Hard-to-heal wounds cause significant patient suffering and place a heavy burden on healthcare systems, accounting for about 4 % of the entire healthcare budget globally. In hospital settings, up to 40% of beds are occupied by patients with wounds, and substantial resources are allocated to outpatient care. Wounds are highly susceptible to bacterial colonization, which can lead to infection and further delay healing. With the rise of multidrug-resistant bacteria, this challenge has become a global healthcare concern.

This thesis explores new antimicrobial strategies and techniques to modify nanocellulose wound dressings to enable both detection and treatment of infections in hard-to-heal wounds. Nanocellulose, and in particular bacterial nano-cellulose (BC), is an attractive wound dressing material that provides a moist wound microenvironment and a protective barrier against external pathogens but lacks inherent antimicrobial properties. Antimicrobial peptides (AMPs) have recently gained attention for wound treatment due to their potent antimicrobial effects and low risk of antibiotic resistance.

This thesis explores the possibility to functionalize BC wound dressings with AMPs, focusing on: i) enhancing AMPs loading, ii) preventing uncontrolled release, and iii) enabling triggered AMPs release to develop stimuli-responsive wound dressings. Initially, the physical adsorption of the AMPs PLNC8 αβ in BC was investigated. However, this approach resulted in low loading efficiency and uncontrolled release. To improve AMPs loading, mesoporous silica nanoparticles (MSNs) were self-assembled in the BC dressings, resulting in a BC-MSN composite with a threefold increase in specific surface area. The nanocomposites retained attractive wound dressing properties and AMPs loading proved to be more efficient, achieving up to a fourfold increase in AMPs concentrations. In a second approach loading of AMPs micelles and AMPs-loaded MSNs in a hyaluronic acid hydrogel that was subsequently grafted to BC was developed. This strategy ensured efficient AMPs loading and high antimicrobial activity while stimulating healing.

To attain a more precise AMPs release control, a strategy to coat AMPs-loaded MSNs with proteins was developed. Protein capped MSNs retained the AMPs in absence of proteases, while protease-triggered degradation of the capping led to a ~85% AMPs release. Protease activity is typically highly upregulated in infected wounds and the protease responsive AMPs release can thus facilitate efficient infection control. In addition to controlled AMPs delivery, this thesis explores novel AMPs designs. Starting with PLNC8 β, iterations of truncations, amino acid substitutions, and lipidation, lead to the discovery of a new class of sequence-optimized antimicrobial peptides (SOAPs) with broad-spectrum activity and potent antimicrobial action in the micromolar concentration range. To further enhance their efficacy and reduce the required dosage, SOAP peptides were loaded onto self-assembled BC-silver nanoparticle (AgNP) composite dressings, allowing for the simultaneous delivery of AMPs and silver ions.

Beyond infection treatment, this work also explores the potential of BC-MSN composite materials for wound infection detection. The high surface area of the composite was utilized to immobilize a pH-responsive molecule capable of detecting pH changes in the wound microenvironment indicative of infection. The immediate color change of the dressings in infected wounds could enable early and non-invasive wound status monitoring.

Overall, the functionalization strategies presented in this thesis provide a highly adaptable platform for development of multifunctional antimicrobial nano-cellulose wound dressings, offering new possibilities for advanced wound care.