Corynebacterium Diphtheria - Lecture Notes PDF

Medical bacteriology / Lecture 5 / Dr. Sahand K Arif  Corynebacteria are Gram-positive, aerobic, nonmotile, rod-shaped bacteria related to the Actinomycetes.  They do not form spores or branch as do the actinomycetes, but they have the characteristic of forming irregular sh...

Medical bacteriology / Lecture 5 / Dr. Sahand K Arif  Corynebacteria are Gram-positive, aerobic, nonmotile, rod-shaped bacteria related to the Actinomycetes.  They do not form spores or branch as do the actinomycetes, but they have the characteristic of forming irregular shaped, club-shaped or V-shaped arrangements in normal growth.  They undergo snapping movements just after cell division which brings them into characteristic arrangements resembling Chinese letters. The "barred" appearance is due to the presence of polyphosphate inclusions called metachromatic granules.  The genus Corynebacterium consists of a diverse group of bacteria including animal and plant pathogens, as well as saprophytes. Some corynebacteria are part of the normal flora of humans,  Three strains of Corynebacterium diphtheriae are recognized, gravis, intermedius and mitis.  They are listed here by falling order of the severity of the disease that they produce in humans.  All strains produce the identical toxin and are capable of colonizing the throat.  The differences in virulence between the three strains can be explained by their differing abilities to produce the toxin in rate and quantity, and by their differing growth rates.  gravis strain has a generation time (in vitro) of 60 minutes; the  intermedius strain has a generation time of about 100 minutes; and  mitis stain has a generation time of about 180 minutes.  The faster growing strains typically produce a larger colony on most growth media.  In the throat (in vivo), a faster growth rate may allow the organism to deplete the local iron supply more rapidly in the invaded tissues, thereby allowing earlier or greater production of the diphtheria toxin.  Corynebacterium diphtheriae is spread by droplets, secretions, or direct contact. In situ lysogenic conversion of nontoxigenic strains to a toxigenic phenotype has been documented. Infection is spread solely among humans, although toxigenic strains have been isolated from horses. Asymptomatic nasopharyngeal carriage is common in regions where diphtheria is endemic. the diphtheritic lesion is often covered by a pseudomembrane composed of fibrin, bacteria, and inflammatory cells. Diphtheria toxin can be proteolytically cleaved into two fragments: an N- terminal fragment A (catalytic domain), and fragment B (transmembrane and receptor binding domains). Fragment A catalyzes the NAD+-dependent ADP-ribosylation of elongation factor 2, thereby inhibiting protein synthesis in eukaryotic cells. Fragment B binds to the cell surface receptor and facilitates the delivery of fragment A to the cytosol. pseudomembrane bull's neck  Two factors have great influence on the ability of Corynebacterium diphtheriae to produce the diphtheria toxin:  (1) low extracellular concentrations of iron  (2) the presence of a lysogenic prophage in the bacterial chromosome.  The gene for toxin production occurs on the chromosome of the prophage, but a bacterial repressor protein controls the expression of this gene.  The repressor is activated by iron, and it is in this way that iron influences toxin production. High yields of toxin are synthesized only by lysogenic bacteria under conditions of iron deficiency.  DiphtheriaToxin u Blocks protein synthesis u Protein 63Kd u controlled by Tox gene u lysogenic phage Beta-corynephage u expressed if [iron] low u 2 components A-B  The A subunit then terminates host cell protein synthesis by inactivating elongation factor-2 (EF-2) which is required for translocation of polypeptidyl- transfer RNA from the acceptor to the donor site on the eukaryotic ribosome. Toxin Fragment A inactivates EF-2 by catalyzing a reaction that yields free nicotinamide plus an inactive adenosine diphosphate-ribose-EF-2 complex (ADP- ribosylation).  For primary isolation, a variety of media may be used: Loeffler's serum medium Tinsdale tellurite agar  diphtheritic lesions are often covered with a pseudomembrane, Tinsdale agar  The most common in vitro assay for toxigenicity is the Elek immunodiffusion test  This test is based on the double diffusion of diphtheria toxin and antitoxin in an agar medium.  A sterile, antitoxin-saturated filter paper strip is embedded in the culture medium, and C diphtheriae isolates are streak-inoculated at a 90° angle to the filter paper.  The production of diphtheria toxin can be detected within 18 to 48 hours by the formation of a toxin-antitoxin precipitin band in the agar.  Alternatively, many eukaryotic cell lines (e.g., African green monkey kidney, Chinese hamster ovary) are sensitive to diphtheria toxin  Corynebacterium urealyticum Urinary tract infections (UTI’s); rare but important Urease hydrolyzes urea; release of NH4+, increase in pH, alkaline urine, renal stones  Corynebacterium jeikeium Opportunistic infections in immunocompromised (e.g., patients with blood disorders, bone marrow transplants, intravenous catheters) Multiple antibiotic resistance common (MDR) Carriage on skin of up to 40% of hospitalized patients (e.g., marrow t-plants)  Diseases L. monocytogenes causes meningitis and sepsis in newborns and immunosuppressed adults. It also causes outbreaks of febrile gastroenteritis.  important Properties L. monocytogenes is a small gram-positive rod arranged in V, L- shaped formations similar to corynebacteria.  The organism exhibits an unusual tumbling movment that distinguishes it from the corynebacteria, which are nonmotile.  Colonies on a blood agar plate produce a narrow zone of beta-hemolysis that resembles the hemolysis of some streptococci.  L. monocytogenes infections occur primarily in two clinical settings:  (1) in the fetus : newborn as a result of transmission across the placenta or during delivery;  (2) in immunosupressed adults, especially renal transplant patients. The organism is distributed worldwide in animals, plants, and soil. From these reservoirs, it is transmitted to humans by contact with animal or their feces, by unpasteurized milk, and by contaminated vegetables. In the United States, listeriosis primarily a food-borne disease associated with eating unpasteurized cheese. The pathogenesis of Listeriois dependent upon the organism's ability to invade mononuclear phagocytic cells. Because Li.steria preferentially grows intracellularly, cell-mediated immunity is more important host defense than humoral immunity. Suppression of cell-mediated immunity predisposes to Listeria infections.  L. monocytogenes can move from cell to cell by means of "actin rockets," a filament of actin that contracts and propels the bacteria through the membrane of one human cell and into another.  Listeriolysin O is an important virulence factor. It is a cytolysin/hemolysin similar to streptolysin O that acts by degrading cell membranes. Laboratory diagnosis  Laboratory diagnosis is made primarily by Gram stain and culture  The appearance of gram-positive rods  on blood agar, it produces small, grey, translucent drop-like colonies surrounded by a small zone of indistinct beta hemolysis.  It may require incubation up to 48 hours to produce visible growth.  The isolation of lesteria is confirmed by the presence of motile organisms, which differentiate them from the nonmotile corynebacteria.