Six strains belonging to five species of Polyporus ( P. arcularius, P. arcularioides , P. tricholoma, P . cfr. tricholoma , and P. varius ), collected from an Atlantic Forest area in Misiones (Argentina), where species usually grow exposed to high temperatures and humidity, were identified by morphological and molecular analyses. P. tricholoma (BAFC 4536) and P. arcularioides (BAFC 4534) were selected by their lignin‐degrading enzyme production, their ability to produce primordial of basidiomes under submerged fermentation, and the decrease in lignin content caused in Poplar wood (up to 29% after 45 days). Among several variables evaluated with a Plackett–Burman design (glucose, copper, vanillic acid and manganese concentration, incubation period, and light incidence), the most important factor affecting laccase and Mn‐peroxidase (MnP) production by both strains, was light incidence. Light induced fruit body development but diminished laccase and MnP production. Moreover, a modified isoenzymatic laccase pattern was observed, showing additional isoenzymes when fungi were cultivated under darkness and differences in optimal temperature. Although the studied strains did not produce high laccase and MnP titers (uppermost detected 4230 and 90 U L −1 , respectively), their laccases showed thermal stability and optimal temperature above 70 °C, representing an interesting source in the search of thermo‐tolerant enzymes for biotechnological applications.