The impact of pooling on the observed microbiome profile of preweaned piglet feces

Pooling individual samples could be an efficient approach for large-scale population-based microbiome studies. However, it is unknown whether pooled samples accurately reflect the microbiome composition and diversity obtained from individual samples. This study investigated the impact of various pooling methods on the observed fecal microbiome of preweaned piglets. Individual fecal samples were collected from 10 litters of day-old piglets (N = 137) and 10 litters of 20-day-old piglets (N = 121), as well as pen-floor samples from the same litters. The individually collected samples were processed individually and also used to create pools of both raw feces and extracted DNA. Individual samples, raw feces pools, DNA pools, and pen-floor samples were subjected to 16S rRNA gene sequencing. The microbial profile in pen-floor samples from litters of preweaned piglets was very different from individual piglet samples within the pen; thus, they may not be suitable for litter-level piglet microbiome studies. However, overall microbial diversity and composition from DNA and feces pools were comparable to individual samples, despite potentially underestimating some low-abundance or low-prevalence taxa. These results suggest that pooling can be used as an efficient and cost-effective approach to characterize litter-level microbial profiles for current and future population-level microbiome research in preweaned piglet populations.