P gingivalis

P GINGIVALIS
Dr p mikitha
Ii mds
Deptof periodontology
Aecsmaaruti collegeof dentalsciencesandresearchcentre

Session 1
CONTENTS
 Introduction
 Taxonomy
 Biochemical characteristics
 Bacterial structure of p. gingivalis
 Microbiological tests
 Oral ecology
 Transmission
 P. gingivalis in human periodontal disease: occurrence & treatment
 Virulence factors

INTRODUCTION
 Periodontal diseases comprise a group of infections involving the
supporting tissues of the teeth, these range in severity from mild and
reversible inflammation of the gingiva to chronic destruction of
periodontal tissues with eventual exfoliation of teeth.
 The bacterial etiology is complex, with a variety of organisms responsible
for the initiation and progression of disease.
 The classification of the various manifestations of periodontal diseases is
continually changing.
 The nature of the pathogenic agents varies among these disease entities,
as well as among patients and even between different disease sites within
a patient.

TAXONOMY
 Family Bacteroidaceae
genera Bacteriods
Fusobacterium
Leptotrichia
 Emergence of the genus Porphyromonas and taxonomic position of
P. gingivalis
How KH et al. Life below the gum line: Pathogenic mechanisms of Pg. Microbiology and Molecular Biology Reviews, December 1998, 1244-1263.

BIOCHEMICAL PROPERTIES
 Asaccharyolytic & anaerobic
 Saliva and mucous membranes – temporary transition phase
 Subgingival ecosystem is ideal
Nutrients- Redox potential
Peptides & aa
Vit K, Vit B12 & heme

Samplingtechniques
Subgingival plaque -
 Mombelli et al (1991) showed that 4 individual subgingival
specimens, each from the deepest periodontal pocket in each
quadrant, should be included to detect Pg
 Christersson et al (1992) also showed that sampling 3 pockets with 5
mm or greater pocket depth provided a probability of 95% of
detecting Pg
Whole saliva sample seems to be a reasonably good substitute for
subgingival samples

Current methods for detection
Method Molecular target Detection limit
Culture 0.1-0.2% of total viable
count
Immunodiagnostic
Indirect immunofluorescence Polyclonal
antibodies to
whole-cell
antigens
500 cells/ml

Flow cytometry
Evalusite
Bacterial
concentration flow
cytometry
Monoclonal
antibodies to
LPS
Polyclonal
antibodies to
serotype
antigens
Monoclonal
antibodies to
whole-cell
antigens
100 cells/ml
106 cells
104 cells

Nucleic acid probe
Digoxigenin-labled
whole genomic probe Genomic DNA 103 cells
DMDx Genomic DNA
Radiolabeled
oligonucleotide
16SrRNA genes 103 cells
PCR 16SrRNA genes 25-100 cells
Nested PCR Space region bw
16SrRNA & 23rRNA
genes
10 cells

Multiplex PCR 16SrRNA genes 5-50 cells
Immunodiagnostic Specific Pg DNA
region of unknown
functions
16 cells

CULTURE
 Non selective
 Selective
Grow in culture media forming convex, smooth glossy colonies of 1-
2mm diameter, which demonstrate a progressive darkening in the
center, because of production of protoheme, the substance
responsible for the typical color of these colonies

BANA
 Tf, Pg , Td and capnocytophaga species have a trypsin-like enzyme in
common
 Activity of the enzyme can be measured with the hydrolysis of the
colorless substrate BANA
 Diagnostic kit – Perioscan
 Loesche et al (1992) and Beck et al (1990) used this assay in their study

 Real-time PCR technology has demonstrated a high degree of sensitivity and
specificity
 Culture, checkerboard analysis and real-time quantitative PCR require
sophisticated lab facilities, labor intensive and expensive
 Only culture enables to study of antibacterial susceptibilities and detection of
unexpected bacteria
 To date, there is no ideal microbial diagnostic for adjunctive clinical use in
periodontics

ORAL ECOLOGY
&
PERSON-TO-PERSON TRANSMISSION

The natural habitat
 No evidence that they live free in nature
 Human isolates phenotypically different from those of other mammals
 Occurs in pockets & mucosal surfaces
 Not found in edentulous babies & elderly people
 Do not belong to the indigenous microbiota of any other body site
 Can cause nonoral infections

Initial colonization on the tooth surfaces
 Does not seem to colonize clean tooth surface
 Prefers sites showing inflammation, poor OH, & sites harboring G+
bacteria
 Absent/infrequent in primary dentition of primary teeth in healthy
children
 Efficiently equipped to colonize supragingival tooth surfaces,
probably at any area of the dentition

Distribution pattern within individuals
 Widely distributed, which might be related to the lack of efficacy in the
antibody response
 Occurrence of Pg +ve patients increases with age
 The noted distribution may be related to the efficacy of antibody
response against the in.org. It has been suggested that the IgG response
to Aa is protect and able to limit infection arounded a few teeth,
whereas elevated antibody titers to Pg are not capable of controlling it,
which may lead to generalized periodontal destruction

Proportions of flora, balance-imbalance
 In periodontal healthy sites, present in very low proportions
 High proportions in disease- breakdown in homeostasis
 Difference bw Aa & Pg colonization was demonstrated by Torkko &
Asikainen (1996) and Rams et al (1997) in AP patients. Aa- 4%in both,
whereas Pg 16% & 23% respectively

Genetic diversity of Pg in nature
 The population genetic study of bacteria arose from interest in evolutionary
processes of organisms, but has also provided important insight into bacterial
pathogenesis
 Studies on pathogenic bacteria have suggested that many, but not all,
bacterial pathogens exist as discrete clones in nature: that is, the genetic
changes due to recombination occur so infrequently that the genotypes of
bacteria remain stable over significantly long periods of time. Moreover,
specific clones may be responsible for a majority of infectious diseases
ascribed to a pathogen

 Loos et al. (1993) examined the population genetic structure of Pg by using
multilocus enzyme electrophoresis on 88 human Pg strains.
 Most strains originated in the subgingival sites of AP or other oral infections
and only two isolates in non-periodontitis individuals.
 The results suggested that the population structure of Pg is clonal, with
certain clones being wide spread in geographically diverse regions. No
association was found between specific clusters and disease types
(periodontal diseases, endodontic infections or other oral infections) or the
ability of the strains to cause spreading or localized infections in the mouse
model.

Genetic diversity of Pg within individuals
 Studies on Pg show that most individuals are infected with a single
clone of this species

Transmission
 The behaviour of a pathogenic microorganism depends upon the
interaction between the host, the pathogen and the environment.
Changes in any of these factors will affect the likelihood of
transmission
 Host factors include, among others, sexual behaviour, hygiene,
occupation, nutrition, immunity and general health.
 Microbial factors include infectiousness, pathogenicity, virulence and
survival in human and animal hosts, including resistance towards
immune responses and drugs.
Van Winkelhoff, A. J., & Boutaga, K. Transmission of periodontal bacteria and models of infection. Journal of Clinical Periodontology. 2005;
32(s6), 16–27.

 Environmental factors such as temperature, humidity and oxygen
tension are determinants of the survival of microbes outside the host.
 When transmission is directly from parents to offspring (via ovum,
placenta, milk, blood, saliva) it is referred to as vertical.
Transmission is called horizontal when an individual infects
unrelated individuals by contact, respiratory or faecal–oral spread.
32(s6), 16–27.

Vertical transmission
 Vertical transmission of P. gingivalis may occur but has only rarely been
observed based on genotyping of the isolates from parents and children. The
Tuite-McDonnell et al.’s (1997) study has suggested that vertical
transmission of P. gingivalis can occur, but this study does not provide
conclusive evidence as genotyping has not been performed.
 A study conducted by Tiute-McDonnell et al (1997) used PCR assay to
detect oral Pg from members of 104 randomly selected multigeneration
families. Results showed that finding an infected family member
substantially, increased the relative risk of detecting Pg from other family
members.
32(s6), 16–27.

 This implies intrafamilial transmission of Pg, although it does not
prove it. It is possible that members share habits, such as poor OH
and deficient dental care, which may enhance colonization of this
species and lead to acquisition strains from extrafamilial sources as
well.
32(s6), 16–27.

Horizontal transmission
 Can occur bw siblings or spouses
 Two studies, Petit et al (1993) & Saarrela (1996) both reported that siblings
harbored identical genotypes of P gingivalis. The reasons for the strain
identity in siblings may be that the siblings have transmitted the strain to
each other or that each sibling has acquired the strain directly from
bacterium-positive parent. However, the study designs do not permit
distinction between the horizontal and vertical modes of transmission
32(s6), 16–27.

 Transmission of Pg is rather uncommon between spouses in regard to their
intimate co-habitance of several years
 P. gingivalis among cohabiting adults may be interpreted in several ways.
Transmission occurred more frequently, but did not lead to persistent
colonization or detectable levels of the organism, methods did not allow
recovery of the organism or of all colonizing clones, colonization is host-
dependent, colonization depends on the characteristics of the strain, the
numbers of bacteria in the transfer inoculation were too low or exposure to
infection occurred too rarely
The results also suggest that established oral microbiota does not easily accept
new bacterial invaders.
32(s6), 16–27.

 Associations between bacteria and disease have been well established for A.
actinomycetemcomitans and P gingivalis .Recent evidence suggested the
possibility that people with periodontitis may cause periodontal breakdown
in their spouses (Von Troil-Linden et al 1995).
 The study design included 20 married couples. Two kinds of probands were
selected according to periodontal status. One group of probands exhibited
advanced periodontitis and the other group was periodontally healthy.
 The spouses of these probands volunteered to participate in the study. The
inclusion criteria were that the couples were in their middle ages and had
been married for more than ten years.
Clinical significance of transmission
32(s6), 16–27.

 The results showed that the spouses of the diseased probands had more
frequently deep periodontal pockets, radiographic attachment loss and
periodontal pathogens than the spouses of the healthy probands. The
difference could not be explained by sex, age or social status. Neither was
there a difference in the level of oral hygiene between the two spouse groups.
 However, different results were obtained in a seven-year longitudinal study
on young Indonesian couples who had been married for on the average of ten
years (Van der Velden et al 1996). The results showed that the cohabitation
did not lead to change in the periodontal condition of the spouse.

32(s6), 16–27.

Route of infection from person to person
 The role of saliva as a transport vehicle has been supported by the
findings that A. actinomyceterncomitans and P gingivalis can be cultured
from salivary samples, which indicates that these bacteria are able to
survive in saliva during transportation to a new host.
 However, compared with salivary inoculation, mucosal contact or
sharing a toothbrush may allow direct implantation of bacteria to
potential growth locales.

 Saliva is a likely transport vehicle for transmitting strains of Pg, along with
many other organisms. This may occur especially easily to children. Saliva of
the caregiver can be brought to the child's mouth when caring for a young
child. It is not known what the size of the inoculum is or how many events of
exposure are needed to maximize the possibility for success- fully
transmitting Pg.
 Probability that the higher their load in saliva, the greater the risk of
colonization of the recipient. Suppression of the organisms in saliva may
prevent their spread among individuals.
 Periodontal treatment can suppress salivary Aa & Pg under detection level or
strongly decrease their numbers in patients with periodontitis for atleast 6
months.

Pg IN HUMAN DISEASE: OCCURRENCE & TREATMENT
 The designation of Aa & Pg as periodontal pathogens presupposes
that destructive periodontal disease is more prevalent in periodontal
sites exposed to the organisms than in non-exposed periodontal sites.
 It also assumes that future destruction takes place more frequently in
periodontal sites exposed to the organisms than in those not exposed
Griff en AL, Becker MR, Lyons SR, Moeschberger ML, Leys EL. Prevalence of porphyromonas gingivalis and periodontal health status. J Clin Microbiol 1998;36:3239-
42.

Subgingival prevalence of Pg
 Most authors agree that periodontally healthy children and adolescents
harbor few or no P gingivalis in the subgingival microbiota.
 However, based on DNA probe analysis, Bimstein et al. (1996) described
subgingival P gingivalis in many healthy children.
 Chen & Slots (1999) used DNA probe identification may overestimate
the prevalence of the organism due to cross-hybridization with other
subgingival species.
42.

 P gingivalis has been described in 37-63% of LJP patients. However, the
organism is rarely found at the debut of the disease and tends to comprise
only a small part of the microbiota in early disease stages.
 In contrast, P gingivalis is a predominant organism in GJP and may assess
pathogenetic significance in the disease.
 Adults having a healthy and minimally diseased periodontium reveal
subgingival P gingivalis in less than 10% of study sites.
42.

 On the other 40-100% of AP patients may yield the organism .
Furthermore, P g comprises a considerably higher proportion of
subgingival microbiota in deep than in shallow periodontal pockets .
 The high occurence of P gingivalis in AP lesions together with the
organism's pathogenic potential are main reasons for designating the
species as a prime pathogen of AP
42.

P gingivalis virulent strains
 P gingivalis strains vary in pathogenicity .
 Based on experimental infections in animals, P gingivalis strains of high
virulence cause disseminated infections with severe tissue damage and death
whereas less virulent or noninvasive strains prod localized abscess with
milder tissue damage.
 Laine & van Winkelhoff (1998), studying the virulence of capsular serotypes
of P gingivalis in a mouse model, found encapsulated strains to produced
phlegmonous, spreading type of infections, whereas less virulent non-
encapsulated strains produced localized abscesses
42.

 Dahlen and Slots (1989) comparing the growth and pathogenicity of P
gingivalis strains in subcutaneous tissue chambers implanted in rabbits,
found the organism to be particular virulent in co-infection with
A.actinomycetemcomitans. Differences in collagenolytic activity among P
gingivalis strains may account for some of the observed variation in P
gingivalis virulence.
 The search for highly virulent genotypes of P gingivalis has not been very
rewarding
42.

Pg in ailing or failing implants & in periodontal infection
 P gingivalis are rare isolates from healthy mucosal sites around integrated
dental implants.
 Mengel et al. (1996) studied 35 healthy implant sites in five partially
edentulous patients who had been treated for severe periodontitis. None of
the implants in the study yielded Aa or Pg
 Pg was related to implant failure in of studies. In animal studies black-
pigmented anaerobic rods including Pgingivalis-like organisms were
associated with development of ligature-induced peri-implantitis Apparently
P gingivalis comprise major pathogen in infectious implant failure
42.

 A actinomycetemcomitans and P gingivalis can attach to barrier
membranes and P gingivalis can penetrate porous barrier membranes
one side to the other. Both organisms have been implicated in failing
regenerative periodontal therapy.
 Demolon et al. (1993) demonstrated an association between P
gingivalis on barrier membranes and clinical signs of gingival
inflammation.
42.

Effect of periodontal treatment on subgingival Pg
 Scaling and root planing may cause temporary decrease in levels of
subgingival P gingivalis but is not capable of eradicating the organism from
subgingival sites.
 Location of P gingivalis in inaccessible areas, such as furcations or the base
of periodontal pockets, is probably responsible for the failure of mechanical
therapy to remove the organism.
42.

 Similarly, non-resective periodontal surgery is not effective in removing P
gingivalis from adult periodontitis lesions. Surgery aimed at eliminating
periodontal pockets provides a more predictable suppression of P gingivalis,
especially when followed by diligent oral hygiene.
 Systemic antimicrobial therapy in combination with nonsurgical periodontal
scaling and root planing may not ensure subgingival eradication of P
gingivalis. Neither is topical antimicrobial therapy very useful in eliminating
P gingivalis from deep periodontal pocket . The most effective control of P
gingivalis seems to be achieved by employing a combination of resective
periodontal surgery, systemic antibiotic therapy and good oral hygiene.
42.

 Regeneration of periodontal connective attachment is contingent upon the
absence of P gingivalis and other periodontal pathogens in treated sites.
Zarkesh et al (1999) described a method of coating PTFE barrier membranes
with tetracycline. Compared with non-coated barrier membranes,
tetracycline-coated membranes permitted less P gingivalis colonization and
more clinical attachment gain.
42.

CAPSULE
 Considered an important virulence factor by many
 Some strains have either a thin layer or are devoid of it
 It is formed by a polysaccharide heteropolymer on the outer
membrane of the bacterial cell (Woo et al., 1979).
Holt SC, Kesavalu L, Walker S, Genco CA. Virulence factors of Porphyromonas gingivalis. Periodontol 2000 1999;28:168-238.

CHEMICAL COMPOSITION
 Differ bw strains
 Mansheim & Kasper (1977) -galactose, glucose and galactosamine
 Okuda et al (1987) - rhamnose, glucose, galactose, mannose and
methyl pentose

BIOLOGICAL FUNCTION
 Highly encapsulated strains exhibit decreased autoagglutination,
lower bouyant densities and were more hydrophilic than the less
encapsulated strains
 Increased encapsulation was correlated with increased resistance to
phagocytosis, serum resistance and decreased induction of PMN
chemiluminiscence
Pg capsules conjugated to bovine serum albumin and to the fimbriae
protein have been examined as vaccine candidates

OUTER MEMBRANE PROTEINS
Structure–
Complex multilayered structure
 The inner, cytoplasmic membrane
 A thin, peptidoglycan
 Assymetrical, outer membrane
-The outer membrane contains complex LPS, lipoproteins and
peripheral & transport proteins, Porin proteins, short fimbriae
-Made of atleast 20 proteins

OUTER MEMBRANE PROTEINS &
COAGGREGATION
 Pg may play an important role in the formation and maintenance of the
periodontal biofilm
 The benchmark report of Gibbons and Nygaard (1970) demonstrated that
there were specific interactions bw members of the oral microbiota and for
the first time revealed that bacteria do attach to both hard and soft surfaces as
well as to each other.
 In Pg, this interaction with selected G+ & G- bacteria was mediated by
specific outer membrane proteins in the whole cell-associated outer
membrane or in the outer membrane vesicles

LIPOPOLYSACCHARIDE
 The outer membrane of G- bacteria is assymetric, the outer leaflet of
which contains the LPS
 Very large molecule
 3 parts –
-The O antigen
-Core region
-Lipid A

CHEMICAL COMPOSITION
 Neutral sugars – Rhamnose, mannose, galactose and glucose
 Glucosamine and galactosamine represent the amino sugars
 Fatty acids
 Phosphorus

BIOLOGICAL PROPERTIES
Endotoxic activity is confined to lipid A, while the immunological activity
is contained within the O antigen.
 Ogawa et al (1994) demonstrated induction of IL-1 ra, IL-6, IL-8, IFN-
γ, GM-CSF secretion by cultured mononuclear cells
 Bramati et al (1989) and Sismey Durrant & Hopps (1991) –PGE2
 Tamura et al (1992) and Takada et al (1991) – IL-1β and IL-8
The preponderence of evidence indicates that LPS, especially the Lipid A,
is capable of stimulating the host inflammatory response directly via the
induction of host derived cytokine production

Bacterial fimbriae
 Thin, straight appendage originally referred to as Pili
 2 MAJOR CLASSES-
-Type-specific fimbriae
-F- or sex pili
 Approx 3 to 25nm in diameter & 3 to 25 µm long
 Vary in distribution & number from 10 to 100/cell

P GINGIVALIS FIMBRIAE
 Peritrichous
 Comprised of atleast 1000 protein subunits (fimbrillin subunits)
 Sex pili not present
Atleast 2 distinct fimbriae molecules –
 Major
 Minor

MAJOR FIMBRIAE
 Composed of fimbrillin
F/n –
1. Act in bacterial interactions with host tissue by mediating bacterial
adhesion & colonization in targeted sites
2. Are capable of binding specifically to and activating various host cells,
resulting in release of cytokines, as well as cell adhesion molecules
3. Have been shown to be necessary for bacterial invasion of host cells

 Lee et al (1991) first reported the variation of Fim A proteins and
divided a number of Pg strains into 4 types based on their N-terminal
aa sequences. Pg fim A genes have been further classified into 6
variants (type I to V , and Ib) on the basis of nucleotide sequence
 Nakagawa & coworkers (2000) found a majority of patients were
found to carry type II fim A organisms, followed by type IV. In
healthy it was type I (2001)

MINOR FIMBRIAE
 Was found in 1996 (Hamada et al) and was shown to be short fimbria-
like appendages in an fim A deficient mutant
 mfa 1 gene encodes for the subunit protein of the minor fimbriae (Mfa
1)
It is thought that production of both major & minor fimbriae is required
for the expression of pathogenic traits by Pg
- Amano et al (2004)

HEMAGGLUTININS
 Pg produces atleast 5 hemagglutinating molecules
 When expressed on the bacterial cell surface, hemagglutinins may promote
colonization by mediating binding of bacteria to receptors on human cells
 Since Pg utilizes heme for growth, binding of bacterial cells to erythrocytes
may also serve a nutritional function
 Evidence does implicate a role for fimbrillin

 Hemaggluttin activities expressed by Pg include those complexed with LPS
on the cell surface and a released form designated exohemagglutinin
 4 hemagglutinin genes from Pg strainms have been identified: hag A, hag B,
hag C & hag D
 Several studies have demonstrated that the hemagglutination events mediated
by Pg might be due to the combined effects of atleast 3 enzymes formed into
a large protein complex

Session 2
contents
 Invasiveness of P gingivalis
 Evidence supporting a role for P gingivalis as pathogen in
periodontal disease
 Vaccines for P gingivalis
 Conclusion
 References

PROTEINASES
 One of the potentially significant virulence characteristics of Pg is the large
number of hydrolytic, proteolytic and lytic enzymes that are produced by
essentially all known strains
Many of the enzymes are either-
 Exposed at the surface (in the outer membrane) of the bacterium where
they are able to come in contact with the host cells and tissues
 Within the periplasmic space capable of being transported to the cell
surface
 In the outer membrane vesicles, which are sloughed from the outer surface
during growth

Serine
Aspartate
Proteinases Thiol
Metalloproteinases
Secretory pattern is type IV
 Of these, the collagenases, aminopeptidases and trypsin like
proteinases are vertical to Pg Pathogenesis.
 Trypsin like proteinases – the Aeg and Lys proteinases all cysteine
proteinases, called gingipanis, the cytokine proteinases are thiol
proteinases and comprise the group of endopeptidases whose
members rely for their catalytic activity on the presence of the thiol
group of cysteine residue in the enzymes molecule

I. TRYPSIN-LIKE PROTEASE (THE
GINGIPAINS)
 These proteases occur in multiple forms that are found extra cellularly on the
bacterial cell surface and in some cases are associated with protein regions or
“domain” involved in adherence properties.
2 proteases-
 Arg gingipains (Rgp) encoded by rgpA & rgpB
 Lys gingipains (Kgp) encoded by kgp

FUNCTIONS OF GINGIPAINS
 Adherence and colonization:
 Gingipains in hemoglobin binding and heme acquisition
 Production of nutritious peptides
 Degradation of antibacterial peptides
 Exploiting complement
 Direct degradation of extracellular matrix proteins

II. AMINOPEPTIDASES
 The aminopeptidases have been found in several members of the oral
microbiota, many of which exhibit some arylaminopeptidase activity..
 Pg is probably the only periodontopathic microbiota that exhibits
strong dipeptidyl arylaminopeptidase activity
 Nakamura et al. (274), defined a “pure enzyme”, further studies of this
aminopeptidase revealed that it contained at least two additional
aminopeptidases, N-CBz-glycyl-arginyl peptidase (N-CBz-Gly-Gly-
Arg), which was both cell associated and extracellular, and an
extracellular peptidase, glycyl-prolyl peptidase (Gly-Pro peptidase).
Both aminopeptidases were thiol and serine activated proteinases,
respectively

III. Caseinases
 Hydrolyse the protein casein
 Exists in atleast 3 isoforms called Pase A, B & C
 B & C display a cleavage pattern similar to that of trypsin (ie
cleavage of carboxy side of arginine)
 Pase A has its preferred cleavage site as the carboxyl side of lysine
 Immunological characterization suggested that Pase-C is located on
the cell surface and monoclonal antibodies against the protein
inhibited the hemagglutinating activity, hydrolyzed BApNA and
strongly degraded native type I collagen

IV. COLLAGENASE
 Collagen has been found to comprise a significant percentage of
gingival connective tissues. The major portion of the gingival
connective tissue is the highly organized collagen fibers, which have
as their primary function the maintenance of tissue integrity and
function.
 It was almost 35 years ago that Shultz-Haudt & Schrep (1955)
demonstrated that a mixed culture of bacteria isolated from the
gingival cavity displayed collagenolytic activity. This activity was
demonstrated to be collagenase from Bacteroides melanogenicus.

 Data support the participation of bacterially derived collagenase;
however these data are equivocal.
 Collagenase is perhaps the most important of the P gingivulis
proteolytic enzymes, and if expressed in vivo, would be a major
destructive enzyme (virulence factor) associated with the soft tissue
destruction characteristic of human periodontitis.
 Several investigators have postulated that the P gingivulis
collagenase may participate with host-derived collagenase in the
destruction of gingival collagen

OTHER ENZYMES
 Alkaline phosphatase
 Superoxide dismutase
 Sulfatase, heparinase and chondroitinase

EVIDENCESUPPORTINGA ROLE FOR P. GINGIVALIS AS PATHOGENIN
PERIODONTALDISEASE
Criterion
 Association
 Elimination
 Host response
 Increased in periodontitis lesions, found
associated with crevicular epithelium
 Suppressed or eliminated in successful
therapy, found in recurrent lesions.
 Increased systemic and local antibody
levels in LAP

Criterion
 Animal studies
 Found to be important in experimental
mixed infections and in periodontitis in
the cynomolgus monkey
 Host tissues cell adherence and invasion,
collagenase, trypsin like enzyme
(gingipains), fibrinolysin, Phospholipase
A, phosphates, endotoxin (LPS), factors
that affect PMN function

INVASIVENESS OF P GINGIVALIS
 As part of its strategies for survival into the host, P. gingivalis is able
to invade cells and tissues (Yilmaz, 2008), thus avoiding the immune
surveillance.
 Porphyromonas gingivalis can actively invade gingival epithelial
cells, where it can maintain viability and replicate (Belton et al.,
1999; Tribble et al., 2006).
 This invasive property is dependent on its major fimbriae, which
bind to b1 integrin on the surface of host cells, an event that causes
rearrangements of the actin cytoskeleton to allow internalization
(Yilmaz et al., 2002, 2003)
42.

 Porphyromonas gingivalis can also invade macrophages, but within
these cells its replication is less active (Wang et al., 2007).
 once P. gingivalis has invaded intracellularly, there are no signs of
apoptosis or necrosis (Nakhjiri et al., 2001). It can then actively
secrete an ATP-hydrolysing enzyme, thus suppressing ATPdependent
apoptosis (Yilmaz et al., 2008) and allowing its survival in host cells.
 Subsequently, it can disseminate from cell to cell, through actin
cytoskeleton bridges without causing cell death, and spread while
avoiding immune surveillance (Yilmaz et al., 2006).
42.

 Once P. gingivalis is established in the cell, it affects cell-cycle
pathways and thus accelerates proliferation of gingival epithelial
cells, in a fimbriae-dependent fashion (Kuboniwa et al., 2008). This
could well constitute a mechanism of expansion of the periodontal
pocket epithelium, which is a histopathological feature of
periodontitis.
42.

Relevance of cell invasion
 Regulation of MMP production by GEC is disrupted and this will
interfere with extracellular matrix repair and reorganization
 Invasive Pg cells inhibit secretion of IL-8 by GEC
42.

Treatment of periodontal infections
 3 factors play a major role in the difficulties encountered in achieving
long-term control of periodontal infections without the need for
continuous maintenance-
a) The first is the numerous reservoirs of these taxa in the oral cavity
coupled with the potential for reacquiring the taxa from other persons in
the environment
b) The second is the stability conferred on communities of species while
living in biofilms
c) The third is the resiliency of the “climax community”
42.

Future directions in the treatment of P gingivalis
 Use of isolated adhesins as adhesins is one approach
 Expression & secretion on the oral comensal S. gordonii of Pg fimbrillin
domains
 Expression of Hag B in live avirulent Salmonella typhimurium induces
primary and recall antibody response in both serum and saliva and may
thus provide an effective means of delivering vaccines capable of
blocking colonization
42.

 As an alternative to vaccine delivery, blocking agents targeted to active
domains of adhesins or proteases
 Other potential targets include molecules involved in the regulation of
virulence gene expression, virulence factors upregulated invivo and
signaling molecules involved in biofilm expression. The availability of Pg
genome can be anticipated to make these approaches more viable
42.

Vaccines for P gingivalis
 Vaccination is a process that induces specific immune resistance to a
bacterial or viral infection.
 A common finding in patients with periodontitis is the presence of P.
gingivalisspecific antibodies in serum and gingival crevicular fluid.
 Immunization with several P. gingivalis-specific antigens has been
shown to enhance the immune response against P. gingivalis, as
demonstrated by the induction of specific antibodies and reduction of
P. gingivalis-induced alveolar bone loss in animal models.
Nitin K, Nitin D, and Swapna M. Periodontal vaccine: A dream or reality. J Indian Soc Periodontol. 2011; 15(2): 115–120.

 The production of antibodies generally indicates the activation of our
major host defense mechanism; these antibodies are insufficient to
clear P. gingivalis infection. Although complete protection through
immunization has not yet been achieved, new knowledge about
specific P. gingivalis antigens holds promising possibilities for the
future

 Active immunization
• Whole bacterial cells
• Sub unit Vaccines
• Synthetic peptides as antigens
 Passive immunization
• Murine monoclonal antibody
• Plantibodies
 Genetic immunization
• Plasmid vaccines
• Live, viral vector vaccines

 Bruce C. Ross & coworkers (2001) used a whole genome sequencing
approach to identify potential vaccine candidates. From a genomic sequence,
120 genes were selected The selected genes were cloned for expression in
Escherichia coli and screened with P. gingivalis antisera before purification
and testing in an animal model. Two of these recombinant proteins (PG32
and PG33) demonstrated significant protection in the animal model, while a
number were reactive with various antisera. This process allows the rapid
identification of vaccine candidates from genomic data.

 Frank A. Roberts & coworkers (2004) measured PGE2 levels in gingival
crevicular fluid of primates immunized with a P. gingivalis vaccine
(formalin-killed P. gingivalis) followed by ligature-induced
periodontitis.PGE2 levels were dramatically suppressed in immunized
animals versus controls. A significant correlation was also found between
PGE2 levels and decreased bone loss scores. This study presents the first
evidence of a potential mechanism involved in periodontitis vaccine-induced
suppression of bone loss in a nonhuman primate model and offers insight into
the role of PGE2 in periodontal destruction.

 Agenta’s Patent Technology (filed 12/6/2003) “Immunization against
periodontitis with the HA2 hemoglobin binding domain of
Porphyromonas gingivalis”
How do we know HA2 domain is important to P. gingivalis?
 HA2 domain acquires iron and porphyrin
 This pathogen has no biological machinery to make the porphyrin
metabolite
 The pathogen grows poorly if HA2 is deleted
Agenta has tested an HA2 domain vaccine in an animal periodontitis model

CONCLUSION
 It is evident that P. gingivalis has developed mechanisms to invade and
persist into the host, by absolutely adapting to its local niche. Its
paradoxically opposing (stimulatory vs. inhibiting) effects on innate
immune and inflammatory responses aim to subvert host defence
mechanisms, in order to facilitate its survival in the tissues
 Finally, as periodontitis is of polymicrobial nature, it is reasonable to
consider the role of different bacterial species within the context of
(subgingival) biofilm communities. Hence, P. gingivalis is likely to
function in concerted action with other species, to their mutual benefit.

REFERENCES
 Socransky SS, Haffajee AD. Dental biofilms: Difficult therapeutic targets.
Periodontol 2000. 2002; 28:12-55.
 Griffen AL, Becker MR, Lyons SR, Moeschberger ML, Leys EL.
Prevalence of porphyromonas gingivalis and periodontal health status. J Clin
Microbiol 1998;36:3239-42.
 Holt SC, Kesavalu L, Walker S, Genco CA. Virulence factors of
Porphyromonas gingivalis. Periodontol 2000 1999;28:168-238.
 Nitin K, Nitin D, and Swapna M. Periodontal vaccine: A dream or reality. J
Indian Soc Periodontol. 2011; 15(2): 115–120.

 How KH et al. Life below the gum line: Pathogenic mechanisms of Pg.
Microbiology and Molecular Biology Reviews, December 1998, 1244-
1263.
 Chen T, Nakayama K, Belliveau L & Duncan MJ. Porphyromonas
gingivalis gingipains and adhesion to epithelial cells. Infect Immun. 2001;
69: 3048–3056.
 Bao, K. et al. Role of Porphyromonas gingivalis gingipains in
multispecies biofilm formation. BMC Microbiol. 2014; 14: 258-260.
 Van Winkelhoff, A. J., & Boutaga, K. Transmission of periodontal
bacteria and models of infection. Journal of Clinical Periodontology.
2005; 32(s6), 16–27.

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