Gewhimica a Cosmochimica Acta Vol. 57. pp. 391 l-3984 0016-7037/93/$6.00f .

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Copyright @ 1993 Pergamon Press Ltd. Printed in U.S.A.

Bi~~hemi~l cycles of carbon, sulfur, and free oxygen in a microbial mat

DONALD E. CANFIELD*and DAVID J. DES MAWS
Ames Research Center, MoffettField, CA 94035, USA

(Received February I I, 1992; accepted in revisedform March 19, 1993)

Abstract-Complete budgets for carbon and oxygen have been constructed for cyanobacterial mats dom-
inated by Microcoleus chthonoplustes from the evaporating ponds of a salt works located in Guerrero
Negro, Baja California Sur, Mexico. Included in the budget are measured rates of O2 production, sulfate
reduction, and elemental exchange across the mat/brine interface, day and night, at various temperatures
and times of the year. We infer from this data the various sinks for Ot, as well as the sources of carbon
for primary production. To summarize, although seasonal variability exists, a major percentage of the
Oz produced during the day did not diffuse out of the mat but was used within the mat to oxidize both
organic carbon and the sulfide produced by sulfate reduction. At night, most of the O2 that diffused into
the mat was used to oxidize sulfide, with Or respiration of minor importance.
During the day, the internal mat processes of sulfate reduction and 02 respiration generated as much
or more inorganic carbon (DIG) for primary production as diffusion into the mat. Also, oxygenic pho-
tosynthesis was the most important process of carbon fixation, althou~ anoxygenic photosynth~is may
have been important at low light levels during some times of the year. At night, the DIC lost from the
mat was mostly from sulfate reduction. Elemental fluxes across the mat/brine interface indicated that
carbon with an oxidation state of greater than zero was taken up by the mat during the day and liberated
from the mat at night. Overall, carbon with an average oxidation state of near zero accumulated in
the mat.
Both carbon fixation and carbon oxidation rates varied with temperature by a similar amount. These
mats are thus closely coupled systems where rapid rates of photosynthesis both require and fuel rapid
rates of heterotroph~~ carbon oxidation.

INTRODUCTION CANFIELDand DES MARAIS, 1991). Numerous measure-

ments can be made over a single die1 cycle, and, thus, mi-
~ANOBA~E~IA,A~OMPANIED bynumerousother~~e~
crobial mats offer a unique opportunity to understand the
flourish in mat communities in many coastal environments,
cycling of carbon, sulfur, and oxygen to a level not earily
lakes, and thermal springs. These mats represent complex
attained in other ecosystems. Our approach is to quantify
ecosystems complete with photoautotrophic, photohetero-
rates of both net (using flux chambers) and gross processes
trophic, chemautotrophic and heterotrophic populations.
(measured directly) of carbon transformations over a die1
Mats contain most, if not all, of the microbial elements and
cycle, at different times of the year where natural variability
biogeochemicai processes that exist in any aquatic ecosystem.
in carbon processing is encountered.
in mats, however, microbial and chemical zonations occur
A prime motivation in studying mats is to understand car-
on millimeter to sub-millimeter depth scales. For example,
bon cycling as it relates to sedimentary (texture, laminations,
primary carbon production by photosynthesis can occur to
etc.) and geochemical (carbon concentration, carbon and
between 0.4-3 mm depth in a mat, with subsequent oxygen
sulfur isotopes, etc.) features that would be preserved poten-
penetration to between 0.5-S mm (see RIWZWK et al.,
tially in a lithified mat. Ancient lithified mats occur as stro-
1983). Below the zone of oxygen penetration, sulfide from
matolites ranging in age from nearly 3.5 Ga to modern. Our
dissimilatory sulfate reduction will accumulate; and at the
expectation is that studies of modern mats will help us to
oxygen/sulfide interface, over a depth range of perhaps 0.2-
interpret ancient stromatolites for evidence of environmental
0.3 mm, a diverse population of sulfide oxidizing organisms
changes such as in ocean and atmospheric chemistry, and
are found (JORGENSENand DES MARAIS, 1986 ).
also biological evolution.
lnv~ti~tions of mat ecosystems are in some ways fairly
straightforward because the photosynthetic community is
physically attached to the underlying heterotrophic com- STUDY SITE
munity. The quite small dimensions associated with the mi-
We~l~evelo~ ( 3-8 cm thick) cy~no~~te~al mats (see
crobial populations pose a special challenge, but new tech-
D’AMELIO et al., 1989, for description) are found in the
nology has emerged which allows the rates of important mi-
evaporating ponds of the Exportadom de Sal salt works, lo-
crobial processes, like primary production and sulfate
cated in Guerrero Negro, Baja California Sur, Mexico. We
reduction, to be made with the needed spatial resolution
studied mats in Pond 5 that were permanently submerged
( REVSBECHand J~RCENSEN, 198 1; REVSBECHet al., 1983;
under about 70 cm of evaporation-concentrated seawater,
whose salinity may vary between about 90- 108L during the
* Present address: School of Earth and Atmospheric Sciences, year. Over the time =scaIeof a field trip (two weeks), however,
Georgia Institute of Technology, Atlanta, GA 30332-0340, USA. salinity remained nearly constant. We have no continuous
3971
3912 D. E. Canfield and D. J. Des Marais
annual temperature record for Pond 5, but from our expe-
rience, pond water temperatures may vary up to 8- 10°C
during the course of a die1 cycle and annually between about
12°C (early morning in late winter and early spring) and
30°C (mid-afternoon in summer and early fall). At our study
site, which encompassed an area of about 600 m*, the mats
were composed almost completely of organic matter, with
only a very minor mineral component. Also, organic matter
deposition was mostly autochthonous.
We have studied Pond 5 mats over the past several years
and report here on the results from field trips during Novem-
her, 1989 (Nov. 89), April, 1990, and November, 1990 (Nov.
90). In general, during all of our trips to Pond 5, we have
found the mats to be dominated by the filamentous, bundle-
forming, cyanobacterium Microcoleus chthonoplastes (see
D'AMELIO et al., 1989; DES MARAIS, 1990), with lesser
numbers of ~~e~ii~or~aspp. and Spi~li~a ~a~~~thl~or~i~
(also a filamentous cyanobacterium) and the unicellular cy-
anobacterium Synechococcus sp. At the mat surface, we have
generally observed a thin layer of the diatoms Nitzschia and
~a~i~ui~ spp. We have also noted some variability in the
surface texture of the mats. With reference to our three sam-
pling times, during both Nov. 89 and Nov. 90, mat topog-
raphy was much as described by J~~RGENSEN and DES MAUAIS
( 1990), with approximately 0.5 mm “hills” spaced at 2-3
mm intervals (features resembling the larger “ridge” described
by these authors could be found but were not sampled by
us). By contrast, in April 90, the mat was very smooth, with
little or no surface relief.
We also note other important ditferences in the mats which
d~tinguish the two November trips from the April trip. Dur-
ing Nov. 89 and Nov. 90, the daytime OZ/HZS interface,
typically found between 1-2 mm depth in the mats (JAR-
GENSENand DES MARAIS, 1986), was heavily populated by
the colorless sulfur oxidizing bacterium Beggiatoa spp., with
low numbers of the green filamentous bacterium Chloroflexus
sp. During April 90, Beggiatoa was virtually absent, and the
Oz/H2S interface was dominated by a dense layer of Chlo-
roflexus. Whereas Beggiatoa is a chemautotrophic organism
gaining energy from the oxidation of sulfide with 0~ (NESON
and JANNASCH, 1983), Chloro~ex~ may oxidize sulfide as
a phototroph using photosystem I (PIERSONand CASTEN-
HOLZ, 1974). These differences in species composition are
stressed as they reflect major differences in the mat environ-
ment and the cycling of chemical species within the mats
during the different trips. This issue will be a major focus of
what follows.
CARBON AND 02 I’RJDGET ~ONSIDE~~ONS
In general, the main pathways and processes of carbon
and O2 cycling in cyanobacterial mats have been identified
(for example, REVSBECHet al., 1983; SKYRING et al., 1983);
these are summarized in Fig. 1. During the day, light strikes
the mat surface, fueling primary carbon fixation by both ox-
ygenic and anox~enic photo~phs. Some secondary carbon
fixation will occur with the growth of nonphotosynthetic au-
totrophic and heterotrophic bacteria. The sources of inorganic
carbon (DIC) for primary production include diffusion from
the overlying water, DIG derived from both O2 and anaerobic
respiration within the photic zone of the mat (see CANFIELD
and DES MARAIS, 199 1)) and DIC from heterotrophic activity
deeper in the mat. Some dissolved organic carbon (DOC),
obtained from either the water column or within the mat,
may also be incorporated into growing biomass (not shown
in Fig. 1 ), or conversely, may be lost from the mat by dif-
fusion.
The only source of 02 within the mat during the day is
primary production by oxygenic photosynthesis. The amount
of Oz produced by this process depends on the rate of carbon
fixation, the oxidation state of the carbon fixed, as well as
the amount of nitrate that must be reduced to ammonia in
forming organic material. Sinks for 02 include both diffusion
out of the mat into the overlying brine and deeper into the
mat to oxidize reduced chemical species such as sulfide and
ammonia. Some 9 will also be used to oxidize both organic
matter within the aerobic zone (02 respiration), as well as
any sulfide produced in the aerobic zone by sulfate reduction
(CANFIELD and DES MARAIS, 199 1; FROND and COHEN,
1992).
At night, the oxidation of organic matter produces DIC
which diffises out of the mat into the overlying water (Fig.
1). Carbon oxidation can occur both by aerobic ( O2 respi-
ration) and anaerobic pathways (sulfate reduction is the most
im~~nt). Again, as during the day, some DIC will also be
used in the growth of nonphotosynthetic bacteria. Also, as
during the day, DOC may be cycled within the mat and/or
exchanged across the mat-brine interface. Oxygen is con-
sumed by the mat at night and is used to oxidize organic
matter, sulfide, and possibly also some ammonia.
Our experiments were designed to constrain the rates of
as many of these processes as possible, although there are
some that we will not consider. We will not, for example,
include nonphototrophic bacterial carbon production in our
budgets. This omission will likely cause only a small error
because much less carbon is fixed by these secondary pro-
cesses. Also, if the population size of the nonphotosynthetic
bacterial community does not change much during a die1
cycle, there is little or no et&ct on the budgets.
Preliminary porewater and flux chamber results (D. J. DES
MARA~Sand D. E. CANFIELD, unpubl. data) indicate that
DOC fluxes into and out of the mats am very small compared
to DIC fluxes, so that Mx: may not be a large net contributor
to the carbon budget. We will not consider DOC in our bud-
gets. We will not include the cycling of nitrogen in our bud-
gets, and this is perhaps our most serious omission. We add,
however, that although ammonia oxidation may be an im-
portant 02-consuming process, we are unsure of the extent
to which ammonia is oxidized in the mats vs. direct incor-
poration into growing biomass. Also, we are not sure of the
extent to which nitrate is reduced during photosynthesis; the
amount may in fact be small because the pond waters in
which these mats grow are very depleted in nitrate ( JAVOR,
1983). We have also assumed that denitrification is a rela-
tively minor carbon oxidation process, as in most sediments
overlain by O&ch water (J~~RGENSEN,1983; CANF~ELD,
1993). As more information on the nitrogen cycle becomes
available, we will adjust our budgets accordingly.
METHODS
In a typicai experiment, several pieces of fresh mat, collected from
about a 20 m2 region in the pond, were transported to 35 L aquaria
 Biogeochemistry of C, S, and O2 in a microbial mat 3913

DAY

Msteurleoe OIC I 4

I
I 02
I respiration
I
I \
I 02 u Primary
I
I prod-n
J
i bpZ20n 1
I Sulfide
I oxidation
I
I
reduction
; ,,,, //, ,,,,, ;;:\t1;;;;,L((,,,4

///~////////~//~~///, Sulttde oxidation

I\ sulfete
nduction
I
I

II
HP-5
I

NIGHT
DC
Met sulfeoe
02

-th

sunste I

i
IZ
“2s
reduction I
I
I
I

FIG. 1. Cartoon depicting the principles of carbon and Or cycling in the cyanobacterial mats of Pond 5. During the
day, Or production from oxygenic photosynthesis is distributed to some depth in the mat. This Oz may diffuse from
the mat, diffuse into the region of the Oz/HzS interface to oxidize sulfide, or be used within the aerobic zone for Oz
respiration or to oxidize any sulfide produced by sulfate reduction in this zone. DIC is used by both oxygenic and
anoxygenic phototrophs in primary carbon production. The source of the DIC includes diffusion from the overlying
brine, diffusion from below the Or/HzS interface (DIG is liberated in this region by sulfate reduction), and DIC
liberated in the oxic zone by Or respiration and sulfate reduction. At night, Or diffises into the mat and is used to
oxidize organic carbon (Or respiration) and the sulfide produced by sulfate reduction. DIC is liberated by both sulfate
reduction and Or respiration and diffuses out of the mat into the overlying brine. Arrows depict net fluxes of DIC (left
side of figure) and 02 (right side).

and maintained at a constant temperature. (between 17-30°C) in
fresh pond water for a preincubation period of 24 h. Mats with min-
imal surface relief (~3 mm) were chosen. Constant temperature was
maintained to within + 1“C by the use of a combined circulator/
heater/cooler which ran tap water through aluminum coils inserted
into the aquaria. After this preincubation period, tank water was
replaced by newly collected pond water, and experiments were begun
with continuous temperature regulation.
Flux Measurements
In one set of experiments, all-glass flux chambers (Fig. 2) were
placed over the mats, beginning at sunrise. The Pyrex glass allowed
unimpeded light penetration to the mat surface, with no appreciable
heating (<I “C) of the chamber. Constant stirring was provided by
an ail-glass stir paddle which revolved at a constant rate of twelve
revolutions per minute. The chambers covered -0.02 m2 of mat
surface, and chamber volumes were varied depending on the antic-
ipated activity of the mat; taller chambers were deployed over more
active mats. A fine-mesh screen was placed over the aquaria to de-
crease the ambient light levels by 40%; this mimicked the light in-
tensity incident on the mats at 70 cm water depth in the ponds.
Mats were incubated under these conditions until noon, when the
flux chambers were sampled for dissolved Or and dissolved total
DIC. Dissolved Or was measured with a microelectrode, and water
for DIC analysis was added to evacuated glass bottles. DIC samples
were acidified in the lab at’NASA-Ames, liberating COr , which was
measured manometrically. The relative standard deviation of the Or
determination was about +3W, and 25% for the CO1 measurement.
Oz-rich gas that had accumulated at the surface of the chamber was
collected and measured for volume and composition. Volumes could
be read to +2%, and replicate flux experiments (for Or and CO,)
generally agreed to within +%I. After the noon sampling, chambers
were redeployed with fresh tank water (to ensure that carbon
3914 D. E. Canfield and D. J. Des Marais
FIG. 2. Schematic diagram of benthic chamber used for mat studies.
Letters designate the foIlowing: (A) chamber made from the base of
a 4 L Pyrex beaker;(B) Pyrex glass paddles and shaft; (C) red rubber
tubing which couples Pyrex stirrer to 12 rpm D.C. gearmotor; (D)
fabricated Teflon shaft seal threaded on the outside to permit at-
tachment to chamber using nylon nut; (E) septum assembly (piece
exposed to chamber interior is fabricated from Teflon and attached
to chamber in same way as part D; silicone septum is held in place
by Swagelok nut); (F) microbial mat.
limitation would not occur during the incubations) and collected
again at sunset. After sunset, the chambers were redeployed with
added O2 so as to not go anoxic during the night. The chambers were
sampled and redeployed at midnight and collected again just before
sunrise. Redeployments required about 30 min.
Oxygen Production and Distribution
Rates of O2 production and O2 depth profiles were determined
throughout the day on mat pieces under conditions of light and tem-
perature identical to those for the flux measurements. Mat pieces
were contained in 7.5 cm diameter core tubes and permanently sub-
merged in a large reservoir (35 L) of fresh pond water. The water
over the mat was constantly agitated with an aquarium bubbler. 02
production rates were measured using Clark-type O2 microsensors
(tip diameter IO-30 pm with minimal stirring sensitivity; REVSBECH
and WARD, 1983) and the light/dark shift technique (REVSBECH
and J~~RRGENSEN, 1981; REVSBECH et al., 1983), where rates of O2
production are assumed to equal rates of Or decrease immediately
after a steady-state mat is placed in darkness.
This method has been treated theoretically (REVSBECHet al., 1986;
GLUD et al., 1992) and provides a good determination of depth in-
tegrated O2 production rates. The method includes 02 used in pho-
torespiration but not in the Mehler reaction (GLUD et al., 1992). In
our experiments, the mat surface was darkened by either shading the
mat with black cardboard, which reduced the ambient light levels by
85% (Nov. 89 experiments), or by covering the aquarium with a
black cloth, causing complete darkening of the mat surface (subse-
quent experiments). The mat surface was left darkened for no more
than 4 set, and the mat was allowed to reattain steady state before a
new measurement was made. The profile of 02 in the mat was ob-
tained simultaneously with the productivity measurements. The
measurement of Oz concentration is very accurate (? l-2%), and
our ability to resolve an Oa profile is limited mostly by our ability to
position accurately the microelectrode with depth. This could be ac-
complished with a precision of at least ?5%. Duplicate determinations
of photosynthetic rate at a given depth were generally reproducible
to better than +5%.
Or production was measured at random spots in the mat, with the
mat/brine interface identified as the first interval where, on darkening,
an 02 decrease was observed without a time lag (see REVSBECHet
al., 1983; GLUD et al., 1992). The electrode was calibrated for zero
O2 (at depth in the mat) and air saturation after each productivity
profile. The saturation concentration of O2 in the pond water was
determined by Winkler titration ( GRASSHOFF,1976). By choosing
random spots, we integrated any spatial variability in the activity of
the photosynthetic community. We typically observed photosynthetic
rate at a given light level to vary by about + 15-20% at different places
in the mat. This may be taken as a measure of the small-scale ( mm2)
variability in the activity of the photosynthetic population. Large-
scale (several cm2) variability in these mats was less, as evidenced
by the good reproducibility of the flux chamber results (see the pre-
ceding text).
Sulfate Reduction
Rates of sulfate reduction were determined at noon and midnight,
at the same temperature and light conditions as for the mats described
here in the preceding section. In some cases, the same mat piece was
used for both microeksctrode and suifate reduction rate measurements.
In other cases, sulfate reduction rates were determined on mats col-
lected from directly adjacent to the mat piece used for microelectrode
work.
To measure rates, cores were injected vertically, in triplicate, with
radiolabeled sulfate (“S) made to approximately 100960salinity in
NaCI. Each injection was about 1.5 MBq, with about 2-3 mm of
brine let? covering the mat (some label was also added to this brine
so that label would not diBi_tseout of the mat). After injection, a
Pyrex beaker was inverted over the mat, and the core plus beaker
were immersed into a constant temperature bath, with sunlight al-
lowed to strike the mat surface. The cores were incubated in this
manner for 20-30 min and then sampled with cut-off sharpened
plastic syringe barrels, which were quickly frozen (less than 5 min)
over dry ice. In time series experiments, similar rates of sulfate re-
duction were found for incubation times ranging from between 10
min to 1 h. Times outside of this interval were not explored. We
could not monitor O2 production and distribution during these in-
cubations; but in separate experiments with similar brine coverage
(2-3 mm), O2 distribution and production rates remained relatively
constant over the time scales of our sulfate reduction rate measure-
ments. The standard deviation of depth-integmted sulfate reduction
rates, from triplicate cores, ranged from fi.5 to +3 l%, with an average
of +I 1.2% (n = 7).
Other Measurements
Mats were collected for elemental sulfur (So) analysis during the
course of a die1 cycle and immediately frozen. Elemental sulfur was
removed from the mat by soxhlet extraction with acetone and pre-
cipitated onto Cu metal wire ( BERNER, 1964). The Cu plus Cu-
sulfide was collected on a filter and digested in an acidic chromous
chloride solution. The liberated sulfur was precipitated as A&S. Total
reduced sulfur in the mat was also liberated with a boiling chromous
chloride solution (CANFIELD et al., 1986) and collected as Ag2S.
Concentrations were determined by weighing the AgzS.
RESULTS AND DISCUSSION
Flux Chamber Results
The results of the flux experiments are given in Table 1
and do not include exchange that occurred while the flux
chambers were being collected and redeployed. During Nov.
89, we did not redeploy the tlux chambers at noon and feel
that carbon limitation during the latter parts of the day se-
verely reduced rates of photosynthesis. For this reason, we
report only nighttime O2 and DIC fluxes for this period.
 Biogeochemistry of C, S, and O2 in a microbial mat 3975

Table1.Measuredcarbooandoxygenfluxes~. appreciable light penetrated to the 0z/H2S interface and was

available for sulfide utilizing phototrophs; we, in fact, ob-
mmolesm-2 served a dense layer of the green sulfur bacterium Chloro-
m !%3Y &&
flexus sp. at this interface in April 90. These observations are
r p1 G Q2 consistent with appreciable anoxygenic photosynthesis during
Nov. 09 2oQc n.d.b n.d 43.8 -26.6 this time. In Nov. 89 and Nov. 90,02 penetrated much deeper
than O2 production. JORGENSEN and DES MARAIS ( 1990)
3ooc nd. n.d. 125 -79
note that when O2 penetrates well below the zone of O2 pro-
April90 1X -34.5 15.3 32.4 -13.9 duction, little or no light reaches the Oz/HrS interface, and
-96 116 -60
this interface will be dominated by nonphotosynthetic sulfur
30% 55
oxidizing bacteria. During Nov. 89 and Nov. 90, we observed
Nov. 90 2093 -69 51 75 -61 a sparse population of Chlorofexus sp., and, instead, the 02/
a Fluxesarecomputed overthetimeintervalof thefluxchamber H2S interface was populated by the colorless sulfur bacterium
deployments. Negativevalueswe for fluxesintothemat,and positive Beggiatoa spp; thus, anoxygenic photosynthesis was unfa-
valuesfor fluxesout of the mat into the overlying brine.
b n.d. = not detemined vorable during these times.
Rates of O2 production, obtained throughout the day in
the different experiments, have been integrated with depth
Sulfate Reduction and Solid Phase Sulfur to yield total rates of O2 production. O2 profiles have been
utilized to calculate the flux of Or into the mat day and night
Rates of sulfate reduction measured in Nov. 89 and Nov.
and out of the mat during the day. The results of these cal-
90,are presented in Fig. 3. The occurrence of high rates of
culations are compared in Fig. 6 with the light incident onto
sulfate reduction in the aerobic zone of the mats during the
the mat surface during the day. The uncertainty in O2 pro-
day has been considered elsewhere (CANFIELD ,and DES
duction rate measurements has been discussed previously.
MARAIS, 199 1). Rates of sulfate reduction decreased expo-
The error in obtaining 02 fluxes from gradients varies de-
nentially with depth, as might be expected for the decom-
pending on the profile but, on average, is better than + 10%.
position .of initially fresh organic biomass ( WESTRICH and
A bigger uncertainty in calculating fluxes is determining the
BERNER, 1984). At 20°C during both Nov. 89 and Nov. 90,
porosity of the mats, which, in turn, controls the sediment
daytime rates of sulfate reduction were higher in the surface
diffusion coefficient (OS). The mats have a water content of
portion of the mat than at night (Fig. 3). FROND and COHEN
90% and, assuming a density of 1.5 g cm-3 for the organic
( 1992) have found a similar stimulation of daytime sulfate
material (the same value as for sugars), a porosity of 0.93 is
reduction in a cyanobacterial mat and have argued that me-
calculated, which yields a D, of 0.86 D r, where D f is the free
tabohzable organic substrate, derived from photosynthesizing
diffusion coefficient (ULLMAN and ALLER, 1982). The flux
autotrophs, could fuel these higher rates. This could explain
can then be calculated as the product of D,, the mat porosity,
our higher daytime rates at 20°C; however, during Nov. 89
and the 02 gradient (Fick’s first law). We use this as an upper
at 30°C near-surface rates of sulfate reduction were similar
limit on the flux. A complication with mats is that both living
both day and night.
and dead cells cause an impedance toward the diffusion of
Depth-integrated rates of sulfate reduction are presented
solutes, but also contain water, such that an artificially high
in Table 2. For April 90, sulfate reduction rates were not
porosity may be calculated from the water content. For a
measured but estimated assuming that all of the nighttime
cyanobacterial mat from the Bay of Arhus, REVSBECH et al.
O2 flux into the mat was used to oxidize the sulfide from
( 1986 ) determined D, to be 70% as great as D fr which would
sulfate reduction to sulfate. Some support for this assumption
correspond to an effective porosity of 0.84, using the rela-
comes from the Nov. 89 and Nov. 90 data, where between
tionships presented in ULLMAN and ALLER ( 1982). We can-
77 and 96% (with an average of 84%) of the measured O2
not be certain as to the similarity between the mats studied
uptake was used to oxidize sulfide (Table 3 ). For April 90,
by REVSBECH et al. ( 1986) and the Baja mats, but we use
the same rates of sulfate reduction were assumed day and
these values as a lower limit on the O2 flux. At night, most
night, based on such a similarity during the other sampling
of the O2 gradient is established within the benthic boundary
times (Table 2).
layer and not in the mat, so we use Df to calculate fluxes.
Solid phase sulfur species over a die1 cycle in April 90 at
Also included in Fig. 6 are the average fluxes of O2 into
30°C are shown in Fig. 4. Elemental sulfur accumulated at
and out of the mat as obtained from the flux chambers, as
the sediment surface during the day and was oxidized at night.
well as the flux of Oz required to oxidize the sulfide produced
Below 2 cm, no significant die1 cycling of solid phase sulfur
by sulfate reduction. These results and their calculation will
was observed.
be discussed in detail below.
Oxygen Microelectrode Results Light and 0, Production Rate
Examples of the depth distributions of dissolved O2 and From Fig. 6, there is a clear correlation between O2 pro-
02 production rate during the different trips are shown in duction rate and light intensity. These two are plotted together
Fig. 5. In April 90, photosynthetic O2 production occurred in Fig. 7; for all of the sampling periods, rates of O2 production
to the full depth of O2 penetration (Fig. Sb). As discussed increased linearly with increasing light intensity, with no ev-
by JORGENSEN and DES MARAIS ( 1986), this signifies that idence for photosaturation or photoinhibition. REVSBECH et
3976 D. E. Canfield and D. J. Des Marais

10

a
90
Nov. 69,20X Nov. 69,30-C
i
g@ 0 1 2 3 4 5 6 ; 10
1 Sulfate red. (pIA min-1) Sulfate red. (PM mln-1)

”
v” 0
fbw ocD
- wo
wo
- 08
0%

0
_ 0

0
-0

Nov. 90,2O’C 0 Nov. 60,3O”C

4oI I I I I I I I
0 4 6 12 16 0 10 20 30
Sulfate red. (PM min-1) Suttate red. (pM min-l)

FIG. 3. Rates of sulfate reduction measured day and night at various times of the year.

Table 2. Integrated rates of sulfate reduction. al. ( 1983) reported a similar correlation for cyanobacterial
mats from Solar Lake (Sinai). It is interesting that even
nmoles a+ min-1
though no photosaturation was observed for the mats at light
ImP !a! m levels up to 800 PE me2 s-l, individual species of cyanobac-
Nov. 69 WC 1.66 (0.36P 1.51 (0.12) teria become photosaturated at light levels of around 50-60
PE m-*s-’ . As shown by STAL et al. ( 1985) and JORGENSEN
3ooc 5.04 (0.026) 4.55 (0.15)
et al. ( 1987), light is considerably attenuated at shallow depths
April90 ET l.olb 1.01" in mats, and the highest rates of photosynthesis are often
304: 4.65b 4.65b found at some depth below the mat surface (REVSBECH et
al., 1983; Fig. 4). Hence, photosaturation may occur in in-
Nov.90 2ow 4.17 (1.28) 3.74 (0.38)
dividual layers in a mat. However, the photic zone generally
3wc 12.7 (0.46) becomes deeper with increasing light ( REVSBECH et al., 1983),
a numbers inparenthesis are one standard deviation
allowing for an overall correlation between light and 02 pro-
b estimated from rates of nighttime Q uptake (see text) duction rate.
 Biogeochemistry of C, S, and Or in a microbial mat 3911

Average Q Flux 9601 used in sulfide

(Nooks an-2 milt-‘) OXi&liOn

Nov.89 20°C 3.5 to.@’ 660

3Ow 11.43 (0.57) 79 (3)

1600
Nov.90 2O“C 6.62 (0.43) 67 (9)

1400
a average 9 flux (from Table 1) is combined with sulfate reduction
rates (Teble 2) to @e the % of the 02 flux into the mat used to oxidii
sulfide to sulfate
b numben, in parenthesisare one standard deviation 1800

During April 90, at 30°C (Fig. 7b), 02 production became

insignificant at light levels less than about 400 PE mm2 s-‘, 0

a light level of about one-half maximum and a level where

considerable O2 production was still observed at other times
of the year. We propose that during April 90, and especially
at ‘higher temperatures (30”(Z), anoxygenic photosynthesis
dominated photosynthetic carbon production at lower light
levels, suppressing oxygenic photosynthesis. This proposal is
consistent, as discussed in the preceding text, with the high
abundance of anoxygenic photosynthetic organisms in April
90. We are, however, unable to explain why this community
was so conspicuous at this time.
(b) April 66,17”C -
C2 Production and Temperature
0 106 206 300 400
Rates of O2 production responded strongly to changes in
temperature. In Nov. 89, O2 production rose by a factor of -200 . .“I . . I

3 when temperature was increased by 10°C. In April, an 0

l 0
increase of 13°C caused O2 production, at maximum light 0 0
0 l
levels, to increase by a factor of 4 (Fig. 7). One factor con- 00
l
tributing to the temperature response of 02 production is l 0”
COZ-fixation by ribulose 1,5 bisphosphate carboxylase /ox- 0 0”
0
0
0

0”

0”
0”
(C) Nov. 66,2O’C

0 406 866 1260

02 WV, 02 production (#A min.‘)

FIG. 5. Representative examples of the depth distributions of Or

production rate and dissolved O2 concentration with depth in the
mat at different times of the year.

+I?/
4
2-4 mm

ygenase (often referred to as RuBPC/O or RUBISCO), the

COz-fixing enzyme of green plants and cyanobacteria.
0 4 ~ 8 12 16 ~ 2, 24
Sunliee Time (hr)
BADGER ( 1980) isolated pure RuBPC/O from the cyano-
bacterium Anabaena variabilis and showed that, between 8
FIG. 4. The concentrations of elemental sulfur over a full diurnal and 40°C the carboxylase activity (at or near carbon satu-
cycle are shown for the O-2 mm depth interval for a mat held at ration of the enzyme) increased by a factor of 2.3 for a 10°C
30°C during April 90. The concentration of total reduced sulfur (in- increase in temperature. Other factors external to the cell,
cluding pyrite) is shown over the same diurnal cycle for the 2-4 mm
depth interval. The concentrations of total sulfur in the 2-4 mm such as CO2 and nutrient availability, are also temperature-
interval were similar to the concentrations of pyrite sulfur (data not dependent (see the following) and might contribute to the
shown) from the O-2 mm interval. temperature dependence of O2 evolution.
3978 D. E. Canfield and D. J. Des Marais

I 0 0 I
oo” 0 April 90 17uC 0 0
0
April90 3OeC
00
0 750 0
I 0,
0
00 500
0
0
0
250

0 %
0
0 0
80
0
0 0
I 60
0 0 02Prcd
0 0 I A TOk3loz nux

I
I
A
-_--

T 7i-a-
Flux Chamber
El
0
FIUX
Flux
out
I”

r 0 .--_------_ ---A

LL -2
0” -4
-6

-0
9 II 13 15 17 I9 21 23

Time (hrs) Time (hrs)

FIG. 6. Light intensity and microelectrode results at various temperatures during Nov. 89, April 90, and Nov. 90.
Light intensity incident on the mat surface during the course of the day is shown in the top panel. In the middle panel
are the rates of O2 production measured with 02 microelectrodes. During Nov. 89, rates of Q production underestimated
the true rates (see text for details). Also in the middle panel is the flux of O2 diffised from the photic zone calculated
from O2 gradients (for April 90 and Nov. 90). Included are both diffusion into the mat and diffusion out of the mat
into the brine (see Fig. 5 ). These diffusional fluxes are presented individually in the lower panel and are positive for
O2 diffused from the mat into the brine and negative for O2 diffused into the mat. Points represent the average of high
and low fluxes (shown by vertical bars) considering extreme values for net porosity (see text). Also in the lower panel
are the O2 fluxes (into the mat at night and out of the mat during the day) measured by the flux chambers (dashed
lines) and the flux of 02, both day and night, used to oxidize sulfide produced from sulfate reduction (thatched areas).

COMPARISON OF MICROELECTRODE
CHAMBER RESULTS
In constructing the budgets to follow, we need to incor-
porate results obtained from the following three different
methodologies: namely, the microelectrodes, the flux cham-
bers, and sulfate reduction rate measurements. Although the
same light and temperature regimes were reproduced in these
experiments, we must consider whether experimental artifacts
as imposed by potentially different hydrologic regimes could
compromise our budget calculations. The concern is that the
thickness of the benthic boundary layer (BBL; see BOUDREAU
and GUINASSO, 1982 ) may have varied, potentially a&cting
rates of biological processes requiring dissolved species from
the overlying water (photosynthesis, for example). We draw
on several lines of evidence to show that we can, in fact,
reasonably combine our different results.
As mentioned previously, the flux of 02 into the mat at
night (Table 1) was similar to the flux necessary to oxidize
the nighttime sulfide production to sulfate (Table 3, Fig. 6).
This observation is rationalized by observing that in the ab-
sence of O2 production (at night, for example), 9 penetrates
to between only 50- 100 Nrn in Pond 5 mats (see JORGENSEN
AND FLUX and DES MARAIS, 1990). Thus, O2 may be utilized in carbon
oxidation over a depth comprising only about 5-10% of the
normal photic zone thickness and O. l-0.2% of the total mat
thickness. With such a thin zone for O2 respiration, the an-
aerobic process of sulfate reduction must dominate carbon
oxidation at night; and without a mineral repository for sulfide
(pyrite concentrations are low and do not increase with depth;
Fig. 4 (D. E. Canfield, unpubl. data), the sulfide must diffuse
to the sediment surface to be oxidized by Oz. Hence, we
would expect the nighttime O2 flux to match nearly rates of
sulfate reduction, and the good agreement observed (Table
3; Fig. 6) gives us confidence that the flux chambers are mea-
suring material exchange consistent with our independently
determined rates of sulfate reduction.
As a further comparison, during Nov. 90, where we have
the most complete data set, the nighttime flux of 02 into the
mat as determined Born microelectrode 02 gradients is similar
to, though a little lower than, the flux chamber 02 uptake
rate (Fig. 6). The somewhat lower fluxes from the 02 profiles
are not unexpected as a flat surface was assumed in calculating
fluxes, probably underestimating the true mat surface area
and, hence, the actual O2 flux into the mat (JORGENSEN and
DES MARAIS, 1990).
 Biogeochemistry of C, S, and OZin a microbial mat

0 cPo Nov 90 2O’JC

7
n 0

E
oo”
ul
0 oo” 0 0
3 500. 60 500
0 0
E
0,
250. o 8%
3
0

O.@

6 -
2 00
‘E. 12 30-
00 OoO
u2
0 0 0 0 02Prcd
e 8- 0 A Total 02 flux 20. 0 0
0
[L’;. 0
80

0” p- 0

E
0 o-

o_--_--_---_---_----_

-10
:hamber Iamber

-15i-. . . *. . . . . . . . . I
7 9 I I 13 15 I7 19 21 23 7 9 II 13 15 I7 I9 21 23 9 II 13 15 17 I9 21 23

Time (hrs) Time (hrs) Time (hrs)

FIG. 6. (Continued)

Lastly, the daytime O2 release from the mat, as intercepted Nighttime O2 Budget
by the flux chambers, is compared with the diffusional flux
of 02 from the mat calculated from O2 profiles. This com- The details of the nighttime 02 budget were outlined in
parison is used as a surrogate for comparing rates of primary the previous section and show that sulfide oxidation is the
production in the two different experiments, as the release major sink for O2 at night (Table 3). During both Nov. 89
rate of 02 from the mat scales with primary production rate and Nov. 90, there was a small additional flux of O2 in excess
(see the following). During Nov. 90, a relatively complete of that required to oxidize sulfide. We have attributed this
set of Or profiles allows for our most accurate estimate of O2 flux to a small amount of O2 respiration, though it is also
diffusion from the mat. Assuming a flat mat, and considering possible that ammonia oxidation could have acted as a sink
only the time interval over which the flux chamber was de- for 02. The nighttime 02 budget is compiled in Fig. 8.
ployed, a diffisional flux of 4 1.5 +- 6.5 mmol 02 m-’ d-’ is
calculated (with a mat porosity of 0.89 + 0.04; see the pre-
ceding text). Due to irregular surface topography, the true Daytime O2 Budget
value will be somewhat higher (JORGENSEN and DES MARAIS,
1990)) though by an uncertain amount. This range in values, The only source of Oz during the day is photosynthesis.
then, is not inconsistent with the flux chamber result of 51 Daily rates of O2 production are obtained by first combining
mmol O2 m-* d-’ (Table 1; Fig. 6) and gives further justi- the more abundant light measurements with the P vs. I trends
fication for combining flux chamber and microelectrode re- in Fig. 7 to produce a rather complete estimate of 0~ pro-
sults. For the other sampling periods (Fig. 6), rates of O2 duction during the day (Fig. 9a-c). Daytime O2 production
efflux from the mats were also similar for the two different is the area under the curve when multiplied by the mat po-
methodologies. rosity. We have not included in our calculations the time
interval during which the flux chambers were collected and
BUDGETS FOR CARBON AND OXYGEN redeployed. Since microelectrode O2 production rates in Nov.
89 underestimated actual O2 production (see Methods), they
In what follows, comprehensive budgets will be constructed have not been included in the daytime 02 budget. To deter-
describing carbon and 02 cycling in the mats. With these mine the uncertainty of total daytime O2 production values,
budgets, we will identify the processes controlling carbon fix- we note that the productivity measurements were reproduc-
ation during the day and carbon oxidation day and night. ible to +_S%and were made randomly in the mat to accom-
We will see that although the processes do not change, their modate mat variability, estimated as f20%. Since we have
relative quantitative importance does. included different areas of the mat in our P vs. I trends (Fig.
3980 D. E. Canfield and D. J. Des Marais

40
(a) Nov. 69 Nov. 89,2O’C

30

_ 10 Not determined
7
.E
E 0
9
f
z!
E 100
s
60

(b) April 90 60

40

20 Not determined

0
Orprod Dlff-out S-c&d 02-mSp Diff.in Soxid 02-nsp

April 90,17’C

60

April 90,30X

o’.&‘&.&.&.,(

Light (MMmm25-l)

FIG. 7. Rates of O2 production as a function of light intensity onto

the mat surface at different temperatures and different times of the Not delermined
year.

100
7), our uncertainty in calculating total 02 production will Nov. SD, 20°C
be less than +20%, and we use k 10% as a reasonable estimate. 60

During the day, the O2 produced by photosynthesis can 60

diffuse out of the mat or be consumed within the mat by O2

II
40
respiration and sulfide oxidation (Fig. 1). The best constraints
on the O2 used to oxidize sulfide are from April 90, where 20

the concentrations of solid phase sulfur were followed over 0

a die1 cycle at 30°C (fig. 4). The accumulation of elemental
sulfur during the day was significant and accounted for 57%
of the estimated sulfide production. The amount of O2 used
to oxidize sulfide in April 90, then was calculated assuming
that 57% of the sulfide was oxidized to elemental sulfur and
the rest to sulfate. During Nov. 89 and Nov. 90, elemental
sulfur may also have accumulated during the day, but we
can only speculate as to how much and have chosen instead
to assume complete oxidation of sulfide to sulfate. The fluxes
of O2 out of the mat are from the flux chamber measurements
(Table I), and the only unknown parameter is the amount
-
-20’
Orprod Dltt-out S-Ad Orreap DM-In S-mid 02-nYSP
FIG. 8. A histogram showing the fate of O2 night (shaded area)
and day (open area) iri the mat at dilkent temperaturesand different
times of the year. “Or-Prod” is the rate of O2 production by photo-
synthesis, “Diff-out” is the flux of O2 diffused from the mat into the
overlying brine during the day, “S-oxid” is the 02 flux used to oxidize
sulfide, “02-resp” is the O2 used in O2 respiration, and ‘TM-in” is
the flux of O2 diffused into the mat at night. One-standard-deviation
errors, calculated from propagation of error analysis of individual
results, are also shown. Open bars are daytime fluxes; shaded bars
are nighttime fluxes.
 Biogeochemistry of C, S, and O2 in a microbial mat 3981

gree by temperature. During Nov. 90, about 50% of the O2

(a) April So, 17-c 0 PredICted
production was consumed witbin the mat; in this case, our
A Measumd
n mass balance showed no evidence for appreciable 02 respi-
A
M- A ration. (If solid sulfur was cycled during Nov. 90, as in April
00’ p 90, the amount of Oz used to oxidize sulfide, Soxid, is reduced
OO
A 0,
to 23.8 mmol rnw2, and O2 respiration becomes 14.6
Q O mm01 mm2).
10 0 A
We have independent support for our conclusions regard-
0
ing daytime O2 cycling. In Fig 6 (middle panel), rates of O2

0-l
I . 1 . I . I . I . I .
production are compared to the total diffusional flux of O2
into and out of the mat. In the absence of 02 utilization in
the zone of O2 production, the O2 production rate should
match this total diffusional flux. If O2 is utilized in the region
(b) April 66, WC A
0 0
where it is produced, then less is available to diffuse, and the
production rate will exceed the loss rate by diffusion. During
April 90, at both I7 and 3O”C, far more O2 was produced
4” 0
by photosynthesis than was transported by diffusion (Fig. 6))
supporting our earlier conclusion of substantial O2 respiration.
In addition, the flux of 02 into the mat in April 90 was greater
than the flux required to oxidize sulfide, giving further evi-
dence for O2 respiration. This difference, however, represents
only a portion of the total O2 respiration in the mat. As men-
tioned in the preceding text, O2 respiration is significant in
the upper photic zone, and only a relatively small amount
40 of the O2 production is transported into the mat by diffusion.
(c) Nov.O0,2O’C A A During Nov. 90, the combined flux of O2 deeper into the
mat and into the overlying water nearly matched rates of O2
30 O0 production (Fig. 6). This shows that, by contrast with April
A% A’ ’
EoA”
90, very little 02 consumption occurred in the region of O2
0 A production, with most O2 transported away from this zone
20
by diffusion. A small amount of O2 respiration was evident
by a slight excess O2 flux into the mat compared with that
10 required to oxidize sulfide from sulfate reduction (Fig. 6).
q”’ Overall, however, consistent with mass balance results, mi-
croelectrode results gave no indication of appreciable O2 res-
A
0 piration during this time period.
6 6 10 12 14 16 16
lime (hr) Taken together, our results show that O2 respiration can
be a significant process in cyanobacterial mats, with several
FIG. 9. Measured and predicted rates of O2 production during the
possible respiration pathways, including “dark” respiration,
course of a day are shown for different temperatures and different
times of the year. Nov. 89 is not included because the measured rates photorespiration, and aerobic heterotrophic bacterial metab-
underestimated the actual rates, and these data have not been used olism. In “dark” respiration, photosynthetic organisms use
in the O2 budget calculations. See text for details. the chemical energy stored in carbohydrates to produce ATP
for growth and maintenance. In photorespiration, the oxy-
genase activity of the enzyme RuBPC/O catalyzes the oxi-
of Oz used in O2 respiration, which has been calculated as dation of RuBP with 02. Aerobic bacteria use O2 to oxidize
the difference between the total amount of 02 produced and nonliving organic substrate (including mucilage, dead bac-
the amount used to oxidize sulfide and diffused from teria, sheath material, and DOC), and they are very important
the mat. organic carbon oxidizers in nonphotosynthetic marine sed-
A summary of the budget is shown in Fig. 8, with errors iments (JORGENSEN, 1982; BENDER and HEGGIE, 1984;
derived from standard propagation of error analysis. During CANFIELD, 1989).
April 90, only about 20% (at both 17 and 30°C) of the 02 Because rates of O2 respiration during the day in April 90
produced in the mat escaped by diffusion across the mat- (Fig. 8, Table 3) were much higher than could be possible
brine interface. Most of the 02 was cycled internally within at night (see the preceding text), we suggest that there is a
the mat, and most of this 02 was used in 0s respiration; probable link between daytime O2 respiration and photosyn-
sulfide oxidation was a relatively minor sink. Although tem- thesis. Photorespiration would be a likely process linking
perature had a strong effect on the rates of biological processes, photosynthesis to O2 respiration; however, it is generally
there was little effect on how 02 was utilized. Hence, the considered to be suppressed in cyanobacteria (see COLMAN,
processes that produce oxygen (photosynthesis) and the pro- 1989, for a review). As another possibility, cells undergoing
cesses that consume Oz in the mat (sulfide oxidation and 02 photosynthesis might excrete dissolved photosynthate for use
respiration) were influenced during April 90 to the same de- by aerobic bacteria. We must conclude, however, that the
3982 D. E. Canfield and D. J. Des Marais

relative importance of the process( es) controlling O2 respi- Considering sources of DIC, from sulfate reduction (Table
ration in these mats remains unresolved. 2 ) we assume that two moles of organic carbon are oxidized
(producing DIC) for each mole of sulfate reduced. The DIC
Carbon Budget liberated from 02 respiration (Fig. 8) is estimated assuming
that one organic carbon is oxidized to CO2 for each O2 re-
A budget for carbon may be constructed in much the same duced. The DIC diffused into the mat is taken from the flux
manner as for 02. During the day, DIC is fixed within the measurements (Table 1). These data are also summarized
mat by both oxygenic and anoxygenic photosynthesis; these in Table 4.
comprise the important DIC sinks. The sources of DIC for From the compilation of DIC sources, in Nov. 90, about
fixation include the DIC diffused into the mat from the over- 60% of the DIC fixed within the mat was derived by diffusion
lying water, plus the DIC derived from within the mat by O2 from the overlying water, and about 40% was derived from
respiration and sulfate reduction. At night, the DIC lost from internal cycling within the mat, mostly from sulfate reduction.
the mat comes from sulfate reduction and O2 respiration. By contrast, in April 90, at both 17 and 30°C most of the
There is enough information to compute the daytime car- DIC used in photosynthesis was derived from within the mat,
bon budget from both the DIC sources and DIC sinks. Be- with O2 respiration being the most significant source. We
ginning with the DIC sinks, rates of DIC fixation by oxygenic conclude that the mat itself is always a major source of DIC
photosynthesis are estimated from rates of 9 production, for photosynthesis and, at some times of the year, the most
assuming that one carbon is fixed for each O2 produced. DIC significant source. Also, as with the O2 budget, the April 90
fixation by anoxygenic photosynthesis is considered impor- data suggest that temperature had little effect on the relative
tant only during April 90, with the rates assumed to be con- importance of the different carbon sources.
trolled by the diffusional supply of sulfide from within the Total rates of DIC fixation estimated from the DIC sinks
mat and the stoichiometry of the suhide oxidation reaction. are always lower than are rates calculated from the sum of
Four times more organic carbon can be produced if sulfide carbon sources. In other words, there is apparently more DIC
is oxidized to sulfate (Eqn. 1) rather than So (Eqn. 2 ), both available for fixation (sources) than is used (sinks). A similar
of which are potential products of anoxygenic photosynthesis discrepancy is also observed at night, during which more DIC
(COHENet A., 1975; TROPERet al., 1982), as follows: diffuses from the mat (by direct measurement) than is ap-
parently made available by respiration, calculated as the sum
2CO2 + H2S + 2H20
of O2 respiration and sulfate reduction (Table 2, Fig. 8) and
--* 2CH20 + SO:- + 2H+; and (1) assuming the same stoichiometries as described in the pre-
ceding text (results summarized in Table 5).
1/2CO, + H2.S + 1/2CHz0 + So + 1/2HzO. (2)
Ultimately, the mismatch in the DIC budget, both day and
Assuming that all sulfide oxidation was accomplished by night, arises from an imbalance between the DIC and 02
anoxygenic phototrophs, with 57% of the daytime sulfate re- fluxes across the mat/brine interface, as measured by the
duction accumulated as So and the rest oxidized to sulfate flux chambers. During the day, more DIC is diffused into the
(see the preceding text), rates of carbon fixation by anoxy- mat than O2 is released by the mat; at night, more DIC is
genie photosynthesis can be computed. These rates, and rates lost than O2 is consumed. These results are well outside of
of oxygenic photosynthesis, are summarized in Table 4. the analytical uncertainties in both the DIC and O2 mea-
surements. Flux chamber results demonstrate then that dur-
ing the day, carbon is incorporated into the mat with an
Table 4. Daytime DIC Budget
oxidation state greater than zero, and carbon with a similarly
ApriI90170C ApriIW3O”C Nov 90 20°C
high oxidation state is lost from the mat at night. The mis-
match between the sources and sinks in the DIC budget arises
p1c siis:
from the assumption that the carbon fixed in the mat during
OXypk 75.7 (8.3)a 236 (26) 84.2 (9.8) the day and released by the mat at night has an oxidation
Photosynthesis state of zero.
Anoxygenic 5.6 (1.4) 23 (5.8) 0 Over a full die1 cycle, the mat is at a steady state, with
Photosynthesis daytime DIC and O2 fluxes opposite but nearly balancing
the nighttime fluxes (Table 1). We can conclude that most
Total 81.3 (8.4) 259 (27) 84.2 (9.8)
of the more positively charged carbon incorporated into the
mat during the day is liberated from the mat at night. In this
DIC Sources: way, the organic carbon accumulating in the mat (a very
Diffusion from 34.5 (1.8) 98 (4.9) 69 (3.5) small percentage of the amount fixed during a day) may have
overlying water an oxidation state more similar to zero, as expected from the
Sulfate 10.9 (1.3) 45 (5.4) 41.6 (13) carbohydrates, lipids, and amino acids which comprise the
Reduction bulk of mat organic carbon (BOON, 1984). Elemental analysis
oxic 53.5 (8.4) 158 (27) -3.2 (16) of Pond 5 mat (C,H,N,S,O, D. E. Canfield, unpubl. data)
Respiration confirms this and reveals that the average oxidation state of
mat carbon is near zero.
Total 98.9 9.7) 301 (28) 107.4 01)
Just how Pond 5 mats accumulate relatively oxidized car-
a one standard deviation bon during the day is not certain. It is well known that to
 Biogeochemistryof C, S, and O2 in a microbial mat
Table 5. NighttimeIX budget.
mmoles m-2s
Temp S=@ sulf redc Tot&
Nov89 2OW 3.5 (2.5)f 25.1 (2.5) 28.6 (3.5)
30% 16.2 (2.0) 62.8 0.0) 79.0 0.8)
April 90 17X -g 13.9 (1.5) 13.9 (1.5)
JBC % 60.0 (7.2)
60.0 Q.2)
Nov. 90 2OoC 8.0 (0.8) 53.0 (15.9) 61.0 (15.9)
a ratescalculatedover time period of flux experiments
b DIG from oxicrespiration
c DIC from sulfatereduction
d sum of DIC flux from respiration
e measuredDIC flux from mat
f one standarddeviation
g not measured,but assumedzero:see textfor details.
provide more COZ for rapid photoayntheais, cyanobacterial
cells accumulate DIC from their surrounding medium (e.g,
BERRY et al., 1976; BADGER et al., 1978; COLEMAN and
COLMAN, 1981; BADGER, 1985). This accumulation can be
quite impressive; cells of Coccochloris peniocystis, for ex-
ample, attain total DIC concentrations of up to 4 mM, with
an external DIC concentration of only 0.1 mM (COLEMAN
and COLMAN, 198 1). A large store of DIC in cyanobacterial
cells could constitute a sink for oxidized carbon during the
day. However, not enough DIC can be stored by this mech-
anism. Assuming that one-half of the mat volume to a depth
of 1 mm is cell cytoplasm, a high estimate, an internal cell
concentration of 4 mM would result in the accumulation of
about 2 mmol of C m-’ in the mat. This is a small amount
of carbon compared to the daily fluxes into and out of the
mat (Table 1).
We suggest that the imbalance between C and O2 in the
mats may result from the accumulation of the initial oxidized
products of photosynthesis or photorespiration during the
day and from their subsequent oxidation at night. Under
conditions of nutrient stress in cyanobacteria, the Cq dicar-
boxylic acids, malic acid, aspartic acid, and oxaloacetic acid
(carbon oxidation states from + 1 to + 1.5 ) may be significant
early products of COZ fixation. These acids are formed from
phosphoenolpyruvate (PEP) with the enzyme PEP carbox-
ylase, the same as in Cq plants (COLMAN et al., 1976; FEUIL-
LADE et al., 1982; COLMAN, 1989). Experimental evidence
suggests that in cyanobacteria, by contrast with C, plants, Cq
acids are not further involved in the C, carbon reduction
cycle (COLEMAN and COLMAN, 198 1; COLMAN, 1989). In-
stead, they may be used in secondary metabolic reactions,
or, in the case of aspartic acid, in protein synthesis (COLMAN,
1989), though their function is not well understood. Signif-
icant storage of these acids during the day, and reoxidation
at night, could account for the observed imbalance between
the carbon and O2 cycles.
Another possible source of relatively oxidized carbon is
photorespiration. In this process, RuBP is oxygenated and
oxidized first to glycolate (carbon oxidation state of + 1).
Although photorespiration may be inhibited in cyanobacteria
( CHENGand COLMAN,1974; COLMAN,1989), glycolate for-
3983
mation has been demonstrated in cyanobacteria under similar
conditions of high O2 and low COz as experienced during the
day in microbial mats (HAN and ELEY, 1973; BERGMAN et
Measured C flu+ al., 1984). To account for the C02/02 imbalance that we
observe in the mats, glycolate would need to be stored during
43.8 (2.2) the day and oxidized at night. However, we know of no ex-
perimental or field evidence to support this hypothesis.
125 (6.3)
Although we cannot identify the mechanism accounting
32.4 (1.62) for the 02/C02 imbalance, we must acknowledge that some
such mechanism does exist and is likely related to the early
118 (5.9)
products of photosynthesis.
75 (3.8)
SUMMARY
We have identified and quantified the sources and sinks
of carbon, sulfur, and O2 in a cyanobacterial mat and have
constructed budgets to account for the cycling of these ele-
ments. Some process, anoxygenic photosynthesis coupled
with O2 respiration, for example, dominates at certain times
of the year and not at other times. We do not fully understand
this but speculate that microbial populations may be adjusting
to variable light regimes within the mats.
A goal of this study was to document the factors controlling
primary production in these mats, and we advance the fol-
lowing as the most important: light, temperature, and the
availability of nutrients and carbon. Light and temperature
had a demonstrated effect on primary production, but we
did not explore specifically the effects of carbon and nutrient
limitation. We did find that much of the carbon for primary
production was derived from within the mat, and we expect
that the same is also true for the nutrient elements, perhaps
even more so. This is because the ponds in which these mats
grow are very nutrient-poor ( JAVOR, 1983).
It is intriguing that primary production in these mats may
be controlled mostly by internal mat cycling. We note that
rates of primary production responded to temperature
changes by a similar amount as the heterotrophic processes
that make nutrients available for primary production. Also,
the mats are nearly at steady state, such that over 24 h, the
amount of carbon fixed by the mat during the day is nearly
balanced by the carbon lost at night. In principle, then, the
nutrients liberated by the mat over 24 h cycle are sufficient
to fuel primary production during the day (with a small ad-
ditional flux to account for the carbon preserved in the mat).
This scenario requires that nutrients (N and P) lost during
the night are effectively trapped by the mats and not lost to
the overlying water. This matter is currently under investi-
gation.
We suggest that cyanobacterial mats such as those found
in Pond 5 are closely coupled systems where rapid rates of
photosynthesis both require and fuel rapid rates of carbon
oxidation processes. This hypothesis becomes more com-
pelling if mat bacteria can store (at least for a short time)
nutrients made available by oxidative processes and allocate
them to carbon fixation, as a function of the available light.
Acknowledgments-We acknowledge laboratory assistance by Anne
Tharpe; field assistance by Brad Bebout, Jim Bauer, and Bruno Risati,
as well as careful reviews by Linda Jahnke, Larry Hochstein, N. P.
Revsbech, Rick Jahnke, Cindy Lee, and an anonymous reviewer.
This work was supported by a grant to DJD from NASA’s Planetary
3984 D. E. Canfield and D. J. Des Marais
Biology Program and by a fellowship to DEC from the National
Research Council.
Editorial haloing: C. J_ee
REFERENCES
BADGERM. R. ( 1980) Kinetic properties of rib&se l,%isphosphate
carboxylase/oxygenase. Arch. B&hem. Biophys. 201,247-254.
BADGERM. R. ( 1985) The fluxes of inorgslnic carbon species during
photosynthesis in cyanobacteria with particular reference to Sy-
nechococcus sp. In inorganic Carbon Uptake by Aquatic Photo-
synthetic Org~isms (ed. W. J. LUCASand 3. A. BERRY), pp. 39-
52. Amer. Sot. Plant Physiol.
BADGERU. R., KAPLAN A., and BERRYJ. A. ( 1978) A mechanism
for concentrating CO, in Chlamydomonasreinardtiiand Anabaena
variabilis and its role in photosynthetic CO* fixation. Carnegie
Inst. Wash. Yearb. 77,25 I-26 1.
BENDER M. L. and HEGGIE D. T. ( 1984) Fate of ortquric carbon
reaching the deep sea floor: A status report. Geochim. Cosmochim.
Acta 4&977-986.
BERGMANB., CODD G. A., and HALLBOML. ( 1984) Glycolate ex-
cretion by Ns-fixing cyanobacteria treated with photorespiratory
inhibitors. 2. P~anzenphysiol.113,45 l-460.
BERNERR. A. ( 1964) ~st~bution and diagenesis of sulfur in same
sediments from the Gulf of California. Mar. Chem. 1, 17-140.
BERRYJ. A., BOYNTONJ., KAPLANA., and BADGERM. R. ( 1976)
Growth and photosynthesis of Chlamydomonas reinardtii as a
function of CO? concentration. Carnegie Inst. Wash. Yearb. 75,
423-432.
Boon J. J. ( 1984) Tracing the origin of chemicai fossils in microbia1
mats: BiogeochemicaI investigations of Solar Lake cyanobacteriai
mats using analytical pyrolysis methods. In Micobial Mats: Stro-
matolites (ed. Y. COHEN, R. W. CASTENHOLZ,and H. 0. HAL-
VORSON), pp. 3 13-342. Alan R. Liss, New York.
BOUDREAUB. P. and GUINASSON. L. I 1982) The infhrence of a
diffusive sublayer on accretion, dissolution, &id diagenesis at the
sea&or. In The Dynamic Environment of the Ocean Floor (ed.
K. A. FANNING and F. T. MANHEIM), pp. 115-145. Lexir@on.
CANF~ELDD. E. ( 1989) Sulfate reduction and oxic respiration in
marine sediments: Implications for organic carbon preservation
in euxinic environments. Deep-Sea Res. 36, 12 t - 138.
CANFIELDD. E. ( 1993) Organic matter oxidation in marine sedi-
ments. In interactionsofC,N,P, and S Biogeochemical Cycles (ed.
R. WOLLAST, L. CHOU, and F. MACKENZIE), pp. 333-363.
Springer-Verlag.
CANFIELDD. E. and DES MARAIS D. J. ( 199 1) Aerobic sulfate re-
duction in microbial mats. Science 251, 1471-1473.
CANFIELDD. E., RAISWEU R., WE~TRICHJ. T., REAVESC. M., and
BERNERR. A. ( 1986) The use ofchromium reduction in the anal-
ysis of reduced sulfur in sediments and shales. Chem. Geol. 54,
149-155.
CHENC K. H. and COZMANB. ( 1974) Measurement of photores-
pimtion in some microscopic aI8ae. Flanfa 115,207-2 12.
COHEN Y., JORGENSEN8. B., PADAN E., and SHILOM. ( 1975) Sui-
fide-dependent anoxygenic photosynthesis in the cyanobacterium
Oscillatorialimnetica. Nature 257,489-492.
COLMAN B. (1989) Photosynthetic carbon assimilation and the
suppression ofphotorespiration in the cyanobacteria. Aquatic Bot-
any34.21 l-231.
COLMAN B., CHENG K-H., and INGLE R. K. ( 1976) The relative
activities of PEP carboxylase and RuBP carboxylase in blue-green
algae. Plant Sci. Left. 6, 123-127.
COLEMANJ. R. and COLMANB. ( 198 1) Inorganic carbon accumu-
lation and photosynthesis in a blue-green alga as a function of
external PH. Plant Physiol. 67, 9 17-92 I.
D’AMELIOE. D., COHEN Y., and DES MARAIS D. J. ( 1989) Com-
parative functional ultrastructure of two hypersaline submerged
cyanobacterial mats: Guerrero Negro, Baja California Sur, Mexico,
and Solar Lake, Sinai, Egypt. In Microbial Mats, Physiological
Ecology of Benthic Microbial Communities (ed. Y. COHEN and
E. ROSENEERG),pp. 97-113. Amer. Sot. Microbial.
DES MARAISD. J. ( 1990) Microbial mats and the early evolution of
life. Trends Ecol. Evol. 5, 140-144.
EEUILLADEJ., I%UI~_.LADE M., and JOLIVETE. ( 1982) Photosyn~etic
metabolism in the cyanophyte. Oscillatoria rubescens D.C. II.
Carbon metabolism under nitrogen starvation. Arch. Microbial.
131, 107-l 1 I.
FROND C. and COHENY. ( 1992) Diurnal cycles of sulfate reduction
under oxic conditions in cyano~cte~~ mats. Appi. Environ. Mi-
crobiol. 58, 70-77.
GLUD R. N., RAMSING N. B., and REVSBECHN. P. ( 1992) Photo-
synthesis and photosynthesis-coupled tespimtion in natural bioliIms
measured by use of oxygen micmsensors. J. Phycol. 28,5 I-60.
GRASSHOFF K. ( 1976) Method of SeawaterAnafysis.Verlag Chemie.
HAN T. W. and ELEY J. H. ( 1973) GlycoIate excretion by Anacystis
nidulans: Effect of HCO, concentration, oxygen concentration,
and light intensity. Plant Cell Physiol. 14, 285-291.
JAVORB. J. ( 1983) Planktonic standing crop and nutrients in a sahern
ecosystem. Limnol. Oceanogr. 28, 153- 159.
J@RGENSENB. B. ( 1982 ) Mineralization of organic matter in the sea
bed: The role of sulfate reduction. nature 2%,643-645.
JORGENSENB. B. ( 1983) Processes at the sediment-water interface.
In The Major Biogeochemical Cycles and Their Interactions(ed.
B. l!bLIN and R. B. COOK), pp. 477-5 15. J. Wiley & Sons.
JORGENSENB. B. and DES MARAIS D. J. ( 1986) Competition for
sulfide among coforless and purple sulfur bacteria in cyanobacterial
mats. FEMS Mi&rob~ol.Ecok 38, 179- 186.
J~~RGENSENB. B. and DES MARAIS D. J. ( 1990) The diffusive
boundary layer of sediments: Oxygen microgradients over a mi-
crobial mat. Limnol. Oceanogr. 35, 1343-l 355.
J~~RGENSEN 13. B., COHEN Y., and DES MARAIS D. J. ( 1987) Pho-
tosynthetic action spectra and adaptation to spectral light distri-
bution in a benthic cyanob~te~aI mat. Appt. Environ. Mier~iol.
53,879-886.
MARTIN J. H., KNAUER G. A., KARL D. M., and BROENKOW
W. W. ( 1987) VERTEX: Carbon cycling in the northeast Pacific.
Deep-Sea Res. 34,267-285.
NELSOND. C. and JANNASCHH. W. ( 1983) Chemauto~p~c~o~h
of marine B~gi~oa in suI~~~dient cultures. Arch. ~icrobiol.
136.262-269.
PIERSONB. K., and CASTENHOLZR. W. ( 1974) Phototrophic gliding
filamentous bacterium of hot springs, ChloroJlexus-aurantiacus,
gen and sp.-nov. Arch. Microbial. 100, 5-24.
REVSBECHN. P. and JORGENSEN 8. B. ( 198 1 f Primary production
of microalgae in sediments measured by oxygen microprofile,
Hr4CO; fixation, and oxygen exchange methods. Limnoi. Ocean-
ogr. 26, 7 I Y-730.
REVSBECHN. P. and WARD D. M. ( 1983) Oxygen microelectrode
that is insensitive to medium chemical composition: Use in an
acid microbial mat dominated by Cyanidium caldarium. Appf
Environ. Microbioi. 45, 755-759.
REVSBECHN. P., JORGENSENB. B., and BLACKEURNT. H. ( 1983)
Microelectrode studies of the photosynthesis and 02, H2S, and pH
profiles of a microbial mat. Limnol. Oceunogr. 28, 1062- 1074.
REVSBECHN. P., MADSENB., and J~~RGENSEN B. B. ( 1986) Oxygen
production and consumption in sediments at high spatiaI resolution
by computer simulation of oxygen data. Limnol. Oceanogr. 31,
293-304.
SKYRING G. W., CHAMBERSL. A., and BAULD J. (1983) Sulfate
reduction in sediments colonized by cyanobacteria, Spencer Gulf,
South Australia. Australian.I. Mar. Freshwaier Res. 34,359-374.
STAL L. J., VANGEMERDENH., and KRUMBEINW. E. f 1985) Stnsc-
ture and developm~t of a benthic marine microbiaI mat. FEhfS
h4icrobiof. &of. 31, .I I- 125.
TROPERH. G., FISCHERU., and KELLY D. P. ( 1982) Anaerobic
oxidation of sulfur compounds as electron donors for bacterial
photosynthesis. Phil. Trans. Roy. Sot. London 2!%,529-542.
ULLMAN W. J., and ALLER R. C. ( 1982) Diffusion coefficients in
near-shore sediments. Limnol. Oceanogr. 27,552~5.56.
WESTRICHJ. T. and BERNERR. A. ( 1984) The role of sedimentary
organic matter in bacterial sulfate reduction: The G model tested.
Limnol. Oceanogr. 29,236-249.