Previous studies showed that inclusion of polyphenol-rich plant extracts in feed improves performance in ruminants, but the effect may vary depending on the dosage used. It has been shown that polyphenols modify the digestive process through modulation of gastrointestinal microbiota and bypass of protein digestion. The rumen houses a highly complex microbiota which is able to convert indigestible plant mass into energy mainly through production of short-chain fatty acids (SCFA).
High-throughput sequencing of 16S rRNA gene was used to study the temporal changes in microbiota composition of eight rumen-fistulated steers supplemented with four doses of chestnut and quebracho plant extracts (0, 0.075, 0.15 and 0.30 kg per ton) in a 4x4 Latin-square design. In each sample, the pre- and post- prandial pH of the ruminal liquor was measured and the SCFA profile was determined by HPLC. Bioinformatics and statistical analysis of microbiota variability was performed using QIIME2 software. 6.724.119 reads of 16S rRNA gene V3-V4 region were obtained in total. A significant variation in rumen microbiota beta diversity was detected between animals and between samplings (p<0.001), but pre- and post-prandial samples showed very limited variation, with virtually identical patterns in both series of data. A smaller but significant effect was observed between polyphenol doses on alpha and beta diversity parameters. Steers with higher doses tend to have lower richness (p=0.09) and lower Shannon’s diversity index (p=0.03), which indicates a more even and balanced microbial community in treated animals. When comparing with the not-supplemented control group, all three polyphenol doses reduced the relative abundance of phylum Euryarchaeota, which includes the methanogenic archaea responsible for greenhouse gas production. The two higher doses also increased the relationship between Firmicutes and Bacteroidetes (F/B). The dose of 0.15% was the one with the highest F/B ratio and the lowest relative abundance of methanogenic bacteria. Regarding SCFAs production and pH, a strong correlation was observed between these parameters and rumen microbiota diversity. Steers with lower number of bacterial species had a more acidic ruminal pH (p=0.0004), and higher ratio of C2/C3 in their SCFA profile (p=0.0013).
The results showed that the groups supplemented with chestnut and quebracho polyphenols have similar changes compared to the control group, but the modulatory effect on the diversity and composition of rumen microbiota and SCFA production profile varies depending on the dosage of extract used, reaching an optimal effect at intermediate doses. Therefore, the dose of polyphenols is a parameter that can be optimized since differentially alters ruminal physiology.
Keywords: microbiota, phytochemicals, rumen, bovines, polyphenols