Mycobacteriology Flashcards
What bacterial killing agent is used for processing AFB samples?
Sodium hydroxide (NaOH)
What mucolytic agent is used for processing AFB samples?
N-acetyl-L-cysteine (NALC)
What is the classic smear and culture presentation for TB?
Smear positive
Culture negative
What is the sensitivity of AFB smears?
5,000 - 10,000 AFB/mL sputum
In what media does cording occur?
Liquid (MIGT)
Important things to remember about cording?
Not all TB cord, and not all cording is TB
What specimens get direct SecA1 sequencing?
All first time smear positive patients
What stain for AFB is used for direct patient samples?
Auramine rhodamine (AR)
What stain for AFB is used from media (solid or liquid)?
Kinyon
Which orgs may be Kinyon negative? Why? What do we do for confirmation?
Rapid growers
Have less mycolic acid in cell wall (less developed)
Modified acid-fast stain
How do we differentiate TB from m. bovis?
PCR
What are initial steps in identification of Mycobacteria?
Growth rate (fast 7 days) Colony morphology (not rough, slow growing - rule out TB) Color (non-pigmented, photo, scoto)
Mycobacteria - Rapid Growers
M. abscessus
M. chelonae
M. fortuitum
M. mucogenicum
M. smegmatis
Mycobacteria - Slow Growers
M. TB complex (tuberculosis, bovis, BCG, etc.) M. avium M. intracellulare M. haemophilum M. genavense M. kansasii M. marinum M. xenpoi M. gordonae M. scrofulaceum
Mycobacteria - Photochromogens
M. kansasii
M. marinum
M. asiaticum
M. simiae
Mycobacteria - Scotochromogens
M. scrofulaceum
M. gordonae
M. szuldai
M. flavescens
M. fortuitum - Colony Morphology
Rapid grower
Smooth (or rough), shiny
Irregular edge
M. fortuitum - Disease
Localized traumatic wound infections
Catheter infections
Surgical/Cosmetic surgery wound infections (breast augmentation)
Rarely a respiratory pathogen
M. abscessus group - Species
M. abscessus
M. massiliense
M. bolletii
M. abscessus group - Colony Morphology
Rapid grower
??
M. abscessus group - Disease
Chronic lung infections (CF, CGD patients)
Localized traumatic would infections
Surgical infections
Disseminated skin infections (pts on corticosteroids, organ transplants)
Catheter infections
Eye infections
M. chelonae - Colony Morphology
Rapid grower
??
M. chelonae - Disease
Localized traumatic wound infections
Post-traumatic or post-surgical corneal infections
Catheter infections
Disseminated skin infections (pts on corticosteroids, organ transplants)
Sinusitis
M. mucogenicum - Colony Morphology
Off white, mucoid, shiny, smooth?
M. mucogenicum - Disease
Normal Host - sputum contaminant
Immunocompromised Host -
Catheter infections (well-documented)
Lung infections? (unclear)
M. TB Complex - Species
M. tb M. bovis M. bovis BCG M. caprae M. africanum M. microti M. canetii M. pinnipedii
M. TB Complex orgs that are Niacin/Nitrate positive? And how are they distinguished?
MTB and M. canetii
MTB - rough, M. canetii - smooth
M. tb standard susceptibilities - drugs tested
SIRE + PZA Streptomycin (STR) Isoniazid (INH) Rifampin (RIF) Ethambutol (EMB) Pyrazinamide (PZA)
Which member of the TB complex is inherently resistant to Pyrazinamide (PZA)?
M. bovis
Definition of MDR TB
Resistant at least to both Isoniazid (INH) and Rifampin (RIF)
Definition of XDR TB
Additional resistance to any fluoroquinolone (the “floxacins”) and resistant to at least one of the three injectable drugs for TB (capreomycin, amikacin, kanamycin).
M. avium Complex - Species
M. avium
M. intracellulare
M. chimaera
“X cluster” - MAC probe pos, avium and intra. probe neg), includes M. colombiense, M. mantenii
M. avium Complex - Colony Morphology
Non-pigmented
Smooth, flat, transparent colonies
Rough variants can sometimes form
M. avium Complex - Disease
Frequently isolated slow growing NTM Chronic lung infections (CF, etc) Disseminated infections (usually AIDS pts) Lymphadenitis Cutaneous infections
M. xenopi - Colony Morphology
Classic “birds nest” colony morphology
Enhanced growth at 42 deg. C
Non-pigmented
M. xenopi - Disease and Treatment
Pulmonary infection
Optimal treatment for M. xenopi infection is not well established - in vitro sensis do not correlate with clinical response
Where is M. xenopi commonly found?
Mat be found in hospital water taps, hot water storage tanks, and contaminated bronchoscopes
When do we perform sensis for MAC?
Only when treatment failure is suspected
M. kansasii - Colony Morphology
Photochromogen
Flat, spready, dull colonies
Initially off-white then becoming yellow with exposure to light
M. kansasii - Disease
Chronic respiratory infection
Disseminated disease in AIDS
Isolated from liver/spleen (hairy cell leukemia)
What Mycobacteria cross-react with a PPD?
M. kansasii
What Mycobacteria cross-react with a Quanterferon Gold (QFN)?
M. kansasii
M. marinum
M. kansasii - AST and observed resistance
Initially test for Rifampin - then others
Resistance observed to Ethanmbutol, Ciprofloxacin, Doxycyclin, and Capreomycin
M. marinum - Colony Morphology
Photochromogen
Grows at 30 deg. C
Irregular edged colonies, off-white turing bright yellow
M. marinum - Disease
“Swimming pool” or “fish tank granuloma”
Is found in fresh and salt water, diagnosis often delayed due to uncommon org and failure to recall aquatic exposure
M. marinum - Susceptibility profile
Generally S to: Rifampin Rifabutin Etahmbutol Clarithromycin Trim/Sulfa
Generally R to:
Isoniazid (INH)
Pyrazinamide (PZA)
M. ulcerans - Colony Morphology and Culture
Extremely slow growing (6-12 wk inc. at 32 deg. C)
Colonies are yellowish, rough, with well-demarcated edges
Culture fails in over half of cases, is confirmed by molecular methods
M. ulcerans - Disease
Ulcerative skin disease known as Buruli ulcer
most frequently seen in children in rural tropical environments near wetlands
M. gordonae - Colony Morphology
Scotochromogen
Orange, smooth, shiny colonies
M. gordonae - Disease
Not significant, ubiquitous in the environment
Which AFB do not grow in culture?
M. leprae
M. tilburgii (rare species)
M. genavense (grows in liquid media only)
Which AFB require special growth supplements?
M. haempohilum (hemin)
M. avium ssp paratuberculosis, M. genavensae (Mycobactin J - siderophore)
What are the cutoffs for positive PPD tests set by the CDC?
5mm - high risk (HIV+, exposure toe active TB pt)
10mm - increased probability (recent immigrants, IVDU, healthcare workers)
15 mm - low risk (everyone else)
What do DNA probes target?
The ribosomal RNA sequences (becuase have ~10,000 copies, provides natural amplification)
What Mycobacteria species have available probes?
M. TB complex (TB, bovis, africanum, etc)
M. avium complex
M. kansasii
M. gordonae
How is DNA fingerprinting for epidemiology done in TB?
Restriction fragment length polymorphism
DNA is fragment using a restriction endonuclease, run on a gel, and then probed for IS6110 (has many copies inserted throughout genome)
Mechanism of INH resistance?
Deletion/mutation in katG gene, isolates become catalase negative or have decreased catalase activity
Resistance also associated with changes in inhA gene encoding for an enzyme in mycolic acid synthesis
Mechanism of Streptomycin resistance?
Mutations in rpsL gene (ribosomal S12 protein) and rrs (16s RNA)
Mechanism of RIF resistance?
Mutations in rpoB gene (B subunit of RNA polymerase)
Mechanism of fluoroquinolone resistance?
Mutations in gyrA (DNA gyrase)
What is preferred initial therapy for MAC?
Clarithromycin or Azithromycin PLUS ethambutol
What is the specimen of choice for TB diagnosis in kids?
Gastric aspirate, should be 5-10ml, same 3 consecutive day rule applies, neutralization recommended (1% sodium bicarb) for specimens >4 hrs from processing
Acceptable contamination rate for AFB processing?
3-5%
What can be added to specimens from CF patients for AFB work up?
5% oxalic acid added to conc sediments
What proteins are detected by IGRAs?
ESAT-6, CFP-10, and TB7.7
Which organisms can give false positive IGRA results?
M. marinum, M. kansasii, M. szulgai, M. flavescens
What are the types of leprosy and how do they differ clinically/diagnostically?
Lepromatous - many AFB in lesions (no CMI), biopsy nodules and plaques
Tuberculoid - very few AFB, biopsy rims of lesions
What causes buruli ulcer? How is it cultured?
M. ulcerans, biopsy or swabs cultured for 6 weeks at 30 degrees
Which NTM can exhibit cording?
M. gordonae, M. chelonae, M. marinum
If isolate is Nicain positive - think?
M. tuberculosis
If isolate is Nicain positive and a photochromagen - think?
M. simiae
If isolate is R to pyrazinamide and a slow grower - think?
M. bovis of M. bovis BCG
If isolate is associated with pt with bladder cancer and intravesicular immunotherapy - think?
M. bovis BCG
What is critical concentration resistance?
Growth of >1% of the inoculum in a the presence of the concentration of the drug that is the lowest conc that is 95% of “wild” strains
Which RGM have an inducible erm gene?
M. fortuitum
M. abbscessus (subsp. absecssus and bolletti)
How do you test for inducible (erm) resistance in RGM?
Read Clarithromycin result at 3 days, then reincubate and read again at 14 days
