Vitrification is essential for anhydrobiosis in an African chironomid, Polypedilum vanderplanki

doi:10.1073/pnas.0706197105

Vitrification is essential for anhydrobiosis in an African chironomid, Polypedilum vanderplanki

*Center for Biological Resources and Informatics, Tokyo Institute of Technology, B-62 Nagatsuta-cho, Midori-ku, Yokohama 226-8501, Japan;
^†Spectroscopic Instruments Division, JASCO Corporation, Hachioji, Tokyo 192-8537, Japan; and
^‡Anhydrobiosis Research Unit, National Institute of Agrobiological Sciences, Ohwashi 1-2, Tsukuba, Ibaraki 305-8634, Japan

Edited by David L. Denlinger, Ohio State University, Columbus, OH, and approved February 1, 2008 (received for review July 2, 2007)

Abstract

Anhydrobiosis is an extremely dehydrated state in which organisms show no detectable metabolism but retain the ability to revive after rehydration. Thus far, two hypotheses have been proposed to explain how cells are protected during dehydration: (i) water replacement by compatible solutes and (ii) vitrification. The present study provides direct physiological and physicochemical evidence for these hypotheses in an African chironomid, Polypedilum vanderplanki, which is the largest multicellular animal capable of anhydrobiosis. Differential scanning calorimetry measurements and Fourier-transform infrared (FTIR) analyses indicated that the anhydrobiotic larvae were in a glassy state up to as high as 65°C. Changing from the glassy to the rubbery state by either heating or allowing slight moisture uptake greatly decreased the survival rate of dehydrated larvae. In addition, FTIR spectra showed that sugars formed hydrogen bonds with phospholipids and that membranes remained in the liquid-crystalline state in the anhydrobiotic larvae. These results indicate that larvae of P. vanderplanki survive extreme dehydration by replacing the normal intracellular medium with a biological glass. When entering anhydrobiosis, P. vanderplanki accumulated nonreducing disaccharide trehalose that was uniformly distributed throughout the dehydrated body by FTIR microscopic mapping image. Therefore, we assume that trehalose plays important roles in water replacement and intracellular glass formation, although other compounds are surely involved in these phenomena.

Footnotes

^§To whom correspondence should be addressed. E-mail: oku{at}affrc.go.jp
Author contributions: M.S. and T.F. contributed equally to this work; M.S., T.F., and T.O. designed research; T.F., K.-i.A., D.T., Y.N., T.K., and M.W. performed research; M.S., T.F., and K.-i.A. analyzed data; and M.S., T.F., Y.N., and T.O. wrote the paper.
The authors declare no conflict of interest.
This article is a PNAS Direct Submission.
This article contains supporting information online at www.pnas.org/cgi/content/full/0706197105/DCSupplemental.
↵ ¶ In recent years, values of ≈115 ± 2°C have been widely accepted as the onset or midpoint value of the glass-transition temperature of anhydrous trehalose, Tg ₁ (51, 59–62), though observed value have ranged from 73°C (10) to 117°C (60, 61). Lower values could be due to the effects of residual water or impurities. There has been more consistent agreement between measurements of the glass-transition temperature of pure water (Tg ₂), giving −135°C as the onset value (52, 59). The value of the parameter, k, in the Gordon–Taylor equation is sensitive to the values of Tg ₁ and Tg ₂, which should be determined by fitting onset glass transition temperatures measured at various water contents, with careful choice of the value of Tg ₁. The Gordon–Taylor equation parameterized in ref. 51 satisfies these requirements for confidence.
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