An easy and inexpensive method for determining the rate of individual phoretic events of nematodes.

Considering their limited locomotory capabilities, the cosmopolitan distribution of free-living nematodes may rely on phoretic dispersal. We describe a new, inexpensive device to investigate individual phoretic events of the nematode Caenorhabditis elegans using the pomace flies Drosophila melanogaster and Drosophila hydei over short time periods. Using our device, we replicated previous findings demonstrating that phoresis requires C. elegans to be in the dauer stage and capable of nictation. Additionally, we find that phoresis can happen on the order of seconds, and does not increase linearly with time of interaction. Using this approach can facilitate the investigation of nematode biogeography, which could provide useful insight into their, and their vector's, control.

Learning Deficits and Attenuated Adaptive Stress Response After Early-Life Seizures in Zebrafish.

Early-life seizures (ELS) are often associated with the development of cognitive deficits. However, methods to predict and prevent these deficits are lacking. To increase the range of research models available to study cognitive consequences of ELS, we investigated whether seizures in larval zebrafish (Danio rerio) lead to behavioral deficits later in life. We thus modified the existing pentylenetetrazole (PTZ)-induced seizure model in larval zebrafish, exposing zebrafish to PTZ daily from 5 to 7 days post-fertilization (dpf). We then compared later-life learning, social behavior (shoaling), and behavioral and chemical measures of anxiety in the PTZ-exposed zebrafish (PTZ group) to that of naïve clutchmates (untouched controls, UC) and to a second control group (handling control, HC) that experienced the same handling as the PTZ group, but without PTZ exposure. We observed that only the PTZ group displayed a significant deficit in a y-maze learning task, while only the HC group displayed a social deficit of decreased shoaling. HC fish also showed an increased frequency of behavioral freezing and elevated cortisol responses to netting, heightened stress responses not seen in the PTZ fish. Since mild stressors, such as the handling the HC fish experienced, can lead to learned, advantageous responses to stress later in life, we tested escape response in the HC fish using an acoustic startle stimulus. The HC group showed an enhanced startle response, swimming significantly farther than either the PTZ or UC group immediately after being startled. Taken together, these results indicate that seizures in larval zebrafish impair learning and the development of an adaptive, heightened stress response after early-life stress. These findings expand the behavioral characterization of the larval zebrafish seizure model, strengthening the power of this model for ELS research.

Boldness, Aggression, and Shoaling Assays for Zebrafish Behavioral Syndromes.

A behavioral syndrome exists when specific behaviors interact under different contexts. Zebrafish have been test subjects in recent studies and it is important to standardize protocols to ensure proper analyses and interpretations. In our previous studies, we have measured boldness by monitoring a series of behaviors (time near surface, latency in transitions, number of transitions, and darts) in a 1.5 L trapezoidal tank. Likewise, we quantified aggression by observing bites, lateral displays, darts, and time near an inclined mirror in a rectangular 19 L tank. By dividing a 76 L tank into thirds, we also examined shoaling preferences. The shoaling assay is a highly customizable assay and can be tailored for specific hypotheses. However, protocols for this assay also must be standardized, yet flexible enough for customization. In previous studies, end chambers were either empty, contained 5 or 10 zebrafish, or 5 pearl danios (D. albolineatus). In the following manuscript, we present a detailed protocol and representative data that accompany successful applications of the protocol, which will allow for replication of behavioral syndrome experiments.

Interactions between aggression, boldness and shoaling within a brood of convict cichlids (Amatitlania nigrofasciatus).

A behavioral syndrome is considered present when individuals consistently express correlated behaviors across two or more axes of behavior. These axes of behavior are shy-bold, exploration-avoidance, activity, aggression, and sociability. In this study we examined aggression, boldness and sociability (shoaling) within a juvenile convict cichlid brood (Amatitlania nigrofasciatus). Because young convict cichlids are social, we used methodologies commonly used by ethologists studying social fishes. We did not detect an aggression-boldness behavioral syndrome, but we did find that the aggression, boldness, and possibly the exploration behavioral axes play significant roles in shaping the observed variation in individual convict cichlid behavior. While juvenile convict cichlids did express a shoaling preference, this social preference was likely convoluted by aggressive interactions, despite the small size and young age of the fish. There is a need for the development of behavioral assays that allow for more reliable measurement of behavioral axes in juvenile neo-tropical cichlids.

Sex differences in a shoaling-boldness behavioral syndrome, but no link with aggression.

A behavioral syndrome is observed in a population when specific behaviors overlap at the individual level in different contexts. Here, we explore boldness and aggression personality spectra, the repeatability of shoaling, and possible associated correlations between the behaviors in a population of lab-reared zebrafish (Danio rerio). Our findings describe a sex-specific boldness-shoaling behavioral syndrome, as a link between boldness and shoaling behaviors is detected. The results indicate that bold males are likely to have a stronger shoaling propensity than shy males for unfamiliar conspecifics. Conversely, bold females are more likely to shoal than shy females, but only when presented with heterospecific individuals. Additionally, aggression does not correlate with boldness or shoaling propensity for either sex. A positive relationship between boldness and shoaling that differs by sex is contrary to most of the present literature, but could help to explain population dynamics and may also have evolutionary implications.

A comparison of methodologies to test aggression in zebrafish.

Aggression is a psychological construct that is commonly used to classify zebrafish behavior. Aggression is a complex trait that can be difficult to accurately measure. The literature on fish behavior describes many different methodologies to examine aggression, which, we believe, have not been compared in a formal manner. In this study we observed 19 individual zebrafish (Danio rerio) and quantified bites, lateral displays, charges, darts, and time near the stimulus in six common assays used to measure aggression. The methodologies included an inclined mirror assay, two flat mirror assays with different acclimation periods, a live conspecific assay, a clay model stimulus assay, and a video recording assay. Our results indicate high repeatability in most aggressive behaviors over time, which confirms the value of each assay to measure personality. However, our results also indicate significant differences between the assays. Specifically, assays using a flat mirror or live conspecific as a stimulus for aggression elicited more attempted bites than an inclined mirror, a clay model stimulus, or a video recording stimulus. Furthermore, the inclined mirror stimulus provoked more darts than any other assay. The results suggest the need for researchers to consider specific research goals when selecting the appropriate stimulus to provoke aggression in zebrafish.

Fish cam: an online tool for introducing shoaling behavior to the classroom.

Fish Cam is an on-line educational resource that enables students to participate in behavioral research projects without ever leaving their classroom. By linking onto the Fish Cam site, students will observe an experimental tank in which fish choose shoal-mates in dichotomous choice tests. In these experiments, a test fish, in the central compartment, displays its shoaling preference by swimming near small shoals of fish in either of two side compartments. Assays are designed to examine the effects of phenotype, shoal size, and other factors known to influence shoaling. Students monitor Fish Cam in real time, and students collect data simply by running timers when the test fish crosses into the preference zones at each end of the central compartment. The times are logged onto data sheets that we provide, and we assist the students with their analysis. The simplicity of shoaling behavior makes it an ideal model system for data collection that is accessible to students of all ages and, in its first few years of operation, Fish Cam studies have been performed by fifth-, seventh- and eleventh-grade students. Sample lesson plans and handouts are available online to enhance the Fish Cam experience. The ultimate goals of this project are to make scientific research accessible in the classroom and promote science education.

Shoaling preferences and the effects of sex ratio on spawning and aggression in small laboratory populations of zebrafish (Danio rerio).

Owing to a lack of basic information on the biology of zebrafish (Danio rerio), lab managers must often base decisions regarding the care and use of this species on anecdotal information. In an effort to provide researchers with context-specific behavioral information, the authors evaluated shoaling and spawning behaviors in small groups of zebrafish. In each shoaling assay, a fish was given a choice to shoal with either a single fish or a group of three fish. Females preferred to shoal with a group of three individuals rather than with a single individual, regardless of the sex of the other fish. Males preferred groups of three males over single males but preferred single females to groups of three females. In spawning assays, zebrafish were placed in breeding tanks in one of three sex ratios (1 male:1 female; 3 males:1 female; 1 male:3 females). Reproductive efficiency did not differ among groups, but aggression (evaluated according to presence of shed scales) was more frequently observed in the male-dominated treatment group.

Shoaling preferences of two common killifish (Fundulus heteroclitus and F. diaphanus) in the laboratory and in the field: a new analysis of heterospecific shoaling.

Heterospecific grouping behavior of mummichogs (Fundulus heteroclitus) and banded killifish (Fundulus diaphanus) was analyzed in the laboratory and in a freshwater tidal marsh in Cremona, Maryland. Several parameters of wild, intact shoals were measured, including species composition, body length, parasite load, gender, and any physical abnormalities. Fish collected were used for laboratory analysis of shoaling preferences. When size was equal, banded killifish and mummichogs preferred conspecific shoals to heterospecific shoals, consisting of mummichogs, banded killifish, and sheepshead minnows (Cyprinodon variegates). Shoal collection from the field resulted in mixed species shoals with individuals predominantly unaffected by parasites or other physical abnormalities. Size appeared to be a sorting mechanism. A temporal shift in lengths was evident. Initial shoals caught contained significantly smaller fish compared to the final shoals caught. Results are compared with previous studies on heterospecific shoaling in killifish and new characteristics of heterospecific shoals inhabiting freshwater tidal marshes are discussed.

Methodologies for the care, maintenance, and breeding of tropical poison frogs.

The Biodiversity Laboratories at Saint Joseph's University house more than 100 exotic and endangered animal species-including insects, fish, amphibians, and reptiles. Research in the laboratories is devoted, primarily, to understanding the genetic and ecological bases of animal behavior. In addition, a common theme of the work is the development of techniques for the successful care of exotic animals within a laboratory setting. Such techniques may be critical to studies aimed at understanding the life histories of endangered species and saving them from extinction. Current projects to study the biology of tropical poison frogs have methodologies to improve captive care, maintenance, and breeding. In this article, I present these methodologies and their impact on the welfare of captive poison frogs.

Mutations in raised Drosophila melanogaster affect experience-dependent aspects of sexual behavior in both sexes.

Many aspects of the reproductive behavior of Drosophila melanogaster are modified dramatically by experience and age. Males' courtship of immature males and fertilized females decreases over time. Females' receptivity to copulation, and the behaviors that females perform and elicit, are affected by their age and sexual experience. We show that mutations in a raised stock affect all of these age- and experience-dependent aspects of male and female sexual behavior. Experience has no effect on raised males' courtship of immature males and has opposite effects on raised and wild-type males' courtship of fertilized females. In comparison to controls, raised females become sexually mature at an earlier age, and sexually mature raised virgin females copulate more quickly. Following mating, raised females elicit more courtship and remate faster and more frequently than control females.

CHEMICAL COMMUNICATION IN HAWAIIAN DROSOPHILA.

We are interested in elucidating the extent to which lekking Hawaiian Drosophila species have diverged from their continental counterparts, which engage in sexual behavior at communal food sources, with regard to the chemical communication systems that the flies employ. Accordingly, we have analyzed flies from three closely related Hawaiian Drosophila species in the adiastola subgroup. These species are of interest because the males engage in a unique behavior: while courting, they raise their abdomens over their heads and emit anal droplets. Analysis of the flies' behavior, the hydrocarbons in males' anal droplets, and males' cuticular hydrocarbons suggest that females' responses to males may be mediated by cuticular pheromones and/or pheromones in males' extruded droplets that enable the females to distinguish conspecific from heterospeciflc males. Conversely, perception of cuticular hydrocarbons from conspecific females enables D. adiastola males to distinguish females from a closely related species from conspecific females. On the basis of these observations, we suggest that the adiastola subgroup species are unique among drosophilids in that they utilize an anal droplet-mediated pheromone communication system, some or all components of which are species specific. However, the lekking Hawaiian Drosophila species are similar to D. melanogaster and related continental species in that the Hawaiian flies employ a cuticular pheromone communication system, some components of which are sex and species-specific.

INCOMPLETE BEHAVIORAL ISOLATION BETWEEN TWO DISTANTLY RELATED DROSOPHILA SPECIES.

Two sympatric, distantly related Drosophila species, D. affinis and D. melanogaster, interact sexually in the laboratory and in the field. When mature males from one species are tested with females or sexually attractive males from the other species, the mature males perform courtship that, in most cases, is indistinguishable from the courtship that attractive conspecific flies elicit. Moreover, some of the D. affinis females that elicit courtship from D. melanogaster males copulate with them.