Using AlphaFold Predictions in Viral Research.

Elucidation of the tertiary structure of proteins is an important task for biological and medical studies. AlphaFold, a modern deep-learning algorithm, enables the prediction of protein structure to a high level of accuracy. It has been applied in numerous studies in various areas of biology and medicine. Viruses are biological entities infecting eukaryotic and procaryotic organisms. They can pose a danger for humans and economically significant animals and plants, but they can also be useful for biological control, suppressing populations of pests and pathogens. AlphaFold can be used for studies of molecular mechanisms of viral infection to facilitate several activities, including drug design. Computational prediction and analysis of the structure of bacteriophage receptor-binding proteins can contribute to more efficient phage therapy. In addition, AlphaFold predictions can be used for the discovery of enzymes of bacteriophage origin that are able to degrade the cell wall of bacterial pathogens. The use of AlphaFold can assist fundamental viral research, including evolutionary studies. The ongoing development and improvement of AlphaFold can ensure that its contribution to the study of viral proteins will be significant in the future.

Potential Applications of Thermophilic Bacteriophages in One Health.

Bacteriophages have a wide range of applications such as combating antibiotic resistance, preventing food contamination for food safety, and as biomarkers to indirectly assess the quality of water. Additionally, bacteriophage components (endolysins and coat proteins) have a lot of applications in food processing, vaccine design, and the delivery of cargo to the body. Therefore, bacteriophages/components have a multitude of applications in human, plant/veterinary, and environmental health (One Health). Despite their versatility, bacteriophage/component use is mostly limited to temperatures within 4-40 °C. This limits their applications (e.g., in food processing conditions, pasteurization, and vaccine design). Advances in thermophilic bacteriophage research have uncovered novel thermophilic endolysins (e.g., ΦGVE2 amidase and MMPphg) that can be used in food processing and in veterinary medicine. The endolysins are thermostable at temperatures > 65 °C and have broad antimicrobial activities. In addition to thermophilic endolysins, enzymes (DNA polymerase and ligases) derived from thermophages have different applications in molecular biology/biotechnology: to generate DNA libraries and develop diagnostics for human and animal pathogens. Furthermore, coat proteins from thermophages are being explored to develop virus-like particle platforms with versatile applications in human and animal health. Overall, bacteriophages, especially those that are thermophilic, have a plethora of applications in One Health.

Efficacy evaluation of an air-assisted electrostatic disinfection device for the effective disinfection and sanitization against the spread of pathogenic infections

This research aims to check the chargeability of sodium hypochlorite and the efficacy evaluation of an air-assisted electrostatic disinfection device. Five different inanimate surfaces i.e., wood, glass, stainless steel, plastic and fabric were considered to examine the performance in terms of efficacy, survival time, off-target losses, spray coverage and the volume of disinfectant consumed. A significant charge-to-mass level of 2.43 mC/kg was achieved for sodium hypochlorite at an applied voltage of 2.0 kV, a liquid flowrate of 253 ml/min and applied air pressure of 4.0 bar. The experimental results found that 1000 mg/L of sodium hypochlorite concentration effectively eliminated Pseudomonas aeruginosa, Clostridium perfringens and Bacteriophage MS2 colonies. © 2023 Elsevier B.V.

Emergent crisis of antibiotic resistance: A silent pandemic threat to 21st century.

Antibiotic resistance has become an indispensably alarming menace to the global community. The primary factors are overuse and abuse of antibiotics, lack of novel medicines under development, the health care industry's focus on profit, and the absence of diagnostic testing prior to the prescription of antibiotics. Additionally, over the past few decades, the main factors contributing to the global spread of antibiotic resistance have been the overuse of antibiotics in livestock and other animals, drug efficacy, development of fewer new vaccines, environmental toxicity, transmission through travel, and lack of funding for healthcare research and development. These factors have accelerated resistance in microorganisms through structural and functional modifications in bacteria such as reduced drug permeability, increased efflux pumps, enzymatic antibiotic modification, and change in drug target, intracellular infection, and biofilm creation. There has been an increase in resistance during the pandemic and among cancer patients due to improper prescriptions. A number of modern therapeutic alternatives have been developed to curb widespread antibiotic resistance such as nanoparticle, bacteriophage, and antimicrobial biochemical approaches. It is high time to explore new alternatives to curtail enormous increase in resistant pathogens which could be an incurable global confrontation. This review highlights the complete insight on the global drivers of resistance along with the modes of action and impacts, finally discussing the latest therapeutic alternatives.

Potential Use of a Combined Bacteriophage-Probiotic Sanitation System to Control Microbial Contamination and AMR in Healthcare Settings: A Pre-Post Intervention Study.

Microbial contamination in the hospital environment is a major concern for public health, since it significantly contributes to the onset of healthcare-associated infections (HAIs), which are further complicated by the alarming level of antimicrobial resistance (AMR) of HAI-associated pathogens. Chemical disinfection to control bioburden has a temporary effect and can favor the selection of resistant pathogens, as observed during the COVID-19 pandemic. Instead, probiotic-based sanitation (probiotic cleaning hygiene system, PCHS) was reported to stably abate pathogens, AMR, and HAIs. PCHS action is not rapid nor specific, being based on competitive exclusion, but the addition of lytic bacteriophages that quickly and specifically kill selected bacteria was shown to improve PCHS effectiveness. This study aimed to investigate the effect of such combined probiotic-phage sanitation (PCHSφ) in two Italian hospitals, targeting staphylococcal contamination. The results showed that PCHSφ could provide a significantly higher removal of staphylococci, including resistant strains, compared with disinfectants (-76%, p < 0.05) and PCHS alone (-50%, p < 0.05). Extraordinary sporadic chlorine disinfection appeared compatible with PCHSφ, while frequent routine chlorine usage inactivated the probiotic/phage components, preventing PCHSφ action. The collected data highlight the potential of a biological sanitation for better control of the infectious risk in healthcare facilities, without worsening pollution and AMR concerns.

Virus Elimination Using High-Voltage Pulses in Aqueous Solutions

This article presents the results of the research conducted on bacteriophages inactivation using high-voltage (HV) short electric pulses. The bacteriophages are bacterial viruses, which share similar basic features in their structure to other viruses, so that they may be considered a first choice model for experimental studies on viral pathogens, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV2). The aqueous solution of bacteriophages was subjected to the influence of high electric-field pulses, provided by a Marx generator. It had been observed that the electric discharge was a two-stage process where the initial inrush charging current was followed by a formation of a discharge channel due to local charge trapping and a secondary burst of current. The possible explanation of inactivation of bacteriophages could be due to the formation of discharge plasma observed during the experiments, followed by likely creation of radical species. Other plausible additional mechanisms include irreversible electroporation. Two different electrode configurations with different electric-field spatial distributions were examined. A complete 100% inactivation of the bacteriophages was achieved in the system featuring the uniform field distribution. This article novelty lies in the fact that it demonstrates 100% effectiveness of the HV pulses treatment in elimination of viruses, and proposes a method of capturing these pathogens from air. IEEE

Increased Handpiece Speeds without Air Coolant: Aerosols and Thermal Impact.

This study assessed the impact of increased speed of high-speed contra-angle handpieces (HSCAHs) on the aerosolization of a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) surrogate virus and any concomitant thermal impact on dental pulp. A bacteriophage phantom-head model was used for bioaerosol detection. Crown preparations were performed with an NSK Z95L Contra-Angle 1:5 (HSCAH-A) and a Bien Air Contra-Angle 1:5 Nova Micro Series (HSCAH-B) at speeds of 60,000, 100,000, and 200,000 revolutions per minute (rpm), with no air coolant. Bioaerosol dispersal was measured with Φ6-bacteriophage settle plates, air sampling, and particle counters. Heating of the internal walls of the pulp chambers during crown preparation was assessed with an infrared camera with HSCAH-A and HSCAH-B at 200,000 rpm (water flows ≈15 mL min-1 and ≈30 mL min-1) and an air-turbine control (≈23.5 mL min-1) and correlated with remaining tissue thickness measurements. Minimal bacteriophage was detected on settle or air samples with no notable differences observed between handpieces or speeds (P > 0.05). At all speeds, maximum settled aerosol and average air detection was 1.00 plaque-forming units (pfu) and 0.08 pfu/m3, respectively. Irrespective of water flow rate or handpiece, both maximum temperature (41.5°C) and temperature difference (5.5°C) thresholds for pulpal health were exceeded more frequently with reduced tissue thickness. Moderate and strong negative correlations were observed based on Pearson's correlation coefficient, between remaining dentine thickness and either differential (r = -0.588) or maximum temperature (r = -0.629) measurements, respectively. Overall, HSCAH-B generated more thermal energy and exceeded more temperature thresholds compared to HSCAH-A. HSCAHs without air coolant operating at speeds of 200,000 rpm did not increase bioaerosolization in the dental surgery. Thermal risk is variable, dependent on handpiece design and remaining dentine thickness.

Virus Occurrence and Survival in Reusable Resources: A Minireview

This work covers important aspects of the occurrence and viability of various viruses in the two most common reusable waste resources: wastewater and biomass waste. Detection of human, bacterial and plant viruses in these wastes are summarized. Historically, human viruses have been monitored in wastewater for decades. Evidence suggests that wastewater mostly contains fecal-orally transmitted viruses, which are abundant and diverse. Recently, an increasing occurrence of SARS-CoV2 in sewage water with the spreading epidemics has been confirmed but lacking biological proof of infectivity yet. Besides human pathogens, wastewater is shown to be rich in bacteriophages and plant viruses as well, which supposedly enter the water from human guts. Viruses serving as water quality indicators are also discussed here. Lastly, we focus on biomass waste treatment, showing the presence of some common and stable plant viruses which may supposedly survive the technological process. Copyright © Publisher PH <<Akademperiodyka>> of the NAS of Ukraine, 2022.

Ultrashort Peptides for the Self-Assembly of an Antiviral Coating

Antiviral compounds are important for generating sterile surfaces. Here, two extremely short peptides, DOPA-Phe-NH2 and DOPA-Phe(4F)-NH2 that can self-assemble into spherical nanoparticles with antiviral activity are presented. The peptide assemblies possess excellent antiviral activity against bacteriophage T4 with antiviral minimal inhibitory concentrations of 125 and 62.5 µg mL−1, for DOPA-Phe-NH2 and DOPA-Phe(4F)-NH2, respectively. When the peptide assemblies are applied on a glass substrate by drop-casting, they deactivate more than 99.9% of bacteriophage T4 and Canine coronavirus. Importantly, the peptide assemblies have low toxicity toward mammalian cells. Overall, the findings can provide a novel strategy for the design and development of antiviral coatings for a decreased risk of viral infections. © 2023 The Authors. Advanced Materials Interfaces published by Wiley-VCH GmbH.

Virus Occurrence and Survival in Reusable Resources: A Minireview

This work covers important aspects of the occurrence and viability of various viruses in the two most common reusable waste resources: wastewater and biomass waste. Detection of human, bacterial and plant viruses in these wastes are summarized. Historically, human viruses have been monitored in wastewater for decades. Evidence suggests that wastewater mostly contains fecal-orally transmitted viruses, which are abundant and diverse. Recently, an increasing occurrence of SARS-CoV2 in sewage water with the spreading epidemics has been confirmed but lacking biological proof of infectivity yet. Besides human pathogens, wastewater is shown to be rich in bacteriophages and plant viruses as well, which supposedly enter the water from human guts. Viruses serving as water quality indicators are also discussed here. Lastly, we focus on biomass waste treatment, showing the presence of some common and stable plant viruses which may supposedly survive the technological process. Copyright © Publisher PH <<Akademperiodyka>> of the NAS of Ukraine, 2022.

Virus inactivation by sequential ultraviolet-chlorine disinfection: Synergistic effect and mechanism.

The COVID-19 outbreak has raised concerns about the efficacy of the disinfection process followed in water treatment plants in preventing the spread of viruses. Ultraviolet (UV) and chlorine multi-barrier disinfection processes are commonly used in water treatment plants; however, their effects on virus inactivation are still unclear. In this study, the effects of different disinfection processes (i.e., UV, free chlorine, and their combination) on waterborne viruses were analyzed using bacteriophage surrogates (i.e., MS2 and PR772) as alternative indicators. The results showed that the inactivation rates of PR772 by either UV or free chlorine disinfection were higher than those of MS2. PR772 was approximately 1.5 times more sensitive to UV disinfection and 8.4 times more sensitive to chlorine disinfection than MS2. Sequential UV-chlorine disinfection had a synergistic effect on virus inactivation, which was enhanced by an increase in the UV dose. As compared with single free chlorine disinfection, UV irradiation at 40 mJ cm-2 enhanced MS2 and PR772 inactivation significantly with a 2.7-fold (MS2) and a 1.7-fold (PR772) increase in the inactivation rate constants on subsequent chlorination in phosphate buffered saline. The synergistic effect was also observed in real wastewater samples, in which the MS2 inactivation rate increased 1.4-fold on subsequent chlorination following UV irradiation at 40 mJ cm-2. The mechanism of the synergistic effect of sequential UV-chlorine disinfection was determined via sodium dodecyl sulfate-polyacrylamide gel electrophoresis, using MS2 as an indicator. The results showed that the synergistic effect was due to damage to MS2 surface proteins caused by previous UV disinfection, which enhanced the sensitivity of MS2 to chlorination. This study provides a feasible approach for the efficient inactivation of viruses in water supply and drainage.

Bacteriophage Cocktails in the Post-COVID Rehabilitation.

Increasing evidence suggests that gut dysbiosis is associated with coronavirus disease 2019 (COVID-19) infection and may persist long after disease resolution. The excessive use of antimicrobials in patients with COVID-19 can lead to additional destruction of the microbiota, as well as to the growth and spread of antimicrobial resistance. The problem of bacterial resistance to antibiotics encourages the search for alternative methods of limiting bacterial growth and restoring the normal balance of the microbiota in the human body. Bacteriophages are promising candidates as potential regulators of the microbiota. In the present study, two complex phage cocktails targeting multiple bacterial species were used in the rehabilitation of thirty patients after COVID-19, and the effectiveness of the bacteriophages against the clinical strain of Klebsiella pneumoniae was evaluated for the first time using real-time visualization on a 3D Cell Explorer microscope. Application of phage cocktails for two weeks showed safety and the absence of adverse effects. An almost threefold statistically significant decrease in the anaerobic imbalance ratio, together with an erythrocyte sedimentation rate (ESR), was detected. This work will serve as a starting point for a broader and more detailed study of the use of phages and their effects on the microbiome.

Symbiotic microbiota: A class of potent immunomodulators

Microbiome is known to exist as symbiotic commensals in humans, domestic and wild animals, birds, fishes, reptiles, insects etc. DNA sequencing and metagenomic platforms have deciphered the complex role played by communities of microbiota (bacteria, fungi, viruses, protozoa and other eukaryotic species) in survival and regulation of host physiology, metabolism and regulation of host immune system. Any alteration in the microbial population or breach in the symbiotic alliance with the host may ultimately lead to development of different kinds of pathologies. Realization of the enormous role played by the microbiome in health and diseases of human and domestic livestock led researchers to find ways to modulate these resident microbiomes for improvement in health and management of diseases. Theoretically there are several ways that can be employed for manipulating the composition and functional capacity of the resident microbiome, which may lead to improvements in human and livestock health. Though studies have shown therapeutic potential of the microbiome, considerable challenges exist in the actual implementation of these strategies in clinical settings. This review discusses the symbiotic relationship between microbiome and host and strategies to modulate host immune responses by manipulating microbiome profile. Paper also highlights how to overcome existing obstacles for successful implementation of microbiome manipulation techniques. In this era of COVID-19, it would be worth analysing the role of resident microbiome in the magnitude of COVID-19 severity which may have occurred through immunomodulation.

Particulate Matter versus Airborne Viruses-Distinctive Differences between Filtering and Inactivating Air Cleaning Technologies

The current pandemic of the SARS-CoV-2 virus requires measures to reduce the risk of infection. In addition to the usual hygiene measures, air cleaners are a recommended solution to decrease the viral load in rooms. Suitable technologies range from pure filters to inactivating units, such as cold plasma or UVC irradiation. Such inactivating air cleaners, partly combined with filter technology, are available on the market in various designs, dimensions and technical specifications. Since it is not always clear whether they may produce undesirable by-products, and the suitability for particular applications cannot be assessed on the basis of the principle of operation, the effectivity of six inactivating devices was investigated in a near-real environment. The investigations were based on a standard method published by the VDI. The procedure was extended in such a way that a permanent virus source was simulated, which corresponds to the presence of a person suffering from COVID-19 in a room. The study addresses the difference of the mere presence of viruses to the determination of the virulence. As a result, a deep understanding is provided between the behavior of a virus as a pure aerosolized particle and its real infectivity in order to enable the assessment of suitable air cleaners.

VIRAL PRODUCTION AND TRANSDUCTION FOR GENE-EDITING USING MICROFLUIDICS

Viral production and transduction have seen a considerable increase in use for gene therapies and especially vaccines (e.g., COVID-19) due to their abilities to deliver a significant amount of genetic information and integrate it into the genome. However, challenges associated with viral production/transduction involve steps that are very time consuming, manually intensive, and laborious. In efforts to expedite this process, we have created a microfluidic methodology that will provide a “hands-off” workflow in the genome engineering pipeline. In this work, we developed a platform which can generate lentiviral particles on-demand containing the gene-editing machinery that will be able to modify target breast cancer cell lines. © 2021 MicroTAS 2021 - 25th International Conference on Miniaturized Systems for Chemistry and Life Sciences. All rights reserved.

Rapid SARS-CoV-2 testing with duplexed recombinase polymerase amplification and a bacteriophage internal control

Without global mass vaccination, COVID-19 will continue to infect and cause serious illness, disproportionately in low- and middle-income countries. Point-of-care and home-based nucleic acid amplification tests (NAATs) are valuable tools to control COVID-19 transmission. Here we present a rapid isothermal NAAT for duplexed detection of SARS-CoV-2 and an MS2 bacteriophage internal control. This assay amplifies RNA in less than 15 minutes, utilizes a low temperature of 39°C, and has fluorescence or visual lateral flow readout. This positions our assay for use in low-cost paper-based nucleic acid diagnostic devices for ultrasensitive and reliable COVID-19 detection in POC or home-based settings. © 2021 MicroTAS 2021 - 25th International Conference on Miniaturized Systems for Chemistry and Life Sciences. All rights reserved.

Effectiveness of air-purifying devices and measures to reduce the exposure to bioaerosols in school classrooms.

The SARS-CoV-2 pandemic, which suddenly appeared at the beginning of 2020, revealed our knowledge deficits in terms of ventilation and air pollution control. It took many weeks to realize that aerosols are the main route of transmission. The initial attempt to hold back these aerosols through textile masks seemed almost helpless, although there is sufficient knowledge about the retention capacity of fabric filters for aerosols. In the absence of a sufficient number of permanently installed heating, ventilation, and air conditioning systems, three main approaches are pursued: (a) increasing the air exchange rate by supplying fresh air, (b) using mobile air purifiers, and (c) disinfection by introducing active substances into the room air. This article discusses the feasibility of these different approaches critically. It also provides experimental results of air exchange measurements in a school classroom that is equipped with a built-in fan for supplying fresh air. With such a fan and a window tilted at the appropriate distance, an air exchange rate of 5/h can be set at a low power level and without any significant noise pollution. Heat balance calculations show that no additional heat exchanger is necessary in a normal classroom with outside temperatures above 10°C. Furthermore, a commercial mobile air purifier is studied in a chamber and a test room setup in order to examine and evaluate the efficiency of such devices against viable viruses under controlled and realistic conditions. For this purpose, bacteriophages of the type MS2 are used. Both window ventilation and air purifiers were found to be suitable to reduce the concentration of phages in the room.

Dynamics of nasopharyngeal tract phageome and association with disease severity and age of patients during three waves of COVID-19.

In December 2019, several patients were hospitalized and diagnosed with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, which subsequently led to a global pandemic. To date, there are no studies evaluating the relationship between the respiratory phageome and the SARS-CoV-2 infection. The current study investigated the phageome profiles in the nasopharyngeal swabs collected from 55 patients during the three different waves of coronavirus disease 2019 (COVID-19) in the Campania Region (Southern Italy). Data obtained from the taxonomic profiling show that phage families belonging to the order Caudovirales have a high abundance in the patient samples. Moreover, the severity of the COVID-19 infection seems to be correlated with the phage abundance.

Structurally Modified Plant Viruses and Bacteriophages with Helical Structure. Properties and Applications.

Structurally modified virus particles can be obtained from the rod-shaped or filamentous virions of plant viruses and bacteriophages by thermal or chemical treatment. They have recently attracted attention of the researchers as promising biogenic platforms for the development of new biotechnologies. This review presents data on preparation, structure, and properties of the structurally modified virus particles. In addition, their biosafety for animals is considered, as well as the areas of application of such particles in biomedicine. A separate section is devoted to one of the most relevant and promising areas for the use of structurally modified plant viruses - design of vaccine candidates based on them.

Design and Feature Analysis of a Thermally Coupled Corona Discharge Air Disinfection Device

The COVID-19 caused by the novel coronavirus is still spreading globally, and blocking its airborne transmission route is of great significance to control the pandemic. The conventional plasma air disinfection devices show advantages in their dynamic and rapid capabilities, but the disinfection performance is limited by a single method, besides, there exists the risk of secondary infection during maintenance. In this work, according to the physiological characteristics of the novel coronavirus, an air disinfection device based on thermally coupled corona discharge was proposed for the improvement of conventional plasma air disinfection technology, which adopted the wire-plate array electrode structure to initiate corona discharge, and utilized heating wires embedded in the collection plate to achieve centralized heating. The discharge para-meters were measured, and a discharge power at stable operation was discovered to be as high as 5.6 W, for which the discharge law was found to obey the Townsend relationship. Measurement and simulation of the thermal parameters showed that, compared with the overall air heating, the efficiency of centralized heating was increased by 17 times, with minimal impact on the ambient temperature. Bacillus amyloliquefaciens and Bacillus subtilis bacteriophages were used as model bacteria and virus to verify the disinfection performance. Results demonstrate that the killing performance is effectively enhanced via thermally coupled corona discharge, with a removal rate of residual virus on the collection plate increasing by 99.97%, thereby reducing the risk of secondary infection. This work lays a device foundation for killing the airborne novel coronavirus, and also provides a technical reference for cutting its airborne transmission. © 2022, High Voltage Engineering Editorial Department of CEPRI. All right reserved.