Mycobacterium tuberculosis is the etiological agent of tuberculosis. The World Health Organization has estimated that 8 million of people develop active TB every year and the situation is complicated by an increase of Mycobacterium tuberculosis strains resistant to drugs used in antitubercular therapy: MDR and XDR-TB. Myrtle leaf extracts, used as an antiseptic in Sardinian traditional medicine, have strong antibacterial activity as several investigations showed. In this study we investigated the antimicrobial properties of the essential oil of Myrtus communis against clinical strains of M. tuberculosis and M. paratuberculosis

Bua, Alessandra

Cannas, Sara

Cubeddu, Marina

Marongiu, Bruno

Molicotti, Paola

Porcedda, Silvia

Sechi, Leonardo Antonio

Zanetti, Stefania Anna Lucia

UnissResearch

Hindawi Publishing Corporation
Interdisciplinary Perspectives on Infectious Diseases
Volume 2010, Article ID 931530, 3 pages
doi:10.1155/2010/931530
Research Article
Evaluation of the Antimicrobial Properties of
the Essential Oil of Myrtus communis L. against
Clinical Strains of Mycobacterium spp.
Stefania Zanetti,1 Sara Cannas,1 Paola Molicotti,1 Alessandra Bua,1 Marina Cubeddu,1
Silvia Porcedda,2 Bruno Marongiu,2 and Leonardo Antonio Sechi1
1Department of Biomedical Science, Microbiology, University of Sassari, 07100 Sassari, Italy
2Department of Chemical Science, University of Cagliari, 09042 Monserrato, Italy
Correspondence should be addressed to Stefania Zanetti, zanettis@uniss.it
Received 9 June 2009; Revised 23 November 2009; Accepted 15 April 2010
Academic Editor: Jonathan Mayer
Copyright © 2010 Stefania Zanetti et al. This is an open access article distributed under the Creative Commons Attribution
License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly
cited.
Mycobacterium tuberculosis is the etiological agent of tuberculosis. The World Health Organization has estimated that 8 million of
people develop active TB every year and the situation is complicated by an increase of Mycobacterium tuberculosis strains resistant
to drugs used in antitubercular therapy: MDR and XDR-TB. Myrtle leaf extracts, used as an antiseptic in Sardinian traditional
medicine, have strong antibacterial activity as several investigations showed. In this study we investigated the antimicrobial
properties of the essential oil of Myrtus communis against clinical strains of M. tuberculosis and M. paratuberculosis.
1. Introduction
Myrtle (Myrtus communis L.) is an evergreen shrub belong-
ing to the family of Mirtaceae that grows spontaneously
throughout the Mediterranean area. In Italy it grows along
the coast and in the inner hills, and it is spread especially
in the islands, where it is one of the most characteristic
species. Myrtus communis had history in the popular and
traditional medicine: the essential oil obtained from leaves
and, sometimes, flowers and berries has been used for its
tonic and balsamic properties, and it is used in flavour and
fragrance industries. In Sardinia natural formations are still
the main source for the production of a traditional liqueur,
that every year reaches 3 million bottles. Essential oils are
gaining remarkable interest for their potential multipurpose
use as antioxidant, antibacterial, and antiseptic agent [1–4];
the essential oil obtained from the leaves was used in the past
for the treatment of lung disorders.
The isolation of essential oils from Myrtus communis
leaves is usually obtained by hydrodistillation method with
a Clevenger-type apparatus, according to the Italian Oﬃcial
Pharmacopoeia. The chemical composition of the essen-
tial oils, analysed by Gas/Cromatography (G/C), generally
exhibits α-pinene, 11%; 1,8-cineole, 16%; linalool, 12%; α-
terpineol, 7%; and limonene, 5%. The Sardinian myrtle oil is
characterized by the lack of myrtenyl acetate and by a higher
content of limonene.
Romani et al. [5] showed the biological activities of tan-
nins, including anticancer and antioxidant. In our previous
studies (data not published) the antimicrobial properties of
myrtus essential oil against several clinical strains and in
particular against Helicobacter pylori [6] were studied, and
we obtained encouraging results. Considering these results,
in this study, we have used this essential oil towards strains
of M. tuberculosis and strains of Mycobacterium avium subsp.
paratuberculosis.
2. Materials and Methods
In this study we investigated the antimicrobial properties of
the essential oil of Myrtus communis against two reference
strains: M. tuberculosis H37Rv (virulent strain) and M.
tuberculosis H37Ra (avirulent strain) and 8 clinical isolates
of M. tuberculosis resistant to one or more drugs, collected in
the Department of Biomedical Sciences, Microbiology of the
University of Sassari, Italy.
2 Interdisciplinary Perspectives on Infectious Diseases
Table 1: Activity of antitubercular drugs and Myrtus communis against Mycobacterium tuberculosis strains.
Strains MIC Antitubercular Drug∗
M. tuberculosis Myrtus communis % (v/v) SM INH RIF ETH PZA CLA CIP
H37Rv 0.17% S S S S — — —
H37Ra 0.17% S S S S — — —
M. tuberculosis 0.17% S S R S — — —
M. tuberculosis 0.17% S S R S — — —
M. tuberculosis 0.17% S R R S — — —
M. tuberculosis 0.17% R S R S — — —
M. tuberculosis 0.17% R R S S — — —
M. tuberculosis 0.17% R R S R — — —
M. tuberculosis 0.17% R R R S R R R
M. tuberculosis 0.17% S S S S — — —
M. paratuberculosis (1515) >2% — — — — — — —
M. paratuberculosis (1517) >2% — — — — — — —
∗
SM: streptomicin; INH: isoniazid; RIF: rifampicin; ETH: ethambutol; PZA: pyrazinamide; CLA: clarithromycin; CIP: ciprofloxacin.
We also studied the antimicrobial properties of the
essential oil towards two human strains of Mycobacterium
avium subsp. paratuberculosis (1515 and 1517).
The antibacterial activity of the oil was assessed by the
proportional method used for Mycobacterium tuberculosis
as described in NCCLS-National Committee for Clinical
Laboratory Standards [7].
Briefly, isolate suspensions of M. tuberculosis in 7H9
broth were adjusted to an optical density of 1McFarland, and
two dilutions, 10−2 and 10−4, were plated onto 7H10 agar
with a diﬀerent concentration of essential oil (16%, 14%,
12%, 10%, 8%, 4%, 2%, 1%, and 0.17% v/v).
We also investigated the antimicrobial properties of some
components of M. communis and, in particular, limonene, 1-
8 cineole and α-pinene, using the proportional method [7].
3. Results and Discussion
In Table 1 we report the data obtained with the essential oil
in toto.
Towards all the strains of M. tuberculosis, including the
extensively drug-resistant (XDR), the oil of M. communis
tested showed an MIC of 0.17% (v/v), whereas against the
two M. paratuberculosis strains showed an MIC of 2% (v/v).
Subsequently we tested each diﬀerent compound
(limonene, 1-8 cineole, α-pinene) to see if it had diﬀerent
antimicrobial properties.
As far as limonene concerned, it showed towards all
the strains an MIC of 2% (v/v); about 1-8 cineole, for 4
strains, including H37Rv, the MIC was of 2% (v/v), while
the MIC shown for other 4 strains was of 16% (v/v); α-
pinene showed an MIC of 1% (v/v) for 3 strains, and for
one strain an MIC of 2% (v/v), for 3 others an MIC of 8%
(v/v), and for only one MIC was of 16% (v/v) (Table 2).
Limonene and α-pinene are monoterpenic that, according
to the literature, are used as expectorant, antalgic, revulsive,
antitussive, mucolitic, and decongestant; about 1-8 cineole it
is an oxide used as expectorant, mucolitic, and decongestant
[8].
We compared the activity of our essential oil with
four standard antitubercular drugs: streptomycin, isoniazid,
rifampin, and ethambutol, performing according to inter-
national protocols [7]. About the mycobacterial strains, one
clinical strain was susceptible to all the drugs. Two clinical
strains were rifampicin resistant, three strains were resistant
to 2 drugs (one streptomicin and rifampicin resistant,
one streptomicin and isoniazid resistant, and one isoniazid
and rifampicin resistant), one was resistant to 3 drugs
(streptomicin, isoniazid, and ethambutol). The last one was
a XDR strains. The essential oils screened in toto have a
better antimicrobial activity than each single compound
against all mycobacteria tested. The results presented here
may contribute to the knowledge of the antimicrobial
properties of myrtle and our aim is to carry on further
studies.
4. Conclusions
The chemical composition of the essential oil of M. com-
munis exhibited qualitative diﬀerences that depended on
diﬀerent geographical areas and from the season in which
the leaves were picked up. The essential oil that we used for
antimicrobical in vitro assay contained a high quantity of
monoterpenic and oxide that, according to literature, do not
have antimicrobical activity. From our studies emerged that
the essential oil in toto might have a good activity towards
M. tuberculosis, although the individual compounds (except
α-pinene) showed in all the strains a higher MIC. It may be
the consequence of a sinergic eﬀect of all the compounds
together.
The results from the myrtle oil in toto showed a
good activity towards M. tuberculosis but not toward M.
paratuberculosis. The MIC registered against M. tuberculosis
was 0.17% (v/v) in comparison with an MIC of 2% (v/v)
observed toward M. paratuberculosis.
A limit of our study is the small amount of the essential
oil that did not allow to perform the MIC towards all the
strains used. These data are encouraging even if further
Interdisciplinary Perspectives on Infectious Diseases 3
Table 2: Activity of Myrtus communis and single compound against Mycobacterium tuberculosis strains.
Strains MIC MIC % (v/v)
M. tuberculosis Myrtus communis % (v/v)
Extracts of Myrtus communis oil
Limonene 1-8 Cineole α Pinene
H37Rv 0.17% 0.17% 2 1
H37Ra 0.17% — — —
M. tuberculosis 0.17% 2 2 1
M. tuberculosis 0.17% 2 >16 8
M. tuberculosis 0.17% 2 2 1
M. tuberculosis 0.17% 2 2 16
M. tuberculosis 0.17% 2 >16 8
M. tuberculosis 0.17% 2 >16 2
M. tuberculosis 0.17% 2 >16 8
M. tuberculosis 0.17% — — —
M. paratuberculosis (1515) >2% — — —
M. paratuberculosis (1517) >2% — — —
additional “in vitro” testing and large clinical studies are
necessary to verify the potential use of the essential oils of
myrtle as antitubercular drug. Given the excellent results that
we obtained in this study, we would expand the research with
further studies, to value the possible cytotoxic eﬀects, and
eventually to perform tests using “in vivo” mouse model.
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Evaluation of the antimicrobial properties of the essential oil of <i>Myrtus communis L.</i> against clinical strains of <i>Mycobacterium spp.</i>

Evaluation of the antimicrobial properties of the essential oil of <i>Myrtus communis L.</i> against clinical strains of <i>Mycobacterium spp.</i>

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