Investigating the effect of foliar spraying of zinc nanoparticles and biostimulants on modulating the effect of water deficit stress in sugar beet by using Integrated Biomarker Response Version 2

The effects of foliar applications of zinc oxide nanoparticles and various biostimulants were studied to alleviate water stress in sugar beet. The experiment used a split-split-plot layout based on a Randomized Complete Block Design (RCBD) with three replications over two growing seasons (2022–2024). The main plot consisted of two irrigation levels: Irrigation after 60 and 120 mm of evaporation was considered normal irrigation (WW) and water deficit stress (WD). Zinc nanoparticle (ZnO-NPs) levels (control, 0.2, and 0.4 mg L−1) were assigned to subplots, and biostimulants (control, chitosan, proline, and chitosan + proline) were assigned to sub-subplots. The results shoewd under WD conditions foliar spraying of 4 mg L−1 of ZnO-NPs increased the chlorophyll b content (22.85%), carotenoid (9.58%), proline content (18.42%), beta-glycine (13.20%), stomatal conductance (33.53%), gibberellin (GA) (9.09%), cytokinin (CK) (13.07%), catalase enzyme activity (CAT) (7.86%), superoxide dismutase (SOD) (25.56%), ascorbate peroxidase (APX) (5.34%), and root yield (RY) (10.48%) and decreased abscisic acid (ABA) (11.24%), malondialdehyde (MAD) (17.33%), and hydrogen peroxide (17.18%) compared to the control. Among biostimulants treatments, application of chitosan + proline under WD conditions increased the content of chlorophyll a (37.44%), chlorophyll b (16.23%), proline (4.87%), beta-glycine (18.09%), GA (7.00%), auxin (IAA) (35.40%), CK (18.03%), CAT (11.42%), APX (19.6%), and RY (11.46%) compared to control, and decreased the content of ABA (24.16%), MAD (9.03%), and hydrogen peroxide (11.50%). In this experiment, the combination of 2 mg L-1 ZnO-NPs with chitosan and proline exhibited a synergistic effect, increasing the content of chlorophyll a and b, relative water content (RWC), SOD, and RY, while reducing ABA. The lowest IBRv2 values were recorded for control + proline, control + chitosan, 2 mg L−1 ZnO-NPs + Control, 4 mg L−1 ZnO-NPs + Control, and 4 mg L−1 ZnO-NPs + Proline treatments. 4 mg L−1 ZnO-NPs + chitosan + proline (I = 0.803) and 4 mg L−1 ZnO-NPs + proline (I = 0.809) showed the smallest increases in MAD content. In terms of RY, the least decrease was observed in the treatments of 4 mg L−1 ZnO-NPs + Chitosan and Proline (I = − 0.492) and 4 mg L−1 ZnO-NPs + Proline (I = − 1.014).