Responses of chlorophyll fluorescence to CO ₂ elimination as an indicator of Crassulacean acid metabolism photosynthesis

Crassulacean acid metabolism (CAM) is found in a wide variety of vascular plant species, mainly those inhabiting water-limited environments. Identifying and characterizing diverse CAM species enhances our understanding of the physiological, ecological, and evolutionary significance of CAM photosynthesis. In this study, we examined the effect of CO ₂ elimination on chlorophyll fluorescence-based photosynthetic parameters in two constitutive CAM Kalanchoe species and six orchids. In CAM-performing Kalanchoe species, the effective quantum yield of photosystem II showed no change in response to CO ₂ elimination during the daytime but decreased with CO ₂ elimination at dusk. We applied this method to reveal the photosynthetic mode of epiphytic orchids and found that Gastrochilus japonicus , Oberonia japonica , and Bulbophyllum inconspicuum , but not Bulbophyllum drymoglossum , are constitutive CAM. Although B. drymoglossum had relatively high malate content in leaves, they did not depend on it to perform photosynthesis even under water deficient or high light conditions. Anatomical comparisons revealed a notable difference in the leaf structure between B. drymoglossum and B. inconspicuum ; B. drymoglossum leaves possess the large water storage tissue internally, unlike B. inconspicuum leaves, which develop pseudobulbs. Our data propose a novel approach to identify and characterize CAM plants without labor-intensive experimental procedures.