Seasonal differences in freezing stress resistance of needles of Pinus nigra and Pinus resinosa: evaluation of the electrolyte leakage method.

Seasonal changes in freezing stress resistance of needles of red pine (Pinus resinosa Ait.) and Austrian pine (Pinus nigra Arnold) trees were measured by an electrolyte leakage method and by visual observation. During most of the year, freezing stress resistance determined by the two methods gave similar results. The electrolyte leakage method provided a good estimate of seasonal changes in freezing stress resistance except for red pine needles in their most winter-hardy state. To obtain a reliable estimate of freezing stress resistance in winter-hardy red pine needles it was necessary to combine the electrolyte leakage method with visual observations. When red pine needles survived exposure to -80 degrees C or lower, electrolyte leakage was never more than 30% even when the needles were exposed to a slow freeze-thaw stress of -196 degrees C. However, rapid freezing of red pine needles to -196 degrees C resulted in electrolyte leakage of over 80%. Red pine needles attained a much higher freezing stress resistance during the winter than Austrian pine. Red pine needles also acclimated and deacclimated faster than Austrian pine needles. An index of injury was developed based on the electrolyte leakage method ((R(2) + R(1))/2, where R(1) is the minimum % electrolyte leakage from noninjured tissue and R(2) is the maximum % electrolyte leakage at the highest injury) that reliably predicted freezing stress resistance of pine needles for most of the year. Important aspects for developing a successful index of injury for pine needles are: use of cut needles, vacuum infiltration and shaking during incubation in water.We conclude that: (1) during cold acclimation the cell wall properties of the pine needles changed and these changes, which appeared to differ in the two species, might explain the very low leakage of electrolytes from winter-hardy needles of red pine; (2) pine needles survive winter by developing the ability to tolerate extracellular ice formation, because after rapid freezing the needles were severely injured; and (3) red pine is adapted to a shorter growing season and colder winters than Austrian pine.