<Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> PAO1 Resistance to Zinc Pyrithione: Phenotypic Changes Suggest the Involvement of Efflux Pumps

The aim of this study is to investigate the involvement of an efflux pump in the development of Pseudomonas aeruginosa resistance to zinc pyrithione (ZnPT). In the presence of efflux inhibitor carbonyl cyanide m-chlorophenyl-hydrazone (CCCP), the minimum inhibitory concentration of ZnPT for P. aeruginosa resistant cells is reduced significantly (p < 0.05). In addition, the concentration of ZnPT excluded by the resistant bacteria was reduced significantly (p < 0.01). However, the above reductions did not reach the levels measured for P. aeruginosa PAO1 sensitive strain. Furthermore, such changes in P. aeruginosa resistant cells were correlated with the overexpression of outer membrane proteins, reduced sensitivity toward imipenem (p < 0.01) and increased sensitivity toward sulphatriad and chloramphenicol (p < 0.05). In a continuation to a previous study, we conclude that P. aeruginosa resistance to ZnPT is multifactorial and involves induced efflux systems. Suzanne Abdel Malek is currently on leave from Petra University, and a member at the Faculty of Applied Medical Sciences, King Abdulaziz University, Jeddah, Saudi Arabia.     

Antimicrobial susceptibility changes and T-OMP shifts in pyrithione-passaged planktonic cultures of Pseudomonas aeruginosa PAO1

AIMS: The aim of this study was to determine whether passaging Pseudomonas aeruginosa PAO1 with sub-MICs of the pyrithione biocides results in both the induction of decreased susceptibility towards these antimicrobials and associated outer membrane profile changes. METHODS AND RESULTS: Previous work by this group has shown that it is possible to induce susceptibility changes towards the isothiazolone biocides in Ps. aeruginosa PAO1 by successive passages in the presence of increasing sub-MICs of biocide. This procedure was accompanied by the loss of a 35 kDa outer membrane protein, T-OMP. In this experiment, this process was repeated with the biocides sodium pyrithione (NaPT), zinc pyrithione (ZnPT) and cetrimide. The pattern of susceptibility was similar to that observed with the isothiazolone biocides. Upon removal of biocide, the observed MIC did not return to the original pre-exposure value. The onset and development of resistance was accompanied by the loss of T-OMP from outer membrane profiles, which suggests that this is a non-specific membrane channel whose production within the cell is sensitive to biocide presence. The T-OMP reappeared when the cells were passaged in the absence of pyrithione. Cross-resistance studies indicated that induced resistance to one biocide yields partial resistance towards other members of the group and the positive control. CONCLUSIONS: These results indicate that the pyrithione biocides have similar susceptibility profiles in Ps. aeruginosa to those exhibited by the isothiazolones, but that the acquired changes in susceptibility to the pyrithiones is largely irreversible. SIGNIFICANCE AND IMPACT OF THE STUDY: This study indicates that acquired susceptibility changes towards sub-MICs of selected biocides are multifactorial in nature.     

Microemulsions are membrane-active, antimicrobial, self-preserving systems

Microemulsions are physically stable oil/water systems that have potential use as delivery systems for many pharmaceuticals which are normally of limited use due to their hydrophobicity, toxicity or inability to access the site of action. It has been suggested that microemulsions are self-preserving antimicrobials in their own right, although there is little evidence to support this. In this experiment, microemulsions of various compositions were formulated and tested for their stability and antimicrobial action. The physical stability of the different microemulsions was assessed by centrifugation at 4000g and by storage in a water bath at 37 degrees C for one month, during which no phase separation was observed. The antimicrobial activity of the microemulsions was tested using the compendial method, observation of the kinetics of killing, and transmission electron microscopy (TEM) of microemulsion-exposed cultures of Pseudomonas aeruginosa PA01. These latter experiments on Ps. aeruginosa indicated distinct signs of membrane disruption. The results indicated that the microemulsions are self-preserved, and that their killing of microbial cultures is very rapid and may be the result of membrane activity.     

A review of the antiviral activity of Chitosan,including patented applications and its potential use against COVID-19

Chitosan is an abundant organic polysaccharide, which can be relatively easily obtained by chemical modification of animal or fungal source materials. Chitosan and its derivatives have been shown to exhibit direct antiviral activity, to be useful vaccine adjuvants and to have potential anti-SARS-CoV-2 activity. This thorough and timely review looks at the recent history of investigations into the role of chitosan and its derivatives as an antiviral agent and proposes a future application in the treatment of endemic SARS-CoV-2.     

Microemulsions are highly effective anti-biofilm agents

AIMS: The demonstration of the antibiofilm effects of pharmaceutical microemulsions. METHODS AND RESULTS: Microemulsions were prepared as physically stable oil/water systems. Previous work by this group has shown that microemulsions are highly effective antimembrane agents that result in rapid losses of viability in planktonic populations of Pseudomonas aeruginosa and Staphylococcus aureus. In this experiment a microemulsion preparation was used upon established biofilm cultures of Ps. aeruginosa PA01 for a period of 4 h. The planktonic MIC of sodium pyrithione and the planktonic and biofilm MICs of cetrimide were used as positive controls and a biofilm was exposed to a volume of normal sterile saline as a treatment (negative) control. Results indicate three log-cycle reductions in viability within the microemulsion treated biofilm, as compared to those observed in control treatments of similar biofilms (one log-cycle reduction in viabilities). CONCLUSIONS: The results indicate that the microemulsions are highly effective antibiofilm agents. SIGNIFICANCE AND IMPACT OF THE STUDY: This study suggests that microemulsions may have a role in the treatment of industrial and environmental biofilms.     

Outer membrane protein shifts in biocide-resistant Pseudomonas aeruginosa PAO1

Benzisothiazolone (BIT), N-methylisothiazolone (MIT) and 5-chloro-N-methylisothiazolone (CMIT) are highly effective biocidal agents and are used as preservatives in a variety of cosmetic preparations. The isothiazolones have proven efficacy against many fungal and bacterial species including Pseudomonas aeruginosa. However, some species are beginning to exhibit resistance towards this group of compounds after extended exposure. This experiment induced resistance in cultures of Ps. aeruginosa exposed to incrementally increasing sub-minimum inhibitory concentrations (MICs) of the isothiazolones in their pure chemical forms. The induced resistance was observed as a gradual increase in MIC with each new passage. The MICs for all three test isothiazolones and a thiol-interactive control compound (thiomersal) increased by approximately twofold during the course of the experiment. The onset of resistance was also observed by reference to the altered presence of an outer membrane protein, designated the T-OMP, in SDS-PAGE preparations. T-OMP was observed to disappear from the biocide-exposed preparations and reappear when the resistance-induced cultures were passaged in the absence of biocide. This reappearance of T-OMP was not accompanied by a complete reversal of induced resistance, but by a small decrease in MIC. The induction of resistance towards one biocide resulted in the development of cross-resistance towards other members of the group and the control, thiomersal. It has been suggested that the disappearance of T-OMP from these preparations is associated with the onset of resistance to the isothiazolones in their Kathon form (CMIT and MIT).     

Pyrithione biocides as inhibitors of bacterial ATP synthesis

Sodium pyrithione and zinc pyrithione (NaPT and ZnPT, respectively) are widely used as cosmetic preservatives and general antimicrobial agents. They have been shown to be active against fungal cell walls, associated membranes and bacterial transport processes. Investigations were undertaken into the effect of these antimicrobial agents on substrate catabolism and intracellular ATP levels using an oxygen electrode and luciferin-luciferase technology, respectively. Results indicate that, while both compounds are poor inhibitors of substrate catabolism, sub-inhibitory concentrations of biocide greatly reduces intracellular ATP levels in both Escherichia coli NCIMB 10000 and Pseudomonas aeruginosa NCIMB 10548. This is thought to be due to the action of NaPT and ZnPT on the Gram-negative bacterial membrane.