Exposed subcutaneous tissue provides a favorable medium for microorganisms DUPLICATE

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1 ORIGINAL RESEARCH In-vitro Comparison of Antimicrobial Efficacy of Various Wound Dressing Materials WOUNDS 2010;22(7): From the 1 Department of Plastic Reconstructive and Aesthetic Surgery and 2 Department of Infection Diseases, Mersin University, Mersin, Turkey Address correspondence to: Yavuz Basterzi, MD Mersin University, Medical Faculty Zeytinlibahce cd Mersin, Turkey basterziyavuz@yahoo.com Yavuz Basterzi, MD; 1 Gulden Ersoz, MD; 2 Guliz Sarac, MD; 2 Alper Sari, MD; 1 Ferit Demirkan, MD 1 Abstract: Many new antimicrobial dressings have been used for the treatment of or protection against wound infection since the early 1980s. There are many different wound dressings such as silver, povidone iodine, and chlorhexidine hexidine impregnated materials on the market today. Various manufacturers acturers assert that their dressings are the most effective and therefore should be preferentially employed. However, it is difficult to find a study that clearly identifies the most effective antimicrobial wound dressing. Methods. Eight different commercially available wound dressing materials were compared in terms of their antimicrobial al effectiveness on 18 different microorganisms via disk diffusion test (Kirby-Bauer Method) on Mueller-Hinton (MH) agar. Among the 16 bacterial and 2 yeast species grown on MH agar plates, Contreet was thd the most effective antimicrobial dressing tested (P < 0.001). 01). Results. A statistical difference was not found concerning efficacy against gram-positive and -negative bacteria among any of the materials except for Aquacel Ag and Inadine, the activity of which were found to be higher on gram-positive bacteria compared to gramnegative bacteria (P = 0.029, P = 0.030). In-vitro data suggest that Contreet is the most effective for topical treatment of contaminated wounds. Conclusion. Further methods of assessment, including the Nw use of infected animal models and clinical studies, will be necessary to better understand the antimicrobial efficacy of these dressings. ud DUPLICATE Exposed subcutaneous tissue provides a favorable medium for microorganisms to contaminate and colonize. The conditions become optimal for microbial growth if the involved tissue is devitalized and/or the host immunity is compromised. This situation presents a considerable problem for both the patient and caregiver. Oral and systemic antibiotics are often used to control infection. While systemic antibiotheraphy is mandatory for advanced skin infections, from a basic wound management perspective, the three methods of eliminating a local wound infection are debridement, wound cleansing, and topical antimicrobial application. 1,2 Many different topical antimicrobial agents are available such as silver, povidone iodine, chlorhexidine, and honey. Vol. 22, No. 7 July

2 Silver has been used as an antimicrobial for thousands of years either as liquid, cream, or more recent- Bactigras (Smith and Nephew) Product ly, silver-coated wound Inadin (Johnson & Johnson) dressings. Its antimicrobial MelDra (MedEllens) activity is attributed to Textus (Biocell) Acticoat (Smith & Nephew) its ability to block the Aquacel Ag. (ConvaTec) transmembranous energy Contreet (Coloplast) metabolism in bacteria. 3 Ultec (Kendall, Covidien) Chlorhexidine is a persistent antimicrobial agent that is active against a wide range of gram-positive and gram-negative bacteria. It has been widely used for surgical hand and skin antisepsis. Povidone iodine is another commonly used antimicrobial agent, which has been demonstrated to be effective in killing a broad range of the microorganisms typically associated with wound infection. 4 The antimicrobial properties of honey in relation to wound management date back to ancient times. Recently, studies have been published regarding the antimicrobial activity of honey against methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa. 5,6 Advancements in science and technology have spawned significant improvements in wound dressings. Some newer dressings incorporate antimicrobials and overcome the disadvantages associated with topical antimicrobial application. Currently, various different antimicrobial wound dressings are available on the market. However, studies that clearly identify which materials are superior do not exist. The aim of the present study was to compare the antimicrobial properties pertie of different commercially available wound dressing materials. Materials and Methods Organisms. Eighteen microorganisms were evaluated in this study and all clinical isold isolates plus one standard strain S aureus ATCC were collected from routine clinical species submitted to the Mersin University Hospital (Mersin, Turkey). All isolates were identified morphologically and biochemically by a standard laboratory procedure. The following species were used in the experiments: extended spectrum beta-lactamase (ESBL), producing and non-producing Escherichia coli, Acinetobacter baumannii (carbapenem resistant), Stenotrophomanas maltophilia, ESBL producing and non-producing Klebsiella pneumonia, Pseudomonas aeruginosa, Enterobacter cloacae, methicillin-sensitive Staphylococcus aureus (MSSA), methicillin-resistant 166 WOUNDS Table 1. Antimicrobial contents of dressing materials were tested. pertien singsḋ DUPLIC DUP PLICATE Antimicrobial contents Chlorhexidine acetate Povidone-iodine Medical honey Silver Nanocrystalline silver crystals Ionic silver Ionic silver -- (used as control) E- TE- Figure 1. Pieces of dressings were placed on the inoculated Mueller-Hinton (MH) agar plates and observed for the zone of inhibition around the dressings. Staphylococcus aureus (MRSA), methicillin-sensitive coagulase-negative staphylococci (MSCNS), methicillinresistant coagulase-negative staphylococci (MRCNS), penicillin susceptible enterococci, vancomycin resistant enterococci (VRE), and two strains of Candida albicans. Dressing materials. The wound dressings tested were: Bactigras (Smith and Nephew, Hull, UK), Inadine (Systagenix, North Yorkshire, UK), MelDra (Dermagenics BV, The Netherlands), Textus (Biocell, Engelskirchen, Germany), Acticoat (Smith & Nephew, St. Petersburg, FL), Aquacel Ag (ConvaTec, Skillman, NJ), Contreet (Coloplast, Humlebaek, Denmark), Ultec (Covidien, Mansfield, MA). Ultec was used as a control (Table 1). All dressing materials were cut into 4-mm diameter discs under sterile conditions. Methods. The antimicrobial efficacy of the wound dressings was evaluated against various microorganisms using the zone of inhibition test. The tested organisms were sub-cultured on agar plates with 5% blood over an 18-hour period after which a suspension representing

3 Table 2. Diameter of the inhibition zones around the dressing discs for all microorganisms. Microorganisms The size of the zones of clearance around the discs (mm) Bactigras Inadine Meldra Textus Acticoat Aquacel Ag Contreet Ultec* E coli ESBL (-) E coli ESBL (+) Klebsiella pneumonia ESBL (-) Klebsiella pneumonia ESBL (+) Enterobacter cloacae Pseudomonas aeruginosa Acinetobacter baumannii Acinetobacter baumannii Stenotrophomanas maltophilia S aureus ATCC MSSA MRSA MSCNS MRCNS Enterococci VRE Candida albicans Candida albicans *Control group ESBL: Extended spectrum beta lactamase; MSSA: Methicillin sensitive Staphylococcus aureus; MRSA: Methicillin resistant Staphylococcus aureus; MSCNS: Methicillin sensitive coagulase negative Staphylococci; MRCNS: Methicillin resistant coagulase negative Staphylococci; VRE: Vancomycin resistant Enterococci t approximately 10 6 to 10 7 CFU/mL (0.5 Table 3. Comparison of the mean size of the inhibition zones McFarland) was prepared. The dressing discs around the dressing materials. were placed on the inoculated Mueller- n Minimum Maximum Mean ± SD P Hinton (MH) agar plates and observed for the (mm) (mm) (mm) zone of inhibition around the dressings (Figure 1). Plates were incubated for 24 Contreet ± hours at 37 C. A metric ruler was used to Acticoat ± Bactigras ± measure the diameter of the zone of inhibi- Aquacel Ag ± tion for each disc used (Kirby-Bauer Inadine ± method). MelDra Textus ± ± < Statistical Analysis SPSS for Windows (SPSS, Inc., Chicago, IL) Ultec ± 0 -- software was used for data management and statistical analysis. The zone of inhibition diameter from each dressing disc was compared using a paired sample test. Each dressing material was analyzed using a chi-squared test in regards to efficacy against the different microorganisms. The level of significance was set at 0.05 for all statistical tests. Results The present study included 9 gram-negative, 7 grampositive bacteria, and 2 yeast isolates. An inhibition zone PLICA AT DUPL PLI LIC CATE was not detected for Textus and Ultec (control dressing). Therefore, these materials were excluded from the statistical evaluation and regarded as most ineffective against microorganisms. The results for the size of the zones of clearance around the discs for all microorganisms on MH agar are shown in Table 2. Comparisons of the zones of inhibition are shown in Table 3. Among the 16 bacterial and 2 yeast species grown on MH agar plates, Contreet was the most effective antimicrobial dressing tested (P < 0.001). Textus and Ultec were found to be ineffective. The bactericidal activities of Vol. 22, No. 7 July

4 Figure 2. Efficacies of wound dressings against gram-positive bacteria. Figure 3. Efficacies of wound dressings against gram-negative bacteria. Acticoat, Aquacel Ag, and Bactigras were similar and more effective than Inadine and MelDra (P < 0.05). MelDra had the most variable bactericidal activity. It was interesting that the bactericidal activity of MelDra was more rapid and pronounced against MRSA than it was against MSSA. A statistical difference was not found among any of the dressings concerning efficacy against gram-positive and gram-negative bacteria, except for Aquacel Ag and Inadine. Both Aquacel Ag and Inadine were effective against both gram-positive and gram-negative microorganisms, yet their antibacterial activity was found to be 168 WOUNDS higher on gram-positive bacteria compared to gram-negative bacteria (P = 0.029, P = 0.030; [Figures 2 and 3]). Discussion Intense bacterial colonization is commonly associated with acute and chronic wounds. Several factors determine whether wounds colonized with microorganisms will become infected. These factors include the amount of bacteria per gram of tissue, species of microorganism, and the host immune response. Colonization can be defined d as the presence of replicating microorganisms in the wound that does not damage tissue. Colonization alone does not delay the wound healing process. Wound infection is characterized by an invasion of microorganisms into the tissue (more than 10 5 organisms per gram of tissue) and has clinical signs and symptoms such as hyperemia, fever, pain, and cellulitis. An intermediate stage, critical colonization, is increasingly being recognized as part of the wound infection process. This is a relatively new UPc description and refers to a transitional state between colonization and invasive wound infection. In this situation, granulation tissue is locally infected without any clinical signs of infection. Both wound infection and critical colonization can interfere with the healing process. Therefore, preventing the progression from colonization to infection is a major concern in wound care. 7 The primary treatment of a wound infection is copious irrigation with normal saline, drainage, debridement, and appropriate antibiotic therapy. Topical antimicrobial agents such as silver, povidone iodine, and chlorhexidine have been widely used in the presence of local infection (critical colonization) after meticulous debridement of all non-viable tissue. Bacterial biofilm formation is especially problematic since it is associated with resistance to host defenses and renders microorganisms more resistant to topical antimicrobials. Surgical removal of the biofilm layer is essential to facilitate the effects of topical antimicrobials and to eliminate local infection. More recently, several wound dressing materials with varying antimicrobial properties have become available for clinical use. The considerable variations in the structure, composition, and antimicrobial content of these contemporary dressings make the selection of an appropriate DUPL UPLICATE

5 Table 4. P values of each compared dressing material. material had strong bacterial inhibitive properties and Bactigras Inadine MelDra Acticoat Aquacel Contreet reduced wound infection. Ag Heggers et al 3 compared the efficacy of Acticoat, Silvasorb Bactigras < Inadine < (Medline, Mundelein, IL), and MelDra < Silverlon (Argentum Medical, Acticoat < Chicago, IL) against S aureus Aquacel Ag - < and P aeruginosa infections. Contreet - Heggers et al found that these The paired sample test was used to compare the measurements. The level of signifi- silver dressings were equally cance was set at 0.05 for all statistical tests. effective in reducing the microbial colonization in wounds. 3 TEHegg TEsilv dressing more difficult. Numerous studies have evaluated the advantages of using these modern dressings However, a comprehensive study has yet to clearly identify which antimicrobial dressing materials are superior. In the present study, inhibition zone sizes were used to compare the antimicrobial efficacy of various dressing preparations. Inhibition zone sizes in this in-vitro test are In the present study, Acticoat had the second most obvious inhibition zone. However, when it was compared with the Aquacel Ag and Bactigras, a statistically significant difference was not found (P > 0.05). Aquacel Ag is a silver-impregnated dressing with hydrofiber (sodium carboxymethylcellulose) and con- tains 21 µg ionic silver per cm This dressing material dependent upon two factors: the ability of the antimi- i- had a mean inhibition zone of 9.61 mm ± 4.15 mm crobial agent to diffuse into the agar, and the antimicro- among the tested microorganisms in the present study. bial activity of the agent against the microorganism. m. The Although Aquacel Ag was proven to have a moderate diffusion kinetics of the tested antimicrobial agents is not antibacterial activity against the tested microorganisms, the same in the agar gel this might be regarded lim-d d as a limitation of the study design. However, the diffusion kinet- dressing materials. Antibacterial activity of Aquacel Ag this activity was not the strongest among the tested DUAlt ics are also not equal in vivo on actual wounds among these agents. To standardize the study design, all of the dressings were tested in the same agar plate. Contreet is a newer-generation silver-based dressing that contains 13 µg silver per cm 2 that provides sustained was higher for gram-positive bacteria compared to gramnegative bacteria. The in-vitro antimicrobial efficacy of Textus silver dressing was found to be inefficient. Textus did not have a remarkable inhibition zone for any of the microorganisms tested. release of ionic silver to exert an antibacterial effect within the wound. 11 Ip et al 12 demonstrated that Contreet Bactigras is a dressing made of cotton impregnated provided maximal bactericidal al activity for Enterococcus with 0.5 % chlorhexidine acetate. Chlorhexidine is faecalis, Enterobacter cloacae,, Proteus vulgaris, P known as a persistent antimicrobial agent that is active aeruginosa, and Acinetobacter baumanii. Castellano et against a wide range of gram-positive and gram-negative al 13 reported similar findings ind in 2007 when they compared eight silver containing dressings to determine Bactigras was as effective as some of the silver-based bacteria. In the present study it was observed that Pr their in-vitro antimicrobial efficacy. Their findings suggest that Contreet was the most effective dressing. In the present study when Contreet was compared with other dressing materials, it displayed a superior antimicrobial activity on all microorganisms tested (P < 0.001). Significant differences were not detected concerning the bactericidal activities of Contreet against gram-negative and gram-positive bacteria. Acticoat is another silver-containing wound dressing that utilizes nanotechnology to release nanocrystalline dressings (Acticoat and Aquacel Ag). A significant difference was not found in terms of its effectiveness against gram-negative or gram-positive bacteria. Inadine uses povidone-iodine as its antimicrobial agent. Povidone-iodine has been demonstrated to be effective at killing a wide spectrum of microorganisms generally associated with wound infection. In the present study, although it demonstrated potential to reduce microorganism growth, it was unsatisfactory compared to the silver impregnated dressings. silver crystals. 12 Many studies have reported that this MelDra contains medical honey and had the most vari- antiban DUPLICATE Vol. 22, No. 7 July

6 able bactericidal activity. Interestingly, its killing effect was more evident against MRSA than MSSA. Conclusion In-vitro data suggest that Contreet is the most effective dressing for the topical treatment of contaminated wounds. In the present study, a diffusion technique was used because it is known that antimicrobial agents within dressings exert their effect through direct contact with the microorganisms via diffusion. The diffusion kinetics of the tested antimicrobial agents is not the same in the agar gel, which might be regarded as a limitation of the study design. However, the diffusion kinetics among these agents is also not equal to in vivo clinical wounds. To standardize the study design, all of the dressings were tested in the same agar plate. However, further methods of assessment, including the use of infected animal models and clinical studies, will be necessary to better understand the antimicrobial efficacies of these dressings. References 1. Galiano RD, Mustoe TA. Wound Care. In: Aston SJ, Beasley RW, Thorne CH, Grabb WC, Smith JW, eds. Grabb & Smith s Plastic Surgery. Philadelphia, PA: Lippincott incott Williams & Wilkins; 2007: Stotts NA, Hunt TK. Pressure ulcers. Managing aging bacterial colonization and infection. Clin Geriatr Med. 1997;13(3): Heggers J, Goodheart RE, Washington J, et al. Therapeutic efficacy of three silver dressings s in an infected animal model. J Burn Care Rehabil. 2005;26(1): (1):5 4. Bibbo C, Patel DV, Gehrmann n RM, Lin SS. Chlorhexidine provides superior skin decontamination in foot and ankle surgery: a prospective pective randomized study. Clin Orthop Relat Res. 2005;438: ;438: Maeda Y, Loughrey A, Earle JA,D JA, et al. Antibacterial activity of honey against community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA). Complement Ther Clin Pract. 2008;14(2): George NM, Cutting KF. Antibacterial honey (Medihoney ): in-vitro activity against clinical isolates of MRSA, VRE, and other multiresistant gram-negative organisms including Pseudomonas aeruginosa. WOUNDS. 2007;19(9): Edwards R, Harding KG. Bacteria and wound healing. Curr Opin Infect Dis. 2004;17(2): Boateng JS, Matthews KH, Stevens HN, Eccleston GM. Wound healing dressings and drug delivery systems: a 170 WOUNDS 26(1):5N incottd review. J Pharm Sci. 2008;97(8): Basterzi Y, Bagdatoglu C, Sari A, Demirkan F. Aplasia cutis congenita of the scalp and calvarium: conservative wound management with novel wound dressing materials. J Craniofac Surg. 2007;18(2): Ulkür E,, Oncul O, Karagoz H, Yeniz E, Celiköz B. Comparison of silver-coated dressing (Acticoat), chlorhexidine acetate 0.5% (Bactigras), and fusidic acid 2% (Fucidin) for topical antibacterial effect in methicillin-resistant Staphylococci-contaminated, full-skin thickness rat burn wounds. Burns. 2005;31(7): Burd A, Kwok CH, Hung SC, et al. A comparative study of the cytotoxicity of silver-based dressings in monolayer cell, tissue explant, and animal models. Wound Repair Regen ;15(1): Ip M, Lui SL, Poon VK, Lung I, Burd A. Antimicrobial activ- ities of silver dressings: an in-vitro comparison. J Med Microbiol. 2006;55(Pt 1): Castellano JJ, Shafii SM, Ko F, et al. Comparative evaluation of silver containing antimicrobial dressings and drugs. Int UPW Wound J. 2007;4(2): DUPL PLICAT ATE