: The study aimed to examine physiological responses of firefighters performing a firefighting simulation test (FST) and to determine the relationship between physical fitness parameters and FST performance. Aerobic fitness, muscular strength, muscular endurance, and anaerobic capabilities were evaluated in 37 firefighters (21-profesionals and 16-volunteers firefighters). Furthermore, participants carried out the FST during which we measured performance, respiratory gas exchange, heart rate (HR), perceived exertion and blood lactate concentrations. Males were significantly faster than females for all tasks of the FST (p < 0.01); however, final performance score (16.5 ± 2.9 and 14.5 ± 2.6 points for males and females, respectively), HR (94.0 ± 2.0% and 93.7 ± 2.3% of HRmax) and perceived exertion (8.1 ± 0.9 and 7.1 ± 1.3) were not significantly different. Prediction of FST performance by LASSO regression revealed a model that included mainly aerobic capacity and maximal strength. In conclusion, FST challenged both aerobic and anaerobic energy metabolisms for both males and females and requires various physiological abilities to perform.