Einsatz molekularer Methoden zur Identifizierung und Charakterisierung gram-negativer nonfermentativer Stäbchenbakterien von Mukoviszidosepatienten

Abstract

Phenotypic identification of gram-negative bacteria from Cystic Fibrosis patients carries a high risk of misidentification. Therefore, the results of biochemical identification by API 20NE with 16S rRNA gene sequencing were compared in 88 gram-negative, oxidase-positive rods, other than morphologically and biochemically typical Pseudomonas aeruginosa, from respiratory secretions of CF patients. The API 20NE allowed correct identification of the bacterial species in 15 out of 88 (17%) isolates investigated. Agreement between API and the 16S rRNA gene sequencing results was high only in isolates with an API result classified as "excellent identification". Even API results classified as "very good identification" or "good identification" showed a high rate of misidentification (67% and 84%). 52 isolates of morphological and biochemical nontypical Pseudomonas aeruginosa, representing 59% of all isolates investigated, were not identifiable or misidentified in the Api 20NE. Misidentification of Pseudomonas aeruginosa, the clinically most relevant pathogen, has remarkable consequences for the patients. Furthermore, the 16S rRNA gene sequencing led to the detection of rarely or even newly described species in respiratory secretions of CF patients. For example, Inquilinus limosus, a recently described bacterium was identified in Germany for the first time. With molecular typing using PFGE, a chronic Burkholderia multivorans infection involving persistency of one strain and temporary, consecutive coinfection with two different strains was detected in a Cystic Fibrosis patient. Comparison of the colony morphology with the genotype revealed no correlation. These data are important for interpretation of clinical outcome and transmission studies in CF patients. In conclusion, molecular methods are superior over biochemical tests for identification of gram-negative, oxidase-positive rods in CF patients.