Lugol's solution and Gentian violet eradicate methicillin-resistant Staphylococcus aureus biofilm in skin wound infections.

The study aimed to evaluate the antibacterial efficacy of Lugol's solution 5% and Gentian violet 1% against methicillin-resistant Staphylococcus aureus (MRSA) biofilm in vivo. The bactericidal efficacy for treatment of MRSA-biofilm skin wound infection was tested in a murine model. Luciferase-tagged S. aureus Xen31, a MRSA-strain derived from S. aureus ATCC-3359130, was used for infection. Wounds were made in the skin of mice and infected with MRSA. The mice were treated with Lugol's solution and Gentian violet. Application of the antimicrobial agents started 24 hours post infection and was repeated daily for five-days. The antimicrobial effect on the biofilm bacteria was evaluated by measuring bioluminescence from MRSA daily for seven-days. Lugol's solution and Gentian violet showed a significant reduction in luminescent signals from the first assessment day to all subsequent days (P < .001). Lugol's solution and Gentian violet effectively eradicated MRSA in biofilm in vivo and could be alternatives or in addition to topical antibiotics when MRSA-biofilm wound infection is suspected.

From 3D printing to 3D bioprinting: the material properties of polymeric material and its derived bioink for achieving tissue specific architectures.

The application of 3D printing technologies fields for biological tissues, organs, and cells in the context of medical and biotechnology applications requires a significant amount of innovation in a narrow printability range. 3D bioprinting is one such way of addressing critical design challenges in tissue engineering. In a more general sense, 3D printing has become essential in customized implant designing, faithful reproduction of microenvironmental niches, sustainable development of implants, in the capacity to address issues of effective cellular integration, and long-term stability of the cellular constructs in tissue engineering. This review covers various aspects of 3D bioprinting, describes the current state-of-the-art solutions for all aforementioned critical issues, and includes various illustrative representations of technologies supporting the development of phases of 3D bioprinting. It also demonstrates several bio-inks and their properties crucial for being used for 3D printing applications. The review focus on bringing together different examples and current trends in tissue engineering applications, including bone, cartilage, muscles, neuron, skin, esophagus, trachea, tympanic membrane, cornea, blood vessel, immune system, and tumor models utilizing 3D printing technology and to provide an outlook of the future potentials and barriers.

Localization of progenitor/stem cells in the human tympanic membrane.

BACKGROUND: The outer epithelium of the tympanic membrane (TM) initiates the closure of a perforation. Embryonic stem cells have been used in attempts to enhance the healing capacity of induced perforations in experimental animals. More knowledge about epithelial cell proliferation and cell migration is needed for a better understanding of the TM healing process. This includes the identification of regenerative zones within the outer epithelial layer of the TM where progenitor cells may be present. METHODS: Normal human TMs from translabyrinthine surgery were investigated using immunohistochemistry and immunofluorescence to detect the progenitor/stem cell markers α6-integrin, ß1-integrin and cytokeratin 19 (CK19). RESULTS: α6-Integrin was detected in the basal layer of the keratinizing epithelium in the umbo, in the annular region and along the malleus but not in the intermediate portion of the pars tensa. ß1-Integrin and CK19 were found in the same locations not only in the basal layer but also in the suprabasal layers of the keratinizing epithelium. CONCLUSIONS: Possible progenitor cells are found in the umbo, the annular region and along the malleus. Further studies are needed to identify the source of these cells.

Bioactive glass S53P4 eradicates Staphylococcus aureus in biofilm/planktonic states in vitro.

Background: Increasing antimicrobial resistance to antibiotics is a substantial health threat. Bioactive glass S53P4 (BAG) has an antimicrobial effect that can reduce the use of antibiotics. The aim of this study was to evaluate the antimicrobial efficacy of BAG in vitro on staphylococci in biofilm and in planktonic form. Secondary aims were to investigate whether supernatant fluid primed from BAG retains the antibacterial capacity and if ciprofloxacin enhances the effect.Methods: BAG-S53P4 granules, <45 µm, primed in tryptic soy broth (TSB) were investigated with granules present in TSB (100 mg/mL) and after removal of granules (100, 200, and 400 mg/mL). The efficacy of BAG to eradicate Staphylococcus aureus biofilm in vitro was tested using 10 different clinical strains and 1 reference strain in three test systems: the biofilm-oriented antiseptic test based on metabolic activity, the biofilm bactericidal test based on culturing surviving bacteria, and confocal laser scanning microscopy (CLSM) combined with LIVE/DEAD staining.Results: Exposure to 48 h primed BAG granules (100 mg/mL) produced bactericidal effects in 11/11 strains (p = 0.001), and CLSM showed reduction of viable bacteria in biofilm (p = 0.001). Supernatant primed 14 days, 400 mg/mL, reduced metabolic activity (p < 0.001), showed bactericidal effects for 11/11 strains (p = 0.001), and CLSM showed fewer viable bacteria (p = 0.001). The supernatant primed for 48 h, or in concentrations lower than 400 mg/mL at 14 days, did not completely eradicate biofilm.Conclusion: Direct exposure to BAG granules, or primed supernatant fluid, effectively eradicated S. aureus in biofilm. The anti-biofilm effect is time- and concentration-dependent. When BAG had reached its full antimicrobial effect, ciprofloxacin had no additional effect.

A New, Promising Experimental Ossicular Prosthesis: A Human Temporal Bone Study With Laser Doppler Vibrometry.

OBJECTIVE: We compared the sound transmission using different types of total ossicular replacement prostheses (TORP); we then studied the performance of a new TORP that we designed inspired by the columella, the single ossicle found in birds. METHODS: Stapedial vibrations were measured on nine freshly frozen human temporal bones with laser Doppler vibrometry. We then compared the performances of eight common TORP positions or designs as well as the new silver prototype of bird-type prosthesis, designed also according to our digital holography patterns of the human tympanic membrane (TM). RESULTS: The TORPs placed in lateral contact with both the TM and the malleus handle outperformed, at most frequencies, those placed only in contact with the TM.The new bird-type prosthesis performed equally well or better than all other prostheses. CONCLUSION: If the malleus handle can be retained when placing a TORP, the best sound transmission can be achieved by placing the TORP in contact with both the distal part of the malleus handle and the TM. The good performance of our bird-type prosthesis suggests that there is still room for future improvement of prosthesis design to further optimize hearing outcomes after surgery.

Evaluation of Artificial Fixation of the Incus and Malleus With Minimally Invasive Intraoperative Laser Vibrometry (MIVIB) in a Temporal Bone Model.

BACKGROUND: A significant number of adults suffer from conductive hearing loss due to chronic otitis media, otosclerosis, or other pathologies. An objective measurement of ossicular mobility is needed to avoid unnecessarily invasive middle ear surgery and to improve hearing outcomes. METHODS: Minimally invasive intraoperative laser vibrometry provides a method that is compatible with middle ear surgery, where the tympanic membrane is elevated. The ossicles were driven by a floating mass transducer and their mobility was measured using a laser Doppler vibrometer. Utilising this method, we assessed both the absolute velocities of the umbo and incus long process as well as the incus-to-umbo velocity ratio during artificial fixation of the incus alone or incus and malleus together. RESULTS: The reduction of absolute velocities was 8 dB greater at the umbo and 17 dB at the incus long process for incus-malleus fixations when compared with incus fixation alone. Incus fixation alone resulted in no change to the incus-to-umbo velocity ratio where incus-malleus fixations reduced this ratio (-11 dB). The change in incus velocity was shown to be the most suitable parameter to distinguish between incus fixation and incus-malleus fixation. When the whole frequency range was analyzed, one could also differentiate these two fixations from previously published stapes fixation, where the higher frequencies were less affected. CONCLUSION: Minimally invasive intraoperative laser vibrometry provides a promising objective analysis of ossicular mobility that would be useful intraoperatively.

Functional effects of repeated pressure loads upon the tympanic membrane: mechanical stiffness measurements after simulated habitual sniffing.

In experimental studies it was found that otitis media causes stiffness loss in the tympanic membrane, possible precursors to retraction pockets and cholesteatoma. Besides otitis media habitual sniffing behaviour is associated with the development of retractions. The present study aims to test the hypothesis that repeated sniffing manoeuvre may cause not only structural, epithelial tympanic membrane changes presumed to be possible precursors to retractions, but also tympanic membrane stiffness loss, another possible mediator for the development of retractions. An experimental model with a pressure chamber was used to mimic the pressure conditions for the tympanic membrane in habitual sniffers' ears. The stiffness properties of twelve Mongolian gerbil tympanic membranes were measured with moiré interferometry after varying time up to 12 days with repeated pressure loading. Three days later, lower overall displacement were obtained in two ears; after 7-12 days the displacement readings were normal. This study with maximum of 12 days of pressure loading did not verify the hypothesis that habitual "sniffing" impairs the stiffness of the tympanic membrane.

Surgery for Chronic Otitis Media Causes Greater Taste Disturbance Than Surgery for Otosclerosis.

OBJECTIVES: Patients with otosclerosis more often complain about postoperative taste disturbance than patients with chronic otitis media, which seems paradoxical. We aim to investigate if and potentially why this seems to be the case, since the chorda tympani nerve (CTN) is thought to be severely traumatized less frequently during surgery in the former than in the latter. STUDY DESIGN: Prospective cohort study. SETTING: Department of Otorhinolaryngology at Hospital of Vastmanland, Vasteras, Sweden. PATIENTS: Sixty-five adults undergoing primary middle ear surgery were included. Thirty-seven were operated on for chronic suppurative otitis media with or without cholesteatoma (CSOM) and 28 for otosclerosis. INTERVENTIONS: Middle ear surgery due to otosclerosis or CSOM. Subjective and objective taste measurements and quality of life (QoL) questionnaire. MAIN OUTCOME MEASURES: Taste was assessed using electrogustometry (EGM) and the filter paper disc (FPD) method before and up to 1 year after surgery. Questionnaires on taste disturbance, including a visual analogue scale (VAS), and QoL were completed before and up to 1 year after surgery. RESULTS: Subjective taste disturbance anytime during the 1-year follow-up were reported by 62 and 46%, respectively. The difference in EGM 1 week after surgery compared with preoperative EGM was significantly greater among CSOM patients than otosclerosis. One year postoperatively, the difference is non-significant. CONCLUSION: Surgery for CSOM causes greater initial and more long-lasting taste disturbances as compared with surgery for otosclerosis. One-year postoperative taste normalizes for both CSOM and otosclerosis patients according to VAS and EGM measurements. No real change in QoL was seen 1-year postoperatively. LEVEL OF EVIDENCE: Level 2 evidence is prospective observational research with an experimental design.

Stem cells and enhanced healing of chronic tympanic membrane perforation.

CONCLUSIONS: Important information about the basic reparative process of tympanic membrane (TM) healing is shown, which can be incorporated for further clinical understanding. This provides a basis for the exploration of stem cell treatment for TM perforations and holds promise for future improvements. OBJECTIVES: This study aimed to analyse the healing of TM perforation by using stem cells and the stiffness of the membrane was tested in an acute and long-term study. MATERIALS AND METHODS: Sprague-Dawley rats were used in a model of TM perforation. The perforation was performed with a laser system. Stem cells were applied and the healing time and morphological analysis were performed with light and transmission electron microscope. Stiffness was examined by moiré interferometry. RESULTS: The stiffness of the perforated and healed TM was restored after just 2 weeks. In the chronic perforation model, mesenchymal stem cells enhanced the healing.

A randomized study of four different types of tympanostomy ventilation tubes - Full-term follow-up.

OBJECTIVE: To evaluate the impact of tympanostomy ventilation tube material (silicone vs fluoroplastic) and shape (short vs long) regarding time to extrusion, occurrence of otorrhea, occlusion, tube removal and occurrence of persistent perforation. METHODS AND MATERIAL: Four different types of ventilation tubes were used; Long Armstrong tubes, Donaldson tubes, Shepard tubes and straight tubes, representing four specific combinations of VT material (silicone or fluoroplastic) and shape (short, double flanged or long, single flanged). Four hundred children scheduled for bilateral tube insertion were included in a randomized trial. The patients received one type of tube in the right ear and another type in the left ear. The incidence of tube extrusion and complications were monitored postoperatively every third month by an otolaryngologist. RESULTS: Twenty-two children were excluded during surgery. Out of the studied 378 children the mean age was 35.3 months. 63.8% were boys. Short tubes extruded earlier than long tubes; hazard ratio (HR) 4.84 (95% CI 3.50-6.69, p < 0.001). Long Armstrong tubes were least prone to extrude. Silicone tubes resulted in significantly longer time to first infection in a VT ear, HR 1.68 (95% CI 1.03-2.76, p = 0.039). Donaldson tubes rendered the longest mean time to first infection (p = 0.025). Infections did not affect tube extrusion rates significantly (p = 0.879). No significant differences were found regarding tube occlusion, tube extraction or persistent perforation. CONCLUSIONS: Long tubes are less prone to extrude early. Long Armstrong tubes have the least propensity to extrude early. Silicone tubes render significantly longer time to first infection. Donaldson tubes result in least infections. Infection does not affect extrusion rates significantly. LEVEL OF EVIDENCE: 1b.

An Optimal Partial Ossicular Prosthesis Should Connect Both to the Tympanic Membrane and Malleus: A Temporal Bone Study Using Laser Doppler Vibrometry.

OBJECTIVE: To compare stapes vibrations in different partial ossicular replacement prosthesis (PORP) applications. METHODS: Stapedial vibrations were measured on fresh frozen human temporal bones with laser Doppler vibrometry. Eight different types of common ossiculoplasty methods were compared regarding recovery of stapes vibrations in relation with the normal ossicular chain. The PORPs were divided into three groups: 1) PORPs with the lateral contact only with the tympanic membrane, 2) PORPs with lateral contact only to the malleus handle, and 3) PORPs with lateral contact with both the malleus handle and the tympanic membrane. RESULTS: The PORPs with lateral contact only to the malleus handle performed better than the PORPs with lateral contact to the tympanic membrane only at 2 kHZ, but the best recovery was found in the group with contact both to the malleus handle and the tympanic membrane. CONCLUSION: The best sound transmission might be achieved by placing a PORP in contact with both the tympanic membrane and the handle of the malleus.

Tympanic Membrane Derived Stem Cell-Like Cultures for Tissue Regeneration.

Epidermal cells with stem cell-like characteristics have been identified in the tympanic membrane (TM) and localized specifically to the umbo and annulus regions. While they have been proposed to play a role in the regeneration of both acute and chronic TM perforations, evidence for the mechanism and regulation of their contribution is not yet described. Indeed, the behavior of these putative stem cells is largely unknown, in part due to a lack of refined methods for efficient cell isolation. In this study, we compared different explant techniques using normal and perforated rat TM tissues and investigated their ex vivo characteristics. TM after perforation in vivo showed increased staining for epidermal stem cell markers integrin ß1 and cytokeratin (CK) 19, and for proliferation Ki-67, indicating activation of the proliferative centers. A mixed population of fibroblasts and epithelial cells were isolated from explant cultures. Excised TM umbo implanted on a culture well insert was the most effective technique. Explants made from perforated TM produced cells before those from unperforated TM. More importantly, the implanted TM umbo organoid was capable of producing cells in a continuous manner, allowing subsequent harvest using trypsin. Primary rat TM epithelial cell cultures positive for pancytokeratin had colony forming activity and could be enriched for CK 19-positive cells that were capable of culture expansion by proliferation and cell migration when subject to a wound assay. Taken together, trauma to the TM activated the proliferative centers and prompted early cell production from TM umbo organoid cultures, which produced TM stem cell-like cultures that proved suitable for tissue engineering of the TM.

Minimally invasive laser vibrometry (MIVIB) with a floating mass transducer - A new method for objective evaluation of the middle ear demonstrated on stapes fixation.

Ossicular fixation through otosclerosis, chronic otitis media and other pathologies, especially tympanosclerosis, are treated by surgery if hearing aids fail as an alternative. However, the best hearing outcome is often based on knowledge of the degree and location of the fixation. Objective methods to quantify the degree and position of the fixation are largely lacking. Laser vibrometry is a known method to detect ossicular fixation but clinical applicability remains limited. A new method, minimally invasive laser vibrometry (MIVIB), is presented to quantify ossicle mobility using laser vibrometry measurement through the ear canal after elevating the tympanic membrane, thus making the method feasible in minimally invasive explorative surgery. A floating mass transducer provides a clinically relevant transducer to drive ossicular vibration. This device was attached to the manubrium and drove vibrations at the same angle as the longitudinal axis of the stapes and was therefore used to assess ossicular chain mobility in a fresh-frozen temporal bone model with and without stapes fixation. The ratio between the umbo and incus long process was shown to be useful in assessing stapes fixation. The incus-to-umbo velocity ratio decreased by 15 dB when comparing the unfixated situation to stapes fixation up to 2.5 kHz. Such quantification of ossicular fixation using the incus-to-umbo velocity ratio would allow quick and objective analysis of ossicular chain fixations which will assist the surgeon in surgical planning and optimize hearing outcomes.

Transcription and microRNA Profiling of Cultured Human Tympanic Membrane Epidermal Keratinocytes.

The human tympanic membrane (TM) has a thin outer epidermal layer which plays an important role in TM homeostasis and ear health. The specialised cells of the TM epidermis have a different physiology compared to normal skin epidermal keratinocytes, displaying a dynamic and constitutive migration that maintains a clear TM surface and assists in regeneration. Here, we characterise and compare molecular phenotypes in keratinocyte cultures from TM and normal skin. TM keratinocytes were isolated by enzymatic digestion and cultured in vitro. We compared global mRNA and microRNA expression of the cultured cells with that of human epidermal keratinocyte cultures. Genes with either relatively higher or lower expression were analysed further using the biostatistical tools g:Profiler and Ingenuity Pathway Analysis. Approximately 500 genes were found differentially expressed. Gene ontology enrichment and Ingenuity analyses identified cellular migration and closely related biological processes to be the most significant functions of the genes highly expressed in the TM keratinocytes. The genes of low expression showed a marked difference in homeobox (HOX) genes of clusters A and C, giving the TM keratinocytes a strikingly low HOX gene expression profile. An in vitro scratch wound assay showed a more individualised cell movement in cells from the tympanic membrane than normal epidermal keratinocytes. We identified 10 microRNAs with differential expression, several of which can also be linked to regulation of cell migration and expression of HOX genes. Our data provides clues to understanding the specific physiological properties of TM keratinocytes, including candidate genes for constitutive migration, and may thus help focus further research.

Distribution of different collagen types in the rat's tympanic membrane and its suspending structures.

BACKGROUND: The objective of the study was to investigate the histological distribution of collagens in the healthy rat's tympanic membrane. METHODS: Immunohistochemical analysis of collagen type I, II, III, and IV in the tympanic membranes in healthy adult female Sprague-Dawley rats. The staining was semiquantified using light microscopy in a blinded fashion, not knowing what type of collagen the slide had been stained for. RESULTS: The pars tensa of the tympanic membrane was mainly stained for collagen type II and IV. The fibrous annulus could on immunohistochemistry be subdivided into an inner and an outer portion. The inner portion of the fibrous annulus was mainly stained for collagen type II, whereas the outer portion was most strongly stained for collagen type III and collagen type IV. The test-retest reliability of the semiquantative method was 81%. CONCLUSION: Collagen type II and IV are the major collagen constituents of the pars tensa of the tympanic membrane. The outer portion of the fibrous annulus has collagen type III and IV as its major constituents, whereas the inner portion is made up of collagen type II.

Structural and functional properties of the healed tympanic membrane: a long-term follow-up after laser myringotomy.

HYPOTHESIS: The short-term healing scar that forms after experimental laser myringotomy will revert to a normal lamina propria in the long run. The mechanical stiffness will stay normal. BACKGROUND: Recent studies have shown severe structural changes in the fibrous layer in the early course after experimental laser myringotomy, whereas the scar quickly restored the strength of the tympanic membrane (TM). A reorganization of the fiber layer is expected to occur. METHODS: Potassium titanyl phosphate laser myringotomy was made on one side of the TM in Sprague-Dawley rats. The ear of the other side was untouched and used for control. After half a year of observation, the stiffness and strength of the healed TMs were measured with moiré interferometry and examined with otomicroscopy and light and electron microscopy. RESULTS: The interferometry readings showed a slightly reduced strength in the myringotomized and healed TMs. After half a year, still there were immense structural changes including increased thickness over a wide area of the pars tensa with increased amounts of fibers. An obvious reorganization of the fiber layer was lacking. CONCLUSION: Laser myringotomy causes profound, long-standing, or permanent structural changes in the lamina propria of the pars tensa, whereas the strength of the TM may become slightly reduced.

Healing time, long-term result and effects of stem cell treatment in acute tympanic membrane perforation.

OBJECTIVE: The incidence of otitis media in children between the age of 2 and 6 years is well documented. Repeated attacks may cause acute and chronic perforations. The surgical treatment for repairing chronic perforation is quite uncomfortable for the patients of this age group because of the invasiveness of this treatment. The aim of this study was to determine the long-term influence of embryonic stem cells on acute perforations and the effect of gelatin as a vehicle for applied stem cells. The possibility of teratogenic effects of the stem cells was also observed. METHODS: Bilateral laser myringotomy was performed in 17 adult Sprague-Dawley rats, divided into two groups. Gelatin, a substance suitable as vehicle for bioactive material was used bilaterally around the perforation in group A, to serve as a scaffold for repairing tissue. The stem cells were used in the right tympanic membrane perforation leaving the left tympanic membrane as a control. The animals in group B received the same treatment except for the use of gelatin and in addition received an immuno-suppressive agent. After half a year of observation the mechanical stiffness of the tympanic membrane was measured by moiré interferometry for group B and the morphological study was performed by light microscopy for both groups A and B and electron microscopy for group A. RESULTS: Stem cell treated ears did not show any enhanced healing of the perforation although a marked thickening of the lamina propria was observed compared with control group. After half a year the strength and the stiffness of the tympanic membrane was almost the same for both treated and untreated ears. No evidence of teratoma was found after half a year. CONCLUSION: This study suggests that the stem cells stimulate the proliferation of connective tissue and fibers in the lamina propria, possibly mediated by secreted substances, although the stiffness properties do not seem to be altered. The use of gelatin does not seem to enhance the healing process of the tympanic membrane perforation.

Clinical and audiological short-term and long-term outcomes of fat graft myringoplasty.

INTRODUCTION: Results of fat graft myringoplasty are often reported with only short-term follow-up. Audiological results are less commonly reported, as well as long-term follow-up results. MATERIALS AND METHODS: One hundred consecutive patients scheduled for fat graft myringoplasty were included in a prospective cohort study. Clinical and audiological outcomes were assessed at six weeks and one year postoperatively. RESULTS: Perforation sizes ranged from 0.5 to 4 mm. The six-week follow-up showed a total perforation closure rate of 72.9% with a statistically significant (p = .03) higher rate for the pediatric age group (83.0%). 64.4% of all patients were healed at one-year follow-up. Statistical analyses for background factors did not reveal any significant difference in healing rates with regard to patient sex or location or cause of the perforation. The mean preoperative air-conduction (AC) threshold was 25 dB with an air-bone gap of 12 dB. At the one-year follow-up the mean air conduction threshold for healed ears was improved to 16.6 dB, still 54.2% of them had a type B tympanogram. CONCLUSIONS: Children had a higher perforation closure rate at six-week follow-up than adult patients. Recurrent tympanic membrane perforations were common after initially successful fat graft myringoplasties. Long-term hearing was improved after successful fat graft myringoplasty, resulting in a mean AC threshold of 16.6 dB.

Effects of mechanical trauma to the human tympanic membrane: an experimental study using transmission electron microscopy.

HYPOTHESIS: To examine the cellular events following induced superficial lesions of the human tympanic membrane (TM). Such information could lead to enhanced appreciation of repair mechanisms and novel strategies to restore TM perforations. BACKGROUND: Persistent perforation of the TM in chronic otitis media is a major global health problem and frequently necessitates surgical intervention. Most TM perforations heal spontaneously and swiftly, but sometimes healing fails. The underlying mechanisms and the reason for incomplete repair are often elusive, although some mechanisms have been proposed. METHODS: Here, five healthy adult human TMs were sampled during vestibular schwannoma surgery. Three days before harvesting, three TMs were superficially lesioned, including the epithelial and sub-epithelial layers, using a needle and two TMs served as controls. Light and transmission electron microscopy were performed. RESULTS: Surrounding lesion showed distinct ultrastructural changes. This included a keratinocyte frontier with electron-dense cells with abundant ribosomes and nuclei metamorphosis. Beneath, were activated fibroblasts and invaded/transformed free cells and signs of increased transcellular activity of adjacent blood vessels. CONCLUSIONS: The study describes dynamic morphological events of a human lesioned TM. The human model may be used for further investigations and understanding of TM healing mechanisms.

Lugol's solution eradicates Staphylococcus aureus biofilm in vitro.

OBJECTIVES: The aim of the study was to evaluate the antibacterial efficacy of Lugol's solution, acetic acid, and boric acid against Staphylococcus aureus biofilm. METHODS: The efficacy of Lugol's solution 1%, 0.1%, and 0.05%, acetic acid 5% or boric acid 4.7% for treatment of Staphylococcus aureus biofilm in vitro was tested using 30 clinical strains. Susceptibility in the planktonic state was assessed by disk diffusion test. Antiseptic effect on bacteria in biofilm was evaluated by using a Biofilm-oriented antiseptic test (BOAT) based on metabolic activity, a biofilm bactericidal test based on culturing of surviving bacteria and confocal laser scanning microscopy combined with LIVE/DEAD staining. RESULTS: In the planktonic state, all tested S. aureus strains were susceptible to Lugol's solution and acetic acid, while 27 out of 30 tested strains were susceptible to boric acid. In biofilm the metabolic activity was significantly reduced following exposure to Lugol's solution and 5% acetic acid, while boric acid exposure led to no significant changes in metabolic activities. In biofilm, biocidal activity was observed for Lugol's solution 1% (30/30), 0.1% (30/30), and 0.05% (26/30). Acetic acid and boric acid showed no bactericidal activity in this test. Confocal laser scanning microscopy, assessed in 4/30 strains, revealed significantly fewer viable biofilm bacteria with Lugol's solution (1% p < 0.001, 0.1% p = 0.001 or 0.05% p = 0.001), acetic acid 5% for 10 min (p = 0.001) or 30 min (p = 0.015), but not for acetic acid for 1 min or boric acid. CONCLUSION: Lugol's solution 1.0% and 0.1% effectively eradicated S. aureus in biofilm and could be an alternative to conventional topical antibiotics where S. aureus biofilm is suspected such as external otitis, pharyngitis and wounds.