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Bacteria residing in the human gastrointestinal tract has a symbiotic relationship with its host. Animal models have demonstrated a relationship between exercise and gut microbiota composition. This was the first study to explore the relationship between cardiorespiratory fitness (maximal oxygen consumption, VO 2 max) and relative gut microbiota composition (Firmicutes to Bacteroidetes ratio [F/B]) in healthy young adults in a free-living environment. Twenty males and 17 females (25.7 ± 2.2 years), who did not take antibiotics in the last 6 months, volunteered for this study. VO 2 max was measured using a symptom-limited graded treadmill test. Relative microbiota composition was determined by analyzing DNA extracted from stool samples using a quantitative polymerase chain reaction that specifically measured the quantity of a target gene (16S rRNA) found in Firmicutes and Bacteroidetes. Relationships between F/B and potentially related dietary, anthropometric, and fitness variables were assessed using correlation analyses with an appropriate Bonferroni adjustment ( p  < .004). The average F/B ratio in all participants was 0.94 ± 0.03. The F/B ratio was significantly correlated to VO 2 max ( r  = .48, p  < .003), but no other fitness, nutritional intake, or anthropometric variables ( p  > .004). VO 2 max explained ∼22% of the variance of an individual's relative gut bacteria as determined by the F/B ratio. These data support animal findings, demonstrating a relationship between relative human gut microbiota composition and cardiorespiratory fitness in healthy young adults. Gastrointestinal bacteria is integral in regulating a myriad of physiological processes, and greater insight regarding ramifications of exercise and nutrition on gut microbial composition may help guide therapies to promote human health.