Virus inactivation in groundwater in a postglacial lava field in arctic climate.

Outbreaks of viral gastroenteritis are often connected to contaminated drinking water. The assessment of the water quality relies on the cultivation of indicator bacteria, and little is known of the fate of viruses in groundwater, especially in arctic regions. In Iceland, the groundwater temperature is between 3 and 6°C. The aim of this study was to determine virus inactivation at low temperature in a groundwater microcosm and in a borehole in a postglacial lava field. Two phage species that are commonly used as surrogates for norovirus were used, MS2 and PhiX174. Dialysis bags were used for the samples, and a device was constructed to hold many samples at a time and protect the samples in the borehole. No significant decrease of infective PhiX174 phages in the borehole or of the MS2 phages in the microcosm was observed. A slightly significant decrease of PhiX174 in the microcosm was noticed, with one log reduction time of 476 days. On the other hand, a significant reduction in MS2 was found in the field test, where the time needed for 90% reduction was 12·5 days. The results showed that an infective virus can exist in groundwater for months or years in arctic regions and a great difference may exist between results from microcosm and field tests. SIGNIFICANCE AND IMPACT OF THE STUDY: This study reveals that arctic regions are highly sensitive to virus contamination as an infective virus may exist in groundwater for years at low temperature. The results also show that the virus inactivation observed in field tests may differ considerably from the inactivation observed in laboratory microcosms. The results emphasize the importance of large protection zones around drinking water intakes as well as good wastewater treatment so that the likelihood of faecal contamination of groundwater is reduced.

Effect of polymer and glass physicochemical properties on MS2 recovery from food contact surfaces.

Viruses are transmissible via their interaction with contact surfaces of food containers or tools. This study evaluated the recoveries of MS2 coliphage, a virus surrogate, from polypropylene (PP), polyvinyl chloride (PVC), polyethylene (PE), and glass (borosilicate and soda lime), as influenced by the surface chemistry and topography. MS2 (5-6 logs) in PBS with 1% TSB was inoculated onto each of 9 different surfaces, 24-h cold-incubated, and recovery was quantified by infectivity. The order of MS2 recovery efficiency from smooth surfaces was PP > PE ≥ soda lime glass, which classified into 3 ANOVA groups, p = 0.05. The MS2 recovery ratios of smooth vs. rough surfaces were 1.4-1.5. Atomic force microscopy revealed 21-nm diam pinholes (<28-nm of MS2 size) in the borosilicate glass. The lowest and highest MS2 recoveries among the 9 surfaces were demonstrated by the hole-bearing borosilicate glass (34 ±â€¯8%) and smooth PP (69 ±â€¯14%) respectively. Generally greater MS2 recovery was obtained from smooth PP and PE surfaces compared to glass, but topographic alterations (pinholes or increased roughness) decreased recovery possibly by trapping the viruses.

Measurements of the self-assembly kinetics of individual viral capsids around their RNA genome.

Self-assembly is widely used by biological systems to build functional nanostructures, such as the protein capsids of RNA viruses. But because assembly is a collective phenomenon involving many weakly interacting subunits and a broad range of timescales, measurements of the assembly pathways have been elusive. We use interferometric scattering microscopy to measure the assembly kinetics of individual MS2 bacteriophage capsids around MS2 RNA. By recording how many coat proteins bind to each of many individual RNA strands, we find that assembly proceeds by nucleation followed by monotonic growth. Our measurements reveal the assembly pathways in quantitative detail and also show their failure modes. We use these results to critically examine models of the assembly process.

A new approach to testing the efficacy of drinking water disinfectants.

New disinfection procedures are being developed and proposed for use in drinking-water production. Authorising their use requires an effective test strategy that can simulate conditions in practice. For this purpose, we developed a test rig working in a flow-through mode similar to the disinfection procedures in waterworks, but under tightly defined conditions, including very short contact times. To quantify the influence of DOC, temperature and pH on the efficacy of two standard disinfectants, chlorine and chlorine dioxide, simulated use tests were systematically performed. This test rig enabled quantitative comparison of the reduction of four test organisms, two viruses and two bacteria, in response to disinfection. Chlorine was substantially more effective against Enterococcus faecium than chlorine dioxide whereas the latter was more effective against the bacteriophage MS2, especially at pH values of >7.5 at which chlorine efficacies already decline. Contrary to expectation, bacteria were not generally reduced more quickly than viruses. Overall, the results confirm a high efficacy of chlorine and chlorine dioxide, validating them as standard disinfectants for assessing the efficacy of new disinfectants. Furthermore, these data demonstrate that the test rig is an appropriate tool for testing new disinfectants as well as disinfection procedures.

Microbial inactivation and cytotoxicity evaluation of UV irradiated coconut water in a novel continuous flow spiral reactor.

A continuous-flow UV reactor operating at 254nm wave-length was used to investigate inactivation of microorganisms including bacteriophage in coconut water, a highly opaque liquid food. UV-C inactivation kinetics of two surrogate viruses (MS2, T1UV) and three bacteria (E. coli ATCC 25922, Salmonella Typhimurium ATCC 13311, Listeria monocytogenes ATCC 19115) in buffer and coconut water were investigated (D10 values ranging from 2.82 to 4.54mJ·cm-2). A series of known UV-C doses were delivered to the samples. Inactivation levels of all organisms were linearly proportional to UV-C dose (r2>0.97). At the highest dose of 30mJ·cm-2, the three pathogenic organisms were inactivated by >5 log10 (p<0.05). Results clearly demonstrated that UV-C irradiation effectively inactivated bacteriophage and pathogenic microbes in coconut water. The inactivation kinetics of microorganisms were best described by log linear model with a low root mean square error (RMSE) and high coefficient of determination (r2>0.97). Models for predicting log reduction as a function of UV-C irradiation dose were found to be significant (p<0.05) with low RMSE and high r2. The irradiated coconut water showed no cytotoxic effects on normal human intestinal cells and normal mouse liver cells. Overall, these results indicated that UV-C treatment did not generate cytotoxic compounds in the coconut water. This study clearly demonstrated that high levels of inactivation of pathogens can be achieved in coconut water, and suggested potential method for UV-C treatment of other liquid foods. INDUSTRIAL RELEVANCE: This research paper provides scientific evidence of the potential benefits of UV-C irradiation in inactivating bacterial and viral surrogates at commercially relevant doses of 0-120mJ·cm-2. The irradiated coconut water showed no cytotoxic effects on normal intestinal and healthy mice liver cells. UV-C irradiation is an attractive food preservation technology and offers opportunities for horticultural and food processing industries to meet the growing demand from consumers for healthier and safe food products. This study would provide technical support for commercialization of UV-C treatment of beverages.

A Light-Activated Antimicrobial Surface Is Active Against Bacterial, Viral and Fungal Organisms.

Evidence has shown that environmental surfaces play an important role in the transmission of nosocomial pathogens. Deploying antimicrobial surfaces in hospital wards could reduce the role environmental surfaces play as reservoirs for pathogens. Herein we show a significant reduction in viable counts of Staphylococcus epidermidis, Saccharomyces cerevisiae, and MS2 Bacteriophage after light treatment of a medical grade silicone incorporating crystal violet, methylene blue and 2 nm gold nanoparticles. Furthermore, a migration assay demonstrated that in the presence of light, growth of the fungus-like organism Pythium ultimum and the filamentous fungus Botrytis cinerea was inhibited. Atomic Force Microscopy showed significant alterations to the surface of S. epidermidis, and electron microscopy showed cellular aggregates connected by discrete surface linkages. We have therefore demonstrated that the embedded surface has a broad antimicrobial activity under white light and that the surface treatment causes bacterial envelope damage and cell aggregation.

Evaluation of Chlorine Treatment Levels for Inactivation of Human Norovirus and MS2 Bacteriophage during Sewage Treatment.

This study examined the inactivation of human norovirus (HuNoV) GI.1 and GII.4 by chlorine under conditions mimicking sewage treatment. Using a porcine gastric mucin-magnetic bead (PGM-MB) assay, no statistically significant loss in HuNoV binding (inactivation) was observed for secondary effluent treatments of ≤25 ppm total chlorine; for both strains, 50 and 100 ppm treatments resulted in ≤0.8-log10 unit and ≥3.9-log10 unit reductions, respectively. Treatments of 10, 25, 50, and 100 ppm chlorine inactivated 0.31, 1.35, >5, and >5 log10 units, respectively, of the norovirus indicator MS2 bacteriophage. Evaluation of treatment time indicated that the vast majority of MS2 and HuNoV inactivation occurred in the first 5 min for 0.2-µm-filtered, prechlorinated secondary effluent. Free chlorine measurements of secondary effluent seeded with MS2 and HuNoV demonstrated substantial oxidative burdens. With 25, 50, and 100 ppm treatments, free chlorine levels after 5 min of exposure ranged from 0.21 to 0.58 ppm, from 0.28 to 16.7 ppm, and from 11.6 to 53 ppm, respectively. At chlorine treatment levels of >50 ppm, statistically significant differences were observed between reductions for PGM-MB-bound HuNoV (potentially infectious) particles and those for unbound (noninfectious) HuNoV particles or total norovirus particles. While results suggested that MS2 and HuNoV (measured as PGM-MB binding) behave similarly, although not identically, both have limited susceptibility to chlorine treatments of ≤25 ppm total chlorine. Since sewage treatment is performed at ≤25 ppm total chlorine, targeting free chlorine levels of 0.5 to 1.0 ppm, these results suggest that traditional chlorine-based sewage treatment does not inactivate HuNoV efficiently.IMPORTANCE HuNoV is ubiquitous in sewage. A receptor binding assay was used to assess inactivation of HuNoV by chlorine-based sewage treatment, given that the virus cannot be routinely propagated in vitro Results reported here indicate that chlorine treatment of sewage is not effective for inactivating HuNoV unless chlorine levels are above those routinely used for sewage treatment.

Minimizing Bias in Virally Seeded Water Treatment Studies: Evaluation of Optimal Bacteriophage and Mammalian Virus Preparation Methodologies.

One key assumption impacting data quality in viral inactivation studies is that reduction estimates are not altered by the virus seeding process. However, seeding viruses often involves the inadvertent addition of co-constituents such as cell culture components or additives used during preparation steps which can impact viral reduction estimates by inducing non-representative oxidant demand in disinfection studies and fouling in membrane assessments. The objective of this study was therefore to characterize a mammalian norovirus surrogate, murine norovirus (MNV), and bacteriophage MS2 at sequential stages of viral purification and to quantify their potential contribution to artificial oxidant demand and non-representative membrane fouling. Our results demonstrate that seeding solvent extracted and 0.1 micron filtered MNV to ~105 PFU/mL in an experimental water matrix will result in additional total organic carbon (TOC) and 30 min chlorine demand of 39.2 mg/L and 53.5 mg/L as Cl2, respectively. Performing sucrose cushion purification on the MNV stock prior to seeding reduces the impacts of TOC and chlorine demand to 1.6 and 0.15 mg/L as Cl2, respectively. The findings for MNV are likely relevant for other mammalian viruses propagated in serum-based media. Thus, advanced purification of mammalian virus stocks by sucrose cushion purification (or equivalent density-based separation approach) is warranted prior to seeding in water treatment assessments. Studies employing bacteriophage MS2 as a surrogate virus may not need virus purification, since seeding MS2 at a concentration of ~106 PFU/mL will introduce only ~1 mg/L of TOC and ~1 mg/L as Cl2 of chlorine demand to experimental water matrices.

The toxicity of cationic surfactant HDTMA-Br, desorbed from surfactant modified zeolite, towards faecal indicator and environmental microorganisms.

Surfactant Modified Zeolite (SMZ) represents a versatile, cost-effective permeable reactive material, capable of treating multiple classes of contaminants. The potential for HDTMA-Br, a cationic surfactant commonly used to modify zeolite, to desorb from the zeolite surface has been identified as a potential issue for the ongoing use of SMZ in water remediation contexts. This paper investigates the toxicity of HDTMA-Br towards enteric virus surrogates, F-RNA bacteriophage MS2 and E. coli, Bacillus subtilis, and soil microflora. The concentration of surfactant desorbing from SMZ was quantified through a bioassay using E. coli. Results showed HDTMA-Br concentrations of ≥10-5M were toxic to MS2, ≥10-4M were toxic to E. coli and ≥10-6M were toxic to B. subtilis. No toxic relationship was established between HDTMA-Br and soil microflora. Desorption of ≥10-4M of HDTMA-Br was shown for the two SMZ samples under the mixing conditions used. Effects of this surfactant on total soil microflora were ambiguous since no toxic relationship could be established, however, HDTMA-Br, at concentrations desorbing from SMZ, were shown to impact the soil bacterium B. subtilis. Further research is required to determine the effect of this surfactant on microbial populations and species diversity in soils.

MS2 Lysis of Escherichia coli Depends on Host Chaperone DnaJ.

The L protein of the single-stranded RNA phage MS2 causes lysis of Escherichia coli without inducing bacteriolytic activity or inhibiting net peptidoglycan (PG) synthesis. To find host genes required for L-mediated lysis, spontaneous Ill (insensitivity to Llysis) mutants were selected as survivors of L expression and shown to have a missense change of the highly conserved proline (P330Q) in the C-terminal domain of DnaJ. In the dnaJP330Q mutant host, L-mediated lysis is completely blocked at 30°C without affecting the intracellular levels of L. At higher temperatures (37°C and 42°C), both lysis and L accumulation are delayed. The lysis block at 30°C in the dnaJP330Q mutant was recessive and could be suppressed by Lovercomes dnaJ (Lodj ) alleles selected for restoration of lysis. All three Lodj alleles lack the highly basic N-terminal half of the lysis protein and cause lysis ∼20 min earlier than full-length L. DnaJ was found to form a complex with full-length L. This complex was abrogated by the P330Q mutation and was absent with the Lodj truncations. These results suggest that, in the absence of interaction with DnaJ, the N-terminal domain of L interferes with its ability to bind to its unknown target. The lysis retardation and DnaJ chaperone dependency conferred by the nonessential, highly basic N-terminal domain of L resembles the SlyD chaperone dependency conferred by the highly basic C-terminal domain of the E lysis protein of ϕX174, suggesting a common theme where single-gene lysis can be modulated by host factors influenced by physiological conditions.IMPORTANCE Small single-stranded nucleic acid lytic phages (Microviridae and Leviviridae) lyse their host by expressing a single "protein antibiotic." The protein antibiotics from two out of three prototypic small lytic viruses have been shown to inhibit two different steps in the conserved PG biosynthesis pathway. However, the molecular basis of lysis caused by L, the lysis protein of the third prototypic virus, MS2, is unknown. The significance of our research lies in the identification of DnaJ as a chaperone in the MS2 L lysis pathway and the identification of the minimal lytic domain of MS2 L. Additionally, our research highlights the importance of the highly conserved P330 residue in the C-terminal domain of DnaJ for specific protein interactions.

Inactivation modeling of human enteric virus surrogates, MS2, Qß, and ΦX174, in water using UVC-LEDs, a novel disinfecting system.

In order to assure the microbial safety of drinking water, UVC-LED treatment has emerged as a possible technology to replace the use of conventional low pressure (LP) mercury vapor UV lamps. In this investigation, inactivation of Human Enteric Virus (HuEV) surrogates with UVC-LEDs was investigated in a water disinfection system, and kinetic model equations were applied to depict the surviving infectivities of the viruses. MS2, Qß, and ΦX 174 bacteriophages were inoculated into sterile distilled water (DW) and irradiated with UVC-LED printed circuit boards (PCBs) (266nm and 279nm) or conventional LP lamps. Infectivities of bacteriophages were effectively reduced by up to 7-log after 9mJ/cm2 treatment for MS2 and Qß, and 1mJ/cm2 for ΦX 174. UVC-LEDs showed a superior viral inactivation effect compared to conventional LP lamps at the same dose (1mJ/cm2). Non-log linear plot patterns were observed, so that Weibull, Biphasic, Log linear-tail, and Weibull-tail model equations were used to fit the virus survival curves. For MS2 and Qß, Weibull and Biphasic models fit well with R2 values approximately equal to 0.97-0.99, and the Weibull-tail equation accurately described survival of ΦX 174. The level of UV-susceptibility among coliphages measured by the inactivation rate constant, k, was statistically different (ΦX 174 (ssDNA)>MS2, Qß (ssRNA)), and indicated that sensitivity to UV was attributed to viral genetic material.

Influence of solution chemistry on the inactivation of particle-associated viruses by UV irradiation.

MS2 inactivation by UV irradiance was investigated with the focus on how the disinfection efficacy is influenced by bacteriophage MS2 aggregation and adsorption to particles in solutions with different compositions. Kaolinite and Microcystis aeruginosa were used as model inorganic and organic particles, respectively. In the absence of model particles, MS2 aggregates formed in either 1mM NaCl at pH=3 or 50-200mM ionic strength CaCl2 solutions at pH=7 led to a decrease in the MS2 inactivation efficacy because the virions located inside the aggregate were protected from the UV irradiation. In the presence of kaolinite and Microcystis aeruginosa, MS2 adsorbed onto the particles in either 1mM NaCl at pH=3 or 50-200mM CaCl2 solutions at pH=7. In contrast to MS2 aggregates formed without the presence of particles, more MS2 virions adsorbed on these particles were exposed to UV irradiation to allow an increase in MS2 inactivation. In either 1mM NaCl at pH from 4 to 8 or 2-200mM NaCl solutions at pH=7, the absence of MS2 aggregation and adsorption onto the model particles explained why MS2 inactivation was not influenced by pH, ionic strength, and the presence of model particles in these conditions. The influence of virus adsorption and aggregation on the UV disinfection efficiency found in this research suggests the necessity of accounting for particles and cation composition in virus inactivation for drinking water.

Efficient collection of viable virus aerosol through laminar-flow, water-based condensational particle growth.

AIMS: State-of-the-art bioaerosol samplers have poor collection efficiencies for ultrafine virus aerosols. This work evaluated the performance of a novel growth tube collector (GTC), which utilizes laminar-flow water-based condensation to facilitate particle growth, for the collection of airborne MS2 viruses. METHODS AND RESULTS: Fine aerosols (<500 nm) containing MS2 coliphage were generated from a Collison nebulizer, conditioned by a dilution dryer and collected by a GTC and a BioSampler. The GTC effectively condensed water vapour onto the virus particles, creating droplets 2-5 µm in diameter, which facilitated collection. Comparison of particle counts upstream and downstream revealed that the GTC collected >93% of the inlet virus particles, whereas the BioSampler's efficiency was about 10%. Viable counts of the GTC-collected viruses were also one order of magnitude higher than those of the BioSampler (P = 0·003). CONCLUSION: The efficiency of the GTC for the viable collection of MS2 viruses exceeds that of industry standard instrument, the BioSampler, by a factor of 10-100. SIGNIFICANCE AND IMPACT OF THE STUDY: This study reveals that the GTC is an effective collector of viable MS2 aerosols, and concludes the instrument will be an effective tool for studying viable virus aerosols and the inhalation risks posed by airborne viruses.

Enteric Viral Surrogate Reduction by Chitosan.

Enteric viruses are a major problem in the food industry, especially as human noroviruses are the leading cause of nonbacterial gastroenteritis. Chitosan is known to be effective against some enteric viral surrogates, but more detailed studies are needed to determine the precise application variables. The main objective of this work was to determine the effect of increasing chitosan concentration (0.7-1.5% w/v) on the cultivable enteric viral surrogates, feline calicivirus (FCV-F9), murine norovirus (MNV-1), and bacteriophages (MS2 and phiX174) at 37 °C. Two chitosans (53 and 222 kDa) were dissolved in water (53 kDa) or 1% acetic acid (222 KDa) at 0.7-1.5%, and were then mixed with each virus to obtain a titer of ~5 log plaque-forming units (PFU)/mL. These mixtures were incubated for 3 h at 37 °C. Controls included untreated viruses in phosphate-buffered saline and viruses were enumerated by plaque assays. The 53 kDa chitosan at the concentrations tested reduced FCV-F9, MNV-1, MS2, and phi X174 by 2.6-2.9, 0.1-0.4, 2.6-2.8, and 0.7-0.9 log PFU/mL, respectively, while reduction by 222 kDa chitosan was 2.2-2.4, 0.8-1.0, 2.6-5.2, and 0.5-0.8 log PFU/mL, respectively. The 222 kDa chitosan at 1 and 0.7% w/v in acetic acid (pH 4.5) caused the greatest reductions of MS2 by 5.2 logs and 2.6 logs, respectively. Overall, chitosan treatments showed the greatest reduction of MS2, followed by FCV-F9, phi X174, and MNV-1. These two chitosans may contribute to the reduction of enteric viruses at the concentrations tested but would require use of other hurdles to eliminate food borne viruses.

Use of a Hand Sanitizing Wipe for Reducing Risk of Viral Illness in the Home.

This study determined whether a hand sanitizing wipe can reduce virus transmission in households, and could reduce the probability of infection by rhinovirus and rotavirus. Bacteriophage MS-2 (a marker virus) was used to assess viral transmission in five households having at least two children of ages 2-18. Hands of one female adult were inoculated with ~10(8) PFU MS-2 bacteriophages in each home, and after 8 h, hands of all family members and select fomites were sampled to determine baseline contamination without intervention. This sequence was repeated with the intervention, where all family members were instructed to use a quaternary ammonium compound-based sanitizing wipe at least once per day. A significant reduction of virus after the intervention occurred on inoculated hands (95.3%; p = 0.0039), all fomites combined (74.5%; p < 0.005), and non-inoculated hands and fomites combined (73.5%; p < 0.005). However, viral reduction on non-inoculated hands was not significant, likely due to small sample size. Using rhinovirus and rotavirus as models it was estimated that infection risk was reduced by ~30 to 89% with the use of sanitizing wipes once per day depending on the starting concentration of these viruses on hands of susceptible individuals. Therefore, using a hand sanitizing wipe can significantly reduce viral transmission and risk of illness in homes. Previous studies have shown other hand hygiene interventions, such as alcohol-based hand sanitizers, are even more effective for reducing risk of illness in homes; however the sanitizing wipe used in this study is appropriate to use for microbial reduction.

Aerosol-phase activity of iodine captured from a triiodide resin filter on fine particles containing an infectious virus.

AIMS: To avoid interference by water-iodine disinfection chemistry and measure directly the effect of iodine, captured from a triiodide complex bound to a filter medium, on viability of penetrating viral particles. METHODS AND RESULTS: Aerosols of MS2 coli phage were passed through control P100 or iodinated High-Efficiency Particulate Air media, collected in plastic bags, incubated for 0-10 min, collected in an impinger containing thiosulphate to consume all unreacted iodine, plated and enumerated. Comparison of viable counts demonstrated antimicrobial activity with an apparent half-life for devitalization in tens of seconds; rate of kill decreased at low humidity and free iodine was captured by the bags. CONCLUSIONS: The results support the mechanism of near-contact capture earlier proposed; however, the disinfection chemistry in the aerosol phase is very slow on the time scale of inhalation. SIGNIFICANCE AND IMPACT OF THE STUDY: This study shows that disinfection by filter-bound iodine in the aerosol phase is too slow to be clinically significant in individual respiratory protection, but that it might be of benefit to limit airborne transmission of infections in enclosed areas.

A compact point-of-use water purification cartridge for household use in developing countries.

Simple, low-cost household interventions are known to be effective in lowering the incidence of waterborne diseases in developing countries. However, high costs along with operational and maintenance issues have prevented the successful adoption of these interventions among the affected communities. To address these limitations, a cost-effective, gravity-driven water purification cartridge has been developed by employing the synergistic disinfection action of low concentrations of silver and chlorine on bacteria and viruses. The silver and chlorine treatment components within the cartridge have been developed using inexpensive materials and integrated with a life indicator and auto-shut-off-mechanism within a compact form factor. The antibacterial as well as antiviral performance of the cartridge was tested by using ground water spiked with Escherichia coli and MS2 bacteriophage. The results show that, although individually, the silver and chlorine treatment systems were unable to inactivate the test strains, the integrated cartridge inactivates both bacteria as well as viruses up to the log reduction requirement of the USEPA guide standard for microbiological water purifiers over its designated life of 2,000 liters.

Comparison of high-volume air sampling equipment for viral aerosol sampling during emergency response.

OBJECTIVE: This study compared the performance of two high-volume bioaerosol air samplers for viable virus to an accepted standard low-volume sampler. In typical bioaerosol emergency response scenarios, highvolume sampling is essential for the low infective concentrations and large air volumes involved. DESIGN: Two high-volume air samplers (XMX/2LMIL and DFU-1000) were evaluated alongside a lowvolume sample (BioSampler). Low and high concentrations (9.3-93.2 agent containing particles per liter of air [ACPLA]) of male-specific coliphage 2 (MS2) virus were released into a 12 m3 aerosol test chamber and collected using the air samplers. The collection media from the samplers were then processed and viable virus was assessed via plaque assay. SETTING: Aerosol test chamber. SUBJECTS, PARTICIPANTS: None. INTERVENTIONS: Collection media and flow rate were modified for the XMX/2L-MIL sampler for viable analysis. MAIN OUTCOME MEASURES: Concentration estimates in units of plaque forming units per liter of air (PFU/liter) assessed by the samplers as compared to the levels inside the chamber as evaluated with a slit to agar plate in units of ACPLA. Comparison was made via one-way analysis of variance. RESULTS: Both the XMX/2L-MIL and DFU-1000 achieved collection effectiveness equal to or greater than the low-volume air sampler for the evaluated MS2 concentrations. The XMX/2L-MIL reliably collected quantifiable low concentrations of MS2, but the DFU-1000 was unable to do so. CONCLUSIONS: For emergency response to suspected bioaerosols, the evaluated high-volume samplers are as effective as the standard low-flow sampler and should be considered in conducting a health risk assessment. If low concentrations are expected, then high-flow samplers using liquid collection are preferred.

Survival of airborne MS2 bacteriophage generated from human saliva, artificial saliva, and cell culture medium.

Laboratory studies of virus aerosols have been criticized for generating airborne viruses from artificial nebulizer suspensions (e.g., cell culture media), which do not mimic the natural release of viruses (e.g., from human saliva). The objectives of this study were to determine the effect of human saliva on the infectivity and survival of airborne virus and to compare it with those of artificial saliva and cell culture medium. A stock of MS2 bacteriophage was diluted in one of three nebulizer suspensions, aerosolized, size selected (100 to 450 nm) using a differential mobility analyzer, and collected onto gelatin filters. Uranine was used as a particle tracer. The resulting particle size distribution was measured using a scanning mobility particle sizer. The amounts of infectious virus, total virus, and fluorescence in the collected samples were determined by infectivity assays, quantitative reverse transcription-PCR (RT-PCR), and spectrofluorometry, respectively. For all nebulizer suspensions, the virus content generally followed a particle volume distribution rather than a number distribution. The survival of airborne MS2 was independent of particle size but was strongly affected by the type of nebulizer suspension. Human saliva was found to be much less protective than cell culture medium (i.e., 3% tryptic soy broth) and artificial saliva. These results indicate the need for caution when extrapolating laboratory results, which often use artificial nebulizer suspensions. To better assess the risk of airborne transmission of viral diseases in real-life situations, the use of natural suspensions such as saliva or respiratory mucus is recommended.

High-throughput assay for temporal kinetic analysis of lytic coliphage activity.

Plaque analysis allows the determination of phage titer and multiplicity of infection. Yet, this overnight assay provides only endpoint results, ignoring kinetic aspects. We introduce an alternative high-throughput and rapid method for kinetic analysis of lytic coliphage activity. Escherichia coli was infected with serial dilutions of MS2 coliphage, and bacterial growth was monitored using a multi-well plate reader providing within hours the equivalent data as obtained overnight. Additional information is yielded, including phage replication rate, progeny size per cycle, and viral propagation during bacterial growth. This method offers further insights into physicochemical mechanisms of lytic coliphage infection and temporal control. It also provides a virus-host interaction acumen.