A microbiological and Molecular perspective of Vibrio cholerae.

1. NU-JAHAT-JABIN
DEPARTMENT OF GENETIC ENGINEERING AND
BIOTECHNOLOGY.
UNIVERSITY OF CHITTAGONG

2. Vibrio cholerae &CHOLERA :
MICROBIOLOGICAL AND MOLECULAR PERSPECTIVE

3. CONTENT
 Introduction of Cholera.
 Epidemics and Pandemics of Cholera.
 Causative agent of Cholera.
 Cholera and V. cholerae in the view of microbiological
perspective.
 Molecular perspective of V. cholerae
 Possible Future Threat and Research
 References 3

4. CHOLERA; A BLUE DEATH
What is
Cholera?
Acute
Secretory
diarrheal
disease
Results rice
water stools
Agents
Gram
negative
bacteria
Vibrio
cholerae
Age
group
All age
group
Children <5
years are
more
susceptible.
4

5. MICROBIOLOGY OF V. cholerae
This section deals with-
The Causative agent of Cholera
Classification of Vibrio cholerae.
Virulence factors.
5

6. Vibrio Cholerae- THE CAUSETIVE AGENT
OF CHOLERA
• Gram negative
• Facultative anaerobic
• Non-Spore forming
Morphology
• 1.4 to 2.6µm long
• Coma shapedStructure
• ˜ 105 to108 bacteria is required for
successful infection
• Natural habitat is brackish or salt
water
Other
feature 6

7. CLASSIFICATION OF V. cholerae
Classified depending on “O” antigen
7
Non epidemic

8. SEROGROUPS OF V. CHOLERAE
8
 Produce Cholera toxin
(CT) and Toxin co
regulated pillus (TCP)
 Involve in epidemics
 Mechanism of
pathogenesis is known
 CT negative, but
contain toxin like Cholix
toxin.
 Involve in sporadic and
localized out break
 Pathogenic mechanism
is not clear.
O1 &O139 Non O1 & Non O139

9. EPIDEMIOLOGY OF CHOLERA
• Both epidemic and endemic
• Both seasonal and off seasonal
Pattern
• 1.3 to 4 million affected people.
• 143000 to 210000 deaths.
(WHO,2016)
Mortality
rate/year
• Poor sanitation.
• Transmission – either human to
human or environment to human
Reason
9

10. EPIDEMICS OF CHOLERA
10
Figure- Cholera outbreak , 2010 -2015

11. PANDEMICS OF CHOLERA
Fig-Historical change of population of V. cholerae .
11
Ref- Kim et al.,2015

12. 12
Biotype Phenotypes Genotype
CCA PB VP Phage
IV
Phage
5 tcpA rstR ctxB
Classic
al
- s - s r Cla Cla B1
El Tor + r + r s El El B3
PANDEMICS; CLASSICAL VERSES El TOR
Abbreviations: CCA- Chicken Cell Agglutination
PB – Polymixin
VP- Voges-Proskaure
Key : +, positive ; -, negative; s , sensitive; r, resistance
Ref- Safa et al,.2010

13. PANDEMICS; CLASSICAL VERSES El TOR
Classical – Ah my past !!!!
El Tor – He He, I am the new
king!!
13

14. VIRULENCE FACTORS
Biological
Function
Types
Virulence
factors
Virulence
factors
Cholera
Toxin
Hyper secretion
and loss of water,
electrolytes,
Also Act as
colonization
factors
Toxin
Coregulated
pillus (TCP)
Adherence
to mucosal
cell
tcpA
Responsible
gene
14
ctxA and
ctxB

15. OTHER VIRULENCE FACTORS INCLUDES
Factors Biological Function Responsible
gene
Accessory colonization
factors
Adhesion to mucosal cell acf
Hemagglutination
protease
Helps to dissolve
glycoprotein coating of
intestinal cell
Hap
Accessory cholera
enterotoxin
Release exotoxin ace
Other membrane protein Colonization assisting
mechanism
ompW
15

16. V. cholerae: MOLECULAR PERSPECTIVE
 Genome organization
 Cholera toxin
 Molecular mechanism of action
 Antibiotics resistance mechanism
 Evolution of new variants
16

17. GENOME ORGANIZATION
2 Circular chromosome.
The largest one is 2.4 Mb and the smallest one is about 1.6
Mb in length.
17
Ref- Heidelberg J.F et al., 2000

18. GENETIC ORGANIZATION OF CTXΦ
18Figure- Bacteriophage CTXφ organization
Ref- Safa et al., 2010

19. CHOLERA TOXIN
A subunit
A1 segment-ADP
ribosylates of G
protein
A2 Segment-
Facilitate binding A
with B subunit by
forming a alpha
helix
B subunit
Attachment of Toxin
to the host cell.
19
Figure- Structure of CT
Ref: Naomi LB et al.,

20. MOLECULAR MECHANISM OF ACTION
Figure- Molecular mechanism of actions of Cholera toxin
20

21. ANTIBIOTIC RESISTANCE MECHANISM
• Widespread and
inappropriate use of
antibiotics
Causes
• Major threat to public health
because it results treatment
failure
Concern
• For cholera treatment –
Azithromycine.
Drug of
choice 21

22. FIGHT AGAINST ANTIBIOTICS
22
Type of
Antibiotic
resistance
Intrinsic
mechanism
Efflux pump
Spontaneous
mutation
Acquired
mechanism
HGT

23. INHERENT RESISTANCE
23
Spontaneous
mutation
Cell wall
biosynthesis
inhibitor.
Alafosfalin
resistance
DNA
replication
inhibitor
Quinolone
resistance
Efflux pump Spontaneous mutation
Ref- Anderson et al,. 2015
Ref-Pukatzki et al., 2011

24. EVOLUTION OF V. CHOLERA
24
Ref- O’Shea et al., 2004.

25. UNDERSTANDING EVOLUTION BY USING
MOLECULAR FINGERPRINTING
Pre
genomic
era
• Pulsed Field Gel electrophoresis
• RAPD
• MLVA
Genomic
era
• Sanger Sequencing
• Pyrosequencing
Post
Genomic
era
• Second generation sequencing
(Illumina)
• Third Generation sequencing (Taqbio) 25
Ref- Rahman et al., 2015

26. FUTURE THREAT
Don’t ever think that I am
simple I can make big
TROUBLE!!
Strains of V. cholerae.
26

27. REFERENCES
 Safa, A., Nair, G.B. and Kong, R.Y., 2010. Evolution of new variants
of Vibrio cholerae O1. Trends in microbiology, 18(1), pp.46-54.
 Rahaman, M., Islam, T., Colwell, R.R. and Alam, M., 2015. Molecular
tools in understanding the evolution of Vibrio cholerae. Frontiers in
microbiology, 6, p.1040.
 Kim, E.J., Lee, C.H., Nair, G.B. and Kim, D.W., 2015. Whole-genome
sequence comparisons reveal the evolution of Vibrio cholerae
O1. Trends in microbiology, 23(8), pp.479-489.
 Mackay, D.M., 1980. Cholera: The present pandemic. Public
health, 94(5), pp.283-287.
 Li, W., Raoult, D. and Fournier, P.E., 2009. Bacterial strain typing in
the genomic era. FEMS microbiology reviews, 33(5), pp.892-916.
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28. REFERENCES
 O'shea, Y.A., Reen, F.J., Quirke, A.M. and Boyd, E.F., 2004.
Evolutionary genetic analysis of the emergence of epidemic Vibrio
cholerae isolates on the basis of comparative nucleotide sequence
analysis and multilocus virulence gene profiles. Journal of clinical
microbiology, 42(10), pp.4657-4671.
 Li, W., Raoult, D. and Fournier, P.E., 2009. Bacterial strain typing in
the genomic era. FEMS microbiology reviews, 33(5), pp.892-916.
 Banerjee, R., Das, B., Nair, G.B. and Basak, S., 2014. Dynamics in
genome evolution of Vibrio cholerae. Infection, Genetics and
Evolution, 23, pp.32-41.
 Reidl, J. and Klose, K.E., 2002. Vibrio cholerae and cholera: out of
the water and into the host. FEMS microbiology reviews, 26(2),
pp.125-139.
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