Transcript root-canal treatment

Microbiology of Root Canal
Infections
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Please answer these questions:
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1.What classes of microorganisms are involved with root canal infections?
How do microorganisms get into the root canal?
2.How can we prevent root canal infections?
3.What is the importance of mechanical instrumentation in root canal
treatment?
4.Describe the microbial shifts in the root canal as the periapical lesion
develops.
5.What species of bacteria are commonly found in primary root canal
infections
6.What are the common bacteria found in root canal treated teeth with
persistent p.a. lesions?
7. Describe an experiment to show that bacteria is the main cause of root
canal treatment.
8. Describe an experiment to show that the collection of tissue fluids is not
responsible for the development of p.a. radiolucencies.
9. What is the distribution of bacteria in the root canal?
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Classes of microorganisms found in the
root canal
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Bacteria – prokaryotes, no membrane bound
organelles
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Fungi – eukaryote
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no nucleus, cell wall – peptidoglycan,
cell wall - chitin
Viruses - exist as a protein coat or capsid,
sometimes enclosed within a membrane, either
DNA or RNA which codes for the virus
elements, depends on host cell for replication
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Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Bacteria isolated from root canals may
themselves be infected….
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Stevens et al.,Oral Microbiol Immunol. 2009;24(4):278-84.
Bacteriophages induced from lysogenic root
So, if E.
faecalis were
to be cultured from
a root canal –
canal
isolates
of Enterococcus
faecalis.
was the periapical lesion due to Enterococcal infection?
 Lysogeny was demonstrated in 4 of the 10 E.
faecalis strains.
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 the
condition of a host bacterium that has incorporated a
phage into its own genetic material; "when a phage
infects a bacterium it can either destroy its host or be
incorporated in the host genome in a state of lysogeny”
http://www.thefreedictionary.com/lysogeny
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Cox CF et al., J Prosthet Dent. 1987 Jan;57(1):1-8.
How
do microorganisms enter the root
Biocompatibility of surface-sealed dental materials against
canal?
exposed pulps.
If you want a bacteria tight seal – use ZnOE –at least 2mm thick.
Carious exposure most common cause,
must realise when
treating such a tooth
leakage is from all
possible sources.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Bacteria can invade pulp through
exposed dentine…
However bacteria inward movement into pulp space
often impeded or blocked by:
1.Outflow of dentinal fluid
2.Presence of odontoblastic process
3.Presence of caries crystals – formed by presence of supersaturated
solution of Calcium Phosphate in the presence of acids.
4.Macromolecules in fluids carried into tubules
Arch Oral Biol 42 :835; 1997
Even if pulp exposed for 2 weeks <2 mm into pulp (J Endod 8:391)
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
How can we prevent root canal
infection?
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Keep the protective coverings of dentine on the tooth !
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Prevent Decay – Oral hygiene, flouride, CCP-ACP
Reduce gum disease – Oral hygiene
Reduce enamel wear – Good occlusion (ortho), Bruxers give night
guard, ensure proper reduction of crowns so that opaque does not show
through porcelain in PFMs
Reduce trauma to dental tissue – sports, CL IIdivII,
Keep the pulp alive !
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Use ZOE as a base below decay to kill bacteria
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Dental pulp can mount an immune response
Does not affect composites in a bad way see He et al. ,J Dentistry 2009
When there is caries detect and treat early – regular XR’s, careful reading
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Dental Pulp Can Mount Immune
Response – Both Innate and Adaptive
Hahn et al., J Endod 2007;33:773–781
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Pulp Immune Response
Adapted from Hahn et al., J Endod 2007;33:773–781
As acid diffuses ahead of bacteria
Macrophages
blocked by
dentine –
cannot get at
bacteria –
overactive
secretion –
damage pulp
– pulpitis.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Evidence that bacteria is the main
cause of pulp disease
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Historical believe
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– pulp irritation from materials caused pulpitis
− periapical irritation of tissue fluids cause periapical lesion
(Hollow tube theory)
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Proven incorrect by Torneck CD Oral Surg Oral Med Oral Pathol. 1966
Mar;21(3):379-87
Experimental Evidence – bacteria is the cause
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gnotobiotic rats with pulp exposure no pulp necrosis,
material thought to be irritating to pulp
had dentine bridges when placed against pulp (AR)
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Updated 2010 Jan
Kakehashi, S et al.. 1965. Oral Surg Oral Med Oral Pathol 20, 340–349
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Evidence that bacteria is the main
cause of pulp disease
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Teeth pulps extirpated with sterile instruments in sterile field.
Some were sealed using ZnOE to prevent bacteria leakage.
others left open to mouth.
Teeth could be sealed and remain sterile (for up to 6 months)
did not develop infections.
Teeth with infected pulp tissue showed inflammatory reactions
clinically and in histologically.
Teeth with infection showed clinical signs in 12/52 teeth and
47/52 on radiograph.
Anaerobic strains increased at six months, some initial samples
could not be cultured in the final culture.
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Updated 2010 Jan
Moller AJR et al., Scand J Dent Res 1981:89:475-84
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Evidence that bacteria is the main
cause of pulp disease
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Bacteria are regularly cultured from root canals with
periapical lesions and pulp necrosis.
Using culture techniques, of the 500 taxa of bacteria
commonly found in the mouth only about 20-30
genera are regularly isolated from root canals.
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Fusobacterium nucleatum,Streptococcus spp, Porphyromonas spp,
Prevotella intermedia, Peptostreptoccus spp, Actinomyces spp,
Eubacterium spp.
Using molecular techniques e.g. 16S ribosomal RNA
possible to identify and infer phylogeny of all
organisms on earth !
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
How are bacteria cultured from root
canals?
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Tooth isolated. Prophy with pumice. Wash water. H2O2 applied on tooth.
Wash sterile water. Tincture Iodine applied to tooth( 5 mins). Washed
sterile water. Dry with sterile gauze. Tooth opened with high speed drill
using sterile water spray. Just before entry into tooth, culture taken –
(negative control). Enter into tooth. Sterile paper points used to pick up
Have
cultured
all bacteria on earth?
liquid we
inside
root canal.
Transfer to reducing transport medium immediately. Incubate 1) in
anaerobic conditions for 7 days(enriched with CO2 and H2), if no growth
till 14 days; 2) in aerobic conditions for at least 4 days
Colour and Morphology study of isolated colony, Biochemical studies ( e.g.
blood agar etc.)
Subculture in various selection media.
Microscopy to look for movement (dark field for spirocheates) etc
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Colonization of the pulp
Ingress of bacteria
 Inflammatory response generates exudate rich
in plasma proteins – favours proteolytic
bacteria capable of evading host response
 Prevotella spp.
 Porphyromonas spp
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Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Colonization of the pulp
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When pulp is completely necrotic
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Range of bacteria invades the space (no more defence)
Initially predominant facultative anaerobes
Gradient of oxygen tension and nutrient supply develops
from coronal to apical
Coronal zone: Predominantly facultative anaerobes
Apical zone: Predominantly obligate anaerobes
Eventually obligate anaerobes dominate.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
How are bacteria isolated using PCR?
Use reagent to wash and remove nucleic
material from paper points
 Go through automated PCR cycler –
denaturing, annealing, extension; analyse DNA
sequence ( gel electrophoresis)
 Analyse base sequence online.
 Identification of closest match as the
bacterium
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Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Koch’s Postulate
The microorganism must occur in every case of the
disease in question and under circumstances that can
account for the pathological changes and clinical
course of disease.
Root
Canal
infections
do
not
fit
Koch’s
 The microorganism must be able to grow in pure
culture
postulate
! It is usually a polymicrobial
 infection.
After isolation from the diseased host and growth in
pure culture, the microorganism must be able to
induce a similar disease in experimental animals
 The microorganism should be isolated again from the
experimentally inoculated host.
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Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Shortcomings of Koch’s Postulate
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Emphasis is on bacteria causing disease neglecting
host response
It places emphasis on culturable bacteria and neglects
that many spp cannot be yet cultured and some exist
in VBNC states
Imply that all strains of the bacteria are equally
virulent
Presupposes that only a single spp causes disease
Requirement of reinoculation of the bacteria in an
animal to cause similar disease – some bacteria do not
cause disease in animals
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Microbial Shifts in the Root Canal
Fusobacterium spp is also a link between primary colonizers and
late colonizers in dental plaque. In the root canal, they help to
reduce oxygen and allow anaerobic bacteria to thrive.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Microbial Shifts in the Root Canal
Microbial Ecol Health Dis 1989;2:1-10.
Nutrition is the main engine of change of bacteria flora in root canals .
It starts as a mixed infection but overtime the relative component of
anaerobic infection increases as the REDOX potential becomes more
anaerobic.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Microbial Shifts in Root Canal
Infections
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Experiment , monkey teeth, open to oral environment
for 6-7 days after pulp amputation, tooth sealed and
opened after 7days, 90 days ,180 days and 1060 days.
Looked at ratio obligate anaerobes : facultative
anaerobes . Initial ratio between 1.4 and 1.7
After 90 days 3.9; after 180 days 6.5 after 3 years
>11.3 ( increase in strain number and cell number)
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Updated 2010 Jan
Fabricius et al., Scand J Dent Res 1982:90:134-144
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Microbial Shifts in Root Canal
Infections
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Initial gram+ve rods and cocci and gram-ve rods;
shifted to obligate anaerobes gram +ve and gram –
ve rods; shift most pronounced in the apical region,
though few facultative anaerobes still present.
Hence, in the apical part – fastidious, slow growing
obligate anaerobes outnumber more rapidly growing
facultatively anaerobic bacteria
Enterococci slight reduction in number of strains over
3 year duration
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Updated 2010 Jan
Fabricius et al., Scand J Dent Res 1982:90:134-144
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Microbial Shifts in Root Canal
Infections
 Why
does the bacteria type shift?
 Oxygen
sensitivity of species
 Different available nutrients in coronal vs
apical part of canal
 Interaction of bacteriocins
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Microbial Shifts in the Root Canal
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Sterile pulp extirpation, sealed in sterile or left open to be
infected by oral bacreria
Sealed sterile necrotic pulp no pa leesion after 6-7 months
Also when combinations of bacterial strains originally isolated
from an infected root canal were inoculated into further canals,
dominance of anaerobic bacteria was established.
Some species found in initial samplings not found in final
samplings
Interestingly, the original proportion of bacterial strains was
reproduced in the canals despite the fact that they were
inoculated in equal quantities into the canals.
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Fabricius L et al., Influence of combinations of oral bacteria on periapical tissues of monkeys. Scand J
Dent Res 1981;89:475-484.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Bacteria Shifts - Oxygen and oxygen products are likely
to play an important role as ecologic determinants
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That selective and interactive mechanisms operate in a root
canal is further supported by the finding that a Prevotella
oralis strain inoculated separately was unable to survive on its
own in the root canal.
On the other hand the P. oralis strain survived and dominated
This
is but onewhen
selection
nutrient.
somebacteria.
situations,
the
the established
inoculated
withInother
These
by
product of
oneshown
bacterium
is the
nutrientmilieu
of another.
experiments
have
that the
endodontic
is a
selective habitat that supports the development of specific
proportions of the anaerobic microflora.
It is likely that bacteria adapted to the specific conditions that
exist in different parts of the canal form aggregates.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Combinations of certain bacteria may
make abscesses persist !
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Abscess - the presence of P. intermedia or P. endodontalis in
the combinations resulted in a failure of abscess resolution
with a gradually increasing accumulation of leukocytes.
The results suggested that purulent inflammation in the apical
region in certain cases may be induced by specific
combinations of bacteria in the root canal and that the presence
of P. intermedia or P. endodontalis in such combinations is
essential. However, these bacteria need the support of
additional microorganisms to achieve pathogenicity.
Important to realise that bacteria communicate ! They can also cooperate !G et al., Capacity of anaerobic bacteria from necrotic dental pulps to
Sundqvist
induce purulent infections. Infect Immun 1979;25:685-93.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Combinations of certain bacteria may
make abscesses persist !
These results were later corroborated by van
Winkelhoff et a1.who found that all abscesses
of endodontic origin harbored one or more
species of the genera Prevotella and
Porphyromonas.
Prevotella intermedia was found in 63%,
Porphyromonas endodontalis in 53%, and
Porphyromonas gingivalis in 12% of the
examined abscesses.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Bacteria isolated from Primary infections
Sundqvist G. Oral Surg Oral M ed Oral Pathol. 1994 Oct;78(4):522-30.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
What is the distribution of bacteria
within root canals?
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Bacteria found in the lumen of the root canal and also within the tubules of dentine.
At the beginning of root canal infection, bacteria are distributed nearer the coronal
end of the root canal and mostly about 100-150mm from the lumen. (Also
dependent on bacteria species E. faecalis to 400 mm)
Bacteria and their toxic products can get out of the lumen into the periapical tissues
and cause inflammation and infection.
With progression of disease, bacteria penetration advances in all directions and
have been reported at the canal leading into the foramen.
Response by body brings a concentration of macrophages to the periapical region to
prevent spread of infection – this accumulation of macrophages contribute to bone
resorption and causes the periapical radioluncency. When there is a sinus
discharging related to an infected tooth, bacteria is likely to be widely distributed
within the root canal.
Each mm2 of dentine has about 40,000 tubular openings, except in the apical region,
there are about 8,000 per mm2
Crit Rev Oral Biol Med 13(2):171-183 (2002)
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Bacteria in Dentinal Tubules – Brown
and Brenn stain
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
What are bacteria of deep caries?
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Gram +ve rods (facultative and anaerobes)
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Gram +ve Cocci (facultative and anaerobes)
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Lactobacillus
Actinomyces
Peptococcus
Streptopeptococcus
Gram−ve rods (anaerobic)
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Prevotella
Porphyromonas
Fusobacterium
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
How do Gram+ve and Gram –ve
Bacteria Differ?
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Bacteria causes p.a. lesion
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The purpose of this study was to evaluate 50 human periapical
lesions for bacteria and epithelium in a case study in dental
practice.
Bacteria were found in all teeth, colonizing necrotic tissue in
the main canal, dentinal tubules, or apical ramifications, and in
the body of the periapical lesion in 18 abscesses or cysts.
Twenty-one lesions were epithelialized (14 abscesses, 20
granulomas, and 16 cysts were distinguished).
Granulomas were most common, and most epithelialized
lesions were cysts.
Bacteria were only detected periapically in abscesses or cysts.
Inflamed tissue was present in the apical root canal in one
third of cases.
Ricucci D et al., Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2006;101:239-49
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Wound healing of apical tissues after root canal therapy:
a long-term clinical, radiographic, and histopathologic
observation study
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77 root canal treated teeth, which were deemed to be
unrestorable, with no evidence of periapical bone lesion
evaluation of the radiographs, 51 cases that 3 independent evaluators
assessed as having normal periapical conditions were selected.
In the majority of the cases, complete healing was observed, with no signs
of acute or chronic inflammatory processes in the remaining apical tissue or
periodontal tissue fragments.
Some cases showed moderate inflammation in the root canal tissue.
Narrowing of the apical root canal by cementum was a common finding in
most cases, but total closure was not observed.
Debris intermixed with necrotic tissue and sealer particles was a common
finding in the pulp stump.
Bacteria were present in the coronal portion of the root in almost all cases,
but in only 1 case could bacteria be demonstrated in the coronal and apical
portions of the root.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
What are Cultured bacteria from apical
aspect of infected teeth
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Bacteria in Apical 5mm of Infected Root Canals
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Baumgartner and Falkler 1991 JOE 17:380-383
Coronal caries with periapical lesions
10 most common species from 10 teeth
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Actinomyces(6/10), Lactobacillus(7/10), Black-pigmented
Bacteroides species(5/10), non-pigmented Bacteroides
species(6/10),Peptostreptococcus(6/10), Veillonella(5/10),
Enterococcus faecalis(4/10), Fusobacterium
nucleatum(3/10), Streptococcus mutans(4/10).
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
What Bacteria are commonly found in
Root canals after root canal treatment
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Chávez De Paz et al.Int Endod J. 2003 Jul;36(7):500-8.
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Following removal of the interappointment temporary
cement, the canal was flushed with sampling fluid
(VMG I, Möller 1966) and instrumented to remove
Ca(OH)2 if present. Then and in cases where IKI had
been used for interappointment medication (Table 1),
VMGI was added and the canal walls were scaled.
The suspension was absorbed into charcoal
impregnated paper points (Möller 1966), which were
transferred to VMGA III. This sample was designated
Sample 1
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Bacteria in Root canals after root canal
treatment
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Chávez De Paz et al.Int Endod J. 2003 Jul;36(7):500-8.
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A total of 248 strains were isolated from 107 teeth giving
bacterial growth. Gram-positives predominated (85%).
Lactobacillus spp. (22%), non-mutans streptococci (18%), and
Enterococcus spp. (12%) were the most common isolates.
Gram-negative anaerobes were relatively sporadic. Large
radiographic bone lesions, persistent pain and use of intracanal
calcium hydroxide dressing correlated with bacterial presence
(P < 0.05).
CONCLUSIONS: Once established, non-mutans streptococci,
enterococci and lactobacilli appear to survive commonly
following root-canal treatment of teeth with clinical and
radiographical signs of apical periodontitis.
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Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Results
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Chávez De Paz et al.Int Endod J. 2003 Jul;36(7):500-8
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200 cases analysed, 107 presented with bacterial growth in the
root-canal sample designated as Sample 1
Non-mutans Streptococci and Enterococci were more
commonly recovered from teeth with diagnosis of necrotic
pulps (n = 19 and n = 10, respectively), their presence was
significantly more common in the retreatment cases
Gram-negative anaerobic rods (Prevotella, Fusobacterium,
lactose positive Enterobacteria and Porphyromonas), were
mainly present in teeth originally diagnosed with necrotic pulp
and were infrequent in re-treatment cases
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Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Results
Chávez De Paz et al.Int Endod J. 2003 Jul;36(7):500-8
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In the present study, Gram-positive bacteria were
recovered from root canals of teeth receiving rootcanal treatment for either symptomatic or nonsymptomatic apical periodontitis.
In several cases the organisms remained or new ones
appeared in subsequent samples, despite chemomechanical treatment and antimicrobial intracanal
dressings, suggesting that conditions for their survival
and growth prevailed in these root canals. Byström &
Sundqvist (1981), Gomes et al. (1996) and Peters
et al. (2002) have reported similar findings.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Results
Chávez De Paz et al.Int Endod J. 2003 Jul;36(7):500-8
Radiographic analyses demonstrated 6 teeth
with no lesions, five of them belonging to the
NG (no growth) group.
 Large bone lesions were the most common
(64%), and correlated significantly with
presence of bacteria (P = 0.001).
 Occurrence of small lesions was positively
correlated with no bacterial growth
(P = 0.0001).
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Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Definition of Large and Small Lesion
< 2 = small; >2 = large
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Comments
Root canal disinfection is not perfect ( not this
paper alone, many other papers)
 Just because we stir some chlorox and even if
we apply CH in the root canals does not mean
that they will be sterilized !
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Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Bacteria in root canal of refractory
lesions
SoNon-mutans
did these studies:
Streptococci and Enterococci were more
commonly recovered from teeth with diagnosis of necrotic
Sundqvist et al 1998 Oral Surg ;85(1):86-93. Scandinavian study, Umea
pulps
(n = Sweden
19 and n- of=54
10,teeth
respectively),
their presence was
University,
most had E faecalis
significantly more common in the retreatment cases
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Molander et al 1998 Int Endod J;31;1-7 Goteborg University, Sweden Enterococci were the most frequently isolated genera
The same bacteria is found commonly
among
failed
cases
in
many
Peciuliene et al 2000 J Endod.; 26(10):593-5. - in a Lithuanian population
(Vilnius, Lithuania)geographical
– Enterococcus the most
often isolated species
regions.
Hancock et al 2001 Oral Surg. ;91(5):579-86. - in a North American
population 30% of positive cultures were E faecalis
In Singapore ?
Pinheiro et al 2003 Int Endod J. Jan;36(1):1-11. Microorganisms from canals
of root-filled teeth with periapical lesions. (South American) –
Enterococcus was the most frequently isolated species
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
E faecalis – survives when buried by
GP in root canals
Sedgley et al. Int Endod J. 2005 Oct;38(10):735-42
The root canals in groups 1 and 2 were
inoculated with 10 6 bacteria, incubated for 48
h at 37 degrees C then filled with gutta-percha
and zinc-oxide eugenol sealer.
 E faecalis maintained viability for 12-months
ex vivo. The clinical implications are that
viable E. faecalis entombed at the time of root
filling could provide a long-term nidus for
subsequent infection.
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Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
What do they look like?
cocci
spirochaete
rods
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
What bacteria cause root canal infections?
Peptostreptococcus
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Mouth 500 species bacteria
Found in root canals about 150
Individual canals yield about 5-7 species
Most common :
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Peptostreptococcus – anaerobic, Gram +, non-sporulating;
small cells, spherical, short chains, pairs or individually;
the only genus among anaerobic gram-positive cocci
encountered in clinical infections;
part of the normal florae of human mucocutaneous
surfaces, including the mouth, intestinal tract, vagina,
urethra, and skin
http://en.wikipedia.org/wiki/Peptostreptococcus
Updated 2010 Jan
Ann Intern Med. Aug 1980;93(2):244-8.; Clin Infect Dis. Jun 1993;16 Suppl 4:S248-55.
J Clin Microbiol. Jun 1988;26(6):1181-8. ;Mayo Clin Proc. May 1974;49(5):300-8.
Scand J Infect Dis. 1994;26(5):503-10.
Microbiology of Root Canal Infections
Dr. Sum CheePeng
What bacteria cause root canal infections?
Prevotella
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

Prevotella spp –( Bacteroides melaninogenicus) anaerobic. Gramnegative, non-motile, rod-shaped, singular cells, colonize by binding
other bacteria and epithelial cells; breakdown of protein and
carbohydrate foods.
Prevotella cells' natural antibiotic resistant genes, which prevent
extermination
Prevotella species cause infections such as abscesses, bacteraemia,
wound infection, bite infections, urinary tract and vagina infections,
and periodontitis.
Specific infections caused by Prevotella include periodontitis.
Symptoms of Prevotella infections can include pain, swelling, and in
some cases a "wet" canal.
Antibiotics – metronidazole, amoxicillin/clavunalate.
Updated 2010 Jan
focus.hms.harvard.edu/.../research_briefs.shtml
Microbiology of Root Canal Infections
Dr. Sum CheePeng
What bacteria cause root canal infections?
Porphyromonas





Gram-negative, non-sporulating, anaerobic, rod-shaped bacteria
that produce porphyrin pigments (dark brown/black pigments).
closely related to Gram-positive bacteria than other Gramnegative bacteria.
The black pigmentation of P. gingivalis is from the accumulation
of hemin used as an iron source for bacterial growth.
(people with higher metal intakes, such as iron, at higher risk of
getting gingivitis and periodontitis.)
cell surface adhesion molecules interact with other bacteria,
epithelial cells, and extracellular matrix proteins, assist the
bacteria in living in their human host.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
What bacteria cause root canal infections?
Fusobacterium
Gram-negative non-sporulating ;exceptional ability to adhere
with both Gram-ve and Gram+ve plaque microorganisms in
biofilms , has made it a highly invasive pathogen. Primarily
given attention for its periodontal implications, strains of
Fusobacterium have been identified as pathogen to many parts
of the body.
Courtesy University of Adelaide
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Candida albicans
Gram negative rods
http://www.asm.org/Division/c/fungi.htm
•Candida albicans is a common
commensal yeast that colonizes
the gastrointestinal or genital
tract of 15 – 30 % of healthy
humans.
•Serious infections such as lifethreatening invasive infections
in immunocompromised hosts
(chemotherapy,
transplantation).
•Invasive candidiasis mainly
occur in hospitalized patients
http://calbicans.mlst.net/misc/info.asp
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
What bacteria cause root canal infections?
Eubacterium
Eubacterium (Includes E coli) currently includes a
heterogeneous group of gram-positive, nonsporulating, anaerobic bacilli,
slow growing, fastidious and generally unreactive
in biochemical tests - cultivation and identification
difficult and the taxonomy of the group remains
indifferent.
commtechlab.msu.edu/sites/dlc-me/zoo/zah0700.html
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
What bacteria cause root canal infections?
Actinomyces
Gram-positive, can be either anaerobic or facultatively anaerobic .
Actinomyces species do not form endospores;
while individual bacteria are rod-shaped, morphologically Actinomyces
colonies form fungus-like branched networks of hyphae
http://en.wikipedia.org/wiki/Actinomyces
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
What then are the objectives of Root
Canal Disinfection
Remove all substratum nutrient sources
 Remove as much of the infection as possible
 Leave a surface that is easy for bacteria to be
flushed out
 Leave behind a substratum that is nonconducive for bacteria survival
 Attempt to leave behind a bactericidal surface

Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Sum et al.,
What we leave behind_
Aust Endo J 2005 31_3 pg 1-7
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
How deep do bacteria penetrate into
tubules?
Infection of tubules is random
 Tubule infection occurs as a late event in
What will happen when you remove 1500m of dentine?
development
of pulpal infection
 Depth of bacteria invasion may depend on
substrate available (collagen helps), but
usually is observed in the third of dentine
nearer the lumen of the root canal (300m to
1500m Love and Jenkinson Crit Rev Oral Biol Med 2002;13:171–83;George et al., J

Endod 2005;31:867–72.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Bacteria infection in dentinal tubules
Love and Jenkinson Crit Rev Oral Biol Med. 2002;13(2):171-83

Recent evidence suggests that streptococci
may recognize components present within
dentinal tubules, such as collagen type I, which
stimulate bacterial adhesion and intra-tubular
growth. Specific interactions of other oral
bacteria with invading streptococci may then
facilitate the invasion of dentin by select
bacterial groupings.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Can we kill E. faecalis in tubules with
CH?
Cwikla et al. J Endod. 2005 Jan;31(1):50-2



Dentin powder samples collected with ISO 018 burs showed a
statistically significant reduction in E. faecalis for all three
experimental groups in comparison with untreated control
specimens.
You basically have to use another chemical – not CH.
I use CHX insignificant
Hibiscrub. HBibiscrub
alsowere
contains
Statistically
differences
alsomany
found between
–tensioactivegroups.
to allow fluid
penetration.
(Intomost effective
thesurfactants
three experimental
Metapex
was the
tubules ?
dentinal
tubule disinfectant, followed by CH+IKI and then
Cationic in nature – high rate of charge changes kills bacteria!
CH.
I also use ultrasound.
Similar results were observed at greater dentin tubule depths
(ISO 021 burs) with the exception that intracanal treatment
with CH resulted in significantly higher numbers of E. faecalis
in comparison with untreated control specimens.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
E. faecalis
About 30-50% failed cases harbour this
bacterium (PCR >70%)
 It can survive very high pH =pH12.



CH treatment ?
This contrasts with primary lesions

Mixed infection 4-6 strains, anaerobes.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
E. faecalis
What concentrations would kill this bacterium in a clinically
meaningful time?
 3% NaOCl - “biofilm or planktonic suspension phenotype. A
cell-dense pellet "presentation“ prepared from planktonic
phenotype was also tested. …The difference in gradients of
This is just to give you an idea. There are many factors complicating
bacterial killing among the biofilm, planktonic suspension or
the killing action of disinfectants; amongst which anatomical factors,
pellet presentation
waspresence
significant
(p < 0.05)
formation
of biofilms and
of dentine
debrisand
are dependent
major
upon the test agent except in the case of NaOCl and calcium
factors.
where
nothe
difference
could
be detected.
Ishydroxide
there a need
to soak
canal longer
in NaOCl?
 NaOCl was the most effective agent and achieved 100% kills
for all presentations of E. faecalis after a 2 min contact time.”


Updated 2010 Jan
J Endod. 2005 Jan;31(1):30-6.
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Bubble implosion into dentinal tubule when High Intensity
Focused Ultrasound was used
J Endod. 2009 Jul;35(7):1028-33
Research conducted at NUS.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Adhesion of E. faecalis to chemically
treated dentine
Hence,
sequence
is important.
What gives
rise to collagen
in root canals ?
Ending with NaOCl means that collagen
EDTA
exposed by EDTA
would be dissolved.
How long do we need to dissolve
How do
we get rid by
of collagen
?
collagen
exposed
EDTA?
NaOCl
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Sequence is important !
After Marraige
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Bacteria adhesion

Very complicated matter

Some variables:
DLVO Theory
Medium of bacteria culture
Ionic Strength
Valence of ions
Salt Concentration
Surface conformation of
proteins and adhesins
Presence of other bacteria
Phase of growth
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Can we irrigate the canals without
instrumentation?
Need to widen canal so that surface tension
will not impede ingress of fluids
 Instrumentation of
root canal helps to remove
from George and Kishen 2008
tissue and to break up biofilms.
 Biofilms can be calcified. Bacteria in such
biofilms may not get killed even if the biofilm
is contacted by disinfectants.
 Whatever we do, remnant bacteria will exist.

Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Can we prevent reinfection?



Other than remnant bacteria, ingress after treatment
may be from tubules exposed to oral cavity or from
leaking restorations.
Insurance ?
Seal canal orifice with IRM. Will not affect
composite bonding.


(J Dent. 2009 Dec 2.A suitable base material for composite resin
restorations: Zinc oxide eugenol. He LH, Purton DG, Swain MV)(
Supports idea of IRM seal over canal orifice, some problems
applying this to Black’s Class II cavities)
Eugenol at 10-4 mols/l kills bacteria
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Intracanal bacteria and periapical
lesions
Bacteria usually limited within root canals
 Few cases of reported bacteria in the apical
tissues are with infected cysts and abscesses
 Periapical lesions are formed by chemotaxis of
macrophages which are attracted to the region
by bacteria and bacteria products diffusing to
the area
 Bacteria products include LPS, enzymes –
proteases and other cytotoxic metabolites

Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Nair,PNR.
Crit. Rev. Oral Biol. Med.
2004;15:348-381
NG= neutrophillic
granules
EP=epithelium
GR=granuloma
A granuloma is defence against spread of bacteria
into the body
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
BA=bacteria
Summary



Prevent entry – OH, perio,caries control
RDI
Scrub tooth with disinfectant before working





Use of cotton swab is important ( fibres are like bacteria peritrichous
fibrils
and fimbriae
overcomes
repulsive
barrier) it !
Don’t
let it– get
in. If
it isenergy
in KILL
Use disinfectants judiciously to kill bacteria in canals and
tubules
Use of ultrasound may be helpful.
Do not leave collagen behind !
Seal the tooth to prevent bacteria re-entry (IRM at orifice,
appropriate long term restoration, surface tubules sealing?)
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Epilogue


Giving systemic antibiotics cannot kill bacteria in root canals. Besides:
1.”reduction of commensal bacteria after antibiotic treatment may interfere
with nutrient availability and impair beneficial stimulation of GI immune
response. This impairment may be associated with continued colonization
by opportunistic microbes and inflammatory immune response that could
lead to malabsorption and malnutrition.”

Nutrition and the Immune System Nutrition 1998; 14:573-579 ( Cornell University
Immunology Research Laboratory)

2.Accumulating evidence showing that bacteria surface carbohydrates
(symbiotic bacteria) drive the modulation of the intestinal immune system,
resulting in mature, balanced immune responses and oral tolerance.

Curr Opin Gastroenterol 24:720-724(Channing Lab, Harvard Med Sch)
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Epilogue

The gut of newborn infant is sterile. In full-term infants a diet of breast
milk induces the development of a flora rich in Bifidobacterium spp. Other
obligate anaerobes, such as Clostridium spp. and Bacteroides spp., are
more rarely isolated and also enterobacteria and enterococci are relatively
few. During the corresponding period, formula-fed babies are often
colonized by other anaerobes in addition to bifidobacteria and by
facultatively anaerobic bacteria; the development of a "bifidus flora" is
unusual.
Acta Paediatr Suppl. 2003 Sep;91(441):48-55

bacterial DNA of streptococci, staphylococci, lactic acid bacteria and
bifidobacteria in the samples of human milk, which confirms that breast
milk can be an important source of bacteria and bacterial DNA to the infant
gut.

Lett Appl Microbiol. 2009 Feb 18.
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Epilogue




After birth, each human being may be colonized by over 100 trillion
bacteria, representing over 500 bacterial species. The ratio of bacterial
to human cells in a normal adult may exceed 10:1.
The nature and the species of microflora acquired in the first few
months of life is determined by many factors including, external
environmental microflora, introduction of cow's milk, use of antibiotics
and immunomodulatory agents, and use of breastfeeding.
Recent investigations have shown that the nature of mucosal microflora
acquired in early infancy determines the outcome of mucosal
inflammation and the subsequent development of mucosal disease,
autoimmunity and allergic disorders later in life. It appears that altered
mucosal microflora in early childhood alters signaling reactions which
determine T cell differentiation and/or the induction of tolerance.
Nestle Nutr Workshop Ser Pediatr Program. 2008;61:145-81
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Epilogue
After birth, each human being may be colonized by over 100 trillion
bacteria, representing over 500 bacterial species. The ratio of bacterial
to human cells in a normal adult may exceed 10:1.
 The Bacteria
nature and
of microflora
acquired
in the
first few
thatthe
arespecies
commensals
are vital to
the health
of humans.
months of life is determined by many factors including, external
They aremicroflora,
just opportunistic
pathogens,
doingmilk,
harmuse
where
environmental
introduction
of cow's
of antibiotics
they should
normally
and immunomodulatory
agents,
and usenot
ofbelong.
breastfeeding.
It is investigations
of utmost importance
that rubber
damnature
isolation
be used and
 Recent
have shown
that the
of mucosal
microflora
the working
fieldinfancy
be scrubbed
free ofthe
bacteria
before
acquired
in early
determines
outcome
of starting
mucosalwork.
inflammation
and the subsequent
development
mucosal
disease,
As
health care professionals
– we should
not abuseof
use
of antibiotics
and
autoimmunity
and
allergic
disorders laterappropriate
in life. It appears
altered
encourage
vaginal
birth
and breastfeeding(=
bacteriathat
seeding).
mucosal microflora in early childhood alters signaling reactions which
determine T cell differentiation and/or the induction of tolerance.


Nestle Nutr Workshop Ser Pediatr Program. 2008;61:145-81
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng
Who are we feeding?

‘Acquisition of enteral access and provision of a
sufficient volume of enteral nutrients early in the
hospital course of a critically ill patient afford an
opportunity to improve the outcome of that patient
through the progression of his or her disease process.
Failure to use the enteral route of feeding not only
squanders this opportunity, but may, in addition,
promote a proinflammatory state, which exacerbates
disease severity and worsens morbidity’

J Clin Gastroenterol. 2002 Sep;35(3):209-13
Updated 2010 Jan
Microbiology of Root Canal Infections
Dr. Sum CheePeng