Electrocorticographic patterns dominated by low-frequency waves in camphor-induced seizures.

Camphor is an aromatic terpene compound found in the essential oils of many plants, which has been used for centuries as a herbal medicine, especially in children. However, many studies have shown that camphor may have major side effects, including neurological manifestation, such as seizures. In the present study, we investigated the electrocorticographic patterns of seizures induced by camphor in male adult Wistar rats. Each rat received 400 mg/kg (i.p.) of camphor prior to monitoring by electrocorticography. The application of camphor resulted a rapid evolution to seizure and marked changes in the electrocorticographic readings, which presented characteristics of epileptiform activity, with an increase in the total power wave. The decomposition of the cerebral waves revealed an increase in the delta and theta waves. The analysis of the camphor traces revealed severe ictal activity marked by an increase in the polyspike wave. Our data thus indicate that camphor may cause seizures, leading to tonic-clonic seizures. Clearly, further studies are necessary to better elucidate the mechanisms through which camphor acts on the brain, and to propose potential treatments with anticonvulsant drugs that are effective for the control of the seizures.

American Civil War plant medicines inhibit growth, biofilm formation, and quorum sensing by multidrug-resistant bacteria.

A shortage of conventional medicine during the American Civil War (1861-1865) spurred Confederate physicians to use preparations of native plants as medicines. In 1863, botanist Francis Porcher compiled a book of medicinal plants native to the southern United States, including plants used in Native American traditional medicine. In this study, we consulted Porcher's book and collected samples from three species that were indicated for the formulation of antiseptics: Liriodendron tulipifera, Aralia spinosa, and Quercus alba. Extracts of these species were tested for the ability to inhibit growth in three species of multidrug-resistant pathogenic bacteria associated with wound infections: Staphylococcus aureus, Klebsiella pneumoniae, and Acinetobacter baumannii. Extracts were also tested for biofilm and quorum sensing inhibition against S. aureus. Q. alba extracts inhibited growth in all three species of bacteria (IC50 64, 32, and 32 µg/mL, respectively), and inhibited biofilm formation (IC50 1 µg/mL) in S. aureus. L. tulipifera extracts inhibited biofilm formation (IC50 32 µg/mL) in S. aureus. A. spinosa extracts inhibited biofilm formation (IC50 2 µg/mL) and quorum sensing (IC50 8 µg/mL) in S. aureus. These results support that this selection of plants exhibited some antiseptic properties in the prevention and management of wound infections during the conflict.

Delivery of adapalene using a novel topical gel based on tea tree oil nano-emulsion: Permeation, antibacterial and safety assessments.

The aim of present study was to design and optimize 0.1% adapalene loaded nano-emulsion to improve the drug efficacy and increase its user compliance. Effect of type and concentration of surfactants was studied on size of 0.1% adapalene loaded nano-emulsion. Optimized formulation was then evaluated for particle size, polydispersity index, morphology, viscosity, and pH. Subsequently, 1% carbopol® 934 was incorporated to the optimized formulation for preparation of its gel form. The efficacy and safety of 0.1% adapalene loaded nano-emulsion gel was assessed compared to marketed gel containing 0.1% adapalene. In-vitro studies showed that adapalene permeation through the skin was negligible in both adapalene loaded nano-emulsion gel and adapalene marketed gel. Furthermore, drug distribution studies in skin indicated higher retention of adapalene in the dermis in adapalene loaded nano-emulsion gel compared with adapalene marketed gel. Antibacterial activity against Propionibacterium acnes showed that adapalene loaded nano-emulsion is effective in reducing minimum inhibitory concentration of the formulation in comparison with tea tree oil nano-emulsion, and pure tea tree oil. In vivo skin irritation studies showed absence of irritancy for adapalene loaded nano-emulsion gel. Also, blood and liver absorption of the drug, histological analysis of liver and liver enzyme activity of rats after 90 days' treatment were investigated. No drug was detected in blood/liver which in addition to an absence of any adverse effect on liver and enzymes showed the potential of adapalene loaded nano-emulsion gel as a novel carrier for topical delivery of adapalene.

Efficacy and safety of a therapeutic apparatus using hydrogen peroxide photolysis to treat dental and periodontal infectious diseases.

The present study aimed to evaluate the acute locally injurious property of our most current hydroxyl radical generation system by hydrogen peroxide (H2O2) photolysis. This system, which releases 3% H2O2 with a 405-nm laser, was developed in our laboratory for the treatment of dental and periodontal infectious diseases. First, the hydroxyl radical yield generated by H2O2 photolysis was examined by applying an electron spin resonance-spin trapping technique. Second, the bactericidal effect of the device was examined under a simulant condition in which Streptococcus mutans, a pathogenic bacterial species that causes caries, was irrigated with running 3% H2O2 concomitantly with laser irradiation. Finally, the acute topical effect of the model apparatus on rat palatal mucosa was evaluated by histological examination. We found that the hydroxyl radical yield was dependent upon laser output power. The bacterial count was substantially reduced within as little as 3 min. No abnormal findings were observed in the palatal mucosa, even when rats received three treatments of 3% H2O2 with laser irradiation at an output power of 40 mW. These results suggest that our apparatus has the ability to kill bacteria via hydroxyl radical generation and is safe to use at the lesion site of dental and periodontal infectious diseases.

Neurotoxicant exposure during pregnancy is a confounder for assessment of iodine supplementation on neurodevelopment outcome.

CONTEXT: The developing brain is vulnerable to iodine deficiency (ID) and environmental neuro-toxicants. OBJECTIVES: To assess neurocognitive development of children whose mothers have received (or not) iodine supplementation during pregnancy, in an area of borderline ID, while assessing in utero exposure to environmental neuro-toxicants. DESIGN/PATIENTS: Among 86 children born from normal euthyroid women who participated in our prospective interventional study on iodine supplementation (150 µg/day) started early in pregnancy, 44 (19 with iodine supplementation, 25 controls) were assessed at two years using the Bayley test. Information on parents' education and habits (smoking), and on child development was recorded. Thyroid tests at each trimester of pregnancy and on cord blood (CB) were available, as well as milk concentrations of selected environmental compounds known for their neurotoxicity, including heavy metals and PCBs. RESULTS: There was no difference in Bayley tests for children born to mothers with and without iodine supplementation, but sample size was small. Language and Social-Emotional Scales were negatively correlated with TBG at all times tested, while PCB 118 correlated negatively with all Language scales. Among maternal and CB thyroid tests, only CB thyroglobulin, the best marker of iodine status, correlated (negatively) with neurodevelopment scales (Motor and Expressive Language). CONCLUSIONS: This pilot study suggests that PCB118 has a negative impact on neurocognitive development, possibly mitigating the benefit of iodine supplementation in an area of borderline ID. We propose that exposure to environmental neurotoxicants should be taken into account when designing studies on the benefit of iodine supplementation in pregnancy. The potential interactions between TBG, environmental neurotoxicants and brain development warrant further studies.

Cytokine expression in human osteoblasts after antiseptic treatment: a comparative study between polyhexanide and chlorhexidine.

PURPOSE/AIM OF THE STUDY: Chlorhexidine and polyhexanide are frequently used antiseptics in clinical practice and have a broad antimicrobial range. Both antiseptics are helpful medical agents for septic wound treatment with a high potential for defeating joint infections. Their effect on human osteoblasts has, so far, not been sufficiently evaluated. The aim of this study was to investigate the activating potential of polyhexanide and chlorhexidine on inflammatory cytokines/chemokines in human osteoblasts in vitro. MATERIALS AND METHODS: Human osteoblasts were isolated and cultivated in vitro and then treated separately with 0.1% and 2% chlorhexidine and 0.04% polyhexanide as commonly applied concentrations in clinical practice. Detection of cell structure and cell morphology was performed by light microscopic inspection. Cytokine and chemokine secretion was determined by using a multiplex suspension array. RESULTS: Cell shrinking, defective cell membrane, and the loss of cell adhesion indicated cell damage of human osteoblasts after treatment with both antiseptics was evaluated by using light microscopy. Polyhexanide, but not chlorhexidine, caused human osteoblasts to secrete various interleukins (1ß, 6, and 7), interferon γ, tumor necrosis factor α, vascular endothelial growth factor, eotaxin, fibroblast growth factor basic, and granulocyte macrophage colony-stimulating factor as quantified by multiplex suspension array. CONCLUSIONS: Both antiseptics induced morphological cell damage at an optimum exposure between 1 and 10 min. But only polyhexanide mediated a pronounced secretion of inflammatory cytokines and chemokines in human osteoblasts. Therefore, we recommend a preferred usage of chlorhexidine in septic surgery to avoid the induction of an inflammatory reaction.

[Microbicides for preventing sexually transmitted infections: Current status and strategies for preclinical evaluation of new candidates]. / Microbicidas para la prevención de infecciones de transmisión sexual: estado actual y estrategias para la evaluación preclínica de nuevos candidatos.

Microbicides are a new tool, still under investigation, which could help prevent infection by the human immunodeficiency virus (HIV) and other sexually transmitted infections (STIs). Increasing evidence shows that the complexity of sexual transmission of viral pathogens requires the identification of compounds able to block the early events during the cycle of viral infection. In this manuscript we provide a comprehensive review of the different microbicide strategies that have been studied or are currently being considered for STI prevention, particularly emphasizing those having the potential to block HIV infection. The manuscript also reviews the complex process that is required to conduct future clinical studies in humans and concludes with a brief discussion of the strategies that could be part of the immediate future in microbicide research.

Development of HIV-1 rectal-specific microbicides and colonic tissue evaluation.

The gastrointestinal tract is structurally and functionally different from the vagina. Thus, the paradigm of topical microbicide development and evaluation has evolved to include rectal microbicides (RMs). Our interest was to create unique RM formulations to safely and effectively deliver antiretroviral drugs to mucosal tissue. RMs were designed to include those that spread and coat all surfaces of the rectum and distal colon rapidly (liquid) and those that create a deformable, erodible barrier and remain localized at the administration site (gel). Tenofovir (TFV) (1%) was formulated as an aqueous thermoreversible fluid and a carbopol-based aqueous hydrogel. Lipid-based liquid and gel formulations were prepared for UC781 (0.1%) using isopropyl myristate and GTCC (Caprylic/Capric Triglycerides), respectively. Formulations were characterized for pH, viscosity, osmolality, and drug content. Pre-clinical testing incorporated ex vivo colonic tissue obtained through surgical resections and flexible sigmoidoscopy (flex sig). As this was the first time using tissue from both sources side-by-side, the ability to replicate HIV-1 was compared. Efficacy of the RM formulations was tested by applying the products with HIV-1 directly to polarized colonic tissue and following viral replication. Safety of the formulations was determined by MTT assay and histology. All products had a neutral pH and were isoosmolar. While HIV-1BaL and HIV-1JR-CSF alone and in the presence of semen had similar replication trends between surgically resected and flex sig tissues, the magnitude of viral replication was significantly better in flex sig tissues. Both TFV and UC781 formulations protected the colonic tissue, regardless of tissue source, from HIV-1 and retained tissue viability and architecture. Our in vitro and ex vivo results show successful formulation of unique RMs. Moreover, the results of flex sig and surgically resected tissues were comparable suggesting the incorporation of both in pre-clinical testing algorithms.

Influence of different additives and their concentrations on corneal toxicity and antimicrobial effect of benzalkonium chloride.

PURPOSE: The aim of this study was to examine the ophthalmic additives responsible for modulating acute corneal epithelial toxicity induced by benzalkonium chloride (BAC) and investigate the ability of polyoxyethylene hydrogenated castor oil 40 (HCO-40) and polysorbate 80 (PS-80) to reduce the corneal toxicity and antimicrobial effects of BAC. METHODS: Cytotoxicity of the additives, which included glycerin, polyvinyl alcohol, propylene glycol, polyethylene glycol, and PS-80, on rabbit corneal epithelial cells was examined using the cell proliferation assay in the presence and absence of 0.02% BAC. The corneal transepithelial electrical resistance change after a 60-second exposure to HCO-40 or PS-80 mixed with 0.02% BAC was measured in living rabbits. Corneal damage was examined using scanning electron microscopy. The antimicrobial activities of HCO-40 and PS-80 with 0.02% BAC against Staphylococcus aureus, Propionibacterium acnes, Pseudomonas aeruginosa, Escherichia coli, and Streptococcus pneumoniae were assessed. RESULTS: Of all the tested additives, only PS-80 could prevent the BAC-induced cytotoxicity. Corneal epithelial barrier function disorder caused by 0.02% BAC was significantly alleviated by either PS-80 or HCO-40 in a concentration-dependent manner. Scanning electron microscopy images showed an improvement of BAC-induced corneal epithelial toxicity after the addition of HCO-40 or PS-80. The antimicrobial effect of the BAC against P. aeruginosa, E. coli, and S. pneumoniae was reduced after adding HCO-40 or PS-80. CONCLUSIONS: HCO-40 and PS-80 reduce acute corneal toxicity and the antimicrobial effect of BAC. Possible interactions between BAC and other additives should be taken into consideration when evaluating the toxicity and antibacterial properties of BAC.

Microbicidas para la prevención de infecciones de transmisión sexual: estado actual y estrategias para la evaluación preclínica de nuevos candidatos. / [Microbicides for preventing sexually transmitted infections: Current status and strategies for preclinical evaluation of new candidates].

Microbicides are a new tool, still under investigation, which could help prevent infection by the human immunodeficiency virus (HIV) and other sexually transmitted infections (STIs). Increasing evidence shows that the complexity of sexual transmission of viral pathogens requires the identification of compounds able to block the early events during the cycle of viral infection. In this manuscript we provide a comprehensive review of the different microbicide strategies that have been studied or are currently being considered for STI prevention, particularly emphasizing those having the potential to block HIV infection. The manuscript also reviews the complex process that is required to conduct future clinical studies in humans and concludes with a brief discussion of the strategies that could be part of the immediate future in microbicide research.

Diaryltriazine non-nucleoside reverse transcriptase inhibitors are potent candidates for pre-exposure prophylaxis in the prevention of sexual HIV transmission.

OBJECTIVES: Pre-exposure prophylaxis and topical microbicides are important strategies in the prevention of sexual HIV transmission, especially since partial protection has been shown in proof-of-concept studies. In search of new candidate drugs with an improved toxicity profile and with activity against common non-nucleoside reverse transcriptase inhibitor (NNRTI)-resistant HIV, we have synthesized and investigated a library of 60 new diaryltriazine analogues. METHODS: From this library, 15 compounds were evaluated in depth using a broad armamentarium of in vitro assays that are part of a preclinical testing algorithm for microbicide development. Antiviral activity was assessed in a cell line, and in primary human cells, against both subtype B and subtype C HIV-1 and against viruses resistant to therapeutic NNRTIs and the candidate NNRTI microbicide dapivirine. Toxicity towards primary blood-derived cells, cell lines originating from the female reproductive tract and female genital microflora was also studied. RESULTS AND CONCLUSIONS: We identified several compounds with highly potent antiviral activity and toxicity profiles that are superior to that of dapivirine. In particular, compound UAMC01398 is an interesting new candidate that warrants further investigation because of its superior toxicity profile and potent activity against dapivirine-resistant viruses.

Animal models for microbicide safety and efficacy testing.

PURPOSE OF REVIEW: Early studies have cast doubt on the utility of animal models for predicting success or failure of HIV-prevention strategies, but results of multiple human phase 3 microbicide trials, and interrogations into the discrepancies between human and animal model trials, indicate that animal models were, and are, predictive of safety and efficacy of microbicide candidates. RECENT FINDINGS: Recent studies have shown that topically applied vaginal gels, and oral prophylaxis using single or combination antiretrovirals are indeed effective in preventing sexual HIV transmission in humans, and all of these successes were predicted in animal models. Further, prior discrepancies between animal and human results are finally being deciphered as inadequacies in study design in the model, or quite often, noncompliance in human trials, the latter being increasingly recognized as a major problem in human microbicide trials. SUMMARY: Successful microbicide studies in humans have validated results in animal models, and several ongoing studies are further investigating questions of tissue distribution, duration of efficacy, and continued safety with repeated application of these, and other promising microbicide candidates in both murine and nonhuman primate models. Now that we finally have positive correlations with prevention strategies and protection from HIV transmission, we can retrospectively validate animal models for their ability to predict these results, and more importantly, prospectively use these models to select and advance even safer, more effective, and importantly, more durable microbicide candidates into human trials.

Toxicity of antiseptics against chondrocytes: what is best for the cartilage in septic joint surgery?

In septic joint surgery, the most frequently used antiseptics are polyhexanide, hydrogen peroxide and taurolidine. The aim of this study was to examine the effects of these antiseptics on viability of human chondrocytes. Our hypothesis was that antiseptics and supplemental irrigation with sodium chloride lavage are less toxic on human chondrocytes than treatment with antiseptics only. Primary human chondrocytes were isolated and cultured from six donated human knee joints. Polyhexanide, hydrogen peroxide or taurolidine were added to the cultures. Toxicity analysis was performed by visualisation of cell structure using light microscopy and LDH activity. The determination of vital cells and total cell numbers of chondrocytes treated with antiseptics partly followed by irrigation with sodium chloride solution was performed by using Casy Cell-Counter. Light microscopic data revealed a defect in cell structure after addition of antiseptics. We showed a significant increase of LDH enzyme activity after the treatment with polyhexanide or taurolidine. After treatment with antiseptics followed by sodium chloride solution a significant increase of vital and total cell numbers resulted in comparison with the chondrocytes that were only treated with antiseptics. The data show that treatment with polyhexanid, hydrogen peroxide or taurolidine induces cell death of human chondroctes in vitro. The application of sodium chloride solution after the treatment with polyhexanide and hydrogen peroxide possibly has a protective effect on chondrocyte viability.

Properties of a new mouthrinse for patients receiving radiation therapy.

INTRODUCTION: Patients receiving radiation therapy due to oral cancer develop complications such as hyposalivation, mucositis, oral infections, dental hypersensitivity and caries. Mouthrinses can alleviate some of these problems. AIMS AND OBJECTIVES: To investigate the in vitro antimicrobial properties and cytotoxicity of an experimental mouthrinse. METHODS: The mouthrinse contained 30% hexylene glycol (glycerine), 7% potassium nitrate and 0.025% sodium fluoride. The minimal inhibitory concentration (MIC) of these ingredients and the mixture was determined for C. albicans, S. aureus and S. mutans over 24 hours at different concentrations. The MICs of two commercial mouthrinses, Corsodyl and Plax, were also determined using the same organisms. All mouthrinses were then tested to determine the percentage kill over 1, 2, and 3 minutes. RESULTS: The MICs for hexylene glycol were 10%, 30% and 10% for C. albicans, S. aureus and S. mutons respectively. Potassium nitrate and sodium fluoride had no antimicrobial effects. The MIC of Corsodyl was 0.016 mg/ml for all the test organisms. The MIC for Plax varied from 0.0002 mg/ml to 0.001 mg/ml. The kill rates for all mouthrinses were acceptable, with no statistical differences between them. The experimental mouthrinse was not toxic to human oesophageal SCC cells after 1 minute exposure. At the time of the experiment, the costs of a similar quantity of the experimental mouthrinse, Corsodyl and Plax were R5.24, R30.00 and R10.00 respectively. CONCLUSIONS: The experimental mouthrinse was cost-effective and proved to have an antimicrobial effect and could be used safely to alleviate oral infections, desensitize teeth, improve oral hygiene and control dental caries in cancer patients after radiation therapy.

Eicosapentaenoic and docosahexaenoic acids enriched polyunsaturated fatty acids from the coastal marine fish of Bay of Bengal and their therapeutic value.

Eicosapentaenoic acid (EPA)/docosahexaenoic acid (DHA) enriched polyunsaturated fatty acids (PUFA) significantly present in marine fish oil emerge as preventive agents for combating many health problems specially in chronic or metabolic disorders. The fish in the coastal area of Bay of Bengal has remained unexplored with respect to EPA/DHA enriched PUFA content in its oils, although it may be a potential source in harnessing the health benefit. In this study, seven varieties of the coastal fish were analysed for the content of EPA/DHA. The one locally known as lotte, (Harpadon nehereus) though has low content of total lipids, was found to have high EPA/DHA in its oil. The phospholipids rich fraction was extracted from the total fish oil. The EPA/DHA enriched PUFA was isolated to investigate the potential use for health benefits. EPA/DHA is found to act as protective agent against mercury poisoning studied in cell culture as well as in animal mode. It is found to be highly preventive in diabetes. The lotte is available in the coastal area of Bay of Bengal adjoining West Bengal, India in large scale and it is the first report showing EPA/DHA enriched PUFA in these fish oil that can be availed to harness in important health benefits.

Cytotoxicity evaluation of Persica mouthwash on cultured human and mouse cell lines in the presence and absence of fetal calf serum.

BACKGROUND AND AIMS: The effectiveness of an ideal antimicrobial agent depends on its ability to kill microbes while causing minimal toxicity to host cells. Several studies have been reported on the antimicrobial effects of chewing sticks (Salvadora persica) on oral bacteria. The purpose of this study was to evaluate the cytotoxic effects of Persica and chlorhexidine (CHX) mouthwashes on cultured human and mouse cell lines. MATERIALS AND METHODS: This was an experimental study. The toxic effects of four dilutions of Persica and CHX mouthwashes on KB, Saos-2, J744 A1, and gingival fibroblast cells were evaluated by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay. The effect of fetal calf serum (FCS) components on the cytotoxicity of these mouthwashes was also investigated. STATISTICAL ANALYSIS: Analysis of variance and the Kruskal-Wallis test were used to evaluate the results. RESULTS: The results indicated that Persica , at concentrations higher than 0.1%, exerted a very significant cytotoxic effect on all the cell lines (P < or = 0.01). CHX, at a concentration of 0.001%, exerted toxic effects only on gingival fibroblasts; concentrations higher than 0.001% were required to produce significant cell death in the other cell lines. At all the concentrations under study, both Persica and CHX exerted significantly greater cytotoxic effects in the absence of FCS than in its presence (i.e., in control culture medium). The toxicities of both mouthwashes were attenuated in the presence of FCS (10%). CONCLUSION: Our results indicate that both Persica and CHX mouthwashes are toxic to macrophage, epithelial, fibroblast, and osteoblast cells in a concentration-dependent manner.

Influence of nitrogen and phosphorus concentrations and ratios on Lemna gibba growth responses to triclosan in laboratory and stream mesocosm experiments.

The effects of co-occurring nutrient and contaminant stressors are very likely to interact in aquatic systems, particularly at the level of primary producers. Site-specific nitrogen (N) and phosphorus (P) concentrations are often much lower and differ in relative availability than those used in nutrient-saturated laboratory assays for aquatic plants, which can introduce uncertainty in prospective ecological hazard and risk assessments. Because triclosan, an antimicrobial agent included in personal care products, potentially presents high relative risk among antimicrobial agents to aquatic plants and algae, we performed laboratory experiments with the model aquatic macrophyte Lemna gibba across a gradient of environmentally relevant N:P levels with and without triclosan co-exposure. Frond numbers (7 d) were significantly higher in N:P treatments of 16 and 23 but were lower in N:P of 937 and 2,500 treatments relative to standardized control media (N:P=3). When triclosan co-exposure occurred at high nutrient concentrations, frond number median effective concentration values at N:P 0.75, 3, and 16 were more than twofold lower than triclosan median effective concentration values in low nutrient media N:P ratios. However, a triclosan median effective concentration for frond number was twofold lower at N:P of 2,500 than at other N:P ratios in low concentration media. Influences of P enrichment on triclosan toxicity to L. gibba were further explored during a 14-d outdoor experimental stream mesocosm study. Effects of 2.6 and 20.8 microg L(-1) triclosan on L. gibba growth rates were more pronounced with increasing P treatment levels, which was generally consistent with our laboratory observations. Findings from these laboratory and field studies indicate that site-specific nutrient concentrations and ratios should be considered during assessments of primary producer responses to chemical stressors.

Changes in antibiotic susceptibility in staphylococci habituated to sub-lethal concentrations of tea tree oil (Melaleuca alternifolia).

AIMS: To investigate the effect of sub-lethal challenge with tea tree oil (TTO) on the antibiotic resistance profiles of staphylococci. METHODS AND RESULTS: Isolates of methicillin-resistant/-sensitive Staphylococcus aureus (MRSA and MSSA) and coagulase-negative staphylococci (CONS) were habituated to sub-lethal concentrations of TTO (72 h). Following habituation, the minimum inhibitory concentrations (MIC) of antibiotics and TTO were determined. Habituated MRSA/MSSA cultures had higher (P < 0.05) MIC values than control cultures for the examined antibiotics. Habituated MRSA/MSSA cultures also displayed decreased susceptibility to TTO. Although the MIC of habituated MRSA/MSSA for the examined antibiotics reverted to control values after subsequent culture in the absence of TTO, the increased MIC against TTO were maintained. When compared with control cultures, habituated CoNS cultures had higher (P < 0.05) MIC values against three-fifths of the antibiotics examined; no changes in TTO MIC were observed. CONCLUSIONS: TTO habituation 'stress-hardens' MRSA and MSSA, evidenced by transient decreased antibiotic susceptibility and stable decreased TTO susceptibility. Although TTO habituation did not decrease susceptibility of CoNS to TTO, such cultures showed transient decreased antibiotic susceptibility. SIGNIFICANCE AND IMPACT OF THE STUDY: Application of TTO at sub-lethal concentrations may reduce the efficacy of topical antibiotics used with TTO in combination therapies.