Detection of Legionella pneumophila in water samples by species-specific real-time and nested PCR assays.

AIMS: Legionella pneumophila is a contaminant of man-made water systems, including potable water, cooling towers, water systems of large buildings, etc. It is the most common causative agent of legionellosis, a respiratory infection, which may give rise to restricted outbreaks. To survey environmental water samples from hospitals and private habitations in Bologna, we developed a species-specific nested and a TaqMan real-time PCR for the detection of L. pneumophila. We compared the two assays and both to cultural isolation. METHODS AND RESULTS: The targeted gene was macrophage infectivity potentiator (mip), conserved in L. pneumophila, and divergent in other legionellae. One assay was based on a nested PCR and the other on a TaqMan real-time PCR protocol. Their sensitivities were 14 % or 5% higher than that of cultural isolation respectively. The detection limits were 1-2 genome equivalents per 50-microl reaction. Specificity was assessed using DNA from nine target and 20 nontarget organisms. CONCLUSIONS: When applied to water samples, both assays detected L. pneumophila at 80% or higher frequency. SIGNIFICANCE AND IMPACT OF THE STUDY: The species-specific molecular diagnosis of L. pneumophila by means of nested PCR does not require a specific instrumentation, exhibits a high sensitivity, and is advantageous over the cultural isolation and real-time PCR detection. It allows to quickly monitor water samples for the risk assessment of environmental contaminations.

Prevalence of Legionella spp. in swimming pool environment.

A study was carried out to evaluate the prevalence of bacteria of the Legionella genus in the water from the pools and showers of 12 swimming pools in the city of Bologna (Italy). Sampling took place four times for every establishment, once for each season of the year. Legionella spp. were found in 2 of the 48 pool water samples (L. micdadei and L. bozenanii) and in 27 of the 48 samples taken from the hot water of the showers: 19 were positive for L. pneumophila (10-19,250 cfu/l) and 18 for other species (20-6000 cfu/l). The contamination was seen to be consistent throughout the year, since the same species of legionellae were isolated at different samplings and in concentrations that did not reveal any seasonal trend. The legionellae and the Gram negative bacteria were not found in shower water with a temperature above 43 degrees C. However, in the samples of shower water with a lower temperature a statistically significant inverse correlation was seen between legionellae and Pseudomonas aeruginosa (r = -0.51; p<0.01) as well as between legionellae and Gram negative bacteria (r = -0.70; p<0.01). The potential risk of contracting infections from Legionella spp. in the swimming pool environment does not seem to be linked to the pool water, but to that of the showers. The water temperature of showers should therefore be maintained at a level high enough to prevent the reproduction of these bacteria.

[In vitro cytotoxicity in the immunological study of human glioblastomas]. / Citotossicità in vitro per lo studio immunologico die glioblastomi umani.

An immunological study about the lymphocytes of nineteen patients affected by glioblastoma has been executed by using CML and ADCC tests. Eleven healthy subjects and nine ones affected by bladder carcinoma have been studied for control. The CML test has demonstrated an increase of citotoxic activity of lymphocytes in the totality of the patients affected by glioblastoma (37,09 +/- 3,67)% versus (4,00 +/- 1,78)% of controls. The ADCC test has demonstrated diminution of citotoxic activity of lymphocytes of the patients affected by glioblastoma in comparison with controls (20,57 +/- 9,77)% versus (29,18 +/- 6,67)% of the healthy controls versus (27,66 +/- 8,51)% of bearers of bladder carcinomas.