Introduction of a functionally novel consumer to a low diversity system: Effects of the mud crab Rhithropanopeus harrisii on meiobenthos

Estuarine, Coastal and Shelf Science xxx (2015) 1e8

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Introduction of a functionally novel consumer to a low diversity system: Effects of the mud crab Rhithropanopeus harrisii on meiobenthos €rnoja Külli Lokko*, Jonne Kotta, Helen Orav-Kotta, Kristiina Nurkse, Merli Pa €ealuse St, 12618 Tallinn, Estonia Estonian Marine Institute, University of Tartu, 14 Ma

a r t i c l e i n f o

a b s t r a c t

Article history: Received 12 March 2015 Received in revised form 4 November 2015 Accepted 16 November 2015 Available online xxx

The Harris mud crab Rhithropanopeus harrisii recently expanded into much of the Baltic Sea. This invasion is expected to have significant effects on the structure and functioning of benthic ecosystems due to the lack of native crabs. Habitat type potentially modulates the effects as crabs are expected to behave differently in different habitats. In this study we experimentally evaluated the effect of R. harrisii on the species composition and dominance structure of shallow water meiobenthos within common habitat types of the north-eastern Baltic Sea. Among the studied environmental variables R. harrisii had by far the strongest effects on meiobenthos. The effects of R. harrisii varied among different habitats with the crab mostly modifying taxonomic composition and species abundances of meiobenthic communities on unvegetated soft bottom sediments. Our experiment also showed that boulders provided shelter for R. harrisii and thereby reduced their burrowing activity and effects on the adjacent soft bottom meiobenthos. © 2015 Elsevier Ltd. All rights reserved.

Keywords: Meiobenthos Brackish water Introduced species Sediment properties Mud crab Baltic Sea

1. Introduction Invasive non-native species are one of the biggest threats to nature as they can alter ecosystems and economies (Simberloff, 2011; Strayer, 2012). This is particularly true to ocean ecosystems, primarily near shorelines where human use is exponentially increasing and diversifying (Galil et al., 2014). Although there is no doubt that invasive species have modified marine ecosystems, evidence for most of the reported impacts is still weak (e.g. Ojaveer and Kotta, 2015). However, the impacts of these changes on the structure and functioning of ecosystems and the social and economic consequences that then arise need to be well understood to inform prioritization of management interventions. Until recently, vast areas of the Baltic Sea hosted no native crabs or other organisms with similar functional roles (Bonsdorff and Pearson, 1999). However, after an invasion of the Harris mud crab Rhithropanopeus harrisii (Gould, 1841) the situation changed dramatically. In the late 2000s and the early 2010s R. harrisii was discovered in the northern and eastern Baltic Sea including the Archipelago Sea along the southwestern coast of Finland and the

* Corresponding author. E-mail address: [email protected] (K. Lokko).

northeastern Gulf of Riga in Estonia. Since then the species has been exponentially increasing its range and density (Kotta and Ojaveer, 2012; Fowler et al., 2013). Owing to the prior absence of any large burrowers and epibenthic predators this introduced species is expected to form a new functional link within existing assemblages and cause pronounced impacts on community structure, food webs and ecosystem functioning. This natural experiment offers an excellent opportunity to observe how communities that evolved without this functional guild react to the presence of R. harrisii and thereby better understand the role of crabs in coastal ecosystems in general. The introduction of R. harrisii will likely cause changes in the abundance of organisms across many trophic levels (e.g. Carpenter and Kitchell, 1993). For example, crabs may control the abundance of grazers (Silliman et al., 2004) and thereby modify the growth patterns of micro- and macroalgae. Crabs may also prey on deposit and suspension feeding bivalves and thereby affect the trophic state of sediment (Norkko et al., 2001; Giles et al., 2006; Cranford et al., 2009). Burrowing crabs can lower snails' growth rates due to increased snail burial (Armitage and Fong, 2006) and disturb bivalves' feeding behaviour and damage their shells (Lomovasky et al., 2006). Moreover, chemical cues caused by crab predation may induce prey populations to change their behaviour and such

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Please cite this article in press as: Lokko, K., et al., Introduction of a functionally novel consumer to a low diversity system: Effects of the mud crab Rhithropanopeus harrisii on meiobenthos, Estuarine, Coastal and Shelf Science (2015), http://dx.doi.org/10.1016/j.ecss.2015.11.017

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K. Lokko et al. / Estuarine, Coastal and Shelf Science xxx (2015) 1e8

alterations may have dramatic consequences for the functioning of benthic ecosystems (Premo and Tyler, 2013). Finally, the mud crab as an effective burrower effectively reworks sediment through bioturbation that affects sediment granulometry and organic matter content (Botto and Iribarne, 2000), sediment biogeochemistry, pore water chemical characteristics and nutrient benthic fluxes (Gilbert et al., 1998; Fanjul et al., 2007, 2011). Most of these issues have not yet been addressed in the Baltic Sea. There exists only one experimental study from the northern Baltic Sea suggesting that R. harrisii is already decreasing species richness of €m littoral hard bottom macroinvertebrate community (Forsstro et al., 2015). Meiofauna (i.e. benthic organisms that can pass through a 500 mm mesh sieve but are retained on a 44 mm sieve; Giere, 2009) represents one of the most abundant and diverse groups of animals and have important functions in marine food webs; nevertheless, they are relatively understudied in many parts of the world (Giere, 2009). Crabs can influence meiobenthos through different mechanisms. Although this is not known for R. harrisii, the soldier crab Mictyris longicarpus can directly feed on meiofauna (Spilmont et al., 2009). The burrowing activity of crabs can cause physical disturbance that directly alter the abundance and species composition of meiobenthos (Warwick et al., 1990; Rosa and Bemvenuti, 2005). However, the effects are known to vary among taxa. For example, crab burrowing reduces the abundance of ostracods, copepods and turbellarians (Rosa and Bemvenuti, 2005) but not nematodes (Warwick et al., 1990) and in some instances their abundance may even be increased (Shimanaga et al., 2012). Alternatively, crab faecal pellets together with associated bacteria might serve as a new food resource for some meiofaunal taxa. Studies involving R. harrisii suggest that their diet consists of detritus to a considerable extent (e.g. Czerniejewski and Rybczyk, 2008) and thereby it is expected that R. harrisii may compete strongly for food with some taxa of meiofauna. On the other hand, R. harrisii preys to great extent on deposit feeding bivalves (Czerniejewski and Rybczyk, 2008; Ojaveer and Kotta, 2015) thus, they may likely reduce competitive interactions for food among detritus feeding meiobenthos. Moreover, the reduction of macrofauna due to crab predation (e.g. Milke and Kennedy, 2001) may have severe repercussions for meiofauna, as macrofauna are known to have an important structuring effect on meiobenthic communities  (Olafsson, 2003). The evidence above indicates that the potential effects of the crab on meiofauna are diverse and the introduction of R. harrisii is likely to disrupt natural meiobenthic communities by establishing new ecological relationships, displacing or completely disassembling native communities, but likely facilitating some native species through the provision of alternative food to native consumers. Considering the vast array of potential impacts and the prior lack of crabs in the northeastern Baltic Sea basin, the effects of R. harrisii on meiobenthos are difficult to predict. Moreover, there are virtually no studies on the effects of any invasive species on the Baltic Sea meiobenthos (but see Urban-Malinga et al., 2013). In its native range R. harrisii is commonly found in habitats that provide some type of shelter from predators either in the form of plants or debris (Ryan, 1956). In its invasive range, however, the species is even more flexible occurring from exposed hard bottoms to sheltered soft bottoms either unvegetated or vegetated (Fowler et al., 2013). In the Baltic Sea R. harrisii is abundant both in soft sediment and mixed bottom habitats. On mixed bottom habitats, R. harrisii is known to avoid soft sediment and hide either under boulders or among boulders covered with macroalgae (Nurkse et al., 2015). On the other hand, on soft sediment the invasive crab constantly reworks sediment deposits through bioturbation and thereby posing a significant disturbance to the meiobenthic communities. Therefore, the influence of R. harrisii on meiobenthos

is expected to vary among habitats and such habitat specific interactions need to be experimentally assessed. The aim of this study was to experimentally investigate the effect of R. harrisii on the taxonomic composition and abundance of a shallow water meiofauna community in the north-eastern Baltic Sea. The following questions were asked: (1) Does habitat type modify the effects of R. harrisii on meiofauna? Potentially, R. harrisii has stronger effect on unvegetated soft bottoms than on other types of environments as such habitat is characterised by low availability of food and microhabitats and thus higher feeding and sediment reworking activity rates. (2) Does R. harrisii have variable effects on different meiobenthic taxa? We expect that R. harrisii enriches sediment with organic matter and thereby enhances abundances of nematodes, ciliates and disfavours oligochaetes, turbellarians and Gastrotricha. (3) Does R. harrisii have variable effects on meiobenthos species composition and total abundance? We expect that R. harrisii has a larger effect on meiobenthos species composition than on total abundances and this is because sensitivities of species vary among taxa whereas total abundance is often a function of nutrient availability, the latter being largely unaffected by the presence of crabs. 2. Materials and methods 2.1. Experiment design and set-up In order to investigate the effect of R. harrisii on meiobenthos, a ~ iguste Bay, the manipulative experiment was conducted in Ko north-eastern Baltic Sea in June 2013 (58
22.100 N, 22
58.690 E) using 10 l plastic buckets with a diameter of 24 cm. The sediment (fine sand) together with meiobenthos was collected from a shallow water embayment (0.5e1 m depth) adjacent to the experiment site. In order to obtain similar starting conditions, sediment was homogenized prior to the experiment. In the homogenized sediment the medium particle size of sand varied within 2 mm and the content of silt varied less than 2%. Then the buckets were filled with a 6 cm layer of homogenized sand and natural seawater of salinity 6 and were allowed to settle for 6 h. The filled buckets were placed into an outdoor mesocosm facility, assuring ambient light and seawater temperature for all experimental buckets. Coastal waters of the north-eastern Baltic Sea are dominated by soft and mixed bottom habitats. Owing to habitat-specific behavioural differences the role of R. harrisii on meiobenthos is expected to vary among habitats (Nurkse et al., 2015). We used four different habitat types in the experiment. Soft sediments contained homogenized sand only, whereas mixed sediments were generated by adding two similar-sized small stones on the homogenized sands. For vegetated soft sediments the charophyte Chara aspera were seeded and on mixed sediment stones overgrown with the green alga Cladophora glomerata were placed on sand instead of unvegetated stones. Stones and macroalgae were collected adjacent to the experiment site. In order to start the experiments with similar numbers of species, all stones and macroalgae were inspected for all macroinvertebrates which were then removed from the substrates. In addition, the substrates were carefully rinsed in a bucket of freshwater in order to remove associated motile fauna before introducing them into the experimental buckets. The density of R. harrisii is not well known in the Baltic Sea as current monitoring programs do not target such epibenthic predators due to prior lack of such a function. A pilot experiment deploying artificial reefs in the north-eastern Baltic Sea area has indicated that the number of R. harrisii per 1 m2 seafloor surface area ranges from a few crabs to 1000 crabs and these values (range and density) are still increasing (Fowler et al., 2013; J. Kotta

Please cite this article in press as: Lokko, K., et al., Introduction of a functionally novel consumer to a low diversity system: Effects of the mud crab Rhithropanopeus harrisii on meiobenthos, Estuarine, Coastal and Shelf Science (2015), http://dx.doi.org/10.1016/j.ecss.2015.11.017

K. Lokko et al. / Estuarine, Coastal and Shelf Science xxx (2015) 1e8

unpublished data). In order to avoid intraspecific competition between crabs only one crab was added per bucket. This value is within a range the natural density of crabs in the study area. The crabs used in the experiment were all of similar size and their carapace length were estimated at 12.1 ± SE 0.5 mm. In summary, the experiment consisted of the following treatments and treatment levels: crab presence (present, absent) and habitat type (soft unvegetated sediment, soft sediment with Chara, mixed unvegetated sediment, mixed sediment with Cladophora); resulting in 8 treatment combinations, each replicated five times. 2.2. Sampling and sample analyses After incubation for 4 days one core sample was taken from each bucket 5 cm down into the sediment using a sharp-edged corer with an inner diameter of 2.7 cm. Care was taken that samples did not contain benthic macroalgae and macroinvertebrates. Each sample was transferred to plastic container, immediately rinsed three times with carbonated drinking water, every time the supernatant was poured off into plastic bottle, and then fixed with Lugol's solution. The samples were condensed to 100 ml by sedimentation for 4 days and siphoning the excess water. From each sample, five 2 ml subsamples were taken wherein all organisms were counted under a microscope in a Bogorov chamber. For the identification, the specimens were slide-mounted in glycerol and determined down to the genus level when possible, using an Olympus BX 50 microscope. In each sample 25 nematode individuals were randomly chosen, slide-mounted and identified to the lowest possible taxonomic level. Additional core samples for sand grain size, chlorophyll a and organic content of the sediment were collected from each bucket. Sediment grain size was determined using sieving techniques. The following sieves were used: 0.5, 0.25 and 0.125 mm. The software Gradistat, version 8 (Blott and Pye, 2001) was used to calculate sorting and to obtain the mean percentages of different sediment fractions. The content of sediment organic matter was measured as a loss of ignition at 500
C during 3 h. Chlorophyll a samples were extracted in 96% ethanol overnight. Chlorophyll a was quantified fluorometrically correcting for phaeopigments (Strickland and Parsons, 1972). 2.3. Data analysis In order to describe the relationship between environmental variables and the taxonomic composition and dominance structure of meiofauna community a canonical correspondence analysis (CCA) was carried out on complete environmental data set. CCA is one of the most widely used ordination techniques today. Like other ordination techniques CCA orders objects (communities) that are characterized by values on multiple variables (species presences or abundances) so that similar objects are near each other and dissimilar objects are farther from each other. The results of ordination are typically viewed as 2-dimensional graphs. Moreover, CCA attempts to explain differences in community structure between sites in terms of differences in environmental variables. The analysis was completed using in the statistical software R (The R Foundation for Statistical Computing, 2013). Then a crossed two-factor non-parametric permutational analysis of variance (PERMANOVA) was performed to investigate the separate and interactive effects of the Harris mud crab and habitat type on the structure of meiobenthic communities. Prior to analysis, a BrayeCurtis similarity matrix was calculated using raw data (untransformed) and presence/absence transformation to detect whether the potential differences between the meiobenthic

3

communities were due to differences in relative abundances or species composition. When an interaction was identified as significant (p < 0.05), post-hoc PERMANOVA pair-wise tests were conducted to detect which levels of the factors involved differed from each other. Taxa responsible for observed differences were identified by the similarity percentages test (SIMPER). PERMANOVA and SIMPER analyses were conducted using PRIMER 6 PERMANOVAþ software (Clarke and Gorley, 2006). In addition, a crossed two-way analysis of variance (ANOVA) was performed to investigate the separate and interactive effects of R. harrisii and habitat type on sediment properties (the content of sediment organic matter, chlorophyll a and silt) using the package anova.lm in the R environment (The R Foundation for Statistical Computing, 2013). When a factor or interaction was identified as significant (p < 0.05), post-hoc Bonferroni tests were conducted to determine which treatment levels were significantly different. As the total abundance of broad groups of meiobenthic taxa (turbellarians, nematodes, rotifers, ciliates, gastrotrichs, insect larvae, juvenile bivalves, copepods and copepod nauplii) did not follow the assumptions of ANOVA analysis, the permutational factorial ANOVA was performed instead to investigate the separate and interactive effects of R. harrisii and habitat type on these aggregated measures using the package Vegan in the R environment (Oksanen et al., 2015). 3. Results The sediment used in the experiment was classified as moderately sorted fine sand. The content of fine sand (125e250 mm) varied from 80 to 89% and that of silt from 8 to 16%. The sediment organic content was similar in all mesocosms estimated at 0.17e0.61%, while chlorophyll a content varied from 0 to 10.78 mg corer1. R. harrisii had no significant effect on any measured sediment properties (2-way ANOVA p > 0.05 for the separate and interactive effects of crab). The meiofaunal communities in the buckets consisted of rotifers, nematodes, ciliates, testaceans, small crustaceans, gastrotrichs, turbellarians, settled bivalves and insect larvae (Table 1). Nematodes were the most abundant taxa in all treatment groups. The CCA analysis showed that R. harrisii contributed more to variation in meiobenthic communities than any other studied environmental variable (CCA pcrab ¼ 0.002; Fig. 1). Apart from R. harrisii, the content of sediment silt and organic matter explained a large fraction of variability in the species composition and abundance of meiofauna. The effects of R. harrisii and soft unvegetated sediment co-varied (Hab1 in Fig. 1) whereas other habitat types affected meiobenthos largely independently of R. harrisii. The CCA analysis also showed that R. harrisii had clear taxon-specific effects on meiofauna. In general, the presence of R. harrisii favoured testaceans, turbellarians and rotifers and disfavoured copepod nauplii and juvenile bivalves. PERMANOVA complemented the results of CCA analysis and showed that R. harrisii had no overall effect on the taxonomic composition and dominance structure of meiofauna communities (p ¼ 0.134 and p ¼ 0.234 respectively). However, in interaction with habitat type R. harrisii affected taxonomic composition (p ¼ 0.048; Table 2), but not dominance patterns of meiofauna (p ¼ 0.667). R. harrisii modified meiofauna community in habitats with unvegetated soft bottoms (post-hoc PERMANOVA pair-wise test; p ¼ 0.004) but not in other habitat types (soft sediment with Chara: p ¼ 0.100; mixed unvegetated sediment: p ¼ 0.946; mixed sediment with Cladophora: p ¼ 0.231). In unvegetated soft bottoms R. harrisii promoted abundance and probability of occurrence of ciliates, rotifers and insect larvae as well as abundance of nematodes. On the other hand the abundance and probability of

Please cite this article in press as: Lokko, K., et al., Introduction of a functionally novel consumer to a low diversity system: Effects of the mud crab Rhithropanopeus harrisii on meiobenthos, Estuarine, Coastal and Shelf Science (2015), http://dx.doi.org/10.1016/j.ecss.2015.11.017

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K. Lokko et al. / Estuarine, Coastal and Shelf Science xxx (2015) 1e8

Table 1 Meiofaunal taxa in the studied habitats. Taxa Testatcea Ciliata Cercozoa Turbellaria Gastrotricha Rotifera

Bivalvia (juv) Nematoda

Ostracoda Copepoda Diptera (juv)

Centropyxis Difflugia Cyphoderia ampulla

Colurella Lecane Trichocerca Adoncholaimus Anoplostoma Axonolaimus cf. Chromadora Chromadorina Chromadorita cf. Doliolaimus Enoplolaimus Leptolaimus papilliger Leptolaimus sp2 Linhomoeidae sp1 Linhomoeidae sp2 Microlaimidae sp1 Microlaimidae sp2 Monhysteridae sp1 cf. Prochromadorella Viscosia Undetermined sp1 Undetermined sp2 Undetermined sp3 Undetermined sp4 Undetermined sp5 Undetermined sp6

Soft unvegetated

Soft with Chara

x

x

x x x x x x

x x x x x

x x

x x x x x x

x x

x x

x x

x x x x x

x x

x x x x x x x

x x x

x x x x

Mixed unvegetated

Mixed with Cladophora

x

x x x x x

x x

x x x x

x x

x x

x x x

x

x x

x

x

x

x x

x x x

x x

x x x x x

x x x

x x x

occurrence of gastrotrichs, copepods and settled bivalves was higher in buckets without R. harrisii (SIMPER analysis). PERMANOVA analyses performed separately for different broad taxonomic groups of meiofauna showed that R. harrisii modified

x x

species composition and abundances of turbellarians (PERMANOVA, p ¼ 0.014, Table 3) and juvenile bivalves (PERMANOVA, p ¼ 0.017). In interaction with habitat type R. harrisii also modified species composition and abundances of insect larvae in unvegetated soft sediment (PERMANOVA, p ¼ 0.049). For other taxa main effects and interactions were not significant. However, the presence of crab was associated with elevated numbers of nematode taxa. Univariate tests of permutational ANOVA on the total abundance of broad groups of meiobenthic taxa fully matched the community level effects of R. harrisii described in the paragraph above. The permulational ANOVA tests showed that R. harrisii

Table 2 The results of PERMANOVA analyses on the effects of habitat type and R. harrisii to the taxonomic composition (presence/absence data) and dominance structure (untransformed abundance data) of meiofauna communities. Model/Factor

Fig. 1. Results of CCA ordination demonstrating differences in responses of meiofaunal taxa to R. harrisii and environmental variables. Hab1 e soft unvegetated sediment; Hab2 e soft sediment with Chara aspera; Hab3 e mixed unvegetated sediment; Hab4 e mixed sediment with Cladophora glomerata. Filled circles represent buckets with macroalgae and empty circles represent mesocosms without macroalgae.

df

SS

Untransformed data Habitat 3 3767 Crab 1 1256 Habitat*Crab 3 2088 Res 32 27,778 Total 39 34,888 Presence/absence data Habitat 3 3670 Crab 1 2182 Habitat*Crab 3 6929 Res 32 37,673 Total 39 50,454

MS

Pseudo-F

P

No permutations

1255.6 1255.9 695.9 868.1

1.446 1.447 0.802

0.146 0.234 0.667

998 999 999

1223.2 2182.4 2309.6 1177.3

1.039 1.854 1.962

0.427 0.134 0.048

998 999 999

Bold text indicates a statistically significant relationship.

Please cite this article in press as: Lokko, K., et al., Introduction of a functionally novel consumer to a low diversity system: Effects of the mud crab Rhithropanopeus harrisii on meiobenthos, Estuarine, Coastal and Shelf Science (2015), http://dx.doi.org/10.1016/j.ecss.2015.11.017

K. Lokko et al. / Estuarine, Coastal and Shelf Science xxx (2015) 1e8

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Table 3 The results of PERMANOVA analyses on the effects of habitat type and R. harrisii to the dominance structure (untransformed data) of various groups of meiofauna. Model/Factor

df

Rotifers Habitat 3 Crab 1 Habitat*Crab 3 Res 32 Total 39 Ciliates Habitat 3 Crab 1 Habitat*Crab 3 Res 32 Total 39 Turbellarians Habitat 3 Crab 1 Habitat*Crab 3 Res 32 Total 39 Nematodes Habitat 3 Crab 1 Habitat*Crab 3 Res 32 Total 39 Gastrotrichs Habitat 3 Crab 1 Habitat*Crab 3 Res 32 Total 39 Copepods Habitat 3 Crab 1 Habitat*Crab 3 Res 32 Total 39 Copepod nauplii Habitat 3 Crab 1 Habitat*Crab 3 Res 32 Total 39 Juvenile bivalves Habitat 3 Crab 1 Habitat*Crab 3 Res 32 Total 39 Insect larvae Habitat 3 Crab 1 Habitat*Crab 3 Res 32 Total 39

SS

MS

Pseudo-F

P (perm)

Unique perms

232 198 1099 6492 8020

77.2 197.9 366.3 202.9

0.380 0.975 1.806

0.839 0.332 0.169

999 999 998

164 337 2486 9808 12,795

54.6 337.0 828.6 306.5

0.178 1.100 2.704

0.957 0.302 0.063

999 999 999

114 232 114 1327 1787

37.9 231.8 37.9 41.5

0.913 5.589 0.913

0.470 0.014 0.465

999 998 998

1180.2 596.0 119.5 398.0

2.836 1.498 0.300

0.047 0.220 0.905

999 999 999

629 35 138 4501 5303

209.6 34.7 46.1 140.7

1.490 0.247 0.327

0.228 0.660 0.831

998 999 998

36 165 426 2610 3236

11.9 164.6 141.9 81.5

0.146 2.019 1.740

0.945 0.156 0.168

999 998 999

291 269 132 3114 3805

96.9 269.1 43.9 97.3

0.996 2.765 0.451

0.409 0.100 0.702

998 998 999

145 410 449 1864 2868

48.5 409.7 149.5 58.3

0.832 7.031 2.566

0.477 0.017 0.063

998 999 999

241 3 1039 3747 5030

80.3 2.7 346.4 117.1

0.686 0.023 2.958

0.579 0.938 0.049

997 999 999

3152 596 358 12,735 16,841

Bold text indicates a statistically significant relationship.

raised the abundance of turbellarians (2-way permutational ANOVA, main effect of R. harrisii: p ¼ 0.043) and decreased the abundance of juvenile bivalves (2-way permutational ANOVA, main effect of R. harrisii: p ¼ 0.009). In interaction with habitat type, the mud crab significantly promoted the abundance of insect larvae in unvegetated soft sediment (2-way permutational ANOVA, interactive effect of R. harrisii and habitat type: p ¼ 0.044). For other taxa main effects and interactions were statistically not significant at p > 0.05 (Fig. 2). 4. Discussion The results of this experiment demonstrate that the invasive

Fig. 2. Average abundance (ind cm3 ± SE) of turbellarians, juvenile bivalves and insect larvae with and without R. harrisii in different habitats.

R. harrisii potentially affects shallow water meiobenthic communities in the north-eastern Baltic Sea. As seen from the results of multivariate analyses, R. harrisii shaped the community of meiofauna more than other studied environmental variables. Only sediment silt content had a comparable effect on meiofauna. This is because silt content is the key environmental factor for meiofauna

Please cite this article in press as: Lokko, K., et al., Introduction of a functionally novel consumer to a low diversity system: Effects of the mud crab Rhithropanopeus harrisii on meiobenthos, Estuarine, Coastal and Shelf Science (2015), http://dx.doi.org/10.1016/j.ecss.2015.11.017

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and sediments with high silt content are often considered as unfavourable for many meiobenthic organisms (Nogrady et al., 1993; Giere, 2009). As compared to R. harrisii, habitat type had no separate effects on meiobenthos. However, habitats in interaction with R. harrisii strongly contributed to the variability of meiobenthic communities. The experiment confirmed that R. harrisii has potential to significantly alter the taxonomic composition of meiobenthos in unvegetated soft bottom habitats but not in other habitat types. Small stones in treatments with mixed sediments as well as vegetation provide shelter for R. harrisii and the crabs only rarely go into soft sediment (Nurkse et al., 2015). On unvegetated soft sediment habitats, however, R. harrisii constantly reworks sediment deposits through bioturbation, thereby posing a significant disturbance to the meiobenthic communities. Moreover, vegetated sediments may also provide more microhabitats for meiobenthos (Giere, 2009) and with an increasing number of microhabitats the elevated diversity of meiobenthos including stress tolerant species is expected (Gingold et al., 2010). Consequently, such an elevated species diversity may promote higher meiobenthic community stability against various stressors including those associated with the presence of R. harrisii. PERMANOVA analysis showed that the mud crab modified taxonomic composition of meiobenthos rather than abundance. While R. harrisii had an effect on the abundance of turbellarians and settled juvenile bivalves, no significant effects were observed for nematodes, a dominant component of the studied community. This contrasts to an earlier observation (Warwick et al., 1990) that burrowing crabs are expected to reduce species richness, diversity and evenness of nematode community without actually changing the overall density of nematodes. In our experiment, R. harrisii significantly reduced the abundance of juvenile bivalves. The mechanisms behind the observed decline may be related to direct predation, mechanical damage due to crab burrowing and/or the transport of organic matter into the deeper layers of sediment i.e. a reduction of food availability in the sediment surface (Strasser and Günther, 2001; Lomovasky et al., 2006; Whitton et al., 2012). R. harrisii significantly reduced the mortality of turbellarians. Turbellarians are known to prefer well oxygenated fine sandy sediments rich of food supply (Giere, 2009); the majority are predatory, but seldom themselves preyed upon (Martens and Schockaert, 1986). Thus, crab burrowing and consequent increases in organic matter (Botto and Iribarne, 2000) in sediment can be beneficial to turbellarians. Similarly, Urban-Malinga et al. (2013) have recently shown that bioturbation by macrofauna increases abundance of turbellarians and this is also in line with a recent finding that sand structure is the key environmental variable for turbellarians (Lokko et al., 2014). In some cases, however, a burrowing crab may also have negative impacts on turbellarians  (Rosa and Bemvenuti, 2005) or no impact at all (Olafsson and Ndaro, 1997). In unvegetated soft sediment, insect larvae, mostly chironomids, were much more abundant in the presence than in the absence of R. harrisii. It has been shown that detritus is an important food source for chironomid larvae (Hirabayashi and Wotton, 1999; Henriques-Oliveira et al., 2003). While R. harrisii had no significant effect on the content of sediment organic matter, it may be possible that the faecal material of crabs together with associated bacteria serves as a rewarding food resource in habitat with poor feeding conditions. Our experiment demonstrated that within a few days only, the R. harrisii had several direct and interactive effects on meiobenthos. However, no effects on the sediment properties were detected except for a slight increase in sediment silt content in the presence

of R. harrisii. Long-term exposure to R. harrisii will likely be more severe but less predictable due to the presence of many indirect effects. For example, R. harrisii is known to feed on adult bivalves (Hegele-Drywa and Normant, 2009) and therefore the abundance of deposit feeders may be severely reduced in benthic communities in the presence of the mud crab. This can relax interspecific competition for food between meio- and macrofauna and ultimately boost up the density of meiobenthos. This statement is supported by existing experimental evidence on the increased mortalities of meiobenthic ostracods either through physical smothering and/or intensified food competition induced by e.g. the native bivalve Macoma balthica (Nascimento et al., 2011). The presence of M. balthica also reduces the abundance of nematodes, copepods and oligochaetes by preventing the development of  microalgal mats in sediment surface (Olafsson et al., 2005). Thus, meiobenthic taxa that are not disfavoured by the crab burrowing activity may become more abundant. Alternatively, some suspension-feeding bivalves enhance meiobenthos by organic-rich biodeposits that provide an excellent food source (Castel et al., 1989; Kotta et al., 2009). As the distribution of R. harrisii is not limited by reduced salinity (Boyle et al., 2010; Roche et al., 2009; Patton et al., 2010), the mud crab may invade the whole northeastern Baltic Sea including adjacent inland waterbodies and rivers. Some specimens are already reported from the Rhine, hundreds of kilometres upstream from the sea (Freyhof and Steinmann, 1998). Given its large habitat range and lack of natural enemies in the invaded waterbodies (e.g. Fowler et al., 2013), R. harrisii has the potential to affect communities of meiobenthos, not only in the north-eastern Baltic Sea, but likely also in the adjacent fresh-water basins. It is important to stress that our study is a short term experiment and care should be taken with extrapolating experimental results to the field. For example, we did not detect any significant effect of R. harrisii on the sediment properties whereas such effects may appear in the longer run. The actual outcome of the establishment of R. harrisii depends on abiotic environmental settings, the synergistic effects of the mud crab on meio- and macrofauna as well as the indirect effects of macrobenthos on meiobenthos. Thus, further long-term and broad-scale in situ experimental research is needed to tackle all the possible impacts of R. harrisii and its implications for the whole shallow water benthic ecosystem and services provided. However, based on the existing evidence and this study, this species very likely strengthens top-down control of macrophytes, meio- and macrofauna in the shallow water ecosystem of the Baltic Sea.

Acknowledgements Many thanks to Dr. Taavi Virro for help in identifying several taxa. Funding for this research was provided by Institutional research funding IUT02-20 of the Estonian Research Council. The study has been also supported by the projects “The status of marine biodiversity and its potential futures in the Estonian coastal sea” No 3.2.0801.11-0029 of Environmental protection and technology program of European Regional Fund. The project has received funding from the BONUS project BIO-C3, the joint Baltic Sea research and development programme (Art 185), funded jointly from the European Union's Seventh Programme for research, technological development and demonstration and from the Estonian Research Council. The study received funding also from the European Community's Seventh Framework Programme (FP7/ 2007e2013) under Grant Agreement No. 266445 for the Vectors Change in Oceans and Seas marine Life, Impact on Economics Sectors (VECTORS).

Please cite this article in press as: Lokko, K., et al., Introduction of a functionally novel consumer to a low diversity system: Effects of the mud crab Rhithropanopeus harrisii on meiobenthos, Estuarine, Coastal and Shelf Science (2015), http://dx.doi.org/10.1016/j.ecss.2015.11.017

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