**2. Wound infections**

conditions [1]. The wound healing is a dynamic process consisting of four continuous and precisely programmed phases, namely haemostasis, inflammation, proliferation and remodelling. Multiple factors, such as infections, stress, diabetes, smoking and obesity, can lead to impaired wound healing by interfering with one or more of these phases [2]. Once injured, the skin loses many of the protective defence mechanism of the intact skin and is colonized by the microorganisms on its surface. According to the replication status of the microorganisms, a wound can be classified as contaminated, colonized, locally infected and/or with spreading invasive infection [3]. The main bacterial mode of living in an infected wound is biofilm, which can be defined as a confluent community of adherent bacteria characterized by high cell densities and encased in an extracellular polymeric matrix that acts as physical barrier for biological and pharmaceutical antimicrobials [4, 5]. The presence of bacterial biofilm is associated with impaired epithelialization and granulation tissue formation and promotes a low-grade inflammatory response that interferes with wound healing [6]. The biofilm matrix plays an important role in the increased antibiotic resistance and has an enormous impact on medicine in terms of both therapeutic options and costs. Biofilm has been estimated to be associated with 65% of nosocomial infections, and the treatment costs associated with biofilm infection and chronic wounds have been estimated to be more than 1 billion USD annually in

The increasing resistance of bacteria to antibiotics represents a huge concern, so that the World Health Organization recently has described the problem as 'so serious that it threatens the achievements of modern medicine' [8, 9]. Moreover, the large number of wound dressings and the limited guidelines available have induced an undesirable inconsistency in wound-care practice [10]. The local treatment of wounds is crucial for preventing infections, controlling exudates and providing the moist environment necessary for wound healing. At this purpose, efforts have been made by many research groups in the development of bioactive dressings, which can play an active role in wound protection and healing, and/or are able to release biomolecules and antimicrobials for prevention and treatment of wound infections [8, 11, 12]. A strategy for the treatment of infected wounds with increased resistance to traditional antibiotic therapy is the use of specific antibacterial agents immobilized on the surface of a material, thus providing a wide spectrum of activity in terms of bacterial toxicity and destruc-

This chapter aims to provide the reader with an overview of the most promising routes to develop advanced biomaterials with antimicrobial properties for the management of wound infections through nanotechnology approaches. The new generation and application of nanomaterials with novel properties are one of the century's key technology developments, which offer extraordinary opportunities in the pharmaceutical and medical field [14]. The great potential of nanometals such as zinc, copper and silver in wound dressing formulations and their use as antimicrobial agent in wound infections will be presented, along with the most recent efforts and results achieved by several research group in the definition of effective strategies for prevention of wound infection and for enhanced wound healing. Moreover, the most relevant results obtained by the authors of this chapter in the field of silver-based antibacterial treatments for wound-healing application will be presented and discussed.

the United States [4, 7].

436 Wound Healing - New insights into Ancient Challenges

turation of the bacterial biofilm matrix [13].

The skin represents a complex and effective barrier between the organism and the environment, preventing invasion of pathogens, chemical and physical insults and unregulated loss of water and solutes [15]. From a microbiological point of view, the primary function of normal and intact skin is to control the microbial populations that live on the skin surface and to prevent the underlying tissues from invasion and colonization by potential pathogens [16]. A wound, which represents the loss of skin integrity and following exposure of subcutaneous tissue, provides a moist, warm and nutritious environment for microbial colonization and proliferation. The abundance and diversity of microorganisms in any wound depend on different factors such as wound type, depth, location and quality, the level of tissue perfusion and the antimicrobial efficacy of the host immune response [16]. As all open wounds lack the protective covering of skin, microorganisms from endogenous or exogenous sources can be introduced onto the wound surface and can lead to colonization [17, 18]. Colonization is defined as the presence of proliferating bacteria on the surface of a wound, without a noticeable host response and without clinical signs and symptoms. Differently, wound infection depends on the pathogenicity of the microorganisms and on the immune competency of the host, and it is characterized by the presence of the clinical signs of infection such as erythema, pain, tenderness, heat, oedema, cellulites and abscess or pus [19, 20]. Within an infected wound, the main bacterial mode of living is a biofilm [4]. Bacterial biofilm consists of a complex microenvironment of single or mixed bacterial species encased within an extracellular polymeric substance (EPS) produced by bacteria. The moist, adhesive and proteinaceous wound surface represents the ideal environment for biofilm development [21]. If microbes attach to the wound surface and proliferate, the biofilm begins to develop and, when it is well established, it exhibits resistance to the host immune system and antimicrobials. At this stage, the biofilm is considered mature and difficult to eradicate, thus requiring specialized management practices and increasing the risk of non-healing and clinically infected wound (i.e. showing signs of inflammation or purulence) [17]. Biofilm infections compromise wound closure and contribute to wound chronicity. Persistent infections may arrest the growth of the repairing tissue and significantly [22] impairing the key healing processes such as the inflammatory immune response, granulation tissue formation and epithelialization. Although a moist environment is necessary for optimal wound healing, poor moisture/exudate control within a wound environment promotes the development of biofilm. Consequently, moisture balance is essential to optimize the wound environment for healing and minimize the opportunity for biofilms to develop [23–25]. Preventing biofilm is fundamental for faster and more effective treatment of chronic wounds [17]. However, despite the evidence for the presence of biofilm in wounds, research studies are required to detect biofilm and to determine the exact role played by multispecies biofilms in terms of delayed wound-healing process [26]. Different biofilms can be identified within a wound environment, such as aggregates of cells dispersed within the wound exudate, in slough or on necrotic tissue or on the wound dressing [27]. The microbial community presents multiple difficulties for clinicians in attempting to heal a chronic wound. Biofilms are resistant to many biocides, antibiotics and wound-care products. So, managing biofilm often involves its physical removal from the wound surface with sharp or surgical debridement [28].

The control of biofilm is a key part of chronic wound management, but the use of antiseptic dressings for preventing and managing biofilm and infection still needs further research involving well-designed, randomized controlled trials [29]. The concept of a bacterial contamination, colonization and biofilm-related infection is now widely accepted in wound care, and the recognition of the biofilm and the evolution of topical antiseptics to control bioburden in wounds are considered strictly related to the concept of TIME (tissue, infection/inflammation, moisture balance and edge of wound) and to its relation with the current best practice [30]. In healthcare, infections lead to longer hospital stays for patients, specifically wound dressings and increased hospital costs [12]. Also worsened by an ageing population and the incidence of diabetes and obesity, the huge economic and social impact of wounds requires higher level of attention and resources to understand biological mechanisms underlying cutaneous wound complications [31].

Infections of the dermis, including burns, surgical site infections and non-healing diabetic foot ulcers affect over a million people. Individuals with diabetes represent a particularly vulnerable category because many of them develop foot ulceration during the course of their disease and undergo amputation. In addition to diabetics, several other groups of immune-compromised patient populations are plagued by slow-healing and non-healing wounds, such as trauma and burn victims, cancer patients and pressure ulcers in the elderly [32]. The incidence, morbidity, mortality and costs associated with non-healing of chronic skin wounds are dramatic. Chronic wounds cost millions of dollars annually in the healthcare industry of the United States, and biofilm significantly contributes many billions of dollars to the global cost of chronic wounds because of its role in delaying the wound-healing process and extending the inflammatory phase of repair [19, 33–35].

Along with the direct medical costs borne by the hospital or insurer, also indirect costs including lost patient productivity and diminished functional status should be considered [36]. The control of bioburden is recognized as an important aspect of wound management, which requires new solutions against microbes and their biofilms. Octenidine dihydrochloride and polyhexanide are effective and tolerated antiseptics used in wound management today, but antiseptics alone may not be able to achieve wound healing without addressing other factors such as the general health of patients or the wound's physical environment [37, 38]. Next generation of wound treatment strategies for non-healing chronic wounds can be achieved by adopting a biofilm-based management approach to wound care, in order to kill and prevent reattachment of microorganisms [26].
