Advances in Environmental Biology

Advances in Environmental Biology, 8(13) August 2014, Pages: 506-511
AENSI Journals
Advances in Environmental Biology
ISSN-1995-0756
EISSN-1998-1066
Journal home page: http://www.aensiweb.com/AEB/
In Vitro comparison of the Effects of Ginger Extract, Fluconazole and Nystatin on
Candida glabrata and Candida kruzei
1Ali
Taghavi zenouz, 2Masoumeh Mehdipour,
Abdollahian
1Solmaz
Pourzare*,
3Mohammad
Adibpour,
1Tahmoores
1
Department of Oral Medicine, Faculty of Dentistry, Tabriz University of Medical Sciences, Tabriz, Iran.
Department of Oral Medicine, Faculty of Dentistry, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
3
Department of Mycology, Tabriz University of Medical Sciences, Tabriz, Iran.
2
ARTICLE INFO
Article history:
R Received 25 April 2014
Received in revised form
8 May 2014
Accepted 20 July 2014
Available online 18 August 2014
Keywords:
Ginger extract, candida glabrata,
candida kruzei, growth inhibition
diameter
ABSTRACT
Objective: At recent years, some studies reported the microbial resistance of candida
specie against antifungal agents. Therefore this study was aimed to evaluate the
antifungal property of ginger extract on candida glabrata and kruzei and compare its
effect with common antifungal agents. Materials & Methods: In this experimental
study, after preparation of ginger extract in MIC concentration, four discs containing
ginger extract, nystatin, fluoconazole, and the blank disc were placed on plates
containing cultured candida glabreta and kruzei. Then the diameter of growth inhibition
zone was measured. The collected data was reported by descriptive statistics. Collected
data was analyzed using SPSS 16. P<0.05 was considered statistically significant.This
study was done in Tabriz medical university grant number 2013472975 between 2012
till 2013. Results: The mean growth inhibition zone diameter against candida glabrata
was the greatest in ginger extract (27.5 ± 0.6) and the least in nystatin (17 ± 0.5). The
mean growth inhibition zone diameter against candida kruzei was the greatest in ginger
extract (31.06 ± 0.7) and the least in nystatin (17.5 ± 0.5). Kruskal-Wallis analysis
revealed that there was a significant difference among the growth inhibition zone
diameter against both fungal species (P<0.05). Mann-Whitey U test indicated that the
growth inhibition zone diameter of ginger extract was more that nystatin and
flouconazole in respect of two candidal species. (P=0.000). Conclusions: Antifungal
effect of ginger extract against candida glabrata and candida kruzei was more than
nystatin and flouconazole. These fungi were resistant to nystatin and fluconazole but
sensitive to ginger extract.
© 2014 AENSI Publisher All rights reserved.
To Cite This Article: Ali Taghavi zenouz, Masoumeh Mehdipour, Solmaz Pourzare, Mohammad Adibpour, Tahmoores Abdollahian, In
Vitro comparison of the Effects of Ginger Extract, Fluconazole and Nystatin on Candida glabrata and Candida kruzei. Adv. Environ. Biol.,
8(13), 506-511, 2014
INTRODUCTION
Candidiasis is one of the most prevalent and important opportunistic fungal infections in human beings
which caused by candida yeast species like albicans, glabrata, and kruzei. Different forms of candidiasis, in
acute and chronic forms, affect various parts of body such as skin, oral and genital mucosa, bronchus, GI tract
and lungs. These infections diffuse usually in immunosuppression status like HIV and other contributing factors
and strike internal organs such as kidney and liver [1]. Antifungal drugs, in various formulations, are being used
topically (like nystatine and clotrimazole) and systemically (azoles and amphotericin B) [2]. In recent years,
numerous studies have been reported the failure of treatment in patients with different clinical types of
candidiasis. Long-term consumption of antifungal agents has been caused adverse effects and also drugresistance against these against leaded to the limitations in antifungal therapies [3, 4]. Drug resistance is
reported more in individuals with compromised immune system and long-term use of antifungal agents [5]. The
number of infections caused by candidal species other than albicans (like kruzei and glabrata) is increasing and
these microorganisms respond poorly to azole treatments such as fluconazole which is the most effective azole
in management of fungal infections in immunosuppress and HIV positive individuals). This drug resistance
leads to the increase of infection prevalence [6]. Side effects of the common antifungal agents include nausea,
vomiting, hepatic dysfunction, arrhythmias, neuropathies, and etc. Therefore, recent researches are directed in
finding effective antifungal agents with natural origin and less side effects [5]. Among herbal extracts, inhibitory
Corresponding Author: Solmaz Pourzare, Department of Oral Medicine, Faculty of Dentistry, Tabriz University of
Medical Sciences, Tabriz, Iran,
Tel: 00989148125118, E-mail: [email protected]
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Solmaz Pourzare, 2014
Advances in Environmental Biology, 8(13) August 2014, Pages: 506-511
effect of ginger extract on microorganisms has been evaluated by researchers. Antimicrobial effect of ginger
extract has been investigated against staphylococcus aureus, pseudomonas aeruginosa, and escherichia coli and
reveled that this extract has certain inhibitory effect on these species [7]. In another study antifungal activity of
ginger extract was assessed against fluconazole-resistant candida albicans species isolated from patients with
genital candidiasis. It revealed that ginger extract has a inhibitory impact on all candida albicans isolates [8].
In traditional medicine ginger is administrated for cure of movement inabilities, nausea and vomiting during
pregnancy, and etc. numerous studies did not report any side effects for ginger except for sedation and
drowsiness [6-8]. Antimicrobial and antifungal effects of ginger extract has been proved against some of
bacterial and fungal species as candida albicans in vitro, and since no study has been performed on other
candidal species like glabrata and kruzei which are increasing with raising immunosuppress and HIV affected
patients, therefore this study was aimed to evaluate the antifungal property of ginger extract on candida glabrata
and kruzei and compare its effect with common antifungal agents.
MATERIALS AND METHODS
In this in-vitro experimental study, disc diffusion method was employed for comparison of the effect of
ginger extract with two commonly used antifungal agent, nystatin and fluconazole. Standard species of candida
glabrata (BSM 11226) and candida kruzei (BSM 70079) were tested in this study. 0.5 McFarland suspension
was prepared by 24-hour cultured microorganisms in sterile physiologic serum. For each fungal species,
antibiogram test was carried out on 15 sabouraud dextrose agar. Plates were inoculated using sterile swab
soaking in prepared fungal suspension and dispersed on gar medium. After that quaternary discs containing
ginger extract, nystatin, fluconazole, and a blank one were placed on each plate with a same distance from each
other. After completion of these processes, all plates were incubated at 35°C for 18-24 hours and then diameter
of growth inhibition zone around discs were measured using a millimeter ruler. There should not be any growth
inhibition around blank disc to confirm test validity.
For preparation of ginger extract, 500 grams of arid herb was grinded and wetted in 100 milliliters of 99%
ethanol for 24 hours. Prepared solution was distillated and eventually 45 grams of arid extract were obtained. In
a 96-well cell culture microplate, 100 microliters of sabouraud dextrose medium were added to each well,
subsequently, 100 microliters of ginger extract were added to first well and later dilutions were prepared in next
wells. Then, 100 microliters of fungal specimen were added to each well and after 18-24 hours incubation at
35°C, wells were evaluated in respect of turbidity. The well before the first turbid well was considered as MIC.
This was performed for both of fungal species. Obtained MIC for both fungal species was 25 µg/ml.
Antibiogram discs of fluconazole and nystatin were prepared from Padtan Teb, Tehran, Iran. Fluconazole discs
each contained 25 µg fluconazole in a 6.4 mm disc and nystatin discs contained 100 units or 20 µg pure
nystatin. Ginger extract discs were prepared by solving 25 mg of extract in 1ml of total ethylic alcohol (99.6%).
Sterile disks were placed on a sterile glass and 10 µl of prepared 25 mg/ml of extract were poured on them to
absorb completely. Subsequently, discs were transferred into 37 °C incubator for 30-60 minutes in order to
evaporation of alcohol and obtaining arid discs.
Collected data first was reported by descriptive statistics. Kolmogorov-Smirnov analysis was used for
evaluation of data normality. Comparison of mean growth inhibition zone of three experimental groups was
done by Kruskal-Wallis test in both fungal species. U-Mann Whitney analysis was used for pair-wise
comparisons. P<0.05 was considered significant in this study. This study was done in Tabriz medical university
grant number 2013472975 between 2012 till 2013.
RESULTS AND DISCUSSION
Results:
The greatest growth inhibition diameter in candida kruzei and glabrata was recorded around ginger extract
discs (table 1).
Table 1: descriptive data regarding growth inhibition diameter around tested antifungal agents
Candida
Experimental
Mean (std. deviation)
antifungal agent
Glabrata
Ginger extract
31.06 (0.7)
Nystatin
17 (0.5)
Fluconazole
18.3 (0.5)
Kruzei
Ginger extract
27.5 (0.6)
Nystatin
17.5 (0.5)
Fluconazole
19.5 (0.5)
minimum
Maximum
30
16
18
26
17
19
32
18
19
28
18
20
Kolmogorov-Smirnov analysis revealed nan-parametric data distribution (P>0.05). Kruskal-Wallis test
indicated that mean of growth inhibition zone of tested materials are statistically significant in case of both
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fungal species (P=0.000). U-Mann-Whitey analysis depicted that all pair-wise comparisons are statistically
significant (P=0.000). In other words, mean growth inhibition zone diameter of ginger extract was more than
nystatin and fluconazole (Fig. 1 and 2).
Fig. 1: Error bar of growth inhibition zone diameter of experimental antifungal agents against Candida glabrata
Fig. 2: Error bar of growth inhibition zone diameter of experimental antifungal agents against Candida Kruzei
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Fig. 3: Growth inhibition zone against Candida glabrata
N: nystatin, Z: ginger extract, F: fluconazole
Fig. 4: Growth inhibition zone against Candida kruzei
N: nystatin, Z: ginger extract, F: fluconazole
Discussion:
In this study antifungal effects of ginger extract were investigated against candida kruzei and glabrata. For
comparison of antifungal effect of ginger extract, effects of two commonly used antifungal agents (nystatin and
fluconazole) were assessed, too. For this, disc diffusion agar method was used since it a simple and reliable
method and it was employed in similar studies [7, 8]. In our study antifungal effect of ginger extract was greater
than nystatin and fluconazole in respect of both candida species.
In previous studies antifungal effects of protein in ginger rhizome was evaluated and revealed that this
protein had inhibitory effect on some of fungi such as fusarium oxysporum [9]. Taechowisan et al isolated a
material called CMUAC 130 from ginger which had inhibitory effect on fytopathogen fungi growth as fusarium
[10]. Nguefack et al indicated that ginger extract could prevent the proliferation fusarium moniliform,
aspergillus flavus, and aspergillus fumigatus and can inhibit these fungi growth in-vitro [11]. Ficker et al
evaluated antifungal properties of 36 herbal extracts on 13 human fungal pathogens and reported that among
these extracts, ginger and jimpijapa extract had inhibitory effect on various fungal species. Also it was revealed
that ginger extract is one of the extracts which precluded the growth of fungi which were resistant to
amphotericin B and ketoconazole [12]. Agarwal et al depicted this extract’s inhibitory effect on Spilosoma
insect species [13]. Other researches have evaluated the antifungal effects of its rhizome and assigned it as an
effective extract on aspergillus and fyopathogens [14]. Mohammadi et al assessed its antifungal properties
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against clinical isolates of fluconazole-resistant candida albicans. Their findings indicated that ginger extract
had inhibitory effect on all tested species and they declared ginger as an effective agent on candida albicans in
laboratory setting [8]. There have not been performed any studies on antifungal effect of finger extract on nonalbicans candida species. Therefore, our study aimed this matter. Similar to previous studies, our findings also
depicted that its antifungal properties is stronger than nystatin and fluconazole.
Although, antimicrobial effects of common antimicrobial agents on different microorganisms are compared
to the standards deducted by CLSI (Clinical and Laboratory Standard Institute), there is not any reference
standards for other materials such as herbal extracts. Former studies just have reported antifungal properties of
these extracts descriptively and have not compared with any references. In this study, we compared the
antifungal effects on ginger extract with two commonly used antifungal agents (nystatin and fluconazole), for
the first time and indicated that antifungal effect of this extract is much more than nystatin and fluconazole
against candida kruzei and glabrata species. According to CLSI standards in 2011, growth inhibition zone
diameter of nystatin against candida kruzei and glabrata is 25 mm and this is 22 mm for fluconazole [15, 16].
The mean inhibition diameter obtained for nystatine against candida kruzei and glabrata was 17 mm and this
was 19 mm for fluconazole in our study which illustrates these antifungal agents’ resistance to nystatin and
fluconazole. But these diameters were 31 and 27.5mm for ginger extract against candida kruzei and glabrata,
respectively. These diameters shows that these microorganisms were sensitive to ginger extract in MIC
concentration.
Ginger is administrated for cure of movement inabilities, nausea and vomiting during pregnancy, and etc.
numerous studies did not report any side effects for ginger except for sedation and drowsiness [6-8].
Antimicrobial and antifungal effects of ginger extract has been proved against some of bacterial and fungal
species as candida albicans in vitro, and in our study its antifungal effect were proved on candida glabrata and
kruzei which are increasing with raising immunosuppress and HIV affected patients, therefore because of
ginger’s inexpensiveness and less side effects, it’s administration is suggested for fungal infections. However,
further animal and human studies are needed to confirm this.
Conclusions:
Antifungal inhibitory effect of ginger extract against candida kruzei and glabrata was greater than nystatin
and fluconazole. These fungi were resistant to nystatin and fluconazole but sensitive to ginger extract.
Suggestions: Regarding greater antifungal effect of ginger extract in comparison with two commonly used
antifungal agents, and also because of ginger’s inexpensiveness and less side effects, its administration is
suggested for fungal infections. However, further animal and human studies are needed to confirm this.
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