scholarly journals Individual or Combined Effects of Meropenem, Imipenem, Sulbactam, Colistin, and Tigecycline on Biofilm-Embedded Acinetobacter baumannii and Biofilm Architecture

2016 ◽  
Vol 60 (8) ◽  
pp. 4670-4676 ◽  
Author(s):  
Yung-Chih Wang ◽  
Shu-Chen Kuo ◽  
Ya-Sung Yang ◽  
Yi-Tzu Lee ◽  
Chun-Hsiang Chiu ◽  
...  
Keyword(s):  
Acinetobacter Baumannii ◽  
Laser Scanning ◽  
Bactericidal Effect ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Roughness Coefficient ◽  
Content Type ◽  
Biofilm Thickness ◽  
Carbapenem Resistant ◽  
Biofilm Architecture

ABSTRACTAcinetobacter baumanniibiofilms are difficult to eradicate. We investigated the effects of meropenem (2 mg/liter), imipenem (2 mg/liter), sulbactam (4 mg/liter), colistin (2 mg/liter), and tigecycline (2 mg/liter), alone or in combination, on biofilm-embedded carbapenem-resistant and carbapenem-susceptibleA. baumannii(CRAb and CSAb, respectively) cells, as well as on the architecture of the biofilms.A. baumanniiATCC 15151 (Ab15151) and its OXA-82-overproducing transformant, along with two clinical CSAb and two clinical CRAb isolates of differing clonalities, were used. The minimal bactericidal concentrations for biofilm-embedded cells of the six tested isolates were >50-fold those of their planktonic cells. When used individually, meropenem exhibited a higher killing effect than the other four antimicrobials on biofilm-embedded CSAb cells in the colony biofilm assay. For two clinical CRAb isolates, meropenem plus sulbactam or sulbactam plus tigecycline showed >100-fold the bactericidal effect exhibited by these agents used alone after 48 h of treatment. The effect of antimicrobials on the architecture of Ab15151 biofilm emitting green fluorescence was determined by confocal laser scanning microscopy using COMSTAT software. Significant decreases in the maximum biofilm thickness were observed after exposure to meropenem and imipenem. Meropenem plus sulbactam significantly decreased the biomass and mean thickness and increased the roughness coefficient of biofilms, but sulbactam plus tigecycline only decreased the maximum and mean biofilm thickness compared to any of these agents used alone. Meropenem was active against biofilm-embedded CSAb, whereas meropenem plus sulbactam exhibited synergism against biofilm-embedded CRAb and caused significantly more damage to the biofilm architecture than did any of the agents used alone.

scholarly journals Sequential Treatment of Biofilms with Aztreonam and Tobramycin Is a Novel Strategy for Combating Pseudomonas aeruginosa Chronic Respiratory Infections

2016 ◽  
Vol 60 (5) ◽  
pp. 2912-2922 ◽  
Author(s):  
Estrella Rojo-Molinero ◽  
María D. Macià ◽  
Rosa Rubio ◽  
Bartolomé Moyà ◽  
Gabriel Cabot ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Laser Scanning ◽  
Bactericidal Effect ◽  
Cell System ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Antibiofilm Activity ◽  
Content Type ◽  
Inhaled Antibiotics ◽  
Resistant Mutants

ABSTRACTTraditional therapeutic strategies to control chronic colonization in cystic fibrosis (CF) patients are based on the use of a single nebulized antibiotic. In this study, we evaluated the therapeutic efficacy and dynamics of antibiotic resistance inPseudomonas aeruginosabiofilms under sequential therapy with inhaled aztreonam (ATM) and tobramycin (TOB). Laboratory strains PAO1, PAOMS (hypermutable), PAOMA (mucoid), and PAOMSA (mucoid and hypermutable) and two hypermutable CF strains, 146-HSE (Liverpool epidemic strain [LES-1]) and 1089-HSE (ST1089), were used. Biofilms were developed using the flow cell system. Mature biofilms were challenged with peak and 1/10-peak concentrations of ATM (700 mg/liter and 70 mg/liter), TOB (1,000 mg/liter and 100 mg/liter), and their alternations (ATM/TOB/ATM and TOB/ATM/TOB) for 2 (t= 2), 4 (t= 4), and 6 days (t= 6). The numbers of viable cells (CFU) and resistant mutants were determined. Biofilm structural dynamics were monitored by confocal laser scanning microscopy and processed with COMSTAT and IMARIS software programs. TOB monotherapy produced an intense decrease in CFU that was not always correlated with a reduction in biomass and/or a bactericidal effect on biofilms, particularly for the CF strains. The ATM monotherapy bactericidal effect was lower, but effects on biofilm biomass and/or structure, including intense filamentation, were documented. The alternation of TOB and ATM led to an enhancement of the antibiofilm activity against laboratory and CF strains compared to that with the individual regimens, potentiating the bactericidal effect and/or the reduction in biomass, particularly at peak concentrations. Resistant mutants were not documented in any of the regimens at the peak concentrations and only anecdotally at the 1/10-peak concentrations. These results support the clinical evaluation of sequential regimens with inhaled antibiotics in CF, as opposed to the current maintenance treatments with just one antibiotic in monotherapy.

scholarly journals Microscopic and Spectroscopic Analyses of Chlorhexidine Tolerance in Delftia acidovorans Biofilms

2014 ◽  
Vol 58 (10) ◽  
pp. 5673-5686 ◽  
Author(s):  
Tara Rema ◽  
John R. Lawrence ◽  
James J. Dynes ◽  
Adam P. Hitchcock ◽  
Darren R. Korber
Keyword(s):  
Laser Scanning ◽  
Cell Types ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Content Type ◽  
Flow Cells ◽  
Biofilm Thickness ◽  
Delftia Acidovorans ◽  
Scanning Transmission ◽  
Negative Effect

ABSTRACTThe physicochemical responses ofDelftia acidovoransbiofilms exposed to the commonly used antimicrobial chlorhexidine (CHX) were examined in this study. A CHX-sensitive mutant (MIC, 1.0 μg ml−1) was derived from a CHX-tolerant (MIC, 15.0 μg ml−1)D. acidovoransparent strain using transposon mutagenesis.D. acidovoransmutant (MT51) and wild-type (WT15) strain biofilms were cultivated in flow cells and then treated with CHX at sub-MIC and inhibitory concentrations and examined by confocal laser scanning microscopy (CLSM), scanning transmission X-ray microscopy (STXM), and infrared (IR) spectroscopy. Specific morphological, structural, and chemical compositional differences between the CHX-treated and -untreated biofilms of both strains were observed. Apart from architectural differences, CLSM revealed a negative effect of CHX on biofilm thickness in the CHX-sensitive MT51 biofilms relative to those of the WT15 strain. STXM analyses showed that the WT15 biofilms contained two morphochemical cell variants, whereas only one type was detected in the MT51 biofilms. The cells in the MT51 biofilms bioaccumulated CHX to a similar extent as one of the cell types found in the WT15 biofilms, whereas the other cell type in the WT15 biofilms did not bioaccumulate CHX. STXM and IR spectral analyses revealed that CHX-sensitive MT51 cells accumulated the highest levels of CHX. Pretreating biofilms with EDTA promoted the accumulation of CHX in all cells. Thus, it is suggested that a subpopulation of cells that do not accumulate CHX appear to be responsible for greater CHX resistance inD. acidovoransWT15 biofilm in conjunction with the possible involvement of bacterial membrane stability.

scholarly journals Role of Exopolysaccharides in Pseudomonas aeruginosa Biofilm Formation and Architecture

2011 ◽  
Vol 77 (15) ◽  
pp. 5238-5246 ◽  
Author(s):  
Aamir Ghafoor ◽  
Iain D. Hay ◽  
Bernd H. A. Rehm
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Biofilm Formation ◽  
Laser Scanning ◽  
Bacterial Biofilm ◽  
Laser Scanning Microscopy ◽  
Extracellular Dna ◽  
Confocal Laser ◽  
Content Type ◽  
Biofilm Architecture

ABSTRACTPseudomonas aeruginosais an opportunistic human pathogen and has been established as a model organism to study bacterial biofilm formation. At least three exopolysaccharides (alginate, Psl, and Pel) contribute to the formation of biofilms in this organism. Here mutants deficient in the production of one or more of these polysaccharides were generated to investigate how these polymers interactively contribute to biofilm formation. Confocal laser scanning microscopy of biofilms formed in flow chambers showed that mutants deficient in alginate biosynthesis developed biofilms with a decreased proportion of viable cells than alginate-producing strains, indicating a role of alginate in viability of cells in biofilms. Alginate-deficient mutants showed enhanced extracellular DNA (eDNA)-containing surface structures impacting the biofilm architecture. PAO1 ΔpslAΔalg8overproduced Pel, and eDNA showing meshwork-like structures presumably based on an interaction between both polymers were observed. The formation of characteristic mushroom-like structures required both Psl and alginate, whereas Pel appeared to play a role in biofilm cell density and/or the compactness of the biofilm. Mutants producing only alginate, i.e., mutants deficient in both Psl and Pel production, lost their ability to form biofilms. A lack of Psl enhanced the production of Pel, and the absence of Pel enhanced the production of alginate. The function of Psl in attachment was independent of alginate and Pel. A 30% decrease in Psl promoter activity in the alginate-overproducing MucA-negative mutant PDO300 suggested inverse regulation of both biosynthesis operons. Overall, this study demonstrated that the various exopolysaccharides and eDNA interactively contribute to the biofilm architecture ofP. aeruginosa.

scholarly journals Identification and Characterization of an Acinetobacter baumannii Biofilm-Associated Protein

2007 ◽  
Vol 190 (3) ◽  
pp. 1036-1044 ◽  
Author(s):  
Thomas W. Loehfelm ◽  
Nicole R. Luke ◽  
Anthony A. Campagnari
Keyword(s):  
Cell Surface ◽  
Acinetobacter Baumannii ◽  
Laser Scanning ◽  
Time Course ◽  
Surface Reactivity ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Specific Cell ◽  
Military Health ◽  
Biofilm Thickness

ABSTRACT We have identified a homologue to the staphylococcal biofilm-associated protein (Bap) in a bloodstream isolate of Acinetobacter baumannii. The fully sequenced open reading frame is 25,863 bp and encodes a protein with a predicted molecular mass of 854 kDa. Analysis of the nucleotide sequence reveals a repetitive structure consistent with bacterial cell surface adhesins. Bap-specific monoclonal antibody (MAb) 6E3 was generated to an epitope conserved among 41% of A. baumannii strains isolated during a recent outbreak in the U.S. military health care system. Flow cytometry confirms that the MAb 6E3 epitope is surface exposed. Random transposon mutagenesis was used to generate A. baumannii bap1302::EZ-Tn5, a mutant negative for surface reactivity to MAb 6E3 in which the transposon disrupts the coding sequence of bap. Time course confocal laser scanning microscopy and three-dimensional image analysis of actively growing biofilms demonstrates that this mutant is unable to sustain biofilm thickness and volume, suggesting a role for Bap in supporting the development of the mature biofilm structure. This is the first identification of a specific cell surface protein directly involved in biofilm formation by A. baumannii and suggests that Bap is involved in intercellular adhesion within the mature biofilm.

scholarly journals Gentian Violet Exhibits Activity against Biofilms Formed by Oral Candida Isolates Obtained from HIV-Infected Patients

2011 ◽  
Vol 55 (6) ◽  
pp. 3043-3045 ◽  
Author(s):  
Rana S. Traboulsi ◽  
Pranab K. Mukherjee ◽  
Jyotsna Chandra ◽  
Robert A. Salata ◽  
Richard Jurevic ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Laser Scanning ◽  
Gentian Violet ◽  
Dry Weight ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Content Type ◽  
Biofilm Thickness

ABSTRACTThe effect of gentian violet againstCandida albicansand non-Candida albicansbiofilms formed on polymethylmethacrylate strips was evaluated using a dry weight assay and confocal laser scanning microscopy. The ability of gentian violet to inhibitCandida albicansgermination was also assessed. Gentian violet activity againstCandidabiofilms was demonstrated by a reduction in dry weight, disruption of biofilm architecture, and reduced biofilm thickness. Additionally, gentian violet inhibitedCandidagermination in a concentration-dependent manner.

scholarly journals Anti-biofilm and anti-virulence effects of zerumbone against Acinetobacter baumannii

Microbiology ◽  
2020 ◽  
Vol 166 (8) ◽  
pp. 717-726
Author(s):  
Hye-Rim Kim ◽  
Da-Seul Shin ◽  
Hye-In Jang ◽  
Yong-Bin Eom
Keyword(s):  
Acinetobacter Baumannii ◽  
Biofilm Formation ◽  
Type Species ◽  
Laser Scanning ◽  
Multidrug Resistant ◽  
Laser Scanning Microscopy ◽  
Transcriptional Analysis ◽  
Confocal Laser ◽  
Content Type ◽  
Link Type

Acinetobacter baumannii is a multidrug-resistant opportunistic pathogen that affects patients with a compromised immune system and is becoming increasingly important as a hospital-derived infection. This pathogen is difficult to treat owing to its intrinsic multidrug resistance and ability to form antimicrobial-tolerant biofilms. In the present study, we aimed to assess the potential use of zerumbone as a novel anti-biofilm and/or anti-virulence agent against A. baumannii . The results showed that zerumbone at sub-inhibitory doses decreased biofilm formation and disrupted established A. baumannii biofilms. The zerumbone-induced decrease in biofilm formation was dose-dependent based on the results of microtitre plate biofilm assays and confocal laser scanning microscopy. In addition, our data validated the anti-virulence efficacy of zerumbone, wherein it significantly interfered with the motility of A. baumannii . To support these phenotypic results, transcriptional analysis revealed that zerumbone downregulated the expression of biofilm- and virulence-associated genes (adeA, adeB, adeC and bap) in A. baumannii . Overall, our findings suggested that zerumbone might be a promising bioactive agent for the treatment of biofilm- and virulence-related infections caused by multidrug-resistant A. baumannii .

The effect of N-acetylcysteine in a combined antibiofilm treatment against antibiotic-resistant Staphylococcus aureus

2020 ◽  
Vol 75 (7) ◽  
pp. 1787-1798
Author(s):  
Arthika Manoharan ◽  
Theerthankar Das ◽  
Gregory S Whiteley ◽  
Trevor Glasbey ◽  
Frederik H Kriel ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Dna Cleavage ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Combination Treatments ◽  
Resazurin Assay ◽  
Biofilm Architecture ◽  
Biofilm Disruption ◽  
The Uk

Abstract Background The WHO declared Staphylococcus aureus as a ‘pathogen of high importance’ in 2017. One-fifth of all bloodstream-related infections in Australia and 12 000 cases of bacteraemia in the UK (2017–18) were caused by the MRSA variant. To address the need for novel therapies, we investigated several permutations of an innovative combination therapy containing N-acetylcysteine (NAC), an antibiotic and an enzyme of choice in eradicating MRSA and MSSA biofilms. Methods Biofilm viability (resazurin assay) and colony count methods were used to investigate the effect of NAC, antibiotics and enzymes on S. aureus biofilm disruption and killing. The effects of NAC and enzymes on the polysaccharide content of biofilm matrices were analysed using the phenol/sulphuric acid method and the effect of NAC on DNA cleavage was determined using the Qubit fluorometer technique. Changes in biofilm architecture when subjected to NAC and enzymes were visualized using confocal laser scanning microscopy (CLSM). Results NAC alone displayed bacteriostatic effects when tested on planktonic bacterial growth. Combination treatments containing 30 mM NAC resulted in ≥90% disruption of biofilms across all MRSA and MSSA strains with a 2–3 log10 decrease in cfu/mL in treated biofilms. CLSM showed that NAC treatment drastically disrupted S. aureus biofilm architecture. There was also reduced polysaccharide production in MRSA biofilms in the presence of NAC. Conclusions Our results indicate that inclusion of NAC in a combination treatment is a promising strategy for S. aureus biofilm eradication. The intrinsic acidity of NAC was identified as key to maximum biofilm disruption and degradation of matrix components.

scholarly journals Bactericidal compounds controlling growth of the plant pathogen pseudomonas syringae pv. actinidiae, which forms biofilms composed of a novel exopolysaccharide

2020 ◽  
Author(s):  
S Ghods ◽  
Ian Sims ◽  
MF Moradali ◽  
BHA Rehma
Keyword(s):  
Pseudomonas Syringae ◽  
Chlorine Dioxide ◽  
Laser Scanning ◽  
Extracellular Polymeric Substances ◽  
Virulent Strain ◽  
Bactericidal Effect ◽  
Cell System ◽  
Laser Scanning Microscopy ◽  
Base Layer ◽  
Confocal Laser

© 2015, American Society for Microbiology. Pseudomonas syringae pv. actinidiae is the major cause of bacterial canker and is a severe threat to kiwifruit production worldwide. Many aspects of the disease caused by P. syringae pv. actinidiae, such as the pathogenicity-relevant formation of a biofilm composed of extracellular polymeric substances (EPSs), are still unknown. Here, a highly virulent strain of P. syringae pv. actinidiae, NZ V-13, was studied with respect to biofilm formation and architecture using a flow cell system combined with confocal laser scanning microscopy. The biofilm formed by P. syringae pv. actinidiae NZ V-13 was heterogeneous, consisting of a thin cellular base layer 5 μm thick and microcolonies with irregular structures. The major component of the EPSs produced by P. syringae pv. actinidiae NZ V-13 bacteria was isolated and identified to be an exopolysaccharide. Extensive compositional and structural analysis showed that rhamnose, fucose, and glucose were the major constituents, present at a ratio of 5:1.5:2. Experimental evidence that P. syringae pv. actinidiae NZ V-13 produces two polysaccharides, a branched α-D-rhamnan with side chains of terminal α-D-Fucf and an α-D-1,4-linked glucan, was obtained. The susceptibility of the cells in biofilms to kasugamycin and chlorine dioxide was assessed. About 64 and 73% of P. syringae pv. actinidiae NZ V-13 cells in biofilms were killed when kasugamycin and chlorine dioxide were used at 5 and 10 ppm, respectively. Kasugamycin inhibited the attachment of P. syringae pv. actinidiae NZ V-13 to solid surfaces at concentrations of 80 and 100 ppm. Kasugamycin was bacteriostatic against P. syringae pv. actinidiae NZ V-13 growth in the planktonic mode, with the MIC being 40 to 60 ppm and a bactericidal effect being found at 100 ppm. Here we studied the formation, architecture, and composition of P. syringae pv. actinidiae biofilms as well as used the biofilm as a model to assess the efficacies of bactericidal compounds.

scholarly journals Calcite Biomineralization by Bacterial Isolates from the Recently Discovered Pristine Karstic Herrenberg Cave

2011 ◽  
Vol 78 (4) ◽  
pp. 1157-1167 ◽  
Author(s):  
Anna Rusznyák ◽  
Denise M. Akob ◽  
Sándor Nietzsche ◽  
Karin Eusterhues ◽  
Kai Uwe Totsche ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Extracellular Polymeric Substances ◽  
Large Fraction ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Rrna Gene ◽  
Seepage Water ◽  
Bacterial Cells ◽  
Content Type ◽  
Rich Media

ABSTRACTKarstic caves represent one of the most important subterranean carbon storages on Earth and provide windows into the subsurface. The recent discovery of the Herrenberg Cave, Germany, gave us the opportunity to investigate the diversity and potential role of bacteria in carbonate mineral formation. Calcite was the only mineral observed by Raman spectroscopy to precipitate as stalactites from seepage water. Bacterial cells were found on the surface and interior of stalactites by confocal laser scanning microscopy. Proteobacteria dominated the microbial communities inhabiting stalactites, representing more than 70% of total 16S rRNA gene clones. Proteobacteria formed 22 to 34% of the detected communities in fluvial sediments, and a large fraction of these bacteria were also metabolically active. A total of 9 isolates, belonging to the generaArthrobacter,Flavobacterium,Pseudomonas,Rhodococcus,Serratia, andStenotrophomonas, grew on alkaline carbonate-precipitating medium. Two cultures with the most intense precipitate formation,Arthrobacter sulfonivoransandRhodococcus globerulus, grew as aggregates, produced extracellular polymeric substances (EPS), and formed mixtures of calcite, vaterite, and monohydrocalcite.R. globerulusformed idiomorphous crystals with rhombohedral morphology, whereasA. sulfonivoransformed xenomorphous globular crystals, evidence for taxon-specific crystal morphologies. The results of this study highlighted the importance of combining various techniques in order to understand the geomicrobiology of karstic caves, but further studies are needed to determine whether the mineralogical biosignatures found in nutrient-rich media can also be found in oligotrophic caves.

scholarly journals Alginate Oligosaccharide-Induced Modification of the lasI-lasR and rhlI-rhlR Quorum-Sensing Systems in Pseudomonas aeruginosa

2018 ◽  
Vol 62 (5) ◽  
Author(s):  
Alison A. Jack ◽  
Saira Khan ◽  
Lydia C. Powell ◽  
Manon F. Pritchard ◽  
Konrad Beck ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Quorum Sensing ◽  
Laser Scanning ◽  
Homoserine Lactone ◽  
Laser Scanning Microscopy ◽  
Extracellular Dna ◽  
Mass Spectrometry Analysis ◽  
Confocal Laser ◽  
Steric Interaction ◽  
Content Type

ABSTRACT Pseudomonas aeruginosa plays a major role in many chronic infections. Its ability to readily form biofilms contributes to its success as an opportunistic pathogen and its resistance/tolerance to antimicrobial/antibiotic therapy. A low-molecular-weight alginate oligomer (OligoG CF-5/20) derived from marine algae has previously been shown to impair motility in P. aeruginosa biofilms and disrupt pseudomonal biofilm assembly. As these bacterial phenotypes are regulated by quorum sensing (QS), we hypothesized that OligoG CF-5/20 may induce alterations in QS signaling in P. aeruginosa . QS regulation was studied by using Chromobacterium violaceum CV026 biosensor assays that showed a significant reduction in acyl homoserine lactone (AHL) production following OligoG CF-5/20 treatment (≥2%; P < 0.05). This effect was confirmed by liquid chromatography-mass spectrometry analysis of C 4 -AHL and 3-oxo-C 12 -AHL production (≥2%; P < 0.05). Moreover, quantitative PCR showed that reduced expression of both the las and rhl systems was induced following 24 h of treatment with OligoG CF-5/20 (≥0.2%; P < 0.05). Circular dichroism spectroscopy indicated that these alterations were not due to steric interaction between the AHL and OligoG CF-5/20. Confocal laser scanning microscopy (CLSM) and COMSTAT image analysis demonstrated that OligoG CF-5/20-treated biofilms had a dose-dependent decrease in biomass that was associated with inhibition of extracellular DNA synthesis (≥0.5%; P < 0.05). These changes correlated with alterations in the extracellular production of the pseudomonal virulence factors pyocyanin, rhamnolipids, elastase, and total protease ( P < 0.05). The ability of OligoG CF-5/20 to modify QS signaling in P. aeruginosa PAO1 may influence critical downstream functions such as virulence factor production and biofilm formation.

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