Attachment, spreading and growth of VERO cells on microcarriers for the optimization of large scale cultures

The VERO cell attachment, spreading and growth were measured as a function of the substrate and temperature used for cell cultivation, the presence of fetal calf serum (FCS) in the medium and the initial cell inoculum used for cultivation on MCs. The data show that the cell attachment kinetics were comparable at RT or 37v°C, a higher rate of cell attachment occurred to MCs and the presence of FCS inhibited the cell attachment to glass or plastic but not to MCs. The cell spreading, in general higher at 37v°C, was dependent on the presence of FCS, comparable on glass or plastic substrate and lower on MCs. The spread of VERO cells over MCs was fully dependent on the presence of FCS and decreases progressively with a delayed addition of FCS into the medium. The cell detachment by trypsin was slower from MCs and the cells recovered showed lower viability and reattachment. Better results of detachment, viability and reattachment were obtained by treatment with the trypsin at pH of 8 instead of 7. The lower was the number of cells/MC for the initial inoculum, the higher was the percent of unoccupied MCs (with 1 cell/MC we had 35.6% of unoccupied MCs), which were shown to remain uncovered during the whole period of culture. With an initial inoculum of 4, 6 and 8 VERO cells/MC, respectively 46%, 76% and 83% of the MCs were totally covered by cells after 7 days, the cultures showing at this time, respectively, 5.1 2 10⁵, 8.8 2 10⁵ and 1.8 2 10⁶ cells/ml, which represented a biomass production of respectively 8.5x, 9.7x and 15.5x. When compared to 175 cm² T-flasks, using the same amount of medium, a VERO cell culture on 2 mg/ml of MCs offers about 10 times more available surface for cell growth and allowed the obtention of 7 times more cells. The optimization procedures concerning initial steps of VERO cell cultures, such as the attachment, spreading and growth as a function of parameters like initial cell inoculum and medium supplementation are of special interest mainly due to the perspective of a large use of VERO cell cultures for human viral vaccine production.     

Elevated pCO2 affects the lactate metabolic shift in CHO cell culture processes

The shift from lactate production to consumption in CHO cell metabolism is a key event during cell culture cultivations and is connected to increased culture longevity and final product titers. However, the mechanisms controlling this metabolic shift are not yet fully understood. Variations in lactate metabolism have been mainly reported to be induced by process pH and availability of substrates like glucose and glutamine. The aim of this study was to investigate the effects of elevated pCO₂ concentrations on the lactate metabolic shift phenomena in CHO cell culture processes. In this publication, we show that at elevated pCO₂ in batch and fed‐batch cultures, the lactate metabolic shift was absent in comparison to control cultures at lower pCO₂ values. Furthermore, through metabolic flux analysis we found a link between the lactate metabolic shift and the ratio of NADH producing and regenerating intracellular pathways. This ratio was mainly affected by a reduced oxidative capacity of cultures at elevated pCO₂. The presented results are especially interesting for large‐scale and perfusion processes where increased pCO₂ concentrations are likely to occur. Our results suggest, that so far unexplained metabolic changes may be connected to increased pCO₂ accumulation in larger scale fermentations. Finally, we propose several mechanisms through which increased pCO₂ might affect the cell metabolism and briefly discuss methods to enable the lactate metabolic shift during cell cultivations.     

Productivity gains do not compensate for reduced calcification under near‐future ocean acidification in the photosynthetic benthic foraminifer species Marginopora vertebralis

Changes in the seawater carbonate chemistry (ocean acidification) from increasing atmospheric carbon dioxide (CO₂) concentrations negatively affect many marine calcifying organisms, but may benefit primary producers under dissolved inorganic carbon (DIC) limitation. To improve predictions of the ecological effects of ocean acidification, the net gains and losses between the processes of photosynthesis and calcification need to be studied jointly on physiological and population levels. We studied productivity, respiration, and abundances of the symbiont‐bearing foraminifer species Marginopora vertebralis on natural CO₂ seeps in Papua New Guinea and conducted additional studies on production and calcification on the Great Barrier Reef (GBR) using artificially enhanced pCO₂. Net oxygen production increased up to 90% with increasing pCO₂; temperature, light, and pH together explaining 61% of the variance in production. Production increased with increasing light and increasing pCO₂ and declined at higher temperatures. Respiration was also significantly elevated (~25%), whereas calcification was reduced (16–39%) at low pH/high pCO₂ compared to present‐day conditions. In the field, M. vertebralis was absent at three CO₂ seep sites at pH_Total levels below ~7.9 (pCO₂ ~700 μatm), but it was found in densities of over 1000 m^?2 at all three control sites. The study showed that endosymbiotic algae in foraminifera benefit from increased DIC availability and may be naturally carbon limited. The observed reduction in calcification may have been caused either by increased energy demands for proton pumping (measured as elevated rates of respiration) or by stronger competition for DIC from the more productive symbionts. The net outcome of these two competing processes is that M. vertebralis cannot maintain populations under pCO₂ exceeding 700 μatm, thus are likely to be extinct in the next century.     

The Response of Antarctic Sea Ice Algae to Changes in pH and CO2

Ocean acidification substantially alters ocean carbon chemistry and hence pH but the effects on sea ice formation and the CO₂ concentration in the enclosed brine channels are unknown. Microbial communities inhabiting sea ice ecosystems currently contribute 10–50% of the annual primary production of polar seas, supporting overwintering zooplankton species, especially Antarctic krill, and seeding spring phytoplankton blooms. Ocean acidification is occurring in all surface waters but the strongest effects will be experienced in polar ecosystems with significant effects on all trophic levels. Brine algae collected from McMurdo Sound (Antarctica) sea ice was incubated in situ under various carbonate chemistry conditions. The carbon chemistry was manipulated with acid, bicarbonate and bases to produce a pCO₂ and pH range from 238 to 6066 µatm and 7.19 to 8.66, respectively. Elevated pCO₂ positively affected the growth rate of the brine algal community, dominated by the unique ice dinoflagellate, Polarella glacialis. Growth rates were significantly reduced when pH dropped below 7.6. However, when the pH was held constant and the pCO₂ increased, growth rates of the brine algae increased by more than 20% and showed no decline at pCO₂ values more than five times current ambient levels. We suggest that projected increases in seawater pCO₂, associated with OA, will not adversely impact brine algal communities.     

A micro pCO2 electrode

By utilizing a previously developed micro pH glass electrode it has been possible to make a micro pCO₂ electrode with a tip diameter of about 10 μm. This was accomplished by placing the micro pH electrode in a conical tube containing a weak NaHCO₃ solution. The tip of the conical tube was closed with Teflon^® oil wax mixture. This closure prevented the flow of solution, but allowed CO₂ to pass into the NaHCO₃ solution thus altering the pH of this solution. Changes in pH were seen and measured by the micro pH electrode and could be related to the pCO₂ of gas or solution in which the total electrode system was placed. This electrode, principally because of its small size, has many possible applications in biological research.     

A continuous-flow culture system for organ culture of large explants of adult tissue: Effect of oxygen tension on mouse molar periodontium

Summary A modified continuous-flow culture system (CFCS) was developed to maintain large explants of periodontium from adult mouse in organ culture. The culture medium was stored in a reservoir outside of the incubator, pumped via polyvinyl tubing into small glass culture chambers that were placed in the oxygenator and then collected in a waste flask. Medium was analyzed for pO₂, pCO₂ and pH during the culture period. Three-molar and singlemolar explants of periodontium were maintained for 48 hr in the CFCS at two different pO₂ ranges: 100 to 120 mm Hg and 400 to 420 mm Hg. [³H]Proline was added 24 hr prior to sacrifice. Light-microscope morphological and radioautographic observations suggested that cell viability and incorporation of [³H]proline, probably into newly synthesized protein, increased with an increase in pO₂ and was related to a pO₂ gradient extending from the periphery to the center of the explants.     

The effects of pCO2 on yeast growth and metabolism under continuous fermentation

Summary The effects of pCO₂ were investigated by changing the aeration rate, the purging gas and the total pressure in a chemostat cultivatioa Under glucose supply limitation, an increase in pCO₂ from 44 kPa to 195 kPa resulted in 25 % decrease in cell concentration, 8 % increase in ethanol concentration, and 50 % decrease in glycerol concentration. Under oxygen supply limitation, similar dependency of ethanol and glycerol on pCO₂ was observed, however, no influence of pCO₂ on the cell yield was observed. The change in ethanol yield by pCO₂ appeared to be caused by the equilibrium shift of pyruvate dehydrogenase system.On leave from Kasetsart Univ., Bangkok 10903, ThailandOn leave from JGC Corporation, Bessho 1-14-1, Minami-Ku, Yokohama, Kanagawa, 232 Japan     

Natural variation and the capacity to adapt to ocean acidification in the keystone sea urchin Strongylocentrotus purpuratus

A rapidly growing body of literature documents the potential negative effects of CO₂‐driven ocean acidification (OA) on marine organisms. However, nearly all this work has focused on the effects of future conditions on modern populations, neglecting the role of adaptation. Rapid evolution can alter demographic responses to environmental change, ultimately affecting the likelihood of population persistence, but the capacity for adaptation will differ among populations and species. Here, we measure the capacity of the ecologically important purple sea urchin Strongylocentrotus purpuratus to adapt to OA, using a breeding experiment to estimate additive genetic variance for larval size (an important component of fitness) under future high‐pCO₂/low‐pH conditions. Although larvae reared under future conditions were smaller than those reared under present‐day conditions, we show that there is also abundant genetic variation for body size under elevated pCO₂, indicating that this trait can evolve. The observed heritability of size was 0.40 ± 0.32 (95% CI) under low pCO₂, and 0.50 ± 0.30 under high‐pCO₂ conditions. Accounting for the observed genetic variation in models of future larval size and demographic rates substantially alters projections of performance for this species in the future ocean. Importantly, our model shows that after incorporating the effects of adaptation, the OA‐driven decrease in population growth rate is up to 50% smaller, than that predicted by the ‘no‐adaptation’ scenario. Adults used in the experiment were collected from two sites on the coast of the Northeast Pacific that are characterized by different pH regimes, as measured by autonomous sensors. Comparing results between sites, we also found subtle differences in larval size under high‐pCO₂ rearing conditions, consistent with local adaptation to carbonate chemistry in the field. These results suggest that spatially varying selection may help to maintain genetic variation necessary for adaptation to future OA.     

The O2, pH and Ca2+ Microenvironment of Benthic Foraminifera in a High CO2 World

Ocean acidification (OA) can have adverse effects on marine calcifiers. Yet, phototrophic marine calcifiers elevate their external oxygen and pH microenvironment in daylight, through the uptake of dissolved inorganic carbon (DIC) by photosynthesis. We studied to which extent pH elevation within their microenvironments in daylight can counteract ambient seawater pH reductions, i.e. OA conditions. We measured the O₂ and pH microenvironment of four photosymbiotic and two symbiont-free benthic tropical foraminiferal species at three different OA treatments (∼432, 1141 and 2151 µatm pCO₂). The O₂ concentration difference between the seawater and the test surface (ΔO₂) was taken as a measure for the photosynthetic rate. Our results showed that O₂ and pH levels were significantly higher on photosymbiotic foraminiferal surfaces in light than in dark conditions, and than on surfaces of symbiont-free foraminifera. Rates of photosynthesis at saturated light conditions did not change significantly between OA treatments (except in individuals that exhibited symbiont loss, i.e. bleaching, at elevated pCO₂). The pH at the cell surface decreased during incubations at elevated pCO₂, also during light incubations. Photosynthesis increased the surface pH but this increase was insufficient to compensate for ambient seawater pH decreases. We thus conclude that photosynthesis does only partly protect symbiont bearing foraminifera against OA.     

Seasonal changes of pCO2 over a subantarctic Macrocystis kelp bed

The partial pressure of carbon dioxide (pCO₂), calculated from pH and total alkalinity measurements, was monitored together with chlorophyll a and bacterioplankton biomass in shallow coastal water located inside and outside a giant kelp bed (Macrocystis pyrifera) situated in the Kerguelen Archipelago, Southern Ocean. In spite of large changes over a short time-scale, pCO₂ variations over the year are large and exhibit a seasonal pattern in which the different stages of the annual biological turnover are well marked. The overall pattern of pCO₂ variations is related to biological activity (development of both photosynthesis and respiration) during almost the whole year. However, physical and thermodynamical constraints exert a strong influence on pCO₂ at meso time-scale (10 days) and/or when biological activity is weak. Macrocystis acts to maintain pCO₂ below saturation almost the whole year and large undersaturations (pCO₂ as low as 20 μatm) were observed within the kelp bed. Furthermore, primary production of Macrocystis covers a period of 8 ∼ 9 months a year from winter to late summer and the kelp bed seems to favour the spring phytoplanktonic bloom. The buffer factor β indicates that, outside the kelp bed, inorganic carbon dynamics are mainly influenced by air-sea exchange and photosynthesis without calcification. Inside the kelp bed, β suggests calcification by the epiphytic community. Accepted: 1 April 2000     

Ultra-low carbon dioxide partial pressure improves the galactosylation of a monoclonal antibody produced in Chinese hamster ovary cells in a bioreactor

Objective

To explore the influence of ultra-low carbon dioxide partial pressure (pCO₂) on the monoclonal antibody (mAb) N-glycosylation profile in Chinese hamster ovary (CHO) cell culture.

Results

In fed-batch bioreactor cultures, lowering the pCO₂ in the medium (<?25 mmHg) via increasing headspace aeration decreased the cell viability and mAb production in CHO cells. Additionally, mAb galactosylation under low pCO₂ was approximately 27.45?±?2.13%, noticeably higher than that observed under normal pCO₂ (21.36?±?1.66%) at harvest. However, all of the relevant intracellular nucleotide sugar concentrations were dramatically decreased to approximately 50% of the levels found under normal pCO₂ on day 7. Real-time PCR revealed that the upregulation of galactosylation-related glycosyltransferase genes and substrate transporter genes played a critical role in the improved galactosylation under the ultra-low pCO₂ condition.

Conclusions

In the bioreactor culture processes, ultra-low pCO₂ demonstrated a positive effect on mAb galactosylation.

     

High CO2 levels impair alveolar epithelial function independently of pH

Background

In patients with acute respiratory failure, gas exchange is impaired due to the accumulation of fluid in the lung airspaces. This life-threatening syndrome is treated with mechanical ventilation, which is adjusted to maintain gas exchange, but can be associated with the accumulation of carbon dioxide in the lung. Carbon dioxide (CO₂) is a by-product of cellular energy utilization and its elimination is affected via alveolar epithelial cells. Signaling pathways sensitive to changes in CO₂ levels were described in plants and neuronal mammalian cells. However, it has not been fully elucidated whether non-neuronal cells sense and respond to CO₂. The Na,K-ATPase consumes ∼40% of the cellular metabolism to maintain cell homeostasis. Our study examines the effects of increased pCO₂ on the epithelial Na,K-ATPase a major contributor to alveolar fluid reabsorption which is a marker of alveolar epithelial function.

Principal Findings

We found that short-term increases in pCO₂ impaired alveolar fluid reabsorption in rats. Also, we provide evidence that non-excitable, alveolar epithelial cells sense and respond to high levels of CO₂, independently of extracellular and intracellular pH, by inhibiting Na,K-ATPase function, via activation of PKCζ which phosphorylates the Na,K-ATPase, causing it to endocytose from the plasma membrane into intracellular pools.

Conclusions

Our data suggest that alveolar epithelial cells, through which CO₂ is eliminated in mammals, are highly sensitive to hypercapnia. Elevated CO₂ levels impair alveolar epithelial function, independently of pH, which is relevant in patients with lung diseases and altered alveolar gas exchange.     

Rising CO2 Interacts with Growth Light and Growth Rate to Alter Photosystem II Photoinactivation of the Coastal Diatom Thalassiosira pseudonana

We studied the interactive effects of pCO₂ and growth light on the coastal marine diatom Thalassiosira pseudonana CCMP 1335 growing under ambient and expected end-of-the-century pCO₂ (750 ppmv), and a range of growth light from 30 to 380 µmol photons·m⁻²·s⁻¹. Elevated pCO₂ significantly stimulated the growth of T. pseudonana under sub-saturating growth light, but not under saturating to super-saturating growth light. Under ambient pCO₂ susceptibility to photoinactivation of photosystem II (σ_i) increased with increasing growth rate, but cells growing under elevated pCO₂ showed no dependence between growth rate and σ_i, so under high growth light cells under elevated pCO₂ were less susceptible to photoinactivation of photosystem II, and thus incurred a lower running cost to maintain photosystem II function. Growth light altered the contents of RbcL (RUBISCO) and PsaC (PSI) protein subunits, and the ratios among the subunits, but there were only limited effects on these and other protein pools between cells grown under ambient and elevated pCO₂.     

Effects of carbon dioxide and methane on methanogenesis

Summary A mixed microbial culture, enriched from sewage sludge, was cultivated on a glucose medium in a high turbulence fermentor at constant temperature, pH and loading rate. Effects of high pCO₂ (0.95 atm) and pCH₄ (0.90 atm) were compared to those of low pCO₂ (0.05 atm) and pCH₄ (0.05 atm). Generally, rapid in creases in pCO₂ resulted in rapid decreases in methane production and, conversely rapid decreases in pCO₂ by N₂-sparging resulted in rapid increases in methane production. Decreased values of methane production were also accompanied by acetic acid accumulation. No inhibiting effects of high pCH₄ were detected. A method to obtain low pCO₂ by gas recycling combined with CO₂-absorption is proposed.     

Adhesion of murine cells to glass microbeads: substrate-adsorbed serum proteins.

Small (1 mm diameter) glass beads treated with acid and alkali provided a satisfactory substratum for the attachment and growth of normal and SV-40 transformed Balb/c 3T3 murine cells. Cells attached to the beads with similar, though slower kinetics as to flat glass surfaces, and spread and grew with their usual morphology; when detached by EGTA treatment, they left behind cellular substrate-attached material (SAM) which was similar electrophoretically to that obtained from tissue culture plastic. Because of their small size and rough etched surfaces, the beads have a large surface area and high adsorptive capacity, and so are a useful tool to isolate specific serum proteins adsorbed from the culture medium that may be important for cell attachment and spreading. The adsorbed serum proteins were solubilized with SDS and analyzed by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) under reducing and non-reducing conditions. They included all reduced species adsorbed to tissue culture plastic, and only small amounts of one other major protein extracted from a “bacteriological” polystyrene surface on which cells could not grow. Profiles of unreduced samples differed considerably. The profile of adsorbed proteins varied little with tiem (5 minutes–4 days), temperature (4°–37°C), pH (5–9), presence of the protease inhibitor PMSF, or serum concentration (0.1–10%). Much of the adsorbed protein, qualitatively similar to the SDS-extracted material, could be eluted with H₂O or phosphate-buffered saline. Purified albumin and fibrinogen bound avidly to the beads; the material adsorbed from serum contained a large amount of albumin, however, little fibrinogen and no cold-insoluble globulin (as a 220 K protein) could be detected by Coomassie blue stained SDS-PAGE gels.     

Growth Attenuation with Developmental Schedule Progression in Embryos and Early Larvae of Sterechinus neumayeri Raised under Elevated CO2

The Southern Ocean, a region that will be an ocean acidification hotspot in the near future, is home to a uniquely adapted fauna that includes a diversity of lightly-calcified invertebrates. We exposed the larvae of the echinoid Sterechinus neumayeri to environmental levels of CO₂ in McMurdo Sound (control: 410 µatm, Ω = 1.35) and mildly elevated pCO₂ levels, both near the level of the aragonite saturation horizon (510 µatm pCO₂, Ω = 1.12), and to under-saturating conditions (730 µatm, Ω = 0.82). Early embryological development was normal under these conditions with the exception of the hatching process, which was slightly delayed. Appearance of the initial calcium carbonate (CaCO₃) spicule nuclei among the primary mesenchyme cells of the gastrulae was synchronous between control and elevated pCO₂ treatments. However, by prism (7 days after the initial appearance of the spicule nucleus), elongating arm rod spicules were already significantly shorter in the highest CO₂ treatment. Unfed larvae in the 730 µatm pCO₂ treatment remained significantly smaller than unfed control larvae at days 15–30, and larvae in the 510 µatm treatment were significantly smaller at day 20. At day 30, the arm lengths were more differentiated between 730 µatm and control CO₂ treatments than were body lengths as components of total length. Arm length is the most plastic morphological aspect of the echinopluteus, and appears to exhibit the greatest response to high pCO₂/low pH/low carbonate, even in the absence of food. Thus, while the effects of elevated pCO₂ representative of near future climate scenarios are proportionally minor on these early developmental stages, the longer term effects on these long-lived invertebrates is still unknown.     

Cell metabolism and medium perfusion in VERO cell cultures on microcarriers in a bioreactor

Parameters of VERO cell growth and metabolism were studied in cultures performed on microcarriers (MCs) using a bioreactor with a working capacity of 3.7?l. Kinetic studies of VERO cell growth in batch, semi-batch and perfusion cultures using concentrations of 2 and 10?mg/ml of MCs showed that a high concentration of MCs (10?mg/ml) and the use of medium perfusion allowed the attainment of higher final yields of VERO cells (6?×?10⁶ cells/ml after 10 days of culture). Perfusion also allowed better use of MCs as indicated by the observation of about 100% of MCs totally covered by cells and the appearance of multilayered cells on 64% of MCs after 13 days of VERO cell culture with 2?mg/ml of MCs. Concerning the concentration of nutrients in the cultures, the medium perfusion was able to sustain suitable levels of galactose and glutamine, which quickly decreased after 4 days in batch cultures. The air inlet in the batch cultures was capable of eliminating the NH₄ ⁺ which accumulated in the medium culture. Lactate accumulated during the first days of culture but then was utilized by the cells and decreased along the culture time. The optimization of VERO cell cultures on microcarriers as indicated by the concentration of MCs, medium perfusion and air inlet is discussed.     

The relative availability of inorganic carbon and inorganic nitrogen influences the response of the dinoflagellate Protoceratium reticulatum to elevated CO2

This work originates from three facts: (i) changes in CO₂ availability influence metabolic processes in algal cells; (ii) Spatial and temporal variations of nitrogen availability cause repercussions on phytoplankton physiology; (iii) Growth and cell composition are dependent on the stoichiometry of nutritional resources. In this study, we assess whether the impact of rising pCO₂ is influenced by N availability, through the impact that it would have on the C/N stoichiometry, in conditions of N sufficiency. Our experiments used the dinoflagellate Protoceratium reticulatum, which we cultured under three CO₂ regimes (400, 1,000, and 5,000 ppmv, pH of 8.1) and either variable (the NO₃^? concentration was always 2.5 mmol · L^?1) or constant (NO₃^? concentration varied to maintain the same C_i/NO₃^? ratio at all pCO₂) C_i/NO₃^? ratio. Regardless of N availability, cells had higher specific growth rates, but lower cell dry weight and C and N quotas, at elevated CO₂. The carbohydrate pool size and the C/N was unaltered in all treatments. The lipid content only decreased at high pCO₂ at constant C_i/NO₃^? ratio. In the variable C_i/NO₃^? conditions, the relative abundance of Rubisco (and other proteins) also changed; this did not occur at constant C_i/NO₃^?. Thus, the biomass quality of P. reticulatum for grazers was affected by the C_i/NO₃^? ratio in the environment and not only by the pCO₂, both with respect to the size of the main organic pools and the composition of the expressed proteome.     

Stress-tolerant corals of Florida Bay are vulnerable to ocean acidification

In situ calcification measurements tested the hypothesis that corals from environments (Florida Bay, USA) that naturally experience large swings in pCO₂ and pH will be tolerant or less sensitive to ocean acidification than species from laboratory experiments with less variable carbonate chemistry. The pCO₂ in Florida Bay varies from summer to winter by several hundred ppm roughly comparable to the increase predicted by the end of the century. Rates of net photosynthesis and calcification of two stress-tolerant coral species, Siderastrea radians and Solenastrea hyades, were measured under the prevailing ambient chemical conditions and under conditions amended to simulate a pH drop of 0.1–0.2 units at bimonthly intervals over a 2-yr period. Net photosynthesis was not changed by the elevation in pCO₂ and drop in pH; however, calcification declined by 52 and 50 % per unit decrease in saturation state, respectively. These results indicate that the calcification rates of S. radians and S. hyades are just as sensitive to a reduction in saturation state as coral species that have been previously studied. In other words, stress tolerance to temperature and salinity extremes as well as regular exposure to large swings in pCO₂ and pH did not make them any less sensitive to ocean acidification. These two species likely survive in Florida Bay in part because they devote proportionately less energy to calcification than most other species and the average saturation state is elevated relative to that of nearby offshore water due to high rates of primary production by seagrasses.     

Extreme Variations of pCO2 and pH in a Macrophyte Meadow of the Baltic Sea in Summer: Evidence of the Effect of Photosynthesis and Local Upwelling

The impact of ocean acidification on benthic habitats is a major preoccupation of the scientific community. However, the natural variability of pCO₂ and pH in those habitats remains understudied, especially in temperate areas. In this study we investigated temporal variations of the carbonate system in nearshore macrophyte meadows of the western Baltic Sea. These are key benthic ecosystems, providing spawning and nursery areas as well as food to numerous commercially important species. In situ pCO₂, pH (total scale), salinity and PAR irradiance were measured with a continuous recording sensor package dropped in a shallow macrophyte meadow (Eckernförde bay, western Baltic Sea) during three different weeks in July (pCO₂ and PAR only), August and September 2011.The mean (± SD) pCO₂ in July was 383±117 µatm. The mean (± SD) pCO₂ and pH_tot in August were 239±20 µatm and 8.22±0.1, respectively. The mean (± SD) pCO₂ and pH_tot in September were 1082±711 µatm and 7.83±0.40, respectively. Daily variations of pCO₂ due to photosynthesis and respiration (difference between daily maximum and minimum) were of the same order of magnitude: 281±88 µatm, 219±89 μatm and 1488±574 µatm in July, August and September respectively. The observed variations of pCO₂ were explained through a statistical model considering wind direction and speed together with PAR irradiance. At a time scale of days to weeks, local upwelling of elevated pCO₂ water masses with offshore winds drives the variation. Within days, primary production is responsible. The results demonstrate the high variability of the carbonate system in nearshore macrophyte meadows depending on meteorology and biological activities. We highlight the need to incorporate these variations in future pCO₂ scenarios and experimental designs for nearshore habitats.