The multi-factorial modes of action of urease in the pathogenesis of incontinence associated dermatitis.

Background: Incontinence Associated Dermatitis (IAD) is a type of skin inflammation caused by chronic exposure to urine and/or faeces. Current treatment strategies involve creating a barrier between the skin and urine/faeces rather than targeting specific irritants. Urease expressing pathogens catalyse the conversion of urea, present in urine, into ammonia. The accumulation of ammonia causes an elevation in skin pH which is believed to activate faecal enzymes which damage skin, and opportunistic pathogens, which lead to secondary infections. Objectives: To develop a better, multi-factorial model of IAD pathogenesis, including the effect of urease-expressing bacteria on skin, mechanism of damage of urease and urease-triggered activity of faecal enzymes and secondary pathogens. To study the effect of urease inhibition on preventing IAD skin damage. Methods: Five separate studies were made using ex vivo porcine skin and in vivo human skin models. Measurements of the change in skin barrier function were made using skin impedance, trans-epidermal water loss (TEWL), stratum corneum moisture and pH. Skin was exposed to artificial urine, inoculated with various microbes, enzymes and chemicals to examine the influence of: 1) urease-positive Proteus mirabilis 2) ammonia, 3) combination of P. mirabilis and a faecal enzyme, trypsin, 4) combination of P. mirabilis and opportunistic pathogens, Candida albicans and Staphylococcus aureus, 5) inhibition of urease using acetohydroxamic acid (AHA) on barrier function. Results: The urease-mediated production of ammonia had two principal effects: it elevated skin pH and caused inflammation, leading to significant breakdown in skin (stratum corneum) barrier function. Urease was found to further increase the activity of faecal enzymes and opportunistic pathogens, due to elevated skin pH. The urease inhibitor, AHA, was shown to have significantly reduced damage to skin barrier function, measured as its electrical resistance. Conclusions: Targeted therapeutic strategies should be developed to prevent the manifestation of IAD, rather than creating a generic barrier between skin and urine/faeces. Urease has been identified as a crucial component in the manifestation of IAD, due to its role in the production of ammonia. Urease inhibition provides a promising therapeutic target to halt the progression of IAD.

A TCF-based fluorescent probe to determine nitroreductase (NTR) activity for a broad-spectrum of bacterial species.

A nitroreductase (NTR) responsive fluorescent probe with long wavelength fluorescence emission was used to determine the NTR activity of a selection of bacterial species under a range of different bacterial growth conditions ensuring applicability under multiple complex clinical environments, where sensitivity, reaction time, and the detection accuracy were suitable for planktonic cultures and biofilms.

Cold Atmospheric Plasma-Activated Composite Hydrogel for an Enhanced and On-Demand Delivery of Antimicrobials.

We present the concept of a versatile drug-loaded composite hydrogel that can be activated using an argon-based cold atmospheric plasma (CAP) jet to deliver both a drug and CAP-generated molecules, concomitantly, in a tissue target. To demonstrate this concept, we utilized the antibiotic gentamicin that is encapsulated in sodium polyacrylate (PAA) particles, which are dispersed within a poly(vinyl alcohol) (PVA) hydrogel matrix. The final product is a gentamicin-PAA-PVA composite hydrogel suitable for an on-demand triggered release using CAP. We show that by activating using CAP, we can effectively release gentamicin from the hydrogel and also eradicate the bacteria effectively, both in the planktonic state and within a biofilm. Besides gentamicin, we also successfully demonstrate the applicability of the CAP-activated composite hydrogel loaded with other antimicrobial agents such as cetrimide and silver. This concept of a composite hydrogel is potentially adaptable to a range of therapeutics (such as antimicrobials, anticancer agents, and nanoparticles) and activatable using any dielectric barrier discharge CAP device.

Detecting and monitoring incontinence associated dermatitis: Does impedance spectroscopy have a part to play?

In this review, current understanding of the prevention and treatment of Incontinence Associated Dermatitis (IAD) is discussed. The need for preventative measures which target specific faecal/urinary irritants is highlighted, including the role of urease inhibitors. There is no existing internationally and clinically accepted method to diagnose and categorise the severity of IAD. Diagnosis currently relies on visual inspection; non-invasive techniques to assess skin barrier function could remove subjectiveness, particularly in darker skin tones. Impedance spectroscopy is a non-invasive technique which can be used to monitor skin barrier function, supporting visual assessments. Six studies (2003-2021) which used impedance to assess dermatitis were reviewed; inflamed skin was distinguishable from healthy skin in each case. This suggests that impedance spectroscopy could be useful in diagnosis early-stage IAD, potentially enabling earlier intervention. Finally, the authors present their initial findings on the role of urease in skin breakdown in an in vivo IAD model, using impedance spectroscopy.

TCF-based fluorescent probe for monitoring superoxide anion produced in bacteria under chloramphenicol- and heat-induced stress.

We report on a superoxide anion (O2˙-) responsive fluorescent probe called TCF-OTf. TCF-OTf is able to monitor O2˙- production when the bacterial species Pseudomonas aeruginosa, Staphylococcus aureus, Escherichia coli, and Enterococcus faecalis are exposed to chloramphenicol and heat shock at 50 and 58 °C.

Optimisation of a lozenge-based sensor for detecting impending blockage of urinary catheters.

Catheter-associated urinary tract infections resulting from urease-positive microorganisms are more likely to cause a urinary catheter blockage owing to the urease activity of the microbes. Catheter blockage can be dangerous and increases the risk of severe infections, such as sepsis. Ureases, a virulence factor in Proteus mirabilis, cause an increase in urine pH - leading to blockage. An optimised biosensor "lozenge" is presented here, which is able to detect impending catheter blockage. This lozenge has been optimised to allow easy manufacture and commercialisation. It functions as a sensor in a physiologically representative model of a catheterised urinary tract, providing 6.7 h warning prior to catheter blockage. The lozenge is stable in healthy human urine and can be sterilized for clinical use by ethylene oxide. Clinically, the lozenge will provide a visible indication of impending catheter blockage, enabling quicker clinical intervention and thus reducing the morbidity and mortality associated with blockage.

A small-molecular inhibitor against Proteus mirabilis urease to treat catheter-associated urinary tract infections.

Infection and blockage of indwelling urinary catheters is significant owing to its high incidence rate and severe medical consequences. Bacterial enzymes are employed as targets for small molecular intervention in human bacterial infections. Urease is a metalloenzyme known to play a crucial role in the pathogenesis and virulence of catheter-associated Proteus mirabilis infection. Targeting urease as a therapeutic candidate facilitates the disarming of bacterial virulence without affecting bacterial fitness, thereby limiting the selective pressure placed on the invading population and lowering the rate at which it will acquire resistance. We describe the design, synthesis, and in vitro evaluation of the small molecular enzyme inhibitor 2-mercaptoacetamide (2-MA), which can prevent encrustation and blockage of urinary catheters in a physiologically representative in vitro model of the catheterized urinary tract. 2-MA is a structural analogue of urea, showing promising competitive activity against urease. In silico docking experiments demonstrated 2-MA's competitive inhibition, whilst further quantum level modelling suggests two possible binding mechanisms.