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Vibrio cholerae and vibrio parahemolyticus toxins
1. Vibrio cholerae and Vibrio
parahemolyticus toxins
Dr Ravi Kant Agrawal, MVSc, PhD,
Senior Scientist (Veterinary Microbiology)
Food Microbiology Laboratory
Division of Livestock Products Technology
ICAR-Indian Veterinary Research Institute
Izatnagar 243122 (UP) India
2. 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
3. Recent Cholera Pandemics
1-6th 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
4.
5. 500-400 BC: Sanskrit writings
500 BC: Hippocrates
200 AD: Galen
900 AD: Rhazes, Islamic physician
Sanskrit, Arabic, and Chinese writings dating back 2,000 years
Ancient Texts Describe Cholera
6. Started in by Ganges in Calcutta - Kumbh festival
Polluted water, crowded camps
10,000 in British army and hundreds of thousands of natives
dead
Spread by trade routes – Iran, Baku, Astrakhan, Russia
Cold winter kept it from reaching western Europe
1st
Pandemic: 1817-1823
7. England’s attempt to control spread of infectious disease
Tried to prevent international movement
Eventually repealed (based on flawed scientific understanding)
Quarantine Act of 1825
8. Bengal, Afghanistan, Asia, Moscow, England, US
William Brooke O’Shaughenessy
Industrial Revolution
England’s Cholera Prevention Act of 1832
Entered US through NY and New Orleans ports – spread by
railway and troop movement after Civil War
2nd
Pandemic: 1829-1852
13. Began in Bengal
Britain and
Europe affected
Dr. John Snow
Mapped cases
to find cause
Broad Street
Pump
3rd
Pandemic: 1852-1859
14. Map led Snow to believe that Broad Street pump was cause of
outbreak
Those affected drank from pump
Sewage probably contaminated well
Removal of pump handle - end of outbreak
Skepticism about Snow’s findings
Broad Street Pump
15. Compared deaths from Cholera between 2 groups
Group A: Southwark and Vauxhall Water Co. – 70 deaths per 10,
000 (London source of Thames)
Group B: Lambeth Water Co. – 5 deaths per 10,000 (source
upstream from London
The “Grand Experiment”
16. Massive public health reforms
Much smaller outbreak in 1866
Results
17. From Egypt to Europe by returning pilgrims from the
Haj at Mecca
Imported into NY by ship
Last time cholera in England
Third and Fourth International Sanitary Conferences
(Paris and Vienna)
International Health Regulations
International Sanitary Commission – precursor of PAHO (Pan
American Health Organization)
4th
Pandemic: 1863-1879
18. Began in India, spread east and west
1883 - Robert Koch cultured V. cholerae
Good sanitation – did not affect much of Europe
Diagnosis and quarantine – kept it out of US
Prevented contact between those with exposure to
unsanitary conditions (on ships) and those on mainland
5th
Pandemic: 1881-1896
19. Spread through Asia
Did not affect Europe or US
6th
Pandemic: 1899-1923
20. 1959: cholera enterotoxin by S.N. De in Calcutta
Cholera bacillus is not harmful – toxin is what induces
outpouring of fluid and inhibits sodium transport
Treatment by rehydration (oral or intravenously) of fluid and
electrolytes
How to measure rapid fluid loss
Discoveries
21. Caused by El Tor strain
From Pacific Islands to Asia, Bangladesh, India, USSR, Iran, Iraq
1970: reemerged in Africa after 100 years
1991: Latin America (4,000 dead of 400,000 cases)
1993: O139 serogroup (“Bengal”) – may be start of 8th
pandemic
7th Pandemic: 1961-present
22. Aug 2000: published complete DNA sequence of V. cholerae, El
Tor strain
Unusual - 2 distinct chromosomes
Hope that genome will be useful in creating an effective vaccine
Genome
24. Grows in salt and fresh water (Brackish rivers, coastal waters)
Can survive and multiply in brackish water by infecting
copepods
Associate with plankton and algae
Endemic in areas of poor sanitation (India and Bangladesh)
Transmitted by fecal-oral route
Has over 150 identified serotypes based on O-antigen (206
serogroups)
ONLY O1 AND O139 ARE TOXIGENIC AND CAUSE CHOLERA.
V. cholerae
25. Classification: Serogroups and Biotypes
The species V. cholerae can be sub-classified into > 200
serogroups based on the O antigen of LPS (lipopolysaccharide).
Only O1 and O139 strains have been implicated in the cholera
syndrome.
26. Classification: O1 Serogroup
2 Biotypes: El Tor and Classical
V. cholerae O1 are further divided
into 2 major sub-serotypes (Inaba
Ogawa and Hikojima ).
The basis for sub-typing is 3
antigenic determinants of the O
antigen structure of their LPS.
These sub-serotypes are
differentiated in agglutination
and vibriocidal antibody tests on
the basis of their dominant heat-
stable lipopolysaccharide somatic
antigens.
The serotypes share one
determinant known as the A
antigen.
In addition, Inaba strains express
the C antigen whereas Ogawa
strains express the B antigen .
27. Classification: O1 Serogroup
O1 cholera almost always fall into the Heiberg I fermentation
pattern; that is, they ferment sucrose and mannose but not
arabinose, and they produce acid but not gas.
Hiiberg (1934) divided Vibrios in 6 groups based on fermentation
of sucrose, mannose and arabinose.
Vibrio cholera also possesses lysine and ornithine
decarboxylase, but not arginine dihydrolase.
Freshly isolated agar-grown vibrios of the El Tor biotype, in
contrast to classical V. cholerae, produce a CELL-ASSOCIATED
MANNOSE-SENSITIVE HEMAGGLUTININ which is found active in
chicken erythrocytes.
Strains of the El Tor biotype, however, produce less cholera
toxin, but appear to colonize intestinal epithelium better than
vibrios of the classical variety.
Also, El Tor biotype, seem some what more resistant to
environmental factors.
Thus, El Tor strains have a higher tendency to become endemic
and exhibit a higher infection-to-case ratio than the classical
biotype.
28. Classification: Other antigens
O139 Serogroup
In 1993, the emergence of an entirely new serogroup (O139)
was the cause an epidemic in Bangladesh.
O139 organisms produce a POLYSACCHARIDE CAPSULE but
do not produce O1 LPS or O1 antigen.
Toxigenic O139 cholera arose through the acquisition of a
large block of genes encoding the O139 antigen by O1 El Tor.
Non-O1, Non-O139 Serogroup
Most are CT (cholera toxin) negative and are not associated
with epidemic disease.
29. Strains Causing Epidemics
2 main serogroups carry set of virulence genes necessary for
pathogenesis
O1-two major biotypes - classical and E1 Tor - distinction based on
biochemical properties and susceptibility to bacteriophages
E1 TOR produce hemolysins
Classical: 1 case per 30-100 infections
El Tor: 1 case per 2-4 infections
O39 Contained /restricted to Bangladesh, India: 1993 outbreak in
Bangladesh and India
O39 genetically derived from E1 tor, but antigenic structure
different enough—no existing immunity—all susceptible
Serogroup - group of microorganims serotype having 1/more
antigens in common
30. Profile of Vibrio cholerae
Gram-negative rods
CCurved or comma shapedurved or comma shaped
Highly motile; single polar flagella
Non-spore forming
Associated with salt water
Oxidase positive
Facultative anaerobe
1.4-2.6 μm length x .5-.8 μm width
Chemo-organotroph
Optimal growth 20-30 degrees
RReadily cultivatedeadily cultivated,, ssimple nutritional requirementsimple nutritional requirements
Tolerate alkaline conditions to pH9.0Tolerate alkaline conditions to pH9.0
Sensitive to low pH and die rapidly in solutions below pH
6
Proliferate in summers
Produces cholera toxin
Pathogenic and non-pathogenic strains
206 serogroups
lipopolysaccharide coat which provides protection
against hydrophobic compounds
provides a surface for immune recognition
31. 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
32. Divisions of V. Cholerae
Biotype (biovar)
different strains of the same bacterial species
distinguished by a group of phenotypic or genetic traits
Serogroup
bacteria of the same species with different antigenic
determinants on the cell surface
V. Cholera has more than 150 (206) different serogroups, only
two of which cause epidemic disease
33. V. Cholerae
01 serogroup
Classic genome: 3.2-3.6 MbClassic genome: 3.2-3.6 Mb
El Tor (El) genome: 4 MbEl Tor (El) genome: 4 Mb
O1 antigen is divided into 3O1 antigen is divided into 3
types: A,B,Ctypes: A,B,C
• A antigen:A antigen: made of 3-deoxy-made of 3-deoxy-
L-glycerotetronic acidL-glycerotetronic acid
• B, C antigen:B, C antigen: not beennot been
characterizedcharacterized
34. Broad temperature & pH range for growth on media
• 18-37°C
• 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
35. Vibrio cholerae: Antigenic structure
– Common heat-labile flagellar H antigen
– O lipopolysaccharide confers serologic specificity
• More than 150 O antigen serogroups
• Only O-1 and 0139 serogroups cause Asiatic cholera
• Three serotypes; Ogawa, Inaba, Hikojima
• Two biovars of O1 derogroup; classic and El Tor
37. Vibrio cholerae: Epidemiology
– Epidemic cholera - spread by contaminated water under
conditions of poor sanitation
– Endemic - consumption of raw seafood
– Copepods
38. People with low gastric acid levels
Children: 10 × more susceptible than adults
Elderly
Blood types
O>> B > A > AB
People Most at Risk
39. Period of Communicability
During acute stage
By end of week, 70% of patients non-infectious
By end of third week, 98% non-infectious
A few days after recovery
40. Vibrio cholerae - Clinical manifestations
Asymptomatic colonization to fatal diarrhea
Onset 2-3 days after ingestion
Abrupt onset of watery diarrhea and vomiting
RICE WATER STOOLS
Severe fluid and electrolyte loss-dehydration, metabolic acidosis,
hypovolemic shock, renal failure
Death 60% if untreated, 1% if treated for fluid loss
cause of death: Dehydration and shock
41. Symptoms
Occur 2-3 days after consumption of contaminated food/water
No fever-not invasive
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
Clever-viable even after exit body-take host’s liquid
42. Cholera Gravis
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
43. Vibrio-Prevention and Control
Improved sanitation
Fluid and electrolyte replacement
Antibiotic prophylaxis
Improved food handling
44. Virulence factors of V. cholerae O1 and O139
Virulence factor Biological effect
Cholera toxin Hypersecretion of electrolytes and water
Co-regulated pilus Adherence to mucosal cells adhesin
Accessory colonization
factor
adhesin
Hemagglutination
protease
Releases bacteria from mucosal cells
Zona occludens Exotoxin
Accessory cholera
enterotoxin
Exotoxin
Flagellum Motility
Siderophores Iron sequestration
45. Vibrio cholerae: Pathogenesis
Ingest 108
-1010
organisms
Non invasive infection of small intestine
Organisms secrete enterotoxin
Watery diarrhea
46. Cholera disease begins with ingestion of contaminated water
or food.
The bacteria that survive the acidic conditions of the stomach
colonize in the small intestine.
The cholera toxin (CT) is responsible for the severe diarrhea
characteristic of the disease.
Other virulence factors- heat stable endotoxin
Cholera Toxin
CT is a proteinaceous enterotoxin (exotoxin) secreted by V.
cholerae.
CT is antigenically and pharmacologically identical in all
serotypes and biotypes.
Pathogenesis of V. cholerae
47. Horizontal Gene Transfer
1. Acquisition of VPI1. Acquisition of VPI
2. lysogenic conversion by phage2. lysogenic conversion by phage
3. exchange of genes leads to expression of O-antigen and capsule3. exchange of genes leads to expression of O-antigen and capsule
48. V. Cholera
• The 01 strain and
the recent 0139
strain have
different antigens
expressed in the
polysaccharide
capsule
• The change in
structure is
thought to have
arisen from a
recombination
event.
49. V. Cholerae
two circular chromosomestwo circular chromosomes
Chromosome 1 is larger
(2.96 million base pairs)
and carries many genes
for essential cell
functions and
housekeeping.
Chromosome 2 is smaller
(about 1.07 million base
pairs) and carries the
integron island.
50. Chromosome 1
Carries many genes for essential cell
functions and housekeeping.
It also contains important virulence
genes, most of which have been
acquired by lateral gene transfer from
other species
Chromosome one carries two
bacteriophages.
ONE VIRUS is called the V. cholera
pathogenicity island phage (VPI),
which infects and inserts its DNA into
the bacterial chromosome and allows
the synthesis of a pilus which the
bacteria uses to attach to the host
intestine.
SECOND VIRUS is called the cholera-
toxin phage phi (CTXΦ). The CTX
phage inserts itself into chromosome
one and the bacterium is then capable
of secreting a powerful enterotoxin.
51. Chromosome 2
The integron region is often found
on plasmids and serves as a "gene
capture system."
This region may contain antibiotic
resistance genes.
52. Genetics of Cholera Toxin
Genes encoding CT ctxAB-
recognized to be the genome
of a filamentous phage CTXΦ
(ctxA and ctxB).
Transcription of ctxAB is
regulated by several proteins.
CTXΦ genome can integrate
into the host genome at a
specific site, attRS.
The CTX genetic element also
has a “core” region carrying
several phage morphogenesis
genes.
These entire CTX gene set is
flanked repeated sequences,
the attRS1 site.
The entire genetic element is
6.9kb.
The receptor for CTXThe receptor for CTXΦΦ is Toxin-Coregulated Pili (TCP)is Toxin-Coregulated Pili (TCP)
53. Toxin-Coregulated Pili
Efficient colonization of V. cholera in the small intestine
requires the expression of TCP’s
• TCP’s are expressed on the surface of V. cholera
• TCP’s are long laterally associated filaments
• The major pilin subunit is TcpA
Genes for TCP production are clustered on the pathogenicity
island located on chromosome 1
54. Recap of phage movement
V. cholerae did not always cause disease.
Infection with the CTX phage (a temperate and filamentous
phage) gives the bacterium its toxinogenicity.
The phage recognizes a pilus on the surface of the bacterium
and uses it to enter the cell.
Once inside the cell, the CTX phage integrates into the
chromosome and the lysogen expresses cholera toxin.
The CTX phage has received special attention because it is the
first filamentous phage found to transfer toxin genes to its
host.
The important lesson from this discovery is that many different
types of phage may carry virulence factors, and transfer of
virulence genes by phage may be a major mechanism of
evolution of new bacterial diseases.
55. Genomic Structure: Bacteriophage
In 1996 Matthew K. Waldor and John J. Mekalanos reported a
stunning discovery about the toxin.
The toxin was for the first time shown to be not a part of the
bacterium but actually that of a virus that got integrated into
the V. cholerae genome.
Normally this virus remains silent within V. cholerae but during
infection it gets activated.
The major virulence factor of cholera, CT (cholera toxin) is
encoded on a filamentous phage (ctxΦ) that is capable of
transducing the ctx gene into other cholera strains.
The released phages specifically attach to the bacterium and
enter it.
Vigorous viral multiplication results in the production of large
amounts of toxin causing severe diarrhea.
56. Genomic Structure: Pathogenicity Islands (PAI)
Upon transduction, the
bacteriophage (ctxΦ)
brings the toxin and a
specific pilus called toxin-
co-regulated pilus (TCP).
The important genes
involved in intestinal
colonization (tcp) and
virulence gene regulation
(toxT) are encoded in a
40Kb pathogenicity island.
This PAI is present in
pathogenic cholera
strains.
tcp gene
ctx gene
57. Pathogenesis: Overview
To establish disease, V. cholerae must
be ingested in contaminated food or
water and survive passage through
the gastric barrier of the stomach.
On reaching the lumen of the small
intestine, the bacteria must overcome
the clearing mechanism of the
intestine (peristalsis), penetrate the
mucous layer and establish contact
with the epithelial cell layer.
58. Pathogenesis: Overview
Colonization of the intestinal microvilli and the subsequent
production and release of cholera toxin, lead to the purging
diarrhea.
This complex progression of events appears to involve tightly
regulated differential gene expression by the bacteria.
This is because expression of intestinal colonization factors is
unlikely to be of advantage to the bacterium in its salt/fresh
water environment niche.
59. Pathogenesis: Cholera Toxin (CT)
In 1983, by administering purified CT to volunteers, Levin et al.
were able to conclusively demonstrate that the toxin is the
major mediator of the cholera syndrome.
Ingestion of only 5μg of purified toxin resulted in production of
1-6 L of diarrheal stool.
CT elicits vigorous mucosal immune responses in the absence of
a conventional adjuvant.
Direct immunomodulatory effects of CT on leukocytes include
induction of CD25 and class II MHC on B cells, apoptosis of CD8+
T cells, and activation of macrophages with release of IL-10.
60. Pathogenesis: Cholera Toxin (CT) Structure
CT is a prototype A/B subunit toxin,
consisting of one A subunit and 5 B
subunits.
The B subunit weighs 11.6kDa each and
multimerize to form a pentameric ring,
which binds the holotoxin to a eukaryotic
cell surface receptor.
61. Pathogenesis: Cholera Toxin (CT) Structure
• The A subunit contains an intracellular ADP-ribosyltransferase
activity.
• The mature A subunit is proteolytically cleaved to produce a
21.8kDa A1 polypeptide, which contains the intracellular
enzymatic activity, and a 5.4kDa A2 polypeptide.
• After cleavage, the A1 and A2 polypeptides remain linked by a
disulphide bond.
• The crystal structure of CT revealed that the 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.
62. Pathogenesis: Cholera Toxin (CT) Structure
CT must be assembled for activity, as neither the A nor B
subunit individually can cause secretory diarrhea.
CT holotoxin is assembled in the periplasmic space.
The subunits are exported individually into the periplasm
through the cytoplasmic membrane via the general secretion
pathway; both the A and B protein subunits contain normal
sequences at their N-terminus.
63. Pathogenesis: Cholera Toxin (CT) Structure
Once in the periplasm, both
subunits must undergo
modification by the
periplasmic enzyme DsbA,
which is responsible for
disulphide bond formation.
Again, once the holotoxin is
secreted from the bacterium,
the A subunit must be cleaved
to generate separate A1 and
A2 peptides for maximum
toxin activity.
64. Cholera Toxin (CT) Structure
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.
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.
65. 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.
66. • The biological activity of CT is dependent on binding of B
pentamer to specific receptors on enterocytes: GM1 ganglioside.
• Internalization is initiated once CT-GM1 complexes cluster which
then invaginate to form apical endocytic vesicles.
How Does Cholera Toxin Work?
67. 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)).
68. 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
69. 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
70. 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-
and bicarbonates into the
intestinal contents.
How Does Cholera Toxin Work?
71. • 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?
72. Encoded by a prophage
Molecular mass of 84,000 daltons
A subunit-ADP-ribosylating toxin
B subunit-bind GM1-gangliosides on enterocytes
A subunit ADP ribosylates Gs-alpha which regulates activation
of adenlyate cyclase which inactivates GTPase function of G-
protein coupled receptors in intestinal cells
G proteins stuck in “On” position
Result is persistent increase in cAMP levels-100 fold increase in
cAMP
Activation of ion channels
Ions flow out- Hyper secretion of Na, Cl, K, bicarbonate
The change in ion concentrations leads to the secretion of large
amounts of water into the lumen, known as diarrhea
How Does Cholera Toxin (ctxAB) Work?
76. 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
Bacterial food poisoning
Viral gastroenteritis
77. 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)
78. Laboratory Diagnosis
Visualization by dark field or phase
microscopy
Look like “shooting stars”
Gram Stain: gram – curved rods
Isolate V. cholerae from patient’s
stool/vomitus - Specimens
Culture:
Transport medium - Cary-Blair semi-
solid agar
Enrichment medium - alkaline
peptone broth: Vibrios survive and
replicate at high pH while other
organisms are killed or do not
multiply
Selective Medium: TCBS agar plate,
sucrose agar: Yellow colonies form
(sucrose fermentation)
Quick immunological methods:
immunofluorescent “ball” test and
PCR
81. Vibrio Prevention & Control
• Disrupt fecal-oral transmission by
improved sanitation
• Fluid and electrolyte replacement
• Antibiotic prophylaxis
• Improved food handling
82. Treatment
*Even before identifying cause of disease, rehydration therapy
must begin Immediately because death can occur within hours*
Oral rehydration
Intravenous rehydration
Antimicrobial therapy
83. Treatment: Oral Rehydration
Reduces mortality rate from over 50% to less than 1%
Recover within 3-6 days
Should administer at least 1.5x amount of liquid lost in stools
Use when less than 10% of bodyweight lost in dehydration
84. Treatment: Oral Rehydration Salts (ORS)
Reduces mortality from over 50% to less
than 1%
Packets of Oral Rehydration Salts
Distributed by WHO, UNICEF
Dissolve in 1 L water
NaCl, KCl, NaHCO3, glucose
85. Treatment: How ORS Works
Na+
transport coupled to
glucose transport in small
intestine
Glucose enables more efficient
absorption of fluids and salts
Potassium passively absorbed
86. Treatment: ORS in United States?
American doctors skeptical of such simple, inexpensive
treatment
Cost
ORS: $270/infant
IV: $2,300/infant
$1 billion/year for IV treatment for rehydrating children
Insurance companies do not reimburse for ORS
600 American children die unnecessarily from dehydration each
year
Hospitals consider IV more time efficient
Less personal attention required
87. Treatment: Intravenous Rehydration
Used when patients have lost more than 10% bodyweight from
dehydration
Unable to drink due to vomiting
Only treatment for severe dehydration
88. Treatment: Intravenous Rehydration
Ringer’s Lactate
Commercial product
Has necessary
concentrations of
electrolytes
Alternative options
Saline
Sugar and water
Do not replace
potassium, sodium,
bicarbonate
89. Treatment: Antibiotics
Adjunct to oral rehydration
Reduce fluid loss by half
Reduce recovery time by half
2-3 days instead of 4-6
Tetracycline, Doxycycline
Not recommended
Short duration of illness
Antibiotic resistance
Limited gain from usage
90. Boil or treat water with chlorine or iodine
No ice
Cook everything
Rule of thumb: “Boil it, cook it, peel it, or forget it.”
Wash hands frequently
Traveling Precautions
91. Vaccines
Need localized mucosal immune response
Oral Vaccine
Not recommended
Travelers have very low risk of contracting disease: 1-2
cases per million international trips
Not cost-effective to administer vaccines in endemic
regions
Brief and incomplete immunity
Two types approved for humans:
Killed whole-cell
Live-attenuated
92. Vaccines: Killed Whole-cell Vaccines
Provides antigens to evoke protective antitoxic and
antibacterial immunity
Contains:
1 x 1011
heat inactivated bacteria
Mixture of V. cholerae O1 El Tor and classical strains
1 mg of B subunit of cholera toxin
93. Killed Whole-cell Vaccines: Disadvantages
50% protection for 6 months to adults
Gives less than 25% protection to children aged 2-5
Need for multiple doses of nonliving antigens
94. Vaccines: Live-Attenuated
Eliminates need for multiple doses of non-living antigens
Ensures that crucial antigens potentially altered during killing
process would be retained
Expected to mimic broad immunity conferred by natural
infection
85-90% protection against classical biovar
65-80% protection against El Tor biovar
95. Live Attenuated Vaccines: Disadvantages
In children, protection rapidly declines after 6 months
In adults, only receive 60% protection for 2 years
Live vaccine induces mild cholera symptoms
Mild diarrhea, abdominal cramping
97. Precautions Taken in US
Environmental Protection Agency (EPA) works closely with
water and sewage treatment operators
FDA
Tests imported shellfish
Controls US shellfish sanitation program
98. 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.
Major source: raw sea-food.
99. 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
104. Thanks
Acknowledgement: All the presentations available online on the
subject are duly acknowledged.
Disclaimer: The author bear no responsibility with regard to the
source and authenticity of the content.
Editor's Notes
1)In industrialized with reported cases cholera--imported
Serogroup- group of microorg serotype having 1/more antigens in common
O1-two major biotypes
O39 restricted to Bangladesh, India
O1-South America
--what does O MEAN???
two biotypes—classicla and E1 Tor—dist based on biochem prop and susceptibility to bacteriophages
1993 outbreak in Bangladesh and India
E1TOR prod hemolysins
9) O39 genetically derived from E1 tor, but antigenic structure different enough—no existing immunity—all agues susceptible
1) Gram neg-rod shaped
a)cell walls don’t retain dye—appear red
b)pos-thick cell walls-retain dye
1)Will grow on syntehtic media with glucose as sole source of crabon and energy
2)summers-water temps exceed 20 C
a)summers—illnesses inc
3)Zoo plankton?
Not sure why O more susceptible—allows bacteria to adhere to gut lining—guess
a)e1tor and 0139—cholera gravis—mostly O blood type
endemic regions
a)bangladesh—low attack rates in adults
b) Antibodies– resistance--- intestinal IgA prevent attachment to mucosal surface, neutralize cholera toxin
b)Cmmon in children
1)No fever—not invasive
2) clever-viable even after exit body—take host’s liquid
1)Bact prod toxin that….
2)Prevents hydrolysis of GTP to GDP
1
Define dark field and phase microscopy!
1) Treatment depends on severity of dehydration
1)Distr to cholera inflicted areas
2) Fluid therapy
a) electrolyte solution with glucose for energy
3)Add glucose to therapy—elminates need for sterility, IV
1)Sodium transport coupled to glucose transport in small intestine
a)glucose accelerates absorption of glucose and water
2)Glucose Enables small intestine to absorb fluids and salts more efficiently
3)Boil water for at least 10 minutes
4)Bicarbonate—corrects acidosis
5)Sports drinks
a)excess sugar—worsen condition bc of osmolosis
1)cotransport- coupling system
2)Glucose Enables small intestine to absorb fluids and salts more efficiently
3)Boil water for at least 10 minutes
4)Bicarbonate—corrects acidosis
5)Sports drinks
a)excess sugar—worsen condition bc of osmolosis
6) $2300 per infant vs $270
a)1 billion dollars on children in US
b)600.year die from dehydration
c)doctors not reimbursed for ORT bc not widely accepted practice in industrailized countries—use IV
d)ORT requires mom to hold/feed—helathcare profs won’t give that attn
2)Doctors don’t trust bc it is so simple/cheap
3)Time consuming
4)Insurance companies don’t reimburse
1)Underused in children with dehydration
1)Need intuitive system
2)epidemic-local people help each other
no medical personel, illiterate
3)Doctors without borders
4) IV drip treatment of cholera victims at Refugee camp mozambique
1)Saline can be used because it’s better than nothing