Antimicrobial effects of Melaleuca alternifolia (tea tree) essential oil against biofilm-forming multidrug-resistant cystic fibrosis-associated Pseudomonas aeruginosa as a single agent and in combination with commonly nebulized antibiotics.

Broth microdilution assays were used to determine minimum inhibitory concentrations (MICs) and fractional inhibitory concentration indices (FICIs) of tea tree oil (TTO), tobramycin, colistin and aztreonam (ATM) against clinical cystic fibrosis-associated Pseudomonas aeruginosa (CFPA) isolates (n = 20). The minimum biofilm eradication concentration (MBEC) and fractional biofilm eradication concentration index (FBECI) were also determined using a similar microbroth dilution checkerboard assay, with biofilms formed using the MBEC device® . TTO was effective at lower concentrations against multidrug-resistant (MDR) CFPA isolates (n = 3) in a biofilm compared to in a planktonic state (MBEC 18·7-fold lower than MIC). CFPA within biofilm was less susceptible to ATM, colistin and tobramycin compared to planktonic cells (MBEC 6·3-fold, 9·3-fold, and 2·1-fold higher than MIC respectively). All combinations of essential oil and antibiotic showed indifferent relationships (FICI 0·52-1·72) when tested against planktonic MDR CFPA isolates (n = 5). Against CFPA isolates (n = 3) in biofilm, combinations of TTO/aztreonam and TTO/colistin showed indifferent relationships (mean FBECI 0·85 and 0·60 respectively), whereas TTO/tobramycin showed a synergistic relationship (mean FBECI 0·42). The antibiofilm properties of TTO and the synergistic relationship seen between TTO and tobramycin against CFPA in vitro make inhaled TTO a promising candidate as a potential therapeutic agent.

Natural products show diverse mechanisms of action against Clostridium difficile.

AIMS: To investigate the mechanisms of action of natural products with bactericidal (cinnamon root powder, peppermint oil, trans-cinnamaldehyde, menthol and zingerone) or bacteriostatic (fresh garlic bulb extract, garlic clove powder, Leptospermum honey and allicin) activity against two Clostridium difficile strains. METHODS AND RESULTS: Bactericidal products significantly reduced intracellular ATP after 1 h (P ≤ 0·01), quantified using the BacTiter-Glo reagent, and damaged the cell membrane, shown by the leakage of both 260-nm-absorbing materials and protein, and the uptake of propidium iodide. Bacteriolysis was not observed, determined by measuring optical density of treated cell suspensions at 620-nm. The effect of three bacteriostatic products on protein synthesis was quantified using an Escherichia coli S30 extract system, with Leptospermum honey (16% w/v) showing significant inhibition (P < 0·01). Lastly, no products showed elevated minimum inhibitory concentrations against antimicrobial-resistant C. difficile, determined by broth microdilution. CONCLUSIONS: Cytoplasmic membrane damage was identified as a mechanism of action that may contribute to the activity of several natural products against C. difficile. SIGNIFICANCE AND IMPACT OF THE STUDY: This study describes the possible mechanisms of action of natural products against C. difficile, yet the efficacy in vivo to be determined.

Effects of natural products on several stages of the spore cycle of Clostridium difficile in vitro.

AIMS: To investigate the effect of natural products on the spore cycle of Clostridium difficile in vitro. METHODS AND RESULTS: Twenty-two natural products were investigated using four C. difficile strains. Effects on sporulation, determined using microscopy and a conventional spore recovery assay, showed that fresh onion bulb extract (6·3% v v-1 ) and coconut oil (8% v v-1 ) inhibited sporulation in all four isolates by 66-86% and 51-88%, respectively, compared to untreated controls. Fresh ginger rhizome extract (25% v v-1 ) was also inhibitory, although to a lesser extent. Using a standard spore germination and outgrowth assay, germination was unaffected by the 22 products, whereas outgrowth was significantly reduced by artichoke extract (18·8 mg ml-1 ), fresh onion bulb extract (25% v v-1 ), Leptospermum honeys (8% w v-1 ) and allicin (75 mg ml-1 ; P < 0·01). Sporicidal activity, investigated using a standard plate recovery assay, was minimal. CONCLUSIONS: Three of the 22 natural products (13%) showed inhibitory effects on sporulation of C. difficile and six products (27%) reduced vegetative outgrowth of C. difficile. SIGNIFICANCE AND IMPACT OF THE STUDY: This study shows the potential of natural products to inhibit different stages of C. difficile sporulation and encourages further investigation in this field.

Antimicrobial activity of natural products against Clostridium difficile in vitro.

AIMS: To investigate the antimicrobial activity of various natural products against Clostridium difficile in vitro. METHODS AND RESULTS: The antibacterial activity of 20 natural products was determined by the agar well diffusion and broth microdilution assays against four C. difficile strains, three comparator organisms and four gastrointestinal commensal organisms. Of the raw natural products, garlic juice had the highest activity. The most active processed products were peppermint oil and the four pure compounds trans-cinnamaldehyde, allicin, menthol and zingerone. Furthermore, Bacteroides species had similar susceptibility to C. difficile to most natural products; however, Lactobacillus casei was less susceptible. The combined effect of natural products with vancomycin or metronidazole was determined using the conventional checkerboard titration method and the fractional inhibitory concentration index was calculated. The results showed a possible synergism between trans-cinnamaldehyde and vancomycin and partial synergy between trans-cinnamaldehyde and metronidazole. CONCLUSIONS: The study indicates a range of antimicrobial activity of natural products against C. difficile and suggests that they may be useful as alternative or complementary treatments for C. difficile infection (CDI), particularly as most are able to be given orally. SIGNIFICANCE AND IMPACT OF THE STUDY: This study encourages further investigation of natural products for treatment of CDI.

Antimicrobial activity of honey from the stingless bee Trigona carbonaria determined by agar diffusion, agar dilution, broth microdilution and time-kill methodology.

AIMS: The aim of this study was to determine the spectrum of antimicrobial activity of 11 samples of stingless bee honey compared to medicinal, table and artificial honeys. METHODS AND RESULTS: Activity was assessed by agar diffusion, agar dilution, broth microdilution and time-kill viability assays. By agar dilution, minimum inhibitory concentration (MIC) ranges were 4% to >10% (w/v) for Gram-positive bacteria, 6% to >16% (w/v) for Gram-negative bacteria and 6% to >10% (w/v) for Candida spp. By broth microdilution, all organisms with the exception of Candida albicans and Candida glabrata were inhibited at 3 log for Pseudomonas aeruginosa and <1 log for C. albicans. Similar treatment with each control honey resulted in decreases of <1 log for all organisms. CONCLUSIONS: Stingless bee honey has broad-spectrum antibacterial activity although activity against Candida was limited. Stingless bee honey samples varied in activity and the basis for this remains to be determined. SIGNIFICANCE AND IMPACT OF THE STUDY: Stingless bee honey had similar activity to medicinal honey and may therefore have a role as a medicinal agent.

A review of the toxicity of Melaleuca alternifolia (tea tree) oil.

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.

Assessment of the antibacterial activity of tea tree oil using the European EN 1276 and EN 12054 standard suspension tests.

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.

Effectiveness of hand-cleansing formulations containing tea tree oil assessed ex vivo on human skin and in vivo with volunteers using European standard EN 1499.

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.

Treatment of acne with tea tree oil (melaleuca) products: a review of efficacy, tolerability and potential modes of action.

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.

Susceptibility of transient and commensal skin flora to the essential oil of Melaleuca alternifolia (tea tree oil).

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 (MIC90) of 0.25%. The highest MIC90 was 3% for Pseudomonas aeruginosa. The lowest minimum bactericidal concentration (MBC90) 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 staphylococci and micrococci. These results suggest that tea tree oil may be useful in removing transient skin flora while suppressing but maintaining resident flora.

Gastric partitioning for morbid obesity: postoperative weight loss, technical complications, and protein status.

The study of 325 patients who underwent gastric partitioning (stapling) was undertaken to assess the complications and weight-loss records over a 5-year period. A subgroup of 15 patients was studied to determine protein status preoperatively and 2, 4, and 6 months after surgery. Laboratory tests, anthropometric measures, and food records were used to assess patient health and nutritional status. Four operative techniques have been used by a surgeon in Reno, NV, since 1979. The main postoperative complication in the first two procedures, a nonreinforced horizontal staple line (Groups 1 and 2), was staple line disruption and, therefore, poor weight-loss results. The staple line was reinforced in the next 193 patients (Group 3), and the patients followed a blenderized diet for 8 weeks postoperatively. Persistent vomiting was the most common problem in that group; 24% required dilatation of the stoma via gastroscopy. The vertical staple procedure was adopted for the next 48 patients (Group 4) to further minimize disruption and severe vomiting. Other operative complications were relatively few. At 48 and 18 months, respectively, Groups 3 and 4 showed an average weight loss of 27% of the preoperative weight. Eating patterns and food tolerances changed dramatically postoperatively, and protein intake dropped significantly. Mean protein intakes below standard recommended dietary allowance (RDA) were observed in a subgroup of Groups 3 and 4. However, the mean total lymphocyte count and albumin values demonstrated that patients were not compromising visceral proteins. The mean creatinine height indexes and arm muscle areas showed no significant difference in somatic protein status.(ABSTRACT TRUNCATED AT 250 WORDS)

Chemical characteristics and antimicrobial effects of some Eucalyptus kinos.

ETHNOPHARMACOLOGICAL RELEVANCE: Eucalyptus kinos are tannin-rich, mostly red-coloured wood exudates. They have played an important role in the traditional medicines of Australian Aboriginal people and were also a valued source of antibacterial and astringent agents for early European settlers. MATERIALS AND METHODS: Nineteen different Eucalyptus kinos were collected and analysed for their total phenolics and total tannin content as well as their relative amounts of hydrolysable and condensed tannins. They were also classified in accordance with Maiden's traditional kino categories. Well plate diffusion assays using three Gram positive and two Gram negative bacteria and a yeast species were carried out to assess antimicrobial properties. RESULTS: The investigated kino samples differ strongly in their total phenolics and overall tannin as well as their relative hydrolysable and condensed tannin contents. All but one could be assigned to one of the traditional Maiden kino classes. The samples, in particular those collected from Corymbia maculata and Eucalyptus ficifolia, demonstrated a strong antibacterial activity towards Gram positive bacteria but were inactive against the Gram negative strains and the yeast. No obvious correlation seems to exist between a particular Maiden class and antibacterial activity but there is a positive correlation between total phenolics/tannin content and antibacterial effect although two of the investigated kinos (Eucalyptus flocktoniae and Eucalyptus sargentii) deviated from this trend. The relative amounts of hydrolysable and condensed tannins in a kino sample do not seem to determine the antibacterial effect. CONCLUSION: Eucalpytus kinos present an interesting class of natural products which should be investigated further, not only to contribute to the growing field of tannin chemistry but to also learn more about the individual role played by the various hydrolysable and condensed tannins that determine a kino's antibacterial activity and to contribute to a better understanding of the use of some of these kinos in traditional systems of medicine. In particular samples like Eucalyptus flocktoniae kino, which recorded a higher antibacterial activity than predicted based on total tannin content, warrant more detailed chemical and antimicrobial analyses.

Melaleuca alternifolia (Tea Tree) oil: a review of antimicrobial and other medicinal properties.

Complementary 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.

Precipitate production by some Malassezia species on Dixon's agar.

The production of a precipitate by Malassezia species on Dixon's agar was observed. Malassezia furfur (n = 12), M. obtusa (n = 2) and M. slooffiae (n = 3) were precipitate negative, while M. sympodialis (n = 32) and M. globosa (n = 6) were precipitate positive. This test may be useful in differentiating Malassezia species.

The participative method of subject definition as used in the quantitative modelling of hospital laundry services.

The objectives for developing the participative method of subject definition were to gain all the relevant information to a high level of fidelity in the earliest stages of the work and so be able to build a realistic model at reduced labour cost. In order to better integrate the two activities--information acquisition and mathematical modelling--a procedure was devised using the methods of interactive management to facilitate teamwork. This procedure provided the techniques to create suitable working relationships between the two groups, the informants and the modellers, so as to maximize their free and accurate intercommunication, both during the initial definition of the linen service and during the monitoring of the accuracy and reality of the draft models. The objectives of this project were met in that the final model was quickly validated and approved, at a low labour cost.

In vitro susceptibility of Malassezia furfur to the essential oil of Melaleuca alternifolia.

The susceptibility of 64 Malassezia furfur isolates to Melaleuca alternifolia oil was determined. The minimum inhibitory concentration for 90% of isolates was 0.25% by agar dilution and 0.12% by broth dilution. These data indicate that tea tree oil may be useful in the treatment of skin conditions involving M. furfur.

Antifungal effects of Melaleuca alternifolia (tea tree) oil and its components on Candida albicans, Candida glabrata and Saccharomyces cerevisiae.

OBJECTIVES: The aim of this study was to investigate the mechanism of action of tea tree oil and its components against Candida albicans, Candida glabrata and Saccharomyces cerevisiae. METHODS: Yeast cells were treated with tea tree oil or components, at one or more concentrations, for up to 6 h. During this time, alterations in permeability were assessed by measuring the leakage of 260 nm absorbing materials and by the uptake of Methylene Blue dye. Membrane fluidity was measured by 1,6-diphenyl-1,3,5-hexatriene fluorescence. The effects of tea tree oil on glucose-induced medium acidification were quantified by measuring the pH of cell suspensions in the presence of both tea tree oil and glucose. RESULTS: The treatment of C. albicans with tea tree oil and components at concentrations of between 0.25 and 1.0% (v/v) altered both permeability and membrane fluidity. Membrane fluidity was also increased when C. albicans was cultured for 24 h with 0.016%-0.06% (v/v) tea tree oil, as compared with control cells. For all three organisms, glucose-induced acidification of the external medium was inhibited in a dose-dependent manner in the presence of 0.2%, 0.3% and 0.4% tea tree oil. CONCLUSIONS: Data from this study support the hypothesis that tea tree oil and components exert their antifungal actions by altering membrane properties and compromising membrane-associated functions.

In-vitro activity of essential oils, in particular Melaleuca alternifolia (tea tree) oil and tea tree oil products, against Candida spp.

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.

Antimicrobial activity of essential oils and other plant extracts.

The antimicrobial activity of plant oils and extracts has been recognized for many years. However, few investigations have compared large numbers of oils and extracts using methods that are directly comparable. In the present study, 52 plant oils and extracts were investigated for activity against Acinetobacter baumanii, Aeromonas veronii biogroup sobria, Candida albicans, Enterococcus faecalis, Escherichia col, Klebsiella pneumoniae, Pseudomonas aeruginosa, Salmonella enterica subsp. enterica serotype typhimurium, Serratia marcescens and Staphylococcus aureus, using an agar dilution method. Lemongrass, oregano and bay inhibited all organisms at concentrations of < or = 2.0% (v/v). Six oils did not inhibit any organisms at the highest concentration, which was 2.0% (v/v) oil for apricot kernel, evening primrose, macadamia, pumpkin, sage and sweet almond. Variable activity was recorded for the remaining oils. Twenty of the plant oils and extracts were investigated, using a broth microdilution method, for activity against C. albicans, Staph. aureus and E. coli. The lowest minimum inhibitory concentrations were 0.03% (v/v) thyme oil against C. albicans and E. coli and 0.008% (v/v) vetiver oil against Staph. aureus. These results support the notion that plant essential oils and extracts may have a role as pharmaceuticals and preservatives.

Broth micro-dilution method for determining the susceptibility of Escherichia coli and Staphylococcus aureus to the essential oil of Melaleuca alternifolia (tea tree oil).

A broth micro-dilution method was used to examine the susceptibility of Escherichia coli (n = 110) and Staphylococcus aureus (n = 105) to the essential oil of Melaleuca alternifolia (tea tree oil). The detergent Tween 80 was used successfully to enhance the solubility of tea tree oil in the test medium. The MIC90 of tea tree oil for E. coli was 0.25% while for S. aureus it was 0.50%.