In attempt to develop an in vitro assay system of antileprosy drug, fusidic acid and
rifampin, we have applied DNA and RT PCR for the detection of M. leprae infection
into murine peritoneal macrophages. The evalutation of efficacy of antileprosy drugs and
follow-up of MDT were hindered by lack of molecular biological methods to substitute
AFB stain, which stains lipid coat of M. leprae could not judge the viability and status
of M. leprae proteins, DNA and RNA.
Fusidic acid, known as an inhibitor of protein translocation, and rifampin, the most
microbicidal agent of MDT components and inhibitor of DNA-dependent RNA
polymerase of M. leprae, were injected into M. leprae-infected murine macrophages.
Freezing and thawing method reveals most effective way of DNA separation from M.
leprae-infected macrophages. Infectivity of M. leprae was confirmed by AFB staining.
In spite of successful DNA and RT PCR for the 18 kDa gene of M. leprae, We could
not find any difference of bands in agarose gel depending on the concentration and
duration of treatments of fusidic acid and rifampin against M. leprae-infected
macrophages in vitro. It means that in vitro assays of antileprosy drugs by DNA and
RT PCR are so sensitive to detect any bactericidal effects on very slow growing M.
leprae. At present, in vitro assay system using DNA and RT PCR appears not eligible
to substitute present expensive methods such as [14C-palmitic acid]
respirometry with Buddemeyer system or BACTEC 460. It needs further study to settle
down as an in vitro assay system to follow up the efficacy of antileprosy drugs such as
fusidic acid and rifampin.
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