Effects of road salt, egg predation and alterations to the egg-mass jelly layers on the embryos of spotted salamander Ambystoma maculatum

The field of toxicology has traditionally assessed the risk of contaminants by using laboratory experiments and a range of pesticide concentrations that are held constant for short periods of time (1-4 days). From these experiments, one can estimate the concentration that causes no effect on survival. However, organisms in nature frequently experience multiple, applications of pesticides over time rather than a single constant concentration. In addition, organisms are embedded in ecological communities that can propagate indirect effects through a food web. Using outdoor mesocosms, we examined how low concentrations (10-250 microg/L) of a globally common insecticide (malathion) applied at various amounts, times, and frequencies affected aquatic communities containing zooplankton, phytoplankton, periphyton, and larval amphibians (reared at two densities) for 79 days. All application regimes caused a decline in zooplankton, which initiated a trophic cascade in which there was a bloom in phytoplankton and, in several treatments, a subsequent decline in the competing periphyton. The reduced periphyton had little effect on wood frogs (Rana sylvatica), which have a short time to metamorphosis. However, leopard frogs (Rana pipiens) have a longer time to metamorphosis, and they experienced large reductions in growth and development, which led to subsequent mortality as the environment dried. Hence, malathion (which rapidly breaks down) did not directly kill amphibians, but initiated a trophic cascade that indirectly resulted in substantial amphibian mortality. Importantly, repeated applications of the lowest concentration (a "press treatment" consisting of seven weekly applications of 10 microg/L) caused larger impacts on many of the response variables than single "pulse" applications that were 25 times as great in concentration. These results are not only important because malathion is the most commonly applied insecticide and is found in wetlands, but also because the mechanism underlying the trophic cascade is common to a wide range of insecticides, offering the possibility of general predictions for the way in which many insecticides impact aquatic communities and the populations of larval amphibians.

A common challenge that oviparous animals face is securing survivorship during the vulnerable embryonic stage. One of the parental investment strategies to improve survivorship is providing physical structures to protect the embryos. In amphibians, there is a notable diversity in jelly‐layer structures surrounding eggs. Previous studies show that these jelly layers provide eggs with protection against egg predators, egg pathogens, and desiccation. However, few studies examined the cost–benefit relationship of the jelly‐layer structures. By using the predator–prey interaction between wood frog (Lithobates sylvaticus) tadpoles and spotted salamander (Ambystoma maculatum) eggs as a model system, we tested three hypotheses: (1) having the outer jelly layers would be costly to the embryos, (2) the relative benefit of the structural egg defense would become apparent and increase as the intensity of egg predation increases, and (3) a certain degree of predation would increase the hatching success of salamander embryos by mechanically thinning the thick outer jelly layers and increasing oxygen diffusion throughout an egg mass. To test these hypotheses, we conducted a factorial experiment in which we crossed four egg‐predation levels with two jelly‐layer conditions, intact or removed. We found that the jelly layers were essential in protecting spotted salamander embryos from wood frog tadpoles but that the associated cost was apparent in no‐predation treatments. The differential survivorship between intact eggs and eggs without jelly layers showed that the fitness advantage of jelly layers increased as the level of predation increased. Finally, the hatching success of intact egg masses was highest under the high predation conditions. These results imply that the evolution of the jelly‐layer thickness occurred under constant egg‐predation pressure. Given this predator–prey coevolution, egg predators may play a critical role in improving the hatching success of salamander embryos under certain conditions.

Intraguild predation (IGP) is common in most communities, but many aspects of density-dependent interactions of IG predators with IG prey are poorly resolved. Here, we examine how the density of an IG predator can affect feeding group size, IG egg predation, and the growth responses of IG prey. We used laboratory feeding trials and outdoor mesocosm experiments to study interactions between a social intraguild predator (larvae of the wood frog; Rana sylvatica) and its prey (spotted salamander; Ambystoma maculatum). Larvae of R. sylvatica could potentially affect A. maculatum by consuming shared larval food resources or by consuming eggs and hatchlings. However, successful egg predation requires group feeding by schooling tadpoles. We established from five to 1,190 hatchlings of R. sylvatica in mesocosms, then added either 20 A. maculatum hatchlings to study interspecific competition, or a single egg mass to examine IGP. Crowding strongly suppressed the growth of R. sylvatica, and IGP was restricted to the egg stage. In the larval competition experiment, growth of A. maculatum was inversely proportional to R. sylvatica density. In the predation experiment, embryonic mortality of A. maculatum was directly proportional to the initial density of R. sylvatica and the mean number of tadpoles foraging on egg masses. IGP on eggs reduced A. maculatum hatchling density, which accelerated larval growth. Surprisingly, the density of R. sylvatica had no overall effect on A. maculatum growth because release from intraspecific competition via egg predation was balanced by increased interspecific competition. Our results demonstrate that the density of a social IG predator can strongly influence the nature and intensity of interactions with a second guild member by simultaneously altering the intensity of IGP and intra- and interspecific competition.

Previous research on habitat associations of pond-breeding amphibians has used community assemblages as response variables because the intensive labor required to quantify population size is usually prohibitive. However, wood frogs (Rana sylvatica) and spotted salamanders (Ambystoma maculatum) oviposit egg masses that can be surveyed rapidly; thus, we were able to quantify the influence of within-pond parameters on their annual breeding effort. During 2000–2001, we assessed the effects of hydroperiod, within-pond vegetation, canopy closure, hydrologic isolation, fish occurrence, and pond size on egg mass counts of wood frogs and spotted salamanders at 124 ponds in western Rhode Island. Study sites were stratified by road density, which served as an indirect measure of non-breeding habitat quality and quantity. Hydroperiod had a significant influence on annual breeding effort. Egg masses of both species were most abundant in seasonal ponds that dried between 1 August and 30 November. Breeding populations of spotted salamanders were smaller in ponds that dried before the end of July, whereas breeding populations of wood frogs were reduced in ponds that did not dry during our two-year study. Both species usually attached egg masses to shrubs or branches in shallow water near the water surface, which may explain why both species oviposited more egg masses in ponds with substantial vegetation complexity, including extensive coverage by shrubs and persistent non-woody vegetation. Although wood frogs were more likely than spotted salamanders to avoid ovipositing in ponds with fish, egg mass counts of both species were substantially reduced in ponds containing fish. Numbers of egg masses for spotted salamanders and wood frogs were similar between larger ponds and those as small as 0.05 ha and 0.15 ha, respectively. Previously, researchers proposed protecting isolated wetlands > 0.2 ha to sustain populations of pond-breeding amphibians, but our research suggests that wetlands as small as 0.05 ha may provide critical breeding habitat for some species in southern New England.

Egg-Laying Activities and Reproductive Traits in Females of Oplurus cuvieri cuvieri

Deicing agents, primarily road salt, are applied to roads in 26 states in the United States and in a number of European countries, yet the scale of impacts of road salt on aquatic organisms remains largely under-studied. The issue is germane to amphibian conservation because both adult and larval amphibians are known to be particularly sensitive to changes in their osmolar environments. In this study, we combined survey, experimental, and demographic modeling approaches to evaluate the possible effects of road salt on two common vernal-pond-breeding amphibian species, the spotted salamander (Ambystoma maculatum) and the wood frog (Rana sylvatica). We found that in the Adirondack Mountain Region of New York (USA), road salt traveled up to 172 m from the highway into wetlands. Surveys showed that egg mass densities of spotted salamanders (A. maculatum) and wood frogs (R. sylvatica) were two times higher in forest pools than roadside pools, but this pattern was better explained by road proximity than by increased salinity. Experiments demonstrated that embryonic and larval survival were reduced at moderate (500 muS) and high conductivities (3000 muS) in A. maculatum and at high conductivities in R. sylvatica. Demographic models suggest that such egg and larval stage effects of salt may have important impacts on populations near roads, particularly in the case of A. maculatum, for which salt exposure may lead to local extinction. For both species, the effect of road salt was dependent upon the strength of larval density dependence and declined rapidly with distance from the roadside, with the greatest negative effects being limited to within 50 m. Based on this evidence, we argue that efforts to protect local populations of A. maculatum and R. sylvatica in roadside wetlands should, in part, be aimed at reducing application of road salt near wetlands with high conductivity levels.

Wildlife biologists and managers are concerned about the effects of forest fragmentation and habitat loss on pond-breeding amphibian populations. Most research has assessed the effects of habitat composition at multiple spatial scales on the presence or absence of amphibians at breeding ponds. We were interested in the effects of habitat characteristics on amphibian population size and used Wood Frog (Rana sylvatica) and Spotted Salamander (Ambystoma maculatum) egg mass counts as an index. Between 2001 and 2005, we monitored 65 seasonal ponds within forested landscapes in the Pawcatuck River watershed of Rhode Island. Both species were detected in at least 88% of the ponds sampled. Egg mass counts for both species were highest in ponds that usually dried between early October and late November. Wood Frog egg mass numbers were positively associated with pond hydroperiod, size, and depth; location on glacial fluvial deposits; the extent of persistent nonwoody plant cover in ponds; and the area of upland forest within 1 km of the pond edge. Egg mass numbers were negatively associated with location on alluvium or dense till deposits, percent canopy cover, the number of buildings within 1 km, and the area of residential development within 1 km. Spotted Salamander egg mass counts were positively associated with pond hydroperiod, size, and depth, and upland forest area within 1 km. They were negatively related to location on alluvium. Multivariate models developed from within-pond variables explained more variation in egg mass counts for both species than those developed from landscape-level factors, but the best combined models suggested that habitat characteristics at both scales are useful in the prediction of breeding effort at individual sites. Given the continuing urbanization of southern New England and the ineffectiveness of wetland regulations in protecting required terrestrial habitat around seasonal ponds, proactive techniques also are required to assure the maintenance of pond-breeding amphibian populations.

Summary Egg predation is seldom considered in life‐history studies of freshwater insects, but could be an important source of mortality with potential to limit population numbers. Costs of egg predation to prey can be considered at two levels: (i) fitness costs to individuals via reduced reproductive output; and (ii) population costs via reduced recruitment of benthic larvae. Larvae of Orthotrichia armata (Trichoptera: Hydroptilidae) feed on the egg masses of caddisflies in the Little River, central Victoria, Australia, but the small size of O. armata suggests that they may be unable to penetrate egg masses that are covered with a thick layer of spumaline jelly. We predicted that predation by O. armata would be more severe for egg masses with thinner spumaline. With field surveys throughout the peak caddisfly oviposition period (summer), we compared predation by O. armata on the egg masses of nine caddisfly taxa. Egg masses were categorised, according to the thicknesses of spumaline, into bulbous (thickly jellied), thinly jellied and jelly‐free morphotypes, with three species per morphotype. Bulbous egg masses were rarely consumed. Thinly jellied egg masses comprised just 4–13% of all egg masses in each survey, but attracted 52–93% of all egg predators. Jelly‐free egg masses were readily consumed, but O. armata were disproportionately scarce on them. In a second survey, we compared the proportions of eggs that were consumed from egg masses with small and large clutch sizes, by monitoring the thinly jellied egg masses of Taschorema sp. (598 ± 95 eggs per mass) and Ethochorema turbidum (1195 ± 192 eggs per mass) to eclosion. More than 25% of Taschorema sp. egg masses were entirely consumed, causing complete reproductive failure of parent females (fitness costs to individuals). Consumption rates were < 25% for most egg masses of E. turbidum. Orthotrichia armata consumed 48% of all Taschorema sp. eggs and 20% of E. turbidum eggs, suggesting that egg predation may substantially curb larval recruitment (costs to populations), particularly for species with small clutch sizes. Our results demonstrate that egg mortality varies for aquatic insect species that package their eggs in different ways. Combined with recent data from the same system, our results suggest egg mass morphology–mortality trade‐offs, with bulbous egg masses particularly vulnerable to flow forces (shear) and thinly jellied/jelly‐free egg masses particularly vulnerable to predation. Given the wide array of aquatic invertebrate species that lay single egg masses on hard substrata, it is feasible that this is a common trade‐off that warrants more research on the impacts of egg predation on aquatic species.

Ranaviruses have been associated with rising numbers of mass die-offs in amphibian populations around the globe. However, most studies on ranaviruses to date focused on larval amphibians. To assess the role of postmetamorphic amphibians in the epidemiology of ranaviruses and to determine the role of viral immune-suppression genes, we performed a bath-exposure study on post-metamorphic wood frogs ( Rana sylvatica) using environmentally relevant concentrations of wild-type Frog virus 3 (WT FV3), and a gene-knockout mutant (KO FV3), deficient for the putative immune-suppression gene vIF-2α. We observed a 42% infection rate and 5% mortality across the virus challenges, with infection rates and viral loads following a dose-dependent pattern. Individuals exposed to the knockout variant exhibited significantly decreased growth and increased lethargy compared with wild-type treatments. Although 85% of exposed individuals exhibited common signs of ranavirosis throughout the experiment, most of these individuals did not exhibit signs of infection by 40 d post-exposure. Overall, we showed that even a single short time exposure to environmentally relevant concentrations of ranavirus may cause sublethal infections in postmetamorphic amphibians, highlighting the importance of this life stage in the epidemiology of ranaviruses. Our study also supports the importance of the vIF-2α gene in immune-suppression in infected individuals.

Annual counts of egg masses have been promoted as an appropriate state variable for monitoring populations of some amphibian species. However, if some egg masses are not detected and detectability changes over time, the use of egg mass counts is unreliable. Variation in counts of egg masses may be indicative of variation in actual abundance or variation in detectability. I used closed capture–recapture models to estimate detection probability and evaluate potential sources of variation in the detectability of Wood Frog (Rana sylvatica) egg masses in a pond in Rocky Mountain National Park in 2003 and 2004. Model selection results and model-averaged estimates provided evidence that detection probability varied between years. However, I found no evidence of variation between observers within each year. The results of this study provide additional evidence that detection probability needs to be accounted for if egg mass counts are to be used to infer abundance and population trends in amphibian popul...

With rare exceptions, anuran larvae have traditionally been considered to occupy lower trophic levels in aquatic communities where they function as microphagous suspension feeders. This view is being challenged by studies showing that tadpoles with generalized morphology often function as macrophagous predators. Here, we review the literature concerning macrophagy by tadpoles and provide two additional examples involving generalized tadpoles. In the first, we demonstrate with laboratory and field experiments that wood frog (Rana sylvatica) tadpoles are major predators of macroinvertebrates in ponds. In the second, we show that green frog (R. clamitans) tadpoles can cause catastrophic reproductive failure of the wood frog via egg predation. These results and data from other studies challenge the assumption that generalized tadpoles function as filter-feeding omnivores, and question the general applicability of community organization models which assume that predation risk increases with pond permanence. We suggest that predation risk is greater in temporary ponds than in more permanent ponds for many organisms that are vulnerable to predation by tadpoles. This being so, a conditional model based upon interactions that are species-specific, life-stage-specific, and context-dependent may better explain community organization along hydrological gradients than models which assume that temporary ponds have few or no predators.

Many amphibian populations are in decline worldwide. Surprisingly, few studies have examined how such declines may benefit mosquitoes. Amphibian larvae may compete with and prey upon mosquito larvae, and may alter oviposition habitat selection (OHS) of mosquito adults. However, often overlooked, observed among-pool egg distributions attributed to OHS may additionally or alternatively be explained by egg predation. Temporary pools of mountainous areas of the Mediterranean serve as larval habitat for both the mosquito, Culiseta longiareolata, and the salamander, Salamandra infraimmaculata. We found Culiseta larvae and egg rafts to be highly vulnerable to predation by pre-metamorphosing Salamandra larvae, but not to metamorphosing ones. In outdoor mesocosm experiments, oviposition avoidance by Culiseta females in response to caged Salamandra was not demonstrated regardless of salamander developmental stage. Egg raft abundance was significantly reduced in free-roaming, pre-metamorphosing Salamandra but not by metamorphosing ones. Thus, Salamandra larvae may have little deterrence on Culiseta oviposition. Instead, fewer egg rafts are attributed largely to egg predation. This study highlights the importance of egg raft predation in addition to OHS when interpreting the influence of predators on prey egg distributions. It also highlights that a cost of declining amphibian populations is their reduced impacts on mosquito populations.

The amount of genetic diversity within a population can affect ecological processes at population, community, and ecosystem levels. However, the magnitude, consistency, and scope of these effects are largely unknown. To investigate these issues, we conducted two experiments manipulating the amount of genetic diversity and environmental factors in larval amphibians. The first experiment manipulated wood frog genetic diversity, the presence or absence of caged predators, and competition from leopard frogs to test whether these factors affected survival, growth, and morphology of wood frogs and leopard frogs. The second experiment manipulated wood frog genetic diversity, the presence or absence of uncaged predators, and resource abundance to test whether these factors affected wood frog traits (survival, morphology, growth, development, and behavior) and other components of the ecological community (zooplankton abundance, phytoplankton, periphyton, and bacterial community structure). Genetic diversity did not affect wood frog survival, growth, and development in either experiment. However, genetic diversity did affect the mean morphology of wood frog tadpoles in the first experiment and the abundance and distribution of zooplankton in the second experiment. It did not affect phytoplankton abundance, periphyton abundance, or bacterial community structure. While effect sizes (Cohen's d) of genetic diversity were approximately half those of environment treatments, the greatest effect sizes were for interaction effects between genetic diversity and environment. Our results indicate that genetic diversity can have a large effect on ecological processes, but the direction of those effects is highly dependent upon environmental conditions, and not easily predicted from simple measures of traits.

We studied the life history of the caddisfly Neophylax rickeri Milne (Trichoptera: Uenoidae) in two coastal northern California (Marin Co.) streams, Redwood Creek (a third order stream) and Webb Creek (a first order stream). Features were described for egg (oviposition location, predation, and egg mass phenology), larval (instar determination, instar phenology, voltinism, and case building), pupal (prepupal diapause, pupation phenology, mortality, and predation), and adult (emergence phenology, sex ratios, protandry, size, and longevity) stages. Oviposition on the downstream-facing sides of rocks occurred primarily in November in both streams; however, egg masses were also present from April through November in Webb Creek. Egg masses in Redwood Creek, but not Webb Creek, sometimes contained an egg predator, Acanthocnema sp. (Diptera: Scathophagidae). N. rickeri has five instars. The Redwood Creek population exhibits a synchronous univoltine pattern of development, but in Webb Creek a more variable pattern is present, with the majority of the population being univoltine with a small bivoltine summer generation. The Redwood Creek population had a longer prepupal diapause, but a shorter emergence period than Webb Creek. Prepupal mortality was highest in the summer in Redwood Creek, but it was overall low in Webb Creek. Pupal and adult sex ratios indicated protandry, with males emerging before females by ≈2 wk in both streams. The differences in the life histories of N. rickeri between streams for voltinism, life cycle timing, mortality, and pupal diapause suggest that physical habitat differences likely influence the variation in life history of this species.

Embryos of most anurans develop within a special microhabitat consisting of the jelly secreted around the eggs by the female parent. The jelly envelope covering each embryo effectively provides a buffer between it and the surrounding water; moreover, the three-dimensional structure of the gelatinous egg mass is thought to represent an adaptation to thermal stresses and gas exchange requirements likely to be encountered by the developing embryos. Jelly masses of North American ranid frogs can be categorized into two species-specific types related to these environmental factors: globular-shaped and surface film forms (Moore, 1940). Frogs that breed early in the spring deposit their eggs in globular masses, which may facilitate the concentration and retention of heat (obtained from solar radiation) within the egg masses. Such traits may be particularly adaptive in the cold water characteristic of ponds in early spring. In contrast, frogs breeding in late spring and summer generally deposit their eggs in monolayer surface film masses. Because embryos of these species develop in much warmer conditions, their metabolic rates are higher than those of early-breeding species and the oxygen tension in the water is lower; thus this egg mass form may represent an adaptation to maximize surface area for gas exchange (reviewed in Salthe and Mecham, 1974). In some instances, this surface film structure may also functionally cool the eggs (Ryan, 1978). Wood frogs (Rana sylvatica) are the most northerly distributed ectothermic tetrapod in North America, with a range extending from north of the Arctic Circle to Georgia. In temperate regions, wood frogs breed in early spring, frequently before ice has completely melted from breeding ponds. Wood frogs deposit their egg masses in communal clumps within limited areas, thereby compounding insulation effects provided by their individual egg masses. Although several investigators have suggested that clumping of egg masses warms embryos (R. sylvatica: Herreid and Kinney, 1967; Hassinger, 1970; Howard, 1980; Seale, 1982; R. temporaria: Savage, 1950, 1961; Guyetant, 1966; Beattie, 1980; R. pipiens: Zweifel, 1968; Hassinger, 1970; Merrell, 1977; R. aurora, R. pretiosa: Licht, 1971), differences in temperatures of egg masses at various positions within clumps have not been investigated. We studied egg masses in two woodland ponds near Ithaca, New York during April 1978 and 1980. The ponds are shallow (less than 0.8 m depth) permanent pools, with little submerged or emergent vegetation. Most egg masses were deposited in clumps in the shallowest water (less than 0.25 m depth), attached to submerged vines, reeds, and twigs. Generally, all egg masses in a pond were deposited at one site, leading to the formation of one clump per pond. However, in 1978 at one pond, six clumps consisting of variable numbers of egg masses were formed, allowing us to compare characteristics of egg masses based upon clump size within the same pond. Clumps consisted of between 3 and 150 egg masses. To investigate the temperature of egg masses in relation to their positions within a clump, we measured temperatures of some egg masses (approx. 3 cm within each egg mass) both at the center and at the periphery of clumps, as well as those of single egg masses deposited outside clumps. We then compared these temperatures with that of the surrounding water, 10 cm outside the clump (5 cm below the water surface). All temperature data were obtained with a Schultheis quick-reading thermometer. To quantify the of each egg mass to the surrounding water, we visually estimated the percentage surface area of each egg mass not in direct contact with surrounding egg masses (on its lateral and ventral surfaces). Observations were made on clumps consisting of various numbers of egg masses, and in clumps on both sunny and overcast days. Although most egg masses within clumps were warmer than the surrounding water, egg masses in central locations within a clump were warmer than those at the edge of that clump. Temperature elevation of egg masses was inversely correlated with their exposed surface area. Moreover, egg mass temperatures increased (in all clump positions) as the size of the clump increased (Fig. 1). This finding suggests that the presence of additional egg masses in a clump provides added insulation, improving the heat retention capacity of the clumped egg masses. The correlation of temperature elevation of an egg mass with its surface area exposure was apparent both on sunny and on cloudy days. Figure 2 shows temperature elevation of egg masses as a function of their position within a clump of 36 egg masses on two successive afternoons. The first day was sunny and central egg masses were 3.0-3.2?C above ambient. Weather conditions the