The Research Project AEROPORE

In the FFG-funded research project “Aeropore,” the Austrian Research Institute for Chemistry and Technology (OFI) together with the Center for Electron Microscopy Graz (ZFE), both members of ACR (Austrian Cooperative Research), have developed a novel filter test rig. This enables filter systems to be assessed not only on the basis of their permeability to particles in terms of number and size, but also according to their biological risk. Starting in autumn 2023, OFI expert Bernadette Führer will be researching in the project “AeroMobil” how this novel test facility can also be used in mobile applications.

not infect humans, but just bacteria. Thus, a high level of operational safety can be ensured. Bacteriophages might support and facilitate aerosolization studies and subsequently, prediction and estimation of behavior and reactivity of highly pathogenic airborne viruses.

As biological particles such as viruses behave differently and comprise detectable infectious potential compared to inert non-biological particles, it is important for the design and testing of filtration and ventilation systems to investigate the interactions between bioaerosols and filters as well as air conditioning and cleaning devices. These non-biological particles may represent the size of e.g. viruses but do not necessarily reflect the separation mechanisms on nanometric scale, which are often connected to physio-chemical characteristics of viruses, which cannot be simulated by an e.g. salt particle.

In general, it is known, that nonenveloped viruses are more persistent towards environmental stress such as temperature, relative humidity, irradiation via UV or disinfection agents, whereas in the case of enveloped viruses, additionally possessing a lipid membrane carrying, e.g. characteristic membrane spike proteins, are more sensitive towards environmental influencing factors. Furthermore, bacteria or fungal spores may change size and shape depending on cell membrane and wall characteristics as reaction environmental factors such as desiccation, temperature, humidity or the presence of surrounding carrier fluids, droplets or particles. The potential of applied bioaerosols lies furthermore in serving as extension tool to commonly used non-biological aerosols such as salt or DEHS particles for evaluation of potential reduction of biological risk by e.g. air filters or Heating, Ventilation and Air Conditioning (HVAC) systems.

Breathe Easy: Insight into the Research Project Aeropore

Within the research project “Aeropore” conducted at OFI, an independent testing and research institute in Austria and member of ACR (Austrian Cooperative Research), the subject was intensified. In the course of the project, it was possible to develop a pilot-scale simulation filter test rig (at right) for testing air filters with airborne biological hazards belonging to biological safety level 1 (BSL1).

Even before COVID-19 pandemic the topic air hygiene and biological safety played a major role for OFI. Thus, the development of reproducible and comparable methods allowing the performance of single-pass tests using bioaerosols (determination of filter passage, fresh “air mode”) for evaluation of air filters from a biological and immunological perspective could be enabled.

Whereas everything started with a focus on allergens, pollen and fungal spores, the testing scope has been extended further during the course of the project. Working with bioaerosols containing bacteria and viruses used for filter testing was enabled at OFI within a second step, whereas the COVID-19 pandemic definitely enhanced transparency and attention on the topic.

It is known, that handling airborne particles functioning as test bioaerosol can be challenging. Each microbial strain or bioparticle requires proper preparation, sampling, aerosolization and detection techniques due to different characteristics and structural features of the biological particles, especially in airborne state. In addition, capability to survive as well as to maintain specific biological activity in airborne state is required by applied microbes for being able to detect them.

As there are very strict safety regulations when working with highly patho-

 For the investigation of air filters, this filter test rig was developed at OFI, which can also be used to classify filter systems in terms of their separation performance for biological hazardous substances. OFI genic and infectious microorganisms such as SARS-CoV-2, influenza viruses, Ebola or Mycobacterium tuberculosis at laboratory, many research studies recommend safe and non-pathogenic models for human viruses.

To give an example, commonly used virus surrogates for aerosol experiments are MS2 bacteriophage and phi6 bacteriophage, which are also used at OFI for filter and air cleaner evaluation. MS2 bacteriophage, a well-known bacterial virus with an average size of 27 nm, was chosen as representative for a robust non-enveloped virus comparable to noroviruses. On the other hand, phi6 bacteriophage with an average size of 86 nm was selected as enveloped virus posing as a surrogate for Ebola virus or Coronavirus.

During method development, both viral surrogates were compared regarding their stability and recovery as airborne particles for testing air filters within

What is a Bioaerosol?

If an aerosol mixture contains biological material or microorganisms, the term bioaerosol or biological aerosol is used. Such a biological aerosol consists of living and non-living components or fragments such as pollen, fungi, bacteria and viruses. These differ in size:

• Size of fungal spores: most commonly observed size range of spores is 2 to 10 μm but can vary from around 1 to 50 μm

• Size of bacteria: 0.5 up to 4 μm is assumed depending on bacteria species as well as sampling and analysis techniques

• Size of viruses: 15-500 nm

• Size of pollen: average between 25 and 40 µm, subpollen particles (“Ubisch bodies” ~0.5 µm)

Testing with real biologically active viruses, bacteria or allergenic particles, especially at large scales can be challenging and complex. However, they could support better understanding of infection control and pathogen transmission, especially for risk minimization instruments such as filtration systems in health care settings.

OFI filter test rig. For checking the suitability of the selected strains as a stable bioaerosol recovery and viability after aerosolization into the test rig were investigated. Liquid aerosol generator from Topas GmbH was used for aerosolization of selected bioaerosols at different volume flow rates ranging from 40-300 m³/h for defined period of time per run. The nebulized airborne viruses sprayed into the test duct were sampled via collection filter media mounted at the end of the test duct followed by liquid extraction, which is just one of several bioaerosol sampling techniques.

The quantification of the viruses is usually done via double layer method by optically counting plaque forming units (PFU) being the golden standard for detection of bacteriophages. The method requires incubation of the host bacteria together with the sample solution containing the target virus at specific conditions. So-called plaques are formed by present bacteriophages as a result of infecting host bacteria, grown

References: on agar plates containing nutrient media. For evaluation of retention capacity or filtration efficiency of air filters regarding hazardous bioparticles, tests with and without a test filter, (a cabin air filter) are done.

Following general issues were important to consider during development of test bioaerosols applied within a filter test rig:

1. Stability and recovery of viruses, bacteria, fungal spores or allergenic pollen on surfaces.

2. Stability and recovery of airborne particles in airborne state after aerosolization within whole test duct.

3. Checking suitability of bioaerosol for single-pass tests for air filters, filter media and air cleaners.

More Research is Needed

The application of bioaerosols for a standardized air filter evaluation is still a black box within aerosol research and filter testing, which could be further opened for supporting estimation and

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In the future, OFI will deal with this topic even more intensively: in the autumn of 2023, the research project “AeroMobil” will start. The aim is to facilitate the assessment of air purification measures in different room concepts by developing a mobile test station.

Biotechnologist Bernadette Führer has been working in applied research at Austrian Research Institute for Chemistry and Technology (OFI) since 2017, where she specializes in filter analysis and air hygiene. She contributes her know-how to various RDI projects: Within the project "Aeropore", she was instrumental in setting up a new methodology with which air filters can be evaluated using allergens, viruses and bacteria in a particularly realistic way. Visit www.ofi.at. Reach Bernadette Führer at +43 798 16 01 633 or bernadette.fuehrer@ofi.at.

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