REVIEW: Evidence supporting the 'preparation for oxidative stress' (POS) strategy in animals in their natural environment.

Hypometabolism is a common strategy employed by resilient species to withstand environmental stressors that would be life-threatening for other organisms. Under conditions such as hypoxia/anoxia, temperature and salinity stress, or seasonal changes (e.g. hibernation, estivation), stress-tolerant species down-regulate pathways to decrease energy expenditures until the return of less challenging conditions. However, it is with the return of these more favorable conditions and the reactivation of basal metabolic rates that a strong increase of reactive oxygen and nitrogen species (RONS) occurs, leading to oxidative stress. Over the last few decades, cases of species capable of enhancing antioxidant defenses during hypometabolic states have been reported across taxa and in response to a variety of stressors. Interpreted as an adaptive mechanism to counteract RONS formation during tissue hypometabolism and reactivation, this strategy was coined "Preparation for Oxidative Stress" (POS). Laboratory experiments have confirmed that over 100 species, spanning 9 animal phyla, apply this strategy to endure harsh environments. However, the challenge remains to confirm its occurrence in the natural environment and its wide applicability as a key survival element, through controlled experimentation in field and in natural conditions. Under such conditions, numerous confounding factors may complicate data interpretation, but this remains the only approach to provide an integrative look at the evolutionary aspects of ecophysiological adaptations. In this review, we provide an overview of representative cases where the POS strategy has been demonstrated among diverse species in natural environmental conditions, discussing the strengths and weaknesses of these results and conclusions.

Sex ratio in the mother's environment affects offspring population dynamics: maternal effects on population regulation.

Classic population regulation theories usually concern the influence of immediate factors on current populations, but studies investigating the effect of parental environment factors on their offspring populations are scarce. The maternal environments can affect offspring life-history traits across generations, which may affect population dynamics and be a mechanism of population regulation. In cyclical parthenogens, sexual reproduction is typically linked with dormancy, thereby providing a negative feedback to population growth. In this study, we manipulated population sex ratios in the mother's environment to investigate whether this factor affected future population dynamics by regulating offspring sexual reproduction in the rotifer Brachionus calyciflorus. Compared with females in male-biased environments, those in female-biased environments produced fewer mictic (sexual) offspring, and their amictic (asexual) offspring also produced a lower proportion of mictic females at a gradient of population densities. Moreover, populations that were manipulated under male-biased conditions showed significantly smaller population sizes than those under female-biased conditions. Our results indicated that in cyclical parthenogens, mothers could adjust the sexual reproduction of their offspring in response to the current population sex ratio, thus providing fine-scale regulation of population dynamics in addition to population density.

Antioxidant defense response during hibernation and arousal in Chinese soft-shelled turtle Pelodiscus sinensis juveniles.

Antioxidant defense is essential for animals to cope with homeostasis disruption during hibernation. The present study aimed to investigate the antioxidant defense response of juvenile soft-shelled turtle Pelodiscus sinensis during hibernation and following arousal. Turtle brain, liver, and kidney samples were collected at pre-hibernation (17 °C mud temperature; MT), during hibernation (5.8 °C MT) and after arousal (20.1 °C MT) in the field. Transcript levels of NF-E2-related factor 2 (Nrf2) decreased significantly during hibernation and recovered after arousal in all tissues. Cerebral and nephric copper-zinc superoxide dismutase (Cu/Zn SOD), catalase (CAT), glutathione peroxidase 3 (GPx3) and nephric GPx4 mRNA showed similar changing patterns as Nrf2. Cerebral Mn SOD, GPx1 and nephric GPx1 up-regulated after arousal. Hepatic Cu/Zn SOD, GPx1 and GPx3 mRNA kept stable, except hepatic GPx4 increased during hibernation. Hepatic Mn SOD and CAT increased after arousal. In the GSH system, mRNA levels of glutathione synthetases (GSs) kept stable during hibernation and up-regulated after arousal in most tissues except nephric GS2 mRNA remained unchanged. Gene expressions of glutathione reductase (GR) exhibited a tissue specific changing pattern, while those of glutathione-S-Transferase (GST) shared a similar pattern among tissues: remained stable or down-regulated during hibernation then recovered in arousal. In contrast to these diverse responses in gene expressions, most of the antioxidant enzyme activities maintained high and stable. Overall, no preparation for oxidative stress (POS) strategy was found in enzymatic antioxidant system in P. sinensis juveniles during hibernation, the Chinese soft-shelled turtles were able to stay safe from potential oxidative stress during hibernation by maintaining high level activities/concentrations of the antioxidant enzymes/antioxidants.

Immune defense parameters of wild fish as sensitive biomarkers for ecological risk assessment in shallow sea ecosystems: A case study with wild mullet (Liza haematocheila) in Liaodong Bay.

Environmental monitoring is important to the health management of an ecosystem. Biomarkers are particularly relevant because they are direct indicators of any toxic effects on organisms and are cheaper to use compared with chemical indicators, especially for extremely low-level organic contaminants. Fish can be significantly affected by pollutants, given their high trophic levels in aquatic food chains. Their immune function is closely related to their survival. The present study compared immune function-related parameters of wild mullet (Liza haematocheila) samples from low (Jinzhou) and high (Yingkou) polluted sites during the pre-winter (PW) and pre-breeding (PB) periods in Liaodong Bay, to evaluate the effect of water pollution on fish health and to explore potential biomarkers of coast water pollution. Compared with Jinzhou mullet, there was a significantly higher level of hematocrit in Yingkou mullet, but a significantly lower serum lysozyme level (P < 0.001), indicating that these fish were immunosuppressed. Significant differences occurred in the spleen between the two site populations. The abnormal: normal fish ratio in Yingkou L. haematochila was significantly higher than that of Jinzhou L. haematochila (2.5 times of that of Jinzhou during PB and nine times during PW). The splenic index of male Yingkou L. haematochila was 47.2% higher than that of Jinzhou L. haematochila in PW (P = 0.001). Moreover, histological observations showed that the spleen of the former was more congestive, with increased numbers (39.6% more) of melanomacrophage centers (MMCs) and changes in pigments (hemosiderin 8.3% higher and melanin 29.4% higher), compared with the latter. The splenic MMC area of Yingkou L. haematochila was significantly smaller than that of Jinzhou L. haematochila (P < 0.05) in PB, but showed no clear difference in PW (P > 0.05). Splenic MMC number was significantly higher in individual Yingkou L. haematochila with abnormal livers compared with normal Yingkou L. haematochila during both sampling periods. The splenic MMC area in abnormal livers was approximately four times those of normal individuals during PB in Yingkou L. haematochila. The number of splenic melanomacrophages (MM) in abnormal livers was approximately nine times those of the normal livers during PW. There were also differences in pigments in normal Yingkou individuals compared with normal Jinzhou samples during PW (melanin 29.4% higher and hemosiderin 8.3% higher). Based on these results, we suggest that serum lysozyme activity, splenic MM number and MMC (both number and area), and melanin of local fish have potential as sensitive biomarkers for the assessment of coastal water pollution.

Positive or negative? The shell alters the relationship among behavioral defense strategy, energy metabolic levels and antioxidant capacity in freshwater turtles.

BACKGROUND: The relationships among energy metabolic levels, behavioral and other physiological traits help to determine the trade-off of energy allocation between different traits and the evolution of life-history driven by natural selection. However, these relationships may be distinctive in selected animal taxa because of their unique traits. In the present study, the relationships among energy metabolic levels, behavioral defense strategies, and antioxidant capacity were explored in three freshwater turtle species with different shell morphologies, by assessing responses to attack, righting time, shell morphology, whole-organism metabolic rates, tissue metabolic enzyme activities and antioxidant levels. RESULTS: The Chinese three-keeled pond turtles, Chinemys reevesii, showed a passive defense strategy, relatively larger shells, a higher resting metabolic rate (RMR) and higher antioxidant levels compared to the snapping turtle, Chelydra serpentina, or the Chinese soft-shelled turtle, Pelodiscus sinensis. These latter two species both showed an active defense strategy, a higher factorial aerobic scope and better muscle anaerobic metabolic capacity but relatively smaller shells, lower RMR and antioxidant capacity. CONCLUSION: Our results indicate a negative relationship between RMR and activity levels in behavioral defense strategies along small-big shell continuum among the three turtle species. We also found a positive relationship between antioxidant capacity and energy metabolism but a negative one between antioxidant capacity and activity levels in defense strategies. The present study indicated a role of turtle shell in forming unique relationship between energy metabolic levels and behaviors in freshwater turtle taxa and a possible trade-off between the maintenance of physiological homeostasis and activity levels in energy allocation.

Antioxidant response to acute cold exposure and during recovery in juvenile Chinese soft-shelled turtles (Pelodiscus sinensis).

The antioxidant defense protects turtles from oxidative stress caused by adverse environment conditions, such as acute thermal fluctuations. However, it remains unclear how these defenses work. The present study examined changes in key enzymes of the enzymatic antioxidant system and the glutathione (GSH) system at both the mRNA and enzyme activity levels during acute cold exposure and recovery in juvenile Chinese soft-shelled turtles, Pelodiscus sinensis Transcript levels of the upstream regulator NF-E2-related factor 2 (Nrf2) were also measured. Turtles were acclimated at 28°C (3 weeks), then given acute cold exposure (8°C, 12 h) and finally placed in recovery (28°C, 24 h). The mRNA levels of cerebral and hepatic Nrf2 and of genes encoding downstream antioxidant enzymes did not change, whereas levels of nephric Nrf2, manganese superoxide dismutase (MnSOD) and glutathione peroxidase 4 (GPx4) mRNA decreased upon cold exposure. During recovery, Nrf2 mRNA remained stable in all three tissues, hepatic Cu/ZnSOD, MnSOD and catalase (CAT) mRNA levels increased, and nephric MnSOD and GPx4 mRNAs did not change from the values during cold exposure. In the GSH system, mRNA levels of most enzymes remained constant during cold exposure and recovery. Unmatched with changes in mRNA level, high and stable constitutive antioxidant enzyme activities were maintained throughout, whereas GPx activity significantly reduced in the kidney during cold exposure, and in liver and kidney during recovery. Our results suggest that the antioxidant defense regulation in response to acute cold exposure in P. sinensis may not be achieved at the transcriptional level, but may rely mainly on high constitutive antioxidant enzyme activities.

Twenty years of the 'Preparation for Oxidative Stress' (POS) theory: Ecophysiological advantages and molecular strategies.

Freezing, dehydration, salinity variations, hypoxia or anoxia are some of the environmental constraints that many organisms must frequently endure. Organisms adapted to these stressors often reduce their metabolic rates to maximize their chances of survival. However, upon recovery of environmental conditions and basal metabolic rates, cells are affected by an oxidative burst that, if uncontrolled, leads to (oxidative) cell damage and eventually death. Thus, a number of adapted organisms are able to increase their antioxidant defenses during an environmental/functional hypoxic transgression; a strategy that was interpreted in the 1990s as a "preparation for oxidative stress" (POS). Since that time, POS mechanisms have been identified in at least 83 animal species representing different phyla including Cnidaria, Nematoda, Annelida, Tardigrada, Echinodermata, Arthropoda, Mollusca and Chordata. Coinciding with the 20th anniversary of the postulation of the POS hypothesis, we compiled this review where we analyze a selection of examples of species showing POS-mechanisms and review the most recent advances in understanding the underlying molecular mechanisms behind those strategies that allow animals to survive in harsh environments.

Maternal Effects Via Resting eggs in Predator Defense of the Rotifer Brachionus calyciflorus.

Maternal effects play important evolutionary and ecological roles. Amictic female mothers of monogonont rotifer Brachionus calyciflorus can transmit predatory information of Asplanchna brightwellii in their environment to their offspring by changing the offspring's defensive morphology to increase their fitness. However, it remains unclear whether such maternal effects also exist during sexual reproduction of a mictic mother. This study explored the maternal effect in mictic mothers using the B. calyciflorus and A. brightwellii as a prey-predator model. We collected resting eggs from two groups of mictic mothers that previously experienced environments with (P environment) or without (NP environment) Asplanchna kairomones. Stem females from the resting eggs of each maternal group were also hatched and reared in P and NP environments. The population growth rate of offspring who experienced the same environment as their mictic mothers was significantly higher than those that experienced a different environment. When exposed to a gradient of predator kairomone levels, the posterolateral spine of the offspring elongated with increasing kairomone concentration. Offspring from the P mictic mother showed significantly shorter posterolateral spines than those from the NP mictic mother at each predator kairomone level. Offspring originating from NP mictic mothers clearly elongated their posterolateral spines at low concentrations of predator kairomones, while those from P mothers elongated their posterolateral spines only at the highest levels of predator kairomone. Our findings highlight the existence of anticipatory maternal effects during the sexual process via resting eggs of B. calyciflorus in response to predator kairomone.

Digital Gene Expression Profiling reveals transcriptional responses to acute cold stress in Chinese soft-shelled turtle Pelodiscus sinensis juveniles.

Turtles are well known for their stress tolerance, including an ability to deal with temperature extremes or rapid thermal change. To know more about the comprehensive molecular basis of thermal stress responses in turtles, we assessed differentially expressed genes (DEGs) in the brain, liver and kidney of juvenile soft-shelled turtles, Pelodiscus sinensis, after acute cold stress (28 °C-8 °C acute transfer and held for 12 h) and following recovery (back to 28 °C and held for 24 h) by digital gene expression profiling. Selected DEGs were also validated via real-time PCR. We found the fewest DEGs in the brain, only one-tenth of the number seen in liver, indicating a tissue-specific gene expression pattern. The DEGs indicated the potential activation of several important functions in response to cold stress and recovery in P. sinensis. This included response to oxidative stress or regulation of reactive oxygen species metabolism in the brain and liver, cerebral inositol metabolism, hepatic monosaccharide metabolism, hepatic complement system, renal DNA repair mechanisms, and TNF and PI3K-Akt signaling pathways in the kidney. These functions likely responded to cold stress in different tissues of P. sinensis to help minimize or repair cell damage as well as enhance innate immunity. The outcomes of this study provide some fundamental insight into the tissue specific complex mechanisms underlining cold stress responses in the soft-shelled turtle P. sinensis.

Antioxidant responses in hibernating Chinese soft-shelled turtle Pelodiscus sinensis hatchlings.

The antioxidant defense system protects turtles from oxidative stress during hibernation. The present study examined changes of the antioxidant enzymes both on mRNA level and enzyme activity level during hibernation of Chinese soft-shelled turtle Pelodiscus sinensis hatchlings. The upstream regulator NF-E2 related factor 2 (Nrf2) mRNA was also measured. Samples were taken at pre-hibernation (17.0°C, Mud temperature (MT)), hibernation (5.8°C, MT) and arousal (20.1°C, MT). Nrf2 exhibited a tissue-specific pattern of expression with a decrease in the brain, slight increase in the liver and heart during hibernation, and significant increase during arousal in all the three tissues. Superoxide dismutase (SOD) mRNA, catalase (CAT) mRNA, and glutathione peroxidase 3 (GPx3) mRNA exhibited a similar pattern as Nrf2 in the brain and liver during the entire hibernation period. Hepatic GPx4 mRNA level increased during hibernation and decreased during arousal, whereas it did not change in the heart. Cerebral SOD and CAT activities kept stable during the experimental period, but GPx activity decreased significantly during hibernation and arousal. Hepatic GPx enzyme activity did not change, whereas those of SOD and CAT exhibited a notable decrease during arousal. Malondialdehyde concentration did not increase during the hibernation process, indicating an effective protection of the antioxidant defense system.

Dimorphic expression of sex-related genes in different gonadal development stages of sterlet, Acipenser ruthenus, a primitive fish species.

Molecular mechanism of sex determination and differentiation of sturgeon, a primitive fish species, is extraordinarily important due to the valuable caviar; however, it is still poorly known. The present work aimed to identify the major genes involved in regulating gonadal development of sterlet, a small species of sturgeon, from 13 candidate genes which have been shown to relate to gonadal differentiation and development in other teleost fish. The sex and gonadal development of sterlets were determined by histological observation and levels of sex steroids testosterone (T), 11-ketotestosterone (11-KT), and 17ß-estradiol (E2) in serum. Sexually dimorphic gene expressions were investigated. The results revealed that gonadal development were asynchronous in 2-year-old male and female sterlets with the testes in early or mid-spermatogenesis and the ovaries in chromatin nucleolus stage or perinucleolus stage, respectively. The levels of T and E2 were not significantly different between sexes or different gonadal development stages while 11-KT had the higher level in mid-spermatogenesis testis stage. In all the investigated gonadal development stages, gene dmrt1 and hsd11b2 were expressed higher in male whereas foxl2 and cyp19a1 were expressed higher in female. Thus, these genes provided the promising markers for sex identification of sterlet. It was unexpected that dkk1 and dax1 had significantly higher expression in ovarian perinucleolus stage than in ovarian chromatin nucleolus stage and in the testis, suggesting that these two genes had more correlation with ovarian development than with the testis, contrary to the previous reports in other vertebrates. Testicular development-related genes (gsdf and amh) and estrogen receptor genes (era and erb) differentially expressed at different testis or ovary development stages, but their expressions were not absolutely significantly different in male and female, depending on the gonadal development stage. Expression of androgen receptor gene ar or rspo, which was supposed to be related to ovarian development, presented no difference between gonadal development stages investigated in this study whenever in male or female.

Ascorbic acid regulation in stress responses during acute cold exposure and following recovery in juvenile Chinese soft-shelled turtle (Pelodiscus sinensis).

Intense temperature change often leads to increased oxidative stress in many animals with a few exceptions, including the turtle. To date, little is known about the mechanism of protective antioxidative defenses in turtles during acute temperature change, specifically the role that the antioxidant ascorbic acid (AA) plays. In this study, Chinese soft-shelled turtles (Pelodiscus sinensis) were initially acclimated at 28°C (3 wks), exposed to acute cold condition (8°C, 8 h) and finally placed in recovery (28°C, 24 h). L-Gulonolactone oxidase (GLO) mRNA exhibited a stable transcription pattern during the intense thermal fluctuation. GLO activity also remained stable, which validated the mRNA expression pattern. The similar Q10 values for GLO activity in the different treatment groups at incubation temperatures of 28°C and 8°C indicated that the GLO activity response to thermal change exhibited a temperature-dependent enzymatic kinetic characteristic. The AA storage was tissue-specific as well as the AA re-supply in the recovery period, with brain as the priority. Despite the insufficient transport during cold exposure, the plasma AA reservoir greatly contributed to the redistribution of AA during recovery. Depending on the prominent GLO activity, the high level of tissue-specific AA storage and the extraordinary plasma AA transport potential, the Chinese soft-shelled turtle endured severe thermal fluctuations with no apparent oxidative stress. However, the significant decrease in AA concentration in the brain tissue during acute cold exposure suggested that such a strategy may not be sufficient for prolonged cold exposure.

Spatial heterogeneity of stream environmental conditions and macroinvertebrates community in an agriculture dominated watershed and management implications for a large river (the Liao River, China) basin.

Understanding the effects of watershed land uses (e.g., agriculture, urban industry) on stream ecological conditions is important for the management of large river basins. A total of 41 and 56 stream sites (from first to fourth order) that were under a gradient of watershed land uses were monitored in 2009 and 2010, respectively, in the Liao River Basin, Northeast China. The monitoring results showed that a total of 192 taxa belonging to four phyla, seven classes, 21 orders and 91 families were identified. The composition of macroinvertebrate community in the Liao River Basin was dominated by aquatic insect taxa (Ephemeroptera and Diptera), Oligochaeta and Molluscs. The functional feeding group GC (Gatherer/Collector) was dominant in the whole basin. Statistical results showed that sites with less watershed impacts (lower order sites) were characterized by higher current velocity and habitat score, more sensitive taxa (e.g., Ephemeroptera), and the substrate was dominated by high percentage of cobble and pebble. The sites with more impacts from agriculture and urban industry (higher order sites) were characterized by higher biochemical (BOD5) and chemical oxygen demand (COD), more tolerant taxa (e.g., Chironominae), and the substrate was dominated by silt and sand. Agriculture and urban-industry activities have reduced habitat condition, increased organic pollutants, reduced macroinvertebrate abundance, diversity, and sensitive taxa in streams of the lower Liao River Basin. Restoration of degraded habitat condition and control of watershed organic pollutants could be potential management priorities for the Basin.

Diverse Response Pattern to Anoxia in Three Freshwater Turtle Species.

With increasing water eutrophication and global warming, anoxia and hypoxia are becoming more and more common in water environments. Most vertebrates have a limited tolerance to anoxia of only a few minutes, but some species, such as turtles, can survive for months being exposed to anoxia. Antioxidant defense systems may have a potential role in resisting anoxia stress in freshwater turtles. The three-keeled pond turtle Chinemys reevesii, the snapping turtle Chelydra serpentina and the soft-shelled turtle Pelodiscus sinensis are three popular aquaculture species and share similar habitats in China. While C. reevesii and C. serpentina are hard-shelled turtles with poor skin permeability, P. sinensis is soft-shelled turtle whose skin permeability is good. We examined the antioxidant defense responses in different tissues of the three turtle species under acute anoxia stress for 10 h and subsequently recovered for 24 h in order to reveal the response patterns of the antioxidant defense system of the three turtle species that differed in morphological structure and life history strategy. We found that the antioxidant response patterns to acute anoxia stress were tissue- and species-specific. The soft-shelled turtle was more sensitive to anoxia than the hard-shelled turtles. Under anoxia stress, the three species kept the activities of most antioxidant enzymes stable. C. reevesii and P. sinensis were highly dependent on vitamin C in antioxidant defense, while high activities of structural antioxidant enzymes were found in the tissues of C. serpentina. The above diverse patterns may be related with adaptive evolution of morphological structure and physiological functions of the three turtle species.

Effects of Extreme Light Cycle and Density on Melatonin, Appetite, and Energy Metabolism of the Soft-Shelled Turtle (Pelodiscus sinensis).

Constant darkness and constant light exposure often disturb the circadian rhythm in the behavior and energy metabolism of vertebrates. Melatonin is known as the hormonal mediator of photoperiodic information to the central nervous system and plays a key role in food intake and energy balance regulation in vertebrates. The popularly cultured soft-shelled turtle Pelodiscus sinensis has been reported to grow better under constant darkness; however, the underlying physiological mechanism by which darkness benefits turtle growth is not clear yet. We hypothesized that increased melatonin levels induced by darkness would increase appetite and energy metabolism and thus promote growth in P. sinensis. In addition, in order to elucidate the interaction of photoperiod and density, juvenile turtles were treated under three photoperiods (light/dark cycle: 24L:0D, 12L:12D, 0L:24D, light density 900 lux) and two stocking densities (high density: 38.10 ind./m2, low density: 6.35 ind./m2) for 4 weeks, and then the blood and brain tissues of turtles were collected during the day (11:00-13:00) and at night (23:00-1:00) after 2 days of fasting. We examined changes in plasma melatonin levels, food intake (FI), and appetite-related hormones (plasma ghrelin and leptin), as well as growth and energy metabolism parameters such as specific growth rate (SGR), standard metabolic rate (SMR), plasma growth hormone (GH), and thyroid hormone/enzyme activity (plasma triiodothyronine T3, thyroxine T4, and T45'-deiodinase activity). Moreover, we also assessed the responses of mRNA expression levels of food intake-related genes (kisspeptin 1 (Kiss1); cocaine amphetamine-regulated transcript (CART); neuropeptide Y (NPY)) in the brain. The results showed that under high density, SGR was the lowest in 24L:0D and the highest in 0L:24D. FI was the highest in 0L:24D regardless of density. Plasma melatonin was the highest in 0L:24D under high density at night. SMR increased with decreasing light time regardless of density. Most expressions of the measured appetite-related genes (Kiss1, CART, and NPY) were not affected by photoperiod, nor were the related hormone levels, such as plasma leptin, ghrelin, and GH. However, thyroid hormones were clearly affected by photoperiod. T3 level in 0L:24D under high density during the day was the highest among all treatment groups. T4 in 24L:0D under high density during the day and T45'-deiodinase activity in 24L:0D under low density at night were significantly reduced compared with the control. Furthermore, the energy metabolism-related hormone levels were higher under higher density, especially during the day. Together, melatonin secretion is not only modulated by light but also likely to be regulated by unknown endogenous factors and density. Altered plasma melatonin induced by constant darkness and density seems to be involved in the modulation of energy metabolism rather than appetite in the soft-shelled turtle.

Molecular cloning and characterization of Foxp3 in Atlantic salmon (Salmo salar).

Foxp3 is a T cell-specific transcription factor and plays a key role in the development of Treg cells and in the immune regulatory process during inflammation. Here we report cloning and characterization of the full-length cDNA of Atlantic salmon Foxp3, which possesses a Forkhead domain, a zinc finger domain and a leucine-zipper domain as its counterpart in mammals. Foxp3 is highly expressed in thymus. Furthermore, regulated expression was observed in head kidney cells in response to ß-glucan and mitogens (LPS and ConA), and in the head kidney, spleen and liver after intraperitoneal injection of live Aeromonas salmonicida. In addition, transfection of CHSE-214 cells with salmon Foxp3 fused with a C-termial RFP tag, resulted in the expression of the transgene in and close to the nuclei upon stimulation. Taken together, these results suggest the presence of a Foxp3 gene in Atlantic salmon that may be an important transcription factor in immune regulation, and further research may reveal the existence of Treg-like T cells in this species.

Comparison of Aeromonas salmonicida resistant and susceptible salmon families: a high immune response is beneficial for the survival against Aeromonas salmonicida challenge.

Selective breeding has been employed to improve resistance to infectious diseases in aquaculture and it is of importance to investigate the expression profiles of immune genes together with complement activity of Atlantic salmon with different genetic background in response to pathogens, in particular against Aeromonas salmonicida. This study examined acute phase products, and several central T cell cytokines and a transcription factor in different tissues, namely head kidney, spleen and liver, in two families of Atlantic salmon with high and low mortalities, after challenge by A. salmonicida. The results showed that the expression pattern of target genes differed in lymphoid and non-lymphoid organs in the two families. Generally, in lymphoid organs, higher expression of pro-inflammatory genes, such as TLR5M, TLR5S, GATA3, IFN-γ, IL-17D, as well as the pleiotropic cytokine gene IL-10 in the resistant family was observed at the same time point. One may speculate that a relatively high immune response is a pre-requisite for increased survival in a A. salmonicida challenge test. In addition, the resistant fish possessed higher complement activity pre-challenge compared to susceptible fish. Complement activity may be applied as an indicator in selective breeding for enhanced disease resistance.

Effect of maternal ammonia stress on population dynamics of the rotifer Brachionus calyciflorus offspring.

Maternal effects play important roles in phenotypic variations among individuals and are thus considered to regulate population performance in responses to environmental stress. High ammonia levels are known to suppress population growth of the rotifer Brachionus calyciflorus. However, it remains unclear whether maternal environmental ammonia stress influences the offspring phenotypic variation and, if so, how it affects the offspring population dynamics in the rotifer. The present work examined variations in life history, morphology, feeding and digestive activities of B. calyciflorus offspring affected by maternal ammonia stress and the effect of the above variations on offspring population dynamics. We observed increased fitness in the offspring population affected by the cumulative maternal effect. There was a trade-off between offspring (F1) survival and reproductive investment under maternal (F0) ammonia stress. Population growth of the offspring possibly increased via decreasing body size and posterolateral spine length while enhancing cellulase activity. Moreover, the absence of the posterolateral spine of the rotifer was a sensitive response to maternal ammonia stress. These findings underscore maternal environmental stress as an important source of phenotypic variations and highlight these multiple responses work together to affect population dynamics.

Effects of chronic exposure to nonylphenol on locomotor activity and social behavior in zebrafish (Danio rerio).

Nonylphenol (NP), a xenoestrogen ubiquitously found in aquatic ecosystems, is of high environmental concern. The present work assessed the effects of exposure to NP on locomotor activity and social behavior in male and female zebrafish (Danio rerio), to evaluate the possible hazard of NP to fish behavior and to pursue a potential biomarker of NP contamination. Fish were randomly divided into six groups. Five groups were receiving 0.1, 1, 10, 50 and 100 microg/L nominal concentrations of NP for 60 days, respectively. A sixth control group was given the same treatment as the other five groups, but no NP. Locomotor activity, aggressive behavior, group preference and leaving shoal were examined. NP exposure showed marked influence on locomotor activity of the male zebrafish, whereas that of the female was not significantly affected by NP. Aggressive behavior and group preference were significantly affected by NP exposure in both male and female groups. Locomotor activity and aggressive behavior of the male and group preference of both male and female zebrafish were clearly inhibited at 100 microg/L NP. No significant alteration in leaving shoal was observed under NP exposure. The results suggested that changes in locomotor activity and aggressive behavior of the male and group preference of both male and female may be used as an ecologically relevant integrative biomarker of NP contamination.

A New IL6 Isoform in Chinese Soft-Shelled Turtle (Pelodiscus sinesis) Discovered: Its Regulation during Cold Stress and Infection.

The Chinese soft-shelled turtle (Pelodiscus sinesis) is a widely cultured commercial species in East and Southeast Asian countries. The turtles frequently suffer from acute cold stress during farming in China. Stress-induced factor such as Interleukin-6 (IL6) is a multifunctional molecule that plays important roles in innate and adaptive immune response. In the present study, we found that the turtle possessed two IL6 transcripts, where one IL6 transcript contained a signal peptide sequence (psIL6), while the other IL6 transcript (psIL6ns) possessed no such signal peptide gene. To test any differential expression of the two isoforms during temperature and microbial stress, turtles were adapted to optimal environmental water temperature (25 °C), stressed by acute cooling for 24 h, followed with the challenge of Aeromonas hydrophila (1.8 × 108 CFU) or Staphylococcus aureus (5.8 × 108 CFU). Gene characterization revealed that psIL6ns, a splicer without codons encoding a signal peptide and identical to the one predicted from genomic sequence, and psIL6, a splicer with codons encoding a signal peptide, were both present. Inducible expression was documented in primary spleen cells stimulated with ConA and poly I: C. The splenic and intestinal expression of psIL6ns and psIL6 was increased in response to temperature stress and bacterial infection.