MYCOBACTERIA

Mycobacterium are aerobic, non–spore-

forming (except for Mycobacterium
marinum and M. avium subsp.
paratuberculosis), nonmotile, very thin,
slightly curved or straight rods
Mycobacterium is the only genus in the
Mycobacteriaceae
Genera that are closely related to
Mycobacterium include Corynebacterium,
Nocardia, Rhodococcus, Segniliparus,
Tsukamurella, and Gordonia
Mycobacterium spp. have an unusual cell I. MYCOBACTERIUM TUBERCULOSIS
wall; it contains N-glycolylmuramic acid COMPLEX (MTC)
instead of N-acetylmuramic acid and has a TUBERCULOSIS / COMSUMPTION
very high lipid content, which creates a EPIDEMIOLOGY AND PATHOGENESIS
hydrophobic permeability barrier. EPIDEMIOLOGY
Mycobacteria are difficult to stain with Gram
staining, but are generally considered  M. tuberculosis is the cause of most cases
gram positive. of human tuberculosis, particularly in
acid fastness; this characteristic developed countries.
distinguishes mycobacteria from other  M. tuberculosis complex organisms are not
genera. able to replicate in the environment and are
Rapid-growing mycobacteria (RGM), in therefore isolated to growth in tissues of
which growth is apparent sooner than 7 humans and other warm-blooded animals.
days after subculture to LowensteinJensen Tuberculosis continues to be a public health
medium, may partially or completely lose problem in the United States. An additional
this characteristic as a result of their growth complicating factor in the management of
characteristics. tuberculosis is the increasing incidence of
Another important feature of many species coinfection with HIV. HIV-associated
is that they grow more slowly than most tuberculosis
other human pathogenic bacteria because In the United States, tuberculosis is typically
of their hydrophobic cell surface. found among the poor, the homeless,
Slow-growing mycobacteria, by definition, intravenous drug users, alcoholics, the
require more than 7 days to produce elderly—in general, medically underserved
colonies on solid media. populations.
The genus Mycobacterium includes several Another area of concern is the increased
highly pathogenic species included in the M. identification of multidrug-resistant
tuberculosis complex—M. uclerans and the tuberculosis (MDR-TB) and extensively
nonculturable M. leprae. There are also drug-resistant tuberculosis (XDR-TB).
more than 180 species of nonculturable
nontuberculosis mycobacteria (NTM), many Pathogenesis
of which are increasingly found in Inhalation of a single viable organism has
opportunistic pathogens that are found been shown to lead to infection, although
throughout the environment. close contact is usually necessary.
For the most part, mycobacteria can be Of those who become infected with M.
divided into two major groups based on tuberculosis, 15% to 20% develop disease.
fundamental differences in epidemiology The disease usually occurs some years
and association with disease: those after the initial infection, when the patient’s
belonging to the M. tuberculosis complex immune system breaks down for some
and the others to the NTM group. reason other than the presence of
tuberculosis bacilli in the lung. In a small
percentage of infected hosts, the disease
 becomes systemic, affecting a variety of complex may range from asymptomatic to
organs. acutely symptomatic.
INGESTION OF UNPASTEURIZED MILK  Cases of pulmonary disease caused by M.
cows, M. bovis inhalation of infectious tuberculosis complex organisms are
droplets from infected cattle. clinically, radiologically, and pathologically
An attenuated strain of M. bovis, bacillus indistinguishable
Calmette-Guérin (BCG), has been used  Primary tuberculosis typically is
extensively in many parts of the world to considered a disease of the respiratory
immunize susceptible individuals against tract. Common presenting symptoms
tuberculosis. include low-grade fever, night sweats,
M. africanum demonstrates physiologic and fatigue, anorexia (loss of appetite), and
biochemical properties that place it weight loss.
intermediately between M. tuberculosis and  These organs include the following:
M. bovis. This organism has been primarily o Genitourinary tract
identified as the cause of approximately half o Lymph nodes (cervical lymphadenitis)
of the cases of tuberculosis in West Africa, o Central nervous system (meningitis)
but it has also been found in the United o Bone and joint (arthritis and osteomyelitis)
States in patients who previously resided in o Peritoneum
Africa. o Pericardium
M. caprae can be identified by its o Larynx
susceptibility to pyrazinamide. This o Pleural lining (pleuritis)
organism is associated with approximately  Disseminated tuberculosis can be
31% of the cases of human tuberculosis. diagnosed by a positive tuberculin skin
Reservoir hosts for the organism include test
goats, cattle, sheep, pigs, wild boars, deer,  Patients may also have latent tuberculosis
and fox.  A patient with latent tuberculosis is not
M. microti, typically found in rodents, guinea infectious and does not have active
pigs, rabbits, cats, llamas, and meerkats, disease, although the organism is present in
usually fails to grow in culture. granulomas. Patients with latent
It has been identified in tuberculosis in both tuberculosis may progress to active disease
immunocompetent and immunosuppressed ( reactivation tuberculosis) at any time.
patients. M. canettii has been primarily  Reactivation tuberculosis typically occurs
identified in cases of lymphadenitis and after an incident in which cellular immunity
generalized tuberculosis in is suppressed or damaged as a result of a
immunocompromised individuals. change in lifestyle or other health condition.
M. pinnipedii is transmitted from sea lions to Individuals infected with HIV are particularly
humans and has been associated with susceptible to developing active
granulomatous lesions in the lymph nodes, tuberculosis.
lungs, pleura, and spleen. It is also Diagnosing tuberculosis is more difficult in
pathogenic in guinea pigs, rabbits, people infected with HIV because are
camels, and possibly cattle. anergic (lack a biologic response) to
The two additional species in the M. tuberculin skin testing, a primary means
tuberculosis complex reservoir hosts include of identifying individuals infected with M.
the banded mongoose (M. mungi) or large tuberculosis.
mammals such as gazelles, antelopes,
waterbucks, and oryxes (M. orygis).
SPECTRUM OF DISEASE

 Tuberculosis may mimic other diseases,

such as pneumonia, neoplasm, or fungal
infections.
 In addition, clinical manifestations in
patients infected with M. tuberculosis
 The tuberculin skin test, or purified
protein derivative (PPD) test, is based on
the premise that after infection with M.
tuberculosis, an individual develops a
delayed hypersensitivity cell-mediated
immunity to certain antigenic components of
the organism.
After 48 to 72 hours, an infected individual
will show a delayed hypersensitivity reaction
to the PPD, characterized by erythema
(redness) and, most important, induration
(firmness because of the influx of immune
cells
The PPD test is not 100% sensitive or
specific, and a positive reaction to the skin
test does not necessarily signify the
presence of disease.

The T-Spot TB test

enzyme-linked immunosorbent assay
(ELISA) called QuantiFERON-TB Gold Plus
2. NONTUBERCULOUS MYCOBACTERIA
a. Slow-Growing
b. Rapidly Growing NTM and;
c. Non-cultivatable NTM
The NTM include all mycobacterial species
that do not belong to the M. tuberculosis
complex. Currently approximately 180
species
 The members of this large group of
mycobacteria are often opportunistic
pathogens.
 Significant geographic variability is seen
both in the prevalence of and the species
responsible for NTM disease.
 present everywhere in the environment and
sometimes colonize the skin and respiratory RUNYON GROUPS I TO IV
and gastrointestinal tracts of healthy
RUNYON GROUPS SLOW-GROWING
individuals
I TO III NTM
 Interpretation of a positive NTM culture is RUNYON GROUP IV rapid growers NTM
complicated.
  Mycobacterium spp. synthesize
carotenoids (a group of yellow to red
pigments) in varying amounts and thus can
be categorized into three groups—
photochromogens, scotochromogens,
and nonphotochromogens—based on the
production of those pigments
The SLOW-GROWING NTM CAN BE
SUBDIVIDED INTO THREE GROUPS based
on the production of those pigments

A. PHOTOCHROMOGENS
The photochromogens (Table 42.3) are slow-
growing NTM that produce colonies requiring
light to form pigment.
B. SCOTOCHROMOGENS
The scotochromogens (Table 42.4) are slow-
growing NTM that produce pigmented
colonies whether grown in the dark or the
light
B. NONPHOTOCHROMOGENS
The nonphotochromogens (Table 42.5) are
slow-growing NTM that produce unpigmented
colonies whether grown in the dark or the
A. SLOW-GROWING NONTUBERCULOUS
light
MYCOBACTERIA
MYCOBACTERIUM AVIUM COMPLEX EPIDEMIOLOGY AND PATHOGENESIS
The introduction of highly active  MAC is an important pathogen in both
antiretroviral therapy (HAART) has reduced immunocompromised and immunocompetent
the infections caused by M. avium complex in populations.
patients with AIDS.  MAC is particularly noteworthy for its
GENERAL CHARACTERISTICS. potentially pathogenic role in pulmonary
infections in patients with AIDS and in those
 MAC comprises M. avium, Mycobacterium who are not infected with HIV. T
intracellulare, M. avium subsp. avium, M.  organisms are ubiquitous in the environment
avium subsp. paratuberculosis, M. avium and have been isolated from natural water,
subsp. silvaticum (wood pigeon bacillus), M. soil, dairy products, pigs, chickens, cats, and
avium subsp. hominissuis, M. arosiense, M. dogs.
vulneris, M. marseillense, M.  Infections caused by MAC are acquired by
bouchedurhonense, M. chimaera, M. inhalation or ingestion.
colombiense, M. yogonense, and M.  NTM have extraordinary starvation survival.
timonense.  MAC cultures can have an opaque glossy-
white colony morphology or can produce a
smaller translucent colony morphology
 CLINICAL SPECTRUM OF DISEASE.

B. RAPIDLY GROWING  The common human pathogens in this

NONTUBERCULOUS MYCOBACTERIA group include Mycobacterium abscessus
subsp. abscessus, Mycobacterium
 The large group of organisms that constitute
chelonae, and Mycobacterium fortuitum.
the RGM is divided into six major groups of
On Gram staining, these organisms appear
potentially pathogenic species based on
as weakly gram-positive rods resembling
pigmentation and molecular studies.
diphtheroids.
 EPIDEMIOLOGY AND PATHOGENESIS water) and in marine and terrestrial life
forms.
 The rapidly growing mycobacteria
 may be commensals on the skin.
considered potentially pathogenic can
 They can also be health care–associated
cause disease in either healthy or
infections
immunocompromised patients.
 Like many other NTM, these organisms are SPECTRUM OF DISEASE
ubiquitous in the environment and are
present worldwide.  The most common infection associated with
RGM is posttraumatic wound infection.
 They have been found in soil, marshes,
rivers, and municipal water supplies (tap
 C. NONCULTIVATABLE
NONTUBERCULOUS MYCOBACTERIA—
MYCOBACTERIUM LEPRAE

 The nontuberculous mycobacterium M.

leprae is a close relative of M. tuberculosis.
This organism causes leprosy (also called
Hansen disease)
 Hansen disease/ Leprosy is a chronic
disease of the skin, mucous membranes,
and nerve tissue. Leprosy remains a
worldwide public health concern owing to
the development of drug-resistant isolates.
 M. leprae has not yet been cultivated SPECTRUM OF DISEASE
in vitro, although it can be cultivated in the
 spectrum of disease caused by M. leprae
armadillo and in the footpads of mice.
ranges from subclinical infection to
 Molecular biologic techniques have
intermediate stages of disease to full-blown
provided most of the information about this
and serious clinical manifestations involving
organism’s genomic structure and its
the skin, upper respiratory system, testes,
various genes and their products.
and peripheral nerves. T
 Routine diagnosis of leprosy is based on
 Two major forms of the disease are a
distinct clinical manifestations, such as
localized form, called 1. tuberculoid
hypopigmented skin lesions and peripheral
leprosy, and a more disseminated form,
nerve involvement, in conjunction with a
called 2. lepromatous leprosy.
skin smear that tests positive for acid-fast
Patients with lepromatous leprosy are
bacilli (AFB).
anergic to M. leprae because of a defect in
EPIDEMIOLOGY AND PATHOGENESIS their cell-mediated immunity. Because the
organisms’ growth is unimpeded, these
Understanding of the epidemiology and individuals develop extensive skin lesions
pathogenesis of leprosy is hampered by the containing numerous AFB; the organisms
inability to grow the organism in culture. can spill over into the blood and
EPIDEMIOLOGY disseminate.
In contrast, individuals with tuberculoid
 The primary reservoir for M. leprae is leprosy do not have an immune defect; the
infected humans. disease is localized to the skin and nerves.
 The disease is transmitted person to person
through inhalation or contact with infected LABORATORY DIAGNOSIS OF
skin. MYCOBACTERIAL INFECTION

PATHOGENESIS Purpose of AFB Test

 leprosy is both a bacterial and an  TB diagnosis

immunologic disease.  Other mycobacterial infection detection
Two primary phases are a silent phase,  Monitoring treatment response
during which the leprosy bacilli multiply in  Identifying the cause of symptoms
the skin in macrophages, and an
SPECIMEN COLLECTION AND TRANSPORT
intermediate phase, in which the bacilli
multiply in peripheral nerves and begin to 1. Pulmonary spx
cause sensory impairment. 2. Gastric Lavage spx
3. Urine Spx
4. Fecal spx
5. Tissue and Body Fluid
6. Blood spx
7. Wounds, Skins lesions, and Aspirates
 SPECIMEN PROCESSING INADEQUATE SPECIMEN AND REJECTION
CRITERIA
CONTAMINATED SPECIMENS
 Specimens should be rejected according to
 Most specimens submitted for
the following guidelines: (1) insufficient
mycobacterial culture consist of organic
volume. (2) contamination with saliva, (3)
debris, such as mucin, tissue, serum, and
dried swabs, (4) pooled sputum or urine, (5)
other proteinaceous material contaminated
container has been compromised or is
with organisms.
broken or leaking, and (6) length of time
 Rapidly growing mycobacteris are from collection to processing is too long
especially susceptible to high or prolonged
 NaOH method
exposure to 2% or more sodium hydroxide
 Zephiran-trisodium phosphate method
(NaOH)
 N-acetyl-L-cysteine (NALC)-2% NaOH
 Digestion-decontamination procedures
method
should be as gentle as possible.
 Oxalic acid
 SPECIAL CONSIDERATIONS SPECIMENS NOT REQUIRING
DECONTAMINATION
 Aerosol-induced sputum should be treated
as sputum  Tissues or body fluids collected aseptically
 Gastric lavages should be processed within usually do not require the digestion and
4 hours of collection or neutralized with 10% decontamination methods used with
sodium carbonate and refrigerated until contaminated specimens.
processed as for sputum.  + CSF should be handled aseptically and
 Urine specimens should be divided into a centrifuged for 30 minutes at 36003 g to
maximum of four 50-mL centrifuge tubes concentrate the bacteria.
and centrifuged at 36003 g for 30 minutes.  Pleural fluid should be collected in sterile
 For fecal specimens, approximately 0.2 g of anticoagulant (1 mg/mL
stool is emulsified in 11 mL of sterile, ethylenediaminetetraacetic acid [EDTA] or
filtered, distilled water. 0.1 mg/mL, heparin).
 Swabs and wound aspirates should be  Joint fluid and other sterile exudates can be
transferred to a sterile, 50-ml. conical handled aseptically and inoculated directly
centrifuge tube containing a liquid medium to media.
at a ratio of 1 part specimen to 5 to 10 parts  Bone marrow aspirates may be injected into
liquid medium. Pediatric Isolator tubes (Alere, Waltham,
 Large pieces of tissue should be finely MA), which help prevent clotting
minced with a sterile scalpel and scissors.
DIRECT DETECTION
1. Microscopy
2. Acid Fast Stain
3. Fluorochrome and Fuschin and Acid Fast
4. Antigen-Protein Detection
5. Immunodiagnostic Testing
6. Genetic Sequence and Nuclear Acid
Amplification
CULTIVATION
1. SOLID MEDIA
 2. LIQUID MEDIA
LIQUID MEDIA

 use of a liquid media system reduces the

turnaround time for isolation of acid-fast
bacilli to approximately 10 days, compared
with 17 days or longer for conventional solid
media.
 Growth of mycobacteria in liquid media,
regardless of the type, requires 5% to 10%
CO2.
 Gram staining can also be performed if
contamination is suspected.

APPROACH TO IDENTIFICATION

CONVENTIONAL PHENOTYPIC TEST

BIOCHEMICAL TESTING
 BIOCHEMICAL TESTS  ability to reduce tellurite in 3 to 4 days
distinguishes members of MAC from
1. Niacin
most other nonchromogenic species
 plays an important role in the oxidation-
6. Arylsulfatase
reduction reactions that occur during
 Enzyme arylsulfatase is present in most
mycobacterial metabolism.
mycobacteria.
 A positive niacin test is preliminary
 Test conditions can be varied to
evidence that an organism that exhibits a
distinguish the different forms of the
buff colored, slow-growing, rough colony
enzyme.
may be M. tuberculosis.
7. Growth Inhibitory by Thiophene-2-
 test is not sufficient to confirm
Carboxylic Acid Hydrazide
identification
 Test is used to distinguish M. bovis from
M. tuberculosis.

2. Nitrate Reduction
 is valuable for identifying M.
tuberculosis, M. kansasil, M. szolgai,
and M. fortuitum.
 The ability of acid-fast bacilli to reduce
nitrate is influenced by the age of the
colonies, temperature, pH, and enzyme
inhibitors.
 Commercially available nitrate strips
yield acceptable results with strongly
nitrate-positive organisms
ANTIMICROBIAL SUSCEPTIBILITY TESTING

 In vitro drug susceptibility testing should

be performed on the first isolate of M.
tuberculosis from all patients.
 Susceptibility testing of M. tuberculosis
requires meticulous care in the
3. Catalase preparation of the medium, selection of
 Most species of mycobacteria, except adequate samples of colonies,
for certain strains of M. tuberculosis standardization of the inoculum, use of
complex (some Isoniazid-resistant appropriate controls, and interpretation
strains) and M. gastri, produce the of results.
intracellular enzyme catalase, which  Direct Versus Indirect Susceptibility
splits hydrogen peroxide into water and Testing
oxygen. CONVENTIONAL METHODS
4. Tween 80 Hydrolysis
 is useful for differentiating species of  The development of primary drug
photochromogens, nonchromogens, and resistance in tuberculosis represents an
scotochromogens. increase in the proportion of resistant
5. Tellurite Reduction organisms.
 Some species of mycobacteria reduce  A poor clinical outcome is predicted with
potassium tellurite at variable rates. an agent when more than 1% of bacilli in
the test population are resistant.
THERAPY: ISONIAZID
PREVENTION: BCG VACCINE
NONTUBERCULOUS MYCOBACTERIA

 In general, the treatment of patients

infected with NTM requires more
Individualized therapy than does the
treatment of patients with tuberculosis.
 Currently, sufficient data exist to allow
general recommendations for
susceptibility testing of MAC, M.
kansasii, and M. marinum, Pulmonary
infections with MAC are often treated
with clarithromycin, rifampin, and
ethambutol (or streptomycin or amikacin
for severe disease).
 If the infection is disseminated,
clarithromycin, ethambutol, and rifabutin
may be prescribed.
 Pulmonary infections with M. kansasii
are treated with isoniazid, rifampin, and
ethambutol
 M. marinum skin and soft tissue
infections may be treated with either
clarithromycin and ethambutol,
clarithromycin and rifampin, or rifampin
and ethambutol