Nitrogen efficiency (Neff; milk N/N intake) in dairy cows is limited and most of the consumed N is excreted in manure. Despite the crucial role of the gastrointestinal microbiome on N metabolism, associations between bacterial communities at different sections and Neff are not fully elucidated. Enhanced understanding of host-microbiome interactions can provide insights to improve Neff in dairy cows. Twenty-three Holstein cows were selected, and their Neff were determined using a N balance approach. From the cohort of cows, six cows were classified as low Neff and five cows as high Neff and their rumen and fecal bacterial communities were profiled using amplicon sequence variants (ASV) based on 16S rRNA gene sequencing. Then, relationships between differentially abundant bacterial features and Neff were evaluated. Neff in low and high cows averaged 22.8 and 30.3%, respectively. With similar N intake, high Neff cows wasted less N in manure compared to low Neff cows (P< 0.01, 11.0 ± 0.59 vs 14.3 ± 0.54 g of N/kg of milk). Rumen fermentation and plasma profiles were similar between Neff groups, but for plasma Gln which was greater (P= 0.02) in high compared to low Neff cows. In both rumen and feces, the phylogenetic composition of the bacterial communities was similar (P≥ 0.65) between Neff groups, but differences were observed at the species -level (amplicon sequence variants). In the rumen, differentially abundant species from the genusPrevotellashowed strong positive correlations with Neff, whereas in feces, differentially abundant species from the classClostridiashowed strong negative correlations with Neff. Our results revealed that Holstein cows with divergent Neff display distinctive bacterial community structure at the species-level in both the rumen and feces. Strong correlations between differentially abundant species and Neff in both sample sites, support the importance of the rumen bacterial composition on productive responses and suggest a more relevant role of the hindgut microbiome. Targeting both pre- and post-gastric bacterial communities may provide novel opportunities to enhance Neff in dairy cows.