Factors contributing to ice nucleation and sequential freezing of leaves in wheat

D P Livingston,T Tuong,L V Gusta,T Artlip,M Wisniewski,R Tisdale,A Bertrand

doi:10.1007/s00425-021-03637-w

Abstract

Main conclusionAnatomical, metabolic and microbial factors were identified that contribute to sequential freezing in wheat leaves and likely contribute to supercooling in the youngest leaves and potentially meristematic regions.Infrared thermography (IR) has been used to observe wheat leaves freezing independently and in an age-related sequence with older leaves freezing first. To determine mechanisms that might explain this sequence of freezing several analytical approaches were used: (1) The size of xylem vessels, in proximity to where freezing initiated, was measured to see if capillary freezing point depression explained sequential freezing. The sequence of freezing in the four youngest leaves was correlated, with the largest vessels freezing first. (2) Carbohydrate and amino acids were analyzed to determine if solute concentrations as well as interactions with membranes explained the freezing sequence. Sucrose was highly correlated to the freezing sequence for all leaves suggesting a prominent role for this sugar as compared to other simple sugars and fructans. Among individual free amino acids proline and serine were correlated to the freezing sequence, with younger leaves having the highest concentrations. (3) Microflora within and on leaf surfaces were determined to measure potential freezing initiation. Levels of bacteria and fungi were correlated to the freezing sequence for all leaves, and species or genera associated with high ice nucleation activity were absent in younger leaves. Moisture content and transcript expression of ice binding proteins were also measured. As expected, our results show that no single mechanism explains the freezing sequence observed via infrared analyses. While these multiple mechanisms are operative at different levels according to the leaf age, they seem to converge when it comes to the protection of vital meristematic tissues. This provides potential phenotypic characters that could be used by breeders to develop more winter-hardy genotypes.

Highlights

Winter cereals such as wheat, barley, rye and oats are planted in the fall and harvested the following spring or summer depending on growing conditions
A previous study using IR thermography demonstrated that freezing in wheat occurs in two stages with a third stage that was not visible in IR but was implied, since plants fully recovered after stages 1 and 2 but died if the temperature was reduced beyond the second stage (Livingston et al 2018)
The goal of the present study was to identify anatomical, metabolic and microbial factors that contribute to the sequential freezing in wheat leaves to protect young meristematic tissues by supercooling

Summary

Introduction

Winter cereals such as wheat, barley, rye and oats are planted in the fall and harvested the following spring or summer (or fall in northern regions) depending on growing conditions. Much of the complexity involved in freezing in plants has been underscored by observations of freezing using infrared (IR) thermographic imaging that can monitor ice propagation between and within plants (Wisniewski et al 1997, 2015; Livingston 2018). This technology has demonstrated the role of different factors in extrinsic and intrinsic ice nucleation, including the role of ice-nucleating bacteria, thick, waxy cuticles, stomata location, and antifreeze proteins

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