Canadian Journal of Zoology
The role of feeding strategy in the tolerance of a terrestrial salamander (Plethodon cinereus) to biogeochemical changes in northern hardwood forests
Journal: Canadian Journal of Zoology
Manuscript ID cjz-2017-0302.R2
Manuscript Type: Article
Date Submitted by the 13-Aug-2018 Author:
Complete List of Authors: Bondi, Cheryl; SUNY College of Environmental Science and Forestry Beier, Colin; SUNY College of Environmental Science and Forestry DepartmentDraft of Forest and Natural Resources Management Fierke, Melissa; SUNY College of Environmental Science and Forestry Ducey, Peter; SUNY Cortland Department of Biology
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Eastern red-backed salamander, Food web, Plethodon cinereus, Keyword: salamander diet, trophic ecology, calcium depletion, ACID RAIN < Discipline
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The role of feeding strategy in the tolerance of a terrestrial salamander (Plethodon cinereus) to biogeochemical changes in northern hardwood forests
Bondi, C.A.1, C.M. Beier1, M.K. Fierke1, and P.K. Ducey2
1College of Environmental Science and Forestry, State University of New York 1 Forestry Drive Syracuse New York 13210 Email: [email protected]
2Biological Sciences Department State University of New York, Cortland P.O. Box 2000 Cortland New York 13045 Email: [email protected]
Corresponding author Cheryl A Bondi, 232 Eastern Ave, Apt. 202 Manchester NH 03104 Draft Phone: (617) 838-1224 Email: [email protected]
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C.A. Bondi, C.M. Beier, M.K. Fierke, and P.K. Ducey
We investigated whether the trophic ecology of an apex predator is influenced by ecosystem-
level nutrient depletion. The feeding behavior and nutrient assimilation of a terrestrial
salamander (Plethodon cinereus (Green,1818)) was surveyed along a gradient of forest
biogeochemistry. Recent studies have documented populations of these salamanders in forests
with low-pH soils that were long thought to be fatal. One mechanism that may enable P. cinereus
to tolerate acid-impaired habitats is its generalist life history. We sampled diet, invertebrate prey
abundance, and tissue composition of P. cinereus from sites that range in calcium availability
and soil pH in northern forests of North America. We found P. cinereus consistently exhibited a generalist feeding strategy, having diverseDraft diets closely represented resource availability. Prey abundances were unrelated to the biogeochemical gradient (excluding gastropods), indicating
relatively intact food webs. Although P. cinereus at the two most acid-impaired sites consumed more prey, overall trophic strategies were consistent across the gradient. Salamander tissue composition was unrelated to variation in forest biogeochemistry, although manganese levels were elevated in the most acid-impaired forests. We suggest that a generalist feeding strategy, combined with diverse and compositionally stable food webs, facilitates tolerance by this abundant predator of the challenges imposed by acid-impaired habitats.
Keywords calcium depletion, Eastern Red-Backed Salamander, Food web, Plethodon cinereus, salamander diet, trophic ecology,
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Introduction
The eastern deciduous forest biome of North America has been shaped over the last
century by many anthropogenic drivers of environmental change, but air pollution in its many
forms has arguably been its most chronic and widespread stressor (McNulty et al. 2007; Sullivan
et al. 2018). Atmospheric deposition of sulfur and nitrogen as “acid rain”, and its effects on soil
nutrients, plant species composition, and forest health, are well documented (Likens et al. 1970;
Federer et al. 1989; DeHayes et al. 1999; Driscoll et al. 2001; Lovett et al. 2009), while the direct
and indirect impacts of these changes on forest biodiversity and trophic interactions are still
under investigation (Driscoll et al. 2003; Pabian and Brittingham 2011; Beier et al. 2012; Bondi
et al. 2015). Deposition of strong acids where soil buffering capacity is insufficient (DeHayes et
al. 1999) results in both the acidificationDraft (lowered pH) of soil habitats and leaching of essential
nutrients such as calcium (Ca) and other base cations (e.g., Mg, Na) from soils (Driscoll et al.
2001). Chronic depletion of base cations also drives the mobilization of soluble aluminum (Al3+)
and manganese (Mn2+) in forest soils, which can be toxic to plants and animals in high
concentrations (Lawrence et al. 1995). Therefore, soil acidification is a potential direct stressor to
soil fauna, related to species tolerance of changes in habitat quality (e.g., decreased substrate pH,
increased toxic metals), and may be an indirect stressor via its impacts on biochemical cycling of
essential nutrients and nutritional deficits (particularly Ca) in food webs (Hames et al. 2002).
Due to base-poor parent materials in soils at many locations, recovery of Ca pools and cycles in
most acid-impaired forests will proceed very slowly, even under today’s rates of N and S
deposition, which remain at historically low levels since monitoring began (Lawrence et al.
2015; Sullivan et al. 2018). This suggests that the legacy of acid impairment, which may include
deficiencies of essential nutrients in forest food webs, could persist for centuries.
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Yet such stressors, legacies, and their bottom-up effects in terrestrial food webs may be mediated in some cases by species with traits that confer the ability to tolerate and/or compensate for changes in habitat and nutritional quality. Species that are both highly abundant and widely distributed have evolved generalist traits for persisting in spatially and temporally dynamic landscapes that may also be of benefit in tolerating novel anthropogenic stressors (Bonier et al.
2007). If these species also play unique or ‘keystone’ roles, their ability to persist may confer some degree of stability to community assembly, trophic interactions, and higher-order processes such as nutrient cycling (Richmond et al 2005). To explore this question, we investigated the feeding ecology of a widely distributed, highly abundant ‘keystone’ species of eastern deciduous forests– the Eastern Red-Backed Salamander (Plethodon cinereus Green, 1818), recently found to persist in severely acidified forests ofDraft the eastern US and Canada – across a wide biogeochemical gradient of pH and Ca availability in northern hardwood forests (Wyman and
Moore 2010; Bondi et al. 2016).
Terrestrial salamanders are among the most abundant predators in forest ecosystems and play significant ecological roles in food webs (Burton and Likens 1975; Davic and Welsh 2004;
Semlitsch et al. 2014). Plethodontid salamanders are particularly influential in forest food webs because of their high abundance, generalist feeding strategy, ability to efficiently convert biomass to energy, and trophic position as a prey source to higher order consumers (Burton and
Likens 1975; Burton 1976; Best and Welsh, Jr. 2014; Semlitsch et al. 2014). These salamanders act as top-down regulators of the detritivore community, influencing ecosystem processes such as leaf litter decomposition and assimilation of nutrients into the food web (Davic and Welsh
2004; Walton 2013; Hickerson et al. 2017). Furthermore, terrestrial salamanders are prey for secondary consumers including ground-feeding birds, small mammals, snakes, and larger
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amphibians (Cochran 1911; Brodie et al. 1979; Ducey et al. 1994; Fenster and Fenster 1996;
Lancaster and Wise 1996), making them facilitators of nutrient and energy flows between below
and above-ground components of the food web.
Chronic acid deposition could reduce Ca availability for terrestrial salamanders via
several mechanisms. In general, terrestrial vertebrates must acquire Ca2+ ions from their
environment and then store and mobilize reserves as needed (Srivastav et al. 1995, 2000). A
major source of Ca for amphibians is from diet, via absorption through the small intestine
(Robertson 1975). A second source is transport of Ca2+ ions from the environment (water or
substrate) through the epithelial layers. Adult salamanders (Ambystoma tigrinum (Green, 1825))
and frogs (Lithobates pipiens (Schreber, 1782)) actively transport Ca2+ across the skin in a
saturable process that is proportional to DraftCa2+ available in external solution (Stiffler 1996; Zerella
and Stiffler 1999). It is unknown whether ion uptake by terrestrial amphibians is limited by the
availability of nutrients in the substrate or if, when environmental sources of Ca are deficient
(due to low concentration, low pH, and/or uptake inhibition by Al cations), amphibians adjust
their feeding behavior to supplement their diet with additional Ca from prey.
Invertebrate prey groups vary widely in elemental composition of their body tissues and,
in turn, differ in their nutritional value for consumers (Riechle et al. 1969; Gist and Crossley Jr.
1975; Seastedt and Tate 1981; Studier and Sevick 1992). For example, the shells of gastropods
contain high amounts of Ca (200-396 ppt; Reichle et al. 1969; Gist and Crossley 1975; Blum et
al. 2000; Bondi 2015), and therefore, as a functional group, land snails represent the largest
reservoir of nutritional Ca in forest soil food webs (Bondi 2015). Oribatid mites are miniscule
but highly abundant in forest soils (3,000-22,000/m2; Gist and Crossley 1975; Bondi 2015) and
their cuticle is very Ca-rich, in both carbonate and oxalate forms (Norton and Behan-Pelletier
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1991). By contrast, millipedes are relatively large soil invertebrates that have high Ca concentrations in their cuticle (~123-327 ppt; Reichle et al. 1969; Gist and Crossley 1975; Bondi
2015), but are not edible to most predators due to noxious secretions used for defense (Blum and
Woodring 1962; Kuwahara et al. 2002), making them a much less available source of Ca to consumers.
The Eastern Red-Backed Salamander (Plethodon cinereus (Green,1818)) may be the most abundant vertebrate in the northern hardwood forests of eastern North America (Burton and
Likens 1976) and is considered a ‘keystone’ species for its influence on soil food webs and ecosystem processes. Early research showed that P. cinereus exposed to acidic soils in the laboratory (pH <3.8) exhibited lower growth rates and higher rates of sodium loss, and actively avoided low pH substrates in situ (WymanDraft and Hawksley-Lescault 1987; Sugalski and Claussen
1997). However, recent studies in the northern hardwood forests of the eastern US and Canada have observed P. cinereus populations in very acidic forest soils (pH < 3.5), including severely impaired soils (pH < 3.0) with base cation deficits, where toxic Al3+ has been mobilized in soil solution (Moore and Wyman 2010; Bondi et al. 2016). The mechanism(s) by which populations of P. cinereus can thrive in such habitats, which were long thought to be unsuitable, if not lethal, are unknown. One hypothesis is that a generalist feeding strategy (Burton 1976) enables P. cinereus to acquire the requisite nutrition for growth and reproduction, even in forests where, at an ecosystem level, such nutrients are very scarce or have been greatly depleted from their primary sources (soils and vegetation; Bondi et al. 2016). However, for this hypothesis to be supported, we must assume that the food web that P. cinereus relies upon is sufficiently intact, from a functional perspective – i.e., that the available prey are sufficient in both abundance and nutritional value – despite orders-of-magnitude differences in soil and plant nutrition, as well as
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soil acidity, between acid-impaired and non-acidified (well-buffered) forests. An alternative
hypothesis is that P. cinereus in acid-impaired forests is locally adapted to preferentially
consume Ca-rich invertebrate prey to offset any Ca deficiencies in the overall food web and/or
greatly reduced Ca ion availability in soil solution.
In this study we evaluated these alternative hypotheses by determining whether diet,
feeding behavior, and tissue assimilation (body concentrations of nutrients) of P. cinereus varied
among extant populations in forests representing a regional gradient in Ca availability and soil
pH, including severely acid-impaired ecosystems. Given their ecological role, it is important to
understand how environmental changes, such as calcium depletion, may affect P. cinereus
feeding behavior and nutrient assimilation. We assessed soil invertebrate communities at the
level of functional groups to determine whetherDraft the overall composition – and by proxy, their
nutritional quality in terms of Ca content – of the food web differed between acid-impaired and
more nutrient-rich forest ecosystems. Based on intensive sampling of the stomach contents of P.
cinereus, we evaluated whether feeding behavior was consistently generalist (reflecting the
relative availability of prey) or was more specialized (i.e., indicating a preference for Ca-rich
prey groups) in acid-impaired forests. We also assayed elemental concentrations of Ca and other
nutrients in P. cinereus tissue to characterize changes along the study gradient and to identify
any nutritional deficiencies in populations living on acid-impaired soils. Given their ecological
role, it is important to understand how environmental changes, such as calcium depletion, may
affect feeding behavior and nutrient assimilation of a top predator. Moreover, any acid-mediated
changes in the trophic ecology of P. cinereus could have cascading trophic affects in both
directions— shifts in invertebrate prey consumption can alter detritivore community structure
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and subsequently leaf litter decomposition rates, while changes in nutrient assimilation will influence higher trophic levels that feed on salamanders.
Materials and Methods
Study sites
To investigate whether P. cinereus diet and body composition vary among sites with contrasting levels of Ca availability and pH, we collected salamanders and their stomach contents from nine study sites in hardwood forests in New Hampshire and Vermont (Table 1). The study sites, selected from a more extensive gradient where P. cinereus were sampled (Bondi et al. 2016) span a range of soil Ca availability, Ca content of foliage, and soil pH, representative of the northern hardwood forests of the easternDraft US and Canada (Table 1). Soils of the study region where the sites are located are dominated by Spodosols and surficial deposits are glacial till.
Vegetation composition consisted of mature hardwood forests with sugar maple (Acer saccharum Marshall), American beech (Fagus grandifolia Ehrh), and yellow birch (Betula alleghaniensis Britton) dominant in the overstory, with red spruce (Picea rubens Sarg) and balsam fir (Abies balsamea Mill.) present at higher elevations. Soil Ca availability and pH (Oa/A soil horizons), and Ca concentrations of the foliage of A. saccharum, were previously measured at these sites (see methods in Horsley et al. 2008). Because A. saccharum is the dominant tree species and constitutes a majority of the leaf litter at these sites, we used foliar Ca concentrations as a proxy for Ca availability in the detritus of the forest floor. The terrestrial salamander community at these sites is dominated by P. cinereus, with their numbers ranging from 74% at site LOW2 to 100% at sites VLOW2, HIGH1 and HIGH 2, of the overall salamander abundance.
The other species found at these sites were Eurycea bislineata (Green, 1818), Ambystoma
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maculatum (Shaw, 1802), and the Notophthalmus viridescens (Rafinesque, 1820) eft stage in
relatively low numbers compared to P. cinereus. Because differences in the physiology and
ecology of the unstriped and striped morph have been described (Anthony et al. 2008), the nine
sites we selected for this study consisted mainly of the striped morph of P. cinereus, with only a
single salamander at the MOD2 site identified as an unstriped morph.
Prey availability
In the summer of 2011 we surveyed invertebrates of the forest floor to assess prey availability.
We collected four 0.25-m2 leaf litter samples (Oi, Oe, and Oa horizons) from random locations
within the salamander survey area, for aDraft total composite sample of 1-m2 per site. Leaf litter
arthropods were extracted using Berlese funnels and stored in 70% ethanol. Macroarthropods (>1
mm) were counted and identified to functional group (Table 2). Microarthropods (<1 mm) were
subsampled using a modified Stempel method designed for sampling zooplankton in aqueous
solution (Edmondson and Winberg 1971). The sample was put in 100 ml of 70% ethanol,
homogenized by stirring, and five 2-ml subsamples were withdrawn with a broad-based pipette.
In each subsample we counted the number of Collembola, Oribatida, and non-oribatid mites
(Acari) and calculated the mean and standard deviation of the five subsamples. We collected a
second set of four 0.25-m2 leaf litter samples (Oi, Oe, and Oa horizons) at each site for extraction
of litter snails. The samples were dried, sieved, and sorted under 2.5x magnification to remove
all snails. Nonparametric Spearman-rank correlations were used to determine if there was an
association (ρ > 0.5) between abundance of each prey group, prey richness (number of functional
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groups), and total invertebrate abundance with soil Ca availability and pH (Oa/A horizon), and foliar Ca (A. saccharum Ca concentration).
Salamander diet
Surveys for salamanders were conducted at each site from 17 June ― 4 August, 2011 and 23
June ― 6 July, 2012 and encompassed the area where previous soil and foliar sampling had occurred (Horsley et al. 2008). We conducted active cover searches for salamanders including under logs, rocks, fern mats, and bark. Salamanders were weighed to the nearest 0.1 g, and snout to vent (SVL; measured at anterior angle) and total length (TL) were measured to the nearest 0.1 mm. Stomach contents were obtained from salamanders > 28.0 mm SVL using gastric lavage
(Fraser 1976; Bondi et al. 2015). StomachDraft contents were stored in 70% ethanol and identified to functional group using a stereoscopic dissecting microscope with digital camera (Leica, Buffalo
Grove, USA). We measured the length and width of each prey item and calculated the volume as a cylinder.
A Multiple Response Permutation Procedure (MRPP) was used to test whether salamander diet composition (proportion of count for each given prey type of the total prey consumed; n/N) was different between the two years of data collection (2011 and 2012). For each prey group we calculated the number (total count of a given prey type), frequency (number of stomachs in which a given prey type was present), and total volume for each site. Importance values (Ix) were calculated for each prey group for all the stomach samples pooled per site using the equation:
Ix = [{(nx/N) + (vx/V) + (fx/F)}/3] x 100
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where nx = count of prey type x, N = total count of all prey items, vx = volume of a prey type x, V
= sum of volumes of all prey items, fx= the number of stomachs with prey type x, and F = sum of
the frequencies of all prey types (Anderson and Mathis 1999; Wheeler et al. 2007; Anthony et al.
2008; Milanovich et al. 2008). Importance values range between 0 and 100 and represent the
relative importance of a prey type to the overall diet. This index takes into account all 3 measures
of diet (count, volume, and frequency) so it reduces the bias of large and rare prey items.
Trophic strategy
Prey richness (number of prey groups), Shannon index of prey diversity (H’), total count of prey,
and total volume were calculated at the site level (all salamanders pooled) and for each
individual salamander. We calculated theDraft frequency of occurrence (FO; f/number of stomachs
analyzed) for each prey group at the site level. We used a Tukey HSD procedure to test whether
salamanders consumed different amounts of prey (count and volume), as well as richness and
diversity of prey, among the study sites representing a soil [Ca] gradient.
To evaluate the feeding strategy of salamanders, we used an approach proposed by
Costello (1990) and modified by Amundsen et al. (1996). For this method, the frequency of
occurrence (FO) is plotted on the x-axis and represents the number (Ni) of salamander stomachs
in which prey group i was present, divided by total number of stomachs (N). Prey-specific
abundance (Pi) is the sum (total number) of individuals consumed in prey group i, divided by the
sum of all stomach contents (total number) in only those predators that consumed prey i. This
graphical approach indicates feeding specialization for certain prey groups by their higher Pi
values along the y-axis — a generalist feeding behavior is indicated when most or all prey
groups have lower Pi values. Prey groups plotted at the upper left quadrant of the Costello plot
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(high abundance and low occurrence) are indicative of specialization by individual predators,
while those plotted in the upper right quadrant indicate specialization by the entire predator
population. We assessed Costello plots to determine whether salamanders at nutrient-poor sites
adopted a specialized diet on Ca-rich prey, with snails and oribatid mites having higher values
along the y-axis.
To investigate whether salamanders are selecting specific prey groups in greater
proportion to their availability, we calculated an index of selectivity at the site level as follows:
Li = ri – pi
where Li is the index of prey selectivity for prey group i, ri is the relative abundance of prey
group i in the salamander diet, and pi is the relative abundance of prey group i in the availability
sample (Strauss 1979). The selectivity indexDraft can have values ranging from 1.0 to -1.0. As Li approaches 1.0, the predator is actively selecting prey group i, and as Li approaches -1.0, the salamander is avoiding prey group i. As Li approaches 0, prey i is being consumed in proportion to its abundance in the environment. We expect that if salamanders require more dietary Ca at nutrient-poor sites, they will actively select for Ca-rich prey indicated by higher Li values.
Tissue nutrient analysis
To identify any nutrient deficiencies or differences among P. cinereus populations due to site- level biogeochemistry, we assayed elemental concentrations of whole salamanders. We collected
10 adult P. cinereus from each site in 2012. Salamanders were euthanized in the field using 20%
Benzocaine gel (Chen and Combs 1994), rinsed with distilled water, and transported to the lab on ice. Salamanders were freeze dried at -40°C and the dry mass of each individual was measured to the nearest .001 g on an analytical microbalance (Mettler Toledo, Columbus, OH).
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Each individual was acid digested in 10 ml 6N HN03 in an open, acid-washed crucible at 100°C
for 30 minutes. The digestate was filtered through #40 Whatman paper (Pittsburgh, PA, USA)
and diluted to 50 ml with distilled deionized water. We analyzed aliquots of samples for Ca, Mg,
K, Na, Mn, P, and S using inductively coupled plasma optical emission spectrometry (Perkin
Elmer Optima 3300 DV, Perkin Elmer, Waltham, MA, USA) following procedures described in
EPA method 200.7 (EPA 2004). We used nonparametric Spearman-rank correlations to
determine if there was an association (ρ > 0.5) between the P. cinereus tissue concentrations of
Ca, Mg, K, Na, Mn, P, and S, and the site-level measures of soil [Ca], foliar [Ca] and soil pH.
The methods used in this research to capture, handle, and collect salamanders meet the animal
ethics approval of the Institutional Animal Care and Use Committee of the State University of
New York, College of Environmental ScienceDraft and Forestry (IACUC #120403) and followed the
animal care guidelines provided in the Guide to the Care and Use of Experimental Animals
(Olfert et al. 1993).
Results
Prey availability
We collected and identified a total of 6,101 macroarthropods, 4,282 leaf litter snails, and
estimated a total abundance of 180,980 microarthropods from the forest floor samples (Table 3).
The most abundant prey groups at all the sites were microarthropods, which included oribatid
mites, non-oribatid mites, and springtails. The minimum estimated abundance of
microarthropods was 9,450 per m2 at site HIGH1 and the maximum was 29,220 per m2 at
VLOW1 (Table 3, Fig. 1A). Macroarthropod abundance ranged from 194 per m2 at HIGH1 to
1,108 per m2 at MOD2 (Fig. 1B) with spiders, other larvae (all insect groups excluding
Lepidoptera and beetles), and millipedes abundant at all sites (Fig. 1C).
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Overall there was no association between total invertebrate prey abundance or functional group richness and site-level measures of Ca availability and soil pH (Fig. 2). However, the abundances of some prey groups were significantly correlated with the Ca and/or pH gradient, including positive associations for snails, centipedes, and wasps, and negative associations for true bugs and Lepidoptera larvae. The remaining functional groups, including Ca-rich prey such as oribatid mites and millipedes, did not indicate sensitivity (in terms of changing abundance) to the Ca / pH gradient.
Salamander diet composition and trophic strategy
We collected stomach contents from 215 P. cinereus individuals over two sampling seasons: 104 in 2011, and 111 in 2012. From these dietDraft assays, a total of 2,402 prey items were identified to functional group (usually Order), with an additional 18 that were too degraded to be identified and excluded from further analysis. Using the MRPP procedure, we did not detect a significant difference in salamander diet composition between 2011 and 2012, all sites combined (A =
0.0022, P = 0.08). Given this result, and because we are describing the general diet of P. cinereus at these sites, we combined both years of data for analysis.
We found little or no evidence that P. cinereus trophic stagey, based on metrics of prey count, volume, richness, and diversity of prey, varied consistently across the nine sites along the gradient of soil-Ca availability (Fig. 3). Salamanders at the two Ca-poor sites (VLOW1 and
VLOW2) ate the highest number of prey with a higher average prey count per stomach compared with five other sites (F8,206 = 4.54, P < 0.001). Site had no effect on mean prey volume (F8,206 =
0.91, P = 0.51), although salamanders at MOD2 had the highest mean prey volume (25.9 mm3) due to the high consumption of larvae at this site (Table 4). While there was an effect of site on
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prey richness, (F8,206 = 2.3, P = 0.02,), there was no trend in pair-wise differences associated
with site-level Ca availability. Prey diversity of salamander diets was similar across the nine sites
(F8,206 = 1.13, P = 0.34), in part because of high within-site (across individual) variance observed
in this index (Fig. 3D).
Eastern Red-Backed Salamanders had a diverse diet including 21 prey functional groups
(Table 3). Overall, the prey groups most often present in salamander stomachs (F) and consumed
in highest abundance (n) were microarthropods including springtails, non-oribatid mites, and
oribatid mites. Springtails were found on average in 70% (SD = 25.5 and range of 34% to 88%)
of all salamander stomachs and accounted for 24% of the total prey consumed. Non-oribatid
mites were found on average in 69% (SD = 30.7), and oribatid mites in 59% (SD = 14.6), of all
the salamander diet samples collected. TerrestrialDraft snails, a Ca-rich prey source, were present in
an average of 31% of salamander stomachs (SD = 17.3), but only accounted for ~5% of total
prey consumed (range of 2% - 11%), suggesting that gastropods are commonly present in P.
cinereus diet but in low numbers. Adult beetles, beetle larvae, and other larvae contributed the
most to prey volume, and at one site (HIGH2) Lepidoptera larvae also contributed significantly
to prey volume. The relative importance of prey groups (Ix) to salamander diet among the sites
was variable, but overall, springtails, oribatid mites, other larvae, adult beetles, and non-oribatid
mites were the most important (Fig. 4). Calcium-rich prey (oribatid mites and snails) were not a
more important component of salamander diet at sites with low soil Ca versus moderate-Ca and
high-Ca sites.
Plethodon cinereus exhibited a generalist feeding strategy that was unrelated to site
measures of Ca availability or pH, based on Costello-Admunsen plots used to visualize patterns
in frequency of prey consumption (Fig. 5). We observed no evidence of a more specialized diet
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on Ca-rich invertebrates at the nutrient-poor sites, as all prey groups had Pi values < 0.5 at all sites. Most prey groups had low frequency of occurrence and low prey-specific abundance, and therefore are found in the lower left quadrant of the plot (Fig. 5), indicating few individuals consumed these prey and in small numbers. The plots confirm that 3 functional groups – mites
(oribatid and non-oribatid) and springtails – are consistently found in salamander stomachs. For all sites, non-oribatid mites were projected in the lower right of the graph, indicating they were frequently found in salamander stomachs (>0.5 F) and consistently eaten by many salamanders in a population, even if in low numbers. Oribatid mites and springtails were also frequently projected in the lower right corner of the plot, indicating they were present in the stomachs of many salamanders at all sites.
Prey selection Draft
Plethodon cinereus did not select for Ca-rich prey groups (oribatid mites and snails) in relation to Ca availability or soil pH. All of the macroinvertebrate groups, apart from other insect larvae, were eaten in proportion to their availability, based on Li values near zero (Fig. 6). Insect larvae
(excluding Lepidoptera and beetle larvae) were selected for at most of the sites with Li values between 0.1-0.2, except sites LOW1, MOD3, and HIGH2 with Li values < 0.1. Salamanders selected against oribatid mites at all of the sites, with negative selection indices ranging from -
0.4 at the lowest Ca site (VLOW1) to -0.61 at a low-ca site (LOW2). Selection behavior varied among sites for the other microarthropod groups (non-oribatid mites and springtails), with Li values fluctuating between negative and positive values across the sites. For example, salamanders selected for springtails (Li > 0.1) at five sites, and at four sites salamanders ate springtails in proportion to their availability. Salamanders consumed non-oribatid mites in
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proportion to their availably at seven sites, whereas moderately selected for them (Li > 0.1) at
LOW1, and exhibited negative selection at the lowest-Ca site (VLOW1).
Salamander body composition
Elemental concentrations of P. cinereus bodies (Table 4) were closely similar to those reported
from individuals collected in New Hampshire (Burton and Likens 1975) and P. serratus
(Grobman, 1944) in Missouri (Semlitsch et al. 2014). For all but one of the elements analyzed
(Mg, K, Ca, Na, S, and P), there was no association between elemental concentrations and soil or
foliar Ca, or soil pH (Table 4). We did observe a negative association between Oa/A soil Ca and
tissue Mn concentration (ρ = -0.57), and foliar Ca concentrations and salamander Mn
concentrations (-0.60) indicating that theDraft P. cinereus populations at nutrient poor (lowest Ca
availability in both the soils and leaf litter) sites (VLOW1, VLOW2, and LOW1) had the highest
Mn concentrations in whole-body assays.
Discussion
Our study is the first to assay salamander diets along a regional gradient in nutrient (Ca)
availability and habitat condition (substrate pH), and using this approach, the first to consider
dietary behavior among the potential mechanisms allowing P. cinereus populations to persist in
severely acid-impaired forests with substrate pH < 3.5 where, until very recently, it was thought
they could not survive. Our findings confirm that plethodontid salamanders are dietary
generalists that rely on a diverse food base (Burton 1976; Maerz et al. 2005; Wheeler et al. 2007;
Snyder 2011), in which microarthopods, insect larvae, and beetles were the most important prey
(Jaeger 1972; Maglia 1996; Adams and Rohlf 2000), and that this generalist predation strategy is
maintained even in chronically acidified forests where Ca has been severely depleted at the
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ecosystem level, from soils and vegetation. Our results provide support for the hypothesis that a generalist feeding behavior confers some degree of tolerance of P. cinereus populations to habitat degradation in an acidified landscape (Bondi et al. 2016).
Yet this generalist strategy, in which salamanders consume a variety of prey, can only function to help P. cinereus tolerate acid-impaired habitats if the food web available remains sufficiently intact. Our results confirm this important corollary – we found little evidence that prey availability changes in quantity or quality across our large study gradient. If prey availability and/or nutritional quality were lower in forests where Ca has been anthropogenically depleted, perhaps P. cinereus would be unable to persist, given that Ca transport from soil solution via the epithelium is likely driven by environmental concentrations (Zerella and Stiffler
1999). We did observe that P. cinereus Draftdwelling in the most Ca-poor, low-pH forests on average consume more prey items than P. cinereus in more moderate and Ca-rich forests. Salamanders relied heavily on smaller prey, especially mites and springtails, but likely can do so efficiently because of their great abundance in the soil. Microarthropods are extremely abundant (> 10,000 individuals/m2) and miniscule (< 0.5 mm3) and would require a very low energy expenditure and low risk of injury to capture due to their high density and small size (Jaeger and Barnard 1981).
Jaeger et al. (1990) found P. cinereus will indiscriminately forage on most abundant prey types during stressful foraging conditions (dry litter), whereas during wet conditions when salamanders are able to more actively forage they avoid larger, chitinous prey. It may be that during stressful conditions associated with low nutrient availability and acidic soils, P. cinereus are more apt to consume the highly abundant microarthropods that are a minimal energy expenditure to capture and low digestive time.
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Species traits of prey groups also illustrate how the soil food web may function to sustain
predator populations in acid-impaired, Ca-deprived forests. Land snails were the only prey group
that were both important to salamander diets, and sensitive to soil and foliar Ca availability at the
site-level. However, land snails are adapted to a wide range of Ca availability and pH, including
generalist taxa and acidophilic specialists (Tattersfield et al. 2001; Hotopp 2002; Nekola 2010).
Even at the most acidic, Ca-poor sites, we estimated snail densities between 38–140 individuals
per m2. Given that there are typically 0.05-0.4 salamanders per m2 in hardwood forests
(Semlitsch et al. 2014), with maximum estimates of 2.8 salamanders per m2 in Virginia (Mathis
1991), snail abundance should be sufficient to supplement diets as an occasional, but highly
concentrated Ca source. By contrast, we found no associations between the abundance of
oribatid mites – overall the most importantDraft prey group that is known to be nutritionally rich in Ca
– and site-level measures of Ca availability or substrate pH. Although Ca is a substantial
component of the cuticle of oribatid mites (Gist and Crossley 1975; Norton and Behan-Pelletier
1991), their overall abundance is not linked with environmental Ca, although some species are
known to be acidophilic (Hågvar and Amundsen 1981; Hagvar 1984; Hågvar 1990; Fisk et al.
2006). The Ca carbonate of oribatid mite cuticle may derive from consumption of calcium
oxalate produced by wood-decaying fungi in the presence of oxalic acid and so may not be
linked to soil Ca availability or litter quality (Cromack et al. 1977; Norton and Behan-Pelletier
1991).
Salamanders consumed most prey groups in proportion to their availability and exhibited
moderate selection at some sites for insect larvae, Collembolans, and non-oribatid mites.
Selection for these prey types are in line with optimal foraging choices for easily digestible
(insect larvae and Collembolans) or easily captured (mites) prey compared with larger, more
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active prey (Jaeger 1990). While the selectivity index showed that salamanders did not select for
oribatid mites, we caution the interpretation that this prey group is actively avoided. Given their
overwhelming abundance in the soil and leaf litter samples (~6,000 – 18,000 individuals/meter2) a salamander’s diet would have to consist solely of oribatid mites for the proportion of stomach contents to be higher than the proportion they are available in the environment. In addition, although they are highly abundant, it is likely that oribatid mites are encountered less frequently by a foraging salamander than other microarthropods and larger invertebrates. Oribatid mites are less mobile than Collembolans and spend more time sedentary within the minute interstices of the fragmentation layer of leaf litter (R. Norton personal communication). A large predator moving through the forest floor, such as a salamander, cannot access these tiny spaces where most oribatid mites reside. Therefore, theDraft mites extracted from our soil and leaf litter samples likely include many oribatid mites that are never encountered by a foraging salamander.
We sampled salamander stomach contents only during the summer months and therefore did not capture seasonal variation in feeding behavior. Yet because our study sites were at the northern extent of the P. cinereus range, and located at moderate to higher elevations, the study populations have a relatively short active season that coincides with the summer months.
Therefore, the summer diet likely reflects their overall diet because of the shorter foraging season. Because surveys did not coincide with rainy nights, salamander diets were documented under relatively warm and dry conditions when salamanders do not travel far from cover objects to forage. During dry conditions salamanders spend more time under cover objects (e.g. logs, rocks, and burrows) and move deeper into the soil to avoid desiccation (Heatwole 1960; Taub
1961). Fraser (1976) and Jaeger (1972) observed salamanders with full stomachs following rain events and empty stomachs in between, and concluded that predator mobility, not prey
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availability, is the primary cause of low prey ingestion during dry weather. Therefore, the diets
described in this study likely represent what salamanders eat when they are limited spatially by
drier substrate conditions, and not constrained by food availability.
We found no indication that salamanders on Ca-poor sites suffered any deficiency of Ca
or other nutrients, such as Na or Mg, in their body tissues; except for elevated manganese (Mn)
levels on acid-impaired sites, which is discussed below. Our results for Ca, K, Mg, Na, P, and S
are very similar to those reported by Burton and Likens (1976) for P. cinereus in the White
Mountains of New Hampshire, indicating there is little temporal or spatial variation in
salamander body composition. Although our results for tissue Na may be inconsistent with
earlier work that found higher Na efflux rates for P. cinereus on low pH substrates (Wyman and
Hawksley-Lescault 1987), we did not measureDraft Na efflux and cannot make a direct contrast.
Elevated levels of manganese (Mn) in tissues of salamanders dwelling on more acidic
sites is a potential indication of diet-mediated stress for P. cinereus in acid-impaired forests. We
observed nearly five-times higher tissue [Mn] in salamanders dwelling at the low Ca end-
member site LOW1 (0.14 mg/g or 140 ppm), compared to the high Ca end-member site HIGH2
(0.03 mg/g or 30 ppm). Manganese is naturally abundant in the environment and its
biogeochemistry is complex and poorly understood in forests (Bradl 2004; Millaleo et al. 2010).
As an essential micronutrient to plants and animals, low levels of Mn are required for many
metabolic processes; however, it becomes toxic in excessive amounts. In acidic soils, Mn oxides
are reduced on cation exchange sites and the concentration of soluble Mn2+ (the mineralized
form available for plant uptake) in the soil solution is increased (Kogelmann and Sharpe 2006).
Manganese has been shown to bioaccumulate in tissues of organisms at lower trophic levels and
animal tissue concentrations are associated with Mn concentrations in the environment (ATSDR
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2012). Because salamanders are consuming larger numbers of invertebrates at the base of the detritus food web (e.g., fungivore and detritivore Acari and Collembola), they may ingest higher levels of Mn than populations in more base-rich forests. Together, these points suggest that Mn may be more bioavailable, both in the food web and via passive adsorption, to P. cinereus and other upper-level consumers in more acidic forests, but further study is needed to evaluate the role of dietary Mn as a potential concern.
Our findings support that the trophic ecology of eastern redback salamanders contributes to their ability to persist in acid-impaired forest habitats, based on four lines of evidence. First, we confirmed that P. cinereus were generalist predators across a broad regional gradient in calcium availability and soil pH, from well-buffered to heavily acidified sites. Second, P. cinereus consumed most prey groups inDraft direct proportion to their abundance, except for one functional group (oribatid mites) that was so highly abundant, a proportional representation in P. cinereus diet would have excluded all other prey groups. Third, we observed that the environmental abundance (or dietary availability) of nearly all prey groups, including Ca-rich microfauna such as oribatid mites, did not vary consistently across the broad range of soil Ca and pH found in northern hardwood forests. Lastly, extant P. cinereus populations in severely acid- impaired forests exhibited no Ca or nutrient deficiencies in their body tissues, aside from elevated Mn levels, which may be of concern. Overall these findings help to explain recent observations of P. cinereus populations persisting in acid-impaired habitats, with substrate pH <
3.5, that for many decades have been thought to be fatal to plethodontid salamanders (Bondi et al. 2016; Moore and Wyman 2010). The generalist strategy of this keystone predator is well established, but we have shown that not only is this trait conserved by populations in degraded habitats, but that the strategy ‘works’ for P. cinereus because its food webs remain
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compositionally intact, providing ample prey, including Ca-rich taxa, for it to consume without
specializing on certain prey groups. Of the ten most important functional groups in salamander
diets, only did snails (Gastropoda) exhibit any association with the soil Ca or pH gradient, yet
these were still in significant – and likely more than sufficient – abundance in even the most
acidified, Ca-poor forests. Therefore P. cinereus does not need to specialize on catching large or
very Ca-rich, and potentially risky, prey in habitats where its other biological pathway for Ca
assimilation (from aqueous soil solution through the skin) is limited or negated. Our findings
suggest that Plethodon cinereus is buffered from direct impacts of acid pollution by their own
generalist traits, as well as the persistence of a functional food web, despite dramatic changes in
the biogeochemistry of forest habitats. A diversity of traits within soil functional groups, such as
pH sensitivity and nutrient assimilation Draftphysiology, may therefore play a significant ‘bottom-up’
role in mediating impacts of an environmental stressor on apex predators like the Eastern Red-
Backed Salamander.
Acknowledgements
This research was supported by a grant from the US Forest Service Northeastern States
Research Cooperative (www.nsrc.org). Chelsea Geyer, Jamie Wahls, Amanda Temple, Drew
Smith, and Sabrina Green provided field and laboratory assistance, and Scott Bailey, Steve
Horsley, Robert Long, and Richard Hallett provided soil chemistry data from their research sites.
The salamander body tissues analysis was done with assistance from Chuck Schirmer and
Deborah Driscoll at the SUNY College of Environmental Science and Forestry Analytical and
Technical Services and Forest Soils Laboratories. This research was conducted on the property
of the Green Mountain National Forest, White Mountain National Forest, and the Equinox
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Preservation Trust. Scientific Collecting permits were acquired from New Hampshire Fish and
Game and Vermont Fish and Wildlife to collect salamanders and obtain diet samples, and methods were approved by the Institutional Animal Care and Use Committee of the State
University of New York, College of Environmental Science and Forestry (IACUC #120403).
Draft
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Figure 1. Abundance of microarthropods with standard deviation bars from subsamples (A), counts of macroinvertebrates (B), and other invertebrate prey groups (C) at nine sites in New Hampshire and Vermont. Figure 2. Spearman rank coefficients of pairwise comparisons between Oa/A soil Ca (A), foliar Ca (B) and Oa/A pH (C) and abundance of 21 invertebrate prey groups at nine sites in New Hampshire and Vermont. Significance of correlation was assessed at ρ > 0.5. Figure 3. Interval plots of mean (values shown to the right of symbol) and 95% confidence intervals for total count of prey (A), total prey volume, (B) richness (C) and diversity (D) of prey groups, per a salamander stomach. Sites that do not share the same letter are significantly different from one another using Tukey pairwise comparisons.
Figure 4. Importance values (converted to percentages) for the 21prey groups found in Plethodon cinereus stomachs from nine sites in New Hampshire and Vermont. Symbol colors represent the sites grouped as low (white), moderate (grey), and higher (black) Oa/A soil-Ca availability.
Figure 5. Modified Costello plots of the feeding strategy for P. cinereus at nine sites in New Hampshire and Vermont. Sites are arranged by level of Oa/A soil Ca and indicated on the top left of each graph. On the y-axis is prey-specific abundance (Pi), which represents the total number of prey group i divided by the sum of all stomach contents in only those predators that consumed prey i. Frequency of occurrence (FO) isDraft plotted on the x-axis and is the number (Ni) of salamander stomachs in which prey group i was found out of the total number of stomachs analyzed. Prey groups are symbolized as: ants = 1, bark lice = 2, adult beetles = 3, beetle larvae = 4, centipedes = 5, adult flies = 6, other larvae = 7, lepidoptera larvae = 8, millipedes = 9, non- oribatid mites = 10, oribatid mites = 11, pseudoscorpions = 12, snails = 13, spider = 14, springtails = 15, true bugs = 16, and wasps = 17.
Figure 6. Plethodon cinereus prey selectivity index from nine sites in New Hampshire and Vermont that represent a range of Oa/A soil Ca availability. Plotted along the y-axis are Li values, for which >0 indicate selection, <0 indicate avoidance and values at or around 0 indicate salamanders consume a particular prey group in proportion to its availability in the environment
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Abundance of microarthropods with standard deviation bars from subsamples (A), counts of macroinvertebrates (B), and other invertebrate prey groups (C) at nine sites in New Hampshire and Vermont.
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Spearman rank coefficients of pairwise comparisons between Oa/A soil Ca (A), foliar Ca (B) and Oa/A pH (C) and abundance of 21 invertebrate prey groups at nine sites in New Hampshire and Vermont. Significance of correlation was assessed at ρ > 0.5.
152x304mm (300 x 300 DPI)
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Interval plots of mean (values shown to the right of symbol) and 95% confidence intervals for total count of prey (A), total prey volume, (B) richness (C) and diversity (D) of prey groups, per a salamander stomach. Sites that do not share the same letter are significantly different from one another using Tukey pairwise comparisons.
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Importance values (converted to percentages) for the 21prey groups found in Plethodon cinereus stomachs from nine sites in New Hampshire and Vermont. Symbol colors represent the sites grouped as low (white), moderate (grey), and higher (black) Oa/A soil-Ca availability.
254x190mm (300 x 300 DPI)
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Modified Costello plots of the feeding strategy for P. cinereus at nine sites in New Hampshire and Vermont. Sites are arranged by level of Oa/A soil Ca and indicated on the top left of each graph. On the y-axis is prey- specific abundance (Pi), which represents the total number of prey group i divided by the sum of all stomach contents in only those predators that consumed prey i. Frequency of occurrence (FO) is plotted on the x-axis and is the number (Ni) of salamander stomachs in which prey group i was found out of the total number of stomachs analyzed. Prey groups are symbolized as: ants = 1, bark lice = 2, adult beetles = 3, beetle larvae = 4, centipedes = 5, adult flies = 6, other larvae = 7, lepidoptera larvae = 8, millipedes = 9, non-oribatid mites = 10, oribatid mites = 11, pseudoscorpions = 12, snails = 13, spider = 14, springtails = 15, true bugs = 16, and wasps = 17.
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Plethodon cinereus prey selectivity index from nine sites in New Hampshire and Vermont that represent a range of Oa/A soil Ca availability. Plotted along the y-axis are Li values, for which >0 indicate selection, <0 indicate avoidance and values at or around 0 indicate salamanders consume a particular prey group in proportion to its availability in the environment.
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1 2 Table 1. Soil (Oa/A horizons) and foliar chemistry, topographic, and environmental characteristics for nine sites in the White (NH-) 3 and Green (VT-) Mountains of New Hampshire and Vermont where Eastern Red-Backed Salamander (Plethodon cinereus) diet and 4 tissue samples were collected in the summers of 2011 and 2012.
Site Region Latitude Longitude Soil pH Soil Ca* Foliar Ca† Elevation‡ Slope‡ Aspect‡ Basal Area⸹ VLOW1 NH 43.90723 -71.61177 3.30 0.52 3,871 578 23 SE 30.58 VLOW2 VT 43.36156 -72.93236 3.44 0.89 4,383 515 15 SW 35.44 LOW1 VT 44.09015 -73.05123 2.73 3.43 3,505 589 15 SW 34.55 LOW2 NH 44.03436 -71.89082 3.22 3.70 8156 720 26 SW 23.47 MOD1 NH 43.97367 -71.18802 3.89 5.30 9,410 250 7 S 44.92 MOD2 VT 43.16293 -73.09455 3.68 7.71 11,956 481 15 E 28.00 MOD3 VT 43.36733 -72.9128 2.81 8.40 5,592 654 13 SE 34.51 HIGH1 VT 43.38507 -72.84328 3.40Draft 16.54 5,636 672 33 N 49.32 HIGH2 NH 43.98785 -71.90552 3.84 17.32 11,705 444 28 NE 62.91 5 -1 6 *Soil Ca is in cmol+∙kg 7 †Foliar content is mg∙kg-1 8 ‡Elevation (m) and slope (%) are based on USGS digital elevation models (Horsley et al., 2008). Aspect was measured in the field. 9 ⸹Total basal area (BA; m2∙ha-1) of overstory. 10
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11 Table 2. Invertebrate prey availability (individuals/m2) extracted from leaf litter samples collected from sites in New Hampshire and 12 Vermont that represent a gradient of Oa/A soil Ca availability; sites are ordered from left to right with increasing Oa/A soil Ca. Prey group VLOW1 VLOW2 LOW1 LOW2 MOD1 MOD2 MOD3 HIGH1 HIGH2 Total Oribatid mites* 15,920 8,690 9,710 14,830 15,380 15,620 14,380 6,220 17,630 118,380 Non-oribatid mites* 5,950 1,300 3,480 2,970 4,980 5,590 2,150 1,040 5,230 32,690 Springtails* 7,350 1,770 2,760 2,310 2,090 6,360 3,030 2,190 2,050 29,910 Snails 140 40 38 1,517 484 660 257 215 931 4,282 Other larvae 131 70 181 76 121 518 210 59 97 1,463 Spiders 219 49 133 159 146 37 38 44 249 1,074 Millipedes 92 19 48 207 169 62 15 20 89 721 Beetles, larvae 125 42 92 55 101 100 33 16 131 695 Beetles, adults 23 23 183 13 147 132 28 14 35 598 Pseudoscorpiones 56 22 18 115 36 25 27 2 49 350 Bark lice 116 0 43 Draft47 26 15 3 3 34 287 Flies 29 17 35 35 46 30 19 21 20 252 Ants 12 6 39 0 39 110 22 5 17 250 Centipedes 18 7 29 47 43 55 6 4 33 242 Annelid worms 15 4 69 1 16 40 1 12 12 170 Lepidoptera larva 8 2 4 1 1 2 3 1 2 24 Slugs 2 4 2 4 2 4 4 2 0 24 True bugs 4 2 4 6 2 1 2 0 2 23 Thrip 0 0 2 0 9 5 0 0 0 16 Harvestmen 0 4 1 0 2 2 1 0 0 10 Wasps 12 2 6 6 23 10 1 3 9 72 Totals Macroinvertebrates 1,002 313 927 2,289 1,413 1,808 670 421 1,710 10,553 Microinvertebrates* 29,220 11,760 15,950 20,110 22,450 27,570 19,560 9,450 24,910 180,980 All invertebrates 30,022 12,073 16,877 22,399 23,863 29,378 20,230 9,871 26,620 191,533 13 *Numbers represent means derived from 5–1ml subsamples. 14
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15 Table 3. Diet composition of Eastern Red-Backed Salamander (Plethodon cinereus) from New Hampshire and Vermont. n = 16 percentage of total prey consumed by all salamanders and v = percentage of total prey volume of all salamanders at a site. F = 17 percentage of salamander stomachs containing a given prey group at a site. Sites are organized left to right in increasing order of Oa/A 18 soil Ca.
VLOW1 VLOW2 LOW1 LOW2 MOD1 MOD2 MOD3 HIGH1 HIGH2 Prey Group n v F n v F n v F n v F n v F n v F n v F n v F n v F Annelid worms 3 5 6 0 0 0 0 0 0 0 0 0 1 0 6 1 1 3 0 0 0 0 0 0 0 0 0 Ants 1 4 10 0 1 4 0 0 0 0 0 0 16 24 66 5 7 44 6 10 17 0 0 0 1 1 8 Bark lice 1 1 16 0 0 0 0 0 0 7 4 36 1 0 6 1 0 6 1 0 9 0 0 0 3 1 17 Beetles, Adults 2 12 29 4 16 42 10 54 36 5 26 32 2 7 16 12 27 78 1 11 13 2 15 23 3 6 17 Beetles, larvae 2 11 32 2 16 38 1 1 9 5 8 36 2 12 19 2 4 19 3 6 26 2 6 23 3 23 21 Centipedes 0 0 3 0 0 0 0 0 0 0 0 4 0 3 3 2 1 19 0 0 0 0 0 0 1 0 4 Flies, Adults 2 4 32 4 6 33 7 11 36 2 2 20 6 12 38 1 1 13 6 8 39 7 17 46 2 2 13 Harvestmen 0 0 0 0 0 0 0 0 0 Draft 0 0 0 0 1 3 0 0 0 0 0 0 0 0 0 0 0 0 Larvae, other 25 30 61 12 19 42 10 11 27 9 21 36 15 16 22 28 40 72 2 3 17 12 32 46 5 2 25 Lepidoptera larva 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 9 3 0 38 4 0 0 0 2 45 17 Millipedes 1 1 16 1 1 8 0 0 0 3 3 24 0 0 3 1 0 6 1 2 9 3 4 31 2 1 13 Mites, non oribatid 7 1 65 14 3 79 33 4 73 13 1 68 19 1 69 23 3 75 12 1 65 15 3 77 27 1 54 Oribatid mites 13 2 81 25 3 83 9 0 45 5 0 32 8 0 41 13 1 59 18 1 74 13 2 69 13 1 50 Pseudoscorpions 1 0 13 1 0 17 0 0 0 0 0 4 0 0 0 0 0 3 0 0 0 1 0 8 0 0 0 Slugs 0 0 0 0 10 4 0 0 0 0 0 0 0 8 3 0 0 0 0 0 0 0 0 0 0 0 0 Snails 3 5 23 5 8 42 4 5 27 3 2 16 12 5 47 5 1 41 7 5 35 2 4 23 7 4 29 Spiders 3 5 42 3 8 38 4 8 27 5 14 48 3 3 19 1 1 13 4 4 26 5 7 38 4 5 25 Springtails 31 14 87 26 7 83 20 4 55 38 17 84 8 3 47 3 1 34 36 9 74 38 9 77 24 3 88 Thrips 0 0 3 0 0 0 0 0 0 0 0 0 0 0 0 0 0 3 0 0 0 0 0 0 0 0 0 True bugs 1 1 16 0 0 4 1 1 9 2 1 12 0 0 3 0 0 0 0 1 4 0 0 0 1 0 4 Wasps, Adult 2 2 26 1 2 17 0 0 0 3 1 20 5 3 22 3 3 28 1 1 13 1 1 8 3 3 17 Stomach samples 31 24 11 25 32 32 23 13 24 19 20
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Table 4. Mean and standard deviation (in italics) for elemental analysis of Eastern Red-Backed Salamander (Plethodon cinereus) body composition for Ca, K, Mg, Mn, K, P, and S (ppt; mg∙g-1) collected from nine sites in the White (NH) and Green (VT) Mountains of New Hampshire Vermont in order of increasing Oa/A soil Ca from top to bottom. Spearman correlation coefficients (ρ) of body tissue concentrations with Oa/A soil Ca and pH, and foliar Ca concentrations are listed on the bottom, along with body tissue concentrations reported for other Plethodon populations.
Site Region N Calcium Potassium Magnesium Manganese Sodium Phosphorous Sulfur
VLOW1 NH 10 25.42 7.19 1.23 0.11 2.36 17.55 5.86 (6.14) (1.26) (0.11) (0.04) (0.62) (2.84) (0.42) VLOW2 VT 9 27.97 7.55 1.46 0.12 2.90 19.64 6.24 (3.91) (1.38) (0.29) (0.06) (0.57) (2.17) (0.64) LOW1 VT 10 28.39 7.54 1.49 0.14 3.02 20.41 6.65 (6.16) (1.80) (0.18) (0.08) (0.59) (2.40) (0.66)
LOW2 NH 10 26.10 7.98 1.26 0.04 2.70 18.07 5.69 (2.94) (1.27) (0.15) (0.02) (0.56) (1.50) (0.62)
MOD1 NH 10 26.39 7.09 1.35 0.08 2.54 17.99 5.92 (3.27) (1.12) (0.26) (0.08) (0.35) (1.55) (0.62)
MOD2 VT 10 29.02 6.67 1.36 Draft0.05 2.50 19.37 5.90 (3.63) (0.86) (0.21) (0.03) (0.46) (1.82) (0.58)
MOD3 VT 10 28.28 8.96 1.40 0.05 3.04 19.70 6.11 (4.36) (1.85) (0.13) (0.02) (0.55) (2.04) (0.55)
HIGH1 VT 9 25.40 7.95 1.33 0.07 2.79 18.46 6.06 (5.01) (0.81) (0.17) (0.02) (0.56) (1.92) (0.80)
HIGH2 NH 10 26.26 6.75 1.30 0.03 2.29 18.59 5.92 (2.61) (1.35) (0.11) (0.01) (0.46) (1.42) (0.54)
All sites 98 27.03 7.51 1.35 0.08 2.68 18.86 6.04 Spearman ρ- OA/A soil Ca 0.00 0.00 0.01 -0.57 -0.09 0.01 -0.10
Spearman ρ- OA/A soil pH -0.03 -0.33 -0.17 -0.26 -0.32 -0.15 -0.19 Spearman ρ- Foliar Ca 0.02 -0.24 -0.16 -0.60 -0.28 -0.10 -0.26
P. cinereus- New Hampshire* 28.5 7.4 1.3 NA 2.5 18.6 5.9 P. serratus- Missouri* 29.9 8.1 1.3 NA 6.1 21.6 6.3
*Values reported in Burton and Likens (1976) and Semlitsch et al. (2014)
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