Fecal microbiota transplantation (FMT) has been used empirically for decades in equine medicine to treat intestinal dysbiosis but evidence-based information is scarce. This in vitro study aimed at assessing the effect of a commonly used pre-FMT processing method on the bacterial composition and viability of the fecal filtrate. Three samples of fresh equine manure (T0) were processed identically: the initial manure was mixed with 1 L of lukewarm water and chopped using an immersion blender to obtain a mixture (T1), which was left uncovered during 30 min (T2) and percolated through a sieve to obtain a fecal filtrate (T3). Samples were taken throughout the procedure (Tn) and immediately stored at 4 °C until processing. The 16S rDNA amplicon profiling associated with propidium monoazide treatment was performed on each sample to select live bacteria. Analyses of α and β diversity and main bacterial populations and quantitative (qPCR) analysis were performed and statistically compared (significance p < 0.05) between time points (T0–T3). No significant differences in ecological indices or mean estimated total living bacteria were found in the final fecal filtrate (T3) in regard to the original manure (T0); however, relative abundances of some minor genera (Fibrobacter, WCHB1-41_ge and Akkermansia) were significantly different in the final filtrate. In conclusion, the results support the viability of the major bacterial populations in equine feces when using the described pre-FMT protocol.

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