This prospective observational study conducted in a neonatal intensive care unit aimed to evaluate echocardiographic changes provoked by anemia and transfusion of packed red blood cells (pRBCs) in premature infants. In this study, 32 anemic premature infants had serial echocardiographic assessment of left ventricular (LV) systolic performance, LV preload, and afterload immediately before, within 48 h, and up to 120 h after the transfusion of pRBCs. Pretransfusional evaluations also were compared with similar assessments of 71 nonanemic inpatient premature infants analogous for sex, gestational age at birth, and postnatal age. Left ventricular systolic performance was estimated from fractional shortening, LV output, and LV myocardial performance index (LVMPI). The LV preload was estimated from the LV end-diastolic dimension and the ratio of left atrium-to-aortic root dimension (LA/Ao ratio). The LV afterload was estimated from end-systolic wall stress. The LVMPI was found to decrease with increasing corrected gestational age in both the nonanemic (R = 0.173; p = 0.03) and anemic (R = 0.460; p = 0.007) infants. The LVMPI was the only index that changed after transfusion of pRBCs, decreasing in the younger anemic infants (p = 0.011) and increasing in the older anemic infants (p = 0.012). Finally, a significant inverse relationship between pre- and posttransfusional LVMPI values (R = 0.730; p < 0.001) was noted. The LVMPI may allow for identification of preterm infants more likely to be helped by transfusion of pRBCs.