Macrofauna collected on colonization panels at Snail Vent Field on the Mariana Back-arc in 2014 RAPID: Larval Abundance, Behavior and Dispersal at Deep-sea Hydrothermal Vents in the Southern Mariana Trough (Mariana Back-arc Vents)

This dataset indicates organisms, identified to varying levels of taxonomic granularity, present on colonization surfaces deployed at deep-sea vents in the West Pacific. Description The "Submarine Ring of Fire 2014 – Ironman" Expedition was a cooperative venture with two science parties with overlapping interests and goals. Craig Moyer from Western Washington University led a group funded by the National Science Foundation to conduct research on iron-oxidizing bacteria at hydrothermal vents. Bill Chadwick led a NOAA/PMEL group funded by the NOAA Ocean Exploration and Research Program and focused on the chemical and biological impacts of hydrothermal vent emissions from active submarine volcanoes in the Mariana arc. Additional cruise information and original data are available from the NSF R2R data catalog. NSF Award Abstract: Summary: Since the discovery of deep-sea hydrothermal vents over thirty years ago, scientists have been perplexed by the question: How are these vent sites colonized and, more specifically, How are the faunal populations established and maintained at these very discrete and often ephemeral habitats. For animals that are sessile or have limited mobility as adults, dispersal to these habitats occurs early in the life cycle, as planktonic larvae in the water column. Due to the difficulties in sampling deep-sea larvae, including low abundances (dilute concentrations), we have very few quantitative estimates of larval dispersal between or larval supply to hydrothermal vents. We also have little to no knowledge of the behavior of vent larvae. The PIs will use large-volume plankton pumps to collect larvae near vents in the southern Mariana Trough in a collaborative effort to quantify larval abundance, behavior, and dispersal in this little-studied region. The collaboration combines the PI's strengths in the collection and morphological identification of larvae and quantifying and modeling dispersal between deep-sea vents, and those of Japanese partners in rearing larvae of hydrothermal vent fauna, molecular genetic identification of larvae, and population genetics of vent fauna. Intellectual merit: The southern Mariana Trough is a very interesting region in which to study dispersal of vent-endemic fauna, due to the proximity of vents in the back-arc spreading center to vents along the Mariana Arc. These two tectonic settings create different habitat conditions and support vent communities with different species composition. Vent sites the PIs will visit, in the axis and just off-axis of the back-arc spreading center are as close as 25 km to vents on the arc, yet 600 km south of the other known vents in the back-arc. In addition to the new information on larval abundance, diversity, behavior, and dispersal that will be gained for this little-studied region of the world's ridge system, this project has direct relevance to the integration and synthesis goals of the U.S. Ridge 2000 Program. The PI's lab group has conducted previous work at Integrated

sandwiches during recovery was 17.36 deg C. Sandwiches were initially placed into a biobox onboard the ROV, then transferred to another biobox on an elevator for recovery. During this transfer, as well as during the recovery of the elevator at the surface, some specimens may have been lost.

Shipboard Sample Processing
Sandwiches were removed from the biobox into individual bags and the elevator biobox washings were siphoned into a jar. The bags and jar were filled with ammonium sulfate solution buffered with EDTA and sodium citrate. We want to thank Shawn Arellano for this sample recovery and shipboard sample processing.

Laboratory Sorting and Morphological Identification
Samples remained in the ammonium sulfate solution for 1 to 1.5 years. Samples were rinsed with TE buffer prior to placement in 95% ethanol. The sandwiches were manually disassembled, and each entire plate-inclusive of both sides and edges-was examined for all attached macrofauna under a dissecting microscope at 25x. Washings were passed over a 63 micron sieve, and also examined at 25x.
Macrofauna, including foraminifera, were sorted to lowest taxonomic level, tallied, and placed into separate vials. Ciliates were tallied, but not usually retained; some were placed into sorted vials. Morphological identification was limited due to the degradation of the sample. We consulted with international experts to identify many of the morphotypes. All morphotypes, except the ciliates, were photographed. Potentially, ciliates could be identified through metabarcoding of the sorted vials. All photographs were by Mary Toner, with the exception of barnacles by Hiromi Watanabe.
This dataset is complete for one of the three "sandwiches", and for the biobox washings.

Genetic Barcoding
A subset of eight individuals, representing eight morphotypes, was prepared for the Canadian Centre for DNA Barcoding. We used sterile techniques to provide snips of larger specimens.
The five submitted in 2017 used the CCDB regular protocol for extraction, amplification, (with two primer sets, C_LepFoIF/C_LepFOIR and ZplankF1/ZplankR1) and sequencing for the Cytochrome Oxidase Subunit 1 -5 prime region (COI). The three submitted in 2018 used the CCDB protocol for Next Generation Sequencing for COI. Results may be viewed in the Barcode of Life Data System (BOLD) Public Data Portal using the project search SNLBE. We also consulted with international experts on genetic sequences.

Problem Report
One set of colonization "sandwiches" could not be recovered, as it was buried by a partial collapse of surrounding rock. Some specimens may have been lost during a transfer of samples between bioboxes at the (ZIP Archive (ZIP), 160.08 MB) MD5:4f468227727ebc3f03fce6cd3f354e61 seafloor, as well as upon recovery at the surface. As such, our counts cannot be used for quantitative analysis of hydrothermal macrofauna.

Data Processing
Identification information and counts were stored in tables, including manual assignment to WoRMS AphiaIDs. These tables were cleaned, organized, and joined in R (v. 3.4.1, "Single Candle") using R Studio (v. 1.0.153), then manually verified. We used the R packages worrms, taxize, tidyr, dplyr, and tibble.  instruments. Also called a "light microscope".
[  No other such comparison has been possible due to the lack of sampling effort for larvae with large-volume pumps. Also, they are proposing the first experiments with live vent larvae (to the best of our knowledge -with the exception of brachyuran megalopae at 1 atm) to estimate swimming and sinking rates that are important for adding behavioral information to models of larval dispersal. Broader Impacts: The project involves reciprocal training and cultural exchange -the PIs will learn field and laboratory research techniques from the Japanese PIs, and they will learn from the U.S. PIs. The project will also benefit the career development of a junior researcher (Beaulieu). The proposed activity broadens the participation of both U.S. and Japanese women scientists in sea-going, oceanographic research. The PIs will broadcast the cruise activities in a web log posted by the international InterRidge Program Office, and they anticipate at least three scientific publications will emerge.
New species will be added to the online photographic identification guide for vent larvae and included in the second edition of the printed guide. Additional cruise data and information are available from MGDS: http://www.marine-geo.org/tools/search/entry.php?id=YK10-11 [