Modern highly insulated greenhouses are more energy efficient than conventional types. Furthermore applying dynamic greenhouse climate control regimes will increase energy efficiency relatively more in modern structures. However, this combination may result in higher air and crop temperatures. Too high temperature affects the plant photosynthetic responses, resulting in a lower rate of photosynthesis. To predict and analyse physiological responses as stress indicators, two independent experiments were conducted, to detect the effect of high temperature on photosynthesis: analysing photosystem II (PSII) and stomatal conductance (gs). A combination of chlorophyll a fluorescence, gas exchange measurements and infrared thermography was applied using Chrysanthemum (Dendranthema grandiflora Tzvelev) ‘Coral Charm’ as a model species. Increasing temperature had a highly significant effect on PSII when the temperature exceeded 38 °C for a period of 7 (±1.8) days. High temperature decreased the maximum photochemical efficiency of PSII (Fv/Fm), the conformation term for primary photochemistry (Fv/Fo) and performance index (PI), as well as increased minimal fluorescence (Fo). However, at elevated CO2 of 1000 μmol mol−1 and with a photosynthetic photon flux density (PPFD) of 800 μmol m−2 s−1, net photosynthesis (Pn) reached its maximum at 35 °C. The thermal index (IG), calculated from the leaf temperature and the temperature of a dry and wet reference leaf, showed a strong correlation with gs. We conclude that 1) chlorophyll a fluorescence and a combination of fluorescence parameters can be used as early stress indicators as well as to detect the temperature limit of PSII damage, and 2) the strong relation between gs and IG enables gs to be estimated non-invasively, which is an important first step in modelling leaf temperature to predict unfavourable growing conditions in a (dynamic) semi closed greenhouse.