CHAPTER
1

INTRODUCTION

1.1       Background of Study

           The
Mud crab from the genus Scylla  (Forsskal, 1775) are the largest edible
crab that can be found from South Africa, Coast of Indian Ocean, Malay
Archipelago, south of Japan all the way to eastern Australia. Mud crabs can be found
in estuaries and mangrove areas. They are highly tolerant of variations in
water salinity and temperature. Juvenile and adult mud crabs occupies burrows
in the Intertidal zone. They occupy just below the low tide mark, where they would
bury themselves in the mud during the day. Mating occur in estuaries. Mature Scylla sp. migrates offshore to spawn. Spawned eggs are attached to the
pleopod hairs of the abdominal flap.

Biofloc are aggregates
(flocs) of algae, bacteria, protozoans, organic matter such as faeces and
uneaten food. These flocs are held together in a mucus that is secreted by
bacteria, bounded together by filamentous microorganisms or held together by an
electrostatic attraction. Secondly, these biofloc systems are developed to
prevent the introduction of foreign disease to an aquaculture farm from
incoming water. Biofloc are also used as a water treatment system for water
improve environmental control over production and provides nutrition to
cultured animals. There are two basic biofloc system, which are ‘green water’
and ‘brown water’. The green water are composed of both algal and bacterial
compound while brown water is composed of only bacterial compound. Biofloc
systems are normally used on suitable culture species which can tolerate high
solid concentrations, poor water quality and able to consume and digest
biofloc.

1.2     Problem statement

         According to the Food and Agriculture
Organization of the United Nations, under the section of Fisheries and
Aquaculture Department (FAO), the main issues that are faced all over the world
is the lack of seed stock for expanding aquaculture industry. Development of
mud crab aquaculture has started for some time but not much success has been
achieved due to mass mortality of crab seed especially during the zoea and
megalopa stage. Therefore, it is necessary to identify the certain factors
affecting survival and development of mud crab seed (Kasetsart J, 2002).

The over-exploitation
of wild crab stocks, destruction of mangrove swamps and poor quality of wild
production to support the increasing demand are the factors that have driven
the development of hatchery technology for mud crabs (Lindner, 2005). The
supply of of crab seed stocks from the wild will varies over time as
recruitment to the fishery is seasonal (Walton et al., 2006), as reflected in
the variation of zoeal abundance in near shore waters (Sara et al., 2006).

Threats to mud crab and
seed supply alike includes algal blooms, industrial and urban run-off, and the
over-exploitation of wild adult mud crabs. Furthermore, supplies for mud crab
seeds can also be limited by the number of crabbers targeting certain species
of mud crabs (Say and Ikhwanuddin, 1999). The need for a reliable supply of mud
crab seed stock year-round to support farm expansion is essential and will
boost the future significance of mud crab hatcheries.

1.3     Significance of Study

         Due to the fast growth of human
population, the more effective way must be developed to increase the yield of
aquaculture to fulfil the demand of the aquatic product.  Now, the aquaculture sector were developed
and became more important so that more effective ways must be searched to make
sure that the sector run parallel to the increasing of world’s human
population. Scylla sp. is now farmed
in the Indo-West Pacific region, which covers from East and South Africa to the
southeast and east Asia, and  Northeast
of Australia (FAO, 2001). Due to the main issues of low crab seed stocks, the
application of biofloc in Scylla sp.
hatching tanks is highly beneficial if it increases the survival rate and
decrease the rate of mortality in larval zoea stage. Thus, the amount of yield
that will be harvested can be increased.

1.4      Objectives

          The main objectives of this project is
to apply Biofloc agents into Scylla sp. hatching tanks and examine the effects
of biofloc on the growth and survival rates of Scylla sp. hatchlings
(Zoea to juvenile stage).

CHAPTER
2

LITERATURE
REVIEW

2.1      Biology of Mud crab

2.1.1   Taxanomy status

           The taxanomy status of Scylla paramamosain are as follows;

Kingdom:                                         Animalia

Phylum:                                            Arthopoda

Subphylum:                                      Crustacea

Class:                                               
Malacostraca

Subclass:                                         
Eumalacostraca Grobben

Superorder:                                      Pleocyemata
Burkenroad

Order:                                               Decapoda

Suborder:                                         
Pleocyemata

Infraorder:                                       
Brachyura

Superfamily:                                     Portunoidea

Family:                                             Portunidae

Subfamily:                                       
Portuninae

Genus:                                              Scylla

Species:                                            Scylla paramamosain

Common names:                              Mud crab,
Mangrove crab,

                                                                    
Ketam nipah (Malaysia)

Figure
2.1 : Scylla paramamosain (Estampador,
1949)

2.1.2   Morphology

            Scylla
paramamosain hatchlings called zoea
is about one millimetre long with undeveloped limbs. It has a dorsal spine,
rostral spine and carapace. The antennules or first antenna is an unsegmented
rod. The second antenna has two apical setae, one is long and the other is
short. The first maxilliped has no spines, the basipodite has two plumose setae
on the outer margin and has five segments. The second maxilliped has one setae
on the inner margin and four setae along the inner edge of the basipodite. The
endopodite has three segments. The telson consists of two points and is forked
shape. Has three setae on each side. The abdomen has five segments. The third
and fourth has a pair of short lateral spines.

Figure
2.1.2(a) : External feature of Scylla sp.
zoea

2.1.3   Water Quality Parameter

          The optimum range of water quality
parameters for production of seed of Scylla
paramamosain are given below:

·        
Water
temperature                           – 28-31°C

·        
Salinity                                            –
26-31 ppt

·        
pH                                                    –
7-8

·        
Dissolved oxygen                            – >5 ppm

·        
Nitrite                                              –
< 0.02 ppm (Source: International Journal of Fisheries and Aquatic Studies, 2014)   2.1.4   Life Cycle             The life cycle of Scylla sp. involves six distinct phases, the egg, zoea, megalopa, young adult and adult. As juveniles, the female and male crabs are difficult to be differentiated. Adult female and male crabs can be differentiated by observing the abdominal flap. An adult female crab have a larger and broader abdominal flap compared to the male crab.           Mating occurs about four days and happens only when the shell of the adult female crab has been shed. The adult male crab can sense when the female crab is about to moult. The male crab will climb on top by clasping using it's legs. Once the female crab has shed its shell, the male will turn the female crab upside down. The male will then transfer its sperms into the female's oviducts. The sperm of the male can be kept for a few months while waiting for the eggs to ripen for fertilization. Egg incubation takes about ten to seventeen days, depending on the temperature of water (Ong, 1996; DuPlessis, 1971; Arriola, 1940). Mud crab eggs are orange in colour when ripe. After fertilized, the eggs will turn to brownish black in colour when it is about to hatch. A single batch of eggs may contain from one to two million eggs. Although the number of hatchlings are high, the mortality rate are high as well.           Scylla sp. starts as a zoea (hatchling) after hatching from an egg. In three weeks, a zoea will undergo moulting stages up to five times (stage I-V; Delathiere, 1990). The length of the first zoeal stage is about 1 millimetre long and has undeveloped limbs. A zoea will float along with planktons in the sea. The survival rate and growth development strongly depends on water temperature and salinity (Hill, 1974) At the fifth moult, it will turn into a megalopa. A megalopa has a pair of functional claws that are used for feeding and grasping. During this stage, cannibalism normally occurs as the bigger megalopa will try to devour smaller megalopae. Addition to that, a megalopa will move inshore. Within ten days, the megalopa will moult and turn into a juvenile crab. Juvenile crabs has fully functional limbs and claws.   2.2     Biofloc technology          Bioflocs technology is the use of microbial components to metabolize organic residues in water. It consists of flocs of algae, protozoans, bacterias, and other types of particulate organic matter (feces and uneaten feed) that are held together by filamentous microorganisms. Biofloc is one of the innovative methods for waste management and nutrient retention that offers a possible solution to solve environmental problems in aquaculture. Application of biofloc also offers benefits in improving aquaculture productions in aquaculture which proved nutritious food source and able to improve feed utilization efficiency. Equally important, biofloc can significantly reduce the quantity of water used in an aquaculture farm. The consumption of biofloc by cultured animals has shown positive results on growth rate. Growth enhancements has been attributed to both bacterial and algae nutritional components, which up to 30% of conventional feeding ration can be lowered due to consumption of biofloc (Emerencianno et al., 2013).   CHAPTER 3     MATERIALS AND METHODS     3.1     Location of experiment             This experiment will be run at the marine hatchery of Institute of Tropical Aquaculture (AKUATROP), University Malaysia Terengganu (UMT).   3.2      Source of female broodstock           The female broodstock will be obtained from local fisherman at Setiu wetlands. They catch it from estuarine and mangrove swamps.   3.3     Experimental design and Tank setup            Tank preparation starts from preparing 18 units of 500L tanks with total 5 triplicate treatments and control tanks (without biofloc). The inoculum will be injected into biofloc of treatment tanks and add in molasses, for helping rapid floc formation. The inoculum being chosen is Bacillus species. Approximately after 2 weeks, newly hatched larvae will be placed in all tanks with gentle aeration. During this time, the volume of biofloc will be monitored and measured by using Imhoff cone. The water samples collected every 2 days to undergo water quality tests and analysis. The data collected then undergo ANOVA to compare the significant differences between water quality parameters among treatments.       Replication 1 2 3 Control     2 mL/ L 4 mL/ L   All tanks rearing larvae and feeding with rotifers (60 mL-1) daily 6 mL/ L 8 mL/ L 10 mL/ L                                Figure 3.3 : Experimental design and tank setup                                                           .   3.4     Biofloc formation and harvest             The treatments, in triplicate, consisted in five volumes of biofloc in the water: 2, 4, 6, 8 and 10 mL/L. Whereas, the source of bioflocs will be collected from AQUATROP hatchery in UMT. Inject inoculum with Bacillus sp. into biofloc for helping rapid floc formation. The volume of biofloc in the experimental tanks will be monitoring for 14 days interval?so that the required biofloc volume in each treatment can be maintained.   3.4.1   Biofloc measurement and monitoring Biofloc volume will be monitored with experiment, the floc volume is measured every week by using Imhoff cone. Imhoff cone used to determine settleable solids and estimate biofloc volume (Hargreaves, 2013). Pouring 1 litre of water taking from each tank into Imhoff cones and settling the biofloc particles for 1 hour then record the measured data. As the biofloc continuously forming, excess volume of biofloc should be sludge by siphoning out from the treatment tanks (Chaignon et al., 2002).               3.5     Stocking of larvae and tank management               The mud crab larvae will stock to a density of 80 larvae L-1 in 18 units of 500L tanks (Nghia et al., 2007). The larvae will be observed start from zoeal stage 1 until first moult of juvenile, and this approximately will take 28 days. 70% of tank will be filled with brackish water whereas the remaining are substrates. Rotifers (Branchionus sp.) are added daily with density 60 mL-1 as feed for larval developmet (Nghia et al., 2007).       3.6     Feeding regime              The hatchlings will be fed with newly hatched Artemia once a day in the morning. Artemia will be used through the zoeal stage up to the juvenile stage. The hatchlings are fed with the rate of 10-15 nauplii of Artemia.   3.7     Water quality parameter             In this project, sea water will be used in all tanks. The use of sea water should be properly set up for the broodstock and hatchlings; salinity of 26-31 ppt, optimum temperatures of 28-31°C, pH value of 7-8, dissolved oxygen of >5 ppm
and nitrate level <0.02 ppm. The water parameters will be measured regularly by using YSI meter.   3.8      Survival rate              The survival rate of the hatchlings will be obtained at the end of the project. The number of survived hatchlings that reaches to juvenile stage will be counted manually and carefully. The counting process will be a total of three times and average number will be calculated.   3.9      Larvae stage index (LSI)              During the culture period, the larvae will undergo 8 stages before morphing into juvenile stage. The stages of the larvae will be noted with Larvae Stage Index (LSI). To collect sample, the aeration must be removed a while from the water to make sure the larvae is evenly distributed to the entire tank. Then, the larvae are collected randomly; on the surface, on bottom, and middle of the water. Manzi et al. (1977) shows that the LSI is calculated as follows;   LSI = Where,  = Larvae stage  = Number of larvae in Stage S1 n   = Total number of larvae observed   3.10      Data collection             During the project, the  data on water quality parameter, growth development and survival rate of the hatchlings will be recorded and determined.                   CHAPTER 4     EXPECTED RESULTS     4.0      Expected results              With the application of Biofloc in the culture tanks, the mud crab broodstock will release the eggs earlier compared to broodstock that is not applied with biofloc. The eggs from the tank that is applied with biofloc will also hatch earlier compared to the tank that is not applied. As for the hatchling; the survival rate is expected to increase and growth rate will be enhanced when biofloc is applied.