Some plants are able to thrive and complete their life cycle in (extreme) saline environment. We call them halophytes, as oppose to glycophytes, plants with no or very poor tolerance to salts. There is much scientific effort put into uncovering mechanisms of salt tolerance in halophytes and finding ways to develop salt tolerant crops.

I’ve had opportunity to study one of the halophytes, namely Bassia indica. The work was performed within SPIRIT project (http://www.spirit-ion.eu/) through collaboration with prof. Avi Golan from Israel. The B. indica plant exhibits extraordinary root growth toward saline patches in soil [1] and has been proposed as salt phytoremediation plant [2]. In this post I want to share some of the outcomes of the work performed on the B. indica leaves, additional to that already published [3].

B. indica leaves have Kranz anatomy, meaning that they consist of two photosynthetic cell types: bundle sheath cells, which surround the vascular centres, and the mesophyll cells, which surround the bundle sheath cells. In B. indica, the bundle sheath cells create a semi-circle to surround the vascular bundle on one side and they border the mesophyll cells on the other. Water storage tissue is located in the centre of the leaf lamina and indicates the succulent nature of the leaves. You can clearly see these tissues on the leaf cross-sections (photos taken and samples prepared at the Chair of Botany and Plant Physiology, Department of Biology, Biotechnical Faculty, University of Ljubljana, Slovenia):

morph
Cross-section of B. indica leaves. “UV” and “Blue” images are auto-fluorescence after excitation with 470 nm for “UV” and with 546 nm for “Blue”.

Freeze-dried cross-sections of leaves were analysed using micro-Proton Induced X-Ray Emission (Micro-PIXE) at Jožef Stefan Institute, Ljubljana, Slovenia. Quantitative mineral element distribution maps were generated in PyMca (http://pymca.sourceforge.net/) based on numerical matrices from GeoPIXE II software (http://nmp.csiro.au/Geopixe.html).

distribu
Quantitative mineral element spatial distribution in B. indica leaves.

Cell-type specific distribution is easy to extract from these quantitative distribution maps. Water storage tissue and epidermis contain the highest concentrations of Na and K, the mesophyll is rich in Cl and K, bundle sheath cells contain most of Mg, while vascular bundles have the highest concentrations of P, S, Cl and Fe. Hotspots in Ca distribution map could be assigned to Ca oxalate crystals.

Note the extreme numbers in the colour bars; these represent concentrations. These concentrations alone are proof of exceptional salt tolerance in this plant species.

In some cases it is difficult to discern specific mineral element distributions, especially between neighbouring tissues (e.g. bundle sheath cells and vascular bundle). Thus, we often use co-localisation maps, where individual mineral element distribution maps are overlain. In these co-localisation maps each mineral element is represented by one colour (red, green or blue). The level of co-localisation is indicated by a change of colour when two basic colours are mixed in the following manner:

trikotnik
This colour triangle is helpful for understanding co-localisation maps with co-localised pixels having intermediate colours.

Here is a combination of Na distribution (red), K distribution (green) and different mineral elements (blue) as indicated. Through these co-localisation it is much easier to appreciate  cell-type specific distribution of mineral elements.

coloc_Na_K
Mineral element co-localisation in B. indica leaves.

And other combinations:

coloc_Na and others

I hope these photos convinced you of the beautiful world of the plants on the inside I like to explore so much.

References:

[1] Shelef et al. 2010, Plant Biosystems 144, 471-478.

[2] Shelef et al. 2012, Water Research 46(13), 3967–3976.

[3] Pongrac et al. 2013, Nuclear Instruments and Methods in Physics Research Section B: Beam Interactions with Materials and Atoms 306, 144-149.

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