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Cording, a virulence-related characteristic of Mycobacteria , analysis using Cord formation in Mycobacterium tuberculosis has been traditionally related to These structures consist of bacilli aligned in parallel, end to end, and side to side, along the long axis of the cord.
However, similar structures were recently found in non-pathogenic and in opportunistic Mycobacterium species such as M. Nevertheless, the observation of these formations at high resolution is crucial for two essential applications. Firstly, to identify the unknown mycobacterial components responsible for cording formation, an important step to understand virulence.
Secondly, to demonstrate that cording is not specific of M. Thus, patients would be exposed to an ineffective drug regimen with many adverse side effects. Here, we focused on sample treatment in order to certainly observe cords by using scanning electron microscopy SEM. The fast method of impregnation with osmium vapours was compared with the conventional method of sample treatment chemical fixation, dehydration and critical-point drying.
Both techniques require the use of spreading pellicles formed by mycobacteria grown on liquid medium, to ensure enough material to visualize this fragile structure. Osmium impregnation is an interesting method for a first approach to visualize more native structures, since mycobacteria are not immersed in liquid solutions and architectural details are preserved.
As a disadvantage, bacilli are covered by an extracellular matrix making their observation difficult Figure 1. Using the conventional method, where highly hydrophobic mycobacteria are immersed in several aqueous solutions and thereafter in alcoholic solutions, the extracellular matrix is mostly lost and bacilli are easily observed forming cords Figures 1, 2. By using the latter method we have demonstrated the formation of cords in rough M.
Mycobacterium abscessus is an opportunistic emergent pathogen that causes lung infections and it is possible to misidentify it as M. Thus, these findings imply crucial information for clinical microbiology routine protocols since the observation of cords by light microscopy still remain as the unique method for tuberculosis diagnosis in wide developing areas.
We concluded that it is necessary to stress on particular protocols to observe mycobacterial cords at ultrastructural level. Moreover, observation at ultrastructural level with SEM is the most effective method to clearly visualize cords in Mycobacterium. References 1.
Hall-Stoodley et al. Clement et al. Williams et al. Representative micrographs for comparing conventional treatment B and osmium fixation A in three non-tuberculous Mycobacterium species: M. Bar sizes, medium magnification large images 7. Figure 2. Representative micrographs for comparing rough and smooth strains of M.
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