Comparative chlorophyll a fluorescence and thylakoid system ultrastructure in chloroplasts of two Italian Cichorium intybus L. cultivars

Red chicory (Cichorium intybus L., Asteraceae) is a prominent and widely grown winter crop in Northern Italy, with Chioggia Precoce and Treviso Precoce as two main varieties cultivated along the eastern coast of Emilia Romagna region. The strong varietal selection has led to different leaf arrangement and orientation in the two cultivars, i.e., mostly plagiotropic in cv. Chioggia and orthotropic in cv. Treviso. These variations influence the angle of solar irradiation of the leaves and, therefore, could hint at possible morpho-functional specializations in the chloroplasts as a side effect of the varietal selection. To verify whether this was the case, we combined chloroplast ultrastructural, chlorophyll a fluorescence and the δ13C isotopic analyses of the two aforementioned varieties. Plants grown in neighbouring fields in Ferrara coast were sampled between December 2023 and January 2024. Transmission electron microscopy revealed that, in mature newly expanded leaves, the chloroplasts in cv. Chioggia were typically elliptical, with a well-developed thylakoid system. In contrast, chloroplasts in cv. Treviso were more globular, exhibiting an extremely heterogeneous internal structure, even within the same cell. The thylakoid system in cv. Treviso was characterized by long single thylakoid arrays and grana of highly variable size, along with small 'thylakoid circles.' The morphometric analysis of grana showed that the granum height (h) in cv. Treviso was greater than in cv. Chioggia, owing to more thylakoids per granum and higher stacking repeat distance (SRD). The cv. Chioggia showed, instead, a higher granum cross-sectional irregularity (GSI), i.e., a prominent tendency of thylakoid disks to slide apart on the lateral plane. Despite such structural features, in-field analysis using a Handy-PEA fluorimeter and a MultispeQ spectrometer, along with modulated chlorophyll a fluorescence quenching, showed no signs of impaired photosynthetic functionality in cv. Treviso. Remarkably, cv. Treviso appeared somehow even more efficient than cv. Chioggia. To support independently this inference, the δ13C isotopic ratio was examined across roots, stems, and leaves, confirming more favourable values in cv. Treviso compared to cv. Chioggia. Overall, the chloroplast structural abnormalities observed in cv. Treviso leaves do not seem to impact negatively the plant’s photosynthetic performance. Instead, these changes may reflect a specific adaptation that arose during the cultivar selection process.