Aims: To isolate predominant ruminal bacteria that produce trans -10, cis -12 conjugated linoleic acid (CLA) from linoleic acid (LA). Methods and Results: Mixed bacteria from ruminal contents of a cow fed grain were enriched with DL-lactate and trypticase. They produced more trans -10, cis -12 CLA than those that were not enriched (7 vs 2 mug mg protein(-1) , P < 0.05). Enrichments had an abundance of large cocci that produced trans -10, cis -12 CLA from LA. Strain YJ-4 produced the most trans -10, cis -12 CLA (approx. 7 μg mg protein(-1) ) and 16S rDNA sequencing indicated that YJ-4 was a strain of Megasphaera elsdenii. Megasphaera elsdenii T81 produced approx. 4 μg trans -10, cis -12 CLA mg protein(-1) while strains B159, AW106 and JL1 produced < 0.5 mug mg protein(-1) . The trans -10, cis -12 CLA production of YJ-4 was first order with respect to cell concentration (0-800 mug protein ml(-1) ), but kinetics were not first order with respect to substrate concentration. Conclusions: Some M. elsdenii strains produce significant amounts of trans -10, cis -12 CLA. Significance and Impact of the Study: Trans- 10, cis -12 CLA appears to cause milk fat depression in cattle fed diets supplemented with grain and polyunsaturated fatty acids, but predominant ruminal bacteria that produced trans -10, cis -12 CLA from LA had not previously been isolated.