Phenotypic and Genotypic Analysis of Two Rare Cases of --SEA/-α4.3 HbH Disease.

Abstract

Background: To report two cases of hemoglobin H (HbH) disease with routine genetic testing missed rare variants, where the phenotype was inconsistent with routine genetic testing results, explore their true genotypes and clinical characteristics, and evaluate the application value of single-molecule nanopore sequencing (CycloneSEQ platform) in detecting rare thalassemia variants.

Methods: Common thalassemia gene testing was performed on two adult male patients who presented with different complications. The results showed --^SEA/αα, which was significantly inconsistent with their hematological parameters. Subsequently, long-read sequencing of the α-globin gene cluster was conducted using the CycloneSEQ platform (single-molecule nanopore sequencing technology), and specific primers were designed based on the results for PCR verification.

Results: The two patients were ultimately diagnosed with rare deletional HbH disease with the genotype --^SEA/-α^4.3 (NC_000016.10:g.176935-181274DEL). The deletion range includes the α1 gene and is accompanied by an X-Y-Z segment recombination, which is reported here for the first time. Both patients exhibited varying degrees of anemia and complications, suggesting that the same genotype can manifest significant phenotypic heterogeneity.

Conclusion: Conventional thalassemia testing fails to identify rare deletional HbH disease, frequently causing genotype-phenotype discordance and diagnostic bias. The newly reported --^SEA/-α^4.3 genotype with X-Y-Z recombination expands the HbH disease genotypic spectrum and confirms obvious phenotypic heterogeneity. CycloneSEQ nanopore sequencing effectively detects rare complex variants undetectable by routine methods, supporting precise diagnosis of atypical thalassemia with favorable clinical prospects.