Browsing by Author "Stoodley, P."

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    Cephalosporin-3’-diazeniumdiolate NO-donor prodrug PYRRO-C3D is active against Streptococcus pneumoniae biofilms despite the absence of -lactamases
    (Elsevier, 2017-02-21) Allan, Raymond N.; Kelso, M.J.; Rineh, A.; Yepuri, N.R.; Feelisch, M.; Soren, O.; Brito-Mutunayagam, S.; Salib, R.J.; Stoodley, P.; Clarke, S.C.; Webb, J.S.; Hall-Stoodley, L.; Faust, S.N.
    Bacterial biofilms show high tolerance towards antibiotics and are a significant problem in clinical settings where they are a primary cause of chronic infections. Novel therapeutic strategies are needed to improve anti-biofilm efficacy and support reduction in antibiotic use. Treatment with exogenous nitric oxide (NO) has been shown to modulate bacterial signaling and metabolic processes that render biofilms more susceptible to antibiotics.We previously reported on cephalosporin-3’-diazeniumdiolates (C3Ds) as NO-donor prodrugs designed to selectively deliver NO to bacterial infection sites following reaction with b-lactamases. With structures based on cephalosporins, C3Ds could, in principal, also be triggered to release NO following b-lactam cleavage mediated by transpeptidases/penicillin-binding proteins (PBPs), the antibacterial target of cephalosporin antibiotics. Transpeptidase-reactive C3Ds could potentially show both NO-mediated anti-biofilm properties and intrinsic (b-lactam-mediated) antibacterial effects. This dual-activity concept was explored using Streptococcus pneumoniae, a species that lacks b-lactamases but relies on transpeptidases for cell-wall synthesis. Treatment with PYRRO-C3D (a representative C3D containing the diazeniumdiolate NO donor PYRRO-NO) was found to significantly reduce viability of planktonic and biofilm pneumococci, demonstrating that C3Ds can elicit direct, cephalosporin-like antibacterial activity in the absence of b-lactamases. While NO release from PYRRO-C3D in the presence of pneumococci was confirmed, the anti-pneumococcal action of the compound was shown to arise exclusively from the b-lactam component and not through NO-mediated effects. The compound showed similar potency to amoxicillin against S. pneumoniae biofilms and greater efficacy than azithromycin, highlighting the potential of C3Ds as new agents for treating pneumococcal infections.
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    D-methionine interferes with non-typeable Haemophilus influenzae peptidoglycan synthesis during growth and biofilm formation
    (Microbiology Society, 2017-07-12) Dawe, H.; Berger, E.; Sihlbom, C.; Angus, E. M.; Howlin, R. P.; Laver, J. R.; Tebruegge, M.; Hall-Stoodley, L.; Stoodley, P.; Faust, S. N.; Allan, Raymond N.
    Non-typeable Haemophilus influenzae (NTHi) is an opportunistic pathogen that plays a major role in a number of respiratory tract infections, including otitis media, cystic fibrosis and chronic obstructive pulmonary disease. Biofilm formation has been implicated in both NTHi colonization and disease, and is responsible for the increased tolerance of this pathogen towards antibiotic treatment. Targeting metabolic pathways that are important in NTHi biofilm formation represents a potential strategy to combat this antibiotic recalcitrance. A previous investigation demonstrated increased expression of a putative D-methionine uptake protein following exposure of NTHi biofilms to the ubiquitous signalling molecule, nitric oxide. We therefore hypothesized that treatment with exogenous D-methionine would impact on NTHi biofilm formation and increase antibiotic sensitivity. Treatment of NTHi during the process of biofilm formation resulted in a reduction in biofilm viability, increased biomass, changes in the overall biofilm architecture and the adoption of an amorphous cellular morphology. Quantitative proteomic analyses identified 124 proteins that were differentially expressed following D-methionine treatment, of which 51 (41 %) were involved in metabolic and transport processes. Nine proteins involved in peptidoglycan synthesis and cell division showed significantly increased expression. Furthermore, D-methionine treatment augmented the efficacy of azithromycin treatment and highlighted the potential of D-methionine as an adjunctive therapeutic approach for NTHi biofilm-associated infections.
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    Low concentrations of nitric oxide modulate Streptococcus pneumoniae biofilm metabolism and antibiotic tolerance
    (American Society for Microbiology, 2016-03-25) Allan, Raymond N.; Morgan, S.; Brito-Mutunayagam, S.; Skipp, P.; Feelisch, M.; Hayes, S. M.; Hellier, W.; Clarke, S. C.; Stoodley, P.; Burgess, A.; Ismail-Koch, H.; Salib, R. J.; Webb, J. S.; Faust, S. N.; Hall-Stoodley, L.
    Streptococcus pneumoniae is one of the key pathogens responsible for otitis media (OM), the most common infection in children and the largest cause of childhood antibiotic prescription. Novel therapeutic strategies that reduce the overall antibiotic consumption due to OM are required because, although widespread pneumococcal conjugate immunization has controlled invasive pneumococcal disease, overall OM incidence has not decreased. Biofilm formation represents an important phenotype contributing to the antibiotic tolerance and persistence of S. pneumoniae in chronic or recurrent OM. We investigated the treatment of pneumococcal biofilms with nitric oxide (NO), an endogenous signaling molecule and therapeutic agent that has been demonstrated to trigger biofilm dispersal in other bacterial species. We hypothesized that addition of low concentrations of NO to pneumococcal biofilms would improve antibiotic efficacy and that higher concentrations exert direct antibacterial effects. Unlike in many other bacterial species, low concentrations of NO did not result in S. pneumoniae biofilm dispersal. Instead, treatment of both in vitro biofilms and ex vivo adenoid tissue samples (a reservoir for S. pneumoniae biofilms) with low concentrations of NO enhanced pneumococcal killing when combined with amoxicillin-clavulanic acid, an antibiotic commonly used to treat chronic OM. Quantitative proteomic analysis using iTRAQ (isobaric tag for relative and absolute quantitation) identified 13 proteins that were differentially expressed following low-concentration NO treatment, 85% of which function in metabolism or translation. Treatment with low-concentration NO, therefore, appears to modulate pneumococcal metabolism and may represent a novel therapeutic approach to reduce antibiotic tolerance in pneumococcal biofilms.
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    Low-Dose Nitric Oxide as Targeted Anti-biofilm Adjunctive Therapy to Treat Chronic Pseudomonas aeruginosa Infection in Cystic Fibrosis
    (Elsevier, 2017-07-24) Howlin, R.; Cathie, K.; Hall-Stoodley, L.; Cornelius, V.; Duignan, C.; Allan, Raymond N.; Fernandez, B.O.; Barraud, N.; Bruce, K.D.; Jefferies, J.; Kelso, M.; Kjelleberg, S.; Rice, S.A.; Rogers, G.B.; Pink, S.; Smith, C.; Sukhtankar, P.S.; Salib, R.; Legg, J.; Carroll, M.; Daniels, T.; Feelisch, M.; Stoodley, P.; Clarke, S.C.; Connett, G.; Faust, S.N.; Webb, J.S.
    Despite aggressive antibiotic therapy, bronchopulmonary colonization by Pseudomonas aeruginosa causes persistent morbidity and mortality in cystic fibrosis (CF). Chronic P. aeruginosa infection in the CF lung is associated with structured, antibiotic-tolerant bacterial aggregates known as biofilms. We have demonstrated the effects of non-bactericidal, low-dose nitric oxide (NO), a signaling molecule that induces biofilm dispersal, as a novel adjunctive therapy for P. aeruginosa biofilm infection in CF in an ex vivo model and a proof-of-concept double-blind clinical trial. Submicromolar NO concentrations alone caused disruption of biofilms within ex vivo CF sputum and a statistically significant decrease in ex vivo biofilm tolerance to tobramycin and tobramycin combined with ceftazidime. In the 12-patient randomized clinical trial, 10 ppm NO inhalation caused significant reduction in P. aeruginosa biofilm aggregates compared with placebo across 7 days of treatment. Our results suggest a benefit of using lowdose NO as adjunctive therapy to enhance the efficacy of antibiotics used to treat acute P. aeruginosa exacerbations in CF. Strategies to induce the disruption of biofilms have the potential to overcome biofilm-associated antibiotic tolerance in CF and other biofilm-related diseases.
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    Targeting microbial biofilms: current and prospective therapeutic strategies
    (Macmillan Publishers Limited, 2017-09-25) Koo, H.; Allan, Raymond N.; Howlin, R. P.; Stoodley, P.; Hall-Stoodley, L.
    Biofilm formation is now recognized as a key virulence factor for a wide range of chronic microbial infections. While it has been well known for decades that bacteria and fungi in biofilms become highly tolerant of antibiotics, the development of effective therapeutics has lagged behind our growing understanding of biofilm biology. The multifactorial nature of biofilm development and drug tolerance imposes significant challenges to conventional antimicrobials, and indicates the need for multi-targeted or combinatorial therapies. In light of the discrepancy between the explosion of papers presenting multitude of methods to control biofilms and the sparsity of biofilm specific treatments available to the clinician, in this review, we focus on current therapeutic strategies and those in development for the treatment of biofilm infections, which target vital structure-function traits and drug tolerance mechanisms, including the extracellular matrix and dormant cells. We emphasize strategies that are supported by in vivo or ex vivo studies, highlight emerging anti-biofilm technologies, and provide a rationale for multi-targeted therapies aimed at disrupting the complex biofilm microenvironment.