Understanding the response of grassland production and carbon exchange to intra-annual variation in precipitation and nitrogen addition is critical for sustainable grassland management and ecosystem restoration. We introduced growing-season drought treatments of different lengths (15, 30, 45 and 60 day drought) by delaying growing-season precipitation in a long-term nitrogen addition experiment in a low diversity meadow steppe in northeast China. Response variables included aboveground biomass (AGB), ecosystem net carbon exchange (NEE), and leaf net carbon assimilation rate (A). In unfertilized plots drought decreased AGB by 13.7% after a 45 day drought and 31.7% after a 60 day drought (47.6% in fertilized plots). Progressive increases in the drought response of NEE were also observed. The effects of N addition on the drought response of productivity increased as drought duration increased, and these responses were a function of changes in AGB and biomass allocation, particularly root to shoot ratio. However, no significant effects of drought occurred in fertilized or unfertilized plots in the growing season a year after the experiment, N addition did limit the recovery of AGB from severe drought during the remainder of the current growing season. Our results imply that chronic N enrichment could exacerbate the effects of growing-season drought on grassland productivity caused by altered precipitation seasonality under climate change, but that these effects do not carry over to the next growing season.

Vegetation sampling

Vegetation surveys were conducted four times: (1) at the beginning of the drought treatments, on or around May 22, 2017, (2) immediately after each 15, 30, 45, or 60 day drought treatment ended, (3) at the end of the 2017 growing season (Sep. 4), and (4) approximately one year later in mid-August of 2018. In each plot, we surveyed the number of plant species, and counted the number of individuals for each species in 3 randomly placed 0.5 m × 0.5 m quadrats. For each survey, twenty live shoots of L. chinensis were randomly collected from each plot to calculate the proportion of dead leaves.

After the vegetation survey, one of the quadrats was randomly selected for the measurement of aboveground biomass (AGB) by harvesting all aboveground live plant shoots. Harvest quadrats were not allowed to overlap. For belowground biomass (BGB), we washed the roots out of three soil cores with a diameter of 10 cm and a depth of 30 cm. The soil cores were randomly collected in the quadrat used for the harvesting of AGB. The harvested plant and root materials were oven-dried at 70°C to a constant weight (48 hours). Leaf C/N ratio was determined by an elemental analyzer (vario EL cube, Ele mentar, Langenselbold, Germany). The root:shoot (R/S) ratio was calculated as BGB/AGB.

The densities of buds and tillering nodes were measured at the end of the 2017 growing season (Sep. 14). A soil sample with an area of 0.25 m × 0.25 m and a depth of 0.2 m was collected from each plot. The collected soil was gently crushed by hand, and the numbers of buds and tillering nodes of L. chinensis were counted directly in the field to avoid damage of the buds by dehydration.

Leaf carbon exchange measurements

The measurements of leaf net carbon assimilation rate (A) and transpiration rate (E) were conducted from 08:00 to 10:00 am on a sunny day at the end of each drought treatment using a portable infrared gas analyzer (LI-6400, LiCor Inc., Lincoln, NE, USA). Five representative intact L. chinensis leaves (uppermost, fully expanded) were randomly selected in each plot for leaf gas exchange measurements. Photon flux density and CO₂ concentration in the leaf chamber were set at 1500 μmol m^-2 s^-1 and 400 μmol mol^-1, respectively. Leaf chamber temperature was set to match the ambient value.

Ecosystem carbon exchange measurements

Concurrently with leaf-level measurements, we measured short-term net ecosystem CO₂ exchange (NEE) and evapotranspiration (ET) using an infrared gas analyzer (LI-6400, LiCor Inc., Lincoln, NE, USA) attached to a transparent chamber (0.5 m × 0.5 m × 1 m polymethyl methacrylate). To stabilize the transparent chamber, an iron frame (0.5 m × 0.5 m) was fixed in the center of each plot in early 2017. For each measurement, 11 consecutive recordings of CO₂ and H₂O concentration were logged at 10 s intervals during a 2 min period after the chamber was mounted on top of the iron frame, then the slope of the linear regression of CO₂ and H₂O concentration was used to determine NEE and ET . Within the chamber, four small electric fans ran continuously to mix air during the measurement.

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