Normal Flora Vs Pathogens

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Normal Flora

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10 to 13 human cells and 10 to 14th bacterial cells
Definition
 microorganisms that colonize the body but do not normally produce diseases
Locations
 parts of the body that are exposed or communicate with the outside environment
Semi-permanent
 acquired during and shortly after birth
 varies continuously throughout life
Three types of symbiosis
 Commensalism
 Beneficial to one partner while the other is unaffected
 Mutualism
 Interactions between two organisms in which both organisms benefit
 Parasitism
 Beneficial to one partner while harmful to the other

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Some important microorganisms of the normal flora and our interaction with these organisms.

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 Skin: few residents in dry areas; large population in moister places
 Mouth & nose: high microbial density
 Teeth: plaque = biofilm of bacteria
 Lungs: normally quite sterile
 Urethra: lightly colonized
 Vagina: extensive flora; age-related changes (lactobacillus)

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What are the advantages and disadvantages of the normal flora.

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Advantage of Normal Flora:
• Antigenic stimulation Æ helps development of immune system
• Preventing colonization by pathogens through microbial antagonism (i.e. competitive exclusion) by
o competing with pathogens for nutrients & space
o producing toxic factors or environmental conditions harmful to pathogens
• Other:
o production of vitamins B and K by gut flora

Disadvantage of normal flora:
• Harmful when overgrown or misplaced due to use of broad-spectrum antibiotics or when fecal flora are transferred to urinary tract

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What are nosocomial infections and some of the major microorganisms causing nosocomial infections.

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 Hospital-acquired infections (≠ community acquired infections)
 Hospitals = major reservoir of opportunistic pathogens and antibiotic-resistant pathogens
 Compromised status of host, results from:
 invasive procedures (surgery, anesthesia)
 suppressed immune systems (drugs, radiations)

 Transmission routes
 person-to-person
 fomite (i.e. through non-living objects: e.g. needles, respiratory aids…)

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1. Infection
2. Etiologic Agent
3. Pathogenesis
4. Predisposition factors

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Infection
 colonization of body by pathogenic microbes
 may exist without symptoms of a disease
Etiologic agent
 the cause of disease
Pathogenesis
 how the disease develops
Predisposition factors
 make the host susceptible to a disease
 e.g. gender, genetic background, geographical location, age, life style, immune status etc…

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What bacterial pathogens are and the difference between opportunistic and primary pathogens.

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Strict pathogens
 always associated with human diseases
 e.g. Mycobacterium tuberculosis (tuberculosis), Neisseria gonorrhoeae (gonorrhea)…
Opportunistic pathogens
 take advantage of preexisting conditions that enhance susceptibility of patient to cause a disease or a more serious disease
 many of them are members of the patient’s normal microbial flora – e.g. Pseudomonas aeruginosa, Candida albicans

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What are the mechanisms of bacterial pathogenesis

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 Enter the environments (us)
 Remain in the niche – colonization
 Gain access to food sources
 Escape clearance by host protective responses
 Problem Æ cause damage to human host

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What are the portals of entry and exit?

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 Natural openings
 Respiratory tract – inhalation (e.g. Mycobacterium tuberculosis, S. pneumoniae)
 Gastrointestinal tract – ingestion (e.g. Salmonella, E. coli)
 Genitourinary tract – sexual transmission (e.g. Neisseria gonorrhoeae, Chlamydia trachomatis)
 Skin
 trauma (e.g. Clostridium tetani)
 arthropod bite (e.g. Rickettsia, Yersinia pestis)
 Exit from the host
 transmission to a fresh host
 shed in large numbers in secretions and excretions
 available in blood for uptake, e.g. blood-sucking arthropods or needles

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What determines host suseptability?

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Organism factors overcome host factors!!!
Host Factors: Immune system
Organism Factors: Dose and Virulence (ability to cause disease by host colonization, evation of host defenses and damage=disease)

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What are the factors involved in colonization?

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 Adhesion
 to epithelial or endothelial cells of bladder, intestine, respiratory tract, blood vessels…
 major mechanisms
 pili
 adhesin proteins
 teichoic acid
 biofilm

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Invasion

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After adhesion, invasive micro-organisms have the ability to penetrate the target tissue
 Some invasive bacteria use normal phagocytic entry mechanisms to gain access
 Other invasive bacteria can enter host cells which are not naturally phagocytic, by
 specific attachment to the host cell
 production of surface proteins called invasins
 induction of local rearrangements of the cytoskeleton ð formation of pseudopod-like structures which engulf the pathogen into the host cell

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Legionnaire’s Disease pathogenesis

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1) Attaches to alveolar macrophages Æ coiling phagocytosis
2) Prevents phagosome-lysosome fusion
3) Replication inside the phagosome
4) Lysis of phagocytes
 Hydrolytic enzymes
Æ Lung damage & Inflammatory response

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Salmonella enteric pathogenesis

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1. Adherence to mucosal surface induce changes in enterocyte or M cell Æ triggers surface ruffles
2. Ruffles = plasma membrane sites with filamentous actin cytoskeletal rearrangement
3. Uptake of organisms within host cell membrane-bounded vesicles

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Evasion

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 Bacteria have developed ways to avoid the major antibacterial defenses by inactivating or evading the host system and antibody
 evading recognition and killing by phagocytes
 growing inside cells to hide from these host responses
Ways to evade complement or antibody response:
 Intracellular growth (e.g. Mycobacteria, Chlamydia)
 Antigenic variation, mimicry or masking (e.g. Neisseria gonorrhoeae)
 Inactivation of antibody or complement (e.g. IgA-specific protease)
 Capsule and Biofilm - prevents antibody and complement from reaching the bacteria
 Protection of the membrane from attack (e.g. LPS O antigen prevents; M protein in S. pyogenes)
Ways to escape phagocytosis:
 Preventing contact with phagocyte (e.g. capsule)
 Preventing opsonization
 Production of enzymes capable of lysing phagocytes
 streptolysin produced by S. pyogenes
 α-toxin produced by C. perfringens
Ways to evade phagocytic killing: Inhibition of phagolysosome fusion; Resistance to lysosome products; Escape from phaogosome and propagation into cytoplasma

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Mechanisms of Host Damage

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1. By-products of bacterial growth – tissue destruction
 acids or gas from fermentation
2. Degradative enzymes
 break down host tissues & facilitate development of infection and spread into the tissue
 phospholipase, collagenase, protease, hyaluronidase...
3. Toxins
 directly harm tissue or trigger destructive biologic activities
 endotoxins: cell wall components freed when cells die and break up.
 exotoxins: soluble proteins released by living bacterial cells
4. By inducing hypersensitivity reactions

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Endotoxins

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• Endotoxins: cell wall components freed when cells die and break up
o gram(-) lipopolysaccharide = endotoxin
o in gram(+): teichoic and lipoteichoic acids – endotoxin-like effects
o peptidoglycan and its breakdown product
• Lipopolysaccharide (LPS) activates almost every immune mechanisms as well as the clotting pathway
• Low concentrations
Æ fever, vasodilatation, and activation of immune and inflammatory responses
• High concentrations
o Æ excessive response – shock & death

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Exotoxins

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• Proteins released by living bacterial cells
• Local effect or effect at sites distant from the site of infection
• Often receptor-binding proteins that alter a function and/or kill the cell
• Toxin genes often on plasmid or lysogenic phage
• Cytolysins: enzymatic lysis and pore forming

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How do intracellular-acting toxins access their substrates with the host cells?

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• AB toxins
• released into the host environment
• may act locally or diffuse to act distally
• B subunit = binding component; interacts with receptors on the cell membrane & promotes entry
• A subunit = active component
• Targets: ribosomes, transport mechanisms, intracellular signaling (cAMP production, G protein function)
• Direct injection of toxins through flagella or pilus into host cells by Type III or IV secretion systems, respectively
• Finally, Bordetella adenylate cyclase toxins directly enter the cytosol from the plasma membrane

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Examples of intracellular-acting toxins

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1. Diphtheria toxin
2. Cholera toxin
3. Neurotoxins

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Diphtheria toxin

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• Inhibition of protein synthesis
• Inactivation of elongation factor-2 Æinactivates transfer of amino acids from tRNA to the polypeptide chain during translation of mRNA by ribosomes
• Infection of mucosa of oropharynx
• Multiplication of bacteria and production of toxin
• Exotoxin activity Æ death of surrounding cells
• È
• Inflammatory reaction leads to collection of a gray exudate which evolves into thick pseudomembrane
• This coating adheres strongly to mucosa and may extend into larynx/trachea ð obstruction and suffocation

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Cholera toxin

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• Hyperactivation
• A1 portion ADP-ribosylates a regulatory molecules involved in production of cAMP Æ increase in intracellular cAMP

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Neurotoxins

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• Both are AB toxins
• Tetanus toxin:
• blocks release of inhibitory neurotransmitters Æ unopposed firing of the motor neurons Æ constant contraction
• Botulinum toxin:
• blocks release of acetylcholine from peripheral nerves Æ preventing contraction Æ flaccid paralysis

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Superantigens

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• Bind to the external regions of the T-cell receptor and the major histocompatibility complex class II (MHC II) on macrophage
• nonspecific interact
• Æ massive release of cytokines
• E.g. S. aureus TSST-1 (Toxic Shock Syndrome Toxin)

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S. aureus and Toxic Shock Syndrome

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• Often associated with use of tampons, but can result from focal infections at any body site by toxin-producing S. aureus
• The organism does not disseminate; the toxin does and is responsible for the clinical features*****
• Sudden onset: fever, vomiting, diarrhea
• Then: red rash resembling a sunburn and desquamation 1-2 wks later
• Hypotensive shock, impaired renal and hepatic functions, occasional deaths
• 3% of all cases are fatal

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Important Bacterial Virulence Mechanisms

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1. Colonization
• Adherence
• Biofilm
2. Damage
• By-products of growth (gas, acid)
• Endo/Exotoxins
• Degradative enzymes
• Cytotoxic proteins
• Superantigens
3. Evading host defence
• Evasion of phagocytic and immune clearance
• Capsule / Biofilm
• Intracellular growth
4. Resistance to antibiotics

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Classify infectious diseases

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• Epidemic diseases: Infectious diseases that attacks many people at the same time in the same geographical area.
• Endemic diseases: Diseases that constantly present in a local population, e.g. Malaria
• Pandemic diseases: Worldwide occurrence, e.g. plague, AIDS

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Koch’s Postulates

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• Purpose: to establish the etiology (cause) of an infectious disease.
• Experimental procedures:
• Same pathogen must be present in every disease case.
• The pathogen is isolated from the diseased host and grown in pure culture.
• The pathogen from the pure culture is inoculated into a susceptible animal and must be shown to reproduce the same disease.
• The pathogen must be isolated from the animal and be shown to be the same as the original organism.
• Exceptions to Koch's Postulates
• Pure culture cannot be obtained using artificial media, e.g. Treponema pallidum (syphilis).
• No experimental animal (so Step 3 and 4 cannot be carried out).