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Fulltext A rare case of alpha-thalassaemia intermedia in a Malay patient double heterozygous for alpha(+)-thalassaemia and a mutation in alpha1 globin gene CD59 (GGC --> GAC)

George E, Jama T, Azian AS, Rahimah A, Zubaidah Z

Med J Malaysia, 2009 Dec;64(4):321-2.
PMID: 20954559

A rare case of thalassaemia-intermedia involving a non-deletion alpha thalassemia point mutation in the alpha1-globin gene CD59 (GGC --> GAC) and a deletion alpha+ (-alpha(3.7)) thalassaemia in which use of high performance liquid chromatography (HPLC) C-gram Hb subtype profile and DNA molecular analysis helped establish the diagnosis.
Fulltext Pre-transplantation serum ferritin as a prognostic marker in allogeneic haemopoietic stem cell transplant patients in a tertiary care hospital in Malaysia

Thambiah SC, George E, Samsudin IN, Hong LH, Chuo LL, Ramli N, et al.

Natl Med J India, 2016 May-Jun;29(3):136-140.
PMID: 27808061

BACKGROUND: The principal cause of iron overload in patients with haematological malignancies is recurrent red cell transfusions for anaemia. The serum ferritin level reflects the iron burden in the body, in the absence of inflammation or liver disease. In Malaysia, data are lacking on the association between pre-transplant serum ferritin levels and outcome after allogeneic haemopoietic stem cell transplant.

METHODS: We did a cross-sectional study using retrospective data of 106 post-allogeneic haemopoietic stem cell transplant patients (HLA-matched sibling) with haematological malignancies at Hospital Ampang to determine the relationship between pre-transplant serum ferritin levels and post-transplant outcome, post-transplant complications and survival time. Patients were divided into two groups according to the iron status: serum ferritin level >1000 μg/L (iron overload) and <1000 μg/L.

RESULTS: The median age for patients was 30.5 (18-58) years. The median pre-transplantation serum ferritin level and the prevalence of pre-transplantation iron overload were 2423 (408.2-7664) μg/L and 87.5%, respectively. No significant association was found between iron status and demographic factors, type of haematological malignancy and post-transplant complications. Although insignificant, patients with iron overload had a shorter survival time (36 months) compared to those with no iron overload (40 months). There was also no significant association between the iron status and post-transplant outcome. Significant post-transplant complications associated with post-transplant outcome were the need for total parenteral nutrition (TPN) (p=0.014) and chronic graft-versus-host disease (GVHD) (p=0.008). Similarly, significant associations were found between age group (p=0.003), TPN (p=0.035) and chronic GVHD (p=0.012) with survival time using Kaplan-Meir analysis. However, after Cox regression, only age group was found to be significantly associated with survival time (p=0.014).

CONCLUSION: Serum ferritin is an acute phase reactant and its levels increase in the presence of tissue necrosis and inflammation. Both these events occur in haematological malignancies. Although serum ferritin level is a non-invasive, relatively cost-effective, widely available and practical indicator of iron status, it is not specific to iron overload. Therefore, a true association between the serum ferritin level and iron burden is problematic in patients with haematological malignancies.
Molecular defects in the beta-globin gene identified in different ethnic groups/populations during prenatal diagnosis for beta-thalassemia: a Malaysian experience

Tan JA, George E, Tan KL, Chow T, Tan PC, Hassan J, et al.

Clin Exp Med, 2004 Dec;4(3):142-7.
PMID: 15599663 DOI: 10.1007/s10238-004-0048-x

Beta-thalassemia is the most-common genetic disorder of hemoglobin synthesis in Malaysia, and about 4.5% of the population are heterozygous carriers of the disorder. Prenatal diagnosis was performed for 96 couples using the Amplification Refractory Mutation System and Gap-Polymerase Chain Reaction. We identified 17 beta-globin defects-initiation codon for translation (T-G), -29 (A-G), -28 (A-G), CAP +1 (A-C), CD 8/9 (+G), CD 15 (G-A), CD 17 (A-T), CD 19 (A-G), Hb E (G-A), IVS1-1 (G-T), IVS1-5 (G-C), CD 41/42 (-CTTT), CD 71-72 (+A), IVS2-654 (CT), poly A(A-G), 100-kb Ggamma(Agammadeltabeta) degrees and 45-kb Filipino deletions. The 192 beta-alleles studied comprised Chinese (151 patients), Malay (21), Orang Asli from East Malaysia (15), Filipino (1), Indian (1), Indonesian Chinese (2), and Thai (1). In the Chinese, 2 beta-globin defects at CD 41/42 and IVS2-654 were responsible for 74% of beta-thalassemia. beta-mutations at CD 19, IVS1-1 (G-T), IVS1-5, poly A, and hemoglobin E caused 76% of the hemoglobin disorders in the Malays. The Filipino 45-kb deletion caused 73.3% of bthalassemia in the Orang Asli. Using genomic sequencing, the rare Chinese beta-mutation at CD 43 (G-T) was confirmed in 2 Chinese, and the Mediterranean mutation IVS1-1 (G-A) was observed in a Malay beta-thalassemia carrier. The beta-globin mutations confirmed in this prenatal diagnosis study were heterogenous and 65 (68%) couples showed a different globin defect from each other. The use of specific molecular protocols has allowed rapid and successful prenatal diagnosis of beta-thalassemia in Malaysia.
HbA2 levels in β-thalassaemia carriers with the Filipino β0-deletion: are the levels higher than what is found with non-deletional forms of β0-thalassaemia?

George E, Teh LK, Tan J, Lai MI, Wong L

Pathology, 2013 01;45(1):62-5.
PMID: 23222244 DOI: 10.1097/PAT.0b013e32835af7c1

AIMS: Classical carriers of β-thalassaemia are identified by a raised HbA2 level. Earlier studies indicated that the Filipino β-deletion has high raised HbA2 levels. The introduction of automated high performance liquid chromatography (HPLC) for thalassaemia screening is an important advance in technology for haematology laboratories. The BioRad Variant II Hb analyser is a common instrument used to quantify HbA2 levels in thalassaemia screening. This study aimed to determine HbA2 levels in carriers of Filipino β-mutation using the BioRad Variant II Hb analyser.
METHODS: The Filipino β-deletion was identified using gap-polymerase chain reaction (PCR) in the parents of transfusion dependent β-thalassaemia patients who were homozygous for the Filipino β-deletion in the indigenous population of Sabah, Malaysia. Hb subtypes were quantified on the BioRad Variant II Hb analyser. Concurrent α-thalassaemia was identified by multiplex gap-PCR for deletions and amplification refractory mutation system (ARMS)-PCR for non-deletional mutations.
RESULTS: The mean HbA2 level for Filipino β-thalassaemia trait was 5.9 ± 0.47 and with coinheritance of α-thalassaemia was 6.3 ± 0.44 (-α heterozygous) and 6.7 ± 0.36 (-α homozygous). The HbA2 levels were all >4% in keeping with the findings of classical β-thalassaemia trait and significantly higher than levels seen in non-deletional forms of β-thalassaemia.
CONCLUSION: The HbA2 level measured on the BioRad Variant II Hb analyser was lower than the level in the first description of the Filipino β-thalassaemia. β-thalassaemia trait with coinheritance of α-thalassaemia (-α) is associated with significantly higher HbA2 level.
Fulltext Hb E beta +-thalassaemia in west Malaysia: clinical features in the most common beta-thalassaemia mutation of the Malays [IVS 1-5 (G-->C)]

George E, Wong HB

Singapore Med J, 1993 Dec;34(6):500-3.
PMID: 8153710

Patients with the Hb beta + [IVS 1-5 (G-->C)] clinically presented as beta-thalassaemia intermedia and remained asymptomatic in the absence of blood transfusions. With or without blood transfusions the patients were short and had moderate to marked thalassaemia facies. Children who received blood transfusions showed progressive iron loading with age. The serum ferritin and serum alanine transaminase levels were significantly raised in the patients who were given blood transfusions. In the presence of blood transfusions, and absence of adequate iron chelation therapy, splenectomy became an inevitable event at some stage of the disease because of increasing transfusing requirements.
Fulltext Hereditary ovalocytosis in Malays

George E, Mohandas N, Duraisamy G, Adeeb N, Zainuddin ZA, Teng MS, et al.

Med J Malaysia, 1988 Dec;43(4):327-31.
PMID: 3241598
Fulltext Inhibitory action of gonadotropin-inhibitory hormone on the signaling pathways induced by kisspeptin and vasoactive intestinal polypeptide in GnRH neuronal cell line, GT1-7

Son YL, Ubuka T, Soga T, Yamamoto K, Bentley GE, Tsutsui K

FASEB J, 2016 06;30(6):2198-210.
PMID: 26929433 DOI: 10.1096/fj.201500055

Gonadotropin-inhibitory hormone (GnIH) acts as a negative regulator of reproduction by acting on gonadotropes and gonadotropin-releasing hormone (GnRH) neurons. Despite its functional significance, the molecular mechanism of GnIH action in the target cells has not been fully elucidated. To expand our previous study on GnIH actions in gonadotropes, we investigated the potential signal transduction pathway that conveys the inhibitory action of GnIH in GnRH neurons by using the GnRH neuronal cell line, GT1-7. We examined whether GnIH inhibits the action of kisspeptin and vasoactive intestinal polypeptide (VIP), positive regulators of GnRH neurons. Although GnIH significantly suppressed the stimulatory effect of kisspeptin on GnRH release in hypothalamic culture, GnIH had no inhibitory effect on kisspeptin stimulation of serum response element and nuclear factor of activated T-cell response element activities and ERK phosphorylation, indicating that GnIH may not directly inhibit kisspeptin signaling in GnRH neurons. On the contrary, GnIH effectively eliminated the stimulatory effect of VIP on p38 and ERK phosphorylation, c-Fos mRNA expression, and GnRH release. The use of pharmacological modulators strongly demonstrated the specific inhibitory action of GnIH on the adenylate cyclase/cAMP/protein kinase A pathway, suggesting a common inhibitory mechanism of GnIH action in GnRH neurons and gonadotropes.-Son, Y. L., Ubuka, T., Soga, T., Yamamoto, K., Bentley, G. E., Tsutsui, K. Inhibitory action of gonadotropin-inhibitory hormone on the signaling pathways induced by kisspeptin and vasoactive intestinal polypeptide in GnRH neuronal cell line, GT1-7.
A molecular marker associated with mild hemoglobin H disease

George E, Ferguson V, Yakas J, Kronenberg H, Trent RJ

Pathology, 1989 Jan;21(1):27-30.
PMID: 2762043

The clinical spectrum of HbH disease varies from a benign disorder to a severe anemia which is blood-transfusion dependent. Heterogeneity at the clinical level is now being understood in terms of the underlying molecular defects. In this study a mild phenotype found in a group of patients with HbH disease is associated with two types of alpha-thalassemia. These are: alpha+-thalassemia (-alpha 3.7/) and alpha 0-thalassemia (--SEA/). In contrast, a second group with more severe HbH disease has a non-deletional alpha-thalassemia defect instead of alpha+-thalassemia (genotype alpha alpha T/--SEA). In the majority of cases, the basis for non-deletional alpha-thalassemia is Hb Constant Spring.
Fulltext Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1

Klionsky DJ, Abdel-Aziz AK, Abdelfatah S, Abdellatif M, Abdoli A, Abel S, et al.

Autophagy, 2021 Jan;17(1):1-382.
PMID: 33634751 DOI: 10.1080/15548627.2020.1797280

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.
Fulltext Genetic variants in HBS1L-MYB with Hb subtypes level among Filipino β°-deletion carriers coinherited with -α3.7 deletion

Lim, L. N., Yu, K. S., Chua, S. M., George, E., Lai, M. I., Wong, L., et al.

Malaysian Journal of Medicine and Health Sciences, 2019;15(102):47-47.
MyJurnal

Introduction: Filipino β°-deletion is predominant among the β-thalassaemia patients in the indigenous population of Sabah, Malaysia particularly among the Kadazandusun. Individuals who co-inherit with α- and β-thalassaemia will demonstrate milder clinical symptoms with modified complete blood count (CBC) and Hb subtype parameters. HBS1L-MYB variants act as one of the key regulator of haematopoiesis and erythropoiesis and display strong association
with variation of HbF levels. Therefore, this study aims to evaluate the association between genetic variants in HBS1L-MYB with Hb subtypes level among Filipino β°-deletion carriers co-inherited with -α3.7 deletion. Methods: Filipino β°-deletion and -α3.7 deletion were identified using gap-polymerase chain reaction (PCR). A total of 34 subjects found with coinheritance of Filipino β°-deletion and -α3.7 deletion were subjected for HBS1L-MYB intergenic polymorphisms (HMIP) analysis. Hb subtypes level were quantified using BioRad Variant II Hb analyser. Genotyping of HBS1L-MYB variants rs9399137 and rs11759553 was done using own designed tetra primer ARMS-PCR. Results: The minor allele frequencies (MAF) of the two HMIP is found more than 0.05 (rs11759553, MAF=0.18 and rs9399137, MAF=0.15), indicating the significance of these variants among the study subjects. Significant difference was found between HbF level and HBS1L-MYB variant rs11759553 with p-value less than 0.05 (p=0.001). Subjects with homozygous genotype for rs11759553 (T/T) was found with higher HbF, followed by heterozygous (A/T) and wild type (A/A). rs11759553 and rs9399137 was found did not influence the level of HbA and HbA2. HMIP of rs11759553 and rs9399137 are found significant among Filipino β°-deletion carriers co-inherited with -α3.7deletion with its high minor allelic frequency and high HbF level. Strong association with HbF level was demonstrated when
coinheritance of rs11759553. Conclusion: This study demonstrates that there are significant associations between certain genetic variants in HBS1L-MYB with Hb subtypes level among Filipino β°-deletion carriers co-inherited with -α3.7 deletion.
Fulltext Identification of αo-thalassaemia (–SEA) using an Enzyme- Linked Immunosorbent Assay (ELISA) for embryonic zeta-globin chain detection – a preliminary study

Tan, J. A. M. A., George, E., Lim, E. J., Zakaria, Z., Hassan, R., Wee, Y. C., et al.

Malaysian Journal of Medicine and Health Sciences, 2006;2(2):81-87.
MyJurnal

Objectives: This study aimed to evaluate the UBI MAGIWELTM ζ-GLOBIN ELISA Kit for the presumptive diagnosis of αo-thalassaemia. The ELISA results obtained were confirmed by molecular characterisation of αo-thalassaemia using a Duplex-PCR. Methods: Routine peripheral blood counts and red cell indices were determined in 94 blood samples sent for Hb analysis. Hb subtypes were quantified by high performance liquid chromatography (HPLC) and Hb electrophoresis conducted on agarose gel at pH 8.5. Zeta-globin chain levels were determined using the UBI MAGIWELTM ζ-GLOBIN ELISA Kit. Molecular analysis was performed using a duplex-PCR which simultaneously amplifies
a normal 136 bp sequence between the ψα−α2-globin genes and a 730 bp Southeast Asian deletion-specific sequence (–SEA) between the ψα2−θ1-globin genes. Results: Using the ELISA assay kit, 20 blood samples were presumptively identified as α-thalassaemia carriers from elevated ζ-globin chains (OD>0.3) while the remaining 74 blood samples showed OD
Fulltext Beta thalassaemia mutations in Malays: a simplified cost-effective strategy to identify the mutations

George, E., Teh, L.K., Rosli, R., Lai, M.I., Tan, J.A.M.A.

Malaysian Journal of Medicine and Health Sciences, 2012;8(1):45-53.
MyJurnal

Beta (β)- thalassaemia is a public health problem in Malaysia. The carrier rate is estimated to be 4.5% by micro-mapping studies particularly among Malays who comprise 53.5% of the population in Malaysia. The common diagnostic method in Malaysia for mutation detection is by amplification refractory mutation system (ARMS). It allows single mutation detection in each reaction but is labour intensive and time consuming when many mutations need to be identified. The purpose of this study was to develop a diagnostic tool for effective mutation detection of beta thalassaemia in Malay patients and compare its efficacy with ARMS-PCR, the current method in use. Methods: Reverse dot blot hybridization (RDBH) technique was incorporated in the development of two strip assays [RDBH-Strip M(6) and RDBH-Strip C(6)] to identify common beta thalassaemia mutations in the Malays. The panels of selected mutations were based on the mutation frequencies in Malaysia reported in previous studies. RDBH-Strip M(6) was applied as step 1 and RDBH-Strip C(6) was applied as step 2 for unidentified mutations. The strips were validated with the gold standard method, ARMS- PCR. Results: One hundred and thirty seven Malay patients with 274 alleles were studied. In Step 1 mutation detection, 238 alleles (86.9%) were identified in 119 of patients by RDBH-Strip M(6). Step 2 resulted in a further detection of 20 alleles in another 10 patients by RDBH-Strip C(6). The combination of both strips resulted in the identification of 258 alleles in 129 (94.6%) of 137 Malay patients. The strip assays were 100% sensitive and specific when compared with ARMS-PCR method. Conclusion: Two strip assays utilising the RDBH technique were developed to identify common β-thalassaemia mutations in Malays. The RDBH Strip M(6) identified 86.9% of the mutations and the RDBH-Strip C (6) detected further 7.3% alleles. This two step strategy was found to be rapid and cost effective for the direct diagnosis of β-thalassaemia mutations in the Malays. The remaining unidentified mutations would require DNA sequencing. It can serve as a specific molecular diagnostic tool for effective diagnosis of
β-thalassaemia mutations in this ethnic group.
Fulltext Public perceptions and attitudes toward thalassaemia: Influencing factors in a multi-racial population

Wong LP, George E, Tan JA

BMC Public Health, 2011;11:193.
PMID: 21447191 DOI: 10.1186/1471-2458-11-193

Thalassaemia is a common public health problem in Malaysia and about 4.5 to 6% of the Malays and Chinese are carriers of this genetic disorder. The major forms of thalassaemia result in death in utero of affected foetuses (α-thalassaemia) or life-long blood transfusions for survival in β-thalassaemia. This study, the first nationwide population based survey of thalassaemia in Malaysia, aimed to determine differences in public awareness, perceptions and attitudes toward thalassaemia in the multi-racial population in Malaysia.
Alpha-thalassaemia in association with beta-thalassaemia patients in Malaysia: a study on the co-inheritance of both disorders

Wee YC, Tan KL, Kuldip K, Tai KS, George E, Tan PC, et al.

Community Genet, 2008;11(3):129-34.
PMID: 18376108 DOI: 10.1159/000113874

BACKGROUND/AIMS: Individuals with double heterozygosity for alpha- and beta-thalassaemia and heterozygous beta-thalassaemia show a similar haematological picture. Co-inheritance of alpha- and beta-thalassaemia in both partners may result in pregnancies with either Hb Bart's hydrops foetalis or beta-thalassaemia major, or pregnancies with both disorders.
METHODS: The co-inheritance of alpha-thalassaemia in 322 beta-thalassaemia carriers in Malaysia was studied.
RESULTS: The frequency of alpha-thalassaemia in the beta-thalassaemia carriers was 12.7% (41/322), with a carrier frequency of 7.8% for the SEA deletion, 3.7% for the -alpha(3.7) deletion, 0.9% for Hb Constant Spring and 0.3% for the -alpha(4.2) deletion.
CONCLUSION: Double heterozygosity for alpha- and beta-thalassaemia was confirmed in 5 out of the 41 couples and the risk of the fatal condition Hb Bart's hydrops foetalis was confirmed in two of these couples. Detection of the Southeast Asian (SEA) deletion in the Malaysian Malays in this study confirms that Hb Bart's hydrops foetalis can occur in this ethnic group. Results of this study have provided new information on the frequency and different types of alpha-thalassaemia (--(SEA), -alpha(3.7) and -alpha(4.2) deletions, Hb Constant Spring) in Malaysian beta-thalassaemia carriers.
Fulltext Molecular characterisation and frequency of Ggamma Xmn I polymorphism in Chinese and Malay beta-thalassaemia patients in Malaysia

Wong YC, George E, Tan KL, Yap SF, Chan LL, Tan JA

Malays J Pathol, 2006 Jun;28(1):17-21.
PMID: 17694955

The molecular basis of variable phenotypes in P-thalassaemia patients with identical genotypes has been associated with co-inheritance of alpha-thalassaemia and persistence of HbF production in adult life. The Xmn I restriction site at -158 position of the Ggamma-gene is associated with increased expression of the Ggamma-globin gene and higher production of HbF This study aims to determine the frequency of the digammaferent genotypes of the Ggamma Xmn I polymorphism in P-thalassaemia patients in two ethnic groups in Malaysia. Molecular characterisation and frequency of the Ggamma Xmn I polymorphism were studied in fifty-eight Chinese and forty-nine beta-thalassaemia Malay patients by Xmn I digestion after DNA amplification of a 650 bp sequence. The in-house developed technique did not require further purification or concentration of amplified DNA before restriction enzyme digestion. The cheaper Seakem LE agarose was used instead of Nusieve agarose and distinct well separated bands were observed. Genotyping showed that the most frequent genotype observed in the Malaysian Chinese was homozygosity for the absence of the Xmn I site (-/-) (89.7%). In the Malays, heterozygosity of the Xmn I site (+/-) was most common (63.3%). Homozygosity for the Xmn I site (+/+) was absent in the Chinese, but was confirmed in 8.2% of the Malays. The ratio of the (+) allele (presence of the Xmn I site) to the (-) allele (absence of the Xmn I site)) was higher in the Malays (0.66) compared to the Chinese (0.05). The (+/-) and (+/+) genotypes are more commonly observed in the Malays than the Chinese in Malaysia.
Fulltext Specific and straightforward molecular investigation of β-thalassemia mutations in the Malaysian Malays and Chinese using direct TaqMan genotyping assays

Kho SL, Chua KH, George E, Tan JA

Genet. Mol. Res., 2013;12(3):2409-15.
PMID: 23479149 DOI: 10.4238/2013.February.28.4

Beta-thalassemia is a life-threatening inherited blood disorder. Rapid characterization of β-globin gene mutations is necessary because of the high frequency of Malaysian β-thalassemia carriers. A combination real-time polymerase chain reaction genotyping assay using TaqMan probes was developed to confirm β-globin gene mutations. In this study, primers and probes were designed to specifically identify 8 common β-thalassemia mutations in the Malaysian Malay and Chinese ethnic groups using the Primer Express software. "Blind tests" using DNA samples from healthy individuals and β-thalassemia patients with different genotypes were performed to determine the specificity and sensitivity of this newly designed assay. Our results showed 100% sensitivity and specificity for this novel assay. In conclusion, the TaqMan genotyping assay is a straightforward assay that allows detection of β-globin gene mutations in less than 40 min. The simplicity and reproducibility of the TaqMan genotyping assay permit its use in laboratories as a rapid and cost-effective diagnostic tool for confirmation of common β-thalassemia mutations in Malaysia.
Fulltext High throughput molecular confirmation of β-thalassemia mutations using novel TaqMan probes

Kho SL, Chua KH, George E, Tan JA

Sensors (Basel), 2013;13(2):2506-14.
PMID: 23429513 DOI: 10.3390/s130202506

β-Thalassemia is a public health problem where 4.5% of Malaysians are β-thalassemia carriers. The genetic disorder is caused by defects in the β-globin gene complex which lead to reduced or complete absence of β-globin chain synthesis. Five TaqMan genotyping assays were designed and developed to detect the common β-thalassemia mutations in Malaysian Malays. The assays were evaluated with 219 "blinded" DNA samples and the results showed 100% sensitivity and specificity. The in-house designed TaqMan genotyping assays were found to be cost- and time-effective for characterization of β-thalassemia mutations in the Malaysian population.
Fulltext A novel gap-PCR with high resolution melting analysis for the detection of α-thalassaemia Southeast Asian and Filipino β°-thalassaemia deletion

Kho SL, Chua KH, George E, Tan JA

Sci Rep, 2015;5:13937.
PMID: 26365497 DOI: 10.1038/srep13937

Homozygosity for the α-thalassaemia Southeast Asian (α-SEA) and Filipino β°-thalassaemia (β-FIL) deletions can cause serious complications leading to foetal death or life-long blood transfusions. A rapid and accurate molecular detection assay is essential in populations where the deletions are common. In this study, gap-polymerase chain reaction (PCR) with high resolution melting (HRM) analysis was developed to detect both the large deletions. Melting curves at 86.9 ± 0.1 °C were generated by normal individuals without the α-SEA deletion, 84.7 ± 0.1 °C by homozygous α-SEA deletion individuals and two melting curves at 84.7 ± 0.1 °C and 86.9 ± 0.1 °C by α-SEA deletion carriers. Normal individuals without the β-FIL deletion produce amplicons with a melting temperature (Tm) at 74.6 ± 0.1 °C, homozygous β-FIL individuals produce amplicons with Tm at 73.6 ± 0.1 °C and heterozygous β-FIL individuals generate two amplicons with Tm at 73.6 ± 0.1 °C and 74.6 ± 0.1 °C. Evaluation using blinded tests on 220 DNA samples showed 100% sensitivity and specificity. The developed assays are sensitive and specific for rapid molecular and prenatal diagnosis for the α-SEA and β-FIL deletions.
Fulltext A holistic approach to education programs in thalassemia for a multi-ethnic population: consideration of perspectives, attitudes, and perceived needs

Wong LP, George E, Tan JA

J Community Genet, 2011 Jun;2(2):71-9.
PMID: 22109791 DOI: 10.1007/s12687-011-0039-z

Hemoglobin disorders which include thalassemias are the most common heritable disorders. Effective treatment is available, and these disorders can be avoided as identification of carriers is achievable using simple hematological tests. An in-depth understanding of the awareness, attitudes, perceptions, and screening reservations towards thalassemia is necessary, as Malaysia has a multi-ethnic population with different religious beliefs. A total of 13 focus group discussions (70 participants) with members of the general lay public were conducted between November 2008 and January 2009. Lack of knowledge and understanding about thalassemia leads to general confusions over differences between thalassemia carriers and thalassemia major, inheritance patterns, and the physical and psychologically impact of the disorder in affected individuals and their families. Although most of the participants have not been tested for thalassemia, a large majority expressed willingness to be screened. Views on prenatal diagnosis and termination of fetuses with thalassemia major received mixed opinions from participants with different religions and practices. Perceived stigma and discrimination attached to being a carrier emerged as a vital topic in some group discussions where disparity in the answers exhibited differences in levels of participants' literacy and ethnic origins. The two most common needs identified from the discussion were information and screening facilities. Participants' interest in knowing the severity of the disease and assessing their risk of getting the disorder may imply the health belief model as a possible means of predicting thalassemia public screening services. Findings provide valuable insights for the development of more effective educational, screening, and prenatal diagnostic services in the multi-ethnic Asian society.
Fulltext Molecular characterisation of Haemoglobin Constant Spring and Haemoglobin Quong Sze with a Combine-Amplification Refractory Mutation System

Wee YC, Tan KL, Chua KH, George E, Tan JA

Malays J Med Sci, 2009 Jul;16(3):21-8.
PMID: 22589661 MyJurnal

BACKGROUND: The interaction of the non-deletional α(+)-thalassaemia mutations Haemoglobin Constant Spring and Haemoglobin Quong Sze with the Southeast Asian double α-globin gene deletion results in non-deletional Haemoglobin H disease. Accurate detection of non-deletional Haemoglobin H disease, which is associated with severe phenotypes, is necessary as these mutations have been confirmed in the Malaysian population.
METHODS: DNA from two families with Haemoglobin H disease was extracted from EDTA-anticoagulated whole blood and subjected to molecular analysis for α-thalassaemia. A duplex polymerase chain reaction was used to detect the Southeast Asian α-globin gene deletion. Polymerase chain reaction-restriction fragment length polymorphism analysis was then carried out to determine the presence of Haemoglobin Constant Spring and Haemoglobin Quong Sze. A combine-amplification refractory mutation system protocol was optimised and implemented for the rapid and specific molecular characterisation of Haemoglobin Constant Spring and Haemoglobin Quong Sze in a single polymerase chain reaction.
RESULTS AND CONCLUSIONS: The combine-amplification refractory mutation system for Haemoglobin Constant Spring and Haemoglobin Quong Sze, together with the duplex polymerase chain reaction, provides accurate pre- and postnatal diagnosis of non-deletional Haemoglobin H disease and allows detailed genotype analyses using minimal quantities of DNA.
KEYWORDS: Combine-ARMS; Hb Constant Spring; Hb Quong Sze; medical sciences

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