Quantifying microbial growth and carbon use efficiency in dry soil environments via ¹⁸ O water vapor equilibration

Soil microbial physiology controls large fluxes of C to the atmosphere, thus, improving our ability to accurately quantify microbial physiology in soil is essential. However, current methods to determine microbial C metabolism require liquid water addition, which makes it practically impossible to measure microbial physiology in dry soil samples without stimulating microbial growth and respiration (namely, the "Birch effect"). We developed a new method based on in vivo ¹⁸ O-water vapor equilibration to minimize soil rewetting effects. This method allows the isotopic labeling of soil water without direct liquid water addition. This was compared to the main current method (direct ¹⁸ O-liquid water addition) in moist and air-dry soils. We determined the time kinetics and calculated the average ¹⁸ O enrichment of soil water over incubation time, which is necessary to calculate microbial growth from ¹⁸ O incorporation in genomic DNA. We tested isotopic equilibration patterns in three natural and six artificially constructed soils covering a wide range of soil texture and soil organic matter content. We then measured microbial growth, respiration and carbon use efficiency (CUE) in three natural soils (either air-dry or moist). The proposed ¹⁸ O-vapor equilibration method provided similar results as the current method of liquid ¹⁸ O-water addition when used for moist soils. However, when applied to air-dry soils the liquid ¹⁸ O-water addition method overestimated growth by up to 250%, respiration by up to 500%, and underestimated CUE by up to 40%. We finally describe the new insights into biogeochemical cycling of C that the new method can help uncover, and we consider a range of questions regarding microbial physiology and its response to global change that can now be addressed.