Northeastern Naturalist Vol. 21, No. 3 L.A. Auker, A.L. Majkut, and L.G. Harris 2014 479 2014 NORTHEASTERN NATURALIST 21(3):479–494 Exploring Biotic Impacts from Carcinus maenas Predation and Didemnum vexillum Epibiosis on Mytilus edulis in the Gulf of Maine Linda A. Auker1,*, Alison L. Majkut1,2, and Larry G. Harris3 Abstract - Mytilus edulis (Blue Mussel) is an ecologically important species in the Gulf of Maine. However, many introduced species that have a direct negative impact on the Blue Mussel have entered this system, some as predators (e.g., Carcinus maenas [Green Crab]) and others as aggressive epibionts (e.g., Didemnum vexillum [Carpet Sea Squirt]). Didemnum vexillum has been increasing in abundance throughout the Gulf for the past 10 years and form large mat-like growths on mussel beds, covering individual mussels completely. The first part of our study used a predator-exclusion experiment to determine the impact of predators on the plantigrade stage of the Blue Mussel life cycle. During this stage, no epibiosis occurs due to a protective periostracum layer on the mussel shell. The second part of our study used laboratory trials to assess how overgrowth by D.vexillum impacts predator choice, handling time, and consumption of mussels. There were a significantly greater number of Blue Mussel plantigrades on exclusion panels than on the exposed-cage control panels. Green Crab and Nucella lapillus (Dog Whelk) predators were present on our nonexclusion panels. In laboratory trials, Green Crab handling time of Blue Mussels was not significantly different between mussels that were clean and mussels that were overgrown, but crab behavior and overall consumption showed a greater selection for clean mussels. This selection indicates an associational predator-resistance effect of D.vexillumt epibiont on Blue Mussels. The results of our study, while focused on one specific predator species, suggest that while young Blue Mussels with no epibionts are preyed upon heavily, D.vexillum likely deters predators from older mussels. Because D.vexillum form large matlike colonies that can cover a large area, their presence may have a significant impact on community structure in the Gulf of Maine. Introduction Epibiosis and predation The overgrowth of one living organism by another is known as epibiosis. Both the overgrown organism (the basibiont) and the overgrowing organism (the epibiont) are impacted by this relationship, as are other organisms that attempt to interact with the basibiont (Buschbaum et al. 2007, Enderlein et al. 2003, Wahl 1989). Basibionts may gain advantages from the relationship (mostly in the form of protection from predation), but frequently suffer significant disadvantages (Burlakova et al. 2000; Buschbaum and Saier 2001; Haag et al. 1993; Ricciardi et al. 1995; Thieltges 2005; Wahl 1989, 1997). Such disadvantages may include decreased buoyancy 1Department of Biology, Siena College, 515 Loudon Rd., Loudonville, NY 12211. 2Boston Heart Diagnostics, 175 Crossing Boulevard, Framingham, MA 01702. 3Department of Biological Sciences, University of New Hampshire, 38 College Road, Durham, NH 03824. Corresponding author - lauker@stlawu.edu. Manuscript Editor: Melisa Wong Northeastern Naturalist 480 L.A. Auker, A.L. Majkut, and L.G. Harris 2014 Vol. 21, No. 3 of the basibiont, hindered motion, damage to surfaces, reduction of diffusion of soluble materials, and direct competition with the epibiont (Wahl 1989). Epibiosis can alter predator–prey relationships by creating a new interface on the prey (Wahl et al. 1997). That is, predators may attack or avoid the epibiont leading to profound positive or negative effects on the basibiont. When epibiosis changes predator responses, it does so through either associational resistance or shared doom (Wahl and Hay 1995). Associational resistance occurs when an epibiont deters predator attacks, thus reducing the risk to itself and the basibiont host (Laudien and Wahl 1999, Marin and Belluga 2005, Thieltges 2005, Thornber 2007, Vance 1978, Wahl and Hay 1995). This situation typically occurs when epibiont species mask chemical cues from the basibiont (Wahl et al. 1997), or repels the predator through chemical deterrence (Laudien and Wahl 2004, Wahl et al. 1997). Shared doom occurs when predators consume the epibiont, resulting in either incidental or deliberate consumption of the basibiont host (Buschbaum et al. 2007, Enderlein et al. 2003, Farren and Donovan 2007, Wahl et al. 1997, Wahl and Hay 1995). In the case of shared doom, epibionts may enhance attractive chemical cues (Wahl et al. 1997) or improve prey handling (Enderlein et al. 2003). Mytilus edulis L. (Blue Mussel) is an important ecological species and a dominant member of consumer-regulated stable communities in the Gulf of Maine (Bertness et al. 2002). Blue Mussels are both a food source for multiple organisms (Clark et al. 2006, DeGraaf and Tyrrell 2004, Field 1922, Norberg and Tedengren 1995, Shumway and Stickney 1975), and ecosystem engineers that form complex mussel-bed habitats (Jones et al. 1994, Tsuchiya and Nishihira 1986). A positive relationship has been shown between habitat complexity and biotic diversity (Dean and Connell 1987). The diversity of organisms associated with mussel beds includes several epibionts, such as attached barnacles, hydroids, and algae (Suchanek 1978). Epibionts become more common on individual Blue Mussels as they age and the antifouling shell layer, or periostracum, sloughs off (Bers and Wahl 2004, Bers et al. 2006). This sloughing typically occurs after juvenile Blue Mussels move from filamentous algae, where they develop from plantigrade larvae, to mussel beds where they grow into the adult stage (Bayne 1964). The invasive ascidian Didemnum vexillum (Kott) (Carpet Sea Squirt) is an abundant invasive species in the Gulf of Maine that readily colonizes living organisms, including Blue Mussels, as an epibiont (Auker 2006, 2010; Auker and Oviatt 2008). This species likely came from Asia as an epibiont on Crassostrea gigas (Thunberg) (Pacific Oyster) imported to the Damariscotta River estuary in the 1950s (Dijkstra et al. 2007, Lambert 2009). Didemnum vexillum was first documented in the estuary in 1993, but anecdotal evidence indicated its presence since the late 1970s (USGS 2013; L.G. Harris, pers. observ.). Didemnum vexillum has also colonized other temperate coastal environments on the east coast of the United States, including Cape Cod (Carman and Grunden 2010) and Rhode Island (Auker and Oviatt 2008), and continues to spread worldwide (USGS 2013). Didemnum vexillum has been increasing in abundance throughout the Gulf for the past 10 years, forming large mat-like Northeastern Naturalist Vol. 21, No. 3 L.A. Auker, A.L. Majkut, and L.G. Harris 2014 481 growths on Blue Mussel beds, covering individual Blue Mussels completely (L. Auker, pers observ). Didemnum vexillum is in a family of ascidians (Didemnidae) that are known to possess antipredatory chemical defenses, either in the form of secondary metabolites (Blunt et al. 2006, Lindquist et al. 1992, Prado et al. 2004, Vervoort et al. 1998) or inorganic acids (Stoecker 1978, Stoecker 1980, but see Parry 1984). As a result, fouling by D. vexillum may reduce predation on organisms that they overgrow. There is concern that invasive species may reduce, or otherwise alter, the role of ecosystem engineers, which in turn may negatively impact the surrounding ecosystem (Crooks 2002). Blue Mussels in the Gulf of Maine are controlled by predators including introduced crab species, e.g., Hemigrapsus sanguineus (De Haan) (Asian Shore Crab) and Carcinus maenas L. (Green Crab) (DeGraaf and Tyrrell 2004, Tyrrell et al. 2006). If overgrowth of Blue Mussels by D. vexillum prevents predation on Mussels, this could have severe implications for both the mussels and the Gulf of Maine ecosystem. This paper will focus on how mussels are impacted by predation at two different life stages—plantigrade Blue Mussels that have no epibionts and adults that have been colonized by the invasive D. vexillum. The first goal of this study was to examine the impacts of predation on a life-history stage of Blue Mussels that have no epibionts, the plantigrade stage. Our second goal was to understand the impacts that D. vexillum has on predation by a common Gulf of Maine predator, Green Crab, and to determine whether the effect of D. vexillum overgrowth is one of associational resistance or shared doom. Methods Predator-exclusion experiment To test the effects of predation on newly settled plantigrade Blue Mussels, we designed and deployed collectors (artificial turf doormats covering 5-cmlong PVC pipes with a 2-cm diameter; modified from collectors used in Brenner and Buck 2010, Harris et al. 2004, Walter and Liebezeit 2003) with and without predator-exclusion cages. Exclusion panels consisted of five 50-cm-long artificial turf mussel-collectors covered with 5-mm mesh. Control panels consisted of 5 uncovered turf panels. Cage controls consisted of five 5-mm mesh-covered turf panels with large openings (approximately 200 cm2) cut in the mesh. We deployed collector panels at the Hampton River Marina in Hampton, NH, off the side of a floating dock where the water depth ranges 1.5–2.1 m, the average salinity ranges 29–31 psu (Deacon and Nash 2002), and the average water temperature was 1.7–2.8 °C (NOAA 2014). We suspended the panels about 1 m below the dock from 25 February to 17 March 2012. After 3 weeks, we retrieved and disassembled the cages, and thoroughly rinsed the turf panels with tap water in order to remove organisms present on the panels. We placed all objects removed from the turf into a gridded petri dish, and identified and counted all organisms. Our data satisfied the assumptions for normality and equal variances, so we used a one-way analysis of variance (ANOVA) and Tukey’s post-hoc test to determine if there were differences in newly Northeastern Naturalist 482 L.A. Auker, A.L. Majkut, and L.G. Harris 2014 Vol. 21, No. 3 settled (postlarval) Blue Mussel abundance among treatments. We also scored the types and numbers of predators present, and we used a one-way ANOVA to determine differences in predator presence among treatments. Epibiont experiment We collected Blue Mussels (mean shell length = 26.4 mm, SD = 3.1 mm) and D. vexillum from underneath floating docks in New Castle, NH. We collected female Green Crabs (mean carapace width = 37.1 mm SD = 5.2 mm) from intertidal areas in New Hampshire and Rhode Island. We kept all animals except D. vexillum in a closed, temperature-controlled (15 °C) system at the University of New Hampshire. We fed the Green Crabs a maintenance diet of Blue Mussels every 2–3 days until one week before the experiments, at which time we stopped feeding them. We collected D. vexillum from floating docks immediately before all experiments to ensure the organisms’ optimal health. Our experiments took place from late summer to mid-autumn when D. vexillum is metabolically active in the Gulf of Maine (Dijkstra 2007). Handling time and prey selection. For each trial, we filled a large basin (34 cm x 43 cm x 11.5 cm deep) with sea water (salinity = 32 psu), placed a Sony® Handycam DCR-SR47 digital video camera on a tripod (55.5 cm to base of camera), and aimed it at the basin. We placed 1 Blue Mussel with 90–100% D. vexillum cover and 1 clean Mussel without D.vexillumcover on opposite corners at the far end of the basin from the camera. Recording began as soon as a naïve Green Crab was placed in the basin. Once we added the Green Crab, the set-up was left undisturbed and the Green Crab was allowed to explore the basin at will for 30 minutes. At the end of the 30-minute period (one trial), we recorded the type of Blue Mussel ultimately consumed—overgrown or clean. If both Blue Mussels were consumed, then we recorded the outcome as consumed. We completed 29 trials, all of which were conducted during the day. We played back videos in a VLC Media Player (VideoLAN 2009) and recorded measurements for each of the following variables for each trial: (1) initial selection of Mussel as indicated by which the Green Crab first approached; (2) handling time for each clean and each overgrown mussel, and for D. vexillum alone (if it was removed from the Blue Mussel); and, (3) final Blue Mussel selection as indicated by Blue Mussel consumed. Our data satisified the assumptions for normal distribution and equal variances; thus, we conducted a one-way ANOVA and Tukey’s post-hoc test to determine if significant differences existed in handling time. Consumption. For each trial, we filled two 10-gallon aquaria with seawater. One aquarium contained 30 Blue Mussels free of epibionts. The second aquarium contained 30 Blue Mussels that were overgrown with D. vexillum. We added 6 Green Crabs to each aquarium and left them undisturbed for 24 hours at 15 °C. After 24 hours, we removed the Green Crabs, isolated them in their respective groups, counted the Blue Mussels consumed in each aquarium, and replenished each aquarium with Blue Mussels to the original sum of 30 individuals. To see if the Green Crabs limited their feeding due to satiation or because they were deterred by the overgrowth of D. vexillum, we placed the Green Crabs in the treatment tanks opposite the one Northeastern Naturalist Vol. 21, No. 3 L.A. Auker, A.L. Majkut, and L.G. Harris 2014 483 in which they fed in the previous 24 hours (i.e., control crabs were placed with overgrown mussels, and vice versa). We again left these crabs to feed for 24 hours undisturbed. At the end of this second period, we counted the number of Blue Mussels consumed. We completed 5 trials and used new Green Crabs for each trial. We conducted a paired t-test on the proportion of clean and overgrown Blue Mussels consumed by each group of crabs. Results Predator exclusion We observed significantly more Mussel plantigrades on exclusion panels than on control or cage control panels (F2,12 = 6.78, P = 0.011; Fig. 1). A Tukey’s HSD post-hoc test showed that there were significant differences between the cage control treatment and the exclusion treatment (P = 0.015) and between the control treatment and the exclusion treatment (P = 0.027). The Figure 1. Effects of predator exclusion on settlement of Blue Mussel plantigrades in Hampton, NH, in spring 2012. Error bars represent ± 1 standard error. The control cage (n = 5) and no cage (control) treatments (n = 5) showed similar results (P = 0.95), indicating that the cage construction had little effect on Blue Mussel settlement on the turf panels. The panels in which predators were excluded (n = 5) showed significantly more settlement of larvae (F2,12 = 6.78 P = 0.011), indicating that predators may be having a significant impact on Blue Mussel survival at an early stage of the bivalve’s life-history. Northeastern Naturalist 484 L.A. Auker, A.L. Majkut, and L.G. Harris 2014 Vol. 21, No. 3 control and cage control treatments showed no significant differences in postlarval abundance (P = 0.95). Therefore, the cage did not impact predation on the postlarval mussels. Mussel predators found on the panels in the cage control and control treatments were Green Crabs and Nucella lapillus L. (Dog Whelk) (Fig. 2). Predators were abundant on both the control and cage control panels but rare on the exclusion panels (F2,12= 3.411, P = 0.067). Green Crabs (n = 1) and Atlantic Dog Winlkes (n = 5) found on the exclusion panels were small enough to enter the mesh surrounding the panel. Figure 2. The number of predators found on turf panels in Hampton, NH, in spring 2012. Error bars represent ± 1 standard error. The predators found were the Green Crab (crabs) and the Dog Whelk (snails). The control (n = 5) and cage control treatments (n = 5) were similar in having the greatest number of predators per panel upon retrieval. While the exclusion cages (n = 5) had fewer or no predators, the difference between these panels and the control panels was not significant, due to a large amount of variation among the panels (F2,12= 3.411, P = 0.067). Northeastern Naturalist Vol. 21, No. 3 L.A. Auker, A.L. Majkut, and L.G. Harris 2014 485 Handling time and selection experiments More overgrown Blue Mussels were initially approached in selection experiments, but more clean Blue Mussels were ultimately consumed; some overgrown mussels (5 out of 29) were also consumed (Table 1). Green Crabs that initially approached clean Blue Mussels only consumed clean Mussels, whereas Green Crabs that approached overgrown Blue Mussels consumed either clean or overgrown Blue Mussels (Table 2). In 13 trials, Green Crabs approached the overgrown Blue Mussel first. However, instead of attacking the Blue Mussel immediately (which Green Crabs did when they approached clean Blue Mussels), the Green Crabs picked up D. vexillum and moved it towards its own mouthparts as if it were assessing the ascidian. The ascidian was never actually consumed, because within seconds after picking it up, the Green Crab discarded it The result of this action was that the Green Crab removed the ascidian from the Blue Mussel, creating a Blue Mussel identical to the control, or clean, Blue Mussel. Every consumed Blue Mussel that was initially overgrown had its ascidian epibiont removed by the Green Crab. Green Crabs in 9 trials of our study showed no interest in either Blue Mussel during either initial approach or final consumption. In 5 of these trials, they only responded to the ascidian epibiont. In one of these trials, the Green Crab spent 39 s handling D. vexillum, and the rest of the trial attempting to escape the basin. In other cases, the Green Crab spent from 12–161 s handling the ascidian, while ignoring the Blue Mussels for the rest of the trial. In 4 trials, the Green Crab showed no response to either Blue Mussel or D. vexillum. The average time Green Crabs spent handling clean Blue Mussels (mean = 417 s, SD = 563 s) was not significantly different than time spent handling overgrown Blue Mussels (mean = 251 s, SD = 425 s) (t56 = 1.265, P = 0.211; Fig. 3). An average of 62 s (SD = 96.4 s) was spent handling just D. vexillum; this was significantly less handling time than for either the clean or overgrown Blue Mussel (F2,85 = 3.269, P = 0.043). Consumption experiments Crabs consumed more clean Blue Mussels than overgrown Blue Mussels (Fig. 4). A paired t-test indicated a significant difference between the proportion of clean Blue Mussels consumed in the first 24 hours and the proportion of overgrown Blue Mussels consumed in the second 24 hours by the same crab group (t4 = 3.328, P = 0.029). Green Crabs that were initially placed with overgrown Blue Mussels in the first 24 hours of the experiment consumed significantly more clean Blue Mussels in the second 24 hours (t4 = 3.766, P = 0.020). Green Crab behavior in this experiment was consistent in each trial. Green Crabs placed in tanks with overgrown Blue Mussels typically pulled off D. vexillum before consuming the Blue Mussel. If the ascidian was growing in such a way that it was difficult to remove, then that Blue Mussel was generally ignored. Many of the untouched Blue Mussels in the overgrown tanks were heavily covered with the ascidian. Northeastern Naturalist 486 L.A. Auker, A.L. Majkut, and L.G. Harris 2014 Vol. 21, No. 3 Table 2. Outcomes of Green Crab initial selection of clean or overgrown Blue Mussels. Crabs initially approaching clean mussels finally consumed clean mussels or no mussels, whereas crabs that initially approached overgrown mussels finally consumed nothing, clean mussels, overgrown mussels, or assessed the ascidians for an extended period of time. Numbers indicate the total number of Green Crabs observed. Some crabs consumed both mussels during their trial (n = 3 [10.3%]). Crab choice Nothing Clean mussel Overgrown mussel Ascidian only Crabs initially approaching clean mussels finally consumed 2 (6.9%) 6 (20.7%) 0 (0.0%) 0 (0.0%) Crabs initially approaching overgrown mussels finally consumed 8 (27.6%) 3 (10.3%) 4 (13.8%) 1 (3.4%)* *The ascidian was not completely consumed, although observations indicate the Green Crab in one trial held the ascidian to its mouthparts for a prolonged period of time for a taste. Table 1. Green Crab selection of clean and overgrown Blue Mussels. Initial approach refers to the first Blue Mussel the Green Crab contacted during the trial. Final choice means the type of mussel consumed by the end of the trial. Numbers indicate the total number of Green Crabs observed. Some crabs consumed both mussels during their trial (n = 3 [10.3%]). Crab choice Nothing Clean mussel Overgrown mussel Ascidian only Initial approach 4 (14.8%) 8 (27.6%) 13 (44.8%) 5 (17.2%) Final choice 18 (62.1%) 9 (31.0%) 5 (17.2%) 1 (3.4%)* *The ascidian was not completely consumed, although observations indicate the Green Crab in one trial held the ascidian to its mouthparts for a prolonged period of time for a taste. Northeastern Naturalist Vol. 21, No. 3 L.A. Auker, A.L. Majkut, and L.G. Harris 2014 487 Discussion Predators appear to have a significant impact on Blue Mussels at an early life stage in which no epibiosis occurs. Postlarval Blue Mussels that are newly settled on filamentous substrate during primary settlement lack epibionts; this age class of Blue Mussels possesses a periostracum that has antifouling properties and keeps the Blue Mussel clear of epibionts (Bers and Wahl 2004, Bers et al. 2006). Later, this periostracum wears down and disappears, allowing epibionts to settle on the mussel shell. In our experiments, we found significantly more Blue Mussel plantigrades on predator-exclusion panels than on control or cage control panels after 3 weeks. Thus, predators appear willing and capable of consuming unfouled plantigrades, contributing to increased mortality at this life stage. These results support the findings of previous studies that have found that predation is the single-most important source of natural mortality in Mytilus (Seed Figure 3. Mean handling time of clean Blue Mussels and those overgrown by D. vexillum (Carpet Sea Squirt) by the Green Crab in a laboratory experiment (n = 29). Error bars represent ± 1 standard error. There was no significant difference in handling time between clean Blue Mussels and those overgrown with D. vexillum in this study (t56 = 1.265, P = 0.211), though the least amount of time was spent on handling Blue Mussels overgrown with D. vexillum (F2,28 = 3.269 P = 0.043) . n = 16 trials in which clean Mussels were handled, n = 16 trials in which overgrown Blue Mussels were handled, and n = 14 trials in which D. vexillum was handled. Northeastern Naturalist 488 L.A. Auker, A.L. Majkut, and L.G. Harris 2014 Vol. 21, No. 3 and Suchanek 1992). Our study assessed predation on very small (less than 1 mm) Blue Mussels, and several previous studies have focused on predator preference for different size classes of Blue Mussels. For example, Asterias forbesii L. (Sea Star) prefers Blue Mussels less than 70 mm in shell length (Campbell 1983). The Green Crab has shown a similar preference in previous studies (Elner and Hughes 1978, Juanes 1992). Strongylocentrotus droebachiensis (Müller) (Sea Urchin) preys on Blue Mussels less than 16 mm in length (Briscoe and Sebens 1988), and Somateria mollissima L. (Eider Duck) prefers Blue Mussels between 10–25 mm long (Raffaeli et al. 1990). The few predators found on our exclusion panels were small enough to fit through the mesh. Interestingly, the most common predator we identified on the exclusion panels was the predatory Atlantic Dog Winkle. This snail is consumed by Green Crabs (Hughes and Elner 1979) and its relative abundance on the exclusion panel may have been due to the near-complete exclusion of Green Crabs from these Figure 4. Effect of overgrowth of Didemnum vexillum on Blue Mussel consumption by Green Crabs in the laboratory. Error bars represent ± 1 standard error. Group 1 Green Crabs were fed clean Blue Mussels in the first 24 hours of the experiment, and then overgrown Blue Mussels in the second 24 hours of the experiment. They consumed significantly fewer overgrown Mussels (t4 = 3.328, P = 0.029). Group 2 Green Crabs were fed overgrown Blue Mussels in the first 24 hours, then fed clean Blue Mussels in the second 24 hours. These crabs also consumed significantly fewer overgrown Blue Mussels (t4 = 3.766, P=0.020). n = 5 separate trials for each group. Northeastern Naturalist Vol. 21, No. 3 L.A. Auker, A.L. Majkut, and L.G. Harris 2014 489 panels. The significantly greater quantities of Blue Mussels found on the exclusion panels compared to the controls indicate that the low density of small Atlantic Dog Winkles found on the predator-exclusion panels had little to no effect on Blue Mussel plantigrade numbers. In our study, because the Blue Mussel panels were left undisturbed for 3 weeks, we cannot determine exactly which predators fed on the Blue Mussels present on our panels. However, because we found both Dog Whelks and Green Crabs on our cages when we retrieved them, we assumed these were some of the common predators active during this period of time. Although we also cannot verify that all Blue Mussels on the non-exclusion panels were actually consumed and not dislodged (without observing the panels for the full 3 weeks), we can assume that any potential physical contact with the panel itself did not dislodge significant numbers of plantigrades. Plantigrade Blue Mussels strongly attach to their substrate, and considerable effort is required to remove them from panels (L.A. Auker, pers. observ.). The protected cage control panels and the exposed control panels were also statistically similar in the abundance of Blue Mussels found on each; therefore, we were assured that removal from panels is most likely due to predation rather than removal due to dislodging. In consumption assays, Green Crabs consumed fewer overgrown Blue Mussels than control Blue Mussels, suggesting an associational resistance effect of D. vexillum epibiosis on Blue Mussels. Wahl et al. (1997) identified 4 stages of predator activity: encounter, recognition, capture-handling, and consumption. They hypothesized that epibiosis only affects recognition and capture-handling. In our study, more overgrown Blue Mussels than control Blue Mussels were approached first, and the ascidian did not instantly repel the predator; therefore, encounter was not affected. However, the consumption stage was negatively affected because Green Crabs consumed more clean Blue Mussels. Didemnum vexillum deterred Green Crab predation indicating that the symbiont provides an associational resistance to predation, a positive aspect of epibiosis for the Blue Mussel in terms of providing a refuge from predation. It is unclear what specifically deterred predation, although D. vexillum possesses an acidic tunic and may possess additional chemical defenses. When D. vexillum’s tunic is disturbed, surface-test cells break apart and release acid (S. Bullard, University of Hartford, West Hartford, CT, pers. comm.). However, Parry (1984) suggested that this acid is quickly neutralized by calcium spicules in the test or is buffered by seawater. The Green Crabs in our choice experiment picked up and handled D. vexillum for relatively long time periods (up to 161 s), so it is unclear if the Green Crab was affected by the release of any acid. In the consumption experiments, Green Crabs that were placed in tanks with control Blue Mussels ate more Blue Mussels than Green Crabs placed in tanks with overgrown Blue Mussels. This proved true for both Green Crabs that were initially placed with clean Blue Mussels and for Green Crabs initially placed with overgrown Blue Mussels; in all trials, the presence of D. vexillum reduced Blue Mussel consumption. This result supports earlier studies that have shown that chemical extracts from members of the family Didemnidae Northeastern Naturalist 490 L.A. Auker, A.L. Majkut, and L.G. Harris 2014 Vol. 21, No. 3 contain predator deterrents (Blunt et al. 2006, Lindquist et al. 1992, Prado et al. 2004, Stoecker 1978, 1980, Vervoort et al. 1998, Wright et al. 2002). Our results for the selection and consumption laboratory studies indicated that D. vexillum decreases predator selection for the Blue Mussel (decreased handling time, decreased final choice, and decreased overall consumption). The selection experiments allowed observation of individual Green Crab behavior, but contained chemical cues of both the Blue Mussel and ascidian, whereas the consumption experiments provided a free-for-all scenario in which Green Crabs were given a greater amount of prey of one type. In these experiments, the clean Blue Mussel tanks contained Blue Mussel chemosensory cues, but the overgrown Blue Mussel tanks contained both ascidian and mussel cues. Based on our findings from the selection experiment, that handling time was not significantly decreased in overgrown Blue Mussels, the results from the consumption study indicate that the presence of the D. vexillum chemosensory cues have an impact on Blue Mussel consumption. The anti-predator resistance provided by D. vexillum to Blue Mussels may vary with time of year. We used D. vexillum and Blue Mussels covered with the ascidian collected within one to two days of the feeding trials for our study. During the winter months, D. vexillum senesces and several potential predator species have been observed feeding on the ascidian (Valentine et al. 2007). At this time, D. vexillum may not provide any resistance to potential Blue Mussel predators; the ascidian may even provide an additional source of food for predators, potentially resulting in a shared doom scenario for Blue Mussels, in which predators are attracted to an epibiont, and the basibiont is consequently consumed (Wahl et al. 1997). Blue Mussel populations are controlled by several predators in the Gulf of Maine, including Atlantic Dog Winkle, Sea Stars, and several native and invasive species of crabs (Bordeau and O’Connor 2003, Seed and Suchanek 1992). If D. vexillum reduces predation on Blue Mussels through associational resistance as indicated in this study, several predatory species may be negatively affected. In the top-down predator-controlled systems seen in our study area (Donahue et al. 2009), community dynamics could be affected by this associational resistance (Wahl et al. 1997). For example, predators would consume fewer Blue Mussels when the latter are overgrown, and resort to other species for food, or the predator populations may decrease due to lack of food. Areas covered in these large mat-like D. vexillum colonies would likely be most dramatically impacted. Although our study focused on one predator, the Green Crab, we previously attempted this experiment with Sea Stars in the laboratory and did not observe any instance of their consuming Blue Mussels. Expanding this study into the field to look at other common predators’ preferences would be beneficial in understanding the implications of overgrowth by a dominant invasive ascidian. As for the individual Blue Mussel, the associational resistance effect from overgrowth provides a trade-off for negative effects on growth. During the time of year in which D. vexillum is most abundant, gamete production, shell-lip thickness, and tissue production decrease in Blue Mussels (Auker 2010). The Blue Northeastern Naturalist Vol. 21, No. 3 L.A. Auker, A.L. Majkut, and L.G. Harris 2014 491 Mussel may not grow as quickly when overgrown, but will likely be protected from predation; this tradeoff may be particularly advantageous for Blue Mussels that colonize the benthos and have both benthic (e.g., Sea Stars and crabs) and pelagic (Tautogolabrus adspersus [Wahlbaum] [Cunner] and other fish species) predators. Although Laudien and Wahl (2004) predicted that the decrease in growth of Blue Mussels caused by an epibiont may prolong its susceptibility to predation because smaller Blue Mussels are preferred over larger Blue Mussels (e.g., Murray et al. 2007), our results suggest otherwise. Epibiotic D. vexillum’s mat-like morphology, which tends to overgrow Blue Mussels completely and deter predators, protects small Blue Mussels (less than 5 cm) from predation. Overall, the results from this study suggest that predation is a significant factor in plantigrade Blue Mussel-population abundance, a Blue Mussel life stage that does not yet possess epibionts. We also observed that D. vexillum has a positive effect on Blue Mussels by providing an anti-predator defense against the Green Crab, a common predator in the Gulf of Maine. Protection from predation could have strong implications for predators in areas dominated by D. vexillum growth and has the potential to alter community dynamics in the Gulf of Maine. Acknowledgments The Hampton River Marina (Hampton, NH) provided floating-dock space for the deployment and retrieval of our predator-exclusion panels. We wish to thank Emily Gamelin for help in collecting animals for the laboratory experiments. 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