Naive Fish Research Articles

Zbtb46 Controls Dendritic Cell Activation by Reprogramming Epigenetic Regulation of cd80/86 and cd40 Costimulatory Signals in a Zebrafish Model.

The establishment of an appropriate costimulatory phenotype is crucial for dendritic cells (DCs) to maintain a homeostatic state with optimal immune surveillance and immunogenic activities. The upregulation of CD80/86 and CD40 is a hallmark costimulatory phenotypic switch of DCs from a steady state to an activated one for T cell activation. However, knowledge of the regulatory mechanisms underlying this process remains limited. In this study, we identified a Zbtb46 homolog from a zebrafish model. Zbtb46 deficiency resulted in upregulated cd80/86 and cd40 expression in kidney marrow-derived DCs (KMDCs) of zebrafish, which was accompanied with a remarkable expansion of CD4+/CD8+ T cells and accumulation of KMDCs in spleen of naive fish. Zbtb46 -/- splenic KMDCs exhibited strong stimulatory activity for CD4+ T cell activation. Chromatin immunoprecipitation-quantitative PCR and mass spectrometry assays showed that Zbtb46 was associated with promoters of cd80/86 and cd40 genes by binding to a 5'-TGACGT-3' motif in resting KMDCs, wherein it helped establish a repressive histone epigenetic modification pattern (H3K4me0/H3K9me3/H3K27me3) by organizing Mdb3/organizing nucleosome remodeling and deacetylase and Hdac3/nuclear receptor corepressor 1 corepressor complexes through the recruitment of Hdac1/2 and Hdac3. On stimulation with infection signs, Zbtb46 disassociated from the promoters via E3 ubiquitin ligase Cullin1/Fbxw11-mediated degradation, and this reaction can be triggered by the TLR9 signaling pathway. Thereafter, cd80/86 and cd40 promoters underwent epigenetic reprogramming from the repressed histone modification pattern to an activated pattern (H3K4me3/H3K9ac/H3K27ac), leading to cd80/86 and cd40 expression and DC activation. These findings revealed the essential role of Zbtb46 in maintaining DC homeostasis by suppressing cd80/86 and cd40 expression through epigenetic mechanisms.

The emergence of collective knowledge and cumulative culture in animals, humans and machines.

The emergence of collective knowledge and cumulative culture in animals, humans and machines.

Vaccination of Gilthead Seabream After Continuous Xenoestrogen Oral Exposure Enhances the Gut Endobolome and Immune Status via GPER1.

In fish culture settings, the exogenous input of steroids is a matter of concern. Recently, we unveiled that in the gilthead seabream (Sparus aurata), the G protein-coupled estrogen receptor agonist G-1 (G1) and the endocrine disruptor 17α-ethinylestradiol (EE2) are potent modulators in polyreactive antibody production. However, the integral role of the microbiota upon immunity and antibody processing in response to the effect of EE2 remains largely unexplored. Here, juvenile seabreams continuously exposed for 84 days to oral G1 or EE2 mixed in the fish food were intraperitoneally (i.p.) immune primed on day 42 with the model antigen keyhole limpet hemocyanin (KLH). A critical panel of systemic and mucosal immune markers, serum VTG, and humoral, enzymatic, and bacteriolytic activities were recorded and correlated with gut bacterial metagenomic analysis 1 day post-priming (dpp). Besides, at 15 dpp, animals received a boost to investigate the possible generation of specific anti-KLH antibodies at the systemic and mucosal interphases by the end of the trial. On day 43, EE2 but not G1 induced a significant shift in the serum VTG level of naive fish. Simultaneously, significant changes in some immune enzymatic activities in the serum and gut mucus of the EE2-treated group were recorded. In comparison, the vaccine priming immunization resulted in an attenuated profile of most enzymatic activities in the same group. The gut genes qPCR analysis exhibited a related pattern, only emphasized by a significant shift in the EE2 group’s il1b expression. The gut bacterial microbiome status underwent 16S rRNA dynamic changes in alpha diversity indices, only with the exposure to oral G1, supporting functional alterations on cellular processes, signaling, and lipid metabolism in the microbiota. By the same token, the immunization elevated the relative abundance of Fusobacteria only in the control group, while this phylum was depleted in both the treated groups. Remarkably, the immunization also promoted changes in the bacterial class Betaproteobacteria and the estrogen-associated genus Novosphingobium. Furthermore, systemic and mucosal KLH-specific immunoglobulin (Ig)M and IgT levels in the fully vaccinated fish showed only slight changes 84 days post-estrogenic oral administration. In summary, our results highlight the intrinsic relationship among estrogens, their associated receptors, and immunization in the ubiquitous fish immune regulation and the subtle but significant crosstalk with the gut endobolome.

Immersion vaccines against Yersinia ruckeri infection in rainbow trout: Comparative effects of strain differences

The protective effects of autogenous and commercial ERM immersion vaccines (bacterins based on Yersinia ruckeri, serotype O1, biotypes 1 and 2) for rainbow trout (Oncorhynchus mykiss) were compared in order to evaluate whether the use of local pathogen strains for immunization can improve protection. In addition, the effect of the bacterin concentration was established for the commercial product. Following sublethal challenge of vaccinated and non‐vaccinated control fish with live bacteria, we followed the bacterial count in the fish (gills, liver and spleen). The expression of genes encoding immune factors (IL‐1β, IL‐6, IL‐8, IL‐10, IFN‐γ, MHCI, MHCII, CD4, CD8, TCRβ, IgM, IgT, IgD, cathelicidins 1 and 2, SAA and C3) and densities of immune cells in organs were recorded. Both vaccines conferred protection as judged from the reduced bacterial load in exposed fish. Innate immune genes were upregulated in all groups following bacterial challenge but significantly more in non‐vaccinated naive fish in which densities of SAA‐positive immune cells increased. Immunoglobulin genes were upregulated on day 5 post‐challenge, and fish vaccinated with the high commercial bacterin dosage showed increased IgM levels by ELISA on day 14 post‐challenge, reflecting that the vaccine dosage was correlated to protection. In conclusion, both vaccine types offered protection to rainbow trout when exposed to live Y. ruckeri and no significant difference between commercial and autogenous vaccines was established.

The effect of temperature on the spatial learning rate of zebrafish (Danio rerio)

AbstractTemperature has a wide range of effects on the biology and ecology of ectotherms, including fish. However, the literature provides only a few studies assessing its effect on their cognitive abilities. We tested the effect of temperature on the spatial learning rate of zebrafish by comparing the daily rate of change in the number of fish choosing the correct arm first and behavioural performance (i.e., distance travelled, time needed to locate the daily food reward and swimming speed) in two temperatures (21 and 31°C) in 7 successive sessions. The daily rate of change in behavioural performance was expressed as the percentage difference between the value in a given session and the value in the previous session. Two experiments were performed in a T‐maze with single feeders in the arms, one empty and the other with a food reward. In each of the experiments, we used 10 naive fish, which were placed in the T‐maze individually once a day. The fish were fed between sessions to avoid increasing differences in hunger levels and, in turn, increasing the differences in motivation to find the food between the temperature treatments. The results revealed that the learning rate was greater at the higher temperature, which was apparent in a greater percentage reduction in the time and distance travelled needed to locate the daily food reward between successive sessions at the higher temperature than at the lower temperature. The results show there was a significant effect of temperature on the daily rate of change in the number of fish choosing the correct arm, distance travelled and time needed to locate the daily food reward, which may be attributed to the positive effect of temperature on learning rate, and may indicate the importance of temperature for the cognitive testing of zebrafish.

Amino acid availability of processed Brassica carinata meals in hybrid striped bass Morone chrysops ♀ x M. saxatilis ♂

An earlier study showed that hybrid striped bass (Morone chrysops ♀ x M. saxatilis ♂) diets containing 20 % animal meals and up to 30 % aerobically fermented (AF) carinata meal (Brassica carinata, CM) followed by a single wash with water (AFCM) or 30 % CM double-washed with water without AF (WCM), resulted in similar growth. However feed utilization and Fulton’s condition factor of 30 % WCM were better. The current study investigated effects of AFCM and WCM on amino acid digestibilities using naive fish. Effects of AFCM and WCM on trypsin activity, and amino acid availability were determined using hybrid striped bass fed experimental diets for 127 and 148 days, respectively (after a growth trial). High (30 %) AFCM reduced all essential amino acids (EAA) and conditionally EAA (CEAA) except histidine, methionine and threonine. However, all EAA and CEAAs except lysine and cysteine were digested better in 30 %WCM than 30 % AFCM. High (30 %) CMs resulted in generally high serum free methionine and tryptophan, but less leucine for the first 9 h. However, 30 % WCM resulted in generally more serum free arginine and valine than 30 % AFCM. High CMs increased muscle free histidine. Generally, WCM resulted in more muscle lysine and valine than AFCM. Muscle methionine was increased by CMs. Both serum and muscle free total EAAs in hybrid striped bass fed 30 % WCM were elevated continuously over a longer period above the baseline levels. Using EAA:lysine, only muscle histidine were adequate for muscle synthesis over the 36-h period. Overall, amino acids were more available in WCM than AFCM.

Impact of vaccination and selective breeding on the transmission of Infectious salmon anemia virus

A transmission trial was carried out using 420 Atlantic salmon (Salmo salar L., parr stage) to simultaneously test the impact of vaccination and selective breeding on the transmission of Infectious salmon anemia virus (ISAV). Genetic difference in disease resistance was based on mortality differences across 15 families (defined by a genomic breeding value (GEBV)) and organised into three groups (low GEBV (LBV), mid GEBV (MBV) and high GEBV (HBV)). Three different shedder groups were infected with ISAV before being placed in tanks (T1 – T18) with naive contact fish (n = 15), one representative from each of the 15 families. The shedder groups included LBV vaccinated (LBVv), LBV not vaccinated (LBVnv) and HBV not vaccinated (HBVnv). The trial was run with two consecutive sets of nine tanks so the infectiousness of the shedder fish could be tested at different stages in their infection process (early (3–9 days post-infection (dpi) versus late (9–15 dpi)). Infection and mortality data of the contact fish were analysed using a Generalised Linear Mixed Model (GLMM) and a Bayesian epidemiological model. Neither vaccination nor genetic resistance prevented transmission, but both lowered the probability of infection in contact fish. Though not statistically significant, the effect of genetic resistance was larger (LBVNV vs HBVNV: odds ratio: 8.35 (0.75–93.36)) than vaccination (LBVNV vs LBVV: odds ratio: 4.52 (0.43–46.99)). There was no difference in the susceptibility of fish with different resistance breeding values, however, significant differences were found in their endurance to ISAV infection, with LBV fish dying 14 days earlier than HBV fish. Mortality as a resistance phenotype in breeding programmes appears to simultaneously improve survival of ISAV infected fish and reduce ISA transmission. However, it would be beneficial to evaluate mucus viral load (MVL) as an additional phenotype to more effectively reduce ISA transmission.

Follow the straggler: zebrafish use a simple heuristic for collective decision-making.

Animal groups often make decisions sequentially, from the front to the back of the group. In such cases, individuals can use the choices made by earlier ranks, a form of social information, to inform their own choice. The optimal strategy for such decisions has been explored in models which differ on, for example, whether or not agents take into account the sequence of observed choices. The models demonstrate that choices made later in a sequence are more informative, but it is not clear if animals use this information or rely instead on simpler heuristics, such as quorum rules. We show that a simple rule 'copy the last observed choice', gives similar predictions to those of optimal models for most likely sequences. We trained groups of zebrafish to choose one arm of a Y-maze and used them to demonstrate various sequences to naive fish. We show that the naive fish appear to use a simple rule, most often copying the choice of the last demonstrator, which results in near-optimal choices at a fraction of the computational cost.

Temperature training improves transcriptional homeostasis after heat shock in juvenile Atlantic sturgeon (Acipenser oxyrinchus).

Exposure to high temperatures can lead to thermotolerance in fish, which is hypothesized to potentially improve post-release survival in species under restocking programs, like Atlantic sturgeon. The aim of this study was to determine whether Atlantic sturgeon juveniles exposed to a 4-week temperature treatment respond differently to a subsequent heat shock than juveniles exposed to heat shock for the first time (naive fish). Response to heat shock was assessed by mapping the liver transcriptome. In total, 838 unique contigs were differentially expressed between the trained and the control group (592 downregulated, 261 upregulated, and 15 down- or upregulated, depending on the condition), corresponding to genes involved in the response to heat, tissue damage, proteolysis, and metabolism. Temperature-trained fish showed 2-4-fold fewer dysregulated contigs than naive fish, indicating their ability to maintain and recover homeostasis faster. During heat shock, hspc1 was upregulated in both experimental groups, while hspa1 and dnaja4 were exclusively upregulated in the control. Overall, compensatory mechanisms were observed in addition to the heat shock response. Only two genes, fgg and apnl, were upregulated at nearly all timepoints in both groups. Peptidases were more strongly downregulated in control fish, which also showed a reduction in lipid metabolism during recovery. Keratins, pck1, gadd45ga, and gadd45gb were differentially expressed between trained and control fish, and due to their roles in tissue protection and ER stress reduction, they might be responsible for the maintenance of the transcriptional homeostasis observed in trained fish.

Training fish for restocking: refuge and predator training in the hatchery has limited benefits for a marine fish.

Large numbers of hatchery-reared fish are released in stocking programmes; however, success is limited by high mortality. Predation is seen as the main cause of deaths but might be reduced by training fish before release to avoid predators and/or use refuge. In this study on a potential restocking species, yellowfin bream Acanthopagrus australis, the effects of predator training and refuge on the behaviour of fish in the hatchery were tested. In the first experiment, juvenile bream were exposed to predatory mangrove jack (Lutjanus argentimaculatus) fed exclusively on bream flesh while housed in tanks with and without refuge. Predator training altered fish behaviour when fish were re-exposed to predators, but the effects were subtle and varied between groups of fish. In contrast, refuge created strong and consistent changes in behaviour, significantly slowing down the amount of time that fish took to consume food. A second experiment focused on the effects of refuge. Bream were trained to use artificial seagrass or house bricks as refuge and then exposed to mangrove jacks in a laboratory predation experiment. When refuge was available, fish significantly slowed down their feeding rate. There was a small, transient increase in survival for fish given seagrass refuges, but this was irrespective of whether the bream were trained to use refuge. The results of this study indicate that the use of refuge may be innate and the benefits of refuge may be available to naive hatchery-reared fish or fish trained to use refuge shortly before release. This suggests that there is potential to improve post-release survival of fingerlings without time-consuming and expensive hatchery training.

Reversible morphological changes in a juvenile marine fish after exposure to predatory alarm cues.

Chemical cues from predators induce a range of predator-induced morphological defences (PIMDs) observed across fish taxa. However, the mechanisms, consistency, direction and adaptive value of PIMDs are still poorly studied. Here, we have tested if predatory cues can induce changes in the body shape of the juvenile marine fish Sparus aurata reared under controlled conditions without the presence of predators by exposing individuals to the olfactory stimulus of a fish predator. We tested our hypothesis using a nested replicated before-after-control-impact experiment, including recovery (potential reversibility) after the cessation of the predator stimulus. Differences in the size-independent body shape were explored using landmark-based geometric morphometrics and revealed that, on average, individuals exposed to a predatory cue presented deeper bodies and longer caudal regions, according to our adaptive theoretical predictions. These average plastic responses were reversible after withdrawal of the stimulus and individuals returned to average body shapes. We, therefore, provide evidence supporting innate reversible PIMDs in marine naive fish reared under controlled conditions. The effects at the individual level, including fitness and the associated applied implications, deserve further research.

Together We Stand – Analyzing Schooling Behavior in Naive Newborn Guppies through Biorobotic Predators

A major advantage of animal aggregations concerns cooperative antipredator strategies. Schooling behavior emerges earlier in many fish species, especially in those cannibalizing their offspring. Experience is fundamental for developing schooling behavior. However, the cognitive ability of naive newborn fish to aggregate remains unclear. Herein, Poecilia reticulata, was selected as model organism to investigate how combinations of biomimetic robotic agents and adult conspecific olfactory cues affect collective responses in newborns. The role of white and brown backgrounds in evoking aggregations was also assessed. Olfactory cues were sufficient for triggering aggregations in P. reticulata newborns, although robotic agents had a higher influence on the group coalescence. The combination of robotic agents and olfactory cues increased schooling behavior duration. Notably, schooling was longer in the escape compartment when robotic agents were presented, except for the combination of the male-mimicking robotic fish plus adult guppy olfactory cues, with longer schooling behavior in the exploring compartment. Regardless of the tested cues, newborn fish aggregated preferentially on the brown areas of the arena. Overall, this research provides novel insights on the early collective cognitive ability of newborn fish, paving the way to the use of biomimetic robots in behavioral ecology experiments, as substitutes for real predators.

A sense of place: pink salmon use a magnetic map for orientation.

The use of 'map-like' information from the Earth's magnetic field for orientation has been shown in diverse taxa, but questions remain regarding the function of such maps. We used a 'magnetic displacement' experiment to demonstrate that juvenile pink salmon (Oncorhynchus gorbuscha) use magnetic cues to orient. The experiment was designed to simultaneously explore whether their magnetic map is used to direct fish (i) homeward, (ii) toward the center of their broad oceanic range or (iii) along their oceanic migratory route. The headings adopted by these navigationally naive fish coincided remarkably well with the direction of the juveniles' migration inferred from historical tagging and catch data. This suggests that the large-scale movements of pink salmon across the North Pacific may be driven largely by their innate use of geomagnetic map cues. Key aspects of the oceanic ecology of pink salmon and other marine migrants might therefore be predicted from magnetic displacement experiments.

Effects of SCUBA bubbles on counts of roving piscivores in a large remote marine protected area.

This study examined the effects of SCUBA bubbles on fish counts in underwater visual surveys conducted in the Papahānaumokuākea Marine National Monument (PMNM). Specifically, paired fish surveys were conducted at each survey site, utilizing two different gear types: open-circuit SCUBA (OC) and closed-circuit rebreather (CCR). Bubble exhaust released from the OC equipment is a potential source of bias for in-situ fish observations, as the associated audio and visual disturbances could either attract or repel fishes depending on whether their behavior is more driven by curiosity or caution. The study area, is a large (~1.5 million km2) and extremely remote marine protected area in which the response of coral reef fishes to divers represent natural behavior of naive fishes with little or no previous contact with humans. Historically, surveys conducted on OC in this area have shown an abundance of large roving piscivores and this study set out to determine the extant, if any, the audible and visual disturbances of OC bubbles have. The species typically seen in these prior surveys were Caranx ignobilis, Caranx melampygus, Aprion virescens, and a couple of species of sharks. We found differences in counts for some roving piscivores, including significantly more jacks observed on OC than CCR (Caranx ignobilis 57% more, and Caranx melampygus 113% more). Instance of first encounter, i.e. the time when a fish was first observed during a survey, also varied for some species. Higher numbers of Aprion virescens (p = 0.04), and C. melampygus (p = <0.001) were observed in the first 5-minutes of counts by divers on OC (i.e. when they were using breathing apparatus that produced noises that could be heard over long distances). Although not the focus of the study, we also assessed differences between OC and CCR counts for other groups of fishes. Estimated abundance of benthic damselfish was higher on OC than CCR, and counts of butterflyfish were lower on OC; but there were no significant differences for the other groups considered. This is an important control study that documents the natural responses of coral reef fishes to SCUBA bubbles generated by in-situ surveys.

Is Habituation Measurable in Lumpfish Cyclopterus lumpus When Used as Cleaner Fish in Atlantic Salmon Salmo salar Aquaculture?

To investigate how lumpfish interact in Atlantic salmon aquaculture, physiological stress responses and changes in behaviour were analysed in experienced and naive lumpfish. Experienced lumpfish (30.2 ± 7.93 g, mean ± SD) coexisted with a commercial scale production unit of Atlantic salmon (1258.5 ± 152.12 g) for 30 to 60 days, while naive lumpfish (38.2 ± 12.37 g) were kept with conspecifics only. Ten trials from each background were tested. For each trial, 10 lumpfish were tagged and transferred to a video monitored experimental tank (2 × 2 × 0.7 m). In each trial, swimming behaviour was mapped for all lumpfish every 60 s in 20 min, 10 min before, and 10 min after the introduction of four Atlantic salmon. Naive lumpfish expressed significantly increased burst swimming activity and maintained longer interspecific distance to Atlantic salmon in comparison with experienced fish. In addition, mean plasma cortisol levels were significantly elevated in naive fish after exposure to Atlantic salmon. We argue that naive lumpfish expressed innate physiological and behavioural stress responses during first encounter with Atlantic salmon, while reduced responses in experienced individuals indicated habituation. The effect from behavioural and physiological stress in newly deployed naive lumpfish–before and during habituation–should be taken account for when lumpfish are introduced in commercial sea cages to improve welfare for the species. In addition, we suggest that habituation could be applicable during the rearing phase to moderate the transition from a simple tank environment with conspecifics only to interspecies interaction with Atlantic salmon in sea cages.

Nurture is above nature: nursery experience determines habitat preference of red sea bream Pagrus major juveniles

Habitat preference is thought to be genetically programmed in fishes. However, fishes can choose habitat based on their personal experience of an environment. We investigated whether the environment in which fish are raised affects habitat preference in red sea bream Pagrus major juveniles, and tested if the formed preference lasts until later life stages. Juveniles were reared in tanks with a substrate of either sand or artificial seaweed for 40 days. Naive fish were raised without either type of substrate. In the preference test, individual fish were allowed to choose either a sand or artificial seaweed microhabitat. The tested fish were then kept in barren tanks, and similar tests conducted again on days 30 and 100. Sand and seaweed treatment fish preferred the corresponding habitat immediately after the rearing treatment, whereas naive fish did not exhibit any preference. These preferences were maintained when fish were tested on day 30, but not on day 100. The present study suggests that habitat preference is acquired through the rearing environment at the nursery stage, and that this preference lasts for at least 30 days. The formation of habitat preference should help juveniles to choose an optimal microhabitat in a fluctuating environment.

Winning the arms race: host-parasite shared evolutionary history reduces infection risks in fish final hosts.

Parasite manipulation of intermediate hosts evolves to increase parasite trophic transmission to final hosts, yet counter selection should act on the final host to reduce infection risk and costs. However, determining who wins this arms race and to what extent is challenging. Here, for the first time, comparative functional response analysis quantified final host consumption patterns with respect to intermediate host parasite status. Experiments used two evolutionarily experienced fish hosts and two naive hosts, and their amphipod intermediate hosts of the acanthocephalan parasite Pomphorhynchus tereticollis. The two experienced fish consumed significantly fewer infected than non-infected prey, with lower attack rates and higher handling times towards the former. Conversely, the two naive fish consumed similar numbers of infected and non-infected prey at most densities, with similar attack rates and handling times towards both. Thus, evolutionarily experienced final hosts can reduce their infection risks and costs via reduced intermediate host consumption, with this not apparent in naive hosts.

Susceptibility of the topmouth gudgeon (Pseudorasbora parva) to CyHV-3 under no-stress and stress conditions

Cyprinid herpesvirus 3 (CyHV-3), also known as koi herpesvirus, is the causative agent of the highly contagious koi herpesvirus disease, which is restricted to koi and common carp and causes significant losses in both fish stock. Some experimental investigations have shown that other cyprinid or non-cyprinid species may be asymptomatically susceptible to this virus and might play roles as potential carriers of CyHV-3 or might contribute to persistence of this virus in environment. Therefore, it seems important to verify not only the susceptibility of other cyprinid or non-cyprinid species, but also their ability to transmit CyHV-3 infection to susceptible species. Our previous investigation of the susceptibility of the topmouth gudgeon (Pseudorasbora parva) did not reveal the presence of CyHV-3 DNA in the tissues of this species after cohabitation with infected koi. Consequently, we changed the experimental conditions and applied two stress factors (removal of skin mucus and scaring) which would presumably mimic the stress most commonly encountered in the wild. Both experiments (without and with stress factors) consisted of primary and secondary challenges. In both the no-stress and stress experiments, the first challenge was focused only on testing the susceptibility of the topmouth gudgeon to the virus. With the secondary challenge, we investigated potential viral transmission from the topmouth gudgeon to healthy naive koi after exposure to stress factors. All fish (dead, surviving and sacrificed) were tested for the presence of CyHV-3 DNA using nested PCR (no-stress experiment) and real-time PCR (stress experiment). After the primary challenge of the no-stress experiment, PCR did not reveal the presence of CyHV-3 DNA in any specimen of cohabitated topmouth gudgeon, but all specimens of dead koi were CyHV-3 DNA-positive. PCR of fish tissues subjected to the secondary challenge did not show the transfer of virus to naive fish. After exposure to stress (removal of skin mucus), qPCR revealed four out of five samples (80%) of topmouth gudgeon to be positive for CyHV-3 DNA. Two out of five samples (40%) of topmouth gudgeon treated by scaring were found to be positive for the presence of viral DNA. Real-time PCR after the secondary challenge did not reveal any viral DNA positivity in specimens of topmouth gudgeon from groups previously exposed to stress. The stress experiments show that removal of skin mucus might potentially lead to susceptibility of topmouth gudgeon to CyHV-3 infection, but the transmission of the virus to koi carp was not observed.

Evaluation of an antiparasitic compound extracted from Polygonum cuspidatum against Ichthyophthirius multifiliis in grass carp

Ichthyophthirius multifiliis is a ciliated parasite that infests almost all freshwater fish species and causes great economic losses to the aquaculture industry. In this study, a compound with anti-I. multifiliis activity was isolated from Polygonum cuspidatum and identified as emodin. In vitro anti-I. multifiliis results showed that emodin at 1 mg/L killed all I. multifiliis theronts for 96.0 min, and at 0.5 mg/L or lower concentrations could not kill all I. multifiliis theronts, but could significantly reduce the infectivity of theronts after pretreatment with emodin at the low concentrations mentioned above for 2 h. Additionally, emodin at 1 mg/L and 2 mg/L completely terminated the reproduction of nonencysted and encysted tomonts, respectively. In vivo tests, emodin at 0.5 mg/L could cure infected grass carp and protect naive fish from I. multifiliis infection by continuous adding emodin for 10 days. The 96 h median lethal concentration (LC50) of emodin to grass carp was 3.15 mg/L, which were approximately 18 and 7 times the median effective concentration (EC50) of emodin for killing theronts (0.18 mg/L) and nonencysted tomonts (0.45 mg/L), respectively. On the basis of these results, emodin is an effective compound for the development of a new drug against I. multifiliis.

Antiparasitic efficacy of commercial curcumin against Ichthyophthirius multifiliis in grass carp (Ctenopharyngodon idellus)

Ichthyophthiriasis, caused by Ichthyophthirius multifiliis, is a prevalent parasitic disease in freshwater fish, it leads to a significant economic loss in fish farming industry. Following the ban of malachite green in usage of treating food fish, new antiparasitic agents for the treatment of ichthyophthiriasis is being studied. The aims of the present study were to investigate in vitro and in vivo antiparasitic efficacy of a commercial curcumin against I. multifiliis and evaluate its acute toxicity in grass carp. In vitro tests, curcumin killed all theronts at 1mg/L for 38.7min, caused 100.0% mortality of nonencysted tomonts at 8mg/L for 47.3min, and terminated the reproduction of encysted tomonts at 4mg/L for 16h, respectively. In vivo trials, curcumin at 4mg/L for 10days exposure eradicated all parasitic trophonts in situ, and protected both infected and naive fish from I. multifiliis infections. The results of in vitro and in vivo experiments proved curcumin to be effective against the parasite. The 96h median lethal concentration (LC50) of curcumin to grass carp (58.5mg/L) was about 146.3 times median effective concentration (EC50) of curcumin to theronts (0.4mg/L), and 18.9 times EC50 of curcumin to nonencysted tomonts (3.1mg/L). Therefore, the commercially available curcumin can be used as a potential lead compound for the development of commercial drugs against I. multifiliis.