Chlorophyll Pigment and Leaf Macronutrient Trait Variation of Four <i>Salix</i> Species in Elevated CO₂, under Soil Moisture Stress and Fertilization Treatments

Leaf chlorophyll pigment and macronutrient concentrations were quantified for four willow species (Salix cordata (COR), S. discolor (DIS), S. eriocephala (ERI) and S. interior (INT)) while growing under 2 × 2 factorial of ambient and elevated CO2 and well-watered and drought-stressed soil moisture treatments (Expt. 1). After the first year, we saw no difference in stem biomass between CO2 treatments. In the following year, a second experiment was conducted on a subset of well-watered willows as a 2 × 2 factorial of atmospheric CO2 and soil fertilization (FERT). For both years of Expt. 1, chlorophyll a, b, a + b (TCC) and carotenoids (CAR) significantly downregulated in response to elevated CO2 (eCO2) and upregulated in response to drought (DRT). In Expt. 2, FERT mitigated CO2 downregulation of TCC and CAR, and upregulated TCC and CAR. Across species, ERI had the greatest pigment concentrations followed by either COR or DIS. Except for one case, INT had the lowest pigment concentrations. A significant species x FERT interaction was due to magnitude effects. The CHLa:b ratio was not affected by CO2 or DRT but did increase in response to FERT. INT had the greatest CHLa:b ratio followed by DIS, then either ERI or COR. In the second year, TCC:CAR ratio decreased in response to eCO2 and increased in response to DRT and FERT. In Expt. 1, leaf N was the only nutrient to significantly downregulate in response to eCO2; whereas all other nutrient levels remained unchanged. In response to DRT, leaf N and Mg upregulated; whereas leaf P, K, and Ca were downregulated. In response to eCO2 in Expt. 2, again only leaf N downregulated; whereas all other nutrients remained unchanged. All leaf nutrients upregulated in response to FERT. Of the four species, INT had the greatest leaf N and K, and the lowest Ca. Species variation was important, but so to was clonal variation in response to change. Indeed, INT leaf chlorophyll and macronutrients are significantly different or segregated from the other three willow species and this may be related to the evolutionary origins of INT, and other species of the taxonomic section Longifoliae, in the arid southwest USA and Mexico. Furthermore, under low nutrient conditions, it may be necessary to fertilize the plants to see a biomass response to eCO2