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IN   THE NAME OF GOD
BIOFILM FORMATION IN
PATHOGENS BACTERIA
Profosser: Dr.ahmad ali PORBABAEI


by: ABBAS MOROVVATI
CONTENT
 Definition of Biofilm
 History

 Biofilm information

 Biofilm information in vitro

 Where biofilms are found?

 Advantage of biofilm

 Biofilm and antibiotic resistance

 Dis advantage of biofilm

 Biofilms and infectious diseases

 References
DEFINITION OF BIOFILM

 Biofilm: An aggregate of microbes with a distinct
  architecture. A biofilm is like a tiny city in which
  microbial cells, each only a micrometer or two long,
  form towers that can be hundreds of micrometers
  high. The "streets" between the towers are really
  fluid-filled channels that bring in nutrients, oxygen
  and other necessities for live biofilm communities.
 The biofilm is held together and protected by a
  matrix of excreted polymeric compounds called
  EPS .This matrix protects the cells within it and
  facilitates communication among them through
  biochemical signals
 Attachment by pili and etc
BIOFILM
 Biofilms consist of:
1-microbes
2-EPS(poly sacarid--glycoprotein)
3-water(85-98%)
 Biofilm can contain many different types of
  microorganism, e.g. bacteria, archaea, protozoa,
  fungi and algae; each group performing specialized
  metabolic functions.
 Fungal biofilms also frequently contaminate medical
  devices. They cause chronic vaginal infections and
  lead to life-threatening systemic infections in people
  with hobbled immune systems
HISTORY

 Van lion Hook
 zobel

 1913    Sohngen
 1970    enviromental
  places
 1971    Marshal
 1973     charakis
 1976      koshreton
BIOFILM INFORMATION
HOW IS BIOFILM FORMED?

Bacteria continue to grow, outer cells provide a
 physical barrier to protect inner cells




                      surface
  Planktonic bacteria,                              in
  the presence of water,                       attach
  to a surface.
BIOFILM INFORMATION IN VITRO
 1_batch culture model
 2_cotinuous culture model




 NOTE
 Study population of archie bacteria use of
  coenzyme F420 and use of OR NAD and ATP
  production and so
 MONOCULTURE:THIS BIOFILM FORMED BY ONE
    TYPE OF BACTERIA
.    WHERE BIOFILMS ARE FOUND
   . 1. on solid substratums in
    contact with moisture,
    2. on soft tissue surfaces in living
    organisms
    3. at liquid-air interfaces.
   4-Biofilms form on the surface of
    catheter lines and contact
    lenses. They grow on
    pacemakers, heart valve
    replacements, artificial joints and
    other surgical implants
   5-Biofilms grow in hot, acidic
    pools in Yellowstone National
    Park (USA)
   6-In industrial environments,
    biofilms can develop on the
    interiors of pipes
BIOFILM IMAGE BY ESEM
ADVANTAGE OF BIOFILM
   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.
   One benefit of this environment is increased resistance to
    detergents
   Biofilms can help eliminate petroleum oil from contaminated
    oceans or marine systems.
   Bioreactors for water infiltration
BIOFILM AND ANTIBIOTIC RESISTANCE
                                  Little or no effect
                                   because the bacteria in
                                   biofilm are in a different
   Bacteriostatic
                                   phase than most
    Antibiotics
                                   antibiotics target




                     Surface
DIS ADVANTAGE OF BIOFILM
   In industrial environments, biofilms can develop on the
    interiors of pipes, which can lead to clogging and corrosion. Biofilms
    on floors and counters can make sanitation difficult in food
    preparation areas
 Infiltration of wast water
 Oil industrial
 Attache to hulk and reduce speed of ship
 Envelope Destruction such as color and epoxi
  envelopes
 Reduce of heat transfer in condenser
 Biological erosion
 Biofoulding
 Reduce speed of water in pipe
BIOFILMS AND INFECTIOUS DISEASES

   Biofilms have been found to be involved in a wide
    variety of microbial infections in the body, by one
    estimate 80% of all infections.Infectious processes
    in which biofilms have been implicated include
    common problems such as urinary tract infections,
    catheter infections, middle-ear infections, formation
    of dental plaque --gingivitis, coating contact lenses,
    and less common but more lethal processes such
    as endocarditis, infections in cystic fibrosis, and
    infections of permanent indwelling devices such as
    joint prostheses and heart valves-chronic sinusitis-
DENTAL PLAQUE

 Dental plaque is the material that adheres to the
 teeth and consists of bacterial cells (mainly
 Streptococcus mutans and Streptococcus sanguis),
 salivary polymers and bacterial extracellular
 products. Plaque is a biofilm on the surfaces of the
 teeth. This accumulation of microorganisms subject
 the teeth and gingival tissues to high concentrations
 of bacterial metabolites which results in dental
 disease
ROLES FOR FLAGELLAR STATORS IN BIOFILM
FORMATION BY PSEUDOMONAS AEROGINOSA

 pseudomonas
  aeroginosa caused
  sistic fibrosis It has
  only a single flagellum
 Genom of bacteria
  encode 2 flagellar
  stators called MotAB or
  the MotCD
 Flagellar stator
  produce energy
MYCOBACTERIUM AVIUM
 A biofilm-forming
  opportunistic human
  pathogen found in the
  environment
 Several strains have
  been isolated from
  AIDS patients and
  others with
  compromised immune
  systems
PAST WORK
 Past work on genes associated with biofilm
  formation in M. avium has yielded approximately 12
  genes that are up-regulated in biofilm formation.
 Several of these genes are important for
  glycopeptidolipid (GPL) biosynthesis, while others
  play a key role in fatty acid metabolism or the citric
  acid cycle.
CURRENT WORK
   Using primers for previously identified genes
    associated with biofilm formation, quantify gene
    expression M. avium strains MAC A5, MAC 101,
    and MAC 104 in the presence and absence of three
    different antibiotics at sub-inhibitory concentrations
    using Real-Time PCR.
GENES / GENE PRODUCTS
Biofilm genes
 Glycosyl Transferase, essential for the expression
  of mature GPLs.
 GuaB2 (IMP dehydrogenase), catalyzes the first
  reaction in GMP biosynthesis
 PmmB (Phosphomannose mutase), converts D-
  Mannose 1-Phosphate TO D-Mannose 6-
  Phosphate
Control gene
 16S RNA, not involved in biofilm formation, to be
  used as a control
TIME COURSE OF BIOFILM FORMATION BY
STAPHYLOCOCCUS AUREUS AND STAPHYLOCOCCUS
EPIDERMIDIS MASTITIS ISOLATES

 Isolation mastitis staphylococcal
 Loci gene

 Isolatin with fluorescent in situ hybridisation(FISH)

 TSB

 Fixation by paraformaldehyde

 Change in pentration by ethanol,lysostaphin

 Hybridation with 16SrRNA

 Probes :sta and sau
EFFECT OF SHEAR STRESS ON GROWTH ,ADHESION AND BIOFILM
FORMATION OF PSEUDOMONAS AEROGINOSA WITH
ANTIBIOTIC_INDUCED MORPHLOGICAL CHANGES


 Effect of morphological changes and stress on
  growth and attachment biofilm formation
 pseudomonas aeroginosa ATCC27853

 Microtitre plate assay

 Effect of .5 piperacillin tazobactam imipenem
  meropenem
 Orbital shaking 250rpm

 Decrease of attachment and biofilm information

 Use of sub_mic antibiotic(minimal inhibitory
  concentration)
BILE SALTS ENHANCE BACTERIAL CO-AGGREGATION,
BACTERIAL-INTESTINAL EPITHELIAL CELL
ADHESION,BIOFILM FORMATION AND ANTIMICROBIAL
RESISTANCE OF BACTEROIDES FRAGILIS

 bacteroides fragilis
  anearobic bacteria
 Bile as detergenet

 Bile salt hydrolase
  enzyme
 bacteroides fragilis
  NCTC9343
TRYPANOSOMA CRUZI: ULTRA STRUCTUREAL
STUDIES OF ADHESION,LYSIS AND BIOFILM
FORMATION BY SERRATIA MARCESCENS

 trypanosoma cruzi
  cused chagas disease
 Serratia SM365

 Attach by D_MANOSE

 Serratia DB11
STRUCTURE AND FUNCTION OF ESHERICHIA COLI
PROTEIN YMGB:APROTEIN CRITICAL FOR BIOFILM
FORMATION AND ACID RESISTANCE

 YMgB gene in biofilm
 In richment culture of
  glocose ,decrease
  motolity of cellular
 Resistance to acid
REFERENCES
   1-Christian m,toutain(2007) Roles of flagellar stators in
    biofilm formation by pseudomonas aeruginosa
 2-Staffan kjellberg and micheal givskov(2007) the
  biofilm mode of life :mechanism and adaptations
 3-M.oliveira s.f nunes (2007)Time course of biofilm
  formation by staphylococcus aureus and
  staphylococcus epidermidis mastitis isolates
 4-Daniele p.castro Sergio H.seabra (2007)
  trypanosoma cruzi: ultra structureal studies of
  adhesion,lysis and biofilm formation by serratia
  marcescens
   5-Lilian pumbwe christopher A.skilbeck(2007) bile salts
    enhance bacterial co-aggregation, bacterial-intestinal
    epithelial cell adhesion,biofilm formation and
    antimicrobial resistance of bacteroides fragilis
   6-Kristina mojica danielle Elsey (2007) Quantitative
    analysis of biofilm EPS uronic acid content
   7-A.P.FONSECA J.C SOUSA(2007) effect of shear
    stress on growth ,adhesion and biofilm formation of
    pseudomonas aeroginosa with antibiotic_induced
    morphlogical changes
   8-JINTAE LEE Rebecca pag (2007)
   9-Structure and function of esherichia coli protein
    YmgB:Aprotein critical for biofilm formation and acid
    resistance
Biofilm information in pathogen bacteria

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Biofilm information in pathogen bacteria

  • 1. IN THE NAME OF GOD
  • 2. BIOFILM FORMATION IN PATHOGENS BACTERIA Profosser: Dr.ahmad ali PORBABAEI by: ABBAS MOROVVATI
  • 3. CONTENT  Definition of Biofilm  History  Biofilm information  Biofilm information in vitro  Where biofilms are found?  Advantage of biofilm  Biofilm and antibiotic resistance  Dis advantage of biofilm  Biofilms and infectious diseases  References
  • 4. DEFINITION OF BIOFILM  Biofilm: An aggregate of microbes with a distinct architecture. A biofilm is like a tiny city in which microbial cells, each only a micrometer or two long, form towers that can be hundreds of micrometers high. The "streets" between the towers are really fluid-filled channels that bring in nutrients, oxygen and other necessities for live biofilm communities.  The biofilm is held together and protected by a matrix of excreted polymeric compounds called EPS .This matrix protects the cells within it and facilitates communication among them through biochemical signals  Attachment by pili and etc
  • 5. BIOFILM  Biofilms consist of: 1-microbes 2-EPS(poly sacarid--glycoprotein) 3-water(85-98%)  Biofilm can contain many different types of microorganism, e.g. bacteria, archaea, protozoa, fungi and algae; each group performing specialized metabolic functions.  Fungal biofilms also frequently contaminate medical devices. They cause chronic vaginal infections and lead to life-threatening systemic infections in people with hobbled immune systems
  • 6. HISTORY  Van lion Hook  zobel  1913 Sohngen  1970 enviromental places  1971 Marshal  1973 charakis  1976 koshreton
  • 8. HOW IS BIOFILM FORMED? Bacteria continue to grow, outer cells provide a physical barrier to protect inner cells surface Planktonic bacteria, in the presence of water, attach to a surface.
  • 9. BIOFILM INFORMATION IN VITRO  1_batch culture model  2_cotinuous culture model  NOTE  Study population of archie bacteria use of coenzyme F420 and use of OR NAD and ATP production and so  MONOCULTURE:THIS BIOFILM FORMED BY ONE TYPE OF BACTERIA
  • 10. . WHERE BIOFILMS ARE FOUND  . 1. on solid substratums in contact with moisture, 2. on soft tissue surfaces in living organisms 3. at liquid-air interfaces.  4-Biofilms form on the surface of catheter lines and contact lenses. They grow on pacemakers, heart valve replacements, artificial joints and other surgical implants  5-Biofilms grow in hot, acidic pools in Yellowstone National Park (USA)  6-In industrial environments, biofilms can develop on the interiors of pipes
  • 12. ADVANTAGE OF BIOFILM  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.  One benefit of this environment is increased resistance to detergents  Biofilms can help eliminate petroleum oil from contaminated oceans or marine systems.  Bioreactors for water infiltration
  • 13. BIOFILM AND ANTIBIOTIC RESISTANCE  Little or no effect because the bacteria in biofilm are in a different  Bacteriostatic phase than most Antibiotics antibiotics target Surface
  • 14. DIS ADVANTAGE OF BIOFILM  In industrial environments, biofilms can develop on the interiors of pipes, which can lead to clogging and corrosion. Biofilms on floors and counters can make sanitation difficult in food preparation areas  Infiltration of wast water  Oil industrial  Attache to hulk and reduce speed of ship  Envelope Destruction such as color and epoxi envelopes  Reduce of heat transfer in condenser  Biological erosion  Biofoulding  Reduce speed of water in pipe
  • 15. BIOFILMS AND INFECTIOUS DISEASES  Biofilms have been found to be involved in a wide variety of microbial infections in the body, by one estimate 80% of all infections.Infectious processes in which biofilms have been implicated include common problems such as urinary tract infections, catheter infections, middle-ear infections, formation of dental plaque --gingivitis, coating contact lenses, and less common but more lethal processes such as endocarditis, infections in cystic fibrosis, and infections of permanent indwelling devices such as joint prostheses and heart valves-chronic sinusitis-
  • 16. DENTAL PLAQUE Dental plaque is the material that adheres to the teeth and consists of bacterial cells (mainly Streptococcus mutans and Streptococcus sanguis), salivary polymers and bacterial extracellular products. Plaque is a biofilm on the surfaces of the teeth. This accumulation of microorganisms subject the teeth and gingival tissues to high concentrations of bacterial metabolites which results in dental disease
  • 17. ROLES FOR FLAGELLAR STATORS IN BIOFILM FORMATION BY PSEUDOMONAS AEROGINOSA  pseudomonas aeroginosa caused sistic fibrosis It has only a single flagellum  Genom of bacteria encode 2 flagellar stators called MotAB or the MotCD  Flagellar stator produce energy
  • 18. MYCOBACTERIUM AVIUM  A biofilm-forming opportunistic human pathogen found in the environment  Several strains have been isolated from AIDS patients and others with compromised immune systems
  • 19. PAST WORK  Past work on genes associated with biofilm formation in M. avium has yielded approximately 12 genes that are up-regulated in biofilm formation.  Several of these genes are important for glycopeptidolipid (GPL) biosynthesis, while others play a key role in fatty acid metabolism or the citric acid cycle.
  • 20. CURRENT WORK  Using primers for previously identified genes associated with biofilm formation, quantify gene expression M. avium strains MAC A5, MAC 101, and MAC 104 in the presence and absence of three different antibiotics at sub-inhibitory concentrations using Real-Time PCR.
  • 21. GENES / GENE PRODUCTS Biofilm genes  Glycosyl Transferase, essential for the expression of mature GPLs.  GuaB2 (IMP dehydrogenase), catalyzes the first reaction in GMP biosynthesis  PmmB (Phosphomannose mutase), converts D- Mannose 1-Phosphate TO D-Mannose 6- Phosphate Control gene  16S RNA, not involved in biofilm formation, to be used as a control
  • 22. TIME COURSE OF BIOFILM FORMATION BY STAPHYLOCOCCUS AUREUS AND STAPHYLOCOCCUS EPIDERMIDIS MASTITIS ISOLATES  Isolation mastitis staphylococcal  Loci gene  Isolatin with fluorescent in situ hybridisation(FISH)  TSB  Fixation by paraformaldehyde  Change in pentration by ethanol,lysostaphin  Hybridation with 16SrRNA  Probes :sta and sau
  • 23. EFFECT OF SHEAR STRESS ON GROWTH ,ADHESION AND BIOFILM FORMATION OF PSEUDOMONAS AEROGINOSA WITH ANTIBIOTIC_INDUCED MORPHLOGICAL CHANGES  Effect of morphological changes and stress on growth and attachment biofilm formation  pseudomonas aeroginosa ATCC27853  Microtitre plate assay  Effect of .5 piperacillin tazobactam imipenem meropenem  Orbital shaking 250rpm  Decrease of attachment and biofilm information  Use of sub_mic antibiotic(minimal inhibitory concentration)
  • 24. BILE SALTS ENHANCE BACTERIAL CO-AGGREGATION, BACTERIAL-INTESTINAL EPITHELIAL CELL ADHESION,BIOFILM FORMATION AND ANTIMICROBIAL RESISTANCE OF BACTEROIDES FRAGILIS  bacteroides fragilis anearobic bacteria  Bile as detergenet  Bile salt hydrolase enzyme  bacteroides fragilis NCTC9343
  • 25. TRYPANOSOMA CRUZI: ULTRA STRUCTUREAL STUDIES OF ADHESION,LYSIS AND BIOFILM FORMATION BY SERRATIA MARCESCENS  trypanosoma cruzi cused chagas disease  Serratia SM365  Attach by D_MANOSE  Serratia DB11
  • 26. STRUCTURE AND FUNCTION OF ESHERICHIA COLI PROTEIN YMGB:APROTEIN CRITICAL FOR BIOFILM FORMATION AND ACID RESISTANCE  YMgB gene in biofilm  In richment culture of glocose ,decrease motolity of cellular  Resistance to acid
  • 27. REFERENCES  1-Christian m,toutain(2007) Roles of flagellar stators in biofilm formation by pseudomonas aeruginosa  2-Staffan kjellberg and micheal givskov(2007) the biofilm mode of life :mechanism and adaptations  3-M.oliveira s.f nunes (2007)Time course of biofilm formation by staphylococcus aureus and staphylococcus epidermidis mastitis isolates  4-Daniele p.castro Sergio H.seabra (2007) trypanosoma cruzi: ultra structureal studies of adhesion,lysis and biofilm formation by serratia marcescens
  • 28. 5-Lilian pumbwe christopher A.skilbeck(2007) bile salts enhance bacterial co-aggregation, bacterial-intestinal epithelial cell adhesion,biofilm formation and antimicrobial resistance of bacteroides fragilis  6-Kristina mojica danielle Elsey (2007) Quantitative analysis of biofilm EPS uronic acid content  7-A.P.FONSECA J.C SOUSA(2007) effect of shear stress on growth ,adhesion and biofilm formation of pseudomonas aeroginosa with antibiotic_induced morphlogical changes  8-JINTAE LEE Rebecca pag (2007)  9-Structure and function of esherichia coli protein YmgB:Aprotein critical for biofilm formation and acid resistance