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Year : 2016  |  Volume : 59  |  Issue : 1  |  Page : 102-103
The pursuit of rare hemoglobins

Department of Hematology, Postgraduate Institute of Medical Education and Research, Chandigarh, India

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Date of Web Publication9-Mar-2016

How to cite this article:
Sharma P, Das R. The pursuit of rare hemoglobins. Indian J Pathol Microbiol 2016;59:102-3

How to cite this URL:
Sharma P, Das R. The pursuit of rare hemoglobins. Indian J Pathol Microbiol [serial online] 2016 [cited 2020 Nov 28];59:102-3. Available from: https://www.ijpmonline.org/text.asp?2016/59/1/102/178223

In this issue of the IJPM, Sharma, et al. report the case of a 10-year-old boy who presented with anemia and mild splenomegaly and in whom hemoglobin high-performance liquid chromatography (Hb HPLC) revealed a peak in the HbC window. [ 1] The case, which was finally diagnosed after family studies and direct gene sequencing as the rare asymptomatic Hb Agenogi, is both informative and intriguing at several levels.

The technology behind hemoglobinopathy screening has progressively evolved over the last few decades. The labor-intensive cellulose acetate or agarose gel electrophoretic techniques are no longer tenable as the first-line screening techniques in busy departments, although they remain invaluable for the confirmation of variants detected by other tests. [2] Cation-exchange HPLC has been the technique of choice for over 10 years due to its attributes of being accurate, relatively high-throughput, and amenable to complete automation. HPLC resolves most cases, with only a few issues, like borderline HbA2 values and potential interference by iron deficiency and α-thalassemia. [3],[4]

A reality of HPLC-based diagnosis in India has been the dominance of one instrument manufacturer in the market, which is not an ideal situation for both innovation and pricing. Capillary electrophoresis (CE), which has recently gained prominence, is a high-throughput (up to 38 samples per hour in high-end instruments) and price competitive (at least two different companies sell CE instruments) alternative. CE picks up variants that may be missed by HPLC and separates HbE from HbA2. [5] It can be used to measure glycated hemoglobin and perform serum protein electrophoresis. [6],[7] CEs use in hemoglobinopathy diagnosis is growing, the only major hurdle in its routine application seemingly being the learning curve involved in using this new technology.

Hb Agenogi mimics HbC on both HPLC and alkaline pH electrophoresis. HbC is extremely rare in India, to the point of nonexistence. [8] In fact, a variant Hb peak eluting in the HbC window in an Indian will finally, most likely, not turn out to be HbC. [9] It is interesting to speculate as to why HbS, which is also common in African populations like HbC, traveled across continents to reach India in the Arab-Indian haplotype but HbC did not. Plausible reasons, which are still unraveling, include variable chromosomal recombination and gene conversion events at various time points (younger mutations being more geographically localized), human migrations (HbC has peak frequencies in Western Africa and seems to have diffused more Northward and Westward, rather than towards Asia in the East), geospatial isolation (the river Niger sharply limits the zone of HbE toward its East bank) and selection pressures from malaria as well as HbS (thalassemia and other hemoglobinopathies being rare wherever very high frequencies of βS are found in the Central and East Africa). [10]

The origin of the Agenogi mutation in this Indian family remains a mystery. Sporadic asymptomatic hemoglobinopathies arising from de novo mutations are not unusual. These individuals are often detected incidentally during thalassemia screening programs. It must be emphasized that the next logical step for such abnormal peaks found by HPLC screening is Hb electrophoresis at alkaline pH [2] to narrow down the differential diagnosis and spousal Hb HPLC if clinically pertinent. This should be followed, if required, by DNA sequencing of the β/α-globin genes to characterize the mutation as was done in the reported case.

Moreover, on a final note, this particular report illustrates the importance of inter-institutional cooperation, and the value of seeking specialist opinions when faced with diagnostically challenging patients. Despite the global explosion in sequencing technologies, it will take time before all large institutions acquire CE-based sequencers. Till then, such collaborations will remain necessary to finally resolve similar structural rarities.

   References Top

Sharma S, Sharma G, Chandra J, Colah R. Hemoglobin Agenogi - A rare abnormal beta globin chain variant. Indian J Pathol Microbiol 2016;59:99-101.  Back to cited text no. 1
  Medknow Journal  
Sreedharanunni S, Sharma P, Murgai P, Chhabra S, Das R. Automated alkaline-pH electrophoresis followed by densitometry does not correlate with cation-exchange (CE)-HPLC in quantification of HbA2 and variant hemoglobins. Clin Chem Lab Med 2015;53:e301-3.  Back to cited text no. 2
Rangan A, Sharma P, Dadu T, Saxena R, Verma IC, Bhargava M. ß-Thalassemia mutations in subjects with borderline HbA2 values: A pilot study in North India. Clin Chem Lab Med 2011;49:2069-72.  Back to cited text no. 3
Sharma P, Das R, Trehan A, Bansal D, Chhabra S, Kaur J, et al. Impact of iron deficiency on hemoglobin A2% in obligate ß-thalassemia heterozygotes. Int J Lab Hematol 2015;37:105-11.  Back to cited text no. 4
Keren DF, Hedstrom D, Gulbranson R, Ou CN, Bak R. Comparison of Sebia Capillarys capillary electrophoresis with the Primus high-pressure liquid chromatography in the evaluation of hemoglobinopathies. Am J Clin Pathol 2008;130:824-31.  Back to cited text no. 5
Urrechaga E. High-resolution HbA(1c) separation and hemoglobinopathy detection with capillary electrophoresis. Am J Clin Pathol 2012;138:448-56.  Back to cited text no. 6
Gay-Bellile C, Bengoufa D, Houze P, Le Carrer D, Benlakehal M, Bousquet B, et al. Automated multicapillary electrophoresis for analysis of human serum proteins. Clin Chem 2003;49:1909-15.  Back to cited text no. 7
Piel FB, Howes RE, Patil AP, Nyangiri OA, Gething PW, Bhatt S, et al. The distribution of haemoglobin C and its prevalence in newborns in Africa. Sci Rep 2013;3:1671.  Back to cited text no. 8
Badyal RK, Chhabra S, Sharma P, Das R. An intriguing high performance liquid chromatogram of a double heterozygosity for Hb Q-India/Hb D-Punjab. Hemoglobin 2014;38:440-3.  Back to cited text no. 9
Livingstone FB. Who gave whom hemoglobin S: The use of restriction site haplotype variation for the interpretation of the evolution of the β-globin gene. Am J Hum Biol 1989;1:289-302.  Back to cited text no. 10

Correspondence Address:
Prashant Sharma
Department of Hematology, Level 5, Research Block A, Postgraduate Institute of Medical Education and Research, Sector 12, Chandigarh - 160 012
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/0377-4929.178223

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