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Medical Microbiology II - 4336 > Unit 2: Mycobacteria, Acid-fast Bacteria > Flashcards

Unit 2: Mycobacteria, Acid-fast Bacteria Flashcards

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1
Q

How do bacilli strains differ in their morphology?

A

to grow as solitary rods or long strands serpentine cords

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2
Q

What causes bacteria to be acid-fast?

A

mycolic acid

lipid-rich cell wall resistant to digesting agents such
as strong acids and bases

can be strong or weak acid-fast

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3
Q

Acid -fast bacteria do not stain readily, but once stained with ____________ dyes, resist decolorization with tenacity.

A

arylmethane

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4
Q

Should it be reported if only one acid-fast bacteria is seen?

A

Yes! Report if you seen ANY.

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5
Q

What cell wall component makes the classification of bacteria?

A

peptidoglycan

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6
Q

What group of people are more likely to get active TB?

A

HIV/AIDS patients

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7
Q

What are the four clinical significant Mycobacteria?***

A

-Mycobacterium tuberculosis complex
-Mycobacterium ulcerans
-Mycobacterium leprae
-Nontuberculosis mycobacteria

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8
Q

What is part of the Mycobacterium tuberculosis complex?**

A

-M. tuberculosis
-M. bovis
-M. microti
-M. africanum

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9
Q

What are the nontuberculosis mycobacteria that fall under the Runyon classification?***

A

-Runyon I: M. Kansasii & M. marinum
-Runyon II: M. scrofulaceum
-Runyon III: M. avium complex
-Runyon IV: M. chelonei and M. fortuitum

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10
Q

Mycobacterium tuberculosis (MTB) is a pathogenic bacterial species in the genus Mycobacterium and the causative agent of most cases of tuberculosis (TB) First discovered in 1882 by ____________.

A

Robert Koch

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11
Q

TB is a pandemic disease, tuberculosis is especially common in the developing world owing to HIV infection (_______% of individuals with HIV disease may have tuberculosis).

A

15-20

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12
Q

a chronic, granulomatous, slowly progressive infection, usually of the lungs; eventually, many other organs and tissues may be affected.

A

Mycobacterium tuberculosis

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13
Q

Microbiologic characteristics of TB?

A

Aerobic and nonmotile.
No endospores or capsules
Gram-positive.
Cell wall, thicker, hydrophobic, waxy, and rich in mycolic acids/mycolates. Some species are fastidious.

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14
Q

TB adapts readily to growth on very simple substrates, using ___________ or amino acids as nitrogen sources and __________as a carbon source in the presence of mineral salts.

A

ammonia, glycerol

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15
Q

What is the optimum growth for TB?

A

25 to over 50 degrees Celsius

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16
Q

General symptoms of TB?

A

Fever, fatigue, night sweats, weight loss & malaise
Septicemia, multiple organ failure
Weight loss despite increased appetite – “consumption”

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17
Q

Symptoms of pulmonary tuberculosis?

A

pneumonia, chronic cough and coughing up blood

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18
Q

Symptoms of digestive tuberculosis?

A

abdominal pain and diarrhea, fever, weight loss. Bowel obstruction or bleeding may occur.

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19
Q

Symptoms of renal tuberculosis?

A

dysuria, hematuria and flank pain. WBCs in urine.

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20
Q

What are the virulence factors of TB?

A

-Waxy cell wall.
-Major factor is ability to invade and survive within macrophages* as surface protein called “exported repetitive protein” prevents phagosome from joining with lysosome.
-It produces no exotoxins or no LPS.

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21
Q

How are TB mycobacterial antigens classified as?

A

1- Soluble (cytoplasmic) and insoluble (cell wall lipid bound).
2- Carbohydrates or proteins
3- by their distribution within the genus

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22
Q

How is TB transmitted? and how does the disease progress?

A

by droplets from person with active case of tuberculosis.
The microorganism is very stable in sputum droplets and can remain viable in very even dry sputum for up 6 days.
M. tuberculosis in droplets is then inhaled and reach the highly aerobic environment of the lung where it produces non-specific pneumonitis
-inflammation —> “exudative lesion” and end up with granulomas

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23
Q

In TB, Granuloma becomes surrounded by fibrin which calcifies witch is called a ___________, it can be seen by chest X – ray. The infection may stop at this point and the individual may have no more symptoms.

A

tubercle

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24
Q

What are granulomas composed of?

A

collection of macrophages

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25
Q

What is a tubercle bacillus?

A

-macrophage bacteria complex.
-calcified
-enclosed —> no infection unless it bursts

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26
Q

How can TB become reactivated?
What is the mortality rate?

A

-The tubercle can disseminate throughout the body – becomes systemic infection
-Tubercle can be coughed up and swallowed, becoming systemic via the gastrointestinal tract
-Tubercle can burst years after primary infection

50%

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27
Q

What are the two ways TB can be diagnosed?

A
  1. AFB in sputum smear: indicative of tuberculosis.
  2. M.tuberculosis culture: egg yolk or oleic acid albumin containing media agar; 2-8 wks (Löwenstein–Jensen medium, Liquid BACTEC?)
  3. Chest X – ray
  4. Tuberculin skin test (Mantoux Test)
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28
Q

What is the special media for M. tuberculosis?

What agar does the bacillus not grown on?

A

TheLöwenstein–Jensen medium*

blood agar

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29
Q

medium used for TB that contains radioactive metabolites and growth can be detected by the presence of radioactive carbon dioxide production in about 2 weeks

A

Liquid BACTEC medium*

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30
Q

How does TB appear when grown on Löwenstein–Jensen medium?

A

brown, granular colonies (sometimes called “buff, rough and tough”)*

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31
Q

How long must TB be incubated onLöwenstein–Jensen medium?

A

incubated for a significant length of time, usually four weeks, due to the slow doubling time ofM. tuberculosis(15–20 hours) compared with other bacteria.

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32
Q

Purified protein derivative (PPD), part of cell wall is injected under the skin, 24 – 48 hours measure the size of the welt that forms. Cell have been primed and recognize the PPD.

A

Tuberculin skin test (Mantoux Test)

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33
Q

What does a positive Mantoux test mean?

A

the pt. has been exposed to the organism

  • Could have active infection
  • Could have been infected but were one of the 90% who are Asymptomatic.
  • You have been immunized
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34
Q

Mycobacterium leprae staining morphology is virtually indistinguishable from __________.

A

M. tuberculosis

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35
Q

Mycobacterium leprae has not been cultured on routine mycobacteria media, what media is used instead?

A

-mouse footpad
-Nine-banded armadillo

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36
Q

-Chronic disease of the skin, mucous membranes, and nerves
-Infectivity very low (requires close contact)

A

Leprosy (Hansen’s Disease)

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37
Q

What bacteria causes Leprosy (Hansen’s Disease)?

A

Mycobacterium leprae

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38
Q

Leprosy:

Most pathology is associated with organism’s tropism for…

A

peripheral nerves

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39
Q

Leprosy:

localized; confined to skin and nerves.

A

Tuberculoid

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40
Q

Leprosy:

disseminated. Associated with a impaired cell-mediated immunity

A

Lepromatous

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41
Q

How is Nontuberculous Mycobacteria transmitted?

A

not usually person to person. Frequently environmental
Isolated from culture: may not indicate disease

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42
Q

What are other names for nontuberculous mycobacteria?

A

MOTT (Mycobacteria Other Than Tubercle Bacilli), Atypical or Opportunistic

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43
Q

Runyon Classification:

Group I?

A

Photochromogens (pigment
on exposure to light)

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44
Q

Runyon Classification:

Group II?

A

Scotochromogens (pigment in dark)

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45
Q

Runyon Classification:

Group III:?

A

Non-photochromogens

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46
Q

Runyon Classification:

Group IV:?

A

Rapid Growers growth at 4-6 days

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47
Q

What are two examples in Runyon Group I?

A

-M. kansasii
-M. marinum

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48
Q

Where is M. kansasii (group I) found?

A

water

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49
Q

What temp does M. marinum (group I) grow better at?

A

at 30 degrees Celsius

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50
Q

Where is M. marinum (group I) found?

A

Found in salt water and fresh water—contamination from infected fish and other wildlife

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51
Q

What diseases does M. kansasii (group I) cause?

A

Causes chronic pulmonary disease and extrapulmonary diseases such as cervical lymphadenitis and cutaneous disease

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52
Q

What diseases does M. marinum (group I) cause?

A

Causes cutaneous disease (usually enters through open wounds or traumatic injury)

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53
Q

What are the Runyon Group II examples?

A

*rarely isolated

-M. gordonae (biosafety level I)
-M. scrofulaceum
-M. szulgai

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54
Q

Runyon Group II:

Water contaminant. Rarely causes disease

A

M. gordonae (biosafety level I)

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55
Q

Runyon Group II:

Raw milk, soil, water, and other dairy products. Cervical adenitis in children, pulmonary disease, skin infections and bacteremia

A

M. scrofulaceum

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56
Q

Runyon Group II:

Water and soil. Pulmonary disease, cervical adenitis, and bursitis

A

M. szulgai

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57
Q

Nonphotochromogens
-Non-pigmented
->7 days to appear on solid media
-Frequently isolated in laboratory

A

Runyon Group III

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58
Q

Examples of Runyon group III

A

-M. avium complex
-M. xenopi
-M. ulcerans
-M. haemophilum

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59
Q

-Most common NTM in the USA
-Ubiquitous: water, soil, animals
-Includes many species and subspecies

A

M. avium Complex

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60
Q

M. avium and ___________ are different organisms but closely resemble each other.

A

M. intracellulare

-Distinction cannot be made by routine laboratory testing or on clinical grounds

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61
Q

M. avium Complex is frequently called _______.

A

MAI

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62
Q

What diseases does M. avium complex cause?

A

-Pulmonary infections in patients with preexisting pulmonary disease
-Cervical lymphadenitis
-Disseminated disease in HIV & AIDS

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63
Q

Runyon Group III –> M. avium complex:

-Buruli ulcer, a neglected tropical disease reported in >30 countries.
-Causes scarring and can progress to bone.
-Grows at lower temperatures.

A

M. ulcerans

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64
Q

Runyon Group III –> M. avium complex:

-Associated with water and can survive high temperature.
-May cause slowly progressing pulmonary infection

A

M. xenopi

65
Q

Runyon Group III –> M. avium complex:

-Skin infections but can disseminate.
-Requires hemin.
-Sub chocolate plate

A

M. haemophilum

66
Q

Runyon IV characteristics?

A

-Grow in <7 days.
-Grow on routine media
-Stain weakly on Gram-stain. -Ubiquitous
-Cause disease in immunocompromised persons

67
Q

Examples of Runyon IV (account for 90% of clinical diseased caused by RGM)

A

-M. fortuitum
-M. chelonae
-M. abscessus

68
Q

Causes post-operative infections of the chest, skin and soft tissue. Localized, pulmonary, and CNS diseases

A

M. fortuitum (Runyon IV)

69
Q

Causes skin and soft tissue, postoperative wound infections, and keratitis

A

M. chelonae (Runyon IV)

70
Q

causes skin and soft tissue, pulmonary, postoperative infections

A

M. abscessus (Runyon IV)

71
Q

__________ ranks high among laboratory-acquired infections.

A

Tuberculosis

72
Q

Tuberculosis has a low infective dose, ____% chance of infection with fewer than 10 AFB.

A

50

73
Q

What safety measures are made for TB specimen processing?

A

(Level 2), minimum:
BSCI or II, gloves, limited access, slides heat-fixed within BSC.

74
Q

What safety measures are made for positive TB cultures?

A

(Level 3): Lab separated by interlock anteroom and has negative pressure, Type II BSC with UV/HEPA, materials for disposal decontaminated before removal (usually autoclaved)
PPE: gloves, gown, respirator

75
Q

What type of mycobacteria specimens are acceptable and why?

A

almost any specimen is acceptable because mycobacteria present with a wide rang of infections.

swabs should not be accepted

76
Q

Gastric aspirate mycobacteria samples should be processed within ____ hours of collection or neutralized with ____________.

A

4, 10% sodium carbonate

77
Q

All mycobacteria samples except blood should be stored how?

A

refrigerated within 1 hour of collection if not immediately processed

78
Q

Some specimens can be directly inoculated to media; others must be processed. Processing means?

A

mucus digestion and decontamination.

79
Q

What are three ways specimens are processed?

A

-NaOH
-(N-acetyl-L-cysteine) NALC
-Zephiran-trisodium phosphate & Oxalic acid

80
Q

________ acid is sometimes used for specimens contaminated with Pseudomonas or Proteus.

A

Oxalic

81
Q

digestant (liquefies the sample)

A

(N-acetyl-L-cysteine) NALC
-NALC-NaOH (most
common)

82
Q

Used for decontamination (mucolytic as well)

A

NaOH

-too much NaOH can also kill Mycobacteria

83
Q

What is the disadvantage of the strong alkaline reagent usually used for the decontamination step?

A

it is also toxic to mycobacteria

84
Q

Steps of specimen processing?

A

-Add NaOH/NALC-digestion and decontamination
-Vortex, let sit for 15 min
-Add buffer to stop the reaction; centrifuge at high speed
-Pour off the supernatant and make smears and inoculate solid and liquid media.
-monitor

85
Q

What are the steps for quality control for Decontamination Process?

A

-Prepare a simulated sputum sample with 102, 103, and 104 CFU/mL.
-Add to sterile sputum.
-Process as usual and inoculate media.
-Inoculate a set of media without processing.

86
Q

unprocessed samples would also show __________ growth

A

reduced

87
Q

What does the MGIT™ Mycobacteria Growth Indicator Tubes contain?

A

4 ml of Middlebrook 7H9 Broth base with a fluorescent indicator.

88
Q

Modified Middlebrook 7H9 broth Growth & Antibiotic supplement _________ sponges provide unique growth matrix

A

cellulose

89
Q

Modified Middlebrook 7H9 broth:

broth is contained in a plastic tube with screw-top and is supplemented with _____________ enrichment and _____________ antibiotic mixture.

A

BD MGIT™ OADC

BD MGIT™ PANTA™

90
Q

What does the Löwenstein Jensen agar contain?

A

Contains eggs and
malachite green (inhibits most other bacteria)

91
Q

How should tubed solid media be incubated?

A

Slanted position with screw caps loose for 5–7 days, after 7 days caps should be tightened and tubes may be positioned upright. 6-8wks. Liquid media: 6wks

92
Q

How should plated solid media be incubated?

A

CO2-permeable plastic bags, medium-side down (5–10% CO2)

93
Q

What is the optimum temperature for MTBC?

A

35–37°C. Some NTM grow best at 25–33°C. M. xenopi requires 40–42°C.

94
Q

How should culture be incubated for skin, bone, and joint biopsies?

A

inoculate two sets of media including chocolate, one at 35-37°C and the other at the lower optimal temperature (usually 30°C)
-M. marinum, M. ulcerans, M. haemophilium

95
Q

What are two preliminary identificators on culture?

A
  1. rate of growth on solid media
  2. colonial pigmentation
96
Q

What is the appearance of M. tuberculosis colonies?

A

crumbly, and buff colored

97
Q

What are the microscopy detection methods for AFB?

A

A. Light microscopy stains
Ziehl-Nielson
Kinyon
B. Fluorescent stain
Fluorochrome
(Auramine-Rhodamine)

98
Q

Microscopically, Mycobacterium avium-intracellulare infection is marked by numerous acid fast organisms growing within ____________.

A

macrophages

99
Q

What are the identification tests based off of?

A

-Growth rate
-Pigment
-Colony morphology (do acid fast stain to make sure it is an AFB).
-Complex media requirements

100
Q

The three examples of identification tests?

A

-The Niacin Test: Mycobacterium tuberculosis produces large amounts of niacin.
-The Catalase Test: M. tuberculosis produces a heat sensitive catalase.
-The Nitrate Test: M. tuberculosis can reduce nitrate.

101
Q

The niacin test is detected by the addition of what to a suspension of the culture?

A

10% cyanogen bromide and 4% aniline in 96% ethanol

102
Q

What color is a positive niacin test?

A

canary yellow

103
Q

An example that would be positive for the niacin test? negative?

A

M. tuberculosis

M. bovis

104
Q

Is the catalase test quantitative?

A

semiquantitative

105
Q

Most mycobacteria have a lipase that can split Tween 80. What are the exceptions?

A

Mtb, M. avium, M. bovis

106
Q

What color is a positive Tween 80 test? negative?

A

positive- pink
negative-orange

107
Q

What distinguish M. tuberculosis from M. bovis?

A

TCH (Thiopene-2-carboxylic acid hydrazide)

-M. bovis is susceptible

108
Q

_______ reduces tellurite in 3 to 4 days, most other nonchromogenic species do not.

A

MAC

109
Q

Do most nonchromogenic species reduce tellurite?

A

No

110
Q

All rapid growers reduce tellurite in ____ days.

A

3

111
Q

What does a positive and negative Tellurite reduction test look like?

A

positive- cloudy
negative- clear

112
Q

What organism is strongly positive for the Arylsulfatase test?
What is negative?

A

M. fortuitum-chelonei

M-avium

113
Q

What does a positive arylsulfatase test look like?

A

red/pink

114
Q

What type of bacteria does the arylsulfatase test detect?

A

fast growers

115
Q

What is the principle of the Arylsulfatase test?

A

Tripotasium phenolphthaleindisulfide/sulfate producing free phenolphthalein by arylsulfatase enzyme

116
Q

What can have a positive or negative result for Iron uptake?

A

Fortuitum/chelonae

117
Q

-Identifies mycobacteria by analysis of mycolic acids
-test is based on separation of mycolic acids
-It is the primary means of ID in health dept/CDC for non-TB mycobacteria

A

HPLC

118
Q

-Range of speciation and sensitivity greater than DNA probe (without amplification) and biochemical testing.
-Can be performed directly on smear-positive specimen or on cultures.

A

HPLC

119
Q

identification method:

-Limited number of species (MTB complex and MAV complex)
-Uses single-strand DNA probe
-Probe is labeled – often with a chemiluminescent label.
-DNA:RNA hybrid
-A selection reagent removes unbound probe
-Read in luminometer

A

DNA probe

120
Q

Most laboratories that identify mycobacteria use automated instruments to detect growth and for susceptibility testing.
Two examples?

A

BacTec and the mycobacteria growth indicator tube (MGIT).

121
Q

Automated detection systems:

detects CO2 production produced by viable bacteria.

A

BacTec

122
Q

Automated detection systems:

a fluorescence-based system relying on oxygen consumption rather than carbon dioxide production.

A

MGIT (mycobacteria growth indicator tube)

123
Q

Bactec NAP test:

if growth is detected in the control bottle but not in the NAP bottle, it can be identified as…

A

M. tuberculosis

124
Q

A rapid method of identification, that inhibits Mtb complex

A

Bactec NAP test

125
Q

Identification method:

Requires extraction (special sample processing)
-Extract proteins and
inactivate sample

A

MALDI-TOF

ex: Bruker

126
Q

What are the advantages of the MALDI-TOF method?

A

-Rapid identification
-Good identification to species level for rapid growers and most other groups
-MALDI-TOF instrumentation is becoming more commonly available

127
Q

What are the disadvantages of the MALDI-TOF method?

A

-Time-consuming extraction protocol
-Cannot differentiation species in MTB complex
-Must have culture growth on solid media
-Many groups currently working on methods for detection directly from liquid media

128
Q

If results are inconclusive by TB/MAC PCR and MALDI-TOF what should done next?

A

DNA sequencing

129
Q

Both Bruker BioTyper and bioMerieux VITEK MS
take ______ hours to process prior to loading on the MALDI-TOF instrument

A

2-2.5

130
Q

Bruker BioTyper OR bioMerieux VITEK MS?

-30 minute heat lysis inactivation
-Ethanol wash
-Silica bead disruption with acetonitrile/formic acid extraction

A

Bruker BioTyper

131
Q

Bruker BioTyper OR bioMerieux VITEK MS?

-Kit based
-Ethanol/bead inactivation/disruption
-No heat lysis
-Formic acid/acetonitrile extraction

A

bioMerieux VITEK MS

132
Q

-Multiplex real-time PCR
-Detection directly from patient sample: Sputum
-2 hour TAT
-Sample-to-answer
-Moderate complexity

A

GeneXpert – MTB/RIF

133
Q

GeneXpert – MTB/RIF:

Simultaneous detection of both MTB and _________ resistance mutations, which are markers for MDR-TB strain.

A

rifampin

134
Q

A single Xpert MTB/RIF Assay result detected approximately ___% of patients who were acid-fast bacilli (AFB) smear–positive and culture-confirmed as infected with Mycobacterium tuberculosis complex (MTBC)

A

97

135
Q

Two serial Xpert MTB/RIF Assay results detected ____ % of AFB smear–positive/MTBC culture-positive patients.

A

100

136
Q

One or two Xpert MTB/RIF Assay tests detected ____% and ____%, respectively, of sputum specimens that were AFB smear–negative but culture-positive for MTBC.

A

55, 69

137
Q

What is the treatment for acid-fast bacteria?

A

Long-term therapy (6–9 months) with antituberculous drugs (isoniazid, rifampicin, pyrazinamide, ethambutol).

-Combination therapy should always be given*

138
Q

What is a growing and a persistent problem with treatment?

A

Drug Resistance

139
Q

What is a prevention method for TB?

A

bacille Calmette–Guérin (BCG) vaccination containing live attenuated organisms, in childhood.

140
Q

What does MDR-TB stand for?

A

Multi-drug Resistance tuberculosis

141
Q

A global program that manages regimentation of drug delivery as a cornerstone of managing TB disease

A

directly observed therapy (DOT) ?

142
Q

Some cases of human contracting ______ are from vaccine, or from drinking unpasteurized milk from infected cow.

A

M. bovis

143
Q

What is the vaccine for TB?

A

BCG (Bacilli Calmete Guerin) vacine

144
Q

The BCG vaccine uses live attenuated ________, causes TB in cows.

A

M. bovis

145
Q

How does the BCG vaccine work?

A

-Made to stimulate the cell-mediated immune response as M. tuberculosis is largely an intracellular pathogen.
-Most effective in children, given as part of early childhood regime in endemic regions.

146
Q

What is used for prevention of TB?

A

Chemoprophylaxis

-Isoniazid given to people who were in contact

147
Q

What is used for treatment of latent TB?

A

Isoniazid and/or Rifampin (used together, 12 dose min)

148
Q

What is the first “new” TB antibiotic in 40 years?

A

Diarylquinolines

-Inhibit ATP production
-Novel mechanism
-Shorter course of therapy

149
Q

What does DOTS stand for?

A

direct observation treatment system

150
Q

What is the proposed reference method for MTB susceptibility testing with pyrazinamide?

A

radiometric BACTEC TB 460 (BD Biosciences, Sparks, MD)

151
Q

What is the reference MTB susceptibility testing method for all drugs except pyrazinamide?

A

Agar proportion

152
Q

What is the TAT for MTB susceptibility testing?

A

3 weeks

153
Q

Resistance definition…

A

therapy is less likely to be clinically successful when >1% of the bacterial population being tested in vitro is resistant.

154
Q

What are the two FDA-cleared systems for MTB susceptibility testing?

A

-Radiometric BACTEC TB 460
-ESP Culture System II (Trek Diagnostics, Westlake, OH)

155
Q

How is susceptibility testing done for resistant MTB strains?

A

Molecular testing

156
Q

What is the most frequently used method for susceptibility testing of MTBC in the U.S.?

A

Radiometric BACTEC TB 460

157
Q

Susceptibility testing:

What are the primary drugs that are recommended by the NCCLS subcommittee for testing against isolates of MTBC?

A

isoniazid (at 2 concentrations)

and rifampin, ethambutol, and pyrazinamide (1 concentration of each).

158
Q

What is the TAT for susceptibility testing of MTBC?

A

7-10 days

Medical Microbiology II - 4336 (20 decks)

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