Spore-Forming Clostridium (Clostridioides) difficile in Wastewater Treatment Plants in Western Australia.

There is growing evidence that shows Clostridium (Clostridioides) difficile is a pathogen of One Health importance with a complex dissemination pathway involving animals, humans, and the environment. Thus, environmental discharge and agricultural recycling of human and animal waste have been suspected as factors behind the dissemination of Clostridium difficile in the community. Here, the presence of C. difficile in 12 wastewater treatment plants (WWTPs) in Western Australia was investigated. Overall, C. difficile was found in 90.5% (114/126) of raw sewage influent, 48.1% (50/104) of treated effluent, 40% (2/5) of reclaimed irrigation water, 100% (38/38) of untreated biosolids, 95.2% (20/21) of anaerobically digested biosolids, and 72.7% (8/11) of lime-amended biosolids. Over half of the isolates (55.3% [157/284]) were toxigenic, and 97 C. difficile ribotypes (RTs) were identified, with RT014/020 the most common (14.8% [42/284]). Thirteen C. difficile isolates with the toxin gene profile A+ B+ CDT+ (positive for genes coding for toxins A and B and the binary C. difficile transferase toxin [CDT]) were found, including the hypervirulent RT078 strain. Resistance to the antimicrobials fidaxomicin, vancomycin, metronidazole, rifaximin, amoxicillin-clavulanate, meropenem, and moxifloxacin was uncommon; however, resistance to clindamycin, erythromycin, and tetracycline was relatively frequent at 56.7% (161/284), 14.4% (41/284), and 13.7% (39/284), respectively. This study revealed that toxigenic C. difficile was commonly encountered in WWTPs and being released into the environment. This raises concern about the possible spillover of C. difficile into animal and/or human populations via land receiving the treated waste. In Western Australia, stringent measures are in place to mitigate the health and environmental risk of recycling human waste; however, further studies are needed to elucidate the public health significance of C. difficile surviving the treatment processes at WWTPs. IMPORTANCE Clostridium difficile infection (CDI) is a leading cause of antimicrobial-associated diarrhea in health care facilities. Extended hospital stays and recurrences increase the cost of treatment and morbidity and mortality. Community-associated CDI (CA-CDI) cases, with no history of antimicrobial use or exposure to health care settings, are increasing. The isolation of clinically important C. difficile strains from animals, rivers, soil, meat, vegetables, compost, treated wastewater, and biosolids has been reported. The objective of this study was to characterize C. difficile in wastewater treatment plants (WWTPs) in Australia. We found that C. difficile can survive the treatment processes of WWTPs, and toxigenic C. difficile was being released into the environment, becoming a potential source/reservoir for CA-CDI.

Clostridioides difficile infection and One Health: an equine perspective.

Clostridioides (Clostridium) difficile presents a significant health risk to humans and animals. The complexity of the bacterial-host interaction affecting pathogenesis and disease development creates an ongoing challenge for epidemiological studies, control strategies and prevention planning. The recent emergence of human disease caused by strains of C. difficile found in animals adds to mounting evidence that C. difficile infection (CDI) may be a zoonosis. In equine populations, C. difficile is a known cause of diarrhoea and gastrointestinal inflammation, with considerable mortality and morbidity. This has a significant impact on both the well-being of the animal and, in the case of performance and production animals, it may have an adverse economic impact on relevant industries. While C. difficile is regularly isolated from horses, many questions remain regarding the impact of asymptomatic carriage as well as optimization of diagnosis, testing and treatment. This review provides an overview of our understanding of equine CDI while also identifying knowledge gaps and the need for a holistic One Health approach to a complicated issue.

Cationic biaryl 1,2,3-triazolyl peptidomimetic amphiphiles targeting Clostridioides (Clostridium) difficile: Synthesis, antibacterial evaluation and an in vivo C. difficile infection model.

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.

Non-conventional antimicrobial and alternative therapies for the treatment of Clostridium difficile infection.

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.

Tea tree oil gel for mild to moderate acne; a 12 week uncontrolled, open-label phase II pilot study.

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.

Infection with Toxin A-Negative, Toxin B-Negative, Binary Toxin-Positive Clostridium difficile in a Young Patient with Ulcerative Colitis.

Large clostridial toxin-negative, binary toxin-positive (A(-) B(-) CDT(+)) strains of Clostridium difficile are almost never associated with clinically significant C. difficile infection (CDI), possibly because such strains are not detected by most diagnostic methods. We report the isolation of an A(-) B(-) CDT(+) ribotype 033 (RT033) strain of C. difficile from a young patient with ulcerative colitis and severe diarrhea.

Effect of habituation to tea tree (Melaleuca alternifolia) oil on the subsequent susceptibility of Staphylococcus spp. to antimicrobials, triclosan, tea tree oil, terpinen-4-ol and carvacrol.

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 µg/mL) to intermediate susceptibility (MIC=3 µg/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.

Tea tree oil (5%) body wash versus standard care (Johnson's Baby Softwash) to prevent colonization with methicillin-resistant Staphylococcus aureus in critically ill adults: a randomized controlled trial.

OBJECTIVES: To determine whether the daily use of 5% tea tree oil (TTO) body wash (Novabac 5% Skin Wash) compared with standard care [Johnson's Baby Softwash (JBS)] had a lower incidence of methicillin-resistant Staphylococcus aureus (MRSA) colonization. PATIENTS: The study setting was two intensive care units (ICUs; mixed medical, surgical and trauma) in Northern Ireland between October 2007 and July 2009. The study population comprised 391 patients who were randomized to JBS or TTO body wash. METHODS: This was a Phase 2/3, prospective, open-label, randomized, controlled trial. TRIAL REGISTRATION: ISRCTN65190967. The primary outcome was new MRSA colonization during ICU stay. Secondary outcomes included the incidence of MRSA bacteraemia and maximum increase in sequential organ failure assessment score. RESULTS: A total of 445 patients were randomized to the study. After randomization, 54 patients were withdrawn; 30 because of a positive MRSA screen at study entry, 11 due to lack of consent, 11 were inappropriately randomized and 2 had adverse reactions. Thirty-nine (10%) patients developed new MRSA colonization (JBS n = 22, 11.2%; TTO body wash n = 17, 8.7%). The difference in percentage colonized (2.5%, 95% CI - 8.95 to 3.94; P = 0.50) was not significant. The mean maximum increase in sequential organ failure assessment score was not significant (JBS 1.44, SD 1.92; TTO body wash 1.28, SD 1.79; P = 0.85) and no study patients developed MRSA bacteraemia. CONCLUSIONS: Compared with JBS, TTO body wash cannot be recommended as an effective means of reducing MRSA colonization.

Topically applied Melaleuca alternifolia (tea tree) oil causes direct anti-cancer cytotoxicity in subcutaneous tumour bearing mice.

BACKGROUND: Melaleuca alternifolia (tea tree) oil (TTO) applied topically in a dilute (10%) dimethyl sulphoxide (DMSO) formulation exerts a rapid anti-cancer effect after a short treatment protocol. Tumour clearance is associated with skin irritation mediated by neutrophils which quickly and completely resolves upon treatment cessation. OBJECTIVE: To examine the mechanism of action underlying the anti-cancer activity of TTO. METHODS: Immune cell changes in subcutaneous tumour bearing mice in response to topically applied TTO treatments were assessed by flow cytometry and immunohistochemistry. Direct cytotoxicity of TTO on tumour cells in vivo was assessed by transmission electron microscopy. RESULTS: Neutrophils accumulate in the skin following topical 10% TTO/DMSO treatment but are not required for tumour clearance as neutrophil depletion did not abrogate the anti-cancer effect. Topically applied 10% TTO/DMSO, but not neat TTO, induces an accumulation and activation of dendritic cells and an accumulation of T cells. Although topical application of 10% TTO/DMSO appears to activate an immune response, anti-tumour efficacy is mediated by a direct effect on tumour cells in vivo. The direct cytotoxicity of TTO in vivo appears to be associated with TTO penetration. CONCLUSION: Future studies should focus on enhancing the direct cytotoxicity of TTO by increasing penetration through skin to achieve a higher in situ terpene concentration. This coupled with boosting a more specific anti-tumour immune response will likely result in long term clearance of tumours.

Effects of Melaleuca alternifolia (tea tree) essential oil and the major monoterpene component terpinen-4-ol on the development of single- and multistep antibiotic resistance and antimicrobial susceptibility.

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.

Survey of the antimicrobial activity of commercially available Australian tea tree (Melaleuca alternifolia) essential oil products in vitro.

OBJECTIVES: The aim of this study was to investigate the antimicrobial activity of a range of commercially available tea tree oil (TTO) products and to evaluate whether formulation plays a significant part in their antiseptic activity. METHODS: The antimicrobial activity of the purchased products and control TTO solutions was assessed against Escherichia coli, Staphylococcus aureus, Salmonella typhimurium, Pseudomonas aeruginosa, and Candida albicans using well diffusion, broth microdilution, and broth macrodilution assays. RESULTS: Zone sizes obtained by the agar well diffusion assay ranged from 0 to 49.8 mm, with the more viscous and lipophilic products producing the smallest zones. Micro- and macrodilution methods showed that eight products had minimum inhibitory concentrations that were lower than the nonformulated TTO control. The remaining three products showed activity equivalent to the TTO control. CONCLUSIONS: In general, the commercially available antiseptic TTO products showed antimicrobial activity that was equivalent to, or greater than the nonformulated TTO control. This suggests that the TTO within these products has retained its antimicrobial activity. Furthermore, the enhanced activity of the products may be attributed to other antimicrobial excipients within the products such as preservatives, or to synergistic antimicrobial interactions between the TTO and other product excipients. The observation that the commercially available antiseptic TTO products tested in this study retained adequate antimicrobial activity emphasizes the importance of considering how product bases and excipients may interact with the active compound during formulation to ensure efficacy of the final product. Finally, the current data suggest that these TTO products may also be active in vivo. However, this can only be determined through further studies and in clinical trials.

Uncontrolled, open-label, pilot study of tea tree (Melaleuca alternifolia) oil solution in the decolonisation of methicillin-resistant Staphylococcus aureus positive wounds and its influence on wound healing.

Many complementary and alternative products are used to treat wounds. The essential oil of Melaleuca alternifolia, tea tree oil, has proven antimicrobial and anti-inflammatory properties, may be useful in methicillin-resistant Staphylococcus aureus (MRSA) decolonisation regimens and is reputed to have 'wound-healing' properties, but more data are required to support these indications. The primary aim of this uncontrolled case series was to assess whether a tea tree oil solution used in a wound cleansing procedure could decolonise MRSA from acute and chronic wounds of mixed aetiology. The secondary aim was to determine if the tea tree oil solution influenced wound healing outcomes. Nineteen participants with wounds suspected of being colonised with MRSA were enrolled in a pilot study. Seven were subsequently shown not to have MRSA and were withdrawn from the study. As many as 11 of the remaining 12 participants were treated with a water-miscible tea tree oil (3·3%) solution applied as part of the wound cleansing regimen at each dressing change. Dressing changes were three times per week or daily as deemed necessary by the study nurse following assessment. One participant withdrew from the study before treatment. No participants were MRSA negative after treatment. After treatment had been implemented, 8 of the 11 treated wounds had begun to heal and reduced in size as measured by computer planimetry. Although this formulation and mode of delivery did not achieve the primary aim of the study, tea tree oil did not appear to inhibit healing and the majority of wounds reduced in size after treatment.

Inhibition of established subcutaneous murine tumour growth with topical Melaleuca alternifolia (tea tree) oil.

PURPOSE: Systemic toxicity coupled with long treatment regimes of approved topical chemotherapeutic agents such as imiquimod and 5-fluorouracil (5-FU) are limiting. There is now more focus on the potential use of topical terpene agents as skin cancer treatments. Here, we show for the first time that topical Melaleuca alternifolia (tea tree) oil (TTO), abundant in terpenes, has in vivo antitumour activity. METHOD: Topical TTO formulations applied to immunocompetent tumour-bearing mice were assessed for antitumour efficacy by monitoring tumour growth and by histological analysis following treatment. RESULTS: Four, daily, topical treatments of 10% TTO/DMSO regressed subcutaneous AE17 mesotheliomas in mice for a period of 10 days and significantly retarded the growth of subcutaneous B16-F10 melanomas. The antitumour effect of topical 10% TTO/DMSO was accompanied by skin irritation similar to other topical chemotherapeutic agents, but unlike other approved topical agents, quickly and completely resolved. Furthermore, we show that topical 10% TTO/DMSO caused an influx of neutrophils and other immune effector cells in the treated area, with no evidence of systemic toxicity. CONCLUSION: TTO combined with an effective carrier significantly inhibited the growth of aggressive, subcutaneous, chemo-resistant tumours in immunocompetent mice. Taken together, these findings highlight the potential of topical TTO as an alternative topical antitumour treatment.

Bacterial and dye penetration through interim restorations used during endodontic treatment of molar teeth.

INTRODUCTION: The purpose of this study was to investigate the association between dye and bacterial penetration through interim restorations used during endodontic treatment. METHODS: Sixty-four extracted human teeth were used, with 2 teeth each as positive and negative controls. Endodontic access with a mesio-occluso-distal cavity was prepared. Palatal cusps of maxillary molars and buccal cusps of mandibular molars were removed. Cotton was placed over the canals and covered with Cavit. Thirty teeth were restored with Ketac Silver (KS) and 30 with KS reinforced with a stainless steel band (KSSB). Samples were submersed in India ink mixed with brain heart infusion broth containing Streptococcus gordonii. After 3 months of simulated chewing, structural integrity and dye and bacterial penetration were assessed. RESULTS: Positive controls had both dye and bacterial penetration. Negative controls had no dye or bacterial penetration. All KS restorations debonded, whereas 18 KSSB restorations (60%) debonded. KS restorations were 1.67 times more likely to debond than KSSB restorations (Fisher exact test). KS was 1.3 times more likely to have dye penetration than KSSB (Fisher exact test) and 3 times more likely to have bacterial penetration, although not statistically significant (chi(2) test). Overall, 88.3% of specimens had dye penetration, and 20% had bacterial penetration. This 68.3% difference indicated no association between dye and bacterial penetration (exact McNemar test). CONCLUSIONS: Stainless steel bands helped maintain structural integrity of KS restorations under masticatory function. Bands helped prevent dye penetration but not bacterial penetration. There was no association between dye and bacterial penetration.

Antimicrobial activity of commercial Olea europaea (olive) leaf extract.

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.

Antimicrobial and anti-inflammatory activity of five Taxandria fragrans oils in vitro.

The antimicrobial activity of five samples of Taxandria fragrans essential oil was evaluated against a range of Gram-positive (n= 26) and Gram-negative bacteria (n= 39) and yeasts (n= 10). The majority of organisms were inhibited and/or killed at concentrations ranging from 0.06-4.0% v/v. Geometric means of MIC were lowest for oil Z (0.77% v/v), followed by oils X (0.86%), C (1.12%), A (1.23%) and B (1.24%). Despite differences in susceptibility data between oils, oils A and X did not differ when tested at 2% v/v in a time kill assay against Staphylococcus aureus. Cytotoxicity assays using peripheral blood mononuclear cells demonstrated that T. fragrans oil was cytotoxic at 0.004% v/v but not at 0.002%. Exposure to one or more of the oils at concentrations of

Frequencies of resistance to Melaleuca alternifolia (tea tree) oil and rifampicin in Staphylococcus aureus, Staphylococcus epidermidis and Enterococcus faecalis.

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 <10(-9). Single-step mutants resistant to tea tree oil were undetectable at two times the minimum inhibitory concentration (MIC) for S. aureus RN4220 and derivative mutator strains or at 3 x MIC for the remaining S. aureus strains, including a clinical meticillin-resistant S. aureus isolate. Similarly, no mutants were recovered at 2x MIC for S. epidermidis or at 1x MIC for E. faecalis. Resistance frequencies determined in vitro for rifampicin (8 x MIC) ranged from 10(-7) to 10(-8) for all isolates, with the exception of the S. aureus mutator strains, which had slightly higher frequencies. These data suggest that Gram-positive organisms such as Staphylococcus and Enterococcus spp. have very low frequencies of resistance to tea tree oil.

Role of the MexAB-OprM efflux pump of Pseudomonas aeruginosa in tolerance to tea tree (Melaleuca alternifolia) oil and its monoterpene components terpinen-4-ol, 1,8-cineole, and alpha-terpineol.

Using a series of efflux mutants of Pseudomonas aeruginosa, the MexAB-OprM pump was identified as contributing to this organism's tolerance to the antimicrobial agent tea tree (Melaleuca alternifolia) oil and its monoterpene components terpinen-4-ol, 1,8-cineole, and alpha-terpineol. These data show that a multidrug efflux system of P. aeruginosa can extrude monoterpenes and related alcohols.

Use of deception to achieve double-blinding in a clinical trial of Melaleuca alternifolia (tea tree) oil for the treatment of recurrent herpes labialis.

Double-blinding is an important and widely implemented feature of clinical trials although its success is rarely assessed. In a randomized, placebo-controlled trial of tea tree oil, an aromatic essential oil, for the treatment of recurrent herpes labialis (RHL), or cold sores, deception was used to prevent volunteers from identifying their treatment allocation. Volunteers received placebo (n=102) or tea tree oil (n=112) ointment in preparation for their next episode of RHL and were told, falsely, that the aroma of the ointments had been changed to prevent identification of the treatment group. At the trial's end, of the volunteers who had used their ointment and presented for treatment assessment (n=100), approximately 50% correctly guessed their treatment allocation (P=0.774). Amongst volunteers that had not presented for treatment assessment (n=114), 12 volunteers did not provide blinding data and 46 did not open their tube. For the 56 volunteers who opened their tube, less than half of those receiving tea tree oil (44.4%) and only a small proportion of those on placebo (17.2%) were able to correctly identify their treatment allocation. Among the volunteers that were not treated, the P-value was 0.083. This study showed that the ethical use of deception may provide effective blinding in challenging circumstances.