Acid fast / Auramine-rhodamine

Stains & molecular markers
Acid fast / Auramine-rhodamine

Author: Nat Pernick, M.D.

Topic Completed: 1 August 2015

Minor changes: 19 January 2021

Copyright: 2003-2021, PathologyOutlines.com, Inc.

PubMed Search: Acid fast[title]

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Table of Contents

Definition / general | Methods | Microscopic (histologic) images | Videos

Cite this page: Pernick N. Acid fast / Auramine-rhodamine. PathologyOutlines.com website. https://www.pathologyoutlines.com/topic/stainsacidfast.html. Accessed May 11th, 2021.

Definition / general

Acid fast refers to microorganisms whose cell wall has a high lipid content of mycolic acids and long chain fatty acids, which traditionally is considered to cause them to bind and retain the complex basic dye carbol-fuchsin even after strong decolorization with acid-alcohol (thus "acid-fast") (Wikipedia)
- Hänscheid et. al. believe auramine O actually binds to nucleic acids, not to the cell wall (see Lancet Infect Dis 2007;7:236, J Microbiol Methods 2008;74:119)
Partially acid fast organisms exhibit both acid fast and non-acid fast bacilli and filaments in a single strain
Standardization recommended for interpretation (Hum Pathol 2012;43:1845)
Sputum smears may misidentify acid-fast bacilli as Mycobacterium tuberculosis in HIV+ patients (J Acquir Immune Defic Syndr 2013;63:168)
Acid fast organisms include Mycobacteria, oocysts of Cryptosporidium parvum, Cyclospora, Isospora; also hooklets of cysticerci
Partially acid fast organisms include Actinomyces: Dietzia (Int J Syst Evol Microbiol 2006;56:1667), Gordonia (Emerg Infect Dis 2000;6:382), Nocardiae (Surg Infect (Larchmt) 2012;13:163), Rhodococcus (South Med J 1991;84:1217), Tsukamurella (J Med Case Rep 2008;2:207); also rarely Mycobacterium peregrinum (J Clin Microbiol 2005;43:2015)
Note: nucleic acid based tests can rapidly detect and speciate mycobacteria (Arch Pathol Lab Med 2008;132:1333, Thorax 2008;63:317)

Methods

Ziehl-Neelsen (classic): common method; bacteria stain bright red due to retention of carbol-fuchsin dye; background is methylene blue counterstain; procedure involves heat (#1, #2)

Ziehl-Neelsen (modified bleach): may be more sensitive than classic stain (Acta Cytol 2008;52:325,J Cytol 2012;29:165)

Ziehl-Neelsen (modified for stool specimens): does not require heating (Centers for Disease Control)

Kinyoun: common method; uses more concentrated fuchsin dye and lipid solvent, but no heat; bacteria stain bright red against green background (#1, #2)

Fite: to detect M. leprae (leprosy) and Rhodococcus (Diagn Cytopathol 2001;24:244); combines peanut / vegetable oil with xylene to minimize exposure of bacteria cell wall to organic solvents and protect precarious acid-fastness of organism (#1, #2)

Ellis and Zabrowarny: protocol excludes phenol; procedure (J Clin Pathol 1993;46:559)

Auramine-rhodamine: mixture of Auramine O and Rhodamine B dyes, auramine binds to mycolic acid in cell wall; detection requires a fluorescence microscope (mercury vapor lamp or LED), but is most sensitive stain for mycobacteria (Hum Pathol 1984;15:1085, PLoS One 2011;6:e22495); saves time in searching for microorganisms (Clin Infect Dis 2008;47:203); procedure

Water filters are recommended to reduce false positives due to non-TB mycobacteria (Appl Environ Microbiol 2007;73:6296)

Microscopic (histologic) images

Images hosted on other servers:

Cryptosporidium: