Synthesis of Antimicrobial Gallium Nanoparticles Using the Hot Injection Method.

Antibiotic resistance continues to be an ongoing problem in global public health despite interventions to reduce antibiotic overuse. Furthermore, it threatens to undo the achievements and progress of modern medicine. To address these issues, the development of new alternative treatments is needed. Metallic nanoparticles have become an increasingly attractive alternative due to their unique physicochemical properties that allow for different applications and their various mechanisms of action. In this study, gallium nanoparticles (Ga NPs) were tested against several clinical strains of Pseudomonas aeruginosa (DFU53, 364077, and 365707) and multi-drug-resistant Acinetobacter baumannii (MRAB). The results showed that Ga NPs did not inhibit bacterial growth when tested against the bacterial strains using a broth microdilution assay, but they exhibited effects in biofilm production in P. aeruginosa DFU53. Furthermore, as captured by atomic force microscopy imaging, P. aeruginosa DFU53 and MRAB biofilms underwent morphological changes, appearing rough and irregular when they were treated with Ga NPs. Although Ga NPs did not affect planktonic bacterial growth, their effects on both biofilm formation and established biofilm demonstrate their potential role in the race to combat antibiotic resistance, especially in biofilm-related infections.

N-acetylcysteine prevents catheter occlusion and inflammation in catheter associated-urinary tract infections by suppressing urease activity.

Introduction: Proteus mirabilis is a key pathobiont in catheter-associated urinary tract infections (CA-UTIs), which is well known to form crystalline biofilms that occlude catheters. Urease activity alkylates urine through the release of ammonia, consequentially resulting in higher levels of Mg2+ and Ca2+ and formation of crystals. In this study, we showed that N-acetyl cysteine (NAC), a thiol antioxidant, is a potent urease inhibitor that prevents crystalline biofilm formation. Methods: To quantify urease activity, Berthelot's method was done on bacterial extracts treated with NAC. We also used an in vitro catheterised glass bladder model to study the effect of NAC treatment on catheter occlusion and biofilm encrustation in P. mirabilis infections. Inductively-coupled plasma mass spectrometry (ICP-MS) was performed on catheter samples to decipher elemental profiles. Results: NAC inhibits urease activity of clinical P. mirabilis isolates at concentrations as low as 1 mM, independent of bacterial killing. The study also showed that NAC is bacteriostatic on P. mirabilis, and inhibited biofilm formation and catheter occlusion in an in vitro. A significant 4-8log10 reduction in viable bacteria was observed in catheters infected in this model. Additionally, biofilms in NAC treated catheters displayed a depletion of calcium, magnesium, or phosphates (>10 fold reduction), thus confirming the absence of any urease activity in the presence of NAC. Interestingly, we also showed that not only is NAC anti-inflammatory in bladder epithelial cells (BECs), but that it mutes its inflammatory response to urease and P. mirabilis infection by reducing the production of IL-6, IL-8 and IL-1b. Discussion: Using biochemical, microbiological and immunological techniques, this study displays the functionality of NAC in preventing catheter occlusion by inhibiting urease activity. The study also highlights NAC as a strong anti-inflammatory antibiofilm agent that can target both bacterial and host factors in the treatment of CA-UTIs.

Ascorbic acid modulates the structure of the Pseudomonas aeruginosa virulence factor pyocyanin and ascorbic acid-furanone-30 combination facilitate biofilm disruption.

The production of pyocyanin by Pseudomonas aeruginosa increases its virulence, fitness and biofilm formation. Pyocyanin is also a redox molecule and we hypothesize that ascorbic acid being an antioxidant will interact with pyocyanin. The main objective of this study was to investigate the potential interaction of ascorbic acid with pyocyanin, and also to investigate the impact of ascorbic acid in combination with Furanone-30 on quorum sensing and biofilm formation of P. aeruginosa. When incubated with ascorbic acid, hyperchromic and hypsochromic shifts in pyocyanin absorbance peaks at 385 nm and 695 nm were observed. In the presence of dehydroascorbic acid and citric acid, these shifts were absent, indicating that the intrinsic antioxidant property of ascorbic acid was probably essential in binding to pyocyanin. NMR spectroscopy showed shifts in 1H NMR pyocyanin peaks between 8.2 to 5.8 ppm when incubated in the presence of ascorbic acid. Density Functional Theory (DFT) supported potential interactions between the -CH2OH or -OH moieties of ascorbic acid with the -C=O moiety of pyocyanin. The pyocyanin-ascorbic acid complex impaired pyocyanin binding to DNA. Ascorbic acid combined with furanone-30 elevated quorum-sensing inhibition in P. aeruginosa, which was directly associated with significantly reduced P. aeruginosa virulence, adhesion, aggregation and biofilm formation and enhanced antibiotic-mediated bacterial killing. This study demonstrated that the antioxidant ascorbic acid directly binds to pyocyanin, modulates its structure and results in disruption of biofilm formation and associated tolerance to antibiotics.

Novel quinazolinone disulfide analogues as pqs quorum sensing inhibitors against Pseudomonas aeruginosa.

It is well established that the quorum sensing (QS) in Pseudomonas aeruginosa is primarily responsible for the synthesis and the release of several virulence factors including pyocyanin and are involved in biofilm formation. In the Pseudomonas quinolone signal (PQS) system, autoinducers such as PQS and HHQ bind and activate the transcription regulator protein receptor PqsR (MvfR). Targeting PqsR with competitive inhibitors could be a promising strategy to inhibit QS in P. aeruginosa to overcome antimicrobial resistance. In this study, we have designed and synthesized a series of novel quinazolinone disulfide-containing competitive inhibitor of PqsR. The most potent analogue 8q efficiently inhibited the pqs system with an IC50 value of 4.5 µM. It also showed complete suppression of pyocyanin production and a significant reduction in biofilm formation by P. aeruginosa (PAO1) with low cytotoxicity. Additionally, 8q produced synergy in combination with known antibiotics such as ciprofloxacin and tobramycin. Finally, molecular docking analysis suggested that compound 8q could bind with the ligand-binding domain of PqsR in a similar fashion to the native ligand.

The Efficacy of an N-Acetylcysteine-Antibiotic Combination Therapy on Achromobacter xylosoxidans in a Cystic Fibrosis Sputum/Lung Cell Model.

Cystic fibrosis (CF) is a disorder causing dysfunctional ion transport resulting in the accumulation of viscous mucus. This environment fosters a chronic bacterial biofilm-associated infection in the airways. Achromobacter xylosoxidans, a gram-negative aerobic bacillus, has been increasingly associated with antibiotic resistance and chronic colonisation in CF. In this study, we aimed to create a reproducible model of CF infection using an artificial sputum medium (ASMDM-1) with bronchial (BEAS-2B) and macrophage (THP-1) cells to test A. xylosoxidans infection and treatment toxicity. This study was conducted in three distinct stages. First, the tolerance of BEAS-2B cell lines and two A. xylosoxidans strains against ASMDM-1 was optimised. Secondly, the cytotoxicity of combined therapy (CT) comprising N-acetylcysteine (NAC) and the antibiotics colistin or ciprofloxacin was tested on cells alone in the sputum model in both BEAS-2B and THP-1 cells. Third, the efficacy of CT was assessed in the context of a bacterial infection within the live cell/sputum model. We found that a model using 20% ASMDM-1 in both cell populations tolerated a colistin-NAC-based CT and could significantly reduce bacterial loads in vitro (~2 log10 CFU/mL compared to untreated controls). This pilot study provides the foundation to study other bacterial opportunists that infect the CF lung to observe infection and CT kinetics. This model also acts as a springboard for more complex co-culture models.

Editorial for Special Issue "Antibacterial Resistance and Novel Strategies to Eradicate Bacterial Biofilms".

Bacterial resistance to antimicrobial agents, including antibiotics, disinfectants, and detergents, is on the rise, with consequences associated with morbidity, mortality, and economic loss in the healthcare sector [...].

Antimicrobial and Anti-inflammatory Gallium-Defensin Surface Coatings for Implantable Devices.

Emerging and re-emerging infections are a global threat driven by the development of antimicrobial resistance due to overuse of antimicrobial agents and poor infection control practices. Implantable devices are particularly susceptible to such infections due to the formation of microbial biofilms. Furthermore, the introduction of implants into the body often results in inflammation and foreign body reactions. The antimicrobial and anti-inflammatory properties of gallium (Ga) have been recognized but not yet utilized effectively to improve implantable device integration. Furthermore, defensin (De, hBD-1) has potent antimicrobial activity in vivo as part of the innate immune system; however, this has not been demonstrated as successfully when used in vitro. Here, we combined Ga and De to impart antimicrobial activity and anti-inflammatory properties to polymer-based implantable devices. We fabricated polylactic acid films, which were modified using Ga implantation and subsequently functionalized with De. Ga-ion implantation increased surface roughness and increased stiffness. Ga implantation and defensin immobilization both independently and synergistically introduced antimicrobial activity to the surfaces, significantly reducing total live bacterial biomass. We demonstrated, for the first time, that the antimicrobial effects of De were unlocked by its surface immobilization. Ga implantation of the surface also resulted in reduced foreign body giant cell formation and expression of proinflammatory cytokine IL-1ß. Cumulatively, the treated surfaces were able to kill bacteria and reduce inflammation in comparison to the untreated control. These innovative surfaces have the potential to prevent biofilm formation without inducing cellular toxicity or inflammation, which is highly desired for implantable device integration.

Halogenated Dihydropyrrol-2-One Molecules Inhibit Pyocyanin Biosynthesis by Blocking the Pseudomonas Quinolone Signaling System.

Quorum-sensing (QS) systems of Pseudomonas aeruginosa are involved in the control of biofilm formation and virulence factor production. The current study evaluated the ability of halogenated dihydropyrrol-2-ones (DHP) (Br (4a), Cl (4b), and F (4c)) and a non-halogenated version (4d) to inhibit the QS receptor proteins LasR and PqsR. The DHP molecules exhibited concentration-dependent inhibition of LasR and PqsR receptor proteins. For LasR, all compounds showed similar inhibition levels. However, compound 4a (Br) showed the highest decrease (two-fold) for PqsR, even at the lowest concentration (12.5 µg/mL). Inhibition of QS decreased pyocyanin production amongst P. aeruginosa PAO1, MH602, ATCC 25619, and two clinical isolates (DFU-53 and 364707). In the presence of DHP, P. aeruginosa ATCC 25619 showed the highest decrease in pyocyanin production, whereas clinical isolate DFU-53 showed the lowest decrease. All three halogenated DHPs also reduced biofilm formation by between 31 and 34%. The non-halogenated compound 4d exhibited complete inhibition of LasR and had some inhibition of PqsR, pyocyanin, and biofilm formation, but comparatively less than halogenated DHPs.

Effect of N-Acetylcysteine in Combination with Antibiotics on the Biofilms of Three Cystic Fibrosis Pathogens of Emerging Importance.

Cystic fibrosis (CF) is a genetic disorder causing dysfunctional ion transport resulting in accumulation of viscous mucus that fosters chronic bacterial biofilm-associated infection in the airways. Achromobacter xylosoxidans and Stenotrophomonas maltophilia are increasingly prevalent CF pathogens and while Burkholderia cencocepacia is slowly decreasing; all are complicated by multidrug resistance that is enhanced by biofilm formation. This study investigates potential synergy between the antibiotics ciprofloxacin (0.5-128 µg/mL), colistin (0.5-128 µg/mL) and tobramycin (0.5-128 µg/mL) when combined with the neutral pH form of N-Acetylcysteine (NACneutral) (0.5-16.3 mg/mL) against 11 cystic fibrosis strains of Burkholderia, Stenotrophomonas and Achromobacter sp. in planktonic and biofilm cultures. We screened for potential synergism using checkerboard assays from which fraction inhibitory concentration indices (FICI) were calculated. Synergistic (FICI ≤ 0.5) and additive (0.5 > FICI ≥ 1) combinations were tested on irreversibly attached bacteria and 48 h mature biofilms via time-course and colony forming units (CFU/mL) assays. This study suggests that planktonic FICI analysis does not necessarily translate to reduction in bacterial loads in a biofilm model. Future directions include refining synergy testing and determining further mechanisms of action of NAC to understand how it may interact with antibiotics to better predict synergy.

N-Acetylcysteine Protects Bladder Epithelial Cells from Bacterial Invasion and Displays Antibiofilm Activity against Urinary Tract Bacterial Pathogens.

Introduction: Urinary tract infections (UTIs) affect more than 150 million individuals annually. A strong correlation exists between bladder epithelia invasion by uropathogenic bacteria and patients with recurrent UTIs. Intracellular bacteria often recolonise epithelial cells post-antibiotic treatment. We investigated whether N-acetylcysteine (NAC) could prevent uropathogenic E. coli and E. faecalis bladder cell invasion, in addition to its effect on uropathogens when used alone or in combination with ciprofloxacin. Methods: An invasion assay was performed in which bacteria were added to bladder epithelial cells (BECs) in presence of NAC and invasion was allowed to occur. Cells were washed with gentamicin, lysed, and plated for enumeration of the intracellular bacterial load. Cytotoxicity was evaluated by exposing BECs to various concentrations of NAC and quantifying the metabolic activity using resazurin at different exposure times. The effect of NAC on the preformed biofilms was also investigated by treating 48 h biofilms for 24 h and enumerating colony counts. Bacteria were stained with propidium iodide (PI) to measure membrane damage. Results: NAC completely inhibited BEC invasion by multiple E. coli and E. faecalis clinical strains in a dose-dependent manner (p < 0.01). This was also evident when bacterial invasion was visualised using GFP-tagged E. coli. NAC displayed no cytotoxicity against BECs despite its intrinsic acidity (pH ~2.6), with >90% cellular viability 48 h post-exposure. NAC also prevented biofilm formation by E. coli and E. faecalis and significantly reduced bacterial loads in 48 h biofilms when combined with ciprofloxacin. NAC visibly damaged E. coli and E. faecalis bacterial membranes, with a threefold increase in propidium iodide-stained cells following treatment (p < 0.05). Conclusions: NAC is a non-toxic, antibiofilm agent in vitro and can prevent cell invasion and IBC formation by uropathogens, thus providing a potentially novel and efficacious treatment for UTIs. When combined with an antibiotic, it may disrupt bacterial biofilms and eliminate residual bacteria.

Natural Product Rottlerin Derivatives Targeting Quorum Sensing.

Rottlerin is a natural product consisting of chalcone and flavonoid scaffolds, both of which have previously shown quorum sensing (QS) inhibition in various bacteria. Therefore, the unique rottlerin scaffold highlights great potential in inhibiting the QS system of Pseudomonas aeruginosa. Rottlerin analogues were synthesised by modifications at its chalcone- and methylene-bridged acetophenone moieties. The synthesis of analogues was achieved using an established five-step synthetic strategy for chalcone derivatives and utilising the Mannich reaction at C6 of the chromene to construct morpholine analogues. Several pyranochromene chalcone derivatives were also generated using aldol conditions. All the synthetic rottlerin derivatives were screened for QS inhibition and growth inhibition against the related LasR QS system. The pyranochromene chalcone structures displayed high QS inhibitory activity with the most potent compounds, 8b and 8d, achieving QS inhibition of 49.4% and 40.6% and no effect on bacterial growth inhibition at 31 µM, respectively. Both compounds also displayed moderate biofilm inhibitory activity and reduced the production of pyocyanin.

Disruption of biofilms and killing of Burkholderia cenocepacia from cystic fibrosis lung using an antioxidant-antibiotic combination therapy.

Cystic fibrosis (CF) is a disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR). The resulting chloride and bicarbonate imbalance produces a thick, static lung mucus. This mucus is not easily expelled from the lung and can be colonised by bacteria, leading to biofilm formation. CF lung infection with Burkholderia cepacia complex (BCC), particularly the subspecies B. cenocepacia, results in higher morbidity and mortality. Patients infected with BCC can rapidly progress to "cepacia syndrome", a fatal necrotising pneumonia. The aim of this study was to identify whether a combination therapy (CT) of selected antioxidants and antibiotics significantly disrupts B. cenocepacia biofilms and to determine the optimum CT level for treatment. Using controlled in vitro spectrophotometry, colony-forming unit and microscopy assays, three antioxidants (N-acetylcysteine [NAC], glutathione and vitamin C) and three antibiotics (ciprofloxacin, ceftazidime and tobramycin) were screened and assessed for their ability to disrupt the early and mature biofilms of six B. cenocepacia CF isolates. A combination of NAC and ciprofloxacin produced a statistically significant biofilm disruption in all strains tested, with growth inhibition (>5-8 log10) observed when exposed to 4890 or 8150 µg/mL NAC in combination with 32 or 64 µg/mL ciprofloxacin. NAC-mediated biofilm disruption may be aided by the acidic pH of NAC at higher concentrations. This study showed that NAC is an effective disruptor that reduces the necessity for high concentrations of antibiotic. Further research will focus on the host toxicity and efficacy in ex vivo CF models.

Novel Seleno- and Thio-Urea Containing Dihydropyrrol-2-One Analogues as Antibacterial Agents.

The quorum sensing (QS) system in multi-drug-resistant bacteria such as P. aeruginosa is primarily responsible for the development of antibiotic resistance and is considered an attractive target for antimicrobial drug discovery. In this study, we synthesised a series of novel selenourea and thiourea-containing dihydropyrrol-2-one (DHP) analogues as LasR antagonists. The selenium DHP derivatives displayed significantly better quorum-sensing inhibition (QSI) activities than the corresponding sulphur analogues. The most potent analogue 3e efficiently inhibited the las QS system by 81% at 125 µM and 53% at 31 µM. Additionally, all the compounds were screened for their minimum inhibitory concentration (MIC) against the Gram-positive bacterium S. aureus, and interestingly, only the selenium analogues showed antibacterial activity, with 3c and 3e being the most potent with a MIC of 15.6 µM.

Thioether-linked dihydropyrrol-2-one analogues as PqsR antagonists against antibiotic resistant Pseudomonas aeruginosa.

The Pseudomonas quinolone system (pqs) is one of the key quorum sensing systems in antibiotic-resistant P. aeruginosa and is responsible for the production of virulence factors and biofilm formation. Thus, synthetic small molecules that can target the PqsR (MvfR) receptor can be utilized as quorum sensing inhibitors to treat P. aeruginosa infections. In this study, we report the synthesis of novel thioether-linked dihydropyrrol-2-one (DHP) analogues as PqsR antagonists. Compound 7g containing a 2-mercaptopyridyl linkage effectively inhibited the pqs system with an IC50 of 32 µM in P. aeruginosa PAO1. Additionally, these inhibitors significantly reduced bacterial aggregation and biofilm formation without affecting planktonic growth. The molecular docking study suggest that these inhibitors bind with the ligand binding domain of the MvfR as a competitive antagonist.

Covalent Immobilization of N-Acetylcysteine on a Polyvinyl Chloride Substrate Prevents Bacterial Adhesion and Biofilm Formation.

Biofilm formation and antimicrobial resistance at surgical implant sites result in high morbidity and mortality. Identifying novel molecules that inhibit biofilm formation to coat surgical biomaterials is essential. One such compound is N-acetylcysteine (NAC), a potent antioxidant precursor for glutathione, necessary in mammalian cells and known to disrupt/prevent biofilms. In this study, NAC was covalently immobilized onto functionalized polyvinyl chloride surfaces using plasma immersion ion implantation (PIII) treatment that achieves covalent binding without the need for linker groups. NAC immobilization was characterized using water contact angles, Fourier-transform infrared, and X-ray photoelectron spectroscopy techniques. Bacterial viability and biofilm formation on NAC surfaces were assessed using resazurin assays, phase contrast microscopy, and colony counting experiments. Effect of NAC on bacterial polysaccharide production and DNA cleaving was investigated using the phenol-sulfuric acid method and the Qubit fluorometer. Surface thermodynamics between the NAC coating and bacterial cells were measured using the Lewis acid-base method. Surface characterization techniques demonstrated superficial changes after PIII treatment and subsequent covalent NAC immobilization. NAC-coated surfaces significantly reduced biofilm viability and the presence of Gram-negative and Gram-positive bacteria. NAC also decreased polysaccharide production and degraded DNA. This led to unfavorable conditions for biofilm formation on NAC-coated surfaces, as demonstrated by surface thermodynamic analysis. NAC-coated surfaces showed no cytotoxicity to human fibroblast cells. This study has successfully utilized NAC as an antibiofilm coating, which may pave the way for improved prophylactic coatings on medical implant devices in the future.

Design, Synthesis and Biological Evaluation of Novel Anthraniloyl-AMP Mimics as PQS Biosynthesis Inhibitors Against Pseudomonas aeruginosa Resistance.

The Pseudomonas quinolone system (PQS) is one of the three major interconnected quorum sensing signaling systems in Pseudomonas aeruginosa. The virulence factors PQS and HHQ activate the transcription regulator PqsR (MvfR), which controls several activities in bacteria, including biofilm formation and upregulation of PQS biosynthesis. The enzyme anthraniloyl-CoA synthetase (PqsA) catalyzes the first and critical step in the biosynthesis of quinolones; therefore, it is an attractive target for the development of anti-virulence therapeutics against Pseudomonas resistance. Herein, we report the design and synthesis of novel triazole nucleoside-based anthraniloyl- adenosine monophosphate (AMP) mimics. These analogues had a major impact on the morphology of bacterial biofilms and caused significant reduction in bacterial aggregation and population density. However, the compounds showed only limited inhibition of PQS and did not exhibit any effect on pyocyanin production.

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

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.

Conditions Under Which Glutathione Disrupts the Biofilms and Improves Antibiotic Efficacy of Both ESKAPE and Non-ESKAPE Species.

Bacterial antibiotic resistance has increased in recent decades, raising concerns in hospital and community settings. Novel, innovative strategies are needed to eradicate bacteria, particularly within biofilms, and diminish the likelihood of recurrence. In this study, we investigated whether glutathione (GSH) can act as a biofilm disruptor, and enhance antibiotic effectiveness against various bacterial pathogens. Biological levels (10 mM) of GSH did not have a significant effect in inhibiting growth or disrupting the biofilm in four out of six species tested. However, exposure to 30 mM GSH showed >50% decrease in growth for all bacterial species, with almost 100% inhibition of Streptococcus pyogenes and an average of 94-52% inhibition for Escherichia coli, Methicillin-resistant Staphylococcus aureus (MRSA) and Methicillin-sensitive S. aureus (MSSA) and multi-drug resistant Acinetobacter baumannii (MRAB) isolates, respectively. Klebsiella pneumoniae and Enterobacter sp. isolates were however, highly resistant to 30 mM GSH. With respect to biofilm viability, all species exhibited a >50% decrease in viability with 30 mM GSH, with confocal imaging showing considerable change in the biofilm architecture of MRAB isolates. The mechanism of GSH-mediated biofilm disruption is possibly due to a concentration-dependent increase in GSH acidity that triggers cleaving of the matrix components. Enzymatic treatment of MRAB revealed that eDNA and polysaccharides are essential for biofilm stability and eDNA removal enhanced amikacin efficiency. Combination of GSH, amikacin and DNase-I showed the greatest reduction in MRAB biofilm viability. Additionally, GSH alone and in combination with amikacin fostered human fibroblast cell (HFF-1) growth and confluence while inhibiting MRAB adhesion and colonization.

Bacteriophage PEV20 and Ciprofloxacin Combination Treatment Enhances Removal of Pseudomonas aeruginosa Biofilm Isolated from Cystic Fibrosis and Wound Patients.

Antibiotic resistance in Pseudomonas aeruginosa biofilms necessitates the need for novel antimicrobial therapy with anti-biofilm properties. Bacteriophages (phages) are recognized as an ideal biopharmaceutical for combating antibiotic-resistant bacteria especially when used in combination with antibiotics. However, previous studies primarily focused on using phages against of P. aeruginosa biofilms of laboratory strains. In the present study, biofilms of six P. aeruginosa isolated from cystic fibrosis and wound patients, and one laboratory strain was treated singly and with combinations of anti-Pseudomonas phage PEV20 and ciprofloxacin. Of these strains, three were highly susceptible to the phage, while one was partially resistant and one was completely resistant. Combination treatment with PEV20 and ciprofloxacin enhanced biofilm eradication compared with single treatment. Phage and ciprofloxacin synergy was found to depend on phage-resistance profile of the target bacteria. Furthermore, phage and ciprofloxacin combination formulation protected the lung epithelial and fibroblast cells from P. aeruginosa and promoted cell growth. The results demonstrated that thorough screening of phage-resistance is crucial for designing phage-antibiotic formulation. The addition of highly effective phage could reduce the ciprofloxacin concentration required to combat P. aeruginosa infections associated with biofilm in cystic fibrosis and wound patients.

Spray-Dried Particles of Nitric Oxide-Modified Glutathione for the Treatment of Chronic Lung Infection.

Antibiotic resistance in pathogenic bacteria has emerged as a big challenge to human and animal health and significant economy loss worldwide. Development of novel strategies to tackle antibiotic resistance is of the utmost priority. In this study, we combined glutathione (GSH), a master antioxidant in all mammalian cells, and nitric oxide, a proven biofilm-dispersing agent, to produce GSNO. The resazurin biofilm viability assay, crystal violet biofilm assay, and confocal microscopy techniques showed that GSNO disrupted biofilms of both P. aeruginosa PAO1 and multidrug resistant A. baumaunii (MRAB 015069) more efficiently than GSH alone. In addition, GSNO showed a higher reduction in biofilm viability and biomass when combined with antibiotics. This combination treatment also inhibited A. baumaunii (MRAB 015069) growth and facilitated human foreskin fibroblast (HFF-1) confluence and growth simultaneously. A potentially inhalable GSNO powder with reasonable aerosol performance and antibiofilm activity was produced by spray drying. This combination shows promise as a novel formulation for treating pulmonary bacterial infections.