Effects of cadmium chloride on the development of rainbow trout Oncorhynchus mykiss early life stages

The sub-chronic (28–56 days) effects of exposure to low concentrations of cadmium (Cd; 0·05, 0·25, 0·50 and 2·50 μg l⁻¹) shortly following fertilization on embryos, larvae and juvenile rainbow trout Oncorhynchus mykiss were examined. Premature hatching occurred at lower concentrations (0·05 and 0·25 μg l⁻¹ Cd), however, delayed hatching was seen in the 2·50 μg l⁻¹ Cd group, with >90% of hatching occurring on the last day of the hatching period. Larval growth was negatively affected by Cd exposure in a concentration-dependent manner. Larvae exposed to 2·50 μg l⁻¹ Cd were 13·9 ± 0·8% shorter in total length ( L _T) and weighed 22·4 ± 3·5% (mean ± s . e .) less than controls at the end of the exposure period. Plasma sex steroid concentrations (oestradiol in juvenile females and 11-ketotestosterone in juvenile males) were elevated (four- to 10-fold over controls) in exposed fish in both males and females, following 28 days of exposure to 0·05, 0·25 and 0·50 μg l⁻¹ Cd, respectively. These results suggest that environmentally realistic concentrations (in the μg l⁻¹ range) of Cd can affect the development of O. mykiss impacting embryos, larvae and juvenile fish.     

Assessment of water and sediment contamination in small streams by means of cytological and biochemical alterations in isolated rainbow trout (Oncorhynchus mykiss) hepatocytes

Isolated hepatocytes of rainbow trout (Oncorhynchus mykiss) were exposed to (1)native waters and acetone sediment extracts ofthe Krähenbach and the Körsch, twosmall streams in southwest Germanycharacterized by different levels of chemicalcontamination, as well as (2) differentconcentrations (1 ×, 10 ×, 50 ×) of anartificial mixture of 20 contaminants (PAHs,PCBs, pesticides) typical of the chemicalburden of the more heavily contaminatedKörsch. Following exposure for up to 3 d,hepatocytes were analyzed by means of electronmicroscopy and enzyme biochemistry to evaluatesublethal cytotoxic effects. Results documentthat dilutions of Krähenbach and Körschsurface water and sediment extracts inducemultiple time- and dose-dependent structuraland functional changes in hepatocytes. Betweenboth rivers as well as between free water phaseand sediment, significant differences in thetoxic burden could be detected. Consistently,however, both morphological and biochemicalresponses induced by samples from the moreseverely polluted river Körsch were morepronounced than those of the moderatelycontaminated river Krähenbach, whichindicates that the differential pollution ofthe two rivers directly translates into theextent of biochemical and morphological changesin hepatocytes. Cytopathological effectsincluded deformation of nuclear envelopes andincrease of heterochromatin, heterogeneity ofmitochondria, vesiculation of cisternae of therough endoplasmic reticulum, as well asproliferation of the smooth endoplasmicreticulum. Peroxisomal and lysosomalproliferation could be correlated tostimulation of the corresponding markerenzymes, catalase and acid phosphatase. In bothrivers, activities of lactate dehydrogenase andalanine aminotransferase as markers ofcytosolic glycolysis and protein metabolismwere increased indicating a general stimulationof cellular metabolism. Similar changes wereinduced by exposure to the artificial chemicalmixture resembling the chemical contaminationof the Körsch. Cytological changes inisolated hepatocytes thus not only serve todiscriminate between different levels ofcontamination of water, but may also help todifferentiate between pollution levels ofsediments.     

PFOS对斑马鱼胚胎及仔鱼的生态毒理效应

全氟辛烷磺酸（Perfluorooctane sulfonate, PFOS）是一种广泛存在于水生生态系统的新型持久性有机污染物（Persistent Organic Pollutants, POPs）,其对鱼类健康的影响以及水生生态系统安全的潜在威胁是当前人们高度关注的水环境问题。为探究PFOS对斑马鱼（Danio rerio）胚胎及仔鱼的生态毒理效应,本文研究了不同浓度（0,0.1,1,10 mg/L）PFOS暴露对斑马鱼胚胎孵化率、仔鱼畸形率与死亡率、仔鱼心率、仔鱼运动行为以及生长的影响。结果表明：PFOS暴露对斑马鱼胚胎孵化率、孵出仔鱼死亡率与畸形率的影响显著（P﹤0.05）,10 mg/L PFOS暴露导致胚胎孵化率下降,孵化延迟,仔鱼死亡率与畸形率升高;PFOS暴露4 dpf（day post-fertilization,dpf）或8 dpf 对斑马鱼仔鱼心率影响显著（P﹤0.05）,心率随PFOS暴露浓度升高而增加;PFOS 暴露6 dpf 或9 dpf 对斑马鱼仔鱼的运动行为影响显著（P﹤0.05）,10 mg/L PFOS暴露6 dpf 导致运动斑马鱼仔鱼比例和仔鱼最大持续运动距离增加（P﹤0.05）,PFOS暴露9 dpf,单位时间内仔鱼的运动距离、停顿频率、平均每次运动距离随PFOS暴露浓度升高而减少（P﹤0.05）,最大持续运动距离随PFOS暴露浓度升高而增加（P﹤0.05）,呈剂量依赖的毒理学效应;PFOS暴露导致斑马鱼仔鱼体长和吻宽下降（P﹤0.05）或有下降的趋势,并对吻宽/体长、吻宽/头长影响显著（P﹤0.05）。以上研究结果提示：PFOS对斑马鱼胚胎及仔鱼具有显著的发育与行为毒性,仔鱼心率、运动行为、吻宽/体长以及吻宽/头长等是评估水体PFOS污染敏感而有效的生物标志物。     

Differences in growth rates among cohorts of Encrasicholina punctifer and Engraulis japonicus larvae in the coastal waters off Tanshui River Estuary, Taiwan, as indicated by otolith microstructure analysis

The hatching dates of Encrasicholina punctifer and Engraulis japonicus larvae collected in the coastal waters off Tanshui River Estuary during the fishing seasons of 1992 and 1993 indicated that these two anchovies had protracted spawning seasons, which resulted in multiple recruitment cohorts. Encrasicholina punctifer larvae recruited to the estuary from October to March, while the majority of E. japonicus larvae came in March-May and to a lesser extent in October and November. The E. punctifer larvae on arrival to the estuary were 17·4–35·6 mm in length, 167ndash;89 days old and had growth rates of 0·4–1·0 mm day⁻¹, E. japonicus larvae were 12·1–32·7 mm in length, 19–62 days old and had growth rates of 0·7–0·9 mm day⁻¹. Growth rates were significantly different among cohorts and positively correlated to water temperature.     

Effect of delayed first feeding on development and feeding ability of Paralabrax maculatofasciatus larvae

The impact of delayed feeding early in development on late feeding ability and development of spotted sand bass Paralabrax maculatofasciatus larvae was examined. Larvae were sampled from hatching until day 19 after delayed feeding for zero (control), 1, 2, or 3 days. Feeding incidence was evaluated as the percentage of larvae with food in the gut and feeding intensity was measured by direct counting of prey after dissection of the gut. Delayed feeding effects due to starvation were observed early in development. By day 5, significant differences ( P  < 0·05) were observed between controls and larvae submitted to degrees of delayed feeding, including total length ( L _T), eye diameter, and the size of the head, liver, gut, muscles and body. Differences were still apparent in L _T, and body, liver and muscle heights at the end of the studied period. Larvae under total starvation did not survive beyond day 5. Initial feeding incidence was 35, 60, 90 and 10% for larvae first fed on day 2, 3, 4 or 5, respectively. Mean ±  s . d . feeding intensity was 3·6 ± 0·8, 2·8 ± 1·3, 5·2 ± 0·3, and 10·2 ± 1·5 rotifers per larva depending on whether larvae were initially fed on day 2, 3, 4 or 5, respectively. The point of no return occurred between 4 and 5 days after hatching.     

DNA damage induced by genotoxicants in zebrafish (Danio rerio) embryos after contact exposure to freeze-dried sediment and sediment extracts from Laguna Lake (The Philippines) as measured by the comet assay

Laguna Lake, Philippines, is utilised as a resource for drinking and irrigation water and a major source of animal protein for more than 11 million residents. However, it also serves as a huge sink for anthropogenic pollutants from a variety of direct or indirect discharges. Given their well-known properties to accumulate organic pollutants, Lake Laguna sediments have been investigated by use of a newly developed sediment-contact assay to determine the bioavailable fraction of the total genotoxic hazard potential. Freshly fertilized zebrafish eggs (2 h after fertilization) were allowed to develop while exposed to different concentrations of freeze-dried sediments (exposure to bioavailable compounds) as well as organic extracts (complete genotoxic hazard potential including bioaccumulation) prepared from sediments collected at five sites within the lake. Following a 96 h exposure, single-cell suspensions were prepared from macerated larvae and analyzed for DNA strand-breaks by use of the comet assay. Genotoxicity could be identified in both solid-phase exposure scenarios as well as after exposure to organic sediment extracts. Only the South Bay (SB) site did not show a significant genotoxic burden relative to controls in tests of both freeze-dried sediment and organic extracts. In contrast, the Northwest Bay site displayed pronounced genotoxicity to the zebrafish embryos in both exposure scenarios. Perylene and copper in the sediments could be identified as potential causes of the genotoxic response. To compare the results (maximum induction coefficients) of zebrafish embryos with an established comet protocol, rainbow trout liver cells (RTL-W1) were exposed to the same extracts. The findings correlated well (Spearman correlation r=0.90), proving a good reliability of the results from zebrafish primary cells. In conclusion, the present study demonstrates that the bioavailable fraction of the genotoxic pollutants may pose a threat for both benthic organisms and human health via drinking-water and fish consumption.     

The ontogeny of haematopoiesis in the marine teleost Leiostomus xanthurus and a comparison of the site of initial haematopoiesis with Opsanus tau

The ontogeny of haematopoiesis in the perciform fish, spot Leiostomus xanthurus , differed from that reported as the norm for fishes, as exemplified by the cypriniform zebrafish Danio rerio , and observed in the batrachoidiform oyster toadfish Opsanus tau . Erythropoiesis in spot was first evident in the head kidney of yolk‐sac larvae 3 days after hatching (DAH). No embryonic intermediate cell mass (ICM) of primitive stem cells or blood islands on the yolk were apparent within embryos. Erythrocytes were first evident in circulation near the completion of yolk absorption, c . 5 DAH, when larvae were c . 2·0 mm notochord length ( L _N). Erythrocyte abundance increased rapidly with larval development for c . 14 to 16 DAH, then became highly variable following changes in cardiac chamber morphology and volume. Erythrocytic haemoglobin (Hb) was not detected within whole larvae until they were 12 DAH or c . 3·1 mm L _N, well after yolk and oil‐globule absorption. The Hb was not quantified until larvae were >47 DAH or >7 mm standard length. The delayed appearance of erythrocytes and Hb in spot was similar to that reported for other marine fishes with small embryos and larvae. In oyster toadfish, a marine teleost that exhibits large embryos and larvae, the ICM and Hb were first evident in two bilateral slips of erythropoietic tissue in the embryos, c . 5 days after fertilization. Soon thereafter, erythrocytes were evident in the heart, and peripheral and vitelline circulation. Initial haematopoiesis in oyster toadfish conformed with that described for zebrafish. While the genes that code for the development of haematopoiesis are conserved among vertebrates, gene expression lacks phylogenetic pattern among fishes and appears to conform more closely with phenotypic expression related to physiological and ecological influences of overall body size and environmental oxygen availability.     

Growth of young-of-the-year mackerel in the Bay of Biscay

The first growth season of young-of-the-year (0+ year) mackerel Scomber scombrus , sampled in the Bay of Biscay, was parameterized to determine growth patterns. Daily increments were identified on sagittae otoliths, for calculation of age and growth of 92 larvae and 54 juveniles over the range 3·6–215·0 mm standard length ( L _S). A Gompertz curve was fitted to the length-at-age data. At the end of the first year of growth L _S was 194·2 mm, with a maximum growth increment of c . 2 mm day⁻¹, observed 62 days after hatching. Backcalculated growth increments for mackerel juveniles, during their larval stage, were higher than those observed for sampled larvae; only 10·9% of sampled larvae were estimated to survive. Growth for north-eastern Atlantic mackerel was slower than that published for north-western Atlantic mackerel. Backcalculated hatching dates for mackerel were consistent with the typical temporal distribution of mackerel spawning in the Bay of Biscay.     

The importance of measuring biotic and abiotic factors in the lower egg pocket to predict coho salmon egg survival

Based on results from simulated redds of coho salmon Oncorhynchus kisutch , the amount of fine sediment <0·5 mm in the lower half of the egg pocket, rather than the entire egg pocket of the redd, was a strong predictor of egg survival to hatching (  r ²  = 0·62). The relationship was much stronger than observed in other studies, which typically ignore egg pocket structure. Abundance of a fish egg-eating worm, Haplotaxis ichthyophagous , an oligochaete that may have been attracted to fine sediment and dead eggs in the egg pocket, was also associated with a decrease in egg survival. The worm, however, accounted for little of the variance in survival compared to fine sediment. Only 10% fine sediment (<0·5 mm) in the lower pocket was required to decrease survival from 100 to 5%. Other abiotic factors had weaker (gravel permeability) or non-existent (dissolved oxygen) correlations with survival.     

Lack of gas bladder inflation by the larvae of zebrafish in the absence of an air-water interface

Compared to a control gas bladder inflation rate of 95·1±1·9%, zebrafish Danio rerio larvae 72 h post-fertilization maintained in closed chambers had an inflation rate of just 19·1±7·7%. Larval survivorship through 10 days in closed chambers (32%) was significantly less than that in open chambers (76%), and the extent of spinal curvature was significantly higher among larvae maintained in closed chambers. Larvae which failed to inflate their gas bladder showed very little change in body length, and had a final dry weight c. 14% of that for control larvae. The small number of larvae with inflated gas bladders found in closed chambers might be attributed to the inadvertent introduction of small bubbles into two replicates. These results indicate that access to an air-water interface is critical for the normal development of zebrafish larvae, but also that a small, submerged, spherical gas volume may to a limited extent be used for initial gas bladder inflation.     

Characteristics of egg and larval production in captive bluespotted gobies

Spawning of the Hawaiian coral-reef goby Asterropteryx semipunctata was diurnal, occurring at various times throughout the day. Mean length of eggs deposited in nests was 0·76 mm (range 0·67–0·84); mean egg width was 0·47 mm (range 0·41–0·52). Clutch size varied from 296 to 1552 eggs (mean=886±309), and was independent of standard length, total body weight, and body condition. Mean relative clutch size was 1·59 eggs mg^-1 total body weight (range 0·84–2·43). Clutches hatched 4–5 nights after being deposited in a nest. Mean notochord length of newly-hatched larvae was 1·88 mm (range 1·60–2·04). The minimum period of time that elapsed between egg deposition and subsequent growth of a new batch of oocytes to spawning size was 5–6 days, providing a reasonable estimate of minimum spawning interval. Compared with other gobiids, tropical species tend to have shorter incubation periods, smaller eggs and smaller larvae at hatching.     

Drinking rate, uptake of bacteria and microalgae in turbot larvae

The drinking rate of turbot larvae increased from 14 to 120 nl larva^-1 h^-1 from day 2 to 11 after hatching, which gave a slightly increased specific drinking rate (calculated per biomass) from day 2 to 7 (0·8–1·9 nl μg carbon^-1 h^-1. The clearance rate of both algae and bacteria was 10–100 times higher than the drinking rate, which indicated that the larvae had an active uptake of both algae and bacteria. On day 2 and 4 after hatching highest clearance rate was observed for Tetraselmis sp. On day 6 about the same clearance rate was observed for bacteria, Isochrysis galbana and Tetraselmis sp. Until day 4 the turbot larvae had a higher ingestion rate of Tetraselmis sp. than of I. galbana , whereas on day 6 the rates were similar (28–41 ng carbon larvae^-1 h^-1). The assimilation efficiency was somewhat higher for I. galbana than for Tetraselmis sp., and on day 6 the assimilated algae constituted 1·5 and 0·9% of the larval biomass for I. galbana and Tetraselmis sp., respectively.     

Nerve Growth Factor Stimulates Cardiac Regeneration via Cardiomyocyte Proliferation in Experimental Heart Failure

Although the adult heart likely retains some regenerative capacity, heart failure (HF) typically remains a progressive disorder. We hypothesise that alterations in the local environment contribute to the failure of regeneration in HF. Previously we showed that nerve growth factor (NGF) is deficient in the failing heart and here we hypothesise that diminished NGF limits the cardiac regenerative response in HF. The capacity of NGF to augment cardiac regeneration was tested in a zebrafish model of HF. Cardiac injury with a HF phenotype was induced in zebrafish larvae at 72 hours post fertilization (hpf) by exposure to aristolochic acid (AA, 2.5 µM, 72–75 hpf). By 168 hpf, AA induced HF and death in 37.5% and 20.8% of larvae respectively (p<0.001). NGF mRNA expression was reduced by 42% (p<0.05). The addition of NGF (50 ng/ml) after exposure to AA reduced the incidence of HF by 50% (p<0.01) and death by 65% (p<0.01). Mechanistically, AA mediated HF was characterised by reduced cardiomyocyte proliferation as reflected by a 6.4 fold decrease in BrdU+ cardiomyocytes (p<0.01) together with features of apoptosis and loss of cardiomyocytes. Following AA exposure, NGF increased the abundance of BrdU+ cardiomyocytes in the heart by 4.8 fold (p<0.05), and this was accompanied by a concomitant significant increase in cardiomyocyte numbers. The proliferative effect of NGF on cardiomyocytes was not associated with an anti-apoptotic effect. Taken together the study suggests that NGF stimulates a regenerative response in the failing zebrafish heart, mediated by stimulation of cardiomyocyte proliferation.     

Comparison of the Long-Term Risks of Removal and In Situ Management of Contaminated Sediments in the Fox River

The long-term potential risks of environmental dredging vs. in situ contaminated sediment management practices are discussed and compared for the Lower Fox River, Wisconsin. The risks are identified as being largely associated with the residual sediment contamination associated with either approach. The integral of the surface area-weighted average contaminant concentration in surface sediment is proposed as a metric to compare these risks. Capping is shown to exhibit significantly reduced exposure and risk relative to the dredging scenarios, even if potential undetected erosion of 5% of the cap is considered. Even with the improbable event of undetected failure of 25% of the cap, the exposure and risk associated with capping is approximately equal to or below all dredging scenarios. A preference for dredging due to the perception that it eliminates the long-term risk of in situ capping is not supported by this analysis. Although strictly applicable only to the Lower Fox River, the results suggest sitespecific analyses must be conducted to determine which sediment management approaches minimize the potential for long-term exposure and risk.     

Development of bile salt-dependent lipase in larval turbot

Pancreatic bile salt-dependent lipase (BSDL) was present with 0·5 μg BSDL larva⁻¹ from hatching in turbot larvae. The enzyme content increased during the yolk sac phase to 1·1 μg BSDL larva⁻¹. This suggests that larval turbot are able to digest lipids from the start of exogenous feeding. The BSDL synthesis was stimulated first by food about 5 days after the onset of first feeding. The content per larva increased exponentially in fed larvae to 20 μg BSDL larva⁻¹ on day 23 after hatching and decreased in starved larvae. In contrast, the specific content decreased during the first feeding phase, meaning that smaller larvae had a higher content of enzyme related to their biomass than did bigger larvae.     

Maternal transfer of cadmium tolerance in larval Oreochromis mossambicus

Exposing female tilapia to Cd did not affect reproduction and egg quality, nor the survival of their offspring. The LT ₅₀ of Cd-exposed larvae (within 24 h post hatching) from Cd-treated female Oreochromis mossambicus (mean ±S.E.=10·9±0·5 days, n =4) was greater than that of Cd-exposed larvae from control females (6·9± 1·2 days, n =4). A substantial amount of metallothionein mRNA (MT mRNA) was found in both oocytes and newly hatched larvae from Cd-treated females, but MT mRNA levels were low in newly fertilized eggs. The amount of MT mRNA was very low in all these stages in the control group. Cadmium could not be detected at any stage for either control or Cd-treated group, and metallothionein protein levels were also very low in both groups. The early appearance of MT mRNA in oocytes and newly hatched larvae from Cd-treated females may result in early translation after hatching and explain their tolerance to Cd.     

Oxygen requirements of zebrafish (Danio rerio) embryos in embryo toxicity tests with environmental samples

The zebrafish embryo test is a widely used bioassay for the testing of chemicals, effluents and other types of environmental samples. Oxygen depletion in the testing of sediments and effluents is especially important and may be a confounding factor in the interpretation of apparent toxicity. In order to identify oxygen levels critical to early developmental stages of zebrafish, oxygen consumption of zebrafish embryos between 0 and 96h post-fertilization, minimum oxygen levels required by the embryos for survival as well as the effects of oxygen depletion following exposure to model sediments were determined. No significant effects on zebrafish embryo development were observed for oxygen concentrations between 7.15 and 3.33mg/L, whereas at concentrations between 3.0and 2.0mg/L minor developmental retardations were observed, yet without any pathological consequences. Oxygen concentrations lower than 0.88mg/L were 100% lethal. In the sediment contact tests with zebrafish embryos, native sediments rich in organic materials rapidly developed strongly hypoxic conditions, particularly at the sediment-water interface (0 to 500μm distance to the sediment).