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  • 98
    ATCC biofilms
    Effects of COL and EGCg against Sm biofilm formation. Reference strain ATCC13637 and two clinical isolates (Sm1, and Sm2) were used. <t>Biofilms</t> were stained with crystal violet and their biomasses were determined by optical density (OD) measurement at 620 nm. Compared to untreated control cells, samples exposed to EGCg and COL exhibited a significant reduction in the number of Sm sessile cells of ATCC13637 and Sm1. Results are expressed as average OD ± standard deviation (SD). Experiments were performed in triplicate. * P
    Biofilms, supplied by ATCC, used in various techniques. Bioz Stars score: 98/100, based on 1351 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Bio-Rad biofilm a
    Effects of COL and EGCg against Sm biofilm formation. Reference strain ATCC13637 and two clinical isolates (Sm1, and Sm2) were used. <t>Biofilms</t> were stained with crystal violet and their biomasses were determined by optical density (OD) measurement at 620 nm. Compared to untreated control cells, samples exposed to EGCg and COL exhibited a significant reduction in the number of Sm sessile cells of ATCC13637 and Sm1. Results are expressed as average OD ± standard deviation (SD). Experiments were performed in triplicate. * P
    Biofilm A, supplied by Bio-Rad, used in various techniques. Bioz Stars score: 88/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Rayto Life biofilm
    Effects of COL and EGCg against Sm biofilm formation. Reference strain ATCC13637 and two clinical isolates (Sm1, and Sm2) were used. <t>Biofilms</t> were stained with crystal violet and their biomasses were determined by optical density (OD) measurement at 620 nm. Compared to untreated control cells, samples exposed to EGCg and COL exhibited a significant reduction in the number of Sm sessile cells of ATCC13637 and Sm1. Results are expressed as average OD ± standard deviation (SD). Experiments were performed in triplicate. * P
    Biofilm, supplied by Rayto Life, used in various techniques. Bioz Stars score: 93/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    MJ Research biofilm a
    Effects of COL and EGCg against Sm biofilm formation. Reference strain ATCC13637 and two clinical isolates (Sm1, and Sm2) were used. <t>Biofilms</t> were stained with crystal violet and their biomasses were determined by optical density (OD) measurement at 620 nm. Compared to untreated control cells, samples exposed to EGCg and COL exhibited a significant reduction in the number of Sm sessile cells of ATCC13637 and Sm1. Results are expressed as average OD ± standard deviation (SD). Experiments were performed in triplicate. * P
    Biofilm A, supplied by MJ Research, used in various techniques. Bioz Stars score: 85/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Johnson & Johnson biofilms
    An overview of the high-throughput protocol for metal susceptibility testing using the MBEC assay . (A) Frozen stocks of bacteria were streaked out on the appropriate agar medium to obtain a first- and a subsequent second-subculture. (B) Colonies were collected from second-subcultures and suspended in broth medium to a 1.0 McFarland Standard. (C) This suspension was diluted 30-fold in broth, and the 1 in 30 dilution was used to inoculate the MBEC assay. (D) The inoculated device was placed on a rocking table in an incubator. (E) Serial dilutions of metal cations and oxyanions were set up along length of a microtiter plate along (the challenge plate). (F) The <t>biofilms</t> were rinsed to remove loosely adherent planktonic bacteria. (G) The first peg from each row was removed. These pegs were used to verify growth of the biofilms on the pegs. The peg lid was then inserted into the challenge plate. (H) During exposure, metals diffuse into the biofilm while planktonic cells are shed from the surface of the biofilm. Sloughed cells serve as the inoculum for planktonic MIC and MBC determinations. (I) The exposed biofilms were rinsed twice and the peg lid was inserted into fresh recovery medium containing the appropriate neutralizing agent (the recovery plate). The biofilms were disrupted into the recovery medium by sonciation on a water table sonicator. (J) Aliquots of planktonic cultures were transferred from the challenge plate to a microtiter plate containing the appropriate neutralizing agents (the neutralizing plate). (K) An aliquot from the recovery and neutralizing plates were spotted onto rich agar media. (L) MIC values are determined by reading the optical density at 650 nm (OD 650 ) of the challenge plate after the desired period of incubation using a microtiter plate reader. Spot plates were qualitatively scored for growth to obtain MBC and MBEC values. MBEC values were redundantly determined by determining the A 650 of the recovery plates after incubation.
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    94
    Millipore biofilms
    Rapid development of nfxB mutants in a 24-h-old PAO1 flow cell biofilm treated with low-dose CIP. The <t>biofilms</t> of PAO1- mCherry -P CD - gfp + were grown in flow cell chambers with a continuous flow of minimal medium for 24 h at 37°C and then grown for an additional 24 h with 0.2 μg/ml CIP (24 h +CIP for 24 h) or without CIP (24 h −CIP for 24 h). Red shows wild-type cells (elongated or filamentous) due to the constitutive expression of mCherry, and green shows nfxB mutants due to the expression of GFP via the P CD - gfp + reporter. z-stacks were generated by using a Zeiss LSM 710 microscope and processed with Imaris 8.2 software (Bitplane). Top-row images show orthogonal 3D views, and middle and bottom (for treated biofilms only) rows show perspective 3D views of biofilms with an overlay of red and green fluorescence. The images shown are representative of results for two independent flow cell channels.
    Biofilms, supplied by Millipore, used in various techniques. Bioz Stars score: 94/100, based on 1410 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Denville Scientific blue biofilm
    Rapid development of nfxB mutants in a 24-h-old PAO1 flow cell biofilm treated with low-dose CIP. The <t>biofilms</t> of PAO1- mCherry -P CD - gfp + were grown in flow cell chambers with a continuous flow of minimal medium for 24 h at 37°C and then grown for an additional 24 h with 0.2 μg/ml CIP (24 h +CIP for 24 h) or without CIP (24 h −CIP for 24 h). Red shows wild-type cells (elongated or filamentous) due to the constitutive expression of mCherry, and green shows nfxB mutants due to the expression of GFP via the P CD - gfp + reporter. z-stacks were generated by using a Zeiss LSM 710 microscope and processed with Imaris 8.2 software (Bitplane). Top-row images show orthogonal 3D views, and middle and bottom (for treated biofilms only) rows show perspective 3D views of biofilms with an overlay of red and green fluorescence. The images shown are representative of results for two independent flow cell channels.
    Blue Biofilm, supplied by Denville Scientific, used in various techniques. Bioz Stars score: 85/100, based on 13 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Innovotech biofilm inoculator
    Rapid development of nfxB mutants in a 24-h-old PAO1 flow cell biofilm treated with low-dose CIP. The <t>biofilms</t> of PAO1- mCherry -P CD - gfp + were grown in flow cell chambers with a continuous flow of minimal medium for 24 h at 37°C and then grown for an additional 24 h with 0.2 μg/ml CIP (24 h +CIP for 24 h) or without CIP (24 h −CIP for 24 h). Red shows wild-type cells (elongated or filamentous) due to the constitutive expression of mCherry, and green shows nfxB mutants due to the expression of GFP via the P CD - gfp + reporter. z-stacks were generated by using a Zeiss LSM 710 microscope and processed with Imaris 8.2 software (Bitplane). Top-row images show orthogonal 3D views, and middle and bottom (for treated biofilms only) rows show perspective 3D views of biofilms with an overlay of red and green fluorescence. The images shown are representative of results for two independent flow cell channels.
    Biofilm Inoculator, supplied by Innovotech, used in various techniques. Bioz Stars score: 93/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Fisher Scientific biofilm substratum
    The S-layer and type IV pili are required for <t>biofilm</t> formation by Synechocystis . The “attached” data series shows crystal violet binding measured at the OD 600 . The “suspended” data series shows planktonic growth measured at OD 730 . Wza, SD517; Slyr, SD523; PilC, SD519; WT, SD100. Both data series are shown on the same y axis. Error bar corresponds to one standard deviation from the sample mean.
    Biofilm Substratum, supplied by Fisher Scientific, used in various techniques. Bioz Stars score: 93/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    GraphPad Prism Inc biofilm chemostats
    The S-layer and type IV pili are required for <t>biofilm</t> formation by Synechocystis . The “attached” data series shows crystal violet binding measured at the OD 600 . The “suspended” data series shows planktonic growth measured at OD 730 . Wza, SD517; Slyr, SD523; PilC, SD519; WT, SD100. Both data series are shown on the same y axis. Error bar corresponds to one standard deviation from the sample mean.
    Biofilm Chemostats, supplied by GraphPad Prism Inc, used in various techniques. Bioz Stars score: 93/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Olympus biofilm distribution
    Percentage <t>biofilm</t> formed and inhibited in Staphylococcus aureus. (A) Expression levels of SarA in the bacterial strains taken in this study at the log and late phases of growth. (B) Percentage biofilm formation of SarA mutant ALC637 (ΔsarA::795 Tn917LTV1) on treatment and no treatment with 2-[(Methylamino)methyl]phenol by crystal violet method. (C,D) Percentage biofilm inhibition in clinical S. aureus isolates P1966 and AB459 by 2-[(Methylamino)methyl]phenol. All the assays were done in triplicates and the values were expressed as mean ± SD. ∗ Indicates significantly different ( p ≤ 0.05) when compared to untreated (control) with 2-[(Methylamino)methyl]phenol. NS denotes not significant ( P > 0.05).
    Biofilm Distribution, supplied by Olympus, used in various techniques. Bioz Stars score: 91/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Waters Corporation biofilm formation
    <t>Biofilm</t> formation by various kinase mutants on tomato roots
    Biofilm Formation, supplied by Waters Corporation, used in various techniques. Bioz Stars score: 93/100, based on 77 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Innovotech biofilms biofilms
    Persister cells in H. volcanii <t>biofilms.</t> Comparison of percentage survival of planktonic and biofilm cultures of H. volcanii , following incubation for 6 h in biocidal concentrations of H 2 O 2 , NaClO, and chlorhexidine. Plotted values are the mean of triplicate measurements and error bars represent ± SD. Asterisks denote significance values as determined by paired t -tests: ∗ p
    Biofilms Biofilms, supplied by Innovotech, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Innovotech calgary biofilm plate
    Liposomal clarithromycin activity on P. aureginosa PA-13572 <t>biofilm</t> (MBEC assay). Free (F-CAM) or liposomal formulations were introduced to mature biofilm at concentrations of 32 mg/liter (a), 64 mg/liter (b), and 128 mg/liter (c). Untreated biofilm acted as a control. The data represent three independent experiments in triplicate and are shown as means ± SEM. P values were considered significant compared with the control: ***, P
    Calgary Biofilm Plate, supplied by Innovotech, used in various techniques. Bioz Stars score: 85/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    Thermo Fisher filmtracer green biofilm
    Liposomal clarithromycin activity on P. aureginosa PA-13572 <t>biofilm</t> (MBEC assay). Free (F-CAM) or liposomal formulations were introduced to mature biofilm at concentrations of 32 mg/liter (a), 64 mg/liter (b), and 128 mg/liter (c). Untreated biofilm acted as a control. The data represent three independent experiments in triplicate and are shown as means ± SEM. P values were considered significant compared with the control: ***, P
    Filmtracer Green Biofilm, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 26 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    ATCC biofilm formation
    PelA H Bc (BCE_5582) is a predicted glycoside hydrolase that is capable of disrupting pre-formed Pel-dependent biofilms. (A) <t>Biofilm</t> formation of the indicated strains of B . cereus ATCC 10987 assessed by the crystal violet assay. VC, empty vector control. Error bars represent the standard error of the mean of six independent trials. Statistical significance, compared to WT + VC, was determined using one-way analysis of variance with Dunn’s multiple comparison. ns, no significant difference; ****, p
    Biofilm Formation, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 2160 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Bitplane biofilm analysis xtension
    PelA H Bc (BCE_5582) is a predicted glycoside hydrolase that is capable of disrupting pre-formed Pel-dependent biofilms. (A) <t>Biofilm</t> formation of the indicated strains of B . cereus ATCC 10987 assessed by the crystal violet assay. VC, empty vector control. Error bars represent the standard error of the mean of six independent trials. Statistical significance, compared to WT + VC, was determined using one-way analysis of variance with Dunn’s multiple comparison. ns, no significant difference; ****, p
    Biofilm Analysis Xtension, supplied by Bitplane, used in various techniques. Bioz Stars score: 93/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Agar Scientific biofilm microbial biofilms
    Correlations between the number of microorganisms forming <t>biofilm</t> ( S. mutans ) colony forming units and the optical density (the biofilm mass) at different time points (( A )–18 h; ( B )–20 h; ( C )–22 h; ( D )–24 h) after Lactobacillus salivarius (HM6 Paradens) administration.
    Biofilm Microbial Biofilms, supplied by Agar Scientific, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Corning Life Sciences biofilm construction biofilms
    Viable counts of each bacterial species present in the initial inoculum (per well) and in the 3‐day old <t>biofilms</t> ( n = 7). Fn = Fusobacterium nucleatum ; Pg = Porphyromonas gingivalis ; Aa = Aggregatibacter actinomycetemcomitans . Error bars: standard error
    Biofilm Construction Biofilms, supplied by Corning Life Sciences, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    89
    Becton Dickinson biofilm formation biofilms
    Confocal Laser Scanning Microscopy of <t>biofilm</t> formed by S. pseudintermedius strain DSM 25713. Biofilm was allowed to form for 48 h at 37 °C, in absence of serum, under both ( a ) static, and ( b ) dynamic (flow cell chamber) conditions. Static <t>biofilms</t> were further treated for 24 h with increasing gentamicin concentrations (1x-128xMIC). Representative images of biofilm exposed at ( c ) 1x and ( d ) 128xMIC gentamicin are shown. Orthogonal images z are projections of x and y planes, collected within the biofilm as indicated by the green and red lines in the top view. Image capture was set for simultaneous visualization of red (Propidium iodide-stained dead cells), green (Syto-9-stained viable cells), and blue (Concanavalin A-stained EPS) fluorescence. Magnification, x100
    Biofilm Formation Biofilms, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 89/100, based on 20 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    Millipore nthi biofilms
    Confocal Laser Scanning Microscopy of <t>biofilm</t> formed by S. pseudintermedius strain DSM 25713. Biofilm was allowed to form for 48 h at 37 °C, in absence of serum, under both ( a ) static, and ( b ) dynamic (flow cell chamber) conditions. Static <t>biofilms</t> were further treated for 24 h with increasing gentamicin concentrations (1x-128xMIC). Representative images of biofilm exposed at ( c ) 1x and ( d ) 128xMIC gentamicin are shown. Orthogonal images z are projections of x and y planes, collected within the biofilm as indicated by the green and red lines in the top view. Image capture was set for simultaneous visualization of red (Propidium iodide-stained dead cells), green (Syto-9-stained viable cells), and blue (Concanavalin A-stained EPS) fluorescence. Magnification, x100
    Nthi Biofilms, supplied by Millipore, used in various techniques. Bioz Stars score: 90/100, based on 17 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    Amresco biofilm minimal media
    <t>Biofilm</t> elimination by MNP-enhanced oxidative cleavage
    Biofilm Minimal Media, supplied by Amresco, used in various techniques. Bioz Stars score: 86/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    89
    Kodak Corp xar biofilm
    <t>Biofilm</t> elimination by MNP-enhanced oxidative cleavage
    Xar Biofilm, supplied by Kodak Corp, used in various techniques. Bioz Stars score: 89/100, based on 59 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    SAS institute biofilm control sas
    <t>Biofilm</t> elimination by MNP-enhanced oxidative cleavage
    Biofilm Control Sas, supplied by SAS institute, used in various techniques. Bioz Stars score: 90/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Illumina Inc biofilm cells
    Quantification of <t>biofilm</t> formation by the wild-type strain (ATCC 17978), a stable knockout mutant strain lacking the gene A1S_0114 (ATCC Δ0114), the same mutant strain containing the pET-RA plasmid (ATCC Δ0114 + PETRA), and a mutant strain containing the pET-RA plasmid harboring the A1S_0114 gene (ATCC Δ0114 + PETRA + 0114).
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    Image Search Results


    Effects of COL and EGCg against Sm biofilm formation. Reference strain ATCC13637 and two clinical isolates (Sm1, and Sm2) were used. Biofilms were stained with crystal violet and their biomasses were determined by optical density (OD) measurement at 620 nm. Compared to untreated control cells, samples exposed to EGCg and COL exhibited a significant reduction in the number of Sm sessile cells of ATCC13637 and Sm1. Results are expressed as average OD ± standard deviation (SD). Experiments were performed in triplicate. * P

    Journal: PLoS ONE

    Article Title: Effects of Green Tea Compound Epigallocatechin-3-Gallate against Stenotrophomonas maltophilia Infection and Biofilm

    doi: 10.1371/journal.pone.0092876

    Figure Lengend Snippet: Effects of COL and EGCg against Sm biofilm formation. Reference strain ATCC13637 and two clinical isolates (Sm1, and Sm2) were used. Biofilms were stained with crystal violet and their biomasses were determined by optical density (OD) measurement at 620 nm. Compared to untreated control cells, samples exposed to EGCg and COL exhibited a significant reduction in the number of Sm sessile cells of ATCC13637 and Sm1. Results are expressed as average OD ± standard deviation (SD). Experiments were performed in triplicate. * P

    Article Snippet: After 24 h exposure to EGCg, both 24-h-old and 7-day-old biofilms from ATCC 13637, Sm1, Sm2 showed a mean viability decrease in comparison to untreated biofilms.

    Techniques: Staining, Standard Deviation

    Optical sections of 48-h-old Sm biofilms (reference strain ATCC13637 and clinical isolates: Sm1 and Sm2) treated with EGCg and COL at 0.25×MIC, 0.5×MIC, 1×MIC. Biofilms were treated with formalin as killing control. Live bacteria are stained in green (Syto9), dead bacteria in red (propidium iodide [PI]) or yellow (overlapping regions). Experiments were performed in duplicates (image data: 1024 ×1024 pixel with a pixel-size of 0.284 μm; z-step-size: 2 μm). Length of size bar: 50 μm.

    Journal: PLoS ONE

    Article Title: Effects of Green Tea Compound Epigallocatechin-3-Gallate against Stenotrophomonas maltophilia Infection and Biofilm

    doi: 10.1371/journal.pone.0092876

    Figure Lengend Snippet: Optical sections of 48-h-old Sm biofilms (reference strain ATCC13637 and clinical isolates: Sm1 and Sm2) treated with EGCg and COL at 0.25×MIC, 0.5×MIC, 1×MIC. Biofilms were treated with formalin as killing control. Live bacteria are stained in green (Syto9), dead bacteria in red (propidium iodide [PI]) or yellow (overlapping regions). Experiments were performed in duplicates (image data: 1024 ×1024 pixel with a pixel-size of 0.284 μm; z-step-size: 2 μm). Length of size bar: 50 μm.

    Article Snippet: After 24 h exposure to EGCg, both 24-h-old and 7-day-old biofilms from ATCC 13637, Sm1, Sm2 showed a mean viability decrease in comparison to untreated biofilms.

    Techniques: Staining

    Effects of COL and EGCg on 24-h- and 7-day-old established biofilms of Sm. Reference strain ATCC13637 and clinical isolates (Sm1 and Sm2) had their biofilm metabolic activity defined by XTT viability assay. OD measurement was determined at 492 nm, and results are expressed as average OD ± standard deviation (SD). Experiments were performed in triplicate. * P

    Journal: PLoS ONE

    Article Title: Effects of Green Tea Compound Epigallocatechin-3-Gallate against Stenotrophomonas maltophilia Infection and Biofilm

    doi: 10.1371/journal.pone.0092876

    Figure Lengend Snippet: Effects of COL and EGCg on 24-h- and 7-day-old established biofilms of Sm. Reference strain ATCC13637 and clinical isolates (Sm1 and Sm2) had their biofilm metabolic activity defined by XTT viability assay. OD measurement was determined at 492 nm, and results are expressed as average OD ± standard deviation (SD). Experiments were performed in triplicate. * P

    Article Snippet: After 24 h exposure to EGCg, both 24-h-old and 7-day-old biofilms from ATCC 13637, Sm1, Sm2 showed a mean viability decrease in comparison to untreated biofilms.

    Techniques: Activity Assay, Viability Assay, Standard Deviation

    An overview of the high-throughput protocol for metal susceptibility testing using the MBEC assay . (A) Frozen stocks of bacteria were streaked out on the appropriate agar medium to obtain a first- and a subsequent second-subculture. (B) Colonies were collected from second-subcultures and suspended in broth medium to a 1.0 McFarland Standard. (C) This suspension was diluted 30-fold in broth, and the 1 in 30 dilution was used to inoculate the MBEC assay. (D) The inoculated device was placed on a rocking table in an incubator. (E) Serial dilutions of metal cations and oxyanions were set up along length of a microtiter plate along (the challenge plate). (F) The biofilms were rinsed to remove loosely adherent planktonic bacteria. (G) The first peg from each row was removed. These pegs were used to verify growth of the biofilms on the pegs. The peg lid was then inserted into the challenge plate. (H) During exposure, metals diffuse into the biofilm while planktonic cells are shed from the surface of the biofilm. Sloughed cells serve as the inoculum for planktonic MIC and MBC determinations. (I) The exposed biofilms were rinsed twice and the peg lid was inserted into fresh recovery medium containing the appropriate neutralizing agent (the recovery plate). The biofilms were disrupted into the recovery medium by sonciation on a water table sonicator. (J) Aliquots of planktonic cultures were transferred from the challenge plate to a microtiter plate containing the appropriate neutralizing agents (the neutralizing plate). (K) An aliquot from the recovery and neutralizing plates were spotted onto rich agar media. (L) MIC values are determined by reading the optical density at 650 nm (OD 650 ) of the challenge plate after the desired period of incubation using a microtiter plate reader. Spot plates were qualitatively scored for growth to obtain MBC and MBEC values. MBEC values were redundantly determined by determining the A 650 of the recovery plates after incubation.

    Journal: BMC Microbiology

    Article Title: High-throughput metal susceptibility testing of microbial biofilms

    doi: 10.1186/1471-2180-5-53

    Figure Lengend Snippet: An overview of the high-throughput protocol for metal susceptibility testing using the MBEC assay . (A) Frozen stocks of bacteria were streaked out on the appropriate agar medium to obtain a first- and a subsequent second-subculture. (B) Colonies were collected from second-subcultures and suspended in broth medium to a 1.0 McFarland Standard. (C) This suspension was diluted 30-fold in broth, and the 1 in 30 dilution was used to inoculate the MBEC assay. (D) The inoculated device was placed on a rocking table in an incubator. (E) Serial dilutions of metal cations and oxyanions were set up along length of a microtiter plate along (the challenge plate). (F) The biofilms were rinsed to remove loosely adherent planktonic bacteria. (G) The first peg from each row was removed. These pegs were used to verify growth of the biofilms on the pegs. The peg lid was then inserted into the challenge plate. (H) During exposure, metals diffuse into the biofilm while planktonic cells are shed from the surface of the biofilm. Sloughed cells serve as the inoculum for planktonic MIC and MBC determinations. (I) The exposed biofilms were rinsed twice and the peg lid was inserted into fresh recovery medium containing the appropriate neutralizing agent (the recovery plate). The biofilms were disrupted into the recovery medium by sonciation on a water table sonicator. (J) Aliquots of planktonic cultures were transferred from the challenge plate to a microtiter plate containing the appropriate neutralizing agents (the neutralizing plate). (K) An aliquot from the recovery and neutralizing plates were spotted onto rich agar media. (L) MIC values are determined by reading the optical density at 650 nm (OD 650 ) of the challenge plate after the desired period of incubation using a microtiter plate reader. Spot plates were qualitatively scored for growth to obtain MBC and MBEC values. MBEC values were redundantly determined by determining the A 650 of the recovery plates after incubation.

    Article Snippet: In general, biofilms formed using a trough have a 5- to 10-fold greater cell density than those formed using the microtiter plate format (J.J. Harrison, H. Ceri and C. Stremick, unpublished data).

    Techniques: High Throughput Screening Assay, Incubation

    Growth of bacterial biofilms in the MBEC assay . (A) Mean cell density of Pseudomonas aeruginosa ATCC 27853 biofilms on the pegs in different rows of the MBEC assay. Each value is expressed as the mean and standard deviation of 4 to 6 trials. There is no significant difference between cell density of biofilms in the different rows ( p = 0.842 using one-way ANOVA). (B) SEM photomicrograph of a P. aeruginosa biofilm on the peg surface. (C) Mean cell density of Escherichia coli TG1 on the pegs in different rows of the MBEC assay. Each value is expressed as the mean and standard deviation of 4 to 6 trials. There is no significant difference between cell density of biofilms in the different rows ( p = 0.274 using one-way ANOVA). (D) SEM photomicrograph of an E. coli biofilm on the peg surface. The bar represents 5 μm.

    Journal: BMC Microbiology

    Article Title: High-throughput metal susceptibility testing of microbial biofilms

    doi: 10.1186/1471-2180-5-53

    Figure Lengend Snippet: Growth of bacterial biofilms in the MBEC assay . (A) Mean cell density of Pseudomonas aeruginosa ATCC 27853 biofilms on the pegs in different rows of the MBEC assay. Each value is expressed as the mean and standard deviation of 4 to 6 trials. There is no significant difference between cell density of biofilms in the different rows ( p = 0.842 using one-way ANOVA). (B) SEM photomicrograph of a P. aeruginosa biofilm on the peg surface. (C) Mean cell density of Escherichia coli TG1 on the pegs in different rows of the MBEC assay. Each value is expressed as the mean and standard deviation of 4 to 6 trials. There is no significant difference between cell density of biofilms in the different rows ( p = 0.274 using one-way ANOVA). (D) SEM photomicrograph of an E. coli biofilm on the peg surface. The bar represents 5 μm.

    Article Snippet: In general, biofilms formed using a trough have a 5- to 10-fold greater cell density than those formed using the microtiter plate format (J.J. Harrison, H. Ceri and C. Stremick, unpublished data).

    Techniques: Standard Deviation

    Rapid development of nfxB mutants in a 24-h-old PAO1 flow cell biofilm treated with low-dose CIP. The biofilms of PAO1- mCherry -P CD - gfp + were grown in flow cell chambers with a continuous flow of minimal medium for 24 h at 37°C and then grown for an additional 24 h with 0.2 μg/ml CIP (24 h +CIP for 24 h) or without CIP (24 h −CIP for 24 h). Red shows wild-type cells (elongated or filamentous) due to the constitutive expression of mCherry, and green shows nfxB mutants due to the expression of GFP via the P CD - gfp + reporter. z-stacks were generated by using a Zeiss LSM 710 microscope and processed with Imaris 8.2 software (Bitplane). Top-row images show orthogonal 3D views, and middle and bottom (for treated biofilms only) rows show perspective 3D views of biofilms with an overlay of red and green fluorescence. The images shown are representative of results for two independent flow cell channels.

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: Real-Time Monitoring of nfxB Mutant Occurrence and Dynamics in Pseudomonas aeruginosa Biofilm Exposed to Subinhibitory Concentrations of Ciprofloxacin

    doi: 10.1128/AAC.02292-16

    Figure Lengend Snippet: Rapid development of nfxB mutants in a 24-h-old PAO1 flow cell biofilm treated with low-dose CIP. The biofilms of PAO1- mCherry -P CD - gfp + were grown in flow cell chambers with a continuous flow of minimal medium for 24 h at 37°C and then grown for an additional 24 h with 0.2 μg/ml CIP (24 h +CIP for 24 h) or without CIP (24 h −CIP for 24 h). Red shows wild-type cells (elongated or filamentous) due to the constitutive expression of mCherry, and green shows nfxB mutants due to the expression of GFP via the P CD - gfp + reporter. z-stacks were generated by using a Zeiss LSM 710 microscope and processed with Imaris 8.2 software (Bitplane). Top-row images show orthogonal 3D views, and middle and bottom (for treated biofilms only) rows show perspective 3D views of biofilms with an overlay of red and green fluorescence. The images shown are representative of results for two independent flow cell channels.

    Article Snippet: To isolate nfxB mutants for further characterization, PAO1- mCherry -PCD - gfp + biofilms were harvested by pumping 1 to 1.5 ml of a glass bead (212 to 300 μm; Sigma)-saline suspension through the flow cell channels and collected into sterile Eppendorf tubes.

    Techniques: Flow Cytometry, Expressing, Generated, Microscopy, Software, Fluorescence

    Development of nfxB mutants in a 72-h-old PAO1 flow cell biofilm during treatment with low-dose CIP. The biofilms of PAO1- mCherry -P CD - gfp + were grown in three independent channels of flow cell chambers with a continuous flow of minimal medium for 72 h and then treated with 0.2 μg/ml CIP for a total of 96 h. Imaging by CLSM was done every 24 h. At least 3 images were taken per channel at every time point. Red represents wild-type cells due to the constitutive expression of mCherry, and green shows nfxB mutants due to the expression of GFP+ via the P CD - gfp + reporter. z-stacks were generated by using a Zeiss LSM 710 microscope and processed with Imaris 8.2 software (Bitplane). The images show orthogonal 3D biofilm views (left) or a perspective view (right) with an overlay of red and green channel fluorescence.

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: Real-Time Monitoring of nfxB Mutant Occurrence and Dynamics in Pseudomonas aeruginosa Biofilm Exposed to Subinhibitory Concentrations of Ciprofloxacin

    doi: 10.1128/AAC.02292-16

    Figure Lengend Snippet: Development of nfxB mutants in a 72-h-old PAO1 flow cell biofilm during treatment with low-dose CIP. The biofilms of PAO1- mCherry -P CD - gfp + were grown in three independent channels of flow cell chambers with a continuous flow of minimal medium for 72 h and then treated with 0.2 μg/ml CIP for a total of 96 h. Imaging by CLSM was done every 24 h. At least 3 images were taken per channel at every time point. Red represents wild-type cells due to the constitutive expression of mCherry, and green shows nfxB mutants due to the expression of GFP+ via the P CD - gfp + reporter. z-stacks were generated by using a Zeiss LSM 710 microscope and processed with Imaris 8.2 software (Bitplane). The images show orthogonal 3D biofilm views (left) or a perspective view (right) with an overlay of red and green channel fluorescence.

    Article Snippet: To isolate nfxB mutants for further characterization, PAO1- mCherry -PCD - gfp + biofilms were harvested by pumping 1 to 1.5 ml of a glass bead (212 to 300 μm; Sigma)-saline suspension through the flow cell channels and collected into sterile Eppendorf tubes.

    Techniques: Flow Cytometry, Imaging, Confocal Laser Scanning Microscopy, Expressing, Generated, Microscopy, Software, Fluorescence

    The S-layer and type IV pili are required for biofilm formation by Synechocystis . The “attached” data series shows crystal violet binding measured at the OD 600 . The “suspended” data series shows planktonic growth measured at OD 730 . Wza, SD517; Slyr, SD523; PilC, SD519; WT, SD100. Both data series are shown on the same y axis. Error bar corresponds to one standard deviation from the sample mean.

    Journal: Applied and Environmental Microbiology

    Article Title: Axenic Biofilm Formation and Aggregation by Synechocystis sp. Strain PCC 6803 Are Induced by Changes in Nutrient Concentration and Require Cell Surface Structures

    doi: 10.1128/AEM.02192-18

    Figure Lengend Snippet: The S-layer and type IV pili are required for biofilm formation by Synechocystis . The “attached” data series shows crystal violet binding measured at the OD 600 . The “suspended” data series shows planktonic growth measured at OD 730 . Wza, SD517; Slyr, SD523; PilC, SD519; WT, SD100. Both data series are shown on the same y axis. Error bar corresponds to one standard deviation from the sample mean.

    Article Snippet: Three milliliters of culture was added to each well of a 12-well plate (Corning Costar, catalog no. 07–200–82; Fisher Scientific) that contained a 22-mm-thick glass coverslip as a biofilm substratum (12–540–B; Fisher Scientific).

    Techniques: Binding Assay, Standard Deviation

    Biofouling of a nonaxenic rooftop photobioreactor during growth of WT Synechocystis . (A) During lag phase, no biofilm growth was evident. (B to G) Images taken every 24 h during rapid growth (approximate doubling every 24 h over a period of 5 days). Biofouling such as in the representative images shown was correlated with using hard tap water to prepare BG11 medium; no biofouling was evident when softened tap water was used. Scale bar, ~2 cm. The glass PBR tubes are ~20 cm in diameter.

    Journal: Applied and Environmental Microbiology

    Article Title: Axenic Biofilm Formation and Aggregation by Synechocystis sp. Strain PCC 6803 Are Induced by Changes in Nutrient Concentration and Require Cell Surface Structures

    doi: 10.1128/AEM.02192-18

    Figure Lengend Snippet: Biofouling of a nonaxenic rooftop photobioreactor during growth of WT Synechocystis . (A) During lag phase, no biofilm growth was evident. (B to G) Images taken every 24 h during rapid growth (approximate doubling every 24 h over a period of 5 days). Biofouling such as in the representative images shown was correlated with using hard tap water to prepare BG11 medium; no biofouling was evident when softened tap water was used. Scale bar, ~2 cm. The glass PBR tubes are ~20 cm in diameter.

    Article Snippet: Three milliliters of culture was added to each well of a 12-well plate (Corning Costar, catalog no. 07–200–82; Fisher Scientific) that contained a 22-mm-thick glass coverslip as a biofilm substratum (12–540–B; Fisher Scientific).

    Techniques:

    Percentage biofilm formed and inhibited in Staphylococcus aureus. (A) Expression levels of SarA in the bacterial strains taken in this study at the log and late phases of growth. (B) Percentage biofilm formation of SarA mutant ALC637 (ΔsarA::795 Tn917LTV1) on treatment and no treatment with 2-[(Methylamino)methyl]phenol by crystal violet method. (C,D) Percentage biofilm inhibition in clinical S. aureus isolates P1966 and AB459 by 2-[(Methylamino)methyl]phenol. All the assays were done in triplicates and the values were expressed as mean ± SD. ∗ Indicates significantly different ( p ≤ 0.05) when compared to untreated (control) with 2-[(Methylamino)methyl]phenol. NS denotes not significant ( P > 0.05).

    Journal: Frontiers in Microbiology

    Article Title: Staphylococcus aureus Quorum Regulator SarA Targeted Compound, 2-[(Methylamino)methyl]phenol Inhibits Biofilm and Down-Regulates Virulence Genes

    doi: 10.3389/fmicb.2017.01290

    Figure Lengend Snippet: Percentage biofilm formed and inhibited in Staphylococcus aureus. (A) Expression levels of SarA in the bacterial strains taken in this study at the log and late phases of growth. (B) Percentage biofilm formation of SarA mutant ALC637 (ΔsarA::795 Tn917LTV1) on treatment and no treatment with 2-[(Methylamino)methyl]phenol by crystal violet method. (C,D) Percentage biofilm inhibition in clinical S. aureus isolates P1966 and AB459 by 2-[(Methylamino)methyl]phenol. All the assays were done in triplicates and the values were expressed as mean ± SD. ∗ Indicates significantly different ( p ≤ 0.05) when compared to untreated (control) with 2-[(Methylamino)methyl]phenol. NS denotes not significant ( P > 0.05).

    Article Snippet: Two and three-dimensional images were captured using a confocal laser scanning microscope with a 40× objective lens to show biofilm distribution (Olympus FLUOVIEW, FV1000).

    Techniques: Expressing, Mutagenesis, Inhibition

    Confocal laser scanning microscopy imaging. Representative images showing the biofilm inhibition effects of 2-[(Methylamino)methyl]phenol at varying concentrations on Staphylococcus aureus P1966 and AB459 during log phase of growth (6 h). The results were in concordance with microtitre plate quantitative assays where biofilm inhibition was more at 1.25 μM. Scale bar in images represents 50 μm.

    Journal: Frontiers in Microbiology

    Article Title: Staphylococcus aureus Quorum Regulator SarA Targeted Compound, 2-[(Methylamino)methyl]phenol Inhibits Biofilm and Down-Regulates Virulence Genes

    doi: 10.3389/fmicb.2017.01290

    Figure Lengend Snippet: Confocal laser scanning microscopy imaging. Representative images showing the biofilm inhibition effects of 2-[(Methylamino)methyl]phenol at varying concentrations on Staphylococcus aureus P1966 and AB459 during log phase of growth (6 h). The results were in concordance with microtitre plate quantitative assays where biofilm inhibition was more at 1.25 μM. Scale bar in images represents 50 μm.

    Article Snippet: Two and three-dimensional images were captured using a confocal laser scanning microscope with a 40× objective lens to show biofilm distribution (Olympus FLUOVIEW, FV1000).

    Techniques: Confocal Laser Scanning Microscopy, Imaging, Inhibition

    Biofilm formation by various kinase mutants on tomato roots

    Journal: Molecular microbiology

    Article Title: A Bacillus subtilis Sensor Kinase Involved in Triggering Biofilm Formation on the Roots of Tomato Plants

    doi: 10.1111/j.1365-2958.2012.08109.x

    Figure Lengend Snippet: Biofilm formation by various kinase mutants on tomato roots

    Article Snippet: To identify the tomato root-released chemical(s) that induce biofilm formation, we first applied the root exudates to a Sep-Pak Plus C-18 column (Waters Corporation, MA, USA) and separated compounds by eluting them with aqueous methanol (from a gradient of 10% to 100%).

    Techniques:

    l -Malic acid stimulates biofilm formation

    Journal: Molecular microbiology

    Article Title: A Bacillus subtilis Sensor Kinase Involved in Triggering Biofilm Formation on the Roots of Tomato Plants

    doi: 10.1111/j.1365-2958.2012.08109.x

    Figure Lengend Snippet: l -Malic acid stimulates biofilm formation

    Article Snippet: To identify the tomato root-released chemical(s) that induce biofilm formation, we first applied the root exudates to a Sep-Pak Plus C-18 column (Waters Corporation, MA, USA) and separated compounds by eluting them with aqueous methanol (from a gradient of 10% to 100%).

    Techniques:

    B. subtilis forms biofilms on the tomato root surfaces

    Journal: Molecular microbiology

    Article Title: A Bacillus subtilis Sensor Kinase Involved in Triggering Biofilm Formation on the Roots of Tomato Plants

    doi: 10.1111/j.1365-2958.2012.08109.x

    Figure Lengend Snippet: B. subtilis forms biofilms on the tomato root surfaces

    Article Snippet: To identify the tomato root-released chemical(s) that induce biofilm formation, we first applied the root exudates to a Sep-Pak Plus C-18 column (Waters Corporation, MA, USA) and separated compounds by eluting them with aqueous methanol (from a gradient of 10% to 100%).

    Techniques:

    Persister cells in H. volcanii biofilms. Comparison of percentage survival of planktonic and biofilm cultures of H. volcanii , following incubation for 6 h in biocidal concentrations of H 2 O 2 , NaClO, and chlorhexidine. Plotted values are the mean of triplicate measurements and error bars represent ± SD. Asterisks denote significance values as determined by paired t -tests: ∗ p

    Journal: Frontiers in Microbiology

    Article Title: Archaeal Persisters: Persister Cell Formation as a Stress Response in Haloferax volcanii

    doi: 10.3389/fmicb.2017.01589

    Figure Lengend Snippet: Persister cells in H. volcanii biofilms. Comparison of percentage survival of planktonic and biofilm cultures of H. volcanii , following incubation for 6 h in biocidal concentrations of H 2 O 2 , NaClO, and chlorhexidine. Plotted values are the mean of triplicate measurements and error bars represent ± SD. Asterisks denote significance values as determined by paired t -tests: ∗ p

    Article Snippet: Persister Cell Formation in Biofilms Biofilms of H. volcanii were grown using the MBEC device (Innovotech).

    Techniques: Incubation

    Liposomal clarithromycin activity on P. aureginosa PA-13572 biofilm (MBEC assay). Free (F-CAM) or liposomal formulations were introduced to mature biofilm at concentrations of 32 mg/liter (a), 64 mg/liter (b), and 128 mg/liter (c). Untreated biofilm acted as a control. The data represent three independent experiments in triplicate and are shown as means ± SEM. P values were considered significant compared with the control: ***, P

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: Efficacy and Safety of Liposomal Clarithromycin and Its Effect on Pseudomonas aeruginosa Virulence Factors

    doi: 10.1128/AAC.00235-13

    Figure Lengend Snippet: Liposomal clarithromycin activity on P. aureginosa PA-13572 biofilm (MBEC assay). Free (F-CAM) or liposomal formulations were introduced to mature biofilm at concentrations of 32 mg/liter (a), 64 mg/liter (b), and 128 mg/liter (c). Untreated biofilm acted as a control. The data represent three independent experiments in triplicate and are shown as means ± SEM. P values were considered significant compared with the control: ***, P

    Article Snippet: In order to assess the minimum biofilm eradication concentration (MBEC), P. aeruginosa strain PA-13572 was allowed to form a biofilm in a Calgary biofilm plate (Innovotech, Edmonton, AB, Canada) as previously reported ( ).

    Techniques: Activity Assay, Chick Chorioallantoic Membrane Assay

    PelA H Bc (BCE_5582) is a predicted glycoside hydrolase that is capable of disrupting pre-formed Pel-dependent biofilms. (A) Biofilm formation of the indicated strains of B . cereus ATCC 10987 assessed by the crystal violet assay. VC, empty vector control. Error bars represent the standard error of the mean of six independent trials. Statistical significance, compared to WT + VC, was determined using one-way analysis of variance with Dunn’s multiple comparison. ns, no significant difference; ****, p

    Journal: PLoS Pathogens

    Article Title: Discovery and characterization of a Gram-positive Pel polysaccharide biosynthetic gene clusterA systematic pipeline for classifying bacterial operons reveals the evolutionary landscape of biofilm machineries

    doi: 10.1371/journal.ppat.1008281

    Figure Lengend Snippet: PelA H Bc (BCE_5582) is a predicted glycoside hydrolase that is capable of disrupting pre-formed Pel-dependent biofilms. (A) Biofilm formation of the indicated strains of B . cereus ATCC 10987 assessed by the crystal violet assay. VC, empty vector control. Error bars represent the standard error of the mean of six independent trials. Statistical significance, compared to WT + VC, was determined using one-way analysis of variance with Dunn’s multiple comparison. ns, no significant difference; ****, p

    Article Snippet: To assess whether cdgF and cdgE play a role in biofilm formation by ATCC 10987, we generated cdgF and cdgE deletion mutant strains and analyzed biofilm formation using the crystal violet assay ( ).

    Techniques: Crystal Violet Assay, Plasmid Preparation

    The pel operon of B . cereus ATCC 10987 is involved in the generation of a Pel-like extracellular material. (A) Brightfield and epifluorescence microscopy of P . aeruginosa (PAO1 Δ wspF Δ psl P BAD pel ) wild-type and Δ pelF mutant, and B . cereus ATCC 10987 wild-type and Δ pelF mutant biofilms grown for 24 h on poly-L-lysine coated glass coverslips and stained with DAPI, to detect DNA, and Wisteria floribunda lectin (WFL) conjugated to Texas red to detect Pel-like extracellular material. Scale bar = 250 μm. (B) Dot blot of extracellular material from B . cereus ATCC 10987 or the Δ pelF mutant grown for 24 h at 30°C with shaking, separated into crude cell-associated and secreted fractions. Extracellular material was detected using WFL conjugated to horseradish peroxidase (HRP; left ) or α-Pel primary antibody with HRP-conjugated secondary antibody ( right ).

    Journal: PLoS Pathogens

    Article Title: Discovery and characterization of a Gram-positive Pel polysaccharide biosynthetic gene clusterA systematic pipeline for classifying bacterial operons reveals the evolutionary landscape of biofilm machineries

    doi: 10.1371/journal.ppat.1008281

    Figure Lengend Snippet: The pel operon of B . cereus ATCC 10987 is involved in the generation of a Pel-like extracellular material. (A) Brightfield and epifluorescence microscopy of P . aeruginosa (PAO1 Δ wspF Δ psl P BAD pel ) wild-type and Δ pelF mutant, and B . cereus ATCC 10987 wild-type and Δ pelF mutant biofilms grown for 24 h on poly-L-lysine coated glass coverslips and stained with DAPI, to detect DNA, and Wisteria floribunda lectin (WFL) conjugated to Texas red to detect Pel-like extracellular material. Scale bar = 250 μm. (B) Dot blot of extracellular material from B . cereus ATCC 10987 or the Δ pelF mutant grown for 24 h at 30°C with shaking, separated into crude cell-associated and secreted fractions. Extracellular material was detected using WFL conjugated to horseradish peroxidase (HRP; left ) or α-Pel primary antibody with HRP-conjugated secondary antibody ( right ).

    Article Snippet: To assess whether cdgF and cdgE play a role in biofilm formation by ATCC 10987, we generated cdgF and cdgE deletion mutant strains and analyzed biofilm formation using the crystal violet assay ( ).

    Techniques: Epifluorescence Microscopy, Mutagenesis, Staining, Dot Blot

    B . cereus ATCC 10987 forms biofilms that are dependent on the pelDEA DA FG operon. (A) B . cereus ATCC 10987 pel operon architecture. Open reading frames are represented as arrows, with the directionality of transcription indicated by the arrow direction. Open reading frames and the overall operon architecture is drawn to scale. Arrow colours correspond to predicted protein functions as defined in Fig 1 . Gene IDs are indicated above or below each gene. (B) Air-liquid interface biofilm formation by ATCC 10987 in a 96-well microtitre plate (left) and a borosilicate glass tube (right). (C) Biofilm formation of the indicated strains of B . cereus ATCC 10987 assessed by the crystal violet assay. Error bars represent the standard error of the mean of six independent trials. Statistical significance, compared to WT, was determined using one-way analysis of variance with Dunn’s multiple comparison. ****, p

    Journal: PLoS Pathogens

    Article Title: Discovery and characterization of a Gram-positive Pel polysaccharide biosynthetic gene clusterA systematic pipeline for classifying bacterial operons reveals the evolutionary landscape of biofilm machineries

    doi: 10.1371/journal.ppat.1008281

    Figure Lengend Snippet: B . cereus ATCC 10987 forms biofilms that are dependent on the pelDEA DA FG operon. (A) B . cereus ATCC 10987 pel operon architecture. Open reading frames are represented as arrows, with the directionality of transcription indicated by the arrow direction. Open reading frames and the overall operon architecture is drawn to scale. Arrow colours correspond to predicted protein functions as defined in Fig 1 . Gene IDs are indicated above or below each gene. (B) Air-liquid interface biofilm formation by ATCC 10987 in a 96-well microtitre plate (left) and a borosilicate glass tube (right). (C) Biofilm formation of the indicated strains of B . cereus ATCC 10987 assessed by the crystal violet assay. Error bars represent the standard error of the mean of six independent trials. Statistical significance, compared to WT, was determined using one-way analysis of variance with Dunn’s multiple comparison. ****, p

    Article Snippet: To assess whether cdgF and cdgE play a role in biofilm formation by ATCC 10987, we generated cdgF and cdgE deletion mutant strains and analyzed biofilm formation using the crystal violet assay ( ).

    Techniques: Crystal Violet Assay

    Binding of c-di-GMP to PelD is required for B . cereus ATCC 10987 biofilm formation. (A) Structure of P . aeruginosa PelD 156-455 (PDB code: 4DN0), and model of B . cereus PelD 170-403 . Inset: close up of the I-site and residues involved in the binding of c-di-GMP. The equivalent B . cereus residues to Arg367, Asp370, and Arg402 of P . aeruginosa were identified as Arg363, Asp366, and Arg395, and are represented as green sticks for clarity. The sequence containing these residues is indicated below the inset. (B) Isothermal titration calorimetry experiments for the wild-type PelD Bc 150-407 and the indicated mutants. Each experiment displays the heats of injection (top panel) and the normalized integration data as a function of the molar syringe and cell concentrations (bottom panel). The calculated dissociation constant (K D ) for the wild-type protein is indicated on the figure. (C) Biofilm formation of the indicated strains of B . cereus ATCC 10987 assessed by the crystal violet assay. Error bars represent the standard error of the mean of six independent trials. Statistical significance was determined using one-way analysis of variance with Dunn’s multiple comparison. ns, no significant difference; ****, p

    Journal: PLoS Pathogens

    Article Title: Discovery and characterization of a Gram-positive Pel polysaccharide biosynthetic gene clusterA systematic pipeline for classifying bacterial operons reveals the evolutionary landscape of biofilm machineries

    doi: 10.1371/journal.ppat.1008281

    Figure Lengend Snippet: Binding of c-di-GMP to PelD is required for B . cereus ATCC 10987 biofilm formation. (A) Structure of P . aeruginosa PelD 156-455 (PDB code: 4DN0), and model of B . cereus PelD 170-403 . Inset: close up of the I-site and residues involved in the binding of c-di-GMP. The equivalent B . cereus residues to Arg367, Asp370, and Arg402 of P . aeruginosa were identified as Arg363, Asp366, and Arg395, and are represented as green sticks for clarity. The sequence containing these residues is indicated below the inset. (B) Isothermal titration calorimetry experiments for the wild-type PelD Bc 150-407 and the indicated mutants. Each experiment displays the heats of injection (top panel) and the normalized integration data as a function of the molar syringe and cell concentrations (bottom panel). The calculated dissociation constant (K D ) for the wild-type protein is indicated on the figure. (C) Biofilm formation of the indicated strains of B . cereus ATCC 10987 assessed by the crystal violet assay. Error bars represent the standard error of the mean of six independent trials. Statistical significance was determined using one-way analysis of variance with Dunn’s multiple comparison. ns, no significant difference; ****, p

    Article Snippet: To assess whether cdgF and cdgE play a role in biofilm formation by ATCC 10987, we generated cdgF and cdgE deletion mutant strains and analyzed biofilm formation using the crystal violet assay ( ).

    Techniques: Binding Assay, Sequencing, Isothermal Titration Calorimetry, Injection, Crystal Violet Assay

    Model for B . cereus ATCC 10987 Pel biosynthesis. PelF is a glycosyltransferase that polymerizes UDP-GalNAc to generate the Pel polysaccharide (light pink triangles are acetyl groups, red hexagons are GalN monosaccharide units, dark red teardrops are UDP). Once synthesized, Pel is exported across the cytoplasmic membrane via PelD, PelE, PelA DA , and/or PelG. PelA DA , corresponding to the deacetylase domain of P . aeruginosa PelA, partially deacetylates the polymer to generate mature Pel. PelA H , corresponding to the hydrolase domain of P . aeruginosa PelA, performs a role in regulating overall biofilm formation levels of ATCC 10987. Pel biosynthesis requires binding of the nucleotide second messenger c-di-GMP to the I-site of the cytoplasmic GGDEF domain of PelD. CdgF is a diguanylate cyclase that synthesizes c-di-GMP from two GTP molecules and thus positively regulates Pel biosynthesis. CdgE is a phosphodiesterase that converts c-di-GMP to 5’-pGpG and thus negatively regulates Pel biosynthesis. GalE epimerizes UDP-GlcNAc to UDP-GalNAc, which is presumably utilized by PelF for Pel polymerization. C, cytoplasm; CM, cytoplasmic membrane; EC, extracellular space; PG, peptidoglycan; UDP, uridine di-phosphate; GlcNAc, N- acetylglucosamine; GalNAc, N -acetylgalactosamine; GTP, guanosine tri-phosphate; c-di-GMP, cyclic-3’,5’-dimeric guanosine monophosphate; 5’-pGpG, phosphoguanylyl-3',5'-guanosine.

    Journal: PLoS Pathogens

    Article Title: Discovery and characterization of a Gram-positive Pel polysaccharide biosynthetic gene clusterA systematic pipeline for classifying bacterial operons reveals the evolutionary landscape of biofilm machineries

    doi: 10.1371/journal.ppat.1008281

    Figure Lengend Snippet: Model for B . cereus ATCC 10987 Pel biosynthesis. PelF is a glycosyltransferase that polymerizes UDP-GalNAc to generate the Pel polysaccharide (light pink triangles are acetyl groups, red hexagons are GalN monosaccharide units, dark red teardrops are UDP). Once synthesized, Pel is exported across the cytoplasmic membrane via PelD, PelE, PelA DA , and/or PelG. PelA DA , corresponding to the deacetylase domain of P . aeruginosa PelA, partially deacetylates the polymer to generate mature Pel. PelA H , corresponding to the hydrolase domain of P . aeruginosa PelA, performs a role in regulating overall biofilm formation levels of ATCC 10987. Pel biosynthesis requires binding of the nucleotide second messenger c-di-GMP to the I-site of the cytoplasmic GGDEF domain of PelD. CdgF is a diguanylate cyclase that synthesizes c-di-GMP from two GTP molecules and thus positively regulates Pel biosynthesis. CdgE is a phosphodiesterase that converts c-di-GMP to 5’-pGpG and thus negatively regulates Pel biosynthesis. GalE epimerizes UDP-GlcNAc to UDP-GalNAc, which is presumably utilized by PelF for Pel polymerization. C, cytoplasm; CM, cytoplasmic membrane; EC, extracellular space; PG, peptidoglycan; UDP, uridine di-phosphate; GlcNAc, N- acetylglucosamine; GalNAc, N -acetylgalactosamine; GTP, guanosine tri-phosphate; c-di-GMP, cyclic-3’,5’-dimeric guanosine monophosphate; 5’-pGpG, phosphoguanylyl-3',5'-guanosine.

    Article Snippet: To assess whether cdgF and cdgE play a role in biofilm formation by ATCC 10987, we generated cdgF and cdgE deletion mutant strains and analyzed biofilm formation using the crystal violet assay ( ).

    Techniques: Synthesized, Histone Deacetylase Assay, Binding Assay

    Biofilm formation by B . cereus ATCC 10987 is regulated by the diguanylate cyclase CdgF and the phosphodiesterase CdgE. (A) Biofilm formation by cdgF deletion and complementation strains assessed by the crystal violet assay. (B) Biofilm formation by cdgE deletion and complementation strains assessed by the crystal violet assay. VC, empty vector control; xyl, xylose. Error bars represent the standard error of the mean of six independent trials. Statistical significance was determined using one-way analysis of variance with Dunn’s multiple comparison. ns, no significant difference; *, p

    Journal: PLoS Pathogens

    Article Title: Discovery and characterization of a Gram-positive Pel polysaccharide biosynthetic gene clusterA systematic pipeline for classifying bacterial operons reveals the evolutionary landscape of biofilm machineries

    doi: 10.1371/journal.ppat.1008281

    Figure Lengend Snippet: Biofilm formation by B . cereus ATCC 10987 is regulated by the diguanylate cyclase CdgF and the phosphodiesterase CdgE. (A) Biofilm formation by cdgF deletion and complementation strains assessed by the crystal violet assay. (B) Biofilm formation by cdgE deletion and complementation strains assessed by the crystal violet assay. VC, empty vector control; xyl, xylose. Error bars represent the standard error of the mean of six independent trials. Statistical significance was determined using one-way analysis of variance with Dunn’s multiple comparison. ns, no significant difference; *, p

    Article Snippet: To assess whether cdgF and cdgE play a role in biofilm formation by ATCC 10987, we generated cdgF and cdgE deletion mutant strains and analyzed biofilm formation using the crystal violet assay ( ).

    Techniques: Crystal Violet Assay, Plasmid Preparation

    Correlations between the number of microorganisms forming biofilm ( S. mutans ) colony forming units and the optical density (the biofilm mass) at different time points (( A )–18 h; ( B )–20 h; ( C )–22 h; ( D )–24 h) after Lactobacillus salivarius (HM6 Paradens) administration.

    Journal: Nutrients

    Article Title: Effect of a Lactobacillus Salivarius Probiotic on a Double-Species Streptococcus Mutans and Candida Albicans Caries Biofilm

    doi: 10.3390/nu9111242

    Figure Lengend Snippet: Correlations between the number of microorganisms forming biofilm ( S. mutans ) colony forming units and the optical density (the biofilm mass) at different time points (( A )–18 h; ( B )–20 h; ( C )–22 h; ( D )–24 h) after Lactobacillus salivarius (HM6 Paradens) administration.

    Article Snippet: Scanning Electron Microscopic Analysis of Biofilm Microbial biofilms were grown on 13 mm diameter round basic slides (Agar Scientific, Stansted, UK) in the wells of a 24-well plate, according to the protocol described above.

    Techniques:

    S. mutans , C. albicans , and oral streptococci/yeast biofilm formation: changes in microorganism count (logCFU/mL), before and after administration of Lactobacillus salivarius (HM6 Paradens) after 24 h. Wilcoxon’s test was used for dependent (repeated) measurements (paired Wilcoxon’s test; p

    Journal: Nutrients

    Article Title: Effect of a Lactobacillus Salivarius Probiotic on a Double-Species Streptococcus Mutans and Candida Albicans Caries Biofilm

    doi: 10.3390/nu9111242

    Figure Lengend Snippet: S. mutans , C. albicans , and oral streptococci/yeast biofilm formation: changes in microorganism count (logCFU/mL), before and after administration of Lactobacillus salivarius (HM6 Paradens) after 24 h. Wilcoxon’s test was used for dependent (repeated) measurements (paired Wilcoxon’s test; p

    Article Snippet: Scanning Electron Microscopic Analysis of Biofilm Microbial biofilms were grown on 13 mm diameter round basic slides (Agar Scientific, Stansted, UK) in the wells of a 24-well plate, according to the protocol described above.

    Techniques:

    The protocol of biofilm generation and measurement. BHI: Brain Heart Infusion broth, CFU: colony forming units, liq: liquid, FBS: Fetal Bovine Serum, PBS: phosphate-buffered saline.

    Journal: Nutrients

    Article Title: Effect of a Lactobacillus Salivarius Probiotic on a Double-Species Streptococcus Mutans and Candida Albicans Caries Biofilm

    doi: 10.3390/nu9111242

    Figure Lengend Snippet: The protocol of biofilm generation and measurement. BHI: Brain Heart Infusion broth, CFU: colony forming units, liq: liquid, FBS: Fetal Bovine Serum, PBS: phosphate-buffered saline.

    Article Snippet: Scanning Electron Microscopic Analysis of Biofilm Microbial biofilms were grown on 13 mm diameter round basic slides (Agar Scientific, Stansted, UK) in the wells of a 24-well plate, according to the protocol described above.

    Techniques:

    SEM images of the mono-species biofilm generated by C. albicans , S. mutans and the double-species oral streptococci/yeasts biofilm—untreated and treated with the Lactobacillus salivarius (HM6 Paradens)—after 24 h of biofilm formation. ( A ) The 24 h C. albicans biofilm formed on a flat agar surface (Agar Scientific, Stansted, UK); ( B ) The 24 h C. albicans biofilm formed on a flat agar surface treated with the Lactobacillus salivarius (HM6 Paradens); ( C ) The 24 h S. mutans biofilm formed on a flat agar surface (Agar Scientific, UK); ( D ) The 24 h S. mutans biofilm formed on a flat agar surface treated with the Lactobacillus salivarius (HM6 Paradens); ( E ) Co-culture oral streptococci/yeasts biofilm—untreated with the Lactobacillus salivarius (HM6 Paradens)—after 24 h of biofilm formation; ( F ) Co-culture oral streptococci/yeasts biofilm—treated with the Lactobacillus salivarius (HM6 Paradens)—after 24 h of biofilm formation. The Culture was maintained at 36 °C, pH 7.0 and pCO 2 5%, in bovine serum as a medium additive promoting growth of the culture in the presence of a sucrose substrate (5%). The occurrence of a co-culture biofilm at this stage may depend on the C. albicans morphotypes showing a twofold nature: buds and C. albicans hyphae may colonize mucous membranes and constitute physiological microflora (commensal) or may lead to infection under favorable conditions (opportunistic pathogens). There was no clear, compact structure for the C. albicans biofilm and single loosely located budding cells. Other pathological forms of yeasts are invisible. There was an apparent change in the C. albicans morphotype in the S. mutans common culture and visible pleomorphic forms were true hyphae, blastoconidia, which in the mixed culture also produce mycelial forms, whose role is related to damage to immune cells (macrophages) leading to microorganism invasion. There was abundant extracellular matrix between cells and covering bacterial and yeast cells. Bacterial cells were visible in chains adhering to yeast cells and wrapped around them. There was no clear, compact S. mutans / C. albicans biofilm structure or single loosely located budding cells. Other morphological forms of C. albicans were invisible. There was no visible extracellular matrix ( D , F ), which is formed by S. mutans alone and S. mutans with C. albicans treated with the Lactobacillus salivarius ( C , E ). Original magnification: 400×, 4000× and 6000×.

    Journal: Nutrients

    Article Title: Effect of a Lactobacillus Salivarius Probiotic on a Double-Species Streptococcus Mutans and Candida Albicans Caries Biofilm

    doi: 10.3390/nu9111242

    Figure Lengend Snippet: SEM images of the mono-species biofilm generated by C. albicans , S. mutans and the double-species oral streptococci/yeasts biofilm—untreated and treated with the Lactobacillus salivarius (HM6 Paradens)—after 24 h of biofilm formation. ( A ) The 24 h C. albicans biofilm formed on a flat agar surface (Agar Scientific, Stansted, UK); ( B ) The 24 h C. albicans biofilm formed on a flat agar surface treated with the Lactobacillus salivarius (HM6 Paradens); ( C ) The 24 h S. mutans biofilm formed on a flat agar surface (Agar Scientific, UK); ( D ) The 24 h S. mutans biofilm formed on a flat agar surface treated with the Lactobacillus salivarius (HM6 Paradens); ( E ) Co-culture oral streptococci/yeasts biofilm—untreated with the Lactobacillus salivarius (HM6 Paradens)—after 24 h of biofilm formation; ( F ) Co-culture oral streptococci/yeasts biofilm—treated with the Lactobacillus salivarius (HM6 Paradens)—after 24 h of biofilm formation. The Culture was maintained at 36 °C, pH 7.0 and pCO 2 5%, in bovine serum as a medium additive promoting growth of the culture in the presence of a sucrose substrate (5%). The occurrence of a co-culture biofilm at this stage may depend on the C. albicans morphotypes showing a twofold nature: buds and C. albicans hyphae may colonize mucous membranes and constitute physiological microflora (commensal) or may lead to infection under favorable conditions (opportunistic pathogens). There was no clear, compact structure for the C. albicans biofilm and single loosely located budding cells. Other pathological forms of yeasts are invisible. There was an apparent change in the C. albicans morphotype in the S. mutans common culture and visible pleomorphic forms were true hyphae, blastoconidia, which in the mixed culture also produce mycelial forms, whose role is related to damage to immune cells (macrophages) leading to microorganism invasion. There was abundant extracellular matrix between cells and covering bacterial and yeast cells. Bacterial cells were visible in chains adhering to yeast cells and wrapped around them. There was no clear, compact S. mutans / C. albicans biofilm structure or single loosely located budding cells. Other morphological forms of C. albicans were invisible. There was no visible extracellular matrix ( D , F ), which is formed by S. mutans alone and S. mutans with C. albicans treated with the Lactobacillus salivarius ( C , E ). Original magnification: 400×, 4000× and 6000×.

    Article Snippet: Scanning Electron Microscopic Analysis of Biofilm Microbial biofilms were grown on 13 mm diameter round basic slides (Agar Scientific, Stansted, UK) in the wells of a 24-well plate, according to the protocol described above.

    Techniques: Generated, Co-Culture Assay, Infection

    Intergroup differences between biofilm mass (OD), before and after Lactobacillus salivarius (HM6 Paradens, LS) administration. Kruskal–Wallis test was used for dependent (repeated) measurements; p

    Journal: Nutrients

    Article Title: Effect of a Lactobacillus Salivarius Probiotic on a Double-Species Streptococcus Mutans and Candida Albicans Caries Biofilm

    doi: 10.3390/nu9111242

    Figure Lengend Snippet: Intergroup differences between biofilm mass (OD), before and after Lactobacillus salivarius (HM6 Paradens, LS) administration. Kruskal–Wallis test was used for dependent (repeated) measurements; p

    Article Snippet: Scanning Electron Microscopic Analysis of Biofilm Microbial biofilms were grown on 13 mm diameter round basic slides (Agar Scientific, Stansted, UK) in the wells of a 24-well plate, according to the protocol described above.

    Techniques:

    S. mutans , C. albicans , and oral streptococci/yeast biofilm formation: changes in optical density (OD) of biofilm mass, before and after administration of Lactobacillus salivarius (HM6 Paradens) after 24 h. Wilcoxon’s test was used for dependent (repeated) measurements (paired Wilcoxon’s test; p

    Journal: Nutrients

    Article Title: Effect of a Lactobacillus Salivarius Probiotic on a Double-Species Streptococcus Mutans and Candida Albicans Caries Biofilm

    doi: 10.3390/nu9111242

    Figure Lengend Snippet: S. mutans , C. albicans , and oral streptococci/yeast biofilm formation: changes in optical density (OD) of biofilm mass, before and after administration of Lactobacillus salivarius (HM6 Paradens) after 24 h. Wilcoxon’s test was used for dependent (repeated) measurements (paired Wilcoxon’s test; p

    Article Snippet: Scanning Electron Microscopic Analysis of Biofilm Microbial biofilms were grown on 13 mm diameter round basic slides (Agar Scientific, Stansted, UK) in the wells of a 24-well plate, according to the protocol described above.

    Techniques:

    Correlations between the number of microorganisms forming a biofilm ( S. mutans ) (log(CFU/mL) and the optical density (biofilm mass) after 18 ( A ), 20 ( B ), 22 ( C ), and 24 ( D ) h of incubation, before Lactobacillus salivarius probiotic administration.

    Journal: Nutrients

    Article Title: Effect of a Lactobacillus Salivarius Probiotic on a Double-Species Streptococcus Mutans and Candida Albicans Caries Biofilm

    doi: 10.3390/nu9111242

    Figure Lengend Snippet: Correlations between the number of microorganisms forming a biofilm ( S. mutans ) (log(CFU/mL) and the optical density (biofilm mass) after 18 ( A ), 20 ( B ), 22 ( C ), and 24 ( D ) h of incubation, before Lactobacillus salivarius probiotic administration.

    Article Snippet: Scanning Electron Microscopic Analysis of Biofilm Microbial biofilms were grown on 13 mm diameter round basic slides (Agar Scientific, Stansted, UK) in the wells of a 24-well plate, according to the protocol described above.

    Techniques: Incubation

    SEM images of the mono-species biofilm, generated by S. mutans , C. albicans and the double-species oral streptococci/yeasts biofilm—untreated and treated with the Lactobacillus salivarius (HM6 Paradens)—after 24 h of biofilm formation. ( A ) The 14 h S. mutans biofilm formed on a flat agar surface (Agar Scientific, Stansted, UK). S. mutans adheres to the polystyrene surface, mainly using a sucrose-dependent mechanism; ( B ) The 24 h S. mutans biofilm formed on a flat agar surface; visible polymeric Extracellular Matrix (ECM), which has an open architecture with nutrient channels, and other properties; ( C ) The 24 h C. albicans biofilm formed on a flat agar surface (Agar Scientific, Stansted, UK). C. albicans adheres to the polystyrene surface, mainly using mycelial forms, visible pseudohyphae, budding yeast, and the so-called germ tube, considered to be the key features of the pathogenicity of the fungus. The culture was maintained at 36 °C, pH 7.0 and pCO 2 5%, in bovine serum as a medium additive promoting growth of the culture in the presence of a sucrose substrate (5%); ( D ) The 24 h C. albicans biofilm formed on a flat agar surface (Agar Scientific, Stansted, UK) under the influence of Lactobacillus salivarius (HM6 Paradens). There was no clear, compact structure for the C. albicans biofilm and single loosely located budding cells. Other morphological forms of yeasts were invisible; ( E ) The 24 h double-species oral streptococci/yeasts biofilm formed on a flat agar surface (Agar Scientific, Stansted, UK). There was an apparent change in the C. albicans morphotype in the S. mutans common culture and visible pleomorphic forms were true hyphae and, blastoconidia, which in the mixed culture also produce mycelial forms, whose role is related to damage to immune cells (macrophages), leading to microorganism invasion. There was abundant extracellular matrix between cells and covering bacterial and yeast cells. Bacterial cells were visible in chains adhering to yeast cells and wrapped around them; ( F ) The 24 h double-species S. mutans / C. albicans biofilm formed on a flat agar surface (Agar Scientific, Stansted, UK) under the influence of Lactobacillus salivarius (HM6 Paradens). There was no clear, compact S. mutans / C. albicans biofilm structure or single loosely located budding cells. Other morphological forms of C. albicans were invisible. There was no visible extracellular matrix. Original magnification: 4000× and 6000×.

    Journal: Nutrients

    Article Title: Effect of a Lactobacillus Salivarius Probiotic on a Double-Species Streptococcus Mutans and Candida Albicans Caries Biofilm

    doi: 10.3390/nu9111242

    Figure Lengend Snippet: SEM images of the mono-species biofilm, generated by S. mutans , C. albicans and the double-species oral streptococci/yeasts biofilm—untreated and treated with the Lactobacillus salivarius (HM6 Paradens)—after 24 h of biofilm formation. ( A ) The 14 h S. mutans biofilm formed on a flat agar surface (Agar Scientific, Stansted, UK). S. mutans adheres to the polystyrene surface, mainly using a sucrose-dependent mechanism; ( B ) The 24 h S. mutans biofilm formed on a flat agar surface; visible polymeric Extracellular Matrix (ECM), which has an open architecture with nutrient channels, and other properties; ( C ) The 24 h C. albicans biofilm formed on a flat agar surface (Agar Scientific, Stansted, UK). C. albicans adheres to the polystyrene surface, mainly using mycelial forms, visible pseudohyphae, budding yeast, and the so-called germ tube, considered to be the key features of the pathogenicity of the fungus. The culture was maintained at 36 °C, pH 7.0 and pCO 2 5%, in bovine serum as a medium additive promoting growth of the culture in the presence of a sucrose substrate (5%); ( D ) The 24 h C. albicans biofilm formed on a flat agar surface (Agar Scientific, Stansted, UK) under the influence of Lactobacillus salivarius (HM6 Paradens). There was no clear, compact structure for the C. albicans biofilm and single loosely located budding cells. Other morphological forms of yeasts were invisible; ( E ) The 24 h double-species oral streptococci/yeasts biofilm formed on a flat agar surface (Agar Scientific, Stansted, UK). There was an apparent change in the C. albicans morphotype in the S. mutans common culture and visible pleomorphic forms were true hyphae and, blastoconidia, which in the mixed culture also produce mycelial forms, whose role is related to damage to immune cells (macrophages), leading to microorganism invasion. There was abundant extracellular matrix between cells and covering bacterial and yeast cells. Bacterial cells were visible in chains adhering to yeast cells and wrapped around them; ( F ) The 24 h double-species S. mutans / C. albicans biofilm formed on a flat agar surface (Agar Scientific, Stansted, UK) under the influence of Lactobacillus salivarius (HM6 Paradens). There was no clear, compact S. mutans / C. albicans biofilm structure or single loosely located budding cells. Other morphological forms of C. albicans were invisible. There was no visible extracellular matrix. Original magnification: 4000× and 6000×.

    Article Snippet: Scanning Electron Microscopic Analysis of Biofilm Microbial biofilms were grown on 13 mm diameter round basic slides (Agar Scientific, Stansted, UK) in the wells of a 24-well plate, according to the protocol described above.

    Techniques: Generated

    Scanning electron microscopy (SEM). ( A – C ) images of the double-species biofilm formed by C. albicans and S. mutans , after 24 ( A ) and 48 ( B , C ) h of biofilm formation. Culture was maintained at 37 °C, pH 7.0 and pCO 2 5%, in bovine serum as a medium additive promoting the growth of the culture in the presence of a sucrose substrate (5%). Original magnification: 400× and 6000×.

    Journal: Nutrients

    Article Title: Effect of a Lactobacillus Salivarius Probiotic on a Double-Species Streptococcus Mutans and Candida Albicans Caries Biofilm

    doi: 10.3390/nu9111242

    Figure Lengend Snippet: Scanning electron microscopy (SEM). ( A – C ) images of the double-species biofilm formed by C. albicans and S. mutans , after 24 ( A ) and 48 ( B , C ) h of biofilm formation. Culture was maintained at 37 °C, pH 7.0 and pCO 2 5%, in bovine serum as a medium additive promoting the growth of the culture in the presence of a sucrose substrate (5%). Original magnification: 400× and 6000×.

    Article Snippet: Scanning Electron Microscopic Analysis of Biofilm Microbial biofilms were grown on 13 mm diameter round basic slides (Agar Scientific, Stansted, UK) in the wells of a 24-well plate, according to the protocol described above.

    Techniques: Electron Microscopy

    Viable counts of each bacterial species present in the initial inoculum (per well) and in the 3‐day old biofilms ( n = 7). Fn = Fusobacterium nucleatum ; Pg = Porphyromonas gingivalis ; Aa = Aggregatibacter actinomycetemcomitans . Error bars: standard error

    Journal: Clinical and Experimental Dental Research

    Article Title: The effect of metronidazole plus amoxicillin or metronidazole plus penicillin V on periodontal pathogens in an in vitro biofilm model. The effect of metronidazole plus amoxicillin or metronidazole plus penicillin V on periodontal pathogens in an in vitro biofilm model

    doi: 10.1002/cre2.96

    Figure Lengend Snippet: Viable counts of each bacterial species present in the initial inoculum (per well) and in the 3‐day old biofilms ( n = 7). Fn = Fusobacterium nucleatum ; Pg = Porphyromonas gingivalis ; Aa = Aggregatibacter actinomycetemcomitans . Error bars: standard error

    Article Snippet: 2.2 Biofilm construction Biofilms were prepared in 24‐well plates (Corning Inc., NY, USA) by adapting a previously described protocol (Sanchez et al., ).

    Techniques:

    CLSM images of 3‐day old biofilm after antibiotic exposure. Overlapping of images collected from green and red channel. (a) 3‐day‐old biofilm (negative control); (b) 3‐day old biofilm CHX‐treated for 2 hr; (c) 3‐day old biofilm treated with AMX + MET in high concentration for 2 hr; (d) 3‐day old biofilm treated with PV + MET in high concentration for 2 hr. Z‐stacks were taken in xyz projection with 63× objective, oil immersion, at 10 μm from the biofilm bottom. Scale bar: 10 μm

    Journal: Clinical and Experimental Dental Research

    Article Title: The effect of metronidazole plus amoxicillin or metronidazole plus penicillin V on periodontal pathogens in an in vitro biofilm model. The effect of metronidazole plus amoxicillin or metronidazole plus penicillin V on periodontal pathogens in an in vitro biofilm model

    doi: 10.1002/cre2.96

    Figure Lengend Snippet: CLSM images of 3‐day old biofilm after antibiotic exposure. Overlapping of images collected from green and red channel. (a) 3‐day‐old biofilm (negative control); (b) 3‐day old biofilm CHX‐treated for 2 hr; (c) 3‐day old biofilm treated with AMX + MET in high concentration for 2 hr; (d) 3‐day old biofilm treated with PV + MET in high concentration for 2 hr. Z‐stacks were taken in xyz projection with 63× objective, oil immersion, at 10 μm from the biofilm bottom. Scale bar: 10 μm

    Article Snippet: 2.2 Biofilm construction Biofilms were prepared in 24‐well plates (Corning Inc., NY, USA) by adapting a previously described protocol (Sanchez et al., ).

    Techniques: Confocal Laser Scanning Microscopy, Negative Control, Concentration Assay

    Viable counts of each bacterial species in the 3‐day old biofilms (negative controls) and subjected to antiseptic or antibiotic treatment ( n = 6). No live CFU were retrieved from biofilms treated with CHX (positive controls). Single green bars: number of live Aggregatibacter actinomycetemcomitans retrieved from biofilms exposed to antibiotic combinations in high (H) or low (L) concentrations. * shows statistical significance between biofilms exposed to PV + MET in high and low concentration ( p = .041, t test). Fn = Fusobacterium nucleatum ; Pg = Porphyromonas gingivalis ; Aa = A . actinomycetemcomitans . Error bars: standard error

    Journal: Clinical and Experimental Dental Research

    Article Title: The effect of metronidazole plus amoxicillin or metronidazole plus penicillin V on periodontal pathogens in an in vitro biofilm model. The effect of metronidazole plus amoxicillin or metronidazole plus penicillin V on periodontal pathogens in an in vitro biofilm model

    doi: 10.1002/cre2.96

    Figure Lengend Snippet: Viable counts of each bacterial species in the 3‐day old biofilms (negative controls) and subjected to antiseptic or antibiotic treatment ( n = 6). No live CFU were retrieved from biofilms treated with CHX (positive controls). Single green bars: number of live Aggregatibacter actinomycetemcomitans retrieved from biofilms exposed to antibiotic combinations in high (H) or low (L) concentrations. * shows statistical significance between biofilms exposed to PV + MET in high and low concentration ( p = .041, t test). Fn = Fusobacterium nucleatum ; Pg = Porphyromonas gingivalis ; Aa = A . actinomycetemcomitans . Error bars: standard error

    Article Snippet: 2.2 Biofilm construction Biofilms were prepared in 24‐well plates (Corning Inc., NY, USA) by adapting a previously described protocol (Sanchez et al., ).

    Techniques: Concentration Assay

    CLSM image in maximum projection of the series taken in xzy axis of the 3‐day old biofilm. Viable and nonviable bacterial cells are depicted in green and red, respectively. Scale bar: 10 μm

    Journal: Clinical and Experimental Dental Research

    Article Title: The effect of metronidazole plus amoxicillin or metronidazole plus penicillin V on periodontal pathogens in an in vitro biofilm model. The effect of metronidazole plus amoxicillin or metronidazole plus penicillin V on periodontal pathogens in an in vitro biofilm model

    doi: 10.1002/cre2.96

    Figure Lengend Snippet: CLSM image in maximum projection of the series taken in xzy axis of the 3‐day old biofilm. Viable and nonviable bacterial cells are depicted in green and red, respectively. Scale bar: 10 μm

    Article Snippet: 2.2 Biofilm construction Biofilms were prepared in 24‐well plates (Corning Inc., NY, USA) by adapting a previously described protocol (Sanchez et al., ).

    Techniques: Confocal Laser Scanning Microscopy

    Confocal Laser Scanning Microscopy of biofilm formed by S. pseudintermedius strain DSM 25713. Biofilm was allowed to form for 48 h at 37 °C, in absence of serum, under both ( a ) static, and ( b ) dynamic (flow cell chamber) conditions. Static biofilms were further treated for 24 h with increasing gentamicin concentrations (1x-128xMIC). Representative images of biofilm exposed at ( c ) 1x and ( d ) 128xMIC gentamicin are shown. Orthogonal images z are projections of x and y planes, collected within the biofilm as indicated by the green and red lines in the top view. Image capture was set for simultaneous visualization of red (Propidium iodide-stained dead cells), green (Syto-9-stained viable cells), and blue (Concanavalin A-stained EPS) fluorescence. Magnification, x100

    Journal: BMC Microbiology

    Article Title: New insights in Staphylococcus pseudintermedius pathogenicity: antibiotic-resistant biofilm formation by a human wound-associated strain

    doi: 10.1186/s12866-015-0449-x

    Figure Lengend Snippet: Confocal Laser Scanning Microscopy of biofilm formed by S. pseudintermedius strain DSM 25713. Biofilm was allowed to form for 48 h at 37 °C, in absence of serum, under both ( a ) static, and ( b ) dynamic (flow cell chamber) conditions. Static biofilms were further treated for 24 h with increasing gentamicin concentrations (1x-128xMIC). Representative images of biofilm exposed at ( c ) 1x and ( d ) 128xMIC gentamicin are shown. Orthogonal images z are projections of x and y planes, collected within the biofilm as indicated by the green and red lines in the top view. Image capture was set for simultaneous visualization of red (Propidium iodide-stained dead cells), green (Syto-9-stained viable cells), and blue (Concanavalin A-stained EPS) fluorescence. Magnification, x100

    Article Snippet: Time course of biofilm formation Biofilms were allowed to form in each well of a 24-well flat-bottom polystyrene tissue-treated microtiter plate (BD Company), as described above.

    Techniques: Confocal Laser Scanning Microscopy, Flow Cytometry, Staining, Fluorescence

    Kinetic of biofilm formation, through 72 h-incubation, by S. pseudintermedius strain DSM 25713 onto polystyrene. ( a - f ) Representative SEM images of biofilm formation after 1, 4, 8, 24, 48, and 72 h of incubation, respectively. Magnification (x1.000). ( g , h ) Magnification (x20.000) of ( e ) and ( f ), respectively. Cocci are surrounded by EPS appearing as an extensive network of filaments stretched among cells and between these and the substratum. ( i ) Kinetic of biofilm formation as assessed by viable count. Maximum, median, and minimum values are shown in each box (n = 6)

    Journal: BMC Microbiology

    Article Title: New insights in Staphylococcus pseudintermedius pathogenicity: antibiotic-resistant biofilm formation by a human wound-associated strain

    doi: 10.1186/s12866-015-0449-x

    Figure Lengend Snippet: Kinetic of biofilm formation, through 72 h-incubation, by S. pseudintermedius strain DSM 25713 onto polystyrene. ( a - f ) Representative SEM images of biofilm formation after 1, 4, 8, 24, 48, and 72 h of incubation, respectively. Magnification (x1.000). ( g , h ) Magnification (x20.000) of ( e ) and ( f ), respectively. Cocci are surrounded by EPS appearing as an extensive network of filaments stretched among cells and between these and the substratum. ( i ) Kinetic of biofilm formation as assessed by viable count. Maximum, median, and minimum values are shown in each box (n = 6)

    Article Snippet: Time course of biofilm formation Biofilms were allowed to form in each well of a 24-well flat-bottom polystyrene tissue-treated microtiter plate (BD Company), as described above.

    Techniques: Incubation

    ESEM images of biofilm formed by S. pseudintermedius strain DSM 25713 onto polystyrene following 72 h-incubation. ( a ) Biofilm exhibited spatially heterogeneous organization, as suggested by the presence of “mushroom-like” structures (as indicated by arrows). Magnification: x3.000. ( b , c ) Multilayered organization with the presence of bacteria under EPS matrix (as indicated by arrows). Magnification: x12.500 and x20.000, respectively

    Journal: BMC Microbiology

    Article Title: New insights in Staphylococcus pseudintermedius pathogenicity: antibiotic-resistant biofilm formation by a human wound-associated strain

    doi: 10.1186/s12866-015-0449-x

    Figure Lengend Snippet: ESEM images of biofilm formed by S. pseudintermedius strain DSM 25713 onto polystyrene following 72 h-incubation. ( a ) Biofilm exhibited spatially heterogeneous organization, as suggested by the presence of “mushroom-like” structures (as indicated by arrows). Magnification: x3.000. ( b , c ) Multilayered organization with the presence of bacteria under EPS matrix (as indicated by arrows). Magnification: x12.500 and x20.000, respectively

    Article Snippet: Time course of biofilm formation Biofilms were allowed to form in each well of a 24-well flat-bottom polystyrene tissue-treated microtiter plate (BD Company), as described above.

    Techniques: Incubation

    Standardization of experimental conditions for biofilm formation by S. pseudintermedius strain DSM 25713 on polystyrene surface. Effect of dynamic (filled squares) or static (filled triangles) incubation, incubation time (24, 48, and 72 h), and inoculum concentration (10 5 , 10 6 , and 10 7 CFU/mL) on biofilm biomass formation, as assessed by spectrophotometric assay. Values are means ± SDs (n = 6). *** p

    Journal: BMC Microbiology

    Article Title: New insights in Staphylococcus pseudintermedius pathogenicity: antibiotic-resistant biofilm formation by a human wound-associated strain

    doi: 10.1186/s12866-015-0449-x

    Figure Lengend Snippet: Standardization of experimental conditions for biofilm formation by S. pseudintermedius strain DSM 25713 on polystyrene surface. Effect of dynamic (filled squares) or static (filled triangles) incubation, incubation time (24, 48, and 72 h), and inoculum concentration (10 5 , 10 6 , and 10 7 CFU/mL) on biofilm biomass formation, as assessed by spectrophotometric assay. Values are means ± SDs (n = 6). *** p

    Article Snippet: Time course of biofilm formation Biofilms were allowed to form in each well of a 24-well flat-bottom polystyrene tissue-treated microtiter plate (BD Company), as described above.

    Techniques: Incubation, Concentration Assay, Spectrophotometric Assay

    Effect of serum and pH on biofilm formation and growth by S. pseudintermedius strain DSM 25713. ( a ) Serum was tested against biofilm formation at various dilutions (1:2, 1:10, and 1:100), as free or adsorbed to polystyrene, under different pH (5.5, 7.1, and 8.7). Control wells contained bacteria but not serum. Biofilm biomass amount was measured by crystal violet assay, then normalized on bacterial growth by calculating the specific biofilm formation (SBF) index (see Materials and Methods). ( b ) The effect of free serum against bacterial growth was assessed by measuring OD 600 of cell grown in broth following 24 h-incubation. Results are means + SDs (n = 9). * p

    Journal: BMC Microbiology

    Article Title: New insights in Staphylococcus pseudintermedius pathogenicity: antibiotic-resistant biofilm formation by a human wound-associated strain

    doi: 10.1186/s12866-015-0449-x

    Figure Lengend Snippet: Effect of serum and pH on biofilm formation and growth by S. pseudintermedius strain DSM 25713. ( a ) Serum was tested against biofilm formation at various dilutions (1:2, 1:10, and 1:100), as free or adsorbed to polystyrene, under different pH (5.5, 7.1, and 8.7). Control wells contained bacteria but not serum. Biofilm biomass amount was measured by crystal violet assay, then normalized on bacterial growth by calculating the specific biofilm formation (SBF) index (see Materials and Methods). ( b ) The effect of free serum against bacterial growth was assessed by measuring OD 600 of cell grown in broth following 24 h-incubation. Results are means + SDs (n = 9). * p

    Article Snippet: Time course of biofilm formation Biofilms were allowed to form in each well of a 24-well flat-bottom polystyrene tissue-treated microtiter plate (BD Company), as described above.

    Techniques: Crystal Violet Assay, Incubation

    In vitro effect of antibiotics against preformed biofilm by S. pseudintermedius strain DSM 25713. Biofilms allowed to form following 48 h-incubation were exposed for further 24 h to each antibiotic at concentrations equal or multiple of MIC. Results are expressed as percentage of biofilm’s viability – as assessed by viable colony count - compared to control (unexposed, 100 % viability) (n = 6). The dotted line indicates a reduction in biofilm viability of at least 20 % vs control ( p

    Journal: BMC Microbiology

    Article Title: New insights in Staphylococcus pseudintermedius pathogenicity: antibiotic-resistant biofilm formation by a human wound-associated strain

    doi: 10.1186/s12866-015-0449-x

    Figure Lengend Snippet: In vitro effect of antibiotics against preformed biofilm by S. pseudintermedius strain DSM 25713. Biofilms allowed to form following 48 h-incubation were exposed for further 24 h to each antibiotic at concentrations equal or multiple of MIC. Results are expressed as percentage of biofilm’s viability – as assessed by viable colony count - compared to control (unexposed, 100 % viability) (n = 6). The dotted line indicates a reduction in biofilm viability of at least 20 % vs control ( p

    Article Snippet: Time course of biofilm formation Biofilms were allowed to form in each well of a 24-well flat-bottom polystyrene tissue-treated microtiter plate (BD Company), as described above.

    Techniques: In Vitro, Incubation

    In vitro activity of antibiotics at sub-inhibitory concentrations against biofilm formation by S. pseudintermedius strain DSM 25713. Biofilm biomass formed during 24 h-incubation was measured, using the crystal violet assay, in the presence of antibiotics at concentrations equal to 1/2x, 1/4x, and 1/8xMIC. Results were plotted as percentage of biofilm biomass formed in the presence of antibiotic, compared to controls (not exposed, 100 % biofilm biomass) (n = 6). The dotted line indicates a reduction in biofilm biomass of at least 20 % vs control ( p

    Journal: BMC Microbiology

    Article Title: New insights in Staphylococcus pseudintermedius pathogenicity: antibiotic-resistant biofilm formation by a human wound-associated strain

    doi: 10.1186/s12866-015-0449-x

    Figure Lengend Snippet: In vitro activity of antibiotics at sub-inhibitory concentrations against biofilm formation by S. pseudintermedius strain DSM 25713. Biofilm biomass formed during 24 h-incubation was measured, using the crystal violet assay, in the presence of antibiotics at concentrations equal to 1/2x, 1/4x, and 1/8xMIC. Results were plotted as percentage of biofilm biomass formed in the presence of antibiotic, compared to controls (not exposed, 100 % biofilm biomass) (n = 6). The dotted line indicates a reduction in biofilm biomass of at least 20 % vs control ( p

    Article Snippet: Time course of biofilm formation Biofilms were allowed to form in each well of a 24-well flat-bottom polystyrene tissue-treated microtiter plate (BD Company), as described above.

    Techniques: In Vitro, Activity Assay, Incubation, Crystal Violet Assay

    Biofilm elimination by MNP-enhanced oxidative cleavage

    Journal: Nanoscale

    Article Title: Ferromagnetic nanoparticles with peroxidase-like activity enhance the cleavage of biological macromolecules for biofilm elimination

    doi: 10.1039/c3nr05422e

    Figure Lengend Snippet: Biofilm elimination by MNP-enhanced oxidative cleavage

    Article Snippet: Pseudomonas aeruginosa (PA01) was cultured in biofilm minimal media (M63, Amresco) supplemented with 0.2% Glucose, 0.5% Casamino acids (BD), and 1mM MgSO4 , in a 96 well plate overnight at 37°C.

    Techniques:

    Quantification of biofilm formation by the wild-type strain (ATCC 17978), a stable knockout mutant strain lacking the gene A1S_0114 (ATCC Δ0114), the same mutant strain containing the pET-RA plasmid (ATCC Δ0114 + PETRA), and a mutant strain containing the pET-RA plasmid harboring the A1S_0114 gene (ATCC Δ0114 + PETRA + 0114).

    Journal: PLoS ONE

    Article Title: Whole Transcriptome Analysis of Acinetobacter baumannii Assessed by RNA-Sequencing Reveals Different mRNA Expression Profiles in Biofilm Compared to Planktonic Cells

    doi: 10.1371/journal.pone.0072968

    Figure Lengend Snippet: Quantification of biofilm formation by the wild-type strain (ATCC 17978), a stable knockout mutant strain lacking the gene A1S_0114 (ATCC Δ0114), the same mutant strain containing the pET-RA plasmid (ATCC Δ0114 + PETRA), and a mutant strain containing the pET-RA plasmid harboring the A1S_0114 gene (ATCC Δ0114 + PETRA + 0114).

    Article Snippet: The complete mRNA transcriptomic profiles of exponentially growing and stationary-phases cultures and from biofilm cells were obtained by Illumina procedures.

    Techniques: Knock-Out, Mutagenesis, Positron Emission Tomography, Plasmid Preparation

    Sequence distribution of the 1621 genes identified in the present work as up-regulated in biofilm vs. stationary phase cells. Genes involved in: A) biological processes, B) cellular components, and C) molecular functions. The results were filtered by the number of sequences (cutoff = 40, 5, and 80, respectively).

    Journal: PLoS ONE

    Article Title: Whole Transcriptome Analysis of Acinetobacter baumannii Assessed by RNA-Sequencing Reveals Different mRNA Expression Profiles in Biofilm Compared to Planktonic Cells

    doi: 10.1371/journal.pone.0072968

    Figure Lengend Snippet: Sequence distribution of the 1621 genes identified in the present work as up-regulated in biofilm vs. stationary phase cells. Genes involved in: A) biological processes, B) cellular components, and C) molecular functions. The results were filtered by the number of sequences (cutoff = 40, 5, and 80, respectively).

    Article Snippet: The complete mRNA transcriptomic profiles of exponentially growing and stationary-phases cultures and from biofilm cells were obtained by Illumina procedures.

    Techniques: Sequencing

    Sequence distribution of genes expressed only in biofilm-associated cells and inhibited in planktonic cells. Genes involved in A) biological processes, B) molecular functions, and C) cellular components. The results were filtered by the number of sequences (cutoff = 1, 4, and 1, respectively).

    Journal: PLoS ONE

    Article Title: Whole Transcriptome Analysis of Acinetobacter baumannii Assessed by RNA-Sequencing Reveals Different mRNA Expression Profiles in Biofilm Compared to Planktonic Cells

    doi: 10.1371/journal.pone.0072968

    Figure Lengend Snippet: Sequence distribution of genes expressed only in biofilm-associated cells and inhibited in planktonic cells. Genes involved in A) biological processes, B) molecular functions, and C) cellular components. The results were filtered by the number of sequences (cutoff = 1, 4, and 1, respectively).

    Article Snippet: The complete mRNA transcriptomic profiles of exponentially growing and stationary-phases cultures and from biofilm cells were obtained by Illumina procedures.

    Techniques: Sequencing

    Quantification of biofilm formation by the wild-type strain (ATCC 17978) and strains with chromosomal disruptions in the genes A1S_0114, A1S_0302, A1S_1507, A1S_3168 and A1S_2042.

    Journal: PLoS ONE

    Article Title: Whole Transcriptome Analysis of Acinetobacter baumannii Assessed by RNA-Sequencing Reveals Different mRNA Expression Profiles in Biofilm Compared to Planktonic Cells

    doi: 10.1371/journal.pone.0072968

    Figure Lengend Snippet: Quantification of biofilm formation by the wild-type strain (ATCC 17978) and strains with chromosomal disruptions in the genes A1S_0114, A1S_0302, A1S_1507, A1S_3168 and A1S_2042.

    Article Snippet: The complete mRNA transcriptomic profiles of exponentially growing and stationary-phases cultures and from biofilm cells were obtained by Illumina procedures.

    Techniques: