Bacteriology Microbiology

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Microbial Mechanisms of Pathogenicity

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Microbial Mechanisms of Pathogenicity
In this chapter we will take a look at some of the specific properties
of microorganisms that contribute to:
– Pathogenicity: The ability of microorganism to cause disease by
overcoming the defense of the host (refer to humans).
– Virulence: The degree or extent of pathogenicity
• To cause disease, most pathogens must:
– gain access to host,
– adhere to host tissues,
– penetrate or evade host defenses,
– and damage the host tissues.
• However, some microbes do not cause diseases by directly damaging
host tissue, instead disease is due to the accumulation of microbial
waste products.
• Also, some microbes, such as those that cause dental caries, can
cause disease without penetrating the body (Streptococcus mutans)

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Mucous membranes, Skin, Parenteral route (direct deposition beneath
the skin or membranes).
1. Mucous membranes: lining the respiratory tract, gastrointestinal tract,
genitourinary tract, and conjuctiva a delicate membrane that covers
the eye balls.
• Respiratory tract (RT): is the easiest and frequent. Microbes are
inhaled into the nose or mouth in drops of moisture and dust particles.
– Diseases that are commonly contracted via the respiratory tract include the
common cold, pneumonia, tuberculosis, influenza, measles, and small pox.
• Gastrointestinal (GI) tract: microorganisms can gain access to GI in
food and water and via contaminated fingers. Most microbes that
enter the body in these ways are destroyed by hydrochloric acid (HCL)
and enzymes in the stomach, or by bile and enzymes in the small
intestine. Those that survive can cause disease.
– Microbes in the GI tract can cause hepatitis A, typhoid fever, amoebic
dysentry, giardiasis, shigellosis, and cholera (These pathogens are then
eliminated with feces and can be transmitted to other host via contaminated
water, food, and fingers).
• Genitourinary tract: is a portal of entry for pathogens that are
contracted sexually .
– Some microbes can cause sexually transmitted diseases, for example, HIV,
syphilis, chlamydia, gonorrhea.
Portals of Entry

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Treponema pallidum : syphilis
amoebic dysentry

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2. Skin: Unbroken skin is impenetrable by most microorganisms. Some
microbes gain access to the body through opening in the skin, such as
hair follicles, and sweat glands ducts.
– In addition, larvae of the hookworm actually bore through intact skin, and
some fungi grow on the keratin in skin or infect skin itself.
3. Parenteral route: other microorganisms can access to the body when
they are deposited directly into the tissue beneath the skin or into
mucous membranes when these barriers are penetrated or injured.
– For example, punctures, injections, bites, cuts, wounds, surgery all establish
parenteral routes.
Portals of Entry

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The virulence of a microbe is often expressed as the ID50:
Infectious dose for 50% of a sample population.
Numbers of Invading Microbes
Bacillus anthracis
Portal of entry ID50
Skin (cutaneous anthrax) 10-50 endospores
Inhalation 10,000-20,000 endospores
Ingestion 250,000-1,000,000 endospores
• These data show that cutaneous anthrax is significantly easier to
acquire that either inhalation or ingestion.
• A study or V. cholerae showed that the ID50 is 108 cells.

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• Bacillus anthracis, gram-positive, endospore-forming aerobic rod
• Found in soil and water
• Cattle are routinely vaccinated
• Treated with ciprofloxacin or doxycycline
Anthrax affects human in three forms:
• Cutaneous anthrax
– Endospores enter through minor cut
– 20% mortality
Anthrax
• Gastrointestinal anthrax
– Ingestion of undercooked
food contaminated food
– 50% mortality
• Inhalational anthrax
– Inhalation of endospores
– 100% mortality

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The potency of a toxin is often expressed as the LD50: Lethal
dose of a toxin for 50% of a sample population.
Toxin LD50
Botulinum toxin 0.03ng/kg
Shiga toxin 250 ng/kg
Staphylococcal enterotoxin 1350ng/kg
• These data show that a much smaller dose of botulinum toxin is needed
to cause symptoms. .

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• The attachment between the pathogen and the host is accomplished by
means of surface molecules on the pathogen called adhesins or ligands that
bind specifically to receptors on the cells of certain host tissue. The
majority of adhesins on the microorganisms studied are glycoproteins or
lipoproteins. The receptors on host cells are typically sugars, mannose.
• Adhesins may be located on a microbe’s glycocalyx or on other microbial
surface structures, such as pili, fimbriae, and flagella.
– Streptococcus mutans (tooth decay)
• Glycocalyx
– Escherichia coli (gastrointestinal disease)
• Fimbriae
– Treponema pallidium (cause syphilis)
• Used its tapered end as a hook to attach to the host cell.
– Lesteria monocytogenes (causes meningitis, spontaneous abortion,
stillbirths)
• produces an adhesin for a specific receptor on host cells.
– Neisseria gonorrhoeae (grows inside human epithelial cells and
leukocytes).
• Fimbriae containing adhesins and an outer membrane protein (Opa protein).
Adherence

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– Streptococcus pyogenes
• M protein (heat resistant and acid resistant protein). The M
protein mediates the attachment of the bacterium to epithelial
cells of the host and helps the bacterium resist phagocytosis by
white blood cells.
– Neisseria gonorrhoeae (grows inside human epithelial cells
and leukocytes).
• Fimbriae and an outer membrane protein (Opa protein).
– The waxes that make up the cell wall of M. tuberculosis also
increase virulence by resisting digestion by phagocytes.
• In fact, M. tuberculosis can even multiply inside phagocytosis.
Certain bacteria contain chemical substances that contribute to
virulence.

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– Coagulase (produced by some member of genus Staphylococcus)
• Coagulate the fibrinogen in blood. Fibrinogen is a plasma protein
produced by the liver, is converted by coagulases into fibrin, the thread
that form a blood clot.
– Kinases (Streptococcus pyogenes)
• Digest fibrin clots (successfully used to dissolve some types of blood
clots in cases of heart attacks due to obstructed coronary arteries).
– Hyaluronidase (Streptococci and clostridium spp.)
• Hydrolyzes hyaluronic acid, a type of polysaccharide that holds together
certain cells of the body, particularly cells of connective tissue and help
the microorganism spread from its initial site of infection.
• This digestion action is thought to be involved in the tissue blackening
of infected wounds.
– Collagenase (Clostridium spp.)
• Hydrolyzes collagen which forms the connective tissue of muscles and
other body organs and tissues. (facilitate the spread of gas gangrene).
– IgA proteases (N. gonnorrhoeae and N. meningitides)
• Destroy IgA antibodies (a class of antibodies that produced to defend
against adherence of pathogens to mucosal surfaces)
The virulence of some bacteria is thought to be aided by the production
of extracellular enzymes (exoenzymes and related substance)

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• In the presence of antigen the body produces proteins called
antibodies, which bind to the antigens and inactivate and destroy
them.
• However, some pathogens can alter their surface antigens, by a
process called antigenic variation. Thus by the time the body
mounts an immune response against a pathogen, the pathogen has
already altered its antigens and is unaffected by the antibodies.
• Some microbes can activate alternative genes resulting in
antigenic changes. For example,
• N. gonorrhoeae has several copies of the Opa-coding gene, resulting in
cells with different antigens and in cells that express different
antigens overtime.
Example of other bacteria and protozoa that are capable of antigenic
variation:
• Influenzavirus, the causative agent of influenza (flu)
• N. gonorrhoeae, the causative agent of gonorrhea
• Trypanosma brucei gambiense, the causative agent of African
trypanosomiasis (sleeping sickness).
The virulence of some bacteria is thought to be aided by the
Antigenic variation (Alter surface proteins)

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Penetration into the Host Cell cytoskeleton
Salmonella entering epithelial
cells.
A major component of cytoskeleton is a
protein called actin, which is used by some
microbes to penetrate host cells and by
others to move through and between host
cells.
• Salmonella strains and E. coli contact with
the host cell plasma membrane and lead to
changes in the membrane at the point of
contact. The microbes produce surface
proteins called invasins that rearrange
nearby actin filaments of the cytoskeleton.
This cause cytoplasmic structures to
project from the host cell like a pedestal
under the Salmonella.
• Certain bacteria such as Shigella species
and Listeria species can actually use actin
to propel themselves through the host cell
cytoplasm and from one host to another.

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Sexually-transmitted diseases (STDs):
disease causative agent reported cases/yr
In USA
bacterial
gonorrhea Neisseria gonorrhoeae 350,000; true incidence
much higher
chlamydial
infections
Chlamydia trachomatis 527,000; true incidence
much higher
syphilis Treponema pallidum 51,600
chancroid Haemophilus ducreyi 356; true incidence much
higher
viral
genital herpes
simplex
Herpes simplex virus (HSV) Around 30 million infected
papillomavirus
infections
human papilloma virus (HPV) Around 40 million infected
AIDS human immunodeficiency
virus (HIV)
60,860

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Syphilis:
Treponema pallidum: motile spirochete; cannot be grown in vitro;; strictly
human pathogen
transmission: direct person-person contact
T. pallidum in testis

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stage symptoms pathogenesis
initial contact,
incubation period
(2-10 wks)
multiplication at infection
site
primary syphilis
(1-3 months)
primary chancre; enlarged nodes
(groin), spontaneous healing
(within 4-6 wks)
[silent phase (~2-10 wks)]
proliferation in regional
lymph nodes and blood
secondary syphilis
(weeks-months)
flu-like (muscle ache, headache,
fever), rash,
multiplication; lesions in
lymph nodes, joints, muscle,
skin, mucous membranes, liver
latent syphilis
(3-30 yrs)
none T. pallidum dormant
→ eventual new cell growth
tertiary syphilis neurological (paralysis, insanity)
cardiovascular (aortic lesions,
rupture; stroke)
progressive destructive disease
further dissemination,
invasion,
gumma formation (skin, bone,
joints, testes)
progression of syphilis:

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general paresis:
Altered personality, emotional instability, delusions, memory loss,
impaired judgement, abnormalities of eyes, speech defects
congenital syphilis:
• fetus susceptible after 4th month of gestation.
• risk greatest if mother has primary, secondary syphilis (less if latent).
~40% of affected fetuses miscarried or stillborn
• neonates may develop secondary syphilis within few weeks.
• characteristic deformities (face, teeth) may appear in early
childhood.
treatment:
• penicillin; most effective for primary, secondary syphilis.
• no known antibiotic resistance problems.
syphilis screening:
• used to involve series of serological tests, potentially multiple blood
samplings.

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Gonorrhea:
Neisseria gonorrhoeae: gram negative diplococci, nonmotile, fastidious; most
strains susceptible to cold, drying; humans
transmission: direct person-person contact
symptoms: may be mild, absent (especially in females)
• incubation period: 2-7 days
male: urethritis, painful urination, thick, pus-containing penile discharge
female: increased vaginal discharge, painful urination, abdominal pain,
menstrual abnormalities .
pathogenesis:
- attachment to epithelia: urethra, cervix, pharynx, conjunctiva
- avoidance of phagocytosis & host immune response (due to hypervariable
surface antigens)
- capable of invading of host cells
treatment:
penicillin resistance is a problem, especially in Asia
cephalosporins, fluoroquinones, azithromycin
vaccine - much effort, little success (hypervariability of surface proteins,
intracellular mode)

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Chlamydia trachomatis
• obligate intracellular parasite; infection mimics gonorrhea in
several ways
symptoms: 7-14 d post-exposure
male: thin, grey-white penile discharge, testicular pain
female: increased vaginal discharge, may be painful urination, vaginal
bleeding, abdominal pain
inclusion conjunctivitis: - eye infection in neonates
tissue damage:
• attaches to sperm (like N. gonorrheae)
• testicular damage, fallopian tube damage may lead to sterility
treatment: early use of azithromycin, tetracycline, erythromycin will
prevent serious complications
• condom use is effective in prevention

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How bacterial pathogens damage host cells?
• If the pathogen overcomes the host defense, the microorganism can
damage host cells in four ways:
• By using the host’s nutrients
• By causing direct damage in the immediate vicinity of the invasion
• By producing toxins, transported by blood and lymph
• By inducing hypersensitivity reactions

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Using the host’s nutrients
• Iron is required for the growth of most pathogenic bacteria.
• The concentration of free iron in human body is fairly low because iron in
human body is tightly bound to iron-transport proteins, such as
lactoferrin, transferrin, and ferritin, as well as hemoglobin;
• In order to obtain free iron, some pathogens produce protein called
siderophores.
• When iron is needed by a pathogen, siderophores are released into the
medium where they take the iron away from iron-transport proteins. Once
the iron-siderophore complex is formed, it is taken up by a siderophores
receptors on the bacterial surface. Then the iron is brought into the
bacterium.
• Some pathogens have receptors that bind directly to iron-transport
proteins and hemoglobin.

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Direct Damage
• Once pathogen attach to host cells, they can cause direct damage as the
pathogens use the host cell for nutrients and produce waste products.
• As pathogen metabolize and multiply in cells, the cells usually rupture and
the pathogens spread to other tissue. (Many viruses, intercellular
bacteria, protozoa that grow in host cells are released when the host cell
rupture).
• Some bacteria such as E. coli, Shigella, Sallmonella, and Nisseria
gonorrhoeae, can induce host epithelial cells to engulf them by a process
that resembles phagocytosis. These pathogen can disrupt host cells.
• Other bacteria can penetrate host cells by excreting enzymes and by
their motility; such penetration can itself damage the host cell.

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The production of toxins
• Toxins: Are poisonous substances that are produced by certain
microorganisms and that contribute to pathogenicity.
• Toxigenicity: Ability of microorganisms to produce a toxin
Toxins transported by the blood or lymph can cause serious, and
sometimes fatal effects. Some toxins produce fever, cardiovascular
disturbances, diarrhea, and shock.
Toxins can also inhibit protein synthesis, destroy blood cells and blood
vessels, and disrupt the nervous system by causing spasms.
• Toxemia: Presence of toxins in the host's blood
• Antitoxin: Antibodies against a specific toxin (Antibodies produced by the
body provide immunity to exotoxins).
• Toxoid: Inactivated toxin used in a vaccine. (When exotoxins inactivated
by heat or by formaldehyde, iodine, or other chemicals, they no longer
cause the disease but can still stimulate the body to produce antitoxins).

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Endotoxins:
are lipopolysacchrides
Toxins are of two general types, based on their position relative to the
microbial cell: Exotoxins and endotoxins.
Exotoxins:
are proteins

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Exotoxin
Source Mostly Gram +
Metabolic product By-products of growing cell
Chemistry Protein
Fever? No
Neutralized by antitoxin Yes
LD50 Small
The genes for most exotoxins are carried on bacterial phages or plasmids.
Found on phages, toxin genes for: Diphtheria, Botulism, Scarlet fever, Toxic
streptococci (“flesh-eating”)
Toxin genes found on plasmids: E. coli toxin causes diarrhea, S. aureus toxin causes
“scalded skin syndrome”.
• Because exotoxins are soluble in body fluid, they can easily diffuse into the
blood and are rapidly transported throughout the body.
• Exotoxins are disease-specific, because it is the exotoxins that produce
the specific signs and symptoms of the disease (not a bacterial infection).

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Exotoxins are divided into three
principle types on the basis of their
structure and function:
1. A-B toxins or type III toxins
2. Membrane-disrupting toxin or type II
toxins
3. Superantigens or type I toxins.
1. A-B toxins:
- Are consists of two parts designated A
and B, both of which are polypeptides.
- Most exotoxins are A-B toxin.
An example of A-B toxin is the
diphtheria toxin, which illustrated in the
figure.
Exotoxins

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2. Membrane-disrupting toxins or type II toxins:
– Causes Lysis of host cells by disrupting their plasma membrane:
• Some do this by making protein channels in the plasma
membrane (e.g., leukocidins, hemolysins). Example,
Staphylococcus aureus .
• Others disrupting phospholipid bilayer. Example, Clostridium
perfringens.
– Contribute to virulence by killing host cells especially phagocytes.
Membrane-disrupting toxins that kill phagocytic leukocytes (WBC)
are called leukocidins.
• Most leukocidins are produced by staphylococci, streptococci, and
pneumococci.
Membrane-disrupting toxins that destroy erythrocytes (RBC) are
called hemolysins.
• Important producers of hemolysins include staphylococci and
streptococci.
Hemolysins produced by streptococci are called streptolysins.
Cont’d Exotoxins

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3. Superantigens or type I toxins:
– They are bacterial protein
– Superantigen stimulate the proliferation of immune cells called T
cells. These cells are types of WBC that act against foreign
organisms and tissues and regulate the activation and proliferation of
other cells of the immune system.
– In resposne to superantigens, T cells are stimulated to release
enormous amounts of chemical called cytokines from host cells.
Cytokines are small protein hormone that stimulate or inhibit many
normal function.
– The excessively high levels of cytokines released by the T cells
enters the bloodstream and give rise to a number of symptoms,
including fever, nausea, vomiting, diarrhea, and sometimes shock, and
even death.
– Bacterial superantigens include staphylococcal toxins that cause food
poising and toxic shock syndrome.
Exotoxins

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Exotoxins are named on the basis of several characteristics.
One is the type of the host cell that is attached:
– Neurotoxins: attack nerve cells
– Hepatotoxins: attack liver cells
– Cardiotoxins: attack heart cells
– Leukotoxins: attack leukocytes
– Enterotoxins: attack the lining of the gastrointestinal tract
– Cytotoxins : attack a wide variety of cell
Some exotoxins are named for the disease with which they are associated:
– Diphtheria toxins: cause of diphtheria
– Tetanus toxins: cause of tetanus
Other exotoxins are named for the specific bacterium that produces them:
– Botulinum toxin: Clostridium botulinum
– Vibrio entertoxin: Vibrio cholerae

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Exotoxins
Species Exotoxin
Corynebacterium diphtheriae
A-B toxin. Inhibits protein
synthesis.
Streptococcus pyogenes Membrane-disrupting. Erythrogenic.
Clostridium botulinum A-B toxin. Neurotoxin
C. tetani A-B toxin. Neurotoxin
Vibrio cholerae A-B toxin. Enterotoxin
Staphylococcus aureus Superantigen. Enterotoxin.

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Endotoxins
• Endotoxins are part of the outer portion of the cell wall of Gram –ve
bacteria.
• The lipid portion of the lipopolysaccharides, called lipid A, is the
endotoxins. Thus, endotoxins are lipopolysaccharides, whereas exotoxins
are proteins.
• Endotoxins are released when Gram-ve bacteria are lysed, or during
bacterial multiplication.
 Endotoxins exert their effects by stimulating macrophages to release
cytokines in very high concentrations. At these levels, cytokines are toxic.
Endotoxins give rise to a number of symptoms, including chills, fever,
weakness, generalized aches, and sometimes shock, and even death, and can
also induce miscarriage.
 Another consequence of endotoxins is the activation of blood-clotting
proteins, causing formation of small blood clots and as a result induces the
death of the tissue. This condition is referred to as disseminated
intravascular clotting.

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Endotoxins
The fever (pyrogenic response) caused by endotoxins is believed to occur as
depicted in fig below.
Bacterial cell death caused by lysis or antibiotics can also produce fever by
this mechanism.
Both aspirin and acetaminophen reduce fever by inhibiting the synthesis of
prostaglandins.

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Endotoxins
Source Gram–ve bacteria
Metabolic product Present in LPS of outer membrane
Chemistry Lipid
Fever? Yes
Neutralized by antitoxin No
LD50 Relatively large

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• Respiratory tract
– Coughing, sneezing
• Gastrointestinal tract
– Feces, saliva
• Genitourinary tract
– Urine, vaginal secretions
• Skin
• Blood
– Biting, needles/syringes
Portals of Exit

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Mechanisms of Pathogenicity