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Mycobacteriaceae P.O. Box 131375, Bryanston, 2074 Ground Floor, Block 5 Bryanston Gate, Main Road Bryanston, Johannesburg, South Africa www.thistle.co.za Tel: +27 (011) 463-3260 Fax: +27 (011) 463-3036 OR + 27 (0) 86-538-4484 e-mail : mail@thistle.co.za Please read this section first The HPCSA and the Med Tech Society have confirmed that this clinical case study, plus your routine review of your EQA reports from Thistle QA, should be documented as a “Journal Club” activity. This means that you must record those attending for CEU purposes. Thistle will not issue a certificate to cover these activities, nor send out “correct” answers to the CEU questions at the end of this case study. The Thistle QA CEU No is: MT00025. Each attendee should claim THREE CEU points for completing this Quality Control Journal Club exercise, and retain a copy of the relevant Thistle QA Participation Certificate as proof of registration on a Thistle QA EQA. MICROBIOLOGY LEGEND CYCLE 29 – ORGANISM 2 Mycobacteriaceae Mycobacterium is a genus of Actinobacteria, given its own family, the Mycobacteriaceae. The genus includes pathogens known to cause serious diseases in mammals, including tuberculosis (Mycobacterium tuberculosis) and leprosy (Mycobacterium leprae). The Latin prefix "myco—" means both fungus and wax; its use here reflects the "waxy" compounds that compose parts of the cell wall. Mycobacteria are aerobic and nonmotile bacteria (except for the species Mycobacterium marinum, which has been shown to be motile within macrophages) that are characteristically acid-alcohol fast. Mycobacteria do not contain endospores or capsules and are usually considered Gram-positive. However, this has been strongly argued by other scientists. While Mycobacteria do not seem to fit the Gram-positive category from an empirical standpoint (i.e. they generally do not retain the crystal violet stain well), they are classified as an acid-fast Gram-positive bacterium due to their lack of an outer cell membrane. All Mycobacterium species share a characteristic cell wall, thicker than in many other bacteria, which is hydrophobic, waxy, and rich in mycolic acids/mycolates. The cell wall consists of the hydrophobic mycolate layer and a peptidoglycan layer held together by a polysaccharide, arabinogalactan. The cell wall makes a substantial contribution to the hardiness of this genus. Mycobacterium tuberculosis bacteria using Mycobacterium tuberculosis acid-fast Ziehl-Neelsen stain Many Mycobacterium species adapt readily to growth on very simple substrates, using ammonia or amino acids as nitrogen sources and glycerol as a carbon source in the presence of mineral salts. Optimum growth temperatures vary widely according to the species and range from 25 °C to over 50 °C. Some species can be very difficult to culture sometimes taking over two years to develop in culture. Furthermore, some species also have extremely long reproductive cycles — Thistle QA is a SANAS accredited organisation, No: PTS0001 Accredited to ISO guide 43 and ILAC G13 Certificate available on request or at www.sanas.co.za P.O. Box 131375, Bryanston, 2074 Ground Floor, Block 5 Bryanston Gate, Main Road Bryanston, Johannesburg, South Africa www.thistle.co.za Tel: +27 (011) 463-3260 Fax: +27 (011) 463-3036 OR + 27 (0) 86-538-4484 e-mail : mail@thistle.co.za M. leprae, may take more than 20 days to proceed through one division cycle (for comparison, some E. coli strains take only 20 minutes), making laboratory culture a slow process. In addition, the availability of genetic manipulation techniques still lags far behind that of other bacterial species. A natural division occurs between slowly– and rapidly–growing species. Mycobacteria that form colonies clearly visible to the naked eye within seven days on subculture are termed rapid growers, while those requiring longer periods are termed slow growers. Mycobacteria cells are straight or slightly curved rods between 0.2-0.6 µm wide and 1.0-10 µm long. Staining characteristics Mycobacteria are classical acid-fast organisms. Stains used in evaluation of tissue specimens or microbiological specimens include Fite's stain, Ziehl-Neelsen stain, and Kinyoun stain. Mycobacteria appear phenotypically most closely related to members of Nocardia, Rhodococcus and Corynebacterium. Ecological characteristics Mycobacteria are widespread organisms, typically living in water (including tap water treated with chlorine) and food sources. Some, however, including the tuberculosis and the leprosy organisms, appear to be obligate parasites and are not found as free-living members of the genus. Pathogenicity Mycobacteria can colonize their hosts without the hosts showing any adverse signs. For example, billions of people around the world have asymptomatic infections of M. tuberculosis. Mycobacterial infections are notoriously difficult to treat. The organisms are hardy due to their cell wall, which is neither truly Gram negative nor positive. Additionally, they are naturally resistant to a number of antibiotics that disrupt cell-wall biosynthesis, such as penicillin. Due to their unique cell wall, they can survive long exposure to acids, alkalis, detergents, oxidative bursts, lysis by complement, and many antibiotics. Most Mycobacteria are susceptible to the antibiotics, clarithromycin and rifamycin, but antibiotic- resistant strains have emerged. As with other bacterial pathogens, surface and secreted proteins of M. tuberculosis contribute significantly to the virulence of this organism. There is an increasing list of extracytoplasmic proteins proven to have a function in the virulence of M. tuberculosis. Medical classification Mycobacteria can be classified into several major groups for purpose of diagnosis and treatment: M. tuberculosis complex, which can cause tuberculosis: M. tuberculosis, M. bovis, M. africanum, and M. microti; M. leprae, which causes Hansen's disease or leprosy; Nontuberculous Mycobacteria (NTM) are all the other Mycobacteria, which can cause pulmonary disease resembling tuberculosis, lymphadenitis, skin disease, or disseminated disease. Phenotypic testing Various phenotypic tests can be used to identify and distinguish different Mycobacteria species and strains. References 1. Ryan KJ, Ray CG (editors) (2004). Sherris Medical Microbiology (4th ed.). McGraw Hill. 2. http://en.wikipedia.org/wiki/Mycobacterium Questions 1. Discuss the morphological characteristics of Mycobacteria. 2. How would you identify Mycobacteria in your laboratory? 3. Discuss the pathogenicity of Mycobacteria. Thistle QA is a SANAS accredited organisation, No: PTS0001 Accredited to ISO guide 43 and ILAC G13 Certificate available on request or at www.sanas.co.za .
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