Repurposing mitomycin C in combination with pentamidine or gentamicin to treat infections with multi-drug resistant (MDR) 3 Pseudomonas aeruginosa

The aims of this study were (i) to determine if the combination of mitomycin C with pentamidine or existing antibiotics resulted in enhanced efficacy versus infections with MDR P. aeruginosa in vivo; and (ii) to determine if the doses of mitomycin C and pentamidine in combination can be reduced to levels that are non-toxic in humans but still retain antibacterial activity. Resistant clinical isolates of P. aeruginosa, a mutant strain over-expressing the MexAB-OprM resistance nodulation division (RND) efflux pump and a strain with three RND pumps deleted, were used. MIC assays indicated that all strains were sensitive to mitomycin C, but deletion of three RND pumps resulted in hypersensitivity and over-expression of MexAB-OprM caused some resistance. These results imply that mitomycin C is a substrate of the RND efflux pumps. Mitomycin C monotherapy successfully treated infected Galleria mellonella larvae, albeit at doses too high for human administration. Checkerboard and time–kill assays showed that the combination of mitomycin C with pentamidine, or the antibiotic gentamicin, resulted in synergistic inhibition of most P. aeruginosa strains in vitro. In vivo, administration of a combination therapy of mitomycin C with pentamidine, or gentamicin, to G. mellonella larvae infected with P. aeruginosa resulted in enhanced efficacy compared with monotherapies for the majority of MDR clinical isolates. Notably, the therapeutic benefit conferred by the combination therapy occurred with doses of mitomycin C close to those used in human medicine. Thus, repurposing mitomycin C in combination therapies to target MDR P. aeruginosa infections merits further investigation.Peer reviewe

Modelling the transmission of infectious diseases inside hospital bays:implications for COVID-19

Healthcare associated transmission of viral infections is a major problem that has significant economic costs and can lead to loss of life. Infections with the highly contagious SARS-CoV-2 virus have been shown to have a high prevalence in hospitals around the world. The spread of this virus might be impacted by the density of patients inside hospital bays. To investigate this aspect, in this study we consider a mathematical modelling and computational approach to describe the spread of SARSCoV-2 among hospitalised patients. We focus on 4-bed bays and 6-bed bays, which are commonly used to accommodate various non-COVID-19 patients in many hospitals across the United Kingdom (UK). We investigate the spread of SARS-CoV-2 infections among patients in non-COVID bays, in the context of various scenarios: placing the initially-exposed individual in different beds, varying the recovery and incubation periods, having symptomatic vs. asymptomatic patients, removing infected individuals from these hospital bays once they are known to be infected, and the role of periodic testing of hospitalised patients. Our results show that 4-bed bays reduce the spread of SARS-CoV-2 compared to 6-bed bays. Moreover, we show that the position of a new (not infected) patient in specific beds in a 6-bed bay might also slow the spread of the disease. Finally, we propose that regular SARS-CoV-2 testing of hospitalised patients would allow appropriate placement of infected patients in specific (COVID-only) hospital bays.</p

Repurposing the anti-viral drug zidovudine (AZT) in combination with meropenem as an effective treatment for infections with multi-drug resistant, carbapenemase-producing strains of Klebsiella pneumoniae

Funding: University of St Andrews.Multi-drug resistant (MDR) Klebsiella pneumoniae represent a global threat to healthcare due to lack of effective treatments and high mortality rates. The aim of this research was to explore the potential of administering zidovudine (AZT) in combination with an existing antibiotic to treat resistant K. pneumoniae infections. Two MDR K. pneumoniae strains were employed, producing either the NDM-1 or KPC-3 carbapenemase. Efficacy of combinations of AZT with meropenem were compared with monotherapies against infections in Galleria mellonella larvae by measuring larval mortality and bacterial burden. The effect of the same combinations in vitro was determined via checkerboard and time-kill assays. In vitro, both K. pneumoniae strains were resistant to meropenem but were susceptible to AZT. In G. mellonella, treatment with either AZT or meropenem alone offered minimal therapeutic benefit against infections with either strain. In contrast, combination therapy of AZT with meropenem presented significantly enhanced efficacy compared to monotherapies. This was correlated with prevention of bacterial proliferation within the larvae but not elimination. Checkerboard assays showed that the interaction between AZT and meropenem was not synergistic but indifferent. In summary, combination therapy of AZT with meropenem represents a potential treatment for carbapenemase-producing MDR K. pneumoniae and merits further investigation.PostprintPeer reviewe

Rapid determination of antimicrobial susceptibility of Gram-negative bacteria from clinical blood cultures using a scattered light integrated collection (SLIC) device

Funding: This work was funded by the University of St Andrews.Background A bloodstream infection (BSI) presents a complex and serious health problem, a problem that is being exacer- bated by increasing antimicrobial resistance (AMR). Gap statement The current turnaround times (TATs) for most antimicrobial susceptibility testing (AST) methods offer results retrospective of treatment decisions, and this limits the impact AST can have on antibiotic prescribing and patient care. Progress must be made towards rapid BSI diagnosis and AST to improve antimicrobial stewardship and reduce preventable deaths from BSIs. To support the successful implementation of rapid AST (rAST) in hospital settings, a rAST method that is affordable, is sustainable and offers comprehensive AMR detection is needed. Aim To evaluate a scattered light-integrated collection (SLIC) device against standard of care (SOC) to determine whether SLIC could accelerate the current TATs with actionable, accurate rAST results for Gram-negative BSIs. Methods Positive blood cultures from a tertiary referral hospital were studied prospectively. Flagged positive Gram-negative blood cultures were confirmed by Gram staining and analysed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, Vitek 2, disc diffusion (ceftriaxone susceptibility only) and an SLIC device. Susceptibility to a panel of five antibiotics, as defined by European Committee on Antimicrobial Susceptibility Testing breakpoints, was examined using SLIC. Results A total of 505 bacterial–antimicrobial combinations were analysed. A categorical agreement of 95.5 % (482/505) was achieved between SLIC and SOC. The 23 discrepancies that occurred were further investigated by the broth microdilution method, with 10 AST results in agreement with SLIC and 13 in agreement with SOC. The mean time for AST was 10.53±0.46 h and 1.94±0.02 h for Vitek 2 and SLIC, respectively. SLIC saved 23.96±1.47 h from positive blood culture to AST result. Conclusion SLIC has the capacity to provide accurate AST 1 day earlier from flagged positive blood cultures than SOC. This significant time saving could accelerate time to optimal antimicrobial therapy, improving antimicrobial stewardship and man- agement of BSIs.Peer reviewe

Rapid determination of antimicrobial susceptibility of Gram-negative bacteria from clinical blood cultures using a scattered light-integrated collection device

Background. A bloodstream infection (BSI) presents a complex and serious health problem, a problem that is being exacerbated by increasing antimicrobial resistance (AMR). Gap Statement. The current turnaround times (TATs) for most antimicrobial susceptibility testing (AST) methods offer results retrospective of treatment decisions, and this limits the impact AST can have on antibiotic prescribing and patient care. Progress must be made towards rapid BSI diagnosis and AST to improve antimicrobial stewardship and reduce preventable deaths from BSIs. To support the successful implementation of rapid AST (rAST) in hospital settings, a rAST method that is affordable, is sustainable and offers comprehensive AMR detection is needed. Aim. To evaluate a scattered light-integrated collection (SLIC) device against standard of care (SOC) to determine whether SLIC could accelerate the current TATs with actionable, accurate rAST results for Gram-negative BSIs. Methods. Positive blood cultures from a tertiary referral hospital were studied prospectively. Flagged positive Gram-negative blood cultures were confirmed by Gram staining and analysed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, Vitek 2, disc diffusion (ceftriaxone susceptibility only) and an SLIC device. Susceptibility to a panel of five antibiotics, as defined by European Committee on Antimicrobial Susceptibility Testing breakpoints, was examined using SLIC. Results. A total of 505 bacterial-antimicrobial combinations were analysed. A categorical agreement of 95.5% (482/505) was achieved between SLIC and SOC. The 23 discrepancies that occurred were further investigated by the broth microdilution method, with 10 AST results in agreement with SLIC and 13 in agreement with SOC. The mean time for AST was 10.53±0.46 h and 1.94±0.02 h for Vitek 2 and SLIC, respectively. SLIC saved 23.96±1.47 h from positive blood culture to AST result. Conclusion. SLIC has the capacity to provide accurate AST 1 day earlier from flagged positive blood cultures than SOC. This significant time saving could accelerate time to optimal antimicrobial therapy, improving antimicrobial stewardship and management of BSIs.</p

Combination therapy with ciprofloxacin and pentamidine against Multidrug-Resistant Pseudomonas aeruginosa : assessment of in vitro and in vivo efficacy and the role of Resistance-Nodulation-Division (RND) efflux pumps

Funding: This research was funded by the University of St Andrews.The aim of this work was to (i) evaluate the efficacy of a combination treatment of pentamidine with ciprofloxacin against Galleria mellonella larvae infected with an MDR strain of P. aeruginosa and (ii) determine if pentamidine acts as an efflux-pump inhibitor. Resistant clinical isolates, mutant strains overexpressing one of three RND efflux pumps (MexAB-OprM, MexCD-OprJ, and MexEF-OprN), and a strain with the same three pumps deleted were used. MIC assays confirmed that the clinical isolates and the mutants overexpressing efflux pumps were resistant to ciprofloxacin and pentamidine. The deletion of the three efflux pumps induced sensitivity to both compounds. Exposure to pentamidine and ciprofloxacin in combination resulted in the synergistic inhibition of all resistant strains in vitro, but no synergy was observed versus the efflux-pump deletion strain. The treatment of infected G. mellonella larvae with the combination of pentamidine and ciprofloxacin resulted in enhanced efficacy compared with the monotherapies and significantly reduced the number of proliferating bacteria. Our measurement of efflux activity from cells revealed that pentamidine had a specific inhibitory effect on the MexCD-OprJ and MexEF-OprN efflux pumps. However, the efflux activity and membrane permeability assays revealed that pentamidine also disrupted the membrane of all cells. In conclusion, pentamidine does possess some efflux-pump inhibitory activity, in addition to a more general disruptive effect on membrane integrity that accounts for its ability to potentiate ciprofloxacin activity. Notably, the enhanced efficacy of combination therapy with pentamidine and ciprofloxacin versus MDR P. aeruginosa strains in vivo merits further investigation into its potential to treat infections via this pathogen in patients.Publisher PDFPeer reviewe

A computational investigation of COVID-19 transmission inside hospital wards and associated costs

The COVID-19 pandemic has placed a particular burden on hospitals: from intra-hospital transmission of the infections to reduced admissions of non-COVID-19 patients. There are also high costs associated with the treatment of hospitalised COVID-19 patients, as well as reductions in revenues due to delayed and cancelled treatments. In this study we investigate computationally the transmission of COVID-19 inside a hospital ward that contains multiple-bed bays (with 4 or 6 beds) and multiple single-bed side rooms (that can accommodate the contacts of COVID-19-positive patients). The aim of this study is to investigate the role of 4-bed bays vs. 6-bed bays on the spread of infections and the hospital costs. We show that 4-bed bays are associated with lower infections only when we reduce the discharge time of some patients from 10 days to 5 days. This also leads to lower costs for the treatment of COVID-19 patients. In contrast, 6-bed bays are associated with reduced hospital waiting lists (especially when there are also multiple side rooms available to accommodate the contacts of COVID-19-positive patients identified inside the 6-bed bays).</p

Community antibiotic prescribing in patients with COVID-19 across three pandemic waves:a population-based study in Scotland, UK

Objectives: This study aims to examine community antibiotic prescribing across a complete geographical area for people with a positive COVID-19 test across three pandemic waves, and to examine health and demographic factors associated with antibiotic prescribing.Design: A population-based study using administrative data.Setting: A complete geographical region within Scotland, UK.Participants: Residents of two National Health Service Scotland health boards with SARS-CoV-2 virus test results from 1 February 2020 to 31 March 2022 (n=184 954). Individuals with a positive test result (n=16 025) had data linked to prescription and hospital admission data ±28 days of the test, general practice data for high-risk comorbidities and demographic data.Outcome measures: The associations between patient factors and the odds of antibiotic prescription in COVID-19 episodes across three pandemic waves from multivariate binary logistic regression.Results: Data included 768 206 tests for 184 954 individuals, identifying 16 240 COVID-19 episodes involving 16 025 individuals. There were 3263 antibiotic prescriptions ±28 days for 2395 episodes. 35.6% of episodes had a prescription only before the test date, 52.3% of episodes after and 12.1% before and after. Antibiotic prescribing reduced over time: 20.4% of episodes in wave 1, 17.7% in wave 2 and 12.0% in wave 3. In multivariate logistic regression, being female (OR 1.31, 95% CI 1.19 to 1.45), older (OR 3.02, 95% CI 2.50 to 3.68 75+ vs &lt;25 years), having a high-risk comorbidity (OR 1.45, 95% CI 1.31 to 1.61), a hospital admission ±28 days of an episode (OR 1.58, 95% CI 1.42 to 1.77) and health board region (OR 1.14, 95% CI 1.03 to 1.25, board B vs A) increased the odds of receiving an antibiotic.Conclusion: Community antibiotic prescriptions in COVID-19 episodes were uncommon in this population and likelihood was associated with patient factors. The reduction over pandemic waves may represent increased knowledge regarding COVID-19 treatment and/or evolving symptomatology

Carbapenem-only combination therapy against multi-drug resistant Pseudomonas aeruginosa : assessment of in vitro and in vivo efficacy and mode of action

Funding: This research was funded by the University of St Andrews.The aim of the study was to determine the efficacy of carbapenem-only combination treatments derived from four approved drugs (meropenem, doripenem, ertapenem and imipenem) against a MDR strain of P. aeruginosa in a Galleria mellonella larvae infection model. G. mellonella larvae were infected with P. aeruginosa NCTC 13437 (carrying the VIM 10 carbapenamase) and the efficacy of the six possible dual, four triple, and one quadruple carbapenem combination(s) were compared to their constituent monotherapies. Four of these combinations showed significantly enhanced survival compared to monotherapies and reduced the bacterial burden inside infected larvae but without complete elimination. Bacteria that survived combination therapy were slower growing, less virulent but with unchanged carbapenem MICs—observations that are consistent with a persister phenotype. In vitro time-kill assays confirmed that the combinations were bactericidal and confirmed that a low number of bacteria survived exposure. Mass spectrometry was used to quantify changes in the concentration of carbapenems in the presence of carbapenemase-carrying P. aeruginosa. The rate of degradation of individual carbapenems was altered, and often significantly reduced, when the drugs were in combinations compared with the drugs alone. These differences may account for the enhanced inhibitory effects of the combinations against carbapenem-resistant P. aeruginosa and are consistent with a ‘shielding’ hypothesis. In conclusion, carbapenem combinations show promise in combating MDR P. aeruginosa and are worthy of additional study and development.Publisher PDFPeer reviewe