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		Quiz Me!	Parasitism. Mechanisms of Pathogenicity
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Last revised: Monday, April 24, 2000
Ch. 29 (p. 581-591) in Prescott et al, Microbiology, 4th Ed.

Note: These notes are provided as a guide to topics the instructor hopes to cover during lecture. Actual coverage will always differ somewhat from what is printed here. These notes are not a substitute for the actual lecture!

Copyright 2000. Thomas M. Terry

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Pathogens

• World Health Organization (WHO) in Geneva maintains records on health statistics, infection rates, epidemics in most of World.
• View WHO information by individual disease categories

  Class Exercise: Identify major causes of Mortality (deaths) and Burden (acutely diseased individuals) from most recent WHO statistics (from class handout).

• Pathogens are parasitic organisms with specialized adaptations that allow them to interact directly or indirectly with hosts so they can grow, reproduce, and spread. In the process they cause host damage ranging from mild annoyance to death.
• To understand pathogens, we must understand how they can overcome host obstacles (including the presence of other microbes already present), and how they can cause disease through specific virulence factors.

Terminology

• Disease is a process (verb), not a state (noun)
• Every disease is a race between pathogen trying to gain a foothold and host defenses trying to prevent pathogen. Many factors involved: virulence and numbers of pathogen, health and age of host, etc.
• Wrong to equate "one pathogen" = disease. Much more complex.
• Disease = state of being not in good health (not at ease, "dis-ease").
• Parasite often used to refer to protozoans or worms; the term "Pathogen" is typically used when referring to bacteria, virus, or fungus that causes disease. All are parasitic.
• Pathogen = organism with potential to cause disease
• Infection = pathogen is growing in or on host
• Virulence = degree or intensity of pathogenicity
• Invasiveness = ability of pathogen to spread to other tissues in body
• Infectivity = ability of pathogen to establish infection
• Toxigenicity = ability of pathogen to secrete toxins
• Septicemia = infection in which pathogen grows massively in the body, being found in blood and throughout organs. Usually leads to death.

Intracellular vs Extracellular Life: choices & consequences

Most pathogens have evolved to live either inside or outside of host cells, rarely if ever in both habitats.

Intracellular life

• Poses special problems for host; can't easily attack pathogen without harming its own tissues. Many pathogens are adapted for intracellular life, including all viruses, certain bacteria (e.g. TB, plague),
• Since white blood cells (macrophages, lymphocytes) are major components of defense system, many successful pathogens target these cells specifically for intracellular growth.
• Problem: to be successful, pathogen at some point must leave cells, exit host. Best chance to prevent infection is sometime during exit -- transmission -- entry to new host, before it has a chance to hide in new cells.
• Some intracellular parasites are so highly evolved that they can't survive at all outside their host's cells. Ex: Chlamydia, Rickettsia. To be successful, these must rely on mechanisms such as sexual contact or animal bites to transmit them to new hosts.

Bacterial Disease Case Study: Tuberculosis and Mycobacterium tuberculosis

M. tuberculosis is a strictly aerobic bacterium, with a very slow doubling time (12-18 hours) View electron micrograph of M. tuberculosis Mycobacteria have very waxy coats (made of mycolic acid compounds). They resist destaining with acid and alcohol, and are called the acid-fast bacteria. Their waxy coats resist uptake of many antibiotics. TB has a long latent period; first detectable signs of antibody response are 8-12 weeks after infection. TB is usually asymptomatic; only 10-20% of infected persons become diseased. How does M. tuberculosis cause disease? Almost any tissue can become infected, but lung is common focus of infection, so consider TB in lung: Bacterium is taken up inside phagosome by macrophage (first stage of phagocytosis) Bacterium secretes proteins that block fusion of phagosome with lysosome Bacterium slowly grows and replicates inside macrophage host cell, gradually turning cell into a bacterial replicator. Macrophages migrate to regional lymph nodes Other macrophages are attracted by chemotaxis to the site of infection, forming an early tubercule. Activated macrophages kill M. tuberculosis, stop spread. Host develops cell-mediated immunity, delayed hypersensitivity to tuberculin, a protein secreted by M. tuberculosis. In some cases, mature tubercules are formed as firm outer layer of macrophages "wall off" inner mass of infected macrophages. Center of tubercule liquiefies, forms air-filled cavity that promotes growth of bacteria, liquefaction of tubercule contents. Rupture of tubercule can allow bacteria to enter bronchioles, spread throug respiratory system and other tissues Patients with pulmonary TB have respiratory problems, cough up mucus secretions frequently. TB can attack many other sites in body as well as lungs. View cross-section of normal lung View cross-section of lung with TB View cross-section of lung with "miliary" TB (so called because of extensive white abcesses resembling grains of millet) TB is one of the most common diseases world-wide. The following annual estimates are probably too low: Worldwide annual deaths from TB: 3 million (98% in developing countries) Worldwide annual reported disease cases: 8 million Worldwide incidence of infection: somewhere between 1 in 10 to 1 in 3 people U.S. incidence in 1995: 22,860 cases (8.7 cases per 100,000 population) View further information about TB Want to play doctor? Practice your diagnostic skill with a Web-based case study of a patient with TB. Since you started reading this page people have been infected with TB.

Extracellular life

• Pathogen must deal with host's defensive strategies: white blood cells, immune system, etc.
• But does provide greater opportunities for grown, reproduction, and spreading than living inside cells.
• Can rapidly colonize a habitat; Ex. when cholera invades intestine, can quickly multiply, spread to cover large surface area
• Typical bacterial pathogens that act extracellularly: E. coli, Pseudomonas sp., Vibrio cholera

Facultative intracellular pathogens

• Some bacteria can grow either inside host cells or outside, depending on circumstances.
• Examples: Shigella, Salmonella sp.,

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Virulence Factors

• Virulence Factors are specific adaptations that allow pathogen to:
  1. attach selectively to host tissues
  2. gain access to nutrients by invading or destroying host tissues
  3. avoid host defenses
• Many examples of virulence factors:
  1. Specific attachment & entry factors
     • Pathogen must be able to bind to some receptor molecule on cell surfaces. These typically have necessary functions for cell.
     • Most diseases are tissue specific, because only certain tissues have receptor molecule needed. Ex: HIV binds to cells that have CD4 receptor (only certain lymphocytes).
     • Fimbriae or pili are used by some bacteria to attach selectively to certain tissues. Ex: Neisseria gonorrhaea binds to genital epithelium by fimbriae. In mutant cells w/o fimbriae, infectivity and pathogenicity are lost.
  2. Invasive enzymes
     • Many bacteria have specific enzymes that allow cells to penetrate host tissues
     • Example 1: collagenase produced by Clostridium perfringens. Enzyme degrades collagen, the primary structural fiber of connective tissue (25% of body's protein), allows penetration deeper into tissues ---> fulminating gangrene. Strictly anaerobic process, only occurs when tissue is damaged so blood can't supply oxygen (e.g. serious wounds, frostbite).
     • Example 2: hemolysin enzymes produced by Streptococcus pyogenes dissolves cell membranes of tissues, produces typical symptoms of "strep throat".
  3. Tricks to avoid host defenses.

     Phagocytosis:

     Phagocytosis is one of the most important functions the nonspecific host defenses, and is carried out by most white blood cells except lymphocytes, including polymorphonuclear leukocytes (neutrophils) and mononuclear phagocytes (monocytes, macrophages). View time-lapse movie of phagocytosis (watch the yeast cell coated with red dye be taken into the cell) Phagocytosis involves a series of steps: Contact between phagocyte and microbial cell Engulfment and invagination Formation of a phagosome Fusion of phagosome with a lysosome (--> phagolysosome) Hydrolytic enzymes can destroy most organic molecules. Additionally, "oxygen burst" takes up oxygen, not for respiration but to produce toxic oxidizing agents (hydrogen peroxide and superoxide) inside phagolysosome.

     1. Capsules
        • Many pathogens have thick extracellular polysaccharide capsules. Capsules inhibit phagocytosis, prevent quick disposal of bacterium by WBCs. Loss of capsule typically causes loss of infectivity.
     2. "Nasty Enzymes"
        1. Leukocidins
           • Some pathogens secrete chemicals that specifically kill WBCs.
           • Ex: Staphylococcus aureus. Accumulation of pus at infected site is caused by dead WBCs.
        2. Coagulase
           • Staph. aureus produces enzyme that coagulates blood.
           • Result: WBCs, other body defenses can't reach site of infection. Staph typically remains localized, "walled off" from defenses, produces many nasty types of localized infections such as boils, abcesses, etc.
             
     3. Exotoxins
        • Most exotoxins are proteins, secreted from cell, often damaging tissues at some distance. Very potent, small amounts are very toxic.
        • Often coded by plasmid DNA (ex. E. coli) or lysogenic phage DNA (ex. botulism, diphtheria)
        • Almost always inactivated by heat. Most are good antigens when inactive, can make toxoids (antigens without poison activity) = strong immune response.
        • Ex. 1: Diphtheria toxin. From Corynebacterium diphtheriae. Enters cell, inactivates elongation factor needed for protein synthesis. Result = cell gradually loses ability to make proteins (same toxin molecule keeps inactivating more and more factors), shuts down.
        • Ex. 2: Botulin toxin, a neurotoxin (attacks nervous system). FromClostridium botulinum, anerobic soil bacterium, endospore former. Most potent toxin known --- 1 gram could kill 10 million people. Toxin interferes with synaptic transmission at nerve-muscle junctions ---> flaccid paralysis. Occurs most typically in improperly canned
          • View movie showing botulin toxin activity (requires free Shockwave plug-in)
        • Ex. 3: Tetanus toxin, another neurotoxin. FromClostridium tetanus, anerobic soil bacterium, endospore former. Blocks synaptic transmission to inhibitory neurons needed to relax one muscle when the other in paired muscle contracts (e.g. biceps must relax when triceps contracts, vice versa), leads to rigid paralysis. Common from deep wounds, pulled teeth. (But in 20% of cases, no history of injury!). Kills about 1 million infants/year by infecting umbilical stump. Treatment: antitoxin. Prevention: toxoid immunization (lasts 5-10 yrs).
        • Ex. 4: Cholera toxin = an enterotoxin (attacks enteric tract). From Vibrio cholerae. Binds to receptors on intestinal cells, chemically alters molecule involved in c-AMP production, leaves cAMP stuck in the "on" position. Causes massive outflow of water (chasing outflow of Na+/Cl-). Similar mode of action for other enterotoxin. Epidemic in S. America currently. Pathogen is free-living in fresh water, only causes infection in humans. Can be spread by drinking water, food (shellfish common). Untreated, mortality is ~50%. With fluid replacement, <1%. Prevention: clean drinking water.
          • Visit cholera (Vibrio cholerae) web page from Bacteriology 330, by Kenneth Todar, University of Wisconsin Department of Bacteriology
     4. Endotoxins
        • Endotoxins are integral parts of Gram-negative outer membrane (= LPS, lipolysaccharide). Unlike Exotoxins, they are typically heat resistant, active only in sizable amounts, and remain bound to cells.
        • Mechanism of action is very diverse. "When we sense LPS, we are likely to turn on every defence at our disposal" (Lewis Thomas), including fever, decrease in iron, inflammation, blood clotting, reduced sugar in blood, etc. Most important clinical problems are fever and shock.
        • Typical scenario: Gram-negative bacteria (e.g. E. coli, Pseudomonas) enter body via clinical procedure (improperly sterilized kidney dialysis tubing, catheter, etc.), cause sudden decrease in blood pressure (hypotension) = "septic shock". Can be lethal.
     5. Siderophores
        • Iron plays special role in control of infection. Most bacteria require iron to synthesize cytochromes. Iron in human body is tightly bound, either in hemoglobin (blood cells), on transferrin proteins (serum and lymph), or lactoferrin (milk, tears, saliva, mucus, etc.).
        • Some bacteria (Streptococci) do not require iron -- metabolism is strictly fermentative, no respiratory system. But most bacteria have to find way to get iron or cannot grow.
        • Siderophores = iron-binding factors that allow some bacteria to compete with the host for iron. Ex: Enterochelin produced by enteric bacteria (E. coli, Salmonella). Mutants that cannot synthesize enterochelin lose virulence. These mutants can regain virulence if pure enterochelin is injected along with mutant bacteria.

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