIOCIN-LIKE ACTIVITY OF BUTYRIVIBRIO FIBRISOLVENS AND ITS EFFECT ON OTHER RUMINAL BACTERIA AND ON AMMONIA PRODUCTION. APPLIED AND ENVIRONMENTAL MICROBIOLOGY. 2002. Interpretive Summary: Cattle in the U.S. are often fed antibiotics, but the widespread use of antibiotics in animal feed has been criticized. Antibiotics are primarily targeted against gram-positive gut bacteria. Gram-positive ruminal bacteria produce large amounts of hydrogen, a precursor of methane ammonia, a wasteful end-product of amino acid degradation, and lactic acid, an acid that causes ruminal acidosis, ruminal ulcers, founder and even death of the animal. Some gram-positive bacteria produce peptides (bacteriocins) that can inhibit other gram-positive bacteria, and bacteriocins have been proposed an alternative to antibiotics. We isolated a ruminal bacterium that inhibits ruminal bacteria that wastefully degrade amino acids, and we have purified the bacteriocin. Research on bacteriocins has the potential to decrease the need for antibiotic in animal feed and lessen the risk of antibiotic resistance in human medicine.
Technical Abstract: When mixed ruminal bacteria from a cow fed hay were serially diluted into medium that had only peptides and amino acids as energy sources, little growth or ammonia production was detected at dilutions greater than 1 million. The high dilutions did, however, have bacteria that could ferment carbohydrates, and some of these bacteria inhibited Clostridium sticklandii iSR, an obligate amino acid fermenting bacterium. Phylogenetic analysis indicated that the most active isolate (JL5) was closely related to other strains of Butyrivibrio fibrisolvens. Strain JL5 inhibited a variety of Gram-positive species, but it had no effect on Gram-negative ruminal bacteria. Bacteriocin production did not occur until JL5 had reached stationary phase, and glucose was exhausted. The JL5 bacteriocin did not cause the lysis of C. sticklandii SR, but viability decreased dramatically. Bacteriocin-treated cell suspensions of C. sticklandii SR lost intracellular potassium, but these non-viable cells retained their ability to produce ammonia. The bacteriocin was resistant to several proteases, and it could be precipitated with 60% ammonium sulfate and dialyzed (3500 Da cut-off). The bacteriocin was separated by PAGE, and C. sticklandii SR overlays indicated that the molecular weight was approximately 3600 Da. Because ruminal fluid inhibited the growth of Clostridium aminophilum F and Peptostreptococcus anaerobius C (as well as C. sticklandii SR), bacteriocins may play a role in regulating ammonia production in vivo.