In vitro methods for assessing gas production and rumen digestibility: Methodological considerations, advantages, and...

Although animal-based studies provide information with the highest physiological and metabolic relevance, in vitro methods are essential for assessing diets and feedstuffs during initial screening. These methods have accelerated our understanding of digestion and fermentation in the rumen ecosystem, contributing to the optimization of animal production and the mitigation of greenhouse gas emissions. This paper reviews the characteristics, strengths, weaknesses, and standardization procedures for the current in vitro rumen fermentation methods. Overall, these methods reduce animal-to-animal variation and are faster than in vivo trials; however, their main limitations are the closed system's characteristics and inability to cover host-microbe-environment interactions. Thus, selecting the appropriate method requires careful evaluation of the investigator's aims. The in vitro batch culture (IVBC) and the ANKOM RF Gas Production System (AGPS) are valuable methods for assessing gas production and digestibility; both also measure environmental impacts, such as methane production. The rumen simulation Technique (RUSITEC) is closest to rumen conditions and could have a greater impact on long-term rumen microbiota research. On the other hand, the ANKOM Daisy^II incubator is an efficient apparatus for forage digestibility determination in livestock species. Finally, we highlight guidelines to standardize and increase the robustness and reproducibility of these methods.