Omid Teymournejad(Medical Bacteriology, PhD)

A biofilm is an assemblage of surface-associated microbial cells that is enclosed in an extracellular polymeric substance matrix

Biofilms may form on a wide variety of surfaces, including living tissues, indwelling medical devices, industrial or potable water system piping

Biofilms are a major cause of human infections

The majority of hospital-acquired infections are due to biofilms because they can be life-threatening colonizers of biomedical devices

The first step is the attachment of the bacterial cells to the selected abiotic or biotic surface

Initial attachment is mediated through weak reversible van der Waals interactions between the cell surface and the substratum, which can lead to stronger adhesion receptor mediated attachment

Bacterial cell surface structures such as flagella, fimbriae, LPS, and exopolysaccharidesparticipate in irreversible interactions

The second step corresponds to the development of micro-colonies promoted by the growth and division of the first attached cells (primary colonizers)

The micro-colonies progressively enlarge and coalesce to form the first layer of cells covering the surface

When multiple layers of cells pile up on the surface, the third step of the formation is obtained

indicated by the presence of a mature biofilmcharacterized by the presence of macro-colonies surrounded by water channels that help distribute nutrients and signaling molecules

Finally, to survive when nutrients become limited, or simply to spread and colonize to other niches, some biofilm cells can detachindividually or in clumps

biofilms display a common attribute, the biofilmmatrix.

Contrary to free-floating planktonic cells, biofilmcells are embedded in a self produced extracellular matrix, the extracellular polymeric substance (EPS), that holds them together

Biofilms are composed of about 80–85% EPS (by volume) and only 15–20% cells (by volume)

Although the EPS may vary in chemical and physical properties, its major components are polysaccharides, proteins, and extracellular DNA

The EPS plays a major role in maintaining the integrity of the biofilm and can confer other beneficial properties.

Since the EPS is also highly hydrated, it can prevent desiccation in some natural biofilms

The EPS can also act as a diffusion barrier, preventing toxic substances such as antibiotics and disinfectants from reaching their target

Most biofilms found in nature are polymicrobials, where diverse species expressing multiple phenotypes are involved

The most amazing fact is that even in a mono-species biofilm, phenotypic heterogeneity exists

Cell differentiation in biofilms may depend on the local environmental conditions surrounding the cells

Different concentration gradients of oxygen, nutrients, ions, and chemicals create a wide variety of microhabitats providing conditions suitable for bacterial colonization

Cells located in the upper biofilm layers consume all available oxygen and grow aerobically, while an anaerobic micro-niche developed underneath the aerobic layer

Oxygen- and nutrient depleted regions are found at the bottom layers of the biofilmstructure and under these circumstances, most of the sessile cells are metabolically inactive or dead

A few years ago, Watnick & Kolter proposed the interesting idea that biofilms can be compared to cities

In these cities of microbes, microorganisms are considered “social” organisms able to communicate with one another

Using different chemical languages, bacteria learn about their current cell population and determine when they have reached a critical mass

Using that information, bacteria can thus modify their behavior to carry out processes that would require many cells acting in conjunction to be effective

Cell-to-cell communication is generally carried out by diffusible signal molecules produced and released by bacteria

When bacteria are growing within a biofilm, they secrete signaling molecules (auto-inducers) that increase in concentration as a function of bacterial cell density

In a process called quorum sensing, bacteria communicate with one another by using auto-inducers to regulate their gene expression in response to fluctuations in the cell population density

Two types of quorum-sensing systems are recognized in bacteria:

intra-species communication and inter-species communication

During intra-species communication, several auto-inducers have been identified

Gram-negative bacteria usually use acylhomoserine lactone (AHL) as signal molecules, while Gram-positive bacteria utilize small peptides

During inter-species communication, bacteria use autoinducer-2 (AI-2)

In some cases, quorum sensing does not seem to be involved in biofilm structural development

while for other species, there is evidence that quorum sensing is important for the attachment of bacteria to the surface, the maturation of the biofilm, or the control of events leading to the dispersion of cells

A significant characteristic of microbial biofilms is their high-level drug tolerance

Bacterial biofilms have been shown to have a 100- to 1,000- fold increased tolerance toward antibiotics in comparison to their free-swimming counterparts

Diffusion barrier imparted by the EPS matrix

Phenotypic heterogeneity

Gene Transferring

Secretion of antibiotic degrading enzymes

A primary function that has been attributed to the biofilm matrix is protection,

Several studies have shown that the EPS matrix can act as an impermeable barrier to limit antimicrobial penetration, thereby protecting the biofilm cells

Such protection can be due either to physical hindrance in antimicrobial diffusion or to direct binding of the antibiotics by the EPS matrix

Upon antibiotic treatment, cells at the top of the liquid– biofilm interface die due to their closer exposure, while bacteria embedded deep inside the biofilm are able to survive

Anionic EPS matrix can bind and sequester toxic cationic heavy metals, cationic antimicrobial peptides, and positively-charged antibiotics (e.g. aminoglycosides)

Phenotypic heterogeneity occurs within biofilms notably due to different concentration gradients

Cells localized at the bottom layers of the biofilm are normally found in a growth stage analogous to stationary-phase planktoniccells, while the physiology of cells localized at the top surface layers is similar to exponentially growing planktonic cells

Microbial cells, especially those in the deeper layers of the biofilms where nutrients and oxygen are limited, are associated with a lower growth rate

This reduced metabolic activity might account for the enhanced tolerance toward antibiotics that target bacterial cellular processes such as DNA replication or translation

Conventional antibiotics used to treat infections are mostly effective at killingrapidly growing cells

The decreased metabolic activity of cells found within the deeper biofilm layers may thus contribute to antibiotic tolerance and the persistence of biofilm infections

Biofilms also provide an ideal niche for the exchange of extrachromosomal DNA (plasmids)

Conjugation (the mechanism of plasmid transfer) occurs at a greater rate between cells in biofilms than between planktonic cells