Zebrafish Facilities Research Articles

A survey of water chemistry used in zebrafish facilities and their effects on early zebrafish development.

There are a variety of published standard methods and water chemistry recommendations for zebrafish ( Danio rerio) husbandry, but empirical evidence for their justification is often lacking, as is information on some variables that have important biological effects on fish. Importantly, these different recommendations could contribute to variability in results and fish welfare between or within institutions. Here we document the current range of water chemistry used by various research institutions around the world and report initial findings on their effects on the development and growth of zebrafish. Over 40 institutes responded to a survey that revealed a large variation in water chemistry used for zebrafish husbandry including differences in the set-points and acceptable ranges for temperature, pH and conductivity. In subsequent experiments, zebrafish ( D. rerio, WIK) embryos/larvae exposed to a large range of salt concentrations (50μM to 10mM Na + or 30 - 2500 μS/cm) and CO 2 levels (400 - 8,000 μatm). Larvae exposed to the lowest salt concentration (5 μM Na + or < 30μS/cm) had a slower response to touch and their swim bladders were not inflated. Larvae exposed to 5-100 μM Na + were 5 % shorter in total body length than those exposed to higher salt concentrations (>100 μM Na +). Zebrafish embryo/larvae exposed to intermediate pCO 2 values (~2000 μatm) were 1 to 3.5% longer than those exposed to either ambient (400 μatm) or higher (4000 μatm) pCO 2, but pCO 2 did not affect developmental endpoints up to 4 dpf. Overall, we highlight the magnitude of variation in water chemistry used within zebrafish research and provide some empirical evidence to show that not all of these water conditions might be optimal for developing zebrafish and reproducibility of research, although further research is necessary to determine longer-term effects of water chemistry on older larvae, juveniles and adults.

Review of Pseudoloma neurophilia (Microsporidia): A common neural parasite of laboratory zebrafish (Danio rerio).

Zebrafish (Danio rerio) is now the second most used animal model in biomedical research. As with other vertebrate models, underlying diseases and infections often impact research. Beyond mortality and morbidity, these conditions can compromise research end points by producing nonprotocol induced variation within experiments. Pseudoloma neurophilia, a microsporidium that targets the central nervous system, is the most frequently diagnosed pathogen in zebrafish facilities. The parasite undergoes direct, horizontal transmission within populations, and is also maternally transmitted with spores in ovarian fluid and occasionally within eggs. This transmission explains the wide distribution among research laboratories as new lines are generally introduced as embryos. The infection is chronic, and fish apparently never recover following the initial infection. However, most fish do not exhibit outward clinical signs. Histologically, the parasite occurs as aggregates of spores throughout the midbrain and spinal cord and extends to nerve roots. It often elicits meninxitis, myositis, and myodegeneration when it infects the muscle. There are currently no described therapies for the parasite, thus the infection is best avoided by screening with PCR-based tests and removal of infected fish from a facility. Examples of research impacts include reduced fecundity, behavioral changes, transcriptome alterations, and autofluorescent lesions.

A survey of water chemistry used in zebrafish facilities and their effects on early zebrafish development

Background There are a variety of published standard methods and water chemistry recommendations for zebrafish (Danio rerio) husbandry, but empirical evidence for their justification is often lacking, as is information on some variables that have important biological effects on fish. Importantly, these different recommendations could contribute to variability in results and fish welfare between or within institutions. Methods Here we document the current range of water chemistry used by various research institutions around the world and report initial findings on their effects on the development and growth of zebrafish. Over 40 institutes responded to a survey that revealed a large variation in water chemistry used for zebrafish husbandry including differences in the set-points and acceptable ranges for temperature, pH and conductivity. In subsequent experiments, zebrafish (D. rerio, WIK) embryos/larvae exposed to a large range of salt concentrations (50μM to 10mM Na+ or 30 – 2500 μS/cm) and CO2 levels (400 – 8,000 μatm). Results Larvae exposed to the lowest salt concentration (5 μM Na+ or &lt; 30μS/cm) had a slower response to touch and their swim bladders were not inflated. Larvae exposed to 5-100 μM Na+ were 5 % shorter in total body length than those exposed to higher salt concentrations (&gt;100 μM Na+). Zebrafish embryo/larvae exposed to intermediate pCO2 values (~2000 μatm) were 1 to 3.5% longer than those exposed to either ambient (400 μatm) or higher (4000 μatm) pCO2, but pCO2 did not affect developmental endpoints up to 4 dpf. Conclusions Overall, we highlight the magnitude of variation in water chemistry used within zebrafish research and provide some empirical evidence to show that not all of these water conditions might be optimal for developing zebrafish and reproducibility of research, although further research is necessary to determine longer-term effects of water chemistry on older larvae, juveniles and adults.

Disentangling the link between zebrafish diet, gut microbiome succession, and Mycobacterium chelonae infection

BackgroundDespite the long-established importance of zebrafish (Danio rerio) as a model organism and their increasing use in microbiome-targeted studies, relatively little is known about how husbandry practices involving diet impact the zebrafish gut microbiome. Given the microbiome’s important role in mediating host physiology and the potential for diet to drive variation in microbiome composition, we sought to clarify how three different dietary formulations that are commonly used in zebrafish facilities impact the gut microbiome. We compared the composition of gut microbiomes in approximately 60 AB line adult (129- and 214-day-old) zebrafish fed each diet throughout their lifespan.ResultsOur analysis finds that diet has a substantial impact on the composition of the gut microbiome in adult fish, and that diet also impacts the developmental variation in the gut microbiome. We further evaluated how 214-day-old fish microbiome compositions respond to exposure of a common laboratory pathogen, Mycobacterium chelonae, and whether these responses differ as a function of diet. Our analysis finds that diet determines the manner in which the zebrafish gut microbiome responds to M. chelonae exposure, especially for moderate and low abundance taxa. Moreover, histopathological analysis finds that male fish fed different diets are differentially infected by M. chelonae.ConclusionsOverall, our results indicate that diet drives the successional development of the gut microbiome as well as its sensitivity to exogenous exposure. Consequently, investigators should carefully consider the role of diet in their microbiome zebrafish investigations, especially when integrating results across studies that vary by diet.

Modeling of Zebrafish Housing Temperatures During Power Outage and Instrument Failure.

Power outages can happen anywhere and anytime for various reasons. This threat affects also scientific work of biologists. Especially problematic area is aquatic animal husbandry, where life support of the animals is dependent on continuous electricity supply and years of scientific work may depend on the well-being of these animal stocks. Therefore, tools to estimate and control these risks are needed. In this study, I have used modeling to estimate aquarium water temperature changes during power outages and constructed simplified models for zebrafish aquaria. A calculation worksheet is also provided to help to model kinetics of water temperature changes in zebrafish facilities.

Evaluation of Disinfection Methods for Artificial Plants in Zebrafish (Danio rerio) Recirculating Water Systems.

The use of artificial plants as environmental enrichment for zebrafish (Danio rerio) in biomedical research facilities has been shown to provide benefits in animal welfare and care. Despite the benefits of artificial plants to zebrafish welfare, some research facilities are hesitant to incorporate them into their routine husbandry practices due to concerns about disease transmission and a lack of guidance on effective disinfection practices between tanks. Limited published information is available on how to adequately disinfect artificial plants, which creates concerns regarding their reuse between tanks in recirculating water systems. Proper sanitation and disinfection of these items is crucial to preventing the spread of disease in the system. We evaluated 2 disinfection methods- a commercial-grade laboratory glassware dishwasher and an ethylene oxide (ETO) sterilizer-by using ATP detection and bacterial culture of the artificial plants before and after the disinfection process. Plants were placed in the dirty sump of 2 separate recirculating systems (2,500 to 3,000 fish per system) for 2 wk before the start of the study. High ATP levels and various bacterial organisms were detected prior to disinfection. The commercial-grade labo- ratory glassware dishwasher and ETO sterilizer both significantly reduced ATP levels and resulted in complete eradication of live bacteria that were present before treatment. This study demonstrates 2 effective methods for disinfecting artificial plants in zebrafish facilities.

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.

Timing matters: female receptivity and mate choice in the zebrafish (Danio rerio)

Female choice has been documented in many animal taxa, and how we test it has been refined through years of studies on the topic. However, when designing mate choice experiments some variables, surprisingly, often remain overlooked, including receptivity and reproductive stage. Here, we aimed to assess whether the female reproductive stage influences strength and direction of mate choice in the zebrafish, Danio rerio. Females were offered a choice between two males differing in body size. We found that female choice in our experimental setup was significantly repeatable and that females preferred larger males. Nonetheless, the level of choosiness of females was affected by the time since the last spawning. Females spent more time choosing when tested 7 and 10 days after spawning rather than 4 days, indicating a higher receptivity to males from one week after the last spawning. Moreover, females preferred larger males only when tested 7 and 10 days after spawning. Our results suggest that female mate choice should take female receptivity into account, by standardizing time since the last spawning across females. More broadly, this suggests that 7–10 days since the previous spawning is the ideal time interval for zebrafish female receptivity to peak, with implications for facilities and researchers to increase egg production in natural spawning events and manual egg collection.Significance statementThe role of pre-copulatory female mate preference has long been recognized in sexual selection. Nonetheless, female receptivity often remains overlooked in mate choice experiments especially in external fertilizing species. In the present study, we investigated if the female reproductive stage affects the strength and direction of female mate choice in an external fertilizing fish, the zebrafish, Danio rerio. We found that, when tested 7 and 10 days after spawning rather than 4 days, females spent more time choosing, demonstrating an increased receptivity to males from 1 week following the last spawning. Furthermore, only at 7 and 10 days after spawning females exibith a clear preference for the bigger males. Our study highlights the importance of considering the female receptivity in future studies assessing mate choices in this and other externally fertilizing species, and also for zebrafish facilities to increase egg production in natural spawning events and manual egg collection.

Rederivation of a mutant line (prop 1) of zebrafish Danio rerio infected with Pseudoloma neurophilia using in vitro fertilization with eggs from pathogen-free wild-type (AB) females and sperm from prop 1 males.

Along with the growing number of laboratories that work with zebrafish (Danio rerio), it is necessary to have animals with good sanitary quality. Specific pathogens can interfere with the experimental results and in the life quality of the animals. Pseudoloma neurophilia is a parasite with high potential for interference in behavioural, morphology, toxicological and genetic research, and is very common in zebrafish facilities. With that, we implemented a protocol for the pathogen elimination in a genetically modified lineage (prop 1) using eggs from specific pathogen-free (SPF) wild-type fish (AB line) for in vitro fertilization, along with water recirculation equipment disinfection, appropriate PCR screening and back crossing protocols. This resulted in SPF prop 1 heterozygotes, which allowed us to move forward with subsequent crossings to develop homozygote prop 1 mutants for our research. Hence, this demonstrates a useful strategy for an individual research laboratory to rederive a specific mutant free line that is not available from other SPF laboratories.

A Systematic Analysis of Metal and Metalloid Concentrations in Eight Zebrafish Recirculating Water Systems.

Metals and metalloids are integral to biological processes and play key roles in physiology and metabolism. Nonetheless, overexposure to some metals or lack of others can lead to serious health consequences. In this study, eight zebrafish facilities collaborated to generate a multielement analysis of their centralized recirculating water systems. We report a first set of average concentrations for 46 elements detected in zebrafish facilities. Our results help to establish an initial baseline for trouble-shooting purposes, and in general for safe ranges of metal concentrations in recirculating water systems, supporting reproducible scientific research outcomes with zebrafish.

Pseudocapillaria tomentosa, Mycoplasma spp., and Intestinal Lesions in Experimentally Infected Zebrafish Danio rerio.

Intestinal neoplasms and preneoplastic lesions are common in zebrafish research facilities. Previous studies have demonstrated that these neoplasms are caused by a transmissible agent, and two candidate agents have been implicated: a Mycoplasma sp. related to Mycoplasma penetrans and the intestinal parasitic nematode, Pseudocapillaria tomentosa, and both agents are common in zebrafish facilities. To elucidate the role of these two agents in the occurrence and severity of neoplasia and other intestinal lesions, we conducted two experimental inoculation studies. Exposed fish were examined at various time points over an 8-month period for intestinal histopathologic changes and the burden of Mycoplasma and nematodes. Fish exposed to Mycoplasma sp. isolated from zebrafish were associated with preneoplastic lesions. Fish exposed to the nematode alone or with the Mycoplasma isolate developed severe lesions and neoplasms. Both inflammation and neoplasm scores were associated with an increase in Mycoplasma burden. These results support the conclusions that P. tomentosa is a strong promoter of intestinal neoplasms in zebrafish and that Mycoplasma alone can also cause intestinal lesions and accelerate cancer development in the context of nematode infection.

Reviewing Pseudoloma neurophilia infections in the popular zebrafish model

AbstractZebrafish is a worldwide used animal model to better understand cultured fish species; thus, in the last two decades, zebrafish facilities have been created in many aquaculture research centres. However, and despite the attempts of rigorous aseptic conditions for animal husbandry in wet laboratory facilities, the presence of parasites cannot be underestimated. The microsporidium Pseudoloma neurophilia, which mostly affects the central nervous system, has been listed as the most prevalent parasite in reared zebrafish. The emergence of P. neurophilia is usually detected when the individual presents irreversible morphological and/or behavioural symptoms, and consequently, the spread of the disease is inevitable, indicating that the infection is expanded across rearing tanks. This review attempts, for the first time, to collect and discuss the current information on P. neurophilia life cycle, target tissues and symptomatology of the infection occurred in zebrafish. Due to its vertical transmission together with the increasing presence of zebrafish as a model in reproduction‐related problems found in aquaculture, it is highlighted, among others, the sexual dimorphism of the pathogenic effects and the possible transgenerational implications of parasitic infections. Further, to date none effective treatments have been described, so here we provide in detail available diagnostic methods, discussing the importance of the early detection, giving safety‐rearing recommendations to decrease the presence of P. neurophilia. Our intention is to minimize parasite outbreaks in zebrafish facilities, which, on the one hand, can compromise zebrafish welfare, and on the other hand, could interfere furtively in the resulting research previously designed for improving aquaculture production but also other biomedical‐related research.

The importance of pH: How aquarium water is affecting behavioural responses to drug exposure in larval zebrafish

There has been rapid growth in the use of larval zebrafish as a complementary vertebrate model for drug discovery, abuse liability and pharmacological toxicology, resulting in a huge increase in zebrafish facilities worldwide. However, many research groups working with zebrafish do not typically report the pH of husbandry conditions in methodologies, nor are the pH of drug treatments reported in many research articles. This unknown factor can be a major contributor in the differential effects of drug treatments. Therefore, as a case study, we tested the impact of altering pH of several drugs of abuse and assessed locomotor changes associated with a single drug concentration delivered at different pHs. We found that a change of a single pH unit, within the pH ranges commonly used in zebrafish husbandry, was enough to alter locomotor activity at a fixed drug concentration. Many pharmacological agents are dependent on environmental factors, such as pH, to determine bioavailability. Efficaciousness for many classes of drug is dependent on their ionization state in which shifts towards uncharged species can influence the easy of a drug crossing biological membranes. Thus, we urge users to report pH in husbandry methods and drug treatments to improve replicability and inter-study comparisons.

Behavioural effects of the common brain-infecting parasite Pseudoloma neurophilia in laboratory zebrafish (Danio rerio)

Research conducted on model organisms may be biased due to undetected pathogen infections. Recently, screening studies discovered high prevalence of the microsporidium Pseudoloma neurophilia in zebrafish (Danio rerio) facilities. This spore-forming unicellular parasite aggregates in brain regions associated with motor function and anxiety, and despite its high occurrence little is known about how sub-clinical infection affects behaviour. Here, we assessed how P. neurophilia infection alters the zebrafish´s response to four commonly used neurobehavioral tests, namely: mirror biting, open field, light/dark preference and social preference, used to quantify aggression, exploration, anxiety, and sociability. Although sociability and aggression remained unaltered, infected hosts exhibited reduced activity, elevated rates of freezing behaviour, and sex-specific effects on exploration. These results indicate that caution is warranted in the interpretation of zebrafish behaviour, particularly since in most cases infection status is unknown. This highlights the importance of comprehensive monitoring procedures to detect sub-clinical infections in laboratory animals.

Zebrafish: Housing and husbandry recommendations.

This article provides recommendations for the care of laboratory zebrafish (Danio rerio) as part of the further implementation of Annex A to the European Convention on the protection of vertebrate animals used for experimental and other scientific purposes, EU Commission Recommendation 2007/526/EC and the fulfilment of Article 33 of EU Directive 2010/63, both concerning the housing and care of experimental animals. The recommendations provide guidance on best practices and ranges of husbandry parameters within which zebrafish welfare, as well as reproducibility of experimental procedures, are assured.Husbandry procedures found today in zebrafish facilities are numerous. While the vast majority of these practices are perfectly acceptable in terms of zebrafish physiology and welfare, the reproducibility of experimental results could be improved by further standardisation of husbandry procedures and exchange of husbandry information between laboratories. Standardisation protocols providing ranges of husbandry parameters are likely to be more successful and appropriate than the implementation of a set of fixed guidance values neglecting the empirically successful daily routines of many facilities and will better reflect the wide range of environmental parameters that characterise the natural habitats occupied by zebrafish.A joint working group on zebrafish housing and husbandry recommendations, with members of the European Society for Fish Models in Biology and Medicine (EUFishBioMed) and of the Federation of European Laboratory Animal Science Associations (FELASA) has been given a mandate to provide guidelines based on a FELASA list of parameters, ‘Terms of Reference’.

Viability of Pseudocapillaria tomentosa Eggs Exposed to Heat, Ultraviolet Light, Chlorine, Iodine, and Desiccation

Pseudocapillaria tomentosa is an important pathogen in zebrafish facilities. We investigated heat, ultraviolet (UV) light, chlorine, iodine, and dessciation for killing the parasite's eggs. Eggs released with feces larvate in about 5-10 days, and treatments were evaluated by exposing fresh eggs and subsequently comparing larvation to untreated eggs as an indication of survival. Collectively, untreated eggs in all trials showed high levels of survival. Eggs were exposed to elevated temperatures (40°C, 45°C and 50°C) for 1, 8, or 24 h, which resulted in substantial reduction in viability of eggs. UV radiation was effective, with no larvation at 50-300 mWs/cm2 and <2% at 20 mWs/cm2. Three chlorine products (JT Baker, Clorox®, and Bi-Mart) were tested at 25, 50, 100, 500, and 3,000 ppm (pH 7.0-7.3) with 10 min exposure. All were effective at 500 or 1,000 ppm. There was variability between three products and trials at lower concentrations, but overall chlorine was not very effective at 25-100 ppm except for Bi-Mart brand at 100 ppm. Povidone-iodine was not effective at 25 or 50 ppm for 10 min, but was effective at 200 ppm for 1 h. Desiccation was effective, and no eggs larvated after 2 h drying.

Overview of a Disease Outbreak and Introduction of a Step-by-Step Protocol for the Eradication of Mycobacterium haemophilum in a Zebrafish System.

In 2017, the zebrafish unit at University of Glasgow experienced a detrimental outbreak of pathogenic bacterium, Mycobacterium haemophilum. The presence of other bacterial species was also confirmed by bacteriology growth in the same unit. The affected individuals composed of a wild-origin parental population sourced from India and their F1 offspring generation. Bacteria were diagnostically confirmed to be present systemically in fish and within the water and biofilm of the recirculating zebrafish system. In the absence of a publicly accessible step-by-step disinfectant protocol for these difficult-to-eliminate pathogens, we devised a successful procedure to eradicate mycobacteria and Aeromonas species after colony removal using Cleanline Chlorine tablets (active ingredient Sodium dichloroisocyanurate) and Virkon Aquatic®. Postdisinfection diagnostics did not detect pathogens in the system or in the new fish inhabiting the system that were tested. Newly established fish colonies have not shown similar clinical signs or disease-induced mortality in the 1-year period following system disinfection and repopulation. We present a historical background of the bacterial outbreak and a disinfection method which can be replicated in other zebrafish facilities—at small or large scales—for reliable mycobacterium removal. This procedure can be implemented as a disinfection protocol before the introduction of a new fish population to a previously contaminated system.

A longitudinal assessment of host-microbe-parasite interactions resolves the zebrafish gut microbiome\u2019s link to Pseudocapillaria tomentosa infection and pathology

BackgroundHelminth parasites represent a significant threat to the health of human and animal populations, and there is a growing need for tools to treat, diagnose, and prevent these infections. Recent work has turned to the gut microbiome as a utilitarian agent in this regard; components of the microbiome may interact with parasites to influence their success in the gut, meaning that the microbiome may encode new anthelmintic drugs. Moreover, parasite infections may restructure the microbiome’s composition in consistent ways, implying that the microbiome may be useful for diagnosing infection. The innovation of these utilities requires foundational knowledge about how parasitic infection, as well as its ultimate success in the gut and impact on the host, relates to the gut microbiome. In particular, we currently possess limited insight into how the microbiome, host pathology, and parasite burden covary during infection. Identifying interactions between these parameters may uncover novel putative methods of disrupting parasite success.ResultsTo identify interactions between parasite success and the microbiome, we quantified longitudinal associations between an intestinal helminth of zebrafish, Pseudocapillaria tomentosa, and the gut microbiome in 210 4-month-old 5D line zebrafish. Parasite burden and parasite-associated pathology varied in severity throughout the experiment in parasite-exposed fish, with intestinal pathologic changes becoming severe at late time points. Parasite exposure, burden, and intestinal lesions were correlated with gut microbial diversity. Robust generalized linear regression identified several individual taxa whose abundance predicted parasite burden, suggesting that gut microbiota may influence P. tomentosa success. Numerous associations between taxon abundance, burden, and gut pathologic changes were also observed, indicating that the magnitude of microbiome disruption during infection varies with infection severity. Finally, a random forest classifier accurately predicted a fish’s exposure to the parasite based on the abundance of gut phylotypes, which underscores the potential for using the gut microbiome to diagnose intestinal parasite infection.ConclusionsThese experiments demonstrate that P. tomentosa infection disrupts zebrafish gut microbiome composition and identifies potential interactions between the gut microbiota and parasite success. The microbiome may also provide a diagnostic that would enable non-destructive passive sampling for P. tomentosa and other intestinal pathogens in zebrafish facilities.

Improved protocol for rapid zebrafish growth without reducing reproductive performance

Despite widespread use of zebrafish Danio rerio (Hamilton) as a model organism for finfish aquaculture, genetics and molecular biology, husbandry methods and nutrition have not been standardised. We characterise a protocol for rearing zebrafish using a recently developed combination of environmental parameters that yield rapid growth rates: high densities of live food organisms (marine rotifers and brine shrimp nauplii), low salinity, continuous light and algal turbidity during larval and early juvenile stages (5–21 days post fertilisation, dpf) followed by a transition to conventional rearing methods (dark/light photoperiod, freshwater). We assessed a range of larval fish densities in a stagnant water system with continuous ad libitum feeding until 21 dpf and periodic water changes. Fish stocking densities of 12–22 fish/L gave maximum growth rate and growth rate declined as stocking density increased past this threshold. Survival rates during 5–21 and 21–42 dpf were 87 ± 9 and 98 ± 3% respectively. Mean ± SD weight and length were 64 ± 9 mg and 19.4 ± 1.0 mm at 21 dpf and 245 ± 28 mg and 30.4 ± 1.5 mm at 42 dpf. Fertility was assessed at 66 ± 3 dpf. Mean clutch size was 232 ± 124 oocytes/female and fertilisation rate was 72 ± 16%. This protocol will significantly increase productivity in zebrafish facilities.

Pseudocapillaria tomentosa in laboratory zebrafish Danio rerio: patterns of infection and dose response.

Parasites in wild populations almost always exhibit aggregation (overdispersion), in which relatively few hosts are infected with high numbers of the parasites. This pattern of infection has also been observed in laboratory studies, where many of the sources of natural variation are removed. Pseudocapillaria tomentosa (Nematoda) is common in zebrafish (Danio rerio) facilities. We describe here patterns of infections in zebrafish experimentally infected with larvated P. tomentosa eggs in various trials with defined numbers of eggs. One trial with eggs delivered in a gelatin diet is also included. Fish were exposed at 25, 75, and 200 eggs fish-1, and the minimal infectious dose was estimated to be 1.5 eggs fish-1. The ID50 (50% infective dose) was calculated to be 17.5 eggs fish-1. We also included data from a trial and 2 previously published experiments with undefined doses in which zebrafish were exposed to infectious water and detritus from a tank that previously contained infected fish. All doses resulted in a high prevalence of infection (>70%), except at the 25 eggs fish-1 dose, where the prevalence was 43-46%. Mean abundance of worms corresponded to dose, from 0.57 worms fish-1 at 25 eggs fish-1 to 7 worms fish-1 at 200 eggs fish-1. Variance to mean ratios (V/M) and the k parameters showed aggregation across the 8 separate trials, including the gelatin diet. Aggregation increased with increased parasite abundance. Given the consistent observation of aggregation across our experiments, the zebrafish/P. tomentosa system provides a potentially robust, high-throughput model to investigate factors that influence differences in host susceptibility within defined populations.