WORLD AQUACULTURE 49 to detect if viral pathogens are present in these products even when they are purchased from reliable sources. Freshly harvested yellow fin tuna roe samples (23) were collected from a local tuna processing company during dry and wet seasons. Molecular diagnoses were performed on six pooled tuna roe samples for presence of shrimp viruses, following published PCR protocols for white spot syndrome virus (WSSV) (Lo et al. 1996), infectious hypodermal and hematopoietic necrosis virus (IHHNV) (Nunan et al. 2000), and RT-PCR protocols for Taura syndrome virus (TSV) (Nunan et al. 1998), yellowhead virus (YHV) (Tang and Lightner 1999) and infectious myonecrosis virus (IMNV) (Poulos and Lightner 2006). Diagnostic results verified that tuna roe was free of these viral pathogens. Diet Formulation and Evaluation A semi-moist diet was formulated based on the nutritional values of tuna roe and other feed ingredients to achieve suitable texture and water stability (Table 3). Proximate analysis of the semi-moist diet was compared to that of a proprietary commercial maturation diet containing 55.3 percent protein and 9.8 percent lipid. A three-phase evaluation was performed to compare the tuna roe-based diet with different control feeding regimes. For each phase, SPF L. vannamei adults (>30 g) produced from the same cross were stocked at 10 per m2 in 30m3 maturation fiberglass outdoor tanks (23.6 m2 bottom area) with a male to female ratio of 1.4:1. The fish roe maturation diet was made daily (Fig. 2) and stored at -20 °C until feed was used. Twenty minutes prior to each feeding, the diet was removed from storage and cut into small pieces (~2 g) for feeding shrimp broodstock. Shrimp were conditioned by feeding ad libitum with the experimental diet and control diet four times per day. After feeding diets for one month, female shrimp reached an average of more than 45 g (Fig. 3) and overall reproductive traits were then evaluated. For the first two phases, ovarian maturation was assessed by external observation of ovarian size and color as described by Vaca and Alfaro (2000): Stage I. The ovary is transparent with no distinguishable outline. Stage II. The ovary is visible as a thin opaque line along the dorsal central axis. Stage III. The ovary is visible as a thick and yellow band. Stage IV. The ovary is turgid, broad and dark orange. Mating and spawning are imminent. Females that reached ovarian developmental stages III and IV were identified and recorded from each tank during daytime. Each evening around 2000 h, females were checked for natural mating during first two phases of each comparison. Artificial insemination was performed on stage IV females during the phase 3 comparison for each dietary treatment. After insemination, each female was transferred to an individual spawning tank (121 L). Spawned females were removed from spawning tanks around midnight. The number of eggs per spawn, percentage of fertilized eggs, hatching rate and number of nauplii per spawn were recorded. Between 10 and 12 hours after spawning, three 100 mL samples were taken to assess the total number of eggs. Microscopic investigation of the proportion of normal embryos was used to calculate fertilization rate. Thirty-four hours after spawning, three samples of 100 mL were randomly taken to count number of nauplii. Hatching rate was (CONTINUED ON PAGE 50) FIGURE 1. A tuna processing plant on Guam. FIGURE 2. The semi-moist diet containing tuna roe. FIGURE 3. Female shrimp fed with the semi-moist diet. TABLE 1. Proximate analysis of tuna roe (mean and SD; n=6). Sample # Ash (%) Protein(%) Lipid (%) P (%) K (%) Ca(%) Mg (%) Na (%) B (ppm) Cu (ppm) Fe (ppm) Mn(ppm) Zn (ppm) Mean 6.8 74.5 10.0 1.25 0.94 0.05 0.14 1.03 1.2 5.0 61 1.7 360 SD 0.8 1.7 3.9 0.09 0.17 0.01 0.01 0.13 0.4 0.6 11 0.5 36
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