The aim of the present study was to analyze the growth and physiological performance of two coexisting species, Trifolium fragiferum, and Trifolium repens, under the effect of NaCl and rhizobial symbiosis. Seeds of T. fragiferum and T. repens were collected from populations in the wild, and plants were cultivated in an automated greenhouse, two plants per container. Three basic types of planting were performed: (1) both plants were T. fragiferum (single species), (2) one T. fragiferum and one T. repens (species coexistence), (3) both plants were T. repens (single species). For every basic type, three subtypes were made: (1) non-inoculated, (2) inoculated with rhizobia taken from T. fargiferum, (3) inoculated with rhizobia taken from T. repens. For every subtype, half of the containers were used as control, and half were treated with NaCl. Shoot fresh mass of plants was significantly (p < 0.001) affected by species coexistence, inoculant, and NaCl. Three significant two-way interactions on plant shoot growth were found: between species coexistence and NaCl (p < 0.001), inoculant and species (p < 0.05), and NaCl and species (p < 0.001). A significant three-way interaction between inoculant, NaCl, and species (p < 0.001) indicated different responses of shoot growth of the two species to inoculant type and NaCl. NaCl treatment was an important factor for T. fragiferum, resulting in better growth in conditions of species coexistence, but the positive effect of bacterial inoculant was significantly more pronounced. A decrease in peroxidase activity in leaves was a good indicator of relative NaCl tolerance, while the absence/presence of rhizobial inoculation was reflected by changes in leaf chlorophyll concentration and photochemical activity of photosystem II. It can be concluded that interaction between biotic and abiotic factors affected the outcome of the coexistence of the two Trifolium species. Distribution of T. fragiferum in sea-affected habitats seems to be related to a higher competitive ability with allied species at increased substrate salinity, based on better physiological salinity tolerance.

The aim of the present study was to establish an experimental system in controlled conditions to study the physiological effect of abiotic/biotic interaction using a rare wild leguminous plant species from coastal sand dunes, Anthyllis maritima. The particular hypothesis tested was that there is an interaction between sand burial, rhizobial symbiosis and salt treatment at the level of physiological responses. Experiment in controlled conditions included 18 treatment combinations of experimental factors, with two intensities of sand burial, rhizobial inoculation and two types of NaCl treatment (soil irrigation and foliar spray). Shoot biomass was significantly affected both by burial and by inoculation, and by interaction between burial and NaCl in the case of shoot dry mass. For plants sprayed with NaCl, burial had a strong significant positive effect on shoot growth irrespective of inoculation. General effect of inoculation with rhizobia on shoot growth of plants without NaCl treatment was negative except for the plants buried 2 cm with sand, where significant stimulation of shoot dry mass by inoculant was found. The positive effect of burial on shoot growth was mainly associated with an increase in leaf petiole height and number of leaves. Performance index significantly increased in buried plants in all treatment combinations, and leaf chlorophyll concentration increased in buried plants independently on burial depth, and only in plants not treated with NaCl. Inoculation led to significant increase of leaf peroxidase activity in all treatment combinations except NaCl-irrigated plants buried for 2 cm by sand. Sand burial stimulated peroxidase activity, mostly in non-inoculated plants, as inoculation itself led to increased enzyme activity. In conclusion, strong interaction between sand burial and NaCl treatment was evident, as the latter significantly affected the effect of burial on growth and physiological indices. Moreover, rhizobial symbiosis had a significant effect on physiological processes through interaction with both sand burial and NaCl treatment, but the effect was rather controversial; it was positive for photosynthesis-related parameters but negative for growth and tissue integrity indices.

Saussurea esthonica is an endangered plant species typical for wet inland habitats such as calcareous fens. Due to its limited population size and distribution, non-invasive sampling of is important in the research of S. esthonica. The aim of the present study was to assess the effect of mineral nutrient availability and substrate moisture on the growth, physiological status, and mineral nutrition of S. esthonica. The non-destructive measurement of physiological parameters was performed in native habitats during three vegetative seasons, followed by two experiments in controlled conditions. Soil at the two Estonian sites had a relatively larger similarity in the composition of plant-available mineral nutrients in comparison to the two Latvian sites. The chlorophyll a fluorescence parameter Performance Index correlated with the total precipitation in the respective month before measurement, but no significant relationship with other environmental variables was found. For mineral nutrient experiments, plants were grown in four substrates with different mineral nutrient composition, resembling that of soil at different S. esthonica sites. Plant growth and physiological indices were significantly affected by the mineral composition of the substrate. Differences in leaf and root mineral nutrient concentrations of S. esthonica plants in part reflected differences in substrate mineral concentration. To evaluate the effect of soil moisture on growth and photosynthesis-associated parameters of S. esthonica, plants were cultivated in “Pope+” substrate at four different moisture treatments (dry, normal, wet, and waterlodged). The most intense growth of S. esthonica plants was evident in waterlodged conditions, which decreased with a decrease in soil moisture. The biomass of leaves increased by 106% and that of the roots increased by 72% as soil moisture increased from dry to normal. For waterlodged plants, leaf biomass increased by 263% and root biomass increased by 566%, in comparison to that for plants cultivated in dry substrate. Substrate drying had a more negative effect on the growth of S. esthonica plants in comparison to that of waterlodging, and this can be directly linked to prevalent hydrological conditions of an alkaline fen habitat native to the species. Therefore, the preservation of the natural water regime in natural habitats is critical to the conservation of the species.

An understudied problem in plant heavy metal biology is the effects of acute versus gradual or chronic metal exposure. The aim of the present study was to compare the growth and physiological responses of Rumex hydrolapathum Huds. plants subjected to gradual or acute Mn stress treatment in controlled conditions. Heavy metal was applied to substrate either as one 1.00 g L⁻¹ Mn dose (acute treatment) or the same dose in four steps of increasing amounts within 12 days (gradual treatment). Peroxidase activity in actively photosynthesizing leaves was used for monitoring induced biochemical changes resulting from Mn treatment. The number of leaves per plant significantly increased in the case of gradual treatment with Mn, but this effect was not statistically significant for acute treatment. Leaf fresh mass significantly decreased in both cases due to the decrease in leaf water content, but dry biomass of leaves was not affected, with no significant differences between the two types of treatments. A significantly lower chlorophyll fluorescence parameter Performance Index in large leaves of plants under the acute Mn treatment than in plants under the gradual treatment was evident. An increase in leaf peroxidase activity by Mn treatment was proportional to the metal dose received, but plants in the acute treatment with 1.00 g L⁻¹ Mn had a significantly lower peroxidase response in comparison to the gradual treatment with 1.00 g L⁻¹ Mn. In conclusion, under gradual treatment, biochemical changes related to the induction of tolerance to the heavy metal are expressed, as indicated by the continuous increase in leaf peroxidase activity after each treatment step.