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

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 H 2 S 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

- 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.