Zebrafish最新文献

The skin mucus of teleost fish harbors a complex microbial community that is continually interacting with the aquatic environment. Despite zebrafish, Danio rerio, serving as a model organism in a myriad of research fields, very little is known about the composition and role of the skin mucus microbiome. The purpose of this study was to determine a simple sampling method for the skin mucus microbiome, identify prominent bacterial members, and compare its composition to the microbial community of the surrounding environment. Next-generation sequencing of the V3-V4 region of the 16S rRNA gene was performed on skin mucus and filtered tank water samples. Results show that prominent bacterial members of the skin mucus in zebrafish include Actinobacteria (Mycobacteriaceae) and Gammaproteobacteria (Aeromonadaceae), followed by Alphaproteobacteria and Betaproteobacteria. The tank water contained much higher bacterial diversity and was clearly different from the skin mucus microbiome, despite continuous interaction. This study identifies a straightforward sampling method for the zebrafish skin mucus microbiome, enabling hypothesis generation on the role of ectosymbionts on host and microbiome health.

Brain imaging requires mounting of zebrafish larvae in a vertical position, but anesthetized or fixed larvae tend to fall on their sides without external support. Current solution is to manually hold individual larva until liquid agarose solidifies, which is time consuming, labor intensive, and unfriendly to beginners. We developed a method to form larva-shaped slots in agarose gel using a computer numerical controlled manufactured mold. Each slot nearly perfectly fits a larva in its upright position, and larvae can be easily mounted by inserting into the slots. On average, each larva can be mounted in <1 min using this method.

Genetically encoded fluorescent tags such as green fluorescent protein fused to protein have revolutionized cell biology as they permit high-resolution protein imaging in live systems. Split fluorescent proteins, with a small fragment of 16 amino acids, can be inserted in the coding sequence to label proteins. We demonstrate successful integration of two bright and fast maturing split fluorescent proteins, mNeon green and sfCherry2, in zebrafish, and show that they are suitable for live imaging, including time-lapse series, and that they have a high signal-to-noise ratio. Furthermore, we show that CRISPR/Cas9 can be used to generate fluorescently tagged proteins in vivo.

Setting nutritional standards for larval zebrafish (Danio rerio) that maximize growth, survival, and reproductive success is challenging. We evaluated the effects of different feeding regimens on larval zebrafish by comparing Gemma Micro 75 pelleted diet and live-type L rotifers (Brachionus plicatilis) in 3 feeding regimens starting at 9 days postfertilization (dpf): bolus feeding of live diet (BL), continuous feeding of live diet (CL), and pelleted diet (PD). Animals in the PD and CL groups were longer than the BL group at 4-5 weeks postfertilization. The PD group was also greater in body depth than both live diet groups. There was no significant difference in weight between the groups. There were also no significant differences in fecundity or sex ratios indicating that all feeding methods successfully promote growth of a useful breeding stock of fish. In addition, we quantified the equipment, consumable, and labor costs associated with these methods, and found that the PD regimen was superior to both live diet regimens. These data suggest that providing a high nutrient-density pelleted diet to larval and juvenile zebrafish is an effective means to increase early growth and to decrease cost and labor associated with nursery care.

The use of proper anesthesia in zebrafish research is essential to ensure fish welfare and data reliability. However, anesthesia long-term side effects remain poorly understood. The purpose of this study was to assess anesthesia quality and recovery in adult zebrafish using different anesthetic protocols and to determine possible long-term effects on the fish activity and anxiety-like behaviors after anesthesia. Mixed-sex adult AB zebrafish were randomly assigned to five different groups (Control, 175 mg/L of tricaine methanesulfonate [MS222], 45 mg/L of clove oil, 2 mg/L of etomidate, and 5 mg/L of propofol combined with 150 mg/L of lidocaine) and placed in the respective anesthetic bath. Time to lose the equilibrium, response to touch and to caudal fin pinch stimuli, and recovery after anesthesia administration were evaluated. In addition, after stopping anesthesia, respiratory rate, activity, and anxiety-like behaviors in the novel tank test were studied. Overall, all protocols proved to be adequate for zebrafish anesthesia research as they showed full recovery at 1 h, and only etomidate had minor effects on fish behavior in the novel tank, a validated test for anxiety.

Animals communicate with each other through a variety of behavioral interactions, many of which are often complex due to the interplay of several ecological factors. Observations on dyadic interactions can help throw light on the more complex interactions observed among group living organisms and can help in understanding mechanisms of behaviors related to mating strategies, dominance hierarchies, and decision-making. This study focused on the assessment of several generally observed interactions among dyads of different sexes (female-female, male-male, and male-female) in wild zebrafish (Danio rerio). Temporal dynamics of these interactive behaviors were observed in 45 dyads across 3 time intervals of the day. We used generalized linear mixed models to investigate the effect of time, sex of dyad, and their interaction on specific behaviors. While the frequency of occurrence of some behaviors showed clear variation across time intervals of the day, these were further found to depend on the composition of the dyad. Contrary to previous reports, we found that same-sex dyads are equally aggressive and aggressive interactions did not vary temporally. Mating-associated interactions, as expected, were significantly higher in mixed-sex dyads and declined significantly from early morning to afternoon. Interestingly, we also found some mating-associated interactions in same-sex dyads. A fine line exists between social and mating-associated interactions in many organisms and so we speculate that these interactions could also be social interactions and not mating-related behavior. Our findings shed light on complex interactive behaviors among zebrafish, that are likely to be affected by time as well as sex composition of interacting individuals and thus has important implications for groups varying in sex ratios in the wild.

The zebrafish is a valuable model organism that is widely used in studies of vertebrate development. In the laboratory, zebrafish embryonic development is normally carried out at 28.5°C. In this study, we sought to determine whether it was possible to modify the speed of embryonic development through the use of short- and long-term variations in incubation temperature. After incubation at 20°C-32°C, most early-stage embryos survived to the epiboly stage, whereas more than half of the embryos died at <20°C or >32°C. The rate of development differed between embryos incubated at the lowest (18°C) and highest (34°C) temperatures: a difference of 60 min was observed at the 2-cell stage and 290 min at the 1k-cell stage. When blastulae that had developed at 28°C were transferred to a temperature lower than 18°C for one or more hours, they developed normally after being returned to the original 28°C. Analyses using green fluorescent protein-buckyball mRNA and in situ hybridization against vasa mRNA showed that primordial germ cells increase under low-temperature culture; this response may be of use for studies involving heterochronic germ cell transplantation. Our study shows that embryonic developmental speed can be slowed, which will be of value for performing time-consuming, complicated, and delicate microsurgical operations.

The zebrafish is becoming an increasingly popular research animal around the world. Its welfare is affected by an array of environmental factors, such as food access and water quality. Holding density is an important welfare aspect, not least due to its interaction with other housing conditions. Despite the extensive use of zebrafish in research, little is known of how densities affect its welfare. In this systematic review, we have performed a large literature search, compiled, and evaluated all publications regarding zebrafish holding density. We have analyzed how density effects growth, reproduction, and stress response, including behavior, water quality, and pathogenic outbreaks in young and adult fish. Our review shows that the holding densities tested vary largely depending on the research focus, for example, body growth or behavior. In fact, research indicates that future recommendations on holding density could depend on which welfare aspects are considered. Overall, there is a need for more studies investigating the interactive effects of density on welfare indicators, such as reproduction coupled with stress response. We stress the necessity of including holding density in universal housing guidelines and reporting information on holding conditions of larvae and adults when publishing zebrafish work.