The encysted dormant embryo proteome of Artemia sinica

The possibility of the brine shrimp Artemia to produce dormant embryo (cysts) in diapause is a key feature in its life history. In the present study, we obtained a proteomic reference map for the diapause embryo of Artemia sinica using two-dimensional gel electrophoresis with a pH range of 4-7 and a molecular weight range of 10-100 kDa. Approximately 233 proteins were detected, and 60 of them were analyzed by capillary liquid chromatography tandem mass spectrometry (LC-MS/MS). Of these, 39 spots representing 33 unique proteins were identified, which are categorized into functional groups, including cell defense, cell structure, metabolism, protein synthesis, proteolysis, and other processes. This reference map will contribute toward understanding the state of the diapause embryo and lay the basis and serve as a useful tool for further profound studies in the proteomics of Artemia at different developmental stages and physiological conditions.

中华卤虫滞育卵发育和重金属刺激相关蛋白质组学研究

卤虫(Artemia)是一种广温、耐高盐的小型甲壳动物，广泛分布于内陆盐湖和沿海盐田中。卤虫的无节幼体作为重要的蛋白优质饵料，被广泛的应用于水产养殖生产。卤虫具有特殊的生物学特性，是研究甲壳动物胚胎发育的良好的实验材料，同时也是一种研究动物抗逆机制的模式动物。卤虫有卵生和卵胎生两种繁殖后代的方式，当环境条件适宜时，卤虫倾向于采取卵胎生方式，即直接产生无节幼体；而在恶劣的环境条件下，卵生方式占主要地位，产生处于滞育状态的、具有复杂外壳的休眠卵。卤虫的滞育卵具有独特的生物学特性和特殊的生理生化特点。其发育停滞，细胞分裂停止，酶活力下降，代谢活动受到抑制并可耐受各种极端恶劣环境，如缺氧、低温、紫外线、干燥等。即使在最适的环境中滞育卵的孵化率也很低，只有受到某些特定的非生物信号的刺激才自能终止这种滞育状态，恢复生理代谢；当环境条件适宜时，能够继续发育孵化成无节幼体。因此，卤虫的滞育卵在卤虫的整个生活史中占有重要的地位。另一方面，卤虫是极端环境生物，能够抵抗各种恶劣环境胁迫刺激，因此是研究抗逆机理的良好的实验动物。 本论文利用蛋白质组学技术，研究了卤虫滞育卵及滞育卵发育过程中的蛋白质组表达情况，并研究了卤虫幼体在重金属刺激后蛋白表达的变化情况。得到如下结果： 建立了中华卤虫滞育卵可溶性总蛋白的双向凝胶电泳对照图谱。在pH 4–7、分子量10-100 kDa范围内，检测到约 233个蛋白点，并利用高效液相色谱-质谱联用（LC-ESI-MS/MS）技术鉴定了其中的48个丰度较大及感兴趣的蛋白点，根据这些蛋白的生物学功能进行分类，功能类别包括细胞防御蛋白、抗氧化蛋白、细胞骨架蛋白、代谢相关蛋白等。在卤虫滞育卵中共分离鉴定到6个分子量和等电点存在差异的小热休克蛋白p26的异构体，生物信息学分析表明该蛋白有三种不同的功能位点，分别是蛋白激酶C磷酸化位点，Casein 激酶II磷酸化位点及 N-myristoylation 位点。 采用低温脱水的方法对滞育卵进行激活刺激，并对活化卵和滞育卵蛋白表达图谱进行了对比分析。结果表明对卤虫滞育卵的激活刺激引起了其蛋白表达的明显变化。活化卵图谱中蛋白点总数比滞育卵中明显增多，特别是在pI<5.5范围内。约70个蛋白点在激活刺激后上调表达，包括部分只在激活卵中表达的蛋白；25个下调表达，包括部分只在滞育卵中表达的蛋白；其余约60％（占滞育卵蛋白点数目百分比）的蛋白点表达量基本恒定。热休克蛋白家族、抗氧化蛋白家族成员等蛋白变化明显，小热休克蛋白p26、小热休克蛋白ArHsp21蛋白以及过氧化物还原酶异构体在激活卵中特异表达。 活化卵孵化过程中不同发育时期的蛋白表达又呈现出不同的特点，分别在孵化后6h、12h、18h和24h的蛋白质组学图谱上检测到267、285、195和210个蛋白点。孵化后6h和12h休眠卵蛋白表达个数相对较多，与胚胎发育过程中的器官发生和剧烈的形态变化相适应；孵化后18h和24h休眠卵蛋白表达明显下降，部分蛋白的表达关闭，部分蛋白开始富集表达。 利用双向凝胶电泳技术分析了中华卤虫幼体受到急性硫酸铜刺激后的蛋白表达变化情况。通过图谱对比分析，检测到了5mM硫酸铜刺激24h后，卤虫幼体中14个差异表达的蛋白点。利用LC-ESI-MS/MS技术鉴定了其中的7个蛋白，其中3个蛋白上调表达，分别是热休克蛋白70（7.5倍）, 肌动蛋白（2.3倍）和伴侣分子亚基1（3.0倍）。3个蛋白下调表达，分别是：精氨酸激酶（2.8倍）, 延伸因子2 (2.0倍） 和富含甘氨酸蛋白(2.0倍）。硫酸铜刺激后特异表达的一个蛋白被鉴定为过氧化物还原酶(Peroxiredoxin，Prx)。根据质谱检测提供的蛋白肽段信息和其他生物过氧化物还原酶保守氨基酸序列设计简并引物，结合RACE技术，从中华卤虫幼体中克隆到了过氧化物还原酶基因，该基因的cDNA全长为756个碱基，其中开放阅读框为594个碱基，编码198个氨基酸，其蛋白理论分子量为22.0 kDa，理论等电点为6.98。多序列比对结果显示中华卤虫Prx基因的推导氨基酸序列与美国卤虫和中国对虾的同源性高达98%和94%。实时荧光定量PCR结果显示，硫酸铜刺激后，该基因在卤虫无节幼体中的转录水平明显升高，在24h达到正常水平的3.0倍。

Eclosión de huevos de artemia (Artemia franciscana, Kellogg) a nivel de Laboratorio en la Universidad Nacional Agraria Managua, Niaragua

En el estudio de eclosión de Artemia franciscana a nivel de laboratorio, tuvo como objetivo determinar el tipo de tratamiento que se adapta a condiciones de laboratorio brindando buenos resultados para la eclosión del crustáceo Artemia ( Artemia franciscana Kellogg). La metodología utilizada consistió en la toma de datos en el laboratorio evaluando los siguientes tratamientos: Toma de pH de los ensayos; Toma de temperatura; Salinidad; Medición de Sobrevivencia. Una vez establecido el estudio se procedió a poner el huevo de Artemia, en lo cual se pusieron a eclosionar durante 48 horas para iniciar el proceso eclosivo del crustáceo. El manejo que se aplicó en el estudio consistió en tomar datos de temperatura y del pH para llevar un control y seguimiento a la eclosión de Artemia así como el control de la alimentación que consistía a base de una dieta de pastilla de alga S pirulina máxima triturada. De acuerdo al análisis estadístico realizado con metodología de DUNCAN, correlación de Pearson y procesado en el programa estadístico INFOSTAT indican que la mejor eclosión se obtuvo en el tratamiento D con un 65.40% dando pautas de ser el tratamiento que mejor se adapto a las condiciones de laboratorio. La sobre vivencia de los otros tratamientos es positiva y adaptable a las condiciones de laboratorio donde es recomendable utilizar recipientes con mayor capacidad de almacenamiento de agua. Recomendándose para futuros cultivos de Artemia se puede llevar a cabo el manejo ya implementado en el Laboratorio de Ciencias Biológicas de la Universidad Nacional Agraria para un mayor rendimiento en la sobre vivencia del estudio. Y seguir proporcionando información para la eclosion de dicho crustáceo que representa una opción alimenticia de bajo impacto a productores pesqueros del país.

Post-larval feeding of Clarias gariepinus (Burchell, 1802) on cultured zooplankton and artemia diets

The findings are presented of a study undertaken to provide baseline information that may be employed to develop a simplified hatchery technique for postlarval feeding of Clarias gariepinus on zooplankton cultured indoors in small aquaria. Three series of experiments were undertaken to identify the type and quantity of organic manure that would produce maximum densities of zooplankton in the shortest time in indoor cultures. Results suggest the possibility of the culture of natural zooplankton populations for use in larval feeding of C. gariepinus fry. Its effective hatchery application would bring about reduction in the unit cost per fingerling and ensure continuous hatchery operations even in the absence of Artemia cysts

El coste de la osmorregulación: Efecto de la salinidad sobre la tasa metabólica del crustáceo Artemia salina

El efecto de diferentes salinidades en la tasa metabólica de la artemia

Toxicity of heavy metals to brine shrimp Artemia

Bioassay were carried out on 48h cultured nauplii of brine shrimp Artemia by exposing them to seven trace elements viz. copper (Cu), zinc (Zn), cadmium (Cd), nickel (Ni), lead (Pb), iron (Fe) and manganese (Mn). Synergistic effects of all these elements and additive effects of Cu and Zn, Cd and Pb, and Ni and Fe were also investigated. Comparatively, the degree of toxicity for compound bioassays was higher than individual simple tests. The values were averaged and the expected median lethal concentration [LC sub(50)] of tested heavy metals was obtained by probit analysis on the basis of cumulative numbers of dead organisms after 24 and 48h. The order of toxicity of the metals to Artemia was Pb>Cd>Cu>Ni>Zn>Fe>Mn. Potency ratios of the seven metals were also calculated. The 24 and 48h variations obtained in LC sub(50) values were significantly different and relative implications of these are discussed.

Successful inoculation of Artemia and production of cysts in the coastal saltpans of Bangladesh II

Bangladesh has no naturally occurring Artemia, and all the growing shrimp hatcheries of the country depend entirely on import of cysts from foreign countries. Following successful inoculation of Artemia and production of cysts for the first time in this country in a coastal saltpan (at Chanua, Banskhali) by the senior author (in 1989-90), a similar second attempt was made under this programme in a saltpan (1000 m super(2)) of Demoshia, Chakaria, Cox's Bazar, Bangladesh between January and April 1992. A total of 1639.9 g (dry weight) of cysts (i.e. 5.46 kg DW/ha/month) have been produced using the Red Jungle Brand, whereas the previous attempt obtained 517 g of cysts (i.e. 2.07 kg DW/ha/month) using the Great Salt Lake Brand.

Effect of decapsulation on viability and hatching performance of Artemia cysts at different salinity levels

Artemia cysts were produced from the traditional solar salt works of Bangladesh through different fertilization treatments were tested for viability and hatching performance in different forms, such as processed and preserved, processed and decapsulated and unprocessed and undecapsulated. Decapsulated cysts performed maximum hatching (86.0%) in 20ppt salinity during 48 hours of incubation. The hatching percentage by the unprocessed and undecapsulated cysts were very low (12.0- 18.7%) in all the tested salinity grades.

Effects of culture treatments on HUFA levels in Artemia cysts

Artemia cysts (of GSL, Utah, USA origin) were produced from the modified traditional solar salt works of Bangladesh during winter months through different feeding/fertilization treatments (T1, T2, T3, T4 ) were analyzed to understand the effects of treatments on their fatty acid profile. Palmitic, Linolenic, Eicosapantaenonic and Docohexaenoic acids (mg/g. DW) were found highest for the cysts in T1 (16.0% ±1 .36), T2 (14.7% ±0.47), T3 (4.7% ±0.40) and T4 (0.7% ±0.06) treatments, respectively. High amount of 18:3(n-3) acids in the cysts of all sources proves to be freshwater type of the cysts. The presence of marine type essential fatty acids in the cysts of all sources were found low for 20:5n-3 (3.7-4.7%) and very low for 22:6n-3 (0.09-0.7%). No significant variation was observed for 16:0 acids within the treatments, but for 18:3(n-3) acid, the variation was found highly significant (P= 0.0052) between T2 and T4 treatments. For 20:5(n-3), only variation between T2 and T4 was found insignificant (P=0.1161), but between other treatments, significant variation was observed between T2 and T4 (P=0.0241), T2 and T4 (P=0.0022) and T1 andT4 (P=0.0161). No significant variation was found in other treatments.

Substitution of Artemia with Brachionus in different ratios during larval rearing of Macrobrachium rosenbergii and evaluation of their feed efficacy

Larval growth during stage I-VIII was studied in Macrobrachium rosenbergii. Duration in moult periodicity were recorded-during larval development period, larvae were fed with Brachionus (grown on Baker's yeast and also Brachinous raised through organic manuring in outdoor culture containers). The performance of the feed was evaluated through substitution of Brachionus in the feeding protocol, in lieu of Artemia 1st instar. The Artemia, Brachionus substitution ratio of 75:25 was found to be most efficient. The study also indicates that the comparative growth rate of Brachionus plicatilis is higher in manure loaded tanks than with Baker's yeast. Growth rate "Y'' in culture tank being 0.245 and 0.112 and corresponding duplicating time (Td) too was found to be 2.855 and 6.365 respectively in tanks manured/enriched with pig manure.

Effect of different feeds and water salinities on the cyst production of brine shrimp, Artemia sp.

Effect of different feeds and salinities on cyst production in Artemia sp. was studied. Among the 12 different feeds used, ragi powder suspension showed the best growth and cyst production. A wide range of salinity from 90 to 210‰ was used to study its effect on cyst production. Among all these treatments, 130‰ salinity was found suitable for induction of oviparity and cyst production in the San Francisco Bay strain of Artemia sp.

Note on the occurrence of Artemia in Sri Lanka

A brief account is given of the Artemia populations occurring in Sri Lanka with respect to inland and brackishwater aquaculture activities.

Pond production of Artemia in Pakistan

Currently our government and the private sectors are very much interested in the establishment of marine aquaculture. For the successful operation in aquaculture of finfishes and shellfishes, the basic requirement is the suitable diet, apart from proper environment. For the larvae, juveniles and adult stages of the culturing organisms the live Artemia is the ideal food. The aquaculturists the worldover are using live food for their culturing organisms, as the live food played an important role in the dietary management of aquaculture of finfishes and shellfishes (Sorgeloos and Kulasekarapandian, 1984), particularly during larval stages. The live nauplii of Artemia are used in aquaculture of finfishes and shellfishes due to being nutritionally balanced, non polluting, economically bearable, viable and readily acceptable to the culturing species. The adult Artemia is also used for feeding the aquarium fishes particularly so when there is a clear abundance of this resource which is cheaper and can economically compete with alternative artificial diet. By the use of Artemia the aquaculturists may obtain optimum growth and survival rate of the organisms. The life cycle of Artemia is very short, which is completed within two weeks especially during dry season in highly saline waters, the two weeks old Artemia starts producing cysts. These cysts become ready to harvest within a week.