Biofilms • Communities of organisms attached to a solid surface

– Can be nonliving or living tissue surface• Evolve over time consisting of many species• Most important, they are a multiorganism cooperative population

• Two main types of biofilms– Sessile

• Permanently anchored to a surface• Covalently bonded to the surface

• Planktonic– Free floating

• Movement to new habitats

Biofilms (Cont’d) • Examples of biofilms

– Water pipes – Ventilator system of airplanes or convention centers – Wine casks causing spoilage – Serious lung infections of CF patients

• Problems with susceptibility

Natural Biofilm on a Metal Surface

Environmental and Cultural Factors That Affect Biofilm Development

Biofilms: Community of Cells • Most important characteristics

– Attachment efficiency– Nutritional resources– Substrata– Environment shear stress or force

Variables Important in Cell Attachment and Biofilm Formation

Stage I – Development I • Reversible binding to

surface • Increased attachment

via fimbriae and pili

Stage II – Development II • Irreversible binding and aggregate

formation • Decreased motility • Exopolysaccharide (EPS) trap nutrients

and planktonic bacteria

Stage III – Maturation I • Colony thickness of

greater than 10 um thick

Stage IV – Maturation II • Colony thickness of greater than

100 um thick • Some bacteria detach but are

trapped in the film

Stage V • Breaking off of bacteria

leads to start of new biofilms

Stage 0 • Planktonic state

Stages in Biofilm Formation (Cont’d) • Active growing cells • Persister cells

– Cells in a dormancy-like state • Importance

– Cells not actively growing may not be affected by drugs » Cell wall inhibitors » Ribosome inhibitors

• Communication between bacteria

– Quorum sensing (QS) • Pheromones

– Gram positive – low-molecular-weight homoserines – Gram negative – peptides and proteins

Architecture of Biofilms • Outer layer

– Most dynamic and metabolically active cells • Intermediate layer

– Still active but less so – Genetic reservoir for genes involving nutrient utilization and drug

resistance • Inner surface layer

– Persister cells

• Dynamic system – Defends itself as a group

• Freely exchanging traits and retaining resistance

Mechanisms of Pathogenicity

Biofilms as a Defense Mechanism • When culturing organisms

– Catheter tips, artificial joints, etc. – Isolation of individual organisms can be hard to culture

• Sessile – Isolated colonies may not reflect the colonies permanently attached to

the plastic surface • Planktonic

– Isolated colonies may not contain antibiotic resistance, but other colonies in the group may contain resistance

• Look susceptible in a dish but not in the patient – Treatment failure

Biofilms as a Defense Mechanism (Cont’d)

• Protect against pH changes • Interference with immune function

– Prevent phagocytosis – Prevent antigen exposure to antibodies

• Sticky EPS glues biofilm together; stops clearance • Organization of biofilm

– Slow-growing organisms attached to surface show increase resistance to antibiotics

Biofilms as a Defense Mechanism (Cont’d)

• Gene transfer – Transformation – Conjugation

• Greater genetic potential as a group than alone – Eventually the virulence factors cluster, causing a worsening of disease

Diseases Associated with Biofilms • Primarily indwelling medical devices

– Examples include • Artificial heart valves • Prostheses • Catheters

• Can be tissue and vessels as well – Some disease as it progresses from acute to chronic diseases

Dental Biofilms • Plaque

– Caries (cavities) – Periodontal disease

• Dental cleaning removes plaque – Biofilm develops again

• Acquired pellicle – Organisms produce

glycans to produce slime layer

• Sugars – Broken down to acids that damage

teeth

Laboratory Consequences Associated with Biofilms

• Cultures – Require growth to get colonies

• Problem is colonies won’t grow under normal conditions • False negatives

– Improper sample collection • Swabs or culturing outer surface of equipment

• Aggregates of organisms – Single colonies can represent up to 100,000 bacteria of mixed origin

• Thus amounts of each organism are greatly underestimated or not considered significant

• Antibiotic susceptibility – Single isolates that are members of a biofilm may not represent the

genetic potential or resistance of a community

Detection of Biofilms • PCR with pathogen-specific probes • Confocal laser scanning microscopic imaging

– CLSM

Potential Interventions • Establishing biofilms in 96 well plates

– Minimal biofilm elimination concentration (MBEC) • Help select successful concentrations of drugs and appropriate

concentration • Treatment outcomes

– Prevent metastasis – Reduce bioburden – Prevent attachment

• Other treatments – Sonication to disrupt biofilm – Toxic compounds (silver)