Biofilms in

Infection
Dr.T.V.Rao M D

Beginning of Microbes

Bacteria first appeared

on earth about 3.6
billion years ago, long
before the appearance
of Homo sapiens around
100,000 years ago..
Van Leeuwenhoek
was the first person to
visualize, graphically
illustrate, and label
"animalcules" (bacteria)
that he found in plaque
scraped from his own
teeth.

Bio film constitutes

A

biofilm is an aggregate of
microorganisms in which cells are stuck
to each other and/or to a surface. These
adherent cells are frequently embedded
within a self-produced matrix of
extracellular polymeric substance (EPS).
Biofilm EPS, which is also referred to as
"slime," is a polymeric jumble of DNA,
proteins and polysaccharides.

Biofilm is a complex
substance.

A biofilm is a complex
aggregation of
microorganisms growing
on a solid substrate.
Biofilms are
characterized by
structural heterogeneity,
genetic diversity,
complex community
interactions, and an
extracellular matrix of
polymeric substances.

Biofilms found in Nature

everywhere where is there is
moisture

More

properly known as biofilm,

slime cities thrive wherever there is
water - in the kitchen, on contact
lenses, in the gut linings of animals.
When the urban sprawl is extensive,
bio films can be seen with the naked
eye, coating the inside of water pipes
or dangling slippery and green from
plumbing." (Coghlan 1996)

Biofilm supports the

Bacterial growth

Biofilm are a common mode of bacterial

growth in nature and their presence has an
enormous impact on many aspects of our
lives, such as sewage treatment, corrosion of
materials, food contamination during
processing, pipe collapse, plantmicroorganisms interaction in the biosphere,
the formation of dental plaque, the
development of chronic infections in live
tissue (mastitis, Otitis, pneumonia, urinary
infections, osteomyelitis) or problems related
to medical implants.

Formation of Biofilms
Biofilms

may form
on living or nonliving surfaces,
and represent a
prevalent mode of
microbial life in
natural, industrial
and hospital
settings

Biofilms increases Antibiotic

resitance
With
microorganisms
are highly
resistant to
antimicrobial
treatment and are
tenaciously bound
to the surface

Mechanisims of Biofilm
formation

Formation of a biofilm
begins with the
attachment of freefloating microorganisms
to a surface. These first
colonists adhere to the
surface initially through
weak, reversible van der
Waals forces. If the

colonists are not

immediately
separated from the
surface, they can
anchor themselves
more permanently
using cell adhesion
structures such as pili

Factors Influencing Rate and

Extent of Biofilm Formation

Indwelling medical device when contaminated

with microorganisms, several variables
determine whether a biofilm develops. First
the microorganisms must adhere to the
exposed surfaces of the device long enough to
become irreversibly attached. The rate of cell
attachment depends on the number and types
of cells in the liquid to which the device is
exposed, the flow rate of liquid through the
device, and the physicochemical
characteristics of the surface

Technology understands
Biofilms better

Technological progress in
microscopy, molecular
genetics and genome
analysis has significantly
advanced our
understanding of the
structural and molecular
aspects of biofilms,
especially of extensively
studied model
organisms such as
Pseudomonas
aeruginosa.

Steps in Biofilm
Development

Biofilm development can

be divided into several
key steps including
attachment, micro
colony formation, biofilm
maturation and
dispersion; and in each
step bacteria may recruit
different components
and molecules including
flagella, type IV pili, DNA
and exo polysaccharides.

Stages of biofilm
development.

Steps in Biofilm
formation

Bacteria associated with

Biofilms differ
Bacteria

living in a biofilm can have

significantly different properties from
free-floating bacteria, as the dense and
protected environment of the film allows
them to cooperate and interact in various
ways. One benefit of this environment is
increased resistance to detergents and
antibiotics, as the dense extracellular
matrix and the outer layer of cells protect
the interior of the community.

Biofilms major cause of

Nosocomial infections

Microbial biofilms,
which often are
formed by
antimicrobialresistant
organisms, are
responsible for
65% of infections
treated in the
developed world.

Biofilms a Great threat to

Implants
A

significant number of people are

affected by biofilm infections which
develop on medical devices implanted in
the body such as catheters (tubes used to
conduct fluids in or out of the body),
artificial joints, and mechanical heart
valves. When implanted material
becomes colonized by microorganisms, a
slow developing but persistent infection
results.

Biofilms a Grwoing concern in

Modern Medicine
Prosthetic

device infections, such as

those involving artificial heart valves,
intravascular catheters, or prosthetic
joints, are prime examples of biofilmassociated infections. With the
increasing use of such devices in the
modern practice of medicine, the
prevalence of these infections is
expected to increase.

Dental plaque

Dental plaque is a
yellowish biofilm
that build up on
the teeth. If not
removed regularly,
it can lead to
dental caries.

Dental plaques

The formation of
dental plaque bio
films includes a
series of steps that
begins with the
initial colonization
of the pellicle and
ends with the
complex formation
of a mature bio film.

Formation of Dental
Biofilms

Additionally, through
the growth process of
the plaque bio film, the
microbial composition
changes from one that
is primarily grampositive and
streptococcus-rich to a
structure filled with
gram-negative
anaerobes in its more
mature state.

Cell-cell signaling (ex. quorum

sensing), and communication with
different bacteria enhance Biofilm
formation

Biofilms everywhere

They're everywhere:
on your shower
curtain, on medical
devices implanted in
patients, on rocks in
rivers and streams,
and in your nose.
While the sheer
number of different
organisms a biofilm
may contain makes it
a challenge to study,

CDC on Biofilms
Biofilms

form on the surface of

catheter lines and contact lenses.
They grow on pacemakers, heart
valve replacements, artificial joints
and other surgical implants. The CDC
(Centers for Disease Control)
estimate that over 65% of
Nosocomial (hospital-acquired)
infections are caused by biofilms.

Biofilms interfere in Antibiotic

Therapy

Bacteria growing in a
biofilm are highly
resistant to antibiotics,
up to 1,000 times more
resistant than the same
bacteria not growing in
a biofilm. Standard
antibiotic therapy is
often useless and the
only recourse may be
to remove the
contaminated implant.

Biofilm and Antibiotic

resistance

A key property of bio

films is that individual
microorganisms are
bound together by a
polymeric substance
excreted by the
microorganisms.. This
protective
encapsulation is
believed to play a role
in some antibioticresistant infection.

Bacterial resitance and

Biofilms
Another

area of great importance from a

public health perspective is the role of
biofilms in antimicrobial-drug resistance.
Bacteria within biofilms are intrinsically
more resistant to antimicrobial agents
than plank tonic cells because of the
diminished rates of mass transport of
antimicrobial molecules to the biofilm
associated cells or because biofilm cells
differ physiologically from plank tonic
cells

Biofilms in Cystic fibrosis

Biofilms are
involved in
numerous
diseases. In cystic
fibrosis patients
have Pseudomonas
infections that
often result in
antibiotic resistant
biofilms.

Endocarditis and Biofilms

Microorganisms may attach and develop

biofilms on components of mechanical heart
valves and surrounding tissues of the heart,
leading to a condition known as prosthetic
valve endocarditis. The primary organisms
responsible for this condition are S.
epidermidis, S. aureus, Streptococcus spp.,
gram-negative bacilli, diphtheroids,
enterococci, and Candida spp. These
organisms may originate from the skin, other
indwelling devices such as central venous
catheters, or dental work.

Eye and Biofilms

The presence of
bacterial biofilms has
been demonstrated on
many medical devices
including intravenous
catheters, as well as
materials relevant to the
eye such as contact
lenses, scleral buckles,
suture material, and
intraocular lenses. Many
ocular infections often
occur when such
prosthetic devices come
in contact with or are
implanted in the eye.

Biofilms and Contact lenses

Bacterial biofilm
formation on contact
lenses and contact lens
storage cases may be
a risk factor in contact
lens-associated corneal
infections. Studies
have shown that
contamination of lens
cases by bacteria,
fungi, and amoebae is
common with 20% to
80% of lens wearers
having a contaminated
lens case.

Urinary catheters and

Biofilms

Urinary catheters are tubular latex or

silicone devices, which when inserted may
readily acquire biofilms on the inner or
outer surfaces. The organisms commonly
contaminating these devices and
developing biofilms are
S.
epidermidis, Enterococcus faecalis, E.
coli, Proteus mirabilis, P. aeruginosa,
K. pneumoniae, and other gram-negative
organisms. The longer the urinary catheter
remains in place, the greater the tendency
of these organisms to develop biofilms and
result in urinary tract infections.

Biofilms and indwelling medical

devices

Biofilms on indwelling medical devices may

be composed of gram-positive or gramnegative bacteria or yeasts. Bacteria
commonly isolated from these devices
include the gram-positive Enterococcus
faecalis, Staphylococcus aureus,
Staphylococcus epidermidis, and
Streptococcus viridians; and the gramnegative Escherichia coli, Klebsiella
pneumoniae, Proteus mirabilis, and
Pseudomonas aeruginosa.

Indwelling catheters and

Biofilms*

Central venous catheters, the reference

method for quantification of biofilms on
catheter tips is the roll-plate technique, in
which the tip of the catheter is removed
and rolled over the surface of a
nonselective medium. Quantification of the
biofilm depends on the number of
organisms recovered by contact with the
agar surface. Biofilm-associated cells on the
inner lumen of the device are not detected
with this method, which has low diagnostic
sensitivity and low predictive value for
catheter-related bacteraemia.

Indwelling catheters and

Biofilms*

In addition, this method cannot detect

more than 1,000 colony-forming units
(CFU) per tip. A method that used
sonication plus vortexing as a means
of quantifying biofilms on catheter tips
showed that a level of 104 CFU per tip
is predictive of catheter-related
septicaemia

* Biofilms and Device-Associated Infections

Rodney M. Donlan Centers for Disease Control and Prevention
Atlanta, Georgia, USA

Antibiotic therapy alone may

not cure ?

Antimicrobial agents are

administered during valve
replacement and
whenever the patient has
dental work to prevent
initial attachment by
killing all microorganisms
introduced into the
bloodstream. As with
biofilms on other
indwelling devices,
relatively few patients
can be cured of a
biofilm infection by
antibiotic therapy alone

Biofilms a concern in
Antimicrobial Therapy

Microbial biofilms may pose a public health

problem for persons requiring indwelling
medical devices. The microorganisms in
biofilms are difficult or impossible to treat
with antimicrobial agents; detachment from
the device may result in infection. Although
medical devices may differ widely in design
and use characteristics, specific factors
determine susceptibility of a device to
microbial contamination and biofilm
formation.

Biofilms need higher

concentration of Antibiotics
Biofilms

are remarkably difficult to treat

with antimicrobials. Antimicrobials may
be readily inactivated or fail to penetrate
into the biofilm. In addition, bacteria
within biofilms have increased (up to
1000-fold higher) resistance to
antimicrobial compounds, even though
these same bacteria are sensitive to
these agents if grown under plank tonic
conditions.

Biofilms help Gene

transfer

Biofilms increase the

opportunity for gene
transfer
between/among
bacteria.. Gene
transfer can convert
a previous a virulent
commensals
organism into a
highly virulent
pathogen.

Biofilms Quorum
sensing

Certain species of
bacteria communicate
with each other within
the biofilm. As their
density increases, the
organisms secrete low
molecular weight
molecules that signal
when the population
has reached a critical
threshold. This process,
called quorum
sensing, is responsible
for the expression of
virulence factors.

Quorum sensing helps the

survival of pathogens

Biofilms contribute for new

phenotypes
Bacteria

express new, and sometimes

more virulent phenotypes when growing
within a biofilm. Such phenotypes may
not have been detected in the past
because the organisms were grown on
rich nutrient media under plank tonic
conditions. The growth conditions are
quite different particularly in the depths
of biofilms,

Biofilms protects from

Immune responses
Bacteria

embedded within biofilms are

resistant to both immunological and
non- specific defence mechanisms of
the body. Contact with a solid surface
triggers the expression of a panel of
bacterial enzymes which catalyze the
formation of sticky polysaccharides that
promote colonization and protection.

Biofilms Protects from

Phagocytosis
Phagocytes

are unable to effectively

engulf a bacterium growing within a
complex polysaccharide matrix
attached to a solid surface. This
causes the phagocyte to release
large amounts of pro-inflammatory
enzymes and cytokines, leading to
inflammation and destruction of
nearby tissues.

Current objectives on
Biofilm research

o Development of improved imaging of

biofilms in situ;
o Development of improved clinically
relevant in vitro and in vivo models of
biofilms under specific in vivo conditions such
as flow rate, nutrient content, and
temperature;
o
Development of better probes (genetic,
metabolic, and immunological) for real- time
analysis;
o
Studies of quorum sensing/signaling
molecules;

Current objectives on Biofilm

research

o Further characterization of biofilm-specific

gene expression;
o
Studies of the exchange of genetic material
within biofilms;
o
Studies of organic contaminants on substrata,
and their influence on biofilm structure;
o Development
of novel approaches to control pathogenic
bacteria by, for example, devising strategies
to favour growth of non-pathogenic
microorganisms in biofilm communities;

Current objectives on Biofilm

research

o Studies of pathogenic
mechanisms of microbes
growing in biofilms;
o Elucidation of
mechanisms of
resistance of biofilms to
antimicrobial agents;
o Studies of
host immune
responses, both
innate and adaptive
to biofilms;

Current objectives on Biofilm

research

In studies of infectious lung disease in cystic

fibrosis;
o Studies on the potential of diagnostic
procedures such as Bronchoalveloar lavage
and bronchoscopy to disturb local biofilm
flora and inoculate distant locations;
o Development of
mathematical models and computer
simulations of biofilms;
o Development of the
methodology for the prevention and control
of biofilms from catheters, water unit lines,
and other clinically important solid surfaces;.

Searching for alternatives

Tissue engineering

Role of biofilms in multiple pathologies and the

difficulty in resolving these pathologies speaks
to the importance of developing means of
replacing or enhancing the therapies already
in use. The use of synthetic materials in the
body ranges from catheters to mesh to stents
to heart valves and beyond. Until the
development of viable and practical tissue
engineering, then number and types of
applications in which synthetic materials are
used will continue to increase.

Emerging Methods

Several researchers
are finding solutions
for the cure of Biofilms
, yet it is experimental,
with advances in
molecular biology
better model
treatments can be
identified to reduce
the problem of Biofilm
interference in
Antibiotic therapy.

Created for Dr.T.V.Rao MDs e

learning series.
Email
doctortvrao@gmail.com