Enhancement of antibiotic efficacy against multi-drug resistant Pseudomonas aeruginosa infections via combination with curcumin and 1-(1-Naphthylmethyl)-Piperazine

Objective: The aim of this study was to determine if the plant phenolic curcumin (CUR) and the arylpiperazine 1-(1-naphthylmethyl)-piperazine (NMP) could restore antibiotic efficacy versus MDR P. aeruginosa infection. Methods: The MICs of piperacillin, meropenem and levofloxacin in the presence or absence of CUR or NMP against a MDR strain that over-expresses the MexAB-OprM efflux-pump and the isogenic parent strain were compared. The efficacy of the same combination treatments was also tested in a Galleria mellonella in vivo infection model and larval survival and bacterial burden compared. Results: In vitro, CUR restored the activity of piperacillin, meropenem and levofloxacin versus the MDR strain of P. aeruginosa only weakly. There was no evidence in vitro of a similar effect with NMP. In vivo, treatment of G. mellonella larvae infected with the MDR strain with a combination of NMP or CUR plus levofloxacin, and piperacillin plus CUR, resulted in enhanced therapeutic benefit compared to the monotherapies. When compared with monotherapies, the enhanced efficacy of the combination treatments correlated with reduced bacterial burden. Conclusion: CUR and NMP restored the efficacy of antibiotic therapy in vivo versus MDR P. aeruginosa infection.Publisher PDFPeer reviewe

Exponential Mixing for a Stochastic PDE Driven by Degenerate Noise

We study stochastic partial differential equations of the reaction-diffusion type. We show that, even if the forcing is very degenerate (i.e. has not full rank), one has exponential convergence towards the invariant measure. The convergence takes place in the topology induced by a weighted variation norm and uses a kind of (uniform) Doeblin condition.Comment: 10 pages, 1 figur

Scoping study on the prospects for sustainable tree crop development in Sub-Saharan Africa. A paper prepared for the World Bank

The objectives of this short scoping study are to identify key issues for development of the tree crop sector particularly smallholder production, and to draw attention to opportunities for World Bank funding of tree crop initiatives as a focus for possible productive investment in Sub-Saharan Africa, and for in-depth examination in a wider study. The study looks particularly at issues relevant to new crop development/replanting and those arising from liberalisation

Wax moth larva (Galleria mellonella): An in vivo model for assessing the efficacy of antistaphylococcal agents

Objectives - To investigate whether the wax moth larva, Galleria mellonella, is a suitable host for assessing the in vivo efficacy of antistaphylococcal agents against Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) infections. Methods - Wax moth larvae were infected with increasing doses of S. aureus to investigate the effect of inoculum size on larval survival. In addition, infected wax moth larvae were treated with daptomycin, penicillin or vancomycin to examine whether these agents were effective against S. aureus and MRSA infections in vivo. Results - Increasing inoculum doses of live S. aureus cells resulted in greater larval mortality, but heat-killed bacteria and cell-free culture filtrates had no detrimental effects on survival. Larval mortality rate also depended on the post-inoculation incubation temperature. After larvae were infected with S. aureus, larval survival was enhanced by administering the antistaphylococcal antibiotics daptomycin or vancomycin. Larval survival increased with increasing doses of the antibiotics. Moreover, penicillin improved survival of larvae infected with a penicillin-susceptible methicillin-susceptible S. aureus (MSSA) strain, but it was ineffective at similar doses in larvae infected with MRSA (penicillin resistant). Daptomycin and vancomycin were also effective when administered to the larvae prior to infection with bacteria. Conclusions - This is the first report to demonstrate that antibiotics are effective in the wax moth larva model for the treatment of infections caused by Gram-positive bacteria. The new wax moth larva model is a useful preliminary model for assessing the in vivo efficacy of candidate antistaphylococcal agents before proceeding to mammalian studies, which may reduce animal experimentation and expense

Enhanced efficacy of putative efflux pump inhibitor/antibiotic combination treatments versus MDR strains of Pseudomonas aeruginosa in a Galleria mellonella in vivo infection model

This work was supported by the University of St Andrews.Objectives: The objectives of this study were to compare the antibiotic susceptibility of Pseudomonas aeruginosa strains with increased efflux pump expression in vitro and in vivo and to use these same strains to evaluate the efficacy of combinations of antibiotics with putative efflux pump inhibitors in vivo. Methods: A collection of P. aeruginosa strains that overexpress three efflux pumps (MexAB-OprM, MexCD-OprJ and MexEF-OprN), in addition to a strain with all three Mex pumps deleted, were used. The virulence of these strains and their antibiotic susceptibility was measured in vivo using a Galleria mellonella larval infection model. The inhibitory effect of combinations of putative efflux pump inhibitors (trimethoprim and sertraline) with antibiotics on the strain overexpressing MexAB-OprM was also measured in vitro and compared with their efficacy in vivo in terms of larval survival and bacterial burden. Results: Increased expression of the individual efflux pumps, or deletion of all three, had no significant effect on the virulence of P. aeruginosa in vivo. Expression levels of the efflux pumps clearly influenced antibiotic efficacy in vivo. The efficacy of levofloxacin, piperacillin and meropenem against larvae infected with the efflux pump mutants reflected susceptibility to the same drugs in vitro. Treatment of G. mellonella larvae infected with a strain that overexpressed MexAB-OprM with a combination of putative efflux pump inhibitors and levofloxacin resulted in enhanced therapeutic benefit compared with the constituent monotherapies. Conclusions: This study has demonstrated the utility of using G. mellonella to screen for novel therapeutic options for MDR P. aeruginosa and has shown that antibiotic/efflux pump inhibitor combinations should be further investigated for clinical application.PostprintPeer reviewe

Effective immunosuppression with dexamethasone phosphate in the Galleria mellonella larva infection model resulting in enhanced virulence of Escherichia coli and Klebsiella pneumoniae

MPT was the recipient of an ERASMUS training grant. FE is supported by the University of St Andrews.The aim was to evaluate whether immunosuppression with dexamethasone 21-phosphate could be applied to the Galleria mellonella in vivo infection model. Characterised clinical isolates of Escherichia coli or Klebsiella pneumoniae were employed, and G. mellonella larvae were infected with increasing doses of each strain to investigate virulence in vivo. Virulence was then compared with larvae exposed to increasing doses of dexamethasone 21-phosphate. The effect of dexamethasone 21-phosphate on larval haemocyte phagocytosis in vitro was determined via fluorescence microscopy and a burden assay measured the growth of infecting bacteria inside the larvae. Finally, the effect of dexamethasone 21-phosphate treatment on the efficacy of ceftazidime after infection was also noted. The pathogenicity of K. pneumoniae or E. coli in G. mellonella larvae was dependent on high inoculum numbers such that virulence could not be attributed specifically to infection by live bacteria but also to factors associated with dead cells. Thus, for these strains, G. mellonella larvae do not constitute an ideal infection model. Treatment of larvae with dexamethasone 21-phosphate enhanced the lethality induced by infection with E. coli or K. pneumoniae in a dose- and inoculum size-dependent manner. This correlated with proliferation of bacteria in the larvae that could be attributed to dexamethasone inhibiting haemocyte phagocytosis and acting as an immunosuppressant. Notably, prior exposure to dexamethasone 21-phosphate reduced the efficacy of ceftazidime in vivo. In conclusion, demonstration of an effective immunosuppressant regimen can improve the specificity and broaden the applications of the G. mellonella model to address key questions regarding infection.Publisher PDFPeer reviewe

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

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

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

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