Cold tolerance of Littorinidae from southern Africa: Intertidal snails are not constrained to freeze tolerance

All intertidal gastropods for which cold tolerance strategies have been assessed have been shown to be freeze tolerant. Thus, freeze tolerance is considered an adaptation to the intertidal environment. We investigated the cold tolerance strategies of three species of subtropical and temperate snails (Gastropoda: Littorinidae) to determine whether this group is phylogenetically constrained to freeze tolerance. We exposed 'dry' acclimated and 'wet' rehydrated snails to low temperatures to determine temperature of crystallisation (Tc), lower lethal temperature and LT₅₀ and to examine the relationship between ice formation and mortality. Tc was lowest in dry Afrolittorina knysnaensis (-13.6±0.4°C), followed by dry Echinolittorina natalensis (-10.9±0.2°C) and wet A. knysnaensis (-10.2±0.2°C). The Tc of both A. knysnaensis and E. natalensis increased with rehydration, whereas Tc of dry and wet Afrolittorina africana did not differ (-9.6±0.2 and -9.0±0.2°C respectively). Wet snails of all species exhibited no or low survival of inoculative freezing, whereas dry individuals of A. knysnaensis could survive subzero temperatures above -8°C when freezing was inoculated with ice. In the absence of external ice, Afrolittorina knysnaensis employs a freeze-avoidance strategy of cold tolerance, the first time this has been reported for an intertidal snail, indicating that there is no family-level phylogenetic constraint to freeze tolerance. Echinolittorina natalensis and A. africana both showed pre-freeze mortality and survival of some internal ice formation, but were not cold hardy in any strict sense.