A PCR-METHOD FOR THE DETECTION OF THE ASS OCIATION BETWEEN LEGIONELLASP AND AMEBA SP

With two pairs of primers for the amplification of the MIP- (macrophag e infectivity potentiator) and the SS rDNA-fragment, it was possible t o establish a DNA extraction and a PCR method for the detection of Leg ionella sp. in water-samples and, after cultivation, in Amoeba sp.. Th erefore, water-samples from a warmwater-system in a hospital were take n. In all samples, legionellae were detected by the PCR method and ide ntified by cultivation and a direct immunfluorescence-method as L. pne umophila (serogroup 1). Legionellae and amoebae of the same water samp le were cocultured. Legionellae were also adherent at the outer-membra ne. To separate the amoebae from the legionellae, the amoebae were sed imented selectively by centrifugation at 200 x g. This washing procedu re had to be repeated seven times in order to eliminate the extraamoeb ale legionellae for sure. After DNA-extraction of water samples and he at treatment of the intraamoebale legionellae respectively, the amplif ication was performed with the MIP- and 5S rDNA-primers. In 14 of 16 c ocultivations growth of legionellae was found. This result and the det ection of legionellae and amoebae in the same water samples suggest th at an infection of amoebae may also take place in the watersystem of t he hospital. This is important for the disinfection as a procedure to eliminate legionellae, since intraamoebale bacteria are more resistant to environmental manipulation. Because in two of the cocultivations n o growth of legionellae in amoebae was found, it can be assumed that o nly specific subtypes of legionellae can infect amoebae species.