Structure and Classification of Haemocytes in the Bivalve Mollusc Meretrix meretrix

Light and electron microscopic studies were carried out in order to characterize haemocytes in the bivalve mollusc Meretrix meretrix. According to nucleus and cytoplasm characters, four types of haemocytes were recognized： agranular haemocytes, lymphoid haemocyte, large granular and small granular haemocytes. Agranular hamocyte is the main cell type, accounting for 75%. It is agranular with rich organelles in cytoplasm, including mitochondria, golgi body and endoplasmic reticulum. Glycogen deposits were usually found in this cell type. The number of lymphoid haemocyte accounts for 1% - 2%. This cell type is agranular and shows a high ratio of nucleus to cytoplasm. A few organelles were found. High electrondense granules with diameters of 0.2 - 0.5μm and rich organelles were found in small granular haemocyte. The proportion of this cell type is about 15%. Rich granules of high electron-dense with diameters of 0.8- 2.4μm were found in large granular haemocyte. The proportion of this cell type is about 10%, and the quantity of organelles is the least.

Effects of Temperature on Haemocyte and Granulocyte Counts and Expressions of Immunity-Related Genes in Haemocytes of Scallop Chlamys farreri After Vibrio anguillarum Infection

Bivalve live in aquatic environment and the water temperature can affect their immunity directly.In this research,the scallop Chlamys farreri was injected with 10^4 or 10^7 CFU mL^-1 Vibrio anguillarum and cultured at 11℃,17℃,23℃,and 28℃,respectively.For the control scallop,only phosphate-buffered saline(PBS)was injected.Then total haemocytes and granulocytes were measured by ELISA using monoclonal antibodies.In the meantime,expressions of six immunity-related genes,including lipopolysaccharide andβ-1,3-glucan binding protein(CfLGBP),C-type lectin(CfLec-2),Toll-like receptor(Cf TLR),Lysozyme(CfLYZ),superoxide dismutase(SOD),and phenoloxidase(CfPO)in haemocytes were measured using quantitative real-time PCR.The results showed that total haemocytes counts in 10^4 CFU mL^-1 injection groups showed no differences compared to the control group at all temperatures.However,they varied significantly in 10^7 CFU mL^-1 injection groups at 3 h at 11℃,6–12 h at 17℃,3–48 h at 23℃,and 12–48 h at 28℃.Granulocytes counts in 10^4 CFU mL^-1 injection groups showed no variance compared to the control group at all temperatures,except for 12 h at 23℃,and 24–36 h at 28℃.They were significantly decreased in 10^7 CFU mL^-1 injection groups during 6–48 h at 11℃,12–48 h at 17℃,3–48 h at 23℃,and 3–72 h at 28℃.The expression levels of six immunity-related genes in haemocytes of 10^7 CFU mL^-1 injection groups were significantly higher than those of control group and 10^4 CFU mL^-1 injection groups at all temperatures.The results indicated that infected with high concentration of vibrios,haemocyte counts,granulocyte counts and the expressions of immunity-related genes in scallop C.farreri were significantly affected by environmental temperature.

Classification and morphology of circulating haemocytes in the razor clam Sinonovacula constricta

The razor clam Sinonovacula constricta is widely distributed in the intertidal zones and estuarine waters along the coast of western Pacific Ocean and is extensively cultured.Even though haemocytes are known to play an important role in the immune mechanisms of bivalves,these cells are poorly studied in S.constricta.We researched the morphology and immunological activities of haemocytes in S.constricta using light and electron microscopy and flow cytometry.Three major subpopulations of haemocytes were identified in the haemolymph:granulocytes,semigranulocytes,and hyalinocytes.These subpopulations were divided using flow cytometry,but not satisfactorily.Therefore,the flow cytometry findings were combined with the light and electron microscopy findings,as well as Percoll density-gradient centrifugation findings,to classify and distinguish between the cell types more effectively.The combined findings showed that granulocytes was larger cells,while semigranulocytes was smaller and more abundant.Further,granulocytes had numerous granules in the cytoplasm,semigranulocytes contained fewer and smaller granules,and hyalinocytes was smaller and less abundant with no or a few granules.Both granulocytes and semigranulocytes had greater phagocytotic activity and a higher lysosomal content than hyalinocytes.The results declared that granulocytes and semigranulocytes were the main haemocytes involved in the cellular defence mechanism in S.constricta.

Influence of two different dietary zinc sources in freshwater prawn Macrobrachium rosenbergii post larvae

A feeding experiment was conducted to evaluate the ef fects of bulk zinc(B-Zn) and zinc nanoparticles(Zn-NPs) on survival, growth, compositions of amino acid and fatty acid, and nonspeci?c immune responses of the freshwater prawn, Macrobrachium rosenbergii post larvae. The B-Zn(size 10 μm)and Zn-NPs(size 50 nm) were supplemented separately with the basal feeds of M. rosenbergii at 60 mg/kg and feed without supplementation of any Zn source was served as a control. M. rosenbergii were fed these feeds for 90 days and the results showed that signi?cant( P <0.05) improvements in survival, growth, feed intake,speci?c growth rate, essential amino acids, unsaturated fatty acids, nonspeci?c immune responses(total haemocytes and dif ferential haemocytes count) of M. rosenbergii fed with B-Zn and Zn-NPs supplemented feeds when compared to control. Among these Zn sources, Zn-NPs supplemented feeds fed prawns showed signi?cantly( P <0.05) better performance than that of B-Zn and control. Hence, present study suggests that the 60 mg/kg Zn-NPs can be supplemented with basal feeds of M. rosenbergii for regulating better survival and growth.

Physiological, histopathological and cellular immune effects of Pergularia tomentosa extract on Locusta migratoria nymphs

The migratory locust Locusta migratoria(Orthoptera,Acrididae)is one of the most important pests due to its extensive and severe damage to crops in large parts of Africa and Asia.Biodegradable and ecologically natural products such as bota nical insecticides are emergi ng can didates for replaceme nt of usually applied chemical pesticides.The crude methanolic extract of Pergularia tomentosa(PME)was investigated for their toxicity and physiological aspects on L.migratoria nymphs.Results showed that treatment of newly emerged fourth and fifth instar nymphs resulted in significant mortality and significant repellent activity with an LC50 value of 0.18 and 0.38%,respectively,after seven days of treatment.The PME toxicity was also dem on strated by histopathological cha nges in the alime ntary canal resulting in con siderable disorganization and severe damage of the caeca and proventriculus structure.The extract induced cellular immune reactions which manifested by a significant decrease in the number of the differential haemocyte counts(prohemocytes and plasmatocytes)and important cell lysis.Data of biochemical analyses showed that the PME reduced the activity of acetylcholinesterase and induced the glutathione S-transferases.The neurotoxic effect was confirmed by the histological alterations in the brain structure,particularly in the neurosecretory cells showing typical signs of cell necrosis.

Immunotoxicity Effect of Benzo[α]Pyrene on Scallop Chlamys farreri

The toxic effects of benzo[α]pyrene （B[α]P） at different concentrations （0.1, 0.5, 1, 2.5 and 7.5 μgL^-1） on scallop （Chlamys farreri） immune system were studied. The results showed that B[α]P had significant toxic effects on the haemocyte counts, neutral red uptake, phagocytosis, bacteriolytic and antibacterial activity （P〈0.05）, while the seawater control and acetone control had no significant differences. The haemocyte counts, neutral red uptake, phagocytosis and bacteriolytic activity in all B[α]P treatment groups as well as antibacterial activity in groups of 0.5, 1, 2.5 and 7.5 μgL^-1·B[α]P decreased significantly （P〈0.05）. Some of these indices tended to be stable on the sixth day and others on the ninth day, and the indices showed clear time- and concentration-response to B[α]E Bactenolytic activity in 0.1 μgL^-1 B[α]P treatment group and antibacterial activity in 0.1 ggLl and 0.5 μgL^-1 B[α]P treatment groups increased at the beginning of exposure and reached their peaks on day 1 and day 6, respectively. Following that, both activities decreased gradually and became stable after day 9. When all the indices reached stability, they were significantly lower than those in control group （P〈0.05）, except for antibacterial activity in 0.1 μgLl B[α]P treatment group （P〉0.05）. Thus, B[α]P has evident toxic effects on scallop immune system, which supports the view that a relationship exists between pollution and lmmunomodulation in aquatic organisms.

Responses of Prophenoloxidase System and Related Defence Parameters of Litopenaeus vannamei to Low Salinity

In this study,we investigated the effects of low salinity(26 and 21) on the prophenoloxidase(proPO) system and related defence parameters in the shrimp Litopenaeus vannamei.The results showed that low salinity induced a significant increase of dopamine(DA) concentration in haemolymph at 6 h of the experiment;on the other hand,total haemocyte count(THC),differential haemocyte count(DHC) and PO activity decreased over time to the lowest level at 24 h and remained low thereafter.Serine Protease(SP) and Proteinase Inhibitor(PI) activity in the two lower salinity treatments decreased to the lowest level at 12 and 24 h,respectively,and both recovered to the control level at 72 h.In contrast,α2-macroglobulin(α2M) activity in the two lower salinity treatments peaked at 24 h and then decreased to the control level at 72 h.Therefore,it may be concluded that stress-induced DA plays an important temporary role in neurotransmission and causes immune response in L.vannamei in adapting to salinity changes.

Effect of salinity changes on haemocyte of pearl oyster Pinctada martensii

Objective:To evaluate the impact of salinity changes in the rearing environment on the haemocyte activities of pearl oyster Pinctada martensii(P.martensii).Methods:Three salinity treatments including 12‰,22‰,and 32‰were tested.Haemocyte mortality rate and neutral red retention(NRR)assay were used to investigate the impact of rearing salinity on the haemocyte activities of pearl oyster.Results:Results from the present study indicated that salinity significantly affected the haemocyte mortality rate of P.martensii.The highest haemocytes mortality rate was observed in the 12‰treatment,and the lowest mortality rate was observed in the 32‰treatment.NRR time was also significantly affected by the salinity.The highest NRR time was observed in the 32‰treatment,and the lowest NRR time was observed in the 12‰treatment.Conclusions:Results from the present study indicated that salinity<22‰had significant impact on the haemocyte mortality rate and NRR time.Reducing the environmental salinity will pose an additional stress and may also reduce the defense capacities of P.martensii and make them more susceptible to parasites and bacteria.

Growth and immune response of pond-reared giant freshwater prawn Macrobrachium rosenbergii post larvae fed diets containing Chlorella vulgaris

A 70-day feeding trial was conducted to evaluate the efficacy of Chlorella vulgaris supplementation in diets of giant freshwater prawn(Macrobrachium rosenbergii)post larvae.Resistance of the prawns against Aeromonas hydrophila and several immune parameters(total hemocyte count and prophenoloxidase activity)were also assessed in the study.Iso-nitrogenous,iso-lipidic,and iso-caloric prawn feeds were prepared utilizing a fishmeal based positive control diet(F0)and four dietary treatments with C.vulgaris inclusion levels of 2(F2),4(F4),6(F6),and 8(F8)%.Post larvae of M.rosenbergii were stocked randomly(mean initial body weight of 0.39�0.38 g)in fifteen net cages(8.1 m�8.9 m with an average depth of 1 m)in an earthen pond for the assessment of growth parameters.Overall growth indices for prawn fed Chlorella containing diets were enhanced in comparison to the control treatment.Prawn fed diets with Chlorella showed significantly higher(P<0.05)prophenol oxidase activity,total hemocyte counts,and survival rates post bacterial infection.These findings demonstrated that inclusion of 4%,6%,and 8%C.vulgaris in prawn diets provided optimal growth rates and improved immunity of the post larvae.