Elsevier

Infection, Genetics and Evolution

Volume 34, August 2015, Pages 153-159
Infection, Genetics and Evolution

The evolutionary origins of Southeast Asian Ovalocytosis

https://doi.org/10.1016/j.meegid.2015.06.002Get rights and content

Highlights

  • SAO is among the oldest human malaria-resistance alleles discovered to date.

  • Considerable haplotype variation exists among chromosomes bearing the SAO mutation.

  • Selection operating on SAO was historically much weaker than it is at present day.

  • SAO likely was the target of a partial “soft sweep”.

Abstract

Southeast Asian Ovalocytosis (SAO) is a common red blood cell disorder that is maintained as a balanced polymorphism in human populations. In individuals heterozygous for the SAO-causing mutation there are minimal detrimental effects and well-documented protection from severe malaria caused by Plasmodium vivax and Plasmodium falciparum; however, the SAO-causing mutation is fully lethal in utero when homozygous. The present-day high frequency of SAO in Island Southeast Asia indicates the trait is maintained by strong heterozygote advantage. Our study elucidates the evolutionary origin of SAO by characterizing DNA sequence variation in a 9.5 kilobase region surrounding the causal mutation in the SLC4A1 gene. We find substantial haplotype diversity among SAO chromosomes and estimate the age of the trait to be approximately 10,005 years (95% CI: 4930–23,200 years). This date is far older than any other human malaria-resistance trait examined previously in Southeast Asia, and considerably pre-dates the widespread adoption of agriculture associated with the spread of speakers of Austronesian languages some 4000 years ago. Using a genealogy-based method we find no evidence of historical positive selection acting on SAO (s = 0.0, 95% CI: 0.0–0.03), in sharp contrast to the strong present-day selection coefficient (e.g., 0.09) estimated from the frequency of this recessively lethal trait. This discrepancy may be due to a recent increase in malaria-driven selection pressure following the spread of agriculture, with SAO targeted as a standing variant by positive selection in malarial populations.

Introduction

Southeast Asian Ovalocytosis (SAO) is a human red blood cell deformity that confers broad-spectrum protection against multiple malaria-causing parasites and represents a canonical example of balancing selection operating via heterozygote advantage in humans. The trait is caused by a 27 base-pair deletion in SLC4A1, the gene encoding the erythrocyte membrane Band 3 protein (Jarolim et al., 1991). SAO is associated with heterozygous carriers of the deletion, and when homozygous the mutation is fully lethal in utero in the absence of extraordinary medical intervention (Genton et al., 1995, Liu et al., 1994, Picard et al., 2014). Evidence for a heterozygote advantage for individuals with SAO in malarial areas comes from case-control studies that have demonstrated nearly complete protection conferred by SAO against severe clinical malaria (including cerebral malaria) and malaria-related mortality caused by Plasmodium falciparum (Allen et al., 1999, Genton et al., 1995, Rosanas-Urgell et al., 2012). This protection may be due to altered cytoadherence properties of infected SAO red blood cells that make cerebral sequestration and associated death less likely (Cortés et al., 2005). Additionally, in a study of children in Papua New Guinea the trait was shown to protect against Plasmodium vivax parasitemia, infection prevalence, and incidence of severe malarial disease (Rosanas-Urgell et al., 2012). These results suggest that SAO provides a broad spectrum of defense against disease caused by multiple Plasmodium species. Further evidence of the role of malaria-driven natural selection acting to maintain the trait comes from a positive correlation between SAO frequency and endemicity of P. falciparum in Island Southeast Asia and Melanesia (Mgone et al., 1996). Indeed, based on its high frequency, a fitness advantage of approximately 9% has been estimated for individuals with SAO in strongly malarial areas of Papua New Guinea (Genton et al., 1995).

SAO is widespread in Island Southeast Asia and neighboring regions on the Malay Peninsula as far north as southern Thailand (Kimura et al., 1998, Kimura et al., 2003, Mgone et al., 1996, Ngouprommin et al., 2012). Individuals with SAO are characterized as having oval-shaped red blood cells with increased membrane rigidity and decreased anion transport, but no clinical symptoms beyond sporadic associations with anemia in both adults and neonates (Amato and Booth, 1977, Coetzer et al., 1996, Laosombat et al., 2005, Laosombat et al., 2010, O’Donnell et al., 1998, Reardon et al., 1993, Schofield et al., 1992). The deletion mutation that causes SAO removes 9 amino acids from within the 11th exon of SLC4A1 and distinguishes SAO from other forms of ovalocytosis in Southeast Asia (Kimura et al., 2006, Patel et al., 2001).

While the protective effects of SAO against malaria are well established, the evolutionary history of this trait is poorly understood. Although it is most common in Island Southeast Asia, it has also been found at low frequency in widely dispersed locations, including Madagascar, Mauritius, South Africa and Sri Lanka (Coetzer et al., 1996, Rabe et al., 2002, Tanner et al., 1991, Vidyatilake and Gooneratne, 2004). One explanation for this pattern is that the trait originated in Island Southeast Asia and then spread across the Indian Ocean via dispersal of speakers of Austronesian languages (Rabe et al., 2002). Austronesian settlement of Madagascar is estimated to have occurred approximately 1200 years ago and originated in the Java–Kalimantan–Sulawesi area (Cox et al., 2012, Pierron et al., 2014), suggesting the trait was present in the Austronesian source population at this time. In communities of Papua New Guinea associations between SAO frequency and Austronesian language have been demonstrated, again suggesting a link between the spread of Austronesian-speaking individuals and the distribution of SAO (Kimura et al., 2003, Tsukahara et al., 2006). However, while many uniquely Austronesian genetic markers have affinities with present-day aboriginal Taiwanese, this pattern is not observed for SAO (Wilder et al., 2009). This discrepancy suggests that the association between SAO and speakers of Austronesian-languages developed only in the later dispersal stages of the Austronesian expansion. As such, the origins of SAO remain unclear and it is not known how long populations in Southeast Asia have possessed this trait.

This study elucidates the origins of SAO as well as the historical selection coefficient affecting the trait by examining patterns of nucleotide diversity and linkage disequilibrium among chromosomes bearing the causal mutation from across its geographical range. Island Southeast Asia harbors a diversity of unique alleles that confer resistance to malarial disease, but few have been examined from an evolutionary perspective. The case of SAO is particularly intriguing, as it represents an outstanding example of a rare balanced polymorphism in humans.

Section snippets

DNA samples and amplification

We examined a total of 60 SAO chromosomes, sampled from four geographically disparate sources: Southern Thai individuals sampled in Thailand (n = 18); one self-identified Chinese and five self-identified Malays sampled in Malaysia (n = 6); and individuals of diverse indigenous Indonesian ancestry from the provinces of North Maluku (n = 19) and East Nusa Tenggara, Indonesia (n = 18). All individuals included in this study were confirmed by diagnostic DNA amplification to be from heterozygous carriers of

Nucleotide variation

We successfully resequenced and experimentally phased a 9472 base-pair portion of the SLC4A1 gene in 60 SAO heterozygotes from Southeast Asia, spanning Thailand to eastern Indonesia. Our phased DNA sequence data uncovered a total of 31 separate haplotypes, including 5 among SAO chromosomes and 26 from non-SAO chromosomes (Fig. 1). These haplotypes are defined by 36 unique event polymorphisms, including 33 single nucleotide polymorphisms (SNPs), 2 single-base indels, and the 27 bp SAO-causing

Discussion

The evolution of malaria resistance in human populations from Island Southeast Asia is poorly understood. Our study demonstrates that SAO is the result of a single mutation event that we estimate to have occurred between 170 and 800 generations ago (4930–23,200 years bp), with point estimates from two different methods spanning a relatively narrow range from 294 to 345 generations (8526–10,005 years bp). During the time since SAO arose in human populations there has been opportunity for substantial

Acknowledgements

We would like to thank John Chase, Milly Chandler and Dylan Funk for assistance with data collection and members of the Wilder laboratory and two anonymous reviewers for comments on the manuscript. This study was funded by a grant from the National Science Foundation (BCS-1062258) to J.A.W., and the funding source had no role in the design or implementation of the research.

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