Dose-response effects of Poncirus trifoliata extract on in vitro ruminal methane production, fermentation, and microbial abundance

The effects of different inclusion rates of Poncirus trifoliata (PT) extract on in vitro ruminal fermentation parameters were investigated. Six dose levels were referred to as follows: PT0, control (without PT); PT1, (3 mg/L); PT3, (9 mg/L); PT5, (15 mg/L); PT7, (21 mg/L); PT9, (27 mg/L), each incubated with 50 mL of buffered rumen fluid and 300 mg of timothy hay as a substrate for 72 h of incubation using an in vitro batch culture system. The in vitro dry matter digestibility (IVDMD) was significantly affected by the PT dose, except after 6 and 48 h of incubation. Total volatile fatty acids (VFAs) increased quadratically (12 h of incubation) within the range of PT1–PT7, whereas decreased at PT9 dose. Methane (CH4) production and the proportion of CH4 in the total gas produced decreased linearly after 12 h of incubation. The relative abundance of Ruminococcus albus and Ruminoccocus flavefaciens was greater at the doses PT1 than PT0, whereas that of Fibrobacter succinogenes was greater at PT0 than at PT9 after 24 h of incubation. Methanogenic archaea decreased linearly with increasing PT dose after 12 h of incubation. The abundance of ciliate-associated methanogens decreased linearly over 24 h of incubation under PT7 and PT9 treatments. These results suggest that the dose of PT can modulate ruminal fermentation to alter VFA concentration and reduce CH4 production by altering ruminal bacterial abundance.Highlights This study evaluated the dose-response effects of Poncirus trifoliata as a methane mitigation agent in ruminants. Inclusion of Poncirus trifoliate lowered in vitro dry matter digestibility and total volatile fatty acid concentration. Inclusion of Poncirus trifoliata extract reduced methane production after 12 h of incubation but did not maintained. Inclusion of Poncirus trifoliata extract reduced methanogenic archaea and ciliate-associated methanogens abundance