The cryopreservation techniques proposed for embryos of the fruit fly Drosophila melanogaster are not yet ready for practical use. Alternative methods for long-term storage of D. melanogaster strains, although urgently needed, do not exist. Herein, we describe a narrow interval of low temperatures under which the larvae of D. melanogaster can be stored in quiescence for up to two months. The development of larvae was arrested at the pre-wandering stage under fluctuating thermal regime (FTR), which simultaneously resulted in diminishing the accumulation of indirect chill injuries. Our physiological, metabolomic, and transcriptomic analyses revealed that compared to larvae stored at constant low temperatures, the larvae stored under FTR conditions were able to decrease the rates of depletion of energy substrates, exploited brief warm episodes of FTR for homeostatic control of metabolite levels, and more efficiently exerted protection against oxidative damage.

Faculty of Science University of South Bohemia Branišovská 31 37005 České Budějovice Czech Republic

Institut für Populationsgenetik Vetmeduni Vienna Vienna Austria

Institute of Entomology Biology Centre CAS Branišovská 31 37005 České Budějovice Czech Republic

Université de Rennes 1 UMR CNRS 6553 ECOBIO 263 Avenue du Général Leclerc 35042 Rennes France

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