Electronic Supplementary Material to:
Ecological effects of ocean acidification and habitat
complexity on reef-associated macro-invertebrate
communities
K. E. Fabricius, G. De’ath, S. Noonan, S. Uthicke
Australian Institute of Marine Science, PMB 3, Townsville Qld 4810, Australia
Methods
Table S1: Survey methods. (a) Operational taxonomic units (OTUs) differentiated in the
fine-scale and echinoderm surveys, and their classification into mobile and sessile, and
calcifying and non-calcifying groups; (b) Definitions of six levels of fine-scale structural
complexity of substrata; (c) Categories of substratum types.
(a) Operational taxonomic units
Phylum
Operational taxonomic units
PlathelPlathelminthes.
minthes
Echiura
Bonellia (spoon worm).
Mollusca Bivalvia: Subclass Pteriomorphia, Order Ostreoida, Pectinia
(scallop), Ostreidae (true oyster); Order Mytiloida, Mytilidae:
differentiating Lithophaga (date mussel), Gastrochaena,
other;
Subclass Heterodonta (burrowing clam, cockle, other).
Gastropoda: the groups 'Opisthobranchia' (differentiating sea
hare, Nudibranchia) and 'Prosobranchia' (differentiating Cone,
Conch, Cowry, Turbo, Drupella, Coralliphyllia, and ‘other’).
Gastropoda: Vermetidae.
Annelida
Polychaeta: Errantia like forms (bristle worm).
Polychaeta: Serpulidae (christmas tree worm).
Polychaeta: Sabellidae (feather duster worm), Terebellidae.
Arthropod
a
Crustacea – Malacostraca – Decapoda: differentiating
Dendrobranchiata (true prawn);
Macrura: Achelata (spiny lobster, slipper lobster, rock lobster,
Moreton Bay bug), Astracidea (reef lobster); Pleocyemata:
Caridae (true shrimp), Stenopodidea (barber pole shrimp),
Anomura - Galatheidea (squat lobster, porcelain crab),
Anomura - Paguroidae (hermit crab); Brachyura (true crab).
1
Mobility
Mobile
Calcifier
No
Sessile
Predomi
nantly
sessile
No
Yes
Mobile
Yes
Sessile
Mobile
Sessile
Sessile
Yes
No
Yes
Mostly
No
Yes
Mobile
Echinodermata
Chordata:
Tunicata
Crustacea – Maxillopora (barnacle)
Fine-scale surveys (20 m2 belt transects):
Crinoidea (feather star); Ophiuroidea (brittle star);
Holothuroidea (sea cucumber); Asteroidea (seastar);
Echinoidea (sea urchin).
Echinoderm surveys (100 m2 belt transects):
Holothuroidea (sea cucumber): Pearsonothuria graeffei,
Thelenota anax, Stichopus horrens, Holothuria coluber,
Holothuria sp., Holothuria edulis, Euapta. sp.,
Asteroidea (seastar): Culcita novaeguinea, Gomphia watsoni,
Nardoa novaecaledoniae;
Acanthaster planci, Echinaster sp., Fromia indica, Fromia
sp., Linckia guildingii, Linckia laevigata, Linckia multifora;
Echinoidea (sea urchin): Eucidaris sp., Echinostrephus sp.,
Diadema savignyi, Echinothrix spp., Echinometra sp.
Ascidiacea (sea squirt).
Sessile
Mobile
Yes
Yes
Sessile
No
(b) Definitions of the six ordinal rating scales to visually estimate fine-scale structural
complexity of substrata [1].
Level
Definition
0
Flat (e.g. coralline algal pavement, upper surface of smooth massive Porites colonies)
1
Few cracks or crevices (e.g. coral rubble; massive Porites with few surface
irregularities; Fig. 1c)
2
Few medium-scale structures, massive colonies and low relief substrata, no branching,
foliose or tabulate skeletons
3
Moderate number or <10 cm crevices and branches (e.g. mixed community with
relatively low relief and few branching colonies; Fig. 1b)
4
Complex fine-scale structure (many crevices, plates and branches)
5
Extremely complex structure (the whole quadrat covered by tall thickets of living or
dead branching hard corals or the fire coral Millepora spp.; Fig. 1a)
(c) Categories within each of the three substratum types
Substratum Categories
HC
Living hard corals (branching, foliose, corymbose, tabulate, encrusting,
submassive, massive), and Millepora spp.
DS
Substrata devoid of living corals or other macro-benthos, and dominated by
coralline algae, turf algae, rubble or sand.
Other
Soft corals, sponges, calcareous macro-algae, non-calcareous macro-algae.
2
Table S2: Frequencies of the number of survey quadrats across CO2, substratum type and
complexity levels.
Substratum
HC
DS
Control
251
263
High.CO2
328
289
Substratum type = HC
Complexity
0
1
2
3
4
5
Control
0
5
52
53
64
77
High.CO2
1
97
137
39
28
26
Substratum type = DS
Complexity
0
1
2
3
4
5
Control
6
39
96
80
39
3
High.CO2
11
108
123
33
12
2
Seawater chemistry and oceanographic data
The three seep locations (Dobu, Esa’Ala, and Upa Upasina) are located along an active
tectonic fault line, representing the western-most extension of the Woodlark Rift where the
Australian and Solomon continental plates are spreading apart [2].
A total of 968 discrete seawater samples of pH were taken at the six sites from ~0.5 m above
the benthos (Table 1, Fig. S1). The data include all samples collected between 2010 and
2012, representing a range of tidal, irradiance and wave conditions to characterise the ranges
encountered by the organisms. The samples were immediately analysed for pH and
temperature with a Mettler Toledo InLab Expert Pro pH electrode and SG78 pH/temperature
meter, calibrated with NBS buffers, and using a TRIS seawater pH standard as a reference.
Subsamples were analysed for salinity with a handheld salinity probe (Hach), and 450 of
these samples were fixed with mercuric chloride and stored in PET bottles for later
determination of TA with a 855 Robotic Titrosampler (Metrohm), or for TA and dissolved
inorganic carbon (DIC) with a VINDTA 3D (Marianda). pH (presented at the total pH scale)
was derived from millivolt readings [3]. Other relevant seawater carbonate parameters
(aragonite saturation state, DIC, pCO2) were calculated from pH, TA, DIC, salinity and
temperature using the R program Seacarb v2.4.8 [4]. A CTD (Sea-Bird SBE 16Plus) with a
3
SBE 18 pH Sensor mounted was also deployed at various locations to assess the temporal and
spatial variability in pH (Fig. S2). Mean seawater temperature did not vary between sites [5].
Figure S1: Seawater chemistry at the six study sites.
4
Figure S2: Example of temporal variability in seawater pH (total scale) at a high CO2 site
(red) and control site (blue) of Upa Upasina (27th – 30th April 2012). Median seawater pH at
the high CO2 site was 7.86, and at the control site 8.00 during this observation period.
8.2
8
pH (total)
7.8
7.6
7.4
7.2
7
6.8
5
CO2 Effects
Table S3: Densities and number of taxonomic units of benthic macro-invertebrates at the
three PNG seeps (35 surveys, Fig. 2). Shown are ratios (high CO2 over control transects) of
mean densities (m-2), and number of lower operational taxonomic units (OTUs), classes and
phyla (number per transect). Also shown are lower and upper 95% confidence intervals (CI)
of the ratios, and mean values at the high CO2 and control sites. Groups that show significant
increases or declines at high CO2 (i.e., ranges of lower to upper 95% CI do not include 1.0)
are highlighted in bold. Ratios were estimable only for groups with densities >0 at all six
sites.
ratio high CO2 /
control
(lower,
upper 95% CI)
0.48
0.43
0.59
0.51
0.38
0.77
0.79
0.85
(0.28, 0.83)
(0.25, 0.74)
(0.31, 1.16)
(0.30, 0.85)
(0.17, 0.87)
(0.68, 0.87)
(0.71, 0.88)
(0.78, 0.93)
5.18
3.31
1.86
4.19
0.983
9.06
6.39
3.61
10.85
7.72
3.13
8.25
2.60
11.76
8.12
4.24
0.22
1.29
0.60
0.69
0.10
(0.08, 0.57)
(0.82, 2.04)
(0.33, 1.06)
(0.23, 2.05)
(0.03, 0.31)
0.922
1.63
1.64
0.878
0.181
4.24
1.26
2.75
1.27
1.85
Crustaceans - Crab
Crustaceans – Hermit crab
Crustaceans - Shrimp
0.08
0.49
0.24
(0.01, 0.62)
(0.22, 1.08)
(0.11, 0.56)
0.158
0.406
0.358
1.93
0.829
1.48
Gastropods
Bivalves
1.33
1.25
(0.72, 2.49)
(0.71, 2.20)
0.847
0.783
0.635
0.626
Crinoids
Ophiuroids
Holothurians
Asteroids
Echinoids
Linckia multifora
Diadema savignyi
Echinothrix spp.
0.25
0.83
1.26
1.15
2.20
2.97
3.81
9.50
(0.09, 0.69)
(0.40, 1.72)
(0.26, 6.19)
(0.50, 2.64)
(1.06, 4.57)
(1.24, 7.12)
(1.60, 9.06)
(1.08, 83.8)
0.353
0.883
0.022
0.094
0.146
0.016
0.059
0.026
1.415
1.071
0.018
0.082
0.066
0.005
0.015
0.003
Sabellids
Terebellids
0.85
0.44
(0.22, 3.34)
(0.22, 0.88)
1.331
0.231
1.564
0.529
Summary Variables:
Total Count
Mobiles
Sessiles
Calcified groups
Non-calcified groups
Nr. OTUs
Nr. Classes
Nr. Phyla
mean
high CO2
mean
control
Highest taxonomic groups:
Decapod crustaceans
Molluscs
Echinoderms
Polychaetes
Ascidians
Subordinate taxonomic groupings:
6
Combined effects of CO2, structural complexity and substratum types
Figure S3: Mean structural complexity of reef substrata at the control and high CO2 sites of
the three seep locations Dobu, Esa’Ala and Upa Upasina. Each circle represents a survey
averaging complexity data from 62–80 quadrats.
7
Table S4: Effects of CO2, complexity and substratum type on the densities and number of operational taxonomic units of macro-invertebrates
(GLMM results for the ten main responses, Fig. 4). Data based on the surveys conducted during the 2012 expedition.
Response
Total Count
Family
Mobiles
Sessiles
Calcified groups
Non-calcified groups
Neg.Binomial (1.686) Neg.Binomial (2.559) Neg.Binomial (0.257) Neg.Binomial (1.612) Neg.Binomial (0.3143)
df
F
p-value
F
p-value
F
p-value
F
p-value
F
p-value
CO2
1
3.15
0.076
20.21
<0.001
1.38
0.24
1.39
0.238
11.04
0.001
poly(Complexity, 2)
2
20.67
<0.001
73.44
<0.001
7.05
<0.001
24.01
<0.001
5.12
0.006
Substratum
1
9.89
0.002
1.43
0.232
13.52
<0.001
12.66
0.001
0.28
0.597
CO2:poly(Complexity,2)
2
2.17
0.114
1.33
0.264
1.82
0.162
1.24
0.29
0.16
0.851
CO2:Substratum
1
3.11
0.078
1.32
0.249
1.73
0.189
2.75
0.097
0.51
0.473
Response
Family
df
Nr. Classes
Decapod Crustaceans
quasipoisson
Neg.Binomial (1.0431) Neg.Binomial (0.5232) Neg.Binomial (1.3716) Neg.Binomial (0.0971)
F p-value
Molluscs
Echinoderms
Ascidians
F
p-value
F
p-value
F
p-value
F
p-value
CO2
1
2.71
0.1
18.55
<0.001
0.536
0.464
2.236
0.135
10.247
0.001
poly(Complexity, 2)
2 27.17
<0.001
41.24
<0.001
1.386
0.251
34.233
<0.001
5.9
0.003
Substratum
1
5.35
0.021
1.14
0.286
14.471
0.001
0.032
0.858
0.108
0.743
CO2:poly(Complexity,2)
2
6.88
0.001
1.89
0.152
2.915
0.055
1.29
0.276
0.618
0.539
CO2:Substratum
1
8.19
0.004
0.01
0.905
0.509
0.476
0.011
0.916
2.382
0.123
8
Table S5: Partial effects of CO2, complexity and substratum type on macro-invertebrate
communities: proportional changes in the densities and number of operational taxonomic
units per transect (OTUs, classes and phyla) at high CO2 compared to control sites
(CO2/Control), along the complexity gradient (lowest vs highest complexity), and in quadrats
devoid of live coral and other macro-fauna compared with those dominated by live coral
(DS/HC). In brackets: lower and upper 95% confidence intervals. Groups that show
significant increases or declines (i.e., with ranges of their lower and upper 95% confidence
intervals not including 1.0) are highlighted in bold.
CO2/control
complexity
(lower,
substratum (DS/HC)
(lower,
(lower,
mean
upper 95% CI)
mean
upper 95% CI)
mean
upper 95% CI)
Total Count
0.61
(0.43, 0.85)
0.37
(0.26, 0.52)
0.58
(0.50, 0.67)
Mobiles
0.58
(0.46, 0.72)
0.13
(0.09, 0.18)
0.96
(0.83, 1.10)
Sessiles
0.52
(0.29, 0.96)
2.61
(1.16, 5.84)
0.26
(0.18, 0.37)
Calcified groups
0.69
(0.50, 0.97)
0.4
(0.29, 0.56)
0.55
(0.48, 0.64)
groups
0.29
(0.18, 0.47)
0.11
(0.03, 0.36)
0.75
(0.48, 1.18)
Nr. OTUs
0.66
(0.53, 0.81)
0.35
(0.28, 0.44)
0.73
(0.67, 0.81)
Nr. Classes
0.69
(0.55, 0.86)
0.42
(0.33, 0.53)
0.72
(0.65, 0.79)
Nr. Phyla
0.66
(0.54, 0.80)
0.45
(0.36, 0.56)
0.74
(0.67, 0.82)
crustaceans
0.37
(0.24, 0.55)
0.10
(0.06, 0.17)
0.83
(0.66, 1.04)
Molluscs
1.26
(0.80, 1.99)
1.46
0.76, 2.79)
0.34
(0.26, 0.45)
Echinoderms
0.61
(0.37, 1.01)
0.11
(0.06, 0.18)
1.02
(0.82, 1.27)
Polychaetes
0.64
(0.24, 1.69)
6.64
(2.04, 21.6)
0.26
(0.15, 0.45)
Ascidians
0.13
(0.06, 0.29)
0.06
(0.01, 0.34)
0.73
(0.39, 1.37)
Gastropods
1.02
(0.59, 1.76)
0.59
(0.24, 1.44)
0.68
(0.46, 0.99)
Bivalves
1.84
(0.88, 3.83)
4.67
(1.90, 11.5)
0.18
(0.12, 0.26)
Crinoids
0.06
(0.01, 0.32)
0.04
(0.02, 0.09)
1.21
(0.89, 1.64)
Ophiuroids
0.72
(0.39, 1.31)
0.12
(0.06, 0.26)
0.59
(0.43, 0.83)
Holothurians
3.92
(0.40, 38.8)
13.0
(0.42, 401)
1.26
(0.54, 2.95)
Asteroids
4.85
(1.56, 15.1)
0.13
(0.03, 0.54)
2.03
(1.06, 3.88)
Echinoids
1.45
(0.64, 3.29)
0.47
(0.14, 1.59)
2.03
(1.19, 3.49)
Sabellids
0.66
(0.25, 1.79)
7.43
(2.26, 24.4)
0.25
(0.14, 0.44)
Terebellids
0.49
(0.29, 0.86)
0.25
(0.09, 0.74)
1.02
(0.67, 1.56)
Non-calcified
Decapod
9
References
1.
2.
3.
4.
5.
Wilson S.K., Graham N.A.J., Polunin N.V.C. 2007 Appraisal of visual assessments of
habitat complexity and benthic composition on coral reefs. Marine Biology 151,
1069-1076.
Little T.A., Baldwin S.L., Fitzgerald P.G., Monteleone B. 2007 Continental rifting
and metamorphic core complex formation ahead of the Woodlark spreading ridge,
D’Entrecasteaux Islands, Papua New Guinea. Tectonics 26. 10.1029/2005TC001911.
Dickson A.G., L. S.C., Christian J.R.E. 2007 Guide to best practices for ocean CO2
measurements. North Pacific Marine Science Organization (PICES) Special
Publication 3, 176 pp.
Lavigne H., Gattuso J.P. 2013 Package ‘seacarb’: Seawater carbonate chemistry with
R. Version 2.4.8. (ed. R_Development_Core_Team). http://cran.rproject.org/web/packages/seacarb/index.html.
Fabricius K., Langdon C., Uthicke S., Humphrey C., Noonan S., De’ath G., Okazaki
R., Muehllehner N., Glas M., Lough J. 2011 Losers and winners in coral reefs
acclimatized to elevated carbon dioxide concentrations. Nature Climate Change 1,
165–169.
10