Original Communication - Detection of two rare β-thalassemia mutations [-90 (C → T) and CD 26 (C →T)] among Indians

BACKGROUND: β -Thalassemia (β-thal) is present in practically every caste group in Indians. Molecular characterization of β-thal in these groups has revealed an extremely heterogeneous picture. AIM: To identify all the mutations and to detect the novel mutations using a versatile mutation detection technique. MATERIALS AND METHODS: Denaturing gradient gel electrophoresis (DGGE) has been established to scan the entire β-globin gene to localize the mutation followed by DNA sequencing for characterization. The DNA samples from two families referred to us either for prenatal diagnosis or for DNA studies were studied. RESULTS: Atypical DGGE patterns in fragments B & A indicating the presence of the mutation, have been detected in both the families. DNA sequencing revealed two rare patterns fragments with patterns in fragments β-thal mutations [CD 26 (C→T) and -90 (C →T)]. CONCLUSION: DGGE is a useful mutation detection technique to identify β-thal mutations among the heterogeneous Indian population

Detection of fetal mutations causing hemoglobinopathies by non-invasive prenatal diagnosis from maternal plasma

Background: Prenatal diagnosis of hemoglobinopathies enables couples at risk to have a healthy child. Currently used fetal sampling procedures are invasive with some risk of miscarriage. A non-invasive approach to obtain fetal deoxyribonucleic acid (DNA) for diagnosis would eliminate this risk. Aim: To develop and evaluate a non-invasive prenatal diagnostic approach for hemoglobinopathies using cell-free fetal DNA circulating in the maternal plasma. Settings and Design: Couples referred to us for prenatal diagnosis of hemoglobinopathies where the maternal and paternal mutations were different were included in the study. Materials and Methods: Maternal peripheral blood was collected at different periods of gestation before the invasive fetal sampling procedure was done. The blood was centrifuged to isolate the plasma and prepare DNA. A size separation approach was used to isolate fetal DNA. Nested polymerase chain reaction (PCR)-based protocols were developed for detection of the presence or absence of the paternal mutation. Results and Conclusions: There were 30 couples where the parental mutations were different. Of these, in 14 cases the paternal mutation was absent and in 16 cases it was present in the fetus. Using cell-free fetal DNA from maternal plasma, the absence of the paternal mutation was accurately determined in 12 of the 14 cases and the presence of the paternal mutation was correctly identified in 12 of the 16 cases. Thus, this non-invasive approach gave comparable results to those obtained by the conventional invasive fetal sampling methods in 24 cases giving an accuracy of 80.0%. Although the nested PCR approach enabled amplification of small quantities of cell-free DNA from maternal plasma at different periods of gestation after size separation to eliminate the more abundant maternal DNA, an accurate diagnosis of the presence or absence of the paternal mutation in the fetus was not possible in all cases to make it clinically applicable

Analysis of Complement Receptor Type 1 (CR1) Polymorphisms and Its Association With Malaria in Rural Population of Maharashtra

The interaction between human host and the Plasmodium parasite is complex. The factors affecting the causality of infection and its severity are yet not completely understood. Single Nucleotide Polymorphisms (SNP) associated with CR1 may be associated with patho-physiology of malaria and its susceptibility to the disease. Methods: The objective of the present study was to calculate the incidence of various antigens of Knops blood group system and CR1 Exon22 polymorphisms in rural population from Chiplun Taluka of Ratnagiri district. The study included 112 malaria positive cases and 909 healthy controls, which were screened for CR1 Exon22 polymorphism. Knops (Kna/b), McCoy (McCa/b), Swain-Langley (Sl1/2) polymorphisms were screened in 93 cases and 321 healthy controls. The frequencies were determined using a PCR-RFLP technique. Results: Only wild types of the allele form were observed in Knops blood group system in malaria cases and healthy control. CR1 exon22 polymorphism was seen in the study population with all the 3 allele type distributed in the cases and control samples. No significant allelic or genotypic differences were found between the controls and the disease groups. Conclusion: The results of the present study demonstrate that common CR1 Exon22 and Knops blood group system are not associated with malaria in the endemic area

Red Cell Genetic Abnormalities, b-Globin Gene Haplotypes, and APOB Polymorphism in the Great Andamanese, A Primitive Negrito Tribe of Andaman and Nicobar Islands, India

The Great Andamanese are a primitive Negrito tribe of the Andaman and Nicobar Islands, India, with a total population of 37. We studied 29 individuals from eight families from this population for abnormal hemoglobins, G6PD deficiency, DNA haplotypes, and apolipoprotein B (APOB, gene) polymorphism. Hb E was detected in five individuals, the prevalence of Hb E heterozygotes being 14.3%. One individual had b-thalassemia trait. One female was G6PD deficient and showed the G6PD Orissa mutation. Haplotype analysis of the b-globin gene cluster showed that the bE chromosomes were linked to two haplotypes (– – – – – + + and + + – + + + +) representing the framework 1 gene, whereas the bA chromosomes showed eight different haplotypic patterns corresponding to framework 1 and 3 genes. APOB polymorphism analysis showed that the 631-base-pair (bp) allele was the predominant one with a high homozygosity rate, which could be due to the higher rate of inbreeding in this isolated group. The presence of Hb E and our findings on haplotype analysis supports the hypothesis that the Great Andamanese are reasonably believed to be the surviving representatives of the Negrito race that once flourished in the entire Southeast Asian region in ancient times

A holistic approach to education programs in thalassemia for a multi-ethnic population: consideration of perspectives, attitudes, and perceived needs

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