Leishmania (Viannia) braziliensis is the main etiological agent of cutaneous leishmaniasis in Brazil. The Thor strain, isolated in the state of Rio de Janeiro, presents subpopulations with distinct biological profiles. Considering the role of surface proteases in parasite virulence and adaptation, this study aimed to compare the enzymatic profiles of promastigotes and axenic amastigotes of the Thor strain and three subpopulations: Thor03, Thor10, and Thor22.

Promastigote and axenic amastigote forms were treated with phospholipase C (PLC) for selective release of GPI-anchored surface proteases. Proteolytic activity was analyzed by gelatin zymography, fluorimetry using substrates specific for metalloproteases, cysteine and serine proteases, and assays with selective inhibitors. Quantification was based on the rate of hydrolysis of fluorescent substrates.

All subpopulations showed distinct proteolytic activity profiles. Metalloprotease activity was predominant, especially in Thor03 amastigotes. Significant cysteine and serine protease activities were also observed, with variable patterns among forms and subpopulations. The enzymatic profiles suggest a functional role in early infection and host adaptation.

Differences in surface protease activity among Thor strain subpopulations highlight the relevance of these enzymes as virulence factors. Subpopulation Thor03 showed greater enzymatic potential in the amastigote form, indicating a likely contribution to infectivity. These findings reinforce the use of proteases as molecular markers and potential therapeutic targets in cutaneous leishmaniasis.