From diel photosynthesis to crop growth in the crassulacean acid metabolism (CAM) orchid Phalaenopsis

Crassulacean acid metabolism (CAM) is a photosynthetic adaption that has evolved in response to water stress and/ or high temperatures. CAM can be found in plants that grow in (semi)-arid habitats, but CAM also appears in plants in the humid tropics, for example, in epiphytes that have no direct access to soil. CAM plants are remarkable at conserving water, because they take up CO2 during the night, which allows them to keep their stomata closed during the hottest part of the day. The aim of this thesis was to increase understanding of CAM physiology by studying the economically important ornamental orchid Phalaenopsis. To reach this objective, Phalaenopsis plants were grown at several combinations of temperature and light. The response to these treatments was determined on different biological scales (from leaf to plant growth) and on different temporal scales (from seconds to the entire period of cultivation).Chapter 1 introduces CAM and describes how the diel cycle of CO2 uptake and CO2 refixation can be captured in four phases. CAM plants are often not considered when looking for future-proof solutions in agriculture, but one option that is being explored is the bio-engineering of CAM into C3 plants, which is also discussed here. The second half of this general introduction focuses on CAM in the orchid Phalaenopsis. The cultivation practice of the economically important Phalaenopsis pot plant is outlined, and the effects of temperature, light and CO2 on Phalaenopsis growth and development are summarized.Chapter 2 introduces a conceptual framework for CAM plants. A literature study revealed that currently no mechanistic models for CAM growth exist. Therefore, a conceptual framework was developed, which consists of three modules, covering 1) CAM photosynthesis, 2) allocation of assimilates among carbon pools, and 3) biomass accumulation among plant organs. The framework covers processes related to CAM physiology on different biological (from leaf to plant) and temporal (from hours to a full cultivation period) scales. It can function as the basis for development of a mechanistic crop growth model for CAM. Development of the framework also showed that Tt is still unclear how carbon is processed and allocated within CAM plants, when diel carbon fluxes are linked to plant growth.Chapter 3 demonstrates that genotypic variation is large between Phalaenopsis hybrids. In two experiments with 19 and 14 genotypes, the effects of temperature and light on vegetative growth traits were studied. Furthermore, the after-effects of these treatments on flowering traits were determined. Hierarchal component analysis provided insight in how traits correlated and how they contributed to growth and development of Phalaenopsis. Plants that were grown at 31°C showed a strong reduction of plant and root growth (biomass accumulation) compared to plants grown at 27°C, while leaf initiation rates increased. An increase in light intensity (from 60 µmol m-2 s-1 to 140 µmol m-2 s-1) accelerated both vegetative plant growth (biomass accumulation) and development (leaf initiation). Both an increase in light intensity, as well as an increase in temperature during the vegetative phase resulted in an increased number of flower spikes, and number of flowers and buds. In this chapter it is demonstrated that vegetative plant traits can be used to predict flowering quality despite genotypic and phenotypic variation. Traits like number of leaves and biomass of the vegetative plant are a good proxy for, respectively, number of inflorescences and inflorescence biomass.Chapter 4 describes physiological differences between genotypes that underpin plant growth and development of Phalaenopsis. Plants from two genotypes were exposed to light and temperature treatments comparable to those in chapter 3. Measurements on gas exchange and carbohydrate analysis within a diel CAM-cycle were combined with measurements of plant growth and development. Per genotype, principal component analysis (PCA) was used to identify which traits explained most of the variation that occurred. Genotypes were found to differ in processes related to all three modules of the framework described in chapter 2. This chapter elucidated that a correlation exists between cumulative diel CO2 uptake and vegetative plant dry weight of Phalaenopsis, but only when leaf area of the plant is considered, not when expressed on a m2 basis.Phase IV is considered important for its substantial contribution to CO2 uptake and to the productivity of CAM plants. Therefore, chapter 5 examines CO2 uptake via C3 and C4 carboxylation in phase IV in the CAM species Phalaenopsis ‘Sacramento’ and Kalanchoe blossfeldiana ‘Saja’. Short blackout periods, switching to 2% O2 and measurements of chlorophyll fluorescence during phase IV all indicated that in Phalaenopsis ‘Sacramento’, PEPC might be the main carboxylase. This is unlike what is known from other CAM species, where Rubisco is the main carboxylase in phase IV. Indeed, results of K. blossfeldiana ‘Saja’ confirmed that Rubisco was the main carboxylase. Additionally, carbohydrate analysis showed that starch accumulated in Phalaenopsis during phase IV which indicated that Rubisco also active as a carboxylase, alongside PEPC. This chapter discusses that having both carboxylases simultaneously active may lead to double carboxylation and futile cycling of CO2, but that it might also serve as a mechanism for photoprotection.Chapter 6 summarizes and reflects upon the concepts and conclusions of the previous chapters. This discussion focusses on how processes that are determined within diel cycles link to plant growth, which is determined over a longer period of time. The methodology that was used for gas exchange measurements is reflected upon. In this chapter, the effect of other environmental factors such as red: far-red and CO2 on Phalaenopsis cultivation are examined. This chapter furthermore described perspectives of the results of this thesis for commercial Phalaenopsis cultivation. Linking together the results of different chapters, it is demonstrated that diel CO2 uptake measured on a leaf can be used as a proxy for flowering quality, and could thus function as an early selection criterium when phenotyping in Phalaenopsis breeding. Additionally, some suggestions are given for future research, building on knowledge produced in this thesis, e.g. how to improve the conceptual framework.