Considerations for determining the performance of ultraviolet light emitting diode fluid disinfection systems.

The International Ultraviolet Association (IUVA) Task Force was formed to develop guidelines regarding testing and reporting on performance of UV LED water disinfection systems. The goal was to provide clarity in a guidance document on measuring system performance across the global UV LED water disinfection system market. A review of current performance measurement protocols for mercury lamp based systems shows that the common elements of UV LED system performance measurement protocols should be as follows: specified standard for the amount of pathogen reduction required, the requirement that the validation testing be conducted by a competent facility, and that the system be continually monitored by UV sensors while in use to verify system performance unless pathogen reduction is not claimed. UV LEDs have selectable peak wavelengths, as opposed to mercury lamps that have fixed emission wavelength values. As a result of this difference, the following changes to protocols used to test mercury lamp systems are recommended. First, the use of disinfection benchmarks other than 254 nm dose, such as direct inactivation values, dose benchmarks referenced to 254 nm, and/or dose benchmarks at the UV LED emission wavelength that give the same inactivation as the original 254 nm UV dose benchmark. Second, the use of 254 nm UV water transmittance values as a placeholder, rather than an assumed correct value, for systems under test with LED wavelengths >250 nm and water transmittance values ≥87%. More research is needed for lower wavelengths and UVTs. Third, the recommendation that germicidal response UV sensors be used in UV LED based systems to ensure that the validated disinfection is delivered. Finally, additional LED-specific considerations were also noted. UV LEDs are also instant-on devices, making them ideal light sources for systems operated intermittently. Performance testing of systems operated intermittently should include a test to insure that pathogens do not migrate past the UV LEDs while the LEDs are off. UV LED devices have recognized protocols for determining the lifetime of the devices, as well as for measuring other device properties. Caution should be exercised in using these lifetime values for devices in UV disinfection systems, since the thermal environment of the devices may be different for protocol testing and disinfection system operation. PRACTITIONER POINTS: Validation of UVC LED fluid disinfection is necessary for point of use, point of entry, and municipal applications. The emission spectrum properties, considerations and measurements for output over lifetime, and unique system design considerations of UVC LEDs as light sources are factors that must be considered when evaluating fluid disinfection performance. The instant-on operation, system geometry, validation benchmarks, system sensing, water transmittance, and fouling must also be considered for UVC LED devices when evaluating fluid disinfection performance.

The Many Hosts of Mycobacteria 9 (MHM9): A conference report.

The Many Hosts of Mycobacteria (MHM) meeting series brings together basic scientists, clinicians and veterinarians to promote robust discussion and dissemination of recent advances in our knowledge of numerous mycobacterial diseases, including human and bovine tuberculosis (TB), nontuberculous mycobacteria (NTM) infection, Hansen's disease (leprosy), Buruli ulcer and Johne's disease. The 9th MHM conference (MHM9) was held in July 2022 at The Ohio State University (OSU) and centered around the theme of "Confounders of Mycobacterial Disease." Confounders can and often do drive the transmission of mycobacterial diseases, as well as impact surveillance and treatment outcomes. Various confounders were presented and discussed at MHM9 including those that originate from the host (comorbidities and coinfections) as well as those arising from the environment (e.g., zoonotic exposures), economic inequality (e.g. healthcare disparities), stigma (a confounder of leprosy and TB for millennia), and historical neglect (a confounder in Native American Nations). This conference report summarizes select talks given at MHM9 highlighting recent research advances, as well as talks regarding the historic and ongoing impact of TB and other infectious diseases on Native American Nations, including those in Southwestern Alaska where the regional TB incidence rate is among the highest in the Western hemisphere.

Fluence-based QMRA model for bacterial photorepair and regrowth in drinking water after decentralized UV disinfection.

Ultraviolet disinfection is a promising solution for decentralized drinking water systems such as communal water taps. A potential health risk is enzymatic photorepair of pathogens after UV disinfection, which can result in regrowth of pathogens. Even though photorepair is a known issue, no formal risk assessments have been conducted for photorepair after UV disinfection in drinking water. The main objective was to construct a quantitative microbial risk assessment (QMRA) of photorepair after UV disinfection of drinking water in a decentralized system. UV disinfection and photorepair kinetics for E. coli were modelled using reproducible fluence-based determinations. Impacts of water collection patterns, and wavelength-dependent water container material transmittance, sunlight intensity, and photorepair enzyme absorbance were quantified. After UV disinfection by 16 or 40 mJ/cm2 of < 5-log microorganisms per L, risk of infection did not exceed 1-in-10,000 under conditions permitting E. coli photorepair. Risk from photorepair was less than 1-in-10,000 for photorepair light exposure < 0.75 h throughout the day for UV fluence 16 mJ/cm2 or greater. UV disinfection followed by solar disinfection surpassing photoreactivation during storage reduced risk below 1-in-10,000 for photorepair light exposure > 2.5 h between modelled times of 9 AM - 3 PM. The model can be expanded to other pathogens as UV fluence and photorepair fluence response kinetics become available, and this QMRA can be used to inform the placement of community water access points to reduce risk of photorepair and ensure adequate shelf life of UV disinfected water under safe storage conditions.

UV222 disinfection of SARS-CoV-2 in solution.

There is an urgent need for evidence-based engineering controls to reduce transmission of SARS-CoV-2, which causes COVID-19. Although ultraviolet (UV) light is known to inactivate coronaviruses, conventional UV lamps contain toxic mercury and emit wavelengths (254 nm) that are more hazardous to humans than krypton chlorine excimer lamps emitting 222 nm (UV222). Here we used culture and molecular assays to provide the first dose response for SARS-CoV-2 solution exposed to UV222. Culture assays (plaque infectivity to Vero host) demonstrated more than 99.99% disinfection of SARS-CoV-2 after a UV222 dose of 8 mJ/cm2 (pseudo-first order rate constant = 0.64 cm2/mJ). Immediately after UV222 treatment, RT-qPCR assays targeting the nucleocapsid (N) gene demonstrated ~ 10% contribution of N gene damage to disinfection kinetics, and an ELISA assay targeting the N protein demonstrated no contribution of N protein damage to disinfection kinetics. Molecular results suggest other gene and protein damage contributed more to disinfection. After 3 days incubation with host cells, RT-qPCR and ELISA kinetics of UV222 treated SARS-CoV-2 were similar to culture kinetics, suggesting validity of using molecular assays to measure UV disinfection without culture. These data provide quantitative disinfection kinetics which can inform implementation of UV222 for preventing transmission of COVID-19.

Persistence of viable MS2 and Phi6 bacteriophages on carpet and dust.

Resuspension of dust from flooring is a major source of human exposure to microbial contaminants, but the persistence of viruses on dust and carpet and the contribution to human exposure are often unknown. The goal of this work is to determine viability of MS2 and Phi6 bacteriophages on cut carpet, looped carpet, and house dust both over time and after cleaning. Bacteriophages were nebulized onto carpet or dust in artificial saliva. Viability was measured at 0, 1, 2, 3, 4, 24, and 48 h and after cleaning by vacuum, steam, hot water extraction, and disinfection. MS2 bacteriophages showed slower viability decay rates in dust (-0.11 hr-1 ), cut carpet (-0.20 hr-1 ), and looped carpet (-0.09 hr-1 ) compared to Phi6 (-3.36 hr-1 , -1.57 hr-1 , and -0.20 hr-1 , respectively). Viable viral concentrations were reduced to below the detection limit for steam and disinfection for both MS2 and Phi6 (p < 0.05), while vacuuming and hot water extraction showed no significant changes in concentration from uncleaned carpet (p > 0.05). These results demonstrate that MS2 and Phi6 bacteriophages can remain viable in carpet and dust for several hours to days, and cleaning with heat and disinfectants may be more effective than standard vacuuming.

Indoor Dust as a Matrix for Surveillance of COVID-19.

Ongoing disease surveillance is a critical tool to mitigate viral outbreaks, especially during a pandemic. Environmental monitoring has significant promise even following widespread vaccination among high-risk populations. The goal of this work is to demonstrate molecular severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) monitoring in bulk floor dust and related samples as a proof of concept of a noninvasive environmental surveillance methodology for coronavirus disease 2019 (COVID-19) and potentially other viral diseases. Surface swab, passive sampler, and bulk floor dust samples were collected from the rooms of individuals positive for COVID-19, and SARS-CoV-2 was measured with quantitative reverse transcription-PCR (RT-qPCR) and two digital PCR (dPCR) methods. Bulk dust samples had a geometric mean concentration of 163 copies/mg of dust and ranged from nondetects to 23,049 copies/mg of dust detected using droplet digital PCR (ddPCR). An average of 89% of bulk dust samples were positive for the virus by the detection methods compared to 55% of surface swabs and fewer on the passive sampler (19% carpet, 29% polystyrene). In bulk dust, SARS-CoV-2 was detected in 76%, 93%, and 97% of samples measured by qPCR, chip-based dPCR, and droplet dPCR, respectively. Detectable viral RNA in the bulk vacuum bags did not measurably decay over 4 weeks, despite the application of a disinfectant before room cleaning. Future monitoring efforts should further evaluate RNA persistence and heterogeneity in dust. This study did not measure virus infectivity in dust or potential transmission associated with dust. Overall, this work demonstrates that bulk floor dust is a potentially useful matrix for long-term monitoring of viral disease in high-risk populations and buildings.IMPORTANCE Environmental surveillance to assess pathogen presence within a community is proving to be a critical tool to protect public health, and it is especially relevant during the ongoing COVID-19 pandemic. Importantly, environmental surveillance tools also allow for the detection of asymptomatic disease carriers and for routine monitoring of a large number of people as has been shown for SARS-CoV-2 wastewater monitoring. However, additional monitoring techniques are needed to screen for outbreaks in high-risk settings such as congregate care facilities. Here, we demonstrate that SARS-CoV-2 can be detected in bulk floor dust collected from rooms housing infected individuals. This analysis suggests that dust may be a useful and efficient matrix for routine surveillance of viral disease.

Attenuation of Helper T Cell Capacity for TH1 and TH17 Differentiation in Children With Nontuberculous Mycobacterial Infection.

Nontuberculous mycobacteria (NTM) infect children with increasing frequency worldwide. Using blood and lymph node tissue from children with NTM lymphadenitis, and uninfected lymph node tissue from community controls, we evaluated helper T (TH) cells in functional assays of TH1/TH17 differentiation and measured the concentration of their associated cytokines at the site of infection. Circulating TH cells from infected children were attenuated in their TH1/TH17 differentiation capacity and expressed less interferon γ and interleukin 17 after polyclonal stimulation. Similar differences were observed at the site of infection, where most cytokine concentrations were unchanged relative to controls. Our data are consistent with a model wherein TH1/TH17 differentiation is attenuated in NTM-infected children.

Drinking Water Microbiome Project: Is it Time?

Now is an opportune time to foster collaborations across sectors and geographical boundaries to enable development of best practices for drinking water (DW) microbiome research, focusing on accuracy and reproducibility of meta-omic techniques (while learning from past microbiome projects). A large-scale coordinated effort that builds on this foundation will enable the urgently needed comprehensive spatiotemporal understanding and control of DW microbiomes by engineering interventions to protect public health. This opinion paper highlights the need to initiate and conduct a large-scale coordinated DW microbiome project by addressing key knowledge gaps and recommends a roadmap for this effort.

Synergy of MS2 disinfection by sequential exposure to tailored UV wavelengths.

The advantages of polychromatic ultraviolet (UV) light for viral disinfection can be optimized for disinfection using emerging tailored wavelength sources including KrCl excimer lamps and light emitting diodes (LEDs). Disinfection of the common viral surrogate MS2 bacteriophage was measured after exposure to these emerging sources and conventional low pressure (LP) mercury UV lamps in individual or sequential exposures. The first dose response for any virus (MS2) exposed to a KrCl excimer lamp is reported, showing the high efficiency of fluence-based disinfection because of increased viral susceptibility at the low wavelengths emitted by the excilamp. Sequential exposure dose responses indicated synergy from sequential exposures of LP and excimer lamps, which were competitive on an electrical basis at worst-and best-case scenarios of wall plug efficiency with current medium pressure (MP) disinfection. Best-case scenarios for electrical efficiency also showed all sequential exposures to be competitive with MP UV disinfection. Predictive models for sequential exposure dose responses were assessed to support the current feasibility of incorporating sequential UV exposures to optimize tailored wavelength viral disinfection.

Succession of toxicity and microbiota in hydraulic fracturing flowback and produced water in the Denver-Julesburg Basin.

Hydraulic fracturing flowback and produced water (FPW) samples were analyzed for toxicity and microbiome characterization over 220 days for a horizontally drilled well in the Denver-Julesberg (DJ) Basin in Colorado. Cytotoxicity, mutagenicity, and estrogenicity of FPW were measured via the BioLuminescence Inhibition Assay (BLIA), Ames II mutagenicity assay (AMES), and Yeast Estrogen Screen (YES). Raw FPW stimulated bacteria in BLIA, but were cytotoxic to yeast in YES. Filtered FPW stimulated cell growth in both BLIA and YES. Concentrating 25× by solid phase extraction (SPE) revealed significant toxicity throughout well production by BLIA, toxicity during the first 55 days of flowback by YES, and mutagenicity by AMES. The selective pressures of fracturing conditions (including toxicity) affected bacterial and archaeal communities, which were characterized by 16S rRNA gene V4V5 region sequencing. Conditions selected for thermophilic, anaerobic, halophilic bacteria and methanogenic archaea from the groundwater used for fracturing fluid, and from the native shale community. Trends in toxicity echoed the microbial community, which indicated distinct stages of early flowback water, a transition stage, and produced water. Biota in another sampled DJ Basin horizontal well resembled similarly aged samples from this well. However, microbial signatures were unique compared to samples from DJ Basin vertical wells, and wells from other basins. These data can inform treatability, reuse, and management decisions specific to the DJ Basin to minimize adverse environmental health and well production outcomes.

Wavelength-Dependent Damage to Adenoviral Proteins Across the Germicidal UV Spectrum.

Adenovirus, a waterborne pathogen responsible for causing bronchitis, pneumonia, and gastrointestinal infections, is highly resistant to UV disinfection and therefore drives the virus disinfection regulations set by the U.S. Environmental Protection Agency. Polychromatic UV irradiation has been shown to be more effective at inactivating adenovirus and other viruses than traditional monochromatic irradiation emitted at 254 nm; the enhanced efficacy has been attributed to UV-induced damage to viral proteins. This research shows UV-induced damage to adenoviral proteins across the germicidal UV spectrum at wavelength intervals between 200 and 300 nm. A deuterium lamp with bandpass filters and UV light-emitting diodes (UV LEDs) isolated wavelengths in approximate 10 nm intervals. Sodium dodecyl sulfate polyacrylamide gel electrophoresis and image densitometry were used to detect signatures for the hexon, penton, fiber, minor capsid, and core proteins. The greatest loss of protein signature, indicating damage to viral proteins, occurred below 240 nm. Hexon and penton proteins exposed to a dose of 28 mJ/cm2 emitted at 214 nm were approximately 4 times as sensitive and fiber proteins approximately 3 times as sensitive as those exposed to a dose of 50 mJ/cm2 emitted at 254 nm. At 220 nm, a dose of 38 mJ/cm2 reduced the hexon and penton protein quantities to approximately 33% and 31% of the original amounts, respectively. In contrast, a much higher dose of 400 mJ/cm2 emitted at 261 and 278 nm reduced the original protein quantity to between 66-89% and 80-93%, respectively. No significant damage was seen with a dose of 400 mJ/cm2 at 254 nm. This research directly correlates enhanced inactivation at low wavelengths with adenoviral protein damage at those wavelengths, adding fundamental insight into the mechanisms of inactivation of polychromatic germicidal UV irradiation for improving UV water disinfection.

Longitudinal and Source-to-Tap New Orleans, LA, U.S.A. Drinking Water Microbiology.

The two municipal drinking water systems of New Orleans, LA, U.S.A. were sampled to compare the microbiology of independent systems that treat the same surface water from the Mississippi River. To better understand temporal trends and sources of microbiology delivered to taps, these treatment plants and distribution systems were subjected to source-to-tap sampling over four years. Both plants employ traditional treatment by chloramination, applied during or after settling, followed by filtration before distribution in a warm, low water age system. Longitudinal samples indicated microbiology to have stability both spatially and temporally, and between treatment plants and distribution systems. Disinfection had the greatest impact on microbial composition, which was further refined by filtration and influenced by distribution and premise plumbing. Actinobacteria spp. exhibited trends with treatment. In particular, Mycobacterium spp., very low in finished waters, occurred idiosyncratically at high levels in some tap waters, indicating distribution and/or premise plumbing as main contributors of mycobacteria. Legionella spp., another genus containing potential opportunistic pathogens, also occurred ubiquitously. Source water microbiology was most divergent from tap water, and each step of treatment brought samples more closely similar to tap waters.

Factors Influencing Bacterial Diversity and Community Composition in Municipal Drinking Waters in the Ohio River Basin, USA.

The composition and metabolic activities of microbes in drinking water distribution systems can affect water quality and distribution system integrity. In order to understand regional variations in drinking water microbiology in the upper Ohio River watershed, the chemical and microbiological constituents of 17 municipal distribution systems were assessed. While sporadic variations were observed, the microbial diversity was generally dominated by fewer than 10 taxa, and was driven by the amount of disinfectant residual in the water. Overall, Mycobacterium spp. (Actinobacteria), MLE1-12 (phylum Cyanobacteria), Methylobacterium spp., and sphingomonads were the dominant taxa. Shifts in community composition from Alphaproteobacteria and Betaproteobacteria to Firmicutes and Gammaproteobacteria were associated with higher residual chlorine. Alpha- and beta-diversity were higher in systems with higher chlorine loads, which may reflect changes in the ecological processes structuring the communities under different levels of oxidative stress. These results expand the assessment of microbial diversity in municipal distribution systems and demonstrate the value of considering ecological theory to understand the processes controlling microbial makeup. Such understanding may inform the management of municipal drinking water resources.

Molecular analysis of single room humidifier bacteriology.

Portable, single-room humidifiers are commonly used in homes for comfort and health benefits, but also create habitats for microbiology. Currently there is no information on home humidifier microbiology aside from anecdotal evidence of infection with opportunistic pathogens and irritation from endotoxin exposure. To obtain a broader perspective on humidifier microbiology, DNAs were isolated from tap source waters, tank waters, and biofilm samples associated with 26 humidifiers of ultrasonic and boiling modes of operation in the Front Range of Colorado. Humidifiers sampled included units operated by individuals in their homes, display models continuously operated by a retail store, and new humidifiers operated in a controlled laboratory study. The V1V2 region of the rRNA gene was amplified and sequenced to determine the taxonomic composition of humidifier samples. Communities encountered were generally low in richness and diversity and were dominated by Sphingomonadales, Rhizobiales, and Burkholderiales of the Proteobacteria, and MLE1-12, a presumably non-photosynthetic representative of the cyanobacterial phylum. Very few sequences of potential health concern were detected. The bacteriology encountered in source waters sampled here was similar to that encountered in previous studies of municipal drinking waters. Source water bacteriology was found to have the greatest effect on tank water and biofilm bacteriology, an effect confirmed by a controlled study comparing ultrasonic and boiler humidifiers fed with tap vs. treated (deionized, reverse osmosis, 0.2 µm filtered) water over a period of two months.