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Nitrate and ammonium are the most important nitrogen sources for phytoplankton growth. Differential utilization of inorganic nitrogenous compounds by phytoplankton has been observed and may have significant impacts on primary productivity at local scales. We used enrichment experiments with natural phytoplankton populations from the freshwater tidal zone of the Guadiana estuary, a coastal ecosystem increasingly subjected to anthropogenic influences, to study the effects of nitrate and ammonium on N-consumption and phytoplankton growth. In addition, we used combined additions of nitrate and ammonium to understand the inhibitory effect of ammonium over nitrate uptake. Ammonium concentrations in the freshwater tidal reaches of the Guadiana estuary throughout the sampling period were too low to exert an inhibitory effect on nitrate uptake or a toxic effect on phytoplankton growth. Nitrate was clearly the main nitrogen source for phytoplankton at the study site. Overall, nitrate seemed to become limiting at concentrations lower than 20 μM and N-limitation was particularly significant during summer. A trend of decreasing nitrate uptake with increasing ammonium concentrations and uptake suggested an overall preference for ammonium. However, preference for ammonium was group-specific, and it was observed mainly in green algae and cyanobacteria. In fact, cyanobacteria relied only on ammonium as their N-source. On the contrary, diatoms preferred nitrate, and did not respond to ammonium additions. The increasing eutrophication in the Guadiana estuary and particularly increased inputs of nitrogen as ammonium due to urban waste effluents may result in a shift in phytoplankton community composition, towards a dominance of cyanobacteria and green algae.

Hematopoietic stem cells (HSCs) produce all essential cellular components of the blood. Stromal cell lines supporting HSCs follow a vascular smooth muscle cell (vSMC) differentiation pathway, suggesting that some hematopoiesis-supporting cells originate from vSMC precursors. These pericyte-like precursors were recently identified in the aorta-gonad-mesonephros (AGM) region; however, their role in the hematopoietic development in vivo remains unknown. Here, we identify a subpopulation of NG2 + Runx1 + perivascular cells that display a sclerotome-derived vSMC transcriptomic profile. We show that deleting Runx1 in NG2 + cells impairs the hematopoietic development in vivo and causes transcriptional changes in pericytes/vSMCs, endothelial cells and hematopoietic cells in the murine AGM. Importantly, this deletion leads also to a significant reduction of HSC reconstitution potential in the bone marrow in vivo. This defect is developmental, as NG2 + Runx1 + cells were not detected in the adult bone marrow, demonstrating the existence of a specialised pericyte population in the HSC-generating niche, unique to the embryo. Hematopoietic stem cells are supported by niche cells that help balance stem cell self-renewal and differentiation. Here they show that Runx1 deletion in the embryonic perivascular HSC niche impairs hematopoietic development in vivo and causes transcriptional changes in pericytes/vSMCs, endothelial cells and hematopoietic cells in the murine AGM.

It is generally assumed that the larger the bottle volume, the longer the duration of phytoplankton microcosm experiments. We hypothesize that volume and duration are independent, as volume does not regulate the extension of the exponential growth phase. We conducted two microcosm experiments using 1, 2, and 8 L bottles, inoculated with phytoplankton collected in the Ria Formosa lagoon (SE Portugal) and incubated for 1, 2, 4, and 8 days. Phytoplankton net growth rates were estimated using chlorophyll a concentration and cell abundance, determined with epifluorescence and inverted microscopy. Results show that the experimental duration significantly affected net growth rates, independently of volume, with decreasing net growth rates with time. Regarding volume, we found significant, but weak, differences in net growth rates, and significant two-way interactions only for the larger-sized cells. No significant differences in net growth rates across the different volumes were detected for the smaller, most abundant taxa and for the whole assemblage. We conclude that duration, not volume, is the main factor to consider in microcosm experiments, and it should allow the measurement of responses during the exponential growth phase, which can be detected through daily sampling throughout the duration of the experiment.

Planktonic heterotrophic prokaryotes (HProks) are a pivotal functional group in marine ecosystems and are highly sensitive to environmental variability and climate change. This study aimed to investigate the short-term effects of increasing carbon dioxide (CO2), ultraviolet radiation (UVR), and temperature on natural assemblages of HProks in the Ria Formosa coastal lagoon during winter. Two multi-stressor microcosm experiments were used to evaluate the isolated and combined effects of these environmental changes on HProk abundance, production, growth, and mortality rates. The isolated and combined effects of increased CO2 on HProks were not significant. However, HProk production, cellular activity, instantaneous growth rate, and mortality rate were negatively influenced by elevated UVR and positively influenced by warming. Stronger effects were detected on HProk mortality in relation to specific growth rate, leading to higher HProk net growth rates and abundance under elevated UVR and lower values under warming conditions.

The Ria Formosa coastal lagoon is a highly productive shallow ecosystem in southern Portugal, subjected to nutrient inputs from anthropogenic and natural sources. Nutrients are major abiotic drivers of phytoplankton in this system, but their effects on phytoplankton assemblages and the occurrence of nutrient limitation are still poorly understood. The main goal of this study was, thus, to evaluate the occurrence, type, and effects of nutrient limitation on phytoplankton community and specific functional groups in the Ria Formosa coastal lagoon. We conducted nutrient enrichment experiments with factorial additions of nitrogen (N) and phosphorus (P) using natural phytoplankton assemblages from distinct locations in the Ria Formosa, throughout a yearly cycle. Phytoplankton composition and abundance were evaluated using inverted and epifluorescence microscopies, and spectrophotometric methods were used for biomass. Limitation was defined as higher phytoplankton growth following enrichment with a particular nutrient in relation to the non-enriched control. The most common type of phytoplankton limitation was simultaneous co-limitation by N and P; diatoms, as r-strategists, were the most frequently limited group. Single N and P limitation, and serial P limitation were also observed, as well as negative responses to nutrient enrichment. Group-specific responses to nutrient enrichment were not reflected in the relative abundance of phytoplankton groups within the whole assemblage, due to the numerical dominance of pico-sized groups (cyanobacteria and eukaryotic picophytoplankton). Ambient nutrient ratios and concentrations did not predict phytoplankton nutrient limitation, given the different nutrient utilisation traits among phytoplankton functional groups. Therefore, nutrient ratios should not be used as indicators of nutrient limitation in eutrophication assessment.

The use of several light-related variables, such as the Secchi disc depth, the euphotic depth, and in particular, the diffuse attenuation coefficient (Kd), is deeply rooted in phytoplankton research, but these are not the most appropriate indicators of the amount of light available for photosynthesis. We argue that the variable of interest for phytoplankton is the mean light intensity in the mixed layer (Im), which represents the mean light to which phytoplankton cells are exposed throughout their life cycle, while being continuously mixed in the mixed layer. We use empirical data collected in different coastal ecosystems in southern Portugal to demonstrate why Im should be the preferred metric instead of the deeply rooted Kd. We show that, although the relationship between Im and Kd is inversely proportional, it is not always strong or even significant. Different Im values can be associated with the same Kd, but distinct Im have different physiological effects of phytoplankton. Therefore, Kd does not capture the amount of light available for photosynthesis, given that, unlike Im, Kd calculation does not consider the depth of the mixed layer. Therefore, we urge phytoplankton researchers to consider the measurement and calculation of Im when evaluating light-related processes in phytoplankton ecology.

Changes in temperature and CO2 are typically associated with climate change, but they also act on shorter time scales, leading to alterations in phytoplankton physiology and community structure. Interactions among stressors may cause synergistic or antagonistic effects on phytoplankton dynamics. Therefore, the main goal of this work is to understand the short-term isolated and interactive effects of warming and high CO2 on phytoplankton nutrient consumption, growth, production, and community structure in the Ria Formosa coastal lagoon (southern Portugal). We performed microcosm experiments with temperature and CO2 manipulation, and dilution experiments under temperature increase, using winter phytoplankton assemblages. Phytoplankton responses were evaluated using inverted and epifluorescence microscopy. Overall, phytoplankton growth and microzooplankton grazing on phytoplankton decreased with warming. Negative antagonist interactions with CO2 alleviated the negative effect of temperature on phytoplankton and cryptophytes. In contrast, higher temperature benefited smaller-sized phytoplankton, namely cyanobacteria and eukaryotic picophytoplankton. Diatom growth was not affected by temperature, probably due to nutrient limitation, but high CO2 had a positive effect on diatoms, alleviating the effect of nutrient limitation. Results suggest that this winter phytoplankton assemblage is well acclimated to ambient conditions, and short-term increases in temperature are detrimental, but can be alleviated by high CO2.

Phytoplankton are dominant primary producers and key indicators in aquatic ecosystems. Understanding the controlling factors on the structure of phytoplankton assemblages is fundamental, but particularly challenging at the land–ocean interface. To identify the patterns and predictors of phytoplankton assemblage structure in the Ria Formosa coastal lagoon (south Portugal), this study combined phytoplankton abundance along a transect between the discharge point of a wastewater treatment plant and a lagoon inlet, over two years, with physico-chemical, hydrographic, and meteo-oceanographic variables. Our study identified 147 operational taxonomic units (OTUs), and planktonic diatoms (60–74%) and cryptophyceans (17–25%) dominated the phytoplankton in terms of abundance. Despite strong lagoon hydrodynamics, and the lack of spatial differences in the phytoplankton abundance and most diversity metrics, the multivariate analysis revealed differences in the assemblage structure between stations (p < 0.001) and seasons (p < 0.01). Indicator analysis identified cryptophyceans as lagoon generalists, and 11 station-specific specialist OTUs, including Kryptoperidinium foliaceum and Oscillatoriales (innermost stations) and potentially toxigenic species (Pseudo-nitzschia and Dinophysis; outer lagoon station, p < 0.05). Water temperature, pH, and nutrients emerged as the variables that best explained the changes in the phytoplankton assemblage structure (p < 0.001). Our findings provide insight into the relevance of local anthropogenic and natural forcings on the phytoplankton assemblage structure and can be used to support the management of RF and other coastal lagoons.

Jacob, J.; Correia, C.; Torres, A.F.; Xufre, G.; Matos, A.; Ferreira, C.; Reis, M.P.; Caetano, S.; Freitas, C.S.; Barbosa, A.B., and Cravo, A., 2020. Impacts of decommissioning and upgrading urban wastewater treatment plants on the water quality in a shellfish farming coastal lagoon (Ria Formosa, South Portugal). In: Malvárez, G. and Navas, F. (eds.), Global Coastal Issues of 2020. Journal of Coastal Research, Special Issue No. 95, pp. 45–50. Coconut Creek (Florida), ISSN 0749-0208. Ria Formosa is a productive coastal lagoon, located on the south coast of Portugal, and represents the largest national producer of shellfish bivalves (ca. 90% production). This ecosystem is subjected to various anthropogenic pressures, including the discharge of urban wastewater treatment plants (UWWTP), which impacts the lagoon water quality. This study aimed to assess the impact of alterations in the functioning of two UWWTP on the water quality of Ria Formosa, based on chemical variables, phytoplankton composition (including potential harmful species) and faecal contamination. During the period September 2018 - October 2019, water sampling was conducted along dominant longitudinal gradients of the effluent dispersion from the discharge point (1-2 km), for two sites: a decommissioned (OP) and a modified (FO) UWWTP. After modification, the later started receiving a higher influent volume (ca. 40%), under an innovative technology system (biological treatment in aerobic granular sludge). Based on chemical water quality variables, phytoplankton and indicators of faecal contamination, a significant improvement along the longitudinal gradient from the discharge point was observed after OP decommissioning. This improvement was fast, being detected two months after decommissioning, positively affecting areas used as shellfish farming grounds. However, distribution patterns of bacteriological indicators and regular shellfish harvesting interdictions suggested an alternative source of faecal contamination after OP decommissioning. At FO, both chemical variables and bacteriological indicators of faecal contamination revealed a slower improvement, only six-months after the UWWTP alteration. Before that, increased and highly variable ammonium, chlorophyll a concentration, phytoplankton abundances and Escherichia coli densities, revealed an unstable phase. Overall, a lower water quality at FO in respect to OP reflected not only a higher effluent volume but also more restricted water circulation for the former.

Phytoplankton seasonal and interannual variability in the Guadiana upper estuary was analyzed during 1996-2005, a period that encompassed a climatic controlled reduction in river flow that was superimposed on the construction of a dam. Phytoplankton seasonal patterns revealed an alternation between a persistent light limitation and episodic nutrient limitation. Phytoplankton succession, with early spring diatom blooms and summer-early fall cyanobacterial blooms, was apparently driven by changes in nutrients, water temperature, and turbulence, clearly demonstrating the role of river flow and climate variability. Light intensity in the mixed layer was a prevalent driver of phytoplankton interannual variability, and the increased turbidity caused by the Alqueva dam construction was linked to pronounced decreases in chlorophyll a concentration, particularly at the start and end of the phytoplankton growing period. Decreases in annual maximum and average abundances of diatoms, green algae, and cyanobacteria were also detected. Furthermore, chlorophyll a decreases after dam filling and a decrease in turbidity may point to a shift from light limitation towards a more nutrient-limited mode in the near future.