## 2 Materials and Methods
### Isolation of Bradyrhizobium Bacteria from Commercial Preparations  
In order to isolate nitrogen-fixing bacteria, 10 g of inoculated peat and 10 g of gel were weighed from the commercial preparations Nitragina and Nitroflora, respectively, and transferred into 100 mL of pyrophosphate. The resulting suspensions were shaken on a shaker at 150 rpm for 20 min and then diluted. 0.2 mL of the dilution of 10\({}^{8}\) of each preparation was applied with a cell spreader on a YMA medium [18] and incubated at 28 \({}^{\circ}\)C for 7 days. A single colony was collected with a sterile inoculation loop from each of the Petri dishes and it was streaked onto the YMA medium again. The resulting isolates were then inoculated onto YMA slants and used for further laboratory investigations.  
Interactions between Symbiotic Bacteria (Bradyrhizobium sp.) and Endophytes (Pseudomonas fluorescens or Bacillus subtilis)  
In order to determine interactions between the rhizobia and endophytic bacteria, liquid cultures of all strains were prepared in the first stage. Three-day-old _Bradyrhizobium_ bacterial cultures (isolated from the Nitragina or Nitroflora preparations) as well as _Pseudomonas fluorescens_ and _Bacillus subtilis_ were transferred several times on agar slants dedicated to each bacterial species--YMA [18] for _Bradyrhizobium_ sp., agars for _Bacillus subtilis_ bacteria according to [PERSON] [16] and King B [17] for _Pseudomonas fluorescens_, were used to make a suspension by adding 5 mL of saline into each test tube. Next, 0.1 mL of the bacterial suspension was used to inoculate 100 mL of the YMA liquid medium for _Bradyrhizobium_ sp., the medium prepared according to [PERSON] for _Bacillus subtilis_, and [PERSON] for _Pseudomonas fluorescens_ (five flasks for each variant).  
The bacterial cultures were incubated at 28 \({}^{\circ}\)C and shaken on a shaker at 70 rpm for 96 h, 48 h, and 48 h for _Bradyrhizobium_ sp., _Bacillus subtilis_, and _Pseudomonas fluorescens_, respectively.  
The ring method was used for the analysis of interactions between the microorganisms used in the research (28) in eight experimental variants (five replicates in each):  
1. _Bradyrhizobium_ sp. (isolated from the Nitragina preparation) on _Pseudomonas fluorescens_;
2. _Bradyrhizobium_ sp. (isolated from the Nitragina preparation) on _Bacillus subtilis_;
3. _Pseudomonas fluorescens_ on _Bradyrhizobium_ sp. (isolated from the Nitragina preparation);
4. _Bacillus subtilis_ on _Bradyrhizobium_ sp. (isolated from the Nitragina preparation);
5. _Bradyrhizobium_ sp. (isolated from the Nitroflora preparation) on _Pseudomonas fluorescens_;
6. _Bradyrhizobium_ sp. (isolated from the Nitroflora preparation) on _Bacillus subtilis_;
7. _Pseudomonas fluorescens_ on _Bradyrhizobium_ sp. (isolated from the Nitroflora preparation);
8. _Bacillus subtilis_ on _Bradyrhizobium_ sp. (isolated from the Nitroflora preparation).  
A total of 5 mL of 2% agar was poured onto sterile Petri dishes for each of the abovementioned experimental variants. After it solidified, three sterile rings (diameter 10 mm) were applied with sterile tweezers into each Petri dish. Next, 10 mL of the mixture of the dissolved medium characteristic of a given bacterial species with their appropriate dilution (10 \(\times\) 10\({}^{5}\)) was poured so that it would not get inside the rings. When it cooled down, the rings were removed from the dishes under sterile conditions. A total of 0.1 mL of the prepared bacterial suspension was applied into the places of two holes, while saline was placed in the third hole as the control variant. After one hour, the plates were placed in a thermostat at 28 \({}^{\circ}\)C and incubated for 72 h without inversion. After this time interactions between the bacteria were read.