Table of Contents

• Coverage
• Dataset Description
  • Methods & Sampling
  • BCO-DMO Processing Description
• Data Files
• Related Publications
• Parameters
• Instruments
• Project Information
• Program Information
• Funding

Sample collection:
Sponges were photographed in situ and collected from a variety of habitats including: Autonomous Reef Monitoring Structures (ARMS), floating docks, pier pilings, marine mammal pens, and shallow (0.3 to 3 meters (m)) natural reef environments in Kāne'ohe Bay, Ke'ehi Harbor and Shark's Cove on the island of O'ahu, Hawai'i. Collections from ARMS were conducted monthly throughout a 2-year sampling period (July 2016 through June 2018) following methods in (Timmers et al. 2020) and Vicente et al. (2022). Sponge species were mapped using the ggmap v.3.0.901 package (Kahle & Wickham 2013) in R v.4.3.2 (R Core Team 2023). Field observations of morphology, color, consistency, surface, and oscules for each specimen were recorded. Samples were preserved in 95% ethanol and when enough material was available, were also fixed in 4% paraformaldehyde (PFA) for 24 hours and then transferred to 70% ethanol. Type and other specimens were deposited in the Florida Museum of Natural History (catalogue numbers beginning with acronym UF) in Florida, USA, and the Bernice Pauahi Bishop Museum (catalogue numbers beginning with acronym BPBM) in O'ahu, USA. Samples from Kāneʻohe Bay were collected under special activities collection permits SAP2018-03 and SAP2019-06 (covering the period of January 13, 2017, through April 10, 2019) issued by the State of Hawai'i Division of Aquatic Resources. Samples from Pūpūkea Marine Life Conservation District were collected under permit SAP2023-20 (covering the period of March 11, 2022, through March 10, 2023). Samples from Kewalo and Ke'ehi harbors were collected by the Division of Aquatic Resources Aquatic Invasive Species team, who do not require special activities permits.

DNA extraction, sequencing, and assembly:
Subsamples of sponge tissue (30 milligrams (mg)) were removed from type material preserved in 95% ethanol and were processed for DNA extraction. Two different approaches were utilized for DNA extraction and downstream analysis. First, we followed protocols in Vicente et al. (2022) for DNA extractions, polymerase chain reactions (PCR) for amplifying partial fragments of both 28S rRNA and COI genes and for Sanger sequencing. Forward and reverse reads were assembled, trimmed, and edited by eye using Geneious 10 (Kearse et al. 2012). Sequences were checked for contamination using the online BLAST server (Altschul et al. 1990) and results that showed at least 85% sequence identity to sponges were used for subsequent analysis. All assembled chromatograms resulted in >90% high-quality base pair reads with a mean Phred quality score 40. Second, total DNA from holotypes of Haliclona (Gellius) loe BPBM C1523, Haliclona (Reniera) kahoe BPBM C1539, Haliclona (Rhizoniera) pahua BPBM C1518, vouchers BPBM C1519, BPBM C1510 for Haliclona (Soestella) caerulea and Gelliodes conulosa, respectively, were extracted with a phenol-chloroform method modified from Saghai-Maroof et al. (1984) and used directly for library preparation with the Illumina True-Seq PCR free kit and low-coverage of whole genome sequencing on Illumina NovaSeq 6000 at the Iowa State University DNA facility. A detailed protocol for mitochondrial and ribosomal genome sequences for these samples can be found in Lavrov et al. (2025).

Phylogenetic analysis:
Haplosclerida sequences within GenBank closely resembling (>90% sequence identity) new sequences in our study were selected for the phylogenetic analysis. These only included sequences from species associated with voucher specimens associated with a peer-reviewed publication authored or coauthored by taxonomists. ClustalW with default parameters was used for aligning partial and complete 28S rRNA and COI sequences. Alignments consisted of 300 bp of the 28S and 480 bp of the COI gene sequence. RaxML (Stamatakis 2006) included Geneious 10 was used for maximum likelihood (ML) analysis with the GTR+GAMMA model of nucleotide substitution, 100 starting maximum parsimony trees, and 1,000 bootstrap replicates. Resulting bootstrap values of >50 from the ML posterior probabilities are shown on the tree. Phylogenetic trees were rooted on Ephydatia fluviatilis OX175335.1 and ON000190.1 for 28S and COI, respectively. Sequences of holotypes and other specimens for each species were deposited to GenBank under accession numbers: MW016123, MT452542, MT586742, MW016124, MW059074, MW143255, MW016168, MW059064, MW059059, MW016133, MT586743, MW016360, MW016153, MW016155, MW016154, MW059075, MW016154. 

Sectioning and spicule preparation:
Sponge pieces (3 to 5 cubic millimeters) containing both ectosomal and choanosomal tissue fixed in either 4% PFA or 95% ethanol were transferred to 70% ethanol. Sponge pieces were dehydrated in an alcohol series of 35%, 50% and 70%, 100% and embedded in paraffin. Sections >100 micrometers (μm) thick were cut perpendicular to the surface through the ectosome and choanosome with a microtome. In specimens where the ectosome was specialized, tangential sections across the sponge surface were made at 100 μm thickness. Small pieces were also boiled in nitric acid for 1 to 2 minutes or until solution turned clear. Spicules were let to settle, and the acid was discarded. Spicules were then rinsed two times with distilled water to remove the acid; water was then changed to 95% ethanol for storage. Spicules were suspended and a few drops were observed under light microscopy, photographed, and measured using ImageJ (Abràmofff et al. 2005) http://imagej.nih.gov/ij/). Fifty oxeas and, if present, ten sigmas per species were measured [lengths and widths, expressed herein as minimum–mean [±1 standard deviation (SD)]–maximum length / width in μm (n)]. A few drops of the spicule suspension were added to a stub, air dried, and imaged under a Hitachi S-4800 FESEM Scanning Electron Microscope (SEM) at the Biological Electron Microscope Facility at the University of Hawai'i Mānoa.

Summarizing morphological characters of congeneric comparative material:
Sponge species found throughout the Hawaiian Archipelago are shared with the Central Indo-Pacific (Australia, Philippines, the Mariana Archipelago), Temperate Australasia (New Zealand), Temperate Northern Pacific (Japan), the Eastern Mexican Pacific, Caribbean, Mediterranean and the Northeast Atlantic (Bergquist 1967; Carballo et al. 2013; de Laubenfels 1950; van Soest et al. 2021). We used the World Porifera database to include a summary of morphological characters for 51 Haliclona spp. closely resembling the new species in this study. These species were selected from geographic locations with shared species with the Hawaiian Archipelago. Species from the Temperate Atlantic, Black Sea, Arctic, or Southern Ocean were considered improbable species due to geographic barriers and temperate climates. Species from these ecoregions were therefore disregarded as comparative material.

- Converted original file "Vicenteetal2025_Zootaxa_primary file.xls" to CSV format.
- Imported file "Vicenteetal2025_Zootaxa_primary file.csv" into the BCO-DMO system.
- Marked "NaN" as a missing data value (missing data are empty/blank in the final CSV file).
- Renamed fields to comply with BCO-DMO naming conventions.
- Replaced non-standard character em dash ("−") with hyphen ("-") in the length and width columns.
- Replaced non-standard character "±" with "+/-" in the length and width columns.
- Converted Date format to YYYY-MM-DD.
- Saved the final file as "986889_v1_haplosclerid_sponges_hi.csv".

NSF Award Abstract:
Coral reefs are among the most species-rich ecosystems on the planet, occupying only about 1% of the seafloor, but housing more than a quarter of known marine biodiversity. Sometimes called the rainforests of the sea, coral reefs have great intrinsic biological, cultural and economic value. Nearly a billion people across the planet rely on coral reef ecosystems as a significant source of their diet, and the annual economic benefits of coral reefs are estimated to be around $9.9 Trillion USD. Thus, the global decline of coral reefs by an estimated 30-50% since the 1980s is of considerable concern as scientists struggle to understand whether species are being lost before they are even discovered. While coral reefs are spectacularly diverse, the majority of this biodiversity actually lives hidden deep within the three-dimensional framework of the reef itself. This hidden (or cryptic) community of organisms are both dramatically understudied and fundamentally important for the persistence of coral reefs. Sponges are a dominant group among these cryptic organisms within the reef which provide food from the bottom of the food chain and help sustain coral reef biodiversity. Despite the vital ecological role of sponges on coral reefs, little is known about their diversity, abundance or species ranges across the Indo-Pacific. For example, the most striking marine biodiversity gradient on the planet is described from several of the visibly dominant groups on coral reefs, including corals and reef fishes. From the global hotspot of species richness in the Indo-Pacific Coral Triangle there is a sharp eastward decline in species numbers to more remote oceanic islands in the Central Pacific, such as the Hawaiian Archipelago. However, no survey to date has evaluated whether the diversity of poorly known cryptic coral reef species, such as sponges, show the same pattern as the visible species that dominate the surface of the reef. Summer training modules introduce at-risk Pacific Islander youth to coral reef biodiversity to recruit and train a new generation of sponge taxonomists. Identification guides are being produced to help resource managers in establishing a baseline of sponge diversity, which allows resource managers to identify and protect native species, improves detection of alien species introductions and serves as a tool for monitoring changes in the ecosystem in response to human impacts. The work is being disseminated widely through scientific literature, public and professional presentations, popular press articles, and an educational display about sponges and coral reef biology in collaboration with the Waikīkī Aquarium.

This important knowledge gap is addressed by analyzing an existing backlog of standardized sampling devices (ARMS) collected from throughout the Pacific Ocean to determine whether sponges that live largely unseen within the reef framework follow the same diversity gradient as has been previously reported for fish and corals. By integrating taxonomy with multi-locus DNA barcoding and metabarcoding, this project is documenting species richness and biodiversity patterns among the cryptic sponge community across five ecoregions spanning over 10,000 km of the tropical Pacific. These collections include many new species and are providing vouchered DNA barcodes to existing reference databases that currently include fewer than 1% of sponge species across the planet. Sponges are a rich source for pharmaceutical development, so discovery of new species also provides opportunity for exploration of natural products from both the sponges and culturable microbes associated with them. By examining sponge species occurrence and diversity along both environmental and anthropogenic gradients in each ecoregion, the data also address whether coral reef sponges can serve as indicators of human impacts. Collectively, these results are transforming our knowledge of tropical Pacific sponge biodiversity, species ranges, and providing much-needed reference barcodes to global sequence databases. By determining whether sponges show the same Indo-Pacific richness gradient as reported in fishes and corals, this project is testing how well generalizations made from the visible subset of species that live on the surface of coral reefs apply to rest of coral reef biodiversity. This study is greatly advancing our knowledge of Pacific coral reef sponges and will ultimately inform the scale over which vital ecological roles performed by this understudied taxon, such as the production of nutrients at the bottom of the food chain, are acting across the Pacific.

This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

Description from NSF award abstract:
The objective of this Research Coordination Network project is to develop an international network of researchers who use genetic methodologies to study the ecology and evolution of marine organisms in the Indo-Pacific to share data, ideas and methods. The tropical Indian and Pacific Oceans encompass the largest biogeographic region on the planet, the Indo-Pacific. It spans over half of the Earth's circumference and includes the exclusive economic zones of over 50 nations and territories. The Indo-Pacific is also home to our world's most diverse marine environments. The enormity and diversity of the Indo-Pacific poses tremendous logistical, political and financial obstacles to individual researchers and laboratories attempting to study the marine biology of the region. Genetic methods can provide invaluable information for our understanding of processes ranging from individual dispersal to the composition and assembly of entire marine communities.

The project will:
(1) assemble a unique, open access database of population genetic data and associated metadata that is compatible with the developing genomic and biological diversity standards for data archiving,
(2) facilitate open communication and collaboration among researchers from across the region through international workshops, virtual communication and a collaborative website,
(3) promote training in the use of genetic methodologies in ecology and evolution for researchers from developing countries through these same venues, and
(4) use the assembled database to address fundamental questions about the evolution of species and the reservoirs of genetic diversity in the Indo-Pacific.

The network will provide a model for international collaborative networks and genetic databasing in biodiversity research that extends beyond the results of this Research Coordination Network effort.