Biofouling Monitoring for Aquatic Invasive Species (AIS) in DFO Maritimes, southwest New Brunswick:

Transcription

1 Biofouling Monitoring for Aquatic Invasive Species (AIS) in DFO Maritimes, southwest New Brunswick: Dawn Sephton, Jennifer Martin, Murielle LeGresley and Kelsi Godin Science Branch Coastal Ecosystem Research Division 2016 Canadian Technical Report of Fisheries and Aquatic Sciences 3140

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3 i Canadian Technical Report of Fisheries and Aquatic Sciences No Biofouling Monitoring for Aquatic Invasive Species (AIS) in DFO Maritimes Region, southwest New Brunswick: by Dawn Sephton, Jennifer Martin, Murielle LeGresley and Kelsi Godin Science Branch Maritimes Region Coastal Ecosystem Science Division Fisheries and Oceans Canada Bedford Institute of Oceanography PO Box 1006 Dartmouth, Nova Scotia, B2Y 4A2

4 ii Think Recycling! Pensez à recycler Her Majesty the Queen in Right of Canada, Cat. No. Fs. 97-6/3140E ISBN ISSN Correct citation for this publication: Sephton, D., J.L. Martin, M. LeGresley and K. Godin Biofouling monitoring for aquatic invasive species (AIS) in DFO Maritimes Region, southwest New Brunswick: Can.Tech. Rep. Fish. Aquat. Sci. 3140: vi + 68 p.

5 iii TABLE OF CONTENTS ABSTRACT v RÉSUMÉ. vi 1.0 INTRODUCTION MATERIALS AND METHODS STATION SELECTION MONITORING COLLECTORS MONITORING PROTOCOL DETERMINATION OF PRESENCE AND PERCENTAGE COVERAGE OF TUNICATES RESULTS GENERAL OCCURRENCE, TUNICATE COVERAGE (DEGREE OF INFESTATION): Ciona intestinalis Botryllus schlosseri Botrylloides violaceus DISCUSSION TUNICATE ESTABLISHMENT AND SPREAD, Ciona intestinalis Botryllus schlosseri Botrylloides violaceus Non-detection of Styela clava and Didemnum vexillum SPATIAL AND TEMPORAL VARIATION IN TUNICATE DISTRIBUTIONS Spatial Variation Variation among collectors Range Expansions Temporal Variation Seasonal Variation Year-to-year Variation CONCLUDING REMARKS.. 36 ACKNOWLEDGEMENTS.. 37 REFERENCES. 38 APPENDIX 1. Monitoring details and tunicate age by station, APPENDIX 2. Monitoring details and tunicate age by station,

6 APPENDIX 3. Monitoring details and tunicate age by station, APPENDIX 4. Monitoring details and tunicate age by station, APPENDIX 5. Monitoring details and tunicate age by station, APPENDIX 6. Monitoring details and tunicate age by station, iv

7 v ABSTRACT Sephton, D., Martin, J.L., LeGresley, M. and Godin, K Biofouling monitoring for aquatic invasive species (AIS) in DFO Maritimes Region, southwest New Brunswick; Can. Tech. Rep. Fish. Aquat. Sci. 3140: vi + 68 p. The establishment of four species of invasive tunicates: Ciona intestinalis, Botryllus schlosseri, Botrylloides violaceus and Styela clava has had detrimental impacts on the shellfish culture industry in Atlantic Canada, and several additional species have been identified as potential new invaders to the region (Locke 2009). A surveillance and monitoring program for invasive, biofouling tunicates was initiated on the southwest coast of New Brunswick by DFO Maritimes Region in 2006 at geo-referenced coastal monitoring stations. Monitoring collectors were deployed at 11 to 21 stations annually during three deployment intervals; First (May - August), Second (August - October) and Full (May October). In 2008, 14 stations were designated as sentinel stations to be monitored every year thereafter. Ciona intestinalis was the most widespread tunicate in this region, found at 64% to 95% of all stations monitored between 2006 and 2011, and at 86% to 93% of sentinel stations between 2008 and Botryllus schlosseri was also present at many stations: 36% to 81% of all stations monitored between 2006 and 2011, and at 71% to 93% of sentinel stations between 2008 and Botrylloides violaceus was found in this region for the first time in 2009 at a single station: Head Harbour (7% of sentinel stations). It was recorded at two (14%) and three (21%) sentinel stations in 2010 and 2011, respectively. While the number of sentinel stations where C. intestinalis has been detected has not increased during the course of this work, both B. schlosseri and B. violaceus have spread to new stations in the area. Styela clava, and Didemnum vexillum were not detected in southwest NB between 2006 and Musquash was the only tunicate-free sentinel station, while tunicates were not found at Kennebecasis Yacht Club and Saint John during the earlier years (2006 and 2007) of the monitoring program.

8 vi RÉSUMÉ Sephton, D., Martin, J.L., LeGresley, M.M. et Godin, K Surveillance de biosalissures pour les espèces aquatiques envahissantes (EAE) dans la région des Maritimes du MPO, sud-ouest Nouveau Brunswick : Can. Tech. Rep. Fish. Aquat. Sci : vi + 60 p. L établissement de quatre espèces de tuniciers envahissants: Ciona intestinalis, Botryllus schlosseri, Botrylloides violaceus et Styela clava a eu des effets néfastes sur l'industrie de la conchyliculture au Canada atlantique, et plusieurs espèces supplémentaires ont été identifiées (Locke 2009) en tant que nouveaux envahisseurs potentiels dans la région. Un programme de surveillance et de contrôle pour les tuniciers envahissants intrinsèques aux salissures biologiques a été lancé en 2006 dans la partie Nouveau-Brunswick de la Région des Maritimes du MPO, basé sur plusieurs stations côtières de surveillance géoréférencées. Des collecteurs de surveillance (plaques) ont été déployés à stations chaque année au cours de plusieurs périodes de déploiement; première période (mai - août), deuxième période (août - octobre) et période complète (mai - octobre). Quatorze stations sentinelles ont été identifiées en 2008 pour faire partie du programme de surveillance de façon annuelle. Ciona intestinalis était l'ascidie la plus répandue dans cette région, et se retrouve dans 64% à 95% de toutes les stations surveillées entre 2006 et 2011, et dans 86% à 93 % des stations sentinelles entre 2008 et Botryllus schlosseri était également présent dans de nombreuses stations: 36% à 81% de toutes les stations surveillées entre 2006 et 2011, et 71% à 93% des stations sentinelles entre 2008 et Botrylloides violaceus a été détecté pour la premiere fois en 2009 à une seule station : Head Harbour (7% des stations sentinelles). Il a été trouvé à deux (14%) et trois (21%) stations sentinelles stations en 2010 et 2011, respectivement. Pendant cette travaille, le nombre de stations n était pas augmenté on C. intestinalis était localizé, mais les deux espèces B. schlosseri et B. violaceus était déplacé vers nouveaux stations dans cette locale. Les espèces S. clava et Didemnum vexillum n'ont pas été détectées dans le sud-ouest du NB entre 2006 et Musquash était la seule station sentinelle libre de tuniciers de 2006 à 2011 et aucune espèce de tuniciers n a été trouvée au Kennebecasis Yacht Club ou à Saint Jean, NB au cours des années préliminaires (2006 et 2007) du programme de surveillance.

9 1 1.0 INTRODUCTION Non-indigenous species (NIS) pose great risk to native species, and biodiversity, and can negatively impact native ecosystems and their function (Sala et al. 2000). Among NIS of global concern, ascidian tunicates, commonly known as sea-squirts, have affected marine ecosystems through their negative impacts on native species (Lambert 2001; Lutz-Collins et al. 2009) and communities (Blum et al. 2007; Dijkstra et al. 2007a; Lambert and Lambert 1998, 2003; Lengyel et al. 2009). Tunicates pose a serious threat to shellfish aquaculture operations as they overgrow bivalves and gear (Carver et al. 2003; Bullard et al. 2007a; Valentine et al. 2007a), resulting in increased operation and production costs (MacNair et al. 2006; McKindsey et al. 2007). Non-indigenous tunicate infestations have resulted in decreased productivity and growth of blue mussels, Mytilus edulis (Linnaeus, 1758) (Daigle and Herbinger 2009) in Nova Scotia (NS), and significant losses to the mussel culture industry in Prince Edward Island (PEI) and NS since the mid 1990 s (Boothroyd et al. 2002; Clarke and Therriault 2007; Howes et al. 2007; Ramsay et al. 2008). Although shellfish aquaculture is not established in southwestern New Brunswick (swnb), this is a pilot research area (DFO 2013) for integrated multitrophic aquaculture (IMTA), so the presence of tunicates may impact this developing industry. Four species of non-indigenous, fouling tunicates are now well established in the coastal waters of Atlantic Canada (Carver et al. 2006a, b; Clarke and Therriault, 2007; Sargent et al. 2013; Sephton et al. 2011, 2014, 2015; Vercaemer and Sephton 2014); two solitary species, the vase tunicate, Ciona intestinalis (Linneaus, 1776), and the clubbed tunicate Styela clava (Herdmann, 1881), and two colonial species; the golden star tunicate Botryllus schlosseri (Pallas,1766), and the violet tunicate, Botrylloides violaceus (Oka, 1927). Since 2012, three potential invaders identified by Locke (2009) have been detected in Nova Scotia: the solitary European sea squirt Ascidiella aspersa (Muller, 1776) and two colonial species, the compound sea squirt Diplosoma listerianum (Milne-Edwards, 1841), and pancake batter tunicate, Didemnum vexillum (Kott, 2002) (Moore et al. 2014; Vercaemer et al. 2015). Of these species, three present in nearby NS waters at the onset of monitoring in the Bay of Fundy region of southwest New Brunswick (NB), C. intestinalis and B. schlosseri have been recorded previously (Van Name 1912, 1945; Gosner, 1971; Linkletter et al., 1977), while B. violaceus has not (LeGresley et al. 2008; Martin et al. 2011). Didemnum vexillum, A. aspersa, and D. listerianum have also not been detected previously (LeGresley et al. 2008; Martin et al. 2011). The arrival and establishment of D. vexillum in this region is of particular concern, as it has been present on the United States (US) side of George s Bank since

10 (Valentine et al. 2007b) and in US waters, at Eastport, Maine, since 2003 (Vercaemer et al. 2015). In response to the growing threat posed by aquatic invasive species (AIS) to native coastal communities, fisheries and shellfish aquaculture in Atlantic Canadian waters, Fisheries and Oceans Canada (DFO) developed and initiated an AIS Biofouling Monitoring Program in 2006 (Sephton et al., 2011), including the Bay of Fundy region of swnb. LeGresley et al. (2008) and Martin et al. (2011) reported on the presence of tunicates, based on settlement on Full season (May to November) plates only, at most stations in this region from Here we provide additional information on trends in tunicate distribution based on data obtained during all deployment periods from 2006 to 2009, as well as the results of biofouling monitoring conducted in 2010 and MATERIALS AND METHODS 2.1. STATION SELECTION Coastal general monitoring stations were selected for the DFO AIS Monitoring program based on the presence of potential or existing risk factors for the introduction of spread of AIS. These included; (1) presence of shellfish or mussel processing facilities, (2) mussel or shellfish aquaculture sites, (3) important commercial port with international traffic, (4) marina of yacht club with US traffic, (5) commercial fishing harbours, and (6) harbours with herring or US lobster processing facilities. As noted above, shellfish culture and processing (factors 1 and 2) are not yet established in this region, however, there is considerable fishing and recreational traffic into this region from US destinations (factors 3 and 4), as well as considerable exchange of water (Chang et al. 2005). Sites with air-exposure at low tide without floating docks were not included. Twenty-five geo-referenced stations were established between 2006 and 2011 (Table 1), and of these, 11 to 21 stations were monitored each year. Fourteen sentinel stations (Table 2) were designated for ongoing annual monitoring in These were identified as critical stations due to ecological considerations, proximity to established populations of AIS or elevated risk by the presence of vectors of AIS introduction.

11 Table 1: Aquatic Invasive Species biofouling monitoring stations in southwest New Brunswick (swnb), , and including Passamaquoddy Bay, and Deer, Campobello and Grand Manan Islands. Stn Location Region Latitude, Longitude, Station Description and Deployment Stucture No. o N o W 164 Kennebecasis Ya swnb Yacht Club, floating dock 165 Saint John swnb Large shipping harbour, floating dock 132 Musquash (Five swnb Small fishing harbour, Marine Protected Area, Fathom Harbour) public wharf, floating dock 133 Dipper Harbour swnb Medium fishing and processing harbour, public wharf, floating dock 135 Beaver Harbour swnb Medium fishing and processing harbour, public wharf, floating dock 139 Lime Kiln Bay swnb Salmon aquaculture site, cages 141 Bay Bay swnb Integrated multitrophic aquaculture site, cages 138 Wallace Cove swnb Small cove, floating dock 142 Charlie Cove swnb Integrated multitrophic aquaculture site, cages 143 L'Etete swnb Passenger Ferry landing, floating dock 153 Crow Island swnb Integrated multitrophic aquaculture site, cages 148 Hog Island Passamaquoddy Salmon aquaculture site, cages 151 St. Andrews Passamaquoddy Medium, mixed use port, floating dock 152 SABS Passamaquoddy Government Research Station wharf, floating dock 149 Fairhaven Dock Deer Island Small fishing harbour, public wharf, floating dock 150 Fairhaven MF Deer Island Salmon aquaculture site, cages 146 Leonardville Deer Island Small fishing harbour, public wharf, floating dock 147 Indian Island Fundy Isles Salmon aquaculture site, cages 144 Head Harbour Campobello Medium fishing harbour, public wharf, floating dock 3

12 Table 1, continued. Stn Location Region Latitude, Longitude, Station Description and Deployment Stucture No. N W 163 Harbour de LoutreCampobello Small harbour, mussel cages and floating dock 215 Roosevelt ProvincCampobello Small cove, floating dock 145 Wilson's Beach Campobello Small fishing harbour, public wharf, floating dock 136 North Head Grand Manan Medium, mixed use harbour, public wharf floating dock 137 Ingalls Head Grand Manan Medium fishing harbour, public wharf, floating dock 140 Seal Cove Grand Manan Medium fishing harbour, public wharf, floating dock 4

13 5 Table 2: Biofouling monitoring stations monitored in swnb, (S) = sentinel station. Stn Location Year No Kennebecasis Yacht Club x 165 Saint John x x 132 Musquash (Five Fathom Hrb) (S) x x x x 133 Dipper Harbour (S) x x x x x x 135 Beaver Harbour (S) x x x x x 139 Lime Kiln Bay x x x x 141 Back Bay (S) x x x x x 138 Wallace Cove x x x x x 142 Charlie Cove x x x x 143 L'Etete (S) x x x x x 153 Crow Island x 148 Hog Island x x x 151 St. Andrews (S) x x x x x x 152 SABS (S) x x x x x x 149 Fairhaven Dock (S) x x x x x 150 Fairhaven MF x x x x x lost 146 Leonardville (S) x x x x x 147 Indian Island x x x x 144 Head Harbour (S) x x x x x x 163 Harbour de Loutre x x x x 215 Roosevelt Provincial Park x 145 Wilson's Beach (S) x x x x 136 North Head (S) x x x x x x 137 Ingalls Head (S) x x x x x 140 Seal Cove (S) x x x x x x Total # of Stations Monitored MONITORING COLLECTORS The saucer-petri dish-polyvinyl chloride (PVC) plate monitoring collector (Figure 1) and described by Martin et al. (2011) was used in monitoring in Beginning in 2009, collector saucers had only three Petri dishes, and not four as shown in Figure 1.

14 6 Figure 1: AIS plate collectors used in biofouling monitoring in southwest New Brunswick, Beginning in 2009, only 3 Petri dishes were attached to the saucer MONITORING PROTOCOL A total of 25 geo-referenced stations were established in the region and monitored at least once between 2006 and Of these, 11 to 21 were monitored in each year (Table 2) by staff from DFO Science Branch, Maritimes Region staff, based at the St. Andrews Biological Station (SABS). Fourteen sentinel stations were established in 2008, and of these, 6 and 12 were monitored in 2006 and 2007, respectively. Collectors were suspended at a depth of 1 m below the water surface so that they remained submerged throughout the tidal cycle (Martin et al. 2011). The number of collectors deployed at and retrieved from each site during First (May deployment and August retrieval), Second (August deployment and October retrieval) and Full (May deployment and October retrieval) deployments differed from year to year (Table 3). Table 3. Number of monitoring collectors deployed annually at each monitoring station. Deployment First Second Full

15 7 Collectors were deployed predominantly from floating docks, but also from buoys and cages on aquaculture sites (Martin et al. 2011). Collectors were deployed in such a way that multiple areas representative of differing habitats (sheltered or higher current) of the station were sampled. Details of collector deployments and species ages for all stations from 2006 to 2011 are shown in Appendices 1 to 6, respectively. Following the prescribed monitoring period (see Appendices 1-6), collectors were removed and brought to the laboratory. Collector ropes, tags and weights, as well as plates and petri dishes, were examined for the presence of tunicates (Table 4). Table 4: List of non-indigenous tunicate species subject to detection and monitoring in Scientific Name Ciona intestinalis Botryllus schlosseri Botrylloides violaceus Styela clava Didemnum vexillum Common Name Vase tunicate Golden star tunicate Violet tunicate Clubbed tunicate Pancake batter tunicate Individual Petri dishes and monitoring plates from each collector were placed bottom-side up in sequence (top plate = left (1), middle plate = centre (2) and bottom plate = right (3)) on a white, labelled background and photographed using a digital Camera (2006: Canon Powershot SD700 IS; : Nikon D80). Each Petri dish and plate was examined under a Nikon SMZ1000 dissecting scope and the presence of invasive tunicates was noted. These data were recorded and entered into a Microsoft ACCESS database for further analysis. 2.4 DETERMINATION OF PRESENCE AND PERCENTAGE COVERAGE OF TUNICATES In 2006, the results of microscopic plate and petri dish analysis were used to determine the percentage for each tunicate species for each plate. In , Image J version 1.37 analysis [ImageJ 1.37 Wayne Rasbund, National Institute of Health, USA _23(32 bit)] was used to determine the percent of plates and petri dishes from the digital photographs. Categories for the percentage were 0 (absent); 1: < 25% age (minimal) of the under plate surface; 2: 25 50% age (moderate); 3: 51 75% age (abundant), and 4: % age (very abundant). Median numerical values for each age category were 0: 0, 1: 12.5%, 2: 37.5%, 3: 62.5%, and 4: 87.5%.

16 8 The average percentage category for each species of tunicate at each station was determined separately for each deployment period (First, Second and Full). An average value for a deployment period was calculated by dividing the sum of the median numerical values by the total number of plates and petri dishes reed (i.e. four petri dishes and four plates per collector in , and three petri dishes and three plates per collector thereafter). The number of collectors deployed in each deployment period in each year is given in Table 5. The average numerical value obtained was then converted to a category value (i.e., 0-4). Similarly, the year average percentage category was determined as the sum of all median values divided by the total number of plates and petri dishes reed in all deployment periods (Table 5). Median values were converted to the appropriate category value. Table 5: Maximum numbers of petri dish and PVC plates used to calculate the year average percentage age of tunicates at a monitoring station, in each year # of Collectors Deployed # of Petri Dishes # of PVC Plates RESULTS 3.2 GENERAL OCCURRENCE, General results for the presence of tunicate species monitored annually at all stations from , and at the 14 sentinel stations from 2008 to 2011, are given in Tables 6 and 7, respectively. Locations where C. intestinalis, B. schlosseri, and B. violaceus were present in 2006 through 2011 are shown in Figures 2 to 7, respectively. Ciona intestinalis was the most frequently recorded tunicate overall; present at 64 95% of all stations from 2006 through 2011 (Table 6), and at one or more stations in Passamaquoddy Bay (PQB), at coastal locations (Charlie Cove L Etete Beaver Harbour), and on Deer, Campobello and Grand Manan Islands throughout the study period (Figures 2 7). While B. schlosseri was present at many stations between 2006 and 2011 (Figures 2 7), ranging from 36% to 81%, it was consistently second in dominance compared to C. intestinalis. While C. intestinalis occurred at about the same number of stations from , B. schlosseri appeared at more stations during this time. Botrylloides violaceus was the least frequently recorded tunicate during

17 9 this period. It was noted for the first time in 2009 only at Head Harbour on Campobello Island (Figure 5). It was found at Beaver Harbour, along the Bay of Fundy (BoF) coast, and at Wilson s Beach on Campobello Island (Figure 6) in 2010, and at all three of the aforementioned stations in 2011 (Figure 7). Table 6: Prevalence of non-indigenous tunicates in swnb at all monitoring stations, The number of stations where a tunicate was present, and its percentage occurrence, are given for each year. Species Ciona intestinalis 7 (64%) 15 (71%) 19 (90%) 19 (90%) 18 (95%) 14 (88%) Botryllus schlosseri 4 (36%) 10 (48%) 10 (48%) 12 (57%) 13 (68%) 13 (81%) Botrylloides violaceus (5%) 2 (10%) 3 (19%) Styela clava Didemnum vexillum No tunicates 3 (27%) 4 (19%) 1 (5%) 1 (5%) 1 (5%) 1 (6%) # of Stations Monitored Styela clava, and D. vexillum, were not detected in the region between 2006 and 2011, although the native D. albidum was (Martin et al. 2011). Only Musquash, a very shallow, silty, high current with low and variable salinity where collectors were often exposed to air, was free of tunicates from Tunicates were not found in Saint John Harbour or in Head Harbour during monitoring in 2006 and 2007, nor in North Head in 2006, or at the Kennebecasis Yacht Club or in Lime Kiln Bay in The occurrence of non-indigenous tunicates at sentinel stations from 2008 to 2011 gives a more accurate picture of their spread (Table 7). While C. intestinalis was consistently present at 12 or 13 stations during this period, B. schlosseri was detected at more stations with time, as was B. violaceus after its initial appearance in 2009.

18 Figure 2: Presence of Ciona intestinalis, Botryllus schlosseri and Botrylloides violaceus in southwest New Brunswick in

19 Figure 3: Presence of Ciona intestinalis, Botryllus schlosseri and Botrylloides violaceus in southwest New Brunswick in

20 Figure 4: Presence of Ciona intestinalis, Botryllus schlosseri and Botrylloides violaceus in southwest New Brunswick in

21 Figure 5: Presence of Ciona intestinalis, Botryllus schlosseri and Botrylloides violaceus in southwest New Brunswick in

22 Figure 6: Presence of Ciona intestinalis, Botryllus schlosseri and Botrylloides violaceus in southwest New Brunswick in

23 Figure 7: Presence of Ciona intestinalis, Botryllus schlosseri and Botrylloides volaceaus in southwest New Brunswick in

24 16 Table 7: Presence of non-indigenous tunicates in swnb at sentinel stations (n = 14), The number of stations where a tunicate was present, as well as the percentage of the total number of sentinel stations, are given. Species Ciona intestinalis 12 (86%) 12 (86%) 13 (93%) 12 (86%) Botryllus schlosseri 10 (71%) 11 (79%) 11 (79%) 13 (93%) Botrylloides violaceus 0 1 (7%) 2 (14%) 3 (21%) Styela clava Didemnum vexillum No tunicates present 2 (14%) 1 (7%) 1 (7%) 1 (7%) 3.3 TUNICATE COVERAGES (DEGREE OF INFESTATION) Ciona intestinalis Average annual age of C. intestinalis for 2006 through 2011 is shown in Figures 8 through 13, respectively. Coverage by C. intestinalis was low (<25%) where it was present in 2006 and 2007, except for moderate age (26-50%) at Lime Kiln Bay in 2006 (Figure 8). In 2008, moderate ages of C. intestinalis were noted at 5 of 21 stations, located in PQB, along the BoF coast and on Campobello Island (Figure 10), with low age at all other stations. Low age was again evident at all stations in 2009 (Figure 11). Heaviest ages were noted in 2010 (Figure 12) when age was moderate at 6 of 19 stations, and abundant at 4 of 10 stations, in several areas in swnb. Coverage was heaviest (abundant) at Back Bay in 2011 (Figure 13), and moderate at 6 stations in three areas of swnb in Grand Manan Island was the only area monitored in the region where ages of C. intestinalis were consistently low (<25%) Botryllus schlosseri Average annual age of B. schlosseri was low (<25%) in all years (Figures 14 19), with a few exceptions. In 2008, age was moderate (26-50%) at one station only on Grand Manan Island (Ingalls Head) (Figure 16), while in 2009, age was also moderate at Beaver Harbour, on the BoF shore, and at Harbour de Loutre, on Campobello Island (Figure 17) Botrylloides violaceus Average annual age of B. violaceus was low (<25%) whenever ( ) and wherever it occurred (Appendix 4; station 144: present on one Petri dish only, Appendix 5; stations 135 and 145: present on one or two plates only, and Appendix 6: stations 135, 144 and 145: present on one or two plates only).

25 Figure 8: Average annual plate age of Ciona intestinalis in southwest New Brunswick in

26 Figure 9: Average annual plate age of Ciona intestinalis in southwest New Brunswick in

27 Figure 10: Average annual plate age of Ciona intestinalis in southwest New Brunswick in

28 Figure 11: Average annual plate age of Ciona intestinalis in southwest New Brunswick in

29 Figure 12: Average annual plate age of Ciona intestinalis in southwest New Brunswick in

30 Figure 13: Average annual plate age of Ciona intestinalis in southwest New Brunswick in

31 Figure 14: Average annual plate age of Botryllus schlosseri in southwest New Brunswick in

32 Figure 15: Average annual plate age of Botryllus schlosseri in southwest New Brunswick in

33 Figure 16: Average annual plate age of Botryllus schlosseri in southwest New Brunswick in

34 Figure 17: Average annual plate age of Botryllus schlosseri in southwest New Brunswick in

35 Figure 18: Average annual plate age of Botryllus schlosseri in southwest New Brunswick in

36 Figure 19: Average annual plate age of Botryllus schlosseri in southwest New Brunswick in

37 DISCUSSION 4.2 TUNICATE ESTABLISHMENT AND SPREAD, Two species of non-native tunicates, C. intestinalis and B. schlosseri, were known to be present in this region (Van Name, 1912, 1945; Gosner, 1971; Linkletter et al. 1977) at the onset of the AIS Biofouling Monitoring program in Botryllus violaceus and D. vexillum were noted in nearby US waters in the Gulf of Maine (Osman and Whitlatch, 1995; Dijkstra et al. 2007b), while S. clava has been present in New England since 1970 (Moore et al. 2014). Monitoring results from 2006 to 2011 confirm the presence of C. intestinalis and B. schlosseri, and of a previously unrecorded species, B. violaceus, in this region. Given the potential for anthropogenic introduction by commercial and recreational vessel traffic from the US (Martin et al. 2011; Sephton and Vercaemer 2015; Vercaemer et al. 2015), and the movement of vessels and gear among salmon aquaculture sites in the region (Martin et al. 2011), it is possible that new tunicate species present in the Gulf of Maine and along the US eastern seaboard may arrive, survive and spread in the future (Locke 2009). There is also an increasing risk of arrival via circulating currents or vessel traffic from nearby NS for D. vexillum, confirmed as present in the upper Bay of Fundy in 2013 (Moore et al. 2014; Vercaemer et al. 2015), and S. clava, now present in Lunenburg and Halifax harbours and in Chedabucto Bay (Vercaemer and Sephton, 2014; Sephton et al., In prep.) Ciona intestinalis Ciona intestinalis was recorded at 7 of 11 monitoring stations in 2006, and by 2008 it was established at almost all monitoring sites as the most prevalent tunicate species in the region. The Musquash station, located in a Marine Protected Area, was the only station free of C. intestinalis. Environmental conditions (high current, low and variable salinity, silty water, low tidal levels and substrate exposure) may be unsuitable for tunicate establishment. Re-analysis of all the monitoring plate pictures, and not just those from Full deployment collectors, indicated that C. intestinalis was present, but not reported (see Martin et al. 2011) at Seal Cove in 2006 and at Dipper Harbour in Failure to detect C. intestinalis at Dipper Harbour in 2009 is probably due the low incidence of this species at the station, as only one individual was found on monitoring plates here in Settlement of C. intestinalis larvae is also highly aggregated (i.e., patchy) (Osman and Whitlatch 1995), so it is possible to miss the presence of species if their populations are small or if only a few monitoring collectors are deployed Botryllus schlosseri Botryllus schlosseri was present at 4 of 11 monitoring stations in 2006, and 10 of 21 stations in 2007, throughout the region. Stepping-stone like introductions, probably

38 30 resulting from movement of fouled boat hulls or salmon aquaculture gear between monitoring stations, were evident on Deer and Campobello Islands in 2008 and 2009, and in the L Etete-Wallace Cove area and St. Andrews area in By 2011 only 3 stations (Musquash, Wallace Cove and Indian Island) were free of this species. Given the high degree of current and tidal exchange (Chang et al. 2005) in the region, B. schlosseri, and other colonial tunicates such as B. violaceus and D. vexillum, may arrive at new locations as tissue fragments carried by currents that may reattach and grow (Bullard et al. 2007b; Carmen et al. 2014), in addition to the anthropogenic vectors of introduction mentioned previously. The non-detection of B. schlosseri in Back Bay (Stn 141) and at Fairhaven (Stn 149) from year-to-year may be indicative of its low age at most stations in the region, and failure of the limited number of monitoring plates deployed at stations to reliably detect the presence of small tunicate populations Botrylloides violaceus The presence of B. violaceus in this region was documented for the first time in 2009 (Martin et al. 2011), at Head Harbour on Campobello Island. By 2010, it had spread to another site, Wilson s Beach on Campobello Island, and was also noted at Beaver Harbour on the southwestern coast of NB. It was detected at all three sites in Annual monitoring through the summer of 2015 indicates that B. violaceus has spread and very small populations are present throughout the region: at Beaver Harbour, and further east on the coastline to Dipper Harbour, in Back Bay, at Leonardville on Deer Island, at both monitoring sites on Campobello Island, and at all monitoring sites on Grand Manan (Sephton et al. In prep.). As noted above, there are many vectors of introduction and spread in the region, so there is potential for the arrival of B. violaceus at the four sentinel sites (Musquash, L Etete, St. Andrews and Fairhaven) currently free of the species Non-detection of Styela clava and Didemnum vexillum Martin et al. (2011) noted that the ranges of salinity and water temperature at the sites investigated between 2006 and 2009 were well within the environmental tolerances of the five species monitored, so it is possible for S. clava and D. vexillum to survive and establish if they are introduced. While both species are present in New England (Moore et al. 2014; Dijkstra et al. 2007a) and NS waters (Moore et al. 2014, Vercaemer et al. 2015) neither were observed during the course of this study. Styela clava has been present in PEI, on mussel aquaculture sites since 1998 (Clarke and Therriault 2007). Shellfish aquaculture is not established in swnb other than at experimental IMTA sites in the Lime Kiln Bay area, so there is little opportunity

39 31 for the species to be introduced through contaminated mussel seed or gear. This species was detected in NS in 2012 (Moore et al., 2014) in three high-traffic ports: Halifax Harbour (Vercaemer et al. 2014), Lunenburg Harbour and near Mulgrave, in Chedabucto Bay, so there is now potential for introduction by shipping and recreational vessels from NS (see Lacoursiere-Roussel et al. 2012), as well as from US waters. The potential introduction of D. vexillum into the region is a more immediate concern, given its presence nearby at Eastport, Maine, the existence of suitable habitat throughout the region (Therriault and Herborg 2007), and ecological concerns associated with the establishment of this species (Herborg et al. 2009; Valentine et al. 2007b; Vercaemer et al. 2015). Rapid assessment surveys conducted annually in the waters of swnb in 2009 (Martin et al 2010) through 2012 (Sephton and Vercaemer 2015) in an attempt to broaden detection efforts for D. vexillum beyond plate monitoring in nearshore areas did not record the species, nor did ongoing plate monitoring from 2012 through 2015 (Sephton et al., In prep.), although a native species, D. albidum, was present (Sephton and Vercaemer 2015). Following its disy in the upper BoF, in NS waters in 2013 (Moore et al. 2014), surveys were conducted throughout the BoF, and on Georges, German and northern Browns Banks (Vercaemer et al. 2015) to delineate its presence in eastern Canadian waters. No D. vexillum was found off Grand Manan, Campobello or Deer Islands during these surveys. It is possible that populations may be absent, or very small in this area (Vercaemer et al. 2015). It is also possible that the current DFO monitoring protocols are insufficient for the detection of small, isolated patches of D. vexillum or that the shallow intertidal habitats where monitoring efforts are concentrated are not prime habitat for this species. Future monitoring efforts in the region will incorporate the use of monitoring ground-lines successfully tested in the Minas Basin in 2014 (Vercaemer et al. 2015) on gravel and rock substrates in deeper water. 4.2 SPATIAL AND TEMPORAL VARIATION IN TUNICATE DISTRIBUTIONS Spatial Variation Variation Among Collectors Variation in the presence and age of C. intestinalis, B. schlosseri and B. violaceus was evident during monitoring conducted between 2006 to 2009 (LeGresley et al. 2008; Martin et al. 2011) and in 2010 and 2011, among plates of the same collector, and between duplicate collectors deployed during the same time period. This type of spatial variation has also been noted in monitoring in NS (Sephton et al., 2011, 2014, 2015) and reflects the localized (clumped) settlement patterns of tunicates that result from their short larval lifespans (<24hrs), and dependence on water current for dispersal due to their poor swimming abilities (Carver et al. 2006a, b). Thus, it is important that

40 32 duplicate collectors are deployed during each monitoring period to facilitate species detection. This change in the monitoring protocol was implemented in the region in 2012 with the deployment of duplicate collectors during the Full (May October) and Second (August October) deployment periods. Given the patchy nature of small tunicate populations, it is also important to examine submerged structures and surfaces on site, increasing the potential for detection of small populations Range Expansions While some inconsistencies in the number and locations of stations monitored in 2006 and 2007 make it difficult to track potential range expansions in this region, C. intestinalis has been present at most stations (~90%), and all areas (PQB, BoF coast, Deer, Campobello and Grand Manan Islands) monitored from 2008 to Botryllus schlosseri has also been present at one or more stations in all areas since 2006, but it was noted at more stations on the BoF coast from 2009 onward. Range expansion is evident in the distribution of B. violaceus, first detected on Campobello Island in 2009, followed by one site on the BoF coast (Beaver Harbour) in 2010 and a second site on Campobello in Indeed this species has continued to spread: to Grand Manan and on the BoF coast in 2012, to Deer Island and a second location on both Campobello and Deer Islands in 2013, and finally to SABS in PQB in 2015 (Sephton et al., In prep.). This type of range expansion may reflect vessel or gear (aquaculture) mediated bay-tobay spread al Variation Seasonal Variation First deployment collectors (May August) were deployed between 2006 and 2009 during this study (Martin et al. 2011), but this practice was discontinued in 2010 due to no, or low, tunicate settlement (see Appendices 1-6). During 2006, 2007 and 2009, less than 8 % of all first deployment plates or petri collectors had tunicate settlement. This may reflect colder spring and early summer temperatures in this region (LeGresley et al. 2008, Martin et al. 2011) compared to the warmer waters of the southwest and south shores of NS nearby (Sephton et al. 2011, 2014, 2015), where tunicates are frequently observed on First deployment collectors. Many First deployment collectors were also lost or damaged as weather conditions were often harsh early in the season in swnb. In 2008, however, the incidence of settlement on First deployment plates was 28%, and C. intestinalis was detected at three stations (Beaver Harbour, St. Andrews, and SABS) and B. schlosseri was detected at Beaver Harbour. Martin et al. (2011) also noted increased age by C. intestinalis on Full deployment collectors in 2008 compared with other years, and attributed this to a change in the water temperature regime in 2008, with a peak in September instead of the usual August peak, allowing a longer growth period for the tunicates. It is possible then, that waters were also warmer

41 33 early in 2008, and this may have contributed to the settlement of tunicates during the First deployment period. The removal of first deployment collector from the protocol in 2010 should not influence the annual detection of tunicates, based on Second and Full deployment collectors only, however, as there were no instances between 2006 and 2009 where C. intestinalis or B. schlosseri were detected on First Deployment collectors, and not on Second deployment and/or Full deployment collectors (Appendices 1-6). Tunicate age on Second deployment collectors (August October) was similar to, or a bit lower than on Full deployment (May October) collectors (Appendices 1-6), indicating that most settlement by tunicates in this region occurs between August and October. In NS, Full deployment collectors at many locations on the southwest and south shores are saturated with the dominant tunicate species present, usually C. intestinalis (Sephton et al. 2011, 2014, 2015), and less abundant species present may not be detected on collector surfaces due to a lack of available settlement space. The deployment of fresh plates in August may facilitate the detection of these less abundant tunicates, or those which settle later when the waters are warmer, and on surfaces not already fouled by species able to tolerate cooler waters Year-to-Year Variation There was year-to-year variation in the presence, and age, of C. intestinalis, B. schlosseri and B. violaceus, similar to that noted between 2006 and 2009 by Martin et al (2011). Much of this variation, or absence, for example; C. intestinalis at the Fairhaven Aquaculture site in 2008, may be a reflection of the small populations present, and consequent low ages seen on monitoring plates. Non-detection may also result from inadequacies of the monitoring protocol itself, such as the deployment of four monitoring collectors only, although collectors used in this region had both petri dish and plates as settlement surfaces, which enhanced the detection of less abundant species (Martin et al. 2011). It is important to deploy duplicate collectors at mid-season (Second deployment) facilitating the detection of less abundant species on fresh plates, and to examine a variety of submerged structures to detect the presence of tunicates when visiting monitoring stations. Finally, all collector data gathered should be used in the reporting of a species presence at a monitoring site. Martin et al. (2011) based their reporting on Full deployment collectors only, overlooking the very low presence of C. intestinalis at Seal Cove in 2006 and at Dipper Harbour in 2008 on Second deployment collectors.

42 34 Annual variation in the presence and age of tunicates in a region may also be related to environmental factors such as temperature and salinity (see references in Carver et al. 2006a, b). Coverage of C. intestinalis, at all stations monitored between 2006 and 2011 (Figures 8-13) was generally low in 2006, 2007 and 2009, low-tomoderate in 2008 and low-to-moderate-to-abundant in 2010 and Examination of the year to year trends at sentinel stations (Table 8) indicates that 2010 was the year of highest age at the most stations, followed by 2011 and then Coverage by B. schlosseri, which was generally low in all years (Figures 14-19, Table 6) including 2010, was a bit higher in 2008 and 2009 when some moderate ages were noted. It is possible that different species populations react differently to increases in temperature, or that some species with lower infestation levels require several consecutive warmer years to display increases in abundance. Martin et al. (2011) noted a change in the temperature regime in 2008 when temperature peaked later in the season in September. Robinson et al. (1996) characterized monthly temperature and salinity in this region between 1990 and 1995, and noted that there were four groupings of similar temperatures in the area, two of which encompassed sites monitored during the present study. The St. Croix Estuary cluster (Stns 151 and 152) was characterized by greater extremes and higher variability in temperature (and salinity), while the cluster offshore and outside Passamaquoddy Bay (Stns 138, , 142, 143 and possibly 144) was characterized by lower extremes and less variability. For example, winter water temperature was often below zero in the St. Croix estuary, but rarely below zero outside Passamaquoddy Bay. Additional environmental factors may facilitate heavier tunicate settlement at specific stations or within an area. The warmer area outside Passamaquoddy Bay, which includes Back Bay, is also an important site for salmon aquaculture (LeGresley et al. 2008), which may provide additional artificial substrates (Tyrell and Byers 2007) for tunicate settlement, and high nutrient loads (Lander et al. 2004) to enhance tunicate growth. Water currents can also influence tunicate settlement; C. intestinalis thrives in areas of low flow or minimal wave exposure (see Carver et. al. 2006a), giving it a competitive advantage in areas such as harbours and marinas (Carver et al. 2006a). This may explain its higher ages on monitoring plates in sheltered harbours such as Beaver and Head Harbours and at Leonardville, and the absence of tunicates at Musquash, where current is high, salinity is low, and water is silty. In comparison with the ages seen nearby on the southwest and south coasts of NS (Sephton et al. 2011, 2014, 2015), year-to year ages of C. intestinalis and B. schlosseri are generally lower in swnb. Whether this is a result of the harsher winters, cooler summer water temperatures, or greater current activity (tidal and regional circulation) in the region, or a combination of these factors, is not clear.

43 Table 6: Year average prevalence of Ciona intestinalis, and Botryllus schlosseri at sentinel stations in NB in 2006 through nm= not monitored. Categories for the percentage were: 0: (absent), 1: <25% (low), 2: 25-50% (moderate), 3: 51-75% (heavy), % and 4 (very heavy). Red and green lines indicate increasing and decreasing abundance trends, respectively. SW NB = southwest New Brunswick, Deer I. = Deer Island, Camp. I. = Campobello Island and Grand Manan I. = Grand Manan Island. 35

44 CONCLUDING REMARKS Biofouling monitoring conducted in southwest NB between 2006 and 2011 has enabled the following conclusions: Ciona intestinalis and B. schlosseri are well established throughout the region, and by 2011, only the monitoring station at Musquash, where environmental conditions are unfavorable, was free of these tunicates. There was a pattern of increasing age of these species at many stations in all areas of the region from 2006 to 2011, indicative of spread and establishment. Stations with highest age of C. intestinalis, the dominant tunicate in the region, are at sheltered sites at Beaver Harbour, Back Bay, SABS, Fairhaven, Leonardville and Head Harbour. Coverage by B. schlosseri is generally low in the monitoring area. Botrylloides violaceus was recorded for the first time in the region in 2009 on Campobello Island, and was present at three monitoring stations on Campobello and Deer Island in Its age is very low at these stations, often only single, small colonies were observed on monitoring plates. Didemnum vexillum and S. clava, present in nearby waters, have not yet been detected in the area, but there is a risk of introduction and spread for both species given the presence of suitable habitat and vectors of introduction. The following recommendations should be addressed in future work in this region: Given the high risk of introduction of invasive tunicates to the region, based on vectors of introduction and spread, biofouling monitoring should continue at all sentinel stations. The deployment of additional monitoring collectors during Second and Full deployment periods should be considered to facilitate the detection of small tunicate populations at many stations Due to the high likelihood of the introduction and establishment of D. vexillum in the abundant, suitable habitat in the region, alternate monitoring devices, such as ground-lines, should be deployed to strengthen and expand detection efforts for this species. If IMTA and shellfish aquaculture develops in the area, it will be important to expand monitoring efforts as new sites are approved, due to the risk of introduction of S. clava from other aquaculture areas.

45 37 ACKNOWLEDGEMENTS We are grateful to the following individuals and organizations who monitored stations and provided data: - Bruce Thorpe and Kathy Cleghorn; New Brunswick Department of Agriculture, Aquaculture and Fisheries - Wayne Miner, Danny Loveless, Perry Smith, Paul McCurdy, Joanne Power and Gillian Forbes; DFO, SABS - IMTA site operators at Fairhaven and Charlie Cove Thomas Sephton, Benedikte Vercaemer, Angelica Silva and Claudio DiBacco reviewed an early version of this report and provided many useful suggestions for its improvement. Funding for this monitoring survey was provided by Fisheries and Oceans Canada, Aquatic Invasive Species program.

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51 43 scallop fisheries. J. Exp. Mar. Biol. Ecol. 342: Van Name, W.G Simple ascidians of the coasts of New England and neighboring British provinces. Proc. Boston Soc. Nat. Hist. 34: Van Name, W.G The North and South American Ascidians. Bull. Am. Mus. Nat. Hist. 84: Vercaemer, B., Sephton, D., Clement, P., Harman, A., Stewart-Clark, S., and DiBacco, C Distribution of the non-indigenous colonial ascidian Didemnum vexillum (Kott, 2002) in the Bay of Fundy and on offshore banks, eastern Canada. Man. Biol. Invas. 6: Vercaemer, B. and Sephton, D Rapid assessment and early monitoring of Halifax harbour and Bedford Basin following the detection of three new marine invasive species. Can. Tech. Rep. Fish. Aquat. Sci., 3063: v + 28 p.

52 Appendix 1: Details of monitoring and tunicate age on individual collectors at each station in Stations are grouped by geographical region or Bay. Species ages are given for top, middle and bottom plates, (x,y, z) followed by four petri plates (A, B,C and D) (x,y,z,a,b,c,d), where 0 = no tunicate age, 1 = <25% age, 2 = 26-50% age, 3 = 51-75% age, abd 4 = % age. C.i. = Ciona intestinalis, B.s. = Botryllus schlosseri and B. v. = Botrylloides violaceus, and L = collector, plate or Petri dish lost. 44

53 Appendix 1, continued. Region Deer Island Campobello Island Grand Manan Island Stn Location No. Monitor By Deployment Dates Deploy Period (days) C.i. B.s. D.v. 150 Fairhaven MF NBDAAF First (1 Jun Aug.) 77 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (17 Aug Oct.) 71 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (1 Jun Oct.) 148 1,1,1,2,0,1,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (1 Jun Oct.) 148 1,2,0,1,2,2,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 144 Head Harbour NBDAAF First (1 Jun Aug.) 77 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (17 Aug Oct.) 71 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (1 Jun Oct.) 148 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (1 Jun Oct.) Harbour de Loutre NBDAAF First (1 Jun Aug.) 77 Sec. (17 Aug Oct.) 71 0,0,0,1,1,L,L 0,0,0,1,0,L,L 0,0,0,0,0,L,L Full (1 Jun Oct.) 148 Full (1 Jun Oct.) North Head NBDAAF First (5 Jul Aug.) 49 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (23 Aug Nov.) 90 Full (5 Jul Nov.) 139 Full (5 Jul Nov.) Seal Cove NBDAAF First (5 Jul Aug.) 49 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 * = on saucer Sec. (23 Aug Nov.) 90 1*,0,0,0,0,0, 1,0,0,1,0,0,0 0,0,0,0,0,0,0 Full (5 Jul Nov.) 139 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (5 Jul Nov.) 139 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 45

54 Appendix 2: Details of monitoring and tunicate age on individual collectors at each station in Stations are grouped by geographical region or Bay. Species ages are given for top, middle and bottom plates, (x,y,z) followed by four petri plates (A, B,C and D) (x,y,z,a,b,c,d), where 0 = no tunicate age, 1 = <25% age, 2 = 26-50% age, 3 = 51-75% age and 4 = % age. C.i. = Ciona intestinalis, B.s. = Botryllus schlosseri and B. v. = Botrylloides violaceus, and L = collector, plate or Petri dish lost. Region sw NB Stn. No. Location Monitor By Deployment Dates Deploy Period (days) C.i. B.s. B.v. 164 Kennebecasis YC DFO First (20 Jun Aug.) 57 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (16 Aug. - 1 Oct.) 46 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (10 Jun. - 1 Oct.) 103 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 165 Saint John DFO First (20 Jun Aug.) 57 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (16 Aug. - 1 Oct.) 46 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (10 Jun. - 1 Oct.) 103 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 133 Dipper Harbour DFO First (12 Jun Aug.) 65 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (16 Aug Oct.) 67 0,0,0,0,0,0,0 2,1,2,2,1,1,3 0,0,0,0,0,0,0 Full (12 Jun Oct.) 132 0,0,0,0,0,0,0 1,1,2,1,1,1,1 0,0,0,0,0,0,0 139 Lime Kiln Bay NBDAAF First (22 Jun Aug) Sec. (17 Aug - 17 Oct.) Full (22 Jun Oct.) ,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 135 Beaver Harbour DFO First (12 Jun Aug.) 65 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (16 Aug Oct.) 67 0,0,0,0,1,0,0 1,1,2,1,1,1,1 0,0,0,0,0,0,0 Full (12 Jun Oct.) 132 0,0,1,1,0,1,0 0,1,1,2,1,3,1 0,0,0,0,0,0,0 141 Back Bay NBDAAF First (14 Jun. - 1 Aug.) 48 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (1 Aug Oct.) 78 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (14 Jun Oct.) 126 1,0,0,1,1,1,1 0,0,0,1,1,0,1 0,0,0,0,0,0,0 138 Wallace Cove NBDAAF First (14 Jun. - 1 Aug.) Sec. (1 Aug Oct.) Full (14 Jun Oct.) ,1,0,1,1,1,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 142 Charlie Cove DFO First (7 Jun Aug.) 64 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (10 Aug Oct.) 81 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (7 Jun Oct.) 145 1,1,1,1,1,2,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 46

55 Appendix Appendix 2, continued 1, continued. Region sw NB Deer Island Campobello Island Stn No. Location Monitor By Deployment Dates Deploy Period (days) C.i. B.s. D.v. 143 L'Etete NBDAAF First (14 Jun. - 1 Aug.) 48 Sec. (1 Aug Oct.) 78 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (14 Jun Oct.) 126 0,0,0,1,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 148 Hog Island NBDAAF First (19 Jun Aug.) Sec. (21 Aug Oct.) Full (19 Jun Oct.) ,0,1,1,1,1,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 151 St. Andrews DFO First (11 Jun Aug.) 60 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (10 Aug Oct.) 74 1,1,1,0,0,1,0 0,0,0,0,1,0,1 0,0,0,0,0,0,0 Full (11 Jun Oct.) 134 1,0,1,0,0,0,1 0,0,0,2,1,1,0 0,0,0,0,0,0,0 152 SABS DFO First (11 Jun Aug.) 60 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (10 Aug Oct.) 74 1,0,1,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (11 Jun Oct.) 134 1,1,1,0,0,1,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 149 Fairhaven Dock NBDAAF First (19 Jun. - 1 Aug.) Sec. (1 Aug Oct.) Full (19 Jun Oct.) 150 Fairhaven MF NBDAAF First (19 Jun. - 1 Aug.) Sec. (1 Aug Oct.) Full (19 Jun Oct.) ,1,1,2,1,1,0 0,0,0,1,1,1,0 2,1,0,2,1,0,1 0,1,0,1,0,1,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 146 Leonardville NBDAAF First (22 Jun. - 1 Aug.) 40 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (1 Aug Oct.) 77 Full ( Jun Oct.) 117 1,1,1,1,1,1,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 144 Head Harbour NBDAAF First (19 Jun. - 1 Aug.) 43 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (1 Aug Oct.) 77 Full (19 Jun Oct.) 120 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 47

56 Appendix 1, continued Region Stn No. Location Monitor By Deployment Dates Deploy Period C.i. B.s. D.v. Campobello Island Grand Manan Island 215 Roosevelt Park NBDAAF First (7 Jun. - 1 Aug.) 55 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (1 Aug Oct.) 76 Full ( 7 Jun Oct.) 131 0,0,0,1,1,1,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 163 Harbour de Loutre DFO First (7 Jun. - 1 Aug.) 55 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (1 Aug Oct.) 76 Full (7 Jun Oct.) 131 0,0,0,1,1,0,0 2,1,1,3,1,2,2 0,0,0,0,0,0,0 136 North Head DFO First (25 Jun Aug.) 59 Sec. (23 Aug Oct.) 68 1,1,0,0,0,0,0 1,1,1,1,1,1,1 0,0,0,0,0,0,0 Full (25 Jun Oct.) 126 1,1,0,0,0,0,0 1,1,2,1,1,1,1 0,0,0,0,0,0,0 137 Ingalls Head DFO First (25 Jun Aug.) 59 Sec. (23 Aug Oct.) 68 0,0,0,0,0,0,0 2,1,2,1,1,1,0 0,0,0,0,0,0,0 Full (25 Jun Oct.) 126 0,0,0,0,0,0,0 1,1,1,0,1,0,1 0,0,0,0,0,0,0 140 Seal Cove DFO First (25 Jun Aug.) 59 Sec. (23 Aug Oct.) 68 0,0,0,0,0,0,0 2,1,0,1,1,1,1 0,0,0,0,0,0,0 Full (25 Jun Oct.) 126 1,1,1,0,0,1,0 1,1,0,2,2,1,1 0,0,0,0,0,0,0 48

57 Appendix 3: Details of monitoring and tunicate age on individual collectors at each station in Stations are grouped by geographical region or Bay. Species ages are given for top, middle and bottom plates, (x,yz) f ollowed by four petri plates (A, B,C and D) (x,y,z,a,b,c,d), where 0 = no tunicate age, 1 = <25% age, 2 = 26-50% age, 3 = 51-75% age, and 4 = % age. C.i. = Ciona intestinalis, B.s. = Botryllus schlosseri and B. v. = Botrylloides violaceus, and L = lost. Region sw NB Stn. No. Location Monitor By Deployment Dates Deploy Period (days) C.i. B.s. B.v. 132 Musquash DFO First (18 Jul Aug.) 34 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (21 Aug. - 6 Nov.) 77 Full (18 Jul. - 6 Nov.) 111 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 133 Dipper Harbour DFO First (6 Jun Aug.) 76 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (21 Aug. - 6 Nov.) 77 0,1,0,0,0,1,0 1,0,1,1,1,0,1 0,0,0,0,0,0,0 Sec. (21 Aug. - 6 Nov.) 77 0,0,0,0,0,0,0 1,1,1,1,1,1,1 0,0,0,0,0,0,0 135 Beaver Harbour DFO First (6 Jun Aug.) 76 0,0,0,1,1,1,1 1,2,1,3,2,1,3 0,0,0,0,0,0,0 Sec. (21 Aug. - 6 Nov.) 77 0,1,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (6 Jun. - 6 Nov.) 153 0,0,0,1,1,1,1 0,0,0,1,1,1,1 0,0,0,0,0,0,0 139 Lime Kiln Bay NBDAAF First (23 Jun Aug.) Sec. (14 Aug. - 4 Nov.) Full (23 Jun. - 4 Nov.) ,1,2,3,2,3,2 0,0,0,0,0,0,0 0,0,0,0,0,0,0 141 Back Bay NBDAAF First (10 Jun Aug.) 62 Sec. (11 Aug Oct.) 79 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (10 Jun Oct.) 141 0,0,0,1,1,1,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 138 Wallace Cove NBDAAF Sec. (11 Aug Oct.) 79 0,0,0,3,4,3,3 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (10 Jun Oct.) 141 0,0,0,4,4,2,4 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (10 Jun Oct.) 141 1,1,1,4,4,2,4 0,0,0,0,0,0,0 0,0,0,0,0,0,0 142 Charlie Cove NBDAAF First (25 Jun Aug.) 55 Sec. (19 Aug. - 4 Nov.) 77 0,1,1,3,3,1,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (25 Jun. - 4 Nov.) 132 2,1,1,3,3,2,2 0,0,0,0,0,0,0 0,0,0,0,0,0,0 143 L'Etete NBDAAF Sec. (11 Aug Oct.) 79 0,1,0,1,1,1,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (10 Jun Oct.) 141 0,0,1,L,L,1,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (10 Jun Oct.) 141 1,1,1,1,1,2,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 49

58 Appendix 3, continued. Region sw NB Deer Island Stn Location No. Monitor By Deployment Dates Deploy Period (days) C.i. B.s. D.v. 148 Hog Island First (20 Jun Aug.) 59 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (18 Aug Oct.) 73 Full (20 Jun Oct.) 132 0,0,0,0,0,0,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 151 St. Andrews DFO First (6 Jun Aug.) 76 1,1,1,1,1,1,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (21 Aug. - 6 Nov.) 77 1,0,0,0,0,0,0 0,1,0,0,0,0,0 0,0,0,0,0,0,0 Full (6 Jun. - 6 Nov.) 153 2,2,3,2,2,2,1 1,0,0,0,0,0,0 0,0,0,0,0,0,0 152 SABS DFO First (6 Jun Aug.) 76 1,1,0,L,L,L,L 0,0,0,L,L,L,L 0,0,0,L,L,L,L Sec. (21 Aug. - 6 Nov.) 77 1,1,0,1,1,1,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (6 Jun. - 6 Nov.) 153 1,2,4,4,2,3,3 0,0,0,0,0,0,0 0,0,0,0,0,0,0 149 Fairhaven Dock NBDAAF Sec. (19 Aug Oct.) 72 0,1,1,1,1,1,1 1,1,1,1,1,1,1 0,0,0,0,0,0,0 Full (10 Jun Oct.) 142 1,1,1,1,1,1,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (10 Jun Oct.) 142 1,1,1,1,1,1,3 1,1,1,1,1,1,0 0,0,0,0,0,0,0 150 Fairhaven MF NBDAAF First (10 Jun Aug.) Sec. (19 Aug Oct.) Full (10 Jun Oct.) ,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 146 Leonardville NBDAAF Sec. (18 Aug Oct.) 73 0,1,1,0,0,1,1 0,1,1,0,0,0,0 0,0,0,0,0,0,0 Full (10 Jun Oct.) 142 2,1,1,1,3,2,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (10 Jun Oct.) 142 2,2,2,1,3,3,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 147 Indian Island NBDAAF First (13 Jun Aug.) 66 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (18 Aug. - 4 Nov.) 78 0,0,0,1,2,1,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Full (13 Jun. - 4 Nov.) 144 1,0,1,1,1,1,1 0,0,0,0,0,0,0 0,0,0,0,0,0,0 50

59 Appendix 3, continued Region Campobello Island Grand Manan Island Stn No. Location Monitor By Deployment Dates Deploy Period (days) C.i. B.s. D.v. 144 Head Harbour NBDAAF First (13 Jun Aug.) 67 Sec. (19 Aug. - 4 Nov.) 77 0,0,0,1,1,1,0 0,0,0,1,1,1,0 0,0,0,0,0,0,0 Full (13 Jun. - 4 Nov.) 144 2,2,2,3,4,2,1 1,0,0,1,0,0,0 0,0,0,0,0,0,0 163 Harbour de Loutre NBDAAF First (13 Jun Aug.) 67 no data no data no data Sec. (19 Aug. - 4 Nov.) 77 1,1,1,3,2,1,1, 1,1,2,0,1,1, 0,0,0,0,0,0,0 Full (13 Jun. - 4 Nov.) 144 1,1,1,2,3,2,1 1,0,1,1,0,0,1 0,0,0,0,0,0,0 145 Wilson's Beach NBDAAF First (13 Jun Aug.) Sec. (18 Aug. - 4 Nov.) Full (13 Jun. - 4 Nov.) ,0,0,1,1,1,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 136 North Head NBDAAF Sec. (13 Aug Oct.) 78 0,0,0,1,1,1,0 1,1,1,1,1,1,1 0,0,0,0,0,0,0 Full (26 Jun Oct.) 126 0,1,0,1,0,0,1 1,1,1,1,1,1,2 0,0,0,0,0,0,0 Full (26 Jun Oct.) 126 0,1,0,1,0,0,1 0,1,1,0,0,0,1 0,0,0,0,0,0,0 137 Ingall's Head NBDAAF Sec. (13 Aug Oct.) 78 0,0,0,0,0,0,0, 1,0,1,3,3,1,3 0,0,0,0,0,0,0 Full (26 Jun Oct.) 126 1,1,0,0,0,0,0 3,2,4,0,0,0,0 0,0,0,0,0,0,0 Full (26 Jun Oct.) 126 0,1,1,1,0,1,1 4,1,4,1,1,10 0,0,0,0,0,0,0 140 Seal Cove NBDAAF First (26 Jun Aug.) 48 0,0,0,0,0,0,0 0,0,0,0,0,0,0 0,0,0,0,0,0,0 Sec. (13 Aug Oct.) 78 Full (26 Jun Oct.) 126 0,1,1,1,0,1,1 1,2,1,1,1,1,1 0,0,0,0,0,0,0 51

60 Appendix 4: Details of monitoring and tunicate age on individual collectors at each station in Stations are grouped by geographical region or Bay. Species ages are given for top, middle and bottom plates, (x,y, z) followed by three petri plates (A, B, and C) (x,y,z,a,b,c), where 0 = no tunicate age, 1 = <25% age, 2 = 26-50% age, 3 = 51-75% age, and 4 = % age. C.i. = Ciona intestinalis, B.s. = Botryllus schlosseri and B.v.= Botrylloides violaceus, and L = collector, plate or Petri dish lost. Region sw NB Stn. No. Location Monitor By Deployment Dates Deploy Period (days) C.i. B.s. B.v. 132 Musquash DFO First (21 May - 12 Aug.) 83 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 First (21 May - 12 Aug.) 83 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (12 Aug. - 9 Nov.) 89 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (12 Aug. - 9 Nov.) 89 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Full (21 May - 9 Nov.) 172 Full (21 May - 9 Nov.) 172 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 133 Dipper Harbour DFO First (21 May - 12 Aug.) 83 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 First (21 May - 12 Aug.) 83 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (12 Aug. - 9 Nov.) 89 0,0,0,0,0,0 2,2,2,2,3,4 0,0,0,0,0,0 Sec. (12 Aug. - 9 Nov.) 89 0,0,0,0,0,0 1,1,1,2,2,2 0,0,0,0,0,0 Full (21 May - 9 Nov.) 172 0,0,0,0,0,0 2,1,1,2,3,2 0,0,0,0,0,0 Full (21 May - 9 Nov.) 172 0,0,0,0,0,0 3,2,2,3,2,3 0,0,0,0,0,0 135 Beaver Harbour DFO First (21 May - 12 Aug.) 83 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 First (21 May - 12 Aug.) 83 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (12 Aug. - 9 Nov.) 89 Sec. (12 Aug. - 9 Nov.) 89 1,0,1,1,1,1 3,4,4,3,3,3 0,0,0,0,0,0 Full (21 May - 9 Nov.) 172 Full (21 May - 9 Nov.) 172 3,2,1,1,3,2 2,2,3,3,1,2 0,0,0,0,0,0 139 Lime Kiln Bay NBDAAF First (26 May - 6 Aug.) 72 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 First (26 May - 6 Aug.) 72 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (6 Aug Oct.) 77 Sec. (6 Aug Oct.) 77 0,1,1,3,4,3 0,0,0,0,0,0 0,0,0,0,0,0 Full (26 May - 22 Oct.) 149 3,3,2,3,1,2 0,0,0,0,0,0 0,0,0,0,0,0 Full (26 May - 22 Oct.)

61 Appendix 4, continued Region sw NB Stn. No. Location Monitor By Deployment Dates Deploy Period (days) C.i. B.s. B.v. 141 Back Bay NBDAAF First (26 May - 6 Aug.) 72 0,0,0,L,L,L 0,0,0,L,L,L 0,0,0,L,L,L First (26 May - 6 Aug.) 72 0,0,0,L,L,L 0,0,0,L,L,L 0,0,0,L,L,L Sec. (6 Aug Oct.) 77 1,0,0,1,0,0 1,1,0,1,1,1 0,0,0,0,0,0 Sec. (6 Aug Oct.) 77 1,0,0,4,4,2 0,0,0,0,01 0,0,0,0,0,0 Full (26 May - 22 Oct.) 149 1,1,0,L,L,L 1,1,1,L,L,L 0,0,0,L,L,L Full (26 May - 22 Oct.) 149 L,L,L,L,2,L L,L,L,L,1,L L,L,L,L,0,L 138 Wallace Cove NBDAAF First (10 Jun. - 6 Aug.) 57 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 First (10 Jun. - 6 Aug.) 57 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (6 Aug Nov.) 116 Sec. (6 Aug Nov.) 116 0,0,0,1,1,1 0,0,0,0,0,0 0,0,0,0,0,0 Full (10 Jun Nov.) 173 0,0,1,1,2,1 0,0,0,0,0,0 0,0,0,0,0,0 Full (10 Jun Nov.) 173 1,1,1,2,1,2 0,0,0,0,0,0 0,0,0,0,0,0 142 Charlie Cove NBDAAF First (26 May - 11 Aug.) 77 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 First (26 May - 11 Aug.) 77 1,1,1,0,1,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (11 Aug Oct.) 70 0,1,1,0,1,1 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (11 Aug Oct.) 70 0,1,0,1,1,1 0,0,0,0,0,0 0,0,0,0,0,0 Full (26 May - 20 Oct.) 147 1,1,1,0,1,1 0,0,0,0,0,0 0,0,0,0,0,0 Full (26 May - 20 Oct.) 147 1,1,1,1,1,1 0,0,0,0,0,0 0,0,0,0,0,0 143 L'Etete NBDAAF First (26 May - 4 Aug.) 70 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 First (26 May - 4 Aug.) 70 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (4 Aug Oct.) 79 0,0,0,1,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (4 Aug Oct.) 79 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Full (26 May - 22 Oct.) 149 0,0,0,0,1,0 0,0,0,0,0,0 0,0,0,0,0,0 Full (26 May - 22 Oct.) 149 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 148 Hog Island NBDAAF First (29 May - 4 Aug.) 67 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 First (29 May - 4 Aug.) 67 L,0,0,0,L,L L,0,0,0,L,L L,0,0,0,l,L Sec. (4 Aug Nov.) 115 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (4 Aug Nov.) 115 0,0,0,L,L,0 0,0,0,L,L,0 0,0,0,L,L,0 Full (29 May - 27 Nov.) 182 1,0,0,L,L,L 0,0,0,L,L,L 0,0,0,L,L,L Full (29 May - 27 Nov.) 182 0,0,0,0,L,0 0,0,0,0,L,0 0,0,0,0,L,0 53

62 Appendix 4, continued. 54

63 Appendix 4, continued. Region Deer Island Campobello Island Stn No. Location Monitor By Deployment Dates Deploy Period (days) C.i. B.s. B.v. 147 Indian Island NBDAAF First (29 May - 4 Aug.) 67 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 First (29 May - 4 Aug.) 67 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (4 Aug. - 2 Nov.) 90 0,1,1,1,2,2 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (4 Aug. - 2 Nov.) 90 1,1,1,1,3,1 0,0,0,0,0,0 0,0,0,0,0,0 Full (29 May - 2 Nov.) 157 0,0,0,1,1,1 0,0,0,0,0,0 0,0,0,0,0,0 Full (29 May - 2 Nov.) 157 0,0,0,2,1,1 0,0,0,0,0,0 0,0,0,0,0,0 144 Head Harbour NBDAAF First (29 May - 4 Aug.) 67 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 First (29 May - 4 Aug.) 67 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (4 Aug. - 5 Nov.) 93 1,1,1,1,1,1 3,4,3,2,3,3 0,0,0,0,0,0 Sec. (4 Aug. - 5 Nov.) 93 1,2,1,2,1,2 3,1,1,1,1,1 0,0,0,0,0,1 Full (29 May - 5 Nov.) 160 1,1,1,1,1,1 0,1,1,1,0,1 0,0,0,0,0,0 Full (29 May - 5 Nov.) 160 2,1,2,1,1,1 0,0,1,0,1,0 0,0,0,0,0,0 163 Harbour de Loutre NBDAAF First (29 May - 4 Aug.) 67 First (29 May - 4 Aug.) 67 Sec. (4 Aug. - 2 Nov.) 90 0,1,0,1,1,1 2,1,4,1,1,2 0,0,0,0,0,0 Sec. (4 Aug. - 2 Nov.) 90 Full (29 May - 2 Nov.) 157 Full (29 May - 2 Nov.) Wilson's Beach First (29 May - 4 Aug.) 67 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 First (29 May - 4 Aug.) 67 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (4 Aug. - 2 Nov.) 90 0,0,0,1,1,3 1,1,1,1,1,3 0,0,0,0,0,0 Sec. (4 Aug. - 2 Nov.) 90 1,1,1,1,1,1 1,1,1,1,1,1 0,0,0,0,0,0 Full (29 May - 2 Nov.) 157 0,0,0,1,1,0 0,1,0,1,0,0 0,0,0,0,0,0 Full (29 May - 2 Nov.)

64 Appendix 4, continued. Region Grand Manan Island Stn No. Location Monitor By Deployment Dates Deploy Period (days) C.i. B.s. B.v. 136 North Head NBDAAF First (3 Jun. - 5 Aug.) 65 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 First (3 Jun. - 5 Aug.) 65 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (5 Aug Oct.) 86 0,0,0,0,1,0 1,2,1,0,1,1 0,0,0,0,0,0 Sec. (5 Aug Oct.) 86 0,0,0,0,0,0 0,0,1,2,0,1 0,0,0,0,0,0 Full (3 Jun Oct.) 151 1,1,1,1,1,L 1,1,1,1,0,L 0,0,0,0,0,L Full (3 Jun Oct.) 151 1,1,1,1,0,0 1,1,1,1,1,1 0,0,0,0,0,0 137 Ingall's Head NBDAAF First (3 Jun. - 5 Aug.) 65 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 First (3 Jun. - 5 Aug.) 65 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (5 Aug Oct.) 86 Sec. (5 Aug Oct.) 86 1,1,0,1,1,1 1,1,1,3,2,1 0,0,0,0,0,0 Full (3 Jun Oct.) 151 0,0,1,1,1,1 1,1,3,1,2,1 0,0,0,0,0,0 Full (3 Jun Oct.) 151 0,0,0,0,1,1 2,4,2,1,1,1 0,0,0,0,0,0 140 Seal Cove NBDAAF First (3 Jun. - 5 Aug.) 65 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 First (3 Jun. - 5 Aug.) 65 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 Sec. (5 Aug Oct.) 86 0,0,0,0,1,0 4,2,2,3,2,2 0,0,0,0,0,0 Sec. (5 Aug Oct.) 86 0,0,0,1,1,L 1,1,1,0,2,L 0,0,0,0,0,L Full (3 Jun Oct.) 151 1,0,0,L,L,L 0,0,0,L,L,L 0,0,0,0,0,0 Full (3 Jun Oct.) 151 1,1,1,L,L,L 0,0,0,L,L,L 0,0,0,0,0,0 56

65 Appendix 5: Details of monitoring and tunicate age on individual collectors at each station in Stations are grouped by geographical region or Bay. Species ages are given for top, middle and bottom plates, (x,y,z) followed by four three plates (A, B, and C) (x,y,z,a,b,c), where 0 = no tunicate age, 1 = <25% age, 2 = 26-50% age, 3 = 51-75% age, and 4 = % age. C.i. = Ciona intestinalis, B.s. = Botryllus schlosseri and B.v.= Botrylloides violaceus, and L = collector, plate or Petri dish lost. * = Present on site. Region sw NB Stn. No. Location Monitor By Deployment Dates 132 Musquash DFO Sec. (12 Aug Nov.) Full (10 Jun Nov.) Full (10 Jun Nov.) 133 Dipper Harbour DFO Sec. (12 Aug. - 5 Nov.) Full (10 Jun. - 5 Nov.) Full (10 Jun. - 5 Nov.) 135 Beaver Harbou r DFO Sec. (12 Aug. - 5 Nov.) Full (10 Jun. - 5 Nov.) Full (10 Jun. - 5 Nov.) 139 Lime Kiln Bay DFO Sec. (27 Aug. - 2 Nov.) Full (15 Jun. - 2 Nov.) Full (15 Jun. - 2 Nov.) 141 Back Bay NBDAAF Sec.(13 Aug. - 2 Nov.) Full (7 Jun. - 2 Nov.) Full (7 Jun. - 2 Nov.) 138 Wallace Cove NBDAAF Sec. (13 Aug. - 2 Nov.) Full (7 Jun. - 2 Nov.) Full (7 Jun. - 2 Nov.) 143 L'Etete NBDAAF Sec. (13 Aug. - 2 Nov.) Full (7 Jun. - 2 Nov.) Full (7 Jun. - 2 Nov.) 153 Crow Island NBDAAF Sec. (13 Aug. - 2 Nov.) Full (7 Jun. - 2 Nov.) Full (7 Jun. - 2 Nov.) Deploy Period (days) C.i. 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,1,1,1 0,1,1,0,0,0 2,1,1,3,4,2 4,4,4,4,4,4 4,4,4,4,4,4 0,0,0,1,1,0 2,1,1,4,3,4 2,0,0,3,L,2 1,0,1,L,2,4 1,1,0,2,3,3 1,1,1,1,1,1 0,0,0,2,1,2 1,1,1,1,1,1 1,1,0,1,1,1 1,1,1,1,1,1 B.s. 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 4,2,2,1,2,2 3,2,3,1,1,1 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 3,1,1,1,1,1 1,1,0,0,L,0 1,0,1,L,1,0 1,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 B.v. 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 1*,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 0,0,0,0,0,0 57

66 Appendix 5, continued. 58