Pasteur Institute of Madagascar
Standard Diagnostics
Mycobacterium Unity
Intermediary Report (11/07/2008)
Evaluation of the SD TB Ag MPT64 kit
I. Context
The biological, immunological and molecular characterizations of Mycobacterium Tuberculosis lead to the identification of different antigens useful for the elaboration of better diagnosis methods in order to make the distinction between tuberculosis complex, responsible for tuberculosis, and other mycobacterium not belonging to this complex.
MPT64 is one of the predominant proteins secreted by Mycobacterium Tuberculosis. This protein is secreted during active growing of strains of tuberculosis complex. It is an immunogenic protein of 24 kDa specific to tuberculosis complex. This antigen has been suggested as potential candidate of a vaccine composition because it is preferentially recognized by Tcells CD4+.
II. Problem of the research and objectives
Recently, Standard Diagnostics, Inc. has developed an easy and rapid method using mouse monoclonal antibodies anti-MPT64. The kit can be easily used in laboratories for a rapid identification of tuberculosis bacilli from solid or liquid culture.
The principal objective of this study is to evaluate the ‘SD TB Ag MPT64 Rapid test’ for identification of Mycobacterium of tuberculosis complex.
More specifically, this study is done to;
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evaluate performances of the test for identification of tuberculosis bacillus isolates from solid culture (Lowenstein Jensen medium)
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Compare results of the new test with results of classical tests for identification of mycobacterium (biochemical tests) used in laboratory.
III. Methodological and experimental approach
The study was done on strains of the Laboratory of Mycobacterium of the Pasteur Institute of Madagascar. These strains contained mycobacterium of tuberculosis complex (M. Tuberculosis and M. Bovis) and other atypical mycobacterium. Other bacterial species were tested to evaluate the specificity of the new test. These species were collected from the Clinical Biology Center of the Pasteur Institute of Madagascar.
A first evaluation is done on 81 strains (all species mixed). The second step of the evaluation will be done with other strains according to the number of tests available (88 tests).
For the first step, strains of M. Tuberculosis , M. Bovis , atypical mycobacterium and other non-mycobacterium pathogen strains have been selected to be tested with the kit:
For each strain, SD Ag MPT64 has been tested in parallel with specific identification of each strain.
The results of the identification of mycobacterium of tuberculosis complex were compared to biochemical identification results.
During manipulations, the reference strain H37 Rv (mycobacterium laboratory-IPM) was used as positive sample and distilled water as negative sample.
Table: Repartition of strains used
Strain
Number
M. Tuberculosis
36
M. Bovis
12
Atypical Mycobacterium
13
other non-mycobacterium pathogen strains
20
IV. Results
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From 02 to 09 July 2008, the kit has been evaluated on:
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36 strains of M. Tuberculosis
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12 strains of M. Bovis
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13 strains of Atypical Mycobacterium
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20 strains of other non-mycobacterium pathogen strains
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Biochemical identification
The identification of the mycobacterium strains is done following the algorithm of the Mycobacterium Laboratory based on an assembly of morphologic, cultural and biochemical criteria. The specific identification of the other pathogen bacteria is done following the algorithm of the Biological Institute of Madagascar.
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Identification by SD Ag TB MPT64
For each strain a sample of the colony on the solid culture is taken and diluted in an Eppendorf containing 200 ?l of extraction buffer included in the kit. After strong vortexing, 100 ?l of this mix is taken and add on the device. If two lines (Test (T) and Control (C)) in the window, the result is positive. If one single line appears on the Control line, the test is negative. The results have been interpreted in double-blind.
The results obtained with SD TB Ag MPT64 were concordant with the results of the biochemical identification. All the samples of the non tuberculosis complex gave a negative result with the kit.
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Result analysis
In total 48 strains of tuberculosis complex and 33 non-tuberculosis strains have been tested by the SD Ag MPT64 kit.
Sensibility = 48 / (48+0) = 1 ie 100%
Specificity = 33/33 = 1 ie 100%
PPV=48 / (48+0) = 1 ie 100%
NPV= 33/(3+0)3 = 1 ie 100%
Remark: Two strains in collection identified as M. Bovis (01-3236) and M. mucogenicum gave respectively a negative and positive result with TB Ag MPT64 test. The parallel biochemical identification of these strains showed that the first strain was an atypical mycobacterium and the second one was a M.Tuberculosis strain.
Conclusion
According to the results of this first evaluation, SD TB Ag MPT64 presents a good sensibility (100%) and a good specificity (100%) to identify mycobacterium of the tuberculosis complex (M.Tuberculosis and M. Bovis). The evaluation of the kit will be continued on a prospective manner in the Laboratory of Mycobacterium in parallel with biochemical identification of the strains.
September 17th, 2009 at 10:57 am
This kit sounds amazing