5 IN T E R N A L m E d Ic IN E ISSN 2413-6077. IJmmR 2022 Vol. 8 Issue 2 doI 10.11603/ijmmr.2413-6077.2022.2.13152 SICKLE CELL THALASSEMIA IN AN ADULT FROM CENTRAL INDIA SHOWING MASSIVE SPLENIC INFARCTION AND GAMNA-GANDY BODIES IN SPLENIC PARENCHYMA WITH CONCOMITANT PLASMODIUM FALCIPARUM INFECTION (case report) *T. S. Singh1, R. Singh2, R. Pandey3 1 – 155 BASE HOSPITAL, TEZPUR, INDIA 2 – INDIAN NAVAL HOSPITAL SHIP ASVINI, MUMBAI, INDIA 3 – COMMAND HOSPITAL, PUNE, INDIA Background. Sickle cell thalassemia is a heterozygous state of HbS/β+ or HbS/β0 manifested clinically either as an asymptomatic carrier or have features akin to sickle cell anemia. Objective. The aim of the study is to review the literature and discuss the varied clinical manifestations and diagnosis of a case report of haemoglobinopathy in an adult from Central India. Methods. A case of haemoglobinopathy from central part of India is being investigated. Results. A case of haemoglobinopathy in an adult presenting to a tertiary hospital with chronic back ache was reported. The patient was found to have massive splenomegaly with evidence of splenic infarction. Gross examination of spleen revealed multiple soft yellowish pasty areas, which on microscopic examination showed significant necrosis with presence of great amount pale amorphous yellow substance, Gamna-Gandy bodies (GGBs) and massive foreign body cell reaction in splenic parenchyma. Post-splenectomy peripheral blood smear (PBS) showed thrombocytosis and plasmodium falciparum gametocytes. The Hb electrophoresis revealed both elevation of Hb S (49.7%), Hb F (46.7%); Hb A2 (3.0%) and (Hb A 0.7%) consistent with as Sickle Cell Thalassemia Hb S/β+ Thalassemia. Conclusion. Sickle cell thalassemia with long standing huge splenomegaly, splenic infarction with GGBs deposition and concomitant falciparum malaria in clinically stable patient is rare indeed. KEYWORDS: sickle-cell anemia; beta-thalassemia; malaria; plasmodium falciparum; Gamna-Gandy bodies. *Corresponding author: Thongam Sachin Singh, Assistant professor (Pathology), 155 Base Hospital, Tezpur, 784001, India. E-mail: sachinthongam@gmail.com International Journal of Medicine and Medical Research 2022, Volume 8, Issue 2, p. 5-10 copyright © 2022, TNMU, All Rights Reserved T. S. Singh et al. Introduction Hemoglobinopathies are the most common inherited RBC disorders, thalassemia and sickle cell disease – the most frequent. Co-existent both sickle cell mutation and thalassemia mu- tation, commonly beta thalassemia, facilitating development to a compound heterozygous state known as sickle cell beta thalassemia, was first reported and described by Silvestroni and Bianco in 1944 [1]. The incidence of this con- dition is very common in Mediterranean region. Its prevalence in Indian subcontinent is under- reported, perhaps due to underdiagnosis as most of these patients are normal and asymp- tomatic. Documented prevalence of sickle cell beta thalassemia in India is less than 1% in various studies [2]. Inherited as autosomal recessive disorder it occurs when one abnormal gene for produc- tion of hemoglobin S is inherited from one parent and one abnormal gene for production of beta thalassemia is inherited from the other parent. The genes for both hemoglobin and beta thalassemia are both located on chro- mosome 11. Manifestation of the condition was studied in 1973 by Serjeant at el [3]. Clinical mani- festations vary depending upon heterozygosity from being completely asymptomatic (Hb S/ β+) to signs and symptoms very much akin to sickle cell anemia (Hb S/ β0) [4]. Case Report A 29-year-old serving soldier, the resident of central India, while being employed in high altitude area complained of chronic pain abdo- men, which was found to be due to massive splenomegaly (23 cm in span) with sonographic evidence of splenic infarction. Approximately 7 years earlier, he had similar episode of abdo- men pain for which he was worked up to rule 6 IN T E R N A L m E d Ic IN E ISSN 2413-6077. IJmmR 2022 Vol. 8 Issue 2 any hematological disorder. He had microcytic hypochromic anemia with normal TLC and DLC. Thrombocytosis was noted. There was no documented evidence of any organomegaly in the past. Past Hb electrophoresis reports revealed HbS – 49.7%, Hb A – 0.7%, Hb A2 – 3.0% and Hb F – 46.7%; consistent with as Sickle Cell Thalassemia (Hb S/β+ Thalassemia). His general, systemic examinations were within norm during pre-anesthetic check-up. He was taken up for the splenectomy operation and specimen sent to Dept of Pathology for histopathological examination. Lab received a grossly enlarged specimen of splenectomy weighing 1.2 kg and having apparently normal architecture externally. Serial cut section of the fresh specimen showed multiple soft yellowish areas of splenic infarction. Formalin-soaked cotton were placed in between these cut surfaces and whole specimen was put in 10 % formalin for 24 hours. Multiple sections from these necrosed soft areas were submitted along with normal appearing areas of spleen. H&E stains slides were studied by microscopy. Histopathological examination revealed significant necrosis in infarcted area and there were multiple areas showing deposition of yellowish granular substances in splenic tissue which were Congo-red negative and biref rin- gence negative under polarized light. Sickle cells in the sinusoids were also seen. Post splenectomy period was uneventful with no fever, however, there was incidental findings of different forms of plasmodium fal­ ciparum including gametocytes in the peripheral blood. Thrombocytosis was also noted as expected. Discussion Sickle cell thalassemia, described originally as micro-drepanocytic disease by Silvertroni and Bianco [1], is a clinical condition where mutations in both HbS gene and Thalssemia gene (commonly beta gene) occur resulting in two heterozygous states of either Hb S/ β+ or Hb S/ β0. Heterogenicity in the beta thalassemia mutations leads to different beta globin syn- thesis and hence different amounts of HbA are synthesized which result in variable clinical manifestations ranging from nearly asympto- matic to a severe condition similar to sickle cell anemia (homozygous HbS) [4]. Laboratory investigations reveal microcytic hypochromic red cells with target cells and occasionally sickled forms. Hemoglobin elec t- rophoresis may reveal 60-90% Hb S, 0-30% Hb A, 1–20% Hb F and an increased HbA2 level. In Hb S/β+Thalassemia, as in our case, variable amounts of Hb A dilute Hb S and consequently inhibit polymerization-induced cellular damage. Increased levels of Hb A are usually associated with a milder phenotype [2, 5, 6, 7]. In our case, in the pre-splenectomy period, the patient was consistently having microcytic anemia as shown in Table 1 with leukopenia and increased LDH indirectly suggesting he- molysis. Platelet was adequate. No malarial parasite was seen in the peripheral smear. Post splenectomy, anemia improved but mild leuko- cytosis and thrombocytosis was evidenced as expected. LDH was found decreasing. Inci- dentally, peripheral smear showed numerous gametocytes of Plasmodium falciparum. Mo- lecular tests for confirmation of sickle cell thalassemia were unavailable in the lab. Hence diagnosis was purely based on tests for sickling and Hb electrophoresis. Role of PBS examination and relevant histopathological sections could not be undermined. Individually, both thalassemia and sickle cell trait offer protection from malarial infection, however when both occurs in the same indi- vidual protection offered by thalassemia can be reversed by another additional genetic polymorphism such as HbS mutation. The two mutations essentially cancel each other and the individual becomes susceptible to malaria [8]. Heterozygotes for the sickle gene (Hb genotype AS) are relatively protected against death due Table 1. Relevant present investigations Sr No. Pre splenectomy Post splenectomy Hb (gm/dL) 6.6 10.1 MCV (fL) 68 69 TLC 2450 11220 DLC P-69%, L-23%, M-07%, E-01% P-82%, L-11%, M-04%, E-03% Platelets (105/μL) 1.1 5.56 LDH (U/L) >1000 634 PBS No malarial parasites Malarial parasites evidenced T. S. Singh et al. 7 IN T E R N A L m E d Ic IN E ISSN 2413-6077. IJmmR 2022 Vol. 8 Issue 2 Fig. 1. Gross specimen before and after fixation showing areas of infarction. Fig. 2. Photomicrographs from H&E sections showing deposition of yellowish fine granular to amorphous (inset) substances in spleen (Gamna-Gandy bodies) and sickle cells. Fig. 3. Photomicrographs showing numerous gametocytes and trophozoites of Plasmodium falciparum and thrombocytosis. Fig. 4. Photomicrographs showing positive sickling with malarial gametocytes. T. S. Singh et al. 8 IN T E R N A L m E d Ic IN E ISSN 2413-6077. IJmmR 2022 Vol. 8 Issue 2 to malaria, probably through accelerated clearance by macrophages of Plasmodium fal- ciparum-infected erythrocytes and by inhibiting growth of parasites in RBCs by increasing intracellular potassium, reduced PH and increasing endothelial adherence of infected cells [9]. On the other hand, homozygous (Hb SS) sickle cell anemia patients are not protected from malaria or any other infections [10,11] and, hence, there is increased chance of dying from malaria which is additionally aggravated by having dysfunctional spleen [12,13] or in the post-splenectomy period as removal of ab nor- mal RBCs and other intracytoplasmic inclusion bodies is the primary function of spleen [14,15]. In our case, the pre-splenectomy period blood did not show any malarial parasites due to effective hyperactive enlarged spleen but once patient’s spleen was removed, numerous forms of plasmodium falciparum were seen in the blood as the individual was devoid of major protection offered by spleen including removal of infected RBCs and other inclusion bodies. People living in the malaria endemic areas have some degree of acquired immunity against malaria which may weaken over ab- sence of recurrent exposures or when indivi- duals moved to non-endemic areas [16]. This partial acquired immunity against malaria develops during the first 5 years of life and is dependent on the intensity of transmission, on exposure frequency and so it decreases with time if re-exposure does not occur [17]. In sickle cell disease, auto-splenectomy or functional asplenia occurs in early adulthood in homo- zygous state, however, in sickle beta-thalassemia the spleen may persist and remain enlarged even in adults. This splenomegaly may cause massive pooling of blood in the spleen (splenic sequestration), which leads to its enlargement and resulting in severe anemia and hence a more compelling indication for splenectomy in sickle beta-thalassemia. This reduces substan- tially the rate of blood trans fusion [18]. Also, multiple areas in splenic parenchyma with deposition of yellow­brownish fine gra­ nular to amorphous substance with associated foreign-body giant cell reaction were found [19]. They were Gamna-Gandy bodies. Initially described by Marini as siderotic nodules in 1904, French physician Charles Gandy in 1907 found it in biliary cirrhosis patient. In 1921, the etiology was better described by an Italian pathologist Carlos Gamna, who observed de- posits of amorphous material composed of iron and calcium sulfate in the spleen of a patient, who died of chronic hemolytic anemia; it was named Splenogranulomatosisiderotica. The name “Gamna-Gandy bodies” has been widely used ever since [20]. It is formed in sickle cell disease (SCD) due to chronic episodes of vaso- occlusion and hemolysis in the central arteriole of the white pulp with periarteriolar hemorrha- ges followed by mineral elements of the blood. It can be seen as either fine granular deposits in red pulp or as perivascular deposits or sub- capsular deposits. GGBs can vary in size, ranging from 10 to 49 microns [21]. The GGDs is not pathognomonic of sickle cell disease as it can be found in many other diseases and in several other organs. Conclusion Splenomegaly in a case of sickle cell thalas- semia is an established association, however to such extent that infarction with deposition of Gamna-Gandy bodies deposition is rare indeed. Loss of protective mechanism due splenectomy also predispose the individual to manifest subclinical malarial infection. Our case highlighted patient’s high endurance and acquired immunity against all the conditions mentioned by virtue of his birth with inherited mutations in an endemic area of malaria. Diagnosis in the era of molecular tests still count on Hb electrophoresis, sickling test and peripheral blood smear. Conflict of Interests Authors declare no conflict of interest. Acknowledgements Sincere thanks to all the staff of the De- partment of Pathology, 155 base Hospital, Tezpur, India Fig. 5. Hb electrophoresis at alkaline pH showing a prominent band in SDG region, Hb F and faint band in HbA2 (Hb S – 47.7%, Hb F – 30.7%, Hb A – 18.5 % and Hb A2 – 3.1 %). T. S. Singh et al. 9 IN T E R N A L m E d Ic IN E ISSN 2413-6077. IJmmR 2022 Vol. 8 Issue 2 Author’s Contributions Thongam Sachin Singh – conceptualization, methodology, formal analysis, writing – original draft, writing – re viewing and editing; Raj Singh – data curation, writing – reviewing and editing; Rahul Pandey – investigation, formal analysis. СЕРПОВИДНО-КЛІТИННА ТАЛАСЕМІЯ З МАСИВНИМ ІНФАРКТОМ СЕЛЕЗІНКИ ТА ВУЗЛАМИ ГАМНА-ГАНДІ В ПАРЕНХІМІ СЕЛЕЗІНКИ З СУПУТНЬОЮ МАЛЯРІЙНОЮ ІНФЕКЦІЄЮ У ДОРОСЛОГО З ЦЕНТРАЛЬНОЇ ІНДІЇ (клінічний випадок) T. S. Singh1, R. Singh2, R. Pandey3 1 – 155 BASE HOSPITAL, TEZPUR, INDIA 2 – INDIAN NAVAL HOSPITAL SHIP ASVINI, MUMBAI, INDIA 3 – COMMAND HOSPITAL, PUNE, INDIA Вступ. Серпоподібноклітинна анемія – це спадкова гемоглобінопатія, яка характеризується гетерозиготним станом Hb S/β+ або Hb S/β0 і клінічно може не проявлятися у гетерозиготних носіїв Мета. Метою дослідження є огляд літератури та обговорення різноманітних клінічних проявів і діагностики клінічного випадку гемоглобінопатії у пацієнта із Центральної Індії. Методи. Описано випадок гемоглобінопатії у пацієнта із Центральної Індії. Результати. Ми повідомляємо про випадок гемоглобінопатії у дорослого, який звернувся до лікарні третього рівня з хронічним болем у спині. Було виявлено масивну спленомегалію з ознаками інфаркту селезінки. При загальному дослідженні селезінки виявлені множинні м’які жовтуваті тістоподібні ділянки, які при мікроскопічному дослідженні показали виражений некроз з наявністю великої кількості блідо-аморфної жовтої субстанції, тілець Гамна–Ганді (GGBs) і виражену реакцію гігантських клітин на чужорідне тіло в паренхімі селезінки. Мазок периферичної крові після спленектомії показав тром- боцитоз і гаметоцити Plasmodium falciparum. Електрофорез гемоглобіну виявив підвищення Hb S (49,7%), Hb F – 46,7%; Hb A2 – 3,0% & Hb A – 0,7%, що відповідає серповидноклітинній таласемії Hb S/β+ таласемії. Висновки. Серповидноклітинна таласемія з тривалою спленомегалією, інфарктом селезінки з тільцями Гамна-Ганді і супутньою малярією у клінічно стабільних пацієнтів є справді рідкісним явищем. КЛЮЧОВІ СЛОВА: серповидно-клітинна анемія; бета-таласемія; малярія; малярійний плазмодій; Тільця Гамна–Ганді. Information about the authors Thongam Sachin Singh – Assistant Professor (Pathology), 155 Base Hospital, Tezpur, India https://orcid.org/ 0000­0002­5029­8066, e­mail: sachinthongam@gmail.com Raj Singh – Associate Professor (Pathology), Indian Naval Hospital Ship Asvini, Mumbai, India https://orcid.org/0000­0001­8423­6296, e­mail: rajsingh2u@yahoo.com Rahul Pandey – Associate Professor (Surgery), Command Hospital, Pune, India https://orcid.org/ 0000­0003­2624­5113, e­mail: rahuladviksimpy@gmail.com References 1. Silvestroni E, Bianco I. Microdrepanocito- anemia in un soggetto di razza Bianca. Boll A Accad Med Roma 1944; 70:347. 2. Mohanty D, Colah RB, Gorakshakar AC, Patel RZ, Master DC, Mahanta J, Sharma SK, Chaud- hari U, Ghosh M, Das S, Britt RP, Singh S, Ross C, Jagannathan L, Kaul R, Shukla DK, Mu thuswamy V. Prevalence of β­thalassemia and other haemo­ globinopathies in six cities in India: a multicentre study. J Community Genet.2013; 4(1):33­42. https://doi.org/10.1007/s12687­012­0114­0 3. Serjeant GR, Ashcroft MT, Serjeant BE, Milner PF. The clinical features of sickle-cell- tha- lassaemia in Jamaica. Br J Haematol. 1973; 24(1): 19-30. https://doi.org/10.1111/j.1365­2141.1973. tb05723.x 4. Kinney TR, Ware RE. Compound heterozygous states. Sickle cell disease: basic principles and clinical practice. 1994; 29:437­52. 5. Steinberg MH. Compound heterozygous and other sickle hemoglobinopathies. Disorders of T. S. Singh et al. 10 IN T E R N A L m E d Ic IN E ISSN 2413-6077. IJmmR 2022 Vol. 8 Issue 2 hemoglobin: genetics, pathophysiology, and clinical management. 2001:786­810. 6. Schmugge M, Waye JS, Basran RK, Zurbriggen K, Frischknecht H. THE Hb S/β+-Thalassemia Phenotype Demonstrates that the IVS­I (− 2)(A>C) Mutation is a Mild β­Thalassemia Allele. Hemoglobin. 2008; 32(3):303­7. https://doi.org/10.1080/03630260802004459 7. Divoky V, Baysal E, Schiliro G, Dibenedetto SP, Huisman TH. A mild type of Hb S‐β+‐thalassemia [‐92 (C→T)] in a sicilian family. American journal of hematology. 1993; 42(2):225­6. https://doi.org/10.1002/ajh.2830420216 8. Leffler EM, Band G, Busby GB, Kivinen K, Le QS, Clarke GM, Bojang KA, Conway DJ, Jallow M, Sisay- Joof F, Bougouma EC. Resistance to malaria through structural variation of red blood cell invasion re- ceptors. Science. 2017; 356(6343). https://doi.org/10.1126/science.aam6393 9. Luzzatto L. Sickle cell anaemia and malaria. Mediterranean journal of hematology and infectious diseases. 2012; 4(1). https://doi.org/10.4084/mjhid.2012.065 10. McAuley CF, Webb C, Makani J, Macharia A, Uyoga S, Opi DH, Ndila C, Ngatia A, Scott JA, Marsh K, Williams TN. High mortality from Plasmodium falciparum malaria in children living with sickle cell anemia on the coast of Kenya. Blood, The Journal of the American Society of Hematology. 2010; 116(10):1663­8. https://doi.org/10.1182/blood­2010­01­265249 11. Makani J, Komba AN, Cox SE, Oruo J, Mwamtemi K, Kitundu J, Magesa P, Rwezaula S, Meda E, Mgaya J, Pallangyo K. Malaria in patients with sickle cell anemia: burden, risk factors, and outcome at the outpatient clinic and during hospitalization. Blood, The Journal of the American Society of Hematology. 2010; 115(2):215­20. https://doi.org/10.1182/blood­2009­07­233528 12. Tubman VN, Makani J. Turf wars: exploring splenomegaly in sickle cell disease in malaria‐ endemic regions. British journal of haematology. 2017; 177(6):938­46. https://doi.org/10.1111/bjh.14592 13. Brousse V, Buffet P, Rees D. The spleen and sickle cell disease: the sick (led) spleen. British journal of haematology. 2014; 166(2):165­76. https://doi.org/10.1111/bjh.12950 1 4 . C h o t i v a n i c h K , U d o m s a n g p e t c h R , McGready R, Proux S, Newton P, Pukrittayakamee S, Looareesuwan S, White NJ. Central role of the spleen in malaria parasite clearance. Journal of Infectious Diseases. 2002; 185(10):1538­41. https://doi.org/10.1086/340213 15. Demar M, Legrand E, Hommel D, Esterre P, Carme B. Plasmodium falciparum malaria in sple- nectomized patients: two case reports in French Guiana and a literature review. The American journal of tropical medicine and hygiene. 2004; 71(3):290­3. https://doi.org/10.4269/ajtmh.2004.71.290 16. Doolan DL, Dobaño C, Baird JK. Acquired immunity to malaria. Clinical microbiology reviews. 2009; 22(1):13­36. https://doi.org/10.1128/CMR.00025­08 17. Färnert A, Wyss K, Dashti S, Naucler P. Duration of residency in a non-endemic area and risk of severe malaria in African immigrants. Clinical Microbiology and Infection. 2015; 21(5):494­501. https://doi.org/10.1016/j.cmi.2014.12.011 18. Olaosebikan R, Ernest K, Bojang K, Mokuolu O, Rehman AM, Affara M, Nwakanma D, Kiechel JR, Ogunkunle T, Olagunju T, Murtala R. A randomized trial to compare the safety, tolerability, and effec- tiveness of 3 antimalarial regimens for the prevention of malaria in Nigerian patients with sickle cell di- sease. The Journal of infectious diseases. 2015; 212(4):617­25. https://doi.org/10.1093/infdis/jiv093 19. Kleinschmidt-DeMasters BK. Gamna-Gandy bodies in surgical neuropathology specimens: ob­ servations and a historical note. Journal of Neuro pa- thology & Experimental Neurology. 2004; 63(2):106­ 12. https://doi.org/10.1093/jnen/63.2.106 20. Piccin A, Rizkalla H, Smith O, McMahon C, Furlan C, Murphy C, Negri G, Mc Dermott M. Com- position and significance of splenic Gamna­Gandy bodies in sickle cell anemia. Human pathology. 2012; 43(7):1028­36. https://doi.org/10.1016/j.humpath.2011.08.011 21. Tedeschi LG. The Gamna nodule. Hum Pathol. 1971;2(1):182­3. https://doi.org/10.1016/S0046­8177(71)80030­8 Received 27 August 2022; revised 20 October 2022; accepted 1 November 2022. This is open access article distributed under the Creative Com- mons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. T. S. Singh et al.