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The evolution of genes for pigmentation in African cichlid fishes.

The cichlid fishes in the East African Great Lakes exhibit a great diversity of color patterns, presumably as adaptations to species-specific habitats and/or due to the action of sexual selection on color for species discrimination or female mate choice. To elucidate the mechanisms underlying such pigment pattern diversity, we cloned the cichlid homologs of tyrosinase, endothelin receptor b1, mitf, and Aim1 that previously had been cloned and characterized from pigmentation mutants of zebrafish and medaka fish. Gene sequence analysis among five cichlid species from the Great Lakes shows that the evolutionary rate of amino acid replacement in mitf is the highest of these four genes. We then compared the mitf amino acid replacement rates between species from the lacustrine and tilapiine/steatocranus lineages, and between Lake Malawi- and Victoria-haplochromine cichlids and Lake Tanganyika Lamprologini. We show that the evolutionary rate within the lacustrine lineage is twice that of the tilapiine/steatocranus lineage, but that rates for the Malawi-Victoria haplochromine and Lamprologini lineages are almost the same. These results suggest that the accelerated evolution of mitf might have occurred concomitantly with pigment pattern diversification in Great Lakes species, but not necessarily correlated with species under intense sexual selection on male mating color via female mate choice. Finally, we characterized a novel alternatively spliced variant of cichlid mitf that is similar to a mammalian mitf splice variant generated using alternate splice sites. We suggest that this new variant in cichlids, like that in mammals, encodes an MITF transcriptional factor having higher relative DNA binding affinity. These data provide a novel example of functional convergence in which a particular splice variant is independently generated via alternative splicing of a specific gene in different lineages.

Alternative Splicing↗

Genetic and developmental basis of cichlid trophic diversity.

Cichlids have undergone extensive evolutionary modifications of their feeding apparatus, making them an ideal model to study the factors that underlie craniofacial diversity. Recent studies have provided critical insights into the molecular mechanisms that have contributed to the origin and maintenance of cichlid trophic diversity. We review this body of work, which shows that the cichlid jaw is regulated by a few genes of major additive effect, and is composed of modules that have evolved under strong divergent selection. Adaptive variation in cichlid jaw shape is evident early in development and is associated with allelic variation in and expression of bmp4. Modulating this growth factor in the experimentally tractable zebrafish model reproduces natural variation in cichlid jaw shape, supporting a role for bmp4 in craniofacial evolution. These data demonstrate the utility of the cichlid jaw as a model for studying the genetic and developmental basis of evolutionary changes in craniofacial morphology.

Animals↗

Function of a key morphological innovation: fusion of the cichlid pharyngeal jaw.

The pharyngeal jaw of cichlids may represent a key innovation that facilitated their unparalleled trophic divergence. In cichlids, 'fusion' of the lower pharyngeal jaw (LPJ) results from suturing between the two lower ceratobranchials. To examine, what novel abilities a more extensively fused pharyngeal jaw may confer, the function of LPJ suturing was examined in Heroine cichlids. Greater LPJ suturing, pharyngeal jaw splitting under compression and the forces used to crush molluscs in the wild suggest increased LPJ fusion in the trophically polymorphic Herichthys minckleyi operates to strengthen the pharyngeal jaw. Among Heroine cichlid species, the presence of an external LPJ suture and feeding specialization on molluscs was evolutionarily quite variable, but greater LPJ fusion estimated from the amount of external suturing was highly correlated with molluscivory. Throughout cichlid diversification, increased pharyngeal jaw fusion via suturing has likely helped to reinforce the LPJ during pharyngeal processing thereby facilitating the ability of cichlids to exploit durable prey.

Animals↗

Phylogenetic utility of the tyrosine kinase gene X-src for assessing relationships among representative cichlid fishes.

The nuclear gene X-src is a member of the tyrosine-kinase class of proto-oncogenes whose normal product is localized within the cytoplasm of the cell. The X-src gene has been used in only a few phylogenetic studies, each focusing on systematics of killifishes (Cyprinodontiformes). The present study is an attempt to examine the phylogenetic utility of X-src for uncovering relationships of representative cichlid fishes, especially the cichlids of Middle America. The family Cichlidae is a species-rich group of tropical freshwater fish made up of more than 1000 species which show a Gondwanan pattern of distribution. Cichlid fish have been the focus of numerous studies ranging from behavioral to biogeographical to systematic in nature. Particular emphasis has focused on the cichlids of the African Great Lakes and the explosive adaptive radiation of this group. However, Neotropical cichlids have received considerably less attention than their African counterparts. Our findings regarding the utility of X-src concur with those of previous phylogenetic analyses showing the exons of X-src to be highly conserved and useful mostly for revealing deep relationships among taxa. Like previous X-src studies, we also found the intron sequences of the gene to be variable in length and difficult to align across distantly related taxa but they provided useful information for resolving relationships among more closely related taxa. The X-src phylogeny supports the monophyly of Neotropical cichlids and cichlasomines groups A (=heroines) + B (=cichlasomines). A highly resolved tree is obtained within the heroines but little support is evident for most nodes based on the low number of unambiguous substitutions. The X-src gene is likely to be quite useful for resolving deep phylogenetic relationships such as those among major groups of actinopterygian fishes.

Animals↗

Mitochondrial phylogeny of the endemic mouthbrooding lineages of cichlid fishes from Lake Tanganyika in eastern Africa.

Of the three cichlid species flocks in eastern Africa, Lake Tanganyika harbors the oldest species assemblage, which is also the most diverse morphologically and behaviorally. For 12 species (20 individuals) of 12 genera of the tribe Ectodini, 852 bp from two segments (cytochrome b and control region) of the mitochondrial genome were sequenced. In addition, orthologous sequences were obtained from eight species (11 individuals) representing other mouthbrooding lineages from Lake Tanganyika. Comparisons of sequence divergences revealed that the single Tanganyikan tribe Ectodini appears to be approximately five times older than the whole Lake Malawi cichlid species flock, suggesting that the radiation of the Tanganyikan mouthbrooding lineages took place long before the species flocks of Lakes Malawi and Victoria evolved. Seven of nine surveyed tribes of Tanganyikan cichlids appear to be approximately equally divergent, and this seems to corroborate the hypothesis of a rapid simultaneous formation of lineages at an early stage in the history of the Lake Tanganyika species flock. The close genetic relationship between the endemic Tropheus lineage and a nonendemic "Haplochromine," Astatotilapia burtoni, indicates that members of the tribe Tropheini may be the sister group of the cichlid flocks of Lakes Malawi and Victoria. The phylogenetic analyses demonstrate the monophyly of the Ectodini and identify the Cyprichromini as their sister group among the Tanganyikan cichlids. Within the tribe Ectodini the molecular data suggest both a branching pattern different than that previously proposed and a subdivision of the Ectodini into four clades, instead of the two originally described. The previously postulated model of morphological transformations believed to be responsible for the drastically different types of ecological specialization found among the Ectodini might therefore be in need of reinterpretation. Characters immediately related to foraging and nutrition seem to be particularly prone to homoplasy, even among members of a single lineage of cichlid fishes.

Africa, Eastern↗

Phylogenetic analysis of cichlid fishes using nuclear DNA markers.

The recent explosive adaptive radiation of cichlids in the great lakes of Africa has attracted the attention of both morphologists and molecular biologists. To decipher the phylogenetic relationships among the various taxa within the family Cichlidae is a prerequisite for answering some fundamental questions about the nature of the speciation process. In the present study, we used the random amplification of polymorphic DNA (RAPD) technique to obtain sequence differences between selected cichlid species. We then designed specific primers based on these sequences and used them to amplify template DNA from a large number of species by the polymerase chain reaction (PCR). We sequenced the amplified products and searched the sequences for indels and shared substitutions. We identified a number of such characters at three loci--DXTU1, DXTU2, and DXTU3--and used them for phylogenetic and cladistic analysis of the relationships among the various cichlid groups. Our studies assign an outgroup position to Neotropical cichlids in relation to African cichlids, provide evidence for a sister-group relationship of tilapiines to the haplochromines, group Cyphotilapia frontosa with the lamprologines of Lake Tanganyika, place Astatoreochromis alluaudi to an outgroup position with respect to other haplochromines of Lakes Victoria and Malawi, and provide additional support for the monophyly of the remaining Lake Victoria haplochromines and the Lake Malawi haplochromines. The described approach holds great promise for further resolution of cichlid phylogeny.

Africa, Eastern↗

Incipient speciation in sympatric Nicaraguan crater lake cichlid fishes: sexual selection versus ecological diversification.

The growing body of empirical evidence for sympatric speciation has been complemented by recent theoretical treatments that have identified evolutionary conditions conducive to speciation in sympatry. The Neotropical Midas cichlid (Amphilophus citrinellum) fits both of the key characteristics of these models, with strong assortative mating on the basis of a colour polymorphism coupled with trophic and ecological differentiation derived from a polymorphism in their pharyngeal jaws. We used microsatellite markers and a 480 bp fragment of the mitochondrial DNA control region to study four polymorphic populations of the Midas cichlid from three crater lakes and one large lake in Nicaragua in an investigation of incipient sympatric speciation. All populations were strongly genetically differentiated on the basis of geography. We identified strong genetic separation based on colour polymorphism for populations from Lake Nicaragua and one crater lake (Lake Apoyo), but failed to find significant genetic structuring based on trophic differences and ecological niche separation in any of the four populations studied. These data support the idea that sexual selection through assortative mating contributes more strongly or earlier during speciation in sympatry than ecological separation in these cichlids. The long-term persistence of divergent cichlid ecotypes (as measured by the percentage sequence divergence between populations) in Central American crater lakes, despite a lack of fixed genetic differentiation, differs strikingly from the patterns of extremely rapid speciation in the cichlids in Africa, including its crater lakes. It is unclear whether extrinsic environmental factors or intrinsic biological differences, e.g. in the degree of phenotypic plasticity, promote different mechanisms and thereby rates of speciation of cichlid fishes from the Old and New Worlds.

Animals↗

Nucleotide sequence and genomic organization of cichlid fish minisatellites.

We have cloned, sequenced, and determined the genomic organization of minisatellites from the African cichlid fish, Oroechromis niloticus. We estimate that minisatellites related in sequence to the Jeffreys' core probes 33.6 and 33.15 occur approximately every 1000 kilobase pairs in the cichlid fish genome. Sequencing of three minisatellites revealed that the size of the monomer units of the tandem arrays ranged from 7 to 24 base pairs (bp). One minisatellite appeared to contain a higher ordered periodicity of 90-120 bp superimposed on the apparent 15 bp monomer repeat, indicating a particularly large unit of homogenization for a minisatellite array. Sequence heterogeneity of repeat units within tandem arrays varied considerably from one minisatellite to another. Hybridization of cloned minisatellites to genomic DNA of cichlid fishes generated, in most instances, multilocus fingerprint patterns, indicating that families of minisatellites related by sequence exist in the cichlid genome. Two minisatellite clones, however, generated polymorphic single locus fingerprints, suggesting that these loci are conserved in closely related African cichlids. The cross hybridization of these cichlid minisatellites within and between related taxa, as well as to unrelated fish species such as Atlantic salmon (Salmo salar), complement the human minisatellite sequences for the study of genetic relationships among individuals from a wide range of fish species.

Animals↗

Functional morphology of extreme jaw protrusion in Neotropical cichlids.

The New World cichlids Petenia splendida and Caquetaia spp. possess extraordinarily protrusible jaws. We investigated the feeding behavior of extreme (here defined as greater than 30% head length) and modest jaw-protruding Neotropical cichlids by comparing feeding kinematics, cranial morphology, and feeding performance. Digital high-speed video (500 fps) of P. splendida, C. spectabile, and Astronotus ocellatus feeding on live guppy prey was analyzed to generate kinematic and performance variables. All three cichlid taxa utilized cranial elevation, lower jaw depression, and rotation of the suspensorium to protrude the jaws during feeding experiments. Extreme anterior jaw protrusion in P. splendida and C. spectabile resulted from augmented lower jaw depression and anterior rotation of the suspensorium. Morphological comparisons among eight cichlid species revealed novel anterior and posterior points of flexion within the suspensorium of P. splendida and Caquetaia spp. The combination of anterior and posterior loosening within the suspensorium in P. splendida and Caquetaia spp. permitted considerable anterior rotation of the suspensorium and contributed to protrusion of the jaws. Petenia splendida and C. spectabile exhibited greater ram distance and higher ram velocities than did A. ocellatus, resulting primarily from increased jaw protrusion. Petenia splendida and C. spectabile exhibited lower suction feeding performance than A. ocellatus, as indicated by lower suction-induced prey movements and velocities. Thus, extreme jaw protrusion in these cichlids may represent an adaptation for capturing elusive prey by enhancing the ram velocity of the predator but does not enhance suction feeding performance.

Adaptation, Biological↗

Mix and match color vision: tuning spectral sensitivity by differential opsin gene expression in Lake Malawi cichlids.

Cichlid fish of the East African Rift Lakes are renowned for their diversity and offer a unique opportunity to study adaptive changes in the visual system in rapidly evolving species flocks. Since color plays a significant role in mate choice, differences in visual sensitivities could greatly influence and even drive speciation of cichlids. Lake Malawi cichlids inhabiting rock and sand habitats have significantly different cone spectral sensitivities. By combining microspectrophotometry (MSP) of isolated cones, sequencing of opsin genes, and spectral analysis of recombinant pigments, we have established the cone complements of four species of Malawi cichlids. MSP demonstrated that each of these species predominately expresses three cone pigments, although these differ between species to give three spectrally different cone complements. In addition, rare populations of spectrally distinct cones were found. In total, seven spectral classes were identified. This was confirmed by opsin gene sequencing, expression, and in vitro reconstitution. The genes represent the four major classes of cone opsin genes that diverged early in vertebrate evolution. All four species possess a long-wave-sensitive (LWS), three spectrally distinct green-sensitive (RH2), a blue-sensitive (SWS2A), a violet-sensitive (SWS2B), and an ultraviolet-sensitive (SWS1) opsin. However, African cichlids determine their spectral sensitivity by differential expression of primarily only three of the seven available cone opsin genes. Phylogenetic analysis suggests that all percomorph fish have similar potential.

Animals↗

Molecular phylogeny and biogeography of the Malagasy and South Asian cichlids (Teleostei: Perciformes: Cichlidae).

Phylogenetic relationships of the Malagasy and South Asian cichlids are investigated using nucleotide characters from two mitochondrial genes, a 544 bp region of the large ribosomal subunit (16S) and a 649 bp region of cytochrome c oxidase subunit I (COI). This is the first molecular analysis to include a thorough taxonomic sampling of all Malagasy-South Asian genera, and a near complete taxonomic inventory of all valid species. Parsimony analysis of the combined data set (1193 aligned nucleotide positions) results in a single, completely resolved phylogenetic hypothesis. Results of this analysis, and that based on more comprehensive taxonomic sampling across Cichlidae for 16S alone (554 bp for 73 taxa), indicate that the Malagasy cichlids are paraphyletic, whereas the Malagasy and South Asian cichlids comprise a monophyletic group. In both analyses, the African and Neotropical assemblages are monophyletic. The Malagasy-South Asian cichlids are not recovered as plesiomorphic members of the family in either analysis. Two major clades are recovered within the Malagasy-South Asian assemblage and given subfamilial rank, Etroplinae, comprising Paretroplus (Madagascar) and Etroplus (southern India and Sri Lanka), and Ptychochrominae, comprising Ptychochromis, Ptychochromoides, and Oxylapia, all endemic Malagasy genera. Placement of the endemic Malagasy genus Paratilapia is equivocal depending on the gene fragment(s) analyzed. Inter- and intrageneric relationships within Ptychochrominae and Etroplinae are presented and discussed. The hypothesis of relationships for Cichlidae based on nucleotide characters from 16S alone, arguably the most comprehensive and broadly sampled data set across all major geographic assemblages to date, is congruent with prevailing hypotheses regarding the sequence of Gondwanan fragmentation and a vicariance scenario to explain the current distribution of cichlid fishes.

Animals↗

Hybrid origin of a cichlid population in Lake Malawi: implications for genetic variation and species diversity.

The importance of species recognition to taxonomic diversity among Lake Malawi cichlids has been frequently discussed. Hybridization - the apparent breakdown of species recognition - has been observed sporadically among cichlids and has been viewed as both a constructive and a destructive force with respect to species diversity. Here we provide genetic evidence of a natural hybrid cichlid population with a unique colour phenotype and elevated levels of genetic variation. We discuss the potential evolutionary consequences of interspecific hybridization in Lake Malawi cichlids and propose that the role of hybridization in generating both genetic variability and species diversity of Lake Malawi cichlids warrants further consideration.

Animals↗

On the origin of Lake Malawi cichlid species: a population genetic analysis of divergence.

The cichlid fishes of Lake Malawi are famously diverse. However, phylogenetic and population genetic studies of their history have been difficult because of the great amount of genetic variation that is shared between species. We apply a recently developed method for fitting the "isolation with migration" divergence model to a data set of specially designed compound loci to develop portraits of cichlid species divergence. Outgroup sequences from a cichlid from Lake Tanganyika permit model parameter estimates in units of years and effective population sizes. Estimated speciation times range from 1,000 to 17,000 years for species in the genus Tropheops. These exceptionally recent dates suggest that Malawi cichlids as a group experience a very active and dynamic diversification process. Current effective population size estimates range form 2,000 to near 40,000, and to >120,000 for estimates of ancestral population sizes. It appears that very recent speciation and gene flow are among the reasons why it has been difficult to discern the phylogenetic history of Malawi cichlids.

Animals↗

Using SINEs to probe ancient explosive speciation: "hidden" radiation of African cichlids?

Cichlid fishes of the east African Great Lakes represent a paradigm of adaptive radiation. We conducted a phylogenetic analysis of cichlids including pan-African and west African species by using insertion patterns of short interspersed elements (SINEs) at orthologous loci. The monophyly of the east African cichlids was consistently supported by seven independent insertions of SINE sequences that are uniquely shared by these species. In addition, data from four other loci indicated that the genera Tilapia (pan-African) and Steatocranus (west African) are the closest relatives to east African cichlids. However, relationships among Tilapia, Steatocranus, and the east African clade were ambiguous because of incongruencies among topologies suggested by insertion patterns of SINEs at six other loci. One plausible explanation for this phenomenon is incomplete lineage sorting of alleles containing or missing a SINE insertion at these loci during ancestral speciation. Such incomplete sorting may have taken place earlier than 14 MYA, followed by random and stochastic fixation of the alleles in subsequent lineages. These observations prompted us to consider the possibility that cichlid speciation occurred at an accelerated rate during this period when the African Great Lakes did not exist. The SINE method could be useful for detecting ancient exclusive speciation events that tend to remain hidden during conventional sequence analyses because of accumulated point mutations.

Base Sequence↗

Phylogeography of Lake Malawi cichlids of the genus Pseudotropheus: significance of allopatric colour variation.

One of the most compelling features of the cichlid fishes of the African Great Lakes is the seemingly endless diversity of male coloration. Colour diversification has been implicated as an important factor driving cichlid speciation. Colour has also been central to cichlid taxonomy and, thus, to our concept of species diversity. We undertook a phylogeographical examination of several allopatric populations of the Lake Malawi cichlid Pseudotropheus zebra in order to reconstruct the evolutionary history of the populations, which exhibit one of two dorsal fin colours. We present evidence that populations with red dorsal fins (RT) are not monophyletic. The RT population defining the northern limit of the distribution has evidently originated independently of the southern RT populations, which share a common ancestry. This evidence of species-level colour convergence is an important discovery in our understanding of cichlid evolution. It implies that divergence in coloration may accompany speciation, and that allopatric populations with similar coloration cannot be assumed to be conspecific. In addition to this finding, we have observed evidence for introgression, contributing to current evidence that this phenomenon may be extremely widespread. Thus, in species-level phylogenetic reconstructions, including our own, consideration must be given to the potential effects of introgression.

Alleles↗

Construction of a BAC library for Haplochromis chilotes, a cichlid fish from Lake Victoria.

Cichlid fishes in Lake Victoria are model organisms for studying rapid radiation and speciation. On the way to examine the molecular basis of how these cichlid fishes achieved such a remarkable morphological diversification, we constructed a bacterial artificial chromosome (BAC) library derived from a cichlid species, Haplochromis chilotes, from Lake Victoria. The library includes 157,056 clones with the average insert size of 128 kb, corresponding to a 10-fold coverage of the H. chilotes genome. Given that the cichlid fishes endemic to Lake Victoria are closely related to one another phylogenetically and their genomes are nearly identical, this BAC library can be utilized to isolate genes from the more than 200 Haplochromine cichlid species in Lake Victoria.

Animals↗

Genetic architecture sets limits on transgressive segregation in hybrid cichlid fishes.

The role of hybridization in the evolution of animal species is poorly understood. Transgressive segregation is a mechanism through which hybridization can generate diversity and ultimately lead to speciation. In this report we investigated the capacity of hybridization to generate novel (transgressive) phenotypes in the taxonomically diverse cichlid fishes. We generated a large F2 hybrid population by crossing two closely related cichlid species from Lake Malawi in Africa with differently shaped heads. Our morphometric analysis focused on two traits with different selective histories. The cichlid lower jaw (mandible) has evolved in response to strong directional selection, and does not segregate beyond the parental phenotype. The cichlid neurocranium (skull) has likely diverged in response to forces other than consistent directional selection (e.g., stabilizing selection), and exhibits marked transgressive segregation in our F2 population. We show that the genetic architecture of the cichlid jaw limits transgression, whereas the genetic basis of skull shape is permissive of transgressive segregation. These data suggest that natural selection, acting through the genome, will limit the degree of diversity that may be achieved via hybridization. Results are discussed in the context of the broader question of how phenotypic diversity may be achieved in rapidly evolving systems.

Animals↗

Micro- and macroevolutionary decoupling of cichlid jaws: a test of Liem's key innovation hypothesis.

The extent to which elements of functional systems can change independently (modularity) likely influences the diversification of lineages. Major innovations in organismal design, like the pharyngeal jaw in cichlid fishes, may be key to a group's success when they relax constraints on diversification by increasing phenotypic modularity. In cichlid fishes, pharyngeal jaw modifications that enhanced the ability to breakdown prey may have freed their oral jaws from serving their ancestral dual role as a site of both prey capture and prey processing. This functional decoupling that allowed the oral jaws to become devoted solely to prey capture has been hypothesized to have permitted the two sets of cichlid jaws to evolve independently. We tested the hypothesis that oral and pharyngeal jaw mechanics are evolutionarily decoupled both within and among Neotropical Heroine cichlids. In the trophically polymorphic species Herichthys minckleyi, molariforms that exhibit enlarged molarlike pharyngeal jaw teeth were found to have approximately 400% greater lower jaw mass compared to H. minckleyi with the alternative papilliform pharyngeal morphology. However, oral jaw gape, lower jaw velocity ratios, anterior jaw linkage mechanics, and jaw protrusion did not differ between the morphotypes. In 40 other Heroine species, there was a weak correlation between oral jaw mechanics and pharyngeal jaw mass when phylogenetic history was ignored. Yet, after expansion of the cytochrome b phylogeny for Heroines, change in oral jaw mechanics was found to be independent of evolutionary change in pharyngeal jaw mass based on independent contrasts. Evolutionary decoupling of oral and pharyngeal jaw mechanics has likely played a critical role in the unparalleled trophic diversification of cichlid fishes.

Animals↗