Lignin is a complex aromatic polymer that constitutes a major fraction of plant biomass and represents a valuable renewable carbon resource. Naturally decaying wood serves as an environmental reservoir of microorganisms capable of degrading lignin. In this study, we isolated and characterized sixteen bacterial strains from decaying Tilia cordata wood using an enrichment culture technique with lignin as the sole carbon source. Taxonomic identification via 16S rRNA gene sequencing revealed microbial diversity spanning the genera Bacillus, Pseudomonas, Stenotrophomonas, and several members of the Enterobacteriaceae family, including Raoultella terrigena isolates. Metagenomic sequencing of the wood substrate revealed an exceptionally rich and balanced bacterial community (Shannon index H′ = 5.07), dominated by Streptomyces, Bradyrhizobium, Bacillus, and Pseudomonas, likely reflecting a specialized consortium adapted to lignin rich late-stage decay. Functional phenotyping demonstrated that all isolates possess ligninolytic potential, evidenced by peroxidase/laccase-type activity through methylene blue decolorization. Dynamic Light Scattering (DLS) and HPLC analyses showed that some isolates, such as Raoultella terrigena MGMM806, effectively depolymerized lignosulfonate into low molecular weight fragments (1.23 nm), while others accumulated intermediate metabolites or completely mineralized the substrate. Growth profiling on monolignol substrates revealed a broad spectrum of catabolic specialization in lignin monomer degradation. The results demonstrate a complex system of metabolic partitioning within a natural bacterial consortium. This collection represents a foundational genetic resource for developing engineered biocatalysts and synthetic microbial communities aimed at the efficient conversion of lignin into valuable aromatic compounds.

Multi-strain consortia are expected to protect plants more reliably than single strains, yet this is rarely tested directly, and the behavior of introduced strains in the field is difficult to follow. We assembled a consortium of Stenotrophomonas rhizophila MGMM118, Bacillus halotolerans MGMM119, Pseudomonas grimontii MGMM120 and Pseudomonas viciae MGMM121 strains isolated from the winter wheat rhizosphere and characterized their hydrolytic activities, auxin production, and analyzed their genomes. The consortium increased shoot and root length of wheat. In biocontrol experiments on barley, the consortium reduced the root rot disease index caused by Fusarium culmorum by approximately ten-fold (5.6% vs. 59.7%), outperforming each individual strain (11.9–17.9%). The consortium also reduced Fusarium root rot in tomato by two-fold. In a field experiment on wheat, strain-specific qPCR detected all introduced microorganisms in the wheat rhizosphere at up to 106 CFU per gram of soil, demonstrating different dynamics for each individual strain. Metabarcoding showed that community structure depended on plant developmental stage rather than on the consortium application. Since metabarcoding could not reliably identify the introduced strains, we suppose that strain-specific qPCR is a more appropriate tool for monitoring of inoculants, particularly in complex soil microbial communities.

Objective: Bacterial biofilms, as a lifestyle strategy for surviving hostile environments, are influenced by their architecture, which is mainly composed of exopolysaccharides. Alginate ensures the structure and biofilm stability, and its biosynthesis is governed by multiple genes, including algA, which encodes enzymes for the alginate precursor. In this study, we evaluated the impact of algA gene deletion on functional biocontrol properties of Pseudomonas putida, including extracellular polymeric substance (EPS), alginate exopolysaccharide production, plant colonization ability, and biocontrol efficacy against Fusarium oxysporum f.sp. radicis-lycopersici (Forl) ZUM2407 in tomato plants. Methods: The extracellular polymeric substance (EPS) was evaluated after precipitation from a 48-h culture grown statically in LB medium covered with cellophane. Alginate content was quantified calorimetrically at 485 nm using a dissolved EPS dissolved in milli-Q water, filtered, and reacted with 80% phenol and H2SO4. The colonization ability was assessed in competitive and non-competitive environments using tomato plants in a gnotobiotic system. The biocontrol efficiency against Forl ZUM2407 was assayed under laboratory conditions. Results: The results obtained demonstrated that algA deletion significantly reduced EPS and alginate production by up to 12.8 ± 1.4% and 5.8 ± 0.2%, respectively. The root colonization ability evaluated in competitive and non-competitive environments demonstrated substantially reduced ability of P. putida PCL1760-ΔalgA, with 69.0 ± 0.4% and 82.5 ± 0.1% decreases in basal and apical tomato part-plants, respectively. Most notably, P. putida PCL1760-ΔalgA was up to 47.01 ± 7.51% less effective in suppressing disease caused by Forl ZUM2407 compared to its wild-type strain, which was up 49.52 ± 8.2% effective. Conclusion: The algA as a critical gene for alginate-mediated biofilm architecture, contributes to root colonization and biocontrol ability in P. putida. These findings extended the critical role for algA in maintaining the plants colonization capacity of PCL1760 and consequent biocontrol performance against Fusarium wilt.

Monitoring individual strains in complex microbial communities remains a fundamental challenge in microbial ecology and biotechnology. Here, we present an integrated pipeline for identifying and validating strain-specific loci (SSL) in four biotechnologically relevant plant growth promoting strains from three genera (Stenotrophomonas, Bacillus, and Pseudomonas). The pipeline applies a two-round specificity-filtering strategy combining whole-genome comparison and high-sensitivity BLASTn validation of revealed strain-specific loci (SSL) against the NCBI nucleotide database. SSL count decreased with increasing Average nucleotide identity (ANIb) of the strains used for the analysis, ranging from one locus in B. halotolerans (ANIb = 98.91%) to 15 loci in S. rhizophila (ANIb = 86.49%). All 25 SSL were universally AT-rich, mainly accessory-genome-associated, with flanking regions enriched in genes of unknown function (34.6%) and mobile genetic elements (19.2%). TaqMan qPCR assays targeting SSL demonstrated high specificity—no target sequences were detected across ten geographically distinct soil samples, nor in a native rhizosphere metagenome—and sensitivity, with limits of detection of 0.01–0.1 pg of genomic DNA. Spike-in experiments in soil yielded method detection limits (MDL) of 850–15,000 CFU/g. All four strains were detected in the wheat rhizosphere seven days after consortium application in a field experiment, validating the pipeline for multi-strain field monitoring.

Microorganisms are fundamental drivers of soil productivity, mediating nutrient cycling and pathogen suppression. In this study, we evaluated changes in the fungal community in the soil of barley (Hordeum vulgare L.) in a field experiment involving the application of a consortium of Paenibacillus pabuli, Priestia megaterium, Pseudomonas koreensis, and Pseudomonas orientalis. Seed pretreatment and seed pretreatment followed by rhizosphere drenching at different growth stages were implemented. Regarding fungal communities in bulk soil, the rhizospheres of untreated and treated plants were characterized based on full-length ribosomal RNA gene (18S-5.8S-28S) metabarcoding sequencing. Despite the compositional shifts, no statistical differences were observed among the alpha diversity metrics. Seed treatment resulted in long-term, targeted suppression of Fusarium graminearum, Fusarium fujikuroi, Fusarium musae, and Fusarium verticillioides from the booting through flowering and dough development stages, outperforming seed pretreatment followed by rhizosphere drenching. A low-modularity network was observed in the rhizosphere of untreated plants. Seed treatment fostered a highly interconnected and uniform network with low hub-betweenness scores. Rhizosphere drenching of pretreated seeds shifted the network topology toward higher hub-betweenness scores, reducing their connectivity by up to 10% in the rhizosphere and bulk soil. These findings provide a framework for optimizing the soil ecosystem for sustainable agriculture.

Bacillus licheniformis species produce a wide array of secondary metabolites and are recognized for their high tolerance to toxins and heavy metals, making them promising bacterial biocontrol agents in agriculture, industry, and medicine. In this study, the complete genome of the Bacillus licheniformis strain MGMM137 isolated from the cooling drilling mud of an oil well was examined. The bioinformatics analysis revealed 4439 predicted coding sequences. The assembled genome was 4269212 bp in size and had a GC content of 45.9 %. During the genomic analysis, numerous genes encoding enzymes homologous to known proteins and capable of breaking down aromatic compounds and their derivatives were found. The strain was also shown to contain open reading frames with high sequence similarity to genes of known proteases, lipases, chitinases, cellulases, and amylases. Some of the identified gene clusters turned out to be homologous to known operons encoding genes involved in the synthesis of metabolites with antimicrobial and fungicidal properties. Furthermore, genes encoding proteins of resistance to many antibiotics were detected. Thus, the isolated and characterized strain can be used as a potential agent for the bioremediation of soils contaminated with sewage or oil waste.