Infection-competent monkeypox virus contamination identified in domestic settings following an imported case of monkeypox into the UK.

An imported case of monkeypox was diagnosed in December 2019 in a traveller returning from Nigeria to the UK. Subsequently, environmental sampling was performed at two adjoining single-room residences occupied by the patient and their sibling. Monkeypox virus DNA was identified in multiple locations throughout both properties, and monkeypox virus was isolated from several samples 3 days after the patient was last in these locations. Positive samples were identified following the use of both vacuum and surface sampling techniques; these methodologies allowed for environmental analysis of potentially contaminated porous and non-porous surfaces via real-time quantitative reverse transcriptase PCR analysis in addition to viral isolation to confirm the presence of infection-competent virus. This report confirms the potential for infection-competent monkeypox virus to be recovered in environmental settings associated with known positive cases and the necessity for rapid environmental assessment to reduce potential exposure to close contacts and the general public. The methods adopted in this investigation may be used for future confirmed cases of monkeypox in order to establish levels of contamination, confirm the presence of infection-competent material and to identify locations requiring additional cleaning.

Characterisation of Particle Size and Viability of SARS-CoV-2 Aerosols from a Range of Nebuliser Types Using a Novel Sampling Technique.

Little is understood about the impact of nebulisation on the viability of SARS-CoV-2. In this study, a range of nebulisers with differing methods of aerosol generation were evaluated to determine SARS-CoV-2 viability following aerosolization. The aerosol particle size distribution was assessed using an aerosol particle sizer (APS) and SARS-CoV-2 viability was determined after collection into liquid media using All-Glass Impingers (AGI). Viable particles of SARS-CoV-2 were further characterised using the Collison 6-jet nebuliser in conjunction with novel sample techniques in an Andersen size-fractioning sampler to predict lung deposition profiles. Results demonstrate that all the tested nebulisers can generate stable, polydisperse aerosols (Geometric standard deviation (GSD) circa 1.8) in the respirable range (1.2 to 2.2 µm). Viable fractions (VF, units PFU/particle, the virus viability as a function of total particles produced) were circa 5 × 10-3. VF and spray factors were not significantly affected by relative humidity, within this system where aerosols were in the spray tube an extremely short time. The novel Andersen sample collection methods successfully captured viable virus particles across all sizes; with most particle sizes below 3.3 µm. Particle sizes, in MMAD (Mass Median Aerodynamic Diameters), were calculated from linear regression of log10-log10 transformed cumulative PFU data, and calculated MMADs accorded well with APS measurements and did not differ across collection method types. These data will be vital in informing animal aerosol challenge models, and infection prevention and control policies.

Persistence of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Virus and Viral RNA in Relation to Surface Type and Contamination Concentration.

The transmission of SARS-CoV-2 is likely to occur through a number of routes, including contact with contaminated surfaces. Many studies have used reverse transcription-PCR (RT-PCR) analysis to detect SARS-CoV-2 RNA on surfaces, but seldom has viable virus been detected. This paper investigates the viability over time of SARS-CoV-2 dried onto a range of materials and compares viability of the virus to RNA copies recovered and whether virus viability is concentration dependent. Viable virus persisted for the longest time on surgical mask material and stainless steel, with a 99.9% reduction in viability by 122 and 114 h, respectively. Viability of SARS-CoV-2 reduced the fastest on a polyester shirt, with a 99.9% reduction within 2.5 h. Viability on the bank note was reduced second fastest, with 99.9% reduction in 75 h. RNA on all surfaces exhibited a 1-log reduction in genome copy number recovery over 21 days. The findings show that SARS-CoV-2 is most stable on nonporous hydrophobic surfaces. RNA is highly stable when dried on surfaces, with only 1-log reduction in recovery over 3 weeks. In comparison, SARS-CoV-2 viability reduced more rapidly, but this loss in viability was found to be independent of starting concentration. Expected levels of SARS-CoV-2 viable environmental surface contamination would lead to undetectable levels within 2 days. Therefore, when RNA is detected on surfaces, it does not directly indicate the presence of viable virus, even at low cycle threshold values. IMPORTANCE This study shows the impact of material type on the viability of SARS-CoV-2 on surfaces. It demonstrates that the decay rate of viable SARS-CoV-2 is independent of starting concentration. However, RNA shows high stability on surfaces over extended periods. This has implications for interpretation of surface sampling results using RT-PCR to determine the possibility of viable virus from a surface, where RT-PCR is not an appropriate technique to determine viable virus. Unless sampled immediately after contamination, it is difficult to align RNA copy numbers to quantity of viable virus on a surface.

Evolution of 2009 H1N1 influenza viruses during the pandemic correlates with increased viral pathogenicity and transmissibility in the ferret model.

There is increasing evidence that 2009 pandemic H1N1 influenza viruses have evolved after pandemic onset giving rise to severe epidemics in subsequent waves. However, it still remains unclear which viral determinants might have contributed to disease severity after pandemic initiation. Here, we show that distinct mutations in the 2009 pandemic H1N1 virus genome have occurred with increased frequency after pandemic declaration. Among those, a mutation in the viral hemagglutinin was identified that increases 2009 pandemic H1N1 virus binding to human-like α2,6-linked sialic acids. Moreover, these mutations conferred increased viral replication in the respiratory tract and elevated respiratory droplet transmission between ferrets. Thus, our data show that 2009 H1N1 influenza viruses have evolved after pandemic onset giving rise to novel virus variants that enhance viral replicative fitness and respiratory droplet transmission in a mammalian animal model. These findings might help to improve surveillance efforts to assess the pandemic risk by emerging influenza viruses.

Biofilm formation in an experimental water distribution system: the contamination of non-touch sensor taps and the implication for healthcare.

Hospital tap water is a recognised source of Pseudomonas aeruginosa. U.K. guidance documents recommend measures to control/minimise the risk of P. aeruginosa in augmented care units but these are based on limited scientific evidence. An experimental water distribution system was designed to investigate colonisation of hospital tap components. P. aeruginosa was injected into 27 individual tap 'assemblies'. Taps were subsequently flushed twice daily and contamination levels monitored over two years. Tap assemblies were systematically dismantled and assessed microbiologically and the effect of removing potentially contaminated components was determined. P. aeruginosa was repeatedly recovered from the tap water at levels above the augmented care alert level. The organism was recovered from all dismantled solenoid valves with colonisation of the ethylene propylene diene monomer (EPDM) diaphragm confirmed by microscopy. Removing the solenoid valves reduced P. aeruginosa counts in the water to below detectable levels. This effect was immediate and sustained, implicating the solenoid diaphragm as the primary contamination source.

Comparative investigations for adenovirus recognition and quantification: Plastic or natural antibodies?

Comparative and comprehensive investigations for adenovirus recognition and detection were conducted using plastic and natural antibodies to compare three different strategies. The implementation of molecularly imprinted polymer (MIP) technology for specific and sensitive recognition of viruses with the combination of biosensors was reported. Plastic antibodies (MIPs nanoparticles) were produced for adenovirus by employing a novel solid phase synthesis method. MIP receptors were then characterised using dynamic light scattering (DLS) and transmission electron microscopy (TEM) techniques prior to immobilisation on a surface plasmon resonance (SPR) sensor as affinity receptor for adenovirus detection. Two different templates were also imprinted as control MIPs (vancomycin-MIP and MS2-MIP). The specific recognition of adenovirus was investigated in the concentration range of 0.01-20 pM and the limit of detection was achieved as 0.02 pM. As an alternative to MIP receptors, direct and sandwich assays were developed for adenovirus quantification using natural antibodies. The detection limit of direct and sandwich assays were found as 0.3 pM and 0.008 pM, respectively. The kinetic data analyses were performed for three different adenovirus recognition methods and cross-reactivity studies were also conducted using MS2 phage as control virus and an excellent specificity was achieved with all assays types. This work confirmed the suitability of the MIPs SPR sensor for the detection of viruses.

Detection of Waterborne Viruses Using High Affinity Molecularly Imprinted Polymers.

Molecularly imprinted polymers (MIPs) are artificial receptor ligands which can recognize and specifically bind to a target molecule. They are more resistant to chemical and biological damage and inactivation than antibodies. Therefore, target specific-MIP nanoparticles are aimed to develop and implemented to biosensors for the detection of biological toxic agents such as viruses, bacteria, and fungi toxins that cause many diseases and death due to the environmental contamination. For the first time, a molecularly imprinted polymer (MIP) targeting the bacteriophage MS2 as the template was investigated using a novel solid-phase synthesis method to obtain the artificial affinity ligand for the detection and removal of waterborne viruses through optical-based sensors. A high affinity between the artificial ligand and the target was found, and a regenerative MIP-based virus detection assay was successfully developed using a new surface plasmon resonance (SPR)-biosensor which provides an alternative technology for the specific detection and removal of waterborne viruses that lead to high disease and death rates all over the world.

Reaerosolization of Spores from Flooring Surfaces To Assess the Risk of Dissemination and Transmission of Infections.

The aim of this study was to quantify reaerosolization of microorganisms caused by walking on contaminated flooring to assess the risk to individuals accessing areas contaminated with pathogenic organisms, for example, spores of Bacillus anthracis. Industrial carpet and polyvinyl chloride (PVC) floor coverings were contaminated with aerosolized spores of Bacillus atrophaeus by using an artist airbrush to produce deposition of ∼10(3) to 10(4) CFU · cm(-2). Microbiological air samplers were used to quantify the particle size distribution of the aerosol generated when a person walked over the floorings in an environmental chamber. Results were expressed as reaerosolization factors (percent per square centimeter per liter), to represent the ratio of air concentration to surface concentration generated. Walking on carpet generated a statistically significantly higher reaerosolization factor value than did walking on PVC (t = 20.42; P < 0.001). Heavier walking produced a statistically significantly higher reaerosolization factor value than did lighter walking (t = 12.421; P < 0.001). Height also had a statistically significant effect on the reaerosolization factor, with higher rates of recovery of B. atrophaeus at lower levels, demonstrating a height-dependent gradient of particle reaerosolization. Particles in the respirable size range were recovered in all sampling scenarios (mass mean diameters ranged from 2.6 to 4.1 µm). The results of this study can be used to produce a risk assessment of the potential aerosol exposure of a person accessing areas with contaminated flooring in order to inform the choice of appropriate respiratory protective equipment and may aid in the selection of the most suitable flooring types for use in health care environments, to reduce aerosol transmission in the event of contamination.

Staphylococcus aureus dispersal from healthy volunteers.

BACKGROUND: Understanding Staphylococcus aureus dispersal from human carriers is vital for preventing transmission and colonization of this organism in health care settings. This study investigated the S aureus supershedder hypothesis in relation to attributes of healthy volunteers. METHODS: Microbial aerosol generation from volunteers was quantified within a controlled environmental chamber during walking or sitting activities. Biological air samplers were used to determine numbers of total S aureus colony-forming units disseminated during these activities. RESULTS: A total of 17 volunteers was sampled on 3 occasions. Hairstyle (long hair tied up or a shaved head) was the only significant predictor of dissemination of S aureus (5% significance level). No other significant effects were found at the 5% level. A negative binomial distribution provides the best fit with respect to S aureus. CONCLUSION: We found that, in the context of our small sample size, hairstyle (long hair tied up or a shaved head) statistically affected levels of bacteria shed from volunteers. However, we found no evidence for "supershedders" or "cloud adults," suggesting they are at an extreme end of a continuous distribution.

Testing the efficacy of homemade masks: would they protect in an influenza pandemic?

OBJECTIVE: This study examined homemade masks as an alternative to commercial face masks. METHODS: Several household materials were evaluated for the capacity to block bacterial and viral aerosols. Twenty-one healthy volunteers made their own face masks from cotton t-shirts; the masks were then tested for fit. The number of microorganisms isolated from coughs of healthy volunteers wearing their homemade mask, a surgical mask, or no mask was compared using several air-sampling techniques. RESULTS: The median-fit factor of the homemade masks was one-half that of the surgical masks. Both masks significantly reduced the number of microorganisms expelled by volunteers, although the surgical mask was 3 times more effective in blocking transmission than the homemade mask. CONCLUSION: Our findings suggest that a homemade mask should only be considered as a last resort to prevent droplet transmission from infected individuals, but it would be better than no protection.

Influenza aerosols in UK hospitals during the H1N1 (2009) pandemic--the risk of aerosol generation during medical procedures.

BACKGROUND: Nosocomial infection of health-care workers (HCWs) during outbreaks of respiratory infections (e.g. Influenza A H1N1 (2009)) is a significant concern for public health policy makers. World Health Organization (WHO)-defined 'aerosol generating procedures' (AGPs) are thought to increase the risk of aerosol transmission to HCWs, but there are presently insufficient data to quantify risk accurately or establish a hierarchy of risk-prone procedures. METHODOLOGY/PRINCIPAL FINDINGS: This study measured the amount of H1N1 (2009) RNA in aerosols in the vicinity of H1N1 positive patients undergoing AGPs to help quantify the potential risk of transmission to HCWs. There were 99 sampling occasions (windows) producing a total of 198 May stages for analysis in the size ranges 0.86-7.3 µm. Considering stages 2 (4-7.3 µm) and 3 (0.86-4 µm) as comprising one sample, viral RNA was detected in 14 (14.1%) air samples from 10 (25.6%) patients. Twenty three air samples were collected while potential AGPs were being performed of which 6 (26.1%) contained viral RNA; in contrast, 76 May samples were collected when no WHO 2009 defined AGP was being performed of which 8 (10.5%) contained viral RNA (unadjusted OR = 2.84 (95% CI 1.11-7.24) adjusted OR = 4.31 (0.83-22.5)). CONCLUSIONS/SIGNIFICANCE: With our small sample size we found that AGPs do not significantly increase the probability of sampling an H1N1 (2009) positive aerosol (OR (95% CI) = 4.31 (0.83-22.5). Although the probability of detecting positive H1N1 (2009) positive aerosols when performing various AGPs on intensive care patients above the baseline rate (i.e. in the absence of AGPs) did not reach significance, there was a trend towards hierarchy of AGPs, placing bronchoscopy and respiratory and airway suctioning above baseline (background) values. Further, larger studies are required but these preliminary findings may be of benefit to infection control teams.

A nationwide survey of the quality of antimalarials in retail outlets in Tanzania.

INTRODUCTION: Retail pharmaceutical products are commonly used to treat fever and malaria in sub-Saharan African countries. Small scale studies have suggested that poor quality antimalarials are widespread throughout the region, but nationwide data are not available that could lead to generalizable conclusions about the extent to which poor quality drugs are available in African communities. This study aimed to assess the quality of antimalarials available from retail outlets across mainland Tanzania. METHODS AND FINDINGS: We systematically purchased samples of oral antimalarial tablets from retail outlets across 21 districts in mainland Tanzania in 2005. A total of 1080 antimalarial formulations were collected including 679 antifol antimalarial samples (394 sulfadoxine/pyrimethamine and 285 sulfamethoxypyrazine/pyrimethamine), 260 amodiaquine samples, 63 quinine samples, and 51 artemisinin derivative samples. A systematic subsample of 304 products was assessed for quality by laboratory based analysis to determine the amount of the active ingredient and dissolution profile by following the published United States Pharmacopoeia (USP) monogram for the particular tablet being tested. Products for which a published analytical monogram did not exist were assessed on amount of active ingredient alone. Overall 38 or 12.2% of the samples were found to be of poor quality. Of the antifolate antimalarial drugs tested 13.4% were found to be of poor quality by dissolution and content analysis using high-performance liquid chromatography (HPLC). Nearly one quarter (23.8%) of quinine tablets did not comply within the tolerance limits of the dissolution and quantification analysis. Quality of amodiaquine drugs was relatively better but still unacceptable as 7.5% did not comply within the tolerance limits of the dissolution analysis. Formulations of the artemisinin derivatives all contained the stated amount of active ingredient when analysed using HPLC alone. CONCLUSIONS: Substandard antimalarial formulations were widely available in Tanzania at the time of this study. No products were detected that did not contain any amount of the stated active ingredient. Quinine and sulfadoxine/pyrimethamine products were the most widely available and also the most likely to be of poor quality. Substandard products were identified in all parts of the country and were labeled as made by both domestic and international manufacturers. With the expansion of the retail pharmaceutical sector as a delivery channel for antimalarial formulations the need for regular nationwide monitoring of their quality will become increasingly important.

Anti-trypanosomal activities of DNA topoisomerase inhibitors.

Only four drugs are available for chemotherapy of human African sleeping sickness with undesirable toxic side effects. The development of new anti-trypanosomal drugs is, therefore, urgently required. In this study, 15 DNA topoisomerase inhibitors, including approved anti-cancer drugs, were tested for in vitro activity against bloodstream forms of Trypanosoma brucei and human leukaemia HL-60 cells. All compounds exhibited anti-trypanosomal activity, with ED50 values ranging between 3 nM and 30 microM, and MIC values between 100 nM and >100 microM. The trypanocidal activities of the most effective DNA topoisomerase inhibitors, aclarubicin, doxorubicin and mitoxantrone, were comparable with those of commercial anti-trypanosomal drugs. These data support the use of DNA topoisomerase inhibitors as lead compounds for anti-trypanosomal drug development.