Mesophyll conductance (g_m) and stomatal conductance (g_s) are two crucial components of the diffusive limitation of photosynthesis. Variation of g_m in response to CO₂ concentration was evaluated by using two independent methods based on measurements of variable electron transport rate (J) and instantaneous carbon isotope discrimination, respectively. Both methods of g_m estimation showed a very similar shape of the g_m/C_i relationship, with an initial increase at low substomatal CO₂ concentrations (C_i), a peak at 180–200 μmol mol⁻¹C_i, and a subsequent decrease at higher C_i. A good correlation was observed between values of g_m estimated from the two methods, except when C_i <200 μmol mol⁻¹, suggesting that the initial increase of g_m at low C_i was probably due to unreliable estimates over that range of C_i. Plants were also treated with abscisic acid (ABA), which induced a reduction in g_s without significantly affecting the rate of photosynthesis, g_m or the photosynthetic capacity. The present results confirm, using two independent methods, that g_m is strongly sensitive to C_i, and that the relationship between g_s and g_m is not conservative, differing between control and ABA-treated plants.

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 ¹⁸O‐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 ¹⁸O‐liquid water addition) in moist and air‐dry soils. We determined the time kinetics and calculated the average ¹⁸O enrichment of soil water over incubation time, which is necessary to calculate microbial growth from ¹⁸O 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 ¹⁸O‐vapor equilibration method provided similar results as the current method of liquid ¹⁸O‐water addition when used for moist soils. However, when applied to air‐dry soils the liquid ¹⁸O‐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.