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 gram  M. tuberculosis is the cause of most cases
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
susceptibility to pyrazinamide. This o Larynx
organism is associated with approximately o Pleural lining (pleuritis)
31% of the cases of human tuberculosis.  Disseminated tuberculosis can be
Reservoir hosts for the organism include diagnosed by a positive tuberculin skin
goats, cattle, sheep, pigs, wild boars, deer, test
and fox.  Patients may also have latent tuberculosis
M. microti, typically found in rodents, guinea  A patient with latent tuberculosis is not
pigs, rabbits, cats, llamas, and meerkats, infectious and does not have active disease,
usually fails to grow in culture. although the organism is present in
It has been identified in tuberculosis in both granulomas. Patients with latent
immunocompetent and immunosuppressed tuberculosis may progress to active disease
patients. M. canettii has been primarily ( reactivation tuberculosis) at any time.
identified in cases of lymphadenitis and  Reactivation tuberculosis typically occurs
generalized tuberculosis in after an incident in which cellular immunity
immunocompromised individuals. is suppressed or damaged as a result of a
M. pinnipedii is transmitted from sea lions to change in lifestyle or other health condition.
humans and has been associated with Individuals infected with HIV are particularly
granulomatous lesions in the lymph nodes, susceptible to developing active
lungs, pleura, and spleen. It is also tuberculosis.
pathogenic in guinea pigs, rabbits, Diagnosing tuberculosis is more difficult in
camels, and possibly cattle. people infected with HIV because are
The two additional species in the M. anergic (lack a biologic response) to
tuberculosis complex reservoir hosts include tuberculin skin testing, a primary means
the banded mongoose (M. mungi) or large of identifying individuals infected with M.
mammals such as gazelles, antelopes, tuberculosis.
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 I SLOW-GROWING
individuals 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
The introduction of highly active
antiretroviral therapy (HAART) has
EPIDEMIOLOGY AND PATHOGENESIS
reduced the infections caused by M. avium
complex in patients with AIDS.  MAC is an important pathogen in both
immunocompromised and
GENERAL CHARACTERISTICS. immunocompetent populations.
 MAC is particularly noteworthy for its
 MAC comprises M. avium, Mycobacterium potentially pathogenic role in pulmonary
intracellulare, M. avium subsp. avium, M. infections in patients with AIDS and in those
avium subsp. paratuberculosis, M. avium who are not infected with HIV. T
subsp. silvaticum (wood pigeon bacillus), M.  organisms are ubiquitous in the
avium subsp. hominissuis, M. arosiense, M. environment and have been isolated from
vulneris, M. marseillense, M. natural water, soil, dairy products, pigs,
bouchedurhonense, M. chimaera, M. chickens, cats, and dogs.
colombiense, M. yogonense, and M.  Infections caused by MAC are acquired by
timonense. inhalation or ingestion.
 NTM have extraordinary starvation survival.  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 potentially pathogenic species based on

NONTUBERCULOUS MYCOBACTERIA pigmentation and molecular studies.
 The common human pathogens in this
 The large group of organisms that constitute
group include Mycobacterium abscessus
the RGM is divided into six major groups of
 subsp. abscessus, Mycobacterium  Like many other NTM, these organisms are
chelonae, and Mycobacterium fortuitum. ubiquitous in the environment and are
On Gram staining, these organisms appear present worldwide.
as weakly gram-positive rods resembling  They have been found in soil, marshes,
diphtheroids. rivers, and municipal water supplies (tap
water) and in marine and terrestrial life
forms.
EPIDEMIOLOGY AND PATHOGENESIS  may be commensals on the skin.
 They can also be health care–associated
 The rapidly growing mycobacteria
infections
considered potentially pathogenic can
cause disease in either healthy or SPECTRUM OF DISEASE
immunocompromised patients.
 The most common infection associated with
RGM is posttraumatic wound infection.
 C. NONCULTIVATABLE  Hansen disease/ Leprosy is a chronic
NONTUBERCULOUS MYCOBACTERIA— disease of the skin, mucous membranes,
MYCOBACTERIUM LEPRAE and nerve tissue. Leprosy remains a
worldwide public health concern owing to
 The nontuberculous mycobacterium M. the development of drug-resistant isolates.
leprae is a close relative of M. tuberculosis.
 M. leprae has not yet been cultivated
This organism causes leprosy (also called
in vitro, although it can be cultivated in the
Hansen disease)
armadillo and in the footpads of mice.
 Molecular biologic techniques have  Two major forms of the disease are a
provided most of the information about this localized form, called 1. tuberculoid
organism’s genomic structure and its leprosy, and a more disseminated form,
various genes and their products. called 2. lepromatous leprosy.
 Routine diagnosis of leprosy is based on Patients with lepromatous leprosy are
distinct clinical manifestations, such as anergic to M. leprae because of a defect in
hypopigmented skin lesions and peripheral their cell-mediated immunity. Because the
nerve involvement, in conjunction with a organisms’ growth is unimpeded, these
skin smear that tests positive for acid-fast individuals develop extensive skin lesions
bacilli (AFB). containing numerous AFB; the organisms
can spill over into the blood and
EPIDEMIOLOGY AND PATHOGENESIS disseminate.
Understanding of the epidemiology and In contrast, individuals with tuberculoid
pathogenesis of leprosy is hampered by the leprosy do not have an immune defect; the
inability to grow the organism in culture. disease is localized to the skin and nerves.

EPIDEMIOLOGY LABORATORY DIAGNOSIS OF

MYCOBACTERIAL INFECTION
 The primary reservoir for M. leprae is
infected humans. Purpose of AFB Test
 The disease is transmitted person to person  TB diagnosis
through inhalation or contact with infected  Other mycobacterial infection detection
skin.
 Monitoring treatment response
PATHOGENESIS  Identifying the cause of symptoms

 leprosy is both a bacterial and an SPECIMEN COLLECTION AND TRANSPORT

immunologic disease.
1. Pulmonary spx
Two primary phases are a silent phase,
2. Gastric Lavage spx
during which the leprosy bacilli multiply in
3. Urine Spx
the skin in macrophages, and an
4. Fecal spx
intermediate phase, in which the bacilli
5. Tissue and Body Fluid
multiply in peripheral nerves and begin to
6. Blood spx
cause sensory impairment.
7. Wounds, Skins lesions, and Aspirates
SPECIMEN PROCESSING
CONTAMINATED SPECIMENS

 Most specimens submitted for

mycobacterial culture consist of organic
debris, such as mucin, tissue, serum, and
other proteinaceous material contaminated
with organisms.
 Rapidly growing mycobacteris are
especially susceptible to high or prolonged
exposure to 2% or more sodium hydroxide
(NaOH)
SPECTRUM OF DISEASE  Digestion-decontamination procedures
should be as gentle as possible.
 spectrum of disease caused by M. leprae
ranges from subclinical infection to INADEQUATE SPECIMEN AND REJECTION
intermediate stages of disease to full-blown CRITERIA
and serious clinical manifestations involving
the skin, upper respiratory system, testes,  Specimens should be rejected according to
and peripheral nerves. T the following guidelines: (1) insufficient
 volume. (2) contamination with saliva, (3)  NaOH method
dried swabs, (4) pooled sputum or urine, (5)  Zephiran-trisodium phosphate method
container has been compromised or is  N-acetyl-L-cysteine (NALC)-2% NaOH
broken or leaking, and (6) length of time method
from collection to processing is too long  Oxalic acid

SPECIAL CONSIDERATIONS  For fecal specimens, approximately 0.2 g of

stool is emulsified in 11 mL of sterile,
 Aerosol-induced sputum should be treated
filtered, distilled water.
as sputum
 Swabs and wound aspirates should be
 Gastric lavages should be processed within
transferred to a sterile, 50-ml. conical
4 hours of collection or neutralized with 10%
centrifuge tube containing a liquid medium
sodium carbonate and refrigerated until
at a ratio of 1 part specimen to 5 to 10 parts
processed as for sputum.
liquid medium.
 Urine specimens should be divided into a
 Large pieces of tissue should be finely
maximum of four 50-mL centrifuge tubes
minced with a sterile scalpel and scissors.
and centrifuged at 36003 g for 30 minutes.
 1. SOLID MEDIA

SPECIMENS NOT REQUIRING DECONTAMINATION

 Tissues or body fluids collected aseptically usually

do not require the digestion and decontamination
methods used with contaminated specimens.
 + CSF should be handled aseptically and
centrifuged for 30 minutes at 36003 g to
concentrate the bacteria.
 Pleural fluid should be collected in sterile
anticoagulant (1 mg/mL
ethylenediaminetetraacetic acid [EDTA] or 0.1
mg/mL, heparin).
 Joint fluid and other sterile exudates can be
handled aseptically and inoculated directly to
media.
 Bone marrow aspirates may be injected into
Pediatric Isolator tubes (Alere, Waltham, MA),
which help prevent clotting

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
2. LIQUID MEDIA

CULTIVATION
APPROACH TO IDENTIFICATION