Colonization of Raphanus sativus by human pathogenic microorganisms.

Contamination of vegetables with human pathogenic microorganisms (HPMOs) is considered one of the most important problems in the food industry, as current nutritional guidelines include increased consumption of raw or minimally processed organic vegetables due to healthy lifestyle promotion. Vegetables are known to be potential vehicles for HPMOs and sources of disease outbreaks. In this study, we tested the susceptibility of radish (Raphanus sativus) to colonization by different HPMOs, including Escherichia coli PCM 2561, Salmonella enterica subsp. enterica PCM 2565, Listeria monocytogenes PCM 2191 and Bacillus cereus PCM 1948. We hypothesized that host plant roots containing bactericidal compounds are less prone to HPMO colonization than shoots and leaves. We also determined the effect of selected pathogens on radish growth to check host plant-microbe interactions. We found that one-week-old radish is susceptible to colonization by selected HPMOs, as the presence of the tested HPMOs was demonstrated in all organs of R. sativus. The differences were noticed 2 weeks after inoculation because B. cereus was most abundant in roots (log10 CFU - 2.54), S. enterica was observed exclusively in stems (log10 CFU - 3.15), and L. monocytogenes and E. coli were most abundant in leaves (log10 CFU - 4.80 and 3.23, respectively). The results suggest that E. coli and L. monocytogenes show a higher ability to colonize and move across the plant than B. cereus and S. enterica. Based on fluorescence in situ hybridization (FISH) and confocal laser scanning microscopy (CLSM) approach HPMOs were detected in extracellular matrix and in some individual cells of all analyzed organs. The presence of pathogens adversely affected the growth parameters of one-week-old R. sativus, especially leaf and stem fresh weight (decreased by 47-66 and 17-57%, respectively). In two-week-old plants, no reduction in plant biomass development was noted. This observation may result from plant adaptation to biotic stress caused by the presence of HPMOs, but confirmation of this assumption is needed. Among the investigated HPMOs, L. monocytogenes turned out to be the pathogen that most intensively colonized the aboveground part of R. sativus and at the same time negatively affected the largest number of radish growth parameters.

Endophytic microbiota and ectomycorrhizal structure of Alnus glutinosa Gaertn. at saline and nonsaline forest sites.

The tolerance of European alder (Alnus glutinosa Gaertn.) to soil salinity can be attributed to symbiosis with microorganisms at the absorptive root level. However, it is uncertain how soil salinity impacts microbial recruitment in the following growing season. We describe the bacterial and fungal communities in the rhizosphere and endosphere of A. glutinosa absorptive roots at three tested sites with different salinity level. We determined the morphological diversity of ectomycorrhizal (ECM) fungi, the endophytic microbiota in the rhizosphere, and the colonization of new absorptive roots in the following growing season. While bacterial diversity in the rhizosphere was higher than that in the absorptive root endosphere, the opposite was true for fungi. Actinomycetota, Frankiales, Acidothermus sp. and Streptomyces sp. were more abundant in the endosphere than in the rhizosphere, while Actinomycetota and Acidothermus sp. dominated at saline sites compared to nonsaline sites. Basidiomycota, Thelephorales, Russulales, Helotiales, Cortinarius spp. and Lactarius spp. dominated the endosphere, while Ascomycota, Hypocreales and Giberella spp. dominated the rhizosphere. The ECM symbioses formed by Thelephorales (Thelephora, Tomentella spp.) constituted the core community with absorptive roots in the spring and further colonized new root tips during the growing season. With an increase in soil salinity, the overall fungal abundance decreased, and Russula spp. and Cortinarius spp. were not present at all. Similarly, salinity also negatively affected the average length of the absorptive root. In conclusion, the endophytic microbiota in the rhizosphere of A. glutinosa was driven by salinity and season, while the ECM morphotype community was determined by the soil fungal community present during the growing season and renewed in the spring.

Selection of microorganisms capable of polyethylene (PE) and polypropylene (PP) degradation.

Microorganisms degrade microplastics, but their potential is still not fully exploited, e.g., due to inadequate selection of microorganisms. We developed an effective selection method of microorganisms capable of polyethylene (PE) and polypropylene (PP) degradation and assessed the scale of polymer degradation by microbial populations. We isolated seven bacterial strains (three Priestia megaterium strains, Klebsiella pneumoniae, Pseudomonas fluorescens, Enterobacter ludwigii, Chryseobacterium sp.) and seven fungal strains (four Fusarium spp., two Lecanicillium spp. and Trichoderma sp.) with PE degradation potential, as well as seven bacterial strains (five Serratia marcescens and two Enterobacter spp.) and six fungal strains (four Aspergillus spp., Fusarium oxysporum and Penicillium granulatum) with PP degradation ability. Scanning electron microscopy (SEM) analysis confirmed the presence of a biofilm and revealed surface changes in both PE and PP pellets, but the greatest changes (microcracks and corrugations) were observed for PP incubated with bacteria. Fourier transform infrared (FTIR) spectroscopy confirmed the structural changes on the studied polymer surfaces. In conclusion, the isolation of plastic-degrading bacteria and fungi from waste landfills represents an effective strategy for the collection of microorganisms with high potential for PE and PP degradation. The bacteria and fungi revealed better potential for PP degradation and PE degradation, respectively.

Pseudomonas stutzeri and Kushneria marisflavi Alleviate Salinity Stress-Associated Damages in Barley, Lettuce, and Sunflower.

Soil salinity is one of the most important abiotic factors limiting plant productivity. The aim of this study was to determine the effect of selected halotolerant plant growth-promoting endophytes (PGPEs, Pseudomonas stutzeri ISE12 and Kushneria marisflavi CSE9) on the growth parameters of barley (Hordeum vulgare), lettuce (Lactuca sativa), and sunflower (Helianthus annuus) cultivated under salt stress conditions. A negative effect of two higher tested salinities (150 and 300 mM NaCl) was observed on the growth parameters of all investigated plants, including germination percentage and index (decreasing compared to the non-saline control variant in the ranges 5.3-91.7 and 13.6-90.9%, respectively), number of leaves (2.2-39.2%), fresh weight (24.2-81.6%); however, differences in salt stress tolerance among the investigated crops were observed (H. annuus > H. vulgare > L. sativa). Our data showed that the most crucial traits affected by endophyte inoculation under salt stress were chlorophyll concentration, leaf development, water storage, root development, and biomass accumulation. Thus, the influence of endophytes was species specific. K. marisflavi CSE9 promoted the growth of all tested plant species and could be considered a universal PGPEs for many plant genotypes cultivated under saline conditions (e.g., increasing of fresh weight compared to the non-inoculated control variant of barley, lettuce, and sunflower in the ranges 11.4-246.8, 118.9-201.2, and 16.4-77.7%, respectively). P. stutzeri ISE12 stimulated growth and mitigated salinity stress only in the case of barley. Bioaugmentation of crops with halotolerant bacterial strains can alleviate salt stress and promote plant growth; however, the selection of compatible strains and the verification of universal plant stress indicators are the key factors.

Choosing source of microorganisms and processing technology for next generation beet bioinoculant.

The increase of human population and associated increasing demand for agricultural products lead to soil over-exploitation. Biofertilizers based on lyophilized plant material containing living plant growth-promoting microorganisms (PGPM) could be an alternative to conventional fertilizers that fits into sustainable agricultural technologies ideas. We aimed to: (1) assess the diversity of endophytic bacteria in sugar and sea beet roots and (2) determine the influence of osmoprotectants (trehalose and ectoine) addition during lyophilization on bacterial density, viability and salt tolerance. Microbiome diversity was assessed based on 16S rRNA amplicons sequencing, bacterial density and salt tolerance was evaluated in cultures, while bacterial viability was calculated by using fluorescence microscopy and flow cytometry. Here we show that plant genotype shapes its endophytic microbiome diversity and determines rhizosphere soil properties. Sea beet endophytic microbiome, consisting of genera characteristic for extreme environments, is more diverse and salt resistant than its crop relative. Supplementing osmoprotectants during root tissue lyophilization exerts a positive effect on bacterial community salt stress tolerance, viability and density. Trehalose improves the above-mentioned parameters more effectively than ectoine, moreover its use is economically advantageous, thus it may be used to formulate improved biofertilizers.

Mixture of Salix Genotypes Promotes Root Colonization With Dark Septate Endophytes and Changes P Cycling in the Mycorrhizosphere.

The roots of Salix spp. can be colonized by two types of mycorrhizal fungi (ectomycorrhizal and arbuscular) and furthermore by dark-septate endophytes. The fungal root colonization is affected by the plant genotype, soil properties and their interactions. However, the impact of host diversity accomplished by mixing different Salix genotypes within the site on root-associated fungi and P-mobilization in the field is not known. It can be hypothesized that mixing of genotypes with strong eco-physiological differences changes the diversity and abundance of root-associated fungi and P-mobilization in the mycorrhizosphere based on different root characteristics. To test this hypothesis, we have studied the mixture of two fundamentally eco-physiologically different Salix genotypes (S. dasyclados cv. 'Loden' and S. schwerinii × S. viminalis cv. 'Tora') compared to plots with pure genotypes in a randomized block design in a field experiment in Northern Germany. We assessed the abundance of mycorrhizal colonization, fungal diversity, fine root density in the soil and activities of hydrolytic enzymes involved in P-mobilization in the mycorrhizosphere in autumn and following spring after three vegetation periods. Mycorrhizal and endophytic diversity was low under all Salix treatments with Laccaria tortilis being the dominating ectomyorrhizal fungal species, and Cadophora and Paraphaeosphaeria spp. being the most common endophytic fungi. Interspecific root competition increased richness and root colonization by endophytic fungi (four taxa in the mixture vs. one found in the pure host genotype cultures) more than by ectomycorrhizal fungi and increased the activities of hydrolytic soil enzymes involved in the P-mineralization (acid phosphatase and ß-glucosidase) in mixed stands. The data suggest selective promotion of endophytic root colonization and changed competition for nutrients by mixture of Salix genotypes.

Bacterial microbiome of root-associated endophytes of Salicornia europaea in correspondence to different levels of salinity.

The halophytes have evolved several strategies to survive in saline environments; however, an additional support from their associated microbiota helps combat adverse conditions. Hence, our driving interests to investigate the endophytic bacterial community richness, diversity, and composition associated to roots of Salicornia europaea from two test sites with different origins of soil salinity. We assumed that salinity will have a negative effect on the diversity of endophytes but simultaneously will permit the high occurrence of halophylic bacteria. Further, to establish the role of the host and its external environment in determining the endophytic diversity, we analyzed the physico-chemical parameters of root zone soil and the concentration of salt ions in the plant roots. The results based on the Miseq Illumina sequencing approach revealed a higher number of endophytic bacterial OTUs at naturally saline test site with a higher level of soil salinity. Proteobacteria and Bacteriodetes were the dominant endophytic phyla at both analyzed sites; additionally, the high occurrence of Planctomycetes and Acidobacteria at more saline site and the occurrence of Firmicutes, Actinobacteria, and Chloroflexi at less saline site were recorded. The salinity in the root zone soil was crucial in structuring the endophytic community of S. europaea, and the significant prevalence of representatives from the phyla Deltaproteobacteria, Acidobacteria, Caldithrix, Fibrobacteres, and Verrucomicrobia at the more saline test site suggest domination of halophylic bacteria with potential role in mitigation of salt stress of halophytes.

Cadmium-induced changes in the production of siderophores by a plant growth promoting strain of Pseudomonas fulva.

The impact of increasing Cd2+ exposure on the quality and quantity of siderophores produced by a plant growth promoting Pseudomonas fulva strain was tested to gain insight into the degree of change. P. fulva was cultured in the increasing concentrations of Cd2+ (0, 0.5, 1.0, 2.0 mM). The secreted siderophores were separated by HPLC and characterized by UHPLC-QTOF/MS. In the presence of 2 mM Cd2+ synthesis of siderophores (hydroxamates, catecholates, phenolates) was mitigated compared to the treatments with lower concentrations of Cd2+ (0.5 and 1 mM). Increased synthesis of catecholates in 0.5 and 1 mM Cd2+ and of phenolates in 0.5-2 mM Cd2+ was revealed compared to the variant without Cd2+ . Out of seven different hydroxamates, the secretion of ferrioxamine E was significantly decreased in the highest Cd2+ concentration. Two additional ferrioxamines, X2 and D2, were secreted independent of the presence or absence of Cd2+ . Exposure to Cd2+ change the composition of siderophores secreted by P. fulva with selective promotion of catecholates and phenolates at the expense of hydroxamates. Successful adaptation in a Cd-contaminated soil in the frame of practical applications to promote phytoremediation can be assumed.

Endophytic and rhizosphere bacteria associated with the roots of the halophyte Salicornia europaea L. - community structure and metabolic potential.

The main objective of our study was to assess density and diversity of rhizosphere (R) and endophytic (E) microorganisms associated with the halophyte S. europaea. Microorganisms were isolated from two saline sites (S1: 55dSm(-1), anthropogenic origin; S2: 112dSm(-1), natural salinity) located in central Poland and compared with microbial populations in the soil (S) using culture-independent (phospholipid fatty acids analysis, PLFA) and culture-dependent techniques. The endophytic and rhizosphere bacteria were identified and screened for nifH and acdS genes, and their metabolic properties were assessed. Strains with the potential to promote plant growth were selected for further study. PLFA analysis revealed that Gram-negative bacteria were dominant at both saline test sites; the total microbial biomass depended on the site (S1E>S). In contrast, culture-dependent techniques revealed that Gram-positive bacteria (Actinobacteria and Firmicutes) were dominant (S1: E-77.3%, R-86.3% and S2: E-59.1%, R-87.5%). Proteobacteria were observed in the rhizosphere at the lowest frequency (S1: 13.7% and S2: 12.5%). Greater salinity decreased the range and specificity of metabolic activity among the endophytes. These conditions also resulted in a broader spectrum of metabolic abilities in rhizobacteria; however, these metabolic processes were present at lower levels.

Metabolic potential and community structure of endophytic and rhizosphere bacteria associated with the roots of the halophyte Aster tripolium L.

The submitted work assumes that the abundance and diversity of endophytic and rhizosphere microorganisms co-existing with the halophytic plant Aster tripolium L. growing in a salty meadow in the vicinity of a soda factory (central Poland) represent unique populations of cultivable bacterial strains. Endophytic and rhizosphere bacteria were (i) isolated and identified based on 16S rDNA sequences; (ii) screened for nifH and acdS genes; and (iii) analyzed based on selected metabolic properties. Moreover, total microbial biomass and community structures of the roots (endophytes), rhizosphere and soil were evaluated using a cultivation-independent technique (PLFA) to characterize plant-microbial interactions under natural salt conditions. The identification of the isolated strains showed domination by Gram-positive bacteria (mostly Bacillus spp.) both in the rhizosphere (90.9%) and roots (72.7%) of A. tripolium. Rhizosphere bacterial strains exhibited broader metabolic capacities, while endophytes exhibited higher specificities for metabolic activity. The PLFA analysis showed that the total bacterial biomass decreased in the following order (rhizosphere

Ectomycorrhizal Community Structure of Salix and Betula spp. at a Saline Site in Central Poland in Relation to the Seasons and Soil Parameters.

Saline stress is one of the most important abiotic factors limiting the growth and development of plants and associated microorganisms. While the impact of salinity on associations of arbuscular fungi is relatively well understood, knowledge of the ectomycorrhizal (EM) fungi of trees growing on saline land is limited. The main objective of this study was to determine the density and diversity of EM fungi associated with three tree species, Salix alba, Salix caprea and Betula pendula, growing in saline soil during two seasons, autumn and spring. The site was located in central Poland, and the increased salinity of the soil was of anthropogenic origin from soda production. The degree of EM colonisation of fine root tips varied between 9 and 34 % and depended on the tree species of interest (S. caprea < S. alba < B. pendula) and season (spring < autumn). Moreover, the ectomycorrhizal colonisation of B. pendula was positively correlated with pH and CaCO3, while for S. caprea and S. alba, colonisation was associated with most of the other soil parameters investigated; e.g. salinity, Corg and N. Analysis of EM fungi revealed four to five different morphotypes per each season: Tomentella sp. Sa-A, Hebeloma collariatum Sc-A, Geopora sp. Sc-A, Helotiales sp. Bp-A in the autumn and Tomentella sp. Sa-S, Tomentella sp. Sc-S and three morphotypes from the families Thelephoraceae and Pyronemataceae in the spring. In conclusion, the density of EM is related to the level of salinity (ECe), season and tree species. Tomentella spp., Hebeloma sp., Geopora sp. and Helotiales sp. are groups of species highly adapted to saline conditions.