Ambystoma Maculatum Research Articles

Asymmetric Introgression in a Spotted Salamander Hybrid Zone.

Before the establishment of reproductive isolation, deeply diverged intraspecific lineages can experience complex genetic and behavioral interactions as they come into secondary contact. Divergent selective and demographic processes mediate gene flow among lineages, resulting in hybrid zones with complex biogeographic structure. Discordance in the biogeographic patterns of autosomal and maternally inherited loci provides a useful window to infer the processes mediating admixture and introgression across hybrid zones. Here, we sampled 489 genotypes across a hybrid zone between 2 phylogeographic lineages of the spotted salamander, Ambystoma maculatum, and characterize discordant patterns of nuclear and mitochondrial introgression across the contact boundary. Our results indicate asymmetric introgression of nuclear DNA beyond the contact boundary from the western to eastern lineage, with introgression of eastern mitochondrial DNA into the western lineage. We discuss alternative mechanisms for this pattern and attribute this result to neutral patterns of population expansion of the western lineage into the east in combination with female mate choice for larger-bodied western males. Our results underscore the complexity of interacting mechanisms that give rise to reproductive asymmetries in the earliest stages of the speciation process.

Comparación de la morfología de los eritrocitos de Triturus marmoratus con la de otras especies de anfibios

In this work we obtained the main morphometric values of marbled newt (Triturus marmoratus) erythrocytes: major axis length (dy), minor axis length (dx), nucleus length (dn), nucleus width (an), nucleus surface (Sn) and erythrocyte surface (Se), as well as hematocrit values. These morphometric values from T. marmoratus are compared with those of several species of anurans and urodeles. With this purpose, we conduct an inter-specific comparison of the relationship between the number of erythrocytes (N) and the major axis length or the erythrocyte surface, determining that T. marmoratus has large erythrocytes, which place it within the group of species very linked to the aquatic environment like Ambystoma maculatum, Proteus anguinus, Necturus maculosus, or Cryptobranchus alleganiensis, among others.

A Test of Local Enhancement in Amphibians

AbstractLocal enhancement is an underexplored social learning mechanism that is often observed in organisms that live in groups. This mechanism occurs when individuals are attracted to areas where conspecifics have previously been, but which are not present when the animal actually moves into the area. We tested for local enhancement in wood frog tadpoles (Lithobates sylvatica) and spotted salamander larvae (Ambystoma maculatum) in three experiments that exposed individuals to one side of a test chamber which was empty and another that contained a group of three conspecifics. Side preference of the focal individual was recorded once the conspecifics were removed. Tadpoles showed a clear preference for moving to areas where a group of tadpoles had previously been located. Conversely, this preference was not observed in salamander larvae. In addition, salamander larvae took significantly more time to initially choose a side. These results indicate that tadpoles exhibit local enhancement, whereas aquatic salamander larvae do not. This difference in social learning could be largely due to differences in aquatic ecology between tadpoles and salamander larvae.

Life-history costs of reproductive behaviors in a wetland-breeding amphibian

Studies examining the association between life-history strategies and behavior can help inform our understanding of the fitness consequences of reproductive behaviors. Breeding migrations are common in numerous taxa and often represent an important reproductive cost. We examined body mass changes of spotted salamanders (Ambystoma maculatum) during their seasonal breeding migrations to determine the influence of breeding behaviors on reproductive costs. We sampled salamanders entering and exiting a 0.92-ha ephemeral wetland in the North Carolina Piedmont using a 400-m drift fence completely encircling the wetland. Traps were checked daily during annual breeding migrations in mid-January to early March of 2013 and 2014. The relationship between body mass changes and explanatory factors including sex, duration of stay at wetland, and distance between site entrance and exit were examined using a set of candidate linear mixed models selected a priori and compared using Akaike's information criterion. Female salamanders lost a greater percentage of body mass during the time in the wetland than males as a result of oviposition. Model selection identified duration of stay as the most important predictor of female body mass change with greater duration of stay associated with lower mass loss. In males, percentage mass loss was most closely associated with site entrance–exit correspondence with greater distance between entrance and exit sites associated with greater mass loss. These data demonstrate both the importance of wetland entrance–exit sites and duration of stay at the wetland in determining mass change during spotted salamander reproduction and also that these relationships can differ as a function of sex. Taken together, our findings provide a potential link between behavior, life-history cycles, and costs experienced during reproduction.

Phylogenetic analysis of algal symbionts associated with four North American amphibian egg masses.

Egg masses of the yellow-spotted salamander Ambystoma maculatum form an association with the green alga “Oophila amblystomatis” (Lambert ex Wille), which, in addition to growing within individual egg capsules, has recently been reported to invade embryonic tissues and cells. The binomial O. amblystomatis refers to the algae that occur in A. maculatum egg capsules, but it is unknown whether this population of symbionts constitutes one or several different algal taxa. Moreover, it is unknown whether egg masses across the geographic range of A. maculatum, or other amphibians, associate with one or multiple algal taxa. To address these questions, we conducted a phylogeographic study of algae sampled from egg capsules of A. maculatum, its allopatric congener A. gracile, and two frogs: Lithobates sylvatica and L. aurora. All of these North American amphibians form associations with algae in their egg capsules. We sampled algae from egg capsules of these four amphibians from localities across North America, established representative algal cultures, and amplified and sequenced a region of 18S rDNA for phylogenetic analysis. Our combined analysis shows that symbiotic algae found in egg masses of four North American amphibians are closely related to each other, and form a well-supported clade that also contains three strains of free-living chlamydomonads. We designate this group as the ‘Oophila’ clade, within which the symbiotic algae are further divided into four distinct subclades. Phylogenies of the host amphibians and their algal symbionts are only partially congruent, suggesting that host-switching and co-speciation both play roles in their associations. We also established conditions for isolating and rearing algal symbionts from amphibian egg capsules, which should facilitate further study of these egg mass specialist algae.

Photosynthetic carbon from algal symbionts peaks during the latter stages of embryonic development in the salamander Ambystoma maculatum.

BackgroundIt was recently discovered that symbiotic algae in the eggs of the salamander Ambystoma maculatum translocate fixed carbon from photosynthesis to developing embryos. Fixed carbon translocation was shown in embryos at one time point during development, however, it was unknown if fixed carbon translocation occurs throughout all developmental stages.FindingsIn this study, fixed carbon translocation was measured in salamander eggs at six time points over the latter half of development. Fixed carbon translocation did not occur until the middle tailbud portion of development (stages 26–30), and translocation was measured in 20% or less of eggs sampled. Peak carbon translocation occurred during the late tailbud phase of development (stages 31–35), where as much as 87% of eggs sampled showed translocation, and average percent translocation was 6.5%. During the final stages of development, fixed carbon translocation declined, and translocation was not detected in embryos five days prior to hatching.ConclusionsThe onset of fixed carbon translocation from Oophila to A. maculatum embryos during the second half of embryonic development is likely due to the corresponding settlement and concentration of Oophila in the inner egg envelope. In addition, carbon translocation ceases in late stage embryos as the inner egg envelope thins and ruptures in preparation for hatching.

Landscape context influences the abundance of amphibians and the strength of their food web interactions in small ponds

Many species, including most amphibians, undergo an ontogenetic niche shift (ONS) from an aquatic larval stage to a terrestrial adult stage. We use the ratio of aquatic to terrestrial habitat in a landscape as a tool to understand the influence of landscape context on the population growth of ONS species. The aquatic to terrestrial ratio (ATR) of habitats can be viewed as an analog to the influence of resource ratios on the population growth of consumers and depends on the degree to which each habitat type limits the growth of a given population. Population growth rates of shorter‐lived species tend to be more limited by demographic rates in early (aquatic) life stages. As a result, increasing the ATR should lead to a higher total population size in the landscape (and higher densities in the terrestrial habitat), but have little influence on the density of individuals in any given aquatic habitat. Alternatively, population growth rates of longer‐lived species tend to be more limited by demographic rates in later (terrestrial) life stages and increasing the ATR should have little influence on the total population size in the landscape, but decrease the density of individuals in any given aquatic habitat. We show that among‐landscape variation in the breeding‐pond densities of three widespread amphibians with contrasting life histories is consistent with this framework. Within‐pond densities of Pseudacris crucifer, a species with short‐lived adults, were not influenced by ATR, whereas within‐pond densities of Hyla versicolor, a longer‐lived member of the same family (Hylidae), declined as ATR increased. Ambystoma maculatum, a long‐lived salamander, also had lower densities in ponds with higher ATR. Because A. maculatum larvae are important predators in ponds, we use structural equation modeling to show that landscape context (ATR) can moderate community structure via direct (amphibian abundances) and indirect (prey species richness) effects.

Effects of fine-scale forest habitat quality on movement and settling decisions in juvenile pond-breeding salamanders.

A better understanding of how individuals respond to variation in habitat quality while moving through heterogeneous habitats is needed to predict ecological phenomena at larger scales, such as local population and metapopulation dynamics. We sought to identify how fine-scale habitat quality affects the decisions of juvenile pond-breeding salamanders (Ambystoma maculatum and A. annulatum) to cease dispersive movements away from their natal pond, select a refuge, and settle. Because of the acute susceptibility of juvenile amphibians to evaporative water loss in terrestrial habitats, we predicted that they possess mechanisms for adjusting their behavior in response to variations in fine-scale habitat quality. We used experimental field enclosures to isolate the effects of habitat quality on settling behavior and employed generalized linear mixed models to examine how manipulations in canopy cover (closed or open) and microhabitat (control, compacted soils, high coarse woody debris, high burrow density), along with environmental variables (rainfall and air temperature), affect the individual's probability of settling. Our results indicated that A. maculatum and A. annulatum had a 10% and 30% decreased probability of settling in open-canopy clearcut habitat, respectively, compared to closed-canopy forest habitat. In addition, A. annulatum were 24% less likely to settle in compacted soil treatments. Although the settlement probability of A. annulatum did not depend on refuge availability, A. maculatum were 18% and 25% more likely to settle under conditions of high burrow density and high coarse woody debris, respectively. These findings make a unique contribution to our understanding of amphibian movement ecology by demonstrating how the interplay of external factors and individual behavior produce observed patterns of movement and habitat selection.

Landmark learning by juvenile salamanders (Ambystoma maculatum)

Learning to use a landmark as a beacon to locate resources is one of the simplest forms of spatial learning. We tested whether landmark learning occurs in a semifossorial salamander that migrates annually to breeding ponds as adults. Juvenile spotted salamanders (Ambystoma maculatum) were tested in square containers with a plastic feeding dish in each corner, and a piece of earthworm was placed in one randomly-chosen dish. For landmark-trained salamanders, a rock was placed beside the dish containing the prey. For control salamanders, the rock was placed beside a randomly selected feeding dish. Each salamander was trained once every 2 days for 30 days. Significantly more landmark-trained salamanders than control salamanders entered the landmark area first, and landmark-trained individuals had faster latencies to enter the landmark area and longer stay-times. These results suggest that spotted salamanders are able to locate resources by associating their positions with landmarks.

Optimization of culturing conditions for toxicity testing with the alga Oophila sp. (Chlorophyceae), an amphibian endosymbiont.

Eggs of the yellow-spotted salamander (Ambystoma maculatum) have a symbiotic relationship with green algae. It has been suggested that contaminants that are preferentially toxic to algae, such as herbicides, may impair the symbiont and, hence, indirectly affect the development of the salamander embryo. To enable testing under near-standard conditions for first-tier toxicity screening, the authors isolated the alga from field-collected eggs and identified conditions providing exponential growth rates in the apparent asexual phase of the alga. This approach provided a uniform, single-species culture, facilitating assessment of common toxicity end points and comparison of sensitivity relative to other species. Sequencing of the 18s ribosomal DNA indicated that the isolated alga is closely related to the recently described Oophila amblystomatis but is more similar to other known Chlamydomonas species, suggesting possible biogeographical variability in the genetic identity of the algal symbiont. After a tiered approach to culturing method refinement, a modified Bristol's media with 1 mM NH4 (+) as nitrogen source was found to provide suitable conditions for toxicity testing at 18 °C and 200 µmol m(-2) s(-1) photosynthetically active radiation (PAR) on a 24-h light cycle. The validity of the approach was demonstrated with Zn(2+) as a reference toxicant. Overall, the present study shows that screening for direct effects of contaminants on the algal symbiont without the presence of the host salamander is possible under certain laboratory conditions.

The roles of oxygen and ammonia in the symbiotic relationship between the spotted salamander Ambystoma maculatum and the green alga Oophila amblystomatis during embryonic development

Some of the positive benefits of the symbiotic relationship between the yellow spotted salamander Ambystoma maculatum and the green alga Oophila amblystomatis on salamander embryonic development have been well documented. However, the fundamental basis of the relationship, in terms of the exchange of metabolic and photosynthetic products, is under debate, especially as it relates to newly discovered endosymbiotic aspects of the relationship. We developed a novel experimental approach in which we liberated egg capsules from egg masses and compared de-capsulated embryos to capsulated embryos. We were able to discern the contribution of salamander embryo respiration and ammonia excretion and algal photosynthesis to oxygen and ammonia dynamics within the egg capsules under light and dark conditions. Algal photosynthesis drove oxygen production under light conditions, while salamander respiration caused hypoxia during dark conditions. Ammonia was consumed to such an extent that it may be the limiting factor for algal growth. Algal cells actively penetrated the developing gut of embryos through the blastopore from an early developmental stage, and some algae came to reside in the vicinity of liver primordia towards later embryonic development, perhaps explaining the lack of detectable ammonia production by de-capsulated embryos. We suggest that oxygen and ammonia may influence A. maculatum growth and O. amblystomatis population dynamics. We have also demonstrated the extent to which algal cells penetrate internal spaces of embryos, which may be the precursor to endosymbiosis between these two species.

Development of 31 new microsatellite loci for two mole salamanders (Ambystoma laterale and A. jeffersonianum)

Ambystoma salamanders are amphibians that due to limited dispersal abilities and reliance on wetlands for breeding are susceptible to population declines and local extinctions (Blaustein et al. 2011). Species identification within Ambystoma is especially difficult due to the presence of unisexual Ambystoma that consist of multiple all-female lineages in which clones can have between two and five nuclear genomes from up to five other Ambystoma species (Bogart et al. 2007). The majority of these unisexual Ambystoma are composed of nuclear genomes from two species, A. laterale (Blue Spotted Salamander) and A. jeffersonianum (Jefferson Salamander). We developed species-specific microsatellite markers for these two species as a tool for the identification and investigation of the genetic interactions between sexual and unisexual groups in areas where either sexual species is endangered or of special conservation concern (Ohio, Indiana, and Ontario).

Population of origin and environment interact to determine oomycete infections in spotted salamander populations

Spatial variation in disease risk in wild populations can depend both on environmental and genetic factors. Understanding the various contributions of each factor requires experimental manipulation of both the environment and genetic composition of populations under natural field conditions. We first examined natural patterns of oomycete composition and infection in the eggs of 13 populations of the spotted salamander Ambystoma maculatum. We then performed a fully factorial field transplant of the eggs of six populations to separate the contributions from population of origin and the environment on oomycete resistance in spotted salamanders. Among wild ponds, we found strong variation in oomycete infections in spotted salamander populations and differences in the composition of oomycete communities. In transplant experiments, salamander populations differed in their resistance to oomycete infections via a significant interaction between population of origin and environment. However, not all populations were locally adapted to local conditions. One population was significantly adapted to its home environment, and another one was significantly maladapted. These population effects could originate from differential adaptation of salamander populations to local oomycete communities or environmental conditions that mediate resistance, local adaptation and maladaptation of oomycetes to hosts, or from maternal transmission. Accounting for both environment and population of origin will often be necessary to understand disease dynamics in wild populations.

Six New Host Records forMesocestoidessp. Tetrathyridia (Cestoidea: Cyclophyllidea) from Amphibians and Reptiles of Arkansas, U.S.A

ABSTRACT: Forty-two individual amphibians and 37 reptiles from Arkansas, U.S.A., were examined for tetrathyridia of the cyclophyllidean cestode genus, Mesocestoides. The following herpetofauna were found to be infected with encapsulated or free tetrathyridia of this tapeworm, 1 of 25 (4%) spotted salamanders, Ambystoma maculatum; 1 of 17 (6%) western slimy salamanders, Plethodon albagula; 1 of 21 (5%) 5-lined skinks, Plestiodon fasciatus; 1 of 5 (20%) black racers, Coluber constrictor priapus; 1 of 10 (10%), western rat snakes, Pantherophis obsoletus; and 1 of 1 (100%) eastern garter snake, Thamnophis sirtalis sirtalis. We report 6 new host records for Mesocestoides sp., including the fifth and sixth report of this parasite from caudate amphibians of the world. This is also only the second time this tapeworm has been reported from snakes of the genus Thamnophis.

Response of the green alga Oophila sp., a salamander endosymbiont, to a PSII‐inhibitor under laboratory conditions

In a rare example of autotroph-vertebrate endosymbiosis, eggs of the yellow-spotted salamander (Ambystoma maculatum) are colonized by a green alga (Oophila sp.) that significantly enhances salamander development. Previous studies have demonstrated the potential for impacts to the salamander embryo when growth of the algae is impaired by exposure to herbicides. To further investigate this relationship, the authors characterized the response of the symbiotic algae (Oophila sp.) alone to the photosystem II (PSII) inhibitor atrazine under controlled laboratory conditions. After extraction of the alga from A. maculatum eggs and optimization of culturing conditions, 4 toxicity assays (96 h each) were conducted. Recovery of the algal population was also assessed after a further 96 h in untreated media. Average median effective concentration (EC50) values of 123 µg L(-1) (PSII yield), 169 µg L(-1) (optical density), and 299 µg L(-1) (growth rate) were obtained after the 96-h exposure. Full recovery of exposed algal populations after 96 h in untreated media was observed for all endpoints, except for optical density at the greatest concentration tested (300 µg L(-1) ). Our results show that, under laboratory conditions, Oophila sp. is generally less sensitive to atrazine than standard test species. Although conditions of growth in standard toxicity tests are not identical to those in the natural environment, these results provide an understanding of the tolerance of this alga to PSII inhibitors as compared with other species.

No evidence of natal habitat preference induction in juveniles with complex life histories

Individual variation in physiology and behaviour can have strong effects on ecological and evolutionary processes. Natal habitat, one source of individual variation, can influence individual phenotype, behaviour and fitness through effects on eventual habitat selection. Natal habitat preference induction occurs when individuals match stimuli in their selected habitat to those of their natal habitat. Natal habitat can also affect habitat selection through its influence on body condition (silver spoon effect). We tested for natal habitat preference induction and body-condition-dependent habitat selection in two species with complex life histories, the spotted salamander, Ambystoma maculatum, and the small-mouthed salamander, Ambystoma texanum. We reared salamanders from hatchlings in mesocosms with leaf, grass or control substrate, and tested juvenile habitat selection through two behavioural assays. We found weak evidence of larger salamanders having decreased latency and sampling more habitats, lending support to the body-condition-dependent habitat selection hypothesis in these species. Juveniles preferred grass litter cues regardless of the substrate in their natal mesocosm, suggesting natal habitat preference induction may not occur in species with complex life histories. We propose that species with complex life histories use simple movement rules, such as moving along habitat gradients, to select postnatal habitat when moving through a novel environment.

The Spots of the Spotted Salamander Are Sexually Dimorphic

Brilliant and conspicuous coloration in animals is often thought to signal quality to conspecifics (sexual selection) or to signal unpalatability to predators (aposematic selection). Ambystoma maculatum (spotted salamanders) have bilateral rows of conspicuous bright yellow dorsal spots against dark skin. Although this coloration has been long thought a classic example of warning coloration, to date, there are no quantitative measures of spot coloration. We captured adult male and female A. maculatum as they entered breeding ponds and measured body condition, spectral reflectance of the yellow spots, spot size, and coverage of the dorsal area in spots. We found evidence of sexual dichromatism; males had a larger area of their dorsum covered in spots, and tended to have more chromatic, but less bright yellow color. Moreover, our data suggest a tradeoff between coloration and body condition. Animals in better body condition expressed lower chroma and those with greater spot coverage expressed brighter spots ...

Dynamics of the growth, life history transformation and photosynthetic capacity of Oophila amblystomatis (Chlorophyceae), a green algal symbiont associated with embryos of the northeastern yellow spotted salamander Ambystoma maculatum (Amphibia)

The recent discovery that the unicellular green alga Oophila amblystomatis, invades embryonic tissues and cells of the salamander Ambystoma maculatum prompted us to investigate the growth and life history transformations of the algal symbionts in egg capsules. During embryonic development, symbionts were first detected microscopically as a cohesive population of swimming cells in the vicinity of the blastopore around embryonic stage 17. This population of cells grew and at embryonic stage 25, a fraction of the population began to affix to the inside of the egg capsule. Cells then underwent syngamy, lost flagella, and transformed into non-motile cells. We observed a linear increase in the accumulation of such capsule-associated cells from embryonic stage 25 to 40. The population of zoospores did not grow over this period and showed a declining trend between stage 39 and 40. We verified the population growth by measuring relative chlorophyll a content and also measured quantum yield (QY) of photosystem II (PS II) using pulse amplitude modulated (PAM) fluorometry. The population, but not the cell size, of non-motile capsule membrane-bound cells increased modestly during a one-month period after hatching, and continued to contain high levels of chlorophyll a and photosynthetic capacity. We conclude that O. amblystomatis undergoes a life history transition in egg capsules and speculate that many of these symbionts become zygotes, rather than invading the embryo.

Going green with algae

Going green with algae

Modifications of the genital kidney proximal and distal tubules for sperm transport inNotophthalmus viridescens(Amphibia, Urodela, Salamandridae)

Male salamanders use nephrons from the genital kidney to transport sperm from the testicular lobules to the Wolffian duct. The microstructure of the epithelia of the genital kidney proximal tubule and distal tubule was studied over 1 year in a population of Notophthalmus viridescens from Crawford and Pike counties in central Missouri. Through ultrastructural analysis, we were able to support the hypothesis that the genital kidney nephrons are modified to aid in the transportation of sperm. A lack of folding of the basal plasma membrane, in both the genital kidney proximal and distal tubules when compared to the pelvic kidney proximal and distal tubules, reduces the surface area and thus likely decreases the efficiency of reabsorption in these nephron regions of the genital kidney. Ciliated epithelial cells are also present along the entire length of the genital kidney proximal tubule, but are lacking in the epithelium of the pelvic kidney proximal tubule. The exact function of these cilia remains unknown, but they may aid in mixing of seminal fluids or the transportation of immature sperm through the genital kidney nephrons. Ultrastructural analysis of proximal and distal tubules of the genital kidney revealed no seasonal variation in cellular activity and no mass production of seminal fluids throughout the reproductive cycle. Thus, we failed to support the hypothesis that the cellular activity of the epithelia lining the genital kidney nephrons is correlated to specific events in the reproductive cycle. The cytoplasmic contents and overall structure of the genital and pelvic kidney epithelial cells were similar to recent observations in Ambystoma maculatum, with the absence of abundant dense bodies apically in the epithelial cells lining the genital kidney distal tubule.