evaluate potential use of gut weed (enteromorpha sp.) as a food source for tilapia (oreochromis niloticus): affect on survival and growth

The results in the first experiment showed that growth performance of the experimental fish fed 1 day pellet feed_1 day dried gut weed was comparable to the group fed single pellet feed.

CAN THO UNIVERSITY COLLEGE OF AQUACULTURE AND FISHERIES

EVALUATE POTENTIAL USE OF GUT WEED (Enteromorpha sp.) AS A FOOD SOURCE FOR TILAPIA (Oreochromis niloticus):

AFFECT ON SURVIVAL AND GROWTH

By DAM PHUOC HIEN

A thesis submitted in partial fulfillment of the requirements for

The degree of Bachelor of Aquaculture

Can Tho City, December 2012

CAN THO UNIVERSITY COLLEGE OF AQUACULTURE AND FISHERIES

EVALUATE POTENTIAL USE OF GUT WEED (Enteromorpha sp.) AS A FOOD SOURCE FOR TILAPIA (Oreochromis niloticus):

AFFECT ON SURVIVAL AND GROWTH

By DAM PHUOC HIEN

A thesis submitted in partial fulfillment of the requirements for

The degree of Bachelor of Aquaculture

Promoter

Dr NGUYEN THI NGOC ANH

Can Tho City, December 2012

I would like to express my deep gratitude to my promoter Dr Nguyen Thi Ngoc Anh for constant guidance and enthusiastic help during conducting experiment and her patience in correcting thesis

Special acknowledgements to teachers of College of Aquaculture and Fisheries, Can Tho University have taught me the experiences during study

I especially thanks to my classmates from Advanced Aquaculture course 34 and Aquaculture course 35

Always facilitating and enthusiastically helping me complete the thesis

of 3.04g and 0.33 g in the first and second experiment, respectively Tilapia were stocked in the 120L-composite tanks for the first experiment and in the 70L-plastic tanks for the second experiment at a density of 30 fish per tank and in seawater at salinity of 5 ppt Fish were fed to satiation twice a day for 42 days At the end of the feeding trials, survival of experimental fish in two experiments was not affected by the feeding treatments ranging from 82.0 to 84.4% and 100% for the first and the second experiments, respectively The results in the first experiment showed that growth performance of the experimental fish fed 1 day pellet feed_1 day dried gut weed was comparable to the group fed single pellet feed Moreover, using gut weed as direct feed can maintain better water quality and reduce the feed costs The results of the second experiment showed that mean final body weight and specific growth rate and Daily weight gain increased with increasing gut weed protein level in the fish diet up to 20% Increasing the level of gut weed protein in the diet from 30 to 40% did not exert any additional advantage in terms of growth of fish and an inclusion level of 50% led to negative effects on growth Based on the present study, it is concluded that gut weed has

a great potential as alternative ingredients in diets and inclusion level up to 40% have no adverse effect on growth, feed utilization

TABLE OF CONTENTS Contents Page

ACKOWLEDGE i

ABSTRACT ii

TABLE OF CONTENTS iii

LIST OF TABLES vii

LIST OF FIGURES viii

LIST OF ABBREVIATIONS viiii

Chapter 1 1

INTRODUCTION 1

1.1 Introduction 1

1.2 Research objectives 2

1.3 Research contents 2

Chapter 2 3

LITERATURE REVIEW 3

2.1 Gut weed 3

2.1.1 Classification 3

2.1.2 Morphology of Enteromorpha sp 3

2.1.3 Distribution 4

2.1.4 Nutritional value of gut weed Enteromorpha 4

2.1.5 Use of gut weed in aquaculture 6

2.2 Tilapia 7

2.2.1 Production of Tilapia in the world and in Vietnam 7

2.2.2 Classification and habitat 8

2.2.3 Nutritional requirement 9

Chapter 3 11

MATERIALS AND METHODS 11

3.1 Study subject 11

3.2 Materials and chemicals 11

3.3 Research methodologies 11

3.3.1 Experiment 1 Evaluating the effect of dried gut weed (Enteromorpha sp.) as direct food to replace pellet feed for feeding fingerling Tilapia 11

3.3.2 Experiment 2: Evaluating the effect of fishmeal replacement with gut weed (Enteromorpha sp.) as a protein source in practical diets for Tilapia 15

Chapter 4 20

RESULTS AND DISCUSSION 20

4.1 Experiment 1: Evaluating the effect of dried gut weed (Enteromorpha sp.) as direct food to replace pellet feed for feeding fingerling Tilapia 20

4.2 Effects of using dried gut weed (Enteromorpha sp.) as direct food to replace pellet feed on Tilapia performance 21

4.2.1 Survival 21

4.2.2 Growth performance 22

4.2.3 Feed conversion ratio (FCR) 24

4.2.4 Cost reduction of replacing gut weed Enteromorpha sp in diets 25

4.3 Experiment 2: Effect of protein fishmeal replacement with gut weed protein on Tilapia performance 26

4.3.1 Water quality 26

4.3.2 Survival and Growth performance 27

4.3.3 Feed conversion ratio (FCR) 29

4.3.4 Economic comparison of feed cost 30

Chapter 5 31

CONCLUSION AND RECOMMENDATION 31

5.1 Conclusion 31

5.2 Recommendation 31

REFERENCES 32

LIST OF TABLES

Table 1: Proximate composition (% dry matter) of the experimental diets 13Table 2: Proximate composition of ingredients used for experimental diets (% dry matter)………17Table 3: Ingredients and proximate analysis of the experimental diets (% dry matter) 18Table 4: Average water temperature and pH during 42 days in experiment 1 20Table 5: Average concentration of TAN and NO2in experiment 1 21Table 6: Mean initial weight, mean final weight, weight gain, specific growth rate (SGR) and daily weight gain (DWG) of Tilapia in experiment 1 23Table 7: Feed conversion ratio in the Experiment 1 24Table 8: The expenditure of commercial feed combined feed with dried gut weeds to replace in diets of tilapia in the Experiment 1 25Table 9: Temperature and pH during 42 days of experiment 2 26Table 10: Total Ammonia Nitrogen (TAN) and Nitrite (NO2) during 42 days of Experiment 2 27Table 11: Survival and growth performance of Tilapia fry fed different diets over 42 days……… 28Table 12: Feed cost for Tilapia growth in the Experiment 2 30

LIST OF FIGURES

Figure 1: Morphology of Enteromorpha species 3

Figure 2: Experimental system 12

Figure 3: Experimental system 16

Figure 4: Survival rate of Tilapia fed the first experiment diets after 42 days 22

Figure 5: Feed conversion ratio in the Experiment 2 29

LIST OF ABBREVIATIONS

SGR Specific growth rate

FCR Feed conversion ratio

Chapter 1 INTRODUCTION 1.1 Introduction

In aquaculture, feed cost is a highest proportion and it accounts for more than 50% of the total production costs (Tran Thi Thanh Hien and Nguyen Anh Tuan, 2009) In addition, most feed manufactures are using expensive imported fishmeal as a protein source for aqua feeds resulting in high price Therefore, assessment of cheaper or more readily available alternative plant protein sources such as soybean meal, seaweed meal

or other sources that may reduce the use of fishmeal in feeds is necessary (FAO, 2011) Among alternative plant protein sources, seaweeds may be considered as a suitable ingredient for replacing fishmeal in fish and shrimp diets because of its high nutritional value (FAO, 2003; Dhargalkar and Pereira, 2005)

Like other seaweeds, gut weed Enteromorpha sp has a high nutritional value; it contains

9–14% protein; 2–3.6% lipid; 32–36% ash, and n-3 and n-6 fatty acids 10.4 and 10.9 g/100 g of total fatty acid, respectively; the protein of this seaweed has a high

digestibility up to 98% (Fleurence, 1999; Aguilera-Morales, et al., 2005) Hence, gut

weed can be used as a direct feed or as ingredient in diets for fish and shrimp Recent

investigates revealed that gut weed (Enteromorpha sp.) belonging to green algae

distribute abundantly in the extensive shrimp farms and other brackish water bodies of the Mekong delta (SUDA, 2009) This indicates large quantity of gut weed is available for aquaculture feeds So far, in Vietnam study on the utilization of gut weed for fish has not been greatly considered

In the other hand,one of the great advantages of Tilapia for aquaculture is that they feed

on a low trophic level The members of the genus Oreochromis are all omnivores,

feeding on algae, aquatic plants, small invertebrates, detritus material (KevinFitzsimmons, 2009) In addition, tilapia can be applied in poly-culture with other species

or integrated aquaculture system They have a good alternative to culture conditions, are

species of high economic and can live in fresh and brackish water (Sayed, et al., 2006).

Consequently, investigating the potential use of gut weed (Enteromorpha sp.) as feed for

Tilapia was implemented

1.2 Research objectives

Determine suitable percentage of fish meal protein replaced by gut weed protein

(Enteromorpha sp.) in a practical diet and gut weed as direct feed to substitute a

commercial feed for Tilapia fingerlings This study could encourage farmers using local availability of gut weed as food source for tilapia or other fish in the Mekong delta and itcould contribute to reduce feed costs and improve profits for farmers

Chapter 2 LITERATURE REVIEW 2.1 Gut weed

Species within the genus Enteromorpha are very difficult to identify as differences

between species are small and hard to spot (Budd & Pizzola, 2002)

Figure 1: Morphology of Enteromorpha species

They are green seaweeds, with tubular and elongate fronds that may be branched, flattened or inflated (Kirby, 2001) They are bright green in color and may occasionally

be bleached white, particularly around rock pools They attach to the substrate by means

of a minute disc-like holdfast (Gibson, et al., 2001) The fronds of a species may vary in

appearance due to changes in environmental conditions, which further confuses identification, and microscopic examination of cell details is often required to identify a species with certainty (Budd, & Pizzola, 2002)

2.1.3 Distribution

Enteromorpha species are distributed throughout the world, in a wide variety of

environments They can tolerate different salinities ranging from freshwater to seawater and can be found in salt streams They can grow on the ocean coast, in the brackish and

fresh water inland Enteromorpha can also grow on many types of substrate: sand, mud

or rock, even wood, concrete or metal type or free development without substrates

Enteromorpha is also able to grow throughout the intertidal littoral plane It can often be

found growing with Ulva, it also grow with other algae, in a variety of different habitats

(Kirby, 2001)

In Vietnam, according to Nguyen Van Tien (2007), gut weed commonly distribute instatic brackish water bodies from Hai Phong, Nam Dinh, Nghe An, Ninh Thuan, Binh Thuan, Khanh Hoa, Thanh Hoa, Kien Giang provinces Other study revealed that in the

Mekong delta gut weed (Enteromorpha spp.) was found abundantly in the extensive

farms, abandoned ponds, discharge canals and rice field from Bac Lieu, Soc Trang, Ca Mau and Ben Tre provinces (ITB-Vietnam, 2011) Similar investigation was reported by Nguyen Minh Tien (2012)

2.1.4 Nutritional value of gut weed Enteromorpha

Several studies confirmed that the nutritional value of seaweed varied depending on species, developmental stages, seasonal change and geographic regions and were affected by the environmental factors such as salinity, temperature and nutrient contents

in the habitats ((Haroon et al., 2000; Banerjee et al., 2009; Nguyen Minh Tien, 2012).

Haroon et al (2000) investigated the biochemical composition of Enteromorpha spp

from the Gulf of Gdańsk coast on the southern Baltic Sea Authors concluded that the

nutritional value of Enteromorpha spp is higher during spring and autumn (high protein

and lipid content) than during summer Carbohydrate contributes the relatively largest proportion (29.09-39.81%) of the biochemical composition and its values vary considerably with sampling station and season; followed by protein (9.42-20.60%) and

lipid (3.47-4.36%) According to Aguilera-Morales et al (2005), chemical analysis indicated that Enteromorpha spp has 9–14% protein; 2–3.6% ether extract; 32–36%

ash, and n-3 and n-6 fatty acids 10.4 and 10.9 g/100 g of total fatty acid, respectively The protein of this seaweed has a high digestibility (98%)

The findings of Banerjee et al (2009) on biochemical composition of three kinds of seaweeds Ulva lactuca, Enteromorpha intestinalis, and Catenella repens Indian river,

they reported that the results of biochemical composition of these seaweeds seem to be strongly affected by ambient hydrological parameters (surface water salinity, temperature and nitrate content) in the present geographical locale The protein content

of E intestinalis varied in the range of 5.18-13.84%; lipid: 0.07-0.30%, carbonhydrate

33.53-57.03% and astaxanthin: 112.72-186.11ppm

According to Nguyen Minh Tien (2012), the nutritional composition of gut weeds

Enteromorpha sp from Bac Lieu and Soc Trang and provinces have marked changes in

different growth stages in which the nutritional values of young and adult stages were significantly higher compared to that of the senescent gut weed Furthermore, the proximate composition (protein, lipid, ash, fiber and carbohydrates) and amino acid compositions of gut weed varied with the season and strongly influenced by salinity and

temperature Author suggested that gut weeds (Enteromorpha sp.) in the brackish water

bodies of the Mekong delta have high nutritional values; they can be used as feeds for aquaculture species or applied in the integrated aquaculture system

2.1.5 Use of gut weed in aquaculture

Previous studies revealed that gut weed Enteromorpha spp can be used as a food source for fish (Yousif, et al., 2004; Asino et al., 2010; Nguyen Thi Ty Ni, 2012)

El Sayed (1999), evaluated seaweeds as food source for Tilapia, author revealed that tilapia is able to absorb 70-80% of green algae that some fish species are unlikely to digest

Yousif, et al (2004) evaluated the dehydrated Enteromorpha incorporated at a rate of 0 (control), 10, 20 and 30% in the diets for rabbitfish, Siganus canaliculatus (Park) They

found that the survival, growth performance and feed utilization efficiency of experimental fish were observed to decrease with increasing inclusion levels of dehydrated algae The best results of all parameters were achieved by fish fed control

diet and fresh Enteromorpha Carcass protein was not affected by the different

treatments while lipid content was observed to increase in the group of fish

supplemented with fresh Enteromorpha.

Asino et al (2010) assessed the diets contain graded levels (0%,5%,10% and 15%) of

Enteromorpha prolifera for feeding juvenile large yellow croaker (Pseudosciaena crocea), they found that When the supplementation of E prolifera was >5%, the specific

growth rate (SGR) and feed efficiency ratio (FER) were significantly higher compared with the control group (0%) Based on SGR and FER, author suggested that

supplementation levels of E prolifera can reach at least 15% without affecting the

growth and still maintain a high survival rate for this fish species

Swain and Padhi (2011), evaluated the nutritional value of four seaweeds such as

Enteromorpha intestinalis, Grateloupia filicina, Gracilaria verrucosa and Polysiphonia sertularioides which were added at various induction levels in the

diets fed to juvenile Rohu (Labeo rohita) and Mrigal (Cirrihinus mrigala ) fingerlings The result showed that these fish fed Enteromorpha intestinalis included at a

rate of 30% in the experimental diet had similar growth performance compared to those fed three other seaweed substitution and all seaweed diets was better than the fish

received the control diet Author also concluded the difference may be attributed not only to the additional protein content, but to the value addition in terms of certain indispensable amino acid and fatty acids

Nguyen Thi Ty Ni (2012) assessed the potential use of gut weed (Enteromorpha sp.) as

a feed for spotted scat (Scatophagus argus) The results indicated that survival of

experimental fish was not affected by the feeding treatments Growth rate and feed efficiency of spotted scat fed the diets replaced from 10% to 40% of fishmeal protein by gut weed protein had no significant differences compared to the control diet Moreover, growth performance of the experimental fish fed solely fresh gut weed was comparable

to the group fed single pellet feed (control) and its protein efficiency ratio was highest and significantly different from the control group The proximate composition of fish carcass after feeding trials did not change except the lipid content was significantly higher in fish receiving the control than in other fish groups

Similar findings was reported by Nguyen Thi Ngoc Anh et al (2012), mudskipper (Pseudapocryptes elongatus) fingerlings fed the diets with fishmeal protein replaced by

increasing dietary levels of gut weed protein 10%, 20%, 30%, 40% and 50% Survival of experimental fish was similar among treatment When comparing the control diet with the treatments of 10%, 20%, 30% and 40% gut weed protein replacement, no significant differences were observed for growth performances and feed efficiency Particularly, fish group received the 10% gut weed protein substitution showed slightly higher growth than those in the control treatment The proximate composition (water content, protein, ash and phosphorus) of fish carcass in all treatments was similar except the lipid and calcium levels in the treatments of 40 and 50% substitution were significantly lower compared to the control treatment

2.2 Tilapia

2.2.1 Production of Tilapia in the world and in Vietnam

Tilapia is one of the most widely cultured fish in the world such as China, Taiwan, Brazil, Africa, and Costa Rica… Currently, farmed tilapia represents more than 75% of

world tilapia production, and this contribution has been exponentially growing in recent years (FAO, 2009) According to FAO (2010), world tilapia production has been booming during the last decade, with output doubling from 830000 tones in 1990 to 1.6 million tones in 1999 and to 3.5 million tones in 2008 Asia represents about three quarter of world tilapia production China is by far the main tilapia producing country, alone produces 1.2 million, while the rest of Asia contributes 0.9 million tones Total

tilapia production is mainly Nile tilapia (Oreochromis niloticus) All new countries

entering tilapia production concentrate on this species, which is easy to grow In 2008,about three quarters of world Tilapia production were Nile Tilapia

Production of Tilapia in Vietnam has been increasing year by year; the farming area has been expanded In 2009, the area of tilapia reached 29,717 ha, production reached 50 thousand tons, were cultured in brackish water and fresh water with farming forms extensive and semi-intensive Tilapia products for domestic consumption are 95-98%, while exports reached only 869 tones with a value reaching U.S $ 1.9 millions According to the 2015 planning of Ministry of Agriculture and Rural Development, the total output of Tilapia is 300-350 thousand tons, 60 thousand hectares of Tilapia farming, and 30% with export value reaching 100 millions (website:

http://www.rial.org)

2.2.2 Classification and habitat

Tilapia Oreochromis niloticus (Linnaeus, 1758) are classified as follows:

According to Doan Khac Do (2008), Tilapia is euryhaline species; they can live in freshwater, brackish and saline water They are highly tolerant of low dissolved oxygen concentration (below 1 mg/L) even down to 0.1 mg/L but optimum growth is obtained at levels greater than 3 mg/L Tilapia can live at pH range of 5-10 and suitable pH range for their growth is 7,0 -8.5 Optimal temperature for the growth of Tilapia is 25-32°C, when temperature is below 20°C, fish reduce or stop eating and the fish will die when the temperature is less than 11°C (Harrison, 2006) Other study also confirm that Tilapia can tolerate a wider range of environmental conditions including factors such as salinity, dissolved oxygen, temperature, pH, and ammonia levels than most cultured freshwater fishes can In general, most tilapia is highly tolerant of saline waters, although salinity

tolerance differs among species (Mjoun, et al., 2010).

and animal protein sources (Medri et al., 2000: Mjoun, et al., 2010).

The protein requirement of Tilapia decreases with age and size with higher dietary crude protein concentrations required for fry (30–56%) and juvenile (30–40%) but lower protein levels (28–30%) for larger Tilapia (Winfree and Stickney, 1981; Twibell and

Brown, 1998) Tilapia (O aureus) at weight of 0.3-0.5g with protein source from soy

flour or 36% fish meal diet (Davis and Stickney, 1978) The juveniles (individual weight

of 2.5g) of monosexual Tilapia (Oreochromis niloticus) have optimal protein diet is 30%

with protein supplied mainly soybean meal, rice bran, and wheat flour (Bahnasawy, 2009) Tilapia species require 10 essential amino acids that need to be supplied by the diet Essential amino acid requirements can be met by the use of a balance of both plant and animal proteins, and if necessary, by the inclusion of synthetic amino acids in the

complete feed (Mjoun, et al., 2010).

Winfree and Stickney (1981) revealed that for tilapia up to 2.5 g, the optimum dietary lipid concentration was 5.2%, decreasing to 4.4% for fish up to 7.5 g Additianally, Jauncey (2000) suggested that to maximize protein utilization, dietary fat contents should be between 8 and 12% for tilapia up to 25 g, and 6 to 8% for larger fish As with most fish, tilapia seems to have a requirement for n-6 (linoleic) fatty acids, and to a lesser extent, a requirement for n-3 (linolenic) fatty acids Dietary lipids should supply at

least 1% of n-6 fatty acids (Teshima et al., 1982)

Tilapia can effectively utilize carbohydrate levels up to 30 to 40% in the diet, which is

considerably more than most cultured fish (Anderson, et al., 1984; Teshima, et al.,

1985) Fiber is usually considered indigestible, as tilapia do not have the required enzymes for fiber digestion Maximum growth for Tilapia, crude fiber levels in diets

should probably not exceed 5% (Anderson et al., 1984).

Chapter 3 MATERIALS AND METHODS 3.1 Study subject

1 Tilapia (Oreochromis niloticus)

2 Gut weeds (Enteromorpha species)

3.2 Materials and chemicals

-Refract meter, temperature and pH meter, electronic scale, pumps, aerator, fridge, aeration pipe, plastic tanks…

-Chemicals used in water treatment: chlorine, iodine, natrithiosulfat, test kits

3.3 Research methodologies

3.3.1 Experiment 1 Evaluating the effect of dried gut weed (Enteromorpha sp.) as

direct food to replace pellet feed for feeding fingerling Tilapia

Experimental setup

The feeding trial was performed in the experimental hatchery of the College of Aquaculture and Fisheries, Can Tho University The dried gut weed was used as a direct feed to replace commercial feed for feeding Tilapia juvenile Everyday fish was fed only one kind of feed namely either commercial feed or dried gut weed Four treatments in triplicate were conducted as follows (Fig 2):

- Treatment 1 CF: commercial feed (control)

- Treatment 2 DGW: dried gut weed

- Treatment 3 CF_1DGW: 1 day commercial feed_1 day dried gut weed

- Treatment 4 CF_2DGW: 1 day commercial feed_2 day dried gut weed

Figure 2: Experimental system

Culture conditions

- Tank volume: 120L composite tank

- Culture system: open clear water system

- Fish density: 30 fish per tank

- Salinity in culture tank: 5 ppt

- Aeration: Each tank was provided with slightly continuous aeration

- Feeding rate: up to satiation (10-15% body weight per day)

- Feeding frequency: 2 times per day at 8:00 am and 4:00 pm

- Experimental period: 6 weeks

Experimental fish

-Source of fish: Fingerlings of Tilapia were purchased from a reliable hatchery in Can Tho city, and visually checked for signs of disease and parasites Before starting the feeding trial, fish were reared in a 1-m³ tank for 1 week in order to acclimate to the laboratory conditions and to get them acquainted with the feeding trays

-Size of fish: Fingerlings with initial individual weight of 3.04±0.48 g were stocked in each tank