Technical Abstract: The objective of the current study was to examine the effect of pasteurization of waste milk used to feed dairy calves on the bacterial diversity of their lower gut. Using 16S rDNA bacterial tag-encoded FLX amplicon pyrosequencing (bTEFAP), fecal samples from dairy calves aging from 1 week to 6 months old and fed either pasteurized or non-pasteurized waste milk were analyzed for bacterial diversity. Calves were maintained on two separate farms and aside from how the waste milk was treated, were housed and cared for similarly. Fifteen calves were sampled from each age group (1, 2, and 4 weeks, and 2, 4, and 6 months of age; n = 90 samples per milk treatment, 180 total samples) on each farm via rectal palpation and the samples shipped and frozen prior to analysis. In general, bacterial diversity as represented by the total number of different species was greater for the calves fed pasteurized waste milk at all ages (except 1 week of age) and increased with increasing age in both treatments. Proteobacteria, Bacteroidetes, and Firmicutes were the predominant phyla. Differences in phyla and class were observed among treatments and age of calf but with no consistent trends. Salmonella were detected in 9 out of 14 (64%) of the week-old calves fed non-pasteurized milk. Treponema, an important beneficial bacteria in cattle rumen, was more prevalent in the pasteurized animals and became higher (P < 0.001) in the older animals from this group. Escherichia-Shigella were detected among treatments at all ages and highest at one week of age, averaging approximately 21 and 20% of all bacteria for calves fed pasteurized and non-pasteurized waste milk, respectively and declining as calves aged (2.6 and 1.3%). The consistent detection of Salmonella in the younger animals fed non-pasteurized milk and its absence in all other groups is an important finding related to this feeding practice.