Deciphering the swordtail's tale: a molecular and evolutionary quest

The power of sexual selection to influence the evolution of morphological traits was first proposed more than 130 years ago by Darwin. Though long a controversial idea, it has been documented in recent decades for a host of animal species. Yet few of the established sexually selected features have been explored at the level of their genetic or molecular foundations. In a recent report, Zauner et al.1 describe some of the molecular features associated with one of the best characterized of sexually selected traits, the male-specific tail "sword" seen in certain species of the fish genus Xiphophorus. Zauner et al. find that the msxC gene, a gene previously implicated in fin development from work in zebrafish, is dramatically and specifically upregulated in the development of the ventral caudal fin rays, which give rise to the sword, in males. The results provide the first molecular insight into the development of this sexually selected trait while prompting new questions about the structure of the entire genetic network that underlies this trait. To fully understand the molecular-genetic and evolutionary history of this network, however, it will be essential to determine whether sword-development is a basal or derived trait in Xiphophorus.     

An organizer controls the development of the "sword," a sexually selected trait in swordtail fish

SUMMARY Male swordtail fish of the genus Xiphophorus (Poeciliidae) possess a "sword" that is composed of several colored elongated ventral fin rays of the caudal fin. The sword is a secondary sexual trait that evolved through sexual selection by female preference. To uncover the developmental mechanisms underlying the metamorphosis from a juvenile caudal fin to the sword, we have devised a transplantation protocol to assay the fate of single transplanted fin rays and their interactions with flanking rays. These experiments provide evidence for the existence of a previously unrecognized inductive signal that originates in those rays that develop into the two longest sword rays. This "sword organizer" causes adjacent fin rays to grow and become integrated into the sword and induces the development of an additional, typically pigmented sword in grafts to the dorsal part of the caudal fin. We show that the potential to develop a sword is restricted to certain parts of the caudal fin. Our findings suggest that the evolution of swords in swordtails required the acquisition of two developmental mechanisms: the establishment of signaling competence in prospective sword rays in the embryo or early larva, and its activation through androgen signaling in adult male fish.     

Retinoic Acid Is Involved in the Metamorphosis of the Anal Fin into an Intromittent Organ,the Gonopodium,in the Green Swordtail (Xiphophorus hellerii)

In poeciliid fish the male anal fin has been transformed into a gonopodium, an intromittent organ required for internal fertilization. Elevated testosterone levels induce metamorphosis of a subset of anal fin rays to grow and form the specialized terminal structures of the gonopodium. The molecular mechanisms underlying these processes are largely unknown. Here, we investigated whether retinoic acid (RA) signaling is involved in gonopodium development in the swordtail Xiphophorus hellerii. We showed that aldh1a2, a RA synthesizing enzyme, and the RA receptors, rar-ga and rar-gb, are expressed in anal fins during metamorphosis. aldh1a2 expression is regulated by testosterone in a concentration-dependent manner and is up-regulated in both hormone-induced and naturally developing gonopodia. Androgen receptor (ar), a putative regulator of gonopodial development, is co-expressed with aldh1a2 and the RA receptors in gonopodial rays. Importantly, experimental increase of RA signaling promoted growth of the gonopodium and increased the number of new segments. Based on gene expression analyses and pharmacological manipulation of gonopodium development, we show that the RA signaling pathway is activated in response to androgen signaling and promotes fin ray growth and development during the metamorphosis of the anal fin into the gonopodium.     

Microsatellites in the genus Xiphophorus,developed in Xiphophorus montezumae

Species of the genus Xiphophorus (swordtails and platies) are of great interest for the study of evolution of sexually selected traits like the sword, which is an elongation of ventral fin rays of the male caudal fin, that has evolved in several species within this genus. The detection of 10 microsatellites within the genus Xiphophorus will enable studies about the correlation of this trait with sexual reproductive success of males possessing swords of different lengths. These microsatellites will also be useful in determining population structure and enable paternity analysis in these species, where sperm storage is widespread.     

The cost of the sword: escape performance in male swordtails

The handicap theory of sexual selection posits that male display traits that are favored in mate choice come at a significant cost to performance. We tested one facet of this hypothesis in the green swordtail (Xiphophorus helleri). In this species, the lower ray of male caudal fin is extended into a 'sword', which serves to attract potential mates. However, bearing a long sword may increase drag and thus compromise a male's ability to swim effectively. We tested escape performance in this species by eliciting C-start escape responses, an instinctive escape behavior, in males with various sword lengths. We then removed males' swords and retested escape performance. We found no relationship between escape performance and sword length and no effect of sword removal on escape performance. While having a large sword may attract a predator's attention, our results suggest that sword size does not compromise a male's escape performance.     

Androgen dependent development of a modified anal fin, gonopodium, as a model to understand the mechanism of secondary sexual character expression in vertebrates

Male external genitalia show structural variations among species. Androgenic hormones are essential for the morphological specification of male type copulatory organs, while little is known about the developmental mechanisms of such secondary sexual characters. Western mosquitofish Gambusia affinis may offer a clue to the sexual differentiation researches, because they show a prominent masculine sexual character for appendage development, anal fin to gonopodium (GP) transition, and its formation could be induced in early juvenile fry by exogenously supplied androgens. We show that GP development is promoted by androgen dependent augmentation of sonic hedgehog (Shh) expression. Two AR cDNAs were cloned and identified as ARalpha and ARbeta from western mosquitofish. Both ARs were predominantly expressed in the distal region of outgrowing anal fin rays. Exposure of fry to androgen caused anal fin outgrowth concomitant with the Shh induction in the distal anal fin ray epithelium. When AR signaling was inhibited by its antagonist flutamide in fry, the initial induction of the Shh was suppressed accompanying retarded anal fin outgrowth. Similar suppression of anal fin outgrowth was induced by treatment with cyclopamine, an inhibitor of Shh signaling. These observations indicate that androgen dependent Shh expression is required for anal fin outgrowth leading to the formation of a genital appendage, the GP in teleost fishes. Androgen-induced GP formation may provide insights into the expression mechanism regulating the specification of sexual features in vertebrates.     

Strong reproductive skew among males in the multiply mated swordtail Xiphophorus multilineatus (Teleostei)

Male swordtails in the genus Xiphophorus display a conspicuous ventral elongation of the caudal fin, the sword, which arose through sexual selection due to female preference. Females mate regularly and are able to store sperm for at least 6 months. If multiple mating is frequent, this would raise the intriguing question about the role of female choice and male-male competition in shaping the mating system of these fishes. Size-dependent alternate mating strategies occur in Xiphophorus; one such strategy is courtship with a sigmoid display by large dominant males, while the other is gonopodial thrusting, in which small subordinate males sneak copulations. Using microsatellite markers, we observed a frequency of multiple paternity in wild-caught Xiphophorus multilineatus in 28% of families analyzed, but the actual frequency of multiple mating suggested by the correction factor PrDM was 33%. The number of fathers contributing genetically to the brood ranged from one to three. Compared to other species in the family Poeciliidae, both frequency and degree of multiple paternity were low. Paternity was found to be highly skewed, with one male on average contributing more than 70% to the offspring. Hence in this Xiphophorus mating system, typically one male dominates and sneaker males do not appear to be particularly effective. Postcopulatory mechanisms, however, such as sperm competition, are also indicated by our data, using sex-linked phenotypes among the offspring.     

Covariation between predation risk, body size and fin elaboration in the green swordtail, Xiphophorus helleri

Natural and sexual selection can have either opposing or synergistic effects on the evolution of traits. In the green swordtail Xiphophorus helleri , sexual selection arising from female choice is known to favour larger males and males with longer swords. We examined variation in male and female size and fin morphology among 15 populations that varied in their predation environments. Males and females from populations in which piscivorous fishes were present had longer and deeper bodies than did males and females from populations in which piscivorous fishes were absent. Controlling for a positive effect of body size on sword length, males from populations in which piscivores were present had relatively shorter swords than did males from populations in which piscivores were absent. The associations between morphology and predation environment may be due to direct effects of predation, indirect effects of predation, other sources of selection that covary with predator presence, or other environmental effects on trait expression. These results suggest that while sexual selection favours longer swords, natural selection may have an opposing effect on sword length in populations with predators. Natural selection on body size, however, may act synergistically with sexual selection in populations with predators; both may favour the evolution of larger body size. The body size results for X. helleri contrast with related taxa that have become model systems for the study of life history evolution.  © 2004 The Linnean Society of London, Biological Journal of the Linnean Society , 2004, 83 , 87–100.     

17α-甲基睾酮对剑尾鱼的影响

目的研究17α-甲基睾酮暴露对产后雌性剑尾鱼(Xiphophorus helleri)性逆转组织形态学变化的影响,探讨剑尾鱼第二性征变化作为水环境风险评价(ERA)的有效生物学标记的可能性。方法使用浸浴法以10μg/L 17α-甲基睾酮为剑尾鱼染毒持续7周,对实验鱼的体形、腹鳍、臀鳍、尾鳍及性腺等组织器官的变化进行观察;同时对幼鱼在实验室养殖条件下的性别分化进行统计。结果 17α-甲基睾酮对产后雌鱼有明显的影响,受诱导后出现性逆转,逐渐发育形成雄性第二性征:体形变纤细;腹鳍第1鳍条变短、第2和第3鳍条延长,臀鳍第3、4、5鳍条末端钩状化且第3鳍条变粗壮,尾鳍上下缘出现增生;体内与臀鳍相连的骨骼合并生长;受诱导的雌鱼性腺呈现退化趋势并伴有卵细胞坏死现象,生殖能力受到负面影响。结论剑尾鱼臀鳍和尾鳍变化可作为水环境雄激素物质污染监测的有效生物学标记。     

Relationships among msx gene structure and function in zebrafish and other vertebrates

The zebrafish genome contains at least five msx homeobox genes, msxA, msxB, msxC, msxD, and the newly isolated msxE. Although these genes share structural features common to all Msx genes, phylogenetic analyses of protein sequences indicate that the msx genes from zebrafish are not orthologous to the Msx1 and Msx2 genes of mammals, birds, and amphibians. The zebrafish msxB and msxC are more closely related to each other and to the mouse Msx3. Similarly, although the combinatorial expression of the zebrafish msx genes in the embryonic dorsal neuroectoderm, visceral arches, fins, and sensory organs suggests functional similarities with the Msx genes of other vertebrates, differences in the expression patterns preclude precise assignment of orthological relationships. Distinct duplication events may have given rise to the msx genes of modern fish and other vertebrate lineages whereas many aspects of msx gene functions during embryonic development have been preserved.      

The turn of the sword: length increases male swimming costs in swordtails

Sexual selection via female mate choice can result in the evolution of elaborate male traits that incur substantial costs for males. Despite increased interest in how female mating preferences contribute to the evolution of male traits, few studies have directly quantified the locomotor costs of such traits. A sexually selected trait that could affect movement costs is the sword exhibited by male swordtail fishes: while longer swords may increase male mating success, they could negatively affect the hydrodynamic aspects of swimming activities. Here, we examine the energetic costs of the sword in Xiphophorus montezumae by experimentally manipulating sword length and measuring male aerobic metabolism during two types of activity, routine swimming and courtship swimming. Direct measurements of oxygen consumption indicate that males with longer swords expend more energy than males with shortened swords during both types of swimming. In addition, the sword increases the cost of male courtship. Thus, while sexual selection via female choice favours long swords, males with longer swords experience higher metabolic costs during swimming, suggesting that sexual and natural selection have opposing effects on sword evolution. This study demonstrates a hydrodynamic cost of a sexually selected trait. In addition, this study discriminates between the cost of a sexually selected trait used in courtship and other courtship costs.     

Female disdain for swords in a swordtail fish

Studies of mate choice evolution tend to focus on how female mating preferences are acquired and how they select for greater elaboration of male traits. By contrast, far less is known about how female preferences might be lost or reversed. In swordtail fish Xiphophorus, female preference for the sword ornament is an ancestral trait. Xiphophorus birchmanni, however, is one species that has secondarily lost the sword. Using synthetic animation playback of "virtual" males, we found that female X. birchmanni preferred a swordless conspecific over a sworded heterospecific. Moreover, when offered the choice between a conspecific without a sword and one with a digitally attached sword, females preferred the former. These results suggest female preferences need not always select for elaboration of male traits, and they provide a plausible explanation for the lack of introgression of a sexual trait in a naturally occurring hybrid zone.     

Testis and gonopodium development in Anableps Dowi (Pisces: Anablepidae) correlated with pituitary gonadotropic zone area

Testicular development in the viviparous fish, Anableps dowi, is described from embryonic stages through sexual maturation and is correlated with development of the gonadotropic zone of the pituitary. Only isolated spermatogonia or cysts of early spermatogonia are found in embryos 1.8 to 4.4 cm in length and postnatal specimens 5.4 to 8.8 cm in length. Cysts with more advanced spermatogonia are seen in specimens from 7.9 to 10.8 cm in length. Spermatogenetic meioses are first observed in a 10.2 cm male. Specimens greater than 12.0 cm have mature spermatozoa (partial spermatozeugmata) within the main testicular ducts, indicating gonadal maturation. The morphological transformation of the anal fin into the intromittent gonopodium is also described, using both whole and cleared specimens. Even in the smallest postnatal male available (5.4 cm), the anal fin can be distinguished from that of an equivalent sized female. Anal fin changes proceed until the mature gonopodium is developed, when body lengths greater than 16 cm are reached. Testicular maturation, defined as the presence of partial spermatozeugmata in the main testicular ducts, precedes the completion of gonopodial development. Measurements of ventral gonadotropic zone area in pituitary mid-sagittal sections show a continuous increase from neonatal through sexually mature specimens. The results point to a role for testicular androgens in early testis development.     

Male Biases for Male Characteristics in Females in Priapella olmecae and Xiphophorus helleri (Family Poeciliidae)

Preexisting receiver biases can affect the evolution of sexually selected traits once traits favored by such biases arise. Female guayacón olmeca, Priapella olmecae , and green swordtail, Xiphophorus helleri , possess a bias favoring sworded conspecific males, despite phylogenetic evidence that the sexually selected sword expressed by male swordtails arose in Xiphophorus after the divergence of the two genera. In this study, we investigated the state of the bias favoring a sword in male Priapella and Xiphophorus to determine whether males also possess a bias that could operate in an intersexual selection context. Male P. olmecae preferred conspecific females with swords to those without swords. Thus, males and females in this unsworded species appear to share a preexisting bias favoring individuals of the opposite sex with swords. Male X. helleri , however, did not express a bias favoring sworded females; instead, in this species in which the sword is restricted to males, males discriminated against conspecific females with swords. Previous work suggests that female mate choice and male–male competition likely contribute to the maintenance of the sword in X. helleri . The sword may also play a role in sex recognition in swordtails. The absence of a sword preference in male green swordtails could reflect the current function of the sword.     

Rapid and socially induced change of a badge of status

This paper reports a new aspect of male signalling in the green swordtail Xiphophorus hellerii. Males are highly ornamented and possess a sword (an extension of the caudal fin), as well as a lateral stripe that has been shown to be a badge of status. Males of one natural population, however, were found to quickly change the colour of their lateral stripe to red when socially dominant over other males and black when subdominant, thereby exploiting the female preference for red stripes while dominant, and avoiding the costs of male aggression while subdominant.     

Zebrafish msxB, msxC and msxE function together to refine the neural-nonneural border and regulate cranial placodes and neural crest development

The zebrafish muscle segment homeobox genes msxB, msxC and msxE are expressed in partially overlapping domains in the neural crest and preplacodal ectoderm. We examined the roles of these msx genes in early development. Disrupting individual msx genes causes modest variable defects, whereas disrupting all three produces a reproducible severe phenotype, suggesting functional redundancy. Neural crest differentiation is blocked at an early stage. Preplacodal development begins normally, but placodes arising from the msx expression domain later show elevated apoptosis and are reduced in size. Cell proliferation is normal in these tissues. Unexpectedly, Msx-deficient embryos become ventralized by late gastrulation whereas misexpression of msxB dorsalizes the embryo. These effects appear to involve Distal-less (Dlx) protein activity, as loss of dlx3b and dlx4b suppresses ventralization in Msx-depleted embryos. At the same time, Msx-depletion restores normal preplacodal gene expression to dlx3b-dlx4b mutants. These data suggest that mutual antagonism between Msx and Dlx proteins achieves a balance of function required for normal preplacodal differentiation and placement of the neural-nonneural border.     

Secondary reduction of preference for the sword ornament in the pygmy swordtail Xiphophorus nigrensis (Pisces: Poeciliidae)

Female mating preferences can be secondarily lost for a number of reasons. We examined the preference of female pygmy swordtails, Xiphophorus nigrensis, for the sword, a conspicuous extension of the caudal fin in some males. Females failed to show a preference for conspecific males with swords when presented with live males naturally varying in sword length, with live males of manipulated sword length, and with synthetic animations of males expressing natural variation in sword length. Females showed a significant bias against swords when presented with synthetic animations bearing supernormal sword characteristics. The reduced preference for swords, relative to closely related fish, may result from an increase in the cost of choice due to predation risk, selection against mating with heterospecifics, or changes in the spatial and contrast properties of the conspecific signal.     

The anal fin complex in a weakly discharging electric fish, Gnathonemus petersii(Mormyridae)

An examination of the permanent bony structures of the anal fin complex in the mormyrid fish, Gnathonemus petersii , revealed two new structural sexual dimorphisms: longer proximal pterygiophores and wider anal fin rays in males than in females. Both structures are thought to facilitate the male's courtship‐associated anal fin reflex. Adult male mormyrid fishes are characterized by a dorsally directed indentation of the posterior body wall (anal fin indentation). The expression of this indentation in males, presumably driven by anal fin musculature, was correlated with the fish's gonadal state: large indentations were associated with high gonado‐somatic indices and small indentations with low indices.