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Journal of Experimental Marine Biolog
Biomechanical properties and holdfast morphology of coenocytic
algae (Halimedales, Chlorophyta) in Bocas del Toro, Panama
Kim Anderson a, Lisa Close b, Robert E. DeWreede c,*, Brandon J. Lynch d,
Carlos Ormond e, Matt Walker f
a University of Central Oklahoma,100 N. University Drive, Edmond, OK 73034, United Statesb University of California, Davis; School of Veterinary Medicine, One Shields Avenue, Davis, CA 95616, United Statesc University of British Columbia, Department of Botany, 3529-6270 University Blvd., Vancouver, BC, Canada V6T 1Z4
d University of Arizona, Tucson, AZ 85721, United Statese The University of British Columbia, Vancouver, BC, Canada V6T 1Z4
f Texas A and M University-Kingsville, 700 University Blvd. MSC 114, Kingsville, TX 78363 * 361.593.2111, United States
Received 15 April 2005; received in revised form 4 July 2005; accepted 11 July 2005
Abstract
For attached marine organisms, specific biomechanical properties may result in detachment or in tissue loss, when sufficient
tensile force is applied. Algae experience such forces through water movement, which may thus act to limit size, abundance,
and species composition, of populations of algae.
Coenocytic construction is uncommon in the algae, but it occurs relatively more frequently in green algae found in
shallow subtidal sediments associated with coral reefs, e.g., at our study site of Isla Colon, Bocas del Toro, Panama. We
studied the biomechanical properties of some tropical coenocytic algae (Udotea flabellum (Ellis et Solander) Lamouroux,
Penicillus capitatus Lamarck, P. pyriformis A. and E.S. Gepp, and Halimeda gracilis Harvey) anchored in sediments. We
compare our results with published data on other coenocytic algae, as well as with multicellular algae. Our results show that
properties of sand-dwelling coenocytes, such as mean force to dislodge (4.9–12.7 N), mean force to break (6.6–22.1 N), and
mean strength (1.0–7.0 MN m�2), are all within the range reported for temperate, multicellular, algae. In contrast, the
coenocytes differed markedly from the temperate non-coenocytes in the consequences of applied tensile force: coenocytes
were removed whole, while most temperate algae attached to rocks break within the thallus. Some multicellular algae can
regrow from tissue left on the substratum; three of the four coenocytic species we examined had rhizoids connecting closely
adjacent (0.1–0.15 m) individuals, and these rhizoids may serve to regrow a new individual. While our experiments indicated
that sufficient tensile force results in dislodgment, calculations using the experimentally determined variables led us to
conclude that water velocities sufficient to dislodge individuals are unlikely to occur. Since dislodgment is usually fatal for
algae, the role of the holdfast is a critical one. All of the species we investigated had similar holdfast morphology, a mass of
rhizoids which entrained sand, the entire unit forming a hemispherical to cylindrical mass. Despite the consistency in holdfast
0022-0981/$ - s
doi:10.1016/j.jem
* Correspondi
E-mail addre
y and Ecology 328 (2006) 155–167
ee front matter D 2005 Elsevier B.V. All rights reserved.
be.2005.07.005
ng author. Tel.: +1 604 822 6785; fax: +1 604 822 6089.
ss: dewreede@interchange.ubc.ca (R.E. DeWreede).
K. Anderson et al. / J. Exp. Mar. Biol. Ecol. 328 (2006) 155–167156
form, and the initial prediction that this was an optimal form for anchoring these algae, our data suggest that this is not the
case.
D 2005 Elsevier B.V. All rights reserved.
Keywords: Algae; Biomechanics; Coenocyte; Detachment; Holdfast
1. Introduction
Green algae, some of which are both coenocytic
and calcified, are common in tropical shallow water
marine habitats, where they function as primary
producers and generate calcium carbonate sediments.
Unlike most temperate zone marine macroalgae,
which are predominantly multicellular, attached to
solid substrata (e.g., rocks), and not calcified, tropi-
cal coenocytes are unicellular, are frequently
anchored in sediments such as coral rubble, sedi-
ments, and mangrove detritus, and many are calci-
fied. Despite the internal structural simplicity
(connected filaments lacking separating walls), gen-
era such as Caulerpa, Halimeda, Udotea and Peni-
cillus attain a complex morphology consisting of a
rhizoidal holdfast(s), a stipe or stolon, and an
astounding morphological variation across species
and genera. For these algae, holdfast morphology
and sediment cohesiveness are important determi-
nants of the maximum tensile force they are able
to withstand. This contrasts with algae attached to
rock surfaces, where adhesion of the algal tissue to
rock is critical. At our two study sites in the vicinity
of Isla Colon, Bocas del Toro (N 98 24V 53.53, W828 19V 49.90), Panama, Udotea, Penicillus, and
Halimeda co-exist, often intermixed in habitats ran-
ging from shallow wave exposed coral sand to finer
sediment areas adjacent to the red mangrove, Rhizo-
phora mangle Linnaeus.
Among the many environmental factors determin-
ing the abundance, size, and overall morphology of
algae are forces generated by water motion, e.g., tidal
currents and wave impact (e.g., Shaughnessy et al.,
1996; Neushul, 1972; Blanchette, 1997; Denny et al.,
1997; Duggins et al., 2003). When such forces are
sufficiently great, algae may be removed in their
entirety from the rocks and sediments (Duggins et
al., 2003; Collado-Vides et al., 1998), or break within
the thallus, e.g., losing surface area to decrease drag
(Carrington, 1990; Shaughnessy et al., 1996; Blanch-
ette, 1997; Milligan and DeWreede, 2004). While
there have been many studies detailing biomechanical
properties, and effects of wave impact, on temperate
algae attached to rocks (Koehl and Wainwright, 1977;
Armstrong, 1987; Carrington, 1990; Gaylord et al.,
1994; Johnson and Koehl, 1994; Gaylord and Denny,
1997; Milligan and DeWreede, 2000; Duggins et al.,
2003; Milligan and DeWreede, 2004), few studies
have examined such features for tropical sand-dwell-
ing coenocytes (Collado-Vides et al., 1998; Padilla,
1989). We undertook this research to compare biome-
chanical properties between coenocytic and multicel-
lular algae and to examine the biomechanical
properties of sand-dwelling coenocytes in relation to
water movement they might reasonably be expected to
encounter.
To date, detailed biomechanical properties (relating
to attachment) of calcified coenocytes have only been
reported for a single species, Udotea flabellum (Col-
lado-Vides et al., 1998). Here, by investigating these
properties for three additional species, and also for
one of the same species but at a geographically distant
site, we seek to place the limited prior data into a
broader context and thus reveal any patterns that may
exist. Furthermore, given the structural differences
between multicellular and coenocytic algae, our
more extensive knowledge of the former may not be
applicable to coenocytes. For example, it has been
reported that some multicellular algae survive forces
generated through water movement by loosing thallus
tissue (e.g., Blanchette, 1997; Milligan and
DeWreede, 2004); it seems unlikely that such a
mechanism is a viable one for coenocytic algae,
given that repeatedly torn tissue could result in a
significant loss of cytoplasm. As noted by Ross et
al. (2005), Vroom et al. (2003) for Halimeda tuna,
and Vroom and Smith (2003) more generally, some
coenocytes are able to tolerate severe injury, whereas
others die as a result.
K. Anderson et al. / J. Exp. Mar. Biol. Ecol. 328 (2006) 155–167 157
For algae in general, holdfast morphology varies
from a hemispherical rhizoidal mass, to creeping
rhizomes, to solid disks. The holdfast shape of
the individuals we examined and tested was con-
sistently bowl-shaped to cylindrical, unless attached
to coral fragments; in which case the holdfast forms
a flat rhizoidal mass along the upper surface of the
coral. The bowl/cylinder form arises as the algal
filaments enclose the sediments and form a cohe-
sive mass. Some tropical sand-dwelling coenocytic
algae (e.g., some species of Udotea) have subsur-
face lateral rhizoids arising from the holdfast area
that connect to other individual(s) of the same
species (Fig. 1; and Littler and Littler, 2000);
these lateral rhizoids may give rise to new indivi-
duals, at least in culture (Hillis-Colinvaux, 1972).
If these rhizoidal connections occur generally, and
if they are able to give rise to new individuals in
situ, a dislodged individual may die, but a new
one may be regenerated. We examined the algae
we studied for the presence of these rhizoidal
connections.
Collado-Vides et al. (1998) described biomecha-
nical properties of Udotea flabellum, found at
Fig. 1. Halimeda gracilis, Udotea flabellum, and Penicillus pyr-
iformis; scale bar=4 cm. Lateral rhizoids connect two individuals of
U. flabellum and two of P. pyriformis, such rhizoids were not found
for H. gracilis neighbors.
Puerto Morelos, Mexico. The authors reported that
95% of U. flabellum individuals are removed from
the substratum in their entirety (i.e., do not break)
after applying a mean tensile force of 8.6 N (95%
C.I.=1.6), break primarily at the stipe–holdfast
junction in the laboratory (19.3 N; 95% C.I.=2.1),
and have a mean strength of 4.8 MN (95%
C.I.=0.63). Here, we report the biomechanical prop-
erties of four species of coenocytic green algae,
including Udotea flabellum. We tested the hypoth-
eses detailed below, extend the number of coenocy-
tic species for which biomechanical properties are
known, and compare these properties for the differ-
ent species we tested.
Based on the species studied to date (Collado-
Vides et al., 1998; Padilla, 1989), we hypothesized
that when sufficient tensile force is applied, coeno-
cytic individuals will not break within the thallus but
will be pulled whole from the substratum. If the
preceding holds, then the force to break an indivi-
dual exceeds the force required to remove an entire
individual. Both these predictions differ from what is
common in multicellular algae attached to rocky
surfaces, which frequently break first within the
stipe and stipe/holdfast region (e.g., Shaughnessy et
al., 1996; Utter and Denny, 1996; Johnson and
Koehl, 1994; Carrington, 1990) or in the blade
(Blanchette et al., 2002; Blanchette, 1997); hold-
fast/substratum breaks occur less frequently (Milli-
gan and DeWreede, 2000, 2004; but see Duggins et
al., 2003).
The holdfast is crucial in anchoring algae, and
we predicted that the tensile force required to
remove algae from the substratum will increase as
holdfast volume increases. For multicellular algae
attached to rocky substrata, this relationship is
equivocal, as a positive correlation between hold-
fast biomass and tenacity was reported for the
kelp Agarum fimbriatum, but no such correlation
was found for Costaria costata (Duggins et al.,
2003).
Drag on algae increases with water velocity and
blade surface area; in flexible organisms, blade
surface area will change with increasing water velo-
city, and this change is encapsulated by the coeffi-
cient of drag. We predicted that the force required
to remove algae from the substratum increases as
blade surface area increases. Such a relationship
K. Anderson et al. / J. Exp. Mar. Biol. Ecol. 328 (2006) 155–167158
was found for both Agarum fimbriatum and Cost-
aria costata (Duggins et al., 2003). In contrast, we
predicted no correlation of strength with the tensile
force needed to remove a coenocytic individual,
with blade surface area, or with holdfast volume.
Finally, given the consistency of the cylindrical to
bowl shape of the holdfasts in all the coenocytes
we studied, we predicted that these shapes provide
maximum tenacity, compared to other shapes con-
structed from an equal mass of tissue.
2. Materials and methods
2.1. Study area
Our research was done using the facilities of the
Institute for Tropical Ecology and Conservation
(ITEC) in Isla Colon, Bocas del Toro, Panama. Our
study sites were at Lime Point (N 9 24V, W 82 19V) andConch Point (N 9 22V, W 82 17V).
We attempted to measure water motion over the
course of this study, but we were unsuccessful.
However, we judge Lime Point as relatively wave
exposed compared to Conch Point (about 6 km
from Lime Point). Near shore and shallow waters
(b0.50 m) at Lime Point are characterized by
patches of Thalassia testudinum Koenig (turtle
grass), Syringodium filiforme Kuetzing (manatee
grass), and sand. The shallower sandy areas have
Penicillum capitatus and Udotea flabellum, as well
as species of Caulerpa (e.g., C. racemosa (Forsskal)
J. Aghard and C. sertularioides (S.G. Gmelin)
Howe), all species intermixing with the sea grasses.
Sites closer to the reef edge, and deeper (N1 m)
have a greater abundance of Udotea, and less of
Penicillus, and an increased abundance of live coral.
Here, the urchins Diadema antillarum Philippi and
Echinometra lucunter lucunter Linnaeus are first
seen, and the latter increase in abundance over the
reef crest where live coral and greater water depths
(0.5 m to 5 m) occur.
The second study site, Conch Point, is along a
more wave-protected shore lined with mangroves
(Rhizophora mangle). Water depth here ranges
from 0.5 to 1 m at the mangrove edge and
increases to approximately 7 m at some 20 m
from shore. The substratum immediately adjacent
to the mangrove edge is populated with sea grass
(Thalassia testudinum) and green algae, and in
deeper water with various coral species and
sponges. The mangroves are inhabited by small
groups of juvenile fish, and the sea grass beds
and coral clusters are inhabited by small stingrays,
other fish, feather duster worms (e.g., Anamobaea
spp.), brittle stars (Ophiuroidea spp.), various urch-
ins (e.g., Echinometra and Tripneustis), and other
invertebrates. Along the mangroves, sand comprised
60.8% of the substratum, followed by algae
(21.9%), sea grass (8%), coral rubble (4.9%), live
coral (3.5%), and sponge (0.9%). The density of
the algae investigated in this study varied from 161
per square meter (Halimeda gracilis), 1.7 indivi-
duals per square meter for Udotea flabellum, to 0.4
per square meter for Penicillus spp.
The two sites from which we collected data
differed in wave exposure during our stay, with
Lime Point more wave exposed than Conch Point.
That this is a persistent difference is re-enforced by
the observations that Lime Point has a much longer
fetch (reaching to the open waters of the Carib-
bean), lacks the mangroves which border the reef
at Conch Point, and a coarser surface sediment than
at Conch Point. We assume that they can attain, if
rarely, 16–20 m s�1 (Jones and Demetropoulos,
1968, Denny et al., 1985).
2.2. Species tested
We studied four of the most common species of
green coenocytic algae in the shallow subtidal zone
at Bocas del Toro, Udotea flabellum (Ellis et Solan-
der) Lamouroux, Penicillus capitatus Lamarck, P.
pyriformis A. and E.S. Gepp, and Halimeda gracilis
Harvey. All four species are patchy, and Penicillus
spp. and H. gracilis both occur intermixed in places
with U. flabellum. Given this distribution, we
assume that for a given site, these algae are all
exposed to similar water movement. All of the
experimental species tested were not visibly
damaged and non-reproductive; however, some
Halimeda gracilis developed gametangia after
August 1, 2004 (a night with a full moon). At
this time about 2% of the individuals turned white
and showed the bright green gametangia typical of
the species.
K. Anderson et al. / J. Exp. Mar. Biol. Ecol. 328 (2006) 155–167 159
2.3. Species size distribution and holdfast connections
The size class distribution for each species was
obtained by measuring, from substratum to blade
tip, about 100 in situ individuals. To ascertain whether
individuals were connected by underground lateral
rhizoids, we carefully removed 15–20 pairs (within
10–15 cm of each other) from the substratum.
2.4. Biomechanical procedures
Biomechanical procedures were as described in
Collado-Vides et al. (1998). In brief, thalli were
clamped in situ either between wooden blocks
lined with neoprene (and held together with a
clamp), or with the industrial clamp itself. The
blocks or clamps were attached by 16 kg test line
to a spring device (tensiometer—Bell and Denny,
1994; Milligan and DeWreede, 2000), which mea-
sured the force required to detach or break the
thallus. Since most individuals detached whole,
this process was repeated in the laboratory to deter-
mine break location (e.g., blade, stipe/holdfast junc-
tion, etc.) and force-to-break. Strength was
determined by measuring the cross-sectional area
of the break using calipers and the formula for
the closest shape (e.g., an oval or a circle), and
dividing the force-to-break (in newtons) by the
cross-sectional area (m2); the result is given in
mega-newtons (=106 N). Each spring scale used
was calibrated using weights of known mass.
Areas of blades and volumes of holdfasts were
determined using techniques appropriate to their
structure, e.g., the single flat surface of Udotea
blades and the small discs comprising the dbladeTof Halimeda. Blade surface area of Udotea was
estimated by cutting a 4 cm by 4 cm piece from a
Udotea blade, weighing it, and then weighing the
remainder of the blade to determine its (approximate)
surface area. For Penicillus, the relevant surface area
(the projected surface area) was obtained by flatten-
ing the fibrils comprising the blade, tracing the out-
line on a piece of paper and cutting along the
outline. By weighing a 4 cm by 4 cm piece of
paper and then the cut-out paper, the approximate
surface area of each Penicillus blade was calculated.
The surface area of Halimeda was determined by
filling a 4 cm by 4 cm piece of paper with the
individual ddiscs,T weighing the filled paper, and
then weighing the complete blade (comprised of
many discs) and converting this weight to surface
area. Holdfast volume for all species was determined
by measuring water displacement in a graduated
cylinder.
We calculated approximate coefficient of drag
(Cd) values for two species, Halimeda gracilis
and Penicillus capitatus. For Udotea flabellum,
we used previously calculated Cd values (Col-
lado-Vides et al., 1998). We assumed that values
for P. capitatus were similar to those for P. pyr-
iformis, based on the similar size, level of calcifi-
cation, and morphology of these two species.
Values for Cd were determined using a Pesolan0–600 g scale, a PVC pipe to hold the scale, a
connecting fish-line, a lead sinker, and by attaching
the seaweed to the end of the line; this equipment
held the seaweed over the side of the boat and
about 35 cm below the surface of the water. These
tests were done at slack tide, and always in the
same direction, thus minimizing effects of residual
water flow. Following a series of measurements at
known velocities, control values were obtained by
repeating the procedure without the seaweed. Velo-
cities were determined by timing the boat run over
a known distance (250 m) marked on the shore by
flagging tape tied to coconut trees; we were able to
measure drag forces at velocities ranging from 1.1
to 3.3 m s�1.
We indirectly tested the relationship between hold-
fast tenacity and shape by forming wooden blocks
into different holdfast shapes (bowl, short cylinder,
cylinder, and inverted mushroom—the first two are
most similar to the holdfast morphology we encoun-
tered), and measuring their resistance to tensile forces
with and without a coating of the dloopT surface of
Velcro (n) (the Velcro was used to simulate rhizoidal
filaments).
3. Results
3.1. Size class distribution
Size class distribution for each of the species we
tested is given in Fig. 2A–E and indicates that the
maxima differ among species. The most common
A. B.Udotea flabellum: Size Class Distribution
(Lime Point)
0
2
4
6
8
10
12
14
16
18
Above-ground Length (cm)
Per
cent
in S
ize
Cla
ss
Udotea flabellum: Size Class Distribution (ConchPoint)
0
5
10
15
20
25
1 2 3 4 5 6 7 8 9
3 4 5 6 7 8 9
10 11 12 13 14 15 16 171 2 3 4 5 6 7 8 9 10 11 12
1 2 3 4 5 6 7 8 9
1 2 3 4 5 6 7 8 9
10 11 12
13 14 15 16 17
Above-ground Length (cm)
Per
cent
in S
ize
Cla
ss
C. D.Halimeda gracilis: Size Class Distribution (Conch Point)
0
5
10
15
20
25
30
Above-ground Length (cm)
Penicillus capitatus: Size Class Distribution (Lime Point)
0
5
10
15
20
25
30
35
40
1 2Above-ground Length (cm)
Per
cent
in S
ize
Cla
ss
E. Penicillus pyriformes: Size Class Distribution
(Conch Point)
0
5
10
15
20
25
30
35
Above-Ground Length (cm)
Per
cent
in S
ize
Cla
ssP
erce
nt in
Siz
e C
lass
Fig. 2. (A) Size class distribution of Udotea flabellum at Lime Point; (B) size class distribution of Udotea flabellum at Conch Point; (C) size
class distribution of Halimeda gracilis at Conch Point; (D) size class distribution of Penicillus capitatus at Lime Point; (E) size class distribution
of Penicillus capitatus at Conch Point. N =100 for all species.
K. Anderson et al. / J. Exp. Mar. Biol. Ecol. 328 (2006) 155–167160
size class for Udotea flabellum at Lime Pt. is the 8–
8.9 cm class, as it is for Halimeda gracilis. The
other species are generally shorter, with both Peni-
cillus species predominating in the 4–4.9 cm size
class. The absence of the smallest size classes is
real, not an artifact of search techniques.
3.2. Morphological variables
The morphological variables (e.g., blade surface
area and holdfast volume) varied for the different
species (Table 1). Udotea flabellum had the largest
mean blade surface areas and holdfast volumes, and
Penicillus spp. the smallest. Halimeda gracilis dif-
fers from the other species tested in that its dbladeTconsists of a series of calcified, discoid segments,
each disc joined to its neighbor by non-calcified and
adhering filaments.
We found rhizoidal connections for three of our
test species, but not for Halimeda gracilis (Fig. 1);
however, we were only able to find these connections
for two or three individuals of each species (we
excavated 10–15 neighboring pairs for each species).
This absence could indicate that we broke such con-
nections, as they are extremely fragile, or that they
occur infrequently.
3.3. Biomechanical measurements
We found that 100% of Udotea flabellum and
98% of Penicillus spp. individuals tested came out
of the sediment as complete thalli when sufficient
Table 3
Correlations of biomechanically related morphological features for
Udotea flabellum at Lime Pt., Bocas del Toro, Panama
Lime Pt.
Correlation of: With: Significance of correlation
Strength Force to NS p =0.612 r2=0.009
Table 1
Summary of means (with 95% confidence interval) of blade surface
area in square meters (m2) and holdfast volume in milliliters (ml) for
the species tested
Species Site Blade surface
area (m2)
Holdfast
volume (ml)
Udotea flabellum Lime Pt. 0.0049
(0.0017)
6.84 (1.94)
Udotea flabellum Conch Point 0.0023
(0.0034)
4.17 (5.83)
Penicillus capitatus Lime Pt. 0.00031
(0.000072)
0.96 (0.273)
Penicillus pyriforme Conch Point 0.00043
(0.000061)
1.17 (0.144)
Halimeda gracilis Conch Point 0.00261
(0.00058)
2.03 (0.702)
K. Anderson et al. / J. Exp. Mar. Biol. Ecol. 328 (2006) 155–167 161
tensile force was applied. Halimeda gracilis, how-
ever, always broke at the dpads,T immediately adja-
cent to the location where the clamp was applied.
The force needed to break these pads was so small
that we were unable to obtain a reading on our
instruments. However, when the clamp was placed
on the stipe of H. gracilis, the entire plant was
pulled from the substratum.
Results of the biomechanical measurements
(removal force, break force, and strength) are in
Table 2. For the species we tested, Udotea flabel-
lum requires the greatest mean force to remove,
requires the greatest force-to-break, but has inter-
Table 2
Summary of mean removal force, break force, and strength in mega-
newtons per square meter (MN/m2); values in parentheses are 95%
confidence intervals
Species Site Removal
force
(newtons)
Break
force
(newtons)
Strength
(MN/m2)
Udotea flabellum Lime Pt. 12.7 (3.0) 22.1 (3.4) 3.5 (0.8)
Udotea flabellum Conch Point 8.6 (2.8) 22.3 (3.2) 5.7 (1.3)
Udotea flabelluma Puerto
Morelos
8.6 (1.6) 19.3 (2.1) 4.8 (0.6)
Penicillus
capitatus
Lime Pt. 5.0 (0.6) 8.1 (0.9) 7.0 (1.6)
Penicillus
pyriforme
Conch Point 4.9 (0.4) 6.6 (0.9) 4.5 (0.5)
Halimeda gracilis Conch Point 7.1 (1.7)b 12.3 (1.8) 1.0 (0.2)
a Collado-Vides et al., 1998.b Clamp placed at stipe, as individual discs broke at forces too low
to measure.
mediate strength. The values measured for U. fla-
bellum at Bocas del Toro are similar to those
reported by Collado-Vides et al. (1998) for Puerto
Morelos in the Mexican Caribbean (Table 2). Tables
3 and 4(A–D) show that for each species we tested,
the force required to remove it from sediments is
always significantly less than the force required to
break it.
Estimates of the coefficient of drag (Cd) were
obtained at four water velocities, for Penicillus and
Halimeda; the Cd measured at the highest velocity
(3.3 m s�1) was used, in conjunction with surface
area and the force required to pull an individual
from the substratum, to calculate the water velocity
at which sufficient drag was generated to pull it
from the sediments (Table 5). The highest water
velocities so calculated were for the most weakly
attached individuals, of Penicillus capitatus (42.7 m
s�1), and Udotea flabellum (26.1 m s�1). The low-
est water velocity to dislodge was 5.0 m s�1 for
Halimeda gracilis; in fact, this species required the
remove
Blade
surface
area
NS p =0.670 r2=0.007
Force to
break
NS p =0.259 r2=0.045
Holdfast
volume
NS p =0.355 r2=0.349
Holdfast
Volume
Force to
remove
NS p =0.116 r2=0.086
Blade Surface
Area
Force to
remove
S p b0.0001 r2=0.522
Holdfast
volume
S p =0.0006 r2=0.349
Means
significantly
different?
(dtT-test):Force to remove
compared to
force to break
S p b0.0001
NS—not significant at p =0.05; S=significant at a probability level
(=p) of b0.05; r2 is the coefficient of determination.
Table 4
Correlations of biomechanically related morphological features for Udotea, Halimeda, and Penicillus, at Bocas del Toro, Panama
Correlation of: With: Significance of Correlation
A. Conch point: Udotea flabellum
Blade surface area Force to remove S p b0.0001 r2=0.756
Holdfast volume S p b0.0001 r2=0.776
Means significantly different? (dtT-test):Force to remove compared to force to break S p b0.0001
B. Conch point: Halimeda sp.
Blade Surface Area Force to remove S p =0.035 r2=0.280
Holdfast volume NS p =0.954 r2=0.0003
Means significantly different? (dtT-test):Force to remove compared to force to break S p b0.0001
C. Lime point: Penicillus capitatus
Blade surface area Force to remove S p =0.008 r2=0.2000
Holdfast volume NS p =0.463 r2=0.0169
Means significantly different? (dtT-test):Force to Remove compared to Force to break S p =6.13E-8
D. Conch point: Penicillus pyriforme
Blade Surface Area Force to remove NS p =0.209 r2=0.0556
Holdfast volume NS p =0.136 r2=0.0777
Means significantly different? (dtT-test):Force to Remove compared to Force to break S p =0.036
NS—not significant at p =0.05; S—significant at a probability level (=p) of b0.05; r2 is the coefficient of determination. Correlations tested
were the same as in Table 3, but only significantly different comparisons (for any species) are listed below.
K. Anderson et al. / J. Exp. Mar. Biol. Ecol. 328 (2006) 155–167162
lowest water velocity for dislodgment for each of
the three categories in Table 5.
To determine tenacity of different holdfast shapes,
we tested four wooden blocks carved into holdfast
Table 5
Calculation of the water velocity in meters per second (m s�1) req
Species Uf Lime Pt Uf Conch Pt
Minimum RF (N) 3.2 2.4
Cd 0.021 0.021
Area (m2) 0.00044 0.00065
V to remove (m s�1) 26.1 18.5
Mean RF (N) 12.7 11.6
Cd 0.021 0.021
Area (m2) 0.0049 0.0023
V to remove (m s�1) 15.5 21.7
Maximum RF (N) 34.4 27.2
Cd 0.021 0.021
Area (m2) 0.016 0.013
V to Remove (m s�1) 14.4 14.0
Species are Uf—Udotea flabellum; Hg—Halimeda gracilis; Pc—Penici
velocity required to remove an individual is based on field measurements
species from the substratum (minimum, medium, and maximum removal
thallus surface area (area) in square meters (m2 ).
shapes, each with and without a Velcro cover. The
addition of Velcro significantly increased the resis-
tance to tensile forces (Table 6), apparently due to a
better adherence to the sediment particles. Of the four
uired to remove (V to remove) a species from the substratum
Hg Conch Pt Pc Lime Pt. Pp Conch Pt.
1.7 3.1 2.8
0.073 0.12 0.12
0.00043 0.000027 0.000067
10.4 42.7 25.7
7.1 5.0 4.9
0.100 0.12 0.12
0.0026 0.0003 0.0004
7.3 15.8 13.4
12.1 10.0 9.4
0.13 0.12 0.12
0.0071 0.0009 0.0011
5.0 11.8 11.8
llus capitatus; Pp—Penicillus pyriforme. The calculation of water
of the minimum, medium, and maximum force found to remove a
force in newtons (N)), coefficient of drag (Cd) at 3.3 m s� 1 , and
Table 6
Mean removal force (N), and 95% confidence interval (95% C.I.) of
wooden models of holdfasts, with and without Velcro
Shape No Velcro With Velcro
Mean (N) 95% C.I. Mean (N) 95% C.I.
Bowl 142.0 36.6 222.0 41.6
Cylinder 142.0 36.6 176.7 38.0
Mushroom 202.5 40.9 280.0 46.5
Short cylinder 198.0 36.6 290.0 41.6
K. Anderson et al. / J. Exp. Mar. Biol. Ecol. 328 (2006) 155–167 163
shapes, the dmushroom and dshort cylinderT morphol-
ogies had significantly greater resistance to removal
than did either the dbowlT or dcylinderT shape.
4. Discussion
4.1. Detachment and force-to-break.
Almost all individuals of Udotea flabellum and
Penicillus spp. pulled out of the substratum whole
rather than breaking; these results were similar to
those found for U. flabellum in Puerto Morelos
(Collado-Vides et al., 1998), but contrast with the
pattern seen for temperate algae attached to rocks.
For algae that leave tissue on the substratum as a
result of a break (e.g., holdfast portions, holdfast and
stipe, etc.), these dremaindersT provide an opportunity
for regrowth; however, little is known about this
capacity in algae, including the tropical coenocytes.
The relationship between location of a break and
capacity for regrowth provides an interesting area
of future research with important ecological ramifi-
cations for individual survivorship in response to
water motion.
The breaking force of the algae we tested varies
from about 1.8 times greater than the removal force
(Udotea flabellum) to 1.2 times (Penicillus pyri-
forme). As noted above, the exception is Halimeda
gracilis, which broke initially at the dpads,T with
minimal force; however, after clamping below the
pads, these same individuals pulled out completely
from the substratum. In contrast, and as discussed
above, multicellular algae attached to rocks frequently
break within the thallus.
The force required to break a seaweed must be
clearly distinguished from its strength; the former is
measured in newtons, strength in newtons per unit
area. Break force can thus be increased by adding
tissue to a potential break-point or by increasing
strength. Measured break forces for algae vary widely;
for temperate algae, we measured forces ranging from
means of 2.6 N for Mastocarpus papillatus, to 92.6 N
for Egregia menziesii (personal observations). The
coenocytes we report on here had mean break forces
ranging from 6.6 N (Penicillus pyriforme; range: 1.6–
16.9 N) to 22.3 N (Udotea flabellum; range: 8.0–48.8
N). We conclude that the force necessary to break
coenocytes is similar to that of multicellular algae
attached to rocks, and that the coenocytic nature of
the species we tested does not significantly affect this
property.
4.2. Relationship between holdfast volume and
removal force
We predicted that larger holdfasts have a greater
resistance to tensile forces, and we expected a positive
and significant correlation. However, none of the
species showed a significant correlation between
holdfast volume and force-to-remove. In multicellular
algae, this relationship is also equivocal (Duggins et
al., 2003). The absence of such a correlation for algae
anchored in sediments may be because holdfast tena-
city is determined by a combination of factors such as
sediment shear strength, holdfast surface area, and
rhizoid strength. Thus, localized compaction of sedi-
ments may result in a smaller holdfast dplugT but onewith similar tenacity as a larger volume holdfast in
looser sediments; this would obscure relationships
between holdfast volume and tenacity. These factors
are addressed for vascular plants (those with flexible
stems and fibrous roots) by Ennos (1993) and Russell
(1977), but we know of no such data for algae
anchored in sediments.
4.3. Blade surface area and holdfast tenacity
As the blade surface area (=BSA) of algae
increases, so does drag generated by water flow and
we anticipated that attachment tenacity would increase
with BSA. We found a significant correlation between
BSA and the force required to remove an individual
from the substratum for all species except Penicillus
pyriforme. The mechanism behind this correlation is
not known but, except for Udotea, it does not result
K. Anderson et al. / J. Exp. Mar. Biol. Ecol. 328 (2006) 155–167164
from an increase in holdfast volume as blade surface
area increases.
A similar significant and positive correlation
between BSA and attachment tenacity was found for
two temperate subtidal brown algae attached to rocks,
Agarum fimbriatum and Costaria costata (Duggins et
al., 2003). For A. fimbriatum Duggins et al. (2003)
also found a significant and positive relationship
between force to dislodge and holdfast dry mass,
but this was absent for Costaria costata at 3 of the
4 sites they studied. Once again, the question of
mechanism arises for C. costata, as it did above for
the coenocytic sand-dwelling algae. For C. costata, an
increase in detachment force as thallus wet mass
increases does not translate to a positive relationship
between dislodgment force and holdfast dry mass.
What accounts for the increased attachment force? Is
it a tighter adherence due to greater penetration by
holdfast rhizoids into the substratum, or is it due to the
generation of a stronger dglueT with age? Once again,
the biomechanical properties give rise to additional
research questions.
4.4. The relationship between strength and
morphology
None of the species we tested showed a significant
relationship between strength and any of the other
measured variables, hence, we conclude that our
hypothesis (of no correlation between strength and
our other measured variables) is supported. Our
results do indicate that mean strength differs signifi-
cantly among species, ranging from a low of 1 MN
m�2 in Halimeda gracilis to a high of 7 MN m�2 for
Penicillus capitatus. These values are comparable to
those for multicellular species, e.g., 6.7 MN m�2 for
Mastocarpus papillatus (Carrington, 1990), and 0.8–
1.3 MN m�2 for three non-calcified multicellular
algae and also for three non-calcified coenocytes
(Padilla, 1993); for calcified coenocytic and multi-
cellular algae Padilla (1993) found mean values of
1.4–2.9 MN m�2. Even the large kelps from tempe-
rate waters show similar strengths, e.g., 3.5 MN m�2
for Alaria marginata and 1.7 MN m�2 for Egregia
menziesii (DeWreede, personal observations). The
similarity in strength for a wide range of morpholo-
gical types of algae argues for a fundamental con-
straint on altering strength, possibly a function of the
biological materials available for wall composition.
This, despite the fact that temperate algae tend to
break rather than dislodge whole, presumably re-
enforcing selection for greater strength.
Padilla (1985) found that calcified tropical algae (a
coenocyte and two multicellular species) were signif-
icantly stronger than similar non-calcified algae; all of
the algae we tested were calcified. In a biomechanical
context calcification thus acts to strengthen the thallus
against tensile forces, in addition to other possible
functions such as influencing herbivory (Padilla,
1985, 1989; Littler and Littler, 1980). However,
these results raise the question that if greater strength
assures that an individual coenocyte is removed whole
from the substratum, rather than breaking, of what
advantage is this?
4.5. Holdfast form and tenacity
The consistency in holdfast form (a short cylinder
or hemispherical bowl) arising from the rhizoids and
the sediment they entrain suggests that this morphol-
ogy ensures minimal dislodgment, compared to other
possible holdfast shapes. In contrast to available lit-
erature on vascular plant roots (e.g., Ennos, 1993;
Stokes et al., 1996; Mickovski, 2002), we found no
literature addressing the question of doptimal formT forholdfasts of sand-dwelling coenocytes. For macroal-
gae attached to a rocky substratum, Vogel (1988)
concluded that b. . . the common response seems to
be a tapering disk of attachment that is increasingly
flexible toward the periphery—the whole thing dis-
torts a little to avoid stress concentrations at an edge.QAny comparison of algal holdfasts with vascular plant
roots must be undertaken with the caveat that while
algal holdfasts function only in anchorage (but see
Chisholm et al., 1996), vascular plant roots also
absorb nutrients. This latter function may alter root
structure and morphology compared to that if only
biomechanical constraints apply.
Both algae and vascular plant anchoring systems
are subjected to two major forces, vertical tensile
forces (as are generated during grazing) and lateral
forces (as from wind in vascular plants, and moving
water in algae). Ennos (1993) points out that to neu-
tralize lateral loads (vascular) plants are expected to
have at least one rigid element that will resist with its
bending resistance, while the surrounding soil resists
K. Anderson et al. / J. Exp. Mar. Biol. Ecol. 328 (2006) 155–167 165
with its compressive resistance. Vertical forces, how-
ever, are b. . . best resisted by a large number of thin
fibrous roots which have a large surface area over
which tension can be quickly transferred to the soilQ(Stokes et al., 1996; Ennos, 1990). The plug formed in
our coenocytic algae by the adherence of sand and
filaments may serve both functions, resisting tensile
forces by the frictional resistance between the holdfast
plug and the surrounding sand, and lateral forces by
the rigidity of the plug itself.
4.6. Water velocities and detachment
Water movement has been shown to be an impor-
tant physical variable affecting both size and abun-
dance of algae (Denny et al., 1985; Gaylord et al.,
1994). Carrington (1990) concluded that the interplay
of stipe diameter, blade surface area, and water velo-
city determined the maximum size of Mastocarpus
papillatus blades. Similarly, Blanchette (1997) and
Blanchette et al. (2002) showed that water motion
affected the size of Fucus gardneri blades and survi-
val of Egregia menziesii, respectively. Also, Milligan
and DeWreede (2000) found that holdfast attachment
strength was critical for survival of Hedophyllum
sessile, and that only more strongly attached indivi-
duals survived winter storms. However, for Udotea
flabellum, Collado-Vides et al. (1998) showed that
water velocities sufficient to remove this species
would be rarely encountered.
The morphological and biomechanical variables
we measured were used to calculate the water velocity
required to remove species from the substratum. As
discussed, on-site evidence led us to conclude that
Lime Point is more wave exposed than Conch Point.
These observations coincide with the lower mean
removal forces found for Udotea flabellum at Conch
Point and for Penicillus pyriforme (Conch Point)
compared to Penicillus capitatus from Lime Point.
Halimeda gracilis is only found at Conch Point and
exhibited the lowest removal force of any of the algae
we tested. However, at Lime Point (Fig. 2A), the size
class distribution of Udotea is shifted to the right
compared to Conch Point (Fig. 2B), suggesting that
blade surface area does not limit these algae.
If the full range of water velocities suggested by
Denny et al. (1985) and Gaylord (1999) occurs at our
study sites, none of the species we tested are immune
from the possibility of dislodgment. Based on our
calculations, all four species were consistent in that
individuals with the largest blade surface area required
the lowest water velocity for removal. For Halimeda
gracilis, all three categories (Table 5) are predicted to
be dislodged at velocities from 5 to 11 m s�1. Since
H. gracilis was abundant at Conch Point, we conclude
that water velocities exceeding 11 m s�1 had not
occurred at this location since the establishment of
this population, but that the higher wave impacts
observed at Lime Point may contribute to the absence
of this species at this site.
The importance of water movement in structuring
these populations of algae cannot be answered in full.
The smaller individuals of each species required velo-
cities of at least 10 m s�1 (Halimeda gracilis), and the
other species in excess of 18 m s�1, for dislodgment.
In Bocas del Toro higher winds occur in January–
February, with a brief increase in storms during July.
If reproduction in these species is limited to a parti-
cular time, then it is possible that the juveniles occur
during the season in which storms are most likely, and
that these grow to adults in the calmer weather. For
Panama, time specific, and synchronous, reproduction
has been reported by Clifton (1997) for each of the
genera studied by us; the most frequent reproductive
activity occurred from April to June, but varied within
this time frame for a specific species. We found no
juveniles in our July (2004) sampling, except for a
small percentage (b3%) in the population of Penicil-
lus pyriformes. In the absence of growth rate data on
any of these species, or of longevity, the importance of
waves in eliminating the larger size classes is
unknown. However, it can be concluded that for all
but H. gracilis, water velocities necessary to dislodge
the individuals we encountered (those that are left?)
are highly unlikely to occur.
4.7. Dislodgment and survival
For all of the species we tested, individuals were
pulled from the sand with less force than is required to
break that same individual. Is it advantageous to these
individuals that they retain their morphological integ-
rity even though they loose their attachment? With
one exception, we know of no evidence that any of the
tropical sand-dwelling coenocytic algae are able to re-
attach after removal from the substratum. The excep-
K. Anderson et al. / J. Exp. Mar. Biol. Ecol. 328 (2006) 155–167166
tion is among species of Halimeda (Walters et al.,
2002) where field and laboratory evidence points to
re-attachment and subsequent growth of vegetative
fragments, e.g., individual discs. We suggest that the
small force required to detach Halimeda gracilis frag-
ments contributes to this phenomenon. As was con-
cluded by Collado-Vides et al. (1998) for Udotea
flabellum, removal of H. gracilis from the substratum
results in its death, as re-attachment is unlikely. We
propose that the same applies to individuals of Peni-
cillus spp.
However, it appears that death of an individual
may not result in the death of the genet. As we use
the term here, an dindividualT can be either a ramet
(if individuals are connected via rhizoids), or the
entire genet (if there are no connections). We looked
for and found rhizoidal connections between indivi-
duals (of the same species) of Penicillus capitatus,
P. pyriforme, and Udotea flabellum, but these con-
nections were not found for Halimeda gracilis. Lit-
tler and Littler (2000) suggest that these rhizoids
function to proliferate ramets of a genet and may
also give rise to a new individual after dislodgment
of an earlier one. In this case, the small force
required to detach individuals of Halimeda, and
the absence of regenerating dstolons,T is offset by
the vegetative reproduction made possible by the
reattachment and subsequent growth of fragments.
Penicillus and Udotea apparently lack the ability to
regenerate from fragments, but require a greater
force for detachment than does Halimeda, and may
be able to regenerate at least one individual from the
dstolonT left behind.
5. Conclusions
Coenocytic construction is unusual in the algae,
but it occurs relatively more frequently in green algae
found in shallow subtidal sediments associated with
coral reefs. The combination of this relatively unique
construction and poor anchorage in sediments led us
to investigate their attachment and other biomechani-
cal properties. We compared the biomechanical prop-
erties we found to those of non-coenocytic temperate
algae already in the published literature. Our results
show that properties of coenocytes, such as force to
dislodge, force to break, and strength, are all within
the range reported for temperate, multicellular, algae.
In contrast, the sand-dwelling coenocytes differed
markedly from the latter in the consequences of
applied tensile force: coenocytes were removed
whole, while most temperate algae attached to rocks
break within the thallus. Thus, it appears that loosing
tissue to survive hydrodynamic forces is not an option
for species of Udotea and Penicillus. Some temperate
algae can also regrow from the tissue left on the
substratum, but whether coenocytes, in situ, can
regrow from remnant underground rhizoids is not
known. Nevertheless, three of the four species we
examined had such rhizoids, and the one that did
not (Halimeda gracilis) is capable of growing from
detached thallus fragments. What has not been tested
is the attachment and breaking force of coenocytes
attached to solid surfaces such as coral.
Since dislodgment is usually fatal for algae, the
role of the holdfast is a critical one. All of the species
we investigated had similar holdfast morphology, a
mass of rhizoids which entrained sand, the entire unit
forming a hemispherical to cylindrical mass. Despite
the consistency in holdfast form, and the initial pre-
diction that this was an optimal form for anchoring
these algae, our preliminary data suggest this is not
the case. Our data suggest that a longer cylinder
(longer than we commonly encountered in these
algae) provides more resistance to tensile forces.
The problems of optimal holdfast morphology for
sand-dwelling algae, and how this relates to both
sediment coherence and drag forces encountered, are
worthy of a detailed investigation.
Acknowledgements
A portion of this study was supported by a Dis-
covery Grant (NSERC 589872) to R. DeWreede. We
also thank the staff and director of ITEC for use of a
boat and laboratory facilities. [AU]
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