Fundamentals of Bioaerosols Science: From Physical to Biological Dimensions of Airborne Biological Particles

By Naomichi Yamamoto

Fundamentals of Bioaerosols Science: From Physical to Biological Dimensions of Airborne Biological Particles covers both the physical and biological aspects of bioaerosol science. It is assumed that researchers with a physics background are often unfamiliar with the biological aspects (e.g., molecular biology, PCR, and DNA sequencing), and researchers with a biology background are often unfamiliar with the physical aspects (e.g., aerosol physics, air sampling, and aerodynamic diameter) of bioaerosol science. This book aims to bridge the interdisciplinary gap between the fields of bioaerosol science. Fundamentals of Bioaerosols Science include topics such as bioaerosol physical properties, sampling and monitoring methods, analytical methods, control techniques, and relationship to climate.

• Presents an in-depth explanation of the fundamentals of bioaerosols science
• Includes an introduction to the latest knowledge and technologies related to bioaerosol science
• Features interdisciplinary contents that are useful even for those without specialized knowledge

• Language: English
• Publisher: Elsevier Science
• Release date: May 24, 2023
• ISBN: 9780323859707

Naomichi Yamamoto

Dr. Naomichi Yamamoto is a Professor in the Department of Environmental Health Sciences at Seoul National University (SNU). His current research interests are in the areas of i) aerobiology, ii) environmental microbiome, and iii) One Health. His research team uses genomics approaches to analyze environmental DNA (eDNA) to investigate the ecological roles of organisms, including airborne organisms, in the environment and their impacts on human health. Before joining to SNU faculty in 2012, Dr. Yamamoto worked at Yale School of Engineering as a postdoctoral fellow supported by the Japan Society for the Promotion of Science (JSPS). He received his B.Eng. degree in applied physics from Waseda University, M.S. degree in environmental health sciences from UCLA, and Ph.D. degree in environmental studies from the University of Tokyo.

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Front Cover for Fundamentals of Bioaerosols Science - From Physical to Biological Dimensions of Airborne Biological Particles - 1st edition - by Naomichi Yamamoto

Fundamentals of Bioaerosols Science

From Physical to Biological Dimensions of Airborne Biological Particles

Naomichi Yamamoto

Department of Environmental Health Sciences, Seoul National University (SNU), Seoul, South Korea

Table of Contents

Cover image

Title page

Copyright

Chapter 1. Introduction

Abstract

1.1 Overview

1.2 Definitions of terms

1.3 Historical background

References

Chapter 2. Taxonomy

Abstract

2.1 Overview

2.2 Basic taxonomy

2.3 Viruses

2.4 Bacteria

2.5 Fungi

2.6 Plants

2.7 Arthropods

2.8 Vertebrates

References

Chapter 3. Physical properties

Abstract

3.1 Overview

3.2 Physical quantities

3.3 Equivalent diameters

3.4 Gravitational settling

3.5 Aerodynamic equivalent diameter

3.6 Inertia

3.7 Diffusion

3.8 Respiratory aerosols

3.9 Droplet physics

3.10 Human respiratory deposition

Appendix 1–3 Supporting information

References

Chapter 4. Human health impacts

Abstract

4.1 Overview

4.2 Basics of airborne and droplet-borne infectious diseases

4.3 Viral infections

4.4 Bacterial infections

4.5 Fungal infections

4.6 Drug resistance in bioaerosols

4.7 Basics of airborne allergic diseases

4.8 Pollen allergy

4.9 Fungal allergy

4.10 Arthropod allergy

4.11 Vertebrate allergy

4.12 Toxins

References

Chapter 5. Principles of sampling and monitoring methods

Abstract

5.1 Overview

5.2 Impactor

5.3 Filter

5.4 Air inlet

5.5 Sedimentation sampling

5.6 Precipitation sampling

5.7 Real-time monitoring

References

Chapter 6. Analytical methods

Abstract

6.1 Overview

6.2 Microscopy

6.3 Culture

6.4 Bioassays

6.5 Nucleic acid-based methods

6.6 Hyphenated methods

References

Chapter 7. Control technologies

Abstract

7.1 Overview

7.2 Filtration

7.3 Respirators and face masks

7.4 Ventilation

7.5 Ultraviolet germicidal irradiation

7.6 Photocatalytic oxidation

7.7 Electrostatic precipitator and ionizer

References

Chapter 8. Bioaerosols in built and natural environments

Abstract

8.1 Overview

8.2 Bioaerosols in indoor environment

8.3 Bioaerosols from industrial sources

8.4 Antimicrobial-resistant microorganisms and antimicrobial resistance genes

8.5 Genetically modified plants and herbicide-resistant weeds

8.6 Biogeochemical cycles

8.7 Climate system

References

Index

Copyright

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Chapter 1

Introduction

Abstract

This chapter defines key terms related to bioaerosols and introduces the historical background of bioaerosol research. Historical advances in bioaerosol research are closely linked to the invention of microscopy in the late 16th century and the establishment of microbial culture methods in the late 19th century. In addition, early research on airborne pollen and fungal spores around the 19th century to identify the causes of hay fever also contributed greatly to advances in bioaerosol research. More recently the rise of molecular biology and the invention of related techniques, such as polymerase chain reaction and DNA sequencing, have also revolutionized it. This chapter reviews the history of bioaerosol research and the historical events that influenced it.

Keywords

Aerobiology; bioaerosols; Charles Harrison Blackley; life; primary biological aerosol particles (PBAPs); Richard Leach Maddox; Walter Hesse

1.1 Overview

This chapter defines key terms related to bioaerosols and introduces the historical background of bioaerosol research. Historical advances in bioaerosol research are closely linked to the invention of microscopy in the late 16th century and the establishment of microbial culture methods in the late 19th century. In addition, early research on airborne pollen and fungal spores around the 19th century to identify the causes of hay fever contributed greatly to advances in bioaerosol research. More recently the rise of molecular biology and the invention of related techniques, such as polymerase chain reaction (PCR) and DNA sequencing, have also revolutionized it. This chapter reviews the history of bioaerosol research and the historical events that influenced it.

1.2 Definitions of terms

Below are definitions of key terms that appear frequently in the area of bioaerosols science. The terms may be used with different meanings depending on the context, but unless otherwise noted, the following definitions shall apply.

1.2.1 Bioaerosols

The term bioaerosol can be divided into two parts: bio and aerosol. The word aerosol can be further divided into two parts: aero and sol. The prefix aero- means air in Latin. The suffix -sol is German for a dispersion system in which a solid or liquid is the dispersoid and a fluid, such as a liquid, or gas is the dispersion medium. Therefore the term aerosol refers to a dispersion system in which solid and/or liquid particles are the dispersoids and the air is the dispersion medium. In the context of aerobiology, the air here can be defined as a gas composed of ~78% N2, ~21% O2, ~1% trace gases, and water vapor.

The term bioaerosol can be defined as an aerosol originating from life, as the prefix bio- comes from the Greek word bios, which means life. Therefore the term bioaerosol refers to a dispersion system in which particles derived from life (biological particles, see below for their definition) are the dispersoids and the air is the dispersion medium. Examples of biological particles include virus particles, bacterial cells, fungal spores, and pollen grains. The size of bioaerosol particles typically ranges from 0.5 to 100 μm (Hirst, 1995). Particles of this size can exist stably in the air for at least a few seconds.

1.2.2 Life

As defined above, bioaerosols are aerosols of particles derived from life. In this book, as particles derived from life, I will include viruses, bacterial cells, fungal spores, pollen grains, and so on. However, it may also be worth knowing that there is no consensus definition of life, and several definitions have been proposed. One famous definition is based on the concept of negative entropy proposed by the physicist Erwin Schrödinger (1887–1961) in his book What Is Life? (1944). Entropy is a physical quantity that describes the disorder of things. Entropy usually increases (according to the second law of thermodynamics), which means that disorder also increases. In life, an increase in entropy, or an increase in disorder, means decay and death. According to Schrödinger, life is defined as something that evades decay by maintaining organization by extracting order from the environment, exerting negative entropy. He also wrote that to avoid decay, metabolism is necessary. In fact, the ability of metabolisms is often cited as one of the requirements of life along with others (e.g., replication). However, by that definition, viruses are considered nonliving because they lack metabolic ability, and in fact, some researchers argue that viruses are nonliving (Moreira and López-García, 2009; López-García, 2012). On the other hand, some researchers consider viruses as life based on their ability to replicate (Koonin and Starokadomskyy, 2016). It has also been proposed by researchers to define viruses as capsid-encoding organisms and eukaryotes and prokaryotes as ribosome-encoding organisms (Raoult and Forterre, 2008). As such, there is no consensus on whether viruses should be considered life, but in this book, like many other bioaerosol-related books ever published (American Conference of Governmental Industrial Hygienists, 1999; Burge, 1995; Cox and Wathes, 1995; Delort and Amato, 2017; Jonsson et al., 2014; Kowalski, 2006), viruses will be considered life.

1.2.3 Biological particles

In this book, the term biological particles refers to particles that are directly dispersed from a source in the form of solid particles derived from life or droplets containing them without undergoing a significant chemical reaction. This term will be used synonymously with the term primary biological aerosol particles (PBAPs) commonly used by researchers (Matthias-Maser and Jaenicke, 1995; Després et al., 2012). According to Matthias-Maser and Jaenicke (1995), PBAPs are defined as airborne solid particles derived from organisms including microorganisms (dead or alive) and their fragments. In this book, I define biological particles as equivalent to PBAPs. Thus examples of biological particles include viruses, bacteria, fungal spores, pollen grains, arthropod feces and fragments, and animal hair and skin fragments.

Organisms, including humans, produce organic gases. For example, humans are known to emit organic gases, such as acetaldehyde and acetone, from their skin (Sekine et al., 2007). Similarly, microorganisms, such as bacteria and fungi, emit volatile organic compounds, such as alcohols, esters, ethers, ketones, terpenoids, geosmin, and dimethyl disulfide, which are collectively called microbial volatile organic compounds (Korpi et al., 2009). Furthermore, plants emit organic gases, such as isoprene, monoterpenes, and sesquiterpenes, which are collectively called biogenic volatile organic compounds (Hallquist et al., 2009). These volatile organic compounds emitted from living organisms can undergo chemical reactions in the air and transform into particulate matter called biogenic secondary organic aerosol (BSOA). BSOA can also be considered biological particles but are not covered in this book. In addition, particles generated by burning biomass, such as wood, may be considered biological, as they are of biogenic origin. However, they are also not covered in this book, as they undergo significant chemical reactions.

1.2.4 Droplets

In this book, the term droplet refers to liquid water particles. Droplets may contain solid matter and/or other impurities, which may be biological. The residue left after the droplet water evaporates is called the droplet nucleus. In the context of infectious diseases, for example, according to the World Health Organization (2014), respiratory aerosols with a diameter of >5 µm are defined as droplets, and those with a diameter of ≤5 µm are defined as droplet nuclei. In this book, regardless of their size, liquid water particles are called droplets. This is due to the fact that the droplet size changes rapidly due to evaporation (see Chapter 3). Therefore it seems difficult to define droplets based on their size.

1.2.5 Aeroallergens

Aeroallergens are inhalable allergens that are suspended in the air. Allergens are antigens that cause allergies mediated by immunological mechanisms (Johansson et al., 2001). For a definition of allergies, see Chapter 4. The term inhalant allergen is also used almost synonymously with aeroallergen. However, in the literature, aeroallergens tend to refer to outdoor allergens (e.g., pollen and fungus), while inhalant allergens tend to refer to indoor allergens (e.g., house dust mite, cat, and dog).

1.2.6 Aerobiology

Aerobiology is the study of life and its derivatives in the air. Its scope includes studies on their behavior and impacts, as well as on their sampling, analysis, and control methods. The impacts include human health impacts and environmental impacts. Due to its interdisciplinary nature, the study of aerobiology requires knowledge of a wide range of disciplines, such as physics and biology.

1.3 Historical background

Behind the historical evolution of the field of aerobiology are the inventions of microscopy in the late 16th century and microbial culture in the late 19th century. In addition, early research on airborne pollen and fungal spores, which began around the 19th century to identify the causative agents of hay fever, contributed greatly to the progress of aerobiology. More recently the advent of molecular biology and the invention of related technologies, such as PCR and DNA sequencing, also revolutionized it. Below, we outline the relationship between these historical events and advances in the field of aerobiology.

1.3.1 Early research on airborne organisms by microscopy

Microscopes are analytical instruments that magnify objects that are too small to be seen with the naked eye, and the invention of microscopes has contributed greatly to research in the field of aerobiology. Historically, the invention of microscopes has been associated with the advancement of the field of optics from ancient times, as they use properties of light, such as refraction to magnify objects. For example, historical figures who have contributed to the advancement of the field of optics include the Greek mathematician Euclid (c.300 BCE) who taught the straightness of light, the ancient Roman scholar Claudius Ptolemy (c.100–c. 170) and the Arab scholar Ibn al-Haytham (c.965–c.1040) who explained the phenomenon of refraction (Croft, 2006), and the English philosopher Roger Bacon (c.1220–c.1292) who explained the mechanism of magnification by optical lenses. Correspondingly, optical lenses have also been used in a variety of applications, such as spectacles, telescopes, and microscopes. One theory holds that the first compound microscope (one with multiple lenses) was invented around 1595 by the Dutch spectacle-maker Sacharias Jansen (1585–1632) and his father Hans Jansen (Croft, 2006).

Microscopes began to be used to observe microscopic organisms. Notable early microscopists who observed microscopic organisms included the English scientist Robert Hooke (1635–1703). Hooke, who is also famous for his law of elasticity in physics, commonly known as Hooke’s law, is known as a person who discovered the cell by observing the structure of cork microscopically and calling its microscopic porous structure a cell (Hooke, 1665). Hooke also microscopically observed and illustrated aerobiologically related organisms, such as mold and mite (Fig. 1.1). The mold, which he called blue Mould (Fig. 1.1A), was found on the red covers of a small book as white spots like hairy sheep’s skin (Hooke, 1665) and is now known to be a species of Mucor (Gest, 2004). Mucor, which causes a fungal infection called mucormycosis, is known to be found in indoor air (Takahashi, 1997; Shelton et al., 2002). Another indoor microscopic creature illustrated by Hooke included mites (Fig. 1.1B). Hooke explains that the mite resides on all kinds of moldy substances, which is consistent with the mold–mite interactions uncovered today. Also, as trivia, the mite that Hooke illustrated is now known to be a species of Tyrophagus, which is known to inhabit buildings, such as barns and houses (Jervis, 2014). Tyrophagus is a type of mite called storage mite and is thought to be a possible cause of allergies, especially in agricultural farmers (Solarz and Pająk, 2019). The remaining indoor microscopic organism illustrated by Hooke included a small insect that was walking on the book he was reading, which he called Crab-like Insect. The Crab-like Insect is now known to be the house pseudoscorpion (Chelifer cancroides), which is commonly found in indoor environments, such as barns and houses (Jervis, 2014). In addition to molds and mites, pollen grains, which are also aerobiologically important, were illustrated under microscopes by early researchers, such as the Italian physician and biologist Marcello Malpighi (1628–94) and the English physician and plant anatomist Nehemiah Grew (1641–1712) (de Klerk, 2018). Pollen grains illustrated by Grew (1682) are shown in Fig. 1.2.

Figure 1.1 Micrographs of (A) blue Mould and (B) mites. Source: Reprinted from Hooke, R., 1665. Micrographia, or, some physiological descriptions of minute bodies made by magnifying glasses: with observations and inquiries thereupon, printed by J. Martyn and J. Allestry, printers to the Royal Society, London, UK, contributed by Missouri Botanical Garden, Peter H. Raven Library, in public domain.

Figure 1.2 Micrographs of pollen grains. Source: Reprinted from Grew, N., 1682. The Anatomy of Plants, printed by W. Rawlins, for the author, London, contributed by Missouri Botanical Garden, Peter H. Raven Library, in public domain.

Microscopy has also been used to observe airborne microscopic organisms, with early attempts to observe those in atmospheric deposits (both wet and dry deposits). For example, the Dutch scientist Antonie van Leeuwenhoek (1632–1723) used his unique single-lens microscope to study microscopic organisms (he called them animalcula and living atoms) in rainwater. In his famous letter sent to the Royal Society on 9 October 1676, van Leeuwenhoek reported a series of his observations on microscopic organisms in water, including rainwater (van Leeuwenhoek, 1677). In one of his observations, he collected fresh rainwater by placing a large cleaned porcelain dish on top of a wooden vessel about a foot and a half above the ground to prevent splashing mud from the ground. He reported that no living creatures were found in the rainwater on the day of collection, but that there were small, highly transparent animalcula in the same rainwater left for several days. His observation implied that animalcula were present in the rainwater and multiplied over several days. Of course, the possibility of microbial contamination originating from the dish or elsewhere cannot be ruled out since there was no sterilization technology at that time. Nevertheless, his contribution is enormous. In fact, Antonie van Leeuwenhoek is considered the discoverer of bacteria, because the little animalcula he observed were so small and reported to be 1/1000 the size of a daphnid (van Leeuwenhoek, 1677), so they were likely bacteria. He is considered the first person to observe bacteria.

In addition to van Leeuwenhoek’s observations of animalcula in rainwater (wet deposits), the German microscopist Christian Gottfried Ehrenberg (1795–1876) also reported observations of microscopic organisms in the fine dust (dry deposits) that Charles Darwin (1809–82) collected on board the Beagle, which arrived off the coast of Cape Verde in January 1833. Cape Verde is an island nation in the Atlantic Ocean off the west coast of Northwest Africa. According to Darwin (1846), it was reported that large amounts of fine dust fell on vessels navigating the neighborhood at that time. Darwin also reported that the atmosphere was often hazy, with the fine dust falling almost constantly. He collected the fine dust that fell on the Beagle and reported it to be excessively fine and of a reddish-brown color. Then, the fine dust sample was sent to Ehrenberg and was found to contain large amounts of infusoria and phytoliths (Darwin, 1846). The fine dust from Northwest Africa that fell on the Beagle is now commonly referred to as Saharan dust. Interestingly, Ehrenberg (1849) reported, in his book, observations of microscopic organisms not only in Saharan dust (which he called Atlantic trade wind dust) but also in sirocco dust. Sirocco dust refers to dust that reaches Mediterranean countries, such as Italy, across the Mediterranean Sea from the Sahara desert. As a side note, the fine dust collected by Darwin, which was kept in a museum in Germany, was recently analyzed by geochemical and molecular biological methods (Gorbushina et al., 2007). The study confirmed that the fine dust did indeed come from Western Sahara and revealed that microbes, such as bacteria and fungi, that are still alive adhere to the dust. The results of this study indicate that microbes in Saharan dust can be transported long distances from the continent of Africa to, for example, the Americas, while staying alive for long periods of time.

Another interesting early study on microorganisms in atmospheric deposits was reported by the American physician James Henry Salisbury (1823–1905). In his paper, he reported that in May of 1862 patients with intermittent fevers began appearing in the areas of Ohio and Mississippi valleys (Salisbury, 1866). He also reported the detection of algoid-type cells in the patients’ salivary secretions and mucus. To confirm that this algoid-type cell was responsible, he placed glass plates horizontally about 1 ft above over the areas’ marshy grounds. Under the microscope, he observed these algoid cells on the plates. This disease-causing microalga he discovered is known to be a species of Palmella, and he named it Gemiasma, meaning Earth miasma.

Not only microorganisms in atmospheric deposits but also microorganisms in the air collected using air samplers were reported, for example, by the English photographer and physician Richard Leach Maddox (1816–1902) in the 1870s. Maddox is famous for his invention of gelatino-bromide emulsion and dry photographic plate and made significant contributions to the field of photomicrography, photography using microscopy (Eyre, 1904). However, Maddox also made enormous contributions to the field of aerobiology, such as development of an air sampler called the Aëroconiscope to study airborne microorganisms. Fig. 1.3 shows his drawing of the Aëroconiscope (Maddox, 1870). The Aëroconiscope is based on the impactor principle (see Chapter 5) but uses wind instead of drawing air with a pump for the inertial impaction of aerosol particles. The Aëroconiscope has the structure of a hollow wind vane, in which there is an air acceleration nozzle and a clean thin cover glass for inertial impaction of accelerated aerosol particles. The cover glass is coated with a viscous substance, such as glycerin or purified treacle containing potassium acetate, and the collected microorganisms are observed with or without culture under a microscope. Fig. 1.4 shows an illustration of uncultured and cultured fungi collected from the air by the Aëroconiscope (Maddox, 1871). As we notice, there is a significant difference in the types of microscopic organisms observed by Maddox and Ehrenberg, with Ehrenberg observing mostly infusoria and phytoliths but few fungal spores in atmospheric deposits he analyzed, while Maddox observing numerous fungal spores in samples taken directly from the air. This is likely due in part to sampling bias due to the difficulty of small particles, such as fungal spores, to settle to the ground by gravity and demonstrates the importance of direct air sampling for efficient collection of small particles, such as fungal spores, in the air. Also, the Aëroconiscope developed by Maddox was used by other researchers. For example, the Scottish doctor David Douglas Cunningham (1843–1914) used a modified Aëroconiscope to study the spread of epidemics, such as cholera, and their relationship with airborne microorganisms in India and reported that there was no relationship (Cunningham, 1873). As an aside, in Cunningham (1873), in addition to the Aëroconiscope developed by Maddox, an air sampler called the Aëroscope is also introduced. The Aëroscope was developed by the French naturalist Félix Archimède Pouchet (1800–72) earlier and differs from the Aëroconiscope in that it uses a water aspirator as the driving force for air suction (Olivier, 1883).

Figure 1.3 Air sampler called the Aëroconiscope developed by Maddox (1870). Source: Reprinted from Maddox, R.L., 2011. On an apparatus for collecting atmospheric particles. Monthly Microscopical Journal 3, 286, Copyright (2011), with permission from John Wiley and Sons.

Figure 1.4 Microscopic illustration of uncultured and cultured fungi collected from the air by the Aëroconiscope by Maddox (1871). Source: Reprinted from Maddox, R.L., 2011. Observations on the use of the Aëroconiscope, or air-dust collecting apparatus. Monthly Microscopical Journal 5, 48, Copyright (2011), with permission from John Wiley and Sons.

The 20th century saw the development of various air samplers for microscopy. One example is the Hirst spore trap (Hirst, 1952) (Fig. 1.5). The Hirst spore trap is a type of slit-to-glass impactor (see Chapter 5 for details). In the Hirst spore trap, air is accelerated as it passes through a narrow orifice (slit), and the accelerated aerosol particles are inertially collected on a Vaseline-coated glass slide. After air sampling, the pollen and fungal spores collected on the glass slide are examined under a microscope. Slit-type impactors, such as the Hirst spore trap, have a structure in which the position of the collection substrate (e.g., slide glass) relative to the orifice (slit) moves over time. This allows the particles to be collected more uniformly on the substrate. Another historical air sampler for microscopy is the Rotorod samplers (Leighton et al., 1965). The Rotorod samplers have also been used to investigate airborne pollen and fungal spores (Magill et al., 1968). The Rotorod sampler collects airborne particles inertially by spinning a thin rod collector connected to a motor at high speed. The rod collector is greased. After air sampling, pollen and fungal spores collected on the collector are examined under a microscope.

Figure 1.5 Hirst spore trap. The part m represents the orifice that draws in air and accelerates it. The part n represents a glass slide, where aerosol particles drawn through the orifice are inertially collected. The glass slide moves slowly over time. Source: Reprinted from Hirst, J.M., 2008. An automatic volumetric spore trap. Annals of Applied Biology 39, 259, Copyright (2008), with permission from John Wiley and Sons.

1.3.2 Early research on aeroallergens and respiratory allergies

Early research on aeroallergens and respiratory allergies has also contributed significantly to the field of aerobiology. It was not until the 20th century that the word allergy was introduced and its mechanisms became clearer. However, research into respiratory allergies has been conducted since at least the early 19th century in the form of examining the relationship between hay fever and airborne pollen and fungal spores. Briefly, the word allergy is thought to be introduced by the Austrian pediatrician Clemens von Pirquet (1874–1929) early 20th century, originating from the words ergeia meaning reactivity and allos meaning altered (von Pirquet, 1911). Currently allergy is defined as hypersensitivity reactions triggered by the immune system and includes both immunoglobulin E (IgE)-mediated and non-IgE-mediated reactions (Johansson et al., 2001). Historically, IgE was discovered by Ishizaka et al. (1966) and Johansson and Bennich (1967) in the 1960s, and IgE-mediated hypersensitivity, known to include allergic asthma and allergic rhinitis (e.g., hay fever), is commonly referred to as type I hypersensitivity. In addition to type I hypersensitivity, those associated with respiratory allergies include types III and IV hypersensitivities that cause hypersensitivity pneumonitis (e.g., farmer’s lung and bird fancier’s lung). Hypersensitivity pneumonitis is mainly mediated by IgG, and IgG was first reported by Tiselius and Kabat (1939) in the late 1930s. However, even before these discoveries, research on respiratory allergies had been conducted, and research on hay fever in particular had been conducted since the 19th century.

Hay fever has been scientifically reported since at least the 19th century. For example, the English physician John Bostock (1772–1846) reported a patient with the initials J.B. (himself) who suffered from eye irritation, runny nose, sneezing, and chest tightness from early or mid-June to late July each year (Bostock, 1819). He also reported that in a year when the patient was confined to his home for 6 weeks during the summer months, the patient had far fewer symptoms than in other years. About a decade later, Bostock (1828) reported trends in about 30 patients with similar symptoms in his second paper. He proposed naming the disease catarrhus æstivus (summer catarrh) because it occurs in summer. In addition, Bostock reported that although he himself believed that exposure to heated air and sunlight and physical activity were the causes of the summer catarrh, many patients felt that the effluvium from hay was the cause and it had hence had the popular name of the hay fever.

After that, the English physician Charles Harrison Blackley (1820–1900), who himself suffered from hay fever for more than two decades, conducted a series of systematic experiments on hay fever and clarified that grass pollen was the cause (Blackley, 1873). In one of his experiments, pollen or its decoction was intentionally applied or inhaled to the respiratory mucosa of patients (such experiments are now called challenge trials) and observed for responses. He reported that patients developed symptoms similar to summer catarrh when exposed to pollen of grasses, such as rye grass (Lolium perenne), meadow foxtail (Alopecurus pratensis), rye (Secale cereale), wheat (Triticum sativum), oat (Avena sativa), and barley (Hordeum distichum). These grasses belong to the family Poaceae (Gramineae). He also devised a pollen trap (settle plate) to investigate pollen abundance in the air (Fig. 1.6). Four slips of glass coated with fluid were placed on the pollen trap, and the deposited pollen was observed under a microscope after 24 h. Using this device, Blackley investigated the seasonality of pollen dispersal and reported that it began in early June, peaked around the end of June, and ended in late July and that the observed seasonal trend corresponded well with the seasonal trend in the intensity of disease symptoms. He also reported that about 95% of the pollen observed under the microscope belonged to the family Poaceae. These findings strongly suggested that grass pollen, among many other environmental factors (e.g., heated air, sunlight, ozone, and dust) that Blackley investigated, was responsible for summer catarrh. In addition to these discoveries, Blackley made unique attempts such as developing a personal air sampler that collects airborne pollen by holding it in the mouth like a pipe and inhaling the air, and a kite equipped with glass slips to measure the amount of pollen at high altitudes in the atmosphere.

Figure 1.6 Pollen trap devised by Blackley (1873). Four slips of glass coated with a fluid are placed at locations A. Source: Reprinted from Blackley, C.H., 1873. Experimental Researches on the Causes and Nature of Catarrhus æstivus (Hay-Fever or Hay-Asthma), Bailliere, Tindall & Cox, London, contributed by Royal College of Physicians in Edinburgh, in public domain.

At about the same time in the 1870s as Blackley (1873) reported the summer catarrh, that is, hay fever, in England, the American physician Morrill Wyman (1812–1903) reported the catarrhus autumnalis (autumnal catarrh), different from the summer catarrh also known as hay fever, rose catarrh, or June cold, in the northern part of the United States (Wyman, 1872). Wyman, who also suffered from symptoms, reported that the autumnal catarrh occurs in a different season than the summer catarrh. He reported that the symptoms of autumnal catarrh start around August 20th every year, gradually diminish in the fourth week of September, and almost disappear at the end of September. He noticed that the season of symptoms nearly coincided with the flowering of the Roman wormwood (Ambrosia artemisiaefolia, synonymous with A. artemisiifolia). The Roman wormwood, now better known as the common ragweed, flowers in mid-August or a little later, constantly shedding large amounts of fine pollen until it ceases in late September. Wyman conducted challenge trials in which multiple subjects, including his son and himself, were exposed to pollen of the Roman wormwood. He and his son showed symptoms of the autumnal catarrh, but, overall, the results were mixed. These findings led him to a general impression that the Roman wormwood was the cause of the disease, but he also noted that it may not be a cause of the whole disease.

In the 20th century pollen was recognized as the cause of these catarrhs (Dunbar, 1913). Shortly thereafter, hay fever came to be considered a type of allergy (Sormani, 1916). In addition to grass (e.g., rye grass) and weed (e.g., ragweed) pollens, tree pollen has been reported to cause allergies. For example, in California, a patient was reported to be allergic to pine pollen (Rowe, 1939). In Sweden, in addition to pollen allergy caused by grass pollen in summer, pollen allergy caused by tree pollen, such as birch (Betula verrucosa) and alder (Alnus glutinosa), in spring has been reported (Juhlin-Dannfelt, 1948). In Japan, there have been many reports of allergies to cedar (Cryptomeria japonica) pollen since the 1960s (Araki, 1960; Araki, 1961). In the 1980s an increase in pediatric asthma caused by house dust mites was reported in countries, such as the United Kingdom (Smith et al., 1969), Japan (Kabasawa et al., 1976), and Australia (Clarke and Aldons, 1979). It is thought that the increase in asthma prevalence in these countries was attributed to improved insulation and airtightness of homes, resulting in indoor environments that are warmer and therefore favorable for the growth of house dust mites and that are more likely to accumulate their feces and debris (Platts-Mills, 2015). Moreover, the problem of allergic asthma in children in urban cities, known as inner-city asthma, caused by cockroaches has become apparent, particularly since the 1990s (Rosenstreich et al., 1997). Since the beginning of the 21st century, although the ownership rates of pet animals, such as cats and dogs, have not changed, it has been reported that the rates of allergic sensitization to them have increased in countries, such as Sweden (Rönmark et al., 2009). This indicates an increase in allergic disease, even though exposure to allergens remains unchanged. As suspected causes, the increased use of vaccinations and antibiotics and lifestyle changes are pointed out (Platts-Mills, 2015).

1.3.3 Early research on culturing airborne microorganisms

According to Koch’s postulates (see Chapter 4), isolation and pure culture of the pathogen is essential to identify the disease-causing agent. This is because a one-to-one causal relationship between a disease and a pathogen cannot be established without isolating the pathogen. Efforts to culture and isolate microorganisms, including pathogens, were actively carried out by researchers in the latter half of the 19th century. One of the backgrounds behind their research on microbial culture during this period was that fermentation or putrefaction were thought to be related to human diseases, which is known as zymotic theory. Examples of researchers who were interested in fermentation include the French chemist and microbiologist Louis Pasteur (1822–95). He discovered that the growth of brewer’s yeast is associated with the conversion of sugar into alcohol and carbonic acid (Pasteur, 1857a). He also discovered that sugar is converted to alcohol in the predominance of brewer’s yeast, but it is converted to lactic acid in the predominance of a certain microorganism (which he called lactic yeast) (Pasteur, 1857b). All of this demonstrates how important it is to grow microorganisms in a pure form to achieve the desired fermentation. However, although Pasteur was able to obtain a culture pure enough to obtain the desired fermentation, it is believed that he was unable to obtain microorganisms in a truly pure form (Hitchens and Leikind, 1939). The pure culture of clonal microorganisms is believed to have been first accomplished by the British surgeon Joseph Lister (1827–1912) in 1878 using a method called limiting dilution in liquid culture (Santer, 2010; Weiss, 2005).

The problem with liquid media used by Pasteur and Lister is that it is difficult, if not impossible, to obtain pure cultures, as they are prone to background and carry-over contaminations. This led to the invention and introduction of solid media by other researchers. Solid media have the advantage of facilitating pure culture because colonies consisting of conspecific microorganisms are formed on the medium and a conspecific microorganism can be precisely isolated from each colony using an inoculation needle or the like. Several researchers have contributed to the development of solid media. One of them was the German mycologist Joseph Schröter (1837–94). Schröter reported in 1872 that he observed colonies of pigmented bacteria on the surface of sliced potatoes (Schröter, 1872), which is considered to be the first solid medium used. Subsequently, the German physician and microbiologist Robert Koch (1843–1910) and his colleagues contributed to the development of more convenient solid media. The problem with using potatoes as a culture medium was that bacteria, such as micrococci and bacilli, proliferate in large numbers, whereas bacteria that have been shown to be pathogenic in animal experiments (except for some pathogenic bacteria, such as Bacillus anthracis) do not grow. To overcome this, in 1881 Koch (1881) devised nutrient gelatin media for culturing pathogenic bacteria. The nutrient gelatin media are a medium to which gelatin is added to solidify known nutrient solutions, such as meat extracts, that have been confirmed to be able to culture the target pathogen. In his study on tuberculosis, published a year later, Koch (1882) used a solid medium added with agar instead of gelatin. It is thought that agar was used, as gelatin has problems, such as being liquefied by certain microorganisms and not solidifying at temperatures above 37°C (Hitchens and Leikind, 1939). The idea of using agar as a solid medium is thought to have originated with Fanny Hesse (1850–1934), the wife of Walter Hesse (1846–1911) who conducted aerobiological research under Robert Koch (Hitchens and Leikind, 1939). In addition, Julius Richard Petri (1852–1921), assistant to Robert Koch, devised in 1887 a shallow dish for solid media, now known as the Petri dish (Petri, 1887). All of these groundbreaking inventions by Koch and others contributed greatly to subsequent progress in microbiology.

The accomplishments of microbial research by Koch and others have also been applied and contributed to the field of aerobiology. For example, Hesse (1884) devised an apparatus for collecting and culturing airborne microorganisms. Fig. 1.7 shows the Hesse’s apparatus used by Frankland (1886). Briefly, the Hesse’s apparatus is a glass tube about 70 cm long and 3.5 cm wide, the inner wall of which is coated with nutritional gelatin. The tube mounted on the tripod at the sampling site is connected to an aspirator consisting of two 10-L Erlenmeyer flasks with a rubber hose, and air is drawn into the glass tube by removing water from the aspirator. The aspiration rate is adjusted around 1 L per 2–3 min, and a total of 20 L of air is drawn (Hesse, 1884). Airborne microorganisms drawn into the glass tube deposit on the solid medium by gravitational settling or other mechanisms. Microorganisms deposited on the gelatin medium in the glass tube form colonies (Fig. 1.8). Colonies formed are then counted. Hesse adjusted the aspiration rate to a maximum of about 1 L per 2–3 min to ensure sufficient residence time of aerosol particles in the glass tube. He pointed out that higher aspiration rates lead to more breakthrough of the particles. Also, in Hesse’s paper published in 1884 (Hesse, 1884), gelatin instead of agar was used. He pointed out the problem that bacteria that liquefy gelatin interfere with colony observation. This also highlights the superiority of agar as a solid substrate. The Hesse’s apparatus was also used by the British microbiologist Grace Frankland (1858–1946) and the chemist Percy F. Frankland (1858–1946), who were wife and husband, to investigate airborne microorganisms. For example, they reported the distribution of airborne microorganisms in different parts of the United Kingdom (Frankland, 1886), and also the seasonal variation of airborne microorganisms with low concentrations observed in winter and high concentrations observed in summer (Frankland et al., 1887b). They also reported the identification of bacterial and yeast species isolated from air by the Hesse’s apparatus and other methods (Frankland et al., 1887a). As another method, Frankland (1886) used the so-called settle plate method (see Chapter 5) in which nutritive gelatin was added to a glass dish like a Petri dish (Fig. 1.9). The Hesse’s apparatus was also used by the British chemist Thomas Carnelley (1854–90). He measured airborne microorganisms in homes and schools and reported that opening windows widely for ventilation (see Chapter 7 for ventilation) was important in eliminating them (Carnelley et al., 1887).

Figure 1.7 Hesse’s glass tube whose inner wall is coated with nutrient gelatin used by Frankland (1886). Source: Reprinted from Frankland, P.F., 1886. The distribution of micro-organisms in air. Proceedings of the Royal Society of London 40, 512, Copyright (1886), with permission from the Royal Society (U.K.).

Figure 1.8 Colonies of airborne microorganisms collected and grown on a nutrient medium in a Hesse’s glass tube. Source: Adapted from Carnelley, T., Haldane, J.S., Anderson, A.M., 1887. The carbonic acid, organic matter, and micro-organisms air, more especially of dwellings and schools. Philosophical Transactions of the Royal Society of London B 178, Fig. 1.2, Copyright (1887), with permission from the Royal Society (U.K.).

Figure 1.9 Glass dish filled with nutrient gelatin to collect airborne falling microorganisms. Source: Reprinted from Frankland, P.F., 1886. The distribution of micro-organisms in air. Proceedings of the Royal Society of London 40, 514, Copyright (1886), with permission from the Royal Society (U.K.).

Since the invention of the Hesse’s glass tube, many new air samplers for culture have been developed. For example, Grace Frankland and Percy F. Frankland devised a glass tube air sampler with plugs of glass wool or sugar-coated glass wool at both ends of the glass tube (Frankland and Frankland, 1887b; Frankland and Frankland, 1887a). In this method, a certain volume of air is passed through the glass tube with an aspirator, and airborne microorganisms are collected in the glass wool by a mechanism similar to filtration. After air sampling, the glass wool plug is immersed in sterile water or melted gelatin peptone and agitated to elute the microorganisms into solution. The solution containing the eluted microorganisms is then added with gelatin, poured into a flask or plate, and cultured. They reported that their flask method and Hesse’s method yield virtually identical results under calm air conditions (Frankland and Frankland, 1887a). Julius Richard Petri also developed a glass tube containing sand filter to trap airborne microorganisms (Petri, 1888) (Fig. 1.10). After air sampling, sand is mixed with gelatin and placed in a Petri dish to examine cultured colonies. Carnelley and Wilson (1888) also reported a method for collecting airborne microorganisms in a flask. In their method, gelatin peptone is added in a flask, sterilized at 100°C, and then solidified. The flask is attached with tubes, and air is sucked by an aspirator. Airborne microorganisms drawn into the flask settle on the jelly and the colonies formed are counted. They considered that the collection of airborne microorganisms in the flask is related not only to gravity but also to the initial velocity. Therefore the Carnelley’s apparatus may be considered the first viable impactor of its kind.

Figure 1.10 Petri’s apparatus consisting of a glass tube containing sand filter. Source: Reprinted from Petri, R.J., 1888. Eine neue Methode Bacterien und Pilzsporen in der Luft nachzuweisen und zu zählen. Zeitschrift für Hygiene 3, 47, Copyright (1888), with permission from Springer Nature.

In the 20th century many air samplers for culture were developed. For example, various types of viable impactors have been developed. Viable impactors are inertial impactors using solid nutrient media as particle collection substrates (see Chapter 5 for impactors). One example is the slit impactor developed by Bourdillon et al. (1941). In the slit impactor, air is accelerated through a narrow slit, and the accelerated airborne microorganisms are inertially collected on a Petri dish containing nutrient agar medium. The Petri dish rotates on a turntable relative to the position of the slit, allowing particles to be evenly collected on the Petri dish. Another example is the Andersen impactors. Instead of a narrow slit, the Andersen impactors use multiple circular nozzles to accelerate the air. There are several models of Andersen impactors, the original being a so-called cascade impactor with six impactors arranged in series, with 400 circular jet nozzles in each impactor stage (Andersen, 1958). Each impactor stage has a different nozzle diameter and is designed to collect particles of different sizes onto each stage, thus allowing the measurement of particle size distribution. In addition to the 6-stage impactor, 1- and 2-stage impactors were also later developed (Jones et al., 1985; Turner and Hill, 1975). These Andersen impactors are viable impactors that inertially collect airborne microorganisms on nutrient agar media. Furthermore, a type of air samplers, now called impingers, that collect airborne microorganisms into filtering liquid was reported by Rettger (1910). Rettger (1910) used sterile saline as the filtration liquid, which was mixed with liquefied gelatin after air sampling and poured into a sterile Petri dish to count cultured colonies. After that, impingers called the Greenberg–Smith impinger (Greenburg and Bloomfield, 1932) and all-glass impingers (U.S. Public Helath Service Engineering Unit, 1944) were developed, and the all-glass impinger was also tested to measure airborne microorganisms (Tyler and Shipe, 1959). Also, although the original purpose was not to collect airborne microorganisms, dust removal technologies, such as cyclones (Morse, 1886) and electrostatic precipitators (Cottrell, 1908), were invented from the late 19th century to the early 20th century. These technologies are also used in bioaerosol research today. For more information on cyclones and electrostatic precipitators, see Chapters 5 and 7, respectively.

1.3.4 Emergence of molecular biology and related technologies and their applications to aerobiology

In addition to microscopy and culture methods, molecular biological methods are applied in the field of aerobiology. Therefore it seems meaningful to look back on those historical backgrounds. Below, we review the emergence of molecular biology and related technologies and their applications to aerobiology.

Table 1.1 presents a chronology of key discoveries and related technical inventions in the field of molecular biology. Today, DNA is well known as the molecule that controls heredity, but it was the Swiss physician Friedrich Miescher (1844–95) in 1869 who first isolated the compound that makes up DNA (Dahm, 2005). Specifically, he isolated a novel compound, different from proteins, from leukocytes and named it nuclein because it exists in the nucleus of cells. After that, the German biochemist Albrecht Kossel (1853–1927) discovered that nuclein consists of four types of bases (five types if RNA is included). He discovered adenine (A) in 1885, guanine (G) in 1891, thymine (T) and cytosine (C) in 1893, and uracil (U) in 1901 (Jones, 1953). About half a century later, the American biochemist Erwin Chargaff (1905–2002) discovered that the ratio of adenine to thymine and the ratio of guanine to cytosine were always the same (base pairing rules or Chargaff’s rules) (Chargaff et al., 1951). Two years later, three consecutive papers were published in the journal Nature, revealing that DNA has a double helix structure (Watson and Crick, 1953; Wilkins et al., 1953; Franklin and Gosling, 1953). Also earlier, the British bacteriologist and epidemiologist Frederick Griffith (1877–1941) showed in 1928 that the traits of dead virulent bacteria can be transferred to live nonvirulent bacteria (he called it the transforming principle) (Griffith, 1928). Later, the Canadian-American physician and researcher Oswald Avery (1877–1955) and his colleagues discovered that DNA, not proteins, was responsible for transformation, proving that DNA is a hereditary molecule (Avery et al., 1944).

Table 1.1

Along with the discoveries in molecular biology, related technologies have also been developed. For example, in 1985 a technique called PCR for exponentially amplifying DNA was developed by the American biochemist Kary Mullis (1944–2019) and his colleagues (Saiki et al., 1985). However, in the original PCR, an enzyme called the Klenow fragment of Escherichia coli DNA polymerase I was used, but because the reaction is carried out at high temperatures, there was the problem of thermal degradation of the DNA polymerase. To solve this problem, PCR using thermostable Taq polymerase was later developed (Saiki et al., 1988). Prior to the invention of PCR, Taq polymerase was purified by Chien et al. (1976) from Thermus aquaticus, a thermotolerant bacterium Brock and Freeze (1969) discovered in thermal springs such as Yellowstone National Park. However, conventional PCR has the limitation of not being able to quantify DNA. To solve this problem, a method to monitor the amplification of DNA by PCR in real time (real-time PCR) was developed (Higuchi et al., 1992), and a technique called quantitative PCR to quantify the initial amount of DNA by observing the progress of PCR amplification in real time was introduced (Higuchi et al., 1993; Heid et al., 1996; Gibson et al., 1996).

In addition to PCR, the technologies of DNA sequencing were developed. For example, in the 1970s the so-called first-generation sequencing (FGS) technologies were developed by Sanger et al. (1977) and Maxam and Gilbert (1977). In the field of aerobiology, especially until about 2010, Sanger sequencing was frequently used in combination with DNA cloning, a technique of making large numbers of copies of a DNA molecule. DNA cloning is based on recombinant DNA technology originally invented by Jackson et al. (1972) and Cohen et al. (1973) in the 1970s. However, FGS, including Sanger sequencing, suffers from low throughput (i.e., the amount of sequences processed per unit time) as it reads only one DNA sequence per sample. In the 1990s new sequencing methods that became the bases for what is now called next-generation sequencing or high-throughput sequencing (HTS) were invented. Examples include pyrosequencing (Nyren et al., 1993) used in 454 sequencers and reverse-terminator sequencing (Canard and Sarfati, 1994) used in Illumina sequencers, which have been commercially available since the mid-2000s. These sequencing technologies are also called massively parallel sequencing because they can read millions to billions of sequences of short DNA molecules (less than 1000 bases) in parallel at once. They are also called second-generation sequencing (SGS). However, a drawback of SGS was the short length of each read. The development of third-generation sequencing (TGS), with read lengths typically exceeding 1000 bases, has overcome this problem. One example of TGS is nanopore sequencing, the underlying technology of which was invented by Kasianowicz et al. (1996) in the 1990s and commercialized in the 2010s. For more information on DNA-based methods including DNA sequencing, see Chapter 6.

In many fields of biology, including aerobiology, instead of sequencing the whole genome of an organism or a collection of the whole genomes of organisms contained in an environmental sample (called metagenome), a method called DNA barcoding to sequence a short DNA region, such as genetic marker, and identify species is often used. DNA barcoding has the advantages, such as sensitive detection of organisms because the targeted barcode region can be amplified by PCR and minimal resources required for sequencing. As DNA barcode markers, bacterial 16S ribosomal RNA (rRNA) gene and fungal internal transcribed spacer (ITS) region are often used. The method of identifying the species of organisms based on the sequences of DNA barcode markers, such as 16S rRNA gene, originated from the idea of Woese and Fox (1977), who attempted to classify organisms based on rRNA gene sequences in 1977. Until then, organisms were classified into two domains, prokaryotes and eukaryotes, but they demonstrated that by analyzing rRNA genes, prokaryotes were further divided into bacteria and archaea, and organisms are therefore composed of three domains, which is now widely accepted by researchers. For bacteria, among several rRNA genes, 16S rRNA gene is used as a DNA barcode marker. For fungi, the DNA barcode markers, such as the ITS region, 18S rRNA gene, and 28S rRNA gene, are used. Among them, the ITS region was formally proposed by the Consortium for the Barcode of Life as a universal DNA barcode marker for fungi (Schoch et al., 2012). For accurately making taxonomic assignments based on DNA barcode markers, accurate reference databases are essential. For such purposes, for example, the National Center for Biotechnology Information (NCBI) provides accurate reference databases called Reference Sequence (RefSeq), and the data for loci, such as bacterial and archaeal 16S rRNA gene, fungal ITS region, fungal 18S rRNA gene, and fungal 28S rRNA gene, are covered by the RefSeq Targeted Loci Projects of NCBI.

From around 2000 research on bioaerosols using Sanger sequencing in combination with DNA cloning began to appear in the literature. Examples of early studies reported and the types of airborne organisms and DNA barcode markers analyzed in those studies include 16S rRNA gene of bacteria (Radosevich et al., 2002; Angenent et al., 2005; Maron et al., 2005; Després et al., 2007; Polymenakou et al., 2008), 18S rRNA gene of fungi (Boreson et al., 2004), ITS region of fungi (Després et al., 2007; Fröhlich-Nowoisky et al., 2009), ribulose-bisphosphate carboxylase gene (chloroplast) of plants (Després et al., 2007), and small subunit rRNA gene of organisms of all three domains of life (Fierer et al., 2008). However, since the introduction of HTS into the market around the mid-2000s, aerobiological studies using Sanger sequencing have begun to be replaced by those using HTS. Many of the early HTS-based aerobiology studies analyzed DNA barcode markers, such as the bacterial 16S rRNA gene. As examples of early aerobiology research using HTS, studies analyzing the 16S rRNA gene of bacteria in indoor and outdoor air were reported in the early 2010s (Bowers et al., 2011; Bowers et al., 2012; Hospodsky et al., 2012). Similarly, in the 2010s, aerobiological studies using HTS targeting the ITS regions of fungi (Yamamoto et al., 2012; Adams et al., 2013) and plants (Núñez et al., 2017; Dong et al., 2019) were also reported. These HTS-based studies have revealed the diversity of airborne organisms that could not be elucidated by conventional methods, such as culture, microscopy, and Sanger sequencing. In addition, especially in the latter half of the 2010s, studies using a method called metagenomic sequencing have been reported in the field of aerobiology (Yooseph et al., 2013; Aalismail et al., 2019; Gusareva et al., 2019; Gusareva et al., 2020). Metagenomic sequencing is a method that analyzes all genomes extracted from organisms present in an environmental sample. One of the great advantages of metagenomic sequencing is the ability to analyze a set of multiple different types of antimicrobial resistance genes (ARGs) that do not have common conserved sequences. The set of ARGs contained in a population is called a resistome. In recent years studies on the analysis of air resistomes using metagenomic sequencing have been reported in China (Pal et al., 2016; Hu et al., 2018; Qin et al., 2020) and other countries (Leung et al., 2021). Uncovering atmospheric resistomes using metagenomic sequencing may reveal more about the environmental spread of antimicrobial resistance, which is likely to become one of the major challenges facing humanity in the future. For more information on ARGs in bioaerosols, see Chapter 8.

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