The prevalence of Th1/Th2 response, spleen changes and megakaryocytes were investigated in BALB/c mice (n = 138) infected with Leishmania infantum, and treated with Leishmania infantum 30× (10−30) biotherapy – BioLi30×. We performed controlled experiments using 8-to-12-week-old mice, infected with 5 × 107L. infantum promastigotes, divided into eight groups: G1 (healthy), G2 (infected with L. infantum), G3 (BioLi30× pre-treated), G4 (BioLi30× pre/post-treated), G5 (BioLi30× post-treated), G6 (Water 30× post-treated), G7 (Antimonium crudum 30× post-treated) and G8 (Glucantime® post-treated). G3–G7 groups were orally treated with their respective drugs diluted in filtered water (1:10), and G8 received Glucantime® (0.6 mg/100 μl of PBS), intraperitoneally. Spleen fragments were submitted to double blind histopathological evaluation and the number of megakaryocytes was counted. Besides, animals’ serum was measured after 49 days of infection, and cytokines (IFN-γ, IL-4, IL-10, IL-12), as well as the Th1/Th2 correlation (IFN-γ/IL-4 and IFN-γ/IL-10), were analyzed. Spleen histological parameters were classified as: healthy appearance (G1); discreet (G3–G7), moderate (G2) and moderate to severe (G8) white pulp hyperplasia; proliferation of megakaryocytes (G2–G8), and intense disruption (G2–G8). All groups, except for G7, showed higher percentages of megakaryocytes per field ranging from 87% to 15%, when compared to healthy animals (G1). Th1 predominance in IFN-γ/IL-4 ratio (comparing to G2) was detected in G4, G5, G6 and G7. Finally, pre/post (BioLi30x) and post-treatment (Antimonium crudum 30x) presented reduction of megakaryocytes/spleen changes due to immunomodulation animal process, controlling the infection process, probably by the Th1 cytokine predominance.