蛭弧菌海水菌株Bdh5221裂解特性及生长影响因子研究

对利用双层琼脂平板法分离纯化获得的蛭弧菌海水菌株Bdh5221裂解特性及生长影响因子进行了初步研究,为其在生物防治上应用提供理论依据。研究结果表明,蛭弧菌海水菌株Bdh5221对嗜水气单胞菌、副溶血弧菌、鳗弧菌、溶藻弧菌、哈维氏弧菌、大肠杆菌、荧光假单胞菌、迟钝爱德华氏菌等革兰氏阴性菌有较好的裂解作用;对八叠球菌、金黄色葡萄球菌、枯草芽胞杆菌等革兰氏阳性菌的裂解能力较弱;对木糖葡萄球菌和啤酒酵母菌不裂解;可在超声波破碎后的G+菌和酵母菌上生长。蛭弧菌Bdh5221在宿主菌浓度106~109cfu/ml下均可正常生长,高浓度宿主菌更有利于蛭弧菌生长;Bdh5221最适生长的pH在7.0~7.5之间,盐度为20~30左右;Bdh5221增殖速度与起始接种浓度影响不大,起始接种浓度101~103pfu/ml下经过5~7d生长可增殖104左右。     

3株噬菌蛭弧菌裂解特性及在水体中的应用研究

对从海水中分离的3株噬菌蛭弧菌Blb-1、Blb-2、Blb-3在不同条件下的裂解特性进行了初步研究,并进行了Blb-2裂解水体哈维氏弧菌的应用试验。结果表明,3株菌在盐度为0~30时均能良好生长,具有广盐性;3株菌对灭活后的宿主菌具有更强的裂解作用,形成噬菌斑的时间普遍提前,且噬菌斑较大;3株蛭弧菌的裂菌谱试验以及不同蛭弧菌菌龄的裂解效果试验表明,3株菌对气单胞菌属和弧菌属细菌均有很好的裂解能力,而对水产益生菌芽孢杆菌、乳酸菌等裂解能力很弱;应用试验表明,噬菌蛭弧菌对水体中的哈维氏弧菌具有较强的的清除能力。     

夏季鲆人工育苗技术研究

1噬菌蛭弧菌微生物生态制剂的研究历程 1979年，我国首次开展对噬菌蛭弧菌（以下简称蛭弧菌）的研究。研究中发现蛭弧菌对许多动物（包括人体）和植物的致病菌，如霍乱弧菌、不凝集弧菌、伤寒沙门菌、副溶血弧菌、福氏志贺菌及大肠杆菌、嗜水气单胞菌、鳗弧菌、溶藻弧菌、荧光假单胞菌等均具有显著的裂解作用。中国水产科学研究院黄海水产研究所、中国海洋大学、上海水产大学、江苏省海洋渔业研究所等单位的研究证明，蛭弧菌对水生动物养殖中常见的副溶血弧菌、溶藻弧菌等革兰氏阴性菌的裂解率可达97．33％。     

蛭弧茵裂解作用的初步研究

蛭弧菌是一种寄生性细菌,广泛分布干土壤及备种水域之中,对诸多革兰氏阴性菌尤其是水产动物病原菌具有吞噬裂解活性.本文选取七株细菌为宿主菌,对蛭弧菌的裂解作用进行了初步研究.结果表明:除金黄色葡萄球菌外.蛭弧菌对其他六株菌均具有不同程度的裂解活性,其中以对红色假单胞菌的裂解能力更强.     

Ca2+、Mg2+、盐度对凡纳滨对虾存活、生长及风味的影响

采取L49(78)安排7水平Ca2+、Mg2+、盐度正交试验,开展60d凡纳滨对虾养殖试验,通过比较凡纳滨对虾成活率、日均增长值、日均增重量及鲜味氨基酸含量,分析养殖水体中Ca2+、Mg2+、盐度三因子对凡纳滨对虾存活、生长及虾体风味的影响;采取L8(27)安排2水平Ca2+、Mg2+、盐度正交试验,分析养殖水体中Ca2+、Mg2+、盐度三因子间交互作用对凡纳滨对虾存活、生长及虾体风味的影响。结果表明:水体中Ca2+、Mg2+、盐度对成活率和虾体鲜味氨基酸都有显著影响(P<0.05),Ca2+、Mg2+对凡纳滨对虾生长具有显著影响(P<0.05),其中Mg2+对成活率影响最大,Ca2+对生长影响最大,而鲜味氨基酸受盐度影响最大。Ca2+浓度为100mg/L和400mg/L,Mg2+为1200mg/L,盐度为10时,成活率最高;Ca2+≥200mg/L与Mg2+≥300mg/L时,生长速度无明显变化,Ca2+浓度为100mg/L,Mg2+浓度为150mg/L,盐度为10~20时,体长和体质量增加最快;Ca2+、Mg2+含量与盐度越高,鲜味氨基酸含量越高,Ca2+浓度为400mg/L,Mg2+浓度为750mg/L,盐度为35和20时,风味氨基酸含量最高;低盐、低钙、低镁水体显著降低了凡纳滨对虾的生长存活;Ca2+与Mg2+对成活率及体质量日均增长具有显著的交互作用影响,Ca2+与盐度对成活率具有显著的交互作用影响,Mg2+与盐度对各试验指标均没有显著影响。     

蛭弧菌裂解作用的初步研究

蛭弧菌是一种寄生性细菌．广泛分布干土壤及各种水域之中．对诸多革兰氏阴性菌尤其是水产动物病厚菌具有吞噬裂解活性。本文选取七株细菌为宿主菌．对蛭弧菌的裂解作用进行了初步研究。结果表明：除金黄色葡萄球菌外，蛭弧菌对其他六株菌均具有不同程度的裂解活性．其中以对红色假单胞菌的裂解能力更强。     

应用蛭弧菌清除海产品潜在致病弧菌的研究

以从海洋环境中分离到4株蛭弧菌作为生物净化因子,对16株常见海产品潜在食源性致病弧菌进行裂解(消除)试验。结果表明,Bh04-41a、Bh04-4、Bh04-A 和Bh04-1f等4株蛭弧菌分别可裂解4、7、11、12株致病弧菌,裂解率为24%、43.8%、68.8%、75%;4株一起,则可裂解15株弧菌,裂解率高达93.8%。研究结果展示了蛭弧菌在消除海产品中潜在食源性致病弧菌的可行性。     

鳗鲡肠道蛭弧菌的分离及其对水产动物细菌性疾病的预防作用研究

以多发性水产动物疾病的病原菌嗜水气单胞菌为宿主，从鳗鲡肠道中分离到一株噬菌力较强的蛭弧菌，命名为Bdm4。通过光学显微镜、相差显微镜和电子显微镜对蛭弧菌Bdm4进行革兰氏染色观察显示，蛭弧菌Bdm4为短杆状和长杆状的革兰氏阴性细菌，其大小约为0．3—0．4μm×1．2～1．5μm；菌有单根的极生长鞭毛。Bdm4对试验所选取的多种海淡水致病菌均有较强的裂解能力，而对所选用的益生菌则不能裂解；且其在鱼体表、鳃和肠道中能有效地消减致病菌，抑制致病菌的繁殖，同时有利于肠道乳酸杆菌数量的增长。     

蛭弧菌在水产养殖中应用研究进展

正蛭弧菌是一类专门寄生于其他细菌并能导致其裂解的革兰氏阴性菌,对多种致病菌具有良好的裂解效果~[1],属于蛭弧菌科,比一般细菌小,能通过细菌滤器,广泛存在于土壤、河流、下水道污泥等环境中。蛭弧菌不侵染真核细胞,对人和动物无致病性~[2-3],薛恒平等~[4-6]报道了蛭弧菌对仔鸡、小鼠、鲫鱼(Carassius auratus)的生长没有毒害作用。蛭弧菌作为"活性抗生素"在农牧业、医学以及环保方面     

养殖水体主要化学因子对蛭弧菌Bdh5221生长的影响

研究了养殖水体中NO-3N、NO-2-N、NH 4-N、PO-4-P和S2-等主要化学因子对蛭弧菌海水菌株Bdh5221生长的影响,为蛭弧菌在生物防治上的应用提供理论依据.试验结果发现,NO-3-N的浓度在O～10 mg/L之间、NH 4-N的浓度在0～5 mg/L之间,NO-3-N、NH 4-N的浓度越高越有利于蛭弧菌Bdh5221的生长;NO-2-N的浓度在0～0.5 mg/L之间,随着浓度的增高,对蛭弧菌Bdh5221的生长有一定的抑制作用;PO3-4-P的浓度在0～10 mg/L之间、S2-的浓度在0～5 mg/L之间,未见它们对蛭弧菌Bdh5221的生长有影响.     

罗非鱼皮营养成分分析及鱼皮明胶提取工艺的探讨

以罗非鱼皮为原料，对其营养成分进行分析，并通过正交试验方法探讨罗非鱼皮明胶的提取工艺，同时对明胶的理化及微生物指标进行分析。结果表明，罗非鱼皮中粗蛋白含量高达33．14％，其中胶原蛋白含量为27．8％，占其粗蛋白的83．9％；鱼皮提取明胶的适宜工艺条件为在2．5％（W／V）NaOH溶液中浸泡3．5h，然后在0．02％（V／V）HCl浸泡约3h，熬胶温度为55℃，提取得率为26．5％；所得鱼皮明胶的凝胶强度达468g，粗蛋白含量为82．46％，菌落总数为1．3×10^5CFU·g^-1，大肠菌群结果报告为阴性。     

Evaluation of the AQUARAPID-Va, AQUAEIA-Va and dot-blot assays for the detection of Vibrio anguillarum in fish tissues

The comparative accuracy of the serological assays AQUARAPID-Va, AQUAEIA-Va (BIONOR AS), and dot-blot for a rapid diagnosis of vibriosis in fish was evaluated. Twenty-one Vibrio anguillarum strains, representative of pathogenic and environmental serotypes, and 13 strains of other fish pathogenic bacteria were used to assess the sensitivity and specificity of the detection methods. The serological assays tested detected all the strains of V. anguillarum serotypes O1 and O2. The dot-blot assay was the most specific and sensitive method, detecting almost all isolates from serotypes O1, O2 and O3, with an average sensitivity of 1 x 10(6) bacteria g(-1) of fish tissue. The AQUARAPID-Va and the AQUAEIA-Va systems were able to detect 5 x 10(6) and 5 x 10(7) bacteria g(-1) of fish tissue, respectively. The simplicity, effectiveness and speed of the AQUARAPID-Va system confirmed this method as the most suitable serological test for the detection of V. anguillarum in field analysis and small-scale laboratory studies.     

Non-specific immune response of turbot,Scophthalmus maximus (L.), experimentally infected with a pathogenic Vibrio pelagius

The effect of a pathogenic Vibrio pelagius, isolated during a mass mortality of turbot larvae, on the non-specific immune response of turbot, Scophthalmus maximus (L.), macrophages was studied both in vitro and in vivo. The in vitro treatment of head kidney (HK) macrophages with viable V. pelagius caused a significant inhibition of the chemiluminescence (CL) response in comparison with untreated macrophages, while incubation with heat-killed bacteria did not affect this response. In vivo, the intraperitoneal injection of V. pelagius resulted in a significant inhibition of the CL response in infected fish at days 1 and 4 post-infection compared with the control fish response. The HK macrophage nitric oxide (NO) production was enhanced by in vitro incubation with intermediate doses of viable V. pelagius (5 x 10(3) and 5 x 10(4) bacteria mL(-1)) and higher doses of the heat-killed bacteria (5 x 10(4)-5 x 10(6) bacteria mL(-1)). In both cases, the NO inhibitor N-omega -nitro-L-arginine was capable of down-regulating the specific NO induction caused by incubation with the bacterial treatments. In contrast, incubation with ECPs at higher doses caused a reduction in NO production. In vivo, a significant enhancement in NO production was also observed in macrophage supernatants at day 10 post-infection. Lysozyme concentration in the serum was also significantly increased in the experimentally infected fish at days 4 and 10 post-injection. In addition, viable V. pelagius and its ECPs significantly reduced HK macrophage viability in vitro, whereas no significant differences in viability were observed during the incubation with heat-killed bacteria. As NO production was enhanced in the experimentally infected fish, the inhibitory effect of the NO donor, S-nitroso-acetyl-penicillamine (SNAP), was tested in vitro in a cell-free assay. The results showed that growth of V. pelagius was significantly inhibited using SNAP at a high concentration (1 mM).     

罗非鱼循环水养殖系统中一株肺炎克雷伯氏菌的分离鉴定及其脱氮性能研究

实验从山东德州市一罗非鱼(Oreochromis niloticus)养殖场循环水养殖系统的脱氮池中分离到一株具有高效脱氮特性的菌株(编号DZYC02),分别以葡萄糖、蔗糖、可溶性淀粉、丁二酸钠、乙酸钠、柠檬酸钠为碳源,研究了碳源种类对菌株DZYC02脱氨氮效果的影响;同时以蔗糖为碳源、NH4Cl为氮源,研究了不同碳氮比、初始pH及盐度对该菌株脱氮效果的影响。结果显示:菌株DZYC02在以柠檬酸钠为碳源、C∶N≥15、pH 5~7、盐度0~15的条件下具有良好的脱氮效果,24 h内对浓度为20 mg/L的NH+4的去除率达100%,48 h内对浓度为20 mg/L的NO-2去除率高达100%;将该菌采用浸泡方式感染斑马鱼(Danio rerio)进行生物安全试验,结果显示菌株DZYC02对斑马鱼表现出较好的安全特性。分别用Biolog细菌鉴定方法和16S rDNA序列分析比对法对该菌进行鉴定,结果显示菌株DZYC02为一株肺炎克雷伯氏菌(Klebsiella pneumoniae)。     

长鳍真鲨源哈氏弧菌的主要生物学性状研究

从两尾病死的观赏用长鳍真鲨Oceanic whitetip shark，Carcharhinus longimanus L．中分离到的细菌，进行了形态特征、理化特性和对抗菌类药物的敏感性等生物学性状检验。同时，测定了16S rRNA基因序列，分析了相关细菌相应序列的同源性，构建了系统发育树。结果表明，8株供试纯培养菌（编号：HQ050226—1至HQ050226—4、HQ050227-1至HQ050227—4）为弧菌属（Vibrio Pacini 1854）的哈氏弧菌（Vibrio harveyi Johnson and Shunk 1936）。所扩增的16S rRNA基因序列长度（不包括引物结合区），HQ050226—1株为1463bp（GenBank登录号：EF635305）、HQ050227—1株为1464bp（GenBank登录号：EF635306），与GenBank数据库中哈氏弧菌的同源性在98％及99％。药敏试验结果显示，对供试37种抗菌药物中的青霉素G等4种耐药，对头孢唑啉等33种敏感。     

Vaccine development for winter ulcer disease, Vibrio viscosus, in Atlantic salmon, Salmo salar L.

Coldwater Vibrio species isolated from Atlantic salmon, Salmo salar L., during winter ulcer disease outbreaks at saltwater sites in Norway and Iceland were characterized phenotypically, tested for virulence, and used to evaluate the efficacy of multivalent, oil-adjuvanted vaccines. The intraperitoneal (i.p.) injection of rainbow trout, Oncorhynchus mykiss (Walbaum), in fresh water with one bacteria species isolated during winter ulcer outbreaks, V. ‘viscosus’, produced rapid mortality and disease signs which resembled those observed during natural outbreaks [10⁵ colony-forming units (cfu) fish^??1]. Another species, V. ‘wodanis’, was not virulent to rainbow trout (10³–10⁶ cfu fish^??1). Although vaccination of rainbow trout with a mineral-oil-adjuvanted, injectable vaccine containing V. anguillarum (serotypes 01 and 02), V. salmonicida and Aeromonas salmonicida did not provide protection against injection challenge with V. viscosus, vaccines which included V. viscosus produced significant protection in Atlantic salmon and rainbow trout. Atlantic salmon vaccinated with an oil-adjuvanted vaccine containing V. viscosus, V. wodanis and atypical A. salmonicida produced a relative percentage survival (RPS) of 97% when challenged i.p. with V. viscosus, demonstrating cross-protection between strains from Iceland and Norway. Short-term efficacy was demonstrated in rainbow trout by injection challenge at 21 and 43 days post-vaccination with an oil-adjuvanted vaccine containing V. viscosus, V. anguillarum (01/02), V. salmonicida and A. salmonicida, which produced an RPS of 96–99%. Rainbow trout challenged with V. viscosus at 52 and 362 days post-vaccination produced an RPS of 93% and 79%, indicating that vaccination provided long-term protection. In a similar manner, rainbow trout injected i.p. with 0.2 mL of a vaccine containing the five bacteria species and infectious pancreatic necrosis virus produced a 90% RPS when challenged with V. viscosus 66 days later. The high RPS under a severe challenge burden, along with disease signs in experimental freshwater challenges which resembled the saltwater disease condition, indicated that V. viscosus is a contributing factor to winter ulcer and that vaccination will protect against the disease.     

对虾养殖池中一株弧菌拮抗菌的分离鉴定

从对虾养殖池中分离出1株编号为2013082515(简称菌株15)的菌株,以鳗弧菌(Vibrio anguillarum)、哈维氏弧菌(Vibrio harveyi)和副溶血弧菌(Vibrio parahaemolyticus)为指示菌,分析了菌株15对指示菌的抑菌效果及最低抑菌浓度,同时,分析了菌株15对其他7株弧菌的抑菌效果.结果显示,菌株15对3株指示菌均具有抑菌效果,对鳗弧菌、哈维氏弧菌及副溶血弧菌的最低抑菌浓度分别为2.50×104、2.50×105、2.50×105 CFU/ml;对其他弧菌也具有一定的抑菌效果.采用注射及浸泡感染方法分析了菌株15对对虾的生物安全性,结果显示,在高浓度时,菌株15对对虾具有潜在毒性.分别用细菌全细胞脂肪酸气相色谱法和16S rDNA序列分析比对法对该菌进行分类鉴定,表明菌株15为一株假交替单胞菌(Pseudoalteromonas sp.).     

海带_江蓠浸渍光合细菌菌液后喂养九孔鲍的效果

使用海带、江蓠浸渍光合细菌的不同浓度和时间对大小两种不同规格九孔鲍的生长效果进行试验.结果表明,光合细菌促进养成鲍的生长效果,因海带、江蓠浸渍光合细菌的浓度和时间而异,浸渍浓度越大,浸渍时间越长,则其促进生长作用越明显;海带、江蓠浸渍光合细菌的浓度和时间分别为150×10-6(V/V)和3h～4h或200×l0-6(V/V)和2h～3h的试验组鲍的平均壳长增长率都比对照组提高了5%以上,最高提高11.5%左右(P＜0.05).     

中国对虾中抗菌短肽的分离纯化与功能分析

以水产病害的病原菌为目标菌，从中国对虾（Penaeuschinesis）体液中分离筛选抗菌肽，通过Sephadex G-50、RP-HPLC等分离纯化技术，得到一种抑制水产病原菌的抗菌肽PC-V-2。经初步鉴定，该抗菌肽具有广谱抗性，不仅抗鲁克氏耶尔森氏菌（Yersinia ruckeri），而且对金黄色葡萄球菌（Staphyloccus auleus）、枯草杆菌（Bacillus subttilis）等革兰氏阳性菌，大肠杆菌（Escherichia coli）等革兰氏阴性菌和白色念珠菌（Candida albicans）等真菌也有较强的抗性。另外，该抗菌肽还有一定的抑制凝血的活性，但没有检测到溶血和丝氨酸蛋白酶抑制活性。经MALDI-TOF质谱分析，其分子量为1848D，但其序列及其他生物特性还有待进一步研究。     

网箱养殖青石斑鱼的溃疡病病原

2002年夏季厦门同安湾一带气候炎热,降雨量少,海水盐度偏高,同安湾刘五店的网箱养殖石斑鱼大面积暴发溃疡病.本研究调查了石斑鱼溃疡病暴发的特征和症状,主要表现为肌肉溃疡坏死、眼球脱落、鱼骨暴露等.从病鱼体表及内脏分离出优势菌群命名为TS-628,经回归感染证实TS-628就是引发本次石斑鱼溃疡病的病原菌.对病原菌进行鉴定发现,该菌革兰氏染色呈阴性,电镜下观察菌体呈短杆状,极端单鞭毛,综合研究该菌在形态、生理生化、16S rDNA同源性及药物敏感性等方面的特性,基本确认分离到的病原菌为哈维氏弧菌(Vibrio harveyi),该菌对氯霉素、壮观霉素等多种抗生素敏感,对万古霉素、青霉素G等抗生素不敏感.哈维氏弧菌是水产养殖病害常见病原菌,但作为养殖石斑鱼的病原菌在国内属首次报道.