BACKGROUND: Leishmaniasis remains a significant global health concern, ranking among the top ten infectious diseases and causing substantial mortality and socioeconomic burden. Effective and accessible treatments are needed. This study investigated the potential of a hydroalcoholic extract from readily available urban green algae as an anti-leishmanial agent, focusing on its impact on key weight-related indicators of Leishmania major infection in BALB/c mice. To evaluate the in vivo anti-leishmanial activity of the hydroalcoholic extract from the common green algae genus Spirogyra against Leishmania major in BALB/c mice, specifically by assessing its effects on weight loss, lesion size, liver weight, and spleen weight-key indicators of disease progression. METHODS: Spirogyra algae were collected and identified in Yazd Province, Iran. A hydroalcoholic extract was prepared and administered via intraperitoneal injection into Leishmania major-infected BALB/c mice at doses of 3, 6, and 12 mg/kg/day, starting after lesion development. The control groups included untreated infected mice (negative control), healthy uninfected mice (control), and infected mice treated with Glucantime (positive control). We assessed treatment efficacy by monitoring weight loss, lesion diameter, liver weight, and spleen weight. RESULTS: Treatment with the highest concentration of Spirogyra extract (12 mg/kg/day) significantly mitigated weight loss in infected mice, demonstrating comparable efficacy to Glucantime. Both the 12 mg/kg/day algae extract and Glucantime significantly controlled lesion growth. Importantly, both treatments significantly reduced liver and spleen weight compared with the negative control group, indicating a reduction in organomegaly. Specifically, the negative control and 3 mg/kg extract groups exhibited the highest liver weights, whereas the negative control group showed significantly higher spleen weights than the other groups. The 12 mg/kg extract and Glucantime groups showed liver and spleen sizes comparable to the healthy control group, demonstrating effective control of organ size changes associated with leishmaniasis. CONCLUSION: The hydroalcoholic extract of urban Spirogyra green algae, particularly at a dose of 12 mg/kg/day, exhibited significant in vivo anti-leishmanial activity in BALB/c mice. Evaluated through weight indicators such as reduced weight loss, controlled lesion growth, and normalized liver and spleen weights, the extract showed promise in mitigating the detrimental effects of Leishmania major infection and warrants further investigation as a potential source for novel anti-leishmanial therapeutics.