Polarimetric imaging microscopy for advanced inspection of vegetal tissues

Abstract

Optical microscopy techniques for plant inspection benefit from the fact that at least one of the multiple properties of light (intensity, phase, wavelength, polarization) may be modified by vegetal tissues. Paradoxically, polarimetric microscopy although being a mature technique in biophotonics, is not so commonly used in botany. Importantly, only specific polarimetric observables, as birefringence or dichroism, have some presence in botany studies, and other relevant metrics, as those based on depolarization, are underused. We present a versatile method, based on a representative selection of polarimetric observables, to obtain and to analyse images of plants which bring significant information about their structure and/or the spatial organization of their constituents (cells, organelles, among other structures). We provide a thorough analysis of polarimetric microscopy images of sections of plant leaves which are compared with those obtained by other commonly used microscopy techniques in plant biology. Our results show the interest of polarimetric microscopy for plant inspection, as it is non-destructive technique, highly competitive in economical and time consumption, and providing advantages compared to standard non-polarizing techniques.

Polarimetric observables for the enhanced visualization of plant diseases

“…A suitable tool for studying depolarizing information are Mueller polarimeters 23 , 24 , that commonly performs macroscopic analysis of samples. Recent works 4 , 25 , 26 have shown and discussed the advantages of the depolarization-related observables to enhance image contrast between different plant structures of interest, as midrib, secondary veins, stomata and raphides. This is because their constituent units (cellulose, pectin, water content, among others) present different polarimetric features (retardance or dichroism) and/or different spatial organization, these situations inducing depolarization at macroscopic scale observation.…”

“…This is because their constituent units (cellulose, pectin, water content, among others) present different polarimetric features (retardance or dichroism) and/or different spatial organization, these situations inducing depolarization at macroscopic scale observation. Importantly, depolarization methods have proved to be a very interesting tool for the inspection of plant tissues, independently or in combination to other optical instrumentation, as spectroscopic instrumentation, phase or fluorescent microscopy, among others 25 , 27 .…”

“…This metric has been used, for instance, to estimate the chlorophyll content 29 , the plant stress 30 and for vegetation classification purposes 31 . The description of the depolarization provided by the DoP, can be refined with the use of the so-called depolarization metric sets as it has been discussed in recent works 25 , 26 . A depolarization space is a combination of at least three depolarization-related metrics which are derived from the Mueller matrix, to enhance the image contrast in plant structures 4 , 25 , 26 , 32 .…”

“…The description of the depolarization provided by the DoP, can be refined with the use of the so-called depolarization metric sets as it has been discussed in recent works 25 , 26 . A depolarization space is a combination of at least three depolarization-related metrics which are derived from the Mueller matrix, to enhance the image contrast in plant structures 4 , 25 , 26 , 32 . This is the case of the so-called Indices of Polarimetric Purity (IPPs) 33 , 34 , that are connected with the type of polarimetric randomness that a system induces to incident light (they give information of the depolarization anisotropy of the system, i.e., dependence of the input polarization with the resultant depolarization response).…”

“…In particular we show how depolarizing metrics are suitable for the discrimination between healthy tissues and different type and stages of plant infections. The depolarization set of metrics chosen for the present study is the IPPs set because, as stated before, it provides three suitable metrics for plant visualization 25 , 26 . In addition to IPPs, we include in the study, for completeness, a second set of depolarizing observables: the so-called Components of Purity (CPs) 36 .…”