Light-exposed coastal sediments in shallow waters and intertidal areas are often colonized by benthic microalgae and cyanobacteria, which under the absence of animal grazing (typically under environmental extremes such as desiccation, high salinity or sulfide levels) can form complex stratified microbial biofilm communities, i.e., microbial mats (Stal, 1995), that stabilize the sediment by excretion of exopolymers. Microbial mats are densely populated and highly compacted, vertically stratified microbial communities characterized by steep gradients of physical (light and temperature) and chemical parameters (Kühl et al., 1996; Dillon et al., 2009; Al-Najjar et al., 2012; De Beer and Stoodley, 2013). The uppermost layers of coastal microbial mat layers are typically dominated by diatoms on top of a dense green cyanobacterial layer that is often dominated by Microcoleus chtonoplastes and various other motile, filamentous cyanobacteria (Wieland et al., 2003; Fourcans et al., 2004; Dillon et al., 2009). Often, purple sulfur bacteria and green filamentous anoxygenic phototrophs are found below the cyanobacteria followed by a reduced black layer of precipitated iron sulfide (Jørgensen, 1982). Besides light-driven sulfide oxidation by anoxygenic phototrophs, sulfide can also be oxidized efficiently by colorless sulfur bacteria such as filamentous Beggiatoa spp. (Nelson and Castenholz, 1981) that are motile and produce white patches in the microbial mat at the oxygen-sulfide interface (Jørgensen and Revsbech, 1983).

Light is the primary energy source for photosynthetic microbial mats. Due to the high density of photopigments, organic matter, and sediment particles, light is subject to intense scattering and absorption within microbial mats (Kühl and Jørgensen, 1994; Kühl et al., 1994). This can lead to an extremely narrow photic zone (Kühl et al., 1997) and a rapid change in spectral composition with depth (Lassen et al., 1992; Cartaxana et al., 2016b). Ploug et al. (1993) related changes in light quality in a coastal microbial mat to the vertical zonation of a population of diatoms over a dense filamentous cyanobacteria layer that largely sustained their oxygenic photosynthesis via phycobiliproteins with absorption characteristics complementary to chlorophylls. Similarly, complementary use of visible and near-infrared radiation by chlorophylls/phycobilins vs. bacteriochlorophylls enables coexistence of dense populations of oxygenic phototrophs on top of anoxygenic phototrophs (Kühl and Fenchel, 2000). Apart from light, other parameters such as nutrient availability or the presence of sulfide may vertically limit photosynthesis in microbial mats (Stal, 1995; Kühl et al., 1996; Wieland et al., 2003).

The ecological success of benthic microbes in optically dense and vertically stratified communities has recurrently been linked to cell motility allowing individual microbes to search for optimal environmental conditions regarding crucial parameters such as light, temperature, O₂ or nutrient availability (Whale and Walsby, 1984; Bebout and Garcia-Pichel, 1995; Bhaya, 2004; Serôdio et al., 2006). Complex migratory rhythms determined by day/night cycles, tidal regimes, UV exposure and changes in irradiance levels have been described for both diatom- and cyanobacteria-dominated phototrophic mat communities (Bebout and Garcia-Pichel, 1995; Serôdio et al., 2006; Coelho et al., 2011). Similar strategies to optimize photon capture are known in terrestrial plants, where the position of chloroplast in palisade and mesophyll layers in leaves can change depending on light levels and light field directionality, i.e., diffuse versus collimated light (Vogelmann, 1993; Gorton et al., 1999; Wada et al., 2003). Raphidic diatoms, filamentous cyanobacteria and Beggiatoa spp. are able to glide within an extracellular polymeric matrix at speeds of 0–10 μm s^–1 (Glagoleva et al., 1980; Richardson and Castenholz, 1987; Hoiczyk, 2000; Gupta and Agrawal, 2007; Kamp et al., 2008; Tamulonis et al., 2011). Because of the steep light gradient, migration and the resultant vertical redistribution of the productive biomass have important consequences for both the photobiology of the phototrophs and the net primary productivity of the microbial mat ecosystem (Bebout and Garcia-Pichel, 1995; Cartaxana et al., 2016b).

Recent studies have focused on the efficiency with which light is utilized and converted to chemical energy via photosynthesis in cyanobacterial mats and mixed cyanobacteria-diatom biofilms (Al-Najjar et al., 2010, 2012). In these studies, relatively low photosynthetic efficiencies were estimated for microbial mats compared with ecosystems with a more open canopy-like organization such as coarse sediments (Lichtenberg et al., 2017), macroalgal stands (Sand-Jensen et al., 2007), corals (Brodersen et al., 2014), or terrestrial forest communities (Terashima et al., 2016), where light propagation is not hindered to the same extent by self-shading and photosynthetic inactive components. How the photosynthetic efficiency of biofilms and microbial mats is modulated by the migration of motile phototrophic populations remains to be studied in detail. In this study, we used fiber-optic scalar irradiance and field radiance microprobes in combination with O₂ and temperature microsensors to investigate the radiative energy budget for the euphotic zone in a coastal microbial mat dominated by motile cyanobacteria and colorless sulfur bacteria. We investigated how photosynthetic efficiency of oxygenic phototrophs in the microbial mat was affected by changes in vertical biomass distribution imposed by acclimating the microbial mat to different light conditions.

Radiative energy budgets in microbenthic systems have previously been studied in cyanobacterial mats and biofilms (Al-Najjar et al., 2010, 2012), sediments (Lichtenberg et al., 2017), and corals (Brodersen et al., 2014), albeit under the assumption of a homogenous depth distribution of biomass. In the studied microbial mat, dense populations of phototrophic microalgae and cyanobacteria reside in steep gradients of resource stratification shaped by communities of photo-, chemo-, and heterotrophic microorganisms creating a steep redox gradient (van Gemerden, 1993). Consequently, the physical and chemical landscape in such microbial mats can change, within less than one mm, from intense sun exposure (>1000 μmol photons m^–2 s^–1) and O₂ supersaturation (up to 2 mM) to complete darkness and a reduced anoxic sediment high in H₂S (Jørgensen, 1982).

The studied microbial mat exhibited an extremely high optical density and attenuation of PAR, where >99% of the surface irradiance was effectively absorbed <0.4 mm below the microbial mat surface. Light was thus the primary limiting resource for microbial phototrophs in the mat. One strategy to remain competitive under light limitation is to complement light absorption by Chl a, the main light harvesting pigment in oxygenic photosynthesis, by metabolic investment in producing a range of accessory pigments absorbing a broader part of the available light spectrum and channeling it to the photosynthetic reaction centers (Stomp et al., 2007; Trampe and Kühl, 2016; Kühl et al., 2020). HPLC pigment analysis of the upper 0.5 mm of the microbial mat revealed that very efficient light absorption was achieved by a mixture of light harvesting pigments from cyanobacteria (Chl a, mixoxanthophyll, zeaxanthin, oscillaxanthin and β,ε-carotene) and anoxygenic phototrophs (BChl a), able to absorb light from the UV-B well into the NIR region (Figure 3). We did not find any signs of typical diatom light harvesting pigments (fucoxanthin, diadinoxanthin or Chl c) or the far-red absorbing chlorophylls d or f in either measurements of scalar irradiance or by HPLC analysis.

The attenuation of light was stratified with depth and varied between the different acclimation states. In all acclimations, we found strong spectral attenuation from typical cyanobacterial pigments confirmed by the HPLC pigment analysis. In low light acclimated mats the strongest spectral attenuation was in the top 0.2 mm while the strongest spectral attenuation was found deeper (0.3–0.5 mm) in the high light acclimated mats.

Consequently, low surface reflection was observed in the low and high light-acclimated microbial mats. In the low light-acclimated mat, the cyanobacterial population was located near the surface and only 0.75% of the incident irradiance was backscattered, while in the high light-acclimated state, the motile cyanobacteria migrated downward changing the surface biomass composition leading to higher reflectance. During dark-acclimation, the surface of the biofilm became anoxic and a layer of the motile, filamentous colorless sulfur bacteria Beggiatoa formed on the mat surface. Beggiatoa spp. are sulfide oxidizing bacteria known to store granules of elemental sulfur (Nelson and Castenholz, 1981), which make the filaments appear white due to strong light scattering, which in our measurements increased the surface reflectance to 9.2% of the incident irradiance. Multiple scattering in sediments and coral tissues have previously been shown to increase the scalar irradiance in top layers (Kühl and Jørgensen, 1994; Wangpraseurt et al., 2012) which could explain the very low attenuation of light in the top layers of the dark-acclimated mat (Figure 2). Light-dependent migration patterns of cyanobacteria and Beggiatoa thus clearly modulated the radiative energy input to the microbial mat.

The strong light absorption in the uppermost mat layers, resulted in very high local photosynthetic rates reaching >30 nmol O₂ cm^–3 s^–1. This is about 2–5 fold higher than rates measured in most other cyanobacteria-dominated microbial mats (Al-Najjar et al., 2012), but e.g., comparable to rates measured in a highly stratified intertidal mudflat community dominated by motile diatoms (Cartaxana et al., 2016b) and in a cultivated diatom biofilm (Jensen and Revsbech, 1989). We investigated photosynthesis under high light exposure of microbial mats exhibiting different distributions of motile microbes in response to different light history. Investigations of photosynthesis under increasing irradiances in motile microbial systems are complex as migrating cells under changing light can change the spatial characteristics of the biofilm/mat and thereby potentially affect light exposure and distribution in different mat layers (Kühl et al., 1997). In the present study, the main aim was thus to investigate how the migration of motile cyanobacteria affected photosynthetic efficiency under different spatial organization of phototrophs in the microbial mat.

This is experimentally challenging, and we note that these measurements were not performed under steady-state conditions. The change to higher light than the acclimation irradiance will start to move the mat toward a new steady-state and rates and profiles will thus be measured under transient “quasi steady-state” conditions. The measurements of scalar irradiance and gross photosynthesis reflect the community’s response to high light in the current distribution of biomass. The distribution of O₂ and temperature is, however, more complex to interpret, as the distribution of O₂ is determined by the interplay between diffusion, production and consumption, while the temperature profiles are determined by the local heat dissipation and heat transfer properties. These are slower processes and will move toward a steady state on the same time scale as motility driven biomass redistribution in such compact systems as the studied mats.

Areal rates of oxygenic photosynthesis, integrated across the entire euphotic zone, did not differ markedly between different light acclimations (0.99, 1.11, and 0.84 nmol O₂ cm^–2 s^–1 for dark-, low light- and high light-acclimated samples, respectively) under the high saturating photon irradiance used for the energy budget measurements. It is however noted that the lowest areal photosynthetic rates were observed in the high light-acclimated state. One could speculate that the longer acclimation period to high light could have given the phototrophs, in this treatment, time to adjust their balance between photosynthesis and non-photochemical quenching (NPQ) processes. One of the most important short-term regulatory mechanisms to avoid photo damage is NPQ, where excess energy decays from singlet excited chlorophyll (¹Chl^∗) into heat dissipation (Müller et al., 2001). This mechanism significantly lowers the effective quantum yield of photosynthesis but avoids the formation of reactive singlet oxygen (¹O₂^∗) by the triplet state of chlorophyll (³Chl^∗) (Müller et al., 2001), which can have long term detrimental effects on the photosystems by degradation of the D1 protein; an important component in photosystem II (Nymark et al., 2009). The higher rates displayed by the top population for the dark- and low-light acclimated mats could therefore be a product of the more open reaction centers before regulating the different NPQ components that operate on time scales from a few minutes to a few hours (Kress and Jahns, 2017).

Photophobic responses or phototaxis enabling movement along a light gradient (Jekely, 2009) are alternative strategies of photoprotection, enabling motile phototrophs to align their position at optimum irradiance in light gradients (Tamulonis et al., 2011) depending on the status of the electron transport chain and time of the day (Burns and Rosa, 1980). Microcoleus chtonoplastes displays light dependent migration preferring moderate to low light levels (Ramsing and Prufert-Bebout, 1994), and other studies have found that as little as a 4% difference in light intensity between “head” and “tail” of a cyanobacterial filament can trigger their movement (Häder, 1987). From our measured depth profiles of gross oxygenic photosynthesis, it appears that the distance of vertical migration was limited to only 0.2 to 0.3 mm. However, due to the steep light gradients in the investigated microbial mat, the changes in the light field over such short distances are dramatic, and cyanobacteria migrating 200 μm deeper into the mat experienced a >84-fold decrease in scalar irradiance (Figure 2C; 0.4–0.2 mm) from >350 μmol photons m^–2 s^–1 to < 10 μmol photons m^–2 s^–1. Such strong changes in light exposure upon migration distances of a few 100 μm have previously been demonstrated in hypersaline cyanobacterial mats (Kühl et al., 1997). Consequently, the spatial distribution of gross photosynthesis varied with light acclimation, probably as a result of migrations of the cyanobacteria and Beggiatoa.

Other triggers for migration could also play a role in the observed redistribution of biomass. Sulfide is well known to cause migration due to its cytotoxicity (Whale and Walsby, 1984). Sulfide was most likely present in high concentrations in the studied mats due to i) the black FeS layer below the photic zone, ii) the presence of BChl a, indicative of anoxygenic phototrophs that oxidize hydrogen sulfide to sulfur, and the presence of a dense population of Beggiatoa known to thrive at the interface of strong O₂ and sulfide concentration gradients. Earlier microsensor measurements in microbial mats from the same sampling site also demonstrated significant sulfide production and light-driven dynamics of O₂, pH and sulfide in the uppermost mat layers (Nielsen et al., 2015). As stated above, Microcoleus prefer moderate to low light levels and their migratory behavior is thus probably due to a combination of photophobic movement and sulfide avoidance, although they can grow in H₂S concentrations up to 974 μM (de Wit and van Gemerden, 1987).

Compared to earlier studies of heat dissipation in corals and microbial mats (Jimenez et al., 2008; Al-Najjar et al., 2010; Brodersen et al., 2014), we found a relatively small heat dissipation from the biofilm surface into the water column and the surface temperature of the microbial mat increased by only ∼0.3°C. However, the heat dissipation in deeper mat layers was larger than expected. The zone of maximum temperature increase was expected to be found in the same zone as the largest energy deposition, i.e., in the upper few hundred micrometers of the microbial mat, where >90% of the incident light was attenuated. However, the heat dissipation below 0.5 mm depth was in the same order of magnitude as the upward heat dissipation and reached higher temperatures (Figure 4). We note that below the dense photic zone, a black layer of precipitated iron sulfide was found, and absorption of light energy in this sediment layer apparently contributed significantly to the heating of the microbial mat, but this remains speculative. While we did not measure temperature deep enough in the sediment to be able to calculate the downward transport of heat, we estimated from the principle of energy conservation that the downward heat flux contributed about 20–50% of the total heat dissipation.

The amount of photochemically conserved energy did not change markedly between different light acclimation states of the microbial mat when investigated at similar high irradiance, despite the higher reflection and lower heat dissipation observed in the dark-acclimated biofilms. Consequently, the photosynthetic energy conservation was 2.1, 2.4, and 1.8% of the incident energy, for dark-, low light-, and high light-acclimated biofilms, respectively. This indicates that by migration, the phototrophic community can apparently sustain similar energy conservation efficiencies, while avoiding the detrimental effects of excessive light when situated near the surface.

The vertical stratification of relative photosynthetic efficiencies (Figure 6) can either be ascribed to (i) a higher density of biomass contributing to photosynthesis, or (ii) an overall higher efficiency caused by higher cell specific pigment content and/or higher absorption cross section, in that area (Iglesias-Prieto and Trench, 1994, 1997; Falkowski and Raven, 2007; Al-Najjar et al., 2010). However, in our measurements we found differences in the vertical positions of the peaks of relative photosynthetic efficiency depending on light acclimation. At the cellular level, regulation of the pigment density and the absorption cross section of antennae pigments occurs on a longer time scale than the acclimation time used in these experiments, although the formation of zeaxanthin from β-carotene is a short term photoprotective mechanism (Falkowski and Raven, 2007). Given the time scale, the difference in vertical position of the peaks in relative photosynthetic efficiencies, can thus primarily be ascribed to migration.

In a fluctuating light environment, phototrophs can modulate light exposure/harvesting either by regulating pigment densities or by moving to a different light environment. For this mechanism to be energetically successful, the cost of motility must present an advantage compared to employment of regulation of light harvesting or photoprotective pigments. The cost of motility was estimated for Oscillatoria to be 0.2–5% of the energy generated by oxidative phosphorylation (Halfen and Castenholz, 1971). This is a relatively low cost considering that in a dense microbial system, where oxygenic phototrophs not only have to cope with alternating light environments but also steep and dynamic chemical gradients of e.g., pH and sulfide. Motility may thus be an important trait for phototrophs in microbial mats enabling rapid optimization of their light exposure and chemical microenvironment. Migration of cyanobacterial populations can also mitigate effects of continuous sedimentation and overgrowth by other microbes (Whale and Walsby, 1984).

Previously, photosynthetic quantum efficiencies in microbial mats have been calculated by applying a model that estimates the local quantity of absorbed light, which is then correlated to the local rates of gross photosynthesis (Al-Najjar et al., 2010). However, this model assumes a homogenous distribution of light absorption (i.e., biomass), which makes the application of such a model problematic in a motile community exhibiting photomovement. Calculations using the model of Al-Najjar et al. (2010) on our data showed apparent photosynthetic quantum efficiencies, about 2-fold higher than the theoretical maximum of 0.125 (i.e., 8 photons needed to produce 1 O₂ molecule). We speculate that the presence of a black ferrous sulfide layer in close proximity to the phototrophs also interfered with such estimations of the local density of absorbed light. However, the distribution of sand particles, organic matter and photosynthetic biomass was not further quantified, and a correction for this possible artifact was not attempted. Therefore, we estimated relative quantum efficiencies of photosynthesis by relating the depth specific gross photosynthesis rates to the scalar irradiance incident to that point following the approach of Lassen et al. (1992), who also found that the position of maximum quantum efficiencies changed with light exposure. The relative quantum efficiencies estimated in our mat samples were, however, up to three times higher than values reported by Lassen et al. (1992) for a less compacted microbial mat.

The photosynthetic efficiency depends on photo-pigmentation and other biotic/abiotic substances contributing to light absorption (Al-Najjar et al., 2012) but also on complex interplay between cyanobacteria and sulfur oxidizers (Klatt et al., 2016). Thus, the microscale 3D structure of the microbial mat will have an influence on the light availability and the energy balance of the system, where more open systems will display larger photic zones and different responses to changes in light environment as compared to more compacted systems (Lichtenberg et al., 2017). How such microscale canopy-like effects in microbial mats are affected by the presence of much higher densities of abiotic matter in comparison to e.g., plant canopies remain to be studied in more detail.

We conclude that cyanobacteria can sustain high relative photosynthetic efficiencies by vertical migration in response to shifting light conditions, thereby optimizing the overall capacity of light absorption and photosynthetic activity against a backdrop of dynamic chemical gradients of O₂, pH and sulfide. This was evident from the overall photosynthetic energy conservation efficiency that did not change with light acclimation but showed vertical differences in the distribution of maximum relative photosynthetic efficiency. Further studies could investigate how distribution and efficiency of gross photosynthesis respond to changing light environment in microbial mats with vertically fixed populations of phototrophs in different depth horizons, e.g., by applying a motility inhibitor (Cartaxana et al., 2016a). In very compact systems such as the mats studied here it is, however, difficult to resolve local differences due to the spatial resolution of the microsensors and methodology to non-invasively monitor biomass distributions. In less compacted systems (e.g., mats like studied by Wieland and Kühl, 2000; Saenger et al., 2006; Lichtenberg et al., 2017) with more open structures and thereby less steep gradients of the physico-chemical parameters, local differences in energy deposition would be easier to describe in more detail.

Our study adds novel insight to the regulation of photosynthesis in compacted microbial mats and biofilms. In terms of energy conservation and photosynthetic efficiency, such systems are often regarded less efficient due to their more or less flat topography (e.g., Al-Najjar et al., 2012). However, there is increasing evidence that microstructural changes can modulate light harvesting and photosynthesis in dense photosynthetic biofilms and tissues, and that such systems can exhibit canopy-like properties at a microscale, where a more open microstructure and/or modulation of the scattering and lateral distribution of incident light seems to govern higher photosynthetic efficiency (e.g., Wangpraseurt et al., 2014; Lichtenberg et al., 2016, 2017; Lyndby et al., 2016). In the present study, we show that reorganization of the phototrophs via migration is yet another functional trait that helps optimize overall light harvesting and photosynthesis in biofilms and microbial mats.

The datasets generated for this study are available on request to the corresponding author.

ML, PC, and MK designed the experiments and analyzed the data. ML and PC performed the experiments. ML wrote the manuscript with editorial input from PC and MK.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.