• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of geneticsGeneticsCurrent IssueInformation for AuthorsEditorial BoardSubscribeSubmit a Manuscript
Genetics. Nov 2005; 171(3): 1219–1229.
PMCID: PMC1456824

The Extent of Linkage Disequilibrium Caused by Selection on G6PD in Humans


The gene coding for glucose-6-phosphate dehydrogenase (G6PD) is subject to positive selection by malaria in some human populations. The G6PD A− allele, which is common in sub-Saharan Africa, is associated with deficient enzyme activity and protection from severe malaria. To delimit the impact of selection on patterns of linkage disequilibrium (LD) and nucleotide diversity, we resequenced 5.1 kb at G6PD and ~2–3 kb at each of eight loci in a 2.5-Mb region roughly centered on G6PD in a diverse sub-Saharan African panel of 51 unrelated men (including 20 G6PD A−, 11 G6PD A+, and 20 G6PD B chromosomes). The signature of selection is evident in the absence of genetic variation at G6PD and at three neighboring loci within 0.9 Mb from G6PD among all individuals bearing G6PD A− alleles. A genomic region of ~1.6 Mb around G6PD was characterized by long-range LD associated with the A− alleles. These patterns of nucleotide variability and LD suggest that G6PD A− is younger than previous age estimates and has increased in frequency in sub-Saharan Africa due to strong selection (0.1 < s < 0.2). These results also show that selection can lead to nonrandom associations among SNPs over great physical and genetic distances, even in African populations.

RECENT studies have focused on describing and understanding the general structure of linkage disequilibrium (LD) in the human genome, primarily to provide a sound basis for mapping disease loci in association studies (Kruglyak 1999; Goldstein 2001). Patterns of LD are expected to be complicated because LD is affected by many factors, including genetic drift, population structure, migration, admixture, selection, mutation, gene conversion, and recombination (Ardlie et al. 2002). Moreover, some of these factors, such as recombination, are not constant across the genome (McVean et al. 2004), and thus LD is expected to vary in different genomic regions. Despite this expected complexity, several general results have emerged from empirical studies of LD in humans. First, the human genome is divided into haplotype blocks, with regions of high LD over fairly long stretches, separated by regions with little LD (Daly et al. 2001; Gabriel et al. 2002; Phillips et al. 2003; Wall and Pritchard 2003). There is clear evidence that the spaces between these blocks correspond to recombination hotspots in some cases (e.g., Jeffreys et al. 2001), although simulations suggest that a block-like pattern may be expected even in the absence of recombination hotspots (Phillips et al. 2003). Recombination hotspots may occur in the human genome roughly every 200 kb (McVean et al. 2004). Second, there appears to be less LD in African populations than in non-African populations (Tishkoff et al. 1996; Reich et al. 2001). This observation is consistent with the presumed larger long-term effective population size for African populations. Third, selection on individual genes can elevate levels of LD in a given genomic region (e.g., Huttley et al. 1999; Sabeti et al. 2002; Saunders et al. 2002; Toomajian and Kreitman 2002; Swallow 2003). In fact, this expectation has motivated several statistical tests of a neutral model of molecular evolution (e.g., Hudson et al. 1994; Kelly 1997; Slatkin and Bertorelle 2001; Sabeti et al. 2002; Toomajian et al. 2003). It is difficult to predict exactly how far the effects of selection will extend because the observed patterns will depend on multiple factors, including the type of selection (e.g., balancing, purifying, directional), the time over which selection has acted, the strength of selection, the local recombination rate, and various demographic factors. To study this problem empirically, we have chosen to focus on the genomic region surrounding G6PD, a gene known to be subject to selection in humans.

Glucose-6-phosphate dehydrogenase (G6PD) is a housekeeping enzyme that catalyzes a critical step in the pentose monophosphate shunt of glycolysis. G6PD deficiency mutations cause hemolytic anemia and neonatal jaundice (Beutler 1994); however, some deficiency mutations also confer resistance to severe malaria (Allison 1960; Motulsky 1961; Ruwende et al. 1995). Many human populations exhibit G6PD deficiency alleles at frequencies that range between 0.05 and 0.20 as a consequence of selection. G6PD A− is a common deficiency allele in sub-Saharan Africa that reaches frequencies of ~0.2 in populations living in malarial areas (Livingstone 1985; Cavalli-Sforza et al. 1996). This allele is characterized by two nonsynonymous changes relative to the normal allele (G6PD B) (Figure 1), which decrease enzyme activity to ~12% of normal (Hirono and Beutler 1988) and confer ~50% reduction in risk of severe malaria in both females and males (Ruwende et al. 1995). It follows that the G6PD A− allele is beneficial in the presence of malaria caused by Plasmodium falciparum, while in the absence of malaria this allele is deleterious. The wealth of knowledge and the clear understanding of genotype-phenotype connections for G6PD make it a useful model for studying the effects of selection on patterns of LD in humans.

Figure 1.
Ideogram of the human X chromosome and the genomic regions surveyed in this study. Approximate distances between each of the surveyed windows including G6PD are marked on the scale. Transcription orientations of the genic regions are marked with solid ...

Recently, several studies have investigated patterns of nucleotide variability at G6PD and at loci relatively close to G6PD (Tishkoff et al. 2001; Sabeti et al. 2002; Saunders et al. 2002; Verrelli et al. 2002). All of these studies documented LD associated with the G6PD A− allele. In particular, Sabeti et al. (2002) and Saunders et al. (2002) showed that LD extended over ~550 kb in an African sample. Neither of these studies surveyed loci beyond this distance, and therefore they were unable to delimit the full extent of LD caused by selection on G6PD. Here, we extend these results to delimit the genomic region over which selection at G6PD has created LD. We resequenced ~3-kb windows from each of eight loci in a 2.5-Mb region, centered roughly on G6PD in a panel of 51 individuals from sub-Saharan Africa. In this panel, we also resequenced 5.1 kb at G6PD and ~2 kb at an unlinked “control” locus, situated 19 Mb proximal to G6PD. Our data show that selection at G6PD has affected a region that spans >1.6 Mb of the human X chromosome, demonstrating that selection can have considerable effects on nucleotide variability over remarkably long genomic distances in humans.



DNA sequences were determined in a sample of 51 human males of African descent (Table 1) that includes 20 G6PD A− alleles, 11 G6PD A+ alleles, and 20 G6PD B alleles. The G6PD A− allele is defined by two mutations: an A → G nonsynonymous mutation at G6PD coding site 376, co-occurring with a G → A nonsynonymous mutation at coding site 202 (Figure 1) (Hirono and Beutler 1988). G6PD A+ is defined by the A → G nonsynonymous mutation at G6PD coding site 376 that reduces enzyme efficiency to 80% of normal (Figure 1). This mild deficiency allele does not confer resistance to malaria (Ruwende et al. 1995) and is found in sub-Saharan Africa at a frequency of ~0.2 (Takizawa et al. 1987). All G6PD functional alleles were determined a priori by restriction fragment length polymorphism analysis of a FokI restriction site at coding position 376 and a NlaIII restriction site at coding position 202 (Xu et al. 1995). As G6PD A− is believed to be of a single origin (Saunders et al. 2002; Verrelli et al. 2002) we selected individuals to represent diverse localities in sub-Saharan Africa. By studying X-linked loci in males we were able to PCR amplify single alleles and directly recover haplotypes over long genomic distances to study patterns of LD. Homologous sequences from a chimpanzee (Pan troglodytes) and an orangutan (Pongo pygmaeus) were also determined at each locus for divergence estimates. All sampling protocols were approved by the Human Subjects Committee at the University of Arizona.

Individuals sampled in this study

Loci surveyed:

G6PD and nine flanking loci (G18MC, G1.5MC, IDH3G, BGN, L1CAM, TAZ, GAB3, F8, and G0.9MT) were surveyed for nucleotide variability (Figure 1). Loci G18MC, G1.5MC, and G0.9MT (our nomenclature) are anonymous intergenic regions while the remainder of the regions surveyed are primarily introns. All loci were chosen because of their physical distance from G6PD, and there is no evidence that any of these loci are themselves targets of selection. Approximately 30 other genes are found within 1 Mb on either side of G6PD and none of these genes is known to be recent targets of positive directional selection in sub-Saharan Africa. All loci chosen are single copy in the genome and are situated outside the pseudoautosomal region. G0.9MT is situated near the boundary of the pseudoautosomal region, and we did not survey loci distal to this locus.

PCR amplification and sequencing:

Single PCR fragments (2–5 kb) were amplified for each individual at each locus using a long-range PCR system (Invitrogen HiFi Taq). Amplification primers for G6PD and L1CAM are found in Saunders et al. (2002) and primers for all other loci can be found at www.genetics.org/supplemental (Table S1). Internal primers were used to generate overlapping sequence runs on an ABI 3730 automated sequencer. Contiguous sequence that included coding and noncoding regions was assembled for each individual for each locus, using the computer program SEQUENCHER (Gene Codes, Ann Arbor, MI). Sequences have been submitted to GenBank under accession nos. DQ173562DQ173642.

Nucleotide variability data analysis:

Nucleotide variability and the frequency spectrum of alleles in an unbiased African sample have been reported in detail elsewhere for G6PD (Saunders et al. 2002; Verrelli et al. 2002). To gain insight into the long-range effects of selection on nucleotide variability we analyzed three subsets of our data: G6PD A−, G6PD A+, and G6PD B alleles. We calculated haplotype diversity (Hd), θπ (Nei and Li 1979), and θw (Watterson 1975) at each locus using dnaSP 4.0 (Rozas et al. 2003). Under neutral equilibrium conditions both θπ and θw for a random sample estimate the neutral parameter 3Neμ for X-linked loci, where Ne is the effective population size and μ is the neutral mutation rate, assuming a sex ratio of one. However, the structure of the sample in the present study is nonrandom, and therefore θπ and θw are used simply as measures of nucleotide variability. We also analyzed nucleotide variability in a constructed random sample (CRS) (Hudson et al. 1994). This subset of chromosomes (n = 26) contains G6PD alleles at frequencies that are representative of a typical sub-Saharan African population subject to malarial selection, on the basis of extensive allele frequency surveys (G6PD A−, 0.11; G6PD A+, 0.20; G6PD B, 0.69) (Livingstone 1985). We calculated θπ, θw, Tajima's D (Tajima 1989), and Fu and Li's D (Fu and Li 1993) at each locus for the CRS to compare to nonbiased African samples that are available for different X-linked loci. Detailed statistics of nucleotide variability for the CRS are available at www.genetics.org/supplemental (Table S2). Divergence data were derived for each of these loci by calculating the average of all pairwise comparisons between the homologous sequence from an outgroup and the samples in the CRS. LD between pairs of polymorphic sites was measured using the statistic |D′| (Lewontin 1964). This measure of linkage disequilibrium is standardized to equal 0 when there is random association among polymorphisms (i.e., no disequilibrium) and to equal 1 when there is complete association among polymorphisms (i.e., complete disequilibrium).

The age of the G6PD A− allele:

The age of the G6PD A− allele and the intensity of past selection it experienced were estimated by combining the method of Slatkin (2001) for generating intraallelic genealogies of selected alleles with the method of Garner and Slatkin (2002) for estimating the probability of haplotypes at two linked loci. All analyses were performed on the basis of long-range haplotypes, using the intralocus combination of sites 55, 59, and 60 at L1CAM; SNP 90 at G6PD (i.e., coding site 202); and the intralocus combination of sites 99, 100, and 101 at G0.9MT (Figure 2). The computer program described by Slatkin (2001) was used to generate sample paths of allele frequency from the time of the mutation (t1, the allele age) until the present (t = 0), with the constraint that the frequency at t = 0 is the observed frequency, 0.1. An additive dominance model was used. We assumed a constant population size of Ne = 10,000 and Ne = 20,000 individuals. For each sample path, a neutral coalescent model was used to generate an intraallelic genealogy of G6PD A− since it arose by mutation. The intraallelic coalescence times from this genealogy were then passed as parameters to a program that estimates the probability of obtaining the observed configuration of the data (the numbers of the four haplotypes found on the 20 A−-bearing chromosomes), given the recombination rates and haplotype frequencies on non-A−-bearing chromosomes (assumed constant). That probability is the likelihood of the data, given the intraallelic genealogy. For each value of s, the selective advantage of A− was considered, 90,000 sample paths and intraallelic genealogies were generated, and 10 replicates of the Garner-Slatkin program were used to estimate the likelihood for each sample path. Likelihoods were averaged across sample paths, using the weighting method described by Slatkin (2001). For each parameter value, this method provided an estimate of the likelihood of the data under the model and an estimate of the posterior distribution of allele age.

Figure 2.
Table of polymorphism for G6PD and surrounding loci. Fifty-one unrelated human males of sub-Saharan African descent were surveyed for nucleotide variability at G6PD and nine surrounding loci. Individual samples were selected on the basis of a priori allele ...

To allow analysis of the multisite data set by this method, we used the fact that all 20 A− chromosomes carried the same haplotype for 925 kb telomeric to G6PD (to locus G0.9MT) and that 14 of 20 chromosomes carried the same haplotype for 556 kb centromeric to G6PD (see SNPs 55, 59, and 60 at L1CAM; Figure 2). The recombination parameters in the two directions were assumed to be c = 0.01675 and 0.00555 M for L1CAM and G0.9MT, respectively (Kong et al. 2002). Therefore, the two-locus data set was 14 AMB, 6 aMB, 0 AMb, 0 aMb (in the notation of Garner and Slatkin 2002), where A represents the haplotype of rare alleles at L1CAM SNPs 55, 59, and 60 (which are very rare on non-A− chromosomes), M represents the site under selection, and B represents the multilocus haplotype at G0.9MT, which is rare on non-A− chromosomes.


Nucleotide diversity at G6PD:

Polymorphic sites at G6PD are presented in Figure 2. We calculated nucleotide variability for three subset groups: (i) individuals bearing the G6PD A− allele (n = 20), (ii) individuals bearing the G6PD A+ allele (n = 11), and (iii) individuals bearing the G6PD B allele (n = 20). At G6PD we observed 18 segregating sites (excluding three INDELs) in the entire sample, consistent with previous findings (Saunders et al. 2002; Verrelli et al. 2002). Among the G6PD A− individuals (n = 20) there was no nucleotide variability in 5109 bp of contiguous DNA sequence. Noncoding nucleotide variability among G6PD A+ and G6PD B alleles was θπ = 0.024 and 0.04%, respectively (Figure 3a). Nucleotide variability for the CRS was θπ = 0.068% and θw = 0.090% (Figure 3a; Table S2 at http://www.genetics.org/supplemental/), consistent with previously reported levels of nucleotide variability at G6PD in sub-Saharan Africa (Sabeti et al. 2002; Saunders et al. 2002; Verrelli et al. 2002) and with the average of 15 other X-linked loci (θπ = 0.0755%, θw = 0.0815%) (Hammer et al. 2004). Tajima's D and Fu and Li's D for the CRS were −0.794 and −0.142, respectively (Table S2 at http://www.genetics.org/supplemental/), also consistent with the previous studies at G6PD.

Figure 3.
Nucleotide variability for G6PD and nine surrounding loci for subset groups of the data set (G6PD A− alleles, G6PD A+ alleles, G6PD B alleles, and CRS): (a) nucleotide diversity (θπ); (b) haplotype diversity.

Nucleotide diversity around G6PD:

The segregating sites for nine loci flanking G6PD are presented in Figure 2. At GAB3, F8, and G0.9MT, loci distal to G6PD, we found no nucleotide variability in the A− group (Figure 3a). This portion of the data includes a cumulative survey of 13,582 bp, thus exhibiting a remarkable degree of nucleotide homogeneity among 20 unrelated individuals of African decent. At these same loci, the average noncoding nucleotide variability for the A+ group, the B group, and the CRS was θπ = 0.027, 0.024, 0.022% respectively (Figure 3a). Estimates of haplotype diversity for the different allele classes also demonstrate a significant contrast between the A− alleles and the other allele classes. At loci distal to G6PD, the average haplotype diversity was Hd = 0.0, 0.435, and 0.879 for A−, A+, and B, respectively (Figure 3b).

This general pattern of reduced variability among A− individuals is also seen proximal to G6PD; however, the pattern is not as extreme as that on the distal side, and the pattern decays beyond L1CAM (~556 kb from G6PD; Figure 3, a and b). At TAZ, L1CAM, and IDH3G the average noncoding nucleotide variability for the A− group, A+ group, B group, and CRS was θπ = 0.032, 0.037, 0.040, and 0.041%, respectively (Figure 3a; Table S2 at http://www.genetics.org/supplemental/). At BGN, G1.5MC, and G18MC, loci mapping 0.9–19 Mb from G6PD, the average noncoding nucleotide variability for the A− group, A+ group, B group, and CRS was θπ = 0.093, 0.072, 0.091, and 0.082%, respectively (Figure 3a; Table S2 at http://www.genetics.org/supplemental/). At these three loci the average level of nucleotide variability among the A− individuals is not reduced relative to that of the other subsets of the data. Together our results demonstrate that the A− chromosomes exhibit reduced variability relative to other allelic classes at loci around G6PD over a region that spans ~1.5 Mb (roughly from L1CAM to G0.9MT). This effect may extend further distally, but we were unable to survey loci beyond G0.9MT, which lies near the border of the pseudoautosomal region (see materials and methods).

Linkage disequilibrium:

To examine patterns of linkage disequilibrium we calculated |D′| (Lewontin 1964) for all pairwise comparisons of segregating sites for which the minor allele was found in five or more individuals (Figure 4). Intragenic pairwise comparisons show strong LD within each of the loci surveyed, consistent with expectations over short distances. However, a striking feature of the data is seen in intergenic comparisons. Within G6PD, all A− individuals share a common haplotype that differs from the consensus G6PD B allele at six sites (segregating sites 82, 83, 84, 87, 90, and 91). These sites exhibit strong LD (significant by Fisher's exact test) with sites at L1CAM (sites 55, 59, and 60), IDH3G (site 41), GAB3 (sites 92 and 94), and G0.9MT (site 99). Furthermore, site 99 at G0.9MT exhibits complete LD (D′ = 1) with the aforementioned sites at L1CAM and IDH3G. Together, this pattern defines a conserved G6PD A− haplotype that spans >1.6 Mb encompassing G6PD. Although site 21 at BGN also exhibits strong LD (D′ = 1) with three sites associated with the common G6PD A− haplotype (sites 83, 84, and 90), this pattern does not represent conservation of the extended A− haplotype, as sites such as 17, 19, 23, 25, and 36 at BGN do not exhibit strong LD with G6PD. When G6PD A− individuals are excluded from the analysis, few intergenic associations in significant LD are found. At IDH3G a haplotype that consists of the minor alleles at sites 38 and 48 is found in significant LD with site 36 of BGN and site 54 of L1CAM. Significant intergenic LD is also found between site 75 of G6PD and sites 92 and 94 of GAB3. This LD is not associated with any known functional alleles. As the minor allele polymorphisms in each of these cases are not associated with the extended G6PD A− haplotype, this LD remains intact when G6PD A− individuals are excluded from analyses. In summary, these data demonstrate that a majority of the intergenic LD in the surveyed region is due to the extended G6PD A− haplotype.

Figure 4.
Patterns of LD between segregating sites at G6PD and flanking loci. The table of polymorphism includes only segregating sites at which the less common allele (minor allele) is found in five or more individuals. At each segregating site, one allele is ...

Age of the G6PD A− allele and strength of selection:

Figure 5A shows the likelihood of the two-locus data set described in materials and methods as a function of s, the assumed selective advantage of A−-bearing chromosomes. Three curves are shown, one based on the recombination rates (c = 0.01675 and 0.00555 for L1CAM and G0.9MT, respectively), one based on assuming half those values (c = 0.008375 and 0.002775), and one based on assuming twice those rates (c = 0.0335 and 0.0111). Although the choice of recombination rate affects the estimated likelihoods, the qualitative results are the same. For all three sets of values, s is likely to be at least 0.1. This method does not allow us to place an upper bound on s, but on other grounds we can exclude values >0.2, which is roughly the selective advantage of individuals heterozygous for the S allele at the β-globin locus (HB-S) in malarial regions, which is thought to have a higher selection coefficient than G6PD deficiency with respect to malarial protection (Allison 1964).

Figure 5.
Results of the evolutionary analysis of the G6PD A− allele. Results were obtained by combining the importance sampling method of Slatkin (2001) for averaging over replicate sample paths with the method of Garner and Slatkin (2002) for computing ...

The posterior distribution of the age of A− depends on s. Figure 5B shows the distributions for two values of s, assuming the estimated recombination rates. For s = 0.1, which is the smallest value consistent with the observations, the estimated age is ~100 generations with an upper bound of <150 generations. The posterior distribution depends only slightly on the recombination rates, as shown in Figure 5C. Most of the information about the age of a strongly advantageous allele is contained in the frequency, not in the extent of linkage disequilibrium with nearby marker alleles.


Nucleotide variability around G6PD:

We investigated patterns of nucleotide variability at G6PD, a locus known to be under natural selection by malaria in Africa, and at nine flanking loci at varying distances from G6PD. Previous studies have shown that in sub-Saharan Africa levels of nucleotide variability at G6PD are typical of those at other X-linked loci, and tests of neutrality based on the frequency spectrum of alleles do not deviate from neutral equilibrium expectations (Sabeti et al. 2002; Saunders et al. 2002; Verrelli et al. 2002). Our data corroborate these results. Nonetheless, we find that the levels of nucleotide variability among G6PD A− alleles are reduced over ~1.5 Mb of the X chromosome, roughly from L1CAM to G0.9MT, demonstrating the strong impact of selection at G6PD on neighboring genes.

Conserved extended haplotype among G6PD A− chromosomes:

The effect of selection on G6PD is also seen in the extent of LD. Previous studies documented LD between G6PD and SNPs within ~600 kb around G6PD in Africa (Sabeti et al. 2002; Saunders et al. 2002). Here we have shown that the ancestral G6PD A− extended haplotype spans >1.6 Mb. A genome-wide survey of the half distance of |D′| (the distance at which D′ decays to half its maximal value, |D′| = 0.5) was ~100 and ~5 kb for a Caucasian and an African population, respectively (Reich et al. 2001). Short-range LD in African populations relative to non-African populations is common for most human data sets (e.g., Tishkoff et al. 1996; Wall and Pritchard 2003), making the finding of such extensive LD associated with G6PD A− in Africa highly unusual.

The pattern of extensive LD seen in these data is consistent with recent strong selection at G6PD accompanied by hitchhiking of SNPs that preexisted on the ancestral G6PD A− chromosome. However, patterns of LD may also be created by population admixture and/or underlying population subdivision in a sample. This is a potential concern in this study because 11 of the 20 G6PD A− individuals are African-American. However, 4 of the African-American samples (VA088, VA076, VA025, and M115) have a disrupted G6PD A− ancestral extended haplotype at IDH3G (contributing to more than half of the G6PD A− extended haplotype recombinants at this locus). Furthermore, the African-American G6PD A− samples considered alone have levels of nucleotide variability similar to those of the non African-American G6PD A− samples considered alone (data not shown). Along with the detection of a portion of this extended G6PD A− haplotype by Sabeti et al. (2002) that included 252 sub-Saharan African (i.e., non-African-American) samples, these results suggest that an overrepresentation of African-American G6PD A− samples is not a major factor contributing to the long-range LD seen here, and that selection is the most likely explanation for the atypical pattern of LD.

The effect of selection on LD has been studied at several other genes. The HLA region exhibits long-range LD in general, and in a non-African panel Sanchez-Mazas et al. (2000) detected LD in this region spanning ~1.3 Mb. However, given the local recombination rate in this region, the genetic distance over which LD is found is not significantly higher than the genome average (Walsh et al. 2003). The long-range LD found in the HLA region might not be due to physical linkage, but instead may be a result of epistatic interactions that create nonrandom combinations of alleles that are advantageous for immune response (Meyer and Thomson 2001). Significant LD was detected over 20 kb around FY (Duffy) in a non-African sample, consistent with recent selection by P. vivax (Hamblin et al. 2002). At HB-globin), long-range LD was detected among SNPs spanning nearly 100 kb in association with HB-E alleles in a Thai population, consistent with selection by malaria (Ohashi et al. 2004). And at LCT, significant LD has been reported, spanning >800 kb in association with lactase persistence alleles in a European-American population (Bersaglieri et al. 2004).

The decay of the G6PD A− extended haplotype (EH) is slightly asymmetrical around the target of selection. The EH decays between 705 kb (at IDH3G) and 991 kb (at BGN) proximal to G6PD, whereas it remains fully conserved among all 20 G6PD A− chromosomes at 925 kb (at G0.9MT) distal to G6PD. Genetic hitchhiking around a target of selection is not necessarily expected to exhibit a symmetrical pattern, even in the face of homogeneous recombination rates across the affected region (Kim and Stephan 2002). Nonetheless, we note that the extended G6PD A− haplotype spans a region exhibiting heterogeneity in recombination rate. For example, the sex-averaged local recombination rate for the region spanning from G6PD to BGN is ~2.0 cM/Mb, while the estimated recombination rate between G6PD and G0.9MT is ~0.6 cM/Mb (UCSC human map viewer based on Kong et al. 2002). The relatively low local recombination rate distal to G6PD is consistent with the absence of recombinant G6PD A− extended haplotypes in this region. It seems unlikely, however, that the conserved G6PD A− EH extends much further in the distal direction, since G0.9MT is adjacent to the q-arm pseudoautosomal region of the X chromosome, where recombination rates are substantially higher.

The observation that the extended G6PD A− haplotype spans >1.6 Mb has interesting implications given that >60 additional genes (including 38 Online Mendelian Inheritance in Man, OMIM, loci) have been identified in this region. In the event that a functional trait (not related to G6PD deficiency) is associated with an ancestral G6PD deficiency EH, this trait could increase in frequency along with the target of selection at G6PD. For example, the gene OPN1MW (OMIM no. 303800) that is responsible for green color blindness is located within this region (~350 kb proximal to G6PD). A study by Filosa et al. (1993) demonstrated that in a region of Calabria that bears the Mediterranean-type G6PD deficiency allele (G6PDmed coding site 563 C → T), all individuals with the 563 C → T mutation (n = 7) were also deutan (green) color-blind on the basis of a visual acuity test. This suggests that in this population a chromosome that carried a G6PDmed allele also harbored a mutation causing a clinical condition of deutan color blindness. Presumably, as the G6PDmed mutation was favorably selected in this population, the deutan color blindness trait hitchhiked on the EH. In our data, the polymorphism at site 41 in IDH3G (Figure 2) causes a nonconservative amino acid change (Arg → Cys) that is found on 14 of the 20 G6PD A− chromosomes and is rarely found on any other G6PD allelic background. The phenotypic consequences of this polymorphism, if any, are unknown. However, given that this polymorphism is in significant LD with G6PD site 202, our data suggest that it may be at its current frequency in Africa due to a hitchhiking event with the G6PD A− allele.

Age of the G6PD A− allele and magnitude of selection:

Previous estimates for the age of G6PD A− suggest that the allele is young (<20,000 years) on the basis of closely linked microsatellite variability (Tishkoff et al. 2001), coalescent-based analysis of a G6PD gene tree (Coop and Griffiths 2004), and intergenic LD (Saunders et al. 2002). Our current analysis suggests that the likely age of the G6PD A− allele is ~100 generations with an upper bound of 150 generations given a selection coefficient against the normal (G6PD B) homozygotes of s ≈ 0.1. This age estimate (2500–3750 years, assuming a 25-year generation time) is somewhat younger than a previous age estimate based on intraallelic microsatellite variability (3840–11,760 years with s = 0.044) (Tishkoff et al. 2001). The discrepancy between these two age estimates may be due to the different selection coefficients that were estimated or to uncertainty in microsatellite mutation rate and/or recombination rates in Xq28. Coalescent-based analyses utilizing ~5 kb from G6PD provide a relatively old age estimate of >9500 years (Verrelli et al. 2002; Coop and Griffiths 2004). Although this analytical method is generally powerful, in this case the structure of the data (i.e., homogeneity among G6PD A− alleles) precludes a precise estimate of the age of the A− allele.

Our likelihood analysis suggests that the selection coefficient for the G6PD A− allele is large; however, it is similar in magnitude to other selection coefficients estimated in humans. For example, selection coefficients of s = 0.26, 0.30, and ~0.15 have been proposed for HB-S (Allison 1956), CCR5Δ32 (Schliekelman et al. 2001), and some HLA alleles (Satta et al. 1994), respectively, for protection from infectious diseases in humans. It is possible that these large values may reflect an ascertainment bias toward recognizing loci under strong selection.


Selection at G6PD is strong and recent, consistent with an adaptive response to a recent increase in virulence of P. falciparum in sub-Saharan Africa (Ruwende et al. 1995; Tishkoff et al. 2001; Sabeti et al. 2002; Saunders et al. 2002). The rapid increase in frequency of G6PD A− has resulted in retention of the ancestral haplotype among the majority of G6PD A− chromosomes spanning >1.6 Mb (~1% of the human X chromosome). This provides a dramatic example of the extent to which recent positive selection can generate long-range LD in human populations. Moreover, if selection is specific to particular geographic regions, as is the case here, it may lead to large differences in patterns of LD for the same genomic region in different populations. This highlights the need for population-specific haplotype maps in association studies.


We thank J. Kim, D. Garrigan, and A. Indap for technical assistance. Some human DNA samples were kindly donated by L. Luzzatto, K. Nafa, Rex Riis, and Jeffrey Ban. R. O. Ryder provided chimpanzee and orangutan samples. B. A. Payseur, E. T. Wood, H. E. Hoekstra, and A. J. Redd provided helpful discussion. D. Begun and two anonymous reviewers provided helpful comments. This material is based on work supported by the National Science Foundation under grant no. 0206756.


  • Allison, A. C., 1956. The sickle cell and haemoglobin C genes in some African populations. Ann. Hum. Genet. 21: 67–89. [PubMed]
  • Allison, A. C., 1960. Glucose-6-phosphate dehydrogenase deficiency in red blood cells of East Africans. Nature 186: 531. [PubMed]
  • Allison, A. C., 1964. Polymorphism and natural selection in human populations. Cold Spring Harbor Symp. Quant. Biol. 29: 137–149. [PubMed]
  • Ardlie, K. G., L. Kruglyak and M. Seielstad, 2002. Patterns of linkage disequilibrium in the human genome. Nat. Rev. Genet. 3: 299–309. [PubMed]
  • Bersaglieri, T., P. C. Sabeti, N. Patterson, T. Vanderploeg, S. F. Schaffner et al., 2004. Genetic signatures of strong recent positive selection at the lactase gene. Am. J. Hum. Genet. 74: 1111–1120. [PMC free article] [PubMed]
  • Beutler, E., 1994. G6PD deficiency. Blood 84: 3613–3636. [PubMed]
  • Cavalli-Sforza, L. L., P. Menozzi and A. Piazza, 1996. The History and Geography of Human Genes. Princeton University Press, Princeton, NJ.
  • Coop, G., and R. C. Griffiths, 2004. Ancestral inference on gene trees under selection. Theor. Popul. Biol. 66: 219–232. [PubMed]
  • Daly, M. J., J. D. Rioux, S. E. Schaffner, T. J. Hudson and E. S. Lander, 2001. High-resolution haplotype structure in the human genome. Nat. Genet. 29: 229–232. [PubMed]
  • Filosa, S., V. Calabro, G. Lania, T. J. Vulliamy, C. Brancati et al., 1993. G6PD haplotypes spanning Xq28 from F8C to red-green color-vision. Genomics 17: 6–14. [PubMed]
  • Fu, Y. X., and W.-H. Li, 1993. Statistical tests of neutrality of mutations. Genetics 133: 693–709. [PMC free article] [PubMed]
  • Gabriel, S. B., S. F. Schaffner, H. Nguyen, J. M. Moore, J. Roy et al., 2002. The structure of haplotype blocks in the human genome. Science 296: 2225–2229. [PubMed]
  • Garner, C. P., and M. Slatkin, 2002. Likelihood-based disequilibrium mapping for two-marker haplotype data. Theor. Popul. Biol. 61: 153–161. [PubMed]
  • Goldstein, D. B., 2001. Islands of linkage disequilibrium. Nat. Genet. 29: 109–111. [PubMed]
  • Hamblin, M. T., E. E. Thompson and A. DiRienzo, 2002. Complex signatures of natural selection at the Duffy blood group locus. Am. J. Hum. Genet. 70: 369–383. [PMC free article] [PubMed]
  • Hammer, M. F., D. Garrigan, E. T. Wood, J. A. Wilder, Z. Mobasher et al., 2004. Heterogeneous patterns of variation among multiple human X-linked loci: the possible role of diversity-reducing selection in non-Africans. Genetics 167: 1841–1853. [PMC free article] [PubMed]
  • Hirono, A., and E. Beutler, 1988. Molecular cloning and nucleotide sequence of cDNA for human glucose-6-phosphate dehydrogenase variant (A)−. Proc. Natl. Acad. Sci. USA 85: 3951–3954. [PMC free article] [PubMed]
  • Hudson, R. R., K. Bailey, D. Skarecky, J. Kwiatowski and F. J. Ayala, 1994. Evidence for positive selection in the superoxide dismutase (Sod) region of Drosophila melanogaster. Genetics 136: 1329–1340. [PMC free article] [PubMed]
  • Huttley, G. A., M. W. Smith, M. Carrington and S. J. O'Brien, 1999. A scan for linkage disequilibrium across the human genome. Genetics 152: 1711–1722. [PMC free article] [PubMed]
  • Jeffreys, A. J., L. Kauppi and R. Neumann, 2001. Intensely punctate meiotic recombination in the class II region of the major histocompatibility complex. Nat. Genet. 29: 217–222. [PubMed]
  • Kelly, J. K., 1997. A test of neutrality based on interlocus associations. Genetics 146: 1197–1206. [PMC free article] [PubMed]
  • Kim, Y., and W. Stephan, 2002. Detecting a local signature of genetic hitchhiking along a recombining chromosome. Genetics 160: 765–777. [PMC free article] [PubMed]
  • Kong, A., D. F. Gudbjartsson, J. Sainz, G. M. Jonsdottir, S. A. Gudjonsson et al., 2002. A high-resolution recombination map of the human genome. Nat. Genet. 31: 241–247. [PubMed]
  • Kruglyak, L., 1999. Prospects for whole-genome linkage disequilibrium mapping of common disease genes. Nat. Genet. 22: 139–144. [PubMed]
  • Lewontin, R. C., 1964. Interaction of selection + linkage. I. General considerations—heterotic models. Genetics 49: 49–67. [PMC free article] [PubMed]
  • Livingstone, F. B., 1985. Frequencies of Hemoglobin Variants: Thalassemia, the Glucose-6-Phosphate Dehydrogenase Deficiency, G6PD Variants and Ovalocytosis in Human Populations. Oxford University Press, Oxford.
  • McVean, G. A. T., S. R. Myers, S. Hunt, P. Deloukas, D. R. Bentley et al., 2004. The fine-scale structure of recombination rate variation in the human genome. Science 304: 581–584. [PubMed]
  • Meyer, D., and G. Thomson, 2001. How selection shapes variation of the human major histocompatibility complex: a review. Ann. Hum. Genet. 65: 1–26. [PubMed]
  • Motulsky, A. G., 1961. Glucose-6-phosphate dehydrogenase haemolytic disease of the newborn, and malaria. Lancet 1: 1168.
  • Nei, M., and W.-H. Li, 1979. Mathematical model for studying genetic variation in terms of restriction endonucleases. Proc. Natl. Acad. Sci. USA 76: 5269–5273. [PMC free article] [PubMed]
  • Ohashi, J., I. Naka, J. Patarapotikul, H. Hananantachai, G. Brittenham et al., 2004. Extended linkage disequilibrium surrounding the hemoglobin E variant due to malarial selection. Am. J. Hum. Genet. 74: 1198–1208. [PMC free article] [PubMed]
  • Phillips, M. S., R. Lawrence, R. Sachidanandam, A. P. Morris, D. J. Balding et al., 2003. Chromosome-wide distribution of haplotype blocks and the role of recombination hot spots. Nat. Genet. 33: 382–387. [PubMed]
  • Reich, D. E., M. Cargill, S. Bolk, J. Ireland, P. C. Sabeti et al., 2001. Linkage disequilibrium in the human genome. Nature 411: 199–204. [PubMed]
  • Rozas, J., J. C. Sanchez-DelBarrio, X. Messeguer and R. Rozas, 2003. DnaSP, DNA polymorphism analyses by the coalescent and other methods. Bioinformatics 19: 2496–2497. [PubMed]
  • Ruwende, C., S. C. Khoo, A. W. Snow, S. N. R. Yates, D. Kwiatkowski et al., 1995. Natural selection of hemizygotes and heterozygotes for G6PD deficiency in Africa by resistance to severe malaria. Nature 376: 246–249. [PubMed]
  • Sabeti, P. C., D. E. Reich, J. M. Higgins, H. Z. P. Levine, D. J. Richter et al., 2002. Detecting recent positive selection in the human genome from haplotype structure. Nature 419: 832–837. [PubMed]
  • Sanchez-Mazas, A., S. Djoulah, M. Busson, I. L. De Gouville, J. C. Poirier et al., 2000. A linkage disequilibrium map of the MHC region based on the analysis of 14 loci haplotypes in 50 French families. Eur. J. Hum. Genet. 8: 33–41. [PubMed]
  • Satta, Y., C. Ohuigin, N. Takahata and J. Klein, 1994. Intensity of natural-selection at the major histocompatibility complex loci. Proc. Natl. Acad. Sci. USA 91: 7184–7188. [PMC free article] [PubMed]
  • Saunders, M. A., M. F. Hammer and M. W. Nachman, 2002. Nucleotide variability at G6PD and the signature of malarial selection in humans. Genetics 162: 1849–1861. [PMC free article] [PubMed]
  • Schliekelman, P., C. Garner and M. Slatkin, 2001. Natural selection and resistance to HIV. Nature 411: 545–546. [PubMed]
  • Slatkin, M., 2001. Simulating genealogies of selected alleles in a population of variable size. Genet. Res. 78: 49–57. [PubMed]
  • Slatkin, M., and G. Bertorelle, 2001. The use of intra-allelic variability for testing neutrality and estimating population growth rate. Genetics 158: 865–874. [PMC free article] [PubMed]
  • Swallow, D. M., 2003. Genetics of lactase persistence and lactose intolerance. Annu. Rev. Genet. 37: 197–219. [PubMed]
  • Tajima, F., 1989. Statistical method for testing the neutral mutation hypothesis by DNA polymorphism. Genetics 123: 585–595. [PMC free article] [PubMed]
  • Takizawa, T., Y. Yoneyama, S. Miwa and A. Yoshida, 1987. A single nucleotide base transition is the basis of the common human glucose-6-phosphate dehydrogenase variant (A)+. Genomics 1: 228–231. [PubMed]
  • Tishkoff, S. A., E. Dietzsch, W. Speed, A. J. Pakstis, J. R. Kidd et al., 1996. Global patterns of linkage disequilibrium at the CD4 locus and modern human origins. Science 271: 1380–1387. [PubMed]
  • Tishkoff, S. A., R. Varkonyi, N. Cahinhinan, S. Abbes, G. Argyropoulos et al., 2001. Haplotype diversity and linkage disequilibrium at human G6PD: recent origin of alleles that confer malarial resistance. Science 293: 455–462. [PubMed]
  • Toomajian, C., and M. Kreitman, 2002. Sequence variation and haplotype structure at the human HFE locus. Genetics 161: 1609–1623. [PMC free article] [PubMed]
  • Toomajian, C., R. S. Ajioka, L. B. Jorde, J. P. Kushner and M. Kreitman, 2003. A method for detecting recent selection in the human genome from allele age estimates. Genetics 165: 287–297. [PMC free article] [PubMed]
  • Verrelli, B. C., J. H. McDonald, G. Argyropoulos, G. Destrol-Bisol, A. Froment et al., 2002. Evidence for balancing selection from nucleotide sequence analyses of human G6PD. Am. J. Hum. Genet. 71: 1112–1128. [PMC free article] [PubMed]
  • Wall, J. D., and J. K. Pritchard, 2003. Haplotype blocks and linkage disequilibrium in the human genome. Nat. Rev. Genet. 4: 587–597. [PubMed]
  • Walsh, E. C., K. A. Mather, S. F. Schaffner, L. Farwell, M. J. Daly et al., 2003. An integrated haplotype map of the human major histocompatibility complex. Am. J. Hum. Genet. 73: 580–590. [PMC free article] [PubMed]
  • Watterson, G. A., 1975. Number of segregating sites in genetic models without recombination. Theor. Popul. Biol. 7: 256–276. [PubMed]
  • Xu, W. M., B. Westwood, C. S. Bartsocas, J. J. Malcorraazpiazu, K. Indrak et al., 1995. Glucose-6-phosphate-dehydrogenase mutations and haplotypes in various ethnic groups. Blood 85: 257–263. [PubMed]

Articles from Genetics are provided here courtesy of Genetics Society of America
PubReader format: click here to try


Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...


Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...