Genetic similarity between Boccardia proboscidea from Western North America and
cultured abalone, Haliotis midae, in South Africa
Carol A. Simon
⁎, Daniel J. Thornhill
, Fernanda Oyarzun
, Kenneth M. Halanych
Department of Zoology and Entomology, Rhodes University, Grahamstown 6140, South Africa
Department of Biological Sciences, Auburn University, 101 Rouse Life Sciences Building, Auburn University, Auburn, AL 36849, USA
Department of Biology, Bowdoin College, 6500 College Station Rd., Brunswick, ME 04011, USA
Friday Harbor Laboratories, University of Washington, Seattle, Box 351800, Seattle, WA 98195-1800, USA
Department of Biology, University of Washington, Seattle, Box 351800, Seattle, WA 98195-1800, USA
Received 12 February 2009
Received in revised form 26 May 2009
Accepted 26 May 2009
South African cultured abalone, Haliotis midae, are commonly infested by the non-indigenous spionid
polychaete, Boccardia proboscidea. This annelid species occurs naturally along the west coast of North
America and around Japan, but has also been introduced in Hawai i, Australia, New Zealand and perhaps the
Iberian Peninsula. Reportedly, worms were inadvertently transported to South Africa on Haliotis rufescens
imported from California in the late 1980s. To test this hypothesis, populations from six abalone farms on the
west, south and east coasts of South Africa were compared with populations from California (Alamitos Bay
and La Jolla), Washington State (False Bay Harbour) and British Colombia (Vancouver Island). Sequence data
of 16S rRNA and cytochrome b(Cyt b) mitochondrial genes show a genetic similarity between worms from
South Africa and the west coast of North America with identical haplotypes for each gene found among these
populations. The data also indicate that worms were spread among farms in South Africa primarily through
the transport of infested abalone.
© 2009 Elsevier B.V. All rights reserved.
Recently, increased attention has been paid to infestations of non-
indigenous species such as shell-boring polychaetes on commercially
harvested molluscs (Kuris and Culver, 1999; Bailey-Brock, 20 00; Read
and Handley, 2004; Bertrán et al., 2005; Radashevsky and Olivares,
2005; Vargas et al., 2005; Moreno et al., 2006; Sato-Okoshi et al.,
2008). These annelid worms may reduce the ﬂesh condition of
cultured molluscs through delayed growth rates and increased
mortality, thereby increasing production costs (e.g., Martin and
Britayev, 1998; Lleonart, 2001; Simon et al., 2006). Furthermore,
animal transportation for aquaculture is considered a leading vector
for introduction of non-indigenous marine species, including poly-
chaetes, which are inadvertently transferred with their hosts (Naylor
et al., 2001). Thus, putative origins of some non-indigenous species
may be inferred based on the origin of imported aquaculture stocks
(Bailey-Brock and Ringwood, 1982; Kuris and Culver, 1999; Bailey-
Brock, 2000; Radashevsky and Olivares, 2005; Moreno et al., 2006).
However, when a non-indigenous pest living on cultured species does
not infest economically important shellﬁsh in its native range,
establishing its putative origin becomes more problematic. In such
instances, vectors of transportation other than the importation of
organisms for aquaculture, such as ballast water (Carlton and Geller,
1993; Carlton, 1996), must also be considered.
South Africa, the second largest supplier of cultured abalone in the
world, with just less than 850 MT exported in 2008 (Troell et al., 2006;
Wayne Barnes, Abalone Farmers Association of Southern Africa, pers.
comm.). Increased infestation of abalone by spionid polychaetes has
negatively impacted some farms. In one instance, approximately
500,000 abalone were culled at a single farm due to spionid
infestations (R. Clark, Wild Coast Abalone, pers. comm.). The cultured
abalone, Haliotis midae Linnaeus 1758, in South Africa are infested by a
number of spionids, including indigenous Dipolydora capensis (Day
1955) and Polydora hoplura Claparède 1870, and by non-indigenous
Boccardia proboscidea Hartman 1940 which were ﬁrst reported in
South Africa in 2004 (Simon et al., 2006; Simon and Booth, 2007). To
date, B. proboscidea has not been detected in naturally occurring
shellﬁsh in South Africa (Simon et al., in press).
The known native range of B. proboscidea includes Japan and the
western coast of North America, from British Columbia to southern
California, with unconﬁrmed records extending the distribution even
further south (Hartman, 1940; Woodwick, 1963; Fauchald, 1977;
Petch, 1995; Sato-Okoshi, 2000; F. X. Oyarzun et al., unpublished data).
In its native range, B. proboscidea occupies a wide ecological niche
burrowing into soft rock and in crevices, among encrusting algae and
Aquaculture 294 (2009) 18–24
⁎Corresponding author. Current address: Department of Botany and Zoology,
Stellenbosch University, Matieland, Private Bag X01, Stellenbosch 7602, South Africa.
Tel.: +27 21 808 3068 ; fax: +27 21 808 2405.
E-mail addresses: email@example.com (C.A. Simon), firstname.lastname@example.org
(D.J. Thornhill), email@example.com (F. Oyarzun), firstname.lastname@example.org
0044-8486/$ –see front matter © 20 09 Elsevier B.V. All rights reserved.
Contents lists available at ScienceDirect
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in muddy and sandy sediments (Hartman, 1940; Woodwick, 1963;
Gibson et al., 1999). Although it occurs on gastropod shells and mud
deposits in the crevices of Crassostrea gigas Thunberg 1793 shells, B.
proboscidea has never been recorded on abalone shells in its natural
range (Woodwick, 1963; Blake and Evans, 1973; Martin and Britayev,
1998; Sato-Okoshi, 2000). B. proboscidea has been introduced to
Hawai i with cultured oysters (Bailey-Brock, 2000) and to Australia via
either ballast water or mariculture (where it has now spread into
natural environments and cultured molluscs; Blake and Kudenov,
1978; Lleonart, 2001; Hewitt et al., 2004; Sato-Okoshi et al., 2008). It
has also been recorded on abalone in New Zealand (Read, 2004) and
living amongst Zostera, calcareous algae, Mytilus and in rock crevices
on the Iberian Peninsula (Martínez et al., 2006).
Over the last 70 years, South African aquaculturists have imported
several bivalve and gastropod species that could have served as vectors
for B. proboscidea, from Europe, California and Chile (Robinson et al.,
2005). For example, the abalone, Haliotis rufescens Swainson 1822, was
imported to Saldanha Bay from California in the 1980s; however, all
individuals died within a month and were never re-imported (Grifﬁths
et al.,1992). Additionally, since the 1970s, the bulk of the oyster industry
has been maintained by spat of C. gigas imported regularly from Chile,
the United Kingdom and France (Grifﬁths et al., 1992; Robinson et al.,
2005). Based solely on the history of molluscan imports to South Africa
and the known distribution of B. proboscidea, it is impossible to
determine the source or vector of transportation of the South African
B. proboscidea population. As a result, alternative approaches are
required to address this question.
In this study, we utilized a molecular approach based on 16S rRNA
and cytochrome b(Cyt b) mitochondrial gene sequence data to infer the
source population of non-indigenous B. proboscidea infesting farmed H.
midae in South Africa. The west coast of the United States was
hypothesized to be the source population of B. proboscidea,asitwas
the source of the abalone suspected to have introduced this infestation.
Previous studies have successfully used molecular techniques to
establish the (presumed) origin of non-indigenous species, including
spionids (Bastrop et al., 1998; Bastrop and Blank, 2006) and molluscs
(Dupont et al., 2003; Martel et al., 2004; McGlashan et al., 2008).
Therefore, patterns of evolutionary relatedness based on Bayesian and
coalescent analyses may elucidate whether infesting B. proboscidea in
South Africa originated on the west coast of the US.
2. Methods and materials
2.1. Data collection
B. proboscidea was collected from infested abalone from South
African farms near Saldanha Bay on the west coast (Jakobsbaai Sea
Products [Pty] Ltd), in Walker Bay on the south coast (Abagold [Pty]
Fig. 1. Localities of collection sites of Boccardia proboscidea and the major harbours in South Africa with the ﬂow direction of the predominant currents indicated. Localiti es included:
JB (Jakobsbaai Sea Products [Pty] Ltd), Ab (Abagold [Pty] Ltd), AF (Atlantic Fishing), RB (Roman Bay Sea Farm [Pty] Ltd), IJ (Irvin and Johnson Ltd), and HH (Wild Coast Abalone [Pty]
Ltd at Haga Haga).
19C.A. Simon et al. / Aquaculture 294 (2009) 18–24
Ltd, Atlantic Fishing, Roman Bay Sea Farm [Pty] Ltd, and Irvin and
Johnson Ltd), and north of Haga Haga on the east coast (Wild Coast
Abalone [Pty] Ltd) (Fig. 1; see Table 1 for the codes that will be used
throughout the paper) of South Africa from March to April 2007 and
preserved in 70% ethanol. DNA was extracted using a Qiagen DNeasy®
Tissue Kit according to the manufacturer's instructions. An approxi-
mately 376 bp fragment of the cytochrome b(Cyt b) mitochondrial
gene (Table 1) was ampliﬁed using the primers of Boore and Brown
(2000):‘Cyt b424F’(5′-GGW TAY GTW YTW CCW TGR GGW CAR AT-
3′) and ‘Cyt b876R’(5′-GCRT AWG CRA AWA RRA ART AYC TC-3′),
with the cycling conditions: initial denaturation 94 °C, 3 min; 40
cycles of 94 °C, 30 s, annealing 45 °C, 30 s, extension 72 °C,1 min; ﬁnal
extension 72 °C, 7 min. An approximately 439 bp fragment of the 16S
rRNA mitochondrial gene (Table 1) was ampliﬁed using the primers of
Palumbi et al. (1991):‘16SarL’(5′-CGC CTG TTT ATC AAA AAC AT-3′)
and ‘16SbrH’(5′-CCG GTC GAA CTC AGA TCA GCT-3′), with the cycling
conditions: initial denaturation 94 °C, 3 min; 40 cycles of 94 °C, 30 s,
annealing 48 °C, 30 s, extension 72 °C, 1 min; ﬁnal extension 72 °C,
7 min. All PCR products were veriﬁed by 1× sodium borate (SB) agarose
gel electrophoresis. 16S PCR products were gel-puriﬁed using the
QIAquick® PCR Puriﬁcation Kit, while Cyt bPCR products were puriﬁed
with the Montage™PCR Filter Units (Millipore). Puriﬁed PCR products
were bidirectionally sequenced using Genome Lab™Quick Start Mix
(Beckman Coulter) on a Beckman CEQ 8000 using the same primers as
for PCR, with one exception: ‘Cyt b876R’was replaced by Cyt bBP 876R
(5′-RAA WAR RAA GTA TCA YTC AGG-3′). Sequences were edited in
Sequencher v4.6 (Gene Codes Corporation, Ann Arbor, Michigan). The
Cyt bsequences were translated using the Drosophila mitochondrial
code in McClade v4.06 (Maddison and Maddison, 2003) to ensure that
stop codons were not present. All sequences were deposited in GenBank
(Accession nos. FJ434476–FJ434482, FJ434486; Table 2).
Available sequence data for B. proboscidea from the western coast
of North America included free-living samples from the intertidal of
Vancouver Island (Accession nos. FJ434483–FJ434485 and FJ434487–
FJ434489), California and Washington State (Accession nos.
FJ972541–FJ972569, see Appendix A). Initial data collection revealed
that Cyt bsequences were more variable than 16S, and therefore more
informative to investigate the putative source population. Therefore,
subsequent data collection efforts emphasized collecting Cyt b, rather
than 16S, sequence data.
2.2. Data analysis
Sequence data from South Africa was compared to North American
data to determine whether haplotypes were shared between regions.
Sequences were aligned using Clustal W (Thompson et al., 1994)in
Bioedit (Hall, 1999). Sequence data were not concatenated as 16S data
of the South African samples possessed no variation and thus provides
limited information about the South African samples. Thus, analyses
herein focused on Cyt b. Alignments are provided in TreeBase (www.
To further explore the relationship between B. proboscidea
populations, phylogenetic relationships among haplotypes were
estimated using Bayesian inference in MrBayes v3.1 (Ronquist et al.,
2005). Based on the availability of sequence data, Marenzallaria
neglecta Sikorski and Bick, 2004 (GenBank Accession no. DQ309261)
was selected as the outgroup for Cyt banalyses, while Dipolydora
giardi (Mesnil, 1896) (GenBank Accession no. DQ779632) was
selected for 16S analyses. The HKY + Γmodel of substitution, as
suggested by a hierarchical Likelihood Ratio Test (hLRT) in MrMo-
deltest v2.2 (Nylander, 2004), was implemented for Cyt bsequence
data. Two sets of four chains (3 hot, 1 cold) were run simultaneously to
1.0 × 10
generations and sampled every 100 generations. In initial
analysis using M. neglecta as an outgroup, the ﬁrst 1000 burn-in
generations were discarded. Burn-in values were based on the
convergence of likelihood scores in the chains. None of the nodes
was well-supported. Further analysis, using only the unique haplo-
types, was run for 2 × 10
generations and sampled every 100
generations without selection of an outgroup. The ﬁrst 305 burn-in
generations were discarded. Phylogenetic relationships were also
estimated from 16S sequence data via Bayesian analysis implementing
the K80+ I model of substitution suggested by hLRT in MrModeltest
v2.2 (Nylander, 2004). Two sets of four chains (3 hot, 1 cold) were run
simultaneously to 2× 10
generations and sampled every 100
generations, with the ﬁrst 450 burn-in generations being discarded.
In all cases, 50% majority-rule consensus trees were computed.
Finally, haplotype networks representing intraspeciﬁc relation-
ships were constructed in TCS v1.21 (Clement et al., 2000) with gaps
treated as missing data with other options set to their default values.
Connectivity levels were examined at 92%–95% for Cyt bsequence
data and at 95% for 16S. Reticulations between haplotypes were
resolved following Crandall et al. (1994).
Forty-eight individuals from South Africa were sequenced for Cyt b
(376 bp) and 26 for 16S (439 bp) resulting in seven Cyt bhaplotypes
but only one 16S haplotype (Table 2). For Cyt bthere was a single
haplotype that represented 40 individuals (haplotype 1), and four
haplotypes (n=6) that each differed from this haplotype by a single
nucleotide substitution (Table 2;Fig. 2). The Cyt bgene had ﬁve (1.3%)
variable characters, of which two (0.5%) were parsimony informative.
The percentage GC content was 47.8% for Cyt band 44.9% for 16S.
With the inclusion of sequences from North American sites, there
were 111 and 50 sequences for Cyt band 16S genes, respectively. In
this expanded dataset, the Cyt bgene had 37 (9.8%) variable
characters, of which 19 (5.1%) were parsimony informative, whereas
GC content was 47.5%. For the 16S gene, there were eleven (2.5%)
variable characters, of which three (0.7%) were parsimony informa-
tive, and GC content was 44.8%. All characters could be aligned
Distribution of South African haplotypes, with respective accession numbers, at the six
Population (farm) Total
West coast South coast East coast
JB Ab AF RB IJ HH
1 FJ434476 6 8 8 7 7 4 40
2 FJ434477 1 1
3 FJ434478 1 1 2
4 FJ434479 1 1
5 FJ434480 1 1
6 FJ434481 1 1 2
7 FJ434482 1 1
A FJ434486 8 2 1 7 6 2 26
Collection locations and number of individuals sequenced per sampling site.
Locality Code Number of 16S
Number of Cyt b
Jakobsbaai Sea Products [Pty] Ltd JB 8 6
Abagold [Pty] Ltd Ab 2 10
Atlantic Fishing AF 1 10
Roman Bay Sea Farm [Pty] Ltd RB 7 10
Irvin and Johnson Ltd IJ 6 8
Wild Coast Abalone [Pty] Ltd HH 2 4
Total 26 48
20 C.A. Simon et al. / Aquaculture 294 (2009) 18–24
unambiguously for both genes. The maximum uncorrected genetic
distances between haplotypes were p=0.01258 and p= 0.04740 for
16 S and Cyt b, respectively.
3.1. Phylogenetic analyses
Bayesian analysis of Cyt bwith M. neglecta as the outgroup
produced a clade which included all samples from South Africa,
Washington (False Bay Harbour), Canada (Vancouver Island), and
three samples from California (Alamitos Bay). Most of the Californian
samples were outside of this clade. However, support for these groups
was poor, with posterior probabilities of 0.50 and 0.64 for the
geographically mixed and California clades, respectively (data not
shown). When reanalysed with only the 32 unique haplotypes (and
excluding the outgroup due to its extreme branch length), nodal
support increased to a posterior probability value of 1.00 (Fig. 2).
Bayesian analysis of 16S data (including 14 unique haplotypes and
outgroup) produced a tree with no resolution (data not shown). This
difference in resolution between 16S and Cyt bmitochondrial genes
has been previously observed in marine invertebrates (e.g. Wilson
et al., 2007; Hunter and Halanych, 2008), with 16S being consistently
less variable than Cyt b. The patterns observed here highlight the
importance of selecting the appropriate gene for the question being
addressed in intraspeciﬁc genetic studies.
3.2. Coalescent analyses
Consistent with relationships inferred from the Cyt bhaplotype-only
Bayesian analysis, coalescent analysis of Cyt bby TCS at a 95%
connectivity level generated two haplotype networks with a total of
29 haplotypes. When the connectivity level was reduced to 92%, the two
networks fused into a single haplotype network. For the sake of clarity,
hereinwe focus on results using only the 95% connectivelylevel. The two
resulting networks, designated Networks 1 and 2 (Fig. 3a and b),
corresponded to the mixed clade and California grouping generated by
the Bayesian analysis. In Network 1, a single haplotype was common to
South Africa (n=40), Vancouver Island (n=5), Washington (n=3),
and Alamitos Bay (California, n=2) (Fig. 3a). This network also
included six Cyt bhaplotypes (1 or 2 individuals per haplotype) unique
to South Africa (Fig. 3a). Populations at Abagold [Pty] Ltd (Ab) and
Roman Bay Sea Farm [Pty] Ltd (RB) each had two unique haplotypes,
while one haplotype each was shared by populations at Atlantic Fishing
(AF), RB, and Irvin and Johnson Ltd (IJ), respectively (Tab le 2). Three
more haplotypes represented specimens from California (Alamitos Bay),
Vancouver Island, and Washington State (Fig. 3). Network 2 comprised
19 haplotypes composed exclusively of individuals from southern
California (Alamitos Bay and La Jolla; Fig. 3b).
As mentioned previously, sequence diversity of 16S ribosomal
mtDNA was low compared to that of Cyt bmtDNA. Parsimony network
analysis of all sequences resulted in a single network with 10 haplotypes
(Fig. 3c). The South African haplotype was common to both California
(Alamitos Bay) and Washington State. The difference in the number of
haplotypes generated by Bayesian and parsimony network analysis is a
consequence of missing data at the sequence ends and the way in which
these data are treated by the different analyses (Joly et al. 2007).
Molecular analyses of two mitochondrial genes suggest that B.
proboscidea infestations on South African abalone farms were
genetically similar to worms from the west coast of North America.
Identical 16S and Cyt bhaplotypes were found in both South Africa
and the North American west coast, indicating a common genetic
history between these geographically disparate locations.
Fig. 2. Bayesian inference phylogeny of unique Boccardia proboscidea Cyt bmtDNA haplotypes. The tree was rooted using the outgroup Marenzallaria neglecta in analyses including all
sampled individuals. Posterior probabilities were obtained from analyses that included only ingroup taxa, as the number of substitutions along the outgroup branch appeared to have
saturated a number of informative nucleotide positions. Nodal support values (≥0.5) indicated as a posterior probability next to the relevant nodes. Haplotype names correspond to
the results presented in Table 2 for South African samples and Appendix A for American samples. Names including lower case letters indicate that that haplotype has data missing
relative to the primary haplotype and are considered a single haplotype in the parsimony analysis (see Fig. 3).
21C.A. Simon et al. / Aquaculture 294 (2009) 18–24
4.1. Putative origin of infestation
The most common Cyt bhaplotype observed in South Africa is
widespread across the North American Paciﬁc coast. This haplotype
may, however, also occur in other unsampled B. proboscidea
populations. Although B. proboscidea is native to Japan (Sato-Okoshi,
2000), to the best of our knowledge, South Africa does not import live
Japanese shellﬁsh. Furthermore 16S data show that Vancouver Island
samples are distinct from more southern samples in North America
suggesting that Japanese samples are also likely to be distinct. While
conﬁrmed reports of these worms occurring in aquaculture stocks in
either Europe or Chile (Ruellet, 2004; Moreno et al., 2006)are
lacking, there are unconﬁrmed reports of this worm occurring in
claybeds in the intertidal of Harwich (Essex, England, T. Worsfold,
pers. comm.) and among Sabellaria alveolata tubes in Clarach
(Ceregidon, Central Wales, V. Cole, pers. comm.). These reports are
of particular interest as oyster spat imported to South African farms
were sourced from suppliers situated close by (A. Antonin, Striker
Oyster Fishing Company, pers. comm.). Thus, until these reports are
conﬁrmed and other populations are more comprehensively
sampled, we cannot deﬁnitely rule out alternative origins for the
South African populations. For instance, it is possible that the South
African B. proboscidea populations are a secondary introduction from
either European or Chilean aquaculture stocks that were originally
Fig. 3. (a) and (b) TCS networks of B. proboscidea based on Cyt bmtDNA haplotypes using the 95% connectivity level. Networks 1 and 2 correspond with the mixed clade and
Californian group of haplotypes, respectively (see Fig. 2). (c) TCS networks of B. proboscidea based on 16S rRNA data using the 95% connectivity level. Sampled haplotypes are
indicated by shaded circles; missing or unsampled haplotypes are indicated by black dots. Each branch indicates a single mutational difference. Circle size is proportional to observed
haplotype frequency. Haplotypes are shaded according to the geographic region from which the sample was collected (see legend). Haplotype names correspond to the results
presented in Table 2 for South African samples and Appendix A for American samples.
22 C.A. Simon et al. / Aquaculture 294 (2009) 18–24
infected by North American shellﬁsh, but where infestation has gone
undetected. However, given the notable genetic variation observed
among the natural western North American populations, it seems
unlikely that Chilean or European populations would have a genetic
signature similar to those of central California if they are in fact native
4.2. Current biogeography of haplotypes on South Africa
Populations from farms on the west and east coasts of South Africa
were each represented only by the common Cyt bhaplotype while
those from the farms in the Walker Bay area were represented by two
to four haplotypes. This difference in genetic diversity between farms
may be an artefact of uneven sampling. There were only four and six
samples from Wild Coast Abalone [Pty] Ltd (HH) and Jakobsbaai Sea
Products [Pty] Ltd (JB), respectively, compared to eight or ten from the
other four farms. Alternatively, if this diversity is real, the populations
in the South coast area may 1) be the oldest, 2) have been subjected to
more than one introduction, 3) have had the greatest haplotype
diversity at the time of introduction, or 4) have experienced fewer
population bottlenecks since introduction. Of course, several of these
factors could have worked in concert to produce the current
The genetic similarity between farms suggests that worms are
ultimately derived from the same source. These animals have
planktotrophic larvae that could potentially be released through the
seawater system and serve as a vector for dispersal to other farms.
However, B. proboscidea has not yet been detected in the surrounding
natural environment (Simon, et al., in press). A more plausible
explanation for genetically similar infestations on multiple farms is
that the worms have been spread primarily through the movement of
abalone between farms (cf., Dupont et al., 2003; McGlashan et al.,
4.3. Possible vectors of transportation
There has been a signiﬁcant increase in the incidence of marine
biological invasions as the number of human-mediated vectors,
particularly shipping ballast water and aquaculture, has increased
(e.g., Carlton and Geller, 1993; Carlton, 1996; Naylor et al., 2001). H.
rufescens, which occurs in overlapping distribution ranges with B.
proboscidea (Woodwick, 1963), were imported on a single occasion to
Saldanha Bay from California (Grifﬁths et al., 1992). Although B.
proboscidea has not been recorded on H. rufescens, it has been
recorded on other gastropods and sediment (Woodwick, 1963) which
may have been inadvertently transported with the abalone. By
contrast, South African aquaculturists regularly import the spat of C.
gigas, a known host of B. proboscidea (Bailey-Brock, 2000; Sato-
Okoshi, 2000), from Chile, the United Kingdom and France (Robinson
et al., 2005). While this polychaete has not been recorded from Chile
or France (Ruellet, 2004; Moreno et al., 2006) there are unconﬁrmed
records of this species from the United Kingdom (see above). Spat
larger than 25 mm have been imported to South Africa from the
United Kingdom where they would have been exposed to the natural
environment before transport; such spat are also large enough to host
spionid larvae or juveniles (A. Antonin, Striker Oyster Fishing
Company, pers. comm.). Imported oyster spat may therefore have
been an alternative or additional vector for the introduction of this
5. Conclusions: implications for mariculturists
Several studies (e.g., Kuris and Culver, 1999; Dupont et al., 20 03;
Martel et al., 2004; Moreno et al., 2006; McGlashan et al., 2008;this
study) have raised important issues concerning the management
and movement of aquaculture stocks and their pests. These include
the infestation of naturally occurring molluscs by non-indigenous
species that escape from aquaculture facilities, the spread of non-
indigenous species among farms and the negative economic impact
that they have on cultured shellﬁsh. Simply inspecting molluscs for
epibionts before or immediately after moving between facilities
(both international and local) might not be enough. For example,
many molluscs are infested by non-problematic spirorbid worms
which build calcium tubes on the surface of the shell. Empty
spirorbid burrows often provide a refuge for Polydora-type larvae
(Lleonart, 2001; C. A. Simon pers. obs.). These larvae would easily be
missed during a perfunctory inspection for epibionts and may
therefore go undetected during a limited quarantine period. Regular
and intensive examination of mollusc stocks is imperative to detect
the presence of potentially problematic organisms. These considera-
tions are especially important when stocks are transported around
This work was funded by the National Research Foundation and
Marine and Coastal Management of South Africa (Frontier Program
for Mariculture). All authors contributed to the generation and
analyses of data and the preparation of the manuscript. Special
thanks are due to A.R. Mahon for supplying us with North American
B. proboscidea sequences and T. MacDonald, A. Mouton, and abalone
farmers for supplying the worms. Helpful discussions and comments
on the manuscript were provided by A.R. Mahon, S.R. Santos, G.
Zardi, B. Jansen van Vuuren and M.H. Villet. USA National Science
Foundation funds are gratefully acknowledged (EAR-0120646, OCE-
0425060). This work is AU Marine Biology Program contribution
Haplotype names and accession numbers of sequences of North
American samples provided by A. Mahon (haplotypes 8–10; C, E and F)
and F. Oyarzun. Localities: Al, Ca = Alamitos Bay, California; Lj, Ca = La
Jolla, California; FBH, Wa = False Bay Harbour, Washington; Wl, Va =
Witty's Lagoon, Vancouver Island, Canada.
1 Al, Ca; FBH,
FJ972548 A Al, Ca; FBH,
8 WL, Va;
B Al, Ca FJ972542
9 WL, Va;
C WL, Va FJ434487
10 WL, Va FJ434485 D Al, Ca FJ972543
11 Al, Ca; Lj, Ca FJ972551 E WL, Va FJ434488
12 Lj, Ca FJ972552 F WL, Va FJ434489
13 Al, Ca FJ972553 G FBH, Wa FJ972544
14 Al, Ca FJ972554 H FBH, Wa FJ972545
15 Al, Ca FJ972555 J Al, Ca FJ972546
16 Lj, Ca FJ972556 K Lj, Ca FJ972547
17 Al, Ca FJ972557
18 Al, Ca FJ972558
19 Al, Ca FJ972559
20 Al, Ca FJ972560
21 Al, Ca FJ972561
22 Lj, Ca FJ972562
23 Lj, Ca FJ972563
24 Al, Ca FJ972564
25 Al, Ca; Lj, Ca FJ972565
26 Al, Ca FJ972566
27 Lj, Ca FJ972567
28 Lj, Ca FJ972568
29 Lj, Ca FJ972569
23C.A. Simon et al. / Aquaculture 294 (2009) 18–24
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