The experiment was conducted in nine earthen ponds for a period of 6 months from May to November 2013 at the Department of Aquaculture, Bangladesh Agricultural University (BAU), Mymensingh-2202, Bangladesh.
2.1 Description of experimental ponds Nine earthen ponds were used to conduct the experiment. All of them were situated to the Southwest side of wet laboratory complex, Faculty of Fisheries, BAU, Mymensingh. The ponds were rectangular in shape, 40 m2 in size, 1 m in depth, and having strong dykes, flat bottom etc. All of the ponds were well exposed to sunlight, free from unwanted aquatic vegetation and well managed and equipped with inlets and outlets. However, the inlets and outlets were sealed with polythene sheets.
2.2 Experimental design The experiment consisted of three treatments (T1, T2, and T3) with three replications for each treatment: T1 - carps and shing; T2 - carps, shing and snail; and T3 - carps, shing, snail, tilapia, and water spinach. Eighty carps (catla, silver carp, rui, mrigal = 3:1:2:2) and 100 shing in a 1 m3 cage were stocked per 40 m2 pond area in all the three treatments. An additional cage of same volume stocking with 100 tilapia was set in T3. Snail was stocked at the rate of 250 g/40m2 of pond area in T2 and T3. Water spinach was cultivated on four floating trays in T3 only.
2.3 IMTA pond preparation All the ponds were dried out completely. The undesired small fish, aquatic weeds etc. were removed from the ponds. Excess bottom mud was removed and used to repair the broken and uneven dykes. Lime and compost were applied at 250 and 680 kg/ha during pond preparation. Compost was prepared in a pit of a dyke using mustard oil cake (36.5%), cow dung (36.5%), urea (9%) and water hyacinth (18%), and applied as basal manure for feeding snail and enhancement of primary productivity.
2.4 Sample collection All of the samplings were done in the morning between 9.00 and 10.00 AM for the estimation of phytoplankton. A 500 ml glass jar was used for the collection of 1000 ml water. After collection of water from the surface of ponds, it was concentrated to 90 ml passing through a bolting silk plankton net and then kept in a vial. Then the collected samples were preserved in 10% formalin and transferred to the laboratory as soon as possible for further analyses.
2.5 Phytoplankton Study Two types of studies were done in the laboratory using the collected samples: qualitative and quantitative. A qualitative study gives an idea on the types of phytoplankton present in the experimental ponds; on the other hand, a quantitative study estimates the number of various phytoplankton in water of the experimental ponds. Finally, primary productivity of the experimental ponds was determined by the quantitative study. 2.5.1 Qualitative study The qualitative analyses of phytoplankton were done up to the genus level according to Pennak (1953) [18] and Bellinger (1992). The identification of phytoplankton was done using a digital microscope (ANOVA 950 ES). 2.5.2 Quantitative study Estimation of phytoplankton was done with a plankton counting cell named Sedgewick- Rafter cell (SR cell) which is a special type of thick slide having a cell in the center, which is divided into 1000 small squares on the surface. Phytoplankton was counted by placing the cell under a high power microscope with the projection of 10×15 and the number of phytoplankton was expressed as cells/l. 2.5.2.1 Preparation of slide The Sedgewick-Rafter (S-R) counting cell is 55 mm long, 20 mm wide and 1 mm deep, and volume of the chamber is 1 ml, the counting chamber is equally divided into 1000 fields, each of the fields having a capacity of 1 micro liter. One ml from the concentrated volume of the phytoplankton samples was taken on the S-R cell with a dropper. Then the counting chamber was covered with a cover slip so as to eliminate the air bubbles and left to stand for a few minutes to allow the phytoplankton settle down. Further analyses were done placing the cell under the microscope. 2.5.2.2 Study under microscope One ml sub-sample was transferred to the Sedgewick-Rafter (S-R) counting cell and placed under a binocular microscope (ANOVA 950 ES). Phytoplanktons were counted from 10 random fields of the S-R counting cell, and was expressed numerically cell per liter of water according to APHA (1992).
2.6 Studies on water quality parameters The water quality parameters namely temperature (°C) - was recorded using a Celsius thermometer; pH and dissolved oxygen (mg/l) - were measured using HACH kit (Model: FF1A) fortnightly during the experimental period. Samples were collected from 09.00 to 10.00 AM and analyses were performed in the Laboratory of the Department of Aquaculture, BAU, Mymensingh, Bangladesh.
2.7 Data analysis The collected data were entered into a Microsoft Excel spreadsheet and then analyses were done using a statistical software, SPSS (Statistical Package for Social Sciences) Version-16.0. One-way analysis of variance (ANOVA) was employed to analyze any significance of difference among the treatment means followed by Duncan’s multiple range test (Duncan, 1955) and the level of significance was assigned at 0.05 (Zar, 1999).