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Cocci

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I. Gram Positve Cocci

Genus staphylococci
Genus streptococci

A. Genus staphylococci
General Characteristic
Gram-positive cocci
Non-sporulating, non-motile
Grape-like clusters
Facultative anaerobe
Round colony in media
Some of them are normal flora of the skin and mucus membrane
It can produce catalase, (d/t from streptococcus)
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The genus Staphylococcus includes over 30 species and


subspecies, but only three are human pathogen:
Staphylococcus aureus
Staphylococcus epidermides
Staphylococcus saprophytics

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1. Staphylococcus aureus

Gram-positive spherical bacteria


Clusters resembling grapes
Facultative anaerobes
Catalase-positive
Oxidase-negative
Produce yellow colony in culture
Coagulase positive and
Can grow in temperature ranging from 15C to 45C

Habitat

S. aureus is normal inhabitant of human oropharynx with a carrier rate


of 15% to 25%

Transmission

Contact with skin lesions, hands of carrier


Ingestion of contaminated food with enterotoxins
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Virulence factors

Surface proteins that promote colonization of host tissues


Invasins that promote bacterial spread in tissues (leukocidin, kinases,
hyaluronidase)
Surface factors that inhibit phagocytic engulfment (capsule, Protein A)
Biochemical properties that enhance their survival in phagocytes
(carotenoids, catalase production);
Immunological mask (Protein A, coagulase, clotting factor)
Membrane-damaging toxins that lyse cell membranes (hemolysins,
leukotoxin, leukocidin)
Exotoxins that damage host tissues or otherwise provoke symptoms of
disease
Inherent and acquired resistance to antimicrobial agents
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1. Membrane damaging Toxins


1. -toxin (-hemolysin)
Most potent membrane-damaging toxin
Platelets and monocytes are particularly sensitive to toxin
Susceptible cells have a specific receptor for -toxin
which allows the toxin to bind causing small pores
through which monovalent cations can pass.
2. -toxin
Sphingomyelinase which damages membranes rich in
this lipid
The majority of human isolates of S. aureus do not
express -toxin
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3. d-toxin
Very small peptide toxin produced by most strains of S.
aureus
It is also produced by S. epidermidis.
The role of d-toxin in disease is unknown
4. Leukocidin
Is hemolytic, but less than alpha hemolysin
5. Coagulase and clumping factor
Coagulase is an extracellular protein which binds to
prothrombin in the host to form a complex called
staphylothrombin
It is reasonable to speculate that the bacteria could
protect themselves from phagocytic and immune
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defenses by causing localized clotting. 10/30/16

6. Staphylokinase
Lyses fibrin
Localized fibrinolysis might aid in bacterial spreading
7. Other extracellular enzymes
Proteases
Lipase
Deoxyribonuclease (DNase)
Fatty acid modifying enzyme (FAME).
The first three provide nutrients for the bacteria, and it is
unlikely that they have anything but a minor role in
pathogenesis
o The FAME enzyme important in abscesses
Modify anti-bacterial lipids and prolong bacterial survival
o

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Clinical Manifestation
It causes superficial skin lesions such as boils and
furunculosis
More serious infections such as:
Pneumonia
Mastitis(inflammation
Meningitis,

of the breast )

and

UTI,

and
Deep-seated infections, such as: Osteomyelitis and endocarditis

S. aureus is a major cause of hospital acquired


(nosocomial) infection of surgical wounds and infections
associated with indwelling medical devices.
Causes food poisoning by releasing enterotoxins into food,
and TSS by release of superantigens into the blood stream.
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2. Avoidance of Host Defenses


S. aureus expresses a number of factors that have the
potential to interfere with host defense mechanisms.
1. Capsular polysaccharide
The majority of clinical isolates of S.aureus express a
surface polysaccharide of either serotype 5 or 8
Although it does impede phagocytosis in the absence of
complement, it also impede colonization of damaged
heart valves, perhaps by masking adhesions.
2. Protein A
Protein A is a surface protein of S. aureus which binds
IgG molecules by their Fc region.
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The bacteria will bind IgG molecules in the wrong


orientation on their surface which disrupts opsonization
and phagocytosis.
Mutants of S. aureus lacking protein A are more
efficiently phagocytosed in vitro, and mutants in
infection models have diminished virulence.
3. Leukocidin
S. aureus can express a toxin that specifically acts on
polymorphonuclear leukocytes
Phagocytosis
is an important defense against
staphylococcal infection so leukocidin should be a
virulence factor
Leukocidin causes membrane damage to leukocytes

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4. Exotoxins
Systemic release of -toxin causes septic shock, while
enterotoxins and TSST-1 are superantigens that may cause
toxic shock.
Staphylococcal enterotoxins cause emesis (vomiting) when
ingested and the bacterium is a leading cause of food
poisoning
Exfoliatin toxin causes the scalded skin syndrome in
neonates, which results in widespread blistering and loss of
the epidermis
There are two antigenically distinct forms of the toxin, ETA and ET-B
The toxins have esterase and protease activity and
apparently target a protein which is involved in maintaining
the integrity of the epidermis
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Superantigens
S. aureus secretes two types of toxin with superantigen
activity, enterotoxins, of which there are six antigenic types
(named SE-A, B, C, D, E and G), and toxic shock syndrome
toxin (TSST-1)

Enterotoxins

Cause diarrhea and vomiting when ingested and are


responsible for Staphylococcal food poisoning

TSST-1

Is expressed systemically and is the cause of toxic shock


syndrome (TSS).
When expressed systemically, enterotoxins can also cause
toxic shock syndrome.
TSST-1 is weakly related to enterotoxins, but it does not have
emetic activity
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TSST-1 is responsible for 75% of TSS, including all


menstrual cases
Superantigens stimulate T cells non-specifically without
normal antigenic recognition
Up to one in five T cells may be activated, whereas only 1
in 10,000 are stimulated during a usual antigen presentation
Cytokines are released in large amounts, causing the
symptoms of TSS and toxic shock
Clinical Feature /diseases
Skin infections, folliculities, wound infections
Food poisoning due to entertoxins
Toxic shock syndrome

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Clinical significance

S. aureus causes
disease by infecting
tissues
typically
creating
abscesses
and/or by producing
toxins.
The
localized
host
response
to
Staphylococcal
infection
is
inflammation,
characterized
by
swelling, accumulation
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of pus, and necrosis of

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1. Localized skin infections


The most common S. aureus infections are small,
superficial abscesses involving hair follicles (folliculitis)
or sweat or sebaceous glands
Subcutaneous abscesses called furuncles (boils) often form
around foreign bodies, such as splinters.
These generally respond to local therapy, that is, removal
of the foreign body, soaking, and drainage as indicated.
Carbuncles are larger, deeper, multiloculated
skin
infections that can lead to bacteremia and require
antibiotic therapy.
Impetigo is usually a localized, superficial, spreading
crusty skin lesion generally seen in children. It can be
caused by S. aureus, although more commonly by
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Streptococcus pyogenes, or both organisms together.

2. Deep localized infections


These may be metastatic from superficial
infections or skin carriage, or may result
from trauma.
S. aureus is the most common cause of
acute and chronic infection of bone marrow.
S. aureus is also the most common cause of
acute infection of joint space in children
(septic joint). [Note: Septic joints are
medical emergencies because pus can
rapidly cause irreparable cartilage damage.
They must be treated promptly with
drainage and an antibiotic.]
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3. Acute Endocarditis
Generally associated with intravenous drug
abuse
Caused
by injection of contaminated
preparations or by needles contaminated
with S. aureus.
S. aureus also colonizes the skin around the
injection site, and if the skin is not sterilized
before injection, the bacteria can be
introduced into soft tissues and the
bloodstream, even when a sterilized needle
is used.
4. Septicemia
Is a generalized infection with sepsis
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5. Pneumonia

6. Nosocomial infections
S. aureus is one of the most common
causes of hospital-acquired infections,
often of wounds (surgical, decubital) or
bacteremia associated with catheters
7.Toxinoses

Are diseases caused by the action of a


toxin, frequently when the organism that
secreted the toxin is undetectable.
Toxinoses caused by S. aureus include:
Toxic shock syndrome
Which
results in high fever, rash
(resembling a sunburn, with diffuse
erythema followed by desquamation)
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Staphylococcal gastroenteritis
Is caused by ingestion of food contaminated
with enterotoxin-producing S. aureus.
Often contaminated by a food-handler,
these foods tend to be protein-rich and
improperly refrigerated. Symptoms, such as
NVD
Scalded skin syndrome
Involves the appearance of superficial
bullae resulting from the action of an
exfoliative
toxin
that
attacks
the
intercellular adhesive of the stratum
granulosum, causing marked epithelial
desquamation.
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22The bullae may be infected or
may result

Diseases
Clinical symptoms
Pathogenicity factors
Gastroenteritis 2-6 hours after ingesting
Enterotoxins A-E
(food poisoning) toxin: nausea, abdominal
preformed in food
- toxin ingested pain, vomiting, followed
preformed in
by diarrhea
food
Infective
Fever, malaise,
Fibrin-platelet mesh,
endocarditis leukocytosis, heart murmur
cytolytic toxins
(may be absent initially)
Abscesses/furun Subcutaneous tenderness, Coagulase, probably
cles/carbuncles redness and swelling; hot
the cytolysins
Toxic Shock
Fever, hypotension,
TSST-1
Syndrome
scarlatiniform rash which
desquamates (particularly
on palms and soles,
multiorgan failure
Impetigo
Erythematous papules to Coagulase, exfoliatins
bullae
Pneumonia
Productive pneumonia with
?, all
rapid onset, high rate of
necrosis and high fatality;
nosocomial, ventilator,
postinfluenza, IV drug
abuse, CF, CGD, etc.
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Surgical
Fever with cellulitis and/or
Coagulase,

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Resistance of Staphylococci to Antimicrobial Drugs


Hospital strains of S. aureus are usually resistant to a
variety of different antibiotics.
A few strains are resistant to all clinically useful
antibiotics except vancomycin.
But,
vancomycin-resistant strains are increasinglyreported
The term MRSA refers to Methicillin resistant
Staphylococcus aureus. It is resistant to all currently
available
-lactam
antibiotics
(penicillins,
cephalosporins)

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A plasmid associated with vancomycin resistance has


been detected;
In Enterococcus faecalis which can be transferred to S.
aureus in the laboratory
It is speculated that this transfer may occur naturally
(e.g. in the gastrointestinal tract)
Resistance is due to:
Chromosomal genes mutation followed by selection of
resistant strains, and
Acquisition of resistance genes as extrachromosomal
plasmids

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Coagulase-Negative Staphylococci
2. Staphylococcus epidermidis
Of twelve coagulase-negative Staphylococcal species

that have been recovered as normal commensals of


human skin and anterior nares, the most abundant
and important is S. epidermidis.
Despite its low virulence, it is a common cause of
hospital-acquired infections associated with the use
of implanted prosthetic devices and catheters.

3. Staphylococcus saprophyticus
The

second most important coagulase-negative


Staphylococcus
Is a frequent cause of cystitis(inflammation of
urinary bladder) in women, probably related to its
occurrence as part of normal vaginal flora.
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Sensitive to most antibiotics, even penicillin G.

Laboratory DX
Specimen- surface swabs of wound, pus,
blood sputum, CSF, Feaces
Gram stains- gram positive cocci in cluster
Culture-grow well aerobically and in CO2
enriched media at 37 oC
Colony appearance
S.aureus Golden colony
S. epidermids- White colony
S. saprophytics-white or yellow colony

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Biochemical Test

Catalase test
Staphylococici- positive
Streptococci -negative
Coagulase test
S. aureus positive, Salt tolerant; ferments
mannitol on mannitol salt agar
S. epidermidis- negative
S. saprophytics - negative
Prevention
Prevention of contact with S-aureus is almost
impossible
Treatment
Ampicillin- for penicillin sensitive staphylococci.
???? Cloxacillin, Floxacillin- for penicillin resistance
staphylococci????
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Catalse test- a positive test


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B. Streptococcus
Gram positive
Facultative anaerobic
Non spores, Non motile
Catalase negative
Chain like
Lancefield grouping
Species-specific CHO cell wall antigens
Groups designated A-H, K-V
Some are not grouped

Classification of Streptococcus based on;


Hemolytic reaction
Lance field carbohydrate group
Biochemical reaction
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2. Classification based on lance field system


Many Streptococci possess a polysaccharide
in their cell wall which is chemically and
antigenically distinct from the cell wall
polysaccharide in other bacteria in their genus
This discovered by Rebecca Lance field enable
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the use of specific antisera to identify specific

It is designated A-H and K-V


Clinically important are A,B,C,D,F,G
The main species and groups of medical

importance
1. S. pyogens ----------- Lance field group
A
2. S.agalactial ----------- Lance field group
B
3. Enterococci ----------- Lance field group
D
NB.Viridan
Streptococci
and
anaerobic
Streptococci are not grouped under Lance field
classification
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1. S. pyogen

Nonmotile
Non sporeforming
Chains
Catalase-negative
Facultative anaerobe
Gram-positive
Group A, beta hemolytic
Streptococcus
Have a capsule composed of
hyaluronic acid????

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Streptococcus
pyogenes.
Uper.
Gram
stain
of
Streptococcus pyogenes in a
clinical
specimen.
Lower.
Colonies
of
Streptococcus

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Pathogenesis
Virulance factor
M protein: fibronectin-binding protein (Protein F) and
lipoteichoic acid for adherence, inhibit phagocytosis.
Hyaluronic acid capsule: as an immunological cover
and to inhibit phagocytosis
Invasins: such as streptokinase, hyaluronidase, and
streptolysins O & S
Exotoxins: such as pyrogenic (erythrogenic) toxin
which causes the rash of scarlet fever and systemic
toxic shock syndrome.

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Extracellular toxins and enzymes


The most important in the context of pathogenicity are:
Streptolysin O, streptolysin S: destroy the membranes
of erythrocytes and other cells
Pyrogenic streptococcal exotoxins (PSE) A, B, C;
responsible for fever
Streptokinase: dissolves fibrin
DNases: breakdown of DNA, producing runny pus
Hyaluronidase: breaks down a substance that cements
tissues together

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Clinical significance
Acute pharyngitis(inflammation of pharynx) or
pharyngotonsilitis
Impetigo
Erysipelas
Puerperal sepsis
Acute rheumatic fever
Acute glomerulonephritis
Streptococcal TSS

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Laboratory diagnosis
Detection of the bacteria from clinical specimen
Gram stain
Culture
Catalase test negative
Identification from non pyogen B-hemolytic is by
bacitracin test, S. pyogen is more sensitive
Serology
Group A antigen can be detected using particles coated
with antibodies (latex agglutination)
ASO

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Treatment
Penicillin is still uniformly effective in treatment of
Group A Streptococcal disease
No effective vaccine has been produced, but specific Mprotein vaccines are being tested.
Prevention
Rheumatic fever is prevented by rapid eradication of the
infecting organism
Prolonged prophylactic antibiotic therapy is indicated
after an episode of rheumatic fever

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2. Streptococcus agalactiae

Approximately 25% of healthy women harbor S. agalactiae in


their vaginal flora and suffer no resulting ill effects

But immunosuppressed patients and new bornes are susceptible


to S. agalactiae infection

Transmission occurs from an infected mother to her infant at


birth and venereally (propagated by sexual contact) among adults

Group B Streptococci are a leading cause of meningitis and


septicemia in neonates, with a high mortality rate

They are also an occasional cause of infections in postpartum


women (endometritis) and individuals with impaired immune
systems, in whom the organism may cause septicemia or
pneumonia.
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Laboratory diagnosis
Smear- non motile gram positive cocci in chains
Culture- grow in aerobic and anaerobic environment at
temp 35-37oC
Grow in ordinary media with shiny or dry colonies with
gray white or colorless appearance
Shows clear zone of hemolysis(beta hemolysis) on blood Agar

Biochemical Test and Sensitivity Test

Catalase: Negative
Bile solubility test: Negative
cAMP test: Positive
Bacitracin: Negative/resistant

Treatment: Penicillin families


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3. Streptococcus pneumoniae (pneumococcus)


Gram positive, but old cultures: negative
Catalase negative

Encapsulated, spherical

Anaerobic

Orientation: pairs or chains

Lancet-shaped cocci

Fastidious

Ferment glucose to lactic acid

Alpha hemolytic

Transmission

Aerosols are major means of dissemination


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Virulance
Capsule
Choline binding proteins

Hemolysins

PspA (protective antigen): inhibit complement-mediated


opsonization
Autolysin LytA: responsible lysis in stationary phase as well as in
the presence of antibiotics
Autolysin LytB is a glucosaminidase involved in cell separation and
LytC exhibits lysozyme-like activity
Pneumolysin is stored intracellularly and is released upon lysis of
pneumococci by autolysin

IgA protease
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Clinical significance

Acute bacterial pneumonia


Otitis media
Bacteremia/sepsis
Meningitis

Laboratory diagnosis
1. Catalase negative; 2 H202 2 H20 + 02
2.Optochin test (ethylhydrocupreine HCl), inhibits
Optochin
growth of pneumococci but not viridans positive

Optochin negative
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3. Bile solubility test


Bile or bile salts (surface active agents) activate an
autolytic amidase which cleaves the bond between
alanine & muramic acid in the peptidoglycan resulting in
lysis of MOs.
Amidase is present in pneumococcus but not in viridans
4. Quellung (capsular precipitation) reaction
Most rapid & most useful
Pneumococcal specimen mixed with antipneumococcal
serum & methylene blue
Positive result: refractile & swollen capsules on oil
immersion
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5. Animal Inoculation
Fatal infection within 16-48 hours to mice injected
pneumococci antigen intraperitoneally
For experimental purposes

Serologic Diagnosis
Detection

of pneumococcal antibodies
Radioimmunoassay
Detection of capsular polysaccharide

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Treatment
Multiple antibiotic resistant strains of S. pneumoniae emerged in the
early 1970s
Increases in penicillin resistance have been followed by resistance to
cephalosporins and multidrug resistance
Antibiotic susceptibility testing is important
Prevention
There are two types of pneumococcal vaccine:
pneumococcal polysaccharide vaccine (PPV) and
pneumococcal conjugate vaccine (PCV7)
PPV: immunizes against 23 serotypes of
S. pneumoniae and is
indicated for the protection of high-risk individuals older than two
years

PCV7: is effective in infants and toddlers (six weeks to five years of age)

It is made up of seven pneumococcal antigens conjugated to CRM197


a47mutant nontoxic diphtheria toxin
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II. Gram Negative Diplococci

Include Neisseria spps.


Gram-negative cocci
Kidney bean appearance
Nonmotile, Non-spore forming, and Non-acid fast.
Oxidase positive
The genus contains two pathogenic and many commensal species,
most of which are harmless inhabitants of the upper respiratory and
alimentary tracts
The pathogenic species are Neisseria meningitidis (meningococcus), a
major cause of meningitis and bacteremia, and Neisseria gonorrhoeae
(gonococcus), the cause of gonorrhea
The structural elements of N. meningitidis and N. gonorrhoeae are the
same, except that the meningococcus has polysaccharide capsule
external to the cell wall
Gonococci
are more fastidious than meningococci 10/30/16
48

1. Neisseria gonorrhoeae
N.gonorrhoeae
is
frequently
observed
inside
polymorphonuclear leukocytes of clinical samples
obtained from infected patients
Usually transmitted during sexual contact or, more rarely,
during the passage of a baby through an infected birth
canal
It does not survive long outside the human body because
it is highly sensitive to dehydration
Gonococci are unencapsulated
Structure
1.Pili
Enhance attachment and resistance to phagocytosis
Among the most important virulence factors
At least twenty gonococcal genes code for pilin

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By shuffling and recombining chromosomal regions of


these genes, a single strain of N. gonorrhoeae can, at
different times, synthesize (express) multiple pilins that
have different amino acid sequences
This process, known as gene conversion, allows the
organism to produce antigenically different pilin
molecules over time
2. Lipooligosaccharide
Gonococcal lipooligosaccharides (LOS) have shorter,
more highly branched, nonrepeat O-antigenic side chains
than do lipopolysaccharides found in other gram-negative
bacteria (LPS).
The bactericidal antibodies in normal human serum are
IgM molecules directed against LOS antigens???

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3. Outer membrane proteins (OMPs)


OMP I, functioning as a porin in complex with OMP III,
is antigenically diverse in different strains of gonococci
OMP II (opacity protein ), along with pili, mediates
attachment of the organism to a host cell
Because of OMP II's ability to undergo extensive
antigenic variation, it also contributes significantly to the
ability of the organism to evade the immune response and
cause repeated infections
Classification and Antigenic Types
There are at least 70 different strains, characterized by the
absence or presence of pili, opacity of colonies,
auxotyping (nutritional requirements), serotyping and
genotyping
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Pathogenesis
It is associated with the acute onset of symptoms and
purulent mucosal drainage due to the organisms ability to
recruit polymorphonuclear leukocytes (PMNs)
Pili and OMP II facilitate adhesion of the gonococcus to
epithelial cells of the urethra, rectum, cervix, pharynx, or
conjunctiva and, therefore, make colonization possible
Pili also enable the bacterium to resist phagocytosis
Gonococci produce an IgA protease that cleaves IgA,
helping the pathogen to evade immunoglobulins
After gonococci attach to the nonciliated epithelial cells
of the fallopian tube, they are surrounded by the
microvilli, which draw them to the surface of the mucosal
cell
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The gonococci appear to enter the epithelial cells by a


process called parasite-directed endocytosis??
This process seems to be initiated by microbial factors
because it does not occur unless the gonococci are viable
and because it involves host cells that are not normally
phagocytic
Gonococci are not destroyed within the phagocytic vacuole;
it is not clear whether they replicate in the vacuoles
The major porin protein of the gonococcal outer membrane,
por (protein I), has been proposed as a candidate invasin (a
substance that helps mediate invasion into a host cell)
Gonococci can produce one or several outer membrane
proteins called Opa proteins (proteins II)
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These proteins are subject to phase variation and are


usually found on cells from colonies possessing an opaque
phenotype (O+).
At any one time, a gonococcus may express zero, one, or
several different Opa proteins, though each strain has 10 or
more genes for different Opas.
Trypsin-like proteases present in cervical mucus may help
select for protease-resistant transparent (O-) colony
phenotypes
O+ colony phenotypes (protease sensitive) predominate in
cultures taken during the middle portion of the menstrual
cycle?
Cervical proteases increase during the second half of the
cycle,
resulting in an increase in the O- phenotype
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Rmp (Protein III) is an outer membrane protein found in


all strains of N. gonorrhoeae
It does not undergo phase variation and is found in a
complex with Por and LOS
It shares partial homology with the Omp A protein of
Escherichia coli. Antibodies to Rmp, induced either by a
neisserial infection or by colonization with E.coli, block
bactericidal antibodies directed against Por and LOS.
LOS has a profound effect on the virulence and
pathogenesis of N gonorrhoeae.
Gonococcal LOS produces mucosal damage in fallopian
tube organ cultures and brings about the release of
enzymes, such as proteases and phospholipases, that may
be55important in pathogenesis
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More recent evidence suggests that gonococcal LOS


stimulates the production of tumor necrosis factor (TNF) in
fallopian tube organ cultures; inhibition of tumor necrosis
factor with specific antiserum prevents tissue damage
Thus, gonococcal LOS appears to have an indirect role in
mediating tissue damage
Gonococcal LOS is also involved in the resistance of N.
gonorrhoeae to the bactericidal activity of normal human
serum
Oligosaccharides containing epitopes defined by specific
monoclonal antibodies are associated with a serumresistant phenotype
Gonococci are highly autolytic and release peptidoglycan
fragments during growth
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These fragments, released by bacterial and/or host


peptidoglycan hydrolases, are toxic for fallopian tube
mucosa and may contribute to the intense inflammatory
reactions characteristic of gonococcal disease.
Gonococci (and meningococci) bind only human
transferrin and lactoferrin
This specificity is thought to be the reason these
organisms are exclusively human pathogens
Nevertheless, the role of transferrin- and lactoferrinbound iron in in vivo growth is unknown

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Host Defenses
Not everyone exposed to N. gonorrhoeae acquires the disease
This may be due to variations in the size or virulence of the
inoculum, to nonspecific resistance, or to specific immunity
Nonspecific factors have been implicated in natural resistance
to gonococcal infection
In women, changes in the genital pH and hormones may
increase resistance to infection at certain times of the menstrual
cycle
The variability in the susceptibility of gonococcal strains to the
bactericidal and bacteriostatic properties of urine is thought to
be one of the reasons some men do not develop a gonococcal
infection when exposed
Most uninfected individuals have serum antibodies that react
with
58 gonococcal antigens????
10/30/16

Infection with N. gonorrhoeae stimulates both mucosal and


systemic antibodies to a variety of gonococcal antigens
Mucosal antibodies are primarily IgA and IgG
In genital secretions, antibodies have been identified that
react with Por, Opa and LOS, and some of the ironregulated proteins
Gonococcal antigens such as pili, Por, Opa, Rmp, and LOS
elicit a serum antibody response during an infection
Antipilus antibody levels tend to be higher in women than
in men??
The predominant IgG subclass that reacts with a variety of
gonococcal antigens is IgG3, followed by IgG1 and IgG4.
IgG2 is minimal, suggesting that polysaccharides are not
important in the immune response to gonococcal infection
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Antigenic Variation
It is among several microorganisms whose surface
structures are known to change antigenically from
generation to generation during growth of a single strain
The antigenic structures of major interest are pili, Opa
proteins and LOS, for which there is evidence of antigenic
variation both in vitro and in vivo.

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Epidemiology
The only natural host for N. gonorrhoeae is the human
Gonorrhea is transmitted almost exclusively by sexual
contact
The highest rates occur in women between the ages of 15
and 19 years and in men 20 and 24 years of age
Gonorrhea is usually contracted from a sex partner who
is either asymptomatic or has only minimal symptoms
It is estimated that the efficiency of transmission after one
exposure is about 35% from an infected woman to an
uninfected man and 50 to 60% from an infected man to
an uninfected woman
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Clinical Significance
The organisms may cause a localized infection with the
production of pus, or may lead to tissue invasion, chronic
inflammation, and fibrosis.
A higher proportion of females than males are generally
asymptomatic; these individuals act as the reservoir for
maintaining and transmitting gonococcal infections.
Genitourinary tract infection presents with a yellow,
purulent urethral discharge and painful urination.
A greenish-yellow cervical discharge is most common,
often accompanied by intermenstrual bleeding.

63

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The disease may progress to the uterus, causing salpingitis


(inflammation of the fallopian tubes), pelvic inflammatory
disease (PID), and fibrosis.
Infertility occurs in approximately 20% of women with
gonococcal salpingitis, as a result of tubal scarring.
Rectal infections, prevalent in male homosexuals, are
characterized by constipation, painful defecation, and
purulent discharge.
Pharyngitis is contracted by oral-genital contact. Infected
individuals may show a purulent exudate, and the
condition may mimic a mild viral or a streptococcal sore
throat
64

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Ophthalmia neonatorum is an infection of the


conjunctival sac that is acquired by a newborn during
passage through the birth canal of a mother infected with
gonococcus.
If untreated, acute conjunctivitis may lead to blindness.
Treatment is with erythromycin because the antibiotic
also eradicates Chlamydia trachomatis, if present.
Disseminated Infection
Most strains of gonococci have a limited ability to
multiply in the bloodstream.
Therefore, bacteremia with gonococci is rare. (Note: In
contrast, meningococci multiply rapidly in blood )

65

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Diagnosis
Specimens for the laboratory diagnosis of gonorrhea
should be collected before treating the patient
N. gonorrhoeae grows best under aerobic conditions, and
most strains require enhanced CO2
N. gonorrhoeae ferments glucose but not maltose, lactose,
or sucrose.
Thayer-Martin medium (chocolate agar supplemented
with several antibiotics that suppress the growth of
nonpathogenic neisseriae and other normal and abnormal
flora) is typically used to isolate gonococcus

66

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Control
There is no effective vaccine to prevent gonorrhea
Candidate vaccines consisting of pilus protein or Por are of little
benefit
The development of an effective vaccine has been hampered by
the lack of a suitable animal model and the fact that an effective
immune response has never been demonstrated
Condoms are effective in preventing the transmission of
gonorrhea
The evolution of antimicrobial resistance in N. gonorrhoeae may
ultimately affect its control
Strains with multiple chromosomal resistance to penicillin,
tetracycline, erythromycin and cefoxitin have been identified in
different geographical areas
Sporadic
high-level resistance to spectinomycin and
fluoroquinolones
have been reported
67
10/30/16

2. Neisseria meningitidis

Twelve serogroups have been defined on the basis of the


antigenic specificity of a polysaccharide capsule
The most important disease-producing serogroups are A,
B, C, W-135, and Y
In addition to the group polysaccharides, individual N.
meningitidis strains may contain two distinct classes of
pili and multiple classes of OMPs
Nasopharyngeal carrier rate is 10%
Spreading is by respiratory droplets
Sero -groups C and A are associated with epidemic
disease

68

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Virulence Factors
Meningoccal endotoxin (LOS) is responsible for many toxic
effects
Capsule
protects bacteria from antibody mediated
phagocytosis
Pili allow to colonization of nasopharynx
Pathogenesis
The meningococcus is an exclusively human parasite
It can either exist as an apparently harmless member of the
normal flora or produce acute disease
Meningococci range from carrier state to bacteremia
Pili attach to microvilli as introduction to invasion
Polysaccharide capsules are antiphagocytic
Spread to blood and CNS produce systemic endotoxemia
69
LPS and PG trigger cytokine release 10/30/16

Manifestations
Meningitis is most frequent infection
Meningococcemia and rash may progress to disseminated
intravascular coagulation /DIC/
Meningococcal encephalitis
Pneumonia
Endocardiatis
Urethritis

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Diagnosis
Laboratory diagnosis

Specimen: CSF, blood


Smear: Gram-negative
Culture: Transparent or grey, shiny, mucoid colonies in
chocolate agar after incubation at 35-37Oc in a CO2
enriched atmosphere.
Intracellular

diplococci

Serology: Antibodies to meningococcal polysacharides


can be measured using: latex agglutination or
hemagglutination tests

71

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Biochemical reactions
Species

Glucos Lacto Malto Sucro


e
se
se
se
Neisseria
Positive Negati Negati Negati
gonorrhoeae
ve
ve
ve
Neisseria
Positive Negati Positiv Negati
meningitidis
ve
e
ve
Treatment
Penicillin and 3rd generation cephalosporins
such as cefotaxime are effective because of
good BBB(blood brain barrier) penetration
72

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Prevention
Rifampin is now the primary chemoprophylactic agent,
but ciprofloxacin has also been effective
A, C,Y, and W-135 polysaccharide vaccines are useful in
high-risk populations
Protein
conjugate vaccines(PCV) may enhance
immunogenicity in children
Nonimmunogenic serogroup B polysaccharide remains a
problem

73

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III. Enterobacteriaceae

Gram-negative, Non-sporing, rod shaped bacteria


Oxidase negative
Ferment glucose and may or may not produce gas
Reduce nitrate to nitrite (there are a few exceptions)
Are facultative anaerobes
If motile, motility is by peritrichous flagella
Many are normal inhabitants of the intestinal tract of man
and other animals
Some are enteric pathogens and others are urinary or
respiratory tract pathogens
Differentiation is based on biochemical reactions and
differences in antigenic structure
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The antigenic structure is used to differentiate organisms


within a genus or species
Three major classes of antigens are found:
o Somatic
O antigens: these are the heat stable
polysaccharide part of the LPS

Variation from smooth to rough colonial forms is accompanied by


progressive loss of smooth O antigen

Flagellar H antigens: are heat labile


o Envelope or capsule K antigens: overlay the surface O
antigen and may block agglutination by O specific antisera
Boiling for 15 minutes will destroy the K antigen and
unmask O antigens
The K antigen is called the Vi (virulence) antigen in
Salmonella typhi
o

75

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Classification

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1. Escherichia coli (E.coli)


Gram-negative
Non-spore forming
Rod-shaped
Oxidase negative
facultative anaerobe
Lactose fermenter
Motile

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Grow

well on the usual laboratory media in both the presence


and absence of oxygen
Normal flora of the mouth and intestine:Protects

the intestinal tract from bacterial infection


Produces small amounts of vitamins B12 and K
Lives symbiotically with us (we help them to live, and they help us to
live)
There

are more than 700 different serotypes of E. coli


Distinguished by different surface proteins and polysaccharides

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Transmission
Ingestion of contaminated food or water
Main sources of E.coli include:

Undercooked meat
Unpasteurized milk, apple juice and orange juice
Swimming in or drinking water contaminated with sewage
Unwashed vegetables and fruits
Bacteria in the diarrhea of infected persons can be passed on if
their hands are not washed well

Epidemiology
E.coli have world wide distribution
Many countries around the world are constantly suffering
from E. coli contamination
Is most common cause of diarrhea in tourists
79
10/30/16

Antigens of E.coli
O antigen
Somatic (on LPS)
171 antigens

H antigen( protein )

Flagella
56 antigens

K antigen

Capsule and (polysaccharides)


80 antigens

80

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Virulence Factors
Fimbriae (Pili)
Hemolysins
Flagella
Exotoxins(Thermolabile toxin (LT) & Thermostable toxin
(ST)
Endotoxin LPS
Capsules
K antigens
Drug resistance plasmids

81

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Infections caused by E. coli


While most E. coli are good for us, there are a few strains
of E. coli that are harmful to humans
E. coli is responsible primarily for three types of
infections in humans

Urinary tract infections


Neonatal meningitis and
Intestinal diseases

These conditions depend on a specific array of pathogenic


(virulence) determinants possessed by the organism
E. coli is best known for its ability to cause intestinal
diseases
Five classes of E. coli that can cause diarrheal diseases are
recognized:
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Enterotoxigenic E. coli (ETEC)


Enteroinvasive E. coli (EIEC)
Enterohemorrhagic E. coli (EHEC)
Enteropathogenic E. coli (EPEC) and
Enteroaggregative E. coli (EAggEC)

1. Enterotoxigenic E. coli (ETEC)


Is an important cause of diarrhea in infants and travelers
in underdeveloped countries or regions of poor
sanitation-travelers diarrhea
Diseases vary from minor discomfort to a severe
cholera-like syndrome
The disease requires colonization and elaboration of one
or more enterotoxins.
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Pathogenesis
Heat-labile (LT) enterotoxins: an A-B toxin which increases intracellular
levels of cAMP.
Subunit A causes intense and prolonged hyper-secretion of chloride ions
and inhibits the reabsorption of sodium and chloride
The gut lumen is distended with fluid and hypermotility and secretory
diarrhea occur, lasting for several days
It stimulates the production of neutralizing antibodies, and cross-reacts
with cholera toxin
The pathogenesis of LT is initiated by binding of LT-B subunits to the
ganglioside receptor GM1 found in caveolae on the host cell surface

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After binding, the LT holotoxin is internalized in vesicles


through the Golgi and the endoplasmic reticulum (ER)
In the Golgi, the holotoxin is disassembled
The A subunit is transported to the ER
Subunit A1 is the active molecule whereas subunit A2
anchors the subunit A to the B pentamer
The A1 subunit translocates through the intracellular
membrane to interact with ADP-ribosylating factors
(ARFs) in the cytoplasm
The activated A1 subunit then ADP-ribosylates the
subunit of a regulatory Gs, an intracellular guanine
nucleotide protein
85

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The inhibition of Gs GTPase activity results in the


irreversible activation of the transmembranar adenylate
cyclase complex leading to cAMP accumulation in the
cell
Elevated levels of cAMP activate cAMP-dependent
protein kinase A (PKA), inducing phosphorylation of the
cystic fibrosis transmembrane regulator (CFTR)
Activation of CFTR provokes the secretion of Cl- and
HCO3
PKA is also responsible for inhibition Na+ re-absorption
by the Na+/H+-exchanger 3 (NHE3).
Overall, the result is an osmotically-driven increased in
penetration of water and electrolytes resulting in fluid
accumulation
in the intestine.
86
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ST (heat-stable) toxin: causes an increase in cGMP in the


host cytoplasm - leads to secretion of fluid and electrolytes
resulting in diarrhea.
Binds to the extracellular domain of guanylate cyclase C
(GC-C) found on the brush border of the intestinal epithelium
Activation of the GC-C intracellular catalytic domain leads
to the conversion of GTP to cGMP and its intracellular
accumulation
This increase activates cGMP-dependent protein kinase II
(cGMPKII) leading to the phosphorylation of the CFTR
Also, elevated cGMP levels inhibit phosphodiesterase 3
leading to cAMP increase and activation of PKA.
Then, PKA act on the CFTR and also inhibits the reabsorption of sodium.
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ST produces secretory diarrhea via stimulation of the


CFTR channel and causing over-secretion of Cl- and
HCO3 which is followed by an osmosis-driven
electrolytes and water release by the cells.
Colonization factors (CFAs): facilitate the attachment of
E. coli strains to intestinal epithelium, usually are pili in
nature
Preventing their rapid removal by intestinal peristalsis
Epidemiology

World-wide; both adults and children


Incubation 1-2 days; persists 3-4 days
Mild symptoms including cramps, nausea, vomiting, watery
diarrhea
88

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2. Enteroinvassive E. coli (EIEC)

EIEC closely resemble Shigella in their pathogenic


mechanisms and the kind of clinical illness they produce
These bacteria can penetrate into the colonic mucosa,
where they cause ulcerous inflammatory lesions
Like Shigella, EIEC are invasive organisms
Not produce LT or ST toxin and, unlike Shigella, they do
not produce the shiga toxin

89

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Pathogenesis
In EIEC the plasmid encode a gene for a K surface
antigen which enables to:

Penetrate and multiply within epithelial cells of the colon


causing widespread cell destruction

Lyses the phagosomal vacuole, spread through the

cytoplasm and infect adjacent cell


Invade and destroy colonic epithelium

Clinical syndrome is identical to Shigella dysentery


and includes a dysentery-like diarrhea with fever
Cramps with blood and leukocytes in stool
Uncommon; often food-borne
90

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3. Enteropathogenic E. coli (EPEC)


Induce a watery diarrhea similar to ETEC, but they do not
possess the same colonization factors and do not produce ST or
LT toxins
Produce a non fimbrial adhesin designated intimin, an outer
membrane protein that mediates the final stages of adherence
Adhesiveness is mediated by plasmid-encoded intimin
Pathogenesis
Diarrhea and other symptoms of EPEC infections probably are
caused by bacterial invasion of host cells and interference with
normal cellular signal transduction, rather than by production of
toxins
Cause childhood diarrhea(infantile diarrhea)

91

Infants < 1 year affected


Watery diarrhea

10/30/16

4. Enteroaggregative E. coli (EAggEC)

Have the ability to attach to tissue culture cells in an aggregative


manner
Cause persistent diarrhea in young children
Resemble ETEC strains in that the bacteria adhere to the
intestinal mucosa and cause non-bloody diarrhea without
invading or causing inflammation

Pathogenesis

Bacterial cells autoagglutinate, stick to one another


Then produce a hemolysin related to the hemolysin produced by
E. coli strains involved in urinary tract infections.
Watery diarrhea, with vomiting, dehydration and low grade fever
are significant features

92

Infants < 6 months


AIDS patients

10/30/16

5. Enterohemorrhagic E. coli (EHEC)


EHEC are represented by a single strain (serotype
O157:H7), which causes a diarrheal syndrome distinct
from EIEC (and Shigella) in that there is copious bloody
discharge and no fever
A frequent life-threatening situation is its toxic effects on
the kidneys (hemolytic uremia).
E.g. Pediatric diarrhea caused by this strain can be fatal due to
acute kidney failure (hemolytic uremic syndrome [HUS]).

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Pathogenesis
Do not invade mucosal cells as readily as Shigella, but EHEC

strains produce a toxin that is virtually identical to the Shiga toxin.


The shiga toxin / cytotoxin plays a role in the intense

inflammatory response produced by EHEC strains and may


explain the ability of EHEC strains to cause HUS
The toxin is phage encoded unlike shigella (chromosome encoded)
Results in severe abdominal pain, bloody diarrhea, no fever
Affects children < 5 years

Diagnosis of E. coli

Clinical
Laboratory
Serology, culture, mcroscopy
94

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Treatment
Antibiotic therapy must take into consideration the resistance pattern

of the pathogen
Aminopenicillins
Ureidopenicillins
Cephalosporins,
Floroquinolones, or
Cotrimoxazole

Severe diarrhea necessitates oral replacement of fluid and electrolyte

losses

Prevention & Control


Sanitary: diligent adherence to hygiene is a paramount
Prophylactic antibiotics. eg cotrimoxazole

95

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2. Shigella

Enteric rods
Gram-negative
Facultative anaerobes
Non-spore forming
Oxidase negative
Non motile
Possess O and some have K antigens
Do not produce gas from glucose
Do not produce H2S on TSI
Non-lactose fermenters
Four species

Shigella sonnei (most common in industrial world)


Shigella flexneri (most common in developing countries)
Shigella boydii, and
Shigella dysenteriae
96

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Habitat & Transmission


Shigella species are found only in the human intestinal tract;
humans are the only reservoir
Transmitted by the feco-oral route
Mode of spread: person-to-person, contaminated fingers,
food, flies, fomites etc.
Infective dose: 10-100 viable bacilli
Incubation period: 1 to 7 days
Carriers of pathogenic strains can excrete the organism up to
two weeks after infection and occasionally for longer periods.
Duration of illness:

Untreated: severe symptoms for about two weeks


Antibiotic treatment shortens illness and prevent spread to
others

97

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Pathogenesis
Two-stage disease:
Early stage:

Watery diarrhea attributed to the enterotoxic activity of


Shiga toxin following ingestion and noninvasive
colonization, multiplication, and production of
enterotoxin in the small intestine

Fever attributed to neurotoxic activity of toxin


Second stage:

Adherence to and tissue invasion of large intestine with


typical symptoms of dysentery

Cytotoxic activity of Shiga toxin increases severity


98

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Virulence Factors
1.Invasiveness
Attachment (adherence) and internalization with complex
genetic control
Large multi-gene virulence plasmid regulated by multiple
chromosomal genes
2. Exotoxin (Shiga toxin)
3. Intracellular survival & multiplication

99

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Invasiveness in Shigella-Associated Dysentery


Penetrate through mucosal surface of colon and invade and
multiply in the colonic epithelium
Preferentially attach to and invade into M cells in Peyers
patches (lymphoid tissue, i.e., lymphatic system) of small
intestine
M cells typically transport foreign antigens from the
intestine to underlying macrophages, but Shigella can lyse
the phagocytic vacuole and replicate in the cytoplasm

Note: This contrasts with Salmonella which multiplies in the


phagocytic vacuole

Actin filaments propel the bacteria through the cytoplasm


and into adjacent epithelial cells with cell-to-cell passage

Effectively avoiding antibody-mediated humoral immunity

100

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Characteristics of Shiga Toxin


Enterotoxic, neurotoxic and cytotoxic
Encoded by chromosomal genes
Similar to the Shiga-like toxin of enterohemorrhagic E.
coli (EHEC)
Note:

except that Shiga-like toxin is encoded by lysogenic


bacteriophage

Clinical Features of Shigellosis


Persistent diarrhea with frequent and painful passage of
stools consisting mostly of blood, mucus and pus;
accompanied by fever and stomach cramps
Case fatality rates of 5-15%
101

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Cause breaks (ulcers) in mucous membrane lining of


intestine:Inflammation and tissue damage
Causes painful straining to pass stools; can lead to rectal
prolapse
Ulcers commonly in the rectum
Results in increased production of mucus
Loss of blood and serum proteins into intestinal cavity

Physical signs are those of dehydration beside fever, lower


abdominal tenderness & normal or increased bowel sounds.
The severity of the disease depends upon the species one is
infected
S. dysenteria is the most pathogenic followed by S. flexneri,
S.102sonnei and S. boydii.
10/30/16

Complications

Severe anorexia
Hypoproteinaemia
Dilation of the large intestine
Seizures
Anaemia
Hemolytic uremic syndrome(HUS)
Disseminated intravascular coagulation (DIC)
Reiter syndrome
Persistent diarrhoea
Rectal prolapse
Weight loss and malnutrition
Arthritis, conjunctivitis & urethritis
Myocarditis
103
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Laboratory diagnosis
Direct microscopy
Culture
Slide agglutination test
Macroscopy
Bright

red stool
Adherent to container
Odorless
small quantity

104

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Treatment
Replace fluids: Oral or IV rehydration with fluids and
electrolytes
Treat with antibiotics: trimethoprim/sulfamethoxazole,
Ampicillin, Ciprofloxacin,
Prevention and Control:
Hand washing, especially after defecation
Improved sanitation and hygiene
improve

water, waste treatment/disposal and food

sanitation
reduce overcrowding, etc.

No effective vaccine
105

10/30/16

3. Salmonella
Gram-negative
Oxidase negative
Non-sporulated
Facultatively anaerobic
Motile
Produce gas from glucose
Produce H2S on TSI media
Non lactose fermenter
Ubiquitous/very common human and animal pathogens
Salmonellosis in humans usually takes the form of a selflimiting food poisoning (gastroenteritis), but occasionally
manifests
as a serious systemic infection (enteric
fever)
106
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Possessing three major antigens


H

or flagellar antigen
O or somatic antigen and
Vi antigen

Vi antigen is a superficial antigen overlying the O antigen


It is present in a few serovars, the most important being S.
typhi.

107

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Habitat
The principal habitat of the salmonellae is the intestinal
tract of humans and animals
Typhi and Paratyphi A are strictly human serovars that
may cause grave diseases often associated with invasion
of the bloodstream.
Salmonella classification has undergone numerous
revisions; currently, all strains are grouped in two species:
S. enterica and S. bongori
2500 different serotypes
S.typhi, S.choleraesuis, and perhaps S.paratyphi A and
S.paratyphi B are primarily infective for humans, and
infection with these organisms implies acquisition from a
human
source
108
10/30/16

S. enterica is further divided into 6 subspecies and


contains different serotypes differentiated on the O and
H- Antigens. E.g.
Salmonella

serotype (serovar) typhimurium


Salmonella serotype enteritidis
Salmonella serotype typhi
Salmonella serotype paratyphi

109

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Patogenesis
Salmonella infections in humans vary with;

Strain
Infectious dose
Nature of the contaminated food and
Host status

An oral dose of at least 105 S.typhi cells are needed to cause typhoid
in 50% of human volunteers

Whereas at least 109 S. typhimurium cells (oral dose) are


needed to cause symptoms of a toxic infection
Infants, immunosuppressed patients, and those affected
with blood disease are more susceptible to Salmonella
infection than healthy adults

110

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In humans, Salmonella are the cause of two diseases called


salmonellosis:

Enteric fever (typhoid), resulting from bacterial


invasion of the bloodstream E.g; S. typhi and S.
paratyphi

Acute gastroenteritis (Non-typhoidal), resulting from a


food borne infection/intoxication.E.g; S.typhimurium, S.
enteriditis

Salmonella invade epithelial cells of the small intestine

Disease may remain localized or become systemic,


sometimes with disseminated foci
111

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1. Gastroenteritis

This localized disease is caused primarily by serotypes


enteriditis and typhimurium (nontyphoidal salmonella)

Transmission is via consumption of contaminated food

Infectious dose:
Typically about 1,000,000 bacteria
But much lower if the stomach pH is raised and if the
vehicle for infection is chocolate (about 100 bacteria)

112

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Virulence factors

Fimbriae or pili: attach the bacteria to cells lining the


intestinal lumen

The Vi antigen is a capsule that affords salmonellae some


protection from phagocytosis

Endotoxic LPS, which is responsible for septic shock in


patients with bacteriemia

Enterotoxins: responsible for many of the clinical signs of


Gastroenteritis, binds to GM1 gangliosides and cause
hyper secretion of fluids and electrolytes by elevating
levels of cAMP.
113

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Pathogenesis
Pathogenic salmonellae ingested in food survive passage
through the gastric acid barrier
Invade intestinal mucosa
Invasion of epithelial cells stimulates the release of
proinflammatory cytokines
Induces an inflammatory reaction
Causes diarrhoea and may lead to ulceration and
destruction of the mucosa

114

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Clinical Features of
Gastroenteritis
Symptoms usually begin

within 6 to 48 hours

NVD, Abdominal pain,


Headache, Fever

Duration varies, usually


2-7 days
Seldom fatal, elderly or
immunocompromised

115

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2. Typhoid salmonellosis

Caused by the serovars typhi and paratyphi A, B and C


Salmonellae are taken up orally
Enter lymphatic tissue
First spreading lymphogenously, then hematogenously
A generalized septic clinical picture results
Human carriers are the only source of infection

Asymptomatic Carriage

Chronic carriage in 1-5% of cases following S. typhi or S.


paratyphi infection
Gall bladder usually the reservoir
Chronic carriage with other Salmonella spp occurs in <<1% of
cases and does not play a role in human disease transmission
116

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Laboratory Diagnosis
Specimen:
Blood, stool, urine, serum for typhoid fever
Blood =80% positive in the first week
Stool=70-80% positive in the second and 3rd week
Urine =20% positive in the third and fourth week
Serum for widal test Positive after second week of illness

Culture and biochemical tests

117

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Serology
Widal test
The patient serum is tasted for O and H antibodies against
following antigen suspension
Serum agglutinins rise sharply during the second and third
weeks
High or rising titer of O ( 1:160) suggests that active
infection is present
High titer of H ( 1:160) suggests past immunization or past
infection
High titer of antibody to the Vi antigen occurs in some carriers

Treatment
Chloramphenicol (CAF)
Norfloxaclin???? Cipro
118

10/30/16

4. Vibrio cholerae
Vibrios are curved, Gram-negative rods commonly found
in saltwater
Cells may be link end to end, forming S shapes and spirals
They are rapidly motile by means of a single polar
flagellum
Pathogenic vibrios include:
1. V. cholerae serogroup O1 strains -epidemic cholera;
2. Non-O1 V. cholerae and related strains that cause sporadic
cases of cholera like and other illnesses; and
3. V.parahaemolyticus and other halophilic vibrios, which
cause gastroenteritis and extraintestinal infections
119

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There are over 150 O antigen serotypes, only two of


which (O1 and O139) cause cholera
Cholera is a life-threatening secretory diarrhea induced
by an enterotoxin secreted by V. cholerae
An O1 variant, V. cholerae biogroup El Tor is
distinguished by biochemical reactions
O139 strains resemble O1 El Tor strains but possess a
unique O antigen and have a polysaccharide capsule

120

10/30/16

Structure

The vibrios that cause epidemic cholera (O1) is subdivided into


two biotypes: classical and El Tor
Both biotypes (El Tor and classical) contain two major serotypes,
Inaba and Ogawa
These serotypes are differentiated in agglutination and vibriocidal
antibody tests on the basis of their dominant heat-stable LPS
somatic antigens
The cholera group has a common antigen, A, and the serotypes are
differentiated by the type-specific antigens, B (Ogawa) and C
(Inaba)
An additional serotype, Hikojima, which has both specific
antigens (rare)
V. cholerae O139 appears to have been derived from the pandemic
El Tor biotype but has lost the characteristic O1 somatic antigen
121

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Epidemiology
Transmission is through feco oral
Incubation period is hours-2 days
122

10/30/16

Pathogenesis
To produce disease, V. cholerae must reach the small
intestine in sufficient numbers to multiply and colonize
Large doses required to pass stomach acid barrier
Pili mediate epithelial adherence
CT-stimulated intestinal hyper secretion; causes diarrhea
Small intestine loses liters of fluid
K+ and bicarbonate losses cause hypokalemia and
acidosis

123

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Cholera Toxin
CT is an A-B type ADP-ribosylating toxin
B subunit receptor is a ganglioside on cell surface
Its molecule is an aggregate of multiple polypeptide
chains organized into two toxic subunits (A1, A2) and
five binding (B) units.
A1 enters cytoplasm and ADP ribosylates regulatory G
protein
Adenylate cyclase becomes locked in active state
The cAMP causes the active secretion of Na, Cl, K,
HCO3 and water out of the cell into the intestinal lumen

124

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Manifestations
Extreme watery diarrhea causes large fluid loss
Dehydration and electrolyte imbalance are the major
problems
Diagnosis
The initial suspicion of cholera depends on recognition of the
typical clinical features in an appropriate epidemiologic
setting.
Bacteriologic diagnosis is accomplished by isolation of V.
cholerae from the stool.
The organism grows on common clinical laboratory media
such as blood agar and MacConkey agar
But its isolation is enhanced by the use of a selective
medium
(thiosulfate-citrate-bile salt-sucrose10/30/16
agar).
125

Treatment
Oral or intravenous fluid and electrolyte replacement is
crucial
Antimicrobic therapy can reduce duration and severity
Tetracyclines
Trimethoprim and
Erythromycin

Prevention
Water sanitation and cooking shellfish prevent infection
Vaccines prepared from whole cells, LPS, and CT B
subunit have been disappointing, providing protection that
is not long-lasting
Other Vibrios ?????
126

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5. Helicobacter Pylori
Helicobacter pylori is a small, curved, gram-negative,
rod-shaped bacterium.
H. pylori causes more than 90% of duodenal ulcers and
up to 80% of gastric ulcers
Growth requires a microaerophilic atmosphere and is
slow (3 to 5 days)
Epidemiology
H.pylori infection is one of the most common bacterial
infections in the world
Humans are the only known reservoir
Transmission by feco-oral pathway
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Pathogenesis
Virulance factor
Urease: ammonia production neutralizes acid
Flagella: allows the organisms to swim to the less acid pH
locale beneath the gastric mucus
Surface proteins (Cag protein)
Neutrophil-activating protein (NAP)
H. pylori colonization is virtually always accompanied by a
cellular infiltrate ranging from minimal mononuclear
infiltration of the lamina propria to extensive inflammation
with neutrophils, lymphocytes, and microabscess formation
This inflammation may be due to toxic effects of the urease or
the VacA

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The Cag protein may contribute by stimulation of


cytokines (interleukin-8), and a neutrophil-activating
protein (NAP) has been shown to recruit neutrophils to
the gastric mucosa
Added together urease, Cag, and NAP provide ample
explanation for the gastritis that is universal in H. pylori
infection.
A prolonged and aggressive inflammatory response
could lead to epithelial cell death and ulcers, but other
virulence factors play a more direct role.
The chief of these is VacA, which is responsible for
much of the epithelial cell erosion seen in human
infection.
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Clinical manifestation
Primary infection with H. pylori is either silent or causes
an illness with nausea and upper abdominal pain lasting
up to 2 weeks
Years later, the findings of gastritis and peptic ulcer
disease include nausea, anorexia, vomiting, epigastric
pain, and even less specific symptoms such as belching
Many patients are asymptomatic for decades, even up to
perforation of an ulcer
Perforation can lead to extensive bleeding and peritonitis
due to the leakage of gastric contents into the peritoneal
cavity
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Diagnosis
The most sensitive means of diagnosis is endoscopic
examination, with biopsy and culture of the gastric mucosa
Active infection can also be diagnosed with a 13C- or 14Clabeled urea breath test or with a fecal antigen detection assay
H. pylori specific IgG (serum or salivary antibody) is a useful
marker for epidemiologic studies of past or current infection,
but its sensitivity is suboptimal
Treatment
H. pylori is susceptible to a wide variety of antimicrobial
agents
Requires combination therapy with two or more antibiotics
A typical regimen includes amoxicillin plus clarithromycin
plus a proton pump inhibitor, such as omeprazole
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IV. Mycobacterium genus


1. M. tuberculosis
Contain over 17,000 different strains
TB is a disease of humans and humans are the only
reservoir for the bacterium

Rod-shaped
Neither Gram+ve nor Gram -ve
Non-motile and non-sporulated
Obligate aerobe
Facultative intracellular parasite
Slow generation time (15-20hrs)
Lack exotoxins or endotoxins
Classified
as acid-fast bacteria
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Classification
Family: Mycobacteriaceae
Order : Actinomycetales
Genus: Mycobacterium
Four species under this genus:
M. tuberculosis complex
The non-tuberculosis mycobacteria
M. leprae and
M.ulcerance

Human pathogens under mycobacterium genus include:

M.tuberculosis
M.bovis------consumption of unpasteurized milk
M. leprae
M.aviumTB like disease (HIV Pts)

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The disease tuberculosis


TB infection means that M.TB is in the body but the
immune system is keeping the bacteria under control
People who have TB infection but not TB disease are
NOT infectious
The immune system does this by producing macrophage
TB infection is not considered a case of TB
Predisposing factors for TB infection
Over crowded
Poor nutrition
HIV infection is the #1 predisposing factors for M.TB
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Tuberculosis: Infection vs Disease


TB Infection

TB disease in lungs

M.TB. present

M.TB. present

Tuberculin skin test positive

Tuberculin skin test positive

Chest X-ray normal

Chest X-ray usually reveals lesion

Sputum smears and cultures negative

Sputum smears and cultures positive

No symptoms

Symptoms such as cough, fever, weight


loss

Not infectious

Often infectious before treatment

Not defined as a case of TB

Defined as a case of TB

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Virulance
M. tuberculosis do not produce any toxins
Its ability to survive and multiply within macrophage is
its main virulence factor
The organism is slow growing and tolerates the
intracellular environment
It is known to prevent the acidification of the phagosome
that is needed for effective killing of microbes by
lysosomal enzymes
Lipoarabinomannan is a heteropolysaccharide that
inhibits macrophage activation by IFN-gamma

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Pathogenesis
Mycobacterium tuberculosis is spread by small airborne
droplets generated by:
Coughing
Sneezing

Talking or
Tinging

Leads to infection of the respiratory system


The infection may not be progress in to
disease( immunity, strain and infectious dose)
The majority of the bacilli are trapped in the upper parts
of the airways
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Diagnosis of Pulmonary Tuberculosis


1. Clinical diagnosis
2. Laboratory diagnosis

Microscopy

Culture

Molecular diagnosis

Skin test

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Tuberculin Skin Test


is used globally in screening for M.TB infection
The skin test works by injecting Purified Protein
Derivative (PPD) into the skin
PPD test measures hypersensitivity to tuberculoprotein
PPD test interpreted by area of induration
Positive PPD indicates past or current infection

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Interpretation
Read 48 to 72 hours later
An area of measured induration of 10 mm or more
constitutes a positive reaction
No induration indicates a negative reaction
A positive PPD test indicates that the individual has been
infected at some time with M. tuberculosis or with a
strongly cross-reacting mycobacterium of another species
A negative PPD
test in a healthy
individual
indicates that he
or she has not
been infected
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with M.

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Radiographic procedures
The initial suspicion of pulmonary TB is often based on
abnormal chest radiographic findings in a patient with
respiratory symptoms
Treatment

Rifampicin
Isoniazide
Bactericidal, first line
Pyrazinamide
Streptomycin
Ethambutol
Ethionamide
Thiacetazone
Bacteriostatic
Paraminosalicylic acid
141

Second line

Aminosalicylic

acid
Ethionamide
Cycloserine
Fluoroquinolon

es
Kanamycin
10/30/16

Ionized and rifampin are active against both intra- and


extracellular organisms
Pyrazinamide acts at the acidic pH found within cells
Streptomycin does not penetrate into cells and is thus active
only against extracellular organisms
Rifampin (RIF)(inhibit transcription)
Isoniazid (INH)( mycolic acids synthesis inhibitor)
Pyrazinamide (PZA )( bactericidal drug active)
Ethambutol (EMB)(affects the biosynthesis of the cell wall, it
contributes towards increasing the susceptibility of M.
tuberculosis to other drugs)
Streptomycin (SM)(inhibits the initiation of mRNA
translation)
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Drug resistant M.TB

Drug resistance in tuberculosis (TB) is a matter of great concern for


TB control programs

Resistance is usually acquired by alteration of the drug target


through mutation or by titration of the drug through overproduction
of the target

Mono Resistance: Resistance to single drug

Poly-Resistance: Resistance to more than one drug other than


Rifampicin and Isoniazid together

Multi-Drug Resistance (MDR): Resistance to at least Isoniazid and


Rifampicin

Extensive Drug Resistance (XDR-TB): MDR plus Resistance to:


Fluoroquinolone

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Aminoglycosides (Kanamycin, Amikacin)

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2. M.leprea
Typical acid-fast bacilli-singly, in parallel bundles, or in
globular masses
Are regularly found in scrapings from skin or mucous
membranes (particularly the nasal septum) in lepromatous
leprosy
The bacilli are often found within the endothelial cells of
blood vessels or in mononuclear cells
Pathogenicity
M. leprae is transmitted from human to human through
prolonged contact
Obligate intracellular parasite of macrophages

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Clinical significance
Leprosy is a chronic granulomatous condition of peripheral
nerves and mucocutaneous tissues, particularly the nasal
mucosa.
It occurs between two clinical extremes: tuberculoid and
lepromatous leprosy
In tuberculoid leprosy, the lesions occur as large maculae
(spots) in cooler body tissues such as skin (especially the nose,
outer ears, and testicles), and in superficial nerve endings
Neuritis leads to patches of anesthesia in the skin
The lesions are heavily infiltrated by lymphocytes and giant and
epithelioid cells, but caseation does not occur.
The patient mounts a strong cell-mediated immune response
and develops delayed hypersensitivity which can be shown by
skin test with lepromin, a tuberculin-like extract of lepromatous
tissue.
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The course of lepromatous leprosy is slow but


progressive.
Large numbers of organisms are present in the lesions
and reticuloendothelial system, and immunity is severely
depressed

Lepromatous
Leprosy

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Tuberculoid
Leprosy

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Laboratory identification
M. leprae is an acid-fast bacillus.
It has not been successfully maintained in artificial
culture, but can be grown in the footpads of mice, which
may also be a natural host although playing no role in
human disease.
Laboratory diagnosis of lepromatous leprosy, where
organisms are numerous, involves acid-fast stains of
specimens from nasal mucosa or other infected areas.
In tuberculoid leprosy, organisms are extremely rare, and
diagnosis depends on clinical findings and the histology
of biopsy material

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Treatment and prevention


Several drugs are effective in the treatment of leprosy,
including sulfones such as dapsone, rifampin, and
clofazamine.
Treatment is prolonged, and combined therapy is
necessary to ensure the suppression of resistant mutants.
The fact that vaccination with BCG has shown some
protective effect in leprosy has encouraged further
interest in vaccine development.

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V. Spirochetes
are

spiral shaped
Many are difficult to see by routine microscopy
Many are thin and take stains poorly
Only darkfield microscopy, immunofluorescence, or special staining
techniques that effectively increase their diameter can demonstrate
these spirochetes.
Some are strict anaerobes, others require low concentrations of
oxygen and still others are aerobic
Some have not been isolated in culture
Spirochetes that are important human pathogens are confined to three
genera:
Treponema (T. pallidum causes syphilis)
Borrelia (B.burgdorferi causes Lyme disease, B. recurrentis causes
relapsing fever) and
Leptospira (L. interrogans causes leptospirosis)
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1.Treponema pallidum

Nonpathogenic treponemes may be part of the normal


flora of intestinal tract, oral cavity or genital tract
Some of the oral treponemes have been associated with
gingivitis and periodontal disease
fastidious
has not been successfully cultured in vitro (limited and
slow growth)
The lipid composition of T. pallidum is complex,
consisting of several phospholipids, including cardiolipin,
and a poorly characterized glycolipid which is
biochemically and immunologically distinct from LPS
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Epidemiology
is an exclusively human pathogen
Transmission is by contact with mucosal surfaces or blood
In most cases, infection is acquired from direct sexual
contact with an individual who has an active primary or
secondary syphilitic lesion
Less commonly, the disease may be spread by non genital
contact with a lesion (eg, of the lip), sharing of needles by
intravenous drug users, or transplacental transmission to
the fetus within approximately the first 3 years of the
maternal infection
Modern screening procedures have essentially eliminated
blood transfusion as a source of the disease
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Pathogenesis
The spirochete reaches the subepithelial tissues through in
apparent breaks in the skin /passage between the epithelial
cells, where it multiplies slowly with little initial tissue
reaction

May be due to the relative paucity of exposed antigens on the


surface of the organism, but no specific reasons are known

As lesions develop, the basic pathologic finding is an


endarteritis
The small arterioles show swelling and proliferation of
their endothelial cells
This reduces or obstructs local blood supply, probably
accounting for the necrotic ulceration of the primary lesion
and subsequent destruction at other sites
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Slow multiplication produces endarteritis, granulomas


Ulcer heals but spirochetes disseminate
Syphilis is then silent/unknown reason/ until the
disseminated secondary stage develops and then silent
again with entry into latency
Although evasion of host defenses is clearly taking place,
the mechanisms involved are unknown.
Latent

periods may be due to surface binding of host


components

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The inflammatory response to:


immune complexes
spirochetal lipoproteins and
complement in arteriolar walls accounts for some of the
injury in syphilitic lesions.

The granulomatous nature of the lesions in late syphilis is


consistent with injury caused by delayed-type
hypersensitivity responses

No toxins, virulence factors, or other molecules can yet be


linked with specific features of syphilis

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Manifestations
I. Primary stage
Spirochetes multiply locally at the site of entry, and some
spread to nearby lymph nodes and then reach the
bloodstream
In 210 weeks after infection, a papule develops at the
site of infection and breaks down to form an ulcer with a
clean, hard base ("hard chancre")
The inflammation is characterized by a predominance of
lymphocytes and plasma cells
This "primary lesion" always heals spontaneously, but 2
10 weeks later the "secondary" lesions appear
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II. Secondary stage


Manifest with generalized Maculopapular rash and white
patches in the mouth
There may also be syphilitic meningitis, hepatitis,
nephritis
The secondary lesions also subside spontaneously
Both primary and secondary lesions are rich in spirochetes
and highly infectious
III. Latent stage
In about 30% of cases, early syphilitic infection progresses
spontaneously to complete cure without treatment
In another, the untreated infection remains latent
(principally evident by positive serologic tests)
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Patients are symptom free but relapse can occur


In early stage patient is infectious but in late stage of
latent syphilis patients are none infectious
IV. Tertiary stage
Characterized by the development of granulomatous
lesions (gummas) in skin, bones, and liver; degenerative
changes in the CNS (meningovascular syphilis); or
cardiovascular lesions
In all tertiary lesions, treponemes are very rare, and the
exaggerated tissue response must be attributed to
hypersensitivity to the organisms
However, treponemes can occasionally be found in the
eye or CNS in late syphilis

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Diagnosis
Specimens include tissue fluid expressed from early
surface lesions for demonstration of spirochetes; blood
serum for serologic tests
1. Darkfield Examination
A drop of tissue fluid or exudate is placed on a slide and
a cover slip pressed over it to make a thin layer.
The preparation is then examined under oil immersion
with dark field illumination for typical motile spirochetes

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2. Serologic Tests
These tests use either nontreponemal or treponemal antigens
Nontreponemal includes:
VDRL (Venereal Disease Research Laboratory) and
RPR (rapid plasma reagin)

Treponemal tests includes


Fluorescent Treponemal Antibody (FTA-ABS) Test
Treponema pallidum-Particle Agglutination (TP-PA) Test (T.
pallidum hemagglutination (TPHA))

Treatment
Doxycycline and amoxicillin are the first-line antimicrobics
for the treatment of early Lyme disease and arthritis
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VI. Rickettsiae

Obligate intracellular gram-negative coccobacilli


occurring in single, pairs, short rods and filaments
Poorly stained in gram reaction
Grow in yolk sac of embryonated eggs, cell culture and
laboratory animals
Destroyed by heat, drying and bactericidal chemicals
With the exception of C. burnetii (Coxiella burnetii) , the
organisms are transmitted by arthropods
C. burnetii is transmitted exclusively by inhalation of
dust containing the pathogens
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Clinical Features

Clinical illness is due to the invasion and multiplication


of rickettsiae in the endothelial cells of small blood
vessels
It manifests with fever, headache, malaise, skin rash and
enlargement of liver and spleen
The genus rickettsia has three main groups based on their
antigenic structure
These are:

Typhus group

Scrub typhus group

Spotted fever group

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Hosts and vectors of the medically important


rickettsiae
Organism
Typhus group
R.
prowazeckii
R. typhi
Scrub typhus
R.tsutsugamu
shi

Disease
Louse-borne
typhus
Murine typhus
Scrub typhus

Spotted fever
group
Rocky Mountain
R. conorii
Spotted fever
R. rickettis
Rickettsial pox
R. akaris
162

Host

Vector

Man
Rat

Body
louse
Rat flea

Rodents

Mite

rodents,
dogs
Rodents,
dogs
Mice

Tick
Tick
Mite

10/30/16

R. prowazeckii and R. typhi are common in Ethiopia

1. Rickettisia Prowazeckii

It causes epidemic or louse-borne typhus


Clinical Features

It is transmitted by self-inoculation of the organism by


scratching after bite by infected louse

The illness manifests with sudden onset of fever,


headache, malaise and skin rash.

Epidemics of the disease are associated with


overcrowding ,cold weather, lack of washing facilities,
famine and war
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2. Rickettsia Typhi

It causes endemic or flea-borne typhus

Clinical Features:

It is transmitted to man when bitten by an infected rat-flea

The disease is milder than louse-borne typhus and occurs in


those individuals living or working in highly rat-infested
area.

Laboratory Diagnosis

Specimen: Serum for serological test

The serological tests to diagnose typhus are Weil-felix


Treatment

Tetracycline

Chloramphenicol
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VII. Chlamydia

Chlamydiae are obligate intracellular parasites


Cell envelope consists of two lipid bilayers with associated
cell wall material that resembles a gram-negative envelope
Contains no PG and muramic acid
Two stages of reproductive cycle:

Elementary bodies (EB) (outsideof host cells)


Initial bodies (IB) (inside the host cells)

The EB is taken up by phagocytosis into susceptible host


cells
Once inside the cell, the elementary body prevents fusion of
the phagosome and lysosome, protecting itself from
enzymatic destruction
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The particle reorganizes over the next eight hours into a


larger, noninfectious reticulate body, which becomes
metabolically active and divides repeatedly by binary
fission within the cytoplasm of the host cell
As the reticulate body divides, it fills the endosome with
its progeny, forming an inclusion body
After 48 hours, multiplication ceases and reticulate bodies
condense to become new infectious elementary bodies
The elementary bodies are then released from the cell by
cytolysis, ending in host cell death
The three human pathogen species of chlamydiae are:

C. psittaci
C. trachomatis and
C. pneumoniae

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1. Chlamydia psittaci
The natural hosts of C. psittaci are birds
This species causes infections of the respiratory organs,
the intestinal tract, the genital tract and the conjunctiva of
parrots and other birds
Humans are infected by inhalation of dust (from bird
excreta containing the pathogens)
After an incubation period of one to three weeks,
psittacosis /ornithosis presents with fever, headache, and
a pneumonia that often takes an atypical clinical course
The infection may, however, also show no more than the
symptoms of a common cold, or even remain clinically
silent
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Diagnosis
Diagnosis is primarily serologic
The pathogen can be grown from sputum in special cell
cultures
Therapy: Tetracyclines (doxycycline)
2. Chlamydia trachomatis
It is a pathogen that infects only humans
There are 15 serotypes of C trachomatis

C. trachomatis serotype A, B, Ba, C causes trachoma


C. trachomatis serotype D-K causes genital infection
C. trachomatis serotype L1-L3 causes lymphogranuloma venereum
(LGV)

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