Chronic fatigue/physical exhaustion (FPE) impacts combat readiness but is difficult to identify. We tested the hypothesis that resting heart rate variability (HRV), including both time- and frequency-domain assessments, would correlate with hydration status and aerobic capacity in military recruit-age men and women with varying fitness levels. Cardiac interbeat intervals were recorded using a heart R-R monitor during 20 minutes of quiet, supine rest with paced breathing (0.25 Hz). HRV metrics included average R-R interval (RRIavg), R-R interval standard deviation (RRISD), the percentage of adjacent R-R intervals varying by > or = 50 ms (pNN50), and integrated areas of R-R interval spectral power at the high (0.15-0.4 Hz) (RRIHF) and low (0.04-0.15 Hz) (RRILF) frequencies. Treadmill maximal oxygen uptake (VO2 max), segmental bioimpedance estimates of total body water (TBW), and urine specific gravity (USG) were also assessed. All dependent variables of interest were within expected ranges, although absolute ranges of individual values were considerable. RRI correlated with VO2 max (r = 0.49; p < 0.001), with TBW (r = 0.38; p < 0.001), and inversely with USG (r = -0.23; p = 0.02). RRISD correlated with VO2 max (r = 0.21; p = 0.03), but not with TBW or USG. pNN50 correlated inversely with USG (r = -0.21; p = 0.03) but not with VO2 max or TBW. R-R interval spectral power at the high and low frequencies did not correlate with VO2 max, TBW, or USG. We have demonstrated that fitness level and hydration status may affect cardiac function via changes in autonomic tone, highlighting the potential of field-based assessment of heart rate variability metrics to identify FPE and other aspects of combat readiness.