High temperatures represent a limitation for growth and development of many crop species. Several studies have demonstrated that the yield reduction of tomato under high temperatures and drought is mainly due to a photosynthetic decline. In this paper, a set of 15 tomato genotypes were screened for tolerance to elevated temperatures by cultivating plants under plastic walk-in tunnels. To assess the potential tolerance of tomato genotypes to high temperatures, measurements of chlorophyll fluorescence, pigments content and leaf functional traits have been carried out together with the evaluation of the final yields. Based on the greenhouse trials, a group of eight putative heat-sensitive and heat-tolerant tomato genotypes was selected for laboratory experiments aimed at investigating the effects of short-term high temperatures treatments in controlled conditions. The chlorophyll fluorescence induction kinetics were recorded on detached leaves treated for 60 min at 35 • C or at 45 • C. The last treatment significantly affected the photosystem II (PSII) photochemical efficiency (namely maximum PSII quantum efficiency, F v /F m, and quantum yield of PSII electron transport, Φ PSII ) and the non-photochemical quenching (NPQ) in the majority of genotypes. The short-term heat shock treatments also led to significant differences in the shape of the slow Kautsky kinetics and its significant time points (chlorophyll fluorescence levels minimum O, peak P, semi-steady state S, maximum M, terminal steady state T) compared to the control, demonstrating heat shock-induced changes in PSII functionality. Genotypes potentially tolerant to high temperatures have been identified. Our findings support the idea that chlorophyll fluorescence parameters (i.e., Φ PSII or NPQ) and some leaf functional traits may be used as a tool to detect high temperatures-tolerant tomato cultivars.Plants 2020, 9, 508 2 of 16 and quality [5,6]. Tomato (Solanum lycopersicum), being an excellent source of health-promoting compounds, is one of the most important crops cultivated worldwide and its heat sensitivity varies among different genotypes [4,7,8]. Generally, the optimal temperature range for photosynthesis is considered to be between 25 • C and 30 • C [6]. The rising of average temperatures due to the ongoing climate change will cause extensive productivity losses in Mediterranean areas, where tomato is traditionally cultivated [9][10][11]. In this framework, it becomes important to perform studies that are able to identify the most promising genotypes able to face heat stress.The relationship between gas exchange and crop yield has been largely studied in tomato, suggesting leaf transpiration as the most reliable indicator for yield prediction under drought [12]. However, beside gas exchange, other photosynthesis related parameters [13] should be taken into account to build a "eco-physiological identity card" for different genotypes.Chlorophyll fluorescence represents a good tool to rapidly and accurately detect plant health status, the occurrence of...