Previous Article | Next Article 
Journal of Clinical Microbiology, November 1999, p. 3693-3697, Vol. 37, No. 11
0095-1137/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
Simple and Rapid Identification of the
Mycobacterium tuberculosis Complex by Immunochromatographic
Assay Using Anti-MPB64 Monoclonal Antibodies
Chiyoji
Abe,1,*
Kazue
Hirano,1 and
Tetsuo
Tomiyama2
Research Institute of Tuberculosis, Japan
Anti-Tuberculosis Association, Kiyose-shi, Tokyo
204-0022,1 and Tomiyama
Laboratories, Nerima-ku, Tokyo 178-0061,2 Japan
Received 10 March 1999/Returned for modification 22 June
1999/Accepted 23 July 1999
 |
ABSTRACT |
A newly developed immunochromatographic assay (MPB64-ICA) for
identification of the Mycobacterium tuberculosis complex
was evaluated with 20 reference strains of mycobacterial species and 111 clinical isolates. MPB64-ICA displayed a very strong reaction band
with organisms belonging to the M. tuberculosis complex but not with mycobacteria other than M. tuberculosis (MOTT
bacilli), except for one of four M. marinum strains tested
and one M. flavescens strain, both of which gave very weak
signals. The effectiveness of MPB64-ICA in combination with two liquid
culture systems (MB-REDOX and MGIT) was tested. A total of 108 of 362 sputum specimens processed were positive for acid-fast bacilli. Samples
taken from the cultures on the same days when either of the two culture
systems became positive for mycobacteria were assayed with MPB64-ICA.
Of 108 cultures with mycobacteria, 51 showed a positive signal with the test, in which the presence of the M. tuberculosis complex
was demonstrated later by the Accuprobe for M. tuberculosis
complex. In addition, MPB64-ICA could correctly detect the M. tuberculosis complex in mixed cultures of the M. tuberculosis complex and MOTT bacilli. These results indicate
that MPB64-ICA can be easily used for rapid identification of the
M. tuberculosis complex in combination with culture systems
based on liquid media without any technical complexity in clinical laboratories.
| |
INTRODUCTION |
Mycobacterium
tuberculosis is a facultative intracellular bacterium that causes
tuberculosis. Presently, tuberculosis affects 1.7 billion people
worldwide. There are 8 million new cases and around 3 million deaths
per year (30). The situation is further complicated by the
emergence of multidrug-resistant strains (24, 31).
Therefore, there is a need for rapid identification of mycobacteria and
rapid drug susceptibility testing for effective treatment of the disease.
To identify mycobacteria, conventional biochemical tests are
traditionally used (4, 14, 15, 29, 33). Key tests can be
used to identify species, or further preliminary grouping may be used.
Other approaches to identifying some species of mycobacteria are
available. They include the p-nitrobenzoic acid (23,
28) and
p-nitro-
-acetylamino-
-hydroxypropiophenone (17,
23) tests for discrimination of the M. tuberculosis
complex from mycobacteria other than M. tuberculosis (MOTT
bacilli); DNA probe methods for identification or confirmation of the
M. tuberculosis complex, M. avium complex,
M. kansasii, and M. gordonae (10, 11);
and gas-liquid chromatography or high-performance liquid chromatography analyses for recognizing the patterns of the mycobacterial cell wall
fatty acids or mycolic acids (26). The advantages of the last three methods are that they are capable of providing definitive identification within 2 to 4 h after adequate growth.
The immunogenic protein MPB64 has been found in unheated culture fluids
of M. tuberculosis H37Rv and in some strains of M. bovis BCG (3, 12, 21). This antigen induced a strong
delayed-type hypersensitivity reaction similar to that induced by
purified protein derivatives in guinea pigs sensitized with these
strains, whereas no reaction to MPB64 was observed with M. kansasii or M. intracellulare (12). The
MPB64 antigen has been shown to be specific for the M. tuberculosis complex (12). Thus, MPB64 could be useful
in studies on the pathogenesis and cell-mediated immunology of
mycobacteria and in the development of diagnostic tests. In the present
study, a newly developed (27) immunochromatographic assay
(MPB64-ICA) using anti-MPB64 monoclonal antibodies for rapid discrimination between the M. tuberculosis complex and MOTT
bacilli was evaluated with reference strains of 20 mycobacterial
species, 111 clinical isolates, 108 liquid cultures inoculated with
clinical specimens, and 7 M. bovis BCG substrains, and the
results were compared with those of other identification tests.
| |
MATERIALS AND METHODS |
Mycobacterial strains and clinical specimens.
Reference
strains of 20 Mycobacterium species (Table
1), 53 M. tuberculosis
clinical isolates, 55 MOTT clinical isolates (3 strains of M. abscessus, 13 strains of M. avium complex, 2 strains of
M. gordonae, 30 strains of M. kansasii, 3 strains
of M. marinum, 3 strains of M. szulgai, and 1 unidentified strain), 7 M. bovis BCG substrains (Glaxo,
Pasteur, Tice, Brazilian, Japanese, Russian, and Swedish), and 3 mixed
cultures with M. tuberculosis and MOTT bacilli were used for
this study. Clinical specimens, mostly sputum, were obtained from 362 different patients admitted to Fukujuji Hospital (Japan
Anti-Tuberculosis Association, Kiyose-shi, Tokyo) with symptoms of
pulmonary diseases.
All specimens were decontaminated by the
N-acetyl-L-cysteine-NaOH (NALC-NaOH) method,
which was slightly modified from the original (16). Two
volumes of NALC-NaOH solution (2% NaOH, 1.45% Na-citrate, 0.5% NALC)
were mixed with the specimen on a test tube mixer for digestion, and
the mixtures were allowed to stand for 15 min at room temperature. Ten
volumes of 10 mM phosphate buffer (pH 7.4) were added for dilution, and
the mixtures were centrifuged at 3,000 × g for 20 min
at 4°C. After the supernatant fluids were carefully decanted, the
resulting sediments were suspended in 1 ml of the same buffer.
Media and culture methods.
The Mycobacteria Growth Indicator
Tube (MGIT) (Nippon Becton Dickinson Co., Ltd., Tokyo), which is a
recently introduced nonradiometric culture system, uses an
oxygen-quenched fluorescent indicator (1, 6, 8, 22). A
fluorescent compound, ruthenium metal complex, is embedded in silicone
on the bottom of 16- by 100-mm round-bottom tubes. Actively respiring
mycobacteria consume the dissolved oxygen and allow the fluorescence to
be observed with a 365 nm UV transilluminator. The MGIT contains 4 ml
of modified Middlebrook 7H9 liquid medium. The supplement contains
casein peptone, albumin, dextrose, catalase, oleic acid, polymyxin B, amphotericin B, nalidixic acid, trimethoprim, and azlocillin.
MB-REDOX (Biotest AG, Frankfurt, Germany) is serum-supplemented,
modified Kirchner medium (4 ml) containing a colorless tetrazolium
salt, special vitamin complex, and the antibiotic admixture PACT
(polymixin B, amphotericin B, carbenicillin, and trimethoprim)
(
7). In this method, the tetrazolium salt is reduced by the
redox system of the mycobacteria to a brownish formazan. This
formazan
is water insoluble and is easily detected by the naked
eye. The
MB-REDOX medium was kindly provided by Nissui Pharmaceutical
Co., Ltd.,
Tokyo,
Japan.
A 0.2-ml portion of each pretreated specimen was inoculated into MGIT
and MB-REDOX tubes. All media were incubated at 37°C
and checked
twice a week for the first 4 weeks and once a week
for the remaining
culture
time.
Identification of mycobacterial isolates.
All isolates were
differentiated and identified by an RNA-DNA hybridization assay
(4, 10, 11) with commercial kits for culture confirmation
and identification of species belonging to the M. tuberculosis complex and M. avium complex (Gen-Probe, San Diego, Calif.) and by conventional culturing or biochemical testing
(4, 14). Niacin test strips were purchased from Kyokuto Pharmaceutical Co., Ltd., Tokyo, Japan (15, 33).
MPB64-ICA.
MPB64-ICA is a rapid immunochromatographic
identification test for the M. tuberculosis complex that
uses anti-MPB64 monoclonal antibodies (27). Monoclonal
antibodies were produced from hybridomas obtained by the fusion of P3U1
myeloma cells with spleen cells of mice immunized with an MPB64
antigen. The test strip consists of a sample pad, a reagent pad, a
nitrocellulose membrane, and an absorbent pad. The antibodies were
immobilized on the nitrocellulose membrane as the capture reagent (test
line). Secondary antibodies, which recognized another epitope of the
antigen, conjugated with colloidal gold particles were used for antigen
capture and detection in a sandwich-type assay.
As the test sample applied in the sample well flows laterally through
the membrane, the antibody-colloidal gold conjugate
binds to the MPB64
antigen in the sample. The complex then flows
further and binds to the
monoclonal antibodies on the solid phase
in the test zone, producing a
red band. In the absence of MPB64,
there is no line in the positive
reaction zone. The liquid continues
to migrate along the membrane and
produces a red band in the control
zone, demonstrating that the
reagents are functioning properly
(Fig.
1).
View larger version (82K):
[in this window]
[in a new window]
|
FIG. 1.
Identification of the M. tuberculosis complex
by MPB64-ICA. Negative, only one reddish-purple band appears in the
control (C) window. Positive, in addition to the control band, a clear
distinguishable reddish-purple band also appears in the test (T)
window.
|
|
For sample preparation from solid cultures, a loopful (1 µl) of
growth was vortexed for 1 min in a screw-capped tube containing
200 µl of 10 mM phosphate buffer-0.1% Tween 80 and 5 to 10 pieces
of
glass beads. One hundred microliters of each treated sample
was applied
to the sample wells and they were allowed to stand
at room temperature
for 15 min. If the sample contains MPB64,
a red band is produced in the
test zone. Liquid cultures (100
µl) were applied directly to the
sample wells without use of the
sample preparation
procedure.
| |
RESULTS |
Reference strains of 20 mycobacterial species were subjected to
MPB64-ICA for identification of the M. tuberculosis complex. Samples prepared from Ogawa egg slants were added to the sample wells
of the cassette. As shown in Table 1, strong reaction bands against the
bacteria belonging to the M. tuberculosis complex, M. tuberculosis, M. africanum, and M. bovis
were demonstrated, while no positive signal was observed for the MOTT
bacilli tested, except for M. flavescens, which showed a
very weak signal. Next, to further confirm the specificity of the test,
the results with MPB64-ICA were compared with those of the
Accuprobe-M. tuberculosis complex culture confirmation test
(Accuprobe) and the niacin accumulation test with 111 mycobacterial
cultures. These included 53 M. tuberculosis isolates, 55 MOTT isolates, and 3 mixed cultures with M. tuberculosis and
MOTT bacilli (Table 2). All 53 M. tuberculosis isolates and the 3 mixed cultures were positive for
the M. tuberculosis complex by both the MPB64-ICA and
Accuprobe tests. On the other hand, the niacin test failed to obtain a
positive result from the three mixed cultures, which were confirmed to
contain the M. tuberculosis complex by the MPB64-ICA and
Accuprobe tests. MPB64-ICA showed a positive result for one of three
M. marinum clinical isolates tested, although the reaction
was very weak. Of 55 MOTT cultures, one was positive in the niacin
test; this was an M. kansasii culture and did not seem to
contain M. tuberculosis according to the results of the
other two tests.
View this table:
[in this window]
[in a new window]
|
TABLE 2.
Comparison of the results obtained by MPB64-ICA,
Accuprobe for M. tuberculosis complex, and niacin
accumulation tests
|
|
To investigate the sensitivity of the assay, 10-fold dilutions of the
M. bovis BCG Japanese strain grown in Middlebrook 7H9 liquid
medium were made in cold 10 mM phosphate buffer containing 0.1% Tween
80. The organisms were washed once with the same buffer before the
dilution to remove MPB64 antigen secreted in the culture medium. The
number of organisms was determined on Middlebrook 7H11 agar plates.
Each dilution containing M. bovis BCG organisms was vortexed
in the tube containing glass beads and then applied to the test
cassette. The detection limit of MPB64-ICA was estimated as
105 CFU/ml (Table 3).
The usefulness of the culture systems based on liquid media had been
confirmed by many studies. In this study, we tried to do early
confirmation of the M. tuberculosis complex in liquid cultures by MPB64-ICA. Two culture systems, the MGIT and MB-REDOX systems, were used for the evaluation. Samples taken from the cultures
when they showed a positive signal for mycobacteria in the liquid
cultures were assayed with MPB64-ICA. A total of 108 of 362 sputum
specimens processed were positive for mycobacteria (acid-fast bacilli).
Fifty-one cultures tested positive with MPB64-ICA, and the presence of
the M. tuberculosis complex in these cultures was
demonstrated by other confirmation tests (Table
4). One culture was a mixed culture with
M. tuberculosis complex and M. avium complex
bacilli; the M. tuberculosis complex was detected exactly by
the test. All 57 cultures with MOTT bacilli were negative with MPB64-ICA. The mean times for the confirmation of the M. tuberculosis complex were 15 and 16.5 days from processing of the
specimens with the MB-REDOX and MGIT systems, respectively.
It is well known that M. bovis BCG vaccine strains can be
divided into two groups on the basis of several features, such as the
number of IS6110 copies (9, 25) and the presence
of MPB64 antigen (12, 21) and methoxymycolates
(19). As shown in Table 5,
these two groups of the substrains were clearly differentiated by
MPB64-ICA.
| |
DISCUSSION |
The immunochromatographic test used here is of a new generation
and is one of the simplest and fastest tests to perform. The MPB64
antigen was found in the culture fluid of only the M. tuberculosis complex and some strains of M. bovis BCG
(12, 21). The specificity of the antigen for the M. tuberculosis complex was confirmed by a
radioimmunoassay-inhibition assay using anti-MPB64 antibodies (12). The purpose of this study was to evaluate the newly
developed immunochromatographic assay, MPB64-ICA (27), for
identification of the M. tuberculosis complex with the
reference strains of the clinically important mycobacterial species and
with clinical isolates. MPB64-ICA showed a very strong signal with the
organisms belonging to the M. tuberculosis complex but not
with MOTT bacilli, except for one of four strains of M. marinum tested and one M. flavescens strain, which
showed a very weak signal (Tables 1 and 2). These two species may be
easily differentiated from the M. tuberculosis complex by
the following properties: M. marinum has photochromogenic and smooth-type colonies on solid media and is isolated from skin lesions in most cases (5, 29), M. flavescens has
pigmented, smooth-type colonies (29), and both normally grow rapidly.
The MPB64 antigen is found in the culture fluids of the M. tuberculosis complex. The antigen is secreted in significant
amounts during the early period of culturing and decreases with longer cultivation (21). A gene cloning study has shown that the
antigen is synthesized with a signal peptide characteristic of a
secreted protein (32).
During the past decade, culture systems based on liquid media have been
introduced for primary isolation of mycobacteria from clinical
specimens in many countries (1, 2, 6-8, 13, 18, 20, 22).
Here we used two culture systems with different detection systems for
mycobacteria, MB-REDOX and MGIT. A total of 108 cultures of 362 sputum
specimens processed were positive for acid-fast bacilli in these two
systems. When the samples from these positive cultures were assayed
with MPB64-ICA, 51 showed positive results for the MPB64 antigen (Table
4). In the Accuprobe test, all 51 cultures were confirmed to contain
the M. tuberculosis complex bacilli. One culture was a
mixture of M. tuberculosis complex and M. avium
complex, again demonstrating the usefulness of the test in mixed
cultures. The mean detection times for the M. tuberculosis
complex were 15 and 16.5 days with the MB-REDOX and MGIT systems,
respectively. A positive signal for MPB64-ICA was obtained from all of
51 cultures on the same days when these culture systems had become
mycobacterium positive. It is very important to discriminate early
between M. tuberculosis and MOTT bacilli for appropriate
management of the patients and effective treatment of the disease.
MPB64-ICA can be easily used for rapid identification of the M. tuberculosis complex in combination with the culture systems based
on liquid media without any troublesome sample preparation in a laboratory.
Mixed mycobacterial infections with two species occur less frequently
in human immunodeficiency virus negative patients, but they are not
rare and are difficult to diagnose by conventional methods. In this
study, we tested four mixed cultures (Tables 2 and 4). All were
cultures with the M. tuberculosis complex and MOTT bacilli;
two contained the M. avium complex and one each contained
M. kansasii and M. gordonae. MPB64-ICA could
detect the M. tuberculosis complex in all of these cultures.
The Accuprobe test also confirmed exactly the presence of the M. tuberculosis complex, but the niacin test failed to detect it in
all four cultures.
When fresh M. bovis BCG grown in a liquid medium was used
for the evaluation, the analytical sensitivity of MPB64-ICA was calculated as 105 CFU/ml (Table 3), a value similar to that
for the Accuprobe test (10, 11). Therefore, this test should
not be used directly on fresh clinical specimens.
M. bovis BCG is isolated occasionally from individuals with
lymphadenitis occurring after M. bovis BCG vaccination or
from bladder cancer patients with M. bovis BCG instillation
therapy. However, it is not easy to differentiate between M. tuberculosis and M. bovis BCG. Harboe et al. showed
that the Brazilian, Japanese, Russian, and Swedish strains of M. bovis BCG possessed an MPB64 antigen but that the Pasteur, Glaxo,
and Tice strains did not (12). In the present study, it was
found that two groups of M. bovis BCG could be clearly
differentiated by MPB64-ICA (Table 5). In countries where the Pasteur,
Glaxo, or Tice strain of M. bovis BCG was used for
vaccination, M. bovis BCG isolates may be easily
discriminated from M. tuberculosis by MPB64-ICA.
| |
ACKNOWLEDGMENTS |
This work was partly supported by Health Sciences Research grants
from the Ministry of Health and Welfare of Japan (Research on Emerging
and Re-emerging Infectious Diseases) and by the Tuberculosis and
Leprosy panel, U.S.-Japan Cooperative Medical Science Program.
| |
FOOTNOTES |
*
Corresponding author. Mailing address: Research
Institute of Tuberculosis, Japan Anti-Tuberculosis Association,
Kiyose-shi, Tokyo 204-0022, Japan. Phone: 81-424-93-5773, Fax:
81-424-92-4600. E-mail: hirano{at}jata.or.jp.
| |
REFERENCES |
| 1.
|
Abe, C.,
K. Hirano, and M. Wada.
1998.
Evaluation of the Mycobacteria Growth Indicator Tube system using oxygen sensitive fluorescent sensor for rapid detection of mycobacteria, p. 175-180.
In
M. Casal (ed.), Clinical mycobacteriology. Prous Science, Barcelona, Spain
|
| 2.
|
Abe, C.,
S. Hosojima,
Y. Fukasawa,
Y. Kazumi,
M. Takahashi,
K. Hirano, and T. Mori.
1992.
Comparison of MB-Check, BACTEC, and egg-based media for recovery of mycobacteria.
J. Clin. Microbiol.
30:878-881[Abstract/Free Full Text].
|
| 3.
|
Abou-Zeid, C.,
M. Harboe, and G. A. W. Rook.
1987.
Characterization of the secreted antigens of Mycobacterium bovis BCG: comparison of the 46-kilodalton dimeric protein with proteins MPB64 and MPB70.
Infect. Immun.
55:3213-3214[Abstract/Free Full Text].
|
| 4.
|
American Society for Microbiology.
1995.
Section 3. Mycobacteriology.
In
Clinical microbiology procedures handbook. American Society for Microbiology, Washington, D.C
|
| 5.
|
Aronson, J. D.
1926.
Spontaneous tuberculosis in salt fish.
J. Infect. Dis.
39:315.
|
| 6.
|
Badak, F. Z.,
D. L. Kiska,
S. Setterquist,
C. Hartley,
M. A. O'Conell, and R. L. Hopfer.
1996.
Comparison of Mycobacteria Growth Indicator Tube with BACTEC 460 for detection and recovery of mycobacteria from clinical specimens.
J. Clin. Microbiol.
34:2236-2239[Abstract].
|
| 7.
|
Cambau, E.,
C. Wichlacz,
C. Truffot-Pernot, and V. Jarlier.
1999.
Evaluation of the new MB Redox system for detection of growth of mycobacteria.
J. Clin. Microbiol.
37:2013-2015[Abstract/Free Full Text].
|
| 8.
|
Casal, M.,
J. Gutierrez, and M. Vaquero.
1997.
Comparative evaluation of the Mycobacteria Growth Indicator Tube with the BACTEC 460TB system and Löwenstein-Jensen medium for isolation of mycobacteria from clinical specimens.
Int. J. Tuberc. Lung Dis.
1:81-84[Medline].
|
| 9.
|
Fomukong, N. G.,
J. W. Dale,
T. W. Osborn, and J. M. Grange.
1992.
Use of gene probes on the insertion sequence IS986 to differentiate between BCG vaccine strains.
J. Appl. Bacteriol.
72:126-133[Medline].
|
| 10.
|
Gen-Probe, Inc.
1990.
Accuprobe mycobacterium culture confirmation tests.
Gen-Probe, Inc., San Diego, Calif
|
| 11.
|
Goto, M.,
S. Oka,
K. Okuzumi,
S. Kimura, and K. Shimada.
1991.
Evaluation of acridinium ester-labeled DNA probes for identification of Mycobacterium tuberculosis and Mycobacterium avium-Mycobacterium intracellulare complex in culture.
J. Clin. Microbiol.
29:2473-2476[Abstract/Free Full Text].
|
| 12.
|
Harboe, M.,
S. Nagai,
M. E. Patarroyo,
M. L. Torres,
C. Ramirez, and N. Cruz.
1986.
Properties of proteins MPB64, MPB70, and MPB80 of Mycobacterium bovis BCG.
Infect. Immun.
52:293-302[Abstract/Free Full Text].
|
| 13.
|
Isenberg, H. D.,
R. F. D'Amato,
L. Heifets,
P. R. Murray,
M. Scardamaglia,
M. C. Jacobs,
P. Alperstein, and A. Niles.
1991.
Collaborative feasibility study of a biphasic system (Roche Septi-Chek AFB) for rapid detection and isolation of mycobacteria.
J. Clin. Microbiol.
29:1719-1722[Abstract/Free Full Text].
|
| 14.
|
Kent, P. T., and G. P. Kubica.
1985.
Public health mycobacteriology. A guide for the level III laboratory.
Centers for Disease Control, Atlanta, Ga
|
| 15.
|
Konno, K.
1956.
New chemical method to differentiate human-type tubercle bacilli from other mycobacteria.
Science
124:985[Free Full Text].
|
| 16.
|
Kubica, G. P. W.,
E. Dye,
M. L. Cohn, and G. Middlebrook.
1963.
Sputum digestion and decontamination with N-acetyl-L-cysteine-sodiumhydroxide for culture of mycobacteria.
Am. Rev. Respir. Dis.
87:775-779[Medline].
|
| 17.
|
Laszlo, A., and E. Siddiqi.
1984.
Evaluation of a rapid radiometric differentiation test for the Mycobacterium tuberculosis complex by selective inhibition with p-nitro--acetylamino--hydroxypropiophenone.
J. Clin. Microbiol.
19:694-698[Abstract/Free Full Text].
|
| 18.
|
Middlebrook, G.,
Z. Reggiardo, and W. D. Tigertt.
1977.
Automatable radiometric detection of growth of Mycobacterium tuberculosis in selective media.
Am. Rev. Respir. Dis.
115:1066-1069[Medline].
|
| 19.
|
Minnikin, D. E.,
J. H. Parlett,
M. Magnusson,
M. Ridell, and A. Lind.
1984.
Mycolic acid patterns of representatives of Mycobacterium bovis BCG.
J. Gen. Microbiol.
130:2733-2736[Abstract/Free Full Text].
|
| 20.
|
Morgan, M. A.,
C. D. Horstmeier,
D. R. DeYoung, and G. D. Roberts.
1983.
Comparison of a radiometric method (BACTEC) and conventional culture media for recovery of mycobacteria from smear-negative specimens.
J. Clin. Microbiol.
18:384-388[Abstract/Free Full Text].
|
| 21.
|
Nagai, S.,
H. G. Wiker,
M. Harboe, and M. Kinomoto.
1991.
Isolation and partial characterization of major protein antigens in the culture fluid of Mycobacterium tuberculosis.
Infect. Immun.
59:372-382[Abstract/Free Full Text].
|
| 22.
|
Pfyffer, G. E.,
H.-M. Welscher,
P. Kissling,
C. Cieslak,
M. J. Casal,
J. Gutierrez, and S. Rüsch-Gerdes.
1997.
Comparison of the Mycobacteria Growth Indicator Tube (MGIT) with radiometric and solid culture for recovery of acid-fast bacilli.
J. Clin. Microbiol.
35:364-368[Abstract].
|
| 23.
|
Rastogi, N.,
K. S. Goh, and H. L. David.
1989.
Selective inhibition of the Mycobacterium tuberculosis complex by p-nitro--acetylamino--hydroxypropiophenone (NAP) and p-nitrobenzoic acid (PNB) used in 7H11 agar medium.
Res. Microbiol.
140:419-423[Medline].
|
| 24.
|
Snider, D. E., and W. L. Roper.
1992.
The new tuberculosis.
N. Engl. J. Med.
326:703-705[Medline].
|
| 25.
|
Takahashi, M.,
Y. Kazumi,
Y. Fukasawa,
K. Hirano,
T. Mori,
J. W. Dale, and C. Abe.
1993.
Restriction fragment length polymorphism analysis of epidemiologically related Mycobacterium tuberculosis isolates.
Microbiol. Immunol.
37:289-294[Medline].
|
| 26.
|
Tisdall, P. A.,
D. R. DeYoung,
G. D. Roberts, and J. P. Anhalt.
1982.
Identification of clinical isolates of mycobacteria with gas-liquid chromatography: a 10-month follow-up study.
J. Clin. Microbiol.
16:400-402[Abstract/Free Full Text].
|
| 27.
|
Tomiyama, T.,
K. Matsuo, and C. Abe.
1997.
Rapid identification of Mycobacterium tuberculosis by an immunochromatography using anti-MPB64 monoclonal antibodies.
Int. J. Tuberc. Lung Dis.
1(Suppl. 1):S59.
|
| 28.
|
Tsukamura, M., and S. Tsukamura.
1964.
Differentiation of Mycobacterium tuberculosis and Mycobacterium bovis by p-nitrobenzoic acid susceptibility.
Tubercle
45:64-65[Medline].
|
| 29.
|
Wayne, L. G., and G. P. Kubica.
1986.
The mycobacteria, p. 1435-1457.
In
P. H. A. Sneath, N. S. Mair, M. E. Sharp, and J. G. Holt (ed.), Bergey's manual of systematic bacteriology, vol. 2. Williams & Wilkins, Baltimore, Md
|
| 30.
|
World Health Organization.
1996.
Report of the tuberculosis epidemic.
World Health Organization, Geneva, Switzerland
|
| 31.
|
World Health Organization.
1997.
Anti-tuberculosis drug resistance in the world. The WHO/IUATLD global project on anti-tuberculosis drug-resistance surveillance, 1994-1997.
World Health Organization, Geneva, Switzerland
|
| 32.
|
Yamaguchi, R.,
K. Matsuo,
A. Yamazaki,
C. Abe,
S. Nagai,
K. Terasaka, and T. Yamada.
1989.
Cloning and characterization of the gene for immunogenic protein MPB64 of Mycobacterium bovis BCG.
Infect. Immun.
57:283-288[Abstract/Free Full Text].
|
| 33.
|
Young, W. D., Jr.,
A. Maslansky,
M. S. Lefar, and D. P. Kronish.
1970.
Development of a paper strip test for detection of niacin produced by mycobacteria.
Appl. Microbiol.
10:939-945.
|
Journal of Clinical Microbiology, November 1999, p. 3693-3697, Vol. 37, No. 11
0095-1137/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
This article has been cited by other articles:
-
Park, M. Y., Kim, Y. J., Hwang, S. H., Kim, H. H., Lee, E. Y., Jeong, S. H., Chang, C. L.
(2009). Evaluation of an Immunochromatographic Assay Kit for Rapid Identification of Mycobacterium tuberculosis Complex in Clinical Isolates. J. Clin. Microbiol.
47: 481-484
[Abstract]
[Full Text]
-
Abe, C., Kobayashi, I., Mitarai, S., Wada, M., Kawabe, Y., Takashima, T., Suzuki, K., Sng, L.-H., Wang, S., Htay, H. H., Ogata, H.
(2008). Biological and Molecular Characteristics of Mycobacterium tuberculosis Clinical Isolates with Low-Level Resistance to Isoniazid in Japan. J. Clin. Microbiol.
46: 2263-2268
[Abstract]
[Full Text]
-
Hirano, K., Aono, A., Takahashi, M., Abe, C.
(2004). Mutations Including IS6110 Insertion in the Gene Encoding the MPB64 Protein of Capilia TB-Negative Mycobacterium tuberculosis Isolates. J. Clin. Microbiol.
42: 390-392
[Abstract]
[Full Text]
-
Somoskovi, A., Song, Q., Mester, J., Tanner, C., Hale, Y. M., Parsons, L. M., Salfinger, M.
(2003). Use of Molecular Methods To Identify the Mycobacterium tuberculosis Complex (MTBC) and Other Mycobacterial Species and To Detect Rifampin Resistance in MTBC Isolates following Growth Detection with the BACTEC MGIT 960 System. J. Clin. Microbiol.
41: 2822-2826
[Abstract]
[Full Text]
-
Hasegawa, N., Miura, T., Ishii, K., Yamaguchi, K., Lindner, T. H., Merritt, S., Matthews, J. D., Siddiqi, S. H.
(2002). New Simple and Rapid Test for Culture Confirmation of Mycobacterium tuberculosis Complex: a Multicenter Study. J. Clin. Microbiol.
40: 908-912
[Abstract]
[Full Text]
Top
Abstract
Introduction
