Quality assurance for molecular epidemiology of tuberculosis methods in the mycobacterium reference laboratory |
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Authors: | Paul J Freidlin Drora Goldblatt Hasia Kaidar-Shwartz Zeev Dveyrin Efrat Rorman |
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Institution: | (1) National Mycobacterium Reference Laboratory, National Public Health Laboratory, Ministry of Health, Tel Aviv, Israel;(2) National Public Health Laboratory, Ministry of Health, Tel Aviv, Israel |
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Abstract: | In the National Mycobacterium Reference Laboratory of the Israeli National Public Health Laboratory (hereafter referred to
as “the laboratory”), three methods are employed for the molecular epidemiology of Mycobacterium tuberculosis: IS6110 restriction fragment length polymorphism typing (RFLP typing), 43 spacer oligonucleotide typing (spoligotyping),
and 24 loci Mycobacterial Interspersed Repeating Unit—Variable Number of Tandem Repeats typing (MIRU-VNTR typing). In this
article, we describe the main practical aspects concerning quality assurance of these methods that are based on standardized,
internationally agreed upon conditions, including consensus reference strains and markers. All three methods were validated
by classical epidemiology (confirmed transmission) and clinical information. The laboratory has introduced a new 5 colors,
4 primer sets multiplex modification of the optimal 24-miru typing system that includes an easily produced in-house internal
standard for the high-throughput capillary electrophoresis system. Quantitative measurement of the internal standards yielded
statistics for measurement uncertainty that include the frequency distribution, mean, standard deviation, 95% confidence interval
and coefficient of variation. Use of the new internal standard developed in our laboratory allowed us to introduce the first
quantitative evaluation of the system performance of the AB3130xl capillary electrophoresis genetic analyzer for MIRU-VNTR
typing. The results are discussed in terms of expected accuracy and precision of MIRU-VNTR results, and possible implications
for forensic microsatellite typing which may be much more sensitive to the observed intra- and inter-plate variation. |
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