Establishment and drug susceptibility test of isoniazid resistant Mycobacterium smegmatis

With the emergence of drug resistant tuberculosis, it is very urgent to find novel anti-tuberculosis drugs, especially novel anti-drug-resistant tuberculosis drugs.  Because of the slow growth and the need to work in a biosafty environment of Mycobacterium tuberculosis, the development of evaluation of drug effect is severely impeded.  In order to solve these issues, non-pathogenic fast-growing Mycobacterium smegmatis is introduced as test organism.  The inhA is one of a target of isoniazid (INH) overexpression or mutation of this gene in Mycobacterium tuberculosis conferring resistant to INH.  A recombinant plasmid bearing inhA was constructed and electroporated into Mycobacterium smegmatis, using shuttle expression vector pMV261.  Transformants were induced to express a protein of inhA, identified by SDS-PAGE.  Results show that Mycobacterium smegmatis containing inhA plasmids exhibited 100-fold or greater increased resistance to INH, but it conferred no increased resistance to others first-line anti-tuberculosis drugs.  Resazurin microtiter assay plate testing of Mycobacterium smegmatis susceptibility to drugs is a rapid, simple, and inexpensive method and could decrease color background of drugs by detecting fluorescence.  It will be benefit for high- throughout screening of drugs of anti-isoniazid-resistant Mycobacteria.