Frequency of interspecific mating in salamanders of the plethodontid genus Desmognathus: different experimental designs may yield different results

Heterosexual courtship trials were staged between dusky salamanders of two species ( Desmognathus fuscus and D. santeetlah ) from two allopatric populations in the southern Appalachian Mountains of eastern North America. Two experiments were conducted in order to test for effects of experimental design on estimates of the strength of sexual isolation between these species. In Experiment 1, frequencies of interspecific mating were determined when: (1) a male was presented with a conspecific and a heterotypic female successively; and (2) both types of females were presented simultaneously to a male. The frequency of interspecific mating by male D. fuscus was greatest when females were presented successively. The frequency of interspecific mating by male D. santeetlah was not influenced by method of female presentation. An index of sexual isolation of +0.29 was obtained between D. fuscus and D. santeetlah when females were presented successively, and of +0.63 when females were presented simultaneously. In Experiment 2, interspecific mating frequencies were determined in courtship arenas of two different sizes. No significant effect of arena size on frequency of interspecific mating was found for D. fuscus. In D. santeetlah , a trend towards greater mating frequency in small arenas was almost significant. These results suggest that interspecific mating frequencies (and thus estimates of the strength of sexual isolation between populations or species) may be sensitive to the experimental design employed. The validity and generality of this conclusion for other animals is an urgent, empirical issue.     

Population differentiation in plethodontid salamanders: divergence of allozymes and sexual compatibility among populations of Desmognathus imitator and D. ochrophaeus (Gaudata: Plethodontidae)

Plethodontid salamanders of the genus Desmognathus exhibit varying levels of genetic differentiation among and within both allopatric and sympatric taxa. This provides excellent opportunities to study population differentiation and speciation. Two morphologically similar species in this genus, D. imitator and D. ochrophaeus, are genetically well-differentiated from one another and occur in sympatry with no evidence of hybridization and introgression. We report that the degree of sexual isolation between these two species is very high, regardless of whether the populations under comparison are allopatric or sympatric with one another. Neither reinforcement nor reproductive character displacement are required to explain the evolution of sexual incompatibility in sympatry. Sexual behaviour apparently diverges while populations are allopatric with one another. Preliminary study indicates that D. imitator consists of populations among which there may be significant sexual isolation in the absence of detectable genetic differentiation.     

Stifled sex in sympatry: patterns of sexual incompatibility among desmognathine salamanders

Sexual incompatibility due to differences in mate-recognition systems can be an important factor in preventing hybridization and gene flow between animal populations in sympatry. We tested in the laboratory for sexual incompatibility between populations of two species of desmognathine salamanders that occur in sympatry in south-eastern Kentucky, North America, Desmognathus monticola and D. welteri (Caudata: Plethodontidae: Desmognathinae). Spermatophore deposition and insemination (two related measures of sexual success) were frequent in intraspecific encounters, but interspecific encounters never resulted in the deposition of spermatophores by males (Experiment 1). The sexual behaviour patterns of these two species are similar qualitatively, but all encounters staged between them failed to progress beyond the most preliminary of courtship interactions (Experiment 2), perhaps due to species differences in chemosensory communication. Our data on sexual incompatibility between D. monticola and D. welteri are similar to those available for other desmognathine taxa that occur in sympatry. Review of a broader data-set on patterns of sexual incompatibility both within and between species, and in allopatry and in sympatry, provides evidence for both divergence of mate-recognition systems in allopatry and enhancement of sexual incompatibility in sympatry for desmognathine salamanders. We hypothesize that diversification in allopatry may be a consequence of natural and/or sexual selection acting to promote sexual success within populations. Enhancement in sympatry may be a consequence of subsequent selection for accurate species recognition under the threat of hybridization or gene flow between species.  © 2004 The Linnean Society of London, Biological Journal of the Linnean Society , 2004, 82 , 367–375.     

Population and Species Divergence of Chemical Cues that Influence Male Recognition of Females in Desmognathine Salamanders

Growing evidence indicates that males may be more discriminating of mating partners than often has been assumed. In the North American Ocoee dusky salamander, Desmognathus ocoee (Plethodontidae: Desmognathinae), sexual incompatibility among conspecific populations is high in encounters staged in the laboratory, at least in part because males fail to recognize ‘other’ females as appropriate targets for courtship. I used Y-mazes to test the hypothesis that males of D. ocoee discriminate between substrate-borne chemical cues produced by ‘own’ (homotypic) and ‘other’ (heterotypic) females. Males of four populations discriminated in favor of substrates soiled by homotypic females over clean (control) substrates (expt 1), suggesting that females produce chemical cues of sociosexual significance to males. Furthermore, males from these populations discriminated in favor of substrates soiled by homotypic females vs. substrates soiled by heterotypic females (expt 2), both conspecific and heterospecific (D. carolinensis and D. orestes). Thus, differences among populations and species in female chemical cues appear to affect the chemotactic responses of males. I suggest that, together with differences in behavioral signals and responses exhibited during courtship, differences in female chemical cues likely contribute to sexual incompatibility among populations and taxa of desmognathine salamanders.     

Variations in sexual behavior among capuchin monkeys function for conspecific mate recognition: a phylogenetic analysis and a new hypothesis for female proceptivity in tufted capuchins

Researchers of capuchin monkeys have noted stereotyped body postures, facial expressions, and vocalizations that accompany copulations in this genus. Notable variations in these sexual behaviors are observed across capuchin species. Although several hypotheses exist to explain variation in the duration and vigor of sexual behaviors across species, there is no proposed explanation for variation in the forms of these behaviors. I hypothesized that the forms of sexual behaviors function as recognition signals of conspecific mates. Such signals are adaptive when F1 hybrids exhibit reduced fitness compared with nonhybrid offspring. Recent evidence from nonprimate taxa supports the existence of species recognition signals during mating. Using newly observed sexual behaviors for Cebus albifrons and a recent phylogeny of capuchins, I found significant support for a key prediction of the conspecific mate recognition hypothesis: evolutionary changes in sexual behaviors were associated with speciation. Given the resultant best model for evolution of sexual behaviors, I reconstructed the ancestral pattern of sexual behaviors for extant capuchin species (genera Cebus and Sapajus). This reconstruction suggests that the extreme female proceptivity of tufted capuchin monkeys may function to increase female reproductive choices in the context of sperm-limited males (genus Sapajus).     

Reproductive character displacement generates reproductive isolation among conspecific populations: an artificial neural network study

When interactions with heterospecifics prevent females from identifying conspecific mates, natural selection can promote the evolution of mating behaviours that minimize such interactions. Consequently, mating behaviours may diverge among conspecific populations in sympatry and in allopatry with heterospecifics. This divergence in conspecific mating behaviours-reproductive character displacement-can initiate speciation if mating behaviours become so divergent as to generate reproductive isolation between sympatric and allopatric conspecifics. We tested these ideas by using artificial neural networks to simulate the evolution of conspecific mate recognition in populations sympatric and allopatric with different heterospecifics. We found that advertisement calls diverged among the different conspecific populations. Consequently, networks strongly preferred calls from their own population to those from foreign conspecific populations. Thus, reproductive character displacement may promote reproductive isolation and, ultimately, speciation among conspecific populations.     

Rapid evolution of cuticular hydrocarbons in a species radiation of acoustically diverse Hawaiian crickets (Gryllidae: trigonidiinae: Laupala)

Understanding the origin and maintenance of barriers to gene exchange is a central goal of speciation research. Hawaiian swordtail crickets (genus Laupala) represent one of the most rapidly speciating animal groups yet identified. Extensive acoustic diversity, strong premating isolation, and female preference for conspecific acoustic signals in laboratory phonotaxis trials have strongly supported divergence in mate recognition as the driving force behind the explosive speciation seen in this system. However, recent work has shown that female preference for conspecific male calling song does not extend to mate choice at close range among these crickets, leading to the hypothesis that additional sexual signals are involved in mate recognition and premating isolation. Here we examine patterns of variation in cuticular lipids among several species of Laupala from Maui and the Big Island of Hawaii. Results demonstrate (1) a rapid and dramatic evolution of cuticular lipid composition among species in this genus, (2) significant differences among males and females in cuticular lipid composition, and (3) a significant reduction in the complexity of cuticular lipid profiles in species from the Big Island of Hawaii as compared to two outgroup species from Maui. These results suggest that behavioral barriers to gene exchange in Laupala may be composed of multiple mate recognition signals, a pattern common in other cricket species.     

Nest and Egg Recognition by Salamanders in the Genus Desmognathus: A Comprehensive Re‐Examination

We conducted a comparative investigation of nest relocation and egg recognition by four species of streamside salamanders indigenous to eastern North America: Seepage Salamander (Desmognathus aeneus), Santeetlah Salamander (Desmognathus santeetlah), Ocoee Salamander (Desmognathus ocoee), and Northern Dusky Salamander (Desmognathus fuscus). Females of all four species were able to relocate their nest sites following displacement of 1 m. Upon return to a nest site following natural displacement, females must be able to recognize their eggs and, in some instances, choose between their own clutch and the unattended clutch of a conspecific. In two‐choice behavioral tests, female salamanders of all four species moved randomly within the test chamber in the absence of eggs, but preferred their own eggs to a filter paper blank. One species, D. fuscus, exhibited a similar attraction to conspecific eggs suggesting that the presence of eggs, regardless of their origin, may stimulate maternal care in physiologically primed females. We found that egg discrimination is not as infallible as previously reported. All four species spent more time with their own eggs than they did with the eggs of a conspecific female; however, this trend was significant for D. ocoee only. Interspecific variation in egg discrimination is attributed to differential selection associated with the life history of individual species.     

The courtship behaviour of the spotted dusky salamander, Desmognathus fuscus conanti (Amphibia: Caudata: Plethodontidae)

The courtship behaviour of the North American southern dusky salamander, Desmognathus fuscus conanti (Plethodontidae), is described for the first time. The male is the most active participant, as is typical for caudate amphibians, and much of his behaviour can be understood in terms of orientation to and stimulation (or persuasion) of the female. The latter is largely accomplished by the transfer of courtship pheromones from the male's mental (chin) gland. The orientation and persuasion phases of courtship appear necessary if sperm transfer by means of a spermatophore deposited upon the substratum is to be successful. The courtship of D. fuscus conanti is qualitatively identical to that of D. santeetlah , with which it is sexually compatible in the laboratory and with which it may hybridize in nature. However, a broader comparison of species in the genus Desmognathus reveals that the greatest interspecific differences occur during the orientation and persuasion phases of courtship. A complete understanding of the evolution of courtship behaviour in desmognathine salamanders awaits availability of ethograms for all other taxa and a robust, independent phytogeny for the entire subfamily.     

Reinforcement shapes clines in female mate discrimination in Drosophila subquinaria

Reinforcement of species boundaries may alter mate recognition in a way that also affects patterns of mate preference among conspecific populations. In the fly Drosophila subquinaria, females sympatric with the closely related species D. recens reject mating with heterospecific males as well as with conspecific males from allopatric populations. Here, we assess geographic variation in behavioral isolation within and among populations of D. subquinaria and use cline theory to understand patterns of selection on reinforced discrimination and its consequences for sexual isolation within species. We find that selection has fixed rejection of D. recens males in sympatry, while significant genetic variation in this behavior occurs within allopatric populations. In conspecific matings sexual isolation is also asymmetric and stronger in populations that are sympatric with D. recens. The clines in behavioral discrimination within and between species are similar in shape and are maintained by strong selection in the face of gene flow, and we show that some of their genetic basis may be either shared or linked. Thus, while reinforcement can drive extremely strong phenotypic divergence, the long‐term consequences for incipient speciation depend on gene flow, genetic linkage of discrimination traits, and the cost of these behaviors in allopatry.     

Systematics of dusky salamanders, Desmognathus (Caudata: Plethodontidae), in the mountain and Piedmont regions of Virginia and North Carolina, USA

We analysed mitochondrial (cytochrome  b ) nucleotide sequences, nuclear allozyme markers, and morphometric characters to investigate species boundaries and phylogenetic relationships among dusky salamanders ( Desmognathus ) in the southern Blue Ridge and adjacent Piedmont Physiographic Provinces of Virginia and North Carolina. Our results revealed four distinct mitochondrial DNA clades that are also characterized by distinct allozyme markers. One clade consists of sequences derived from populations distributed from New England to south-western Virginia that are referable to Desmognathus fuscus Rafinesque, 1820, although there is considerable sequence and allozyme divergence within this clade. A second clade consists of sequences derived from populations referable to Desmognathus planiceps Newman, 1955, a form that we resurrect from its long synonymy under D. fuscus . Desmognathus planiceps and D. fuscus also differ in mandibular tooth shape. Two other cytochrome  b sequences recovered from populations along the Blue Ridge escarpment in southern Virginia are quite divergent from those of the previous two clades, and these populations may represent yet another distinct species. Sequences from a population in the Brushy Mountains in the Piedmont of northern North Carolina are similar to those of Desmognathus carolinensis . Population groupings indicated by allozyme data generally correspond to the cytochrome  b clades. Cryptic diversity in Appalachian desmognathan salamanders clearly requires further study. © 2008 The Linnean Society of London, Zoological Journal of the Linnean Society , 2008, 152 , 115–130.     

Males,but not females,contribute to sexual isolation between two sympatric species of <Emphasis Type="Italic">Gambusia</Emphasis>

Reproductive isolation restricts genetic exchange between species. Various pre- and post-mating barriers, such as behavior, physiology and gametic incompatibility, have been shown to evolve in sympatry. In certain scenarios, isolation can be asymmetrical, where species differentially prefer conspecifics. We examined sexual isolation via conspecific mate preference between Gambusia affinis and G. geiseri in both sexes. To investigate male contribution to sexual isolation, we compared the number of mating attempts (gonopodial thrusts) directed at either a conspecific or a heterospecific female, in both species. We also examined sperm priming and expenditure in males in the presence of conspecific or heterospecific females. We then measured female preference for either a conspecific or heterospecific male, in both species. We found that males of both species preferred to mate with conspecific females, but showed no difference in sperm production or expenditure between conspecific and heterospecific females. Females of both species did not prefer conspecific over heterospecific males. Our results suggest that sexual isolation might be mediated by male mate choice in this system and not female choice, suggesting that there is asymmetrical reproductive isolation between the sexes in G. affinis and G. geiseri, but symmetrical species isolation.     

Dusky salamanders (Desmognathus, Plethodontidae) from the Coastal Plain: multiple independent lineages and their bearing on the molecular phylogeny of the genus

Recent phylogenetic reassessment of the lungless salamanders (Plethodontidae) confirmed a major life-history reversal-from direct development to an aquatic larval stage-in the dusky salamanders (Desmognathus) of eastern North America. This reversal initiated high rates of lineage accumulation, reputedly generating the species richness and ecological breath that now characterize Desmognathus. Certain important aspects of the radiation, e.g., ecomorphological evolution, have been identified through intense sampling effort of Appalachian Highland lineages. However, the research preoccupation on montane species has left overlooked a significant component of dusky salamander distribution-the Coastal Plain. We present the first molecular phylogeny for Desmognathus to incorporate extensive coverage from the Atlantic and Gulf coastal plains. We examined 38 Coastal Plain populations in conjunction with 45 additional populations, representing 16 of the 19 nominal species. Bayesian analysis of 88 mitochondrial cox1 haplotypes diagnosed eight independent population lineages within the Coastal Plain, a number at odds with the region's three currently recognized species. Desmognathus has apparently experienced a complex biogeographic history in this physiographic region, one involving multiple invasions and several ecological transitions from lotic to lentic habitats.     

Patterns of trombiculid mite (Hannemania dunni) parasitism among plethodontid salamanders in the western Piedmont of North Carolina

Trombiculid mites are known to parasitize a variety of amphibian species, yet few comparisons of mite parasitism among amphibian species have been made. In this study, we investigated patterns of trombiculid mite parasitism among 3 plethodontid salamanders (Desmognathus fuscus, Eurycea cirrigera, and Plethodon cylindraceus) in the western Piedmont of North Carolina. All 3 salamander species were parasitized by a single species, Hannemania dunni. Desmognathus fuscus harbored mites more frequently (60.4% of individuals) than E. cirrigera (11.1%) or P. cylindraceus (14.6%). Desmognathus fuscus also had higher parasite loads than E. cirrigera or P. cylindraceus (P < 0.001). Mites on D. fuscus were found more frequently on the limbs than other body locations (P < 0.001). We found no correlation between salamander size and mite abundance (P = 0.689), but salamander collection sites influenced the abundance of mites on D. fuscus (P = 0.002). We found no effect of season on mite abundance in D. fuscus (P = 0.952). Salamander habitat preferences and edaphic or climatic differences among study sites may influence patterns of Hannemania sp. parasitism of salamanders.     

REPRODUCTIVE CHARACTER DISPLACEMENT OF EPICUTICULAR COMPOUNDS AND THEIR CONTRIBUTION TO MATE CHOICE IN DROSOPHILA SUBQUINARIA AND DROSOPHILA RECENS

Interactions between species can alter selection on sexual displays used in mate choice within species. Here we study the epicuticular pheromones of two Drosophila species that overlap partially in geographic range and are incompletely reproductively isolated. Drosophila subquinaria shows a pattern of reproductive character displacement against Drosophila recens, and partial behavioral isolation between conspecific sympatric versus allopatric populations, whereas D. recens shows no such variation in mate choice. First, using manipulative perfuming experiments, we show that females use pheromones as signals for mate discrimination both between species and among populations of D. subquinaria. Second, we show that patterns of variation in epicuticular compounds, both across populations and between species, are consistent with those previously shown for mating probabilities: pheromone compositions differ between populations of D. subquinaria that are allopatric versus sympatric with D. recens, but are similar across populations of D. recens regardless of overlap with D. subquinaria. We also identify differences in pheromone composition among allopatric regions of D. subquinaria. In sum, our results suggest that epicuticular compounds are key signals used by females during mate recognition, and that these traits have diverged among D. subquinaria populations in response to reinforcing selection generated by the presence of D. recens.     

BEHAVIORAL OBSERVATIONS OF SEXUAL ISOLATION AMONG ALLOPATRIC POPULATIONS OF THE MOUNTAIN DUSKY SALAMANDER,DESMOGNATHUS OCHROPHAEUS

The behavioral basis of sexual isolation was investigated in 11 crosses staged between individuals from nine allopatric populations of the Appalachian mountain dusky salamander, Desmognathus ochrophaeus. Scan-sampled observations of within-population (homotypic) and between-population (heterotypic) heterosexual encounters yielded the following results. 1) Fewer heterotypic encounters than homotypic encounters resulted in insemination of females. Indices of the strength of sexual isolation based on insemination data ranged from 0.26 (lowest) to 0.82 (highest), indicating that mechanisms that confer sexual isolation may evolve during allopatric differentiation. 2) Heterotypic encounters that progressed as far as the stage of courtship during which the male stimulates the female to mate did so at the same time as homotypic encounters, although the number of the former reaching this stage was lower. 3) The time interval from courtship initiation to insemination was longer in heterotypic encounters than homotypic encounters. In some crosses, males did not attempt to initiate heterotypic encounters, indicating that this sex is responsible for sexual isolation in these instances. We use our results to formulate specific hypotheses concerning the behavioral basis of sexual isolation in these salamanders, and some possible experimental approaches are suggested.     

Visual perception of light organ patterns in deep‐sea shrimps and implications for conspecific recognition

Darkness and low biomass make it challenging for animals to find and identify one another in the deep sea. While spatiotemporal variation in bioluminescence is thought to underlie mate recognition for some species, its role in conspecific recognition remains unclear. The deep‐sea shrimp genus, Sergestes sensu lato (s.l.), is one group that is characterized by species‐specific variation in light organ arrangement, providing us the opportunity to test whether organ variation permits recognition to the species level. To test this, we analyzed the visual capabilities of three species of Sergestes s.l. in order to (a) test for sexual dimorphism in eye‐to‐body size scaling relationships, (b) model the visual ranges (i.e., sighting distances) over which these shrimps can detect intraspecific bioluminescence, and (c) assess the maximum possible spatial resolution of the eyes of these shrimps to estimate their capacity to distinguish the light organs of each species. Our results showed that relative eye size scaled negatively with body length across species and without sexual dimorphism. Though the three species appear capable of detecting one another's bioluminescence over distances ranging from < 1 to ~6 m, their limited spatial resolution suggests they cannot resolve light organ variation for the purpose of conspecific recognition. Our findings point to factors other than conspecific recognition (e.g., neutral drift, phenotypic constraint) that have led to the extensive diversification of light organs in Sergestes s.l and impart caution about interpreting ecological significance of visual characters based on the resolution of human vision. This work provides new insight into deep‐sea animal interaction, supporting the idea that—at least for these mesopelagic shrimps—nonvisual signals may be required for conspecific recognition.     

Phylogenetic analysis of ecomorphological divergence, community structure, and diversification rates in dusky salamanders (Plethodontidae: Desmognathus)

An important dimension of adaptive radiation is the degree to which diversification rates fluctuate or remain constant through time. Focusing on plethodontid salamanders of the genus Desmognathus, we present a novel synthetic analysis of phylogeographic history, rates of ecomorphological evolution and species accumulation, and community assembly in an adaptive radiation. Dusky salamanders are highly variable in life history, body size, and ecology, with many endemic lineages in the southern Appalachian Highlands of eastern North America. Our results show that life-history evolution had important consequences for the buildup of plethodontid-salamander species richness and phenotypic disparity in eastern North America, a global hot spot of salamander biodiversity. The origin of Desmognathus species with aquatic larvae was followed by a high rate of lineage accumulation, which then gradually decreased toward the present time. The peak period of lineage accumulation in the group coincides with evolutionary partitioning of lineages with aquatic larvae into seepage, stream-edge, and stream microhabitats. Phylogenetic simulations demonstrate a strong correlation between morphology and microhabitat ecology independent of phylogenetic effects and suggest that ecomorphological changes are concentrated early in the radiation of Desmognathus. Deep phylogeographic fragmentation within many codistributed ecomorph clades suggests long-term persistence of ecomorphological features and stability of endemic lineages and communities through multiple climatic cycles. Phylogenetic analyses of community structure show that ecomorphological divergence promotes the coexistence of lineages and that repeated, independent evolution of microhabitat-associated ecomorphs has a limited role in the evolutionary assembly of Desmognathus communities. Comparing and contrasting our results to other adaptive radiations having different biogeographic histories, our results suggest that rates of diversification during adaptive radiation are intimately linked to the degree to which community structure persists over evolutionary time.     

Alternative reproductive tactics in male freshwater fish influence the accuracy of species recognition

Sexual conflict can result in coercive mating. Because males bear low costs of heterospecific mating, coercive males may engage in misdirected mating attempts toward heterospecific females. In contrast, sexual selection through consensual mate choice can cause mate recognition cues among species to diverge, leading to more accurate species recognition. Some species show both coercive mating and mate choice‐associated courtship behaviors as male alternative reproductive tactics. We hypothesized that if the selection pressures on each tactic differ, then the accuracy of species recognition would also change depending on the mating tactic adopted. We tested this hypothesis in the guppy (Poecilia reticulata) and mosquitofish (Gambusia affinis) by a series of choice experiments. Poecilia reticulata and G. affinis males both showed imperfect species recognition and directed all components of mating behavior toward heterospecific females. They tended to direct courtship displays more frequently toward conspecific than heterospecific females. With male P. reticulata, however, accurate species recognition disappeared when they attempted coercive copulation: they directed coercions more frequently toward heterospecific females. We also found that heterospecific sexual interaction had little effect on the fecundity of gravid females, which suggests that prepregnancy interactions likely underpin the exclusion of G. affinis by P. reticulata in our region.