Effects of Invasive Colonial Tunicates and a Native Sponge on the Growth, Survival, and Light Attenuation of Eelgrass (Zostera Marina)

Aquatic Invasions (2012) Volume 7, Issue 3: 315–326 doi: http://dx.doi.org/10.3391/ai.2012.7.3.003 Open Access

Research Article

Effects of invasive colonial tunicates and a native sponge on the growth, survival, and light attenuation of eelgrass (Zostera marina)

Melisa C. Wong* and Bénédikte Vercaemer Fisheries and Oceans Canada, Bedford Institute of Oceanography, Dartmouth, NS, B2Y 4A2, Canada E-mail: [email protected] (MCW), [email protected] (BV) *Corresponding author

Received: 13 December 2011 / Accepted: 5 March 2012 / Published online: 29 March 2012

Handling Editor: Mark Hanson, Fisheries and Oceans Canada, Canada

Abstract

We examined the effects of invasive colonial tunicates (golden star, Botryllus schlosseri; violet Botrylloides violaceus), and the native breadcrumb sponge (Halichondria panicea) on the growth, survival, and light attenuation of eelgrass (Zostera marina). Eelgrass shoot growth and survival were higher for unfouled shoots than for fouled shoots, and dependent on fouling species identity. Growth was lowest for shoots with violet tunicate fouling, and survival was lowest for shoots with sponge fouling. A large proportion (0.20-0.38) of fouled shoots marked for growth measurements were not found during retrieval compared to unfouled plants (0.08), suggesting that fouling led to premature breaking away of shoots and blades. Transmission of incident photosynthetically active radiation (PAR) through fouled blades decreased exponentially with increasing biomass of most fouling species. Relatively low biomass (1-1.5 dry mg cm-2 of blade) of the fouling organisms reduced light transmission by up to 98 % compared to unfouled blades. The proportion of incident PAR attenuated by the sponge and orange and burgundy morphs of the violet tunicate increased hyperbolically across their biomass, and reached a plateau at ~0.5 dry mg cm-2 of blade. The golden star tunicate and cream morph of the violet tunicate attenuated incident PAR linearly across their biomass. For the range of biomass examined, all fouling species attenuated 65-95 % of incident PAR prior to it reaching the blade. The reduction in light transmission was likely the causal mechanism underlying reduced growth of fouled shoots. Fouling of eelgrass by the invasive colonial tunicates and the native sponge will have numerous ecological consequences, including reduced productivity and coverage of eelgrass beds. Key words: ascidians, Botryllus schlosseri, Botrylloides violaceus, photosynthetically active radiation, Halichondria panicea, seagrass

Introduction coverage because enhanced biomass of phytoplankton, epiphytes, macroalgae, and Seagrass beds are valuable coastal habitats that suspended solids shade and smother seagrass provide numerous important ecological functions plants (Hauxwell et al. 2003; Kemp et al. 2005; and services, including organic carbon Walker et al. 2006). Disease, dredging, vessel production, enhanced biodiversity, nutrient grounding, and threats from global climate cycling, and nursery habitat (Heck et al. 1995; change (e.g., increased sea surface temperature Duarte and Chiscano 1999; Heck et al. 2003; and storm frequency, sea level rise) also have Orth et al. 2006; Schmidt et al. 2011). Over the demonstrable negative impacts on seagrass past century, global seagrass coverage has coverage (Short and Wyllie-Echeverria 1996; experienced widespread declines (Short and Short and Neckles 1999; Orth et al. 2006). Wyllie-Echeverria 1996; Orth et al. 2006, Additionally, new mechanisms of seagrass Waycott et al. 2009), with global loss of areal decline continue to emerge. In particular, recent coverage outpacing gains by more than tenfold reviews suggest that invasive species may be an (Waycott et al. 2009). Multiple stressors on the important factor leading to seagrass loss (Orth et global, regional, and local scales are responsible al. 2006; Williams 2007). for loss of seagrass meadows (Short and Wyllie- The rate of reported species invasions in Echeverria 1996; Duarte 2002; Orth et al. 2006; marine ecosystems have increased exponentially Waycott et al. 2009). Degradation of water over the last two centuries (Ruiz et al. 2000). In quality is well documented to reduce seagrass seagrass ecosystems, at least 56 non-native

315 M.C. Wong and B. Vercaemer species have been introduced worldwide panicea (Pallas, 1766)] (Wong pers. obs.). (Williams 2007). The majority of invasive Invasive colonial tunicates may thus have species in seagrass beds are invertebrates (63 %), important ecological consequences that did not and include cnidarians, ascidians, molluscs, exist previously for eelgrass beds in some areas polychaetes, crustaceans, and ectoprocts of the Maritime provinces. To better understand (Williams 2007; Carmen and Grunden 2010). the effect of fouling on eelgrass by these species, Despite the prevalence of invasive invertebrates, we determined (i) the growth, survival, and most studies of invasive species in seagrass beds chlorophyll a content of unfouled eelgrass blades have focused on effects of non-native and those fouled with the invasive colonial macroalgae (Williams 2007; e.g., den Hartog tunicates (golden star tunicate and violet 1997; Ceccherelli et al. 2000; Eklöf et al. 2005; tunicate) and the native breadcrumb sponge, (ii) Drouin et al. 2011). The few studies of invasive the reduction in transmission of incident PAR invertebrates in seagrass beds have found that through fouled blades across a gradient of they have predominantly negative effects on fouling organism biomass, and (iii) the seagrass coverage either directly through plant attenuation of incident PAR by the fouling fouling, or indirectly, through feeding or habitat organisms themselves across a gradient of their formation. For example, invasive anemones biomass. Using this knowledge, we discuss the (Bunodeopsis sp. Andres, 1881) and bryozoans potential ecological consequences of the invasive [Zoobotryon verticillatum (Della Chiaje, 1822)] colonial tunicates and native sponge for eelgrass can foul eelgrass (Zostera marina Linnaeus, beds that usually experience minimal epiphyte 1758), reducing light reaching the blades and fouling. weighing blades down causing canopy collapse (Sewell 1996). Invasive green crabs [Carcinus Materials and methods maenas (Linnaeus, 1758)] can reduce the survival of transplanted eelgrass shoots through Study site and organisms feeding and bioturbation (Davis et al. 1998), while high abundances of invasive tube-building Field measurements were conducted in August polychaetes [Neodexiospira brasiliensis (Grube, and September 2010 at an eelgrass bed in Port 1872)] may reduce seagrass growth (Critchley et l’Hebert, on the south shore of Nova Scotia, al. 1997). Canada (GPS coordinates: N43.86805, Seagrass beds in the Canadian Maritime W64.96280). The eelgrass bed was located in the provinces are primarily composed of eelgrass inner portion of the bay, which is characterized (Zostera marina), and are often characterized by by low water currents and high sediment well established populations of invasive deposition (M. Wong, unpub. data). Percent silt invertebrates. The three most common invasive content and organic content of sediments was species are the European green crab (Carcinus 52.8 and 18.5%, respectively, and was maenas), the colonial golden star tunicate determined from 6 replicate samples processed [Botryllus schlosseri (Pallas, 1766)], and the as in Bale and Kenny (2005) and Wong et al. colonial violet tunicate [Botrylloides violaceus (2011). The bed was a monospecific stand of (Oka, 1927)] (Wong unpubl. data). Tunicate eelgrass, with shoot densities of ~1000 shoots fouling of eelgrass blades likely has effects m-2 and percent cover of ~90 %, determined from similar to those from epiphyte fouling: the 10 randomly placed quadrats (0.5m × 0.5m) amount of photosynthetically active radiation during the experimental period. Water depth at (PAR) reaching the blade will be reduced, and mean low water ranged from 0 to 0.35 m, and at blades will be weighed down, leading to self mean high water from 1.0 to 1.5 m. Water shading and breaking away of shoots (Bulthuis temperature during the field work ranged from and Woelkerling 1983; Brush and Nixon 2002). 15 to 30°C. Tunicate fouling would thus result in reduced Invasive colonial tunicates and a native shoot growth, photosynthesis, and shoot survival sponge were commonly observed fouling (Short et al. 1995; Moore and Wetzel 2000), with eelgrass blades. Tunicates were the golden star the degree of impact dependent on tunicate tunicate and violet tunicate. The golden star biomass. Although eelgrass beds on the Atlantic colour morph was brown. The violet tunicate was coast of Nova Scotia are rarely heavily fouled by present in 3 colour morphs: orange, burgundy, epiphytes, blades are sometimes fouled by the and cream. No actual violet colour morphs were native breadcrumb sponge [Halichondria observed. The native breadcrumb sponge was

316 Invasive tunicates in seagrass beds pale yellow, and formed smooth thin crusts on Light attenuation of fouled and unfouled eelgrass the eelgrass blades. Other fouling epifauna or blades epiphytes on eelgrass blades were usually not We measured the transmission of photo- observed during the field work or at other times synthetically active radiation (PAR) through of year (M. Wong, pers. obs.). fouled and unfouled eelgrass blades using an underwater quantum sensor (LICOR, LI-192) Effects of tunicates and sponge on eelgrass and data logger (LICOR, LI-1400). Unfouled growth and survival blades of eelgrass and blades with varying Growth of fouled and unfouled eelgrass shoots degrees of tunicate or sponge fouling were was determined using the plastochrone method collected from the eelgrass bed by snorkeling. (Short and Duarte 2001). On three different dates On shore and under natural light, the light sensor between August and September 2010 (i.e., D1: was suspended directly under the water surface August 20, D2: September 2, D3: September 9), in a large plastic tank. Black electrical tape was unfouled shoots, and shoots with either sponge, used to adjust the width of the light sensor to be golden star tunicate, or one of the three violet similar to that of a typical eelgrass blade, tunicate colour morphs on the leaf blades were allowing the sensor to be fully covered when found by snorkelling at low tide. On each date, a determining light transmission through blades. large sewing needle was used to mark the shoots For measurements of the transmission of incident by making a small hole through all the leaves in PAR through each fouled blade, incident the middle of the sheath. Marked plants were irradiance at the water surface (I0) was first located >1 m apart to avoid marking plants on recorded (Figure 1). One side of the fouled blade the same rhizome. Marked plants were tagged was then gently scraped clean, the clean side using plastic tie-warps and flagged with surveyor submersed directly over the light sensor, and the tape to aid in recovery. Individually numbered irradiance transmitted through the blade + flags were inserted into the sediment next to the fouling organism (Transb,f) was recorded. marked plants to allow identification of the Scraping one side clean prior to measurements original fouling organism and the location of was necessary to directly measure light marked plants in case they broke free prior to transmitted through the blade. The fouling harvesting. organism that covered the top side of the blade Marked plants were harvested 19-22 days after was then removed by gently scraping and marking. On retrieval, the number of marked irradiance transmitted through the blade alone plants alive and missing was recorded. Retrieved (Transb) was recorded. The proportion of plants were transported to the laboratory and incident light transmitted through the fouled refrigerated in the dark until processing. Plants blade (i.e., blade + tunicate or sponge) was were rinsed in fresh water, each shoot taken calculated as Transb,f /I0. The proportion of apart at the meristem and the presence of fouling incident light attenuated by the fouling organisms and condition of the leaves (i.e., dead organisms themselves (Attenf /I0) was calculated or alive) was recorded. To determine leaf growth as (I0 – Transb,f – Attenb)/I0, where Attenb is light per shoot, the number of new, unmarked leaves attenuated by an unfouled blade (Figure 1b). The was counted, and the dry weight of the full effect of scraping on light transmission was length third leaf was determined by drying at determined by comparing transmission through 60°C for 24 hours. The leaf plastochrone interval non-fouled blades before and after scraping. (i.e., time required to produce a new leaf) was Fouled blades with patchy cover by the fouling calculated by dividing the growth period by the organism were not used. number of new leaves produced. Leaf growth The tunicate or sponge removed for measure- was then calculated for each marked shoot as: ments of Transb was placed in ethanol, and later Leaf growth (mg shoot-1 d-1) = weight of 3rd leaf dried at 60°C for 24 hours to determine dry mass (mg shoot-1) / plastochrone interval (d) (Short per area of blade. The portion of the blade that and Duarte 2001). The number of shoots marked covered the light sensor was clipped, and frozen for each fouling species ranged from 10-26 on until processed for chlorophyll a content. each sampling date, and depended on our ability Chlorophyll a was extracted from the blade to find appropriate shoots. Violet tunicates were samples using 90% acetone, and chlorophyll a not observed until the end of August, and thus per area of blade determined by the were only marked on the last two marking dates. Welschmeyer technique (Welschmeyer 1994).

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Figure 1. Calculation of PAR attenuated by the fouling organisms, where (a) I0 – Attenb – Transb,f = Attenf, and (b) I0 – Transb = Attenb. I0 = incident PAR, Attenf = PAR attenuated by the fouling organism, Attenb = PAR attenuated by the blade, Transbf = PAR transmitted through the fouled blade, Transb = PAR transmitted through the unfouled blade. Parameters with an asterisk indicate those directly measured with light sensor.

continuous independent variable (i.e., covariate), Statistical analyses and the proportion of incident PAR transmitted as the dependent variable. A significant To examine the effect of different fouling interaction term between the categorical and species on shoot growth, two-way analyses of continuous (covariate) predictor variables would variance (ANOVAs) with fouling species and indicate that the regression coefficients (i.e., marking date as fixed factors and leaf growth per slopes) differ among groups (Quinn and Keough shoot as the dependent variable were used. 2002), meaning that transmission of incident Because violet tunicates were not observed on PAR differs with fouling species (Quinn and the first marking date, two different ANOVAs Keough 2002). The effect of blade scraping on were conducted. The first ANOVA used 3 levels incident PAR transmission was determined using of fouling species (none, golden star, sponge) a one-way ANOVA, with blade scraping (3 and 3 levels of marking date (D1, D2, D3). The levels: no scrape, 1 scrape, 2 scrapes) as the second ANOVA used 4 levels of fouling species independent variable and proportion of incident (none, golden star, sponge, violet) and 2 levels PAR transmitted as the dependent variable. of date (D2, D3). The three colour morphs of To determine the relationship between light violet tunicate were pooled to provide enough attenuation by the sponge or each tunicate data for the analyses. Type III sums of squares species across their respective biomass gradient, based on unweighted marginal means were used we used non-linear least squares regression to fit to account for the unequal number of replicates the hyperbolic function: per fouling species (Quinn and Keough 2002). We used linear least-squares regression to Atten aM determine the relationship between the f  proportion of incident PAR transmitted through I0 1 bM the fouled blades with biomass of the fouling organism. The proportion of incident PAR where Attenf / I0 = the proportion of incident transmitted was log10 transformed to linearize the PAR attenuated by the fouling species, M = dry data which showed exponential decay with mass of the fouling species, and a and b are tunicate or sponge biomass. Tunicate and sponge parameters. biomass were standardized by area of blade Linear least squares regression was used to covering the light sensor. Regressions were fit to determine the relationship between chlorophyll a data for each fouling species separately. Separate concentration in fouled blades with tunicate or regression lines were compared using ANCOVA, sponge biomass. Because no clear relationship with fouling species as the categorical existed, a one-way ANOVA with fouling species independent variable (5 levels: sponge, golden (6 levels) as the independent variable and star tunicate, or the 3 violet tunicate colour chlorophyll a concentration as the dependent morphs), fouling organism biomass as the variable was conducted.

318 Invasive tunicates in seagrass beds

interactions were examined using Tukey’s test. All statistical analyses were done using R v.2.8.1 statistical software.

Results

Field observations The golden star tunicate and the sponge were visible on eelgrass blades beginning in mid-July. All colour morphs of violet tunicates were observed later, beginning in late August. The distribution of tunicate colonies and the sponge was very patchy, with larger patches becoming more prevalent at the end of September. Tunicate colonies and the sponge grew on single eelgrass Figure 2. Growth (mean ± 1 SE) of unfouled eelgrass shoots and blades or spread across numerous blades of those fouled with invasive colonial tunicates or a native sponge. neighbouring plants, and often formed mats Marking occurred on 3 different dates (D1-D3) between August and September 2010. SG = unfouled seagrass, GS = golden star ranging from 5 to 30 cm in diameter. Both alive tunicate, SP = sponge, V = violet tunicate. Data were pooled and dead blades were present in the mats, and the across all violet tunicate colour morphs. ND = no data. n = 3-15. mats weighed the eelgrass blades down towards the sea bottom. Blades were sometimes fouled with several species of organisms, but these blades were not used in our light or growth measurements. Floating eelgrass detritus on the water surface and shore often contained blades fouled with the tunicates or sponge.

Effects of tunicates and sponge on eelgrass survival and growth The proportion of marked shoots that were retrieved and showed growth was highest for unfouled blades, followed in decreasing order by those fouled with golden star tunicate, violet tunicate, and sponge (Table 1). Large proportions (0.205-0.375) of fouled marked plants were not found during retrieval compared

to unfouled marked plants (0.086). The highest Figure 3. Chlorophyll a concentration (mean ± 1 SE) in unfouled proportion of marked plants retrieved that were eelgrass blades and those fouled with invasive colonial tunicates or a native sponge. SG = unfouled seagrass, GS = golden star dead were those fouled by sponge (0.333), while tunicate, SP = sponge, VB = burgundy morph of the violet the proportion of dead plants retrieved that were tunicate, VC = cream morph of the violet tunicate, VO = orange originally fouled by golden star tunicate (0.180) morph of the violet tunicate. n = 22-47. did not differ from unfouled plants (0.171). Interestingly, marked plants that were originally fouled on one or more blade were not always retrieved still fouled, and no marked plants that For all regressions, ANOVAs, and the were originally unfouled were retrieved fouled. ANCOVA, residual plots were examined to Eelgrass shoot growth was significantly determine if the underlying assumptions of affected by both fouling species and marking homogeneity of variance and normality were date (Figure 2). This effect was evident when the violated (Draper and Smith 1998). If violations ANOVA included 3 fouling species (none, occurred, they were corrected by transforming sponge, golden star tunicate) and 3 marking the data using the square root or log10 (Draper dates (square root transformed; fouling species: and Smith 1998). Significant main effects and F2,55 = 3.52, p = 0.037; date: F2,55 = 5.49, p =

319 M.C. Wong and B. Vercaemer

0.007), and when the ANOVA included the 4 PAR transmitted through the fouled blades fouling species and 2 marking dates (log10 ranged from 0.05 to 0.1. Comparison of transformed; fouling species: F3,50 = 8.28, p = regression lines indicated that the proportion of 0.008; date: F1,50 = 1.31, p = 0.021). For both incident PAR transmitted through fouled eelgrass analyses, shoot growth was significantly highest blades differed significantly among fouling for unfouled blades, followed by blades fouled species (interaction term: F4,165 = 6.06, p = by the sponge and golden star tunicate (Tukey’s 0.00014), with regression intercepts and slopes test, p < 0.05). Shoot growth was lowest for for blades fouled with sponge and all violet blades fouled by the violet tunicate when this tunicate morphs differing significantly (p < 0.05) fouling species was included in the analysis from those for golden star tunicate. Scraping of (Tukey’s test, p < 0.05). Growth of shoots fouled eelgrass blades to remove tunicates did not affect by golden star and sponge tended to be similar light transmission (F2,30 = 0.64, p = 0.534). (Tukey’s test, p > 0.05). Shoot growth for plants The proportion of incident PAR attenuated by marked on D1 and D2 was significantly higher the sponge and the orange and burgundy morphs than for plants marked on D3 (Tukey’s test, p < of the violet tunicate increased hyperbolically 0.05). with their dry biomass (Table 2b, Figure 5). For Chlorophyll a concentration of blades was the golden star tunicate and the cream colour significantly affected by fouling species (F5,214 = morph of the violet tunicate, this relationship 14.3, p < 0.0001; Figure 3), and was highest for tended to increase linearly. For all fouling blades fouled with the cream morph of violet species, the maximum proportion of incident tunicate and the unfouled eelgrass, intermediate PAR attenuated was 0.9 to 0.95 for the range of in blades with golden star tunicate and the biomass examined. For the three species fit with orange and burgundy morphs of violet tunicate, the hyperbolic model, the proportion of incident and lowest in blades with the sponge (Tukey’s PAR attenuated tended to plateau at test, p < 0.05). Chlorophyll a concentration was approximately 0.5 dry mg cm-2 blade. not significantly affected by scraping of the blades (F1,45 = 0.99, p = 0.324). Discussion

Light attenuation of fouled and unfouled eelgrass Field observations made during our study blades indicated that invasive colonial tunicates and a native sponge were the main fouling organisms The proportion of incident PAR transmitted of eelgrass at our study site. These fouling through eelgrass blades fouled with invasive organisms had negative effects on eelgrass colonial tunicates or the native sponge decreased growth, survival, and light transmission. Shoot significantly as fouling biomass increased for all growth of fouled plants was clearly reduced fouling species, except for the golden star compared to unfouled plants. Interestingly, this tunicate where the trend was only marginally effect was dependent on the identity of the significant (Table 2a, Figure 4). With the fouling species, with growth lowest for shoots exception of blades fouled by golden star fouled by the violet tunicate. Also, the effects of tunicate, the proportion of incident PAR the fouling organisms on shoot growth were transmitted decreased exponentially across dependent on the growing season of both the fouling organism biomass, and log10 transformed eelgrass and fouling species. In temperate data were used for the linear regressions. The regions, eelgrass growth is strongly influenced proportion of incident PAR transmitted through by seasonal fluctuations in ambient light and eelgrass blades fouled with golden star tunicate temperature (Robertson and Mann 1984; tended to decrease linearly with increased Hemminga and Duarte 2000). Eelgrass biomass, tunicate biomass. For unfouled eelgrass blades, density, and growth on the Atlantic coast of the proportion of incident PAR that was Nova Scotia are usually highest from late May transmitted was 0.30 ± 0.009 (mean ± SE, n = until the end of August, with declines beginning 33). Even blades fouled with minimal biomass by September (Robertson and Mann 1984; Wong (<0.20 dry mg cm-2 blade) of the sponge or unpub. data). Although tunicate zooids and tunicates transmitted less incident PAR sponges are likely present on eelgrass blades compared to unfouled blades (0.05 – 0.25). when shoot growth is at a maximum (Carver et When biomass of fouling organisms was >1.0 al. 2006), colonies are not large enough to detect dry mg cm-2 blade, the proportion of incident until later in the season when eelgrass growth is

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Table 1. Survival of marked unfouled and fouled eelgrass shoots pooled across all marking dates. Data for all colour morphs of the violet tunicate were also pooled. All plants that showed growth were also still alive.

No. of Proportion plants Species fouling marked Proportion plants Proportion plants Proportion shoots retrieved that shoots retrieved fouled retrieved dead plants missing marked grew None 35 0.000 0.743 0.171 0.086 Sponge 39 0.154 0.462 0.333 0.205 Golden star tunicate 50 0.040 0.600 0.180 0.220 Violet tunicate 24 0.250 0.542 0.083 0.375

Figure 4. Proportion of incident PAR transmitted (log10 transformed) through eelgrass blades fouled by invasive colonial tunicates or the native sponge across a gradient of their biomass. n = 23-44.

beginning to slow. Despite the incomplete The most likely mechanism underlying overlap in the major eelgrass growth season with reduced growth of fouled shoots was the heavy fouling by the tunicates and sponge, reduction of incident PAR reaching fouled effects of the fouling species on shoot growth blades. For most fouling species, the proportion were still detected. Tunicate and sponge fouling of incident PAR transmitted through the fouled persisted well into the late fall at the field site eelgrass blades decreased exponentially with (Wong unpub. data), and continued to influence increasing biomass. All fouling species eelgrass growth dynamics late in the growing attenuated high proportions of incident PAR season. prior to the light reaching the eelgrass blades.

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Figure 5. Proportion of incident PAR attenuated by the invasive colonial tunicates or the native sponge across a gradient of their biomass. n = 23-44.

Table 2. (a) Parameter estimates (± SE) and regression statistics for linear regressions of the proportion of incident PAR transmitted through fouled blades (log10 transformed) across a biomass gradient of the fouling organism. df1 = numerator df, df2 = denominator df. (b) Parameter estimates (± SE) for non-linear regressions of the hyperbolic equation for the proportion of incident PAR attenuated by the fouling organism across a gradient of their biomass.

2 Fouling species Coefficient Intercept Fdf1,df2 p Adjusted R

Golden star tunicate -0.14±0.08 -1.05 ± 0.05 3.361,41 0.074 0.053

Sponge -0.39 ± 0.09 -0.67 ± 0.06 17.41,31 0.0002 0.34

Violet tunicate – orange morph -0.41 ± 0.11 -0.71 ± 0.06 14.21,41 0.0005 0.24

Violet tunicate – burgundy morph -0.50 ± 0.09 -0.67 ± 0.04 30.31,21 <0.0001 0.57

Violet tunicate – cream morph -0.39 ± 0.08 -0.64 ± 0.05 23.11,30 <0.0001 0.42

Fouling species a b Sponge 32.4 ± 8.2 35.4 ± 9.6 Violet tunicate – orange morph 32.6 ± 6.8 35.2 ± 7.8 Violet tunicate – burgundy morph 33.0 ± 6.4 35.4 ± 7.4

322 Invasive tunicates in seagrass beds

The sponge and the orange and burgundy portion of the tunicate or sponge structure laid morphs of the violet tunicate attenuated incident across the light sensor (i.e., the amount of zooids PAR hyperbolically across increasing biomass. or calcareous structures vs. matrix covering the An increase in light attenuation that plateaus at sensor) would have influenced measurements of high biomass of the fouling species has also been light attenuation. It is unlikely that additional observed for epiphytes fouling blades of several measurements would have reduced this inherent seagrass species (as reviewed by Brush and variability. Tunicate zooids were often Nixon 2002; Alcoverro et al. 2004; Frankovich overlapping and differed in size, and structures and Zieman 2005). The exact form of the within the sponge integument were not visible to saturating function (i.e., plateau levels off or the naked eye. Thus, it was not possible to continues to increase slightly in height) can vary standardize measurements by the number of (Brush and Nixon 2002), and is influenced in zooids or sponge structures covering the sensor. part by how the data weight the regression When our measurements of light attenuation (Frankovich and Zieman 2005). In our study, a by the tunicates and sponge were compared to thickening of the tunicate colony or sponge at attenuation by erect epiphytes at similar biomass high biomass caused the plateau in light (as reviewed by Brush and Nixon 2002), the attenuation, which begins at ~0.5 dry mg cm-2 of encrusting organisms in our study attenuated blade. At this biomass, 85 - 95% of the incident higher proportions of incident PAR. A similar light is attenuated, so further thickening result was found by Cebrián et al. (1999) when contributes little to further PAR attenuation. This light attenuation of a red encrusting algae and an differs from studies of light attenuated by erect brown algae was directly compared. epiphytes, where the plateau in light attenuation Generally, 1 dry mg cm-2 of blade of seagrass at high biomass results because the excess plants epiphytes attenuates 10-50% of incident PAR tend to grow or float from the sides of the blade, (Brush and Nixon 2002), which is much less than and do not intercept incident PAR that can be the 90-95% attenuated by the same biomass of attenuated by the leaf (Brush and Nixen 2002). tunicates and sponge in our study. Even a In addition to a hyperbolic relationship, we also minimal amount of tunicate or sponge fouling on identified linear relationships between light eelgrass will substantially affect light attenuation attenuation and biomass of the golden star by the blades. The overall effect of reduced light tunicate and the cream morph of the violet will not only be influenced by biomass of the tunicate. Similar relationships have been fouling organisms, but also by their optical observed for bryozoans on eelgrass blades properties. Examination of their optical proper- (Glazer 1999). Linear relationships may indicate ties using a spectrophotometer would elucidate that a larger range in biomass should be sampled how the tunicates and sponge change the light to better capture the plateau in light attenuation spectrum reaching the eelgrass blades, and in at high fouling organism biomass. Clearly, the particular, would indicate if they act as neutral relationship between light attenuation and filter or if they are attenuating the red and violet biomass of fouling organisms is species specific, wavelengths most harvested by eelgrass for pho- and depends on how the species grow on the tosynthesis (Cummings and Zimmerman 2003). leaves as well as their physical structure. Fouling by the colonial tunicates and the Our measurements of light attenuation by sponge cause low light conditions for the fouled colonial tunicates and the sponge were highly plants as well as for neighbouring plants shaded variable compared to measurements of epiphyte by the tunicate or sponge mats. Shading by light attenuation in other studies (Cebrián et al. fouled plants or environmental variables such as 1999; as reviewed by Brush and Nixon 2002; suspended sediment can reduce seagrass Alcoverro et al. 2004; Frankovich and Zieman nutrients, growth, survival, and photosynthetic 2005). The irregular physical structure of the processes (Ralph et al. 2007; Biber et al. 2009; tunicates and sponge contributed to this Collier et al. 2009; Ochieng et al. 2010). variability. Colonial tunicates consist of a Eelgrass plants can acclimate to low light polysaccharide matrix in which individual zooids conditions by increasing chlorophyll content to are embedded in distinct patterns (Hirose et al. enhance photon capture efficiency (Cummings 1991; Carver et al. 2006), while the sponge and Zimmerman 2003; Ralph et al. 2007). skeleton is interspersed with siliceous and However, this mechanism is not completely calcareous spicules (Ruppert et al. 2003). The compensatory, because increased chlorophyll

323 M.C. Wong and B. Vercaemer content does not scale proportionally with density in the bed (Short et al. 1995; Moore and changes in light absorption (i.e., the “package” Wetzel 2000; Ralph et al. 2007). A reduction in effect; Dubinsky et al. 1986). In our study, photosynthetic rate also impairs root functioning, measurements of chlorophyll a content in because flow of oxygen to the belowground eelgrass blades fouled by the cream and orange components is lowered (Hemminga and Duarte morph of the violet tunicate may have indicated 2000). Light limitation may change the ratio of an early response to light reduction, because aboveground to belowground components to chlorophyll a content was similar to unfouled reduce the maintenance cost of non- blades yet higher than in blades with other photosynthetic tissues (Olesen and Sand-Jensen fouling species. However, this may also simply 1993). The degree of impact of reduced reflect the later settlement and growth of violet photosynthesis on aboveground and belowground tunicate colonies compared to the other fouling plant components will depend in part on the species. Chlorophyll a content in blades fouled amount of carbohydrate reserves available for with the golden star tunicate, sponge, and maintenance of plant metabolism (Hemminga burgundy morph of the violet tunicate was and Duarte 2000), and how well the tunicates significantly lower than in unfouled blades, and sponge act as a physical barrier to gas suggesting blade senescence and reduced blade exchange and nutrient uptake. The reduction of lifespan. eelgrass primary production is not compensated In addition to the reduction in light reaching by the tunicates or sponge as it is by fouling fouled eelgrass blades, tunicate and sponge epiphytes, which often contribute significantly to fouling also had a physical impact on eelgrass the total primary productivity of seagrass beds plants. The relatively low retrieval rate of (Moncreiff et al. 1992; Nelson and Waaland marked fouled plants indicates that fouling likely 1997; Wear et al. 1999). Clearly, fouling of caused shoots to break away. Also, many marked eelgrass by invasive colonial tunicates and a fouled plants were retrieved unfouled, indicating native sponge may have numerous negative that fouled blades broke away and were replaced ecological consequences resulting from reduced with new growth. Observations during plant shoot growth, survival, and light attenuation of marking indicated that shoot and blade loss were fouled blades. The severity of these impacts not restricted to the marked plant that was depends on environmental factors controlling the originally fouled, because the fouling organisms population dynamics of the fouling organisms as often spread to neighbouring shoots. The well as the seagrass. These impacts may not have replacement of lost shoots and blades with new previously existed in seagrass beds that are growth may have negative effects on bed usually free of heavy epiphyte fouling, such as productivity, because stored carbohydrate those in the Canadian Maritime provinces. reserves are used that would otherwise be allocated for plant maintenance during the winter Acknowledgements and at the beginning of the growing season (Zimmerman et al. 1995; Lee and Dunton 1997; We thank M. Bravo and J. Ellis for assistance in the laboratory Alcoverro et al. 2001). The fouling organisms and field. M. Dowd provided valuable scientific discussion. Comments from anonymous reviewers improved the manuscript. themselves likely benefited from the breaking Funding was provided by the Aquatic Invasive Species Research away of shoots and blades, because it provided a program of Fisheries and Oceans Canada. transport mechanism (i.e. “rafting”) allowing colonization of new areas in the eelgrass bed and References of new eelgrass beds in neighbouring bays (Worcester 1994). Alcoverro T, Manzanera M, Romero J (2001) Annual metabolic Numerous ecological consequences for carbon balance of the seagrass Posidonia oceanica: the eelgrass beds are likely when shoot growth, importance of carbon reserves. Marine Ecology Progress Series 211: 105-116, http://dx.doi.org/10.3354/meps211105 survival, and light attenuation are reduced by Alcoverro T, Pérez M, Romero J (2004) Importance of within- fouling from the invasive colonial tunicates and shoot epiphyte distribution for the carbon budget of the native sponge. The reduction in incident PAR seagrasses: the example of Posidonia oceanica. Botanica reaching fouled eelgrass blades documented in Marina 47: 307-312, http://dx.doi.org/10.1515/BOT.2004.036 Bale AJ, Kenny AJ (2005) Sediment analysis and seabed our study will lower photosynthesis within all characterization. In: Eleftheriou A, McIntyre A (eds), parts of the plant (Williams and McRoy 1982; Methods for the study of the marine benthos. Blackwell Beer et al. 1998). This will reduce the number of Science Ltd, Oxford, UK, pp 43-86, http://dx.doi.org/ leaves per shoot, shoot growth rate, and shoot 10.1002/9780470995129.ch2

324 Invasive tunicates in seagrass beds

Beer S, Vilenkin B, Weil A, Veste M, Susel L, Eshel A (1998) Duarte CM (2002) The future of seagrass meadows. Measuring photosynthetic rates in seagrasses by pulse Environmental Conservation 29: 192-206, http://dx.doi.org/ amplitude modulated (PAM) fluorometry. Marine Ecology 10.1017/S0376892902000127 Progress Series 174: 293-300, http://dx.doi.org/10.3354/ Duarte CM, Chiscano CL (1999) Seagrass biomass and meps174293 production: a reassessment. Aquatic Botany 65: 159-174, Biber PD, Kenworthy JW, Paerl HW (2009) Experimental http://dx.doi.org/10.1016/S0304-3770(99)00038-8 analysis of the response and recovery of Zostera marina (L.) Dubinsky Z, Falkowski P, Wyman K (1986) Light harvesting and and Halodule wrightii (Ascher.) to repeated light-limitation utilization in phytoplankton. Plant Cell Physiology 27: 1335- stress. Journal of Experimental Marine Ecology and Biology 1349 369: 110-117, http://dx.doi.org/10.1016/j.jembe.2008.10.031 Eklöf JS, de la Torre Castro M, Adelsköld L, Jiddawi NS, Brush ML, Nixon SW (2002) Direct measurements of light Kautsky N (2005) Differences in macrofaunal and seagrass attenuation by epiphytes on eelgrass Zostera marina. Marine assemblages in seagrass beds with and without seaweed Ecology Progress Series 238: 73-79, http://dx.doi.org/10. farms. Estuarine and Coastal Shelf Science 63: 385-396, 3354/meps238073 http://dx.doi.org/10.1016/j.ecss.2004.11.014 Bulthuis DA, Woelkerling WJ (1983) Biomass accumulation and Frankovich TA, Zieman JC (2005) Periphyton light transmission shading effects of epiphytes on leaves of the seagrass, relationships in Florida Bay and the Florida Keys, USA. Heterozostera tasmanica, in Victoria, Australia. Aquatic Aquatic Botany 83: 14-30, http://dx.doi.org/10.1016/j.aquabot. Botany 16: 137-148, http://dx.doi.org/10.1016/0304-3770(83) 2005.05.003 90089-X Glazer BT (1999) Analysis of Physical, Chemical, and Biological Carman MR, Grunden DW (2010) First occurrence of the Factors Inhibiting Growth and Restoration of Submerged invasive tunicate Didemnum vexillum in eelgrass habitat. Vascular Plants in Delaware’s Indian River and Rehoboth Aquatic Invasions 5: 23-29, http://dx.doi.org/10.3391/ai. Bays. MSc thesis, University of Delaware, Lewes, DE, USA 2010.5.1.4 Hauxwell J, Cebrián J, Valiela I (2003) Eelgrass Zostera marina Carver CE, Mallet AL, Vercaemer B (2006) Biological synopsis loss in temperate estuaries: relationship to land-derived of the colonial tunicates, Botryllus schlosseri and nitrogen loads and effect of light limitation imposed by algae. Botrylloides violaceus. Canadian Manuscript Report of Marine Ecology Progress Series 247: 59-73, Fisheries and Aquatic Sciences 2747, pp 1-42 http://dx.doi.org/10.3354/meps247059 Cebrián J, Enríquez, Fortes M, Agawin N, Vermaat JE, Duarte Heck KL, Able KW, Roman CT, Fahay MP (1995) Composition, CM (1999) Epiphyte accrual on Posidonia oceanica (L.) abundance, biomass, and production of macrofauna in a New Delile leaves: Implications for light absorption. Botanica England estuary: comparisons among eelgrass meadows and Marina 42: 123-128, http://dx.doi.org/10.1515/BOT.1999.015 other nursery habitats. Estuaries 18: 379-389, Ceccherelli G, Piazzi L, Cinelli F (2000) Response of the http://dx.doi.org/10.2307/1352320 nonindigenous Caulerpa racemosa (Forsskal) J. Agardh to Heck KL, Hays G, Orth RJ (2003) Critical evaluation of the the native seagrass Posidonia oceanica (L.) Delile: effect of nursery role hypothesis for seagrass meadows. Marine density of shoots and orientation of edges of meadows. Ecology Progress Series 253: 123-136, http://dx.doi.org/ Journal of Experimental Marine Biology and Ecology 243: 10.3354/meps253123 227-240, http://dx.doi.org/10.1016/S0022-0981(99)00122-7 Hemminga MA, Duarte CM (2000) Seagrass ecology. Cambridge Collier CJ, Lavery PS, Ralph PJ, Masini RJ (2009) Shade-induced University Press, Cambridge, UK, http://dx.doi.org/10.1017/ CBO9780511525551 response and recovery of the seagrass Posidonia sinuosa. Hirose E, Saito Y Watanabe H (1991) Tunic cell morphology and Journal of Experimental Marine Biology and Ecology 370: classification in Botryllid ascidians. Zoological Science 5: 89-103, http://dx.doi.org/10.1016/j.jembe.2008.12.003 951-958 Critchley AT, Farnham WF, Thorp CH (1997) On the co- Kemp WM, Boynton WR, Adolf JE, Boesch DF, Bolcourt WC, occurrence of two exotic, invasive marine organisms: the Brush G, Cornwell JC, Fisher TR, Gilbert PM, Hagy JD, brown seaweed Sargassum muticum (Yendo) Fensholt and Harding LW, Houde ED, Kimmel DG, Miller WD, Newell the spirorbid tube worm Janua (Neodexiospira) brasiliensis RIE, Roman MR, Smith EM, Stevenson JC (2005) (Grube), in association with the indigenous eelgrass, Zostera Eutrophication of Chesapeake Bay: historical trends and marina L. and wrack, Fucus serratus L. in the south-west ecological interactions. Marine Ecology Progress Series 303: Netherlands and the Channel Islands, Europe. South African 1-29, http://dx.doi.org/10.3354/meps303001 Journal Botany 63: 474-479 Lee K-S, Dunton KH (1997) Effects of in situ light reduction on Cummings ME, Zimmerman RC (2003) Light harvesting and the the maintenance, growth and partitioning of carbon resources package effect in the seagrasses Thalassia testudinum Banks in Thalassia testudinum Banks ex König. Journal of ex König and Zostera marina L.: optical constraints on Experimental Marine Biology and Ecology 210: 53-73, photoacclimation. Aquatic Botany 75: 261-274, http://dx.doi.org/10.1016/S0022-0981(96)02720-7 http://dx.doi.org/10.1016/S0304-3770(02)00180-8 Moncreiff CA, Sullivan MJ, Daehnick AE (1992) Primary Davis RC, Short FT, Burdick DM (1998) Quantifying the effects production dynamics in seagrass beds of Mississippi Sounds of green crab damage to eelgrass transplants. Restoration – the contributions of seagrass, epiphytic algae, sand Ecology 6: 297-302, http://dx.doi.org/10.1046/j.1526-100X. microflora, and phytoplankton. Marine Ecology Progress 1998.00634.x den Hartog C (1997) Is Sargassum a threat to eelgrass beds? Series 87: 161-171, http://dx.doi.org/10.3354/meps087161 Aquatic Botany 58: 37-41, http://dx.doi.org/10.1016/S0304- Moore KA, Wetzel RL (2000) Seasonal variations in eelgrass 3770(97)00007-7 (Zostera marina L.) responses to nutrient enrichment and Draper NR, Smith H (1998) Applied regression analysis. John reduced light availability in experimental ecosystems. Wiley and Sons, NY, USA Journal of Experimental Marine Biology and Ecology 244: 1- Drouin A, McKindsey CW, Johnson LE (2011) Higher abundance 28, http://dx.doi.org/10.1016/S0022-0981(99)00135-5 and diversity in faunal assemblages with the invasion of Nelson TA, Waaland JR (1997) Seasonality of eelgrass, epiphyte, Codium fragile ssp. fragile in eelgrass meadows. Marine and grazer biomass and productivity in subtidal eelgrass Ecology Progress Series 424: 105-117, http://dx.doi.org/ meadows subjected to moderate tidal amplitude. Aquatic 10.3354/meps08961 Botany 56: 51-74, http://dx.doi.org/10.1016/S0304-3770(96) 01094-7

325 M.C. Wong and B. Vercaemer

Ochieng CA, Short FT, Walker DI (2010) Photosynthetic and Short FT, Neckles HA (1999) The effects of global climate morphological responses of eelgrass (Zostera marina L.) to a change on seagrasses. Aquatic Botany 63: 169-196, gradient of light conditions. Journal of Experimental Marine http://dx.doi.org/10.1016/S0304-3770(98)00117-X Biology and Ecology 382: 117-124, http://dx.doi.org/ Short FT, Wyllie-Echeverria S (1996) Natural and human- 10.1016/j.jembe.2009.11.007 induced disturbance of seagrasses. Environmental Olesen B, Sand-Jensen K (1993) Seasonal acclimatization of Conservation 23: 17-27, http://dx.doi.org/10.1017/S03768929 eelgrass Zostera marina growth to light. Marine Ecology 00038212 Progress Series 94: 91-99, http://dx.doi.org/10.3354/meps Walker DI, Kendrick GA, McComb AJ (2006) Decline and 094091 recovery of seagrass ecosystems - the dynamics of change. Orth RJ, Carruthers TJB, Dennison WC, Duarte CM, Fourqurean In: Larkum AWD, Orth RJ, Duarte CM (eds), Seagrasses: JW, Heck KL, Hughes AR, Kendrick GA, Kenworthy WJ, Biology, Ecology and Conservation. Springer, Dordrecht, Olyarnik S, Short FT, Waycott M, Williams SL (2006) A The Netherlands, pp 551-565 global crisis for seagrass ecosystems. BioScience 56: 987- Waycott M, Cuarte CM, Carruthers TJB, Orth RJ, Dennison WC, 996, http://dx.doi.org/10.1641/0006-3568(2006)56[987:AGC Olyarnik S, Calladine A, Fourqurean JW, Heck KL, Hughes FSE]2.0.CO;2 AR, Kendrick GA, Kenworthy WJ, Short FT, Williams SL Quinn GP, Keough MJ (2002) Experimental design and data (2009) Accelerating loss of seagrasses across the globe analysis for biologists. Cambridge University Press, threatens coastal ecosystems. Proceedings of the National Cambridge, UK Academy of Sciences 106: 12377-12381, http://dx.doi.org/ Ralph PJ, Durako MJ, Enriquez S, Collier CJ, Doblin MA (2007) 10.1073/pnas.0905620106 Impact of light limitation on seagrasses. Journal of Wear DJ, Sullivan MJ, Moore AD, Millie DF (1999) Effects of Experimental Marine Biology and Ecology 350: 176-193, water-column enrichment on the production dynamics of http://dx.doi.org/10.1016/j.jembe.2007.06.017 three seagrass species and their epiphytic algae. Marine Robertson AI, Mann KH (1984) Disturbance by ice and life- Ecology Progress Series 179: 201-213, http://dx.doi.org/ history adaptations of the seagrass Zostera marina. Marine 10.3354/meps179201 Biology 80: 131-141, http://dx.doi.org/10.1007/BF02180180 Welschmeyer NA (1994) Fluorometric analysis of chlorophyll a Ruiz GM, Fofonoff PW, Carlton JT, Wonham MJ, Hines AH in the presence of chlorophyll b and pheopigments. (2000) Invasion of coastal marine communities in North Limnology and Oceanography 39: 1985-1992, America: Apparent patterns, processes, and biases. Annual http://dx.doi.org/10.4319/lo.1994.39.8.1985 Review of Ecology and Systematics 31: 481-531, Williams SL (2007) Introduced species in seagrass ecosystems: http://dx.doi.org/10.1146/annurev.ecolsys.31.1.481 status and concerns. Journal of Experimental Marine Biology Ruppert EE, Fox RS, Barnes RD (2003) Invertebrate Zoology: A and Ecology 350: 89-110, http://dx.doi.org/10.1016/j.jembe. Functional Evolutionary Approach. Brooks Cole, Thompson 2007.05.032 Learning, CA, USA Williams SL, McRoy CP (1982) Seagrass productivity effect of Schmidt AL, Coll M, Romanuk TN, Lotze HK (2011) Ecosystem light on carbon uptake. Aquatic Botany 12: 321-344, structure and services in eelgrass Zostera marina and http://dx.doi.org/10.1016/0304-3770(82)90025-0 rockweed Ascophyllum nodosum. Marine Ecology Progress Wong MC, Peterson CH, Piehler MF (2011) Evaluating estuarine Series 437: 51-68, http://dx.doi.org/10.3354/meps09276 habitats using secondary production as a proxy for food web Sewell AT (1996) Eelgrass Growth and Abundance in an Urban support. Marine Ecology Progress Series 440: 11-25, Estuary: The Negative Effects of Anemone Coverage. MS http://dx.doi.org/10.3354/meps09323 thesis, San Diego State University, San Diego, CA Worcester SE (1994) Adult rafting versus larval swimming: Short FT, Burdick DM, Kaldy JE (1995) Mesocosm experiments dispersal and recruitment of a botryllid ascidian on eelgrass. quantify the effects of eutrophication on eelgrass, Zostera Marine Biology 121: 309-317, http://dx.doi.org/10.1007/ BF00346739 marina L. Limnology and Oceanography 40: 740-749, Zimmerman RC, Reguzzoni JL, Alberte RS (1995) Eelgrass http://dx.doi.org/10.4319/lo.1995.40.4.0740 Short FT, Duarte CM (2001) Methods for the measurement of (Zostera marina L.) transplants in San Francisco Bay: role of seagrass growth and production. In: Short FT, Duarte CM light availability on metabolism, growth and survival. (eds), Global seagrass research methods. Elsevier Science, Aquatic Botany 51: 67-86, http://dx.doi.org/10.1016/0304- 3770(95)00472-C Amsterdam, The Netherlands, pp 156-182, http://dx.doi.org/ 10.1016/B978-044450891-1/50009-8

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