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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.

Background European Society of Cardiology 2015 guidelines approved 18F-fluorodeoxyglucose positron emission tomography/computed tomography (18F-FDG PET/CT) as a useful diagnostic imaging technique in prosthetic valve endocarditis (PVE) and recent evidence seems to suggest a role of nuclear imaging in the follow-up of cardiovascular infections, but nowadays there are no sufficient data available. Case summary A 67-year-old male presented with fever, weight loss, and fatigue. His medical history included ulcerative colitis and a previous Bentall-De Bono surgical procedure in 2014. A previous recent hospitalization to a small community hospital did not reveal a clear aetiology for the fever: transeosophageal echocardiography showed dubious peri-prosthetic tissue alterations, interpreted as post-surgical fibrosis; consequently, the patient was discharged with steroid therapy. At admission in our ward, we repeated transoesophageal echocardiography that confirmed the peri-prosthetic alterations. Moreover, 18F-FDG PET/CT showed two hypermetabolic areas, one around the prosthetic tube in the aortic bulb and the other in relation with the prosthetic aortic valve. Serological test was positive for Coxiella burnetii infection with consequent beginning of a targeted antimicrobial therapy with oral doxicycline and hydroxychloroquine. Echocardiography, serology, and 18F-FDG PET/CT follow-up demonstrated a progressive response to treatment and clinical conditions of the patient gradually improved. Discussion According to guidelines, 18F-FDG PET/CT can be used in ambiguous PVE to improve diagnostic accuracy of standard techniques. In this case, 18F-FDG PET/CT combined with echocardiography and serological tests is used not only to better define diagnosis but also for treatment response monitoring during follow-up.

Background/aims The microbiological surveillance of endoscopes and automated flexible endoscope reprocessing have been proven to be two of the most difficult and controversial areas of infection control in endoscopy. The purpose of this study was to standardize a sampling method for assessing the effectiveness of standard reprocessing operating procedures for flexible fiberoptic laryngoscopes (FFLs). Methods First, the sampling devices were directly inoculated with Bacillus atrophaeus spores; second, tissue non tissue (TNT) wipes were tested on artificially contaminated surfaces and on FFLs. Results Comparison of the sponges, cellulose, and TNT wipes indicated that the TNT wipes were more effective in releasing spores (93%) than the sponges (49%) and cellulose wipes (52%). The developed protocol provides a high efficiency for both collection and extraction from the stainless steel surface (87% of the spores were removed and released) and from the FFL (85% of the spores were removed and released), with relatively low standard deviations for recovery efficiency, particularly for the analysis of the FFL. Conclusion TNT wipes are more efficient for sampling surface areas, thereby aiding in the accuracy and reproducibility of environmental surveillance.

The majority of patients hospitalized for heart failure (HF) are admitted to internal medicine (IM) rather than to cardiology (CA) units, but to date few studies have analyzed the characteristics of these two populations. In this snapshot survey, we compared consecutive patients admitted for HF in six IM units vs. one non-intensive CA unit. During the 6-month survey period, 467 patients were enrolled (127 in CA, 27.2% vs. 340 in IM, 72.8%). IM patients were almost 10 years older (CA 75 ± 10, IM 82 ± 8 years; p

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.

This study was designed to examine the in vitro bactericidal activity of hydrogen peroxide against Legionella. We tested hydrogen peroxide (Peroxy Ag⁺) at 600 ppm to evaluate Legionella survival in a simulated dental treatment water system equipped with Water Hygienization Equipment (W.H.E.) device that was artificially contaminated. When Legionella pneumophila serogroup (sg) 1 was exposed to Peroxy Ag⁺ for 60 min we obtained a two decimal log reduction. High antimicrobial efficacy was obtained with extended periods of exposure: four decimal log reduction at 75 min and five decimal log reduction at 15 h of exposure. Involving a simulation device (Peroxy Ag⁺ is flushed into the simulation dental unit waterlines (DUWL)) we obtained an average reduction of 85% of Legionella load. The product is effective in reducing the number of Legionella cells after 75 min of contact time (99.997%) in the simulator device under test conditions. The Peroxy Ag⁺ treatment is safe for continuous use in the dental water supply system (i.e., it is safe for patient contact), so it could be used as a preventive option, and it may be useful in long-term treatments, alone or coupled with a daily or periodic shock treatment.

Two different real-time quantitative PCR (PMA-qPCR) assays were applied for quantification of Legionella spp. by targeting a long amplicon (approx 400 bp) of 16S rRNA gene and a short amplicon (approx. 100 bp) of 5S rRNA gene. Purified DNA extracts from pure cultures of Legionella spp. and from environmental water samples were quantified. Application of the two assays to quantify Legionella in artificially contaminated water achieved that both assays were able to detect Legionella over a linear range of 10 to 10(5) cells ml(-1). A statistical analysis of the standard curves showed that both assays were linear with a good correlation coefficient (R(2) = 0.99) between the Ct and the copy number. Amplification with the reference assay was the most effective for detecting low copy numbers (1 bacterium per PCR mixture). Using selective quantification of viable Legionella by the PMA-qPCR method we obtained a greater inhibition of the amplification of the 400-bp 16S gene fragment (Δlog(10) = 3.74 ± 0.39 log(10) GU ml(-1)). A complete inhibition of the PCR signal was obtained when heat-killed cells in a concentration below 1 × 10(5) cells ml(-1) were pretreated with PMA. Analysing short amplicon sizes led to only 2.08 log reductions in the Legionella dead-cell signal. When we tested environmental water samples, the two qPCR assays were in good agreement according to the kappa index (0.741). Applying qPCR combined with PMA treatment, we also obtained a good agreement (kappa index 0.615). The comparison of quantitative results shows that both assays yielded the same quantification sensitivity (mean log = 4.59 vs mean log = 4.31).

Evaluation of the potential risk associated with Legionella has traditionally been determined from culture-based methods. Quantitative polymerase chain reaction (qPCR) is an alternative tool that offers rapid, sensitive and specific detection of Legionella in environmental water samples. In this study we compare the results obtained by conventional qPCR (iQ-Check™ Quanti Legionella spp.; Bio-Rad) and by culture method on artificial samples prepared in Page's saline by addiction of Legionella pneumophila serogroup 1 (ATCC 33152) and we analyse the selective quantification of viable Legionella cells by the qPCR-PMA method. The amount of Legionella DNA (GU) determined by qPCR was 28-fold higher than the load detected by culture (CFU). Applying the qPCR combined with PMA treatment we obtained a reduction of 98.5% of the qPCR signal from dead cells. We observed a dissimilarity in the ability of PMA to suppress the PCR signal in samples with different amounts of bacteria: the effective elimination of detection signals by PMA depended on the concentration of GU and increasing amounts of cells resulted in higher values of reduction. Using the results from this study we created an algorithm to facilitate the interpretation of viable cell level estimation with qPCR-PMA.

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 log 10 =2.25), ii) PMA-qPCR results were on average 27-fold higher than the culture results (Δ log 10 =1.43), iii) propidium monoazide–induced signal reduction in qPCR were nearly 10-fold (Δ log 10 =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).