Temporal variations of intestinal microorganisms have been investigated in humans, but limited information is available for other animal species. The aim of the study was to evaluate the abundance and short-term temporal variability of fecal microbiota in dogs. Two fecal samples were collected (15 days apart) from six healthy dogs. The microbiota was evaluated using fluorescence in situ hybridization (FISH) and 454-pyrosequencing targeting the 16S rRNA and its gene. Pyrosequencing revealed 15 families comprising >80% of all microbiota, over time intraindividual coefficients of variation (%CV) ranged from 2% to 141% (median: 55%). In contrast, the interindividual %CV ranged from 62% to 230% (median: 145%). Relative proportions of Faecalibacterium (important for intestinal health) and Subdoligranulum were low (two dogs harbored 4-7% of Subdoligranulum, the remaining dogs had <1% of either genus). Conversely, FISH revealed that Faecalibacterium comprised a median of 5% of total counts (range: 0-8%, probe Fprau645). A novel FISH probe (Faecali 698) was tested that, compared with Fprau645, can detect in silico a similar percentage of Faecalibacterium but higher proportions of Subdoligranulum. This probe revealed a high percentage of Faecalibacterium-Subdoligranulum (median: 16% of total counts). Future studies should consider the observed variability and discrepancies in microbial abundance between FISH and 454-pyrosequencing.