氨氮胁迫对三疣梭子蟹酚氧化酶原系统和免疫指标的影响

以三疣梭子蟹(Portunus trituberculatus)为研究对象,研究氨氮胁迫对其酚氧化酶原系统和免疫指标的影响.实验分0mg/L、1 mg,L、5mg,L、20mg,L氨氮处理组,分别于0h、6h、12 h、24 h、48 h取样并进行相关指标测定.结果表明:氨氮胁迫对三疣梭子蟹血淋巴多巴胺含量、血细胞数量、血细胞酚氧化酶原(proPO)活力和mRNA表达以及抗菌、溶菌活力的影响显著(P<0.05).各处理组血淋巴多巴胺含量于6 h达到最大值,12 h后恢复至对照组水平;各处理组血细胞数量均呈明显下降趋势,除20mg/L氨氮处理组半颗粒细胞、总细胞数量表现为逐渐下降趋势外,其他处理组3类血细胞和总细胞数量分别于12h或24h时降至最低值,然后保持稳定.各处理组上述指标分别呈明显下降、升高、下降和峰值变化趋势,其中1mg/L处理组血细胞proPO活力在12 h后恢复至对照组水平,而mRNA表达与对照组差异不显著,其他处理组血细胞proPO活力和mRNA表达分别于12h、6h后保持稳定;各处理组抗菌活力于6h或12h达到最小值,然后恢复至对照组水平,而溶菌活力在24 h后趋于稳定,且1 mg/L处理组与对照组无明显差异.各处理组血细胞数量、proPO活力、溶菌活力在稳定后均与氨氮浓度呈明显的负相关性,而血细胞proPO mRNA表达呈显著正相关性.由此可见,三疣梭子蟹在氨氮胁迫下血细胞数量、proPO活力和mRNA表达以及免疫指标表现出明显的时间剂量效应性,具有显著的免疫毒性效应.本研究旨在为三疣梭子蟹养殖水环境调控和病害防治提供基础依据.     

双齿围沙蚕与赤子爱胜蚓对凡纳滨对虾生长和免疫指标的影响

以鲜活饵料双齿围沙蚕（Perinereis aibuhitensis）、赤子爱胜蚓（Eisenia foetida）和人工配合饵料投喂凡纳滨对虾（Litopenaeus vannamei），并测定各种饵料对其生长和免疫指标的影响。结果表明，沙蚕和蚯蚓单独投喂或与人工配合饵料配合投喂都可显著提高凡纳滨对虾的生长速率，但用沙蚕单独投喂的对虾成活率较低。蚯蚓与人工配合饵料配合投喂可显著提高对虾血清中抗菌活力、溶菌活力、酚氧化酶活力和血细胞数量；蚯蚓单独投喂可显著提高血清中抗菌活力、酚氧化酶活力和血细胞数量，对血清中溶菌活力作用不明显。沙蚕与人工配合饵料配合投喂可显著提高对虾血清中抗菌和溶菌活力．但对血清中酚氧化酶活力和血细胞数量变化作用不明显；单独投喂沙蚕对所测各项免疫指标影响均不显著。研究结果证实：利用蚯蚓和沙蚕可显著提高对虾的生长速率，但蚯蚓的饵料效果优于沙蚕。[中国水产科学，2006，13（4）：561—565]     

脂多糖、多巴胺与蛋白激酶抑制剂联合作用对凡纳滨对虾血细胞吞噬率、胞吐酚氧化酶活力的影响

研究了脂多糖(LPS)、多巴胺(DA)对凡纳滨对虾血细胞吞噬、胞吐及信号通路的影响。结果表明,LPS、DA对凡纳滨对虾血细胞数量、吞噬率和胞吐酚氧化酶活力影响显著(P<0.05),LPS、DA与血细胞孵育30min后,高浓度(1～10mg/L或μmol/L)处理组对虾血细胞数量均随作用浓度增大而明显下降,胞吐酚氧化酶活力则显著升高,LPS处理组对虾血细胞吞噬率显著升高,而DA处理组对虾血细胞吞噬率则表现出明显的下降趋势。同时在LPS作用下,分别加入蛋白激酶C(PKC)、酪氨酸蛋白激酶(TPK)抑制剂chelerythrine、genistein后,凡纳滨对虾血细胞吞噬和胞吐作用均受到明显的抑制作用,抑制剂对吞噬的抑制效果为genistein>chelerythrine,对胞吐的抑制效果则为chelerythrine>genistein;在DA作用下,加入蛋白激酶A(PKA)抑制剂H-89后,血细胞吞噬作用得到增强,但对胞吐作用无明显影响,chelerythrine、genistein均抑制对虾血细胞的胞吐作用,抑制剂的作用效果为chelerythrine>genistein,两种抑制剂对吞噬均无显著影响。     

氨氮促使携带白斑综合症病毒凡纳滨对虾发病及其血细胞、酚氧化酶和过氧化氢酶变化

通过白斑综合症病毒(WSSV)感染得到携带WSSV病毒的凡纳滨对虾,它们由0.05 mg/L氨氮水体进入不同氨氮浓度水体,在7 d内,1.75 mg/L,2.25 mg/L,3.00 mg/L氨氮水体内个体发生病毒病,并产生死亡。在14 d内,0.75 mg/L氨氮以及大于这一浓度水体内个体发生病毒病,并产生死亡,其中3.00 mg/L氨氮水体内死亡100%,2.25 mg/L氨氮水体内死亡80%,1.75 mg/L水体内死亡60%。在0.35 mg/L氨氮水体,7-14 d内对虾开始发病,没产生死亡。发病个体血细胞数量下降迅速,血清酚氧化酶先增高再迅速下降,其中3 mg/L氨氮水体中的携带病毒对虾只有下降。同时氨氮浓度越大,携带病毒凡纳滨对虾血清过氧化氢酶活力下降越快,越多。     

细菌J-10对凡纳滨对虾非特异性免疫指标的影响

以凡纳滨对虾(Litopenaeus vannamei boone)为试验材料,将细菌J-10添加到饲料中投喂对虾,测定其对凡纳滨对虾血清酚氧化酶(PO)、抗菌活力(Ua)和溶菌活力(U1)、溶血素活性和血清中蛋白含量的影响。结果显示,试验组各项指标均优于对照组,用副溶血弧菌进行人工攻毒,免疫保护率达到40％。结果表明,细菌J-10可以增强凡纳滨对虾的非特异性免疫功能。     

一种微生物絮团的生化分析及其对凡纳滨对虾免疫力的影响

利用红糖与尿素为碳氮源在自然海水中培养微生物絮团,获得絮团产物,其离心后进行初步的生化分析表明,絮团产物上清液中微生物胞外产物重均分子量为213 281 u.絮团沉淀物中多糖含量占29.6％,氨基酸含量占12.6％.将絮团产物按0、0.02％、0.1％、0.5％、2.5％的比例添加至低蛋白饵料中投喂凡纳滨对虾,14 d后分别测定实验对虾血清溶菌活力、抗菌活力和酚氧化酶活力,结果显示,在饵料中添加微生物絮团浓度为2.5％的对虾血清中抗菌与溶菌活力最高,添加微生物絮团浓度为0.5％与2.5％的对虾血清中酚氧化酶活力较低蛋白饵料对照组显著提高.用哈维氏弧菌感染实验对虾后,饵料中添加0.1％微生物絮团产物组对虾的死亡率最低.综合分析认为,凡纳滨对虾摄食微生物絮团后,能够显著提高对虾的非特异免疫力,抗微生物感染的能力得到增强.     

一种微生物絮团的生化分析及其对对虾免疫力的影响

利用红糖与尿素为碳氮源在自然海水中培养微生物絮团,获得絮团产物,对该产物离心后进行初步的生化分析表明,絮团产物上清液中微生物胞外产物重均分子量为213281Da。絮团沉淀物中多糖含量占29.6%,氨基酸含量占12.6%。将絮团产物按0、0.02、0.1、0.5、2.5%的比例添加至低蛋白饲料中投喂凡纳滨对虾 (Litopenaeus vannamei),14d后分别测定实验对虾血清溶菌活力、抗菌活力和酚氧化酶活力,结果显示在饲料中添加微生物絮团浓度为2.5%的对虾血清中抗菌与溶菌活力最高(P<0.05),添加微生物絮团浓度为0.5%与2.5%的对虾血清中酚氧化酶活力较低蛋白饵料对照组显著提高(P<0.05)。用哈维氏弧菌(Vibrio harveyi)感染实验对虾后,结果显示饵料中添加0.1%微生物絮团产物组对虾的死亡率最低。综合分析认为对虾摄食微生物絮团后,能够显著提高对虾的非特异免疫力,抗微生物感染的能力得到增强。     

颤藻对凡纳滨对虾生长和免疫相关酶活力的影响

颤藻(Oscillatoria sp.)是对虾养殖水体中的一种常见蓝藻。通过在凡纳滨对虾养殖水体中投加不同浓度的颤藻,测定凡纳滨对虾的成活率与体长、体重,以及不同颤藻浓度下凡纳滨对虾的免疫相关酶活力,研究颤藻对凡纳滨对虾生长和免疫酶活力的影响。结果表明:颤藻对对虾的成活率、体长和体重有显著影响(P<0.05),随藻浓度增加对虾的成活率、体长和体重呈下降的趋势。颤藻对对虾超氧化物歧化酶(SOD)、过氧化物酶(POD)、酚氧化酶(PO)、碱性磷酸酶(ALP)、谷胱甘肽S-转移酶(GST)、抗菌活力影响显著(P<0.05),低浓度的颤藻诱导酶活力增大,当浓度大于12.5 mg/L时,其酶活力呈下降的趋势。当颤藻浓度为2.5 m g/L和12.5 mg/L时,GST、ALP活力分别达到最大值(39.05 U/mg、73.62 U/g)。以上研究结果说明,颤藻具有一定的毒性,低浓度的颤藻可诱导酶活性增加,高浓度的颤藻能抑制酶的活性和对虾生长。为了保证凡纳滨对虾的健康养殖,养殖水体中的颤藻浓度应该控制在12.5 mg/L以下。     

亚硝酸氮和副溶血弧菌对凡纳滨对虾部分免疫指标的影响

在养殖水体中含不同浓度亚硝酸氮的情况下,用副溶血弧菌浸泡感染的方式对凡纳滨对虾(Litopenaeus vannamzei)进行毒性实验;并检测了凡纳滨对虾在亚硝酸氮和弧菌胁迫下的部分免疫指标的变化,研究亚硝酸氮和副溶血弧菌对凡纳滨对虾免疫力的影响.结果表明,实验Ⅰ:亚硝酸氮浓度为0.75、1.50、3.00 mg/L组分别胁迫健康凡纳滨对虾,对虾的血细胞数、血清酚氧化酶活力、酸性磷酸酶活力、超氧化物歧化酶活力和过氧化氢酶活力均显著低于对照组(0.005 mg/L)(P<0.05);10 d时,0.005、0.75、1.50、3.00 mg/L组的对虾死亡率分别达到0%、16.7%、20.0%、26.7%.实验Ⅱ:亚硝酸氮胁迫同时,用副溶血弧菌感染对虾,与实验Ⅰ相比,对虾的各项免疫指标受到的影响更大,其各项免疫指标与相应的实验Ⅰ相比,均有显著的差异(P<0.05);第10天时,0.005、0.75、1.50、3.00 mg/L对虾死亡率分别达到33.3%、40.0%、46.7%、50.0%.可见亚硝酸氮浓度越高,弧菌对对虾免疫力的破坏性越大,对虾的死亡率也越高.     

非离子氨胁迫对淡水和海水养殖凡纳滨对虾呼吸代谢酶活力影响的比较

为了探讨非离子氨胁迫对淡水和海水两种养殖条件的凡纳滨对虾呼吸代谢酶活力的影响,在实验室条件下研究了非离子氨胁迫(0.1 mg/L和0.5 mg/L)后,两种养殖条件凡纳滨对虾己糖激酶(HK)、丙酮酸激酶(PK)、乳酸脱氢酶(LDH)和琥珀酸脱氢酶(SDH)活力的变化规律,并将两种养殖条件对虾的相关指标进行了比较。结果显示:(1)非离子氨胁迫后,两种养殖条件凡纳滨对虾鳃HK活力变化显著,而肌肉HK活力变化则不显著。(2)非离子氨胁迫后,两种养殖条件凡纳滨对虾鳃PK活力先升高,后逐渐恢复到正常水平;淡水养殖对虾肌肉PK活力则显著升高,而海水养殖对虾肌肉PK活力变化则不显著。(3)0.1 mg/L非离子氨胁迫后,两种养殖条件对虾鳃和肌肉LDH活力变化均不显著,而0.5 mg/L非离子氨对两种养殖条件对虾鳃和肌肉的LDH活力均具有显著影响。(4)非离子氨胁迫后,两种养殖条件对虾鳃和肌肉SDH活力均显著降低。研究表明,非离子氨胁迫对淡水养殖凡纳滨对虾呼吸代谢酶活力具有显著影响;非离子氨胁迫后,凡纳滨对虾有氧代谢迅速减弱,而无氧代谢在胁迫初期略有升高,随后减弱,推测非离子氨胁迫可能使对虾机体主要供能物质发生改变。     

免疫增强剂对河鲫鱼白细胞吞噬和溶菌酶活性的影响

在饵料中添加不同含量的免疫增强剂(0.2%,0.5%,0.8%)连续投喂21d。在0,1,4,7,14,21d检测河鲫鱼(Carassius auratus)血液白细胞的吞噬活性、血清和体表粘液的溶菌酶活性。结果表明,该免疫增强剂可使河鲫鱼血液中白细胞吞噬活性、血清和体表粘液的溶菌酶活性都有明显提高。在停止投喂后7d,血清溶菌酶的活性逐渐降低与对照组无显著差异(P>0.05),但白细胞吞噬活性依然显著差异于对照组(P<0.05)。     

Seminal plasma proteins of Atlantic halibut (Hippoglossus hippoglossus L.)

Protein content and properties in the seminal plasma of Atlantic halibut (Hippoglossus hippoglossus) were assayed using spectrophotometric and electrophoretic methods. The protein concentration ranged from 6.4 ± 3.1 to 19.4 ± 3.4 mg ml⁻¹ and anti-proteolytic activity from 585.2 ± 104.6 to 2912.4 ± 367.4 U l^−l. A high correlation between anti-proteolytic activity and protein concentration (r = 0.95), and between sperm concentration and osmolality was found (r = 0.92). There was a significant decrease in anti-proteolytic activity from the first to the second sampling, but not in protein concentration. Anti-proteolytic activity and protein concentration were significantly affected by variations in individual males. Electrophoresis revealed four anti-proteolytic bands and individual differences in bands of proteolytic activity, which were subsequently characterized as metalloproteases and serine proteases.     

不同海洋弧菌中5类溶血素基因的分布及其与溶血活性和磷脂酶活性的相关性

用57株海洋弧菌,包括26株弧菌标准菌株、20株哈维氏弧菌、11株副溶血弧菌(从不同宿主和不同地理环境中分离得到),用PCR法合成5类相应的地高辛标记的溶血素基因探针,利用其进行Southern Blot,检测这5类溶血素基因在57株弧菌中的分布。结果显示,在57海洋株弧菌中,含有TDH、HlyA、TLH、δ-VPH和HLX溶血素基因的菌株分别为2株、2株、49株、3株和30株。另外,1株霍氏格里蒙菌(Grimontia hollisae)和1株副溶血弧菌(V.parahaemolyticus)中分别含有2个TDH溶血素基因。用鱼血平板和卵磷脂平板检测57株弧菌的溶血活性和磷脂酶活性,结果表明,弧菌溶血活性和磷脂酶活性与TLH溶血素基因具有显著相关性,与另外4类溶血素基因的关系不明显。     

北极微生物的分离及抗菌抗肿瘤活性筛选

采用多种培养基对北极地区(海洋沉积物、湖泊沉积物及土壤)微生物进行分离,并采用琼脂平板打孔法和MTT法进行了抗菌、抗肿瘤活性菌株的筛选.共分离极地微生物151株,筛选结果显示,12.4%的菌株有抗菌活性,6.2%菌株有抗肿瘤活性,其中3株同时具有抗菌、抗肿瘤活性,表明极地有良好的微生物药用资源.     

蜈蚣藻多糖降血糖及降血脂活性的研究

本文通过建立四氧嘧啶高血糖小鼠模型和高脂饲料高血脂小鼠模型,进行摄入不同剂量的蜈蚣藻多糖的动物实验,以测定的小鼠空腹血糖值、大鼠血脂水平[胆固醇（TC）、甘油三酯（TG）、高密度脂蛋白胆固醇（HDL—C）、低密度脂蛋白胆固醇（LDL—C）]以及肝脏的生化指标值,探讨蜈蚣藻多糖的降血糖、降血脂活性效果。结果表明蜈蚣藻多糖能显著降低高血糖小鼠的空腹血糖值及高血脂小鼠的TC,TG,LDL—C含量,提高HDL—C含量。蜈蚣藻多糖具有降血糖、降血脂等功效。     

甲基盐霉素对鲤白细胞吞噬活性和溶菌酶活力的影响

选用60 g左右健康正常的鲤540尾,随机平均分为6组,每组设3个重复。试验设6种处理,即基础饲料中分别添加甲基盐霉素0(对照组)、10、20、40、80、160 mg/kg。研究了甲基盐霉素对鲤白细胞吞噬活性和溶菌酶活性的影响。试验结果表明,饲料中添加10 mg/kg甲基盐霉素对鲤白细胞吞噬活性和溶菌酶活性影响不显著,而添加20~160 mg/kg甲基盐霉素对鲤白细胞吞噬活性和溶菌酶活性有一个从促进到抑制的转变,剂量越高,这个转变越快,并且随饲喂时间的延长,其抑制作用越来越强。     

Interaction of grammistins with lipids and their antibacterial activity

ABSTRACT: Grammistins are hemolytic and ichthyotoxic peptides in the skin secretion of soapfishes and are structurally characterized by their abundance in amphiphilic α-helicity. In the present study, their interaction with lipids and lipid vesicles as well as antibacterial activity were examined using four grammistins (Gs 1 and Gs 2 from Grammistes sexlineatus and Pp 1 and Pp 3 from Pogonoperca punctata ). The hemolytic activity of grammistins was inhibited by phospholipids but not by cholesterol. Moreover, grammistins released carboxyfluorescein entrapped within liposomes made of phosphatidylcholine. In contrast, grammistins were found to have antibacterial activity with a broad spectrum against nine species of bacteria, including both Gram-negative and Gram-positive groups. The potency of their antibacterial activity was not related to that of hemolytic activity, suggesting that grammistins bind to membrane phospholipids but lyse erythrocyte and bacterial membranes via different mechanisms. Conclusively, grammistins are new members of the family of cell non-selective membrane-lytic peptides with amphiphilic α-helices, being similar to pardaxins, which are secreted from the skin of soles, and to melittin, which is derived from bee venom.     

Biological activities of synthetic grammistins and analogous peptides

Grammistins, peptide toxins isolated from the skin secretions of two species of soapfishes Grammistes sexlineatus and Pogonoperca punctata, are new members of the family of linear membrane-lytic antibacterial peptides characterized by the abundance of amphiphilic α-helices. As the first step toward future use of grammistins to examine the mode of membrane lysis and design new antibiotics, C-terminally free and amidated forms were chemically synthesized for each of five grammistins (Gs 1, Gs 2, Pp 1, Pp 2b and Pp 3), and evaluated for hemolytic and antibacterial activities. No difference in biological activities was observed between C-terminally free and amidated forms of Gs 1, while the C-terminally amidated forms of the other grammistins exhibited much higher activities than the C-terminally free forms. Comparison with the biological activities of natural grammistins, together with our preliminary cloning experiments, showed that natural grammistins, except for Gs 1, are C-terminally amidated. As for Pp 1, six analogous peptides were further synthesized. Determination of their biological activities revealed that the amphiphilic α-helical structure and positive charge are important for the hemolytic and antibacterial activities, respectively, of Pp 1.     

Salinity regulates antioxidant enzyme and Na+K+‐ATPase activities of juvenile golden pompano Trachinotus ovatus (Linnaeus 1758)

Effects of salinity on the growth, survival, superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX) and sodium‐potassium adenosine triphosphatase (Na⁺K⁺‐ATPase) activities of juvenile golden pompano Trachinotus ovatus were studied under a laboratory condition. Experimental fish were reared at the salinities of 10‰, 18‰, 26‰ and 34‰ for 30 days. Growth and survival of juvenile golden pompano were significantly affected by the rearing salinity. Fish reared at 34‰ achieved the highest specific growth rate, while the highest survival was obtained when fish were cultured at the salinity of 26‰. The highest GPX activity was obtained when fish were cultured at 26‰, and the lowest GPX activity was observed when fish reared at 34‰ salinity. The SOD activities of fish reared at 18‰ and 34‰ were significantly higher than those reared at 10‰ and 26‰. The lowest of Na⁺K⁺‐ATPase activity was obtained in fish reared at 34‰, while the highest Na⁺K⁺‐ATPase activity was obtained when fish reared at 18‰. Results from present study indicate that juvenile golden pompano can be reared above 18‰ without sacrificing fish survival, and the best growth can be achieved when fish is reared at the salinity of 34‰. The salinity of 10‰ may be too low for juvenile golden pompano as the growth, survival and SOD activity were reduced.