Cyanobacteria community in two tropical eutrophic reservoirs in northeastern Brazil

This study investigated the effect of seasonality on the Cyanobacteria community in two tropical eutrophic reservoirs in northeastern Brazil. Monthly samplings were conducted in the sub-surface of reservoirs. The following abiotic variables were analyzed: apparent color, turbidity, conductivity, alkalinity, calcium, magnesium, sodium, potassium, sulfate, total phosphorous, ammonia, nitrate, nitrite, chlorides, total hardness and chlorophyll-a, according to APHA (2012). Cyanobacteria density was quantified through biomass. Data were analyzed using one-way ANOVA and Redundancy Analysis (RDA). Higher mean values of Cyanobacteria biomass occurred in the dry (9.9mm L) and rainy (19.0mm L) season in Jucazinho and Carpina reservoirs, respectively, especially Planktothrix agardhii, Cylindrospermopsis raciborskii and Geiterinema amphibium, which occurred in 100% of the samples. RDA analysis revealed that the variables nitrite, ammonia, conductivity, calcium, sulfate, chlorides and alkalinity influenced most the Cyanobacteria community in both reservoirs. High trophic state index was observed throughout the year. Seasonality affected both biotic and abiotic variables.


Introduction
The occurrence of Cyanobacteria has been observed in various environments (freshwater, brackish, marine and terrestrial), but prevalent in freshwater ecosystems (LEE, 2008). Certain characteristics make some Cyanobacteria genera particularly suitable for blooming, such as presence of sheath and mucilage; specialized cells for nitrogen fixation (heterocysts) and resistance (akynetes); aerotopes; tolerance to large fluctuations in temperature; phosphorus storage capacity, among others (DOKULIL; TEUBNER, 2000).
In tropical reservoirs, temperature has no significant influence on phytoplankton dynamics, due to little variations with values usually high above, the limiting ones to growth, but seasonal patterns of phytoplankton communities cannot be neglected (FIGUEREDO;GIANI, 2009). In these ecosystems, environmental conditions are influenced by rainfall events (BORGES et al., 2008, CHELLAPPA et al., 2008, which alter the volume and level of the ecosystem, being especially important to the dynamics of the phytoplankton community (ALMEIDA et al., 2012 Plankton communities depend on adaptations to biotic and abiotic characteristics of each environment. Changes in the trophic status of reservoirs can cause serious damages to the system, especially when eutrophication takes place, since this process interferes with ecological balance and leads to alterations in limnological conditions (ALMEIDA et al., 2012, BARBOSA et al., 2014. Climate changes and eutrophication have provided ecological conditions for the development of cyanobacteria in a large number of Brazilian reservoirs. In works carried out over the past five years in eutrophic reservoirs in Northeastern Brazil, there are frequent reports of cyanobacterial blooms (LIRA et al., 2009, ALMEIDA et al., 2012. This region has relatively uniform seasonal characteristics, such as a dry season (summer), with high temperatures and low rainfall index, and a rainy season (winter), with milder temperatures and a more intense rainfall regime .
Studies have correlated the dynamics of cyanobacterial communities with environmental variables in an attempt to propose models and patterns for eutrophic reservoirs. However, the spatial and temporal heterogeneity in reservoirs allow, even for a short period, sudden changes in the composition of biological communities and also in the limnological parameters, thus producing distinct ecological dynamics. The Capibaribe River basin is considered one of the largest and most important of the Pernambuco State, used for multiple purposes and constantly subjected to anthropogenic eutrophication along its mainly urban stretch.
Accordingly, the hypothesis of this study is that the seasonal variation in rainfall promotes significant fluctuations in hydrological variables, and hence modifies the structure and composition of the Cyanobacteria community in two tropical eutrophic reservoirs in Northeastern Brazil.
Thus, this work aims to contribute to better understanding of the role of environmental variables on cyanobacterial ecology in two large eutrophic reservoirs in the Capibaribe River basin (Pernambuco State, Brazil), as well as contribute to a survey on the cyanobacteria species recorded.

Sampling and laboratory analysis
Samples for qualitative and quantitative analysis of the cyanobacteria community were collected monthly between January 2013 and December 2014 at three sampling sites in the limnetic zone, during two seasons: rainy (March to August) and dry (January to February and September to December) seasons. All samples were obtained in duplicate under the water surface (approximately 30 cm deep), collected, preserved and stored in accordance with APHA (2012).
Data on air temperature (°C) and rainfall (mm) were obtained from the National Institute for Space Research (INPE) and from the Pernambuco Agency of Water and Climate (APAC), respectively, in meteorological stations located at a maximum of 2 km distance from the sampling sites.
TN: TP molar ratio was determined according to Downing et al. (1992). For the estimation of nutrient limitation, TN: TP < 20 was considered as limited by nitrogen; TN: TP > 38 was considered as limited by phosphorus (KOSTEN et al., 2009 The trophic state index was calculated based on Carlson (1977).
Analysis of variance (ANOVA) was used to test differences in abiotic and biotic variables (p < 0.05) between rainy and dry seasons using the Statistica software (test version). Redundancy Analysis (RDA) was applied to assess associations between Cyanobacteria and environmental variables, using the R software. The matrix with biotic data was constructed only with species of cyanobacteria with higher biomass than 5% of the total biomass.

Results
Data of air temperature and rainfall confirmed the occurrence of two distinct seasons in the regions where the Jucazinho and Carpina reservoirs are located. The dry and rainy seasons were characterized by average temperatures above 23ºC. Rainfall was higher in the rainy season in comparison to the dry season, which showed an average rainfall of 3mm (Jucazinho Reservoir).
Limitation of nitrogen throughout the study was observed in both reservoirs. Although the average concentrations of the three nitrogen inorganic forms (ammonia, nitrite and nitrate) have not shown significant differences between the seasons, higher values were verified in the rainy season in both reservoirs, especially the nitrate concentrations with average concentrations above 0.3 μmol in both reservoirs. Regarding pH, alkaline values occurred throughout the study, but without significant differences between the seasons, as well as for the other parameters.
In the reservoirs, values of chlorophyll-a and total phosphorous classified these waters as eutrophic (Jucazinho) or hypereutrophic (Carpina) throughout the study.
In this study, total biomass of cyanobacteria exceeded 1.0 mm 3 L -1 in all samples. Greater biomass of all cyanobacteria species occurred during the rainy and dry seasons in Carpina and Jucazinho reservoirs, respectively, as shown in Table 1. Planktothrix agardhii (Gomont) Anagnostidis and Komárek (1988) occurred on 100% of samples, with greater mean biomass in Carpina (mean: 9.9 ± 2.1 mm 3 L -1 ) than in Jucazinho (mean: 5.3 ± 1.0 mm 3 L -1 ). This species represented 71.8 and 88.6% of the total biomass of Jucazinho and Carpina reservoirs, respectively, considered a dominant species in both reservoirs.
There was a significant difference in biomass between dry and rainy seasons in both reservoirs (ANOVA: F = 7.84 and F = 8.72, p < 0.05 for Jucazinho and Carpina, respectively). Table 1. Average values of Cyanobacteria biomass (mm 3 L -1 ) in Carpina and Jucazinho reservoirs during rainy and dry seasons; + = occurrence of biomass lower than 0.1 mm 3 L -1 ; -= absence; DS = dry season; RS = rainy season.

Taxon
Carpina Jucazinho DS RS DS RS Oscillatoriales Geitlerinema amphibium (C. Agardh) Anagnostidis (1989) Planktothrix agardhii (Gomont) Anagnostidis and Komárek (1988)  The results of the Redundancy Analysis (RDA) are illustrated in Figure 1. The Monte Carlo test proved significant (p < 0.01). There were relationships between environmental variables and biological factors. Eigenvalues of Axis 1 and 2 explained 55.3% of the variance in biological data. The correlation of species biomass with environmental conditions was high in both axes.
RDA Axis 1 separated the reservoirs. The sampling units in Jucazinho Reservoir were positively related to this axis, whereas those in the Carpina Reservoir were negatively related. Regarding to the intra-set correlation, the variables associated positively to Axis 1 were nitrite and ammonia and a group of sampling units of the Jucazinho Reservoir was observed. Calcium, alkalinity, sulfate, chloride and conductivity were negatively associated with Axis 1 and a group of sampling units of the Carpina reservoir was verified. Cylindrospermopsis raciborskii (Woloszynska) Seenaya and Suba Raju (1972), P. agardhii and Geitlerinema amphibium (C. Agardh) Anagnostidis (1989) were inversely related with Axis 1 and were the most abundant taxa in the Carpina Reservoir. C. raciborskii was also associated with higher conductivity and lower values of nitrite and ammonia. P. agardhii and G. amphibium were associated with higher values of calcium, sulfate, chloride and alkalinity. These three  The second axis showed temporal variations between years (2012 and 2013) and seasons (dry and rainy) both in Jucazinho and Carpina reservoirs. C. raciborskii, P. agardhii and G. amphibium were mostly associated with sampling units of the Carpina Reservoir. Non-N-fixing species (P. agardhii and G. amphibium) were positively related to this axis, whereas the N-fixing species (C. raciborskii) was negatively related. The variables positively associated to Axis 2 were chloride, sulfate, alkalinity and calcium and conductivity was negatively associated with this axis. As observed in the Carpina Reservoir, P. agardhii, C. raciborskii and G. amphibium were also abundant in the Jucazinho reservoir.

Discussion
Most limnological parameters analyzed, both biotic and abiotic, showed higher concentrations in the Carpina Reservoir, than Jucazinho. It was possibly because the Carpina reservoir is downstream of Jucazinho, and receives the load of organic matter from the stretch between the two reservoirs.
Both reservoirs showed high TSI, classifying the reservoirs as eutrophic (Jucazinho) and hypereutrophic (Carpina). Moura et al. (2007b), and Lira et al. (2011), studying Brazilian reservoirs, observed that these environments remained eutrophic or hypereutrophic throughout the year, mainly in the drier months.
The electrical conductivity showed high levels in both reservoirs throughout the study period, ranging from 1,146 to 1,961 μS cm -1 . This parameter is considered a good indicator of water quality, since high values may indicate eutrophication, possibly due to the decay of organic matter that release large amount of ions in the water body. This report is supported by studies in eutrophic reservoirs performed by Gemelgo et al. (2009) Alkaline pH values were registered throughout the study in both reservoirs, as well as in studies carried out in eutrophic reservoirs in northeastern Brazil (MOURA et al., 2007b. The aquatic communities can interfere with pH and algae can raise the pH through photosynthesis. The preference of Cyanobacteria from neutral to alkaline environments is due to their ability to use bicarbonate as a form of inorganic carbon and their limited ability to regulate and maintain neutral the internal pH in acidic ecosystems (LEE, 2008).
High concentrations of phosphorus and nitrogen forms were verified in all samples, corroborating studies in eutrophic ecosystems undertaken by Moura et al. (2007a), Gemelgo et al. (2009, Teixeira de Oliveira et al.(2011), Dantas et al., 2012, Piccin-Santos and and . However, seasonality has some influence on these macronutrient concentrations. The phosphorous showed the highest concentration in the dry season, a result also found by  in studies in eutrophic reservoirs. Nevertheless, higher concentrations of this macronutrient and nitrogen forms were found in the rainy season, confirming Moura et al. (2007b), Teixeira de Oliveira et al. (2011), Dantas et al. (2012) and Bezerra et al. (2014). The profile for nitrogen was confirmed in this study and may be due to the input of sediments and nutrients from the reservoir margins. Finally, studies performed by Moura et al. (2007a) and Dellamano-Oliveira et al. (2008) detected smaller concentrations of both nutrients during the rainy season, possibly as a result of the dilution effect of rainfall. The characterization of the studied areas shows that the upstream stretch of Carpina and Jucazinho reservoirs is subjected to intense human activities, considering the agglomeration of several municipalities along the banks of the Capibaribe River. Both regions lack sewage treatment in urban and rural areas, which contribute with the input of undesirable chemicals in the reservoir water, as well as accelerate eutrophication process.
Although phosphorus is frequently considered as the most important limiting factor for algal growth (DELAZARI-BARROSO et al., 2007), nitrogen limitation is recognized as the ideal condition for the occurrence of cyanobacterial blooms, reported in several studies in eutrophic ecosystems, highlighting the predominance of heterocystous species (MOURA et al., 2007a, 2007b, DELAZARI-BARROSO et al., 2007BICUDO, 2008, BEZERRA et al., 2014, COSTA et al., 2014, including reservoirs of Northeastern Brazil, probably due the high availability of phosphorous . Nevertheless, studies also demonstrated the predominance of cyanobacteria under phosphorus limiting conditions (CHELLAPPA et al., 2008, FIGUEREDO;GIANI, 2009) or found no relationship between cyanobacterial dominance and TN: TP ratio in eutrophic reservoirs .
Analyzing the RDA (Figure 1), it was observed that the nitrogen forms NO 2 and NH 3 were negatively correlated with C. raciborskii (N-fixing species), P. agardhii and G. amphibium (non-N-fixing species). This result was similar to that reported by Fonseca and Bicudo (2008), considering negative correlation with cyanobacteria total density and Willame et al. (2008), who considered negative correlation with cyanobacteria efficient in competing for N. This result suggests that these filamentous species preferred the oxidized form of nitrogen (nitrate). However, studies highlighted that C. raciborskii abundance decreases with the decreasing inorganic nitrogen (TONETTA et al., 2014).
With respect to conductivity, there was a positive correlation with the cyanobacteria species identified, corroborating a study carried out in the same reservoir by Lira et al. (2011). As shown in Figure 1, P. agardhii, C. raciborskii and G. amphibium had positive correlation, meaning that the same environmental conditions are favorable for the growth of these three taxa. Fonseca and Bicudo (2008) pointed out that species of Nostocales were positively correlated with conductivity values.
Despite high throughout the year, higher cyanobacteria biomass values were found during the dry season in the Jucazinho Reservoir and in the rainy season in the Carpina Reservoir. Permanent cyanobacterial dominance is regarded as the ultimate phase of eutrophication (DOKULIL; TEUBNER, 2000). The results found for the Carpina Reservoir are in accordance to Lira et al. (2011), in a study carried out in the same reservoir. Teixeira de Oliveira et al. (2011), Lira et al. (2011 and Oliveira et al. (2011) showed a predominance of cyanobacteria during the rainy season, due to the displacement of allochthonous nutrients, which enriched the ecosystem. Nevertheless, higher biomass, mainly of filamentous species, was found during the dry season in many reservoirs (MOURA et al., 2007b, DELLAMANO-OLIVEIRA et al., 2008, TUNDISI et al., 2008. Reduced biomass in the rainy season can occur due to the hydrological instability caused by the intense rainfall events, characteristic of the onset of the rainy season in tropical regions. The influence of seasonality on cyanobacteria biomass may occur by the regulation of light penetration into water bodies. In the present study, cyanobacteria biomass had negative correlation with turbidity in both reservoirs. Although with low values, the turbidity was higher in the dry season in the Carpina Reservoir, and in the rainy season in the Jucazinho Reservoir; some studies indicate that greater values of turbidity can be more advantageous to Oscillatoriales and Nostocales (FIGUEREDO;GIANI, 2009). Moura et al. (2007a) found higher turbidity values in the rainy season in a reservoir of the Pernambuco State, corroborating the results found in Jucazinho Reservoir.
Blooms and coexistence of P. agardhii and C. raciborskii have been frequently reported in tropical eutrophic reservoirs (FIGUEREDO; GIANI, 2009)  period in both reservoirs. They are associated with the highest values of turbidity (R-strategist species), temperature, conductivity, pH and phosphorous (GEMELGO et al., 2009). In the present study, there were higher values of Oscillatoriales biomass (P. agardhii and G. amphibium), followed by Nostocales (C. raciborskii). According to Moura et al. (2007a), in the Carpina Reservoir, higher densities of Oscillatoriales than Nostocales were verified. In eutrophic ecosystems in northeastern Brazil, the dominance of a single species throughout the year has been recorded (MOURA et al., 2007b).
C. raciborskii may dominate some eutrophic reservoirs for long periods, especially under nitrogen limiting conditions . C. raciborskii blooms have been reported (DOKULIL;TEUBNER, 2000, TONETTA et al., 2014, including in the Pernambuco State . Studies in eutrophic reservoirs have registered P. agardhii blooms (DOKULIL; TEUBNER, 2000, MOURA et al., 2007b, GEMELGO et al., 2009GIANI, 2009. In agreement with Reynolds (2002), this species is classified as a S1 functional group, characterized by inhabiting turbid and mixed environments with low light penetration. Furthermore, it was shown that Planktothrix compete well under N-limitation and is able to survive long periods of nitrogen deficiency (WILLAME et al., 2008). The conditions in the Jucazinho Reservoir were conducive to the occurrence of S1 associations often accompanied by C. raciborskii (Sn functional group) and Aphanizomenon gracile (Lemmermann) Lemmermann (H1 group) .
The persistence of cyanobacteria is probably favored by the high nutrient concentrations in eutrophic systems (DELLAMANO-OLIVEIRA et al., 2008) or due to the synergism of several factors (BORGES et al., 2008, FIGUEREDO;GIANI, 2009. Long hydraulic retention times, allied to the other physical and chemical parameters, may have favored the development of cyanobacteria, resulting in high densities in the reservoirs, as observed by Gemelgo et al. (2009). For Jucazinho and Carpina reservoirs, the theoretical hydraulic retention time is about 401 days and 109 days, respectively. Short residence times cause biomass loss by hydraulic washout and mechanical shock in phytoplankton cells (TEIXEIRA DE OLIVEIRA et al., 2011).
Seasonal models for many physical and chemical parameters are common in tropical reservoirs, which are basically defined by the strong influence of rainfall. It should be stressed that the majority of species that coexist in cyanobacterial blooms also form part of this group and are capable of living in adverse conditions and grow competitively when conditions are favorable (LIRA et al., 2011).
In the present study, in the Carpina Reservoir, the rainy season provided conditions of competitive equality among the opportunistic species, leading to a reduction in P. raciborskii dominance by 5%. A. gracile was sensitive to the increased rainfall, but is opportunists and establishes rapid growth under situations of increased light penetration. Although relatively low, the change in the dominance of P. agardhii in the rainy season was sufficient for the establishment of opportunistic species, which began to coexist. Nevertheless, in Jucazinho Reservoir, the transition from dry to the rainy season increased the dominance of P. agardhii in the cyanobacteria community about 4%, due to the disappearance of Microcystis aeruginosa (Kützing) Kützing (1846) and Merismopedia tenuissima Lemmermann (1898), which were sensitive to the increased rainfall in the reservoir.
The cyanobacteria community in the reservoirs is strongly influenced by biotic and abiotic factors, undergoing the effects of seasonality. This study represents a significant contribution to the determination of ecology of cyanobacteria in eutrophic aquatic ecosystems, used for multiple purposes, taking into account the effect of seasonality.

Conclusion
This study showed higher values of cyanobacteria biomass in the dry (9.9 mm 3 L -1 ) and rainy (19.0 mm 3 L -1 ) seasons in Jucazinho and Carpina reservoirs, respectively, especially P. agardhii, C. raciborskii and G. amphibium, which occurred in 100% of the samples. RDA analysis revealed that the variables nitrite, ammonia, conductivity, calcium, sulfate, chlorides and alkalinity influenced most the Cyanobacteria community in both reservoirs. Seasonality affected both biotic and abiotic variables.