M.A. Almeida

Factors Influencing Bacterial Production in a Shallow Estuarine System.

The bacterioplankton of the marine and brackish water zones of the complex system Ria de Aveiro was characterized as profiles of bacterial abundance and biomass productivity. During the warm season, total bacteria ranged from 0.2 to 8.5 × 109 cells L-1 and active bacteria number from 0.1 to 3.1 × 109 cells L-1. Total and active bacterial numbers were, on average, three times higher in brackish than in marine water. Bacterial productivity on different dates and different tides in the marine zone varied from 0.05 to 4.5 mg C L-1 h-1. Here the average productivity (1.1 mg C L-1 h-1) was 3.5 times less than in brackish water (average 3.8 mg C L-1 h-1; range 0.7-14.2 mg C L-1 h-1). Specific productivity varied from 0.05 to 2.61 fg C cell-1 h-1, a range that was similar throughout the ecosystem. However, specific productivity per active cell was 19% higher in brackish water. Bacterial production variation was best explained by the number of active bacteria, which, in turn, was highly associated with total bacterial number, temperature, and particulate organic carbon. In the marine zone, bacterial production was also influenced by depth and salinity. In the brackish zone, the set of independent variables explained a smaller percentage of bacterial production variation than in marine zone, suggesting greater importance of other variables. In the marine zone, and mainly near low tide, productivity was significantly higher (average 3.3 times) at the surface (down to 0.5 m) than in the deeper layers of the water column. This stratification of bacterial productivity was linked to the increased specific productivity per active cell, as no modification in the proportion of active cells in the population could be detected. The vertical profile of bacterial production in the deeper zone of this estuarine ecosystem, in which no clear salinity or thermal stratification occurs throughout the tidal cycle, seemed to reflect a biochemical stratification generated by increased phytoplankton exudation and/or by photochemical transformation of semilabile or recalcitrant organic compounds. Shallower water masses tend to blur this surface effect. The relative importance of photochemical transformation in the pattern of estuarine bacterial production will therefore tend to vary with the bathymetry of the system.

Loss of Estuarine Bacteria by Viral Infection and Predation in Microcosm Conditions

The bacterioplankton density in Ria de Aveiro, a shallow estuarine ecosystem, varied in the broad range of 1.9-10.6 × 109 cells L-1. The range of values was about 2 times higher in brackish water than in marine water. At high tide bacterial abundance was 2-3 times lower than at low tide. The overall variation in virioplankton was in the range of 2.4-25.0 × 1010 particles L-1. Brackish water was about 2 times richer in viral particles than the marine water. Near low tide the virioplankton was 2-3 times higher that at high tide. Viral density followed the pattern of bacterial abundance (it explained 40% of virioplankton variation). The viruses to bacterium ratio varied, throughout tidal cycles, by a factor of about 10 establishing the range 4.7-55.6 (average 17.6). This ratio was rather similar in the two estuarine zones. We compared the effects of infection and predation on the control of bacterioplankton size in the two zones of the estuary. The approach to this question was conducted in experimental microcosms, set up in six combinations of plankton variables affecting the presence/absence of predators, virus-to-bacterium ratio (10-fold increase), virus-to-bacterium distance (2.2-fold increase), and bacterial growth rate. The results showed that predation was similar, in a percent basis, in marine (69%) and brackish water (73%). Viral infection was, however, higher in brackish water (59%) than in the marine water (36%). We conclude that the bacterioplankton along the salinity gradient evolves under biological pressures that are in different balance in the marine and brackish water zones. The effect of viral lysis on bacterial communities with enhanced growth (after yeast extract addition) was masked even when the initial ratio was 10-fold greater than in the natural samples. The high density of the virioplankton did not preclude the large and rapid increase in bacterial density. We suggest that the dynamics of the equilibrium between bacteria and viruses in the environment is driven to higher numerical levels during periods of intensive bacterial growth. On the contrary, at low bacterial growth rates the temporarily increased virus-to-bacterium ratio may drive the equilibrium to its lowest levels.

Short-term responses of the natural planktonic bacterial community to the changing water properties in an estuarine environment : Ectoenzymatic activity, glucose incorporation, and biomass production

The possibility that two principlal bacterial communities expressing different levels of heterotrophic activity might coexist in an estuarine ecosystem (Ria de Aveiro, Portugal) and could quickly respond to tidal fluctuations of environmental factors was experimentally tested in diffusion chambers by swapping the dissolved components of the natural water between the two communities and comparing their reactivity against the unaltered controls. The results for ectoenzymatic activity (Leuaminopeptidase and β-glucosidase), glucose incorporation and biomass production after transference of the marine bacterial community to brackish water showed maxima in the range of 241–384% of the control values. The opposite transference of the brackish-water bacterial community to marine water produced maximal decreases to 0.14–0.58% of the control values. In a reverse experiment, designed as the return to the initial conditions after 2 hours of the first exposure, the marine community rapidly re-acquired the characteristic low profile of activity. Contrastingly, the negative effects of 2 hours of exposure to marine water on the activity of the brackish water bacteria persisted, at least for 4 hours, after return to their own water. The apparent short-term irreversibility of the decline in activity of the brackish water bacteria when exposed to marine water, in parallel with the quick and reversible positive response of the marine water bacteria to the brackish water, suggests the development of two distinct bacterioplankton communities adapted to the environmental conditions prevailing at distinct sections of the estuary. The reactivity to environmental changes demonstrated by the two communities allows the prediction of estuarine profiles of bacterial activity steeper than those expected from the conservative transport of bacterial cells associated with tidal currents.

Compartments of oxygen consumption in a tidal mesotrophic estuary (Ria de Aveiro, Portugal)

Oxygen consumption rates were determined, in parallel with primary production and bacterial biomass production, as an approach to the analysis of carbon cycling in the estuarine community of the Ria de Aveiro. The water column of the marine zone was the major contributor (64-99 %) to the total aerobic carbon remineralisation in which O 2 uptake rates averaged from 80 to 127 mg O 2 .m -2 .h -1 , respectively at low tide and high tide. The planktonic consumption of O 2 varied from 0.010 to 0.041 mg O 2 .L -1 .h -1 with the highest values in the brackish zone. Small water column depths in this zone, however, reduced the integrated average consumption of the plankton, per unit of surface area, to 57 (LT) and 66 % (HT) of that observed in the marine zone. Benthic O 2 consumption rates, 5.1 to 22.0 mg O 2 .m -2 .h -1 , were two to four times higher in the brackish zone when compared to the rates in the marine zone. It represented 1-31 % of the total surface integrated values in different areas and at different tides. From the ratios of the primary production and bacterial biomass production, on a per surface unit basis. it is concluded that, in late autumn, the Ria de Aveiro was mostly a heterotrophic system with a feeble recovery of primary production at HT in the marine zone and at LT in the brackish water zone.

Fluxes of bacterioplankton between a tidal estuary and the sea: returning to the "Outwelling Hypothesis"

The tidal dynamics of bacterioplankton communities at the outer part ofa shallow estuary (Ria de Aveiro, Portugal) were studied during 6 tidal cyclesat a fixed sampling site. Bacterial numbers (0.2–8.1 ×109 cells l−1), aminopeptidase activity(189–1662 nmol l−1 h−1),β-glucosidase activity (1.7–67.0 nmoll−1 h−1) and potential glucoseincorporation (0.48–3.99 nmoll−1 h−1) followed a consistent patternof increase during ebb and decrease during flood.Fluxes of bacterioplankton populations and associated heterotrophic activitiesbetween the estuary and the coastal area during a tidal cycle were estimatedfrom the water flux as derived from a two-dimensional vertically-integratednumerical model. The net fluxes estimated for a tidal cycle ranged from−26.0 to −2.5 to × 1016 bacterial cells. The nettidal fluxes of potential heterotrophic activities ranged from −10 to−80 mol h−1 for aminopeptidase, −0.33to −1.10 mol h−1 for β-glucosidase and−0.18 to +0.03 mol h−1 for glucoseincorporation. Net fluxes were generally negative in sign indicating thetransfer of phyto- and bacterioplankton, as well as potential capacities for thedegradation and recycling of organic matter, from the outer estuarinecompartment to the sea.

Is bacterioplankton production in the Ria de Aveiro influenced by salt marshes and bed sediment

The contribution of potential export of materials from bottom sedimentsand salt marshes into the water column of a shallow estuarine system of Ria deAveiro to the observed high bacterial productivity in the mid section of thisestuary was evaluated. Vertical profiles of physical, chemical and bacterialvariables were studied in the marine and brackish water zones, and oftransversal profiles in the brackish zone only. Although the concentrations ofseston (17–241 mg l−1), particulate organiccarbon (3–15.5 mg l−1) and chlorophyll(1.2–7.0 μg l−1) varied widely, thevertical and transversal profiles were without much variation. Total bacterialnumber (0.2–8.5 × 109 cellsl−1) and the number of particle-attached bacteria(0.02–2.50 × 109 cellsl−1)along vertical and transversal profiles did not differ much, but the rate ofbacterial production (0.05–14.2 μg C l−1h−1) and dissolved organic carbon concentration(6.0–69.2 mg l−1) were frequently highernear the salt marsh margin at the brackish water transect. The increase inproductivity could not be associated with runoff of particulate matter butcoincided with the inputs of dissolved organic carbon. The results of verticaland transversal profiles point to a minor role of particulate matter additionsfrom the salt marsh area or from bed sediments.

Ectoenzymatic activity and glucose heterotrophic metabolism in a shallow estuary (Ria de Aveiro, Portugal): influence of bed sediments and salt marshes

Abstract Bacterioplankton abundance and activity were studied in the estuarine system of Ria de Aveiro (Portugal) to test if tidal resuspension of sediments and transport of particles from the salt marshes may act as factors of variability of bacterial communities. The total and attached cell abundance, ectoenzymatic activity and the heterotrophic metabolism of glucose, as well as seston, chlorophyll a and particulate organic carbon (POC) were monitored during four 10-h periods along the tidal cycle at four sampling sites across a transect. The variation of particulate materials (seston, POC and chlorophyll a ) along the transect was not significantly correlated with either distance to the margin or distance to the sediment surface. Nevertheless, proximity to the salt marsh or to the bottom sediment surface favoured glucose incorporation and aminopeptidase activity. A multiple stepwise linear regression analysis using temperature, salinity, seston, POC, chlorophyll a , distance to sediment surface and distance to the margin as independent variables explained 66.5% of the variability of the fraction of particle-attached bacteria and only a very small proportion (12–43%) of the observed variability of total bacterial abundance, ectoenzymatic activity and glucose utilization. The spatial patterns of variation of the concentration of particulate material (seston, POC and chlorophyll a) do not clearly indicate the occurrence of sediment resuspension and runoff from the margins. This, together with the poor contribution of these parameters to the transversal and tidal variability of bacterial activity, dismisses the importance of inputs of suspended material across the sediment/water interface and from neighbouring salt marshes in the control of bacterial density and activity.

Virological quality of the Ria de Aveiro: Validity of potential microbial indicators

The summer occurrence of human enteroviruses and rotaviruses in the bacteriologically clean area of the Ria de Aveiro, a coastal marine lagoon, prompted the question of the assessment of the virological quality of recreational waters and shellfish raising beds. Enteroviruses were present in surface water at a density of 3 pfu 10 l−1 and were accumulated in sediments and, especially, in cockles where they reached concentrations 2 to 310log units greater. Rotaviruses were detected at one10log unit below the density of enteroviruses in sediments and cockles and were not detected in water. Four bacteriophage systems were assayed as indicators of human enteric viruses: somatic coliphages ofE.coli C, sexual and sexual-RNA coliphages plated onSalmonella WG49 and phages againstBacteroides fragilis HSP40. The results obtained from 2 lagoon stations sampled in summer, autumn and winter showed that the four systems failed to indicate the presence of enteroviruses and rotaviruses in water, sediment and shellfish samples. The absence of phages ofB. fragilis HSP40 in all types of samples taken from the lagoon, but not from the residual waters of the treatment station, suggests that they may suffer a strong negative pressure in this ecosystem as their proportion to the coliphages in the cockles deviated strongly from the ratio of 1∶100 to 1∶1000 observed at the sewage outfall. In fact, no correlation was observed between these phages and enteric viruses or coliphages. Alternatively, it is possible that the importance of diffuse faecal pollution and the interference of faecal pollutants of animal origin, including migratory sea birds which are abundant in winter, can alter the proportions of the faecal bacteriophages beyond recognition. It is apparent that bacteriophage monitoring of the health risk linked to the occurrence of viruses in the marine environment is not yet fully resolved, what may leave viral quality assessment dependent on direct detection of human enteric virus.