Microbrial Biofilms

Question 1

What is a Biofilm?

Answer 1

Biofilms are communities of microorganisms that adhere to surfaces and form a
protective matrix (EPS).
●
EPS is a complex mixture of polymers produced by the microorganisms within
the biofilm, often referred to as a “slime layer” or “glycocalyx,” is composed of a
variety of substances, including polysaccharides, proteins, nucleic acids (such
as DNA), and other organic and inorganic molecules.
●
Biofilms are found in various environments, including natural and man
made
settings.
●
Biofilms can have both positive and negative impacts on human health and
industry.

Question 2

Where do biofilms appear?

Answer 2

Biofilms can form on a wide range of
surfaces, including medical devices,
natural and industrial surfaces, and living
tissues. Understanding biofilm
composition and the mechanisms of
biofilm formation is crucial in various
fields, including medicine, environmental
science, and industrial engineering,
where biofilms can have both beneficial
and detrimental effects

Question 3

What are the advantages o biofilms?

Answer 3

The biofilm matrix provides several advantages to the microorganisms living
within it:
●
Protection
: The EPS protects microorganisms from external threats, such
as antimicrobial agents, immune system responses, and physical removal.
●
Nutrient Availability
: The matrix helps retain and concentrate nutrients,
promoting the growth of microorganisms within the biofilm.
●
Adhesion
: The EPS facilitates the adhesion of microorganisms to surfaces,
initiating the formation of the biofilm

Question 4

What are the key components of biofilm?

Answer 4

Microorganisms
Extracelleular Polymeric substances
Water channels

Question 5

Key component - microorganisms

How

Answer 5

Bacteria are the most common microorganisms found in
biofilms, but they can also include fungi and algae.
●
The microorganisms within a biofilm can belong to different
species and can form complex, synergistic communities.

Question 6

Key component - EPS

How?

Answer 6

Polysaccharides: They contribute to the slimy texture and provide
structural support to the biofilm.
●
Proteins: Produced by the microorganisms, proteins in the EPS can
have various functions, including adhesion to surfaces and cohesion
within the biofilm.
●
Nucleic Acids: DNA and RNA are part of the EPS, and they can
contribute to the stability of the biofilm structure

Question 7

Key component - water channels

How

Answer 7

Biofilms often have a network of water channels that allow
the flow of nutrients, gases, and waste products within the
biofilm structure.

Question 8

How are they formed and structured?

Answer 8

Biofilms are formed through a process called attachment, where microorganisms
adhere to a surface. They then grow and develop into complex structures with a
protective matrix.
●
Biofilm formation involves a series of key stages, and the process can vary depending
on the type of microorganisms involved and the environment.

Question 9

What are the stages of biofilm formation?

Answer 9

Reversible adhesion of planktonic (free
floating) cells to a surface
●
Irreversible attachment
Production of cell adhesion structures permanently using EPS components
●
Early development
Cells grow and divide to form microcolonies, begin synthesising EPS matrix enclosing cells
●
Maturation
Biofilm architecture develops into a complex, heterogeneous 3D structure containing
pores, water channels, and cell clusters
●
Dispersal
Active dispersal through enzymatic breakdown of matrix or shearing forces or passive
dispersal by single cell detachment

Question 10

What is antimicrobial resistance?

Answer 10

Antimicrobial resistance (AMR) is a significant
concern in the context of biofilms. Biofilms provide
a protective environment for microorganisms, and
the extracellular polymeric substances (EPS) matrix
acts as a barrier that hinders the penetration and
effectiveness of antimicrobial agents.
This protective mechanism contributes to the
increased resistance of biofilm associated
microorganisms to various antibiotics and
disinfectants.

Question 11

Key Factors Contributing to Antimicrobial
Resistance in Biofilms
1.

Answer 11

Physical Barrier
reduced penetration
Nutrient limitation
Quorum sensing/gene expression
persisters
Genetic adaptations
Bio-film associated infections

Question 12

Key Factors Contributing to Antimicrobial
Resistance in Biofilms
1. Physical Barrier

Answer 12

The EPS matrix physically restricts the diffusion of antimicrobial agents, preventing them
from reaching the microorganisms embedded within the biofilm.

Question 13

Key Factors Contributing to Antimicrobial
Resistance in Biofilms

2. Reduced Penetration:

Answer 13

The dense and three
dimensional structure of biofilms hinders the penetration of
antibiotics, making it difficult for these agents to reach all microorganisms in the biofilm.

Question 14

Key Factors Contributing to Antimicrobial
Resistance in Biofilms
3. Nutrient Limitation:

Answer 14

Biofilms create microenvironments with different nutrient availability, oxygen levels, and
pH. These variations can induce changes in microbial physiology, potentially leading to
reduced susceptibility to antimicrobial agents.

Question 15

Key Factors Contributing to Antimicrobial
Resistance in Biofilms

4.
Quorum sensing / Gene Expression:

Answer 15

Microorganisms within biofilms often communicate, leading to the upregulation of genes
associated with antimicrobial resistance, making biofilm associated bacteria more
resilient

Question 16

Key Factors Contributing to Antimicrobial
Resistance in Biofilms

Persisters

Answer 16

Some microorganisms within biofilms exist in a dormant or
persister state, which makes
them less susceptible to antimicrobial agents that target actively dividing cells.

Question 17

Key Factors Contributing to Antimicrobial
Resistance in Biofilms

Genetic adaptions

Answer 17

6. Genetic Adaptations:
   Biofilm
   associated microorganisms may undergo genetic changes that confer
   resistance to antimicrobial agents. These adaptations can be the result of
   selective pressures imposed by the presence of antimicrobials.

Question 18

Key Factors Contributing to Antimicrobial
Resistance in Biofilms

7. Biofilm
   Associated Infections:

Answer 18

In a clinical context, biofilm formation on medical devices or within the human
body can lead to chronic infections that are challenging to treat. The resistant
nature of biofilms contributes to persistent infections.

Question 19

Facts about biofilms in disease

Answer 19

Biofilms associated
with over 80% of
microbial infections

Provide reservoir for
chronic infection
persistence and
biomaterial related
infections

1000x more resistant
to antibiotics than
planktonic cells

Question 20

Why are biofilms in disease a concern?

Answer 20

Biofilm
associated infections are a major concern in medical microbiology.
Biofilms can cause chronic and recurring infections that are difficult to treat.
They can also lead to the development of antimicrobial resistance.
●
Biofilms contribute to the persistence of infections by providing a protected
environment for microorganisms. The matrix protects bacteria from the
immune system and antimicrobial agents, leading to chronic and recurrent
infections.

Question 21

what diseases are biofilms related to?

Answer 21

Dental
Medical Devices
Resp infections
Chronic wounds
Chronic otitis media
Infectious endocarditis
GI infections
UTI

Question 22

How does biofilms affect

dental

Answer 22

Dental Plaque and Tooth Decay: Dental biofilms, commonly known as dental plaque,
form on teeth and contribute to tooth decay (caries) and gum disease (gingivitis and
periodontitis).

Question 23

how does biofilms affect medical devices

Answer 23

Catheter
Associated Infections: Biofilms often form on indwelling medical devices
such as urinary catheters, central venous catheters, and prosthetic joints, leading
to infections that are challenging to treat.

Question 24

How does biofilms effect rest infections?

Answer 24

Cystic Fibrosis: Patients with cystic fibrosis often experience chronic respiratory
infections due to the formation of biofilms in the airways. Pseudomonas aeruginosa
is a common pathogen associated with biofilm formation in the lungs of individuals
with cystic fibrosis.

Question 25

How does biofilms affect chronic wounds?

Answer 25

Diabetic Ulcers: Chronic wounds, particularly in diabetic individuals, can
harbor
biofilms, contributing to delayed wound healing and an increased risk of infection.

Question 26

How does biofilms effect otitis media?

Answer 26

Middle Ear Infections: Biofilms have been implicated in chronic otitis media, where
persistent infections in the middle ear can lead to hearing loss.

Question 27

How does biofilms effect infectious endocarditis?

Answer 27

Heart Valve Infections: Biofilms on heart valves can cause infectious endocarditis, a
serious condition that may require surgical intervention.

Question 28

How does biofilms effect GI infections?

Answer 28

Inflammatory Bowel Diseases: Biofilms have been observed in the gastrointestinal
tract of individuals with inflammatory bowel diseases (IBD), potentially contributing
to the chronic inflammation seen in conditions like Crohn’s disease.

Question 29

How does biofilms effect UTI

Answer 29

Cystitis: Biofilms can form on the uroepithelium and catheters, contributing to the
persistence of urinary tract infections.

Question 30

How do you address bio-film associate diseases?

Answer 30

Addressing biofilm
associated diseases requires a comprehensive
understanding of the mechanisms involved in biofilm formation, persistence,
and resistance to treatment. Researchers are actively exploring strategies to
disrupt biofilms, enhance the effectiveness of antibiotics, and develop targeted
therapies to mitigate the impact of biofilm related infections on human health.

Question 31

How to prevent/control biofilms?

Answer 31

Surface Modification
Antimicrobial agents
Enzymatic disruption
Quorum sensing inhibition
Biofilm disrupting compounds
physical removal
innovative drug delivery system
probiotics and competitive exclusion
electrochemical approaches
education and awareness

Question 32

Summary of prevention and control Strat for biofilms

Answer 32

Preventing and controlling biofilms is a complex challenge due to the resilient
nature of these microbial communities. However, various strategies have been
developed to mitigate biofilm formation and its associated issues
●
Here are some common approaches to biofilm prevention and control:

Question 33

How do we prevent?

Surface modification

Answer 33

Surface Modification
Altering the surface properties of materials can make it less favourable for
microbial attachment. This includes using antimicrobial coatings, modifying surface
roughness, or applying materials with inherent resistance to biofilm formation.

Question 34

Prevent how
Antimicrobroial agents

Answer 34

Using antimicrobial agents, such as antibiotics, antiseptics, and disinfectants, can
help control biofilm formation. However, it’s important to note that biofilm
associated microorganisms may exhibit increased resistance to traditional
antimicrobial agents.

Question 35

Prevent how

enzymatic disruption

Answer 35

Enzymes that break down the biofilm matrix, such as
dispersin B and DNase, can be
used to disrupt biofilms. These enzymes target the extracellular polymeric
substances (EPS), weakening the biofilm structure.

Question 36

Prevent how

quorum sensing inhibition

Answer 36

Interfering with quorum sensing, the communication system used by
microorganisms within biofilms, can disrupt the coordinated behavior of bacteria
and reduce biofilm formation. Quorum sensing inhibitors are being explored as
potential therapeutic agents.

Question 37

Prevent how

Biofilm disrupting compounds

Answer 37

Developing compounds specifically designed to disrupt biofilm formation and
maintenance is an active area of research. These compounds may target the matrix
components or interfere with microbial adhesion.

Question 38

prevent how
physical removal

Answer 38

Regular cleaning and physical removal of biofilms from surfaces can help prevent
their establishment. This is important in both industrial settings and healthcare
environments.

Question 39

Prevent how
Innovative drug delivery systems

Answer 39

Innovative drug delivery systems

Developing drug delivery systems that improve the penetration of antimicrobial
agents into biofilms is an area of ongoing research. These systems aim to enhance
the effectiveness of therapeutic agents against biofilm associated infections.

Question 40

How to prevent

probiotics and competitive exclusion

Answer 40

Introducing beneficial microorganisms or probiotics that can outcompete
pathogenic bacteria for resources can be a preventive strategy. This approach is
being explored in various contexts, including oral health and gut microbiota
management.

Question 41

How to prevent

electrochemical approaches

Answer 41

Electrochemical methods, such as the use of electric fields, have been investigated
as potential tools for disrupting biofilms. These approaches can interfere with
microbial attachment and biofilm formation.

Question 42

Prevent how
education and awareness

Answer 42

Promoting hygiene practices and raising awareness about the risks of biofilm
associated infections are important preventive measures, especially in healthcare
settings.

Question 43

How do we analyse biofilms?

Methods

Answer 43

Microscopy techniques
Flurescene in situ hybdrisation (FISH)
Live/dead staining methods
crystal violet staining
quantitative PCR
Protein and carb assays
micro electrode measurements
raman spectroscopy

Question 44

Biofilm Analysis Methods

Microscopy Techniques:

Answer 44

Confocal Laser Scanning Microscopy (CLSM): CLSM provides detailed three
dimensional images of biofilms. Fluorescent dyes can be used to stain different
components of the biofilm, such as microbial cells, extracellular polymeric
substances (EPS), and specific proteins.
2.
Scanning Electron Microscopy (SEM): SEM offers high
resolution images of biofilm
surfaces. After fixation and dehydration, biofilm samples are coated with a thin
layer of metal and visualized using electron beams.
3. Transmission Electron Microscopy (TEM): TEM provides detailed internal images
of biofilms by transmitting electrons through thin sections of the sample. It allows
for the visualization of microbial cells and the biofilm matrix.
4. Cryogenic Electron Microscopy (Cryo
EM): Cryo EM is used to image biofilms at
cryogenic temperatures, preserving the native structure of the biofilm. This
technique is especially useful for visualizing dynamic processes within biofilms.

Question 45

Biofilm analysis methods

Fluorescence In Situ Hybridization (FISH):

Answer 45

FISH allows for the visualization of specific microbial species within a biofilm by
using fluorescently labeled DNA probes that bind to target nucleic acid sequences.
This technique helps identify the distribution of different microorganisms.
●
Microbial Viability Staining:

Question 46

Biofilm analysis methods

live/dead staining methods

Answer 46

Use fluorescent dyes to
differentiate between live and dead
microbial cells within biofilms. This
helps assess the overall viability
and metabolic activity of biofilm
communities.

Question 47

Biofilm Analysis Methods
Crystal Violet Staining:

Answer 47

Crystal violet is a common dye used to quantify biofilm biomass. After staining, the
dye is solubilized, and the absorbance is measured to estimate the amount of
biomass formed.

Question 48

Biofilm Analysis Methods
Quantitative Polymerase Chain Reaction (qPCR):

Answer 48

qPCR is used to quantify the abundance of specific microbial species or genes
within biofilms. This molecular technique provides information about the
composition and dynamics of biofilm communities.

Question 49

Biofilm Analysis Methods
Protein and Carbohydrate Assays:

Answer 49

Biochemical assays can quantify the amounts of proteins and carbohydrates
present in biofilms, providing insights into the composition of the extracellular
matrix.

Question 50

Biofilm Analysis Methods
Microelectrode Measurements:

Answer 50

Microelectrodes can be used to measure changes in pH, oxygen concentration, and
other parameters within biofilms. These measurements help understand the
microenvironmental conditions within the biofilm.

Question 51

Biofilm Analysis Methods
Raman Spectroscopy:

Answer 51

Raman spectroscopy provides information about the chemical composition of
biofilms by measuring the scattering of laser light. It can be used to analyse the
distribution of specific molecules within biofilms.

Question 52

Overview of the analysis methods of biofilm why its good

Answer 52

Combining multiple techniques allows researchers to obtain a
comprehensive understanding of biofilm structure, composition, and
behaviour.
●
The choice of methods depends on the specific research questions and
the characteristics of the biofilm being studied.