Insufflation of ozonized oxygen into the peritoneum (O3/O2-pneumoperitoneum [O3/O2-PP]) of rats reduced the lethality of peritonitis. We evaluated the prophylactic effect of O3/O2-PP combined with tazobactam/piperacillin (TZP) in polymicrobial lethal peritonitis. Wistar rats were conditioned by daily repeated insufflation of ozone for 5 days, and hematologic parameters were determined. Sepsis was induced by i.p. injection of cecal material derived from donor rats. Simultaneously, TZP was applied at a single dosage of 65 mg/kg or at two dosage schedules of 65 mg/kg each at an interval of 1 h. The conditioning effect of O3/O2-PP on the number of blood cells was measured before inoculation of bacteria. The mRNA levels of proinflammatory cytokine IL-lbeta and TNF-alpha were determined at 4 h post infection in spleen and liver by semiquantitative in situ hybridization analysis. Preconditioning of rats by O3/O2-PP enhanced the number of blood leukocytes and granulocytes and increased the survival rate of septic rats up to 33%. The combination of O3/O2-PP and TZP further enhanced the survival rate up to 93%. This effect was accompanied by a reduced amount of IL-1beta and TNF-alpha mRNA in spleen and liver. In contrast, in non-infected animals the combination of O3/O2-PP and TZP enhanced IL-1beta and TNF-alpha mRNA in the spleen and IL-1beta mRNA in liver when compared with TZP- and sham-treated controls. The preconditioning effect of O3/O2-PP seems to support the biological effectiveness of TZP by altering the immune status before and during the onset of sepsis. The combined therapy could be a simple, preoperative intervention for abdominal surgery to reduce postoperative morbidity and mortality.