Sha-Yen Cheng

The immune response of the white shrimp Litopenaeus vannamei and its susceptibility to Vibrio infection in relation with the moult cycle

The white shrimp Litopenaeus vannamei (8.0-14.4 g) was examined for haemocyte count, phenoloxidase activity, respiratory burst (release of superoxide anion), phagocytic activity, and clearance efficiency to the pathogen Vibrio alginolyticus in relation with moult cycle (postmoult, A, B; intermoult, C; premoult, D(0)/D(1)D(2)/D(3)). Granular cells were the highest at C and D(0)/D(1)stage, and the lowest at A stage. Hyaline cells and THC (total haemocyte count) were higher at C stage, but lower at postmoult stages. Phenoloxidase activity was the highest at C stage, and the lowest at A stage. Respiratory burst was lower at A stage. Phagocytic activity of shrimps against V. alginolyticus decreased significantly at postmoult and premoult stages. Additionally, the clearance efficiency of shrimps to V. alginolyticus was significantly lower for shrimps at A stage than those at C stage. In another experiment, L. vannamei at different moult stages were injected with tryptic soy broth (TSB)-grown V. alginolyticus (1x10(5)cfu shrimp(-1)) and then held in 34% seawater. After 10 h, the mortality of V. alginolyticus-injected shrimps was significantly higher for shrimps at postmoult stage than those at intermoult stage. Over 48-120 h, the mortality of V. alginolyticus-injected shrimps was 50.0%, 33.3% and 40.0% at postmoult, intermoult and premoult stage, respectively. It is concluded that L. vannamei showed a decrease in resistance at A stage through a reduction of its haemocyte count, phenoloxidase activity, respiratory burst, phagocytic activity and clearance efficiency against V. alginolyticus.

Studies on haemocyanin and haemolymph protein levels of Penaeus japonicus based on sex, size and moulting cycle

1. 1. Mean (± SE) haemolymph protein level was 64.78 ± 4.06 and 62.24 ± 4.13 mg/ml, while mean haemocyanin level was 0.72 ± 0.05 mol/l for male (12.45 ± 0.60 g, N = 27) and female (12.41 ± 0.68 g, N = 21) Penaeus japonicus, with no significant difference between the sexes. 2. 2. Protein and haemocyanin in the haemolymph of shrimp significantly increased (P < 0.05) with increased weight in the range of 7.56–21.22 g (N = 70). 3. 3. Among the moulting cycles tested, the shrimp had the highest mean (± SE) protein (74.90 ± 4.61 mg/ml) and haemocyanin (0.73 ± 0.04 mol/l) at the D0 stage, and had the lowest protein (41.37 ± 3.11 mg/ml) and haemocyanin (0.42 ± 0.02 mol/l) at the B stage. 4. 4. Haemocyanin accounts for 64% (C stage) to 72% (D3 stage) of protein in the haemolymph of the shrimp (9.21 ± 0.36 g), and accounts for 76% (females) to 80% (males) of protein in the haemolymph of the shrimp (12.43 ± 1.17 g), respectively.

Hemolymph oxygen content, oxyhemocyanin, protein levels and ammonia excretion in the shrimp Penaeus monodon exposed to ambient nitrite

Subadult Penaeus monodon (21.03±3.19 g) were exposed individually in sea water (30 mg·ml-1) to 0.02 (control), 1.04, 5.02, 10.11 and 20.06 mg·l-1 nitrite-N for 24h. Hemolymph pH, partial pressures of oxygen and carbon dioxide, bicarbonate concentration, oxyhemocyanin and protein levels, and whole animal ammonia-N excretion and nitrite-N uptake were determined. Ammonia-N excretion and hemolymph oxygen partial pressure increased, whereas hemolymph pH, HCO3-, oxyhemocyanin, protein and the ratio of oxyhemocyanin/protein levels decreased with increasing ambient nitrite-N. It is suggested that accumulated nitrite of P. monodon following exposure to ambient nitrite causes reduction of oxyhemocyanin, protein and the ratio of oxyhemocyanin/protein in the hemolymph, and affects nitrogen metabolism and acid-base balance at low hemolymph pH.

Study on the oxyhemocyanin, deoxyhemocyanin, oxygen affinity and acid-base balance of Marsupenaeus japonicus following exposure to combined elevated nitrite and nitrate.

Marsupenaeus japonicus (11.47+/-0.71 g) exposed individually to six different nitrite and nitrate regimes [(0.002 (control), 0.359 and 1.456 mM nitrite combined with 0.005 (control) and 7.458 mM nitrate)] in salinity of 30 ppt (parts per thousand) were examined for hemocyanin oxygen affinity, the fractionation of oxyhemocyanin and deoxyhemocyanin, and the acid-base balance after 24 h. Ambient nitrite at concentration of 0.359 mM caused reduction of oxyhemocyanin and protein by 27 and 11%, respectively, whereas ambient nitrate as high as 7.458 mM caused reduction of oxyhemocyanin and protein by 10 and 7%. Ambient nitrite at concentration of 1.456 mM caused increases of P(50) (indicating reduced oxygen affinity) and pO(2), but caused reduction in hemolymph pCO(2), pH, HCO(3)(-) and TCO(2). Following exposure to combined solutions of 1.456 mM nitrite and 7.458 mM nitrate there were no further changes in oxyhemocyanin, protein, hemolymph P(50), pO(2), pCO(2), HCO(3)(-) and TCO(2), but there was a significant reduction of pH.

Effects of nitrite exposure on the hemolymph electrolyte, respiratory protein and free amino acid levels and water content of Penaeus japonicus

Abstract Penaeus japonicus which had been exposed individually for 3, 6, 12, 24 and 48 h to 0.001 (control), 0.07, 0.36, 0.72 and 1.43 mM nitrite at 30 ppt of salinity were examined for hemolymph electrolyte, oxyhemocyanin, protein, free amino acid (FAA) and urea levels and whole shrimps nitrite uptake and ammonia excretion. Hemolymph oxyhemocyanin, protein, FAA, osmolality, Cl − and Na + levels were inversely related to ambient nitrite and exposure time. However, hemolymph urea level, osmotic differential (medium osmolality−hemolymph osmolality), nitrite uptake, ammonia excretion and water content increased directly with ambient nitrite and exposure time. It is suggested that when P. japonicus are exposed to nitrite, water uptake and nitrite uptake result in reductions of hemolymph Cl − , Na + and protein levels leading to a decrease of hemolymph osmolality. An increase of hemolymph urea and a decrease of arginine may indicate an occurrence of ureogenesis for the intracellular osmoregulation of P. japonicus under nitrite stress.

Hemolymph PCO2, hemocyanin, protein levels and urea excretions of Penaeus monodon exposed to ambient ammonia

Abstract Subadult Penaeus monodon (21.985 ± 3.546 g) were exposed individually in 30 ppt seawater to 0.060 (control), 0.994, 4.631, 9.593 and 20.728 mg/l ammonia-N for 24 h. Hemolymph pH, PO2, PCO2, hemocyanin and protein levels, and whole animal ammonia-N, urea-N and nitrite-N excretions were determined. Urea-N excretion and nitrite-N excretion increased, whereas ammonia-N excretion and hemolymph PCO2 level decreased with increased ambient ammonia-N. P. monodon exposed to ambient ammonia-N greater than 4.631 mg/l, net ammonia-N uptake occurred, and caused significant reduction of hemocyanin and hemolymph protein and their ratio. In addition, there was a shift from ammonotelic to ureotelic excretory pattern of shrimps following exposure to ambient ammonia-N greater than 4.631 mg/l.

Hemocyanin oxygen affinity, and the fractionation of oxyhemocyanin and deoxyhemocyanin for Penaeus monodon exposed to elevated nitrite

Abstract Hemocyanin oxygen affinity and the fractionation of oxyhemocyanin and deoxyhemocyanin in the hemolymph of Penaeus monodon were studied for individual shrimps exposed to 0.001 (control), 0.07, 0.36, 0.72 and 1.44 mmol l−1 nitrite for 3 h up to 48 h in a salinity of 30 ppt at 25°C. The hemolymph P50 increased (indicating reduced affinity) and the hemolymph pO2 increased directly with ambient nitrite and exposure time. Hemolymph pH, oxyhemocyanin and hemolymph protein were inversely related to ambient nitrite and exposure time. P. monodon, following 6 h exposure to ambient nitrite at 0.36 mmol l−1, decreased significantly the fraction of oxyhemocyanin and increased that of deoxyhemocyanin. The increase of P50 and the increased fraction of deoxyhemocyanin suggests that hemocyanin oxygen binding decreases with a concomitant decrease of hemolymph pH, when P. monodon is subjected to nitrite.

Changes of haemocyanin, protein and free amino acid levels in the haemolymph of Penaeus japonicus exposed to ambient ammonia

Abstract Haemolymph ammonia-N of Penaeus japonicus (15.6 ± 2.17 g) increased with increased ambient ammonia-N, as they were exposed individually in 30 ppt seawater to 0.003 (control), 0.367, 0.731, 1.439 and 3.665 mmol/1 ammonia-N after 24 hr. Ammonia-N excretion was inhibited and net ammonia-N uptake occurred, as shrimps were exposed to 0.367 mmol/1 ammonia-N or greater. Haemocyanin and protein levels in the haemolymph of shrimp decreased, whereas free amino acid levels increased with increased ambient ammonia-N in the range of 0.003–1.439 mmol/1 ammonia-N. Exposure of shrimp to ambient ammonia-N at 1.439 mmol/1 caused accumulation of haemolymph ammonia and urea, and caused catabolism of haemocyanin and protein to free amino acids. Urea, taurine and glutamine are the major organic constituents in the haemolymph of shrimp under ammonia stress.

Changes of oxyhemocyanin and protein levels in the hemolymph of Penaeus japonicus exposed to ambient nitrite

Abstract The concentrations of oxyhemocyanin, protein and urea in the hemolymph, and ammonia-N excretion of Penaeus japonicus (15.08 ± 1.56 g) were examined, after the shrimp had been exposed individually in 30 ppt to 0.01 (control), 5.12, 10.46, 20.57 and 50.86 mg/l nitrite-N at 25 °C for 24 h. Accumulation of nitrite-N and urea-N in the hemolymph, and ammonia-N excretion of shrimp increased with increased ambient nitrite-N over the entire range. Nitrite-N uptake occurred, and increased with increased ambient nitrite-N in the range of 5.12–50.86 mg/l. Exposure of P. japonicus to elevated nitrite-N affected nitrogen metabolism, and decreased the oxyhemocyanin and the ratio of oxyhemocyanin/protein. It is suggested that nitrite incorporated into the hemolymph of penaeids may decrease the oxygenation state of the hemolymph, and decrease the fraction of oxyhemocyanin.

The time-course change of nitrogenous excretion in the Kuruma shrimp Penaeus japonicus following nitrite exposure

Penaeus japonicus (12.83 +/- 1.24 g) which were exposed individually to 0.002 (control), 0.076, 0.362, 0.719 and 1.433 mM nitrite at 30 ppt of salinity were examined for hemolymph ammonia, urea and nitrite, and nitrogenous excretion after 3, 6, 12, 24 and 48 h, respectively. Hemolymph nitrite and hemolymph urea increased directly with ambient nitrite and exposure time, whereas hemolymph ammonia was inversely related to ambient nitrite and exposure time. Specific excretions of total-N (total nitrogen), ammonia-N, urea-N and organic-N (organic nitrogen) increased directly with ambient nitrite and exposure time. The contribution of ammonia-N excretion and urea-N excretion in the total-N excreted by the control shrimp was 41.7-90.8 and 2.8-10.5%, respectively. The contribution of ammonia-N in the total-N excreted by P. japonicus decreased to 10.0 and 3.8%, when they were exposed for 24 h to 0.076 and 1.433 mM nitrite, respectively. The contribution of urea-N excretion increased to 58.7 and 21.4%, and the organic-N excretion increased to 31.3 and 74.8% in the total-N excreted by the shrimp following 24 h exposure to 0.076 and 1.433 mM nitrite, respectively. It was concluded that P. japonicus following 24 h exposure to nitrite as low as 0.076 mM increased its ammonia-N excretion by a factor of 1.9, its urea-N excretion by 200, and its organic-N excretion by 37, as compared to those in the control solution.