Effects of different stocking densities and diets on the growth and survival rate of black apple snail (pila polita)

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CAN THO UNIVERSITY COLLEGE OF AQUACULTURE AND FISHERIES EFFECTS OF DIFFERENT STOCKING DENSITIES AND DIETS ON THE GROWTH AND SURVIVAL RATE OF BLACK APPLE SNAIL (Pila polita) By DANG MINH QUAN A thesis submitted in partial fulfillment of the requirements for the degree of Bachelor of Aquaculture Science Can Tho, December 2013 CAN THO UNIVERSITY COLLEGE OF AQUACULTURE AND FISHERIES EFFECTS OF DIFFERENT STOCKING DENSITIES AND DIETS ON THE GROWTH AND SURVIVAL RATE OF BLACK APPLE SNAIL (Pila polita) By DANG MINH QUAN A thesis submitted in partial fulfillment of the requirements for the degree of Bachelor of Aquaculture Science Supervisor Assoc.Prof.Dr. NGO THI THU THAO Can Tho, December 2013 Acknowledgements First of all, I wish to give my honest thank to Rectorate Board of Can Tho University, lecturers and instructors of CAF and Auburn University who have facilitated for my studying during 4.5 years in Can Tho city. I would like to thank Assoc. Prof. Dr. Ngo Thi Thu Thao and Mr. Le Van Binh who have instructed me enthusiastically to finish this graduating thesis. For other valuable help and guide, thanks are extended to all my friends, Mr. Le Van Binh, Mr. Nguyen Chi, Ms. Nguyen Kim Cuong, Ms. Tran Thi Be Gam, Ms. Nguyen Khanh Linh and friends in AAP course 35. I wish to express my sincere gratitude to my advisor, Dr. Duong Thuy Yen for her constant guidance, and thanks to all my beloved classmates in Advanced Aquaculture Program class for all great encouragement and kind help during 4.5 years in CAF. Finally, I thank my family and all my friends who have supported and encouraged me to study and finish my course. DANG MINH QUAN i ABSTRACT This study consists of two experiments on Black apple snail Pila polita . The first experiment was aimed to evaluate the effects of different densities on the growth and survival rate. There were 3 replicates in each treatment and densities as follow: 300, 600, 900, 1200 ind./m2. Snails with initial weight (0.03g) and shell height (4.5 mm) were reared in the composite tanks and fed with industrial pellet. After 42 days, the survival rate at 300 ind./m2 (94.00%) was higher than at 600 ind./m2 (89.44%), 900 ind./m2 (82.30%) and 1200 ind./m2 (80.89%), there was significant difference (P<0.05). Rearing at 300 ind./m2, snails reached highest body weight and shell height (0.33 g and 11.54 mm) compared to 600 ind./m2 (0,29 g and 10.29mm), 900 ind./m2 (0.23 g and 9.84 mm) and 1200 ind./m2 (0.21 g and 9.70 mm). At the stocking density of 600 ind/m2, snails also presented highest feed efficiency (644.4 %) and that was significant difference (p<0.05) compared to other treatments. In results, snails were reared at density of 600 ind./m2 had a good growth performance, survival, productivity and economic effectiveness. The second experiment was conducted to evaluate the effect of different diets on the growth and survival rate of snail Pila polita. Snails were reared at density of 600 ind./m2. A triplicate experiment with 4 different treatments as follow: Rice bran incubated with Bacillus subtilus (RB), RB + B. subtilus supplemented directly into cultured tank, Industrial pellet powder incubated with Bacillus subtilus (IPB) and IPB + B. subtilus supplemented directly into cultured tank. After 42 days, survival rate of treatment 2 (88.00±5.17%) were higher than treatment 1 (81.89±11.03%), treatment 3 (75.00±11.355) and treatment 4 (79.44±7.65%) but no significant difference (p>0.05). Treatment 1 reached highest body weight and shell height (0.20g and 9.51mm) compared to treatment 2 (0.19g and 9.14mm), treatment 3 (0.18g and 9.05mm) and treatment 4 (0.18g and 9.06mm), however there were significant differences (p<0.05). Treatment 2 had the lowest FCR value (0.36) but no significant difference (p<0.05) compare to other treatments. Feeding with treatment 1 had the highest value in biomass growth rate (106.74%), FE (414.22%) and yield (82.14g/m2) compare to other treatments, but no significant difference (p>0.05). Results indicated that Pila polita were reared at 600 ind./m2 and fed with RB fulfilled the requirements on interested aspects such as growth performance, survival, productivity and economic effectiveness. TABLE OF CONTENTS Acknowledgements ..................................................................................................... i ABSTRACT .............................................................................................................. ii LIST OF TABLES ......................................................................................................v LIST OF ABBREVIATIONS .................................................................................. vii CHAPTER 1 ...............................................................................................................1 INTRODUCTION ......................................................................................................1 1.1. Introduction .........................................................................................................1 1.2. Objectives of the study .........................................................................................2 1.3. Contents of research .............................................................................................2 CHAPTER II ..............................................................................................................3 LITERATURE REVIEW............................................................................................3 2.2. Biological characteristics .....................................................................................4 2.2.1. External morphology .........................................................................................4 2.2.2. Habitat and distribution................................................................................4 2.2.3. Food and nutrition........................................................................................5 2.2.4. Reproduction ...............................................................................................6 2.3. Seed production and grow out ..............................................................................6 2.3.1. Seed production ...........................................................................................6 2.3.2. Grow out......................................................................................................6 2.4. International and domestic research activities on Pila polita ................................7 2.4.1. International research activities ....................................................................7 2.4.2. Domestic research activities .........................................................................7 2.5. Probiotics in aquaculture ......................................................................................8 CHAPTER III ...........................................................................................................10 MATERIALS AND METHODS ..............................................................................10 3.1. Time and location ..............................................................................................10 3.2. Materials ............................................................................................................10 iii 3.2.1. Equipment .......................................................................................................10 3.2.2. Feeds .........................................................................................................10 3.2.3. Water resource ...........................................................................................11 3.2.4. Incubated feed making proceduce ..............................................................11 3.3 Methods ..............................................................................................................13 3.3.1. Experimental design ..................................................................................13 3.3.2. Sampling and data collection .....................................................................15 3.3.3. Determine the total density of bacteria, Vibrio and Bacillus sp. in the tank 17 3.4. Statistical analysis ..............................................................................................17 CHAPTER IV ...........................................................................................................18 RESULTS AND DISCUSSIONS .............................................................................18 4.1 Effects of different stocking densities on the growth and survival of Pila polita..18 4.1.1 Water quality parameters..................................................................................18 4.1.2 Growth of Black apple snail .............................................................................21 4.1.3 Growth rates of snails.......................................................................................23 4.1.4. Biomass growth rate and feed efficiency of snail.............................................24 4.2. Effects of different diets on the growth and survival rate of Pila polita ...............25 4.2.1. Water quality parameters .................................................................................25 4.2.2. Total bacteria counts and density of Bacillus in different treatments ..............29 4.2.3.1 Growth rate in shell height .......................................................................30 4.2.3.2 Growth rate in body weight ......................................................................32 CHAPTER 5 .............................................................................................................35 CONCLUSIONS AND RECOMMENDATIONS .....................................................35 5.1. Conclusions .......................................................................................................35 Effects of stocking density on the growth and survival rate of Pila polita ..................35 Effects of different diets on the growth and survival rate of Pila polita......................35 5.2. Recommendations ..............................................................................................35 REFERENCES .........................................................................................................36 APPENDIX ..............................................................................................................41 iv LIST OF TABLES Table 2. 1 Characteristics of the genus Pila .................................................................4 Table 3.1 Physical and chemical parameter collection………….…………………...15 Table 4. 1 Mean values of environmental parameters during culture period 21 Table 4. 2 The specific growth rate in shell length during experiment (%/day) .........23 Table 4.3 The specific growth rate in snail body weight (SGRweight, %/day) ..............24 Table 4. 4 Survival rate, biomass growth rate, Feed conversion ratio (FCR) in different stocking densities .......................................................................................25 Table 4. 5 Mean values of environmental factors in the treatments ..........................28 Table 4. 6 The Specific Growth Rate (%/day) of snail body weight during rearing period........................................................................................................................33 Table 4. 7. Survival rate, biomass growth rate, feed conversion ratio (FCR) in the same snail treatments ................................................................................................34 v LIST OF FIGURES Figure 2. 1 Snail from side view and abdominal view .................................................3 Figure 3. 1 Biosubtyl DL and Biosubtyl-II………………………………………….11 Figure 3. 2 Rice bran incubated with Bacillus subtilus (RB) .....................................12 Figure 3. 3 Industrial pellet powder incubated with Bacillus subtilus (IPB)...............12 Figure 3. 4 Experimental system ...............................................................................13 Figure 4. 1 Variation of temperature during experimental period.…………………..18 Figure 4. 2 Variation of pH during experimental period ............................................19 Figure 4. 3 Variation of TAN during the culture period.............................................20 Figure 4. 4 Variation of NO2 during the culture period..............................................20 Figure 4. 5 Variation of Alkalinity during the culture period .....................................21 Figure 4. 6 Variation of shell length of snails during the culture period.....................22 Figure 4. 7 Variation of snail body weight during the culture period .........................23 Figure 4. 8 Variation of temperature during experimental period ..............................26 Figure 4. 9 Variation of pH during experimental period ............................................27 Figure 4. 10 Variation of TAN during experimental period ......................................27 Figure 4. 11 Variation of NO2 during experimental period .......................................28 Figure 4. 12 Variation of a total bacteria counts in different treatments during 7 days .................................................................................................................................29 Figure 4. 13 Variation of Bacillus density in different treatments during 7 days ......30 Figure 4. 14 Variation of average shell height of snail during experimental period ..31 Figure 4. 15 Variation of specific growth rate in shell height during experimental period........................................................................................................................31 Figure 4. 16 Total weight of snail in the treatments during experimental period ........32 Figure 4. 17 Variation of survival rate of snail during experimental period ...............33 vi LIST OF ABBREVIATIONS DLG Daily length gain DWG Daily weight gain SGRlength Specific growth rate in shell length SGRgrowth Specific growth rate in body weight IPP Industrial pellet powder RB Rice bran incubated with Bacillus subtilus (RB) IPB Industrial pellet powder incubated with Bacillus subtilus (IPB) SL Shell length W initial Weight at initial time of experiment L initial Length at initial time of experiment W final weight at the end of experiment L initial length at the end of experiment t Experiment duration (days) SR Survival rate Nt Number of alive snails No Initial number of released snails FE Feed efficiency FCR Feed conversion ratio vii CHAPTER 1 INTRODUCTION 1.1. Introduction In recent years, with the development trend of the world economy, the domestic aquaculture industry has been developing rapidly in both quantity and quality, as well as expanding the model culture. Beside providing food, aquaculture products are also exported with high economic value, so aquaculture have become a key sector of the economy of Vietnam. Vietnam has many advantages and suitable conditions for aquaculture development. With a coastline of 3620 km, 112 of the river system of rivers, canals, lagoons and dense, large water surface field is a favorable condition to develop aquaculture. To promote the advantages which require investment in research planning and supply of seed for the needs of the farmers. In freshwater aquaculture recent years has become a professional manufacturer to bring high economic efficiency. Freshwater aquaculture training products help improve people's lives and contribute to poverty reduction. In addition to the traditional freshwater species such as common carp, silver carp, catfish, a species as black apple snails (Pila polita) is relatively new, but very promising because it is relatively high economic value. The dish is made from Black apple snails with delicious quality, nutritious (containing 11.9% proteins; 0.7% lipid; vitamins B1, B2, PP; the mineral Ca, P; provide energy of 86 calo/100g meat), amino acids and in particular contain unsaturated healthy fats. (Do Huy Bich et al, 2003) Due to the current supply of Black apple snails mostly from by wild-caught therefore it may be not stable, also it will increase the fishing pressure lead to a decline of biodiversity in fresh waters. Currently, a few studies conducted in breeding Black apple snails with seed collecting from the wild. The results are quite satisfactory, and could bring high economic efficiency for the farmers. On the other hand, this study also contribute a solution to reduce the fishing pressure and conserve natural resources of local species. Based on research and practical demands, this study on “Effects of different stocking densities and diets on the growth, and survival rate of black apple snail Pila polita” was be carried out. 1 1.2. Objectives of the study To find out the appropriate foods and stocking density for rearing Black apple snails. These results could contribute to improve culture technique of Pila polita. 1.3. Contents of research Experiment 1: evaluate the effect of different stocking densities on the growth and survival rate of Black apple snails. Experiment 2: evaluate the effect of different diets on the growth and survival rate of Black apple snails. 2 CHAPTER II LITERATURE REVIEW 2.1 Classification Phylum: Mollusca Class: Gastropoda Order: Architaenioglossa Family: Ampullariidae Genus: Pila Specific name: polita Scientific name: Pila polita English Name: Black Apple Snail Vietnamese name: Ốc bươu đồng, Ốc nhồi In the world, so far 23 species of Pila.sp has classified, including: P. africana, P. Africana martens, P. ampulacea, P. ampullacea, P. angelica, P. cecillei, P. congoensis, P. conica, P. globosa, P. gracilis, P. leopoldvillensis, P. letourmenxi, P. luzonica, P. occidentalis, P. ovata, P. pesmei, P. pesmi, P. polita, P. saxea, P. scutata, P. speciosa, P. virens, P. wernei Figure 2. 1 Snail from side view and abdominal view 3 2.2. Biological characteristics 2.2.1. External morphology Large snails, glossy shell surface, blue yellow or brown colour outside, the inside have light violet colour. The number of whorls is 5.5 - 6, slightly bulging and shallow twisted grooves. Long narrow operculum with the width is at half height, sharp top shell. The last spire is large, up to 5/6 of shell height while the spirals above are small. Table 2. 1 Characteristics of the genus Pila (http://www.applesnail.net/content/pila.htm) Surface SHELL Shape Direction Whorls Shell-opening (aperture) Umbilicus Colour OPERCULUM Structure Head (cephalic) tentacles BODY smooth to rough (growth lines) egg-shaped, ovoid to globose right (dextral) round oval to egg-shaped wide, narrow to closed yellow, dark brown to almost black, with or without spiral bands corneous outside, calcified inside Long Labial tentacles Breathing siphon Colour Long Medium grey-yellow to grey, with dark spots Position above the waterline EGGS 2.2.2. Habitat and distribution Dillon (2000) reported that Pila polita are present in fresh waters in Indochina, Indonesia, China, Thailand and Vietnam. They live in ponds, fields, plains and midlands. When moving, snails open operculum, spread the abdominal muscles as flexible blade on the bottom or on the wall, and secretes a mucous layer to reduce friction. While moving the head protruding, mouth lobe is in the middle and two siphon trunks. 4 The feet withdraw into the shell when operculum closes and stretching when moving, at this time operculum fold on the back side. Snails usually float to the surface to breathe, when there is noise, snails immediately withdraw into the shell and drive to the bottom. In hot or cold weather, they float to the surface. Snails have both gills and lungs so they can live in water and on land. Being as fresh water snail, but some Ampullariids species may be able to tolerate at low levels of salinity (Prashad, 1925; Hunt, 1961; Fujio et al., 1991; Santos et al., 1987), however they generally do not live in brackish waters. Most species are amphibious, able to spend significant lengths of time out of water for breathing air. Many species, especially Pila, Pomacea, Marisa and Lanistes, inhabit slow-moving or stagnant waters in lowland swamps, marshes, ditches, lakes, and rivers (Pain, 1950, 1960; Andrews, 1965; Robins, 1971; Louda and McKaye, 1982; Keawjam, 1986). 2.2.3. Food and nutrition Black apple snail are not selective feeding and eat almost everything available in their environment. In general, they prefer soft and digestible vegetation. Tougher plants and algae are consumed as long as they are able to grasp pieces of with their radula (rasp tongue). When there is not enough food available in the water, Black apple snails can profit from their amphibian life style to leave the water in search for food. An interesting strategy used by Black apple snails to attain food is exhibited when the food is floating on the surface (surface film feeding). In that case the snail crawls to the surface and forms a foot-funnel in to trap particles from the surface. To attract more floating food, the snail makes the same movement with its foot as it does for walking with the front part of the foot. The middle part and the tail of the foot, the snail remain attached to the side or an object near the surface. Once the funnel is filled, the snail brings its head in the funnel and eats the collected material. Behaviour is known as ciliary feeding. Black apple snails are opportinustic and even consume all kinds of dead animals like dead fish, frogs, crustaceans and insects and eggs (fish, frogs, snails etc.). Since 5 the high nutritional value of this alternative food source is high, this behaviour fits well in their survival strategy (http://www.applesnail.net/content/ecology.php). 2.2.4. Reproduction Ampullariids are dioecious, internally fertilizing and oviparous (not reciprocally – fertilizing hermaphrodites as stated by Chang, 1985). Species of Pila have been reported to change sex (Keawjam, 1987; Keawjam and Upatham, 1990). The sex change is from male to female (protandry) and takes place during aestivation. The larger size of females in Pila has therefore been attributed to continuing growth following this change. 2.3. Seed production and grow out 2.3.1. Seed production Ponds with soft muddy bottoms, organic humus, average water levels around 0,5m and slightly flow are suitable for seed production. Ponds were fertilized with chicken manure, dung - cow mixed with chopped straw (1/3). Fertilizing the snail pond was done before stocking at least 3 days. Stocking density is 15-20 snails/m2 with male/female ratio is 1:1. Broodstock snails were released before breeding season and baby snails could be collected after 10-15 days. Care should be taken to avoid broken snail shells. (http://www.kinhtenongthon.com.vn) 2.3.2. Grow out Snails are stocked at rice field, with 0.7-1m in the ditch and 0.2-0.3m in the platform. Normally, snails were integrated culture in ditches, ponds, fields with fish. Lotus were planted to cool down the temperature and to be shelters for snails. Water would be better with slightly flow, must not contaminated with pesticides or rich in organic matter. Ponds should be fertilized with chopped straw and manure before stocking 10 days at a rate of 2 kg/m2. Stocking density varied from 100-150 snails/m2 and at 80 - 120 snails/m2 if bigger snail size was selected. Snails were fed daily with rice bran, cassava, sweet potatoes, vegetables, meat, or trash fish. Foods were fed 6 once a day with daily amount was 10% body weight of cultured snails (http://www.kinhtenongthon.com.vn) 2.4. International and domestic research activities on Pila polita 2.4.1. International research activities Dillon (2000) studied the distribution of snails Pila polita, the authors identified Pila polita commonly distributed in Indonesia, China, Thailand and Vietnam. The author also reported that Pila polita lived in the farm ponds and midland plains. Keawjam (1986) and Thaewnon-ngiw et al. (2003) studied on medical effects of Pila polita. The authors mentioned that the snail is one of eight species of freshwater snails which have an important role in medicine. It is used to treat skin diseases for local community in southern Thailand. 2.4.2. Domestic research activities Nguyen Thi Dat (2010) studied on the effects of different densities and feeds on the growth and survival rate of the snail Pila polita in grow-out period. Snails were fed with cassava leaves, homemade feed (40% rice bran, 20% corn, 10% lighter fish meal and 30% soybean meal). Stocking density was 100 snails/m2 and 150 snails/m2. Results showed that feeds and densities significantly affected the growth and survival rates of the snails. Vegetables combined with homemade diets resulted in higher growth performance, survival rate and economic efficiency. Moreover, stocking with density of 100 snails/m2 showed higher growth, survival rate and economic efficiency than stocking of 150 snails/m2. Nguyen Thi Binh (2011) studied on reproductive and biology characteristics of the snail Pila polita. The results showed that Pila polita was dioecious species and the ratio of male and female was 1:1.67. Snails were often pairing and mating several times before spawning and female snails laid eggs at night time. The main spawning season was from April to June with the mature proportion from 62.2 to 93.3%. The authors also found that stocking snails in the muddy bottom would be better than others and the survival rate varied from 88.15 to 90.93% after 28 days of cultured 7 period. Le Van Binh et al. (2013) investigated the effects of different foods on the growth and survival rate of Pila polita. The results showed that after 35 days of nursing period, the survival rate of snails in rice bran diet (94.4%) was no significant difference (P>0.05) from cassava powder (93.3%) or industrial pellet (93.7%). Highest increased biomass was obtained when feeding snails with pellet (2,027%) and significant difference compared to rice bran (727%) and cassava powder (992%). Feeding with pellet, snail also reached highest body weight and shell height (0.71g and 14.79mm) compared to feeding with rice bran (0.26g and 10.55mm) or cassava powder (0.36g and 11.65mm). Snails were fed with pellet also presented highest feed efficiency (723%) and that was significant difference compared to feeding with rice bran (473%) or cassava powder (529%). 2.5. Probiotics in aquaculture A widely accepted definition is taken from Fuller (Fuller R, 1987), who considered that a probiotic is a cultured product or live microbial feed supplement, which beneficially affects the host by improving its intestinal (microbial) balance. The important components of this definition reflect the need for a living microorganism and application to the host as a feed supplement (Sahu, 2008). Moriarty (1998) and Rengpipat et al. (1998) indicated that probiotics may prevent the luminous bacteria Vibrio species effectively. Intervention mechanism may be a combination of competition between bacteria and different antibiotic compounds by Bacillus spp created. According to some recent researches in aquaculture, the mechanism of probiotics can be divided into following aspects: (1) production of inhibitory compounds, (2) competition for nutrients maintenance, energy, shelter for harmful bacteria, (3) enhance the immune response, (4) improve water quality (Pham Thi Tuyet Ngan, 2010). Several studies have conducted to investigate the effects of probiotics on mollusc. Ngo Thi Thu Thao & Pham Thi Tuyet Ngan (2011) studied the effect of probiotics on larvae of Balylonia areolata. Results showed that treatment with probiotics 8 supplement, the survival and growth rate was higher than a control treatment. Ngo Thi Thu Thao et al. (2012) studied on the effect of different applied methods of the probiotics in juvenile of clam Meretrix lyrata. The results showed that supplementation of probiotics into algae biomass and added directly into the environment led to faster growth in weight and length compared to non-probiotics treatment (p<0.05). Results from these studies showed that the probiotics application has been effective in the nursery as well as in aquaculture. In recent years, the Ministry of Fisheries of Vietnam has allowed the application of microbial products, farmers have become familiar with the biological products and get good results. However, it should be a comprehensive assessment of economic efficiency and probiotic using methods. 9 CHAPTER III MATERIALS AND METHODS 3.1. Time and location Experiments were be carried out from May to December, 2013 at Department of Coastal Aquaculture, College of Aquaculture and Fisheries, Can Tho University. 3.2. Materials 3.2.1. Equipment Tanks: 24 square tanks (60cm × 60cm); Electronic balance (0.01g readability), vernier caliper, thermometer, aeration; pH, NH3/NH4+, NO2-, and alkalinity test kits; subtracts (water lettuce, Pistia stratiotes) 3.2.2. Feeds Pellet feed (18% protein), fine rice bran. 3.2.3. Probiotics Biosubtyl DL: contain Bacillus subtilus and Lactobacillus acidophilus. Biosubtyl-II: contain Bacillus subtilus. 10 Figure 3. 1 Biosubtyl DL and Biosubtyl-II 3.2.4. Water resource Fresh water is taken from fish ponds at College of Aquaculture and Fisheries, Can Tho University. Water is pumped to the settling tanks for 2-3 days and then filtered through a plankton net (50 µm mesh size) into the incubation tank. 3.2.5. Incubated feed making proceduce Firstly, well mix 1kg fine bran (pellet feed) and 5 Biosubtyl-II packages together, then put 500gr molasses and little amount of water to dissolve the sugar and create adhesion for the mixture. After the mixture is mixed well, exposing the mixture at room temperature until completely dry. Finally, crushed and filtered through a mesh to a fine powder (Figure 3.2, 3.3). 11
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