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Gut dysbiosis in patients with chronic kidney disease (CKD) may induce chronic inflammation and increase morbidity. Phosphate-binding agents, generally used in patients with CKD, may potentially change the composition of the gut microbiota. This study aimed to compare the microbiota composition in hemodialysis patients treated with ferric citrate or calcium carbonate. The stool microbiota was investigated in hemodialysis patients treated with ferric citrate (n = 8) and calcium carbonate (n = 46) using 16S rRNA gene amplicon sequencing profiling using linear discriminant analysis of effect size. Further predictive functional profiling of microbial communities was obtained with Tax4Fun in R. Hemodialysis patients treated with calcium carbonate had a significantly reduced microbial species diversity (Shannon index and Simpson index) and an increased microbial alteration ratio compared with patients treated with ferric citrate. A distinct microbial community structure was found in patients treated with ferric citrate, with an increased abundance of the Bacteroidetes phylum and a decreased abundance of the phylum Firmicutes. Members of the order Lactobacillales were enriched in patients treated with calcium carbonate, whereas taxa of the genera Ruminococcaceae UCG-004, Flavonifractor, and Cronobacter were enriched in patients treated with ferric citrate phosphate binder. In conclusion, Ferric citrate therapy results in a more diverse microbiome community compared to calcium carbonate therapy in hemodialysis patients with phosphate binder treatment. The gut microbiome reflects the phosphate binder choice in hemodialysis patients …