A clinical triage score for MLPA testing in children with microcytosis suspected of deletional alpha-thalassemia.

UNLABELLED: Alpha-thalassemia lacks an HbA2-like hematological surrogate to indicate which children with microcytosis warrant molecular testing. We developed and internally validated a Clinical Triage Score to prioritize multiplex ligation-dependent probe amplification (MLPA) referral to detect deletional alpha-thalassemia carriers. In this retrospective single-center study, 187 of 201 children referred for alpha-globin MLPA testing (2020-2025) were analyzed. The endpoint was an alpha-globin deletion detected by MLPA. Pre-specified predictors were entered by forced entry into multivariable logistic regression, with points assigned by Sullivan rescaling. Internal validation used bootstrap optimism correction (B = 1000), and decision curve analysis assessed clinical utility. Alpha-globin deletions were detected in 70 children (37.4%), with α⁰-thalassemia trait predominating (62.9%), and - (α)20.5 was the predominant deletion (45.7%). Family history (adjusted OR 5.68), age-adjusted erythrocytosis (8.96), and normal ferritin (3.55) were independent predictors; erythrocytosis received two points, the others one each (range 0 to 4). The AUROC was 0.814 (95% CI 0.739-0.890). At the Youden-optimal threshold ≥ 2, sensitivity was 87.1% and specificity 53.8%, missing 9 (12.9%); at ≥ 1, sensitivity reached 95.7%. Sensitivity was highest for the 48 clinically significant two-gene deletions (89.6%). Decision curve analysis suggested net benefit over test-none, and over test-all above an 11% threshold.

CONCLUSION

This preliminary, internally validated score may help prioritize MLPA referral in children with microcytosis and performs best for clinically significant two-gene genotypes. Because HBA1/HBA2 sequencing was not routinely performed, a low score does not exclude alpha-thalassemia, particularly non-deletional forms. External prospective validation is required before clinical adoption.

WHAT IS KNOWN: • Alpha-thalassemia has no HbA2-like hematological marker, so carrier detection depends on molecular testing. • No validated triage tool exists to prioritize MLPA referral in children with microcytosis.

WHAT IS NEW: • A TRIPOD-compliant three-variable score (family history, erythrocytosis, normal ferritin) predicted MLPA-detected deletions (AUROC 0.814), with net benefit over test-none and, above an 11% threshold, over test-all. • The spectrum was α⁰-dominant, and sensitivity was highest (89.6%) for the clinically significant two-gene deletions.