Chapter 23 : Acid-Fast Positive and Acid-Fast Negative : The Koch Paradox

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possesses a unique cell wall architecture that is distinct from both Gram-negative and Gram-positive bacteria. The cell wall consists of a thick, lipid-rich outer layer composed primarily of mycolic acids ( ) ( Fig. 1 ). This lipid layer lies on top of a layer of peptidoglycan and the polysaccharide arabinogalactan, which, in turn, are anchored to the inner lipid membrane common to all bacteria ( ). The overall thick waxy coat renders acid-fast (AF) mycobacteria resistant to Gram staining. When stained with alternative dyes, the cell wall is resistant to decolorization with acid alcohol, thus giving these bacteria their sobriquet “acid-fast.” This unique AF property has been the basis for the continuous development of staining procedures over the past century and remains the cornerstone for the diagnosis of tuberculosis (TB), especially in low-income and middle-income countries where more than 90% of TB cases occur ( ). The Ziehl-Neelsen (ZN) stain, also known as the AF stain, which is used in microscopic detection of , was originally developed independently by Ziehl and Neelsen, who improved on the early work of Koch, Rindfleisch, and Ehrlich (see below).

Citation: Vilchèze C, Kremer L. 2017. Acid-Fast Positive and Acid-Fast Negative : The Koch Paradox, p 519-532. In Jacobs, Jr. W, McShane H, Mizrahi V, Orme I (ed), Tuberculosis and the Tubercle Bacillus, Second Edition. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.TBTB2-0003-2015
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Figure 1

Chemical structures of the major mycolic acids of . Cyclopropane rings and methyl branches are shown and annotated with the -adenosyl methionine-dependent methyl transferases responsible for their synthesis. , ; , .

Citation: Vilchèze C, Kremer L. 2017. Acid-Fast Positive and Acid-Fast Negative : The Koch Paradox, p 519-532. In Jacobs, Jr. W, McShane H, Mizrahi V, Orme I (ed), Tuberculosis and the Tubercle Bacillus, Second Edition. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.TBTB2-0003-2015
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Figure 2

Staining of using ZN (left) and auramine O (right). Magnification, ×100.

Citation: Vilchèze C, Kremer L. 2017. Acid-Fast Positive and Acid-Fast Negative : The Koch Paradox, p 519-532. In Jacobs, Jr. W, McShane H, Mizrahi V, Orme I (ed), Tuberculosis and the Tubercle Bacillus, Second Edition. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.TBTB2-0003-2015
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Figure 3

Ser/Thr kinase-dependent signaling cascade resulting in phosphorylation of KasB and loss of acid-fastness. Modification of the cell wall composition in response to exogenous cues is central for adaptation to different environmental conditions. In response to an external signal, mycobacterial Ser/Thr kinases phosphorylate the different FAS-II components, including the β-ketoacyl ACP synthase KasB involved in the addition of the last carbon atoms during the mycolic acid elongation step. Phosphorylation on Thr334 and Thr336 decreases the condensation activity of KasB, resulting in the production of shorter mycolic acids, which probably affects the packing of the lipid layer and also results in the loss of the AF property and severe attenuation in mice.

Citation: Vilchèze C, Kremer L. 2017. Acid-Fast Positive and Acid-Fast Negative : The Koch Paradox, p 519-532. In Jacobs, Jr. W, McShane H, Mizrahi V, Orme I (ed), Tuberculosis and the Tubercle Bacillus, Second Edition. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.TBTB2-0003-2015
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Figure 4

Loss of AF staining coincides with the accumulation of TAG-containing intracellular lipid inclusions. Dual staining of grown under multiple stress conditions, using auramine O for AF-staining (green) and Nile red as a neutral lipid stain (red). Bacilli were observed by confocal laser scanning microscopy. Overlaid images of the dual-stained bacteria are shown. Bar = 4 μm. Quantification of the number of AF-positive and lipid-stain-positive bacilli grown as in (A). Auramine O-stained and Nile red-stained positive cells were counted from multiple scans. (Adapted from Deb et al. 4(6):e6077 with permission of the publisher.)

Citation: Vilchèze C, Kremer L. 2017. Acid-Fast Positive and Acid-Fast Negative : The Koch Paradox, p 519-532. In Jacobs, Jr. W, McShane H, Mizrahi V, Orme I (ed), Tuberculosis and the Tubercle Bacillus, Second Edition. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.TBTB2-0003-2015
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