Benthic dinitrogen fixation in a Northern Red Sea coral reef under seasonally changing environmental conditions

Tropical coral reefs are among the most productive ecosystems on this planet, despite being surrounded by very oligotrophic waters. Effective recycling processes of the limiting nutrients, particularly nitrogen (N), and input of new bioavailable N via dinitrogen (N2) fixation are essential to sustain such high gross primary production. In fact, several benthic reef organisms and substrates are associated with diverse communities of N2 fixing microbes (diazotrophs), but the respective contribution of the different benthic groups to total benthic N2 fixation and the effect of changing environmental conditions on N2 fixation have not been investigated yet. Therefore, this thesis, through a series of interconnected studies carried out in a seasonally dynamic coral reef system, the Northern Red Sea, and in a number of manipulative experiments, contributes to the understanding of benthic N2 fixation in coral reefs by answering the following key questions: 1) How much N2 is fixed by the dominant benthic groups in the Northern Red Sea? 2) What is the relative contribution of the benthic groups to total benthic N2 fixation within the reef? 3) What is the effect of seasonally changing environmental conditions and of single environmental factors on key metabolic processes, particularly N2 fixation, associated with the different benthic groups? Findings revealed that all investigated benthic groups showed N2 fixation activity, whereof bare coral rock, turf algae, carbonate sands and living hard corals were the main N2 fixing components contributing ~ 90% to benthic N2 fixation in investigated Red Sea coral reefs. Soft corals revealed the lowest N2 fixation activity among all investigated groups and released significantly less organic matter to the surrounding water compared to hard corals indicating that N2 fixation may also influence their role as allogenic ecosystem engineers. N2 fixation by most benthic groups was usually highest when nutrient availability was lowest, and water temperature as well as light intensity highest suggesting that the N2 fixation products fuel the metabolic N requirements of reef organisms, particularly during nutrient-depleted conditions (i.e. summer). The manipulation of single environmental factors revealed a stimulation of N2 fixation activities under global warming conditions and a reduction under ocean acidification scenarios indicating that global climate change will affect the nutrient status of reef organisms. In summary, this thesis underlines the ubiquity of N2 fixation associated with different benthic coral reef organisms and substrates, and highlights its importance in sustaining coral reef productivity. Both, the benthic community structure and the prevailing environmental conditions appear to be important in controlling the amount of N2 fixation in coral reefs. Finally, as coral reefs are increasingly and simultaneously exposed to global and local stressors, it is necessary to understand how important metabolic processes on coral reefs (i.e. organic matter fluxes and N2 fixation activity) will be affected, and this thesis provides first quantitative insights into these processes from physiological, biogeochemical and ecological points of view

Studies With Soft Corals – Recommendations on Sample Processing and Normalization Metrics

Soft corals (Octocorallia) often constitute the second most abundant macrobenthic group on many tropical and temperate reefs. However, the gelatinous and leather-like nature of their tissue and their variable hydroskeleton entails a number of problems for tissue homogenization and data normalization. An easy and fast protocol for tissue homogenization, as well as a normalization metric that can be used to perform inter-studies or inter-species comparisons, are thus needed. In this study, we tested whether the tissue sample state before processing (frozen vs. freeze-dried samples) and the media used for tissue homogenization (0.2 μm filtered seawater; FSW vs. Milli-Q water; DI) affect the quantitative measurements of tissue descriptors (chlorophyll, protein, and Symbiodinium concentrations) in the model species Heteroxenia fuscescens. Furthermore, the suitability of dry weight (DW) and ash-free dry weight (AFDW) as size-normalizing metric was investigated across different soft coral species. Our results reveal that freeze-drying the samples and homogenizing them in DI water exhibited several benefits, namely enhancing chlorophyll and protein concentrations up to 50%, saving processing time and providing a more accurate determination of DW and AFDW. Overall, this optimized tissue processing protocol offers a more reliable quantification of tissue descriptors and reduces the chance of underestimating these parameters in soft corals. Finally, since the contribution of sclerites to the total DW of the colony can highly differ between species, we demonstrate that AFDW is a reliable metric for normalizing soft coral data, particularly when inter-species comparisons are made

Effects of Water Column Mixing and Stratification on Planktonic Primary Production and Dinitrogen Fixation on a Northern Red Sea Coral Reef

The northern Red Sea experiences strong annual differences in environmental conditions due to its relative high-latitude location for coral reefs. This allows the study of regulatory effects by key environmental parameters (i.e., temperature, inorganic nutrient, and organic matter concentrations) on reef primary production and dinitrogen (N2) fixation, but related knowledge is scarce. Therefore, this study measured environmental parameters, primary production and N2 fixation of phytoplankton groups in the water overlying a coral reef in the Gulf of Aqaba. To this end, we used a comparative approach between mixed and stratified water column scenarios in a full year of seasonal observations. Findings revealed that inorganic nutrient concentrations were significantly higher in the mixed compared to the stratified period. While gross photosynthesis and N2 fixation rates remained similar, net photosynthesis decreased from mixed to stratified period. Net heterotrophic activity of the planktonic community increased significantly during the stratified compared to the mixed period. While inorganic nitrogen (N) availability was correlated with net photosynthesis over the year, N2 fixation only correlated with N availability during the mixed period. This emphasizes the complexity of planktonic trophodynamics in northern Red Sea coral reefs. Comparing mixed and stratified planktonic N2 fixation rates with those of benthic organisms and substrates revealed a close seasonal activity similarity between free-living pelagic and benthic diazotrophs. During the mixed period, N2 fixation potentially contributed up to 3% of planktonic primary production N demand. This contribution increased by ca. one order of magnitude to 21% during the stratified period. Planktonic N2 fixation is likely a significant N source for phytoplankton to maintain high photosynthesis under oligotrophic conditions in coral reefs, especially during stratified conditions

Spatio-temporal patterns in the coral reef communities of the Spermonde Archipelago, 2012–2014, II: Fish assemblages display structured variation related to benthic condition

The Spermonde Archipelago is a complex of ~70 mostly populated islands off Southwest Sulawesi, Indonesia, in the center of the Coral Triangle. The reefs in this area are exposed to a high level of anthropogenic disturbances. Previous studies have shown that variation in the benthos is strongly linked to water quality and distance from the mainland. However, little is known about the fish assemblages of the region and if their community structure also follows a relationship with benthic structure and distance from shore. In this study, we used eight islands of the archipelago, varying in distance from 1 to 55 km relative to the mainland, and 3 years of surveys, to describe benthic and fish assemblages and to examine the spatial and temporal influence of benthic composition on the structure of the fish assemblages. Cluster analysis indicated that distinct groups of fish were associated with distance, while few species were present across the entire range of sites. Relating fish communities to benthic composition using a multivariate generalized linear model confirmed that fish groups relate to structural complexity (rugosity) or differing benthic groups; either algae, reef builders (coral and crustose coralline algae) or invertebrates and rubble. From these relationships we can identify sets of fish species that may be lost given continued degradation of the Spermonde reefs. Lastly, the incorporation of water quality, benthic and fish indices indicates that local coral reefs responded positively after an acute disturbance in 2013 with increases in reef builders and fish diversity over relatively short (1 year) time frames. This study contributes an important, missing component (fish community structure) to the growing literature on the Spermonde Archipelago, a system that features environmental pressures common in the greater Southeast Asian region

Spatio-temporal patterns in coral reef communities of the Spermonde Archipelago, 2012-2014, I: Comprehensive reef monitoring of water and benthic indicators reflect changes in reef health

Pollution, fishing, and outbreaks of predators can heavily impact coastal coral reef ecosystems, leading to decreased water quality and benthic community shifts. To determine the main environmental drivers of coral reef status in the Spermonde Archipelago, Indonesia, we monitored environmental variables and coral reef benthic community structure along an on-to-offshore gradient annually from 2012 to 2014. Findings revealed that concentrations of phosphate, chlorophyll a-like fluorescence, suspended particulate matter, and light attenuation significantly decreased from on-to-offshore, while concentrations of dissolved O2 and values of water pH significantly increased on-to-offshore. Nitrogen stable isotope signatures of sediment and an exemplary common brown alga were significantly enriched nearshore, identifying wastewater input from the city of Makassar as primary N source. In contrast to the high temporal variability in water quality, coral reef benthic community cover did not show strong temporal, but rather, spatial patterns. Turf algae was the dominant group next to live coral, and was negatively correlated to live coral, crustose coralline algae (CCA), rubble and hard substrate. Variation in benthic cover along the gradient was explained by water quality variables linked to trophic status and physico-chemical variables. As an integrated measure of reef status and structural complexity, the benthic index, based on the ratio of relative cover of live coral and CCA to other coral reef organisms, and reef rugosity were determined. The benthic index was consistently low nearshore and increased offshore, with high variability in the midshelf sites across years. Reef rugosity was also lowest nearshore and increased further offshore. Both indices dropped in 2013, increasing again in 2014, indicating a period of acute disturbance and recovery within the study and suggesting that the mid-shelf reefs are more resilient to disturbance than nearshore reefs. We thus recommend using these two indices with a selected number of environmental variables as an integral part of future reef monitoring

Benthische Stickstofffixierung in einem Korallenriff des nördlichen Roten Meeres unter saisonal sich verändernden Umweltbedingungen

Tropical coral reefs are among the most productive ecosystems on this planet, despite being surrounded by very oligotrophic waters. Effective recycling processes of the limiting nutrients, particularly nitrogen (N), and input of new bioavailable N via dinitrogen (N2) fixation are essential to sustain such high gross primary production. In fact, several benthic reef organisms and substrates are associated with diverse communities of N2 fixing microbes (diazotrophs), but the respective contribution of the different benthic groups to total benthic N2 fixation and the effect of changing environmental conditions on N2 fixation have not been investigated yet. Therefore, this thesis, through a series of interconnected studies carried out in a seasonally dynamic coral reef system, the Northern Red Sea, and in a number of manipulative experiments, contributes to the understanding of benthic N2 fixation in coral reefs by answering the following key questions: 1) How much N2 is fixed by the dominant benthic groups in the Northern Red Sea? 2) What is the relative contribution of the benthic groups to total benthic N2 fixation within the reef? 3) What is the effect of seasonally changing environmental conditions and of single environmental factors on key metabolic processes, particularly N2 fixation, associated with the different benthic groups? Findings revealed that all investigated benthic groups showed N2 fixation activity, whereof bare coral rock, turf algae, carbonate sands and living hard corals were the main N2 fixing components contributing ~ 90% to benthic N2 fixation in investigated Red Sea coral reefs. Soft corals revealed the lowest N2 fixation activity among all investigated groups and released significantly less organic matter to the surrounding water compared to hard corals indicating that N2 fixation may also influence their role as allogenic ecosystem engineers. N2 fixation by most benthic groups was usually highest when nutrient availability was lowest, and water temperature as well as light intensity highest suggesting that the N2 fixation products fuel the metabolic N requirements of reef organisms, particularly during nutrient-depleted conditions (i.e. summer). The manipulation of single environmental factors revealed a stimulation of N2 fixation activities under global warming conditions and a reduction under ocean acidification scenarios indicating that global climate change will affect the nutrient status of reef organisms. In summary, this thesis underlines the ubiquity of N2 fixation associated with different benthic coral reef organisms and substrates, and highlights its importance in sustaining coral reef productivity. Both, the benthic community structure and the prevailing environmental conditions appear to be important in controlling the amount of N2 fixation in coral reefs. Finally, as coral reefs are increasingly and simultaneously exposed to global and local stressors, it is necessary to understand how important metabolic processes on coral reefs (i.e. organic matter fluxes and N2 fixation activity) will be affected, and this thesis provides first quantitative insights into these processes from physiological, biogeochemical and ecological points of view

Diazotrophs: Overlooked Key Players within the Coral Symbiosis and Tropical Reef Ecosystems?

International audienceCoral reefs are highly productive ecosystems thriving in nutrient-poor waters. Their productivity depends largely on the availability of nitrogen, the proximate limiting nutrient for primary production. In reefs, the major nitrogen pathways include regeneration, nitrification, ammonification and dinitrogen (N 2) fixation. N 2 fixation is performed by prokaryotes called " diazotrophs " that abound in coral rubbles, sandy bottoms, microbial mats, or seagrass meadows. Corals, which are the main reef builders, have developed a partnership with dinoflagellates which transform dissolved inorganic nitrogen into amino acids and protein, and with diazotrophs to gain diazotrophically-derived nitrogen (DDN). Pioneering studies found active diazotrophic cyanobacteria in the corals' mucus and/or tissue, and later high throughput sequencing efforts have described diverse communities of non-cyanobacterial diazotrophs associated with scleractinian corals. Yet, the metabolic processes behind these associations and how they benefit corals are currently not well understood. While genomic studies describe the diversity of diazotrophs and isotopic tracer experiments quantify N 2 fixation rates, combined advanced methods are needed to elucidate the mechanisms behind the transfer of DDN to the coral holobiont and whether these mechanisms change according to the identity of the diazotrophs or coral species. Here we review our current knowledge on N 2 fixation in corals: the diversity and localization of diazotrophs in the coral holobiont, the environmental factors controlling N 2 fixation, the fate of DDN within the coral symbiosis as well as its potential role in coral resilience. We finally summarize the unknowns: are the diversity, abundance and localization of diazotrophs within the holobiont species-and/or site-specific? Do they have an impact on DDN production? What are the metabolic mechanisms implicated? Do they change spatially, temporally or according to environmental factors? We encourage scientists to undertake research efforts to tackle these questions in order to shed light on nitrogen cycling in reef ecosystems and to understand if coral-associated N 2 fixation can improve coral's resilience in the face of climate change

Ocean acidification rapidly reduces dinitrogen fixation associated with the hermatypic coral Seriatopora hystrix

Since productivity and growth of coral-associated dinoflagellate algae is nitrogen (N)-limited, dinitrogen (N2) fixation by coral-associated microbes is likely crucial for maintaining the coral-dinoflagellate symbiosis. It is thus essential to understand the effects future climate change will have on N2 fixation by the coral holobiont. This laboratory study is the first to investigate short-term effects of ocean acidification on N2 fixation activity associated with the tropical, hermatypic coral Seriatopora hystrix using the acetylene reduction assay in combination with calcification measurements. Findings reveal that simulated ocean acidification ( pCO2 1080 µatm) caused a rapid and significant decrease (53%) in N2 fixation rates associated with S. hystrix compared to the present day scenario ( pCO2 486 µatm). In addition, N2 fixation associated with the coral holobiont showed a positive exponential relationship with its calcification rates. This suggests that even small declines in calcification rates of hermatypic corals under high CO2 conditions may result in decreased N2 fixation activity, since these 2 processes may compete for energy in the coral holobiont. Ultimately, an intensified N limitation in combination with a decline in skeletal growth may trigger a negative feedback loop on coral productivity exacerbating the negative long-term effects of ocean acidification

Divergent Capacity of Scleractinian and Soft Corals to Assimilate and Transfer Diazotrophically Derived Nitrogen to the Reef Environment

Corals are associated with dinitrogen (N-2)-fixing bacteria that potentially represent an additional nitrogen (N) source for the coral holobiont in oligotrophic reef environments. Nevertheless, the few studies investigating the assimilation of diazotrophically derived nitrogen (DDN) by tropical corals are limited to a single scleractinian species (i.e., Stylophora pistillata). The present study quantified DDN assimilation rates in four scleractinian and three soft coral species from the shallow waters of the oligotrophic Northern Red Sea using the N-15(2) tracer technique. All scleractinian species significantly stimulated N, fixation in the coral-surrounding seawater (and mucus) and assimilated DDN into their tissue. Interestingly, N-2 fixation was not detected in the tissue and surrounding seawater of soft corals, despite the fact that soft corals were able to take up DDN from a culture of free-living diazotrophs. Soft coral mucus likely represents an unfavorable habitat for the colonization and activity of diazotrophs as it contains a low amount of particulate organic matter, with a relatively high N content, compared to the mucus of scleractinian corals. In addition, it is known to present antimicrobial properties. Overall, this study suggests that DDN assimilation into coral tissues depends on the presence of active diazotrophs in the coral's mucus layer and/or surrounding seawater. Since N is often a limiting nutrient for primary productivity in oligotrophic reef waters, the divergent capacity of scleractinian and soft corals to promote N-2 fixation may have implications for N availability and reef biogeochemistry in scleractinian versus soft coral-dominated reefs