First records of marine invasive non-native Bryozoa in Norwegian coastal waters from Bergen to Trondheim

From 29 June to 25 July 2014, scientists from Heriot Watt University and the Natural History Museum, London, aboard the vessel MV Halton, undertook a research cruise along the Norwegian coast. The cruise started in Bergen and over the two-week period surveys were conducted at intervals along the coastline, heading northwards, and including the major ports of Ålesund, Kristiansund, and Trondheim. When the vessel moored up in each harbour, surveys of the local pontoons were conducted to identify fouling species and to ascertain whether any non-native Bryozoa were present. Seven species of fouling Bryozoa were identified. Two of these were the non-native species Tricellaria inopinata d’Hondt and Occhipinti Ambrogi, 1985 and Schizoporella japonica Ortmann, 1890. This study represents the first confirmed records for these species from Norwegian coastal waters. It is likely, given the locations of occurrence, that small boat traffic was a vector for the introduction of these species. Recommendations are given for the amendment and addition of species to the Norwegian Black List.


Introduction
The introduction of a non-indigenous species into a new geographic area can affect local biodiversity by causing a restructuring of the trophic web (Vitousek 1990) as well as directly affecting society through causing damage to marine infrastructures, such as the fouling of pipes, vessels and other submerged infrastructure, and by having negative effects on fisheries or aquaculture (Occhipinti Ambrogi et al. 2011).
In 2012, a comprehensive assessment of the alien species present in Norway was undertaken by researchers at the Norwegian Biodiversity Information Centre, Trondheim, and was published in a document entitled the Norwegian Black List (Gederaas et al. 2012). At this time, there were no records of non-native or invasive species of marine Bryozoa recorded in Norwegian coastal waters. In Table 10 of the report, four species of bryozoans were listed as 'Doorknockers'; i.e., species that were expected to arrive but had not yet been recorded. These species were: Watersipora subtorquata (d'Orbigny, 1852); Tricellaria inopinata d' Hondt and Occhipinti Ambrogi, 1985; Bugula neritina (Linnaeus, 1758); and Bugula stolonifera Ryland, 1960. From 29 June to 25 July 2014, scientists from Heriot Watt University and the Natural History Museum, London, undertook a research cruise aboard the vessel MV Halton. The cruise covered the region between Bergen (60°23.963'N, 05°18.7 40'E) and Trondheim (63°26.318'N,10°23.976'E). The aim was to survey the bryozoan communities and to identify whether any non-native species were present, in a range of different habitats. One part of the survey focused specifically on marinas and pontoons, to establish whether any of the 'Doorknocker' species had arrived in Norway. An open circle means that no fouling species were found, a closed circle means that invasive non-native species were found and a cross hatched circle means that fouling species were present but not invasive non-native species.

Methods
Visual surveys of pontoons and fenders were conducted at the seven locations ( Figure 1). The survey methodology was adapted from that used by Arenas et al. (2006) for rapid assessments of marinas. In this study, two expert bryozoan taxonomists (JSP and MSJ) conducted 30-minute searches at each site. Submerged surfaces of floating plastic pontoons and submerged fenders were inspected for the presence of marine bryozoans. Samples were identified in situ where possible; where this was not possible or in order to confirm identification by microscope, samples were collected and subsequently examined under a Wild M5 stereo dissection microscope. Representative material was selected for preparation for imaging on a LEO 1455-VP Scanning Electron Microscope (SEM) at the EMMA unit of the Natural History Museum, London. Selected specimens were deposited with the NHM Bryozoa section and these are listed with location information and accession numbers in Table 1.
Where identifications of fouling and non-native species were confirmed, the significance of the findings was investigated to quantify significant extensions to the known range.

Results and discussion
In this survey, seven species of bryozoans ( Figure 2) were identified from the study locations:
The first three of these species have been shown, using molecular genetic techniques, to exist as cryptic species complexes (Schwaninger 2008, Gomez et al. 2007Waeschenbach et al. 2012;Nikulina et al. 2007). The cyclostome Patinella verrucaria was growing on a colony of the bryozoan Tricellaria inopinata, which was growing attached to the plastic pontoons. The known invasive species observed were Tricellaria inopinata and Schizoporella japonica. The latter species was not present in the 'Doorknocker' list, whereas the former species was anticipated. These new records represent significant expansions to the known species' distributions and are discussed more fully below.
Tricellaria inopinata (Figure 3) We found Tricellaria inopinata growing on plastic pontoons at Florø and Kristiansund. These are the first known records for this species in Norway and represent a significant northerly extension to the range in North-west Europe based on records in the Orkney Isles, Scotland (Cook et al. 2013;Nall et al. 2015).  Tricellaria inopinata was first described in 1985 (d' Hondt and Occhipinti Ambrogi 1985) from specimens collected in May 1982 from waterways connected to the central region of the Lagoon of Venice. Earlier surveys did not document this species, leading to the hypothesis that it was a recent invader and it subsequently spread rapidly to other parts of the lagoon (Occhipinti Ambrogi 1991; Occhipinti Ambrogi et al. 2011). Tricellaria inopinata is thought to have originated in the north Pacific Ocean (Dyrynda et al. 2000;De Blauwe and Faasse 2001) and was later introduced into Australia, New Zealand, Japan, Taiwan, the West Pacific, and Venice (Occhipinti Ambrogi and d 'Hondt 1994). Just a few years later, it was reported from locations on the European Atlantic coast, including: France (Breton and d'Hondt 2005); Belgium; the Netherlands (De Blauwe and Faasse 2001); and the northwest of Spain and Portugal (in 2004;Marchini et al. 2007). By 1998, this invasive species was present in south-central England (Dyrynda et al. 2000). In the Netherlands it was found to be abundant on: natural surfaces; artificial surfaces such as ropes and buoys; and on other sessile fauna (De Blauwe and Faasse 2001). More recently, it was reported from Woods Hole, Massachusetts, USA, in September 2010 (Johnson et al. 2012). In 2012, it was observed in Dublin Bay, Ireland (Kelso and Wyse Jackson 2012).
Schizoporella japonica (Figure 4) In our survey, Schizoporella japonica was found for the first time in Norwegian coastal waters at the ports of Florø, Ålesund, and Kristiansund. It was growing on the plastic floats of the pontoons and also on the shells of Mytilus edulis that also were attached to the pontoons. Schizoporella japonica was originally described from Japan, and is known from the Hokkaido area (Grischenko et al. 2007). S. japonica was subsequently introduced on Pacific oysters to the Pacific coast of North America, where it is now well established (Powell, 1970;Dick et al. 2005;Blum et al. 2007;Crooks et al. 2011). It was recently recorded in the UK from Holyhead marina, North Wales, the south coast of England, and at several localities around the Scottish coastline including the mainland, Orkney and Shetland Isles (Ryland et al. 2014;Bishop et al. 2015;Nall et al 2015). This species should now be added to the Norwegian Blacklist.

Timing of arrival in Norway of the two invasive species
During this survey, Tricellaria inopinata and Schizoporella japonica were collected and identifications confirmed by SEM imaging (Figures 2 and 3). Examination of the published literature and museum collections has revealed no previous records of these species in Norway. This suggests that they have been introduced relatively recently. Vectors of transmission could be leisure or commercial craft travelling between ports in Norway, and between Norway, Shetland, the Faroes, and Iceland.
In the case of Tricellaria inopinata (a temperate waters species), it is likely that the species has spread northwards from more southerly locations in mainland Europe (De Blauwe and Faasse 2001) or by hopping across the North Sea from Scotland (Cook et al 2013). These are not mutually exclusive hypotheses.
In contrast, Schizoporella japonica is an Arctic/ Boreal species that has been documented on the east and west coasts of North America and more recently in the north mainland of Scotland, Orkney, and Shetland (Ryland et al 2014;Nall et al 2015). The species appears to be colonizing into Europe from the Atlantic in a west to east direction and then spreading along coastlines.
The marinas surveyed along the Norwegian coast fell into three categories: i) those with freshwater input and no bryozoans present; ii) fully saline with native or naturalized fouling species; or iii) fully saline with a mixture of native or naturalized fouling species and invasive nonnative species (Figure 1).

Update on other 'Doorknocker' invasive Bryozoa for Norway
Bugulina simplex (Hincks, 1886) ( Figure 5) Ryland et al. (2011) provide a comprehensive historical review of the distribution of Bugulina simplex in North west Europe, including localities around the British coastline, France, Belgium and the Netherlands. The most northerly record at that time was from Ouddorp in the Netherlands. The species is also present in the Eel Pond at Woods Hole, New England (Ryland 1960). Recently (September 2014), Bugulina simplex has been collected by Rachel Shucksmith, in Lerwick Harbour, Shetland (NHMUK 2014.11.1.12). While there are no records of this species as yet in Norway, with regular vessel traffic between Lerwick and Norwegian ports, it seems likely that it is only a matter of time before an introduction is made. This species is not currently listed as a Doorknocker in the Norwegian Blacklist (2012); however, given its current proximity and the rate of spread of the species, we suggest that it be added to the Blacklist.
Bugulina stolonifera (Ryland, 1960) (Figure 5) The distribution and historical records of B. stolonifera are reviewed in Ryland et al. (2011), and the status of the species in Western Europe is unclear. Its distribution appears to be very patchy and one of the more recent records is from the Outer Harbour, Harwich (Ashelby 2005). This species has been recorded from other regions, mainly in warm temperate waters and is stated by Ryland et al (2011) to be manifestly a warm water species. Given its current distribution and its preference for warmer water, we suggest that it does not represent a significant threat in terms of its chances of colonizing Norwegian coastal waters. However, with sea water temperatures rising, it could become established in the future and should, therefore, remain on the Norwegian Blacklist.
Bugula neritina (Linnaeus, 1758) (Figure 5) The original locality of Bugula neritina (Linnaeus, 1758) has been seemingly lost in the mists of time, though the first known reference to it in the literature (Ellis 1755) gives the localities as Mediterranean and America in the legend to P1 XIX. In the text of the book Ellis refers to a letter about the specimen from America. Unfortunately there is no indication on what the reference to the Mediterranean locality is based. Linnaeus (1758) gave the Mediterranean as a locality, and Pallas (1766) mentions Bermuda. Today, Bugula neritina has a global distribution in temperate, subtropical and tropical waters, which has probably been accomplished by ship fouling or with oyster imports. New molecular genetics studies by Fehlauer-Ale et al. (2013), however, have shown that what was previously thought to have been a complex of three cryptic species of Bugula neritina now relates to three distinct biological species; two of which are considered highly invasive.
In the NHM collections, the northernmost historical record of Bugula neritina in the British Isles appears to be from Scarborough collected by William Bean (1787-1866) in the collection of Dr George Johnston (NHMUK 1847.9.24.133). In Gray (1848) are five specimens listed for Bugula neritina labeled a-e. Four of those (b-e) were slender varieties from Tynemouth and other localities unknown. Specimen 'a' however refers to the material from Scarborough, W Bean Esq figured by Johnston in British Zoophytes plate 60 figures 3,4 ( Figure 5, G,H,I). Hincks (1880) did not mention specimen 'a' under the name Cellularia peachii, Busk. Johnston (1838) also quoted two other early British records given by Fleming (1828) and Templeton (1836) from the "coast of Cheshire". More recently, the species has been recorded from the east coast of the British Isles as far north as Lowestoft as part of Seasearch surveys (September 2014) and as far north as the Netherlands by Hans De Blauwe (Ryland et al 2011). This species is already listed in the Norwegian Blacklist (2012), and as sea temperatures rise, and the likelihood of it reaching Norway increases. We advise that it remains on the Blacklist. Previous European records of Watersipora subtorquata (d'Orbigny, 1852) have recently been shown to be the species Watersipora subatra (Ortmann, 1890) (Vieira et al. 2014). This highly invasive species is not currently known to be present in Norwegian waters; however, it has recently been documented as far north as Helgoland (Kuhlenkamp and Kind 2013). This species is likely to spread further north and, therefore, should remain on the Norwegian Blacklist. The taxon name should, however, be changed from Watersipora subtorquata to Watersipora subatra.

Horizon scanning for future shifting taxa
The traditional paradigm about 'cosmopolitan' species is currently under strong pressure, as it seems to have been proven wrong in many instances. Molecular studies have shown that many cosmopolitan species are actually complexes of cryptic species. On the other hand, some species recently acquired a widespread distribution pattern because of the increases in marine traffic and stepping stone structures being put into place on the seabed. This has allowed some species to invade into areas that would not have previously been possible. For taxonomists and people working in the field to monitor introductions this mixture of possibilities can cause real problems as was highlighted in Harmelin et al. 2012. In Norway, the Blacklist is used to highlight the Doorknocker species that have not yet arrived, but may be on the verge of doing so. Winston, Hayward and Craig, 2000 (Figure 6) Fenestrulina delicia was first described in 2000 (Winston et al. 2000) from the Damariscotta River, Maine (Holotype AMNH no.713). Colonies form dense white patches encrusting shells or algae. This species was recently recorded in north western Europe by De . They report the detecting Fenestrulina delicia on the French coast as far south as Pleneuf-Val-Andre (May, 2008). It was also reported as far north as Helgoland, NE Germany (August, 2008) where it was found on floating strands of the brown alga Himanthalia that were thought to have originated from the English Channel. Colonies of F. delicia were found on offshore natural and artificial substrates, and the species seems to be spreading along the coastlines rather than hopping from one marina to another. Part of the material collected was encrusting empty bivalve shells at locations known for mariculture of oysters and mussels; therefore, it was suggested that shellfish importation is a primary vector for introduction. Beached plastic waste with colonies of F. delicia on it was suggested to be a secondary vector of spread. The bases of offshore wind farm developments form an additional substrate for the species and are providing stepping-stones for the further spread of this exotic species. Fenestrulina delicia was reported as far north as Shetland (Wasson and De Blauwe 2014); however, it was not found as part of the harbour surveys during the present study. We suggest it is a likely 'Doorknocker' species because it may reach Norway by means of drifting seaweed and warrants addition to the Norwegian Blacklist.

Fenestrulina delicia
Pacificincola perforata (Okada and Mawatari, 1937) This species was described from Japanese waters, and has since been recorded in the Netherlands (De Blauwe and Faase 2004). It is possible that this species could spread northwards into Norwegian waters; therefore, we suggest it be added to the Norwegian Blacklist as a Doorknocker species. We included an SEM image of Japanese material from the NHM collection (NHMUK 2006.2.27.75-76; Figure 6) as an aid to future identification. Osburn, 1952 (Figure 6) De Blauwe and Faase (2004) and more recently Faasse et al. (2013) reported that the Pacific bryozoan Smittoidea prolifica Osburn, 1952 has been introduced to the Northeast Atlantic Ocean. Its currently known distribution in the Atlantic is restricted to The Netherlands. They report that all recent records of autochthonous Smittoidea reticulata (Macgillivray, 1842) in The Netherlands probably refer to S. prolifica. The most likely route of introduction is suggested to be via shellfish imports and the SEM images herein were provided to aid future identification. We recommend that this species be added to the Norwegian Blacklist as a Doorknocker species.

Recommendations
We recommend additional surveys be undertaken to the south of Bergen and to the North of Ålesund to obtain a wider baseline of the distributions of potentially invasive non-native bryozoan species in Norwegian waters. In the future, regular monitoring would provide information useful for understanding the spread of the invasive nonnative species along the Norwegian coastline and also to better understand the environmental conditions conducive to their growth and spread. The main vector of transport is highly likely to be leisure craft, as evidenced by the frequent observation of non-native and invasive species in recreational marinas. Some of the doorknocker species may enter Norwegian waters via the aquaculture route; consequently, a different type of monitoring would be required. We suggest that the Norwegian Blacklist 2012 be updated immediately by adding the following species: Watersipora subatra (instead of Watersipora subtorquata), Schizoporella japonica, Bugulina simplex, Fenestrulina delicia, Smittoidea prolifica, and Pacificincola perforata.