Effects of Ocean Acidification and Warming on Sperm Activity and Early Life Stages of the Mediterranean Mussel (Mytilus galloprovincialis)

Larval stages are among those most vulnerable to ocean acidification (OA). Projected atmospheric CO2 levels for the end of this century may lead to negative impacts on communities dominated by calcifying taxa with planktonic life stages. We exposed Mediterranean mussel (Mytilus galloprovincialis) sperm and early life stages to pHT levels of 8.0 (current pH) and 7.6 (2100 level) by manipulating pCO2 level (380 and 1000 ppm). Sperm activity was examined at ambient temperatures (16-17 °C) using individual males as replicates. We also assessed the effects of temperature (ambient and = 20 °C) and pH on larval size, survival, respiration and calcification of late trochophore/early D-veliger stages using a cross-factorial design. Increased pCO2 had a negative effect on the percentage of motile sperm (mean response ratio R= 71%) and sperm swimming speed (R= 74%), possibly indicating reduced fertilization capacity of sperm in low concentrations. Increased temperature had a more prominent effect on larval stages than pCO2, reducing performance (RSize = 90% and RSurvival = 70%) and increasing energy demand (RRespiration = 429%). We observed no significant interactions between pCO2 and temperature. Our results suggest that increasing temperature might have a larger impact on very early larval stages of M. galloprovincialis than OA at levels predicted for the end of the century.

Seawater carbonate chemistry and larval development and settlement of the sea urchin Arbacia lixula in a laboratory experiment

We studied the effects of temperature and pH on larval development, settlement and juvenile survival of a Mediterranean population of the sea urchin Arbacia lixula. Three temperatures (16, 17.5 and 19 °C) were tested at present pH conditions (pHT 8.1). At 19 °C, two pH levels were compared to reflect present average (pHT 8.1) and near-future average conditions (pHT 7.7, expected by 2100). Larvae were reared for 52-days to achieve the full larval development and complete the metamorphosis to the settler stage. We analyzed larval survival, growth, morphology and settlement success. We also tested the carry-over effect of acidification on juvenile survival after 3 days. Our results showed that larval survival and size significantly increased with temperature. Acidification resulted in higher survival rates and developmental delay. Larval morphology was significantly altered by low temperatures, which led to narrower larvae with relatively shorter skeletal rods, but larval morphology was only marginally affected by acidification. No carry-over effects between larvae and juveniles were detected in early settler survival, though settlers from larvae reared at pH 7.7 were significantly smaller than their counterparts developed at pH 8.1. These results suggest an overall positive effect of environmental parameters related to global change on the reproduction of A. lixula, and reinforce the concerns about the increasing negative impact on shallow Mediterranean ecosystems of this post-glacial colonizer.

Acid-base regulation in the Dungeness crab (Metacarcinus magister)

Homeostatic regulation allows organisms to secure basic physiological processes in a varying environment. To counteract fluctuations in ambient carbonate system speciation due to elevated seawater pCO2 (hypercapnia), many aquatic crustaceans excrete/accumulate acid-base equivalents through their gills; however, not much is known about the role of ammonia in this response. The present study investigated the effects of hypercapnia on acid-base and ammonia regulation in the Dungeness crab, Metacarcinus magister on the whole animal and isolated gill levels. Hemolymph pCO2 and [HCO3]- increased in M. magister acclimated to elevated pCO2 (330 Pa), while pH remained stable. Additionally, hemolymph [Na+], [Ca2+], and [SO4]2- were significantly increased. When challenged with varying pH during gill perfusion, the pH of the artificial hemolymph remained relatively unchanged. Overall, ammonia production and excretion, as well as oxygen consumption, were reduced in crabs acclimated to elevated pCO2, demonstrating that either (amino acid) oxidation is reduced in response to this particular stress, or nitrogenous wastes are excreted in an alternative form.

Photophysiological responses of Southern Ocean phytoplankton to changes in CO2 concentrations: Short-term versus acclimation effects

The present study examines how different pCO2 acclimations affect the CO2- and light-dependence of photophysiological processes and O2 fluxes in four Southern Ocean (SO) key phytoplankton species. We grew Chaetoceros debilis (Cleve), Pseudo-nitzschia subcurvata (Hasle), Fragilariopsis kerguelensis (O'Meara) and Phaeocystis antarctica (Karsten) under low (160 µatm) and high (1000 ?atm) pCO2. The CO2- and light-dependence of fluorescence parameters of photosystem II (PSII) were determined by means of a fluorescence induction relaxation system (FIRe). In all tested species, nonphotochemical quenching (NPQ) is the primary photoprotection strategy in response to short-term exposure to high light or low CO2 concentrations. In C. debilis and P. subcurvata, PSII connectivity (p) and functional absorption cross-sections of PSII in ambient light (sigma PSII') also contributed to photoprotection while changes in re-oxidation times of Qa acceptor (tQa) were more significant in F. kerguelensis. The latter was also the only species being responsive to high acclimation pCO2, as these cells had enhanced relative electron transport rates (rETRs) and sigma PSII' while tQa and p were reduced under short-term exposure to high irradiance. Low CO2-acclimated cells of F. kerguelensis and all pCO2 acclimations of C. debilis and P. subcurvata showed dynamic photoinhibition with increasing irradiance. To test for the role and presence of the Mehler reaction in C. debilis and P. subcurvata, the light-dependence of O2 fluxes was estimated using membrane inlet mass spectrometry (MIMS). Our results show that the Mehler reaction is absent in both species under the tested conditions. We also observed that dark respiration was strongly reduced under high pCO2 in C. debilis while it remained unaltered in P. subcurvata. Our study revealed species-specific differences in the photophysiological responses to pCO2, both on the acclimation as well as the short-term level.

Impacts of elevated CO2 on particulate and dissolved organic matter production: microcosm experiments using iron-deficient plankton communities in open subarctic waters

Response of phytoplankton to increasing CO2 in seawater in terms of physiology and ecology is key to predicting changes in marine ecosystems. However, responses of natural plankton communities especially in the open ocean to higher CO2 levels have not been fully examined. We conducted CO2 manipulation experiments in the Bering Sea and the central subarctic Pacific, known as high nutrient and low chlorophyll regions, in summer 2007 to investigate the response of organic matter production in iron-deficient plankton communities to CO2 increases. During the 14-day incubations of surface waters with natural plankton assemblages in microcosms under multiple pCO2 levels, the dynamics of particulate organic carbon (POC) and nitrogen (PN), and dissolved organic carbon (DOC) and phosphorus (DOP) were examined with the plankton community compositions. In the Bering site, net production of POC, PN, and DOP relative to net chlorophyll-a production decreased with increasing pCO2. While net produced POC:PN did not show any CO2-related variations, net produced DOC:DOP increased with increasing pCO2. On the other hand, no apparent trends for these parameters were observed in the Pacific site. The contrasting results observed were probably due to the different plankton community compositions between the two sites, with plankton biomass dominated by large-sized diatoms in the Bering Sea versus ultra-eukaryotes in the Pacific Ocean. We conclude that the quantity and quality of the production of particulate and dissolved organic matter may be altered under future elevated CO2 environments in some iron-deficient ecosystems, while the impacts may be negligible in some systems.

European sea bass, Dicentrarchus labrax, in a changing ocean

Ocean acidification, caused by rising concentrations of carbon dioxide (CO2), is widely considered to be a major global threat to marine ecosystems. To investigate the potential effects of ocean acidification on the early life stages of a commercially important fish species, European sea bass (Dicentrarchus labrax), 12 000 larvae were incubated from hatch through metamorphosis under a matrix of two temperatures (17 and 19 °C) and two seawater pCO2 levels (ambient and 1,000 µatm) and sampled regularly for 42 days. Calculated daily mortality was significantly affected by both temperature and pCO2, with both increased temperature and elevated pCO2 associated with lower daily mortality and a significant interaction between these two factors. There was no significant pCO2 effect noted on larval morphology during this period but larvae raised at 19 °C possessed significantly larger eyes and lower carbon:nitrogen ratios at the end of the study compared to those raised under 17 °C. Similarly, when the incubation was continued to post-metamorphic (juvenile) animals (day 67-69), fish raised under a combination of 19 °C and 1000 µatm pCO2 were significantly heavier. However, juvenile D. labrax raised under this combination of 19 °C and 1000 µatm pCO2 also exhibited lower aerobic scopes than those incubated at 19 °C and ambient pCO2. Most studies investigating the effects of near-future oceanic conditions on the early life stages of marine fish have used incubations of relatively short durations and suggested that these animals are resilient to ocean acidification. Whilst the increased survival and growth observed in this study supports this view, we conclude that more work is required to investigate whether the differences in juvenile physiology observed in this study manifest as negative impacts in adult fish.

Responses of the metabolism of the larvae of Pocillopora damicornis to ocean acidification and warming

Ocean acidification and warming are expected to threaten the persistence of tropical coral reef ecosystems. As coral reefs face multiple stressors, the distribution and abundance of corals will depend on the successful dispersal and settlement of coral larvae under changing environmental conditions. To explore this scenario, we used metabolic rate, at holobiont and molecular levels, as an index for assessing the physiological plasticity of Pocillopora damicornis larvae from this site to conditions of ocean acidity and warming. Larvae were incubated for 6 hours in seawater containing combinations of CO2 concentration (450 and 950 µatm) and temperature (28 and 30°C). Rates of larval oxygen consumption were higher at elevated temperatures. In contrast, high CO2 levels elicited depressed metabolic rates, especially for larvae released later in the spawning period. Rates of citrate synthase, a rate-limiting enzyme in aerobic metabolism, suggested a biochemical limit for increasing oxidative capacity in coral larvae in a warming, acidifying ocean. Biological responses were also compared between larvae released from adult colonies on the same day (cohorts). The metabolic physiology of Pocillopora damicornis larvae varied significantly by day of release. Additionally, we used environmental data collected on a reef in Moorea, French Polynesia to provide information about what adult corals and larvae may currently experience in the field. An autonomous pH sensor provided a continuous time series of pH on the natal fringing reef. In February/March, 2011, pH values averaged 8.075±0.023. Our results suggest that without adaptation or acclimatization, only a portion of naïve Pocillopora damicornis larvae may have suitable metabolic phenotypes for maintaining function and fitness in an end-of-the century ocean.

Effects of ultraviolet radiation and CO2 increase on winter phytoplankton assemblages in a temperate coastal lagoon

Increases in ultraviolet radiation (UVR) and CO2 affect phytoplankton growth and mortality in a variety of different ways. However, in situ responses of natural phytoplankton communities to climate change, as well as its effects on phytoplankton annual cycles, are still largely unknown. Although temperature and UVR have been increasing in temperate latitudes during winter, this season is still particularly neglected in climate change studies, being considered a non-active season regarding phytoplankton growth and production. Additionally, coastal lagoons are highly productive ecosystems and very vulnerable to climate change. This study aims, therefore, to evaluate the short-term effects of increased UVR and CO2 on the composition and growth of winter phytoplankton assemblages in a temperate coastal lagoon. During winter 2012, microcosm experiments were used to evaluate the isolated and combined effects of UVR and CO2, under ambient and high CO2 treatments, exposed to ambient UV levels and photosynthetically active radiation (PAR), or to PAR only. Phytoplankton composition, abundance, biomass and photosynthetic parameters were evaluated during the experiments. Significant changes were observed in the growth of specific phytoplankton groups, leading to changes in community composition. The cyanobacterium Synechococcus was dominant at the beginning of the experiment, but it was negatively affected by UVR and CO2. Diatoms clearly benefited from high CO2 and UVR, particularly Thalassiosira. Despite the changes observed in specific phytoplankton groups, growth and production of the whole phytoplankton community did not show significant responses to UVR and/or CO2.

Trichodesmium's strategies to alleviate phosphorus limitation in the future acidified oceans

Global warming may exacerbate inorganic nutrient limitation, including phosphorus (P), in the surface-waters of tropical oceans that are home to extensive blooms of the marine diazotrophic cyanobacterium, Trichodesmium. We examined the combined effects of P limitation and pCO2, forecast under ocean acidification scenarios, on Trichodesmium erythraeum IMS101 cultures. We measured nitrogen acquisition, glutamine synthetase activity, C uptake rates, intracellular Adenosine Triphosphate (ATP) concentration and the pool sizes of related key proteins. Here, we present data supporting the idea that cellular energy re-allocation enables the higher growth and N2 fixation rates detected in Trichodesmium cultured under high pCO2. This is reflected in altered protein abundance and metabolic pools. Also modified are particulate organic carbon and nitrogen production rates, enzymatic activities, and cellular ATP concentrations. We suggest that adjusting these cellular pathways to changing environmental conditions enables Trichodesmium to compensate for low P availability and to thrive in acidified oceans. Moreover, elevated pCO2 could provide Trichodesmium with a competitive dominance that would extend its niche, particularly in P-limited regions of the tropical and subtropical oceans.

The effects of thermal and high-CO2 stresses on the metabolism and surrounding microenvironment of the coral Galaxea fascicularis

The effects of elevated temperature and high pCO2 on the metabolism of Galaxea fascicularis were studied with oxygen and pH microsensors. Photosynthesis and respiration rates were evaluated from the oxygen fluxes from and to the coral polyps. High-temperature alone lowered both photosynthetic and respiration rates. High pCO2 alone did not significantly affect either photosynthesis or respiration rates. Under a combination of high-temperature and high-CO2, the photosynthetic rate increased to values close to those of the controls. The same pH in the diffusion boundary layer was observed under light in both (400 and 750 ppm) CO2 treatments, but decreased significantly in the dark as a result of increased CO2. The ATP contents decreased with increasing temperature. The effects of temperature on the metabolism of corals were stronger than the effects of increased CO2. The effects of acidification were minimal without combined temperature stress. However, acidification combined with higher temperature may affect coral metabolism due to the amplification of diel variations in the microenvironment surrounding the coral and the decrease in ATP contents.

Ocean acidification rapidly reduces dinitrogen fixation associated with the hermatypic coral Seriatopora hystrix

Since productivity and growth of coral-associated dinoflagellate algae is nitrogen (N)-limited, dinitrogen (N2) fixation by coral-associated microbes is likely crucial for maintaining the coral-dinoflagellate symbiosis. It is thus essential to understand the effects future climate change will have on N2 fixation by the coral holobiont. This laboratory study is the first to investigate short-term effects of ocean acidification on N2 fixation activity associated with the tropical, hermatypic coral Seriatopora hystrix using the acetylene reduction assay in combination with calcification measurements. Findings reveal that simulated ocean acidification ( pCO2 1080 µatm) caused a rapid and significant decrease (53%) in N2 fixation rates associated with S. hystrix compared to the present day scenario ( pCO2 486 µatm). In addition, N2 fixation associated with the coral holobiont showed a positive exponential relationship with its calcification rates. This suggests that even small declines in calcification rates of hermatypic corals under high CO2 conditions may result in decreased N2 fixation activity, since these 2 processes may compete for energy in the coral holobiont. Ultimately, an intensified N limitation in combination with a decline in skeletal growth may trigger a negative feedback loop on coral productivity exacerbating the negative long-term effects of ocean acidification.

Shell condition and survival of Puget Sound pteropods are impaired by ocean acidification conditions

We tested whether the thecosome pteropod Limacina helicina from Puget Sound, an urbanized estuary in the northwest continental US, experiences shell dissolution and altered mortality rates when exposed to the high CO2, low aragonite saturation state (Omega a) conditions that occur in Puget Sound and the northeast Pacific Ocean. Five, week-long experiments were conducted in which we incubated pteropods collected from Puget Sound in four carbon chemistry conditions: current summer surface (460-500 µatm CO2, Omega a=1.59), current deep water or surface conditions during upwelling (760 and 1600-1700 µatm CO2, Omega a=1.17 and 0.56), and future deep water or surface conditions during upwelling (2800-3400 µatm CO2, Omega a=0.28). We measured shell condition using a scoring regime of five shell characteristics that capture different aspects of shell dissolution. We characterized carbon chemistry conditions in statistical analyses with Omega a, and conducted analyses considering Omega a both as a continuous dataset and as discrete treatments. Shell dissolution increased linearly as aragonite saturation state decreased. Discrete treatment comparisons indicate that shell dissolution was greater in undersaturated treatments compared to oversaturated treatments. Survival increased linearly with aragonite saturation state, though discrete treatment comparisons indicated that survival was similar in all but the lowest saturation state treatment. These results indicate that, under starvation conditions, pteropod survival may not be greatly affected by current and expected near-future aragonite saturation state in the NE Pacific, but shell dissolution may. Given that subsurface waters in Puget Sound's main basin are undersaturated with respect to aragonite in the winter and can be undersaturated in the summer, the condition and persistence of the species in this estuary warrants further study.

Offspring sensitivity to ocean acidification changes seasonally in a coastal marine fish

Experimental assessments of species vulnerabilities to ocean acidification are rapidly increasing in number, yet the potential for short- and long-term adaptation to high CO2 by contemporary marine organisms remains poorly understood. We used a novel experimental approach that combined bi-weekly sampling of a wild, spawning fish population (Atlantic silverside Menidia menidia) with standardized offspring CO2 exposure experiments and parallel pH monitoring of a coastal ecosystem. We assessed whether offspring produced at different times of the spawning season (April to July) would be similarly susceptible to elevated (1100 µatm, pHNIST = 7.77) and high CO2 levels (2300 µatm, pHNIST = 7.47). Early in the season (April), high CO2 levels significantly (p < 0.05) reduced fish survival by 54% (2012) and 33% (2013) and reduced 1 to 10 d post-hatch growth by 17% relative to ambient conditions. However, offspring from parents collected later in the season became increasingly CO2-tolerant until, by mid-May, offspring survival was equally high at all CO2 levels. This interannually consistent plasticity coincided with the rapid annual pH decline in the species' spawning habitat (mean pH: 1 April/31 May = 8.05/7.67). It suggests that parents can condition their offspring to seasonally acidifying environments, either via changes in maternal provisioning and/or epigenetic transgenerational plasticity (TGP). TGP to increasing CO2 has been shown in the laboratory but never before in a wild population. Our novel findings of direct CO2-related survival reductions in wild fish offspring and seasonally plastic responses imply that realistic assessments of species CO2-sensitivities must control for parental environments that are seasonally variable in coastal habitats.

The potential of ocean acidifi cation on suppressing larval development in the Pacifi c oyster Crassostrea gigas and blood cockle Arca infl ata Reeve

We evaluated the effect of pH on larval development in larval Pacific oyster (Crassostrea gigas) and blood cockle ( Arca inflata Reeve). The larvae were reared at pH 8.2 (control), 7.9, 7.6, or 7.3 beginning 30 min or 24 h post fertilization. Exposure to lower pH during early embryonic development inhibited larval shell formation in both species. Compared with the control, larvae took longer to reach the D-veliger stage when reared under pH 7.6 and 7.3. Exposure to lower pH immediately after fertilization resulted in significantly delayed shell formation in the Pacific oyster larvae at pH 7.3 and blood cockle larvae at pH 7.6 and 7.3. However, when exposure was delayed until 24 h post fertilization, shell formation was only inhibited in blood cockle larvae reared at pH 7.3. Thus, the early embryonic stages were more sensitive to acidified conditions. Our results suggest that ocean acidification will have an adverse effect on embryonic development in bivalves. Although the effects appear subtle, they may accumulate and lead to subsequent issues during later larval development.