Life-history evolution in livebearing fishes (Teleostei: Poeciliidae)

Life histories lie at the heart of biology, because the tradeoffs each organism faces concerning the distribution of limited resources into either reproduction or maintenance and growth determine that organism's fitness. Generally speaking, an organism can choose to either invest in traits reducing age-specific mortality or to invest in traits increasing age-specific fecundity. Each decision in this regard leads to tradeoffs between current versus future reproduction, somatic maintenance or growth versus reproduction, or number, size and sex of offspring. Because of this, basic life history data provides the backbone for any organism-oriented research, effectively linking behavior, ecology, morphology, and physiology together.One family of teleosts fishes, the livebearing fishes (Poeciliidae), has proven to be excellent models for studying life history adaptations. Within Poeciliidae, at least three characters evolved to give rise to the present diversity within the family: (1) internal fertilization using a transformed anal fin, referred to as the gonopodium, for sperm transfer, (2) livebearing, and (3) different degrees of maternal provisioning for the developing young.Over the years, certain aspects of poeciliid life-history evolution have received particular attention. First and foremost, different predatory regimes have been demonstrated to drive rapid microevolutionary change in guppies, although evidence for macroevolutionary processes is as yet missing. Second, placental evolution and maternal provisioning strategies have been extensively studied, with the main focus of these studies on interspecific rather than intraspecific differences.The research detailed in my dissertation investigates several specific aspects of life-history evolution in livebearing fishes (Poeciliidae) that have so far been largely neglected. The first three chapters broaden our understanding of the importance of life-history evolution in the colonization of and ecological speciation in extreme habitats. Taken together, they highlight the relevance of life-history evolution in maintaining and even driving ecological speciation processes. The fourth chapter emphasizes the significance of environmental affects and intraspecific population differences on maternal provisioning strategies in livebearing fishes. Thus, cautioning against the traditional approach of investigating few ecologically similar populations and ascribing the identified patterns as rigid strategies for that particular species across its natural distribution. Finally, my last chapter opens the door for research on life histories as regulatory mechanisms for stability in the unisexual/bisexual mating system of the Amazon molly.Extremophile poeciliidsAlthough mechanisms that can lead to speciation are of fundamental importance to evolutionary biology, the actual process of speciation is still one of the least understood aspects of evolution. Recently, the idea that reproductive isolation and ultimately speciation can be the result of ecologically-based divergent selection (i.e., ecological speciation) has received a lot of attention. Two main components of ecological speciation are currently recognized: different sources of divergent selection, and different forms of reproductive isolation.Basically, sources of divergent selection can stem from differences between environments, ecological interactions, and/or sexual selection. Under the first scenario, populations inhabit environments that, for example, differ in resource availability or habitat structure, and as populations adapt to their specific environment, the may begin to diverge from one another. Under the second scenario, divergent selection between populations arises due to ecological interactions, for example via competition for shared resources. Finally, under the third scenario, mate preferences differ between populations from ecologically different environments, thus leading to divergent sexual selection.Different forms of reproductive isolation are usually distinguished based on whether they occur before (premating isolation) or after mating (postmating isolation). Premating isolation can for example evolve if populations are separated in time or space, or if populations are sexually isolated, i.e., populations simply differ in their mating signals and preferences. Postmating isolation on the other hand arises due to reduced hybrid fitness, which can originate from genetic incompatibilities, a mismatch between hybrid phenotypes and the environment, or a mismatch between hybrid phenotypes and the mating preferences of conspecifics.Poeciliids inhabit a wide variety of different habitats, ranging from small creeks to large streams, freshwater lakes to coastal (brackish) lagoons, and even subterranean to toxic environments. In particular, poeciliid species of at least three genera (i.e., Gambusia, Limia, and Poecilia) are known to have independently colonized various toxic habitats in the Dominican Republic, Mexico, and the United States. Some species, like Gambusia eurystoma, Limia sulphurophila, and Poecilia sulphuraria are even endemic to sulfidic waters, and via ecological speciation, are actually derived from poeciliids that can be found in sulfidic and nonsulfidic waters of adjacent habitats. In these systems, toxicity stems from naturally occurring hydrogen sulfide, which is acutely toxic to most metazoans, because it competes with oxygen in the respiratory chain. Nonetheless, extremophile poeciliids thrive in these habitats and often occur in high local population densities.A plateau in Tabasco, southern Mexico, provides an even more extreme suit of habitats. Here, divergent selection stems from drastic differences between the physio-chemical characteristics of environments inhabited by populations of the Atlantic molly (Poecilia mexicana), and habitats can be grouped into four distinct classes: toxic/cave, nontoxic/cave, toxic/surface, and nontoxic/surface. All habitat types are interconnected (i.e., no physical barriers prevent movement between environments) and merely several hundred meters apart; nonetheless, locally adapted populations are characterized by profound genetic differentiation coupled with strong behavioral, morphological, and physiological divergence. Premating isolation seems to be largely driven by natural selection in the cave molly system, because translocation experiments revealed high mortalities in fish transferred to waters of a different adjacent habitat type. But even a weak form of sexual isolation has been demonstrated: females prefer males from their own populations over males from adjacent environments. However, the role of life histories in population divergence in this and similar systems has so far remained unstudied.In my first chapter, I therefore began to address this issue by conducting a preliminary study that investigated differences in life-history evolution (i.e., fecundity) between Poecilia mexicana from the sulfidic Cueva del Azufre and from a benign surface habitat. Using a combination of data from field-collected and laboratory-reared animals, I was able to demonstrate that cave mollies not only exhibit reduced fecundity, but also that fecundity in cave mollies is less plastic than in their surface-dwelling counterparts. Although this clearly suggests a heritable component to fecundity divergence in cave mollies, the extent of total life history divergence between cave and surface mollies remained unexplored. Furthermore, these results led me to ask the following question: Was reduced fecundity a response to permanent darkness, to toxicity, or to the combination of both selective forces?In my second chapter, I attempted to address these new questions. To do so, I conducted a full life-history analysis on females collected from four types of habitats, which provided me with a natural 2x2 design, where the same species inhabits environments characterized by all possible combinations of the two selective forces darkness and toxicity (i.e., dark/toxic vs. dark/nontoxic vs. light/toxic vs. light/nontoxic). The data demonstrate a habitat-specific divergence in P. mexicana life histories, and both darkness and toxicity seem to select for the same trait dynamic: low fecundity and large offspring size. This particular trait dynamic most likely arose as a strategy to avoid cannibalism in the extreme habitats, and to create a more efficient body-volume-to-body-surface-area ratio with regards to the amount of body surface exposed to toxicity. Thus, my third chapter adds further evidence to the notion of ecological speciation driving population differentiation between populations from these different habitat types. However, are any of these life history patterns more generally applicable to life-history evolution and ecological speciation processes in poeciliids and other organisms, or is this type of divergence unique to the cave molly system?In order for me to focus on this, I needed to take a broader approach. Hence, for my third chapter, I turned to another system of poeciliids in toxic waters: the Banos del Azufre system, in Tabasco, southern Mexico. Here, two species (Gambusia eurystoma and Poecilia sulphuraria) have diverged, and eventually speciated, in sulfidic waters. Both of these sulfide-endemics derived from two more widespread poeciliids that inhabit surrounding nontoxic and toxic habitats (Gambusia sexradiata and Poecilia mexicana, respectively). In chapter 3, I report on a comparative analysis, in which I contrast life histories of all four species from different habitat types. Even though I also found evidence for genus-specific responses to toxicity, the most pronounced pattern was the same as in the cave molly: hydrogen sulfide induced low fecundity and large offspring size, and the higher the toxicity, the lower the fecundity and the larger the offspring. Overall, my first three chapters therefore provide me with evidence that poeciliid fishes colonizing sulfidic habitats exhibit convergent life history evolution, with the potential to eventually result in complete speciation (as is the case with P. sulphuraria and G. eurystoma). I therefore propose that after an initial period of trait divergence, life-history evolution may provide an additional mechanism for further divergence, because dispersers between habitats will suffer from reduced fitness due to the wrong life history strategy in the new environment. Hence, life histories can be an important mechanism for further divergence in the advanced stages of ecological speciation processes.In my fourth chapter, I am turning to another important life-history aspect of livebearing animals; namely the question of maternal strategies for embryo provisioning. Basically, a mother has two choices: she can either store all the resources required for successful embryo development within the yolk prior to fertilization (lecithotrophy), or she can supplement yolk-stored nutrients with additional provisions via direct transfer after fertilization (matrotrophy). Even though recent theoretical models and empirical studies have stressed that maternal provisioning strategies are resource-dependent, mainstream life history research still attempts to classify an organism as being either exclusively lecithotrophic or matrotrophic. I therefore compared maternal provisioning strategies between two populations of Poecilia mexicana that inhabit vastly different environments: a toxic, resource-poor limestone cave and a benign, resource-rich surface habitat. Furthermore, I directly compared two different techniques that are widely used to quantify maternal provisioning: the indirect matrotrophy index analysis and the direct radio-tracer assay of maternal provisioning. According to the matrotrophy index analysis, both populations of P. mexicana are purely lecithotrophic, while according to the radio-tracer assay, both populations provide similar levels of postfertilization nutrient transfer (i.e., matrotrophy). Together with results from chapter 2, this suggests that P. mexicana is at least capable of incipient matrotrophy and that, to avoid misclassification, both techniques of quantifying nutrient transfer should ideally be employed together. Finally, I propose that current theories on the evolution of matrotrophy in poeciliids need to be revised, since this chapter and other recent research suggest that most livebearing fishes are probably characterized by dual provisioning rather than distinct strategies of either lecithotrophy or matrotrophy.Unisexual poeciliidsUnisexual vertebrates have long been used as model systems to study the maintenance of recombination and all species described to date originated as hybrids. Among unisexual vertebrates, fishes are of central interest because of their peculiar reproductive mechanisms. Several unisexual fishes reproduce via sperm-dependent parthenogenesis (gynogenesis), where sperm is required to initiate embryogenesis, but inheritance is strictly maternal.In such mating systems involving a bisexual and a unisexual species we find two competing types of females, which rely on the same resource, sperm, but have fundamentally different population dynamics. According to theoretical models, the unisexuals should quickly outcompete the bisexuals, thus driving them to extinction. However, this would inevitably lead to their own subsequent demise. Hence, the stability of bisexual/unisexual mating systems and the factors underlying the maintenance of this stability are currently of great interest in evolutionary ecology.The Amazon molly, Poecilia formosa, is a gynogenetic, all-female poeciliid. Poecilia formosa resulted from a single natural hybridization of the two sexual species Poecilia latipinna and Poecilia mexicana, and uses sperm from males of the two parental species for gynogenetic reproduction. Ecological differences and behavioral regulation have been proposed as two possible mechanisms explaining the stability in complexes of gynogens and sexuals, because at certain stages in their life history the gynogen could suffer a significant reduction of fitness. Since P. latipinna and P. mexicana males are not related to the resulting offspring from matings with Amazon mollies, they should be under selection to avoid them. Accordingly, both P. latipinna and P. mexicana males have been shown to (a) discriminate between heterospecific and conspecific females, and (b) to prefer to mate with conspecific females under most circumstances. However, these mating preferences are only of importance if they translate into reduced mating success of gynogens relative to conspecific females.For my fifth and final chapter, I therefore investigated the mate choice:life history interface by documenting the presence of sperm in five natural populations. I extracted sperm from female fish of five syntopic populations of P. formosa and P. latipinna in Texas by flushing out the genital tract. A higher proportion of bisexual females had sperm than unisexuals. Also, among those females that had sperm, bisexuals had more sperm than unisexuals. Even though the results gained from this analysis cannot be used to infer the amount of sperm transferred per copulation or the number of heterospecific matings, they nonetheless show that Amazon mollies receive less sperm in the wild than Sailfin molly females. This represents the first study to investigate the ultimate effects of male mate choice as a stabilizing factor in natural populations, and my results suggest that P. formosa may in fact be sperm limited; however, future studies on life-history differences will have to determine whether this actually results in a fitness reduction

Shared and unique patterns of embryo development in extremophile poeciliids

Background: Closely related lineages of livebearing fishes have independently adapted to two extreme environmental factors: toxic hydrogen sulphide (H2S) and perpetual darkness. Previous work has demonstrated in adult specimens that fish from these extreme habitats convergently evolved drastically increased head and offspring size, while cave fish are further characterized by reduced pigmentation and eye size. Here, we traced the development of these (and other) divergent traits in embryos of Poecilia mexicana from benign surface habitats (“surface mollies”) and a sulphidic cave (“cave mollies”), as well as in embryos of the sister taxon, Poecilia sulphuraria from a sulphidic surface spring (“sulphur mollies”). We asked at which points during development changes in the timing of the involved processes (i.e., heterochrony) would be detectible. Methods and Results: Data were extracted from digital photographs taken of representative embryos for each stage of development and each type of molly. Embryo mass decreased in convergent fashion, but we found patterns of embryonic fat content and ovum/embryo diameter to be divergent among all three types of mollies. The intensity of yellow colouration of the yolk (a proxy for carotenoid content) was significantly lower in cave mollies throughout development. Moreover, while relative head size decreased through development in surface mollies, it increased in both types of extremophile mollies, and eye growth was arrested in mid-stage embryos of cave mollies but not in surface or sulphur mollies. Conclusion: Our results clearly demonstrate that even among sister taxa convergence in phenotypic traits is not always achieved by the same processes during embryo development. Furthermore, teleost development is crucially dependent on sufficient carotenoid stores in the yolk, and so we discuss how the apparent ability of cave mollies to overcome this carotenoid-dependency may represent another potential mechanism explaining the lack of gene flow between surface and cave mollies

Predator avoidance in extremophile fish

Extreme habitats are often characterized by reduced predation pressures, thus representing refuges for the inhabiting species. The present study was designed to investigate predator avoidance of extremophile populations of Poecilia mexicana and P. sulphuraria that either live in hydrogen sulfide-rich (sulfidic) springs or cave habitats, both of which are known to have impoverished piscine predator regimes. Focal fishes that inhabited sulfidic springs showed slightly weaker avoidance reactions when presented with several naturally occurring predatory cichlids, but strongest differences to populations from non-sulfidic habitats were found in a decreased shoaling tendency with non-predatory swordtail (Xiphophorus hellerii) females. When comparing avoidance reactions between P. mexicana from a sulfidic cave (Cueva del Azufre) and the adjacent sulfidic surface creek (El Azufre), we found only slight differences in predator avoidance, but surface fish reacted much more strongly to the non-predatory cichlid Vieja bifasciata. Our third experiment was designed to disentangle learned from innate effects of predator recognition. We compared laboratory-reared (i.e., predator-naïve) and wild-caught (i.e., predator-experienced) individuals of P. mexicana from a non-sulfidic river and found no differences in their reaction towards the presented predators. Overall, our results indicate (1) that predator avoidance is still functional in extremophile Poecilia spp. and (2) that predator recognition and avoidance reactions have a strong genetic basis

Divergent Evolution of Male Aggressive Behaviour: Another Reproductive Isolation Barrier in Extremophile Poeciliid Fishes?

Reproductive isolation among locally adapted populations may arise when immigrants from foreign habitats are selected against via natural or (inter-)sexual selection (female mate choice). We asked whether also intrasexual selection through male-male competition could promote reproductive isolation among populations of poeciliid fishes that are locally adapted to extreme environmental conditions [i.e., darkness in caves and/or toxic hydrogen sulphide (H2S)]. We found strongly reduced aggressiveness in extremophile P. oecilia mexicana, and darkness was the best predictor for the evolutionary reduction of aggressiveness, especially when combined with presence of H2S. We demonstrate that reduced aggression directly translates into migrant males being inferior when paired with males from non-sulphidic surface habitats. By contrast, the phylogenetically old sulphur endemic P. sulphuraria from another sulphide spring area showed no overall reduced aggressiveness, possibly indicating evolved mechanisms to better cope with H2S

Survival in an extreme habitat: the roles of behaviour and energy limitation

Extreme habitats challenge animals with highly adverse conditions, like extreme temperatures or toxic substances. In this paper, we report of a fish (Poecilia mexicana) inhabiting a limestone cave in Mexico. Several springs inside the cave are rich in toxic H2S. We demonstrate that a behavioural adaptation, aquatic surface respiration (ASR), allows for the survival of P. mexicana in this extreme, sulphidic habitat. Without the possibility to perform ASR, the survival rate of P. mexicana was low even at comparatively low H2S concentrations. Furthermore, we show that food limitation affects the survival of P. mexicana pointing to energetically costly physiological adaptations to detoxify H2

Life on the edge: hydrogen sulfide and the fish communities of a Mexican cave and surrounding waters

Most eucaryotic organisms classified as living in an extreme habitat are invertebrates. Here we report of a fish living in a Mexican cave (Cueva del Azufre) that is rich in highly toxic H2S. We compared the water chemistry and fish communities of the cave and several nearby surface streams. Our study revealed high concentrations of H2S in the cave and its outflow (El Azufre). The concentrations of H2S reach more than 300μM inside the cave, which are acutely toxic for most fishes. In both sulfidic habitats, the diversity of fishes was heavily reduced, and Poecilia mexicana was the dominant species indicating that the presence of H2S has an all-or-none effect, permitting only few species to survive in sulfidic habitats. Compared to habitats without H2S, P. mexicana from the cave and the outflow have a significantly lower body condition. Although there are microhabitats with varying concentrations of H2S within the cave, we could not find a higher fish density in areas with lower concentrations of H2S. We discuss that P. mexicana is one of the few extremophile vertebrates. Our study supports the idea that extreme habitats lead to an impoverished species diversit

Locally adapted fish populations maintain small-scale genetic differentiation despite perturbation by a catastrophic flood event

Background: Local adaptation to divergent environmental conditions can promote population genetic differentiation even in the absence of geographic barriers and hence, lead to speciation. Perturbations by catastrophic events, however, can distort such parapatric ecological speciation processes. Here, we asked whether an exceptionally strong flood led to homogenization of gene pools among locally adapted populations of the Atlantic molly (Poecilia mexicana, Poeciliidae) in the Cueva del Azufre system in southern Mexico, where two strong environmental selection factors (darkness within caves and/or presence of toxic H2S in sulfidic springs) drive the diversification of P. mexicana. Nine nuclear microsatellites as well as heritable female life history traits (both as a proxy for quantitative genetics and for trait divergence) were used as markers to compare genetic differentiation, genetic diversity, and especially population mixing (immigration and emigration) before and after the flood. Results: Habitat type (i.e., non-sulfidic surface, sulfidic surface, or sulfidic cave), but not geographic distance was the major predictor of genetic differentiation. Before and after the flood, each habitat type harbored a genetically distinct population. Only a weak signal of individual dislocation among ecologically divergent habitat types was uncovered (with the exception of slightly increased dislocation from the Cueva del Azufre into the sulfidic creek, El Azufre). By contrast, several lines of evidence are indicative of increased flood-induced dislocation within the same habitat type, e.g., between different cave chambers of the Cueva del Azufre. Conclusions: The virtual absence of individual dislocation among ecologically different habitat types indicates strong natural selection against migrants. Thus, our current study exemplifies that ecological speciation in this and other systems, in which extreme environmental factors drive speciation, may be little affected by temporary perturbations, as adaptations to physico-chemical stressors may directly affect the survival probability in divergent habitat types

Whistle communication in mammal-eating killer whales (Orcinus orca): further evidence for acoustic divergence between ecotypes

Public signaling plays an important role in territorial and sexual displays in animals; however, in certain situations, it is advantageous to keep signaling private to prevent eavesdropping by unintended receivers. In the northeastern Pacific, two populations of killer whales (Orcinus orca), fish-eating “resident” killer whales and mammal-eating “transient” killer whales, share the same habitat. Previous studies have shown that residents use whistles as private signals during close-range communication, where they probably serve to coordinate behavioral interactions. Here, we investigated the whistling behavior of mammal-eating killer whales, and, based on divergent social structures and social behaviors between residents and transients, we predicted to find differences in both whistle usage and whistle parameters. Our results show that, like resident killer whales, transients produce both variable and stereotyped whistles. However, clear differences in whistle parameters between ecotypes show that the whistle repertoire of mammal-eating killer whales is clearly distinct from and less complex than that of fish-eating killer whales. Furthermore, mammal-eating killer whales only produce whistles during “milling after kill” and “surface-active” behaviors, but are almost completely silent during all other activities. Nonetheless, whistles of transient killer whales may still serve a role similar to that of resident killer whales. Mammal-eating killer whales seem to be under strong selection to keep their communication private from potential prey (whose hearing ranges overlap with that of killer whales), and they appear to accomplish this mainly by restricting vocal activity rather than by changes in whistle parameters

Predator-induced changes of female mating preferences: innate and experiential effects

Background: In many species males face a higher predation risk than females because males display elaborate traits that evolved under sexual selection, which may attract not only females but also predators. Females are, therefore, predicted to avoid such conspicuous males under predation risk. The present study was designed to investigate predator-induced changes of female mating preferences in Atlantic mollies (Poecilia mexicana). Males of this species show a pronounced polymorphism in body size and coloration, and females prefer large, colorful males in the absence of predators. Results: In dichotomous choice tests predator-naïve (lab-reared) females altered their initial preference for larger males in the presence of the cichlid Cichlasoma salvini, a natural predator of P. mexicana, and preferred small males instead. This effect was considerably weaker when females were confronted visually with the non-piscivorous cichlid Vieja bifasciata or the introduced non-piscivorous Nile tilapia (Oreochromis niloticus). In contrast, predator experienced (wild-caught) females did not respond to the same extent to the presence of a predator, most likely due to a learned ability to evaluate their predators' motivation to prey. Conclusions: Our study highlights that (a) predatory fish can have a profound influence on the expression of mating preferences of their prey (thus potentially affecting the strength of sexual selection), and females may alter their mate choice behavior strategically to reduce their own exposure to predators. (b) Prey species can evolve visual predator recognition mechanisms and alter their mate choice only when a natural predator is present. (c) Finally, experiential effects can play an important role, and prey species may learn to evaluate the motivational state of their predators. Keywords: Sexual selection; female choice; non-independent mate choice; predator recognition; Poecilia mexican

Does personality affect premating isolation between locally-adapted populations?

Background: One aspect of premating isolation between diverging, locally-adapted population pairs is female mate choice for resident over alien male phenotypes. Mating preferences often show considerable individual variation, and whether or not certain individuals are more likely to contribute to population interbreeding remains to be studied. In the Poecilia mexicana-species complex different ecotypes have adapted to hydrogen sulfide (H2S)-toxic springs, and females from adjacent non-sulfidic habitats prefer resident over sulfide-adapted males. We asked if consistent individual differences in behavioral tendencies (animal personality) predict the strength and direction of the mate choice component of premating isolation in this system. Results: We characterized focal females for their personality and found behavioral measures of ‘novel object exploration’, ‘boldness’ and ‘activity in an unknown area’ to be highly repeatable. Furthermore, the interaction term between our measures of exploration and boldness affected focal females’ strength of preference (SOP) for the resident male phenotype in dichotomous association preference tests. High exploration tendencies were coupled with stronger SOPs for resident over alien mating partners in bold, but not shy, females. Shy and/or little explorative females had an increased likelihood of preferring the non-resident phenotype and thus, are more likely to contribute to rare population hybridization. When we offered large vs. small conspecific stimulus males instead, less explorative females showed stronger preferences for large male body size. However, this effect disappeared when the size difference between the stimulus males was small. Conclusions: Our results suggest that personality affects female mate choice in a very nuanced fashion. Hence, population differences in the distribution of personality types could be facilitating or impeding reproductive isolation between diverging populations depending on the study system and the male trait(s) upon which females base their mating decisions, respectively