| Litopenaeus vannamei is one of the most important farmed products in the world.The introduction of L. vannamei in shrimp farming is step by step from seawater tofreshwater. With rapid growth of shrimp farming, more and more diseased problems ofshrimps cause a major loss in shrimp aquaculture. The virus is a main pathogenic factorin explosive epidemic shrimp disease. Presently, experts research on immunology of L.vannamei in order to finding several ways to improve the immunity against disease.And they try to find a biochemical marker of immunological mechanism. Therefore,theimmune-related enzymes such as phosphatase (ACP), alkaline phosphatase (AKP),superoxide dismutase (SOD), peroxidase (POD), polyphenoloxidase (PPO) and so onhave been paid more attention. Research on these enzymes in L. vannamei is importantreference value in theory to understand the technology of pathogens diagnosis, thetreatment of disease problems and especially the prevention studies on shrimp.1 Study on Electrophoretograms of Five Isozymes of LitopenaeusvannameiVertical polyacrylamide gel electrophoresis (PAGE) and biochemical staining wereused to analyze five isozymes including ACP, AKP, SOD, POD and PPO in the eye,liver, heart, gill and muscle of L. vannamei. The electrophoretic patterns of theisozymes showed differences between the healthy shrimps and the diseased shrimpsinfected with virus. The results showed that the five isozymes were all expressed infive kinds of tissues of L. vannamei, and activities of the five isozymes were tissuespecific. Except SOD was the highest in heart of L. vannamei. The activities of theother isozymes were all the highest in liver. After shrimps were infected with virus,physiological metabolism in diseased shrimps was apparently beginning to fail. Thephenotypes of five isozymes had some obviously changes, and some characteristicbands of ACP, AKP, POD and PPO decreased or got weaker in liver and gill. ACPdecreased 2 characteristic bands in liver in diseased shrimps. Furthermore, ACPdecreased 1 characteristic band in gill, and 2 bands got weaker. AKP decreased 1 and 3characteristic bands in liver and gill respectively. POD decreased 4 characteristic bands in liver, and decreased 1 band which was deeper and thicker band in gill of healthyshrimps. PPO decreased 1 characteristic band which was the deepest and thickest bandin healthy shrimps in liver, and in gill also decreased 1 band. But on the downtrend ofSOD activity was in heart and gill. SOD decreased 1 band in heart and gill respectivelyin diseased shrimps. Furthermore, higher activities of ACP, AKP, POD and PPO weredetected in heart of diseased shrimps. ACP, POD and PPO presented 1 new bandrespectively, and AKP presented 3 new ones in heart. Higher activity of SOD wasdetected in liver. Although there was no new band of SOD in heart, 2 bands whichwere detected in healthy shrimps got deeper and thicker. Due to theelectrophoretogram of fiver isozymes changed most obviously in liver and heart ofdiseased shrimps with virus, the shrimp isozymes in liver and heart can be used asbiochemical marker for diagnosis of L.vannamei with virus.2 Study on Cytochemical Location of Five Isozymes of LitopenaeusvannameiThe activities and location of ACP, AKP, SOD, POD and PPO in the liver, muscle,heart, gill and eye of L. vannamei were studied with enzyme cytochemical techniqueof electronic microscope, compared it with the infected shrimp. The results showedthat the ACP activities were mainly deposited in nucellar cell, endoplasmic reticulumand lysosome, and in liver ACP were detected in microvilli, around lipid droplet,mitochondrium and cellular endomembrane. AKP activities were mainly deposited innucellar cell, nuclear membrane, endoplasmic reticulum, cellular membrane andsecondary lysosome. And in liver AKP were detected in microvilli, around lipiddroplet, mitochondrium and cellular endomembrane, and mitochondrial membrane ingill of the healthy shrimps. The SOD activity was site in mitochondrium, cytoplasm incells and around lipid droplet. And the POD and PPO activities were site in specificorganelle respectively: peroxisome and prophenoloxidase. In infected shrimp withvirus, the pathological changes were obviously in tissues and cells. These changeswere showed as follows: The dilatation of endoplasmic reticulum (sarcoplasmicreticulum) and mitochondrium became small vacuoles. Cytoplasm was concentrationand turbidity. More myelin figures appeared in cytoplasm of cells. Muscle fiberbecame loose, very relaxed and obviously ruptured. Lipid droplet was isolated smallgranule in the cytoplasm. Microvilli were fewer. And Nucellar cell became abnormal.At the same time, the activities site of five enzymes had changes in cells. ACP positivereaction granules were only observed in myelin figure, mitochondrial membrane, secondary lysosome, around small lipid droplet and cytoplasm. AKP positive reactiongranules were site in endoplasmic reticulum, myelin figure, mitochondrial membraneand cytoplasm. SOD positive reaction was found in mitochondrial membrane whichwas damaged seriously and densification of mitochondrial matrix. But there were fewSOD positive granules in the cytoplasm. POD positive reaction was still found inperoxisome which decreased in the number. And part of peroxisome was disaggregated.The activity site of PPO was not only in prophenoloxidase, but also in myelin figureand densification of mitochondrial matrix. But PPO positive activity wasn't observedin liver. The results were indicated that lysosomes in which ACP, AKP and PPOactivities were located destroyed and digested membrane structure in immunologicaldefense of L. vannamei. SOD and POD played an important role in eliminatingexcessive superoxide anion in shrimps. From the regularity changes of cytochemicallocation of ACP, AKP, SOD, POD and PPO of L. vannamei, we think that the fiveisozymes have positive effects on the growth and immunoreaction of L. vannamei.
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