Objectives: This study investigated the effects of breathing at resonance frequency (RF) on the autonomic nervous system (ANS) and adaptability to acute stress. Stress is known to lead to autonomic system dysfunction, with adverse effects on physiological and mental well-being. Resonance breathing is well documented to have positive effects on heart rate variability (HRV), an important biomarker representing autonomic balance. To date, a smaller volume of research has studied whether the short-term practice of the breathing exercise can influence the ANS response to acute stress, measured via HRV parameters. Methods: An independent measures design was conducted on 20 healthy adults after obtaining approval from the University Ethics Committee and written informed consent from the participants. Participants were randomly split into two groups, RF breathing group (n=10) and control group (n=10). The intervention group breathed at RF (standardised cycle of 6 breaths/minute) for 3 minutes, whereas the control sat quietly for the relaxation period. After 3 minutes, all groups participated in a mental arithmetic test for 5-minutes, and then sat quietly during a 5-minute recovery period. Mean heart rate (HR) and HRV parameters (RMSSD, LF, HF, LF/HF) were measured at baseline and throughout the experiment. Results: The two groups were not significantly different in mean HR, RMSSD, LF and HF measures during relaxation and stress periods (p>0.05). The intervention group observed a significantly improved LF/HF ratio during the relaxation phase (p<0.05); however, no significant difference was transferred during the maths task. All participants experienced an increase in mean HR and reduction in RMSSD during this stress phase. Conclusion: Clearly the ANS is adversely affected when placed under a stressful task. Breathing at RF when relaxing can improve balance in the ANS (LF/HF ratio), however no evidence was found to support that RF breathing can improve one’s physiological stress response. Nonetheless, the results during relaxation support future research investigations with greater sample sizes and the exercise implemented for a longer period of time in order to generate more meaningful results on stress adaptation.