博士 / 國立海洋大學 / 水產養殖學系 / 87 / A gram-positive, ovoid, diplococoid bacterium tentatively identified as Enterococcus sp. was isolated from diseased Macrobrachium rosenbergii farms in the southern Taiwan from July to October, 1994. The diseased prawns displayed poor growth, anorexia, inactivity, opaque and whitish musculature, and mortality. In histological preparations, melanized hemocytic granulomas were seen in the connective tissue around hemal sinuses together with hemocytic aggregation in necrotic musculature. Five isolates of diplococci were collected from diseased prawns at four farms. They were evaluated for 93 characteristics including morphology, physiology, biochemistry and sensitivity to antibiotics. The results indicated that the isolates belonged to an Enterococcus sp. closely related to Enterococcus seriolicida (synonymy with Lactococcus garvieae). Experimental injection of 3×105 cells of this strain (KM002) into the ventral sinus of the prawn cephalothorax caused 100% mortality in 11 days, and induced muscular necrosis and hepatopancreatitis, gross signs and histopathology similar to those observed in the naturally infected prawns. It was concluded that this Enterococcus sp. was the etiological agent associated with mortality of the farmed diseased prawns.The growth of this pathogenic strain (KM002) of Enterococcus sp. was examined in brain heart infusion broth (BHIB) at different pH levels, temperature, and NaCl concentrations. It grew at pH of 3-10, temperature of 5-45℃ and 0.5-6.5% of NaCl concentration with optimal level of pH 7-8, 25-30℃ and 0.5-1.0% NaCl concentration. M. rosenbergii was challenged by injection of either 1×107 or 2×106 cfu/prawn of KM002 previously incubated under these various conditions for 24 h, and monitored for pathology for 168 h. The results indicated that the pathogenic strain incubated at pH level of 7 and 8, temperature of 27℃ and 30℃ and NaCl concentration of 0.5% and 1.0% caused higher mortality of M. rosenbergii. M. rosenbergii were challenged with an Enterococcus sp. (KM002). Challenge doses and conditions of pH, salinity and temperature were varied to determine the influence of environmental factors on the development of disease and mortality. In pH tests, the onset of mortality was earlier and the cumulative mortality was higher at pH 8.8-9.5 than at pH 4.6-5.2 and 7.5-7.7. In salinity tests at two challenge doses (1×106 and 2×107 cfu/prawn), onset of mortality was earliest at 15 ppt and cumulative mortality was low at 5 ppt and 10 ppt. In temperature tests at pH 7.2-7.5 and at two challenge doses (2×107 and 4×107 cfu/prawn), the onset of mortality was earliest at 33-34℃ and total mortality occurred at 27-28℃ and 33-34℃. Reducing the challenge dose to 5×104 cfu/prawn gave lowest survival at 33-34℃ group. In similar temperature tests but at pH 8.8-9.5, onset of mortality was accelerated and there was 100% death for all the high challenged groups. At low challenge doses, mortality was lower but still highest in the 33-34℃ group. Results indicated that mortality of M. rosenbergii caused by this Enterococcus sp. was exacerbated by higher temperature of 33-34℃ and higher pH level of 8.8-9.5. By contrast, low salinity appeared to have a beneficial effect on survival.The total hemocyte count and phenoloxidase activity were examined for M. rosenbergii that was reared at different conditions of pH, salinity and temperature and feeding rate. In pH tests, the total hemocyte count was significantly higher at pH 7.5-7.7 than at pH 9.0-9.5 (p<0.05) and pH 4.6-5.0 (p<0.1). The hemocyte phenoloxidase activity was significantly different among different pH levels (p<0.05) with an order of pH 7.5-7.7 > pH 4.6-5.0 > pH 9.0-9.5. In temperature tests, the total hemocyte count was significantly lower at 33-34℃ than 27-28 and 30-31℃ (p<0.05). The hemocyte phenoloxidase activity was significantly different among different temperature levels (p<0.05) and was 30-31℃ > 27-28℃ > 33-34℃. In salinity tests, total hemocytes count was directly related to salinity level. Hemocyte phenoloxidase activity was significantly higher at 5 and 10 ppt than at 0 and 15 ppt (p<0.05). In temperature tests at a same feeding rate (0.6%), the total hemocyte count was significantly higher at 27-28℃ than at 20-21℃ and 33-34℃ (p<0.05). Hemocyte phenoloxidase activity was significantly higher at 27-28℃ than at 20-21℃ and 33-34℃ (p<0.05). In the test of different feeding rate at 27-28℃, the total hemocyte count was significantly lower at 0.1% than at 0.5% (p<0.05), 1.0% and 1.5% (p<0.1). Hemocyte phenoloxidase activity was significantly lower at 0.1% of feeding rate than those above 0.5% (p<0.05). Results indicated that the total hemocyte count and phenoloxidase activity of M. rosenbergii varied at different pH, temperature, salinity levels and different feeding rates. It was concluded that the phenoloxidase activity rather than total hemocyte count is considered to be a healthy parameter of M. rosenbergii.
| Identifer | oai:union.ndltd.org:TW/087NTOU0086028 |
| Date | January 1998 |
| Creators | CHeng Winton, 鄭文騰 |
| Contributors | Chen Jiann-Chu, 陳建初 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 93 |
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