VIBRIO

Dr. Priyadarshini Shanmugam

Professor
Department of Microbiology

 Vibrios - Gram Negative

rigid curved rods
Family Vibrionaceae
Actively motile by a polar
flagellum
Most important member is
Vibrio cholerae
First isolated by Koch from
Cholera patients in Egypt

Vibrio cholerae
MORPHOLOGY
Short, curved, cylindrical rod
comma shaped
Curvature lost in subculture
Measures 1.5 m x 0.2-0.4 m
ARRANGEMENT
Arranged in parallel rows fish in
stream appearance

Motility
Actively Motile with a single sheathed
polar flagellum darting motility

Swarm of gnats appearance in stool

samples from acute cholera cases/young
cultures.

Staining

Gram Negative
Non acid fast, nonsporing

LEIFSONS FLAGELLAR
STAIN

Classification of Vibrios
Classification:
(Gardner & Venkatramans)
Vibrio
GroupA
Vibrio cholerae
(Common H antigens)
O1

NonO1

Classical
Ogawa

Group B
Heterogeneous (biochemically)
(antigenically)

Eltor (biotypes)
Inaba

Hikojima (serotypes)

O 139 (non O1) vibrio has also been isolated from cholera cases.

Classical

El Tor

Hemolysis

VP Test

Chick RBC
agglutination
Polymyxin B

sensitivity
Group IV phage
susceptibilty
Eltor phage V
susceptibilty

HEIBERGS CLASSIFICATION
Based on the fermentation of mannose,
sucrose & arabinose
Classified into 6 groups. Later 2 more groups
were added

HEIBERGS CLASSIFICATION (Contd)

Group

Mannose

Sucrose

Arabinose

I
II
III
IV
V
VI
VII
VIII

A
A
A
A
-

A
A
A
A
-

A
A
A
A

THE CHOLERA TOXIN (CTX):

MW 84,000
Five identical B (binding)
subunits
An A (active) subunit, which
is composed of 2 chains:
A1 and A2
- A2 links A1 to B
- A1 is the active fragment

THE CHOLERA TOXIN (CTX)

Phage encoded
Or plasmid encodedcan be transferred to
other nonpathogenic
vibrios
Similar to LT of Ecoli

Mechanism of action of Cholera toxin

CHOLERA- Pathogenesis:
Vibrio ingested in small amount
Acid barrier
Epithelial cells of SI
Adhere to enterocytes
Adherent bacteria produce potent enterotoxin
(cholera toxin)
B subunit binds to GM1 ganglioside receptors

Pathogenesis Contd
Toxin A passes inside cytoplasm
A1 release
Activation of adenylate cyclase
Overproduction of cAMP
Outpouring of large amount of electrolytes
(K+ and HCO3-) - water follows
Watery diarrhoea

Other enzymes
Vibrio elaborates several enzymes such as Collagenase,
elastase, lipase, mucinase, nucleotidase, neuraminidase
Motility, chemotaxis, mucinase help it to pass
through the mucin and attach to the epithelial cell
surface
Cholera lectin is another enzyme- hemagglutininprotease cleaves mucin.
Toxin coregulated pilus- helps in attachment

CHOLERA
Incubation period: 24 hours-5days
Acute diarrheal disease
Starts slowly as mild vomiting & diarrhea or
abruptly with sudden massive diarrhea.
Cholera stool is typically a colorless, watery
fluid with flecks of mucus-rice water stool.
Characteristic inoffensive, sweetish odour.

 Severe diarrhea leads to diminished ECF

volume, hemoconcentration, hypovolemia,
acidosis, shock.
Complications: muscular cramps, renal failure,
pulmonary edema, cardiac arrhythmias,
paralytic ileus
Death

LABORATORY DIAGNOSIS OF CHOLERA

Sample- Stool during acute stage of disease has vibrios in
large numbers 104-109/ml of stool.
Mode of Collection: Stool/Rectal catheter/Rectal swabs
Transport medium is necessary in tropical areas where
vibrios die in 4 hours.
Direct examination Darting motility/Motility inhibition
by antisera.
Enrichment media- Alkaline peptone water
Media-colonies

CULTURE REQUIREMENTS

Strongly aerobic.
Temperature range 16 -40oC (370C)
Alkaline pH preferred (6.4 -9.2)
Preferred pH is 8.2
0.5 1.0% NaCl is required for optimal growth

CULTURE CHARACTERISTICS
Grows well on ordinary media
Nutrient Agar: colonies are
moist, translucent, round discs,
1-2 mm in diameter
Blood Agar-greenish
discoloration, hemodigestion
Peptone water growth occurs in
6 hours as a fine surface pellicle

Culture characteristics
Mac Conkeys Agar colourless at first,
become reddish on prolonged incubation due
to Late lactose fermentation

HOLDING OR TRANSPORT MEDIA

Venkataraman Ramakrishnan Medium pH: 8.6 -8.8
Cary Blair Medium pH: 8.4 Suitable for Salmonella, Shigella and Vibrio
Autoclaved Sea Water

ENRICHMENT MEDIA
Alkaline Peptone Water pH: 8.6
Monsurs taurocholate tellurite peptone water
pH: 9.2

PLATING MEDIA
TCBS Agar:
Thiosulfate/Citrate/Bile
salts/Sucrose Agar - pH:8.2
Most widely used plating
medium for Vibrios- yellow,
convex colonies
Alkaline Bile Salt Agar:
pH:8.2 - Simple and widely
used medium

VIBRIO CHOLERAE
COLONIES ON TCBS AGAR

PLATING MEDIA
(Contd)
Monsurs Gelatin Taurocholate Trypticase
Tellurite agar (GTTA):
Vibrios form small translucent colonies with a
greyish black centre and a turbid halo.

LABORATORY DIAGNOSIS OF CHOLERA (Contd)

String test
Biochemical tests
Cholera red reaction
Agglutination: O subgroup serum
Ogawa
Inaba

STRING TEST
A loopful of the growth is
mixed with 0.5% Sodium
deoxycholate in saline on a
slide.
Positive reaction: the
suspension loses its turbidity,
becomes mucoid and forms a
string when the loop is slowly
drawn away from the solution

BIOCHEMICAL REACTIONS

Catalase & Oxidase positive

Carbohydrate metabolism is
fermentative producing acid but no
gas.
Glucose, Sucrose, Maltose, Mannitol
are fermented
Lactose is fermented very slowly
(late lactose fermenter)
Indole Positive

BIOCHEMICAL REACTIONS (Contd)

Nitrates are reduced to nitrites
Cholera red reaction
Methyl red is Negative.
Gelatin is liquefied.

34
20

EPIDEMIOLOGY
Sporadic, endemic, epidemic, pandemic
1817-1923 six pandemics
1961-1994 Eltor pandemic-different from other pandemic
Non O1 in Oct 1992(O-139)
Carrier
Transmission

TREATMENT
Prompt replacement of Fluid and electrolytes
Tetracycline

PROPHYLAXIS
General
Improvement of environmental sanitation
Provision of protected water for drinking
Specific
Vaccines: Original-live suspension
Killed vaccine-Ogawa & Inaba
O-139 vaccine
Not very effective, immunity low
Oral vaccine-Killed oral whole vaccine
Live oral vaccines

HALOPHILIC VIBRIOS
V.parahaemolyticus- Food poisoning - Kanagawa
phenomenon positive strains are pathogenichemolytic on Wagatsuma agar (high salt blood
agar)
V.vulnificus- Septicaemia, Cellulitis, Acute
diarrhoea