Phillip G. Lee

Biological Characteristics and Biomedical Applications of the Squid Sepioteuthis lessoniana Cultured Through Multiple Generations

Providing squids--especially their giant axons--for biomedical research has now been achieved in 10 mariculture trials extending through multiple generations. The noteworthy biological characteristics of Sepioteuthis lessoniana are (1) this species is behaviorally and morphologically well suited to the laboratory environment; (2) the life cycle is completed in 4-6 months; (3) growth is rapid (12% and 5% wet body weight d-1 for 100 d and for the life span, respectively), with adult size ranging from 0.4-2.2 kg; (4) feeding rates are high (30% wet body weight d-1), and a variety of live crustaceans and fishes are eaten; (5) crowding is tolerated (about 4 squids m-3); (6) the incidence of disease and cannibalism is low; and (7) reproduction in captivity allows culture through three successive generations. Engineering factors contributed to culture success: (1) physical design (i.e., size, shape, and painted pattern) of the culture tanks; (2) patterns of water flow in the culture tanks; (3) water filtration systems; and (4) spawning substrates. Initial production (a few hundred squids per year) suggests that large-scale culture will be able to supply the needs of the biomedical research community. The size (> 400 microns in diameter) and characteristics of the giant axons of Sepioteuthis are appropriate for experimentation, and other studies indicate that the eye, oculomotor/equilibrium system, olfactory system, blood, and ink are equally suitable for research.

Denitrification in aquaculture systems: an example of a fuzzy logic control problem.

Nitrification in commercial aquaculture systems has been accomplished using many different technologies (e.g. trickling filters, fluidized beds and rotating biological contactors) but commercial aquaculture systems have been slow to adopt denitrification. Denitrification (conversion of nitrate, NO3 to nitrogen gas, N2) is essential to the development of commercial, closed, recirculating aquaculture systems (B 1 water turnover 100 day 1 ). The problems associated with manually operated denitrification systems have been incomplete denitrification (oxidation‐reduction potential, ORP\ 200 mV) with the production of nitrite (NO2 ), nitric oxide (NO) and nitrous oxide (N2O) or over-reduction (ORPB 400 mV), resulting in the production of hydrogen sulfide (H2S). The need for an anoxic or anaerobic environment for the denitrifying bacteria can also result in lowered dissolved oxygen (DO) concentrations in the rearing tanks. These problems have now been overcome by the development of a computer automated denitrifying bioreactor specifically designed for aquaculture. The prototype bioreactor (process control version) has been in operation for 4 years and commercial versions of the bioreactor are now in continuous use; these bioreactors can be operated in either batch or continuous on-line modes, maintaining NO3 concentrations below 5 ppm. The bioreactor monitors DO, ORP, pH and water flow rate and controls water pump rate and carbon feed rate. A fuzzy logic-based expert system replaced the classical process control system for operation of the bioreactor, continuing to optimize denitrification rates and eliminate discharge of toxic by-products (i.e. NO2 , NO, N2 Oo r

The effects of surimi and pelleted diets on the laboratory survival, growth, and feeding rate of the cuttlefish Sepia officinalis L.

The effects of feeding a prepared surimi diet (fish-based) and a prepared pelleted diet (shrimpbased) on the survival, growth and feeding rate of the cuttlefish Sepia officinalis L. were evaluated during a 45-day experiment. One hundred and twenty juveniles of laboratory cultured cuttlefish (74.5 ± 12.36 g) were divided into three treatments and were fed thawed shrimp (control), pellets or surimi. Survival rates on these diets were 95.0%, 67.5% and 22.5%, respectively. Preliminary data indicated that the low survival of cuttlefish fed surimi may have been caused by low levels of copper in their blood (131 vs 244 μg/ml) since copper is required for their respiratory blood pigment, hemocyanin. Instantaneous growth rates were 2.71 % body weight BW/day for cuttlefish fed raw shrimp, 0.33% BW/day for cuttlefish fed pellets, and 0.54% BW/day for cuttlefish fed surimi. The feeding rate of cuttlefish fed shrimp was high (6–8% BW/day). The feeding rate on pellets increased with time (from < 1 to 3% BW/day) but never reached the level for raw shrimp. The feeding rate on surimi increased to equal the rate for raw shrimp during days 1–30 (8 to 9% BW/day) and thereafter decreased (<4% BW/day). In conclusion, there was a major distinction between the palatability of a prepared diet and the ability of that diet to support growth. Surimi was highly palatable but resulted in poor survival, suggesting low nutritional quality. In contrast, pellets were less palatable but produced maintenance growth. Development of practical surimi diets will require supplementation of the surimi with soluble micro- and macronutrients.

The effects of semi-purified diets on growth and condition of Sepia officinalis L. (Mollusca: Cephalopoda)

The effects of five different surimi diets (fish myofibrillar protein concentrate) on growth and condition of Sepia officinalis were evaluated in terms of individual growth using different morphometric and biochemical parameters. A casein supplemented surimi diet produced significant growth (instantaneous growth rate 0.4-0.8% body weight). The digestive gland-to-body weight ratio increased and the cuttlebone-to-body weight ratio decreased significantly in relation to instantaneous growth rate. The RNA content of mantle muscle increased significantly, while the DNA content of mantle muscle did not change in relation to instantaneous growth rate. Mantle muscle protein content was depleted in cuttlefish with instantaneous growth rate < or = 0. No compensatory food consumption, food conversion or growth was observed for cuttlefish fed surimi diets after refeeding them with a natural diet (thawed raw shrimp). Digestive gland and cuttlebone-to-body weight ratios, and RNA content in mantle muscle could be used as short-term indicators of instantaneous growth rate and condition of cuttlefish. Mantle muscle protein content could be used as a long-term indicator.

A review of automated control systems for aquaculture and design criteria for their implementation.

Abstract Agriculture in the United States has become the world leader in productivity through intensification, mechanization and automation. A similar path is appropriate for aquaculture since automation of aquaculture systems will allow the industry to: site production closer to markets; improve environmental control; reduce catastrophic losses; minimize environmental regulations by reducing effluents; reduce production costs; and improve product quality. The history of automated control in aquaculture has been brief; most of the systems have been custom-designed, personal computer systems. The current trend is toward the use of industrial process control systems composed of: sensors/transducers, meters/transmitters, communication multiplexers, actuators/output devices, computer hardware and computer control software. These process control systems can be as simple as one computer or as sophisticated as distributed control systems (multiple networked microcomputers). The choice of the systems architecture should be based on price performance, considering labor, product value, environment and vendor support. Success in designing pragmatic and affordable automated control systems for aquaculture will be widely applicable because it will enhance water management, reduce costs associated with manual monitoring and reduce significantly the chance of catastrophic system failures.

Process control and artificial intelligence software for aquaculture

Abstract Modern research and commercial aquaculture operations have begun to adopt new technologies, including computer control systems. Aquaculturists realize that by controlling the environmental conditions and system inputs (e.g. water, oxygen, temperature, feed rate and stocking density), physiological rates of cultured species and final process outputs (e.g. ammonia, pH and growth) can be regulated. These are exactly the kinds of practical measurements that will allow commercial aquaculture facilities to optimize their efficiency by reducing labor and utility costs. Anticipated benefits for aquaculture process control and artificial intelligence systems are: (1) increased process efficiency; (2) reduced energy and water losses; (3) reduced labor costs; (4) reduced stress and disease; (5) improved accounting; and (6) improved understanding of the process. This review explores the technologies and implementation of the technologies necessary for the development of computer intelligent management systems for enhanced commercial aquaculture production. Today’s artificial intelligence (AI) systems (i.e. expert systems and neural networks) offer the aquaculturist a proven methodology for implementing management systems that are both intuitive and inferential. There have been many successful commercial applications of AI (e.g. expert systems in cameras and automobiles). The major factors to consider in the design and purchase of process control and artificial intelligence software are functionality/intuitiveness, compatibility, flexibility, upgrade path, hardware requirements and cost. Of these, intuitiveness and compatibility are the most important. The software must be intuitive to the user or they will not use the system. Regarding compatibility, the manufacturer should be congruent with open architecture designs so that the chosen software is interchangeable with other software products.

Chemotaxis by Octopus maya Voss et Solis in a Y-maze

Abstract Chemotaxis by Octopus maya Voss et Solis ( n = 20) to chemicals introduced at a distance (teloreception) was measured in a Y-maze. The trials ( n = 195) were conducted in a 40-1 Y-maze system with chemicals entering only one arm of the Y-maze. Octopuses were placed into the base compartment near the convergence of the Y-maze arms and allowed to acclimate to the Y-maze (10 min). A chemical was added to one of the Y-maze arms and octopus movements were recorded (10 min) on videotape. The videotape provided information on the arm first penetrated, the sequence of the penetrations, the number of penetrations into each arm, the duration of time spent in each arm and the base and the mean time spent in each arm per penetration. Eight chemicals (ATP, AMP, alanine, betaine, glutamic acid, glycine, proline and taurine), two crude extracts (crab and shrimp) and the control (no chemical) were introduced at random to only one arm of the Y-maze and each octopus was tested for all chemicals and the control. The final concentration of the chemicals was 10 −4 M except ATP at 10 −5 M and AMP at 10 −5 M, while the crude extracts were prepared by homogenizing crab or shrimp tissue to a final concentration of 5 × 10 −5 g · 1 −1 . Control and chemically exposed octopuses were very active while in the Y-maze, penetrating each arm multiple times. Chemotaxis was demonstrated for octopuses exposed to proline, ATP and crab extract since these octopuses most often penetrated the arm receiving chemical first. Increases in the total amount of activity (number of penetrations into arms and total time spent in arms) were correlated to exposure to proline, alanine, ATP and crab extract; these chemicals functioned as excitants. Betaine and taurine were correlated with lower activity; these chemicals functioned as arrestants. Differences in the number of penetrations per arm and the total duration spent in each arm were affected by both the chemical used and individual octopuses. The sequence of penetrations was not affected by different chemicals. As a result, chemotaxis by octopuses was associated most strongly with first exposure to a chemical, causing initial movement by the octopus into the arm receiving the attractant; octopuses probably habituated to the presence of the chemicals thereafter.

Design and function of closed seawater systems for culturing loliginid squids

The life cycle of loliginid squids has been completed in recirculating seawater systems. Two systems were required: a 2 m diameter circular culture tank (CT) with adjoining 2 m circular filter tank (3000 liters total volume of natural seawater) for culture of hatchlings, 1–60 days old; and a 6 × 2·6 × 1 m raceway culture tank (RW) with a smaller adjoining rectangular filter tank (14 850 liters total volume of artificial seawater) for the grow-out of adults. Both systems were equipped with temperature control apparatus, modular filter units (particle filters and activated carbon), foam fractionators, biological filters (crushed oyster shell) and UV sterilizers. The systems carried low bioloads, < 1·0 g/m3 and as high as 0·8 kg/m3, respectively. Water quality was excellent: NH4N was below 0·01 mg/liter in the CT and 0·10 mg/liter in the RW: NO2N was below 0·01 mg/liter in the CT and 0·03 mg/liter in the RW; NO3N was below 12 mg/liter in the CT and below 50 mg/liter in the RW; and pH was above 8·0 in both systems. The design of the systems proved to be behaviorally and physiologically suitable for squids and two species grew to adult size and produced viable young. These systems are compared to other squid maintenance and rearing systems and marine recirculating seawater systems.

Mariculture of the loliginid squid Sepioteuthis lessoniana through seven successive generations

Abstract Sepioteuthis lessoniana is a commercially important squid throughout the Indo-West Pacific and is a useful species in biomedical research. It has now been cultured through seven successive generations in closed, recirculating seawater systems. Egg viability was highest in the parental generation (34.9%) and significantly decreased in the next six generations with a viability rate of 1.56–13%. The highest hatchling survival was in G 4 (80.3%) and the lowest was in G 3 (26.3%). Water quality was maintained at acceptable levels during each successive generation (NH 4 2 3 8.0). Life spans ranged from 169 to 262 days (5.6–8.7 months) with a mean of 208 days (6.9 months). The seven generations reached mean adult sizes at 500–900 g. The age of sexual maturity for females was recorded for all populations except G 6 and ranged from 146 to 224 days (4.9–7.5 months) with an average of 171 days (5.7 months). Sex ratios did not consistently vary from 1:1 in any generation. Live food (i.e., crustaceans and fish) was the only food supplied throughout their life cycle.

First multi-generation culture of the tropical cuttlefish Sepia pharaonis Ehrenberg, 1831

Sepiapharaonis, the pharaoh cuttlefish was cultured through multiplegenerations in the laboratory (5 consecutive generations) using closed,recirculating water filtration systems. The eggs of the original parentalgeneration (GP) were spawned by a wild caught Gulf of Thailandfemale in alocal fisheries laboratory, then packed and shipped air cargo to Texas wherehatching occurred. The culture temperature ranged 25°–28°C, except for one generation that was chilled intentionallyto21 °C and then warmed to 25 °C after 9.6months. Spawning occurred as early as day 161. Spawning output was high in allgenerations except the group that was cultured at 21 °C. Eggfertility was low in captivity (< 20%), but hatchling survival was high(>70%). The average egg incubation time was 13.6 d at 25–28°C. The largest spawn resulted in 600 viable hatchlings andthesmallest resulted in 11 hatchlings. The cuttlefish ate a wide variety ofestuarine crustaceans and fishes as well as frozen shrimp. There were noapparent disease problems since survival from hatching to maturity was over70%.The average life span for cuttlefish cultured at 25–28°Cwas 8.9 months and 12.3 months at 21 °C. Size at hatching wasmeasured for fourth generation (G4) hatchlings; the mean weight athatching was 0.103 g and the mean mantle length was 6.4mm. The largest cuttlefish cultured was a male 300 mmML and 3,045 g; the oldest cuttlefish lived 340 d.This cuttlefish species presents an excellent choice for commercial mariculturebecause of its rapid growth, short life span, tolerance to crowding andhandling, resistance to disease and feeding habits.