Antibacterial Effects of Iranian Fennel Essential Oil on Isolates of Acinetobacter baumannii
The aim of the present study was the evaluation of the antibacterial activity of Fennel essential oil on isolates of Acinetobacter baumannii. Forty eight isolates were collected from clinical specimens from burn wards of hospitals in Tehran, Iran between April and September, 2006. The susceptibility of isolates was determined using a broth microdilution method. Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) of isolates to Fennel essential oil were determined. The susceptibilities of isolates to different antibiotics were tested using agar disk diffusion method. The rates of resistance were determined to antibiotics as follows: cefazolin 100%, ciprofloxacin 100%, ofloxacin 95.8%, kanamycin 95.8%, carbenicillin 93.7%, ticarcillin 93.7%, piperacillin 88.9%, co-trimoxazole 79.1%, ceftizoxime 75%, gentamicin 70.8%, cefalotin 60.4%, amikacin 52% and imipenem 14.6%. Fennel essential oil possessed antibacterial effect against all isolates of A. baumannii. These results suggest the potential use of the Fennel essential oil for the control of multi-drug resistant A. baumannii infections. However, more adequate studies must be carried out to verify the possibility of using it for fighting bacterial infections in human.
Medicinal plants contain biologically activity compounds, many of which have
been shown to have antibacterial properties (Mohsenzadeh,
2007). Foeniculum vulgare (Fennel) belonging to the family Apiaceae
is a perennial herb native to the Mediterranean Region. It is widely cultivated
and extensively used as a culinary spice. The plant is aromatic and is used
as a pot herb. The leaves have diuretic properties and the roots are regarded
as purgatives. Dried fruits of Fennel possess a pleasant aromatic taste and
used for flavouring soups, meat dishes and sauces. The fruits are considered
to be useful in treatment of diseases of the chest, spleen and kidney (Singh
and Kale, 2008).
The antiinflammatory, analgesic and antioxidant activities of the fruit of
Fennel have been reported previously by Choi and Hwang (2004).
Oral administration exhibited inhibitory effect against acute and subacute inflammatory
diseases and type IV allergic reactions and showed a central analgesic effect.
It significantly decreased the high density lipoprotein-cholesterol level along
with a decrease in the peroxidative damage (Choi and Hwang,
2004). The antibacterial activity of essential oil of Fennel has been reported
previously by Ruberto et al. (2000).
Anethole is the principal active component of Fennel seeds which has exhibited
anticancer activity (Aggarwal et al., 2008).
Also, chemopreventive potential of Fennel against carcinogenesis has been shown
earlier by Singh and Kale (2008).
Acinetobacter baumannii is a Gram-negative, nonmotile, nonfermentative
and oxidase-negative bacillus, whose natural reservoir has not been clearly
determined. It is found in many hospital environments and can be colonize in
human body in the hospital environments. The combination of its environmental
colonization and its very high resistance to antimicrobial renders it as a successful
nosocomial pathogen (Nordmann, 2004). There are many
reports of Multi Drug Resistant (MDR) A. baumannii from hospitals in
Europe, North America, Argentina, Brazil, China, Taiwan, Hong Kong, Japan and
Korea and many other areas (Barbolla et al., 2003;
Houang et al., 2001; Lee
et al., 2004; Liu et al., 2006; Naas
et al., 2006; Nishio et al., 2004;
Quale et al., 2003; Van
Looveren and Goossens, 2004; Yu et al., 2004).
These MDR strains often spread to cause outbreaks throughout hospital wards.
Acinetobacter sp. is usually considered to be opportunistic
pathogens. They cause a wide range of clinical complications, such
as pneumonia, septicemia, urinary tract infection, wound infection
and meningitis, especially in immunocompromised patients. MDR A.
baumannii infections tend to occur in immunosuppressed patients, in patients
admitted in intensive care and burn units and in those subjected
to invasive procedures and treated with antibiotics. In respect of its very
high resistance to antimicrobials, introducing of the new antimicrobial agents
against this bacterium is one of the most important goals in treatment of such
infections (Perez et al., 2007).
In this study we evaluated the antibacterial activity of Fennel on 48 hospital isolates of MDR A. baumannii.
MATERIALS AND METHODS
Essential oil: Fennel essential oil from Barije Essence Pharmaceutical Company, Iran (commercial producer of plant essential oils and aromatic substances) were used in this study. The oil was selected based on literature survey and its use in traditional medicine. Quality of the oil ascertained to be more than 80% pure. The main effective components of the Fennel essential oil were anethole (50-60%) and fenchone (10-20%).
Bacterial strains and culture media: A total of 48 isolates were collected
from clinical specimens from burn wards of hospitals in Tehran, Iran during
a 6 months period between April and September, 2006. The isolates were further
processed by the standard methods to identify as the A. baumannii (Baron
and Finegold, 1990). Isolated bacteria were maintained for long storage
on skimmed milk medium (BBL) by adding 10% glycerol in -60°C, cultures were
maintained for daily use on nutrient agar (BBL) slants at 4°C. The Muller
Hinton Agar (MHA) and Muller Hinton Broth (MHB) medium (Pronadisa) were used
for detection of antibiotic resistance of strains. Acinetobacter calcuaceticus
PTCC 1318 has been used as reference strain.
Determination of antimicrobial activity of Fennel essential oil: The
susceptibility of Acinetobacter isolates to Fennel essential oil was
determined using a broth microdilution method based on CLSI guidelines. Minimum
Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC)
of Fennel essential oil for isolates were determined. Muller-Hinton Broth (MHB;
Oxoid) was supplemented with 0.002% (v/v) Tween 80 (Sigma) (MHB-T) to enhance
dispersion of the Fennel oil (Papadopoulos et al.,
2006). The initial concentration of Fennel essential oil in the first tube
contains MHB-T was ½.
This was used to prepare serial doubling dilutions over the range 0.03-25%
(v/v). 1.5x106 inoculums of the isolates were added to each concentration
in MHB-T. A tube containing growth medium without essential oil and an un-inoculated
tube were used as a positive and negative growth control, respectively. Antibacterial
activity was measured by determining MICs and MBCs. The MIC was the lowest concentration
of essential oil that resulted in a clear tube. Ten microlitres from each tube
was spot-inoculated onto Nutrient Agar (NA) and incubated overnight at 37°C
to determine the MBC. The highest dilution that inhibits bacterial growth on
nutrient agar after overnight incubation was taken as MBC (Baron
and Finegold, 1990; Papadopoulos et al., 2006).
Experiments were repeated for three times and the modal value calculated.
Determination of the strains sensitivity to antibiotics: The susceptibilities of isolates to different antibiotics were tested using agar disk diffusion method. To represents the different classes of antimicrobial agents commonly used for the treatment of Acinetobacter sp. infections, we used piperacillin, ciprofloxacin, gentamicin, ofloxacin, cephalotin, ticarcillin, kanamycin, imipenem, amikacin, co-trimoxazole, ceftizoxime, cefazolin and carbenicillin (Hi-media, Mumbai, India).
The rates of resistance to different antibiotics for 48 isolates of Acinetobacter
baumannii have been shown in Table 1. Cefazolin (100%),
ciprofloxacin (100%) ofloxacin (95.8%) and kanamycin (95.8%) showed the highest
rate of resistance and amikacin (52%) and imipenem (14.6%) demonstrated the
lowest (Table 1). 45.8% of isolates showed resistance to the
11 tested antimicrobials. Results showed that Fennel essential oil possessed
antibacterial effect against all isolates of Acinetobacter sp. (Table
2), 39 isolate were sensitive to all tested dilutions and the remaining
nine isolates showed MIC and MBC values in the range of 3.9x10-3
to 1.56x10-2 mm3 mm-3 (Table
||The rates of resistance to different antibiotics for 48 burn
wound isolates of Acinetobacter baumannii
||Antibacterial activity of Fennel essential oil against 48
burn isolates of Acinetobacter baumannii
Also Acinetobacter baumannii PTCC1318 was sensitive to all tested dilutions.
There was complete growth on the positive growth control tube (containing growth medium without essential oil) and no growth on negative growth control (un-inoculated tube).
Acinetobacter baumannii play an important role in colonization and infection
of patients admitted to hospitals. They have been implicated in a variety of
hospital acquired infections, including bacteremia, urinary tract infection,
meningitis and pneumonia (Perez et al., 2007).
Antibiotic resistance is a major problem in treating infection with
A. baumannii, which has become resistant to almost all available antibacterial
drugs (Longo et al., 2007). The use of broad
spectrum antibiotics in hospital environments for treating such infections results
in promoting infections by multi-antibiotic resistant isolates. Present finding
showed that the most useful antibiotics for infections caused by A. baumannii
were imipenem, amikacin and cefalotin. Resistance to some antibiotics such as
gentamicin, ciprofloxacin and co-trimoxazole showed very high increases in comparison
with earlier studies by Guardabassi et al. (1998).
Also, in present study burn wound isolates of Acinetobacter baumannii
showed high resistance to other tested antibiotics (Table 1),
so it seems reasonable to explore new sources of natural compounds with antibacterial
activity against Acinetonacter sp.
The antimicrobial properties of essential oils obtained from Foeniculum
vulgare have been investigated in different trails. The oil has been shown
to be effective against Streptococcus haemolyticus, Bacillus subtilis,
Pseudomonas aeruginosa, Escherichia coli, Klebsiella
species, Staphylococcus epidermidis (Mohsenzadeh,
2007), Bacillus cereus and Bacillus brevis (Ozcan
et al., 2006). Another study has shown an activity against the food-borne
pathogens like Escherichia coli O157:H7, Listeria monocytogenes,
Salmonella typhimurium and Staphylococcus aureus but in Sagdics
study Fennel extract was inactive against some pathogenic bacteria (Sagdic
and Yasar, 2005).
In present study results showed that Fennel essential oil possessed antibacterial effect against all isolates of Acinetobacter baumannii, furthermore, beside the confirmation of the popular use, the obtained results demonstrate that this herbal drug could represent a new source of antimicrobial agents, for the control of Acinetobacter infections. However, more adequate studies must be carried out to verify the possibility of using it for fighting these bacteria in human body infections.
This study has been supported by a research grant from student research committee of Urmia University of Medical Sciences.
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