MAJOR ARTICLE

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Posttreatment Reactivation of Tuberculosis in Mice
Caused by Mycobacterium tuberculosis Disrupted
in mce1R
Chan-Ick Cheigh,1 Ryan Senaratne,1 Yujiro Uchida,1 Nicola Casali,1 Lon V. Kendall,2 and Lee W. Riley1
1
Division of Infectious Diseases, School of Public Health, University of California, Berkeley; 2Laboratory Animal Resources, Colorado State
University, Fort Collins

Background. The reactivation of tuberculosis arises in persons who are latently infected and in those who
have been previously treated. The mechanism of the reactivation of tuberculosis in either situation is not well
understood. A 13-gene mce1 operon of Mycobacterium tuberculosis was previously shown to be associated with
latent infection in mice and may also play a role in reactivation.
Methods. We tested mce1 operon M. tuberculosis mutants in a Cornell mouse model to examine disease
progression and reactivation.
Results. In BALB/c mice, the wild-type, mce1 operon mutant, and mce1R (negative transcriptional regulator
of the mce1 operon) mutant M. tuberculosis strains were equally susceptible to orally administered isoniazid and
pyrazinamide. However, after cessation of the treatment, the mce1R mutant rapidly and progressively proliferated
in mouse lungs and spleens, whereas the other strains remained latent. The reactivation of the mce1R mutant was
associated with disease progression in the mouse lungs.
Conclusion. This observation demonstrates that the constitutive expression of the mce1 genes by M. tuberculosis
in the latent state can cause a reactivation of tuberculosis. The constitutive expression of the mce1 genes in the
mce1R mutant may allow this mutant to maintain its lipid metabolism, enabling it to survive long-term and
proliferate inside granulomas.

A large proportion of the ∼8 million new cases of tu-
berculosis each year arise in individuals who have latent
tuberculosis [1]. The lifetime risk of reactivation of
tuberculosis in those who have no underlying medical
conditions is 2%–23%, but the risk increases to 5%–
15% annually in persons coinfected with human im-
munodeficiency virus (HIV) [2, 3]. In addition, many
of those who develop a reactivation of tuberculosis have
a history of previous treatment of tuberculosis, which
is often incomplete [4]. Efforts to prevent reactivation
of tuberculosis are hampered by our limited under-
Received 19 October 2009; accepted 7 January 2010; electronically published
14 July 2010.
Potential conflicts of interest: none reported.
Financial support: Korea Research Foundation (grant KRF-2005-214-C00221);
National Institute of Health (grant R21AI063350).
Reprints or correspondence: Dr Lee W. Riley, School of Public Health, Univer-
sity of California, Berkeley, 201 Hildebrand Hall, Berkeley, CA 94720 (lwriley@
berkeley.edu).
The Journal of Infectious Diseases 2010; 202(5):752–759
 2010 by the Infectious Diseases Society of America. All rights reserved.
0022-1899/2010/20205-0014$15.00
DOI: 10.1086/655224
standing of the mechanism of latent tuberculosis and
the progression to active disease.
One major obstacle to studying latent tuberculosis
and reactivation of tuberculosis has been the lack of an
ideal animal model. Two murine models of latent My-
cobacterium tuberculosis infection are recognized. Al-
though neither of these models can be said to truly
represent latent or reactivation of tuberculosis in hu-
mans, they have provided useful information [5–8]. In
the first model, sometimes referred to as the low-dose
model, mice are infected aerogenically with 5–10 colony
forming units (CFUs) of M. tuberculosis. The pulmo-
nary bacterial burden stabilizes at a value of 3–4 log10
CFUs, which is maintained for 15–18 months [5, 8].
After this point, the infection reactivates and the mice
die. Flynn et al [6] and others have used this model to
show that reactive nitrogen intermediates as well as
tumor necrosis factor a play a role in preventing re-
activation of disease [7, 9]. There are variations of this
model that use different aerogenic inoculum doses to
accelerate the disease outcome [10].
The other widely used mouse model to study latent

752 • JID 2010:202 (1 September) • Cheigh et al

M. tuberculosis infection is the Cornell model [11–14]. Scanga
et al [8] reported on the utility of several versions of the Cornell
model and concluded that it is highly dependent on the pa-
rameters used to establish latency and that each version has
certain limitations. The Cornell model has the advantage of
achieving very low or undetectable numbers of bacilli and
maintaining those low levels for many weeks; however, it has
the disadvantage of artificially inducing latency with antibiotics
[8]. This model may be more akin to those human patients
who remain latently infected after completing treatment of ac-
tive disease.
The genome of M. tuberculosis contains 4 related copies of
a cluster of genes called mce operons (mce1–mce4) [15]. A
phylogenomic analysis of the operons has shown that they en-
code possible adenosine triphosphate (ATP)–binding cassette
(ABC) transporters [16]. We previously reported that the dis-
ruption of mce1 renders the M. tuberculosis mutant hypervi-
rulent in BALB/c mice and unable to stimulate a T helper 1–
type immune response [17]. The mce1 operon genes are neg-
atively regulated by Mce1R in intracellular M. tuberculosis [18].
An M. tuberculosis mutant disrupted in mce1R constitutively
expresses the mce1 genes in vivo and causes accelerated im-
munopathological response in the infected animal [19].
On the basis of the above-mentioned observations, we used
the Cornell murine model of latent infection to examine the
outcome of latent infection with the mce1 operon and mce1R
mutants. Surprisingly, each of these mutants caused an unex-
pected course of infection in this model and provided new
insights into the mechanism of reactivation of disease that had
not been previously considered.
MATERIALS AND METHODS
Animals. Female BALB/c mice (Charles River, Rockland,
MA) aged 8–10 weeks were maintained and studied in a Bio-
safety Level 3 animal facility at the University of California,
Berkeley.
Bacterial strains and growth conditions. The M. tuber-
culosis H37Rv wild type (American Type Culture Collection
strain 25618), M. tuberculosis H37Rv mce1 operon mutant, M.
tuberculosis H37Rv mce1R mutant, complemented M. tuber-
culosis H37Rv mce1R mutant, and Erdman wild type were
grown in Middlebrook 7H9 broth supplemented with 10% al-
bumin-dextrose-catalase (Becton Dickinson), 0.2% glycerol,
and 0.05% Tween 80 or on Middlebrook 7H11 agar contain-
ing oleic acid–albumin-dextrose-catalase supplement (Becton
Dickinson), 0.5% glycerol, and the antifungal agent cyclohex-
imide (100 mg/mL; Sigma-Aldrich). The 2 mutant strains (mce1
operon mutant and mce1R mutant) were generated from the
M. tuberculosis H37Rv background strain by means of ho-
mologous recombination with a 2-step counterselection strat-
egy, as described in detail elsewhere [17, 18, 20].
M. tuberculosis Disrupted in mce1R • JID 2010:202 (1 September) • 753
Infection and antibiotic treatment in mice. In the initial
set of experiments, mice were infected via the tail vein with
∼5 ⫻ 10 5 CFUs of mce1 operon and mce1R mutant bacilli in
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accordance with the traditional Cornell model. At 4 weeks after
infection, mice were given isoniazid (100 mg/mL) and pyrazin-
amide (15 mg/mL) in drinking water ad libitum for 4 weeks.
After a period of 4 weeks, some of the mice were treated with
0.08 mg of dexamethasone per day intraperitoneally for 6 d a
week for 20 weeks. In the second set of experiments, mice were
infected via the tail vein with a lower dose (5 ⫻ 10 2 CFUs per
animal). They were otherwise treated similarly to those of the
first set of experiments. In both experiments, mice were fol-
lowed up for clinical symptoms, organ CFU counts, and his-
tological analysis after cessation of the antibiotics.
When we observed an unexpected response with the mce1R
mutant in the initial set of experiments, we studied this mu-
tant further with the aerosol infection route. Mice were in-
fected with 100–200 CFUs of the mce1R mutant, comple-
mented mce1R mutant, or wild-type H37Rv strain per mouse
with an aerosol generation device (Inhalation Exposure Sys-
tem; Glas-Col). The inoculum doses in each group were as-
sessed as described below by harvesting the right lung of 4
mice (from each group of mice) 24 h after infection. At 4 weeks
after infection, the mice initiated treatment with isoniazid (100
mg/mL) and pyrazinamide (15 mg/mL) delivered ad libitum in
drinking water. The duration of treatment was 4 and 8 weeks,
and the mice were followed up without administration of
cortisone or dexamethasone for clinical symptoms, organ
CFU counts, and histological analysis after cessation of the
antibiotics.
Microbiological and histopathological examination. At
designated time points, 4 or 5 animals from each group were
killed and the lungs and spleen were harvested and examined
grossly, histologically, and microbiologically. The number of
tubercle bacilli in the corresponding organ homogenates in
phosphate-buffered saline with Tween 80 (0.05%) was assessed
by plating part of the homogenate on Middlebrook 7H11 agar
(Difco) plates. The CFU counts were enumerated 3–4 weeks
later. Data are presented as the mean value ( standard de-
viation) of 4 or 5 mice per group, and the experiment was
performed twice with similar results. Differences between the
mean of the CFU counts obtained from each treatment group
were analyzed by the Student t test and were considered sig-
nificant at P ! .05.
Tissue samples for histopathological studies were prepared
as described elsewhere [21, 22]. Briefly, the left lung was fixed
in 10% buffered formalin and sectioned. Slides for histopath-
ological analysis were made commercially (Histology Consul-
tation Service), and lung sections were stained either with he-
matoxylin and eosin or by the Ziehl-Neelsen method for
acid-fast bacilli. Lung sections from 4 mice from each group
per time point were analyzed by a veterinary pathologist spe-
cializing in mouse pathology, who was blinded to the sources
of the specimens. The lung sections were evaluated according

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to the following set of criteria: (1) percentage of lung displaying
parenchymal lesion, (2) severity of the organization of the le-
sions (in order of severity: diffuse, coalescing nodular, nodular,
or alveolar septal thickening), (3) type of macrophages present
(foamy or epithelioid), (4) lymphocyte distribution (mild,
moderate, marked, or severe), (5) areas of necrosis (focal, mul-
tiple focal, diffuse, or coalescing), (6) involvement of airways
(small to large numbers of inflammatory cells), and (7) number
of neutrophils present.

RESULTS

High-dose tail vein infection in mice treated for 4 weeks. At

a dose of 5 ⫻ 10 5 CFUs, all mice infected with the mce1R mutant
rapidly progressed to death, and hence this dose could not be
used with this mutant in the traditional Cornell model. Detailed
results of the BALB/c infection with the mce1R mutant are
published elsewhere [20]. However, with the wild-type and
mce1 operon mutant M. tuberculosis, bacilli were recovered
from organs during the course of dexamethasone treatment as
follows: 1 of 3 mice infected with the mutant and killed at 12
weeks of treatment with dexamethasone (or 24 weeks after tail
vein infection) showed 470 CFUs from the spleen homogenate
and 17 CFUs from the lung homogenate. No CFUs were re-
covered from any other mice killed at 4, 8, or 20 weeks of
treatment with dexamethasone. Each of 2 of 3 mice infected
with the wild type and killed at 20 weeks of treatment had 17
CFUs from the spleen homogenate and no CFUs from the lung
homogenate. No CFUs were recovered from mice killed at ear-
lier time points (4, 8, and 12 weeks). Figure 1. Results of tail vein infection in mice with a low dose
Low-dose tail vein infection in mice treated for 4 weeks. (5 ⫻ 102 colony-forming units [CFUs]) of the Mycobacterium tuberculosis
Erdman wild type (diamonds), M. tuberculosis H37Rv wild type (squares),
Because of the high pathogenicity of the mce1R mutant at an
mce1 operon mutant (triangles), and mce1R mutant (circles). Antibiotics
inoculum dose of 5 ⫻ 10 5 CFUs, we repeated the tail vein in- were given for 4 weeks to mice in each group. Mice in panel A were
fection with a lower dose. Surprisingly, at a dose of 5 ⫻ 10 2 treated with dexamethasone, whereas those in panel B were not. The
CFUs, a mean of ∼104 mce1R mutant bacilli were recovered number of CFUs recovered from the lungs is shown as the mean value
from the lungs of 3 mice killed 4 weeks after the antibiotics ( standard deviation) of 3 mice per group at the indicated periods of
were stopped and before the dexamethasone was initiated (Fig- infection. The dotted line indicates the limit of detection.
ure 1). The number of bacilli continued to increase until it
reached ∼106 bacilli per lung after 12 weeks of dexamethasone infected with the M. tuberculosis H37Rv wild type, mce1R mu-
treatment. No wild type or mce1 operon mutants were recov- tant, and complemented mce1R strain via the inhalation route.
ered at any time point from the lungs of mice given dexa- At 24 h after infection, 4 mice were killed from each infected
methasone, but 4 weeks after cessation of the drugs, we did group for CFU recovery in the lungs and spleen. The initial
see recovery of both strains in mice not given dexamethasone inoculum doses were (1.3–1.4) ⫻ 10 2 CFUs per mouse for each
(Figure 1B). group (Figure 2). At 4 weeks after infection, no significant
Aerosol infection in mice treated for 4 weeks. The rapid difference in the number of CFUs recovered from the lungs
reactivation of the mce1R mutant after cessation of the antibiotics ([1.0–2.5] ⫻ 10 6 CFUs) was observed in mice infected with any
was unexpected. We therefore studied mce1R in more detail, using of the M. tuberculosis stains (Figure 2A). The number of CFUs
the aerosol infection model, to compare 2 different durations of recovered from the spleens of mice infected with the mce1R
antibiotic administration (4 and 8 weeks). BALB/c mice were mutant ([9.5  3.16] ⫻ 10 4 CFUs) was similar to that recovered

754 • JID 2010:202 (1 September) • Cheigh et al


Figure 2. Results of aerosol infection in mice with the wild-type My-
cobacterium tuberculosis H37Rv strain (squares), mce1R mutant (circles),
and complemented mce1R mutant (triangles). Infected BALB/c mice in
each group were given antibiotics for 4 weeks after 4 weeks of infection.
The number of colony-forming units (CFUs) recovered from the lungs (A)
and spleens (B ) is shown as the mean value ( standard deviation) of
4 or 5 mice per group at the indicated periods of infection.
from the spleens of mice infected with the wild-type H37Rv
strain ([1.3  0.4] ⫻ 10 4 CFUs) or complemented mce1R strain
([4.3  1.4] ⫻ 10 4 CFUs) (Figure 2B).
After 4 weeks of infection, mice were given treatment with
isoniazid and pyrazinamide. The 4-week course of isoniazid
and pyrazinamide resulted in undetectable numbers of viable
bacilli in the spleen and in low numbers ([3.0–6.0] ⫻ 101 CFUs)
of bacilli in the lungs (Figures 2A and 2B). At 6 weeks after
the 4-week antibiotic course (14 weeks after infection), no sig-
nificant difference in the number of bacterial CFUs recov-
M. tuberculosis Disrupted in mce1R • JID 2010:202 (1 September) • 755
ered from the lungs was observed in mice infected with the
wild-type H37Rv strain ([1.3  0.4] ⫻ 10 4 CFUs), comple-
mented mce1R strain ([6.0  1.8] ⫻ 10 3 CFUs), or mce1R mu-
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tant ([2.6  0.5] ⫻ 10 4 CFUs) (Figure 2A). However, the num-
ber of CFUs recovered from the spleens of mice infected with
the mce1R mutant ([1.7  0.5] ⫻ 10 4 CFUs) became progres-
sively greater than that of mice infected with the H37Rv wild
type ([2.4  0.9] ⫻ 10 3 CFUs) (P ! .005) or the complemented
mce1R strain ([7.5  2.5] ⫻ 10 2 CFUs) (P ! .001) (Figure 2B).
Most importantly, by 25 weeks after infection, the number
of CFUs recovered from the lungs and spleens of mice infect-
ed with the mce1R mutant continued to increase to (1.5 
0.4) ⫻ 10 5 and (1.3  0.3) ⫻ 10 5 CFUs, respectively, whereas the
number of CFUs recovered from the organs of mice infected
with the H37Rv wild type and complemented mce1R strain re-
mained unchanged at (1.0  0.3) ⫻ 10 4 and (4.4  1.6) ⫻ 10 3
CFUs in the lungs and (1.7  0.7) ⫻ 10 3 and (5.8  1.5) ⫻ 10 3
CFUs in the spleen, respectively.
Lung histopathology in aerosol-infected mice treated for 4
weeks. Histopathological examination of lungs of mice was
performed prospectively for all mice aerosol-infected with wild-
type M. tuberculosis H37Rv, the mce1R mutant, and the com-
plemented mce1R strain. First, mice were killed 4 weeks after
infection just before the antibiotic treatment started. A large
proportion of the lung parenchyma (25%–75%) in all of the
mice contained multifocal and coalescing nodular lesions com-
prising a mixed population of foamy and epithelioid macro-
phages (Figure 3, row 1). The histology of these lung sections
was, for the most part, indistinguishable. There was no obvious
change in the proportion of lung parenchymal lesions after 4
weeks of treatment in any of the infected mice. One of the lung
sections from a mouse infected with the mce1R mutant showed
small numbers of degenerate neutrophils in the parenchyma
and mild intralumenal degenerative neutrophils.
Mice treated for 4 weeks were examined 17 weeks after ces-
sation of the treatment (25 weeks after the initial infection)
(Figure 4). The lung parenchymal involvement in mice infected
with the H37Rv strain and the complemented mutant was
25%–50%, whereas that of mice infected with the mce1R mu-
tant was !25%. In all mice, the distribution of the lung lesions
was multifocal and nodular, containing a mixed population of
foamy and epithelioid macrophages. No significant difference
in the number of acid-fast bacilli was observed between the
lung sections of mice infected with the wild-type H37Rv
strain, complemented mce1R strain, or mce1R mutant (data
not shown).
Aerosol infection in mice treated for 8 weeks. Because the
wild type and complemented mutant could still be recovered
from mice treated for 4 weeks, we extended the treatment to
8 weeks. The numbers of bacterial CFUs recovered at time
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Figure 3. Histology (hematoxylin and eosin stain) of lung sections of mice infected for 4 weeks before treatment was initiated (row 1), at the end
of 8 weeks of treatment (row 2 ), and 20 weeks after the end of an 8-week treatment (row 3 ). Mice were infected with the wild-type H37Rv strain
(A), complemented mce1R mutant strain (B ), or mce1R mutant strain (C ) of Mycobacterium tuberculosis via the inhalation route. Original magnification,
⫻200; scale bar, 40 mm.

points before the 8-week medication course were similar to
those recovered before the 4-week medication course.
The 8-week course of isoniazid and pyrazinamide result-
ed in undetectable numbers of viable bacilli in the lungs and
spleens of mice in all infected groups at the end of the treatment
course (Figure 5). At 6 weeks after the 8-week treatment course,
(18 weeks after infection), (1.3  1.0) ⫻ 101 CFUs from the
lungs and (2.0  1.8) ⫻ 101 CFUs from the spleens of mice
infected with the mce1R mutant were recovered, whereas no
CFUs were recovered from any of the organs of mice infected
with the H37Rv wild type or complemented mce1R strain (Fig-
ures 5A and 5B).
At 24 and 32 weeks after infection, the number of CFUs
recovered from mice infected with the mce1R mutant continued
to increase from (7.5  5.5) ⫻ 101 to (5.3  4.0) ⫻ 10 2 CFUs in
the lungs and from (1.1  0.9) ⫻ 10 2 to (4.4  3.8) ⫻ 10 2 CFUs
in the spleen, whereas the number of CFUs recovered from the
organs of mice infected with H37Rv wild type and comple-
mented mce1R strain remained undetectable. Interestingly, no
mce1R mutant was detected in the lungs or spleens of any mice
at 3 weeks after cessation of the 8-week antibiotic course, which
suggests that during this period, this mutant was indeed in a
latent state.
Lung histopathology in aerosol-infected mice treated for 8
weeks. At the end of 8 weeks of treatment, the proportion of
the parenchymal lung involvement in all mice decreased from
that during the period just before the treatment was started
(Figure 3). In those infected with the H37Rv strain or the
complemented mce1R mutant, the proportion of parenchymal
involvement decreased to !25% in most of the lung sections
(Figure 3, row 2). In mice infected with the mce1R mutant, the
parenchymal involvement also decreased to 25%–50% (Figure

756 • JID 2010:202 (1 September) • Cheigh et al

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Figure 4. Histopathology of lung sections from BALB/c mice infected with wild-type Mycobacterium tuberculosis strain H37Rv (A), complemented
mce1R mutant (B ), or mce1R mutant (C ). The mice were infected via the inhalation route; after 4 weeks, they were given a 4-week course of
antibiotics. Lung sections from 4 mice in each group were harvested at 17 weeks after the end of treatment (25 weeks after infection) and stained
with hematoxylin and eosin. Lung tissues in all groups show granulomatous interstitial pneumonia. Original magnifications, ⫻100 (row 1) and ⫻400
(row 2 ); scale bars, 80 mm (row 1) and 20 mm (row 2 ).

3C, row 2). Granulomatous lesions were rare and small in mice
infected with the mce1R mutant, and no such lesions were
observed in mice infected with the other strains.
Lung sections of the remaining mice were harvested at 20
weeks after cessation of the 8-week treatment (32 weeks after
infection) (Figure 3, row 3). At this time point, the lung lesions
were much milder than those in the lungs of mice observed at
17 weeks after 4 weeks of treatment (Figure 4). Nodular lung
lesions were observed only in mice infected with mce1R mutant
(Figure 3C, row 3). Lung sections of mice infected with mce1R
mutant showed mild granulomatous pneumonia with multi-
focal nodular lesions that were characterized by scattered foci
of alveolar macrophages, with occasional foamy alveolar mac-
rophages, and large focal aggregates of lymphocytes within the
nodule (Figure 3C, row 3).
DISCUSSION
In this study, we used a variant of the Cornell mouse model
of latent tuberculosis to study latent infection with M. tuber-
culosis mce1 operon and mce1R mutants. The unexpected ob-
servation was the rapid posttreatment proliferation of the
mce1R mutant. At similar infection inoculum doses, the mce1R
mutant rapidly proliferated in mouse lungs and spleens after
the cessation of antibiotics, regardless of whether the mice were
infected via the tail vein or by the aerosol route. The mce1R
mutant, as well as the other strains, exhibited a similar level of
susceptibility to the drugs during the treatment course, as evi-
denced by the low or absent CFU counts at the end of each
treatment course (Figures 1, 2, and 5). All infected mice initially
responded clinically to the treatment, as evidenced by pro-
gressive resolution of the lung lesions (Figure 3). However,
among mice treated for 8 weeks, only those infected with the
mce1R mutant showed recrudescence of the lung lesions after
treatment cessation. This recrudescence correlated with in-
creasing numbers of CFUs recovered from the lungs (Figure
5). Thus, the mce1R mutant appears to cause a true posttreat-
ment reactivation of tuberculosis in mice. The fact that the
mce1R mutant is regulated in vivo [18, 19] suggests that this
response can occur in a natural course of infection with wild-
type M. tuberculosis in humans. Indeed, posttreatment reacti-
vation of tuberculosis in humans is not uncommon among
those who do not complete a standard 6-month course of
treatment, even with drug-susceptible tuberculosis.
We showed that at 3 weeks after cessation of the drugs, the
mce1R mutant could not be cultured from lungs (Figure 5).
Thus, this phase of infection was consistent with this organism

M. tuberculosis Disrupted in mce1R • JID 2010:202 (1 September) • 757


Figure 5. Results of a modified Cornell model of infection with wild-
type Mycobacterium tuberculosis strain H37Rv (squares), mce1R mutant
(circles), and complemented mce1R mutant (triangles). BALB/c mice were
infected via the inhalation route, and antibiotics were given to mice in
each group for 8 weeks after 4 weeks of infection. The number of colony-
forming units (CFUs) recovered from the lungs (A) and spleens (B ) is
shown as the mean value ( standard deviation) of 4 or 5 mice per
group at the indicated periods of infection.
being in a latent state of infection. Here, the drugs may have
reduced the bacterial count to a value below the threshold
recoverable on the Middlebrook 7H11 agar plate. We note that
it is possible that the treatment that resulted in no recovery of
the wild type represents complete sterilization and not latent
infection. However, at least in 1 experiment, we did observe
later recovery of wild-type bacteria from a subset of infected
mice, as depicted in Figure 1B.
Six of the mce1 genes (mce1A–mce1AF) encode cell wall
758 • JID 2010:202 (1 September) • Cheigh et al
proteins that resemble substrate proteins of ABC transporters,
whereas the upstream genes yrbE1A and yrbE1B encode pro-
teins that resemble ABC transporter permeases [16]. ABC trans-
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porters use an adenosine triphosphatase (ATPase) to supply
energy for translocation of a substrate across a membrane [23].
Dassa and Bouige [23] have suggested that RvO655, an ATPase
called Mkl, may serve this function for M. tuberculosis. Joshi
et al [24] have shown that RvO655 and mce1 are functionally
linked and that the mce1 operon may encode an ABC trans-
porter involved in lipid import. Interestingly, the mce1 operon
carries a gene, fadD5, that encodes a fatty acyl co-A synthetase,
which is putatively involved in fatty acid catabolism [15]. Dun-
phy et al [25] recently suggested that FadD5 may be involved
in recycling mycolic acids from dying M. tuberculosis inside
granulomas. Mce1R belongs to a family of GntR-negative tran-
scriptional regulators involved in lipid transport and metab-
olism in other bacterial organisms [18, 26]. Another member
of the mce family of operons, mce4, was recently shown to serve
as a cholesterol importer that is necessary for persistent infec-
tion in mice [27]. These observations together suggest that the
mce operon family is involved in lipid transport across the M.
tuberculosis cell wall and membrane, possibly for the organism
to gain nutrients (carbon as an energy source) during its long-
term survival inside granulomas.
Beste et al [28] recently reported that in Mycobacterium bovis
bacille Calmette-Guérin, the mce1 operon may be involved in
the bacillus switching to a slow growth rate, which supports
the idea that the regulation of the mce1 operon by mce1R can
affect M. tuberculosis replication in vivo. The mce1 genes in the
mce1R mutant are constitutively expressed, whereas in the wild
type, these genes are repressed during the early phase of in-
fection in vivo [18, 19]. The expression of the mce1 genes may
allow M. tuberculosis to import and metabolize lipids for its
survival during the persistent state. When the number of or-
ganisms is reduced by treatment to a level that is undetectable
in culture, wild-type strains that do not express the mce1 genes
may become restricted for replication by the host immune
response, whereas the mce1R mutant strains expressing the mce1
genes would continue to take up and metabolize lipids and
replicate. We have shown elsewhere that wild-type M. tuber-
culosis expresses mce1 genes after ∼8 weeks into infection in
mice [19]. We do not know at this time what triggers expression
or repression of these genes in vivo. In a course of persistent
infection, an infected host may harbor a mixed population of
strains that express these genes and strains that do not express
these genes. The progression to reactivation of disease, as op-
posed to remaining in latent state, may depend on the balance
of these 2 bacterial populations. Factors that disrupt this bal-
ance, such as immunosuppression, old age, large initial infec-
tious inoculum doses, or certain M. tuberculosis strain-related
factors, could lead to the reactivation of tuberculosis.
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