PR

IFY

SG

OL

BA

NG

OR

/ B

AN

GO

R U

NIV

ER

SIT

Y

Genomic islands of speciation separate cichlid ecomorphs in an EastAfrican crater lakeMalinsky, M.; Challis, R.J.; Tyers, A.M.; Schiffels, S.; Terai, Y.; Ngatunga, B.P.;Miska, E.A.; Durbin, R.; Genner, M.J.; Turner, G.F.

Science

DOI:10.1126/science.aac9927

Published: 18/12/2015

Publisher's PDF, also known as Version of record

Cyswllt i'r cyhoeddiad / Link to publication

Dyfyniad o'r fersiwn a gyhoeddwyd / Citation for published version (APA):Malinsky, M., Challis, R. J., Tyers, A. M., Schiffels, S., Terai, Y., Ngatunga, B. P., Miska, E. A.,Durbin, R., Genner, M. J., & Turner, G. F. (2015). Genomic islands of speciation separate cichlidecomorphs in an East African crater lake. Science, 350(6267), 1493-1498.https://doi.org/10.1126/science.aac9927

Hawliau Cyffredinol / General rightsCopyright and moral rights for the publications made accessible in the public portal are retained by the authors and/orother copyright owners and it is a condition of accessing publications that users recognise and abide by the legalrequirements associated with these rights.

• Users may download and print one copy of any publication from the public portal for the purpose of privatestudy or research. • You may not further distribute the material or use it for any profit-making activity or commercial gain • You may freely distribute the URL identifying the publication in the public portal ?

Take down policyIf you believe that this document breaches copyright please contact us providing details, and we will remove access tothe work immediately and investigate your claim.

16. Nov. 2020

RESEARCH ARTICLE◥

GENOMIC EVOLUTION

Genomic islands of speciationseparate cichlid ecomorphs in an EastAfrican crater lakeMilan Malinsky,1,2† Richard J. Challis,3†‡ Alexandra M. Tyers,3 Stephan Schiffels,1

Yohey Terai,4 Benjamin P. Ngatunga,5 Eric A. Miska,1,2 Richard Durbin,1

Martin J. Genner,6* George F. Turner3*

The genomic causes and effects of divergent ecological selection during speciation arestill poorly understood. Here we report the discovery and detailed characterization ofearly-stage adaptive divergence of two cichlid fish ecomorphs in a small (700 meters indiameter) isolated crater lake in Tanzania. The ecomorphs differ in depth preference, malebreeding color, body shape, diet, and trophic morphology. With whole-genome sequencesof 146 fish, we identified 98 clearly demarcated genomic “islands” of high differentiationand demonstrated the association of genotypes across these islands with divergent matepreferences. The islands contain candidate adaptive genes enriched for functions insensory perception (including rhodopsin and other twilight-vision–associated genes),hormone signaling, and morphogenesis. Our study suggests mechanisms and genomicregions that may play a role in the closely related mega-radiation of Lake Malawi.

Understanding the causes and consequencesof speciation, including at the genetic level,requires the investigation of taxa at differ-ent stages on the speciation continuum(1, 2). East African cichlids have repeatedly

undergone rapid adaptive radiation (3). The LakeMalawi radiation has generated over 500 speciesin less than 5 million years, involving divergencein habitat, feeding apparatus, and nuptial color.Thus, these phenomena present an opportunityto observe hundreds of varied, recent, and insome cases ongoing, speciation events (4). How-ever, the investigation of early stages of specia-tion in the large cichlid radiations of Malawi aswell as Lakes Tanganyika and Victoria has beenhampered by difficulties in identifying sister spe-cies relationships, in reconstructing past geograph-ical situations, and in controlling for possibleintrogression from non-sister taxa (5).During 2011, we conducted a survey (table S1)

of fish fauna in six crater lakes in the RungweDistrict of Tanzania (Fig. 1A and table S2). In allsix lakes, we found endemic haplochromine cich-lids of the genus Astatotilapia, closely related toA. calliptera (Fig. 1A), a species widely distributed

in the rivers, streams, and shallow lakemargins ofthe region. Thus, theRungweDistrictAstatotilapiaare close relatives of the of Lake Malawi endemichaplochromine cichlids (5).In LakeMassoko (fig. S1), the benthic zone in

deep waters (~20 to 25 m) is very dimly lit andpopulated by cichlids with phenotypes clearlydifferent from those typical of shallow waters(≲5 m) close to the shore (littoral). Deep-watermales are dark blue-black, whereas most malescollected from the shallow waters are yellow-green, similar to riverine A. calliptera (Fig. 1B,movie S1, and table S3). We also collected small(<65 mm standard length) males that were notreadily field-assigned to either ecomorph (6). Thebenthic and littoral morphs are reminiscent ofthe species pair of Pundamilia cichlids fromLake Victoria (7), but within a potentially sim-pler historical and geographical context. LakeMassoko is steep-sided, has a strong thermo-cline at ~15 m, and an anoxic boundary at ~25 m(8). The estimated time of lake formation is~50,000 years ago (9).

Ecomorph separation

To examine relationships between crater lakeand riverine A. calliptera of southern Tanzania,we obtained restriction site–associated DNA(RAD) data from 30 fish from the Rungwe Dis-trict and 11 outgroup Astatotilapia from thebroader LakeMalawi catchment and aneighboringcatchment (fig. S2 and table S4). A maximum-likelihood phylogeny constructed on the basis ofthese data (6) demonstrates monophyly of allspecimens from Lake Massoko (Fig. 1A). Thus,the RAD phylogeny provides evidence thatMassoko morphs might have evolved in primary

sympatry, as proposed for crater lake cichlidradiations of Cameroon (10) and Nicaragua (11).Morphological analyses of these two colormorphs

revealed significant differences in head and bodyshape, body mass, the shape of pharyngeal teeth,and pharyngeal jawmass [Fig. 1, C to F, and tableS5; analysis of covariance (ANCOVA) tests, all P <0.001]. We also found significant differences instable isotope ratios (Fig. 1Gand table S5;ANCOVAtest, P < 0.001), which are indicative of dietarydifferences. Together these results demonstrateecomorph separation and adaptation to differentecological environments.

Whole-genome evidence

To study the genome-wide pattern of Massokoecomorph divergence and to further clarify itsgeographical context, we obtained whole-genomesequence data at ~15× coverage for 6 individualseach of the yellow littoral and blue benthic eco-morphs and 16 additional A. calliptera from thewider Lake Malawi catchment and two adjacentcatchments (fig. S2), supplemented by lower-coverage (~6×) data from 87 specimens from LakeMassoko (25 littoral, 32 benthic, and 30 smallunassigned) and 30 individuals from Lake Itamba(Fig. 1A and table S6). Sequence data were alignedto theMetriaclima zebra reference assembly (12),from which divergence was 0.2 to 0.3%, and var-iants were called at 4,755,448 sites (1.2 to 1.6millionsites per individual).A maximum-likelihood phylogeny built from

whole-genome sequence data confirmed recip-rocal monophyly of Astatotilapia within LakesMassoko and Itamba and revealed the sister groupof Massoko fish to be an A. calliptera populationfrom the nearby Mbaka River (Fig. 2A). All spe-cimens of the benthic ecomorph formed amono-phyletic clade derived from the littoral ecomorph(Fig. 2A). Principal components analysis (PCA)showed strong population structure (Tracy-Widomstatistics: P < 1 × 10−12), with benthic and littoralindividuals separated by the first eigenvector andforming separate clusters (Fig. 2B). In contrast,within Lake Itamba, PCA did not reveal signifi-cant population structure (Tracy-Widom statis-tics:P= 0.11). Individuals fromMassoko thatwerenot field-assigned to either of the ecomorphs didnot form a monophyletic clade in the phylogeny(Fig. 2A) or a distinct cluster in PCA (Fig. 2B).Analysis of fine-scale genetic relationships

with fineSTRUCTURE (13) supports the mono-phyly of the benthic ecomorph within the lit-toral, but also suggests that compared with thebenthic population, the littoral population hasgreater coancestry with other A. calliptera; inparticular with the Mbaka River sample (fig. S3).Therefore, we tested for evidence of secondarygene flow, as seen in cichlid populations fromCameroonian crater lakes (14). Under the nullhypothesis of no differential gene flow intoMassoko, A. calliptera from the Mbaka Rivershould share derived alleles equally often withthe littoral and with the benthic populations(15, 16). Instead, we found a small excess ofshared derived alleles between A. calliptera fromthe Mbaka River and the littoral population,

RESEARCH

SCIENCE sciencemag.org 18 DECEMBER 2015 • VOL 350 ISSUE 6267 1493

1Wellcome Trust Sanger Institute, Cambridge CB10 1SA, UK.2Gurdon Institute and Department of Genetics, University ofCambridge, Cambridge CB2 1QN, UK. 3School of BiologicalSciences, Bangor University, Bangor, Gwynedd LL57 2UW,UK. 4Department of Evolutionary Studies of Biosystems,SOKENDAI, Kanagawa 240-0193, Japan. 5Tanzania FisheriesResearch Institute, Box 9750, Dar es Salaam, Tanzania.6School of Biological Sciences, Life Sciences Building, 24Tyndall Avenue, University of Bristol, Bristol BS8 1TQ, UK.*Corresponding author. E-mail: george.turner@bangor.ac.uk(G.F.T.); m.genner@bristol.ac.uk (M.J.G.) †These authorscontributed equally to this work. ‡Present address: Institute ofEvolutionary Biology, University of Edinburgh, Charlotte AuerbachRoad, Edinburgh EH9 3FL, UK.

on

July

29,

201

6ht

tp://

scie

nce.

scie

ncem

ag.o

rg/

Dow

nloa

ded

from

when compared with the benthic population(Patterson’s D = 1.1%; 4.86 SD from 0% or P <5.8 × 10−7) (6). The proportion of admixture fwith Mbaka was estimated at 0.9 ± 0.2%. Thisvalue is low, at a proportion that is approxi-mately half of theNeanderthal introgression intonon-Africanhumans (15), andcross-coalescence rateanalysis with themultiple sequentiallyMarkoviancoalescent (MSMC) (6, 17) indicates an averageseparation time of bothMassoko ecomorphs fromother A. calliptera samples (including the MbakaRiver) approximately 10 times earlier than thesplit between the two ecomorphs (fig. S5). Thus,it is unlikely that a secondary invasion from theneighboring river systems (fig. S11B) contributedto the divergence of the ecomorphs.We estimated individual ancestries for all

Massoko and A. calliptera specimens withADMIXTURE (6, 18) (fig. S4). Focusing on theMassoko samples, 11 of the 31 samples field-assigned as littoral were identified as admixed,

with admixture fraction >25% from the benthicgene pool. No individuals identified as benthicwere estimated to be admixed to the same extent;therefore, recent gene flow may be biased fromdeep to shallowwaters. Ten of the 30 unassignedindividuals were also identified as >25% admixed,whereas the remaining 20 unassigned samplesappear to represent subadult individuals of bothbenthic and littoral ecomorphs (fig. S4A). WhenadditionalA. calliptera samples were included inADMIXTURE analysis, a small amount of geneflow into Massoko was apparent with K = 2 an-cestral populations (fig. S4C), consistent with thefineSTRUCTURE and Patterson’s D results de-scribed above. This analysis also suggests similaror even stronger gene flow out of Massoko andinto the Mbaka River (fig. S4C).

Islands of speciation

Interestingly, there are no fixed differences be-tween Massoko benthic and littoral ecomorphs.

Genome-wide divergence FST is 0.038, and almosthalf (47.6%) of the variable sites have zero FST(table S7). Above the low background, a genome-wide FST profile shows clearly demarcated “islands”of high differentiation (Fig. 2, C and E). For sin-gle sites, the maximum FST is 13.6 SD above themean, and 7543 sites have FST over 6 SD abovethe mean. In contrast, comparisons of the com-bined Massoko population and Itamba popula-tion revealed a pattern of consistently high FSTacross the genome (Fig. 2D) and large variance,making it impossible to detect statistical outliers(Fig. 2E). Similar results were obtained whenvarying the window size to include 15, 50, 100,or 500 variants (table S7 and figs. S6 and S7).Comparing observed levels of divergence with

neutral coalescent simulations (6) under a rangeof possible demographic models revealed that thetop 1% of observed FST values (approximately FST ≥0.25) are always higher than the correspondingneutral FST values from simulations, consistent

1494 18 DECEMBER 2015 • VOL 350 ISSUE 6267 sciencemag.org SCIENCE

Fig. 1. Cichlid radiation in the crater lakes of southern Tanzania. (A) Aphylogeny of the crater lake Astatotilapia based on reference-aligned RADdata (7906 SNPs across 5010 polymorphic RAD loci). It demonstrates re-ciprocal monophyly between the populations in each lake except for Itamba,and a close relationship to A. calliptera from rivers and from Lake Malawi.Within Lake Massoko, yellow symbols indicate the littoral morph, blue sym-bols indicate the benthic morph, and gray symbols denote small, pheno-typically ambiguous, and thus unassigned individuals. Additional Astatotilapiaindividuals from other crater lakes are denoted by open circles. A. calliptera from

rivers and Lake Malawi are denoted by black circles. Bootstrap values are dis-played for nodes with >50% support. (B) Breeding males of the yellow littoraland blue benthic morphs of Lake Massoko. The symbols next to the photo-graphs correspond to symbols used in (C) to (G). (C to F) Morphological diver-gence between the two morphs of Lake Massoko. Relative to the littoral morph,the benthic morph has relatively longer head and jaw (C), lower body mass (D),narrower “papilliform” pharyngeal teeth (E), and lighter lower pharyngeal jaws(F). The benthic fish have stable isotope ratios that tend to be more depletedin C13 than the littoral fish, indicative of a more offshore-planktonic diet (G).

RESEARCH | RESEARCH ARTICLE

on

July

29,

201

6ht

tp://

scie

nce.

scie

ncem

ag.o

rg/

Dow

nloa

ded

from

with divergent selection acting on approximatelythis top 1% of variant sites (figs. S8 and S9).We identified genomic regions with observed

benthic-littoral FST ≥ 0.25 (i.e., with FST abovemaximum levels seen in neutral simulations) (6).For this we used windows of 15 variants each,providing a balance between fine genomic res-olution and reducing stochastic variation by av-eraging over variants. Small gaps arising frombrief dips of FST below the threshold were elim-inated by merging regions within 10 kb of oneanother. We found 344 such regions, with totallength of 8.1 Mb (~1% of the genome). Next, tofocus on the more significant outliers, we nar-rowed the list down to a set of 98 highly diver-ged regions (HDRs) for further characterization(table S8) by adding the requirement that at leastone 10-kb window must have reached FST ≥ 0.3.The HDRs varied in length from 4.4 kb to 285 kb(median, 36.1 kb), with a total length of 5.5 Mb.A key prediction of speciation with gene flow

models is that loci participating in speciationshould have both high relative divergence (FST)and high absolute sequence divergence (dXY)(19, 20). However, previous studies [examinedin (20)] revealed low dXY and low nucleotidediversity (p) in regions of high FST. In contrast,we found that dXY in Massoko is significantlyhigher in HDRs relative to the rest of the ge-nome (P < 2.2 × 10−16, two-tailed Mann-Whitneytest; Fig. 3A). In the benthic ecomorph, p inHDRsis not significantly different from the rest of thegenome (P = 0.34, two-tailed Mann-Whitney test;fig. S10A), and in the littoral ecomorph p in HDRsiselevated (P=5.47×10−6, two-tailedMann-Whitneytest; fig. S10B). Individually, 55HDRshavedXYabovethe 90th percentile of the genome-wide distri-bution. The convergence ofFST and dXY measuresin regions of normal p suggests that these 55“islands of speciation” (table S9) may have beeninvolved in reducing gene flow in sympatry andthereby directly causing speciation to progress.In contrast, loci involved in continuing localadaptation after the ecomorphs split sufficientlyto constitute two largely separate gene pools (orduring a period of allopatry) would be expectedto have elevated FST but not dXY (20).Another key prediction of speciation with

gene flowmodels is that loci causing speciationshould be located in relatively few linked clus-ters within the genome (2, 20, 21). Instead of alarge number of scattered islands, the theorypredicts a smaller number of clusters that growin size because of the “divergence hitchhiking”process. We tested this prediction using a re-cently generated linkage map (22) and foundthat at least 27 out of the 55 putative speciationislands are co-localized on five linkage groups(LGs), with 26 of them clustered within theirrespective LGs (Fig. 3B and table S9). These po-tential speciation clusters extended for approxi-mately 25 cM on LG5, 40 cM on LG7, 30 cM onLG12, 5 cM on LG20, and 45 cM on LG 23. Intotal, these regions account for under 7% of thegenome, suggesting that divergence hitchhikingmay play a role in shaping the observed patternof genomic differentiation.

Although genomic islands within these clus-ters are often separated only by a few hundredkilobases, FST divergence between HDRs gen-erally drops to background levels (Fig. 3D), withone exception on scaffold 88 ,where a broader“continent” of divergence has formed (fig. S11).

Further support for sympatric divergence

We next tested whether the HDRs correlatedwith the signal of gene flow into LakeMassoko,as identified using the sample from the nearbyMbaka River. Compared with the rest of the ge-nome, the HDRs do not have elevated values ofPatterson’sD (P = 0.22, two-tailedMann-Whitneytest; fig. S12C), nor elevated f statistics, whichwere recently proposed as an means by whichone could identify introgressed loci (6, 23) (P =0.08, two-tailed Mann-Whitney test; fig. S12D).These results suggest that introgression fromthe Mbaka River did not play a major role ingenerating the HDRs between the benthic andlittoral ecomorphswithin LakeMassoko (fig. S12),and strengthen the evidence that the ecomorphdivergence has taken place within the lake.

Divergent SNPs associated withmate choice

Many recently diverged taxa, particularly thosenot geographically isolated, show stronger pre-

mating isolation than postmating isolation(1, 24, 25). We carried out laboratory experimentsto test for reproductive isolation resulting fromdirect mate choice between the Massoko eco-morphs (6) (movie S2). Fifty Massoko femaleswere given a choice from 16males representingthe variety of male phenotypes. In parallel, wedesigned a single-nucleotide polymorphism (SNP)assay with 117 polymorphic sites representing44 HDRs identified from the first 12 genomessequenced (table S10).We genotyped a reference sample of 18 ben-

thic and 16 littoral males, demonstrating thatthe SNP assay can reliably separate the ecomorphsalong the first principal component (PC1) in PCA(Fig. 4A, top).We then genotyped all females andmales participating in the mate choice experi-ments (Fig. 4A, bottom) and calculated an aver-age of the PC1 distances between each femaleand the males she mated with during the experi-ment, as assayed bymicrosatellite paternity anal-ysis (6). Comparedwith expectation under randommating (6), females had a moderate, but signif-icant (P = 4.3 × 10−5, paired t test), preference formating with males more genetically similar tothemselves (i.e., closer to them along PC1) (Fig.4B), demonstrating direct association betweenHDRvariants andmate choice. Assortativematingby genotype was strong among females with

SCIENCE sciencemag.org 18 DECEMBER 2015 • VOL 350 ISSUE 6267 1495

Genomic location (scaffold)

Fst

Fst

|z-s

core

|

Fig. 2. Whole-genome sequence data. (A) A maximum-likelihood whole-genome phylogenetic tree.Black stars indicate nodes with 100%bootstrap support.The red star highlights the branch that separatesall the Massoko benthic samples from the rest of the phylogeny (benthic ecomorph individuals are mono-phyletic in 50%of bootstrap samples). (B) PCA of genetic variation within LakeMassoko. (C to E) Genome-widepattern of FSTdivergence inwindows of 15 variants each. Darker color indicates greater density of datapoints. (C) Divergence between benthic and littoral ecomorphs within Massoko. (D) Divergence betweencombinedMassoko and Itamba populations. (E) Absolute standard scores of Massoko-Itamba divergence(purple) overlaid on divergence between benthic and littoral ecomorphs (green).

RESEARCH | RESEARCH ARTICLE

on

July

29,

201

6ht

tp://

scie

nce.

scie

ncem

ag.o

rg/

Dow

nloa

ded

from

positive (littoral) PC1 scores (P = 5.9 × 10−9,paired t test), whereas no assortative mating wasdetected among females with negative (benthic)PC1 scores (Fig. 4B).Stronger mating discrimination by ancestral

populations as compared to derived ones has

been previously found in Drosophila and stick-lebacks, possibly because low population densityafter a founder event favors less choosy individ-uals (26). However, it is also possible that thebenthic ecomorph mates assortatively only in thedeep-water environment; given that our experi-

ments used wide-spectrum lighting characteristicof shallow water. Overall, the moderate assorta-tive mating suggests a role for sexual selectionin ecomorph divergence but does not indicatethat it is a primary force causing population-wide divergence.

Signals of adaptation

A reduced level of genetic polymorphism in onesubpopulation may be indicative of a recent se-lective sweep. Overall, the magnitude of differ-ence in nucleotide diversity (p) between benthicand littoral ecomorphs (pdiff) is significantly higherin the HDRs than in the rest of the genome (P <2.2 × 10−16, two-tailed Mann-Whitney test; fig.S13A) (6). Individually 46 HDRs have pdiff abovethe 95th percentile of the genome-wide distribu-tion and are likely to have been under recentpositive selection in one of the two ecomorphs.There is a significant overlap between HDRswith high dXY (putative speciation islands) andHDRs with high pdiff (putative recent selectivesweeps): 35 of 55 high-dXY islands also havehigh pdiff (fig. S13B; P = 3 × 10−5, hypergeometrictest). On the other hand, the 11 putative sweepsthat did not lead to elevated dXY are indicative ofadaptation not directly involved in reproductiveisolation. Reduced nucleotide diversity in highpdiff regions, indicative of selective sweeps, wassignificantly more prevalent in the benthic eco-morph (36 of 46; P < 1.6 × 10−4, two-tailed bi-nomial test; Fig. 3C, top left, and fig. S13C) (6),consistent with the benthic ecomorph being de-rived and undergoing more extensive adaptation.Nevertheless, there are also a few strong outliers,suggesting selective sweeps in the littoral eco-morph (Fig. 3C, bottom right).

Functions of adaptation

To explore the function of candidate adaptivegenes, we performed gene ontology (GO) enrich-ment analysis (6) on three sets: (i) genes in can-didate islands of speciation ±50 kb (enrichedterms in table S11); (ii) genes in all HDRs ±10 kb(table S12); (iii) genes in all HDRs ±50 kb (tableS13). Combining results of all three analyses in anetwork (Fig. 5A) connecting GO terms with highoverlap (i.e., they share many genes) revealedclear clusters of enriched terms related to (i)morphogenesis (e.g., cartilage and pharyngealsystem development or fin morphogenesis), con-sistent with morphological differentiation; (ii)sensory systems (e.g., photoreceptor cell differ-entiation), consistent with previous studies show-ing the role of cichlid vision in adaptation andspeciation (7, 27); and (iii) (steroid) hormonesignaling.We examined in more detail the functions of

candidate genes involved in photoreceptor func-tion (table S14) and two highly diverged allelesof the rhodopsin (rho) gene in Lake Massoko(alleles H4 and H5, separated by four aminoacid changes; FST = 0.39; fig. S14). Blue-shiftedrhodopsin absorption spectra are known to playa role in deep-water adaptation (27). Therefore,we expressed rhodopsins fromH4 andH5 allelesandreconstructed themwith 11-cis-retinal,measured

1496 18 DECEMBER 2015 • VOL 350 ISSUE 6267 sciencemag.org SCIENCE

Fig. 4. Mate choice trials. (A) PCA based on 117 genotyped SNPs. (Top) The first axis of variation(PC1) in PCA reliably separates benthic and littoral males in a reference sample. (Bottom) PC1 po-sitions of females (n = 50) andmales (n = 16) participating inmate- hoice trials. (B) Results: Each pointcompares the average of absolute PC1 distances between a female and the males she mated with(observed PC1 distance) and all males she could have mated with (expected PC1 distance). Points arecolored according to the PC1 score of the female. Females below and to the right of the dasheddiagonal line on average mate with males more like themselves in terms of PC1 score than would betrue if they mated at random.

Fig. 3. Islands of speciation between benthic and littoral ecomorphs. (A) Elevated dXY in HDRs.(B) Clustering of putative speciation islands on five linkage groups. (C) Nucleotide diversity (p) withinHDRs (red points) and outside HDRs (blue with shading corresponding to density). Each point cor-responds to a 10-kbwindow (therefore, theremay bemultiple points per HDR).Overall, 95%of observationslie between the two curves (y = x ± 4.1 × 10−4). Putative sweeps in the benthic ecomorph are in the top leftcorner, and putative sweeps in the littoral are in the bottom right corner. (D) Patterns of FST, dXY, and pdiff in aspeciation cluster on scaffold 15.

RESEARCH | RESEARCH ARTICLE

on

July

29,

201

6ht

tp://

scie

nce.

scie

ncem

ag.o

rg/

Dow

nloa

ded

from

their absorption spectra (6), and demonstratedthat theH5 allele, associatedwith the deep-waterbenthic ecomorph, has a blue-shifted absorptionspectrum (Fig. 5B). The retina-specific retinoldehydrogenase rdh5 (table S14) produces 11-cis-retinal, the visual pigment binding partner ofrhodopsin (28) and thus is likely to have a directrole in dark adaptation. Finally, a mouse orth-olog of rp1l1b affects photosensitivity and mor-phogenesis of the outer segment (OS) of rodphotoreceptor cells, locating to the axoneme ofthe OS and of the connecting cilia (29) (Fig. 5C).Together, these results suggest that divergentselection on rho, rdh5, and rp1l1bmay facilitatethe adaptation of scotopic (twilight) vision tothe darker conditions experienced by the benthicecomorph.

Comparisons to other systems

Overall, our results suggest a pair of incipientspecies undergoing divergence with gene flowwithin crater lake Massoko. Their overall levelof divergence (FST = 0.038) is low compared withbackground FST observed in other recent studiesof speciation with gene flow in Anopheles mos-quitoes (S andM forms; FST = 0.21) (20), Ficedula

flycatchers (FST = 0.36) (30), and Heliconiusbutterflies (FST = 0.18) (31), highlighting thatwe are looking at an early stage of divergence.The MSMC analysis suggests that median ef-fective divergence occurred within the past 500to 1000 years (~200 to 350 generations), afterthe separation of lake fish from the Mbaka Riverpopulation around 10,000 years ago (fig. S5). How-ever, divergence may have started considerablyearlier than these times and be masked by sub-sequent gene flow.Among populations at similar levels of diver-

gence to that of the Lake Massoko ecomorphsare Timema stick insects (FST = 0.015 for adja-cent and FST = 0.03 for geographically isolatedpopulation pairs), in which thousands of re-gions of moderately elevated divergence werefound all across the genome (32), and Germancarrion and Swedish hooded crows (FST = 0.017),which have strongly diverged with fixed differ-ences, but at fewer than five loci (33). InMassoko,we observe an intermediate pattern betweenthese two extremes, with a few dozenmoderatelyelevated islands clustering within the genome,indicating close linkage and no fixed differences.A genome-wide patternwithmultiple loci ofmod-

erate divergence suggests a genomic architecturesimilar to the ecological divergence of a sympat-ric threespine stickleback pair in Paxton Lake,Canada (34), and the sympatric divergence ofdune-specialist sunflowers, Helianthus (35).The ecomorphs of Lake Massoko show clear

differences in traits normally associated withadaptive radiation in cichlid fishes, including bodyshape, pharyngeal jaw morphology, diet, micro-habitat preference, retinal pigment sensitivity,male color, and mate preference (3, 4, 11, 12, 27).Therefore, our study suggests processes and spe-cific genomic regions to investigate to determinewhether they are involved in speciation eventswithin the great cichlid radiations of LakesMalawi,Victoria, and Tanganyika.

REFERENCES AND NOTES

1. J. A. Coyne, H. A. Orr, Speciation (Sinauer, Sunderland, MA, 2004).2. J. L. Feder, S. P. Egan, P. Nosil, Trends Genet. 28, 342–350

(2012).3. C. E. Wagner, L. J. Harmon, O. Seehausen, Nature 487,

366–369 (2012).4. T. D. Kocher, Nat. Rev. Genet. 5, 288–298 (2004).5. D. A. Joyce et al., Curr. Biol. 21, R108–R109 (2011).6. Materials and methods are available as supplementary

materials on Science Online.7. O. Seehausen et al., Nature 455, 620–626 (2008).

SCIENCE sciencemag.org 18 DECEMBER 2015 • VOL 350 ISSUE 6267 1497

Fig. 5. Characterizing function ofgenes in HDRs. (A) Enrichment mapfor significantly enriched GO terms.The level of overlap between GOenriched terms is indicated by thethickness of the edge between them.The size of the node indicates thebest P value for the term, and thecolor of the node indicates the genegroup for which the term was foundsignificant (i.e., has P < 0.05 incandidate speciation islands ±50 kb,blue; in all HDRs ±10 kb or ±50 kb,red; or in both groups, green). Broadfunctional groupings (morphogenesis,sensory systems, etc.) were derivedusing automatic clustering followedby manual editing. (B) The absorp-tion spectrum of the H5 allele ofrho, more prevalent in the benthicecomorph, is shifted toward bluewavelengths. (C) The joint roles ofrho, rdh5, and rp1l1b in photoreceptorrod cells. rdh5 produces thechromophore 11-cis-retinal thatbinds rho, whereas rp1l1b, locatedat the axoneme of the outer segmentand connecting cilia, also contributesto photosensitivity.

RESEARCH | RESEARCH ARTICLE

on

July

29,

201

6ht

tp://

scie

nce.

scie

ncem

ag.o

rg/

Dow

nloa

ded

from

8. M. Delalande, Hydrologie et géochimie isotopique du lacMasoko et de lacs volcaniques de la province active du Rungwe(Sud-Ouest Tanzanie), www.theses.fr (2008).

9. P. Barker, D. Williamson, F. Gasse, E. Gibert, Q Res 60,368–376 (2003).

10. U. K. Schliewen, D. Tautz, S. Pääbo, Nature 368, 629–632 (1994).11. M. Barluenga, K. N. Stölting, W. Salzburger, M. Muschick,

A. Meyer, Nature 439, 719–723 (2006).12. D. Brawand et al., Nature 513, 375–381 (2014).13. D. J. Lawson, G. Hellenthal, S. Myers, D. Falush, PLOS Genet. 8,

e1002453 (2012).14. C. H. Martin et al., Evolution 69, 1406–1422 (2015).15. R. E. Green et al., Science 328, 710–722 (2010).16. E. Y. Durand, N. Patterson, D. Reich, M. Slatkin, Mol. Biol. Evol.

28, 2239–2252 (2011).17. S. Schiffels, R. Durbin, Nat. Genet. 46, 919–925 (2014).18. D. H. Alexander, J. Novembre, K. Lange, Genome Res. 19,

1655–1664 (2009).19. M. A. F. Noor, S. M. Bennett, Heredity 103, 439–444 (2009).20. T. E. Cruickshank, M. W. Hahn, Mol. Ecol. 23, 3133–3157 (2014).21. S. Via, Philos. Trans. R. Soc. London Ser. B 367, 451–460 (2012).22. C. T. O’Quin, A. C. Drilea, M. A. Conte, T. D. Kocher,

BMC Genomics 14, 287 (2013).23. S. H. Martin, J. W. Davey, C. D. Jiggins, Mol. Biol. Evol. 32,

244–257 (2015).24. T. D. Price, M. M. Bouvier, Evolution 56, 2083–2089 (2002).25. R. B. Stelkens, K. A. Young, O. Seehausen, Evolution 64,

617–633 (2010).26. K. Y. Kaneshiro, Evolution 34, 437 (1980).27. T. Sugawara et al., Proc. Natl. Acad. Sci. U.S.A. 102,

5448–5453 (2005).28. G. Duester, Eur. J. Biochem. 267, 4315–4324 (2000).29. T. Yamashita et al., J. Neurosci. 29, 9748–9760 (2009).30. H. Ellegren et al., Nature 491, 756–760 (2012).31. N. J. Nadeau et al., Philos. Trans. R. Soc. London Ser. B 367,

343–353 (2012).32. V. Soria-Carrasco et al., Science 344, 738–742 (2014).33. J. W. Poelstra et al., Science 344, 1410–1414 (2014).34. M. E. Arnegard et al., Nature 511, 307–311 (2014).35. R. L. Andrew, L. H. Rieseberg, Evolution 67, 2468–2482

(2013).

ACKNOWLEDGMENTS

This work was funded by Royal Society–Leverhulme Trust AfricaAwards AA100023 and AA130107 (M.J.G., B.P.N. and G.F.T.),Wellcome Trust Ph.D. studentship grant 097677/Z/11/Z (M.M.),Wellcome Trust grant WT098051 (S.S. and R.D.), Wellcome Trustand Cancer Research UK core support and a Wellcome TrustSenior Investigator Award (E.A.M.), Leverhulme Trust ResearchFellowship RF-2014-686 (M.J.G.), a University of Bristol ResearchCommittee award (M.G.), a Bangor University Anniversary Ph.D.studentship (A.M.T.), and a Fisheries Society of the British Islesaward (G.F.T.). Raw sequencing reads are in the Sequence ReadArchive: RAD sequencing (BioProject: PRJNA286304; accessionnos. SAMN03768857 to SAMN03768912) and whole-genomesequencing (BioProject PRJEB1254: sample accessions listed intable S16). The RAD-based phylogeny and alignments have beendeposited in TreeBase (TB2:S18241). Whole-genome variant callsin the Variant Call Format (VCF), phylogenetic trees, and primersequences for Sequenom genotyping are available from the DryadDigital Repository (http://dx.doi.org/10.5061/dryad.770mc).R.D. declares his interests as a founder and non-executive directorof Congenica Ltd., that he owns stock in Illumina from previousconsulting, and is a scientific advisory board member of DovetailInc. We thank R. Schley for generating pharyngeal jaw data;S. Mzighani, J. Kihedu, and the staff of the Tanzanian FisheriesResearch Institute for logistical support; A. Smith, H. Sungani,A. Shechonge, P. Parsons, J. Swanstrom, G. Cooke, and J. Bridlefor contributions to sampling and aquarium maintenance; theSanger Institute sequencing core for DNA sequencing; andH. Imai (Kyoto University) for the use of the spectrometer inhis laboratory.

SUPPLEMENTARY MATERIALS

www.sciencemag.org/content/350/6267/1493/suppl/DC1Materials and MethodsFigs. S1 to S16Tables S1 to S16References (36–67)Movies S1 and S2

11 July 2015; accepted 10 November 201510.1126/science.aac9927

REPORTS◥

QUANTUM SIMULATION

Connecting strongly correlatedsuperfluids by a quantumpoint contactDominik Husmann,1 Shun Uchino,2 Sebastian Krinner,1 Martin Lebrat,1

Thierry Giamarchi,2 Tilman Esslinger,1* Jean-Philippe Brantut1

Point contacts provide simple connections between macroscopic particle reservoirs. Inelectric circuits, strong links between metals, semiconductors, or superconductors haveapplications for fundamental condensed-matter physics as well as quantum informationprocessing. However, for complex, strongly correlated materials, links have been largelyrestricted to weak tunnel junctions. We studied resonantly interacting Fermi gasesconnected by a tunable, ballistic quantum point contact, finding a nonlinear current-biasrelation. At low temperature, our observations agree quantitatively with a theoreticalmodel in which the current originates from multiple Andreev reflections. In a wide contactgeometry, the competition between superfluidity and thermally activated transport leadsto a conductance minimum. Our system offers a controllable platform for the study ofmesoscopic devices based on strongly interacting matter.

The effect of strong interactions between theconstituents of a quantum many-body sys-tem is at the origin of several challengingquestions in physics. Although the groundstates of strongly interacting systems are

increasingly better understood (1), the propertiesout of equilibrium and at finite temperature oftenremain puzzling because these are determined bythe excitations above the ground state. In labora-tory experiments, strongly interacting systems arefound in certain materials, as well as in quantumfluids and gases (1). In solid-state systems, a con-ceptually simple and clean approach to probenonequilibrium physics is provided by transportmeasurements through the well-defined geome-try of a quantum point contact (QPC) (2–4). Yet,the technical hurdles to realize a controlled QPC

between strongly correlated materials pose a bigchallenge. Ultracold atomic Fermi gases in thevicinity of a Feshbach resonance, in the so-calledunitary regime, provide an alternative route tostudy correlated systems (5). Superfluidity has beenestablished at low temperature (6), but the finite-temperature properties are only partially under-stood (7–10)—a situation similar to the field ofstrongly correlated materials.Recent progress in the manipulation of cold

atomic gases has enabled the realization of a

1498 18 DECEMBER 2015 • VOL 350 ISSUE 6267 sciencemag.org SCIENCE

1Institute for Quantum Electronics, EidgenössischeTechnische Hochschule (ETH) Zürich, CH–8093 Zürich,Switzerland. 2Department of Quantum Matter Physics,Université de Genève, CH-1211 Genève, Switzerland.*Corresponding author. E-mail: esslinger@phys.ethz.ch

Fig. 1. Concept of the experiment. (A) Schematics of the two atom reservoirs (green) connected by aQPC.Two repulsive beams (not shown) confine the center of the cloud in the x and z directions, and theattractive gate beam (red) propagates along z and tunes the density in the QPC. (B) Potential landscapein the plane z = 0. Close to the QPC, the attractive gate creates areas of high density (blue). (C) Theoreticalmodel for the QPC. Both sites of the QPC have a defined atom number, imposing chemical potentialsmL and mR and a pairing gap D. The transparency a is an energy-dependent function describing the trans-mission of single particles from one side to the other. (D) Transport via multiple Andreev reflections.Coherent tunneling of pairs allows for the creation and tunneling of a single particle excitation (pairbreaking), leading to a dc, even for mR – mL = Dm � D.

RESEARCH

on

July

29,

201

6ht

tp://

scie

nce.

scie

ncem

ag.o

rg/

Dow

nloa

ded

from

(6267), 1493-1498. [doi: 10.1126/science.aac9927]350Science 17, 2015) Richard Durbin, Martin J. Genner and George F. Turner (DecemberSchiffels, Yohey Terai, Benjamin P. Ngatunga, Eric A. Miska, Milan Malinsky, Richard J. Challis, Alexandra M. Tyers, StephanEast African crater lakeGenomic islands of speciation separate cichlid ecomorphs in an

 Editor's Summary

   

, this issue p. 1493Sciencespeciation.supporting the idea that genetic changes related to breeding preferences are the first to diverge duringpopulations. Within these islands, the authors found genes likely to be associated with mate choice, the two forms, major regions of genetic divergence, known as genomic islands, separate thein a small lake in Tanzania are diversifying relative to each other. Although there is gene flow between

investigated how two ecomorphs of cichlid fishet al.when they coexist in the same location. Malinsky It is not clear how populations diversify and new species form at the genomic level, especially

Cichlids diverge within a crater lake

This copy is for your personal, non-commercial use only.

Article Tools

http://science.sciencemag.org/content/350/6267/1493article tools: Visit the online version of this article to access the personalization and

Permissionshttp://www.sciencemag.org/about/permissions.dtlObtain information about reproducing this article:

is a registered trademark of AAAS. ScienceAdvancement of Science; all rights reserved. The title Avenue NW, Washington, DC 20005. Copyright 2016 by the American Association for thein December, by the American Association for the Advancement of Science, 1200 New York

(print ISSN 0036-8075; online ISSN 1095-9203) is published weekly, except the last weekScience

on

July

29,

201

6ht

tp://

scie

nce.

scie

ncem

ag.o

rg/

Dow

nloa

ded

from