Two-fold red excess (TREx): a simple and novel digital color index that enables non-invasive real-time monitoring of green-leaved as well as anthocyanin-rich crops.

Background: Digital color indices provide a reliable means for assessing plant status by enabling real-time estimation of chlorophyll (Chl) content, and are thus adopted widely for crop monitoring. However, as all prevalent leaf color indices used for this purpose have been developed using green-leaved plants, they do not perform reliably for anthocyanin (Anth)-rich red-leaved varieties. Hence, the present study investigates digital color indices for six types of leafy vegetables with different levels of Anth to identify congruent trends that could be implemented universally for non-invasive crop monitoring irrespective of species and leaf Anth content. For this, datasets from three digital color spaces, viz., RGB (Red, Green, Blue), HSV (Hue, Saturation, Value), and L*a*b* (Lightness, Redness-greenness, Yellowness-blueness), as well as various derived plant color indices were compared with Anth/Chl ratio and SPAD Chl meter readings of n = 320 leaf samples.

Results: Logarithmic decline of G/R, G-minus-R, and Augmented Green-Red Index (AGRI) with increasing Anth/Chl ratio (R² > 0.8) revealed that relative Anth content affected digital color profile markedly by shifting the greenness-redness balance until the Anth/Chl ratio reached a certain threshold. Further, while most digital color features and indices presented abrupt shifts between Anth-rich and green-leaved samples, the proposed color index Two-fold Red Excess (TREx) did not exhibit any deviation due to leaf Anth content and showed better correlation with SPAD readings (R² = 0.855) than all other color features and vegetation indices.

Conclusion: The present study provides the first in-depth assessment of variations in RGB-based digital color indices due to high leaf Anth contents, and uses the data for Anth-rich as well as green-leaved crops belonging to different species to formulate a universal digital color index TREx that can be used as a reliable alternative to handheld Chl meters for rapid high-throughput monitoring of green-leaved as well as red-leaved crops.