The transcriptomic signature of responses to larval crowding in Drosophila melanogaster.

Intraspecific competition at the larval stage is an important ecological factor affecting life-history, adaptation and evolutionary trajectory in holometabolous insects. However, the molecular pathways underpinning these ecological processes are poorly characterized. We reared Drosophila melanogaster at three egg densities (5, 60, and 300 eggs/mL) and sequenced the transcriptomes of pooled third-instar larvae. We also examined emergence time, egg-to-adult viability, adult mass, and adult sex-ratio at each density. Medium crowding had minor detrimental effects on adult phenotypes compared to low density and yielded 24 differentially expressed genes (DEGs), including several chitinase enzymes. In contrast, high crowding had substantial detrimental effects on adult phenotypes and yielded 2107 DEGs. Among these, upregulated gene sets were enriched in sugar, steroid and amino acid metabolism as well as DNA replication pathways, whereas downregulated gene sets were enriched in ABC transporters, taurine, Toll/Imd signaling, and P450 xenobiotics metabolism pathways. Overall, our findings show that larval crowding has a large consistent effect on several molecular pathways (i.e., core responses) with few pathways displaying density-specific regulation (i.e., idiosyncratic responses). This provides important insights into how holometabolous insects respond to intraspecific competition during development.

Microbiota disruption leads to reduced cold tolerance in Drosophila flies.

It is now acknowledged that bacteria from gut microbiota deeply interact with their host by altering many physiological traits. Such interplay is likely to consequently affect stress tolerance. Here, we compared cold and heat tolerance of Drosophila melanogaster flies with undisrupted (control (Co)) versus disrupted gut microbiota (dechorionated eggs (De)). The disrupting treatment strongly reduced bacterial load in flies' guts, though 16S sequencing analysis did not evidence strong diversity changes in the remaining bacterial community. Both chill coma recovery and acute cold survival were repeatedly lower in De than in Co flies under our experimental conditions. However, heat tolerance was not consistently affected by gut disruption. Our results suggest that microbiota-related effects on the host can alter ecologically relevant traits such as thermal tolerance.

Hormesis-like effect of mild larval crowding on thermotolerance in Drosophila flies.

Crowding is a complex stress that can affect organisms' physiology, especially through decreased food quality and accessibility. Here, we evaluated the effect of larval density on several biological traits of Drosophila melanogaster An increasing gradient, from 1 to 1000 eggs per milliliter of food, was used to characterize life-history traits variations. Crowded conditions resulted in striking decreases of fresh mass (up to 6-fold) and viability, as well as delayed development. Next, we assessed heat and cold tolerance in L3 larvae reared at three selected larval densities: low (LD, 5 eggs ml-1), medium (MD, 60 eggs ml-1) and high (HD, 300 eggs ml-1). LT50 values of MD and, to a lesser extent, HD larvae were repeatedly higher than those from LD larvae, under both heat and cold stress. We investigated potential physiological correlates associated with this density-dependent thermotolerance shift. No marked pattern could be drawn from the expression of stress-related genes. However, a metabolomic analysis differentiated the metabotypes of the three density levels, with potential candidates associated with this clustering (e.g. glucose 6-phosphate, GABA, sugars and polyols). Under HD, signs of oxidative stress were noted but not confirmed at the transcriptional level. Finally, urea, a common metabolic waste, was found to accumulate substantially in food from MD and HD larvae. When supplemented in food, urea stimulated cold tolerance but reduced heat tolerance in LD larvae. This study highlights that larval crowding is an important environmental parameter that induces drastic consequences on flies' physiology and can affect thermotolerance in a density-specific way.