Synchrotron X-Ray Visualisation of Ice Formation in Insects during Lethal and Non-Lethal Freezing

Journal Article · 14 December 2009 · PLoS ONE

DOI: https://doi.org/10.1371/journal.pone.0008259 · OSTI ID:1627392

Sinclair, Brent J. ^[1]; Gibbs, Allen G. ^[2]; Lee, Wah-Keat ^[3]; Rajamohan, Arun ^[4]; Roberts, Stephen P. ^[2]; Socha, John J. ^[5]

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1. Univ. of Western Ontario, London, ON (Canada). Dept. of Biology; DOE/OSTI
2. Univ. of Nevada, Las Vegas, NV (United States). School of Life Sciences
3. Argonne National Lab. (ANL), Argonne, IL (United States). X-Ray Science Division
4. Univ. of Western Ontario, London, ON (Canada). Dept. of Biology
5. Virginia Polytechnic Inst. and State Univ. (Virginia Tech), Blacksburg, VA (United States). Dept. of Engineering Science and Mechanics

Although the biochemical correlates of freeze tolerance in insects are becoming well-known, the process of ice formation in vivo is subject to speculation. We used synchrotron x-rays to directly visualise real-time ice formation at 3.3 Hz in intact insects. We observed freezing in diapausing 3rd instar larvae of Chymomyza amoena (Diptera: Drosophilidae), which survive freezing if it occurs above 214uC, and non-diapausing 3rd instar larvae of C. amoena and Drosophila melanogaster (Diptera: Drosophilidae), neither of which survive freezing. Freezing was readily observed in all larvae, and on one occasion the gut was seen to freeze separately from the haemocoel. There were no apparent qualitative differences in ice formation between freeze tolerant and non-freeze tolerant larvae. The time to complete freezing was positively related to temperature of nucleation (supercooling point, SCP), and SCP declined with decreasing body size, although this relationship was less strong in diapausing C. amoena. Nucleation generally occurred at a contact point with the thermocouple or chamber wall in nondiapausing larvae, but at random in diapausing larvae, suggesting that the latter have some control over ice nucleation. There were no apparent differences between freeze tolerant and non-freeze tolerant larvae in tracheal displacement or distension of the body during freezing, although there was markedly more distension in D. melanogaster than in C. amoena regardless of diapause state. We conclude that although control of ice nucleation appears to be important in freeze tolerant individuals, the physical ice formation process itself does not differ among larvae that can and cannot survive freezing. This suggests that a focus on cellular and biochemical mechanisms is appropriate and may reveal the primary adaptations allowing freeze tolerance in insects.

• Research Organization: Argonne National Lab. (ANL), Argonne, IL (United States)
• Sponsoring Organization: USDOE Office of Science (SC), Biological and Environmental Research (BER). Biological Systems Science Division
• Grant/Contract Number: AC02-06CH11357
• OSTI ID: 1627392
• Journal Information: PLoS ONE, Journal Name: PLoS ONE Journal Issue: 12 Vol. 4; ISSN 1932-6203
• Publisher: Public Library of ScienceCopyright Statement
• Country of Publication: United States
• Language: English