6_Adhesion_271123.pdf

Mondays @14:00-16:00 Introduction to the biology of human pathogenic bacteria 1. Microbes & Humans 1. 2. 3. 4. 5. 2. Koch & Defining Pathogens 1. 2. 3. 3. The microbiota – background Skin microbiota Oral and respiratory tract microbiota Genitourinary tract and gut microbiota Early life microbiota de...

Mondays @14:00-16:00 Introduction to the biology of human pathogenic bacteria 1. Microbes & Humans 1. 2. 3. 4. 5. 2. Koch & Defining Pathogens 1. 2. 3. 3. The microbiota – background Skin microbiota Oral and respiratory tract microbiota Genitourinary tract and gut microbiota Early life microbiota development Koch & some history (+ Anthrax) Koch’s postulates Molecular Koch’s postulates Pathogenesis & Virulence 1. 2. 3. 4. Virulence & ‘measuring’ virulence Opportunistic vs. primary pathogens Stages of pathogenesis Acquisition of virulence/pathogenic factors 1 Mondays @14:00-16:00 Introduction to the biology of human pathogenic bacteria 4. Host Defence Systems 1. 2. 3. 4. 5. 5. Natural/physical barriers Innate immune responses Phagocytosis & complement Adaptive immune responses Microbiota-associated anti-infection responses Defence Systems of Pathogens 1. 2. 3. 4. 5. Avoidance strategies – background Evasion of AMPs and IgA Prevention of phagocytosis and complement Molecular mimicry Antigenic/phase variation 2 Mondays @14:00-16:00 Introduction to the biology of human pathogenic bacteria 6. Adhesion 6. 7. 8. 9. 7. Invasion & Intracellular Growth 1. 2. 3. 4. 8. Significance of bacterial adhesion Pili Adhesins (MSCRAMMS) Other adhesins The intracellular lifestyle Invasion Intracellular survival & cell-to-cell spread Obligate intracellular pathogens Bacterial Toxins 1. 2. 3. 4. Types of toxins (endo vs. exotoxins) Endotoxins (LPS) Exotoxin and modes of action Examples of important bacterial toxins 3 27th November 2023 Introduction to the biology of human pathogenic bacteria Adhesion Prof. Dr. Romana Gerner Chair of Clinical Microbiome 4 Adhesion (1) Successful establishment of infection by bacterial pathogens requires adhesion to host cells: specific ligand-receptor interaction → then colonization of tissues In certain cases also cellular invasion (covered in next lecture!) Followed by (intracellular) multiplication, dissemination to other tissues, and/or persistence Complexity of the bacterial tools used for cell adhesion ranges from single monomeric proteins to more complex multimeric macromolecules 5 Importance of adhesion (1) One of the body's innate immune defenses is the ability to physically remove bacteria from the body via constant shedding of surface epithelial cells from the skin and mucous membranes Removal of bacteria by such means as coughing, sneezing, vomiting, and diarrhea Bacterial removal by bodily fluids such as saliva, blood, mucous, and urine Adhesion types Adhesins are binding proteins, usually on tips of long fibrillary organelles called fimbriae or pili, that are anchored to the cell wall Other important factors involved in bacterial adhesion to host cells include capsules and flagella, and other cell wall-associated molecules Many adhesion factors recognize specific proteins (receptors) on the host cell Adhesins are critical to biofilm formation Biofilms are multicellular communities that are 1000-fold more resistant to antibiotics and immune defense than are planktonic (swimming or drifting) bacteria 7 Pili (fimbriae) for host cell adherence Pili enable some organisms to adhere to receptors on target host cells and thus colonize and resist flushing by the body Pili are thin protein tubes originating from the cytoplasmic membrane and are found in virtually all Gramnegative bacteria, but not in many Gram-positive bacteria 8 Pili (1) Pilus has a shaft composed of the protein pilin Shaft end has an adhesive tip structure shape corresponds to specific glycoprotein or glycolipid receptors on host cells Once attached to host cells, pili can depolymerize and enable adhesions in the bacterial cell wall to make more intimate contact Also evidence that binding of pili to host cell receptors can serve as a trigger for activating the synthesis of additional cell wall adhesins 9 Pili (2) Bacteria are constantly losing and reforming pili They may switch types of adhesive tips of the pili in order to adhere to different cell types and evade immune defenses e.g., E. coli is able to make over 30 different types of pili Some pathogenic bacteria that use pili to adhere to host cells include Neisseria gonorrhoeae Escherichia coli & Salmonella Vibrio cholerae Pseudomonas aeruginosa 10 E. coli and pili E. coli can be a pathogenic and/or a commensal bacterium There are multiple pathotypes (i.e. enteropathogenic E. coli) Different E. coli pathotypes can produce pili that enable adherence to different host cells/tissues e.g., Uropathogenic strains of E. coli can attach to the urinary epithelium (via pili) and cause urinary tract infections also produce afimbrial adhesins (see subsequent slides) for attachment to epithelial cells E. coli in the urinary tract causes more than 85 percent of all UTIs. e.g. Enteropathogenic E. coli (EPEC) use pili to adhere to intestinal epithelial cells Terlizzi et al. Front. Microbiol., 15 August 2017 | https://doi.org/10.3389/fmicb.2017.01566 11 UPEC (Uropathogenic E. coli) and pili In UPEC, the fim operon encodes type 1 pili expresses a hemagglutinin which recognizes mannose The pap operon encodes P- or Pap-pili (pyelonephritis-associated pili) interacts with the di-galactoside unit in the P-blood group antigen In UPEC clinical isolates, the fim operon is constitutive, whereas pap is part of a pathogenicity island that is also responsible for other virulence determinants Mutants lacking type 1 pili or its associated adhesin, FimH, are quickly cleared from the bladder emphasizes importance of type 1 pili mediated hostpathogen interactions 12 UPEC pili (2) Both types of pili are heteropolymeric → Major pilus protein subunit provides pilus stalk → Several minor subunit proteins at distal end → PapG and FimH represent the actual adhesins → Chaperone-usher (CU) pathway assembles pili More than 1,000 copies of the FimA major pilin form the type 1–pilus rod Distal end of pilus tip contains FimH adhesin followed by single copies of the FimG and FimF adaptor subunits Mannosylated proteins that are present on the bladder epithelium bind to FimH Pap (P) pilus Type 1 pilus P pilus is composed of 6 different subunits arranged into two distinct subassemblies (tip fibrillum and pilus rod) At distal end, tip fibrillum is composed of 1x PapG adhesin followed by PapF and PapE subunits Pilus rod is made by >1,000 copies of PapA subunit Adaptor subunit PapK connects above subunits to the PapA rod (superhelical structure at base of the pilum)13 Type IV Pili (1) One class of pili, known as type IV pili, allows attachment to cells, food and fibers and enables twitching motility Located at poles of cell and allows for a gliding motility along a solid surface such as a host cell Extension and retraction of these pili allows the bacterium to drag itself along the solid surface Other functions include biofilm formation and molecule exchange (e.g. DNA) Bacteria with type IV pili include P. aeruginosa, N. gonorrhoeae, Neisseria meningitidis, and V. cholerae 14 Type IV pili (2) These pili are strong, flexible rod-like filaments of 5–8 nm in diameter and 1–2 μm in length Pilin subunits are assembled through interactions between their conserved Nterminal α-helices → forms a hydrophobic core in the filament that is believed to provide extreme mechanical strength Type IV pili bind to the glycolipids asialoGM1 and asialo-GM2 on epithelial cell surfaces In Pseudomonas and Neisseria, type IV pili are believed to be a major virulenceassociated adhesin 15 Adhesins Adhesins are surface proteins found in the cell wall of various bacteria that bind to specific receptor molecules on the surface of host cells → enable bacterium to adhere intimately to that cell in order to colonize and resist physical removal Many, if not most bacteria use one or more adhesins to colonize host cells 16 MSCRAMMS (1) Adhesins: MSCRAMMS – microbial surface components recognising adhesive matrix proteins Cell surface proteins on Staphylococcus aureus called MSCRAMMs bind host proteins may aid adherence to host tissues, and serum-coated plastic provides a critical step to establish infection important in Gram+ve’s MSCRAMMs share a similar structure, with two adjacent IgG-like folded subdomains mediating their attachment to components of the host ECM such as collagen, fibrinogen, or fibronectin (Fn) 17 MSCRAMMS (2) The molecular mechanisms by which MSCRAMMs binds their ligands include ‘dock lock latch’ ‘collagen hug’ tandem β-zipper Very tight binding Deletion of these MSCRAMMs strongly attenuates the ability of S. aureus to colonize tissues, multiply, and cause disease (sepsis) 18 Capsules as adhesins Many bacteria produce a capsular polysaccharide matrix or glycocalyx to form a biofilm on host tissue Many chronic and difficult-to-treat infections are caused by bacteria in biofilms Biofilm of S. aureus on indwelling catheter 19 Flagella as adhesins (1) Many Gram-positive and Gramnegative bacterial species have a flagellum ‘whip’ in Latin Flagellum is primarily a motility organelle that enables movement and chemotaxis However, flagella have also been reported to function as adhesins 20 Flagella as adhesins (2) In several bacterial species, the flagellum is multifunctional and plays a role in motility and is an adhesive organelle; H7 flagellum in E. coli Mediates binding to intestinal epithelial cells Pseudomonas aeruginosa Facilitates binding to mucus Clostridioides difficile Flagella needed for adherence and colonization of the intestinal epithelium during infection 21 Other adhesins: E. coli (1) E. coli O157 (an EPEC strain) utilizes a type 3 secretion system (T3SS) to inject effector proteins into intestinal epithelial cells Some of these cause polymerization of actin at the cell surface → pushes host cell cytoplasmic membrane up to form a pedestal Another effector protein inserts into the membrane of the pedestal to serve as a receptor molecule for E. coli adhesins 500nm Wales et al., J Med Microbiol 2002 22 Other adhesins: E. coli (2) EPEC is a human pathogen that causes acute and chronic pediatric diarrhea Hallmark of EPEC infection is formation of attaching and effacing (A/E) lesions in the intestinal epithelium Formation of A/E lesions is mediated by genes located on the pathogenicity island locus of enterocyte effacement (LEE) encodes the adhesin intimin T3SS six effectors including essential translocated intimin receptor (Tir) → interaction between Tir and intimin anchors the bacterium to host membrane 23 24 Other adhesins: Helicobacter pylori H. pylori use a type 4 secretion system (T4SS) to inject effector proteins into stomach epithelial cells → induces host cells to display more receptors on their surface for H. pylori adhesins Don’t worry you don’t need to know all the components of the H. pylori T4SS! CagA H. pylori causes chronic gastritis, peptic ulcers, and gastric cancer in humans 25 Other adhesins: Bordetella pertussis Filamentous hemagglutinin is an adhesin that allows the bacterium to adhere to galactose residues of glycolipids on respiratory tract ciliated epithelial cells Pertussis toxin also functions as an adhesion One subunit of pertussis toxin remains bound to the bacterial cell wall while another subunit binds to glycolipids on membrane of ciliated epithelial cells of respiratory tract Pertussis also produces an adhesin called pertactin that further enables the bacterium to adhere to cells B. pertussis causes whooping cough 26 Other adhesins: Neisseria gonorrhoeae Neisseria gonorrhoeae produces an adhesin called Opa (protein II) that enables the bacterium to make a more intimate contact with host cell after it first adheres with its pili Like with adhesive tips of pili, N. gonorrhoeae has multiple alleles for Opa protein adhesins enabling the bacterium to adhere to a variety of host cell types N. gonorrhoeae is a bacterial pathogen responsible for gonorrhoea marker for neutrophils 27 Other pathogens with adhesins Tip of the spirochete Borrelia burgdorferi contains adhesins that can bind to fibronectin, laminin etc. of various host cells Causative agent of Lyme disease Treponema pallidum (also spirochete) contains adhesins that are able to bind to fibronectin on epithelial cells Causes syphilis Streptococcus pyogenes (group A beta streptococci) produce a number of adhesins Protein F that binds to fibronectin, a common protein on epithelial cells able to adhere to lymphatics and mucous membranes of the upper respiratory tract and cause streptococcal pharyngitis (strep throat) Lipoteichoic acid binds to fibronectin on epithelial cells M-protein also functions as an adhesin 28 Learning Objectives (1) Describe why adhesion is such an important part of the infection process List the different bacterial factors that are involved in adhesion Explain the term pili (fimbriae) and pili-associated host cell adhesion principles/processes Provide examples of bacterial pathogens that use pili for adherence Describe UPEC-associated pili and how this pathogen uses pili to adhere to host urinary epithelium Briefly describe type IV pili and their role in host cell adherence 29 Learning Objectives (2) Briefly describe how bacteria use adhesins to bind to host cells Explain what is meant by the term MSCRAMMS and their general structure List the 3 molecular mechanisms used by MSCRAMMS to bind their host cell ligands Describe how capsules are involved in adhesion and biofilm formation Provide examples of bacterial pathogens that use flagella for adherence Understand how EPEC uses the LEE to facilitate host cell adherence 30 Learning Objectives (3) Describe how H. pylori uses T4SS for adherence Describe the process of cell attachment by B. pertussis Describe how the adhesin Opa enhances binding of N. gonorrhoeae to host cells Provide additional examples of bacterial pathogens that use adhesins (spirochetes and S. pyogenes) 31