Medical Bacteriology MBIO 460 Lecture 11 Dr. Turki Dawoud 2 nd Semester 1436/1437 H

Virulence Factors and Toxins  Extracellular capsules and slime layers, cell wall and envelope material, and fimbriae and pili are all integral macromolec-ular components of microorganisms that may aid in pathogenesis and act as virulence factors.  However, in addition to these, a number of microbial intracellular and extracellular components function solely as virulence factors. These specialized virulence factors are produced by different pathogens, but many share molecular characteristics and modes of action. Here we examine some key factors

Virulence Factors Many pathogens produce toxins, which we discuss in the following three sections. Many other virulence factors are enzymes that enhance pathogen colonization and growth. For example, streptococci, staphylococci, and certain clostridia produce hyaluronidase (Table 28.4), an enzyme that promotes spreading of organisms in tissues by breaking down the polysaccharide hyaluronic acid, a material that functions in animals as an intercellular cement. Hyaluronidase digests the intercellular matrix, enabling these organisms to spread from an initial infection site. Streptococci and staphylococci also produce proteases, nucleases, and lipases that degrade host proteins, nucleic acids, and lipids. Similarly, clostridia that cause gas gangrene produce collagenase, or κ-toxin (Table 28.4), which breaks down the tissue-supporting collagen network, enabling these organisms to spread through the body.

Fibrin, Clots, and Virulence Fibrin is a protein that functions in the clotting of blood, and thus fibrin clots are often formed by the host at a site of microbial invasion. The clotting mechanism, triggered by tissue injury, isolates the pathogens, limiting infection to a local region. Some pathogens counter this process by producing fibrinolytic enzymes that dissolve the clots and make further invasion possible. One fibrinolytic substance produced by Streptococcus pyogenes is called streptokinase (Table 28.4). By contrast, other pathogens produce enzymes that promote the formation of fibrin clots. These clots localize and protect the organism. The best-studied fibrin-clotting enzyme is coagulase (Table 28.4), produced by pathogenic Staphylococcus aureus. Coagulase causes fibrin to be deposited on S. aureus cells, protecting them from attack by host cells. The fibrin matrix produced as a result of coagulase activity probably accounts for the extremely localized nature of many staphylococcal infections, as in boils and pimples (Figure 34.24). Coagulase-positive S. aureus strains are typically more virulent than coagulase-negative strains.

Toxins 1.Exotoxins 2.Enterotoxin 3.Endotoxin

Exotoxins Exotoxins are toxic proteins released from the pathogen cell as it grows. These toxins travel from a site of infection and cause damage at distant sites. Table 28.4 provides a summary of the properties and actions of some of the known exotoxins as well as other extracellular virulence factors. Exotoxins fall into one of three categories: the cytolytic toxins, the AB toxins, and the super-antigen toxins. The cytolytic toxins work by degrading cytoplasmic membrane integrity, causing lysis. The AB toxins consist of two subunits, A and B. The B component generally binds to a host cell surface receptor, allowing the transfer of the A subunit across the targeted cell membrane, where it damages the cell. The superantigens work by stimulating large numbers of immune cells, resulting in extensive inflammation and tissue damage, as we will discuss later (Section 30.8)

Cytolytic Toxins Various pathogens produce proteins that damage the host cytoplasmic membrane, causing cell lysis and death. Because the activity of these toxins is most easily observed with assays involving the lysis of red blood cells (erythrocytes), the toxins are called hemolysins (Table 28.4). However, they also lyse cells other than erythrocytes. The production of hemolysin is demonstrated in the laboratory by streaking the pathogen on a blood agar plate (a rich medium containing 5% whole blood). During growth of the colonies, hemolysin is released and lyses the surrounding red blood cells, releasing hemoglobin and creating a clearing, called hemolysis (Figure 28.18).

Figure Hemolysis. (a) Zones of hemolysis around colonies of Streptococcus pyogenes growing on a blood agar plate. (b)Action of lecithinase, a phospholipase, around colonies of Clostridium perfringens growing on an agar medium containing egg yolk, a source of lecithin. Lecithinase dissolves the membranes of red blood cells, producing the cloudy zones of hemolysis around each colony

Some hemolysins attack the phospholipid of the host cytoplasmic membrane. Because the phospholipid lecithin (phosphatidylcholine) is often used as a substrate, these enzymes are called lecithinases or phospholipases. An example is the α-toxin of Clostridium perfringens, a lecithinase that dissolves membrane lipids, resulting in cell lysis (Table 28.4, Figure 28.18b; Figure 21.41). Because the cytoplasmic membranes of all organisms contain phospholipids, phospholipases sometimes destroy bacterial as well as animal cytoplasmic membranes. Some hemolysins, however, are not phospholipases. Streptolysin O, a hemolysin produced by streptococci, affects the sterols of the host cytoplasmic membrane. Leukocidins (Table 28.4) lyse white blood cells and may decrease host resistance (Section 29.2).

Staphylococcal α-toxin (Figure and Table 28.4) kills nucleated cells and lyses erythrocytes. Toxin subunits first bind to the lipid bilayer. The subunits then oligomerize into nonlytic heptamers, now associated with the membrane. Following oligomerization, each heptamer undergoes conformational changes to produce a membrane-spanning pore, releasing the cell contents and allowing influx of extracellular material, disrupting cell function and causing cell death.

Protein Synthesis Inhibitor Toxins Several pathogens produce AB exotoxins that inhibit protein synthesis. Diphtheria toxin, produced by Corynebacterium diphtheriae, is a good example of an AB toxin and is an important virulence factor. Rats and mice are relatively resistant to diphtheria toxin, but human, rabbit, guinea pig, and bird cells are very susceptible, with only a single toxin molecule required to kill each cell. Diphtheria toxin is secreted by cells of C. diphtheriae as a single polypeptide. Fragment B specifically binds to a host cell receptor (Figure 28.20). After binding, proteolytic cleavage between fragment A and B allows entry of fragment A into the host cytoplasm. Here fragment A disrupts protein synthesis by blocking transfer of an amino acid from a tRNA to the growing polypeptide chain (Section 7.15). The toxin specifically inactivates elongation factor 2, a protein involved in growth of the polypeptide chain, by catalyzing the attachment of adenosine diphosphate (ADP) ribose from NAD +. Following ADP-ribosylation, the activity of the modified elongation factor 2 decreases dramatically and protein synthesis stops

Diphtheria toxin is formed only by strains of C. diphtheriae that are lysogenized by a bacteriophage called phage β; the tox gene in the phage genome encodes the toxin. Non-toxigenic, non-pathogenic strains of C. diphtheriae can be converted to pathogenic strains by infection with phage β (this is a process called phage conversion, Section 10.10). Exotoxin A of Pseudomonas aeruginosa functions similarly to diphtheria toxin, also modifying elongation factor 2 by ADP- ribosylation (Table 28.4)

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