Genetic evidence for a second domestication of barley (Hordeum vulgare) east of the Fertile Crescent

To identify geographic discontinuities in genetic diversity in wild barley, we used resequencing data from 18 loci and 25 accessions that included a total of 684 SNPs (Fig. 1 and Table 1). A genetic assignment algorithm (34) was applied to the data with K = 2–4 clusters, without using prior information on geographic origin. The model-based algorithm identifies up to K clusters (where K may be unknown), each of which is characterized by distinct allele frequencies at each locus. With K = 2 clusters, the sample is split between the eastern and western portions of the species range, with the transition between predominantly eastern and western assignment occurring among accessions from the northern Zagros Mountains. However, samples from the northeastern extreme of the range cluster partially with western samples. With K = 3 or 4, northeastern samples are differentiated from both western and other eastern samples. This observation is consistent with haplotype composition; northeastern samples carry many private alleles, or haplotype segments, not sampled elsewhere.

Comparison of the percentage of alleles shared among samples to great circle geographic distance also reflects a change in allelic composition between the eastern and western clusters. In SI Fig. 4A, samples are compared both within and between eastern and western clusters. A group of highly differentiated pairs of samples occurs at ≈1,100 km, between accessions from the Mediterranean coast and the Zagros region, reflecting a change in allelic composition (Fig. 1 and SI Fig. 4A). A very similar pattern is also evident in wild barley based on the seven loci also sequenced in barley landraces (SI Fig. 4B) (discussed below).

For STRUCTURE analysis, the data must be treated as unique alleles. Each of the loci was broken into alleles (haplotype segments) based on direct evidence of recombination (see Materials and Methods). Among the 61 haplotype segments from the 18 loci, there are 329 total alleles, 182 of which are private to either eastern of western wild barley. Of these alleles, 40 occur at a frequency of ≥22% within one of the two partitions of the sample. Eighteen of these alleles are private to the eastern cluster, and 22 are private to the western. If the species-wide frequency of an allele is ≥22%, there is >95% probability that the allele would be sampled in accessions from both the eastern and western cluster.

Levels of nucleotide sequence diversity among the eastern and western cluster are similar, with average θ_π for all sites over all 18 loci of 0.00579 for eastern wild barley and 0.00597 for western wild barley.

Taken together, these results suggest that the clusters of wild samples identified in STRUCTURE analysis result from long-standing differences in allelic composition in populations east and west of the Zagros Mountains rather than recent demographic events (e.g., a bottleneck associated with recent founding of eastern populations).

Given that the primary geographic structure in wild barley differentiates western wild barley (from the Fertile Crescent) from eastern wild barley (from the Zagros and further east) we used STRUCTURE to estimate the probability of assignment for each cultivated barley accession to either of these two regions. We found that landrace accessions from east or west of the Zagros have very different probabilities of assignment to the western wild barley cluster (from the Fertile Crescent region). The median probability of assignment for western landraces is 99.4%, with a range of 62.2–99.8% (Fig. 2 and SI Fig. 5). Eastern landraces have a median probability of assignment to the western cluster of only 33.6%, with a range from 0.2% to 78.8%. Three-quarters of eastern landraces have <50% assignment to the western wild barley cluster. Accessions from the Zagros Mountains and Caspian Sea region show the lowest probability of origin in the Fertile Crescent (Fig. 2 and SI Fig. 5). Changes in specification of the data model in STRUCTURE (see Materials and Methods) do not change the primary assignment of individual landrace accessions but do result in less discreet clustering.

As was the case in wild barley, comparison of the percentage of shared alleles with geographic distance within versus between eastern and western landraces indicate differentiation among geographic regions (SI Fig. 6).

At six of the seven loci sequenced in the landrace samples (all but Dhn4), haplotypes that were found exclusively in eastern wild samples also were found to be present in eastern landraces (see SI Fig. 7). The haplotypes include 10 SNPs found only in eastern wild barley and eastern landraces.

These lines of evidence strongly suggest an independent domestication of barley outside the Fertile Crescent. All landrace accessions from east of the Zagros, across Asia to North Korea, have substantial identity to eastern wild barleys (Fig. 2 and SI Fig. 5), with assignment probabilities suggesting a mix of eastern and western origin. These data suggest that eastern landraces have been subject to admixture from imported, western landraces. Archaeological evidence indicates extensive introduction of other Fertile Crescent founder crops into central and southern Asia; for example, einkorn wheat was introduced by ≈6,000 B.C. (4, 5, 11). Thus, it would be surprising if western barley landraces had not been imported along with other domesticates. Introgression with western domesticated barley is likely to have been ongoing; Ordon et al. (19) report high levels of similarity between Japanese barley bred for malting quality and European cultivars.

Which geographic regions have contributed to modern cultivars? To address this question we considered 10 modern cultivars and three barley genetic stocks from North America and Europe. The cultivars have a median 89.7% probability of assignment to the western wild cluster, consistent with the introduction of barley into Europe and ultimately North America from the Fertile Crescent region (Fig. 3) (35). However, two U.S. cultivars and one line of the genetic stocks have <50% probability of assignment to the western cluster, probably because modern barley breeding programs have introduced genetic material from eastern wild barley in the quest to exploit novel germplasm (36). More generally, assignment tests indicate that wild barley from the Fertile Crescent contributed the majority of genetic diversity in present-day European and American cultivars, whereas wild barley from Central Asia contributed half or more of the genetic diversity in barley cultivated from Central Asia to the Far East.

Where could a second barley domestication have occurred? The relatively broad, species-wide sample mesh in the present study suggests an origin of eastern landraces in the western foothills of the Zagros or points farther east. Much of the region immediately east of the Zagros is a high-elevation plateau, where both wild barley populations (4) and known human Neolithic sites are relatively rare (5). However, the locations of early Neolithic agropastoral settlements suggest three general regions in which the secondary domestication could have taken place. In the foothills of the Zagros, at such sites as Ali Kosh and Jarmo (Fig. 1), domesticated barley is dated to ≈7,000–8,000 cal. B.C. Domesticated barley is found at the Indus Valley site of Mehrgarh (in present day Pakistan) from ≈7,000 cal. B.C. Finally, in the piedmont zone between the Kopet Dag mountain range and Kara Kum Desert (east of the Caspian Sea in present day Turkmenistan), cultivated barley was present by ≈6,000 cal. B.C. (see ref. 37). Both naked and hulled six-row barleys were cultivated at Mehrgarh and by the Jeitun culture (in the Caspian Sea region) (5). Mehrgarh lies near the eastern edge of the range of wild barley (1); thus, it is possible that barley found at Mehrgarh was domesticated locally. At Jeitun in southern Turkmenistan (the type site of the Jeitun Culture on the Kopet Dag piedmont), domesticated forms of (probably six-row) barley and einkorn wheat (Triticum monococcum) were being cultivated, and domesticated goats and sheep were being herded, by 6,000 cal. B.C., indicating the presence of a well developed agrarian society (5, 37).

Sample size in the present study limits our ability to compare haplotype composition in various portions of the eastern range of wild barley with that of eastern landraces. However, among the haplotype segments (from six loci) most indicative of eastern origin, i.e., those shared by eastern landraces and eastern wild barley but not sampled in western wild barleys, all but one were found in wild barleys from east of the Caspian Sea, and five segments were exclusive to that region; only one of the 11 segments was sampled only in the western Zagros. Based on a combination of archaeological and genetic data, the most likely location for a second origin of barley is ≈1,500–3,000 km east of sites in the Fertile Crescent where western barley is most likely to have been domesticated.

Evidence for two domestications must be weighed against the alternative, a single domestication in the Fertile Crescent followed by extensive introgression (14). Introgression from Central Asian wild barley into landraces imported from the Fertile Crescent would be necessary because wild barley samples from the Fertile Crescent do not carry all of the haplotype diversity found in eastern landraces. Expanding agrarian cultures could have imported landraces from the Fertile Crescent to Central Asia (5), where introgression with wild barley occurred (14).

There are two principal problems with this scenario (a single domestication followed by introgression). First, in what was perhaps 2,000 or more years before agrarian culture expanded into Central Asia, cultivated barley was subject to human selection for agronomically desirable polygenic traits, such as seed size and loss of seed dormancy (12, 38). After hybridization between wild and landrace barley, repeated back-crossing to the recurrent parent (imported landraces) would be necessary to recover agronomically important traits, diminishing the potential genetic contribution of the donor parent (eastern wild barley). The second issue is that a single domestication cannot explain the two independent origins of the domestication-related traits, nonbrittle ears and kernel row type. As Zohary (13) points out, the fixation of independent mutations at nonallelic, nonbrittle ear loci in cultivated barley is strongly suggestive of at least two domestications. With the addition of multilocus haplotype data demonstrating a strong geographic discontinuity in the probable wild founder populations of barley landraces, both of Zohary's criteria for identifying multiple domestications are satisfied.

Modern analytical methods, combined with high-throughput DNA sequencing at multiple informative loci permit a clearer view of historical events associated with domestication of a major founder crop and reveal at least two initial domestications at the dawn of agriculture. Because both wild and domesticated barleys maintain high levels of informative nucleotide sequence diversity (31), it is probable that more intensive sampling from an appropriate geographic mesh can provide a finer scale view of the history of domestication.

We thank S. Hegde, B. J. Steffenson, and G. Waines for helpful discussion and J. C. Avise, A. H. D. Brown, M. Brandt, B. S. Gaut, D. R. Harris, P. M. Hayes, and N. A. Rosenberg for critical reading of the manuscript. This work was supported by National Science Foundation Grant DEB-0129247.