Electronic Supplementary Material
Definitions
Clonal Structure Indices
Genotypic richness was calculated by dividing the number of unique genotypes by the
total number of samples (Ng/N) [1]. Genotypic diversity was calculated by dividing the observed
genotypic diversity over the expected (Go/Ge) [1]. This measure is equivalent to evenness
corresponding to Nei’s index (eve) as calculated in GENODIVE. Because each individual is
expected to be genetically distinct, Ge is equal to the number of colonies genotyped. Observed
genotypic diversity, Go, was calculated as:
where gi is the relative frequency of the ith of k genotypes estimated as ni (the number of samples
of genotype i found in the total number of samples) divided by N. If colonies in the population
are all sexually produced, Go will be equal to N. Therefore, genotypic diversity is maximized in a
solely sexual population (Go/Ge=1) and approaches 0 when all colonies are asexually produced.
Lastly, a measure of genotypic evenness was calculated by dividing observed genotypic diversity
by the number of unique genotypes (Go/Ng) [1]. In contrast to richness, evenness is more
influenced by genet longevity than recruitment. Evenness will approach 0 when the population is
dominated by one genotype but will approach 1 when each genotype is equally represented in
number of colonies. However, this statistic is meaningless when populations consist of only one
genotype.
Supplemental Results
Clustering Results
The first three principal coordinates explained >80% of the variation. The first principal
component captured the differentiation between the two Porites species, explaining > 60% of the
variation (Fig. 1A).
STRUCTURE HARVESTER supported K = 2 as the most likely number of clusters. Support
for K = 2 clusters was strong; adding a second cluster to the STRUCTURE analysis resulted in a
26% increase in likelihood from assuming just one cluster, while further increases in the number
of clusters resulted in <2% increases in likelihood using this marker set.
Genotyping results
Strong clustering of MLGs into two groups in the PCA and STRUCTURE analysis can be
attributed to the presence of fixed alleles at three loci and low allelic diversity in P. evermanni.
(Fig. S1). Rarefied allelic richness estimates based on a standardized sample size (n=15) resulted
in a mean number of alleles per locus per sampling location of 4.07±0.23 s.d. alleles in P. lobata
and 2.40±0.17 s.d. alleles in P. evermanni. These differences in allelic diversity between species
are likely due to ascertainment bias rather than biology because the markers were developed to
be highly variable in P. lobata [2, 3].
Species distributions
No difference was found in the number of colonies sampled per plot per region
(ANOVA, square root transformed, df = 2, F = 0.574, p = 0.5; post-hoc pairwise tests (Tukey), p
> 0.5 for all comparisons) or in the area sampled (standardized by design). The proportion of P.
evermanni differed among all regions tested (ANOVA, arcsine square root transformed, df = 2, F
= 73.383, p<0.001; post-hoc pairwise tests (Tukey), p < 0.005 for all comparisons; Fishers exact
tests, Bonferonni correction, p<0.001). Mainland Costa Rica had the highest proportion of P.
evermanni, followed by the coastal island and then the oceanic island where P. evermanni was
nearly absent (Fig. 3).
Supplementary References
1.
Stoddart J.A. 1983 A genotypic diversity measure. J. Hered. 74, 489-490.
2.
Polato N.R., Concepcion G.T., Toonen R.J., Baums I.B. 2010 Isolation by distance across the Hawaiian
Archipelago in the reef-building coral Porites lobata. Mol. Ecol. 19, 4661-4677. (doi:10.1111/j.1365294X.2010.04836.x).
3.
Baums I.B., Boulay J.N., Polato N.R., Hellberg M. 2012 No gene flow across the Eastern Pacific Barrier in
the reef-building coral Porites lobata. Mol. Ecol. 21, 5418-5433. (doi:doi: 10.1111/j.1365-294X.2012.05733.x).
Figure S1. Allele frequencies for Porites lobata (Pl) and Porites evermanni (Pe) at 11
microsatellite loci. Circles represent the presence of an allele and are scaled by the frequency of
that allele in the data set. Allele size (bp) is given on the x axis. Species are given on the y axis.
Figure S2. Denaturing gradient gel electrophoresis (DGGE) gel image. Symbiodinium ITS2
sequences from eight representative Porites evermanni (left) and eight Porites lobata samples
(right) were run on a denaturing gradient gel with a ladder and two C15 positive controls (+). All
samples show subclade C15 as their dominant symbiont as evident by the bands to the left of the
arrow. * indicates that the band to the left was excised, re-amplifed, and Sanger sequenced. All
sequences aligned with 100% identity with a published C15 ITS2 sequence (GenBank AY
239369.1).
Table S1. Porites lobata (Pl) and Porites evermanni (Pe) samples (n = 684) were obtained from
three regions (North, Central, and South) and 17 sites in the Eastern Tropical Pacific using
haphazard (H) and/or random (R) sampling methods. Sites are arranged in approximately westto-east and north-to-south order. Given is the ratio of the number of unique multilocus genotypes
by species (genets; NgPl, NgPe) over the total sample size (NPl, NPe). GPS locations are in decimal
degrees (WGS84). The number of polar plots per site is given where applicable. See Table S3 for
polar plot level information.
Region
Subregion
Site
Site Name
NgPl/NPl
NgPe/NPe
Latitude
Longitude
North
Clipperton
CL01
Clipperton
5/51
2/2
10.29989
-109.216
Sampling
Method(s)
H
Mexico
ME01
Ixtapa
0/0
3/17
17.65436
-101.6234
H
Costa Rica
CR01*
San Juanillo
Marino
Ballena
Manuel
Antonio
Caño Island
0/0
5/50
10.31117
-85.74408
H; R (n=1)
22/291
23/34
9.104583
-83.7068
H; R (n=1)
3/31
17/27
9.381867
-84.1438
H
60/791
24/43
8.71067
-83.8911
H; R (n=3)
Central
CR02*
CR03*
CR04*
CR05*
CR06*
Panama†
South
†
Galapagos
6/12
8.6713
-83.7267
29/30
1
43/61
8.727433
-83.3863
H; R (n=2)
1
1/1
5.534834
-87.0875
R (n=3)
9/11
7.8168
-81.75
H
Cocos Island
53/56
PA01*
N Panama2
13/131
PA02*
3
GA01
S Panama
Darwin
4/4
1
7/7
8.63
-79.03
H
36/40
1
4/5
1.616525
-91.9733
H
1
GA02
Wolf
35/41
0/0
1.336935
-91.806
H
GA03
Marchena
Central
Galapagos4
Espanola
24/361
0/0
0.318369
-90.4691
H
12/131
0/0
-0.72846
-90.059
H
0/0
4/10
-1.3501
-89.618
H
1/27
1.476383
-80.7937
H
GA05*
Total
Gulfo Dulce
1/8
CR07*
GA04
Ecuador
Drake Bay
1
EC01
La Llorona
2/20
1
299/377
149/307
1 Samples from Baums et al. 2012
2 Uva, Coibita
3 Contadora, Saboga
4 Santiago, Baltra, Floreana
*Note that site numbers differ from sites described in Baums et al. 2012
†
Collection does not include samples from Forsman et al. 2009
Table S2. Microsatellite loci for Porites lobata (Pl) and Porites evermanni (Pe). The primer
sequences are preceded by the name of the fluorescent dye used (6FAM, VIC, NED or PET;
Applied Biosystems, CA). The type of repeat and the size of the polymerase chain reaction
(PCR) product is given (in basepairs, bp). Loci were amplified in four multiplex and one
singleplex reaction (Plex) using the annealing temperatures (Temp) indicated.
Marker
Name
PL0072
PL0780*
PL0905†
PL1357*
PL1370†
PL1551*
PL1556*
PL1629*
PL1868†
PL2069*
PL2258*
Primer Sequence
F: NED-CACGCCTTTCTATTACGTTGA
R: CACCCTCTTACACTTCATTCATT
F: VIC-GCCAGTAGGTGGATACACTGTT
R: CAAGTACGTTGACGTCGTTG
F: NED-GGTCCAAAGTCCACCATCA
R: TGGTGGAAATAAGTGGTCGA
F:PET- ATGTCCCTGAAACGGAAGTA
R: GATGATGATGTTGTTGATGGTG
F: PET-GCACTGTCTGTAACAAGCGAA
R: CATATTGGAAGGAGGGCTC
F: PET-TGTTTCTGAGTGGCTGTGCT
R: GGTTGGAAAGGGTCCTTCAT
F: PET-CGTTGACGTAACCTTCACCA
R: CACAGGGTAACCTTCCTTGC
F: 6FAM-CCTTGGTTAATTTGCCCTTG
R: ACCAGTCCGGAGTCAAGCTA
F: VIC-TAAGCCACAGCAGGTGTACG
R: AAACGTTCCCTATCCCATCC
F: PET-CGCAGTTCCTTTGATTTGGT
R: GTTTCTTTAGCGGTTGATGGCTTGTTAC
F: NED-ATTAGCGGATGAAGCGAAGA
R: TCCAATGTAACGCCAAATCA
* Polato et al. 2010
† Baums et al. 2012
Repeat
Size (bp)
Temp
(°C)
Plex
(AACG)10
205-407
54
E
(ATT)4 (GTT)7
136-163
52
A
(ATC)9 ACC
(ATC)9
126-183
52
A
(ACC)7…(ATC)
4…(ACC)7
Pl: 252-300
Pe: 100-111
52
D
(GTT)8
189-246
54
E
(GTT)8
178-196
52
A
(ATC)10
153-168
56
B
(GCT)8
168-180
52
D
(AAC)10
Pl: 185-206
Pe: 179
52
D
(GTT)8
Pl:249-267
Pe: 264
52
C
(GAT)10
217-250
56
B
Table S3. Indices of clonal structure as calculated for each polar plot. Colony sizes were
estimated as maximum length multiplied by maximum width from measurements taken in the
field. Go = same as Ne, Ge = N. The average number of ramets (R) over the number of genets (G)
is given as well as average colony size.
Species
Site
P. evermanni
Caño Island
Marino
Ballena
San Juanillo
Gulfo Dulce
Cocos Island
N
Ng
Ng/N
Go
Go/Ge
Go/Ng
R/G
Size (m2) ±
s.d.
Caño1
12
4
0.33
1.71
0.14
0.43
3.0
0.9±1.2
Caño2
10
5
0.50
2.50
0.25
0.50
2.0
0.3±0.3
Caño5
Tres
Hermanas
Punta
Pleito
Aguja
9
4
0.44
3.00
0.33
0.75
2.25
2.9±2.5
10
4
0.40
2.38
0.24
0.60
2.5
1.3±2.6
27
5
0.19
1.48
0.05
0.30
5.4
1.1±3.9
25
17
0.68
11.79
0.47
0.69
1.47
0.3±0.5
25
19
0.76
16.03
0.64
0.84
1.32
0.3±0.3
1
1
1.00
1.00
1.00
1.00
1.0
-
Plot
Sandalo
*Punta
Ulloa
Total
118
58
0.49
38.90
0.33
0.67
2.0
0.8±2.2
Mean*
16.85
8.29
0.47
5.56
0.30
0.59
2.6
1.0
s.d.
8.32
6.68
0.20
5.86
0.20
0.19
1.3
1.4
Caño1
8
8
1.00
8.00
1.00
1.00
1.0
1.2±1.1
Caño2
10
5
0.50
3.57
0.36
0.71
2.0
0.2±0.2
Caño5
Tres
Hermanas
Punta Ulloa
Bahia
Weston
Punta
Maria
11
9
0.82
8.07
0.73
0.90
1.2
1.4±1.5
4
2
0.50
1.60
0.40
0.80
2.0
0.2±0.2
17
16
0.94
15.21
0.89
0.95
1.1
0.1±0.3
20
18
0.90
16.67
0.83
0.93
1.1
0.7±0.8
20
20
1.00
20.00
1.00
1.00
1.0
0.9±1.0
Total
90
78
0.87
73.12
0.81
0.94
1.15
0.7±1.0
Mean
12.85
11.14
0.81
10.45
0.75
0.90
1.3
0.7
s.d.
6.23
6.89
0.22
6.95
0.27
0.11
0.5
0.5
P. lobata
Caño Island
Marino
Ballena
Cocos Island
* P. evermanni means exclude Punta Ulloa