Abstracts: CryoLetters 21 (5), 2000

CryoLetters is a bimonthly, international journal for low temperature science and technology

CryoLetters 21, 261-270 (2000)
CryoLetters, c/o Royal Veterinary College, London NW1 0TU

Recovery of Embryos of Zizania Palustris following Exposure to liquid nitrogen

Darren Touchell and Christina Walters*

USDA-ARS National Seed Storage Laboratory, 1111 South Mason St, Fort Collins Colorado, 80521, USA.


Factors affecting survival of embryos of Zizania palustris after exposure to liquid nitrogen were investigated. Moisture content was the most critical factor. Embryos desiccated to a moisture content between 0.36 and 0.56 g H2O/g dry weight had highest survival after liquid nitrogen exposure (564% and 529% respectively). The recovery of embryos in the light following liquid nitrogen exposure also influenced survival. Survival increased from 354% to 564% when embryos containing 0.36 g H2O/g dry weight were recovered in the dark compared to light. Embryos exposed to desiccation and freezing stresses were subject to increased levels of lipid peroxidation. Light exacerbated lipid peroxidation in recovering embryos. Furthermore, catalase and peroxidase, enzymes involved in reducing reactive oxygen species, increased in activity in response to stress, and were further up-regulated in tissues recovered in the light. This study suggests that there are a number of factors influencing the survival of tissues exposed to liquid nitrogen and recovery procedures that reduce oxidative stress should be employed.

Keywords: cryostorage, dark, light, liquid nitrogen, oxidative stress, recovery, water content, Zizania palustris



CryoLetters 21, 271-278 (2000)
© CryoLetters, c/o Royal Veterinary College, London NW1 0TU

low temperature survival of Slender Naiad (Najas flexilis) seeds

F.R. Hay* and J.S.K. Muir

Millennium Seed Bank, Royal Botanic Gardens Kew, Wakehurst Place, Ardingly, West Sussex, RH17 6TN, UK. E-mail:


The response to drying and storage at -20C or in LN2 was studied in seeds of the freshwater aquatic plant Najas flexilis. The seeds of this species show some desiccation sensitivity, although post-harvest storage in water at 16C resulted in improvements in desiccation tolerance. There was 63% germination of seeds dried to 9.5% moisture content (30% RH) following this maturation period. Optimum moisture contents for seeds stored at -20C for 3 months and in LN2 for 1 week were ~11% and ~15%, respectively.

Keywords: Najas flexilis, seeds, low temperature survival.



CryoLetters 21, 279-288 (2000)
© CryoLetters, c/o Royal Veterinary College, London NW1 0TU


Keith Harding1 and Erica E. Benson2*

1Crop Genetics, Scottish Crop Research Institute, Invergowrie, Dundee DD2 5DA,  Scotland; e-mail:
2*Corresponding author: Plant Conservation Biotechnology Group, Division of Molecular and Life Sciences, School of Science and Engineering, University of Abertay Dundee, Bell Street, Dundee DD1 1HG, Scotland; e-mail:


Shoot-tips derived from S. tuberosum cultivar Brodick were cryopreserved using the encapsulation/dehydration technique. Total DNA was extracted from 30 in vitro plantlets of Brodick regenerated from surviving shoot-tips. DNA was digested with Bam HI and examined using DNA-DNA hybridisation analysis. Fractionation of the digested DNA showed stability of the nuclear-chloroplast genomic complex and identical chloroplast-DNA fragments were detected in all regenerated plantlets. This suggests that the observed nuclear DNA fragments and the chloroplast genome are stable in potato plants regenerated from cryopreserved shoot-tips. This study extends the range of molecular biological techniques which can be applied to examine post-cryogenic storage stability in cryopreserved plant germplasm.

Keywords: cryopreservation, encapsulation, molecular stability, potato, chloroplast DNA.



CryoLetters 21, 289-296 (2000)
©CryoLetters, c/o Royal Veterinary College, London NW1 0TU


Lauritz Smme

University of Oslo, Department of Biology, P.O.Box 1050 Blindern, N-0316 Oslo, Norway


R.A.F. Reaumur in 1736 was the first entomologist to observe that some insects can tolerate freezing, while others cannot. The first reviews on insect cold hardiness were published by P. Bachmetjew who also discovered supercooling in insects. Authors like W. Robinson realised that piercing the cuticle changed the supercooling capacity and N.M. Payne pointed out that there are great seasonal variations in insect cold hardiness. In the early literature, references were frequently made to plant cold hardiness, in which theories on tolerance to freezing were more elaborated. A modern approach to the studies of insect cold hardiness was initiated by R.W. Salt. The number of reports increased enormously in the 1960's and 1970's. During the last 20 to 30 years new insight has appeared from studies on ice nucleators, antifreeze proteins, ice-nucleating bacteria, desiccation and biochemistry of cryoprotectants.

Keywords: arthropods, cold hardiness, history



CryoLetters 21, 297-308 (2000)
© CryoLetters, c/o Royal Veterinary College, London NW1 0TU


H.V. Danks

Biological Survey of Canada (Terrestrial Arthropods), Canadian Museum of Nature, P.O. Box 3443, Station D, Ottawa, Ontario, Canada K1P 6P4


The cold climates and diverse environments of Canada have allowed key studies of insect cold hardiness that developed and widened the understanding of this subject. For example, freezing tolerance, chilling tolerance, freezing resistance, supercooling, cryoprotectants and other features can be combined in many different ways, reflecting a wide range of adaptations. Many other factors interact with and influence cold hardiness, such as habitats and their selection, and water and energy balances. These findings suggest several topics that would be especially fruitful for further study in northern Canada.

Keywords:  Cold hardiness, Insects, Canada, Arctic



CryoLetters 21, 309-314 (2000)
© CryoLetters, c/o Royal Veterinary College, London NW1 0TU


Martin Holmstrup*, Mark Bayley, Heidi Sjursen, Rikke Hjer, Steen Bossen and Kirstine Friis

Department of Terrestrial Ecology, National Environmental Research Institute, Vejlsvej 25, P. O. Box 314, DK-8600 Silkeborg, Denmark Phone +45 8920 1400; fax +45 8920 1414; *e-mail:


The adaptations to climatic stress of invertebrates have been extensively studied and the underlying physiology is relatively well understood. It is therefore possible to predict critical lower temperature and moisture limits at which a species will suffer increased mortality. Environmental pollution can reduce the tolerance to climatic stress considerably but it is not known how significant these synergistic effects are in polluted areas. Results are presented showing examples of synergism between toxic and climatic stress. The physiological mechanisms involved in the synergistic effects are discussed.

Keywords: stress tolerance, cold hardiness, drought tolerance, toxic effects, synergistic effects



CryoLetters 21, 315-324 (2000)
© CryoLetters, c/o Royal Veterinary College, London NW1 0TU


M. Roger Worland1* Peter Convey1 and Alena Lukeov2

1 British Antarctic Survey, Natural Environment Research Council, High Cross, Madingley Road, Cambridge CB3 0ET, UK Fax: 01223 221259; e-mail:
*Corresponding author
Institute of Soil Biology, Czech Academy of Sciences, Na Sdkch 7, 370 05 Česk Budĕjovice, Czech Republic


This study examined the rate of cold hardening of a field population of Antarctic springtails and the effect of eating food with particular levels of ice nucleating activity on the animals whole body freezing point. The SCPs of samples of c. 20, freshly collected, Cryptopygus antarcticus were measured hourly over a 32 hour collection period using differential scanning calorimetry and related to habitat temperature. The mean SCP of the springtails increased from 24 to 10C during which time the habitat temperature warmed slowly from 2.5 to +2.5C.

In laboratory experiments, previously starved, cold tolerant springtails were fed on selected species of algae with measured SCPs but there was no clear correlation between the SCP of food and that of the animals after feeding. Microscopic examination of faecal pellets and guts from springtails showed that algal cells were completely destroyed during digestion.

Keywords: Antarctic, terrestrial algae, food selection, microclimate, springtail, cold tolerance, cyanobacteria, digestion, soil, differential scanning calorimetry


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