corynibacterium

Dr. Huda H. Al-Hasnawy

Gram-Positive Irregular (shape and staining) Non-Spore-Forming Bacilli
1- Corynebacterium:
The non-spore-forming gram-positive bacilli are a diverse group of bacteria. Many members of them are part of the normal flora of skin and mucous membranes of humans. Others are found in animals and plants. Corynebacterium diphtheriae is the most important member of the group.
Corynebacteria are small, slender, pleomorphic, gram-positive rods of distinctive morphology. They are non motile, unencapsulated, and do not form spores. Most species are facultative anaerobes, and those found associated with humans, including the pathogen C. diphtheriae, grow aerobically on standard laboratory media such as blood agar.

Corynebacterium diphtheriae
Diphtheria, caused by C. diphtheriae, is an acute respiratory or cutaneous disease and may be a life-threatening illness. The development of effective vaccination protocols and widespread immunization begin in early childhood has made the disease rare in developed countries. Pathogenicity of diphtheria bacilli is dependent on its ability to produce a powerful exotoxin.

Laboratory identification:
The presumptive diagnosis and treatment must be based on initial clinical observation. Diphtheria should be considered in patients with pharyngitis, low-grade fever, and cervical adenopathy (swelling of the neck). Erythema of the pharynx progressing to adherent gray pseudomembranes increases suspicion of diphtheria.
Definitive diagnosis requires isolation of the organism, which must then be tested for its virulence using an immunologic precipitation reaction to demonstrate toxin production by Elek’s test (in vitro test).
C. diphtheriae can be isolated most easily by a selective medium, such as blood-tellurite medium (Tinsdale agar), which contains potassium tellurite, as an inhibitor of other respiratory flora, and on which the organism produces several distinctive gray-black colonies with halos, also grows on L?ffler s serum medium.
C. diphtheriae from clinical material or culture has a distinctive morphology when stained, for example, with methylene blue. This morphology includes 0.5–1µm in diameter and several micrometers long, with characteristic bands and reddish metachromatic granules. Characteristically, they possess irregular swellings at one end that give them the "club-shaped" appearance.
schick test (in vivo test) is used to a certain population at risk. This test involves the injection of a minute amount of the diphtheria toxin under the skin. The absence of a reaction indicates immunity.
Specimens: swabs from the nose, throat or suspected skin lesions (avoid rupture of the membrane).

Diphtheroids:
Several other corynebacterium species that morphologically resemble
C. diphtheriae, are common commensals of the nose, throat, nasopharynx, skin, urinary tract, and conjunctiva. They are therefore called diphtheroids, and are generally unable to produce exotoxin, but a few cause diseases in rare circumstances, such as in immunosuppressed individuals.

2- Mycobacteria (Acid-Fast Bacilli/AFB):
The mycobacteria are rod-shaped, aerobic bacteria that do not form spores, with lipid-rich cell walls that are resistant to penetration by chemical dyes, such as those used in the Gram stain. They do not stain readily, once stained they resist decolorization by acid or alcohol and are therefore called "acid-fast" bacilli. Mycobacterium tuberculosis causes tuberculosis and is
a very important pathogen of humans, and a leading cause of worldwide death. Mycobacterium leprae causes leprosy. Mycobacterial infections are intracellular, and generally result in the formation of slow-growing granulomatous lesions that are responsible for major tissue destruction. This organism has increasingly become a cause for special concern in immunocompromised patients.

M. tuberculosis
Mycobacteria are long, slender, nonmotile, non-spore forming and strict aerobic rods. Mycobacterial cell walls are unusual in that they are composed of approximately 60 % lipid, including a unique class of very long-chain (75 to 90 carbons), hydroxylated fatty acids (mycolic acids). These complex with
a variety of polysaccharides and peptides, creating a waxy cell surface that makes mycobacteria strongly hydrophobic, and accounts for their acid-fast staining characteristic.
Their unusual cell walls make mycobacteria impermeable to many chemical disinfectants, and result in resistance to the corrosive action of strong acids or alkalis. Mycobacteria are also resistant to drying, but not to heat or ultraviolet irradiation. Most species grow slowly with generation times of 8 to 24 hours.
Laboratory identification:
Early diagnosis and initiation of effective therapy of TB is very important in reducing morbidity and mortality and minimize the spread of infection. Diagnosis of active pulmonary tuberculosis includes demonstration of clinical symptoms and abnormal chest radiographs, and confirmation by isolation of
M. tuberculosis from relevant clinical material.

Screening for prior infection: which is done for high prevalence and high-risk population (HIV) by the use of Mantoux tuberculin test (ie, purified protein derivative or PPD), and determine maximum diameter of induration by palpation.
• Tuberculin skin testing useful for
– Examining person who is not ill but may be infected.
– Determining how many people in group are infected.
– Examining person who has symptoms of TB.

Microscopic Exam: Microscopy and culture are available and appropriate technology, so detection of AFB in stained smears is the easiest & most rapid procedure for evaluating a clinical specimen after staining with Ziehl-Neelsen method (rapid, high specificity and give accurate diagnosis). However, M. tuberculosis cannot be reliably distinguished on morphologic grounds from other occasional pathogens in the genus, from some saprophytic mycobacterial species that may contaminate glassware and reagents in the laboratory, or from those mycobacteria that may be part of the normal flora. Therefore, a definitive identification of M. tuberculosis can only be obtained by culturing the organism on egg-potato based media=Lowenstein-Jensen media (LJ) and result appears after 3-6 wks because of its slow growth on laboratory media.
Additionally, diagnosis can be done by using one of the newer molecular methods such as Polymerase Chain Reaction (PCR) as it has the potential to shorten the time required to detect and identify M. tuberculosis in clinical specimens.

Specimens: consist of fresh early morning sputum (3 samples obtained at least eight hours apart with at least one sample obtained in the early morning), gastric washings, urine, pleural fluid, CSF, joint fluid, biopsy material, blood, or other suspected materials.

Mycobacterium leprae
Causes leprosy, a chronic disease that begins in the skin and mucous membranes and progresses into nerves and known as Hansen’s bacillus or Hansen’s Disease. It is strict parasite – has not been grown on artificial media or tissue culture with slow growth multiplies within host cells in large packets. There are more than 10 million cases of leprosy, mainly in Asia.
Specimens:
Scrapings with a scalpel blade from skin or nasal mucosa or from a biopsy of earlobe skin are smeared on a slide and stained by the Ziehl-Neelsen technique. No serologic tests are of value.

Laboratory identification: M. leprae is an acid-fast bacillus. It has not been successfully maintained in artificial culture, but can be grown in the footpads of mice. Laboratory diagnosis of lepromatous leprosy, where organisms are numerous, involves acid-fast stains of specimens from nasal mucosa or other infected areas. In tuberculoid leprosy, organisms are extremely rare, and diagnosis depends on clinical findings and the histology of biopsy material.