Evolution and role of corded cell aggregation in Mycobacterium tuberculosis cultures

• Published in: Tuberculosis (Edinburgh, Scotland) Vol. 93; no. 6; pp. 690 - 698
• Main Authors: Caceres, Neus, Vilaplana, Cristina, Prats, Clara, Marzo, Elena, Llopis, Isaac, Valls, Joaquim, Lopez, Daniel, Cardona, Pere-Joan
• Format: Journal Article
• Language: English
• Published: Scotland Elsevier Ltd 01.11.2013
• Subjects: Biofilms; Cell Aggregation - physiology; Colony Count, Microbial; Cord Factors - physiology; Corded cell aggregation; Cording; Culture growth characteristics; Freezing; Image Processing, Computer-Assisted - methods; Infectious Disease; Microscopy, Electron, Scanning; Mycobacterium tuberculosis; Mycobacterium tuberculosis - drug effects; Mycobacterium tuberculosis - growth & development; Mycobacterium tuberculosis - physiology; Mycobacterium tuberculosis - ultrastructure; Polysorbates - pharmacology; Pulmonary/Respiratory; Stress, Physiological - physiology; Culture growth characteristics; Mycobacterium tuberculosis; Cording; Corded cell aggregation
• ISSN: 1472-9792; 1873-281X
• DOI: 10.1016/j.tube.2013.08.003

Summary The aim of this study was to evaluate the evolution and role of corded cell aggregation in Mycobacterium tuberculosis cultures according to growth time and conditions. Thus, in standard culture using aerated 7H9 Middlebrook broth supplemented with 0.05% Tween 80, a dramatic CFU decrease was observed at the end of the exponential phase. This phase was followed by a stable stationary phase that led to dissociation between the optical density (O.D.) and CFU values, together with the formation of opaque colonies in solid culture. Further analysis revealed that this was due to cording. Scanning electron microscopy showed that cording led to the formation of very stable coiled structures and corded cell aggregations which proved impossible to disrupt by any of the physical means tested. Modulation of cording with a high but non-toxic concentration of Tween 80 led to a slower growth rate, avoidance of a sudden drop-off to the stationary phase, the formation of weaker cording structures and the absence of opaque colonies, together with a lower survival at later time-points. An innovative automated image analysis technique has been devised to characterize the cording process. This analysis has led to important practical consequences for the elaboration of M. tuberculosis inocula and suggests the importance of biofilm formation in survival of the bacilli in the extracellular milieu.