Background: β-thalassemia is the most common monogenic disease caused by abnormalities in the synthesis of the β-chain of hemoglobin. Methods: From January 2018 to September 2018, 90 patients (age >18 y) with β-thalassemia major or intermedia who referred to Rajaei Cardiovascular, Medical, and Research Center, Tehran, Iran, for the assessment of myocardial iron overload were enrolled. All the patients were receiving regular blood transfusions and chelating therapy. Comprehensive transthoracic echocardiographic studies consisting of 2D echocardiography, tissue Doppler imaging, and real-time 3D echocardiography were performed. Results: A total of 90 patients were enrolled in the study. Cardiac iron toxicity (ie, T2* < 20 ms) was seen in 28 (31%) patients; whereas in 62 (69%) patients, the cardiac iron level was undetectable (ie, T2* > 20 ms). Patients with T2* < 20 ms had significantly higher serum ferritin levels than those with T2* > 20 ms (P = 0. 02). No significant correlation was found between the serum ferritin level and T2* (r = − 0. 08, P = 0. 41). The left ventricular ejection fraction was statistically similar in the 2D and 3D examinations. Left atrial endsystolic and end-diastolic volumes were greater in the patients with iron cardiotoxicity than in those with no detectable cardiac iron deposition (P = 0. 01 and P <0. 001, respectively). Left atrial strain was also significantly lower in the patients with critical iron overload. The patients with T2* < 20 ms also had lower left atrial ejection fractions than those with T2* >20 ms, both in 2D and 3D examinations (both Ps <0. 001). Conclusions: Our study showed that changes in the left atrial structure and function precede impairment in the left ventricular systolic function in thalassemia patients with critical myocardial iron loading.