Effects of nutritional restriction on nitrogen and carbon stable isotopes in growing seabirds

When using stable isotopes as dietary tracers it is essential to consider effects of nutritional state on isotopic fractionation. While starvation is known to induce enrichment of ¹⁵N in body tissues, effects of moderate food restriction on isotope signatures have rarely been tested. We conducted two experiments to investigate effects of a 50–55% reduction in food intake on δ¹⁵N and δ¹³C values in blood cells and whole blood of tufted puffin chicks, a species that exhibits a variety of adaptive responses to nutritional deficits. We found that blood from puffin chicks fed ad libitum became enriched in ¹⁵N and ¹³C compared to food-restricted chicks. Our results show that ¹⁵N enrichment is not always associated with food deprivation and argue effects of growth on diet–tissue fractionation of nitrogen stable isotopes (Δ¹⁵N) need to be considered in stable isotope studies. The decrease in δ¹³C of whole blood and blood cells in restricted birds is likely due to incorporation of carbon from ¹³C-depleted lipids into proteins. Effects of nutritional restriction on δ¹⁵N and δ¹³C values were relatively small in both experiments (δ¹⁵N: 0.77 and 0.41‰, δ¹³C: 0.20 and 0.25‰) compared to effects of ecological processes, indicating physiological effects do not preclude the use of carbon and nitrogen stable isotopes in studies of seabird ecology. Nevertheless, our results demonstrate that physiological processes affect nitrogen and carbon stable isotopes in growing birds and we caution isotope ecologists to consider these effects to avoid drawing spurious conclusions.     

Equations for Lipid Normalization of Carbon Stable Isotope Ratios in Aquatic Bird Eggs

Stable isotope ratios are biogeochemical tracers that can be used to determine the source of nutrients and contaminants in avian eggs. However, the interpretation of stable carbon ratios in lipid-rich eggs is complicated because ¹³C is depleted in lipids. Variation in ¹³C abundance can therefore be obscured by variation in percent lipids. Past attempts to establish an algebraic equation to correct carbon isotope ratios for lipid content in eggs have been unsuccessful, possibly because they relied partly on data from coastal or migratory species that may obtain egg lipids from different habitats than egg protein. We measured carbon, nitrogen and sulphur stable isotope ratios in 175 eggs from eight species of aquatic birds. Carbon, nitrogen and sulphur isotopes were enriched in lipid-extracted egg samples compared with non extracted egg samples. A logarithmic equation using the C∶N ratio and carbon isotope ratio from the non extracted egg tissue calculated 90% of the lipid-extracted carbon isotope ratios within ±0.5‰. Calculating separate equations for eggs laid by species in different habitats (pelagic, offshore and terrestrial-influenced) improved the fit. A logarithmic equation, rather than a linear equation as often used for muscle, was necessary to accurately correct for lipid content because the relatively high lipid content of eggs compared with muscle meant that a linear relationship did not accurately approximate the relationship between percent lipids and the C∶N ratio. Because lipid extraction alters sulphur and nitrogen isotope ratios (and cannot be corrected algebraically), we suggest that isotopic measurement on bulk tissue followed by algebraic lipid normalization of carbon stable isotope ratio is often a good solution for homogenated eggs, at least when it is not possible to complete separate chemical analyses for each isotope.     

Does detritus quality predict the effect of native and non‐native plants on the performance of larval amphibians?

1. The lack of consistent differences between the traits of native and non‐native plant species makes it difficult to make general predictions about the ecological impact of invasive plants; however, the increasing number of non‐native plants in many habitats makes the assessment of the impact of each individual species impracticable. General knowledge about how specific plant traits are linked to their effects on communities or ecosystems may be more useful for predicting the effect of plant invasions. Specifically, we hypothesised that higher carbon‐to‐nitrogen ratio (C:N) and percent lignin in plant detritus would reduce the rate of development and total mass at metamorphosis of tadpoles, resulting in lower metamorph production (total fresh biomass) and amphibian species richness. 2. To test these hypotheses, we raised five species of tadpoles in mesocosms containing senescent leaves of three common native and three common non‐native wetland plants that varied in C:N ratio and % lignin. 3. Leaf mass loss, total metamorph production and the number of species that metamorphosed declined as a function of increasing C:N ratio of plant leaves. Plant lignin content was not related to the production of metamorphs or the number of species that metamorphosed. The percentage of wood frog (Lithobates sylvaticus) and American toad (Anaxyrus americanus) tadpoles reaching metamorphosis declined as a function of increasing plant C:N ratio. Mean time to metamorphosis increased and mean mass at metamorphosis declined as a function of increasing plant C:N ratio. Tadpole performance and metamorph diversity and production (biomass) were similar between native and non‐native plant species with similar C:N ratio in leaves. Percent lignin was not a significant predictor of tadpole performance. 4. Our results show that the impact of a plant invasion on tadpole performance could depend on differences between the quality of the detritus produced by the invading species and that of the native species it replaces. We suggest that plant community changes that lead to dominance by more recalcitrant plant species (those with higher leaf C:N ratio) may negatively affect amphibian populations.     

Populational Changes of the Enteric Protozoans Opalina spp. and Nyctotherus cordiformis During the Ontogeny of Anuran Tadpoles

ABSTRACT. We studied ontogenetic population changes of Opalina and Nyctotherus cordiformis in eight species of tadpoles from 10 sites in east-central Mississippi. Most tadpoles acquired Opalina early in development, while the acquisition of N. cordiformis was variable. Developmental stage, species and collection site explained significant amounts of the variation in Opalina density of tadpoles ( F = 11.6; df = 27, 235; P < 0.0001) and metamorphs ( F = 7.31; df = 24, 84; P < 0.0001). Relationships between Opalina density and host stage showed either (1) a gradual decrease or (2) a gradual increase throughout host ontogeny. Opalina densities declined during metamorphosis. Density variations of N. cordiformis were explained by host species of tadpoles ( F = 9.30; df = 7, 142; P < 0.0001) and by host species and stage of metamorphs ( F = 5.85; df = 8, 62; P < 0.0001).
The length of larval period, habitat duration and generation time of the protozoans are suggested as major modifiers of the protozoan densities. Hosts with long larval periods show a decreasing population density and hosts with short developmental periods show a pattern of increasing density. Neither pattern was detected in tadpoles from temporary sites. Metamorphic declines in protozoan density, but not necessarily the loss of protozoans, reflect metamorphic alterations of the gut common to all hosts.     

Influence of density and predators on metamorphic size in Rhinella schneideri tadpoles raised in mesocosm conditions

Amphibians exhibit extreme plasticity in the timing of metamorphosis, and several species respond to water availability, accelerating metamorphosis when their ponds dry. We analyzed the plasticity of the developmental response to water volume in Rhinella schneideri tadpoles. We raised tadpoles in mesocosm. Covariation between body size at metamorphosis and timing of development was positive. Nevertheless, the first approximately 53% of the metamorphoses finishing the cycle required between 34 and 56 days, and the covariation between body size at metamorphosis and timing of development was negative. For these tadpoles, the larval density and the presence of predators did not significantly affect their mass to metamorphosis. Nevertheless, predators affected time to metamorphosis. For the remainder of the tadpoles that reached metamorphosis at > 56 days, the relationship between body size at metamorphosis and timing of development was positive. For these tadpoles, larval density was important for mass at metamorphosis and presence of predators was also important for time to metamorphosis. Two dominant features were observed: (i) approximately 53% of metamorphs had morphological features similar to individuals developing in desiccating ponds, and (ii) the other individuals had morphological characteristics comparable to metamorphs developing in an unchanging environment.     

Facultative paedomorphosis as a mechanism promoting intraspecific niche differentiation

Organisms with complex life cycles are characterized by a metamorphosis that allows for a major habitat shift and the exploitation of alternative resources. However, metamorphosis can be bypassed in some species through a process called paedomorphosis, resulting in the retention of larval traits at the adult stage and is considered important at both micro‐ and macroevolutionary scales. In facultatively paedomorphic populations of newts, some individuals retain gills and a fully aquatic life at the adult stage (paedomorphs), while others undergo complete metamorphosis (metamorphs), allowing for a terrestrial life‐stage. Because facultative paedomorphosis affects trophic structures and feeding mechanism of newts, one hypothesis is that it may be maintained as a trophic polymorphism, with the advantage to lessen intraspecific competition during the shared aquatic life‐stage. Here, we tested this hypothesis combining stomach content data with stable isotope techniques, using carbon and nitrogen stable isotopes, in facultatively paedomorphic alpine newts Ichthyosaura alpestris. Both stomach content and stable isotope analyses showed that paedomorphs had smaller trophic niches and were more reliant on pelagic resources, while metamorphs relied more on littoral resources, corresponding to a polyphenism along the littoral–pelagic axis and the extension of the population's trophic niche to otherwise ‘underused’ pelagic resources by paedomorphs. Interestingly, stable isotopes revealed that the trophic polyphenism was less marked in males than in females and potentially linked to sexual activity. Although paedomorphosis and metamorphosis are primarily seen as results of tradeoffs between the advantages of using aquatic versus terrestrial habitats, this study provides evidence that additional forces, such as intraspecific trophic niche differences between morphs and trophic niche expansion, may play an important role in the persistence of this dimorphism in heterogeneous environments. Moreover, the different patterns found in males and females show the importance of considering sex to understand the evolutionary ecology of trophic polymorphisms.     

Growth and development of larval green frogs (Rana clamitans) exposed to multiple doses of an insecticide

Our objective was to determine how green frogs (Rana clamitans) are affected by multiple exposures to a sublethal level of the carbamate insecticide, carbaryl, in outdoor ponds. Tadpoles were added to 1,000-l ponds at a low or high density which were exposed to carbaryl 0, 1, 2, or 3 times. Length of the larval period, mass, developmental stage, tadpole survival, and proportion metamorphosed were used to determine treatment effects. The frequency of dosing affected the proportion of green frogs that reached metamorphosis and the developmental stage of tadpoles. Generally, exposure to carbaryl increased rates of metamorphosis and development. The effect of the frequency of carbaryl exposure on development varied with the density treatment; the majority of metamorphs and the most developed tadpoles came from high-density ponds exposed to carbaryl 3 times. This interaction suggests that exposure to carbaryl later in the larval period stimulated metamorphosis, directly or indirectly, under high-density conditions. Our study indicates that exposure to a contaminant can lead to early initiation of metamorphosis and that natural biotic factors can mediate the effects of a contaminant in the environment.     

Parasite-induced changes in nitrogen isotope signatures of host tissues

To estimate isotopic changes caused by trematode parasites within a host, we investigated changes in the carbon and nitrogen isotope ratios of the freshwater snail Lymnaea stagnalis infected by trematode larvae. We measured carbon and nitrogen stable isotopes within the foot, gonad, and hepatopancreas of both infected and uninfected snails. There was no significant difference in the delta13C and delta15N values of foot and gonad between infected and uninfected snails; thus, trematode parasite infections may not cause changes in snail diets. However, in the hepatopancreas, delta15N values were significantly higher in infected than in uninfected snails. The 15N enrichment in the hepatopancreas of infected snails is caused by the higher 15N ratio in parasite tissues. Using an isotope-mixing model, we roughly estimated that the parasites in the hepatopancreas represented from 0.8 to 3.4% of the total snail biomass, including the shell.     

Effects of elemental composition on the incorporation of dietary nitrogen and carbon isotopic signatures in an omnivorous songbird

The use of stable isotopes to infer diet requires quantifying the relationship between diet and tissues and, in particular, knowing of how quickly isotopes turnover in different tissues and how isotopic concentrations of different food components change (discriminate) when incorporated into consumer tissues. We used feeding trials with wild-caught yellow-rumped warblers (Dendroica coronata) to determine delta15N and delta13C turnover rates for blood, delta15N and delta13C diet-tissue discrimination factors, and diet-tissue relationships for blood and feathers. After 3 weeks on a common diet, 36 warblers were assigned to one of four diets differing in the relative proportion of fruit and insects. Plasma half-life estimates ranged from 0.4 to 0.7 days for delta13C and from 0.5 to 1.7 days for delta15N . Half-life did not differ among diets. Whole blood half-life for delta13C ranged from 3.9 to 6.1 days. Yellow-rumped warbler tissues were enriched relative to diet by 1.7-3.6% for nitrogen isotopes and by -1.2 to 4.3% for carbon isotopes, depending on tissue and diet. Consistent with previous studies, feathers were the most enriched and whole blood and plasma were the least enriched or, in the case of carbon, slightly depleted relative to diet. In general, tissues were more enriched relative to diet for birds on diets with high percentages of insects. For all tissues, carbon and nitrogen isotope discrimination factors increased with carbon and nitrogen concentrations of diets. The isotopic signature of plasma increased linearly with the sum of the isotopic signature of the diet and the discrimination factor. Because the isotopic signature of tissues depends on both elemental concentration and isotopic signature of the diet, attempts to reconstruct diet from stable isotope signatures require use of mixing models that incorporate elemental concentration.     

Isotope composition of carbon and nitrogen in tissues and organs of <Emphasis Type="Italic">Betula pendula</Emphasis>

Ratios of ¹³С/¹²C and ¹⁵N/¹⁴N isotopes were identified in different parts and organs of drooping birch (Betula pendula Roth) in preforest-steppe and pine-birch forests of the Middle Urals by mass spectrometry. The data were analyzed and interpreted from the perspective of biochemical processes of carbon and nitrogen metabolism in the leaf, cambial tissue, trunk wood, branches, roots, and in the soil. The lighter isotopic composition of carbon is characteristic for the leaves, trunk cambium as well as fine (<2 mm) roots. The trunk wood is characterized by the basal trend for ¹³C enrichment. The heavier carbon isotopic composition inversely related to metabolic activity of organs and tissues, in addition, ¹³С/¹²C ratio corresponds to the nitrogen content in the organs and tissues, indicating the metabolic control of carbon fractionation in woody plants. The isotopic composition of nitrogen in the aboveground parts of the plant (leaves, trunk cambium, wood) and in the medium and fine roots was significantly depleted in ¹⁵N (δ¹⁵N varies from 0 to–3‰), while main roots (δ¹⁵N = 0.6 ‰) and soil (δ¹⁵N = 2.4–6.7‰) were more enriched. The ratio of stable isotopes of carbon and nitrogen is an integrating index of carbon and nitrogen metabolism in plants.     

PERFORMANCE OF TADPOLES FROM THE HYBRIDOGENETIC RANA ESCULENTA COMPLEX: INTERACTIONS WITH POND DRYING AND INTERSPECIFIC COMPETITION

The performance of three genotypes (LL, LR, RR) of tadpoles resulting from the hybrid mating system of Rana lessonae (phenotype L, genotype LL) and Rana esculenta (phenotype E, genotype LR) was determined in artificial ponds. The effects of interspecific competition and pond drying on growth, development, and survival of tadpoles were used to measure the performance of genotypes and the relative fitness of offspring. Among the three genotypes, tadpoles from the homogametic mating RR had the lowest survival, growth, and development under all environmental conditions. Body size of the LL and LR genotype tadpoles at metamorphosis was reduced by competition and pond drying. Days to metamorphosis were also higher for the LL and LR genotype tadpoles in competition ponds. The proportion of individuals metamorphosing of each genotype was differentially lowered by competition and pond drying. The LL genotype produced more metamorphs than the LR genotype in the constant water level ponds, but the LR genotype produced more in drying ponds. In competition ponds, the LR genotype produced more metamorphs than the LL genotype, but the LL genotype produced more metamorphs in ponds without competition. The RR genotype produced no metamorphs in any of the experimental environments. Increased performance of LR offspring from the heterogametic mating, in harsh conditions, and reduced performance of RR offspring from the homogametic mating, even under favorable conditions, relative to the parental genotype (LL) suggests that the population dynamics of this hybridogenetic system is strongly dependent on mate choice in mixed populations and the subsequent pond environment females select for oviposition and larval development.     

Brain plasticity over the metamorphic boundary: carry-over effect of larval environment on froglet brain development

Brain development shows high plasticity in response to environmental heterogeneity. However, it is unknown how environmental variation during development may affect brain architecture across life history switch points in species with complex life cycles. Previously, we showed that predation and competition affect brain development in common frog (Rana temporaria) tadpoles. Here, we studied whether larval environment had carry-over effects in brains of metamorphs. Tadpoles grown at high density had large optic tecta at metamorphosis, whereas tadpoles grown under predation risk had small diencephala. We found that larval density had a carry-over effect on froglet optic tectum size, whereas the effect of larval predation risk had vanished by metamorphosis. We discuss the possibility that the observed changes may be adaptive, reflecting the needs of an organism in given environmental and developmental contexts.     

Quantitative genetic analysis of larval life history traits in two alpine populations of Rana temporaria

We estimated genetic and maternal variance components of larval life history characters in alpine populations of Rana temporaria (the common frog) using a full-sib/half-sib breeding design. We studied trait plasticity by raising tadpoles at 14 or 20°C in the laboratory. Larval period and metamorphic mass were greater at 14°C. Larval period did not differ between populations, but high elevation metamorphs were larger than low elevation metamorphs. Significant additive variation for larval period was detected in the low altitude population. No significant additive variation was detected for mass at metamorphosis (MM), which instead displayed significant maternal effects. Plasticity in metamorphic mass of froglets was greater in the high altitude population. The plastic response of larval period to temperature did not differ between the populations. Evolution of metamorphic mass is likely constrained by lack of additive genetic variation. In contrast, significant heritability for larval period suggests this trait may evolve in response to environmental change. These results differ from other studies on R. temporaria, suggesting that populations of this broadly distributed species present substantial geographic variation in the genetic architecture and plasticity of tadpole life history traits.     

δ<Superscript>13</Superscript>C and δ<Superscript>15</Superscript>N in tissue of coral polyps and epilithic algae inhabiting damaged coral colonies under the influence of different light intensities

Studies were performed of the carbon and nitrogen stable isotope (δ¹³C and δ¹⁵N) composition (δ¹³C and δ¹⁵N) of the corals Porites cylindrica and P. lutea (5 years after damaging the colonies by the bleaching events) and of epilithic algae settled onto damaged areas of coral colonies. Coral polyps and three epilithic algal communities (‘red algal turf, green algal turf and red calcified crusts’) were sampled along the boundary between communities of coral polyps and algal colonizers from differently illuminated habitats from 2 to 90% of incident surface photosynthetically active radiation (PAR₀). It was found that communities with a predominance of red algae significantly differed from communities with a predominance of green algae in δ¹³C but not in δ¹⁵N values. An influence of habitat irradiance was found only for communities of coral polyps for δ¹³C and δ¹⁵N values: under bright light (70–90% PAR₀) polyp tissues of both coral species were significantly enriched in heavy carbon isotopes and insignificantly in nitrogen isotopes (δ¹³C values difference ~4‰) relative to tissues of corals under lower light 15–50% PAR₀. On the basis of these results we assumed that differences in light intensities in the habitat ranging from 15 to 90% PAR₀ do not influence on accessibility of the main carbon and nitrogen sources for corals and algae, and exchange by these elements between organisms. We also assumed that the relative enrichment in the heavy carbon isotopes of coral tissues in high light is a result of decreased isotope fractionation (or the absence of fractionation in photosynthesis of their zooxanthellae).     

Stable isotope enrichment in laboratory ant colonies: effects of colony age,metamorphosis, diet,and fat storage

Ecologists use stable isotopes to infer diets and trophic levels of animals in food webs, yet some assumptions underlying these inferences have not been thoroughly tested. We used laboratory‐reared colonies of Solenopsis invicta Buren (Hymenoptera: Formicidae: Solenopsidini) to test the effects of metamorphosis, diet, and lipid storage on carbon and nitrogen stable isotope ratios. Effects of metamorphosis were examined in ant colonies maintained on a control diet of domestic crickets and sucrose solution. Effects of a diet shift were evaluated by adding a tuna supplement to select colonies. Effects of lipid content on stable isotopes were tested by treating worker ants with polar and non‐polar solvents. δ¹³C and δ¹⁵N values of larvae, pupae, and workers were measured by mass spectrometry on whole‐animal preparations. We found a significant effect of colony age on δ¹³C, but not δ¹⁵N; larvae, pupae, and workers collected at 75 days were slightly depleted in ¹³C relative to collections at 15 days (Δδ¹³C = ?0.27‰). Metamorphosis had a significant effect on δ¹⁵N, but not δ¹³C; tissues of each successive developmental stage were increasingly enriched in ¹⁵N (pupae, +0.5‰; workers, +1.4‰). Availability of tuna resulted in further shifts of about +0.6‰ in isotope ratios for all developmental stages. Removing fat with organic solvents had no effect on δ¹³C, but treatment with a non‐polar solvent resulted in enriched δ¹⁵N values of +0.37‰. Identifying regular patterns of isotopic enrichment as described here should improve the utility of stable isotopes in diet studies of insects. Our study suggests that researchers using ¹⁵N enrichment to assess trophic levels of an organism at different sites need to take care not to standardize with immature insect herbivores or predators at one site and mature ones at another. Similar problems may also exist when standardizing with holometabolous insects at one site and spiders or hemimetabolous insects at another site.     

Effects of food quality on tissue-specific isotope ratios in the mussel <Emphasis Type="Italic">Perna perna</Emphasis>

Investigations into trophic ecology and aquatic food web resolution are increasingly accomplished through stable isotope analysis. The incorporation of dietary and metabolic changes over time results in variations in isotope signatures and turnover rates of producers and consumers at tissue, individual, population and species levels. Consequently, the elucidation of trophic relationships in aquatic systems depends on establishing standard isotope values and tissue turnover rates for the level in question. This study investigated the effect of diet and food quality on isotopic signatures of four mussel tissues: adductor muscle, gonad, gill and mantle tissue from the brown mussel Perna perna. In the laboratory, mussels were fed one of the two isotopically distinct diets for 3 months. Although not all results were significant, overall δ¹³C ratios in adductor, mantle and gill tissues gradually approached food source signatures in both diets. PERMANOVA analyses revealed significant changes over time in tissue δ¹³C (mantle and gill) with both diets and in δ¹⁵N (all tissues) and C:N ratios (mantle and gill) for one diet only. The percentage of replaced carbon isotopes were calculated for the 3 month period and differed among tissues and between diets. The tissue with the highest and lowest amount of replaced isotopes over 81 days were mantle tissue on the kelp diet (33.89%) and adductor tissue on the fish food diet (4.14%), respectively. Percentages could not be calculated for any tissue in either diet for δ¹⁵N due to the lack of significant change in tissue nitrogen. Fractionation patterns in tissues for both diets can be linked to nutritional stress, suggesting that consumer isotopic signatures are strongly dependent on food quality, which can significantly affect the degree of isotopic enrichment within a trophic level.     

COMPETITION AMONG TADPOLES OF COEXISTING HEMICLONES OF HYBRIDOGENETIC RANA ESCULENTA: SUPPORT FOR THE FROZEN NICHE VARIATION MODEL

Vertebrate animals reproducing without genetic recombination typically are hybrids, which have large ranges, are locally abundant, and live in disturbed or harsh habitats. This holds for the hemiclonal hybridogenetic frog Rana esculenta: it is widespread in Europe and commonly is found in disturbed habitats such as gravel pits. We hypothesize that its widespread occurrence may either be the result of natural selection for a single hemiclone acting as a broadly adapted “general-purpose” genotype, or of interclonal selection, which maintains multiple hemiclones that each are relatively narrowly adapted and perform differently across environments, that is, the Frozen Niche Variation model. We tested these competing hypotheses using 1000-L outdoor artificial ponds to rear tadpoles of the parental species (Rana lessonae [LL] and Rana ridibunda [RR]) alone, and each of three hemiclones of Rana esculenta (GUT1, GUT2, GUT3) alone, and in mixed hemiclonal populations from hatching to metamorphosis. Tadpoles of three coexisting hemiclones from a single natural population (near Gütighausen, Switzerland) were reared in both two- and three-way mixtures in equal total numbers at high and low density. For each species and hemiclone, the proportion of tadpoles metamorphosing decreased as the density of tadpoles increased, with the three hemiclones spanning the range of values exhibited by the two parental species. LL and GUT1 tadpoles produced the highest proportion of metamorphs, whereas tadpoles of RR produced the fewest metamorphs at both densities. GUT1 tadpoles also produced the largest metamorphs at low density, GUT2 and GUT3 tadpoles produced smaller metamorphs than did GUT1 tadpoles at the low density, but the three hemiclones did not differ from each other at high density. The parental species (LL and RR) were intermediate in metamorphic size to the hemiclones at low density, but all genotypes converged on a similar size at high density. Length of the larval period also was affected by density, but its effect was dependent on genotype. GUT1 tadpoles had the shortest larval period at the low density, but larval period was longer and not different between GUT1, GUT3, and LL at high density. RR tadpoles had the longest larval period at both densities. The most dramatic results were that three genotypes (GUT1, GUT2, and RR) maintained rank order and increased days to metamorphosis from low to high density, whereas two genotypes (GUT3 and LL) changed rank order and decreased days to metamorphosis from low to high density. Mixtures of hemiclones in two- and three-way combinations facilitated the proportion of tadpoles metamorphosing for GUT1 and GUT2 at both densities, but only at the low density for GUT3 tadpoles. Results from this experiment are incompatible with the General-Purpose Genotype model as a global explanation of hybrid abundance in these frogs. Alternatively, the Frozen Niche Variation prediction of general performance superiority of clonal mixtures relative to single clone populations is strongly supported. The data confirm that fitness advantages of hemiclones change, depending on the environment, such that in temporally and spatially heterogeneous habitats like ponds, frequency-dependent selection among hemiclones may promote coexistence in hemiclonal assemblages. Yet, differential dispersal or colonization ability and historical factors affecting hemiclone distribution may also be important in shaping patterns of clonal coexistence.     

Osteopenia and stable isotope ratios in bone collagen of Nubian female mummies

Stable carbon and nitrogen isotopes were analysed on bone collagen of 43 Sudanese Nubians from the X-Group period to test dietary hypotheses for the high frequency of osteopenia in this population. Stable carbon isotope ratios indicate that both normal and osteopenic individuals consumed the same mixed diet of C3 and C4 sources, which are assumed to have been constituted by the grain staples wheat/barley and sorghum/millet respectively. Females with osteopenia, however, have significantly elevated δ¹⁵N values. The enrichment effect is greatest in the third and fifth decades of life, and is consistently patterned with microstructural and frequency differences previously reported by other researchers. It is suggested that δ¹⁵N is reflecting differences in urea excretion and the renal processing and clearance of calcium and phosphorus. The study not only alerts us to the susceptibility of stable nitrogen isotopes to non-dietary (i.e. physiological) factors, but also identifies nitrogen isotope ratios as a possible new marker for osteopenia. Am J Phys Anthropol 103:185–199, 1997. © 1997 Wiley-Liss, Inc.     

Sequential predator effects across three life stages of the African tree frog, <Emphasis Type="Italic">Hyperolius spinigularis</Emphasis>

While theoretical studies of the timing of key switch points in complex life cycles such as hatching and metamorphosis have stressed the importance of considering multiple stages, most empirical work has focused on a single life stage. However, the relationship between the fitness components of different life stages may be complex. Ontogenetic switch points such as hatching and metamorphosis do not represent new beginnings—carryover effects across stages can arise when environmental effects on the density and/or traits of early ontogenetic stages subsequently alter mortality or growth in later stages. In this study, I examine the effects of egg- and larval-stage predators on larval performance, size at metamorphosis, and post-metamorphic predation in the African tree frog Hyperolius spinigularis. I monitored the density and survival of arboreal H. spinigularis clutches in the field to estimate how much egg-stage predation reduced the input of tadpoles into the pond. I then conducted experiments to determine: (1) how reductions in initial larval density due to egg predators affect larval survival and mass and age at metamorphosis in the presence and absence of aquatic larval predators, dragonfly larvae, and (2) how differences in mass or age at metamorphosis arising from predation in the embryonic and larval environments affect encounters with post-metamorphic predators, fishing spiders. Reduction in larval densities due to egg predation tended to increase per capita larval survival, decrease larval duration and increase mass at metamorphosis. Larval predators decreased larval survival and had density-dependent effects on larval duration and mass at metamorphosis. The combined effects of embryonic and larval-stage predators increased mass at metamorphosis of survivors by 91%. Larger mass at metamorphosis may have immediate fitness benefits, as larger metamorphs had higher survival in encounters with fishing spiders. Thus, the effects of predators early in ontogeny can alter predation risk even two life stages later.     

Interacting effects of predation risk and food availability on larval anuran behaviour and development

Age and size at metamorphosis are two important fitness components in species with complex life cycles. In anurans, metamorphic traits show remarkable phenotypic plasticity, especially in response to changes in growth conditions. It is also possible that the perception of risk directly determines changes in larval period and the size of metamorphs. This study examines how the perception of predation risk affects the timing of and size at metamorphosis in common frogs (Rana temporaria). I raised tadpoles at two risk levels (fish-conditioned water or unconditioned water) crossed with the availability or lack of food at night (all tadpoles had food available in the day). Tadpoles reacted to chemical cues from predatory fish by decreasing activity. A novel behavioural result was a predation×food interaction effect on refuge use, which also accounted for most of the predator main effect: predation risk only caused increased refuge use in the night-starved treatment. Despite these behavioural modifications, the perception of predation risk did not affect growth rate and mass at metamorphosis in a simple way: the effects of food regime on growth and size at metamorphosis were dependent on the level of predation risk as revealed by significant predation×food interaction effects. Tadpoles who had food withheld at night metamorphosed at the smallest size, suggesting a negative relationship between size at metamorphosis and refuge use. Tadpoles raised in fish-conditioned water had longer larval periods than those in unconditioned water, but these differences were significant only if food was available at night. These results conflict with the hypotheses that tadpoles should reduce their larval period or growth rates (and hence metamorphose at a smaller size) as the risk of predation increases. In contrast to predation risk, food availability strongly affected the length of the larval period: night-starved tadpoles metamorphosed relatively early with or without fish stimulus. Thus, early metamorphosis resulted from periods of low food availability, but not from a heightened ”perceived risk” of predation. This example counters the hypothesis of acceleration of the developmental rate (which shortens the time to metamorphosis) as a mechanism to escape a risky environment. Received: 18 August 1999 / Accepted: 10 January 2000