An integrative identification guide to the Hydrozoa (Cnidaria) of Bocas del Toro, Panama

ABSTRACT

This work is the first attempt to assess the biodiversity of the Hydrozoa in the Archipiélago de Bocas del Toro (Panamá, Caribbean Sea) using morphology and molecular taxonomy, and to produce field identification tools to help future identification and monitoring efforts in the area.

We sampled, identified, vouchered, and barcoded 112 specimens of Hydrozoa from shallow coastal waters (0–22 m depth) in the Archipiélago de Bocas del Toro. The specimens belong to 70 taxa, of which 53 were identified at the species level, and 17 were identified at the genus or family level. We produced 64 sequences of the large ribosomal subunit of the mitochondrial RNA (mt lsu-rRNA, 16S), the genetic marker generally used for barcoding Hydrozoa. We updated the local checklist that now comprises 118 species, and produced 87 detailed taxon identification tables that display species descriptions augmented with pictures, geographic distribution (worldwide and in Bocas del Toro), GenBank accession numbers for the 16S mitochondrial gene, and a synopsis of the families they belong to.

KEYWORDS:

• Hydrozoa
• Bocas del Toro
• identification tools
• barcoding
• Caribbean
• 16S
• biodiversity

Introduction

Hydrozoa are an inconspicuous and often overlooked class of the phylum Cnidaria [1]. Most hydrozoans have a complex life cycle characterized by a progression of three life stages: a short-living larva (the planula), generally metamorphosing into a benthic colonial stage (the polyp), and a pelagic sexual stage (the medusa stage) asexually budded off from the polyp [2]. The life cycle can be shortened into a bi-phasic cycle, by reduction or complete suppression of either the polyp or the medusa stage [3]. With more than 3,700 described species [4] hydrozoans are structurally and functionally important members of benthic and planktonic communities [5; 1]. Work on Hydrozoa has been hampered by the scarcity of taxonomic expertise, which has dramatically declined over the past two decades [6,7]. Also, because polyps and medusae require different expertise and each follow their own identification rules, linking both to a single species has proven difficult and has further hampered cohesive taxonomic revisions. In the last 10 years molecular tools have contributed significantly to hydrozoan taxonomy and have shown that selected gene sequences may be necessary, in combination with traditional taxonomy, to correctly identify cryptic species and disentangle taxonomic confusion [8–12].

Hydrozoa from the Atlantic/Caribbean coast of Panama are abundant but scarcely studied. The Archipiélago de Bocas del Toro, Panamá is located on the NW Caribbean shore of Panama, close to the Costa Rican border. It consists of more than 68 small islands and mangrove keys and is characterized by diverse ecosystems, from mangrove dominated shallow water to coral reefs and sea grass meadows (see Figure 1 for a map). To date, 79 nominal species have been reported in the Bocas del Toro region [13]. However, proper descriptions and species identification tools are lacking, inadequate, or scattered in old and hard to access articles, thus hindering the ability of nonexperts to identify species of interest in this area.

Figure 1. Map of the Bocas del Toro archipelago, with sampling localities.

This work is a result of the Taxonomic Training workshop held in July 2015 in Bocas del Toro, Panama, organized by Smithsonian Tropical Research Institute as part of a National Science Foundation Advancing Revisionary Taxonomy and Systematics (ARTS) grant. During the workshop, in which 14 students participated, we collected shallow water Hydrozoa around the Archipiélago de Bocas del Toro, Panamá. Collections targeted both pelagic medusa with plankton tows and benthic polyps with snorkeling. In this paper, we present an updated checklist of hydrozoan biodiversity in the Archipiélago de Bocas del Toro,, augmented by [I] a DNA barcoding database consisting of sequences of a ~ 600bp gene fragment of the mitochondrial large ribosomal RNA subunit (mt lsu-rRNA, 16S), and [II] taxon identification tables. The 16S gene has been successfully used for taxonomic revisions and is considered the Hydrozoa “barcoding” molecule [10,11,14–17]. Both the barcoding data and the taxonomic identification sheets were produced to aid future biodiversity inventory efforts in the region.

Material and methods

Collection

Hydrozoa were collected during the Tropical Taxonomy Training course on systematics and biology of Hydrozoa (Cnidaria) held at the Smithsonian Tropical Research in Bocas del Toro, Panama from 7/7/2015 to 7/21/2015. A total of 16 people (two instructors and 14 students) were in the field during every sampling effort. A total of 11 sites were sampled. Localities sampled were: Bocas del Toro Marine Station docks/weather station, Punta Hospital, Crawl Cay, Bocatorito Bay vicinity, San Cristobal, Vicinity of Manuguar Cay, Punta Caracol, Swans Cay, Bastimento vicinity of “Casa Verde”, Bocas del Drago, Pandora, Almirante (Quary’s point) (Table 1 and Figure 1). Polyps were collected by snorkeling (0–8 m depth) or SCUBA diving (18–22 m depth), and medusae by plankton towing using a net with 280µm mesh size. Plankton tows were carried out near the Bocas del Toro Station docks and weather station.

Table 1. Samples collected during the 2015 Hydrozoa workshop held in Bocas del Toro, Panama. Date of collections, sample ID, species identified, type of fixative used for preservation (ethanol or formalin), GenBank accession number, and location within the Bocas del Toro archipelago are reported. In the table N = No, Y = Yes.

	Date	Sample ID	Species	Fertile	Ethanol	Formalin	Sequenced/ GenBank accession #	Location
1	7/7/2015	BTH.15.1	Kirchenpaueria halecioides	Y	Y	N	N	STRI BDT docks/weather station
2	7/7/2015	BTH.15.2	Halecium cf. bermudense	Y	Y	N	N	STRI BDT docks/weather station
3	7/7/2015	BTH.15.3	Nemalecium lighti	N	Y	Y	N	STRI BDT docks/weather station
4	7/7/2015	BTH.15.4	Nemalecium lighti	N	Y	Y	MH361321	STRI BDT docks/weather station
5	7/7/2015	BTH.15.5	Kirchenpaueria halecioides	N	Y	Y	N	STRI BDT docks/weather station
6	7/7/2015	BTH.15.6	Clytia linearis	Y	Y	N	MH374630	STRI BDT docks/weather station
7	7/7/2015	BTH.15.7	Dynamena crisioides	Y	Y	N	MH361322	STRI BDT docks/weather station
8	7/7/2015	BTH.15.8	?Cytaeis	N	Y	Y	N	STRI BDT docks/weather station
9	7/7/2015	BTH.15.9	Dynamena crisioides	Y	Y	N	MH361323	STRI BDT docks/weather station
10	7/7/2015	BTH.15.10	Halecium bermudense	Y	Y	N	N	STRI BDT docks/weather station
11	7/8/2015	BTH.15.11	Myrionema amboinense	N	Y	Y	N	Punta hospital
12	7/8/2015	BTH.15.12	Plumularia sp.	N	Y	Y	N	Punta hospital
13	7/8/2015	BTH.15.13	Antennella secundaria	Y	Y	N	MH361324	Punta hospital
14	7/8/2015	BTH.15.14	Halopteris alternata	N	Y	N	MH361325	Punta hospital
15	7/8/2015	BTH.15.15	Eudendrium carneum	N	Y	Y	MH361326	Punta hospital
16	7/8/2015	BTH.15.16	Salacia desmoides	Y	Y	Y	MH361327	Punta hospital
17	7/8/2015	BTH.15.17	Eudendrium bermudense	Y	Y	Y	N	Punta hospital
18	7/8/2015	BTH.15.18	Antennella secundaria	Y	Y	Y	MH361328	Punta hospital
19	7/8/2015	BTH.15.19	Obelia dichotoma	Y	Y	Y	N	Punta hospital
20	7/8/2015	BTH.15.20	Aglaophenia latecarinata	N	Y	Y	N	BDT docks/weather station
21	7/8/2015	BTH.15.21	Kirchenpaueria halecioides	Y	N	Y	N	Punta hospital
22	7/8/2015	BTH.15.22	Plumularia sp.	N	Y	Y	N	Punta hospital
23	7/9/2015	BTH.15.23	Pennaria disticha	Y	Y	Y	MH361329	Crawl Cay
24	7/9/2015	BTH.15.24	Ralpharia gorganiae	Y	Y	N	MH361330	Crawl Cay
25	7/9/2015	BTH.15.25	Stylaster roseus	N	Y	Y	MH361331	Crawl Cay
26	7/9/2015	BTH.15.26	Thyroscyphus marginatus	Y	Y	Y	MH361332	Crawl Cay
27	7/9/2015	BTH.15.27	Gastroblasta raffaelei	N	Y	Y	N	Crawl Cay
28	7/9/2015	BTH.15.28	Pennaria disticha	Y	Y	N	N	Crawl Cay
29	7/9/2015	BTH.15.29	Thyroscyphus marginatus	N	Y	N	MH361334	Crawl Cay
30	7/9/2015	BTH.15.30	?Obelia dichotoma	N	Y	Y	N	Crawl Cay
31	7/9/2015	BTH.15.31	Millepora alcicornis	N	Y	N	MH361335	Crawl Cay
32	7/9/2015	BTH.15.32	Dynamena disticha	N	Y	Y	MH361336	Crawl Cay
33	7/9/2015	BTH.15.33	Dynamena crisioides	N	Y	N	N	Crawl Cay
34	7/9/2015	BTH.15.34	Obelia dichotoma	Y	N	Y	N	Crawl Cay
35	7/9/2015	BTH.15.35	Eudendrium capillare	Y	Y	Y	MH361337	Crawl Cay
36	7/9/2015	BTH.15.36	Clytia hemisphaerica	Y	Y	N	N	Crawl Cay
37	7/9/2015	BTH.15.37	Clytia hemisphaerica	N	N	Y	N	Crawl Cay
38	7/9/2015	BTH.15.38	Halecium sp.2	Y	Y	Y	MH361338	Crawl Cay
39	7/9/2015	BTH.15.39	Hincksella formosa	N	Y	Y	MH361339	Crawl Cay
40	7/10/2015	BTH.15.40	Sphaerocoryne cf. agassizii	N	Y	Y	MH361340	Near Bocatorito Bay
41	7/10/2015	BTH.15.41	Clytia hemisphaerica	N	Y	Y	N	Near Bocatorito Bay
42	7/10/2015	BTH.15.42	Monotheca margaretta	Y	Y	Y	N	Near Bocatorito Bay
43	7/10/2015	BTH.15.43	Sertularia distans	N	Y	Y	MH361341	Near Bocatorito Bay
44	7/10/2015	BTH.15.44	Gastroblasta raffaelei	N	Y	Y	MH361342	Near Bocatorito Bay
45	7/10/2015	BTH.15.45	Halecium cf. nanum	N	Y	Y	MH361343	Near Bocatorito Bay
46	7/10/2015	BTH.15.46	Gastroblasta raffaelei	N	Y	Y	N	Near Bocatorito Bay
47	7/10/2015	BTH.15.47	Kirchenpaueria halecioides	N	Y	N	MH361344	Near Bocatorito Bay
48	7/10/2015	BTH.15.48	Gastroblasta raffaelei	N	Y	N	N	Near Bocatorito Bay
49	7/10/2015	BTH.15.49	Gastroblasta raffaelei	N	Y	N	N	Near Bocatorito Bay
50	7/10/2015	BTH.15.50	Halopteris alternata	N	Y	Y	MH361345	Near Bocatorito Bay
51	7/10/2015	BTH.15.51	Monotheca margaretta	Y	Y	Y	MH361346	Near Bocatorito Bay
52	7/10/2015	BTH.15.52	Halecium sp. [sensu 17]	N	Y	N	MH361347	Near Bocatorito Bay
53	7/10/2015	BTH.15.53	Halopteris alternata	N	Y	N	MH361348	Near Bocatorito Bay
54	7/10/2015	BTH.15.54	Kirchenpaueria halecioides	Y	Y	Y	MH361349	Near Bocatorito Bay
55	7/11/2015	BTH.15.55	Gastroblasta raffaelei	N	Y	Y	N	San Cristobal
56	7/11/2015	BTH.15.56	Oceaniidae indet	N	Y	Y	N	San Cristobal
57	7/11/2015	BTH.15.57	Halecium sp.1	N	Y	N	MH361350	San Cristobal
58	7/11/2015	BTH.15.58	Nemalecium lighti	N	Y	N	MH361351	San Cristobal
59	7/11/2015	BTH.15.59	Oceaniidae indet	N	Y	N	N	San Cristobal
60	7/11/2015	BTH.15.60	Obelia bidentata	N	Y	N	MH361352	San Cristobal
61	7/11/2015	BTH.15.61	Amphinema dinema	Medusa	N	N	N	San Cristobal
62	7/11/2015	BTH.15.62	Clytia gracilis	Y	Y	Y	N	San Cristobal
63	7/11/2015	BTH.15.63	Liriope tetraphylla	Medusa	Y	Y	MH361353	San Cristobal
64	7/11/2015	BTH.15.64	Thecocodium sp.	N	Y	N	MH361354	San Cristobal
65	7/12/2015	BTH.15.65	Plumularia floridana	Y	Y	Y	MH361355	Vicinity of Manuguar Cay
66	7/12/2015	BTH.15.66	Cirrholovenia tetranema	N	N	Y	N	Vicinity of Manuguar Cay
67	7/12/2015	BTH.15.67	Clytia hummelincki	Y	Y	Y	N	Vicinity of Manuguar Cay
68	7/12/2015	BTH.15.68	Clytia linearis	N	Y	N	MH361356	Vicinity of Manuguar Cay
69	7/12/2015	BTH.15.69	Sphaerocoryne cf. agassizii	N	Y	N	N	Vicinity of Manuguar Cay
70	7/12/2015	BTH.15.70	Halecium tenellum	Y	Y	Y	N	Vicinity of Manuguar Cay
71	7/12/2015	BTH.15.71	Egmundella sp. (on Clytia linearis)	N	N	Y	N	Vicinity of Manuguar Cay
72	7/12/2015	BTH.15.72	Clytia hummelincki	Y	Y	N	MH361357	Vicinity of Manuguar Cay
73	7/12/2015	BTH.15.73	Dynamena crisioides	Y	Y	Y	MH361358	Vicinity of Manuguar Cay
74	7/12/2015	BTH.15.74	Dynamena crisioides	N	Y	N	MH361359	Vicinity of Manuguar Cay
75	7/12/2015	BTH.15.75	Obelia bidentata	N	Y	Y	N	Vicinity of Manuguar Cay
76	7/12/2015	BTH.15.76	Clytia linearis	N	Y	Y	MH361361	Vicinity of Manuguar Cay
77	7/12/2015	BTH.15.77	Nemalecium lighti	Y	Y	N	MH361362	Vicinity of Manuguar Cay
78	7/13/2015	BTH.15.78	Pteroclava krempfi	N	Y	Y	MH361363	Punta Caracol
79	7/13/2015	BTH.15.79	Sphaerocoryne cf. agassizi	N	Y	N	MH361364	Punta Caracol
80	7/13/2015	BTH.15.80	Eudendrium capillare	N	Y	N	N	Punta Caracol
81	7/13/2015	BTH.15.81	Pteroclava krempfi	N	Y	Y	N	Punta Caracol
82	7/13/2015	BTH.15.82	Eudendrium capillare	Y	Y	Y	N	Punta Caracol
83	7/13/2015	BTH.15.83	Halopteris alternata	N	Y	Y	N	Punta Caracol
84	7/13/2015	BTH.15.84	Clytia noliformis	Y	Y	N	N	Punta Caracol
85	7/13/2015	BTH.15.85	Bougainvilliidae/?Bimeria	N	Y	Y	MH361365	Punta Caracol
86	7/13/2015	BTH.15.86	Codonorchis sp.	N	Y	N	N	Punta Caracol
87	7/14/2015	BTH.15.87	Solanderia gracilis	Y	Y	Y	MH361366	Swans Cay
88	7/14/2015	BTH.15.88	Eudendrium bermudense	Y	Y	Y	MH361367	Swans Cay
89	7/14/2015	BTH.15.89	Thyroscyphus ramosus	N	Y	Y	MH361368	Swans Cay
90	7/14/2015	BTH.15.90	Pennaria disticha	Y	Y	Y	MH361369	Swans Cay
91	7/14/2015	BTH.15.91	Eudendrium carneum	Y (Female)	Y	Y	MH361370	Swans Cay
92	7/14/2015	BTH.15.92	Stylaster roseus	N	Y	Y	MH361371	Swans Cay
93	7/14/2015	BTH.15.93	Stauridiosarsia nipponica	N	Y	Y	MH361372	Swans Cay
94	7/14/2015	BTH.15.94	Sertularia marginata	N	Y	Y	MH361373	Swans Cay
95	7/14/2015	BTH.15.95	Eudendrium bermudense	Y (male)	Y	Y	N	Swans Cay
96	7/14/2015	BTH.15.96	Pennaria disticha	N	Y	N	MH361374	Swans Cay
97	7/14/2015	BTH.15.97	Macrorhynchia grandis	N	Y	Y	N	Swans Cay
98	7/14/2015	BTH.15.98	Bougainvilliidae 2/?Bimeria	N	Y	Y	MH361375	Swans Cay
99	7/14/2015	BTH.15.99	Clytia sp.	Y	Y	N	MH361376	Swans Cay
100	7/16/2015	BTH.15.100	Antennella secundaria	Y	Y	Y	N	Bastimento vicinity of “Casa Verde”
101	7/15/2015	BTH.15.101	Dentitheca dendritica	N	Y	N	MH361377	The Wall (25mt)/Pandora (20m)
102	7/17/2015	BTH.15.102	Filifera (on hermit crab)/?Turritopsoides	N	Y	N	MH361378	Almirante (Quary’s point)
103	7/17/2015	BTH.15.103	Persa incolorata	Medusa	Y	N	N	Plankton tow near BDT docks
104	7/17/2015	BTH.15.104	Bougainvillia ?pyramidata	Medusa	Y	N	MH361379	Plankton tow near BDT docks
105	7/17/2015	BTH.15.105	Cubaia aphrodite	Medusa	N	Y	N	Plankton tow near BDT docks
106	7/18/2015	BTH.15.106	Sertularia rugosissima	N	Y	Y	N	Bocas del Drago
107	7/18/2015	BTH.15.107	Sertularia rugosissima	N	Y	N	N	Bocas del Drago
108	7/18/2015	BTH.15.108	Rhizogeton sterreri	N	Y	Y	MH361380	Bocas del Drago
109	7/15/2015	BTH.15.109	Sertularella diaphana	N	Y	Y	MH361381	The wall (25mt)/Pandora (20m)
110	7/10/2015	No Voucher	Turritopsis sp.1	Y	N	N	MH029856, MH029857	Close to Bocatorito Bay
111	7/7/2015	No Voucher	Zanclea alba	N	N	N	N	STRI BDT docks/weather station
112	7/7/2015	No Voucher	Millepora complanata	N	N	N	N	STRI BDT docks/weather station
113	7/7/2015	No Voucher	Turritopsis dohrnii	Y	N	N	MH029858, MH029859	Multiple locations

Polyps and medusae were sorted in the laboratory and identified to the lowest taxonomic level using appropriate taxonomic literature [e.g. 16–20]. When possible, polyps from each colony where divided in two vials and preserved in both formalin (for morphological analyses) and 99% ethanol (for genetic analyses). Vouchered specimens were deposited at the Museum of Universidad the Panamá, Panama City, Panama. When little tissue was available, specimens were preserved in ethanol only and used for molecular analyses. DNA samples are currently at the University of Texas A&M at Galveston. Pictures of live specimens featured colony, polyps, medusae (or equivalent reproductive structures), and other morphological structures useful for identification.

Barcoding

Genomic DNA was extracted using standard techniques followed by ethanol precipitation [8]. A ~ 600bp fragment of the large ribosomal subunit of the mitochondrial RNA (lsu-rRNA, 16S) was amplified using SHA and SHB primers [23], Polymerase Chain Reaction (PCR), and conditions as described in Miglietta [15]. The PCR product was run on a 2% agarose gel stained with SybrGreen I nucleic acid gel stain (Sigma-Aldrich) to assay its quantity and quality (i.e. accessory bands). PCR products were purified using exoSAP-it (Affymetrix), following manufacturer’s instruction and used as a template for double stranded sequencing with the amplification primers. The purified DNA was sequenced at the Texas A&M Corpus Christi Genomics Core Laboratory. Sequences were analyzed in Geneious R9 (http://www.geneious.com [24]) and deposited in Genbank (accession numbers MH361321 to MH361381). For definition of anatomical terms used in the ID tables, see online taxonomic glossary for Hydrozoa at https://stricollections.org/portal/misc/glossarycover.php [25].

Results

We collected, vouchered, and fixed in ethanol and/or formalin 112 specimens of Hydrozoa (Table 1). We identified a total of 53 species. An additional 17 taxa could be identified at the genus or family level only due to the lack of fertile structures, small size of the colony, or, in the case of planktonic specimens, early age of the medusae (Table 2 for a complete species list).

Table 2. List of species found in Bocas del Toro during the Hydrozoa Taxonomy course in 2015. For each species we report voucher name(s), number of barcoding sequences (mt lsu-rRNA, 16S) produced per species, and whether their taxonomic description is in the Taxon Identification Tables. A total of 53 taxa were identified at the species level. At the bottom, 17 taxa identified at the genus or higher taxonomic level.

	Species	Voucher Name	Family	Barcoding (n. of sequences)	Taxon Identification Table
1	Turritopsis dohrnii	Yes	Oceaniidae	Yes (2)	Yes
2	Turritopsis sp. 1	BTH.15.110	Oceaniidae	Yes (2)	Yes
3	Turritopsis sp. 4	No	Oceaniidae	No	Yes
4	Rhizogeton sterreri	BTH.15.108	Oceaniidae	Yes (1)	Yes
5	Bougainvillia cf. pyramidata medusae	BTH.15.104	Bougainvilliidae	Yes (1)	No
6	Amphinema dinema	BTH.15.61	Pandeidae	No	Yes
7	Stylaster roseus	BTH.15.25; BTH.15.92	Stylasteridae	Yes (2)	Yes
8	Eudendrium bermudense	BTH.15.17; BTH.15.95; BTH.15.88	Eudendriidae	Yes (1)	Yes
9	Eudendrium capillare	BTH.15.80; BTH.15.82	Eudendriidae	No	Yes
10	Eudendrium carneum	BTH.15.91; BTH.15.15; BTH.15.35	Eudendriidae	Yes (3)	Yes
11	Myrionema amboinense	BTH.15.11	Eudendriidae	No	Yes
12	Ralpharia gorgoniae	BTH.15.24	Tubulariidae	Yes (1)	Yes
13	Millepora alcicornis	BTH.15.31	Milleporidae	Yes (1)	Yes
14	Millepora complanata	Yes Not sampled.	Milleporidae	No	No
15	Solanderia gracilis	BTH.15.87	Solanderiidae	Yes (1)	Yes
16	Pennaria disticha	BTH.15.23; BTH.15.90; BTH.15.96	Pennariidae	Yes (3)	Yes
17	Sphaerocoryne cf. agassizii	BTH.15.40; BTH.15.79; BTH.15.69	Sphaerocorynidae	Yes (2)	Yes
18	Pteroclava krempfi	BTH.15.78; BTH.15.81	Cladocorynidae	Yes (1)	Yes
19	Zanclea alba	Yes Not Vouchered	Zancleidae	No	Yes
20	Aglaophenia latecarinata	BTH.15.20	Aglaopheniidae	No	Yes
21	Macrorhynchia grandis	BTH.15.97	Aglaopheniidae	No	Yes
22	Antennella secundaria	BTH.15.13; BTH.15.100; BTH.15.18	Halopterididae	Yes (2)	Yes
23	Halopteris alternata	BTH.15.50; BTH.15.53; BTH.15.83; BTH.15.14	Halopterididae	Yes (3)	Yes
24	Kirchenpaueria halecioides	BTH.15.1; BTH.15.5; BTH.15.47; BTH.15.21; BTH.15.54	Kirchenpaueriidae	Yes (2)	Yes
25	Dentitheca dendritica	BTH.15.101	Plumulariidae	Yes (1)	Yes
26	Plumularia margaretta	BTH.15.42; BTH.15.51	Plumulariidae	Yes (1)	Yes
27	Plumularia floridana	BTH.15.65	Plumulariidae	Yes (1)	Yes
28	Thyroscyphus marginatus	BTH.15.26; BTH.15.29	Thyroscyphidae	Yes (2)	Yes
29	Thyroscyphus ramosus	BTH.15.89	Thyroscyphidae	Yes (1)	Yes
30	Hincksella formosa	BTH.15.39	Syntheciidae	Yes (1)	Yes
31	Dynamena crisioides	BTH.15.7; BTH.15.9; BTH.15.33; BTH.15.73; BTH.15.74	Sertulariidae	Yes (4)	Yes
32	Dynamena disticha	BTH.15.32	Sertulariidae	Yes (1)	Yes
33	Sertularella diaphana	BTH.15.109	Sertulariidae	Yes (1)	Yes
34	Sertularia rugosissima	BTH.15.106; BTH.15.107	Sertulariidae	No	Yes
35	Sertularia marginata	BTH.15.94	Sertulariidae	Yes (1)	Yes
36	Sertularia distans	BTH.15.43	Sertulariidae	Yes (1)	Yes
37	Salacia desmoides	BTH.15.16	Sertulariidae	Yes (1)	Yes
38	Gastroblasta raffaelei	BTH.15.27; BTH.15.44; BTH.15.46; BTH.15.48; BTH.15.49; BTH.15.55	Campanulariidae	Yes (1)	Yes
39	Clytia gracilis	BTH.15.62	Campanulariidae	No	Yes
40	Clytia hemisphaerica	BTH.15.36; BTH.15.37; BTH.15.41	Campanulariidae	No	Yes
41	Clytia linearis	BTH.15.6; BTH.15.76; BTH15.68	Campanulariidae	Yes (3)	Yes
42	Clytia noliformis	BTH.15.84	Campanulariidae	No	Yes
43	Clytia hummelincki	BTH.15.62; BTH.15.67; BTH.15.72	Campanulariidae	Yes (1)	Yes
44	Obelia bidentata	BTH.15.60; BTH.15.75	Campanulariidae	Yes (1)	Yes
45	Obelia dichotoma	BTH.15.30?, BTH.15.19; BTH.15.34	Campanulariidae	No	Yes
46	Cirrholovenia tetranema	BTH.15.66	Lovenellidae	No	Yes
47	Halecium cf. nanum	BTH.15.45	Haleciidae	Yes (1)	Yes
48	Halecium tenellum	BTH.15.70	Haleciidae	No	Yes
49	Halecium bermudense	BTH.15.2, BTH.15.10	Haleciidae	No	Yes
50	Nemalecium lighti	BTH.15.77; BTH.15.3; BTH.15.4; BTH.15.58	Haleciidae	Yes (3)	Yes
51	Cubaia aphrodite medusa	BTH.15.105	Olindiidae	No	Yes
52	Liriope tetraphylla medusa	BTH.15.63	Geryoniidae	Yes (1)	Yes
53	Persa incolorata medusa	BTH.15.103	Rhopalonematidae	No	Yes
1	Filifera (on hermit Crab: ?Turritopsoides)	BTH.15.102	?	Yes (1)	No
2	?Cytaeis sp.	BTH.15.8	Cytaeididae	No	No
3	Codonorchis sp.	BTH.15.86	Pandeidae	No	Yes
4	Oceaniidae indet	BTH.15.56	Oceaniidae	No	No
5	Oceaniidae indet	BTH.15.59	Oceaniidae	No	No
6	Bougainvilliidae 2/Bimeria?	BTH.15.98	Bougainvilliidae	Yes (1)	No
7	Bougainvilliidae/Bimeria?	BTH.15.85	Bougainvilliidae	Yes (1)	No
8	Stauridiosarsia nipponica	BTH.15.93	Corynidae	Yes (1)	Yes
9	Thecocodium sp.	BTH.15.64	Ptilocodiidae	Yes (1)	No
10	Halecium cf. bermudense	BTH.15.2	Haleciidae	No	No
11	Halecium sp. 1	BTH.15.57	Haleciidae	Yes (1)	No
12	Halecium sp. 2	BTH.15.38	Haleciidae	Yes (1)	No
13	Halecium sp. [sensu 17]	BTH.15.52	Haleciidae	Yes (1)	No
14	Plumularia sp.1	BTH.15.22	Plumulariidae	No	No
15	Plumularia sp.2	BTH.15.12	Plumulariidae	No	No
16	Clytia sp.	BTH.15.99	Campanulariidae	Yes (1)	No
17	Egmundella sp. (on Clytia linearis)	BTH.15.71	Campanulinidae	No	Yes

We sequenced the 16S gene for 64 out of 112 specimens belonging to 44 species. Some of the 112 specimens did not yield sufficient DNA for Polymerase Chain Reaction and thus could not be barcoded. For some species, however, multiple sequences were produced. All sequences were ~ 600bp in length and were deposited in GenBank (accession numbers MH361321-MH361332, MH361334-MH361359, MH361361-MH361381, MH374630).

Morphological, ecological, and barcoding data where assembled in Taxon Identification Tables. We produced 87 tables featuring 28 families (13 in the order Anthoathecata; 12 in the order Leptothecata, 2 in the order Trachymedusae and 1 in the Order Limnomedusae) and 55 species (Appendix 1). These 55 species represent the most common Hydrozoa found in Bocas del Toro during the workshop. For the most speciose taxa we supply identification keys (to genera and/or to species). One of the most abundant families found in the area was the Campanulariidae, with 3 genera and 9 species. Because the polyps of the three genera sampled (Clytia, Gastroblasta, and Obelia) can be easily confused we also supply identification tables for each genus.

Discussion

Check list of Bocas del Toro Hydrozoa

A comprehensive inventory of the Hydrozoa from the Caribbean coast of Panama was previously produced by Calder and Kirkendale [13], who gathered three different collections acquired in 1969, 2002, and 2004, mostly from the Bocas del Toro region. They recorded 79 nominal species (of which were 17 identified at the genus or family level only) belonging to 22 families. We found 53 species, of which only 31 were in common with Calder and Kirkendale [13]. We merged our own inventory with that of Calder and Kirkendale [13],to produce an updated checklist of the Hydrozoa of Bocas del Toro that now comprises 118 taxa, of which 86 identified at the species level and 32 identified at the genus or higher level (Table 3). Given such a small geographical area (250 km²), this is an impressive number. For comparison, 118 species constitutes about ¼ of the total known Hydrozoa species from the Mediterranean Sea [26], and more than a half of the species known from the Arctic [27] or the Antarctic [28]. Seven families and 12 genera are also reported for the first time in Bocas del Toro. Genera added to the checklist are: Amphynema, Cytaeis, Codonorchis, Rhizogeton, Thecocodium, Turritopsoides (?), Gastroblasta, Egmundella, Pteroclava, Cubaia, Lyriope, and Persa. Families new to the Bocas del Toro region are: Pandeidae, Cytaeididae, Cladocorynidae, Ptilocodidae, Olindiidae, Geryoniidae, Rhopalonematidae. Of the new species added to the list, of particular interest is Thecocodium sp. The genus Thecocodium was never reported in the Caribbean and was only recently recorded for the first time in the Atlantic Ocean [29]. Thecocodium sp. (specimen BTH 15.64) presents unique morphological features and may represent a new species. Another species of interest is a species of the genus Coryne (specimen 15.93) found in Swans Key. The colony could not be identified at the species level; however in GenBank its 16S sequence showed 100% identity with Coryne japonica (AY512540) from New Zealand. C. japonica has been reported from the Pacific Ocean but never from the Atlantic Ocean. Our record from Bocas del Toro is the first in the Atlantic Ocean and may represent an introduced species.

Table 3. Updated checklist of the Hydrozoa of Bocas del Toro. The list includes species reported in Calder and Kirkendale [13],and this paper. For the species in this paper, the voucher number is reported. At the bottom, taxa identified at the genus or higher taxonomic level.

	Family	Species	Calder & Kirkerdale [13]	This paper
1	Pandeidae	Amphinema dinema	No	BTH.15.61
2	Cordylophoridae	Corydendrium parasiticum	Yes	No
3	Oceaniidae	Turritopsis dohrnii	No	Yes
4	Oceaniidae	Turritopsis sp. 1	No	BTH.15.110
5	Oceaniidae	Turritopsis sp. 4	No	No
6	Oceaniidae	Turritopsis nutricula	Yes	No
7	Oceaniidae	Rhizogeton sterreri	No	BTH.15.108
8	Bougainvilliidae	Bimeria vestita	Yes	No
9	Bougainvilliidae	Bougainvillia ?pyramidata	No	BTH.15.104
10	Bougainvilliidae	Parawrightia robusta	Yes	No
11	Bougainvilliidae	Silhouetta uvacarpa	Yes	No
12	Stylasteridae	Stylaster roseus	Yes	BTH.15.25; BTH.15.92
13	Eudendriidae	Eudendrium bermudense	Yes	BTH.15.17; BTH.15.95; BTH.15.88
14	Eudendriidae	Eudendrium capillare	Yes	BTH.15.80; BTH.15.82
15	Eudendriidae	Eudendrium carneum	Yes	BTH.15.91; BTH.15.15; BTH.15.35
16	Eudendriidae	Eudendrium sp., aff. album	Yes	No
17	Eudendriidae	Myrionema amboinense	Yes	BTH.15.11
18	Tubulariidae	Ectopleura mayeri	Yes	No
19	Tubulariidae	Ralpharia gorgoniae	Yes	BTH.15.24
20	Tubulariidae	Zyzzyzus calderi	Yes	No
21	Sphaerocorynidae	Sphaerocoryne cf. agassizii	No	BTH.15.40; BTH.15.79; BTH.15.69
22	Sphaerocorynidae	Sphaerocoryne bedoti	Yes	No
23	Cladocorynidae	Pteroclava krempfi		BTH.15.78; BTH.15.81
24	Corynidae	Stauridiosarsia nipponica	No	BTH.15.93
25	Zancleidae	Zanclea alba	Yes	Yes, Not vouchered.
26	Solanderiidae	Solanderia gracilis	Yes	BTH.15.87
27	Pennariidae	Pennaria disticha	Yes	BTH.15.23; BTH.15.28; BTH.15.90; BTH.15.96
28	Milleporidae	Millepora alcicornis	Yes	BTH.15.31
29	Milleporidae	Millepora complanata	Yes	No
30	Milleporidae	Millepora squarrosa	Yes	No
31	Laphoeinidae	Cirrholovenia tetranema	Yes	BTH.15.66
32	Haleciidae	Halecium lightbourni	Yes	No
33	Haleciidae	Halecium nanum	Yes	No
34	Haleciidae	Halecium cf. nanum	No	BTH.15.45
35	Haleciidae	Halecium tenellum	Yes	BTH.15.70
36	Haleciidae	Halecium bermudense	No	BTH.15.2, BTH.15.10
37	Haleciidae	Nemalecium lighti	Yes	BTH.15.77; BTH.15.3; BTH.15.4; BTH.15.58
38	Haleciidae	Sagamihydra dyssymetra	Yes	No
39	Kirchenpaueriidae	Kirchenpaueria halecioides	Yes	BTH.15.1; BTH.15.5; BTH.15.47; BTH.15.21; BTH.15.54
40	Plumulariidae	Dentitheca dendritica	Yes	BTH.15.101
41	Plumulariidae	Monotheca margaretta	Yes	BTH.15.42; BTH.15.51
42	Plumulariidae	Plumularia floridana	Yes	BTH.15.65
43	Plumulariidae	Plumularia setacea	Yes	No
44	Plumulariidae	Plumularia strictocarpa	Yes	No
45	Halopterididae	Antennella curvitheca	Yes	No
46	Halopterididae	Antennella secundaria	Yes	BTH.15.13; BTH.15.100; BTH.15.18
47	Halopterididae	Halopteris alternata	Yes	BTH.15.50; BTH.15.53; BTH.15.83; BTH.15.14
48	Halopterididae	Halopteris carinata	Yes	No
49	Aglaopheniidae	Aglaophenia dubia	Yes	No
50	Aglaopheniidae	Aglaophenia latecarinata	Yes	BTH.15.20
51	Aglaopheniidae	Macrorhynchia philippina	Yes	No
52	Aglaopheniidae	Macrorhynchia grandis		BTH.15.97
53	Thyroscyphidae	Thyroscyphus marginatus	Yes	BTH.15.26; BTH.15.29
54	Thyroscyphidae	Symmetroscyphus intermedius	Yes	No
55	Thyroscyphidae	Thyroscyphus ramosus	Yes	BTH.15.89
56	Syntheciidae	Hincksella formosa	Yes	BTH.15.39
57	Syntheciidae	Synthecium tubithecum	Yes	No
58	Sertulariidae	Diphasia tropica	Yes	No
59	Sertulariidae	Dynamena crisioides	Yes	BTH.15.7; BTH.15.9; BTH.15.33; BTH.15.73; BTH.15.74
60	Sertulariidae	Dynamena disticha	Yes	BTH.15.32
61	Sertulariidae	Dynamena quadridentata	Yes	No
62	Sertulariidae	Sertularella cylindritheca	Yes	No
63	Sertulariidae	Sertularella diaphana	No	BTH.15.109
64	Sertulariidae	Sertularella hartlaubi	Yes	No
65	Sertulariidae	Sertularia rugosissima	No	BTH.15.106; BTH.15.107
66	Sertulariidae	Sertularia loculosa	Yes	No
67	Sertulariidae	Sertularia marginata	Yes	BTH.15.94
68	Sertulariidae	Tridentata subtilis	Yes	No
69	Sertulariidae	Sertularia turbinata	Yes	No
70	Sertulariidae	Sertularia distans	No	BTH.15.43
71	Sertulariidae	Sertularia vervoorti	Yes	No
72	Sertulariidae	Salacia desmoides	No	BTH.15.16
73	Campanulariidae	Gastroblasta raffaelei	No	BTH.15.27; BTH.15.44; BTH.15.46; BTH.15.48; BTH.15.49; BTH.15.55
74	Campanulariidae	Clytia gracilis	Yes	BTH.15.62
75	Campanulariidae	Clytia hemisphaerica	Yes	BTH.15.36; BTH.15.37; BTH.15.41
76	Campanulariidae	Clytia linearis	Yes	BTH.15.6 ; BTH.15.76; BTH15.68
77	Campanulariidae	Clytia paulensis	Yes	No
78	Campanulariidae	Clytia stolonifera	Yes	No
79	Campanulariidae	Clytia gracilis	No	BTH.15.62
80	Campanulariidae	Clytia noliformis	No	BTH.15.84
81	Campanulariidae	Clytia hummelincki	No	BTH.15.62; BTH.15.67; BTH.15.72
82	Campanulariidae	Obelia bidentata	Yes	BTH.15.60; BTH.15.75
83	Campanulariidae	Obelia dichotoma	Yes	BTH.15.30?, BTH.15.19; BTH.15.34
84	Olindiidae	Cubaia aphrodite medusa	No	BTH.15.105
85	Geryoniidae	Liriope tetraphylla medusa	No	BTH.15.63
86	Rhopalonematidae	Persa incolorata medusa	No	BTH.15.103
	Family	Species	Calder	This paper
1	?	Filifera (on Hermit Crab) (?Turritopsoides)	No	BTH.15.102
2	Cytaeididae	?Cytaeis sp.	No	BTH.15.8
3	Pandeidae	Codonorchis sp.	No	BTH.15.86
4	Cordylophoridae	Rhizodendrium sp.	Yes	No
5	Oceaniidae	Oceaniidae indet.	No	BTH.15.56
6	Oceaniidae	Oceaniidae indet.	No	BTH.15.59
7	Bougainvilliidae	Bougainvilliidae 2/?Bimeria	No	BTH.15.98
8	Bougainvilliidae	Bougainvilliidae/?Bimeria	No	BTH.15.85
9	Bougainvilliidae	Bougainvilliidae indet.	Yes	No
10	Eudendriidae	Eudendrium sp.	Yes	No
11	Corynidae	Coryne sp.	Yes	No
12	Corynidae	Corynidae indet.	Yes	No
13	Zancleidae	Zanclea sp.	Yes	No
14	Hydrocorynidae	Hydrocoryne sp.	Yes	No
15	Ptilocodiidae	Thecocodium sp.	No	BTH.15.64
16	Haleciidae	Halecium cf. bermudense	No	BTH.15.2
17	Haleciidae	Halecium sp.	Yes	No
18	Haleciidae	Halecium sp. 1	No	BTH.15.57
19	Haleciidae	Halecium sp. 2	No	BTH.15.38
20	Haleciidae	Halecium sp. [sensu 17]	No	BTH.15.52
21	Haleciidae	Hydranthea sp.	Yes	No
22	Plumulariidae	Plumularia sp.	No	BTH.15.22
23	Plumulariidae	Plumularia sp.	No	BTH.15.12
24	Campanulariidae	Clytia sp., aff. kincaidi	Yes	No
25	Campanulariidae	Clytia sp.	No	BTH.15.99
26	Campanulariidae	Clytia sp. A	Yes	No
27	Campanulariidae	Clytia sp. B	Yes	No
28	Campanulariidae	Clytia sp. C	Yes	No
29	Campanulariidae	Orthopyxis sp.	Yes	No
30	Halammohydridae	Halammohydra sp.	Yes	No
31	Otohydridae	Otohydra sp.	Yes	No
32	Campanulinidae	Egmundella sp. (on Clytia linearis)	No	BTH.15.71

Taxon identification sheets

Taxon Identification Sheets collate the taxonomic description of 56 species found during the Hydrozoa Taxonomy course and confidently identified at the species level. They also include a brief synopsis of the 28 families they belong to (see Tables 1–88 in Supplementary Materials. For each family we included authorship, corresponding Order, number of species (worldwide and in Bocas del Toro), morphologically similar taxa that could be mistakenly identified as member of the family of interest, and their key diagnostic characters. The species identification sheets include author, diagnostic characters of the colony and their reproductive structures (medusae, eumedusoids, or fixed gonophores), species ecology, species distribution in Bocas del Toro, number of specimens collected, pictures and, when available, GenBank accession numbers for their 16S sequence(s). These species identification sheets assemble in simple format information that can be used for a correct identification. The morphological description, pictures, and link to their 16S barcoding sequence represent a comprehensive display of information that integrates traditional and modern taxonomy into a practical tool to aid identification of the most commons species of Hydrozoa in the shallow waters of the Archipiélago de Bocas del Toro, Panamá.

Conclusive remarks

Knowledge on the local biodiversity is an essential pre-requisite for the monitoring and management of environmental assets and ecosystem health worldwide. The present inventory of the marine hydrozoan fauna in the Bocas del Toro shallow water is far to be exhaustive due to the inherent limitation of our sampling efforts, based mostly on snorkeling and more rarely on SCUBA diving collections. However, the high number of recorded taxa suggests that the Caribbean Sea should be considered a region of high hydrozoan diversity. Paradoxically, taxonomy is a science at brink of extinction. The ARTS courses have been devoted not only to increase knowledge on local biodiversity, but towards the conservation and promotion of biodiversity expertise. More generally, training a new generation of taxonomists is a current challenge and a mandatory urge to understanding ecosystem functioning in face of local and global changes, and to address the needs of sustainability of humankind activities.

Author contributions

MPM and S. Piraino designed the experiments; S. Pruski produced the barcoding sequences, all authors collected samples and contributed the Taxon Identification Tables: MPM wrote the paper.

Acknowledgments

The authors would like to thank the National Science Foundation for funding. We would also like to thank the staff at the Smithsonian Tropical Research Institute’s Bocas del Toro Research Station for their support and help during the 2015 taxonomy course, the Panamanian Ministry of the Environment (MiAmbiente) for permission to conduct this research, and two anonymous reviewers for their useful comments.

Associate Editor: Federico Brown

Disclosure statement

No potential conflict of interest was reported by the authors.

Supplementary material

Supplemental data can be accessed here.

Additional information

Funding

This study was funded by the National Science Foundation ARTS grant number DEB-1456501 to MPM and DEB-1456674 to RC, the Texas Sea Grantnumber 02-S170210 and the Texas A&M Pesca Grant to MPM.