Enhanced methodologies for detecting phenotypic resistance in mycobacteria
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Lipid droplets found in algae and other microscopic organisms have become of interest to many researchers partially because they carry the capacity to produce bio-oil for the mass market. They are of importance in biology and clinical practice because their presence can be a phenotypic marker of an altered metabolism, including reversible resistance to antibiotics, prompting intense research.A useful stain for detecting lipid bodies in the lab is Nile red. It is a dye that exhibits solvatochromism; its absorption band varies in spectral position, shape and intensity with the nature of its solvent environment, it will fluoresce intensely red in polar environment and blue shift with the changing polarity of its solvent. This makes it ideal for the detection of lipid bodies within Mycobacterium spp. This is because mycobacterial lipid bodies' primary constituents are nonpolar lipids such as triacylglycerols but bacterial cell membranes are primarily polar lipid species. In this chapter we describe an optimal method for using Nile red to distinguish lipid containing (Lipid rich or LR cells) from those without lipid bodies (Lipid Poor or LP). As part of the process we have optimized a method for separating LP and LR cells that does not require the use of an ultracentrifuge or complex separation media. We believe that these methods will facilitate further research in these enigmatic, transient and important cell states.
Hammond , R J H , Baron , V O , Lipworth , S & Gillespie , S H 2018 , Enhanced methodologies for detecting phenotypic resistance in mycobacteria . in S Gillespie (ed.) , Antibiotic Resistance Protocols . Methods in Molecular Biology , vol. 1736 , Humana Press/Springer , New York, NY , pp. 85-94 . https://doi.org/10.1007/978-1-4939-7638-6_8
Antibiotic Resistance Protocols
© 2018, Springer Science+Business Media, LLC. This work has been made available online by kind permission of the publisher. This is the author created, accepted version manuscript following peer review and may differ slightly from the final published version. The final published version of this work is available at http://doir.org/10.1007/978-1-4939-7638-6_8
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