microbiology of pulp & pathogenesis Flashcards
2 Evidence for role of microorganism in root canal infections
Kakehashi et al 1965
- purposely expose pulp in
- germ free rats -> exposed pulp healed
- conventional rats -> pulp necrosed and apical periodontitis
Experimental study in monkeys (moller et al 1981)
- 26 teeth were kept sterile by sealing (non infected)
- 52 teeth were infected with oral microflora leading to necrotic tissues & inflammation & marked inflammation in apical tissue
what are the difference in species in primary infection and persistent infection
primary infection
- dominated by gram -ve obligate anaerobes with some facultative anaerobes
- initially: facultative bacteria
- after a few days/ weeks: o2 in canal is depleted due to necrosis and bacterial consumption and hence anaerobic envt conducive for growth of obligate anaerobes
persistent/ recurrent infection
- mainly gram +ve facultative anaerobes because these are the ones that are resistant to intracanal Antimicrboials and can endure nutrient deprivation
what is the primary species in recurrent infections
Enterococcus faecalis
- gram +ve facultative anaerobe
- more common in persistent than primary infections
- virulence factors are adhesins, gelatinase, hemolysin
properties of resistance
- survive in high alkaline environment (up to pH 11.5) because it has proton pump so can pump out H+ to neutralise OH- ions and decrease pH
- resistant to most intracanal medicament like CaOH
- survive in nutrient deprived condition
- survive without synergistic support
- forms biofilm, invade dentinal tubules and bind to dentine
what are the species of bacteria in necrotic pulps
1) Streptococci (Gram +ve)
intermedius, mitis, mutans, oralis, sanguinis
2) Obligate anaerobes
- prevotella intermedia
- P gingivalis
- Trep denticola
- Fusobacterium
3) facultative anaerobes
- Lactobacillus
- Actinomyces
what are the ways in which biofilm confers resistance
1) recalcitrance to antimicrobial compounds
- slow penetration: ineffective AB penetration due to ECM -> low conc when reach bacteria
- altered microenvironment in which the bacteria are slow growing which means less cell wall synthesis and reduced target of amoxicillin
2) persistance despite sustained host defences
- some phenotypes gain resistance
- transfer of resistance genes
- presence of efflux pumps to pump out antibiotics
- presence of neutralising enzymes eg B lactamase degrade B lactam ring of pencillin
what are 3 microbial interactions within biofilms
1) cell-to-cell communication
- unicellular bacteria coordinates activities and act similar to multi cellular organisms
- quorum sensing: signalling quorum molecules regulate gene expression
2) metabolic communication
- metabolic product of one organism is used as a nutrient by other organisms
- nutrients for oral bacteria also available from saliva, gcf, foods
3) gene transfer
- biofilm acts as reservoir with transferable mobile genes
- lysed cells in biofilm act as DNA donors and increase opportunity for horizontal gene transfer
what is a normal nervous response of the pulp to bacterial ingress
- microbes diffuse through dentinal tubules and reach pulp
- pulp densely innervated with sensory nerve fibres and release immunomodulatory neuropeptides (eg substance P, CGRP calcitonin gene related peptide) in response to irritation
- give neurogenic inflammation
neurogenic inflammation involves
- vasodilation and hence outflow flow of dentinal fluid
- form tertiary/ reparative dentine and hence decrease dentine permeability
- plasma extravasation will cause increase in immune cellular response
- overall: decelerate ingress of microorganisms
how does healthy pulp progress to reversible pulpitis
Bacterial invasion
- violation of enamel/cementum protecting underlying pulp dentine complex which allow virulence factors to pass through dentinal tubules
- virulence factors like PAMP bind to TLR (PRR) on odontoblasts and trigger innate immune response (releas pro inflam cytokines)
innate immune response (when caries is still >2mm to pulp)
- is the non antigen specific phase
- TLR activates neutrophils, NK cells and macrophages (inflammation & phagocytosis)
- outward flow of dentinal fluid from dentinal tubules and this dentinal fluid contains immune cells which mount pulpal defence before pathogen enters pulp, dilutes toxins, maintains +ve pressure and hence prevents ingress of microorganisms
odontoblasts also play a role in innate imm response
- 1st barrier at pulp dentine junction
- release pro inflam cytokines which leads to cell recruitment
- release nitric oxide, beta defensives to kill pathogen
- release LPS binding protein to decrease pathogenicity of pathogens
adaptive immune response (caries is deep, <2mm to pulp)
- this is the cell mediated immunity one, at cellular level
- APC dendritic cells present antigen to Th cells and activate adaptive immune response
- the greater the degree of bacterial invasion to pulp, the increased accumulation of T cells
- killer T cells can kill microbes directly
how does reversible pulpitis progress to irreversible pulpitis
there is a further adaptive immune response (humoral immunity) at an antibody level
- the greater the degree of bacterial invasion to the pulp, the increased accumulation of B cells
- B cells are activated by T cells and differentiate into plasma cells to produce antigen specific antibodies
critical juncture in pulpitis (Trowbridge 1981)
- bacteria enter pulp
- neutrophils release ROS, NO, lysozymes to eliminate bacteria but this degrades pulp tissue and causes liquefactive necrosis and the formation of microabscesses
- microabscesses coalesce to form localised abscess
how does irreversible pulpitis progress to pulp necrosis
- as inflammation progresses, increased oedema in pulp, increased intrapulpal pressure beyond shunting abilities
- pulp is encased in rigid walls and is a low compliance system with no collateral blood circulation
- compression against the thin walled venules will cause decreased blood flow to pulp and hence hypoxia and pulp necrosis
- lack of circulation also means:
1) immune cells cannot enter the root canal system and hence it protects bacteria from the immune system
2) o2 is depleted by the bacteria and is not replenished and hence is a more conducive envt for anaerobes
defn of apical periodontitis, when does it occur
is an inflammatory lesion in periodontal tissues caused mostly by bacterial elements derived from infected root canal system
in disease, it is characterised by bone resorption at the apical region of infected teeth
it also has a protective function whereby it confines bacteria and prevents spread to adjacent bone marrow spaces and other distant sites
it occurs in
- unsuccessful control of primary infection
- inadequate obturation which leads to re infection
- extrusion of medicaments which will give toxic tissu effects & FB reaction
how does a periapical lesion form
begins with PERIAPICAL INFLAMMATION
- starts before the pulp is completely necrotic
- bacteria colonises root canal space in layers/ aggregates & reaches apical region
- these bacteria produce virulence factors that exit via apical formaen into surrounding periodontal tissues and hence induce periapical inflammation
1) B cells produce IgG against bacteria at apex and the bacteria attaches to IgG antibody and activates complement, giving vascular permeability and oedema
2) activated macrophages produce IL1 & TNFa and these cytokines recruit neutrophils which then activate T lymphocytes and induce proliferation of B lymphocytes
then development of periapical lesion ensues
- activated macrophages produce IL1B & TNFB to recruit and activate osteoclast precursors
- RANKL/RANK Pathway occurs whereby RANKL binds to RANK on osteoclast precursors and induces differentiation into osteoclasts, causing bone resorption
- in early stages of AP, osteoclasts> osteoblasts and hence net bone loss near exit of apical foramen
then theres equilibrium established between bacteria & host
AT EQUILIBRIUM:
- PMNs phagocytose & eliminate bacteria, causing them to spontaneously die
- abundance of macrophages clear up disintegrated PMNs & remaining dead bacteria (macrophages> PMN)
when there is DISEQUILIBRIUM:
- means there is apical abscess
- some bacteria have mechanisms to circumvent killing by the neutrophils
- accumulation of PMNs with no ability to eradicate infectious elements will end up exceeding clearing ability of macrophages and then neutrophils>macrophages
- neutrophils (PMN) continue to release ROS, NO, lysozymes to attempt to eliminate bacteria but end up degrading CT instead, causing liquefactive necrosis and forming pus
- pressure builds up from pus accumulation
- drains through path of least resistance and exits from mucosa by forming chronic abscess with suppurating sinus tract
what happens at the micrbiological level during a flare up
flare up means that there is a disequilibrium
- sudden increase in quantity of bacteria
- increase in chemotactic factors
- increase in number of PMN reaching area (such that it outnumbers the macrophages)
- increase in vascular permability & fluid extravasation (oedema) -> pain and swelling of flare up
when can a flare up occur
spontaneously:
- ecological change in root canal microbiota that favours growth and dispersion of especially aggressive endodontic pathogens
or
after tx:
- when there is iatrogenic apical extrusion of debri
- mechanical opening & widening of apical foramen allows for more nutrients in the root canal space that favours growth of bacteria that avoided elimination during tx
how does periapical healing occur
- periapical lesions contain growth factors like :
1) BMP activate differentiation of mesenchymal stem cells into osteoblasts and hence give bone formation
2) TGF, PDGF, FGF induce proliferation & chemotaxis of osteoprogenitor cells to give osteoblasts
3) FGF also stimulate fibroblast proliferation to give CT formation
why is PA healing able to occur even without complete elimination of microbes
Nair et al 2005
as long as microbes present in quantities and virulence sub critcial to sustain inflammation of periapex
what is condensing osteitis and the histologic changes?
it is the osteoblastic response to low grade irritation from pulp
histologic changes:
- replacement of cancellous bone with dense compact bone
- may have areas of fibrosis & inflammatory infiltrate
what is chronic hyperplastic pulpitis
- exuberant proliferation of chronically inflamed pulp (pulp polyp)
- in open carious lesions (wo pulp chamber roof)
- usually painless
- occurs in children/ young adults
- usually is the permanent 1M
AETIOLOGY OF apical periodontitis
- inflammation will progress apically throughout radicular pulp and affect apical tissues, and the PDL is thin and confined to bony crypt and hence not capable of mounting an immune response that can stop the advancing microbial ingress
- hence will have bone resorption in order to accommodate the formation of a soft tissue lesion, such that concentration of immunologic reactions can occur to stop the advancing infection
- its like a protective mechanism because it attempts to localise and seal off infection
- it also separates irritants from bone to prevent the spread of bacteria from root canal space to neighbouring sites and the development of osteomyelitis
what happens to a periapical lesion after RCT
- elimination of pathogenic bacteria from root canal system and remove source of activation of macrophages and lymphocytes in PA lesion
- over time, there is a gradual elimination of osteolytic signals (IL1B and TNFB cytokines) which were previously released by activated cells
- osteoclast activation in periapical lesion will gradually subside and there is a shift in balance of osteolytic and osteogenic signals, resulting in new bone formation
- remodelling of bone then occurs, gradually the lesion will fill up with new trabecular bone
- any reduction in mobility is due to resolution of PA lesion and gradual bone fill
histological healing outcomes
3 phases - inflammatory, reparative, remodelling
inflammatory:
- influx of immune cells for phagocytosis and removing necrotic debri
- inflammatory cells release cytokines & chemokines to recruit more immune cells to the site
reparative:
- fibroblasts & mesenchymal cells migrate to the PA region to synthesise ECM components for structural support
- angiogenesis
remodelling:
- collagen fibers become more organised and surrounding tissues mature to restore original PA anatomy
how does an endo abscess form
- influx of polymorphonuclear leukocytes (PMN, neutrophils) undergo apoptosis
- hydrolytic enzymes and ROS released
- CT constituents degrade
- tissue in centre is liquefied with granulomatous tissue at the periphery hence forming pus
- for chronic abscesses, pus is discharged through sinus tract through path of least resistance
what is the diffreence between primary and secondary infections and the type of microorganisms
primary infections are those that initially invade and colonise necrotic pulp tissue
the original microorganisms:
- strep
- prevotella
= p gingivalsis
- facultativ anaerobes: Lactobacillus, actinomyces
secondary infections are those that were introduced into the canal during/after initial treatment
- during tx got lack of aspetic tehcnique
- between appt got leakage of temp restor material
- after obtu, can be prep of post space without isolation
what are persistent infections
are microorganisms from primary infections that resisted intracanal antimicrobial procedures and remained in prepared rct system
- flora is less diverse than primary infection
- gram + facultative bacteria PREDOMINANT: no longer anerobic because we expose the canal, and naocl kills off most bacteria liao
so its mainly enterococcus faecalis
- it is actually a gut commensal, not oral.
- tapped in root canal from food or oral fecal contamination
what are the microorganisms in persistent extraradicular infections
extraradicular infection is almost always a sequela of intraradicular infection. and usually if intraradicular is controlled then extra rad can be addressed by host defence and will subside
microorgs:
- actinomyces sp
- propionibacterium pionicum
these can persist in apical lesiosn after endo tx due to their abiltiy to evade host response and survive in apical tissues, preventing apical healing
