The literature review yielded 290 animal-diet discrimination factor estimates of carbon and 268 animal-diet discrimination factor estimates of nitrogen from 66 publications (Supporting Information, Table S1) distributed (for Δ13C and Δ15N, respectively) as follows: mammals (95 and 89), birds (61 and 52), fishes (41 and 47), reptiles (3 and 3), and invertebrates (90 and 77). The overall mean estimates for Δ13C and Δ15N were 0·75‰ (SE = 0·11) and 2·75‰ (SE = 0·10), respectively.
effect of tissue
We initially analysed all taxa combined. Discrimination factors of carbon and nitrogen differed among tissues (F9,220 = 1·93, P = 0·049, and F8,198 = 2·71, P = 0·007, respectively). Consequently, we analysed differences among the consumer classes for muscle, plasma, liver, blood and the whole body (Fig. 2); other tissues did not have sufficient data to carry out this analysis. The carbon discrimination factor for muscle was significantly different among birds, fishes and mammals (F2,21 = 9·15, P = 0·001), but the differences were not significant for the nitrogen discrimination factor (F2,18 = 1·86, P = 0·184). For plasma, both the carbon and the nitrogen discrimination factors were not significantly different between birds and mammals (F1,16 = 3·52, P = 0·079 and F1,16 = 2·53, P = 0·131, respectively). The carbon discrimination factor for liver did not differ significantly among birds, fishes and mammals (F2,16 = 2·92, P = 0·083), but did differ significantly for the nitrogen discrimination factor (F2,16 = 6·67, P = 0·008, Fig. 2). For blood, the carbon discrimination factor was no different between birds and mammals (F1,13 = 0·05, P = 0·834), but the difference was significant for the nitrogen discrimination factor (F1,11 = 5·52, P = 0·039). For the whole body, the carbon discrimination factor was significantly different between invertebrates and fishes (F1,80 = 9·14, P = 0·003), but no significant differences were found for the nitrogen discrimination factor (F1,68 = 0·13, P = 0·721).
Figure 2. Mean (± SE) Δ13C and Δ15N within animal consumer classes among different tissues. Each pictogram above the bar indicates the classes included in each tissue analysis (symbols as in Fig. 1). Numbers inside the bar indicate the sample size. When a tissue had significant differences within classes, the mean for all classes is represented by a bar with a dotted line, and the mean of each class is represented inside by a bar with a solid line. Four tissues (hair, feather, collagen and red blood cells) had no categories or insufficient data for assessment of differences within animal consumer classes.
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effect of diet
Our initial analysis of all taxa combined showed significant negative relationships between discrimination factors and their diet isotopic ratios (F1,230 = 51·58, P < 0·001, R2 = 0·19 and F1,210 = 50·54, P < 0·001, R2 = 0·16, for ΔC and ΔN, respectively). We then performed independent GLMMs for these relationships within each consumer class (birds, mammals, fishes and invertebrate) and, taking into account where possible the type of tissue, the environment and the lipid extraction treatment (we used one of these variables when more than one category was available and valid, as described in the Statistical analysis section). In general, we found the same trend as in the initial analysis (all combined taxa) of significant negative relationships between discrimination factors and their corresponding isotopic ratios (Table 1). However, in some consumer classes, the relationship was not significant, as described below.
Table 1. Factors affecting the carbon and nitrogen discrimination factors in general linear mixed models. The analysed variables are presented for each consumer class and in italics are the ones significant in the full model
|Variables||dfn, dfd||F||P||Variables||dfn, dfd||F||P|
|Mammal||Diet δ13C||1, 64||47·70||< 0·001||Diet δ15N||1, 64||90·23||< 0·001|
|Tissue||5, 64||0·23||0·949||Tissue||5, 64||9·72||< 0·001|
|Lipid||1, 64||0·02||0·901||Lipid||1, 64||0·23||0·632|
|Bird||Diet δ13C||1, 24||5·32||0·030||Diet δ15N||1, 22||1·45||0·242|
|Environment||2, 24||1·83||0·183||Environment||2, 22||5·39||0·012|
|Tissue||4, 24||6·67||< 0·001||Tissue||4, 22||11·82||< 0·001|
|Lipid||1, 24||3·08||0·092||Lipid||1, 22||11·30||0·003|
|Fish||Diet δ13C||1, 18||8·60||0·009||Diet δ15N||1, 21||9·08||0·007|
|Tissue||2, 18||13·39||< 0·001||Tissue||2, 21||14·48||< 0·001|
|Environment||1, 18||1·12||0·304||Environment||1, 21||0·01||0·937|
|Lipid||1, 18||1·31||0·267||Lipid||1, 21||2·01||0·171|
|Invertebrate||Diet δ13C||1, 67||5·85||0·018||Diet δ15N||1, 55||30·36||< 0·001|
|Environment||1, 67||0·82||0·368||Environment||1, 55||0·18||0·675|
For mammals, only two categorical independent variables were added to the GLMM: the lipid extraction treatment and the type of tissue (six categories: blood, red blood cells, hair, liver, muscle and plasma). The carbon discrimination factor was negatively correlated with the diet carbon isotopic ratio, but none of the categorical variables was significant in the full model (Table 1). The nitrogen discrimination factor showed differences among tissues and a significant negative correlation with the diet nitrogen isotopic ratio (Table 1). The GLM on the relationships between discrimination factors and their corresponding diet isotopic ratios confirmed these results, showing significant relationships for both carbon and nitrogen (Δ13C: F1,88 = 91·44, P < 0·001, R2 = 0·51; and Δ15N: F1,78 = 14·25, P < 0·001, R2 = 0·15; Fig. 3a,b). For the nitrogen discrimination factor and within tissues, the GLM between Δ15N and diet isotopic values were only significant for muscle, liver and plasma (F1,13 = 17·74, P = 0·001, R2 = 0·58; F1,14 = 8·17, P = 0·013, R2 = 0·37; and F1,17 = 19·17, P < 0·001, R2 = 0·53, respectively).
Figure 3. Relationship for each taxonomic class between estimates of Δ13C and carbon diet isotopic ratio δ13C, and estimates of Δ15N and nitrogen diet isotopic ratio δ15N. Each taxonomic class is represented by a pictogram as in Fig. 1. Regressions are only shown when significant.
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For birds, three categorical independent variables were added to the GLMM: the lipid extraction treatment, the environment of the bird (three categories: terrestrial, marine and freshwater), and the type of tissue (five categories: blood, feather, liver, muscle and plasma). The carbon discrimination factor was positively correlated with the diet carbon isotopic ratio, and there were significant differences among tissues (Table 1). The nitrogen discrimination factor showed differences among tissues, environments and lipid extraction treatments, but there was no significant relationship with the diet nitrogen isotopic ratio (Table 1). The GLM showed no significant relationships between the carbon discrimination factor and carbon diet isotopic ratio (Δ13C: F1,53 = 0·64, P = 0·425, R2 = 0·012; Fig. 3c); within tissues the GLM between Δ13C and the carbon diet isotopic value was only significant negative for blood (F1,14 = 8·92, P = 0·010, R2 = 0·39). For nitrogen the GLM showed no significant relationships between the discrimination factor and diet isotopic ratio (Δ15N: F1,46 = 0·009, P = 0·924, R2 = 0·00; Fig. 3d). Within tissues and lipid extraction categories, the GLMs between Δ15N and nitrogen diet isotopic value were not significant, and for environment, the relationships were only significant for marine and terrestrial environments (F1,16 = 18·91, P < 0·001, R2 = 0·54 and F1,15 = 18·88, P < 0·001, R2 = 0·56, respectively).
For fishes, three categorical independent variables were added to the GLMM: the lipid extraction treatment, the type of tissue (three different tissues: liver, muscle or whole body) and the environment of the fish (two categories: marine and freshwater). The GLMM for the carbon and nitrogen discrimination factors showed negative relationships with the diet isotopic ratios, and there were also differences among tissues (Table 1). The GLM analysis showed significant relationships between both discrimination factors and the corresponding diet isotopic ratios (Δ13C: F1,39 = 10·69, P = 0·002, R2 = 0·22; and Δ15N: F1,45 = 19·28, P < 0·001, R2 = 0·30; Fig. 3e,f). For the carbon and nitrogen discrimination factors and within tissues, the GLM between discrimination factors and diet isotopic values were only significant for whole body and muscle (carbon: F1,14 = 8·34, P = 0·012, R2 = 0·37, and F1,16 = 7·01, P = 0·018, R2 = 0·30, respectively; nitrogen: F1,15 = 16·65, P < 0·001, R2 = 0·53, and F1,17 = 7·85, P = 0·012, R2 = 0·32, respectively).
For invertebrates, one categorical variable was added to the GLMM model that was not significant in the full model: the environment of the invertebrate (three categories: terrestrial, marine and freshwater). The discrimination factors for nitrogen and carbon were negatively correlated with their corresponding diet isotopic ratios (Table 1). The same trend was evident in the GLM analysis (Fig. 3g,h), which was significant for carbon (F1,84 = 7·97, P = 0·006, R2 = 0·09) and nitrogen (F1,71 = 39·50, P < 0·001, R2 = 0·36) .