Phytoplankton population as alarming warning bioindicator of water pollution in El-Temsah Lake, Egypt

Relative effects of spatio-seasonal variations on the quantitative and qualitative status of phytoplankton in 20 sampling locations of Temsah Lake were examined during winter and summer 2018. The study reveals that the population structure consisted of 105 species from 69 genera, with an annual average of 924155 cell/L. Bacillariophyceae formed the most dominant group with 42.45% of total phytoplankton with an average of 372379 cell/L. Navicula creptocephala was the most abundant species representing 19.16% of total diatoms and 7.78% of the whole standing crop with an annual average of 376879 cell/L. The highest phytoplankton abundance was in the Northern locations. Phytoplankton density showed a negative correlation of pH, temperature and TDS and positively correlated with nutrient content such as nitrate and silicate. In general, summer showed high indices value compared to winter and the diversity index in the current study ranged in moderate pollution range. The polluted state of the aquatic ecosystem in El-Temsah Lake during summer is comparatively higher than winter during the study period. Palmer’s and diversity indices showed that Northern sites were the most ourishing and polluted sites. The study also recorded a number of tolerate algae (13 genera and 8 species), which can be used as an indicator for different degrees of organic pollution. Genus and species Palmer’s Index scores of the northern site were in the range of ≥ 20 during summer, indicating most of the sampling locations showed probable to conrmed high level of organic pollution. polluted sites followed by middle and southern locations of the lowest phytoplankton population. According to Palmer's index and diversity index, most of the sampling locations showed probable to conrmed high level of organic pollution. The study recorded a number of tolerant algae (13 genera and 8 species) which can used as an indicator for different degrees of organic pollution. Therefore, in order to fully understand the response mechanisms associated with phytoplankton ourishing to the climate change phenomenon, the continuity of phytoplankton population research should be considered. These communities could be used as organic pollution indicators for controlling and biomonitoring organic pollution in aquatic environments.


Introduction
Water is the secret of life. It is more important than energy because there are alternative energy sources, but there are no other water sources. World faces rapid deterioration of its owning water. Global population growth and rising living standards contribute to the discharging of tremendous amounts of waste and nutrients into water bodies worldwide (Elshobary et al. 2020). Therefore, it is fundamental to nd environmentally sustainable ways to preserve and protect the valuable water resource. Indeed, continuous monitoring and evaluation of water quality is a crucial concern for sustainable conservation and management. Lakes are often exposed to abrupt environmental vagaries caused by various anthropogenic activities and tourism along their banks. El-Temsah Lake is one of the most common lakes in the Suez Canal of Egypt that receives an immense amount of raw waste discharges (industrial, farming, domestic, leisure, etc.) and tourism activities along its shores. El-Temsah lake is considered a transitional zone connecting two mainly different basins (Red Sea and Mediterranian Sea), affecting ora and fauna distribution (Nassar and Shams El-Din 2006).
Consequently, El-Temsah Lake exhibits serious water quality problems. It is noteworthy that excessive nutrient loading and disruption of water ecological parameters can contribute to eutrophication, resulting in substantial qualitative and quantitative variations in phytoplankton populations and water quality (Xiao et al. 2013;Haque et al. 2019). Phytoplankton blooms cause serious problems in the whole aquatic ecosystems, economy and human survival (Diodato et al. 2005;Hokimoto and Kiyofuji 2014). Furthermore, phytoplankton is directly in uenced by different discharges in addition to human activities and agricultural runoff (Collavini et al. 2001). Also, they have a short life span and rapidly react to any modi cations in the ecosystem, so provide an early warning sign of water deterioration condition and may be helpful for water quality evaluating (El-Kassas and Gharib 2016). Therefore, evaluation of water quality is a critical issue to balance ecological and socio-economic bene ts. Since water quality summarized and de ned forcing variables related to the aquatic ecosystem to be used as a key task for administering water resources on a scienti c basis (Parparov and Hambright 2007). Physicochemical and biological indices are usually used for the assessment of water quality. The former is based on the determination of different physico-chemical parameters in a water sample, whereas the biological indices are dependent on biological evidence such as phytoplankton. Many studies investigated the water quality of Lake El-Temsah till 2016 (Nassar and Shams El-Din 2006;Said and El Agroudy 2006;Donia 2011;Hamed et al. 2012;Kamel 2013;Gaballa 2014;El-Serehy et al. 2018;El-Shoubaky and Hamed 2018;Soliman et al. 2019). All these studies established that the worsening of the lake ecosystem and water quality had been reached a critical level. Thus an urgent action is therefore needed urgently to recover the El-Temsah ecosystem. Therefore, the present study aims to quantify the lake's water quality during 2018 through the physiochemical and biological phytoplankton index. This aim was performed through the numerically mathematical indices of species richness, species diversity, equitability, and Palmer indices to assess these indices' suitability for the maintenance and management of water resources.

Study Areas and sampling locations
This study was carried out in two seasons in August (dry summer season), and February (wet winter season) 2018, where the difference in climate changes was signi cant in these two seasons in Egypt. El-Temsah lake is located near the middle of the Suez Canal about mid-way between the northern city of Port Said and the southern city of Suez at 30°35'46.55''N and 32°19'30.54''E ( Fig. 1). El-Temsah lake is the largest lake of Ismailia city, Egypt, with a total surface area of 14 km with 80 million m 3 . It is a saline shallow water body (3 and 16 m in depth). The lake is connected to a small, shallow lagoon by a narrow passage on the west side. This lagoon is connected with El-Mahsama drains (industrial, agriculture, and sewage e uent). On the north side, the lake occasionally receives input from the Ismailia freshwater canal and agricultural drainage water from the El-Forsan drain. The lake is connected with the Suez Canal from eastern and southern areas (Abd El Samie et al. 2008). Twenty sampling locations were carefully chosen to cover different regions of the lake (Fig.   1).

Environmental Data Collection Water
Water temperature and pH value were measured in situ, using Hydrolab, Model (Multi Set 430i WTW). For the other chemical analysis, water samples were collected in polyethylene bottles and were stored in an icebox upon returning to the laboratory. The collected samples were sieved and ltered through zooplankton net (100 µm mesh size) and Whatman ® GF/F (0.7 µm pore diameter), respectively, and used for chemical analysis. Total Dissolved Solids (TDS), silicate (Si), phosphate (PO3-P), and nitrate NO3-N were measured according to the standard methods of the American Public Health Association (APHA 1998).

Phytoplankton Collection and Enumeration
Triplicate phytoplankton samples with 2 L each were collected in a clean polyethylene bottle from the water surface of the sampling locations. The samples were xed at the sampling site using 10 mL acidic Lugol's iodine and transferred in an icebox to the laboratory. In the laboratory, samples were kept to settle for 48 h. The supernatant was carefully siphoned off via sterilized plastic tubes with plankton net (5µ) and concentrated lower samples stored into 100 mL tight capped bottles in the dark place to prevent light degradation before counting identi cation. Phytoplankton enumeration in triplicate was done by transferring 1 mL into Sedgewick-Rafter counting chamber at 100 and 40 × magni cation using a compound microscope Nikon Eclipse E400 (Nikon, Japan). Phytoplankton was counted (cell/L) and estimated by counting all phytoplankton, whether single cells, colonies, or laments (LeGresley and McDermott 2010). The algal species were identi ed according to standard references (Smith 1950;Desikashary 1959;Hendy 1964;Bourrelly 1981;Prescott 1987). The identi ed species were compared with the latest website of AlgaeBase (http://www.algaebase.org).

Data analysis
Analysis of variance (ANOVA) was assessed to determine the degree of seasonal variation and spatial variation at ≤ 0.05 level of signi cance. Duncan's multiple comparison test was used to compare the average biological and physicochemical parameters using SPSS v.23. The signi cant dissimilarities of phytoplankton communities and environmental conditions in sample locations were tested using one-way Analysis of Similarity (ANOSIM) (Clarke 1993). Similarity percentage analysis (SIMPER) was used to examine genus in uence in the sampling locations using Paleontological Statistics (PAST3).
Phytoplankton richness, equitability indices and biodiversity indices were calculated using Shannon-Wiener diversity index (H'), Margalef (D) and Equitability index (J') to characterize the species diversity in a community across seasons and sampling locations using PAST3. Spatial and temporal physical/chemical patterns were illustrated using contour graphics generated using SURFER 13 (Surfer®, Golden Software, LLC).

Hydrology
Knowledge about different physico-chemical parameters provides a biological activity to assess the ecological quality of an ecosystem. Physicochemical factors play a crucial role in guiding biodiversity in the aquatic environment, contributing to a shift in the diversity and population of aquatic biota (Ramírez et al. 2005;Elshobary et al. 2020). Fig 2 displays the average value of six physicochemical parameters for the twenty locations in Temsah Lake.
The hydrogen ion value (pH) is the leading factor that directs both the chemical and biological activity of the aquatic ecosystem. The pH was slightly alkaline around the year within the standard pH ranged from 6.5 to 8.5 (WHO 2011). The annual mean was 7.55 and uctuated between 6.63 at location 3 during winter to 8.17 at location 13 during summer (Table 1). pH is affected by CO 2 circulation and organic and inorganic contaminants in the water body (Rahman and Huda 2012). The high reported pH value may be owing to the impact of increased biological activity, CO 2 consumption (Govindasamy et al. 2000) and the high photosynthetic rate during summer (Saravanakumar et al. 2008;Sankar et al. 2010).
The water temperature is a key factor as it controls various abiotic and biotic activities of an aquatic ecosystem (Radhika et al., 2004). Stenseth et al. (2005) stated that temperature variation could lead to changes in marine and estuarine ecosystems at all food chain levels, from primary producers (phytoplankton) to top predators. The surface temperature (SWT) uctuated from 24.9 at location 3 to 27.85 ℃ at location 13 (Table 1). The summer season showed the highest SWT might be owing to the intensity of solar radiation, evaporation and water in ux (Kumar et al. 2006;Saravanakumar et al. 2008;Sankar et al. 2010). It was reported that most phytoplankton prefers moderate temperature ranged from 20-35 (Elshobary et al. 2020). Therefore, ve groups of algae were recorded in this study.
Concentrations of total dissolved solids (TDS) in water vary considerably in different geological regions due to differences in minerals' solubilities. The annual TDS uctuated between 2070 ppm at location 2 during summer to 51239 ppm at site 15 during summer (Table 1). The high concentration of TDS is caused mainly by the remnants of inorganic materials and wastewater molecules due to the anthropogenic activities around the water body (Sulistiawati et al. 2020).
The winter season is the leading season, while the concentrations dropped a little bit in the summer. High TDS values during the winter season are attributed to the rains that carry soil and other organic matter from the wetlands to the lake (Sharma and Tiwari 2018). The same nding was observed in the Ismailia canal connected with the lake in 2018 (Elshobary et al. 2020). Ali et al. (2020) demonstrated that TDS in winter exceeded spring, summer and autumn values, respectively. TDS concentrations are very high near the southwestern locations due to the outfall of domestic and industrial wastewater that agreed with previous studies in the same lake during 2011 (Donia 2011) and 2015(El-azim et al. 2018).
Silicate concentration is one of the vital environmental indicators that impact the phytoplankton productivity in the aquatic environment, especially diatoms (Cui et al. 2018). Silicate value was ranged from 0.5 ppm at location 19 during summer to 9.19 ppm at location 3 during winter (Table 1). The availability of silicate in El-Temsah Lake is due to the mixing of agriculture runoff discharged in the lake and connected canals such as Ismailia and Suez canals as well as Bitter Lake. The highest silicate values were observed in winter and reduced in summer; this could be due to the high density of phytoplankton in summer and its high silicate consumption for their biological activity and ourishing during summer (Sankar et al. 2010). These results agreed with (Habib et al. 1997;Elshobary et al. 2020), who showed that silicate concentration in winter was higher than in summer during their studies.
In contrast, autumn is the leader season, followed by spring and summer in Bitter lake (Nassar and Fahmy 2016). These differences have been addressed by Bharathi et al. (2018), who reported that silicate concentration could be varied from season to season according to sampling locations and studying periods in the same water body. Generally, the amount of silica is attributable to the weathering of silicate rocks, in addition to its absorption by diatoms (Wang et al. 2013;El-Otify and Iskaros 2015).
With regards to the trophic conditions of the Temsah lake, nitrate (NO 3 -N) and phosphorus (PO 4 -P) covered a wide range of concentrations, from 1.5 ppm to 4.7ppm and 0.009 ppm to 0.89 ppm, respectively (Table 1). Summer is the leader season in NO 3 -N and PO 4 -P. This nding is agreed with (Abdel-Satar et al. 2017), who demonstrated that NO 3 -N and PO 4 -P values were the maxima in summer compared to winter seasons in the river Nile and Bitter Lakes of Egypt (Nassar and Fahmy 2016). The highest NO 3 -N and PO 4 -P concentrations were observed at location 3 during summer (4. 7 ppm) for nitrate and location 7 during summer (0.89 ppm) for phosphate. The reported highest summer NO 3 -N and PO 4 -P concentration may be mainly due to the organic materials received from the catchment zone (Ashok Prabu et al., 2008). Signi cant spatial and seasonal differences were found for all abiotic variables (p≤0.05) ( Table 1).

phytoplankton composition and percentage frequencies of phytoplankton groups
The quantitative and qualitative status of phytoplankton in Temsah Lake during winter and summer 2018 revealed that its community structure was belonging to ve divisions, namely: Bacillariophyceae, Chlorophyceae, Cyanophyceae, Dinoophyceae and Euglenophyta. A total of 105 species belonging to 69 genera were identi ed during the studying period at different 20 locations covering the whole area of El-Temsah Lake with an annual average of 924155 cell/L. Among those, Bacillariophyceae (26 genera and 39 spp.), Chlorophyceae (24 genera and 33 spp.), Cyanophyceae (9 genera and 15 spp.), Dinophyceae (5 genera and 8 spp.) and Euglenophyceae (3 genera and 10 spp.) as shown in Table (2). This nding is slightly lower than that reported in the recent survey in Temsah lake of Gaballa (2014), who found 116 different genera during 2012. These differences may be associated with spatial and temporal variations in the physiochemical and biological parameters of water bodies (Zhang et al. 2011;Demir et al. 2014; Napiórkowska-Krzebietke and Hutorowicz 2014; Elshobary et al. 2020). The low number of species recorded in the lake can generally be attributed to the persistent discharge of wastewater, leading to increased nutrient supplements hence decreasing predominant species. This nding was a rmed by Ludsin et al. (2001) and Prepas and Charette (2005), who showed that the biodiversity of many aquatic microorganisms reversibly proportions with nutrient content due to increase eutrophication state of the water body.
Bacillariophyceae was the most dominant group and formed about 42.45% of total phytoplankton standing crop with an average of 372379 cell/L. Navicula creptocephala was the most recorded species representing 19.16% of total diatoms and 7.78% of the total standing crop with an annual average of 376879 cell/L. Cyclotella kutzingiana and Diatoma vulgare were also highly recorded during the study period, with an average of 44217.5 and 39460 cell/L, respectively. In a previous study, diatoms were the dominant group in El-Temsah lake, representing 56 and 58%, respectively ( Diatoms exhibit the most dominant group in this study because of their ability to survive under a wide range of environmental factors, such as salinity, temperature and nutrient availability (Huang et al. 2004;Kouhanestani et al. 2019). The same conclusion was observed by Gaballa (2014), who mentioned that phytoplankton's structure was dominated by diatoms followed by Cyanophyta, Dinophyta and Chlorophyta in the same lake during 2012. Elshobary et al.
(2020) have shown that diatoms occupy the rst dominant group, followed by Chlorophyta, Cyanophyta, and Dinophyta in the Ismailia canal connected with Temsah. Otherwise, Euglenophyta, represented by ten species and constituting 0.72% of the total phytoplankton community, is probably represented in this way due to the increase in organic matter pollution in El-Temsah Lake (El-Serehy et al. 2018).

Seasonal variation of total phytoplankton standing crop in El-Temsah Lake
Phytoplankton density recorded in El-Temsah Lake varied greatly from winter to summer, as represented in Table (2). Summer was reported as the most productive season during the study period of 1499563 cell/L representing 80.08% of the total standing crop with phytoplanktonic diversity of 105 species, while winter recorded 348747 cell/L representing only 19.02% of total phytoplankton. Five species (Amphora coffeaeformis, Biddulphia favus, Tetraedron muticum, Scrippsiella spinifera and Trachelomonas hispida) were recorded in summer, which were not represented in winter (Table S1). Elshobary et al. (2020) observed new 11 species were present in summer that were absent in winter in the Ismailia canal. Bacillariophyta was the most dominant phylum in both seasons with 584666 cell/L during summer and 160092 cell/L of winter count. All presented phyla showed the same trend where the maximized productivity was recorded in summer and decreased in winter. While Dinophyta and Euglenophyta showed the lowest community, respectively (Table 2). Adesalu and Nwankwo (2005) demonstrated that Bacillariophyta is ourished in nutrients rich environments of temperate areas. The total phytoplankton and phyla communities at both seasons were signi cantly different at p≤0.001 (Table 2).

Spatial variations
Commonly, the north locations, especially locations 1, 2, 3, and 6, showed the maximum ourishing followed by the central locations (7-13), and the lowest community was observed in the south locations (14-20). Locations 6 and 3 were the most productive and ourishing locations (204,006 and 202,755 cell/L , respectively) followed by locations 2 and 1, respectively. While, the lowest crop was observed at locations 18 and 20 with total average counts of 43,708 and 44,520 cell/L, respectively (Fig. 3). On the other hand, the highest species diversity was recorded in location 6 of the maximum crop with 64 sp. and location 2 with 61 sp. However, the lowest species diversity was detected at site 16 (38 sp.) (Fig. 3). Regarding the phyla diversity, Bacillariophyta was the leading phyla at the most sampling location. Chlorophyta was clearly dominant at 1,2,4,5,6 and 14 locations. While, Cyanophyta was the dominant phyla at location 3 only. The highest Dinophyta community was recorded in location 17, followed by 10. However, the highest Euglenophyta crop was observed at location 16, followed by locations 2 and 3. The ANOVA results show a signi cant spatial difference between the average phytoplankton crop and the number of species at p≤0.001, respectively. The analysis of similarities among 20 locations has been analyzed by One-Way ANOSIM, which pointed out highly signi cant differences (R = 0.982; p = 0.0002), while SIMPER analysis showed overall average dissimilarity of 61.32% among the 20 locations. The most contributing genera with a percentage contribution ≥ 3% were Navicula creptocephala (5.15%), Fragilaria crotonensis (3.39%), Diatoma vulgare (3.18%) and Stephanodiscus astraea (3%). It has been observed that Bacillariophyceae includes most of the contributing taxa. This nding imitates the dominance of these phyla and its direct effect on spatial variation during this study.
3.5. Phytoplankton abundance in relation to some physico-chemical parameters Some physico-chemical parameters were measured, which was observed to be varied spatially and seasonally. Signi cant correlations were observed between phytoplankton abundance and different physico-chemical parameters (Fig.4). Each species inclines to become dominant when the growth conditions meet its speci c requirement. Phytoplankton density showed a negative correlation of pH, temperature and TDS (r = -0.71, -0.62 and -0.56). It was observed that TDS was negatively correlated with total phytoplankton, all algal groups and species diversity. This nding was agreed with previous studies that showed the same negative correlation between phytoplankton population and TDS concentration (Rahi et al. 2013;Elshobary et al. 2020), illustrating the negative effect of inorganic remains materials of TDS on the phytoplankton growth. While Phytoplankton density positively correlated with nutrient content such as Nitrate and silicate. This is due to inorganic nitrogen is one of the main nutrients for algal growth that can improve phytoplankton's growth either in Temsah lake (El-Serehy et al. 2018) or in the connected canals (Elshobary et al. 2020). On the other hand, diatom is the main phytoplankton that can cycle silicon via absorption, dissolution and deposition (Wang et al. 2007). Because the diatom is the dominant group in this study, the total phytoplankton and species number were also correlated positively with silicate content.

Species diversity indices
The diversity indices were proposed to calculate the species richness, species variation, and degree of equitability of relative abundance species (Table 3). In general, summer showed high indices value compared to winter. The diversity index (Shannon-Weaver) of phytoplankton in summer was 2.34-3.61, with an average of 3.0. The high-value area was located in the north of the lake. The value was higher in the north, followed by the middle and the lowest value was located in the south. The highest value was found at locations 2 and 6. This nding is matching with phytoplankton diversity. Noteworthy, Shannon-Weiner diversity index was used as a water pollution indicator, where Wilhm and Dorris (1968) set diversity index<1 for greatly polluted water, intermediate polluted at 1-4 and >4 for unpolluted water. The diversity index in the current study ranged in a moderate pollution range. The richness index (Margalef ) of phytoplankton was quite correlated to the diversity index, where the maximum-value was also observed in the north of the lake and the highest value was found at location 2 and 6. This nding is re ecting the phytoplankton cell abundance. Equitability Index (J') was used by Pielou (1966) to measure species distribution ranged from 0 to 1. However, the average evenness ranged from 0.68 at location 20 to 0.86 at location 6. These values demonstrated that phytoplankton's distributions were quite uniform, and the community structure was unstable (Mahmoud et al. 2018).

Cluster analysis
In order to reveal the similarities among the sampling locations and cluster analysis were done based on the total abundance of the phytoplankton and classes communities as well as environmental parameters (Fig. 5). The results showed the presence of two main clusters with a 100 bootstrap. The rst cluster consists of the northern locations (1-6) and is divided into two subclades of 1-3 locations with 72 bootstraps and 4-6 locations. The second main clade subdivided to rst subclade consisted mostly of middle locations of 7-13 with 91 bootstraps and the second subclade grouped the southern sites of 14-20. This nding re ected that the highest phytoplankton abundance of the northern locations, where these sites are more exposed to pollution discharge than in the middle of the lake, such as oil contamination, domestic outfall, agricultural discharge, and wastewater discharge (El-Serehy et al. 2018). According to the cluster analysis (Fig. 5), a maximum similarity of 93.5% was recorded between locations 1 and 2 followed by 16 and 17. In contrast, the lowest similarity of 72.5% was recorded at locations 1 and 2 and site 3.

Palmer's pollution index
Palmer has published a comprehensive rating of organic pollution tolerate algae, collected by 165 authors, and developed a pollution-tolerant algal index, including 60 most pollution-tolerant algal genera (Palmer 1969). Palmer's index has been calculated seasonally and spatially for genus (AGPI) and species (ASPI). Among these, 13 genera and 8 species were found in El-Temsah Lake (Table 4). Five genera belonging to Bacillariophyta, four genera belonging to Chlorophyta, two genera belonging to Cyanophyta and two genera belonging to Euglenophyta have been recorded. In general, the summer season showed the highest AGPI ranged from 13 to 33 than winter (9-27). Therefore, summer reported the highest pollution levels due to higher pollution loads in summer than winter, indicating the large population and variety of phytoplankton during summer.
In summer, 8 locations (10, 11, 12, 13, 16, 17, 18 and 20) showed probable high pollution and 11 locations (1-9, 14 and 19) showed con rmed high organic pollution most of them are in the north area which showed the highest population and diversity of phytoplankton. Location 15 only showed moderate pollution with score 13. Location 6, 2 and 4 showed the greatest score. These ndings may be due to the fact that northern locations are the closet to people communities, leaving them facing a number of agriculture and industrial contaminations over other locations (El-Serehy et al. 2018).

Conclusion
This study demonstrated that environmental and seasonal uctuations mostly shaped the phytoplankton community of El-Temsah Lake. Bacillariophyta formed the most dominant phyla in both seasons, contributing 42.45% of the total phytoplankton. The polluted state of the aquatic ecosystem in El-Temsah Lake during summer is comparatively higher than winter during the study period. Northern sites were the most fertile and polluted sites followed by middle and southern locations of the lowest phytoplankton population. According to Palmer's index and diversity index, most of the sampling locations showed probable to con rmed high level of organic pollution. The study recorded a number of tolerant algae (13 genera and 8 species) which can used as an indicator for different degrees of organic pollution. Therefore, in order to fully understand the response mechanisms associated with phytoplankton ourishing to the climate change phenomenon, the continuity of phytoplankton population research should be considered. These communities could be used as organic pollution indicators for controlling and biomonitoring organic pollution in aquatic environments.