The direct fluorescent antibody (DFA) test is a rapid microscopic procedure used to identify Legionella pneumophila and other Legionella species. Antibodies specific for Legionella species and serogroups are tagged with a fluorescent dye.  When Legionella is isolated by culture from an environmental source or a patient, the bacteria are placed on a slide and exposed to a panel of DFA reagents. A positive reaction confirms the identification. DFA is sensitive and highly specific when used for species/serotype confirmation.  

DFA staining is not recommended as a method to be applied directly on an environmental sample due to false positive and false negative results. DFA staining will stain both viable (living) and non-viable (dead) Legionella cells.

There are more than 50 Legionella species and 16 serogroups of L. pneumophila, however only a select few species and serogroups are responsible for most illness. Therefore DFA testing is performed to identify or rule out only the most clinically relevant strains.  If requested or when species identification is required such as in investigations of clinical cases, DNA sequencing can be performed for identification of unusual species or those not included in the DFA panels.

Test Method

  • SPL. M12.00  Legionella Identification by Fluorescent Antibody Tests
  • Confirmation using species/serogroup specific reagents:
    Legionella pneumophila serogroups 1-6 and 8  
    Blue white fluorescing Legionella species (L. anisa, L.bozemanii, L. dumoffii, and L. gormanii)  L. pneumophila serogroups not 1-6: confirmed using a pooled reagent of 14 L. pneumophila serogroups. 
  • Legionella species, not pneumophila, are confirmed using a pooled reagent that is specific for 19 Legionella species and serogroups other than L.pneumophila. These include:  L.anisa, L.bozemanii serogroups 1 and 2, L.gormanii, L.longbeacheae serogroups 1 and 2, L.dumoffii, L.micdadei, L.wadsworthii, L.oakridgensis, L.feeleii serogroups 1 and 2, L.sainthelensi, L.jordanis,  L.hackeliae serogroups 1 and 2, L.maceacherniiL.santicrucis.
  • If requested, other non-pneumophila species can be confirmed by DFA and include: L.spiritensis, L.jamestowniensis, L.cherri, L.steigerwaltii, L.rubrilucens /erthra.

Turn Around Time

2–3 days

Test Code



Submit isolate on a slant or agar plate of Buffered Charcoal Yeast Extract (BCYE) in double containment packaging according to IATA regulations for shipping Biological Substances, Category B (UN3373).  Contact SPL if you have questions. 


  1. Procedures for the Recovery of Legionella from the Environment. Centers for Disease Control publication. January 2005.  
  2. DFA manufacturers’ instructions: Remel, Lenexa, KS;  Bio-Rad, Redmond, WA and m-Technologies, Alpharetta, Ga.