ANTIFUNGAL ACTIVITY OF EUCALYPTUS OIL AGAINST CLINICAL ISOLATES OF CANDIDA SPECIES

The majority of mycotic infections are due to Candida species, out of which Candida albicans is being the most frequently isolated species. Eucalyptus is one of the very important and most widely planted genera across the world. It is a tall, evergreen tree, native to Australia and Tasmania, successfully introduced worldwide, now extensively planted in many other countries. Thus, the aim of the present study was to determine the antifungal activity of eucalyptus oil against clinical isolates of Candida spp. We have observed that, clinical isolates of Candida species were not inhibited in any of these dilutions of this oil. The eucalyptus oil does not have antifungal activity against Candida species. However, it is important to include more number of isolates and different dilutions to validate the results.


INTRODUCTION
The majority of mycotic infections are due to Candida species, out of which Candida albicans is being the most frequently isolated species [1] . Various virulence factors plays a crucial role in the pathogenicity of several microorganisms including Candida albicans. [2,3,4] .
Eucalyptus is one of the very important and most widely planted genera across the world. [6] It is a tall, evergreen tree, native to Australia and Tasmania, successfully introduced worldwide, now extensively planted in many other countries . [5] It was introduced in Algeria in 1854 by Ramel. [8] Eucalyptus species are well known as medicinal plants due to their biological and pharmacological properties. In the international pharmacopeia, the most important and represented species, however, is Eucalyptus globulus (E. globulus) which is the main furnisher of essential oils. These essential oils have different applications as anesthetic, anodyne, antiseptic, astringent, deodorant, diaphoretic, disinfectant, expectorant, febrifuge, fumigant, hemostat, inhalant, insect repellant, preventitive, rubefacient, sedative yet stimulant, vermifuge, for a folk remedy for abscess, arthritis, asthma, boils, bronchitis, burns, cancer, diabetes, diarrhea, diphtheria, dysentery, encephalitis, enteritis, erysipelas, fever, flu, inflammation, laryngalgia, laryngitis, leprosy, malaria, mastitis, miasma, pharygnitis, phthisis, rhinitis, sores, sore throat, spasms, trachalgia, worms, and wounds. Sometimes their demand is also high in the soap and cosmetic industries. [7] . Thus, the aim of the present study was to determine the antifungal activity of eucalyptus oil against clinical isolates of Candida spp.

Candida isolates
A total of 20 non repetitive clinical isolates of Candidaspecies were collected from different samples of immunocompromised individuals attending Saveetha Medical College, Thandalam. They were characterized by carbohydrate fermentation and assimilation tests and confirmed. [7] Isolates were preserved in semisolid Sabouraud chloramphenicol semi solid stock and stored at 4˚C until further use.

Characterization of Candida species
Candida species were further characterized by using Hichrom agar (Himedia, Mumbai). gently to dissolve the medium completely. Then it was allowed to cool to 50 o C and rehydrated (one vial) contents of Hichrome Candida selective supplement was added under aseptic precautions. It was mixed well and poured into petridishes. Isolates were identified on Hichrome agar based upon the characteristic color of the colony by subculturing from Sabouraud's chloramphenicol agar plates and the Candida Hichrome plates were incubated at 37 o C for 24-48 hours [9] . Based on colour produced by the isolates speciation have been made.

Detection of antibacterial activity of eucalyptus oil against clinical isolates of Candida spp
Antifungal activity of eucalyptus oil was tested against Candida spp isolates by minimum inhibitory concentration method. Mueller Hinton broth was supplemented with 0.002% (V/V) tween 80 (HiMedia, Mumbai) to enhance the dispersion of the essential oil. Agar dilution method was performed to attain the different concentrations of essential oils such as 0.03%, 0.06%, 0.125%, 0.25%, 0.5%, 1% and 2% in Mueller Hinton Agar (MHA).
Media containing various concentrations of essential oils were poured over the sterile petridishes and allowed to dry. Media without essential oil was served as control plate. Spot inoculation of 0.5 McFarland standard turbidity adjusted isolates were made on the plates and incubated at 37ºC for overnight. The lowest concentration of the essential oils that completely inhibited the growth of isolates was considered as MIC. [10]

Characterization of Candida species by Hichrom Candida agar
A total of 20 clinical isolates of Candidaspp were seeded on to Hichrom Candida agar and results were tabulated based on pigment production.

Result of antifungal activity of eucalyptus oil against clinical isolates of Candida species
We have observed that, clinical isolates of Candida species were not inhibited in any of these dilutions of this oil.
[10]. In contrast, our isolates did not show any inhibitory activity against eucalyptus oil with these dilutions.

CONCLUSION
The eucalyptus oil does not have antifungal activity against Candida species. However, it is important to include more number of isolates and different dilutions to validate the results.