Article

Exhibition of persistent and drug-tolerant L-form habit of Mycobacterium tuberculosis during infection in rats

Central European Journal of Biology (Impact Factor: 0.82). 3(4):407-416. DOI: 10.2478/s11535-008-0032-7

ABSTRACT A model for studying mycobacterial L-form formation in vivo was established to demonstrate the ability of M. tuberculosis to behave as a drug-tolerant L-form persister. Rats were infected by intranasal (i.n.) and intraperitoneal (i.p.) routes
with 1×108 cells/ml of M. tuberculosis. At weekly intervals during a period of five weeks, samples from lung, spleen, liver, kidney, mesenterial and inguinal lymph
nodes, broncho-alveolar and peritoneal lavage liquid were plated simultaneously on Löwenstein-Jensen (LJ) medium or inoculated
into specially supplemented for L-forms Dubos broth (drug-free and drug-containing variants). The use of liquid media enabled
isolation of mycobacterial L-form cultures during the whole period of experiment including the last two weeks, when tubercle
bacilli were not isolated on LJ medium. An unique feature of mycobacterial L-forms was their ability to grow faster than the
classical tubercle bacilli. Isolation and growth of L-form cultures in primary drug-containing media demonstrated their drug-tolerant
properties. Electron microscopy of liquid media isolates showed that they consisted of morphologically heterogenous populations
of membrane-bound and of variable sized L-bodies that completely lack cell walls. The identity of the isolated non-acid fast
and morphologically modified L-forms as M. tuberculosis was verified by specific spoligotyping test. The results contribute to special aspects concerning the importance of mycobacterial
L-form phenomenon for persistence and latency in tuberculosis, phenotypic drug tolerance, as well as for diagnosis of difficult
to identify morphologically changed tubercle bacilli which are often mistaken for contaminants.

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