Figure 13.

Phenotypic characterization was performed in T. marneffei lacking immunogenic genes. (A) Macrophage killing assay was used to determine the involvement of cpeA gene during host cell invasion. Macrophage-derived THP-1 human monocytic cell line was infected with conidia of the wild type, cpeA mutant, and complemented strains at MOI 10. After 2-h of incubation, macrophage-infected cells were lysed, and fungus was harvested by centrifugation. The colony forming units (CFUs) isolated from macrophage-infected cells were counted, and the percentage of T. marneffei killed by macrophageswas calculated. Each bar depicted the average and SD values obtained from triplicate experiments. (B) Stress tolerance was assessed in the Δhsp30 mutant. One thousand conidia from the T. marneffei wild-type and Δhsp30 strains were spotted on the surface of minimal medium containing various stressors, including 1 M sorbitol (osmotic stressor), 0.2 M NaCl (hyper-salinity), and 1 mM hydrogen peroxide, H₂O₂ (oxidative stress). The plates were incubated at 37°C for 10 days and the results were photographed. Experiments were performed in three replicates.