Effect of abiotic factors (Temperature,pH, Heavy Metals) on activity of carbohydrases of food objects of ichthyophages

Separate and complex effect of temperature, pH, and heavy metals (copper, zinc) is investigated on activity of carbohydrases in the whole organism of juvenile fish (carp Cyprinus carpio, golden carp Carassius auratus, roach Rutilus rutilus, perch Perca fluviatilis, common kilka Clupeonella cultriventris, and rotan Perccottus glenii)—potential food objects of typical and facultative ichthyophages. Temperature of 0°C and low pH at separate and combined action noticeably decrease the amylolytic activity in comparison with metals. Enzymes of golden carp, kilka, and rotan are maximally stable to change of temperature and pH. As a rule, the combined effect of low temperature and metals, as well as of low pH and metals, increases the level of separate effect. Maximum decrease of activity of carbohydrases (by two-seven times) is noted under the complex action of temperature 0°C, pH 5.0, and heavy metals (especially copper).     

Activity of Peptidases and Glycosidases: Effect of Light Deprivation and Heavy Metals on the Effects of Serotonin and Cholecystokinin in the Carp <Emphasis Type="Italic">Cyprinus carpio</Emphasis>

The dependence of the effects of serotonin and cholecystokinin on the activity of peptidases and glycosidases in the intestine of the carp Cyprinus carpio from the content of zinc and copper ions in different photoperiods (6: 18 and 0: 24 h) was studied. It was shown that the activity of enzymes in the intestine of fish under the influence of both hormones in conditions of light deprivation significantly increases, especially under the action of copper. The mechanisms of the influence of these factors on the activity of peptidases and glycosidases of fish intestines are discussed.     

Influence of the water heating rate upon thermal tolerance in common carp (Cyprinus carpio L.) during different seasons

For the first time, the dependence of the upper lethal temperature (ULT) level upon the water heating rate was determined in young common carp (Cyprinus carpio) during different seasons and within the wide range of heating rates. The highest ULT values were revealed by using the chronic lethal impact method, with a water heating rate of 1°C/day (or 0.04°C/h). At such a rate, fish can successfully acclimate to a temperature increase. The ULT values determined by that method characterize the fish maximal thermal tolerance. During different seasons these values were differing from each other within a 3°C range. The data on the common carp ULT, along with data on crucian carp (Carassius auratus), evidences that this species belongs to the group of the most thermophilic fish, not only among cyprinids, but also compared to the representatives of other families. Obtained experimental data characterizing the upper thermal limits for fish vital functions may be used for the development of ecological safety criteria near the zones of discharge of heated waters from heating and nuclear power plants, as well as for the forecasting of the potential consequences of global warming.     

The effect of temperature,pH, and heavy metals (copper,zinc) upon proteinase activities in the potential preys for typical and facultative piscivores

Separate and combined action of temperature, pH, and heavy metals (copper and zinc) upon whole body activities of casein-and hemoglobinlytic proteinases were studied in perch (Perca fluviatilis), ruff (Gymnocephalus cernuus), roach (Rutilus rutilus), kilka (Clupeonella cultriventris), and common carp (Cyprinus carpio). Also, these activities were determined in cyprinids’ underyearlings (in sum) dominating the food of typical and facultative piscivores. It was revealed that, following a decrease in temperature to 0°C and in the presence of heavy metals (copper and zinc) at concentrations of 10 mg/l, casein-and hemoglobinlytic proteinase activities decreased statistically significantly in all studied fish. Both decrease and increase in pH values lead to less pronounced changes in the enzymatic activities varied in different species. As a rule, decrease in pH values resulted in considerable increase of the hemoglobinlytic activity, while pH increase caused inconsiderable enzyme activation. Combined action of natural factors and, in some cases, combined action of low temperature and metals, changed the caseinlytic activity to a greater extent than at their separate actions. Magnitude of multifactor effects in most cases is lower than of bifactor effects of temperature and metals.     

Biohydrometallurgical technology of copper recovery from a complex copper concentrate

Leaching of sulfide-oxidized copper concentrate of the Udokan deposit ore with a copper content of 37.4% was studied. In the course of treatment in a sulfuric acid solution with pH 1.2, a copper leaching rate was 6.9 g/kg h for 22 hours, which allowed extraction of 40.6% of copper. At subsequent chemical leaching at 80°C during 7 hours with a solution of ferric sulfate obtained after biooxidation by an association of micro-organisms, the rate of copper recovery was 52.7 g/kg h. The total copper recovery was 94.5% (over 29 hours). Regeneration of the Fe³⁺ ions was carried out by an association of moderately thermophilic microorganisms, including bacteria of genus Sulfobacillus and archaea Ferroplasma acidiphilum, at 1.0 g/L h at 40°C in the presence of 3% solids obtained by chemical leaching of copper concentrate. A flowsheet scheme of a complex copper concentrate process with the use of bacterial-chemical leaching is proposed.     

Critical thermal maximum of Amur sleeper <Emphasis Type="Italic">Perccottus glenii</Emphasis> in different seasons of the year

Seasonal differences in the critical thermal maximum are revealed in Amur sleeper Perccottus glenii at the age of 1+ and 2 years. Maximum values of the critical thermal maximum determined by two criteria (rotation of the body and immobile gill covers) are registered in the summer, and minimum values of this parameter are observed in the winter. Maximum thermal resistance is registered at a low water heating rate (0.2–0.4°C/h), and minimum resistance is reported for a heating rate 5.0–10.5°C/h. The difference between the values of the critical thermal maximum assessed by two criteria reaches 1–2°C. Features of behavior of Amur sleeper are described at different heating rates. Based on the characteristics of thermal adaptation (critical thermal maximum and optimal temperature for growth), Amur sleeper is similar to warm water eurythermal species (goldfish Carassius auratus and carp Cyprinus carpio).     

Assessing the efficacy of corn-based bait containing antimycin-a to control common carp populations using laboratory and pond experiments

Strategic use of oral toxicants could allow for practical and sustainable control schemes for the invasive common carp (Cyprinus carpio, or ‘carp’) if a toxicant selectively targeted carp and not native species. In this study, we incorporated antimycin-a (ANT-A), a known fish toxicant, into a corn-based bait and conducted a series of experiments to determine its toxicity, leaching rate, and species-specificity. Our results showed that ANT-A was lethal to carp at doses ≥ 4 mg/kg and that the amount of ANT-A that leached out of the bait in 72 h was not lethal to carp or bluegill (Lepomis macrochirus). Species-specificity trials were conducted in 227 L tanks, in which carp were stocked with three native species representing families that occur sympatrically with carp in our study region: the fathead minnow (Pimephales promelas), yellow perch (Perca flavescens) and bluegill. These trials showed high mortality of carp (46%) and fathead minnows (76%) but no significant mortality of perch or bluegill. Finally, a pond study, which used the same species composition except for fathead minnows, resulted in 37% morality among adult carp and no mortality among perch or bluegill. Our results suggest that corn-based bait that contains ANT-A could be used to selectively control carp in ecosystems dominated by percids or centrarchids, such as lakes across the Great Plains ecoregion of North America, where carp are especially problematic.     

Thermally induced denaturation and aggregation of BLG-A: effect of the Cu<Superscript>2+</Superscript> and Zn<Superscript>2+</Superscript> metal ions

There is growing evidence that metal ions can accelerate the aggregation process of several proteins. This process, associated with several neuro-degenerative diseases, has been reported also for non-pathological proteins. In the present work, the effects of copper and zinc ions on the denaturation and aggregation processes of β-lactoglobulin A (BLG-A) are investigated by differential scanning calorimetry (DSC), fluorescence, electron paramagnetic resonance (EPR) and optical density. The DSC profiles reveal that the thermal behaviour of BLG-A is a complex process, strongly dependent on the protein concentration. For concentrations ≤0.13 mM, the thermogram shows an endothermic peak at 84.3°C, corresponding to denaturation; for concentrations >0.13 mM an exothermic peak also appears, above 90°C, related to the aggregation of the denaturated BLG-A molecules. The thioflavin T fluorescence indicates that the thermally induced aggregates show fibrillar features. The presence of either equimolar Cu²⁺ or Zn²⁺ ions in the protein solution has different effects. In particular, copper binds to the protein in the native state, as evidenced by EPR experiments, and destabilizes BLG-A by decreasing the denaturation temperature by about 10°C, whereas zinc ions probably perturb the partially denaturated state of the protein. The kinetics of BLG-A aggregation shows that both metal ions abolish the lag phase before the aggregation starts. Moreover, the rate of the process is 4.6-fold higher in the presence of copper, whereas the effect of zinc is negligible. The increase of the aggregation rate, induced by copper, may be due to a site-specific binding of the metal ion on the protein.     

In vitro influence of zinc and magnesium on the deformability of red blood cells artificially hardened by heating

Trace elements have been shown to improve red blood cell (RBC) deformability: zinc in sickle cell disease and magnesium in an in vitro model of chemically rigidified erythrocytes. In this study, we investigated the effect and the influence of incubation time of zinc or magnesium on an in vitro model of rigidified RBCs by heating. Erythrocyte rigidity was determined by viscosimetry at high shear rate by a falling ball viscosimeter MT 90. In the first part of the study, six normal volunteers participated. Viscosimetry was performed on native blood before and after heating the sample for 10 min at 50°C. Therefore, increasing concentrations of zinc gluconate (final concentration: 0.5–4 g/L) or isotonic NaCl as control medium were added to the sample. Heating induced a twofold increase in all indices of RBC rigidity (p<0.05). At all these concentrations of zinc, a highly significant, dose-related fluidifying effect was observed (40–70%): this effect was immediately obtained and did not change over 60 min. Even at the highest concentration, recovery was not complete. In the second part of the study, we studied magnesium’s effects on blood. In a first protocol, whole blood was rigidified by heating at 56°C for 10 min, and the correcting effect of 5 min of incubation at 37°C of RBCs in 150 mmol/L NaCl, MgSO₄, magnesium acetate, and magnesium gluconate was investigated. In a second protocol, the same incubation with NaCl and magnesium salts was made on blood that had not been previously heated. In a third protocol, the correcting effect of magnesium gluconate on heated red blood cells was tested at four concentrations (75, 150, 225, and 300 mmol/L) over 1 h, for evaluating the effects of both concentration and time. Erythrocyte rigidity by heating is corrected by the three salts employed in protocol 1 (compared to sodium). In protocol 2, the deformability of normal (nonheated) red cells is not modified by magnesium. In protocol 3, no marked modification over 1 h is observed. The correcting effect is not complete for 75 mmol/L Mg, but remains the same at the three other concentrations. This study shows that zinc and magnesium at supraphysiological concentration are able to reverse RBC’s rigidification induced by heating, but that magnesium does not modify the flexibility of normal RBCs. This article suggests that zinc and magnesium may be studied in vivo as potential pharmacologic tools for improving hemorheologic disturbances.     

Characterization of two copper/zinc superoxide dismutases (Cu/Zn-SODs) from the desert beetle Microdera punctipennis and their activities in protecting E. coli cells against cold

Superoxide dismutases (SODs) are crucial in scavenging reactive oxygen species (ROS); however, studies regarding SOD functions in insects under cold conditions are rare. In this paper, two novel Cu/Zn-SOD genes in the desert beetle Microdera punctipennis, an extracellular copper/zinc SOD (MpecCu/Zn-SOD) and an intracellular copper/zinc SOD (MpicCu/Zn-SOD), were identified and characterized. The results of quantitative real-time PCR showed that MpecCu/Zn-SOD expression was significantly up-regulated by 4 °C exposure for 0.5 h, but MpicCu/Zn-SOD was not. Superoxide anion radical (O₂•^-) content in beetles under 4 °C exposure for 0.5 h showed an initial sharp increase and fluctuated during the cold treatment period, which was consistent with the relative mRNA level of MpecCu/Zn-SOD. The total SOD activity in the beetle was negatively correlated to the O₂•^- content with a correlation coefficient of −0.437. An E. coli system was employed to study the function of each MpCu/Zn-SOD gene. The fusion proteins Trx-His-MpCu/Zn-SODs were over expressed in E. coli BL21 using pET32a vector, and identified by SDS-PAGE and Western blotting. The transformed bacteria BL21(pET32a-MpecCu/Zn-SOD) and BL21(pET32a-MpicCu/Zn-SOD) showed increased cold tolerance to −4 °C as well as increased SOD activity compared to the control BL21(pET32a). The relative conductivity and malondialdehyde content in the two MpCu/Zn-SODs transformed bacteria under −4 °C were significantly lower than the control BL21(pET32a). Furthermore, BL21(pET32a-MpecCu/Zn-SOD) had significantly higher SOD activity and cold tolerance than BL21(pET32a-MpicCu/Zn-SOD) under −4 °C treatment, and had lower conductivity than BL21(pET32a-MpicCu/Zn-SOD). In conclusion, low temperature led to the accumulation of O₂•^- in M. punctipennis, which stimulated the expression of extracellular MpCu/Zn-SOD gene and the increase of total SOD activity. E. coli overexpressing Trx-His-MpCu/Zn-SODs increased resistance to cold treatment-induced oxidative stress. Our findings will be helpful in further study of Cu/Zn-SOD genes in insect cold-tolerance.     

Effects of abiotic factors (temperature,pH, heavy metals) on activities of glycosidases in invertebrate animals

Differences in amylolytical activity have been established in the whole body of some invertebrate animals (larvae of chironomids Chironomus plumosus, pond snail Limnaea stagnalis, orb snail Planorbis corneus, faucet snail Bithynia tentaculata, and zebra mussel Dreissena polymorpha-potential objects of nutrition of fish benthophages under separate and combined actions of temperature (0, 10 or 20°C), pH (5.0, 7.4 or 8.3), and ions of heavy metals (copper, zinc at a concentration of 10 mg/l) in vitro. The most pronounced inhibitory effect on the amylolytical activity in tissues of the studied animals was produced by a combination of the low temperature, acidic pH values, and the presence of the heavy metal ions. Enzymes of chironomids were more resistant to action of the studied agents as compared with molluscs.     

In vitro storage of unfertilized eggs of the Eurasian perch and its effect on egg viability rates and the occurrence of larval malformations

Ova ageing is the most important factor affecting fish egg quality after ovulation. Long-term storage of fish ova, using cryopreservation and vitrification techniques, has been unsuccessful to date. Instead, short-term in vitro ova storage has been used successfully and optimized in some cultured fish species. In vitro ova storage can drastically improve mass production of larvae and juveniles in the hatcheries by providing the possibility of the synchronous artificial fertilization for different females. To study how long unfertilized eggs of Eurasian perch (Perca fluviatilis L.) can retain their fertilizing ability after stripping, eggs were stored at temperatures of 4°C, 8°C and 12°C for 72 h post-stripping (HPS). The stored eggs of four female perch were separately fertilized at 0 h (i.e. control eggs fertilized before storage) and at 6-hour intervals during the experimental period of 72 h. The embryos reaching the eyed-egg and hatched-larvae stages, eyed-egg mortality and larval malformation rates were recorded as indices of egg quality. The results indicated that the maximum eyed eggs and hatched larvae (86% and 63%, respectively) were observed for eggs fertilized immediately after stripping, whereas the storage of the eggs at 4°C for 48 HPS decreased the eyed-egg and hatched-larvae rates to 46% and 17%, respectively. The use of a higher storage temperature resulted in a more rapid decrease in egg viability: eyed-egg and hatched-larvae rates of 23% and 9%, respectively, were obtained after 48 HPS storage at 8°C and 2% and 1% for eggs stored at 12°C. Eyed-egg mortality and larval malformation rates were not significantly affected by post-stripping ova ageing for at least up to 36 h. Thereafter, both values increased significantly and were measured to be the highest in the most aged ova. The present study demonstrated that stripped Eurasian perch eggs can be stored for at least 12 h at 4°C to 12°C without a significant reduction in their quality.     

Metabolic activity on Biolog FF MicroPlate suggests organic substrate decomposition by Aspergillus terreus NTOU4989 isolated from Kueishan Island Hydrothermal Vent Field,Taiwan

Aspergillus terreus NTOU4989 was isolated from sulfur sediment collected at Kueishan Island Hydrothermal Vent Field, Taiwan, and was shown to be able to grow at 45 °C and pH 3 in seawater, characteristics of hydrothermal vent sites. Fungi are one of the major decomposers in the marine environment, and to find out whether A. terreus NTOU4989 can fulfill such a role at the site, this study investigated its metabolic activity using the Biolog FF MicroPlate™ under different growth conditions (45 °C/pH 3, 45 °C/pH 7, 25 °C/pH 3, 25 °C/pH 7) with/without addition of Al³⁺, Fe²⁺ and Mn²⁺, major metal ions in the vent effluent. The greatest metabolic activity was observed at 25 °C/pH 3 with/without the metal ions based on substrate richness (R_S, metabolic richness), average well colour development (AWCD, metabolic capacity) and average well turbidity development (AWTD, mycelial production). Significant low R_S, AWCD and AWTD were observed at 45 °C, suggesting that A. terreus NTOU4989 is a thermotolerant fungus. The presence of the metal ions did not have a significant effect on its metabolic activity. Monomers of lignin, cellulose and hemicellulose, and common sugars and amino acids of algae were utilized in selected conditions, providing evidence of wood and algal decomposition by the fungus at the hydrothermal vent site.     

Heavy metal uptake capacities by the common freshwater green alga Cladophora fracta

Macroalgae have received much attention for heavy metal removal in treatment of domestic wastewater. In this report, the uptake capacity of a common freshwater green alga, Cladophora fracta, for heavy metal ions (copper, zinc, cadmium, and mercury) was evaluated. The equilibrium adsorption capacities were 2.388?mg Cu²⁺, 1.623?mg Zn²⁺, 0.240?mg Cd²⁺, and 0.228?mg Hg²⁺ per gram of living algae at 18°C and pH?5.0. The removal efficiency for Cu²⁺, Zn²⁺, Cd²⁺, and Hg²⁺ were 99, 85, 97, and 98%, respectively. Greater removal efficiency was achieved when the concentrations of metal ions were at very low level. The results indicated that living algae are suitable for removal and recovery of heavy metal ions from aqueous solutions and can be a potential tool to treat industrial wastewater.     

Effect of daylength and temperature on the larval diapause of the sunflower moth, Homoeosoma electellum

The effect of daylength and temperature on the regulation of the larval diapause of a central Missouri population of the sunflower moth, Homoeosoma electellum, was examined. Fully grown fourth-instar larvae exhibit a facultative diapause. Measurements of the effect of photoperiod on diapause induction revealed critical photoperiods of about 13 h 30 min light/day at 20°C, and between 11 h 45 min and 12 h light/day at 23°C. Third and fourth-instar larvae were shown to be the main sensitive stages for diapause determination. Daylength was also shown to be an important regulator of the rate of diapause development. A short day of LD 10:14 h permitted only a low rate of diapause development, whereas long days of LD 14:10 h and LD 16:8 h accelerated diapause development at 25 and 30°C. When long days were alternated with short days at 30°C the accelerating effect of long days on diapause development was not found. Systematic transfers of chilled diapausing larvae revealed an accelerated diapause development in groups transferred from 10 to 30°C LD 10:14 h, but diapause development was not accelerated in groups transferred from 10 to 30°C LD 16:8 h.     

Temperature effects on aerobic scope and cardiac performance of European perch (Perca fluviatilis)

Several recent studies have highlighted how impaired cardiac performance at high temperatures and in hypoxia may compromise the capacity for oxygen transport. Thus, at high temperatures impaired cardiac capacity is proposed to reduce oxygen transport to a degree that lowers aerobic scope and compromises thermal tolerance (the oxygen- and capacity-limited thermal tolerance (OCLTT) hypothesis). To investigate this hypothesis, we measured aerobic and cardiac performance of a eurythermal freshwater teleost, the European perch (Perca fluviatilis). Rates of oxygen consumption were measured during rest and activity at temperatures between 5 °C and 27 °C, and we evaluated cardiac function by in vivo measurements of heart rate and in vitro studies to determine contractility of myocardial strips. Aerobic scope increased progressively from 5 °C to 21 °C, after which it levelled off. Heart rate showed a similar response. We found little difference between resting and active heart rate at high temperature suggesting that increased cardiac scope during activity is primarily related to changes in stroke volume. To examine the effects of temperature on cardiac capacity, we measured isometric force development in electrically paced myocardial preparations during different combinations of temperature, pacing frequency, oxygenation and adrenergic stimulation. The force-frequency product increased markedly upon adrenergic stimulation at 21 and 27 °C (with higher effects at 21 °C) and the cardiac preparations were highly sensitive to hypoxia. These findings suggest that at (critically) high temperatures, cardiac output may diminish due to a decreased effect of adrenergic stimulation and that this effect may be further exacerbated if the heart becomes hypoxic. Hence cardiac limitations may contribute to the inability to increase aerobic scope at high temperatures in the European perch (Perca fluviatilis).     

Effects of acute and chronic testicular hyperthermia on levels of testosterone and corticosteroids in plasma of boars

Heat is known to depress spermatogenesis in the boar, but there is little quantitative evidence on its effects on testicular steroidogenesis in this species. The studies reported here examine the effects of short-term and chronic testicular hyperthermia on levels of testosterone (T) and corticosteroids (C) in plasma of Large-White (LW) boars.In examining effects of acute heating, three mature LW boars were maintained at 23°, 35° and 23°C ambient during three consecutive 24-h periods. Blood samples were collected hourly and levels of T and C in plasma determined. Prior to heating, plasma T levels varied diurnally (P<0.05) about a 24-h mean value of 2.78 nM. During heating at 35°C, and recovery at 23°C, mean plasma T levels remained unchanged (P>0.05) but there was a loss of diurnal rhythm. Mean 24-h plasma C levels did not change during heating (20.8 nMat 23°C, 20.2 nMat 35°; P>0.05), but fell (P<0.05) to 8.3 nM during the recovery period at 23°C.Effects of chronic heating on testis function were investigated by determining T and C concentrations in peripheral plasma of unilateral cryptorchid boars in which the scrotal testis was removed shortly after birth. Blood samples were drawn hourly, for 24 h, from each animal at about 10 months of age. The boars were then treated, i.v., with 700 IU hCG and blood samples collected frequently for 12 h. Mean plasma T levels before and after hCG treatment were 1.94 and 3.71 nM respectively, the difference between these levels being significant (P<0.05). At the same time, comparison was made with four normal littermates, hemicastrated at 3 days of age and heated to maintain testis temperature near 38°C. Mean plasma T levels in these boars increased (P<0.05) from 5.90 nM before, to 26.5 nM after hCG treatment, both levels being higher (P<0.05) than corresponding values for the hemicastrate cryptorchid animals. Levels of C in plasma increased (P<0.05) in the heated-scrotal boars following hCG treatment but decreased (P<0.05) in the cryptorchid animals. Histological comparison of testicular tissue from the scrotal and cryptorchid animals in this experiment revealed hypertrophy of Leydig cells in the abdominal testes.It is concluded that acute testicular hyperthermia (to c. 38°C) does not result in significant depression in mean plasma T levels of boars. However, chronic heating of testes at 38°C is associated with lower basal levels of T in peripheral plasma and an impaired response of plasma T concentrations following gonadotrophic stimulation.     

Effects of temperature change on cortisol release by common carp Cyprinus carpio

Common carp Cyprinus carpio, stressed by fish handling practices, responded with a decrease in cortisol secretion when temperature was lowered from 20 to 14° C within 3·5 h compared to those kept at 20° C.     

Effects of light on trace elments and triiodothyronine levels in plasma of mirror carp (Cyprinus carpio)

The changes of copper, zinc, iron, magnesium, manganese, and triiodothyronine as a result of short- and long-term isothermal exposure to light were determined in the plasma of mirror carp (Cyprinus carpio). Two groups of fish were subject to a 3-mo treatment that consisted of (1) 8h of light followed by 16 h of darkness and (2) 16 h of light followed by 8h of darkness at 25°C. No significant changes were recorded between the two groups at the beginning of the study. After 3 mo, the triiodothyronine and elements levels as well as the growth rates of fish in the long-time exposure period were significantly higher than those in the short-time exposure (p<0.05).     

Combined effect of temperature and zinc on Caenorhabditis elegans wild type and daf-21 mutant strains

Heavy metal pollution in aquatic ecosystems is a far reaching environmental problem. The possible influences of heavy metal exposure and the potential harm to organisms when combined with other environmental stressors such as temperature have been largely unexplored. An aquatic toxicity test of Caenorhabditis elegans was performed to estimate the 24 h median lethal concentration (LC₅₀) of different zinc concentrations at different temperatures (15 °C, 20 °C, 25 °C, and 30 °C). We also examined the time course thermotolerance on wild type (N2) and daf-21 null (JT6130) adults exposed to 6.1 mM zinc at 37 °C. Hsp90 protein expression level in response to the combined effect of temperature and zinc toxicity was also investigated by both Western blots and ELISA. Our results show that C. elegans wild type nematodes exhibit severe lethal toxicity after a 24 h exposure to zinc at higher temperatures. In addition, the expression level of Hsp90 was highly inhibited in adult worms subjected to zinc stress. This toxicity assay at different temperatures provides insight into organism response to combined effects of temperature and zinc toxicity.