Antileishmanial effect of thymol on Leishmania tropica isolated from Turkey

Objective: Cutaneous leishmaniasis (CL) is a clinical manifestation characterized by long-lasting nodulo-ulcerative wounds and healing with atrophic scars caused by protozoan parasites of the Leishmania species. The high cost and toxic effects of drugs used in the treatment of CL, along with the development of resistance to these drugs, have led to the necessity of investigating innovative therapeutic agents that possess effective antileishmanial activity while stimulating the adaptive immüne system of host cells. The aim of this study was to investigate the antileishmanial activity and immunomodulatory properties of the active substance thymol against Leishmania tropica isolated from Türkiye in in vitro and ex vivo experimental models. Materials and Methods: The Leishmania isolate obtained from a CL patient in Manisa province, stored in the Parasite Bank of Manisa Celal Bayar University Faculty of Medicine, was identified as Leishmania tropica (MHOM/TR/2012/CBCL-LT) through species determination using real-time PCR with ITS1 primers. The cytotoxic activity (CC50) of thymol was determined using the CellTiter-Glo method, while the in vitro antileishmanial activity against L. tropica promastigotes (IC50promastigote) was evaluated using both CellTiter-Glo and hemositometry methods. The ex vivo antileishmanial activity against L. tropica amastigotes (IC50amastigote) was determined through microscopic examination of Giemsa-stained preparations, parasite rescue and transformation assay. Cytokine secretions (IFN-γ, IL-4, IL-10, and IL-12) were evaluated using the ELISA method. Selectivity index (SI) values were calculated using obtained CC50, IC50promastigote, and IC50amastigote values. Results: The CC50 value of thymol was determined to be 160.7 μM/mL, indicating no cytotoxic activity. The in vitro antileishmanial activity against L. tropica promastigotes (IC50promastigote) was found to be 79.41 μM/mL, while the ex vivo antileishmanial activity against L. tropica amastigotes (IC50promastigote) was 105.2 μM/mL. The calculated SI values using CC50, IC50promastigote, and IC50promastigote values were determined to be 2.02 and 1.53 for L. tropica promastigotes and amastigotes, respectively. Additionally, compared to THP-1 macrophage cells infected with L. tropica alone, an increase in IFN-γ and IL-12 cytokines was observed in infected THP-1 macrophage cells treated with thymol, while a decrease was observed in IL-4 and IL-10 cytokines. Conclusion: The demonstration of antileishmanial activity of thymol against L. tropica positions it as a potential candidate for further investigation as a therapeutic agent in CL. The immunomodulatory properties of thymol highlight it as a novel and effective option in the treatment of leishmaniasis. This is regarded as a significant and promising step in the development of new drugs that can be used in the treatment of leishmaniasis.