【作者】 丁森；

【导师】 董双林； 王芳；

【作者基本信息】 中国海洋大学 ， 水产养殖， 2010， 博士

【摘要】 本研究以凡纳滨对虾(Litopenaeus vannamei)稚虾为实验材料,在海水条件和淡水条件下,初步研究比较了蜕壳周期中凡纳滨对虾体组成和血液组成、非特异性免疫、呼吸代谢、消化水平和渗透调节的变化。主要研究结果如下：1海水和淡水条件下不同蜕壳时期凡纳滨对虾体组成和血淋巴组成的比较本文在实验室条件下研究了不同蜕壳时期凡纳滨对虾(Litopenaeus vannamei)(生物学体长4-5 cm)在海水和淡水养殖环境中体组成份、血淋巴成份的变化。实验结果表明：海水与淡水条件下对虾肌肉中粗蛋白含量A期最低,C期最高,且差异显著；而肌肉中粗脂肪含量无显著变化。与海水条件相比,淡水条件下对虾肌肉中粗蛋白含量整体水平显著下降,脂肪和水分含量显著上升。在海水条件下对虾肝胰脏中粗蛋白含量表现出A期较低,C期较高的变化,粗脂肪含量则C期显著低于其它时期；而淡水条件下肝胰脏中粗蛋白含量则表现为蜕壳后期和间期较低,蜕壳前期显著提高的变化,粗脂肪含量无显著变化。对虾肝胰脏粗蛋白、粗脂肪和水分含量整体水平在两种条件下未表现出显著性差异。海水条件下对虾肌肉中蛋氨酸和丙氨酸在不同蜕壳时期出现显著变化,而淡水条件下对虾肌肉中缬氨酸和丝氨酸在不同蜕壳时期发生了显著变化。与海水条件下相比,淡水条件下对虾肌肉中甘氨酸、丙氨酸、半胱氨酸、缬氨酸、蛋氨酸、络氨酸、苯丙氨酸与组氨酸含量整体水平发现了显著变化。在两种条件下,对虾血淋巴中蛋白浓度在不同蜕壳时期均呈现逐渐上升的趋势,即A期蛋白浓度最低,在D3期蛋白浓度最高；而对虾血糖浓度均呈现先下降再升高的趋势,在A和B期血糖浓度较高,由蜕壳间期至D2期逐渐降低,在D3期出现上升。与海水条件相比,淡水条件下对虾血淋巴蛋白和血糖含量整体水平均显著下降。海水条件下,对虾血淋巴中钙和镁元素含量从蜕壳后期到蜕壳前期都呈现逐渐下降的趋势,蜕壳后期含量最高；钾元素含量变化不大；磷元素含量则呈先降低后升高的趋势,在蜕壳间期含量最低。淡水条件下,对虾血淋巴中钙和镁元素含量A期较低,从B期上升,C期后逐渐下降；钾元素含量的变化趋势与钙、镁元素相似；磷元素含量则先升后降,峰值出现在D0期。与海水条件相比,淡水条件下对虾血淋巴中钾和钙元素含量整体水平显著提高,磷元素含量整体水平显著下降,镁元素含量整体水平无显著变化。2海水和淡水条件下不同蜕壳时期凡纳滨对虾非特异性免疫的比较本文在实验室条件下研究了海水和淡水两种条件下不同蜕壳时期(A、B、C、Do-D3)凡纳滨对虾(Litopenaeus vannamei)血细胞数量、酚氧化酶、呼吸爆发、一氧化氮合酶和溶菌酶活力的变化。实验结果表明：1、海水和淡水条件下对虾蜕壳后,血细胞数量均呈逐渐增加的趋势,海水条件下D3期血细胞数量(THC)显著高于蜕壳后期(A期)(P<0.05),而淡水条件下各个时期对虾THC无显著差异。与海水条件相比,淡水条件下对虾THC整体水平显著高于海水条件下的水平(P<0.05)。2、海水和淡水条件下对虾酚氧化酶(PO)活力都在蜕壳间期出现峰值,分别为87.5 U/min和45.0 U/min,蜕壳后期和前期PO活力相对较低。淡水条件下对虾PO活力整体水平显著低于海水条件下的活力(P<0.05)。3、海水和淡水条件下对虾呼吸爆发(RB)均表现出与THC相似的变化规律,在蜕壳前期(尤其是D3期)RB最大。淡水条件下对虾RB整体水平显著低于海水条件下的水平(P<0.05)。4、海水和淡水条件下对虾一氧化氮合酶(NOS)活力均表现出在蜕壳前期高,在蜕壳后期和间期相对较低。淡水条件下对虾一氧化氮合酶活力整体水平显著低于海水条件下的活力(P<0.05)。5、海水和淡水条件下对虾溶菌酶活力各个蜕壳时期的变化都不大,均在蜕壳间期稍高,但海水中对虾溶菌酶活力整体水平显著高于淡水条件下的活力(P<0.05)。3海水和淡水养殖条件下不同蜕壳时期凡纳滨对虾呼吸代谢的比较本文在实验室条件下研究了不同蜕壳时期凡纳滨对虾(Litopenaeus vannamei)(生物学体长4-5 cmm)在海水和淡水养殖环境中耗氧率、排氨率、排尿素率及丙酮酸激酶、乳酸脱氢酶两种代谢酶活力的变化。主要实验结果如下：1、海水养殖条件下,凡纳滨对虾在蜕壳发生前后耗氧率相对上升,D3期和蜕壳后期(A和B期)耗氧率较高,分别为0.667mg·g-1·h-1、0.696mg·g-1·h-1和0.727mg·g-1·h-1, C、D0和D1期对虾耗氧率水平在0.567-0.581mg·g-1·h-1之间,D2期耗氧率最低(0.488mg·g-1·h-1);淡水条件下,对虾耗氧率变化趋势与海水基本相似,在蜕壳前后耗氧率较高,A和B期对虾耗氧率分别为0.651 mg·g-1·h-1和0.650mg·g-1·h-1,其它蜕壳时期对虾耗氧率显著降低(P<0.05)。海水环境中对虾耗氧率的整体水平显著高于淡水环境中对虾耗氧率的整体水平(P<0.05)。2、海水养殖条件下,A和B期对虾的排氨率最高(分别为12.273μg·g-1·h-1和10.644μg·g-1·h-1),显著高于C期的排氨率(4.574μg·g-1·h-1)(P<0.05),在Do-D2期有小幅度的上升,到D3期则降至最低(2.969μg·g-1·h-1);淡水条件下,对虾蜕壳后期的(A和B期分别为40.501μg·g-1·h-1和31.164μg·g-1·h-1)排氨率最高,其它蜕壳时期显著降低(P<0.05),且排氨率整体水平远高于海水条件下的排氨率(P<0.05)。3、海水养殖条件下,对虾排尿素率蜕壳前后较低,D0期较高,且差异显著(P<0.05)；淡水条件下则正好相反,蜕壳前后相对较高,C期较低,且差异显著(P<0.05),海水环境中对虾排尿素率整体水平显著高于淡水环境中对虾排尿素率整体水平(P<0.05)。4、海水养殖条件下,A和B期对虾丙酮酸激酶活力分别为150.67 U/gprot和164.50 U/gprot, C期酶活力下降,在D1和D2达到最高(219.19 U/gprot和233.30 U/gprot), D3期酶活力最低；淡水条件下,对虾丙酮酸激酶活力在B期最高(116.77 U/gprot),随后逐渐下降,在蜕壳前期活力较低,与海水环境相比整体水平显著降低(P<0.05)。5、海水养殖条件下,对虾乳酸脱氢酶活力在B期最高(2587.99 U/gprot),随后逐渐下降,至D3期活力达到最低(1851.02 U/gprot);淡水条件下的整体变化趋势与海水相似,最高酶活力出现在D0期(4376.15 U/gprot),至D3期达到最低(1159.55 U/gprot),整体酶活力较海水环境有所提高。海水与淡水环境中对虾乳酸脱氢酶活力整体水平差异不显著(P>0.05)。4海水和淡水养殖条件下不同蜕壳时期凡纳滨对虾消化生理的比较本文在实验室条件下研究了不同蜕壳时期凡纳滨对虾(Litopenaeus vannamei)在海水和淡水养殖条件下胃蛋白酶活力、类胰蛋白酶活力、脂肪酶活力及淀粉酶活力的变化。主要实验结果如下：1、两种养殖条件下对虾胃蛋白酶活力随蜕壳时期的变化趋势相似,在蜕壳后期酶活力水平较高,蜕壳间期下降,蜕壳前期最低。两种养殖条件下对虾胃蛋白酶活力差异不显著(P>0.05)。2、海水养殖条件下,蜕壳后期(A期)对虾类胰蛋白酶活力最高(4.556 U·mg prot-1),蜕壳间期酶活力也保持较高的水平(4.420 U·mg prot-1),蜕壳前期酶活力在3.462-3.820 U·mg prot-1之间。淡水养殖条件下,蜕壳间期对虾酶活力为1.950U·mgprot-1,显著高于其它时期酶活力(P<0.05)。海水养殖条件下对虾类胰蛋白酶活力整体水平显著高于淡水条件下的活力(P<0.05)。3、海水养殖条件下对虾脂肪酶活力在蜕壳后期(A和B期)最高,分别为6.919 U·mg prot-1和7.531 U·mg prot-1,蜕壳间期略下降,蜕壳前期酶活力有所下降,在4.636-5.258 U·mg prot-1之间。淡水养殖条件下对虾脂肪酶活力在蜕壳后期较高,蜕壳间期最高(8.720 U·mg prot-1),蜕壳前期酶活力下降至3.523-5.813 U·mg prot-1之间。海水与淡水养殖条件下对虾脂肪酶活力整体水平差异不显著(P>0.05)。4、海水养殖条件下,对虾淀粉酶活力在蜕壳后期A和B期分别为1.525 U·mg prot-1和1.398 U·mg prot-1,显著高于其它时期酶活力(P<0.05)。淡水养殖条件下,对虾淀粉酶活力在蜕壳间期最高(2.954 U·mg prot-1),显著高于其它时期酶活力(P<0.05)。海水养殖条件下对虾淀粉酶活力整体水平显著低于淡水条件下的活力(P<0.05)。5海水和淡水养殖条件下不同蜕壳时期凡纳滨对虾渗透调节的比较本文在实验室条件下研究了不同蜕壳时期凡纳滨对虾(Litopenaeus vannamei)在海水和淡水养殖环境中血浆渗透压、鳃丝离子转运酶活力和血蓝蛋白含量的变化。结果发现：1、两种条件下血浆渗透压的调节规律不同,海水条件下蜕壳前期(D2期和D3期)和蜕壳后期(A期和B期)血浆渗透压处于较高水平,蜕壳间期调节至较低水平；淡水条件下蜕壳后期(A期和B期)对虾血浆渗透压最低,之后逐渐升高。受到两种条件下渗透浓度的影响,海水条件下对虾血浆渗透压整体水平显著高于淡水条件下的水平(P<0.05)。2、海水条件下对虾Na+-K+-ATPase活力在蜕壳前期(D2期和D3期)和蜕壳后期(A期和B期)较低,蜕壳间期较高；淡水条件下对虾Na+-K+-ATPase活力在A期最高,后逐渐下降,至D3期达最低水平。两种条件下对虾Na+-K+-ATPase活力和血浆渗透压在蜕壳周期中的变化趋势相反。3、两种条件下对虾碳酸酐酶活力在蜕壳周期中的变化趋势相似,即蜕壳后期较高,其它时期较低,D3期有小幅度地提高。海水与淡水条件下对虾两种离子转运酶活力整体水平均没有显著性差异(P>0.05)。4、两种条件下对虾血蓝蛋白含量在蜕壳周期中变化也不同,海水条件下血蓝蛋白含量在蜕壳后期最低,随后逐渐增加,至蜕壳前期达到最高水平；淡水条件下血蓝蛋白则变化不大,各时期没有显著性差异(P>0.05)。海水条件下对虾血蓝蛋白含量整体水平显著高于淡水条件下的水平(P<0.05)。更多还原

【Abstract】 A series of indoor trials were conducted to investigate the comparison of body composition, nonspecific immunity, respiratory metabolism, digestive physiology and osmoregulation in juvenile Litopenaeus vannamei cultured in seawater and freshwater conditions in relation to molt stages. The primary results were listed below.1 Comparision of body and hemolymph composition of Litopenaeus vannamei under seawater and freshwater conditions in relation to the molt stagesThis experiment was conducted to investigate the changes of body and hemolymph compsitions of Litopenaeus vannamei under seawater and freshwater conditions in relation to the molt stages. The results showed that the crude protein contents in muscle of test shrimp were the lowest and the highest at A stage and C stage, respectively, under both seawater and freshwater conditions. The crude lipid contents in muscle showed no significant differences in a molt cycle under two conditions. Compared with the seawater conditions, the whole level of crude protein contents in muscle increased significantly under freshwater conditions, meanwhile, the whole level of crude lipid and moisture contents decreased significantly. The crude protein contents in hepatopancreas were lower at A stage and were higher at C stage under seawater conditions. The crude lipid contents in hepatopancreas at C stage were significantly lower than those at other stages. Under freshwater conditions, the crude protein contents in hepatopancreas showed lower level at post- and inter-molt stages and exhibited higher at pre-molt stage, while the crude lipid contents showed no significant differences among all stages. The whole level of crude protein, crude lipid and moisture contents exhibited no significant differences between two conditions. Methionine and alanine contents in muscle showed significant differences among all stages under seawater conditions, while valine and serine contents in muscle exhibited significant differences among all stages under freshwater conditions. Compared with the seawater conditions, the whole level of glycine, alanine, cysteine, valine, methionone, tyrosine, phenylalanine and histidine contents in muscle changed significantly under freshwater conditions. Under both conditions, the protein concentration in hemolymph increased gradually from post-molt stage to pre-molt stage, whereas the glucose concentration in hemolymph showed higher at post-molt stage, decreased at inter-molt stage, and increased at D3 stage. Compared with the seawater conditions, the whole level of protein and glucose concentration in hemolymph decreased significantly under freshwater conditions. The calcium and magnesium contents in hemolymph decreased during molt cycle under seawater conditions. The lowest phosphorus contents in hemolymph occurred at C stage, whereas it exhibited high level at premolt and postmolt stages. Under freshwater conditions, the calcium and magnesium contents in hemolymph showed lower at A stage, increased at B stage and decreased gradually after C stage. Kalium contents in hemolymph showed the same trend as calcium and magnesium. The higher contents of Phosphor exhibited at Do stage. Compared with the seawater conditions, the whole level of kalium and calcium contents in hemolymph increased significantly under freshwater conditions, whereas the whole level of phosphor contents in hemolymph decreased significantly, but no significantly changed was found in magnesium contents.2 Comparision of nonspecific immunity of Litopenaeus vannamei under seawater and freshwater conditions in relation to the molt stagesThis experiment was conducted to compare total hemocytes counts (THC), phenol oxidase (PO) activities, respiratory burst (RB), nitric oxide synthase (NOS) activities and Lysozyme (LY) activities of Litopenaeus vannamei under seawater and freshwater conditions in relation to the the molt stages. It was found that:1) THC of experimental shrimp increased from post-molt stage to pre-molt stage in both seawater and freshwater conditions. THC of D3 stage was significantly higher than that of A stage under seawater conditions (P<0.05), whereas no significant differences were found among all stages under freshwater conditions (P>0.05).2) PO activity of experimental shrimp breeding in seawater and freshwater conditions reached peak at inter-molt stage and was 87.5 U/min and 45.0 U/min, respectively, and the lower levels of PO activity occurred at post-molt and pre-molt stages. The whole level of PO activity of experimental shrimp breeding in freshwater conditions was significantly lower than that in seawater conditions (P<0.05).3) The change trend of RB was similar to that of THC under seawater and freshwater conditions, and increased to the peak at pre-molt stage, especially D3 stage. The whole level of RB of experimental shrimp breeding in freshwater conditions was significantly lower than that in seawater conditions (P<0.05).4) NOS activities of experimental shrimp breeding in both seawater and freshwater conditions were higher level at pre-molt stage, whereas lower level occurred at inter-molt and pre-molt stage. The whole level of NOS activity of experimental shrimp breeding in freshwater conditions was significantly lower than that in seawater conditions (P<0.05).5) LY activity of experimental shrimp breeding in seawater and freshwater conditions changed an extent, and the highest levels were found at inter-molt stage. The whole level of LY activity in seawater was significantly higher than in freshwater conditions (P<0.05).3 Comparision of respiratory metabolism of Litopenaeus vannamei under seawater and freshwater conditions in relation to the molt stagesThis experiment was conducted to compare the oxygen consumption rates (OCR), ammonia excretion rates (AER), urea excretion rates (UER), pyruvate kinase activities (PK) and lactate dehydrogenase activities (LDH) of Litopenaeus vannamei under seawater and freshwater conditions in relation to the molt stages. It was found that:1) Under seawater condition, the high level of the OCR occurred around the ecdysis. The OCR of D3 stage and post-molt stage (A and B stages) increased and was 0.667 mg·g-1·h-1,0.696 mg·g-1·h-1 and 0.727 mg·g-1·h-1, respectively, the level of OCR of C, Do and D1 stages were between 0.567-0.581 mg·g-1·h-1, and the lowest level of OCR occurred at D2 stage (0.488 mg·g-1·h-1). Under freshwater conditions, the fluctuation trend of OCR was similar to that one under seawater conditions. The OCR of A and B stages was 0.651 mg·g-1·h-1 and 0.650 mg·g-1·h-1, respectively, and significantly higher than those at other stages (P<0.05). The whole level of OCR of shrimp cultured in seawater conditions was higher than that in freshwater conditions (P<0.05).2) Under seawater conditions, AER of shrimp at A and B stages (was 12.273μg·g-1·h-1 and 10.644μg·g-1·h-1, respectively) and these were significantly higher than that at C stage (4.574μg·g-1·h-1) (P<0.05), and AER increased with a certain extent at Do-D2 stages, then decreased to the lowest level at D3 stage (2.969μg·g-1·h-1). Under freshwater conditions, the highest level (40.501μg·g-1·h-1 and 31.164μg·g-1·h-1) of AER was occurred at post-molt stage, and significantly higher than those at other stages (P<0.05). The whole level of AER of shrimp cultured in seawater conditions was lower than that in freshwater conditions (P<0.05).3) Under seawater conditions, the UER of shrimp decreased around the ecdysis, reached the highest level at DO stage, and there were significant differences between them (P<0.05). Oppositely, under freshwater conditions, the UER of shrimp increased at pre- and post-molt stages, decreased at inter-molt stage and significant differences were found between them (P<0.05). The whole level of UER of shrimp cultured in seawater conditions was significantly higher than that in freshwater conditions (P<0.05).4) Under seawater conditions, PK activity of shrimp at A and B stages was 150.67 U/gprot and 164.50 U/gprot, respectively, decreased at C stage, and then reached the higher level at D1 and D2 stages (219.19 U/gprot and 233.30 U/gprot), finally decreased to the lowest level at D3 stage. Under freshwater conditions, PK activity of shrimp was highest at B stage (116.77 U/gprot), then decreased gradually with the passage of time, the lowest level occurred at pre-molt stage. Compared with the seawater conditions, the whole level of PK of shrimp cultured in freshwater conditions decreased significantly (P<0.05).5) Under seawater conditions, LDH activity of shrimp reached the peak at B stage (2587.99 U/gprot) and decreased gradually, while the lowest level occurred at D3 stage (1851.02 U/gprot). The fluctuation trend under freshwater condition was similar to that under seawater conditions. The highest level occurred at DO stage (4376.15 U/gprot), while the lowest level was showed at D3 stage (1159.55 U/gprot). There were no significant differences of the whole level of LDH activities between the two conditions.4 Comparision of digestive physiology of Litopenaeus vannamei under seawater and freshwater conditions in relation to the molt stagesThis experiment was conducted to compare pepsin activities, tryptase activities, lipase activities and amylase activities of Litopenaeus vannamei under seawater and freshwater conditions in relation to the molt stages. It was found that:1) the changing tendency of pepsin activities of experimental shrimp cultured in seawater and freshwater conditions was similar in a molt cycle. The higher level of pepsin activities occurred in post-molt stage, and then decreased at inter-molt stage, while the lowest level of that was found at pre-molt stage. There were no significant differences of the whole level of pepsin activities between the two conditions (P>0.05).2) Under seawater conditions, tryptase activity of experimental shrimp was the highest at A stage (4.556 U/mg prot), and tryptase activities were 3.462-3.820 U/mg prot at pre-molt stage. Under freshwater condition, tryptase activity at inter-molt stage was 1.950 U/mg prot and was significantly higher than those at other stages (P<0.05). The whole level of tryptase activity of experimental shrimp cultured in seawater conditions was higher than that in freshwater conditions (P<0.05).3) Under seawater conditions, lipase activities of experimental shrimp kept higher level at post-molt stage (A and B stages) and were 6.919 U/mg prot and 7.531 U/mg prot, respectively, and then decreased at inter-molt stage, and the lower level of lipase activities were found at pre-molt stage which were 4.646-5.258 U/mg prot. Under freshwater conditions, lipase activities of experimental shrimp were higher at post-molt stage, and the highest level was found at inter-molt stage (8.720 U/mg prot), and then decreased to 3.523-5.813 U/mg prot at pre-molt stage. No significant differences of the whole level of lipase activities were found between the two conditions (P>0.05).4) Under seawater conditions, amylase activity of experimental shrimp at A and B stages was 1.525 U/mg prot and 1.398 U/mg prot, respectively,and significantly higher than those at other stages (P<0.05). Under freshwater conditions, lipase activity of experimental shrimp reached the peak at inter-molt stage (2.954 U/mg prot), and was significantly higher than those at other stages (P<0.05). The whole level of amylase activity of experimental shrimp breeding in seawater conditions was 1.124 U/mg prot, and significantly higher than that in freshwater conditions.5 Comparision of osmoregulation of Litopenaeus vannamei under seawater and freshwater conditions in relation to the molt stagesThis experiment was conducted to investigate the changes of osmolarity, hemocyanin concentration, Na+-K+-ATPase activity and carbonic anhydrase activity of of Litopenaeus vannamei under seawater and freshwater conditions in relation to the molt stages. The results showed that the changing trends of osmolarity were different under seawater and freshwater conditions. Under seawater conditions, the osmolarity of test shrimp kept higher level at pre-molt stage (D2 and D3 stages) and post-molt stage (A and B stages), whereas lower level occurred at inter-molt stage. Under freshwater conditions, the osmolarity of test shrimp kept lower level at post-molt stage (A and B stages), increased from inter-molt stage. The whole level of osmolarity of L. vannamei cultured in seawater conditions was significantly higher than that in freshwater conditions. There were two different changing trends of hemocyanin concentration of test shrimp under two conditions. Under seawater conditions, the hemocyanin concentration was lower at post-molt stage and increased gradually after that. There were no significant differences of hemocyanin concentration among all molt stages under freshwater conditions. The whole level of hemocyanin concentration of L. vannamei cultured in seawater conditions was also significantly higher than that in freshwater conditions. Under seawater conditions, Na+-K+-ATPase activity in gills kept lower at pre-molt stage (D2 and D3 stages) and post-molt stage (A and B stages), whereas higher level occurred at inter-molt stage. Under freshwater conditions, Na+-K+-ATPase activity in gills was the highest at A stage, and then decreased gradually, and reached the lowest at D3 stage. The trend of Na+-K+-ATPase activity was opposite to the one of osmolarity during the molt cycle under both seawater and freshwater conditions. The changing trends of carbonic anhydrase activity during the molt cycle were similar under two conditions. Carbonic anhydrase activity was higher at post-molt stage, and lower level occurred at other stages and narrowly increased at D3 stage. The whole level of two ion transport enzymes did not show significant differences between seawater and freshwater conditions.更多还原