Shoal Members Research Articles

Relative telencephalon size does not affect collective motion in the guppy (Poecilia reticulata).

Collective motion is common across all animal taxa, from swarming insects to schools of fish. The collective motion requires intricate behavioral integration among individuals, yet little is known about how evolutionary changes in brain morphology influence the ability for individuals to coordinate behavior in groups. In this study, we utilized guppies that were selectively bred for relative telencephalon size, an aspect of brain morphology that is normally associated with advanced cognitive functions, to examine its role in collective motion using an open-field assay. We analyzed high-resolution tracking data of same-sex shoals consisting of 8 individuals to assess different aspects of collective motion, such as alignment, attraction to nearby shoal members, and swimming speed. Our findings indicate that variation in collective motion in guppy shoals might not be strongly affected by variation in relative telencephalon size. Our study suggests that group dynamics in collectively moving animals are likely not driven by advanced cognitive functions but rather by fundamental cognitive processes stemming from relatively simple rules among neighboring individuals.

Use of multimodal sensory cues in predator avoidance by wild-caught zebrafish shoals

Abstract Fishes rely on a multimodal sensory system to assess cues from environmental factors, such as predation, vegetation cover, water flow and food availability. Here, we conducted laboratory-based experiments to examine the immediate response of wild-caught zebrafish (Danio rerio) shoals, comprising ten individuals, to one of the following treatments: (1) olfactory predator cue; (2) visual predator cue; (3) both cues simultaneously; and (4) control with no cue. Experiments were performed in a 75cm × 75cm arena, and test shoals were provided with olfactory cues from a predator and with a model predator as visual information on the presence of a predator. Using the idTracker software, we tracked and analysed shoal responses across these treatments. We found that in comparison to the control treatment, shoals receiving either visual or olfactory cues had significantly greater: (1) cohesion (inter-individual distance); (2) polarization (tendency of shoal members to orient in a common direction); and (3) velocity (directional speed). Interestingly, when the shoals received both visual and olfactory cues simultaneously, the cohesion, polarization and velocity decreased, and such shoals exhibited significantly more individual freezing events. Therefore, our study not only indicates that zebrafish rely comparably on vision and olfaction to escape predation but also shows that the anti-predator response elicited by zebrafish varies based on the number of cues provided.

A mixed-species shoal including fewer heterospecifics is preferred by jack mackerel Trachurus japonicus juveniles

Mixed-species shoals of fishes were reported from various environments such as coral reefs and temperate lakes. The driving mechanism of forming mixed-species shoal can be either haphazard or a preferential choice by an individual. Here, we tested if jack mackerel Trachurus japonicus, a temperate marine pelagic fish, could choose a shoal composition using vision based on a choice experiment. Shoal preference of jack mackerel juveniles was evaluated by presenting shoals comprising one to four heterospecifics (half-lined cardinal Ostorhinchus semilineatus) out of four shoal members to the focal fish. When both conspecific and heterospecific shoals were presented to a single jack mackerel, the focal fish showed a preference for the conspecific rather than heterospecific shoal, indicating that they can choose a conspecific shoal based on the visual information. However, when a focal fish encountered both conspecific and mixed-species shoals including fewer heterospecifics (one out of four individuals), the latter was preferred. The result implies that jack mackerel can judge a preferable composition of shoal using visual information. The inclusion of fewer heterospecifics may be exploiting oddity effects, whereby minority species are exposed to relatively high vulnerability to predators, as well as resulting in reduced competition for food while maintaining predator vigilance in a shoal.

Parasite infection impairs the shoaling behaviour of uninfected shoal members under predator attack

A key benefit of sociality is a reduction in predation risk. Cohesive group behaviour and rapid collective decision making are essential for reducing predation risk in groups. Parasite infection might reduce an individuals’ grouping behaviours and thereby change the behaviour of the group as a whole. To investigate the relationship between parasite infection and grouping behaviours, we studied groups of three-spined sticklebacks, Gasterosteus aculeatus, varying the number of individuals experimentally infected with the cestode Schistocephalus solidus. We studied groups of six sticklebacks containing 0, 2, 3, 4 or 6 infected individuals before and after a simulated bird attack. We predicted that infected individuals would have reduced shoaling and swimming speed and that the presence of infected individuals within a group would reduce group cohesion and speed. Uninfected fish increased shoaling and reduced swimming speed more than infected fish after the bird attack. In groups containing both infected and uninfected fish, the group behaviours were dominated by the more frequent character (uninfected versus infected). Interestingly, groups with equal numbers of uninfected and infected fish showed the least shoaling and had the lowest swimming speeds, suggesting that these groups failed to generate a majority and therefore displayed signs of indecisiveness by reducing their swimming speed the most. Our results provide evidence for a negative effect of infection on a group’s shoaling behaviour, thereby potentially deteriorating collective decision making. The presence of infected individuals might thus have far-reaching consequences in natural populations under predation risk.Significance statementParasite-infected individuals often show deviating group behaviours. This might reduce the anti-predator benefits of group living. However, it is unknown whether such deviations in group behaviour might influence the shoaling behaviour of uninfected group members and thereby the behaviour of the group as a whole. By experimentally infecting sticklebacks and investigating groups varying in infection rates, we show that infected sticklebacks differ in their shoaling behaviours from uninfected sticklebacks. Additionally, the presence of infected sticklebacks within the group affected the behaviour of uninfected shoal members. We show that shoals of infected fish are less cohesive and move slower compared to shoals of uninfected fish. Furthermore, we show that the infection rate of the shoal is crucial for how the group behaves.

Environmental habituation and sexual composition affect juveniles' shoaling activity in a cichlid fish (Pelvicachromis taeniatus).

Group living is widespread in animals, and many fishes form shoals. Examining within-group interactions in fishes may contribute to the general understanding of dynamic social structures in animals. The sex ratio of a group has been shown to influence grouping decisions of fishes and can be expected to affect behaviour at group level. Behavioural experiments usually involve relatively short acclimatisation times, although the establishment of environmental habituation in fishes is understudied. This study tests whether the sex ratio and long-term habituation to experimental conditions influence general shoal performance (activity parameters, density) and responses of shoals to an acoustic-mechanical disturbance cue in juveniles of the cichlid fish Pelvicachromis taeniatus via individual tracking. The disturbance consisted of a defined hit against the experimental tank, which caused sudden noise and water movement. We found that a higher proportion of females increases shoal activity (swimming speed and distance covered), suggesting that female P. taeniatus are more active than males. Furthermore, shoal activity declined when shoals habituated to the experimental settings and with the time that the shoals were grouped together, which may reflect intensified group member familiarity. Moreover, behavioural changes after disturbance were weaker when individuals were kept with their group longer and more familiar to the experimental conditions. For prey species, lower activity might be beneficial under natural conditions due to lower conspicuousness of the group. We did not find any significant effects of the investigated factors on shoal density (mean interindividual distance) and speed synchronisation. The results indicate that sexual composition, familiarity between shoal members and habituation to the experimental environment affect shoal performance in a cichlid fish.

Parasite infection disrupts escape behaviours in fish shoals.

Many prey species have evolved collective responses to avoid predation. They rapidly transfer information about potential predators to trigger and coordinate escape waves. Predation avoidance behaviour is often manipulated by trophically transmitted parasites, to facilitate their transmission to the next host. We hypothesized that the presence of infected, behaviourally altered individuals might disturb the spread of escape waves. We used the tapeworm Schistocephalus solidus, which increases risk-taking behaviour and decreases social responsiveness of its host, the three-spined stickleback, to test this hypothesis. Three subgroups of sticklebacks were placed next to one another in separate compartments with shelter. The middle subgroup contained either uninfected or infected sticklebacks. We confronted an outer subgroup with an artificial bird strike and studied how the escape response spread through the subgroups. With uninfected sticklebacks in the middle, escape waves spread rapidly through the entire shoal and fish remained in shelter thereafter. With infected sticklebacks in the middle, the escape wave was disrupted and uninfected fish rarely used the shelter. Infected individuals can disrupt the transmission of flight responses, thereby not only increasing their own predation risk but also that of their uninfected shoal members. Our study uncovers a potentially far-reaching fitness consequence of grouping with infected individuals.

Cascading effects of predation risk under high organic and inorganic turbidity: impacts on individuals and shoals of a mesopredator fish

We tested whether changes in water transparency caused either by precipitation-mediated sedimentation (inorganic turbidity) or eutrophication (organic turbidity) differently interact with top predator presence to determine predation rates, and individual decisions of mesopredators between foraging and avoiding predators. We tested the hypothesis that fear-induced cascade effects are more pronounced in organic turbid water through an experiment in which we manipulated turbidity (clear water and organic/inorganic turbid water) and predation risk (presence/absence of a top predator) in a 3 × 2 factorial design. We assessed prey consumption, prey selection (benthic and planktonic invertebrates) and food partitioning among individuals within shoals of an invertivorous fish, Moenkhausia forestii. The overall prey consumption by mesopredators was similar with and without top predator in clear water, but the presence of a top predator resulted in decreased invertebrate consumption in turbid waters, with no difference between organic and inorganic turbidity. Also, fear-induced cascade effects caused a strong preference of planktonic prey over benthic in inorganic turbidity and decreased evenness in prey consumption among individuals. Our findings suggest that the interactive effects of turbidity and top predator presence on mesopredator foraging depend on prey type and highlight individual differences in foraging behaviours among shoal members. Increased anthropogenic impacts on aquatic environments could have lasting long-term population impacts for fishes in terms of foraging and predator avoidance behaviour.

Bidirectional interactions facilitate the integration of a robot into a shoal of zebrafish Danio rerio.

Many studies on collective animal behavior seek to identify the individual rules that underlie collective patterns. However, it was not until the recent advancements of micro-electronic and embedded systems that scientists were able to create mixed groups of sensor-rich robots and animals and study collective interactions from the within a bio-hybrid group. In recent work, scientists showed that a robot-controlled lure is capable of influencing the collective decisions of zebrafish Danio rerio shoals moving in a ring and a two-room setup. Here, we study a closely related topic, that is, the collective behavior patterns that emerge when different behavioral models are reproduced through the use of a robotic lure. We design a behavioral model that alternates between obeying and disobeying the collective motion decisions in order to become socially accepted by the shoal members. Subsequently, we compare it against two extreme cases: a reactive and an imposing decision model. For this, we use spatial, directional and information theoretic metrics to measure the degree of integration of the robotic agent. We show that our model leads to similar information flow as in freely roaming shoals of zebrafish and exhibits leadership skills more often than the open-loop models. Thus, in order for the robot to achieve higher degrees of integration in the zebrafish shoal, it must, like any other shoal member, be bidirectionally involved in the decision making process. These findings provide insight on the ability to form mixed societies of animals and robots and yield promising results on the degree to which a robot can influence the collective decision making.

Qingbo (Spinibarbus sinensis) personalities and their effect on shoaling behavior

Group-level behavior has important fitness consequences for group-living fish, which might be affected by the personalities of group members. However, the role of group personality on group behavior has seldom been systematically investigated. To test the effect of the personalities of group members on shoaling behavior and the underlying mechanisms, we investigated the activity and boldness personality traits in qingbo (Spinibarbus sinensis) and grouped qingbo shoals into high personality score, low personality score, and randomly selected control groups and measured the shoal-level behavior traits. Compared with the reactive shoal (low activity and boldness), the proactive (high activity and boldness) shoal showed high activity and increased synchronization of speed, suggesting that the shoal-level behavior was affected by the personalities of the shoal members. Interestingly, the variables in the control shoal were midway between those in the high- and low-activity shoals, suggesting that the average activity score might be the most important factor for group-level behavior (average-determined mechanism). However, when qingbo were divided into high- and low-boldness shoals, the control shoal variables were similar to those of the low-boldness shoals, suggesting that a minority of low-boldness individuals might have a dominant effect on shoal-level behavior (minority-determined mechanism). Cohesion showed a more complicated pattern than the other variables, which might be because the conformity of personalities among shoal members had a profound effect on cohesion. This finding suggests that large interindividual differences in personality play a profound effect on group-level behavior, and different mechanisms might provide the precondition for shoals to exhibit great shoal-level diversity. These findings might have important ecological relevance for the survival of group-living fish in habitats with food resource and predation intensity fluctuations.

Assessing the effects of water flow rate on parasite transmission amongst a social host

Many freshwater habitats are subject to change through time. Specifically, natural flow regimes are substantially modified by not only seasonal climatic change, but also anthropogenic activity. Consequently, freshwater organisms are exposed to variable flow, potentially altering their behaviour and subsequently parasite transmission amongst social hosts. Here, we investigate the effects of flow conditions on the shoaling behaviour of Trinidadian guppies (Poecilia reticulata), and the resulting transmission of a directly transmitted ectoparasite, Gyrodactylus turnbulli. Shoals exposed to continuous flow exhibited significantly greater G. turnbulli peak intensities and abundance when compared to fish subjected to interrupted, but not no-flow conditions. Parasite transmission rate was greater in shoals exposed to interrupted flow, resulting in parasites becoming more distributed amongst shoal members and thus reducing mean intensity in comparison to continuous flow shoals. Furthermore, as prevalence increased, the distance between shoaling conspecifics increased at greater rates in interrupted and no-flow conditions compared to continuous flow: indicating that in the absence of flowing water, parasitism has a greater effect on shoaling decisions. This data highlights how fish behaviourally respond to variable flow conditions and the implications for parasite transmission.

Quantity discrimination in fish species: fish use non-numerical continuous quantity traits to select shoals.

Fish typically prefer to live in big shoals due to the associated ecological benefits. Shoaling is a behavior that depends on the ability to quantitatively discriminate. The fundamental mechanism involved in quantity discrimination determines whether fish can discriminate a shoal using numerical discrete cues (e.g., number of shoal members), non-numerical continuous traits (e.g., total body surface area) or both; however, the mechanism is currently a controversial topic. In the present study, we used a spontaneous choice experiment to test whether guppy (Poecilia reticulata), zebrafish (Danio rerio), Chinese crucian carp (Carassius auratus) and qingbo (Spinibarbus sinensis) rely on continuous (i.e., body surface area) or discrete (i.e., number of shoal members) information for shoal selection by altering the body surface area (cumulative body surface area ratio of 3:2 or 1:1) between two stimulus shoals with a different number of members (2 individuals vs 3 individuals). All four fish species preferred to shoal with the stimulus shoal with the larger cumulative surface area even if the shoal had fewer members; however, fish showed no shoal preference when the cumulative surface body areas of both stimulus shoals were equal. Furthermore, qingbo did not numerically discriminate between a shoal with 1 individual and a shoal with 3 individuals when the cumulative surface areas of both stimulus shoals were equal; however, qingbo showed a preference for the shoal with the larger cumulative surface area when the two stimulus shoals each had 3 individuals. In conclusion, the present study demonstrated that all four fish species relied only on non-numerical continuous quantity information for shoal selection, at least under a difficult task (i.e., 2 vs 3).

Quantity discrimination in angelfish (Pterophyllum scalare) is maintained after a 30-s retention interval in the large but not in the small number range.

The ability to discriminate between sets that differ in the number of elements can be useful in different contexts and may have survival and fitness consequences. As such, numerical/quantity discrimination has been demonstrated in a diversity of animal species. In the laboratory, this ability has been analyzed, for example, using binary choice tests. Furthermore, when the different number of items first presented to the subjects are subsequently obscured, i.e., are not visible at the moment of making a choice, the task requires memory for the size of the sets. In previous work, angelfish (Pterophyllum scalare) have been found to be able to discriminate shoals differing in the number of shoal members both in the small (less than 4) and the large (4 or more) number range, and they were able to perform well even when a short memory retention interval (2-15s) was imposed. In the current study, we increased the retention interval to 30s during which the shoals to choose between were obscured, and investigated whether angelfish could show preference for the larger shoal they saw before this interval. Subjects were faced with a discrimination between numerically small shoals (≤4 fish) and also between numerically large (≥4 fish) shoals of conspecifics. We found angelfish not to be able to remember the location of larger versus smaller shoals in the small number range, but to exhibit significant memory for the larger shoal in the large number range as long as the ratio between these shoals was at least 2:1. These results, together with prior findings, suggest the existence of two separate quantity estimation systems, the object file system for small number of items that does not work with the longer retention interval and the analogue magnitude system for larger number of items that does.

A mechanism mediating inter-individual associations in mixed-species groups

Individuals within groups benefit when they are phenotypically and behaviourally similar. Despite this, mixed-species aggregations are found among a wide range of animal taxa and are known to provide advantages in terms of enhanced foraging efficiency and predator avoidance. However, little is known about individual level interactions within these groups or the mechanisms mediating their structure. Diet-derived chemical cues can mediate within-species associations in fish, but it is not known whether they serve a similar function across species. To investigate this, we created mixed-species groups of sympatric three-spined (Gasterosteus aculeatus) and nine-spined (Pungitius pungitius) sticklebacks and manipulated the diet of the constituent group members. We found that three-spined sticklebacks did not associate more frequently with conspecifics over heterospecifics when all animals received the same diet, suggesting that species-specific cues did not influence association patterns. However, when group members were fed different diets, they interacted with heterospecifics fed on the same diet more than with conspecifics from a different diet treatment. This study demonstrates the importance of chemical cues as a possible mechanism for establishing familiarity, and thereby mediating within-group structure even between individuals in mixed-species aggregations. Understanding these underlying mechanisms has significant implications for our understanding of other social dynamics, including social learning and information transfer. Mixed-species groupings are common among animals, although we have a poor understanding of how these groups are structured, what mediates interactions between group members, and what the function of such groups is. In a study using two co-occurring stickleback species, we show that fish preferred to associate with shoal members fed the same diet as themselves, irrespective of their species. This is most likely mediated by diet-derived chemical cues and may allow fish to avoid detection by predators or forage alongside individuals with similar dietary preferences. Understanding the mechanisms underpinning non-random associations in mixed-species groups has significant implications for our understanding of other social dynamics, including social learning and information transfer.

Turbidity amplifies the non‐lethal effects of predation and affects the foraging success of characid fish shoals

Summary In aquatic systems, many species rely primarily on visual cues to choose optimal foraging sites, capture prey and avoid potential threats. Increases in the turbidity of water reduce visibility and impede animals in determining the precise location of both predators and food. How individuals balance foraging decisions with anti‐predator behaviour in turbid environments is not well understood. We tested the effects of turbidity and predation risk on the foraging behaviour and feeding of an invertivorous fish, Moenkhausia forestii (Characidae), using a mesocosm experiment with a 2 × 2 design, crossing water clarity (clear versus turbid) with predation risk as reflected by the presence or absence of the piscivorous wolf‐fish Hoplias aff. malabaricus (Erythrinidae). We predicted that turbidity and predator presence would interact additively to reduce foraging rates, and that increased turbidity or predator presence would result in disproportionate food partitioning among shoal members. The combination of high turbidity and predator presence resulted in a significant reduction in prey consumption. Foraging success exhibited a skewed distribution in the turbid treatment, i.e. there was a decreased evenness of food partitioning within shoals. Hence, both turbidity and predator presence affect the prey consumption and foraging behaviour of invertivorous fish, with turbidity amplifying the non‐lethal effects of predation risk on foraging success. Our results imply that turbidity‐induced visual obstruction amplifies the negative effects of predator presence on invertivorous fish feeding behaviour, resulting in higher prey survival. Also, our finding that food intake by an intermediate consumer decreased in turbid water with a top predator lends no support to the hypothesis that intermediate consumer fish reduce their anti‐predator behaviour in turbid water. From a management perspective, our findings suggest that the oligotrophication of aquatic systems could dramatically increase predation on basal prey organisms naturally adapted to turbid waters, and reduce within‐shoal differences in feeding behaviour.

Familiarity affects social network structure and discovery of prey patch locations in foraging stickleback shoals.

Numerous factors affect the fine-scale social structure of animal groups, but it is unclear how important such factors are in determining how individuals encounter resources. Familiarity affects shoal choice and structure in many social fishes. Here, we show that familiarity between shoal members of sticklebacks (Gasterosteus aculeatus) affects both fine-scale social organization and the discovery of resources. Social network analysis revealed that sticklebacks remained closer to familiar than to unfamiliar individuals within the same shoal. Network-based diffusion analysis revealed that there was a strong untransmitted social effect on patch discovery, with individuals tending to discover a task sooner if a familiar individual from their group had previously done so than if an unfamiliar fish had done so. However, in contrast to the effect of familiarity, the frequency with which individuals had previously associated with one another had no effect upon the likelihood of prey patch discovery. This may have been due to the influence of fish on one another's movements; the effect of familiarity on discovery of an empty ‘control’ patch was as strong as for discovery of an actual prey patch. Our results demonstrate that factors affecting fine-scale social interactions can also influence how individuals encounter and exploit resources.

Combined effects of flow condition and parasitism on shoaling behaviour of female guppies Poecilia reticulata.

Group living in fish can provide benefits of protection from predators and some parasites, more efficient foraging for food, increased mating opportunities and enhanced energetic benefit when swimming. For riverine species, shoaling behaviour can be influenced by various environmental stressors, yet little is known how flow rate might influence the shoaling of diseased fish shoals. In view of the increasingly unpredictable flow rates in streams and rivers, this study aimed to assess the combined effect of flow condition and parasitism on the shoaling behaviour of a model fish species. Shoal size, shoal cohesion and time spent shoaling of female guppies Poecilia reticulata were compared when infected with the directly transmitted ectoparasite Gyrodactylus turnbulli under flow and static conditions. Flow condition was an important factor in influencing shoaling behaviour of guppies with the fish forming larger shoals in the absence of flow. When a shoal member was infected with G. turnbulli, shoal cohesion was reduced, but the magnitude of this effect was dependent on flow condition. In both flow and static conditions, bigger fish formed larger shoals than smaller counterparts. Future changes to stream hydrology with more frequent flooding and drought events will affect the shoaling tendency of fish. During high-flow events, diseased fish may not be able to keep up with shoal mates and therefore have a higher risk of predation. Additionally, these findings may be important for aquaria and farmed species where an increase in flow rate may reduce aggregation in fish.

The role of body surface area in quantity discrimination in angelfish (Pterophyllum scalare).

Although some fish species have been shown to be able to discriminate between two groups (shoals) of conspecifics differing in the number of members, most studies have not controlled for continuous variables that covary with number. Previously, using angelfish (Pterophyllum scalare) we started the systematic analysis of the potential influence of such continuous variables, and found that they play different roles in shoal discrimination depending on whether large (≥4 fish) or small (<4 fish) shoals were contrasted. Here, we examine the potential role of the overall body surface area of stimulus fish in shoal preference, a prominent variable not yet examined in angelfish. We report that both when numerically large (5 versus 10 fish) and when small (2 versus 3 fish) shoals were contrasted, angelfish were unable to discriminate the numerically different shoals as long as the surface area of the contrasted shoals was equated. Thus, we conclude that body surface may be an important continuous variable in shoal discrimination. This conclusion was further supported by the analysis of preference when shoals of the same numerical size but different body surface area were contrasted. We found subjects to spend significantly more time close to the shoals with the greater overall surface area. Last, we conducted an experiment in which we simultaneously controlled a set of continuous variables, including overall surface area, and found angelfish to use the number of shoal members as a cue only in large shoal contrasts but not in small shoal contrasts. This result suggests the potential existence of different processing systems for large and small numbers in fish.

Mechanisms and extent of information transfer in socially foraging guppies, Poecilia reticulata

Many species in social groups make use of readily available, socially provided information for behaviours including predator avoidance, mate choice and foraging. Not only are actions imitated, but they may also be reapplied in variable future situations, suggesting an explicit knowledge of the purpose of the behaviours. Social learning thus enables animals to learn about their complex environments rapidly and efficiently, aiding survival. However, little is known of the processes underlying information transmission and the complexity of information that can be exchanged. We used shoals of guppies to investigate how animals decide which individual to copy, and the extent of information that can be transferred. Naïve guppies followed the first fish to move. However, although the speed and accuracy of foraging increased significantly during training, the first fish to move was not always the trained (knowledgeable) guppy. No significant difference was found between the number of guppies entering areas of varying food quality, supporting the hypothesis that these fish learn foraging locations by following and suggesting that leaders forage without considering the reduced personal reward arising from sharing a low-quality patch with followers. Animals that learn by copying thus appear limited in the amount of information they can receive, highlighting a selection pressure for more extensive means of transmitting information in complex environments. ► We investigate how and how much information fish can learn from other fish. ► Guppies follow the first individual to leave the shoal: the bold individual. ► Boldness was generally associated with knowledge of where food was to be found. ► Leaders did not consider their shoal members when choosing where to forage. ► Animals that learn by copying are limited in the amount of information they can receive.

Pheromones: Fish Fear Factor

Fish, like many other animals, panic when another individual is injured. Now, the chemical nature of a substance that mediates this reaction has been uncovered.

Inferring the rules of interaction of shoaling fish

Collective motion, where large numbers of individuals move synchronously together, is achieved when individuals adopt interaction rules that determine how they respond to their neighbors' movements and positions. These rules determine how group-living animals move, make decisions, and transmit information between individuals. Nonetheless, few studies have explicitly determined these interaction rules in moving groups, and very little is known about the interaction rules of fish. Here, we identify three key rules for the social interactions of mosquitofish (Gambusia holbrooki): (i) Attraction forces are important in maintaining group cohesion, while we find only weak evidence that fish align with their neighbor's orientation; (ii) repulsion is mediated principally by changes in speed; (iii) although the positions and directions of all shoal members are highly correlated, individuals only respond to their single nearest neighbor. The last two of these rules are different from the classical models of collective animal motion, raising new questions about how fish and other animals self-organize on the move.