Background: Changes that may occur in intestinal microbiota have been shown to result in abnormal levels of intermediates, which can adversely affect the cardiovascular status. Objectives: We aimed to study trimethylamine-N-oxide level (TMAO) in patients with chronic systolic dysfunction as a marker of intestinal dysbiosis and correlate it with cardiac status and echocardiographic findings. Patients and methods: Twenty-five children with chronic cardiac dysfunction underwent sample collection for TMAO level. They have been compared to twenty-five age and sex-matched controls. Patients were assessed with conventional transthoracic echocardiography and three-dimensional speckle-derived strain imaging.  Results: Median age was 4.5 years. Patients had significantly lower ejection fraction (EF) and fractional shortening (FS) (P <0.001), and lower global, radial, circumferential and longitudinal 3-D strain rates (p < 0.001). TMAO was significantly higher in patients (p < 0.05). Best cut-off value was >6.2 um with sensitivity= 96% and specificity= 100%. TMAO correlated with Ross class, hospitalization rate as well as EF, FS, LV dimensions and volumes, and 3 D circumferential and radial strain rates. Linear regression analysis showed circumferential strain being an independent predictor of TMAO levels in the blood. Conclusions: TMAO levels are increased in patients with chronic systolic dysfunction, which is related to their clinical status as well as their echocardiographic parameters. Three-dimensional circumferential strain rate seems to be an independent predictor of TMAO levels.