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Mycofabricated biosilver nanoparticles interrupt Pseudomonas aeruginosa quorum sensing systems.

Singh BR, Singh BN, Singh A, Khan W, Naqvi AH, Singh HB - Sci Rep (2015)

Bottom Line: Transcriptional studies demonstrated that mfAgNPs reduced the levels of LasIR-RhlIR.Further genes quantification analyses revealed that mfAgNPs significantly down-regulated QS-regulated genes, specifically those encoded to the secretion of virulence factors.The results clearly indicated the anti-virulence property of mfAgNPs by inhibiting P. aeruginosa QS signaling.

View Article: PubMed Central - PubMed

Affiliation: Centre of Excellence in Materials Science (Nanomaterials), Z.H. College of Engineering &Technology, Aligarh Muslim University, Aligarh-202002, India.

ABSTRACT
Quorum sensing (QS) is a chemical communication process that Pseudomonas aeruginosa uses to regulate virulence and biofilm formation. Disabling of QS is an emerging approach for combating its pathogenicity. Silver nanoparticles (AgNPs) have been widely applied as antimicrobial agents against human pathogenic bacteria and fungi, but not for the attenuation of bacterial QS. Here we mycofabricated AgNPs (mfAgNPs) using metabolites of soil fungus Rhizopus arrhizus BRS-07 and tested their effect on QS-regulated virulence and biofilm formation of P. aeruginosa. Transcriptional studies demonstrated that mfAgNPs reduced the levels of LasIR-RhlIR. Treatment of mfAgNPs inhibited biofilm formation, production of several virulence factors (e.g. LasA protease, LasB elastrase, pyocyanin, pyoverdin, pyochelin, rhamnolipid, and alginate) and reduced AHLs production. Further genes quantification analyses revealed that mfAgNPs significantly down-regulated QS-regulated genes, specifically those encoded to the secretion of virulence factors. The results clearly indicated the anti-virulence property of mfAgNPs by inhibiting P. aeruginosa QS signaling.

No MeSH data available.


Related in: MedlinePlus

Anti-biofilm activity of mfAgNPs and their effect along with tobramycin on PAO1 biofilms.Images of (A) CV-staining light microscope, (B) SYTO-9 staining CLSM, and (C) uranyl acetate-lead citrate staining SEM biofilms, treated with different mfAgNPs concentrations: (a) 0 μg/mL, (b) 5 μg/mL, (c) 10 μg/mL, (d) 15 μg/mL, and (e) 25 μg/mL for 48 h. (D) Biofilm formation at indicated concentrations of mfAgNPs for 48 h in microtiter plate and detail methodology is mentioned in Supplementary methods. Error bars indicate the SD of 3 measurements. (E,F) Effect of mfAgNPs on tolerance of biofilms to tobramycin was assessed by CLSM and SEM. Images of biofilm formed in flow chambers for 72 h and then treated with indicated concentrations of mfAgNPs for next 24 h and determined the cell viability using Bacterial Cell Viability Kit. (F) Same cells were also processed for SEM analysis and photographed. Treatments include (a) H2O, (b) 350 μg/mL of tobramycin, (c) 25 μg/mL of mfAgNPs, and (d) 350 μg/mL of tobramycin plus 25 μg/mL of mfAgNPs.
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f6: Anti-biofilm activity of mfAgNPs and their effect along with tobramycin on PAO1 biofilms.Images of (A) CV-staining light microscope, (B) SYTO-9 staining CLSM, and (C) uranyl acetate-lead citrate staining SEM biofilms, treated with different mfAgNPs concentrations: (a) 0 μg/mL, (b) 5 μg/mL, (c) 10 μg/mL, (d) 15 μg/mL, and (e) 25 μg/mL for 48 h. (D) Biofilm formation at indicated concentrations of mfAgNPs for 48 h in microtiter plate and detail methodology is mentioned in Supplementary methods. Error bars indicate the SD of 3 measurements. (E,F) Effect of mfAgNPs on tolerance of biofilms to tobramycin was assessed by CLSM and SEM. Images of biofilm formed in flow chambers for 72 h and then treated with indicated concentrations of mfAgNPs for next 24 h and determined the cell viability using Bacterial Cell Viability Kit. (F) Same cells were also processed for SEM analysis and photographed. Treatments include (a) H2O, (b) 350 μg/mL of tobramycin, (c) 25 μg/mL of mfAgNPs, and (d) 350 μg/mL of tobramycin plus 25 μg/mL of mfAgNPs.

Mentions: Microscopic analysis was used to provide initial clue of anti-biofilm property of mfAgNPs, therefore we performed crystal violate (CV) staining assay. A dark CV stained biofilm was observed in untreated control, while a dose-dependent visible reduction in number of microcolonies was examined in mfAgNPs-treated PAO1 biofilm (Fig. 6A-a–e). Additionally, mfAgNPs also disrupted the architecture of biofilm too, as it was more evident from CLSM (Fig. 6B-a–e) and SEM (Fig. 6C-a–e) micrographs. The effect of mfAgNPs on PAO1 biofilm formation was also evaluated using a static biofilm assay. Biofilm formation was inhibited 7–93% by mfAgNPs in a dose dependent manner as shown in Fig. 6D. HF, a positive control was also found to inhibit biofilm formation by 11–96% at the concentrations of 5–25 μg/mL (Supplementary figure 9).


Mycofabricated biosilver nanoparticles interrupt Pseudomonas aeruginosa quorum sensing systems.

Singh BR, Singh BN, Singh A, Khan W, Naqvi AH, Singh HB - Sci Rep (2015)

Anti-biofilm activity of mfAgNPs and their effect along with tobramycin on PAO1 biofilms.Images of (A) CV-staining light microscope, (B) SYTO-9 staining CLSM, and (C) uranyl acetate-lead citrate staining SEM biofilms, treated with different mfAgNPs concentrations: (a) 0 μg/mL, (b) 5 μg/mL, (c) 10 μg/mL, (d) 15 μg/mL, and (e) 25 μg/mL for 48 h. (D) Biofilm formation at indicated concentrations of mfAgNPs for 48 h in microtiter plate and detail methodology is mentioned in Supplementary methods. Error bars indicate the SD of 3 measurements. (E,F) Effect of mfAgNPs on tolerance of biofilms to tobramycin was assessed by CLSM and SEM. Images of biofilm formed in flow chambers for 72 h and then treated with indicated concentrations of mfAgNPs for next 24 h and determined the cell viability using Bacterial Cell Viability Kit. (F) Same cells were also processed for SEM analysis and photographed. Treatments include (a) H2O, (b) 350 μg/mL of tobramycin, (c) 25 μg/mL of mfAgNPs, and (d) 350 μg/mL of tobramycin plus 25 μg/mL of mfAgNPs.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4562228&req=5

f6: Anti-biofilm activity of mfAgNPs and their effect along with tobramycin on PAO1 biofilms.Images of (A) CV-staining light microscope, (B) SYTO-9 staining CLSM, and (C) uranyl acetate-lead citrate staining SEM biofilms, treated with different mfAgNPs concentrations: (a) 0 μg/mL, (b) 5 μg/mL, (c) 10 μg/mL, (d) 15 μg/mL, and (e) 25 μg/mL for 48 h. (D) Biofilm formation at indicated concentrations of mfAgNPs for 48 h in microtiter plate and detail methodology is mentioned in Supplementary methods. Error bars indicate the SD of 3 measurements. (E,F) Effect of mfAgNPs on tolerance of biofilms to tobramycin was assessed by CLSM and SEM. Images of biofilm formed in flow chambers for 72 h and then treated with indicated concentrations of mfAgNPs for next 24 h and determined the cell viability using Bacterial Cell Viability Kit. (F) Same cells were also processed for SEM analysis and photographed. Treatments include (a) H2O, (b) 350 μg/mL of tobramycin, (c) 25 μg/mL of mfAgNPs, and (d) 350 μg/mL of tobramycin plus 25 μg/mL of mfAgNPs.
Mentions: Microscopic analysis was used to provide initial clue of anti-biofilm property of mfAgNPs, therefore we performed crystal violate (CV) staining assay. A dark CV stained biofilm was observed in untreated control, while a dose-dependent visible reduction in number of microcolonies was examined in mfAgNPs-treated PAO1 biofilm (Fig. 6A-a–e). Additionally, mfAgNPs also disrupted the architecture of biofilm too, as it was more evident from CLSM (Fig. 6B-a–e) and SEM (Fig. 6C-a–e) micrographs. The effect of mfAgNPs on PAO1 biofilm formation was also evaluated using a static biofilm assay. Biofilm formation was inhibited 7–93% by mfAgNPs in a dose dependent manner as shown in Fig. 6D. HF, a positive control was also found to inhibit biofilm formation by 11–96% at the concentrations of 5–25 μg/mL (Supplementary figure 9).

Bottom Line: Transcriptional studies demonstrated that mfAgNPs reduced the levels of LasIR-RhlIR.Further genes quantification analyses revealed that mfAgNPs significantly down-regulated QS-regulated genes, specifically those encoded to the secretion of virulence factors.The results clearly indicated the anti-virulence property of mfAgNPs by inhibiting P. aeruginosa QS signaling.

View Article: PubMed Central - PubMed

Affiliation: Centre of Excellence in Materials Science (Nanomaterials), Z.H. College of Engineering &Technology, Aligarh Muslim University, Aligarh-202002, India.

ABSTRACT
Quorum sensing (QS) is a chemical communication process that Pseudomonas aeruginosa uses to regulate virulence and biofilm formation. Disabling of QS is an emerging approach for combating its pathogenicity. Silver nanoparticles (AgNPs) have been widely applied as antimicrobial agents against human pathogenic bacteria and fungi, but not for the attenuation of bacterial QS. Here we mycofabricated AgNPs (mfAgNPs) using metabolites of soil fungus Rhizopus arrhizus BRS-07 and tested their effect on QS-regulated virulence and biofilm formation of P. aeruginosa. Transcriptional studies demonstrated that mfAgNPs reduced the levels of LasIR-RhlIR. Treatment of mfAgNPs inhibited biofilm formation, production of several virulence factors (e.g. LasA protease, LasB elastrase, pyocyanin, pyoverdin, pyochelin, rhamnolipid, and alginate) and reduced AHLs production. Further genes quantification analyses revealed that mfAgNPs significantly down-regulated QS-regulated genes, specifically those encoded to the secretion of virulence factors. The results clearly indicated the anti-virulence property of mfAgNPs by inhibiting P. aeruginosa QS signaling.

No MeSH data available.


Related in: MedlinePlus