ENDANGERED SPECIES RESEARCH
Endang Species Res
Vol. 7: 167?174, 2009
doi: 10.3354/esr00117
Printed July 2009
Published online August 13, 2008
INTRODUCTION
The number of species that exist on Earth is continu-
ally being revised and contested and is by no means a
trivial consideration. It is generally believed that the
stability and persistence of ecosystems is, in part, pred-
icated on species richness (Reusch et al. 2005), and sur-
veys aimed at compiling biodiversity inventories con-
? Inter-Research 2009 ? www.int-res.com*Email: mtcraig@hawaii.edu
How many species of goliath grouper are there?
Cryptic genetic divergence in a threatened marine
fish and the resurrection of a geopolitical species
M. T. Craig1,*, R. T. Graham2, R. A. Torres3, J. R. Hyde4, M. O. Freitas5,
B. P. Ferreira6, M. Hostim-Silva7, L. C. Gerhardinger8, A. A. Bertoncini9,
D. R. Robertson10
1Hawaii Institute of Marine Biology, University of Hawaii at Manoa, PO Box 1346, Kaneohe, Hawaii 96744, USA
2Wildlife Conservation Society, Black Orchid Street, PO Box 37, Punta Gorda, Belize
3Laborat?rio de Gen?mica Evolutiva e Ambiental, Departamento de Zoologia, Universidade Federal de Pernambuco (UFPE),
Av. Prof. Moraes Rego, 1235-Cidade Universit?ria, Recife, Pernambuco 50670-420, Brazil
4Southwest Fisheries Science Center, NMFS, 8604 La Jolla Shores Dr., La Jolla, California 92037, USA
5Universidade Estadual de Santa Cruz (UESC), Programa de P?s-Gradua??o em Sistemas Aqu?ticos Tropicais,
Rodovia Ilh?us-Itabuna km 16, CP 110, Salobrinho, Ilh?us, Bahia 45650-000, Brazil
6Universidade Federal de Pernambuco (UEPE), Av. Prof. Moraes Rego, 1235-Cidade Universit?ria, Recife, 
Pernambuco 50740-550, Brazil
7Universidade do Vale do Itaja? (UNIVALI), CTTMar - Laborat?rio de Ci?ncias Ambientais, Rua Uruguai 458,
Centro Caixa Postal 360, Itaja?, Santa Catarina 88302-202, Brazil
8ECOMAR NGO, Multi-Institutional Program on Local Knowledge and Practices, Caravelas, Bahia, Brazil
9Universidade Federal de S?o Carlos (UFSCar), PPGERN, CxP.676 S?o Carlos, San Paulo 13565-905, Brazil
10Smithsonian Tropical Research Institute, Balboa, Republic of Panam?. Mailing address: STRI, Unit 0948, APO AA 34002, USA
ABSTRACT: The goliath grouper Epinephelus itajara (Epinephelidae) is an exceptionally large
marine fish that inhabits sub-tropical and tropical waters of the Americas and western Africa. Due to
a lack of readily observable morphological variation in specimens across its range, the goliath
grouper has been regarded as a single species. We tested the hypothesis that Pacific and West
Atlantic populations constitute a single species by analyzing nuclear and mitochondrial DNA
sequence data. We found numerous fixed genetic differences for mitochondrial loci between Pacific
and West Atlantic goliath grouper (genetic distance D ? 3.5% at 16S and D ? 6% at cytochrome b; ?st
= 0.98 [p < 0.001] for 16S and ?st = 0.98 [p < 0.001] for cytochrome b). The nuclear S7 intron showed 3
fixed nucleotide differences between Pacific and West Atlantic populations. Within the West Atlantic,
we found few absolute genetic differences (D < 0.01 at 16S and D < 0.02 at cytochrome b), but statis-
tically significant population structure based on haplotype frequency data (?st = 0.04 [p = 0.05] at 16S;
?st = 0.14 [p < 0.001] at cytochrome b). These data indicate that (1) goliath grouper in the West
Atlantic are subdivided into discrete populations, (2) goliath grouper populations in the Pacific and
western Atlantic represent 2 (or more) distinct species, and (3) these distinct populations/species
require separate management and conservation strategies. We resurrect the species Epinephelus
quinquefasciatus (Bocourt 1868) for Pacific goliath grouper.
KEY WORDS:  Epinephelus itajara ? Epinephelus quinquefasciatus ? Phylogeography ? Epinephelidae
Resale or republication not permitted without written consent of the publisher
OPEN
 ACCESS
Contribution to the Theme Section ?Range-wide status and conservation of the goliath grouper?
Endang Species Res: 7: 167?174, 2009
tinue to be at the forefront of conservation efforts. Sur-
prisingly, this basic information is lacking in many
areas, despite its obvious importance for conservation
measures.
In the marine environment, biodiversity inventories
often reflect the fact that marine species tend to be
more broadly distributed than their terrestrial counter-
parts, with some organisms ranging more than
10 000 km (Jablonski & Lutz 1983). With the now wide-
spread use of molecular genetic techniques, it has be-
come clear that cryptic genetic diversity may confound
estimates of both species ranges and biodiversity (re-
viewed in Bickford et al. 2006). Species once thought to
be distributed over immense oceanic expanses are
now known to be comprised of discreet lineages that
may or may not occur in sympatry (reviewed in Rocha
et al. 2007). Although this phenomenon was thought to
occur most commonly in small or difficult-to-study or-
ganisms, recent evidence shows that these cryptic ge-
netic lineages also occur amongst Earth?s largest and
most well-studied marine animals (e.g. Bass et al. 2005,
Quattro et al. 2006, Vianna et al. 2006). In addition,
these genetic studies often elucidate genetic patterns
that reflect demographic connectivity, a critical aspect
of population biology that is often unknown but imper-
ative for effective conservation strategies.
The goliath grouper Epinephelus itajara (Epineph-
elidae) is one of the largest reef fishes on the planet,
reaching over 2 m in length and nearly 450 kg in
weight (Sadovy & Eklund 1999). Although removed
from the US NOAA Species of Concern list in 2006,
goliath grouper are still regulated as a ?no take? species
in the US. This high level of protection for the species
has triggered an increase in juvenile goliath grouper in
the southeastern US. However, goliath grouper popu-
lations are scarce throughout a majority of their range,
and because of their shrinking numbers overall and
the increasing threats they face, goliath grouper have
been placed on the IUCN Redlist in the category ?Crit-
ically Endangered?. Placement of species in this cate-
gory is reserved for the most threatened and impacted
species on the planet, indicating an urgent need to
increase our conservation efforts for this species.
Goliath grouper have a relatively large range for a
marine fish, occurring in sub-tropical and tropical
waters of the Pacific and Atlantic coasts of the Ameri-
cas and West Africa (Heemstra & Randall 1993). No
morphological differences have been identified to date
between these widely distributed populations, even
between those that are separated by the most well-
studied and impassible biogeographic barrier, the Isth-
mus of Panama (Smith 1971, Heemstra & Randall 1993,
M. T. Craig unpubl. data). As a consequence, goliath
grouper continues to be recognized as a single species
across its range.
In an attempt to resolve the specific status of goliath
grouper populations, we sampled goliath grouper from
throughout its amphi-American range to test the
hypothesis that these geographically disjunct popula-
tions are a single species using both nuclear and mito-
chondrial DNA sequences. We show that contrary to
accepted taxonomy, amphi-American populations of
goliath grouper are separated by a high degree of
fixed genetic divergence that is on par with other mor-
phologically identifiable species. Our data indicate
that, despite the absence of a set of diagnosable mor-
phological differences, the goliath grouper population
in the eastern Pacific is best regarded as a separate
and distinct species.
MATERIALS AND METHODS
Tissue samples of Epinephelus itajara were collected
from 4 localities: Panama City, Panama (Pacific), Punta
Gorda, Belize (Caribbean), Goodland, Florida, USA
(Caribbean), and Caravelas, northeastern Brazil (West
Atlantic). Three whole specimens from Panama were
deposited as voucher specimens at the Scripps Institu-
tion of Oceanography Marine Vertebrates Collection
(SIO 00-185). Several attempts were made to obtain
specimens from West Africa, part of the historical
range of E. itajara. None, however, were obtained, and
an informal survey of fisherman from Mauritania,
Senegal, Gambia, Guinea Bissau, Republic of Guinea
and Sierra Leone revealed that few, if any, goliath
grouper have been landed in those areas for some time
(~10 yr), indicating that it may be locally extinct in this
part of its range (B. Seret & J.-D. Durand pers. comm.).
Tissues were taken from specimens captured by com-
mercial and recreational fishermen at the collecting
locality. Tissue samples were preserved in 95%
ethanol or sarcosyl-Urea and stored at ambient tem-
perature in the laboratory. Total genomic DNA was
isolated with the DNEasy isolation kit (Qiagen) follow-
ing manufacturer?s protocols. Extracted DNA was
frozen in TE buffer and archived at ?20?C. Polymerase
chain reaction (PCR) was used to amplify an approxi-
mately 770 base pair (bp) fragment of the mitochondr-
ial cytochrome b (cyt b) gene and an approximately
590 bp fragment of the 16S rRNA gene. In order to
evaluate the utility of nuclear loci, a subset of samples
was screened at the nuclear S7 intron. Sample sizes for
each gene and each population are listed in Table 1.
For cyt b, initial amplification was carried out using
universal primers designed by Song et al. (1998) and
Taberlet et al. (1992). Only weak amplifications were
attained, and species-specific primers were designed
by eye to enhance amplification and sequencing.
Primers used for all genes are listed in Table 2. PCR
168
Craig et al.: How many species of goliath grouper are there?
reaction mixes were prepared using BioMix Red (Bio-
line) following the manufacturer?s instructions with the
addition of 1 ?M of each primer for cyt b and 10 ?M for
16S and S7, and 10 to 100 ng DNA template. The sam-
ples from Florida did not consistently amplify under
the above conditions. Thus, 10 ?l reactions were pre-
pared as follows: 67 mM Tris-HCl pH 8.8, 16.6 mM
(NH4)2SO4, 10 mM ?-mercaptoethanol, 2 mM MgCl2,
800 ?M dNTPs, 0.5 ?M each primer, 0.5 mg ml?1 BSA,
0.5 units Taq DNA polymerase (New England Biolabs)
and 0.5 ?l of stock DNA.
For the Panama, Belize, and Brazil samples, PCR re-
actions were carried out using an initial denaturing step
at 94?C for 2 min, followed by 35 cycles of amplification
(30 s of denaturation at 94?C, 30 s of annealing at 48 to
50?C, and 45 s of extension at 72?C). For the Florida
samples the following conditions were used: 92?C for
2 min, followed by 45 cycles of 92?C for 20 s, 52?C for
90 s, 72?C for 90 s, and a final extension at 72?C for
3 min. Excess oligonucleotide primers and dNTPs were
removed by incubation with Exonuclease I and calf
intestine alkaline phosphatase (ExoCIAP). Direct se-
quencing reactions with fluorescently labeled dideoxy
terminators were performed according
to the manufacturer?s recommenda-
tions and analyzed with an ABI 3100
automated sequencer (Applied Bio-
systems) at the Hawaii Institute of
Marine Biology Sequencing Core Fa-
cility. Unique haplotypes and intronse-
quences were deposited in GenBank
(16S: EU445272?EU445279; cyt b:
EU445280?EU445291, EU823101?
EU823103; S7: EU494944?EU494945).
Sequences for each gene were
aligned using ClustalX (Thompson et
al. 1997) and checked by eye. Phyloge-
netic hypotheses (trees) were con-
structed using PAUP*4.0b10 under
distance, parsimony, and maximum
likelihood optimality criteria. For all
optimality criteria, default settings in
PAUP were used. In the case of the
maximum likelihood analysis, this corresponded to the
HKY85 model of nucleotide substitution. In all cases,
trees were rooted with Epinephelus fuscoguttatus.
Branch support was assessed using bootstrap with 1000
replications. Sequences for outgroups were taken from
GenBank for the species E. fuscoguttatus (AY947561.1)
and E. lanceolatus (AY947588.1) for the 16S gene, and
E. lanceolatus for cyt b (DQ486927, DG486928,
DQ372727). E. lanceolatus has been confirmed as the
sister species to E. itajara by previous molecular and
morphological analysis (Smith 1971, Craig & Hastings
2007).
Population structure was assessed for both mito-
chondrial genes using Analysis of Molecular Variance
(AMOVA) as implemented in the software package
Arlequin (v.3.11; Excoffier et al. 2005). We first eva-
luated population structure using all sample locations,
and then excluded those from Panama to assess
population structure among Atlantic localities. Default
settings in Arlequin were used for all AMOVA, with
the exception that the Kimura 2 parameter model
of nucleotide substitution was used in distance
calculations.
169
Gene/Site N No. of No. of unique Haplotype Nucleotide
haplotypes haplotypes diversity diversity
16S
Panama 25 3 3 0.156 0.0005
Belize 24 2 0 0.083 0.0001
Brazil 20 1 0 0.000 0.0000
Florida 30 4 2 0.303 0.0006
cyt b
Panama 17 5 5 0.684 0.0013
Belize 23 4 3 0.525 0.0007
Brazil 18 5 4 0.601 0.0011
Florida 17 3 2 0.324 0.0004
S7
Panama 4 1 1 0.000 0.0000
Belize 4 1 0 0.000 0.0000
Brazil 5 1 0 0.000 0.0000
Florida 4 1 0 0.000 0.0000
Table 1. Epinephelus itajara. Locations, sample sizes, and genetic diversity
indices. cyt b: cytochrome b
Primer Gene Sequence Source
ItaCBF cyt b 5?-CTACAAAAACCCTATCAATGACC-3? Present study
ItaCBR cyt b 5?-GGTGAAGTTGTCTGGGTC-3? Present study
16SarL 16S rRNA 5?-CGCCTGTTTATCAAAAACAT-3? Palumbi (1996)
16SbrH 16S rRNA 5?-CCGGTCTGAACTCAGATCACGT-3? Palumbi (1996)
S7RPEX1F S7 intron 5?-TGGCCTCTTCCTTGGCCGTC-3? Chow & Hazama (1998)
S7RPEX2R S7 intron 5?-AGCGCCAAAATAGTGAAGCC-3? Chow & Hazama (1998)
Table 2. Sequencing and PCR primers used for mitochondrial and nucleardata analysis
Endang Species Res: 7: 167?174, 2009
RESULTS
Overall, we resolved 745 bp of the cyt b gene from
71 ind., 590 bp of the 16S rDNA gene from 99 ind., and
651 bp of the nuclear S7 intron from 13 ind. of goliath
grouper. We found 15 haplotypes for cyt b, 8 haplo-
types for 16S, and 2 alleles for S7. No haplotypes or
alleles were shared between the Pacific Ocean
(Panama) and Atlantic for any gene, and no indels
were observed in any alignment. Within the West
Atlantic, however, a single common haplotype was
shared between Belize, Brazil and Florida for both
cyt b and 16S.
We found few absolute genetic differences (%
sequence divergence) between Atlantic sampling
localities. However, we found several differences
between Pacific and Atlantic populations. For the mito-
chondrial 16S gene there were 20 fixed nucleotide dif-
ferences between Pacific and western Atlantic sam-
ples, while for cyt b there were 41, and 3 for the S7
intron. These putatively conspecific trans-isthmian
populations represented reciprocally monophyletic
clades with 100% bootstrap support when analyzed
using distance, parsimony, and maximum likelihood
tree-building algorithms (Fig. 1), and were separated
by 3.36 to 3.69% divergence at the 16S locus and 6.05
to 6.46% divergence at cyt b. Descriptive genetic sta-
tistics for each gene at each site are shown in Table 1.
Despite minimal genetic difference, AMOVA ana-
lyses indicated statistically significant genetic struc-
ture within the Atlantic. At cyt b all sampling sites
shared the most common haplotype; however, each
site had 2 or more haplotypes that were restricted to
170
0.01 Sub./Site
BRA2
BRA1
BRA3
BRA4
BRA5
BRA6
BRA7
BRA8
BRA9
BRA10
BRA11
BRA12
BRA13
BRA14
BRA16
BRA17
BRA19
BRA20
FLA436
FLA469
FLA472
FLA480
FLA491
PG21
PG1
PG9
PG3
PG2
PG7
PG6
PG5
PG4
PG10
FLA481
FLA465
FLA445
FLA439
FLA438
BRA18
BRA15
FLA461
PG23
PG22
PG8
PG24
FLA462
FLA463
FLA464
FLA466
FLA467
FLA470
FLA471
FLA473
FLA474
FLA475
FLA476
FLA479
FLA482
FLA483
PG11
PG12
PG13
PG14
PG15
PG16
PG17
PG19
PG18
PG20
FLA437
FLA457
FLA468
FLA478
FLA477
PAN MTC6
PAN34
PAN35
PAN36
PAN37
PAN40
PAN43
PAN44
PAN45
PAN18
PAN20
PAN21
PAN22
PAN23
PAN24
PAN25
PAN29
PAN30
PAN32
PAN33
PAN46
PAN MTC4
PAN MTC5
PAN26
PAN38
E. lanceolatus (AY947588.1)
100
100
100
Fig. 1. Epinephelus spp. Phylogenetic tree
based on the mitochondrial 16S rRNA gene.
With the exception of tips, identical topol-
ogy was obtained for cytochrome b data.
Tree is rooted with E. fuscogutattus (re-
moved for clarity). Numbers above nodes
are bootstrap supports (N = 1000 reps). Lo-
cality abbreviations are: BRA = Caravelas,
Brazil; FLA = Florida, USA; PG = Punta
Gorda; Belize; PAN = Panama City, Panama.
Sub.: substitutions
Craig et al.: How many species of goliath grouper are there?
a particular locality (private haplotypes) and oc-
curred in relatively high frequency (11 to 26% of the
sample size). Among these populations at cyt b ?st =
0.14 (p < 0.001) and at 16S ?st = 0.04 (p = 0.05).
When the Panama location was included, genetic
structure was nearly fixed with ?st = 0.97 (p < 0.001)
for cyt b and ?st = 0.98 (p < 0.001) for 16S. All pair-
wise comparisons of ?st were statistically significant
(Table 3).
DISCUSSION
Uncovering hidden genetic partitions among organ-
isms is one of the most important contributions that
phylogeograhic and population genetic studies can
provide for conservation efforts. Failure to recognize
these cryptic partitions, which often reflect reduced
demographic connectivity, can impact the way in
which populations are managed (Rocha et al. 2007). In
the marine environment, unforeseen genetic structure
has been elucidated across enumerable taxa from
rockfishes (Hyde et al. 2008) to copepods (Goetz 2003)
to bryozoans (Davidson & Haygood 1999). Our results
add another layer to this complexity by adding an
exceptionally large and well-studied marine fish to the
list of genetically partitioned species.
Atlantic population structure
Unexpectedly, we found statistically significant pop-
ulation structuring within the western Atlantic. Our
results demonstrate that, while goliath grouper share a
large fraction of the total genetic diversity, there are
unique haplotypes that are distributed more dispropor-
tionately than expected by chance. This pattern is clear
for both the cyt b data (?st = 0.14, p < 0.001) and the
more slowly evolving 16S locus (?st = 0.04, p = 0.05).
These results corroborate earlier genetic comparisons
of Brazilian and US goliath grouper that identified
divergent genetic signatures of a limited number of
samples (Vaz-Perreira et al. 2007).
Belize and much of coastal Brazil are separated by a
well-recognized biogeographic barrier to dispersal,
the Amazon and Orinoco River outflows (sensu Briggs
1974 and discussed in Floeter et al. 2008). These fresh-
water systems essentially dilute typical saline condi-
tions to the point where many marine taxa cannot sur-
vive. Goliath grouper, however, are known to recruit to
brackish water conditions and spend several years in
this habitat prior to an ontogenetic shift to reef habitat
(Smith 1976, Frias-Torres 2006). Thus, we expected
that goliath grouper should be able to tolerate condi-
tions within the Amazon/Orinoco barrier and would be
able to disperse across, if not live within, this barrier.
Our results suggest that Brazilian and Caribbean
goliath grouper populations are separate and distinct;
however, the role of the Amazon/Orinoco outflow in
shaping this structure is unclear given the biology of
the species. No such barrier exists between Florida and
Belize, however, and there is little to offer in terms of a
physical explanation for the impaired gene flow shown
by our data.
These results imply that goliath grouper should be
managed as separate stocks throughout their Atlantic
range. While the cyt b gene is often considered to
evolve at a sub-optimal rate for assessing contempo-
rary gene flow, our data show a pattern that is consis-
tent with the hypothesis that goliath grouper popula-
tions are not demographically connected. This is
particularly evidenced by the presence of private hap-
lotypes that occur at relatively high frequencies (up to
26% of the sample size at each site). A similar indica-
tion of discrete populations has been indicated
between Florida and Belize based on analysis of vari-
able microsatellite loci (R. Chapman pers. comm.), and
thus we feel that the mtDNA data provide a reliable
and concordant estimate of this structure. These
results highlight the need for a directed stock analysis
of goliath grouper, and ongoing efforts to increase
sample sizes and locations, as well as to include more
variable markers, will further enhance our understand-
ing of the genetic connectivity within this species.
Divergence times and speciation
The amphi-American distribution of goliath grouper,
coupled with the noted absence of morphological dif-
ferentiation, provides a remarkable system to investi-
gate the tempo of speciation in marine fishes. The Isth-
mus of Panama has provided a wealth of information
regarding speciation mechanisms in the sea, and its
role in shaping the biogeography and genetic architec-
ture among goliath grouper is readily apparent. Fossil
records indicate the presence of the immediate ances-
tor of goliath grouper from the upper Miocene that are
171
Panama Brazil Belize Florida
Panama ? < 0.001* < 0.001* < 0.001*
Brazil 0.98 ? 0.001* 0.021*
Belize 0.98 0.16 ? 0.007*
Florida 0.98 0.08 0.15 ?
Table 3. Pairwise population ?st values (below diagonal)
and corresponding p values (above diagonal). *Significance
at p = 0.05
Endang Species Res: 7: 167?174, 2009
virtually indistinguishable from extant species (Aguil-
era & Rodrigues de Aguilera 2004). Goliath grouper,
with their predilection for estuarine habitats as juve-
niles, represent one of the last marine fishes that would
be expected to diverge during the gradual closing of
the Isthmus of Panama (final closure approximately
3.2 million years (Myr); Coates & Obando 1996). Using
this date to calibrate a molecular clock results in a rate
of 1.9?2.0% per Myr for cyt b (range D = 6.0?6.5%
uncorrected p) and 1.0?1.1% per Myr for 16S (range
D = 3.4?3.7% uncorrected p).
One potential cause for the lack of morphological dif-
ference between Pacific and Atlantic populations might
be that they have not been separated for sufficient time
in order for morphological evolution to occur. To ad-
dress this, we compiled a list of epinephelid sister spe-
cies for which genetic data are available and placed
them in the context of geological time to establish when
morphologically distinct species diverged (Table 4).
Applying the molecular evolutionary rate above to the
observed genetic difference between morphologically
distinct epinephelid sister species confirms that trans-
isthmian goliath grouper have had sufficient time for
morphological divergence; the range of divergence
estimates brackets that of goliath grouper. All sister
species that were compared showed morphological
differences ranging from subtle alterations of color
pattern (e.g. Cephalopholis colonus and C. furcifer) to
additional changes in fin-ray elements and other
meristics (e.g. C. panamensis and C. cruentata).
How many species of goliath grouper are there?
While the biological species concept (sensu Mayr
1963) remains as one of the most accepted set of crite-
ria for defining a species relying on actual or potential
interbreeding, taxonomists have historically relied on
one of several versions of the phylogenetic species
concept, relying upon a set of diagnosable morpholog-
ical characters (reviewed in Wheeler & Meier 2000).
These competing criteria have conspired to leave
Pacific goliath grouper as an unrecognized, distinct
species.
Trans-isthmian goliath grouper populations cur-
rently are reproductively isolated; the Isthmus of
Panama physically prevents interbreeding. However,
in the event of a sundering of this barrier, secondary
contact between Pacific and Atlantic populations may
prove that reproductive isolation has not occurred. (It
should be noted that while some authors have sug-
gested that goliath grouper may be able to traverse the
Panama Canal [Heemstra & Randall 1993], our results
do not support this; even though the canal has been
open for a relatively short period of time, shared haplo-
types would be expected if juveniles were traversing
this freshwater system). Simultaneously, the absence
of diagnosable morphology argues that these are truly
populations of one species.
Prior to the present study, regarding goliath grouper
populations as distinct species was not a favored strat-
egy, as to do so would have relied on a species concept
born out of convenience: the ?geopolitical species con-
cept? (sensu Karl & Bowen 1999). Geopolitical species
are defined as:
?groups of individuals [that are] confined to geograph-
ically or politically defined areas and [that] are accorded
species status independent of morphological, genetic,
and reproductive criteria.         (Karl & Bowen 1999, p. 996)
However, in light of the present data, we now have a
diagnosable set of nucleotide characters and an expec-
tation of reproductive isolation that has been main-
tained for millions of years to continue for a consider-
ably long future, both of which provide evidence that
these populations are moving along independent evo-
lutionary trajectories and should be recognized inde-
pendently (sensu Ryder 1986, Moritz 1994, Eizirik
172
Species pair 16S cyt b
% sequence Divergence % sequence Divergence
divergence time divergence time
Alphestes immaculatus/A. multigutattus 3.0 2.7 Myr 6.9 3.5 Myr
Cephalopholis cruentatus/C. panamensis 2.5 2.2 Myr ? ?
Cephalopholis colonus/C. furcifur 2.2 2.0 Myr ? ?
Dermatolepis inermis/D. striolata 1.2 1.0 Myr 7.4 3.7 Myr
Epinephelus clippertonensis/E. labriformis 0.0 <10 kyr 0.0?0.5 <10 kyr
Epinephelus itajara (Pac)/E. itajara (Atl) 3.5 2.9 Myr 6.0 3.0 Myr
Epinephelus lanceolatus/E. itajara (Pac) 3.9 3.5 Myr 11.0 5.5 Myr
Epinephelus lanceolatus/E. itajara (Atl) 3.5 2.9 Myr 11.0 5.5 Myr
Hyporthodus nigritus/H. exsul 3.5 2.9 Myr ? ?
Table 4. Genetic distances (uncorrected p) and estimated divergence times for sister species of grouper. Data from Craig et
al. (2004) and Craig & Hastings (2007). Atl: Atlantic; Pac: Pacific. Myr: million years; kyr: kilo year; ?: no data available
Craig et al.: How many species of goliath grouper are there?
1996). It is thus clear that Pacific and Atlantic goliath
grouper represent 2 distinct species.
The resolution for the taxonomic status of goliath
grouper species is relatively straightforward. The type
locality for Epinephelus itajara (Lichtenstein 1822) is
Brazil (holotype, as Serranus itajara, ZMB 238), and
thus the Atlantic populations would retain this name
with priority. The oldest available name for an eastern
Pacific population is E. quinquefasciatus (Bocourt
1868) (type locality Pacific coast of Guatemala; holo-
type, as S. quinquefasciatus, MNHN 0000-5211). It is
thus our recommendation, following the rules of the
International Commission on Zoological Nomenclature
(ICZN), that this name be applied to Pacific popula-
tions of goliath grouper. We suggest common names of
Atlantic goliath grouper and Pacific goliath grouper,
respectively.
Underestimating biodiversity
The use of traditional species concepts in the docu-
mentation of Earth?s biodiversity remains pervasive
among common practices. For all practical purposes,
the primary criterion for determining what is or is not a
species is morphological distinctiveness. The lack of
attention to the contribution that genetic diversity
makes to overall biodiversity estimates has recently
been highlighted as a potential pitfall of current meth-
ods (reviewed in Bickford et al. 2006). Our results echo
earlier cautions that cryptic genetic lineages may con-
found our ability to adequately estimate biodiversity
(e.g. Fukami et al. 2004) and also add another caution:
large organisms cannot be ignored in this regard. Our
data, along with other recent studies (e.g. Bass et al.
2005, Quattro et al. 2006, Vianna et al. 2006) clearly
show that the phenomenon of hidden genetic diver-
gence is not restricted to understudied or difficult-to-
study organisms, as has been the basis of conventional
wisdom. In fact, goliath grouper are at the forefront of
conservation efforts, as is evidenced by their presence
as the focus of recent symposia, their listing by the
IUCN as ?Critically Endangered,? their prior listing as a
species of concern in US waters by US Fish and
Wildlife Service, and a moratorium on their capture in
both Brazil and the US.
CONCLUSIONS
While the goliath grouper has been recognized as a
single, broadly distributed species, our data clearly
show that this is not the case. We have shown that
there are at least 2 species of goliath grouper, one in
the Pacific (now treated as Epinephelus quinquefascia-
tus [Bocourt]) and one in the Atlantic (E. itajara [Licht-
enstein]), and that there are discrete populations in the
Atlantic. The taxonomic status of eastern Atlantic
goliath grouper is unknown, but best treated as E. ita-
jara until ? or possibly if ? more evidence is brought to
light.
There is now the opportunity to evaluate each of
these species independently in terms of its potential
risk for extinction following IUCN criteria, as well as
the opportunity to develop more realistic conservation
measures for these important marine species. Given
the noted decline in goliath grouper throughout its
range (in particular the absence of the largest individ-
uals once known to be common; L. McClenachan 2009,
this Theme Section), this is not a trivial task. The
absence of goliath grouper from the West African fish-
ery coupled with its uncertain taxonomic status may be
a harbinger of extinction; just as David, we may have
slain an oceanic Goliath.
Acknowledgements. This work was partially funded by Petro-
bras S/A (Programa Petrobras Ambiental) and Conservation
International Brazil to Projeto Meros do Brasil (www.meros-
dobrasil.org), The Summit Foundation to the Belize goliath
grouper project, and the National Science Foundation (grant
nos. OCE-0454873 and EPS-0554657). M.T.C was supported
by the HIMB-NWHI Coral Reef Research Partnership during
the course of this research (NMSP MOA 2005-008/66882). We
are grateful to B. Bowen, S. Karl and L. Rocha for thoughtful
discussion and comments on earlier versions of this manu-
script. We thank H. J. Walker and C. Klepadlo, SIO Marine
Vertebrates Collection for curatorial assistance. This is
SOEST contribution no. 7311 and HIMB contribution no.
1304.
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174
Editorial responsibility: Kevin Rhodes,
Hilo, Hawaii, USA
Submitted: February 19, 2008; Accepted: June 29, 2008
Proofs received from author(s): August 7, 2008