Acellular Dermal Matrix in Plastic and Reconstructive Surgery.

Despite significant advances in medical research, plastic surgeons still face a shortage of suitable patient tissues, and soft tissue reconstruction is no exception. In recent years, there has been a rapid boom in the use of acellular dermal matrix (ADM) in reconstructive and aesthetic surgery. ADM is incorporated into the surrounding tissue and gradually replaced by the host's collagen, thus promoting and supporting the healing process and reducing the formation of scar tissue. The main goal of this article is to provide a brief review of the current literature assessing the clinical applications of ADM across a broad spectrum of applications in plastic and reconstructive surgery.

Preparation and processing of human allogenic dermal matrix for utilization in reconstructive surgical procedures.

AIM: This article presents the development of a novel preparation and processing method as well as indication for clinical applications of human allogeneic acellular dermal matrix, which was developed originally in the Central Tissue Bank (CTB) for use in burn medicine and reconstructive surgery. METHODS: Acellular dermal matrix (ADM) is a biological material assigned for utilization in several surgical procedures due to its unique structure and advantageous properties. The article focuses on a novel preparation and processing method developed by CTB, which differs in its impact on the structure, biological and biomechanical properties of the final ADM compared to the wide range of commercially available ADM products and currently available ADM products of other tissue banks. RESULTS: The ubiquitous acellular allogeneic dermal collagen matrix is the main substance participating in advantageous properties facilitating the use of ADM in numerous indications from dermal replacement and soft tissue augmentation to more extensive surgical reconstructive procedures. Dermal substitutes play an essential role in the reconstruction of full-thickness skin defects, both in acute and chronic wounds, defects of fasciae, peritoneum, etc., and there is a strong evidence that they can improve the final scar quality as well. Differences in preparation methods of ADMs are recently causing concerns among surgeons utilizing the ADMs. We present three different cases with favourable outcomes by using human acellular ADM grafts. CONCLUSIONS: Although ADMs did not fulfil all of the requirements for an ideal dermal substitute, their applications have been advanced for diverse indications in soft tissue reconstructions and augmentations. Early revascularization of the allografts reduces bacterial contamination. Research and development of new generation of acellular dermal matrices with incorporated autologous in vitro cultured cells will likely yield new products and give new hope for continued improve-ments in functional and cosmetic outcomes (Fig. 9, Ref. 60).

Correction: Brain structure in pediatric Tourette syndrome.

In this published article, members of 'The Tourette Association of America Neuroimaging Consortium' were not cited in PubMed. These consortium members are listed in the associated correction.

Adipose derived mesenchymal stem cells harvesting.

BACKGROUND: The lipografting is increasingly used in the field of plastic surgery. Widely used harvesting technique of fatderived stem-cells is lipoaspiration. There exist two big streams of fat harvesting for lipografting: mechanical liposuction and manual liposuction. METHODS: Two harvested specimens were compared in this prospective blind study in the means of stem-cells viability and their ability to grow in cell-cultures. Techniques to compare were: manual lipoaspiration with 50 ml syringe and WAL (water-jet assisted liposuction). RESULTS: Twenty specimens from ten patients were investigated in the tissue bank. There were no differences in the amount of live stem-cells between two groups. Also no differences were found between both harvesting techniques in the mean of cell ability to grow in cell-cultures. CONCLUSION: It can be concluded that there are no statistically significant differences in the number, vitality and viability of stem cells when comparing two ways of mesenchymal stem cell collection, both manual and machine sampling (WAL). When cultured in vitro, both samples collected from each patient also appeared to be able to multiply with no statistical differences (Tab. 2, Fig. 2, Ref. 18).

Toxic epidermal necrolysis data from the CELESTE multinational registry. Part I: Epidemiology and general microbiological characteristics.

INTRODUCTION: Toxic epidermal necrolysis (TEN) is a rare, life-threatening autoimmune disease predominantly manifested in the skin and mucous membranes. Today, infectious complications have the dominant share in mortality of TEN patients. Due to the nature of the therapy and administration of immunosuppressive medications, a wide range of potentially pathogenic microorganisms, which cause infectious complications in different compartments in these patients, is not surprising. MATERIAL AND METHODOLOGY: This is a multicentric study, which included all patients with TEN hospitalized between 2000-2015 in specialized centres in the Czech Republic and Slovakia. The total catchment area was over 12.5 million inhabitants. The actual implementation of the project was carried out using data obtained from the registry CELESTE (Central European LyEll Syndrome: Therapeutic Evaluation), when specific parameters relating to epidemiological indicators and infectious complications in patients with TEN were evaluated in the form of a retrospective analysis. RESULTS: In total, 39 patients with TEN were included in the study (12 patients died, mortality was 31%), who were hospitalized in the monitored period. The median age of patients in the group was 63 years (the range was 4-83 years, the mean was 51 years), the median of the exfoliated area was 70% TBSA (total body surface area) (range 30-100%, mean 67%). SCORTEN was calculated for 38 patients on the day of admission. Its median in all patients was 3 (range 1-6; mean 3). Any kind of infectious complication in the study group was recorded in 33 patients in total (85%). In total, 30 patients (77%) were infected with gram-positive cocci, 27 patients (69%) with gram-negative rods, and yeast cells or fibrous sponge were cultivated in 12 patients (31%). A total of 32 patients (82%) were found to have infectious complications in the exfoliated area, 15 patients (39%) had lower respiratory tract infections, 18 patients (46%) urinary tract infections and 15 patients (39%) an infection in the bloodstream. The most common potentially pathogenic microorganism isolated in our study group was coagulase neg. Staphylococcus, which caused infectious complications in 24 patients. Enterococcus faecalis/faecium (19 patients), Pseudomonas aeruginosa (17 patients), Staphylococcus aureus (11 patients) and Escherichia coli (11 patients) were other most frequently isolated micro-organisms. CONCLUSION: The published data were obtained from the unique registry of TEN patients in Central Europe. In the first part, we have succeeded in defining the basic epidemiological indicators in the group of patients anonymously included in this registry. The study clearly confirms that infectious complications currently play an essential role in TEN patients, often limiting the chances of survival. The study also shows a high prevalence of these complications in the period after 15days from the start of hospitalization, when most patients already have completely regenerated skin cover.

Toxic epidermal necrolysis data from the CELESTE multinational registry. Part II: Specific systemic and local risk factors for the development of infectious complications.

The aim of the study was to identify the most important systemic and local risk factors for the development of infectious complications in patients with toxic epidermal necrolysis (TEN). MATERIAL AND METHODOLOGY: This is a multicentric study that included all patients with TEN who were hospitalized between 2000-2015 in specialized centres in the Czech Republic and Slovakia. The total catchment area included a population of over 12.5 million inhabitants. The actual implementation of the project was carried out using data obtained from the CELESTE (Central European LyEll Syndrome: Therapeutic Evaluation) registry, wherein specific parameters related to epidemiological indicators and infectious complications in patients with TEN were evaluated as a retrospective analysis. RESULTS: A total of 38 patients (97%) of the group were treated with corticosteroids. The comparison of patients with different doses of corticosteroids did not exhibit a statistically significant effect of corticosteroid administration on the development of infectious complications (p=0.421). There was no effect of the extent of the exfoliated area on the development of infectious complications in this area. The average extent of the exfoliated area was 66% TBSA (total body surface area) in patients with reported infectious complications and 71% TBSA (p=0.675) in patients without infectious complications. In the case of the development of an infectious complication in the bloodstream (BSI), the increasing effect of the SCORTEN (SCORe of Toxic Epidermal Necrosis) value was monitored during hospitalization. Within 5days from the beginning of the hospitalization, the average SCORTEN value was 2.7 in 6 patients with BSI and 3.0 in 32 patients without BSI (p=0.588). In the period after the 15th day of hospitalization, 7 patients with BSI had an average SCORTEN value of 3.4, and 16 patients without BSI had an average SCORTEN value of 2.5 (p=0.079). In the case of low respiratory tract infection (LRTI), the effects of the necessity for artificial pulmonary ventilation and the presence of tracheostomy were monitored. The statistically significant effect of mechanical ventilation on the development of LRTI occurred only during the period of 11-15days from the beginning of the hospitalization (p=0.016). The effect of the tracheostomy on the development of LRTI was proven to be more significant. CONCLUSION: We did not find any statistically significant correlation between the nature of immunosuppressive therapy and the risk of developing infectious complications. We failed to identify statistically significant risk factors for the development of BSI. Mechanical ventilation and tracheostomy increase the likelihood of developing LRTIs in patients with TEN.

Development of a new method for the preparation of an acellular allodermis, quality control and cytotoxicity testing.

Demand for use of acellular allodermis is high but commercially appropriate products are not used routinely because of very high price and limited availability. These facts did motivate us to prepare acellular allodermis using a new, simple and less expensive method. We have developed a original method for preparation of acellular allogeneic dermis based on action of a proteolytic enzyme in combination with distilled water. Hypotonic environment in comparison with SDS or Triton ansure no toxicity of the final product. Trials for determination of optimal trypsin concentrations, temperature and time of action were performed. According to our results, the use of 2.5% trypsin/EDTA solution overnight at +4 °C was proving to be optimal. The histology confirmed absence of cells in the prepared dermis. No toxicity of final acellular dermis was confirmed by three independent tests (agar diffusion test contact cytotoxicity test and grow curve). The prepared acellular dermis seems to be suitable not only for direct clinical use, but it can be used as a scaffold for cell cultivation as well.

Cytotoxicity testing of burn wound dressings: first results.

Topical antimicrobial therapy represents an essential part of burn wound care. In order to prevent and treat burn wound infection dressings with antimicrobial properties are applied directly on the wound surface. Not only the infection control but also promotion of healing is very important in burn wound management. It is well known, that a dressing in bactericidal concentration might also delay wound healing. This study was aimed to evaluate the potential toxic effect of topical antimicrobial agents on murine and human dermal cells. For toxicity testing the method by Vittekova et al. was used to evaluate potential toxic effects of 16 agents and 6 control samples on two in vitro cultured cell systems [3T3 cells and dermal fibroblasts] during the first 24 h. Following the 24 h cell culture with the tested agents the live cell counts were evaluated. According to results obtained on both cell systems, the tested samples were divided into three groups-nontoxic, semi-toxic and toxic. Nontoxic samples included Acetic acid 1%, Acticoat®, Dermacyn®, Framykoin®, Silverlon®, gauze, acellular human allodermis and acellular porcine xenodermis. Semi-toxic group included Algivon®Plus, Aquacel®Ag, Betadine®, Nitrofurazone, Octenisept®, Suprasorb® A and a porcine dermal scaffold Xeno-Impl. Finally, the toxic group included Algivon®, Dermazin®, Ialugen®Plus, Prontoderm®, Suprasorb® A Ag and 20% SDS. As the preliminary results of this study have shown, our findings may serve as a potential guide to selection of the most appropriate topical antimicrobial dressings for treatmet of burns. However before they can be translated into clinical practice recommendations, more research on antimicrobial dressings cytotoxicity testing will be necessary.

Impact of Sagittal Balance on Clinical Outcomes in Surgically Treated T12 and L1 Burst Fractures: Analysis of Long-Term Outcomes after Posterior-Only and Combined Posteroanterior Treatment.

Objective. Long-term radiological and clinical outcome retrospective study of surgical treatment for T12 and L1 burst fractures in perspective of sagittal balance measures. Methods. Patients with age of 16-60 years, complete radiographs, early surgical treatment surgery, and follow-up (F/U) > 18 months were included and strict exclusion criteria applied. Regional and thoracolumbar kyphosis angles (RKA and TLA) were measured preoperatively and at final F/U, as were parameters of the spinopelvic sagittal alignment. Clinical outcomes were assessed using validated measures. Results. 36 patients with age mean age of 39 years and F/U of 69 months were included. 61% of patients were treated with bisegmental posterior instrumentation (POST-I) and 39% with combined posteroanterior instrumented fusion (PA-F). At F/U, several indicators for clinical outcomes showed a significant correlation with radiographic measures in the overall cohort with inferior clinical outcomes corresponding with increasing residual deformity and sagittal malalignment. Statistical analysis failed to reach level of significance for the differences between POST-I and PA-F group at final F/U. Only a strong trend towards better restoration of the thoracolumbar alignment was observed for the PA-F group in terms of the RKA and TLA. Conclusions. Results in a surgically treated cohort of T12 and L1 burst fracture patients indicate that superior clinical outcomes depend on restoration of sagittal alignment.

COMPLICATIONS OF LOWER EXTREMITY HEMATOMAS IN PATIENTS WITH PRE-INJURY WARFARINE USE.

INTRODUCTION: The aim of this paper is to ascertain the number of patients with pre-injury warfarin use who developed lower extremity hematomas treated in our facility, to analyse the data, used treatment methods and outcomes in these patients. PATIENTS AND METHODS: We performed a retrospective review, identifying all the patients with pre-injury Warfarin use admitted with hematoma or full-thickness skin loss in the ten years period from January 2006 to December 2015. RESULTS: Overall 9 women and 2 men with mean age of 72 years were identified. All the injuries were sustained in a domestic setting. Except of one female patient primarily admitted to our department, all patients had been hospitalized primarily in local/regional hospitals for an average period of 32.6 days. All the patients transferred to our department required surgical wound closure. The mean wound surface area was 136.3cm² (range 45-525). The duration of hospital stay was 15 days in average. DISCUSSION: The data obtained were compared with the results and findings of similar studies and were discussed. CONCLUSION: The results achieved in the present study showed the beneficial effect of used treatment methods based on the surgical wound closure techniques during hospital stay of the patients. Clinicians, first contact physicians, and also patients alone need to be aware of the vulnerability of this group of patients. The consequences of even minor lower extremity trauma can be serious, with development of a very complex chronic wound that is difficult to manage.

Brain structure in pediatric Tourette syndrome.

Previous studies of brain structure in Tourette syndrome (TS) have produced mixed results, and most had modest sample sizes. In the present multicenter study, we used structural magnetic resonance imaging (MRI) to compare 103 children and adolescents with TS to a well-matched group of 103 children without tics. We applied voxel-based morphometry methods to test gray matter (GM) and white matter (WM) volume differences between diagnostic groups, accounting for MRI scanner and sequence, age, sex and total GM+WM volume. The TS group demonstrated lower WM volume bilaterally in orbital and medial prefrontal cortex, and greater GM volume in posterior thalamus, hypothalamus and midbrain. These results demonstrate evidence for abnormal brain structure in children and youth with TS, consistent with and extending previous findings, and they point to new target regions and avenues of study in TS. For example, as orbital cortex is reciprocally connected with hypothalamus, structural abnormalities in these regions may relate to abnormal decision making, reinforcement learning or somatic processing in TS.

Comparison of structural changes in skin and amnion tissue grafts for transplantation induced by gamma and electron beam irradiation for sterilization.

Sterilization is an important step in the preparation of biological material for transplantation. The aim of the study is to compare morphological changes in three types of biological tissues induced by different doses of gamma and electron beam radiation. Frozen biological tissues (porcine skin xenografts, human skin allografts and human amnion) were irradiated with different doses of gamma rays (12.5, 25, 35, 50 kGy) and electron beam (15, 25, 50 kGy). Not irradiated specimens served as controls. The tissue samples were then thawn and fixed in 10 % formalin, processed by routine paraffin technique and stained with hematoxylin and eosin, alcian blue at pH 2.5, orcein, periodic acid Schiff reaction, phosphotungstic acid hematoxylin, Sirius red and silver impregnation. The staining with hematoxylin and eosin showed vacuolar cytoplasmic changes of epidermal cells mainly in the samples of xenografts irradiated by the lowest doses of gamma and electron beam radiation. The staining with orcein revealed damage of fine elastic fibers in the xenograft dermis at the dose of 25 kGy of both radiation types. Disintegration of epithelial basement membrane, especially in the xenografts, was induced by the dose of 15 kGy of electron beam radiation. The silver impregnation disclosed nuclear chromatin condensation mainly in human amnion at the lowest doses of both radiation types and disintegration of the fine collagen fibers in the papillary dermis induced by the lowest dose of electron beam and by the higher doses of gamma radiation. Irradiation by both, gamma rays and the electron beam, causes similar changes on cells and extracellular matrix, with significant damage of the basement membrane and of the fine and elastic and collagen fibers in the papillary dermis, the last caused already by low dose electron beam radiation.

[AMNIOTIC MEMBRANE APPLICATIONS - OUR EXPERIENCE]. / MOZNOSTI POUZITIA AMNIOVEJ MEMBRÁNY - NASE SKÚSENOSTI.

INTRODUCTION: Amniotic membrane is the innermost part of the fetal and packaging for its exceptional qualities likes to be used in treating many ocular pathologies. Amniotic membrane has improved the ability to treat ocular surface disease. It has unique features like support conjunctival and corneal epithelialization. MATERIAL AND METHODS: Retrospective analysis of group patients who underwent amniotic membrane transplantation at the Department of Ophthalmology Faculty of Medicine and UN Bratislava in 2013-2015. We evaluated indications amniotic membrane transplantation, the percentage, the number of transplants and the number of failures and retransplantation of the membrane. RESULTS: In group of 71 patients (amniotic membrane covering defects of conjunctiva and cornea) male patients formed a slight predominance of males in the number of patients a slightly larger preponderance in 38 women (53.5 %) - 52 surgeries (59.09 %) and 33 male (46.5 %) in 36 interventions (40.91 %). The left eye was affected in 40 interventions (45.45 %), 48 interventions were on the right eye (54.54 %). The most common cause application of 30.68 % in 27 eyes was corneal ulcer, bullous keratopathy followed by the 11.36 % in 10 eyes, and the ulcer herpetic keratitis in 9.10 % in 8 eyes. Injury or vulnus penetrans 6.82 % in 6 eyes, ulcers caused by paresis n. facialis 6.82 % in 6 eyes and sicca syndrome 5.68 % in 5 eyes.In 2015 we applied amniotic membrane covering the defect of eyelids after trauma in one patient. CONCLUSION: Amniotic membrane is the appropriate treatment in a number of diseases of ocular surface when conservative methods of treatment fail. In corneal application can prevent the execution of more aggressive treatment, such as keratoplasty, or to soothe inflammation and keratoplasty is not performed as emergent, but elective.Key words: amniotic membrane transplantation, eye diseases.

Minimally invasive burn care: a review of seven clinical studies of rapid and selective debridement using a bromelain-based debriding enzyme (Nexobrid®).

Current surgical and non-surgical eschar removal-debridement techniques are invasive or ineffective. A bromelainbased rapid and selective enzymatic debriding agent was developed to overcome these disadvantages and compared with the standard of care (SOC). The safety and efficacy of a novel Debriding Gel Dressing (DGD) was determined in patients with deep partial and full thickness burns covering up to 67% total body surface area (TBSA). This review summarizes data from seven studies, four of which were randomized clinical trials that included a SOC or control vehicle. DGD eschar debridement efficacy was >90% in all studies, comparable to the SOC and significantly greater than the control vehicle. The total area excised was less in patients treated with DGD compared with the control vehicle (22.9% vs. 73.2%, P<0.001) or the surgical/non-surgical SOC (50.5%, P=0.006). The incidence of surgical debridement in patients treated with DGD was lower than the SOC (40/163 [24.5%] vs. 119/170 [70.0%], P0.001). Less autografting was used in all studies. Long-term scar quality and function were similar in DGD- and SOCtreated. DGD is a safe and effective method of burn debridement that offers an alternative to surgical and non-surgical SOC.

Les protocoles actuels de détersion d'une brûlure, chirurgicaux et non chirurgicaux, sont soit invasifs soit inefficaces. Un enzyme détersif rapide et spécifique, dérivé de la bromélaïne, a été développé dans le but de palier à ces 2 inconvénients. Il a été comparé aux techniques usuelles (TU). L'efficacité et l'innocuité d'un Gel Topique Détersif (GTD) ont été évaluées chez des patients souffrant de brûlures intermédiaires et profondes atteignant jusqu'à 67% de la Surface Corporelle Totale (SCT). Cette revue compile les données de 7 études cliniques, dont 4, randomisées, faisaient appel aux TU ou à un groupe contrôle. La détersion obtenue avec GTD était toujours > 90%, comparable aux TU et meilleure que dans le groupe contrôle. La surface relative excisée totale était moindre après GTD que chez les contrôles (22.9% VS 73.2%, p<0,001) ou les patients sous TU (50.5%, p=0,006). Le nombre de patients ayant eu besoin de chirurgie a été inférieur dans le groupe GTD que dans le groupe TU (40/163 [24.5%] VS 119/170 [70%], p<0,001). Le recours aux greffes était moins fréquent dans toutes les études. Les qualités cicatricielle et fonctionnelle à distance étaient comparables après TU et GTD. GTD est une technique de détersion efficace et sûre qui offre une alternative au TU, chirurgical ou non.

Structural changes of skin and amnion grafts for transplantation purposes following different doses of irradiation.

An important part of the preparation of biological material for transplantation is sterilization. The aim of our study was to assess the impact of ionizing radiation on three types of biological tissues and the impact of different doses on cells and extracellular matrix. Three types of frozen tissues (porcine skin xenografts, human skin allografts and human amnion) were divided into five groups, control and groups according to the dose of radiation to which these samples were exposed (12.5, 25, 35 and 50 kGy). The tissue samples were fixed by formalin, processed by routine paraffin technique and stained with hematoxylin and eosin, alcian blue at pH 2.5, orcein, periodic acid schiff reaction and silver impregnation. The staining with hematoxylin and eosin showed hydropic degeneration of the cells of epidermis in xenografts by the dose of 12.5 kGy, in human skin it was observed by the dose of 35 kGy. The staining for elastic fibers revealed damage of fine elastic fibers in the xenografts dermis by the dose of 12.5 kGy, in the allografts by 35 kGy. Another change was the disintegration of basement membrane of epithelium, especially in the human amnion at the dose of 50 kGy. The silver impregnation visualized nuclear chromatin condensation mainly in human amnion at the dose of 12.5 kGy. Our results have shown that the porcine xenografts and human amnion were more sensitive to irradiation than the human skin. In the next phase of the project we will focus at more detailed changes in the tissues using immunohistochemical techniques.

[Surgical therapy of wounds and scars]. / Chirurgische Therapie von Wunden und Narben.

Surgery can be very useful in all phases of wound healing and in treating scars. It can be employed along with conservative therapy, either simultaneously or in a sequential manner. The spectrum of surgical measures includes debridement with excision of wound edges or necrotic areas, skin transplantation to cover granulating wounds, and simple excision or complex reconstructive approaches to scars. The advantages of surgical treatment are seen in all phases of wound healing and include rapid onset of action, avoidance of secondary infection and improved function. The shortened healing time helps save personal and material costs.

Hypoglycaemia-induced changes in regional brain volume and memory function.

BACKGROUND: Hypoglycaemic events can be a serious complication of insulin therapy in Type 1 diabetes mellitus. Severe hypoglycaemic exposure can lead to episodic memory impairments, including anterograde amnesia. However, relatively little is known regarding the long-term impact of severe hypoglycaemia on brain structure in Type 1 diabetes mellitus. The goals of the present study were to gain a greater understanding of the long-term effects of severe hypoglycaemia exposure on brain structure and the neural correlates of memory impairments in Type 1 diabetes mellitus. CASE REPORT: Regional grey and white matter volume and total white matter lesion volume were quantified in an individual with long-standing hypoglycaemia-induced anterograde amnesia and compared with age- and gender-matched healthy controls. Our patient has significant reductions in grey matter volume in the hippocampus, thalamus and pallidum, and significant reductions in white matter volume in the splenium, isthmus of the cingulate and cerebellum. He also has a significantly larger total white matter lesion volume than controls. CONCLUSION: This case study highlights the potential of hypoglycaemia for permanent deleterious effects on brain structure and memory function. Our results suggest that subcortical grey matter, periventricular white matter and posterior white matter may be most susceptible to injury from hypoglycaemia exposure, and that structural damage to the hippocampus and isthmus of the cingulate may play a central role in hypoglycaemia-induced memory impairments.

Skin explant cultures as a source of keratinocytes for cultivation.

Cultivated human keratinocytes can be used successfully in the treatment of burn patients, but efforts to heal burns and other wounds can be hampered by the very small skin biopsies available for cultivation of transplantable keratinocyte sheets. A small biopsy (and correspondingly small number of enzymatically isolated keratinocytes for use in classical cultivation techniques) can lead to a low yield of multilayer sheets for clinical application or unacceptably long cultivation times. One way of addressing this is to make use of skin remnants remaining after enzymatic digestion and culture cells migrating out of these skin explants. Sufficient numbers of explant-derived keratinocytes can be obtained to facilitate additional routine cultivation of these cells. Biopsy remnants can be used to initiate explant cultures repeatedly (we were able to re-use pieces of skin 10 times and still obtain useful numbers of keratinocytes) and this "passaging" yields substantially more cells for classical cultivation than would be available from conventional methodology alone, and in a comparable timeframe. Another advantage of this method is that it does not require additional biopsies to be procured from already-compromised patients and overcomes problems associated with contamination of skin samples with resistant hospital-acquired bacterial infections common during prolonged hospitalization.