I A

PROPOSAL IDENTIFICATION Research Title

Sustainable mud crab (Scylla serrata) fishery in Peninsular Malaysia with respect to size at maturity, genetic diversity and population structure.

B Key Words Mud crab, Scylla serrata, genetic diversity, maturity size and population structure. C Student

Noor Mutasha Biha Binti Mohd Din D Degree and Institution

Master of Science (Genetic), Universiti Kebangsaan Malaysia E Supervisor

Dr Yosni Bakar, UKM. F Summary

The serrated swimming crab, Scylla serrata, is the dominant species of mud crab in Malaysia. It is belonging to the family of swimming crabs (Portunidae). It is widespread throughout the Indo-West Pacific (IWP) and typically inhabit the estuaries, mangrove swamps and sheltered bays. It has been targeted for aquaculture industry due to its rapid growth rate, high market prices and form an important source of income for coastal fishing communities. Despite its importance, no great effort has been offered to examine and overcome the impact of harvesting the wild populations. The high demand for S.serrata crab coupled with the absence of any regulations leads to overharvesting of the resource by excessive removal of individuals before they reach maturity. The premium price for mature females with ripe eggs aggravates the problem further. The long-term implication are serious: increased harvest mortality is expected to lower productivity by favoring maturation at younger ages and smaller sizes, reduces reproductive potential, and poses threat to genetic diversity. Therefore, the conservation and management of S.serrata such as prohibiting the capture of immature crab should be done for sustainable utilization of mud crab resources. To maintain the phenotypic and genetic diversity between and within stocks is a critical aspect of conservation and sustainable of management of S.serrata fishery. However, there are significant voids in scientific knowledge on the size at maturity, genetic

diversity, and population structure of S.serrata populations throughout its habitat range in Peninsular Malaysia. Views on population structure of marine species are changing rapidly which was unrecognized microgeographical structure are possible to detect with the advent of polymorphic DNA markers such as microsatellite. This study are aiming to determine levels of genetic diversity to form genetic baseline of wild S.serrata populations, estimate the size at maturity and access its relationship to level of genetic diversity in S.serrata populations, and map the extent of genetically distinct S.serrata populations in Peninsular Malaysia. A panel of ten microsatellite markers that were developed specifically for Scylla species will be utilized to provide sufficient resolution to characterize S.serrata populations. The result gained will provide the scientific information for management recommendations and measures can address the need to achieve sustainable resource continuity while balancing social and economic pressures.

II

OBJECTIVES OF THE RESEARCH

Objectives of the research are as follows:

1. To determine level of genetic diversity of wild mud crab (S.serrata) populations in

Peninsular Malaysia.
2. To estimate the size at maturity and assess its relationships to level of genetic

diversity in populations of mud crab, Scylla serrata.

3. To delineate the population structure of S.serrata in Peninsular Malaysia. 4. To recommend d a minimum size at capture S.serrata based on the size at first

maturity.

III

RESEARCH BACKGROUND

Rationale of the Research

Mud crabs of the genus Scylla are commercially important mangrove residents that are extensively fished throughout their range in the Indo-West Pacific. The information about population dynamics of any Scylla species is limited regardless on their high level of exploitation (Le Vay 2006). Recently, juvenile mud crabs have become sought after as stock for domestication program aimed at producing good-quality captive brood stock. Farming of S.serrata is largely confined to Sabah and Sarawak with a combined annual production of 63 metric tons and valued at RM 1 million (DOF,2005).Nonetheless, Malaysia import of crab products in 2010 stands at over RM3 million. Local prices of wild crabs are varying with season, size, sex and gonad maturity. Size and genetics of S.serrata in Peninsular Malaysia are poorly characterized even though it contributed a lot of benefit in economic sector. No regulation adhered in harvesting the species makes it more vulnerable towards any poses threat such as overfishing. Overfishing will not only reduce the number of reproducing individuals in the populations but also led to reduce genetic diversity due to removal of large number of immature individuals. Each harvest introduces tiny, possible irreversible genetic changes into the population and this will runs the risk of reducing productivity of S.serrata fisheries in the future. In addition to genetic changes, studied done by Kosuge (2001), Trippel (1995) and Jorgenson et al (2007) revealed the increasing number of commercially exploited marine stocks due to the revolutionary response pattern in S.serrata. This is because it bear the strategy to alter its life-history in response to exploitation. The population genetic structure of S.serrata in Malaysia has not been determined but such knowledge is crucial for sustainable management of the fisheries, particularly with respect to development of appropriate management of conservation units. Reproduction of S.serrata takes place in the open sea and the larvae remain plaktoni for 3-4 weeks (Hill 1994) so the dispersal is expexted to be wide, thereby the level of gene flows as well as homogenization of local populations is S.serrata is assumed to be high. However , population genetic studies on highly mobile marine organisms have demonstrated genetic differentiation in not only on a macrogeographic scale but also on microgeographic scale (Fratini and Vannini 2002; Hedgecock 1986; Burton and Feldam 1982), suggest that genetic structure of natural populations of marine invertebrates cannot be reliably inferred from their apparent dispersal capacities. No regulation to limit the collection of juveniles and mature crabs showed that it is

important to have stock management in S.serrata fisheries. This study is consist some some of the issue such as size at maturity, genetic diversity and population structure that are relevant to the development of sustainable management of S.serrata fishery in Peninsular Malaysia.

The Strength and Significant of the Research

The strength of this research study is lies on the levels of genetic diversity of S.serrata in populations in Peninsular Malaysia which can be used to delineate the population of the species. The vulnerable species will always be the priority in this research. With the data gathered in this finding, it will upgrade the level of awareness in consumer and government of the conservation effort and start to limit the exploitation of S.serrata. C References

Burton, R. S. , Feldam, M. W. 1982. Population genetics of coastal and estuarine invertebrates: does larval behavior influence population structure? In:Kennedy, V. S. (Ed.), Estuarine Comparisons, New York Academic, New York, pp, 537-551. DOF, 2005. Annual Fisheries Statistics, Department of Fisheries, Malaysia. Fratini, S. and Vaninni, M. 2002. Genetic differentiation in the mud crab Scylla serrata (Decapoda: Portunidae) within the Indian Ocean Journal of Experimental Marine Biology and Ecology 272:102-116 Hedgecock, D., 1994. Temporal and spatial genetic structure of marine animal populations in the California current. CalCOFI Rep: 35, 73-81. Hill, B. J., 1994. Offshore spawning by the portunid crabs Scylla Serrata (Crustacea:Decapoda). Mar. Biol.120,379-384. Jorgensen, C., Enberg, K., Dunlop, E. S., Arlinghaus, R., Boukal, D. S., Brander, K., Ernande, B., Gardmark, A., Johnston, F., Matsumura, S., Pardoe, H., Raab, K., Silva, A., Vainnika, A,. Dieckmann, U., Heino, M., Rjinsdorp, A. D. 2007. Managing Evolving Fish Stocks. Science 318: 1247-1248.

Kosuge, T. 2001. Brief assessment of stock of mud crabs Scylla serrata spp. in Matang mangrove forest, Malaysia and proposal for resource management. JARQ 35(2):145-148. Ricker, W. E. Changes in the average size and average age of Pacific Salmon. Can. J. Fish. Aquat. Sci. 1981; 38:1636-1656. Trippel, E. A. 1995. Age at maturity as a stress indicator in fisheries. Bioscience; 45:759771

IV A

RESEARCH APPROACH Research Methodology

This research will be carried out in 4 stages (methods), as follows: Method 1: Literature review Literature search includes reviews of the published fishery statistics on landing data and methodological approaches taken by other researchers. Information about abundance of crabs, catch statistics and gears (trap) used are gained from the experienced local fishermen. Method 2: Field study : sampling sites Sampling sites which should cover the entire coasts of Peninsular Malaysia will be identified. The validity of sampling regime depends on the composition of the samples reflecting precisely that are present in the habitats. 100 crab samples from both sexes will be collected or contractpurchased at each study sites. Method 3: i)
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Laboratory study: sexual maturity Measurement and classification of sexual maturity Crabs will be tagged, sexed and measurement will be made on carapace width (CW). The maturity stage of crabs will be determined following standard procedures of Kumar et al. (1999).

ii) -

Size at sexual maturity. The size at sexual maturity will be described graphically using frequency distribution. In addition, the percentage of crabs at different CWs for each sex and sampling site will be subjected to logistic regression to determine the size at which 50% of the crabs are mature (Sommerton, 1980).

Method 4: i)

Laboratory study: Genetic diversity and population structure DNA extraction

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Tissue samples will be removed from a single claw. Qiagen QIAmp tissue kit will be used for total genomic DNA extraction. After precipitation DNA will be stored in Qiagen buffer at 4oC for routine use or at -200oC for long term storage. Amplification

ii) -

PCR will be performed for 10 microsatellites markers developed independently by Gopurenko et al. (2002), Hongyu Ma et al. (2010) and Xu et al. (2009) using their recommended PCR profiles and reaction mixtures. PCR fragments will be size fractionated in polyacrylamide gels and scored. Diversity analysis

iii)
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Basic statistics, including the number of alleles, observed heterozygasity (Ho), gene diversity or expected heterozygosity (He), and inbreeding coefficient (f) will be calculated across loci and populations with 1,000 bootstrap sampling using PowerMarker (Liu 2002). Texts for deviations from Hardy-Weinberg equilibrium (HWE, P<0.05) will be performed. Fixation coefficient (Fis) as a measure of heterozygote deficiency will be evaluated amongst populations using computer program POGPEN (Yeh and Yang, 1999). Population structure analysis

iv)
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Population structure will represent the hierarchical Wrights F-statistics (POGPEN). Pair-wise FST estimates amongst all population combinations will be computed through the construction of co-ancestry matrix by using the software PowerMarker. The assignment of

individual crabs to their most likely populations will be performed by the computer program STRUCTURE (Pritchard et al. 2000).
v) -

Population relationship analysis Genetic distance between populations will be computed. The phylogenetic tree will be constructed on the basis of genetic distance matrix the unweighted pair-group method using arithmetic average (UPGMA) using software package HYLIP (Felsenstein, 1993). a) Importance Value of each species and family. b) Total value of biomass which comprise with above and below ground biomass. c) Species diversity, dominance and richness of the study area. d) Soil type and it relationship with the distribution pattern of tree.

Method 4:

Report writing and submission It is anticipated that the thesis will be comprise the following chapters: (i) Introduction (ii) Objectives and the rational of the study (iii) Literature review (iv) Description of the study area (v) Methodology (vi) Result and discussion (vii) Conclusion and recommendation for future work.

BENEFITS OF THE RESEARCH

It is anticipated that the findings of this research will benefit the following sectors: Body of Knowledge Provide the data and information related to the Scylla serrata species especially in genetic diversity as well as its size at maturity

Government and Public Sectors

Create effective management plan to ensure the sustainable use of mud crab, S.serrata.