Materials and methods Directory UMM :Data Elmu:jurnal:A:Aquaculture:Vol185.Issue1-2.May2000:

1. Introduction

Ž . Ž Farmers of mud crabs Scylla species rely on small crabs or juveniles 4–30 g, . 18–4.0 cm carapace length; CL sourced from intertidal flats and mangroves to stock Ž . grow-out ponds. However, smaller crabs 0.5–5.0 g; 1–2 cm CL can be nursed for 1–2 Ž . weeks in net cages prior to stocking in grow-out ponds SEAFDEC, 1997 . With the increasing demand and declining supply of crab seed, hatchery-reared seed provides an alternate source. Since 1998, researchers of the Aquaculture Department, Southeast Ž . Asian Fisheries Development Center SEAFDECrAQD have been consistently produc- ing mud crab juveniles in the hatchery. There is, however, low survival due primarily to the occurrence of luminescent vibrios at the late zoea and megalopa stages, and at the juvenile stage due to cannibalism. Recently, SEAFDECrAQD researchers successfully cultured hatchery-reared mega- lopae to juveniles in net cages installed in brackishwater ponds for 4 weeks. The Ž . survival and growth rates achieved were greater in ponds 35–53; 2.9–3.4 g than in Ž . hatchery tanks 25–35; 0.70–2.5 g; shooters not included . The culture of megalopae to juveniles in ponds could be a promising alternative to growing them in tanks. With this development, handling and transport of megalopae need to be included in routine stock transfer activities in the hatchery. Handling, packing, and transport activities expose the animals to stress. Crab juveniles are relatively easy to transport by using a wicker basket, a native straw Ž . ‘pandan’ bag or carton lined with moist mangrove or leaves or cheese cloth. Chelae Ž . are tied or cut to prevent fighting among crabs Ladra and Lin, 1991 . In air, mud crab has a life span of 2–18 days when packed with moist marine algae, cotton, or wood Ž . shavings Vasudeo and Kewalramani, 1960 . We observed that megalopae are more sensitive to handling, packing, and transport than juveniles. The optimal conditions during transport of megalopae needed to be determined to reduce mortalities. This study considered the optimal loading density at various durations of transport of hatchery-reared Scylla serrata megalopae. Various temperature levels were also tested to determine the effect of cooling on the survival of megalopae. The taxonomic Ž . classification of the species S. serrata used in this study is based on Keenan et al. Ž . 1998 .

2. Materials and methods

Ž . Ž . There were three experiments: a simulated transport at various loading densities, b simulated transport at various loading densities and durations with and without shaking, Ž . and c simulated transport at various temperature levels. 2.1. Large-scale production of megalopae Newly hatched zoeae were stocked in 10- and 1.5-tanks at 30–50 ind l y1 . Seawater Ž . 32 ppt for rearing was chlorinated with 10–20 ppm hypochlorite and neutralized with sodium thiosulfate. Rearing water was replaced daily at 30, commencing on day 3. The volume replaced was increased from 50 to 80 in the succeeding days. Zoeae were fed Brachionus rotundiformis at 10–15 ind ml y1 and artificial larval diet. Newly hatched Artemia, introduced at zoeae 3 stage, were gradually increased from 1 to 5 ind ml y1 as the larvae progressed. The period of larval development to megalopa normally ranged from 17 to 18 days. 2.2. Simulated transport at three loading densities Three- to five-day old megalopae at loading densities of 50, 100, and 150 ind l y1 Ž . were packed in double polyethylene plastic bags 34 = 25 cm with cooled seawater Ž . 22–24 8C; 28–30 ppt . Bags were inflated with oxygen and sealed tightly. Transport was simulated by loading the bags in a styrofoam box and subjecting them to shaking Ž using an orbit shaker over a period of 6 h. Measured amounts of ice chunks 200 g = 4 . pcs. wrapped in newspaper inside the styrofoam box effectively maintained the temperature in the bags. Immediately after transport, survival of megalopae was determined in each bag. Thereafter, megalopae were stocked in separate 6-l rearing y1 Ž . containers at 10 ind l . Seawater 28 ppt was added gradually to the containers provided with aeration. Newly hatched Artemia served as their feed. Survival of megalopae was monitored 15 h post-transport. Two trials were conducted with three replicates of each loading density. 2.3. Simulated transport, loading densities, duration and shaking Since transport of animals consists of mobile and stationary periods, unshaken and shaken conditions were included as another factor in determining optimal loading densities at various transport durations. Three-day old megalopae stocked at 50 and 100 ind l y1 were subjected to shaking or allowed to remain unshaken for 3, 6, or 9 h. A 2 Ž y1 . Ž . Ž 50 or 100 ind l loading density = 2 shaken or unshaken condition = 3 3, 6, or 9 h . transport time factorial experiment was used. The packing and transport procedures Ž were the same as in the first experiment. Water samples for dissolved oxygen Rain- . Ž . Ž . Ž water and Thatcher, 1960 , ammonia NH -N and nitrite NO -N Strickland and 3 2 . Parsons, 1972 were taken before and after the simulated transport. Three replicates of each treatment were used to determine the survival immediately after transport and another three replicates for 15 h post-transport monitoring. 2.4. Simulated transport at three temperature leÕels Three- to five-day old megalopae were packed in plastic bags at 50 and 100 ind l y1 . Ž . Temperature was adjusted to 20, 24, and 28–29 8C ambient by placing bags in pre-cooled water and allowing the water in plastic bags with megalopae to reach the desired levels. Simulated transport, with shaking, was done over a 6-h period, with the same procedures as in the previous experiments. Temperature was maintained during simulated transport by placing pre-weighed ice blocks wrapped in newspaper. The amounts of ice to maintain 20 8C and 248C were determined in an earlier trial. Temperature was checked every 2 h and controlled by addition or reduction of ice. Survival was determined following simulated transport. There were three replicates for each treatment. In all experiments, arcsine transformations of the percentage survival were analyzed Ž . through analysis of variance ANOVA followed by Duncan’s New Multiple Range Test Ž . Walpole, 1982; Gomez and Gomez, 1984 .

3. Results