Ten Holstein cows were used to investigate the changes in ruminal bacterial populations in response to induction and recovery from diet-induced milk fat depression (MFD). Further, the impact on the ruminal bacterial composition of the cows with MFD inoculated with rumen contents from non-milk fat-depressed donor cows was evaluated. MFD was induced during the first 10 d of each period by feeding a low-fiber and high-polyunsaturated fatty acid (FA) diet [induction diet; 26.1% neutral detergent fiber, 5.8% FA (1.9% C18:2)], resulting in a 30% decrease in milk fat yield. Induction was followed by a recovery phase where all cows were switched to a high-forage, low-polyunsaturated FA diet (31.8% NDF, 4.2% total FA (1.2% C18:2)) and were allocated to (1) control (no inoculation) or (2) ruminal inoculation with donor cow digesta (8 kg/d for 6 d). Ruminal samples were collected at the end of induction (d10) and through recovery (d13, d16 and d28) of each period, separated to solid and liquid fractions, extracted for DNA, PCR-amplified for V1-V2 hypervariable region of the 16S rRNA gene and sequenced using Ion Torrent (PGM) platform for bacterial diversity. The 16S pyrosequence reads were analyzed using the QIIME (v 1.8.0) and all statistical analysis were performed using SAS (V 9.3) and R (V 3.3.1). In both fractions, d10 bacterial communities were observed to be different (P < 0.05) from d13, d16 and d28. MFD (d10) was typified by a reduction (P < 0.05) in Bacteroidetes and Fibrobacteres and an increase (P < 0.05) in Firmicutes and Actinobacteria across both fractions. Both Bacteroidetes and Fibrobacteres recovered as early as 3 d into the recovery period their sensitivity to induction diet. Inoculation of rumen contents from non-MFD donor cows to MFD cows revealed that transfaunation procedure did not result in wholesale shifts in the ruminal microbiota, however, influenced a few genera from Bacteroidetes, Fibrobacteres and Spirochaetes in the liquid fraction. These microbial changes diminished after d16 indicating that the impact of inoculation on the rumen microbiota is a transient effect. It can be concluded that diet-induced MFD is accompanied by microbial dysbiosis in the rumen.