Corynebacterium & Bacillus

- Microscopic appearance

- Colonial morphology

Corynebacterium diphtheriae

- Nasal, nasopharyngeal and tonsillar

diphtheria.

Corynebacterium diphtheriae

- Cutaneous diphtheria.

Corynebacterium diphtheriae

- Throat, nasopharyngeal swabs.

- Skin swab.

Corynebacterium diphtheriae

- Gram positive pleomorphic, long, thin, and curved forms can be seen and also short rods and rods enlarged at one end (clubshaped).

Corynebacterium diphtheriae

- They often appear in clusters, joined at angles like Chinese letters.

- C. diphtheriae often appears beaded due to the

presence

of dark staining granules in the rods.

-These granules, known as volutin or metachromatic

granules, are energy-storing inorganic

polyphosphate

units. In some strains the granules form at the ends

of

the rods.

- The granules are most numerous after the organism

has

been cultured on a protein-rich medium such as

Dorset

egg or Loeffler serum.

Albert Staining of volutin granules

Albert Staining of volutin granules

1- Fix the dried smear using alcohol.

2- Cover the smear with the toluidine blue

malachite green stain for 3–5 minutes.

3-Wash off the stain with clean water.

4- Tip off all the water.

Albert Staining of volutin granules

5- Cover the smear with Albert’s iodine for

1 minute. Wash off with water.

6- Wipe the back of the slide clean, and place it

in

a draining rack for the smear to air-dry.

7- Examine the smear microscopically to look

for

bacteria containing metachromatic granules

Albert Staining of volutin granules

Bacteria cells . . . . . . . . . . . . . . . . . . . . . . Pale green

Metachromatic granules . . . . . . . . . . . . Green-black

Corynebacterium diphtheriae

- They often appear in clusters, joined at angles like Chinese letters.

Tinsdal medium:- grey-black raised colonies surrounded by a dark brown area.

-Brown colour due to formation of H2S which result from interaction between cystine and tellurite.

C. diphtheriae grows rapidly on these media, producing significant growth in 4–6 hours.

Bacillus

Bacillus anthracis

Bacillus anthracis

- Cutaneous ANTHRAX.

Bacillus anthracis

- Pulmonary ANTHRAX.

Bacillus anthracis

- Enteric ANTHRAX.

- Meningoencephalitis.

B. anthracis is a high risk infectious

pathogen, therefore handle specimens

and infected material with care, wearing

protective gloves and face mask, and

following recommended safety

procedures.

Bacillus anthracis

- Fluid aspirated from cutaneous lesions.

- Sputum.

- Blood for culture.

- CSF.

large, 5–8 X 1.5 μm, Gram positive, non-motile bacillus, often appearing joined end to end in chains

In smear from Specimens:- Bacilli are capsulated. The capsular material often appears irregular and

fragmented

In smears from aerobic cultures: Bacilli are non-capsulated

but contain oval spores (same diameter as the bacilli), giving the

organisms a beaded appearance.

They occur in chains.

Fixation of smears:-

B. anthracis is not killed by heat-fixation.

Smears should be chemically fixed by

immersing the dry smears in a container

of potassium permanganate solution for

10–15 minutes.

Bacillus anthracis

Blood agar:- large grey-white 2-5 mm in diameter

irregular with wavy edges colonies (non or slightly

haemolytic).

Bacillus cereus

- Food poisoning.

- Opportunistic infections in

immunocompromised

persons (bacteraemia, pnumonia and

wound

infection).

Bacillus cereus

Blood agar:- large grey-white 2-5 mm in diameter

irregular with wavy edges haemolytic colonies.

B.cereus rapidly liquefies the gelatin along and out from

the line of inoculation.