The effectiveness and mechanisms of antitumor activity of non-pathogenic Escherichia coli EM0 strain using mouse model of Ehrlich ascites carcinoma (EAC) is studied. 48 h after inoculating EAC cells, a single noninvasive treatment of the 2-month-old male white mice’s eyes and mouths with live E. coli isolate increased the life span by 75% (P<0.001). Furthermore, a significant decrease in the volume of ascites fluid (66.5 %, P<0.01) was determined, accompanied by down-regulation of EAC-activated lipid peroxidation processes and changes in the L-arginine metabolic profile in leukocytes as early as within 9 days of post-treatment compared to non-treated EAC-bearing mice. We found EAC-induced stimulation of arginase and nitric oxide synthase (NOS) activity correlated with the levels of their common substrate, L-arginine and products (arginase-derived L-ornithine and NOS-derived NO and L-citrulline) in the cytoplasm and mitochondria of peritoneal and blood leukocytes. The biochemical pattern was differentially modulated by E.coli treatment depending on whether leukocytes were localized in the ascitic fluid or the peripheral blood. E. coli concentrated in the ascitic fluid directly affected the surrounding cells including peritoneal leukocyte in which it decreased the activity of two arginase isoforms and a total NOS in the cytoplasm. Negligible number of E coli at sites remote of tumor suggests its indirect effects particularly via stimulation of LPS-mediated non-specific immune response associated with activation of arginase and NOS in the cytoplasm of blood leukocytes. The data obtained should be taken into account in the further study aimed to use non-pathogenic E. coli strains in the adjuvant therapy of ascites tumors.