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Reproduction of Clinical Isolated Rifampicin-dependent L-form Multidrug-resistant Tuberculosis (MDR-TB)

Jialing Bao, Yiwei Wang, Xiyan Zhang and Min Zhong

The emergence of multi-drug resistant-Mycobacterium tuberculosis (MDR-TB), especially the Rifampicin-dependent (R-) MDR-TB, has become a hot issue. To elucidate the potential mechanism that rifampicin-dependent MDR-TB (latent infection) utilizes for its long-term survival, we studied the morphology and L-form growth pattern of rifampicin-dependent MDR-TB. Rifampicin-dependent MDR-TB was isolated from fresh sputum of patients and showed favorable growth in the Rifampicin-containing conditions. Conversely, when R-MDR-TB was cultured in Rifampicin-depleted medium, bacterial growth ceased and MDR-TB transformed into L-form cells. This transformation process was studied over a span of three weeks by both optical microscope and transmission electron microscopy (TEM). At week one, the R-dependent bacteria cultured under rifampicin-null conditions exhibited acid-fast positive fried egg colonies and mycoplasma cell-like morphology. During week 2, the exhibited morphologies of the R-dependent MDR-TB were: acid-fast positive filamentous, granular sphere, cell membrane sugar-coat (filamentous particles aggregations) and protoplasms with various sizes and shapes. The morphologies of R-MDR-TB at week 3 were: tiny translucent colonies and giant sphere, filaments, protoplasm, fried egg colonies and "mycoplasma (filopodia)". These morphologies are typical and representative characteristics of L-form cells growth. These findings reveal the L-form growth of rifampicin-dependent MDR-TB for the first time, and elucidate the potential mechanism of rifampicin-dependent MDR-TB longterm survival (latent infection). This data will help to provide a foundation for novel early diagnosis and effective treatment of R- MDR-TB.