BACKGROUND: The codling moth (Cydia pomonella) is a major insect pest of apples worldwide. Fully grown last instar larvae overwinter in diapause state. Their overwintering strategies and physiological principles of cold tolerance have been insufficiently studied. No elaborate analysis of overwintering physiology is available for European populations. PRINCIPAL FINDINGS: We observed that codling moth larvae of a Central European population prefer to overwinter in the microhabitat of litter layer near the base of trees. Reliance on extensive supercooling, or freeze-avoidance, appears as their major strategy for survival of the winter cold. The supercooling point decreases from approximately -15.3 °C during summer to -26.3 °C during winter. Seasonal extension of supercooling capacity is assisted by partial dehydration, increasing osmolality of body fluids, and the accumulation of a complex mixture of winter specific metabolites. Glycogen and glutamine reserves are depleted, while fructose, alanine and some other sugars, polyols and free amino acids are accumulated during winter. The concentrations of trehalose and proline remain high and relatively constant throughout the season, and may contribute to the stabilization of proteins and membranes at subzero temperatures. In addition to supercooling, overwintering larvae acquire considerable capacity to survive at subzero temperatures, down to -15 °C, even in partially frozen state. CONCLUSION: Our detailed laboratory analysis of cold tolerance, and whole-winter survival assays in semi-natural conditions, suggest that the average winter cold does not represent a major threat for codling moth populations. More than 83% of larvae survived over winter in the field and pupated in spring irrespective of the overwintering microhabitat (cold-exposed tree trunk or temperature-buffered litter layer).

Institute of Entomology Biology Centre of the Academy of Sciences of the Czech Republic České Budějovice Czech Republic

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Barnes MM (1991) Tortricids in pome and stone fruits. In: Van Der Geestand LPS, Evenhuis HH, editors. Tortricid pests. Their biology, natural enemies and control. Amsterdam: Elsevier Science Publishers, pp. 313–327.

Willett MJ, Neven L, Miller CE (2009) The occurrence of codling moth in low latitude countries: validation of pest distribution reports. Hort Technol 19: 633–637.

Dorn S, Schumacher P, Abivardi C, Meyhöfer R (1999) Global and regional pest insects and their antagonists in orchards: spatial dynamics. Agric Ecosyst Environ 73: 111–118.

Audermard H (1991) Population dynamics of the codling moth. In: Van Der Geestand LPS, Evenhuis HH, editors. Tortricid pests. Their biology, natural enemies and control. Amsterdam: Elsevier Science Publishers, pp. 327–338.

Miller F (1956) Zemědělská entomologie. Praha: Nakladatelství československé akademie věd, 1057 p.

Peterson DM, Hamner WM (1968) Photoperiodic control of diapause in the codling moth. J Insect Physiol 14: 519–528.

Sieber R, Benz G (1980) Termination of the facultative diapause in the codling moth, Laspeyresia pomonella (Lepidoptera, Tortricidae). Entomol Exp Appl 28: 204–212.

Williams DG, Macdonald G (1982) The duration and number of immature stages of codling moth Cydia pomonella (L.) (Tortricidae: Lepidoptera). Aust J Entomol 21: 1–4.

MacLellan CR (1958) Role of woodpeckers in control of the codling moth in Nova Scotia. Can Entomol 90: 18–22.

McLellan CR (1959) Woodpeckers as predators of the codling moth in Nova Scotia. Can Entomol 91: 673–680.

Le Roux EJ (1959) Importance and control of the codling moth, Carpocapsa pomonella (L.) (Lepidoptera: Torticidae), on apple in Quebec. Rep Pomol Fruit Grow Soc Queb 1959: 45–60.

Mailloux M, Le Roux EJ (1960) Further observations on the life-history and habits of the codling moth, Carpocapsa pomonella (L.) (Lepidoptera: Tortricidae), in apple orchards of southwestern Quebec. Rep Pomol Fruit Grow Soc Queb 1960: 45–56.

Solomon ME, Glen DM, Kendall DA, Milsom NF (1976) Predation of overwintering larvae of codling moth (Cydia pomonella (L.)) by birds. J Appl Ecol 13: 341–352.

Glen DM, Milsom NF (1978) Survival of mature larvae of codling moth (Cydia pomonella) on apple trees and ground. Ann Appl Biol 90: 133–146.

Gould E, Geissler GH (1941) Hibernating codling moth larvae. J Econ Ent 34: 445–450.

McLellan CR (1960) Cocooning behaviour of overwintering codling moth larvae. Can Entomol 92: 469–479.

Neven LG (1999) Cold hardiness adaptations of codling moth, Cydia pomonella . Cryobiology 38: 43–50. PubMed

Khani A, Moharramipour S (2007) Seasonal change of cold hardiness in the codling moth, Cydia pomonella (Lepidoptera: Tortricidae). Pak J Biol Sci 10: 2591–2594. PubMed

Khani A, Moharramipour S (2010) Cold hardiness and supercooling capacity in the overwintering larvae of the codling moth, Cydia pomonella . J Insect Sci 10: 83. PubMed PMC

Khani A, Moharramipour S, Barzegar M (2007) Cold tolerance and trehalose accumulation in overwintering larvae of the codling moth, Cydia pomonella (Lepidoptera: Tortricidae). Eur J Entomol 104: 385–392.

Toba HH, Howell JF (1991) An improved system for mass-rearing of codling moths. J Entomol Soc B C 88: 22–27.

Fuková I, Nguyen P, Marec F (2005) Codling moth cytogenetics: karyotype, chromosomal location of rDNA, and molecular differentiation of sex chromosomes. Genome 48: 1088–1092. PubMed

Koštál V, Doležal P, Rozsypal J, Moravcová M, Zahradníčková H, et al. (2011a) Physiological and biochemical analysis of overwintering and cold tolerance in two Central European populations of the spruce bark beetle, Ips typographus . J Insect Physiol 57: 1136–1146. PubMed

Zachariassen KE (1985) Physiology of cold tolerance in insects. Physiol Rev 65: 799–832. PubMed

Sformo T, Walters K, Jeannet K, Wowk B, Fahy GM, et al. (2010) Deep supercooling, vitrification and limited survival to −100°C in the Alaskan beetle Cucujus clavipes puniceus (Coleoptera: Cucujidae) larvae. J Exp Biol 213: 502–509. PubMed

Gessner MO, Neumann PTM (2005) Total lipids. In: Graça MAS, Bärlocher F, Gessner MO, eds.Methods to study litter decomposition: A practical guide. Berlin: Springer, pp 103–107.

Folch AJ, Lees M, Stanley GH (1957) A simple method for the isolation and purification of total lipids from animal tissues. J Biol Chem 226: 497–509. PubMed

Bueding E, Orrell SA (1964) A mild procedure for the isolation of polydisperse glycogen from animal tissues. J Biol Chem 239: 4018–4020. PubMed

Dubois M, Gilles A, Hamilthon JJ, Rebers PA, Smith F (1956) Colorimetric method for determination of sugars and related substances. Anal Chem 28: 350–356.

Newcomer EJ (1920) Winterkilling of codling moth larvae. J Econ Entomol 13: 441–442.

Jegen G (1922) Winter measures against fruit-tree pests. Landw Jahrb Schweiz 36: 83–101.

Headlee TJ (1930) Study of agents for destroying overwintering codling moth larvae. N J Agr Expt Sta Ann Rep 1930: 142–145.

Kot J (1958) Freezing death of codling moth larvae. Ekol Pol Ser B 4: 173–176.

Mailloux M, Leroux EJ (1963) Notes on the distribution of the cocoons, winter mortality and development of the codling moth in the orchards of Quebec in 1962. Ann Soc Entomol Que 8: 48–57.

Sateyev AF (1963) The effect of overwintering conditions on death of codling moth caterpillars under the conditions of central Kazakhstan. Trudy Bot Inst Akad Nauk SSSR 17: 189–194.

MacPhee AW (1964) Cold-hardiness, habitat, and winter survival of some orchard arthropods in Nova Scotia. Can Entomol 96: 617–625.

Sato H (1964) A preliminary report on the response to temperature of the overwintering immature stage of Carpocapsa pomonella . Soc Plant Protect No Japan Ann Rep 15: 113.

Storey KB, Storey JM (1988) Freeze tolerance in animals. Physiol Rev 68: 27–84. PubMed

Lee RE Jr (1991) Principles of insect low temperature tolerance. In: Lee RE Jr, Denlinger DL, editors. Insects at Low Temperature. New York: Chapman and Hall, pp. 17–46.

Lee RE Jr. (2010) A primer on insect cold-tolerance. In: Denlinger DL, Lee RE Jr., editors. Low temperature biology of insects. Cambridge: Cambridge University Press, pp. 3–34.

Sinclair BJ (1999) Insect cold tolerance: How many kinds of frozen? Eur J Entomol 96: 157–164.

Rudolph AS, Crowe JH, Crowe LM (1986) Effects of three stabilizing agents, proline, betaine and trehalose, on membrane phospholipids. Arch Biochem Biophys 245: 134–143. PubMed

Carpenter JF, Crowe JH (1988) The mechanisms of cryoprotection of proteins by solutes. Cryobiology 25: 244–255. PubMed

Koštál V, Zahradníčková H, Šimek P (2011b) Hyperprolinemic larvae of the drosophilid fly, Chymomyza costata, survive cryopreservation in liquid nitrogen. Proc Nat Acad Sci USA108: 13041–13046. PubMed PMC

Koštál V, Šimek P, Zahradníčková H, Cimlová J, Štětina T (2012) Conversion of the chill susceptible fruit fly larva (Drosophila melanogaster) to a freeze tolerant organism. Proc Nat Acad Sci USA 109: 3270–3274. PubMed PMC

Koštál V, Havelka J (2000) Diapausing larvae of the midge Aphidoletes aphidimyza (Diptera: Cecidomyiidae) survive at subzero temperatures in a supercooled state but tolerate freezing if inoculated by external ice. Eur J of Entomol 97: 433–436.

Bale JS (1987) Insect cold hardiness: Freezing and supercooling – an ecological perspective. J Insect Physiol 33: 899–908.

Bale JS (1993) Classes of insect cold hardiness. Funct Ecol 7: 751–753.

Denlinger DL (1991) Relationship between cold hardiness and diapause. In: Lee RE Jr, Denlinger DL, editors. Insects at Low Temperature. New York: Chapman and Hall, pp. 174–198.

Hodková M, Hodek I (1994) Control of diapause and supercooling by the retrocerebral complex in Pyrrhocoris apterus . Entomol Exp Appl 70: 237–245.

Hodková M, Hodek I (1997) Temperature regulation of supercooling and gut nucleation in relation to diapause of Pyrrhocoris apterus (Heteroptera). Cryobiology 34: 70–79. PubMed

Sømme L (1982) Supercooling and winter survival in terrestrial arthropods. Comp Biochem Physiol A 73: 519–543.

Storey KB, Storey JM (1991) Biochemistry of cryoprotectants. In: Denlinger DL, Lee RE Jr., editors. Low temperature biology of insects. Cambridge: Cambridge University Press pp., 64–93.

Telfer WH, Kunkel JG (1991) The function and evolution of insect storage hexameres. A Rev Entomol 36: 205–228. PubMed

Brown JJ (1980) Hemolymph protein reserves of diapausing and non-diapausing codling moth larvae, Cydia pomonella (L) (Lepidoptera, Tortricidae). J Insect Physiol 26: 487–491.

Duman JG (2001) Antifreeze and ice nucleator proteins in terrestrial arthropods. Annu Rev Physiol 63: 327–357. PubMed

Duman JG, Bennett V, Sformo T, Hochstrasser R, Barnes BM (2004) Antifreeze proteins in Alaskan insects and spiders. J Insect Physiol 50: 259–266. PubMed

Raymond JA, DeVries AL (1977) Adsorption inhibition as a mechanism of freezing resistance in polar fishes. Proc Nat Acad Sci USA 74: 2589–2593. PubMed PMC

Zachariassen KE, Kristiansen E (2000) Ice nucleation and antinucleation in nature. Cryobilogy 41: 257–279. PubMed

Pullin AS (1996) Physiological relationship between insect diapause and cold tolerance: Coevolution or coincidence? Eur J Entomol 93: 121–129.

Gehrken U (1985) Physiology of diapause in the adult bark beetle, Ips acccuminatus Gyll., studied in relation to cold hardiness. J Insect Physiol 31: 909–916.

Pullin A, Bale JS, Fontaine LR (1991) Physiological aspects of diapause and cold tolerance during overwintering in Pieris brassicae . Physiol Entomol 16: 447–456.

Chen C-P, Denlinger DL, Lee RE (1991) Seasonal variation in generation time, diapause and cold hardiness in a central Ohio population of the flesh fly, Sarcophaga bullata . Ecol Entomol 16: 155–162.

Watanabe M (2002) Cold tolerance and myo-inositol accumulation in overwintering adults of a lady beetle, Harmonia axyridis (Coleoptera: Coccinellidae). Eur J Entomol 99: 5–9.

Ding L, Li Y, Goto M (2003) Physiological and biochemical changes in summer and winter diapause and non-diapause pupae of the cabbage armyworm, Mamestra brassicae L. during long-term cold acclimation. J Insect Physiol 49: 1153–1159. PubMed

Koštál V, Tamura M, Tollarová M, Zahradníčková H (2004) Enzymatic capacity for accumulation of polyol cryoprotectants changes during diapause development in the adult red firebug, Pyrrhocoris apterus . Physiol Entomol 29: 344–355.

Ma R-Y, Hao S-G, Tian J, Sun J-H, Kang L (2006) Seasonal variation in cold hardiness of the Japanese pine sawyer Monochamus alternatus (Coleoptera: Ceranbycydae). Environ Entomol 35: 881–886.

Vesala L, Salminen TS, Koštál V, Zahradníčková H, Hoikkala A (2002) Myo-inositol as a main metabolite in overwintering flies: seasonal metabolomic profiles and cold stress resistance in a northern drosophilid fly. J Exp Biol 215: 2891–2897. PubMed