title: Non-exponential growth of Mycobacterium leprae Thai-53 strain cultured in vitro.
abstract: In this study, attempts were made to culture this bacterium in media supplemented with a variety of biological materials to determine why cultivation of Mycobacterium leprae in vitro has not this far been successful. A slight increase in the number of cells in medium supplemented with human blood plasma and an extract of nude mouse tissue as observed after more than 3 months of cultivation at 30 °C. To ascertain whether this increase was real growth, the growth was analyzed by droplet digital PCR, which showed a slow increase in the copy number of cell-associated DNA and the release of a large amount of DNA into the culture medium from bacterial cells during cultivation. These results were supported by electron microscopic examination of M. leprae in infected mouse tissues, which showed that most of the replicated bacteria had degenerated and only a few cells survived. Based on these results, it was postulated that many of the replicated cells degenerate during M. leprae growth and that only a few cells remain to participate in the next growth stage. This means that, unlike other cultivable bacteria, the growth of M. leprae is not exponential and the number of cells therefore increase extremely slowly. Thus, accurate judging of the success of M. leprae cultivation requires observation of growth over a long period of time and careful measurement of the increase in number of viable cells.
authors.affiliation: Department of Bacteriology, Faculty of Medical Sciences, Kyushu University, 3-1-1 Maidashi Higashi-ku Fukuoka 812-8582, Japan.
authors.forename: Kazunobu
authors.initials: K
authors.lastname: Amako
authors.affiliation: Department of Bacteriology, Faculty of Medical Sciences, Kyushu University, 3-1-1 Maidashi Higashi-ku Fukuoka 812-8582, Japan.
authors.forename: Ken-Ichiro
authors.initials: KI
authors.lastname: Iida
authors.affiliation: Department of Bacteriology, Faculty of Medical Sciences, Kyushu University, 3-1-1 Maidashi Higashi-ku Fukuoka 812-8582, Japan.
authors.forename: Mitsumasa
authors.initials: M
authors.lastname: Saito
authors.affiliation: Department of Bacteriology, Faculty of Medical Sciences, Kyushu University, 3-1-1 Maidashi Higashi-ku Fukuoka 812-8582, Japan.
authors.forename: Yoshitoshi
authors.initials: Y
authors.lastname: Ogura
authors.affiliation: Department of Bacteriology, Faculty of Medical Sciences, Kyushu University, 3-1-1 Maidashi Higashi-ku Fukuoka 812-8582, Japan.
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Hayashi
authors.affiliation: Department of Bacteriology, Faculty of Medical Sciences, Kyushu University, 3-1-1 Maidashi Higashi-ku Fukuoka 812-8582, Japan.
authors.forename: Shin-Ichi
authors.initials: SI
authors.lastname: Yoshida
mesh_terms: D000818:Animals; D001431:Bacteriological Techniques / Q000379:methods*; D001769:Blood / Q000378:metabolism; D003470:Culture Media / Q000737:chemistry*; D004269:DNA, Bacterial / Q000032:analysis; D006801:Humans; D008819:Mice, Nude; D050296:Microbial Viability; D008854:Microscopy, Electron; D009166:Mycobacterium leprae / Q000254:growth & development* / Q000502:physiology / Q000648:ultrastructure; D013696:Temperature; D014020:Tissue Extracts / Q000378:metabolism
chemical_list: D003470:Culture Media; D004269:DNA, Bacterial; D014020:Tissue Extracts
keywords: cultivation; droplet digital PCR; growth in vitro