c
Forte (2)
ABSTRACT
Measurement of the physiological effects of the red algal
epiphytes Smithora naiadum (Anders.) Hollenberg and Melobesia
mediocris (Fosl.) Setch. and Mason on Phyllospadix torreyi Wats.
were made near Hopkins Marine Station, Pacific Grove, California,
Field studies revealed a significant influence of these epiphytes
on both the breakage incidence and length, with that of Melobesia
being the most pronounced. Analysis of the photosynthetic rate
of this seagrass shows a decrease in the photosynthetic maxima
in both epiphytized samples at a light saturating intensity.
Under light limiting conditions, an increase in photosynthetic
efficiency and a change in chlorophyll a composition in both
epiphytized sample types suggest an adaptive mechanism similiar
to those found in terrestrial and aquatic shade plants.
Forte (3)
INTRODUCTION
The surfgrass Phyllospadix torreyi Wats. grows attached to
rocks or to rocky substrata ranging from the lower part of the
intertidal to a depth of around 30m. It is found as far North as
Oregon and continues South to Baja, California (Munz and Keck 1959).
Strong rhizomes enable the plant to grow in well-aerated, surf-
beaten areas where a rich supply of nutrients are available for
gregarious growth.
The seagrass itself, however, is only one component of the
primary producers in the Phyllospadix commumity. Epiphytic
algae, including Smithora naiadum (Anders.) Hollenberg and
Melobesia mediocris (Fosl.) Setch. and Mason, at maximum develop-
ment amount to as much as 50% of the total leaf-plus-epiphyte
biomass (McRoy and McMillan 1977).
The smooth lamina of P. torreyi provides both a suitable
substrate for the basal cushion of Smithora, and sufficient
surface area to support the calcareous alga Melobesia. Because
Phyllospadix extends high into the water column, both of these
epiphytic red algae receive light of high intensity and nutrients
from the water motion which are essential for growth.
The physiological effects of the epiphytes on Phyllospadix,
however, are not well known. Some work (Harlin 1975) has been
done on P. scouleri Hook. Previous work by Sand-Jensen (1977)
has shown a decrease in the photosynthetic rate of Zostera marina L.
by 31% in the presence of encrusting diatoms, Cocconeis
scuttellum Ehr.
This paper examines the symbiotic relationship between
P. torreyi and two of its epiphytes, Smithora naiadum and Melobesia
mediocris. Specifically, the effect of these epiphytes on the
Forte (4)
chlorophyll content, photosynthetic rate, and maximum load under
field conditions of this surfgrass are examined.
FIELD STUDIES
Materials and Methods
The first part of this project was to determine the maximum
epiphytic load the individual blades of Phyllospadix can with-
stand before breaking. Choosing the intertidal region between
Seal Rock and Agassiz Beach near Hopkins Marine Station as the
sampling site, I marked blades, 50cm in length (+ 2cm) with
color-coded wires. Twenty samples of the oldest lamina from the
new sheath in each of three categories were tagged: 1) non¬
epiphytized Phyllospadix, 2) Phyllospadix with a dense covering
of Melobesia, and 3) of Smithora. After one 25h tidal cycle,
the samples were harvested and examined under a dissecting micro¬
scope for signs of recent breakage. Any breakage, measured in
length, constituted the category "% broken" in Table 1. An
open wound, irregular fibers at the tip, and fresh green color
at the tip were used as indicators.
Results
When compared to the control group of non-epiphytized blades,
2.5 times more blades with Smithora and 115 times more blades with
Melobesia broke (Table 1). Pieces which broke from samples with
Smithora and with Melobesia were 4.3 and 2.5 times longer,
respectively, than pieces which broke from the control group
(p«.06 for both).
Water motion during this 25h tidal cycle was measured with
clod cards (Doty 1971). A diffusion index of .179 was observed.
Forte (5)
Discussion
From these results it is shown that no linear relationship
between epiphytic weight and mean breakage length exists.
Samples with Melobesia have only 15% higher fresh weight on
average than samples without, yet 1.5 times more samples broke,
and the mean breakage length was 2.5 times longer. Samples with
Smithora, on the other hand, have approximately 600% higher
fresh weight than without, but only 2.5 times more samples broke,
and the mean breakage length was only 4 times longer. This
indicates that epiphytic weight is not the only factor determining
the maximum load on Phyllospadix, nor even the most prevalent.
Texture may affect the drag on individual blades and there-
fore influence breakage incidence and length. Smithora, although
having a much larger surface area, is smooth and very flexible!
If presented flat surface to the stream, there would be consider¬
able resistance; if lying parallel to the stream, it would offer
no resistance. On the other hand, the flat, epidermis-hugging
thalli of Melobesia have hard, stoney and rough surface and may
offer considerable resistance to water motion. Another possibil-
ity may be that Melobesia in some way weakens the Phyllospadix
blade making it more susceptible to the water motion charac-
teristic of the environment of Phyllospadix.
The maximum length broken from blades in each of the three
sample types along with the frequency of occurance for this
breakage length are shown in Table 2. For samples with Smithora,
a 20% loss of total blade length in the 25h tidal cycle occurred
three times. Growth rates of epiphytized Phyllospadix as high as
2.5cm per day for the youngest lamina of the new sheath
(Turhollow 1980 - preliminary study) make this finding reasonable.
Forte (6)
Therefore, my results indicate that the presence of Melobesia
and Smithora on P. torreyi does have a significant effect on both
the breakage incidence and length of this surfgrass in its natural
environment.
LAB EXPERIMENTS
Materials and Methods
Laboratory experimentation investigating the physical effects
of the algal epiphytes oncluded measurement of the photosynthetic
rate at saturating and light limiting intensities, and chlorophyll
analysis of P. torreyi.
All samples taken were 8cm long, 46-54cm from the new sheath
on the oldest lamina. Collection took place on cloudy days to
avoid variation in photosynthetic rates due to drastic differences
in light intensity. Samples were kept in aquaria with running
sea water. Fresh weight were taken and photosynthetic rates were
measured within 12 hours of collection as suggested by Drew (1979)
using a Gilson pressure constant differential respirometer. This
procedure has been documented by Umbreit (1972). The side arm
contained approximately 1 ml of standard bicarbonate buffer
solution to keep C0 levels constant throughout the duration of
the experiment. Temperature was set at 15'C, and readings were
taken every 15 min. for 180 min. All rates were calculated in
ul of Oo/cm2. Surface area was chosen to accurately represent
the amount of photosynthetic pigments whichaare located in the
outer epidermis of the Phyllospadix blade. Rates were measured
on all three sample types at a saturating light level of S
256uE/m2/sec, and at a limiting light intensity of 51uE/m/sec.
Evidence that such a reduction of irradiance to 20% of saturating
Forte (7)
levels produces an actual light-limiting condition (Drew 1979).
He investigated the photosynthetic rate of Phyllospadix vs.
irradiance (Figure 1), which I used as a theoretical base.
At 20% of the saturation light level, the relationship between
photosynthetic rate and irradiance is strictly linear, indicating
limiting light conditions.
Pigment analysis was made by extracting chlorophyll with
5% dimethylsulfoxide in methanol. The samples were ground up
with purified sand using a mortar and pestle, and then centrifuged.
Readings were recorded on an ACG #SP2 spectrophotometer (Beckman)
at 650, 665, and 710 nm. The amount of chlorophyll was calculated
using equations provided by Jeffrey and Humphrey (1975).
To investigate the effects of the changes in light quantity
vs. light quality, three aquaria were placed outdoors equipped
with running sea water. Five bare blades with intact rhizomes
collected from the West Beach area were placed in each, and the
individual lamina were kept separate to prevent self-shading.
Only the tops of the aquaria were exposed. A purple cellophane
filter was placed over one tank which allowed passage of wave-
lengths 480nm and 650nm. The light passing through the filter
corresponds to the wavelengths absorbed by chlorophyll a (peaks
at 445 and 665nm). Absorption spectra were run on the filter
as well as on a fresh blade of Smithora for comparison. The
second tank was covered with a double-layer of Nitex screen
corresponding to a neutral density filter. The third tank was
left exposed. In addition to the change in light quality passing
through the purple filter, both the filter and screen decreased
the quantity of light passing through by 75%. After seven days,
the blades were harvested. Chlorophyll was extracted from areas
Forte (8)
0-8, 20-28, and 40-48cm from the meristem.
Results
At saturating light levels, non-epiphytized samples had a
mean photosynthetic rate of 20.2 ul 0/cm/h (Figure 2).
Samples with Melobesia and with Smithora had significantly
lower rates of 16.7((pc.06), and 6.48 (p«.01) respectively.
Under light limiting conditions, however, no significant dir
difference was found. The mean rate for my control group at this
intensity level was 7.61 pl 0/cm/h, while for samples with
Melobesia and Smithora the mean photosynthetic rates were 6.37
and 6.13 ul 0/cm/h respectively.
Total chlorophyll content, measured in ug per unit surface
area, was 26.7 pg/cme for the control group (Figure 3). Samples
with Melobesia and with Smithora had significantly less chlorophyll
with mean values of 15.5 (p4.01) and 20.4 (p2.06) ug/cm respec¬
tively. No significant difference between the chlorophyll a/b
ratios for all three sample types was found. Likewise, the
chlorophyll b content was relatively constant in all three cases,
and the differences found in total chlorophyll content were
reflected in the amount of chlorophyll a between the three sample
types (pg.05 for both).
Analysis of chlorophyll content for the non-epiphytized
blades from the tank experiment showed an increase in the amount
of total chlorophyll with increasing distance from the meristem
in all three cases (Figure 4). Total chlorophyll content was
not significantly different among samples taken from the three
tanks. However, in the 0-8cm region, less total chlorophyll
was found in the blades from tanks covered with the purple filter
and with the Nitex screen. Again, chlorophyll b levels were
Forte (9)
relatively constant in this region, and the differences in total
chlorophyll are reflected in th different amounts of chlorophyll a.
No significant differences were found between the chlorophyll a/b
ratios in samples from all three tanks. These values were in the
same numerical range as the epiphytized and non-epiphytized
samples measured previously.
Discussion
From the differences observed between the photosynthetic
rate at saturating light levels, three different photosynthetic
maxima are apparent: 1) non-epiphytized samples have the highest,
2) samples with Melobesia are intermediate, and 3) those with
Smithora have the lowest maxima. These differences, in addition
to the constant photosynthetic rate between all three sample
types under light limiting conditions, effectively illustrate
that three distinctly different adaptive mechanisms are present.
Analysis of photosynthetic rate per unit of chlorophyll
(Figure 5) reveals that under light limiting conditions both
epiphytized samples are operating more efficiently. Samples
with Melobesia have the least amount of chlorophyll and are
therefore the most efficient. This adaptation is characteristic
of many terrestrial and aquatic shade plants (Bjorkman 1973).
which are able to change their photosynthetic efficiency and/or
chlorophyll content to adapt to changes in light intensity.
On the other hand, the decrease in the photosynthetic rate of the
control samples by 62% when placed under limiting light conditions
is characteristic of sun plants which are not able to adapt to
these environmental changes.
Studies by Wiginton and McMillan (1979) on other seagrasses
Forte (10)
have shown an increase in the a/b ratio with decreasing light
intensities. However, my studies reveal a relatively constant
ratio with respect to light intensities. The changes in total
chlorophyll are reflected in the amount of chlorophyll a found
in each of the three sample types. A list of a/b ratio values
found for other seagrasses have been included for comparative
purposes (Table 3). Of the five species located in tropical
waters, only Halophila decipiens Ostenfeld (Buesa 1975) has
been observed to have a comparable photosynthetic rate with the
non-epiphytized samples in this study. Therefore, other
environmental parameters may affect the chlorophyll a/b ratio
in these tropical seagrasses, and further studies are necessary
before strict comparisons can be made. In addition, Drew (1978)
has found a seasonal variation in the a/b ratios of the seagrasses
Cymodocea nodosa (Ucria) Aschers. and Posidonia oceanica (L.) Delile
with values during the Spring being the highest. This may in part
explain why my values are near the higher end of the range found
for chlorophyll a/b ratios in seagrasses, but seasonal measurements
on P. torreyi have not been done previously.
My observations of a change inhlorophyll a composition with
varying light intensities disputes conclusions by Wiginton and
McMillan (1979). In a study which measured theochlorophydl content
of five seagrasses grown under different light conditions, changes
solely in chlorophyll b levels as an adaption to changes in
irradiance were observed by them.
Changes in total chlorophyll, as well as in chlorophyll a
levels in the tank experiment support my previous observations.
Although similiar changes were not found in either the 20-28 or
Forte (11)
40-48cm regions, the possibility of adaptation throughout the
length of the blade should not be ruled out. The region O-8cm
from the new sheath is the youngest and fastest growing area of
the Phyllospadix blade and perhaps the most sensitive to changes
in light intensity. The relatively short experimentation period
could account for the failure of the more distant segments to
follow similiar adaptive measures. In anynevent, further
experimentation is necessary before any conclusions can be drawn.
In summary, measurement of photosynthetic rates of epiphytized
samples has revealed lower photosynthetic maxima than found for a
control group of bare Phyllospadix blades. At a limiting light
intensity, samples withwaodense covering of Smithora and with
Melobesia are shown to adapt by both increasing photosynthetic
efficiency and changing chlorophyll a composition similiar to
shade plant behavior. This adaptive mechanism is absent in non-
epiphytized samples. Therefore, the presence of one or both of
the red algal epiphytes, Smithora naiadum and Melobesia mediocris,
results in a restructuring of the adaptive mechanisms of this
seagrass, P. torreyi. The degree of the adaptation differs
depending on the type of epiphyte. Factors leading to this ans
adaptation are not obvious, and further investigations including
seasonal variation studies must be done before a full understanding
of this symbiotic association can be obtained.
Forte (12)
ACKNOWLEDGEMENTS
I would like to thank the faculty and students at Hopkins
Marine Station. Special thanks goes to Dr. Robin Barnett for
his helpful advice and suggestions, Celia Smith whose unending
assistance and counseling helped me to appreciate and lovegthe
algae of the sea, and to Dr. Isabella Abbott who gave me the
confidence to stand on my own feet.
Forte (13)
LITERATURE CITED
Buesa, R.J., 1975. Population biomass and metabolic rates of
marine angiosperms on the northwestern Cuban shelf,
Aquat. Bot., 1:11-23.
Bjorkman, 0., 1973. Comparative studies on photosynthesis in
higher plants, pp 1-63, in Giese, A.C. (ed), Photophysiology;
current topics in photobiology and photochemistry, Academic
Press, 269 pp.
Doty, Maxwell S., 1971. Measurement of water movement in reference
to benthic algal growth, Botanica Marina, 14:32-35.
Drew, Edward A., 1978. Factors affecting photosynthesis and its
seasonal variation in the seagrasses Cymodocea nodosa (Ucria)
Aschers. and Posidonia oceanica (L.) Delile in the
Mediterranean, J. exp. Mar. Biol. Ecol., 31:173-194.
Drew, Edward A., 1979. Physiological aspects of primary
production in seagrasses, Aquat. Bot., 7:139-150.
Harlin, M.M., 1975. Epiphyte-host relations in seagrass com-
munities, Aquat. Bot., 1:125-131.
Jeffrey, S.W. and Humphrey, G.F., 1975. New spectrophotometric
equations for determining chlorophylls a, b, ci and c2
in higher plants, algae and natural phytoplankton.
Keast, J.F. and Grant, B.R., 1976. Chlorophyll a:b ratios
in some siphonous green algae in relation to species and
environment, J. Phycol., 12:328-331.
c
Forte (14)
Munz, Phillip A. and Keck, David D., 1959. A California flora,
University of California Press, Berkeley, p 1323.
Sand-Jensen, Kaj, 1977. Effect of epiphytes on ellgrass photo-
synthesis, Aquat. Bot., 3:55-63.
Stirban, M., 1968. Relationship between the assimilatory pigments,
the intensity of chlorophyll flourescence and the level of
the photosynthesis zonesin Zostera marina L., Rev. Roum.
Biol. Botanique, 13:291-295.
Wiginton, John R. and McMillan, Calvin, 1979. Chlorophyll
composition under controlled light conditions as related
to the distribution of seagrasses in Texas and the U.S.
Virgin Islands, Aquat. Bot., 6:171-184.
Forte (15)
FIGURE CAPTIONS
Fig. 1 - Photosynthetic curve of Phyllospadix vs. irradiance
(After Drew, 1979).
Photosynthetic rates measured in lo/m/h at light
Fig. 2
intensities of 256 pE/m2/sec and 51 E/m/sec. Rates
shown are mean values extended over the three hour
period. Standard deviation bars for the bottom
graph have been shifted up for clarity.
Total chlorophyll (top), individual chlorophyll a and
Fig. 3
b levels (middle), and chlorophyll a/b ratios (bottom).
Fig. 4 -
Total chlorophyll (top), individual chlorophyll a and
b levels (middle), and chlorphyll a/b ratios (bottom),
for tank experiment.
Fig. 5 - Total chlorophyll content (top) and photosynthetic rate
per unit of chlorophyll (bottom).
Forte (16)
Phyllospadix
-j
L
100
20
% full sunlight
gure
Forte (17)
3
bare
50
40
30

20
—
50
W/Melobesia
40
30
1
20


50
wiSmithora
40
30
20



L
hours
high intensity
low intensity —
Figure 2
30-
20
10
30
10
51
12 14
N=
12
E.
N=
12 14 14
wiSmithora
Figure 3
N=
.
ILIL
14
MIIIT bare
total chlorophyll
chlorophyll a
chlorophyll b L

alb ratio
wMelobesia
Forte (18)
Forte (19)
30
20
30r
20
10
3
2
filter
F
2


f

ms
„"

„*
O-8
E nopen

...
E
..
. .


20-28
distance from meristem (cm)
Figure 4
Escreen
total chlorophyll




chlorophyll a H
chlorophyll bE
alb ratio
I

.
40-48
Forte (20)
30
20
10
4
total chlorophyll
.

low intensity
high intensity
w/Smithora II bare w/Melobesia
Figure 5
Forte (21)
5
S
s



—
5
15
E
0

29

2


3
11
9
O
Forte (22)
.
2


3—
8
o

18


E

0
10

O

3
Forte (23)
Table 3. Ratios of chlorophyll a to chlorophyll b in 8
seagrass species compared to ratios observed in this
study on Phyllospadix torreyi.
alb ratio.
species
Heterozostera
2.9
(Keast and Grant 1976)
Halophila decipiens
1.48-1.85
(Wiginton and meMillan 1979)
Halodule wright
2.08-2.26
(Wiginton and McMillan 1979)
Syringodium filiforme
1.85-2.03
(Wiginton and Memillan 1979)
Thalassia testudinum
2.38-2.44
(Wiginton and Memillan 1979)
Zostera marina L.
2.81
(Stirban 1968)
Posidonia oceanica
(summer)
2.
(Drew 1978)
3.2
(spring)
Cymodocea nodosa
1.6
(summer)
4.0
(Drew 1978)
(spring)
Phyllospadix torreyi
(control)
3.72
control
3.72
with Melobesis
mediocris
3.32
3.19
with Smithora naiadum