A review of toxicity testing protocols and endpoints with Artemia spp.

Artemia spp. is an historically popular biological model still requiring an official internationally based standardization. Several endpoints are currently available. Short-term acute endpoints include biomarker (acetylcholinesterase; heat stress proteins; lipid peroxidation; thiobarbituric acid reactive substances; thioredoxin reductase; glutathione-peroxidase; glutathione S-transferase; glutathione reductase; aldehyde dehydrogenase; and adenylpyrophosphatase and Fluotox), hatching (dry biomass, morphological disorders and size), behavioral (swimming speed and path length), teratogenicity (growth), and immobilization (meaning mortality after 5–30 s observation). Long-term chronic tests focus on growth, reproduction and survival or mortality after 7–28 d exposure from larval to adulthood stage. We analyzed each test looking at its endpoint, toxicant and experimental design including replicates, exposure time, number of exposed cysts or organisms and their relative life stage, exposure conditions during hatching and testing (salinity, pH, light intensity, aeration dilution media, and food supply), type of testing chambers, and quality assurance and quality control criteria. Similarities and differences between the identified approaches were highlighted. Results evidenced that hatching 24 h short-term and 14 d long-term mortality are the most promising Artemia spp. protocols that should go forward with international standardization.     

Effect of brood size and hatching sequence on prefledging mortality of Sandwich terns: why lay two eggs?

The mortality of Sandwich tern Sterna sandvicensis chicks held in enclosures was studied in colonies on Griend, in the Dutch Wadden Sea, from 1992 to 1999, and on Hirsholm, in the Danish Kattegat, in 1997. Survival of chicks until fledging was 73% for chicks hatching from first-laid eggs or single-egg clutches and 59–64% for partially hatched two-egg clutches, whereas 6% of second hatchlings survived until fledging. Less than 2% of all two-chick broods actually fledged two chicks. Because 18% of the two-egg clutches only hatched one egg, 7% of fledglings of two-egg clutches originated from a second-laid egg. In nests where both eggs hatched, the number of chicks was usually reduced soon after hatching. Within five days of hatching more than 50% of the second hatchlings died of starvation or were preyed upon. It seems that overproduction commonly occurs in Sandwich terns and that investment in a surplus egg mainly serves as an insurance mechanism. On Griend and Hirsholm, chick productivity of two-egg clutches was somewhat higher than for one-egg clutches. Undernourishment was an important cause of death, either directly by starvation or by selective predation of chicks in poor condition. This, in combination with earlier, studies suggests that Sandwich tern parents on Griend are exposed to severe food stress.     

Optimal hatching conditions of Zophobas atratus (Coleoptera: Tenebrionidae) eggs under various culture conditions

Supermealworm, Zophobas atratus (Coleoptera: Tenebrionidae) is a tropical species used for human consumption and as an ingredient in animal feed. We investigated the hatching characteristics of Z. atratus eggs under varying conditions to optimize artificial rearing conditions. Overall, the average egg weight, width, and length decreased by 42.3%, 3.7%, and 10.1%, respectively, on the 6th day post-oviposition. We varied either temperature, photoperiod, illuminance, or humidity, while the other conditions remained constant. Of the four experimental temperatures—25 °C, 27 °C, 30 °C, and 33 °C–30 °C was associated with the highest hatching rate (91.0%) during days 5–7. Although other experimental conditions did not have a significant effect on hatchability, we were able to elucidate the ideal conditions for improved hatchability. For photoperiod, hatchability was the highest (84%) at 12L/12D, followed by 81.0% at 15L/9D, and 70% at 9L/15D. Among the illuminance conditions tested—600, 1800, and 3000 lx—hatchability was the highest at 1800 lx (82.3% on days 4–6). Among the relative humidity (RH) levels tested—50%, 65%, and 80%—hatchability was the highest at 65% RH (88.5% on days 5–6). Thus, the ideal conditions for maximum hatchability were 30 °C, 12L/12D, 65% RH, and 1800 lx. Under these combined conditions, the hatching rate increased by 93.6%, and the hatching time was reduced by two days.     

1-苯基-2-硫脲对斑马鱼胚胎发育与黑色素生成的影响

目的1-苯基-2-硫脲（PTU）可抑制斑马鱼胚胎黑色素的产生,保持斑马鱼透明,便于形态观察和信号检测。本文研究了PTU对斑马鱼胚胎发育的影响和抑制斑马鱼胚胎黑色素生成,保持斑马鱼透明性的最佳浓度。方法用不同浓度PTU处理23hpf（受精后,hourspostfertilization,hpf）斑马鱼胚胎,作用57h后观察80hpf斑马鱼的形态学、生理学改变,计算死亡率和孵化率,测量心率和静脉窦-动脉球之间的距离。结果浓度为0．197mmo]／L、0．296mmol／LPTU可以有效抑制黑色素生成,保持斑马鱼整体透明,对斑马鱼心血管系统结构和生理功能无影响,且不影响斑马鱼正常孵化过程。随着PTU浓度的增加,斑马鱼死亡率增加,孵化率下降,出现心包水肿,心脏畸形等改变,心率下降,静脉窦-动脉球之间的距离增大。结论浓度不高于0．296mmol／L的PTU溶液能有效抑制斑马鱼黑色素生成,对斑马鱼心血管毒性研究无影响。     

Improved Methods of Hatching Heterodera schachtii Larvae for Screening Chemicals

The rate of hatching of Heterodera schachtii larvae was greatly increased by placing cysts in sieves enclosed by small disposable cups. An apparatus that permitted rapid storage of second-stage larvae at 10 C prolonged the viability of the larvae.     

Development of cortical vesicles in Sicyonia ingentis ova: their heterogeneity and role in elaboration of the hatching envelope

In the marine shrimp Sicyonia ingentis, ova lack cortical vesicles at spawning. Previous ultrastructural studies suggested that two different populations of cortical vesicles (dense vesicles and the ring vesicles) appear within 30 min post-spawning. These vesicles undergo sequential exocytosis (exocytosis of the dense vesicles followed by exocytosis of the ring vesicles) that leads to the formation of a hatching envelope around the ovum (see Pillai and Clark: Tissue & Cell 20:941-52, 1988). In the present study, lectins were used as molecular probes to study the development of cortical vesicles subsequent to spawning and the role of these vesicles in formation and elaboration of the hatching envelope. Isolated envelopes were screened with 11 different lectins to determine what group(s) were specific to the envelope glycoconjugates; Concanavalin A (Con A), Griffonia simplicifolia (GS II), Lens culinaris (LCA), and wheat germ agglutinin (WGA) bound to the envelopes. FITC-lectin studies of sectioned ova (fixed at various time points after spawning) utilizing WGA and LCA showed different labelling patterns. Data obtained at the light microscopical level indicated that WGA was specific to the dense vesicles and the outer portion of the envelope, while LCA exhibited specificity for the ring vesicles and the inner portion of the envelope. At the ultrastructural level, gold-LCA labelling was seen associated with the cisternal elements (containing ring-shaped structures), ring vesicles, and the inner layer of the fully formed envelope. These data demonstrated that 1) the ring vesicles are formed by fusion of cisternal elements containing ring-shaped structures; 2) the two species of cortical vesicles are chemically heterogeneous; and 3) the components of each type of vesicle contribute to different integral parts (the outer and inner layers) of the hatching envelope.     

Breeding antarctic krill in captivity

Antarctic krill were maintained in large aquaria at Port of Nagoya Aquarium, Japan, under controlled photoperiod and were fed on phytoplankton and enriched animal feed. Maturation and spawning were induced after the light : dark (L : D) cycle was increased from 8 : 16 or 12 : 12 to 24 : 0, or when the L : D cycle was held constant at 14 : 10. This study is one of the first studies that demonstrate initiation of maturation and spawning events of krill under controlled photoperiod. Out of three experimental batches of krill, a total of 28 spawning events were observed. The mean number of eggs per event was 1424 with a range between 139 and 3458. The mean hatching success per batch was 19.1%. The relation between photoperiod and maturity/spawning is discussed. Furthermore, hatching is compared to previous studies and the reason for the low success is discussed.     

Do temperature fluctuations during incubation always play an important role in shaping the phenotype of hatchling reptiles?

Fluctuating temperatures (FTs) influence hatchling phenotypes differently from constant temperatures (CTs) in some reptiles, but not in others. This inconsistency raises a question of whether thermal fluctuations during incubation always play an important role in shaping the phenotype of hatchlings. To answer this question, we incubated eggs of Naja atra under one CT (28 °C, CT), two temperature-shift [cold first (CF) and hot first (HF) in which eggs were first incubated at 24 or 32 °C and then at the other, each for 20 days, and finally at 28 °C until hatching], and one FT thermal regimes. Female hatchlings were larger in snout–vent length but smaller in tail length, head size than male hatchlings from the same-sized egg; female hatchlings had more ventral scales than did male hatchlings. The FT and HF treatments resulted in shorter incubation lengths. Tail length was greatest in the CT treatment and smallest in the FT treatment, with the CF and HF treatments in between; head width was greater in the CT treatment than in the other three treatments. Other examined hatchling traits did not differ among treatments. The observed morphological modifications cannot be attributed to the effect of thermal fluctuations but to the effect of temperatures close to the upper and lower viable limits for the species. Our results therefore support the hypothesis that hatchling phenotype is not altered by thermal fluctuation in species with no phenotypic response to incubation temperature within some thresholds.     

文县疣螈繁殖初探

2006年4～9月、2007年4～9月,在四川青川初步观察了文县疣螈(Tylototriton wenxianensis)的繁殖习性,并研究了其主要特点,包括繁殖期雌雄差异、繁殖迁徙、交配与产卵、幼体发育等,同时在甘肃文县观察了其幼体的生长发育过程.结果表明,文县疣螈的繁殖期一般为4月上旬至9月上旬,繁殖期成体性比不断变化.该螈具有繁殖迁徙现象,迁徙及求偶行为由雄性占主动.求偶、交配及产卵均在水塘周围岸边的陆地上进行,产卵集中在5月初至7月末,雌螈平均产卵数为42.8枚(n=6),卵的平均孵化期为26.8 d(n=119).卵群的平均孵化率为46.3%(n=6).随着气温的升高、相对湿度和降水量的增加,雌螈产卵量和孵化率升高,卵的平均孵化期缩短.