Abstract Background: Reducing enteric methane emissions from farmed ruminants can be achieved by various nutritional strategies. However, it remains unclear to what extent the effects of diet on rumen microbiome are comparable between different ruminant species. In this work, we compared the effects of starch- and/or lipids-rich diets on the rumen microbiome of cows and goats to enhance our understanding of microbial mechanisms of methanogenesis. The study enrolled four Holstein cows and four Alpine goats, conducted simultaneously in a replicate 4×4 Latin square design, receiving the same ration based on grassland hay and concentrate (CTL) or supplemented with corn oil and wheat starch (COS), marine algae powder (MAP) or hydrogenated palm oil (HPO). The microbiome was studied using non-targeted total RNA sequencing. Results: To identify biologically relevant features, we developed a five-step biostatistical pipeline, combining a network-based approach with clustering and supervised model fitting to associate differentially expressed genes (deKEGGs) and Operational Taxonomic Units (deOTUs) with methane emissions. The COS diet induced the highest methane emissions reduction for cows and goats and most profoundly affected the microbiome. With a focus on the COS vs . CTL comparison, the number of deOTUs and deKEGGs in cows (16 and 381) was higher than in goats (10 and 133). Moreover, clustering analysis revealed network topology and functionality differences between ruminant species. In goats, the reduction in methane emissions was strongly associated with genes involved in Carbohydrate metabolism, and genes coding for methylotrophic and hydrogenotrophic pathways of methanogenesis were overrepresented; in cows, only the hydrogenotrophic pathway was prevalent. Further, sPLS analysis identified potential biomarkers characteristic of each ruminant species, such as tetrahydromethanopterin S-methyltransferase and fructose bisphosphate aldolase for cows, and Methanol: coenzyme M methyltransferase, F420-non-reducing hydrogenase for goats. Conclusions: Overall, these results demonstrated a strong influence of the ruminant species on the responses of the rumen microbial community to dietary changes. We observed that the COS diet reduced similarly enteric methane emissions in cows and goats, but the induced shifts in the rumen microbiome were not the same. These results suggest that the host-animal species conditions microbial interactions within the rumen ecosystem.