Thrust Area 6: Quality Enhancement in Fish

Project title: Integration Of High Value Indigenous Fish With Conventional Carp Culture

Personnel:A. Sinha, B.K.Mahapatra, G.H. Pailan, N.R. Kumar, N.J.Moitra (RAKVK), P. Chatterjee (RAKVK)

Achievements :

Four ponds (each of size 0.1 ha) were renovated at RAKKVK Instructional Farm, Nimpith, South 24-Parganas, W.B. Pond was prepared by clearing the weeds, manuring, liming and netting out the existing fish stock. Carp seed (rohu, catla, mrigal, silver carp, grass carp, common carp, Labeo bata and Puntius javanicus) were stocked @10,000 nos per ha. Small indigenous species viz. Amblypharyngodon mola, Puntius sophore and Salmostoma bacaila were reared separately and introduced in the carp culture ponds @ 20,000nos per ha. One pond where only conventional carp species were being reared had been kept as control pond.

Project title: Mass Scale Breeding and Enhancement of Survivality of Larvae of Magur, Clarias batrachus (Linn.)

Personnel: B.K. Mahapatra, P.K. Roy, P. Sardar, S. Datta, Somdutt, V.K. Tiwari

Achievements :

The diversified congenial habitat for harbouring as well as mass scale natural breeding of magur (Clarias batrachus) had been recorded. A pair of Magur was also successfully bred under controlled condition and studied larval rearing of Magur at CIFE Kolkata Centre for enhancing the survivality. A 30 day experiment was conducted under controlled conditions to rear fry to fingerling stage. Results showed that superior FCR, increased PER, SGR and percent survival were found in fish fed minced tubifex worms followed by fish fed the mixed diet containing artificial feed (D1-42.44% CP) and minced tubifex worms. Though the fish fed zooplankton alone or mixture of zooplankton and artificial diet showed poorer performance than the fish of dietary groups mentioned above, showed better result that the fish fed any kind of artificial diet alone in this experiment.

Project title: Nutritional Strategies To Mitigate Physio-pathological Effects Of Endosulphan In Fish

Personnel: S. Jadhao, S. Gupta, S. Munilkumar, S Dasgupta

Achievements

A sixty five days experiment was conducted to assess the effects of methyl donor supplementation (0.1% Choline, 0.5%betaine and 2% lecithin) in L rohita th exposed to endosulfan(1/10 of endosulfan) alone or concurrently to endosulfan and temperature (34 deg). Compared to control (T1), percent fish weight gain was significantly "(p<0.01)" lowered by endosulfan exposure (T2), but concurrent exposure to high temperature along with endosulfan(T3) has not exacerbated the effect. This effect in concurrently exposed fish was overcome by methyl donor supplementations including choline (T4), but % weight gain in presence of betaine(T5) and lecithin(T6) in particular was even higher than control. The HSI and GSI were not affected "(p>0.05)" by the concurrent exposure to endosulfan and high temperature. The methyl donor supplementations in endosulfan and high temperature exposed fish led to normalization of activities of most of the above enzymes. Post challenge survivability data indicated that the fish exposed to endosulfan (T ) alone and 2 concurrently exposed to temperature (T ) had 27.7 and 22 % less survival than 3 control (T ). Choline supplementation improved the survival that was either at 1 par with control (T ) or 44.45% higher than T . The survival in endosulfan and 1 3 high temperature exposed fish fed betaine (T ) and lecithin (T ) was 52.78 and 5 6 55.56% higher than concurrent endosulfan and high temperature exposed fish (T ) and 13.89 and 16.67% higher than control (T ), respectively. In other 3 1 experiment of 37 days, whereas choline significantly reduced muscle endosulfan residue, no reside was found in betaine and lecithin supplemented group. Assays for testosterone, 11-ketotestosterone, estradiol and vitellogenin have been standardized.

Project title: Studies On Safe And Effective Chemical Control Measures For Floating And Submerged Aquatic Weeds Affecting Aquaculture

Personnel:S. Datta, B.K. Mahapatra, P. Sardar

Achievements

Fluridone was tested at higher dosages at 30, 60, 90 ppb against four submerged weeds e.g. three rooted - Najas, Hydrilla, Vallisneria and one nonrooted - Ceratophyllum. Herbicidal symptoms of fluridone appeared within seven to ten days and appeared as white (chlorotic) growing points in Najas, Hydrilla, Vallisneria or pink growing points in Ceratophyllum. Under optimum conditions, 30 to 90 days were required for the desired level of aquatic weed management was achieved with fluridone. In higher dosages, Najas, Hydrilla and Ceratophyllum were affected fully where as Vallisneria was partially affected. Susceptibility to Fluridone varied depending on time of year, and water movement and stage of growth. Best result was obtained when fluridone was applied prior to initiation of weed growth and when weeds begin active growth. Late spring to summer was ideal time for controlling the weeds as during this time weeds starts growing actively, sunlight and photosynthesis rate is higher. It was observed that when some weed plants were covered with algae (chara, nitella, and filamentous species) the algae and the weed plant were not controlled by fluridone. Amongst the floating weeds - Common duckweed (Lemna minor) was fully controlled, Salvinia (Salvinia spp.) – was partially controlled, pistia and water hyacinth were not controlled by fluridone at any dose.

Project title:Immuno-physiological and Reproductive Responses to Chronic Arsenicosis in Rohu, Labeo rohita (H.) and Magur, Clarias batrachus (L.) and its Remedial Measures Through Dietary Intervention

Personnel:P. Sardar, A. Sinha, G.H. Pailan

Achievements

The acute toxicity of an inorganic arsenic compound sodium arsenite (NaAsO ) 2 was examined in rohu (Labeo rohita H.). The fishes were subjected to static bioassays trial in the laboratory for 96 hours to determine the median lethal concentrations (LC ) of NaAsO . Preliminary trials or range finding tests were 50 2 conducted to figure out the suitable ranges to be used in the final trials of lethality test. In final trials, mortality of fish was recorded at 6, 12, 24, 48, 72 and 96 hours of exposure. The LC values and their 95% confidence limits for 50 different exposure time were calculated by using computer software 'Trimmed Sperman-Karber'. After 96 hours of exposure the LC value of NaAsO 50 2 was 8.55 ppm (95% confidence limit, 5.17 to14.30) for rohu. NaAsO exposed fish showed hyperactivity with rapid swimming and strucking their head at the side of the treated tank after 48 hours exposure of arsenic. Gradually there was a drastic reduction in the activity and slower swimming of the fishes followed by mortality of fishes at 96 hour exposure. At the end of 96 h 6 3 exposure of NaAsO in rohu Hb (g/dl), TEC (x 10 /mm ), MCH (pg), liver ALT (μM 2 pyruvate/30 min/mg protein), AST (μM pyruvate/30 min/mg protein), ACP (μM pyruvate/30 min/mg protein), ALP (μM pyruvate/30 min/mg protein), liver and muscle glycogen (mg/g), total lipid (mg/g), total protein (mg/g), blood lysozyme (μg/ml) and phagocytic acitivity (NBT) (OD at 540 nm) were significantly decreased whereas plasma ALT (IU/l), AST (IU/l), ACP (IU/l), ALP (IU/l), glucose (mg/dl), total lipid (mg/dl) and total protein (g/dl) were significantly increased in comparison to fish of unexposed control. Muscle enzymes did not vary significantly. Trial for finding out the effects of chronic arsenic toxicity on the physiological and immunological profiles in rohu is under progress

Project title:Environmental Impact Of Fipronil : Presence In Water And Soil, Effect Of Fipronil And Its Metabolites On Common Edible Fish Labeo Rohita (rohu) And Bioremediation Studies

Personnel:N. Saharan, Prem Dureja (IARI, New Delhi), P.K.Pandey, G. Tripathi

Achievements

Soil and water samples were collected from six experimental sites near agricultural fields where rice and vegetable crops are grown and fipronil is used as a pesticide. The soil and water samples contained 0.15 – 0.93 ppm residual fipronil and water samples contained 0.21 – 0.97 ppm fipronil. Acute toxicity test on Labeo rohita fingerlings showed that LC value was 0.138 ppm/ litre for Fipronil 0.123ppm/ litre for Fipronil Sulfone and 0.109 ppm/ litre for Fipronil Sulfide after 96 hours experiment. All the exposed animals showed almost similar clinical signs of varying degree depending upon the concentration of fipronil in the acute toxicity test. The signs of hyper activity and restlessness were marked during exposure. Long-term sub lethal toxicity test was conducted for a period of 60 days exposing the animals to 1/10th of LC50 dose of fipronil (0.138ppm/litre) and its metabolites (0.123ppm/litre) for Fipronil Sulfone and 0.109 ppm/ litre for Fipronil Sulfide. Healthy Labeo rohita were fed with artificial purified diets incorporated with different doses of fipronil twice a day. There was 4.9 % growth in body weight in control samples but only 3.2, 3.0 and 2.8 % in samples treated with Fipronil, Fipronil Sulfone and Fipronil Sulphide respectively. These results reveal that metabolites of Fipronil are more toxic than Fipronil. Gill tissues showed marked distal and basal hyperplasia. Fusion of secondary lamellae and hemorrhages were found at some places. Liver tissue exhibited constriction of sinusoids, swelling of cells with pyknotic nuclei and infiltration of inflammatory cells were commonly observed. In the kidney, tubular epithelium showed severe necrotic changes. Multiple granulomatois changes with extensive infiltration of mononuclear cells were discernible in the parenchyma. Moderate necrosis was observed in the myocardium. No abnormality could be detected in the skin and muscle tissue. Marked changes specially in the gill tissue where hyperplasia and fusion of secondary lamellae was observed in almost all the sacrificed animals. Liver tissue showed degenerative and necrotic changes. Bio-remediation of soil: Total 10 isolates were selected based on their morphological characteristics and were further purified for testing their bioremediating capacity. These ten isolates were exposed to different concentrations of fipronil through culture media. The concentration varied 1.0, 3.0 and 5.0 ppm. Three isolates could survive up to 5.0 ppm.

Project title:Characterization And Nanoencapsulation Of Fish Pheromones For Using In Fish Reproduction

Personnel:R. Sharma, S. Munilkumar, S. Gupta, S.D. Singh

Achievements

The water sample where fishes were kept was collected during the breeding season and analyzed through HPLC and LCMS to screen the presence of pheromone. Initial studies showed the presence of 17 α-20β dihydroxy-4- pregnen-3-one (17, 20 β-P) and Prostaglandin F 2α. During the post breeding season, no pheromones were found in the water sample. The testes, ovary and seminal vesicles were collected at different stages of development and various changes were recorded. It was hypothesized that inter tubular septum might be one reason for the fish which prevented magur to ooze out the sperm in captivity. But it was found that inter tubular septum in the testes of Rohu was also thick as that of magur, nullifying the hypothesis. After that gold and chitosan based gold nanoparticles had been developed and were encapsulated with the purified synthetic steroidal pheromones and hormonal pheromone. The particle size was analyzed with Beckman and Coulter particle size analyzer and the particle size were ranged between 65 nm to 215 nm.

Project title:Studies On Oxidative Stability Of Polyunsaturated Fatty Acids During Extrusion Cooking

Personnel:G.Venkateshwarlu, S.Basu, B.B.Nayak

Achievements

In order to understand the extent of amylose-lipid complexation, the lipid was extracted from rice flour and also from rice extrudates after extrusion cooking. The lipid recovered from extrudates was found to be 5.20±0.58 mg/g against the lipid extracted from the rice flour i.e. 10.98±1.20 mg/g. The lipid that became unavailable after extrusion was considered to interact with eitherthe protein, the carbohydrate, or both fractions. Further, the extrudates were digested with α-amylase to release the bound form of lipid which yielded 2.24 ± 0.16 mg/g. This lipid that became available after digestion was considered to interact with the amylose fraction.

Project title:Development Of Improved Quality Ready To Eat Fish Products And Study Of Their Storage Characteristics In Retortable Pouches

Personnel:S.Basu, G. Venkateshwarlu, B.B.Nayak

Achievements

A standard recipe was finalized to prepare Tilapia fish curry in south Indian style. To standardize the heat process schedule (F value), the tilapia steaks of 0 20 mm uniform pieces was soaked in 10% salt solution for 1h and added to curry. The fish curry was subjected to different heat process having F value of 0 6, 7, 8 and 9 for two different temperatures of 116.1 o C and121.1 o C. The 8 samples thus obtained were subjected to organoleptic evaluation and commercial sterility test. It was found that heat process schedule of F =6.7 at 116.1o C yielding a better acceptable product and microbially safe. Tilapia fish curry thus prepared and standardized was fortified with EPA and DHA using cod liver oil at 1% level with a control for comparison. Then the fish curry was 0 subjected to heat process at 116.1 o C for F =6.7. Tilapia fish curry fortified with 0 EPA and DHA with control was analysed for organoleptic, biochemical characteristics, fatty acid profile before and after heat processing in Retort pouch. The products have been in excellent condition for one year. The sandwich paste heat processed at 121.1 o C at F0 4 was analysed for fatty acid profile and other biochemical indices. It was noticed that there was no change in fatty acid content during heat processing. During storage up to one year, there was no significant change in the concentration of EPA and DHA and there was no significant oxidation of the unsaturated fatty acids. The prawn curry fortified with EPA and DHA by adding cod liver oil at 2% level and packed in retortable pouches and heat processed at F value of 6, 7, and 8 at 110 o C, 116 and 121.1 C. F of 7 at 116 o C was found to be organoleptically acceptable.It was observed that upto 1 year the product was acceptable and no significant oxidation of the unsaturated fatty acids occurred.

Project title:Molecular Analysis Of Growth Promoting Peptide Hormone From Food Fish (mullet/seabass)

Personnel:S. D. Singh, R.S. Rana, S. Gupta

Achievements

Oligonucleotide primers as given in previous report were utilized for the synthesis of IGF I and Leptin (Obese) gene. Quality and purity of total RNA isolated from Asian Seabass, Lates calcarifer and rohu ( as prototype) (about 500g live weight) liver as per standard protocol were good and used to synthesize cDNA gene of about 350 base pair for IGF I of Asian seabass and about 450/600 bp leptin obese gene for Rohu/seabass respectively by RT - PCR. These peptide hormones genes were cloned in plasmid vector pTZ57R/T and Escherichia coli DH5α strain by T/A cloning and characterized by restriction enzyme analysis and colony PCR. These specific white recombinant clones specific for leptin hormone i.e. Obese gene and IGF I gene are cryopreserved in glycerol and are also used for nucleotide sequencing to compare their DNA sequences with USA NCBI gene bank database. Presence of Start and stop codons in both of them are to be ascertained before using for further molecular studies. Rohu partial leptin gene after automatic sequencing revealed the maximum homology (97%) with Cyprinus carpio ob 2 gene, followed by Carassius auratus (89%), Ctenopharyngodon idella (85%) and Danio rerio (72%) on BLAST search in the NCBI GenBank database.

Project title:Studies On Viability Of Litopenaeus vannamei Culture In India

Personnel:G. Venugopal, S.S.H. Razvi, Suresh Babu, V. K. Sharma, V. Hari Krishna, K. V. Rajendran, M. Makesh, P.S. Ananthan

Achievements

As per the guidelines Coastal Aquaculture Authority of India the farm was registered. CAAI committee has inspected the Effluent treatment systems and biosecurity measures installed at farm and granted permission to take up culture for 5 years. Water samples from fish ponds have been analyzed for Vibrio sp and TPC load and enumerated the colonies pre & post treatment. On 12-03-2010 L. vannamei PL (8-10 mm) were procured and transported from licensed CP Aquaculture hatchery, Gudur(AP) and after proper acclimatization released in the culture ponds. As per the technical programme of the project for the first year the following stocking densities were adopted viz 10 /m2 in two 0.2 ha ponds, 20 / m2 in two ponds of 0.2 ha each, 30/ m2 in one pond of 0.4 ha. Live seed samples were screened for various viral and bacterial pathogens at CIFE Mumbai. Simultaneously, samples of seed were also screened at CIFE Kakinada centre's Molecular Bio Lab and found WSSV and MBV free as well as free from ecto-parasites. Prestocking and post stocking - Vibrio loads and Total Plate Count of Seed, water and soil were analyzed and has been continued subsequently at fortnight intervals. Soil and plankton samples are analyzed at fortnightly intervals. In all the ponds aeration was given using pond aerators. Commercial grade CP feed is being applied 4 times a day. During 20 days of grow out period the growth of L .vannamei was recorded to be in the range of 2.8 to 3.5 grams and survival was estimated to be in the range of 80- 95 % at different stocking densities.