Strain-dependent variations in replication of European clade 2.3.4.4b influenza A(H5N1) viruses in bovine cells and thermal inactivation in semi-skimmed or whole milk.

We investigated the thermostability of four European avian influenza A(H5N1) viruses in whole and semi-skimmed milk and their replication in bovine kidney and lung cells amid the current influenza A(H5N1) dairy cattle outbreak in the United States. Results showed strain-dependent differences in thermal inactivation, particularly in whole milk, and variable replication efficacy in lung cells. These findings support assessing the inactivation of European H5N1 viruses in milk and their replication in bovine cells, aiding biosafety protocols and public health measures.

Validated Methods for Effective Decontamination and Inactivation of Rift Valley Fever Virus.

The Rift Valley fever virus (RVFV) is an arthropod-borne, zoonotic, hemorrhagic fever virus that can cause severe diseases both in livestock and humans. The spread of RVFV in areas previously considered as non-endemic together with the absence of licensed vaccines for use in humans and animals poses a major health and economic threat worldwide. It is therefore crucial to make major progresses in our understanding and management of this virus and its zoonosis. RVFV is considered a bioterrorism pathogen, and, thus, only a few institutes, facilities, and personnel are legally authorized to detain it and handle it. Moreover, this virus must be manipulated in a biosafety level 3 (BSL3) laboratory following strict biosafety protocols to ensure that biosecurity's highest standards are met. Only certain attenuated strains such as the MP12 strain can be handled in BSL2 laboratories, depending on the country considered. To assist researchers in working with RVFV in the safest possible conditions, this chapter presents validated methods for effective RVFV decontamination and inactivation.

The Influence of Simulated Organic Matter on the Inactivation of Viruses: A Review.

Viruses impose a significant public health burden globally, and one of the key elements in controlling their transmission is the ability to inactivate them using disinfectants. However, numerous challenges to inactivating foodborne viruses exist due to inherent viral characteristics (such as recalcitrance to commonly used inactivation agents) and external factors (such as improper cleaning before application of inactivation agent, improper contact time, etc.). Given the potential for improper application of disinfectants (such as shorter than recommended contact time, improper disinfectant concentration, etc.), understanding the performance of a disinfectant in the presence of an organic load is important. To accomplish this, the introduction of simulated organic loads is often used when studying the efficacy of a disinfectant against different viruses. However, the different types of simulated organic loads used in foodborne virus inactivation studies or their relative effects on inactivation have not been reviewed. The purpose of this review is to survey different simulated organic load formulations used in studying foodborne virus inactivation, as well as present and compare the influence of these different formulations on viral inactivation. The findings included in this review suggest that many simulated organic load formulations can reduce disinfectants' efficacy against viruses. Based on the findings in this review, blood, particularly serum or feces, are among the most commonly used and efficacious forms of simulated organic load in many tests.

Assessing the Efficacy of Three Hatchery Disinfectants for the Inactivation of a Lake Sturgeon Herpesvirus (Family: Alloherpesviridae).

Infectious diseases are a leading cause of losses in the aquaculture industry and conservation programs globally. Simultaneously, infectious diseases pose a substantial risk to fish being hatchery-reared and released into natural habitats for conservation purposes, including the Great Lakes lake sturgeon (Acipenser fulvescens, i.e., GL-LST). Recently, an alloherpesvirus (lake sturgeon herpesvirus 2, i.e., LSHV-2) capable of inducing disease and/or mortality in adult and juvenile GL-LSTs was detected in two adult GL-LST populations. To begin developing disease prevention and/or control methods, in vitro experiments were designed to determine the susceptibility of LSHV-2 to disinfectants commonly used in hatchery and aquaculture facilities (Virkon®-Aquatic: potassium peroxymonosulfate; Ovadine®: polyvinylpyrrolidone iodine complex; and Perox-Aid®: hydrogen peroxide). Cultured LSHV-2 was exposed to each disinfectant at two concentrations (Virkon®-Aquatic: 0.5% and 1%; Ovadine®: 50 and 100 ppm; and Perox-Aid®: 500 and 1000 ppm) in duplicate for durations of 1, 10, and 30 min. Following exposure, the disinfectant was neutralized, and after a 14-day incubation period on a white sturgeon × lake sturgeon hybrid cell line (WSxLS), percent reduction was calculated by comparing the 50% tissue culture infectious doses (TCID50/mL) of the virus with and without disinfectant exposure. When exposed to Perox-Aid®, LSHV-2 percent reduction ranged from 58.7% to 99.5%. When exposed to Ovadine®, the percent reduction ranged from 99.4% to 100%. Lastly, the percent reduction when exposed to Virkon®-Aquatic was 100% for both concentrations and all timepoints. The results herein provide evidence that both Virkon®-Aquatic and Ovadine® are virucidal to LSHV-2 and may represent a means to reduce virus transmission risk under field settings.

Bacillaceae serine proteases and Streptomyces epsilon-poly-L-lysine synergistically inactivate Caliciviridae by inhibiting RNA genome release.

Human norovirus (HuNoV) is an enteric infectious pathogen belonging to the Caliciviridae family that causes occasional epidemics. Circulating alcohol-tolerant viral particles that are readily transmitted via food-borne routes significantly contribute to the global burden of HuNoV-induced gastroenteritis. Moreover, contact with enzymes secreted by other microorganisms in the environment can impact the infectivity of viruses. Hence, understanding the circulation dynamics of Caliciviridae is critical to mitigating epidemics. Accordingly, in this study, we screened whether environmentally abundant secretase components, particularly proteases, affect Caliciviridae infectivity. Results showed that combining Bacillaceae serine proteases with epsilon-poly-L-lysine (EPL) produced by Streptomyces-a natural antimicrobial-elicited anti-Caliciviridae properties, including against the epidemic HuNoV GII.4_Sydney_2012 strain. In vitro and in vivo biochemical and virological analyses revealed that EPL has two unique synergistic viral inactivation functions. First, it maintains an optimal pH to promote viral surface conformational changes to the protease-sensitive structure. Subsequently, it inhibits viral RNA genome release via partial protease digestion at the P2 and S domains in the VP1 capsid. This study provides new insights regarding the high-dimensional environmental interactions between bacteria and Caliciviridae, while promoting the development of protease-based anti-viral disinfectants.

Estimation of the UV susceptibility of aerosolized SARS-CoV-2 to 254 nm irradiation using CFD-based room disinfection simulations.

The recent COVID-19 pandemic has raised interest in efficient air disinfection solutions. The application of germicidal ultraviolet (GUV) irradiation is an excellent contender to prevent airborne transmission of COVID-19, as well as other existing and future infectious airborne diseases. While GUV has already been proven effective in inactivating SARS-CoV-2, quantitative data on UV susceptibility and dose requirements, needed to predict and optimize the performance of GUV solutions, is still limited. In this study, the UV susceptibility of aerosolized SARS-CoV-2 to 254 nm ultraviolet (UV) irradiation is investigated. This is done by employing 3D computational fluid dynamics based simulations of SARS-CoV-2 inactivation in a test chamber equipped with an upper-room UV-C luminaire and comparing the results to previously published measurements performed in the same test chamber. The UV susceptibility found in this study is (0.6 ± 0.2) m2/J, which is equivalent to a D90 dose between 3 and 6 J/m2. These values are in the same range as previous estimations based on other corona viruses and inactivation data reported in literature.

Light-Induced Transformation of Virus-Like Particles on TiO2.

Titanium dioxide (TiO2) shows significant potential as a self-cleaning material to inactivate severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and prevent virus transmission. This study provides insights into the impact of UV-A light on the photocatalytic inactivation of adsorbed SARS-CoV-2 virus-like particles (VLPs) on a TiO2 surface at the molecular and atomic levels. X-ray photoelectron spectroscopy, combined with density functional theory calculations, reveals that spike proteins can adsorb on TiO2 predominantly via their amine and amide functional groups in their amino acids blocks. We employ atomic force microscopy and grazing-incidence small-angle X-ray scattering (GISAXS) to investigate the molecular-scale morphological changes during the inactivation of VLPs on TiO2 under light irradiation. Notably, in situ measurements reveal photoinduced morphological changes of VLPs, resulting in increased particle diameters. These results suggest that the denaturation of structural proteins induced by UV irradiation and oxidation of the virus structure through photocatalytic reactions can take place on the TiO2 surface. The in situ GISAXS measurements under an N2 atmosphere reveal that the virus morphology remains intact under UV light. This provides evidence that the presence of both oxygen and UV light is necessary to initiate photocatalytic reactions on the surface and subsequently inactivate the adsorbed viruses. The chemical insights into the virus inactivation process obtained in this study contribute significantly to the development of solid materials for the inactivation of enveloped viruses.

Does pasteurization inactivate bird flu virus in milk?

Recently, an outbreak of highly pathogenic avian influenza A (H5N1), which carries the clade 2.3.4.4b hemagglutinin (HA) gene and has been prevalent among North American bird populations since the winter of 2021, was reported in dairy cows in the United States. As of 24 May 2024, the virus has affected 63 dairy herds across nine states and has resulted in two human infections. The virus causes unusual symptoms in dairy cows, including an unexpected drop in milk production, and thick colostrum-like milk. Notably, The US Food and Drug Administration reported that around 20% of tested retail milk samples contained H5N1 viruses, with a higher percentage of positive results from regions with infected cattle herds. Data are scant regarding how effectively pasteurization inactivates the H5N1 virus in milk. Therefore, in this study, we evaluated the thermal stability of the H5 clade 2.3.4.4b viruses, along with one human H3N2 virus and other influenza subtype viruses, including H1, H3, H7, H9, and H10 subtype viruses. We also assessed the effectiveness of pasteurization in inactivating these viruses. We found that the avian H3 virus exhibits the highest thermal stability, whereas the H5N1 viruses that belong to clade 2.3.4.4b display moderate thermal stability. Importantly, our data provide direct evidence that the standard pasteurization methods used by dairy companies are effective in inactivating all tested subtypes of influenza viruses in raw milk. Our findings indicate that thermally pasteurized milk products do not pose a safety risk to consumers.

Light exacerbates local and global effects induced by pH unfolding of Ipilimumab.

Monoclonal antibodies (mAbs) are an essential class of therapeutic proteins for the treatment of cancer, autoimmune and rare diseases. During their production, storage, and administration processes, these proteins encounter various stressors such as temperature fluctuations, vibrations, and light exposure, able to induce chemico-physical modifications to their structure. Viral inactivation is a key step in downstream processes, and it is achieved by titration of the mAb at low pH, followed by neutralization. The changes of the pH pose a significant risk of unfolding and subsequent aggregation to proteins, thereby affecting their manufacturing. This study aims to investigate whether a combined exposure to light during the viral inactivation process can further affect the structural integrity of Ipilimumab, a mAb primarily used in the treatment of metastatic melanoma. The biophysical and biochemical characterization of Ipilimumab revealed that pH variation is a considerable risk for its stability with irreversible unfolding at pH 2. The threshold for Ipilimumab denaturation lies between pH 2 and 3 and is correlated with the loss of the protein structural cooperativity, which is the most critical factor determining the protein refolding. Light has demonstrated to exacerbate some local and global effects making pH-induced exposed regions more vulnerable to structural and chemical changes. Therefore, specific precautions to real-life exposure to ambient light during the sterilization process of mAbs should be considered to avoid loss of the therapeutic activity and to increase the yield of production. Our findings underscore the critical role of pH optimization in preserving the structural integrity and therapeutic efficacy of mAbs. Moreover, a detailed conformational study on the structural modifications of Ipilimumab may improve the chemico-physical knowledge of this effective drug and suggest new production strategies for more stable products under some kind of stress conditions.

TLR9-Dependent Activation by Inactivated Parapoxvirus Ovis in Murine Bone Marrow-Derived Dendritic Cells Is Associated with Specific Strain-Dependent Dendritic Cell Subsets.

INTRODUCTION: Inactivated parapoxvirus ovis (iPPVO) exerts strong immunomodulatory effects on innate immune cells, making it an attractive therapeutic candidate. However, little is known about the signaling pathways that are involved in iPPVO-induced immune responses. METHODS: In this study, we systematically analyzed how different types of dendritic cells (DCs) react to iPPVO (Zylexis, strain D1701) in both BALB/c and C57BL/6 mice by flow cytometry and ELISAs, and investigated which signaling pathway is related to DC activation by Western blotting and protein profiling. RESULTS: We demonstrated that bone marrow-derived conventional DCs (BM-cDCs) and bone marrow-derived plasmacytoid DCs (BM-pDCs) matured and secreted type I interferons in response to Zylexis stimulation in both mouse strains. Similarly, Zylexis promoted the secretion of IL-12/23p40 and TNF by pDCs. However, IL-12/23p40 and TNF secretion by cDCs were induced in BALB/c mice but not in C57BL/6 mice. Analyzing the underlying signaling pathways revealed that iPPVO-induced maturation of cDCs was Toll-like receptor 9 (TLR9) independent, while the maturation of pDCs partially depended on the TLR9 pathway. Moreover, the production of proinflammatory cytokines by cDCs and the secretion of IFN-α/ß by pDCs partially depended on the TLR9 pathway in both mouse strains. Therefore, other signaling pathways seem to participate in the response of DCs to iPPVO, supported by protein profiling. CONCLUSION: Our data provide useful insights into the diversity of iPPVO sensors and their varying effects across different strains and species.

Linear Mixed Model of Virus Disinfection by Free Chlorine to Harmonize Data Collected across Broad Environmental Conditions.

Despite the critical importance of virus disinfection by chlorine, our fundamental understanding of the relative susceptibility of different viruses to chlorine and robust quantitative relationships between virus disinfection rate constants and environmental parameters remains limited. We conducted a systematic review of virus inactivation by free chlorine and used the resulting data set to develop a linear mixed model that estimates chlorine inactivation rate constants for viruses based on experimental conditions. 570 data points were collected in our systematic review, representing 82 viruses over a broad range of environmental conditions. The harmonized inactivation rate constants under reference conditions (pH = 7.53, T = 20 °C, [Cl-] < 50 mM) spanned 5 orders of magnitude, ranging from 0.0196 to 1150 L mg-1 min-1, and uncovered important trends between viruses. Whereas common surrogate bacteriophage MS2 does not serve as a conservative chlorine disinfection surrogate for many human viruses, CVB5 was one of the most resistant viruses in the data set. The model quantifies the role of pH, temperature, and chloride levels across viruses, and an online tool allows users to estimate rate constants for viruses and conditions of interest. Results from the model identified potential shortcomings in current U.S. EPA drinking water disinfection requirements.

One-hit kill: On the inactivation of RNA viruses by ultraviolet (UV)-C-induced genomic damage.

Large scale outbreaks of infectious respiratory disease have repeatedly plagued the globe over the last 100 years. The scope and strength of the outbreaks are getting worse as pathogenic RNA viruses are rapidly evolving and highly evasive to vaccines and anti-viral drugs. Germicidal UV-C is considered as a robust agent to disinfect RNA viruses regardless of their evolution. While genomic damage by UV-C has been known to be associated with viral inactivation, the precise relationship between the damage and inactivation remains unsettled as genomic damage has been analyzed in small areas, typically under 0.5 kb. In this study, we assessed genomic damage by the reduced efficiency of reverse transcription of regions of up to 7.2 kb. Our data seem to indicate that genomic damage was directly proportional to the size of the genome, and a single hit of damage was sufficient for inactivation of RNA viruses. The high efficacy of UV-C is already effectively adopted to inactivate airborne RNA viruses.

Inactivation of yellow fever virus by WHO-recommended hand rub formulations and surface disinfectants.

Despite continued outbreaks of yellow fever virus (YFV) in endemic regions, data on its environmental stability or guidelines for its effective inactivation is limited. Here, we evaluated the susceptibility of the YFV 17D vaccine strain to inactivation by ethanol, 2-propanol, World Health Organization (WHO)-recommended hand rub formulations I and II, as well as surface disinfectants. In addition, two pathogenic strains were tested to compare inactivation kinetics by WHO-recommended hand rub formulations I and II. Furthermore, environmental stability of the vaccine strain was assessed. YFV 17D particles displayed infectivity half-life decay profiles of ~13 days at room temperature. Despite this extended environmental stability, YFV was efficiently inactivated by alcohols, WHO-recommended hand formulations, and four out of five tested surface disinfectants. These results are useful in defining disinfection protocols to prevent non-vector borne YFV transmission.

Electrochemical profiling of poliovirus particles inactivated by chemical method and ionizing radiation.

Electrochemical profiling of formaldehyde-inactivated poliovirus particles demonstrated a relationship between the D-antigen concentration and the intensity of the maximum amplitude currents of the poliovirus samples. The resultant signal was therefore identified as electrochemical oxidation of the surface proteins of the poliovirus. Using registration of electrooxidation of amino acid residues of the capsid proteins, a comparative electrochemical analysis of poliovirus particles inactivated by electrons accelerated with doses of 5 kGy, 10 kGy, 15 kGy, 25 kGy, 30 kGy at room temperature was carried out. An increase in the radiation dose was accompanied by an increase in electrooxidation signals. A significant increase in the signals of electrooxidation of poliovirus capsid proteins was detected upon irradiation at doses of 15-30 kGy. The data obtained suggest that the change in the profile and increase in the electrooxidation signals of poliovirus capsid proteins are associated with an increase in the degree of structural reorganization of surface proteins and insufficient preservation of the D-antigen under these conditions of poliovirus inactivation.

Review of factors affecting virus inactivation in aerosols and droplets.

The inactivation of viruses in aerosol particles (aerosols) and droplets depends on many factors, but the precise mechanisms of inactivation are not known. The system involves complex physical and biochemical interactions. We reviewed the literature to establish current knowledge about these mechanisms and identify knowledge gaps. We identified 168 relevant papers and grouped results by the following factors: virus type and structure, aerosol or droplet size, temperature, relative humidity (RH) and evaporation, chemical composition of the aerosol or droplet, pH and atmospheric composition. These factors influence the dynamic microenvironment surrounding a virion and thus may affect its inactivation. Results indicate that viruses experience biphasic decay as the carrier aerosols or droplets undergo evaporation and equilibrate with the surrounding air, and their final physical state (liquid, semi-solid or solid) depends on RH. Virus stability, RH and temperature are interrelated, but the effects of RH are multifaceted and still not completely understood. Studies on the impact of pH and atmospheric composition on virus stability have raised new questions that require further exploration. The frequent practice of studying virus inactivation in large droplets and culture media may limit our understanding of inactivation mechanisms that are relevant for transmission, so we encourage the use of particles of physiologically relevant size and composition in future research.

Tenacity of Animal Disease Viruses on Wood Surfaces Relevant to Animal Husbandry.

The aim of this study was to analyse the hygienic suitability of wood often used in animal husbandry. To this end, the inactivation of viruses (Enterovirus E as a surrogate for non-enveloped viruses and Newcastle disease virus as a surrogate for enveloped viruses) on germ carriers consisting of various types of wood was studied over an extended period to assess the biosafety of wood as an agricultural building material. The study was designed to assess the intrinsic biocidal activity of the wood itself, without the use of a disinfectant. The laboratory tests were based on German test guidelines and current European standards. Five different types of wood germ carriers, i.e., spruce (Picea abies), pine (Pinus sylvestris), poplar (Populus sp.), beech (Fagus sylvatica) and Douglas fir (Pseudotsuga menziesii), as well as stainless-steel carriers, were inoculated with enveloped and non-enveloped viruses and stored for up to four months, and the remaining infectivity of the viruses was continuously assessed. The results showed that intact, finely sawn timber with a low depth of roughness had an inactivating effect on the viruses up to 7.5 decadal logarithmic levels. For the non-enveloped virus, inactivation was fastest on Douglas fir wood, with the target reduction for effective inactivation (reduction by factor 4.0 log10) being achieved after two weeks, and for the enveloped virus on pine wood, it was already achieved from the day of drying. The hygienic effects of the wood carriers may be due to their hygroscopic properties and wood constituents. These effects offer potential for further investigation, including tests with other wood species rich in extractives.

Effects of chemically-reductive trace gas contaminants on non-thermal plasma inactivation of an airborne virus.

Transmission of airborne infectious diseases poses great risk for public health and socio-economic stability, thus, there is a need for an effective control method targeting the spread and transmission of pathogenic aerosols. The existence of chemically-reductive trace air contaminants in animal agriculture may affect the oxidation inactivation process of pathogens. In this study, we report how the presence of such gasses impacts the effectiveness of using non-thermal plasma (NTP) within a packed-bed dielectric barrier discharge reactor to inactivate MS2 bacteriophage. Inactivation of the aerosolized bacteriophage is determined by the combination of viability and polymerase chain reaction assays. Using a plasma power source with a voltage of 20 kV and frequency of 350 Hz, after differentiating and excluding the physical removal effects of viral aerosols potentially caused by plasma, the baseline inactivation of MS2 aerosol in air has been determined based on an overall air flow rate of 200 Liters per minute and plasma discharge power of 1.8 W. When either ammonia or hydrogen sulfide gas is introduced into the airstream at a concentration of 1 part per million, the NTP virus inactivation efficiency is reduced to around 0.5-log from the 1-log baseline inactivation in air alone. Higher concentrations of those gasses will not further inhibit the effectiveness of plasma inactivation.

Systematic investigation and modeling prediction of virus inactivation by ozone in wastewater: Decoupling the matrix effects.

Water disinfection is undoubtedly regarded as a critical step in ensuring the water safety for human consumption, and ozone is widely used as a highly effective disinfectant for the control of pathogenic microorganisms in water. Although the diminished ozone efficiencies in complex water matrices have been widely reported, the specific extent to which individual components of matrix act on the virus inactivation by ozone remains unclear, and effective methodologies to predict the comprehensive effects of various factors are needed. In this study, the decoupled impact of the intricate water matrix on the ozone inactivation of viruses was systematically investigated and assessed from a simulative perspective. The concept of "equivalent ozone depletion rate constant" (k') was introduced to quantify the influence of different species, and a kinetic model was developed based on the k' values for simulating the ozone inactivation processes in complex matrix. The mechanisms through which diverse species influenced the ozone inactivation effectiveness were identified: 1) competition effects (k' = 105∼107 M-1s-1), including organic matters and reductive ions (SO32-, NO2-, and I-), which were the most influential species inhibiting the virus inactivation; 2) shielding effects (k' = 103∼104 M-1s-1), including Ca2+, Mg2+, and kaolin; 3) insignificant effects (k' = 0∼1 M-1s-1), including Cl-, SO42-, NO3-, NH4+, and Br-; 4) promotion effects (k' = ∼-103 M-1s-1), including CO32- and HCO3-. Prediction of ozone disinfection efficiency and evaluation of species contribution under complex aquatic matrices were successfully realized utilizing the model. The systematic understanding and methodologies developed in this research provide a reliable framework for predicting ozone inactivation efficiency under complex matrix, and a potential tool for accurate disinfectant dosage determination and interfering factors control in actual wastewater treatment processes.