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The literature estimates that unknown rumen bacteria with rhodanese activity are key factors in the animal detoxification of cyanogenic glycosides, as they are capable of transforming CN into the less toxic thiocyanate. Therefore, identifying these bacteria will enhance our understanding of how to improve animal health with this natural CN detoxification process. In this study, a rhodanese activity screening assay revealed 6 of 44 candidate rumen bacterial strains isolated from domestic buffalo, dairy cattle, and beef cattle, each with a different colony morphology. These strains were identified as belonging to the species <jats:italic>Enterococcus faecium<\/jats:italic> and <jats:italic>E. gallinarum<\/jats:italic> by 16S ribosomal DNA sequence analysis. A CN-thiocyanate transformation assay showed that the thiocyanate formation capacity of the strains after a 12\u00a0h incubation ranged from 4.42 to 25.49\u00a0mg hydrogen CN equivalent\/L. In addition, thiocyanate degradation resulted in the production of ammonia nitrogen and acetic acid in different strains. This study showed that certain strains of enterococci substantially contribute to CN metabolism in ruminants. 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