Aquacultural Engineering 2 (1983) 287-300

Shipping Studies with Juvenile and Adult Malaysian Prawns Macrobrachium rosenbergii (de Man)*

Theodore I.J. Smith and Allen J. Wannamaker

Marine Resources Research Institute, PO Box 12559, Charleston,
South Carolina 29412, USA

ABSTRACT Four studies examined shipping factors o f packing technique, density, duration, type o f water and use o f habitat material for shipping juvenile and adult prawns Macrobrachium rosenbergii. Prawns were shipped in double polyethylene lined 38 x 38 x 20 cm deep styrofoam boxes containing oxygenated water. A t temperatures o f 19-20C, 1 7 g prawns could be shipped safely for 42 h at a density o f 10-12 prawns per box (12-15g liter-1 shipping water). Juveniles, mean size about 6 g, could be shipped at a density o f 40 per box (18g liter-1) for 24 h or 20-25 per box (9-11 g liter -1) for 48 h. Use o f mesh material to increase surface area in the box did not appear beneficial nor did shipping in brackish water (salinity ~ 8/oo). Adults packed unrestricted resulted in survival rates substantially higher than those obtained from immobilized prawns wrapped in mesh. During the shipment, p H and dissolved oxygen concentrations decreased whereas ammonia concentrations increased. The decreased p H levels may have reduced the ammonia toxicity by decreasing the amount o f toxic unionized ammonia (NHa) in solution. In general, dissolved oxygen concentrations appeared more closely related to survival rates than did other water quality parameters which were measured.

* Contribution no. 174 from the South Carolina Marine Resources Center. This research was sponsored by the Coastal Plains Regional Commission, Grant no. 10740047, a grant from Aquacultura de Guatemala, and the State of South Carolina. Reference to trade names does not imply endorsement. 287 A q uacultural Engineering 0144-8609/83/$03.00 Applied Science Publishers Ltd, England, 1983. Printed in Great Britain

288

T. L J. Smith, A. J. Wannamaker

INTRODUCTION Suitable population densities and water quality parameters have been identified for shipping postlarval Malaysian prawns Macrobrachiurn rosenbergii by commercial hatcheries. These data are not published, but they generally can be obtained from the hatchery managers. Local deliveries are accomplished with live tanks mounted on trucks while long distance shipments require packaging and boxing for air freight transport. The packing procedures for air freight shipment of prawns are similar to those employed in the shipment of tropical fishes. First, two flat bottom polyethylene bags are placed in a styrofoam box (approximately 38 38 20 cm deep) which is then inserted into a cardboard shipping carton. Next, water is added to the inner polyethylene bag and the animals stocked. The shipping water is oxygenated with pure oxygen as is the air space above the water. The inner and outer bags are sealed separately with elastic bands and then the shipping carton is taped closed. Postlarval prawns ( ~ 20 mg in weight) are packed at a density of about 2500-3000 per box for transit times of ~ 24 h while about 1500-2500 per box are packed for shipments of 24-48 h. Prawns are packed in either brackish water (~< 18~o salinity) or freshwater. Prior to shipment water temperature is lowered to 18-23C and the prawns are not fed for 24 h. Sometimes tris(hydroxymethylaminomethane) or other compounds are added to buffer the water while clinoptilolite may be added to freshwater shipments to absorb ammonia. When the above packing and shipping techniques are followed, survival rates of the order of 90% are typically achieved with postlarval prawns (Smith and Sandifer, unpublished data). For a number of years the Marine Resources Research Institute, South Carolina Wildlife and Marine Resources Department, has been a focus for M. rosenbergii research and development activities in the continental US (Sandifer and Smith, 1974, 1976, 1978; Smith et al., 1976, 1981; Liao et al., 1982). As such, live specimens have been shipped to various organizations and investigators throughout the US and other countries. Information and guidelines for shipping juvenile and adult prawns were not available and so studies were undertaken to identify techniques and densities for efficient shipment of larger prawns. The studies summarized here investigated the effects of physical immobilization of prawns, use of habitat materials, shipment in

Shipping studies with juvenile and adult Malaysian prawns

289

brackish water and stocking density on survival o f juvenile and adult prawns.

M A T E R I A L S AND METHODS Prawns were packaged in 38 38 20 cm deep styrofoam boxes containing 13.6 liters o f water following the procedures described above. The smaller prawns were obtained from indoor nursery systems whereas the larger animals were reared in o u t d o o r ponds. During the 24-h period prior to packing, water temperature was reduced to the approximate level used in the studies (18-23C) and the prawns were not fed. To simulate actual shipping conditions, all boxes containing prawns were periodically shaken or moved around during the course o f the various experiments. Prawns reported as stressed were either lying on their lateral surface, had opaque musculature, or displayed impaired mobility. Water quality was generally measured at the beginning, middle and end o f the studies. Shipping bags which were opened to provide water quality data during the study had the air space above the water refilled with oxygen prior to reclosure. Water quality was monitored as follows: temperature - mercury thermometer; dissolved oxygen - Yellow Springs Co. meter model 57; salinity - American Optical refractometer; pH, ammonia-nitrogen, nitrite-nitrogen, and nitrate-nitrogen - LaMotte Chemical Co. test kits, models P-5085 and AG-36, PAN, PLN, and ENA, respectively. Accuracy o f the LaMotte test kits was verified with a Bausch and L o m b Spectronic Mini 20. The freshwater used in the studies consisted o f aged or dechlorinated tap water (river water) or water obtained from a shallow well ( < 3 0 m deep). Water quality characteristics o f the tap and well water are shown in Table 1. Brackish water was obtained by mixing natural seawater or synthetic sea salts with the freshwater. F o u r shipping studies were conducted having different approaches and objectives. For simplicity, the objectives and characteristics o f the various experiments are presented separately below. Experiment I The usual procedure for shipping adult prawns is to individually wrap each animal in mesh or webbing to immobilize it during shipment. This

290

T. L J. Smith, A. J. Wannamaker

~/V/

o,

~6
f2~ r.~

o v~
0

cq C

=.~
Z" C tt~

cS&
o~

0 ,. ('q

b~--.
C

~=~
b

Shipping studies with juvenile and adult Malaysian prawns

291

method helps prevent injury and cannibalism, and puncture of the water bag by the prawn. However, the individual immobilization technique is very time consuming when large numbers o f prawns are to be shipped. In the first study we examined stocking density, packing technique and shipping time. Prawns, average size 17 g, 8.6 cm (orbit o f eye to tip o f telson), were individually immobilized by wrapping in nontoxic plastic mesh (Conweb Corp., product no. XN5175) and stocked in replicate boxes at'densities o f 5, 10 and 15 prawns per box (6.3, 12.5 and 18.8 g liter -1 shipping water, respectively). Two additional boxes were stocked with 10 unwrapped or free prawns. Two other boxes received no prawns and served as controls for comparison of water quality data. Tap water was aged for one week before use in all treatments. Experiment II There were two objectives o f this second study using adults: 1. to compare the effects o f using different sources of water (newly dechlorinated tap water, well water and a mixture of well water and synthetic sea salts); 2. to examine the effects o f shipment time on survival and water quality. Prawns, average size 17.4 g, were stocked at a density o f 12 per box (15.4 g liter -1) and inspected after 24 and 42 h. Sodium thiosulfate was used to dechlorinate the tap water immediately before stocking the prawns. Instant Ocean brand synthetic sea salts were added to the well water to provide brackish water (6%0). Experiment III This study provided information on the shipping of juvenile prawns. Three stocking densities (25, 50 and 75 prawns per box; 10.5, 21.0 and 31.5 g liter q , respectively) were compared using newly dechlorinated tap water. Also, the use o f an inexpensive artificial habitat material was examined to determine its effect on survival. Juvenile prawns, mean weight 5.7 g, were stocked at each density in triplicate. A 0.3 m 2 piece of 10 cm 2 plastic mesh was rolled and placed in one box at each density treatment to provide habitat area for the prawns. Survival and water quality were determined after 24 h.

292

T. L J. Smith, A. J. Wannamaker

Experiment IV
This final study further focused on identifying appropriate shipping parameters for juvenile prawns. The objectives were: 1. to compare effect o f prawn size on survival at a constant shipping biomass; 2. to compare effect o f two types o f water on shipment o f juveniles. Total prawn biomass per b o x was held constant at 224 g per box (16.5 g liter -1) whereas prawn size and shipping density were varied. Mean prawn sizes were 7.4, 5.6 and 4-5 g and densities were 30, 40 and 50 prawns per box, respectively. Water types were freshly dechlorinated tap water and synthetic sea salts mixed with tap water (salinity 4%0). Prawns were inspected at 24 and 48 h. Water quality was analyzed from one replicate o f each treatment at 24 h and from all replicates at 48 h.

RESULTS Experiment I After 24 h all boxes were opened, the adult prawns inspected, and water quality analyzed (Table 2). Water quality in the control boxes remained relatively constant, whereas there were noticeable changes in the boxes containing prawns. The pH decreased in all treatments, and ammonia-nitrogen levels increased. After 24 h, survival was 100% at the lowest density (5 prawns per box, 6.3 g liter -1) and 0% at the highest density (15 prawns per box, 18.8 g liter-a). Survival among the immobilized prawns stocked at 10 per b o x (12.5 g liter -1) averaged 15%, as compared with 100% for the unwrapped or free prawns at the same stocking density. Treatments in which prawns were immobilized at 5 per box and free at 10 per b o x were continued for an additional 24 h. After 48 h, these treatments were again examined. A substantial increase in ammonianitrogen concentrations occurred and oxygen levels were reduced. Survival of the immobilized prawns averaged 10% as compared with 75% for the free animals stocked at double the density (Table 2).

Shipping studies with juvenile and adult Malaysian prawns

293

O 0

O 0

O 0

O 0

O 0

O 0

~o

O ~2

VV
o

VV

VV

VV

7
66
VV
~2 o~

66
VV

06

66

66
VV

oo
V

06
V

e.,

"N
~ O0 O0 O0 O0

294 Experiment II

T. L J. Smith, A. J. Wannamaker

All t y p e s o f w a t e r w e r e suitable f o r s h i p p i n g t i m e s o f 42 h ( T a b l e 3). A v e r a g e survival was 9 8 . 6 % at 24 h a n d 9 7 . 2 % at 4 2 h. T h e least c h a n g e in p H level o c c u r r e d in t h e well water. H o w e v e r , w a t e r q u a l i t y in all t r e a t m e n t s was generally similar. Based o n results o f this s t u d y 2 0 0 0 adult p r a w n s w e r e divided into t w o g r o u p s and s t o c k e d u n w r a p p e d at

TABLE 3 Water Quality and Survival Data for Study Examining Effects of Water Type and Shipping Time on Adult Prawns (12 per box, Experiment II)

Type of water Tempera- Oxygen ture (mg (C) liter-1)

Water quality data pH NO2-N NOa-N NH4-N (mg (mg (mg liter-1) liter-1) liter-1)

Survival (%)

Tap Well Brackisha Tap Tap Well Well Brackisha Brackish a Tap Tap Well Well Brackisha Brackish a a Salinity = 6~oo.

20.3 20.0 20.5 20.0 19.9 19.9 19.9 19.2 19.6 20-0 20-0 20-0 20.0 19.7 19-5

>20.0 > 20,0 > 20.0 17.2 17.7 11.4 14.0 14-6 18.0 13.4 13-8 9.2 15.0 14.2 15.8

Time = 0 h 9.0 <0.1 6-4 <0.1 7.9 <0.1 Time = 24 h 6.5 <0.1 6.3 <0.1
6.7 6.7 6.6 6.9 <0.1 <0-1 <0-1 <0.1

< 1.0 < 1.0 < 1.0 < 1-0 <1.0 <1.0 <1.0 <1.0 < 1.0 < 1.0 <1.0 < 1-0 <1.0 < 1.0 < 1.0

< 1.0 < 1.0 < 1.0 2.0 3.0 3.0 2-0 2.0 1.0 4.0 3.0 5.0 4-0 3-0 2-0

100.0 100.0 91-7 100.0 100-0 100.0 100.0 100.0 83.3 100.0 100.0 100.0

Time = 42 h 5.7 <0.1 5.6 <0.1

6.6 6.7 6.7 6-8 <0.1 <0.1 <0.1 <0.1

Shipping studies with juvenile and adult Malaysian prawns

295

10 per box in either freshly dechlorinated tap water or freshly dechlorinated tap water and Instant Ocean synthetic sea salts (salinity 1-2%o). These prawns were shipped overseas using commercial freight carriers. The average duration o f box confinement was 42 h. In this field trial survival o f adults exceeded 95% in both types o f water. Experiment III After 24 h survival o f juveniles was high (96.0 and 93.3%) at densities o f 25 (10.5 g liter-1) and 50 per box (21.0 g liter-I), respectively, and lower (85.3%) at a density o f 75 per box (31.5 g liter-~) (Table 4).

TABLE 4

Water Quality and Survival Data for Juvenile Prawns (Mean Size, 5.7 g) Stocked at Three Densities. Initial Water Quality Parameters Were: Temperature, 20C; Oxygen, > 20-0 mg liter-1; pH, 8.4; NO2-N, < 0-1 mg liter-~; NO3-N, < 1.0 mg liter- ~; NH4-N, < 1.0 mg liter-1 (Experiment III)

Density (number per box)

Water quality data Tempera-Oxygen ture (mg (C) liter-i) pH NO2-N NOa-N NH4-N (mg (mg (mg liter-1) liter-lj liter-1)

Survival (%)

25, No habitat 25, No habitat 25, Habitat 50, No habitat 50, No habitat 50, Habitat 75, No habitat 75, No habitat 75, Habitat 25, No habitat 25, No habitat 25, Habitat

19.0 19.0 19.0 19-2 19.2 19.1 19.2 19.2 19.7 18.2 18.2 18.2

15-4 15.2 18-2 8.7 3.2 2.6 1.9 2.1 1.6 11.4 16.4 16.4

7~me = 24 h 6.8 <0.1 6-8 <0.1 6.8 <0.1

6-7 6-7 6.7 6.6 6.6 6-6 <0-1 <0-1 <0.1 <0.1 <0.1 <0.1

<1-0 <1.0 <1.0 <1.0 <1.0 <1.0 <1.0 <1.0 <1-0 < 1-0 <1.0 <1-0

2-0 2.0 2.0 4-0 4-0 4.0 6.0 6.0 6.0 5-2 4.0 5.2

96.0 96.0 96.0 96.0 96.0 88.0 85.3 84.0 86-7 96-0 92-0 92-0

Time = 48 h 6.0 <0.1 6.0 <0.1 6.0 <0.1

296

T. L J. Smith, A. J. Wannamaker

Despite the high survival rates, many prawns were stressed after 24 h at the two highest densities. In both of these treatments, dissolved oxygen levels decreased substantially, whereas ammonia concentrations increased. Because of the poor condition of the prawns and deteriorated water quality, the two high density treatments were terminated at 24 h. The low density treatment (25 per box) was continued for an additional 24 h. After 48 h, prawns in this treatment still exhibited a good survival rate (mean, 93-3%). The habitat material had no noticeable effect on survival at any density during the initial 24-h period nor after 48 h in 25 per box density.

Experiment IV
Survival rates of juveniles were excellent and averaged 99.5% for all treatments after 24 h; no stressed prawns were observed at this time. After 48 h, survival rates had declined slightly to 90.8 and 91.9% for densities of 30 and 40 per box, respectively (Table 5). However, approximately 55% of the survivors at both these densities appeared stressed. Survival at the highest density averaged only 21%, and of the survivors, 96% were severely stressed. On average, survival rates in the brackish water were slightly higher than those obtained in freshwater at the two lower densities, but at the highest density survival in brackish water was much lower than that obtained in the freshwater. It should be noted that all boxes of prawns were visually inspected without opening the bags after 43-44 h. At this time survival averaged 99-2% at density 30, 95.6% at density 40, and 96-0% at density 50. Thus, most of the mortalities occurred during the last 4-5 h of the study, suggesting that one or more water quality factors had reached a critical level. The oxygen concentration had substantially decreased in all treatments after 48 h, especially at density 50, and ammonia-nitrogen levels were more than double those recorded at 24 h.

DISCUSSION The various studies have identified changes in water quality which occur during shipment of prawns as well as providing guidelines for shipping M. rosenbergii juveniles and adults. When prawns are shipped in

Shipping studies with/uvenile and adult Malaysian prawns

297

TABLE 5 Water Quality and Survival Data at 48 h for Study Examining Relationship Between Stocking Biomass and Density and Effects of Brackish Water. Initial Water Quality Parameters at Stocking Were: Temperature, 20C; Oxygen, > 20.0 mg liter-l; NO2-N, <0.1 mg liter-l; NO3-N, <1.0 mg liter-l; NH4-N, <1.0 mg liter-l; Freshwater pH, 7.7; Brackish Water pH, 8.5 (Experiment IV)

Treatment Density Salinity (number (~oo) perbox)

30 30 30 30 40 40 40 40 50 50 50 50 0 0 4 4 0 0 4 4 0 0 4 4

Water quality data Tempera-Oxygen ture (mg (C) liter-l)

20.1 20.2 20.1 20.1 20.2 20.1 20.1 20.0 20.1 20.0 20"1 20.0 2.5 3.2 6-9 7.0 2.4 3.0 3.0 2.6 1.0 1.5 1"2 1.4

Survival

(%J
pH NO2-N NO~-N NH4-N (rag (rag (mg liter-1) liter-l) liter-l}
<0.1 <0.1 <0.1 <0.1 <0-1 <0.1 <0.1 <0.1 <0-1 <0.1 <0"1 <0.1 <1-0 <1.0 <1.0 <1-0 <1.0 8.0 2-0 6.0 2.0 2.0 2'0 2.0 7.0 7.0 7-0 7.0 8-0 7;7 8.0 8.0 > 8.0 >8.0 >8-0 >8-0 73.3 96.6 100.0 93-3 82.5 92-5 97.5 95.0 32.0 48.0 0"0 4.0

6.0 6.0 6-0 6.0 6.0 6.0 6.0 6-0 7-0 6.8 6"5 6.5

closed containers, pH and dissolved oxygen concentrations typically decrease whereas ammonia concentrations increase. In aqueous solutions it is chiefly the un-ionized ammonia (NH3) that is toxic to aquatic organisms as ionized ammonia (NH~) cannot pass through tissues (Milne et al., 1958). However, the ratio of N H 3 NH~ in solution is highly dependent on pH and less so on temperature. As pH and temperature decrease so does the a m o u n t o f un-ionized ammonia. Based on the pH and temperature values recorded during the studies, the percent unionized ammonia ranged from 0.04 to 0.40 and the m a x i m u m NH3-N concentration was about 0.03 mg liter-1 (Emerson et al., 1975). Thus, although the pH decreased during shipment, this reduction may have alleviated shipping stress and mortality associated with the accumulation of ammonia.

298

T. L Z Smith, A. J. Wannamaker

Oxygen also appeared to be a major water quality consideration when shipping juveniles and adults. Not only does oxygen level influence the respiration rate but decreased oxygen levels have been shown to increase the toxicity of un-ionized ammonia in some freshwater fishes (Merkins and Downing, 1957). Currently, postlarval prawns are shipped at densities which provide a total biomass of "~ 20-60 g per shipping box or "-- 1.5-4.5 g liter-1 shipping water. When such boxes are opened even after 48 h, dissolved oxygen concentrations are often still at super-saturation levels (/> 15 mg liter-1) (Smith and Sandifer, unpublished data). In the shipment of juveniles and adults, higher biomasses are packed because of cost considerations. In the studies reported herein, packing biomasses ranged from about 85 to 427 g per box (6-3 to 31.5 g liter-I). In some cases, oxygen levels were still elevated after 24 h; by 48 h, however, substantial decreases in oxygen were usually observed. Efforts to ameliorate or prevent changes in water quality were not attempted during these studies. Changes in carbon dioxide and pH can be controlled with buffers (Nemoto, 1957; McFarland and Norris, 1958), and a naturally occurring zeolite (clinoptilolite) has been shown to reduce the build-up of ammonia during shipment of freshwater ornamental fishes (Bower and Turner, 1982). However, granular size, water hardness and pH affect efficiency of ammonia removal (Marking and Bills, 1982). Such procedures, combined with techniques to maintain suitable oxygen levels during shipment could increase the cost effectiveness of shipping larger prawns.

CONCLUSIONS AND RECOMMENDATIONS 1. A variety of water sources appeared equally suitable for shipping prawns including freshly dechlorinated tap water. 2. During shipment, pH and dissolved oxygen concentrations decrease whereas ammonia concentrations increase. 3. In general, the concentration of dissolved oxygen appeared to be more closely related to survival rate than did the other water quality parameters monitored. 4. No benefits resulted from adding plastic habitat material or of shipping in brackish water (2-89~o).

Shipping studies with juvenile and adult Malaysian prawns

299

5. Packing adult prawns unwrapped rather than physically immobilized greatly improved survival and increased the allowable packing density. 6. A stocking density o f about 10-12 prawns per b o x (weight about 17 g) appears safe for shipping times up to 42 h, if temperature is lowered to 19-20C. 7. Juvenile prawns, weight 5.6-6.5 g, can be shipped at a density o f "~40 per b o x for up to 24 h. For shipping times up to 48 h, a density o f ~ 20-25 prawns per b o x (mean size 6.5 g) appears suitable at 19-20C. 8. In general, larger prawns exhibit higher survival rates over a longer period o f time than do smaller prawns packed at similar stocking biomasses.

ACKNOWLEDGEMENTS We thank Frank Taylor, Caroline Gray and Glen Murray for assisting in these studies. Virginia Hargis t y p e d the manuscript.

REFERENCES Bower, C. E. & Turner, D. T. (1982). Ammonia removal by clinoptilolite in the transport of ornamental freshwater fishes. Prog. Fish-Cult., 44 (1), 19-23. Emerson, K., Russo, R. C., Lund, R. E. & Thurston, R. V. (1975). Aqueous ammonia equilibrium calculations: effect of pH and temperature. J. Fish. Res. Bd. Can., 32, 2379-83. Liao, D. S., Smith, T. I. J. & Sandifer, P. A. (1982). Economic, market, and technical considerations for prawn farming in South Carolina. Proc. Trop./Subtrop. Fisheries Tech. Conf. Americas, 7, 139-42. Marking, L. L. & Bills, T. D. (1982). Factors affecting the efficiency of clinoptilolite for removing ammonia from water. Prog. Fish-Cult., 44 (4), 187-9. McFarland, W. N. & Norris, K. S. (1958). The control of pH by buffers in fish transport. Calif. Fish Game, 44, 291-310. Merkins, J. C. & Downing, K. M. (1957). The effect of tension of dissolved oxygen on the toxicity of un-ionized ammonia to several species of fish. Ann. Appl. BiOl., 45,521-7. Milne, M. D., Scribner, B. H. & Crawford, M. A. (1958). Non-ionic diffusion and the excretion of weak acids and bases. Amer. J. Med., 24, 709-29.

300

T. L J. Smith, A. J. Wannamaker

Nemoto, C. M. (1957). Experiments with methods for air transport of live fish. Prog. Fish-Cult., 19 (4), 147-57. Sandifer, P. A. & Smith, T. I. J. (1974). Development of a crustacean mariculture program at South Carolina's Marine Resources Research Institute. Proc. World Maricult. Soc., 5,431-9. Sandifer, P. A. & Smith, T. I. J. (1976). Experimental aquaculture of the Malaysian prawn Macrobrachium rosenbergii (de Man) in South Carolina, USA. FAO Tech. Conf. in Aquaculture, Kyoto, Japan. Sandifer, P. A. & Smith, T. I. J. (1978). Aquaculture of Malaysian prawns in controlled environments. Food Technol., 32 (7), 36-8, 40-2, 44-5, 83. Smith, T. I. J., Sandifer, P. A. & Trimble, W. C. (1976) Pond culture of the Malaysian prawn, Macrobrachium rosenbergii (de Man), in South Carolina, 19741975. Proc. World Maricult. Soc., 7,625-45. Smith, T. I. J., Sandifer, P. A., Jenkins, W. E. & Stokes, A. D. (1981). Effect of population structure and density at stocking on production and commercial feasibility of prawn Macrobrachium rosenbergii farming in temperate climates. Proc. World Maricult. Soc., 12 (1), 233-50.