Airborne disinfection of high-containment facilities before maintenance or between animal studies is crucial. Commercial spore carriers (CSC) coated with 10⁶ spores of Geobacillus stearothermophilus are often used to assess the efficacy of disinfection. We used quantitative carrier testing (QCT) procedures to compare the sensitivity of CSC with that of surrogates for non-enveloped and enveloped viruses including SARS-CoV-2, mycobacteria, and spores to an aerosolized mixture of peroxyacetic acid and hydrogen peroxide (aPAA/HP). We then used the QCT methodology to determine relevant process parameters to develop and validate effective disinfection protocols (≥ 4 log₁₀ reduction) in various large and complex facilities. Our results demonstrate that aPAA/HP is a highly efficient procedure for airborne room disinfection. Relevant process parameters such as temperature and relative humidity can be wirelessly monitored. Furthermore, we found striking differences in inactivation efficacies against some of the tested microorganisms. Overall, we conclude that dry fogging a mixture of aPAA/HP is highly efficient against a broad range of microorganisms as well as material compatible with relevant concentrations. Furthermore, CSC are artificial bio-indicators with lower resistance and thus, should not be used for validating airborne disinfection when microorganisms other than viruses have to be inactivated. IMPORTANCE Airborne disinfection is not only of crucial importance for the safe operation of laboratories and animal rooms where infectious agents are handled, but can also be used in public health emergencies like the current SARS-CoV-2 pandemic. We show that dry fogging a mixture of aPAA/HP is highly microbicidal, efficient, fast, robust, environmentally neutral, and a suitable airborne disinfection method. In addition, the low concentration of dispersed disinfectant, particularly for enveloped viral pathogens like SARS-CoV-2, entails high material compatibility. For these reasons and due to the relative simplicity of the procedure, it is an ideal disinfection method for hospital wards, ambulances, public conveyances and indoor community areas. Thus, we conclude that this method is an excellent choice for control of the current SARS-CoV-2 pandemic.

Manure from animal farms and sewage sludge contain pathogens and opportunistic organisms in various concentrations depending on the health of the herds and human sources. Other than for the presence of pathogens, these waste substances are excellent nutrient sources and constitute a preferred organic fertilizer. However, because of the pathogens, the risks of infection of animals or humans increase with the indiscriminate use of manure, especially liquid manure or sludge, for agriculture. This potential problem can increase with the global connectedness of animal herds fed imported feed grown on fields fertilized with local manures. This paper describes a simple, easy-to-use, low-tech hygienization method which conserves nutrients and does not require large investments in infrastructure. The proposed method uses the microbiotic shift during mesophilic fermentation of cow manure or sewage sludge during which gram-negative bacteria, enterococci and yeasts were inactivated below the detection limit of 3 log₁₀ cfu/g while lactobacilli increased up to a thousand fold. Pathogens like Salmonella, Listeria monocytogenes, Staphylococcus aureus, E. coli EHEC O:157 and vegetative Clostridium perfringens were inactivated within 3 days of fermentation. In addition, ECBO-viruses and eggs of Ascaris suum were inactivated within 7 and 56 days, respectively. Compared to the mass lost through composting (15–57%), the loss of mass during fermentation (< 2.45%) is very low and provides strong economic and ecological benefits for this process. This method might be an acceptable hygienization method for developed as well as undeveloped countries, and could play a key role in public and animal health while safely closing the nutrient cycle by reducing the necessity of using energy-inefficient inorganic fertilizer for crop production.

Background: Technical protection measures for laboratory activities involving biological agents include biological safety cabinets (BSC) that may be contaminated. In the case of diagnostic activities with SARS-CoV-2, this may also affect BSC that are operated at protection level 2; therefore, decontamination of all contaminated surfaces of the BSC may be required. In addition to fumigation with hydrogen peroxide (H2O2), dry fogging of H2O2-stabilized peroxyacetic acid (PM) represents another alternative to fumigation with formalin. However, to prove their efficacy, these alternatives need to be validated for each model of BSC. Methods: The validation study was performed on 4 different BSCs of Class II A2 using the "Mini Dry Fog" system. Results: An aerosol concentration of 0.03% PM and 0.15% H2O2 during a 30 min exposure was sufficient to inactivate SARS-CoV-2. Effective concentrations of 1.0% PAA and 5% H2O2 were required to decontaminate the custom-prepared biological indicators loaded with spores of G. stearothermophilus and deployed at 9 different positions in the BSC. Commercial spore carriers were easier to inactivate by a factor of 4, which corresponded to a reduction of 10(6) in all localizations. Conclusions: Dry fogging with PM is an inexpensive, robust, and highly effective decontamination method for BSCs for enveloped viruses such as SARS-CoV-2. The good material compatibility, lack of a requirement for neutralization, low pH - which increases the range of efficacy compared to H2O2 fumigation - the significantly shorter processing time, and the lower costs argue in favor of this method.

In the case of a notifiable animal disease like salmonellosis, manure is contaminated and must be disinfected. This can be performed using heat measures, chemical disinfectants, or long-term storage. All these measures bring along severe economic, ecological, and logistical problems. The aim of this study was to evaluate lactic acid fermentation (LAF) as an alternative disinfection method. Fermentation was started by adding a carbohydrate source to the manure and creating anaerobic conditions. For testing, cattle manure was enriched with different carbohydrate (CHO) sources and spiked with Salmonella Typhimurium (S. Ty.). The samples were incubated at 10 °C and 21 °C for 111 days (Exp1) and at 21 °C for 50 days (Exp2). The microbial shift was determined using cultural methods and MALDI-TOF. Both the change in pH and Enterococcus spp. were tested as suitable indicators. The results showed the different suitability of the selected CHO for hygienization by LAF. Using squeezed oat as an additive, S. Ty was reduced to below the detection limit under both temperature conditions within 21 days and 14 days. Additional saccharose decreased the reduction time. This study showed that LAF is a valuable alternative for disinfecting cattle manure in the case of bovine salmonellosis. Using this method, both manure and feed residues can be treated in one approach and afterwards be used as fertilizer.

Every year, billions of passengers, pets, and livestock are transported by air, which increases the risk of a global spread of pathogens like severe acute respiratory syndrome coronavirus 1 and 2, H1N1 influenza, the Ebola virus, and foot-and-mouth disease virus. However, due to its complexity and high safety standards, aircraft decontamination is limited to manual surface disinfection, and an approved method for disinfecting otherwise inaccessible areas is still unavailable. To address this issue, we present the development and validation of an effective room disinfection method utilizing low concentrations of aerosolized peroxyacetic acid (aPAA), stabilized by hydrogen peroxide, known as “dry fog.” Therefore, we determined the minimal effective antimicrobial concentration of aPAA for a wide range of agents, including bacteria, viruses, and bacterial endospores. Then, we applied the developed disinfection protocol to a large panel of relevant materials multiple times in the context of the official material compatibility tests (MCTs) requested by the European Union Aviation Safety Agency (EASA) to demonstrate that its application will not cause a negative effect on the aircraft airworthiness. After successfully passing these complex material compatibility tests, we finally validated the disinfection protocol within a narrow-body aircraft A320 for its antimicrobial efficacy by using the most resistant microorganisms to aPAA among our test organisms, the bacteriophage MS2, and spores of Geobacillus stearothermophilus . Thus, we are now able to present the first aerosol-based room disinfection method suitable for aircraft that could easily be adopted for other aircraft types and other means of mass transport as well. IMPORTANCE: Despite numerous outbreaks in recent years, particularly the Ebola virus epidemic and the severe acute respiratory syndrome coronavirus 2 pandemic, there is still no approved method for disinfecting contaminated aircraft. Due to the highly complex nature of aircraft material and technology, it is difficult to balance the antimicrobial efficacy and material compatibility of the biocides used in a way that meets high safety standards. Here, we demonstrate an aerosol-based disinfection protocol using a mixture of peracetic acid and hydrogen peroxide. This protocol passed all material compatibility tests according to relevant aircraft certification specifications established by the European Union Aviation Safety Agency, as well as the manufacturer qualification test programs, and proved to be highly effective against viruses, bacteria, and even bacterial endospores without compromising the airworthiness of the aircraft. By targeting both contaminated surfaces and objects, as well as airborne pathogen reservoirs, it minimizes the spillover risk of zoonotic agents and prevents the mechanical transmission of high-consequence agricultural pathogens into disease-free geographical regions. This makes it the first suitable semi-automatic airborne disinfection method that can be used in addition to routine manual surface disinfection following contamination resulting from contact with people, animals, or goods infected or contaminated with high-consequence pathogens.

Airborne disinfection of high-containment facilities is mandatory for safe operations. However, sewage vent air—particularly in large-animal experimental units—can represent a critical gap and is typically not considered in the design of high-efficiency particulate air (HEPA) filter systems. Here, we present an engineering design enabling HEPA filtration of sewage vent air from primary biological containments, including large-animal facilities and necropsy suites. We additionally evaluated vaporized hydrogen peroxide (vH ₂ O ₂ ) fumigation as an airborne disinfection approach. Disinfection efficacy was determined by quantitative carrier testing using commercial spore carriers (CSC) coated with 10 ⁶ spores of Geobacillus stearothermophilus and compared with germ carriers coated with surrogate viruses, mycobacteria, and other spores. Temperature, relative humidity (RH), and vH ₂ O ₂ concentration were monitored in real time to define process parameters and validate protocols targeting ≥4 log ₁₀ reduction. Inactivation efficacy differed substantially among challenge agents: CSC were least resistant to inactivation, whereas murine norovirus (MNV) on germ carriers was most challenging to inactivate. At medium-high vH ₂ O ₂ concentrations (up to 520 parts per million [ppm]), increasing RH was the dominant driver of inactivation. Based on these data, we developed a biphasic disinfection cycle maintaining a saturated humidity environment with ~35% H ₂ O ₂ for ≥10 h, achieving ≥4 log ₁₀ reduction of MNV infectivity and G. stearothermophilus spore viability on germ carriers. Overall, vH ₂ O ₂ under high-to-saturated RH is an effective means to disinfect technical infrastructure in biological containment settings handling high-consequence pathogens. IMPORTANCE: Vent air from sewage systems in biological containment facilities where large animals are kept should be considered a contaminated waste stream. Therefore, effective high-efficiency particulate air (HEPA) filter systems that prevent mold or other moisture-related deterioration of the filters need to be installed. Airborne vaporized hydrogen peroxide (vH ₂ O ₂ ) is an appropriate method to decontaminate these complex HEPA filter systems. High to saturated humidity is required for virucidal and sporicidal efficacy under medium to high concentration of vH ₂ O ₂ . Commercially available spore carriers are artificial in design, resulting in lower resistance to inactivation compared to custom germ carriers coated with non-enveloped viruses and thus should be avoided as read-out (validation means) by the biosafety community working with high-consequence pathogens in primary containments.