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Background Honey has broad spectrum antibacterial activity against clinically important organisms and may be suitable for treating superficial bacterial infections. However, very little data are available describing potential interactions between honey and other topically applied agents such as antiseptics or essential oils. Methods Interactions between pairs of antibacterial agents were investigated by performing checkerboard assays and determining the fractional inhibitory concentration indices (FICIs). Interactions between the two monofloral honeys marri (from Corymbia calophylla) and manuka, and the antiseptic agents benzalkonium chloride, chlorhexidine digluconate, silver (I) nitrate, tea tree oil, and Eucalyptus polybractea oil were investigated against Staphylococcus aureus ATCC® 43300 and Pseudomonas aeruginosa ATCC® 27853. Results Additive or indifferent interactions (FICI 0.5—2) were observed for all combinations against both organisms tested, with the exception of chlorhexidine and honey. Chlorhexidine and marri honey showed an antagonistic relationship against S. aureus (median FICI 2.00, range 1.25—4.83). Similarly, chlorhexidine and manuka honey showed antagonism against S. aureus (median FICI 2.33, range 2.00—2.67). Conclusions With the exception of chlorhexidine, these data indicate that honey does not interfere with the antimicrobial activity of the tested agents, and that honey may be suitable for combination therapy with other topically applied antibacterial agents for treating superficial bacterial infections.

Multidrug resistant (MDR) P. aeruginosa accounts for 35% of all P. aeruginosa isolated from respiratory samples of patients with cystic fibrosis (CF). The usefulness of β-lactam antibiotics for treating CF, such as carbapenems and later generation cephalosporins, is limited by the development of antibacterial resistance. A proven treatment approach is the combination of a β-lactam antibiotic with a β-lactamase inhibitor. New β-lactam/β-lactamase inhibitor combinations are available, but data are lacking regarding the susceptibility of MDR CF-associated P. aeruginosa (CFPA) to these new combination therapies. In this study we determined MIC values for three new combinations; imipenem-relebactam (I-R), ceftazidime-avibactam (CZA), and ceftolozane-tazobactam (C/T) against MDR CFPA (n = 20). The MIC90 of I-R, CZA, and C/T was 64/4, 32/4, and 16/8 (all µg/mL), respectively. The susceptibility of isolates to imipenem was not significantly improved with the addition of relebactam (p = 0.68). However, susceptibility to ceftazidime was significantly improved with the addition of avibactam (p P. aeruginosa.

Impetigo is a contagious skin disease caused by Staphylococcus aureus and Streptococcus pyogenes. Without treatment, impetigo may be recurrent, develop into severe disease, or have serious, life-threatening sequelae. Standard treatment consists of topical or systemic antibiotic therapy (depending on severity), however, due to antibiotic resistance some therapies are increasingly ineffective. In this study we evaluated the potential for honey as an alternative treatment for impetigo. A broth microdilution assay in 96-well microtitre trays was used to determine the minimum inhibitory concentrations (MICs) of six monofloral honeys (jarrah, marri, red bell, banksia, wandoo, and manuka), a multifloral honey and artificial honey against S. aureus (n = 10), S. pyogenes (n = 10), and coagulase-negative staphylococci (CoNS) (n = 10). The optical density (OD) of all microtitre tray wells was also determined before and after assay incubation to analyse whether sub-MIC growth inhibition occurred. Jarrah, marri, red bell, banksia, and manuka honeys were highly effective at inhibiting S. aureus and CoNS, with MIC

Clostridioides (formerly Clostridium) difficile is a Gram-positive anaerobic bacterial pathogen that causes severe gastrointestinal infection in humans. The current chemotherapeutic options are vastly inadequate, expensive and limited; this results in an exorbitant medical and financial burden. New, inexpensive chemotherapeutic treatments for C. difficile infection with improved efficacy are urgently needed. A streamlined synthetic pathway was developed to allow access to 38 novel mono- and di-cationic biaryl 1,2,3-triazolyl peptidomimetics with increased synthetic efficiency, aqueous solubility and enhanced antibacterial efficacy. The monocationic arginine derivative 28 was identified as a potent, Gram-positive selective antibacterial with MIC values of 4 μg/mL against methicillin-resistant Staphylococcus aureus and 8 μg/mL against C. difficile. Furthermore, the dicationic bis-triazole analogue 50 was found to exhibit broad-spectrum activity with substantial Gram-negative efficacy against Acinetobacter baumannii (8 μg/mL), Pseudomonas aeruginosa (8 μg/mL) and Klebsiella pneumoniae (16 μg/mL); additionally, compound 50 displayed reduced haemolytic activity (13%) in an in vitro haemolysis assay. Membrane-disruption assays were conducted on selected derivatives to confirm the membrane-active mechanism of action inherent to the synthesized amphiphilic compounds. A comparative solubility assay was developed and utilized to optimize the aqueous solubility of the compounds for in vivo studies. The biaryl peptidomimetics 28 and 67 were found to exhibit significant efficacy in an in vivo murine model of C. difficile infection by reducing the severity and slowing the onset of disease.

Synthetic small molecular antimicrobial peptidomimetics represent a promising new class of potential antibiotics due to their membrane-disrupting ability and their decreased propensity for bacterial resistance. A library of 43 mono- and di-cationic biaryl 1,2,3-triazolyl peptidomimetics was designed and synthesized based upon previously established lead biarylpeptidomimetics and a known pharmacophore. A reliable, facile and modular synthetic pathway allowed for the efficient synthesis of multiple unique scaffolds which were subjected to divergent derivatization to furnish the amphiphilic compounds. In vitro testing revealed enhanced antibacterial efficacy against a range of pathogenic bacteria, including bacterial isolates with methicillin, vancomycin, daptomycin, or multi-drug resistance. Preliminary time-kill kinetics and membrane-disruption assays revealed a likely membrane-active mechanism for the tested peptidomimetics. An optimal balance between hydrophobicity and cationic charge was found to be essential for reduced cytotoxicity/haemolysis (i.e. membrane selectivity) and enhanced Gram-negative activity. The cationic biaryl amphiphile 81 was identified as a potent, broad-spectrum peptidomimetic with activity against Gram-positive (methicillin-resistant Staphylococcus aureus - MIC = 2 μg/mL) and Gram-negative (Escherichia coli - MIC = 4 μg/mL) pathogenic bacteria.

The phenol equivalence assay is the current industry-adopted test used to quantify the antibacterial activity of honeys in Australia and New Zealand. Activity is measured based on the diffusion of honey through agar and resulting zone of growth inhibition. Due to differences in the aqueous solubilities of antibacterial compounds found in honeys, this method may not be optimal for quantifying activity. Therefore, a new method was developed based on the existing broth microdilution assay that is widely used for determining minimum inhibitory concentrations (MICs). It utilises the four organisms Staphylococcus aureus ATCC 29213, Enterococcus faecalis ATCC 29212, Escherichia coli ATCC 25922 and Pseudomonas aeruginosa ATCC 27853, and an optical density endpoint to quantify bacterial growth. Decreases in bacterial growth in the presence of honey, relative to the positive growth control, are then used to derive a single value to represent the overall antibacterial activity of each honey. Antibacterial activity was quantified for a total of 77 honeys using the new method, the phenol equivalence assay and the standard broth microdilution assay. This included 69 honeys with undisclosed floral sources and the comparators Manuka, Jarrah (Eucalyptus marginata), Marri (Corymbia calophylla), artificial and multifloral honey. For the 69 honey samples, phenol equivalence values ranged from 0–48.5 with a mean of 34 (% w/v phenol). Mean MICs, determined as the average of the MICs obtained for each of the four organisms for each honey ranged from 7–24% (w/v honey). Using the new assay, values for the 69 honeys ranged from 368 to 669 activity units, with a mean of 596. These new antibacterial activity values correlated closely with mean MICs (R2 = 0.949) whereas the relationship with phenol equivalence values was weaker (R2 = 0.649). Limit of detection, limit of quantitation, measuring interval, limit of reporting, sensitivity, selectivity, repeatability, reproducibility, and ruggedness were also investigated and showed that the new assay was both robust and reproducible.

Background: The efficacy, tolerability and acceptability of a tea tree oil gel (200 mg/g) and face wash (7 mg/g) were evaluated for the treatment of mild to moderate facial acne. Methods: In this open-label, uncontrolled phase II pilot study, participants applied tea tree oil products to the face twice daily for 12 weeks and were assessed after 4, 8 and 12 weeks. Efficacy was determined from total numbers of facial acne lesions and the investigator global assessment (IGA) score. Tolerability was evaluated by the frequency of adverse events and the mean tolerability score determined at each visit. Product acceptability was assessed via a questionnaire at the end of the study period. Results: Altogether 18 participants were enrolled, of whom 14 completed the study. Mean total lesion counts were 23.7 at baseline, 17.2 at 4, 15.1 at 8 and 10.7 at 12 weeks. Total lesion counts differed significantly over time by repeated measures anova (P < 0.0001). The mean IGA score was 2.4 at baseline, 2.2 at 4, 2.0 at 8 and 1.9 at 12 weeks, which also differed significantly over time (P = 0.0094). No serious adverse events occurred and minor local tolerability events were limited to peeling, dryness and scaling, all of which resolved without intervention. Conclusion: This study shows that the use of the tea tree oil products significantly improved mild to moderate acne and that the products were well tolerated.

Clostridium difficile infection is a toxin-mediated disease of the colon. C. difficile virulence is primarily attributed to the production of toxin A and toxin B; thus this study was aimed to investigate the effect of a range of natural products on the production and activity of C. difficile toxins in vitro. Twenty-two natural products were investigated against four C. difficile strains. The activity of products against toxins was determined using Vero and HT-29 cells cytotoxicity and neutral red uptake assays. The indirect effect of products on toxin-mediated cytotoxicity was determined using the same cell lines. The effect of seven products on toxin production by C. difficile was determined using ELISA. Zingerone (0.3 mg/ml) protected both cell lines from C. difficile cytopathic effects, confirmed by the neutral red uptake assay (P Leptospermum honeys (4% w/v), fresh onion bulb extract (12.5% v/v) and trans-cinnamaldehyde (0.005% v/v) all reduced toxin production and activity significantly (P ≤ 0.023). Garlic clove powder (4.7 mg/ml) only reduced toxin activity (P ≤ 0.047). Overall, several natural products had activity against C. difficile toxins in vitro encouraging further investigation against C. difficile toxins in vivo.

Over-the-counter acne treatments containing tea tree oil from the plant Melaleuca alternifolia are widely available, and evidence indicates that they are a common choice amongst those self-treating their acne. The aims of this review were to collate and evaluate the clinical evidence on the use of tea tree oil products for treating acne, to review safety and tolerability and to discuss the underlying modes of therapeutic action.

Clostridium difficile (C. difficile) is a problematic Gram positive bacterial pathogen causing moderate to severe gastrointestinal infections. Based on a lead binaphthyl-tripeptide dicationic antimicrobial, novel mono-, di- and tri-peptidomimetic analogues targeting C. difficile were designed and synthesized incorporating one, two or three d-configured cationic amino acid residues, with a common 1,2,3-triazole ester isostere at the C-terminus. Copper- and ruthenium-click chemistry facilitated the generation of a 46 compound library for in vitro bioactivity assays, with structure-activity trends over the largest compound subset revealing a clear advantage to triazole-substitution with a linear or branched hydrophobic group. The most active compounds were dicationic-dipeptides where the triazole was substituted with a 4- or 5-cyclohexylmethyl or 4,5-diphenyl moiety, providing MICs of 4 μg mL(-1) against three human isolates of C. difficile. Further biological screening revealed significant antimicrobial activity for several compounds against other common bacterial pathogens, both Gram positive and negative, including S. aureus (MICs ≥2 μg mL(-1)), S. pneumoniae (MICs ≥1 μg mL(-1)), E. coli (MICs ≥4 μg mL(-1)), A. baumannii (MICs ≥4 μg mL(-1)) and vancomycin-resistant E. faecalis (MICs ≥4 μg mL(-1)).

Clostridium difficile is an anaerobic, Gram-positive, spore-forming bacillus that causes disease ranging from self-limiting diarrhoea to severe pseudomembranous colitis. C. difficile infection (CDI) commonly affects hospitalised patients and is increasingly identified in patients in the community with no hospital contact. For the last 15 years the incidence of CDI worldwide has been rising, especially in the northern hemisphere. The yearly average number of hospitalizations as a result of this disease is estimated to be over a quarter of a million per year in the United States alone. The main risk factor for CDI is exposure to antimicrobials that affect the gut microflora and, paradoxically, the most common treatments for CDI are the antimicrobials, metronidazole and vancomycin. However, the increasing frequency of highly virulent C. difficile strains, antimicrobial treatment failures, hospital outbreaks, patients with severe complications and cases with multiple recurrences have driven the search for new therapies. Several novel or popular complementary and alternative therapies are self-prescribed for treatment of other diarrheal diseases, and these may also be appropriate for treating CDI. In general, complementary and alternative medicine (CAM) is mainly used by patients when conventional therapeutic agents show limited success against C. difficile and other antimicrobial-resistant bacteria. Among these alternative approaches, a number of treatments, such as herbal remedies, are embraced less by pharmaceutical and medical professions. This review summarises current knowledge of non-conventional antimicrobial and alternative therapies for treatment of CDI. As the demand for non-conventional antimicrobial therapies increases, further studies are required in the field of CAM, especially natural products, for the treatment of CDI.

The aim of this study was to seek additional data on the antimicrobial susceptibility of Staphylococcus spp. after habituation to low levels of the topical antimicrobial agent tea tree (Melaleuca alternifolia) oil. Meticillin-susceptible Staphylococcus aureus (MSSA), meticillin-resistant S. aureus (MRSA) and coagulase-negative staphylococci (CoNS) were habituated to 0.075% tea tree oil for 3 days. Subsequently, the susceptibility of five isolates each of MSSA, MRSA and CoNS to fusidic acid, mupirocin, chloramphenicol, linezolid and vancomycin was determined by Etest, and susceptibility to tea tree oil, terpinen-4-ol, carvacrol and triclosan was determined by agar dilution. Following habituation to 0.075% tea tree oil, antimicrobial MICs differed between control and habituated isolates on 33 occasions (out of a possible 150), with MICs being higher in habituated isolates on 22 occasions. Using clinical breakpoint criteria, one MSSA isolate changed susceptibility category from vancomycin-susceptible (MIC = 2 mu g/mL) to intermediate susceptibility (MIC = 3 mu g/mL) after habituation in one of two replicates. For the non-antibiotic antimicrobial agents, MICs of habituated and control isolates differed on 12 occasions (out of a possible 120); 10 occasions in MRSA and 2 occasions in MSSA. MICs were higher for habituated isolates on five occasions. However, all the differences were one serial dilution only and were not regarded as significant. Habituation to sublethal concentrations of tea tree oil led to minor changes in MICs of antimicrobial agents, only one of which may have been clinically relevant. There is no evidence to suggest that tea tree oil induces resistance to antimicrobial agents. (C) 2013 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

This study investigated the effects of the volatile terpene-rich oil from Melaleuca alternifolia (tea tree oil) on the formation of biofilms and the adhesion of C. albicans cells to both biotic and abiotic surfaces. Biofilm formation on polystyrene was significantly inhibited for 70% of the isolates at the lowest test concentration of 0.016% of tea tree oil (TTO) when quantified by XTT and 40% of isolates when measured by crystal violet staining. Adhesion to polystyrene, quantified by crystal violet staining, was significantly reduced for 3 isolates at 0.031%, 6 isolates at 0.062% and 0.125% and for all 7 isolates at 0.25% TTO. Reductions in adhesion were not due to loss of viability (at concentrations of ≤ 0.125%) or interactions between the TTO and polystyrene. Similarly, adhesion to buccal epithelial and HeLa cells was also significantly reduced in the presence of 0.016-0.062% TTO. Treatment with 0.125% TTO, but not 0.062%, decreased the cell surface hydrophobicity of C. albicans, indicating one potential mechanism by which adhesion may be reduced. These data demonstrate that sub-inhibitory TTO reduces the adhesion of C. albicans to both human cells and polystyrene, inhibits biofilm formation and decreases cell surface hydrophobicity.

This study was conducted to determine the frequencies at which single-step mutants resistant to tea tree oil and rifampicin occurred amongst the Gram-positive organisms Staphylococcus aureus, Staphylococcus epidermidis and Enterococcus faecalis. For tea tree oil, resistance frequencies were very low at

Thirty two new binaphthyl-based, functionalized oxazole and thiazole peptidomimetics and over thirty five novel leucine-containing intermediate oxazoles and thiazoles were prepared in this study. This includes the first examples of the direct C-5 arylation of an amino acid dipeptide-derived oxazole. Moderate to excellent antibacterial activity was observed for all new compounds across Gram positive isolates with MICs ranging from 1-16 μg mL(-1). Results for Gram negative E. coli and A. baumannii were more variable, but MICs as low as 4 μg mL(-1) were returned for two examples. Significantly, the in vitro results with a fluoromethyl-oxazole derivative collectively represent the best obtained to date for a member of our binaphthyl peptide antimicrobials.

The activity of tea tree oil (TTO) and TTO-containing products was investigated according to the EN 1276 and EN 12054 European suspension methods. The activity of different concentrations of TTO, a hygienic skin wash (HSW), an alcoholic hygienic skin wash (AHSW) and an alcoholic hand rub (AHR) was investigated. These formulations were assessed in perfect conditions with the EN 12054 test, and in perfect conditions as well as in the presence of interfering substances with the EN 1276 test, against Staphylococcus aureus, Acinetobacter baumannii, Escherichia coli and Pseudomonas aeruginosa. With the latter test, the activity of the same formulations without TTO was also assessed as a control. With the EN 1276 test, the AHR achieved a >10(5)-fold reduction against all four test organisms within a 1-min contact time. The AHSW achieved a >or=10(5)-fold reduction against A. baumannii after a 1-min contact time and against S. aureus, E. coli and P. aeruginosa after a 5-min contact time. The efficacy of TTO appeared to be dependent on the formulation and the concentration tested, the concentration of interfering substances and, lastly, the organism tested. Nevertheless, 5% TTO achieved a >10(4)-fold reduction in P. aeruginosa cell numbers after a 5-min contact time in perfect conditions. TTO (5%) in 0.001% Tween 80 was significantly more active against E. coli and P. aeruginosa than against S. aureus and A. baumannii. With the EN 12054 test, after a 1-min contact time, 5% TTO in 0.001% Tween 80 and the AHSW achieved a >10(4)-fold reduction in E. coli and A. baumannii cell numbers, respectively, and the AHR achieved a >4 log10 reduction against all organisms tested. The formulations used in this study are now being tested using a novel ex vivo method as well as the in vivo European standard handwashing method EN 1499.

The in vitro activity of Melaleuca alternifolia (tea tree) oil against dermatophytes (n = 106) and filamentous fungi (n = 78) was determined. Tea tree oil MICs for all fungi ranged from 0.004% to 0.25% and minimum fungicidal concentrations (MFCs) ranged from

Globally, scabies affects more than 130 million people at any time. In the developed world, outbreaks in health institutions and vulnerable communities result in a significant economic burden. A review of the literature demonstrates the emergence of resistance toward classical scabicidal treatments and the lack of effectiveness of currently available scabicides in reducing the inflammatory skin reactions and pyodermal progression that occurs in predisposed patient cohorts. Tea tree oil (TTO) has demonstrated promising acaricidal effects against scabies mites in vitro and has also been successfully used as an adjuvant topical medication for the treatment of crusted scabies, including cases that did not respond to standard treatments. Emerging acaricide resistance threatens the future usefulness of currently used gold standard treatments (oral ivermectin and topical permethrin) for scabies. The imminent development of new chemical entities is doubtful. The cumulative acaricidal, antibacterial, antipruritic, anti-inflammatory, and wound healing effects of TTO may have the potential to successfully reduce the burden of scabies infection and the associated bacterial complications. This review summarizes current knowledge on the use of TTO for the treatment of scabies. On the strength of existing data for TTO, larger scale, randomized controlled clinical trials are warranted. Refereed/Peer-reviewed

This study examined the effect of subinhibitory Melaleuca alternifolia (tea tree) essential oil on the development of antibiotic resistance in Staphylococcus aureus and Escherichia coli . Frequencies of single-step antibiotic-resistant mutants were determined by inoculating bacteria cultured with or without subinhibitory tea tree oil onto agar containing 2 to 8 times the MIC of each antibiotic and with or without tea tree oil. Whereas most differences in resistance frequencies were relatively minor, the combination of kanamycin and tea tree oil yielded approximately 10-fold fewer resistant E. coli mutants than kanamycin alone. The development of multistep antibiotic resistance in the presence of tea tree oil or terpinen-4-ol was examined by culturing S. aureus and E. coli isolates daily with antibiotic alone, antibiotic with tea tree oil, and antibiotic with terpinen-4-ol for 6 days. Median MICs for each antibiotic alone increased 4- to 16-fold by day 6. Subinhibitory tea tree oil or terpinen-4-ol did not greatly alter results, with day 6 median MICs being either the same as or one concentration different from those for antibiotic alone. For tea tree oil and terpinen-4-ol alone, day 6 median MICs had increased 4-fold for S. aureus ( n = 18) and 2-fold for E. coli ( n = 18) from baseline values. Lastly, few significant changes in antimicrobial susceptibility were seen for S. aureus and S. epidermidis isolates that had been serially subcultured 14 to 22 times with subinhibitory terpinen-4-ol. Overall, these data indicate that tea tree oil and terpinen-4-ol have little impact on the development of antimicrobial resistance and susceptibility.

The aim of this research was to investigate the activity of a commercial extract derived from the leaves of Olea europaea (olive) against a wide range of microorganisms (n=122). Using agar dilution and broth microdilution techniques, olive leaf extract was found to be most active against Campylobacter jejuni, Helicobacter pylori and Staphylococcus aureus [including meticillin-resistant S. aureus (MRSA)], with minimum inhibitory concentrations (MICs) as low as 0.31-0.78% (v/v). In contrast, the extract showed little activity against all other test organisms (n=79), with MICs for most ranging from 6.25% to 50% (v/v). In conclusion, olive leaf extract was not broad-spectrum in action, showing appreciable activity only against H. pylori, C. jejuni, S. aureus and MRSA. Given this specific activity, olive leaf extract may have a role in regulating the composition of the gastric flora by selectively reducing levels of H. pylori and C. jejuni.

SUMMARYComplementary and alternative medicines such as tea tree (melaleuca) oil have become increasingly popular in recent decades. This essential oil has been used for almost 100 years in Australia but is now available worldwide both as neat oil and as an active component in an array of products. The primary uses of tea tree oil have historically capitalized on the antiseptic and anti-inflammatory actions of the oil. This review summarizes recent developments in our understanding of the antimicrobial and anti-inflammatory activities of the oil and its components, as well as clinical efficacy. Specific mechanisms of antimicrobial and anti-inflammatory action are reviewed, and the toxicity of the oil is briefly discussed.

The essential oil of Melaleuca alternifolia, also known as tea tree or melaleuca oil, is widely available and has been investigated as an alternative antimicrobial, anti-inflammatory and anti-cancer agent. While these properties are increasingly well characterised, relatively limited data are available on the safety and toxicity of the oil. Anecdotal evidence from almost 80 years of use suggests that the topical use of the oil is relatively safe, and that adverse events are minor, self-limiting and occasional. Published data indicate that TTO is toxic if ingested in higher doses and can also cause skin irritation at higher concentrations. Allergic reactions to TTO occur in predisposed individuals and may be due to the various oxidation products that are formed by exposure of the oil to light and/or air. Adverse reactions may be minimised by avoiding ingestion, applying only diluted oil topically and using oil that has been stored correctly. Data from individual components suggest that TTO has the potential to be developmentally toxic if ingested at higher doses, however, TTO and its components are not genotoxic. The limited ecotoxicity data available indicate that TTO is toxic to some insect species but more studies are required.

The efficacy of formulations containing tea tree oil (TTO) has been assessed in vitro in previous studies. Products that passed the European suspension test guidelines were investigated further in this study, in vivo with volunteers using the European handwashing method (EN 1499) and ex vivo using freshly excised human skin samples. The activity of 5% TTO in 0.001% Tween 80, in a hygienic skin wash (HSW) and in an alcoholic hygienic skin wash (AHSW) was investigated and compared with that of a non-medicated soft soap (SS, control). These formulations were assessed against Escherichia coli K12 as recommended by the European standard. In-vivo results showed that 5% TTO in Tween 80 and the AHSW were significantly more active than the SS after 1 min of handwashing. When assessed ex vivo, these two products were also significantly more active than the reference soap after 1 min of rubbing. Both methods showed that 5% TTO in Tween 80 was generally, although not always, more active than a handwash formulation, and that the AHSW was generally more active than the HSW, although this difference was not significant. The formulations tested, as well as the SS, were more active when assessed in vivo than ex-vivo against E. coli, although only the SS and the HSW were significantly more active in vivo. There appeared to be a pattern in the comparison between ex vivo and in vivo results. The antiseptics tested were, on average, 1.28+/-0.06 times more active when assessed in-vivo than when assessed ex vivo. Nevertheless, the main outcome of the European handwashing method is for the formulation tested to be significantly more active than the SS; both 5% TTO in Tween 80 and the AHSW achieved this both in-vivo and ex-vivo. TTO in Tween 80 and in formulations met the European in-vivo method requirements.

The in vitro activities of ketoconazole, econazole, miconazole, and tea tree oil against 54 Malassezia isolates were determined by agar and broth dilution methods. Ketoconazole was more active than both econazole and miconazole, which showed very similar activities. M. furfur was the least susceptible species. M. sympodialis , M. slooffiae , M. globosa , and M. obtusa showed similar susceptibilities to the four agents.

The in-vitro activity of a range of essential oils, including tea tree oil, against the yeast candida was examined. Of the 24 essential oils tested by the agar dilution method against Candida albicans ATCC 10231, three did not inhibit C. albicans at the highest concentration tested, which was 2.0% (v/v) oil. Sandalwood oil had the lowest MIC, inhibiting C. albicans at 0.06%. Melaleuca alternifolia (tea tree) oil was investigated for activity against 81 C. albicans isolates and 33 non-albicans Candida isolates. By the broth microdilution method, the minimum concentration of oil inhibiting 90% of isolates for both C. albicans and non-albicans Candida species was 0.25% (v/v). The minimum concentration of oil killing 90% of isolates was 0.25% for C. albicans and 0.5% for non-albicans Candida species. Fifty-seven Candida isolates were tested for sensitivity to tea tree oil by the agar dilution method; the minimum concentration of oil inhibiting 90% of isolates was 0.5%. Tests on three intra-vaginal tea tree oil products showed these products to have MICs and minimum fungicidal concentrations comparable to those of non-formulated tea tree oil, indicating that the tea tree oil contained in these products has retained its anticandidal activity. These data indicate that some essential oils are active against Candida spp., suggesting that they may be useful in the topical treatment of superficial candida infections.

Objectives: The purpose of this study was to determine the susceptibility of a range of transient and commensal skin flora to the essential oil of Melaleuca alternifolia , or tea tree. Methods: A modified broth microdilution method was used. Polyoxyethylene sorbitan mono-oleate detergent was added to the test medium to enhance solubility of the tea tree oil. Results: Serratia marcescens had the lowest minimum inhibitory concentration (MIC 90 ) of 0.25%. The highest MIC 90 was 3% for Pseudomonas aeruginosa . The lowest minimum bactericidal concentration (MBC 90 ) was 0.25% for S. marcescens and Klebsiella pneumoniae , whereas the highest was 8% for Staphylococcus capitis . Conclusions: S. aureus and most of the gram-negative bacteria tested were more susceptible to tea tree oil than the coagulase-negative staphylocci and micrococci. These results suggest that tea tree oil may be useful in removing transient skin flora while suppressing but maintaining resident flora.

Tea tree oil, an essential oil from the Australian plant Melaleuca alternifolia, has broad antimicrobial activity and is incorporated into a diverse range of pharmaceutical and cosmetic products (2, 3). Blackwell (1) described a patient with typical signs and symptoms of bacterial vaginosis (BV) who treated herself with tea tree oil vaginal pessaries. After treatment, the patient was symptom free and the vaginal flora comprised predominantly gram-positive bacilli. It was suggested that the use of tea tree oil for the treatment of BV be further assessed. Therefore, the aim of this study was to evaluate the activities of tea tree oil against lactobacilli and a range of organisms associated with BV. Reference (n = 9) and stored clinical (n = 24) isolates were obtained from the culture collections of the Western Australian Centre for Pathology and Medical Research (PathCentre) and the Department of Microbiology at The University of Western Australia. Recent clinical isolates (n = 69) were recovered from vaginal swabs submitted to PathCentre and identified according to the Wadsworth Anaerobic Bacteriology Manual (5). Three isolates of Gardnerella vaginalis were kindly provided by Helen MacDonald of the Women’s and Children’s Hospital, Adelaide, Australia. Organisms were tested against doubling dilutions of tea tree oil (batch 93/04, Australian Plantations Pty. Ltd., Wyrallah, Australia) prepared in agar or broth medium. Bacteroides, Prevotella, fusobacteria, and anaerobic gram-positive cocci were tested in supplemented brucella laked-blood agar (5), with 0.5% (vol/vol) Tween 20 added to enhance oil solubility. G. vaginalis and Mobiluncus spp. were tested in Columbia agar base supplemented with 1% Proteose Peptone 3 and 5% chocolatized sheep blood. Lactobacilli (n = 26) were tested by the broth microdilution method using de Man-Rogosa-Sharpe (MRS) broth (Unipath Ltd., Basingstoke, United Kingdom). Selected BV isolates (n = 13) were tested by a broth macrodilution method (5) using brain heart infusion broth (BHIB) supplemented with 1 μg of vitamin K and 5 μg of hemin per ml, with a final test volume of 2 ml. A final concentration of 0.001% (vol/vol) Tween 80 was added to both MRS and supplemented BHIB, to enhance oil solubility. Preparation of inocula, inoculation of susceptibility tests, incubations, and determination of MICs and/or MBCs were carried out as described elsewhere (5). Table ​Table11 lists MIC data obtained by the agar dilution assay. In addition, for five G. vaginalis isolates tea tree oil MICs were 0.06%, and for three of the four Mobiluncus isolates the MICs were 0.03%, with 0.06% for the fourth. By broth macrodilution, MICs for all six Bacteroides isolates were 0.06%. The MBCs for five isolates were 0.06%, while the MBC for the remaining isolate was 0.12%. MICs and MBCs for two Prevotella isolates were 0.03%. The MICs for four of the five Peptostreptococcus anaerobius isolates were 0.06%, with 0.03% for the fifth. MBCs were 0.03% for one isolate, 0.06% for three isolates, and 0.12% for the remaining isolate. MICs for the lactobacilli ranged from 0.12 to 2.0% and the MIC at which 90% of isolates were inhibited (MIC90) was 2.0%. MBCs for lactobacilli ranged from 0.25 to 2.0%, and the MBC90 was 2.0%. TABLE 1 Tea tree oil MICs for organisms associated with BV These MIC data are similar to those obtained in previous studies which indicated that a variety of anaerobic and aerobic bacteria were susceptible to tea tree oil (2–4). The data also show that all lactobacilli tested were appreciably more resistant to tea tree oil than organisms known to be associated with BV, with at least a twofold difference in MIC90 results. Therefore, the clinical success reported by Blackwell (1) may be due, in part, to the susceptibility of BV-associated organisms to tea tree oil and the relative resistance of commensal lactobacilli. This difference in susceptibility may allow formulation of products that will selectively kill or inhibit certain organisms while having a minimal effect on the commensal lactobacilli. Appropriate trials are now urgently needed to determine whether these theoretical benefits will translate into clinical practice.