Linking biofilm spatial structure to real-time microscopic oxygen decay imaging

Two non-destructive techniques, confocal laser scanning microscopy (CLSM) and planar optode (VisiSens imaging), were combined to relate the fine-scale spatial structure of biofilm components to real-time images of oxygen decay in aquatic biofilms. Both techniques were applied to biofilms grown for seven days at contrasting light and temperature (10/20°C) conditions. The geo-statistical analyses of CLSM images indicated that biofilm structures consisted of small (~100 µm) and middle sized (~101 µm) irregular aggregates. Cyanobacteria and EPS (extracellular polymeric substances) showed larger aggregate sizes in dark grown biofilms while, for algae, aggregates were larger in light-20°C conditions. Light-20°C biofilms were most dense while 10°C biofilms showed a sparser structure and lower respiration rates. There was a positive relationship between the number of pixels occupied and the oxygen decay rate. The combination of optodes and CLMS, taking advantage of geo-statistics, is a promising way to relate biofilm architecture and metabolism at the micrometric scale

Delayed response of microbial epipelic biofilm to nutrient addition in a Pampean stream

In streams and rivers, the first organisms which directly receive and respond to nutrients are primary producers (algae, macrophytes) and microbial heterotrophs (bacteria, fungi) since they rely on available inorganic nutrients from the water column. The aim of the present study was to analyze the response of the epipelic microbial biofilm in a Pampean stream submitted to a continuous input of inorganic nutrients (nitrogen and phosphorus). For this purpose, we measured the effects of moderate nutrient addition during 14 mo on the epipelic biofilm community of a meso-eutrophic stream that runs through the Pampean plain. The effects of nutrient enrichment were tested by analyzing the difference in algal and bacterial biomass and 2 extracellular enzymatic activities (β;-glucosidase, phosphatase) at an enriched reach compared with those measured at an unmodified upstream reach. Overall, the response of the epipelon of this Pampean stream produced a slow and delayed effect on algal biomass increases, which might result in a delayed effect on the increase of bacterial densities. Neither phosphatase activity nor β;-glucosidase activity exhibited significant changes due to nutrient addition. This may be due to the fact that the phosphatase activities measured were basal activities, uninhibited by enrichment, and that the epipelic β;-glucosidase activity was regulated more by substrate availability than by any nutrient imbalance. Although changes are slow, if some of these changes were to become chronic, they would affect the functioning and services of the whole stream ecosystem.Instituto de Limnología "Dr. Raul A. Ringuelet

Bilayer infiltration system combines benefits from both coarse and fine sands promoting nutrient accumulation in sediments and increasing removal rates

Infiltration systems are treatment technologies based on water percolation through porous media where biogeochemical processes take place. Grain size distribution (GSD) acts as a driver of these processes and their rates and influences nutrient accumulation in sediments. Coarse sands inhibit anaerobic reactions such as denitrification and could constrain nutrient accumulation in sediments due to smaller specific surface area. Alternatively, fine sands provide higher nutrient accumulation but need a larger area available to treat the same volume of water; furthermore, they are more susceptible to bioclogging. Combining both sand sizes in a bilayer system would allow infiltrating a greater volume of water and the occurrence of aerobic/anaerobic processes. We studied the performance of a bilayer coarse-fine system compared to a monolayer fine one - by triplicate - in an outdoor infiltration experiment to close the C-N-P cycles simultaneously in terms of mass balances. Our results confirm that the bilayer coarse-fine GSD promotes nutrient removal by physical adsorption and biological assimilation in sediments, and further it enhances biogeochemical process rates (2-fold higher than the monolayer system). Overall, the bilayer coarse-fine system allows treating a larger volume of water per surface unit achieving similar removal efficiencies as the fine system. This document is the unedited Author’s version of a Submitted Work that was subsequently accepted for publication in Environmental science and technology

Delayed response of microbial epipelic biofilm to nutrient addition in a Pampean stream

In streams and rivers, the first organisms which directly receive and respond to nutrients are primary producers (algae, macrophytes) and microbial heterotrophs (bacteria, fungi) since they rely on available inorganic nutrients from the water column. The aim of the present study was to analyze the response of the epipelic microbial biofilm in a Pampean stream submitted to a continuous input of inorganic nutrients (nitrogen and phosphorus). For this purpose, we measured the effects of moderate nutrient addition during 14 mo on the epipelic biofilm community of a meso-eutrophic stream that runs through the Pampean plain. The effects of nutrient enrichment were tested by analyzing the difference in algal and bacterial biomass and 2 extracellular enzymatic activities (β;-glucosidase, phosphatase) at an enriched reach compared with those measured at an unmodified upstream reach. Overall, the response of the epipelon of this Pampean stream produced a slow and delayed effect on algal biomass increases, which might result in a delayed effect on the increase of bacterial densities. Neither phosphatase activity nor β;-glucosidase activity exhibited significant changes due to nutrient addition. This may be due to the fact that the phosphatase activities measured were basal activities, uninhibited by enrichment, and that the epipelic β;-glucosidase activity was regulated more by substrate availability than by any nutrient imbalance. Although changes are slow, if some of these changes were to become chronic, they would affect the functioning and services of the whole stream ecosystem.Instituto de Limnología "Dr. Raul A. Ringuelet

Hydrology, light and the use of organic and inorganic materials as structuring factors of biological communities in Mediterranean streams

Hydrological disturbances, light availability and nutrients are the most relevant factors determining the structure of the biological communities in Mediterranean rivers. While some hydrological disturbances are able to induce catastrophic effects, which may cause a complete reset in physical and biological conditions, continued enrichment or changes in light availability are factors leading to the progressive shift in the communities of autotrophs and heterotrophs in the systems. Primary production in Mediterranean streams shows relevant seasonal changes which mainly follows the variations in light availability. In most forested streams, the algal community is shade-adapted. Nutrient enrichment (especially phosphorus) leads to marked increases in primary production, but this increase is not lineal and there is a saturation of algal biomass even in the most enriched systems. The heterotrophs (bacteria, fungi) are related to the pattern of DOC availability (which most depends on the seasonal discharge and leaf fall dynamics) and to the available substrata in the stream. It has been repeatedly observed that shorttime increases of extracellular enzyme activities are related to the accumulation of autochthonous (algal) and/or allochthonous (leaves) organic matter on the streambed during spring and summer, this being more remarkable in dry than in wetter years. Flow reduction favours detritus concentration in pools, and the subsequent increase in the density and biomass of the macroinvertebrate community. In Mediterranean streams collectors are accounting for the highest density and biomass, this being more remarkable in the least permanent systems, in accordance with the effect of floods on the organic matter availability. Nutrients, through the effect on the primary producers, also affect the trophic food web in the streams by favouring the predominance of grazers.Los cambios hidrológicos, la disponibilidad de la luz y los nutrientes son los factores más importantes que determinan la estructura y función de la comunidad en los ríos Mediterráneos. Mientras los cambios hidrológicos pueden causar un total reestablecimiento en las condiciones físicas y biológicas, un aumento continuo de la concentración de nutrientes o cambios en la disponibilidad de la luz, son factores que favorecen un cambio progresivo de las comunidades de autótrofos y heterótrofos. La producción primaria en los ríos Mediterráneos presenta cambios estacionales en función de la luz. En los ríos forestados, las algas están adaptadas a la baja intensidad de la luz. El aumento de la concentración de nutrientes, especialmente el fósforo, favorece la producción primaria pero no de forma lineal, ya que aparece una saturación de la biomasa algal en los sistemas más eutróficos. Los heterótrofos (bacterias y hongos) están más relacionados con la disponibilidad de DOC (que a su vez depende de las variaciones estacionales del caudal y de la dinámica de la caída de las hojas) y de sustrato. Se ha observado, repetidamente, una relación entre el aumento a corto plazo de la actividad exoenzimática y la acumulación de biomasa autóctona algal y/o alóctona (hojas) en el lecho del río, durante la primavera y el verano. Esta relación es más evidente en los años más secos. La reducción del caudal favorece la concentración del detritus en las áreas de menor velocidad de corriente ("pools") y se observa un aumento en la densidad y biomasa de los macroinvertebrados. En los ríos Mediterráneos los recolectores alcanzan las mayores densidades y biomasas, especialmente en los sistemas menos permanentes, en relación con el efecto de las variaciones de caudal sobre la disponibilidad de materia orgánica. Los nutrientes, a través de la producción primaria, tienen un efecto también sobre la estructura de la red trófica, favoreciendo a los ramoneadores

Interaction between physical heterogeneity and microbial processes in subsurface sediments: a laboratory-scale column experiment

Physical heterogeneity determines interstitial fluxes in porous media. Nutrients and organic matter distribution in depth influence physicochemical and microbial processes occurring in subsurface. Columns 50 cm long were filled with sterile silica sand following five different setups combining fine and coarse sands or a mixture of both mimicking potential water treatment barriers. Water was supplied continuously to all columns during 33 days. Hydraulic conductivity, nutrients and organic matter, biofilm biomass, and activity were analyzed in order to study the effect of spatial grain size heterogeneity on physicochemical and microbial processes and their mutual interaction. Coarse sediments showed higher biomass and activity in deeper areas compared to the others; however, they resulted in incomplete denitrification, large proportion of dead bacteria in depth, and low functional diversity. Treatments with fine sediment in the upper 20 cm of the columns showed high phosphorus retention. However, low hydraulic conductivity values reported in these sediments seemed to constraint biofilm activity and biomass. On the other hand, sudden transition from coarse-to-fine grain sizes promoted a hot-spot of organic matter degradation and biomass growth at the interface. Our results reinforce the idea that grain-size disposition in subsurface sandy sediments drives the interstitial fluxes, influencing microbial processes. This document is the unedited Author’s version of a Submitted Work that was subsequently accepted for publication in Environmental science and technolog

A mechanistic model (BCC-PSSICO) to predict changes in the hydraulic properties for bio-amended variably saturated soils

The accumulation of biofilms in porous media is likely to influence the overall hydraulic properties and, consequently, a sound understanding of the process is required for the proper design and management of many technological applications. In order to bring some light into this phenomenon we present a mechanistic model to study the variably saturated hydraulic properties of bio-amended soils. Special emphasis is laid on the distribution of phases at pore-scale and the mechanisms to retain and let water flow through, providing valuable insights into phenomena behind bioclogging. Our approach consists in modeling the porous media as an ensemble of capillary tubes, obtained from the biofilm-free water retention curve. This methodology is extended by the incorporation of a biofilm composed of bacterial cells and extracellular polymeric substances (EPS). Moreover, such a microbial consortium displays a channeled geometry that shrinks/swells with suction. Analytical equations for the volumetric water content and the relative permeability can then be derived by assuming that biomass reshapes the pore space following specific geometrical patterns. The model is discussed by using data from laboratory studies and other approaches already existing in the literature. It can reproduce (i) displacements of the retention curve toward higher saturations and (ii) permeability reductions of distinct orders of magnitude. Our findings also illustrate how even very small amounts of biofilm may lead to significant changes in the hydraulic properties. We, therefore, state the importance of accounting for the hydraulic characteristics of biofilms and for a complex/more realistic geometry of colonies at the pore-scale

Effects of riparian vegetation removal on nutrient retention in a Mediterranean stream

We examined the effects of riparian vegetation removal on algal dynamics and stream nutrient retention efficiency by comparing NH4-N and PO4-P uptake lengths from a logged and an unlogged reach in Riera Major, a forested Mediterranean stream in northeastern Spain. From June to September 1995, we executed 6 short-term additions of N (as NH4Cl) and P (as Na2HPO4) in a 200-m section to measure nutrient uptake lengths. The study site included 2 clearly differentiated reaches in terms of canopy cover by riparian trees: the first 100 m were completely logged (i.e., the logged reach) and the remaining 100 m were left intact (i.e., the shaded reach). Trees were removed from the banks of the logged reach in the winter previous to our sampling. In the shaded reach, riparian vegetation was dominated by alders (Alnus glutinosa). The study was conducted during summer and fall months when differences in light availability between the 2 reaches were greatest because of forest canopy conditions. Algal biomass and % of stream surface covered by algae were higher in the logged than in the shaded reach, indicating that logging had a stimulatory effect on algae in the stream. Overall, nutrient retention efficiency was higher (i.e., shorter uptake lengths) in the logged than in the shaded reach, especially for PO4-P. Despite a greater increase in PO4-P retention efficiency relative to that of NH4-N following logging, retention efficiency for NH4-N was higher than for PO4-P in both study reaches. The PO4-P mass-transfer coefficient was correlated with primary production in both study reaches, indicating that algal activity plays an important role in controlling PO4-P dynamics in this stream. In contrast, the NH4-N mass-transfer coefficient showed a positive relation-ship only with % of algal coverage in the logged reach, and was not correlated with any algal-related parameter in the shaded reach. The lack of correlation with algal production suggests that mechanisms other than algal activity (i.e., microbial heterotrophic processes or abiotic mechanisms) may also influence NH4-N retention in this stream. Overall, this study shows that logging disturbances in small shaded streams may alter in-stream ecological features that lead to changes in stream nutrient retention efficiency. Moreover, it emphasizes that alteration of the tight linkage between the stream channel and the adjacent riparian zone may directly and indirectly impact biogeochemical processes with implications for stream ecosystem functioning

Quality and reactivity of dissolved organic matter in a Mediterranean river across hydrological and spatial gradients.

Understanding DOM transport and reactivity in rivers is essential to having a complete picture of the global carbon cycle. In this study, we explore the effects of hydrological variability and downstream transport on dissolved organic matter (DOM) dynamics in a Mediterranean river. We sampled the main stem of the river Tordera from the source to the sea, over a range of fifteen hydrological conditions including extreme events (flood and drought). By exploring spatial and temporal gradients of DOM fluorescence properties, river hydrology was found to be a significant predictor of DOM spatial heterogeneity. An additional space-resolved mass balance analysis performed on four contrasting hydrological conditions revealed that this was due to a shift in the biogeochemical function of the river. Flood conditions caused a conservative transport of DOM, generating a homogeneous, humic-like spatial profile of DOM quality. Lower flows induced a non-conservative, reactive transport of DOM, which enhanced the spatial heterogeneity of DOM properties. Moreover, the downstream evolution of DOM chemostatic behaviour revealed that the role of hydrology in regulating DOM properties increased gradually downstream, indicating an organised inter-dependency between the spatial and the temporal dimensions. Overall, our findings reveal that riverine DOM dynamics is in constant change owing to varying hydrological conditions, and emphasize that in order to fully understand the role of rivers in the global carbon cycle, it is necessary to take into account the full range of hydrological variability, from floods to droughts