Mycobacterium (Exam 3) Flashcards
(54 cards)
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
2
Q
What are the two major groups of mycobacterium?
A
mycobacterium tuberculosis complex and nontuberculosis mycobacterium
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
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.
5
Q
What are the two staining methods for mycobacterium?
A
Ziehl-Neelson and Kinyoun methods
6
Q
What are the 4 runyoun groups for classification of nontuberculosis mycobacterium (NTM)?
A
- photochromogens
- scotochromogens
- nonphotochromogens
- rapid growers
7
Q
What are photochromogens?
A
require light to form pigment following incubation in the dark.
8
Q
What are scotochromogens?
A
produce pigment in dark or light conditions
9
Q
What are nonphotochromogens?
A
non-pigmented regardless of incubation in dark or light conditions
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).
11
Q
Which NTM species are photochromogens?
A
- M. kansasii
- M. marinum
- M. asiaticum
12
Q
Which NTM species are scotochromogens?
A
- M. scrofulaceum (cause of cervical adenitis in children).
- M. szulgai (at 25º C)
- M. gordonae
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.
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
15
Q
What is the MAC complex an important pathogen in?
A
Aids patients
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
17
Q
Which NTM species are rapid growers?
A
- M. fortuitum
- M. chelonae
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.
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
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
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
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
What are the ATS levels?
- 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
What BLS lab level does the CDC suggest mycobacterium be set at?
-BLS level 3
27
Why do acid fast stains work well with mycobacterium?
– high lipid cell content of Mycobacteria binds fuchsin dye & resists destaining by acid-alcohol.
28
What are the two primary stain methods for mycobacterium?
- 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
What should you use for optimal recovery of mycobacterium?
-at least one solid media and one liquid media
30
What liquid media are used for mycobacterium?
- BACTEC 12B medium
| - Middlebrook 7H9 Broth
31
What solid media are used for mycobacterium?
-one with an agar base and one with an egg base
32
What agar based media is used for mycobacterium?
-Middlebrook 7H10 non-selective.
33
What egg base media are used for mycobacterium???
-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
On average how long does M. Tuberculosis take to grow?
- 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
How should mycobacterium be incubated?
- If skin specimens incubate at 25-30 degrees C
| - 35 degrees C in the dark, 5-10% CO2 and high humidity.
36
How do scotocromogens look on media?
deep yellow – orange in both dark & light (some get darker on light exposure)
37
how do non pigment producing mycobacterium look on media?
– non pigmented in either dark or light (may produce pale yellow, buff, tan colonies not intensifying upon light exposure)
38
What are the important biochemical tests for mycobacterium?
- Niacin accumulation
- Nitrate reduction
- Catalase
- Tween 80 hydrolysis
- Arylsulfatase
- Urease
39
What is the basis behind the niacin accumulation test?
- 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
Which species of mycobacterium are niacin accumulation positive?
- M. tuberculosis
- M. Simiae
- on occassion, strains of M. bovis, M. africanum, and M. marinum lack the enzyme
41
What is the basis behind the nitrate reduction test?
-Only a few species of mycobacteria produce nitroreductase which catalyzes reaction
42
which mycobacterium species are nitrate positive?
- M. tuberculosis
| - M. kansasii
43
How is the tween 80 hydrolysis test performed?
-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
Which mycobacterium are tween 80 positive?
M. Kansasii
-tuberculosis complex tend to be negative.
45
Do M. tuberculosis complex bacteria produce heat stable catalase?
Most M. tuberculosis complex organisms do not produce heat stable catalase
46
What are newer identification methods being used for mycobacterium?
- Broth culture medium/automated continuous growth monitoring systems – BACTEC MGIT
- MALDI-TOF
- Septi-check AFB system
- Molecular
- genotype
47
What is the identification scheme for M. Tuberculosis?
- 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
Does everyone who has been exposed to tuberculosis become infected?
- 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
What population is TB prevalent in?
- Prevalent in elderly, chronic malnourished, alcoholics, homeless, and those living in crowded conditions.
- 1/3 of the worlds population is infected
50
What is primary TB and what are the outcomes?
-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
What is secondary TB?
Secondary (Reactivation) TB- person with latent TB that manifests as disease at later time (immune system becomes compromised for some reason)
52
What is miliary TB?
spread to other organs hematogenously.
53
What is the clinical presentation of pulmonary TB?
- Mimics others – asymptomatic to symptomatic
- Low grade fevers
- Anorexia and weight loss
- Fatigue & night sweats
- Productive cough
- Fever/chills
54
What is the treatment for TB?
- First line: 2 – 3 in combination for 9 months possibly 12.
- Isoniazid
- Rifampin
- Pyrazinamide
- Ethambutol
