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<p>Tuberculosis (TB) still threatens human life despite the availability of childhood vaccination and modern treatment regimens due to the emergence of tuberculosis with extended resistance profiles. The mouse active TB model remains the gold standard for evaluating anti-TB drugs and vaccines. However, <i>in vivo</i> experiments raise ethical concerns and are time-consuming. We established an Akaluc-based bioluminescence platform to enable rapid drug screening in culture media and THP‑1 cells. Codon-optimized Akaluc expression in mycobacteria was accomplished by assortment of vectors and promoters. Bioluminescence kinetics were evaluated in culture media and THP‑1 cells with or without drug treatment, and optimized by adjusting time points and substrate concentrations. TokeOni at a concentration of 10&#xa0;nM/100&#xa0;µL produced the highest bioluminescence compared to other tested concentrations and substrates. Among the tested promoter-plasmid constructs, the Ag85B promoter in pMV261 generated the strongest bioluminescence in <i>Mycobacterium smegmatis</i> and <i>Mycobacterium bovis</i>.&#xa0;Bioluminescence fluctuated with bacterial growth, peaking during the log phase and gradually declining during the stationary phase. A positive correlation was observed between bioluminescence and CFU reduction <i>in vitro</i> upon treatment with sensitive drugs.</p>

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Real-time bioluminescence imaging of mycobacteria with Akaluc: a novel method for monitoring drug efficacy

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年5月15日

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  • Md Shafiul Islam,
  • Atsuki Takeishi,
  • Yoshitaka Tateishi,
  • Akihito Nishiyama,
  • Yuriko Ozeki,
  • Yutaka Yoshida,
  • Amina Kaboso Shaban,
  • Takeshi Annoura,
  • Satoshi Iwano,
  • Takasuke Fukuhara,
  • Sohkichi Matsumoto

摘要

Tuberculosis (TB) still threatens human life despite the availability of childhood vaccination and modern treatment regimens due to the emergence of tuberculosis with extended resistance profiles. The mouse active TB model remains the gold standard for evaluating anti-TB drugs and vaccines. However, in vivo experiments raise ethical concerns and are time-consuming. We established an Akaluc-based bioluminescence platform to enable rapid drug screening in culture media and THP‑1 cells. Codon-optimized Akaluc expression in mycobacteria was accomplished by assortment of vectors and promoters. Bioluminescence kinetics were evaluated in culture media and THP‑1 cells with or without drug treatment, and optimized by adjusting time points and substrate concentrations. TokeOni at a concentration of 10 nM/100 µL produced the highest bioluminescence compared to other tested concentrations and substrates. Among the tested promoter-plasmid constructs, the Ag85B promoter in pMV261 generated the strongest bioluminescence in Mycobacterium smegmatis and Mycobacterium bovis. Bioluminescence fluctuated with bacterial growth, peaking during the log phase and gradually declining during the stationary phase. A positive correlation was observed between bioluminescence and CFU reduction in vitro upon treatment with sensitive drugs.