European Society of Mycobacteriology - Instituto Nacional de Saúde ...

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GENOTYPIC DETECTION OF ISONIAZID AND RIFAMPIN
RESISTANCE IN Mycobacterium tuberculosis CLINICAL ISOLATES
Maitane Aranzamendi Zaldumbide, Carlos Fernandez Mazarrasa , Luis Martinez-Martinez, Jesus Agüero Balbin.
Hospital Universitario Marques de Valdecilla, Servicio de Microbiologia, Santander, Spain.
Background
The emergence of Mycobacterium tuberculosis resistant to first-line drugs underlines the urgent need for new resistanceprofiling
methods that would allow for timely determination of proper treatment. The aim of this study was to evaluate
the development of targeted and fast molecular diagnostic method suitable for specific genome regions responsible for
isoniazid (INH) and rifampin (RAMP) resistance in M. tuberculosis clinical isolates.
Methods
79 strains known to be resistant to INH (n=71) or INH+RAMP (n=8) by the automated system BacT ALERT (bioMérieux)
were selected from a stock collection of clinical isolates (January 2000- March 2009). The genome regions associated
with INH-R (including the codon 315 of the katG gene and the fabG1(mabA)-inhA regulatory region) and RAMP-R (81-bp
hot spot region of the rpoB gene called RRDR) were amplified by PCR and the DNA sequences were studied.
Results
Of the 79 isolates, 22 (27.84%) had the mutation S315T in the katG gene, 5 (6.32%) showed changes at -15 nucleotide of
the fabG-inhA regulatory region and 2 (2.53%) presented both mutations. A significant proportion of strains, 50 (63.29%), had
no detectable alterations at the studied loci. INH + RAMP-R strains were associated with mutations in the RRDR of the rpoB
gene in all cases. From these, majority (5 of 8, 62.5%) presented the mutation S531L, whereas the others involved changes
at the codon 516: mutations D516V and D516F, which were identified in 1 (12.5%) and in 2 (25%) strains respectively.
Conclusions
Our results demonstrated a low sensitivity of this method to detect INH-R strains, and points to the need of finding
out other mutant regions. On the other side, confirm the usefulness of this strategy for fast assessment of resistance to
RAMP, which in turn is a marker for multiresistance. This analysis can also be done with assays based on reverse line blot
hybridization detecting the same mutations, except D516F, which is not targeted.
European Society of Mycobacteriology | 30 th Annual Congress | July 2009 | Porto - Portugal
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