Search

We evaluated the diagnostic performances of 2 media (BCYE, MWY) on 951 Legionella-positive hospital water samples. MWY allowed detecting Legionella in 89.2% of samples, but in 10.8% (103/951) Legionella was found on BCYE plates only. In samples where Legionella was isolated with other microorganisms (663/951), MWY was essential to detect the majority of positive samples (349/663, 52.6%), as fewer plates resulted unreadable; however, in those containing Legionella only, a higher frequency of positive samples was recorded with BCYE (94.8%, 273/288) compared to MWY (85.1%, 245/288). Considering the 484 concordant positive samples, overall Legionella counts were significantly higher on BCYE (P = 0.0029), with 47% of samples showing higher counts on BCYE compared to MWY plates. Furthermore, discordant samples (positive on only one medium) showed different relative proportions between Legionella pneumophila and non-pneumophila, the latter being found more frequently on BCYE only (P = 0.0296).Our findings confirm the appropriateness of the ISO 11731:2017 update., Competing Interests: Declaration of competing interests The authors report no conflicts of interest relevant to this article., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Background: Heater-cooler units (HCUs) used during cardiopulmonary bypass may become colonized with non-tuberculous mycobacteria (NTM), including Mycobacterium chimaera. Recently, a worldwide investigation conducted in hospitalized infected patients has detected M. chimaera in several Stockert 3T HCUs manufactured by LivaNova., Aim: Microbiological surveillance on Stockert 3T (LivaNova) and Maquet HCU40 (Getinge) devices as well as an evaluation of the efficacy of their recommended decontamination protocols., Methods: A total of 308 water samples were collected from 29 HCUs: 264 samples were collected from 17 Stockert 3T HCUs and 44 samples from 12 Maquet HCU40 devices. Samples were tested for total viable counts (TVCs) at both 22 and 36°C, Pseudomonas aeruginosa, coliform bacteria, and NTM. The microbiological surveillance began in June 2017 and ran until October 2019., Findings: A total of 308 HCU water samples were analysed, 65.5% of which yielded NTM. The most frequently colonized device with NTM was the Stockert 3T (88.2%), with a frequency of positive samples of 59.5% (157/264). The Maquet HCU40 devices less frequently yielded NTM (33.3%), with a frequency of positive water samples of 13.6% (6/44). Disinfection procedures were effective in reducing TVCs of bacteria with the exception of NTM species. NTM were detected in both pre-disinfection (50.1%) and post-disinfection (55.7%) samples, and no significant association was found between disinfection and NTM results both in Stockert 3T and Maquet HCU40 devices., Conclusion: This study suggests that manufacturers' procedures for disinfection are ineffective and/or inadequate. Until effective disinfection protocols become available, the only way to minimize the risk of NTM contamination is to closely monitor the water quality in the HCU, keep it as clean as possible, and treat it like any other biohazardous material., (Copyright © 2020 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.)

Background: Invasive non-tuberculous mycobacteria (NTM) infections are emerging worldwide in patients undergoing open-chest cardiac bypass surgery exposed to contaminated heater-cooler units (HCUs). Although this outbreak has been investigated by culturing bacteria isolated from HCU aerosol and water samples, these conventional methods have low-analytic sensitivity, high rates of sample contamination, and long turnaround time., Aim: To develop a simple and effective method to detect NTM in HCUs by real-time polymerase chain reaction (PCR), with a short laboratory turnaround time and reliable culture results., Methods: A total of 281 water samples collected from various HCUs at seven Italian hospitals were simultaneously screened for NTM by a propidium monoazide (PMA)-PCR assay and by conventional culture testing. The results were analysed with culture testing as the reference method., Findings: (i) The agreement between culture testing and PMA-PCR was 85.0% with a cycle threshold (C T ) cut-off value of <38 vs 80.0% with a C T of <43, with a moderate Cohen's κ-coefficient; (ii) the C T cut-off value of <42 was deemed more suitable for predicting positive specimens; (iii) given the low concentration of target DNA in water samples, the minimum volume to be tested was 1 L., Conclusion: The use of PMA-PCR for fast detection of NTM from environmental samples is highly recommended in order to ascertain whether HCUs may represent a potential source of human exposure to NTM. This reliable and simple method reduces laboratory turnaround time compared to conventional methods (one to two days vs eight weeks, respectively), thereby improving control strategies and effective management of HCUs., (Copyright © 2019 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.)

Dental unit waterlines (DUWLs) can be considered one of the possible routes of H. pylori transmission, although its presence in DUWLs has not yet been investigated thoroughly. The present study aimed to discover the prevalence of H. pylori and oral streptococci ( S. oralis and S. mutans ) in DUWLs to evaluate the risk of exposure to human pathogens in dental practices. We collected the output water from 60 dental chair units (DCUs) in 26 private dentistry settings in Turin, searching for H. pylori and oral streptococci (OS) DNA, with a polymerase chain reaction (PCR) technique. At the same time, dentists completed a questionnaire about their DCUs, their main activities, the presence of anti-retraction devices, their attitudes about disinfection, etc. No dental chair unit tested was contaminated with H. pylori or S. mutans ; only one dental chair was contaminated with S. oralis (1.7%). Considering the results, we can state that: (i) the lack of H. pylori DNA in water samples analyzed, suggests that municipal water is presumably treated with a sufficient chlorine level to inactivate DNA over time; (ii) the aspiration of oral fluids is limited by anti-retraction valves fitted distally to hand pieces; (iii) propidium monoazide qPCR (PMA-qPCR) could be a good technique to investigate and monitor potential environmental sources of infections such as DUWLs.

Pseudomonas aeruginosa is an environmental bacterium, ubiquitous in aquatic habitats and water distribution systems, including dental unit waterlines (DUWLs). We investigated the prevalence of P. aeruginosa in DUWLs from private dental settings. We also analyzed the relationship between P. aeruginosa contamination and the presence of Legionella spp. and total viable count (TVC) in order to suggest a simple and inexpensive protocol to test the quality of water from DUWLs. We detected and quantified P. aeruginosa both by culture and by a PMA (propidium monoazide)-qPCR method. Overall, we detected P. aeruginosa in 17 samples using the PMA-qPCR and in 11 samples using the culture. All culture-positive samples were positive with the PMA-qPCR too, with an agreement between the two methods of 93% and a Cohen's kappa coefficient of κ = 0.747 (good concordance). Comparing results with results of our previous study, we noted that (a) P. aeruginosa was isolated only from DUWLs with high TVC and (b) five out of six Legionella-positive samples were negative for Pseudomonas spp. Our final suggestion is that the cleanliness of DUWLs should be assessed by TVC because it is a good indicator of the presence of pathogens such as Legionella spp. and P. aeruginosa.

In this study we evaluated (1) the efficacy of a protocol that combines hydrogen peroxide (shock treatment) and ICX ® tablets (continuous treatment) for the control of microbial contamination in dental unit water lines, and (2) the in vitro antimicrobial activity of ICX ® tablets on collection and wild strains isolated from dental chair output waters. To assess the treatment effectiveness, the microbial load in the output water samples of three dental chairs were investigated: one control chair received only shock treatment. In vitro bactericidal activity was tested against Staphylococcus aureus and Pseudomonas aeruginosa. Data obtained from samples collected from chairs treated with ICX ® and shock treatment and data from the control chair did not differ significantly on the basis of microbial load. In the in vitro study, the product was unable to kill Gram-negative bacteria. These results show that the continuous introduction of ICX ® was not effective in maintaining low counts of the heterotrophic bacteria in the output water of dental devices, and shock treatment may be needed more frequently than monthly.

Legionella spp. are ubiquitous in aquatic habitats and water distribution systems, including dental unit waterlines (DUWLs). The aim of the present study was to determine the prevalence of Legionella in DUWLs and tap water samples using PMA-qPCR and standard culture methods. The total viable counts (TVCs) of aerobic heterotrophic bacteria in the samples were also determined. Legionella spp. were detected and quantified using the modified ISO 11731 culture method. Extracted genomic DNA was analysed using the iQ-Check Quanti Legionella spp. kit, and the TVCs were determined according to the ISO protocol 6222. Legionella spp. were detected in 100% of the samples using the PMA-qPCR method, whereas these bacteria were detected in only 7% of the samples using the culture method. The number of colony forming units (CFUs) of the TVCs in the DUWL and tap water samples differed, with the bacterial load being significantly lower in the tap water samples (p-value = 0). The counts obtained were within the Italian standard range established for potable water in only 5% of the DUWL water samples and in 77% of the tap water samples. Our results show that the level of Legionella spp. contamination determined using the culture method does not reflect the true scale of the problem, and consequently we recommend testing for the presence of aerobic heterotrophic bacteria based on the assumption that Legionella spp. are components of biofilms.

Quantitative polymerase chain reaction (qPCR) offers rapid, sensitive, and specific detection of Legionella in environmental water samples. In this study, qPCR and qPCR combined with propidium monoazide (PMA-qPCR) were both applied to hot-water system samples and compared to traditional culture techniques. In addition, we evaluated the ability of PMA-qPCR to monitor the efficacy of different disinfection strategies. Comparison between the quantification obtained by culture and by qPCR or PMA-qPCR on environmental water samples confirms that the concentration of Legionella estimated by GU/L is generally higher than that estimated in CFU/L. Our results on 57 hot-water-system samples collected from 3 different sites show that: i) qPCR results were on average 178-fold higher than the culture results (Δ log10=2.25), ii) PMA-qPCR results were on average 27-fold higher than the culture results (Δ log10=1.43), iii) propidium monoazide-induced signal reduction in qPCR were nearly 10-fold (Δ log10=0.95), and that iv) different degrees of correlations between the 3 methods might be explained by different matrix properties, but also by different disinfection methods affecting cultivability of Legionella. In our study, we calculated the logarithmic differences between the results obtained by PMA-qPCR and those obtained by culture, and we suggested an algorithm for the interpretation of PMA-qPCR results for the routine monitoring of healthcare water systems using a commercial qPCR system (iQ-check real-time PCR kit; Bio-Rad, Marnes-la-Coquette, France)., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The study aim was to determine retrospectively whether the parallel use of 2 media [buffered charcoal yeast extract (BCYE) and medium of Wadowsky and Yee (MWY)] to isolate Legionella spp. from water samples taken from hospital water supply systems increased the sensitivity of the culture method as compared with methods/protocols in which only seeding on a selective medium is used. We analyzed the results obtained from 931 positive water samples. In 484 of the 931 positive water samples, Legionella spp. was isolated in the presence of other microorganisms; in 83% (400/484), we used MWY to count suspected colonies, which gave a lower number of unreadable plates. In the 447 samples containing only Legionella spp., the highest frequency of positive samples (93%, 418/447) was obtained with BCYE, whereas seeding on MWY yielded 78% (348/447) (P < 0.001). Evaluation of the influence of the media on the Legionella spp. counts obtained by the 2 media showed that BCYE agar produced significantly higher counts than MWY (P < 0.001). The major conclusions that may be drawn from our data are as follows: 1) BCYE gives a high recovery rate of positive samples (93%) and a much greater yield of Legionella spp. than MWY; 2) BCYE was necessary for the detection of non-L. pneumophila spp. which grew poorly on selective media; 3) selective media [MWY or GVPC (glycine, vancomycin, polymyxin B, and cycloheximide)] were necessary for the recovery of Legionella spp. when the non-selective medium (BCYE) was difficult to interpret because of contaminating background flora. The use of different media is recommended for routine water tests in hospitals., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Background: The prevention and control of legionellosis in hospital settings involves environmental sampling, among other measures. The data yielded by sampling constitute an important means of risk assessment and provide a valid basis on which to plan remedial (cleansing and disinfection) and preventive (maintenance) interventions. This retrospective study had 2 objectives: (1) to evaluate the utility of biofilm sampling at distal sites and (2) to identify an efficient environmental sampling strategy., Methods: Samples of hot water and biofilm were collected between June 1999 and March 2008 from 41 hospitals in Italy's Piemonte region. We analyzed results of the samples (water and biofilm) taken from the same site and results of the water samples taken from the recirculation loop and water samples taken from the distal sites during the same sampling run., Results: Microbiological analysis was performed on 3910 pairs of samples (water/biofilm). In 81% of the pairs, the results were concordant; in 17% of the pairs, Legionella was isolated only from the water samples, and in only 2% of the pairs was Legionella isolated from the biofilm sample alone. Data from 299 sampling runs show that 79% (236) of results from the water samples taken from the recirculation loop and water samples taken from the distal sites during the same sampling run were concordant, and 21% (63) were discordant., Conclusions: Our findings suggest that hospitals could safely adopt a simpler (water sampling only without biofilm sampling) and more efficient (monitoring of the entire system through sampling of recirculation loop water) environmental sampling policy., (Copyright 2010 Association for Professionals in Infection Control and Epidemiology, Inc. Published by Mosby, Inc. All rights reserved.)

Objective: To assess the efficacy of an alternative disinfection method for hospital water distribution systems contaminated with Legionella., Methods: Disinfection with peracetic acid was performed in a small hospital contaminated with L. pneumophila serotype 1. The disinfectant was used at concentrations of 50 ppm (first three surveillance phases) and 1,000 ppm (fourth surveillance phase) for 30 minutes., Results: Environmental monitoring revealed that disinfection was maintained 1 week after treatment; however, levels of recontamination surpassing baseline values were detected after approximately 1 month. Comparison of water temperatures measured at the distal outlets showed a statistically significant association between temperature and bacterial load. The circulating water temperature was found to be lower in the two wards farthest away from the hot water production plant than in other wards. It was thought that the lower water temperature in the two wards promoted the bacterial growth even after disinfection., Conclusion: Peracetic acid may be useful in emergency situations, but does not provide definitive protection even if used monthly.