Figure 1
Schematic presentation of the experimental procedures in the study.
Figure 2
Bacterial contamination in in vitro culture of Staphylea pinnata (A); Gram-staining test: Acinetobacter johnsonii colony (B); Methylobacterium rhodesianum colony (C); susceptibility test with agar disc-diffusion method: A. johnsoni (D); M. rhodesioanum (E).
Table 1
PCR reaction mixture for amplification of gene coding 16S
| Volume (μl) | Final concentration | |
|---|---|---|
| GoTaq Green Master mix 2x | 25 | 1× |
| Upstream primer 63F | 5 | 1 μM |
| Downstream primer 1387R | 5 | 1 μM |
| Genomic DNA | 5 | < 250 ng |
| Nuclease-free water | 10 | – |
| Total | 50 |
Table 2
Basic bioinformatics parameters characterised by 16S rRNA sequencing analysis of the endogenous bacteria isolated from the Staphylea pinnata tissue culture
| Sample code | Primer | The strain with the highest compatibility | Compatibility (%) | NCBI access number |
|---|---|---|---|---|
| 16–31 | 63F | Acinetobacter johnsonii | 100 | NR_117624.1 |
| 1387R | Strain ATCC 17909 | 99 | ||
| 16–35 | 63F | Methylobacterium rhodesianum | 100 | NR_041028.1 |
| 1387R | Strain DSM 5687 | 100 |
Table 3
Antibiotics susceptibility test with agar disc-diffusion method performed on bacterial strains isolated from in vitro cultures of Staphylea pinnata
| Antibiotic class | Antibiotic | AJ* | MR |
|---|---|---|---|
| β-lactams | Penicillin | R** | S |
| Amoxicillin | R | S | |
| Ampicillin+sulbactam | R | S | |
| Ampicillin | R | S | |
| Piperacillin | S | S | |
| Tikarcillin | S | S | |
| Cefuroxime | R | S | |
| Tetracyclines | Doksycycline | S | S |
| Tetracycline | S | S | |
| Fluoroquinolones | Ciprofloxacin | S | S |
| Levofloxacin | S | S | |
| Norfloxacin | S | S | |
| Ofloxacin | S | S | |
| Aminoglycosides | Amikacina | S | S |
| Gentamicin | S | S | |
| Netilmicin | S | S | |
| Tobramycin | S | S | |
| Macrolides, Lincosamides, Streptogramins | Azithromycin | R | S |
| Clarithromycin | R | S | |
| Erythromycin | R | S | |
| Clindamycin | R | R | |
| Others | Chloramphenicol | R | S |
| Nitrofurantoin | R | R | |
| Trimethoprim | R | R | |
| Co-trimoxazole | S | R | |
Figure 3
Effect of antibiotics on Staphylea pinnata in vitro culture: D100, D200 and D300—Doxycycline at concentration 100 to 300 mg ∙ dm−3 in the multiplication medium; G100, D200 and G300—Gentamicin at concentration 100–300 mg ∙ dm−3 in the multiplication medium; Control—without antibiotics. *Mean values with the same letter do not differ significantly at α = 0.05, shoots with visible bacteria and shoots without visible bacteria were analysed separately.