Oral Microbiology

Biofilms & Communities

Overview of Biofilms and Infections

  • CDC Report (1999): 65% of infections are associated with biofilms.

  • NIH Report (2002): 80% of infections attributed to biofilms.

  • Historical Perspective:

    • Leeuwenhoek's recognition of polymicrobial 'animalcules'.

    • Pasteur and Koch focused on isolating single species, neglecting the significance of polymicrobial communities and their interconnections within food webs.

Bacterial Entry Points and Infections

Primary Infection Sites

  1. Mouth:

    • Entry through bleeding gums.

  2. Catheter Entries:

    • Example: Subvenous catheter.

  3. Implanted Medical Devices:

    • Example: Artificial hip joint.

Secondary Infection Risks (Highlighted in Green)

  • Possible sites include:

    • Brain

    • Kidneys

    • Intervertebral spaces

    • Bones surrounding implanted medical devices

Medical Problems of Microbial Adhesion

Common Pathogen-Associated Issues

  1. Teeth:

    • Plaque Flora: Malodor; tooth decay; gum disease; loss of teeth.

  2. Stomach:

    • H. pylori: Causes gastric ulcers.

  3. Intestine:

    • E. coli O157: Linked to gastroenteritis.

  4. Colon:

    • C. difficile: Also associated with gastroenteritis.

  5. Upper Respiratory Tract:

    • Pseudomonas aeruginosa: Related to cystic fibrosis, which can lead to death.

  6. Heart Valves:

    • Streptococci; Yeasts; Fungi: Associated with endocarditis, potentially fatal.

  7. Urethral/Cervical Epithelia:

    • Gonococci; Candida: Linked to gonorrhea and thrush.

    • Treponema: Associated with syphilis, which can be fatal.

  8. Bone:

    • Staphylococci; Streptococci; Bacillus; Proteus: Linked to osteomyelitis and sepsis, both potentially deadly.

Dental Health and Microbiology

Importance of Oral Microbiology

  • Interactions:

    • Involves interactions among various oral microbes and their relationship with the host.

    • Complexity of these interactions can lead to a shift from healthy microflora to disease microflora (dysbiosis).

Microbial Composition in Oral Health

  • Bacteria: Over 700 different species.

  • Fungi: Primarily Candida species.

  • **Other Microbes: **Mycoplasma (wall-less bacteria), Protozoa, Viruses (acellular, replicate within living cells).

Normal Healthy Microflora

  • Facultative Bacteria: Streptococci, lactobacilli, staphylococci, Corynebacteria.

  • Anaerobes: Bacteroides species.

Dental Plaque and Disease

Stages of Infection

  • Health Status vs. Disease Progression:

    • Health → Gingivitis → Caries → Periodontitis.

Role of Inflammation in Oral Disease

  • Dental Plaque: Induces inflammation leading to disease such as gingivitis and periodontal disease.

    • Reference: Center for Biofilm Engineering, Montana State University (1999).

Orality and Bacteria Interactions

Dynamics of Oral Health

  • Optimal oral health involves maintaining a balance between endogenous bacteria and the oral defense systems:

    1. Physical Barriers: Keratinized epithelium; mucin production; salivary flow.

    2. Chemical Compounds: Salivary enzymes, antibacterials, and gingival fluid secretions.

    3. Inflammatory Response: Active immune response to microbial presence.

Quorum Sensing and Communication

  • Interaction occurs at a dynamic interface between host cells and microbial cells, facilitated by quorum sensing signals shared among eukaryotic and prokaryotic cells.

Acquisition of Oral Flora

Early Stages of Microbial Colonization

  1. Birth Stage: Initially sterile.

  2. Within Hours: Colonization by Streptococcus spp. (e.g., S. salivarius).

    • Determined by birth method (cesarean or vaginal) and early caregiver exposure.

  3. Around 1 Year: Introduction of various bacteria: Streptococci, Staphylococci, Lactobacilli, Neisseria, Veillonella, Actinomyces, Fusobacterium spp.

    • Colonization by S. mutans and S. sanguis occurs as teeth emerge.

Later Stages and Influences on Microbial Composition

  • Following tooth eruption:

    • Hard tissue favored by: S. sanguis, S. mutans, Actinomyces spp.

    • Colonization of gingival crevices by anaerobes such as Prevotella spp.

  • Loss of teeth leads to re-colonization by a different microflora.

  • Prosthetic appliances (e.g. dentures) may harbor significant yeast populations similar to plaque on enamel.

Bacterial Characteristics and Oral Conditions

Statistics of Oral Bacteria

  1. 108 bacteria are shed daily in saliva.

  2. Plaque bacteria represent 5% of the total salivary flora.

  3. More than 300 species can be isolated from dental plaque.

  4. 1 mg of dental plaque contains approximately 106 bacterial cells.

  5. Healthy gingiva flora mainly consists of aerobic and facultative anaerobic bacteria.

  6. Over 40% of oral bacteria are unculturable.

Key Dental Diseases

Types of Caries and Periodontal Diseases

  1. Caries:

    • Related to a high sugar diet and poor oral hygiene.

    • Symptoms include cavities, malodor, and deterioration of dentition.

  2. Gingivitis (Non-destructive):

    • Caused by poor hygiene; results in inflamed, bleeding gums.

  3. Periodontitis (Destructive):

    • Typically develops with age unless hygiene is exceptionally poor; involves gum and bone loss.

Streptococcal Virulence Factors in Carious Plaque

Metabolic Processes and Their Effects

  1. Saccharolytic:

    • Organisms metabolize sucrose and other sugars.

  2. Glucosyltransferases:

    • Produce EPS glucan (mutan), contributing to plaque biofilm.

  3. Fructosyltransferases:

    • Generate EPS fructan (inulin), also reinforcing plaque biofilm.

  4. Acidogenic:

    • Ferments sugars generating acids (lactic, acetic, formic) that lower pH.

  5. Aciduric:

    • Can grow and metabolize at low pH levels.

    • pH levels below 4 demineralize hydroxyapatite enamel.

Other Species in Plaque

Bacterial Functions

Veillonella

  • Type: Gram-negative anaerobic cocci.

  • Function: Thrives in acidic carious environments; converts lactic acid to less harmful products.

Lactobacillus

  • Type: Gram-positive facultative anaerobes.

  • Role: Typically symbiotic in gut flora but some species increase in plaque associated with caries; acts as an indicator of disease state.

Specific Pathogens in Gingivitis

Fusospirochetes

  • Bacteria Type: Spirochetes and fusiform bacilli, part of normal oral flora.

  • Infection Risk: In bleeding conditions, can lead to infections such as Acute Necrotizing Ulcerative Gingivitis (ANUG), also known as trench mouth or Vincent's angina, caused by anaerobic bacteria like Prevotella intermedia, Fusobacterium, Treponema, and Borrelia spp.

Periodontitis and Its Key Pathogens

The Big Three Pathogens

  1. Treponema denticola:

    • Gram-negative anaerobic spirochete; key player in tissue destruction and elevated in affected patients.

  2. Porphyromonas gingivalis:

    • Gram-negative pathogen associated with chronic adult periodontitis; known for various virulence factors.

  3. Aggregatibacter actinomycetemcomitans:

    • Gram-negative facultative non-motile rod; associated with localized aggressive periodontitis in adolescents and significant bone loss.

    • Virulence Factors: Include leukotoxin (kills PMNs and monocytes), cytolethal distending toxin, and several immunosuppressive factors inhibiting immune responses.

Imaging and Coaggregation of Plaque

Scanning Electron Micrographs

  • Imaging techniques reveal large populations of filamentous organisms and clustered cocci.

Measuring Species Interaction in Plaque

Coaggregation Scoring System

  • Coaggregation scores can quantify interactions:

    • Score 0: No coaggregation.

    • Score 1: Small coaggregates in turbid suspension.

    • Score 2: Easily visible coaggregates in turbid suspension.

    • Score 3: Coaggregates settle slowly, leaving clear supernatant.

    • Score 4: Large coaggregates settle instantly, leaving a clear supernatant.

Colonization Pathways in Plaque Formation

Stages of Bacterial Colonization

  1. Pioneer Species: Initial colonizers that form biofilms on oral surfaces.

  2. Secondary Colonizers: These organisms include those that can cohabitate with or among existing species to create a more complex biofilm.

  3. Tertiary Colonizers: Include late colonizers that depend on the presence of earlier species.

Specific Surface Interactions in Plaque Formation

  • Components Involved: Salivary proteins (e.g., statherin, amylase) interact with bacterial receptors to initiate colonization.

Quorum Sensing in Oral Biofilms

Key Mechanisms of Communication

Intra-species Communication

  • Gram-positive bacteria produce peptides, such as competence stimulating peptides, facilitating biofilm formation.

Inter-species Communication

  • Mediated by Autoinducer-2 (AHL-2), influencing plaque formation and the expression of virulence factors.

Anti-pathogen Strategies Against Biofilms

Mitigation Techniques

  • Biofilm Prevention: Strategies to prevent formation; attachment blockers or detachment signals may inhibit specific pathogens, like Fusobacterium nucleatum, from contributing to established biofilms.

Visualization of Periodontal Tissue Biofilm

Study Findings

  • Filifactor alocis: Specifically identified in periodontal tissue biofilms which are often unculturable.

  • Imaging Techniques: FISH (Fluorescence In Situ Hybridization) utilized on biopsy samples from periodontal surgeries revealed unique structures formed by F. alocis within the mixed bacterial composition of biofilm.

  • Notable formations include:

    • Tree-like structures within coccoid and fusiform bacteria.

    • Palisade arrangements of F. alocis around rod-shaped eubacterial bacteria.

    • Concentrical aggregations of bacteria surrounding F. alocis in the biofilm context.