8. Microbiology of Endodontic Infections

Question 1

• Experiments that set the role of organisms in endo disease
• Took rats > conventional/healthy rats, and also rats that were germ-free (sterile environment, have a defined flora > not as extensive as that in the conv rat)
○ Exposed pulp in both rats
○ In healthy rats > pulp necrosis > apical perio lesions
○ In germ free rats > up to 100 days after exposure > pulp remained ____ (living, blood supply, actively alive)
§ Extended period > no disease in the germ free > tissue ____
• L: germ-free pulp
○ Dentin bridge, and the stained material is the pulp (below)
§ Stained: cells, pulpal contents, no real bacteria
• R: conventional rats
○ Extensive bacterial contamination > infection > abscess formation
• Provided proof that ____ are involved in order to see inflam and disease

Answer 1

vital
repair
bacteria

Question 2

• Group led by Moller
• Took monkey teeth > devitalized the pulps (what you do in a RC > drill through tooth and remove the pulp) > artificially infected w organisms or not infected
○ Group used a mixeture of bacteria
○ Where devitalized pulp was > organisms infecting > tissue pathology > disease was noted
○ In case where pulps were not ifnected > periradicular tissues remained ____
• Proof that ____ were importantt for at least th einitaiation and maybe the full development of disease

Answer 2

normal

organisms

Question 3

• Done by Sundqvist
• Took teeth w pulps that were necrotic but due to trauma
○ Pulps became necrotic, and he sampled the bacteria from the teeth (using end paper points to collect) > xferred to culture medium > bacteria that were dispersed by sonication > important bc they are in biofilms, so they ____ them up > isolate colonies following culturing (in anaerobic and aerobic conditions)
• Spread the samples on various selective/differential media > isolate single colonies
○ Koch’s postulates - identify microorgs assoc w dx > purify them out
• 90% of samples contained ____ bacteria
○ Found only in RC of teeth w apical perio, and the necrotic pulp tissue did not show evidence of dx
○ Some ended up being infected, and some not
§ Those that were infected > contained bacteria (mostly anaerobes)
• Showed that anaerobic bacteria were important in the dx

Answer 3

break

anaerobic

Question 4

Entry of Microorganisms

caries
damage as a result of ____
activity

iatrogenic restorative procedures
damage as a result of ____ treatment

traumatic occlusion
damage as a result of ____ forces on the tooth

Answer 4

bacterial
dental
physical

Question 5

Consequences of Microbial Entry

• Time elapsed between pulp exposure and infection of the entire canal is ____ but is usually a ____ process.
infection

• Colonization of the root canal system allows bacteria to come in contact with the periradicular tissues typically via ____ or root perforations.

Answer 5

unpredictable
slow
apical/lateral foramina

Question 6

Consequences of Microbial Entry

1.____ conditions in
different areas of the root canal

2. ____ of oxygen tension and nutrients (type and availability) is formed
3. ____ residing in different parts can also differ in diversity, density and accessibility to treatment procedures

Apical region
____ oxygen tension
Nutrients from the periradicular tissues: proteins and glycoproteins
Lower bacterial counts
Bacteria less accessible to treatment measures

Coronal region
____ oxygen tension
Nutrients from the oral cavity: carbohydrates
Higher bacterial counts
Microorganisms more accessible to treatment

• Root canal infection is a dynamic process with different bacterial species dominating at different stages.
• Shifts in the composition of the microbiota are largely due to changes in environmental conditions, particularly in regard to ____ tension and ____ availability.

Answer 6

ecologic
gradient
microbiota

lower
higher

oxygen
nutrient

Question 7

• Facultative type of flora
○ ____ species
§ Ferment carbohydrates
• Then necrosis starts > O2 levels drop > loss of blood cirulcation > ____ conditions
○ ____ species
§ Metabolize proteins moreso than carbohydrates
○ ____ bacteria
• Specific species become established
○ Concept known as geography-related pattern > composition of the flora is different among diff patients, and among diff sites/teeth
○ Depend on the ecology of the enviro
§ What’s available and what bac initiate the infection

Answer 7

saccharolytic
anaerobic
asaccharolytic
proteolytic

Question 8

• ____ is set up
• Food chain bt specific bacteria
• Streptomuyces/actino produce formic acid > metabolized by caphlobacter
○ Also produce acetic acid > liked by the pseudoramobacter
• Porp prevotalis > isobutytrate
○ Treponema like them
• Population turns over as the products become more abundant and more species become established
○ Contributes to growht of the bacteria in the infection and estab of a specific type of flora

Answer 8

IC food web

Question 9

Consequences of Microbial Entry

Necrotic Pulp Conditions That Favor Microbial Colonization

• Necrotic pulp provides bacteria with moist, warm, nutritious and anaerobic environment for colonization.
• Environment protected from host defenses because of lack of active ____ circulation in the necrotic pulp tissue.
• Root canal walls are non-shedding surfaces conducive to persistent colonization and formation of ____.
• Major ecologic factors include ____ tension, type and amount of available nutrients and bacterial interactions. (temperature, pH, and receptors for adhesins may also be important)
• About ____ bacterial species, out of about 600 to 700 in the oral cavity, are consistently selected out for growth and survival in necrotic pulp tissue.

Answer 9

blood
complex communities (biofilms)
oxygen
10 to 20

Question 10

Consequences of Microbial Entry

• ____ are the major type of microorganism implicated in the
pathogenesis of apical periodontitis.
• Root canal microbiota is dominated by bacterial morphotypes that include ____, rods, ____ and spirochetes.
• Involved bacteria are usually part of the ____ oral microbiota (“____ pathogens”).

Answer 10

bacteria
cocci
filaments
normal
opportunistic

Question 11

• Stages of biofilm formation
• Biofilms need a supporting substratum > enamel on the surface, or the salivary pellicle coated enamel
○ In RC infection > dentin or the outer surface of a tooth structure
• Conditioning film > allows bac to stick to spec proteins
○ Ind ____ colonizing bacteria come in > stick and form micro colonies > inc in size due to cell div, and produce EC polysacc’s > forms EPS matrix (ECM - grey)
○ Other bacteiral congregations or lightly adherent groups move in (secondary colonizers) > biofilm becomes more complex
○ Bacteria are dividing, growth is away from the surface of the biofilm (lower surface), until the mature biofilm becomes large enough that bac begin to slough off > ____ stage: impt aspect of biofilm formation > allows biofilms to spread
§ Bac get knocked off > free in the enviro (planktonic state) > find a free surface again > biofilms can grow more

Answer 11

primary

dispersal

Question 12

Complex microbial communities

• L: carious lesion
	○ Evidence of disease w radiolucency around the root
• Take the tooth > slice it longitudinally
	○ Clear area = root canal
	○ Pink = invading host cells (neutropihls)
	○ Purple = bacteria
		§ Deposited on the inner wall of the root
• Enlarged that
	○ Structure of the biofilm
		§ Reminiscent of C
			□ Looks same as plaque formation and maturation; a lot of rods, but it's more dense where it's \_\_\_\_ to the wall; as move out from the biofilm the bacteria become more dispersed
			□ Specific interaction bt bacteria, and they're multiplying to form microcolonies
	○ D - more cocci, not as filamentous
		§ See other debris (cellular, etc.) that's sticking to the surface of the biofilm
		§ Distance bt the two walls of the tooth > they're close together > biofilms can look quite different still

Answer 12

adherent

Question 13

Complex microbial communities

____ biofilms generally observed in apical segment of approximately 80% of the root canals of teeth with primary or post-treatment apical periodontitis

Morphology of biofilms differed consistently from individual to individual by ____, morphotypes and bacterial cells/extracellular matrix ratio.

____ underneath biofilms were often invaded by bacterial cells from the bottom of the biofilm community

Answer 13

intraradicular
thickness
dentinal tubules

Question 14

Complex Microbial
Communities (Biofilms) Criteria Used to Establish Causal Link Between Biofilms
and an Infectious Disease

1. Infecting bacteria ____ to or associated with a surface.
2. Direct examination of infected tissue shows bacteria forming ____ or microcolonies encased in an extracellular matrix.
3. Infection is generally ____ to a particular site. Although dissemination may occur, it is a ____ event.
4. Infection is difficult or impossible to eradicate with ____ in spite of the responsible microorganisms being susceptible to killing in the planktonic state.
5. Ineffective host ____ is evident as suggested by the location of bacterial colonies in areas of host tissue associated with host inflammatory cells (polymorphonuclear neutrophils (PMN) and macrophages).

(6. ____ or significant disruption of biofilm structure and ecology leads to remission of the disease process.)

Answer 14

adhered
clusters
confined
secondary
antibiotics
clearance
elimination

Question 15

Involvement of dentinal tubules

• Dentinal tubules have a ____ conformation and traverse the entire width of the dentin.
• Largest diameter (2.5 μm) is near the ____ and smallest diameter (0.9 μm) is near the ____ or cementum.
• Smallest tubule diameter is similar to the ____of most oral bacterial species (0.2 to 0.7 μm).
• When dentin is exposed the pulp is at risk of infection because of the ____ of normal dentin due to its tubular structure.

Answer 15

conical
pulp
enamel
cell diameter
permeability

Question 16

Involvement of Dentinal Tubules

Bacterial Invasion of Dentinal Tubules in a Vital Pulp

• ____ movement of dentinal fluid and tubular contents influence dentinal permeability and may delay intratubular invasion by bacteria.
• Functional or physiologic diameter of the tubules is only 5-10% of the anatomic diameter seen by microscopy because of the presence of tubular contents.
• The few bacteria that reach the pulp may not be significant because the vital pulp can eliminate such a transient infection and rapidly ____ or remove bacterial products due to the presence of host defense molecules, such as ____ and components of the ____ system.

Answer 16

outward
clear
antibodies
complement

Question 17

Involvement of Dentinal Tubules
Bacterial Invasion of Dentinal Tubules in a Non-vital Pulp

• Since most bacteria in the carious process are ____ they may invade dentin by repeated ____ which can “push” cells into the tubules.
• Bacterial cells may be forced into tubules by ____ pressures placed on dentin during mastication.
• Dividing cells can be observed within tubules indicating that bacteria can derive nutrients probably from degrading ____ processes, denatured collagen, bacterial cells that die during the course of infection and intracanal fluids that enter the tubules by capillary action.

Answer 17

nonmotile
cell division
hydrostatic
odontoblastic

Question 18

Involvement of dentinal tubules

• Bacteria aren't actively invading the tubules > they get in indirectly
• In this model (love and jenkinson)
	○ The dentin is lined by \_\_\_\_
		§ Has specific regions that act as receptors for the adhesins on bact
		§ Strepto gordondii > primary colonizer
			□ At end of tubule, receptor will find receptor and stick to the collagen
			□ Can grow in this area > start to divide and form chains > grow into the tubules not by moving as free cells, but as elongating as chains into the tubule
				® Many of oral streptococci grow in \_\_\_\_ (how to diff from staphylococci)
	○ Secondary colonizer comes along > has an \_\_\_\_ that recognizes a protein on the surface of the streptococci
		§ Stick to the primary colonizer > not independently moving into the tubule, but carried when the streptococci chain \_\_\_\_
		§ The streptococci are dragging the secondary colonizers in
	○ This is how tubules become infected

Answer 18

collagen
chains
adhesin
elongates

Question 19

Diversity of endodontic microbiota

• Represents the distribution of various bacteria/members of taxons that are involved in endo infections
	○ The difference > these analysis done from studies that where detection of bac were totally using molecular methods, totally using culturing methods, or using a combination of both methods
• Molecular methods - PCR to 16s rRNA seq, to pyrosequencing (DNA arrays, etc.)
• Culturing methods - automated systems (API), and serotyping to ID bacteria
• Can see diff kjinds of bacteria using \_\_\_\_ methods as opposed to \_\_\_\_ methods
	○ Some taxa (TM7, SR1) > contain bacteria that have never been cultured in the lab, but can now ID in clinical samples
	○ Allow to detect presence even though can't visibly culture them in the lab
• Some of the taxa > solely determined to be in taxa based on DNA/RNA sequence

Answer 19

molecular

culturing

Question 20

Diveristy of the endodontic microbiota

• Shows phylogenetic relationship
	○ Length of lines represent genetic distance
		§ The closer two phyla are, the more closely related they are genetically
		§ The length of the line indicates how far removed they are from earlier connection to another phyla
• Represent the most commonly found types of bacteria present in endo inf
	○ \_\_\_\_ > gram +
	○ \_\_\_\_ > gram -
	○ Bacteroidetes
		§ P gingivalis is in this phylum
• In terms of the species found in absecess, \_\_\_\_ and \_\_\_\_ make up the bulk of the species IDed in abscesses
	○ Depicted in the previous slide
	○ Another way of looking at the istributio of diff phyla in endo infections
	○ Pertains to asbcesses
		§ Three studies that were done w cultural, and three w molecular methods
			□ Bulk of bacteria are members of \_\_\_\_ and \_\_\_\_
				® Significant portion of bac we find in abscesses

Answer 20

firmocutes
proteobacteria
firmucutes
bacteriodetes
firmucutes
bacteriodetes

Question 21

• Representation of the phylogenetic tree of life
○ 3 domains
§ ____
§ ____
§ ____
□ Red ones are the ones he showed us two slides again
® Major phyla found in endo inf
□ Not all the phyla are found in endo infections
® Limitation/restriction

Answer 21

eukarya
archea
bacteria

Question 22

• Divide endo infections into diff types
		○ Intraradicular
			§ Primary
				□ Initial \_\_\_\_
			§ Secondary
				□ Primary inf is cleared up, and a new inf occurs in the same canal
					® May or may not be due to similar or diff bac found in the primary system
		○ Extraradicular
			§ \_\_\_\_
			§ \_\_\_\_

Answer 22

infection
dependent
independent

Question 23

Primary infections w/ sympatomatic (acute) apical periodontitis

• Members of flora found in primary infections
	○ Don't worry ab symptomatic
• Detection of bacteria in a sample
	○ \_\_\_\_
		§ 75% of samples contain this, not 75% of the bacteria in the inf are d denticola
		§ Presence of bac in a sample
	○ \_\_\_\_
		§ Assoc w periodontal dx
	○ \_\_\_\_ is low on the list on being detected in these samples
	○ Diverse group of bacteria; not so diverse in terms of genera, but diverse in species

Answer 23

treponema denticola
porphymonas gingivalis
enterococcus vicalis

Question 24

Primary infections w/ asymptomatic (chronic) apical periodontitis

• Shift in terms of major players
	○ T denticola has moved farther down
	○ \_\_\_\_ has moved up
	○ Enterococcus is still low on the list compared to the others
• Overall, generally you see the same bacteria

Answer 24

dialister

Question 25

Secondary and persistent endodontic infections (post-treatment disease)

• Distribution of bac in secondary and persisinta
•\_\_\_\_ is at the top of the list
	○ Pattern of distributino when compared to primary > a large diff in types of abcteria

Answer 25

enterococcus faecalis

Question 26

prevalence of E. faeclis in post-treatment disease

• Independent studies
	○ Molecular methods
		§ Detection of e faecalis in secondary/later infections
		§ \_\_\_\_
	○ Comparing to e faecalis detected by culture methods > as noted before > the molecular methods showed better detection of the e faecalis
		§ Why are the culturing methods now shown in higher frequenct as the molecular methods
			□ E faecalis is \_\_\_\_
				® Easy to grow in the laboratory, but the culturing methods show low detection when compared to the \_\_\_\_ methods
				® Hard to get the sample, the bacteria are protected in various places in the root > \_\_\_\_ methods may fall apart bc you're not getting the bacteria out 
• Main point: e faeclis has come into prominence in this type of infection

Answer 26

post-treatment
gram + cocci
molecular
culturing

Question 27

Secondary and Persistent Endodontic Infections (Post-treatment Disease)
Enterococcus faecalis

• Most frequently detected (90% of cases) species in root canal–treated teeth.
• Considered a transient species in the human oral cavity (source may be food).
• ____ teeth are about nine times more likely to harbor E. faecalis than cases of primary infections.
• Commonly recovered from teeth treated at multiple ____ or left open for ____.
• ____ penetrates dentinal tubules allowing it to escape the action of endodontic instruments and irrigants.
• Resistant to high ____ (protons internalized by proton pump acidifying the cytoplasm) and ____.
• Forms biofilms in root canals.
• Survives in environments with a scarcity of nutrients (____ state) and then flourish when the nutrient source is re-established.

Answer 27

root canal-treated
visit
drainage
deeply
pH
calcium hydoxide

viable but noncultivable state

Question 28

Secondary and Persistent Endodontic Infections (Post-treatment Disease)
Causation of Disease by E. faecalis is Unproven

• Despite being easily cultivated, E. faecalis is not ____ in all studies evaluating the microbiota of root canal–treated teeth with post-treatment disease.
• Even when present, E. faecalis is rarely one of the most ____ species in pretreatment cases.
• E. faecalis is not more prevalent in root canal–treated teeth with ____ when compared with treated teeth with no lesions.
• Other bacteria found in endodontically treated teeth with apical periodontitis include ____ and some fastidious anaerobic species.
• Uncultivated phylotypes represent up to 55% of the taxa detected in treated canals and can also be in high proportions, corresponding to about half of the 16S rRNA gene sequences retrieved in clone libraries.

Answer 28

detected
dominant
lesions
streptococci

Question 29

Dependent Extraradicular Infections (acute apical abscess)

• Acute apical abscess is dependent on the ____ infection.
• Characterized by ____ inflammation in the periradicular tissues in response to a massive egress of virulent bacteria from the root canal.
• Bacteria adhere to the apical ____ root surface in the form of extraradicular ____ structures.

Answer 29

intraradicular
purulent
external
biofilm

Question 30

Acute Apical Abscesses Dependent on the Intraradicular Infection

• Break down the detection of various samples
	○ \_\_\_\_ at the top
• Extraradicular inf that are dependent on the intraradical inf, the flora looks similar to those in the primary infections
	○ Organisms coming form the primary intraradicular inf that cause the extraradicular infection

Answer 30

t denticola

Question 31

diversity of the endodontic microbiota

microbiota of extraradicular infections (acute apical abscess)

• Independent extraradicular infections no longer maintained by the
intraradicular infection and can persist even after successful eradication of the latter.
• Main bacterial species implicated are ____ species and ____ in a pathologic entity named apical (or periapical, periradicular) actinomycosis.
• Cohesive actinomycotic colonies that may be collectively resistant to ____ are formed within the body of the inflammatory lesion

Answer 31

actinomyces

propionibacterium propionicum

Question 32

Antimicrobial Intervention

• Amoxicillin is the current antibiotic of choice worldwide (____ or
____ recommended if patient is allergic to penicillin)
• ____ antibiotic (bacteriolytic, moderate-spectrum)
• More readily absorbed when administered ____
• Susceptible genera include ____, Actinomyces, ____, Enterococcus, ____, Helicobacter, and Moraxella
• Used for standard ____ (sustained serum levels)
• Included on World Health Organization’s list of essential medicines

Answer 32

clindamycin
erythromycin
b-lactam
orally
streptococcus
bacillus subtilis
haemophilus
prophylaxis

Question 33

Other Microorganisms in Endodontic Infections
Fungi (yeast)

• Gain access as contamination during ____ therapy (secondary infection) or overgrow after inefficient intracanal antimicrobial procedures that cause an imbalance in the ____ endodontic microbiota.
• Ability to colonize and invade dentin and is resistant to ____.
• Detected in 21% of samples from primary intraradicular infections.
• Detected in up to 18% of the cases from root canal–treated teeth.

Answer 33

endodontic
primary
calcium hydroxide

Question 34

Other Microorganisms in Endodontic Infections
Methanobrevibacter oralis-like Archaea

• ____-producing member of the Archaea.
• Detected in 25% of root canals of teeth with chronic apical periodontitis.
• Methanogenic archaea have been detected in samples of ____ plaque associated with periodontal disease.
• However, no member of the Archaea domain to date has been described as a human ____.

Answer 34

methane
subgingival
pathogen

Question 35

Other Microorganisms in Endodontic Infections

Human immunodeficiency virus (HIV)
(detected in ____ pulps of HIV-seropositive patients)

Herpesviruses
(identified in both non-____ and ____ vital pulps)

Human cytomegalovirus (HCMV) and Epstein-Barr virus (EBV) (detected in ____ lesions)

• Direct result of virus infection and replication.
• Result of virally induced ____ of local host defenses which might give rise to overgrowth of pathogenic bacteria in the very apical part of the root canal.
• Bacterial challenge emanating from the canals may cause an influx of virus- infected cells into the periradicular tissues.
• Reactivation of HCMV or EBV, by ____ injury induced by bacteria, may evoke impairment of host ____ response in the periradicular microenvironment.

Answer 35

vital
inflamed
inflamed
apical periodontitis

impairment
tissue
immune

Question 36

Summary of Endodontic Microbiology

• Microorganisms (predominantly bacteria) cause almost ____ pathoses of the pulp and periradicular tissues. A small percentage of these infections may contain fungi, Archaea- like microorganisms or viruses.
• Endodontic infections are usually caused by colonization and invasion of plupal tissue by ____ members of the normal microbiota.
• Endodontic infections contain polymicrobial ____.
• Molecular methods have demonstrated that each patient’s infection contains bacterial species somewhat different in ____ and type than those in infections of other individuals (geography-related patterns).
• Necrotic pulpal tissue is a reservoir of infection that the host immune system is not able to resolve because of the lack of vascular circulation.
• ____ is, by far, the most frequently detected species in secondary or persistent infections. However, the relationship is by ____ rather than causal.
• ____ and ____ species are the most prevalent bacteria in extraradicular infections.
• Knowledge of the microorganisms associated with endodontic disease is necessary to develop a basic understanding of the disease process and a sound rationale for effective management of these patients.
• Antibiotics of choice for treatment of endodontics infections are ____ and ____ or ____ when patient is allergic to penicillin.

Answer 36

all
opportunistic
biofilms
prevalence
e faecalis
actinomyces
propionibacterium
amox
clinda
erythro