Microscopy: Staining Reactions

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The page below is a sample from the LabCE course Medically Important Aerobic Actinomycetes. Access the complete course and earn ASCLS P.A.C.E.-approved continuing education credits by subscribing online.

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Microscopy: Staining Reactions

Specimens may contain a lot of (inflammatory) polymorphonuclear white blood cells, as well as granular tissue and fibrosis. Expected microbiology smear findings for the aerobic actinomycetes are as follows:
Gram stain
Gram-positive organisms appear blue. Gram-negative organisms appear red.
  • Thin, delicate filamentous, beaded, branching organisms (although Rhodococcus appears coccoid and chaining) are present - additional characteristics of the Gram stain will be reviewed in the section on genera and species.
  • Aerobic actinomycetes stain weakly to strongly gram-positive.
  • On a direct smear, bacteria are usually surrounded by polymorphonuclear white blood cells.
Modified acid-fast stain (MAS) - using a weak acid decolorizer
Using the modified Kinyoun method, modified acid-fast organisms appear pink-red (positive). A negative staining reaction is blue-green. Sometimes, organisms stain weakly positive, or variably (some appear positive, some appear negative).
Limitation: Some of the organisms containing mycolic acid in cell walls do not stain acid-fast if grown on blood agar.14
  • Nocardia species stain weakly acid-fast; Rhodococcus, Gordonia, and Tsukamurella vary upon staining with a modified technique.
  • Streptomyces species stain negative. (However, some spores from Streptomyces hyphae may stain acid-fast.)
  • Note: Mycobacterium stain strongly acid-fast.
14. Leber AL, ed. Clinical microbiology procedures handbook. 4th ed. ASM Press; 2016:Section 6.
15. CDC/Hollinger. Image #21042. Under a magnification of 1200X, this photomicrograph of a lung tissue specimen, which had been harvested from a patient, who had been diagnosed with a case of nocardiosis, revealed some of the histopathologic changes caused by the Gram-positive bacterium, Nocardia asteroides. [Now no longer referred to as asteroides, see recent changes in taxonomy.] PHIL public domain. Created 1968. Accessed October 27, 2022. https://phil.cdc.gov/Details.aspx?pid=21042
16. CDC/Ajello. Image #21346. Under a magnification of 1125X, this photomicrograph depicted numerous, Nocardia asteroides [now no longer referred to as asteroides; see recent changes in taxonomy] bacteria, in this acid-fast-stained specimen. PHIL public domain. Created 1968. Accessed October 27, 2022. https://phil.cdc.gov/Details.aspx?pid=21346

15. Stain of lung tissue revealing gram-positive Nocardia bacteria
16. Specimen depicting Nocardia bacteria on MAS