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Identification and control of endophytic bacteria during in vitro cultures of Staphylea pinnata L.

Bożena Szewczyk-Taranek

Bożena Szewczyk-Taranek

Department of Ornamental Plants and Garden Arts, University of Agriculture in KrakowKraków, Poland
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Szewczyk-Taranek, Bożena
, 
Anita Jaglarz

Anita Jaglarz

Moredun Research InstituteMidlothian,
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Jaglarz, Anita
, 
Piotr Pałka

Piotr Pałka

Department of Ornamental Plants and Garden Arts, University of Agriculture in KrakowKraków, Poland
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Pałka, Piotr
, 
Paulina Supel

Paulina Supel

Department of Plant Biology and Biotechnology, University of Agriculture in KrakowKraków, Poland
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Supel, Paulina
, 
Paweł Kaszycki

Paweł Kaszycki

Department of Plant Biology and Biotechnology, University of Agriculture in KrakowKraków, Poland
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Kaszycki, Paweł
, 
Justyna Mazur

Justyna Mazur

Department of Ornamental Plants and Garden Arts, University of Agriculture in KrakowKraków, Poland
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Mazur, Justyna
 and 
Bożena Pawłowska

Bożena Pawłowska

Department of Ornamental Plants and Garden Arts, University of Agriculture in KrakowKraków, Poland
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Pawłowska, Bożena
  
Apr 29, 2020
Identification and control of endophytic bacteria during in vitro cultures of Staphylea pinnata L.'s Cover Image
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Article Category: Research Article
Published Online: Apr 29, 2020
Page range: 47 - 55
Received: May 28, 2019
Accepted: Jan 05, 2020
DOI: https://doi.org/10.2478/fhort-2020-0005
Keywords
© 2020 Bożena Szewczyk-Taranek et al., published by Sciendo
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.

Figure 1

Schematic presentation of the experimental procedures in the study.
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).
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).

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.
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.

Basic bioinformatics parameters characterised by 16S rRNA sequencing analysis of the endogenous bacteria isolated from the Staphylea pinnata tissue culture

Sample codePrimerThe strain with the highest compatibilityCompatibility (%)NCBI access number
16–3163FAcinetobacter johnsonii100NR_117624.1
1387RStrain ATCC 1790999
16–3563FMethylobacterium rhodesianum100NR_041028.1
1387RStrain DSM 5687100

PCR reaction mixture for amplification of gene coding 16S

Volume (μl)Final concentration
GoTaq Green Master mix 2x251×
Upstream primer 63F51 μM
Downstream primer 1387R51 μM
Genomic DNA5< 250 ng
Nuclease-free water10–
Total50

Antibiotics susceptibility test with agar disc-diffusion method performed on bacterial strains isolated from in vitro cultures of Staphylea pinnata

Antibiotic classAntibioticAJ*MR
β-lactamsPenicillinR**S
AmoxicillinRS
Ampicillin+sulbactamRS
AmpicillinRS
PiperacillinSS
TikarcillinSS
CefuroximeRS
TetracyclinesDoksycyclineSS
TetracyclineSS
FluoroquinolonesCiprofloxacinSS
LevofloxacinSS
NorfloxacinSS
OfloxacinSS
AminoglycosidesAmikacinaSS
GentamicinSS
NetilmicinSS
TobramycinSS
Macrolides, Lincosamides, StreptograminsAzithromycinRS
ClarithromycinRS
ErythromycinRS
ClindamycinRR
OthersChloramphenicolRS
NitrofurantoinRR
TrimethoprimRR
Co-trimoxazoleSR
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