Bacillus amyloliquefaciens—Derived Lipopeptide Biosurfactants Inhibit Biofilm Formation and Expression of Biofilm-Related Genes of Staphylococcus aureus
Abstract
:1. Introduction
2. Results
2.1. Genotypic Identification of the Isolate
2.2. Presence of Surfactin, Fengycin and Iturin A Genes
2.3. Ultrahigh-Performance Liquid Chromatography
2.4. MALDI-TOF/MS Analysis
2.5. The Effect of LPs 3/22 on S. aureus CCM 4223 Biofilm Formation
2.6. Analysis of qRT-PCR Results
3. Discussion
4. Materials and Methods
4.1. Microorganisms
4.2. Isolation and Screening of Isolates for BSs Production
4.3. Genotypic Identification
4.4. Detection of the LP Genes
4.5. Isolation of Biosurfactants
4.6. Ultrahigh-Performance Liquid Chromatography
4.7. MALDI-TOF/MS Analysis
4.8. Determination of Growth Inhibition Activity of LPs 3/22 against S. aureus in Planktonic Cells
4.9. Effect of LPs on Biofilm Formation
4.10. Quantification of Gene Expression Using qRT-PCR
4.11. Statistical Analysis
5. Conclusions
Author Contributions
Funding
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Gene | GenBank Sequence Identity/Accession Number |
---|---|
16S rRNA | B. amyloliquefaciens (100%) |
KF811045.1 | |
srfAA | B. subtilis (97.95%) |
KC454625.1 | |
sfp | B. amyloliquefaciens (99.56%) |
KX346253.1 | |
fenB | NA |
fenD | B. amyloliquefaciens (96.63%) |
KP453873.1 | |
ituD | B. amyloliquefaciens (99.36%) |
FJ815155.1 |
Activity [%] | Concentration of LPs 3/22 [mg/mL] | |||
---|---|---|---|---|
15 | 1.5 | 0.15 | 0.015 | |
Inhibition of biofilm formation | 100.19 ± 4.14 | 84.46 ± 4.21 | 60.09 ± 2.06 | 38.84 ± 13.76 |
Biosurfactants | Gene | Sequence | PCR Product Size [bp] | Annealing Temperature [°C] |
---|---|---|---|---|
Surfactin | sfp | F-5′ATGAAGATTTACGGAATTTA 3′ R-5′TTATAAAAGCTCTTCGTACG 3′ | 675 | 50 |
srfAA | F-5′TCGGGACAGGAAGACATCAT 3′ R-5′CCACTCAAACGGATAATCCTGA 3′ | 201 | 60 | |
Fengycin | fenB | F-5′CCTGGAGAAAGAATATACCGTACCY 3′ R-5′GCTGGTTCAGTT KGATCACAT 3′ | 670 | 57 |
fenD | F-5′GGCCCGTTCTCTAAATCCAT 3′ F-5′GTCATGCTGACGAGAGCAAA 3′ | 269 | 60 | |
Iturin A | ituD | F-5′ TTGAAYGTCAGYGCSCCTTT 3′ R-5′ TGCGMAAATAATGGSGTCGT 3′ | 482 | 57 |
Gene | Primer | Primer Sequence (5′-3′) | Reference |
---|---|---|---|
fnbA | fnbA-F | GAAGTGGCACAGCCAAGAAC | This study |
fnbA-R | ACGTTGACCAGCATGTGG | ||
fnbB | fnbB-F | CAATGATCCTATCATTGAGAAGAGTG | This study |
fnbB-R | CCTTCTACACCTTCAACAGCTGTA | ||
srtA | srtA-F | GTGGTACTTATCCTAGTGGCAGC | This study |
srtA-R | GCCTGCCACTTTCGATTTATC | ||
icaA | icaA-F | CTTGCTGGCGCAGTCAATAC | [82] |
icaA-R | GTAGCCAACGTCGACAACTG | ||
icaD | icaD-F | ACCCAACGCTAAAATCATCG | [83] |
icaD-R | GCGAAAATGCCCATAGTTTC | ||
icaB | icaB-F | ATACCGGCGACTGGGTTTAT | [83] |
icaB-R | ATGCAAATCGTGGGTATGTGT | ||
icaC | icaC-F | CTTGGGTATTTGCACGCATT | [83] |
icaC-R | GCAATATCATGCCGACACCT | ||
agrA | agrA-F | TCGTAAGCATGACCCAGTTG | This study |
agrA-R | AAATCCATCGCTGCAACTTT | ||
gyrB | gyrB-F | CCAGGTAAATTAGCCGATTGC | [84] |
gyrB-R | ATCGCCTGCGTTCTAGAGTC |
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Englerová, K.; Bedlovičová, Z.; Nemcová, R.; Király, J.; Maďar, M.; Hajdučková, V.; Styková, E.; Mucha, R.; Reiffová, K. Bacillus amyloliquefaciens—Derived Lipopeptide Biosurfactants Inhibit Biofilm Formation and Expression of Biofilm-Related Genes of Staphylococcus aureus. Antibiotics 2021, 10, 1252. https://doi.org/10.3390/antibiotics10101252
Englerová K, Bedlovičová Z, Nemcová R, Király J, Maďar M, Hajdučková V, Styková E, Mucha R, Reiffová K. Bacillus amyloliquefaciens—Derived Lipopeptide Biosurfactants Inhibit Biofilm Formation and Expression of Biofilm-Related Genes of Staphylococcus aureus. Antibiotics. 2021; 10(10):1252. https://doi.org/10.3390/antibiotics10101252
Chicago/Turabian StyleEnglerová, Karolína, Zdenka Bedlovičová, Radomíra Nemcová, Ján Király, Marián Maďar, Vanda Hajdučková, Eva Styková, Rastislav Mucha, and Katarína Reiffová. 2021. "Bacillus amyloliquefaciens—Derived Lipopeptide Biosurfactants Inhibit Biofilm Formation and Expression of Biofilm-Related Genes of Staphylococcus aureus" Antibiotics 10, no. 10: 1252. https://doi.org/10.3390/antibiotics10101252