Snehalata C. Gupte

HLA Antigen Distribution in Marathi Speaking Hindu Population from Mumbai, Maharastra, India

Three hundred and ninety two unrelated Marathi speaking Hindu people residing in Mumbai, Maharastra, (Western India) were studied for HLA A, B and C locus antigen profiles. The phenotypic frequencies of HLA A1, A2, A9, A11, A24, A33, B5, B7, B35, B40 and Cw4 were increased while frequencies of HLA A10, A23, A28, A31, B14, B16, B18, B21, B37, B39, B50, Cw1, Cw5 and Cw8 were decreased in the Marathi speaking Hindus. The genotype frequencies of HLA A9, A24, A33 and B40 were increased while that of A11, A28, A31, B15, B21, B37, B50, Cw1 and Cw8 were decreased when compared with gene frequencies of other Indians Hindu populations reported. Two loci haplotype analysis revealed that A1-B17, A10-B8 and A19-B12 were common Indian Hindu haplotypes where as A3-B35, A19-B15 and A111-B40 were unique for the Marathi Hindus. Haplotype A2-B4 0 observed in Marathi Hindus were also observed among South Indian Hindus, while A11B35 have been observed among immigrant Indian Hindus. Another halpotype A3-B7 reported from both south Indian and north Indian Hindus was not observed in Marathi Hindus. Significantly negative linkage disequilibrium was observed for haplotypes A19-B40 and A2-B7 in Marathi Hindus. Thus the observed antigen frequencies and linkage disequilibrium in Marathi Hindus suggest the influence of genetic drift caused by selection, geography and culture. Further the study reveals that the Hindu population of India cannot be considered as a single panmictic population with reference to genetic characteristics.

Hb QIndia and its interaction with β-thalassaemia : a study of 64 cases from India

Abstract Haemoglobin Q (Hb Q), a relatively uncommon α-chain structural Hb variant, has been reported either in the heterozygous state or interacting with β-thalassaemia. Individuals inheriting Hb Q generally are asymptomatic and are diagnosed by chance during population screening or as a part of a family study. This paper represents the first large study from India of 64 cases of Hb Q, documenting the haematological and molecular findings on 36 cases of Hb Q trait, 22 of Hb Q β-thalassaemia trait and three of Hb Q β-thalassaemia major, as well as a family of Hb Q homozygous cases. Hb Q is detected by Hb electrophoresis and chromatography. Hb Q levels in homozygous cases ranged from 32% to 35%, while in Hb Q heterozygotes the level was 20%. When there was an interaction of β-thalassaemia heterozygotes the level was 14%, and in interacting β-thalassaemia homozygotes the levels range from 7% to 9%. β-thalassaemia mutations were characterised in cases showing elevated Hb A2 levels, which were markedly reduced in the majority of cases in which β-thalassaemia was absent. Hb Q is rare and not a single homozygous case has been reported. However, Hb Q disease showed wide variation in clinical and haematological presentation in the same family.

Is Cellulose Acetate Electrophoresis a Suitable Technique for Detection of Hb Bart’s at Birth?

Symptomatic alpha-thalassemia (alpha-thal) as found in South-East Asia is uncommon in India. However, the presence of Hb Barts in cord blood samples has been reported from different parts of India and the prevalence of alpha-thal has ranged from 0.5 to 18% by different electrophoretic techniques. The methodology utilised has ranged from paper electrophoresis to isoelectric focussing (IEF). We screened 798 cord bloods for the presence of Hb Barts by cellulose acetate electrophoresis and found a prevalence rate of alpha-thal of 15.3% in a heterogenous population in Bombay. A comparison of four different electrophoretic techniques for detection of Hb Barts in 138 neonates showed that cellulose acetate and starch gel electrophoresis were by and large comparable and only a little less sensitive than IEF. Paper electrophoresis used at many centers in India was most insensitive. As alpha-genotyping is not possible at most centers in the country, it is suggested that a simple cellulose acetate electrophoresis would be the method of choice for screening neonates for alpha-thal in India. As a part of our follow-up study, alpha-genotyping was done by Southern blot hybridization in 24 cases who had shown variable levels of Hb Barts at birth. The rightward deletion (-alpha3.7/) either in a heterozygous or homozygous condition was the only gene defect encountered in this preliminary study. However, 7 of 24 cases (29.17%) showed no correlation between Hb Barts level and alpha-genotypes.

Study of parents of β-thalassemia major children to determine cutoff values of hematological parameters for diagnosis of β-thalassemia trait and assessment of anemia in them

BACKGROUND Because of the overlapping MCV, MCH and HbA 2 values in BTT and non-BTT subjects our laboratory determined own cutoffs. AIMS To establish cutoff values by investigating the parents of thalassemia major children and to assess the degree of anemia in BTT subjects. MATERIALS AND METHODS Study includes 179 parents of thalassemia major children (BTT cases) and 287 non-BTT controls. Samples were analyzed on an electronic hematology analyzer. The samples having MCV ≤ 76 fl and MCH ≤ 27 pg were quantified for HbA 2 by cellulose acetate electrophoresis and grey zone samples were confirmed on HPLC. Statistical Analysis Mean ± SD, sensitivity, specificity, PPV, NPV and accuracy were calculated. The histograms were plotted for MCV, MCH and HbA 2 . RESULTS Cases having MCV ≤ 76 fl and MCH ≤ 27 pg if considered as suspected cases of BTT then we could have missed five known BTT samples. Sensitivity increased to 100% in all three diagnostic parameters when the cutoff values were raised and specificity for MCV and MCH was decreased. But specificity was 100% with raised cutoff for HbA 2 . Hb and HCT mean values were significantly reduced in BTT cases compared to controls (P < 0.001). In 100% females and 84.9% males having BTT, mild to moderate anemia was observed. CONCLUSION In our setup, the cutoff values are MCV (≤78.0 fl), MCH (≤28 pg) and HbA 2 (>3.8%) for BTT diagnosis and there is a mild to moderate anemia in BTT cases.

Molecular characterization of β-thalassemia in four communities in South Gujarat—codon 30 (G → A) a predominant mutation in the Kachhiya Patel community

Different thalassemia mutations have been reported in various ethnic groups and geographical regions in India. In this study, we have investigated Kachhiya Patel, Dhodia Patel, Modh Bania, and Muslim communities of Surat, Gujarat to identify molecular defects causing β-thalassemia in them. Covalent reverse dot blot hybridization technique was used to detect six common Indian β-thalassemia mutations while the seventh mutation (619-bp deletion) was identified by PCR. The less common mutations were detected by amplification refractory mutation and the uncharacterized samples were directly sequenced. Characterization of β-thalassemia mutations was carried out in a total of 175 unrelated β-thalassemia trait cases. We identified IVS 1 nt 5 (G → C) in 31 out of 65 Muslims, codon (Cd) 41/42 (−CTTT) in 14 out of 16 in Modh Banias, Cd 15 (G → A) in 19 out of 24 Dhodia Patels. The most significant observation was an uncommon mutation; Cd 30 (G → A) detected in 61 out of 70 Kachhiya Patels. The 619-bp deletion was detected in 6 out of 10 Muslim-Memons. Many other rare mutations like Cd 15 (−T), Cd 8 (−AA), −88 (C → A), Capsite +1 (A → C), Cd 16(−C), and Cd 5 (−CT) were detected. To our knowledge, our study is the first to characterize β-thalassemia mutations in the Kachhiya Patel community. This study will facilitate molecular analysis and prenatal diagnosis in these four communities.

Viral infections in newborns through exchange transfusion

Preprocedure sera of thirty one neonates requiring exchange transfusion were tested for serological markers of HBV, HCV, CMV, HIV and LFT. All the babies were investigated for these parameters one week and two months after transfusion to evaluate the risk of transmission of viral infection. Serological markers for these viral infections were also studied in the mothers and donors’ blood to establish the route of infection. Donors’ blood used for transfusion was pretested for HBsAg, VDRL and anti-HIV.HBsAg was detected one week post exchange in one baby and two months post exchange in two babies. Exchange transfusion was implicated in two of them, where one donor had HBsAg and the other anti-HBc. Vertical transmission accounted for the remaining one. Out of these HbsAg positive cases, one showed evidence of recently acquired CMV infection. Vertical transmission of anti-HCV was observed in one case. None of the neonates, mothers and donors were positive for anti-HIV.In view of probable serious consequences of HBV and HCV infections, blood used for exchange transfusion ought to be screened for anti-HBc and anti-HCV, besides routine HBsAg, VDRL and anti-HIV screening.

Quantitative variations in A, B, Ht1 and Ht2 antigens on erythrocytes in leukemia and lymphoma

Using an immunoradiometric assay, A, B, Ht1 and Ht2 antigens on erythrocyte membranes were quantitated in 192 leukemia and 23 lymphoma patients. A and B antigens were reduced in 37% and 50% of leukemia cases respectively. A wide range of antigenic reduction was observed for A antigen, while B antigenic reduction was generally 16-20%. Passive absorption of Ht1 antigen from plasma onto erythrocytes was greater in leukemic compared to normal subjects. Among patients with reduced A and B antigens, 27% had increased and 37% decreased Ht2 antigens. Incidence of reduction in antigens was significantly higher (p less than 0.001) in untreated compared to treated patients.

Role of antibody dependent cell mediated cytotoxicity in ABO hemolytic disease of the newborn

Study includes fifty O blood group mothers delivering A or B group infants suffering from jaundice and a control group consisting of thirty one O group mothers of nonjaundiced infants. Lytic ability of maternal IgG anti-A/anti-B was determined by51Cr ADCC assay in which cord blood monocytes were used as effector cells. In control series mean% specific lysis (SL) was 18±3.1 for IgG anti-A and 17.9±3.1 for IgG anti-B. in jaundiced series IgG anti-A was more lytic than IgG anti-B. However, the increase in ADCC lysis was statistically insignificant. Eventhough> 1: 32 titre was more often associated with >35% SL, in general the immune A/B antibody titre showed poor correlation with ADCC lysis. Majority of the severe ABO-HDN cases had >35% ADCC lysis.

Effect of gamma irradiation on cytokines released by platelets during storage

Abstract Most published gamma irradiation studies are on the cytokines secreted by leucocytes than platelet cytokines. If cytokines secretion is suppressed by irradiation then it will be an important application of irradiation. To know this we planned to measure cytokines like Regulated upon activation normal T cells expressed and secreted (RANTES), platelet factor 4 (PF-4), Transforming growth factor βeta1 (TGF-β) and βeta Thromboglobulin (β TG) during the storage of irradiated (IR) and non-irradiated (NI) platelets. Ten platelet concentrate (PCs) were prepared using platelet rich plasma and buffy coat methods each and 10 by apheresis on Trima Cell separator. Each PC was transferred in a transfer bag and IR at about 25 Gy and stored at 22 °C in platelet agitator. Samples were taken from NI and IR bag each on 0 day, 3rd day and 5th day from supernatant for cytokine analysis. The samples were stored below −50 °C for cytokines assays using commercial ELISA kits. RANTES levels were in the range of 12–400 ng/ml on 0 day and increased to 108–800 ng/ml on 5th day in NI platelets and 104–800 ng/ml in IR platelet. PF4 increased from 0 day to 5th day showing levels 300–1500 ng/ml in both types of PC. β-TG range was 748–5258 ng/ml in NI and 878–4638 ng/ml in IR platelets on 5th day. TGF-β1 increased up to 780–38431 pg/ml in NI and 461–50,000 pg/ml in IR PC. The study showed that comparison of cytokine levels during the storage in NI and IR platelets was not significant by Mann–Whitney U- test (p > 0.05). Significant increase was observed in these cytokine levels from 0 day to 3rd and 5th day in NI as well as IR samples by Wilcoxon Signed- Rank test (p < 0.05).

Erythrocyte membrane enzyme acetyl cholinesterase in neonatal jaundice

Erythrocyte membrane enzyme “acetyl cholinesterase” (AChE) was estimated in 200 cases of neonatal jaundice. The enzyme levels were reduced in jaundiced infants in comparison to normal infants (mean 34.5±19.8 units vs 48.8±25.2 units/gm Hb, P<0.05). The levels of the enzyme were not greatly altered in neonatal jaundice due to Rh incompatibility, G-6-PD deficiency and prematurity. Lower levels were observed in cases of ABO hemolytic disease in relation to ABO compatible unexplained cases of neonatal jaundice (27.9±13.4 units vs 37.2±21.2 units/gm Hb, P<0.05). However these findings were not of anv diagnostic value, as low levels of AChE were not associated with spherocytosis and increased osmotic fragility.