Estuarine, Coastal and Shelf Science 250 (2021) 107113

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Ocean acidification increases phytobenthic carbon fixation and export in a
warm-temperate system
Shigeki Wada a, *, Sylvain Agostini a, Ben P. Harvey a, Yuko Omori b, Jason M. Hall-Spencer a, caShimoda Marine Research Center, University of Tsukuba, Shimoda, Japan
Life and Environmental Sciences, University of Tsukuba, Tsukuba, Japanc
School of Biological and Marine Sciences, University of Plymouth, Plymouth, UKb

A R T I C L E I N F O

A B S T R A C T

Keywords:
Carbon cycle
CO2 seep
Diatoms
Macroalgae
Marine productivity

The response of photosynthetic organisms to rising CO2 levels is a key topic in ocean acidification research. Mostof the work in this field has focused on physiological responses in laboratory conditions which lack ecologicalrealism. Studies using seeps as natural analogues for ocean acidification have demonstrated shifts in algal
community composition, but the effect of CO2 on carbon fixation and export remains unclear. Here, we deployed
artificial substrata in a warm-temperate region of Japan to collect algal communities using a CO2 seep off Shikine
Island. Diatoms became dominant on settlement substrata in areas with elevated CO2 levels, whereas macroalgae
dominated at present-day levels of CO2 (reference site). This was supported by pigment composition; fucoxanthin
content, characteristic of diatoms, was higher at the high CO2 site, while more Chlorophyll b, which is char­acteristic of Chlorophyta, was found in the reference site. Algal communities that recruited in water with highlevels of CO2 had elevated rates of photosynthesis. Algal biomass was similar on all settlement panels, regardlessof CO2 concentration. Much of the carbon that was fixed by algae in the high CO2 conditions was exported, likely
due to detachment from the substratum. Diatoms that dominated under high CO2 conditions are more easilytransported away as they have no holdfast, whereas newly settled macroalgae became firmly attached at presentday levels of CO2. These results show that ocean acidification may fundamentally alter coastal carbon cycling,increasing photosynthesis and carbon export from coastal ecosystems in warm-temperate biogeographic regions
due to a shift in community composition from perennial to ephemeral algae.

1. Introduction

acidification increases the amount of dissolved inorganic carbon (DIC)
and this can stimulate the growth of certain photosynthetic organisms(Hendriks et al., 2010; Olischläger and Wiencke, 2013), if sufficientnutrients are available (Celis-Plá et al., 2015). Many algae benefit fromincreased seawater CO2 concentrations in laboratory tests (Gao et al.,
1991; Kubler et al., 1999; Liu et al., 2018) although the response isspecies-specific because some algae have carbon concentrating mecha­
nisms (CCM) adapted to present day low CO2 conditions (Badger et al.,
1998; Cornwall et al., 2017).
To assess changes to coastal carbon cycling, we need to consider boththe physiological responses of algae to ocean acidification, and thechanges in community composition. In areas where CO2 is released fromvolcanic seeps the gas dissolves into seawater providing a natural
analogue for the effects of ocean acidification (Hall-Spencer et al., 2008)
which include profound changes to benthic microalgal (Johnson et al.,2013) and macroalgal communities at both individual and population

Benthic algae are the major primary producers in shallow coastal
regions (Charpy-Roubaud and Sournia, 1990), and they have a numberof other important ecological roles such as a source of energy, carbon
storage and habitat provisioning (Smale et al., 2013; Costanza et al.,2014). Recent work has shown that rising carbon dioxide levels influ­
ence these functions (Hall-Spencer and Harvey, 2019). Uptake ofanthropogenic CO2 is causing surface ocean seawater pH to fall byaround 0.002 units per year with a 100% increase in the concentrationof H+ ions projected for the end of this century (IPCC, 2013). Ocean
acidification is a serious threat to coastal ecosystems because it reduces
the saturation state of carbonate which can make seawater corrosive tocalcifying organisms such as hard corals and shellfish (Doney et al.,2020).
As well as reducing the availability of calcium carbonate, ocean* Corresponding author. Shimoda Marine Research Center, University of Tsukuba, Shimoda, 415-0025, Japan.
E-mail address: swadasbm@shimoda.tsukuba.ac.jp (S. Wada).
https://doi.org/10.1016/j.ecss.2020.107113
Received 26 May 2020; Received in revised form 6 November 2020; Accepted 15 November 2020
Available online 3 December 20200272-7714/© 2020 Elsevier Ltd. All rights reserved.

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Estuarine, Coastal and Shelf Science 250 (2021) 107113

levels (Porzio et al., 2011, 2018, 2020). At a recently characterised
natural analogue site in Japan, calcifying algae and large
habitat-forming macroalgae get replaced by turf algae at elevated CO2
levels (Agostini et al., 2015, 2018; Harvey et al., 2019). This has been
observed at many other shallow-water CO2 seeps worldwide, although
at some seeps seagrasses grow well (Hall-Spencer et al., 2008; Connell
et al., 2018). In the present study, we assess how a CO2 gradient at aJapanese seep affects algal community composition, photosynthesis andbiomass production.
Measurements of photosynthesis at CO2 seeps allow us to better
constrain projections of the response of primary producers to ocean
acidification. In addition, comparison of photosynthesis and biomass
production furthers our understanding of carbon storage by coastal
ecosystems. Previous studies taking this approach have assessed specificalgal taxa (Hofmann et al., 2015; Vogel et al., 2015; Connell et al.,2018), but the effects of ocean acidification on benthic algal community
photosynthesis had not previously been investigated. To tackle this, weused settlement panels which are widely used to easily collect natural
benthic communities (Perkol-Finkel et al., 2006; Sokołowski et al.,2017). This technique has been used to assess shifts in benthic com­munity composition along CO2 gradients in the Mediterranean (Lidbury
et al., 2012; Johnson et al., 2013; Kroeker et al., 2013b). Moreover,settlement panels also allow the investigation of the abundance, di­versity and production of those communities of organisms that recruiton to them.
In the present study, we investigated high CO2 and present-day
conditions off Shikine Island, Japan. After 44 and 72 days wecollected algal communities that had recruited in situ, recorded differ­ences in community composition, incubated them in light conditions tomeasure net photosynthesis, and then analysed the photosynthetic pig­
ments present. Our aim was to assess how high CO2 conditions affectcoastal carbon fixation and storage.

Fig. 1. Study area off the Izu peninsula and archipelago off mainland Japan (a),Shikine Island (b) and the locations for the reference and high CO2 site used forthe deployment of settlement panels (c), land shown in green. (For interpre­
tation of the references to colour in this figure legend, the reader is referred to
the Web version of this article.)high CO2 and reference sites, respectively, corresponding to 34 panels in
the high CO2 and 42 panels in the reference sites. Subsets of the panels
were retrieved in June after 44 days and July (after 72 days). The algaeon the panels were examined using a light microscope.

2. Materials and methods
2.1. Study sites

2.3. Pigment analysis

We deployed algal settlement panels in and away from a CO2 seepthat is well characterised as an analogue for ocean acidification (Agos­
tini et al., 2015, 2018). It is located off Mikawa Bay, Shikine Island(139.2◦ N, 34.3◦ E) in Japan (Fig. 1). Two sites were used, an acidified
area near to the CO2 seep (the ‘high CO2 site’) and a nearby area (the‘reference site’) in an adjacent bay of the island. The high CO2 site
provided an end-of-the-century projection for reductions in pH (IPCC,
2013), and was not confounded by differences in temperature, salinity,
dissolved oxygen, total alkalinity, nutrients or depth relative to thereference site (Agostini et al., 2015, 2018; Harvey et al., 2019). Agostini
et al. (2018) and Harvey et al. (2018) present the environmental andcarbonate chemistry data collected during the present study. Briefly,
seawater pH in total scale (pHT) and other environmental parameters
were monitored using a combination of sensors (interval of measure­ment was 15 or 30 min, recorded from 25th May to July 5th, 2016) anddiscrete water sampling for total alkalinity. For reference and high CO2
sites, pHT was 8.137 ± 0.056 and 7.809 ± 0.093 respectively, thesalinity was 34.5 ± 0.43 and 34.1 ± 0.69, the temperature was 19.7 ±0.71 and 19.5 ± 0.82 ◦ C, the total alkalinity was 2270 ± 15.4 and 2270
± 20.3 μmol kg− 1 and the calculated pCO2 levels were 309 ± 46.4 and769 ± 225 μatm (mean ± SD).

After 44 and 72 days of deployment, 19 and 5 panels from the highCO2, and 18 and 12 panels from the reference site were retrieved (atrandom) for pigment analysis. Each settlement panel was photographedand analysed with imaging software (Adobe, Photoshop) to measure thearea size of the panels, and then stored at − 80 ◦ C for pigment analysis.
The surface of each panel was scraped clean and the settled organisms
were put into 8 ml N’N-dimethylformamide to extract photosynthetic
pigments. Canthaxanthin was added to the extracts as an internal stan­dard, and this was then filtered through a polytetrafluorethylene (PTFE)
filter with pore size of 0.45 μm (GE Healthcare Life Science, Whatman™
syringe filter, 13 mm diameter). Aliquot of extracts was injected into an
isocratic high-performance liquid chromatograph (Agilent, 1100 system
controlled by ChemStation) equipped with a C8 column (Agilent, Zorbax
Eclipse XDB-C8; 4.6 × 150 mm, 3.5 μm) and diode-array and fluorescentdetectors. The peak of each pigment was identified by comparison of
their retention time relative to eight standards; fucoxanthin, prasinox­
anthin, 19′ -hexanoloxyfucoxanthin, diadinoxanthin, alloxanthin, zeax­
anthin, Chlorophyll b and Chlorophyll a (Chl a), as described in moredetail in Hama et al. (2016).

2.2. Deployment of settlement panels

2.4. Net photosynthesis

Settlement panels were held at − 5 m depth Chart Datum within thehigh CO2 and reference sites. The panels were made of transparent PVC
measuring 5 × 2 × 0.2 cm. A total of 76 panels were attached to floats
anchored 1 m above the seafloor and haphazardly spread over a surfaceof ca. 400 m2 in each site. In April 2016, we put four and five floats in the

After 44 and 72 days of deployment, 6 and 4 panels from the highCO2, and 6 and 6 panels from the reference site were retrieved (atrandom) for the measurement of net photosynthesis (NP). Incubation
was carried out using a 250 ml acid-cleaned polycarbonate bottle, with asubset of each panel (0.8 × 2.5 cm) being used. Each bottle was filled
2

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Estuarine, Coastal and Shelf Science 250 (2021) 107113

with seawater collected from the site where the panels were retrieved,
and incubated under light (150 μmol photon s− 1 cm− 2) at 20 ◦ C for 1 h tomeasure rates of NP. Dissolved oxygen (DO) concentrations and pH in
NBS scale (pHNBS) were measured at the start and end of the incubation
period with an optode DO meter (WTW, FD0925) and a multi-sensor(ThermoFisher Scientific, Orion StarTM, equipped with a ROSS pH
electrode, 8156BNUWP). Based on the change in DO concentrations(ΔDO) values, we calculated the NP rates (normalised by unit area)
using the following equations:
NParea (NP per unit area: μg O2 cm− 2 hour − 1) = ΔDO × V/A/T ×
1000.where V, A and T are volume of seawater for incubation (l), area ofthe plate (cm2), and incubation time (hour), respectively (Roth et al.,2019).
Following the incubation, each settlement panel was photographedand analysed with imaging software (Adobe, Photoshop) to measure theexact panel area size. All incubations were completed within 6 h ofcollecting the panels.
2.5. Statistical analysis

Fig. 2. Panels retrieved from high CO2 (a and b on day 44 and 72) andreference sites (c and d on day 44 and 72). Macroalgae dominated on settlement
panels at the reference site (e), diatoms dominated at the acidified site; Tri­ceratium sp. (f) and Biddulphia sp. (g).

The amount of Chl a and the rates of NParea were analysed using a
two-way linear mixed model (LMM) (R package: nlme, function: lme;
v3.1-147: Pinheiro et al., 2020) with ‘Site’ and ‘Days’ as fixed factors.
Since several panels were attached to each float, ‘float’ was treated as arandom factor. Homogeneity of variance (Levene Test) and normality(QQ plot) were assessed for Chl a and NParea. Both Chl a (following log
transformation) and NParea conformed to these assumptions. Weighted
principal component analysis (R package: aroma light, functions: wpca:
Bengtsson et al., 2010), followed by PERMANOVA (R package: vegan,function: adonis; v2.5-6: Oksanen et al., 2019), were used to assess fordifferences in the composition of accessory pigments between ‘Site’ and‘Days’. Weighting for the principal component analysis was used to
improve the robustness to uneven group sample sizes, with weighting
applied to each sample individually as 1/n, where n is the sample size ofthe respective treatment groups (combinations of ‘Site’ and ‘Days’). In
the present study, all statistical analyses were performed using the R
statistical software, version 3.5.1 (R Core Team, 2018), and visual­isations were produced using ggplot 2 (Wickham, 2016) and ggpubr(Kassambara, 2020).

Fig. 3. Mean Chlorophyll a per unit area of the panels (μg cm-2) of panels at
Reference (white) and High CO2 (grey) sites after 44 and 72 days. Error bars
show standard error (n = 18, 18, 12, 5). The statistical significance (linear
mixed model on log-transformed data) of the main effects (Day and Site) andtheir interaction is shown.

3. Results
Visual inspection of the panels revealed biofilm formation and algal
recruitment on all panels across both sites and time points. At the highCO2 site, the panel were covered with a homogenous brown biofilm at
both time points (Fig. 2a and b). Panels at the reference site werecovered with a more diverse range of algal recruits (Fig. 2c and d). After72 days, the panels were more overgrown relative to 44 days, with ju­
venile macroalgal Chlorophyta dominating panels at the reference sitewhereas the diatoms Triceratium sp. and Biddulphia sp. were dominant
on settlement panels at the high CO2 site (Fig. 2e–g).
Chl a was the main pigment and accounted for 31–68% of the total
weight of pigments per sample. The amount of Chl a per unit area of theplates deployed in the high CO2 and reference sites was 0.284 ± 0.048
μg cm− 2 and 0.285 ± 0.022 and on day 44, and increased to 1.86 ± 0.27
μg cm− 2 and 2.03 ± 0.13 and on day 72, respectively (Fig. 3). While Chl
a value significantly increased from day 44–72 (LMM, ‘Days’: F1,5 =93.49, p < 0.001), there was no significant difference between the twosites (LMM, ‘Site’ x ‘Days’, F1,5 = 4.68, p = 0.083) (Table S1). Fuco­
xanthin was the second most abundant pigment, and the ratios of Fuc/Chl a on the panels from high CO2 site were 1.01 ± 0.082 (n = 18) and
0.488 ± 0.041 (n = 5) on day 44 and 72, respectively. These values wererelatively higher than those from reference site (0.724 ± 0.036 (n = 16)
and 0.485 ± 0.070 (n = 12) on day 44 and 72, respectively).
The overall composition of the pigments differed between both the‘Site’ (PERMANOVA: F1, 47 = 27.44, p < 0.001) and ‘Days’(PERMANOVA: F1, 47 = 13.22, p < 0.001), but did not show a significant
interaction (PERMANOVA: F1, 47 = 1.92, p = 0.16) (Table S2). Principal
component analysis showed that the two sites (high CO2 and reference)
were mostly separated along PC1 (accounting for 28.8% of total vari­ance; Fig. S1) with values in a negative and positive direction dis­tinguishing the high CO2 and reference sites, respectively (Fig. 4). The
pigment fucoxanthin showed a loading of − 0.55 on the PC1 andtherefore had a moderate contribution towards explaining the high CO2
site (Table 1). Chl b showed a positive loading of 0.70 and therefore hada moderate contribution towards explaining the reference site (Table 1).
The proportion of fucoxanthin among accessory pigments were gener­
ally higher at the high CO2 site compared to the reference site, whereas
the proportion of chlorophyll b was higher in the reference site, andbelow detection limit at day 44 in the High CO2 site (Table 1). The two‘Days’ (44 and 72) were mostly separated along PC2 (accounting for20.1% of total variance; Fig. S1) with values in a negative and positive
direction distinguishing the earlier (day 44) and latter (day 72) sampling
points, respectively (Fig. 4). Prasinoxanthin, diadinoxanthin, and 19′ hexanoloxyfucoxanthin had a moderate contribution (− 0.70, − 0.53 and− 0.31, respectively) towards explaining the earlier and latter sampling
points (Table 1). Of these pigments, diadinoxanthin represented be­
tween 9 and 19% of the total accessories pigments, although the other3

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Estuarine, Coastal and Shelf Science 250 (2021) 107113

Day 72 compared to Day 44 (LMM: ‘Day’, F1,5 = 34.29, p < 0.01), andalso higher at the high CO2 site relative to the reference site (LMM: ‘Site’,F1,5 = 10.41, p < 0.05) (Fig. 5, Table S3).
4. Discussion
Studies of carbon dioxide seeps worldwide have shown that
increasing levels of ocean acidification cause major shifts in coastal algal
community composition (Hall-Spencer and Harvey, 2019). Here weshow that algal settlement is affected by increasing levels of CO2. Mac­
roalgal recruits dominated settlement panels at a reference site, being
well attached by holdfasts, whereas less firmly attached diatoms domi­
nated on panels at a high CO2 site (Fig. 2). The differences in algal
communities were confirmed through photosynthetic pigment analysis.
The amount of fucoxanthins was higher in the algal communities at highCO2, these pigments are common in heterokont algae such as diatoms
and brown algae (Dring, 1998; Kuczynska et al., 2015).
Microscopy showed that diatoms, rather than brown algae, domi­
nated the early stages of settlement at high CO2, and this was confirmed
by our pigment analyses after 44 and 72 days of settlement on panels.
The pigment composition of brown algae and benthic diatom mats re­ported in previous studies show that the ratio of Fuc/Chl a in mostbrown algae (mean values among 30 species: 0.372; Colombo-Pallotta
et al., 2006; Desmond et al., 2019; Marambio et al., 2017; Seely et al.,
1972; Verma et al., 2017; Méndez et al., 2019) were lower than those ofnatural diatom mats (mean values among 4 reports: 1.168; Sundback
et al., 1996; Wulff et al., 2005, 2008; Stief et al., 2013) (Table S4).

Fig. 4. Weighted principal component analysis biplot of accessory pigments
across site (High CO2, filled circles, Reference - empty circles) and time points(44 days - left, and 72 days - right). The same PCA is used for both panels, buttime points were separated for ease of visualisation, with the loading of eachpigment indicated by vectors. The accessory pigments are fucoxanthin (Fuc),prasinoxanthin (Pra), 19′ -hexanoloxyfucoxanthin (Hex), diadinoxanthin (Dia),
alloxanthin (All), zeaxanthin (Zea) and chlorophyll b (Chl b).

two pigments (Prasinoxanthin and 19′ - hexanoloxyfucoxanthin) repre­
sented less than 5% (Table 1). The two other pigments, Alloxanthin andZeaxanthin were minor components, contributing less than 4.2% each to
the total accessory pigments (Table 1).
The pHNBS values of the seawater collected from the high CO2 and\reference sites (at 20 ◦ C) were 7.92 ± 0.003 and 8.20 ± 0.006 on day 44,
and 7.70 ± 0.009 and 8.16 ± 0.007 (mean ± SD) on day 72, respectively.
The changes in pHNBS (ΔpHNBS) under light incubation for the sample
from high CO2 and reference sites were 0.068 ± 0.007 and 0.043 ±0.008 on day 44, and 0.046 ± 0.020 and 0.052 ± 0.006 (mean ± SD) on
day 72, respectively, reflecting CO2 consumption by photosynthesis.
Since the changes in the pHNBS during the incubations were within
narrow ranges (from 0.023 to 0.081), differences in pHNBS between theseawater collected from the high CO2 and reference sites were main­tained throughout the experiment. The concentrations of DO at the startof light incubations were 8.73–9.48 mg l− 1, and changes in the incu­
bation period were 0.18–0.93 mg l− 1.
Net photosynthesis rates (NParea) of the panels from the high CO2 sitewere 16.5 ± 1.3 (mean ± SE, n = 6) and 31.7 ± 1.8 (mean ± SE, n = 4)
μg O2 cm− 2 hour − 1 on day 44 and 72, and from the reference site were
6.94 ± 0.71, (mean ± SE, n = 6) and 21.8 ± 2.6, (mean ± SE, n = 6) μg
O2 cm− 2 hour− 1 on day 44 and 72, respectively (Fig. 4). Overall, thismeant that NParea of the settlement panels was significantly greater on

Fig. 5. Net photosynthetic rates during laboratory incubation of panels from
Reference (white bars) and High CO2 (grey bars) sites and time points (44 and72 days) in seawater sampled at the same site. Error bars show standard error(n = 6, 6, 6, 4, respectively). The statistical significance (linear mixed model) of
the main effects (Day and Site) and their interaction is shown.

Table 1
PCA loadings of the different pigments (across both sites and time points), and their percentage weights (mean ± se (n)) for each site and time point.
Percentage Weight
PCA loading

44 days after

72 days afterPigment

PC1

PC2

ReferenceHigh CO2

ReferenceHigh CO2

Alloxanthin

0.36

0.21

Chlorophyll b

0.70

0.14

Diadinoxanthin

0.02− 0.53

Fucoxanthin− 0.550.06

19 -Hexanoloxyfucoxanthin

0.02− 0.31

Prasinoxanthin

0.12− 0.70

Zeaxanthin

0.24− 0.23

1.8 ± 0.5(16)7.1 ± 2
(16)
12.4 ± 0.3(16)72.8 ± 2.4(16)0.8 ± 0.6(16)
0.9 ± 0.5(16)4.2 ± 0.7
(16)0.2 ± 0.2
(18)
0±0
(18)
14.9 ± 0.3(18)84 ± 0.4
(18)
0±0
(18)
0.9 ± 0.4
(18)
0±0
(18)

1.2 ± 0.2(12)7.8 ± 1.1
(12)
11.7 ± 0.4(12)72.9 ± 1(12)5 ± 0.6(12)0.3 ± 0.3(12)1.1 ± 0.3(12)2.1 ± 0.1(5)2.9 ± 1.2
(5)
12.4 ± 0.6(5)79.3 ± 1.4(5)3.1 ± 0.3
(5)
0±0
(5)
0.2 ± 0.2
(5)

′

4

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Estuarine, Coastal and Shelf Science 250 (2021) 107113

Considering the higher ratio of Fuc/Chl a on the panels from the highCO2 site, this confirms that benthic diatoms were dominant at the highCO2 site. In contrast, there was significantly more Chl b in the algal
community that settled onto the Reference site panels (Table 1), and thisis the main accessory pigment of Chlorophyta (Dring, 1998). Studies
worldwide show that benthic diatoms seem to be capable of benefittingfrom ocean acidification/high CO2 conditions (Johnson et al., 2013;
Marques da Silva et al., 2017; Harvey et al., 2019). Deployment of set­tlement panels at this site showed that diatom recruitment was more
intense in the high CO2 site than in reference conditions.
Our study sites off Shikine island were very exposed to wave action
and are in the path of a strong ocean current (Kuroshio current), so it isworth comparing our results with those from experiments set up in more
sheltered conditions. Recruitment of algae using artificial substrata havebeen assessed at CO2 seeps off the islands of Vulcano and Ischia in Italy
which are both less exposed to strong water movement than Shikine
Island. Off Vulcano, Johnson et al. (2013) also found higher diatom
abundance at high CO2 on settlement substrata suspended in the watercolumn, but did not report on export of the biomass grown in these
conditions. Off Ischia island CO2 seeps, volcanic stone tiles wereattached to the seafloor to monitor algal succession for 2–14 months(Kroeker et al., 2012; Porzio et al., 2013). Contrary to the present study,
only a minimal biomass of diatoms was reported at high CO2. As thepanels in the present study were suspended off the seafloor, benthic
herbivores, such as gastropods, did not have access to the floats to graze.
Since grazing pressure is an important driving force in the community
composition of benthic flora (Hillebrand et al., 2000), the deployment
position of the artificial substrata (suspended or attached) could lead todifferences in successional processes. In addition to the environmental
condition and experimental set-up, the duration of the experimental
deployment represents another important factor determining algal
community succession. Our experiment examined the phase of early
succession, so there is scope in future studies to investigate the effects ofocean acidification on the long-term development of biofouling com­munities as it takes many months for these communities to mature(Kroeker et al., 2013b; Brown et al., 2018).
Net photosynthetic rates were significantly higher on the high CO2
panels than on the Reference panels (Fig. 5, Table S3), showing an in­crease in carbon fixation under ocean acidification. In the present day,CO2 concentrations in surface seawater are generally less than 15 μmol
kg− 1 (Zeebe and Wolf-Gladrow, 2009) which can limit photosynthetic
rates, through substrate limitation (Badger et al., 1998). Previous lab­
oratory and aquarium-based studies have shown that increased CO2 canenhance photosynthesis in some algae (Gao et al., 1991; Kubler et al.,
1999; Liu et al., 2018). However, a number of recent studies have foundthat increased CO2 can have no or even negative effects on photosyn­
thesis in several algal species (Peach et al., 2017; Porzio et al., 2020),
highlighting that responses to ocean acidification are species-specific.
These interspecific differences lead to shifts in community composi­tion under high CO2 conditions, which complicates the up-scaling oflaboratory studies to ecosystem-level change. In the present study, wedemonstrate that in situ photosynthesis of an algal community increased
with ocean acidification, which is directly relevant for future projectionsof primary production and carbon storage.
Although algal community productivity was enhanced in high-CO2
conditions, the Chl a content on these settlement panels was similar to
those in reference site conditions (Fig. 3) showing that algal biomass did
not increase despite enhanced photosynthesis. This mismatch between
photosynthesis and biomass suggests that a large part of the fixed carbon
was subsequently being lost in the high-CO2 conditions due to thedetachment of algae and export of carbon from the substratum. In thereference site, macroalgae were held in place by holdfasts whichstrongly attached the thalli to hard surfaces and helped them resist dragforces due to waves and currents. Diatoms, which were dominant on thepanels in the high CO2 site, have no holdfast and are only attached bysticky mucus materials exuded extracellularly (Hoagland et al., 1993).

At our seep site, benthic diatoms were so abundant that they formed
turf-like mats in areas with high CO2 (Fig. S2a and Agostini et al., 2018;
Harvey et al., 2019). Detached aggregations of these turf diatoms at thehigh CO2 site occur during the spring-summer season, where they can beseen drifting in clumps across the seabed (Figs. S2b and c). The driftingalgal aggregates are typically a mixture of diatom and drift macroalgae,with the diatom species identified on the panels (Triceratium sp. andBiddulphia sp.) found in large numbers within the aggregates (Figs. S2dand e).
Our findings support other research showing that ocean acidification
changes coastal algal communities, and we provide novel data showing
its effects on coastal carbon fixation and export. Although algae willcontinue to provide a huge amount of energy for coastal ecosystems
through high rates of photosynthesis (Mann, 1973; Graham et al., 2016),
our data show a shift from resilient, long-lived, diverse algal stands that
store large amounts of carbon, to less resilient, short-lived and lessdiverse stands that have increased rates of carbon fixation but greater
rates of carbon export. Such changes in algal communities reduce
habitat complexity and algal biomass in rocky shore habitats. Unlesshumanity is able to minimize and address the impacts of ocean acidifi­
cation we can expect that it will cause major disruption to coastal eco­systems with knock-on effects on nearshore food webs, fisheries andcarbon cycle.
Declaration of competing interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influence
the work reported in this paper.
Acknowledgements
We thank the technical staff of Shimoda Marine Research Center andMr Yasutaka Tsuchiya, for help with field sampling. We are also gratefulto Dr T. Hama for his kind advice. The species identification of diatoms
was supported by Drs K. Ishida and T. Nakayama. Dr J. Hama helpedwith HPLC analysis. The field survey was permitted by Nii-jima Fishery
Cooperatives Shikine-jima Branch. This study was supported by theEnvironment Research and Technology Development Fund (4RF-1701),
the Ministry of Education, Science, Sports and Culture, Grant-in-Aid for
Scientific Research (B) (19H04234) and Graind-in-Aid for Young Sci­
entists (B) (17K17622), and the International Educational and ResearchLaboratory Program of the University of Tsukuba.
Appendix A. Supplementary data
Supplementary data to this article can be found online at https://doi.
org/10.1016/j.ecss.2020.107113.
Author statement
Shigeki Wada: Conceptualization, Methodology, Investigation,
Writing – original draft, Sylvain Agostini: Methodology, Formal anal­ysis, Writing – original draft, Visualisation, Ben P. Harvey: Methodol­
ogy, Formal analysis, Writing – original draft, Visualisation, Yuko
Omori: Investigation, Writing – review & editing, Jason Hall-Spencer:
Methodology, Writing – review & editing
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