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Probiotics and
BiofilmsBy : Tewdros Tesfa (BSc., MSc.) Haramaya University Wondmagegn Demsiss (BSc., MSc.) Wollo University
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Outline
ProbioticsMechanism of action of probioticsIntroduction About biofilmFormation, development and dispersal of
biofilmsProperties of biofilms and Quorum sensingUses and problems of biofilmsBiofilms and infectious diseasesControl and removal of biofilms
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Objectives
At the end of this presentation you will be able to:Define biofilms and ProbioticsUnderstand the mechanism of action of ProbioticsDescribe how biofilms are formed. State the role of biofilms in infectious disease.Understand the antimicrobial drug resistance
mechanisms of biofilms.Know the prevention and control mechanism of
biofilms.
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Definition
Live microorganisms which when administered in adequate amounts confer a health benefit on the host” (FAO/WHO, 2002)
Are live microorganisms; could be fungi or bacteria
Food ingredientsHave beneficial health effectse.g. Lactobacillus, Bifidobacterium,
Saccharomyces boulardii (yeast)
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Definition
Prebiotics- dietary substances (non digestible by human enzymes, non starch carbohydrates) e.g. inulin, lactulose, oligofructose Stimulate favorable growth or
activity of probiotic bacteria
Synbiotics (eubiotics)- Products that contain both probiotics and Prebiotics
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Mechanism of action
Anti cancer effects Are shown to reduce concentration of carcinogenic chemicals (Scavenge superoxide radicals)
Modify epithelial cell kinetics
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Mechanism of action
Anti diarrheal effect Lowering GI pHSecreting bacteriocinsReconstruction of normal flora (especially antibiotics associated and radiation induced diarrhea)
Competing for adhesion sites
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Mechanism of action
Immunomodulation Enhanced barrier
Increased mucus secretion Epithelial membrane integrity
Activation Bacterial surface antigens
Suppression Inducing T-reg cells Inducing regulatory cytokines; TGF-β, IL-10 Counteract pro-inflammatory cytokines
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Mechanism of action
Anti-allergy effectsReducing antigenic challenge; by
enhanced membrane integrity ( as most allergic reactions are thought as a result of large antigenic insult after barrier disintegrates)
Modification of allergen receptor
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Mechanism of action
Stomach and Urogenital health
Inhibitory effect of lactic acid for
H.pylori
An estimated reason for Urogenital
health is maintenance of acidic
environment (lactic acid) and
production of other antibacterial
chemicals (e.g. H2O2)
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Other roles of Probiotics
Reduction of hypertension- by fermentation end products which have inhibitory effect on angiotensin I conversion
Lowering blood cholesterol levelImprovement in Ca absorption Production of some vitamins and
digestive enzymes
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Questions to be answered
Dose of the probiotic bacteria For how long should it be takenCorrelation of specific probiotic
species with its specific action and target
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Summary
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Introduction
What are biofilms?Why should we care about biofilms?What makes them special?What happens as a result of biofilms?How they are formed?How they are removed?
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Introduction about biofilms
The concept of bacterial biofilm was proposed in 1936.
In natural aquatic environments bacteria are predominantly not free floating but grow as multi-species communities attached to submerged surfaces.
Over 90% of all bacteria live in biofilms. In clinical medicine, many environments
provide optimal conditions for the formation of bacterial biofilm, such as contact lenses, urinary catheters, and so on.
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Introduction
What is Biofilm??A biofilm is a community of micro-
organisms, attached to a wet or moist surface. Microorganisms include bacteria, yeasts, algae, fungi.
These microbes make a sticky, slimy, gel-like substance and surround themselves with it.
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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 known as extracellular polymeric
substance (EPS).
Biofilm EPS, which is also referred to as "slime," is a
polymeric jumble of DNA, proteins and polysaccharides.
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Bio film constitutes
Biofilms may form on living or non-living surfaces
and can be prevalent in natural, industrial and
hospital settings.
For example, a pebble in a pond is covered in slimy
green stuff. That slime is a biofilm.
The slippery layer that builds up inside a waste
pipe is a biofilm.
The plaque that builds up between the gum and
the tooth is a biofilm.
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Why Do Bacteria Make Biofilms?
Biofilms protect the pathogenic bacteria, making them hard to kill.
Hiding in biofilms, bacteria can spread throughout the body.
Large, sticky biofilms resist the attack of the immune system and antibiotics.
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Formation and Development of Biofilms
• Biofilm formation is not a random process
Biofilms start as just a few bugs forming a thin layer.
They can develop into complex, three dimensional structures housing millions of individual bacterium.
Like miniature cities, they have towers, columns, bridges and channels for the flow of nutrients.
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Formation and Development of Biofilms
What is the use of understanding the….?
formation, composition, and characteristics of the biofilm assists in
its control and for developing treatment strategies of biofilm diseases.
The greater our understanding of the processes involved in biofilm formation, the greater the chance of developing remedies.
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Formation and Development of Biofilms
The pattern of biofilm formation can be divided into three phases:
I. Attachment of bacteria to a solid surface;
II. Formation of micro colonies on the surface; and
III.Formation of the mature biofilms
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Formation and Development of Biofilms
Neighbors in the biofilm work together to build their niche environment.
Some bugs use their bodies to build support structures, arches, columns.
Others form the foundation and are good at sticking onto the host surface.
Some make the sticky goo or slime that protects them.
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Formation and Development of Biofilms
Formation of a biofilm begins with the attachment of free-floating microorganisms to a solid surface.
These first colonists adhere to the surface initially through weak, reversible adhesion via 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.
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Formation and Development of Biofilms
Once a bacterium attaches to a surface, it activates a whole different set of genes that gives the bacterium different characteristics from those that it had as a free-floating organism.
The first colonists facilitate the arrival of other cells by providing more diverse adhesion sites and beginning to build the matrix that holds the biofilm together.
Some species are not able to attach to a surface on their own but are often able to anchor themselves to the matrix or directly to earlier colonists.
It is during this colonization that the cells are able to communicate via quorum sensing.
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Formation and Development of Biofilms
Once colonization has begun, the biofilm grows through a combination of cell division and recruitment of new bacteria.
Biofilm doubling times are rapid in early development and slower in more mature biofilms.
A second wave of bacterial colonizers adheres to bacteria that are already attached to the surface.
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Formation and Development of Biofilms
The bacteria cluster together to form sessile, mushroom-shaped micro colonies that are attached to the surface at a narrow base.
Co aggregation is the ability of new bacterial colonizers to adhere to the attached cells.
The result of co aggregation is the formation of a complex array of different bacteria linked to one another. This is what we call it mature biofilm
Their Own Little World!!!!
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Formation and Development of Biofilms
Microbes form a biofilm in response to many factors, which may include
cellular recognition of specific or non-specific attachment sites on a surface,
nutritional cues, or in some cases, by exposure of
planktonic cells to sub-inhibitory concentrations of antibiotics.
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Dispersal of biofilms
Dispersal of cells (bacteria) from the biofilm colony is an essential stage of the biofilm life cycle.
Dispersal enables biofilms to spread and colonize new surfaces.
Enzymes that degrade the biofilm extracellular matrix, such as dispersin B and deoxyribonuclease, may play a role in biofilm dispersal.
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Quorum sensing
To work together efficiently, the bugs need to talk to one another. They do so by chemical signals. Bacteria produce diffusible extracellular signalling molecules ( AHLs and oligopeptides)
Bacteria are often considered as simple unicellular organisms, but research has recently shown that many bacteria possess ability to communicate with one another and to organize to communal groups.
to monitor their own population density and to coordinate expression of specific sets of genes in response to the cell density.
This type of cell-density-dependent gene regulation is termed Quorum sensing.
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Properties and structure of biofilms
Biofilms are surface-attached microbial communities with characteristic architecture and phenotypic and biochemical properties distinct from their planktonic counterparts.
One of the best-known of these biofilm specific properties is the development of antibiotic resistance
Biofilms are characterized by • structural heterogeneity, genetic diversity,
complex community interactions, and an extracellular matrix of polymeric substances.
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Properties cont…
The structure of a biofilm community comprises bacterial
A. Micro colonies,B. An extracellular slime layer, C. Fluid channels, and D. A primitive communication system.
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Properties cont…
A. Each micro colony is a tiny, independent community containing thousands of compatible bacteria.
Bacteria in the center --strict anaerobic environment, while other bacteria at the edges -aerobic environment.
Thus, the biofilm structure provides a range of customized living environments (with differing PHs, nutrient availability, and oxygen concentrations) within which bacteria with different physiological needs can survive.
B. The extracellular slime layer is a protective barrier that surrounds the mushroom shaped bacterial micro-colonies.
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Properties cont…
The slime layer protects the bacterial micro colonies from antibiotics, antimicrobials, and host defense mechanisms.
C. A series of fluid channels penetrates the extracellular slime layer.
These fluid channels provide nutrients and oxygen for the bacterial micro colonies and facilitate movement of bacterial metabolites, waste products, and enzymes within the biofilm structure.
D. Each bacterial micro colony uses chemical signals to create a primitive communication system used to communicate with other bacterial micro colonies.
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Depiction of the dynamic nature of a biofilm community.
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USE AND PROBLEMS OF BIOFILMS
BenefitsHelp remove pathogens and reduce the amount of organic
matter in the water, and Eliminate petroleum oil from contaminated oceans or
marine systems. ProblemsThey are associated with persistent infections and They are notorious for their pipe-clogging and -
corroding properties, a costly irritation in industrial settings.
Undesirable taste and odor issues in the water industry; Pipe corrosion within oil and gas industries and souring of
oil in pipelines and storage facilities. Can make sanitation difficult in food preparation areas.
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Biofilms and infectious diseases
• Biofilms have been found to be involved in a wide variety of microbial infections in the body.
It has taken longer to make a clear connection of biofilms to persistent infections in native tissue.
According to the CDC, 65% of all infections in developed countries are caused by biofilms
Biofilms are produced by most if not all pathogens.
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Biofilms and cont…
A well known example of a microbial biofilm in human disease is dental plaque, which is the most known and studied biofilm (secreting acids that destroy teeth and gums).
Pseudomonas aeruginosa, causing an incurable infection in cystic fibrosis patients (Singh et al. 2000),
and Staphylococcus aureus and Staphylococcus epidermidis, infecting indwelling devices (Mack et al. 2004), are probably the best-known biofilm-producing organisms.
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Biofilms and cont…
Biofilms are responsible for a significant portion of acute infections.
A classic case is that of Legionnaire’s disease, an acute respiratory infection resulting from aspiration of clumps of Legionella biofilms detached from air and water heating/cooling systems.
Many food borne pathogens such as E. coli 0157:H7, Listeria monocytogenes, Yersinia enterocolitica, Salmonella spp. and Camphylobacter jejuni can form either single-species or multi-species biofilms on food surfaces and equipment
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Biofilms and cont…
Teeth with Dental Caries The term dental caries refers to the destruction, or necrosis, of teeth usually by bacterial action resulting in a condition commonly known as tooth decay. .
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Examples of diseases associated with biofilmOtitis media (middle ear infections) Periodontal diseaseGingivitisDental CariesChronic bacterial prostatitisEndocarditisSinusitisUTIInfections in cystic fibrosisInfections of all known indwelling devices such as
catheters, orthopedic prostheses, and heart valves;
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Anti-microbial resistance of Biofilm
Bacteria within biofilms are up to 1000 times more resistant to antimicrobials than the same bacteria in suspension.
Extracellular matrix of the biofilm is a passive barrier for antibiotics
To protect the cells against the antibiotics tobramycin and gentamicin P. aeruginosa produces only in biofilms a periplasmic cyclic glucan which complexes antibiotics turning them into harmless compounds.
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Anti-microbial resistance cont…
In biofilms up to 20% of all genes are differently expressed.
Bacteria in biofilms grow slower and with the reduced metabolic activity they are less prone against most antibiotics.
Preliminary evidence suggests that different bacteria within a biofilm can trade genes -- possibly including Abx resistance genes
The infections associated with biofilms may appear to respond to systemic antibiotics because planktonic cells respond and symptoms are reduced, but the persistence of adherent cells leads to recurrent episodes of infection.
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Three hypotheses for mechanisms of antibioticresistance in biofilms
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Prevention and control of biofilm
An effective mechanical control mechanism is biofilm detachment.
Interfere with the bacterial cell to cell communication
A novel alternative to antibiotics are Probiotics.
A unique spray made from stabilized enzymes that can readily dissolve the biofilms.
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Conclusion and recommendation
Biofilms, once considered odd curiosity, are now one of the hottest topics in microbiology. Biofilms occur everywhere: dental plaque is one the most common biofilms that decay teeth,
other biofilms can clog water pipes, others can contaminate
almost any medical device inserted into the body, ranging from contact lenses to catheters and artificial hearts.
Microbiologists have traditionally researched only free-floating, individual bacteria growing in laboratory cultures.“Microbiologists have been barking up the wrong tree since the time of Pasteur,” says Costerton.