养殖大菱鲆病原菌迟缓爱德华氏菌的分离鉴定及其疫苗研制

2006年9月山东胶南某养殖场大菱鲆(Scophthalmus maximus)发生病害,从患病大菱鲆脾脏分离出优势菌株WY28,人工感染试验证实WY28菌株对大菱鲆和斑马鱼(Danio rerio)有较强的致病性,其对大菱鲆和斑马鱼的半数致死剂量(LD50)分别为39cfu/g(Bw)(3.3×102cfu/ind)和2.1×104cfu/g(Bw)(6.4×103cfu/ind).综合该菌在形态与生理生化特征及16S rDNA同源性等方面的实验结果,确定WY28菌株为迟缓爱德华氏菌(Edwardsiella tarda).该菌对先锋霉素V、庆大霉素、氟哌酸、痢特灵等抗生素敏感.WY28菌株经福尔马林灭活制成灭活疫苗,对大菱鲆进行腹腔注射免疫,免疫后第4周测得免疫鱼血清中抗迟缓爱德华氏菌的抗体效价平均为1:1 280.免疫后第6周.免疫鱼血清中抗迟缓爱德华氏菌的抗体效价达到更高水平(平均值为1:3 289.6).攻毒试验表明,受免鱼的相对存活率明显高于对照组.由此可见,利用从病鱼体内分离的迟缓爱德华氏菌菌株制备的灭活疫苗能使大菱鲆较有效抵御迟缓爱德华氏菌强毒株的攻击.     

鲮鱼嗜水气单胞菌病病原的鉴定及药敏试验

从患病鲮鱼的体表、肝、肾分离到3株致病菌(QY1567,QY1569,QY1578),通过培养特性、生化特征与分子鉴定,确定为嗜水气单胞菌(Aeromonas hydrophila)。人工感染试验显示,此3株菌对鲮鱼具有较强的致病力,LD_(50)依次为3.04×10~5、3.8×10~5和1.53×10~5cfu/g。药敏试验表明,3株致病菌对阿米卡星、链霉素、环丙沙星、头孢他啶、米诺环素、妥布霉素、苯唑西林、氨曲南等抗生素高度敏感,对卡那霉素、麦迪霉素、呋喃妥因中度敏感,对左氟沙星、头孢噻吩、庆大霉素、头孢唑啉、头孢噻肟、青霉素G、头孢西丁和头孢哌酮不敏感。     

海南罗非鱼致病性维氏气单胞菌分离鉴定及药敏特性研究

2014年3—5月海南省罗非鱼主养区多个养殖场发生爆发性疾病,从患病尼罗罗非鱼不同组织中分离获得12株优势菌。经人工回归感染试验,从肝脏和脑中分离获得的HN-G-03和HN-B-02菌株具较强的致病力,HN-G-03和HN-B-02菌株对罗非鱼的的半致死密度分别为7.12×105cfu/尾和1.32×105cfu/尾。经16S rRNA和gyr B基因序列分析,同时结合细菌的形态学特征、生理生化特征,菌株HN-G-03和HN-B-02分别被鉴定为维氏气单胞菌的维氏生物型和温和生物型。体外药物药敏试验结果显示,两株菌均对左氧氟沙星、强力霉素、氟苯尼考、恩诺沙星、庆大霉素等5种药物高度敏感,对呋喃妥因等3种药物中度敏感,对青霉素等12种药物较强耐药。     

鲮鱼致病性迟钝爱德华氏菌的鉴定及药敏分析

自广东省清远市某养殖场患病鲮鱼的肝、肾、脾组织分离到两株优势菌QY15814、QY15815,通过细菌生化鉴定管与16SrDNA序列分析对这两株菌进行鉴定,采用纸片扩散法进行药敏试验,并测定了这两株菌对鲮鱼的半致死剂量。试验结果显示,这两株菌为革兰氏阴性短杆菌,兼性厌氧,有动力,能利用蔗糖、甘露糖、麦芽糖,硝酸盐还原、吲哚、MR试验均为阳性;根据其生理生化特性,鉴定为迟钝爱德华氏菌,通过PCR扩增其16SrDNA基因,经测序及NCBI比对,结果与迟钝爱德华氏菌的同源性高达99%。这两株致病菌对头孢噻肟、头孢吡肟、头孢曲松、头孢西丁、头孢哌酮、头孢呋辛等抗生素高度敏感,对链霉素、克拉霉素、红霉素中度敏感,对阿米卡星、复方新诺明、大观霉素、克林霉素不敏感。这两株菌对鲮鱼具有较强的致病力,半致死剂量分别为1.93×105cfu/g和9.65×104cfu/g。     

天峡红鱼回出血病的病原及其防治研究

从患出血病的天峡红蛔进行了病原菌的分离得出 2株细菌 ,经过人工感染实验确立H -I菌株为致病菌株 ,并对该菌形态培养特性 ,生理生化鉴定 ,确定该菌为豚鼠产气单胞菌。药敏　　试验该菌对呋喃妥因 ,氯霉素、丁胺卡那最为敏感 ,经过生产防治试验取得了较好的效果     

大菱鲆源杀鱼爱德华氏菌(Edwardsiella piscicida)的分离鉴定

从患腹水病大菱鲆Scophthalmus maximus的肝脏中分离到一株优势细菌G1,经人工感染实验表明G1为引发大菱鲆腹水病的致病菌,且半致死浓度为LD50=1.21×105 CFU·g-1。采用常规的生理生化鉴定方法及分子生物学方法对G1进行分析,结果表明,G1的16SrRNA基因序列与杀鱼爱德华氏菌Edwardsiella piscicida的同源性达100%,系统发育分析表明菌株G1与杀鱼爱德华氏菌分支聚为一支,结合生理生化鉴定结果确定G1为杀鱼爱德华氏菌。药敏试验表明菌株G1对头孢曲松、环丙氟哌酸、左氧氟沙星等8种抗菌药物高度敏感。     

天峡红Hui出血病的病原及其防治研究

从患出血病的天峡红蛔进行了病原菌的分离得出2株细菌，经过人工感染实验确立H－1菌株为致病菌株，并对该菌形态培养特性，生理生化鉴定，确定该菌为豚鼠产气单胞菌，药敏试验该菌对呋喃妥因，氯霉素，丁胺卡那最为敏感，经过生产防治试验取得了较好的效果。     

中华鳖爱德华氏菌病的病原与防治研究

从患白底板症和肝肿大的病鳖腹水、肝和肠道分离到５株细菌，经人工感染试验证实其中３株为致病菌，生理生化试验鉴定为爱德华氏菌。药敏试验表明病原菌对庆大霉素、卡那霉素、氯霉素、呋喃妥因等药物高度敏感；中草药和开发的新药“菌毒宁”能较好地防治该病。     

鲟源致病性鲁氏耶尔森菌的分离、鉴定及药敏研究

2017年,新疆阜康市某鲟鱼养殖场的西伯利亚鲟出现大批死亡,主要症状为生殖孔红肿外突,腹部有点状出血。自患病西伯利亚鲟肝脏、脾脏分离到一株优势菌株XB2,通过革兰氏染色、生理生化试验、药敏试验,并对其16S rRNA基因序列进行测定,用Mega 6.0对序列进行分析。生理生化试验结果显示,菌株XB2为革兰氏阴性短杆菌,单独或成对出现;产酸不产气,V-P反应呈阳性,能利用蔗糖、果糖、葡萄糖、明胶、枸橼酸;在含1%(m/V)氯化钠中能生长,高于3%(m/V)氯化钠中不生长;在4℃不生长,20～30℃均能生长,但在37℃不生长;不利用乳糖、水杨苷、纤维二糖、硫化氢、酒石酸、阿拉伯糖。进化树显示,菌株XB2与鲁氏耶尔森菌KJ812974同源性达99%。通过回感试验确定其半致死密度为2.37×104 cfu/mL。结合生理生化鉴定结果和16S rRNA基因序列分析结果,确定菌株XB2为鲁氏耶尔森菌。药敏结果显示,鲁氏耶尔森菌XB2对氟苯尼考、左氧氟沙星、哌拉西林、阿洛西林等9种药物敏感,对麦迪霉素、大观霉素、链霉素、氨苄西林等15种药物耐药。     

泥鳅白头病病原菌的分离鉴定、药敏试验及拮抗菌的筛选

为确定连云港市墩尚镇泥鳅(Misgurnus anguillicaudatus)白头病的病原菌,本研究采用常规微生物分离鉴定技术和分子生物学相关方法,对患病泥鳅进行病原菌的分离、鉴定并分析其药物敏感性,采用滤纸片抑菌圈法筛选了该病原菌的拮抗菌。从患病泥鳅病灶部位分离到1株病原菌并命名为W1,经人工感染发现体腔注射浓度为1.6×10~7 CFU/mL 0.1 mL菌液时泥鳅死亡率为80%,在1.6×10~7 CFU/mL浓度下创伤和浸泡感染死亡率为100%。通过形态特征观察、生理生化实验和16S rRNA基因序列分析,将该病原菌初步鉴定为维氏气单胞菌(Aeromonas veronii)。药敏试验结果显示,该菌株对头孢克肟高度敏感,对呋喃唑酮、链霉素、庆大霉素和恩诺沙星中度敏感,对红霉素、环丙沙星、四环素等不敏感。筛选该病原菌的拮抗菌发现,解淀粉芽孢杆菌(Bacillus amyloliquefaciens)X8对维氏气单胞菌W1具有较强的抑制作用。     

Association between the interannual variation in the oceanic environment and catch rates of bigeye tuna (Thunnus obesus) in the Atlantic Ocean

The environmental processes associated with variability in the catch rates of bigeye tuna in the Atlantic Ocean are largely unexplored. This study used generalized additive models (GAMs) fitted to Taiwanese longline fishery data from 1990 to 2009 and investigated the association between environmental variables and catch rates to identify the processes influencing bigeye tuna distribution in the Atlantic Ocean. The present findings reveal that the year (temporal factor), latitude and longitude (spatial factors), and major regular longline target species of albacore catches are significant for the standardization of bigeye tuna catch rates in the Atlantic Ocean. The standardized catch rates and distribution of bigeye tuna were found to be related to environmental and climatic variation. The model selection processes showed that the selected GAMs explained 70% of the cumulative deviance in the entire Atlantic Ocean. Regarding environmental factors, the depth of the 20 degree isotherm (D20) substantially contributed to the explained deviance; other important factors were sea surface temperature (SST) and sea surface height deviation (SSHD). The potential fishing grounds were observed with SSTs of 22–28°C, a D20 shallower than 150 m and negative SSHDs in the Atlantic Ocean. The higher predicted catch rates were increased in the positive northern tropical Atlantic and negative North Atlantic Oscillation events with a higher SST and shallow D20, suggesting that climatic oscillations affect the population abundance and distribution of bigeye tuna.     

Growth performance,digestive enzyme activity and immune response of Japanese sea bass,Lateolabrax japonicus fed with fructooligosaccharide

In this experiment, a feeding trial was performed to determine the effects of fructooligosaccharide (FOS) on growth performance, digestive enzyme activity and immune response of Japanese sea bass, Lateolabrax japonicus juveniles (initial weight 38.3 ± 0.5 g), and the fish were examined following feeding with six levels of FOS (0, 0.5, 1, 2, 4 and 6 g/kg) for 28 days. Significant enhancement of weight gain (WG) and specific growth rate (SGR) was found in fish fed 1 g/kg FOS incorporated diets (p < .05), while the feed conversion ratio (FCR) in the 1, 2 g/kg FOS groups reduced significantly compared with the control (p < .05). Besides, the crude lipid in the 4, 6 g/kg FOS groups increased significantly compared with the control (p < .05). On the other hand, the erepsin and lipase activities significantly elevated in intestine of fish fed 2 g/kg FOS (p < .05) and the lysozyme activity in serum of fish fed 2 g/kg FOS were significantly higher than that in the control (p < .05). Moreover, the alkaline phosphatase activities in serum of fish fed 0.5, 1, 2 g/kg FOS were significantly higher than in control (p < .05). Regression analysis showed that the relationships between dietary FOS levels and either SGR, FCR, erepsin or lysozyme activities were best expressed by regression equations, and the optimal inclusion levels are 1.37, 1.80, 3.06, 3.11, 1.93 and 1.80 g/kg for SGR, FCR, erepsin, lipase, lysozyme and total superoxide dismutase activities, respectively. Overall, this study revealed that FOS incorporated diets could beneficial for L. japonicus culture in terms of increasing the growth, digestion and immune activities. Under the present experimental condition, the optimal supplementary level of FOS in the diet of L. japonicus is 1–3 g/kg.     

Involvement of gonadal steroids in final oocyte maturation of white perch (Morone americana) and white bass (M. chrysops): in vivo and in vitro studies

Plasma estradiol-17 (E₂), testosterone (T), 17,20-dihydroxy-4-pregnen-3-one (DHP) and 17,20,21-tri-hydroxy-4-pregnen-3-one (20-S) levels were measured by radioimmunoassay (RIA) in white perch (Morone americana) and white bass (M. chrysops) that were induced to undergo final oocyte maturation (FOM) with human chorionic gonadotropin (hCG). Plasma DHP levels increased in females of both species in association with oocyte germinal vesicle migration (GVM) and germinal vesicle breakdown (GVBD) and decreased thereafter. Plasma 20-S levels also increased with oocyte GVM in white bass, but were several-fold lower than DHP levels. Circulating E₂ and T levels were greatest during GVM and GVBD in both species and decreased to low levels during oocyte hydration and ovulation. Follicles from white perch and white bass which received a priming injection of hCG in vivo, produced both DHP and 20-S in vitro after exposure to hCG and their oocytes underwent GVBD. Ovarian incubates from unprimed fish of either species produced only E₂ and T and their oocytes did not complete GVBD. Oocytes from unprimed bass, but not perch, matured when follicles were exposed to hCG in vitro. Both trilostane and cycloheximide blocked in vitro production of DHP and 20-S and oocyte GVBD by white perch follices. DHP and 20-S were equipotent inducers of FOM in the GVBD bioassay. None of several other structurally-related steroids tested were effective within a physiological range of concentrations. These results indicate a role for DHP and 20-S in the control of FOM in white perch and white bass.     

Cardiac,ventilatory and metabolic responses of two ecologically dissimilar species of fish to waterborne cyanide

Changes in heart rate, ventilatory activity and oxygen consumption were determined in trout (Salmo gairdneri) and brown bullhead catfish (Ictalurus nebulosus) during exposure to a steadily increasing concentration of waterborne cyanide selected to produce death in 8–9 hours for each species. The lethal cyanide concentration for the bullheads was an order of magnitude higher than for trout. Trout developed an immediate and gradually increasing bradycardia throughout the exposure period. Cyanide produced tachycardia in the bullhead followed by a gradual onset of bradycardia as the concentration of cyanide was raised. Pericardial injection of atropine (a muscarinic cholinergic antagonist) indicated that bradycardia in the trout was due initially to increased vagal tone but later due to the direct effect of cyanide on the heart. Hyperventilation in the trout persisted throughout the exposure period, although the rate and amplitude fluctuated and was variable between individual fish. During the last hour of exposure (highest cyanide concentration), ventilation was characterized by rapid, shallow breaths followed by a sudden respiratory arrest. The bullheads exhibited hyperventilation during the first 3 hours of exposure followed by a gradual, linear drop in ventilation rate and amplitude until death occurred. Cardiac and ventilatory responses in both species were attributed to stimulation of central and peripheral chemoreceptors by cyanide. Evidence is presented which suggests the initial response in the bullheads was due, at least in part, to gustatory stimulation by the cyanide. Oxygen consumption of the trout remained above pre-exposure levels for the majority of the test period. Oxygen consumption in the bullhead paralleled the changes in heart and ventilatory rates. Whole-body lactate levels of fingerlings of both species during cyanide exposure were measured to estimate the extent of anaerobiosis. Whole-body lactate levels were much greater in the bullheads than the trout, indicating a higher capacity for anaerobiosis, possibly due to a greater fuel supply. Overall, the trout responded to cyanide in a manner similar to that produced by environmental hypoxia whereas the bullheads experienced a gustatory stimulus which masked the hypoxia-like response.     

An overview of stress physiology of fish transport: changes in water quality as a function of transport duration

This study brings an integrated analysis about the relationship between water deterioration and its physiological consequences in live fish transport. The analysis was focused on the transport water and its deterioration, and physiological challenges imposed on the fish. Usual commercial handling procedures employed to mitigate fish stress during transport were discussed. Future topics of research for the establishment of safer fish transport protocols were proposed. Transport was classified into short (≤8 h) or long transport (>8 h). The main issue in short transports should be the prevention of water pH reduction, while in long transports it is the increase in ammonia. Plasma cortisol is the most employed marker for stress and is acutely elevated upon short episodes of transport, but remains elevated even in long‐transport events. Plasma glucose is perhaps a better marker for handling stress. Plasma lactate, pH, osmolality CO₂ and ions should be more often evaluated. Plasma Na⁺ and Cl^‐ are very useful markers of acidosis, due to their respective exchange for H⁺ and , for acid–base regulation. The establishment of species‐specific transport protocols should be preceded by such combined analyses of water and physiological parameters.     

Are hatchery‐reared abalone naïve of predators? Comparing the behaviours of wild and hatchery‐reared northern abalone,Haliotis kamtschatkana (Jonas, 1845)

Abalone populations have declined worldwide, generating interest in enhancement using hatchery‐reared individuals. In many cases, such restoration efforts have met with limited success due to high predator‐induced mortality rates. Furthermore, the mortality rates of outplanted hatchery abalone are often considerably higher than for wild individuals. This study uses northern abalone (Haliotis kamtschatkana) as a case study to determine whether hatchery‐reared abalone behave differently than their wild counterparts. In the field, outplanted hatchery‐reared abalone were significantly less responsive than wild abalone, in terms of number of abalone responding and intensity of response, to nearby movement and to physical contact with an inert probe. Also, when encountering a cue to which all abalone responded (a seastar predator), hatchery‐reared individuals remained subdued. Anti‐predator behavioural deficits in hatchery‐reared abalone were more pronounced in 4‐year‐old individuals than in 1‐year‐old individuals, suggesting an influence of either age or amount of time spent in the hatchery environment. These behavioural differences are expected to increase the vulnerability of hatchery‐reared abalone to predators, and are likely a major cause of their elevated predator‐induced mortality when outplanted.     

Effects of cadmium on the kinetics of calcium uptake in developing tilapia larvae, Oreochromis mossambicus

The toxic effects of Cd²⁺ on Ca²⁺ influx kinetics in developing tilapia (Oreochromis mossambicus) larvae were evaluated. Addition of 20 µg l^-1 of Cd²⁺ to the environment of 0 and 3 day-old larvae competitively inhibited the Ca²⁺ uptake within 4h resulting in a great increase in K_m values for Ca²⁺ influx (19.3 and 17.4 fold, respectively) as compared with their respective controls. Consequently, the actual Ca²⁺ influx of larvae in solutions of 0.2 mM Ca²⁺ are suppressed by 32–45%. Also, 3 day-old larvae were more sensitive to internally accumulated Cd²⁺ than 0 day-old larvae. Although the Ca²⁺ influx in 0 and 3 day-old larvae may be restored to the levels of their respective controls with 24h of being transferred to a 20 µg l^-1 Cd²⁺ solution, total body Ca²⁺ content was significantly reduced in 3 day-old larvae. Increased Ca²⁺ uptake efficiency ensures sufficient Ca²⁺ for normal growth. However, rapid increase in Ca²⁺ influx after hatching also leads to higher Cd²⁺ uptake. Exposure to Cd²⁺ will lead to a drop in body Ca²⁺ content resulting in retardation of larval growth. Therefore, we conclude that if Ca²⁺ uptake is interfered with at this critical stage of development, larvae will not be able to maintain normal levels of body Ca²⁺ and will show signs of Cd²⁺ poisoning.     

Migratory dynamics of stream-spawning longnose gar (Lepisosteus osseus)

Abstract– Literature evidence suggests that lake-dwelling longnose gar (Lepisosteus osseus) enter tributary streams to spawn, Until the present study, the dynamics of this breeding migration had never been investigated quantitatively. During the summers of 1991 and 1992, longnose gar were captured as they entered Weaubleau Creek, Missouri, a tributary of Harry S. Truman Reservoir. The in-stream spawning migration began in early April and ended in late May, and was positively correlated with stream flow and water level, and negatively correlated with water temperature. In-stream residence times ranged from 15 to 94 days, with males exhibiting longer residence times than females. Once in-stream, longnose gar travelled as far as 10 km upstream and occupied certain pools at greater relative frequencies. Although the reason for this preferential utilization is not completely understood, it may relate to pool depth and riffle proximity. Longnose gar disperse from the spawning stream great distances, with gar captured in Weaubleau Creek being recaptured up to 48 km away. This information should provide fisheries biologists the means to consider the reproductive ecology of this species in their conservation and management decisions.     

Nutritional regulation of hepatocyte fatty acid desaturation and polyunsaturated fatty acid composition in zebrafish (Danio rerio) and tilapia (Oreochromis niloticus)

The desaturation and elongation of [1-¹⁴C]18:3n-3 was investigated in hepatocytes of the tropical warm freshwater species, zebrafish (Danio rerio) and Nile tilapia (Oreochromis niloticus). The hepatocyte fatty acid desaturation/elongation pathway was assayed before and after the fish were fed two experimental diets, a control diet containing fish oil (FO) and a diet containing vegetable oil (VO; a blend of olive, linseed and high oleic acid sunflower oils) for 10 weeks. The VO diet was formulated to provide 1% each of 18:2n-6 and 18:3n-3, and so satisfy the possible EFA requirements of zebrafish and tilapia. At the end of the dietary trial, the lipid and fatty acid composition was determined in whole zebrafish, and liver, white muscle and brain of tilapia. Both zebrafish and tilapia expressed a hepatocyte fatty acid desaturation/elongation pattern consistent with them being freshwater and planktonivorous fish. The data also showed that hepatic fatty acid desaturation/elongation was nutritionally regulated with the activities being higher in fish fed the VO diet compared to fish fed the FO diet. In zebrafish, the main effect of the VO diet was increased fatty acid Δ6 desaturase activity resulting in the production of significantly more 18:4n-3 compared to fish fed the FO diet. In tilapia, all activities in the pathway were greater in fish fed the VO diet resulting in increased amounts of all fatty acids in the pathway, but primarily eicosapentaenoic acid (EPA; 20:5n-3) and docosahexaenoic acid (DHA; 22:6n-3). However, the fatty acid compositional data indicated that despite increased activity, desaturation of 18:3n-3 was insufficient to maintain tissue proportions of EPA and DHA in fish fed the VO diet at the same level as in fish fed the FO diet. Practically, these results indicate that manipulation of tilapia diets in commercial culture in response to the declining global fish oil market would have important consequences for fish fatty acid composition and the health of consumers. Scientifically, zebrafish and tilapia, both the subject of active genome mapping projects, could be useful models for studies of lipid and fatty acid metabolism at a molecular biological and genetic level. This revised version was published online in August 2006 with corrections to the Cover Date.     

Protein and amino acid composition from the mantle of juvenile O ctopus vulgaris exposed to prolonged starvation

Protein and amino acid composition of the mantle of juvenile O ctopus vulgaris (Cuvier, 1797) during fasting for 27 days were determined. Average protein content of octopus mantle was of 711.19 ± 46.80 g kg^?1 DW, and it decreased with increasing fasting days. The non‐essential amino acids content was higher (486.18 ± 11.08 g kg^?1 protein) than essential amino acids (425.82 ± 9.15 g kg^?1 protein) at the start of the experiment (unstarved animals). The results suggest that the amino acid profile of the mantle where the most abundant amino acids are Arg, His, Lys, Gly, Leu and Pro could indicate a prolonged fasting condition (>20 days) or poor nutrition of O . vulgaris. This study supports the idea of using mantle for metabolic needs of starved O . vulgaris suggesting that the degradation pathway of amino acids to pyruvate and tricarboxylic acid cycle intermediates was favoured contrary to the degradation pathway of ketogenic amino acids. Special considerations should be taken concerning Thr, Ile, Ser, Ala, Asx (Asp, Asn), Glx (Glu, Gln) (because of their fast intake) and Lys and His (due to their stable contents) during a prolonged period of fasting.