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Clinical Microbiology (Exam 3) > Mycobacterium (Exam 3) > Flashcards

Mycobacterium (Exam 3) Flashcards

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1
Q

What are the species in the mycobacterium tuberculosis complex?

A
  • Mycobacterium tuberculosis
  • M. bovis
  • M. bovis BCG (bacillus Calmette- Guerin)-
  • M. africanum
  • M. caprae
  • M. canettii
  • M. microti
  • M. pinnipedii
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2
Q

What are the two major groups of mycobacterium?

A

mycobacterium tuberculosis complex and nontuberculosis mycobacterium

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3
Q

What are the general characteristics of mycobacterium?

A
  • Non-spore forming
  • Slender, slow-growing bacilli
  • Obligate aerobes, CO2 enhances some.
  • Cell wall contains numerous mycolic acids
  • Considered Gram positive/variable
  • Mycobacteria are acid fast
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4
Q

how long does mycobacterium take to grow?

A
  • Visible growth may take 2-60 days
  • A single cells generation time may take 20-36 hours
  • High lipid content of the cell walls, cells tend to clump which makes nutrient uptake more difficult thus slow growth rate.
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5
Q

What are the two staining methods for mycobacterium?

A

Ziehl-Neelson and Kinyoun methods

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6
Q

What are the 4 runyoun groups for classification of nontuberculosis mycobacterium (NTM)?

A
  1. photochromogens
  2. scotochromogens
  3. nonphotochromogens
  4. rapid growers
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7
Q

What are photochromogens?

A

require light to form pigment following incubation in the dark.

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8
Q

What are scotochromogens?

A

produce pigment in dark or light conditions

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9
Q

What are nonphotochromogens?

A

non-pigmented regardless of incubation in dark or light conditions

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10
Q

How are rapid growing NTM differentiated from others?

A

take fewer than 7 days to appear on solid media (groups I-III take longer than 7 days).

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11
Q

Which NTM species are photochromogens?

A
  • M. kansasii
  • M. marinum
  • M. asiaticum
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12
Q

Which NTM species are scotochromogens?

A
  • M. scrofulaceum (cause of cervical adenitis in children).
  • M. szulgai (at 25º C)
  • M. gordonae
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13
Q

which NTM species are nonphotochromogens?

A
  • M. avium/intracellulare complex
  • M. shimoidae
  • M. genavense
  • M. xenopi – found in hot water taps in hospitals, causes pulmonary infections and disseminated disease.
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14
Q

Which mycobacterium species are reported together as a complex and why?

A
  • M. avium complex (MAC),
  • M. avium and M. intracellulare are two separate organisms.
  • On clinical grounds and routine laboratory methods, they cannot be distinguished from each other so reported in combination
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15
Q

What is the MAC complex an important pathogen in?

A

Aids patients

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16
Q

What does the MAC complex present with in AIDS patients and how is this different than TB?

A
  • lymphadentitis
  • does not create granulomas or tissue death in pulmonary infections, differs from TB
  • increased mucus production can lead to bronchiectasis
  • not spread person to person, unlike TB
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17
Q

Which NTM species are rapid growers?

A
  • M. fortuitum

- M. chelonae

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18
Q

What media do the NTM rapids growers grow on?

A
  • MAC
  • They will grow on MacConkey agar (different formulation than that used for enterics, it has crystal violet omitted), most other Mycobacteria spp. do not grow on MacConkey.
  • MacConkey plate is inoculated with a 7 day broth culture of the organism and will see growth in 5 days for both of the species above, they are the only 2 that will grow in 5 days on MacConkey at 37 degrees C.
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19
Q

What is the noncultivatible NTM species and what does it cause?

A
  • M. leprae
  • It is an obligate intracellular bacteria that only survives several minutes in the environment.
  • cause leprosy
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20
Q

What are common mycobacterium tissue sources?

A
  • any tissue or organ
  • sputum is common, taken 3 consecutive days early morning
  • tissue/granuloma
  • urine
  • blood, csf, GI,stool,bone
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21
Q

Why is processing of sputum mycobacterium samples different than other sources?

A

-Processing of sputum specimens for acid fast bacteria requires digestion/ decontamination followed by neutralization and concentration (via centrifugation). These steps kill normal flora bacteria that outgrow TB and also releases the TB bacterium from cells and/or mucin

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22
Q

What are the methods of sputum processing?

A
  • 6% NaOH, 5% Oxalic acid (may be harmful to Mycobacteria – timing critical)
  • Trisodium phosphate (TSP) Zephiran
  • N-acetyl-L-cysteine (NALC) NaOH method – NaOH reduced to 2% when mixed with NALC
23
Q

What is the preferred method of sputum processing and why?

A
  • NALC
  • NALC is a mucolytic agent- liquifies mucus, releasing bacteria
  • NaOH is both mucolytic and a decontaminating agent.
24
Q

What are the steps of sputum processing?

A
  • (digestion/decontamintion) requires 15 minute incubation.
  • Following decontamination/digestion add the phosphate buffer followed by 15-20 minutes of centrifugation.
  • Smears (Acid fast stain) and set up cultures.
25
Q

What are the ATS levels?

A
  • ATS level I grow mycobacterium and stain but do not ID
  • ATS level II Identify TB only.
  • ATS level III Identify all mycobacteria spp. and perform susceptibility testing.
26
Q

What BLS lab level does the CDC suggest mycobacterium be set at?

A

-BLS level 3

27
Q

Why do acid fast stains work well with mycobacterium?

A

– high lipid cell content of Mycobacteria binds fuchsin dye & resists destaining by acid-alcohol.

28
Q

What are the two primary stain methods for mycobacterium?

A
  • Ziehl-Neelsen (hot)
  • Kinyoun (cold) method- uses a higher concentration of phenol in the primary stain to accelerate the staining process and thus does not require heat
29
Q

What should you use for optimal recovery of mycobacterium?

A

-at least one solid media and one liquid media

30
Q

What liquid media are used for mycobacterium?

A
  • BACTEC 12B medium

- Middlebrook 7H9 Broth

31
Q

What solid media are used for mycobacterium?

A

-one with an agar base and one with an egg base

32
Q

What agar based media is used for mycobacterium?

A

-Middlebrook 7H10 non-selective.

33
Q

What egg base media are used for mycobacterium???

A

-Löwenstein-Jensen- coagulated whole eggs, glycerol (enhances growth), potato flour, Malachite green.

-Petragnani – coagulated whole eggs, egg yolks, whole milk, potato, potato flour, Malachite green.
Malachite green dye inhibits other microbes (contaminants)

34
Q

On average how long does M. Tuberculosis take to grow?

A
  • Mycobacterium tuberculosis takes on average 2-4 weeks to grow and susceptibility testing takes and additional 2-4 weeks for a total of 4-8 weeks on average (Ghodbane et al. Nature 2014).
  • New technique by Ghodbane is reducing this time for cultivation to as little as 72 hours.
35
Q

How should mycobacterium be incubated?

A
  • If skin specimens incubate at 25-30 degrees C

- 35 degrees C in the dark, 5-10% CO2 and high humidity.

36
Q

How do scotocromogens look on media?

A

deep yellow – orange in both dark & light (some get darker on light exposure)

37
Q

how do non pigment producing mycobacterium look on media?

A

– non pigmented in either dark or light (may produce pale yellow, buff, tan colonies not intensifying upon light exposure)

38
Q

What are the important biochemical tests for mycobacterium?

A
  • Niacin accumulation
  • Nitrate reduction
  • Catalase
  • Tween 80 hydrolysis
  • Arylsulfatase
  • Urease
39
Q

What is the basis behind the niacin accumulation test?

A
  • all mycobacterium produce niacin
  • Some species lack enzyme necessary to further convert niacin to niacin ribonucleotide resulting in high accumulation of niacin in the culture medium.
40
Q

Which species of mycobacterium are niacin accumulation positive?

A
  • M. tuberculosis
  • M. Simiae
  • on occassion, strains of M. bovis, M. africanum, and M. marinum lack the enzyme
41
Q

What is the basis behind the nitrate reduction test?

A

-Only a few species of mycobacteria produce nitroreductase which catalyzes reaction

42
Q

which mycobacterium species are nitrate positive?

A
  • M. tuberculosis

- M. kansasii

43
Q

How is the tween 80 hydrolysis test performed?

A

-Identifies mycobacteria possessing a lipase that splits the compound (tween 80) into oleic acid and polyoxyethylated sorbitol

-2 drops of Tween 80 are added to 1mL of saline and inoculated with a loopful of organism.
I
-ncubate at 35 degrees C up to 12 days.

-The fluid will change from light orange to pink or red (indicates positive result)

44
Q

Which mycobacterium are tween 80 positive?

A

M. Kansasii

-tuberculosis complex tend to be negative.

45
Q

Do M. tuberculosis complex bacteria produce heat stable catalase?

A

Most M. tuberculosis complex organisms do not produce heat stable catalase

46
Q

What are newer identification methods being used for mycobacterium?

A
  • Broth culture medium/automated continuous growth monitoring systems – BACTEC MGIT
  • MALDI-TOF
  • Septi-check AFB system
  • Molecular
  • genotype
47
Q

What is the identification scheme for M. Tuberculosis?

A
  • Culture growth w/in 4 – 8 weeks
  • Rough, dry, buff, non-pigmented on Lowenstein-Jensen media.
  • Strong Acid-fast +, slightly curved rods, beaded appearance.
  • Positive niacin & nitrate reduction.
  • 68 degree Catalase negative.
48
Q

Does everyone who has been exposed to tuberculosis become infected?

A
  • Not every person who is exposed to the bacteria will become infected (CDC).
  • Contagious - Inhalation of a single bacilli has been shown to lead to infection (not necessarily disease though, minimal infective dose).
49
Q

What population is TB prevalent in?

A
  • Prevalent in elderly, chronic malnourished, alcoholics, homeless, and those living in crowded conditions.
  • 1/3 of the worlds population is infected
50
Q

What is primary TB and what are the outcomes?

A

-Primary TB- the initial infection in a previously unexposed individual, 2 potential outcomes:
-Latent TB infection
Person exposed to TB, became infected and have antibodies against bacterium thus positive skin test but NOT active disease, no symptoms, are not contagious.
-Active Infection/Disease

51
Q

What is secondary TB?

A

Secondary (Reactivation) TB- person with latent TB that manifests as disease at later time (immune system becomes compromised for some reason)

52
Q

What is miliary TB?

A

spread to other organs hematogenously.

53
Q

What is the clinical presentation of pulmonary TB?

A
  • Mimics others – asymptomatic to symptomatic
  • Low grade fevers
  • Anorexia and weight loss
  • Fatigue & night sweats
  • Productive cough
  • Fever/chills
54
Q

What is the treatment for TB?

A
  • First line: 2 – 3 in combination for 9 months possibly 12.
  • Isoniazid
  • Rifampin
  • Pyrazinamide
  • Ethambutol

Clinical Microbiology (Exam 3) (4 decks)

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