ABSTRACT:In order to improve the gut microbiome of calves, probiotic bacteria can be fed as active livingcells (fermented milk), or as live but inactive (freeze-dried) cultures. Ten bifidobacterial strains with suitable probiotic properties (as determined in our previous study) were tested for survival during the freeze-drying process, and screened for their ability to ferment cow's milk. The viability of both freeze-dried and live-cell cultures during storage was also tested. All of the strains tested were able to ferment cow's milk, with average counts of 8.26 ± 0.62 log CFU/ml. Eight out of the ten strains were able to survive in milk for 2 months in counts higher than 10 6 CFU/ml. Bifidobacteria showed high viability following the freeze-drying process, with average numbers of 9.03 ± 0.22 log CFU/vial and did not decrease after 12 months of storage. The mixture of rifampicin-resistant variants of bifidobacteria (RRBs) was fed to 2-day-old dairy Charolais calves in the form of living-cells, or as freeze-dried bacteria. The control group was given no probiotics. Survival of the RRBs administered and the numbers of other bacterial groups in faecal samples was monitored by culturing. Bifidobacteria that were administered passed successfully through the upper parts of the gastrointestinal tract, and were found in numbers higher than 10 9 CFU/g for two weeks. RRBs colonized the intestines of calves for at least 63 days in both treatment groups. Significantly higher total counts of bifidobacteria were found in the treated groups, compared to the control group. Reduction in Escherichia coli and total coliforms numbers, and an increase in lactobacilli counts were observed in both experimental groups following the application of the probiotic mixtures. Our results show that both forms of administering probiotic bifidobacteria to calves are effective, but that the freeze-dried form is more suitable from a practical viewpoint.