Characterization of rumen microbiome and metabolome from an oro-esophageal probe and fluid, particulate and fluid-particulate fractions from rumen fistula in Holstein dairy cows

Less invasive rumen sampling methods such as oro-esophageal probes became widely popular to explore the rumen microbiome and metabolome. However, it remains unclear if such methods represent well the rumen contents from fluid and particulate fractions. Herein, we characterized the microbiome and metabolome in rumen content collected by an oro-esophageal probe and fluid, particulate, and the combined fluid-particulate fractions collected by rumen fistula in ten multiparous Holstein dairy cows. The 16S rRNA gene was amplified and sequenced using the Illumina MiSeq platform. Untargeted metabolome was characterized using gas chromatography of a time-of-flight mass spectrometer. Although the pH of oro-esophageal samples was greater than those of fluid, fluid-particulate, and particulate ones, we found no difference in alpha and beta-diversity of their microbiomes. Bacteroidetes, Firmicutes, and Proteobacteria were consistently the top three most abundant phyla representing ~90% of all detected phyla across all samples. The overall metabolome PLS-DA of oro-esophageal samples was similar to the fluid-particulate samples but differed from fluid and particulate. Enrichment analysis pathways revealed few differences between oro-esophageal and fluid-particulate samples, such as the synthesis of unsaturated fatty acids. The results of the current study suggest that oro-esophageal sampling can be a proxy to screen the rumen microbiome with the 16S platform and overall fluid-particulate metabolome for a single-time and diet context. Nonetheless, studies focusing specifically on fluid and particulate metabolomes and specific metabolic pathways should carefully consider the sampling method used. IMPORTANCE The techniques used to collect the rumen contents (oro-esophageal probe and rumen fistula) suggested potential differences in the populations of rumen microbes, and the implications of these techniques for high throughput studies characterizing the rumen microbiome and metabolome need further elucidation. Ten rumen-fistulated Holstein dairy cows were used to characterize the microbiome and metabolome of samples collected using an oro-esophageal probe and the rumen-fistula fluid, particulate, and fluid-particulate fractions. The results of the current study suggest that oro-esophageal sampling represents well the rumen microbiome and overall fluid-particulate metabolome. However, fluid and particulate metabolomes and specific metabolic pathways across all types of rumen samples differed, indicating that studies focused on the characterization of rumen metabolome variable fractions should carefully consider the sampling method used.