- Email: [email protected]
Direct and indirect effects of an invasive omnivore crayfish on leaf litter decomposition
Science of the Total Environment 541 (2016) 714–720
Contents lists available at ScienceDirect
Science of the Total Environment journal homepage: www.elsevier.com/locate/scitotenv
Direct and indirect effects of an invasive omnivore crayfish on leaf litter decomposition Francisco Carvalho a,b,⁎, Cláudia Pascoal a,b, Fernanda Cássio a,b, Ronaldo Sousa a,b,c a b c
CBMA — Centre of Molecular and Environmental Biology, Department of Biology, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal IB-S, Institute of Science and Innovation for Bio-Sustainability, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal Interdisciplinary Centre of Marine and Environmental Research (CIIMAR/CIMAR), University of Porto, Rua dos Bragas 289, P 4050-123 Porto, Portugal
H I G H L I G H T S
G R A P H I C A L
A B S T R A C T
• Procambarus clarkii is one of the most problematic IAS in European freshwaters. • We assessed the direct and indirect effects of P. clarkii on leaf decomposition. • P. clarkii affected basal resources by direct consumption of leaf litter. • P. clarkii indirectly affected leaf decomposition through invertebrate consumption. • Effects of P. clarkii on other invertebrates may be time dependent.
a r t i c l e
i n f o
Article history: Received 2 September 2015 Received in revised form 24 September 2015 Accepted 24 September 2015 Available online xxxx Editor: D. Barcelo Keywords: Detrital food webs Invasive alien species Litter decomposition Procambarus clarkii Streams
a b s t r a c t Invasive alien species (IAS) can disrupt important ecological functions in aquatic ecosystems; however, many of these effects are not quantified and remain speculative. In this study, we assessed the effects of the invasive crayfish Procambarus clarkii (Girard, 1852) on leaf litter decomposition (a key ecosystem process) and associated invertebrates using laboratory and field manipulative experiments. The crayfish had significant impacts on leaf decomposition due to direct consumption of leaf litter and production of fine particulate organic matter, and indirectly due to consumption of invertebrate shredders. The invertebrate community did not appear to recognize P. clarkii as a predator, at least in the first stages after its introduction in the system; but this situation might change with time. Overall, results suggested that the omnivore invader P. clarkii has the potential to affect detritus-based food webs through consumption of basal resources (leaf litter) and/or consumers. Recognizing that this IAS is widespread in Europe, Asia and Africa, and may attain high density and biomass in aquatic ecosystems, our results are important to develop strategies for improving stream ecosystem functioning and to support management actions aiming to control the invasive omnivore P. clarkii. © 2015 Elsevier B.V. All rights reserved.
⁎ Corresponding author at: CBMA — Centre of Molecular and Environmental Biology, Department of Biology, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal. E-mail address: [email protected] (F. Carvalho).
http://dx.doi.org/10.1016/j.scitotenv.2015.09.125 0048-9697/© 2015 Elsevier B.V. All rights reserved.
F. Carvalho et al. / Science of the Total Environment 541 (2016) 714–720
1. Introduction In freshwaters, particularly in forest streams, allochthonous organic matter from riparian zones is the major source of energy and carbon to aquatic biota (Wallace et al., 1997; Suberkropp, 1998). The decomposition of plant litter is a fundamental process conducted by microbial communities, such as fungi and bacteria, as well as by invertebrate shredders (Gessner et al., 1999; Pascoal et al., 2005). The decomposition of plant litter results in the formation of carbon dioxide, mineral compounds, dissolved organic matter (DOM), and fine particulate organic matter (FPOM) that will be used by other organisms (Gessner et al., 1999). The role that some species, such as omnivores, can play in top-down and bottom-up control of plant litter decomposition is a key question to investigate (Greig & McIntosh, 2006). The biotic interactions between different groups of decomposers plus interactions between organisms within the same group may also have an important role in plant-litter decomposition (Gessner et al., 2007). Because omnivores are able to feed on more than one trophic level, they represent a deviation on trophic-cascade theory that may also lead to large effects on community structure (Diehl, 1993; Polis & Strong, 1996). Omnivores' broad diets allow them simultaneously to fill the trophic position of primary consumers up to top predators (Dorn & Wojdak, 2004). Nevertheless, much debate exists about how the dominance of omnivory changes the energy flow through a food web (Yodzis, 1984; Polis & Strong, 1996; McCann et al., 1998; Thompson et al., 2007). The relationship between biodiversity and ecosystem functioning (BEF) has been one of the most exciting research fields in ecology over the past two decades (Hooper et al., 2005; Cardinale et al., 2012; Tilman et al., 2014). Most studies dealing with BEF, including those using plant litter decomposition in freshwaters, have focused on what happens when species are lost (Pascoal et al., 2010; Geraldes et al., 2012; Fernandes et al., 2013, 2015). However, this can be just part of the problem because at the local scale an increase in species number may also occur due, for example, to the introduction of invasive alien species (IAS) (Sousa et al., 2011). Indeed, many ecosystems show unprecedented rates of species introductions mediated by human activities, and freshwaters are not an exception (Strayer, 2010). IAS modify the structure and functioning of ecosystems because they change abiotic conditions (light availability, nutrient levels, heat transfer, habitat complexity and physical disturbance) and biotic interactions, and thus affect several attributes of native communities, such as diversity, spatial distribution, density and biomass (Grosholz, 2002; Byrnes et al., 2007; Simberloff et al., 2013; Gutiérrez et al., 2014). In addition, IAS impacts depend on the time after invasion, their position in the trophic chain and also on the characteristics of the invaded ecosystem (Strayer, 2012). The red-swamp crayfish Procambarus clarkii (Girard, 1852), from the family Cambaridae, is one well-known IAS in freshwater ecosystems. It is native to the center and south of the United States of America and the northeast of Mexico. This species has been a matter of concern in several countries, including Portugal, due to their ecosystem engineering activities and disruption of several biotic interactions, which ultimately cause several impacts on ecosystem functions and services (Gherardi & Holdich, 1999; Rodriguez et al., 2005). This crayfish species is omnivorous, highly active, and it is well known for playing a key role in the food web (Holdich, 2002). P. clarkii is listed in Europe as one of the 100 worst invasive species (DAISIE database, http://www.europe-aliens.org) with some authors considering this crayfish as one of the ten most problematic IAS (Tablado et al., 2010). In Portugal, P. clarkii is widespread from the north to south and west to east colonizing almost all inland aquatic ecosystems (Sousa et al., 2013). Reductions in invertebrates due to crayfish consumption may have cascade effects on lower trophic levels. This top-down effect can be important not only because of direct consumption but also due to possible
715
indirect non-consumptive interactions, where fierce predators change the behavior of the prey (Lima & Dill, 1990). In this way, prey may consider the risk of being predated as an activity with costs and respond according to that risk (Brown et al., 1999). An interesting hypothesis that has been raised is that the prey behavior and the perception of risk may change if the predator is a native or a non-native species (Sih et al., 2010). A naivety effect has been found in native prey in response to introduced predators, with great declines in density and biomass being reported in distinct organisms (e.g. fishes: McLean et al., 2007; Kuehne & Olden, 2012; invertebrates: Freeman & Byers, 2006; Edgell & Neufeld, 2008). Finally, crayfish may also compete with invertebrates for direct consumption of plant litter due to its omnivore behavior and also compete with other predators for invertebrate prey (Gherardi, 2006). Although P. clarkii is distributed almost worldwide, there is a lack of studies addressing the effects of this IAS on detritus-based food webs and the mechanisms underlying such effects. Given this gap we carried out laboratory and field experiments to test the following hypotheses: i) P. clarkii has a top-down control on plant litter decomposition in forest streams, directly by consuming leaf litter and/or indirectly by consuming invertebrate shredders; and ii) invertebrate shredders change their feeding behavior in the presence of P. clarkii due to the risk of being predated, but the response depends on their naivety (i.e. recognizing or not the crayfish as a potential predator). 2. Materials and methods 2.1. Laboratory experiments We collected males of P. clarkii, with approximately 8 cm of total length (from the rostrum tip to the telson rear edge), in the Minho River (Portugal) near the village of Vila Nova de Cerveira (41°57'N, 8°44'W). We also collected a common invertebrate shredder in streams of North Portugal, Sericostoma sp. larvae (Trichoptera, Sericostomatidae) with approximately 1 cm of total length, in the upper reach of the Cávado River. This site is located 10 km downstream of the town Montalegre, Portugal (41°48'N, 7°51'W) and there are no records of P. clarkii, or any other crayfish species, in this river stretch. Animals of both species were kept under starvation for 24 h before the beginning of the experiments. In a first mesocosm experiment, we assessed the effects of P. clarkii on: i) the consumption of leaf litter in the absence or presence of Sericostoma sp., and ii) the abundance of Sericostoma sp. To this end, we manipulated the presence/absence of the crayfish and the abundance of the invertebrate shredder as follows: control with no Sericostoma sp. and no crayfish; low abundance of Sericostoma sp. (6 individuals) with or without 1 crayfish; high abundance of Sericostoma sp. (12 individuals) with or without 1 crayfish; and 1 crayfish with no Sericostoma sp. Each treatment was replicated 4 times and the experiment ran for 21 days (N = 24) under controlled temperature (15 °C) and photoperiod (12 h in the dark and 12 h with light). For each treatment, aquariums (40 × 23 × 25 cm) were filled with river gravel and pebbles (size 850 μm–60 mm) previously washed and autoclaved (120 °C, 20 min). Aquariums were filled with 3 L of water and equipped with an aeration system. Sets of four grams of alder (Alnus glutinosa Gaertn.) leaves collected in the autumn were weighted, placed in mesh bags and submerged in deionized water for 36 h to promote the leaching of soluble compounds. After that, the leaves were removed from the mesh bags and placed in the aquariums. To ensure the presence of natural microbial communities, 10 disks of alder leaves (12 mm diameter) were previously colonized for one week in a loworder stream and then placed in the aquariums at the beginning of the experiment (following Fernandes et al., 2015). One third of the water volume of each aquarium was renewed every 7 days. The retrieved water was filtered through a 53 μm sieve to collect FPOM. Then, FPOM from each replicate was centrifuged (10 min, 14,000 rpm; Sigma 4–
716
F. Carvalho et al. / Science of the Total Environment 541 (2016) 714–720
16 K), and the pellet lyophilized (Biolblock Scientific-Christ Alpha 2–4 LD Plus) for 48 h, before being weighed to the nearest 0.01 mg. At the end of the experiment, the remaining leaf material was washed, dried at 60 °C for 48 h, and weighed to the nearest 0.01 g. Leaf mass loss was quantified by subtracting the final weight from the initial weight of leaves. In a second mesocosm experiment, we assessed the Sericostoma sp. avoidance behavior to the presence of the predator P. clarkii. To this end, we carried out a three-treatment assay as follows: control (6 Sericostoma sp.); 6 Sericostoma sp. + crayfish water; 6 Sericostoma sp. + 1 crayfish. Sericostoma sp. was able to avoid the predator because the mesocosms were divided transversally by a coarse mesh that separated Sericostoma sp. from the crayfish and the leaves. The mesh size allowed the free movement of Sericostoma sp. but prevented the passage of the crayfish. Therefore, Sericostoma sp. took the risk of being predated when trying to feed on leaves. To assess if the chemical cues putatively released by P. clarkii inhibited the feeding behavior of Sericostoma sp., 10 crayfishes were placed in an aquarium with 15 L of water for 48 h before the beginning of the experiment. In the mesocosm experiment, Sericostoma sp. were placed in aquariums containing water and no crayfish, water that was previously in contact with the crayfish, or water and the crayfish. The mesocosm experiment ran in 3 L aquariums for 21 days, under controlled temperature and photoperiod as above (4 replicates; N = 12). Water was renewed and used for quantification of FPOM production as described above. At the end of the experiment, leaf mass loss was quantified following the procedure described above. 2.2. Field experiment A field manipulative experiment was carried out in the Campos Stream (41°58'N, 8°41'W; Vila Nova de Cerveira, North Portugal). This stream is a tributary of the Minho River with a total basin area of nearly 13 km2, 7.2 km of total length and a maximum altitude of 278.7 m. We selected two sites in a stretch of approximately 200 m where a waterfall divided the upstream from the downstream site. This waterfall acts as a physical barrier to upstream dispersal of the crayfish P. clarkii since there are no records of this IAS upstream of the waterfall. The fish fauna downstream of the waterfall includes the non-native Iberian gudgeon (Gobio lozanoi) as well as native species, such as European eel (Anguilla anguilla), brown trout (Salmo trutta), ruivaco (Achondrostoma arcasii), three-spine stickleback (Gasterosteus gymnurus) and Iberian loach (Cobitis paludica). Above the waterfall, the fish fauna includes brown trout, European eel and ruivaco. The chosen stream stretch is characterized by having typical riparian vegetation of North Portugal dominated by alder and oak (Quercus robur L.) trees. Allochthonous plant litter seems to be the main food resource for stream biota although some submerged vegetation is present. The stream bottom at both sites is similar and mainly constituted by sand, gravel and cobbles. A four-treatment (four replicates for each) experiment was designed to control the presence/absence of P. clarkii and prevent or allow access by invertebrates. To that end, baskets (38 × 29 × 21.5 cm) were filled with four g of alder leaves, two pebbles and gravel; half the baskets were covered by a fine mesh (500 μm mesh size) to prevent the access of invertebrates and the other half by coarse mesh (5 mm mesh size) to allow the entry of invertebrates. The four-treatments in the baskets were: coarse mesh without crayfish; coarse mesh + 1 crayfish; fine mesh without crayfish (control); fine mesh + 1 crayfish. Both mesh sizes excluded the entry of other predators (e.g. fishes) and larger invertebrates (e.g. crayfishes). The experiment was run in parallel at the upstream and downstream sites for 21 days at the end of the summer in 2013. At the end of the experiment, all baskets were transported to the laboratory in separate bags inside cool boxes. Baskets were washed and the invertebrates were separated from the leaves using a battery of sieves from 60 mm to 850 μm. The remaining leaves from each replicate
were washed and dried at 60 °C for 48 h before being weighed to the nearest 0.01 g. Invertebrates were preserved in ethanol (96%, v/v) and then they were counted and identified to the lowest possible taxonomic level following Tachet et al. (2010). For biomass quantification, invertebrates were dried (80 °C) for 48 h and weighed to the nearest 0.01 mg. Physical and chemical stream water parameters were measured at both upstream and downstream sites at the beginning of the experiment. Temperature, pH, conductivity and dissolved oxygen were measured in situ with field probes (Multiline F/set 3 no. 400327, WTW, Weilheim, Germany) and depth was measured with a tape measure. Additionally, stream water samples were collected with sterile dark glass bottles, transported in a cool box (4 °C) to the laboratory to determine the concentrations of ammonium (HACH kit, program 385), nitrate (HACH kit, program 351) and phosphate (HACH kit, program 490) using a HACH DR/2000 photometer (HACH, Loveland, CO). 2.3. Statistical analysis In the laboratory experiment, two-way analysis of variance (ANOVA) was used (Zar, 2009) to test if the presence of crayfish and the abundance of invertebrate shredders affected leaf mass loss and FPOM production. In addition, one-way ANOVA was used (Zar, 2009) to test if the direct or indirect (chemical cues) presence of the crayfish affected leaf mass loss and FPOM production. In the field experiment, a three-way ANOVA was used (Zar, 2009) to test if the presence of invertebrates, the presence of crayfish and the stream site affected leaf mass loss. Two-way ANOVAs were used to test if the presence of crayfish and the stream site affected invertebrate abundance, biomass and Margalef richness index. All ANOVAs were preceded by the Shapiro–Wilk test to check if data had a Gaussian distribution and the Bartlett test to test for the homogeneity of variance (Zar, 2009). All ANOVAs were followed by Tukey's post-tests to search for significant differences between treatments (Zar, 2009). A non-metric Multi-Dimensional Scaling (nMDS) was performed to examine the structure of invertebrate assemblages in the field experiment. In the nMDS ordination, samples are represented as points in a low-dimensional space and the relative distances between points are in the same rank order as the relative dissimilarities of the samples as measured by an appropriate resemblance matrix. The Bray-Curtis index was used to assess the similarity between invertebrate assemblages. Overlay clusters representing a resemblance level of 45% were superimposed to the nMDS diagram. A PERMANOVA test (Anderson, 2001) was performed to test if invertebrate assemblages varied with the stream site (two levels: upstream and downstream) and the presence of crayfish (two levels: presence and absence). In all PERMANOVA tests, the statistical significance of variance (α = 0.05) was tested using 9999 permutations of residuals within a reduced model. When the number of permutations was lower than 150, the Monte Carlo p-value was considered. Analyses of variance were done with STATISTICA 8 (StatSoft, USA). Multivariate analysis and PERMANOVA was done with PRIMER 6 (Primer-E, UK). 3. Results 3.1. Laboratory experiments When testing the effects of P. clarkii on leaf consumption and FPOM production in the presence of the invertebrate shredder Sericostoma sp., all shredders were eaten by the crayfish within a few days. In the laboratory experiment, leaf mass loss varied between 31% in mesocosms without crayfish or Sericostoma sp. and 72% in mesocosms containing crayfish and high Sericostoma sp. abundance (Fig. 1A). The
F. Carvalho et al. / Science of the Total Environment 541 (2016) 714–720
Fig. 1. Percentage of leaf mass loss (A) and FPOM produced during leaf decomposition (B) in the absence (crayfishless) or presence (crayfish) of P. clarkii in mesocosms with different abundance of the invertebrate shredder Sericostoma sp. (no Sericostoma sp.: 0 individuals; low abundance: 6 individuals; high abundance: 12 individuals). FPOM production was not measured in mesocosms without Sericostoma sp. and crayfish. Mean ± SEM, n = 4. Different letters indicate significant differences between treatments (P b 0.05).
717
Fig. 2. Percentage of leaf mass loss (A) and FPOM produced during leaf decomposition (B) in mesocosms with 6 larvae of the invertebrate shredder Sericostoma sp. in the absence of P. clarkii (crayfishless), with indirect presence of P. clarkii (crayfish water) or direct presence of P. clarkii (crayfish). Mean ± SEM, n = 4. Different letters indicate significant differences between treatments (P b 0.05).
3.2. Field experiment: in situ validation presence of crayfish significantly increased leaf mass loss (two-way ANOVA, F = 136.3, P b 0.0001; Fig. 1A). In the presence of the crayfish, Sericostoma sp. abundance did not significantly affect leaf decomposition (Tukey's tests, P N 0.05). In the absence of crayfish, leaf mass loss was higher in the presence than in the absence of Sericostoma sp. (Tukey's tests, P b 0.05), but no differences in leaf mass loss were found between low and high Sericostoma sp. abundance (Tukey's tests, P N 0.05). Mean FPOM production varied between 0.1 g in treatments without crayfish and low shredder abundance and 0.7 g in treatments with crayfish and high Sericostoma sp. abundance (Fig. 1B). FPOM production was significantly higher in the presence of the crayfish (twoway ANOVA, F = 87.1, P b 0.0001). No significant differences in FPOM production were detected between mesocosms with different Sericostoma sp. abundance (two-way ANOVA, F = 0.3, P N 0.05; Fig. 1B). In the experiment for assessing the Sericostoma sp. avoidance behavior to the presence of P. clarkii, leaf mass loss varied between 43.2% in the absence of crayfish and 85.1% in the direct presence of the crayfish (Fig. 2A). Leaf mass loss was significantly higher in mesocosms with P. clarkii (one-way ANOVA, F = 33.1, P b 0.0001). No significant differences were found in leaf consumption by the shredder between mesocosms without P. clarkii or with water previously exposed to the crayfish (Tukey's test, P N 0.05; Fig. 2A). Mean FPOM production varied between 0.2 g in the absence of the crayfish and 1.0 g in the direct presence of the crayfish (Fig. 2B). FPOM production was significantly higher in the presence of crayfish (one-way ANOVA, F = 31.4, P b 0.0001; Fig. 2B). FPOM production did not differ between mesocosms with crayfish water and those without crayfish (Tukey's test, P N 0.05; Fig. 2B).
Physical and chemical stream water parameters were similar at the upstream and downstream sites with the exception of phosphate concentration, which was slightly higher upstream (Table 1). Leaf mass loss was significantly affected by the stream site and the crayfish presence, but not by the presence of invertebrates (three-way ANOVA, F = 4.3, P b 0.05; F = 18.6, P b 0.001 and F = 2.9, P N 0.05, respectively; Fig. 3). At the upstream site, leaf mass loss was lower in the control, intermediate in the treatment with invertebrates only and higher in treatments with the crayfish with or without invertebrates (Tukey's tests, P b 0.05; Fig. 3). At the downstream site, leaf mass loss was lower in the control, intermediate in treatments with invertebrates or crayfish, and higher in the treatment with invertebrates and crayfish (Tukey's tests, P b 0.05; Fig. 3).
Table 1 Minimum and maximum stream water parameters measured at the sampling sites. Parameter
Upstream site
Downstream site
Depth (m) Temperature (°C) pH Conductivity (μS/cm) Dissolved O2 (mg/L) (mg/L) P–PO3− 4 N–NH+ 4 (mg/L) N–NO− 3 (mg/L)
0.30–0.50 15.9–16.6 5.86–6.27 74–78.3 7.98–9.21 0.27–0.28 0.01–0.03 0.07–0.09
0.30–0.50 15.9–16.8 6.16–6.29 77–80 8.14–8.96 0.15–0.16 0.01–0.02 0.06–0.09
718
F. Carvalho et al. / Science of the Total Environment 541 (2016) 714–720
Fig. 3. Leaf mass loss at upstream and downstream sites in the absence or presence of invertebrates and/or P. clarkii. Control: absence of invertebrates and P. clarkii. Mean ± SEM, n = 4. Different letters indicate significant differences between treatments (P b 0.05).
Invertebrate abundance varied between 30 individuals at the upstream site in the presence of crayfish and 167 individuals at the downstream site in the absence of crayfish (Fig. 4A). Invertebrate abundance differed between stream sites and also depended on the presence of the crayfish, with higher values in the absence of the crayfish and at the downstream site (two-way ANOVA, F = 5.1 and F = 8.0, respectively, P b 0.05).
Invertebrate biomass varied between 0.02 g DW at the downstream site in the presence of crayfish and 0.07 g DW downstream in the absence of crayfish (Fig. 4B). Invertebrate biomass was affected by the presence of crayfish and stream site (two-way ANOVA, F = 8.0 and F = 5.1, respectively, P b 0.05 for both factors). At the downstream site, invertebrate biomass was significantly higher in the absence of the crayfish (Tukey's test, P b 0.05). At the upstream site, invertebrate biomass was not affected by the presence of P. clarkii (Tukey's test, P N 0.05). Invertebrate richness measured by the Margalef richness index varied between 2.0 at the upstream site in the presence of crayfish and 3.5 at the downstream site in the absence of crayfish (Fig. 4C). The presence of crayfish and the stream site significantly affected the invertebrate richness (two-way ANOVA, F = 8.3 and F = 6.4, respectively, P b 0.05; Fig. 4C). At the downstream site, the Margalef index was significantly higher in the absence of the crayfish (Tukey's test, P b 0.05) but no differences were detected at the upstream site (Tukey's test, P N 0.05). The nMDS analysis based on the abundance of invertebrates (Fig. 5) discriminated four groups: 1) invertebrate group in the absence of crayfish at the downstream site (DCL2, DCL3, DCL4); 2) invertebrate group in the presence or absence of crayfish at the upstream site (UC1, UC3, UC4, UCL1, UCL3); 3) invertebrate group with four samples from downstream and three samples from upstream in the presence or absence of crayfish (DCL1, DC1, DC2, DC3, UCL2, UCL4, UC2); and 4) invertebrate group with one sample from downstream with crayfish (DC4). The structure of the invertebrate community differed between the upstream and downstream sites (PERMANOVA, Pseudo-F = 1.7, P b 0.05). The presence of crayfish did not affect the structure of the invertebrate community and no interaction between stream site and crayfish presence was found (PERMANOVA, Pseudo-F = 1.4, P N 0.05).
Fig. 4. Invertebrate abundance (A) biomass (B) and Margalef richness index (C) at upstream and downstream sites in the absence or presence of P. clarkii. Mean ± SEM, n = 4. Different letters indicate significant differences between treatments at each site (P b 0.05).
F. Carvalho et al. / Science of the Total Environment 541 (2016) 714–720
Fig. 5. Nonmetric Multi-Dimensional Scaling (nMDS) ordination based on leaf-associated invertebrate community at upstream and downstream sites in the absence or presence of invertebrates and/or P. clarkii. Overlaid clusters correspond to 45% of similarity. (UC (up triangle/filled): Upstream-Crayfish; UCL (up triangle/non-filled): Upstream-Crayfishless; DC (inverted triangle/filled): Downstream-Crayfish; DCL (inverted triangle/non-filled): Downstream-Crayfishless).
4. Discussion Our results suggest that P. clarkii has the potential to affect basal resources in invaded freshwater ecosystems by direct consumption of leaf litter and indirectly by decreasing the abundance of other invertebrates. The direct effects of crayfish on leaf litter decomposition were well established in our laboratory experiments: higher leaf decomposition was observed in the presence of crayfish than of the invertebrate shredder Sericostoma sp., regardless of the abundance of the shredders. This may be explained by the body mass of the crayfish that was much higher than that of Sericostoma sp. or by higher energetic metabolic requirements of P. clarkii in comparison to Sericostoma sp. It is important to point out that the crayfish promptly ate all invertebrate shredders in the first days of laboratory experiments indicating that in the treatments with the crayfish and Sericostoma sp. leaf decomposition was mainly mediated by the crayfish. P. clarkii is well known for its omnivory and shows potentially high predation rates on aquatic invertebrates, mainly insects, crustaceans and gastropods (Momot, 1995; Gherardi, 2006). Therefore, it was expected that P. clarkii would consume first the more energetic resources (shredders) and consumed leaf litter when other resources were unavailable. Interestingly, our results seem to differ from those obtained by Lagrue et al. (2014) with other two crayfish species: Astacus astacus (Linnaeus, 1758) and Pacifastacus leniusculus (Dana, 1852). In that study, both crayfish species did not affect leaf decomposition by direct consumption of basal resources. This discrepancy may happen because i) crayfish diet and trophic niche width vary with species (Usio et al., 2006; Olsson et al., 2009; Jackson et al., 2014), and P. clarkii probably shows high appetency for leaf detritus, and/or ii) the duration of laboratory experiment was longer in our study (21 days vs 6 days), and so the crayfish had the need to feed on other resources after consuming the invertebrates. The production of FPOM followed the pattern found for leaf decomposition described above, with P. clarkii clearly contributing to an increase in the production of FPOM available to other trophic levels. Our results seemed to differ from those of Greig & McIntosh (2006) who showed that an invasive fish (S. trutta) in New Zealand lowered the density of invertebrates and, consequently, the rates of leaf decomposition and FPOM production in mesocosms. In our study, because all Sericostoma sp. were promptly eaten by the crayfish, FPOM production was mainly due to the crayfish activity. Therefore, even if the crayfish eliminates all invertebrate shredders from a particular system, the production of FPOM will not be compromised due to the decomposer
719
behavior of P. clarkii. Nevertheless, if leaf decomposition is driven only by few species, the resilience of the ecosystem may be affected (Peterson et al., 1998). In the same vein, the quality of FPOM produced by P. clarkii may differ from that produced by invertebrate shredders, ultimately affecting other invertebrate feeding groups, namely collector gatherers and collector filterers (Montemarano et al., 2007). However, this was not assessed in our study and further investigation is needed to clarify this hypothesis. To better understand the mechanisms underlying the effects of crayfish on invertebrate shredders, we compared the feeding behavior of Sericostoma sp. in the presence of the predator or waterborne cues from P. clarkii in a laboratory experiment. Our results showed that Sericostoma sp. did not change its shredding behavior on leaf decomposition in mesocosms with water previously exposed to P. clarkii. In the same vein, when shredders had the possibility to avoid direct predation they did not, because after a few moments in the mesocosms, Sericostoma sp. crossed the mesh net to feed on the leaves. This may happen for two reasons: i) the shredders were under starvation and took the risk of being predated, and/or ii) Sericostoma sp. did not recognize P. clarkii as a predator. The last hypothesis is plausible because the shredders used in the experiment were collected from a stream where P. clarkii (or any other crayfish species) was not present. This naivety behavior against invasive predators was already described for many other organisms (e.g. Edgell & Neufeld, 2008; Kuehne & Olden, 2012). In our field experiment, leaf decomposition was higher at the downstream than the upstream stream site in which the presence of P. clarkii was not reported yet. The results obtained at the upstream site were similar to those obtained in the laboratory experiments: decomposition of alder leaves was slightly higher in the presence of crayfish than in the presence of other invertebrates. This can be explained by the size and/or metabolic needs of the crayfish and also because other invertebrates had the possibility to feed inside and outside the baskets, while the crayfish only had the possibility to feed on the leaves enclosed in the baskets. Furthermore, the lack of significant differences between treatments with crayfish in the presence or absence of invertebrates indicates that leaf decomposition was mainly mediated by the crayfish at the upstream site. Moreover, the results obtained in the field support the idea that in streams where diversity and biomass of invertebrate shredders are low, as found at the upstream site, P. clarkii plays an important role in leaf decomposition as shown for other crayfish species (Usio, 2000). At the downstream site (where P. clarkii was already reported), the pattern of leaf decomposition was different from the upstream site: leaf mass loss by the crayfish and invertebrates alone did not differ, and was highest when crayfish and invertebrates were together. Since P. clarkii was already introduced at the downstream site, we cannot discard the hypothesis that invertebrates are able to adjust their behavior after a given time in the presence of P. clarkii. Indeed, at the downstream site the crayfish was well-established and the invertebrate community already had the contact with this species for many years (P. clarkii was introduced in the Minho River basin at least at the beginning of the 1990s; Sousa et al. 2013). Consistently, overall abundance and biomass of invertebrates were higher downstream. This supports the idea that invertebrates may have had enough time to identify the crayfish as a predator, and to adapt their feeding behavior to avoid being predated. In temperate streams, some crayfish species (e.g., Paranephrops zealandicus) affect not only leaf decomposition but also the pattern of colonization by invertebrates (Usio, 2000). Our results in the field seemed to support these observations since the invertebrate community at the downstream site showed higher invertebrate diversity, abundance and biomass. Based on results from the upstream site, we hypothesize that the crayfish may also have an indirect effect on leaf litter due to consumption of native invertebrates. Similar results were described for the crayfish P. leniusculus in Californian (USA) streams (Moore et al., 2012) and other predators in freshwater ecosystems (e.g. Wyman, 1998; Konishi et al., 2001; Mancinelli et al., 2002; Greig
720
F. Carvalho et al. / Science of the Total Environment 541 (2016) 714–720
& McIntosh, 2006), in which predators have an indirect effect on detrital food webs by suppressing the efficiency of leaf litter decomposition. However, our results from the downstream site suggest that indirect effects on leaf decomposition through the consumption of native invertebrates may change through time and invertebrates seem to learn how to avoid the crayfish as a predator. Indeed, in the last years, numerous studies showed that prey may rapidly adapt to the presence of invasive predators evolving anti-predator strategies (e.g. Cox, 2004; Freeman & Byers, 2006). In conclusion, by using a combination of laboratory and field manipulative experiments, we clearly showed that P. clarkii affect detrital food webs by direct consumption of leaf litter and/or by affecting other invertebrate species. Since we are dealing with one of the most widespread IAS that may reach high densities and biomass in aquatic ecosystems, our results are important to develop strategies for improving stream ecosystem functioning and to support management actions aiming to control the invasive omnivore P. clarkii. Acknowledgments The European Regional Development Fund — Operational Competitiveness Program (FEDER-POFC-COMPETE) and the Portuguese Foundation for Science and Technology (FCT) supported this study (PEst-C/ BIA/UI4050/2014, PTDC/AAC-AMB/117068/2010). References Anderson, M.J., 2001. A new method for non-parametric multivariate analysis of variance. Austral Ecology 26, 32–46. Brown, J.S., Laundré, J.W., Gurung, M., 1999. The ecology of fear: optimal foraging, game theory, and trophic interactions. J. Mammal. 80, 385–399. Byrnes, J.E., Reynolds, P.L., Stachowicz, J.J., 2007. Invasions and extinctions reshape coastal marine food webs. PLoS One 3, 295. Cardinale, B.J., Duffy, J.E., Gonzalez, A., Hooper, D.U., Perrings, C., Venail, P., et al., 2012. Biodiversity loss and its impact on humanity. Nature 486, 59–67. Cox, G.W., 2004. Alien species and evolution. Island Press, Washington, United States of America. Diehl, S., 1993. Relative consumer sizes and the strengths of direct and indirect interactions in omnivorous feeding relationships. Oikos 68, 151–157. Dorn, N.J., Wojdak, J.M., 2004. The role of omnivorous crayfish in littoral communities. Oecologia 140, 150–159. Edgell, T.C., Neufeld, C.J., 2008. Experimental evidence for latent developmental plasticity: intertidal whelks respond to a native but not an introduced predator. Biol. Lett. 4, 385–387. Fernandes, I., Duarte, S., Cássio, F., Pascoal, C., 2013. Effects of riparian plant diversity loss on aquatic microbial decomposers become more pronounced with increasing time. Microb. Ecol. 66, 763–772. Fernandes, I., Duarte, S., Cássio, F., Pascoal, C., 2015. Plant litter diversity affects invertebrate shredder activity and the quality of fine particulate organic matter in streams. Mar. Freshw. Res. http://dx.doi.org/10.1071/MF14089 Freeman, A.S., Byers, J.E., 2006. Divergent induced responses to an invasive predator in marine mussel populations. Science 313, 831–833. Geraldes, P., Pascoal, C., Cássio, F., 2012. Effects of increased temperature and aquatic fungal diversity loss on litter decomposition. Fungal Ecol. 5, 734–774. Gessner, M., Chauvet, E., Dobson, M., 1999. A perspective on leaf litter breakdown in streams. Oikos 85, 377–384. Gessner, M.O., Gulis, V., Kueh, K., Chauvet, E., Suberkropp, K., 2007. Fungal decomposers of plant litter in aquatic habitats. In: Kubicek, C.P., Druzhinina, I.S. (Eds.), The Mycota, Vol. IV. Environmental and Microbial Relationships. Springer, Berlin, pp. 301–324. Gherardi, F., 2006. Crayfish invading Europe: the case study of Procambarus clarkii. Mar. Freshw. Behav. Physiol. 39, 175–191. Gherardi, F., Holdich, D.M., 1999. Crayfish in Europe as alien species. How to make the best of a bad situation? A. A. Balkema, Rotterdam Greig, H.S., McIntosh, A.R., 2006. Indirect effects of predatory trout on organic matter processing in detritus-based stream food webs. Oikos 112 (1), 31–40. Grosholz, E., 2002. Ecological and evolutionary consequences of coastal invasions. Trends Ecol. Evol. 17, 22–27. Gutiérrez, J.L., Jones, C.G., Sousa, R., 2014. Toward an integrated ecosystem perspective of invasive species impacts. Acta Oecol. 54, 131–138. Holdich, D.M., 2002. In: Holdich, D.M. (Ed.), Background and functional morphology. Biology of Freshwater Crayfish. Blackwell Science, Oxford. Hooper, D.U., Chapin, F.S.I.I.I., Ewel, J.J., Hector, A., Inchausti, P., Lavorel, S., et al., 2005. Effects of biodiversity on ecosystem functioning: a consensus of current knowledge. Ecol. Monogr. 75, 3–35. Jackson, M.C., Jones, T., Milligan, M., Sheath, D., Taylor, J., Ellis, A., et al., 2014. Niche differentiation among invasive crayfish and their impacts on ecosystem structure and functioning. Freshw. Biol. 59, 1123–1135.
Konishi, M., Nakano, S., Iwata, T., 2001. Trophic cascading effects of predatory fish on leaf litter processing in a Japanese stream. Ecol. Res. 16, 415–422. Kuehne, L.M., Olden, J.D., 2012. Prey naivety in the behavioural responses of juvenile Chinook salmon (Oncorhynchus tshawytscha) to an invasive predator. Freshw. Biol. 57, 1126–1137. Lagrue, C., Podgorniak, T., Lecerf, A., Bollache, L., 2014. An invasive species may be better than none: invasive signal and native noble crayfish have similar community effects. Freshw. Biol. 59 (9), 1982–1995. Lima, S.L., Dill, L.M., 1990. Behavioral decisions made under the risk of predation: a review and prospectus. Can. J. Zool. 68, 619–640. Mancinelli, G., Costantini, M.L., Rossi, L., 2002. Cascading effects of predatory fish exclusion on the detritus-based food web of a lake littoral zone (Lake Vico, central Italy). Oecologia 133, 402–411. McCann, K.S., Hastings, A., Strong, D.R., 1998. Trophic cascades and trophic trickles in pelagic food webs. Proc. R. Soc. Lond. Ser. B Biol. Sci. 265, 205–209. McLean, F., Barbee, N., Swearer, S., 2007. Avoidance of native versus non-native predator odours by migrating whitebait and juveniles of the common galaxiid, Galaxias maculatus. N. Z. J. Mar. Freshw. Res. 41, 175–184. Moore, J.W., Carlson, S.M., Twardochleb, L.A., Hwan, J.L., Fox, J.M., Hayes, S.A., 2012. Trophic tangles through time? Opposing direct and indirect effects of an invasive omnivore on stream ecosystem processes. PLoS One 7 (11), e50687. Momot, W.T., 1995. Redefining the role of crayfish in aquatic ecosystems. Rev. Fish. Sci. 3, 33–63. Montemarano, J.J., Kershner, M.W., Leff, L.G., 2007. Crayfish effects on fine particulate organic matter quality and quantity. Fundamental and Applied Limnology/Archiv für Hydrobiologie 169, 223–229. Olsson, K., Stenroth, P., Nyström, P., Granéli, W., 2009. Invasions and niche width of an introduced crayfish differ from a native crayfish? Freshw. Biol. 54, 1731–1740. Pascoal, C., Cássio, F., Marcotegui, A., Sanz, B., Gomes, P., 2005. Role of fungi, bacteria, and invertebrates in leaf litter breakdown in a polluted river. J. N. Am. Benthol. Soc. 24, 784–797. Pascoal, C., Cássio, F., Nikolcheva, L.G., Bärlocher, F., 2010. Realized fungal diversity increases functional stability of leaf-litter decomposition under zinc stress. Microb. Ecol. 59, 84–93. Peterson, G., Allen, C.R., Holling, C.S., 1998. Ecological resilience, biodiversity, and scale. Ecosystems 1, 6–18. Polis, G.A., Strong, D.R., 1996. Food web complexity and community dynamics. Am. Nat. 147, 813–846. Rodriguez, C.F., Becares, E., Fernandez-Alaez, M., Fernandez-Alaez, C., 2005. Loss of diversity and degradation of wetlands as a result of introducing exotic crayfish. Biol. Invasions 7, 75–85. Sih, A., Bolnick, D.I., Luttbeg, B., Orrock, J.L., Peacor, S.D., Pintor, L.M., Preisser, E., Rehage, J.S., Vonesh, J.R., 2010. Predator–prey naïveté, antipredator behavior, and the ecology of predator invasions. Oikos 119, 610–621. Simberloff, D., Martin, J.-L., Genovesi, P., Maris, V., Wardle, D., Aronson, J., et al., 2013. Impacts of biological invasions: what's what and the way forward. Trends Ecol. Evol. 28, 58–66. Sousa, R., Freitas, F.E., Mota, M., Nogueira, A.J., Antunes, C., 2013. Invasive dynamics of the crayfish Procambarus clarkii (Girard, 1852) in the international section of the river Minho (NW of the Iberian Peninsula). Aquat. Conserv. Mar. Freshwat. Ecosyst. 23, 656–666. Sousa, R., Morais, P., Dias, E., Antunes, C., 2011. Biological invasions and ecosystem functioning: time to merge. Biol. Invasions 13, 1055–1058. Strayer, D.L., 2010. Alien species in fresh waters: ecological effects, interactions with other stressors, and prospects for the future. Freshw. Biol. 55, 152–174. Strayer, D.L., 2012. Eight questions about invasions and ecosystem functioning. Ecol. Lett. 15, 1199–1210. Suberkropp, K., 1998. Microorganisms and organic matter decomposition. In: Naiman, R., Bilby, R. (Eds.), River Ecology and Management: Lessons from the Pacific Coastal Ecoregion. Springer, New York, pp. 120–143. Tablado, Z., Tella, J.L., Sánchez-Zapata, J.A., Hiraldo, F., 2010. The paradox of the long-term positive effects of a North American crayfish on a European community of predators. Conserv. Biol. 24, 1229–1238. Tachet, H., Richoux, P., Bournaud, M., Usseglio-Polatera, P., 2010. Invertébrés d'eau douce: Systématique, biologie, écologie. CNRS éditions, Paris, France. Tilman, D., Isbell, F., Cowles, J.M., 2014. Biodiversity and Ecosystem Functioning. Annu. Rev. Ecol. Evol. Syst. 45 (1), 471. Thompson, R.M., Hemberg, M., Starzomski, B.M., Shurin, J.B., 2007. Trophic levels and trophic tangles: the prevalence of omnivory in real food webs. Ecology 88, 612–617. Usio, N., 2000. Effects of crayfish on leaf processing and invertebrate colonisation of leaves in a headwater stream: decoupling of a trophic cascade. Oecologia 124, 608–614. Usio, N., Suzuki, K., Konishi, M., Nakano, S., 2006. Alien vs. endemic crayfish: roles of species identity in ecosystem functioning. Arch. Hydrobiol. 166, 1–21. Wallace, J.B., Eggert, S.L., Meyer, J.L., Webster, J.R., 1997. Multiple trophic levels of a forest stream linked to terrestrial litter inputs. Science 277, 102–104. Wyman, R.L., 1998. Experimental assessment of salamanders as predators of detrital food webs: effects on invertebrates, decomposition and the carbon cycle. Biodivers. Conserv. 7, 641–650. Yodzis, P., 1984. How rare is omnivory? Ecology 65, 321–323. Zar, J.H., 2009. Biostatistical analysis. Fifth edition. Prentice-Hall, Englewood Cliffs, New Jersey, USA.
Contents lists available at ScienceDirect
Science of the Total Environment journal homepage: www.elsevier.com/locate/scitotenv
Direct and indirect effects of an invasive omnivore crayfish on leaf litter decomposition Francisco Carvalho a,b,⁎, Cláudia Pascoal a,b, Fernanda Cássio a,b, Ronaldo Sousa a,b,c a b c
CBMA — Centre of Molecular and Environmental Biology, Department of Biology, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal IB-S, Institute of Science and Innovation for Bio-Sustainability, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal Interdisciplinary Centre of Marine and Environmental Research (CIIMAR/CIMAR), University of Porto, Rua dos Bragas 289, P 4050-123 Porto, Portugal
H I G H L I G H T S
G R A P H I C A L
A B S T R A C T
• Procambarus clarkii is one of the most problematic IAS in European freshwaters. • We assessed the direct and indirect effects of P. clarkii on leaf decomposition. • P. clarkii affected basal resources by direct consumption of leaf litter. • P. clarkii indirectly affected leaf decomposition through invertebrate consumption. • Effects of P. clarkii on other invertebrates may be time dependent.
a r t i c l e
i n f o
Article history: Received 2 September 2015 Received in revised form 24 September 2015 Accepted 24 September 2015 Available online xxxx Editor: D. Barcelo Keywords: Detrital food webs Invasive alien species Litter decomposition Procambarus clarkii Streams
a b s t r a c t Invasive alien species (IAS) can disrupt important ecological functions in aquatic ecosystems; however, many of these effects are not quantified and remain speculative. In this study, we assessed the effects of the invasive crayfish Procambarus clarkii (Girard, 1852) on leaf litter decomposition (a key ecosystem process) and associated invertebrates using laboratory and field manipulative experiments. The crayfish had significant impacts on leaf decomposition due to direct consumption of leaf litter and production of fine particulate organic matter, and indirectly due to consumption of invertebrate shredders. The invertebrate community did not appear to recognize P. clarkii as a predator, at least in the first stages after its introduction in the system; but this situation might change with time. Overall, results suggested that the omnivore invader P. clarkii has the potential to affect detritus-based food webs through consumption of basal resources (leaf litter) and/or consumers. Recognizing that this IAS is widespread in Europe, Asia and Africa, and may attain high density and biomass in aquatic ecosystems, our results are important to develop strategies for improving stream ecosystem functioning and to support management actions aiming to control the invasive omnivore P. clarkii. © 2015 Elsevier B.V. All rights reserved.
⁎ Corresponding author at: CBMA — Centre of Molecular and Environmental Biology, Department of Biology, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal. E-mail address: [email protected] (F. Carvalho).
http://dx.doi.org/10.1016/j.scitotenv.2015.09.125 0048-9697/© 2015 Elsevier B.V. All rights reserved.
F. Carvalho et al. / Science of the Total Environment 541 (2016) 714–720
1. Introduction In freshwaters, particularly in forest streams, allochthonous organic matter from riparian zones is the major source of energy and carbon to aquatic biota (Wallace et al., 1997; Suberkropp, 1998). The decomposition of plant litter is a fundamental process conducted by microbial communities, such as fungi and bacteria, as well as by invertebrate shredders (Gessner et al., 1999; Pascoal et al., 2005). The decomposition of plant litter results in the formation of carbon dioxide, mineral compounds, dissolved organic matter (DOM), and fine particulate organic matter (FPOM) that will be used by other organisms (Gessner et al., 1999). The role that some species, such as omnivores, can play in top-down and bottom-up control of plant litter decomposition is a key question to investigate (Greig & McIntosh, 2006). The biotic interactions between different groups of decomposers plus interactions between organisms within the same group may also have an important role in plant-litter decomposition (Gessner et al., 2007). Because omnivores are able to feed on more than one trophic level, they represent a deviation on trophic-cascade theory that may also lead to large effects on community structure (Diehl, 1993; Polis & Strong, 1996). Omnivores' broad diets allow them simultaneously to fill the trophic position of primary consumers up to top predators (Dorn & Wojdak, 2004). Nevertheless, much debate exists about how the dominance of omnivory changes the energy flow through a food web (Yodzis, 1984; Polis & Strong, 1996; McCann et al., 1998; Thompson et al., 2007). The relationship between biodiversity and ecosystem functioning (BEF) has been one of the most exciting research fields in ecology over the past two decades (Hooper et al., 2005; Cardinale et al., 2012; Tilman et al., 2014). Most studies dealing with BEF, including those using plant litter decomposition in freshwaters, have focused on what happens when species are lost (Pascoal et al., 2010; Geraldes et al., 2012; Fernandes et al., 2013, 2015). However, this can be just part of the problem because at the local scale an increase in species number may also occur due, for example, to the introduction of invasive alien species (IAS) (Sousa et al., 2011). Indeed, many ecosystems show unprecedented rates of species introductions mediated by human activities, and freshwaters are not an exception (Strayer, 2010). IAS modify the structure and functioning of ecosystems because they change abiotic conditions (light availability, nutrient levels, heat transfer, habitat complexity and physical disturbance) and biotic interactions, and thus affect several attributes of native communities, such as diversity, spatial distribution, density and biomass (Grosholz, 2002; Byrnes et al., 2007; Simberloff et al., 2013; Gutiérrez et al., 2014). In addition, IAS impacts depend on the time after invasion, their position in the trophic chain and also on the characteristics of the invaded ecosystem (Strayer, 2012). The red-swamp crayfish Procambarus clarkii (Girard, 1852), from the family Cambaridae, is one well-known IAS in freshwater ecosystems. It is native to the center and south of the United States of America and the northeast of Mexico. This species has been a matter of concern in several countries, including Portugal, due to their ecosystem engineering activities and disruption of several biotic interactions, which ultimately cause several impacts on ecosystem functions and services (Gherardi & Holdich, 1999; Rodriguez et al., 2005). This crayfish species is omnivorous, highly active, and it is well known for playing a key role in the food web (Holdich, 2002). P. clarkii is listed in Europe as one of the 100 worst invasive species (DAISIE database, http://www.europe-aliens.org) with some authors considering this crayfish as one of the ten most problematic IAS (Tablado et al., 2010). In Portugal, P. clarkii is widespread from the north to south and west to east colonizing almost all inland aquatic ecosystems (Sousa et al., 2013). Reductions in invertebrates due to crayfish consumption may have cascade effects on lower trophic levels. This top-down effect can be important not only because of direct consumption but also due to possible
715
indirect non-consumptive interactions, where fierce predators change the behavior of the prey (Lima & Dill, 1990). In this way, prey may consider the risk of being predated as an activity with costs and respond according to that risk (Brown et al., 1999). An interesting hypothesis that has been raised is that the prey behavior and the perception of risk may change if the predator is a native or a non-native species (Sih et al., 2010). A naivety effect has been found in native prey in response to introduced predators, with great declines in density and biomass being reported in distinct organisms (e.g. fishes: McLean et al., 2007; Kuehne & Olden, 2012; invertebrates: Freeman & Byers, 2006; Edgell & Neufeld, 2008). Finally, crayfish may also compete with invertebrates for direct consumption of plant litter due to its omnivore behavior and also compete with other predators for invertebrate prey (Gherardi, 2006). Although P. clarkii is distributed almost worldwide, there is a lack of studies addressing the effects of this IAS on detritus-based food webs and the mechanisms underlying such effects. Given this gap we carried out laboratory and field experiments to test the following hypotheses: i) P. clarkii has a top-down control on plant litter decomposition in forest streams, directly by consuming leaf litter and/or indirectly by consuming invertebrate shredders; and ii) invertebrate shredders change their feeding behavior in the presence of P. clarkii due to the risk of being predated, but the response depends on their naivety (i.e. recognizing or not the crayfish as a potential predator). 2. Materials and methods 2.1. Laboratory experiments We collected males of P. clarkii, with approximately 8 cm of total length (from the rostrum tip to the telson rear edge), in the Minho River (Portugal) near the village of Vila Nova de Cerveira (41°57'N, 8°44'W). We also collected a common invertebrate shredder in streams of North Portugal, Sericostoma sp. larvae (Trichoptera, Sericostomatidae) with approximately 1 cm of total length, in the upper reach of the Cávado River. This site is located 10 km downstream of the town Montalegre, Portugal (41°48'N, 7°51'W) and there are no records of P. clarkii, or any other crayfish species, in this river stretch. Animals of both species were kept under starvation for 24 h before the beginning of the experiments. In a first mesocosm experiment, we assessed the effects of P. clarkii on: i) the consumption of leaf litter in the absence or presence of Sericostoma sp., and ii) the abundance of Sericostoma sp. To this end, we manipulated the presence/absence of the crayfish and the abundance of the invertebrate shredder as follows: control with no Sericostoma sp. and no crayfish; low abundance of Sericostoma sp. (6 individuals) with or without 1 crayfish; high abundance of Sericostoma sp. (12 individuals) with or without 1 crayfish; and 1 crayfish with no Sericostoma sp. Each treatment was replicated 4 times and the experiment ran for 21 days (N = 24) under controlled temperature (15 °C) and photoperiod (12 h in the dark and 12 h with light). For each treatment, aquariums (40 × 23 × 25 cm) were filled with river gravel and pebbles (size 850 μm–60 mm) previously washed and autoclaved (120 °C, 20 min). Aquariums were filled with 3 L of water and equipped with an aeration system. Sets of four grams of alder (Alnus glutinosa Gaertn.) leaves collected in the autumn were weighted, placed in mesh bags and submerged in deionized water for 36 h to promote the leaching of soluble compounds. After that, the leaves were removed from the mesh bags and placed in the aquariums. To ensure the presence of natural microbial communities, 10 disks of alder leaves (12 mm diameter) were previously colonized for one week in a loworder stream and then placed in the aquariums at the beginning of the experiment (following Fernandes et al., 2015). One third of the water volume of each aquarium was renewed every 7 days. The retrieved water was filtered through a 53 μm sieve to collect FPOM. Then, FPOM from each replicate was centrifuged (10 min, 14,000 rpm; Sigma 4–
716
F. Carvalho et al. / Science of the Total Environment 541 (2016) 714–720
16 K), and the pellet lyophilized (Biolblock Scientific-Christ Alpha 2–4 LD Plus) for 48 h, before being weighed to the nearest 0.01 mg. At the end of the experiment, the remaining leaf material was washed, dried at 60 °C for 48 h, and weighed to the nearest 0.01 g. Leaf mass loss was quantified by subtracting the final weight from the initial weight of leaves. In a second mesocosm experiment, we assessed the Sericostoma sp. avoidance behavior to the presence of the predator P. clarkii. To this end, we carried out a three-treatment assay as follows: control (6 Sericostoma sp.); 6 Sericostoma sp. + crayfish water; 6 Sericostoma sp. + 1 crayfish. Sericostoma sp. was able to avoid the predator because the mesocosms were divided transversally by a coarse mesh that separated Sericostoma sp. from the crayfish and the leaves. The mesh size allowed the free movement of Sericostoma sp. but prevented the passage of the crayfish. Therefore, Sericostoma sp. took the risk of being predated when trying to feed on leaves. To assess if the chemical cues putatively released by P. clarkii inhibited the feeding behavior of Sericostoma sp., 10 crayfishes were placed in an aquarium with 15 L of water for 48 h before the beginning of the experiment. In the mesocosm experiment, Sericostoma sp. were placed in aquariums containing water and no crayfish, water that was previously in contact with the crayfish, or water and the crayfish. The mesocosm experiment ran in 3 L aquariums for 21 days, under controlled temperature and photoperiod as above (4 replicates; N = 12). Water was renewed and used for quantification of FPOM production as described above. At the end of the experiment, leaf mass loss was quantified following the procedure described above. 2.2. Field experiment A field manipulative experiment was carried out in the Campos Stream (41°58'N, 8°41'W; Vila Nova de Cerveira, North Portugal). This stream is a tributary of the Minho River with a total basin area of nearly 13 km2, 7.2 km of total length and a maximum altitude of 278.7 m. We selected two sites in a stretch of approximately 200 m where a waterfall divided the upstream from the downstream site. This waterfall acts as a physical barrier to upstream dispersal of the crayfish P. clarkii since there are no records of this IAS upstream of the waterfall. The fish fauna downstream of the waterfall includes the non-native Iberian gudgeon (Gobio lozanoi) as well as native species, such as European eel (Anguilla anguilla), brown trout (Salmo trutta), ruivaco (Achondrostoma arcasii), three-spine stickleback (Gasterosteus gymnurus) and Iberian loach (Cobitis paludica). Above the waterfall, the fish fauna includes brown trout, European eel and ruivaco. The chosen stream stretch is characterized by having typical riparian vegetation of North Portugal dominated by alder and oak (Quercus robur L.) trees. Allochthonous plant litter seems to be the main food resource for stream biota although some submerged vegetation is present. The stream bottom at both sites is similar and mainly constituted by sand, gravel and cobbles. A four-treatment (four replicates for each) experiment was designed to control the presence/absence of P. clarkii and prevent or allow access by invertebrates. To that end, baskets (38 × 29 × 21.5 cm) were filled with four g of alder leaves, two pebbles and gravel; half the baskets were covered by a fine mesh (500 μm mesh size) to prevent the access of invertebrates and the other half by coarse mesh (5 mm mesh size) to allow the entry of invertebrates. The four-treatments in the baskets were: coarse mesh without crayfish; coarse mesh + 1 crayfish; fine mesh without crayfish (control); fine mesh + 1 crayfish. Both mesh sizes excluded the entry of other predators (e.g. fishes) and larger invertebrates (e.g. crayfishes). The experiment was run in parallel at the upstream and downstream sites for 21 days at the end of the summer in 2013. At the end of the experiment, all baskets were transported to the laboratory in separate bags inside cool boxes. Baskets were washed and the invertebrates were separated from the leaves using a battery of sieves from 60 mm to 850 μm. The remaining leaves from each replicate
were washed and dried at 60 °C for 48 h before being weighed to the nearest 0.01 g. Invertebrates were preserved in ethanol (96%, v/v) and then they were counted and identified to the lowest possible taxonomic level following Tachet et al. (2010). For biomass quantification, invertebrates were dried (80 °C) for 48 h and weighed to the nearest 0.01 mg. Physical and chemical stream water parameters were measured at both upstream and downstream sites at the beginning of the experiment. Temperature, pH, conductivity and dissolved oxygen were measured in situ with field probes (Multiline F/set 3 no. 400327, WTW, Weilheim, Germany) and depth was measured with a tape measure. Additionally, stream water samples were collected with sterile dark glass bottles, transported in a cool box (4 °C) to the laboratory to determine the concentrations of ammonium (HACH kit, program 385), nitrate (HACH kit, program 351) and phosphate (HACH kit, program 490) using a HACH DR/2000 photometer (HACH, Loveland, CO). 2.3. Statistical analysis In the laboratory experiment, two-way analysis of variance (ANOVA) was used (Zar, 2009) to test if the presence of crayfish and the abundance of invertebrate shredders affected leaf mass loss and FPOM production. In addition, one-way ANOVA was used (Zar, 2009) to test if the direct or indirect (chemical cues) presence of the crayfish affected leaf mass loss and FPOM production. In the field experiment, a three-way ANOVA was used (Zar, 2009) to test if the presence of invertebrates, the presence of crayfish and the stream site affected leaf mass loss. Two-way ANOVAs were used to test if the presence of crayfish and the stream site affected invertebrate abundance, biomass and Margalef richness index. All ANOVAs were preceded by the Shapiro–Wilk test to check if data had a Gaussian distribution and the Bartlett test to test for the homogeneity of variance (Zar, 2009). All ANOVAs were followed by Tukey's post-tests to search for significant differences between treatments (Zar, 2009). A non-metric Multi-Dimensional Scaling (nMDS) was performed to examine the structure of invertebrate assemblages in the field experiment. In the nMDS ordination, samples are represented as points in a low-dimensional space and the relative distances between points are in the same rank order as the relative dissimilarities of the samples as measured by an appropriate resemblance matrix. The Bray-Curtis index was used to assess the similarity between invertebrate assemblages. Overlay clusters representing a resemblance level of 45% were superimposed to the nMDS diagram. A PERMANOVA test (Anderson, 2001) was performed to test if invertebrate assemblages varied with the stream site (two levels: upstream and downstream) and the presence of crayfish (two levels: presence and absence). In all PERMANOVA tests, the statistical significance of variance (α = 0.05) was tested using 9999 permutations of residuals within a reduced model. When the number of permutations was lower than 150, the Monte Carlo p-value was considered. Analyses of variance were done with STATISTICA 8 (StatSoft, USA). Multivariate analysis and PERMANOVA was done with PRIMER 6 (Primer-E, UK). 3. Results 3.1. Laboratory experiments When testing the effects of P. clarkii on leaf consumption and FPOM production in the presence of the invertebrate shredder Sericostoma sp., all shredders were eaten by the crayfish within a few days. In the laboratory experiment, leaf mass loss varied between 31% in mesocosms without crayfish or Sericostoma sp. and 72% in mesocosms containing crayfish and high Sericostoma sp. abundance (Fig. 1A). The
F. Carvalho et al. / Science of the Total Environment 541 (2016) 714–720
Fig. 1. Percentage of leaf mass loss (A) and FPOM produced during leaf decomposition (B) in the absence (crayfishless) or presence (crayfish) of P. clarkii in mesocosms with different abundance of the invertebrate shredder Sericostoma sp. (no Sericostoma sp.: 0 individuals; low abundance: 6 individuals; high abundance: 12 individuals). FPOM production was not measured in mesocosms without Sericostoma sp. and crayfish. Mean ± SEM, n = 4. Different letters indicate significant differences between treatments (P b 0.05).
717
Fig. 2. Percentage of leaf mass loss (A) and FPOM produced during leaf decomposition (B) in mesocosms with 6 larvae of the invertebrate shredder Sericostoma sp. in the absence of P. clarkii (crayfishless), with indirect presence of P. clarkii (crayfish water) or direct presence of P. clarkii (crayfish). Mean ± SEM, n = 4. Different letters indicate significant differences between treatments (P b 0.05).
3.2. Field experiment: in situ validation presence of crayfish significantly increased leaf mass loss (two-way ANOVA, F = 136.3, P b 0.0001; Fig. 1A). In the presence of the crayfish, Sericostoma sp. abundance did not significantly affect leaf decomposition (Tukey's tests, P N 0.05). In the absence of crayfish, leaf mass loss was higher in the presence than in the absence of Sericostoma sp. (Tukey's tests, P b 0.05), but no differences in leaf mass loss were found between low and high Sericostoma sp. abundance (Tukey's tests, P N 0.05). Mean FPOM production varied between 0.1 g in treatments without crayfish and low shredder abundance and 0.7 g in treatments with crayfish and high Sericostoma sp. abundance (Fig. 1B). FPOM production was significantly higher in the presence of the crayfish (twoway ANOVA, F = 87.1, P b 0.0001). No significant differences in FPOM production were detected between mesocosms with different Sericostoma sp. abundance (two-way ANOVA, F = 0.3, P N 0.05; Fig. 1B). In the experiment for assessing the Sericostoma sp. avoidance behavior to the presence of P. clarkii, leaf mass loss varied between 43.2% in the absence of crayfish and 85.1% in the direct presence of the crayfish (Fig. 2A). Leaf mass loss was significantly higher in mesocosms with P. clarkii (one-way ANOVA, F = 33.1, P b 0.0001). No significant differences were found in leaf consumption by the shredder between mesocosms without P. clarkii or with water previously exposed to the crayfish (Tukey's test, P N 0.05; Fig. 2A). Mean FPOM production varied between 0.2 g in the absence of the crayfish and 1.0 g in the direct presence of the crayfish (Fig. 2B). FPOM production was significantly higher in the presence of crayfish (one-way ANOVA, F = 31.4, P b 0.0001; Fig. 2B). FPOM production did not differ between mesocosms with crayfish water and those without crayfish (Tukey's test, P N 0.05; Fig. 2B).
Physical and chemical stream water parameters were similar at the upstream and downstream sites with the exception of phosphate concentration, which was slightly higher upstream (Table 1). Leaf mass loss was significantly affected by the stream site and the crayfish presence, but not by the presence of invertebrates (three-way ANOVA, F = 4.3, P b 0.05; F = 18.6, P b 0.001 and F = 2.9, P N 0.05, respectively; Fig. 3). At the upstream site, leaf mass loss was lower in the control, intermediate in the treatment with invertebrates only and higher in treatments with the crayfish with or without invertebrates (Tukey's tests, P b 0.05; Fig. 3). At the downstream site, leaf mass loss was lower in the control, intermediate in treatments with invertebrates or crayfish, and higher in the treatment with invertebrates and crayfish (Tukey's tests, P b 0.05; Fig. 3).
Table 1 Minimum and maximum stream water parameters measured at the sampling sites. Parameter
Upstream site
Downstream site
Depth (m) Temperature (°C) pH Conductivity (μS/cm) Dissolved O2 (mg/L) (mg/L) P–PO3− 4 N–NH+ 4 (mg/L) N–NO− 3 (mg/L)
0.30–0.50 15.9–16.6 5.86–6.27 74–78.3 7.98–9.21 0.27–0.28 0.01–0.03 0.07–0.09
0.30–0.50 15.9–16.8 6.16–6.29 77–80 8.14–8.96 0.15–0.16 0.01–0.02 0.06–0.09
718
F. Carvalho et al. / Science of the Total Environment 541 (2016) 714–720
Fig. 3. Leaf mass loss at upstream and downstream sites in the absence or presence of invertebrates and/or P. clarkii. Control: absence of invertebrates and P. clarkii. Mean ± SEM, n = 4. Different letters indicate significant differences between treatments (P b 0.05).
Invertebrate abundance varied between 30 individuals at the upstream site in the presence of crayfish and 167 individuals at the downstream site in the absence of crayfish (Fig. 4A). Invertebrate abundance differed between stream sites and also depended on the presence of the crayfish, with higher values in the absence of the crayfish and at the downstream site (two-way ANOVA, F = 5.1 and F = 8.0, respectively, P b 0.05).
Invertebrate biomass varied between 0.02 g DW at the downstream site in the presence of crayfish and 0.07 g DW downstream in the absence of crayfish (Fig. 4B). Invertebrate biomass was affected by the presence of crayfish and stream site (two-way ANOVA, F = 8.0 and F = 5.1, respectively, P b 0.05 for both factors). At the downstream site, invertebrate biomass was significantly higher in the absence of the crayfish (Tukey's test, P b 0.05). At the upstream site, invertebrate biomass was not affected by the presence of P. clarkii (Tukey's test, P N 0.05). Invertebrate richness measured by the Margalef richness index varied between 2.0 at the upstream site in the presence of crayfish and 3.5 at the downstream site in the absence of crayfish (Fig. 4C). The presence of crayfish and the stream site significantly affected the invertebrate richness (two-way ANOVA, F = 8.3 and F = 6.4, respectively, P b 0.05; Fig. 4C). At the downstream site, the Margalef index was significantly higher in the absence of the crayfish (Tukey's test, P b 0.05) but no differences were detected at the upstream site (Tukey's test, P N 0.05). The nMDS analysis based on the abundance of invertebrates (Fig. 5) discriminated four groups: 1) invertebrate group in the absence of crayfish at the downstream site (DCL2, DCL3, DCL4); 2) invertebrate group in the presence or absence of crayfish at the upstream site (UC1, UC3, UC4, UCL1, UCL3); 3) invertebrate group with four samples from downstream and three samples from upstream in the presence or absence of crayfish (DCL1, DC1, DC2, DC3, UCL2, UCL4, UC2); and 4) invertebrate group with one sample from downstream with crayfish (DC4). The structure of the invertebrate community differed between the upstream and downstream sites (PERMANOVA, Pseudo-F = 1.7, P b 0.05). The presence of crayfish did not affect the structure of the invertebrate community and no interaction between stream site and crayfish presence was found (PERMANOVA, Pseudo-F = 1.4, P N 0.05).
Fig. 4. Invertebrate abundance (A) biomass (B) and Margalef richness index (C) at upstream and downstream sites in the absence or presence of P. clarkii. Mean ± SEM, n = 4. Different letters indicate significant differences between treatments at each site (P b 0.05).
F. Carvalho et al. / Science of the Total Environment 541 (2016) 714–720
Fig. 5. Nonmetric Multi-Dimensional Scaling (nMDS) ordination based on leaf-associated invertebrate community at upstream and downstream sites in the absence or presence of invertebrates and/or P. clarkii. Overlaid clusters correspond to 45% of similarity. (UC (up triangle/filled): Upstream-Crayfish; UCL (up triangle/non-filled): Upstream-Crayfishless; DC (inverted triangle/filled): Downstream-Crayfish; DCL (inverted triangle/non-filled): Downstream-Crayfishless).
4. Discussion Our results suggest that P. clarkii has the potential to affect basal resources in invaded freshwater ecosystems by direct consumption of leaf litter and indirectly by decreasing the abundance of other invertebrates. The direct effects of crayfish on leaf litter decomposition were well established in our laboratory experiments: higher leaf decomposition was observed in the presence of crayfish than of the invertebrate shredder Sericostoma sp., regardless of the abundance of the shredders. This may be explained by the body mass of the crayfish that was much higher than that of Sericostoma sp. or by higher energetic metabolic requirements of P. clarkii in comparison to Sericostoma sp. It is important to point out that the crayfish promptly ate all invertebrate shredders in the first days of laboratory experiments indicating that in the treatments with the crayfish and Sericostoma sp. leaf decomposition was mainly mediated by the crayfish. P. clarkii is well known for its omnivory and shows potentially high predation rates on aquatic invertebrates, mainly insects, crustaceans and gastropods (Momot, 1995; Gherardi, 2006). Therefore, it was expected that P. clarkii would consume first the more energetic resources (shredders) and consumed leaf litter when other resources were unavailable. Interestingly, our results seem to differ from those obtained by Lagrue et al. (2014) with other two crayfish species: Astacus astacus (Linnaeus, 1758) and Pacifastacus leniusculus (Dana, 1852). In that study, both crayfish species did not affect leaf decomposition by direct consumption of basal resources. This discrepancy may happen because i) crayfish diet and trophic niche width vary with species (Usio et al., 2006; Olsson et al., 2009; Jackson et al., 2014), and P. clarkii probably shows high appetency for leaf detritus, and/or ii) the duration of laboratory experiment was longer in our study (21 days vs 6 days), and so the crayfish had the need to feed on other resources after consuming the invertebrates. The production of FPOM followed the pattern found for leaf decomposition described above, with P. clarkii clearly contributing to an increase in the production of FPOM available to other trophic levels. Our results seemed to differ from those of Greig & McIntosh (2006) who showed that an invasive fish (S. trutta) in New Zealand lowered the density of invertebrates and, consequently, the rates of leaf decomposition and FPOM production in mesocosms. In our study, because all Sericostoma sp. were promptly eaten by the crayfish, FPOM production was mainly due to the crayfish activity. Therefore, even if the crayfish eliminates all invertebrate shredders from a particular system, the production of FPOM will not be compromised due to the decomposer
719
behavior of P. clarkii. Nevertheless, if leaf decomposition is driven only by few species, the resilience of the ecosystem may be affected (Peterson et al., 1998). In the same vein, the quality of FPOM produced by P. clarkii may differ from that produced by invertebrate shredders, ultimately affecting other invertebrate feeding groups, namely collector gatherers and collector filterers (Montemarano et al., 2007). However, this was not assessed in our study and further investigation is needed to clarify this hypothesis. To better understand the mechanisms underlying the effects of crayfish on invertebrate shredders, we compared the feeding behavior of Sericostoma sp. in the presence of the predator or waterborne cues from P. clarkii in a laboratory experiment. Our results showed that Sericostoma sp. did not change its shredding behavior on leaf decomposition in mesocosms with water previously exposed to P. clarkii. In the same vein, when shredders had the possibility to avoid direct predation they did not, because after a few moments in the mesocosms, Sericostoma sp. crossed the mesh net to feed on the leaves. This may happen for two reasons: i) the shredders were under starvation and took the risk of being predated, and/or ii) Sericostoma sp. did not recognize P. clarkii as a predator. The last hypothesis is plausible because the shredders used in the experiment were collected from a stream where P. clarkii (or any other crayfish species) was not present. This naivety behavior against invasive predators was already described for many other organisms (e.g. Edgell & Neufeld, 2008; Kuehne & Olden, 2012). In our field experiment, leaf decomposition was higher at the downstream than the upstream stream site in which the presence of P. clarkii was not reported yet. The results obtained at the upstream site were similar to those obtained in the laboratory experiments: decomposition of alder leaves was slightly higher in the presence of crayfish than in the presence of other invertebrates. This can be explained by the size and/or metabolic needs of the crayfish and also because other invertebrates had the possibility to feed inside and outside the baskets, while the crayfish only had the possibility to feed on the leaves enclosed in the baskets. Furthermore, the lack of significant differences between treatments with crayfish in the presence or absence of invertebrates indicates that leaf decomposition was mainly mediated by the crayfish at the upstream site. Moreover, the results obtained in the field support the idea that in streams where diversity and biomass of invertebrate shredders are low, as found at the upstream site, P. clarkii plays an important role in leaf decomposition as shown for other crayfish species (Usio, 2000). At the downstream site (where P. clarkii was already reported), the pattern of leaf decomposition was different from the upstream site: leaf mass loss by the crayfish and invertebrates alone did not differ, and was highest when crayfish and invertebrates were together. Since P. clarkii was already introduced at the downstream site, we cannot discard the hypothesis that invertebrates are able to adjust their behavior after a given time in the presence of P. clarkii. Indeed, at the downstream site the crayfish was well-established and the invertebrate community already had the contact with this species for many years (P. clarkii was introduced in the Minho River basin at least at the beginning of the 1990s; Sousa et al. 2013). Consistently, overall abundance and biomass of invertebrates were higher downstream. This supports the idea that invertebrates may have had enough time to identify the crayfish as a predator, and to adapt their feeding behavior to avoid being predated. In temperate streams, some crayfish species (e.g., Paranephrops zealandicus) affect not only leaf decomposition but also the pattern of colonization by invertebrates (Usio, 2000). Our results in the field seemed to support these observations since the invertebrate community at the downstream site showed higher invertebrate diversity, abundance and biomass. Based on results from the upstream site, we hypothesize that the crayfish may also have an indirect effect on leaf litter due to consumption of native invertebrates. Similar results were described for the crayfish P. leniusculus in Californian (USA) streams (Moore et al., 2012) and other predators in freshwater ecosystems (e.g. Wyman, 1998; Konishi et al., 2001; Mancinelli et al., 2002; Greig
720
F. Carvalho et al. / Science of the Total Environment 541 (2016) 714–720
& McIntosh, 2006), in which predators have an indirect effect on detrital food webs by suppressing the efficiency of leaf litter decomposition. However, our results from the downstream site suggest that indirect effects on leaf decomposition through the consumption of native invertebrates may change through time and invertebrates seem to learn how to avoid the crayfish as a predator. Indeed, in the last years, numerous studies showed that prey may rapidly adapt to the presence of invasive predators evolving anti-predator strategies (e.g. Cox, 2004; Freeman & Byers, 2006). In conclusion, by using a combination of laboratory and field manipulative experiments, we clearly showed that P. clarkii affect detrital food webs by direct consumption of leaf litter and/or by affecting other invertebrate species. Since we are dealing with one of the most widespread IAS that may reach high densities and biomass in aquatic ecosystems, our results are important to develop strategies for improving stream ecosystem functioning and to support management actions aiming to control the invasive omnivore P. clarkii. Acknowledgments The European Regional Development Fund — Operational Competitiveness Program (FEDER-POFC-COMPETE) and the Portuguese Foundation for Science and Technology (FCT) supported this study (PEst-C/ BIA/UI4050/2014, PTDC/AAC-AMB/117068/2010). References Anderson, M.J., 2001. A new method for non-parametric multivariate analysis of variance. Austral Ecology 26, 32–46. Brown, J.S., Laundré, J.W., Gurung, M., 1999. The ecology of fear: optimal foraging, game theory, and trophic interactions. J. Mammal. 80, 385–399. Byrnes, J.E., Reynolds, P.L., Stachowicz, J.J., 2007. Invasions and extinctions reshape coastal marine food webs. PLoS One 3, 295. Cardinale, B.J., Duffy, J.E., Gonzalez, A., Hooper, D.U., Perrings, C., Venail, P., et al., 2012. Biodiversity loss and its impact on humanity. Nature 486, 59–67. Cox, G.W., 2004. Alien species and evolution. Island Press, Washington, United States of America. Diehl, S., 1993. Relative consumer sizes and the strengths of direct and indirect interactions in omnivorous feeding relationships. Oikos 68, 151–157. Dorn, N.J., Wojdak, J.M., 2004. The role of omnivorous crayfish in littoral communities. Oecologia 140, 150–159. Edgell, T.C., Neufeld, C.J., 2008. Experimental evidence for latent developmental plasticity: intertidal whelks respond to a native but not an introduced predator. Biol. Lett. 4, 385–387. Fernandes, I., Duarte, S., Cássio, F., Pascoal, C., 2013. Effects of riparian plant diversity loss on aquatic microbial decomposers become more pronounced with increasing time. Microb. Ecol. 66, 763–772. Fernandes, I., Duarte, S., Cássio, F., Pascoal, C., 2015. Plant litter diversity affects invertebrate shredder activity and the quality of fine particulate organic matter in streams. Mar. Freshw. Res. http://dx.doi.org/10.1071/MF14089 Freeman, A.S., Byers, J.E., 2006. Divergent induced responses to an invasive predator in marine mussel populations. Science 313, 831–833. Geraldes, P., Pascoal, C., Cássio, F., 2012. Effects of increased temperature and aquatic fungal diversity loss on litter decomposition. Fungal Ecol. 5, 734–774. Gessner, M., Chauvet, E., Dobson, M., 1999. A perspective on leaf litter breakdown in streams. Oikos 85, 377–384. Gessner, M.O., Gulis, V., Kueh, K., Chauvet, E., Suberkropp, K., 2007. Fungal decomposers of plant litter in aquatic habitats. In: Kubicek, C.P., Druzhinina, I.S. (Eds.), The Mycota, Vol. IV. Environmental and Microbial Relationships. Springer, Berlin, pp. 301–324. Gherardi, F., 2006. Crayfish invading Europe: the case study of Procambarus clarkii. Mar. Freshw. Behav. Physiol. 39, 175–191. Gherardi, F., Holdich, D.M., 1999. Crayfish in Europe as alien species. How to make the best of a bad situation? A. A. Balkema, Rotterdam Greig, H.S., McIntosh, A.R., 2006. Indirect effects of predatory trout on organic matter processing in detritus-based stream food webs. Oikos 112 (1), 31–40. Grosholz, E., 2002. Ecological and evolutionary consequences of coastal invasions. Trends Ecol. Evol. 17, 22–27. Gutiérrez, J.L., Jones, C.G., Sousa, R., 2014. Toward an integrated ecosystem perspective of invasive species impacts. Acta Oecol. 54, 131–138. Holdich, D.M., 2002. In: Holdich, D.M. (Ed.), Background and functional morphology. Biology of Freshwater Crayfish. Blackwell Science, Oxford. Hooper, D.U., Chapin, F.S.I.I.I., Ewel, J.J., Hector, A., Inchausti, P., Lavorel, S., et al., 2005. Effects of biodiversity on ecosystem functioning: a consensus of current knowledge. Ecol. Monogr. 75, 3–35. Jackson, M.C., Jones, T., Milligan, M., Sheath, D., Taylor, J., Ellis, A., et al., 2014. Niche differentiation among invasive crayfish and their impacts on ecosystem structure and functioning. Freshw. Biol. 59, 1123–1135.
Konishi, M., Nakano, S., Iwata, T., 2001. Trophic cascading effects of predatory fish on leaf litter processing in a Japanese stream. Ecol. Res. 16, 415–422. Kuehne, L.M., Olden, J.D., 2012. Prey naivety in the behavioural responses of juvenile Chinook salmon (Oncorhynchus tshawytscha) to an invasive predator. Freshw. Biol. 57, 1126–1137. Lagrue, C., Podgorniak, T., Lecerf, A., Bollache, L., 2014. An invasive species may be better than none: invasive signal and native noble crayfish have similar community effects. Freshw. Biol. 59 (9), 1982–1995. Lima, S.L., Dill, L.M., 1990. Behavioral decisions made under the risk of predation: a review and prospectus. Can. J. Zool. 68, 619–640. Mancinelli, G., Costantini, M.L., Rossi, L., 2002. Cascading effects of predatory fish exclusion on the detritus-based food web of a lake littoral zone (Lake Vico, central Italy). Oecologia 133, 402–411. McCann, K.S., Hastings, A., Strong, D.R., 1998. Trophic cascades and trophic trickles in pelagic food webs. Proc. R. Soc. Lond. Ser. B Biol. Sci. 265, 205–209. McLean, F., Barbee, N., Swearer, S., 2007. Avoidance of native versus non-native predator odours by migrating whitebait and juveniles of the common galaxiid, Galaxias maculatus. N. Z. J. Mar. Freshw. Res. 41, 175–184. Moore, J.W., Carlson, S.M., Twardochleb, L.A., Hwan, J.L., Fox, J.M., Hayes, S.A., 2012. Trophic tangles through time? Opposing direct and indirect effects of an invasive omnivore on stream ecosystem processes. PLoS One 7 (11), e50687. Momot, W.T., 1995. Redefining the role of crayfish in aquatic ecosystems. Rev. Fish. Sci. 3, 33–63. Montemarano, J.J., Kershner, M.W., Leff, L.G., 2007. Crayfish effects on fine particulate organic matter quality and quantity. Fundamental and Applied Limnology/Archiv für Hydrobiologie 169, 223–229. Olsson, K., Stenroth, P., Nyström, P., Granéli, W., 2009. Invasions and niche width of an introduced crayfish differ from a native crayfish? Freshw. Biol. 54, 1731–1740. Pascoal, C., Cássio, F., Marcotegui, A., Sanz, B., Gomes, P., 2005. Role of fungi, bacteria, and invertebrates in leaf litter breakdown in a polluted river. J. N. Am. Benthol. Soc. 24, 784–797. Pascoal, C., Cássio, F., Nikolcheva, L.G., Bärlocher, F., 2010. Realized fungal diversity increases functional stability of leaf-litter decomposition under zinc stress. Microb. Ecol. 59, 84–93. Peterson, G., Allen, C.R., Holling, C.S., 1998. Ecological resilience, biodiversity, and scale. Ecosystems 1, 6–18. Polis, G.A., Strong, D.R., 1996. Food web complexity and community dynamics. Am. Nat. 147, 813–846. Rodriguez, C.F., Becares, E., Fernandez-Alaez, M., Fernandez-Alaez, C., 2005. Loss of diversity and degradation of wetlands as a result of introducing exotic crayfish. Biol. Invasions 7, 75–85. Sih, A., Bolnick, D.I., Luttbeg, B., Orrock, J.L., Peacor, S.D., Pintor, L.M., Preisser, E., Rehage, J.S., Vonesh, J.R., 2010. Predator–prey naïveté, antipredator behavior, and the ecology of predator invasions. Oikos 119, 610–621. Simberloff, D., Martin, J.-L., Genovesi, P., Maris, V., Wardle, D., Aronson, J., et al., 2013. Impacts of biological invasions: what's what and the way forward. Trends Ecol. Evol. 28, 58–66. Sousa, R., Freitas, F.E., Mota, M., Nogueira, A.J., Antunes, C., 2013. Invasive dynamics of the crayfish Procambarus clarkii (Girard, 1852) in the international section of the river Minho (NW of the Iberian Peninsula). Aquat. Conserv. Mar. Freshwat. Ecosyst. 23, 656–666. Sousa, R., Morais, P., Dias, E., Antunes, C., 2011. Biological invasions and ecosystem functioning: time to merge. Biol. Invasions 13, 1055–1058. Strayer, D.L., 2010. Alien species in fresh waters: ecological effects, interactions with other stressors, and prospects for the future. Freshw. Biol. 55, 152–174. Strayer, D.L., 2012. Eight questions about invasions and ecosystem functioning. Ecol. Lett. 15, 1199–1210. Suberkropp, K., 1998. Microorganisms and organic matter decomposition. In: Naiman, R., Bilby, R. (Eds.), River Ecology and Management: Lessons from the Pacific Coastal Ecoregion. Springer, New York, pp. 120–143. Tablado, Z., Tella, J.L., Sánchez-Zapata, J.A., Hiraldo, F., 2010. The paradox of the long-term positive effects of a North American crayfish on a European community of predators. Conserv. Biol. 24, 1229–1238. Tachet, H., Richoux, P., Bournaud, M., Usseglio-Polatera, P., 2010. Invertébrés d'eau douce: Systématique, biologie, écologie. CNRS éditions, Paris, France. Tilman, D., Isbell, F., Cowles, J.M., 2014. Biodiversity and Ecosystem Functioning. Annu. Rev. Ecol. Evol. Syst. 45 (1), 471. Thompson, R.M., Hemberg, M., Starzomski, B.M., Shurin, J.B., 2007. Trophic levels and trophic tangles: the prevalence of omnivory in real food webs. Ecology 88, 612–617. Usio, N., 2000. Effects of crayfish on leaf processing and invertebrate colonisation of leaves in a headwater stream: decoupling of a trophic cascade. Oecologia 124, 608–614. Usio, N., Suzuki, K., Konishi, M., Nakano, S., 2006. Alien vs. endemic crayfish: roles of species identity in ecosystem functioning. Arch. Hydrobiol. 166, 1–21. Wallace, J.B., Eggert, S.L., Meyer, J.L., Webster, J.R., 1997. Multiple trophic levels of a forest stream linked to terrestrial litter inputs. Science 277, 102–104. Wyman, R.L., 1998. Experimental assessment of salamanders as predators of detrital food webs: effects on invertebrates, decomposition and the carbon cycle. Biodivers. Conserv. 7, 641–650. Yodzis, P., 1984. How rare is omnivory? Ecology 65, 321–323. Zar, J.H., 2009. Biostatistical analysis. Fifth edition. Prentice-Hall, Englewood Cliffs, New Jersey, USA.