Objective <p>This study aimed to investigate the effects of slow deep breathing (SDB) on heart rate variability (HRV) in patients with type 2 diabetes mellitus (T2DM) patients with cardiovascular autonomic neuropathy (CAN).</p> Methods <p>Sixty patients with CAN secondary to type T2DM were randomly assigned to SDB (<i>n</i> = 30) and control (<i>n</i> = 30) groups. All patients were treated with mecobalamin (0.5&#xa0;mg; times daily). Patients in the SDB group underwent slow deep breathing training against a background of soothing music. Before treatment and 3 months after treatment, height, body weight, heart rate (HR), blood pressure (BP), HbA1c, LDL, TG, heart-rate-variability (HRV), and hart rate recovery (HRR) were measured 1&#xa0;min after the 6-min walk test in the two groups.</p> Results <p>BMI, SBP, DBP, HbA1c, LDL, and TG levels were comparable between the two groups and remained unchanged after treatment in two groups (<i>P</i> &gt; 0.05). In the SDB group, the standard deviation of N-N intervals (SDNN), the square root of the mean squared differences of successive RR (RMSSD), low-frequency power (LF), high-frequency power (HF), and LF/HF ratio changed significantly after treatment (<i>P</i> &lt; 0.05), and these measurements remained statistically significant after adjusting for age (<i>P</i> &lt; 0.05), but HR and the root mean square of the difference between adjacent R-R intervals (PNN50) remained unchanged (<i>P</i> &gt; 0.05). In the control group, HR, SDNN, RMSSD, PNN50, LF, HF, and LF/HF ratio remained unchanged after treatment (<i>P</i> &gt; 0.05). The SBP, SDNN, RMSSD, LF, HF, and LF/HF ratios were significantly different between the two groups after treatment.(<i>P</i> &lt; 0.05), and these measurements remained statistically significant after adjusting for age (<i>P</i> &lt; 0.05). In addition, the HRR1 levels were comparable between the two groups (<i>P</i> &gt; 0.05).</p> Conclusion <p>With soothing music as the background, <b>s</b>ustained slow deep breathing training can improve heart rate variability in patients with type 2 diabetes mellitus and cardiac autonomic neuropathy, independent of age, providing a new approach for the treatment of cardiac autonomic neuropathy in type 2 diabetes.</p> Clinical trial number <p>The study was registered in the Chinese Clinical Trial Registry (ChiCTR2200063260).</p>