This document discusses spore-forming gram-positive rods including Bacillus and Clostridium species. Key points include: - Bacillus and Clostridium are spore-forming, gram-positive rods that can be aerobic or anaerobic. Important medically are B. anthracis, B. cereus, C. botulinum, C. tetani, C. perfringens, and C. difficile. - Laboratory diagnosis involves culturing on blood agar and observing colony morphology, hemolytic patterns, and microscopic appearance including spore formation. Serological and molecular tests can also identify toxins. - Major diseases include anthrax, botulism,

1. Spore-Forming Gram + Rods
Bacillus & Clostridium

2.  Gram positive rods.
 Form endospores. (heat & cold resistance spores)
 Either strict aerobes or aerotolerant anaerobes (that
is, they can grow in the presence of O2 BUT don’t
require it).
 Two medically important bacillus spp. B.anthracis &
B.cereus.
 Most of the bacillus spp. Are found in soil, water and
as airborne contaminants in medical laboratory.
 Bacillus anthracis, cause Anthrax is most important
member of this genus.
General Characteristics of Bacillus

3.  On blood agar
• Large, spreading, gray-white colonies, with irregular
margins
• Many are beta-hemolytic (helpful in differentiating various
Bacillus spp. from B. anthracis)
 Spores seen after several days of incubation, but not
typically in fresh clinical specimens
Laboratory Characteristics of Bacillus

4. Bacillus anthracis

5. Properties of Bacillus anthracis
Large gram + rod.
Square ends, frequently found in chains.
Antiphagocytic capsule is composed of D-glutamate. (This
is unique capsules of other bacteria are polysaccharide).
Nonmotile, other members of the genus are motile.
Culture on blood agar.
3 forms of anthrax:
Cutaneous Painless ulcer with a black eschar.
pulmonary – inhalation of spores
gastrointestinal – ingested spores

6. Laboratory diagnosis
Cultured on blood agar, B.anthracis forms
large, grayish, nonhemolytic colonies with
irregular borders.
A direct immunofluorescence assay aids in
the identification of the organism.
Q. What’s the reason that Spore are usually not
seen in smears of exudate?

7. Laboratory diagnosis
Ans. Because spores form when nutrients are
insufficient and plentiful in infected tissue.
B.anthracis – gram stain. Arrow points to Skin lesion, a necrotic lesion covered

8. Edema and necrosis
around eye
Inhalation anthrax with
bilateral pneumonia, more
severe in left lower lung.

9. Bacillus anthracis
viewed under fluorescence
microscope

11. Bacillus cereus

12. Laboratory diagnosis
and
CLiniCaL features
Cause food poisoning.
Spores on grains such as rice.
Two symptoms a. Short incubation (4hrs)
const. nausea & vomiting. b. Long incubation
(18hrs) watery, nonbloody diarrhea.
Lab. Diagnosis not usually done.
Only symptomatic treatment is given.

13. Quest. Recognize anthracis VS cereus

15. Clostridium spp. characteristics
Gram positive.
Large rods that produce endospores.
Most spp. Are motile.
All clostridia are anaerobic.
Cultured on blood agar.

16. Clostridium botulinum
Cause botulism (food poisoning e.g.
canned meat).
There are eight immunologic type of
toxins (e.g. Botox)
Clinical findings: *flaccid paralysis,
diplopia (double vision), dysphagia and
respiratory muscle failure.
No fever are present.

17. Laboratory diagnosis
Usually organisms not cultured.
Use mouse protection test for botulinum toxin
detection, they’ll die unless protected by antitoxin. Or
by using ELISA assay.
Egg yolk emulsion agar

18. Clostridium tetani

19. Laboratory diagnosis and
CLiniCaL features
• Cause tetanus (lockjaw).
• Germination of spores favored necrotic tissue.
• Has one antigenic type (tetanospasm) unlike
botulinum has eight.
• Clinical findings: *Spastic paralysis, Risus
sardonicus & opisthotonos (back spasm) due
to rigid contraction of the jaw muscle.
• Diagnosis is based on clinical findings.
• Diazepam (valium) as a treatment to prevent
spasm.

20. Tetanus. Note the marked hyperextension
of the back, a position
called opisthotonus. Caused by tetanus
toxin, an exotoxin that inhibits the release
of mediators of the inhibitory neurons in
the spinal cord

21. Clostridium perfringens

22. Laboratory diagnosis and
CLiniCaL features
Cause two distinct disease, food poisoning & gas gangrene
(myonecrosis).
Associated with war wounds, automobile, motorcycle an
accidents and septic abortion (endometritis) inner layer of the
uterus.
Clinical findings pain, edema, cellulitis and necrosis (gangrene).
When cultured unaerobically produces a unique double zone of
B-hemolysis. Egg yolk agar used to demonstrate the presence of
the lecithinase (alpha toxin). Other biochemical test as sugar
fermentation (glucose) and organic acid production such as
acetic acid.

23. Gas gangrene. Note large area of necrosis
on lateral aspect of foot. Necrosis mainly
caused by lecithinase produced
by Clostridium perfringens. Gas in tissue
is a feature of gangrene produced by this
anaerobic bacteria. A large gas and fluid-
filled bulla is seen near the ankle.
Clostridium perfringens
Gram stain
Arrow points to a large gram-
positive rod

25. Clostridium difficile

26. Laboratory diagnosis and
CLiniCaL features
Cause antibiotic associated pseudomembranous
colitis. e.g. clindamycin, cephalosporin and ampicillin.
 Clinical findings: AAD antibiotic associated diarrhea, usually
nonbloody, fever and abdominal cramping often occur.
 Two types of tests usually used to detect the exotoxins, 1st
ELISA for stool samples. 2nd
PCR assay. The presence of a
pseudomembrane in the colon can be detected by
endoscopy.

27. Pseudomembranous colitis. Note
yellowish plaque-like lesions in
sigmoid colon that inhibits a
signal transduction protein,
leading to death of enterocytes.

28. Important Features of Pathogenesis by
Clostridium Species