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  Table of Contents    
CASE REPORT  
Year : 2020  |  Volume : 63  |  Issue : 3  |  Page : 481-484
Case series of HbQ-India, a rare alpha globin variant in a referral laboratory setting in South India


1 Department of Hematopathology, AmPath, Hyderabad, Telangana, India
2 Department of Transfusion Medicine, Citizens Hospitals, Hyderabad, Telangana, India
3 Department of Cytogenetics, AmPath, Hyderabad, Telangana, India

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Date of Submission14-Jun-2019
Date of Decision08-Oct-2019
Date of Acceptance21-Oct-2019
Date of Web Publication7-Aug-2020
 

   Abstract 


HbQ variants are rare alpha globin chain variants commonly found in Sindhi community. It results from a point mutation of α-1 globin gene at position 223 of the coding region of exon 64. It is inherited in an autosomal dominant fashion. HbQ-India is usually clinically silent in heterozygous state unless associated with other conditions like beta thalassemia, alpha thalassemia, HbE disease, or nutritional anemia. High performance liquid chromatography (HPLC) identifies HbQ-India with a prominent peak present just after the Sickle window. We present five cases of HbQ-India from a retrospective analysis of 6034 cases over a period of 3 years, a rarity in a referral setting of South India. Awareness of this entity is important for appropriate recognition to prevent clinically symptomatic hemoglobinopathies. This study also highlights the retention time (RT) and characteristic chromatographic HPLC pattern seen in HbQ-India.

Keywords: Capillary electrophoresis, HbQ India, hemoglobinopathies, HPLC

How to cite this article:
Shaik A, Thekkelakayil ST, Kumawat V, Gupta A, Goyal M. Case series of HbQ-India, a rare alpha globin variant in a referral laboratory setting in South India. Indian J Pathol Microbiol 2020;63:481-4

How to cite this URL:
Shaik A, Thekkelakayil ST, Kumawat V, Gupta A, Goyal M. Case series of HbQ-India, a rare alpha globin variant in a referral laboratory setting in South India. Indian J Pathol Microbiol [serial online] 2020 [cited 2020 Oct 30];63:481-4. Available from: https://www.ijpmonline.org/text.asp?2020/63/3/481/291673





   Introduction Top


Hemoglobin variants arise from mutations in amino acid sequence affecting either α or β globin chains. More than 900 hemoglobin variants have been described of which a quarter are alpha chain variants.[1] Hemoglobin Q (HbQ) is a very rare alpha globin chain variant. Three molecular subtypes of HbQ have been documented, namely HbQ-India, HbQ-Thailand, and HbQ-Iran. These are characterized by distinct point mutations in the α globin chain – HbQ-India (alpha64 Asp to His), HbQ-Thailand (alpha74 Asp to His), and HbQ-Iran (alpha75 Asp to His).[2] HbQ-India was first reported by Sukumaran in 1972 in a Sindhi family in association with beta thalassemia.[3] HbQ-India is inherited in an autosomal dominant fashion, resulting from a point mutation of α-1 globin gene at position 223 of the coding region of exon 64.[2] On agar gel electrophoresis at alkaline pH, the HbQ presents as a band in the position of HbS/D/G, which can commonly be misinterpreted as HbS or HbD. High performance liquid chromatography (HPLC) shows a peak in unknown window with a retention time (RT) of 4.45 ± 0.02 minutes on Bio-Rad D10 and 4.70–4.90 minutes on Bio-Rad variant II.[4]


   Materials and Methods Top


The study was a retrospective observational study conducted in a clinical referral laboratory over a period of 3 years. Complete blood counts were obtained on EDTA anticoagulated whole blood using Coulter LH750 Hematology Analyzer (Beckman Coulter, Fullerton, CA, USA). The data was reviewed by a hematopathologist along with Leishman-stained peripheral blood smear, reticulocyte count, and sickling test.

A total of 6034 cases were screened for beta thalassemias and other hemoglobin variants using BIO-RAD D-10 dual mode instrument (Bio-Rad Laboratories, California, USA) from July 2015 to June 2018, which utilizes the principle of cation-exchange HPLC. Quality control was ensured by running two level controls with every batch. HbA2 cut-off for diagnosing beta Thalassemia trait was 3.8% in our laboratory, based on the verification of the company defined protocols. Any case with borderline HbA2 with normal indices or HbA2 of more than 3.2% with lower red cell indices were recommended for molecular testing. Cases which showed a peak in unknown window after the Sickle window (RT – 4.02-4.30 minutes) after due consideration of red cell indices and clinical profile were diagnosed as HbQ. The HPLC data including the percentage of HbQ-India, it's retention time, and the pattern of the chromatogram were reviewed. Capillary electrophoresis was done in three cases and confirmed HbQ-India. The demographic profile and the indication for testing were recorded for each case, wherever available.


   Results Top


[Table 1] shows demography, clinical indications, RBC indices, and HPLC chromatographic details of the patients. Two patients were infants and other three were adults. Four patients showed altered red cell indices–three had microcytic hypochromic anemia and one had low red cell indices with normal hemoglobin levels and marked erythrocytosis. One case that showed marked erythrocytosis had no history of smoking, alcoholism, or other obvious secondary causes. No further testing was done to evaluate for the underlying cause. Sickling test was negative in all cases.
Table 1: Shows demographic details, indications, red cell indices and chromatographic details of patients with HbQ India

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In all cases HPLC showed a peak in an unknown window after the Sickle window (RT- 4.30-4.44 minutes) with reduced HbA0 and normal HbF levels. Four were diagnosed as heterozygous HbQ-India with normal HbA2 levels with a note for both infants to follow up at three years for definite quantitation of HbA2 and HbQ-India. One showed associated increased HbA2 levels (4.9%) and was diagnosed double heterozygous HbQ/Beta thalassemia trait (BTT). Parental samples of these infants or related members of other patients were not available for further mapping. HPLC chromatogram showed four peaks in all the cases [Figure 1]. These were composed of the main peak (HbQ-India), two peaks just preceding and one immediately following the main peak. Capillary electrophoresis testing performed in three cases showed a peak in the HbD zone and an additional small peak in Z1 zone, consistent with that of presence of HbQ-India [Figure 2]. Three cases belonged to Sindhi community, while ethnicity in two could not be established. Two patients had migrated and settled in the southern part of the country.
Figure 1: HPLC chromatograph of a case of HbQ-India showing a peak in an unknown area with a Retention Time of 4.38 minutes. There are three other peaks associated with the main peak, two just preceding and the one just proceeding

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Figure 2: Capillary electrophoresis of the same case showing main peak in the D-zone and one minor peak in the Z1 zone

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   Discussion Top


Hemoglobinopathies constitute the most common genetic diseases in the world. The spectrum of inherited abnormalities in hemoglobin synthesis ranges from thalassemia to structurally defective hemoglobin variants. HbQ-India is a rare alpha globin chain variant occurring due to point mutations in alpha gene. The prevalence of HbQ-India is 0.4% in India and is found to be lower than HbD-Punjab, HbE, and HbS in Indian population.[5] Over a period of 3 years and we retrospectively analyzed 6034 cases, of which 5 cases of HbQ-India were identified giving overall frequency of 0.08%. Aradhana et al. screened 7530 patients and found the HbQ-India prevalence to be 0.4%, whereas Panigrahi et al. also found the same prevalence in a study of 4500 cases.[6],[7] The largest series studied is by Phanasgaonkar et al. in a referral center, who reported 64 cases of HbQ-India over a period of two decades of which 36 were of HbQ trait, 22 of HbQ-India-BTT, 3 cases of HbQ-India-beta thalassemia major, and 3 cases of HbQ-India homozygous.[1] The frequency in our study was relatively low possibly due to geographical location, difference in referral base of patients, and indications of testing at our center as compared to other specialized centers. Most of the cases screened at our center were asymptomatic. The catchment area for our laboratory was southern and eastern India and lesser from northern and western India, where HbQ-India predominantly exists.

Most often it presents either in the heterozygous state or in association with beta-thalassemia. Normally HbQ-India is clinically silent. This can be explained by replacement of aspartic acid with histidine, which is on the surface of the protein structure and does not affect the protein interchain contacts and electrical charges of the molecule; therefore, it does not cause any changes in hematologic parameters and indices.[8] It becomes symptomatic when it is present in association with other conditions like beta-thalassemia, alpha-thalassemia, HbE, HbH, and nutritional anemia. The degree of erythrocytosis in one case was unusually high as compared to other studies. Secondary causes for the erythrocytosis were excluded; however, the exact cause could not be established and it is desirable to rule out high affinity nature of HbQ although none of the literature describes so.

HbQ percentage in our study ranged from 6.7% to 17.8%. Presence of alpha-thalassemia favors the formation of HbQ, whereas beta-thalassemia reduces the formation of HbQ.[9],[10] In our study a patient with beta-thalassemia showed HbQ levels of 6.7% which was lowest among the five cases. Aradhana et al. showed that levels of HbQ ranged from 13.6 to 24.4% in HbQ-India trait, 7.4 to 9.0% in patients with HbQ-India-BTT, and 32 to 35% in HbQ homozygous.[6] Phanasgaonkar et al. reported HbQ levels of less than 10% (7.2–11.3%) in patients with associated beta–-thalassemia and 13.1–23.5% in heterozygous and 29–36% in homozygous HbQ disease.[1]

There were multiple additional minor peaks seen in the chromatograms. Sharma et al. suggested that these represent post-translation modification of HbQ-India, glycosylation or carry over in case of minor HbS peaks in their patients.[11] Our presumption is that the peak immediately succeeding the HbQ possibly represents αQαββ and peaks preceding HbQ represent post-translational changes. Additional studies have to be done to decipher the exact nature of these peaks. Our study was limited by lack of definitive diagnostic procedures like isoelectric focusing and molecular studies, as these techniques are expensive and not easily available due to rarity of the disorder.


   Conclusion Top


HbQ-India is a very rare alpha chain variant with a very low frequency, usually seen in heterozygous state. Awareness of this entity is important for appropriate recognition to prevent clinically symptomatic hemoglobinopathies. There is more work needed to decipher the exact biochemical and molecular characteristics of other peaks associated with HbQ-India.

Acknowledgements

We acknowledge the efforts of our Technical staff - Mr. Suresh, Mr. Srinivas, Mr. G K Naga Siva Kumar and Mr. Siva Prasad, who contributed in processing of the samples and documentation of the results.

We are also thankful and acknowledge the support of Dr. Asma Niloufer, Consultant Biochemist, Apollo Hospitals, Jubilee Hills, Hyderabad, India for providing support for processing of samples by capillary electrophoresis.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
   References Top

1.
Phanasgaonkar S, Colah R, Ghosh K, Mohanty D, Gupte S. HbQ (India) and its interaction with beta-thalassemia: A study of 64 cases from India. Br J Biomed Sci 2007;64:160-3.  Back to cited text no. 1
    
2.
Abraham R, Thomas M, Britt R, Fisher C, Old J. HbQ-India: An uncommon variant diagnosed in three Punjabi patients with diabetes is identified by a novel DNA analysis test. J Clin Pathol 2003;56:296-9.  Back to cited text no. 2
    
3.
Sukumaran PK, Merchant SM, Desai MP. Hemoglobin Q India (alpha 64 (E13) aspartic acid to histidine) associated with beta thalassemia observed in three Sindhi families. J Med Genet 1972; 9:436-42.  Back to cited text no. 3
    
4.
Joutovsky A, Hadzi-Nesic J, Nardi MA. HPLC Retention time as a diagnostic tool for hemoglobin variants and hemoglobinopathies: A study of 60000 samples in a clinical diagnostic laboratory. Clin Chem 2004;50:1736-47.  Back to cited text no. 4
    
5.
Higgs DR, Thein SL, Wood WG. Distribution and population genetics of the thalassemia's. In: Weatherall DJ, Clegg JB, editors. The Thalassaemia Syndromes. 4th ed. Oxford, England: Blackwell Scientific Publications; 2001. p. 237-84.  Back to cited text no. 5
    
6.
Harrison A, Mashon RS, Kakkar N, Das S. Clinico-hematological profile of Hb Q India: An uncommon hemoglobin variant. Indian J Hematol Blood Transfus 2018;34:299-303.  Back to cited text no. 6
    
7.
Panigrahi I, Bajaj J, Chatterjee T, Sakena R, Mahapatra M, Pati HP. Hb Q India: Is it always benign? Am Hematol 2005;78:245-7.  Back to cited text no. 7
    
8.
Lorkin PA, Charlesworth D, Lehmann H, Rahbar S, Tuchinda S, Eng LI. Two haemoglobins Q, alpha alpha-73(EF3) and alpha-75 (EF4) aspartic acid to histidine. Br J Haematol 1970;19:117-25.  Back to cited text no. 8
    
9.
Qin WB, Baysal E, Wong KF, Molchanova TP, Pobedimskaya DD, Sharma S, et al. Quantities of alpha Q chain variants in heterozygotes with and without a concomitant beta trait. Am J Hematol 1994;45:91-3.  Back to cited text no. 9
    
10.
Felice AE, Webber BB, Huisman TH. Alpha-thalassemia and the production of different alpha chain variants in heterozygotes. Biochem Genet 1981;19:487-98.  Back to cited text no. 10
    
11.
Sharma S, Sharma P, Das R, Chhabra S, Hira JK. HbQ-India (HBA1:c. 193G>C): Hematological profiles and unique CE-HPLC findings of potential diagnostic utility in 65 cases. Ann Hematol 2017;96:1227-9.  Back to cited text no. 11
    

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Correspondence Address:
Manu Goyal
Department of Hematopathology, AmPath, Serilingampally, Nallagandla, Hyderabad - 500 019, Telangana
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/IJPM.IJPM_465_19

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