O'Connell LA, Fontenot MR, Hofmann HA. Characterization of the dopaminergic system in the brain of an African cichlid fish, Astatotilapia burtoni. Journal of Comparative Neurology. 2011;519 (1) :75 - 92.Abstract

Catecholamines, such as dopamine, are evolutionarily ancient neurotransmitters that play an essential role in mediating behavior. In vertebrates, dopamine is central to the nigrostriatal motor and mesolimbic reward systems. Despite its importance, the distribution of the dopaminergic system has not been well studied in the teleost brain. The African cichlid fish Astatotilapia burtoni has become an important model system in social neuroscience and lends itself to uncovering how social decisions are implemented in the brain. To understand better where dopamine acts to regulate social behavior in this species, we have determined the distribution of putative dopaminergic cells and fibers (by tyrosine hydroxylase immunohistochemistry) and dopamine receptors (by in situ hybridization for the D(1A) and D(2) dopamine receptor subtypes) throughout the forebrain and part of the mesencephalon of A. burtoni. Tyrosine hydroxylase immunoreactivity was evident in several regions of the fore- and midbrain, in support of putative homologies to tetrapods. Additionally, the D(1A) and D(2) receptors were identified in brain regions known to modulate social behavior in other vertebrates, including the proposed teleost homologues of the mammalian amygdalar complex, hippocampus, striatum, preoptic area, anterior hypothalamus, ventromedial hypothalamus, and ventral tegmental area/substantia nigra pars compacta. Tyrosine hydroxylase-immunoreactive fibers as well as D(1A) and D(2) receptor expression overlap almost completely in their distribution. These results significantly extend our understanding of the distribution of the dopaminergic system in the teleost brain and suggest a conserved role of dopamine in modulating behavior across vertebrates.

Kline RJ, O'Connell LA, Hofmann HA, Holt JG, Khan IA. The distribution of an avt v1a receptor in the brain of a sex changing fish, epinephelus adscensionis. Journal of Chemical Neuroanatomy. 2011;42 (1) :72 - 88.Abstract

The present study describes the distribution of an arginine vasotocin (AVT) V1a receptor (AVTr) throughout the brain of a sex-changing grouper, rock hind Epinephelus adscensionis. The objectives of this study were to describe the AVTr distribution in the brain of rock hind for potential linkages of the AVT hormone system with sex-specific behaviors observed in this species and to examine sex-specific differences that might exist. An antibody was designed for rock hind AVTr against the deduced amino acid sequence for the third intracellular loop. Protein expression, identified with immunohistochemistry showed high concordance with mRNA expression, identified with in situ hybridization. AVTr protein and mRNA expression was widely distributed throughout the brain, indicating that AVT may act as a neuromodulator via this V1a receptor subtype. AVTr protein and mRNA were present in regions associated with behavior, reproduction and spatial learning, as well as sensory functions such as vision, olfaction and lateral line sensory processing. We observed high AVTr expression in granular cell formations in the internal cellular layer of olfactory bulbs, torus longitudinalis, granular layer of the corpus cerebellum, valvula of the cerebellum, nuclei of the lateral and posterior recesses, and granular eminence. High protein and mRNA expression was also observed in the preoptic area, anterior hypothalamus, and habenular nucleus. No obvious sex differences were noted in any region of the rock hind brain. ?? 2011 Elsevier B.V.

O'Connell LA, Hofmann HA. Genes, hormones, and circuits: An integrative approach to study the evolution of social behavior. Frontiers in Neuroendocrinology. 2011;32 (3) :320 - 335.Abstract

Tremendous progress has been made in our understanding of the ultimate and proximate mechanisms underlying social behavior, yet an integrative evolutionary analysis of its underpinnings has been difficult. In this review, we propose that modern genomic approaches can facilitate such studies by integrating four approaches to brain and behavior studies: (1) animals face many challenges and opportunities that are ecologically and socially equivalent across species; (2) they respond with species-specific, yet quantifiable and comparable approach and avoidance behaviors; (3) these behaviors in turn are regulated by gene modules and neurochemical codes; and (4) these behaviors are governed by brain circuits such as the mesolimbic reward system and the social behavior network. For each approach, we discuss genomic and other studies that have shed light on various aspects of social behavior and its underpinnings and suggest promising avenues for future research into the evolution of neuroethological systems. ?? 2010 Elsevier Inc.

O'Connell LA, Ding JH, Ryan MJ, Hofmann HA. Neural distribution of the nuclear progesterone receptor in the tungara frog, Physalaemus pustulosus. Journal of Chemical Neuroanatomy [Internet]. 2011;41 (3) :137 - 147. Publisher's VersionAbstract

The gonadal steroid hormone progesterone plays an important role across all vertebrates in mediating female reproductive physiology and behavior. Many effects of progesterone are mediated by a nuclear progesterone receptor (PR), which is crucial for integration of external signals and internal physiological cues in the brain to produce an appropriate behavioral output. The t??ngara frog, Physalaemus pustulosus, is an excellent model system for the study of mechanisms by which sensory signals, such as auditory communication, are processed within neural circuits where mate choice decisions are made. To establish a framework for studying the neural basis of mate choice and social behavior in this species, we first describe the cytoarchitecture of the brain using Nissl-stained sections. Then, in order to better understand where progesterone acts to regulate social decisions, we determined the distribution of PR protein throughout the brain of P. pustulosus by immunohistochemistry. We found PR immunoreactivity in key brain regions known to modulate the processing of auditory cues and social behavior in other vertebrates. Due to its widespread distribution, PR likely also plays important roles in non-limbic brain regions that mediate non-social information processing. Further, we have colocalized PR with tyrosine hydroxylase, providing a functional context for the role of progesterone in mediating motivation and motor behavior. Our results significantly extend our understanding of hormonal modulation in the anuran brain and support the important role of the nuclear progesterone receptor in modulating female mate choice and receptivity in amphibians and across vertebrates. ?? 2011 Elsevier B.V.

Oldfield RG, Hofmann HA. Neuropeptide regulation of social behavior in a monogamous cichlid fish. Physiology and Behavior. 2011;102 (3-4) :296 - 303.Abstract

The nonapeptides arginine vasopressin (AVP; including its non-mammalian homolog arginine vasotocin, AVT) and oxytocin (OT; including its non-mammalian homologs mesotocin, MT, and isotocin, IT) regulate social behavior, including aggression and reproduction, via receptors conserved across vertebrates. In monogamous prairie voles, the vasopressin and oxytocin pathways are crucially important for pair-bond formation, specifically by influencing affiliative behavior toward the mate and aggression toward non-mates. Monogamous social systems are found in diverse taxa. We hypothesized that the AVT/IT pathways are associated with mating behavior in monogamous teleost fishes. We used the monogamous convict cichlid, Amatitlania nigrofasciata, to test this idea. In the first experiment, we treated males with a general nonapeptide receptor antagonist during pair-bond formation. Control males were treated with vehicle. On the first day of treatment we observed a significant reduction in both affiliative behavior toward the potential mate and aggression toward neighbors. However, the antagonist did not prevent the pair-bond from forming and the behavioral effects disappeared on subsequent treatment days. In the second experiment, we administered on three consecutive days the AVP/OT receptor antagonist to males that were in an established pair-bond. In established pairs, male affiliation towards the mate and aggressive behavior towards territorial neighbors were not affected by the antagonist. Our results indicate that the basic social behaviors typically mediated by the AVP/OT pathways may provide the building blocks necessary for monogamous mating behavior. ?? 2010 Elsevier Inc.

Dijkstra PD, Wiegertjes GF, Forlenza M, van der Sluijs I, Hofmann HA, Metcalfe NB, Groothuis TGG. The role of physiology in the divergence of two incipient cichlid species. Journal of Evolutionary Biology. 2011;24 (12) :2639 - 2652.Abstract

Sexual selection on male coloration has been implicated in the evolution of colourful species flocks of East African cichlid fish. During adaptive radiations, animals diverge in multiple phenotypic traits, but the role of physiology has received limited attention. Here, we report how divergence in physiology may contribute to the stable coexistence of two hybridizing incipient species of cichlid fish from Lake Victoria. Males of Pundamilia nyererei (males are red) tend to defeat those of Pundamilia pundamilia (males are blue), yet the two sibling species coexist in nature. It has been suggested that red males bear a physiological cost that might offset their dominance advantage. We tested the hypothesis that the two species differ in oxidative stress levels and immune function and that this difference is correlated with differences in circulating steroid levels. We manipulated the social context and found red males experienced significantly higher oxidative stress levels than blue males, but only in a territorial context when colour and aggression are maximally expressed. Red males exhibited greater aggression levels and lower humoral immune response than blue males, but no detectable difference in steroid levels. Red males appear to trade off increased aggressiveness with physiological costs, contributing to the coexistence of the two species. Correlated divergence in colour, behaviour and physiology might be widespread in the dramatically diverse cichlid radiations in East African lakes and may play a crucial role in the remarkably rapid speciation of these fish.

Whitaker KW, Neumeister H, Huffman LS, Kidd CE, Preuss T, Hofmann HA. Serotonergic modulation of startle-escape plasticity in an African cichlid fish: a single-cell molecular and physiological analysis of a vital neural circuit. Journal of neurophysiology. 2011;106 (1) :127 - 137.Abstract

Social life affects brain function at all levels, including gene expression, neurochemical balance, and neural circuits. We have previously shown that in the cichlid fish Astatotilapia burtoni brightly colored, socially dominant (DOM) males face a trade-off between reproductive opportunities and increased predation risk. Compared with camouflaged subordinate (SUB) males, DOMs exposed to a loud sound pip display higher startle responsiveness and increased excitability of the Mauthner cell (M-cell) circuit that governs this behavior. Using behavioral tests, intracellular recordings, and single-cell molecular analysis, we show here that serotonin (5-HT) modulates this socially regulated plasticity via the 5-HT receptor subtype 2 (5-HTR(2)). Specifically, SUBs display increased sensitivity to pharmacological manipulation of 5-HTR(2) compared with DOMs in both startle-escape behavior and electrophysiological properties of the M-cell. Immunohistochemistry showed serotonergic varicosities around the M-cells, further suggesting that 5-HT impinges directly onto the startle-escape circuitry. To determine whether the effects of 5-HTR(2) are pre- or postsynaptic, and whether other 5-HTR subtypes are involved, we harvested the mRNA from single M-cells via cytoplasmic aspiration and found that 5-HTR subtypes 5A and 6 are expressed in the M-cell. 5-HTR(2), however, was absent, suggesting that it affects M-cell excitability through a presynaptic mechanism. These results are consistent with a role for 5-HT in modulating startle plasticity and increase our understanding of the neural and molecular basis of a trade-off between reproduction and predation.

O'Connell LA, Hofmann HA. The Vertebrate mesolimbic reward system and social behavior network: A comparative synthesis. Journal of Comparative Neurology. 2011;519 (18) :3599 - 3639.Abstract

All animals evaluate the salience of external stimuli and integrate them with internal physiological information into adaptive behavior. Natural and sexual selection impinge on these processes, yet our understanding of behavioral decision-making mechanisms and their evolution is still very limited. Insights from mammals indicate that two neural circuits are of crucial importance in this context: the social behavior network and the mesolimbic reward system. Here we review evidence from neurochemical, tract-tracing, developmental, and functional lesion/stimulation studies that delineates homology relationships for most of the nodes of these two circuits across the five major vertebrate lineages: mammals, birds, reptiles, amphibians, and teleost fish. We provide for the first time a comprehensive comparative analysis of the two neural circuits and conclude that they were already present in early vertebrates. We also propose that these circuits form a larger social decision-making (SDM) network that regulates adaptive behavior. Our synthesis thus provides an important foundation for understanding the evolution of the neural mechanisms underlying reward processing and behavioral regulation. J. Comp. Neurol. 519:3599–3639, 2011. © 2011 Wiley-Liss, Inc.

Wong RY, a Hoffmann H. Behavioural Genomics : An Organismic Perspective. Encyclopedia of Life science. 2010 :1 - 9.Abstract

The behavioural patterns observed in many organisms generally resultfromthe integration ofbothexternaland internal cues. Why do animals behave the way they do? The study of the proximate and ultimate mechanisms underlying animal behaviour tries to answer this ques- tion. Although various approaches have been developed for examining – often quantitatively and with increasing specificity and resolution – the roles genes play in the regulation of behaviour, until recently they were limited to individual candidate genes and often neglected ultim- ate mechanisms. Advances in genomic approaches in recent years have made it possible to examine gene expression patterns (in the brain and elsewhere) on a genomicscale even in nontraditional, yet ecologicallyand evolutionarily important model systems. As behavioural genomics begins to integrate proximate and ultimate mechanisms of animal behaviour, we may finally under- stand why animals behave the way they do.

Munchrath LA, Hofmann HA. Distribution of sex steroid hormone receptors in the brain of an african cichlid fish, astatotilapia burtoni. Journal of Comparative Neurology. 2010;518 (16) :3302 - 3326. 2010.munchrath.jcn_.pdf
Hofmann HA. Early developmental patterning sets the stage for brain evolution. Proceedings of the National Academy of Sciences of the United States of America. 2010;107 (22) :9919 - 9920. 2010.hofmann.pnas_.pdf
Kidd CE, Kidd MR, Hofmann HA. Measuring multiple hormones from a single water sample using enzyme immunoassays. General and Comparative Endocrinology [Internet]. 2010;165 (2) :277 - 285. Publisher's VersionAbstract

Many aquatic species, such as teleosts, release into the water and detect multiple bioactive substances to assist in schooling, migration, alarm reactions, and to stimulate behavioral and physiological responses during reproduction and in parent-offspring interactions. Understanding the complex relationship between hormones, behavior and their function in communication requires the simultaneous examination of multiple circulating hormones. However, repeated blood sampling within a short time period is not possible in smaller animals without impacting the very behaviors under investigation. The non-invasive technique of collecting and measuring hormone values in holding water using either radioimmunoassay (RIA) or enzyme immunoassay (EIA) is becoming widely used in teleost research. Commercial assay kits in particular enable rapid and reliable data generation, yet their assay buffers are often specific and potentially incompatible with each other, which can hinder measuring multiple hormones from the same sample. We present here the validation and application of a "nested" elution technique we developed that allows for repeated sampling of multiple reproductive hormones - testosterone (T), 17??-estradiol (E2), progesterone (P), prostaglandin F2?? (PGF) and 11-ketotestosterone (11KT) - from individual samples of animal holding water by using commercial EIA systems. Our results show that when using appropriate controls to account for possible technical and biological confounds, this technique provides a powerful new tool for research in aquatic endocrinology and physiology. ?? 2009 Elsevier Inc. All rights reserved.

Hofmann HA. The neuroendocrine action potential. Winner of the 2008 Frank Beach Award in Behavioral Neuroendocrinology. Hormones and Behavior [Internet]. 2010;58 (4) :555 - 562. Publisher's VersionAbstract

Animals are remarkably well equipped to respond to changes in their environment across different time scales and levels of biological organization. Here, I introduce a novel perspective that incorporates the three main processes the nervous system uses to integrate and process information: electrophysiological, genomic, and neuroendocrine action potentials. After discussing several examples of neuroendocrine action potentials, I lay out the commonalities of these temporally organized responses and how they might be interrelated with electrophysiological activity and genomic responses. This framework provides a novel outlook on longstanding questions in behavioral neuroendocrinology and suggests exciting new avenues for further research that will integrate across disciplines and levels of biological organization. ?? 2010 Elsevier Inc.

O'Quin KE, Hofmann CM, Hofmann HA, Carleton KL. Parallel Evolution of opsin gene expression in African cichlid fishes. Molecular Biology and Evolution. 2010;27 (12) :2839 - 2854.Abstract

Phenotypic evolution may occur either through alterations to the structure of protein-coding genes or their expression. Evidence for which of these two mechanisms more commonly contribute to the evolution of a phenotype can be garnered from examples of parallel and convergent evolution. The visual system of East African cichlid fishes is an excellent system with which to address this question. Cichlid fishes from Lakes Malawi (LM) and Victoria together exhibit three diverse palettes of coexpressed opsins and several important protein-coding mutations that both shift spectral sensitivity. Here we assess both opsin expression and protein-coding diversity among cichlids from a third rift lake, Lake Tanganyika (LT). We found that Tanganyikan cichlids exhibit three palettes of coexpressed opsins that largely overlap the short-, middle-, and long-wavelength-sensitive palettes of LM cichlids. Bayesian phenotypic clustering and ancestral state reconstructions both support the parallel evolution of the short- and middle-wavelength palettes among cichlids from LT and LM. In each case, these transitions occurred from different ancestors that expressed the same long-wavelength palette. We also identified similar but distinct patterns of correlated evolution between opsin expression, diet, and lens transmittance among cichlids from LT and LM as well. In contrast to regulatory changes, we identified few functional or potentially functional mutations in the protein-coding sequences of three variable opsins, with the possible exception of the SWS1 (ultraviolet) opsin. These results underscore the important contribution that gene regulation can make to rapid phenotypic evolution and adaptation.

Carone BR, Fauquier L, Habib N, Shea JM, Hart CE, Li R, Bock C, Li C, Gu H, Zamore PD, et al. Paternally induced transgenerational environmental reprogramming of metabolic gene expression in mammals. Cell [Internet]. 2010;143 (7) :1084 - 1096. Publisher's VersionAbstract

Epigenetic information can be inherited through the mammalian germline and represents a plausible transgenerational carrier of environmental information. To test whether transgenerational inheritance of environmental information occurs in mammals, we carried out an expression profiling screen for genes in mice that responded to paternal diet. Offspring of males fed a low-protein diet exhibited elevated hepatic expression of many genes involved in lipid and cholesterol biosynthesis and decreased levels of cholesterol esters, relative to the offspring of males fed a control diet. Epigenomic profiling of offspring livers revealed numerous modest (∼20%) changes in cytosine methylation depending on paternal diet, including reproducible changes in methylation over a likely enhancer for the key lipid regulator Ppara. These results, in conjunction with recent human epidemiological data, indicate that parental diet can affect cholesterol and lipid metabolism in offspring and define a model system to study environmental reprogramming of the heritable epigenome. © 2010 Elsevier Inc.

Neumeister H, Whitaker KW, Hofmann HA, Preuss T. Social and ecological regulation of a decision-making circuit. Journal of neurophysiology. 2010;104 (October 2010) :3180 - 3188.Abstract

Ecological context, sensory inputs, and the internal physiological state are all factors that need to be integrated for an animal to make appropriate behavioral decisions. However, these factors have rarely been studied in the same system. In the African cichlid fish Astatotilapia burtoni, males alternate between two phenotypes based on position in a social hierarchy. When dominant (DOM), fish display bright body coloration and a wealth of aggressive and reproductive behavioral patterns that make them conspicuous to predators. Subordinate (SUB) males, on the other hand, decrease predation risk by adopting cryptic coloration and schooling behavior. We therefore hypothesized that DOMs would show enhanced startle-escape responsiveness to compensate for their increased predation risk. Indeed, behavioral responses to sound clicks of various intensities showed a significantly higher mean startle rate in DOMs compared with SUBs. Electrophysiological recordings from the Mauthner cells (M-cells), the neurons triggering startle, were performed in anesthetized animals and showed larger synaptic responses to sound clicks in DOMs, consistent with the behavioral results. In addition, the inhibitory drive mediated by interneurons (passive hyperpolarizing potential [PHP] cells) presynaptic to the M-cell was significantly reduced in DOMs. Taken together, the results suggest that the likelihood for an escape to occur for a given auditory stimulus is higher in DOMs because of a more excitable M-cell. More broadly, this study provides an integrative explanation of an ecological and social trade-off at the level of an identifiable decision-making neural circuit.

Renn SCP, Machado HE, Jones A, Soneji K, Kulathinal RJ, Hofmann HA. Using comparative genomic hybridization to survey genomic sequence divergence across species: a proof-of-concept from Drosophila. BMC Genomics [Internet]. 2010;11 :271 - 271. Publisher's VersionAbstract

Genome-wide analysis of sequence divergence among species offers profound insights into the evolutionary processes that shape lineages. When full-genome sequencing is not feasible for a broad comparative study, we propose the use of array-based comparative genomic hybridization (aCGH) in order to identify orthologous genes with high sequence divergence. Here we discuss experimental design, statistical power, success rate, sources of variation and potential confounding factors. We used a spotted PCR product microarray platform from Drosophila melanogaster to assess sequence divergence on a gene-by-gene basis in three fully sequenced heterologous species (D. sechellia, D. simulans, and D. yakuba). Because complete genome assemblies are available for these species this study presents a powerful test for the use of aCGH as a tool to measure sequence divergence.

Aubin-Horth N, Letcher BH, Hofmann HA. Gene-expression signatures of Atlantic salmon's plastic life cycle. General and Comparative Endocrinology [Internet]. 2009;163 (3) :278 - 284. Publisher's VersionAbstract

How genomic expression differs as a function of life history variation is largely unknown. Atlantic salmon exhibits extreme alternative life histories. We defined the gene-expression signatures of wild-caught salmon at two different life stages by comparing the brain expression profiles of mature sneaker males and immature males, and early migrants and late migrants. In addition to life-stage-specific signatures, we discovered a surprisingly large gene set that was differentially regulated-at similar magnitudes, yet in opposite direction-in both life history transitions. We suggest that this co-variation is not a consequence of many independent cellular and molecular switches in the same direction but rather represents the molecular equivalent of a physiological shift orchestrated by one or very few master regulators. ?? 2009 Elsevier Inc. All rights reserved.

Machado HE, Pollen AA, Hofmann HA, Renn SCP. Interspecific profiling of gene expression informed by comparative genomic hybridization: A review and a novel approach in African cichlid fishes. Integrative and Comparative Biology. 2009;49 (6) :644 - 659.Abstract

Modern genomic approaches have facilitated great progress in our understanding of the molecular and genetic underpinnings of ecological and evolutionary processes. Analysis of gene expression through heterologous hybridization in particular has enabled genome-scale studies in many ecologically and evolutionarily interesting species. However, these studies have been hampered by the difficulty of comparing-on a common array platform-gene-expression profiles across species due to sequence divergence altering the dynamics of hybridization. All too often, comparisons of expression profiles across species were limited to contrasting lists of gene or even of just functional categories. Here we review these issues and propose a novel solution. Exploiting the diverse cichlid lineages of East Africa as our model-system, we then present results from an experimental case study that compares the neural gene-expression profiles of males and females of two species that differ in mating system. Using a single microarray platform that contains genes from one species, Astatotilapia burtoni, we conducted a total of 16 direct comparisons for neural gene-expression level between individual males and females from a pair of sister species, the polygynous Enantiopus melanogenys and the monogamous Xenotilapia flavipinnis. Next, we conducted a meta-analysis with previously published data from two different intra-specific expression studies to determine whether sex-specific neural gene expression is more closely associated with behavioral phenotype than it is with gonadal sex. Our results indicate that the gene expression profiles are species-specific to a large extent, as relatively few genes show conserved expression patterns associated with either sex. Finally, we describe how competitive genomic DNA hybridizations between the two focal species allow us to assess the degree to which divergence of sequences biases the results. We propose a masking technique that correlates interspecific expression ratios obtained with cDNA with hybridization ratios obtained with genomic DNA for the same set of species and determines threshold sequence divergence to reduce false positives. Our approach should be applicable to a wide range of interesting questions related to the evolution and ecology of gene expression.

Salzburger W, Renn SCP, Steinke D, Braasch I, Hofmann HA, Meyer A. Annotation of expressed sequence tags for the East African cichlid fish Astatotilapia burtoni and evolutionary analyses of cichlid ORFs. BMC genomics [Internet]. 2008;9 :96 - 96. Publisher's VersionAbstract

BACKGROUND: The cichlid fishes in general, and the exceptionally diverse East African haplochromine cichlids in particular, are famous examples of adaptive radiation and explosive speciation. Here we report the collection and annotation of more than 12,000 expressed sequence tags (ESTs) generated from three different cDNA libraries obtained from the East African haplochromine cichlid species Astatotilapia burtoni and Metriaclima zebra.\n\nRESULTS: We first annotated more than 12,000 newly generated cichlid ESTs using the Gene Ontology classification system. For evolutionary analyses, we combined these ESTs with all available sequence data for haplochromine cichlids, which resulted in a total of more than 45,000 ESTs. The ESTs represent a broad range of molecular functions and biological processes. We compared the haplochromine ESTs to sequence data from those available for other fish model systems such as pufferfish (Takifugu rubripes and Tetraodon nigroviridis), trout, and zebrafish. We characterized genes that show a faster or slower rate of base substitutions in haplochromine cichlids compared to other fish species, as this is indicative of a relaxed or reinforced selection regime. Four of these genes showed the signature of positive selection as revealed by calculating Ka/Ks ratios.\n\nCONCLUSION: About 22% of the surveyed ESTs were found to have cichlid specific rate differences suggesting that these genes might play a role in lineage specific characteristics of cichlids. We also conclude that the four genes with a Ka/Ks ratio greater than one appear as good candidate genes for further work on the genetic basis of evolutionary success of haplochromine cichlid fishes.