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Proposal v. 6: Marine biodiversity survey at St. Eustatius 7-28 June 2015 (Statia Marine Expedition 2015)

Bert W. Hoeksema Department of Marine Zoology, Naturalis Biodiversity Center, Leiden, The

Niels Schrieken MOO Project, ANEMOON Foundation, The Netherlands

Arjen Speksnijder Laboratories, Naturalis Biodiversity Center, Leiden, The Netherlands

General introduction

St. Eustatius, affectionately called “Statia”, is a volcanic in the northern Windward Group of the Lesser in the . Since 10 October 2010 this island is a Dutch municipality in the . It is part of the , which was previously known as the , constituting the ABC Islands (, and Curacao) in the off Venezuela and the SSS Islands (, St. Eustatius, the southern part of St. Maarten, with the submerged Saba Bank) in the western Atlantic (Figure 1).

Figure 1. Position of St. Eustatius in the Dutch Caribbean (ABC Islands and SSS Islands).

The marine biota of St. Eustatius is poorly investigated. The available information is not from recent studies. The Dutch naturalist Dr. Wagenaar Hummelinck (1953) published data on four collection stations where he sampled marine specimens on 10-15 July 1949 (Figure 2B: Stations 1116-1119) besides some fresh and brackish water habitats (Figure 2B: Stations 504- 515).

The phycologist Dr. Vroman (1968: pp. 59-61) published maps (Figure 3) in which he indicated six shore localities where he collected algae (20-21 May 1958). Roos (1971) reported on corals collected at six localities in the period 24-27 July 1965(Figure 4).

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Naturalis Biodiversity Center (previously known as Rijksmuseum van Natuurlijke Historie) was involved in the organization of an expedition to the neighbouring Saba Bank (Van der land 1977), which included two sampling stations (Sta. 29, Sta. 121) at the west coast of St. Eustatius and two at Saba Island (Figure 5). These localities were visited May-June 1972. Samples have been deposited in the Naturalis research collections, where they are available for further research. This material can be used for studies concerning possible changes in the marine fauna and flora (Hoeksema et al. 2011).

Figure 2. Maps published by Wagenaar Hummelinck (1953). A. Windward Group of Antilles where material was collected (name of St. Eustatius is underlined). B. Four collection stations at St. Eustatius: 1116. Southern Part of Gallows Bay, 1117. Downtown near Billy Gut, 1118. Billy Gut near Downtown, 1119. South of Tumble Down Dick Bay.

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Figure 3. Maps published by Vroman (1968). A. Windward Group of Antilles where material was collected (name of St. Eustatius is underlined). B. Six collection stations at St. Eustatius (marked by red circles): 1. Boekaniers Bay, 2. Bay between Boekaniers Bay and Corre Corre Bay, 3. Corre Corre Bay, 4. Concordia Bay, 5. Back-off Bay (Sugarloaf and White Wall), 6. Back-off Bay (sublittoral). Descriptions of the localities are given by Vroman (1968: pp. 59-61)

Figure 4. Collecting stations of Roos (1971) around Saba (S), St Maarten (M) and St Eustatius (E) in 1965. St Eustatius: 1. Cocoluch Bay – Jenkin’s Bay 2. Jenkin’s bay – Tumbledown Dick bay 3. Oranjestad Bay 4. Gallows Bay 5. Compagnie Baai 6. Schildpadden Baai

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Figure 5. Position of St-Eustatius (arrow) in relation to Saba Bank and Saba with sampling stations visited during the Saba Bank Expedition in 1972 (after Van der Land 1977)

Statia Marine 2015

Naturalis Biodiversity Center (‘Naturalis Zeeteam’) plans to organize a marine expedition to St. Eustatius in June 2015 in collaboration with ‘Anemoon Foundation’. The Caribbean Netherlands Science Institute (CNSI) will act as host by offering laboratory space and lodging at Statia. The St. Eustatius National Parks Foundation (STENAPA) will act as local counterpart. This implies that results should be made publicly available for local conservation efforts through scientific reports and publications, websites, and exhibits. Scubaqua will be the local dive center assisting in diving logistics.

The primary goal of this field survey will be to set up a base line, which can be used for future studies on biotic change (marine fauna and flora) after hurricanes or periods of coral bleaching that may cause large-scale coral mortality. The logistic set-up of this expedition will be similar to previous ones in which Naturalis was involved, mostly in Southeast Asia (Hoeksema & Tuti 2001; Hoeksema 2004, Hoeksema & van der Meij 2008, 2010). However, the scientific goals of this expedition may differ because of the baseline aspect.

Goals: 1. A study of the composition and distribution patterns of taxa covered by taxonomic expertise of the research team will show a. marine species that are widely spread or locally distributed b. the variation in species diversity (species richness) among the various localities

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c. the marine benthic diversity of Statia in comparison with other areas in the Caribbean d. whether a baseline can be established for studies concerning changes over time e. whether species can be discovered that are new to science f. the distribution of keystone taxa important for conservation (endangered species, invasive species) 2. Photographic documentation and a reference collection will support the establishment of the baseline. 3. DNA samples will be taken for molecular analyses and DNA bar coding.

Possible expertise: stony corals, soft corals, sponges, associated fauna (shrimps, snails, crabs, etc.), interstitial fauna, macro algae, fish, and metagenomics.

Participants (20) from Naturalis Biodiversity Center (research staff, collection staff), universities (students), Anemoon Foundation (volunteers for long-term monitoring), taxonomic specialists of other institutes.

Sampling strategy: The roving diving technique (presence / absence records per dive with ca. 60 min observation time, including photography and collecting of voucher specimens and dna samples). Max. depth 30 m. Species records will be used in a species richness estimation analysis for species presence-absence comparisons.

Expedition stations: Approximately 35 sites around St.-Eustatius (2 / day; 6 days / week) will be selected during the survey, representing various coral reef environments for covering maximum habitat diversity. A selection will be made from standard dive sites and newly established sites following priorities given by STENAPA (Figure 6). The localities vary with regard to bathymetry (Figure 7), substrate volcanic / limestone (Figures 2B, 3B), wind exposure (Figures 8, 9), coastline morphology (Figure 10), island landscape (Figure 11), and rules and regulations concerning the marine parks (Figure 6).

Time frame: The month June has little wind and rainfall (Figures 8, 9).

Participants (all SCUBA divers except when mentioned otherwise):

Naturalis Biodiversity Center 1. Dr. Bert W. Hoeksema (stony corals and associated fauna) 2. Dr. Ronald Vonk (interstitial bottom fauna) 3. Dr. Arjen Speksnijder (molecular biodiversity bottom fauna) 4. Frank Stokvis, MSc (technician molecular analyses) 5. Mr. Koos van Egmond (collection technician) 6. Ms. Yee Wah Lau, MSc (student UL, octocorals) 7. Ms. Luna van der Loos (student RUG, seaweed) – also member of Anemoon team 8. Dr. Willem Prud'homme van Reine (seaweed) non-SCUBA

Foreign specialists 1. Prof. Jim Thomas, Nova Southeastern University, Fort Lauderdale, USA (amphipods) 2. Dr. Slava Ivanenko, Moscow State University, Russia (coral-associated copepods) 3. Dr. Simone Montano, University of Milano-Bicocca, Milano, Italy (hydrozoans) 4. Dr. James Reimer, University of the Ryukyus, Okinawa, Japan (zoantharians) 5. Mr. Jaaziel E. Garcia-Hernandez, University of (sponges).

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Anemoon Foundation 1. Mr. Niels Schrieken, MSc (coordinator Anemoon; ascideans, sponges) 2. Dr. Godfried van Moorsel (scleractinians, fishes) 3. Mr. Marco Faasse, MSc (hydrozoans, bryozoans) 4. Ms. Marion Haarsma (molluscs, general marine fauna) 5. Ms. Sylvia van Leeuwen (littoral fauna and flora) - 2 weeks non-SCUBA 6. Mr. Peter van Rodijnen (cineast) - 1 week 7. Ms. Susan Hewitt (littoral fauna and flora) - 3 weeks non-SCUBA

STENAPA p.m.

Tasks Participants contribute to the expedition results without financial compensation - a marine species list for Statia (base line) - biodiversity analysis of dive sites around Statia (based on exemplar taxa and species that are protected, rare, invasive (or of other interest) - reference collections (Naturalis and CNSI) - specimens for barcoding - scientific publications - photographic materials, films and documentation for outreach (websites,

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Figure 6. Position of dive sites at St Eustatius. Source: http://www.statiapark.org/parks/marine/img/statia_dive_map.pdf

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Figure 7. Bathymetry around St. Eustatius.

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Figure 8. Wind statistics of St-Eustatius: http://www.windfinder.com/windstatistics/st_eustatius

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Figure 9. Climate of St.-Eustatius.: http://www.whatstheweatherlike.org/netherlands/sint-eustatius.htm

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Figure 10. Coastline of St. Eustatius showing wave exposure at the eastern shore (Google Earth)

References: Hoeksema BW (2004) Marine biodiversity of the coastal area of the Berau region, East Kalimantan, Indonesia. Progress report East Kalimantan Program - Pilot phase (October 2003), Naturalis, Leiden. pp 1-82. Hoeksema BW, Tuti Y (2001) Marine biodiversity of Lombok Strait, Bali: Preliminary research report. NNM Naturalis, Leiden, pp 1-16. Hoeksema BW, van der Meij SET (2008). Cryptic marine biota of the Raja Ampat Islands group, pp. 1-74. Naturalis, Leiden. Hoeksema BW, van der Meij SET (2010) Crossing marine lines at Ternate: Capacity building of junior scientist in Indonesia for marine biodiversity assessments. Naturalis, Leiden. pp. 1-85. Hoeksema BW, van der Land J, van der Meij SET, van Ofwegen LP, Reijnen BT, van Soest RWM, de Voogd NJ (2011) Unforeseen importance of historical collections as baselines to determine biotic change of coral reefs: the Saba Bank case. Marine Ecology 32: 135-141. van der Land J (1977) The Saba Bank – a large atoll in the northeastern Caribbean. FAO Fisheries Report, 200, 469–481. Roos PM (1971) The shallow-water stony corals of the Netherlands Antilles. Studies on the Fauna of Curaçao and other Caribbean Islands 37: 1-108, pls. 1-53. Vroman M (1968) The marine algal vegetation of St. Martin, St. Eustatius and Saba (Netherlands Antilles). Studies on the Flora of Curaçao and other Caribbean Islands 2: 1-120, pls. 1-10.

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Wagenaar Hummelinck P (1953) Description of new localities. Studies on the Fauna of Curaçao and other Caribbean Islands 4: 1-108, pls. 1-8.

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Figure 11. Landscape ecological map of St.-Eustatius: http://www.dcbd.nl/document/landscape-ecological- vegetation-map-sint-eustatius-lesser-antilles

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Main research questions

Species richness around Statia is variable, depending on various environmental elements. It is hypothesized that the following factors play a role: bathymetry (Figure 6), substrate volcanic rock / limestone rock / sand (Figures 2B, 3B), wind exposure (Figures 7, 8), coastline morphology (Figure 9), island landscape and land use (Figure 10), and rules and regulations concerning the marine parks (Figure 5).

Species presence / absence at the dive sites will be used in the survey to find which species are common and which ones are rare. For the monitoring of some species groups the expertise of taxonomic specialists is required and for other species the experience of volunteer naturalist is demanded. The species richness data will be analysed with the help of statistics, for which the help of a statistician will be required (the chairman of ANEMOON is employed by the Central Bureau for Statistics). Species of selected taxa will be used for barcoding. The following species groups will be used. - dominant benthic species groups: stony corals, octocorals, sponges, macro algae. - associated fauna: species living in symbiosis with benthic organsims. - iconic species: species represented by specimens that are relatively easy to identify by non- taxonomists and key / indicator species that are expected to be important for reef management because of their rarity or economic value (queen conch, coral diseases). For more details on the involvement of volunteer biologists (citizen’s science), see the next two pages. - species richness patterns around Statia will also be studied by molecular analyses (see chapter on Barcoding Environment of Statius).

The importance of symbiosis for marine biodiversity is studied by the inclusion of associated fauna. The host dependence may be generalist or specialist (host-specific). Specimens of these species are difficult to find (due to their small size) and recognize by non-taxonomists. Several of these species are expected to be new to science. Examples are coral-associated copepods and symbiotic hydroids on scleractinians and octocorals. Some coral-associated species are well known but their host preference is not well studied, such as Christmas tree worms (Spirobranchus spp.). The results

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Collaboration between Naturalis Biodiversity Center and ANEMOON foundation (ANalyse Educatie en Marien Oecologisch ONderzoek): The marine biodiversity of St Eustatius

Collaboration by Naturalis and the ANEMOON foundation can significantly improve long-term monitoring by volunteer divers around St Eustatius. These improvements include [1] raising public awareness about the marine life, [2] increasing the number of volunteers that monitor St Eustatius, [3] setting up a relatively low cost continuous monitoring program at St Eustatius, and [4] optimizing the validation process of the volunteer monitoring data to enable its use in both peer-reviewed scientific publications and in advisory reports for public and governmental agencies involved in marine nature conservation.

The Monitoring Onderwater Oevers (MOO) project

These above specified goals can be met by promoting the expansion of the MOO project of the ANEMOON Foundation around St Eustatius. The MOO project started in 1994 and is executed in the coastal waters of the Netherlands with a special focus on the Dutch delta region. Serious volunteers are invited to join the project. In the first 3 to 4 years at least 10 workshops were given annually at dive clubs to get the project started and to train the volunteers in how to monitor marine life. In addition, an internet forum was started where anyone can post questions. Both professionals and volunteers can respond to these questions. Nowadays the forum is actively used by volunteers and professional biologists. Unusual records and sightings posted on the forum are almost immediately checked and validated based on reference material (usually photographic evidence). An official collecting permit has been issued to a selection of volunteers for specimens in the Dutch Delta when this is needed for validation of identification. The collected specimen(s) are usually deposited in the collections of Naturalis Biodiversity Center. As a result most species recorded as new to the Netherlands are first reported by volunteers of the ANEMOON foundation.

Over the years a few thousand divers have filled out a MOO form. Around 200 divers do this on a frequent basis. A core group of at least 20 divers with changing composition consistently monitors the underwater life and will probably do so for years to come. They have become specialists of marine species themselves and are actively involved in the internet forum. The uploaded data in the database is validated by experts as far as possible. Because many scuba-divers have underwater cameras, validation of species records has become much easier in recent years. The data has been used in a large number of publications in both peer reviewed scientific journals and reports for the government.

More than 8000 forms (2 volunteers per form) have been filled out over the years, which makes the project one of the most successful marine monitoring projects worldwide that involves scuba-divers. A weak point of the MOO observations is that they does not evenly include all waters in The Netherlands. The coastal waters and especially those of the Dutch Delta are well represented while the number of forms concerning the North Sea is still very limited, i.e. less than a hundred since 1994, regardless of the fact that many dives are made on the North Sea. Most of these divers do not or rarely dive in the Dutch Delta and therefore have no knowledge of or experience with the MOO project.

During two Dogger Bank expeditions, one to the southern North Sea and one to Brown Ridge, baseline studies were performed between 2011 and 2013 to monitor the hard substrata on the Dutch North Sea using the MOO aproach. This resulted in the discovery of over 20 new species for the Dutch North Sea of which several were new to the North Sea as a whole. Part of these results have been published in six peer reviewed articles. They indicate that a lot is still unknown about the diversity of marine species on the Dutch North Sea, and thereby the importance of stimulating monitoring effort in this region.

As the MOO project with volunteer scuba divers has already shown to be very success in coastal waters, this should also be possible in St Eustatius. As both the diversity of species and a large part of

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the group of active divers at St Eustatius will vary, setting up a successful MOO project in this region needs special effort. This includes adjusting MOO forms, giving presentations at St Eustatius, giving directions to volunteers on how to make photographs of specimens in a way that enables the validation of species identifications, etc.

To get this done, ANEMOON foundation and Naturalis Biodiversity Center need to collaborate in any way as much as this is possible and feasible for both parties.

References

Gittenberger E, Gittenberger A, Dekker H, Kronenberg GC (eds.) (2004) New data on Dutch marine . Vita Malacologica 2: 1-72 Gittenberger A (2007) Recent population expansions of non-native ascidians in The Netherlands. Journal of Experimental Marine Biology and Ecology 342: 122-126. Gittenberger A, Schipper C (2008) Long live Linnaeus, Lineus longissimus (Gunnerus, 1770) (Vermes: Nemertea: Anopla: Heteronemertea: Lineidae), the longest worldwide and its relatives occurring in The Netherlands. Zoologische Mededelingen 82: 59-63. Gittenberger A, et al. (2008) Risicoanalyse van de Amerikaanse langlob-ribkwal Mnemiopsis leidyi A. Agassiz, 1865. GiMaRIS. Schonenberg DB, Gittenberger A (2008). The invasive quagga mussel Dreissena rostriformis bugensis (Andrusov, 1879)(Bivalvia: Dreissenidae) in the Dutch Haringvliet, an enclosed freshwater Rhine-Meuse estuary, the westernmost record for Europe. Basteria 72: 345-352. Gittenberger A, et al. (2010) Risk analysis of the colonial sea-squirt Didemnum vexillum Kott, 2002 in the Dutch Wadden Sea, a UNESCO World Heritage Site. GiMaRIS. Gittenberger A, Rensing M, Stegenga H, Hoeksema BW (2010) Native and non-native species of hard substrata in the Dutch Wadden Sea. Nederlandse Faunistische Mededelingen 33: 21-75. Gittenberger A, van Stelt RC (2011) Artificial structures in harbors and their associated ascidian fauna. Aquatic Invasions 6: 413-420. Gittenberger A, Moons JJS (2011) Settlement and possible competition for space between the invasive violet tunicate Botrylloides violaceus and the native star tunicate Botryllus schlosseri in The Netherlands. Aquatic Invasions 6: 435-440. Gittenberger A., et al. (2011) Risk analysis of hull fouling on small to medium sized boats as an import vector of exotic species in the Wadden Sea. GiMaRIS. Gittenberger A, Schrieken N, Lengkeek W (2011) faeroensis Lemche, 1929, and Doto dunnei Lemche, 1976, new for the Dutch fauna and central North Sea (, Nudibranchia). Basteria 75: 111-116. Lindeyer F, Gittenberger A (2011). Ascidians in the succession of marine fouling communities. Aquatic Invasions 6: 421-434. Schrieken N, Gittenberger A, Lengkeek W (2011) First record of Xandarovula patula (Pennant, 1777) in the Dutch North Sea (Gastropoda, Ovulidae). Basteria 75: 107-110. Gittenberger A, Kleukers E. (eds) (2013) Oases of marine biodiversity in the Dutch North Sea. Nederlandse Faunistische Mededelingen 41: 1-114. Gittenberger A, Schrieken N, Coolen JWP, Gittenberger E (2013) Shipwrecks, ascidians and Modiolarca subpicta (Bivalvia, Mytilidae, Musculinae). Basteria 77: 75-82. Gittenberger A, Schrieken N, Coolen J, Vlierhuis W (2013) The Jewel anemone Corynactis viridis, a new order for The Netherlands (Cnidaria: Corallimorpharia). Nederlandse Faunistische Mededelingen 41: 35-41. Lengkeek W, Coolen J, Gittenberger A, Schrieken N (2013) Ecological relevance of shipwrecks in the North Sea. Nederlandse Faunistische Mededelingen 41: 49-57. Schrieken N, Gittenberger A, Coolen J, Lengkeek W (2013) Marine fauna of hard substrata of the Cleaver Bank and Dogger Bank. Nederlandse Faunistische Mededelingen 41: 69-78.

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Barcoding Environment of Statius

In 2010 Naturalis started the Dutch Barcoding project. The aim of this project is to capture the Dutch biodiversity and to create a collection of fully described species backed up by DNA analysis and high resolution photography. The DNA analysis consist of barcoding and phylogenetic analysis of representative genes to support taxonomic classification. Fresh samples are collected to acquire high quality DNA which is also stored for future extended analysis.

Up to now a DNA collection of 40.000 species has been established and 27.000 species have been barcoded. The aim is to contribute to the world wide a database for biodiversity (BOLD). This Barcoding of Life Database consists currently around 3 million barcode sequences describing 144,080 , 52,571 Plants and 15,369 Fungi & Other Life. The BOLD database is functioning as a reference database for (newly) described species and connects this to DNA barcode analysis. This approach is suitable for (environmental) DNA approaches the capture and monitor biodiversity on different trophic levels.

Setting up a barcode database

To enrich our knowledge about Dutch biodiversity in the overseas territories we propose a barcoding project embedded in the baseline biodiversity measurements on the Island of . A DNA and a tissue collection can be set up for future DNA analysis. Barcode sequences are determined from selected species to make a DNA based reference database of the biodiversity of the island (S- BOLD). This reference library can be used for future monitoring. Future monitoring consists of collecting samples and mixed samples for DNA extraction and next generation sequence analysis We want to describe above ground en below water biodiversity based on standardized eDNA analysis. This will be the groundwork of a monitoring tool which can be used in environmental impact studies.

Coordinated sampling using existing barcoding project procedures

A protocol for a reference database is proposed to collect a representation of terrestrial invertebrates like spiders, beetles, flies, bugs, wasps, bees, ants, damselflies, dragonflies, moths, butterflies and other terrestrial invertebrates. We also want to collect vascular plants, mosses and fungi. Below water we want to collect marine invertebrates. Protocols for collecting are already well developed for the Netherlands but should be adapted the tropical environment and we need commitment of the involved researchers for this collection and taxonomic classification. From the collected samples, DNA will be extracted for long term preservation and, if possible, also tissue will be stored. The aim is to acquire as many DNA samples as possible for future research whilst the more costly process of barcoding can be performed on fewer samples answering the current questions. eDNA sampling for metabarcode analysis

Little is known about the interaction of whole community compositions between habitats above and below the surface and to which extend biodiversity is coupled. Modern techniques like metabarcoding of CO1, ITS and matK of environmental DNA by next generation sequencing enables us to analyze total biodiversity in a given dimension and time point. Meta-analysis of eDNA data enables us to relate diversity, interactions and community changes through different dimensions and time. At Statia we will assess above and below surface biodiversity and there is a possibility to sample through time to asses full impact of human practices above ground and the influence of this below the surface. These above and below surface biodiversity assessments can be set up for human impact studies on

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fragile islands and coastal waters in general. The main scientific question lies in interactions between communities.

For monitoring biodiversity changes, an environmental impact a study on terrestrial invertebrates is proposed with Malaise traps in urban, transitional and nature conservation areas during a period of time in which status of the area is changed (e.g. from transitional to conserved or from transitional to urban). A similar sample strategy can be used for marine sediments by sampling cores and in the water column by filtration. Standard dive stations will be used for repetitive sampling in a long-term monitoring programme.

Sampling for microbial interaction

Species can carry a core microbiome which is shared amongst them. This enables us to use the microbial composition as a support for taxonomical classification. The application of microbial communities in the field of marine zoology can resolve of cryptic (coral) species. Therefore targeted sampling of cryptic marine species is proposed. Microbial analysis will be performed by r- RNA barcoding.

Establishing a logistic pipeline for DNA analysis

To maximize the possibilities of DNA analyses in the future we should apply optimized and standardized procedures for tissue preservation and DNA isolation. Therefore it is worthwhile to look on sight for possibilities to conserve samples locally but also to secure DNA by correct preservation and if possible DNA isolation. Local DNA isolation can circumvent phytosanitary risks especially for soil and sediment samples and therefore provides logistically easy transport of samples without need of a licence. A simple laboratory set up with a fridge and centrifuge can be sufficient to secure DNA quality.

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Web sites of main partners ------https://science.naturalis.nl/en/research/systematic-research/marine-zoology/ https://www.facebook.com/zeeteam.naturalis

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http://www.anemoon.org/ https://www.facebook.com/groups/StAnemoonMOO/ ------

http://www.cnsi.nl/ https://www.facebook.com/pages/Caribbean-Netherlands-Science-Institute-at-St-Eustatius- CNSI/381662415272901 ------

STENAPA http://www.statiapark.org/ https://www.facebook.com/pages/Stenapa-St-Eustatius/203334299688441

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http://www.scubaqua.com/

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https://www.facebook.com/pages/Scubaqua-Dive-Center-STATIA/167974488162