Monitoring of airborne biological particles in outdoor atmosphere. Part 2: Metagenomics applied to urban environments

The air we breathe contains microscopic biological particles such as viruses, bacteria, fungi and pollen, some of them with relevant clinic importance. These organisms and/or their propagules have been traditionally studied by different disciplines and diverse methodologies like culture and microscopy. These techniques require time, expertise and also have some important biases. As a consequence, our knowledge on the total diversity and the relationships between the different biological entities present in the air is far from being complete. Currently, metagenomics and next-generation sequencing (NGS) may resolve this shortage of information and have been recently applied to metropolitan areas. Although the procedures and methods are not totally standardized yet, the first studies from urban air samples confirm the previous results obtained by culture and microscopy regarding abundance and variation of these biological particles. However, DNA-sequence analyses call into question some preceding ideas and also provide new interesting insights into diversity and their spatial distribution inside the cities. Here, we review the procedures, results and perspectives of the recent works that apply NGS to study the main biological particles present in the air of urban environments. [Int Microbiol 19(2): 69-80 (2016)]Keywords: airborne biological particles · metagenomics · next-generation sequencing (NGS) · air biomonitoring · urban aerobiolog

Análisis por técnicas morfológicas y secuenciación de ADN del polen atmosférico de la Comunidad de Madrid: estudios preliminares. Morphological analysis and DNA sequencing of atmospheric pollen in Madrid region: preliminary study

Hasta el momento, el estudio de las partículas biológicas en el aire que respiramos, se ha dirigido, principalmente, al conocimiento y control del polen y esporas, aeroalérgenos cuyo impacto en salud es bien conocido. Recientemente la comunidad científica ha sugerido que el aire es un ecosistema en sí mismo, que tendría su propia “aerobiota”, compuesta principalmente por virus, bacterias, esporas de hongos y polen. Para estudiar en conjunto toda esta biodiversidad en el aire urbano en la Comunidad de Madrid, surge el consorcio pluridisciplinar AIRBIOTA-CM, que integra a cinco grupos de investigación de áreas muy diferentes, que pretenden obtener una visión conjunta sobre la composición y dinámica de las partículas biológicas del aire, optimizando los sistemas de muestreo y análisis. Las propuestas más novedosas de las investigaciones iniciadas por este consorcio, son la utilización de técnicas innovadoras de génetica molecular como la secuenciación masiva aplicada en metagenómica (“Next Generation Sequencing”, NGS) y el uso de nuevas estrategias de captación, como el empleo de aeronaves no tripuladas, para muestrear a diferentes alturas y en localizaciones geográficas urbanas que a priori puedan tener una composición diferente de la biota y tengan una actividad humana relevante. El proyecto se inicia en otoño de 2014, y los resultados preliminares que presentamos son los obtenidos mediante el análisis morfológico tradicional y el análisis del ADN del polen de una misma muestra procedente de un captador Burkard. Estos resultados evidencian que los captadores tipo Hirst utilizados por las redes aerobiológicas pueden emplearse también en los estudios de metagenómica, y que los datos obtenidos mediante la aplicación de ambos métodos de análisis coinciden a grandes rasgos, lo que revela que esta nueva metodología constituye una buena aproximación y posible alternativa al análisis morfológico, aunque se necesitan más estudios comparativos para adaptar bien esta tecnología. So far, the study of the biological particles in the air we breathe has been mainly directed at knowing and controlling pollen and spores, aeroallergens with a well-known health impact., It has been recently suggested that the air is an ecosystem in itself, and that it probably has its own biota, which would be composed mainly of viruses, bacteria, fungal spores, and pollen. The main objective of the AIRBIOTA-CM project is to study this diverse set of biological particles present in the urban air in the Community of Madrid using a multidisciplinary, innovative and integrative approach. The project is collaboration between five research groups in very different fields, which aim is to get an overview on the composition and dynamics of biological particles in the air to optimize the methods of sampling and analysis. As a methodological innovation, there is an attempt to apply the breakthroughs in metagenomics to the study of bioaerosols. In addition, new collection strategies have been used, such as the use of unmanned aerial vehicles by designing or adapting new samplers for these vehicles, to sample at different altitudes and in urban geographic locations that might presumably have a different composition of the biota and relevant human activity. The project started in autumn 2014. The preliminary results presented here refer to the comparison of results obtained by means of traditional (light microscopy) and metagenomics methods on atmospheric pollen in the Community of Madrid. The data obtained by both analyses coincide broadly, revealing that the molecular methodology is a good and possible alternative approach to morphological analysis, although more comparative studies to adapt well this technology are needed

Monitoring of the airborne biological particles in outdoor atmosphere. Part 1: Importance, variability and ratios

The first part of this review (“Monitoring of airborne biological particles in outdoor atmosphere. Part 1: Importance, variability and ratios”) describes the current knowledge on the major biological particles present in the air regarding their global distribution, concentrations, ratios and influence of meteorological factors in an attempt to provide a framework for monitoring their biodiversity and variability in such a singular environment as the atmosphere. Viruses, bacteria, fungi, pollen and fragments thereof are the most abundant microscopic biological particles in the air outdoors. Some of them can cause allergy and severe diseases in humans, other animals and plants, with the subsequent economic impact. Despite the harsh conditions, they can be found from land and sea surfaces to beyond the troposphere and have been proposed to play a role also in weather conditions and climate change by acting as nucleation particles and inducing water vapour condensation. In regards to their global distribution, marine environments act mostly as a source for bacteria while continents additionally provide fungal and pollen elements. Within terrestrial environments, their abundances and diversity seem to be influenced by the land-use type (rural, urban, coastal) and their particularities. Temporal variability has been observed for all these organisms, mostly triggered by global changes in temperature, relative humidity, et cetera. Local fluctuations in meteorological factors may also result in pronounced changes in the airbiota. Although biological particles can be transported several hundreds of meters from the original source, and even intercontinentally, the time and final distance travelled are strongly influenced by factors such as wind speed and direction. [Int Microbiol 2016; 19(1):1-1 3]Keywords: airborne biological particles · airbiota · bioaerosols · meteorological factors · air-genome ratio

Monitoring of airborne biological particles in outdoor atmosphere. Part 1: Importance, variability and ratios

The first part of this review ("Monitoring of airborne biological particles in outdoor atmosphere. Part 1: Importance, variability and ratios") describes the current knowledge on the major biological particles present in the air regarding their global distribution, concentrations, ratios and influence of meteorological factors in an attempt to provide a framework for monitoring their biodiversity and variability in such a singular environment as the atmosphere. Viruses, bacteria, fungi, pollen and fragments thereof are the most abundant microscopic biological particles in the air outdoors. Some of them can cause allergy and severe diseases in humans, other animals and plants, with the subsequent economic impact. Despite the harsh conditions, they can be found from land and sea surfaces to beyond the troposphere and have been proposed to play a role also in weather conditions and climate change by acting as nucleation particles and inducing water vapour condensation. In regards to their global distribution, marine environments act mostly as a source for bacteria while continents additionally provide fungal and pollen elements. Within terrestrial environments, their abundances and diversity seem to be influenced by the land-use type (rural, urban, coastal) and their particularities. Temporal variability has been observed for all these organisms, mostly triggered by global changes in temperature, relative humidity, et cetera. Local fluctuations in meteorological factors may also result in pronounced changes in the airbiota. Although biological particles can be transported several hundreds of meters from the original source, and even intercontinentally, the time and final distance travelled are strongly influenced by factors such as wind speed and direction

Monitoring of airborne biological particles in outdoor atmosphere. Part 2: Metagenomics applied to urban environments

The air we breathe contains microscopic biological particles such as viruses, bacteria, fungi and pollen, some of them with relevant clinic importance. These organisms and/or their propagules have been traditionally studied by different disciplines and diverse methodologies like culture and microscopy. These techniques require time, expertise and also have some important biases. As a consequence, our knowledge on the total diversity and the relationships between the different biological entities present in the air is far from being complete. Currently, metagenomics and next-generation sequencing (NGS) may resolve this shortage of information and have been recently applied to metropolitan areas. Although the procedures and methods are not totally standardized yet, the first studies from urban air samples confirm the previous results obtained by culture and microscopy regarding abundance and variation of these biological particles. However, DNA-sequence analyses call into question some preceding ideas and also provide new interesting insights into diversity and their spatial distribution inside the cities. Here, we review the procedures, results and perspectives of the recent works that apply NGS to study the main biological particles present in the air of urban environments

Temporal patterns of variability for prokaryotic and eukaryotic diversity in the urban air of Madrid (Spain)

Although many microorganisms are ubiquitously present in the air, airborne microbial communities have been much less characterized than those in soil or aquatic environments. Besides its ecological importance, detection and monitoring of the wide diversity of these aerosolized microorganisms (bacteria, viruses, fungi and pollen) is relevant for understanding allergy and disease outbreaks, especially in highly populated cities. In this study, we describe the simultaneous biodiversity of bacteria, fungi and plants present in the urban atmosphere of Madrid (Spain) along different seasonal periods, using DNA sequencing. Sampling in two different locations (downtown and peri-urban) we found that changes in the composition of each community are mainly driven by environmental factors, rather than by the features of the specific sampling microenvironments. While pollen particles are dominated by a few taxa characteristic of each season, bacteria and fungi show a high diversity but stable core communities along the year. The prokaryotic core is governed by soil and leaf surface bacteria, with predominance of Actinobacteria (Frankiales and Micrococcales) and Alphaproteobacteria (Sphingomonadales, Rhodobacterales, Rhizobiales and Acetobacterales). Fungal diversity is characterized by the steady presence of members of Capnodiales and Pleosporales. Pathogenic bacterial and fungal taxa were also detected across the year. We also correlated the airborne biodiversity with environmental variables. Air temperature has a strong influence on the community composition of bacteria, while pollen and fungi seasonal variations are mainly correlated with precipitation. Our results contribute to the characterization of airborne prokaryotic and eukaryotic communities in urban areas and show the suitability of this method for biosurveillance strategies

Los avances de la Metagenómica aplicados a la Microbiología del aire (Programa AIRBIOTA-CM)

El proyecto AIRBIOTA-CM nace del interés por conocer y modelizar en profundidad la biodiversidad del aire urbano. Los escasos estudios hasta el momento, a menudo parciales, centrados en un solo grupo taxonómico y abordado mediante técnicas tradicionales, sugieren que la diversidad de este ambiente es mayor de lo pensado, considerándolo como un ecosistema propio y con capacidad de interaccionar e influir sobre los demás presentes en la biosfera

Validation of the Hirst-Type Spore Trap for Simultaneous Monitoring of Prokaryotic and Eukaryotic Biodiversities in Urban Air Samples by Next-Generation Sequencing

Pollen, fungi, and bacteria are the main microscopic biological entities present in outdoor air, causing allergy symptoms and disease transmission and having a significant role in atmosphere dynamics. Despite their relevance, a method for monitoring simultaneously these biological particles in metropolitan environments has not yet been developed. Here, we assessed the use of the Hirst-type spore trap to characterize the global airborne biota by high-throughput DNA sequencing, selecting regions of the 16S rRNA gene and internal transcribed spacer for the taxonomic assignment. We showed that aerobiological communities are well represented by this approach. The operational taxonomic units (OTUs) of two traps working synchronically compiled >87% of the total relative abundance for bacterial diversity collected in each sampler, >89% for fungi, and >97% for pollen. We found a good correspondence between traditional characterization by microscopy and genetic identification, obtaining more-accurate taxonomic assignments and detecting a greater diversity using the latter. We also demonstrated that DNA sequencing accurately detects differences in biodiversity between samples. We concluded that high-throughput DNA sequencing applied to aerobiological samples obtained with Hirst spore traps provides reliable results and can be easily implemented for monitoring prokaryotic and eukaryotic entities present in the air of urban areas