Yearly variation of bacterial production in the Arraial do Cabo protection area (Cabo Frio upwelling region): an evidence of anthropogenic pressure

RESEARCH PAPER

Yearly variation of bacterial production in the Arraial do Cabo protection area (Cabo Frio upwelling region): An evidence of anthropogenic pressure

Sérgio A. Coelho-Souza^{I, II, III}; Gilberto C. Pereira^IV; Ricardo Coutinho^I; Jean R.D. Guimarães^III

^IDepartamento de Biotecnologia Marinha, Instituto de Ciências do Mar Almirante Paulo Moreira, Arraial do Cabo, RJ, Brazil

^IIUniversidade de Santo Amaro, São Paulo, SP, Brazil

^IIILaboratório de Traçadores, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ, Brazil

^IVInstituto Alberto Luiz Coimbra de Pós-Graduação e Pesquisa de Engenharia, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ, Brazil

^{Correspondence}

ABSTRACT

Arraial do Cabo is where upwelling occurs more intensively on the Brazilian coast. Although it is a protection area it suffers anthropogenic pressure such as harbor activities and sporadic sewage emissions. Short-time studies showed a high variability of bacterial production (BP) in this region but none of them evaluated BP during long periods in a large spatial scale including stations under different natural (upwelling and cold fronts) and anthropogenic pressures. During 2006, we sampled surface waters 10 times (5 in upwelling and 5 in subsidence periods) in 8 stations and we measured BP, temperature as well as the concentrations of inorganic nutrients, pigments and particulate organic matter (POM). BP was up to 400 times higher when sewage emissions were observed visually and it had a positive correlation with ammonia concentrations. Therefore, in 2007, we did two samples (each during upwelling and subsidence periods) during sewage emissions in five stations under different anthropogenic pressure and we also measured particles abundance by flow cytometry. The 12 samples in the most impacted area confirmed that BP was highest when ammonia was higher than 2 μM, also reporting the highest concentrations of chlorophyll a and suspended particles. However, considering all measured variables, upwelling was the main disturbing factor but the pressure of fronts should not be neglected since it had consequences in the auto-heterotrophic coupling, increasing the concentrations of non fluorescent particles and POM. Stations clustered in function of natural and anthropogenic pressures degrees and both determined the temporal-spatial variability.

Key words: bacterioplankton, heterotrophic activity, impacted area; extractive reserve, upwelling system.

Introduction

The up-flow of cold and nutrient-rich waters describes upwelling process and it disturbs ecosystem dynamics and increases environmental heterogeneity (Lehmann and Myrberg, 2008). Brazilian upwelling events are more pronounced in the Cabo Frio region (RJ); mainly on the Arraial do Cabo coast (23° S, 42° W), an environmental protection area (Silva, 2004) that is directly impacted by anthropogenic activities (Ferrerira et al., 2004; Lopez and Coutinho, 2010; Cury et al., 2011). In the Cabo Frio region, upwelling occurs with higher frequency and intensity during austral spring and summer seasons, mainly as a function of northeast (NE) winds prevalence. During the autumn/winter season, cold fronts are more common describing a subsidence period (Valentin, 2001; Coelho-Souza et al., 2012). Upwelling events occur intensely off the Cabo Frio embayment, precisely at Focinho do Cabo (Figure 1), a fixed station for many oceanographic studies (eg., Valentin et al., 1987; Guenther et al., 2008).

Sampling in the Focinho do Cabo station, Valentin (1984) showed that upwelling events are the main disturbing factor of the region but did not measure bacterial activity. A few studies focused on bacterioplankton heterotrophic activity and how it was related to upwelling process (Carvalho and Gonzalez-Rodriguez, 2004; Guenther et al., 2008). Measurements of bacterial production (BP) are important to evaluate the cycles of carbon and nutrients in the environment as reported in the experimental and sampling studies in the Focinho do Cabo station associating BP with primary production. These studies showed that carbon, nitrogen and phosphorous cycles were coupled with BP. Herein, we extend the analysis of bacterioplankton heterotrophic activity in space and time to include the interference of cold fronts and anthropogenic activities. Our hypotheses is that natural (upwelling and cold fronts) and anthropogenic (sewage emissions) pressures have different interference scales. We expected a regional interference of upwelling and fronts but punctual anthropogenic impact in function of water circulation.

Materials and Methods

Study area

This study was performed during 2006/7 in 8 sampling stations with different characteristics (Figure 1). Ponta da Cabeça (PC), for instance, is outside the bay and directly influenced by upwelling events and strong waves during cold fronts. The other seven are distributed inside the Cabo Frio embayment as follows: Praia da Ilha (PI) is frequently influenced by upwelling and cold fronts; Pedra Vermelha (PV) and Ilha dos Porcos (IP) are influenced by north waters from Arraial do Cabo; In the Forno inlet (number 2 in Figure 1), upwelling influence is rare and three stations were established: one with oyster and coquille cultivation (Forno Cultivo - FNC); one more exposed to wave action (Fortaleza - FO), and another one near the beach and a floating restaurant (Forno Flutuante - FNF); The eighth site is the Praia dos Anjos (PA) station in the Anjos inlet (number 1 in Figure 1) that also has rare upwelling influence but is directly influenced by harbor activities, as well as by sporadic sewage emissions. These emissions are usually associated with intense rainfalls or with population increase during summer vacations.

Sampling procedure and water variables measurements

All stations were sampled 12 times, 6 during the spring/summer (upwelling period) and 6 during the autumn/winter (subsidence period) seasons, always between 9:00 and 12:00 at morning. At each station, 5 liters of surface water were harvested with a bucket and kept in thermo plastic bottles until laboratory analysis (~3 h). Nutrients (nitrate, nitrite, ammonia and phosphate) were determined according to Strickland and Parsons (1972). The determination of pigment concentrations (chlorophyll a, b, c and pheophytin) followed Richard and Thompson (1952) after filtration through cellulose acetate membranes (Millipore^®, 0.45 mm pore size) and SCOR-UNESCO (1966) equations were used for calculations. Suspended Particulate Matter (SPM) was determined using Whatman^® glass fiber filters with ~1.2 mm mesh and Particulate Organic Matter (POM) was determined through SPM loss upon ignition at 400 °C during 24 h.

Bacterial production

Heterotrophic bacterioplankton activity was measured using the ³H-Leucine incorporation technique (Kirchman et al., 1985). The use of one or two controls was tested using 100% thricloroacetic acid (5% final concentration) in 1.7 mL of sampled water, but one control was found adequate. After 1 h incubations and within a 15 days period after that (Miranda et al., 2007), proteins were extracted using the micro centrifugation method described by Smith and Azam (1992). Protein radioactivity was measured on a Perkin-Elmer TRICARB 1600 liquid scintillation detector during 30 min or after the accumulation of 10000 counts (Coelho-Souza et al., 2013). DPM values were obtained after quenching correction using the external standard of the equipment.

³H-Leucine incorporation rates were calculated using DPM, the specific activity of ³H-Leucine and a conversion rate from DPM to 1 mol of leucine (Kirchman, 1993). Bacterial activity standard error was determined using different leucine concentrations and sub-sampling (Cumming et al., 2007). 5 mL of different substrate concentrations (0.1 to 32 nM) and ³H-Leucine specific activities were used to evaluate the average error associated with sub-sampling (maximum of 7).

Leucine incorporation saturated at 10 nM, and the average errors were usually < 17%, even when adding less than 1 nM leucine. The addition of saturated leucine concentrations could inhibit amino acid biosynthesis but this was not confirmed by Kirchman (2001) and it was observed that 0.5-1.0 nM of leucine was not incorporated by cyanobacteria (Kirchman et al., 1985) while when using 20 nM it was incorporated by picoeukaryotes (Michelou et al., 2007). Hence, for this study we used 0.5 nM leucine and used Simon and Azam (1989) and Simon (1991) conversion factors for isotopic dilution to measure carbon production. Meanwhile, with this empiric correction there was no difference between the measurements using 0.5 or 20 nM of leucine, as used by other authors that studied the region (Carvalho and Gonzalez-Rodriguez, 2004; Guenther et al., 2008).

Flow cytometry

To measure the density of particles in water, 1.5 mL samples were injected in a CytoBuoy Flow Cytometer (Woerden - The Netherlands) equipped with a solid blue laser providing 20 mV at 488 nm, one side scatter (SWS, 446/500nm) detector and three others for red (chlorophyll-a) fluorescence (FL1- 669/725 nm); orange/yellow (FL-2, 601/651 nm), and green/yellow (FL-3, 515/585 nm) fluorescence (Dubelaar et al., 1999). Data acquisition was done through the Cytosiftlab software and the use of the CytoQuick software allowed us to discriminate between fluorescent and non fluorescent particles and estimate their relative size distributions. These analyses were only done two times during sewage emissions observed visually in 2007 and were not included in the statistical analysis.

Statistical analysis

Descriptive statistics were performed using the median value of all measured variables and sampling stations. The similarities between all sampling stations were assessed by cluster analysis (a dendrogram using the Ward method and Euclidean distances). In order to associate the physical-chemical-biological variables, a principal component analysis (PCA) was used (James and McCulloch, 1990). For PCA, we also used wind data from a station situated in Arraial do Cabo (Figure 1), acquired at the web page of the Instituto Brasileiro de Metereologia (www.inmet.gov.br). The used wind variables were the mean force (speed) and direction (NE x SW) during the last 24 h before sampling. Then, we considered BP, winds, temperature, the concentrations of phosphate, nitrate, ammonia, chlorophyll a and POM as variables for analysis and the others (concentrations of nitrite, chlorophylls b and c and pheophytin) as supplementary variables. The number of accepted factors was determined by Eigen values ≥ 1 which justifies more than 10% of variability.

Results

Heterotrophic bacterioplankton production

BP was variable between samplings periods and stations, and its average ranged from 0.6 (PC) to 4.4 (PA). Considering all samples, BP had an average of 1.3 μgC.L^-1.h^-1 and a standard deviation of 2.4 (n = 96). Values above 99% of confidence intervals were observed in PA and FO stations (Figure 2; Table 1). When sewage discharges were visually observed in PA station, BP increased up to almost 400 times and the highest values were associated with ammonia concentrations above 2 μM (Figure 3).

Environmental variability

The dendrogram using physical-chemical-biological water variables (median values) confirms the great pressure of hydrobiological variability in the Cabo Frio upwelling region (Figure 4). The PI station had the highest concentrations of chlorophylls and POM (Table 1), indicating an intense autotrophic production in this area. POM was also high in IP and in the main impacted stations, suggesting an input of allochthonous material from coastal waters and anthropogenic activities, respectively. PC station was different from the other sites since it is under influence of both upwelling and cold fronts, as shown by its lowest minimum temperature and maximum nutrient concentrations (Table 1). Inner the Cabo Frio embayment, the minimum temperatures indicated a low pressure of upwelling, except at PI station. As observed visually, this area was also under high pressure of cold fronts.

Considering the four main components, PCA analysis justified more than 80% of the data variability (data not shown). Factor 1, for instance, accounts for 32.3% and seems to be determined mainly by the contrast between temperature and concentrations of phosphate and nitrate, indicating that upwelling events are the main factor disturbing the system. Factor 2 explained 26.4% of the variability and was associated with wind direction and intensity. As expected, NE wind was associated with upwelling and SW wind was associated with cold fronts increasing the concentrations of POM and pigments. Factor 3 explained 12.4% of the total variability and was associated with BP. On the other hand, factor 4 was associated with the concentration of chlorophylls and justified 10.2% of the data variability. Polling factors 3 and 4, it was possible to speculate on a change of trophic state in the system as another environmental disturbance. Projecting factors 2 and 3 (Figure 5) the association between BP and ammonia was evident.

Table 2 shows data variability in two samples realized in 2007 during sewage discharges. BP, Cl a and suspended particles were highest in PA station. The average of suspended particles decreased from Anjos and Forno inlets to PC station. Meanwhile, POM was highest in PI and PC areas. PC also had the highest ratios between the abundance of non fluorescent and fluorescent (NF/F) particles, as well as the highest ratios between the abundance of non fluorescent particles and the concentration of chlorophyll a (NFpart/Cla). These data indicated that upwelling and cold fronts ressuspended inorganic and organic particles with size bigger than particles inputted during sewage emissions.

The abundance of small particles was highest in inshore stations during upwelling and in the PC station during the subsidence period. The concentration of red fluorescent particles was higher at PO/PV area. Upwelled waters had the highest NF/F ratio and the smallest concentration of particles with orange and yellow/green fluorescence (data not shown). Cold fronts events are expected to decrease the spatial variability of NF/F ratio due to their homogenizing effect in the water column.

Discussion

Our results confirmed that upwelling is the main process determining the abiotic and biotic characteristics of the surface waters in Arraial do Cabo, followed by the winds dynamics, as observed by Valentin and colleagues (1984; 1987) in a fixed station (Focinho do Cabo). Hydrological data were variable during sampling periods and stations (Tables 1 and 2), confirming constant disturbances in the system. The average values of BP was according to previous studies in the region (Carvalho and Gonzalez-Rodriguez, 2004; Guenther et al., 2008; Cury et al., 2011) but its range showed that BP was higher in the stations inner the embayment (Figure 2). Wide coastal area and its extension can maintain the elevated upwelling intensity and chlorophyll concentrations in upwelling systems and the importance of Cabo Frio embayment to the system production should be evaluated in future studies (Coelho-Souza et al., 2012). Furthermore, sewage discharge in the Anjos beach was the main factor increasing bacterial production (Figure 3) and it was coupled with high chlorophyll concentrations.

Abiotic factors such as temperature and nutrient availability usually determine temporal and spatial patterns in the ecosystem and are considered the first selective pressures on growth, survival and metabolic activity of microorganisms (Terrado et al., 2009). BP should be inhibited by low temperatures during upwelling (Kirchman et al., 1995; Tibbles, 1996), and the mixture and warming of the upwelled water is needed to activate the bacteria (Carvalho and Gonzalez-Rodriguez, 2004; Guenther et al., 2008). Cury et al. (2011) reported higher BP values in surface than in deep waters and Zubkoz et al. (2004) observed that amino acid uptake was related to sampling depth and associated it with phytoplankton photosynthetic activity as well as with light intensity and quality (Seymour et al., 2000; Coelho-Souza et al., 2013).

On the other hand, biotic variables, such as abundance, biomass and metabolic activity are also essential for a better understanding of microbial ecology. Nevertheless, some technical limitations impose caution on the interpretation of microbial data. For example, correlations between the number of individuals, their biomass and metabolic activity are not always observed (Andrade et al., 2007). This limits the projection of patterns from other directly measured variables (McArthur, 2006).

Specifically, measurements of bacterial activity are done in closed samples which can modify bacterial metabolism (Pace et al., 2004). The sampling process to evaluate microorganism density and diversity is another challenge for microbial ecology studies. The aggregated distribution of communities requires sub-sampling to be statistically determined by its confidence limits. Thus, the interpretation of results, in addition to the analyzed variables, must also take into account the sampling and sample processing methods (Karl and Dore, 2001).

Bacterial production and the nitrogen cycle

Natural eutrophication by upwelling events is usually associated with inorganic nitrogen inputs (mainly nitrate) that consequently increase the biomass and size of phytoplankton (Guenther et al., 2008). Artificial anthropogenic eutrophication by sewage discharges increased the concentration of ammonia (Figure 3) and, probably, the dissolved organic nitrogen (DON) input in the system. It makes the trophic web longer (Suzuki et al., 1996) since picoplankton prefers to assimilate ammonia than nitrate (Harrison and Wood, 1988).

Many authors suggested that the metabolic balance between autotrophy and heterotrophy indicates the autochthonous and allochthonous inputs in a system (e.g., Cottrell et al., 2006; Thottathil et al., 2008). In this way, Pereira et al. (2010) associated the heterotrophic/autotrophic ratio and the viral abundance with upwelling events in the Cabo Frio system. These authors reported a highest concentration of heterotrophic prokaryotes in the Anjos inlet. They also observed a high abundance of phytoplankton and a highest viral/bacterial ratio close to PI station (Pereira et al., 2009). Our results showed that PI had hydrological difference from all other stations (Figure 4) probably due its influence from coastal, tropical and subtropical waters (Pereira et al., 2009; 2010) that increases the proportion of live cells (e.g., Cl a) in POM (Tables 1 and 2). Therefore, PI should be considered as the main station under autochthonous production due mixing of rich and warm waters. Bacteria use carbon and nutrients from this green web but are also regulated by virus (Pereira et al., 2009). As expected, PC had more pressure of upwelling and cold fronts and stations inner the embayment were divided in function of the highest frequency of north coastal waters in IP and FnC. Unexpectedly, PV grouped with the main anthropogenic impacted stations (PA, FnF and FO) suggesting that water circulation decreases punctual impacts but can disperse pollutants to adjacent areas (Ribeiro, 2002).

Our results indicated a change in the energy source associated with anthropogenic activities; confirming an autochthonous input associated with upwelling and an allochtonous input associated with sewage discharges. We suggest that autochtonous production is associated with ascension of inorganic nutrients and benthic particles from deep water during upwelling (Guenther et al., 2008) that is quickly exported (Valentin et al., 1987) while marine snow is precipitated on the bottom sediment. During cold fronts, carbon and nutrients are ressuspended, water column is homogenized and POM as well as Cl a increases (Figure 5). We hypothesize a regenerated production during cold fronts as a function of the new production during upwelling. However, the equilibrium is broken during sewage discharges since there is a high input of dissolved organic matter that is consumed by autotrophic and heterotrophic microorganisms.

Comparing the lowest (0.2 μgC.L^-1.h^-1) and highest (78.2 μgC.L^-1.h^-1) values of BP in PA station, we observed a ~400-fold increase during sewage discharges (Tables 1 and 2). BP was well associated with ammonium concentrations (Figure 3). Kirchman (1994) suggested that nitrogen is the main element associated with marine bacterioplankton that can also regulate carbon and phosphorus absorption (Azam et al., 1994; Berman and Bronk, 2003). Hoch et al. (2008) observed that organic nitrogen compounds are the main stimulus to bacterial activity. Following up, ammonification of dissolved organic nitrogen (DON) favors the uptake of inorganic nitrogen by bacteria (Kirchman, 1994). Herein, a high range in ammonia concentrations was reported in all stations, but the peaks of nitrate and nitrite were highest in the area directly influenced by upwelling (PC). We suggest different pathways of ammonia production and oxidation in the nitrogen cycling.

Following Cury et al. (2011), Anjos embayment had lower microbial diversity than an area directly influenced by upwelling, mainly for Bacteria and Eukarya domains. Prokaryotes and Eukaryotes are important contributors in NH4⁺ cycling since eukaryotes support prokaryotic needs, including uptake and oxidation processes (Molina et al., 2005). Simulating upwelled waters mixture, Carvalho and Gonzalez-Rodriguez (2004) observed that ammonia concentration was associated with auto and heterotrophic activities, as well as with the water type predominance. Ammonia can also facilitate nitrate assimilation by primary producers (Bode and Varela, 1994) and it is in accordance with the high chlorophyll concentrations reported during sewage discharges observed visually. Blooms of opportunistic and toxic phytoplankton are another risk for the environment (Coelho-Souza et al., 2006).

Ammonifying bacteria were the primary source of ammonium in the Antarctic coastal waters (Serebrennikova et al., 2008). In the Chilean upwelling system, in the oxycline base, NH4⁺ was in high demand by various components of the bacterioplankton community with different metabolisms: aerobic-anaerobic, photoautotrophic, chemoautotrophic, and heterotrophic. Specifically, the aerobic ammonium oxidation was more regulated by NH4⁺ than O₂ availability and contributed to more than 30% of the total dark carbon fixation (Molina and Farías, 2009).

Some planktonic Archaea are able to take up amino acids, being considered as heterotrophic (Alderkamp et al., 2006). In the Cabo Frio system, for Archaea domain, Crenarchaeota was found only in surface waters from the anthropogenic impacted area and this group uses ammonia as its sole energy source (Cury et al., 2011). Following our results obtained during sewage emissions, we suggest that bacteria are producing ammonia from DON and that Crenarchaeota is consuming ammonia. A significant negative relation was observed between Crenarchaeota and NH4⁺ in the Black Sea (Stoica and Herndl, 2007).

Conclusion

The main risk of eutrophication is the establishment of dead zones due to hypoxia when microorganism growth reduces the concentration of dissolved oxygen in the water column (Diaz and Rosenberg, 2008). Though hypoxia can be observed in upwelling regions (Molina and Farías, 2009), no oxygen depletion was observed in the Cabo Frio upwelling system (Valentin, 1984; Gonzalez-Rodriguez et al., 1992; Guenther et al., 2008) probably due to a high water circulation (Seymour et al., 2000). Coastal ecosystems with short residence times tend to export more exogenous nitrogen (Cloern, 2001) but eutrophication can also deteriorate adjacent ecosystems (Hoch et al., 2008) since the fluxes of nutrients and energy are altered (Madigan et al., 2004; Garren et al., 2008). In addition, harbor activities ressuspend sediment and nitrogen that could be exported to the continental shelf and to the open ocean (Fulweiler et al., 2007).

Embayment areas favor the growth of microorganisms but vast sewage dispersion and phytoplankton blooms are rare in the Cabo Frio embayment. On the other hand, there is no information about the compounds present in the Arraial do Cabo sewage and studies associating this discharges with toxicological consequences from domestic chemical products are needed (Schindler, 2006; Soares et al., 2008). Furthermore, bacterial activity is both associated with environmental remediation and pollutants dispersion (Madigan et al., 2004; Coelho-Souza et al., 2006; 2011; Guimaraes et al., 2006).

The view of eutrophication is biased from observations in few systems (eg., Taketani et al., 2003; Araujo and Hagler, 2011). This scientific discipline is still very young to marine systems and a broader view of costal eutrophication will consider how anthropogenic nutrient enrichment interact with other stressors such as species translocation, habitat loss, fishing, inputs of toxic contaminants, manipulation of freshwater flows, aquaculture, and climate change. Ammonium accumulation in water column should influence the communities of benthic primary producers. Consequently, the overgrowth of macroalgae could cause replacement of corals, as well as other changes in benthic invertebrate populations (Cloern, 2001).

In the Arraial do Cabo protection area, we should consider the signs of disturbances that occur before the catastrophic symptoms. These early responses include changes in communities at the species level, indirect responses, changes in the ratios of biogeochemical processes such as nutrient cycling, and shifts in seasonal patterns or in their variability (Cloern, 2001). To avoid ecosystem deterioration, an adequate coastal management is strictly necessary; especially in marine protection areas like Arraial do Cabo (Floeter et al., 2001). Following Nixon (2009), scientists, regulators, politicians, and even activists need to consider coastal marine eutrophication not as a simple "pollution problem" but as a major ecological change that must be viewed through the macroscope.

Acknowledgments

We thank the IEAPM logistic support, especially to the Grupo de Química, the Departament of Oceanography, Bioincrustação Lab team and LEG 10 team. This study would not have been concluded without the help of María Soledad López, Dagles Viana dos Reis, Marco Lourenço, Márcia Sueli Nagethe, Antônio Casarin, Laudemir Martins da Silva, Fábio Araújo and William Romão. An anonymous referee brought significant improvements to the final format of this manuscript. This work was supported by CAPES, CNPq and FAPERJ.

Submitted: August 3, 2012

Approved: April 4, 2013

All the content of the journal, except where otherwise noted, is licensed under a Creative Commons License CC BY-NC.

• Alderkamp AC, Sintes E, Herndl GJ (2006) Abundance and activity of major groups of prokaryotic plankton in the coastal North Sea during the spring and summer. Aquat Microb Ecol 45:237-246.
• Andrade L, Gonzalez AM, Rezende CE, Suzuki M, Valentin JL, Paranhos R (2007) Distribution of HNA and LNA bacterial groups in the Southwest Atlantic ocean. Braz J Microbiol 38:330-336.
• Araujo FV, Hagler AN (2011) Kluyveromices aestuarii, a potential environmental quality indicator yeast for mangroves in the state of Rio de Janeiro, Brazil. Braz J Microbiol 42:954-958.
• Azam F, Smith DC, Steward GF, Hagstrom A (1994) Bacteria - organic-matter coupling and its significance for oceanic carbon cycling. Microb Ecol 28:167-179.
• Berman T, Bronk DA (2003) Dissolved organic nitrogen: a dynamic participant in aquatic ecosystems. Aquat Microb Ecol 31:279-305.
• Bode A, Varela M (1994) Planktonic carbon and nitrogen budgets for the N-NW Spanish shelf: The role of pelagic nutrient regeneration during upwelling events. Scient Mar 58:221-231.
• Carvalho WF, Gonzalez-Rodriguez E (2004) Development of primary and bacterial productivity in upwelling waters of Arraial do Cabo, RJ (Brazil). Braz J Oceanogr 55:35-45.
• Cloern JE (2001) Our evolving conceptual model of the coastal eutrophication problem. Mar Ecol Prog Ser 210:223-253.
• Coelho-Souza AS, Guimaraes JRD, Mauro JBN, Miranda MR, Azevedo S (2006) Mercury methylation and bacterial activity associated to tropical phytoplankton. Sci Tot Environ 364:188-199.
• Coelho-Souza SA, Guimaraes JRD, Miranda MR, Poirier H, Mauro JBN, Lucotte M, Mergler D (2011) Mercury and flooding cycles in the Tapajós river basin, Brazilian Amazon: the role of periphyton of a floating macrophyte (Paspalum repens). Sci Tot Environ 409:2746-2753.
• Coelho-Souza SA, López MS, Guimaraes JRD, Coutinho R, Candella R (2012) Biophysical interactions in the Cabo Frio upwelling system, Southeastern Brazil. Braz J Oceanogr 60:353-365.
• Coelho-Souza SA, Miranda MR, Salgado LT, Coutinho R, Guimaraes JRD (2013) Adaptation of the ³H-Leucine incorporation technique to measure heterotrophic activity associated with biofilm on the blades of the seaweed Sargassum spp. Microb Ecol 65:424-436.
• Cottrell MT, Malmstrom RR, Hill V, Parker AE, Kirchman DL (2006) The metabolic balance between autotrophy and heterotrophy in the western Arctic Ocean. Deep-Sea Res Part I-Oceanogr. Res Pap 53:1831-1844.
• Cumming G, Fidler F, Vaux DL (2007) Error bars in experimental biology. J Cell Biol 177:7-11.
• Cury JC, Araujo FV, Coelho-Souza SA, Peixoto RS, Oliveira JAL, Santos HF, Davila AMR, Rosado AS (2011) Microbial diversity of a Brazilian coastal region influenced by an upwelling system and anthropogenic activity. Plos One. 6(1), e16553. doi:10.1371/journal.pone.0016553.
• Diaz RJ, Rosenberg R (2008) Spreading dead zones and consequences for marine ecosystems. Science 321:926-929.
• Dubelaar GBJ, Gerritzen PL, Beeker AER, Jonker RR, Tangen K (1999) Design and first results of CytoBuoy: A wireless flow cytometer for in situ analysis of marine and fresh waters. Cytometry 37:247-254.
• Ferreira CEL, Goncalves JEA, Coutinho R (2004) Ship hulls and oil platforms as potential vectors to marine species introduction. J Coast Res 3:1340-1345.
• Floeter SR, Guimaraes RZP, Rocha LA, Ferreira CEL, Rangel CA, Gasparini JL (2001) Geographic variation in reef-fish assemblages along the Brazilian coast. Glob Ecol Biogeogr 10:423-431.
• Fulweiler RW, Nixon SW, Buckley BA, Granger SR (2007) Reversal of the net dinitrogen gas flux in coastal marine sediments. Nature 448:180-182.
• Garren M, Smriga S, Azam F (2008) Gradients of coastal fish farm effluents and their effect on coral reef microbes. Environ Microbiol 10:2299-2312.
• Gonzalez-Rodriguez E, Valentin JL, André DL, Jacob SA (1992) Upwelling and downwelling at Cabo Frio (Brazil): comparison of biomass and primary production responses. J Plankton Res 14:289-306.
• Guenther M, Gonzalez-Rodriguez E, Carvalho WF, Rezende CE, Mugrabe G, Valentin JL (2008) Plankton trophic structure and particulate organic carbon production during a coastal downwelling-upwelling cycle. Mar Ecol Progr Ser 363:109-119.
• Guimaraes JRD, Mauro JBN, Meili M, Sundbom M, Haglund AL, Coelho-Souza AS, Hylander L.D. (2006) Simultaneous radioassays of bacterial production and mercury methylation in the periphyton of a tropical and a temperate wetland. J Environ Manag 81:95-100.
• Harrison WG, Wood LJE (1988) Inorganic nitrogen uptake by marine picoplankton - evidence for size partitioning. Limnol Oceanogr 33:468-475.
• Hoch MP, Dillon KS, Coffin RB, Cifuentes LA (2008) Sensitivity of bacterioplankton nitrogen metabolism to eutrophication in sub-tropical coastal waters of Key West, Florida. Mar Poll Bul 56:913-926.
• James FC, McCulloch CE (1990) Multivariate-analysis in ecology and systematics - panacea or pandora box. An Rev Ecol Syst 21:129-166.
• Karl DM, Dore JE (2001) Microbial ecology at sea: Sampling, subsampling and incubation considerations. Met Microbiol 30:13-39.
• Kirchman DL (1993) Leucine incorporation as measure of biomass production by heterotrophic bacteria. In: P.F. Kemp, B.F. Sherr, E.B. Sherr and J.J. Cole (eds.) Handbook of methods in aquatic microbial ecology Lewis, Boca Raton, FL, pp. 509-512.
• Kirchman DL (1994) The uptake of inorganic nutrients by heterotrophic bacteria. Microb Ecol 28:255-271.
• Kirchman D (2001) Measuring bacterial biomass production and growth rates from leucine incorporation in natural aquatic environments. Met Microbiol 30:227-237.
• Kirchman D, Knees E, Hodson R (1985) Leucine incorporation and its potential as a measure of protein-synthesis by bacteria in natural aquatic systems. App Environ Microbiol 49:599-607.
• Kirchman DL, Rich JH, Barber RT (1995) Biomass and biomass production of heterotrophic bacteria along 140-degrees-w in the equatorial pacific - effect of temperature on the microbial loop. Deep-Sea Res Part II-Top St Oceanogr 42:603-619.
• Lehmann A, Myrberg K (2008) Upwelling in the Baltic Sea - A review. J Mar Syst 74:S3-S12.
• Lopez MS, Coutinho R (2010) Positive interaction between the native macroalgae Sargassum sp. and the exotic bivalve Isognomon bicolor? Braz J Oceanogr 58:69-72.
• Madigan MT, Martinko JM, Parker J (2004) Microbiologia de Brock Pearson Prentice Hall, São Paulo, SP, 606 pp.
• McArthur JV (2006) Microbial ecology - an evolutionary approach Elsevier, San Diego, CA, 416 pp.
• Michelou VK, Cottrell MT, Kirchman DL (2007) Light-stimulated bacterial production and amino acid assimilation by cyanobacteria and other microbes in the North Atlantic Ocean. App Environ Microbiol 73:5539-5546.
• Miranda MR, Guimaraes JRD, Coelho-Souza SA (2007) [H-3]Leucine incorporation method as a tool to measure secondary production by periphytic bacteria associated to the roots of floating aquatic macrophyte. J Microbiol Met 71:23-31.
• Molina V, Farías L (2009) Aerobic ammonium oxidation in the oxycline and oxygen minimum zone of the eastern tropical South Pacific off northern Chile (~20°S). Deep-sea Res II 56:1032-1041.
• Molina V, Farías L, Eissler Y, Cuevas LA, Morales CE, Escribano R (2005) Ammonium cycling under a strong oxygen gradient associated with the Oxygen Minimum Zone off northern Chile (~23° S). Mar Ecol Prog Ser 288:35-43.
• Nixon SW (2009) Eutrophication and the macroscope. Hydrobiol 629:5-19.
• Pace ML, del Giorgio P, Fischer D, Condon R, Malcom H (2004) Estimates of bacterial production using the leucine incorporation method are influenced by differences in protein retention of microcentrifuge tubes. Limnol Oceanogr Met 2:55-61.
• Pereira GC, Granato A, Figueiredo AR, Ebecken NFF (2009) Virioplankton abundance in trophic gradients of an upwelling field. Braz J Microbiol 40:857-865.
• Pereira GC, Figueiredo AR, Jabor PM, Ebecken NFF (2010) Assessing the ecological status of plankton in Anjos Bay: a flow cytometry approach. Biogeosci Dis 7:6243-6264.
• Ribeiro VP (2002) Avaliação do potencial da espécie Stramonita haemastoma (Kool, 1987) como indicador biológico de contaminação ambiental por compostos organo-estânicos. Msc thesis Universidade Federal Fluminense, Rio de Janeiro, Brazil, 108 pp.
• Richard TA, Thompson TG (1952) The estimation and characterization of plankton pigments. J Mar Res 11:6.
• Schindler DW (2006) Recent advances in the understanding and management of eutrophication. Limnol Oceanogr 51:356-363.
• SCOR-UNESCO (1966) Determination of photosynthetic pigments in sea water. In: Monographs in oceanographic methods UNESCO, New York, 69 pp.
• Serebrennikova Y, Fanning KA, Walsh JJ (2008) Modelling the nitrogen and carbon cycling in Marguerite Bay, Antarctica: Annual variations in ammonium and net community production. Deep-Sea Res II 55:393-411.
• Seymour JR, Mitchell JG, Pearson L, Waters RL (2000) Heterogeneity in bacterioplankton abundance from 4.5 millimeter resolution sampling. Aquat Microb Ecol 22:143-153.
• Silva PP (2004) From common property to co-management: lessons from Brazil's first maritime extractive reserve. Mar Pol 28:419-428.
• Simon M (1991) Isotope-dilution of intracellular amino-acids as a tracer of carbon and nitrogen-sources of marine planktonic bacteria. Mar Ecol Progr Ser 74:295-301.
• Simon M, Azam F (1989) Protein-content and protein-synthesis rates of planktonic marine-bacteria. Mar Ecol Progr Ser 51:201-213.
• Smith DC, Azam F (1992) A simple, economical method for measuring bacterial protein synthesis rates in seawater using 3H-leucine. Mar Microb Food Webs 6:107-114.
• Soares A, Guieysse B, Jefferson B, Cartmell E, Lester JN (2008) Nonylphenol in the environment: A critical review on occurrence, fate, toxicity and treatment in wastewaters. Environ Internat 34:1033-1049.
• Stoica E, Herndl GJ (2007) Contribution of Cranarchaeota and Euryarchaeota to the prokaryotic plankton in the coastal northwestern Black Sea. J Plankton Res 29:699-706.
• Strickland JD, Parsons TR (1972) A practical handbook of seawater analysis, 167. In: Bulletin Fishery Research Board - Canada. 310 pp.
• Suzuki MT, Sherr EB, Sherr BF (1996) Estimation of ammonium regeneration efficiencies associated with bacterivory in pelagic food webs via a N-15 tracer method. J Plankton Res 18:411-428.
• Taketani RG, Araujo FV, van Weerelt MDM, Sanchez-Nunez ML, Aniszewski, Rosado AS (2003) Use of molecular approach to verify the influence of a eutrophic lagoon in the near-by ocean's bacterioplankton communities. Braz J Microbiol 34:51-52.
• Terrado R, Vincent WF, Lovejoy C (2009) Mesopelagic protists: diversity and succession in a coastal Arctic ecosystem. Aquat Microb Ecol 56:25-39.
• Thottathil SD, Balachandran KK, Gupta GVM, Madhu NV, Nair S (2008) Influence of allochthonous input on autotrophic-heterotrophic switch-over in shallow waters of a tropical estuary (Cochin Estuary), India. Est Coast Shelf Sci 78:551-562.
• Tibbles BJ (1996) Effects of temperature on the incorporation of leucine and thymidine by bacterioplankton and bacterial isolates. Aquat Microb Ecol 11:239-250.
• Valentin JL (1984) Analysis of hydrobiological parameters in the Cabo Frio (Brazil) upwelling. Mar Biol 82:259-276.
• Valentin JL (2001) The Cabo Frio upwelling system, Brazil. In: U. Seeliger and B. Kjerfve (eds). Coastal marine ecosystems of Latin America: Ecological studies Springer, Berlin, pp 97-105.
• Valentin JL, Andre DL, Jacob SA (1987) Hydrobiology in the Cabo Frio (Brazil) upwelling two-dimensional structure and variability during a wind cycle. Cont Shelf Res 7:77-88.
• Zubkov MV, Tarran GA, Fuchs BM (2004) Depth related amino acid uptake by Prochlorococcus cyanobacteria in the Southern Atlantic tropical gyre. FEMS Microbiol Ecol 50:153-161.

Publication Dates

• Publication in this collection
  27 Mar 2014
• Date of issue
  Dec 2013

History

• Received
  03 Aug 2012
• Accepted
  04 Apr 2013