Two selected examples of PIXE microanalysis in ecophysiology are presented. Studies of heavy metal distributions in mycorrhizal and non-mycorrhizal roots of Plantago lanceolata showed different filtration mechanisms of Zn/Pb and Fe/Mn, both enabling plants to cope with metals present in the environment. Studies of the mechanism used by the beetle Chrysolina pardalina to eliminate excessive amounts of Ni revealed that Malpighian tubules are responsible for the elimination of this metal from the hemolymph.In both examples GeoPIXE software was used for true elemental mapping using the Dynamic Analysis method and analysis of spectra from selected micro-areas. Specimen thickness and matrix composition were obtained from proton backscattering spectra.

The aim of this study was to determine elemental composition of sap-feeding insects inhabiting various parts of the Ni hyperaccumulating plant Berkheya coddii Roessl., the endemic species of ultramafic outcrops in Mpumalanga, South Africa. Three species were examined: the aphid Protaphis pseudocardui (Aphididae), abundant on young leaves; the mealybug Orthesia sp. (Ortheziidae) colonizing underground parts of this plant, and the bug Norialsus berkheyae (Cixiidae) living on young shoots. Maps of Ni, K, Ca, Zn, and Fe for selected body areas of these species were generated using Dynamic Analysis method on the basis of particle-induced X-ray emission (micro-PIXE) and proton backscattering (BS) measurements. Atomic absorption spectrometry was used to determine Ni, Zn, Cu, Fe contents in the B. coddii organs, in some sapfeeding insect species including these mentioned above, and in the assassin bug hunting on Chrysolina pardalina, a monophagous beetle of B. coddii. Bioaccumulation factor for Ni in the examined species was below 0.05, and much higher for other metals (Zn ‡ 2; Fe £ 5). Ni distribution within body was species-dependent. It was the highest in the antennae of P. pseudocardui, in the head of Orthesia sp. and in the metathorax of N. berkheyae. Distribution patterns of other metals were different among examined species. Ca was recorded mainly in peripheral parts of the body in all species. Zn showed similar distribution to Ni. Fe distribution was similar to Ni only in the mealybugs. Uneven concentrations of metals within selected body regions indicated their relations with specific organs. Analysis of Ni transfer to higher trophic levels was done on the basis of two food nets: B. coddii-C. pardalina-Rhinocoris neavii and B. coddii-P. pseudocardui-Polyrhachis ant and led to the conclusion that the role of sap-feeding insects in Ni transfer was marginal.

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The middle region of the digestive system, the midgut of freshwater shrimp Neocaridina davidi is composed of a tube-shaped intestine and the hepatopancreas formed by numerous caeca. Two types of cells have been distinguished in the intestine, the digestive cells (D-cells) and regenerative cells (R-cells). The hepatopancreatic tubules have three distinct zones distinguished along the length of each tubule—the distal zone with R-cells, the medial zone with differentiating cells, and the proximal zone with F-cells (fibrillar cells) and B-cells (storage cells). Fasting causes activation of cell death, a reduction in the amount of reserve material, and changes in the mitochondrial membrane potential. However, here we present how the concentration of ROS changes according to different periods of fasting and whether re-feeding causes their decrease. In addition, the activation/deactivation of mitochondrial superoxide dismutase (MnSOD) was analyzed. The freshwater shrimps Neocaridina davidi (Crustacea, Malacostraca, Decapoda) were divided into experimental groups: animals starved for 14 days, animals re-fed for 4, 7, and 14 days. The material was examined using the confocal microscope and the flow cytometry. Our studies have shown that long-term starvation increases the concentration of free radicals and MnSOD concentration in the intestine and hepatopancreas, while return to feeding causes their decrease in both organs examined. Therefore, we concluded that a distinct relationship between MnSOD concentration, ROS activation, cell death activation and changes in the mitochondrial membrane potential occurred.

Autophagy is a natural process that aims to eliminate malfunctioning cell parts, organelles or molecules under physiological conditions. It is also induced in response to infection, starvation or oxidative stress to provide energy in case of an energy deficit. The aim of this 2-dimensional study was to test if, and if so, how, this process depends on the concentration of cadmium in food (with Cd concentrations from 0 to 352 μg of Cd per g of food (dry weight)—D1 dimension) and the history of selection pressure (160 vs 20 generations of exposure to Cd—D2 dimension). For the study, the 5th instar larvae of a unique strain of the moth Spodoptera exigua that was selected for cadmium tolerance for 160 generations (44 μg of Cd per g of food (dry weight)), as well as 20-generation (11, 22 and 44 μg of Cd per g of food (dry weight)) and control strains, were used. Autophagy intensity was measured by means of flow cytometry and compared with life history parameters: survivability and duration of the 3rd larval stage. The highest values of autophagy markers were found in the groups exposed to the highest Cd concentration and corresponded (with a significant correlation coefficient) to an increased development duration or decreased survivorship in the respective groups. In conclusion, autophagy is probably initiated only if any other defense mechanisms, e.g., antioxidative mechanisms, are not efficient. Moreover, in individuals from pre-exposed populations, the intensity of autophagy is lower.