Transcriptome-wide analysis of the response of the thecosome pteropod Clio pyramidata to short-term CO2 exposure

Author Posting. © The Author(s), 2015. This is the author's version of the work. It is posted here by permission of Elsevier for personal use, not for redistribution. The definitive version was published in Comparative Biochemistry and Physiology Part D: Genomics and Proteomics 16 (2015): 1-9, doi:10.1016/j.cbd.2015.06.002.Thecosome pteropods, a group of calcifying holoplanktonic molluscs, have recently become a research focus due to their potential sensitivity to increased levels of anthropogenic dissolved CO2 in seawater and the accompanying ocean acidification. Some populations, however, already experience high CO2 in their natural distribution during diel vertical migrations. To achieve a better understanding of the mechanisms of pteropod calcification and physiological response to this sort of short duration CO2 exposure, we characterized the gene complement of Clio pyramidata, a cosmopolitan diel migratory thecosome, and investigated its transcriptomic response to experimentally manipulated CO2 conditions. Individuals were sampled from the Northwest Atlantic in the fall of 2011 and exposed to ambient conditions (~380 ppm) and elevated CO2 (~800 ppm, similar to levels experienced during a diel vertical migration) for ~10 hrs. Following this exposure the respiration rate of the individuals was measured. We then performed RNA-seq analysis, assembled the C. pyramidata transcriptome de novo, annotated the genes, and assessed the differential gene expression patterns in response to exposure to elevated CO2. Within the transcriptome, we identified homologs of genes with known roles in biomineralization in other molluscs, including perlucin, calmodulin, dermatopontin, calponin, and chitin synthases. Respiration rate was not affected by short-term exposure to CO2. Gene expression varied greatly among individuals, and comparison between treatments indicated that C. pyramidata down-regulated a small number of genes associated with aerobic metabolism and up-regulated genes that may be associated with biomineralization, particularly collagens and C- type lectins. These results provide initial insight into the effects of short term CO2 exposure on these important planktonic open-ocean calcifiers, pairing respiration rate and the gene expression level of response, and reveal candidate genes for future ecophysiological, biomaterial and phylogenetic studies.The Extreme Science and Engineering Discovery Environment (XSEDE), which is supported by National Science Foundation grant number OCI-1053575, provided computing resources for the differential expression analysis. This material is based upon work supported by the National Science Foundation’s Ocean Acidification Program under grant number OCE-1041068 (to Lawson, Wang, Lavery, and Wiebe), the Woods Hole Oceanographic Institution’s Access to the Sea program (to Tarrant, Maas and Lawson) and the WHOI postdoctoral scholarship program (to Maas)

The metabolic response of thecosome pteropods from the North Atlantic and North Pacific oceans to high CO2 and low O2

© The Author(s), 2016. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Biogeosciences 13 (2016): 6191-6210, doi:10.5194/bg-13-6191-2016.As anthropogenic activities directly and indirectly increase carbon dioxide (CO2) and decrease oxygen (O2) concentrations in the ocean system, it becomes important to understand how different populations of marine animals will respond. Water that is naturally low in pH, with a high concentration of carbon dioxide (hypercapnia) and a low concentration of oxygen, occurs at shallow depths (200–500 m) in the North Pacific Ocean, whereas similar conditions are absent throughout the upper water column in the North Atlantic. This contrasting hydrography provides a natural experiment to explore whether differences in environment cause populations of cosmopolitan pelagic calcifiers, specifically the aragonitic-shelled pteropods, to have a different physiological response when exposed to hypercapnia and low O2. Using closed-chamber end-point respiration experiments, eight species of pteropods from the two ocean basins were exposed to high CO2 ( ∼  800 µatm) while six species were also exposed to moderately low O2 (48 % saturated, or  ∼  130 µmol kg−1) and a combined treatment of low O2/high CO2. None of the species tested showed a change in metabolic rate in response to high CO2 alone. Of those species tested for an effect of O2, only Limacina retroversa from the Atlantic showed a response to the combined treatment, resulting in a reduction in metabolic rate. Our results suggest that pteropods have mechanisms for coping with short-term CO2 exposure and that there can be interactive effects between stressors on the physiology of these open ocean organisms that correlate with natural exposure to low O2 and high CO2. These are considerations that should be taken into account in projections of organismal sensitivity to future ocean conditions.This work was funded by the National Science Foundation’s Ocean Acidification Program (grant OCE-1041068), the National Institute of Standards and Technology (NIST-60NANB10D024), and the WHOI postdoctoral scholarship program

Energetic Plasticity Underlies a Variable Response to Ocean Acidification in the Pteropod, \u3cem\u3eLimacina helicina antarctica\u3c/em\u3e

Ocean acidification, caused by elevated seawater carbon dioxide levels, may have a deleterious impact on energetic processes in animals. Here we show that high PCO2 can suppress metabolism, measured as oxygen consumption, in the pteropod, L. helicina forma antarctica, by ~20%. The rates measured at 180–380 µatm (MO2 = 1.25 M−0.25, p = 0.007) were significantly higher (ANCOVA, p = 0.004) than those measured at elevated target CO2 levels in 2007 (789–1000 µatm, = 0.78 M−0.32, p = 0.0008; Fig. 1). However, we further demonstrate metabolic plasticity in response to regional phytoplankton concentration and that the response to CO2 is dependent on the baseline level of metabolism. We hypothesize that reduced regional Chl a levels in 2008 suppressed metabolism and masked the effect of ocean acidification. This effect of food limitation was not, we postulate, merely a result of gut clearance and specific dynamic action, but rather represents a sustained metabolic response to regional conditions. Thus, pteropod populations may be compromised by climate change, both directly via CO2-induced metabolic suppression, and indirectly via quantitative and qualitative changes to the phytoplankton community. Without the context provided by long-term observations (four seasons) and a multi-faceted laboratory analysis of the parameters affecting energetics, the complex response of polar pteropods to ocean acidification may be masked or misinterpreted

The metabolic response of pteropods to acidification reflects natural CO2-exposure in oxygen minimum zones

© The Author(s), 2012. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Biogeosciences 9 (2012): 747-757, doi:10.5194/bg-9-747-2012.Shelled pteropods (Thecosomata) are a group of holoplanktonic mollusks that are believed to be especially sensitive to ocean acidification because their aragonitic shells are highly soluble. Despite this concern, there is very little known about the physiological response of these animals to conditions of elevated carbon dioxide. This study examines the oxygen consumption and ammonia excretion of five pteropod species, collected from tropical regions of the Pacific Ocean, to elevated levels of carbon dioxide (0.10%, 1000 ppm). Our results show that pteropods that naturally migrate into oxygen minimum zones, such as Hyalocylis striata, Clio pyramidata, Cavolinia longirostris and Creseis virgula, were not affected by carbon dioxide at the levels and duration tested. Diacria quadridentata, which does not migrate, responds to high carbon dioxide conditions with reduced oxygen consumption and ammonia excretion. This indicates that the natural chemical environment of individual species may influence their resilience to ocean acidification.Funding of the National Science Foundation (grant OCE-0526502 to Wishner and Seibel, OCE – 0526545 to Daly, and OCE – 0851043 to Seibel), the University of Rhode Island, and the Rhode Island Experimental Program to Stimulate Competitive Research Fellowship Program

Assembly of a reference transcriptome for the gymnosome pteropod Clione limacina and profiling responses to short-term CO2 exposure

© The Author(s), 2017. This is the author's version of the work. It is posted here under a nonexclusive, irrevocable, paid-up, worldwide license granted to WHOI. It is made available for personal use, not for redistribution. The definitive version was published in Marine Genomics 34 (2017): 39-45, doi:10.1016/j.margen.2017.03.003.The gymnosome (unshelled) pteropod Clione limacina is a pelagic predatory mollusc found in polar and sub-polar regions. It has been studied for its distinctive swimming behavior and as an obligate predator on the closely related thecosome (shelled) pteropods. As concern about ocean acidification increases, it becomes useful to compare the physiological responses of closely-related calcifying and non-calcifying species to acidification. The goals of this study were thus to generate a reference transcriptome for Clione limacina, to expose individuals to CO2 for a period of 3 days, and to explore differential patterns of gene expression. Our Trinity assembly contained 300,994 transcripts of which ~26% could be annotated. In total, only 41 transcripts were differentially expressed following the CO2 treatment, consistent with a limited physiological response of this species to short-term CO2 exposure. The differentially expressed genes identified in our study were largely distinct from those identified in previous studies of thecosome pteropods, although some similar transcripts were identified, suggesting that comparison of these transcriptomes and responses may provide insight into differences in OA responses among phylogenetically and functionally distinct molluscan lineages.A. Thabet is grateful for a fellowship from the Egyptian Culture and Education Bureau and for mentoring from Drs. M.M. Sarhan and M.M. Fouda. Funding for this research was provided by a National Science Foundation grant to Lawson, Maas, and Tarrant (OCE-1316040)

Exposure to CO2 influences metabolism, calcification and gene expression of the thecosome pteropod Limacina retroversa

Author Posting. © The Company of Biologists, 2018. This article is posted here by permission of The Company of Biologists for personal use, not for redistribution. The definitive version was published in Journal of Experimental Biology 221 (2018): jeb164400, doi:10.1242/jeb.164400.Thecosomatous pteropods, a group of aragonite shell-bearing zooplankton, are becoming an important sentinel organism for understanding the influence of ocean acidification on pelagic organisms. These animals show vulnerability to changing carbonate chemistry conditions, are geographically widespread, and are both biogeochemically and trophically important. The objective of this study was to determine how increasing duration and severity of CO2 treatment influence the physiology of the thecosome Limacina retroversa, integrating both gene expression and organism-level (respiration and calcification) metrics. We exposed pteropods to over-saturated, near-saturated or under-saturated conditions and sampled individuals at 1, 3, 7, 14 and 21 days of exposure to test for the effect of duration. We found that calcification was affected by borderline and under-saturated conditions by week two, while respiration appeared to be more strongly influenced by an interaction between severity and duration of exposure, showing complex changes by one week of exposure. The organismal metrics were corroborated by specific gene expression responses, with increased expression of biomineralization-associated genes in the medium and high treatments throughout and complex changes in metabolic genes corresponding to both captivity and CO2 treatment. Genes associated with other physiological processes such as lipid metabolism, neural function and ion pumping had complex responses, influenced by both duration and severity. Beyond these responses, our findings detail the captivity effects for these pelagic organisms, providing information to contextualize the conclusions of previous studies, and emphasizing a need for better culturing protocols.Funding for this research was provided by a National Science Foundation grant to G.L.L., A.E.M. and A.M.T. (OCE-1316040). Additional support for field sampling was provided by theWoods Hole Oceanographic Institution, Coastal Ocean Institute and the Pickman Foundation.2019-02-1

The effect of elevated carbon dioxide on the sinking and swimming of the shelled pteropod Limacina retroversa

Author Posting. © The Author(s), 2017. This is the author's version of the work. It is posted here under a nonexclusive, irrevocable, paid-up, worldwide license granted to WHOI. It is made available for personal use, not for redistribution. The definitive version was published in ICES Journal of Marine Science 74 (2017): 1893–1905, doi:10.1093/icesjms/fsx008.Shelled pteropods are planktonic molluscs that may be affected by ocean acidification. Limacina retroversa from the Gulf of Maine were used to investigate the impact of elevated carbon dioxide (CO2) on shell condition as well as swimming and sinking behaviours. Limacina retroversa were maintained at either ambient (ca. 400 μatm) or two levels of elevated CO2 (800 and 1200 μatm) for up to four weeks, and then examined for changes in shell transparency, sinking speed, and swimming behaviour assessed through a variety of metrics (e.g., speed, path tortuosity, wing beat frequency). After exposures to elevated CO2 for as little as four days, the pteropod shells were significantly darker and more opaque in the elevated CO2 treatments. Sinking speeds were significantly slower for pteropods exposed to medium and high CO2 in comparison to the ambient treatment. Swimming behaviour showed less clear patterns of response to treatment and duration of exposure, but overall, swimming did not appear to be hindered under elevated CO2. Sinking is used by L. retroversa for predator evasion, and altered speeds and increased visibility could increase the susceptibility of pteropods to predation.Funding for this research was provided by a National Science Foundation grant to Lawson, Maas, and Tarrant (OCE-1316040). Additional support for field sampling was provided by the WHOI Coastal Ocean Institute, Pickman Foundation, and the Tom Haas Fund at the New Hampshire Charitable Foundation

Life cycle and early development of the thecosomatous pteropod Limacina retroversa in the Gulf of Maine, including the effect of elevated CO2 levels

Author Posting. © The Author(s), 2015. This is the author's version of the work. It is posted here by permission of Springer for personal use, not for redistribution. The definitive version was published in Marine Biology 162 (2015): 2235-2249, doi:10.1007/s00227-015-2754-1.Thecosome pteropods are pelagic molluscs with aragonitic shells. They are considered to be especially vulnerable among plankton to ocean acidification (OA), but to recognize changes due to anthropogenic forcing a baseline understanding of their life history is needed. In the present study, adult Limacina retroversa were collected on five cruises from multiple sites in the Gulf of Maine (between 42° 22.1’–42° 0.0’ N and 69° 42.6’–70° 15.4’ W; water depths of ca. 45–260 m) from October 2013−November 2014. They were maintained in the laboratory under continuous light at 8° C. There was evidence of year-round reproduction and an individual life span in the laboratory of 6 months. Eggs laid in captivity were observed throughout development. Hatching occurred after 3 days, the veliger stage was reached after 6−7 days, and metamorphosis to the juvenile stage was after ~ 1 month. Reproductive individuals were first observed after 3 months. Calcein staining of embryos revealed calcium storage beginning in the late gastrula stage. Staining was observed in the shell gland, shell field, mantle, and shell margin in later stages. Exposure of two batches of larvae at the gastrula stage to elevated CO2 levels (800 and 1200 ppm) resulted in significantly increased mortality in comparison with individuals raised under ambient (~400 ppm) conditions and a developmental delay in the 1200 ppm treatment compared with the ambient and 800 ppm treatments.A. Thabet is grateful for a fellowship from the Egyptian Culture and Education Bureau and for mentoring from Drs. S.A. Saber, M.M. Sarhan and M.M. Fouda. Funding for this research was provided by a National Science Foundation grant to Lawson, Maas, and Tarrant (OCE-1316040). Additional support for field sampling was provided by the WHOI Coastal Ocean Institute and Pickman Foundation to Wang, Maas, and Lawson.2016-10-2

Seasonal controls of aragonite saturation states in the Gulf of Maine

Author Posting. © American Geophysical Union, 2017. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Journal of Geophysical Research: Oceans 122 (2017): 372–389, doi:10.1002/2016JC012373.The Gulf of Maine (GoME) is a shelf region especially vulnerable to ocean acidification (OA) due to natural conditions of low pH and aragonite saturation states (Ω-Ar). This study is the first to assess the major oceanic processes controlling seasonal variability of the carbonate system and its linkages with pteropod abundance in Wilkinson Basin in the GoME. Two years of seasonal sampling cruises suggest that water-column carbonate chemistry in the region undergoes a seasonal cycle, wherein the annual cycle of stratification-overturn, primary production, respiration-remineralization and mixing all play important roles, at distinct spatiotemporal scales. Surface production was tightly coupled with remineralization in the benthic nepheloid layer during high production seasons, which results in occasional aragonite undersaturation. From spring to summer, carbonate chemistry in the surface across Wilkinson Basin reflects a transition from a production-respiration balanced system to a net autotropic system. Mean water-column Ω-Ar and abundance of large thecosomatous pteropods show some correlation, although patchiness and discrete cohort reproductive success likely also influence their abundance. Overall, photosynthesis-respiration is the primary driving force controlling Ω-Ar variability during the spring-to-summer transition as well as over the seasonal cycle. However, calcium carbonate (CaCO3) dissolution appears to occur near bottom in fall and winter when bottom water Ω-Ar is generally low but slightly above 1. This is accompanied by a decrease in pteropod abundance that is consistent with previous CaCO3 flux trap measurements. The region might experience persistent subsurface aragonite undersaturation in 30–40 years under continued ocean acidification.Coastal Ocean Institute at Woods Hole Oceanographic Institution; National Science Foundation Grant Number: (OCE-1316040)2017-07-2

Daily cycle in oxygen consumption by the sea anemone Nematostella vectensis Stephenson

© The Author(s), 2016. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Biology Open (2016): 1-4, doi:10.1242/bio.013474.In bilaterian animals, the circadian clock is intimately involved in regulating energetic metabolism. Although cnidarians exhibit diel behavioral rhythms including cycles in locomotor activity, tentacle extension and spawning, daily cycles in cnidarian metabolism have not been described. To explore a possible circadian metabolic cycle, we maintained the anemone Nematostella vectensis in a 12 h light/dark cycle, a reversed light cycle, or in constant darkness. Oxygen consumption rates were measured at intervals using an optical oxygen meter. Respiration rates responded to entrainment with higher rates during light periods. During a second experiment with higher temporal resolution, respiration rates peaked late in the light period. The diel pattern could be detected after six days in constant darkness. Together, our results suggest that respiration rates in Nematostella exhibit a daily cycle that may be under circadian control and that the cycle in respiration rate is not driven by the previously described nocturnal increase in locomotor activity in this species.Funding was provided by the US–Israel Binational Science Foundation [Grant 201187]. I.T.J. was supported by the WHOI Summer Student Fellow program, which is partially funded by the National Science Foundation Research Experience for Undergraduates program. A.M.R. was supported by National Institutes of Heath [R15GM114740]