The first Chinese case of unstable Hemoglobin Santa Ana detected by capillary electrophoresis: a case report and literature review.

Hemoglobin Santa Ana [ß88(F4)Leu→Pro (CTG > CCG) HBB: c.266T > C] is an unstable hemoglobin variant characterized by a substitution of the amino acid leucine by proline at the 88th position of the ß-globin chain. We for the first time identified this hemoglobin variant in a Chinese patient by capillary electrophoresis (CE). The proband was an 8-year-old boy with chronic anemia, brown urine and splenomegaly. He had been affected by moderate anemia, twice approaching a severe degree, that was attributed to infection. The CE result revealed an abnormal hemoglobin peak at electrophoretic zone 4 that correspond to the hemoglobin Santa Ana peak, and a CTG > CCG mutation at codon 88 of the ß-globin gene was confirmed by DNA sequencing. To avoid misdiagnosis and genetic risks, a literature review of other unstable hemoglobins that migrate similarly to the hemoglobin Santa Ana was performed. Our findings indicate that hemoglobin Santa Ana can be clearly separated by CE, with accurate diagnosis depending on molecular analysis. This information will be useful for providing appropriate genetic counselling and for prenatal diagnosis.

[Differential Diagnosis of Three Commonest Deletion ß-Thalassemia in Chinese].

OBJECTIVE: To analyze the hematological characteristics of Chinese Gγ+(Aγδß)0-thalassemia，SEA-HPFH and Taiwan type ß-thalassemia. METHODS: Hemoglobin electrophoresis and blood routine test were used to analyze the hematological indexes of all peripheral blood samples，PCR-Flow fluorescent hybridization and Gap-PCR were used to detect the globin gene mutations and the data were analyzed statistically. RESULTS: The 3 types of deletion ß- Thalassemia patients were showed as hypochromic small cell anemia. The MCH and MCV values of Taiwan type ß-thalassemia patients were the lowest. The results of hemoglobin electrophoresis showed that the increasing of HbF was found in all of the 3 types. Except for the decreasing of Hb A2 in Chinese Gγ+(Aγδß)0-thalassemia，the levels of Hb A2 in the other two deletion ß-thalassemia patients were significantly increased. Except for Hb, there were significant differences in MCV, MCH, Hb A2 and HbF between Chinese Gγ+(Aγδß)0-thalassemia and SEA-HPFH(P<0.001). CONCLUSION: Through analyze the hematological characteristics, it can be provide that the guidance for the differential diagnosis and genetic consultation of the three commonest deletion ß-thalassemia in Chinese.

[Genetic Effect Analysis of ß-globin Gene 3'UTR+101G>C (HBB:c. *233G>C) Variant].

OBJECTIVE: To investigate whether ß-globin gene 3'UTR+101G>C (HBB:c.*233G>C) variant has genetic effect and provide basis for gene diagnosis and genetic counseling. METHOD: Whole blood cell analysis and capillary zone electrophoresis (CZE) were used to analyze the hematological indexes. The most frequent 23 mutations in southern Chinese individuals were routinely measured by PCR-flow fluorenscence immunmicrobeads assay. Sanger sequencing was used to detect the other variants of ß-globin gene (HBB). RESULTS: In 463 cases, a total of 7 cases with HBB:c.*233G>C variant were detected, among them 4 cases carried other pathogenic variants of HBB gene (2 cases were in trans, 2 cases were in cis), who had typical hematological characteristics of mild ß-thalassemia, and 3 cases also carried abnormal hemoglobin variation, but did not have hematological characteristics of ß-thalassemia. CONCLUSION: The study shows that HBB:c.*233G > C variant has no obvious genetic effect and should be a benign polymorphism.

[Hematological Analysis and Diagnosis of Two Rare Abnormal Hemoglobin].

OBJECTIVE: To analyze the hematological characteristics of Hb Broomhill and Hb Hornchurch, and prenatal diagnosis should be carried out in two families. METHODS: RBC parameters and hemoglobin electrophoretogram were analyzed on the peripheral blood of all patients, and amniotic fluid was collected for prenatal diagnosis. PCR-Flow fluorescent hybridization and Sanger sequencing were performed for gene diagnosis of thalassemia. RESULTS: Three cases of Hb Broomhill were detected, including 2 cases with common SEA α-thalassemia, which was characterized by hypochromic microcytic mild anemia, the capillary electrophoregram revealed a tiny shoulder peak before the Hb A peak; 1 case was diagnosed as Hb Hornchurch combined with ß-thalassemia, which also showed mild anemia. Hemoglobin electrophoretogram showed an abnormal hemoglobin variant peak at Hb A2 zone. CONCLUSION: The carriers of Hb Broomhill and Hb Hornchurch do not have microcytic hypochromic anemia, which do not aggravate the hematological symptoms, such as anemia when being combined with thalassemia of the same type.

Compound heterozygosity for hemoglobin variant Hb-Broomhill and the Southeast Asian α-thalassemia deletion does not worsen outcome: a case report of two unrelated patients.

We report two unrelated cases of compound heterozygosity for hemoglobin (Hb) variant Broomhill and the Southeast Asian (- - SEA/) α-thalassemia deletion, whose clinical features and laboratory findings have never been reported. Hematological analyses revealed abnormal values for both cases as α-thalassemia traits, and capillary electrophoresis suggested an abnormal peak that was incompletely separated from the Hb A peak. A suspension array system and Sanger sequencing were used to characterize the genotypes. Sanger sequencing confirmed the presence of Hb Broomhill [α114(GH2)Pro→Ala; HBA1: c.343C>G]. Eventually, both cases were accurately diagnosed as compound heterozygotes for Hb Broomhill and the (- - SEA/) α-thalassemia deletion, which is the first known report of these variants. This information will be useful when providing appropriate genetic counselling and prenatal diagnosis.

[Gene Diagnosis and Phenotypic Analysis of ß-Thalassemia Caused from a Rare Synonymous Mutation CD29 (C>T)].

OBJECTIVE: To investigate the gene diagnosis and phenotypes analysis for a couple with ß-thalassemia suspected from of blood routine test and hemoglobin electrophoresis, as well as the prenatal gene diagnosis of the fetus. METHODS: The gene mutation of ß-globin in the samples of peripheral blood of pregnant woman and her husband, as well as amniotic fluid of pregnant woman were analyzied and identified by using PCR-RDB and Sanger sequencing. RESULTS: The detection showed that the heterozygote mutation of IVS-Ⅱ-654 (C>T), which is common mutation of ß-globin gene, existed in pregnant woman, while her husband carried a rare mutation CD29 (c.90 C>T) of ß-globin gene. The prenatal diagnosis indicated that the fetus inherited with mutation from the parents, fetus genotype was ßIVS-Ⅱ-6541/ßCD29. CONCLUSION: The CD29(C>T) mutation of ß-globin gene has been identified in Chinese population first. Although this mutation type is symonymous mutation, but its carrier displays phenotype of ß-thalaessmia. Therefore, the attention to this mutation should be paid considering the genetic risk. It contributes to genetic counseling and prenatal gene diagnosis.

Genetic and phenotypic analysis of a rare asymptomatic case of a homozygous Chinese Gγ+(Aγδß)0-thalassemia deletion in a Chinese family.

OBJECTIVE: The clinical and hematologic features of thalassemia are due to different factors, and patients with identical genotypes may regularly exhibit variable severity. In the present work, one homozygous Chinese Gγ+(Aγδß)0-thalassemia case with an asymptomatic phenotype, which is contrary to traditional views, was identified. Analysis of the underlying causes of this rare clinical phenotype involved accurate genetic diagnosis and detection of several genetic modifications. METHODS: Six members of the proband's family were enrolled in the study. Hematological parameters and hemoglobin analysis results were recorded. A suspension-array system, multiplex gap-polymerase chain reaction (gap-PCR) and multiplex ligation-dependent probe amplification (MLPA) were used together to characterize genotypes. Sanger sequencing was utilized to examine the KLF1 gene and four primary fetal hemoglobin (Hb F)-associated single-nucleotide polymorphisms (SNPs). RESULTS: Four family members carried the Chinese Gγ+(Aγδß)0-thalassemia mutation, and a homozygous state was ultimately diagnosed for the proband. All of the Chinese Gγ+(Aγδß)0 mutation-positive cases were coinherited with the Southern Asian α-thalassemia deletion (- - SEA/αα). Two SNP variants, rs7776054 and rs9399137, in the HBS1L-MYB locus were detected in the proband. CONCLUSIONS: Thus far, this is the first study to describe the molecular characterization of a homozygous Chinese Gγ+(Aγδß)0-thalassemia patient who exhibits no clinical symptoms. Our findings suggest that coinheritance of α-thalassemia or HBS1L-MYB locus variants may affect the clinical severity of Chinese Gγ+(Aγδß)0-thalassemia. We conclude that the molecular examination of genetic determinants known to be associated with clinical outcomes in Chinese Gγ+(Aγδß)0-thalassemia should be emphasized.

[Value of Analyzing Hemoglobin A2 by ROC Curve for Screening Thalassemia].

OBJECTIVE: To investigate the value of hemoglobin A2(HbA2) for screening thalassemia. METHODS: A total of 2 000 adults' peripheral blood samples from Guangdong Women and Children Hospital from June 2013 to January 2014 were collected. The hemoglobin A2 (HbA2) level was analyzed by the full automatic capillary electrophoresis technique, and the genotypes of thalassemia were detected. RESULTS: The optimal cutoff values of HbA2 for screening silent α-thalassemia, α-thalassemia trait, intermedia α-thalassemia and ß-thalassemia trait were 2.85%, 2.65%, 2.25% and 3.45%, respectively; the areas under receiver operator characteristic (ROC) curve were 0.709, 0.839, 0.979 and 0.997 respectively; the sensitivities were 0.481, 0.721, 0.953 and 0.994, and the specificities were 0.846, 0.837, 0.929 and 0.969 respectively. CONCLUSION: The optimal cutoff values of HbA2 for screening different type of thalassemia based on our laboratory data are established by using ROC curve. According to the area under ROC curve, a satisfactory accuracy for screening intermedia α-thalassemia and ß-thalassemia trait can be achieved by detecting hemoglobin A2 level.

[Diagnosis and prenatal diagnosis to a family of hemoglobin variant with α-thalassemia].

OBJECTIVE: Diagnosis and prenatal diagnosis to a family of hemoglobin variant with α-thalassemia. METHODS: Whole blood cell analysis, hemoglobin analysis by capillary zone electrophoresis (CZE), Gap-PCR, polymerase chain reaction-reverse dot blot (PCR-RDB) assay and DNA sequencing. RESULTS: Hb Zurich Albisrieden with α°-thalassemia lead to severe anemia. The genotype of fetus is also Hb Zurich Albisrieden with α°-thalassemia. CONCLUSION: Abnormal hemoglobin with α-thalassemia may lead to severe anemia, Prenatal diagnosis of thalassemia has the vital significance for eugenic birth.

The prevalence and molecular spectrum of α- and ß-globin gene mutations in 14,332 families of Guangdong Province, China.

OBJECTIVE: To reveal the familial prevalence and molecular variation of α- and ß-globin gene mutations in Guangdong Province. METHODS: A total of 40,808 blood samples from 14,332 families were obtained and analyzed for both hematological and molecular parameters. RESULTS: A high prevalence of α- and ß-globin gene mutations was found. Overall, 17.70% of pregnant women, 15.94% of their husbands, 16.03% of neonates, and 16.83% of couples (pregnant women and their husbands) were heterozygous carriers of α- or ß-thalassemia. The regions with the highest prevalence were the mountainous and western regions, followed by the Pearl River Delta; the region with the lowest prevalence was Chaoshan. The total familial carrier rate (both spouses were α- or ß-thalassemia carriers) was 1.87%, and the individual carrier rates of α- and ß-thalassemia were 1.68% and 0.20%, respectively. The total rate of moderate-to-severe fetal thalassemia was 12.78% among couples in which both parents were carriers. CONCLUSIONS: There was a high prevalence of α- and ß-thalassemia in Guangdong Province. This study will contribute to the development of thalassemia prevention and control strategies in Guangdong Province.