Serological and molecular identification of Legionella spp. isolated from water and surrounding air samples in Italian healthcare facilities.

BACKGROUND: Legionella is an intracellular microorganism living in natural and artificial aquatic environments. Although its transmission to humans is linked to the inhalation of contaminated aerosols, there is no validated air sampling method for the control and prevention of the disease. The aim of the present study was to provide more information on the distribution of Legionella spp. in indoor environments and to determine whether the same Legionella strains are isolated from air and water samples. METHODS: Ten healthcare facilities located in seven regions of Italy were enrolled. The serological typing of Legionella spp. from water samples and the surrounding air by active and passive sampling was assessed using polyvalent and monovalent antisera. Subsequently, the strains identified as Legionella pneumophila (Lpn) underwent molecular typing by sequence-based typing (SBT) using seven genes (flaA, pilE, asd, mip, mompS, proA, and neuA). The allelic profile number was assigned using the European Working Group for Legionella Infections-SBT database. RESULTS: Lpn serogroup 6 was the most prevalent serogroup; it was found simultaneously in the air and water samples of three different healthcare facilities. In the remaining seven hospitals, Lpn serogroups 1, 6, 7, 9, and 12 were isolated exclusively from water samples. The molecular investigation showed that Lpn strains in the water and air samples of each positive healthcare facility had the same allelic profile. Strains, identified as sequence types (STs) 728 and ST 1638+ST 1324, were isolated in two respective healthcare facilities, and a new strain, identified as ST 1989, was obtained in one healthcare facility. CONCLUSION: The application of the SBT method allowed to verify the homology among Legionella strains from water samples and the surrounding air. The results showed that the same Lpn strains were present in the air and water samples, and a new Legionella strain was identified.

Analytical performance issues: comparison of ATP bioluminescence and aerobic bacterial count for evaluating surface cleanliness in an Italian hospital.

Contaminated hospital surfaces have been demonstrated to be an important environmental reservoir of microorganisms that can increase the risk of nosocomial infection in exposed patients. As a consequence, cleaning and disinfecting hospital environments play an important role among strategies for preventing healthcare-associated colonization and infections. The aim of the present study was to evaluate whether adenosine triphosphate (ATP) presence, measured by bioluminescence methods, can predict microbiological contamination of hospital surfaces. The study was carried out between September and December 2012 at the University Hospital "P. Giaccone" of Palermo. A total of 193 randomly selected surfaces (tables, lockers, furnishings) were sampled and analyzed in order to assess ATP levels (expressed as relative light units or RLU) and aerobic colony count (ACC) or presence of S. aureus. ACC had median values of 1.85 cfu/cm(2)(interquartile range = 4.16) whereas ATP median was 44.6 RLU/cm(2)(interquartile range = 92.3). Overall, 85 (44.0%) surfaces exceeded the established microbial benchmark: 73 (37.8%) exceeded the 2.5 cfu/cm(2)ACC standard, 5 (2.6%) surfaces were positive for S. aureus and 7 (3.6%) showed both the presence of S. aureus and an ACC of more than 2.5 cfu/cm(2). ACC and bioluminescence showed significant differences in the different surface sites (p < 0.001). A significant correlation was found between ACC and RLU values (p-value < 0.001; R(2)= 0.29) and increasing RLU values were significantly associated with a higher risk of failing the benchmark (p < 0.001). Our data suggest that bioluminescence could help in measuring hygienic quality of hospital surfaces using a quick and sensitive test that can be an useful proxy of microbial contamination; however, further analysis will be necessary to assess the cost-efficacy of this methodology before requiring incorporation in hospital procedures.

[Legionella spp. contamination in indoor air: preliminary results of an Italian multicenter study]. / La contaminazione indoor da Legionella spp: risultati preliminari di una indagine multicentrica italiana.

OBJECTIVE: To propose a standardized protocol for the evaluation of Legionella contamination in air. DESIGN: A bathroom having a Legionella contamination in water >1,000 cfu/l was selected in 10 different healthcare facilities. Air contamination was assessed by active (Surface Air System, SAS) and passive (Index of Microbial Air, IMA) sampling for 8 hours, about 1 m away from the floor and 50 cm from the tap water. Two hundred liters of air were sampled by SAS every 12 min, after flushing water for 2 min. The IMA value was calculated as the mean value of colony forming units/16 plates exposed during sampling (2 plates/hour). Water contamination was evaluated at T0, after 4 and 8 hours, according to the standard methods. RESULTS: Air contamination by Legionella was found in three healthcare facilities (one with active and two with passive sampling), showing a concomitant tap water contamination (median=40,000; range 1,100-43,000 cfu/l). The remaining seven hospitals isolated Legionella spp. exclusively from water samples (median=8,000; range 1,200-70,000 cfu/l). CONCLUSIONS: Our data suggest that environmental Legionella contamination cannot be assessed only through the air sampling, even in the presence of an important water contamination.

Can the serological status of anti-HBc alone be considered a sentinel marker for detection of occult HBV infection?

Some individuals have "occult" infection with hepatitis B virus (HBV), defined as presence of HBV genome in the serum or liver tissue without HBV surface antigen (HBsAg) in the serum. The aim of this study was to investigate whether serum antibodies against HBV core antigen in isolation ("anti-HBc alone") are a useful marker of "occult" HBV in patients with or without hepatitis C virus (HCV) infection. "Anti-HBc alone" was detected in the sera of 119/6,544 (1.8%) asymptomatic outpatients referred to the diagnostic laboratory for routine testing for viral hepatitis, 62/607 (10.2%) drug users, and 42/195 (21.5%) patients with hepatocellular carcinoma. Using three in-house nested-PCR amplification assays to detect HBV preS-S (S), precore-core (C), and Pol viral regions, respectively, "occult" HBV sequences were found in 9 of the 223 sera (4.0%) with "anti-HBc alone." The highest prevalence of "occult" HBV sequences (5.9%) was detected in "anti-HBV alone" sera of individuals referred to the diagnostic laboratory without HCV antibodies. Direct sequencing of all PCR products confirmed the specificity of the PCR reactions and revealed the predominance of HBV genotype D. The data presented in this study suggest that detection of "anti-HBc alone" could reflect unrecognized "occult" HBV infection and that physicians should consider investigating such patients with HBV molecular tests.