A Host-Directed Approach to the Detection of Infection in Hard-to-Heal Wounds.

Wound infection is traditionally defined primarily by visual clinical signs, and secondarily by microbiological analysis of wound samples. However, these approaches have serious limitations in determining wound infection status, particularly in early phases or complex, chronic, hard-to-heal wounds. Early or predictive patient-derived biomarkers of wound infection would enable more timely and appropriate intervention. The observation that immune activation is one of the earliest responses to pathogen activity suggests that immune markers may indicate wound infection earlier and more reliably than by investigating potential pathogens themselves. One of the earliest immune responses is that of the innate immune cells (neutrophils) that are recruited to sites of infection by signals associated with cell damage. During acute infection, the neutrophils produce oxygen radicals and enzymes that either directly or indirectly destroy invading pathogens. These granular enzymes vary with cell type but include elastase, myeloperoxidase, lysozyme, and cathepsin G. Various clinical studies have demonstrated that collectively, these enzymes, are sensitive and reliable markers of both early-onset phases and established infections. The detection of innate immune cell enzymes in hard-to-heal wounds at point of care offers a new, simple, and effective approach to determining wound infection status and may offer significant advantages over uncertainties associated with clinical judgement, and the questionable value of wound microbiology. Additionally, by facilitating the detection of early wound infection, prompt, local wound hygiene interventions will likely enhance infection resolution and wound healing, reduce the requirement for systemic antibiotic therapy, and support antimicrobial stewardship initiatives in wound care.

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.

An In Vitro Bladder Model for Studying Catheter-Associated Urinary Tract Infection and Associated Analysis of Biofilms.

Urethral catheters are among the most widely used medical devices, applied to manage a wide range of conditions in hospital, community, and care home settings. In long-term catheterized individuals, infection with Proteus mirabilis frequently complicates the care of patients owing to formation of extensive crystalline biofilms. Here we describe the use of an in vitro bladder model of the catheterized urinary tract and associated analyses to study P. mirabilis crystalline biofilm formation. The model originally described by Stickler et al. (1999, 310:494-501, Methods Enzymol) replicates a complete sterile closed drainage system as used in clinical practice, and permits formation of biofilms directly on catheters under conditions representative of those encountered in vivo. Models may be used to replicate either established infection or early stage colonization, and we describe a range of associated methods for quantification and visualization of biofilms formed on catheters. These methods are also easily adapted to study catheter-associated biofilm formation by other urinary tract pathogens.

Emerging medical and engineering strategies for the prevention of long-term indwelling catheter blockage.

Urinary catheters have been used on an intermittent or indwelling basis for centuries, in order to relieve urinary retention and incontinence. Nevertheless, the use of urinary catheters in the clinical setting is fraught with complication, the most common of which is the development of nosocomial urinary tract infections, known as catheter-associated urinary tract infections. Infections of this nature are not only significant owing to their high incidence rate and subsequent economic burden but also to the severe medical consecutions that result. A range of techniques have been employed in recent years, utilising various technologies in attempts to counteract the perilous medical cascade following catheter blockage. This review will focus on the current advancement (within the last 10 years) in prevention of encrustation and blockage of long-term indwelling catheters both from engineering and medical perspectives, with particular emphasis on the importance of stimuli-responsive systems.

Development of an Infection-Responsive Fluorescent Sensor for the Early Detection of Urinary Catheter Blockage.

Formation of crystalline biofilms following infection by Proteus mirabilis can lead to encrustation and blockage of long-term indwelling catheters, with serious clinical consequences. We describe a simple sensor, placed within the catheter drainage bag, to alert of impending blockage via a urinary color change. The pH-responsive sensor is a dual-layered polymeric "lozenge", able to release the self-quenching dye 5(6)-carboxyfluorescein in response to the alkaline urine generated by the expression of bacterial urease. Sensor performance was evaluated within a laboratory model of the catheterized urinary tract, infected with both urease positive and negative bacterial strains under conditions of established infection, achieving an average "early warning" of catheter blockage of 14.5 h. Signaling only occurred following infection with urease positive bacteria. Translation of these sensors into a clinical environment would allow appropriate intervention before the occurrence of catheter blockage, a problem for which there is currently no effective control method.

Recent advances in therapeutic delivery systems of bacteriophage and bacteriophage-encoded endolysins.

Antibiotics have been the cornerstone of clinical management of bacterial infection since their discovery in the early 20th century. However, their widespread and often indiscriminate use has now led to reports of multidrug resistance becoming globally commonplace. Bacteriophage therapy has undergone a recent revival in battle against pathogenic bacteria, as the self-replicating and co-evolutionary features of these predatory virions offer several advantages over conventional therapeutic agents. In particular, the use of targeted bacteriophage therapy from specialized delivery platforms has shown particular promise owing to the control of delivery location, administration conditions and dosage of the therapeutic cargo. This review presents an overview of the recent formulations and applications of such delivery vehicles as an innovative and elegant tool for bacterial control.

Prevention of encrustation and blockage of urinary catheters by Proteus mirabilis via pH-triggered release of bacteriophage.

The crystalline biofilms of Proteus mirabilis can seriously complicate the care of patients undergoing long-term indwelling urinary catheterisation. Expression of bacterial urease causes a significant increase in urinary pH, leading to the supersaturation and precipitation of struvite and apatite crystals. These crystals become lodged within the biofilm, resulting in the blockage of urine flow through the catheter. Here, we describe an infection-responsive surface coating for urinary catheters, which releases a therapeutic dose of bacteriophage in response to elevated urinary pH, in order to delay catheter blockage. The coating employs a dual-layered system comprising of a lower hydrogel 'reservoir' layer impregnated with bacteriophage, capped by a 'trigger' layer of the pH-responsive polymer poly(methyl methacrylate-co-methacrylic acid) (EUDRAGIT®S 100). Evaluation of prototype coatings using a clinically reflective in vitro bladder model system showed that catheter blockage time was doubled (13 h to 26 h (P < 0.05)) under conditions of established infection (108 CFU ml-1) in response to a 'burst-release' of bacteriophage (108 PFU ml-1). Coatings were stable both in the absence of infection, and in the presence of urease-negative bacteria. Quantitative and visual analysis of crystalline biofilm reduction show that bacteriophage constitute a promising strategy for the prevention of catheter blockage, a clinical problem for which there is currently no effective control method.

An in-situ infection detection sensor coating for urinary catheters.

We describe a novel infection-responsive coating for urinary catheters that provides a clear visual early warning of Proteus mirabilis infection and subsequent blockage. The crystalline biofilms of P. mirabilis can cause serious complications for patients undergoing long-term bladder catheterisation. Healthy urine is around pH 6, bacterial urease increases urine pH leading to the precipitation of calcium and magnesium deposits from the urine, resulting in dense crystalline biofilms on the catheter surface that blocks urine flow. The coating is a dual layered system in which the lower poly(vinyl alcohol) layer contains the self-quenching dye carboxyfluorescein. This is capped by an upper layer of the pH responsive polymer poly(methyl methacrylate-co-methacrylic acid) (Eudragit S100®). Elevation of urinary pH (>pH 7) dissolves the Eudragit layer, releasing the dye to provide a clear visual warning of impending blockage. Evaluation of prototype coatings using a clinically relevant in vitro bladder model system demonstrated that coatings provide up to 12h advanced warning of blockage, and are stable both in the absence of infection, and in the presence of species that do not cause catheter blockage. At the present time, there are no effective methods to control these infections or provide warning of impending catheter blockage.