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AQUATIC MICROBIAL ECOLOGY Vol. 48: 123–130, 2007 Published July 10 Aquat Microb Ecol

Resistance to burial of in stromatolites

Jacco C. Kromkamp1,*, Rupert Perkins2, Nicole Dijkman1, Mireille Consalvey3, Miriam Andres4, R. Pamela Reid4

1Netherlands Institute of Ecology, Centre for Estuarine and Marine Ecology (NIOO-CEME), PO Box 140, 4400 AC Yerseke, The Netherlands 2School of Earth, Ocean and Planetary Sciences, Cardiff University, Cardiff CF10 3YE, UK 3National Institute of Water & Atmospheric Research (NIWA), Greta Point, Private Bag 14-901, Kilbirnie, Wellington, New Zealand 4University of Miami/RSMAS-MGG, 4600 Rickenbacker Causeway, Miami, Florida 33149, USA

ABSTRACT: Stromatolites are complex lithified structures with a well-defined layered structure thought to have been formed by trapping and binding of sediment particles by micro-, especially cyanobacteria. Modern marine stromatolites in live in a high-energy environ- ment (surf zone) and are regularly buried by moving sands. We investigated stromatolite cyanobac- terial photophysiology ex situ, during and after sand burial using variable fluorescence studies. Buried samples inactivated their photosynthetic electron transport, but only when oxygen concentra- tions decreased to low levels. Post-burial, the stromatolite cyanobacterial community reactivated its photosynthetic activity within 1 to 2 h, but this activation was light dependent. It is therefore specu- lated that the redox state of the plastoquinone pool determines the inactivation/reactivation pro- cesses. The ability of cyanobacteria to survive and recover from burial by sediment could be a funda- mental attribute that has contributed to the success of cyanobacteria as stromatolite builders and for the actual existence of stromatolites as organo-sedimentary structures with a putative presence spanning 3500 million yr.

KEY WORDS: Stromatolite burial · Cyanobacteria · fluorescence · PSII · State transition

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INTRODUCTION sedimentation and lithification. Three growth types (with intermediate forms) have been described (Reid et Stromatolites were common prior to the al. 2000). The non-diazotrophic cyanobacterium Schizo- diversity explosion 490 to 540 million yr ago (Awramik trix sp. dominates Pioneer Type 1. The extracellular 1992) and were then widespread in the shallow seas. polymeric substances (EPS) excreted by the cyanobac- Nowadays, modern stromatolites growing in open teria capture the sediment grains (ooids), causing the marine environments are restricted to the margins of comparatively unstable mats to grow. As a result, most of the Exuma Sound in the Bahamas and , the biomass is normally found below the stromatolite . It is thought that the well-laminated micro- surface at variable depths (Fig. 1), although a significant bial structures present here are modern analogues to cyanobacterial biomass can be present above this layer. ancient stromatolites, although the role of eukaryotic Domination of heterotrophic processes leads to the epiphytes has not been evaluated yet. formation of a micritic layer at the surface, typical for a According to Reid et al. (2000), the formation of Type 2 mat. This 20 to 60 µm white layer is formed by stromatolites and stromatolite-like mats found in the precipitation of calcium carbonate, most likely liberated Bahamas depends upon a dynamic balance between by consumption of EPS by heterotrophic activity (Viss-

*Email: [email protected] © Inter-Research 2007 · www.int-res.com 124 Aquat Microb Ecol 48: 123–130, 2007

Fig. 1. Cross-section through a live stromatolite, showing the clearly visi- ble subsurface cyanobacterial layer. The brown layer of extracellular polymeric substances directly below the surface also contains many cyanobacteria, mainly belonging to the genus Schizotrix cher et al. 2000, Paerl et al. 2001). About 15% of the mats MATERIALS AND METHODS in the vicinity of Highborne Cay are of Type 2. Another 15% of the stromatolite cyanobacterial mats are of Sampling. All sampling was performed on High- Type 3, the climax community state. These mats obtain borne Cay (24° 43’ N, 76° 49’ W), a small northerly their stability by the endolithic unicellular cyanobac- island in the Exuma Cays (Fig. 2). Samples of stromato- terium Solentia sp., which bores into the ooids and lite mats were transported in in the shade by this process fuses the sand grains together. The So- and kept in a pool of running seawater collected from lentia spp. are found beneath the Schizotrix surface layer near the sample site. Fresh samples were collected (Macintyre et al. 2000, Reid et al. 2000). daily. All laboratory measurements were made inside Stromatolites in the Bahamas can be found in a high- the the RV ‘Walton Smith’, with rapid transport from energy environment, i.e. in the low intertidal and sub- storage pools to the measurement facilities. tidal waters behind small algal reefs on the east side of Assessment of photosynthetic activity. Photo- the islands, facing Exuma Sound. Despite the shelter synthetic quantum efficiencies were measured with a provided by these small reefs, strong currents, passing DIVING-PAM (Walz, having a red [655 nm] measuring storms and hurricanes move large amounts of sand, light [ML]) or with a WATER-PAM equipped with the regularly exposing and burying the stromatolites and fiberoptic version of an emitter-detector unit (WATER- mats. Burial will most likely induce heterotrophic con- EDF/B, Walz, equipped with a blue [470 nm] ML and ditions leading to a Type 2 mat. This is supported by actinic light source). For both instruments, the saturat- observations that mats dug out from under the sand ing pulse length was set at 0.6 s and the light intensity have the whitish appearance characteristic of the exceeded 3500 to 4000 µmol photons m–2 s–1, as mea- micritic layer characteristic for Type 2 communities sured with the DIVING-PAM light sensor, which was (J. C. Kromkamp & R. Perkins pers. obs.). What is the calibrated against a LiCor LI-190 irradiance sensor. In fate of the buried autotrophic communities? Examina- the dark, the maximum quantum efficiency of photo- tion of naturally buried samples (length of burial not system II (PSII; Fv/Fm) is calculated as Fv/Fm = (Fm – known) indicates a loss of the epiphytic surface com- F0)/Fm, where Fm is the maximum fluorescence yield munity (mainly ), but an apparent persistence and F0 is the minimum fluorescence. In the light, the of the subsurface cyanobacteria. How do the cyano- apparent or operational PSII quantum efficiency is cal- survive this dynamic process of burial and culated as ΔF/Fm’ = (Fm’ – F)/Fm’, where F and Fm’ are exposure? Do they inactivate their photosynthetic the operational and maximum fluorescence yield mea- activity or do they just die, being replaced by new sured in the light (Genty et al. 1989). Relative electron colonisers? When they are exposed, can they reacti- transport rates (rETR, relative due to the inability to vate photosynthetic activity, and if so how quickly? measure the light absorption coefficients) and the These questions are addressed in this paper. exact light level to which the cyanobacteria were Kromkamp et al.: Stromatolite burial 125

curves were fitted using the model of Eilers & Peeters (1988). From the fits the relative max-

imum rate of electron transport (rETRmax) and the initial slope of the rapid light curve (α), a measure of the photosynthetic efficiency, were obtained. See Kromkamp & Forster (2003) for more information about the fluores- cence terminology followed in this paper. The PSII antenna of the cyanobacteria (with only phycocyanine) absorbs little blue light, lead- ing to low fluorescent yields. However, the fiberoptic version of the WATER-PAM is very sensitive due to the photomultiplier detector, and we obtained good signals with relatively low gain and measuring light settings. De- spite the differences in ML colour between the 2 fluorometers, test results showed that quantum efficiencies and RLCs were not sig- nificantly different when the same positions on different stromatolites were measured. Oxygen measurements. The oxygen con- centration was measured using fiberoptic microsensors (with 140 µm tip diameter; Microx TX3, PreSens). To protect the sensi- tive fiberoptic tip, the microsensor was put into a 0.6 mm syringe needle with its tip flush

with the end of the needle, and the O2 con- Fig. 2. Approximate location of the sampling stations (1 to 10) on centrations were measured 1 to 2 mm above Highborne Cay, an island of the Exuma Cays, Bahamas. Most stromato- the stromatolite surface. The optodes were lites were found in subtidal waters and in the surf zone at depths of up calibrated in air-saturated seawater and N2- to 2 m. The grey areas on Highborne Cay are beaches gassed anoxic seawater. In the case of the O2- degassing experiments (by bubbling a 370 ml exposed (Sakshaug et al. 1997, Perkins et al. 2002, bottle with a stromatolite sample which had a 1 cm Forster & Kromkamp 2004, Consalvey et al. 2005) were sand layer on the bottom), discrete water samples were calculated from rapid light curves (RLC) calculated as taken with a 20 ml syringe and emptied carefully into a rETR = ΔF/Fm’ × E, where E is the irradiance generated 20 ml scintillation vial, after which the O2 concentra- by the internal PAM light sources. Depending on the tion was measured immediately. This water was size of the stromatolite samples (approximately 10 to returned to the bottle after the measurement. 30 cm2), 3 to 6 RLCs were made per sample, using a 2 mm spacer to keep the distance to the stromatolite surface as constant as possible. The stromatolites were RESULTS AND DISCUSSION dark adapted for approximately 30 to 45 min. No fluo- rescence measurements were made on spots where Fig. 3 shows the result of a burial experiment. A sam- epiphytic growth was visible. RLCs performed ple was put in a jar, the fiberoptic tip of the fluorome- on reactivated stromatolites were made after 1 min of ter was carefully placed on the stromatolite surface dark adaptation. RLCs were developed to probe the and then the jar was filled with site sand and water (no photosynthetic activity at in situ conditions, and rates stirring). The maximum PSII efficiency (Fv/Fm) of rETR at low irradiances may be influenced by pro- declined gradually until it reached zero, indicating a cesses inducing non-photochemical quenching (Serô- complete inactivation of . After 870 min dio et al. 2005, Perkins et al. 2006). Preliminary results the sample was uncovered, given fresh seawater and showed that when the length of the light steps varied kept in darkness, but this had little effect on Fv/Fm. between 20 and 120 s this did not influence the shape Upon application of low light (upward arrow), the of the RLC. This suggested that acclimation to each effective PSII quantum efficiency (ΔF/Fm’) increased light did not play a role during the RLC. We used a 30 rapidly. Temporarily turning off the light (downward s exposure time for each irradiance value. The light arrow) caused a slight decline in Fv/Fm, prior to further 126 Aquat Microb Ecol 48: 123–130, 2007

0.5 500 0.25

0.4 400 0.20 ’

m 300 0.15

0.3 F , F 200 0.10 0.2 F 100 0.05 Fm’

0.1 ΔF/Fm’ PSII quantum efficiency

PSII quantum efficiency 0 0.00 020406080 0.0 0 200 400 600 800 1000 1200 Time (min)

Time (min) Fig. 5. Changes in F and Fm’ during recovery at an irradiance –2 –1 Fig. 3. Quantum efficiency of a stromatolite sample buried of 77 µmol photons m s . At the downward arrows the under 3 cm of site sand, followed by direct exposure to low light was temporarily turned off and was turned on again at light (open symbols). Upward arrows indicate low light on upward arrows. Note the increase in variable fluorescence (23 µmol photons m–2 s–1); downward arrow indicates light off. (the difference between Fm’ and F) caused by the rise in Closed symbols refer to a control sample kept in the dark Fm’, indicating a recovery of PSII activity as measured by the Δ but not buried. After 870 min (dashed line) the sample was rise in F/Fm’ uncovered. PSII: Photosystem II

in a similar manner as shown in Fig. 3. This corrobo- recovery to initial values when the light was re- rates the idea that light is necessary to reactivate applied. A control sample kept in the dark (not buried cyanobacterial photosynthesis after inactivation. To thus allowing gas exchange with the overlying seawa- test this, 2 experiments were performed. The first ter) showed no change in Fv/Fm over this 20 h period. A experiment showed recovery of PSII efficiency of a repeat of this experiment with a sample that was sample which was buried for 4 d (Fig. 5). Then, 1 d buried for 5 d showed a nearly identical recovery pat- after the start of the burial, we measured the O2 con- tern: after 5 d Fv/Fm had dropped to zero, but when the centration near the stromatolite surface by slowly low- sample was exposed to seawater in the dark a very ering the syringe needle containing the oxygen sensor small increase in Fv/Fm was noted to 0.02 (Fig. 4). With using a manual micromanipulator. Near the surface of the application of a low irradiance of 23 µmol photons the stromatolite the oxygen concentration was 0%. m–2 s–1 recovery increased more rapidly. Brief periods Recovery was followed using a medium low light of darkness arrested the reactivation of photosynthesis intensity of 77 µmol photons m–2 s–1. This sample reac- tivated quickly: fluorescence yields dropped initially, 0.25 but after 10 min Fm’ rose gradually, whereas F decreased slightly, and as a result ΔF/Fm’ recovered rapidly and reached a more-or-less stable value after 0.20 50 to 60 min. Note that on each occasion when the light was turned off for a short period recovery was arrested 0.15 and commenced again when the light was re-applied.

This temporary decrease in Fv/Fm was mainly the 0.10 result of a decrease in Fm. In the second experiment, reactivation was followed for a sample, which had been buried under sand for 4 d (Fig. 6). When the 0.05

PSII quantum efficiency was removed from under the sand and put in fresh seawater and exposed to a very low irradi- 0.00 ance (4 µmol photons m–2 s–1), hardly any recovery 0 20 40 60 80 100 120 140 160 took place during the first hour. Increasing the irra- Time (min) diance to 77 µmol photons m–2 s–1 induced develop- Fig. 4. Post-burial recovery in quantum efficiency of a stroma- ment of variable fluorescence, showing reactivation of tolite sample that had been buried under site sand for 5 d. The photosynthesis, and ΔF/Fm’ stabilised at a rather low sample was uncovered and exposed to darkness followed by value of ~0.06. Turning the irradiance down after low light of 23 µmol photons m–2 s–1 in air-equilibrated sea- –2 –1 water. The upward arrows indicate light on; the downward ~80 min to 4 µmol photons m s caused a decrease in arrows indicate data points when the light was switched off ΔF/Fm’, and, upon re-application of 77 µmol photons Kromkamp et al.: Stromatolite burial 127

550 0.12 this time point both the minimal (F0) and maximal (Fm) F fluorescence yields decreased, but after 0.7 h F0 started Fm’ 0.10 500 to increase slowly, whereas F continued to decrease. ΔF/Fm’ m 0.08 As a result the quantum efficiency (Fv/Fm) decreased ’ 450 ’ steadily. With the exception of the first 20 min, Fv/Fm m m 2 F 0.06 F was linearly correlated to the oxygen concentration (r / , F F = 0.94). The oxygen concentration outside the stroma-

400 Δ 0.04 tolite did not drop below 15 to 20% saturation, due to minor mixing of air with the seawater because we 350 0.02 could not close the lid of the bottle completely. A 300 0.00 repeat of this experiment showed the same result (not 0 20 40 60 80 100 120 shown), and when it was placed in a tray with fresh Time (min) seawater and exposed to the dim light conditions in the laboratory, Fv/Fm recovered from 0.05 to 0.25 in 1.5 h Fig. 6. Changes in fluorescence parameters during recovery (not shown). A control experiment carried out in the in air-equilibrated seawater conditions exposed to a very low Δ irradiance of approximately 4 µmol photons m–2 s–1. At the up- full sunlight showed that F/Fm’ did not change when ward arrows the irradiance was increased to 77 µmol photons the oxygen concentration dropped from 100 to 30%. m–2 s–1. This induced recovery of variable fluorescence as re- However, when the bottle containing the stromatolite Δ flected by a rise in F/Fm’. At the downward arrow the light was placed in the shade, the quantum efficiency was temporarily turned off decreased rapidly, whereas the O2-concentration showed a further, but limited, decrease from 30 to m–2 s–1, the situation reversed as the sample continued 20%. When the bottle was placed in the light again, to reactivate, and ΔF/Fm’ now stabilised at a higher, but ΔF/Fm’ increased once more, while the O2 concentra- still low, value of 0.1. tion remained the same. These results suggest that low These results showed that darkness alone was not oxygen conditions in the light did not lead to inactiva- sufficient to induce inactivation of the photosynthetic tion of photosynthesis (Fig. 8). activity of the stromatolites, because the sample kept Cyanobacteria in stromatolites thus deactivate pho- in aerobic conditions in the dark maintained a high tosynthetic activity after burial due to dark and anaer-

Fv/Fm. As the oxygen concentration in the buried sam- obic conditions. However, they seem to cope with ple decreased due to heterotrophic microbial activity, these conditions quite well, as they reactivate photo- it seems likely that oxygen played a role in the inacti- vation. This was tested by bubbling a sample in a dark- ened bottle with N gas. During the first 20 min no sig- 1200 0.5 120 2 shadesun shade nificant changes were observed, although the oxygen 1000 100 concentration dropped from 100 to 70% (Fig. 7). After 0.4 800 80 ’ ’ 0.3 m

1600 0.35 120 m F F (%) Fo 600 / 60 , 2 F

Fm F Δ 0.30 0.2 O 1400 Fv/Fm 100 [O ] 400 F 40 2 0.25 F ’ m 0.1 80 200 ΔF/F ’ 20 1200 m m 0.20 m O2 F F /

, 60 0 0.0 0 v F 0.15 F 11:00 12:00 13:00 14:00 1000 40 0.10 Time of Day (min) concentration (%) 2 Δ 800 O Fig. 8. Changes in F, F ’ and F/F ’ in full sunlight during a 0.05 20 m m decrease in oxygen conditions induced by bubbling with N2 gas. At the downward arrow the bottle was placed in direct 600 0.00 0 0.0 0.5 1.0 1.5 2.0 2.5 3.0 sunlight and at the upward arrow the bottle was returned to Δ Time (h) the shade. Note that F/Fm’ only reacted to irradiance, but not to [O2]. The O2 concentration is the average of a triplicate, and Fig. 7. Changes in fluorescence parameters and oxygen the standard deviation, although plotted, is generally smaller concentration during inactivation of a microbial mat put in a than the size of the symbols. The fiberoptic tip of the DIVING- bottle in the dark and bubbled with N2 gas. Due to sampling, PAM fluorometer was placed 4 mm above the stromatolite the fiberoptic probe was moved to a slightly different position surface at a small angle, in order to avoid shading of the sur- above the stromatolite surface causing a jump in F0 and Fm face by the fiberoptics. Unfortunately, the light data of the at about 20 min LiCor data logger were lost due to damage to the data logger 128 Aquat Microb Ecol 48: 123–130, 2007

synthesis quickly, although the reactivation seems to schmidt 2000). The high fluorescent state (State 1) is occur in a 2-step process: an initial phase, which is induced when the plastoquinone (PQ) pool is oxidised completed within 1 to 2 h under the conditions we by preferential excitation of PSI. Most cyanobacteria tested and which is characterised by a quick recovery are in the low fluorescent state (State 2) in the dark to a stable but lowered Fv/Fm, and a second phase, (Schreiber et al. 1995, Campbell et al. 1998), because which must follow, as Fv/Fm, ETRmax and α of the reac- respiration causes a reduction of the PQ and the pri- tivated samples were all still below the control values mary electron acceptor of PSII, (QA) pool. During the (Table 1). This second recovery phase was either very inactivation process, oxygen plays an important role: slow or was characterised by a delay and not observed as long as oxygen is present, no inactivation takes by us. On the other hand, it cannot be excluded that place. If microbial activity decreases the oxygen con- part of the cyanobacteria did die during the burial centration slowly after burial, this will prevent re-oxi- event, but that these dead cyanobacteria or their dation of the PQ-pool, thus fixing the PQ and also QA in chlorophyll breakdown products contributed to the a reduced state. This will also initiate 2 simultaneously background fluorescence, thus lowering the measured occurring processes: the reduction of the PQ pool initi- PSII quantum efficiencies. This requires further study. ates a transition from State 1 to State 2, thus a decrease

To our knowledge this is the first study investigating in both F and Fm, whereas the reduction of the QA pool the inactivation/reactivation kinetics of photosynthesis will lead to closure of the PSII reaction centre, causing of the stromatolite cyanobacterial community using a rise in F. Both these processes can be observed in Fig. variable fluorescence techniques, although one similar 7, where the decrease in both F and Fm between 20 and study investigated the reactivation of PSII activity 45 min are dominated by the State 1 to State 2 transi- upon rehydration of cyanobacteria in beach rock tion, whereas the slow rise in F between 0.6 and 2.5 h (Schreiber et al. 2002). Although epi- is caused by a simultaneous reduction of the QA pool, phytic diatoms could be found on the stromatolites, the causing F to rise, and the State 1 to 2 transition, caus- surface community of eukaryotic algae was absent ing F and Fm to drop. from samples that had been buried, suggesting that For the reactivation process, both light and oxygen the epiphytes cannot withstand the abrasive forces are necessary. This can be explained by assuming that imposed by the burial events. The signals we mea- Rubisco needs to be activated by the enzyme Rubisco sured were thus obtained from the cyanobacterial activase (Salvucci et al. 1985). This light-activated component of the stromatolites. The generally low enzyme requires ATP, and its activity is thus related to apparent maximum Fv/Fm values characteristic for the energy charge (Salvucci & Ogren 1996, Jensen cyanobacteria corroborate this. 2000). ATP generation can be achieved by electron

Interpretation of fluorescence kinetics in cyanobac- flow from PSI to O2 via ferredoxin (the Mehler reac- teria is complicated by state transition (Campbell et al. tion), leading to zero net O2 exchange (Asada 2000). 1998) and by the fact that they can use the thylakoid This activation of Rubisco will generate PSI activity, membranes both for photosynthesis and respiration thus inducing re-oxidation of PQ and QA, and this will (Scherer 1990). The PSII antenna consists of phycobili- restore variable fluorescence. However, as not all somes, whereas the majority of the PSI antennae con- cyanobacteria contain the rca gene for this enzyme (Li sists of chlorophyll a. Both photosystems display a dif- et al. 1999), and no published genetic information is ferent excitation spectrum, and, in order to achieve a available for the strains found in stromatolite: this balanced excitation pressure, state transitions occur explanation is our working hypothesis. On the other (Allen 1992, Campbell et al. 1998, Allan & Pfann- hand, Lyngbya spp. do occur in stromatolites, and L. estuarii CCY96106 contains a rca-like gene, which has a very high similarity Table 1. Summary of photosynthetic characteristics obtained using 30 s rapid light curves (RLC, initial slope α); means ± SD are given. Of the control sam- (80%) with the Anabaena rca-gene ples, only 3 RLC had an rETRmax (relative maximum rate of electron transport) (J. C. Kromkamp unpubl. data). Theo- between 10 and 20, and none had an rETRmax < 10. All differences were signifi- retically, the renewed presence of cant (t-test). More measurements were made on the (different) control samples oxygen could also have stimulated than on buried samples respiratory activity (in a form similar to chlororespiration), leading to a partial α Condition Fv/Fm rETRmax re-oxidation of the PQ pool, which will Control 0.40 ± 0.10 46.2 ± 23.4 0.25 ± 0.12 also lead to restoration of some vari- (n = 55) (n = 30) (n = 30) able fluorescence. Both mechanisms After reactivation 0.19 ± 0.09 16.3 ± 3.3 0.10 ± 0.08 were invoked by Schreiber et al. (n = 6, p = 0.002) (n = 6, p = 0.006) (n = 6, p = <0.001) (2002) to be responsible for the restoration of PSII activity in their Kromkamp et al.: Stromatolite burial 129

beach rock community. However, in our case, oxygen tion of reaction centre genes in chloroplasts. Philos Trans alone was not sufficient to restore PSII activity: light R Soc Lond B 355:1351–1360 was necessary as well, and very low intensities caused Asada A (2000) The water–water cycle as alternative photon and electron sinks. Philos Trans R Soc Lond B 355: hardly any reactivation, especially in samples that had 1419–1431 been buried for several days. This reactivation caused Awramik SM (1992) The oldest records of photosynthesis. a State 2 to State 1 transition, which is clearly visible in Photosynth Res 33:75–89 Campbell D, Hurry V, Clarke AD, Gustafsson P, Öquist G Fig. 6 as a rapid initial rise in both F and Fm’ of about –2 –1 (1998) Chlorophyll fluorescence analysis of cyanobacterial 10% at the onset of 77 µmol photons m s . This rise photosynthesis and acclimation. Microbiol Mol Biol Rev in F was followed by a substantial decrease, caused by 62:667–683 a partial re-oxidation of the PQ pool. Theoretically, it Consalvey M, Perkins RG, Paterson DM, Underwood GJC cannot be excluded that toxicity by sulphide might (2005) Pam fluorescence: a beginners guide for benthic have played a role in the inactivation and reactivation, diatomists. Diatom Res 20:1–22 Eilers PHC, Peeters JCH (1988) A model for the relationship as it is known to inhibit photosynthesis. However, mat- between light intensity and the rate of photosynthesis in forming cyanobacteria, which can use sulphide as an . Ecol Model 42:199–215 electron donor for PSI in the light, seem to cope with Forster RM, Kromkamp JC (2004) Modelling the effects of high sulphide concentrations in general rather well chlorophyll fluorescence from subsurface layers on photo- synthetic efficiency measurements in microphytobenthic (Stal 1995), and one might speculate that cyanobacte- algae. Mar Ecol Prog Ser 284:9–22 ria that cannot tolerate sulphide will most likely not Genty B, Briantais JM, Baker NR (1989) The relationship occur in these microbial communities. between quantum yield of photosynthetic electron trans- port and quenching of chlorophyll fluorescence. Biochim Biophys Acta 990:87–92 Jensen RG (2000) Activation of Rubisco regulates photosyn- CONCLUSIONS thesis at high temperature and CO2. Proc Natl Acad Sci USA 97:12937–12938 Stromatolites thus seem well equipped to survive Kromkamp JC, Forster RM (2003) The use of variable fluores- periods of burial, at least of a few days, as we did not cence measurements in aquatic ecosystems: differences between multiple and single turnover measuring proto- investigate burial periods lasting several weeks or cols and suggested terminology. Eur J Phycol 38:103–112 longer. Due to microbial oxygen consumption under Li LA, Zianni MR, Tabita FR (1999) Inactivation of the mono- the sand, PSII will become inactivated, but when the cistronic rca gene in Anabaena variabilis suggests a phys- sand is removed by the currents and stromatolites are iological ribulose bisphosphate carboxylase oxygenase exposed to normal seawater conditions recovery can activase–like function in heterocystous cyanobacteria. Plant Mol Biol 40:467–478 be very quick. The anoxic conditions will reduce PQ Macintyre IG, Prufert-Bebout L, Reid RP (2000) The role of and QA pools, leading to a disappearance of variable endolithic cyanobacteria in the formation of lithified lami- fluorescence and thus of photosynthetic activity. Upon nae in Bahamian stromatolites. Sedimentology 47: exposure to normal oxic conditions, we think it likely 915–921 Paerl HW, Steppe TF, Reid RP (2001) Bacterially mediated that electron donation from PSI to O2 will generate precipitation in marine stromatolites. 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Editorial responsibility: William Li, Submitted: October 18, 2006; Accepted: March 29, 2007 Dartmouth, Nova Scotia, Proofs received from author(s): June 25, 2007