International Journal of Fisheries and Aquaculture Volume 7 Number 5 May 2015 ISSN 2006-9839

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Prof. Upali S. Amarasinghe Dr. Masoud Hedayatifard Department of Zoology, Department of Fisheries Sciences and Aquaculture University of Kelaniya, College of Agriculture and Natural Resources Kelaniya 11600, Sri Lanka. Advanced Education Center Sri Lanka. Islamic Azad University, Ghaemshahr, PO Box: 163, Iran. Dr. V.S. Chandrasekaran Central Institute of Brackishwater Aquaculture (ICAR) Dr. Zhang Xiaoshuan 75, Santhome High Road, R.A.Puram 209#, China Agricultural University(East campus), Chennai-600028, No.17 Qinghua Donglu, India. Beijing, China

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Dr. Dada Adekunle Ayokanmi Dr. Harikrishnan Department of Fisheries and Aquaculture Faculty of Marine Science Technology, Federal University of College of Ocean Sciences Technology, P.M.B 704, Jeju National University, Jeju, 690-756 Akure, Ondo State, South Korea . Nigeria.

Dr. Ramasamy Harikrishnan Prof. Ratha Braja Kishore KOSEF Post Doctoral Fellow, Faculty of Department of Zoology Marine Science, College of Ocean Sciences, Biochemical Adaptation Laboratory Jeju National University, Banaras Hindu University Jeju city, Jeju 690 756, Varanasi 221005 South Korea. India.

Dr. Kawser Ahmed Dr. Esmaile AM Shakman Lab. of Ecology, Environment and Am Vögenteich,13/ 3.09.618057 Rostock Climate Change, Department of Fisheries, Germany . University of Dhaka, Dhaka-1000, Bangladesh. Prof B. Sharma Department of Biochemistry Dr. Maxwell Barson Coordinator, Centre for Biotechnology Biological Sciences Department University of Allahabad University of Zimbabwe PO Box MP 167 Allahabad-U.P., Mount Pleasant Harare, India. Zimbabwe. Dr. Sebastián Villasante Dr. Christopher Marlowe Arandela Caipang Fisheries Economics and Natural Resources Research Unit Faculty of Biosciences and Aquaculture, Bodø University University of Santiago de Compostela, A Coruña. College, Bodø 8049, Spain. Norway. Dr. Mohamed Hamed Yassien Dr. William S. Davidson National Institute of Oceanography and Fisheries, Suez Department of Molecular Biology and Biochemistry branch, Simon Fraser University 8888 University Drive P.O. Box (182), Suez, Burnaby, British Columbia Egypt. Canada V5A 1S6. Dr. Abhay Bhalchandra Thakur Dr. Babak Ghaednia 2/9 Mai Krupa Sagar Society Opp. Catering College Iran Shrimp Research Center ( ISRC) Veer Savarkar Marg Dadar, Mumbai -400 028 Taleghani High Way, P.O.Box 1374 Maharashtra, Bushehr, India. Iran . Dr. Riaz Ahmad Dr. Ramachandra Bhatta Department of Zoology Animal and Fisheries Sciences University, Aligarh Muslim University College of Fisheries, Aligarh- 202002, (UP) Kankanady Mangalore 575 002 India. Indi a.

International Journal of Fisheries and Aquaculture

Table of Contents: Volume 7 Number 5 May 2015

ARTICLES

Research Articles

Effect of dietary replacement of fishmeal with Chlorella vulgaris on 62 growth performance, energy utilization and digestive enzymes in Macrobrachium rosenbergii postlarvae Radhakrishnan S., Saravana Bhavan P., Seenivasan C. and Muralisankar T.

Photosynthetic productivity and biomass of phytoplankton, in Lake 71 Kuriftu, Ormia Region, Ethiopia Zelalem Dessalegn Fayissa

Vol. 7(5), pp. 62-70, May 2015 DOI: 10.5897/IJFA15.0471 Article Number: 736B4E353206 International Journal of Fisheries and ISSN 2006-9839 Copyright ©2015 Aquaculture Author(s) retain the copyright of this article http://www.academicjournals.org/IJFA

Full Length Research Paper

Effect of dietary replacement of fishmeal with Chlorella vulgaris on growth performance, energy utilization and digestive enzymes in Macrobrachium rosenbergii postlarvae

Radhakrishnan S.*, Saravana Bhavan P., Seenivasan C. and Muralisankar T.

Crustacean Biology Laboratory, Department of Zoology, Bharathiar University, Coimbatore – 641046. Tamilnadu, India.

Received 3 January, 2015; Accepted 21 April, 2015

The present study was conducted to assess the growth promoting ability of Chlorella vulgaris on Marobrachium rosenbergii postlarvae. The experimental diets were prepared by fishmeal replacement with C. vulgaris at different levels (25, 50, 75 and 100%). The prepared feeds were offered to the M. rosenbergii postlarvae during 90 days in triplicate. At the end of the feeding experiment, survival rate, weight gain, specific growth rate and feed conversion efficiency were significantly (P < 0.05) higher in 50% C. vulgaris inclusion diet fed PL group. Similarly, total protein, amino acid, carbohydrate and lipid contents were significantly (P < 0.05) higher in specimens fed 50% C. vulgaris supplemented diet. The feeding rate, absorption rate, conversion rate were significantly (P < 0.05) higher in 50% C. vulgaris supplemented diet fed PL group. At this level of replacement, the activity level of digestive protease, amylase and lipase were significantly (P < 0.05) higher in 50% C. vulgaris supplemented group. Among all the experimental groups, the 50% fishmeal replacement with C. vulgaris inclusion diet fed group, showed significant performance. The present results revealed that the partial replacement of fish meal with C. vulgaris is favorable for M. rosenbergii postlarval culture.

Key words: C. vulgaris, digestive enzymes, growth performance, M. rosenbergii.

INTRODUCTION

The culture of freshwater prawn, Macrobrachium predaceous species of fish, and it has a shorter larval rosenbergii (Scampi), has received a great deal of period. The M. rosenbergii breeding, larval culture and attention in India as a preferred crustacean. Under export is one of the important industries in south Indian controlled culture in freshwater and low saline ponds in states of Kerala, Andhra Pradesh and Tamilnadu. In inland, as well as coastal areas, it grows fastest among recent years, the aquaculture industry has succeeded in all freshwater prawns. It shows a wide range of reducing the inclusion rates of ﬁshmeal and ﬁsh oil in the temperature and salinity tolerance, acceptance of a large aquafeeds. However, due to the increase in production of range of formulated diets, culture compatibility with non- all farmed species there is still a growing demand for

*Corresponding author. E-mail: [email protected] Author(s) agree that this article remain permanently open access under the terms of the Creative Commons Attribution License 4.0 International License Radhakrishnan et al. 63

these ingredients (Naylor et al. 2009). Fishmeal is the carbohydrate, lipid, essential amino acids and minerals principal source of protein in commercial aquafeeds. As a (Dawah et al., 2002; Tokusoglu and Unal, 2003; Janczyk et result of the steep increase in price of fishmeal and the al., 2005; Vaikosen et al., 2007). In this study, the effect of decline in fishery resources that goes in to fishmeal fishmeal dietary replacement with C. vulgaris on growth production, there is an interest in developing alternatives performance, energy utilization, carcass composition and to this finite component. Finding and testing alternate digestive enzyme activity in freshwater prawn M. rosenbergii protein and lipid sources is important to the aquatic feed postlarvae (PL) was evaluated. industry (Kiron et al. 2012). As microalgae protein is of good quality, with amino acid profiles comparable to that of other reference feed MATERIALS AND METHODS proteins, it could be a plausible alternative to fishmeal Culture of Chlorella vulgaris protein (Becker 2007). In addition, microalgae, which are the source of all photosynthetic ally fixed carbon in the Collection of pure mother culture of C. vulgaris: C. vulgaris food web of aquatic animals (Kwak and Zedler 1997), mother culture were collected from Vivekananda Institute of Algal may be an ideal replacement for fishmeal in aquatic Technology (VIAT), R.K.M. Vivekananda College, Chennai, Tamil feeds. Meal from the cyanobacterium Spirulina, a Nadu, India. brackishwater genus that is neither a eukaryote nor Preparation of inoculums: The microalgae, C. vulgaris, was marine, has been incorporated into experimental fish inoculated in Bold Basal medium (100 ml mother culture + 900 ml feeds with little success (El-Sayed 1994; Olvera-Novoa et basal medium – as used by Bischoff and Bold (1963) and Schuster al., 1998; Nandeesha et al., 2001; Palmegiano et al. et al. (1990), and the cultures were incubated for 15 days at 24 ± 2005). 1°C in a thermo-statically controlled room and illuminated with cool Protein and vitamin content is a major factor inflorescence lamps (Phillips 40 W, cool daylight 6500 K) at an intensity of 2000 lux in a 12: 12 h light dark regime. determining the nutritional value of microalgae. In addition, polyunsaturated fatty acids (PUFA) e.g. Culture in glass tanks: Culture containers were well cleaned with eicosapentaenoic acid (EPA), arachidonic acid and bleach, rinsed and sun dried for 8 h. Then plastic troughs were filled docosahexaenoic acid (DHA) content is of major with tap water up to 25 L and mixed well with the pure nutrient importance (Catarina et al. 2010). The most frequently media (N-8 medium) (Vonshak1986). 1 L of mother culture of C. used genera are Chlorella, Tetraselmis, Isochrysis, vulgaris was inoculated in the glass tanks. The tanks were vigorously aerated to provide required quantity of oxygen and Pavlova, Phaeodactylum, Chaetoceros, Nannochloropsis, to keep cells and media in suspension. The required Skeletonema and Thalassiosira. Combination of different concentration of algae was developed after 30 days of algal species provides better balanced nutrition and inoculation. The tanks were kept open under 100% outdoor light improves animal growth better than a diet composed of exposures. A constant temperature of 25-30°C was maintained only one algal specie (Spolaore et al. 2006). throughout the growth period.

The main applications of microalgae for aquaculture are Counting of algal cells and filtering method: Sampling was done associated with nutrition or as food additive for coloring once in five days basis using 10 ml capacity vials. Chlorella cells in the ﬂesh of salmonids and for inducing other biological each vial were preserved by adding 2-3 drops of formalin. 1 ml of activities. Microalgae are required for larval nutrition sample was carefully filled in Neubauer Hemocytometer groove during a brief period, either for direct consumption in the (Bauer, 1990) and covered with glass slide. The cells were case of molluscs and peneaid shrimp or indirectly as feed enumerated under compound microscope. Hand tally counter was used for reliable counting. Algal cells were calculated by the for the live prey fed to small fish larvae (Muller-Feuga, following mathematical expression: 2000). In order to be used in aquaculture, a microalgal strain has to meet various criteria, such as ease of culturing, Cells (ml -1) = Total number of cells counted/10×4×10-6 lack of toxicity, high nutritional value with correct cell size and shape and a digestible cell wall to make nutrients Filtering method: Printing polyester/ Nylon fabric cloth with a mesh between 30-60 microns was used as filter. After use, the filter was available (Patil et al. 2007). carefully washed, as quickly as possible, and then dried away from Chlorella vulgaris is unicellular green algae found in direct sunlight. both fresh and marine water and it is widely used as food supplements (Kay 1991). Signiﬁcant attention has recently been drawn to the use of microalgae for Experimental feeds developing functional food, as microalgae produce a The processed feed grade fishmeal was purchased from Rosen great variety of nutrients that are essential for human fisheries (Marathakkara, Thrissur, Kerala, India). The other basal health. The nutritional value of C. vulgaris was initially feed ingredients such as, soybean meal, groundnut oil cake, wheat determined in 1950s-1960s (Lubitz 1963) and first studied bran, binders (Egg and tapioca flour), sunflower oil and vitamin as a possible food source in Japan, United States and capsules were purchased from local markets at Coimbatore Germany after World War II (Miyachi, 1995; Ashraf et al., (Tamilnadu, India). The replacement materials C. vulgaris was cultured in a laboratory as directed by Vonshak (1986) and 2011). The previous researches suggested that C. Schuster et al. (1990). vulgaris contain supreme level of crude protein, The basal ingredients such as processed fishmeal and sundried 64 Int. J. Fish. Aquac.

Table 1. Ingredients and proximate composition of experimental diets.

Ingredients (g kg-1) Control CL-25 CL-50 CL-75 CL-100 Fishmeal 250 187.5 125 62.5 0 Groundnut oil cake 250 250 250 250 250 Soybean meal 250 250 250 250 250 Wheat bran 100 100 100 100 100 Egg albumin 70 70 70 70 70 Tapioca flour 50 50 50 50 50 Sunflower oil 20 20 20 20 20 Vitamin mix* 10 10 10 10 10 C. vulgaris 0 62.5 125 187.5 250

Proximate composition of C. vulgaris incorporated feed (g kg-1) Crude protein 420.20 414.70 409.30 403.80 398.40 Carbohydrate 204.80 206.80 208.80 210.50 212.70 Lipid (%) 147.00 136.30 134.10 132.40 131.30 Ash (%) 111.86 123.30 130.00 137.30 142.00 Moisture (%) 99.30 96.00 93.30 90.46 87.00 Gross energy (kcal kg-1) 3187.71 2930.13 2713.09 2651.96 2379.96

Chemical composition of C. vulgaris: Crude protein- 55.70%; Carbohydrate- 15.28%: Lipid- 10.65%; Ash- 9.00%; Moisture- 6.30%.*Becosules capsules (Each capsule contains);Thiamine mononitrat (IP): 10 mg; Riboflavin (IP): 10 mg; Pyridoxine hydrochloride (IP): 3 mg Vitamin B12 (as tablets 1:100) (IP):15 mcg; Niacinamide (IP): 100 mg; Calcium pantothenate (IP): 50 mg Folic acid (IP): 1.5 mg; Biotin USP (IP):100 mcg; Ascorbic acid (IP):150 mg.

soybean meal, groundnut oil cake and wheat bran were ground 2.90 mm and 2.20 ± 0.39 g were taken for the feeding experiment. separately and filtered with a 60-mesh sieve. The sieved feed Feeding trial was carried out in experimental aquarium (5 x 3 = 15) ingredients were blended with manual blender and the blends were containing 40 L of water, the experimental prawn PLs were used for preparation of experimental diets, one control diet without introduced in the aquarium (30 PLx15 aquarium = 450 PL), and fed microalgae and four diets with C. vulgaris at the concentration of experimental diets for 90 days. The feeding was adjusted to two 25% (CL-25), 50% (CL-50), 75% (CL-75) and 100% (CL-100) of times a day (6:00 am and 6:00 pm) at the ration of 10% of the body fishmeal replacement. The blends were steam cooked for 15 min at weight of PL. The feeding experiment was prolonged for 90 days; 95-100°C and allowed to cool at room temperature. The steam mild aeration was given continuously in order to maintain the cooked blends was mixed with replacement material (C. vulgaris at oxygen level. In each experimental group, 10 prawns sacrificed for the respective concentrations), vitamin tablets, sunflower oil and growth parameter and energy utilization parameters (10 PLx3 =30 binder (egg albumen and tapioca flour). The blends again blended PL/group x 5= 150 PL); 5 PL was sacrificed for proximate with manual blender and added boiled water to prepare a dough composition analysis (5 PLx3=15 PL/group x 5= 75 PL); 5 PL was form. The dough were pelletized with a manual pelletizer fixed with sacrificed for digestive enzyme analysis (5 PLx3=15 PL/group x 5= 3 mm die and the pellets were collected in aluminum trays. Then 75 PL). Therefore, totally 300 PLs (150+75+75 = 300 PL) were the feeds were dried in thermostatic oven until the moisture content sacrificed for these parameters. is less than 10%. The dried pellets were physically examined for visual appearance, uniformity, colour and fragrance. The selected ingredients and feed proximate composition are shown in Table 1. Analyses of the proximate composition of the experimental diets

Animals Analysis of crude protein, moisture, lipid and ash in the formulated

The postlarvae PL of M. rosenbergii (PL 15) were purchased from feed was performed according to AOAC (1995) procedures. The Government prawn hatchery (Azhikkode, Thrissur, Kerala, India). crude protein was analyzed in the Kjeldahl apparatus after acid They were safely brought to the laboratory in well-oxygenated plastic digestion (Pelican equipment; KELPLUS- KES 04 LR with bags. They were stocked in a large cement tank (6’×4’×3’) and automatic digestion system); total lipid was extracted with acclimatized for 2 weeks under laboratory conditions. During the petroleum ether by the Soxhlet method after acid hydrolysis; ash period of acclimatization they were fed with boiled egg albumin (egg was determined using Muffle furnace and moisture was determined custard), Artemia nauplii and crumble feed alternatively twice a day. by thermostatic oven drying method. The formulated feed gross The unused feed and fecal matters were removed daily by the energy was determined using the Oxygen Bomb Calorimeter (230 syphoning method in the acclimatized tank, three fourth of the water VAC; Sl. No. 26036; Advance Research Instrument Company, New was renewed daily and adequately aerated. Delhi, India).

Feeding experiment Determination of growth parameters

M. rosenbergii (PL-30) with the length and weight range of 15.60 ± After the 90 days feeding trial, the growth parameters such as Radhakrishnan et al. 65

survival rate, weight gain, specific growth rate, feed conversion rate, per tank (fifteen per treatment) were randomly sampled and killed condition factor and feed conversion efficiency were individually with a sharp blow in the head. The digestive tract was excised and determined by the following equations (Tekinay and Davies 2001): it was checked that there was no food in any portion. The isolated digestive sections were immediately frozen in liquid nitrogen, and stored at -80°C. Each part was homogenized in nine volumes of ice-cold 100 mM Tris-HCl buffer containing 0.1 mM EDTA and 0.1% (v/v) Triton X-100, pH 7.8. All procedures were performed on ice. Homogenates were centrifuged at 30,000 g for 30 m at 4°C and the resultant supernatants were kept in aliquots and stored at -80°C for further digestive enzyme assays. Total protease activity was determined by the casein-hydrolysis method described by Furne et al. 2005. The assay buffer consisted of 0.1 M glycine-NaOH (pH 10.0). The reaction mixture consisted of casein at 1% (w/v) (0.25 mL), buffer (0.25 mL) and supernatant from the homogenates (0.1 mL). This reaction mixture was incubated for1 h at 37°C. The reaction was stopped by adding 0.6 mL 8% (w/v) trichloroacetic acid solution and kept for 1 h at 2°C

then centrifuged at 1800 g for 10 m and the supernatant absorbance was measured at 280 nm against blanks. For the blank preparation, the supernatant from the homogenates was added at the end of the incubation period, just before adding trichloroacetic

acid. Tyrosine solution was used as standard. One unit of enzyme

activity was defined as the amount of enzyme needed to catalyze Energy utilization the formation of 1.0 μmol of tyrosine per min.

Amylase activity was determined by the starch-hydrolysis method The parameters of energy utilization, such as feeding rate, mean of Bernfeld (1955). The reaction mixture consisted of 2% (w/v) absorption, mean conversion and metabolic rate were calculated. starch solution (0.125 mL), 0.1 M citrate–phosphate buffer at pH 7.5 The energy content of whole prawn, feeds, faeces and exuvia were (0.125 mL) and supernatant from the homogenates (0.5 mL). This calculated by the Oxygen Bomb Calorimetrically derived heat was incubated for 1 h at 37°C. The absorbance was measured at energy value. The scheme of energy budget followed in the 600nm against a blank. For the blank, the supernatant from the present study is that of IBP formula (Petrusewicz and Macfadyen homogenates was added just after the incubation period. Maltose 1970) represented as C = P + R + U + F, where C is the energy solution was used as standard. One unit of amylase activity was consumed, P the growth, R the energy lost as heat due to defined as the amount of enzyme that produced 1.0 μmol of metabolism, F the faeces and U the nitrogenous waste. The daily maltose per min. excretion of ammonia by the prawn was estimated after feeding as Lipase activity was determined following the method of Furne et per the phenol hypochloride method of Solorzano (1969). The al. (2005) by degrading triacylglycerol to free fatty acids. A solution energy loss due to ammonia excretion was calculated using the of 1% polyvinyl alcohol (PVA) and 5 mL of 0.1 N HCl in 1 L of ammonia calorific quotient, 1 mg NH : 5.9 cal. (Elliot 1976). 3 distilled water was heated to 75-85°C, cooled, filtered and adjusted

to pH 8.0 with 0.1 N NaOH. Virgin olive oil was added to an aliquot of the previous solution obtaining a substrate concentration of 0.1 M. This mixture was emulsified for 5 m. In addition, McIlvaine buffer was prepared from 0.1 M citric acid and 0.2 M disodium phosphate. A reaction mixture containing PVA solution-emulsified substrate (1 mL), Mcllvaine buffer at pH 8.0 (0.5 mL), and supernatant from the homogenates (0.5 mL) was incubated for 4 h at 37°C. Afterwards, 3 ml of 1:1 ethanol-acetone solution was added to stop the reaction and break the emulsion. Phenolphthalein in ethanol 1 % (w/v) was added to the reaction mixture and titrated with 0.01 M NaOH. For the blanks, the same procedure was followed but boiled

enzyme was used. One unit of lipase activity was defined as the hydrolysis of 1.0 micro equivalent of fatty acids from triacylglycerols in 1 h at pH 8 and 37°C. Specific activities were expressed in unit per mg of soluble protein (U/mg protein).

Corporal chemical composition of postlarvae

At the end of the experimental period, the experimental prawns were weighed and sacrificed for corporal chemical composition (Where AR= Absorption rate; CR= Conversion rate; ER= Excretion analyses. Five animals per tank (fifteen per treatment) were rate) randomly sampled and killed with a sharp blow in the head. The animals head, alimentary track and exoskeleton were removed. The isolated muscle portions were labeled and kept in -20°C for further Digestive enzymes analyses. Concentration of total protein in enzymatic extracts and At the end of the experimental period, the experimental prawns postlarvae carcasses was estimated by the method of Lowry et al. weighed with no signiﬁcant differences between groups. Prawn (1951) using ethanolic precipitated sample. The blue colour was a feeding was discontinued 24 h before sampling and five animals result of biuret reaction of protein with copper ions in alkali solution 66 Int. J. Fish. Aquac.

and reduction of the phophomolybdic-phosphotungstic of Folin protein sources such as soybean protein for reagent by the tyrosine and tryptophan present in the treated Oncorhyncus mykiss (Kaushik et al., 1995); plant protein protein. This colour intensity was measured at 650 nm against a based diets for tilapia (Goda et al. 2007) and poultry by- blank devoid of protein sample. Bovine serum albumen (BSA) was used as a standard. The content of amino acid was estimated by products for Oreochromis niloticus (Hernandez et al. the method of Moore and Stein (1948). The total amino acid was 2010). extracted with sodium tungstate and H2SO4. When amino acids are In order for protein sources to be considered effective heated with ninhydrine, they undergo deamination. The reaction of replacements for FM, they must be economically amino acid-hydratin complex with ninhydrin produced purple colour, competitive, capable of being produced in large quantities which was measured at 540 nm against a blank. Leucine was used as standard. (Hardy et al. 2002), contain balanced amino acid profiles The carbohydrate was estimated by the method of Roe (1955) and proper crude protein levels, and not compromise the using TCA extracted sample. Carbohydrates were hydrolysed into growth or health of the fish. It is also helpful if they are simple sugars by diluted HCl in hot acidic medium. Glucose is easily handled and stored and do not lead to dehydrated into hydroxyl-methyl furfural. This compound reacts with environmental contamination from release of anthrone and produced green colored product, which was phosphorous and nitrogen. Furthermore, such protein measured at 630 nm against a blank. Glucose was used as standard. sources must be commodity traded (Lunger et al. 2006). Total lipid was extracted with chloroform-methanol mixture Also, the present study represented that the alternative following the method of Barnes and Black-Stock (1973), and protein source is well utilized by the prawn PL, because estimated by the method of Folch et al. (1957). Lipid reacts with the experimental diets fed groups showed significant vanillin in a medium of H2SO4 and phosphoric acid to form a pink improvement in length, weight gain, survival, specific coloured chromogen, which is proportional to the lipid content of the growth rate and feed conversion efficiency. In this feeds sample, which was measured at 540 nm against a blank. Olive oil was used as standard. the 50% fishmeal replaced by C. vulgaris feed fed group showed better performance in growth parameters. These results are in agreement with those obtained by Dawah et Statistical analysis al. (2002) who found that the addition of algae in fish diets improved growth performance of Nile tilapia (O. niloticus). The results were expressed as Mean ± SD. Statistical analysis was carried out by Analysis of Variance (one way ANOVA) followed by Also, Zeinhom (2004) observed that inclusion of algae in DMRT were considered as indicative of significance level of P < fish diets significantly improve the live body weight and 0.05, as compared to the control group. All calculations were SGR. The fresh biomass of Chlorella sp., Tetraselmis sp., performed using: SPSS, version 16.0 for Windows (PSS Inc., 444 Isochrysis sp., Synechococcus sp. and Phormidium sp. N. Michigan Ave., Suite 3000, Chicago, IL 60611). were used as feed for shrimp Penaeus monodon and it enhanced the growth, weight gain and survival rate. Similarly, microalgae (Chlorella sp., Tetraselmis sp., RESULTS AND DISCUSSION Isochrysis sp,) and cyanobacteria (Synechococcus sp.,

Growth, nutritional indices and energy utilization and Phormidium sp.,) live feed fed shrimp had improve the growth, survival and body carcass composition Growth performance, nutritional and energy utilization of (Sivakumar et al. 2011); live C. vulgaris fed Moina M. rosenbergii PL fed experimental diets are presented in micrura had significant growth and survival (Habib et al., Table 2. The initial average body length and weight of PL 2003) and partial replacement of fish meal with dried was 15.6 ± 2.9 mm and 2.2 ± 0.39 g respectively, in all microalgae Chlorella sp and Scenedesmus sp. feed fed the groups. At end of the feeding experiment, the final O. niloticus gain significant improvement in survival, length and weight were significantly (P < 0.05) improved growth and body proximate composition (Tartiel et al. in CL-50 group, followed by the CL-75 and CL-25 groups 2008). Nandeesha et al. (1998) reported that body weight when compared with control treatment. Similarly, the gain of Nile tilapia (O. niloticus) increased linearly with growth parameters and nutritional indices (SR, WG, SGR increasing the level of algae in fish diet at levels less than and FCE) were found to be maximum in specimens fed 20%. on 50% C. vulgaris. The same experiment group (CL-50) In the present study, the feeding rate of C. vulgaris showed a significance increase in feeding, absorption supplemented groups was comparatively higher than in and conversion rates, ammonia excretion and metabolic the control group. The CL-50 showed higher feeding rate, rates. The groups fed on CL-100 showed no significant but the level was slightly decreasing in CL-75 and CL-100 difference when compared with the control group. groups. Part of the food ingested is assimilated in the gut Ferouz et al. (2012) reported in his review study that and the remaining fraction is eliminated as faeces. It is alternative protein sources such as cereals, pulses, oil indicated that the C. vulgaris protein was easily absorbed seeds and some of the animal proteins were present in and assimilated by prawn PL, so the fecal output is lower. aquaculture species. Still, the alternative protein study In this experiment, the groups fed on C. vulgaris needed improvement in proximate composition and anti- supplemented diets showed better feed conversion rate nutritional factors analyses. Previous studies have shown and growth performance. The present study revealed that that fishmeal can be effectively replaced by alternative the experimental feed was well utilized by the PL, and the Radhakrishnan et al. 67

Table 2. Growth, nutritional indices and energy utilization parameters of M. rosenbergii PL fed with experimental diets.

Tests Parameters Control CL-25 CL-50 CL-75 CL-100 F value Initial 15.60 ± 2.90 15.60 ± 2.90 15.60 ± 2.90 15.60 ± 2.90 15.60 ± 2.90 - Length (mm) c b a b c Final 44.20 ± 3.00 50.20 ± 1.60 58.00 ± 2.90 51.60 ± 1.50 43.00 ± 3.10 39.720

Morphometric data Initial 2.20 ± 0.39 2.20 ± 0.39 2.20 ± 0.39 2.20 ± 0.39 2.20 ± 0.39 - Weight (g) Final 16.50 ± 1.00c 21.60 ± 1.60b 28.80 ± 1.50a 23.70 ± 2.40b 16.10± 1.00c 61.091

Survival rate (%) SR 76.66 ± 3.5c 86.66 ± 2.5b 93.33 ± 2.50a 80.00 ± 3.00c 80.00 ± 4.00c 13.399 Weight gain (g) WG 16.10 ± 1.00c 19.40 ± 1.20b 27.30 ± 1.70a 21.50 ± 1.50b 13.90 ± 0.70c 48.125 Specific growth rate (g day -1) SGR 0.997 ± 0.05d 1.102 ± 0.07c 1.252 ± 0.04a 1.146 ± 0.06b 0.960 ± 0.08e 146.54 Condition factor 1.91 ± 0.21a 1.70 ± 0.2ab 1.51 ± 0.23b 1.72 ± 0.13ab 2.02 ± 0.14a 3.339 Nutritional indices Feed conversion rate (%) FCR 1.65 ± 0.19a 1.50 ± 0.12ab 1.21 ± 0.16c 1.25 ± 0.14bc 1.73 ± 0.10a 7.694 Feed conversion efficiency (%) FCE 1.98 ± 0.23b 1.99 ± 0.18b 2.54 ± 0.22a 2.41 ± 0.19a 1.66 ± 0.21b 8.915

Feeding rate 1.65 ± 0.12c 2.20 ± 0.90a 2.31 ± 0.04a 2.16 ± 0.09a 1.78 ± 0.07b 38.91

Absorption rate 1.36 ± 0.07c 1.67 ± 0.06b 1.95 ± 0.09a 1.54 ± 0.10c 1.39 ± 0.50d 42.02

Conversion rate 0.92 ± 0.09c 1.20 ± 0.11b 1.45 ± 0.13a 1.07 ± 0.09bc 0.90 ± 0.10cd 16.89 Energy utilization NH Excretion 0.07 ± 0.01b 0.11 ± 0.01ab 0.12 ± 0.01a 0.10 ± 0.01b 0.08 ± 0.01c 16.80 (kJ day -1) 3 Metabolic rate 0.51 ± 0.03c 0.58 ± 0.02a 0.61 ± 0.06a 0.56 ± 0.02a 0.53 ± 0.02a 0.061

Each value is a mean ± SD of three replicate analysis, within each row means with different superscripts letters are statistically significant P < 0.05 (one way ANOVA and subsequently post hoc multiple comparison with DMRT.

feeds contained nutrients which are easily Corporal chemical composition fed Labeo rohita, gained significant improvement absorbed and converted to energy, because it is of survival and body carcass composition. Also, ultimately producing the significant growth and Total protein, amino acid, carbohydrate and lipid Sivakumar et al. (2011) reported that live three body carcass composition in experimental diet fed content of M. rosenbergii PL was affected by micro algal strains (Chlorella sp., Tetraselmis sp., group. A balanced energy budget is a tool for partial replacement of fishmeal C. vulgaris. The and Isochrysis sp) two cyanobacterial bioenergetics modeling in aquaculture and chemical contents were significantly (P < 0.05) (Synechococcus sp. and Phormidium sp) strains fisheries management. In contrast, for crustacean increased in specimens fed on 50% of inclusion fed P. monodan had significant improvement in species, notably penaeid shrimp information is level, followed by groups fed on CL-25 and CL- growth, survival and body composition. Gouveia limited (Bureau et al., 2000). Similarly, the balanced 75% when compared with the control group. The et al. (1998) suggested that the dry C. vulgaris energy budget study was reported in various studies 100% C. vulgaris inclusion feed fed group showed biomass is an effective and digestible source of of M. rosenbergii PL fed with cereals, pulses based no significant difference when compared with the carotenoid pigments. feed (Bhavan et al. 2010); probiotic incorporated control (Table 3). The present results indicate the C. vulgaris protein was well utilized by the PL of feed (Seenivasan et al., 2012; 2013) and medicinal Digestive enzymes activity herbs incorporated feed (Shanthi et al., 2012; M. rosenbergii. Similarly, Bakhtiyar et al. (2011) Radhakrishnan et al., 2014a, b). reported that Chlorella bioenriched zooplanktons The protease, amylase and lipase activity levels in 68 Int. J. Fish. Aquac.

Table 3. Body chemical composition and activities of digestive enzymes of M. rosenbergii PL fed with experimental diet.

Assays Parameters Control CL-25 CL-50 CL-75 CL-100 F value Protein 563.00 ± 15.80c 640.00 ± 21.00b 698.00 ± 19.50a 636.00 ± 12.60b 552.20 ± 18.60c 34.72 Biochemical d b a c e Amino acid 350.00 ± 11.70 400.60 ± 5.20 438.30 ± 5.80 370.20 ± 6.80 272.30 ± 11.10 155.917 constituents c ab a bc c -1 Carbohydrate 224.10 ± 13.60 258.90 ± 15.70 282.40 ± 17.80 238.70 ± 17.80 213.50 ± 12.30 11.38 dry basis (g kg ) Lipid 174.20 ± 19.80b 179.60 ± 4.10b 212.50 ± 6.40a 190.10 ± 13.40b 140.90 ± 6.10d 15.16

Ash (%) 11.90 ± 0.28b 12.12 ± 0.26b 13.15 ± 0.25a 12.12 ± 0.23b 12.03 ± 0.15b 13.390

Moisture (%) 74.45 ± 1.10ab 73.26 ± 1.15a 72.41 ± 1.50b 73.11 ± 1.65ab 75.2 ± 1.20a 2.103

Initial 0.39 ± 0.09 0.39 ± 0.09 0.39 ± 0.09 0.39 ± 0.09 0.39 ± 0.09 - Protease Final 1.16 ± 0.08bc 1.26 ± 0.07ab 1.31 ± 0.02a 1.21 ± 0.07abc 1.10 ± 0.08c 3.815

Initial 0.27 ± 0.12 0.27 ± 0.12 0.27 ± 0.12 0.27 ± 0.12 0.27 ± 0.12 - Digestive enzyme Amylase Final 0.81 ± 0.05c 0.94 ± 0.04b 1.04 ± 0.03a 0.85 ± 0.06bc 0.78 ± 0.08c 12.389

Initial 0.28 ± 0.07 0.28 ± 0.07 0.28 ± 0.07 0.28 ± 0.07 0.28 ± 0.07 - Lipase Final 0.76 ± 0.05b 0.75 ± 0.06b 0.86 ± 0.03a 0.80 ± 0.04ab 0.61 ± 0.03c 13.195

Values are represented for protease and amylase in Unit mg protein-1; lipase represented in Unit ×102. Each value is a mean ± SD of three replicate analysis, within each row means with different superscripts letters are statistically significant. P < 0.05 (one way ANOVA and subsequently post hoc multiple comparison with DMRT).

enzymatic extracts were assayed in the initial and the protein digestibility in common carp C. carpio. fishmeal-free shrimp feed in a pond trial (Amaya final day of the study (Table 3). The protease Nandeesha et al. (1994) reported that 25, 50, 75 et al. 2007b). Furthermore, beneficial impact of enzyme activity level was significantly higher in and 100% level of fishmeal replacement with S. algal inclusion on shrimp health has been animals of CL-50 fed group, followed by CL- 25 and platensis significantly increased the digestibility in reported recently. L. vannamei fed diets CL-75 groups, but the PL fed on 100% C. vulgaris catla, rohu and common carp mixed culture. supplemented with marine algal meals rich in diet showed no significance when compared to Mustafa and Nagakawa (1995) suggested that the docosahexaenoic acid and arachidonic acid control group. The amylase and lipase activities dietary inclusion of algae contribute to an increase demonstrated significant improvement in immune showed the same tendency with that of protease in protein assimilation and feed utilization. responses (Nonwachai et al. 2010). activity. The statistical analysis revealed that the There is hardly any information on the use of The present study exhibits, dried C. vulgaris can level of digestive enzymatic activities between microalgae as a dry feed component for shrimps, be used as a partial replacement of fishmeal control and experimental feed prawns and though there are ongoing efforts to replace protein in aquafeeds. It was revealed that, the microalgae supplemented groups were statistically fishmeal protein using terrestrial plant proteins. L. growth of M. rosenbergii fed C. vulgaris meal up significant (P < 0.05). vannamei has been successfully grown on a to 50% level increased significantly. At 75% and Similarly, Nandeesha et al. (1998) reported that predominantly plant protein diet containing 100% inclusion level the growth and nutritional Spirulina inclusion diets fed C. carpio had solvent-extracted soybean meal, corn gluten meal utilization was slightly reduced. Hence, the result significantly increased hepatopancreas protease, and corn fermented soluble, which together of this study exposed that the partial replacement amylase and lipase activity when compared to accounted for nearly 98% of the total dietary of fishmeal with C. vulgaris could be beneficial for control. Also, Umesh et al. (1994) reported that 50% protein of 36% (Amaya et al., 2007a). The same nursery maintenance in M. rosenbergii postlarvae of S. platens dietary inclusion significantly improved research group has verified the concept to culture. Radhakrishnan et al. 69

Conflict of Interest Domenzain A, Domezain J, Sanz A (2005). Digestive enzyme activities in Adriatic sturgeon Acipenser naccarii and rainbow trout Oncorhynchus mykiss. A comparative study. Aquacult. 250:391-398 The authors have not declared any conflict of interest. Goda AMAS, Wafa MA, El-Haroun ER, Chowdhary MAK (2007). Growth performance and feed utilization of Nile tilapia Oreochromis niloticus (Linnaeus, 1758) fingerlings and tilapia galilae, ACKNOWLEDGEMENTS Sarotherodon galilaeus (Linnaeus, 1758) fingerlings fed plant protein- based diets. Aquacult. Res. 38:827-837.

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Vol. 6(5), pp. 71-80, May 2015 DOI: 10.5897/IJFA14.0462 Article Number: 9F94B8E53208 International Journal of Fisheries and ISSN 2006-9839 Copyright ©2015 Aquaculture Author(s) retain the copyright of this article http://www.academicjournals.org/IJFA

Full Length Research Paper

Photosynthetic productivity and biomass of phytoplankton, in Lake Kuriftu, Ormia Region, Ethiopia

Zelalem Dessalegn Fayissa

Department of Biology, School of Natural Science, Adama Science and Technology University, Ethiopia.

Received 17 November, 2014; Accepted 9 March, 2015

Photosynthetic productivity of phytoplankton and chemicals of the water column in Lake Kuriftu were studied from August 2005 to April 2006. The light-saturated rate of photosynthesis (Amax), which was -3 -1 positively and strongly correlated with phytoplankton biomass, ranged from 571 to 1136 mg O2 m h . Biomass-specific rate of photosynthetic productivity at light saturation (Pmax) ranged from 18.78 to 33 -1 -1 mg O2 (mg Chl a) h , while the hourly integral photosynthetic production (∑A) which was positively -2 -1 and strongly correlated with biomass and Amax, varied between 0.686 and 1.05 g O2 m h . The factors responsible for the observed spatio-temporal variations in the physical, chemical and biological features of the lake are discussed.

Key words: Photosynthetic productivity, phytoplankton, oxygen.

INTRODUCTION

In order to meet the growing food requirements of the 1981) reported similar values for the phytoplankton of world population, great effort is needed in the Lake Simbi (Kenya). Exceptionally high algal -2 -1 development and utilization of the biological wealth of the photosynthetic rate in the order of 43 to 57 g O2 m d aquatic environment. The study of energy transfer in (Ca. 16 to 21 g C m-2 d-1), has also been reported by lakes and reservoirs is based on the measurement of Talling et al. (1973) for Lake Arenguade, an Ethiopian primary productivity of phytoplankton and the soda lake whose phytoplankton community was environmental variables, which limit or control this dominated by Spirulina platensis. productivity. Primary productivity of aquatic ecosystems Primary productivity and biomass of phytoplankton are is basically dependent upon the photosynthetic activity of affected by an array of chemical, physical, and biological autotrophic organisms (Wetzel and Likens, 1979). factors. There is no doubt that in general, the more Phytoplanktons are the major primary producers in many frequently a lake is stirred by winds to the bottom, the aquatic systems and are an important food for consumers faster the nutrients are recycled from the mud into the (Reynolds, 1984). photosynthetic zone where they may accelerate the rate Melack and Kilham (1974) suggested that in lakes not of productivity (Talling and Lemoalle, 1998). enriched by human activities, gross photosynthetic rates The dynamics of phytoplankton standing stock and -2 -1 -2 -1 of 30 g O2 m d (ca. 11 g C m d ) or greater are productivity in African lakes was reported to vary seldom encountered. More recently, Melack (1979a, intimately with the fluctuation in water level (Lemoalle,

1975; Melack 1976). Melack (1976) found positive meter (Hanna 9024) within a few hours of sample collection and correlation between level of primary productivity and carbonate-bicarbonate alkalinity was determined by titration with changes in water level. Algal cells require elements in HCI to a pH of 4.5. relatively fixed proportions in order to reproduce (Hecky and Kilham, 1988). The various nutrients required by In situ measurement of primary productivity algal cells for growth and multiplication may not always be found in relative proportions required by phytoplankton Primary productivity was measured by the Light and Dark bottle (Hecky and Kilham, 1988). Phosphorous and nitrogen in Technique and the Winkler method of oxygen determination (Mackereth et al., 1978). Composite samples (used just to have all groups and silicon in diatoms are regarded as growth- almost uniform samples for the lake in taking samples at different limiting nutrients. depth of the lake) was produced using water samples collected Ethiopia is endowed with a large number of standing from selected depths distributed within the euphotic zone which water bodies, whose sustainable use can contribute to was used in the estimation of photosynthetic production. Duplicate the economy of the country. The lakes are critical to the light bottles were attached to a suspension line prepared for survival of local communities as they are the actual and incubation purpose, at each of the established depths distributed within the euphotic zones (0.00, 0.25, 0.50, 1.00, 1.50 and 1.75 m). potential sources of food and income. Despite their importance, the limnology of some of the Ethiopian lakes is unexplored. Lake Kuriftu is one such lake, which has Statistical analysis not received attention in spite of its potential economic importance. So, the purpose of this research study was, The relationships between the different physico-chemical and therefore to study the temporal dynamics of biological parameters were tested statistically using Minitab ver.14. photosynthetic primary production of phytoplankton in relation to some physico-chemicals in Lake Kuriftu. RESULTS AND DISCUSSION

Study area Physical parameters

Lake Kuriftu is one of the lakes found in and around the The physical characteristics of Lake Kuriftu during the town of Bishoftu. It is found at an altitude of 1860 m, 47 study period were measured. Lake’s transparency km southeast of Addis Ababa. The lake is located at 8° (vertical visibility) varied between 0.35 m in April 2006 of

47’ N and 39° 00’E. Lake Kuriftu is a shallow ( ≈6 m) the minor rainy season and 0.6 m in December 2005 of (Brook et al., 2001) lake formed by diverting and the dry season at the open water station. damming the tributary of the perennial Mojo River, The surface water temperatures at the central station of Belbela River, for the irrigation practice in the area (Seifu Lake Kuriftu ranged from a minimum of 22.8°C in et al., 2001). February 2006 to a maximum of 33.3°C in January 2005 with most values between 28 and 30°C. The maximum surface water temperatures of Lake Kuriftu are closer to MATERIALS AND METHODS those of Lake Chamo (26 to 30°C; Eyasu, 2004) in Ethiopia and Lake George, in Uganda (26 to 36°C; Ganf Sampling protocol and Horne, 1975) than to those reported for the Ethiopian Rift Valley Lakes including Lakes Ziway (18.5 to 27.5°C; Two stations of samplings were selected from an area of high Girma, 1988), Abijata and Langano (18 to 27°C; human impact (near-shore station) and another from a relatively less impacted area (central station). Water samples were collected Elizabeth and Amha, 1994 ) and Awassa (23.8 to 28.4°C) at least once a month from the two stations with a bottle sampler (Demeke, 1985) and other lakes of the same region as (Ruttner). The water samples were collected from selected depths the present study lake, Lakes Kilole (18.5 to 24°C; Brook, (0.00, 0.50, 1.00, 1.50, 2.00 m) distributed within the euphotic zone. 1994), Babogaya (20.5 to 28.4°C; Yeshiemebet, 2006) The samples collected from different depths were mixed in equal and the Legedadi Reservoir (22.2 to 23.9°C; Adane proportions to produce a composite sample for measurement of biomass, chlorophyll a concentration and photosynthetic primary Sirage, 2006). production. The vertical distribution of temperature in Lake Kuriftu shows the occurrence of small differences between successive depths of the water column down to 4 m. In Measurement of physico-chemical parameters in the field this lake there seemed to be no thermal stratification considering its shallow depth and exposure to wind action The parameters measured in situ include the following: although the oxygen-temperature meter used did not allow the determination of temperature at depths deeper (i) Secchi depth was estimated with a standard Secchi disc of 20 than 4 m. The thermal regime of Lake Kuriftu is probably cm diameter. (ii) pH was measured in situ by a portable digital pH meter (Jenway comparable to the shallowest Crater Lake Kilole and 3200). Lake Ziway (Girma, 1988) which are almost continually (iii) Depth profiles of oxygen were determined with a digital oxygen stirred to their bottoms. Zelalem 73

Table 1. Collective chemical features: pH, total alkalinity (TA) and phenolphthalein alkalinity (PA) and calculated free CO2 of Lake Kuriftu.

Sampling date pH Total alkalinity (PA) (meq/l) Free CO2 (mg/l) 8-08-05 8.2 2.6(0.2) 0.576 27-08-05 8.7 3(0.8) 0.528 8-10-05 8.5 2.9(0.8) 0.504 15-10-05 8.6 2.5(0.2) 0.552 29-10-05 8.6 3(0.8) 0.528 19-11-05 ND 2.7(0.7) 0.480 15-12-05 8.8 3.1(0.9) 0.528 9-01-06 8.42 2.9(0.6) 0.552 5-02-06 8.72 3(0.8) 0.528 1-03-06 8.45 2.4(0.2) 0.528 25-03-06 ND 2.5(0.4) 0.504 26-04-06 8.3 2.3(0.2) 0 .504

ND, Not detected.

Chemical features of Lake Kuriftu January and March, 2006 respectively, with the maximum value coinciding with one of the peaks in phytoplankton Depth profiles of oxygen biomass. The depth distribution of percentage saturation of dissolved oxygen often showed a pattern similar to that Depth profiles of dissolved oxygen determined at the of absolute values of oxygen concentration. Marked central station of Lake Kuriftu during the study period temporal and vertical variations in oxygen concentrations were observed. All depth profiles showed oxygen were observed over the study period. maximum in the upper layer of the water column, with a -1 minimum of 4 mg O2 l at the surface in March 2006 and -1 a maximum of 17.15 mg O2 l in December 2005). pH and alkalinity of Lake Kuriftu Dissolved oxygen was usually lower at the surface of the lake compared to the depth of 0.25 m. The observed Aggregate chemical parameters of Lake Kuriftu lower oxygen concentration at the surface is attributable measured over the study period are given in Table 1. The to the influence of temperature on the solubility of pH of the surface water of Lake Kuriftu at the central oxygen. It could also be the result of the effect of high station ranged from a minimum of 8.20 in August 2005 to light intensity on the photosynthetic generation of a maximum of 8.80 in November 2005. The pH values molecular oxygen as almost all depth profiles of recorded for this lake in the present study are slightly photosynthesis determined in the present study exhibited higher than those reported in an earlier investigation depressed photosynthetic rates at the surface. (7.90 to 8.40; Brook et al., 2001). The pH values of Lake Dissolved oxygen declined with increasing depth, which Kuriftu, which are comparable to those of Lake Awassa is related to the progressively lower oxygen contribution (Makin et al., 1975; Demeke, 1985) were generally lower of photosynthesis as a consequence of the presumably than those recorded in the nearby crater lakes, Lake lower photosynthetic biomass and exponential decline in Bishoftu (9.20; Zinabu, 1994) and Babogaya (8.84 to the level of irradiance and possibly due to the greater 9.09; Yeshiemebet, 2006) and the Rift Valley Lake demand for oxygen for oxidative decomposition of Chamo (8.53 to 9.44; Eyasu, 2004). organic matter by heterotrophs. During the study period, pH values, which are remarkably different from those concentration of dissolved oxygen at the 4 m depth also observed for Lake Kuriftu were reported only from saline - showed temporal variations from a minimum of 2 mg O2 l lakes, including Red Rock Tarn in Australia (Hammer, 1 -1 (August 2005) to a maximum of 7.6 mg O2 l 1981), Mariut in Egypt (Aleem and Samaan, 1969) and (December 2005). The oxygen concentration in the Arenguade in Ethiopia (Talling et al., 1973), which have surface water of Lake Kuriftu was generally higher than pH values between 9.00 and 10.50 that tend to remain at those recorded for the nearby lakes, Lake Kilole (3.4 to high levels owing to the high buffering capacity of the -1 10.6 mg O2 l ; Brook, 1994), and Babogaya (2.75 to 15.8 lake waters (Talling and Lemoalle, 1998). The -1 mg O2 l ; Yeshiemebet, 2006). Percentage saturation of relationship between levels of phytoplankton biomass dissolved oxygen at the surface also showed temporal and Amax and pH was not apparent although a few of the variations, with values ranging from 55 to 305% in high pH values were associated with relatively large algal 74 Int. J. Fish. Aquac.

biomass. High rates of primary productivity allow large maximum rates attained and the extent of surface - daytime CO2 and HCO withdrawal leading to an increase depression of photosynthetic activity. During the study in pH (Maberly, 1996). period, the depth-profiles of gross photosynthesis The high positive correlation between pH and alkalinity exhibited a subsurface maximum rate of gross reported for the combined data of Ethiopian lakes (Wood phtotosynthesis at 0.25 m in all sampling months except and Talling, 1988) and saline lakes worldwide (Hammer, April 2006 when the maximum was observed at the near- 1986) was also observed for Lake Kuriftu (r = 0.814). surface of the water column owing to the cloudy condition Carbonate-bicarbonate alkalinity (in meq/L) at the central of the day of production measurement. The depth profiles station showed marked temporal fluctuations, varying of photosynthetic activity observed for Lake Kuriftu from a low value of 2.3 meq/L (April 2006) to a high value (Figure 1) exhibited depressed rates of photosynthesis at of 3.1 meq/L in December 2005. The high alkalinity value the near-surface of the water columns during most of the was observed during the dry period, which was probably sampling months. Profiles with reduced photosynthetic associated with evaporative concentration of dissolved activity at the near-surface have been reported from ions of this period. The high total alkalinity values many East African lakes including those in Ethiopia recorded in the present study are similar to that observed (Talling et al., 1973; Amha and Wood, 1984; Girma, in a previous study (2.89; Zinabu, 1994). 1988; Demeke and Amha, 1990), Chad (Lemoalle, 1983), The alkalinity of Lake Kuriftu, which is close to that of Kenya (Talling, 1965; Melack, 1981; Vareschi, 1982) and Koka Reservoir (Elizabeth, 2006), is very low compared Tanzania (Melack and Kilham, 1974). Basically, lower to those of the other lakes in the same area including photosynthetic rates of phytoplankton at a lake's surface Lakes Bishoftu (20 meq/l; Wood and Talling, 1988) and are linked to photo-inhibition, which is believed to occur Babogaya (6.4 to 12.1; Yeshiemebet, 2006). Variations in when light exceeds physiological saturation and results in the amount of rainfall can bring about considerable excess of photons that do not become dissipated by differences in lake levels and concentration of dissolved photosynthetic carbon fixation (Long et al., 1994; chemicals either through input of more ions or dilution. Falkowski and Raven, 1997). The decrease in The level of free carbon dioxide was calculated from photosynthetic rates is associated with photo-oxidative pH-alkalinity relationship (Table 1) described in Lind disruption of pigment systems (Amha and Fogg, 1978; (1979). The level of free carbon dioxide ranged from Falkowski and Raven, 1997), inactivation of

0.480 in November 2005 to 0.576 mg l-1 in August 2005. photosynthetic enzymes (Steemann-Nielsen, 1962; The maximum value of free carbon dioxide coincided with Steemann-Nielsen and Jørgensen, 1962) and increased the minimum value of photosynthetic activity and biomass photorespiration (Harris and Lott, 1973; Osmond, 1981). while the minimum value was observed at the time when Considering the significance of inhibition of photosynthetic activity peaked, indicating greater photosynthesis at a lake’s surface for water column photosynthetic removal of dissolved inorganic carbon. productivity, the extent of percentage reduction in gross The calculated concentrations of free carbon dioxide in photosynthesis from Amax due to photo-inhibition was Lake Kuriftu are lower than most of the values reported estimated by calculating the difference between for Legedadi Reservoir (0.26 to 3.15; Adane, 2006). maximum gross photosynthetic rate and gross photosynthetic rate at the near-surface and expressing it as a percentage of the latter. The reduction in Photosynthetic primary productivity photosynthetic rates due to photo-inhibition varied from 0 to 16.70%. Depth profiles of gross photosynthesis As a function of irradiance, the extent of inhibition of photosynthetic productivity was variable. An irradiance -2 -1 The depth profiles of gross photosynthetic rate per unit (PAR) of 11.78 E m h in March 2006 produced 16.60% -3 -1 water volume (A, mg O2 m h ) are shown in Figure 2. reduction from the light-saturated rate (Amax) while a The vertical distribution of photosynthetic activity per unit lower irradiance of 5.849 E m-2 h-1 in August 2005 caused volume of water in Lake Kuriftu was of a typical pattern only 5.07% reduction from Amax. Similar results were for phytoplankton (Talling, 1965; Ganf, 1974; Talling and reported from other water bodies of the tropical (Demeke Lemoalle, 1998). Since composite samples were used for and Amha, 1990) and temperate regions (Jones, 1978). It all incubations, the observed depth profiles were not seems that the extent of surface depression of expressions of varying photosynthetic biomass, but were photosynthetic rates is not a function of only the intensity rather differing responses of uniform algal biomass to of incident irradiance. Experimental studies have shown different irradiances. The depth profiles included three that the extent of photoinhibition varies with photo- main regions on all sampling days except in April 2006. acclimatization state (Kok, 1956; Talling and Lemoalle, The regions were a near-surface region of light-inhibition, 1998) and species-specific differences in photo- a sub-surface region of light-saturation and a lower acclimatization strategies (Jorgensen, 1964; Behrenfield region of light-limitation. The depth profiles of et al., 1998). The difference in the extent of photo- photosynthesis of Lake Kuriftu showed variations in the inhibition between different days of incubation in Zelalem 75

Figure 1. Map of the Lakes in and around Bishoftu town including Lake Kuriftu. 76 Int. J. Fish. Aquac.

Lake Kuriftu might be related to differences in the relative It is interesting to note that the highest biomass (~55) of importance of different species of phytoplankton at Lake Kuriftu was associated with an Amax value of 1044 -3 -1 different times of the study period. mg O2 m h , while the lower phytoplankton biomass -3 -1 The maximum (1136 mg O2 m h ) gross (46.426) yielded the highest Amax (1136). Similarly, Talling photosynthetic rate at light-saturation was observed in et al. (1973) found relatively low light-saturated rate of November 2005 coinciding with a photosynthetic biomass photosynthesis with a high algal crop and the relatively -3 of 46.426 mg Chl a m while the lowest gross low algal crop was found to yield the highest maximum -3 -1 photosynthetic rate (571 mg O2 m h ) was observed in rate (Amax) in Lake Arenguade. The lack of August 2005 to be associated with a biomass of 17.236 correspondence between biomass and Amax was also mg Chl a m-3. The low gross photosynthetic rates at light- reported for phytoplankton of several reservoirs in Sri saturation were associated with a period of the main rainy Lanka (Silva et al., 2002) and in Lake Awassa (Demeke season during which runoff brings particulate materials and Amha, 1990). that reduce light penetration. Net photosynthetic rates According to Talling (1965) and Hammer (1981), high showed a depth-distribution pattern, which was similar to maximum rates associated with low algal biomass are the -1 - that of gross photosynthesis with maximum volumetric results of high specific activity [Pmax, mg O2 (mg Chl a) h rates at a depth of 0.25 m and ranged from a minimum of 1]. It seems that there is a general stimulation of growth -3 -1 429 in August 2005 to a maximum of 1000 mg O2 m h and photosynthetic activity of phytoplankton by nitrate in October 2005. alone or in combination with phosphorus (Reynolds, 1984; Tilman, 1982; Harris, 1986; Smayda, 1990). Algal communities may respond to a decreased supply of a Photosynthetic characteristics limiting nutrient either by decreasing the optimum photosynthetic rate or by producing less efficiently at The light-saturated rate of gross photosynthesis (Amax), suboptimal irradiances (Schindler and Fee, 1975). specific-rate of gross photosynthesis at light-saturation Studies on algal nutrients emphasize the importance of -1 -1 (Pmax, mg O2 (mg Chl a) h ), Percentage reduction from nitrogen and phosphorus (Uku and Mavuti, 1994)). In the -2 Amax due to photo-inhibition, hourly integral (∑A, g O2 m study, the relationship between light-saturated h-1), daily integral rates of gross photosynthesis (∑∑A, g photosynthetic rates and levels of nutrient was not -2 -1 O2 m d ) and photosynthetically active radiation (PhAR) evident. The correlation between Amax and are given in Figure 2. photosynthetically active radiation (PAR) falling on a The maximum rates of phytoplanktonic photosynthesis horizontal surface in the lake's area was positive but -3 -1 (Amax) ranged from 571 to 1136 mg O2 m h . The modest (r = 0.526) although the relation between rates of highest values of these light-saturated rates of gross photosynthesis, magnitude of photoinhibtion and PAR is photosynthesis are higher than those reported for Lake regarded as being controversial because of their Babogaya (106 to 407; Yeshiemebet, 2006), Awassa dependence on the previous life-history of the cells (217 to 425; Demeke and Amha, 1990) and Abijata (960; (Talling and Lemoalle, 1998). Amha and Wood, 1984); all in Ethiopia and Lakes It is necessary to consider the magnitude of the light- Naivasha and Cresent Island Crater (150 to 240 mg O2 saturated rate of photosynthesis per unit of chlorophyll a -3 -1 m h ; Melack, 1979a). Much higher values of Amax have [Photosyntheic capacity or Assimilation number (Pmax), -1 -1 been reported for the Ethiopian crater Lakes Kilole and mg O2 (mg Chl a) h ] when comparing the Arenguade (10,000 to 30,000; Talling et al., 1973), the photosynthetic capacity of phytoplankton communities. shallow rift valley Lake Ziway (1640 to 4670; Girma, Biomass-specific rates at light-saturation ranged from -3 -1 -1 1988); Lake George in Uganda (1900 to 6,000 mg O2 m 18.78 to 33 mg O2 (mg Chl a) h with most values -1 -1 -1 h ; Ganf and Horne, 1975) and Lake Simbi in Kenya (950 between 23 and 30 mg O2 (mg Chl a) h . Pmax of Lake -3 -1 to 12,900 mg O2 m h ; Melack, 1970). Kuriftu is higher than those of Lakes Ziway (9.6 to 22.5; Higher rates of light-saturated photosynthesis were Girma, 1988), Awassa (4 to 19; Demeke and Amha, observed at times of high phytoplankton biomass in Lake 1990), Arenguade (11 to 18; Talling et al., 1973) in Kuriftu. The correlation between Amax and phytoplankton Ethiopia, Lakes Simbi and Sonachi in Kenya (15 to 17 biomass in Lake Kuriftu was positive and strong (r=0.67) and 8 to 14 respectively; Melack, 1981) and Lake George with the latter accounting for about 45% of the variation in in Uganda (17 to 19; Ganf and Horne, 1975). The the former. Pentecost and Happey-Wood (1978) have maximum Pmax value of Kuriftu is closer to those of Lakes also found similar high correlation (r=0.671) between Babogaya (19 to 29) (Yeshiemebet, 2006), Chamo [10 to -1 -1 maximum (light-saturated) rates of photosynthesis and 34 mg O2 (mg Chl a) h ]; Eyasu, 2004] and Lake Kilole -1 -1 chlorophyll a concentration for Welsh lakes. Positive but [16.3 to 33.7 mg O2 (mg Chl a) h ; Talling et al., 1973]. weak correlation was reported between Amax and The correlation between Pmax and Amax is negative and phytoplankton biomass in previous studies in the low (r =-0.34) which may provide an explanation for the Ethiopian Lakes Ziway (r=0.36; Girma, 1988) and Lake association of high light-saturated rates with low algal Chamo (r=0.3; Eyasu, 2004). biomass observed for Lake Kuriftu. In the present study, Zelalem 77

Figure 2. Depth profile of gross photosynthesis per unit volume at open station.

the higher Pmax value was recorded at the time of worldwide and the trend is represented in tropical lakes minimum phytoplankton biomass during the rainy season including Lake George, in Uganda (Ganf, 1972), Lake while the minimum value was observed when the Maciiwaine, in Rhodesia (Robarts, 1979). There are a phytoplankton biomass was at maximum. Pmax is number of factors that determine the photosynthetic inversely proportional to the biomass of phytoplankton as capacity of phytoplankton. It has been shown that the strong and negative correlation (r = -0.90) between temperature (Eppley, 1972), light (Beardall and Morris, the two seem to suggest. This is often encountered 1976; Falkowski, 1981), nutrient regimes (Falkowski and 78 Int. J. Fish. Aquac.

14 0.8 which showed a drop in photosynthetic capacity during 12 0.6 the markedly cooler season supported the same view. As

) 10

-1 8 has been shown by Talling et al. (1973) for Lake

PhAR (E m-2 h-1) m-2 (E PhAR

h 0.4 (m) Depth Secchi Arenguade, Melack (1979) for Lake Simbi, Kenya and

-2 6 4 0.2 Lemoalle (1973) for Lake Chad in Chad, tropical soda PhAR (E m 2 lakes can show a combination of high phytoplankton

0 0.0 Secchi Depth (m)

0 30 60 90 120 150 180 210 240 270 ] A S O N D J F M A standing crop and above-average biomass-specific rates, -1 35 h partly due the large reserve of CO2 for localized 30 -1 photosynthetic activity in condensed photosynthetic 25

20 zones (Talling et al., 1973).

max 15

(mg chl a)

2 5 Production rates per unit area 0 A P [mg O S O N D J F M A 1.2 -2 -1

) Hourly (∑A, g O2 m h ) and Daily integral rates of gross -1 -2 -1

h 0.9 photosynthesis (∑∑A, g O2 m d ) are given in Figure 3. -2 The hourly photosynthetic rates per unit area were 0.6 2 m determined by the Gird Enumeration Analysis (Olson, 0.3 1960). Hourly integral photosynthesis ranged from 0.686

mg O

  -2 -1 0.0 (August 2005) to 1.05 O2 m h (October 2005). The A S O N D J F M A highest hourly integral rate of gross photosynthesis (∑A, 1200 -2 -1 ) g O2 m h ) of Lake Kuriftu is greater than those reported

-1 1000

h for Lakes Ziway (0.0574 to 0.726; Getachew, 2004), 800 -3 Babogaya (0.47 to 0.86; Yeshiemebet, 2006) and

m 600

max 2 Awassa (0.3 to 0.725; Demeke and Amha, 1990) 400 although it is much smaller than those observed in the 200 A (mg O crater lakes Arenguade (1.43 to 2.56; and Simbi (0.62 to 0 A S O N D J F M A 5.22; Melack, 1979b). The seasonal peaks of hourly 60 integral rates were associated with peaks of chlorophyll a ) 50 -3 concentration and light-saturated rate of gross 40 photosynthesis (Amax). The correlation between hourly 30 integral rates and chlorophyll a concentration and Amax 20 was positive and strong (r = 0.71 and 0.69 respectively) 10

Chl a (mg m while its correlation with PAR was positive but weak (r= 0 A S O N D J F M A 0.28). 2005 I 2006 Values of light utilization efficiency of phytoplankton in Months Lake Kuriftu ranged from a minimum of 1% in October Fig. 6. Temporal variations in phytoplankton photosynthetic Figure parametes 3. Temporal in relation variations to biomass in phytoplankton and integral photosynthetic irradiance 2005 to a maximum of 5% in August 2005 during the parameters (closed in circle) relation of to the biomass incubation and integralperiod and irradiance Secchi (closed depth study time. In Lake Kuriftu, efficiency of light utilization circle) (open of the circle) incubation in Lake period Kuriftu. and Secchi depth (open circle) in was high during the rainy season when low Ph.A.R was Lake Kuriftu. recorded. Similar findings were reported for Lakes Ziway, Awassa, and Chamo (Girma, 2006). Keifer and Mitchell (1983) suggested that efficiency of light utilization varies as an inverse function of irradiance with maximum values Stone, 1983) and cell size (Malone, 1971) directly affect occurring at low irradiance. The light utilization efficiency photosynthetic capacity. As temperature and light are values of the phytoplankton in Lake Kuriftu, are much generally high in the tropics, algal type including cell size, higher than those obtained for phytoplankton of Legedadi nutrients and CO2 supply may be considered to be of reservoir (0.16 to 0.98%; Adane, 2006). - greater importance in determining the magnitude of The daily integral rates of photosynthesis (∑∑A g O2 m photosynthetic capacity of phytoplankton. 2 d-1, of the Lake were also determined from the hourly- Comparison of mean photosynthetic productivity in the integrated rates by multiplying with the factor of 0.9 used trophogenic zone of tropical and temperate lakes led by Talling (1965) for other East African Lakes. The Lemoalle (1981) to the conclusion that higher tropical products were then multiplied by the number of hours of rates originate from high photosynthetic capacity, which sunshine often considered for tropical lakes (that is, 10). -2 may be the result of the usual higher temperature in the The calculated values ranged from 6.174 to 9.45 g O2 m -1 tropics. The studies made on Lakes MacIlwaine, d during the sampling period. The highest daily integral Rhodesia (Robarts, 1979) and Chad (Lemoalle, 1983), value of Lake Kuriftu is considerably lower than those Zelalem 79

recorded for Lake Ziway (3.1 to 17.6; Grima, 1988), photosynthetic production of phytoplankton in relation to some Arenguade (11.25 to 44.83 g O m-2 d-1; although it is physico- chemical variables in Lake Chamo Ethiopia. M.Sc. Thesis, 2 Addis Ababa University, Addis Ababa. 72p. close to those of Lake Chamo in Ethiopia (3.8 to 10.86; Falkowski PG (1981). Light-shade adaptation and assimilation numbers. Eyasu, 2004). African Lakes with smaller maximum daily J. Plankton Res. 3:203-216. integrals include Babogaya (1.01 to 5.98; Yeshiemebet, Falkowski PG, Raven JA (1997). Aquatic photosynthesis. Blackwell 2006) and Kilole (1.49 to 2.4; Talling et al., 1973) in sciences, Oxford. 375 p. Falkowski PG, Stone DP (1983). Nitrate uptake in marine Ethiopia. phytoplankton: Energy sources and the interaction with carbon fixation. Mar. Biol. 32:77-84. Ganf GG (1974). Diurnal mixing and vertical distribution of Conflict of Interest phytoplankton in shallow equatorial lake (Lake George, Uganda). Ecol. 62:611-629. Ganf GG, Horne AJ (1975). Diurnal stratification, photosynthesis and The authors have not declared any conflict of interests. nitrogen-fixation in a shallow equatorial lake (Lake George, Uganda). Ecologia 18:165-183. Ganf GG (1972). The regulation of net primary productivity in Lake ACKNOWLEDGEMENTS George, Uganda, East Africa. In: Productivity problems of freshwaters, Z. Kajak and A. Hilbricht- Ilkowska (eds.), Krakow, Polish Scientific Publishers, pp. 693-708. First and foremost, the author would like to express Getachew B (2004). Nutrient and plankton dynamics in the littoral and heartfelt gratitude to Research Advisor, Dr. Demeke Kifle, offshore zones of Lake Ziway. M.Sc Thesis. 67 pp. for his unreserved and committed consistent support and Girma T (1988). A seasonal study on primary production in relation to guidance. Department of Biology, Addis Ababa light and nutrients in Lake Ziway, Ethiopia. M.Sc. Thesis, Addis Ababa University, Addis Ababa, 62 pp. University, deserves special thanks for providing financial Girma T (2006).Temporal Dynamics of the Species Composition, support and vehicle during the study period. Finally, Abundance and Size-Fractionated Biomass and primary production Kuriftu Children’s and Integrated Farm Center are also of Phytoplankton in Lakes Ziway, Awassa and Chamo (Ethiopia). appreciated for allowing the use of their boat throughout PhD. Thesis Addis Ababa University, Addis Ababa. 201 pp. Hammer UT (1981). Primary production in saline lakes. Hydrobiologia the study period. 18:47-78. Hammer UT (1986). Saline Lake Ecosystems of the world. Monogr. Biol. Vol. 59, Dordrecht: W-Junk publ. REFERENCES Harris GP, Lott JNA (1973). Light intensity and photosynthetic rates in phytoplankton. J. Fish. Res. Bd. Can. 20:1771-1778. Adane S (2006). Water quality and Phytoplankton Dynamics in Harris GP (1986). Phytoplankton Ecology: structural function and Legedadi Reservoir. 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