Growth and ecophysiology
At the end of both cultivation periods, we counted all surviving
plants to estimate mortality and measured the height of all surviving
plants. On a subset of five plants per population we determined
exchanges of CO2 and water vapour using an open flow
system carrying air with ambient CO2 pressure (41 ± 1.5
Pa) from the Botanical Garden of Pavia University into an infrared gas
analyser (LCA-4, ADC Co., Hoddesdon, UK) connected to a factory-designed
small (625 mm2) leaf chamber. Illumination was
provided by a glasshouse lamp at 450 ± 20 µmol m−2s−1 PPFD and ambient temperature (25 ± 2 °C). Each
gas-exchange value was the mean of 3-5 measurements per plant. The
gas-exchange measurements were used for determining area-based net
CO2 exchange (net photosynthesis, as µmol
CO2 m−2 s−1),
stomatal conductance ( gs, as mol
H2O m−2 s−1) and
water use efficiency (WUE, as µmol CO2mol−1 H2O m−2s−1). Chlorophyll a fluorescence was determined in
three mature healthy leaves by a portable fluorimeter (OS1-FL,
Opti-Sciences, Hudson, NH, United States). Fm was
obtained using a saturating pulse of actinic light after 1 min of
acclimation under which conditions Fo mas measured. The
ratio Fv/Fm [with Fv =
(Fm – Fo)] was used to assess the
photochemical efficiency of photosystem II as a proxy of stress
experienced by the plants during the experimental cultivation.
Chlorophyll (chl) content was determined (as arbitrary units) on a
further subset of 5 plants with a chl content meter (CCM-200; Opti
Sciences, Tyngsboro, MA, USA).