Zebrafish Populations Research Articles

Multigenerational effects of alcohol: A behavioral study in three zebrafish populations

Fetal alcohol exposure can result in fetal alcohol spectrum disorder (FASD), which encompasses a range of cognitive and behavioral impairments. Although zebrafish have been used as a reliable model to study FASD, little is known about the ontogeny of this disorder and population differences in subsequent generations not directly exposed to alcohol. In this study, we evaluated the behavioral outcomes of zebrafish populations AB, Outbred (OB), and Tubingen (TU), offspring of parents exposed to alcohol during embryonic development. The offspring of adult fish with FASD (exposed to 1 % alcohol at the embryonic stage) was compared to the offspring of unexposed parental fish (0 % alcohol at the embryo phase). The behavioral profile of the offspring was assessed at 6 days post-fertilization (dpf) and 45 dpf. At 6dpf, the AB FASD offspring exhibited hyperactivity and increased time at the edge of the tank, while the TU and OB FASD offspring showed hypoactivity. At 45dpf, TU fish maintained the larval locomotor pattern, characterized by decreased average speed and total distance traveled and increased immobility. However, AB and OB fish did not show alterations in locomotor activity and anxiety-related responses at 45dpf. Our results demonstrate, for the first time, that FASD zebrafish offspring display behavioral differences, which were most evident during the early ontogenetic phase (6dpf) but may vary throughout animal ontogeny. TU fish exhibited the most consistent behavioral pattern across different developmental stages. These findings provide insights into the multigenerational and persistent behavioral consequences of embryonic alcohol exposure in zebrafish. Further research should focus on other features that can be inherited and the development of treatments for the offspring affected by it.

Developmental Delay and Male-Biased Sex Ratio in esr2b Knockout Zebrafish.

The estrogen receptor signaling pathway plays an important role in vertebrate embryonic development and sexual differentiation. There are four major estrogen receptors in zebrafish: esr1, esr2a, esr2b and gper. However, the specific role of different estrogen receptors in zebrafish is not clear. To investigate the role of esr2b in zebrafish development and reproduction, this study utilized TALENs technology to generate an esr2b knockout homozygous zebrafish line. The number of eggs laid by esr2b knockout female zebrafish did not differ significantly from that of wild zebrafish. The embryonic development process of wild-type and esr2b knockout zebrafish was observed, revealing a significant developmental delay in the esr2b knockout zebrafish. Additionally, mortality rates were significantly higher in esr2b knockout zebrafish than in their wild-type counterparts at 24 hpf. The reciprocal cross experiment between esr2b knockout zebrafish and wild-type zebrafish revealed that the absence of esr2b resulted in a decline in the quality of zebrafish oocytes, while having no impact on sperm cells. The knockout of esr2b also led to an abnormal sex ratio in the adult zebrafish population, with a female-to-male ratio of approximately 1:7. The quantitative PCR (qPCR) and in situ hybridization results demonstrated a significant downregulation of cyp19ab1b expression in esr2b knockout embryos compared to wild-type embryos throughout development (at 2 dpf, 3 dpf and 4 dpf). Additionally, the estrogen-mediated induction expression of cyp19ab1b was attenuated, while the estradiol-induced upregulated expression of vtg1 was disrupted. These results suggest that esr2b is involved in regulating zebrafish oocyte development and sex differentiation.

A functional schizophrenia-associated genetic variant near the TSNARE1 and ADGRB1 genes

Recent collaborative genome wide association studies (GWAS) have identified >200 independent loci contributing to risk for schizophrenia (SCZ). The genes closest to these loci have diverse functions, supporting the potential involvement of multiple relevant biological processes; yet there is no direct evidence that individual variants are functional or directly linked to specific genes. Nevertheless, overlap with certain epigenetic marks suggest that most GWAS-implicated variants are regulatory. Based on the strength of association with SCZ and the presence of regulatory epigenetic marks, we chose one such variant near TSNARE1 and ADGRB1, rs4129585, to test for functional potential and assay differences that may drive the pathogenicity of the risk allele. We observed that the variant-containing sequence drives reporter expression in relevant neuronal populations in zebrafish. Next, we introduced each allele into human induced pluripotent cells and differentiated 4 isogenic clones homozygous for the risk allele and 5 clones homozygous for the non-risk allele into neural precursor cells. Employing RNA-seq, we found that the two alleles yield significant transcriptional differences in the expression of 109 genes at FDR <0.05 and 259 genes at FDR <0.1. We demonstrate that these genes are highly interconnected in pathways enriched for synaptic proteins, axon guidance, and regulation of synapse assembly. Exploration of genes near rs4129585 suggests that this variant does not regulate TSNARE1 transcripts, as previously thought, but may regulate the neighboring ADGRB1, a regulator of synaptogenesis. Our results suggest that rs4129585 is a functional common variant that functions in specific pathways likely involved in SCZ risk.

Selective Acoustic Trapping, Translating, Rotating, and Orienting of Organism From Heterogeneous Mixture.

Selective contactless manipulation of organisms with intrinsic mobility from heterogeneous mixture is essential for biomedical engineering and microbiology. Acoustic manipulation, compared to its optical, magnetic, and electrostatic counterparts, provides superior bio-compatibility and additive-free properties. In this study, we present an acoustic manipulation system capable of selectively trapping, translating, rotating, and orienting individual organisms from in-Petri dish organism mixture using a phased transducer array and microscope, by dynamically steering the acoustic field. Specifically, using brine shrimp and zebrafish populations as example, the to-be-manipulated organisms with different sizes or morphologies can be manually designated by the user in microscopic image and interactively localized. Thereafter, the selected organisms can be automatically trapped from the heterogeneous mixture using a multiple focal point-based acoustic field steering method. Finally, the trapped organisms can be translated, rotated, and oriented in regard to the user's distinct manipulation objectives in instant response. In different tasks, closed-loop positioning and real-time motion planning control are performed, highlighting the innovation in terms of automation and accuracy of our manipulation technique. The results demonstrate that our acoustic manipulation system and acoustic field steering method enable selective, stable, precision, real-time, and in-Petri dish manipulation of organisms from heterogeneous mixture.

Exploring the Importance of Environmental Complexity for Newly Hatched Zebrafish.

The effects of an early impoverished social or physical environment on vertebrate neural development and cognition has been known for decades. While existing studies have focused on the long-term effects, measuring adult cognitive phenotypes, studies on the effects of environmental complexity on the early stages of development are lacking. Zebrafish (Danio rerio) hatchlings are assumed to have minimal interaction with their environment and are routinely reared in small, bare containers. To investigate the effects of being raised under such conditions on development of behaviour and cognition, hatchlings housed for 10 days in either an enriched or a standard environment underwent two cognitive tasks. The results were mixed. Subjects of the two treatments did not differ in performance when required to discriminate two areas. Conversely, we found a significant effect in a number discrimination task, with subjects from impoverished condition performing significantly worse. In both experiments, larvae reared in impoverished environment showed a reduced locomotor activity. Given the effects that enrichment appears to exert on larvae, a third experiment explored whether hatchlings exhibit a spontaneous preference for more complex environments. When offered a choice between a bare setting and one with objects of different shapes and colors, larvae spent over 70% of time in the enriched sector. Deepening these effects of an early impoverished environment on cognitive development is crucial for the welfare of captive zebrafish populations and for enhancing the quality and reliability of studies on larval zebrafish.

Innovative high fat diet establishes a novel zebrafish model for the study of visceral obesity

Obesity is a complex chronic condition associated with multiple health risks, including visceral obesity, which is particularly detrimental. To gain insight into the mechanisms underlying obesity and its associated pathologies, a novel zebrafish model was established using an innovative high-fat diet (HFD). The primary goal was to induce visceral obesity in zebrafish and study the associated structural changes. To achieve this, a unique HFD consisting of 40% beef fat (HFD40) was developed and supplemented with magnesium aluminometasilicate to improve stability in a high humidity environment. Feeding regimens were initiated for both juvenile (starting at 2 weeks post-fertilization, lasting 18 weeks) and adult zebrafish (3 months post-fertilization, 8 weeks feeding duration). The innovative dietary approach successfully induced visceral obesity in both juvenile and adult zebrafish. This new model provides a valuable tool to study obesity-related pathologies, metabolic syndrome, and potential therapeutic interventions. Most importantly, the low-cost and easy-to-prepare composition of HFD40 was seamlessly incorporated into the water without the need for separation, was readily absorbed by the fish and induced rapid weight gain in the zebrafish population. In conclusion, this study presents a novel HFD40 composition enriched with a high beef fat concentration (40%), which represents a significant advance in the development of an experimental zebrafish model for the study of visceral obesity and associated metabolic changes.

Systems biology approaches to understand gene specification networks in the zebrafish retina

The ontogeny of the vertebrate retina has been a topic of interest to developmental biologists and human geneticists for many decades. Understanding the unfolding of the genetic program that transforms a field of progenitors' cells into a functionally complex and multi‐layered sensory organ is a formidable challenge. Although classical genetic studies succeeded in identifying the key regulators of retina specification, we are far from understanding the architecture of their gene regulatory networks (GRNs) and thus from predicting their behaviour. A goal in our group during the last years has been to analyse the bifurcation of the GRNs specifying the neural retina and RPE domains, as cells in these two compartments acquire their distinct shape; within a few hours window in zebrafish. We have characterized transcriptome dynamics and chromatin accessibility in segregating NR/RPE populations in zebrafish to identify active cis‐regulatory modules and enriched TF motives within them (Buono et al Nat com 2021). We have shown extensive network redundancy and context‐dependent activity for the core TFs within each domain. In line with this finding, we have shown that the simultaneous mutation of the key specifiers vsx1 and vsx2 by CRISPR/Cas9, which results in severe visual impairment and bipolar cells depletion, does not interfere with neural retina specification. We concluded that although important cis‐regulatory remodelling (ATAC‐seq) occurs in vsxKO retinas during early specification, this has little impact at a transcriptomic level (RNA‐seq). Our observations point to genetic redundancy as an important mechanism sustaining the integrity of the retinal specification network (Letelier &amp; Buono et al Biorxiv 2022). We are currently complementing previous efforts by applying single‐cell methods to the analysis of neural retina and RPE networks. We have generated transcriptome and epigenome (chromium 10x multiome scRNAseq + scATACseq) datasets in zebrafish to obtain predictive information for the identification of novel genes and cis‐regulatory elements involved in the specification of retinal domains. These datasets are a valuable resource to reconstruct the final architecture of the retinal GRNs.

Vimentin Localization in the Zebrafish Oral Cavity: A Potential Role in Taste Buds Regeneration.

The morphology of the oral cavity of fish is related to their feeding habits. In this context, taste buds are studied for their ability to catch chemical stimuli and their cell renewal capacity. Vimentin RV202 is a protein employed as a marker for mesenchymal cells that can differentiate along different lineages and to self-renew, while Calretinin N-18 is employed as a marker of sensory cells, and ubiquitin is a protein crucial for guiding the fate of stem cells throughout development. In this study, a surface morphology investigation and an immunohistochemical analysis have been conducted. The results of the present study reveal, for the first time, the presence of Vimentin RV202 in a taste bud cell population of zebrafish. Some taste bud cells are just Vimentin RV202-immunoreactive, while in other cells Vimentin RV202 and Calretinin N-18 colocalize. Some taste buds are just reactive to Calretinin N-18. Vimentin RV202-immunoreactive cells have been observed in the connective layer and in the basal portion of the taste buds. The immunoreactivity of ubiquitin was restricted to sensory cells. Further studies are needed to elucidate the role of Vimentin RV202 in the maturation of taste bud cells, its potential involvement in the regeneration of these chemosensory organs, and its eventual synergic work with ubiquitin.

Sex-specific DNA methylation and transcription of zbtb38 and effects of gene–environment interactions on its natural antisense transcript in zebrafish

ABSTRACT There is increasing evidence for the involvement of epigenetics in sex determination, maintenance, and plasticity, from plants to humans. In our previous work, we reported a transgenerational feminization of a zebrafish population for which the first generation was exposed to cadmium, a metal with endocrine disrupting effects. In this study, starting from the previously performed whole methylome analysis, we focused on the zbtb38 gene and hypothesized that it could be involved in sex differentiation and Cd-induced offspring feminization. We observed sex-specific patterns of both DNA methylation and RNA transcription levels of zbtb38. We also discovered that the non-coding exon 3 of zbtb38 encodes for a natural antisense transcript (NAT). The activity of this NAT was found to be influenced by both genetic and environmental factors. Furthermore, increasing transcription levels of this NAT in parental gametes was highly correlated with offspring sex ratios. Since zbtb38 itself encodes for a transcription factor that binds methylated DNA, our results support a non-negligible role of zbtb38 not only in orchestrating the sex-specific transcriptome (i.e., sex differentiation) but also, via its NAT, offspring sex ratios.

Assessing the exploratory profile of two zebrafish populations: Influence of anxiety-like phenotypes and independent trials on homebase-related parameters and exploration

Anxiety is a protective behavior when animals face aversive conditions. The open field test (OFT) is used to assess the spatio-temporal dynamics of exploration, in which both homebase formation and recognition of environmental cues may reflect habituation to unfamiliar conditions. Because emotional- and affective-like states influence exploration patterns and mnemonic aspects, we aimed to verify whether the exploratory behaviors of two zebrafish populations showing distinct baselines of anxiety differ in two OFT sessions. Firstly, we assessed the baseline anxiety-like responses of short fin (SF) and leopard (LEO) populations using the novel tank test (NTT) and light-dark test (LDT) in 6-min trials. Fish were later tested in two consecutive days in the OFT, where the spatial occupancy and exploratory profile were analyzed for 30 min. In general, LEO showed pronounced diving behavior and scototaxis in the NTT and LDT, respectively, in which an “anxiety index” corroborated their exacerbated anxiety-like behavior. In the OFT, the SF population spent less time to establish the homebase in the 1st trial, while only LEO showed a markedly reduction in the latency to homebase formation in the 2nd trial. Both locomotion and homebase-related activities were decreased in the 2nd trial, in which animals also revealed increased occupancy in the center area of the apparatus. Moreover, we verified a significant percentage of homebase conservation for both populations, while only SF showed reduced the number of trips and increased the average length of trips. Principal component analyses revealed that distinct factors accounted for total variances between trials for each population tested. While homebase exploration was reduced in the 2nd trial for SF, an increased occupancy in the center area and hypolocomotion were the main factors that contribute to the effects observed in LEO during re-exposure to the OFT. In conclusion, our novel data support the homebase conservation in zebrafish subjected to independent OFT sessions, as well as corroborate a population-dependent effect on specific behavioral parameters related to exploration.

Correlations begin at home: drivers of co-occurrence patterns in personality and cognitive ability in wild populations of zebrafish.

Aquatic habitats are extremely dynamic, with constantly changing ecological factors, which has now been exacerbated due to human-induced rapid environmental change. In such variable environments, it becomes essential to understand how personality and cognition in organisms affect the adaptability of individuals to different habitat conditions. To test this, we studied how personality-related traits as well as cognitive ability differ between populations of wild-caught zebrafish (Danio rerio) from habitats that differed in various environmental factors. We measured emergence into a novel environment as an indicator of boldness, and performance in a spatial task inferred from feeding latencies in a maze over repeated trials to assess learning and memory, as an indicator of cognitive ability. We found that personality affects cognition and although bolder fish are better learners, they show poorer retention of memory across populations. Although personality and cognitive ability varied between habitats, the patterns of their correlations remained similar within each population. However, the individual traits (such as sex and size) that were drivers of personality and cognition differed between the habitats, suggesting that not only do behavioral traits vary between populations, but also the factors that are important in determining them. Personality and cognitive ability and the correlations between these traits determine how well an organism performs in its habitat, as well as how likely it is to find new habitats and adapt to them. Studying these across wild zebrafish populations helps predict performance efficiencies among individuals and also explains how fish adapt to extremely dynamic environments that can lead to variation in behavioral traits and correlations between them. This study not only sheds light on the drivers of interindividual variation and co-occurrence patterns of personality and cognition, but also individual and population factors that might have an effect on them.

Single-cell transcriptomic analysis reveals heterogeneity of the patterns of responsive genes and cell communications in liver cell populations of zebrafish exposed to polystyrene nanoplastics

Nanoplastics (NPs) are a group of emerging environmental pollutants with potential toxicity and health risk on biosystem and ecosystem. Great efforts have been devoted to describing the uptake, distribution, accumulation, and toxicity of NPs at various aquatic organisms; however, the heterogeneous response patterns in zebrafish (Danio rerio) liver cell populations caused by NP exposure have not yet been clarified. Investigation of the heterogeneous response patterns in zebrafish liver cell populations after NPs exposure provides us significances to explore the NP cytotoxicity. In this article, the heterogeneous response patterns in zebrafish liver cell populations after polystyrene (PS)-NPs exposure were studied. Significantly increased content of malondialdehyde and decreased levels of catalase and glutathione were observed, indicating the oxidative damage of zebrafish liver induced by PS-NPs exposure. Afterwards, the liver tissues were enzymatically dissociated and used for single-cell transcriptomic (scRNA-seq) analysis. Nine cell types were identified based on unsupervised cell cluster analysis followed by their marker genes. Hepatocytes were the cell type most impacted by PS-NP exposure, and heterogeneous response patterns of male and female hepatocytes were observed. The PPAR signaling pathway was up-regulated in hepatocytes from both male and female zebrafish. Lipid metabolism-related functions were altered more notably in male-derived hepatocytes, while female-derived hepatocytes were more sensitive to estrogen stimulus and mitochondria. Macrophages and lymphocytes were also highly responsive cell types, with specific immune pathways activated to suggest immune disruption after exposure. Oxidation-reduction process and immune response were significantly altered in macrophages, and oxidation-reduction process, ATP synthesis, and DNA binding were most altered in lymphocytes. Our study not only integrates scRNA-seq with toxicology effects to identify highly sensitive and specific populations of responding cells, revealing highly specialized interactions between parenchymal and non-parenchymal cells and expanding our current understanding of PS-NPs toxicity, but also highlights the importance of cellular heterogeneity in environmental toxicology.

Different housing conditions for zebrafish: What are the effects?

Zebrafish is a popular experimental model in several research areas but little is known about the effects of using different strains or housing conditions. Poor control of genetic background and housing conditions could affect experimental results and data reproducibility. Here we investigated the effects of two possible sources of variation on zebrafish behaviour: fish origin and environmental parameters (light intensity, water temperature and noise). Zebrafish behaviour was then examined using the ‘novel tank test’, one of the most common paradigms used to assess anxiety-like behaviours in zebrafish. Our results show that an increase in light intensity alters fish behaviour, particularly freezing duration and distance from the bottom of the tank, indicating increased anxiety. Swimming activity increased at the lowest temperature (25 °C). However, different levels of background noise did not cause any significant changes in behaviour. Differences were also found between zebrafish strains and populations: while the AB strain from laboratory 1 was minimally influenced by variation in holding conditions, the AB strain from laboratory 2 was highly affected by changes in temperature, light, and background noise. Our study shows that variation in strains and holding conditions can significantly influence the results of behavioural testing and should be carefully considered in the experimental design and properly reported to improve data interpretation and reproducibility.

Long-term behavioral alterations following embryonic alcohol exposure in three zebrafish populations

Fetal alcohol exposure may lead to a condition known as fetal alcohol spectrum disorder (FASD), which comprises a set of consequences, including cognitive and behavioral impairments. Although zebrafish has been applied as a reliable model for studying FASD, there is no approach to the disorder’s ontogeny and population differences. Here, we evaluated the behavioral outcomes of AB, Outbred (OB), and Tübingen (TU) zebrafish populations embryonically exposed to alcohol throughout the development to the adult stage. We exposed 24hpf eggs to 0 %, 0.5 %, or 1.0 % alcohol for 2 h. Fish were let grow and locomotor and anxiety-like behaviors were tested in a novel tank at larval - 6dpf, juvenile – 45dpf, and adult- 90dpf stages. At 6dpf, both AB and OB treated with 1.0 % alcohol showed hyperactivity, while 0.5 % and 1.0 % TU fish exhibited hypolocomotion. At 45dpf, AB and TU fish maintained the larval pattern of locomotion. At the adult stage – 90dpf, both AB and TU populations showed increased locomotor activity and anxiogenic responses, while the OB population did not show altered behavior. Our results show for the first time that zebrafish populations exhibit behavioral differences in response to embryonic alcohol exposure and that it varies along animals' ontogeny. AB fish showed the most consistent behavioral pattern through developmental stages, TU fish showed behavioral changes only in adulthood, and OB population showed high interindividual variability. These data reinforce that different populations of zebrafish are better adapted to translational studies, offering reliable results in contrast to domesticated OB populations obtained from farms, which exhibit more variable genomes.

The kisspeptin-GnIH signaling pathway in the role of zebrafish courtship and aggressive behavior induced by azoxystrobin

Azoxystrobin, a strobilurin widely used to control rice diseases, has raised concerns about possible adverse effects on aquatic ecosystems. At present, very little is known about the effects of azoxystrobin on courtship and aggressive behavior and the potential underlying mechanisms. In the present study, after exposing adult male and female zebrafish to worst-case scenario concentrations of azoxystrobin (0, 2 μg/L, 20 μg/L, and 200 μg/L) for 42 d, we observed a decrease in courtship behavior and an increase in aggressive behavior in both male and female zebrafish. In addition, to elucidate the molecular mechanism of the behavioral effects of azoxystrobin, we quantified the changes in the concentrations of kisspeptin, 5-HT, GnIH, and their corresponding receptor mRNA expression in the brain. The results showed that 200 μg/L azoxystrobin decreased the concentrations of kisspeptin and increased the concentration of GnIH in both male and female zebrafish brain. In addition, azoxystrobin also significantly reduced 5-HT concentration in female zebrafish brain. Further investigation revealed that altered courtship and aggressive behavior were associated with the expression levels of genes (kiss1, kiss2, gnrh3, gnrhr3, 5ht1a, and 5ht2a) involved in kisspeptin-GnIH signaling pathway. In conclusion, our study suggested that azoxystrobin may impair courtship and aggressive behavior in zebrafish by interfering with the kisspeptin-GnIH signaling pathway, which may have more profound effects on natural zebrafish populations.

Analyzing the transmission of SVCV among zebrafish populations by compartment mathematical modeling

There are many studies of freshwater pathogen microorganisms aiming to elucidate the mechanism of infection and the individual immune responses. However, few of these studies focus on the transmission of the pathogens among freshwater fish populations. In this work, we performed experiments by introducing the zebrafish (Danio rerio) infected with spring viremia of carp virus (SVCV) to conduct the infectious process among zebrafish populations. Based on the collected data in the transmission experiment with PCR testing and tissue histology, an SEIR (susceptible-exposed-infected-removed) compartment mathematical model was established to quantitively characterize the propagation of SVCV among fish populations. After deriving the basic reproduction number R0 by this SEIR model, the temperature and density effects were preliminarily investigated. It was found that at lower temperatures between 15 and 21 °C, the transmission rate increased. The culturing density also played a vital role in R0 at certain temperatures. By reducing the culturing density, the determined reproduction number at 18 °C is substantially reduced, implying an effective slowing down of the transmission at this temperature. But the reductions in R0 are less effective for 15 °C and 21 °C. In all, this study proposed a new approach to quantitively analyze the virus transmission in the freshwater aquaculture. By supplementing with the individual immune research tools, it can not only provide effective containment strategies to control epidemics for freshwater fisheries, but also be used to optimize the culturing conditions to avoid the outbreak of transmissible diseases.

Application of an eDNA assay for the detection of Pseudoloma neurophilia (Microsporidia) in zebrafish (Danio rerio) facilities

Environmental DNA (eDNA) water assays are beginning to be implemented for many important pathogens in confined aquaculture systems. Recirculating systems are rapidly being developed for fin fish aquaculture. Zebrafish (Danio rerio) are reared in these systems, and Pseudoloma neurophilia (Microsporidia) represents a serious challenge for zebrafish research facilities. Diagnosis of the pathogen has traditionally used histology or PCR of tissues with lethal sampling. However, with the development of a nonlethal assay to detect P. neurophilia in tank water, facilities will be able to integrate the assay into routine surveillance efforts to couple with their established protocols. Here, we first describe a modified protocol to extract and quantify parasite DNA from the environment for nonlethal detection of P. neurophilia in adult zebrafish populations. Using this modified assay, we then evaluated water samples from a longitudinal experimental infection study, targeting timepoints during initial infection. The parasite was detectable in the water immediately after initial exposure until week 4 post exposure (pe), when the parasite was undetectable until 7 weeks pe. After that time, the parasite was sporadically detected in the water for the 10-month study, likely correlating with the lifecycle of the parasite. Using water samples from the Zebrafish International Resource Center, we also validated the clinical relevance of the assay in a large zebrafish facility. The integration of this assay at ZIRC will significantly compliment surveillance and control efforts for the microsporidian parasite.

Toxicokinetic Modeling of Per- and Polyfluoroalkyl Substance Concentrations within Developing Zebrafish (Danio rerio) Populations.

Per- and polyfluoroalkyl substances (PFAS) are pervasive environmental contaminants, and their relative stability and high bioaccumulation potential create a challenging risk assessment problem. Zebrafish (Danio rerio) data, in principle, can be synthesized within a quantitative adverse outcome pathway (qAOP) framework to link molecular activity with individual or population level hazards. However, even as qAOP models are still in their infancy, there is a need to link internal dose and toxicity endpoints in a more rigorous way to further not only qAOP models but adverse outcome pathway frameworks in general. We address this problem by suggesting refinements to the current state of toxicokinetic modeling for the early development zebrafish exposed to PFAS up to 120 h post-fertilization. Our approach describes two key physiological transformation phenomena of the developing zebrafish: dynamic volume of an individual and dynamic hatching of a population. We then explore two different modeling strategies to describe the mass transfer, with one strategy relying on classical kinetic rates and the other incorporating mechanisms of membrane transport and adsorption/binding potential. Moving forward, we discuss the challenges of extending this model in both timeframe and chemical class, in conjunction with providing a conceptual framework for its integration with ongoing qAOP modeling efforts.

Wild fish beat the heat

Wild animals can adapt to a wide range of temperatures. The ability to perform well in a constantly changing environment is called physiological plasticity. Physiological plasticity allows ectothermic animals, such as fishes, to maintain their metabolism even when the water that the fish is in heats up or cools down wildly. Fishes living in laboratories are less likely to have experienced temperature changes, so will their metabolism still perform well if there is a big temperature shift in their tank? Or have they lost their physiological plasticity because they have only experienced steady temperatures for more than 150 generations? Can living in steady environments affect a fish's behavior, metabolism or gene expression compared with living in a more turbulent environment? To study these questions, Rachael Morgan and colleagues from the Norwegian University of Science and Technology and the University of Greenwich, UK, compared the physiological plasticity of two populations of zebrafish: a wild population collected from India and a population raised in the lab. They aimed to understand whether living in a stable environment can change a fish's ability to perform when there is a big change in temperature.The researchers collected 300 zebrafish from the two populations and exposed each group to a variety of temperatures (from 10 to 38°C) for 1 month. They then measured the fish's swimming ability, escape response, growth rate, metabolic rate and gene expression and compared the measurements of the two groups to learn whether the lab-raised group had more difficulty in adapting to large temperature shifts than the wild group because they have less physiological plasticity.Sure enough, the lab-raised fish were less able to adjust to temperature change than the fish that had come from India. The lab-raised zebrafish were slower swimmers than wild zebrafish at all temperatures, and when escaping, lab-raised zebrafish responded more slowly than wild zebrafish, especially when in colder temperatures. In addition, the lab-raised zebrafish grew faster than the wild zebrafish, likely because domesticated animals are selected for faster growth rates. When comparing metabolism between the groups, the researchers measured the range of temperatures at which the fish could reach their maximum metabolic rates. They found that lab-raised fish could only reach their maximum metabolic rate in water temperatures between 26 and 36°C, but wild fish could reach their maximum metabolic rate over a much broader temperature range (21–38°C). Gene expression was also different among the two groups. The lab fish had greater levels of proteins that deal with stress compared with the wild fish. Interestingly, very cold temperatures were especially difficult for both groups, which may suggest that physiological plasticity is limited at these extreme temperatures.This study tells us that lab domestication affects the ability of animals to adjust to changes in their environment across several biological levels, from behaviour to gene expression. With this knowledge, scientists should take into account that domestication can change animals’ responses to an experiment when choosing study animals. Morgan and colleagues also suggest that there may be a trade-off between high levels of physiological plasticity and growth rates and warn that domestication can quickly lead to a loss of physiological plasticity.

Genetic diversity of wild zebrafish Danio rerio populations available in Bangladesh

The morphometric and genetic variations of 4 wild populations of zebrafish ( Danio rerio ) in Bangladesh were studied using DNA barcoding and RAPD (Random Amplified Polymorphic DNA). The zebrafish populations were as ZMS (zebrafish from the Brahmaputra river, Mymensingh), ZFP (zebrafish from the Kumar river, Faridpur), ZKH (zebrafish from the Moyurriver, Khulna) and ZMG (zebrafish from the Aarial Beel, Munshiganj). The morphometric variations were insignificant to establish the genetic structure and diversity of these populations. Therefore, the genetic variation was analyzed with DNA barcoding and RAPD techniques. FishF1 and FishR1 primers were used for barcoding with amplification of Cytochrome Oxidase C subunit 1(COI) gene region and OPA03, OPA04 and OPF01 primers were used for RAPD analysis. In DNA barcoding analysis, the database provided species matches of 100% sequence similarity for these 4 wild fish populations with D. rerio . The overall sequence alignment similarity among these populations was 99.40%. The neighbor-joining phylogenetic tree was constructed from these COI barcode sequences also revealed genetic relationships and diversity among these populations. In RAPD analysis, a total number of 112 bands were found where 22 bands were polymorphic, indicated 19.64% polymorphism among these 4 wild populations. The percent polymorphic loci and genetic distance varied within a range of 14.00–26.67 and 0.2877–1.6094 respectively. Nei's genetic similarity among these 4 populations across all the primers ranged from 0.2000 to 0.7500. The Dendrogram based on Nei's genetic distance showed 3 clusters indicating the ZFP population is the maximum genetically diversified among them however; the ZMS and ZKH populations were in the same cluster. The knowledge of the molecular variation of wild zebrafish could be helpful for planning more effective and efficient managing strategies for this model species and also provide a significant contribution to zebrafish research all over the world.