Total Coelomocytes Counts Research Articles

Vitamin E requirement of sea cucumber (Apostichopus japonicus) and its' effects on nonspecific immune responses

This feeding trial was conducted to determine the vitamin E requirement of sea cucumber (Apostichopus japonicus) and to study the effects of vitamin E on nonspecific immune parameters. Commercial sea cucumber diets supplemented with 0, 50, 100, 200, 500 or 1000 mg kg−1 vitamin E were fed to juvenile sea cucumber (7.96 ± 0.01 g) for 60 days. Specific growth rate and total coelomocyte counts were maximized in sea cucumber fed diets supplemented with 100 mg kg−1 vitamin E. Reactive oxygen species production was maximized at 50 mg kg−1 vitamin E, whereas superoxide dismutase activity was the highest in sea cucumber fed diets supplemented with 100 mg kg−1 vitamin E. The vitamin E concentration in the body wall of the sea cucumber increased as the vitamin E level in the diet increased. Analysis of specific growth rate and total coelomocyte counts of the sea cucumber indicated that 88–92 mg kg−1 vitamin E is required for maximal growth and nonspecific immune responses of A. japonicus and that 114.7 mg kg−1 vitamin E is required to minimize lipid peroxidation levels.

Effects of Small Peptides on Nonspecific Immune Responses in Sea Cucumber, Apostichopus japonicus

AbstractThe potential immunostimulatory effects of small peptides on sea cucumber, Apostichopus japonicus, were investigated in vivo and in vitro. Small peptides were injected into sea cucumbers and a control group was injected with equal volume of sterile filtered seawater. The nonspecific humoral and cellular responses were determined, respectively, and sea cucumbers were challenged by Vibrio splendidus. The results showed that test groups significantly enhanced most of the immune parameters tested (P < 0.05). Both phagocytic capacity and respiratory burst activity were up to maximum value on the fourth day and the values were 2.3 and 1.4 times as high as the control group, respectively. However, small peptides had no significant effect on total coelomocytes counts. Among the humoral responses, the activities of acid phosphatase, alkaline phosphatase, and superoxide dismutase were up to maximum value on the fourth, fourthth and seventh day, respectively. The values were 2.3, 2.2, and 2.0 times as high as the control group, respectively. At the end of the challenge, the cumulative mortality rate of sea cucumber injected with 0.5 mg small peptides (43.3%) was significantly lower than the control group (66.7%). In conclusion, small peptides as an additive could improve the nonspecific immunity level of Apostichopus japonicus.

Effects of potential probiotic Bacillus subtilis T13 on growth, immunity and disease resistance against Vibrio splendidus infection in juvenile sea cucumber Apostichopus japonicus

A feeding experiment was conducted to determine influences of potential probiotic Bacillus subtilis T13 (isolated from intestine of healthy sea cucumbers) on growth, immunity and disease resistance against Vibrio splendidus infection in juvenile sea cucumbers Apostichopus japonicus. Animals were fed with diets containing B. subtilis T13 at 0, 105, 107 and 109 CFU/g for 30 days, respectively. At the end of the growth trial, fifteen sea cucumbers from each aquarium were sampled for immune indices measurement. Then twenty sea cucumbers from each replicate were challenged with V. splendidus. Results showed that administration of B. subtilis T13 had significant effect on the specific growth rates (SGR) of sea cucumbers (P < 0.05). Phagocytosis, respiratory burst activity and total nitric oxide synthase (T-NOS) activity were significantly improved in coelomocytes of sea cucumbers fed with T13 at 109 CFU/g diet (P < 0.05). The highest values of the total coelomocytes counts (TCC) and superoxide dismutase (SOD) activity were found in sea cucumbers fed diet containing T13 at 109 CFU/g. The cumulative mortality after V. splendidus challenge decreased significantly in sea cucumbers fed with T13 at dose of 109 CFU/g (P < 0.05). The present study confirmed the potential beneficial effects of B. subtilis T13 as dietary probiotic in juvenile A. japonicus.

Interaction of dietary Bacillus subtilis and fructooligosaccharide on the growth performance, non-specific immunity of sea cucumber, Apostichopus japonicus

A feeding experiment was conducted to investigate the interaction of probiotic Bacillus subtilis and prebiotic fructooligosaccharide (FOS) on the growth performance, immunity, intestinal microflora and disease resistance of sea cucumber ( Apostichopus japonicus). Five hundred and forty individuals (initial body weight: 5.06 ± 0.10 g, mean ± S.E) were fed nine practical diets according to a 3 × 3 factorial design: the basal diet as the control diet supplemented with three levels of B. subtilis (0, 1.82 × 10 7 or 4.95 × 10 7 CFU g −1 diet), crossed with 0, 0.25% or 0.50% FOS. After 8 weeks, three sea cucumbers per tank were sampled for bacterial quantification and immunity determination. Then all the sea cucumbers left were challenged by Vibrio splendidus. The results showed that dietary B. subtilis significantly increased the specific growth rate (SGR), total coelomocytes counts (TCC), phagocytosis of sea cucumbers, the counts of total viable bacteria and disease resistance to V. splendidus ( P < 0.05), whereas the counts of Vibrio decreased. However, dietary B. subtilis had no significant effect on phenoloxidase (PO) activity in coelomocyte lysate supernatant (CLS) ( P > 0.05). The SGR, PO activity, total viable bacterial counts (TBC) and Vibrio counts (VBC) were significantly affected by dietary FOS. In the group with 0.50% FOS, TCC, phagocytosis and PO activity significantly increased compared to the group fed without FOS in diet ( P < 0.05). In the groups with 1.82 × 10 7 CFU B. subtilis/g diet, FOS supplementation remarkably decreased VBC. And higher level of FOS (0.50%) resulted in significantly higher TCC and PO activity compared with 0.25% FOS ( P < 0.05). Moreover, the animals fed with diets supplemented with 0.25% and 0.50% FOS at each B. subtilis level had notably lower cumulative mortality after 14 days following V. splendidus exposure ( P < 0.05). Under the experimental conditions, dietary B. subtilis and FOS had a synergistic effect on enhancing immunity and disease resistance of sea cucumber ( P < 0.05).

Immune condition of Apostichopus japonicus during aestivation

Aestivation is an indispensable state in the life history of sea cucumbers, Apostichopus japonicus. The immune characteristics of the coelomic fluid of A. japonicus, were investigated during aestivation. Samples were collected between July and November 2006 from a coastal pond located off the Yellow Sea in Jiaonan, Shandong Province, China. The total coelomocytes counts (TCC), total superoxide dismutase (T-SOD), catalase (CAT), myeloperoxidase (MPO), and lysozyme (LZM) in the coelomic fluid were measured. The activities of catecholamines, [adrenaline (AD), noradrenaline (NA), and dopamine (DOP)] were estimated. TCC decreased from July to September, indicating weakness of the cellular immune activity at that time. Activities of SOD, CAT, MPO, and LZM changed significantly from July to October. Catecholamines AD and NA in coelomic fluid were significantly higher on August 21 and November 27. There was no significant variation in DOP during the sampling period. Thus, immune characters in coelomic fluid of A. japonicus changed significantly during aestivation. Water temperature was significantly and negatively correlated with TCC, and salinity was significantly and positively correlated with AD and NA. The mechanism of aestivation in A. japonicus is complex and might not be attributed only to environmental changes, such as temperature and salinity, as shown in previous studies.

Coelomocyte numbers and expression of HSP70 in wounded sea stars during hypoxia

Hypoxia, mainly caused by eutrophication, is a common and growing problem on marine soft bottoms. Echinoderms are known for their ability to regenerate tissue after wounding but hypoxia has a negative influence on regeneration and also on reproduction in echinoderms. We have investigated the cellular and molecular responses to wounding stress and hypoxia in the sea star Asterias rubens by using the total coelomocyte count (TCC) and the expression of heat shock proteins (HSPs). As early as 1 h after wounding, sea stars under hypoxic conditions show significantly increased TCC and, after 6 h, cell numbers increase approximately two-fold. After a 3-h hypoxia exposure of wounded animals, Western blot analysis reveals highly elevated coelomocyte cytoplasmic HSP70 expression. Non-wounded sea stars exposed to hypoxia and wounded animals kept in normoxia show enhanced HSP70 expression only after 24 h. Immunocytochemical analysis has not demonstrated any translocation of HSP70 from the cytoplasm to the nucleus. We conclude that both wounding and hypoxia elicit a stress response in sea stars and that the combined stress produces synergistic effects that may inhibit the initial processes of wound healing and regeneration.

ECHINODERM IMMUNOLOGY: BACTERIAL CLEARANCE BY THE SEA URCHIN STRONGYLOCENTROTUS PURPURATUS.

Characteristics of bacterial clearance were investigated in the purple sea urchin, Strongylocentrotus purpuratus (Echinodermata: Echinoidea). Primary clearance kinetics were determined for three bacteria, a marine Gram negative motile rod, a marine Gram positive non-motile rod, and a Gram negative freshwater fish pathogen, Aeromonas salmonicida. Clearance kinetics differed for each of the three bacteria. Secondary clearance rates were not significantly different from primary clearance rates for any of the three bacteria, regardless of the time interval between inoculations (9-21 days), implying a probable absence of immunologic memory. During primary clearance, total coelomocyte counts declined 93% by 90 min post injection. All four coelomocyte types declined, however the relative proportions of each type changed during the six-hour sampling period. In cell-free coelomic fluid, viable counts of marine bacteria declined, with different kinetics for the two species. Viable counts in sea water controls did not change. Declines in viable counts may be due to bactericidal activity and/or agglutination, although bacterial agglutination was not observed.