Adaptation of plants and microbes to drought stress affects various biochemical processes such as enzyme activities in the rhizosphere. Nonetheless, it is largely unknown how this adaptation changes the spatial distribution of enzyme activities and catalytic properties of enzymes in the rhizosphere. Also, it is very important to know about the spatial pattern of enzyme activities in seminal and lateral roots regions to clarify how the interaction of drought and root types affects rhizosphere extent and enzyme activities. To fill these knowledge gaps, we grew wheat plants under drought (30% WHC, three weeks) and optimum (70% WHC) conditions and then coupled soil zymography with enzyme kinetics for studying β-glucosidase (GLU), acid phosphatase (ACP), and leucine aminopeptidase (LAP). The results showed that drought considerably decreased the hotspot percentage of GLU and ACP activities by 63% and 67% in comparison with optimum moisture. In the regions of the seminal root, the rhizosphere extent of GLU and LAP, normalized rhizosphere extent, and total activity of GLU, ACP, and LAP were higher in optimum than in drought treatment. For the lateral root regions, no difference in the rhizosphere extent and total enzyme activities was found between drought and optimum moisture. Besides, we found higher normalized total enzyme activity in lateral roots regions than in seminal roots regions. Drought elevated the maximum enzymatic reaction velocity (Vmax) and Michaelis–Menten constant (Km) of GLU whereas dropped the Vmax and Km (non-significant) of ACP and LAP in comparison with optimum moisture. The catalytic efficiency of GLU and ACP were 37% and 27% higher in optimum than in drought treatment. Our results revealed that effects of drought on rhizosphere extent and total enzyme activities in lateral roots regions were weaker than in seminal roots regions. Drought decreased microbial hotspots area, rhizosphere expansion of enzyme activities in seminal roots regions, and catalytic efficiency of enzyme systems. In conclusion, drought decreases root biomass and root exudations in the rhizosphere, leading to narrower microbial hotspots area and rhizosphere extent and thus to less enzyme catalytic efficiency.

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