Yen-Hui Chiu1, Hsiao-Jan Chen2, Ying-Chen Chang3, Yu-Ning Liu4, Shu-Min Kao2, Mei-Ying Liu2, Ying-Yen Weng3, Kwang-Jen Hsiao5, Tze-Tze Liu6. 1. Department of Education and Research, Taipei City Hospital, Taipei, Taiwan; Department of Biotechnology and Pharmaceutical Technology, Yuanpei University of Medical Technology, Hsinchu, Taiwan. 2. Neonatal Screening Center, The Chinese Foundation of Health, Taipei, Taiwan. 3. Cancer Progression Research Center, National Yang-Ming University, Taipei, Taiwan. 4. Department of Education and Research, Taipei City Hospital, Taipei, Taiwan. 5. Department of Education and Research, Taipei City Hospital, Taipei, Taiwan; Preventive Medicine Foundation, Taipei, Taiwan. Electronic address: hsiao@pmf.tw. 6. Department of Education and Research, Taipei City Hospital, Taipei, Taiwan; Cancer Progression Research Center, National Yang-Ming University, Taipei, Taiwan. Electronic address: ttliu@ym.edu.tw.
Abstract
BACKGROUND: Patients with glucose-6-phosphate dehydrogenase deficiency might develop acute hemolytic anemia, chronic hemolytic anemia, and neonatal hyperbilirubinemia when exposed to high levels of oxidative stress. Severe hemolysis may occur in not only patients but also female carriers under certain conditions. However, 80%-85% of female carriers were undetected in an existing newborn screening program because of their wide-ranging levels of enzyme activity. METHODS: We developed a cost- and time-efficient multiplex SNaPshot assay using dried blood spots. RESULTS: By detecting 21 common mutations in Taiwan and Southeast Asia, the assay could determine 98.2% of the mutant alleles in our cohort of Taiwanese newborns. The 9 undetermined mutant alleles were consequently detected by Sanger sequencing, of which 5 unpublished variations-c.187G > A (Pingtung), c.585G > C (Tainan), c.586A > T (Changhua), c.743G > A (Chiayi), and c.1330G > A (Tainan-2)-were detected. Furthermore, 13% of mild mutations were missed in male infants whose enzyme levels at 6.1-7.0 U/gHb in the newborn screening program when set the cutoff value at 6.0 U/gHb. We therefore suggest increasing the cutoff value and applying the multiplex SNaPshot assay as the second tier for neonatal screening. CONCLUSIONS: Our approach could significantly increase the detection rate of male patients and female carriers with a reasonable cost and a reasonable number of clinic referrals.
BACKGROUND:Patients with glucose-6-phosphate dehydrogenase deficiency might develop acute hemolytic anemia, chronic hemolytic anemia, and neonatal hyperbilirubinemia when exposed to high levels of oxidative stress. Severe hemolysis may occur in not only patients but also female carriers under certain conditions. However, 80%-85% of female carriers were undetected in an existing newborn screening program because of their wide-ranging levels of enzyme activity. METHODS: We developed a cost- and time-efficient multiplex SNaPshot assay using dried blood spots. RESULTS: By detecting 21 common mutations in Taiwan and Southeast Asia, the assay could determine 98.2% of the mutant alleles in our cohort of Taiwanese newborns. The 9 undetermined mutant alleles were consequently detected by Sanger sequencing, of which 5 unpublished variations-c.187G > A (Pingtung), c.585G > C (Tainan), c.586A > T (Changhua), c.743G > A (Chiayi), and c.1330G > A (Tainan-2)-were detected. Furthermore, 13% of mild mutations were missed in male infants whose enzyme levels at 6.1-7.0 U/gHb in the newborn screening program when set the cutoff value at 6.0 U/gHb. We therefore suggest increasing the cutoff value and applying the multiplex SNaPshot assay as the second tier for neonatal screening. CONCLUSIONS: Our approach could significantly increase the detection rate of male patients and female carriers with a reasonable cost and a reasonable number of clinic referrals.