Maria Scaturro

Pneumonia associated with a dental unit waterline

Department of Infectious Parasitic and ImmuneMediated Diseases, Istituto Superiore di Sanita, Rome, Italy (M L Ricci BiolD, S Fontana PhD, F Pinci BiolD, M Scaturro PhD); Azienda Unita Sanitaria Locale, Forli, Italy (E Fiumana MD); Azienda Unita Sanitaria Locale, Cesena, Italy (M F Pedna BiolD); Agenzia Regionale Prevenzione e Ambiente, Bologna, Italy (M A Bucci Sabattini BiolD); and “G B Morgagni-Pierantoni” Hospital, Department of Emergency Anaesthesia and Intensive Care Unit, Forli, Italy (P Farolfi MD)

Persistence of the Same Strain of Legionella pneumophila in the Water System of an Italian Hospital for 15 Years

In 2004, an outbreak of legionnaires disease occurred in a hospital in northern Italy with a water system that had been disinfected multiple times since 1990 and equipped with a continuous disinfecting system. Molecular typing linked the outbreak to contamination of the hospital water system and demonstrated the persistence of a predominant strain of Legionella pneumophila for 15 years.

Determination of Legionella pneumophila susceptibility to Melaleuca alternifolia Cheel (tea tree) oil by an improved broth micro-dilution method under vapour controlled conditions

The aim of this study was to determine the in vitro activity of Melaleuca alternifolia Cheel (tea tree) oil (TTO) against 22 strains of Legionella pneumophila of different serogroup and source of isolation. Both a standard broth micro-dilution method, with slight modifications, and a micro-atmosphere diffusion method were used. Furthermore, we have established a simple sealing procedure in the micro-dilution method to determine the antibacterial activity of TTO against Legionella in aqueous phase. The results showed that L. pneumophila, quite irrespective of serogroup and source of isolation, is exquisitely sensitive to TTO, with minimal inhibitory concentration (MIC) ranging from 0.125 to 0.5% v/v, and a bactericidal activity at 0.5% v/v. In addition, we show here that TTO vapours exert critical activity, that must be controlled for reproducible MIC determinations. Overall, our data suggest that TTO could be active as anti-Legionella disinfectant, for control of water system contamination, especially in spas, in small waterlines or in particular respiratory medical devices.

A multicenter study of viable PCR using propidium monoazide to detect Legionella in water samples

Legionella quantification in environmental samples is overestimated by qPCR. Combination with a viable dye, such as Propidium monoazide (PMA), could make qPCR (named then vPCR) very reliable. In this multicentre study 717 artificial water samples, spiked with fixed concentrations of Legionella and interfering bacterial flora, were analysed by qPCR, vPCR and culture and data were compared by statistical analysis. A heat-treatment at 55 °C for 10 minutes was also performed to obtain viable and not-viable bacteria. When data of vPCR were compared with those of culture and qPCR, statistical analysis showed significant differences (P < 0.001). However, although the heat-treatment caused an abatement of CFU/mL ≤1 to 1 log10 unit, the comparison between untreated and heat-treated samples analysed by vPCR highlighted non-significant differences (P > 0.05). Overall this study provided a good experimental reproducibility of vPCR but also highlighted limits of PMA in the discriminating capability of dead and live bacteria, making vPCR not completely reliable.

Legionella pneumophila serogroup 5 infection in the presence of multiple environmental contamination. The importance of a bacteriological diagnosis

Legionella pneumophila is a pathogen that causes severe pneumonia in humans; L. pneumophila serogroup 1 accounts for at least 90% of infections. This is not linked to an environmental predominance of Legionella pneumophila 1, but may be due to a greater virulence of the strain. L. pneumophila sg 5 has also been reported, albeit less frequently, to be a cause of the disease. We report a case of L. pneumophila sg 5 occurring in a large hospital in southern Italy (Apulia region), where both L. pneumophila sg 1 and sg 5 were detected in the water supply; the nosocomial origin was demonstrated by molecular subtyping (PFGE). An environmental investigation, performed immediately after diagnosis of the case of legionellosis, identified a ow L. pneumophila sg 5 contamination level. Our experience highlights that in hospital, risk assessment, in rder to institute control measures for Legionella, should be carried out not only in response to a case of the disease and/or in risk wards only, as described in the Italian Guidelines, but periodically in every ward. The present study confirms that, although in the community L. pneumophila sg 1 is the most frequent strain isolated in both outbreaks and isolated cases, in hospital other serogroups and species may often cause infection because of the high susceptibility of the hosts.

Legionnaires' Disease Pseudoepidemic Due to Falsely Positive Urine Antigen Test Results

Maria Cristina Rota, Stefano Fontana, Carmen Montaño-Remacha, Maria Scaturro, Maria Grazia Caporali, Vincenzo Vullo, Laura Scorzolini, Andreina Ercole, Maria Luisa Ricci National Centre for Epidemiology, Surveillance and Health Promotion, National Institute of Health, Rome, Italy; Infectious, Parasitic and Immune-Mediated Diseases Department, National Institute of Health, Rome, Italy; Public Health and Infectious Diseases Department, Sapienza University of Rome, Rome, Italy; Public Health Prevention Department, Local Health Unit Rome A, Rome, Italy

Characterization of a spontaneous avirulent mutant of Legionella pneumophila Serogroup 6: Evidence of DotA and flagellin involvement in the loss of virulence

The pathogenesis of Legionella pneumophila mainly resides in its ability to inhibit the phagosome-lysosome fusion, which normally prevents the killing of the host cells. In order to characterize the molecular alterations that occurred in a spontaneous avirulent mutant of Legionella pneumophila serogroup 6, named Vir-, we investigated the ability of the mutant to adhere to and multiply in the WI26VA4 alveolar epithelial cell line and in human macrophages, when compared to its parental strain, Vir+. We also determined the colocalization of bacteria with LAMP-1 to gain an insight into the phagosome-lysosome fusion process. Additionally, we determined the flagellin expression and dotA nucleotide sequencing. We observed a lack of expression of flagellin and an in-frame mutation in the dotA. gene. The data obtained strongly suggest the loss of virulence of the mutant could probably be due to the absence of flagellin and the dysfunctional type IV secretion System, resulting from the DotA protein being severely compromised.

Legionnaires' disease contracted from patient workplace: First report of a severe case of coinfection with varicella-zoster virus

A middle-aged immunocompetent woman was diagnosed and treated for a severe pulmonary human herpesvirus 3 infection. During the treatment, an infection from Legionella pneumophila serogroup 1 was also diagnosed. This coinfection threatened the life of the patient and led to serious permanent sequelae. This report highlights the importance of preventing Legionella environmental contamination, suspecting Legionella coinfection in patients with viral pneumonia, and vaccinating susceptible adults against chickenpox.

A Fatal Case of Nosocomial Legionnaires' Disease: Implications From an Extensive Environmental Investigation and Isolation of the Bacterium From Blood Culture.

colonized with CPE. Prospective studies using innovative laboratory techniques are needed on patients with CPE during outbreaks and in endemic settings over periods of time, while in the hospital and after discharge. These studies will help us further understand the natural history of CPE colonization and determine the host/patient, environmental, and microbial factors that may help us predict which patients remain transiently, intermittently, or permanently colonized. This work is required to scientifically inform infection prevention and control strategies on the use of contact precautions and to facilitate risk stratification of patients previously identified as CPE colonized. This research becomes increasingly important as the number of patients with CPE increases, especially in settings where single room/isolation facilities are limited.

Identification and characterization of genes, encoding the 3-hydroxybutyrate dehydrogenase and a putative lipase, in an avirulent spontaneous Legionella pneumophila serogroup 6 mutant.

Legionella pneumophila is a pathogen widespread in aquatic environment, able to multiply both within amoebae and human macrophages. The aim of this study was to identify genes differently expressed in a spontaneous avirulent Legionella pneumophila serogroup 6 mutant, named Vir−, respect the parental strain (Vir+), and to determine their role in the loss of virulence. Protein profiles revealed some differences in Vir− proteomic maps, and among the identified proteins the undetectable 3‐hydroxybutyrate dehydrogenase (BdhA) and a down‐produced lipase. Both Legionella enzymes were studied before and were here further characterized at genetic level. A significant down‐regulation of both genes was observed in Vir− at the transcriptional level, but the use of defined mutants demonstrated that they did not affect the intracellular multiplication. A mutant (MS1) showed an accumulation of poly‐3‐hydroxybutyrate (PHB) granules suggesting a role of bdhA gene in its degradation process. The lipase deduced amino acid sequence revealed a catalytic triad, typical of the ‘lipase box’ characteristic of PHB de‐polymerase enzymes, that let us suppose a possible involvement of lipase in the PHB granule degradation process. Our results revealed unexpected alterations in secondary metabolic pathways possibly linking the loss of virulence to Legionella lack of energy sources.