• The Korean Journal of Zoology
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• v.39 no.1
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• pp.36-46
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• 1996

An endogenous phenoloxidase (EPO) from earthworm, Lumbricus rubellus, has been purified and characterized. The purified EPO using ammonium sulfate fractionation, Blue-2, Phenyl-, and Q-sepharose chromatography steps was revealed in SDS-PAGE as a single protein banri with Mr. of 59 kl)a. A native strudure of the enzyme was examined with an in situ staining of a nondenatudng-PAGE using DL-dopa as a substrate. The result showed that a single band due to the EPO activity was located siighdy above a standard polypeptide with Mr. of 210 kl)a. These fads indicate that the EPO is an oligomeric enzyme. The presence of a monophenolase activity of the purified EPO, which hydroxylates tyrosine to dopa, was confirmed by observing dopachrome accumulation at 475 nm at PH 8.0 with a typical lag phase during 60 mm. of meausrement. A series of inhibition study has been performed for the enzyme with several divalent cation chelators such as phenyithiourea (Flu), 1, lO-phenanthroline, EDTA, and EGTA. Among them, only V'flj inhibited the enzyme with 1C0.5 of 65 MM, which indicated that copper was critical for the catalysis of EPO. The enzyme was maximally active at 35'C and pH 8.0 when L-dopa to dopachrome conversion was spectrophotometricaily monitored at 475 nm. The apparent Km values of P0 for L-opa were obtained as 1.86 mM and 13.8 mM at pH 6.5 and 8.0, respectively. The catalytic efficiencies at both pH were almost identical [(kat/Km)pH8.0/(kcat/Km)pH6.5 = O.92] while the Vmax at p11 8.0 was 6.6-fold higher than that at pH 6.5. This fact may indicate that pH affeds the catalysis at substrate and/or enzyme-substrate complex level rather than the enzyme itself. Taken together, the EPO was an oligomeric enzyme which did not require proteolysis for its activation. These results also indicated that the enzyme can exist, at least, in part as a latent form In vivo, which might be distinct from the prophenoloxidase activating system. Therefore, it is pertinent to consider that there must be certain regulatory molecules or phenomena in L. rubellus which make the 1,0 in a latent form in vivo before the foreign invasions.

• Fisheries and Aquatic Sciences
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• v.26 no.11
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• pp.649-659
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• 2023

The present study compared the structure of mixilliped of Fenneropenaeus chinensis between the larval and adult stage and investigated the effect of the structural difference on the immunity of F. chinensis. A fourteen day and a one-month long culture trial were conducted each with postlarvae and adults of F. chinensis in the biofloc, mixed water (50% biofloc:50% clear seawater) and seawater control. Immune-related genes mRNA expressions of postlarvae was analysed by quantitative reverse transcription polymerase chain reaction (qRT-PCR). And the analysis of adult stage immunity was carried out using phenoloxidase (PO) enzyme activation in haemocyte. In the postlarvae, the final body weights were 51.43 and 58.47 mg for the biofloc water and the control seawater, respectively. On the other hand, the final body weights of the adults were significantly different between biofloc water and seawater. The survival rate showed the opposite trend to the growth rate. Immune related genes mRNA expression levels in the postlarvae in the biofloc water were significantly lower than those in the seawater. While, the adult stage showed significantly higher PO enzyme activations in the biofloc water than in the seawater with the PO enzyme activation increasing proportionally to the biofloc concentration. This result is considerably explained by the observations of setal morphological structures of the third maxilliped: postlarvae have short serrulated setae that compose the small 'net' structure while adults had long and dense plumose setae. It is understood that the morphological difference of the maxilliped structure resulted in the different feeding abilities in the postlarvae and the adult F. chinensis to use bioflocs as food source.

• The Korean Journal of Malacology
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• v.31 no.1
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• pp.1-8
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• 2015

This study was conducted to investigate the effects of elevated water temperature (WT) on biochemical and immunological factors in the hemolymph of the abalones, Haliotis discus hannai and H. discus discus. The abalone were exposed to various WT; 20, 22, 24, 26 and $28^{\circ}C$ for 4 days. In the control and $20^{\circ}C$, total-protein (TP), glucose and calcium (Ca) in hemolymph of H. discus discus were higher than the values in H. discus hannai. The values of magnesium (Mg), alkaline phosphatase (ALP) and lysozyme in H. discus hannai were similar to the H. discus discus in the control. There were no significant alterations in TP, glucose and Mg levels of hemolymph in H. discus hannai and H. discus discus by WT increases. The values of Ca, ALP and lysozyme were increased in H. discus hannai exposed to the high temperature (26 and $28^{\circ}C$) compared to control, while the values in H. discus discus were not significant difference between the WT groups. The phenoloxidase (PO) activity was increased in hemolymph of H. discus hannai exposed to high temperature (${\geq}24^{\circ}C$) compared to the control (P < 0.05). These physiological and immunological parameters were significantly changed in H. discus hannai. However, these parameters in H. discus discus were barely altered at the high WT (P < 0.05). These results suggested that H. discus hannai is considered to be more sensitive than H. discus discus at the high WT.

• International Journal of Industrial Entomology and Biomaterials
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• v.25 no.2
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• pp.139-146
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• 2012

Botanicals have been regarded as sustainable solution of pest control because of their nominal effect on nontarget pest, biodegradability and other environment friendly properties. However, study on impact of botanicals against beneficial insects is less. In this investigation effort has been made to study the effect of essential oils of Ocimum sanctum, Ocimum gratissimum and Ageratum conyzoides against non-mulberry silkworm Antheraea assama in terms of percent mortality, total haemocyte count (THC) and phenol oxidase (PO) enzyme activity. After application of essential oil of the plants by contact residual film technique, age dependent decrease of percent mortality was recorded. Topical treatment of oil at higher dosages ($1{\mu}lg^{-1}$ and $0.5{\mu}lg^{-1}$) caused 100% mortality of late instar larvae at 24 hour but the lowest dose ($0.25{\mu}lg^{-1}$) did not cause any mortality till 48 hour of treatment. Initial increase of total haemocyte count (THC) was recorded at early hours of treatment (0.25 hour) at all dosages (1, 0.5, $0.25{\mu}lg^{-1}$) after topical treatment. A dose dependent increase was observed in phenol oxidase enzyme activity and this might indicate ability of the essential oils to cause induction of immune response in Antheraea assama larvae.

• Korean journal of applied entomology
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• v.42 no.4
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• pp.353-360
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• 2003

Immunodepression can be required for entomopathogenic bacteria to induce their potent pathogenicities to the target insects. Here, we raise a hypothesis that the capacity of a pathogenic bacterium to induce the target insect immunodepression has positive relationship with the degree of pathogenicity. X. nematophilus had 1,200 times as potent as another entomopathogenic bacterium, Staphylococcus gallinarum against the fifth instar larvae of silkworm, Bombyx mori, when they were Injected into the hemocoel. Although both bacteria had significant cytotokic effect on the hemocytes of B. mori, X. nematophilus gave faster and greater cytotoxicity than did S. gallinarum. In cellular immune reactions, B. mori could form 20 hemocyte nodules against the bacterial injection with 5${\times}$10$\^$5/ cells. The number of the hemocyte nodules was significantly depressed when live X. nematophilus was inject-ed, but not in S. gallinarum. Activation of prophenoloxidase (proPO) was depressed in the bacterial injection. The depression of PO activation was significantly greater in X. nematophilus infection than in S. gallinarum injection. Lysozyme activity was induced by the injection of S. gallinarum at 4 h after the treatment, but not induced in X. nematophilus at all the time. These results showed that X. nemato-philus induced greater immunodepression against B. mori and resulted in higher pathogenicity than did S. gallinarum. Therefore, this study suggests that the immunodepression induced by entomopathogenic bacteria has positive relationship with their pathogenicity.

• Fisheries and Aquatic Sciences
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• v.24 no.2
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• pp.89-98
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• 2021

The accumulation of ammonia, a consequence of intensive aquaculture activity, can damage cultured animals. We analyzed the survival rates, growth, antioxidant responses, and immune responses of abalones, Haliotis discus hannai (mean shell length 70.2 ± 4.9 mm; mean body weight 36.9 ± 3.6 g), biweekly for four weeks, to determine the accumulated concentrations of ammonia (0, 0.1, 0.2, 0.4, 0.8, and 1.6 mg/L). In our biweekly measurements, a survival rate of ≥ 95% was observed for all concentrations of ammonia. The specific growth rate (SGR) decreased at ≥ 0.8 mg/L (p < 0.05), but the daily increment in shell length (DISL) showed no significant change (p < 0.05). Regarding antioxidant responses, the superoxide dismutase (SOD) activity of the hepatopancreas was increased at ammonia concentrations over 0.8 and 0.2 mg/L (p < 0.05) at two and four weeks, respectively, and the SOD activity of the gills increased at concentrations over 0.4 and 1.6 mg/L (p < 0.05) at four weeks. The catalase (CAT) activity of the hepatopancreas and gills increased at ammonia concentrations > 0.8 mg/L (p < 0.05). Phenoloxidase (PO) activity increased at ammonia concentrations over 0.8 mg/L (p < 0.05), and Lysozyme (LZM) increased at concentrations over 0.8 and 0.4 mg/L (p < 0.05). Overall, our findings indicated that ammonia concentrations over 0.8 mg/L in seawater might damage the SGR, antioxidant responses, and immune responses in H. discus hannai. Our findings suggest the necessity of improvement or reinstallation for abalone aquaculture systems and can be used to assess the toxic effects of ammonia on H. hannai.