Microbial Pathogenesis 80 (2015) 39e49

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Microbial Pathogenesis
journal homepage: www.elsevier.com/locate/micpath

Review

Drug resistance of bacterial dental biolm and the potential use of

natural compounds as alternative for prevention and treatment
Bochra Kouidhi a, *, Yasir Mohammed A. Al Qurashi a, Kamel Chaieb b
a
b

College of Applied Medical Sciences, Medical Laboratory Department, Yanbu, Taibah University, Saudi Arabia
College of Sciences, Biology Department, Yanbu, Taibah University, Saudi Arabia

a r t i c l e i n f o

a b s t r a c t

Article history:
Received 16 October 2014
Received in revised form
16 February 2015
Accepted 19 February 2015
Available online 21 February 2015

Oral diseases, such as dental caries and periodontal disease are directly linked with the ability of bacteria
to form biolm. The development of dental caries involves acidogenic and aciduric Gram-positive bacteria colonizing the supragingival biolm (Streptococcus, Lactobacillus and Actinomycetes). Periodontal
diseases have been linked to anaerobic Gram-negative bacteria forming a subgingival plaque (Porphyromonas gingivalis, Actinobacillus, Prevotella and Fusobacterium). Cells embedded in biolm are up to 1000fold more resistant to antibiotics compared to their planctonic ones. Several mechanisms have been
proposed to explain biolms drug resistance. Given the increased bacterial resistance to antibiotics
currently used in dentistry, a great importance is given to natural compounds for the prevention of oral
bacterial growth, adhesion and colonization. Over the past decade, interest in drugs derived from medicinal plants has markedly increased. It has been well documented that medicinal plants and natural
compounds confer considerable antibacterial activity against various microorganisms including cariogenic and periodontal pathogens.
This paper provides a review of the literature focusing on the studies on (i) biolm in the oral cavity,
(ii) drug resistance of bacterial biolm and (iii) the potential use of plant extracts, essential oils and
natural compounds as biolm preventive agents in dentistry, involving their origin and their mechanism
of biolm inhibition.
2015 Elsevier Ltd. All rights reserved.

Keywords:
Dental biolm
Oral bacteria
Drug resistance
Medicinal plants

1. Introduction
Oral diseases, such as dental caries, periodontal disease are
directly linked with the ability of bacteria to form biolm [1]. The
development of dental caries involves acidogenic and aciduric
Gram-positive bacteria (Streptococci mutans, Lactobacillus and Actinomycetes). On the other hand, periodontal diseases have been
associated to anaerobic Gram negative bacteria (Porphyromonas
gingivalis, Actinobacillus, Prevotella and Fusobacterium) [2]. Biolm
in the form of supragingival and subgingival plaque is the etiologic
agent in dental caries and periodontal diseases. The emergence and
progression of such pathologies are linked with the increased
resistance to antibiotics by bacterial biolms [3]. Therefore, control
of the dental biolm is a major objective of dental professionals for
the maintenance of oral health. With the increased resistance to
antibiotics and conventional treatment, the discovery of anti-

* Corresponding author.
E-mail address: bochrak@yahoo.fr (B. Kouidhi).
http://dx.doi.org/10.1016/j.micpath.2015.02.007
0882-4010/ 2015 Elsevier Ltd. All rights reserved.

infective agents, active against micro-organisms embedded in

biolms is an important goal in the treatment of oral infection [4].
Several approaches have been used for the control of oral biolm such as preventing biolm formation and disrupting existing
biolms [5]. Moreover, the control of dental biolm requires not
only bacteriostatic but also anti-adhesive actions in order to inhibit
the initial bacterial adherence [5].
Medicinal plants have proven their valuable biological activities
[6,7]. Plant-derived compounds and essential oils could inuence
biolm formation by inhibiting peptidoglycan synthesis [8],
damaging microbial membrane structures [9], and modulating
quorum sensing [8]. In dental care, the use of natural products to
prevent dental caries is receiving large attention [10] and many
studies have reported the effects of medicinal plant extracts on
inhibition of oral pathogen growth, inhibition of bacterial adhesion
to surfaces and reduction of dental plaque.
This paper provides a review of the literature focusing on the
studies on (i) biolm in the oral cavity, (ii) drug resistance of bacteria embedded in biolm and (iii) the potential use of plants extracts as anti-biolm agents in dentistry, involving their origin and

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B. Kouidhi et al. / Microbial Pathogenesis 80 (2015) 39e49

their effect (inhibition of bacterial growth and adherence,

increasing the action of drugs, inhibition of efux pump, inhibition
of quorum sensing systems and inhibition of Glucosyltransferases
activity).
2. Biolm in the oral cavity
A biolm is dened as a sessile microbial community adherent
to a solid surface and surrounded by the microbial extracellular
polysaccharides and proteins [11]. The human oral cavity is a highly
dynamic environment inhabited by more than 750 species [2]. One
cubic millimeter of dental plaque contains about 100 million bacteria and may serve as a persistent reservoir for potential pathogens. The adhesion of oral bacteria is a crucial step in the
development of dental plaque considered as a complex multispecies biolm community [12] resulting in a structurally organized microbial community [13]. Distinct stages in plaque formation include: acquired pellicle formation on teeth surface;
reversible adhesion involving weak long-range physicoechemical
interactions between the cell surface and the pellicle; co-adhesion
of secondary colonizers to already attached cells; multiplication
and biolm formation [14].
Dental plaque, one of the best-studied biolms, is a very
important ecosystem for bacterial growth which affords oral bacteria protection against anti-microbial agents [15] and immune
system [16].

proteins and pathways have been identied leading to its survival

under starved condition [25].
The two-component signal transduction systems (TCS) are
important regulatory systems for bacterial survival and adaptation
to stress [26,27]. The TCS are composed of a transmembrane sensor
kinase that detects environmental changes and a cytosolic response
regulator, which is a DNA binding protein that modulates expression of target genes when phosphorylated by the kinase [28].
3.3. Decreased growth rates and metabolism
Decreased growth and metabolism rates are strategies used by
some bacteria to survive a wide range of environmental threats
such as temperature shifts and extreme pH. Nutrient limitation and
changes in growth rate are often accompanied by modication in
cell envelope components such as polysaccharides [29], envelope
proteins [30], fatty acids and phospholipids [31]. In biolms, cells
located more deeply are exposed to conditions that differ from
those at the surface including nutrient and oxygen tension. These
results were observed in altered phenotypes in terms of growth
rate and gene transcription in Pseudomonas aeruginosa [31]. In
another way, the slow metabolic rate of microorganisms in biolms
can impede effectiveness of many antimicrobial agents [32]. It has
been reported that Enterococcus feacalis forms intracanal biolms,
peri-apical biolms and biomaterial centered infection and the
inhibition of this biolms requires very high concentrations of
antibiotics such as amplicillin, vancomycin, and linezolid [33].

3. Bacterial biolm and drug resistance

3.4. Multiple drug resistance (MDR) pumps
Cells embedded in biolm are more resistant to conventional
antibiotics compared to their planctonic ones [3]. Several mechanisms have been proposed to explain drug resistance of biolm
including: (i) restricted penetration of antimicrobial agents; (ii)
induction of the general stress response; (iii) decreased growth
rates and metabolism; (iv) increasing expression of multiple drug
resistance (MDR) pumps; (v) activating quorum sensing systems;
and (vi) changing proles of outer membrane proteins (OMP).
3.1. Limitation of antimicrobial penetration into bacterial biolm
Antimicrobial drugs must diffuse through the biolm matrix in
order to inactivate the bacterial cells. Bacterial extracellular polymeric substances (EPS) represent a strong barrier (Fig. 1) for these
molecules by inuencing the rate of its transport to the deep biolm layer [17].
EPS limits the diffusion of antimicrobials into a biolm, which
has been extensively studied in oral biolms [18]. There are several
mechanisms involved in this limited penetration of antimicrobials,
such as molecule size of the drugs [19], antimicrobial hydrophobicity [20], interaction with biolm components [21], electrostatic
interactions [22], absorption [23], and neutralization by enzymes
[24]. However, the resistance provided by EPS may be discouraged
by longer exposure time or higher concentrations of antimicrobials,
both are often impossible for oral applications.
3.2. Stress response and biolm drug resistance
Oral biolms are especially subject to a number of environmental uctuations, such as availability of nutrients, pH, oxygen
and presence of bacteriocins. Bacterial cells are protected from the
effects of changes in pH and many chemical agents (Fig. 1) using a
stress response strategy [25]. Antimicrobial drugs targeting the
microbial cells can create reactive oxygen species (ROS), and other
agents causing the accumulation of acid inside the cell, the increase
of abnormal proteins and DNA damage. In S. mutans, many genes,

Pump efux are integral membrane proteins that utilize metabolic energy to expel drugs across the membrane against the concentration gradient [34]. They are one of the most important
mechanism of drug resistance in Gram-positive as well as in Gramnegative bacteria. These efux pumps can be specic for a given
drug or may recognize a wide range of structurally and chemically
unrelated compounds. The latter efux system, which is known as
multidrug resistance (MDR) transporter, is very important for the
emergence of antibiotic resistance among pathogens [34]. MDR
transporters are also important for bacteria to survive under
complex environment and facilitate biolm formation [34,35].
In oral biolm, bacteria are continuously challenged by changes
in the environmental conditions. As a response to such challenges,
S. mutans which can become dominant in oral biolms may rely on
MDR transporter allowing withstanding toxic compounds produced by competing species or present in the plaque environment.
Among the identied pumps, an ABC transporter, the VltA/B was
involved in the efux of viologens and QACs in S. mutans [36].
Recently, MdeA, a multidrug efux pump using the proton motive
force as the energy for efux drugs, was also identied in S. mutans
[37].
A further work demonstrated that S. mutans possesses a
conserved P-type ATPase which enables the bacterium to tolerate
high concentrations of extracellular copper. The expression of the
ATPase appears to be regulated by copper via the activity of two
other members, copY and copZ genes [38].
3.5. Quorum sensing
In most bacteria, cellecell communication and biolm formation depend greatly on quorum sensing (QS) activity [39,40].
However, the role of QS in biocide resistance is not yet clear.
Furthermore, a possible implication of QS in the regulation of
multidrug efux pumps has been described [41]. The over expression of S-ribosylhomocysteinelyase in antibiotic resistance

B. Kouidhi et al. / Microbial Pathogenesis 80 (2015) 39e49

41

Fig. 1. A schematic presentation of mechanisms involved in drugs resistance of bacterial biolms.

enhances its possible role in multi-drug tolerence [42]. Other

studies in aerobic bacteria have demonstrated that QS (Fig. 1) can
modulate levels of Gram negative resistance nodulation division
(RND) family and enhance biolm formation [41,43]. Recently, it
has been reported that the combined use of an antibiotic and a QS
inhibitors may increase the success of antibiotic treatment by
increasing the susceptibility of bacterial biofilms to the tested drugs
[44].
3.6. Changing proles of outer membrane proteins (OMP)
Another resistance mechanism that can be induced in biolm
cells is the alteration of the membrane-protein composition in
response to antimicrobial agents. This change could result in
decreased permeability of the cell to these compounds. Several
antibiotics use porin channels when entering Gram negative bacteria. So, the decreased expression of porins leads to antibiotic
resistance in Gram-negative bacteria [45]. In Gram-positive bacteria the peptydoglycan polymer is very close to the cell surface,
allowing the antibiotic to easily penetrate the cell. In contrast, the
situation is different with Gram-negative bacteria, since they
contain an outer membrane consisting of lipopolysaccharide that
acts as a barrier to hydrophobic compounds and some antibiotics
[45]. However, some small, hydrophilic antibiotics diffuse through
aqueous channels in the outer membrane formed by proteins called
porins.
4. Natural compounds as modulators of bacterial biolms
Medicinal plants have been used as traditional treatments for
numerous human diseases for thousands of years [7]. Plant extracts, phytochemical compounds and essential oils were investigated for their ability to treat or prevent adhesion of oral bacteria to
various surfaces. Table 1 summarizes the inhibitory effects of natural compounds on biolm forming oral bacteria.

4.1. Salvadora persica

Salvadora persica (Salvadoraceae) or Arak, is a commonly plant
used in the middle east [46]. Extracts of S. persica have been
included as ingredients in oral hygiene products [47].
S. persica has been reported to have many pharmacological
properties on cariogenic and periodontal pathogens such as
S. mutans, Streptococcus salivarius, Staphylococcus aureus [48] and
A. actinomycetemcomitans [49]. It has been shown that S. persica
contains bioactive phytochemical compounds (benzyl (6Z,9Z,12Z)6,9,12-octadecatrienoate, 3-benzyloxy-1-nitro-butan-2-ol and 1,3cyclohexane dicarbohydrazide) which inhibit growth and interact
efciently with the bacterial communication QS regulators [50]. In
addition, miswak extracts showed a signicant activity on reducing
the plaque acidogenicity, suggesting a potential role in caries prevention [51] and may inhibits the acid production of Candida albicans [52].

4.2. Juglans regia

Persian walnut, Juglans regia, (Juglandaceae), native to central
Asia grows as a wild or semi-cultivated tree in a wide area from
southeastern Europe to Turkey and Iran. J. regia stem bark contains
chemical constituents, namely, b-sitosterol, ascorbic acid5, juglone,
folic acid, gallic acid, regiolone, and quercetin-3-a-L-arabinoside
[53]. Antibacterial, and antioxidant activities of this plant have been
described elswere [54,55].
Antibacterial activities have been described for different J. regia
cultivars [56]. Study on J. regia locally available in Pakistan developing a signicant activity against S. mutans and play a vital role in
prevention of oral biolm formation as compared to conventional
antibiotics [57]. Recently, antibiolm properties of Tunisian J. regia
extract has been studied for several Human pathogenic bacteria
[58].

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B. Kouidhi et al. / Microbial Pathogenesis 80 (2015) 39e49

Table 1
Inhibitory effects of plant extracts, essential oil on biolm forming oral bacteria.
Plants

Extracts

Salvadora persica

- Miswak

Juglans regia

Compounds

Methanol
Ethanol
Chloroform
Acetone
Aqueous
Water

Inhibition effect

References

- In vivo growth

[48,134]

- Growth
- Biolm
- Quorum sensing

[50]

Biolm
Adherence
Adhesion
Coaggregation
Growth
Biolm
Virulence factors
Growth
Glucosyltransferases
Acid production
Growth
Adherence
Biolm
Glucosyltransferases
Acidogenicity
Growth
VAdherence
Biolm
Hydrophobicity
Growth
Glucosyltransferases
Biolm
Acidogenicity
Acidurity
Specic virulence factors
Adherence
Growth
Adherence
Glucosyltransferase
Glucan-binding lectin activities
Growth

[57]

Growth
Adherence
Biolm
Growth
Adherence
Biolm

[68]

Glucosyltransferase
Glucan production
Biolm
Biolm
Adherence
Virulence factors
Growth
Adherence
EPS formation
Glucosyltransferase
Growth
Biolm
Water-insoluble glucan
Adherence

[77]

Growth
Adherence
Glucosyltransferase
Glucosyltransferases
Glycolytic pH-drop
Dextransucrase
Growth
Adherence
Insoluble glucan synthesis

[91]

- Porphyromonas gingivalis
- Proanthocyanidins
- Licochalcone A

- Porphyromonas gingivalis

- Flavonols
- Proanthocyanidins

- Streptococcus mutans

- Juice

- Streptococcus mutans

- Juice

- Oral streptococci

- Polyphenols

- Streptococcus mutans

- Ethanol

- Porphyromonas gingivalis
- Streptococcus mutans

- Water

Morus alba

Mutans streptococci
Lactobacilli
Staphylococcus aureus
Streptococcus mutans

- Streptococcus mutans

Vaccinium macrocarpon - Non-dialyzable

material (NDM)

Camellia sinensis

Strains

Semi fermented
Non fermented extracts
Ethanol
- 1-deoxynojirimycin (DNJ). Ethanol
-

Streptococcus mutans
Actinobacillus actinomycetemcomitans
Porphyromonas gingivalis
Prevotella intermedia
Streptococcus mutans
Streptococcus mitis
Streptococcus sanguis
Streptococcus mutans
Actinomycete viscosus
Lactobacillus acidophilus
Lactobacillus lactis
Streptococcus mutans

Trachyspermum ammi

- Petroleum ether
- Ethanol

- Streptococcus mutans

Piper betle

- Water

- Streptococcus mutans

- Hydroxychavicol

- Extracts

Vitis Vinifera

- Phenol

- Streptococcus mutans
- Actinomyces viscosus
-

Candida albicans
Candida tropicalis
Candida krusei
Streptococcus mutans

- Streptococcus mutans
Azadirachta indica

- Water

- Streptococcus mutans
- Oral streptococci

[60]
[135]

[63]

[61]

[136]

[66]

[71]
[69]

[67]

[76]

[80,81]

[82]

[83]

[84]

[92]
[99]
[137]

B. Kouidhi et al. / Microbial Pathogenesis 80 (2015) 39e49

43

Table 1 (continued )
Plants

Extracts

Compounds

Strains

- Streptococcus mutans
- Lactobacillus
- Candida albicans

Sanguinaria canadensis
Myristica fragrans

- Streptococcus mutans
Methanol

Kaempferia pandurata

Macelignan

Panduratin A

Isopanduratin A

Pistacia atlantica

- Essential oil

- Water
- Chloroform
- Ethanol

Pistacia vera

- Water
- Chloroform
- Ethanol

Achyranthes aspera

methanol

Artocarpus
lakoocha

- Water

3,12-oleandione

Polygonum cuspidatum - Methanol

Streptococcus sobrinus
Streptococcus salivarius
Streptococcus sanguis
Lactobacillus acidophilus
Lactobacillus casei
Streptococcus mutans
Streptococcus sanguis
Actinomyces viscosus
Streptococcus mutans
Streptococcus sanguis
Actinomyces viscosus
Porphyromonas gingivalis
Prevotella intermedia
Porphyromonas gingivalis
Propionibacterium acnes
Streptococcus mutans
Streptococcus mutans

Streptococcus mutans
Streptococcus salivarious
Streptococcus sobrinus
Streptococcus sanguis
Actinomyces viscosus
Streptococcus mutans
Streptococcus salivarious
Streptococcus sobrinus
Streptococcus sanguis
Streptococcus mutans

Streptococcus mutans
Streptococcus sobrinus
Aggregatibacter
Porphyromonas
Streptococcus mutans
Streptococcus sobrinus

- Ethyl acetate (fraction 1)

- Methanol
- Ethyl acetate

- Methanol

- Fraction F1: resveratrol,

emodin and physcion

- Streptococcus mutans

Helichrysum litoreum

- Ethanol

Rosmarinus ofcinalis

- Water

- Streptococcus mutans
- Actinomyces viscosus
- Streptococcus sobrinus

Mentha spicata

Essential oil

- Streptooccus mutans

Eugenia caryophyllata

- Methanol

Biorin
Kaempferol
Rhamnocitrin
Myricetin
Gallic acid
Ellagic acid
Oleanoic acid

Streptococcus mutans
Actinomyces viscosus
Porphyromonas gingivalis
Prevotella intermedia

Inhibition effect

References

Aggregation
Biolm
Plaque index
Bacterial count
Growth
Adherence
Hydrophobicity
Biolm
number of CFUs
Dental plaque index
Growth

Growth
Adherence
Biolm
Growth
Adherence
Biolm
Growth
Growth

Growth
Adherence
Biolm
Biolm
Adherence
Salivary glycolysis
Acid-producing ability

Growth
Biolm
Adherence
Acid-producing ability
Biolm
Adherence
Glucosyltransferases
Qorum sensing
Growth

[113]

Growth
Biolm
Adherence
Glucosyltransferases
Glycolytic pH-drop
Growth
Biolm
Growth
Biolm
Glucosyltransferases
Glycolytic pH-drop
Acid production
F-ATPase activity
Water-insoluble polysaccharides
Growth
Adherence
Growth
Adherence
Glucosyltransferases
Growth
Biolm
Growth

[119]

[94,138]
[100]

[102]
[104]

[105]

[108,109]

[139]
[107]

[110]

[111]

[116]

[117]

[121]
[120]

[122].

[123]
[126]

[115]
[127]

(continued on next page)

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B. Kouidhi et al. / Microbial Pathogenesis 80 (2015) 39e49

Table 1 (continued )
Plants

Extracts

Compounds

- Essential oil

Eucalyptus

- Essential oil

Strains
-

Inhibition effect

Streptococci spp.
Candida spp.
Streptococci spp.
Actinomyces viscosus
Porphyromonas gingivalis
Prevotella intermedia
- Growth
Porphyromonas gingivalis
Actinobacillus actinomycetemcomitans - Adherence
Fusobacterium nucleatum
Streptococcus mutans
Streptococcus sobrinus

References
[128]
[140]

[114]

4.3. Vaccinium macrocarpon

4.5. Morus alba

Vaccinium macrocarpon (Ericaceae) fruit is one of the best

studied anti-adhesion herbs. The ability of cranberry juice or
cranberry extracts to prevent adhesion of oral pathogens has been
widely investigated [59,60]. Flavonoids and proanthocyanidins
isolated from fresh cranberry fruit were found to inhibit growth
and biolm formation of same periodontal pathogens P. gingivalis
[59,60], S. mutans and other oral streptococci [61,62]. However, the
activities of puried phenolic compounds on S. mutans have been
reported to be moderate [63]. The effects were enhanced by a
combination of these compounds showing that the biological activity of cranberry extracts may be a result from the complex
mixture of avonoids rather than a single active compound [63].
Recently, Xie et al. [64], reported that antibacterial effect of
avonoids against a wide range of pathogenic microorganisms may
be due to the inhibition of nucleic acid synthesis, inhibition of
cytoplasmic membrane function and inhibition of the attachment
and biolm formation.

Morus alba (mulberry) native from Asia, is widely exploited for

its varied medicinal properties. Leaves of M. alba contain avonoids
such as apigenin and quercetin [74]. The root bark of M. alba has
been traditionally used in Asian countries for medicinal purposes. It
exhibits anti-viral, cytotoxic and antimicrobial activity against
various microorganisms [75]. The crude extract of M. alba leaves
showed strong inhibitory action against S. mutans adherence on
glass surface in the presence of high sucrose concentrations [76].
Alcoholic extract of M. alba leaves has shown reduction of the
glucosyltransferase activity in terms of glucan production, reduction in the bacterial density of the biolm and the decrease in
thickness of the preformed biolm (Table 1) of S. mutans [77].

4.4. Camellia sinensis

Camellia sinensis (Theaceae) is a sub-shrub native to China.
Green tea made from C. sinensis is a popular beverage throughout
Asia. It provides a dietary source of biologically active compounds
(Table 1) with benecial properties to human [65]. Studies investigating inhibition of bacterial attachment by phytochemicals to
hard surfaces such as enamel have been reported [66]. Extracts of
green and black tea, revealed inhibitory activity against various oral
pathogens [67]. In addition, non fermented and semi fermented
C. sinensis extracts were able to prevent growth of oral streptococci
and to reduce dental caries [67,68]. Limsong et al. [69], prouved that
ethanol extracts of black tea inhibited S. mutans adhesion in vitro.
Green tea extract contains polyphenols with strong antiglucosyltransferase activity and could inhibit experimental dental
caries in rats [70]. The tea polyphenols, especially epigallocatechin
gallate (EGCG), potently blocked in vitro the adhesion of P. gingivalis
and have been shown to inhibit dental plaque accumulation [71]. It
was found that EGCG (7.8e31.25 mg/ml) exhibited dose-dependent
inhibition of the initial attachment of S. mutans UA159 and did not
induce cellular aggregation of S. mutans [72]. On the other hand,
EGCG at sub-MIC level (15.6 mg/ml) signicantly suppressed the gtf
B, C, D genes of S. mutans UA159 compared with the non-treated
control [72]. Recently it has been reported that EGCG effectively
eradicated E. faecalis biolms and induced the formation of hydroxyl radicals in E. faecalis [73]. Therefore, green tea may be used
in mouthwashes or dentifrices for prevention of dental caries and
periodontal diseases.

4.6. Trachyspermum ammi

Trachyspermum ammi (Ajowan), from the Apiaceae family, is an
important commercial product for the food industry. T. ammi has
been used as medicine, primarily to control indigestion and atulence. It is prescribed for colic and diarrhea [78]. On the other hand,
antimicrobial and antioxidant properties have been attributed to
extracts of ajowan seeds [79]. Anticariogenic activity of the crude
extract and some fraction (the petroleum ether fraction) was
noticed also [80]. This activity was explained on the basis of
potentially active classes of compounds (sterols, terpenes, fatty
acids and their derivatives) present in such fractions [80]. In vitro
evaluation of the anticariogenic potential against S. mutans
concluded a signicant effect of T. ammi seeds on dental caries inhibition by reducing the biolm-forming tendency of S. mutans
[81].
4.7. Piper betle
Piper betle L. (Piperaceae) was traditionally used in India and
China in the prevention of oral malodor. A SEM analysis study
conrmed the inhibitory effects of P. betle leaves extract towards
cell adherence, cell growth and extracellular polysaccharide formation of S. mutans [82].
The hydroxychavicol, a phenolic compound present in the
aqueous extract of the P. betle leaf, showed also inhibitory activity
against oral pathogens and prevent the formation of waterinsoluble glucan. Furthermore, it inhibited the growth of biolms
generated by S. mutans and Actinomyces viscosus [83]. In addition,
Nordin et al. [84], reported that adherence of Candida tropicalis,
C. albicans and Candida krusei was reduced after exposing to P. betle
extract by 86.01%, 61.41% and 56.34%, respectively.
4.8. Vitis vinifera
Vitis vinifera L. (grapevine) is indigenous to southern Europe and

B. Kouidhi et al. / Microbial Pathogenesis 80 (2015) 39e49

Western Asia and is cultivated today in all temperature regions of

the world [85]. Its seeds and leaves are used in herbal medicine and
its fruits are utilized as a dietary supplement. It contain phytochemicals witch were able to suppress oral pathogens associated
with dental caries and periodontal diseases [86]. Wine and grapes
contain a number of phenolic compounds that exhibit antimicrobial activity including avanols [87], gallic acid [88], hydroxycinnamic acids [89] and trans-resveratrol [90]. It has been reported
that grape marc extract was able to affect the formation of a
S. mutans biolm and to signicantly reduce the glucotransferase B
(GTF) activity in a dose-dependent manner [91]. Similar results
were obtained with phenolic extracts prepared from pomace of V.
vinifera showing a highly effective glucotransferase inhibitors more
than 60% inhibition at low concentrations [92].
4.9. Azadirachta indica
Azardirachta indica A., (Meliaceae) known as Neem family is a
tree native to the Indian subcontinent and south east Asia [93]. It
has been used for thousands of years as a tool for the treatment of
several diseases in dentistry [94]. Due to its versatility, all parts of
the tree (leaves, owers, seeds, bark and roots) have been used in
traditional medicine as household remedies against various human
diseases [95].
Teeth brushing with Neem sticks and the chewing of its leaves
and seeds are usually used for mouth hygiene [94]. Neem products
(Table 1) has antibacterial [96] and antifungal activity [97] and a
powerful inhibiting agent against various oral microorganisms [98].
In a parallel way, the in vitro radiolabelled adhesion assays proved
that Neem leaves have a signicant anti-adhesive effect [94,99,100].
4.10. Sanguinaria canadensis
Sanguinaria extract is a mixture of benzophenanthridine alkaloids derived from S. canadensis L. (Papaveraceae). Sanguinarine is
widely used for industrial toothpaste and mouthwash products.
Sanguinarine has a broad antimicrobial activity against most
species of plaque associated bacteria as well as antibiolm activity
[101]. In vitro studies indicate that the anti-plaque action of
Sanguinaria is due to its ability to inhibit bacterial adherence to
newly formed pellicule [102].
4.11. Myristica fragrans

45

Panduratin A and isopanduratin A, isolated from the ethanolic

extract of K. pandurata, exhibited strong activities against cariogenic pathogens and periodontopathogens especially S. mutans,
P. intermedia and P. gingivalis [107,108]. Therefore, Panduratin A,
isolated from the rhizome of K. pandurata, is applicable as antibiolm agent (Table 1) to eliminate oral bacterial colonization
during early dental plaque formation [108,109].

4.13. Pistacia atlantica

Pistacia atlantica subsp. Kurdica (Anacardiaceae) is an Iranian
species. The antimicrobial activities of the diethylether mastic gum
extracts of P. atlantica with their phenolic compounds was tested
against S. mutans biolm [110]. The results showed that the extracts
of P. atlantica resin decreased the total viable S. mutans biolms. At
concentrations of 90%, the extract showed an immediate effect on
the biolm bacteria. This activity was also associated with inhibition of adherence to polystyrene surface. Thus, the extract could
successfully prevent plaque formation on the surface of the tooth.
The inhibition of S. mutans adherence by sub-MIC concentrations of
the extract was attributed to avonoids and tannins in the
P. atlantica extract. Flavonoids are known to have antiglucotransferase activity [110]. A avonoid-rich extract (FEPA) of
P. atlantica was also investigated for the inhibition of growth and
acid production of S. mutans, Streptococcus salivarious, Streptococcus
sobrinus, Streptococcus sanguis and A. viscosus [111]. The antiadherence and the antiacidogenic effect of P. atlantica suggest
that this material could be a useful source for the development of
promising anticariogenic agents.

4.14. Pistacia vera

Pistacia vera tree is native of arid zones of Central and West Asia
and distributed throughout the Mediterranean basin. Recently,
antimicrobial properties of polyphenol-rich fractions derived from
raw shelled and roasted salted pistachios has been reported [112].
The ethanolic extracts of P. vera showed a good anti-adherence
activity against S. mutans to glass surface [113]. Kamrani et al.
[113] suggested that the use of the hull extract of P. vera as a mouth
rinse rich in avonoids and tannins will exhibit an effect on the
growth, adherence and acid-producing ability of S. mutans.

Myristica fragrans Houtt. (Myristicaceae), an endemic plant of

Indonesia, has been used traditionally for spice and medicinal
purposes [103].
M. fragrans (nutmeg) extract showed antibacterial activity
against cariogenic S. mutans [104]. Macelignan, one of the bioactive
compounds from nutmeg has potent anticariogenic activity against
S. mutans compared with other commercial antibacterial agents
[104]. At a concentration of 10 mg/mL, it may reduce a 24 h preformed S. mutans biolm after exposure for 5 min [105]. Considering this rapid bactericidal effect and antibiolm activity (Table 1),
macelignan possesses the potential to be practically applied to
toothpaste or mouthwash products and prevent dental caries by
inhibiting the growth of Streptococcus spp.[104].

4.15. Mentha spicata

4.12. Kaempferia pandurata

4.16. Achyranthes aspera

Kaempferia pandurata Roxb. (Zingiberaceae), known as temu

kunci in Indonesia, has been used traditionally for food and medicinal purposes. Flavonoids, such as boesenbergin A and panduratin A, are recognised as the bioactive components of K. pandurata
[106].

Achyranthes aspera L. (Amaranthaceae), is an ethanomedicinal

herb distributed throughout the tropical world which has been
reported to exhibit a potential anti-biolm potential against the
cariogenic S. mutans. The methanolic extract contain bioactive
compounds with high Quorum Sensing response regulator [116].

Mentha spicata (Lamiaceae) essential oil is largely used for

avoring, perfume production, and medicinal purposes. Its antimicrobial properties against a wide variety of bacteria and fungi
have been reported [114]. In vitro and in vivo antimicrobial and
biolm activities of the essential oils from M. spicata on S. mutans
were described [115]. Antibacterial activity of the essential oils from
M. spicata and its ability to affect biolm formation was noticed.
Accordingly, the essential oils from M. spicata signicantly retard
biolm formation that can contribute to the development of novel
anti-caries (Table 1) treatment [115].

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4.17. Artocarpus lakoocha

Aqueous extract Artocarpus lakoocha Roxb., Moraceae showed
variable degrees of antimicrobial inhibition against both Gramnegative bacteria (A. actinomycetemcomitans, P. gingivalis) and
Gram-positive bacteria (S. mutans, S. sobrinus), with MIC ranging
from 0.10 to 0.39 mg/ml. A. lakoocha extract possesses compounds
with good antimicrobial properties that may be used for the
treatment and prevention of oral infectious [117].
4.18. Polygonum cuspidatum
Polygonum cuspidatum (Polygonaceae) has traditionally been
used in Asia to treat skin burn, gallstone, hepatitis, inammation,
and osteomyelitis. In addition, its root has been used to control oral
diseases, particularly biolm-related infection [118].
Methanolic extract of P. cuspidatum has shown a bactericidal
activity and inhibitory effects on virulence factors of S. mutans and
S. sobrinus [119]. Further study has focused on evaluating the effect
of fractions from P. cuspidatum root (Table 1) on dental cariesrelated factors of S. mutans and S. sobrinus [120,121]. An ethyl acetate fraction noted as F1 demonstrated inhibitory effects on
S. mutans biolm and S. sobrinus at sub-MIC levels [121]. The activity of F1 was suggested to be related to the presence of anthraquinones, cardiac glycosides, terpenoids, and phenolics compounds
[121]. In the same way, Ban et al. [120] showed that Ethyl acetate
fraction from P. cuspidatum root present an inhibitory effects on
glycolytic acid production and GTF activity of S. mutans and
S. sobrinus as well as antimicrobial activities.
Recently, a fraction (F1) separated from P. cuspidatum mainly
composed of resveratrol, emodin and physcion (approximately
16.2%, 18.9% and 2.07% of the weight of F1, respectively) demonstrated an inhibitory effects on acid production and F-ATPase activity of S. mutans in biolms [122].
4.19. Helichrysum litoreum
Helichrysum litoreum Guss (Asteraceae) is a species endemic to
Central-South Italy, Sicilia, and Sardinia [123]. Preliminary research
evidenced bactericidal activity of H. litoreum crude extracts, as also
reported for other Helichrysum species [124]. The ethanolic extract
obtained from H. litoreum was showed effective against S. mutans
and A. viscosus with very low cytotoxicity on broblasts and HaCaT
cells lines [123]. In Vivo Assessment of H. litoreum ethanolic extract
efcacity against S. mutans has shown that a regular daily rinsing
with mouthwash containing H. litoreum ethanol extract could
reduce on 50% the salivary levels of S. mutans. This was suggested to
be probably due to both inhibition of growth and adherence of
S. mutans cells to teeth surfaces [123].
4.20. Rosmarinus ofcinalis
Rosmarinus ofcinalis L. (Rosemary) is a spice and medicinal
herb widely used around the world. The antimicrobial effects of
rosemary essential oil against S. sobrinus was reported [125]. In
addition, aqueous and methanolic extracts of rosemary markedly
inhibited the formation of water-insoluble glucan of S. sobrinus.
This results suggest that rosemary extract may be effective for the
inhibition of the growth of cariogenic oral streptococci [126].
4.21. Eugenia caryophyllata
Eugenia caryophyllata L. (Myrtaceae) is widely used and well
known for its medicinal properties. Clove oil is traditionally used in
dental care as an antiseptic and analgesic. It is active against oral

bacteria associated with dental caries and periodontal disease

[127,128] and effective against a large number of other pathogenic
bacteria [129]. Eugenol, the major phenolic component of clove
essential oil, is widely used in medical and dental practice, due to
its potent fungicidal, bactericidal and anesthetic properties [130].
Other active constituents of clove (biorin, kaempferol, rhamnocitrin, myricetin, gallic acid, ellagic acid and oleanoic acid) showed
antibacterial activities against Gram-negative anaerobic periodontal oral pathogens, including P. gingivalis and P. intermedia
[127].
Recently, expression of biolm- and QS-related genes (gtfB, gtfC,
comDE, smu630, vicR, brpA, ftf, relA, gbpB and spaP) checked by
quantitative real-time PCR revealed that eugenol at a subminimum inhibitory concentration signicantly downregulated
these genes in S. mutans. Thus, the results indicated that eugenol
can inhibit caries-associated biolm and showed its therapeutic
potential against oral biolm [131].
4.22. Eucalyptus
The Eucalyptus (Myrtaceae) is a tree, native to Australia and
Tasmania, successfully introduced worldwide. Its therapeutic uses
have been proved in traditional Chinese and Indian medicine.
Specic activity against periodontal and cariogenic bacteria, such as
P. gingivalis, A. actinomycetemcomitans, Fusobacterium nucleatum,
S. mutans, and S. sobrinus has been reported [114]. Eucalyptol (1, 8cineole), the active ingredient of the eucalyptus oil showed antibiolm activity against C. albicans biolms [132,133].
5. Conclusion
This review present the mechanism of drug resistance developed by bacteria embedded in biolm and the potential use of
medicinal plant (crude extracts, essential oils and puried compounds) as bioactive agents with a signicant antibiolm effect
against oral pathogenic bacteria. By employing such antibiolm
molecules, microbial pathogens would be rendered more susceptible to broad used conventional drugs.
However, further studies at cellular levels and in vivo studies are
needed to elucidate the molecular mechanism by witch these
natural products inhibit bacterial adhesion and minimize biolm
formation.
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