Allee effect in larval resource exploitation in Drosophila: an interaction among density of adults, larvae, and micro-organisms

Abstract 1. Aggregation pheromones can evolve when individuals benefit from clustering. Such a situation can arise with an Allee effect, i.e. a positive relationship between individual fitness and density of conspecifics. Aggregation pheromone in Drosophila induces aggregated oviposition. The aim of the work reported here was to identify an Allee effect in the larval resource exploitation by Drosophila melanogaster, which could explain the evolution of aggregation pheromone in this species. 2. It is hypothesised that an Allee effect in D. melanogaster larvae arises from an increased efficiency of a group of larvae to temper fungal growth on their feeding substrate. To test this hypothesis, standard apple substrates were infested with specified numbers of larvae, and their survival and development were monitored. A potential beneficial effect of the presence of adult flies was also investigated by incubating a varying number of adults on the substrate before introducing the larvae. Adults inoculate substrates with yeast, on which the larvae feed. 3. Fungal growth was related negatively to larval survival and the size of the emerging flies. Although the fungal growth on the substrate was largely reduced at increased larval densities, the measurements of fitness components indicated no Allee effect between larval densities and larval fitness, but rather indicated larval competition. 4. In contrast, increased adult densities on the substrates prior to larval development yielded higher survival of the larvae, larger emerging flies, and also reduced fungal growth on the substrates. Hence, adults enhanced the quality of the larval substrate and significant benefits of aggregated oviposition in fruit flies were shown. Experiments with synthetic pheromone indicated that the aggregation pheromone itself did not contribute directly to the quality of the larval resource. 5. The interaction among adults, micro‐organisms, and larval growth is discussed in relation to the consequences for total fitness.     

Spatial aggregation across ephemeral resource patches in insect communities: an adaptive response to natural enemies?

Although an increase in competition is a common cost associated with intraspecific crowding, spatial aggregation across food-limited resource patches is a widespread phenomenon in many insect communities. Because intraspecific aggregation of competing insect larvae across, e.g. fruits, dung, mushrooms etc., is an important means by which many species can coexist (aggregation model of species coexistence), there is a strong need to explore the mechanisms that contribute to the maintenance of this kind of spatial resource exploitation. In the present study, by using Drosophila-parasitoid interactions as a model system, we tested the hypothesis whether intraspecific aggregation reflects an adaptive response to natural enemies. Most of the studies that have hitherto been carried out on Drosophila-parasitoid interactions used an almost two-dimensional artificial host environment, where host larvae could not escape from parasitoid attacks, and have demonstrated positive density-dependent parasitism risk. To test whether these studies captured the essence of such interactions, we used natural breeding substrates (decaying fruits). In a first step, we analysed the parasitism risk of Drosophila larvae on a three-dimensional substrate in natural fly communities in the field, and found that the risk of parasitism decreased with increasing host larval density (inverse density dependence). In a second step, we analysed the parasitism risk of Drosophila subobscura larvae on three breeding substrate types exposed to the larval parasitoids Asobara tabida and Leptopilina heterotoma. We found direct density-dependent parasitism on decaying sloes, inverse density dependence on plums, and a hump-shaped relationship between fly larval density and parasitism risk on crab apples. On crab apples and plums, fly larvae benefited from a density-dependent refuge against the parasitoids. While the proportion of larvae feeding within the fruit tissues increased with larval density, larvae within the fruit tissues were increasingly less likely to become victims of parasitoids than those exposed at the fruit surface. This suggests a facilitating effect of group-feeding larvae on reaching the spatial refuge. We conclude that spatial aggregation in Drosophila communities can at least in part be explained as a predator avoidance strategy, whereby natural enemies act as selective agents maintaining spatial patterns of resource utilisation in their host communities.     

Fruit,flies and filamentous fungi – experimental analysis of animal–microbe competition using Drosophila melanogaster and Aspergillus mould as a model system

In addition to their fundamental role in nutrient recycling, saprobiotic microorganisms may be considered as typical consumers of food‐limited ephemeral resource patches. As such, they may be engaged in inter‐specific competition with saprophagous animals feeding from the same resource. Bacteria and filamentous fungi are known to synthesise secondary metabolites, some of which are toxic and have been proposed to deter or harm animals. The microorganisms may, however, also be negatively affected if saprophagous animals do not avoid microbe‐laden resources but feed in the presence of microbial competitors. We hypothesised that filamentous fungi compete with saprophagous insects, whereby secondary metabolites provide a chemical shield against the insect competitors. For testing this, we developed a new ecological model system representing a case of animal–microbe competition between saprobiotic organisms, comprising Drosophila melanogaster and species of the fungus Aspergillus (A. nidulans, A. fumigatus, A. flavus). Infestation of Drosophila breeding substrate with proliferating fungal colonies caused graduated larval mortality that strongly depended on mould species and colony age. Confrontation with conidiospores only, did not result in significant changes in larval survival, suggesting that insect death may not be ascribed to pathogenic effects. When confronted with colonies of transgenic fungi that lack the ability to express the global secondary metabolite regulator LaeA (ΔlaeA), larval mortality was significantly reduced compared to the impact of the wild type strains. Yet, also in the ΔlaeA strains, inter‐specific variation in the influence on insect growth occurred. Competition with Drosophila larvae impaired fungal growth, however, wild type colonies of A. nidulans and A. flavus recovered more rapidly from insect competition than the corresponding ΔlaeA mutants (not in A. fumigatus). Our findings provide genetic evidence that toxic secondary metabolites synthesised by saprotrophic fungi may serve as a means to combat insect competitors. Variation in the ability of LaeA to control expression of various secondary metabolite gene clusters might explain the observed species‐specific variation in Drosophila–Aspergillus competition.     

Maternal effects increase survival probability in Drosophila subobscura larvae

Drosophilid flies breeding on ephemeral resource patches (e.g., decaying fruits) are assumed to transfer yeasts to their oviposition sites, presumably in order to positively affect offspring development. We tested this hypothesis with Drosophila subobscura Collin (Diptera: Drosophilidae) by manipulating their nutritional (yeast‐fed vs. non‐yeast‐fed) and reproductive status (mated vs. non‐mated). Flies were then released into vials containing decaying fruits (either sloes, crab apples, or Syrian plums). After a constant residence time in the vials, the flies were removed, 16 first‐instar larvae were transferred to the fruits and their survival probability to the adult stage was recorded. Whereas previous exposure of the larval substrate to yeast‐fed males and virgin females (yeast‐fed and non‐yeast‐fed) had no effect on survivorship, exposure to yeast‐fed and mated females that deposited eggs on the fruits (subsequently removed) led to a significant increase in the survival probability of the transferred larvae to the adult stage. Although the exact mechanism of yeast transmission remains to be determined, we suggest an active inoculation of the breeding substrates with yeast by ovipositing females. In agreement with previous studies, we found a negative effect of mould growth on larval survival, which, however, depended on the fruit type. We discuss various scenarios of yeast involvement in benefits to the insect larvae and suggest that insect–mould interactions should be examined in detail in order to better understand the behavioural and life‐history traits of insects that depend on ephemeral resources.     

Friendly food for fitter flies? – Influence of dietary microbial species on food choice and parasitoid resistance in Drosophila

Nutrition fuels any activity performed by an organism and has been shown to affect its ability to withstand pathogens and parasites. Furthermore, animals over a wide range of taxa have been shown to exhibit a choice of foods and nutrients that are beneficial to their fitness. Saprophagous animals most often feed on microorganisms growing on dead organic matter rather than the organic matter itself. Various yeast species play an important role in both larval and adult nutrition of saprophagous Drosophila melanogaster. We hypothesised the dietary microbial species to affect life‐history traits of D. melanogaster, including the ability to fend offparasitoids, and larvae to prefer to devour those yeast species beneficial to their development and immunocompetence. Particular yeast species known to be associated with D. melanogaster could be shown to have a substantial influence on various larval and adult fitness traits including the ability to encapsulate eggs of the parasitoid wasp Asobara tabida. It also turned out that larvae chose to devour those yeast species which supported their ability to encapsulate parasitoid eggs. Which yeast species was preferred and had a beneficial impact on encapsulation ability, was subject to inter‐individual variability within the investigated population, hinting at the existence of an adaptive heritable variability regarding individual choice and salubriousness of food. The results suggest that the dietary microbial species of saprophagous insects may influence the resistance against parasitoid attacks and thus the outcome of the interaction between a saprophagous host and its parasitoids.     

Are Drosophila preferences for yeasts stable or contextual?

Whether there are general mechanisms, driving interspecific chemical communication is uncertain. Saccharomycetaceae yeast and Drosophila fruit flies, both extensively studied research models, share the same fruit habitat, and it has been suggested their interaction comprises a facultative mutualism that is instigated and maintained by yeast volatiles. Using choice tests, experimental evolution, and volatile analyses, we investigate the maintenance of this relationship and reveal little consistency between behavioral responses of two isolates of sympatric Drosophila species. While D. melanogaster was attracted to a range of different Saccharomycetaceae yeasts and this was independent of fruit type, D. simulans preference appeared specific to a particular S. cerevisiae genotype isolated from a vineyard fly population. This response, however, was not consistent across fruit types and is therefore context‐dependent. In addition, D. simulans attraction to an individual S. cerevisiae isolate was pliable over ecological timescales. Volatile candidates were analyzed to identify a common signal for yeast attraction, and while D. melanogaster generally responded to fermentation profiles, D. simulans preference was more discerning and likely threshold‐dependent. Overall, there is no strong evidence to support the idea of bespoke interactions with specific yeasts for either of these Drosophila genotypes. Rather the data support the idea Drosophila are generally adapted to sense and locate fruits infested by a range of fungal microbes and/or that yeast–Drosophila interactions may evolve rapidly.     

Competition with filamentous fungi and its implication for a gregarious lifestyle in insects living on ephemeral resources

Abstract. 1. Recent studies have demonstrated the existence of positive density dependence in the survival and development of Drosophila (the so‐called Allee effect); however the underlying mechanisms of such Allee effects have remained elusive. Competition with filamentous fungi have often been suggested to be involved in causing high mortality at low larval density, but it has not yet been explicitly tested if the well known spatial aggregation of insect eggs yields a fitness benefit for the developing larvae in the presence of noxious moulds. 2. Using Drosophila melanogaster, the present study tested whether larval survival is greater in aggregations when confronted with various combinations of three representative mould species (Aspergillus, Alternaria, and Penicillium) and a head start for fungal development. 3. High rates of fungal‐dependent mortality and significant positive density‐dependent larval survival (i.e. Allee effects) were observed when larvae were confronted with food resources containing established colonies of Aspergillus or Alternaria. Neither the simultaneous transfer of Aspergillus or Alternaria spores with larvae to food patches nor food infections with Penicillium affected insect larval development. 4. Significant correlations between mould growth and larval survival could be identified, although the patterns that emerged were shown to be inconsistent when the effects were compared between fungal species and fungal priority. Because mould growth only partly explained larval survival, the influence of other fungal‐borne factors, e.g. mycotoxins, needs to be elucidated in order to understand the mechanistic basis of insect–mould interactions. 5. These results are the first to argue convincingly for moulds being involved in mediating Allee effects for insects on ephemeral resources; however they also demonstrate an unexpected diversity in insect–mould interactions. Considering this diversity may be important in understanding insect spatial ecology.     

Drosophila regulate yeast density and increase yeast community similarity in a natural substrate

Drosophila melanogaster adults and larvae, but especially larvae, had profound effects on the densities and community structure of yeasts that developed in banana fruits. Pieces of fruit exposed to adult female flies previously fed fly-conditioned bananas developed higher yeast densities than pieces of the same fruits that were not exposed to flies, supporting previous suggestions that adult Drosophila vector yeasts to new substrates. However, larvae alone had dramatic effects on yeast density and species composition. When yeast densities were compared in pieces of the same fruits assigned to different treatments, fruits that developed low yeast densities in the absence of flies developed significantly higher yeast densities when exposed to larvae. Across all of the fruits, larvae regulated yeast densities within narrow limits, as compared to a much wider range of yeast densities that developed in pieces of the same fruits not exposed to flies. Larvae also affected yeast species composition, dramatically reducing species diversity across fruits, reducing variation in yeast communities from one fruit to the next (beta diversity), and encouraging the consistent development of a yeast community composed of three species of yeast (Candida californica, C. zemplinina, and Pichia kluvyeri), all of which were palatable to larvae. Larvae excreted viable cells of these three yeast species in their fecal pools, and discouraged the growth of filamentous fungi, processes which may have contributed to their effects on the yeast communities in banana fruits. These and other findings suggest that D. melanogaster adults and their larval offspring together engage in 'niche construction', facilitating a predictable microbial environment in the fruit substrates in which the larvae live and develop.     

Patterns of resource distribution among conspecific larvae in two fruit fly species: Anastrepha fraterculus and Ceratitis capitata (Diptera: Tephritidae)

相似文献   

Yeast communities of the cactusPilosocereus arrabidae and associated insects in the Sandy Coastal Plains of Southeastern Brazil

The yeast communities from necrotic tissues, decaying flowers and fruits, and from larval feeding sites of the mothSigelgaita sp. in the cactusPilosocereus arrabidae were surveyed in three restinga ecosystems in Southeastern Brazil. Insects associated with these substrates were sampled to verify the vectoring of yeasts. The cactusPilosocereus arrabidae was shown to have four different yeast communities associated with it. Necrotic stems had a diverse yeast community with the prevalent speciesPichia barkeri, Candida sonorensis, Pichia cactophila, Geotrichum sp.,Myxozyma mucilagina andSporopachydermia sp. A, representing about 80% of the total isolates.Pichia sp. A and aCandida domercqii-like species represented more than 90% of the yeast isolates from decaying flowers. Fruits had a heterogeneous yeast community with typical fruit yeasts of the genusKloeckera, basidiomicetous anamorphs of the genusCryptococcus, the black yeastAureobasidium pullulans, Pichia sp. A, aCandida domercqii-like species, and some cactophilic yeasts, especiallyClavispora opuntiae. The feeding site ofSigelgaita sp. larvae hadClavispora opuntiae as the prevalent species. Insect vectors are suggested as one the most important factors influencing the composition of these yeast communities.     

Do yeasts and Drosophila interact just by chance?

The fruit fly Drosophila melanogaster and the baker's yeast Saccharomyces cerevisiae are classic research model organisms that are also associated in nature, at least around vineyards. Sharing the same ephemeral fruit niche, winged Drosophila feed on immotile yeasts. That a yeast diet is essential for larval development, and that saprophagous fruit flies are attracted to a suite of yeast volatiles, has been well established over the last century. Recently, research has focussed on the potential mutual benefit of this interaction hypothesising yeasts also benefit via dispersal from ephemeral fruits. It now appears that the concept of a co-evolved mutualism between yeasts and Drosophila has permeated the literature. However, until robust evidence regarding the evolution and maintenance of this yeast-fly association has been provided, we suggest there is no compelling evidence to reject the more simplistic null hypothesis that these interactions are due to exaptation, and not a mutualism driven by natural selection.     

Nutrient balance in medfly, Ceratitis capitata, larval diets affects the ability of the developing insect to incorporate lipid and protein reserves

The Mediterranean fruit fly [Ceratitis capitata Wiedemann (Diptera: Tephritidae)], or medfly, is mass produced in many facilities throughout the world to supply sterile flies for sterile insect technique programs. Production of sterile males requires large amounts of larval and adult diets. Larval diets comprise the largest economic burdens in the mass production of sterile flies, and are one of the main areas where production costs could be reduced without affecting quality and efficacy. The present study investigated the effect of manipulating diet constituents on larval development and performance. Medfly larvae were reared on diets differing in the proportions of brewer's yeast and sucrose. We studied the effect of such diets on the ability of pupating larvae to accumulate protein and lipids, and on other developmental indicators. Except for diets with a very low proportion of brewer's yeast (e.g., 4%), pupation and adult emergence rates were in general high and satisfactory. The ability of pupating larvae to accumulate lipid reserves and proteins was significantly affected by the sucrose and yeast in the diet, and by the proportion of protein to carbohydrates (P/C). In contrast to previous nutritional studies conducted with other insects, low P/C in medfly larval diets (with excess dietary carbohydrates) resulted in pupating medfly larvae having a relatively reduced load of lipids; medfly larvae protein contents in these diets were, as expected, relatively low. Similarly, high P/C ratios in the diet produced larvae with high protein and lipid contents. Differences with other insects may be due to differential post‐ingestion regulation where a high dietary carbohydrate diet reduces the lipogenic activity of the larvae, and induces a shift from lipid to glucose oxidation. Larvae reared on low P/C diets spent more time foraging in the diet than larvae maintained on a high P/C diet, suggesting a compensatory mechanism to complement nutrient intake. The results suggest that the content of brewer's yeast, the most expensive diet component, could be fine‐tuned without apparently affecting fly quality.     

Irradiation effect on the structure of bacterial communities associated with the oriental fruit fly,Bactrocera dorsalis

The role of symbiotic microbes in insects, especially the beneficial character of this interaction for insects, has received much attention in recent years as it has been related to important aspects of the host insects' biology such as development, reproduction, survival, and fitness. Among insect hosts, tephritid fruit flies are well known to form beneficial associations with their symbionts. To control these destructive agricultural pests, environmentally friendly approaches, like the sterile insect technique as a component of integrated pest management strategies, remain most effective. In this study, changes in the bacterial profile of mass‐reared oriental fruit flies, Bactrocera dorsalis (Hendel) (Diptera: Tephritidae), were examined in both larval and adult stages and also after irradiation by employing a 16S rRNA gene‐based Illumina sequencing approach. Proteobacteria was the prevalent bacterial phylum in non‐irradiated adults and larvae. Alphaproteobacteria was the most abundant class in larvae but almost absent in adults, which was dominated by Gammaproteobacteria. Firmicutes were present in both developmental stages but at lower relative abundance. At genus level, Acetobacter prevailed in the larval stage and members of the Enterobacteriaceae family in adults. Irradiated samples exhibited higher diversity and richness indices compared to the non‐irradiated oriental fruit flies, whereas no significant changes were observed between the two developmental stages of the non‐irradiated samples. Lactobacillus, members of the Orbacecae family, and Morganella were detected but to a lesser degree upon irradiation, whereas the relative abundance of Lactococcus and Orbus increased. The bacterial profile of larvae appeared to be different compared to that of adult B. dorsalis flies. The subsequent application of irradiation at the pupal stage led to the development of different microbiota between treated and untreated samples, affecting diversity and operational taxonomic unit composition. Irradiated samples of oriental fruit flies were characterized by higher species diversity and richness.     

Differential response to larval crowding of a long‐ and a short‐lived medfly biotype

Response of endophytic fruit fly species (Tephritidae) to larval crowding is a form of scramble competition that may affect important life history traits of adults, such as survival and reproduction. Recent empirical evidence demonstrates large differences in adult life history traits, especially longevity, among Mediterranean fruit fly (Ceratitis capitata; "medfly") biotypes obtained from different regions of the world. However, whether the evolution of long lifespan is associated with response to stress induced by larval crowding has not been fully elucidated. We investigated, under constant laboratory conditions, the response of a short‐ and a long‐lived medfly biotypes to stress induced by larval crowding. Survival and development of larvae and pupae and the size of resulting pupae were recorded. The lifespan and age‐specific egg production patterns of the obtained adults were recorded. Our findings reveal that increased larval density reduced immature survival (larvae and pupae) in the short‐lived biotype but had rather neutral effects on the longed‐lived one. Only larvae of the long‐lived biotype were capable of prolonging their developmental duration under the highest crowding regime to successfully pupate and emerge as adults. Response of emerging adults to larvae crowding conditions was similar in the two medfly biotypes. Those individuals emerging from high larval density regimes had reduced longevity and fecundity. Long‐lived biotype individuals, however, appeared to suffer a higher cost in longevity compared with the short‐lived one. The importance of our findings to understand the evolution of long lifespan is discussed.     

Kin recognition and co‐operative foraging in Drosophila melanogaster larvae

A long‐standing goal for biologists and social scientists is to understand the factors that lead to the evolution and maintenance of co‐operative behaviour between conspecifics. To that end, the fruit fly, Drosophila melanogaster, is becoming an increasingly popular model species to study sociality; however, most of the research to date has focused on adult behaviours. In this study, we set out to examine group‐feeding behaviour by larvae and to determine whether the degree of relatedness between individuals mediates the expression co‐operation. In a series of assays, we manipulated the average degree of relatedness in groups of third‐instar larvae that were faced with resource scarcity, and measured the size, frequency and composition of feeding clusters, as well as the fitness benefits associated with co‐operation. Our results suggest that larval D. melanogaster are capable of kin recognition (something that has not been previously described in this species), as clusters were more numerous, larger and involved more larvae, when more closely related kin were present in the social environment. These findings are discussed in the context of the correlated fitness‐associated benefits of co‐operation, the potential mechanisms by which individuals may recognize kin, and how that kinship may play an important role in facilitating the manifestation of this co‐operative behaviour.     

Weak responses to dietary enrichment in a specialized aphid predator

Some ladybeetles are specialist predators of aphids, coccids or other prey, although they often eat a variety of species from their focal prey taxon. In addition, the diet is often supplemented with alternative prey. How larvae of the aphidophagous Coccinella septempunctata L. utilize a non‐aphid alternative prey (fruit‐fly larvae Drosophila melonogaster Meigen) is compared with adequate (i.e. high‐quality) aphid prey provided alone (monotypic diet) or in mixed diets. The alternative prey are presented either nutrient‐enriched (i.e. raised on dog food supplemented medium) or not (raised on pure medium). Ladybird performance (survival, growth and development) is poor on the pure fly larvae diets, and also reduced when given mixed diets compared with the pure aphid diet. Nutrient enrichment of the fly larvae has no positive effects. The physiological background for the differences in food value, as indicated by performance in life‐history parameters, is a strong pre‐ingestive effect (i.e. reduced consumption of fly larvae compared with aphids) and a post‐ingestive effect (i.e. reduced utilization of assimilated larval fly tissue), whereas the assimilation efficiency of the consumed fly larvae is as high as that of aphids. The results show a physiological trade‐off resulting from prey specialization that reduces the possibility of utilizing alternative prey when the availability of aphids is scarce. Connected with this is a high robustness against variation in prey nutrient diversity and composition; the ladybird shows little positive response to dietary mixing (i.e. neither mixing of adequate aphids, nor of aphids and alternative prey) or to nutrient enrichment of prey. This contrasts with the results from generalist predators (spiders), where similar treatments lead to strong effects on life‐history parameters.     

Chemical signaling and insect attraction is a conserved trait in yeasts

Yeast volatiles attract insects, which apparently is of mutual benefit, for both yeasts and insects. However, it is unknown whether biosynthesis of metabolites that attract insects is a basic and general trait, or if it is specific for yeasts that live in close association with insects. Our goal was to study chemical insect attractants produced by yeasts that span more than 250 million years of evolutionary history and vastly differ in their metabolism and lifestyle. We bioassayed attraction of the vinegar fly Drosophila melanogaster to odors of phylogenetically and ecologically distinct yeasts grown under controlled conditions. Baker's yeast Saccharomyces cerevisiae, the insect‐associated species Candida californica, Pichia kluyveri and Metschnikowia andauensis, wine yeast Dekkera bruxellensis, milk yeast Kluyveromyces lactis, the vertebrate pathogens Candida albicans and Candida glabrata, and oleophilic Yarrowia lipolytica were screened for fly attraction in a wind tunnel. Yeast headspace was chemically analyzed, and co‐occurrence of insect attractants in yeasts and flowering plants was investigated through a database search. In yeasts with known genomes, we investigated the occurrence of genes involved in the synthesis of key aroma compounds. Flies were attracted to all nine yeasts studied. The behavioral response to baker's yeast was independent of its growth stage. In addition to Drosophila, we tested the basal hexapod Folsomia candida (Collembola) in a Y‐tube assay to the most ancient yeast, Y. lipolytica, which proved that early yeast signals also function on clades older than neopteran insects. Behavioral and chemical data and a search for selected genes of volatile metabolites underline that biosynthesis of chemical signals is found throughout the yeast clade and has been conserved during the evolution of yeast lifestyles. Literature and database reviews corroborate that yeast signals mediate mutualistic interactions between insects and yeasts. Moreover, volatiles emitted by yeasts are commonly found also in flowers and attract many insect species. The collective evidence suggests that the release of volatile signals by yeasts is a widespread and phylogenetically ancient trait, and that insect–yeast communication evolved prior to the emergence of flowering plants. Co‐occurrence of the same attractant signals in yeast and flowers suggests that yeast‐insect communication may have contributed to the evolution of insect‐mediated pollination in flowers.     

Dynamics of Social Behavior in Fruit Fly Larvae

We quantified the extent and dynamics of social interactions among fruit fly larvae over time. Both a wild-type laboratory population and a recently-caught strain of larvae spontaneously formed social foraging groups. Levels of aggregation initially increased during larval development and then declined with the wandering stage before pupation. We show that larvae aggregated more on hard than soft food, and more at sites where we had previously broken the surface of the food. Groups of larvae initiated burrowing sooner than solitary individuals, indicating that one potential benefit of larval aggregations is an improved ability to dig and burrow into the food substrate. We also show that two closely related species, D. melanogaster and D. simulans, differ in their tendency to aggregate, which may reflect different evolutionary histories. Our protocol for quantifying social behavior in larvae uncovered robust social aggregations in this simple model, which is highly amenable to neurogenetic analyses, and can serve for future research into the mechanisms and evolution of social behavior.