Fading puppy syndrome (FPS) is a fatal condition in neonatal dogs. Intestinal microbial alterations, although never investigated, may be involved in its pathophysiology. The study examined the occurrence of FPS and its associations with dam, puppy, and husbandry characteristics, compared the intestinal microbial diversity of healthy puppies and those with FPS, and examined whether intestinal microbiomes are predictive of FPS. Day 1 and 8 post-partum (PP) rectal swabs were collected from healthy puppies and puppies which later developed FPS. Microbial compositional structure, including alpha and beta diversities and relative abundance of specific taxa were compared between groups, and microbial data was applied to a machine-learning model to assess the predictive performance of microbial indices of FPS or death. FPS occurred in 22/165 puppies (13%), with a 100% mortality rate. FPS was associated (P < 0.001) with decreased Day 1 PP puppy activity. Day 1 (P = 0.003) and 8 (P = 0.005) PP rectal beta diversities were different in puppies with FPS vs healthy ones. Increased Proteobacteria/Firmicutes ratio, increased relative abundance of Pasteurellaceae, and decreased relative abundance of Clostridia and Enterococcus were associated with FPS. A machine-learning model showed that Day 1 PP rectal microbiome composition accurately predicted FPS-related death. We found that specific rectal microbial phenotypes are associated with FPS, reflecting the significant role of microbiome alterations in this phenomenon. These findings may serve as useful microbial indices for early diagnosis of puppies at risk of FPS and may provide specific therapeutic targets.
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