New microsatellite loci confirm hybrid origin,parthenogenetic inheritance,and mitotic gene conversion in the gynogenetic Amazon molly (Poecilia formosa)

We describe the first microsatellite loci for the gynogenetic Amazon molly, Poecilia formosa, an all‐female species arisen through hybridization of the bisexual species Poecilia mexicana and Poecilia latipinna. The loci showed one to six alleles and an expected heterozygosity between zero and 0.75. As expected with parthenogenetic inheritance, most loci were either constantly homozygous (five loci) or constantly heterozygous (eight loci). For six loci, both heterozygotes and homozygotes occurred. This and the fact that some loci only showed alleles of one of the ancestral species could indicate genome homogenization through mitotic gene conversion. Our new loci conformed to the hybrid origin of Amazon molly and are also applicable to both ancestral bisexual species.     

Male mating behaviour of a molly, <Emphasis Type="Italic">Poecilia latipunctata</Emphasis>: a third host for the sperm-dependent Amazon molly, <Emphasis Type="Italic">Poecilia formosa</Emphasis>

The Tamesí molly, Poecilia latipunctata, has a very limited biogeographical range in northeast Mexico. This area is nested within the ranges of the Atlantic molly, Poecilia mexicana, and the unisexual Amazon molly, Poecilia formosa. Based on morphology, especially fin shape, the Tamesí molly has been considered to be a "short-fin" molly. We describe the courtship sequence of P. latipunctata. The courtship clearly places the species into the clade of "long-fin" mollies, a finding that corroborates earlier studies based on nuclear DNA and mitochondrial DNA. All three species live together in certain habitats. This renders P. latipunctata a potential host species for the sperm-dependent, unisexual Amazon molly. Using behavioural tests, we demonstrate that P. latipunctata males actually copulate with Amazon mollies, despite a pronounced preference for conspecific females. In laboratory experiments P. latipunctata males are capable of triggering embryogenesis in P. formosa females. Field observations support the hypothesis that P. latipunctata is a third host species for P. formosa, indicating that the Amazon molly effectively exploits all available host species for its gynogenetic mode of reproduction. Electronic Publication     

Biogeography of the Amazon molly: ecological niche and range limits of an asexual hybrid species

Aim To understand the relative contributions of environmental factors, dispersal limitations and the presence of sperm donors in determining the distribution of the Amazon molly (Poecilia formosa), a sperm‐dependent unisexual fish species of hybrid origin. To explore niche similarities and/or differences between the hybrid and parental species. To evaluate whether large‐scale abiotic factors can explain a successful introduction of both P. formosa and Poecilia latipinna. Location South‐east United States, Mexico and Central America. Methods We used abiotic variables in ecological niche modelling (ENM) to identify regions with suitable conditions for the presence of the Amazon molly and its two parental species (P. latipinna and Poecilia mexicana). We also used a recently developed metric to calculate the degree of niche overlap between the hybrid and its parental species. Results ENM produced highly significant models [all area under the curve (AUC) > 0.99 for the three species]. Annual mean temperature and minimum temperature of the coldest month were the variables that best explained the distribution of the Amazon molly. With the exception of south Florida, few areas beyond the known distribution of the species were predicted to have suitable environmental conditions. The hybrid species niche overlaps partially with the parental species. However, given the available data, it is neither more similar nor more different than expected by chance. Main conclusions Two different processes are acting to limit the distribution of P. formosa. At the northern limit, although a sperm donor species is present further north, suitable environmental conditions are absent from nearby locations. At the southern limit, a sperm donor species is present and areas with good environmental conditions are present at nearby locations, suggesting that dispersal ability is the limiting factor. We found that the hybrid species overlaps in a similar way with both parental species while still having its own niche identity. This result may be explained by the fact that hybrid species inherit characteristics of two ecologically divergent species, which can result in intermediate or even transgressive phenotypes. These results support recent work on the role of hybridization in diversification.     

Polymorphic MHC loci in an asexual fish,the amazon molly (Poecilia formosa; Poeciliidae)

Genes of the major histocompatibility complex (MHC) encode molecules that control immune recognition and are highly polymorphic in most vertebrates. The remarkable polymorphisms at MHC loci may be maintained by selection from parasites, sexual selection, or both. If asexual species show equal (or higher) levels of polymorphisms at MHC loci as sexual ones, this would mean that sexual selection is not necessary to explain the high levels of diversity at MHC loci. In this study, we surveyed the MHC diversity of the asexual amazon molly (Poecilia formosa) and one of its sexual ancestors, the sailfin molly (P. latipinna), which lives in the same habitat. We found that the asexual molly has polymorphic MHC loci despite its clonal reproduction, yet not as polymorphic as the sexual species. Although the nucleotide diversity was similar between the asexual and sexual species, the sexual species exhibited a greater genotypic diversity compared to the asexual one from the same habitats. Within‐genome diversity was similar for MHC class I loci, but for class IIB, the sexual species had higher diversity compared to the asexual — despite the hybrid origins and higher levels of heterozygosity at microsatellite loci in the asexual species. The level of positive selection appears to be similar between the two species, which suggests that these polymorphisms are maintained by selection. Thus, our findings do not allow us to rule out the sexual selection hypothesis for the evolution of MHC diversity, and although the sexual fish has higher levels of MHC‐diversity compared to the asexual species, this may be due to differences in demography, parasites, or other factors, rather than sexual selection.     

Divergent female mating preference in a clonal fish

In asexual all-female species, such as the Amazon molly, Poecilia formosa, that depend on sperm from “host males” only to trigger embryogenesis, mate choice does not provide any indirect, genetic benefits to the choosing female, although direct benefits are possible. Asexual species are thought to have a low evolutionary potential or evolvability due to the absence of meiotic recombination. Hence, theory predicts that mating preferences in P. formosa for male ornaments—if existent—should resemble those of females of the two parental species (Poecilia latipinna and Poecilia mexicana) involved in the natural hybridization that gave rise to the asexual P. formosa. When examining the female preference for dummy males with or without black vertical bars in the two parental species and in two lineages of P. formosa, a preference was detected in P. latipinna, but not in P. mexicana females. Interestingly, P. formosa living syntopic with P. latipinna also preferred striped males, while others living syntopic with P. mexicana preferred non-striped males. The evolutionary significance of this phenomenon remains largely unexplained, but it might indicate the evolution of mating preferences in a species with low evolutionary potential. Possible mechanisms include introgression and mitotic gene conversion. Females might use male coloration as indicator mechanisms for male traits that matter in terms of direct benefits.     

Juvenile survival in a unisexual/sexual complex of mollies

Unisexual species like the gynogenetic Amazon molly, Poecilia formosa, enjoy a twofold advantage over sexual species, because they do not produce males. Therefore, unisexuals should be able to outcompete and consequently, replace sexual species. For sperm-dependent (gynogenetic) unisexuals this creates a paradox: they cannot replace their sexual hosts without eradicating themselves. Thus, mechanisms must be in place to stabilize such mating systems. We assessed juvenile survivorship between asexual P. formosa and sexual Poecilia latipinna as a possible factor allowing for persistence and coexistence between the two sympatric species. Offspring of gynogenetic Amazon mollies did not differ significantly in survivorship compared to their sexual host, the Sailfin molly, P. latipinna. The presence of an adult female significantly reduced survival in both species, suggesting that filial cannibalisms operates in this system, but does not appear to play a role in stabilizing mixed sexual/asexual populations. Clark Hubbs, who spent 59 years at the University of Texas and was widely regarded as one of the state’s foremost researchers in the field of ichthyology, the study of fish, passed away February 3rd of 2008 after a long battle with colorectal cancer. He was 86.     

Quantifying the threat of extinction from Muller's ratchet in the diploid Amazon molly (<Emphasis Type="Italic">Poecilia formosa</Emphasis>)

Background  

The Amazon molly (Poecilia formosa) is a small unisexual fish that has been suspected of being threatened by extinction from the stochastic accumulation of slightly deleterious mutations that is caused by Muller's ratchet in non-recombining populations. However, no detailed quantification of the extent of this threat is available.     

The effect of rearing temperature on body shape and meristic characters in zebrafish (<Emphasis Type="Italic">Danio rerio</Emphasis>) juveniles

Considering its immediate costs of producing dispensable males, the maintenance of sexual reproduction is a major paradox in evolutionary biology. Asexual lineages that do not face such costs theoretically should replace sexuals over time. Nonetheless, several systems are known in which closely related sexual and asexual lineages stably coexist. In the present study, we studied a sexual/asexual mating complex of a sperm-dependent parthenogenetic fish (amazon molly, Poecilia formosa) and its sexual congeners, the sailfin molly P. latipinna and the Atlantic molly P. mexicana. We asked whether differences in feeding behavior could contribute to their stable coexistence. We conducted a laboratory experiment to compare feeding efficiencies and also measured the competitive abilities between the two reproductive forms. Additionally, we measured gut fullness of fishes caught in natural habitats. Contrary to our predictions, we could not find P. formosa to be less efficient in feeding. We argue that food competition in mollies plays a minor role in mediating coexistence between closely related asexual and sexual mollies.     

Character displacement in sailfin mollies, <Emphasis Type="Italic">Poecilia latipinna</Emphasis>: allozymes and behavior

Synopsis We analyzed variation in allozymes and mating preferences in 12 populations across much of the range of the sailfin molly, Poecilia latipinna. Sailfin mollies can be sympatric with its sexual parasite Amazon mollies, P. formosa. Amazon mollies must co-exist and mate with bisexual males of closely related species (including sailfin mollies) to induce embryogenesis but inheritance is strictly maternal. Where sailfin and Amazon mollies are sympatric there is evidence of reproductive character displacement as males show a significantly stronger mating preference for sailfin molly females over Amazon mollies compared to preferences of males from allopatric populations. From the allozyme data we found a moderate amount of genetic variation across all populations but this variation did not reveal significant partitioning between sympatric and allopatric populations. Additionally, we found no evidence for isolation by distance as genetic distance was not significantly correlated with geographic distance. While allozyme variation also did not significantly correlate with male mating preferences, there was a significant correlation between male mating preferences and geographic distance. This correlation between mating preferences and geographic distance may have arisen from coevolution with Amazon mollies resulting in reproductive character displacement. Taken together, the distribution of genetic and behavioral variation among sympatric and allopatric populations suggests that behavioral evolution has outpaced evolution at the allozyme loci we examined in P. latipinna.     

Do I Know You? Species Recognition Operates Within and Between the Sexes in a Unisexual–Bisexual Species Complex of Mollies

Male sailfin mollies (Poecilia latipinna) can be sexually parasitized by closely related, unisexual, gynogenetic Amazon mollies (Poecilia formosa). This study examined possible cues used by male P. latipinna to distinguish between conspecific females and sympatric, heterospecific P. formosa. Digital photos were used to create models to test male P. latipinna preference for model female P. latipinna and P. formosa with a full suite of traits and altered models of P. latipinna and P. formosa. Male P. latipinna significantly preferred models of either species over no stimulus, demonstrating that models elicit a male response. Males also significantly preferred female P. latipinna models over P. formosa models. We also examined species recognition by female sailfin mollies using the same models, and found that female sailfin mollies significantly preferred to associate with female P. latipinna over P. formosa. These results taken together suggest that the use of fish models yield results similar to those studies using live stimuli. Male preference was then tested for unaltered vs. altered models in the following combinations: (i) P. formosa vs. P. formosa with a female P. latipinna fin; (ii) P. formosa vs. P. formosa with a female P. latipinna lateral spot pattern; (iii) P. formosa vs. P. latipinna with a P. formosa fin and their spotless lateral pattern. Males did not significantly prefer models with any isolated traits over the unaltered P. formosa models. Thus, males may be using traits other than the ones isolated for species recognition or males may be using a suite of multiple traits to recognize conspecific females.     

HYBRIDIZATION LEADS TO SENSORY REPERTOIRE EXPANSION IN A GYNOGENETIC FISH,THE AMAZON MOLLY (POECILIA FORMOSA): A TEST OF THE HYBRID‐SENSORY EXPANSION HYPOTHESIS

Expansions in sensory systems usually require processes such as gene duplication and divergence, and thus evolve slowly. We evaluate a novel mechanism leading to rapid sensory repertoire expansion: hybrid‐sensory expansion (HSE). HSE occurs when two species with differently tuned sensory systems form a hybrid, bringing together alleles from each of the parental species. In one generation, a sensory repertoire is created that is the sum of the variance between parental species. The Amazon molly presents a unique opportunity to test the HSE hypothesis in a “frozen” hybrid. We compared opsin sequences of the Amazon molly, Poecilia formosa, to those of the parental species. Both parental species are homozygous at the RH2–1 locus and each of the four long wavelength sensitive loci, while P. formosa possess two different alleles at these loci; one matching each parental allele. Gene expression analysis showed P. formosa use the expanded opsin repertoire that was the result of HSE. Additionally, behavioral tests revealed P. formosa respond to colored stimuli in a manner similar or intermediate to the parental species P. mexicana and P. latipinna. Together these results strongly support the HSE hypothesis. Hybrid‐sensory repertoire expansion is likely important in other hybrid species and in other sensory systems.     

Trade‐offs between salinity preference and antipredator behaviour in the euryhaline sailfin molly Poecilia latipinna

Salinity preference and responses to predatory chemical cues were examined both separately and simultaneously in freshwater (FW) and saltwater (SW)‐acclimated sailfin mollies Poecilia latipinna, a euryhaline species. It was hypothesized that P. latipinna would prefer FW over SW, move away from chemical cues from a crayfish predator, and favour predator avoidance over osmoregulation when presented with both demands. Both FW and SW‐acclimated P. latipinna preferred FW and actively avoided predator cues. When presented with FW plus predator cues v. SW with no cues, P. latipinna were more often found in FW plus predator cues. These results raise questions pertaining to the potential osmoregulatory stress of salinity transitions in euryhaline fishes relative to the potential fitness benefits and whether euryhalinity is utilized for predator avoidance. This study sheds light on the potential benefits and consequences of being salt tolerant or intolerant and complicates the understanding of the selection pressures that have favoured the different osmoregulatory mechanisms among fishes.     

Cytogenetic studies of Poecilia (Pisces)

Natural populations of triploid females resembling the gynogenetic teleost, Poecilia formosa (Girard), occur in northeastern Mexico where they intermingle with diploid populations of this species and the members of congeneric bisexual species such as P. mexicana or P. latipinna. Mitotic configurations from gill epithelial cells show 46 chromosomes for the diploid fishes, but 69 chromosomes for members of the triploid clones associated with P. formosa. Triploid females have erythrocytes that are significantly larger than those from diploid specimens and also show a roughly 50% elevation in the average DNA content of their somatic nuclei. Similar analyses of two functionally incompetent males of P. formosa, of a number of bisexual F₁ and F₂ hybrid offpsring from P. latipinna x P. mexicana, and of females from several other poeciliid species consistently show only diploid DNA levels and somatic chromosome complements where 22N=46. Demonstration of cytogenetic criteria by which females from triploid clones may be clearly distinguished from sympatric diploid specimens of P. formosa or P. mexicana leaves unresolved, for the present, problems of an appropriate systematic designation for natural populations of triploid gynogenetic fishes. The role of sympatric speciation in the evolution of poeciliid genomes is discussed in terms of alternative mechanisms to account for the persistence in nature of a vertebrate triploid of hybrid origin.This work was supported by grants from the National Science Foundation (GB 7393) and from the U.S. Public Health Service (GM 14644).Recipient of a Research Career Development Award from the U.S. Public Health Service (1 K3 GM 3455).     

Male Mate Choice in Mixed Bisexual/Unisexual Breeding Complexes of Poecilia (Teleostei: Poeciliidae)

The livebearing all-female fish Poecilia formosa reproduces by gynogenesis, a modified form of parthenogenesis. P. formosa forms at least two breeding complexes: in its northern range it exists sympatrically with Poecilia latipinna and in its southern range with Poecilia mexicana. Differences between these complexes and their possible origin are discussed. Embryogenesis is triggered by sperm of males of these closely related sympatric species. Because inheritance is strictly maternal, from the male point of view energy and time invested are totally lost. In this study we wanted to elucidate whether males are able to distinguish between conspecific and parasitic females. It could be shown that males are able to distinguish females optically, but that this ability was obscured as soon as chemical and/or tactile contact was possible. Furthermore, we found that females in an attractive phase of their sexual cycle are always preferred, regardless of species. This is possibly the mechanism by which parasitic females obtain the matings they need to reproduce.     

The effects of various external cations and sodium transport inhibitors on sodium uptake by the sailfin molly,Poecilia latipinna, acclimated to sea water

Summary The effects of various external cations and sodium transport inhibitors on sodium uptake by the sailfin molly,Poecilia latipinna, acclimated to sea water was investigated. While the injection of acetazolamide had no effect on sodium uptake, addition of NH₄ ⁺, H⁺, K⁺ or amiloride to the external solution significantly inhibited the uptake of sodium by the fish. These data support the proposition that sea water acclimatedP. latipinna possess the Na⁺/H⁺ and/or Na⁺/NH₄ ⁺ exchange mechanisms which are normally thought to be present only in fresh water acclimated fish.     

Examination of boldness traits in sexual and asexual mollies (<Emphasis Type="Italic">Poecilia latipinna</Emphasis>, <Emphasis Type="Italic">P. formosa</Emphasis>)

Considering the high costs of sexual reproduction (e.g., the production of males), its maintenance and predominance throughout the Animal Kingdom remain elusive. Especially the mechanisms allowing for a stable coexistence of closely related sexual and asexual species are still subject to a lively debate. Asexuals should rapidly outnumber sexuals due to higher population growth rates, unless they face some disadvantages. Here, we investigate potential differences in feeding behavior in a system of sexual (sailfin mollies, Poecilia latipinna) and coexisting gynogenetic fishes (Amazon mollies, Poecilia formosa). In two different experiments, we tested for differences in behavioral traits associated with boldness. Bold individuals take higher risks for gains in resources, so shyer individuals should be less competitive. Our study was motivated by the recent finding that P. formosa are less likely to be preyed upon by piscine predators than P. latipinna. We asked whether this result is indicative of low boldness in P. formosa. However, no differences between the two species were detectible in our behavioral experiments measuring (a) time to emerge from shelter to explore a novel environment, (b) latency time until feeding in a novel environment, and (c) recovery time until feeding restarted after a simulated predator attack. Furthermore, different boldness measures were not correlated with each other within individuals.     

Frozen F1's amidst a masterpiece of nature: new insights into the rare hybrid origin of gynogenesis in the Amazon molly (Poecilia formosa)

All‐female ‘species’ of fish have been shown to be great models in ecological and evolutionary studies because of the insights they can provide into the origin and evolution of asexuality, the ecology of hybrids, associations between genotype and environment, and the maintenance of sex. Gynogenetic organisms that evolved from sexual ancestors, and combine the disadvantageous traits from sexuality and asexuality, have long baffled evolutionary biologists trying to understand their origin and persistence with their sympatric sexual counterparts. In this issue, a new study using an integrated molecular phylogenetic and classical genetic approach has uncovered compelling evidence regarding the obscure asexual origin of the Amazon molly, Poecilia formosa. By performing an extensive phylogeographic analysis, Stöck et al. (2010) provide evidence that the Amazon molly arose only once within its history, with monophyly being strongly supported by mitochondrial DNA and microsatellite analyses. This result, combined with an elaborate failed attempt to resynthesize the lineage, suggests that vertebrate gynogens such as the Amazon molly are not rare because they are at a disadvantage to their sexual counterparts, but because the genomic conditions under which they arise are rare. Organisms that apparently combine the disadvantages of both sexuality and asexuality remain difficult to understand from both an ecological and an evolutionary perspective, and Stöck et al. (2010) highlight several outstanding important questions. Nonetheless, given that we now have a better knowledge of the origin and history of this unique ‘species’, this should allow researchers to better understand how these frozen F1’s can persist amidst the masterpiece of nature.     

Are you more than the sum of your parents’ genes? Phenotypic plasticity in a clonal vertebrate and F1 hybrids of its parental species

All known vertebrate clones have originated from hybridization events and some have produced distinct evolutionary lineages via hybrid speciation. Amazon mollies (Poecilia formosa) present an excellent study system to investigate how clonal species have adapted to heterogeneous environments because they are the product of a single hybridization event between male sailfin mollies (Poecilia latipinna) and female Atlantic mollies (Poecilia mexicana). Here, we ask whether the hybrid species differs from the combination of its parental species’ genes in its plastic response to different environments. Using a three-way factorial design, we exposed neonates produced by Amazon mollies and reciprocal F1 hybrid crosses to different thermal (24°C and 29°C) and salinity (0/2, 12, and 20 ppt) regimes. We measured various ontogenetic and life history characteristics across the life span of females. Our major results were as follows: (1) Reaction norms of growth and maturation to temperature and salinity are quite similar between the two hybrid crosses; (2) Amazon molly reaction norms were qualitatively different than the P. latipinna male and P. mexicana female (L×M) hybrids for the ontogenetic variables; (3) Amazon molly reaction norms in reproductive traits were also quite different from L×M hybrids; and (4) The reaction norms of net fertility were very different between Amazon mollies and L×M hybrids. We conclude that best locale for Amazon mollies is not the best locale for hybrids, which suggests that Amazon mollies are not just an unmodified mix of parental genes but instead have adapted to the variable environments in which they are found. Hybridization resulting in asexuality may represent an underappreciated mechanism of speciation because the unlikely events required to produce such hybrids rarely occur and is dependent upon the genetic distance between parental species.     

The offspring size/fecundity trade-off and female fitness in the Atlantic molly (<Emphasis Type="Italic">Poecilia mexicana</Emphasis>, Poeciliidae)

Across a variety of taxa, large offspring have been demonstrated to have a fitness advantage over smaller offspring of the same species. However, producing large offspring often comes at the cost of having to produce fewer young, and the payoff (and thus, evolutionary outcome) of this trade-off is expected to vary between environments. Atlantic mollies (Poecilia mexicana: Poeciliidae, Teleostei), inhabiting a sulfidic cave and various non-sulfidic surface habitats in Tabasco (Mexico), are reproductively isolated and evolved divergent female life-history traits: females of the cave ecotype produce considerably fewer, but larger offspring. Stressful (sulfidic) environments may favor the production of larger offspring, as they are better able to cope with chemical stressors. It remains to be determined though if increased offspring survival outweighs the fitness cost of producing fewer but larger offspring even under benign laboratory conditions. We tested 30-day newborn survival of offspring from wild-caught P. mexicana females from diverging populations in a low-density, no predation, no cannibalism, and ad-libitum-food, benign laboratory environment. Survival rates were highly skewed towards larger cave molly offspring; however, surface molly females still had a higher fitness than cave molly females in terms of higher total numbers of surviving offspring. Our study provides evidence for an innate fitness advantage of larger cave molly offspring. Furthermore, the observed differences in life-history strategies could promote further divergence and reproductive isolation among these ecotypes of P. mexicana, because cave molly females immigrating into the adjacent surface habitats would most likely be selected against.