Antifungal Activity of Some Essential Oils against Multi-Resistant Candida Strains

Resistance to oral antifungal drugs, such as fluconazole, ketoconazole, itraconazole for example, becomes a problem when the drugs are used or as prophylaxis or for frequent short-term exposures. Candida is a common fungus that is normally controlled by the immune system, but when immune system is suppressed, Candida can grow on mucous membranes or elsewhere in the body, causing symptoms known as candidiasis. HIV positive and HIV negative people may experience candidiasis and more and more strains become resistant to antifungal drugs. Very recent studies (Luqman et al.,2007) demonstrate that rosemary oil is a potential alternative compound to treat drug-resistant infections. Zambonelli (2004) demonstrated the fungicidal activity of Thymus vulgaris essential oil and the data is confirmed by the Italian Society of Gynecology and Obstetrics. In the present study we performed a screening on a series of essential oils with the aim to find new antifungal agents. Materials and Methods: Essential oils from Anethus graveolens, Mentha piperita, Rosmarinus officinalis and Thymus vulgaris were tested against multi-resistant strains of Candida (albicans, parapsilosis, krusei, glabrata, tropicalis, dubliniensis and guillermondi) from clinical specimens from HIV-seropositive subjects kindly supplied by the Department of Medicine Laboratory-Ospedale Policlinico-Modena. Qualitative analysis of the essential oils was performed by a gas chromatograph 6890-N, quantitative analysis by a gas chromatograph DANI 86-10 and a flame ionization detector (FID). Strains were cultured in Sabouraud Dextrose Liquid Medium (pH 5,6 ± 0,2 at 25°C). Fluconazole was chosen as reference drug. Sensitivity tests were performed in according with the NCCLS method for agar dilution and MIC values (minimal inhibitory concentration) were calculated. Results: Essential oils, characterized by variable percentages of p-cymene, gamma therpinene, tymol, sabinene and 1-8 cyneol, were diluted, added to Sabouraud dextrose liquid medium and evaluated for the antifungal activity in comparison with fluconazole (dilution from 200 to 0.1 mg/l). Thymus, Mentha and Rosmarinus appear to be the most active oils since the antifungal activity is maintained with very low dilution (>1000 of the mother solution) over 24 h and against the most of strains but in particular against C. albicans, C. tropicalis and C. krusei this latter responsible of fungal infections in newborns and small infants. On the same strains, fluconazole MIC is >100mg/l thus showing the presence of strong resistance to the drug. The very high percentage of 1-8 cyneol (14-25%), in Rosmarinus oil could represent a safety index in that the molecule is non toxic on Vero cell cultures, but the data is referred to the isolated compound. Conclusion: At the present we are unable to identify a compound responsible of the anti-fungal activity. No compound is present at the same percentage in all the oils we studied. The results we observed may be due or to the phytocomplex or to a molecule present even at very low concentration. The phytocomplex could reduce the toxicity of isolated compounds as camphora, camphene or alpha pynene. The interesting result is the very good antifungal activity exerted at very low dilutions. Further studies are to be performed to investigate both on the mechanism of action and to the presence of side effects.