In vitro antiviral effect of Mexican and Brazilian propolis and phenolic compounds against human coronavirus 229E.

Propolis is a resinous substance collected by bees (Apis mellifera). It is used for its biological properties. This natural product is available as a safe therapeutic option. Herein, we report the antiviral effects of brown propolis extract from Mexico and green and red propolis extracts from Brazil, as well as their phenolic compounds (quercetin, caffeic acid, and rutin) in preventing infection of MRC-5 cells by HCoV-229E. Normal human fibroblast lung cells (MRC-5) were used to determine the cytotoxicity of the compounds. All samples studied showed antiviral activity. Green and brown propolis extracts, and quercetin exhibited the best EC50 values with values of 19.080, 11.240, and 77.208 µg/mL against HCoV-229E, respectively, and with TC50 of 62.19, 29.192, and 298 µg/mL on MRC-5 cells, respectively. These results are the first in vitro study of the effects of propolis on HCoV-229E and provide the basis for the development of natural formulations against other coronavirus strains.

Inactivation kinetics of benzalkonium chloride and ethanol-based hand sanitizers against a betacoronavirus and an alphacoronavirus.

Background: Hand hygiene is critical to lower the potential for the spread of SARS-CoV-2 and other infectious agents by direct contact. When running water and soap are not available for hand hygiene, ethanol-based hand sanitizers are currently the recommended standard of care [1-3]. Though recently published data showed comparable in vitro effectiveness of benzalkonium chloride (BAK)-based and ethanol-based hand sanitizers against SARS-CoV-2 virus, a paucity of peer-reviewed data on the effectiveness of these formulations against other types of infective coronaviruses remains. This work assessed human coronavirus HCoV-229E (genus Alphacoronavirus) concurrently with SARS-CoV-2, Isolate USA-WA1/2020 (genus Betacoronavirus) to fill this gap. Methods: The test was conducted according to EN14476:2013-A2:2019 [EN14476] Quantitative Suspension Test for the Evaluation of Virucidal Activity in the Medical Area [4]. Two BAK-based hand sanitizers, five ethanol-based hand sanitizers, and an 80% ethanol reference formulation were tested for antiviral activity against SARS-CoV-2 and HCoV-229E at 15- and 30- second contact times. Results: Both SARS-CoV-2 and HCoV-229E were reduced by greater than 4.00-log10 within 15 seconds of contact. Virus decay constants (k) following first-order kinetics were similar for BAK and ethanol-based formulations against both test viruses. The SARS-CoV-2 results reported herein mirrored previous data reported by Herdt et al. (2021). Conclusion: BAK and ethanol hand sanitizer formulations inactivate SARS-CoV-2 and HCoV-229E at similar rates. This data supports previously published effectiveness data for both chemistries and indicates that additional coronavirus strains and variants would demonstrate similar inactivation trends.

Inactivation of SARS-CoV-2 in Water by Chlorination.

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) is present in both respiratory secretions and feces, creating its potential for transmission by swimming pools. Recreational water activity is known to be at increased risk of respiratory infections and respiratory viruses have caused been detected and have caused outbreaks in swimming pools. However, little is known regarding the chlorine inactivation of SARS-CoV-2 in water typical of swimming pools in the USA. In this study, the inactivation of SARS-CoV-2 Isolate hCoV-19/USA-WA1/2020 was observed in water by chlorination. All experiments were conducted within a BSL-3 laboratory at room temperature. Our results show that the virus was reduced by 3.5 log (> 99.9%) after 30 s of 2.05-mg/L free chlorine contact and greater than 4.17 log (limit of detection) (> 99.99%) within 2 min.

Survivability of Delta and Omicron variants of SARS-CoV-2 in wastewater.

Concerns of fecal-aerosol transmission of coronavirus disease 2019 (COVID-2019) coupled with increased transmissibility and disease severity of Delta and Omicron variants of concern (VOC) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), suggest studies on survival of VOC in wastewater are warranted. To the best of our knowledge, this is the first study to investigate the survivability of Delta and Omicron VOC in filtered and unfiltered raw wastewater, and secondary effluent at room temperature (23 °C). The time required for 90 % inactivation (T90) of Delta and Omicron VOC in unfiltered raw wastewater was calculated as 17.7 and 15.3 h, respectively. Rapid inactivation of VOC in wastewater and inability to isolate SARS-CoV-2 in wastewater suggest risks from fecal-aerosol transmission are low. Nevertheless, high transmissibility of VOC cautions overruling fecal-aerosol transmission of COVID-19. Future studies on survival of SARS-CoV-2 in wastewater should attempt viral culture by spiking feces collected from COVID-19 infected patients into wastewater to match the real-world scenario.

Survival of SARS-CoV-2 in wastewater.

The ongoing pandemic of Coronavirus disease 2019 (COVID-19) has affected >600 million people with >6 million deaths. Although Severe Acute Respiratory Syndrome-Coronavirus-2 (SARS-CoV-2), the etiologic agent of COVID-19, is transmitted via respiratory droplets or direct contact, isolation of viable SARS-CoV-2 in feces has been reported. Therefore, there is a need for understanding the persistence of SARS-CoV-2 and emerging variants in wastewater. In this study, the survival of SARS-CoV-2 isolate hCoV-19/USA-WA1/2020 was observed in three wastewater matrices - filtered and unfiltered raw wastewater, and secondary effluent. All experiments were conducted within a BSL-3 laboratory at room temperature. The time required for inactivation of 90 % (T90) of SARS-CoV-2 was 10.4, 10.8, and 18.3 h for unfiltered raw, filtered raw, and secondary effluent, respectively. Progressive decline in infectivity of the virus following first order kinetics was noted in these wastewater matrices. To the best of our knowledge, this is the first study to describe the survival of SARS-CoV-2 in secondary effluent.

Virus rejection and removal in pilot-scale air-gap membrane distillation.

Membrane distillation (MD) is a thermally-driven process that can treat high concentration streams and provide a dual barrier for rejection and reduction of pathogens. Thus, MD has potential applications in treating concentrated wastewater brines for enhancing water recovery and potable water reuse. In bench-scale studies, it was demonstrated that MD can provide high rejection of MS2 and PhiX174 bacteriophage viruses, and when operating at temperatures greater than 55 °C, can reduce virus levels in the concentrate. However, bench-scale MD results cannot directly be used to predict pilot-scale contaminant rejection and removal of viruses because of the lower water flux and higher transmembrane hydraulic pressure difference in pilot-scale systems. Thus far, virus rejection and removal have not been quantified in pilot-scale MD systems. In this work, the rejection of MS2 and PhiX174 at low (40 °C) and high (70 °C) inlet temperatures is quantified in a pilot-scale air-gap MD system using tertiary treated wastewater. Both viruses were detected in the distillate which suggests the presence of pore flow; the virus rejection at a hot inlet temperature of 40 °C for MS2 and PhiX174 were 1.6-log10 and 3.1-log10, respectively. At 70 °C, virus concentrations in the brine decreased and were below the detection limit (1 PFU per 100 mL) after 4.5 h, however, viruses were also detected in the distillate in that duration. Results demonstrate that virus rejection is lower in pilot-scale experiments because of increased pore flow that is not captured in bench-scale experiments.

Transfer efficiency of an enveloped virus, human coronavirus 229E, from various hard surface fomites to finger pads of the hands.

Respiratory viruses can be transmitted by fomite contact, but no data currently exist on the transfer of enveloped viruses. The transfer efficiency of human coronavirus from various hard surfaces ranged from 0.46% to 49.0%. This information can be used to model the fomite transmission of enveloped viruses.

Continuously active disinfectant inactivates severe acute respiratory coronavirus virus 2 (SARS-CoV-2) and human coronavirus 229E two days after the disinfectant was applied and following wear exposures.

The surface environment in rooms of coronavirus disease 2019 (COVID-19) patients may be persistently contaminated despite disinfection. A continuously active disinfectant demonstrated excellent sustained antiviral activity following a 48-hour period of wear and abrasion exposures with reinoculations. Reductions of >4-log10 were achieved within a 1-minute contact time for severe acute respiratory coronavirus virus 2 (SARS-CoV-2) and the human coronavirus, 229E.

New method using a positively charged microporous filter and ultrafiltration for concentration of viruses from tap water.

The methods used to concentrate enteric viruses from water have remained largely unchanged for nearly 30 years, with the most common technique being the use of 1MDS Virozorb filters followed by organic flocculation for secondary concentration. Recently, a few studies have investigated alternatives; however, many of these methods are impractical for use in the field or share some of the limitations of this traditional method. In the present study, the NanoCeram virus sampler, an electropositive pleated microporous filter composed of microglass filaments coated with nanoalumina fibers, was evaluated. Test viruses were first concentrated by passage of 20 liters of seeded water through the filter (average filter retention efficiency was ≥ 99.8%), and then the viruses were recovered using various salt-based or proteinaceous eluting solutions. A 1.0% sodium polyphosphate solution with 0.05 M glycine was determined to be the most effective. The recovered viruses were then further concentrated using Centricon Plus-70 centrifugal ultrafilters to a final volume of 3.3 (±0.3 [standard deviation]) ml; this volume compares quite favorably to that of previously described methods, such as organic flocculation (~15 to 40 ml). The overall virus recovery efficiencies were 66% for poliovirus 1, 83% for echovirus 1, 77% for coxsackievirus B5, 14% for adenovirus 2, and 56% for MS2 coliphage. In addition, this method appears to be compatible with both cell culture and PCR assays. This new approach for the recovery of viruses from water is therefore a viable alternative to currently used methods when small volumes of final concentrate are an advantage.

Efficacy of an antimicrobial surface coating against human coronavirus 229E and SARS-CoV-2.

The COVID-19 pandemic has accelerated the demand for alternatives to standard cleaning and disinfection practices. Antiviral coatingsmay provide an alternative to common surface treatments. A newly developed quaternary ammonium polymer coating was applied to stainless steel coupons and evaluated for efficacy against human coronavirus 229E and SARS-CoV-2. The polymer coating reduced levels of both test viruses by greater than 99.9% relative to non-coated stainless steel coupons during a 2-hour contact time.

Membrane Distillation Provides a Dual Barrier for Coronavirus and Bacteriophage Removal.

The persistence of pathogenic microorganisms in treated wastewater effluent makes disinfection crucial to achieve wastewater reuse. Membrane processes such as ultrafiltration and reverse osmosis (RO) have shown promising results for virus and other contaminant removal from treated wastewater effluents for reuse application. However, RO produces a concentrate stream which contains high concentrations of pathogens and contaminants that often requires treatment and volume reduction before disposal. Membrane distillation (MD) is a treatment process that can reduce RO concentrate volume while augmenting the potable water supply. MD is also a dual barrier approach for virus removal as it operates at a high temperature and permeates only the vapor phase through the membrane interface. The effects of temperature on viable virus concentration and membrane rejection of viruses in MD are investigated in this study using two nonenveloped phages frequently used as enteric virus surrogates (MS2 and PhiX174) and an enveloped pathogenic virus (HCoV-229E). At typical MD operating temperatures (greater than 65 °C), viable concentrations of all three viruses were reduced by thermal inactivation by more than 6-log10 for MS2 and PhiX174 and more than 3-log10 for HCoV-229E. Also, membrane rejection was greater than 6-log10 for MS2 and PhiX174 and greater than 2.5-log10 for HCoV-229E.

Investigation of the effects of an ultraviolet germicidal irradiation system on concentrations of aerosolized surrogates for common veterinary pathogens.

OBJECTIVE: To determine whether exposure to UV germicidal irradiation (UVGI) reduces concentrations of viable aerosolized microorganisms (attenuated strains of common veterinary pathogens) in a simulated heating, ventilation, and air conditioning (HVAC) system. SAMPLE: 42 air samples seeded with bacteriophage MS2 or attenuated strains of Bordetella bronchiseptica, feline calicivirus, feline herpesvirus-1, canine parvovirus, or canine distemper virus (6/microorganism) or with no microorganisms added (6). PROCEDURES: A simulated HVAC unit was built that included a nebulizer to aerosolize microorganisms suspended in phosphate-buffered water, a fan to produce airflow, 2 UVGI bulb systems, and an impinger for air sampling. Ten-minute trials (3 with UVGI, 3 without UVGI, and 1 negative control) were conducted for each microorganism. Impingers collected microorganisms into phosphate-buffered water for subsequent quantification with culture-based assays. Results for samples yielding no target microorganisms were recorded as the assay's lower limit of detection. Statistical analysis was not performed. RESULTS: The UVGI treatment resulted in subjectively lower concentrations of viable MS2, B bronchiseptica, and canine distemper virus (arithmetic mean ± SD log10 microorganism reduction, 2.57 ± 0.47, ≥ 3.45 ± 0.24, and ≥ 1.50 ± 0.25, respectively) collected from air. Feline herpesvirus-1 was detected in only 1 sample without and no samples with UVGI treatment. Feline calicivirus and canine parvovirus were not detectable in any collected samples. CONCLUSIONS AND CLINICAL RELEVANCE: Results for some surrogates of veterinary pathogens suggested a potential benefit to supplementing manual disinfection practices with UVGI-based air cleaning systems in animal care environments. Further research is needed to investigate the utility of UVGI in operating HVAC systems.

Comparative survival of viruses during thermophilic and mesophilic anaerobic digestion.

Micro-scale technology was used to obtain survival data for three animal viruses and two bacteriophages during anaerobic digestion. The data for adenoviruses, MS2 and Φ6 provide the first published reports for survival of these viruses during mesophilic anaerobic digestion. Data were also obtained for thermophilic digestion, which showed greater inactivation of viruses at the higher temperature. Data on the survival of Φ6 are of particular interest since it is a lipid-based virus which has been suggested as a surrogate for the Ebola virus. In contrast, MS2 was found to be an inappropriate surrogate for Ebola.

Distribution of Bacteria in Dental Offices and the Impact of Hydrogen Peroxide Disinfecting Wipes.

Purpose: The purpose of this study was to characterize the occurrence of heterotrophic plate count bacteria (HPCs), fecal bacterial indicators, and methicillin-resistant Staphylococcus aureus (MRSA) on hard, nonporous surfaces (fomites) commonly found in dental offices, and to assess the impact of a hygienic disinfection intervention on the reduction of these bacteria in the office setting.Methods: Samples of various fomites were collected from dental offices located in Arizona (6) and Illinois (4) and assayed for HPCs, total coliforms, Escherichia coli, and MRSA in order to establish baseline bacterial levels. Dental office personnel were then provided with disposable disinfectant wipes containing hydrogen peroxide to use in the office, and the surfaces were subsequently resampled to assess their impact on the specified bacterial populations.Results: The greatest numbers of HPCs were found on the patients' arm rest and office phones, with dental tool handles and the dentist examination lights yielding the lowest levels. Coliform bacteria and E. coli were also detected on those surfaces demonstrating the highest HPCs, and were cultured from other fomites as well. MRSA was also isolated from 5% of the fomites tested. The use of disinfectant wipes significantly reduced the numbers of HPC bacteria detected on fomites (p=0.002). No total coliforms, E. coli, or MRSA were cultured from disinfected fomites following the hygienic intervention.Conclusion: The use of hydrogen peroxide-impregnated towelettes reduced total bacterial numbers on fomites commonly located in dental offices. Total coliforms, E. coli, and MRSA were reduced to levels below detection.

Effectiveness of poliovirus concentration and recovery from treated wastewater by two electropositive filter methods.

Enteric viruses are often present in low numbers in various water matrices. Virus sampling therefore involves multiple concentration steps to condense large samples down to small volumes for detection by cell culture or molecular assays. The NanoCeram® Virus Sampler has been demonstrated to be effective for the recovery of viruses from tap water, surface waters, and seawater. The goal of this study was to evaluate a new method using NanoCeram® filters for the recovery of poliovirus 1 (PV-1) from treated wastewater. Activated sludge effluent samples were spiked with PV-1 and concentrated in side-by-side tests by two methods: (1) NanoCeram® filtration, elution with sodium polyphosphate buffer, secondary concentration via centrifugal ultrafiltration; and (2) 1MDS filtration, elution with beef extract, secondary concentration via organic flocculation. The virus retention and elution efficiencies did not differ significantly between the two methods. In contrast, the secondary concentrate volume was smaller for the NanoCeram® method (8.4 vs. 30 mL) and the secondary concentration efficiencies were different between the two methods with 98 % for centrifugal ultrafiltration (NanoCeram® and 45 % for organic flocculation (1MDS). The overall method efficiencies were significantly different (P ≤ 0.05) with the NanoCeram® method yielding a 57 % and the 1MDS a 23 % virus recovery. In addition, there appeared to be less interference with viral detection via polymerase chain reaction with the NanoCeram® concentrates. This NanoCeram® method therefore is able to efficiently recover PV-1 from large volumes of wastewater and may serve as an inexpensive alternative to the standard 1MDS filter method for such applications.

Concentration and recovery of viruses from water: a comprehensive review.

Enteric viruses are a cause of waterborne disease worldwide, and low numbers in drinking water can present a significant risk of infection. Because the numbers are often quite low, large volumes (100-1,000 L) of water are usually processed. The VIRADEL method using microporous filters is most commonly used today for this purpose. Negatively charged filters require the addition of multivalent salts and acidification of the water sample to effect virus adsorption, which can make large-volume sampling difficult. Positively charged filters require no preconditioning of samples, and are able to concentrate viruses from water over a greater pH range than electronegative filters. The most widely used electropositive filter is the Virosorb 1MDS; however, the Environmental Protection Agency has added the positively charged NanoCeram filters to their proposed Method 1615. Ultrafilters concentrate viruses based on size exclusion rather than electrokinetics, but are impractical for field sampling or processing of turbid water. Elution (recovery) of viruses from filters following concentration is performed with organic (e.g., beef extract) or inorganic solutions (e.g., sodium polyphosphates). Eluates are then reconcentrated to decrease the sample volume to enhance detection methods (e.g., cell culture infectivity assays and molecular detection techniques). While the majority of available filters have demonstrated high virus retention efficiencies, the methods to elute and reconcentrate viruses have met with varying degrees of success due to the biological variability of viruses present in water.

Culturable microbial diversity and the impact of tourism in Kartchner Caverns, Arizona.

Kartchner Caverns in Benson, AZ, was opened for tourism in 1999 after a careful development protocol that was designed to maintain predevelopment conditions. As a part of an ongoing effort to determine the impact of humans on this limestone cave, samples were collected from cave rock surfaces along the cave trail traveled daily by tour groups (200,000 visitors year-1) and compared to samples taken from areas designated as having medium (30-40 visitors year-1) and low (2-3 visitors year-1) levels of human exposure. Samples were also taken from fiberglass moldings installed during cave development. Culturable bacteria were recovered from these samples and 90 unique isolates were identified by using 16S rRNA polymerase chain reaction and sequencing. Diversity generally decreased as human impact increased leading to the isolation of 32, 27, and 22 strains from the low, medium, and high impact areas, respectively. The degree of human impact was also reflected in the phylogeny of the isolates recovered. Although most isolates fell into one of three phyla: Actinobacteria, Firmicutes, or Proteobacteria, the Proteobacteria were most abundant along the cave trail (77% of the isolates), while Firmicutes predominated in the low (66%) and medium (52%) impact areas. Although the abundance of Proteobacteria along the cave trail seems to include microbes of environmental rather than of anthropogenic origin, it is likely that their presence is a consequence of increased organic matter availability due to lint and other organics brought in by cave visitors. Monitoring of the cave is still in progress to determine whether these bacterial community changes may impact the future development of cave formations.