ISSN 10227954, Russian Journal of Genetics, 2011, Vol. 47, No. 6, pp. 744–748. © Pleiades Publishing, Inc., 2011.

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Gene Diversity for Haptoglobin and Transferrin Classical Markers among Hindu and Muslim Populations of Aligarh City, India1 G. Ara, Y. H. Siddique, and M. Afzal Department of Zoology, Aligarh Muslim University, Aligarh, 202002, UP, India email: [email protected]; [email protected] Received June 11, 2010

Abstract—The present paper reports the distribution of serum protein markers viz. haptoglobin and transfer rin in two major groups of Aligarh city of North India. In present study we have undertaken a survey of 538 individuals belonging to eight different populations, four from the Hindu community i.e. Brahmin, Bania, Rajput and Jatav, and the rest four among the Muslim community i.e. Syed, Sheikh, Pathan and Ansari. The heterozygosity ranged from 0.2939 (Ansari) to 0.4873 (Brahmin) for haptoglobin and from 0.000 (Rajput) to 0.1498 (Pathan) for transferrin. The values of DST are 0.4122 and 0.4406, and that of GST are 0.5059 and 0.9726 for haptoglobin and transferrin markers respectively. Through FST test, it has been concluded that there is a high genetic differentiation of populations within Hindu and Muslim groups, though there is absence of any significant differences between these groups. DOI: 10.1134/S1022795411060044 1

India offers immense scope and a variety of situa tions to study the processes of population subdivision and their microevolutionary significance. India’s popu lation structure is remarkable in that the population is divided into a large number of strictly defined endoga mous castes, tribes and religious groups. However, most unique and fundamental to the Indian population structure is the existence of endogamous subcastes within these castes, in any particular region or linguistic area. These subgroups are usually characterized by a high degree of isolation, small effective population size, and a high degree of inbreeding. These conditions are conducive for the process of rapid microdifferentiation. There are indications that in most cases, these subcastes might have evolved from the common parental stock through different fissioning events [1, 2]. In India, the Hindu population constitutes the largest community and the second largest is the Muslim popula tion. In the present study, we took both the populations Muslims as well as Hindus. The history of Hindu popula tion goes back to the Aryans who moved into North West of India. They imposed a caste system to organize the new society created by their arrival. They initially put together a hierarchy of four varnas (i.e. castes), which later was expanded to include a fifth category. The caste system is a rigid class structure based on Hinduism which is confined to India. The caste hierarchies most often quoted include the Brahmins – those engaged in sacri fices and priestly functions; the Kshatriyas – rulers and warriors; the Vaishyas – merchants, farmers and trades

1 The article is published in the original.

men; the Shudras – labourers, craftsmen and service professionals [3–6]. On the other hand Muslims belong to two major sects: Sunnis and Shias, while each sect has different biradaris, grouped under Ashraf and Ajlaf [7]. The former comprise of high rank Muslims like Syeds, Sheikhs, Pathans and Moghuls, while later comprise Qureshis, Ansari and Saifis. Later on Arzals are also reported which comprise sweepers and other groups of lower occupation. Though Islam does not distinguish between the groups on any material grounds, the social isolation might have led to differentiation of the groups over many generations including differences in their gene pools [8, 9]. In the present study, we have used two well known biochemical markers: viz. haptoglobin and transferrin. Haptoglobin (abbreviated as Hp) is a protein, which in humans is encoded by the Hp gene. In blood plasma haptoglobin binds free hemoglobin, released from erythrocytes, with high affinity, and thereby inhibits its oxidative activity [10, 11]. Transferrin is a monomeric glycoprotein, found in plasma at an average concen tration of 250 mg/100 ml. The specific ironbinding protein in plasma, it has a role in the transportation and distribution of iron to the body organs, for iron metabolism and prevention of iron loss via the kid neys. Stored in bone marrow as Tfbound iron, it also possesses bacteriostatic and fungistatic activity. Clini cally, decreases in transferrin are observed in congeni tal disorders, newborns, inflammatory diseases, hypo proteinemias and nephrotic syndrome; increases are found in pregnancy, irondeficiency anemias, and inoculation hepatitis. Transferrin is required by all types of cells in cultures for maximal growth. It is,

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GENE DIVERSITY FOR HAPTOGLOBIN AND TRANSFERRIN CLASSICAL MARKERS

therefore, an important transport factor used in defined culture media [12–16]. The present study is an attempt to investigate the genetic and biochemical variation among two main communities of North Indian population, viz. Hindus and Muslims by using serum protein markers hapto globin and transferrin to find their genetic structure and differentiation. Collection of samples The survey was conducted from February 2009 to May 2009 in Aligarh. The survey was confined to 538 individuals of Aligarh city (UP). Households were selected on random a priori basis during day time by door to door contact by volunteers. This included inhibitants of the city having known biradaris, viz. Populations studied : Syed, Sheikh, Pathan, Ansari (Muslims) and Brahmin, Bania, Rajput, Jatav (Hindus). About 3 ml of blood was col lected from each healthy individual of either sex in a vial containing 0.3 ml of 10% EDTA and the blood was brought to the laboratory and kept in freeze for not more than three hours. Laboratory methods Blood serum was separated from the vial on the day of collection by centrifuging for five minutes at 3000 rpm. The red ceils were washed three times in 0.145 M saline; the washed cells were lysed by the addition of an equal volume of distilled water, and afterwards the haemolysates were frozen. Serum samples were electrophoresed in horizontal 11% (w/v) starch gels using the buffer system of Ash ton and Branden at pH 8.2 [17]. Genetic data analysis The phenotypes were recorded for each tract and each sample, and the gene frequencies were calculated according to Hardy Weinberg law using a gene counting method. Het erozygosity, gene diversity parameters and genetic dis tance were calculated using the following equations: 2

H = 1 – ΣX i ,

(1)

Xi – frequency of i–th allele. Gene diversity was calculated using Nei’s (1973) methods of gene diversity analysis in subdivided popu lations [18]: HT = HS + DST. (2) Genetic distance was determined using Nei’s for mula [19]. The normalized identity of gene between X and Y with respect to all loci is defined as follows (Nei’s 1972). I = Ixy/ IxIy.

(3)

The genetic distance D between X and Y is defined as D = –lnI. (4) The dendrogram was constructed using UPGMA clustering method, phylip version 3.63 [20]. Wahlund effect: FST was calculated by the following formula RUSSIAN JOURNAL OF GENETICS

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745 Syed Bania Sheikh Pathan Brahmin

Jatav Rajput Ansari Dendrogram showing genetic relationship among differ ent populations of Aligarh, North India.

FST = 1 – HS/HT.

(5)

The results of two protein loci namely transferrin and haptoglobin are presented. The phenotypes were counted and gene frequencies were determined as per Hardy–Weinberg law. Phenotypic Frequencies and Allele frequencies (Table 1). Heterozygosity. The pooled heterozygosity for the haptoglobin was found to be 0.4054 and for transferrin it was 0.0255. The highest value of heterozygosity was 0.4873 among Brahmins and lowest among Ansaris (0.2939), while for transferrin the Syeds, Banias and Rajputs have zero heterozygosity. Gene diversity. The coefficient of gene differentia tion relative to total population i.e. GST for total popu lation group was 0.5059 for haptoglobin and 0.9726 for transferrin. The value of GST for Muslim population was 0.0259 and 0.0662 and for Hindu population it was 0.0270 and 0.0105 for the markers haptoglobin and transferrin respectively. Genetic distance. The Nei’s genetic distances were cal culated and on the basis of that dendrogram was con structed using UPGMA method. While the Syed and Bania form one cluster, Sheikh and Pathan form another one, joining with Brahmin, whereas Jatav and Rajput constitute one cluster joining with Ansaris (figure). FST Test – see Table 2. Although 83% of the people are Hindu, India also is the home of more than 120 million Muslim – one of the world’s largest Muslim populations. The Indian popula tion also includes Christians, Sikhs, Jains, Buddhists, and Parsis. The caste system reflects the Indian occupa tionally and religiously defined hierarchies [21]. Tradi 2011

68

3. Pathan

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79

62

3. Rajput

4. Jatav

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Pooled

Hindus

240

298

54

2. Bania

Muslims

45

1. Brahmin

Hindus

95

75

2. Sheikh

4. Ansari

60

No

1. Syed

Muslims

Populations

538

44.61

55.39

11.52

14.68

10.04

8.36

17.66

12.64

13.94

11.5

%

38(61.29)

49(62.02)

29(53.70)

22(48.89)

63(66.31)

22(32.33)

35(46.67)

35(58.33)

2–2 (%)

69(28.75) 138(57.50)

117(39.26) 155(52.01)

18(29.03)

21(26.58)

22(40.74)

8(17.78)

30(31.58)

45(66.17)

25(33.33)

17(28.33)

2–1 (%)

59(24.79) 186(34.57) 293(54.46)

33(13.75)

26(8.72)

6(9.68)

9(11.39)

3(5.56)

15(33.33)

2(2.11)

1(1.47)

15(20)

8(13.33)

1–1 (%)

Phenotype frequency

Haptoglobin

0.2825

0.281

0.284

0.242

0.247

0.259

0.422

0.179

0.346

0.367

0.275

Hp1

0.7175

0.719

0.716

0.758

0.753

0.741

0.578

0.821

0.654

0.634

0.725

Hp2

Allele frequency

–

Tf BC (%)

–

–

–

531(98.70) 5(0.929)

238(99.16) 1(0.42)

293(98.32) 4(1.34)

61(66.30)

79(100)

54(100)

44(97.78) 1(2.22)

93(97.89) 1(1.05)

67(98.53) 1(1.47)

73(97.33) 2(2.67)

60(100)

Tf CC (%)

2(0.3717)

1(0.42)

1(0.33)

1(1.613)

–

–

–

1(1.05)

–

–

–

0.9871

0.9958

0.9915

0.992

1.000

1.000

0.989

0.989

0.993

0.987

1.000

Tf C

Transferrin

Tf CD (%)

Phenotype frequency

Table 1. Distribution of haptoglobin and transferrin phenotypes in various Indian populations (Aligarh)

0.0085

0.0021

0.0067

0.000

0.000

0.000

0.011

0.005

0.007

0.013

0.000

Tf B

Allele frequency

0.0051

0.0021

0.0017

0.0080

0.0000

0.0000

0.0000

0.0053

0.0000

0.0000

0.0000

Tf D

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GENE DIVERSITY FOR HAPTOGLOBIN AND TRANSFERRIN CLASSICAL MARKERS

tionally, there are four broad categories of castes (varnas), including a category of outcastes, earlier called “untouchables” but now commonly referred to as “dal its.” Within these broad categories there are thousands of castes and subcastes, whose relative status varies from region to region. Despite economic modernization pro cess and laws countering discrimination against the lower end of the class structure, the caste system remains an important source of social identification for most of the Hindus and also a potent factor in the political life of the country. The exact origin of the caste system as it is today is still obscure. In the present study, we have taken two protein markers viz. haptoglobin and transferrin to study the genetic diversity among North Indian Population. Significant difference in the allele frequencies of Hp and Tf exist in the North Indian population. The Hp2 allele frequency is higher in general, than the Hp1 allele frequencies in the Aligarh population. They dis play a relative order of the magnitudes, namely, Ansari (0.821) > Jatav (0.758) > Rajput (0.753) > Bania (0.741) > Syed (0.725) > Pathan (0.654) > Sheikh (0.634) > Brahmin (0.578). An excess of Hp2 allele has previously been documented from countries geo graphically as far apart as Jordan and Southeast Asia [22]. Similar to Hp2, the allele Tf C shows higher fre quencies compared to Tf B and Tf D allele. The observed heterozygosity is high for the locus haptoglobin as compared to transferrin. The values of HT, HS, DST and GST are comparatively higher. On the basis of genetic distances between different popula tions, dendrogram was constructed using UPGMA clustering method. The dendrogram shows the higher castes of Hindu and those of Muslim biradari are found closer than the lower castes like Jatav of Hindus and Ansaris of Muslims. The most recent are Sheikh and Pathan, and Syed, Bania, Brahmin, Rajput and Jatav are of approximately the same time on the basis of their origin. The Syeds and Bania form one cluster; Sheikh and Pathan form one cluster, and together join with Brahmin. All these populations are the so called higher castes (like Syed, Sheikh, Pathan among Mus lims and Brahmin and Bania among Hindus). Thus Jatav and Rajput appear to form one cluster, which later on join with Ansaris. This dendrogram also justi fies the migration pattern and history of older Indian populations. There is no significant differentiation between Hindu and Muslim populations, as the higher caste Muslims have so called biradaris and lower caste Muslims (Ajlaf) are quite different from upper caste Muslims, and upper and lower caste Muslims of India are converts from Hindu upper and lower caste, this fact is also favoured by Eaaswarkanth M. et al. [22]. Comparison of genetic divergence between Mus lims and Hindus on the basis of haptoglobin and trans ferrin was found to be insignificant (Table 2), which means that both of the population group Hindu and Muslim have same ancestors, which means most of the Muslims have been converted from Hindus; the differ RUSSIAN JOURNAL OF GENETICS

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Table 2. The values of FST to compare the genetic diver gence between and within Muslims and Hindus on the basis of haptoglobin and transferrin Markers Haptoglobin Transferrin

Muslims

Hindus

Muslims vs. Hindus

0.0259 0.0662

0.0271 0.0105

0.000011 0.000821

ence within the Muslim population and Hindu popu lation was found to be significant. It may be because caste system is rigid among Indians. Though caste sys tem is not present among Muslims but biradaris are there, and between these biradaris marriages are usu ally not allowed, i.e. between Ashrafs (higher caste group) and Ajlafs (lower caste group). Whereas among Hindus caste system is very rigid, they do not marry between the castes. Thus the difference was found to be significant within the populations. One more fact to be noted is that transferrin is bet ter marker to compare the gene diversity between the populations as compared to haptoglobin because the value of FST for haptoglobin is very low i.e. 0.000011 while for transferrin its 0.000821. In a study elsewhere to search for a better marker DNA sequences from Helicobacter pylori, was found to be better than Mito chondrial DNA marker and Microsatellite marker, the value of FST for H. pylori marker was 0.031, for mtDNA was 0.021 and for microsatellite 0.007 [23]. ACKNOWLEDGMENTS Thanks are due to the CSIR, New Delhi for award ing the Fellowship No. 9/112(0421)/2K8EMRI to the first author (Gulshan Ara) and to the Chairman, Department of Zoology, A.M.U., Aligarh, U.P., for laboratory facilities. REFERENCES 1. Malhotra, K.C., Morphological Composition of the People of India, J. Hum. Evol., 1978, vol. 7, no. 1, pp. 45–53. 2. Mukherjee, B.N., Majumder, P.P., Malhotra, K.C., et al., Genetic Polymorphism Analysis among Nine Endogamous Population Groups of Maharashtra, India, J. Hum. Evol., 1979, vol. 8, no. 6, pp. 555–565. 3. Karve, I., Hindu Society—an Interpretation, Poona: Deccan College, 1961. 4. Basu, A., Mukherjee, B.N., Roy, S., et al., Ethnic India: A Genomic View, with Special Reference to Peo pling and Structure, Genome Res., 2003, vol. 13, pp. 2277–2290. 5. Majumder, P.P., Ethnic Populations of India as Seen from an Evolutionary Perspective, J. Biosci., 2001, vol. 26, no. 4, pp.533–545. 2011

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