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0 reviewsSUMMARYMicrobial alterations in different tumor locations of colorectal cancer (CRC) remain unclear. Here, 1,375fecal metagenomes from six in-house and published datasets were analyzed, including 128 right-sidedCRC (rCRC), 168 left-sided CRC (lCRC), 250 rectal cancer (RC), and 829 controls. Firmicutes progressively increase from rCRC, lCRC, to RC, in contrast to the gradual decrease of Bacteroidetes. Tumorlocation-associated fecal microbes are identified, including Veillonella parvula for rCRC, Streptococcusangionosus for lCRC, and Peptostreptococcus anaerobius for RC, while Fusobacterium nucleatum isenriched in all tumor locations. Tumor location-associated bacteria correlate with patient survival. Clinically, we establish a microbial biomarker panel for each tumor location that accurately diagnosesrCRC (area under the receiver operating characteristic curve [AUC] = 91.59%), lCRC (AUC = 91.69%),or RC (AUC = 90.53%) from controls. Tumor location-specific biomarkers also have higher diagnostic accuracy (AUC = 91.38%) than location-non-specific biomarkers (AUC = 82.92%). Overall, we characterizefecal microbes associated with different CRC tumor locations, highlighting that tumor location should beconsidered in non-invasive diagnosis.