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Legionella Agar Enrichment is intended for use in the the preparation of Legionella Agar. The complete medium is based on the Charcoal Yeast Extract (CYE) Agar formula of Feeley et al.,¹ as modified by Edelstein,² and is recommended for use in the isolation and cultivation of Legionella from clinical and non-clinical materials.

Legionella pneumophila was first demonstrated as a causative agent of pneumonia by McDade et al., who isolated this organism from lung tissue of patients who died of Legionnaires Disease.³ Since that time, six additional species of Legionella have been described and have been shown to be either causative agents of pneumonia,^4-7 or to be associated with pneumonia patients by serological procedures.^8,9

Legionellae have been frequently isolated from environmental aquatic sources including air conditioning evaporation condensers,¹⁰ cooling towers¹¹ and potable water sources such as shower heads¹² or water fixtures.¹³ Presumably, these and other aquatic sources serve as reservoirs for this group of organisms and represent the means by which legionellosis is acquired.

Initial isolation of L. pneumophila was performed by McDade et al., using guinea pigs and embryonated chicken eggs.³ In 1978, Weaver reported that Mueller Hinton Chocolate Agar would support good growth of this organism.¹⁴ Feeley et al., subsequently determined that ferric pyrophosphate and L-Cysteine would respectively replace the hemoglobin and enrichment components of Mueller Hinton Chocolate Agar.¹⁴ Feeley et al., also showed that growth of L. pneumophila was optimal at pH 6.9. These observations were incorporated into an improved legionella agar, which they called F-G Agar.

In 1979, Feeley et al., described a modification of F-G Agar.¹ This medium, which they called Charcoal Yeast Extract (CYE) Agar was found to provide better growth of Legionella than obtained on F-G Agar. Subsequently, Pasculle et al., reported that the performance of CYE Agar could be further improved by the addition of ACES buffer.¹⁵ Edelstein described a further modification of CYE Agar in which he incorporated both ACES buffer and alpha ketoglutarate.² Edelstein reported that this further modification, which he referred to as BCYEα Agar, improved the growth and recovery of Legionella. Legionella Agar is a modification of the BCYEα Agar formula of Edelstein. In the formula, the concentration of ACES buffer was reduced from 10.0 g/L to 6.0 g/L.

1. Examine the lyophilized and rehydrated supplement for evidence of deterioration as described under "Product Deterioration".

2. Check the performance of the enrichment by testing in the complete medium. Plates should be inoculated with approximately 100 - 200 colony-forming units of the test cultures and incubated aerobically under humidified conditions at 35°C. Examine plates for growth after 48 and 72 h incubation. Results should be as stated below:

For Laboratory Use.

Use aseptic technique in rehydrating Legionella Agar Enrichment and adding the supplement to the base. Follow proper, established laboratory procedures in handling and disposing of infectious materials. SInce legionellosis is apparently acquired by the airborne route, care should be taken to minimize aerosis. Use of a biological safety cabinet in the handling and inoculation of specimens suspected to contain Legionella organisms and in the handling and examination of inoculated plates is recommended. Work surfaces should be disinfected with 5% phenol or 5% hypochlorite solutions.