Enterococcus mundtii CRL35 as a Dual-Route Treatment for Cutaneous Leishmaniasis: Preclinical Safety and Efficacy Assessment

American tegumentary leishmaniasis (ATL) remains a therapeutic challenge due to the toxicity and limited effectiveness of conventional agents. This study evaluated Enterococcus mundtii CRL35, a food-grade lactic acid bacterium, as a dual-mechanism therapeutic platform against Leishmania (L.) amazonensis. Initial screening identified E. mundtii CRL35 as the most active strain among four enterococci tested, demonstrating high selectivity against promastigotes (SI = 95) and intracellular amastigotes (SI = 25). In a BALB/c murine model of cutaneous leishmaniasis, three treatment modalities were evaluated: topical application of cell-free supernatant (CFS-T), oral viable bacteria (LB-O), and their combination (CFS-T+LB-O). All three treatments achieved significant reductions in lesion parasite burden (approximately 2–3 orders of magnitude versus untreated controls), substantially exceeding the parasitological efficacy of amphotericin B under identical experimental conditions. CFS-T and CFS-T+LB-O additionally produced significant macroscopic lesion improvement. Importantly, no treatment was associated with adverse changes in organ indices or serum biochemical parameters. E. mundtii CRL35-based treatments promoted higher IgG2a/IgG1 antibody ratios compared to untreated controls and amphotericin B, generating an immune profile more favorable for intracellular parasite control. Taken together, these findings support the therapeutic potential of these modalities as safe, effective, and affordable alternatives for ATL, particularly in resource-limited endemic regions where toxicity and logistical barriers limit access to conventional therapy.