Studies on the effect of temperature and pH on the inactivation of fish viral and bacterial pathogens

Disposal of fish by‐products in the European Community must comply with Regulation (EC) No 1069/2009 which categorizes animal by‐products according to risk, and specifies methods of disposal of by‐products according to that risk. There is provision under the regulation for composting or ensiling to be used for by‐products from aquatic animals. Biosecurity considerations require knowledge of the parameters of time and temperature, or time and pH, required to inactivate any fish pathogens that may be present. To provide those data, we undertook laboratory studies on the inactivation of a number of fish pathogenic viruses and bacteria at 60 °C, pH 4.0 and pH 12.0 as a preliminary to conducting subsequent trials with the most resistant viruses and bacteria in fish tissues. The most resistant bacterium to 60 °C, pH 4.0 as well as pH 12.0 was Lactococcus garvieae. Its concentration was reduced to the level of sensitivity of the test after 24–48 h exposure to 60 °C, but it survived for at least 7 days at pH 4.0 and 14 days at pH 12.0. The most resistant virus to 60 °C was infectious pancreatic necrosis virus, and to pH 12.0 was infectious salmon anaemia virus. The majority of the viruses tested survived exposure to pH 4.0 for up to 28 days. The results suggest that the process of acid ensiling alone is not an effective method for the inactivation of many viral and bacterial pathogens, and fish by‐products would need further treatment by a method approved under the regulation following ensiling, whereas alkaline or heat treatment are likely to provide an increased degree of biosecurity for on‐farm processing of mortalities.     

高pH胁迫对克氏原螯虾的急性毒性和鳃、肝胰腺中酶活性及组织结构的影响

研究了高pH胁迫对克氏原螯虾鳃、肝胰腺中酶活性的变化,以及对鳃、肝胰腺组织结构的影响。在得出96 h L C_(50)的基础上,设置对照(pH7.6)和实验(pH10.2)2个pH处理组,进行96 h高pH胁迫,于胁迫后0、2、8、24和96 h测定鳃Na~+-K~+-ATP酶、乳酸脱氢酶(lactate dehydrogenase,LDH)、延胡索酸还原酶(fumarate reductase,FRD)、肝胰腺超氧化物歧化酶(superoxide dismutase,SOD)、过氧化氢酶(catalase,CAT)的活性以及丙二醛(malondialdehyde,MDA)的含量,并采集完整的鳃、肝胰腺做组织切片观察。结果发现,96 h LC50 pH值为10.194。高pH应激后,实验组鳃Na+-K+-ATP酶、LDH和FRD的活性呈上升趋势,pH胁迫8 h后与对照组差异显著(P0.05);随胁迫时间的延长,实验组肝胰腺MDA含量的变化趋势与LDH和FRD相似,CAT活性呈先上升后下降趋势,SOD活性起初变化不大,pH胁迫24 h后出现明显下降(P0.05)。鳃和肝胰腺的组织观察表明,随着pH胁迫时间的延长,鳃呼吸上皮细胞逐渐脱落,角质层受损、破裂,鳃叶受损程度逐渐加剧;肝胰腺小管基膜破损,小管内空泡增多、体积增大,肝细胞细胞数量减少。研究表明,高pH胁迫对克氏原螯虾代谢会产生影响,同时导致氧化应激,并会对鳃和肝胰腺的组织结构造成损伤,影响其生物学功能。     

Controlling the growth of a cyanobacterial contaminant, Synechoccus sp., in a culture of Tetraselmis chui (PLY429) by varying pH: Implications for outdoor aquaculture production

In aquaculture, large volumes of phytoplankton are often grown outdoors to reduce costs. However, growing microalgae in an environment not as well-controlled as in the laboratory can lead to unwanted phytoplankton, including cyanobacteria, contaminating a culture. A cyanobacterial contaminant was isolated from an outdoor culture of Tetraselmis chui (PLY429) at the Milford Laboratory. This study investigated the growth of PLY429 and the cyanobacterium in pure cultures and a mixed culture in a pH range of 6.5-9.5. The division of PLY429 was greater at a pH range of 7.0-8.0; whereas, for the cyanobacterium, higher growth was obtained at pH 8.0-9.0. Results from combined cultures of PLY429 and the cyanobacterium grown at various pHs indicated that maintaining pH near 7.1 yields higher growth of PLY429 than those of the cyanobacterium. These findings suggest that controlling pH may reduce the population of a cyanobacterial contaminant in an aquaculture feed culture.     

在淡水鱼类混养系统中吊养三角帆蚌对养殖产量和水质的影响

通过78 d围隔养殖实验检验了在草鱼、鲫、鲢、鳙混养系统中吊养三角帆蚌对鱼产量和水质的影响。设2个处理,处理Ⅰ混养草鱼、鲫、鲢和鳙,处理Ⅱ在处理Ⅰ基础上按鱼∶蚌=1∶1的比例配养三角帆蚌。实验期间定期采样分析浮游植物种类组成和生物量、初级生产力(P)、群落呼吸(R)、溶氧(DO)、pH、透明度(SD)、主要离子(CO2-3、HCO-3、Cl-、SO2-4、Ca2+、Mg2+)、总碱度、总硬度、氨态氮、亚硝酸态氮、硝酸态氮、活性磷、总氮(TN)、总磷(TP)、总有机碳(TOC)、高锰酸钾指数(COD M n)和生化耗氧量(BOD5)。结果发现,吊养三角帆蚌显著降低水体中Ca2+浓度,但对其他指标均无显著影响。处理Ⅱ草鱼、鲫、鲢产量略高于处理Ⅰ,而鳙产量略低于后者,表明在草鱼、鲫、鲢、鳙混养系统中按1∶1的比例配养三角帆蚌不会导致草鱼和鲫产量下降,但导致鳙产量降低。处理Ⅱ浮游植物多样性(Shannon-Weaver多样性指数、Margalef丰富度指数、Pielou均匀度指数和种类数)、P、P/R、SD和DO略高于处理Ⅰ,而氨态氮、活性磷、TN、TP、COD M n、BOD5和TOC略低于后者,表明在混养系统中配养三角帆蚌可显著降低养殖水体Ca2+浓度,同时可在一定程度上提高浮游植物多样性、P和DO并降低TN、TP、COD M n、BOD5和TOC。结果表明,在淡水鱼类混养系统中适度配养三角帆蚌可提高养殖的经济效益,同时有助于降低养殖系统内氮、磷和有机废物的积累。     

高温下芽孢杆菌对坛紫菜生长及生理的影响

坛紫菜是我国东海区栽培的重要经济海藻,其生长适温为20°C,极限高温为29°C。高温和病原菌均会导致坛紫菜生长受抑、藻体病烂。为研究高温下藻际微生物对坛紫菜生长的影响,本文以坛紫菜叶状体和藻际芽孢杆菌(实验证明该菌株在20°C下为紫菜的益生菌)为材料,检测了在28°C下藻际芽孢杆菌对坛紫菜相对生长率(RGR),抗氧化酶(SOD、POD)活性,可溶性蛋白、游离脯氨酸(Pro)、丙二醛(MDA)和藻胆蛋白含量等生理指标的影响。结果显示,28°C下,在实验初始阶段,菌藻共培养组坛紫菜比纯培养对照组有较高的RGR,但随着培养时间的延长,两组的RGR无显著性差异;4 d后两组坛紫菜均出现腐烂,菌藻共培养组的藻体腐烂程度更为剧烈;培养24 h后,菌藻共培养组的SOD、POD活性和MDA水平均显著高于对照组,而渗透调节物质可溶性蛋白和Pro显著低于对照组;共培养组中,坛紫菜的藻胆蛋白含量均显著高于对照组。该研究表明,28°C下,虽然在短期内藻际芽孢杆菌会促进坛紫菜的生长,但随着培养时间的延长反而不利于坛紫菜的生长,且加剧活性氧和渗透压胁迫,表明该菌的生态功能因环境变化而发生了改变。     

The effects of light and temperature on the photosynthesis of the Asparagopsis armata tetrasporophyte (Falkenbergia rufolanosa), cultivated in tanks

The integrated aquaculture of the tetrasporophyte of Asparagopsis armata Harvey (Falkenbergia rufolanosa) using fish farm effluents may be viable due to the species high capacity of removing nutrients and its content of halogenated organic compounds with applications on the pharmaceutical and chemical industries. In order to optimize the integrated aquaculture of F. rufolanosa, we followed the daily variation of the potential quantum yield (F_v/F_m) of PSII on plants cultivated at different biomass densities and different total ammonia nitrogen (TAN) fluxes to check if they are photoinhibited at any time of the day. Moreover, the photoinhibition under continuous exposure to highly saturating irradiance and its potential for subsequent recovery in the shade was assessed. The potential for year round cultivation was evaluated by measuring rates of O₂ evolution of plants acclimated at temperatures ranging from 15 to 29 °C, the temperature range of a fish farm effluent in southern Portugal where an integrated aquaculture system of F. rufolanosa was constructed.Photoinhibition does not seem to be a major constrain for the integrated aquaculture of F. rufolanosa. Only when cultivated at a very low density of 1.5 g fresh weight (FW) l^− 1 that there was a midday decrease in maximal quantum yield (F_v/F_m). At densities higher than 4 g FW l^− 1, no photoinhibition was observed. When exposed to full solar irradiance for 1 h, F. rufolanosa showed a 33% decrease in F_v/F_m, recovering to 86% of the initial value after 2 h in the shade. A midday decline of the F. rufolanosa F_v/F_m was also observed under the lowest TAN flux tested (∼6 μM h^− 1), suggesting that this fast and easy measurement of fluorescence may be used as a convenient diagnostic tool to detect nutrient-starved unbalance conditions of the cultures. Maximum net photosynthesis peaked at 15 °C with 9.7 mg O₂ g dry weight (DW)^− 1 h^− 1 and remained high until 24 °C. At 29 °C, the net oxygen production was significantly reduced due to a dramatic increase of respiration, suggesting this to be the species' lethal temperature threshold.Results indicate that F. rufolanosa has a considerable photosynthetic plasticity and confirm it as a good candidate for integrated aquaculture at temperatures up to 24° C and cultivation densities of at least 5 g FW l^− 1. When cultivated at these densities, light does not penetrate below the first few centimetres of the surface zone. Plants circulate within the tanks, spending around 10% of the time in the first few centimetres where they are able to use efficiently the saturating light levels without damaging their photosynthetic apparatus.     

Influence of pH and temperature on force development and shortening velocity in skinned muscle fibres from fish

Three species of fish were studied: Atlantic cod (Gadus morhua), sculpin (Myoxocephalus scorpius) (from the North Sea, temperature 2 to 12°C) andNotothenia neglecta (from Antarctica, temperature –2 to +2°C). Single fast muscle fibres were isolated from anterior myotomes and skinned with detergent in order to directly determine the effects of pH and temperature on force production and shortening velocity.In all species maximum force production (Po) was independent of pH over the range 7.3–8.0. Decreasing the pH from 7.3 to 6.6 reduced maximum force by 28% in fibres fromG. morhua andN. neglecta but had no effect on fibres fromM. scorpius. The depression in maximum force with acidosis was accompanied by a proportional decrease in stiffness and an increase in the rate of force recovery after stretch.Unloaded contraction velocity of cod fibres (Vmax) showed a pH optimum at around pH 7.6 decreasing by 31% at pH 6.6. Vmax of fibres from the other species was independent of pH over the range 6.6–8.0.The effects of pH on Po and Vmax were similar at 0 and 10°C. Thus for maximally activated fibres both force and contraction velocity are independent of temperature induced changes in pH. In some species acidosis depresses contractility and is likely to be a contributory factor to muscle fatigue.     

Cryopreservation of dead fish spermatozoa several hours after death of Indian major carp, Labeo rohita and its successful utilization in fish production

Cryopreservation of semen collected from dead fishes showed that it is possible to use it for fish production. We humanely killed individuals of the Indian major carp rohu, Labeo rohita and stored at different temperature regimes of 31 °C, 0 °C, − 10 °C and − 30 °C till 8 h. At every one hour interval semen from these fishes were collected by Pasteur pipette and evaluated for sperm yield/kg body weight, motility, pH, spermatocrit (%) and sperm count. The semen having suitable characteristics such as; 70% spermatocrit or above and motility index of 4 or above was cryopreserved by following a specific protocol. The cryopreserved semen of the dead fishes was stored for 7 days and then thawed in a water bath at 37 °C for 50 s. It was found that up to 8 h, spermatozoa of rohu were viable when stored at 0 °C or − 10 °C. Sperm collected after 8 h of fish death and maintained at 0 °C was the best stored condition that showed 30% larval survival. The spermatozoa collected 8 h after fish death was mostly normal as observed under scanning electron microscope and the total length of rohu spermatozoa was 25-30 μm. The hatchlings produced with this cryopreserved semen grew normally and juvenile fishes of rohu could be produced. This study suggests that germ cells such as spermatozoa of dead fishes can be cryopreserved and utilized for restoration of a species. It has the potential use in cryo-conservation of endangered fishes, restoration of animals through fertilization and genetic manipulation studies.     

抑食金球藻对卤虫和鼠的急性毒性及对兔皮肤和眼的刺激作用

2011年夏季,我国秦皇岛沿岸海域再次暴发了大面积的微微型浮游植物抑食金球藻褐潮。为了弄清该藻的毒性以及评估其对人们生命健康的影响,本实验以卤虫作为受试生物,在室内条件下测试了该褐潮水体对卤虫的急性毒性及卤虫对该藻的摄食作用,并以室内纯种培养的抑食金球藻对小白兔进行急性皮肤刺激和眼刺激实验,对小白鼠进行急性经口毒性实验。结果表明,抑食金球藻褐潮水体对卤虫无急性毒性,卤虫能以抑食金球藻为食,摄食率随着摄食藻密度的升高而上升。抑食金球藻藻液对实验兔的皮肤和眼睛无明显刺激作用,对小白鼠无急性毒性作用。该藻暴发褐潮时,对人们的生命健康安全影响较小。     

Distribution of a fish pathogen Listonella anguillarum in the Japanese flounder Paralichthys olivaceus hatchery

The present study was undertaken to investigate the distribution of Listonella anguillarum in a Japanese flounder (Paralichthys olivaceus) hatchery. A total of 2704 isolates were obtained from the developing fish, live diets and artificial feeds of Japanese flounder and their rearing water, 439 of which were identified as L. anguillarum by the combining incubation on thiosulfate-citrate-bile salt-sucrose (TCBS) agar at 35 °C overnight with polymerase chain reaction (PCR) detection for the VAH1 hemolysin gene. L. anguillarum was detected in all seven rotifer samples, with densities of 2.5 × 10³ to 4.6 × 10⁶ colony forming units (CFU) g^− 1. Both the analyzed samples of Nannochloropsis oculata contained this bacterium at densities of 1.6 × 10⁴ to 1.4 × 10⁵ CFU g^− 1. L. anguillarum was detected in only one of four samples of Artemia nauplii with a density of 4.8 × 10⁵ CFU g^− 1 (35%) and it was not detected in the two analyzed artificial feed samples. L. anguillarum was detected in 11 of 18 specimens of larval and juvenile Japanese flounder at densities of 5.0 × 10¹ to 7.4 × 10⁵ CFU g^− 1, while it was not detected in the two analyzed egg specimens of Japanese flounder. These results indicate that L. anguillarum associated with the developing Japanese flounder is likely derived from rearing water and live diets such as rotifers. Further, it is strongly suggested that L. anguillarum is a transient bacterium of the intestinal microflora for the Japanese flounder but is a permanently indigenous one for the Japanese flounder hatcheries.     

注射LHRH-A2对刀鲚的催产效果及其雌鱼血液生化指标的影响

为了丰富刀鲚人工繁殖理论,探明LHRH-A2对刀鲚的催产效果及其雌鱼血液生化指标的影响,本研究以2冬龄人工养殖刀鲚为研究对象,通过注射LHRH-A2进行人工催产,并分析催产效果;同时应用放射性免疫、化学发光等方法,对人工催产过程中刀鲚雌鱼的血液生化指标进行了系统研究。结果显示,按照雌鱼30μg/kg、雄鱼减半的剂量注射LHRH-A2,刀鲚亲鱼死亡率为16.67%±3.34%;存活刀鲚亲鱼群体中,效应时间为21~24 h,催产率为83.33%±3.34%,受精率为75.06%±6.19%。注射催产素21 h内,血浆中17β-雌二醇(17β-estradiol,E2)、催乳素(prolactin)、甲状腺素(thyroxine,T4)、皮质醇(cortisol)、总蛋白(total protein,TP)、乳酸(lactate)在人工催产过程中表现出先上升后下降的规律;三碘甲腺原氨酸(three iodine thyroid,T3)、甘油三酯(triglyceride,TG)、Cl–呈现持续下降趋势;谷丙转氨酶(alanine transaminase,ALT)、谷草转氨酶(aspartate transaminase,AST)、渗透压(osmotic pressure)、K+、Na+呈现持续上升的趋势。比较以上血浆指标在产卵组和未产卵组的实验结果,产卵组形成高皮质醇、TP、TG、渗透压、Na+、K+的血浆环境和低E2、催乳素、乳酸、T4、T3、Cl-、ALT、AST的血浆环境,而未产卵组的这些指标与产卵组相反。结果表明,人工催产过程对肝脏、肾脏等正常功能造成了影响,从而抑制催产素LHRH-A2对卵子的正向调控作用。本研究初步揭示了人工催产应激对刀鲚雌鱼成功排卵的影响机制,为进一步优化刀鲚的规模化人工繁殖提供科学依据。     

Effect of saponin on hematological and immunological parameters of the giant freshwater prawn, Macrobrachium rosenbergii

Giant freshwater prawns, Macrobrachium rosenbergii (17.9 ± 2.7 g), exposed to different concentrations of saponin at 0, 0.3, 0.6, 0.9 and 1.2 mg l^− 1 for 168 h were examined for osmolality, electrolyte levels, oxyhemocyanin, protein levels, acid-base balance status, total hemocyte count (THC), phenoloxidase activity, and respiratory bursts. Hemolymph oxyhemocyanin, protein, and pO₂ were inversely related to the saponin concentration. Hemolymph oxyhemocyanin, protein, pO₂, pCO₂, and pH of prawns exposed to 1.2 mg l^− 1 saponin were significantly lower than those of prawns exposed to 0.3 mg l^− 1 and control solutions. However, no significant difference was observed in osmolality or electrolyte levels of prawns exposed to different concentrations of saponin for 168 h. The THC of prawns following 168 h of exposure to 0.9 and 1.2 mg l^− 1 saponin increased, but the phenoloxidase activity decreased suggesting that the decrease in phenoloxidase activity under saponin stress was not a consequence of the increase in THC. We concluded that saponin at as low as 0.9 mg l^− 1 decreases the respiratory protein level and acid-base balance, and modulates the immune system of M. rosenbergii.     

温度和pH值对斜带髭鲷蛋白酶、淀粉酶活性的影响

研究了温度和pH值对斜带髭鲷Hapalogenys nitens蛋白酶、淀粉酶活力的影响。结果表明,斜带髭鲷胃内pH值范围为4.9～5.4;肝胰脏pH值为5.8～6.2;肠道pH值为6.5～6.9。蛋白酶活性随温度的上升而增加,在40℃达最大,40℃以上酶活性随温度的上升而下降。在不同消化器官中,蛋白酶活性大小顺序为：胃〉前肠〉幽门盲囊〉后肠〉肝胰脏。在15～50℃范围内,斜带髭鲷消化道不同部位淀粉酶活性的最适温度均为35℃,淀粉酶活性由高到低顺序为：肝胰脏〉幽门盲囊〉前肠〉后肠〉胃。在pH值为2.2～7.6范围内,胃蛋白酶活性的最适pH值为2.8;在pH值为4.8～8.0范围内,其他消化器官蛋白酶的最适pH值均为7.2;在最适pH值下,各消化器官中的蛋白酶活性由高到低顺序为：前肠〉幽门盲囊〉后肠〉肝胰脏。在pH值为4.8～8.0范围内,胃淀粉酶活性的最适pH值为6.0,肠、肝胰脏与幽门盲囊淀粉酶的最适pH值均为6.8,活性由高到低顺序为：肝胰脏〉幽门盲囊〉前肠〉后肠〉胃。在最适温度和pH值下,蛋白酶活性由高到低顺序为：胃〉前肠〉幽门盲囊〉后肠〉肝胰脏;淀粉酶活性由高到低顺序为：肝胰脏〉幽门盲囊〉前肠〉后肠〉胃。     

温度和体质量对军曹鱼生长及氮收支的影响

为调查军曹鱼生长和氮收支与温度和体质量的关系,采用鱼类生物能量学的原理和方法,开展了不同温度(21、27和33 ℃)和体质量(10、20、50和100 g)对军曹鱼生长和氮收支的影响研究,并建立了生长——温度/体质量关系及不同温度和体质量条件下的氮收支方程。结果显示:(1)温度和体质量对军曹鱼生长影响显著,且两者之间存在交互作用。军曹鱼在实验的最高水温(33 ℃)及最小体质量(10 g)时生长最快,在实验的最低水温(21 ℃)及最大体质量(100 g)时生长最慢。(2)实验的体质量范围内,军曹鱼特定生长率均随水温升高呈显著增长趋势;在实验的温度范围内,军曹鱼特定生长率随体质量增加呈减速下降趋势,两者间的关系采用幂函数定量描述。多元回归分析表明,模型LnSGR=a+bLnW+cT+dT²+eTLnW能较好地拟合军曹鱼特定生长率、温度及体质量间的关系。(3)军曹鱼摄食氮、排粪氮、排泄氮和生长氮受温度和体质量影响显著,随温度升高而增加,随体质量增加趋于减少。(4)不同温度和体质量组军曹鱼氮收支方程存在差异,且温度对氮收支方程的影响大于体质量。对同一体质量组,水温27和33 ℃时生长氮占食物氮的比例明显高于水温21 ℃时,而水温27和33 ℃时代谢氮占食物氮的比例则明显低于水温21 ℃时;对同一温度组,军曹鱼氮收支方程变幅较小。(5)在所有氮收支方程中,摄食氮用于排泄的比例大于68%。研究表明,在实验的温度和体质量范围内,环境温度偏低不利于军曹鱼生长;排泄是军曹鱼氮支出的最主要途径;水温27~33 ℃时军曹鱼氮收支方程相对恒定。     

Effect of temperature on growth and energy budget of juvenile cobia (Rachycentron canadum)

Growth and energy budget of juvenile cobia (initial body weight ∼ 22 g) at various temperatures (23, 27, 31 and 35 °C) were investigated in this study. Maximal ration level (RL_max, %/day) increased as temperature (T, °C) increased from 23 °C to 31 °C but decreased at 35 °C, described as a quadratic equation: RL_max = −0.023T² + 1.495T − 17.52. Faecal production (f, mg g^− 1 day^− 1) increased with increased temperature (T, °C), described as a power function: lnf = 0.738lnT − 0.806. As temperature increased, feed absorption efficiency in dry weight (FAE_d, %), protein (FAE_p, %) and energy (FAE_e, %) all increased first and then decreased, but the variation of feed absorption efficiency was small, with ranges of 89.59-91.08%, 92.91-94.71%, 93.92-95.32%, respectively. Specific growth rate in wet weight (SGR_w, %/day), dry weight (SGR_d, %/day), protein (SGR_p, %/day) and energy (SGR_e, %/day) showed a domed curve relative to temperature (T, °C), described as quadratic equations: SGR_w = − 0.068T² + 3.878T − 50.53, SGR_d = − 0.079T² + 4.536T −59.64, SGR_p = − 0.084T² + 4.783T − 63.08 and SGR_e = − 0.082T² + 4.654T − 60.99, and SGR_w, SGR_d, SGR_p and SGR_e maximized at 28.5 °C, 28.6 °C, 28.4 °C, 28.5 °C, respectively, as calculated from the regression equations. The relationships between feed conversion efficiency in wet weight (FCE_w, %), dry weight (FCE_d, %), protein (FCE_p, %), energy (FCE_e, %) and temperature (T, °C) also took on a domed curve described as quadratic equations: FCE_w = − 0.726T² + 39.71T − 473.8, FCE_d = − 0.276T² + 15.31T − 190.6, FCE_p = − 0.397T² + 22.05T − 277.9 and FCE_e = − 0.350T² + 19.39T − 239.9, and FCE_w, FCE_d, FCE_p and FCE_e maximized at 27.4 °C, 27.8 °C, 27.7 °C and 27.7 °C, respectively, as calculated from the regression equations. Energy budget of juvenile cobia fed satiation was 100C = 5F + 67(U + R) + 28G at water temperature 27 °C and 100C = 5F + 70(U + R) + 25G at water temperature 31 °C, where C is food energy, F is faeces energy, (U + R) is excretion energy and metabolism energy, and G is growth energy.     

两种常见淡水鱼在高温储存过程中挥发性盐基总氮和生物胺的含量变化

实验检测了条和红鳍红鲌在30℃下贮存48 h内每隔6 h的挥发性盐基总氮(TVBN)和9种生物胺(尸胺、腐胺、组胺、酪胺、5-羟色胺、亚精胺、精胺、多巴胺和章鱼胺)含量的变化,并对TVBN和生物胺含量与时间的相关性进行分析,以期为淡水水产品类饵料安全投喂和人类食品安全提供基础资料。结果显示,两种鱼在相同贮存条件下TVBN和生物胺含量均存在一定差异。其中,TVBN含量均随着贮存时间的延长而逐渐增加且与时间存在明显的相关性。红鳍红鲌除在42 h时TVBN含量低于■条外,其他时间均高于■条。在30℃贮存下,■条和红鳍红鲌的TVBN含量均在12 h时超过30 mg/100 g,因此,这两种鱼在12 h后不推荐食用。尸胺、腐胺、酪胺和组胺是这两种鱼主要的生物胺类型,且有随贮存时间延长含量显著增高的趋势,并与时间呈显著性相关,可推荐用于评价此两种水产品食用安全的生物胺类型。除此之外,在可测到的生物胺中,红鳍红鲌的尸胺和组胺含量明显高于■条,且红鳍红鲌体内的5-羟色胺与时间也呈显著性相关。     

剑尾鱼IgZ基因的克隆及疫苗免疫对其在组织中表达的影响

为研究剑尾鱼Ig Z基因序列及其在疫苗免疫后的表达规律,本实验根据已知的EST库设计引物,利用RT-PCR等方法,获得剑尾鱼Ig Z基因c DNA全长序列,并进行生物信息学分析和疫苗免疫后组织表达分析。结果显示,剑尾鱼Ig Z基因全长序列为5 420 bp,包含4个外显子和3个内含子;c DNA序列全长1 527 bp,包含一个1 299 bp的完整ORF框,该基因编码432个氨基酸,预测其编码蛋白的分子量大小为47.48 ku,并具有Ig Z的基本结构,与其他硬骨鱼类Ig Z氨基酸序列一致性为28%~54%。荧光定量PCR分析结果显示,Ig Z基因主要在剑尾鱼的头肾、脾脏和肠中分布,且疫苗免疫后11天内,Ig Z基因在头肾、脾脏和肠组织中均表现为先上升后下降的趋势。头肾中Ig Z基因的表达量在第4天时最高,为对照组的2.12倍;脾脏中Ig Z基因在第4天时呈现最高峰,为对照组的4.65倍;肠组织中Ig Z基因的表达量在12 h时有一个小高峰,第2天时最高,为对照组的11.41倍。本研究获得了剑尾鱼Ig Z基因c DNA全长序列,并对其表达规律进行了初步研究,发现Ig Z在肠道黏膜免疫中具有重要作用,这将为进一步验证其在黏膜免疫中的作用以及剑尾鱼作为疾病研究模式动物和疫苗免疫评价模型奠定基础。     

珠江口海域双胞旋沟藻赤潮对卤虫幼体、鱼苗和虾苗的急性毒性

为研究2009年10月下旬在广东珠海海域爆发的双胞旋沟藻赤潮对养殖鱼类及水体中其它生物的影响,实验以卤虫幼体、金鼓鱼苗和凡纳滨对虾苗作为受试生物,在赤潮现场测试双胞旋沟藻对卤虫幼体、鱼苗和虾苗的急性毒性效应。结果显示,24 h双胞旋沟藻对卤虫幼体的LC₅₀(半致死浓度)为9.55×10⁴/mL,藻密度为2.5×10³/mL的双胞旋沟藻对卤虫幼体的LT₅₀(半致死时间)为48.5 h。60 h内该赤潮水体对鱼苗和虾苗的存活无不利影响,卤虫幼体和金鼓鱼苗均可摄食双胞旋沟藻,卤虫幼体对双胞旋沟藻的摄食率低。研究表明,双胞旋沟藻赤潮水体对卤虫幼体有一定的毒性作用,但在低藻密度条件下,卤虫幼体能以该藻为食并维持其生命,双胞旋沟藻对金鼓鱼苗和凡纳滨对虾苗无急性毒性作用。     

纳米ZnO和常规ZnO、ZnSO4对斑马鱼毒性效应的比较

为比较纳米ZnO、常规ZnO和ZnSO_4对斑马鱼氧化应激毒性的强弱,探究纳米ZnO的毒性作用与其释放的Zn2+和本身特性的关系,研究了纳米ZnO、常规ZnO、ZnSO_4对斑马鱼肝脏、肠、鳃组织中抗氧化酶活性及炎症凋亡基因表达的影响。实验将斑马鱼分别暴露于纳米ZnO、常规ZnO、ZnSO_4水体中,在4、24和96 h后,用分光光度法检测斑马鱼肝脏、肠、鳃中过氧化氢酶(CAT)、谷胱甘肽(GSH)、谷胱甘肽硫转移酶(GST)、活性氧自由基(ROS)的变化;采用荧光定量PCR技术,测定实验组中肝脏、肠和鳃中Bax、Bcl-2、TNF-α及IL-6的mRNA相对表达量。结果显示,纳米ZnO、常规ZnO、ZnSO_4均引起斑马鱼各组织的氧化应激反应,且使组织中凋亡基因和炎症基因表达水平发生变化,激活生物体内的细胞坏死和细胞凋亡途径,引起细胞死亡或机体炎症,其中纳米ZnO的致氧化损伤作用最强。研究表明,纳米ZnO对斑马鱼的氧化应激毒性强于常规ZnO和ZnSO_4,而纳米颗粒本身特性是导致纳米ZnO毒性作用的主要原因。     

嗜水气单胞菌引致的金钱鱼细菌性疾病

为了分离鉴定实验室养殖的金钱鱼暴发性疾病的病原,利用传统病原分离的方法,从病鱼肝脏分离得到一株G–短杆菌(Ah201416),对其进行电镜观察和生理生化鉴定,对病鱼组织进行病理切片分析,并根据科赫法则,用分离株对健康金钱鱼进行人工感染。结果显示,该菌株电镜下观察细菌大小为0.8~1.0μm×1.0~2.0μm(宽×长),无芽孢和荚膜,生理生化特性与嗜水气单胞菌基本一致;16S r RNA序列(登录号:KR006248)与Gen Bank中嗜水气单胞菌(Aeromonas hydrophila,Ah ATCC7966)的16S r RNA基因序列的同源性高达99%,系统进化树与Ah聚为一支。病理切片分析发现,发病金钱鱼与正常金钱鱼相比,鳃小片细胞不同程度地脱落,伴有白细胞浸润;肠绒毛结构消失,肌肉层疏松明显;肾脏中肾小管细胞脱落,管腔中有坏死细胞,并出现不同程度的颗粒变性;肝脏细胞形态不规则,伴有血细胞浸润等病理损伤。人工感染实验表明,其对健康金钱鱼96h的半数致死剂量(LD50)为7.35×107 CFU/m L。研究表明,发病金钱鱼中分离的Ah201416菌株为嗜水气单胞菌,丰富了金钱鱼细菌性病原的研究,此结果为金钱鱼细菌性疾病的防治和养殖业的健康发展提供了重要的理论依据。