Plant chitinases (EC 3.2.1.14) are considered as typical defense components under various environmental stresses, including heavy metals. In addition, some of them play crucial role in normal plant growth and development. In this work the profile and activities of these enzymes were analyzed to study the variability of defense within soybean plants. For this, two cultivars with contrasting tolerance to metals were exposed to ecologically relevant doses of arsenic and cadmium. Enzyme profiles revealed a spatial distribution of chitinase activities throughout the individual plants, tending to decrease upwards to the top of the plants. Under metal stress, there was a single responsive isoform detected in roots that behaved opposingly in the studied soybean cultivars. In contrast, several isoforms were activated in aboveground tissue, predominantly in mature (older) leaves. Of these, two were identified (21 and 42 kDa) as more specifically involved in defense against metal stress in soybean. The 21 kDa isoform was concluded as possibly contributing to metal tolerance and deserves further investigations at molecular level. Nevertheless, no sound interaction was detected between leaf developmental stage and responsiveness to metals for either of the chitinase isoforms. Further studying the distribution of induced defense within plants is important in understanding the defense strategy of plants against environmental cues including metals.

Plants have a potential for the uptake and accumulation of essential and non-essential trace elements. The ability to take up and tolerate metals varies between and within species as well as between metals. For most metals, the mechanisms involved in plant tolerance, uptake and accumulation are still not fully known and it is not known to what extent the plant response is metal-specific rather than a general stress response. In the present study, the growth response of soybean to Cd, As, Al and NaCl was compared and contrasted to simple sequence repeat (SSR) marker analysis results for Cda1, a dominant gene located in a major quantitative trait locus that regulates Cd accumulation in soybean, to evaluate the hypothesis that general effect patterns are induced by the individual metals. Principal component analysis revealed that the root growth response was most diverse for Al exposure and decreased in the order of Al > As > Cd > NaCl. NaCl did not exert a differentiating effect, indicating response mechanisms similar, at least partially, to metal exposure. The applied stressors yielded a distinguishable pattern of root responses, indicating the potential of such screens to identify agents acting similarly or differently. The SSR marker analysis also facilitated characterization of the Cd accumulation potential of the 22 soybean cultivars studied, and thereby identification of cultivars with potential health risk under cultivation in Cd-contaminated soils.

Heavy metals perturbate water balance in plants and hence impact stomatal aperture. After longer exposure, stomatal development also is affected, and stomatal density and size can change. Two soybean (Glycine max) varieties were experimentally selected for contrasting tolerance to arsenic and cadmium. For these cultivars, natural variability in leaf epidermal cell parameters was detected. Pot plants at first assimilating leaf stage were exposed for 10 days to 5 mg kg-1 soil As 3+ and 50 mg kg-1 soil Cd 2+ (respectively). Metals accumulated primarily in roots and exerted relatively low impact on biomass. Despite this, we observed diverse adjustments of stomata and pavement cells. In cv. Bólyi 44 the stomatal size decreased upon stress treatment, possibly to avoid further water loss. In contrast, the other cultivar Cordoba uses larger stomata that might be advantageous in gaining further resources. The observed responses varied depending on leaf type. In addition, dorsoventral stomatal responses in width, yet undescribed under metal stress, were observed. Our data show that leaf epidermal cell adjustments are flexible components of plant defense even at low metal doses, and possibly help to compromise the structural and functional needs of plant (tissue) under metal stress.

The chlorophyll fluorescence imaging technique is a valuable tool for studying the impact of heavy metal stress in plants. The toxic effects of cadmium (50 mg/kg soil) and arsenic (5 mg/kg soil) on growth and the photosynthetic apparatus of two soybean cultivars (Glycine max (L.) Merr. cvs. Bólyi 44 and Cordoba) were assessed. After 10 days of growth in the contaminated soil, fresh and dry weights of shoots and maximum quantum yield of photosystem II (F v /F m ) for the three types of leaves (UL-unifoliate leaf, TL1-first fully expanded trifoliate leaf, TL2-newly expanding trifoliate leaf) were determined. No statistically significant change in the growth parameters was recorded. In the youngest leaves (TL2) of cultivar Bólyi 44, arsenic caused decrease in F v /F m by 8.6%. In the cultivar Cordoba we recorded the arsenic impact, conversely, having the highest inhibition rate of fluorescence in the oldest leaves (UL decrease of 5.62%). A similar difference in trend of changes in F v /F m as the impact of cadmium was also recorded. With the Bólyi 44 variety, the TL2 leaves showed most sensitive response (a decrease of 10.75%); while in the case of Cordoba variety TL2 leaves showed the highest tolerance (a decrease of 1.2%). The results suggest possible genotypic differences in defense strategy against cadmium and arsenic in the different types of leaves.