lab diagnosis for third medical student

1. GRAM POSITIVE BACILLI

3. Clinically important Gram positive bacilli
Spore forming
1. Bacillus
2. Clostridium
Non spore forming
1.Corynebacterium
2.Listeria
3.Lactobacillus
Bacilli w/ branching
filaments
1.Actinomyces
2.Nocardia

4. 1- Spore-Forming Gram-Positive Bacilli:
A- Strict aerobic – Genus Bacillus.
B- Strict anaerobic – Genus Clostridium.
Spore- forming gram positive strict aerobic capsulated bacilli, 1x3-4 u size,
arranged in long chains;
• Highly resistant spores
spores may central, subterminal or terminal, depending on the species,seen as
unstained, usually oval spaces, one in each stained mother cell
(sporangium),
• In vitro – prominent capsule(A unique anti-phagocytic capsule is
composed of D-glutamate.
.

5. Most members are saprophytic prevalent in soil,
water, air and vegetation such B. cereus,
and B. subtilius.
Bacillus anthracis non motile while Bacillus
cereus motile.
Most strain
• Catalase positive
• indole negative,
• simmon citrate variable.

6. • Bacillus anthracis
– Human pathogen
– Isolation also considered to be clinically significant
– Zoonosis
• Bacillus cereus
– Environmental organism
– Contaminates food
– Common cause of food poisoning
• Bacillus stearothermophilus
– Tolerates very high temperatures
– Used for quality control of autoclaves
 Bacillus subtilis common lab contaminant

7. Cause the disease Anthrax in animals in which the organism is
transmitted through eating vegetations containing the spores. Human
is infected through contact with animals or their products.
Disease in Human
A- Cutaneous Anthrax(malignant pustule): Generally occurs on exposed surfaces
of the arms, face and neck through wound contamination by the spores of the
organism. About 95% of the cases with a mortality rate 20% .
B- Inhalation Anthrax(wool A sorter disease): About 5% of the cases with 85-90%
mortality.
C- Gastrointestinal Anthrax: Is very rare.

8. Anthrax
Pathogenesis and clinical presentations
Cutaneous anthrax
About 20% mortality
Inhalation anthrax
High mortality
Virulence factors
Capsule
(antiphagocytic)
Toxin
(oedema & death)
Gastrointestinal anthrax
High mortality

9. Anthrax Has Been Used As a Bioweapon
Because it is deadly, noncontagious, and
dispersed by spores, anthrax has always
been considered a good candidate for a
bioweapon. Late in 2001, this possibility
became a reality. Letters containing anthrax
spores were sent to several news reporters
and two United States Senators. Five people
died of inhalational anthrax as a result of
exposure to these spores.

11. Anthrax - Diagnosis
• Specimen
– Aspirate or swab from cutaneous lesion
– Blood culture
– Sputum , Fluid , pus,
• Laboratory investigation
– Gram stain
– Culture
– Identification of isolate

12. Diagnostic Laboratory Tests
Culture on blood agar.
• Gram Stained smears from the local lesion or of blood from dead animals often
show chains of large gram-positive rods.
• Anthrax can be identified in dried smears by immunofluorescence staining
techniques.( Spores not seen in smears of exudate).
• When grown on blood agar plates, the organisms produce nonhemolytic gray to
white colonies with a rough texture and a ground-glass appearance.
• Comma-shaped outgrowths (Medusa head) may project from the colony.
• . In semisolid medium, anthrax bacilli are always nonmotile, whereas related
nonpathogenic organisms (eg, B cereus) exhibit motility by "swarming.“
• Carbohydrate fermentation is not useful
• Virulent anthrax cultures kill mice or guinea pigs upon intraperitoneal injection.
• Demonstration of capsule requires growth on bicarbonate-containing medium in 5–
7% carbon dioxide.
• An enzyme-linked immunoassay (ELISA) has been developed to measure
antibodies against edema and lethal toxins, but the test has not been extensively
studied.

13. Bacillus anthracis.
Gram stain. a "bamboo
appearance”
The cells have characteristic squared ends.
The endospores are ellipsoidal shaped and located centrally in the
sporangium.
The spores are highly refractile to light and resistant to staining.

17. Laboratory Diagnoses

22. Key Characteristics to Distinguish between
B. anthracis & Other Species of Bacillus
Characteristic Bacillus anthracis
Hemolysis
Neg
Motility
Neg
Gelatin hydrolysis
Neg
Other Bacillus spp.
Pos
Pos (usually)
Pos

23. Gram-Variable Stain of B. cereus
with Endospores

24. Cause food poisoning: Two types
1- Emitic type associated with fried rice.
2- Diarrheal type associated with meat dishes and
sauces
Infections in the immunosuppressed hosts
Opportunistic infections of the eye
Meningitis, septicemia, and osteomyelitis
Found as contaminants in drug paraphernalia
Bacillus cereus

25. Bacillus cereus
clinical presentation
EMETIC FORM
Incubation period < 6 hours
Severe vomiting
Lasts 8-10 hours
Gastroenteritis
DIARRHOEAL FORM
Incubation period > 6 hours
Diarrhoea
Lasts 20-36 hours

26. Foodborne Diseases of B. cereus
(Intoxication)
(Foodborne Infection)

27. b.Bacillus cereus
•
•
•
•
Large, motile, saprophytic bacillus
Heat resistant spores
No capsule
Pre formed heat and acid stable toxin
(Emetic syndrome)
• Heat labile enterotoxin (Diarrhoeal disease)
• Lab diagnosis – Demonstation of large
number of bacilli in food

28. Laboratory Identification: Bacillus
anthracis
Characteristics
B. anthracis
B.cereus
Hemolysis on
BAP
Motility
=
+
=
+
String of pearls
+
=
Growth on PEA
=
+
Gelatin hydrolysis
=
+
Susceptibility to
Penicillin (10U/ml)
Susceptible
Resistant

29. Other Bacillus spp.
Bacillus stearothermophilus
• Spores used to test efficiency of killing in autoclaves

30. Other Bacillus species
• Bacillus subtilis
– Common laboratory contaminant

33. Points to Remember
• Differentiating characteristics between B. anthracis and
saprophytic species
• Characteristic, microscopic appearance of Bacillus species
• Clinical forms of infection attributed to B. anthracis
• Clinical significance of Bacillus cereus in healthy as well as
in at-risk populations