Impacts of Protein and Energy Levels on Rumen Fermentation and Microbial Activity Under Different Incubation Temperatures

This study aimed to explore the effects of different incubation temperatures on ruminal fermentation and rumen microorganisms and determine the appropriate protein and energy levels to enhance microbial protein synthesis using an in vitro system. Rumen inoculum was collected from two fistulated Holstein heifers (trial 1: BW: 652.3 kg ± 25.2; trial 2: BW: 683.3 kg ± 30.2) and assessed using a closed-batch culture system. The experimental model employed a 2 × 5 factorial arrangement using incubation temperatures set to 39 and 41 °C, with protein levels set to 12.0, 13.5, 15.0, 16.5, and 18.0% of DM in trial 1 or with energy levels set to 2.4, 2.5, 2.6, 2.7, and 2.8 Mcal/kg of DM in trial 2. The data were analyzed using the MIXED procedure. The results showed increased (<i>p</i> < 0.05) NH₃-N concentrations and total volatile fatty acids (TVFAs) with higher incubation temperatures, while the liquid-associated bacterial (LAB) amounts decreased (<i>p</i> < 0.05) in trials 1 and 2. The interaction between the energy level and incubation temperature affected (<i>p</i> < 0.05) the LAB protein levels in trial 2. Higher protein levels led to increased (<i>p</i> < 0.05) NH₃-N and acetate concentrations, but it decreased (<i>p</i> < 0.05) the propionate percentage. Conversely, higher energy levels decreased (<i>p</i> < 0.05) the amount of acetate and increased the propionate concentration, altering the acetate-to-propionate ratio. However, no interaction involving TVFA and LAB was observed between the incubation temperature and the protein or energy levels. Changes in the NH₃-N, TVFAs, and LAB protein amounts were observed under different incubation temperatures and energy levels. In conclusion, these findings provide insight into the metabolic adaptation under different ruminal temperatures and the impacts of dietary adjustments on rumen fermentation and microbial activity. However, there are limitations to replicating the complex physiological responses that occur within the whole body solely through in vitro experiments.