pew300  (Applied Photophysics)

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Physicochemical properties of the PEW300 peptide. (A) The sequences of PEW300 and Cecropin A (CeA). The red font and underline sites indicate mutation sites of the PEW300 peptide. (B) Predicted alpha-helical wheel of PEW300 peptide ( https://heliquest.ipmc.cnrs.fr/cgi-bin/ComputParams.py ). The hydrophobic residues are yellow, positively charged hydrophilic residues are blue, and non-charged polar residues are gray. (C) Purification of PEW300 peptide. Line 1 and line 2 are purified PEW300 peptides. (D) The secondary structure of PEW300 predicted by SWISS-MODEL ( https://swissmodel.expasy.org/interactive ). (E) CD spectra of the PEW300 peptide.

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques: Mutagenesis, Purification

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Hemolytic activity, cytotoxicity, and drug resistance of PEW300. (A) Hemolytic activity of PEW300 in mouse red blood cells. 0.1% Triton X-100 was used as the positive control (PC); *** p < 0.005 compared with PC group. (B) Cytotoxicity of PEW300 in HEK293 cells was determined by a CCK-8 assay. 0.1% Triton-X 100 and PBS were used as positive control (PC) and negative control (NC), respectively. Error bars represent the standard error from mean cell viabilities from three replicates, *** indicates p < 0.005 compared with PC group. (C) Drug resistance of the PEW300 and gentamicin. GEN, gentamicin.

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques: Activity Assay, Positive Control, CCK-8 Assay, Negative Control

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Antibacterial and antibiofilm properties of PEW300. (A) Antibacterial activity of PEW300, kanamycin (Kan), and gentamicin (GEN) against Pseudomonas aeruginosa JCM5962. (B) Inhibition and (C) dispersion activity of PEW300 against P. aeruginosa JCM5962 biofilms. Error bars represent standard error from mean values as determined by three repeated experiments, *** indicates p < 0.005 and ns means no significant difference compared with Kan-treated groups.

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques: Activity Assay, Inhibition

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Antibiofilm pathway and time-killing kinetic analysis of PEW300 against Pseudomonas aeruginosa . (A) Antibiofilm pathway of PEW300 against P. aeruginosa . Reductive effect of PEW300 against P. aeruginosa biofilm (B) and planktonic cells (C) .

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques:

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: SEM observation of preformed biofilm of Pseudomonas aeruginosa JCM5962 affected by PEW300. The red arrow indicates the damaged JCM5962 cells.

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques:

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Eliminative action of PEW300 on Pseudomonas aeruginosa mature biofilm. (A) Fluorescence analysis of PEW300 on P. aeruginosa mature biofilm components. The scale bar is 200 μm. (B) Quantification analysis of mature biofilms components of P. aeruginosa JCM5962 affected by PEW300. * Indicates statistical significance compared to the control groups ( * p < 0.05; *** p < 0.005).

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques: Fluorescence

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Disruption effect of PEW300 on Pseudomonas aeruginosa membrane. (A) TEM micrographs of P. aeruginosa JCM5962 treated with or without PEW300 (Control). (B) Flow cytometry analysis of the disruption ability of PEW300 on P. aeruginosa membrane.

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques: Flow Cytometry

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Membrane analysis and intracellular disturbance induced by PEW300. (A) Effects of PEW300 on OM permeabilization of Pseudomonas aeruginosa JCM5962. Control represents cells with no PEW300 treatment. (B) LPS-binding affinities of PEW300. Control means the growth of P. aeruginosa JCM5962 with no PEW300 and LPS incubation. * p < 0.05, *** p < 0.005 compared with control. (C) The IM depolarization induced by PEW300. (D) The electrophoretic mobility shift analysis of the interaction between P. aeruginosa JCM5962 genomic DNA (200 ng) and PEW300 peptide. (E) Intracellular ROS intensity produced by P. aeruginosa JCM5962 in the presence of PBS, 1% Triton X-100, and series concentrations of PEW300 peptide. * p < 0.05, *** p < 0.005 compared with PBS-treated group.

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques: Binding Assay, Incubation, Electrophoretic Mobility Shift Assay, Produced

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Analysis of Pseudomonas aeruginosa virulence factors affected by PEW300. PEW300 results in decreased virulence of P. aeruginosa JCM5962 in A549 cells characterized by (A) CCK-8 assay and (B) calcein-AM staining. *** p < 0.005 compared with P. aeruginosa JCM5962-treated group. Scale bar is 100 μm. Comparative analysis of (C) pyocyanin content, (D) elastase activity and (E) alginates content in P. aeruginosa JCM5962 treated with or without PEW300 peptide. (F) qPCR analysis of virulence-related gene expression. Data represent the mean ± SD from three independent experiments performed in triplicate. * indicates p < 0.05, *** indicates p < 0.005; ns means no significant difference compared with control.

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques: CCK-8 Assay, Staining, Activity Assay, Expressing

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Schematic illustration of the (A) antibiofilm pathway and (B) the multiple actions of PEW300 against Pseudomonas aeruginosa .

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques:

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Physicochemical properties of the PEW300 peptide. (A) The sequences of PEW300 and Cecropin A (CeA). The red font and underline sites indicate mutation sites of the PEW300 peptide. (B) Predicted alpha-helical wheel of PEW300 peptide ( https://heliquest.ipmc.cnrs.fr/cgi-bin/ComputParams.py ). The hydrophobic residues are yellow, positively charged hydrophilic residues are blue, and non-charged polar residues are gray. (C) Purification of PEW300 peptide. Line 1 and line 2 are purified PEW300 peptides. (D) The secondary structure of PEW300 predicted by SWISS-MODEL ( https://swissmodel.expasy.org/interactive ). (E) CD spectra of the PEW300 peptide.

Article Snippet: The Circular dichroism (CD) spectra of PEW300 were measured on a Chirascan qCD Spectrometer (Applied Photophysics, United Kingdom) with wavelengths ranging from 190 to 260 nm using a 1 mm path length cuvette in 30 mM SDS and double-distilled water, respectively.

Techniques: Mutagenesis, Purification, Circular Dichroism

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Hemolytic activity, cytotoxicity, and drug resistance of PEW300. (A) Hemolytic activity of PEW300 in mouse red blood cells. 0.1% Triton X-100 was used as the positive control (PC); *** p < 0.005 compared with PC group. (B) Cytotoxicity of PEW300 in HEK293 cells was determined by a CCK-8 assay. 0.1% Triton-X 100 and PBS were used as positive control (PC) and negative control (NC), respectively. Error bars represent the standard error from mean cell viabilities from three replicates, *** indicates p < 0.005 compared with PC group. (C) Drug resistance of the PEW300 and gentamicin. GEN, gentamicin.

Article Snippet: The Circular dichroism (CD) spectra of PEW300 were measured on a Chirascan qCD Spectrometer (Applied Photophysics, United Kingdom) with wavelengths ranging from 190 to 260 nm using a 1 mm path length cuvette in 30 mM SDS and double-distilled water, respectively.

Techniques: Activity Assay, Positive Control, CCK-8 Assay, Negative Control

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Antibacterial and antibiofilm properties of PEW300. (A) Antibacterial activity of PEW300, kanamycin (Kan), and gentamicin (GEN) against Pseudomonas aeruginosa JCM5962. (B) Inhibition and (C) dispersion activity of PEW300 against P. aeruginosa JCM5962 biofilms. Error bars represent standard error from mean values as determined by three repeated experiments, *** indicates p < 0.005 and ns means no significant difference compared with Kan-treated groups.

Article Snippet: The Circular dichroism (CD) spectra of PEW300 were measured on a Chirascan qCD Spectrometer (Applied Photophysics, United Kingdom) with wavelengths ranging from 190 to 260 nm using a 1 mm path length cuvette in 30 mM SDS and double-distilled water, respectively.

Techniques: Activity Assay, Inhibition, Dispersion

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Antibiofilm pathway and time-killing kinetic analysis of PEW300 against Pseudomonas aeruginosa . (A) Antibiofilm pathway of PEW300 against P. aeruginosa . Reductive effect of PEW300 against P. aeruginosa biofilm (B) and planktonic cells (C) .

Article Snippet: The Circular dichroism (CD) spectra of PEW300 were measured on a Chirascan qCD Spectrometer (Applied Photophysics, United Kingdom) with wavelengths ranging from 190 to 260 nm using a 1 mm path length cuvette in 30 mM SDS and double-distilled water, respectively.

Techniques:

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: SEM observation of preformed biofilm of Pseudomonas aeruginosa JCM5962 affected by PEW300. The red arrow indicates the damaged JCM5962 cells.

Article Snippet: The Circular dichroism (CD) spectra of PEW300 were measured on a Chirascan qCD Spectrometer (Applied Photophysics, United Kingdom) with wavelengths ranging from 190 to 260 nm using a 1 mm path length cuvette in 30 mM SDS and double-distilled water, respectively.

Techniques:

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Eliminative action of PEW300 on Pseudomonas aeruginosa mature biofilm. (A) Fluorescence analysis of PEW300 on P. aeruginosa mature biofilm components. The scale bar is 200 μm. (B) Quantification analysis of mature biofilms components of P. aeruginosa JCM5962 affected by PEW300. * Indicates statistical significance compared to the control groups ( * p < 0.05; *** p < 0.005).

Article Snippet: The Circular dichroism (CD) spectra of PEW300 were measured on a Chirascan qCD Spectrometer (Applied Photophysics, United Kingdom) with wavelengths ranging from 190 to 260 nm using a 1 mm path length cuvette in 30 mM SDS and double-distilled water, respectively.

Techniques: Fluorescence

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Disruption effect of PEW300 on Pseudomonas aeruginosa membrane. (A) TEM micrographs of P. aeruginosa JCM5962 treated with or without PEW300 (Control). (B) Flow cytometry analysis of the disruption ability of PEW300 on P. aeruginosa membrane.

Article Snippet: The Circular dichroism (CD) spectra of PEW300 were measured on a Chirascan qCD Spectrometer (Applied Photophysics, United Kingdom) with wavelengths ranging from 190 to 260 nm using a 1 mm path length cuvette in 30 mM SDS and double-distilled water, respectively.

Techniques: Disruption, Membrane, Flow Cytometry

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Membrane analysis and intracellular disturbance induced by PEW300. (A) Effects of PEW300 on OM permeabilization of Pseudomonas aeruginosa JCM5962. Control represents cells with no PEW300 treatment. (B) LPS-binding affinities of PEW300. Control means the growth of P. aeruginosa JCM5962 with no PEW300 and LPS incubation. * p < 0.05, *** p < 0.005 compared with control. (C) The IM depolarization induced by PEW300. (D) The electrophoretic mobility shift analysis of the interaction between P. aeruginosa JCM5962 genomic DNA (200 ng) and PEW300 peptide. (E) Intracellular ROS intensity produced by P. aeruginosa JCM5962 in the presence of PBS, 1% Triton X-100, and series concentrations of PEW300 peptide. * p < 0.05, *** p < 0.005 compared with PBS-treated group.

Article Snippet: The Circular dichroism (CD) spectra of PEW300 were measured on a Chirascan qCD Spectrometer (Applied Photophysics, United Kingdom) with wavelengths ranging from 190 to 260 nm using a 1 mm path length cuvette in 30 mM SDS and double-distilled water, respectively.

Techniques: Membrane, Binding Assay, Incubation, Electrophoretic Mobility Shift Assay, Produced

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Analysis of Pseudomonas aeruginosa virulence factors affected by PEW300. PEW300 results in decreased virulence of P. aeruginosa JCM5962 in A549 cells characterized by (A) CCK-8 assay and (B) calcein-AM staining. *** p < 0.005 compared with P. aeruginosa JCM5962-treated group. Scale bar is 100 μm. Comparative analysis of (C) pyocyanin content, (D) elastase activity and (E) alginates content in P. aeruginosa JCM5962 treated with or without PEW300 peptide. (F) qPCR analysis of virulence-related gene expression. Data represent the mean ± SD from three independent experiments performed in triplicate. * indicates p < 0.05, *** indicates p < 0.005; ns means no significant difference compared with control.

Article Snippet: The Circular dichroism (CD) spectra of PEW300 were measured on a Chirascan qCD Spectrometer (Applied Photophysics, United Kingdom) with wavelengths ranging from 190 to 260 nm using a 1 mm path length cuvette in 30 mM SDS and double-distilled water, respectively.

Techniques: CCK-8 Assay, Staining, Activity Assay, Expressing

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Schematic illustration of the (A) antibiofilm pathway and (B) the multiple actions of PEW300 against Pseudomonas aeruginosa .

Article Snippet: The Circular dichroism (CD) spectra of PEW300 were measured on a Chirascan qCD Spectrometer (Applied Photophysics, United Kingdom) with wavelengths ranging from 190 to 260 nm using a 1 mm path length cuvette in 30 mM SDS and double-distilled water, respectively.

Techniques:

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Physicochemical properties of the PEW300 peptide. (A) The sequences of PEW300 and Cecropin A (CeA). The red font and underline sites indicate mutation sites of the PEW300 peptide. (B) Predicted alpha-helical wheel of PEW300 peptide ( https://heliquest.ipmc.cnrs.fr/cgi-bin/ComputParams.py ). The hydrophobic residues are yellow, positively charged hydrophilic residues are blue, and non-charged polar residues are gray. (C) Purification of PEW300 peptide. Line 1 and line 2 are purified PEW300 peptides. (D) The secondary structure of PEW300 predicted by SWISS-MODEL ( https://swissmodel.expasy.org/interactive ). (E) CD spectra of the PEW300 peptide.

Article Snippet: In this study, high purity of PEW300 peptide was acquired by our previously established peptide expression system ( ; ).

Techniques: Mutagenesis, Purification

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Hemolytic activity, cytotoxicity, and drug resistance of PEW300. (A) Hemolytic activity of PEW300 in mouse red blood cells. 0.1% Triton X-100 was used as the positive control (PC); *** p < 0.005 compared with PC group. (B) Cytotoxicity of PEW300 in HEK293 cells was determined by a CCK-8 assay. 0.1% Triton-X 100 and PBS were used as positive control (PC) and negative control (NC), respectively. Error bars represent the standard error from mean cell viabilities from three replicates, *** indicates p < 0.005 compared with PC group. (C) Drug resistance of the PEW300 and gentamicin. GEN, gentamicin.

Article Snippet: In this study, high purity of PEW300 peptide was acquired by our previously established peptide expression system ( ; ).

Techniques: Activity Assay, Positive Control, CCK-8 Assay, Negative Control

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Antibacterial and antibiofilm properties of PEW300. (A) Antibacterial activity of PEW300, kanamycin (Kan), and gentamicin (GEN) against Pseudomonas aeruginosa JCM5962. (B) Inhibition and (C) dispersion activity of PEW300 against P. aeruginosa JCM5962 biofilms. Error bars represent standard error from mean values as determined by three repeated experiments, *** indicates p < 0.005 and ns means no significant difference compared with Kan-treated groups.

Article Snippet: In this study, high purity of PEW300 peptide was acquired by our previously established peptide expression system ( ; ).

Techniques: Activity Assay, Inhibition

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Antibiofilm pathway and time-killing kinetic analysis of PEW300 against Pseudomonas aeruginosa . (A) Antibiofilm pathway of PEW300 against P. aeruginosa . Reductive effect of PEW300 against P. aeruginosa biofilm (B) and planktonic cells (C) .

Article Snippet: In this study, high purity of PEW300 peptide was acquired by our previously established peptide expression system ( ; ).

Techniques:

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: SEM observation of preformed biofilm of Pseudomonas aeruginosa JCM5962 affected by PEW300. The red arrow indicates the damaged JCM5962 cells.

Article Snippet: In this study, high purity of PEW300 peptide was acquired by our previously established peptide expression system ( ; ).

Techniques:

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Eliminative action of PEW300 on Pseudomonas aeruginosa mature biofilm. (A) Fluorescence analysis of PEW300 on P. aeruginosa mature biofilm components. The scale bar is 200 μm. (B) Quantification analysis of mature biofilms components of P. aeruginosa JCM5962 affected by PEW300. * Indicates statistical significance compared to the control groups ( * p < 0.05; *** p < 0.005).

Article Snippet: In this study, high purity of PEW300 peptide was acquired by our previously established peptide expression system ( ; ).

Techniques: Fluorescence

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Disruption effect of PEW300 on Pseudomonas aeruginosa membrane. (A) TEM micrographs of P. aeruginosa JCM5962 treated with or without PEW300 (Control). (B) Flow cytometry analysis of the disruption ability of PEW300 on P. aeruginosa membrane.

Article Snippet: In this study, high purity of PEW300 peptide was acquired by our previously established peptide expression system ( ; ).

Techniques: Flow Cytometry

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Membrane analysis and intracellular disturbance induced by PEW300. (A) Effects of PEW300 on OM permeabilization of Pseudomonas aeruginosa JCM5962. Control represents cells with no PEW300 treatment. (B) LPS-binding affinities of PEW300. Control means the growth of P. aeruginosa JCM5962 with no PEW300 and LPS incubation. * p < 0.05, *** p < 0.005 compared with control. (C) The IM depolarization induced by PEW300. (D) The electrophoretic mobility shift analysis of the interaction between P. aeruginosa JCM5962 genomic DNA (200 ng) and PEW300 peptide. (E) Intracellular ROS intensity produced by P. aeruginosa JCM5962 in the presence of PBS, 1% Triton X-100, and series concentrations of PEW300 peptide. * p < 0.05, *** p < 0.005 compared with PBS-treated group.

Article Snippet: In this study, high purity of PEW300 peptide was acquired by our previously established peptide expression system ( ; ).

Techniques: Binding Assay, Incubation, Electrophoretic Mobility Shift Assay, Produced

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Analysis of Pseudomonas aeruginosa virulence factors affected by PEW300. PEW300 results in decreased virulence of P. aeruginosa JCM5962 in A549 cells characterized by (A) CCK-8 assay and (B) calcein-AM staining. *** p < 0.005 compared with P. aeruginosa JCM5962-treated group. Scale bar is 100 μm. Comparative analysis of (C) pyocyanin content, (D) elastase activity and (E) alginates content in P. aeruginosa JCM5962 treated with or without PEW300 peptide. (F) qPCR analysis of virulence-related gene expression. Data represent the mean ± SD from three independent experiments performed in triplicate. * indicates p < 0.05, *** indicates p < 0.005; ns means no significant difference compared with control.

Article Snippet: In this study, high purity of PEW300 peptide was acquired by our previously established peptide expression system ( ; ).

Techniques: CCK-8 Assay, Staining, Activity Assay, Expressing

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Schematic illustration of the (A) antibiofilm pathway and (B) the multiple actions of PEW300 against Pseudomonas aeruginosa .

Article Snippet: In this study, high purity of PEW300 peptide was acquired by our previously established peptide expression system ( ; ).

Techniques:

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Physicochemical properties of the PEW300 peptide. (A) The sequences of PEW300 and Cecropin A (CeA). The red font and underline sites indicate mutation sites of the PEW300 peptide. (B) Predicted alpha-helical wheel of PEW300 peptide ( https://heliquest.ipmc.cnrs.fr/cgi-bin/ComputParams.py ). The hydrophobic residues are yellow, positively charged hydrophilic residues are blue, and non-charged polar residues are gray. (C) Purification of PEW300 peptide. Line 1 and line 2 are purified PEW300 peptides. (D) The secondary structure of PEW300 predicted by SWISS-MODEL ( https://swissmodel.expasy.org/interactive ). (E) CD spectra of the PEW300 peptide.

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques: Mutagenesis, Purification

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Hemolytic activity, cytotoxicity, and drug resistance of PEW300. (A) Hemolytic activity of PEW300 in mouse red blood cells. 0.1% Triton X-100 was used as the positive control (PC); *** p < 0.005 compared with PC group. (B) Cytotoxicity of PEW300 in HEK293 cells was determined by a CCK-8 assay. 0.1% Triton-X 100 and PBS were used as positive control (PC) and negative control (NC), respectively. Error bars represent the standard error from mean cell viabilities from three replicates, *** indicates p < 0.005 compared with PC group. (C) Drug resistance of the PEW300 and gentamicin. GEN, gentamicin.

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques: Activity Assay, Positive Control, CCK-8 Assay, Negative Control

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Antibacterial and antibiofilm properties of PEW300. (A) Antibacterial activity of PEW300, kanamycin (Kan), and gentamicin (GEN) against Pseudomonas aeruginosa JCM5962. (B) Inhibition and (C) dispersion activity of PEW300 against P. aeruginosa JCM5962 biofilms. Error bars represent standard error from mean values as determined by three repeated experiments, *** indicates p < 0.005 and ns means no significant difference compared with Kan-treated groups.

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques: Activity Assay, Inhibition

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Antibiofilm pathway and time-killing kinetic analysis of PEW300 against Pseudomonas aeruginosa . (A) Antibiofilm pathway of PEW300 against P. aeruginosa . Reductive effect of PEW300 against P. aeruginosa biofilm (B) and planktonic cells (C) .

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques:

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: SEM observation of preformed biofilm of Pseudomonas aeruginosa JCM5962 affected by PEW300. The red arrow indicates the damaged JCM5962 cells.

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques:

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Eliminative action of PEW300 on Pseudomonas aeruginosa mature biofilm. (A) Fluorescence analysis of PEW300 on P. aeruginosa mature biofilm components. The scale bar is 200 μm. (B) Quantification analysis of mature biofilms components of P. aeruginosa JCM5962 affected by PEW300. * Indicates statistical significance compared to the control groups ( * p < 0.05; *** p < 0.005).

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques: Fluorescence

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Disruption effect of PEW300 on Pseudomonas aeruginosa membrane. (A) TEM micrographs of P. aeruginosa JCM5962 treated with or without PEW300 (Control). (B) Flow cytometry analysis of the disruption ability of PEW300 on P. aeruginosa membrane.

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques: Flow Cytometry

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Membrane analysis and intracellular disturbance induced by PEW300. (A) Effects of PEW300 on OM permeabilization of Pseudomonas aeruginosa JCM5962. Control represents cells with no PEW300 treatment. (B) LPS-binding affinities of PEW300. Control means the growth of P. aeruginosa JCM5962 with no PEW300 and LPS incubation. * p < 0.05, *** p < 0.005 compared with control. (C) The IM depolarization induced by PEW300. (D) The electrophoretic mobility shift analysis of the interaction between P. aeruginosa JCM5962 genomic DNA (200 ng) and PEW300 peptide. (E) Intracellular ROS intensity produced by P. aeruginosa JCM5962 in the presence of PBS, 1% Triton X-100, and series concentrations of PEW300 peptide. * p < 0.05, *** p < 0.005 compared with PBS-treated group.

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques: Binding Assay, Incubation, Electrophoretic Mobility Shift Assay, Produced

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Analysis of Pseudomonas aeruginosa virulence factors affected by PEW300. PEW300 results in decreased virulence of P. aeruginosa JCM5962 in A549 cells characterized by (A) CCK-8 assay and (B) calcein-AM staining. *** p < 0.005 compared with P. aeruginosa JCM5962-treated group. Scale bar is 100 μm. Comparative analysis of (C) pyocyanin content, (D) elastase activity and (E) alginates content in P. aeruginosa JCM5962 treated with or without PEW300 peptide. (F) qPCR analysis of virulence-related gene expression. Data represent the mean ± SD from three independent experiments performed in triplicate. * indicates p < 0.05, *** indicates p < 0.005; ns means no significant difference compared with control.

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques: CCK-8 Assay, Staining, Activity Assay, Expressing

Journal: Frontiers in Microbiology

Article Title: Antibiofilm property and multiple action of peptide PEW300 against Pseudomonas aeruginosa

doi: 10.3389/fmicb.2022.963292

Figure Lengend Snippet: Schematic illustration of the (A) antibiofilm pathway and (B) the multiple actions of PEW300 against Pseudomonas aeruginosa .

Article Snippet: For comparative analysis of virulence production (pyocyanin, elastase, and alginate) with or without PEW300 treatment, P. aeruginosa cells were adjusted to an initial OD 600 at 0.1 and incubated with increasing concentrations (0, 10, and 20 μg/ml) of PEW300 at 37°C for 8 h. Elastolytic activity was assessed using a 1% skimmed milk plate as previously reported ( ); Pyocyanin content was determined by chloroform-hydrochloric acid extraction method as previously described ( ); The amount of alginate was quantified using the borate-carbazole method with sodium alginate (Sigma, United States) as a standard ( ).

Techniques: