VIDEO LECTURE

BACTERIOLOGY - CHAPTER SIXTEEN  

MYCOBACTERIA AND CORYNEBACTERIA

 Dr Alvin Fox

 READING: Murray, 6th Edition Chapter 28 (mycobacteria) Chapter 26 (Corynebacterium diphtheriae)

KEYWORDS
Acid Fast 
Tuberculosis
M. tuberculosis
M. bovis 
Tubercle
PPD
Tuberculin 
Mycobactin
Cord factor
BCG
Mycolic acids
Atypicals
M. avium - M. intracellulare complex
Runyon groups
Leprosy (Hansen's Disease)
M. leprae
Diphtheria
C. diphtheriae
Loeffler's agar
Tellurite agar
Metachromatic bodies
Diphtheria toxin
Schick test
Diphtheroids
Legionella pneumophila
Legionnaires disease
Pontiac Fever
Lepromin
Legionella (Tatlockia) micdadei
Charcoal yeast extract agar
Fatty acid profiles
Hybridization

MYCOBACTERIA 

In the 1980's, many experts felt that the days of tuberculosis as a threat to the US population had passed and the incidence of new cases (around 20,000 a year) was slowly decreasing, even though it was still the leading infectious cause of death world-wide. The situation in the 1990's has changed dramatically. The incidence of tuberculosis has slightly increased and the disease is certainly not going away (This is primarily due to the AIDS epidemic). At the same time multiple drug-resistant strains of M. tuberculosis are appearing regularly. The M. avium - M. intracellulare complex, long considered a group of organisms that only rarely infects man, is now recognized as one of the leading opportunists associated with AIDS. M. leprae is the causative agent of leprosy which remains a major disease in the third world. Due to eradication of infected cattle and pasteurization of milk M. bovis (a zoonotic cause of tuberculosis) is rarely seen in the United States.

Mycobacteria are obligate aerobic, acid-fast rods.

Pathogenesis of tuberculosis

Mycobacterium tuberculosis infects the lung, and is distributed systemically within macrophages and survives intracellularly. Inhibition of phagosome-lysosome fusion and resistance to lysosomal enzymes have both been suggested to play a role. Cell-mediated immunity develops which causes infiltration of macrophages and lymphocytes with development of granulomas (tubercles). The disease can be diagnosed by skin testing for delayed hypersensitivity with tuberculin (also know as protein purified purified from Mycobacterium tuberculosis, PPD). A positive test does not indicate active disease; merely exposure to the organism.

Other pathogenesis factors (of considerably less importance than delayed hypersensitivity) include mycobactin (a siderophore) and cord factor which damages mitochondria.

Diagnosis, identification and treatment

The presence of acid fast bacteria in sputum is a rapid presumptive test for tuberculosis. Subsequently, when cultured, M. tuberculosis will grow very slowly producing distinct non-pigmented colonies after several weeks. M. tuberculosis can be differentiated from most other mycobacteria by the production of niacin. A rapid alternative to culture is polymerase chain amplification (PCR).

Tuberculosis is usually treated for extensive time periods (9 months or longer) since the organism grows slowly and may become dormant. By using two or more antibiotics (including rifampin and isoniazid), the possibility of resistance developing during this extended time is minimized. M. tuberculosis causes disease in healthy individuals and is transmitted man-man in airborne droplets.

Vaccination

The BCG vaccine (Bacillus de Calmette et Guerin, an attenuated strain of M. bovis) has not been effective. In the US, where the incidence of tuberculosis is low, widespread vaccination is not practiced. Indeed immunization (resulting in a positive PPD test) is felt to interfere with diagnosis.

VIDEO
Tuberculosis
patients - symptoms - treatment - prevention - research - parasites
List of sequences
WHO-RealVideo

WEB RESOURCES

CDC Tuberculisis FAQ
 CDC en Español - Tuberculosis 
CDC Major TB guidelines 
WHO Tuberculosis

Estimated HIV/TB co-infection rates, 2008 WHO

Atypicals

The "atypicals" generally infect the immunocompromised host and are thus not transmitted man-man. With the AIDS epidemic, the atypical mycobacteria have taken on new importance with the recognition that the M. avium complex (MAC) results in the most commonly associated systemic bacterial infection. Atypical mycobacteria can cause tuberculosis-like or leprosy-like, diseases, and are not susceptible to certain common anti-tuberculous antibiotics.

Mycobacterium avium. Rod-shaped Bacterium (causes avian tuberculosis)  (SEM x24,000)  

Mycobacterium avium-intracellulare infection of lymph node in patient with AIDS. Ziehl-Neelsen stain. Histopathology of lymph node shows tremendous numbers of acid-fast bacilli within plump histiocytes. CDC/Dr. Edwin P. Ewing, Jr. 

M. avium generally infects AIDS patients when their CD4+ cell count decreases greatly (below 100/mm³). M. tuberculosis infects AIDS patients much earlier in the disease. This clearly demonstrates the much greater virulence of M. tuberculosis. The incidence of systemic disease (versus primarily pulmonary) is much greater in tuberculosis associated with AIDS than in its absence. Furthermore, histologically lesions often appear lepromatous (non-granulomatous, with many organisms). It is rare to find a case of M. avium infection that is not AIDS associated. However, M. tuberculosis is a much more virulent organism. Approximately 20% of the total tuberculosis cases in the US are caused by AIDS. This helps explain why TB is no longer on the decline. Increased homelessness is also suggested to be a factor in the rise of tuberculosis.

Treatment of M. avium also involves a long-term regimen of multiple drug combinations. However, this organism does not always respond to the drug regimens used to treat M. tuberculosis. Appropriate drug combinations are still under investigation in clinical trials. Since M. tuberculosis is the more virulent organism, the drug regimen selected is primarily against M. tuberculosis. If M. avium is suspected other agents effective against this organism are included.

Presence or absence of pigmentation (and its dependence on growth in the light) and slow or fast growth rates of atypical mycobacteria allow some differentiation - the "Runyon Groups". Modern techniques allow ready speciation of mycobacteria based on their cellular fatty acid and/or mycolic acid profiles. This is only performed in reference laboratories. Mycolic acids are components of a variety of lipids found only in mycobacteria, nocardia and corynebacteria. The chain length of these mycolic acids is longest in mycobacteria, intermediate in nocardia and shortest in corynebacteria. This explains why mycobacteria are generally acid fast; nocardia less acid fast; and corynebacteria are non-acid fast.

Global Leprosy Situation 2009 © World Health Organization

Leprosy: New case detection rate 2009
© World Health Organization

Mycobacterium leprae

M. leprae is the causative agent of leprosy (Hansen's Disease), a chronic disease often leading to disfigurement.. It is rarely seen in the U.S. but common in the third world. The organism infects the skin, because of its growth at low temperature. It also has a strong affinity for nerves. In "tuberculoid" leprosy, there are few organisms due to control by active cell-mediated immunity. In "lepromatous" leprosy, due to immunosuppression by the organism, the opposite is found. Although uncommon in the U.S., millions of cases occur worldwide. Treatment with antibiotics (initially dapsone and now multi-drug) is effective and the overall disease incidence worldwide is down. The organism does not grow in culture media. However, it grows well in the armadillo (which has a low body temperature), allowing production of M. leprae antigens and pathogenesis studies. M. leprae has traditionally been identified on the basis of acid-fast stains of skin biopsies and clinical picture. Lepromin is used in skin testing. 

Azadegan Clinic, Teheran: The foot of a woman that has been grossly disfigured through leprosy infection. © World Health Organization/TDR/Crump)

Deformity due to nerve damage with its consequent ulcers and resorption of bone. Such deformities can be worsened by careless use of the hands. © WHO/TDR

The face of a Peruvian man with active lepromatous leprosy. © WHO/ TDR/ McDougall

A young girl, 8 years old, with Burmese-Scots ancestry. The loss of eyebrows is an indication of diffuse lepromatous leprosy ©WHO/ TDR/ McDougall

The face of a patient with active, neglected nodulous lepromatous leprosy. With treatment, all nodules could be reversed. ©WHO/TDR/McDougall

The trunk of a patient showing abnormal lesions of dapsone-resistant leprosy ©WHO/TDR/McDougall

Most patients with leprosy can be cured with multi-drug therapy in just six months as shown in this image © WHO/ TDR

WEB RESOURCES

 Leprosy WHO Fact Sheet 

WHO Action Programme for the Elimination of Leprosy 

Advances in the diagnosis and treatment of leprosy 
Expert Reviews in Molecular Medicine

VIDEO
Leprosy
patients - symptoms - treatment - prevention - research - parasites
List of sequences
WHO-RealVideo

  10-year-old child with severe diphtheria. CDC/NIP/Barbara Rice 

This child has diphtheria resulting in a thick gray coating over back of throat. This coating can eventually expand down through airway and, if not treated, the child could die from suffocation  CDC

CORYNEBACTERIA

C. diphtheriae grows best under strict aerobic conditions It is Gram positive and pleomorphic.

Colonization of the upper respiratory tract (pharynx and nose) and less commonly skin with C. diphtheriae can lead to diphtheria. The organism does not produce a systemic infection. However, in addition to a pseudomembrane being formed locally (which can cause choking), systemic and fatal injury results primarily from circulation of the potent exotoxin (diphtheria toxin). The latter begins over a period of a week. Thus treatment involves rapid therapy with anti-toxin. The gene for toxin synthesis is encoded on a bacteriophage (the tox gene). Corynebacteria that are not infected with phage, thus do not generally cause diphtheria. Diphtheria is now a disease of almost historic importance in the U.S. due to effective immunization of infants (in conjunction with pertussis and tetanus, DPT) with a toxoid (inactive toxin) which causes production of neutralizing antibodies. However, colonization is not inhibited and thus C. diphtheriae is still found in the normal flora (i.e. a carrier state exists). Immunity can be monitored with the Schick skin test. Treatment in non-immune individuals primarily involves injection of anti-toxin. Antibiotics are also administered at this time.

The toxin consists of two types of polypeptide. One binds to host cells; the other then becomes internalized and inhibits protein synthesis. The exotoxin catalyses the covalent attachment of the ADP-ribose moiety of NADH to a rare amino acid, diphthamide, present in EF2 (elongation factor 2). The toxin is not synthesized in the presence of iron as an iron-repressor complex forms which inhibits expression of the tox gene.

C. diphtheriae are identified by growth on Loeffler's medium followed by staining for metachromatic bodies (polyphosphate granules, Babes-Ernst bodies). The term "metachromatic" refers to the color difference of the intracellular polyphosphate granules (pink) compared to the rest of the cell (blue). Characteristic black colonies are seen on tellurite agar from precipitation of tellurium on reduction by the bacteria. Production of exotoxin can be determined by in vivo or in vitro tests.

Other organisms which morphologically resemble C. diphtheriae ("diphtheroids" which include other corynebacteria and also propionibacteria) are found in the normal flora. Isolates should not be confused with these organisms.

WEB RESOURCES

CDC Manual: Diphtheria (pdf)

Return to the Bacteriology Section of Microbiology and Immunology On-line

 

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