Comparative Response of Red and Green Algae to the Quality of Coastal Water of Red Sea, Haql, Saudi Arabia

Aquatic plants are always exposed to various types of stresses and the marine algae have been considered as useful bioindicators for detecting different kinds of environmental alterations. Present investigation was carried out to test the comparative response of red algae (Gracilaria salicornia and Digenea simplex) and green algae (Enteromorpha compressa and Sargassum muticum) to the contaminants present in their surrounding growth medium. The results of the study show that the studied macroalgae responded differently in terms of physiological and biochemical parameters. Green algae exhibited higher concentration of Chl a, b, total chlorophyll content and Chl a:b ratio and carbohydrates content. Whereas, red algae showed higher content of carotenoids, phycocyanin and phycoerythrin and protein content. Moreover, red algae G. salicornia and D. simplex showed lower level of hydrogen peroxide content and TBARS and higher values of proline and glycine betaine content and activities of antioxidant enzymes viz. superoxide dismutase, peroxidase, and catalase than the green algae. Taking all these together, it can be concluded that red algae possess strong defense system than the green algae. Among the studied species, red algae G. salicornia was found best in having a stronger defense system.


Introduction
Water is the prime determinant for the existence of life on any planet of the cosmos and water was the medium where life took its shape. The marine ecosys-tem is considered a reservoir of plant and animal diversity and acts as a vital source of primary production. However, discharge of effluents from industries, sewage, agriculture runoff, and construction sites contains a variety of pollutants including toxic metals, detergents, grease, oil, pesticides, nutrients, suspended particles (Kassem, 1999 [1]), which leads to considerable modifications in the quality of coastal waters (Schulz et al., 1994 [2]; Diez et al., 2019 [3]; Eljaiek-Urzola et al., 2019 [4]). Accumulation of nutrients especially N and P in the marine ecosystem supports algal expansion that results in algal blooms (Khan et al., 2013 [5], 2018 [6]). The presence of these algae on the upper surface prevents the penetration of sunlight which affects marine life at lower surfaces. Moreover, death and decay of these plants require a huge quantity of oxygen which depletes oxygen level and such water exhibits higher values of biochemical oxygen demand (BOD) (Ferreira et al., 2017 [7]; Vigiak et al., 2019 [8]) that adversely affects the aquatic ecosystem. The excess and continuous presence of pollutants in the aquatic ecosystem makes their easy entry in the food chain and causes biomagnification which affects human health (Ahmed et al., 2019 [9]).
Because of submergence in water, seaweeds require more light than other plant groups (Dennison et al., 1993 [10]). In the intertidal zone, the seaweeds are constantly exposed to natural as well as anthropogenic sources which adversely affect marine environment and cause severe losses to seaweeds. These sources induce changes in turbidity, dissolved oxygen and nutrient composition of water and photosynthetic pigments of seaweeds. Moreover, the climatic conditions of the study area such as arid environment, low precipitation and no sources of fresh water also contribute to significant alterations in marine environment. These changes act as stressors and adversely affect pigment concentration of seaweeds leading to reduced growth and biomass production. Excessive generation of reactive oxygen species such as hydrogen peroxide (H 2 O 2 ) is the primary response of plants to various environmental stresses. These ROS cause damage to macromolecules, photosynthetic pigments, leakage of electrolytes and ultimately cell death (Scandalios, 1993 [11]; Mittler, 2002 [12]). Plants cope with the detrimental effects of these ROS through their inbuilt system of antioxidant enzymes such as superoxide dismutase (SOD), peroxidase (POX) and catalase (CAT

Water and Plant Sample Collection
Water samples were collected near the water surface at a distance of about 8 meters from the shoreline from three points of the Haql coast of the Red Sea, Saudi Arabia ( Figure 1). Water quality was assessed using the mean values of three sampling data sets.
The plant samples were comprised of 1) Red algae (Rhodophyta); Gracilaria salicornia (G. salicornia), Digenea simplex (D. simplex) and 2) Green algae (Chlorophyta); Enteromorpha compressa (E. compressa); Sargassum muticum (S. muticum). These plant samples were collected from the same above-mentioned location from where water samples were collected. The collected plant samples were washed to remove surface adhered materials and stored in plastic bottles. Later, the algal samples were washed twice with double distilled water and used to analyze various attributes.

Analyses of Water Samples
Collected water samples were used for the estimation of following quality parameters: turbidity was measured using nephelometer, and pH using portable pH meter. Total dissolved solids (TDS) were estimated using a conductivity meter. Biochemical oxygen demand (BOD) and chemical oxygen demand (COD) were measured by the volumetric titration method (APHA, 1995) [15]. Total Kjeldahl nitrogen (TKN) was estimated by colorimetric method (EPA, 1978a) [16], ammonia [18] using cadmium reduction method. Fluoride was estimated by SPADNS method adapted from standard methods for the examination of water and wastewater (APHA, 1998) [19]. Total phosphorus (P) and total chlorine residual, and iron (Fe) were analyzed using standard methods as described in method 8048-Hach, 8167-Hach, and 8008-Hach, respectively (APHA, 1995) [15]. Oil and grease contents were quantified using gravimetric method 9070 (Blum and Taras, 1968) [20]. Total organic carbon (TOC) was assessed using EPA method 415.1 (EPA, 1999) [21], while phenols were estimated spectrophotometrically using EPA method 420.1 (EPA, 1978b) [22]. Concentration of following HMs

Determination of Photosynthetic Pigments, Total Protein, and Carbohydrates Content
The method of Lichtenthaler and Buschmann (2001) [23] was used to estimate Chlorophyll (Chl) and total carotenoids content. The optical density of the pigment Values are average ± SE of three independent replicates. TDS: total dissolved solids; BOD: biochemical oxygen demand; COD: chemical oxygen demand.

Determination of Stress Markers
To  [27]. The absorbance of the samples was read at 390 nm and H 2 O 2 content was quantified by comparing with a standard curve and was expressed as μmol•g −1 FW. Content of TBARS was determined according to Cakmak and Horst (1991) [28]. The absorbance of the supernatant was measured at 532 nm and 600 nm. The values were corrected for non-specific turbidity by subtracting the absorbance and the values were expressed as nmol•g −1 FW.

Statistical Analysis
The data were analyzed statistically using SPSS-20 statistical software (SPSS Inc., Chicago, IL, USA). Means of three independent replicates were presented ± SE.
When F value was found to be significant at 5% level of probability, critical difference (CD) was calculated.

Photosynthetic Pigments, and Total Protein and Carbohydrates Content
A perusal of the data show that all the studied algal species showed a significant variation in photosynthetic pigments ( life, these pigments are not only responsible for organic food production but also provide protection against high light intensity and also assist in light absorption and energy transfer to the reaction centre. As shown by the results green algae possess a higher concentration of Chl a, Chl b and total Chl, which resulted in enhanced accumulation of carbohydrates ( Figure 2). However, the concentration of carotenoids, phycocyanin and phycoerythrin were found higher in red algae (Table 4) [36]. In addition, variation in these photosynthetic pigments in the studied plants was probably due to the quality of the water (Tables 1-3

Level of Stress Markers in Red and Green Algae
Exposure of plants to various natural stresses as well as to uncontrolled anthropogenic activities, induces excessive generation of ROS which causes damage to biological membranes and adversely affects several plant physiological processes (Khan et al., 2020 [40]). In the present study, we analyzed H 2 O 2 content and TBARS as stress markers of oxidative stress and lipid peroxidation. Marine

Activities of Antioxidant Enzymes and Proline (Pro) and Glycine Betaine (GB) Content in Red and Green Algae
To cope with detrimental effects of oxidative stress plants are fitted with antioxidant system. The antioxidant enzyme SOD is known as the first line of defense against ROS that dismutase  Figure 4(B)). Therefore, lower level of H 2 O 2 in red algae was due to increased activities of these antioxidant enzymes. Whereas in green algae, SOD activity was higher while POX and CAT were lower (Figure 4(A) and Figure 4(B)) that resulted in more accumulation of H 2 O 2 than the red algae. Increase in the activities  The results of the study show that red algae synthesized more Pro and GB than the green algae and highest level of proline was found in red alga D. simplex, whereas lowest value was found in green alga E. compressa ( Figure 5(A) and

Conclusion
The presence of contaminants in water bodies is a menace for the aquatic ecosystem. The present study exhibits that the response of red and green algae to  the contaminants present in water of the Red Sea along the Haql coast. The results of the study show that the two algal groups tested responded differently.
Regarding pigment concentration, Chl a, b, total Chl content and Chl a:b ratio were higher in green algae, whereas, carotenoids, phycocyanin and phycoerythrin were higher in red seaweeds. Higher activities of antioxidant enzymes, and concentration of protein, carbohydrates and protein in red algae show that they possess a strong defense system against the contaminants present in the marine ecosystem. To put all in a nutshell, it can be concluded that red algae were more tolerant of the contaminants than the green algae and the alga G. salicornia was found best in having a stronger defense system.