The influence of intestinal bacterial decontamination on the occurrence of grades II to IV acute graft-versus-host disease (GVHD) was retrospectively analyzed in 194 predominantly adult patients treated by genotypically identical sibling marrow transplantation under conditions of strict protective isolation and intestinal antimicrobial decontamination. Forty-five patients (23%) developed acute GVHD and univariate analysis identified four features that significantly increased the risk for this reaction: chronic myeloid leukemia as the underlying disease, as compared with all other disease categories (P < .0001); female marrow donors for male recipients, as compared with other gender combinations (P < .005); ineffective, as compared with sustained growth suppression of intestinal anaerobic bacteria (P < .006); and methotrexate as the sole immunoprophylactic compound, as compared with cyclosporine containing regimens (P < .05). Using the duration of anaerobic growth suppression as a time-dependent explanatory variable, proportional hazards regression analysis confirmed these features as independent predictors for acute GVHD with relative risk estimates of 1.9 (95% confidence interval [CI], 1.3 to 2.7) for the immunoprophylactic regimen (P < .0004), of 1.8 (95% CI, 1.3 to 2.5) for the underlying disease (P < .0005), of 1.7 (95% CI, 1.2 to 2.5) for anaerobic decontamination (P < .002), and of 1.3 (95% CI, 1.1 to 1.6) for the donor/recipient gender combination (P < .008), respectively. Best subset selection modeling also identified the quality of anaerobic decontamination as the third most important predictor for acute GVHD, when all four significant features were included. Estimates of acute GVHD stratified by the quality of anaerobic bacterial growth suppression showed a strong influence of anaerobic decontamination in patients burdened by at least one of the other unfavorable factors (P < .009). In conclusion, this study provides strong evidence that sustained growth suppression of intestinal anaerobic bacteria after clinical sibling marrow transplantation can independently modulate the occurrence of grades II to IV acute GVHD, which is in concordance with previous results from animal transplantation models. Antimicrobial chemotherapy specifically targeted to the intestinal anaerobic bacterial microflora may be complementarily useful in preventing acute GVHD and should be investigated in a prospective trial.