atcc2001 c glabrata reference strain american type culture collection efg1δδ cph1δδ c albicans yeast  (ATCC)

Journal: MicrobiologyOpen

Article Title: Staphylococcus aureus Extracellular Vesicles Enhance PslE‐Mediated Pathogenesis in Pseudomonas aeruginosa

doi: 10.1002/mbo3.70114

Figure Lengend Snippet: Scheme of double‐crossing over and loxP ‐Gm‐ loxP replacement of pslE gene in Pseudomonas aeruginosa genomic DNA (gDNA) for constructing a pslE gene‐deletion mutant of P. aeruginosa (Δ pslE ) from P. aeruginosa wildtype (WT) by Cre‐ loxP system. Gm, gentamicin.

Article Snippet: Materials and Methods Construction of pslE Gene Deletion in Pseudomonas aeruginosa P. aeruginosa strain American Type Culture Collection (ATCC) 15692, Escherichia coli BW20767 and E. coli DH5α were cultured in Luria–Bertani (LB) medium (Invitrogen, Carlsbad, CA).

Techniques: Mutagenesis

Journal: MicrobiologyOpen

Article Title: Staphylococcus aureus Extracellular Vesicles Enhance PslE‐Mediated Pathogenesis in Pseudomonas aeruginosa

doi: 10.1002/mbo3.70114

Figure Lengend Snippet: Preparation of DNA fragments for constructing pKNOCK AP ::Δ pslE ::Gm plasmid. (A) The upstream region (1200 bp, blue) and downstream region (1160 bp, red) of pslE gene in Pseudomonas aeruginosa wildtype (WT) were amplified using primer Up_F/Up_R and primer Down_F/Down_R, respectively. (B) The loxP ‐Gm‐ loxP fragment (949 bp) was amplified from plasmid pCM351 using primers lxP‐G‐lxP_F and lxP‐G‐lxP_R. (C) The pKNOCK AP plasmid was linearized with Sma I (2066 bp). (D) The DNA fragments were purified and checked by 1% gel electrophoresis. M, 500 bp DNA ladder; U, upstream fragment; Gm, lox P‐Gm‐ lox P; D, downstream fragment; pK, linearized pKNOCK AP plasmid. gDNA, genomic DNA; Gm, gentamicin; PCR, polymerase chain reaction.

Article Snippet: Materials and Methods Construction of pslE Gene Deletion in Pseudomonas aeruginosa P. aeruginosa strain American Type Culture Collection (ATCC) 15692, Escherichia coli BW20767 and E. coli DH5α were cultured in Luria–Bertani (LB) medium (Invitrogen, Carlsbad, CA).

Techniques: Plasmid Preparation, Amplification, Purification, Nucleic Acid Electrophoresis, Polymerase Chain Reaction

Journal: MicrobiologyOpen

Article Title: Staphylococcus aureus Extracellular Vesicles Enhance PslE‐Mediated Pathogenesis in Pseudomonas aeruginosa

doi: 10.1002/mbo3.70114

Figure Lengend Snippet: Generation of Pseudomonas aeruginosa Δ pslE ::Gm. Plasmid pKNOCK AP ::Δ pslE ::Gm was transformed into P. aeruginosa wildtype (WT), and the transformants were selected on Luria–Bertani agar containing 30 μg/mL gentamicin. (A) Scheme of pslE gene in P. aeruginosa WT and replacement of pslE gene with loxP ‐Gm‐ loxP in P. aeruginosa Δ pslE ::Gm. The expected sizes of PCR products using primer pslE _F and pslE _R in P. aeruginosa WT and Δ pslE ::Gm was 2015 and 1102 bp, respectively. (B) Eight Gm‐resistant clones were obtained and the PCR product indicated that pslE gene was replaced with loxP ‐Gm‐ loxP fragment in all clones. M, 500 bp DNA ladder. Gm, gentamicin; PCR, polymerase chain reaction.

Article Snippet: Materials and Methods Construction of pslE Gene Deletion in Pseudomonas aeruginosa P. aeruginosa strain American Type Culture Collection (ATCC) 15692, Escherichia coli BW20767 and E. coli DH5α were cultured in Luria–Bertani (LB) medium (Invitrogen, Carlsbad, CA).

Techniques: Plasmid Preparation, Transformation Assay, Clone Assay, Polymerase Chain Reaction

Journal: MicrobiologyOpen

Article Title: Staphylococcus aureus Extracellular Vesicles Enhance PslE‐Mediated Pathogenesis in Pseudomonas aeruginosa

doi: 10.1002/mbo3.70114

Figure Lengend Snippet: Generation of Pseudomonas aeruginosa Δ pslE . Plasmid pCM157 was transformed into P. aeruginosa Δ pslE ::Gm, and the transformants were selected on Luria–Bertani agar containing 200 μg/mL tetracycline. On the basis of the activity of Cre recombinase from pCM157 plasmid and loxP sequence, Gm‐resistant gene is removed. (A) Scheme of pslE gene in P. aeruginosa wildtype (WT) and remaining pslE gene in P. aeruginosa Δ pslE . The expected sizes of PCR products using primer pslE _F and pslE _R in P. aeruginosa WT and Δ pslE was 2015 and 226 bp, respectively. (B) Ten tetracycline‐resistant clones were randomly selected and the PCR product indicated that pslE and Gm‐resistant genes were successfully deleted in all clones. M1 and M2, 100 and 500 bp DNA ladders, respectively. Gm, gentamicin; PCR, polymerase chain reaction.

Article Snippet: Materials and Methods Construction of pslE Gene Deletion in Pseudomonas aeruginosa P. aeruginosa strain American Type Culture Collection (ATCC) 15692, Escherichia coli BW20767 and E. coli DH5α were cultured in Luria–Bertani (LB) medium (Invitrogen, Carlsbad, CA).

Techniques: Plasmid Preparation, Transformation Assay, Activity Assay, Sequencing, Clone Assay, Polymerase Chain Reaction

Journal: MicrobiologyOpen

Article Title: Staphylococcus aureus Extracellular Vesicles Enhance PslE‐Mediated Pathogenesis in Pseudomonas aeruginosa

doi: 10.1002/mbo3.70114

Figure Lengend Snippet: DNA sequence and predicted amino acids after deletion of pslE gene in PaΔ pslE . (A) DNA sequence demonstrates that 5’‐ and 3’‐ends of pslE gene are joined by linkers and loxP . Boxes indicate start and stop codon of pslE gene. (B) Scheme and predicted amino acid sequence analyzed by Expasy translate tool. Truncated PslE protein consists of 22 and 27 residues from N‐ and C‐terminal of PslE, respectively (blue letters) and 30 residues from linkers and loxP (black letters, loxP is highlighted in grey). (C) Relative mRNA expression of pslE gene in Pseudomonas aeruginosa PaΔ pslE and wildtype (PaWT) determined by RT‐qPCR neutralizing with expression of rpoD gene. The values are from two‐independent experiment ( n = 6). mRNA, messenger RNA; PCR, polymerase chain reaction; RT‐qPCR, reverse transcription quantitative polymerase chain reaction; UD, undetectable.

Article Snippet: Materials and Methods Construction of pslE Gene Deletion in Pseudomonas aeruginosa P. aeruginosa strain American Type Culture Collection (ATCC) 15692, Escherichia coli BW20767 and E. coli DH5α were cultured in Luria–Bertani (LB) medium (Invitrogen, Carlsbad, CA).

Techniques: Sequencing, Expressing, Quantitative RT-PCR, Polymerase Chain Reaction, Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: MicrobiologyOpen

Article Title: Staphylococcus aureus Extracellular Vesicles Enhance PslE‐Mediated Pathogenesis in Pseudomonas aeruginosa

doi: 10.1002/mbo3.70114

Figure Lengend Snippet: Effect of SaEVs on the expression of polysaccharide polymerization‐related genes in psl cluster of pslE gene‐deletion mutant of P. aeruginosa (PaΔ pslE ) and P. aeruginosa wildtype (PaWT). PaΔ pslE and PaWT were treated with 0 and 5 µg/mL SaEVs for 4 h. Relative mRNA expression of pslA, pslE, pslJ, pslK , and pslL was assessed by RT‐qPCR. The values are from two independent experiments ( n = 6). Relative gene expression was calculated by referring to the expression of each target gene in PaWT as 1.0. Statistical analysis was performed using one‐way ANOVA with Tukey's multiple comparison test for relative gene expression. Asterisks indicate p values (* p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001). UD: undetectable.

Article Snippet: Materials and Methods Construction of pslE Gene Deletion in Pseudomonas aeruginosa P. aeruginosa strain American Type Culture Collection (ATCC) 15692, Escherichia coli BW20767 and E. coli DH5α were cultured in Luria–Bertani (LB) medium (Invitrogen, Carlsbad, CA).

Techniques: Expressing, Mutagenesis, Quantitative RT-PCR, Gene Expression, Comparison

Journal: MicrobiologyOpen

Article Title: Staphylococcus aureus Extracellular Vesicles Enhance PslE‐Mediated Pathogenesis in Pseudomonas aeruginosa

doi: 10.1002/mbo3.70114

Figure Lengend Snippet: Effect of SaEVs on LPS production in pslE gene deletion mutant of P. aeruginosa (PaΔ pslE ) and P. aeruginosa wildtype (PaWT). PaΔ pslE and PaWT were treated with or without 5 µg/mL SaEVs for 24 h. LPS were isolated, separated by SDS‐PAGE, and stained using Silver Stain kit. All samples were started with the same number of bacterial cells, prepared by the same process, and applied in equal volumes. The table demonstrates the quantitative intensity of each group, setting the intensity of PaWT without SaEVs as 1.00.

Article Snippet: Materials and Methods Construction of pslE Gene Deletion in Pseudomonas aeruginosa P. aeruginosa strain American Type Culture Collection (ATCC) 15692, Escherichia coli BW20767 and E. coli DH5α were cultured in Luria–Bertani (LB) medium (Invitrogen, Carlsbad, CA).

Techniques: Mutagenesis, Isolation, SDS Page, Staining, Silver Staining

Journal: MicrobiologyOpen

Article Title: Staphylococcus aureus Extracellular Vesicles Enhance PslE‐Mediated Pathogenesis in Pseudomonas aeruginosa

doi: 10.1002/mbo3.70114

Figure Lengend Snippet: Effect of SaEVs on biofilm formation in pslE gene deletion mutant of P. aeruginosa (PaΔ pslE ) and P. aeruginosa wildtype (PaWT). PaΔ pslE and PaWT were cultured with or without 5 µg/mL SaEVs for 16 h. Biofilm biomass was quantified by crystal violet assay. Relative biofilm biomass was calculated by referring PaWT without SaEVs as 100%. The values are from two independent experiments ( n = 6). Statistical analysis was performed using Kruskal–Wallis with Dunn's multiple comparisons. Asterisks indicate p values (* p < 0.05, ** p < 0.01), NS: not significant.

Article Snippet: Materials and Methods Construction of pslE Gene Deletion in Pseudomonas aeruginosa P. aeruginosa strain American Type Culture Collection (ATCC) 15692, Escherichia coli BW20767 and E. coli DH5α were cultured in Luria–Bertani (LB) medium (Invitrogen, Carlsbad, CA).

Techniques: Mutagenesis, Cell Culture, Crystal Violet Assay

Journal: MicrobiologyOpen

Article Title: Staphylococcus aureus Extracellular Vesicles Enhance PslE‐Mediated Pathogenesis in Pseudomonas aeruginosa

doi: 10.1002/mbo3.70114

Figure Lengend Snippet: Effect of SaEVs on invasion of pslE gene deletion mutant of P. aeruginosa (PaΔ pslE ) and P. aeruginosa wildtype (PaWT) into human keratinocytes. PaWT and PaΔ pslE were treated with or without 5 µg/mL SaEVs for 24 h before infection. HaCaT cells were infected with PaΔ pslE and PaWT at MOI = 100 for 90 min. The extracellular bacterial cells were eliminated by gentamicin treatment for 1 h. Then, the intracellular bacterial number was evaluated by plate count assay. Relative intracellular bacterial number was calculated by referring PaWT without SaEVs as 100%. The values are from two independent experiments ( n = 6). Statistical analysis was performed using Kruskal–Wallis with Dunn's multiple comparisons. NS: not significant.

Article Snippet: Materials and Methods Construction of pslE Gene Deletion in Pseudomonas aeruginosa P. aeruginosa strain American Type Culture Collection (ATCC) 15692, Escherichia coli BW20767 and E. coli DH5α were cultured in Luria–Bertani (LB) medium (Invitrogen, Carlsbad, CA).

Techniques: Mutagenesis, Infection

Journal: MicrobiologyOpen

Article Title: Staphylococcus aureus Extracellular Vesicles Enhance PslE‐Mediated Pathogenesis in Pseudomonas aeruginosa

doi: 10.1002/mbo3.70114

Figure Lengend Snippet: Effect of SaEVs on uptake of pslE gene deletion mutant of P. aeruginosa (PaΔ pslE ) and P. aeruginosa wildtype (PaWT) by mouse macrophages. PaΔ pslE and PaWT were treated with or without 5 µg/mL SaEVs for 24 h before infection. RAW 264.7 cells were infected with PaΔ pslE and PaWT at MOI = 10 for 90 min. The extracellular bacterial cells were eliminated by gentamicin treatment for 1 h. Then, the intracellular bacterial number was evaluated by plate count assay. Relative intracellular bacterial number was calculated by referring PaWT without SaEVs as 100%. The values are from two independent experiments ( n = 6). Statistical analysis was performed using Kruskal–Wallis with Dunn's multiple comparisons. Asterisks indicate p values (* p < 0.05, ** p < 0.01), NS: not significant.

Article Snippet: Materials and Methods Construction of pslE Gene Deletion in Pseudomonas aeruginosa P. aeruginosa strain American Type Culture Collection (ATCC) 15692, Escherichia coli BW20767 and E. coli DH5α were cultured in Luria–Bertani (LB) medium (Invitrogen, Carlsbad, CA).

Techniques: Mutagenesis, Infection

Journal: Clinical, Cosmetic and Investigational Dentistry

Article Title: Comparative Efficacy of Sonic, Ultrasonic, and Diode Laser-Activated Irrigation with Combination of NaOCl/EDTA/CHX Solutions Against Fusobacterium nucleatum in Dentinal Tubules: A Confocal Microscopy Study

doi: 10.2147/CCIDE.S567977

Figure Lengend Snippet: Representative images of scanning electron microscope results displaying dental specimens before ( A and B ) and after ( C and D ) F. nucleatum contamination taken at 500x (top) and 5000x (bottom) magnifications.

Article Snippet: Briefly, strain F. nucleatum American Type Culture Collection (ATCC) 25586 was reactivated in trypticase soy broth (TSB) and incubated in an anaerobic chamber for 48 hours.

Techniques: Microscopy

Journal: Clinical, Cosmetic and Investigational Dentistry

Article Title: Comparative Efficacy of Sonic, Ultrasonic, and Diode Laser-Activated Irrigation with Combination of NaOCl/EDTA/CHX Solutions Against Fusobacterium nucleatum in Dentinal Tubules: A Confocal Microscopy Study

doi: 10.2147/CCIDE.S567977

Figure Lengend Snippet: Representative images of confocal laser scanning microscope results after F. nucleatum contamination under SYTO 9 ( A ) and propidium iodide ( B ) staining at 1.25x magnification.

Article Snippet: Briefly, strain F. nucleatum American Type Culture Collection (ATCC) 25586 was reactivated in trypticase soy broth (TSB) and incubated in an anaerobic chamber for 48 hours.

Techniques: Laser-Scanning Microscopy, Staining

Journal: Clinical, Cosmetic and Investigational Dentistry

Article Title: Comparative Efficacy of Sonic, Ultrasonic, and Diode Laser-Activated Irrigation with Combination of NaOCl/EDTA/CHX Solutions Against Fusobacterium nucleatum in Dentinal Tubules: A Confocal Microscopy Study

doi: 10.2147/CCIDE.S567977

Figure Lengend Snippet: Comparative evaluation of irrigation techniques on the elimination of F. nucleatum from dentinal tubules.

Article Snippet: Briefly, strain F. nucleatum American Type Culture Collection (ATCC) 25586 was reactivated in trypticase soy broth (TSB) and incubated in an anaerobic chamber for 48 hours.

Techniques: