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What is Cholera?
A life-threatening secretory diarrhea induced by enterotoxin
secreted by V. cholerae
Water-borne illness caused by ingesting water/food
contaminated by copepods infected by V. cholerae
An enterotoxic enteropathy (a non-invasive diarrheal disease)
A major epidemic disease
Cholera
Recent Cholera Pandemics1-6th pandemic: V. cholerae O1 biotype classical 1817-1923, Asia, Africa, Europe, America and Australia
7th pandemic: V. cholerae O1 biotype El Tor Began in Asia in 1961 Spread to other continents in 1970s and 1980s Spread to Peru in 1991 and then to most of South & Central
America and to U.S. & Canada By 1995 in the Americas, >106 cases; 104 dead
1993: Cholera in Bengal caused by O139
may be cause of 8th pandemic
VibrioVibrio
• Vibrio cholerae --gastroenteritis
• Vibrio parahaemolyticus -- gastroenteritis,
wound infection, bacteremia
• Vibrio vulnificus -- wound infection, bacteremia
Grows in salt and fresh water
Endemic in areas of poor sanitation (India and
Bangladesh ), transmitted by fecal-oral route
Can survive and multiply in brackish water by infecting
copepods
Has over 150 identified serotypes based on O-antigen
Only O1 and O139 are toxigenic and cause Cholera
disease
V. cholerae
Classification: O1 Antigen
小川型 稻叶型 彦岛型
Classification: Other antigens
O139 Serogroup
In 1993, an entirely new serogroup (O139) cause an
epidemic in Bangladesh.
O139 organisms produce a polysaccharide capsule but do
not produce O1 LPS or O1 antigen.
Non-O1, Non-O139 Serogroup
Most are CT (cholera toxin) negative and are not
associated with epidemic disease.
Profile of vibrio cholerae
• G-, curved or comma-shaped rods
• Highly motile; polar flagellum
• Sensitive to low pH and die rapidly in
solutions below pH 6
• Proliferate in summers
• Cholera toxin
• Pathogenic and nonpathogenic strains
• 206 serogroups
Similarities to Enterobacteriaceae
G-, Facultative anaerobes
Fermentative bacilli
Differences from Enterobacteriaceae
Polar flagella
Oxidase positive
Formerly classified together as Vibrionaceae
Primarily found in water sources
Cause gastrointestinal disease
Shown not closely related by molecular methods
Profile of vibrio cholerae
Broad temperature & pH range for growth on media
18-37C
pH 7.0 - 9.0 (useful for enrichment)
Grow on variety of simple media including:
MacConkey’s agar
TCBS (Thiosulfate Citrate Bile salts Sucrose) agar
V. cholerae grow without salt
Most other vibrios are halophilic
Physiology of Vibrio
People with low gastric acid levels (103 -105 CFU )
Children: 10 × more susceptible than adults
Elderly
Blood types
O>> B > A > AB
People Most at Risk
Period of Communicability
During acute stage
A few days after recovery
By end of week, 70% of patients non-infectious
By end of third week, 98% non-infectious
Symptoms
Occur 2-3 days after consumption of contaminated food/water
Usually mild, or no symptoms at all
• 75% asymptomatic
• 20% mild disease
• 2-5% severe Vomiting Cramps Watery diarrhea (1L/h) Without treatment, death in 18 h-several days
Cholera Gravis (霍乱肌无力 )
More severe symptoms
Rapid loss of body fluids
6 liters/hour
107 vibrios/mL
Rapidly lose more than 10% of bodyweight
Dehydration and shock
Death within 12 hours or less
Death can occur within 2-3 hours
Virulent factors
• Toxins and enzymes
- heat stable endotoxin
- enterotoxin (exotoxin [cholera toxin CT])
CT is antigenically and pharmacologically
identical in all sero and bio types
Pathogenesis of V. cholerae
Cholera Toxin (CT) Structure
• The A subunit contains an intracellular ADP-ribosyltransferase activity.
• The mature A subunit is proteolytically cleaved to produce an A1 polypeptide, which contains the intracellular enzymatic activity, and an A2 polypeptide.
• CT is a prototype A/B subunit toxin, 1A+5B
• The B subunit form a pentameric ring, which binds the holotoxin to a eukaryotic cell surface receptor.
Cholera Toxin (CT) Structure
• After cleavage, the A1 and A2 polypeptides remain
linked by a disulphide bond.
• A and B subunits are connected through the C-terminus of the A2 subunit, which is inserted through the central pore of the B pentamer.
• The biological activity of CT is
dependent on binding of B
pentamer to specific receptors
GM1 ganglioside.
How Does Cholera Toxin Work?
• Internalization is initiated once
CT-GM1 complexes cluster which
then invaginate to form apical
endocytic vesicles.
How Does Cholera Toxin Work?
• These vesicles enter cellular
trafficking pathways leading to the
trans-Golgi network (TGN).
• The toxin then moves retrograde
via the Golgi cistern to the ER.
• Once in the ER, CT is processed to
activate the A1 peptide, which then
targets the basolateral membrane
(heterotrimeric GTPase and
adenylate cyclase (AC)).
How Does Cholera Toxin Work?
• Adenylate cyclase (AC) is
activated normally by a
regulatory protein (GS) and
GTP; however activation is
normally brief because
another regulatory protein
(Gi), hydrolyzes GTP.
NORMAL CONDITION
How Does Cholera Toxin Work?
• A1 fragment catalyzes the attachment of ADP-Ribose (ADPR) to the regulatory protein forming Gs-ADPR from which GTP cannot be hydrolyzed.
• Since GTP hydrolysis is the event that inactivates the adenylate cyclase, the enzyme remains continually activated.
CHOLERA
• Thus, the net effect of the
toxin is to cause cAMP to be
produced at an abnormally
high rate which stimulates
mucosal cells to pump large
amounts of Cl- into the
intestinal contents.
How Does Cholera Toxin Work?
• H2O, Na+ and other electrolytes follow
due to the osmotic and electrical gradients
caused by the loss of Cl-.
• The lost H2O and electrolytes in mucosal
cells are replaced from the blood.
• Thus, the toxin-damaged cells become
pumps for water and electrolytes causing
the diarrhea, loss of electrolytes, and
dehydration that are characteristic of
cholera.
How Does Cholera Toxin Work?
• Inactivates GTPase function of G-protein coupled
receptors in intestinal cells
• G proteins stuck in “On” position
• 100 fold increase in cAMP
• Activation of ion channels
• Ions flow out and water follows
How Does Cholera Toxin Work?
Diagnosis
• Cholera should be suspected when patients present
with watery diarrhea, severe dehydration
• Based on clinical presentation and confirmed by
isolation of vibrio cholera from stool
• No clinical manifestations help distinguish
cholera from other causes of severe diarrhea: Enterotoxigenic E. coli
Viral gastroenteritis
Bacterial food poisoning
Diagnosis: Visible Symptoms
• Decreased skin turgor
• Sunken eyes, cheeks
• Almost no urine production
• Dry mucous membranes
• Watery diarrhea consists of:• fluid without RBC, proteins• electrolytes• enormous numbers of vibrio cholera (107 vibrios/mL)
Laboratory Diagnosis
• Visualization by dark field or phase microscopy• Look like “shooting stars”
• Gram Stain• Red, curved rods of bacteria
• Isolate V. cholerae from patient’s stool• Plate on sucrose agar• Yellow colonies form
Vibrio Prevention & Control
• Disrupt fecal-oral transmission
Improved sanitation
• Fluid and electrolyte replacement
• Antibiotic prophylaxis
• Improved food handling
Vibrio parahemolyticus
• One kind of halophilic vibrios;
• optimal NaCl concentration contained in culture media is 3.5%;
• hemolysin related to its pathogenicity, can be detected by human or rabbit RBC test (Kanagawa test);
• cause food poisoning in human beings.
• raw sea-food
• Clinical manifestations– Self-limiting diarrhea to mild cholera-like illness
– 24 hours after ingestion-explosive water diarrhea
• Headache, abdominal cramps, nausea, vomiting, low grade fever for 72 hours or more
• Uneventful recovery
• Wound infections in people exposed to seawater-containing vibrios
Vibrio parahemolyticus
Helicobacter pylori
Helicobacter pylori
•G- with S or spiral-shaped
•Very Motile ---corkscrew motion
•2~6 flagella at one end of HP
•5% O2+10% CO2 at 37 oC
•Reaction of urea hydrolysis and
creates an ammonia cloud
Pathogenesis of Hp
CagA (Cytotoxin associated)
VacA (vacuolationg associated)
LPS also play great importance
Flagellum and urease is necessary
for its adhesion and inhabitation
Adhesin
Duodenal Ulcer (DU) Gastric Ulcer (GU)
Campylobacter jejuni
•G- rods with comma, S, or “gull-wing”
shapes.
•Motive, with a single polar flagellum
•No spore & no capsule
•5% O2+10% CO2
•Two types of colonies:
watery and spreading
round and convex
• Produces a toxin called Cytolethal Distending Toxin (CDT).
• CDT activity requires activation of three genes: cdtA, cdtB, and cdtC.
• CdtB is nuclease that damages DNA and causes cell cycle arrest.
• Causes cell death.
Pathogenesis of Campylobacter
What Are the Symptoms?
• Diarrhea
– Usually watery and sticky
– Can contain blood and fecal leucocytes
• Fever
• Abdominal pain
• Nausea and vomiting
• Headache
• Muscle pain
Who is affected?
• All warm-blooded animals can become affected.
Some animals carry the disease without exhibiting
symptoms.
• Any person can become infected.
• Children under 5 and young adults ages 15-29 are
most often affected.
• Most deaths occur among the elderly and the
immune-suppressed.
Summary
• Properties of Vibrio (stain, culture,
biochemical reaction, antigens and
virulence factors)
• Pathogenesis of V. parahemolyticus, H.
pylori, C. jejuni