RUMINANTS, ESPECIALLY DAIRY COWS AND BEEF CATTLE, ARE AN IMPORTANT SOURCE OF HIGH-QUALITY PROTEIN FOR HUMAN CONSUMPTION. THEY CAN UTILIZE FORAGE AND GRAINS TO MAKE ANIMAL PROTEIN LIKE MILK AND BEEF. HOWEVER, THEY CAN ONLY UTILIZE ABOUT 30% OF THE DIETARY NITROGEN, THE MOST IMPORTANT COMPONENT OF PROTEIN. THE LOWER NITROGEN EFFICIENCY IN RUMINANTS THAN IN NON-RUMINANTS IS ATTRIBUTED TO SOME MICROORGANISMS IN THE STOMACH OF THE FORMER, PARTICULARLY RUMEN PROTOZOA. AS A UNIQUE GROUP OF MICROORGANISMS OF THE MICROBIAL COMMUNITY IN THE RUMEN, RUMEN PROTOZOA ENGULF AND DEGRADE THE MICROBIAL PROTEIN SYNTHESIZED BY THE MICROBIAL COMMUNITY USING THE INGESTED FEED, WHICH CREATES RECYCLING OF MICROBIAL PROTEIN IN THE RUMEN AND DECREASING THE FLOW OF THE PROTEIN TO THE SMALL INTESTINE WHERE PROTEIN IS DIGESTED AND ABSORBED. THIS WASTEFUL INTRARUMINAL RECYCLING OF MICROBIAL PROTEIN SIGNIFICANTLY DECREASES NITROGEN UTILIZATION EFFICIENCY IN ALL RUMINANTS, RESULTING IN HIGH PRODUCTION COST AND ENVIRONMENTAL POLLUTION BY THE EXCRETED NITROGEN. THUS, THERE IS A CRITICAL NEED TO DECREASE THE INTRARUMINAL RECYCLING OF MICROBIAL PROTEIN TO IMPROVE NITROGEN UTILIZATION EFFICIENCY IN RUMINANTS, ESPECIALLY IN DAIRY COWS. RECENTLY, WE FOUND THAT (I) RUMEN PROTOZOA DEPEND ON LIVE BACTERIA FOR THEIR SURVIVAL AND GROWTH, (II) THE GENES ENCODING THEIR DIGESTIVE ENZYMES ARE HIGHLY EXPRESSED, (III) INHIBITION OF THE DIGESTIVE ENZYMES SUBSTANTIALLY INHIBITS RUMEN PROTOZOA AND AMMONIA PRODUCTION, WITHOUT ADVERSELY AFFECTING OTHER ASPECTS OF THE RUMEN ECOSYSTEM, AND (IV) SPECIFIC INHIBITORS AGAINST THE DIGESTIVE ENZYMES CAN BE PREDICTED FROM THE STRUCTURES OF THE DIGESTIVE ENZYMES. IN THIS RESEARCH, WE WILL CONDUCT IN VITRO STUDIES TO IDENTIFY THE BEST INHIBITORS, DETERMINE THE EFFECTIVE DOSES, AND INVESTIGATE THEIR EFFECT ON FEED DIGESTIBILITY, ENGULFMENT AND DEGRADATION OF BACTERIA, ACTIVITIES OF THE DIGESTIVE ENZYMES, AND THE RUMEN MICROBIAL COMMUNITY.WE WILL THEN USE GROWING LAMBS AS A MODEL TO VERIFY THE INHIBITORS AND THEIR EFFECT ON THE ANIMALS AND THE RUMEN MICROBIOME. OUR ULTIMATE GOALS ARE TO FILL CRITICAL GAPS IN KNOWLEDGE ON THE ROLES OF PROTOZOAL DIGESTIVE ENZYMES IN RUMEN FUNCTION AND TO DEVELOP AN INNOVATIVE APPROACH TO IMPROVE NITROGEN UTILIZATION EFFICIENCY, PARTICULARLY IN DAIRY COWS AND BEEF CATTLE, WITH NO ADVERSE EFFECT ON THE ANIMALS OR THEIR RUMEN ECOSYSTEM. THIS PROPOSED RESEARCH WILL BE TRANSFORMATIVE AND TRANSLATIONAL AND CAN POTENTIALLY BENEFIT THE US DAIRY AND BEEF PRODUCERS ECONOMICALLY AND THE SOCIETY ENVIRONMENTALLY IN THE FUTURE.