BACKGROUND: Mountain hiking is an unpredictable activity for firefighters during the search, rescue, and firefighting. Could the quality of the uniform worn by firefighters affect psychophysiological variables? OBJECTIVE: The objective of this study was to monitor the responses of perceived exertion (RPE), musculoskeletal pain (MP), and heart rate variability (HRV) responses in firefighters using different uniforms on a 5 km mountain walking activity. The feeling of comfort, safety, and ergonomic adjustment of the two types of uniforms also was verified. METHODS: Sixteen firefighters were divided into two groups: 4thRUPM"B2" (33% polyester and 67% cotton) vs. GOLD (50% polyamide and 50% cotton). The GOLD group wore a prototype that had not yet been operationally tested. RESULTS: The independent t-test showed that the groups had similar results in RPE at post-march (Omni-Walk/Run = 3.5+/-0.9 vs. 2.9+/-0.6 AU). ANOVA Two-way (2x group and 2x time) was used to verify differences in body mass (BM), MP, and HRV. There was a main effect of time (pre vs. post-march; p < 0.05) for BM, MP, heart rate (HR), mean RR, RMSSD, pNN50, and HF, regardless of group (4thRUPM"B2" vs. GOLD). Rainwater increased post-march BM (92.2+/-9.8 kg vs. 93.5+/-9.9 kg). MP (28.9+/-2.9 vs 33.2+/-7.7 AU), HR (71.6+/-14.2 vs 99.0+/-26.3 bpm), RMSSD (74.8+/-53.7 vs 126.5+/-74.4 ms) and HF (48.5+/-20.2 vs 60.0+/-16.7 nu) also increased post-march. The mean RR (875.2+/-153.3 vs. 673.9+/-155.3 ms) and pNN50 (33.1+/-18.6 vs. 46.3+/-23.9%) reduced post-march. There was a subjective preference for GOLD. CONCLUSION: Different uniforms do not compromise psychophysiological variables during the mountain hike but differentiate ergonomic subjectivity.