Global Spore Sampling Project: a global, standardized dataset of airborne fungal DNA

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Mots-clé

metabarcoding; DNA; MDT; Occurrence

Contacts

Couverture géographique

Global

Couverture taxonomique

Fungi

Couverture temporelle

Données sur le projet

Pas de description disponible

Les personnes impliquées dans le projet:

Méthodes déchantillonnage

Each sampling site was equipped with a cyclone sampler (Burkard Cyclone Sampler for Field Operation, Burkard Manufacturing Co Ltd; http://burkard.co.uk/product/cyclone-sampler-for-field-operation). The sampling sites represent varying climatic zones and altitudes. Most sampling sites were located in natural environments, with a few in urban settings.

Description des étapes de la méthode:

1. Air DNA samples were acquired using cyclone samplers placed at ground level to ensure free airflow through the sampler. The cyclone sampler orientates itself in the direction of the wind and collects all particles from the air with a single reverse flow cyclone. The sampler collects particles with size >1 μm from the air directly into a sampling tube, including spores, pollen, bacteria, and small insects. The sampler's average air throughput is 16.5 liters per minute for a total of 23,800 liters during each 24-h sampling period. Sterile 1.5 ml Eppendorf vials were used as sampling tubes. After sampling, the vial was removed from the cyclone sampler, the lid was closed, and the vials were labeled with the site code and week number. Likewise, the time and duration of the sampling, as well as notes about the presence of rainwater or larger objects (e.g., arthropods), were recorded. Every week, two 24-h samples (henceforth called Sample A and Sample B) were collected from each site. To avoid contamination, gloves were used while handling the samples or the device. Sampling teams were instructed to clean the cyclone part of the device monthly with water and soap and to rinse it with ethanol, or to sterilize it with dry-heat, chlorine or UV when such equipment was available.
2. The samples were stored at −20°C until they were shipped to the University of Helsinki, Finland. Shipping was at room temperature as the shipping time was relatively short and refrigerated transport would be costly. In Helsinki, visible arthropods were removed from the samples. To avoid losing fungal spores attached to the arthropod bodies, their surface was rinsed by adding sterile water into the sample tube and vortexing. After washing, the arthropods were removed with sterile tweezers. Samples with rainwater were dried in a freeze drier (24 h, −80°C, 0.57 mbar) covered with a porous Parafilm to avoid cross-contamination between samples. After drying, all samples were sent to the University of Guelph, Canada, for DNA extraction and sequencing.
3. Upon receipt, each sample tube was accessioned and assigned a unique Process ID. DNA extraction followed Ivanova et al. (2008) with minor modifications. Two hundred seventy microlitre of ILB (700 mM GuSCN, 30 mM EDTA pH 8.0, 30 mM Tris–HCl pH 8.0, 0.5% Triton® X-100, 5% Tween-20) with 30 μl Proteinase K (20 mg/ml) was added to each collection tube before it was gently rotated to wash spores off the tube walls and lid, and the tube was then centrifuged at 11,000 g for 5 s. The resultant pellet was re-suspended by gentle pipetting, and the entire volume was transferred to a Lysing Matrix A tube (MP-BIO). Tissue was ground in a TissueLyser (Qiagen) for 2 min at 28 Hz. Samples were then incubated for 1 h at 56°C, followed by 1 h at 65°C. Lysates were transferred to a MN block containing 600 μl of 5M GuSCN Binding Buffer (5 M GuSCN, 16.66 mM EDTA pH 8.0, 8.33 mM Tris–HCl pH 6.4, 3.33% Triton® X-100), and the entire volume was transferred in two equal aliquots of 350 μl (each followed by centrifugation at 5,000 xg) onto AcroPrep 96 Filter Plates with 3.0 μm glass fiber media/0.2 μm Bio-Inert membrane, followed by two washes with WB buffer (60% ethanol, 50 mM NaCl, 10 mM Tris–HCl pH 7.4, 0.5 mM EDTA pH 8.0). DNA was eluted in 45 μl of 10 mM Tris-HCl pH 8.0.

Citations bibliographiques

1. Ovaskainen, Otso, et al. "Global Spore Sampling Project: A global, standardized dataset of airborne fungal DNA." Scientific data 11.1 (2024): 561. DOI:https://doi.org/10.1038/s41597-024-03410-0
2. Abrego, Nerea, et al. "Give me a sample of air and I will tell which species are found from your region: Molecular identification of fungi from airborne spore samples." Molecular ecology resources 18.3 (2018): 511-524. DOI:https://doi.org/10.1111/1755-0998.12755
3. Ovaskainen, Otso, et al. "Monitoring fungal communities with the global spore sampling project." Frontiers in Ecology and Evolution 7 (2020): 511. DOI:https://doi.org/10.3389/fevo.2019.00511
4. Abrego, Nerea, et al. "Airborne DNA reveals predictable spatial and seasonal dynamics of fungi." Nature 631.8022 (2024): 835-842. DOI:https://doi.org/10.1038/s41586-024-07658-9

Métadonnées additionnelles