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Since <i>E. coli</i> is the most prevalent sepsis bacterium, studying its pathogenic molecular pathways may help with its early diagnosis and individualized treatment. However, few studies have investigated the molecular characterization of <i>E. coli</i> infection only. We extracted <i>E. coli</i> infection-specific genes and indicators from published data and clinical laboratory results in this study. GSE65088 showed 277, 377, and 408 DEGs for <i>E. coli</i> and other bacteria, <i>E. coli</i> and healthy groups, and other bacteria and healthy groups, respectively. DEGs, the MEgreen module with the highest relevance in WGCNA, and the first three MCODE subnetworks were used to find <i>E. coli</i> infection-specific hub genes. HSPA1B and TNF were verified in GSE6269 with ROC-AUCs of 0.7038 and 0.7116, respectively. CIBERSORT showed increased B-cell naive and T-cell CD4 naive infiltration in <i>E. coli</i> infectious sepsis. Patients infected with <i>E. coli</i> were younger than those infected with other pathogens. Compared to the other bacterially infectious sepsis patients, the <i>E. coli</i> patients had low globulin, prealbumin, creatine kinase, and high bilirubin levels. The clinically significant difference indicator IL-2, in combination with hub genes, better differentiated the healthy and <i>E. coli</i> groups, with an ROC-AUC of 0.8793. The study suggested that HSPA1B and TNF may be <i>E.-coli</i>-infection-specific genes, which may help explain the molecular mechanism of infectious sepsis.