ObjectiveThe relationship between intestinal microbiome and colorectal cancer (CRC) progression is unclear. This study aims to identify the intestinal microbiome associated with CRC progression and construct predictive labels to support the accurate assessment and treatment of CRC.MethodThe 192 patients included in the study were divided into stage I-II and stage III-IV CRC patients according to the pathological stages, and preoperative stools were collected from both groups for 16S rDNA sequencing of the intestinal microbiota. Pearson correlation and Spearman correlation coefficient analysis were used to analyze the differential intestinal microbiome and the correlation with tumor microenvironment and to predict the functional pathway. XGBoost model (XGB) and Random Forest model (RF) were used to construct the microbiome-based signature. The total RNA extraction from 17 CRC tumor simples was used for transcriptome sequencing.ResultThe Simpson index of intestinal microbiome in stage III-IV CRC were significantly lower than those in stage I-II CRC.Proteus,Parabacteroides,AlistipesandRuminococcusetc. are significantly enriched genus in feces of CRC patients with stage III-IV. ko00514: Other types of O − glycan biosynthesis pathway is relevant with CRC progression.Alistipes indistinctuswas positively correlated with mast cells, immune activators IL-6 and IL6R, and GOBP_PROTEIN_FOLDING_IN_ENDOPLASMIC_RETICULUM dominantly. The Random Forest (RF) model and eXtreme Gradient Boosting (XGBoost) model constructed with 42 CRC progression-associated differential bacteria were effective in distinguishing CRC patients between stage I-II and stage III-IV.ConclusionsThe abundance and diversity of intestinal microbiome may increase gradually with the occurrence and progression of CRC. Elevated fetal abundance ofProteus,Parabacteroides,AlistipesandRuminococcusmay contribute to CRC progression. Enhanced synthesis of O − glycans may result in CRC progression.Alistipes indistinctusmay play a facilitated role in mast cell maturation by boosting IL-6 production.Alistipes indistinctusmay work in the correct folding of endoplasmic reticulum proteins in CRC, reducing ER stress and prompting the survival and deterioration of CRC, which may owe to the enhanced PERK expression and activation of downstream UPR byAlistipes indistinctus. The CRC progression-associated differential intestinal microbiome identified in our study can be served as potential microbial markers for CRC staging prediction.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12967-023-04119-1.