2. INTRODUCTION
Anthrax (splenic fever), is an acute
infectious disease caused by the
bacteria Bacillus anthracis.
Highly lethal in some forms.
Primarily a disease of domesticated
and wild animals, particularly
herbivores.
Humans become infected when
brought into contact with diseased
animals.
3. Bacillus anthracis
Gram-positive rods with square end
shape, tending to form long chains
(Bamboo cane like)
Non motile
1 - 1.2µm in width x 3 - 5µm in length.
Originates from the Greek word
anthrakis ( (ἄνθραξ), meaning coal.
Normally rest in spore form in the soil,
and can survive for decades in this state.
The spores are ellipsoidal, located in the
middle of the vegetative form, without
enlarging the original form.
Weapon of bioterrorism.
4. TYPES OF ANTHRAX
Cutaneous (skin) anthrax
Inhalational anthrax(also known
pulmonary anthrax)
Gastrointestinal anthrax
8. CONVENTIONAL METHOD
Direct Gram’s-stained smear of a skin
lesion (vesicular fluid or eschar), csf, or
blood showing encapsulated, broad,
gram-positive bacilli.
◦ - Indicators of growth apparent on
sheep’s-blood–agar cultures —
nonhemolytic colonies and large,
nonmotile, nonencapsulated, gram-
positive, spore-forming
◦ rods. Growth does not occur on
MacConkey agar.
9. ◦ Confirmatory diagnostic tests are
performed at a level B laboratory of the
Laboratory Response Network for
Bioterrorism (LRN), where the growth
of
◦ virulent strains on nutrient agar in the
presence of 5 percent carbon dioxide
(or other basal mediums supplemented
with 0.8 percent sodium bicarbonate)
◦ produces heavily encapsulated bacilli
that may be visualized
◦ with India-ink staining.
10. - Susceptibility to lysis by gamma phage
or
◦ - direct fluorescence-antibody staining
of cell-wall polysaccharide
◦ antigen
11.
12. B. anthracis is a non-fastidious
organism and can grow on simple
laboratory media. They are facultative
anaerobes. The optimum temperature
for growth is 37°C and the pH, 7.0–7.4.
13. CULTURE
CHARACTERISTICS
Nutrient agar- After overnight
incubation at 37oC colonies are large
2–3 mm in diameter, irregular, raised,
dull, opaque and grayish white with
‘frosted glass’ (ground glass)
appearance
Sheep blood agar- On sheep blood
agar the colonies are nonhaemolytic
2-3mm in diameter
14.
15. SELECTIVE MEDIA
PLET medium- PLET (polymyxin-
lysozyme-EDTA-thallous acetate) is
a selective media used for the
isolation of Bacillus anthracis from
contaminated specimens. PLET Agar
inhibits most contaminating organisms
and spore-formers closely related to
B.anthracis , such as B. cereus
16. It consists of heart infusion agar with
polymyxin, lysozyme, ethylene diamine
tetra acetic acid (EDTA) and thallous
acetate. After incubation at 37°C for 36–
48 hours, the colonies of B. anthracis
are 1–3 mm, roughly circular, creamy
white with ground-glass texture
17.
18. SPECIAL FEATURES
Susceptibility to penicillin G- B.
anthracis is almost always susceptible to
penicillin, as shown by susceptibility to
penicillin G 10 units discs on Mueller-
Hinton agar. In contrast the non-
pathogenic Bacillus species are more
generally resistant to penicillin.
Susceptibility to gamma bacteriophage-
Gamma Phage has the ability to lyse B.
anthracis grown aerobically on blood or
other nutrient agar and rarely lyses any
other Bacillus species.
19. MOLECULAR METHOD
- In cases of cutaneous anthrax, antibodies to
protective antigen or to the capsule develop
in 68–92% of Patients.
- In one study of 12 patients with confirmed
cutaneous anthrax, 11 had a positive titre (>
1/128) to protective antigen by
electrophoretic immunotransblotting and
- 11 were positive (cut off point, 1/32) for
anticapsule antibodies as measured by
enzyme linked immunosorbent assay.
Samples were taken six weeks after
development of the disease
20.
21. PATHOGENESIS
Major virulence factors are encoded
on two virulence plasmids, pXO1 and
pXO2.
pX01:
184.5 kilobase pairs (kbp) in size
codes for the genes that make up the
secreted exotoxins.
82.7 kDa protective antigen (PA)
90.2 kDa lethal factor (LF)
88.9 kDa oedema factor (OF)
22. pXO2:
95.3 kbp in size
Codes for three genes capB, capC,
and capA
Capsule inhibits phagocytosis of
vegetative anthrax bacilli
23. A skin test using an extract from an
attenuated strain of B.anthracis is
available and is diagnostic in 82% of
patients 1 to 3 days and in 99% of
patients 4 weeks after the onset of
symptoms.
24. PCR METHOD
Sample DNAs were used as template in a
PCR containing primers that
amplify portions of the B. anthracis capA,
capB, capC, cya, lef, and pag genes.
These genes are located on the two large B.
anthracis plasmids, pX01 and pX02, required
for pathogenicity
Samples were also analyzed by using
primers that amplify chromosomal B.
anthracis-specific DNA sequences.