Effect of rhubarb (Rheum spp.) root on in vitro and in vivo ruminal methane production and a bacterial community analysis based on 16S rRNA sequence

Abstract

The objective of this study was to evaluate the anti-methanogenic effect of rhubarb (Rheum spp.) on in vitro, in vivo, and bacterial community composition using Quantitative Insights into Microbial Ecology sequencing. Rhubarb root powder was tested at different concentrations (0, 0.33, 0.67, and 1.33 g/L) in vitro, and all incubations were carried out in triplicate two runs on separate days. Concentrations of 0.67 and 1.33 g/L rhubarb significantly (P < 0.05) reduced methane production and the acetate : propionate ratio compared with those of the Control, without adverse effects on total volatile fatty acids and total gas production. In the second in vivo trial, four Hanwoo (Korean native) steers (live bodyweight, 556 ± 46 kg) with a ruminal cannula were housed individually in metabolic stalls and fed a basal diet twice daily in equal amounts at 0900 hours and 2100 hours. The before rhubarb treatment (before treatment) duration was 24 days for all steers; 14 days were used for diet adaptation and 10 days were used for gas samples collected 1, 2, and 3 h after the morning feeding on Days 3, 5, 7, and 9. We used three syringe needles passed through the ruminal cannula stopper at different time points as a simple and rapid method to sample rumen gas. Thereafter, three mesh bags containing 30 g of sliced rhubarb root each were placed at different depths in the rumen of each steer for 14 days (after treatment), and gas samples were collected on Days 4, 7, 10, 12, and 13. The results showed a significant (P < 0.05) decrease in methane concentration from the rhubarb-treated steers and provide the evidence that this method would be useful for in vivo screening of anti-methanogenic feed additives or plant material. Furthermore, 16s RNA sequencing after treatment showed increases in the numbers of Prevotella, and Lactobacillus, but decreases in Methanobrevibacter. In conclusion, rhubarb had an anti-methanogenic effect in vitro and in vivo, and the increase in the number of Prevotella shifted ruminal fermentation towards propionate production.