title: Resuscitation of viable but nonculturable Legionella pneumophila Philadelphia JR32 by Acanthamoeba castellanii.
abstract: Legionella pneumophila is an aquatic bacterium and is responsible for Legionnaires' disease in humans. Free-living amoebae are parasitized by legionellae and provide the intracellular environment required for the replication of this bacterium. In low-nutrient environments, however, L. pneumophila is able to enter a non-replicative viable but nonculturable (VBNC) state. In this study, L. pneumophila Philadelphia I JR 32 was suspended in sterilized tap water at 10(4) cells/ml. The decreasing number of bacteria was monitored by CFU measurements, acridine orange direct count (AODC), and hybridization with 16S rRNA-targeted oligonucleotide probes. After 125 days of incubation in water, the cells were no longer culturable on routine plating media; however, they were still detectable by AODC and by in situ hybridization. The addition of Acanthamoeba castellanii to the dormant bacteria resulted in the resuscitation of L. pneumophila JR 32 to a culturable state. A comparison of plate-grown legionellae and reactivated cells showed that the capacity for intracellular survival in human monocytes and intraperitoneally infected guinea pigs, which is considered a parameter for virulence, was not reduced in the reactivated cells. However, reactivation of dormant legionellae was not observed in the animal model.
authors.affiliation: Institut für Molekulare Infektionsbiologie, University of Würzburg, Germany.
authors.forename: M
authors.initials: M
authors.lastname: Steinert
authors.forename: L
authors.initials: L
authors.lastname: Emödy
authors.forename: R
authors.initials: R
authors.lastname: Amann
authors.forename: J
authors.initials: J
authors.lastname: Hacker
mesh_terms: D000048:Acanthamoeba / Q000382:microbiology*; D000818:Animals; D001431:Bacteriological Techniques; D001681:Biological Assay; D015169:Colony Count, Microbial; D006168:Guinea Pigs; D006801:Humans; D017403:In Situ Hybridization; D016952:Legionella pneumophila / Q000235:genetics / Q000254:growth & development* / Q000302:isolation & purification; D007877:Legionnaires' Disease / Q000382:microbiology*; D009000:Monocytes / Q000382:microbiology; D015345:Oligonucleotide Probes / Q000235:genetics; D010529:Peritoneal Cavity / Q000382:microbiology; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D013242:Sterilization; D014871:Water Microbiology
chemical_list: D015345:Oligonucleotide Probes; D012336:RNA, Ribosomal, 16S