Combined Gap-Polymerase Chain Reaction and Targeted Next-Generation Sequencing improve alpha- and beta-Thalassemia Carrier Screening in Pregnant Women in Vietnam

Vietnam has a high thalassemia burden. We collected blood samples from 5880 pregnant Vietnamese women during prenatal health checks to assess thalassemia carrier frequency using combined gappolymerase chain reaction (gap-PCR) and targeted next-generation sequencing (NGS). Thalassemia carriers were identified with prevalence of 13.13% (772), including 7.82% (460) carriers of alpha-thalassemia (alpha-thal), 5.31% (312) carriers of beta-thalassemia (beta-that), and 0.63% (37) concurrent alpha-/beta-thal carriers. Deletional mutations (368) accounted for 80.0% of alpha-thal carriers, of which, --(SEA) (Southeast Asian) (n = 254; 55.0%) was most prevalent, followed by the -alpha(3.7) (rightward) (n = 66; 14.0%) and -alpha(4.2) (leftward) (n = 45; 9.8%) deletions. Hb Westmead (H8112: c.369C>G) (n = 53) and Hb Constant Spring (Hb CS or HBA2: c.427T>C) (in 28) are the two most common nondeletional alpha-globin variants, accounting for 11.5 and 6.0% of alpha-thal carriers. We detected 11 different beta-thal genotypes. Hb E (HBB: c.79G>A) (in 211) accounted for 67.6% of beta-thal carriers. The most common beta-thal genotypes were associated with mutations at codon 17 (A>T) (H86: c.52A>T), codons 41/42 (-TTCT) (H88: c.126_129delCTTT), and codon 71/72 (+A) (HBB: c.217_218insA) (prevalence 0.70%, 0.68%, and 0.2%, respectively). Based on mutation frequencies calculated in this study, estimates of 5021 babies in Vietnam are affected with clinically severe thalassemia annually. Our data suggest a higher thalassemia carrier frequency in Vietnam than previously reported. We established that combining NGS with gap-PCR creates an effective large-scale thalassemia screening method that can detect a broad range of mutations.