DETECTION OF LEGIONELLA DNA IN HUMAN AND GUINEA-PIG URINE SAMPLES BY THE POLYMERASE CHAIN-REACTION

A detection system for Legionella DNA in urine samples based on the polymerase chain reaction (PCR) was developed and tested on infected guinea pigs and patients suffering from pneumonia. Results were compared with standard methods for diagnosis of Legionnaires' disease. A primer system was selected which amplifies a 108 bp DNA fragment of the 5S rRNA gene. The sensitivity of the PCR system was one femtogram of extracted Legionella DNA. Three methods were tested for pretreatment of urine samples. Of these, the Geneclean II kit (Bio 101, USA) gave the best results for artificially contaminated urine samples as well as those from infected guinea pigs or patients. Thirty-seven urine samples from 15 guinea pigs intraperitoneally infected with either Legionella pneumophila serogroup 1, 3 and 6 or Legionella micdadei, 26 urine samples of 21 patients suffering from pneumonia, and 30 control samples of patients with urinary tract infection (UTI) were tested. Legionella DNA was detected in 29 of the guinea pig urine samples; whereas, urinary antigen detection using EIA was positive in only 20 of the samples. PCR was also positive in the samples of 11 patients with pneumonia, 9 of which were confirmed by other microbiological methods, such as culture, direct fluorescent antibody test, urinary antigen detection and antibody testing. However, of the 30 control samples from patients with UTI, three samples yielded positive results. The results demonstrate that Legionella DNA is excreted in the urine of infected individuals and that the PCR shows a higher degree of sensitivity than EIA to the detection of soluble Legionella antigen in urine. Although the positive results obtained from control samples are a potential limitation of the test, these preliminary data suggest that PCR can be a sensitive method for the diagnosis of legionellosis from urine samples.