Persistent antigen is essential for sustaining Leishmania -specific memory CD4 ⁺ T cells and long-term immunity

Memory T cells are critical for secondary immunity against pathogens, yet their persistence in the absence of antigen remains unclear. While memory CD8 + T cells are known to persist independently of their cognate antigen, the durability of memory CD4 + T cells in the absence of antigen remains controversial. Recovery from cutaneous leishmaniasis confers lifelong immunity, largely mediated by CD4 + T cells, but the necessity of persistent parasites for sustaining this immunity has not been empirically confirmed. We investigated immunity in mice infected with a dihydrofolate reductase-thymidylate synthase ( dhfr-ts )-deficient Leishmania major , which cannot persist due to their thymidine salvage deficiency. These mice lost protection against wild-type challenge, correlating with a decline in Leishmania (PEPCK)-specific CD4 + T cells. To further dissect this relationship, we generated PEPCK-specific CD4 + TCR transgenic (PEG) mice, enabling precise tracking of Leishmania -specific memory CD4 + T cells. Both in vitro and in vivo -generated memory PEG cells gradually disappeared over time in the absence of antigen, irrespective of the host’s MHC II status, and this loss paralleled the erosion of infection-induced immunity. PEG and endogenous PEPCK-specific memory cells were not maintained in mice infected with dhfr-ts - or PEPCK -deficient L. major , resulting in loss of recall responses and secondary immunity. These findings demonstrate that continuous antigen presence is crucial for maintaining Leishmania -specific memory CD4 + T cells and highlight the role of persistent antigen in sustaining long-term immunity against chronic infections.