Temporal variability is an inherent feature of the healthy canine microbiome assessed by full-length 16S rRNA gene sequencing.

The canine gut microbiome plays a crucial role in host health, yet assessment of its temporal variability remains largely unexplored. Here, we used full-length 16S rRNA gene sequencing to evaluate long-term temporal variability of the fecal microbiomes of 35 healthy dogs sampled monthly over 1–2 years (median samples per dog: 10, range: 5–28 samples per dog, during the years 2020–2023). Analyses revealed that while individual-specific signatures in microbiome composition were observed (PERMANOVA R2 = 0.39, p = 0.001), fecal microbiomes exhibited substantial temporal fluctuations within individuals (9/35 dogs with microbiome dispersion ≥ 75%tile). Microbiome alpha-diversity was also not temporally stable (GAM F = 4.99, p = 0.025). The relative abundances of the 35 bacterial species varied considerably over time with Escherichia coli, and Enterococcus faecium especially dynamic (median CV 243–249%), and Peptacetobacter hiranonis and Collinsella intestinalis more stable (median CV 112–120%). Several taxa exhibited seasonal patterns, such as Fusobacterium spp. peaking in late summer and Peptacetobacter hiranonis declining in July. Collectively, these findings indicate that temporal fluctuations are an inherent feature of the healthy canine microbiome. Lastly, since our sampling captured occasional parasite or pathogen infections, we assessed whether transient C. perfringens, Giardia, or Campylobacter infections were correlated with microbiome alpha- and beta-diversity. We found no correlations with beta-diversity (p > 0.05), but a Giardia infection was associated with lower alpha-diversity (GLMM β= -0.37, p = 0.04).