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Objective The α-globin fusion gene between the HBA2 and HBAP1 genes, is clinically important in thalassemia screening because this fusion gene can cause severe hemoglobin (Hb) H disease when combined with α0 -thalassemia (α0 -thal). In this study, we evaluate the red blood cell parameters of α-thalassemia fusion gene in southern China.Method Study samples suspected of α-thalassemia fusion gene were collected and confirmed by PCR-sequencing from one medical lab center in southern China. Their genotypes and phenotypes were analyzed.Results A total of 266 cases of α-thalassemia fusion gene were confirmed in our lab from 2017 to 2023, most of them were from Hainan province (169 cases) and Huadu district of Guangzhou (21 cases), the nationality of 143 cases from Hainan was identified, with 71.3% (102/143) being from the Li minority. The Hb, MCV, MCH for αα/(αα)fusion in adult males were 143.5±11.83g/L, 81.51±4.39 fl, and 26.26±1.29 pg, respectively; and in females, they were 126.69±12.89 g/L, 80.10±4.05 fl, 25.8±2.04 pg, respectively. All 12 cases (αα) Fusion/ --SEA showed anemia with decreased Hb, MCV and MCH.Conclusion The carriers of α-globin fusion gene heterozygotes are clinically silent and exhibit an α+ phenotype. Individuals with (αα)Fusion/--SEA show apparent anemia. This α-globin fusion gene is relatively common in southern China, specifically among the Li minority of Hainan province. Therefore, it should be taken into account for genetic counseling purposes.

Objectives: This study aimed to establish a droplet digital polymerase chain reaction (ddPCR) assay for South-East Asian (SEA) deletion based on a fully integrated digital PCR system DropXpert S6.Methods: A total of 151 whole blood samples, 10 chorionic villus samples, and 17 amniotic fluid samples were collected, including 106 SEA heterozygotes, 43 normal individuals, 10 Hb Bart's hydrops details, and 19 SEA deletions combined with other genotypes.Genotypes of these samples were determined by the Gap-PCR method. We perform a series of optimizations of the ddPCR system to ensure the performance of the entire ddPCR reaction, such as droplet stability, fluorescence clustering, sensitivity, and accuracy.Results: Our assay exhibited 99.4% (177/178) accuracy compared with the Gap-PCR method, and the minimum detection limit of DNA was 0.1 ng/μL.Both targets have reliable linearity, R2 = 0.9999 for the α-thalassemia SEA deletion allele and R2 = 1 for the wild-type allele. The coefficient of variation for α-thalassemia SEA deletion allele detection at 2 and 10 ng/μL concentrations was 5.42% and 1.91%, respectively. In contrast, the coefficient of variation for wild-type allele detection was 4.06% and 1.83%, demonstrating its high quantitative accuracy. In addition, the DropXpert S6 PCR system showed some advantages over other ddPCR instruments, such as reducing testing costs, simplifying and automating the workflow.Conclusions: The DropXpert S6 PCR system provided a highly accurate diagnosis for α-thalassemia SEA deletion and can be used to detect α-thalassemia as an alternative method.

ABSTRACTBackground Hemoglobin (Hb), a red pigment of red blood cells (RBCs), carries oxygen from the lungs to different organs of the body and transports carbon dioxide back to the lungs. Any fault present in the Hb structure leads to undesirable functional effects of the RBCs, such as sickle cell anemia (SCA), thalassemia, etc. Hemoglobinopathies affect around 7% of people in both developed and developing countries globally. The aim of the present study was to determine the prevalence and carrier frequencies of hemoglobinopathies including SCA, thalassemia, and other abnormal Hb variants among Malayali tribes in the Jawadhu hills of Tiruvannamalai district, Tamil Nadu, India.Methods A community-based cross-sectional study was carried out among 443 Malayali tribes inhabiting the Jawadhu hills of Tiruvannamalai district from July 2022 to September 2022. The RBC indices were analyzed using an automated 5-part hematology analyzer (Mindray, BC-5150) and hemoglobin fractions were done using the HPLC system (Bio-Rad, D-10) following standard protocols.Findings A total of 443 participants were screened, out of whom 14.67% had an abnormal Hb fraction, 83.30% were identified as normal, and 2.03% were borderline. Notably, the study revealed a prevalence of 0.68% for the α-thalassemia trait and 13.99% for the β-thalassemia trait.Interpretation Haemoglobinopathies, specifically the β-thalassemia trait, were most prevalent among the Malayali tribal population of Tamil Nadu residing in the Jawadhu hills of Tiruvannamalai district. Hence, we need special attention for creating awareness, increasing hemoglobinopathies screening programs, and improving the importance of tribal health conditions by the government and non-governmental organizations (NGOs) for the betterment of the ethnic tribes.

Thalassemia, a common autosomal hereditary blood disorder worldwide, mainly contains α- and β-thalassemia. The α-globin gene triplicates allele is harmless for carriers, but aggravates the phenotype of β-thalassemia. Therefore, it is particularly crucial to accurately detect the structural variants of α-globin gene clusters. We reported a 28-year-old man, the proband, with microcytic hypochromic anemia. From pedigree analysis, his mother and sister had hypochromic microcytosis, and his father was normal. Genetic testing of thalassemia identified a novel α-globin gene triplicate named αααanti4.2del726bp (NC_000016.10:g.170769_174300dupinsAAAAAA) by third-generation sequencing (TGS) in the proband and his father, which was further validated by multiplex ligation-dependent probe amplification (MLPA) and Sanger sequencing. The genotypes of the proband's mother and sister were both -α3.7/αα compounded with heterozygous HBB:c.126_129delCTTT. They were categorized as silent α-thalassemia with co-inheritance of β-thalassemia trait. The proband's genotype additionally had the α-globin gene triplicates compared with his mother and sister, which increased the imbalance between α/β-globin, so the proband had more severe hematological parameters. The proband's wife was diagnosed as HBA2:c.427T > C heterozygosis, and his daughter had the novel α-globin gene triplicates compounded with HBA2:c.427T > C, therefore the girl might be asymptomatic. The identification of the novel α-globin gene triplicates provides more insight for the research of thalassemia variants and indicates that TGS has significant advantages on genetic testing of thalassemia for the reliability, accuracy and comprehensiveness. [ABSTRACT FROM AUTHOR]

The 4.2 kb deletion (-α4.2/) is a common a+-thalassemia with a carrier rate, followed by the South-East Asian deletion (–SEA) and the 3.7 kb deletion (-α3.7/). There are few reports about 4.2 kb deletion sub-types. Herein, we present a patient with double heterozygous -α4.2Ⅰ/-α4.2Ⅱwho was identified using third-generation sequencing (TGS). Hematology and hemoglobin fraction analysis were carried out by complete blood count (CBC) and capillary electrophoresis (CE). Gap-PCR was used to detect the common deletional α-thalassemia, and multiple ligation-dependent probe amplification (MLPA) was performed to screen the large deletion. Sanger sequencing identified the variant. The different deletions were confirmed by TGS. CBC showed the patient with microcytic hypochromic anemia, and CE indicated the presence of a Hb variant. Gap-PCR and MLPA detected 4.2 kb deletion homozygotes (-α4.2/-α4.2). The Hb variant was confirmed as Hb Q-Thailand by Sanger sequencing. The patient was identified as compound heterozygous of 4.2 kb deletion and Hb Q-Thailand (-α4.2/-α4.2-Q-Thailand, -α4.2Ⅰ/-α4.2Ⅱ) using TGS. Hb Q-Thailand (-α4.2-Q-Thailand/) complex 4.2 kb deletion heterozygote (-α4.2/) is easily misdiagnosed as 4.2 kb homozygous using Gap-PCR and MLPA. The TGS enables the identification of the two different 4.2 kb deletion sub-types. [ABSTRACT FROM AUTHOR]

In China, conventional genetic testing methods can only detect common thalassemia variants. Accurate detection of rare thalassemia is crucial for clinical diagnosis, especially for children that need long-term blood transfusion. This study aims to explore the application value of third-generation sequencing (TGS) in the diagnosis of rare thalassemia in children with anemia. We enrolled 20 children with anemia, excluding from iron deficiency anemia (IDA). TGS was employed to identify both known and novel thalassemia genotypes, while sanger sequencing was used to confirm the novel mutation detected. Among the 20 samples, we identified 5 cases of rare thalassemia. These included β−4.9 (hg38,Chr11:5226187-5231089) at HBB gene, α−91(HBA2:c.*91delT), αCD30(HBA2:c.91-93delGAG), Chinese Gγ+(Aγδβ)0(NG_000007.3: g.48795-127698 del 78904) and delta – 77(T > C)(HBD:c.-127T>C). Notably, the –SEA/α−91α genotype associated with severe non-deletional hemoglobin H disease (HbH disease) has not been previously reported. Patients with genotypes β654/β−4.9 and –SEA/α−91α necessitate long-term blood transfusions, and those with the –SEA/αCD30α, Chinese Gγ+(Aγδβ)0 and delta thalassemia demonstrate mild anemia. TGS demonstrates promising potential as a diagnostic tool for suspected cases of rare thalassemia in children, especially those suspected to have transfusion-dependent thalassemia (TDT). [ABSTRACT FROM AUTHOR]

To report the hematological and molecular characteristics of a novel α-globin variant found among Chinese families. This study was done on two unrelated families (F1 and F2). Hematological results were obtained by an automated blood cell analyzer. Hemoglobin (Hb) fraction analysis was carried out using capillary electrophoresis (CE) and high-performance liquid chromatography (HPLC). Gap-PCR and reverse dot blot (RDB) methods were used to detect the common α-thalassemia mutations in the Chinese population. The Hb variants were defined by Sanger sequencing. Hb fraction analysis of F2 cord blood using HPLC showed an abnormal peak (3.5%) of the S-window, but CE presented a 12.2% abnormal peak at zone 5(S). Similar results of CE were observed from the F1 twin's cord blood. Compared with newborns, Hb analysis of F2 father using HPLC demonstrated an abnormal peak (16.9%) of S-window and an unknown peak (0.5%) at a retention time of 4.60 min, respectively. In contrast, CE revealed a high Hb F peak at zone 7 and an unknown peak at zone 1. There was no abnormality detected with Gap-PCR and RDB in these patients. However, Sanger sequencing confirmed the presence of a new heterozygous mutation (GAC>GGC) at codon 74 of the HBA2 gene (HBA2:c.224A>G), resulting in a novel Hb variant. We named it Hb Liangqing for the birthplace of the proband. This is the first report of Hb Liangqing detected by HPLC and CE. The normal hematological phenotype suggests that it may be a benign Hb variant. [ABSTRACT FROM AUTHOR]

To investigate the molecular diagnosis of hemoglobin variants in Z region by Capillary electrophoresis in Central Guangxi, Southern China, and analyze their distribution and phenotypic characteristics, to provide a reference for clinical consultation and prenatal diagnosis for couples. A total of 23,709 subjects were collected for blood routine analysis, hemoglobin analysis, and common α- and β-globin gene loci in Chinese population. The hemoglobin electrophoresis components were divided into Zone 1-Zone 15 (Z1-Z15) by Capillary zone electrophoresis (CE). For samples not clearly detected by the conventional technology, Sanger sequencing, and multiplex ligation-dependent probe amplification (MLPA) were used. Single-molecule real-time (SMRT) sequencing technology was used to analyze rare-type genes in a sample with a structural variation. Ten rare hemoglobin variants distributed in Z region were detected in 23,709 samples, including Hb Cibeles, which was reported for the first time in Asia; Hb J-Broussais, Hb G-Honolulu and J-Wenchang-Wuming, they were first reported in Guangxi; 1 case of Hb Anti-Lepore Liuzhou, which was a newly discovered hemoglobin variant; hemoglobin variants Hb G-Siriraj, Hb Handsworth, Hb Q-Thailand, Hb Ube-2, Hb NewYork were also detected. There are a few studies on rare hemoglobin variants in Z region in Southern China. Ten rare hemoglobin variants were found in this study. The hematological phenotype and component content of hemoglobin variants are related to the occurrence of thalassemia. This study enriched the data of rare hemoglobin variants in Southern China and provided a comprehensive data basis for prenatal diagnosis of hemoglobin variants in this area. [ABSTRACT FROM AUTHOR]

ABSTRACTBackground Thalassemia, a common autosomal hereditary blood disorder worldwide, mainly contains α- and β-thalassemia. The α-globin gene triplicates allele is harmless for carriers, but aggravates the phenotype of β-thalassemia. Therefore, it is particularly crucial to accurately detect the structural variants of α-globin gene clusters.Case report We reported a 28-year-old man, the proband, with microcytic hypochromic anemia. From pedigree analysis, his mother and sister had hypochromic microcytosis, and his father was normal. Genetic testing of thalassemia identified a novel α-globin gene triplicate named αααanti4.2del726bp (NC_000016.10:g.170769_174300dupinsAAAAAA) by third-generation sequencing (TGS) in the proband and his father, which was further validated by multiplex ligation-dependent probe amplification (MLPA) and Sanger sequencing. The genotypes of the proband’s mother and sister were both -α3.7/αα compounded with heterozygous HBB:c.126_129delCTTT. They were categorized as silent α-thalassemia with co-inheritance of β-thalassemia trait. The proband’s genotype additionally had the α-globin gene triplicates compared with his mother and sister, which increased the imbalance between α/β-globin, so the proband had more severe hematological parameters. The proband’s wife was diagnosed as HBA2:c.427T > C heterozygosis, and his daughter had the novel α-globin gene triplicates compounded with HBA2:c.427T > C, therefore the girl might be asymptomatic.Conclusion The identification of the novel α-globin gene triplicates provides more insight for the research of thalassemia variants and indicates that TGS has significant advantages on genetic testing of thalassemia for the reliability, accuracy and comprehensiveness.

ABSTRACTObjective The 4.2 kb deletion (-α4.2/) is a common a+-thalassemia with a carrier rate, followed by the South-East Asian deletion (–SEA) and the 3.7 kb deletion (-α3.7/). There are few reports about 4.2 kb deletion sub-types. Herein, we present a patient with double heterozygous -α4.2Ⅰ/-α4.2Ⅱwho was identified using third-generation sequencing (TGS).Methods Hematology and hemoglobin fraction analysis were carried out by complete blood count (CBC) and capillary electrophoresis (CE). Gap-PCR was used to detect the common deletional α-thalassemia, and multiple ligation-dependent probe amplification (MLPA) was performed to screen the large deletion. Sanger sequencing identified the variant. The different deletions were confirmed by TGS.Results CBC showed the patient with microcytic hypochromic anemia, and CE indicated the presence of a Hb variant. Gap-PCR and MLPA detected 4.2 kb deletion homozygotes (-α4.2/-α4.2). The Hb variant was confirmed as Hb Q-Thailand by Sanger sequencing. The patient was identified as compound heterozygous of 4.2 kb deletion and Hb Q-Thailand (-α4.2/-α4.2-Q-Thailand, -α4.2Ⅰ/-α4.2Ⅱ) using TGS.Conclusions Hb Q-Thailand (-α4.2-Q-Thailand/) complex 4.2 kb deletion heterozygote (-α4.2/) is easily misdiagnosed as 4.2 kb homozygous using Gap-PCR and MLPA. The TGS enables the identification of the two different 4.2 kb deletion sub-types.

ABSTRACTObjectives To report the hematological and molecular characteristics of a novel α-globin variant found among Chinese families.Methods This study was done on two unrelated families (F1 and F2). Hematological results were obtained by an automated blood cell analyzer. Hemoglobin (Hb) fraction analysis was carried out using capillary electrophoresis (CE) and high-performance liquid chromatography (HPLC). Gap-PCR and reverse dot blot (RDB) methods were used to detect the common α-thalassemia mutations in the Chinese population. The Hb variants were defined by Sanger sequencing.Results Hb fraction analysis of F2 cord blood using HPLC showed an abnormal peak (3.5%) of the S-window, but CE presented a 12.2% abnormal peak at zone 5(S). Similar results of CE were observed from the F1 twin's cord blood. Compared with newborns, Hb analysis of F2 father using HPLC demonstrated an abnormal peak (16.9%) of S-window and an unknown peak (0.5%) at a retention time of 4.60 min, respectively. In contrast, CE revealed a high Hb F peak at zone 7 and an unknown peak at zone 1. There was no abnormality detected with Gap-PCR and RDB in these patients. However, Sanger sequencing confirmed the presence of a new heterozygous mutation (GAC>GGC) at codon 74 of the HBA2 gene (HBA2:c.224A>G), resulting in a novel Hb variant. We named it Hb Liangqing for the birthplace of the proband.Conclusion This is the first report of Hb Liangqing detected by HPLC and CE. The normal hematological phenotype suggests that it may be a benign Hb variant.

ABSTRACTObjective To investigate the molecular diagnosis of hemoglobin variants in Z region by Capillary electrophoresis in Central Guangxi, Southern China, and analyze their distribution and phenotypic characteristics, to provide a reference for clinical consultation and prenatal diagnosis for couples.Methods A total of 23,709 subjects were collected for blood routine analysis, hemoglobin analysis, and common α- and β-globin gene loci in Chinese population. The hemoglobin electrophoresis components were divided into Zone 1-Zone 15 (Z1-Z15) by Capillary zone electrophoresis (CE). For samples not clearly detected by the conventional technology, Sanger sequencing, and multiplex ligation-dependent probe amplification (MLPA) were used. Single-molecule real-time (SMRT) sequencing technology was used to analyze rare-type genes in a sample with a structural variation.Results Ten rare hemoglobin variants distributed in Z region were detected in 23,709 samples, including Hb Cibeles, which was reported for the first time in Asia; Hb J-Broussais, Hb G-Honolulu and J-Wenchang-Wuming, they were first reported in Guangxi; 1 case of Hb Anti-Lepore Liuzhou, which was a newly discovered hemoglobin variant; hemoglobin variants Hb G-Siriraj, Hb Handsworth, Hb Q-Thailand, Hb Ube-2, Hb NewYork were also detected.Conclusion There are a few studies on rare hemoglobin variants in Z region in Southern China. Ten rare hemoglobin variants were found in this study. The hematological phenotype and component content of hemoglobin variants are related to the occurrence of thalassemia. This study enriched the data of rare hemoglobin variants in Southern China and provided a comprehensive data basis for prenatal diagnosis of hemoglobin variants in this area.

ABSTRACTObjectives To explore the application of third-generation sequencing (TGS) for genetic diagnosis and prenatal genetic screening of thalassemia genes.Methods Two groups of subjects were enrolled in this study. The first group included 176 subjects with positive hematological phenotypes for thalassemia. Thalassemia-associated genes were detected simultaneously in each sample using both the PacBio TGS platform based on single-molecule real-time (SMRT) technology and the conventional PCR-reverse dot blot (PCR-RDB). Sanger sequencing was used for validation when results were discordant between the two methods. The second group included 53 couples with at least one partner having a positive thalassemia hematological phenotype, and they were screened for homotypic thalassemia variants by TGS, and the risk of pregnancies with babies presenting with severe thalassemia, was assessed.Results Of the 176 subjects, 175 had concordant genotypes between the two methods, including 63 normal subjects and 112 α- and/or β-thalassemia gene carriers, with a concordance rate of 99.43%. TGS detected a rare β-thalassemia gene variant −50 (G > A) that was not detected by conventional PCR-RDB. TGS identified seven of the 53 couples as homotypic thalassemia gene carriers, five of whom were at risk of pregnancies with severe thalassemia.Conclusion TGS could effectively detect common and rare thalassemia variants with high accuracy and efficiency. This approach would be suitable for prenatal thalassemia genetic screening in areas with high incidence of thalassemia.

Objectives: Thalassemia, the most common global monogenetic disorder, is highly prevalent in southern China. Epidemiological and molecular characterization of thalassemia is important for designing appropriate prevention strategies in high-risk areas, especially the border area of Guangxi-Yunnan-Guizhou province in southwestern China.Methods: We recruited 38812 reproductive age couples and screened them for thalassemia. Routine blood tests as well as hemoglobin components and levels were evaluated. In addition, suspected thalassemia were identified by gap polymerase chain reaction (Gap-PCR) and PCR-based reverse dot blot (PCR-RDB).Results: The overall prevalence of thalassemia was 26.76%. Specifically, incidences of α-thalassemia, β-thalassemia, and concurrent α- and β-thalassemia were 17.52%, 6.92%, and 2.32%, respectively. The diagnosed α-thalassemia anomalies were associated with six gene mutations and 25 genotypes. The β-thalassemia anomalies were associated with 12 gene mutations and 15 genotypes. Moreover, among the 1799 concurrent mutated α- and β-thalassemia genes, 95 different genotypes were identified. Couples in which both partners were positive for α-thalassemia and β-thalassemia isotypes were 8.80% and 2.08%, respectively. The proportion of couples at a risk of having children with thalassemia major or intermedia was high.Conclusions: This study elucidates on the prevalence and molecular characterization of thalassemia in the border area of Guangxi-Yunnan-Guizhou provinces. These findings provide valuable baseline data for genetic counseling and prenatal diagnosis, with the overarching goal of preventing and controlling severe thalassemia.

Objectives To report the hematolgocial and molecular features of a nove β-globin variant in a Chinese fimaly.Methods The proband was a 19-year-old Chinese man whose Hb analysis by HPLC for thalassemia revealed an abnormal peak. Hb analysis was performed by HPLC and CE. Gap-PCR and PCR-reverse dot blot hybridization (PCR-RDB) were used to detect the common mutations in Chinese population. DNA sequencing was used to determine the Hb variant.Results The Hb variant and Hb A can be separated but co-elutes with Hb F by the CE method. However, the variant can be separated from Hb A0, Hb F, and Hb A2 using HPLC. DNA sequencing showed a mutation of codon 3 in the β-globin gene. His wife’s HPLC revealed a high value of Hb A2, which proved to be the Hb E using PCR-RDB.Conclusion It was the first report of the mutation, so we named it Hb Jiangnan according to the place of residence of the proband. It can be separated by HPLC but not CE. Hb Jiangnan can cause an increased level of Hb A2.

Objectives Shaokwan is one of the inhabitant regions of Hakka population in Guangdong province of southern China. Previous survey has reported that a higher prevalence of abnormal hemoglobin (Hb) in this region. However, large-scale survey on the molecular characteristics of abnormal hemoglobin in this south-north junctional region has not been reported.In this study, we aim to investigate the molecular characteristics of abnormal hemoglobin in this area. Methods Blood samples from medical check-up center in one local hospital were selected for abnormal hemoglobin screening. Hemoglobin electrophoresis and routine blood tests were performed. Hemoglobin variants were further analyzed by PCR and DNA sequencing. Results Among the 9731 study subjects, 45 cases of hemoglobin variants were found by hemoglobin electrophoresis, which gave an incidence of 0.46% (45/9731) in Shaokwan region. 8 kinds of hemoglobin variants were identified by gene sequencing. Hb Q-Thailand (16/45) was the most common hemoglobin variants, followed by HbE (7/45), Hb NewYork (6/45) and Hb G-Chinese (6/45). Discussion and Conclusion Hemoglobin variants had obviously genetic polymorphisms in Chinese. Our study of hemoglobin disorders in this special Hakka Chinese population will contribute considerably to our understanding of the historical, emigrational, and genetic relationships among different ethnic group in this region.

Objective Thalassemia is a monogenic genetic disorder with a high prevalence in populations in the southern region of China. The thalassemia gene prevalence rate in the Baise population in China is high, and several rare gene variants have been detected in the population of this region during routine testing by our study group. To accurately reveal the thalassemia gene variants carried by the population in Baise, and to provide a basis for the formulation of thalassemia prevention and control policies in the region, we conducted a more comprehensive study in a randomly selected population.Results In all, 4,800 randomized individuals were recruited for testing from Baise, and the detection of hot spot thalassemia genetic variants were performed by Gap-PCR and PCR-RDB methods, combined with the relative quantification of homologous fragments and AS-PCR to expand the detection range. The prevalence of thalassemia variants in this population was 24.19%, among which 16.69% of individuals carried α-thalassemia gene variants alone, 5.62% carried β-thalassemia gene variants alone, and 1.88% carried both variants.Conclusions The use of positive primary screening combined with hot spot gene variant detection alone can result in a certain degree of missed detection. In the prevention and control of thalassemia in the region, testing institutions need to pay attention to the detection of rare thalassemia gene variants such as αααanti4.2, αααanti3.7, –α2.4, –α21.9, β−50, β−90, and βIVS-II−5, to provide more accurate genetic counseling advice to subjects.

To explore the application of third-generation sequencing (TGS) for genetic diagnosis and prenatal genetic screening of thalassemia genes. Two groups of subjects were enrolled in this study. The first group included 176 subjects with positive hematological phenotypes for thalassemia. Thalassemia-associated genes were detected simultaneously in each sample using both the PacBio TGS platform based on single-molecule real-time (SMRT) technology and the conventional PCR-reverse dot blot (PCR-RDB). Sanger sequencing was used for validation when results were discordant between the two methods. The second group included 53 couples with at least one partner having a positive thalassemia hematological phenotype, and they were screened for homotypic thalassemia variants by TGS, and the risk of pregnancies with babies presenting with severe thalassemia, was assessed. Of the 176 subjects, 175 had concordant genotypes between the two methods, including 63 normal subjects and 112 α- and/or β-thalassemia gene carriers, with a concordance rate of 99.43%. TGS detected a rare β-thalassemia gene variant −50 (G > A) that was not detected by conventional PCR-RDB. TGS identified seven of the 53 couples as homotypic thalassemia gene carriers, five of whom were at risk of pregnancies with severe thalassemia. TGS could effectively detect common and rare thalassemia variants with high accuracy and efficiency. This approach would be suitable for prenatal thalassemia genetic screening in areas with high incidence of thalassemia. [ABSTRACT FROM AUTHOR]

Thalassemia is a monogenic genetic disorder with a high prevalence in populations in the southern region of China. The thalassemia gene prevalence rate in the Baise population in China is high, and several rare gene variants have been detected in the population of this region during routine testing by our study group. To accurately reveal the thalassemia gene variants carried by the population in Baise, and to provide a basis for the formulation of thalassemia prevention and control policies in the region, we conducted a more comprehensive study in a randomly selected population. In all, 4,800 randomized individuals were recruited for testing from Baise, and the detection of hot spot thalassemia genetic variants were performed by Gap-PCR and PCR-RDB methods, combined with the relative quantification of homologous fragments and AS-PCR to expand the detection range. The prevalence of thalassemia variants in this population was 24.19%, among which 16.69% of individuals carried α-thalassemia gene variants alone, 5.62% carried β-thalassemia gene variants alone, and 1.88% carried both variants. The use of positive primary screening combined with hot spot gene variant detection alone can result in a certain degree of missed detection. In the prevention and control of thalassemia in the region, testing institutions need to pay attention to the detection of rare thalassemia gene variants such as αααanti4.2, αααanti3.7, –α2.4, –α21.9, β−50, β−90, and βIVS-II−5, to provide more accurate genetic counseling advice to subjects. [ABSTRACT FROM AUTHOR]

Objectives: Thalassemia, the most common global monogenetic disorder, is highly prevalent in southern China. Epidemiological and molecular characterization of thalassemia is important for designing appropriate prevention strategies in high-risk areas, especially the border area of Guangxi-Yunnan-Guizhou province in southwestern China. Methods: We recruited 38812 reproductive age couples and screened them for thalassemia. Routine blood tests as well as hemoglobin components and levels were evaluated. In addition, suspected thalassemia were identified by gap polymerase chain reaction (Gap-PCR) and PCR-based reverse dot blot (PCR-RDB). Results: The overall prevalence of thalassemia was 26.76%. Specifically, incidences of α-thalassemia, β-thalassemia, and concurrent α- and β-thalassemia were 17.52%, 6.92%, and 2.32%, respectively. The diagnosed α-thalassemia anomalies were associated with six gene mutations and 25 genotypes. The β-thalassemia anomalies were associated with 12 gene mutations and 15 genotypes. Moreover, among the 1799 concurrent mutated α- and β-thalassemia genes, 95 different genotypes were identified. Couples in which both partners were positive for α-thalassemia and β-thalassemia isotypes were 8.80% and 2.08%, respectively. The proportion of couples at a risk of having children with thalassemia major or intermedia was high. Conclusions: This study elucidates on the prevalence and molecular characterization of thalassemia in the border area of Guangxi-Yunnan-Guizhou provinces. These findings provide valuable baseline data for genetic counseling and prenatal diagnosis, with the overarching goal of preventing and controlling severe thalassemia. [ABSTRACT FROM AUTHOR]

To report the hematolgocial and molecular features of a nove β-globin variant in a Chinese fimaly. The proband was a 19-year-old Chinese man whose Hb analysis by HPLC for thalassemia revealed an abnormal peak. Hb analysis was performed by HPLC and CE. Gap-PCR and PCR-reverse dot blot hybridization (PCR-RDB) were used to detect the common mutations in Chinese population. DNA sequencing was used to determine the Hb variant. The Hb variant and Hb A can be separated but co-elutes with Hb F by the CE method. However, the variant can be separated from Hb A0, Hb F, and Hb A2 using HPLC. DNA sequencing showed a mutation of codon 3 in the β-globin gene. His wife's HPLC revealed a high value of Hb A2, which proved to be the Hb E using PCR-RDB. It was the first report of the mutation, so we named it Hb Jiangnan according to the place of residence of the proband. It can be separated by HPLC but not CE. Hb Jiangnan can cause an increased level of Hb A2. [ABSTRACT FROM AUTHOR]

Shaokwan is one of the inhabitant regions of Hakka population in Guangdong province of southern China. Previous survey has reported that a higher prevalence of abnormal hemoglobin (Hb) in this region. However, large-scale survey on the molecular characteristics of abnormal hemoglobin in this south-north junctional region has not been reported.In this study, we aim to investigate the molecular characteristics of abnormal hemoglobin in this area. Blood samples from medical check-up center in one local hospital were selected for abnormal hemoglobin screening. Hemoglobin electrophoresis and routine blood tests were performed. Hemoglobin variants were further analyzed by PCR and DNA sequencing. Among the 9731 study subjects, 45 cases of hemoglobin variants were found by hemoglobin electrophoresis, which gave an incidence of 0.46% (45/9731) in Shaokwan region. 8 kinds of hemoglobin variants were identified by gene sequencing. Hb Q-Thailand (16/45) was the most common hemoglobin variants, followed by HbE (7/45), Hb NewYork (6/45) and Hb G-Chinese (6/45). Hemoglobin variants had obviously genetic polymorphisms in Chinese. Our study of hemoglobin disorders in this special Hakka Chinese population will contribute considerably to our understanding of the historical, emigrational, and genetic relationships among different ethnic group in this region. [ABSTRACT FROM AUTHOR]

To evaluate any association between pre-transfusion hemoglobin (Hb) levels and thalassemia complications and to identify the Hb cutoff values to predict thalassemia complications. We performed a cross-sectional study in thalassemia patients who attended the Adult Hematology Clinic of the tertiary care University Hospital from October 2017 to October 2018. A point-biserial correlation was used to identify any association between Hb levels and thalassemia complications. A receiver operating characteristic (ROC) curve was used to identify the diagnostic ability of Hb levels to predict thalassemia complications and identify Hb cutoff values. Out of the 102 patients, there were 53 transfusion dependent thalassemia (TDT) patients and 49 non-transfusion dependent thalassemia (NTDT) patients. In theTDT group, Hb levels showed a negative correlation with severe hepatic iron overload and hypogonadism. The cutoff Hb levels to predict severe hepatic iron overload and hypogonadism were ≤7.01 and 6.81 g/dL, respectively, at which points the area under the ROC curve (AUC) were 0.721 and 0.708, respectively. In the NTDTgroup, Hb levels were negatively correlated with hepatic iron overload, osteoporosis, and pulmonary hypertension. The cutoff values of Hb levels to predict hepatic iron overload, osteoporosis, and pulmonary hypertension were ≤8.24, 7.16, and 7.16 g/dL, respectively, at which points the AUC were 0.923, 0.715, and 0.725, respectively. Lower Hb level was associated with more frequent complications in both TDT and NTDT patients. The Hb cutoff levels to predict these complications were identified. [ABSTRACT FROM AUTHOR]

β-Thalassemia (β-thal) is a genetic disease of the blood caused by mutations in the β-globin gene. Conventional methods for detecting thalassemia variants often miss rare and novel variants. Identifying the rare and novel β-thal variants, especially in the high prevalence regions, would enable better disease prevention. A Chinese family who had joined the Thalassemia Prevention Program was recruited in this study. The β-thal carrier screening was performed using next-generation sequencing (NGS), and the results were validated through direct DNA sequencing. Hematological parameters were analyzed, and hemoglobin electrophoresis was performed. Additionally, the presence of thalassemia-associated deletions was determined using gap-polymerase chain reaction. A novel frameshift variant of β-thal, HBB:c.14delC(Codon 4, -C), was identified in a 31-year-old Chinese man. Subsequent genetic investigation showed that his mother also carried this novel variant. Hematological analysis and clinical evaluation suggested that this variant was present in the heterozygous state and might belong to a severe phenotype of β-thal. We identified a novel frameshift variant of β-thal. NGS has the potential for identifying rare and novel thalassemia variants and broadening the spectrum of thalassemia screening and thus may contribute to effective prevention of thalassemia. [ABSTRACT FROM AUTHOR]

Objectives: To investigate the effect of HS-40 regulatory site deletion on α-globin gene expression and its clinical significance. Methods: Venous blood samples of subjects were analyzed using a hematology analyzer and high- performance liquid chromatography; fetal cord blood was analyzed by a capillary electrophoresis analyzer. Gap-polymerase chain reaction (PCR), reverse dot blot (RDB), and multiple-link-dependent probe amplification (MLPA) were used for genotyping of thalassemia. Results: The proband was POLR3 K, HS-40 heterozygous deletion; the proband's wife was –SEA/αα; the fetus was POLR3 K, HS-40 heterozygous deletion combined with –SEA deletion; all of them had microcytic hypochromic anemia. Fetal umbilical cord blood electrophoresis revealed a suspected Hb Bart's band to be 88.4%, and the fetus was, thus, diagnosed as Hb Bart's fetus. The red blood cell parameters of the sporadic case showed that he had microcytic hypochromic anemia. Hemoglobin (Hb) electrophoresis analysis showed Hb H to be 5.3%, leading to a diagnosis of Hb H disease. Gap-PCR and RDB identified the genotype to be –α3.7/αα, βA/βA. MLPA detected heterozygous deletion or –α3.7 deletion on one allele and deletion of the HS-40 regulatory site on the other allele. Conclusion: The deletion of HS-40 regulatory site reduced expression of α-globin. HS-40 heterozygous deletion manifested as mild anemia, which was of microcytic hypochromic type. When compounded with –α3.7/αα, it manifested as Hb H disease; and when compounded with –SEA/αɑ, it manifested as Hb Bart's fetus. [ABSTRACT FROM AUTHOR]

Objectives: Beta-thalassemias are a group of recessively autosomal inherited disorders of hemoglobin synthesis, which, due to mutations of the beta-globin gene, lead to various degrees of defective beta-chain production, an imbalance in alpha/beta-globin chain synthesis, ineffective erythropoiesis, and anemia. Improved survival in thalassemic patients has led to the emergence of previously unrecognized complications, such as renal disease. Methods: A comprehensive literature review through PubMed was undertaken to summarize the published evidence on the epidemiology and pathophysiology of renal disease in thalassemia. Literature sources published in English since 1990 were searched, using the terms beta-thalassemia, renal disease. Results: Renal disease is considered to be the 4th cause of morbidity among patients with transfusion dependent thalassemia. Chronic anemia, hypoxia and iron overload are the main mechanisms implicated in development of renal injury, whereas several studies also suggested a contributive role of iron chelators. Discussion and Conclusion: Kidney disease may develop through progressive renal tubular and glomerular damage; thus, its early recognition is important in order to prevent and/or reverse deterioration. This review will provide an insight on the involved mechanisms implicated in kidney disease in thalassemic patients and will discuss the updates on diagnosis and prevention of renal complications in thalassemia. [ABSTRACT FROM AUTHOR]

Introduction: Noggin is an antagonist of bone morphogenetic proteins (BMPs) and has a strong effect on osteogenesis. Osteoporosis is a common complication of transfusion dependent beta-thalassemia (TDT) and denosumab has been recently emerged as a promising therapeutic option. This was a post hoc investigation of serum noggin levels among TDT patients with osteoporosis who participated in a randomized, placebo-control, phase 2b study. Methods: Patients received either 60 mg denosumab (n = 32) or placebo (n = 31) every 6 months for 12 months. Noggin was measured, for the first time in thalassemia patients, at baseline and at 12 months, using a recently developed high sensitivity fluorescent immunoassay. Results: Both groups showed a significant increase in noggin serum levels (denosumab p < 0.001; placebo p < 0.0001). Interestingly, the increase was higher in the placebo group. Furthermore, we observed a strong correlation between noggin and wrist bone mineral density (r = −0.641, p = 0.002) only in the denosumab group. Conclusion: In conclusion, higher noggin levels reflected more BMP inhibition, since our assay detects free bioactive noggin, which in turn impaired bone formation in placebo group. Therefore, denosumab possibly regulates noggin and favours bone turnover in TDT patients with osteoporosis through a novel mechanism of action. [ABSTRACT FROM AUTHOR]

Objective: Iron overload and oxidative stress are the major causes of serious complications and mortality in thalassemic patients. Our previous work supports the synergistic effects of antioxidant cocktails (curcuminoids or vitamin E, N-acetylcysteine, and deferiprone) in treatment of β-thalassemia/Hb E patients. This further 2-DE-based proteomic study aimed to identify the plasma proteins that expressed differentially in response to antioxidant cocktails. Methods: Frozen plasma samples of ten normal subjects and ten β-thalassemia/Hb E patients at three-time points (baseline, month 6, and month 12) were reduced the dynamic range of proteome using ProteoMiner kit and separated proteins by two-dimensional gel electrophoresis. Differentially expressed proteins were identified using tandem mass spectrometry. Several plasma proteins were validated by ELISA and Western blot analysis. Results: Thirteen and 11 proteins were identified with altered expression levels in the curcuminoids- and vitamin E cocktail groups, respectively. The associations between vitronectin (VTN) expression and total bilirubin levels, as well as between serum paraoxonase/arylesterase 1 (PON1) expression and blood reactive oxygen species were observed. Validation results were consistent with proteomics results. Discussion and conclusions: These plasma proteins may provide better understanding of the mechanisms underlying the therapeutic effects of antioxidant cocktails in thalassemic patients. [ABSTRACT FROM AUTHOR]

The study was to determine the prevalence and clinical significances of red blood cell (RBC)-bound IgG as detected by flow cytometry in polytransfused patients with thalassemias. Relationship of the presence of RBC-bound IgG with RBC alloimmunization was also evaluated. This study included 59 polytransfused patients with β-thalassemia disease. We studied the frequency of RBC autoantibodies and alloimmunization. Direct Coombs test and flow cytometry were performed to detect the presence of RBC autoantibodies while RBC alloantibodies were detected by antibody screening and identification assays. Eight (13.6%) and 34 (57.6%) patients were found a positive direct Coombs test and flow cytometry, respectively. Twenty (33.9%) patients developed RBC alloantibodies. The four most frequent RBC alloantibodies were anti-E (55%), anti-Mia (40%), anti-Di(a) (25%) and anti-c (15%), respectively. There was no significant difference in the presence of RBC-bound IgG between polytransfused with thalassemia patients who developed RBC alloimmunization (13 of 20; 65%) and those without RBC alloantibodies (21 of 39; 53.8%), p = 0.412. Splenectomy and increased transfusion requirement were significantly associated with the presence of RBC-bound IgG but not with RBC alloantibody formation. The overall frequency of RBC alloantibody formation in polytransfused patients with thalassemias was 33.9%. The most common RBC alloantibody was anti-E. RBC autoantibody formation was more frequently detected by flow cytometry (57.6%) than by direct Coombs test (13.6%). Splenectomy was significantly associated with the development of autoreactive RBC-bound IgG antibodies in the polytransfused patients with thalassemias. The presence of the anti-RBC autoantibodies may cause an increase of transfusion requirement. [ABSTRACT FROM AUTHOR]

Objective: Iron overload and oxidative stress are the major causes of serious complications and mortality in thalassemic patients. Our previous work supports the synergistic effects of antioxidant cocktails (curcuminoids or vitamin E, N-acetylcysteine, and deferiprone) in treatment of β-thalassemia/Hb E patients. This further 2-DE-based proteomic study aimed to identify the plasma proteins that expressed differentially in response to antioxidant cocktails. Methods: Frozen plasma samples of ten normal subjects and ten β-thalassemia/Hb E patients at three-time points (baseline, month 6, and month 12) were reduced the dynamic range of proteome using ProteoMiner kit and separated proteins by two-dimensional gel electrophoresis. Differentially expressed proteins were identified using tandem mass spectrometry. Several plasma proteins were validated by ELISA and Western blot analysis. Results: Thirteen and 11 proteins were identified with altered expression levels in the curcuminoids- and vitamin E cocktail groups, respectively. The associations between vitronectin (VTN) expression and total bilirubin levels, as well as between serum paraoxonase/arylesterase 1 (PON1) expression and blood reactive oxygen species were observed. Validation results were consistent with proteomics results. Discussion and conclusions: These plasma proteins may provide better understanding of the mechanisms underlying the therapeutic effects of antioxidant cocktails in thalassemic patients. [ABSTRACT FROM AUTHOR]

The study was to determine the prevalence and clinical significances of red blood cell (RBC)- bound IgG as detected by flow cytometry in polytransfused patients with thalassemias. Relationship of the presence of RBC-bound IgG with RBC alloimmunization was also evaluated. This study included 59 polytransfused patients with β-thalassemia disease. We studied the frequency of RBC autoantibodies and alloimmunization. Direct Coombs test and flow cytometry were performed to detect the presence of RBC autoantibodies while RBC alloantibodies were detected by antibody screening and identification assays. Eight (13.6%) and 34 (57.6%) patients were found a positive direct Coombs test and flow cytometry, respectively. Twenty (33.9%) patients developed RBC alloantibodies. The four most frequent RBC alloantibodies were anti-E (55%), anti-Mia (40%), anti-Di(a) (25%) and antic (15%), respectively. There was no significant difference in the presence of RBC-bound IgG between polytransfused with thalassemia patients who developed RBC alloimmunization (13 of 20; 65%) and those without RBC alloantibodies (21 of 39; 53.8%), p = 0.412. Splenectomy and increased transfusion requirement were significantly associated with the presence of RBC-bound IgG but not with RBC alloantibody formation. The overall frequency of RBC alloantibody formation in polytransfused patients with thalassemias was 33.9%. The most common RBC alloantibody was anti-E. RBC autoantibody formation was more frequently detected by flow cytometry (57.6%) than by direct Coombs test (13.6%). Splenectomy was significantly associated with the development of autoreactive RBC-bound IgG antibodies in the polytransfused patients with thalassemias. The presence of the anti-RBC autoantibodies may cause an increase of transfusion requirement. [ABSTRACT FROM AUTHOR]

Objectives: A multicenter, noninterventional, observational study was conducted in the Latin American countries including Argentina, Brazil, Colombia, Mexico, and Venezuela to assess the prevalence of liver and cardiac iron overload using magnetic resonance imaging (MRI) in patients with chronic anemias except thalassemia. Methods: Patients aged >10 years with transfusion-dependent anemias, except thalassemia, either with <20 units of red blood cell (RBC) transfusions with serum ferritin (SF) levels >2000 ng/mL or with ≥20 units of RBC transfusions regardless of SF level in their lifetime, were enrolled. Iron overload was assessed using MRI. Results: Among 175 patients included, the majority had sickle cell disease (SCD; 52%), followed by aplastic anemia (AA; 17.7%), myelodysplastic syndrome (MDS; 8.6%), Diamond-Blackfan anemia (DBA; 4%), pure red cell aplasia (1.1%), and others (16.6%). Liver iron overload was observed in 76.4% of patients, while cardiac iron overload was seen in 19.2% when assessed by MRI. The prevalence of iron overload was 80.2% in patients with SCD, 73.3% in MDS, 77.4% in AA, 100% in pure red cell aplasia, 71.4% in DBA, and 68.9% in other transfusion-related disorders. A moderate correlation between liver iron concentration (LIC) and SF was observed in patients with SCD and MDS (r = 0.47 and r = 0.61, respectively). All adverse events reported were consistent with the published data for deferasirox or underlying disease. Conclusion: A high prevalence of iron overload in this patient population in Latin American countries indicates that a better diagnosis and management of iron overload is required in these countries. [ABSTRACT FROM AUTHOR]

Objectives: β-Thalassemia disease is caused by mutations in the β-globin gene. This is considered as one of the common genetic disorders in Syria. The aim of this study was to identify the geographical distribution of the β-thalassemia mutations in Syria. Methods: β-Globin gene mutations were characterized in 636 affected patients and 94 unrelated carriers using the amplification refractory mutations system-polymerase chain reaction technique and DNA sequencing. Results: The study has revealed the presence of 38 β-globin gene mutations responsible for β-thalassemia in Syria. Important differences in regional distribution were observed. IVS-I.110 [G > A] (22.2%), IVS-I.1 [G > A] (17.8%), Cd 39 [C > T] (8.2%), IVS-II.1 [G > A] (7.6%), IVS-I.6 [T > C] (7.1%), Cd 8 [−AA] (6%), Cd 5 [−CT] (5.6%) and IVS-I.5 [G > C] (4.1%) were the eight predominant mutations found in our study. The coastal region had higher relative frequencies (37.9 and 22%) than other regions. A clear drift in the distribution of the third common Cd 39 [C > T] mutation in the northeast region (34.8%) to the northwest region (2.5%) was noted, while the IVS-I.5 [G > C] mutation has the highest prevalence in north regions. The IVS-I.6 [T > C] mutation had a distinct frequency in the middle region. Ten mutations −86 [C > G], −31 [A > G], −29 [A > G], 5′UTR; +22 [G > A], CAP + 1 [A > C], Codon 5/6 [−TG], IVS-I (−3) or codon 29 [C > T], IVS-I.2 [T > A], IVS-I.128 [T > G] and IVS-II.705 [T > G] were found in Syria for the first time. Conclusions: These data will significantly facilitate the population screening, genetic counseling and prenatal diagnosis in Syrian population. [ABSTRACT FROM AUTHOR]

Background and objectives: Coinheritance of δβ thalassemia and HPFH with inherited factors is sparsely documented and may affect treatment modalities. So, we screened the presence of α deletion and β mutations in δβ thalassemia and HPFH disorders in 52 cases with high Hb F concentration. Material and methods: Fifty-two individuals with raised HbF levels were study subjects. CZE was done for quantitative assessment of hemoglobin variants. Asian Indian inversion deletion break point type A, B and HPFH-3 were done by GAP-PCR. Results: 18/52 cases of δβ Gγ (Aγδβ)0 thalassemia and 28/52 cases of HPFH-3 deletion were characterized. 6/52 patients with raised HbF levels were negative for δβ Gγ (Aγδβ)0 and HPFH-3 deletion. 9/18 (50%) were heterozygous for Gγ(Aγδβ)0 break point type A, 6/18 (33%) were heterozygous for break point type B and 3/18 (17%) were homozygous. Of the nine patients heterozygous for Gγ(Aγδβ)0 break point type A, three (33%) patients were double heterozygous with alpha 3.7 kb deletion and two (22%) patients showed compound heterozygosity with IVS 1-5(G-C) mutation. 4/9 (45%) patients were Gγ(Aγδβ)0 heterozygous. Discussion and conclusion: We found 5/18(27.β) δβ-thalassemia cases with co-inherited alpha 3.7 deletion and 3/18 (16β) cases with IVS 1-5(G-C) mutation. Patients showed features of thalassemia intermedia phenotype among which those with co-inherited IVS 1-5(G-C) mutation showed severe phenotype as compared to those with co-inherited alpha 3.7 deletion. So, we highlight importance of genotyping of patients with δβ thalassemia or HPFH and coinheritance with inherited factors which plays crucial role in clinicopathological profile and setting up prenatal diagnostic protocol. [ABSTRACT FROM AUTHOR]

Context and Objective: Microcytic anaemia results from defective synthesis of haemoglobin in the erythroid precursors, causing a reduction in its mean corpuscular volume (MCV). The most common causes of microcytosis, without the increase in HbA2 levels, are iron deficiency anaemia (IDA) and α-thalassemia. The aim of this study was to identify the causes of microcytic anaemia and evaluate the haematological parameters from blood donors deemed ineligible (due to the low haematocrit level) that would differentiate the IDA and α-thal, whether isolated or in association. Methods: Genomic DNA was submitted to the polymerase chain reaction multiplex for the diagnosis of the most common allele deletions of α-thal and erythrogram and in order to verify haematological parameters. Iron deficiency (ID) was determined through the measurement of serum ferritin. Results: Of the 204 samples, 82 (40.2%) were identified with ID, 24 (17.8%) with α-thal and 10 (4.9%) with ID associated with α-thal. In the α-thal with ID group haemoglobin (Hb), MCV, mean corpuscular Hb concentration (MCHC) and mean corpuscular Hb (MCH) values were significantly lower compared to the isolated α-thal. In the group with ID Hb, MCV, MCHC and MCH values were significantly lower compared to those with isolated α-thal. The α-thal with ID group, showed Hb, MCV, MCHC and MCH significantly reduced when compared to those with IDA. Conclusions: This study showed that the values of haematological parameters, especially haematocrit, Hb, MCV, MCH, MCHC and red blood cell distribution width (RDW), are lower in patients with IDA, especially when associated with α-thal and therefore it may be useful to discriminate between the different types of microcytic anaemia. [ABSTRACT FROM AUTHOR]

Objectives: A multicenter, noninterventional, observational study was conducted in the Latin American countries including Argentina, Brazil, Colombia, Mexico, and Venezuela to assess the prevalence of liver and cardiac iron overload using magnetic resonance imaging (MRI) in patients with chronic anemias except thalassemia. Methods: Patients aged >10 years with transfusion-dependent anemias, except thalassemia, either with <20 units of red blood cell (RBC) transfusions with serum ferritin (SF) levels >2000 ng/mL or with ≥20 units of RBC transfusions regardless of SF level in their lifetime, were enrolled. Iron overload was assessed using MRI. Results: Among 175 patients included, the majority had sickle cell disease (SCD; 52%), followed by aplastic anemia (AA; 17.7%), myelodysplastic syndrome (MDS; 8.6%), Diamond-Blackfan anemia (DBA; 4%), pure red cell aplasia (1.1%), and others (16.6%). Liver iron overload was observed in 76.4% of patients, while cardiac iron overload was seen in 19.2% when assessed by MRI. The prevalence of iron overload was 80.2% in patients with SCD, 73.3% in MDS, 77.4% in AA, 100% in pure red cell aplasia, 71.4% in DBA, and 68.9% in other transfusion-related disorders. A moderate correlation between liver iron concentration (LIC) and SF was observed in patients with SCD and MDS (r = 0.47 and r = 0.61, respectively). All adverse events reported were consistent with the published data for deferasirox or underlying disease. Conclusion: A high prevalence of iron overload in this patient population in Latin American countries indicates that a better diagnosis and management of iron overload is required in these countries. [ABSTRACT FROM AUTHOR]

Context and Objective: Microcytic anaemia results from defective synthesis of haemoglobin in the erythroid precursors, causing a reduction in its mean corpuscular volume (MCV). The most common causes of microcytosis, without the increase in HbA2 levels, are iron deficiency anaemia (IDA) and α-thalassemia. The aim of this study was to identify the causes of microcytic anaemia and evaluate the haematological parameters from blood donors deemed ineligible (due to the low haematocrit level) that would differentiate the IDA and α-thal, whether isolated or in association. Methods: Genomic DNA was submitted to the polymerase chain reaction multiplex for the diagnosis of the most common allele deletions of α-thal and erythrogram and in order to verify haematological parameters. Iron deficiency (ID) was determined through the measurement of serum ferritin. Results: Of the 204 samples, 82 (40.2%) were identified with ID, 24 (17.8%) with α-thal and 10 (4.9%) with ID associated with α-thal. In the α-thal with ID group haemoglobin (Hb), MCV, mean corpuscular Hb concentration (MCHC) and mean corpuscular Hb (MCH) values were significantly lower compared to the isolated α-thal. In the group with ID Hb, MCV, MCHC and MCH values were significantly lower compared to those with isolated α-thal. The α-thal with ID group, showed Hb, MCV, MCHC and MCH significantly reduced when compared to those with IDA. Conclusions: This study showed that the values of haematological parameters, especially haematocrit, Hb, MCV, MCH, MCHC and red blood cell distribution width (RDW), are lower in patients with IDA, especially when associated with α-thal and therefore it may be useful to discriminate between the different types of microcytic anaemia. [ABSTRACT FROM AUTHOR]

Objectives: Non-transfusion-dependent thalassemia includes a variety of phenotypes and genotypes that rarely require regular transfusions. However, these patients can experience a wide range of complications. The objective of this retrospective study was to verify whether there is a significant difference in non-transfusion-dependent thalassemia-related complications and treatment among males and females. Methods: We performed a re-analysis of samples evaluated in a previously published crosssectional study, regarding 96 non-transfusion-dependent thalassemia patients followed at the 'UOSD Malattie Rare del Globulo Rosso' Centre of the Cardarelli Hospital in Naples, Italy. Results: We found that females were more anemic than males, but there was no significant difference in prevalence of common complications among genders, except for hypogonadism. Furthermore, the transitory regular transfusions regimen in women who had been pregnant does not seem to have a significant impact on overall prognosis. Discussion: In non-transfusion-dependent thalassemia patients, the lower levels of hemoglobin found in females do not seem to indicate a higher prevalence of complications. Conclusion: This data should be considered in studies with experimental treatments aiming to correct anemia in patients with non-transfusion-dependent thalassemia. It should probably also be taken into account in order to set up different transfusion regimens among genders in transfusion-dependent patients. [ABSTRACT FROM AUTHOR]

Introduction: Thalassemia intermedia is a type of anemia which has several treatments modalities. We aimed to study the cost effectiveness of two treatments, including blood transfusion and hydroxyurea, in patients with beta-thalassemia intermedia in south of Iran referred to a referral center affiliated to Iran, Shiraz University of Medical Sciences in 2015. Materials and methods: This was a cost-effectiveness study which was conducted on 122 patients with beta-thalassemia intermedia. The indicator of effectiveness in this study was the reduction of growth disorder (normal BMI). Data analysis was done using SPSS 21, Excel 2010 and Treeage 2011. Finally, the one-way sensitivity analysis was performed to determine the robustness of the results. Results: The average annual costs of blood transfusion and the use of hydroxyurea in 2015 were 20733.27 purchasing power parity (PPP)$ and 7040.29 PPP$, respectively. The effectiveness of blood transfusion was57.4% while in hydroxyurea group was 60.7%. Conclusion: The results showed that the cost effectiveness of using hydroxyurea was more than that of blood transfusion. Therefore, it is recommended that the use of hydroxyurea in the treatment of patients with beta-thalassemia intermedia would become the first priority, and more basic and supplementary insurance coverage for treating such patients using hydroxyurea should be considered. [ABSTRACT FROM AUTHOR]

Objectives: The inherited genetic disorder beta0-thalassemia/Hb E disease is associated with the over-suppression of the master regulator of iron homeostasis, the peptide hormone hepcidin. How developing erythroid cells mediate the suppression of hepcidin remains controversial, although a number of inhibitors have been proposed. Methods: To investigate the ability of erythroid cells to suppress hepcidin expression in liver cells, conditioned media from the culture of in vitro differentiating erythroblasts (from normal controls and beta0-thalassemia/Hb E patients) was used to treat HepG2 cells, and the effects on hepcidin expression were assayed by real-time quantitative PCR and confocal microscopy. Results: Early activation followed by later suppression of hepcidin expression was seen posttreatment. Markedly, however, no significant differences were observed between suppression of hepcidin as mediated by media from the culture of erythroblasts from normal controls and beta0-thalassemia/Hb E patients Discussion: Previous studies investigating the suppression of hepcidin expression in beta0- thalassemia/Hb E disease have used patient-derived serum samples, which are complex fluids with contributions from multiple cell types. This study has developed a simple in vitro system that allows investigation of how a single cell type mediates hepcidin expression. The results support proposals that over-suppression of hepcidin seen in beta-thalassemia/Hb E patients is a consequence of the increased mass of erythropoiesis and not defects in the signaling process per se. Conclusion: The in vitro cell system developed here allows further investigation into the processes mediating erythroid cell suppression of liver hepcidin expression in both normal and pathological states. [ABSTRACT FROM AUTHOR]

Objectives: The aim of this study was to determine the anthropometric measurements in transfusion-dependent β-thalassemia children in Pakistan. The secondary aim was to correlate serum ferritin with the physical growth. Methods: We enrolled 367 children (aged 5-17 years) with transfusion-dependent beta-thalassemia major in the study. Anthropometric measurements, serum ferritin levels, and pre-transfusion hemoglobin levels were measured. Serum ferritin was correlated with the height z-score for age. Results: Laboratory evaluation showed that patients had significantly low mean pre-transfusion hemoglobin of 7.66 ± 1.34 g/dl (range 2.5-10.5) and high median (Q3-Q1) serum ferritin of 5012 ng/ml (6829-3532). The median (Q3-Q1) height-for-age z-score of children was low at −2.69 and (−1.46 to −3.80) and 65.4% children had stunted growth (height for age z-score <−2). There was a significant negative correlation between height for age z-score and serum ferritin levels (p < 0.000). Stunting of growth began early during 5-10 years of age but increased markedly with the progress of time. Conclusions: The study showed that children with beta thalassemia major had delayed physical growth possibly secondary to iron overload. Effective and early iron chelation is needed for preventing growth failure in transfusion-dependent beta thalassemia. [ABSTRACT FROM AUTHOR]

Objectives: To report the hematological and molecular features as well as diagnostic aspects of the hitherto un-described interactions of two rare α-globin chain variants with α0-thalassemia commonly found among Southeast Asian populations. Methods: The study was done on two adult Thai patients (P1 and P2) who had hypochromic microcytic anemia. Hb analysis was carried out using high performance liquid chromatography (HPLC) and capillary electrophoresis (CE). Mutations were identified by PCR and related techniques. Results: Hb analysis of P1 using HPLC showed a normal Hb pattern, but CE demonstrated an abnormal peak at zone 7. DNA sequencing identified a CCG-CTG mutation at codon 95 of the α2 globin gene corresponding to the Hb G-Georgia [α95(G2)Pro→Leu(α2)] previously undescribed in the Thai population. In contrast, Hb analysis of P2 demonstrated an abnormal peak not fully separated from Hb A on HPLC, but not on CE. DNA analysis identified the rarely described Hb Nakhon Ratchasima [α63(E12)Ala→Val(α2)] mutation. Routine DNA analysis detected the SEA deletion α0-thalassemia in trans to the Hb variants in both cases. Hematological parameters were compared with those of patients with compound heterozygote for other α-globin variants and α0-thalassemia previously documented. Conclusions: Identification of the patients confirmed that interaction of these rare Hb variants with α0-thalassemia does not lead to the Hb H disease. Differentiation of these two Hb variants from other clinically relevant hemoglobinopathies in a routine setting is, however, necessary. This can be accomplished using a combined Hb-HPLC and CE analysis followed by PCR-RFLP assays. [ABSTRACT FROM AUTHOR]

Objectives:There are more than 200 known mutations found in patients with β-thalassemia, a possibility to identify an unknown or novel mutation becomes less possible. Here, we report a novel mutation in a patient from Thailand who presented with chronic hemolytic anemia. Methods:A comprehensive hematology and DNA analysis was applied in the index patient and her mother. Results:Hematological and hemoglobin analyses were consistent with the clinical diagnosis of Hb E/β0-thalassemia. However, we could find only Hb E heterozygous mutation using our common polymerase chain reaction-based mutation detection of the β-globin genes. Furthermore, the molecular analysis demonstrated a novel T-deletion at codon 42 of the second exon of the β-globin gene which we named ‘Hb Yala’ according to the origin of this index family. Discussion:This mutation was assumed to generate a truncated β-globin chain terminating at codon 60 with possible unstable variant leading to a ‘null’ or β0-thalassemia. However, the clinical phenotype was surprisingly mild and no other ameliorating genetic factors, including co-inheritance of α-thalassemia and high propensity of Hb F byXmnI polymorphism, were found. Conclusion:This report has provided evidence that genotype–phenotype correlation in thalassemia syndromes is highly complex and a correct clinical severity classification of thalassemia should be mainly based on clinical evaluation. [ABSTRACT FROM AUTHOR]

Introduction:Thalassemia-related complications are one of the main factors that increase morbidity and mortality in aging patients with thalassemia. This study was aimed to report the prevalence and clinical risk factors for the complications in thalassemia. Methods:A multi-center prospective cohort study was conducted in patients with thalassemia aged ≥10 years old. Thalassemia-related complications were heart failure, pulmonary hypertension, extramedullary hematopoiesis, endocrine disorders, infections, thrombosis and leg ulcers. The clinical parameters significantly associated with the complications were analyzed by logistic regression methods. Results:The prevalence of thalassemia-related complications was 60.5% in patients with transfusion-dependent thalassemia (TDT) and 43% in patients with non-transfusion-dependent thalassemia (NTDT). Splenectomy was statistically associated with complications in both TDT and NTDT patients (adjusted odds ratio (AOR) = 7.4,p-value = 0.0001 and AOR = 2.6,p-value = 0.001). Age ≥50 years old (AOR = 2.9,p-value = 0.04) and female gender (AOR = 0.5,p-value = 0.03) were statistically associated with the complications in patients with NTDT. Conclusion:Nearly half of the patients in this cohort had disease-related complications. Splenectomy and advanced age were important factors for complication involvement. Early screening for the complications may reduce the morbidity and mortality in patients with thalassemia. [ABSTRACT FROM PUBLISHER]

Objective:β-Thalassemia is an inherited hemoglobin disorder resulting in chronic hemolytic anemia requiring chronic transfusion therapy. Cardiac involvement is the main cause of death in patients with thalassemia major. The narrow border is between overt myocardial dysfunction and clinically silent left ventricular (LV) dysfunction in patients with thalassemia. Therefore, we need novel parameters in different imaging techniques to discover cardiac involvement in an early and subtle stage. We explore to find a novel, straightforward and informative parameter in echocardiography as a noninvasive, economical and really routine in clinical practice. Methods:In this prospective study, 55 patients, who are known cases of β-thalassemia major, receiving long-term blood transfusions and undergoing iron chelation therapy were enrolled. Ferritin level, cardiac magnetic resonance (CMR) T2 * value, full conventional echocardiography and speckle tracking, LV regional circumferential and longitudinal strain values (%) and time-to-peak strain (ms) of 17 segments cardiac model in eyeball tomogram were measured. Results:There was a significant reduction in global longitudinal strain (GLS) (−20.9% ± 1.9 vs. −22.2 ± 1.03) and also basal segments longitudinal strain compared to normal subjects group (−17.4% ± 2.7 vs. −19.6% ± 1.2). There was no significant difference in circumferential strain value between thalassemia patients and normal control group. Interestingly, there was no significant correlation between GLS and CMR T2 * values showing no association between cardiac iron load and longitudinal strain. Conclusion:Speckle tracking echocardiography could be used as a feasible method for evaluating subclinical myocardial dysfunction in patients with thalassemia major. Echocardiography, using GLS, could predict clinically silent myocardial dysfunction independent of CMR (T2 * value) and extension of iron deposition. Our study also puts forward other causes such as chronic tissue hypoxia resulting from chronic anemia as a root cause and initiating factor for subsequent injury by the iron deposition. Speckle tracking can recognize the cardiac involvement in really early stages. [ABSTRACT FROM AUTHOR]

Background:Infant mortality due to sickle cell disease in sub-Saharan Africa is high, necessitating a better understanding of the modulating factors of the disease in this region. Methods:We assessed the hereditary persistence of foetal haemoglobin and α-thalassemia. We diagnosed 787 subjects, with or without sickle cell trait, by capillary electrophoresis in the Medical Diagnostic Laboratory of the CIRMF (Franceville, Gabon). Results:Heterocellular and pancellular forms of hereditary persistence of foetal haemoglobin occurred at low rates of 10.9 and 2.3%, respectively. The distribution of HbS levels in individuals with sickle cell trait was trimodal, showing a high percentage (52.4%) of heterozygous subjects with α-thalassemia. The distribution of HbA2 levels was bimodal in individuals without sickle cell trait, estimated to be comprised of 12 and 15% of α and β-thalassemic heterozygous subjects, respectively. Conclusions:In sub-Saharan Africa, α-thalassemia is a far more prevalent modulating factor than hereditary persistence of foetal haemoglobin. Our study highlights the need for further investigation of thalassemia, haemoglobinopathies that are neglected in sub-Saharan Africa. [ABSTRACT FROM PUBLISHER]

Objectives: Recent advancements have promoted the use of T2* magnetic resonance imaging (MRI) in the non-invasive detection of iron overload in various organs for thalassemia major patients. This study aims to determine the iron load in the heart and liver of patients with thalassemia major using T2* MRI and to evaluate its correlation with serum ferritin level and iron chelation therapy. Methods:This cross-sectional study included 162 subjects diagnosed with thalassemia major, who were classified into acceptable, mild, moderate, or severe cardiac and hepatic iron overload following their T2* MRI results, respectively, and these were correlated to their serum ferritin levels and iron chelation therapy. Results:The study found that 85.2% of the subjects had normal cardiac iron stores. In contrast, 70.4% of the subjects had severe liver iron overload. A significant but weak correlation (r = −0.28) was found between cardiac T2* MRI and serum ferritin, and a slightly more significant correlation (r = 0.37) was found between liver iron concentration (LIC) and serum ferritin. Discussion:The findings of this study are consistent with several other studies, which show that patients generally manifest with liver iron overload prior to cardiac iron overload. Moreover, iron accumulation demonstrated by T2* MRI results also show a significant correlation to serum ferritin levels. Conclusion:This is the first study of its kind conducted in Indonesia, which supports the fact that T2* MRI is undoubtedly valuable in the early detection of cardiac and hepatic iron overload in thalassemia major patients. [ABSTRACT FROM PUBLISHER]

Objectives:Thalassaemia is a potentially lethal inherited anaemia, caused by reduced or absent synthesis of globin chains. Measurement of the minor adult haemoglobin Hb A2, combining α- with δ-globin, is critical for the routine diagnosis of carrier status for α- or β-thalassaemia. Here, we aim to characterize a novel δ-globin variant, Hb A2Episkopi, in a single family of mixed Lebanese and Cypriot ancestry with mild hypochromic anaemia and otherwise normal globin genotype, which also presents with a coincidental 0.78-Mb sequence duplication on chromosome 1 (1q44) and developmental abnormalities. Methods: Analyses included comprehensive haematological analyses, cation-exchange high-performance liquid chromatography (CE-HPLC), cellulose acetate electrophoresis (CAE), Sanger sequencing and structure-based stability predictions for Hb A2Episkopi. Results: The GCT > GTT missense mutation, underlying Hb A2Episkopi, HBD:c.428C > T, introduces a cd142 codon change in the mature protein, resulting in reduced normal Hb A2amounts and a novel, less abundant Hb A2variant (HGVS: HBD:p.A143V), detectable as a delayed peak by CE-HPLC. The latter was in line with structure-based stability predictions, which indicated that the substitution of a marginal, non-helical and non-interface residue, five amino acids from the δ-globin chain carboxy-terminus, was moderately destabilizing. Discussion: Detection of the new variant depends on the diagnostic set-up and had failed by CAE and on an independent CE-HPLC system, which, in unfavourable circumstances, may lead to misdiagnoses of β-thalassaemia as α-thalassaemia. Given the mixed background of the affected family, the ethnic origin of the mutation is unclear, and this study thus suggests awareness for possible detection of Hb A2Episkopi in both the Cypriot and the Lebanese populations. [ABSTRACT FROM AUTHOR]

Background:Iron chelation therapy is one of the mainstays of the management of the patients with β-thalassemia (BT) major. Deferasirox is an oral active iron chelating agent. Proteinuria is one of the potential renal adverse effects of deferasirox, and monthly follow-up for proteinuria is suggested by Food and Drug Administration and European Medicine Agency. Methods:We aimed to investigate the necessity for monthly monitoring for proteinuria among patients with BT on deferasirox. A retrospective laboratory and clinic data review was performed for patients with BT major or intermedia who were treated with deferasirox chelation therapy. All patients were monitored for proteinuria for every 3 or 4 weeks after the initiation of deferasirox with serum creatinine and spot urine protein/creatinine ratios. Results:The median follow-up time of the 37 (36 BT major and one BT intermedia) patients was 44 months. Seven patients (18.9%) developed significant proteinuria (ratio ≥0.8). Of the 1490 measurements, 12 tests (0.8%) were proteinuric. Urine proteinuria resolved in all of the patients during the follow-up. The risk of proteinuria was higher at ages below a cut-off point of 23 years (p = 0.019). Patients, who were on deferasirox at doses above a cut-off dose of 29 mg/kg/day, were found to have higher risk of proteinuria development (p = 0.004). Conclusion:Proteinuria resolves without any complication or major intervention according to our results. Potentially more risky groups (age below 23 years old and receivers above a dose of 29 mg/kg/day) might be suggested to be followed monthly, besides monitoring all of the patients. [ABSTRACT FROM PUBLISHER]