By Dr.

Nitika Jain
Post Graduate Student

Dental PLAQUE
Physiological approaches to
the control of oral biofilms
Physiological strategies for the control
of oral biofilms
Prevention of low pH

 Bacterial homeostasis in plaque biofilms

would be less likely to be disrupted if the
frequency and depth of acidic conditions
following sugar intake could be reduced. This
could be achieved physiologically by
 (a) inhibitors of acid production,
 (b) consumption of food or drinks containing non-
fermentable sweeteners, and
 (c) the local generation of base (alkali) in plaque.
Virulence Factors of
Periodontopathogens
 Virulence factors of periodontal micro-
organisms can be subdivided into
 (1) factors that promote colonization (adhesins),
(2) toxins and enzymes that degrade host tissues,
(3) mechanisms that protect pathogenic bacteria
from the host.
Adhesive Surface Proteins and Fibrils

 Fimbriae, or pili, are polymeric fibrils composed

of repeating subunits that can extend several
microns from the cell membrane.
 Pili were once thought to be unique to gram-
negative bacteria but have now been identified
in several gram-positive organisms, including
streptococci and actinomyces.
 Strains of P. gingivalis produce two types of
fimbriae, known as the major and minor fimbriae.
Factors that Promote Tissue
Destruction
 The gingipains belong to the cysteine protease
family, which utilize an active site cysteine residue
for catalysis.
 Gingipains are classed as “Arg-gingipains” (RgpA
and RgpB) or “Lys-gingipain” (Kgp), based on their
ability to cleave Arg-Xaa or Lys-Xaa peptide bonds
(Xaa represents any amino acid).
 Gingipains are multifunctional proteins that play
important roles in adhesion, tissue degradation,
and evasion of host responses.
 Hyperleukotoxic strains include the JP2 clone
of A. actinomycetemcomitans, which is
uniquely associated with localized aggressive
periodontitis.
Strategies for Evading Host Immunity

 Pathogenic bacteria have many and varied

strategies for evading or subverting the host
immune system, including
 (1) the production of an extracellular capsule,
 (2) proteolytic degradation of host innate and/or
acquired immunity components,
 (3) modulation of host responses by binding
serum components on the bacterial cell surface,
 (4) invasion of gingival epithelial cells.
Microorganisms Associated
With Periodontal Health
 The bacteria associated with periodontal health are
primarily gram positive facultative species and
members of the genera Streptococcus and
Actinomyces (e.g., S. sanguis, S. mitis, A. viscosus,
and A. naelslundii).
 Small proportions of gram negative species are also
found, most frequently P. intermedia, F. nucleatum,
and Capnocytophaga, Neisseria, and Veillonella spp.
 Microscopic studies indicate that a few spirochetes
and motile rods also may be found
 Certain bacterial species have been proposed to be protective or beneficial
to the host, including S. sanguis, Veillonella parvula, and C. ochracea. They
are typically found in high numbers at periodontal sites that do not
demonstrate attachment loss (inactive sites) but in low numbers at sites
where active periodontal destruction occurs.
 Socransky SS, Haffajee AD; JP 63: 322, 1992.

 These species probably function in preventing the colonization or

proliferation of pathogenic microorganisms.
 One example of a mechanism by which this may occur is the production of
H2O2 by S. sanguis; H2O2 is known to be lethal to cells of A.
actinomycetecomitans.
 (Hillman JD, Socransky SS “the relationships between streptococcal species and
periodontopathogenic bacteria in human dental plaque” Arch Oral Biology 30: 791; 1985)
Microorganisms Associated with
Specific Periodontal Diseases

 Microbiologic procedures clearly demonstrated

that the number and proportions of different
subgingival bacterial groups varied in
periodontal health when compared with the
disease state . The total number of bacteria,
determined by microscopic counts per gram of
plaque, are twice as high in Periodontally
diseased sites than in healthy sites.
Microorganisms Associated with
Specific Periodontal Diseases
 On culturing, bacteria from healthy
periodontal sites it consists mainly of
 Gram positive facultative rods and cocci ( 75%)
 In gingivitis, (44%) and
 periodontitis ( 10 – 13%)

 Increase in proportions of gram negative

rods,13% to 40% in gingivitis and 74% in
advanced periodontitis.
Gingivitis:

 Model system for experimental gingivitis –

described by Loe et al (Loe H; JP 36: 177,
1965) and Theilade et al.
Gingivitis:

 The bacteria found in naturally occurring dental plaque-induced

gingivitis (chronic gingivitis) consist of roughly
 equal proportions of gram-positive (56%) and gram-negative (44%)
species,
 facultative (59%) and
 anaerobic (41%) microorganisms.
 Predominant gram-positive species include S. sanguis, S. mitis, S.
intermedius, S. oralis, A. viscosus, A. naelslundii, and
Peptostreptococcus micros.
 The gram-negative microorganisms are F. nucleatum,P. intermedia,
V. parvula, as well as Hemophilus, Capnocytophaga and
Campylobacter spp
 Pregnancy associated gingivitis :
 This condition is accompanied by increases in
steroid hormones in crevicular fluid and increases
in the P.intermedia, which uses the steroid as
growth factors.( Kornman KS , Loesche WJ ; Infect
Inhuman 35: 256 , 1982)
Chronic Periodontitis:

 Microscopic examination of plaque from sites

with chronic periodontitis have consistently
revealed elevated proportions of
spirochetes .
 Cultivation of plaque microorganisms from
sites of chronic periodontitis reveals high
percentages of anaerobic (90%) gram-
negative (75%) bacterial species
 In chronic periodontitis, the bacteria most often cultivated at high levels
include P. gingivalis, B. forsythus, P. intermedia, C. rectus, Eikenella
corrodens, F. nucleatum, A. actinomycetemcomitans, P. micros, and
Treponema and Eubacterium.

 When Periodontally active sites (i.e., with recent attachment loss) were
examined in comparison with inactive sites (i.e., with no recent
attachment loss), C. rectus, P. gingivalis, P. intermedia, E. nucleatum,
and B. forsythus were found to be elevated in the active sites.

 Detectable levels of P. gingivalis, P. intermedia, B. forsythus, C. rectus, and

A. actinomycetemcomitans are associated with disease progression and
elimination of specific bacterial pathogens with therapy is associated
with an improved clinical response.
 Recent studies have documented an association
between chronic periodontitis and viral
microorganisms of the herpes viruses group, most
notably Epstein-Barr Virus-1 (EBV-1) and human
cytomegalovirus (HCMV).
 ( Contreras A, Slots J: JPR; 35: 3, 2000)
 Presence of subgingival EBV-1 and HCMV are
associated with high levels of putative bacterial
pathogens, including P. gingivalis, B. forsythus,
P. intermedia, and T. denticola
Localized Aggressive Periodontitis:

 Localized aggressive periodontitis develops

around the time of puberty, is observed in
females more often than in males, and typically
affects the permanent molars and incisors.

 The microbiota associated with localized

aggressive periodontitis is predominantly
composed of gram-negative, capnophilic, and
anaerobic rods.
 Microbiologic studies indicate that almost all LJP sites harbor
A. actinomycetemcomitans ( Moore WE ; JPR: 22; 235, 1987)

 Other organisms found in significant levels include P. gingivalis,

E. corrodens, C. rectus, E nucleatum, B. capillus, Eubacterium
brachy, and Capnocytophaga spp. and spirochetes.
 Herpesviruses, including EBV-1 and HCMV, also have been
associated with localized aggressive periodontitis.
 (Michalowicz BS, Ronderos M; “ human herpes virus and P. gingivalis
are associated with juvenile periodontitis” JP 71: 981, 2000)
 After initial colonization of the first permanent teeth
to erupt ( molars and incisors), immune defenses are
stimulated to produce opsonic antibodies.
 Bacteria antagonist to A.a may colonize the
periodontal sites and inhibit A.a from further
colonization.
 A.a may loses its toxin producing capacity for
unknown reasons.
 A defect in cementum formation may be responsible
for the localization of the lesions.
Necrotizing Periodontal Diseases:

 Acute inflammation of the gingival and periodontal

tissues characterized by necrosis of the marginal
gingival tissue and interdental papillae.
 Microbiologic studies indicate that high levels of
 P. intermedia and spirochetes are found in necrotizing
ulcerative gingivitis lesions.
 Spirochetes are found to penetrate necrotic tissue
and apparently unaffected connective tissue.
▪ (Lisgarten MA, Socransky SS; “ relative distribution of bacteria at
clinically healthy and periodontally diseased sites in humans” JCP
5:115; 1978)
Periodontal abscess
 Periodontal abscess are acute lesions that result in very rapid
destruction of the periodontal tissues.
 Often occur in patients with untreated periodontitis but can
be seen in maintenance phase or in the absence of
periodontitis also ( for example associated with impaction of
a foreign objects or with endodontic problems)
 Clinical symptoms :

 Periodontal pathogens are commonly found in significant

numbers in periodontal abscesses include F. nucleatum, P.
intermedia, P. gingivalis, P. micros, and B. forsythus. (Newman
MG, Socransky SS ; JPR 12: 120, 1997)
Periimplantitis
 Periimplantitis refers to inflammatory process affecting tissues around
already osseointegrated implant & results in loss of supporting bone.
 Healthy periimplant pockets are characterized by high proportions of
coccoid cells, low ratio of anaerobic & aerobic species, low numbers
of peiodontal pathogens.
 Implants with periimplantitis reveals species such as A.a, P. gingivalis,
T. forsythia, P. micros, C. rectus, Fusobacterium, & Capnocytophaga
are isolated from failing implants.
 Other species such as p. aeruginosa,
enterobacteriaceae
C. albicans, & staphylococci are
also detected around implants.
 Lee et al. used the cluster method developed by
Socransky et al.to analyze the microbiota
associated with healthy implants and study the
implant and host-related factors able to influence
the microbial film.
 Rams et al. detected “corn cob” morphology around
healthy implants and “brush forms” around failing
fixtures.
▪ (Rams TE, Link CC Jr (1983) Microbiology of failing dental implants
in humans: electron microscopic observations. J Oral Implantol
11:93–100)
Key characteristics of specific
periopathogens
AGGREGATEBACTER
ACTINOMYCETECOMITANS:
 Forms: multiple biotypes
5 serotypes ( a to e)
on the basis of
difference in
polysaccharides.
 Strains from patients in
Africa have an increased
leucotoxin production.
Culture condition & identification

 It grows as white, translucent, smooth, non

hemolytic colony on blood agar.
 Preferably identified on a specific growth
medium ( with vancomycin and bacitracin as
antibiotics to suppress other species)under 5
to 10% CO2.
Small, non-motile, Gram-negative saccharolytic,
capnophilic, round-ended rod that forms small,
convex colonies with a "star-shaped" center
Special pathogenic characterstics

Leukotoxin
it forms the pore
in neutrophil Collegenas
Lipo granulocytes,
monocytes, and e
polysaccha lymphocytes Protease
destruction
ride which able to
consequently die of
Endotoxin of osmotic cleave Ig G
pressure
connective
(Baehni et al. tissue
1979), a cytolethal
distending toxin .
 A. actinomycetemcomitans has been shown,
in vitro, to have the ability to invade cultured
human gingival epithelial cells (Blix et al.
1992, Sreenivasan et al. 1993), human
vascular endothelial cells (Schenkein et al.
2000) and buccal epithelial cells in vivo.
Tannerella forsythia:
 It is a non motile, spindle shaped, highly pleomorphic
rod & gram negative obligate anaeobe.
 Culture condition & identification: It takes 14 days to
form minute colonies, only under anaerobic conditions
& needs several growth factors (N-acetylmuramic acid)
from other species (e.g; F. nucleatum).

 Pathogenicity: It produces several Proteolytic enzymes

that are able to destroy immunoglobulin & factors of
complement system. It also induces apoptotic cell
death.
 This organism was found in higher numbers
in sites of destructive periodontal disease or
periodontal abscesses than in gingivitis or
healthy sites (Lai et al. 1987, Herrera et al.
2000, Papapanou et al. 2000).
Porphyromonas gingivalis:

 Is non motile,
pleomorphic (coccal to
short) rod & gram
negative obligate
anaerobe.
 P. gingivalis is a
member of "black- The black-pigmented colony is an
pigmented isolate of Porphyromonas gingivalis

Bacteroides" group
Culture conditions & identification

 It grows anaerobically , with dark

pigmentation ( brown, dark green, or black)
on blood agar because of a metabolic end
product from blood( hemin)
Virulence factors

proteases
Destruction of Ig,
complement
factors, heme-
sequestering
hemolysin collagenases
proteins,
degradation of
host cell
collagemnase
inhibition
 P. gingivalis can inhibit migration of PMNs
across an epithelial barrier (Madianos et al.
1997) and has been shown to affect the
production or degradation of cytokines by
mammalian cells (Darveau et al. 1998,
Fletcher et al. 1998, Sandros et al. 2000).
Prevotella intermedia & Prevotella
nigrescens:

Prevotella group are

short , round ended
non motile gram
negative rods
They grow anaerobically ,
with dark pigmentation
( brown black colonies) The dark-pigmented
colonies are isolates of Prevotella intermedia
on blood agar.
The levels have been shown to be particularly
elevated in acute necrotizing ulcerative
gingivitis ( Loesche et al. 1982)

in certain forms of periodontitis (Tanner et al.

1979, Dzink et al. 1983, Moore et al. 1985,
Maeda et al. 1998, Herrera et al. 2000,
Papapanou et al. 2000),
Fusobacterium nucleatum:

 F. nucleatum is a Gram-
negative, anaerobic, cigar
shaped bacillus with pointed
ends that has been recognized
as part of the subgingival
microbiota.
 It grows anaerobically on
blood agar.
 Several serotypes are
 F. nucleatum ss nucleatum
 F. nucleatum ss polymorphum F. Nucleatum on specific medium
 F. nucleatum ss vincentii
 F. periodonticum
 F. nucleatum is prevalent in subjects with periodontitis (Papapanou
et al. 2000, Socransky et al. 2002) and periodontal abscesses (Her
rera et al. 2000, Newman MG, Socransky SS ; JPR 12: 120, 1997).

 Invasion of this species into human gingival epithelial cells

in vitro
was accompanied by an increased secretion of IL-8 from the
epithelial cells (Han et al. 2000).

 The species can induce apoptotic cell death in mononuclear and

polymorphonuclear cells (Jewett et al. 2001) and cytokine,
elastase and oxygen radical release from leukocytes (Sheikhi et al.
2000).
Campylobacter rectus:
 C. rectus is a Gram-negative,
short rod, curved ( vibrio) or
helical.The motility is due to polar
flagellum.

 The organism is unusual in that it

utilizes H2 or formate as its
energy source.
 It was first described as a member
of the "vibrio corroders", a group
of short nondescript rods that
formed small convex, "dry
spreading" or "corroding" (pitting)
colonies on blood agar plates.
 C. rectus has been shown to
produce a leukotoxin, and is less
virulent and less proteolytic than
P. gingivalis
Eikenella corrodens:

 E. corrodens is a Gram-negative, capnophilic,

asaccharolytic, regular, small rod with blunt ends.
 E. corrodens has also been found in association
with A. actinomycetemcomitans in some lesions of
LJP (Mandell 1984, Mandell et al. 1987).
 E. corrodens has been shown to stimulate the
production of matrix metalloproteinases (Dahan
et al. 2001) and IL-6 and IL-8 (Yumoto et al. 1999).
Peptostreptococcus micros:

 P. micros is a Gram-positive, anaerobic, small,

asaccharolyticcoccus.
 Two genotypes can be distinguished with the
smooth genotype being more frequently
associated with periodontitis lesions than the
rough genotype (Kremer et al. 2000).
 P. micros in combination with either P.
intermedia or P. nigrescens could produce
transmissible abscesses (van Dalen et al. 1998).
Selenomonas species:
 The organisms may be recognized by their curved shape,
tumbling motility and, in good preparations, by the presence of a
tuft of flagella inserted in the concave side.
 The Selenomonas spp. are Gram-negative, curved, saccharolytic
rods.
 Moore et al. (1987) described six genetically and phenotypically
distinct groups isolated from the human oral cavity.
 Selenomonas noxia was found at a higher proportion of shallow
sites (PD < 4 mm) in chronic periodontitis subjects compared with
similar sites in periodontally healthy subjects (Haffajee et al.
1998). Further, S. noxia was found to be associated with sites that
converted from periodontal health to disease (Tanner et al. 1998).
Eubacterium species:

 E. nodatum, Eubacterium brachy and

Eubacterium timidum are Gram positive,
strictly anaerobic, small, pleomorphic rods.
 They are often difficult to cultivate,
particularly on primary isolation, and appear
to grow better in roll tubes than on blood
agar plates.
 Spirochetes:

 Spirochetes is a group of spiral,

motile organisms.
 They are helical rods 5 to 15µm
long with a diameter of o.5µm.
 They have 3 to 8 spirals.

 Spirochete has been implicated as

the likely etiologic agent of acute
necrotizing ulcerative gingivitis by
its presence in large numbers in The sample is
tissue biopsies from affected sites dominated by large spirochetes with
(Listgarten & Socransky 1964, the typical cork-screw appearance.
Listgarten 1965).
 At least 15 species of subgingival spirochetes have been
described, spirochetes are combined in a single group or
groups based on cell size; i.e. small, medium or large.

 Spirochetes includes T. denticola, T. vincentii, T. socranskii,

often associated with periodontitis.
 Some spirochetes have capacity to migrate within GCF &
penetrate epithelium & connective tissue.
 Some have capacity to degrade even dentin
 T. denticola produces Proteolytic enzymes that can destroy
IgA, IgM, IgG, or complement factors.
 VIRUSES - Contreras & Slots 2000, Kamma et al 2001

12/27/2011 DENTAL PAQUE 57

 Four major viral families are associated with the main viral
oral diseases of adults, as follows:

 1. The group of herpesviruses contains eight different

members that all are enveloped double-stranded DNA
viruses. In the oral cavity, they are related to different ulcers,
tumors, and other oral pathoses.

 2.Human papillomaviruses are grouped within five genera

and are nonenveloped double-stranded DNA viruses. In the
oral cavity, they are related to ulcers, tumors, and oral
pathoses.
12/27/2011 DENTAL PAQUE 58
 3. Picornaviruses are all nonenveloped,
single-stranded RNA viruses. In the oral
cavity, they are related to ulcers and different
oral pathoses
 4. Retroviruses. All retroviruses are
enveloped single-stranded RNA viruses. In
the oral cavity, they are related to different
tumors and oral pathoses.

12/27/2011 DENTAL PAQUE 59

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Yeast

 The majority of isolates are Candida, and the

most prevalent species is C. albicans.
Together with C. albicans, some of the most
common opportunistic fungal pathogens in
humans are C. tropicalis, C. glabrata, C. krusei,
C parapsilosis, C. guilliermondii, and C.
dubliniensis.
Etiologic agents for different types of
oral yeast infection
Protozoa
 Only a few parasites affect the oral cavity, but an increasing
body of literature claims that oral protozoa are more
common than previously appreciated.
 Depending on the type of infection, the parasitic infectious
agents can be divided into two categories: 1.those that
induce local infections and those that induce systemic
infections with indirect effects on the oral cavity. The former
group comprises saprophytes (such as Entamoeba gingivalis
and Trichomonas tenax) 2.that have the potential to turn
into opportunistic pathogens, or free-living amoebae that
occasionally become invasive but seldom present clinically.
Archaea

 Archaea are single-celled organisms that are distinct

from the bacteria as they are from eukaryotes. The
role of archaea in oral diseases is only beginning to
be explored. Methanogenic archaea produce
methane from hydrogen gas (H2)/carbon dioxide
(CO2) and sometimes from formate, acetate,
methanol, or methylamine. These organisms have
been isolated from patients with periodontal disease
by enriching cultures with H2 and CO2.
 Subsequently, a clear correlation between the presence of
archaeal DNA and periodontal disease was established. (Lepp
PW, Brinig MM, Ouverney CC, et al: Methanogenic Archaea
and human periodontal disease. Proc Natl Acad Sci U S A
2004;) 101:61-76).Despite these findings, virtually nothing is
known about the role of archaea in periodontitis.
 Archaea have also been detected in endodontic infections,
and again their contribution to disease is unknown. (Vianna
ME, Conrads G, Gomes BP, et al:
Identification and quantification of archaea
involved in primary endodontic infections. J Clin Microbiol 2006;
44:12-74.)
Conclusion

 The periodontal microbiota is a very complex

ecologic system with many structural and
physiologic interactions among the resident
bacteria and between the bacteria and the host. It
is clearly possible that levels of a particular
species may be elevated as a result of
environmental changes produces by the disease
process and may not be a causative agent. It is
also difficult to ascertain the specific etiologic
pathogens in periodontitis.
References:
 Newman, Takei, Klokkevold, Carranza. Carranza’s
Clinical Periodontology, 10th Edition and 11th Edition
 Lindhe, Lang, Karring. Clinical Periodontology &
Implant Dentistry, 5th Edition.
 Philip D Marsh, Michael V Martin, Oral Microbiology,
5th Edition.
 Perio 2000 - 49:60, 2009
 Perio 2000 - Volume 52 Issue 1, Pages 7 - 121 (Feb 2010)
 “Advnces in dental research” ADR 1997 11: 176
 Philip D. Marsh