FLIP:FLuorescence imaging pipeline for field-based chlorophyll fluorescence images

Authors: Herritt, Matthew; Long, Jacob; Roybal, Michael; Moller Jr, David; MOCKLER, TODD - Danforth Plant Science Center; PAULI, DUKE - University Of Arizona; Thompson, Alison

Submitted to: SoftwareX
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/1/2021
Publication Date: 3/31/2021
Citation: Herritt, M.T., Long, J.C., Roybal, M.D., Moller Jr, D.C., Mockler, T.C., Pauli, D., Thompson, A.L. 2021. FLIP:FLuorescence imaging pipeline for field-based chlorophyll fluorescence images. SoftwareX. 14. Article 100685. https://doi.org/10.1016/j.softx.2021.100685.
DOI: https://doi.org/10.1016/j.softx.2021.100685

Interpretive Summary: Photosynthesis is one of the most important biological reactions on earth providing oxygen and food for humanity. As global populations rise and arable land decreases, crops need to become more efficient at photosynthetic processes, particularly utilizing absorbed light energy. Measuring chlorophyll fluorescence allows researchers to understand how efficiently light is utilized by photosynthesis. The fluorescence imaging pipeline (FLIP) was created to automate the image analysis of he chlorophyll fluorescence imaging(CIF)system, associated with the TERRA-REF field scanalyzer, by extracting the chlorophyll fluorescence traits from plant tissue and combining the data together in a workable format for downstream analysis. FLIP was created specifically for the TERRA-REF CFI system; however, the pipeline is applicable to other CFI systems obtaining canopy level fluorescence images. The automated pipeline removes background pixels from the images and provides an average fluorescence of the total plant tissue. FLIP allowed researchers to process over 1.5 TB of chlorophyll fluorescence image data in approximately 20 hours.

Technical Abstract: Photosynthesis is one of the most important biological reactions on earth providing oxygen and food for humanity. As global populations rise and arable land decreases, crops need to become more efficient at photosynthetic processes, particularly utilizing absorbed light energy. Chlorophyll fluorescence imaging is a rapid, non-destructive measurement that can provide information on the efficiency of the light-dependent reactions of photosynthesis. Over the years chlorophyll fluorescence imaging systems have been developed and improved to capture two critical measurements: minimum fluorescence (F0) and maximum fluorescence (FM). These systems have primarily been utilized in controlled chamber or greenhouse settings focused at the single leaf or small plant level. To improve plant photosynthesis, fluorescence imaging data needs to be obtained from field-grown plants to capture canopy spatial effects. Previously developed software to extract F0 and FM from controlled, leaf level images do not capture the complexity of the light-dependent reactions from field-grown plants. New software is needed that accounts for the canopy spatial effects from images of field grown-plants. FLIP: fluorescence imaging pipeline, was designed specifically for the TERR-REF field scanalyzer located at the University of Arizona’s Maricopa Agricultural Center located in Maricopa, Arizona but could be adapted for other field deployed fluorescence imaging systems. FLIP utilizes open source tools to convert binary images, apply a multi-threshold to extract the fluorescence data from the plant canopy, calculate photosynthetic efficiency, and assign those values to the appropriate experimental plot.