ggregatibacter actinomycetemcomitans (previously Actinobacillus actinomycetemcomitans) is a Gram-negative, facultative nonmotile, rod-shaped oral commensal often found in association with localized aggressive periodontitis, a severe infection of the periodontium, although it is also associated with nonoral infections. Its role in periodontitis was first discovered by Danish-born periodontist Jørgen Slots, a professor of dentistry and microbiology at the University of Southern California School of Dentistry. 'Bacterium actinomycetem comitans' was described by Klinger (1912) as coccobacillary bacteria isolated together with Actinomyces from actinomycotic lesions of man. It was reclassified as Actinobacillus actinomycetemcomitans by Topley & Wilson (1929) and as Haemophilus actinomycetemcomitans by Potts et al. (1985). The species has attracted attention because of its association with localized aggressive periodontitis. is explained here by Dr Harshavardhan Patwal

1. Actinobacillus
Actinomycetemcomitans
Dr Harshavardhan G Patwal

2. Contents
Introduction
Historical perspective
Description
Taxonomy of AA
Serotypes
Factors that influence colonization of AA
Virulence factors
Invasiveness of AA
Transmission of AA

3. AA in dental plaque
AA in non oral infections
Host response to AA invasion
Methods of detection of AA
Treatment modalities of eliminate of AA
summary

4. Introduction
Destructive periodontal disease represents
several disease entities with different clinical
presentation and pathogenic mechanisms
All cases of periodontal disease are infectious
in origin and are initiated and perpetuated by
pathogenic microorganisms in dental plaque.
Dental plaque provides a nutritious ecological
niche to several organisms.
AA – association with AP

5. Historical perspective
 Klinger et al in 1912 – cervicofacial
actinomycotic lesion
 Initially designated , bacterium
actinomycetemcomitans.
 Lieske in 1921 – bacterium comitans
 Topley et al in 1929 – finally designated as
AA.

6. MORPHOLOGICAL
CHARACTERISTICS OF AA
 measures app 0.4 ± 0.1 x 1± 0.4 μm in size.
 Microscopically, cultures appear
predominantly bacillary with few coccal forms
only.
 capnophillic requiring an atmosphere
containing 5-10% co2.
 It is a facultative anaerobe that grows under
anaerobic conditions.
 It is non sporulating and non hemolytic in
nature.

7. Taxonomy of AA
Belongs to pasteurellaceae
Pasteurella Haemophilus Actinobacillus
GENUS:
ACTINOBACILLUS
A.Capsulatese
A.Seminis
A.Sui
A.Ureae
Aa
A.Muris

8. Biochemical properties of AA
Slots – 135 biochemical characters in 6
reference strains and 130 strains of AA
freshly isolated from the oral cavity.
Small, non motile gram negative capnophillic
rods.
They all decomposed hydrogen peroxide, were
oxidase negative, benzidine positive reduced
nitrate, produced strong alkaline and acid
phosphatases and fermented fructose,
glucose and mannose.

9. Tanner et al – in their study found that none of
the isolates fermented glucose, galactose,
lactose, sucrose or trehalose.
Serotypes of AA:
Pulverer and ko – Using tube agglutination
assays detected 24 groups with AA and 1-6
agglutinating Ag on each strain.
King and Tatum – 3 serotypes based on a heat
stable component, were distinguished among
non oral AA.
Taichman et al – 4 serotypes based on surface
Antigen and proteinaceous leukotoxin.

10. Zambon et al – 3 serotypes a, b, & c.
Asikainen et al – relationship betn serotype b
and LJP & c and periodontal health in adults.
Gunsolley et al – Aa serotypes b, c higher in
periodontitis sites
Serotype is that Aa infected individuals with
periodontal disease exhibiting elevated Ab to
multiple serotypes are consistently colonised
by serotype b.
Asikainen et al using selective culture
techniques with immuno diffusion assays – the
parents harbored the same serotype as the
child in 22 out of 23 Aa positive families
suggesting an intra familial transmission of AA.

11. Virulence factors of AA
1. Factors that promote colonization and
persistence in the oral cavity
 Adhesins
 Invasins
 Bacteriocins
 Antibiotic resistance
2. Factors that interfere with host defense:
Leukotoxin
Chemotactic inhibitors
Immunosuppressive proteins


12. 3. Factors that destroy host tissues:
 Cytotoxin
 Collagenase
 Bone resorption agents
 Stimulation of inflammatory mediators
4. Factors that inhibit host repair of tissues:
 Fibroblast proliferation inhibitors
 Inhibitors of bone formation

13. FIMBRIAE
AA fimbriae may occur in peritrichous arrays,
may be more than 2µm in length and 5µm in
diameter and often occurs in bundles.
Fimbriated strains have been found to attach
to hydroxyapatite and saliva coated tooth
surfaces better than non fimbriated strains.
It associated protein, an attachment factor of
AA analyzed genetically, the gene for this
protein is strongly expressed in fimbriated
strains.

14. vesicles
These structures which are LPS in nature,
originate from and continuous with the outer
membrane.
The surface of highly leukotoxic AA strians an
abundance of extracellular membranous
vesicles.
AA vesicles also exhibit adhesive properties.

15. Extracellular amorphous material
Is an amorphous material that frequently
embeds adjacent cells in a matrix.
The material is a protein, most likely a
glycoprotein and has been shown to
demonstrate bone resorption activity.
The amorphous material can easily be from
AA cultures, and can be used to mediate
adhesion of AA to host cells and to
streptococcus parasanguis, a phenomenon
called conveyed adhesion.

16. Surface associated material
Wilon et al showed that a saline extract from
AA, purified by gel filtration and ionic
exchange chromotography caused a dose
dependant stimulation of bone resorption
over concentration range 1µm to 10µm / ml.
Which blocks cell cycle progression in G2 by
a unique mechanism and has a potent
proinflammatory cytokine induction
mechanism with extremely potent induction of
IL6 & IL8.

17. LPS
It contains app 30% carbohydrates, 30% lipid
A, 10-12% hexosamine phosphate phenol
water extracts of LPS from AA are active in
releasing ca from mouse calvaria – bone
resorption.
It can stimulate IL1β & TNFα production from
human peripheral monocytes in blood.
It is cytotoxic to fibroblasts and can result in
reduced incorporation of sulfates into
proteoglycans.

18. Bacteriocins
Actinobacillin is the bacteriocin of AA that is
active agent against S.Sanguis and
A.Viscosus.
It is associated with both the bacterial cell
surface and extracellular vesicles.
Increase the permeability of the cell
membrane which leads to leakage of DNA,
RNA and macromolecules essential for
growth.

19. Bone resorption
mediators
Mediators produced by AA:
LPS
Proteolysis sensitive factor
Microvesicles
Heat shock protein/ Gro EL

20. Collagenase
AA produces collagenase which can degrade
human collagen.
Cytotoxins:
It affects fibroblast viability.
It is a 50 kda protein, inhibit DNA synthesis
and also proliferation of the gingival fibroblast.
Gapstein – 8kda protein is ass with the SAM
of AA which is lethal cytotoxin that can inhibit
cells in the G2 phase of the cell cycle.

21. Fc binding proteins
Fc binding proteins are such competing and
blocking proteins.
The binding of Fc molecules to AA could be
inhibited by biotinylated Fc molecules.
Tolo and hegland demonstrated that such
surface molecules could bind to the Fc
portion of IgG inhibiting the ability of
opsonizing antibodies to bind to PMN and
reduces phagocytosis by 90%.

22. Invasiveness of AA
Goadby – proposed that bacteria can invade
oral tissues.
Listgarten in 1965 – electron microscopy, to
observe gingival tissue from cases of ANUG.
The presence of spirocheates and fusiforms
in the necrotic zone overlying the ulcerated
lesions.
Noiri et al – observed bacteria in gingival
epithelial cells from pts with destructive
periodontitis.

23. AA initially adheres to the epithelial cell
transferrin receptor to start invasion although
binding to integrins may constitute a
secondary entry pathway.
Attachment induces effacement of microvilli
and the bacteria enter through ruffled
apertures in the cell membrane.
Invasion of most strains requires remodeling
of the actin filaments that translocate from the
periphery of the cell to a focus surrounding
the bacteria.
Internal bacteria are initially confined within a
host derived membrane vacuole but this
membrane is soon broken down and the
bacteria resides in the cytoplasm.

24. AA cells induce the formation of surface
membrane protrusion through which the
organism can migrate and enter the
temporarily adjoined adjacent cell.
The invasion of AA into endothelial cells
occurs by a distinct mechanism.
Phosphorycholine – bearing antigens engage
the receptor for PAF to initiate invasion.

25. Implication of AA
invasion
Destruction of periodontal tissues
Sheltered nutritious ecological niche in the
host tissue.
Ideal atmosphere for planktonic organisms
from the biofilm.
AA can down regulate epithelial expression of
NFKβ, endothelial expression of il8.
Can be translocated from cell to cell and to
distant organs like the heart causing extraoral
infection.

26. Person to person transmission of
AA and its implications
Transmission of a pathogen can be of 2 types:
1. Vertical transmission
2. Horizontal transmission
Vertical transmission of AA:
Vertical transmission denotes transmission of
AA to the child from the parent.
Finnish study – Asikainen et al
American study – Preus and zambon
Brazilian study – Tonoco et al

27. Horizontal transmission
May occur betn siblings or between spouses.
Di rienzo et al – 89%
Tinoco et al – 7%
Asikainen et al – evidenced by transmission
between married couples.

28. Clinical significance of transmission of
AA from person to person:
Bacterium positive individuals can easily
transmit disease to their spouses.
Children of affected patients may also harbor
pathogens and hence contract periodontal
disease.
Aerosol mediated transmission may occur.
The role of saliva as a transport vehicle has
been supported by the fact that AA can be
cultured from saliva.

29. AA in dental plaque
The 1996 world workshop in periodontics has
reached a consensus in declaring: AA,
P.Gingivalis, B.Forsythus as portative
periodontal pathogens. There are between a
suspected pathogen and destructive
periodontal disease is provided by AA.
Robert koch’s postulates from 1884 provide
guide lines to determine disease causation by
microorganisms.

30. Sockransky’s postulates - 1979
Association : elevated in lesions of LJP,
prepubertal or adolescent periodontal disease,
lower in health, gingivitis and edentulous
subjects.
Elevated in adult periodontitis lesions, elevated in
active lesions of juvenile periodontitis.
Detected in prospective studies detected in apical
areas of the pocket or in tissue samples from ljp
patients.
Elimination : elimination or suppression resulted in
successful therapy. Recurrent lesions harbor AA.

31. Host response : elevated antibody in serum
or saliva of ljp and patients. Elevated local
antibody in ljp sites.
Virulence factors: leukotoxin, collagenase,
endotoxin, epitheliotoxin, fibroblast inhibitory
factory cytolethal distending toxin
immunosuppressive factors.
Animal studies : induced disease in
gnotobiotic rats. Subcutaneous abscesses in
the murine model.

32. The natural habitat of AA
The oral cavity serves a habitat for a great
variety of bacteria belonging to the human
indigenous microbiota.
AA is usually found in periodontal pockets,
although it can also be recovered from
supragingival plaque, oral mucosal surfaces and
dorsum of the tongue, saliva and pharynx.
From supragingival plaque : Muller et al have
isolated AA
From dorsum of the tongue, saliva : Asikainen et al
From pharynx : Van Steenberg et al

33. Can AA colonise tooth surface?
Kilian et al – supragingival tooth surfaces in
early plaque development
Kilian and Thelaide – could colonize clean
human tooth surfaces
Alalussua S et al – 10% of periodontaly
healthy children with primary dentition.

36. AA in non oral infections
Oral events
Dental plaque
periodontal pathogens
Normal flora Tissue invasionPeriodontal
disease
Systemic exposure to
bacteria and Lps
Hsp60 like molecules
cytokine production
Systemic
events
Damaged
heart valve
Vasculature and
coagulation complications
Infective
endocarditis
Atheroscelorosis,
coronary heart disease
CV events

37. Cardiovascular infections caused
by AA
Endocardidits represents the most frequent non
oral AA infections.
Mortality rate – 33-35%, complication rate
including congestive cardiac feature of 43% and
rate of major mobilization of 50%.
Stauffer and Goldman in 1972 – prosthetic valve
endocariditis due to AA.
Ziljstra – isolated AA from a pt suffering from
pericarditis.
Von winkehoff – AA septicemia from
colonization of a cardiac pacemaker.

38. Intracranial infections due to aa
Originate from dental and periodontal
infections.
Martin et al & Fabiani et al – brain abscess
due to AA.
Renton et al – 19 yr old male with AA cerebral
abscess.

39. Thoracic infections due to aa
Lung abscesses
Septic episodes of dental plaque
Septic episodes of odontogenic infections
Skin infections due to AA:
Apotheloz and Regamy – purelent skin
infections due to aa.
Dommann et Al and Donzis and Rappazo –
wrist abscesses in pts yielding AA on
culturing.

40. Cell mediated responses to aa
The immune response is of 2 kinds
1. Cell mediated
2. Humoral immunity
The cells of the immune system:
Neutrophils
Monocytes
T & B lymphocytes
NK cells

41. The role of neutrophils in tackling
periodontal pathogens
Processes Events / molecules
Enter the blood Bone marrow stromal cell regulation
of hemopoiesis
Transendothelial
migration
L.selectin, p.selectin, chemokine,
proteases
chemotaxis Fmlp, c5a, il8, g protein coupled
receptors, adhesins
Transepitheial
migration
Β2 integrin and il8 interaction
phagocytosis Opsonins, ic3b, cd14, cr3, cr4
killing Annexins, nadph oxidase, mp,
lysozyme

42. AA evade pmn leukocyte
defenses
Impairment of chemotaxis:
Increase epithelial and endothelial expression
of IL8 genes and hence jeopardizes the
chemotactic gradients.
The RTX leukotoxin:
Which binds to LFA-1 on the neutrophil by
interaction with 1 domain of the cd11a
subunit.
Lfa1 is expressed in both myeloid an lympoid
cells and binding to leukotoxin causes death
of the cells.

43. Immuno suppression:
AA produces immuno suppressive factors that
impede the functioning of the immune
system.
Humoral responses to AA:
Individuals colonized by AA generally respond
with humoral response by production of Ig.
There is a rise in local as well as systemic
levels of Ig in response to bacterial antigens
such as fimbriae and adhesins.
Ebersole et al – association betn increased
levels of and increased frequency of
occurrence of antibody to AA in LJP.

44. Later study – showed increased levels of IgG
to AA serotype b in 90% of LJP and 25% of
adult periodontitis.
North American population – association
between antibody to AA and LJP.
A study from brazil – elevated serotype b
antibodies in down’s syndrome patients
suffering from AP.

45. ANTIGENS FINDINGS
LPS EOP
High mol wt LPS
ass antigen
EOP
leokotoxin EOP
100 kda CP
64 kda EOP
fimbriae Genetic control of
humoral response
OMA (40 & 70 kda) Elevated in
generalized EOP
AB response to AG of AA

46. Methods of detection of aa in
periodontal disease
Diagnostic methods to detect the putative
pathogen in deep periodontal pockets which
would help plan appropriate therapy in
especially aggressive forms of periodontal
disease.
The different diagnostic methods :
Culture methods
Immunodianostic methods
Nucleic acid probe
PCR

47. Culture methods
AA is a capnophilic non sporulating, non
motile rod that does not grown on Mac
Conkey’s agar like other members of the
pasteurellaceae family.
Upon primary isolation in bld agar, AA
forms small colonies of .5-1mm
diameter.
The translucent colonies with irregular
edges appear smooth, circular and
convex.

48. The colony morphology of a fresh isolate is
distinctive with the internal star shaped, or
cross cigar morphology from embedding in
agar.
Colonies of fresh isolates are generally
rough.
Repeated subculture yields two types of
colonial variants.
Earliest media developed for AA – MGB
which is selective medium that utilises
trypticase soy broth with malachite green and
bacitracin added as inhibitory agent.

49. Later – composed of trypticse soy agar and
serum with bactracin and vancomycin.
Another medium called the a medium is
essentially tsbv supplemented with
spiramycin, fusidic acid and carbenicillin for
AA.
Tissue culture media have also been used for
AA as the pathogen in fastidious.

53. Immunodiagnostic methods to
detect aa
IDM employ Ab that recognize specific bacterial
Ag to detect that target microorganisms.
S.Bonta et al – detection of AA with a detection
limit of 500 cells/ml with a sensitivity of 82-100%
and specificity of 92%.
Slots et al – evalusite test is a commercially
developed Ab based ELISA for the detection of
AA.
The sample wells are coated first with Ab against
Ag specific for the bacterial specific targeted.
Ag-Ab reactions are then detected by enzyme
linked Ag specific antibodies to the sample wells.
Detection limits are 105 AA cells/sample.

54. Nucleic acid probe
DNA probes entail segments of single stranded
nucleic and labelled with an enzyme or
radioisotope that can locate and bind to their
complementary nucleic acid sequence with low
cross reactivity.
Savdt et al – dixogenin labeled whole genenomic
probes.
Sockransky et al – DNA – DNA checker board
hybridization for the detection of oral bacteria
sample dna from plaque has been released and
immobilized on a nitocellulose membrane by a
process.
The membrane bound DNA is allowed to hybridize
with either digoxigenin labeled whole genomic
DNA or 16s & RNA based olegonucleotide probes.

55. PCR
It involves a reiterate amplification of a region
of DNA flanked by a selective primer pair
specific for the target species.
identifying even 3-5 cells several genes
specific for AA.
AA leukotoxin gene and demonstrated a
sensitivity of detecting even upto 15 cells /μl.
Multiplex PCR can detect several organism
and different primers.

56. Treatment modalities to
eliminate AA
Adjunctive antimicrobial therapy has
been suggested to help eliminating AA.
Genco and collagens – 4 time / day
dose of tetracycline for 14 days every 8th
wkly associated with scaling and RP
was effective in eliminating AA in AP
case.

57. Principles of intervention in AA
elimination
Sloto & Rosing - Scaling and root planning alone
are not effective.
Christerrson - Flap alone is not effective
Adjunctive antibiotic therapy
(Metro with Amox)
1. Mechanical therapy – initial phase – SRP –
microbe samples
2. Reevaluation with assessment of surgical
techniques
3. Systemic antimicrobial therapy started after
surgery or immediately after another round of
mechanotherapy
4. Repeat microbial sampling after 3 months

58. Effectiveness of pdl
treatment on AA
THERAPY STUDY N
O
DISEASE LEVEL OF
AA
PRE
RX
POST
RX
Non surgical Fleming
Renvert
Niemenen
preber
17
15
16
13
46
Periodontitis
Periodontitis
Periodontitis
100
%
63%
50%
24%
40%
92%
9.4%
50%
6%
7%

59. THERA
PY
STUDY NO DISEASE LEVEL OF AA
PRE
RX
POST
RX
Pdl
surgery
Danser
Neiminen
Mombelli
Gunsolleg
kim
15
15
7
23
9
Periodontiits
EOP
LJP
.5%
46.7%
44%
22.5%
66.8%
2%
53.3%
29%
12.9%
0%

60. THERAP
Y
STUDY NO DISEASE LEVEL OF AA
PRE RX PO
ST
RX
Adjuncti
ve
antibiotic
Niminen
Renvert
chrester
sson
25
12
6
Periodon
titis
LJP
71.4%
29%
100%
14.
3%
5%
50
%