A review of Legionella transmission risk in built environments: sources, regulations, sampling, and detection.

The risk of Legionella transmission in built environments remains a significant concern. Legionella can spread within buildings through aerosol transmission, prompting the exploration of airborne transmission pathways and proposing corresponding prevention and control measures based on building characteristics. To this end, a comprehensive literature review on the transmission risk of Legionella in built environments was performed. Four electronic databases (PubMed, Web of Science, Google Scholar, and CNKI) were searched from inception to March 2024 for publications reporting the risk of Legionella transmission in built environments. Relevant articles and gray literature reports were hand-searched, and 96 studies were finally included. Legionella pollution comes from various sources, mainly originates in a variety of built environments in which human beings remain for extended periods. The sources, outbreaks, national standards, regulations, and monitoring techniques for Legionella in buildings are reviewed, in addition to increases in Legionella transmission risk due to poor maintenance of water systems and long-distance transmission events caused by aerosol characteristics. Air and water sampling using various analytical methods helps identify Legionella in the environment, recognize sources in the built environments, and control outbreaks. By comparing the standard regulations of national organizations globally, the authors further highlight gaps and deficiencies in Legionella surveillance in China. Such advancements offer essential insights and references for understanding and addressing Legionella transmission risk in the built environment, with the potential to contribute to safeguarding public health and building environment safety.

Factors Associated with Legionella Detection in the Water Systems of National Lodging Organization Facilities with Water Management Programs in the United States.

A better understanding of risk factors and the predictive capability of water management program (WMP) data in detecting Legionella are needed to inform the efforts aimed at reducing Legionella growth and preventing outbreaks of Legionnaires' disease. Using WMPs and Legionella testing data from a national lodging organization in the United States, we aimed to (1) identify factors associated with Legionella detection and (2) assess the ability of WMP disinfectant and temperature metrics to predict Legionella detection. We conducted a logistic regression analysis to identify WMP metrics associated with Legionella serogroup 1 (SG1) detection. We also estimated the predictive values for each of the WMP metrics and SG1 detection. Of 5435 testing observations from 2018 to 2020, 411 (7.6%) had SG1 detection, and 1606 (29.5%) had either SG1 or non-SG1 detection. We found failures in commonly collected WMP metrics, particularly at the primary test point for total disinfectant levels in hot water, to be associated with SG1 detection. These findings highlight that establishing and regularly monitoring water quality parameters for WMPs may be important for preventing Legionella growth and subsequent disease. However, while unsuitable water quality parameter results are associated with Legionella detection, this study found that they had poor predictive value, due in part to the low prevalence of SG1 detection in this dataset. These findings suggest that Legionella testing provides critical information to validate if a WMP is working, which cannot be obtained through water quality parameter measurements alone.

Legionella colonization and 3D spatial location within a Pseudomonas biofilm.

Biofilms are known to be critical for Legionella settlement in engineered water systems and are often associated with Legionnaire's Disease events. One of the key features of biofilms is their heterogeneous three-dimensional structure which supports the establishment of microbial interactions and confers protection to microorganisms. This work addresses the impact of Legionella pneumophila colonization of a Pseudomonas fluorescens biofilm, as information about the interactions between Legionella and biofilm structures is scarce. It combines a set of meso- and microscale biofilm analyses (Optical Coherence Tomography, Episcopic Differential Interference Contrast coupled with Epifluorescence Microscopy and Confocal Laser Scanning Microscopy) with PNA-FISH labelled L. pneumophila to tackle the following questions: (a) does the biofilm structure change upon L. pneumophila biofilm colonization?; (b) what happens to L. pneumophila within the biofilm over time and (c) where is L. pneumophila preferentially located within the biofilm? Results showed that P. fluorescens structure did not significantly change upon L. pneumophila colonization, indicating the competitive advantage of the first colonizer. Imaging of PNA-labelled L. pneumophila showed that compared to standard culture recovery it colonized to a greater extent the 3-day-old P. fluorescens biofilms, presumably entering in VBNC state by the end of the experiment. L. pneumophila was mostly located in the bottom regions of the biofilm, which is consistent with the physiological requirements of both bacteria and confers enhanced Legionella protection against external aggressions. The present study provides an expedited methodological approach to address specific systematic laboratory studies concerning the interactions between L. pneumophila and biofilm structure that can provide, in the future, insights for public health Legionella management of water systems.

The impact of DNA extraction on the quantification of Legionella, with implications for ecological studies.

Monitoring the levels of opportunistic pathogens in drinking water is important to plan interventions and understand the ecological niches that allow them to proliferate. Quantitative PCR is an established alternative to culture methods that can provide a faster, higher-throughput, and more precise enumeration of the bacteria in water samples. However, PCR-based methods are still not routinely applied for Legionella monitoring, and techniques, such as DNA extraction, differ notably between laboratories. Here, we quantify the impact that DNA extraction methods had on downstream PCR quantification and community sequencing. Through a community science campaign, we collected 50 water samples and corresponding shower hoses, and compared two commonly used DNA extraction methodologies to the same biofilm and water phase samples. The two methods showed clearly different extraction efficacies, which were reflected in both the quantity of DNA extracted and the concentrations of Legionella enumerated in both the matrices. Notably, one method resulted in higher enumeration in nearly all samples by about one order of magnitude and detected Legionella in 21 samples that remained undetected by the other method. 16S rRNA amplicon sequencing revealed that the relative abundance of individual taxa, including sequence variants of Legionella, significantly varied depending on the extraction method employed. Given the implications of these findings, we advocate for improvement in documentation of the performance of DNA extraction methods used in drinking water to detect and quantify Legionella, and characterize the associated microbial community.IMPORTANCEMonitoring for the presence of the waterborne opportunistic pathogen Legionella is important to assess the risk of infection and plan remediation actions. While monitoring is traditionally carried on through cultivation, there is an ever-increasing demand for rapid and high-throughput molecular-based approaches for Legionella detection. This paper provides valuable insights on how DNA extraction affects downstream molecular analysis such as the quantification of Legionella through droplet digital PCR and the characterization of natural microbial communities through sequencing analysis. We analyze the results from a risk-assessment, legislative, and ecological perspective, showing how initial DNA processing is an important step to take into account when shifting to molecular-based routine monitoring and discuss the central role of consistent and detailed reporting of the methods used.

Novel induction of broad-spectrum antibiotics by the human pathogen Legionella.

The majority of antibiotics are natural products, with microorganism-generated molecules and their derivatives being the most prevalent source of drugs to treat infections. Thus, identifying natural products remains the most valuable resource for novel therapeutics. Here, we report the discovery of a series of dormant bacteria in honey that have bactericidal activity toward Legionella, a bacterial pathogen that causes respiratory disease in humans. We show that, in response to bacterial products secreted by Legionella, the honey bacteria release diffusible antimicrobial molecules. Remarkably, the honey bacteria only produce these molecules in response to Legionella spp., when compared to a panel of 24 bacterial pathogens from different genera. However, the molecules induced by Legionella have broad activity against several clinically important pathogens, including many high-priority pathogens. Thus, Legionella spp. are potent drivers of antimicrobial molecule production by uncharacterized bacteria isolated from honey, providing access to new antimicrobial products and an unprecedented strategy for discovering novel antibiotics. IMPORTANCE: Natural products generated by microorganisms remain the most viable and abundant source of new antibiotics. However, their discovery depends on the ability to isolate and culture the producing organisms and to identify conditions that promote antibiotic production. Here, we identify a series of previously undescribed bacteria isolated from raw honey and specific culture conditions that induce the production of antimicrobial molecules that are active against a wide variety of pathogenic bacteria.

Efficacy of chlorine-based disinfectants to control Legionella within premise plumbing systems.

Legionella is an opportunistic waterborne pathogen that causes Legionnaires' disease. It poses a significant public health risk, especially to vulnerable populations in health care facilities. It is ubiquitous in manufactured water systems and is transmitted via inhalation or aspiration of aerosols/water droplets generated from water fixtures (e.g., showers and hand basins). As such, the effective management of premise plumbing systems (building water systems) in health care facilities is essential for reducing the risk of Legionnaires' disease. Chemical disinfection is a commonly used control method and chlorine-based disinfectants, including chlorine, chloramine, and chlorine dioxide, have been used for over a century. However, the effectiveness of these disinfectants in premise plumbing systems is affected by various interconnected factors that can make it challenging to maintain effective disinfection. This systematic literature review identifies all studies that have examined the factors impacting the efficacy and decay of chlorine-based disinfectant within premise plumbing systems. A total of 117 field and laboratory-based studies were identified and included in this review. A total of 20 studies directly compared the effectiveness of the different chlorine-based disinfectants. The findings from these studies ranked the typical effectiveness as follows: chloramine > chlorine dioxide > chlorine. A total of 26 factors were identified across 117 studies as influencing the efficacy and decay of disinfectants in premise plumbing systems. These factors were sorted into categories of operational factors that are changed by the operation of water devices and fixtures (such as stagnation, temperature, water velocity), evolving factors which are changed in-directly (such as disinfectant concentration, Legionella disinfectant resistance, Legionella growth, season, biofilm and microbe, protozoa, nitrification, total organic carbon(TOC), pH, dissolved oxygen(DO), hardness, ammonia, and sediment and pipe deposit) and stable factors that are not often changed(such as disinfectant type, pipe material, pipe size, pipe age, water recirculating, softener, corrosion inhibitor, automatic sensor tap, building floor, and construction activity). A factor-effect map of each of these factors and whether they have a positive or negative association with disinfection efficacy against Legionella in premise plumbing systems is presented. It was also found that evaluating the effectiveness of chlorine disinfection as a water risk management strategy is further complicated by varying disinfection resistance of Legionella species and the form of Legionella (culturable/viable but non culturable, free living/biofilm associated, intracellular replication within amoeba hosts). Future research is needed that utilises sensors and other approaches to measure these key factors (such as pH, temperature, stagnation, water age and disinfection residual) in real time throughout premise plumbing systems. This information will support the development of improved models to predict disinfection within premise plumbing systems. The findings from this study will inform the use of chlorine-based disinfection within premise plumbing systems to reduce the risk of Legionnaires disease.

Two-year evaluation of Legionella in an aging residential building: Assessment of multiple potable water remediation approaches.

Legionella is an opportunistic waterborne pathogen that is difficult to eradicate in colonized drinking water pipes. Legionella control is further challenged by aging water infrastructure and lack of evidence-based guidance for building treatment. This study assessed multiple premise water remediation approaches designed to reduce Legionella pneumophila within a residential building located in an aging, urban drinking water system over a two-year period. Samples (n = 745) were collected from hot and cold-water lines and quantified via most probable number culture. Building-level treatment approaches included three single heat shocks, three single chemical shocks, and continuous low-level chemical disinfection in the potable water system. The building was highly colonized with L. pneumophila with 71 % L. pneumophila positivity. Single heat shocks had a statistically significant L. pneumophila reduction one day post treatment but no significant L. pneumophila reduction at one week, two weeks, and four weeks post treatment. The first two chemical shocks resulted in statistically significant L. pneumophila reduction at two days and four weeks post treatment, but there was a significant L. pneumophila increase at four weeks following the third chemical shock. Continuous low-level chemical disinfection resulted in statistically significant L. pneumophila reduction at ten weeks post treatment implementation. This demonstrates that in a building highly colonized with L. pneumophila, sustained remediation is best achieved using continuous low-level chemical treatment.

Outbreak of Legionnaires' disease in Rzeszów in 2023. / Ognisko choroby legionistów w Rzeszowie w 2023 roku.

BACKGROUND: Legionnaires' disease is a type of severe pneumonia caused by Legionella bacteria. The case fatality rate in this disease is 5-10%. People with various comorbidities, smokers and the elderly are at greater risk of developing the disease. OBJECTIVE: The aim of the work is to present the results of an epidemiological investigation into the outbreak of Legionnaires' disease that occurred in the city of Rzeszów and the surrounding area in August and September 2023 and to present the threat related to the presence of Legionella bacteria in water supply installations and networks. MATERIAL AND METHODS: The material for this publication was data from an epidemiological investigation conducted in the outbreak of Legionnaires disease in Rzeszów in 2023. RESULTS: Epidemiological investigation revealed 165 cases of Legionnaires' disease in the outbreak, including 152 confirmed cases and 13 probable cases. The case fatality rate in a legionellosis outbreak was 15%. Environmental tests were carried out in residential and public buildings and industrial installations during the investigation. As part of environmental tests, 187 water samples were collected, including 87 warm water samples. CONCLUSIONS: The outbreak of Legionnaires' disease in the city of Rzeszów draws attention to the potential threat from the Legionella bacteria to the health and life of especially elderly people suffering from chronic diseases. The environmental tests carried out confirmed the highest number of Legionella bacteria at medium and high levels in water samples taken in the private apartments of sick people. Despite the lack of strict legal regulations clearly specifying the obligations regarding periodic disinfection of internal hot water supply installations, cooperation with their owners should be undertaken to enforce plans and actions in this area.

Adherence to Legionella control regulations and guidelines in Norwegian nursing homes: a cross-sectional survey.

BACKGROUND: Infection by Legionella bacteria is a risk to elderly individuals in health care facilities and should be managed by preventing bacterial proliferation in internal water systems. Norwegian legislation calls for a mandatory Legionella-specific risk assessment with the subsequent introduction of an adapted water management programme. The present study investigates adherence to legislation and guidelines on Legionella control and prevention in Norwegian nursing homes. METHODS: A cross-sectional survey was distributed to Norwegian municipalities to investigate the status of Legionella specific risk assessments of internal water distribution systems and the introduction of water management programmes in nursing homes. RESULTS: A total of 55.1% (n = 228) of the participating nursing homes had performed Legionella-specific risk assessments, of which 55.3% (n = 126) stated that they had updated the risk assessment within the last year. 96.5% introduced a water management programme following a risk assessment, whereas 59.6% of the ones without a risk assessment did the same. Nursing homes with risk assessments were more likely to monitor Legionella levels than those without (61.2% vs 38.8%), to remove dead legs (44.7% vs 16.5%), and to select biocidal preventive treatment over hot water flushing (35.5% vs 4.6%). CONCLUSIONS: This study presents novel insight into Legionella control in Norway, suggesting that adherence to mandatory risk assessment in nursing homes is moderate-low. Once performed, the risk assessment seems to be advantageous as an introduction to future Legionella prevention in terms of the scope and contents of the water management programme.

Microbial Contamination of Dental Unit Waterlines among Dental Clinics of India- An In vitro Study.

BACKGROUND: Dental Unit Water Line (DUWL) deliver water to different handpieces in a dental unit. The water in DUWL circulates in a closed system, where it is taken from a container. The quality of dental water is of considerable importance since patients and dental staff are regularly exposed to water and aerosols generated from dental equipment. Output water from DUWLs may be a potential source of infection for both dental health care personnel and patients. AIM: To assess the microbial contamination in the DUWL among dental clinics in Chennai. MATERIALS AND METHODS: An in vitro study was conducted on 60 water samples from 20 dental clinics in Chennai in December 2019. Water samples were collected from three different sources of the Dental unit according to ADA guidelines. The collected samples were assessed for the presence of Aspergillus, Acinetobacter, Pseudomonas aeruginosa, and Legionella by agar plate method. The data were analysed using SPSS software version 20. RESULTS: Legionella was the most prevalent microorganism with 70% prevalence in a three-way syringe and 50% in scaler and airotor, followed by Pseudomonas aeruginosa and Acinetobacter with 10% prevalence in scaler and airotor and Aspergillus with a prevalence of 10% in the three-way syringe. CONCLUSION: Most of the dental units were contaminated with Aspergillus, Legionella, Pseudomonas aeruginosa and Acinetobacter which pose a serious threat to the patients as well as the dentists.

Automated cooling tower detection through deep learning for Legionnaires' disease outbreak investigations: a model development and validation study.

BACKGROUND: Cooling towers containing Legionella spp are a high-risk source of Legionnaires' disease outbreaks. Manually locating cooling towers from aerial imagery during outbreak investigations requires expertise, is labour intensive, and can be prone to errors. We aimed to train a deep learning computer vision model to automatically detect cooling towers that are aerially visible. METHODS: Between Jan 1 and 31, 2021, we extracted satellite view images of Philadelphia (PN, USA) and New York state (NY, USA) from Google Maps and annotated cooling towers to create training datasets. We augmented training data with synthetic data and model-assisted labelling of additional cities. Using 2051 images containing 7292 cooling towers, we trained a two-stage model using YOLOv5, a model that detects objects in images, and EfficientNet-b5, a model that classifies images. We assessed the primary outcomes of sensitivity and positive predictive value (PPV) of the model against manual labelling on test datasets of 548 images, including from two cities not seen in training (Boston [MA, USA] and Athens [GA, USA]). We compared the search speed of the model with that of manual searching by four epidemiologists. FINDINGS: The model identified visible cooling towers with 95·1% sensitivity (95% CI 94·0-96·1) and a PPV of 90·1% (95% CI 90·0-90·2) in New York City and Philadelphia. In Boston, sensitivity was 91·6% (89·2-93·7) and PPV was 80·8% (80·5-81·2). In Athens, sensitivity was 86·9% (75·8-94·2) and PPV was 85·5% (84·2-86·7). For an area of New York City encompassing 45 blocks (0·26 square miles), the model searched more than 600 times faster (7·6 s; 351 potential cooling towers identified) than did human investigators (mean 83·75 min [SD 29·5]; mean 310·8 cooling towers [42·2]). INTERPRETATION: The model could be used to accelerate investigation and source control during outbreaks of Legionnaires' disease through the identification of cooling towers from aerial imagery, potentially preventing additional disease spread. The model has already been used by public health teams for outbreak investigations and to initialise cooling tower registries, which are considered best practice for preventing and responding to outbreaks of Legionnaires' disease. FUNDING: None.

Legionella Serositis: A Rare Presentation.

BACKGROUND: Legionella has a higher prevalence in India than in the world. Legionaries' disease most commonly involves the lungs but because of increased awareness, extrapulmonary manifestations are also being diagnosed more frequently. CASE DESCRIPTION: We present a case of a young female with acute onset of fever and chest pain. On initial investigation, an electrocardiogram (ECG) reported widespread pulse rate (PR) depression suggestive of pericarditis which was confirmed by ECG. High-resolution computed tomography (HRCT) thorax suggested mild bilateral pleural effusion with normal lung parenchyma. elevated erythrocyte sedimentation rate (ESR), C-reactive protein (CRP) added to the diagnosis of serositis. Serological study for atypical organisms was remarkable for positive immunoglobulin M (IgM) for Legionella. She was treated with a high dose of steroids and azithromycin successfully. CONCLUSION: Isolated extrapulmonary presentation of legionaries disease is often overlooked and is common. So it should be always included in the diagnostic armamentarium as treatment is highly efficacious if started early.

Applications of Quantitative Microbial Risk Assessment to Respiratory Pathogens and Implications for Uptake in Policy: A State-of-the-Science Review.

BACKGROUND: Respiratory tract infections are major contributors to the global disease burden. Quantitative microbial risk assessment (QMRA) holds potential as a rapidly deployable framework to understand respiratory pathogen transmission and inform policy on infection control. OBJECTIVES: The goal of this paper was to evaluate, motivate, and inform further development of the use of QMRA as a rapid tool to understand the transmission of respiratory pathogens and improve the evidence base for infection control policies. METHODS: We conducted a literature review to identify peer-reviewed studies of complete QMRA frameworks on aerosol inhalation or contact transmission of respiratory pathogens. From each of the identified studies, we extracted and summarized information on the applied exposure model approaches, dose-response models, and parameter values, including risk characterization. Finally, we reviewed linkages between model outcomes and policy. RESULTS: We identified 93 studies conducted in 16 different countries with complete QMRA frameworks for diverse respiratory pathogens, including SARS-CoV-2, Legionella spp., Staphylococcus aureus, influenza, and Bacillus anthracis. Six distinct exposure models were identified across diverse and complex transmission pathways. In 57 studies, exposure model frameworks were informed by their ability to model the efficacy of potential interventions. Among interventions, masking, ventilation, social distancing, and other environmental source controls were commonly assessed. Pathogen concentration, aerosol concentration, and partitioning coefficient were influential exposure parameters as identified by sensitivity analysis. Most (84%, n=78) studies presented policy-relevant content including a) determining disease burden to call for policy intervention, b) determining risk-based threshold values for regulations, c) informing intervention and control strategies, and d) making recommendations and suggestions for QMRA application in policy. CONCLUSIONS: We identified needs to further the development of QMRA frameworks for respiratory pathogens that prioritize appropriate aerosol exposure modeling approaches, consider trade-offs between model validity and complexity, and incorporate research that strengthens confidence in QMRA results. https://doi.org/10.1289/EHP12695.

Residual disinfectant effectively suppresses Legionella species in drinking water distribution systems supplied by surface water in Minnesota, USA.

Seven public water systems in Minnesota, USA were analyzed from one to five times over a two-year period to assess temporal changes in the concentrations of total bacteria, Legionella spp., and Legionella pneumophila from source (i.e., raw water) through the water treatment process to the end water user. Bacterial biomass was collected by filtering large volumes of raw water (12 to 425 L, median: 38 L) or finished and tap water (27 to 1205 L, median: 448 L) using ultrafiltration membrane modules. Quantitative PCR (qPCR) was then used to enumerate all bacteria (16S rRNA gene fragments), all Legionella spp. (ssrA), and Legionella pneumophila (mip). Total coliforms, Escherichia coli, and L. pneumophila also were quantified in the water samples via cultivation. Median concentrations of total bacteria and Legionella spp. (ssrA) in raw water (8.5 and 4.3 log copies/L, respectively) decreased by about 2 log units during water treatment. The concentration of Legionella spp. (ssrA) in water collected from distribution systems inversely correlated with the total chlorine concentration for chloraminated systems significantly (p = 0.03). Although only 8 samples were collected from drinking water distribution systems using free chlorine as a residual disinfectant, these samples had significantly lower concentrations of Legionella spp. (ssrA) than samples collected from the chloraminated systems (p = 5 × 10-4). There was considerable incongruity between the results obtained via cultivation-independent (qPCR) and cultivation-dependent assays. Numerous samples were positive for L. pneumophila via cultivation, none of which tested positive for L. pneumophilia (mip) via qPCR. Conversely, a single sample tested positive for L. pneumophilia (mip) via qPCR, but this sample tested negative for L. pneumophilia via cultivation. Overall, the results suggest that conventional treatment is effective at reducing, but not eliminating, Legionella spp. from surface water supplies and that residual disinfection is effective at suppressing these organisms within drinking water distribution systems.

Comparison of culture media for the detection of Legionella spp. in sanitary water samples: Adaptation to ISO 11731:2017.

INTRODUCTION: Water sample culturing is the reference method to detect Legionella spp. in sanitary facilities. Until 2017, UNE-EN ISO 11731 only included the GVPC medium, which inhibits interfering microbiota but hinders the growth of Legionella spp. To improve its recovery, the new standard incorporates the BCYE medium into the working protocol. METHODS: We inoculated 1306 sanitary water samples onto BCYE and GVPC according to an accredited internal procedure. We compared the number of cfu/L of Legionella spp. detected in both media. RESULTS: The median in BCYE was 2000 cfu/L higher than in GVPC (P = .000). In the presence of high amounts of interfering microbiota, both media were similar; in the absence or low interfering microbiota BCYE was four times more sensitive than GVPC (P = .000). CONCLUSION: Including BCYE in the analysis of sanitary water significantly improves the recovery of Legionella spp. in low contaminated samples.

Upstream land use with microbial downstream consequences: Iron and humic substances link to Legionella spp.

Intensified land use can disturb water quality, potentially increasing the abundance of bacterial pathogens, threatening public access to clean water. This threat involves both direct contamination of faecal bacteria as well as indirect factors, such as disturbed water chemistry and microbiota, which can lead to contamination. While direct contamination has been well described, the impact of indirect factors is less explored, despite the potential of severe downstream consequences on water supply. To assess direct and indirect downstream effects of buildings, farms, pastures and fields on potential water sources, we studied five Swedish lakes and their inflows. We analysed a total of 160 samples in a gradient of anthropogenic activity spanning four time points, including faecal and water-quality indicators. Through species distribution modelling, Random Forest and network analysis using 16S rRNA amplicon sequencing data, our findings highlight that land use indirectly impacts lakes via inflows. Land use impacted approximately one third of inflow microbiota taxa, in turn impacting ∼20-50 % of lake taxa. Indirect effects via inflows were also suggested by causal links between e.g. water colour and lake bacterial taxa, where this influenced the abundance of several freshwater bacteria, such as Polynucleobacter and Limnohabitans. However, it was not possible to identify direct effects on the lakes based on analysis of physiochemical- or microbial parameters. To avoid potential downstream consequences on water supply, it is thus important to consider possible indirect effects from upstream land use and inflows, even when no direct effects can be observed on lakes. Legionella (a genus containing bacterial pathogens) illustrated potential consequences, since the genus was particularly abundant in inflows and was shown to increase by the presence of pastures, fields, and farms. The approach presented here could be used to assess the suitability of lakes as alternative raw water sources or help to mitigate contaminations in important water catchments. Continued broad investigations of stressors on the microbial network can identify indirect effects, avoid enrichment of pathogens, and help secure water accessibility.

Virulence Genes and Biofilm Formation Among Legionella pneumophila Isolates Collected from Hospital Water Sources.

Legionella pneumophila can be transmitted to people, especially immunocompromised patients, via hospital water pipe systems and cause severe pneumonia. The aim of our study was to investigate the presence of major virulence factor genes, ability of biofilms formation, and correlation between presence of Legionella isolates and temperature, pH, and residual chlorine of water. Hundred water samples were collected from nine hospitals in Tehran, Iran. Temperature, pH, and residual chlorine were determined during sampling. Different virulence genes and the ability to form biofilms were subsequently analyzed among the L. pneumophila isolates. Results showed that 12 (12%) samples were positive in culture method and all of the isolates were positive as L. pneumophila species (mip). A correlation was found between Legionella culture positivity and temperature and pH of water, but there was no significant correlation between residual chlorine of water samples and the presence of Legionella. The isolation of Legionella rate in summer and spring was higher than winter and autumn. Twelve (100%) isolates were positive for mip genes, 9 (75%) for dot genes, 8 (66.66%) for hsp, 6 (50%) for lvh, and 4 (33.33%) for rtx. All of the isolates displayed strong ability for biofilm production every three days. Two of these isolates (16.6%) displayed weak ability to form biofilm on the first day of incubation. This study revealed that water sources in hospitals were colonized by virulent Legionella and should be continuously monitored to avoid elevated concentrations of Legionella with visible biofilm formation.

Quantitative Microbial Risk Assessment Framework Incorporating Water Ages with Legionella pneumophila Growth Rates.

Water age in drinking water systems is often used as a proxy for water quality but is rarely used as a direct input in assessing microbial risk. This study directly linked water ages in a premise plumbing system to concentrations of Legionella pneumophila via a growth model. In turn, the L. pneumophila concentrations were used for a quantitative microbial risk assessment to calculate the associated probabilities of infection (Pinf) and clinically severe illness (Pcsi) due to showering. Risk reductions achieved by purging devices, which reduce water age, were also quantified. The median annual Pinf exceeded the commonly used 1 in 10,000 (10-4) risk benchmark in all scenarios, but the median annual Pcsi was always 1-3 orders of magnitude below 10-4. The median annual Pcsi was lower in homes with two occupants (4.7 × 10-7) than with one occupant (7.5 × 10-7) due to more frequent use of water fixtures, which reduced water ages. The median annual Pcsi for homes with one occupant was reduced by 39-43% with scheduled purging 1-2 times per day. Smart purging devices, which purge only after a certain period of nonuse, maintained these lower annual Pcsi values while reducing additional water consumption by 45-62%.

Initial waterline contamination by Pseudomonas aeruginosa in newly installed dental chairs.

Water contamination in dental unit waterlines (DUWLs) is a potential source of healthcare-associated infection during dental care. The aim of this study was to evaluate the microbiological quality of DUWLs water from newly installed dental chairs in a French University Hospital. The microbiological quality of water from 24 new DUWLs initially disinfected by ICX Renew-prior to use of the dental units for patient treatment-was assessed for total culturable aerobic bacteria at 22°C and 36°C, Legionella sp., Pseudomonas aeruginosa, and total coliforms. Among the 24 samples analyzed, 21 were compliant with the water quality levels: 19 had no bacteria, and 2 contained only 4 and 1 CFU/mL for total culturable aerobic bacteria at 22°C and 36°C, respectively. Three samples were non-compliant due to contamination by P. aeruginosa (4, 2, and 2 CFU/100 mL). Controlling and preventing the microbiological contamination of DUWLs, especially by pathogenic bacteria, at the time of the installation of the new dental chairs are crucial to prevent healthcare-associated infection in dentistry. IMPORTANCE: Dental unit waterlines (DUWLs) of new dental chairs may be contaminated before their first clinical use, so an initial shock disinfection is crucial at the time of their installation. The microbiological analyses are crucial to control the water quality of DUWLs before their first clinical use because their disinfection does not guarantee the elimination of all bacteria.

Susceptibility of Legionella gormanii Membrane-Derived Phospholipids to the Peptide Action of Antimicrobial LL-37-Langmuir Monolayer Studies.

LL-37 is the only member of the cathelicidin-type host defense peptide family in humans. It exhibits broad-spectrum bactericidal activity, which represents a distinctive advantage for future therapeutic targets. The presence of choline in the growth medium for bacteria changes the composition and physicochemical properties of their membranes, which affects LL-37's activity as an antimicrobial agent. In this study, the effect of the LL-37 peptide on the phospholipid monolayers at the liquid-air interface imitating the membranes of Legionella gormanii bacteria was determined. The Langmuir monolayer technique was employed to prepare model membranes composed of individual classes of phospholipids-phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylglycerol (PG), cardiolipin (CL)-isolated from L. gormanii bacteria supplemented or non-supplemented with exogenous choline. Compression isotherms were obtained for the monolayers with or without the addition of the peptide to the subphase. Then, penetration tests were carried out for the phospholipid monolayers compressed to a surface pressure of 30 mN/m, followed by the insertion of the peptide into the subphase. Changes in the mean molecular area were observed over time. Our findings demonstrate the diversified effect of LL-37 on the phospholipid monolayers, depending on the bacteria growth conditions. The substantial changes in membrane properties due to its interactions with LL-37 enable us to propose a feasible mechanism of peptide action at a molecular level. This can be associated with the stable incorporation of the peptide inside the monolayer or with the disruption of the membrane leading to the removal (desorption) of molecules into the subphase. Understanding the role of antimicrobial peptides is crucial for the design and development of new strategies and routes for combating resistance to conventional antibiotics.