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Spatial and temporal variations in sediment enzyme activities and their relationship with the trophic status of Erhai Lake
Ecological Engineering 75 (2015) 365–369
Contents lists available at ScienceDirect
Ecological Engineering journal homepage: www.elsevier.com/locate/ecoleng
Short communication
Spatial and temporal variations in sediment enzyme activities and their relationship with the trophic status of Erhai Lake Li Zhang a,b , Shengrui Wang a,b, * , Akio Imai c a
State Key Laboratory of Environmental Criteria and Risk Assessment, Chinese Research Academy of Environmental Sciences, Beijing 100012, PR China State Environmental Protection Key Laboratory for Lake Pollution Control, Research Center of Lake Eco-environment, Chinese Research Academy of Environmental Sciences, Beijing 100012, PR China c National Institute for Environmental Studies, Tsukuba, Ibaraki, Japan b
A R T I C L E I N F O
A B S T R A C T
Article history: Received 20 July 2014 Received in revised form 12 November 2014 Accepted 28 November 2014 Available online xxx
The spatial and temporal variations in the activity of six microbial enzymes (invertase, urease, alkaline phosphatase, protease, peroxidase, and polyphenoloxidase) present in the sediment of Erhai Lake, China were investigated. Enzyme activities peaked in summer. This phenomenon can be attributed to high temperature and sediment organic matter accumulation, which could be traced to the breakdown of algal bloom. In addition, sediment enzyme activities exhibited apparent spatial heterogeneity across the lake, but individual enzymes showed similar spatial patterns of activity. Moreover, sediment enzyme activities were found to be closely related to the water quality of Erhai Lake. Enzyme activities were also significantly correlated with sediment total nitrogen and organic matter, implying that the sediment enzyme can be used as a biological indicator that reflects the sediment status. The investigation of sediment enzyme activities can facilitate a comprehensive understanding of the process of nutrient cycling and the mechanisms underlying lake eutrophication. ã 2014 Elsevier B.V. All rights reserved.
Keywords: Erhai Lake sediment Enzyme activity Trophic status Eutrophication
1. Introduction Sediments and soils represent some of the most complex microbial habitats on Earth (Zeng et al., 2010). Microorganisms in soil are important components of the ecosystem (Wu et al., 2014). Sediment microorganisms and their enzymes serve essential functions in lake ecosystems because they are responsible for the decomposition and remineralization of organic matter (OM) into nutrients, thereby strongly influencing the nutrient cycle and energy flux in aquatic ecosystems. Many attempts have been made to investigate the variation in enzyme activity with a focus on the factors that influence this activity; these factors include temperature, pH, humidity, and nutrients (Wallenstein et al., 2012; Corstanje et al., 2007). In recent years, increasing evidence has suggested that microbial properties, such as microorganisms and microbial enzyme activities, can potentially be used to assess ecosystem function and trophic status in various ecosystems, such as that in constructed wetlands (Lei et al., 2012), forests (Pang et al., 2009), and rivers (Hill et al., 2010).
* Corresponding author at: Chinese Research Academy of Environmental Sciences, State Key Laboratory of Environmental Criteria and Risk Assessment, Beijing 100012, PR China. Tel.: +86 010 84915277. E-mail address: [email protected] (S. Wang). http://dx.doi.org/10.1016/j.ecoleng.2014.11.043 0925-8574/ ã 2014 Elsevier B.V. All rights reserved.
Erhai Lake is the second largest fault lake in Yunnan Province, China. Agricultural area source pollution, mainly derived from livestock, farmland, and aquaculture pollution from the north and west, is the main exogenous OM source in this basin. The chemical oxygen demand (COD), total nitrogen (TN), total phosphorus (TP), and NH4–N from agricultural area source pollution into the lake are approximately 9040.1, 2493, 164.8, and 602.8 t/a, respectively. Moreover, the Dali government is located in the southern part of Erhai Basin, in which major human settlements exist, and pollution from wastewater discharge is a serious concern. Activated sludge process, membrane bioreactor, and oxidation ditch are adopted as the main treatment measures for the urban sewage in the basin. The amounts of COD, TN, TP, and NH4–N from urban sewage discharge into the lake are approximately 444.4, 82.1, 6.9, and 56.8 t/a, respectively. Although considerable attention has been directed toward Erhai Lake, the characteristics and variations of sediment enzymes and the effect of sediment enzymes on the eutrophication of this lake remain poorly understood. In this study, the main objectives are (1) to characterize the spatial and temporal variations in the activities of six microbial enzymes (invertase, urease, alkaline phosphatase, protease, peroxidase, and polyphenoloxidase) present in the sediment of Erhai Lake and (2) to discuss the interaction between the sediment enzyme activities and the trophic status of Erhai Lake.
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2. Materials and methods 2.1. Study sites and sampling procedure Erhai Lake can be divided into a northern section, a middle section, and a southern section on the basis of the lake bottom topography. From the standpoint of pollution source, Erhai Lake is seriously impacted by the agricultural area source pollution (northern part and western coast) and human activity (southern part). Therefore, we chose 10 targeted and representational sites (sampled in August, 2011) (Fig. 1), which are representative of the whole lake from the overall characteristics in different regions, to reflect the actual situation of Erhai Lake comprehensively. The detailed descriptions of each sampling site were introduced by Zhang et al. (2014). Sampling was performed using a gravity sediment corer with a length of 30 cm and an internal diameter of 5 cm. To reflect the characteristics of surface sediment accurately, mixed sediment samples from a 10 cm depth of the surface layer (Yang et al., 2010; Chen et al., 2011; Zhang et al., 2013a) were selected as the representative for surface sediment analysis. Then, one portion of the sediment samples was reserved for the measurements of enzyme activity and was placed into a sterile and sealed plastic bag, stored in an icebox (4 C), and analyzed in the laboratory within 24 h. The other portion was reserved for chemical analysis, freeze-dried, and sieved through a standard 100-mesh sieve. 2.2. Physical and chemical analyses The pH and temperature values were determined using a multiparameter water quality monitoring instrument (YSI 6600V2). The TN, TP, OM, and organic phosphorous (OP) contents in sediment, as
well as the TN, TP, and NH4–N levels in overlying water, were measured according to the methods introduced by Zhang et al. (2013a,b). Chlorophyll-a (Chla) in overlying water was measured on the basis of optical density spectrophotometry (State Environmental Protection Administration of China (SEPA), 2002). COD is identified as a national environmental monitoring indicator in China. In this study, we use COD as the key pollution control indicator to reflect the polluted degree of the overlying water. COD was determined using a fast digestion-spectrophotometric method based on the Standard of the People’s Republic of China for Environmental Protection (2007). Sediment enzyme activities were determined as described by Guan (1986). 3. Results 3.1. Temporal and spatial variations in sediment enzyme activities in Erhai Lake Throughout the year, activities of sediment enzymes exhibited similar seasonal patterns. The enzyme activities peaked during spring and summer but remained low during the cold seasons (autumn and winter). Besides, the activities of the six sediment enzymes generally exhibited spatial heterogeneity (Fig. 2), although the spatial patterns of activity of the individual enzymes were similar and significantly correlated (P < 0.01). Sampling sites with high OM content (EH21, EH74, EH132, and EH142) exhibited high enzyme activities. Meanwhile, sediment enzyme activities were low in areas with medium OM content (EH93 and EH105) and relatively deep water. 4. Discussion 4.1. Impact of temperature on the seasonal variation of sediment enzyme activities In our seasonal field study, temperature seemed to affect the seasonal variations of the sediment enzyme activities significantly. Significant positive correlations were observed between sediment enzyme activities and water column temperature (Table 1), which is in accordance with the earlier observations made by Duarte et al. (2008) and Sabine et al. (2005). Increased temperature can enhance the affinity between the enzyme and the substrate, thereby improving the catalytic rate of an enzyme. Simultaneously, the microbial metabolic rate increases with an increased temperature, thereby causing microorganisms to allocate resources to the synthesis and secretion of enzymes (Wallenstein et al., 2012). 4.2. Impact of algae on seasonal variation in sediment enzyme activities
Fig. 1. Geographic location of the sampling sites in Erhai Lake (marked by triangles).
Algae enrichment could help improve enzyme activities in the water column and sediment (Sabine et al., 2005). Fig. 3 shows the Chla concentration in the water column of Erhai Lake throughout the year. In the Erhai ecosystem, the algal biomass in the water column (reflected by the Chla concentration) varies in a manner similar to the activities of sediment enzymes. In autumn and winter (November–February), low temperatures, Chla concentrations, and substrate quantity result in low sediment enzyme activity values. During spring, the increment in water level and the increased temperature and intensity of sunlight in Erhai Basin resulted in marked algal growth. By the time of the summer flood in July, more allochthonous OM is channeled into Erhai Lake, thus, providing energy and nutrient sources that promote microorganism and algal growth. During this growth process, algae in the water column contribute a large amount of fresh autochthonous OM to the sediment. This type of OM contains low C/N levels and is
L. Zhang et al. / Ecological Engineering 75 (2015) 365–369
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Fig. 2. Spatial distributions of enzyme activities (invertase, urease, alkaline phosphatase, protease, peroxidase, and polyphenoloxidase) in the surface sediment of Erhai Lake.
thus easily decomposed and utilized by microorganisms. Consequently, the microbial enzyme activities peak in summer during the annual cycle. This phenomenon is attributed to the high accumulation of sediment OM, which can be traced back to the breakdown of the algal bloom.
4.2.1. Impact of surface sediment enzyme activities on the trophic status of Erhai Lake The enzyme activities in Erhai surface sediment are significantly correlated with TN and COD in the water column (Table 1), indicating that the sediment enzymes may contribute markedly to
Table 1 Correlation analysis of six enzyme activities in sediment and physicochemical parameters in overlying water. Enzymes
Temperature
TN
NH4+
TP
SRP
Chla
COD
Carlson index
Invertase Polyphenoloxidase Peroxidase Alkaline Phosphatase Protease Urease
0.373* 0.554** 0.319* 0.441** 0.535** 0.666**
0.407** 0.518** 0.302 0.454** 0.551** 0.614**
0.105 0.163 0.004 0.056 0.211 0.215
0.109 0.116 0.020 0.149 0.117 0.225
0.376* 0.112 0.298 0.178 0.350* 0.162
0.237 0.299 0.026 0.167 0.266 0.315*
0.336* 0.535** 0.369* 0.379* 0.552** 0.610**
0.688 0.601 0.236 0.611 0.850 0.570
Notes: *, <0.05; **, <0.01.
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Fig. 3. Variation trend of Chla concentration in water column throughout the whole year.
the endogenous load of lake ecosystems. With improving sediment enzyme activities, the decomposition rate of OM by the enzymes is accelerated. More nutrients are released into the interstitial water and enter the overlying water, resulting in an increase in the degree of lake eutrophication. By contrast, the relationship between enzyme activities and NH4+, TP, and SRP is not as significant (Table 1). This phenomenon may be attributed to the abundant factors (e.g., OM content, microbial activity, and nutrient cycling) affecting the autochthonous release from Erhai Lake sediment, which weaken the impact of microbial enzymes relative to that of the other factors. The classical Carlson trophic state index (CTSI) (Carlson, 1977) was used to assess the trophic status of shallow lakes (including algae-dominated and macrophyte-dominated lakes) in China (Zhang et al., 2014). CTSI was calculated with the average values from five limnological parameters, namely, Chla, TP, TN, transparency, and COD. The comprehensive CTSI ranks the lakes and reservoirs using a numerical scale from 0 to 100. The scale values correspond to different trophic state levels: 0 < CTSI 30 is the oligotrophic level, 30 < CTSI 40 is the oligomesotrophic level, 40 < CTSI 50 is the mesotrophic level, 50 < CTSI 60 is the light eutrophic level, 60 < CTSI 70 is the middle eutrophic level, and 70 < CTSI 100 is the hypereutrophic level (Hu et al., 2014). According to this criterion, Erhai Lake is presently at the status of mesotrophication. In this study, positive correlations exist between the Carlson index and the sediment enzyme activities (Table 1). However, the correlation is insignificant. This result may be attributed to the relatively high water depth in Erhai Lake, which results in the decomposition or utilization of nutrient/algae by microorganisms in the water column during sedimentation from water surface to sediment. This phenomenon may weaken the correlation between OM/algae and sediment enzymes, as reflected in the low or even insignificant correlation coefficients between the Carlson index and the sediment enzyme activities. However, the experimental results could still indicate that the sediment enzyme activities are closely related to the lake trophic status. 4.3. Relationship of enzyme activities with the sediment nutrient The enzyme activities are significantly positively correlated with sediment TN and OM, with correlation coefficients of 0.86 0.06 and 0.91 0.06, respectively. This result implies that the sediment enzyme activities can be used as a biological indicator of the sediment status. Protease, urease, and alkaline phosphatase are important extracellular hydrolases and can convert protein, urea, and orthophosphate monoesters into amino acid, ammonium, and orthophosphate, respectively. These compounds can be directly utilized by microorganisms. Humus is believed to have a positive
effect on maintaining the stability of enzymes (Dong et al., 2007). Humic acid can protect urease from being oxidized and degraded or from being inhibited by metal ions under alkaline conditions. In Erhai sediment, OM content is high, particularly at sites EH21, EH73, EH132, and EH142, where the sediment is black. This condition reflects a high degree of humification. Thus, the high OM or humus concentrations also ensure the high stability of protease and urease at these sites, thereby inhibiting their release to the overlying water to some extent. Peroxidase and polyphenoloxidase are extracellular enzymes that markedly contribute to the breakdown of several organic compounds (Oyekola and Pletschke, 2006). In Erhai sediment, OM is high. In addition, significant correlations exist between OM and peroxidase (R2 = 0.909, P < 0.01) and polyphenoloxidase activities (R2 = 0.967, P < 0.01). Hence, the high peroxidase and polyphenoloxidase activities in Erhai sediment are likely caused by the substrate-inducing effect. Moreover, peroxidase and polyphenoloxidase bind to OM and humic substances to form stable compounds (Bolag and Mayers, 1992) in sediment, which is believed to be responsible for the special status of the Erhai ecosystem, i.e., highly polluted sediment, but fairly good water quality. 5. Conclusion Enzyme activities exhibited an apparently seasonal and spatial pattern in Erhai sediment and were closely correlated with the nutrient level in the lake ecosystem. Temperature was believed to affect the seasonal variations of the sediment enzymes activities markedly. A positive correlation existed between the Carlson trophic state index and the sediment enzyme activities, indicating that the sediment enzyme activities are closely related to the lake trophic state. Furthermore, the spatial pattern of sediment enzyme activities was also closely related to the sediment nutrient content. The high sediment OM or humus content helped to stabilize protease and urease. Peroxidase and polyphenoloxidase were verified to be capable of binding to OM and humic substances to form stable compounds in the sediment. Such binding was considered to be responsible for the special status of the Erhai ecosystem, i.e., fairly good water quality despite highly polluted sediment. Acknowledgements This research was supported by the National Natural Science Foundation of China (nos. U1202235, 41103070), the Major Science and Technology Program for Water Pollution Control and Treatment (no. 2012ZX07102-004), the Free Exploration Foundation of State Key Laboratory of Environmental Criteria and Risk Assessment, Chinese Research Academy of Environmental Sciences, and the Scientific Research Foundation for the Returned Overseas Chinese Scholars, State Education Ministry. References Bolag, J.M., Mayers, C., 1992. Detoxification of aquatic and terrestrial sites through binding of pollutants to humic substances. Sci. Total Environ. 117–118, 357–366. Carlson, R.E., 1977. A trophic state index for lakes. Limnol. Oceanogr. 22, 361–369. Chen, J.J., Lu, S.Y., Zhao, Y.K., Wang, W., Huang, M.S., 2011. Effects of overlying water aeration on phosphorus fractions and alkaline phosphatase activity in surface sediment. J. Environ. Sci. 23, 206–211. Corstanje, R., Reddy, K.R., Prenger, J.P., Newman, S., Ogram, A.V., 2007. Soil microbial eco-physiological response to nutrient enrichment in a subtropical wetland. Ecol. Indic. 7, 277–289. Dong, L.H., Yang, J.S., Yuan, H.L., Wang, E.T., Chen, W.X., 2007. Chemical characteristics and influences of two fractions of Chinese lignite humic acids on urease. Eur. J. Soil Biol. 44, 166–171. Duarte, B., Reboreda, R., Caçador, I., 2008. Seasonal variation of extracellular enzymatic activity (EEA) and its influence on metal speciation in a polluted salt marsh. Chemosphere 73, 1056–1063.
L. Zhang et al. / Ecological Engineering 75 (2015) 365–369 Guan, S.Y., 1986. Soil Enzyme and its Research Methods. Agriculture Press, Beijing. Hill, B.H., Elonen, C.M., Jicha, J.M., Bolgrien, D.W., Moffett, M.F., 2010. Sediment microbial enzyme activity as an indicator of nutrient limitation in the great rivers of the Upper Mississippi River basin. Biogeochemistry 97, 195–209. Hu, Z.X., Guo, L.Y., Liu, T., Chuai, X.M., Chen, Q.K., Shi, F., Jiang, L.J., Yang, L.Y., 2014. Uniformisation of phytoplankton chlorophyll a and macrophyte biomass to characterize the potential trophic state of shallow lakes. Ecol. Indic. 37, 1–9. Lei, H., Xu, G., Ming, L., Gang, D., Jinsong, G., Theoneste, N., 2012. Correlation among soil microorganisms, soil enzyme activities, and removal rates of pollutants in three constructed wetlands purifying micro-polluted river water. Ecol. Eng. 46, 98–106. Oyekola, O., Pletschke, B., 2006. ATP-sulphurylase: an enzymatic marker for sulphate reduction. Soil Biol. Biochem. 38, 3511–3515. Pang, X.Y., Ning, W., Liu, Q., Bao, W.K., 2009. The relation among soil microorganism, enzyme activity and soil nutrients under subalpine coniferous forest in Western Sichuan. Acta Ecol. Sinica 29, 286–292. Sabine, W., Helmut, F., Martin, P.T., 2005. Regulation and seasonal dynamics of extracellular enzyme activities in the sediments of a large lowland river. Microb. Ecol. 50, 253–267. Standard of the People’s Republic of China for Environmental Protection (2007). Water Quality—Determination of the Chemical Oxygen Demand—Fast Digestion-Spectrophotometric Method, HJ/T 399-2007. (In Chinese). State Environmental Protection Administration of China (SEPA), 2002. Monitor and Analysis Method for Water and Wastewater, fourth ed. China Environmental Science Press (in Chinese).
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Wallenstein, M.D., Haddix, M.L., Lee, D.D., Conant, R.T., Paul, E.A., 2012. A litter-slurry technique elucidates the key role of enzyme production and microbial dynamics in temperature sensitivity of organic matter decomposition. Soil Biol. Biochem. 47, 18–26. Wu, Q., Wang, X.F., Li, Y., Zhao, H.B., Sen, P., 2014. Response of rhizosphere bacterial diversity to phytoremediation of Ni contaminated sediments. Ecol. Eng. 73, 311–318. Yang, C.X., Wang, S.R., Jin, X.C., Wu, F.C., 2010. Nitrogen and phosphorus mineralization in sediments of Taihu Lake after the removal of light fraction organic matter. Environ. Earth Sci. 59, 1437–1446. Zeng, X., Xiao, X., Wang, F.P., 2010. Response of bacteria in the deep-sea sediments and the Antarctic soils to carbohydrates: effects on ecto enzyme activity and bacterial community. J. Environ. Sci. 22, 1779–1785. Zhang, L., Wang, S.R., Jiao, L.X., Ni, Z.K., Xi, H.Y., Liao, J.Y., Zhu, C.W., 2013a. Characteristics of phosphorus species identified by 31P NMR indifferent trophic lake sediments from the Eastern Plain, China. Ecol. Eng. 60, 336–343. Zhang, L., Wang, S.R., Jiao, L.X., Zhao, H.C., Zhang, Y., Li, Y.P., 2013b. Physiological response of a submerged plant (Myriophyllum spicatum) to different NH4Cl concentrations in sediments. Ecol. Eng. 58, 91–98. Zhang, L., Wang, S.R., Wu, Z.H., 2014. Coupling effect of pH and dissolved oxygen in water column on nitrogen release at water–sediment interface of Erhai Lake, China. Estuarine Coastal Shelf Sci. 149, 178–186.
Contents lists available at ScienceDirect
Ecological Engineering journal homepage: www.elsevier.com/locate/ecoleng
Short communication
Spatial and temporal variations in sediment enzyme activities and their relationship with the trophic status of Erhai Lake Li Zhang a,b , Shengrui Wang a,b, * , Akio Imai c a
State Key Laboratory of Environmental Criteria and Risk Assessment, Chinese Research Academy of Environmental Sciences, Beijing 100012, PR China State Environmental Protection Key Laboratory for Lake Pollution Control, Research Center of Lake Eco-environment, Chinese Research Academy of Environmental Sciences, Beijing 100012, PR China c National Institute for Environmental Studies, Tsukuba, Ibaraki, Japan b
A R T I C L E I N F O
A B S T R A C T
Article history: Received 20 July 2014 Received in revised form 12 November 2014 Accepted 28 November 2014 Available online xxx
The spatial and temporal variations in the activity of six microbial enzymes (invertase, urease, alkaline phosphatase, protease, peroxidase, and polyphenoloxidase) present in the sediment of Erhai Lake, China were investigated. Enzyme activities peaked in summer. This phenomenon can be attributed to high temperature and sediment organic matter accumulation, which could be traced to the breakdown of algal bloom. In addition, sediment enzyme activities exhibited apparent spatial heterogeneity across the lake, but individual enzymes showed similar spatial patterns of activity. Moreover, sediment enzyme activities were found to be closely related to the water quality of Erhai Lake. Enzyme activities were also significantly correlated with sediment total nitrogen and organic matter, implying that the sediment enzyme can be used as a biological indicator that reflects the sediment status. The investigation of sediment enzyme activities can facilitate a comprehensive understanding of the process of nutrient cycling and the mechanisms underlying lake eutrophication. ã 2014 Elsevier B.V. All rights reserved.
Keywords: Erhai Lake sediment Enzyme activity Trophic status Eutrophication
1. Introduction Sediments and soils represent some of the most complex microbial habitats on Earth (Zeng et al., 2010). Microorganisms in soil are important components of the ecosystem (Wu et al., 2014). Sediment microorganisms and their enzymes serve essential functions in lake ecosystems because they are responsible for the decomposition and remineralization of organic matter (OM) into nutrients, thereby strongly influencing the nutrient cycle and energy flux in aquatic ecosystems. Many attempts have been made to investigate the variation in enzyme activity with a focus on the factors that influence this activity; these factors include temperature, pH, humidity, and nutrients (Wallenstein et al., 2012; Corstanje et al., 2007). In recent years, increasing evidence has suggested that microbial properties, such as microorganisms and microbial enzyme activities, can potentially be used to assess ecosystem function and trophic status in various ecosystems, such as that in constructed wetlands (Lei et al., 2012), forests (Pang et al., 2009), and rivers (Hill et al., 2010).
* Corresponding author at: Chinese Research Academy of Environmental Sciences, State Key Laboratory of Environmental Criteria and Risk Assessment, Beijing 100012, PR China. Tel.: +86 010 84915277. E-mail address: [email protected] (S. Wang). http://dx.doi.org/10.1016/j.ecoleng.2014.11.043 0925-8574/ ã 2014 Elsevier B.V. All rights reserved.
Erhai Lake is the second largest fault lake in Yunnan Province, China. Agricultural area source pollution, mainly derived from livestock, farmland, and aquaculture pollution from the north and west, is the main exogenous OM source in this basin. The chemical oxygen demand (COD), total nitrogen (TN), total phosphorus (TP), and NH4–N from agricultural area source pollution into the lake are approximately 9040.1, 2493, 164.8, and 602.8 t/a, respectively. Moreover, the Dali government is located in the southern part of Erhai Basin, in which major human settlements exist, and pollution from wastewater discharge is a serious concern. Activated sludge process, membrane bioreactor, and oxidation ditch are adopted as the main treatment measures for the urban sewage in the basin. The amounts of COD, TN, TP, and NH4–N from urban sewage discharge into the lake are approximately 444.4, 82.1, 6.9, and 56.8 t/a, respectively. Although considerable attention has been directed toward Erhai Lake, the characteristics and variations of sediment enzymes and the effect of sediment enzymes on the eutrophication of this lake remain poorly understood. In this study, the main objectives are (1) to characterize the spatial and temporal variations in the activities of six microbial enzymes (invertase, urease, alkaline phosphatase, protease, peroxidase, and polyphenoloxidase) present in the sediment of Erhai Lake and (2) to discuss the interaction between the sediment enzyme activities and the trophic status of Erhai Lake.
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2. Materials and methods 2.1. Study sites and sampling procedure Erhai Lake can be divided into a northern section, a middle section, and a southern section on the basis of the lake bottom topography. From the standpoint of pollution source, Erhai Lake is seriously impacted by the agricultural area source pollution (northern part and western coast) and human activity (southern part). Therefore, we chose 10 targeted and representational sites (sampled in August, 2011) (Fig. 1), which are representative of the whole lake from the overall characteristics in different regions, to reflect the actual situation of Erhai Lake comprehensively. The detailed descriptions of each sampling site were introduced by Zhang et al. (2014). Sampling was performed using a gravity sediment corer with a length of 30 cm and an internal diameter of 5 cm. To reflect the characteristics of surface sediment accurately, mixed sediment samples from a 10 cm depth of the surface layer (Yang et al., 2010; Chen et al., 2011; Zhang et al., 2013a) were selected as the representative for surface sediment analysis. Then, one portion of the sediment samples was reserved for the measurements of enzyme activity and was placed into a sterile and sealed plastic bag, stored in an icebox (4 C), and analyzed in the laboratory within 24 h. The other portion was reserved for chemical analysis, freeze-dried, and sieved through a standard 100-mesh sieve. 2.2. Physical and chemical analyses The pH and temperature values were determined using a multiparameter water quality monitoring instrument (YSI 6600V2). The TN, TP, OM, and organic phosphorous (OP) contents in sediment, as
well as the TN, TP, and NH4–N levels in overlying water, were measured according to the methods introduced by Zhang et al. (2013a,b). Chlorophyll-a (Chla) in overlying water was measured on the basis of optical density spectrophotometry (State Environmental Protection Administration of China (SEPA), 2002). COD is identified as a national environmental monitoring indicator in China. In this study, we use COD as the key pollution control indicator to reflect the polluted degree of the overlying water. COD was determined using a fast digestion-spectrophotometric method based on the Standard of the People’s Republic of China for Environmental Protection (2007). Sediment enzyme activities were determined as described by Guan (1986). 3. Results 3.1. Temporal and spatial variations in sediment enzyme activities in Erhai Lake Throughout the year, activities of sediment enzymes exhibited similar seasonal patterns. The enzyme activities peaked during spring and summer but remained low during the cold seasons (autumn and winter). Besides, the activities of the six sediment enzymes generally exhibited spatial heterogeneity (Fig. 2), although the spatial patterns of activity of the individual enzymes were similar and significantly correlated (P < 0.01). Sampling sites with high OM content (EH21, EH74, EH132, and EH142) exhibited high enzyme activities. Meanwhile, sediment enzyme activities were low in areas with medium OM content (EH93 and EH105) and relatively deep water. 4. Discussion 4.1. Impact of temperature on the seasonal variation of sediment enzyme activities In our seasonal field study, temperature seemed to affect the seasonal variations of the sediment enzyme activities significantly. Significant positive correlations were observed between sediment enzyme activities and water column temperature (Table 1), which is in accordance with the earlier observations made by Duarte et al. (2008) and Sabine et al. (2005). Increased temperature can enhance the affinity between the enzyme and the substrate, thereby improving the catalytic rate of an enzyme. Simultaneously, the microbial metabolic rate increases with an increased temperature, thereby causing microorganisms to allocate resources to the synthesis and secretion of enzymes (Wallenstein et al., 2012). 4.2. Impact of algae on seasonal variation in sediment enzyme activities
Fig. 1. Geographic location of the sampling sites in Erhai Lake (marked by triangles).
Algae enrichment could help improve enzyme activities in the water column and sediment (Sabine et al., 2005). Fig. 3 shows the Chla concentration in the water column of Erhai Lake throughout the year. In the Erhai ecosystem, the algal biomass in the water column (reflected by the Chla concentration) varies in a manner similar to the activities of sediment enzymes. In autumn and winter (November–February), low temperatures, Chla concentrations, and substrate quantity result in low sediment enzyme activity values. During spring, the increment in water level and the increased temperature and intensity of sunlight in Erhai Basin resulted in marked algal growth. By the time of the summer flood in July, more allochthonous OM is channeled into Erhai Lake, thus, providing energy and nutrient sources that promote microorganism and algal growth. During this growth process, algae in the water column contribute a large amount of fresh autochthonous OM to the sediment. This type of OM contains low C/N levels and is
L. Zhang et al. / Ecological Engineering 75 (2015) 365–369
367
Fig. 2. Spatial distributions of enzyme activities (invertase, urease, alkaline phosphatase, protease, peroxidase, and polyphenoloxidase) in the surface sediment of Erhai Lake.
thus easily decomposed and utilized by microorganisms. Consequently, the microbial enzyme activities peak in summer during the annual cycle. This phenomenon is attributed to the high accumulation of sediment OM, which can be traced back to the breakdown of the algal bloom.
4.2.1. Impact of surface sediment enzyme activities on the trophic status of Erhai Lake The enzyme activities in Erhai surface sediment are significantly correlated with TN and COD in the water column (Table 1), indicating that the sediment enzymes may contribute markedly to
Table 1 Correlation analysis of six enzyme activities in sediment and physicochemical parameters in overlying water. Enzymes
Temperature
TN
NH4+
TP
SRP
Chla
COD
Carlson index
Invertase Polyphenoloxidase Peroxidase Alkaline Phosphatase Protease Urease
0.373* 0.554** 0.319* 0.441** 0.535** 0.666**
0.407** 0.518** 0.302 0.454** 0.551** 0.614**
0.105 0.163 0.004 0.056 0.211 0.215
0.109 0.116 0.020 0.149 0.117 0.225
0.376* 0.112 0.298 0.178 0.350* 0.162
0.237 0.299 0.026 0.167 0.266 0.315*
0.336* 0.535** 0.369* 0.379* 0.552** 0.610**
0.688 0.601 0.236 0.611 0.850 0.570
Notes: *, <0.05; **, <0.01.
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Fig. 3. Variation trend of Chla concentration in water column throughout the whole year.
the endogenous load of lake ecosystems. With improving sediment enzyme activities, the decomposition rate of OM by the enzymes is accelerated. More nutrients are released into the interstitial water and enter the overlying water, resulting in an increase in the degree of lake eutrophication. By contrast, the relationship between enzyme activities and NH4+, TP, and SRP is not as significant (Table 1). This phenomenon may be attributed to the abundant factors (e.g., OM content, microbial activity, and nutrient cycling) affecting the autochthonous release from Erhai Lake sediment, which weaken the impact of microbial enzymes relative to that of the other factors. The classical Carlson trophic state index (CTSI) (Carlson, 1977) was used to assess the trophic status of shallow lakes (including algae-dominated and macrophyte-dominated lakes) in China (Zhang et al., 2014). CTSI was calculated with the average values from five limnological parameters, namely, Chla, TP, TN, transparency, and COD. The comprehensive CTSI ranks the lakes and reservoirs using a numerical scale from 0 to 100. The scale values correspond to different trophic state levels: 0 < CTSI 30 is the oligotrophic level, 30 < CTSI 40 is the oligomesotrophic level, 40 < CTSI 50 is the mesotrophic level, 50 < CTSI 60 is the light eutrophic level, 60 < CTSI 70 is the middle eutrophic level, and 70 < CTSI 100 is the hypereutrophic level (Hu et al., 2014). According to this criterion, Erhai Lake is presently at the status of mesotrophication. In this study, positive correlations exist between the Carlson index and the sediment enzyme activities (Table 1). However, the correlation is insignificant. This result may be attributed to the relatively high water depth in Erhai Lake, which results in the decomposition or utilization of nutrient/algae by microorganisms in the water column during sedimentation from water surface to sediment. This phenomenon may weaken the correlation between OM/algae and sediment enzymes, as reflected in the low or even insignificant correlation coefficients between the Carlson index and the sediment enzyme activities. However, the experimental results could still indicate that the sediment enzyme activities are closely related to the lake trophic status. 4.3. Relationship of enzyme activities with the sediment nutrient The enzyme activities are significantly positively correlated with sediment TN and OM, with correlation coefficients of 0.86 0.06 and 0.91 0.06, respectively. This result implies that the sediment enzyme activities can be used as a biological indicator of the sediment status. Protease, urease, and alkaline phosphatase are important extracellular hydrolases and can convert protein, urea, and orthophosphate monoesters into amino acid, ammonium, and orthophosphate, respectively. These compounds can be directly utilized by microorganisms. Humus is believed to have a positive
effect on maintaining the stability of enzymes (Dong et al., 2007). Humic acid can protect urease from being oxidized and degraded or from being inhibited by metal ions under alkaline conditions. In Erhai sediment, OM content is high, particularly at sites EH21, EH73, EH132, and EH142, where the sediment is black. This condition reflects a high degree of humification. Thus, the high OM or humus concentrations also ensure the high stability of protease and urease at these sites, thereby inhibiting their release to the overlying water to some extent. Peroxidase and polyphenoloxidase are extracellular enzymes that markedly contribute to the breakdown of several organic compounds (Oyekola and Pletschke, 2006). In Erhai sediment, OM is high. In addition, significant correlations exist between OM and peroxidase (R2 = 0.909, P < 0.01) and polyphenoloxidase activities (R2 = 0.967, P < 0.01). Hence, the high peroxidase and polyphenoloxidase activities in Erhai sediment are likely caused by the substrate-inducing effect. Moreover, peroxidase and polyphenoloxidase bind to OM and humic substances to form stable compounds (Bolag and Mayers, 1992) in sediment, which is believed to be responsible for the special status of the Erhai ecosystem, i.e., highly polluted sediment, but fairly good water quality. 5. Conclusion Enzyme activities exhibited an apparently seasonal and spatial pattern in Erhai sediment and were closely correlated with the nutrient level in the lake ecosystem. Temperature was believed to affect the seasonal variations of the sediment enzymes activities markedly. A positive correlation existed between the Carlson trophic state index and the sediment enzyme activities, indicating that the sediment enzyme activities are closely related to the lake trophic state. Furthermore, the spatial pattern of sediment enzyme activities was also closely related to the sediment nutrient content. The high sediment OM or humus content helped to stabilize protease and urease. Peroxidase and polyphenoloxidase were verified to be capable of binding to OM and humic substances to form stable compounds in the sediment. Such binding was considered to be responsible for the special status of the Erhai ecosystem, i.e., fairly good water quality despite highly polluted sediment. Acknowledgements This research was supported by the National Natural Science Foundation of China (nos. U1202235, 41103070), the Major Science and Technology Program for Water Pollution Control and Treatment (no. 2012ZX07102-004), the Free Exploration Foundation of State Key Laboratory of Environmental Criteria and Risk Assessment, Chinese Research Academy of Environmental Sciences, and the Scientific Research Foundation for the Returned Overseas Chinese Scholars, State Education Ministry. References Bolag, J.M., Mayers, C., 1992. Detoxification of aquatic and terrestrial sites through binding of pollutants to humic substances. Sci. Total Environ. 117–118, 357–366. Carlson, R.E., 1977. A trophic state index for lakes. Limnol. Oceanogr. 22, 361–369. Chen, J.J., Lu, S.Y., Zhao, Y.K., Wang, W., Huang, M.S., 2011. Effects of overlying water aeration on phosphorus fractions and alkaline phosphatase activity in surface sediment. J. Environ. Sci. 23, 206–211. Corstanje, R., Reddy, K.R., Prenger, J.P., Newman, S., Ogram, A.V., 2007. Soil microbial eco-physiological response to nutrient enrichment in a subtropical wetland. Ecol. Indic. 7, 277–289. Dong, L.H., Yang, J.S., Yuan, H.L., Wang, E.T., Chen, W.X., 2007. Chemical characteristics and influences of two fractions of Chinese lignite humic acids on urease. Eur. J. Soil Biol. 44, 166–171. Duarte, B., Reboreda, R., Caçador, I., 2008. Seasonal variation of extracellular enzymatic activity (EEA) and its influence on metal speciation in a polluted salt marsh. Chemosphere 73, 1056–1063.
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