Hidden Ecosystems Above: Unraveling Viral-Bacterial Interactions in Cloudwater

Clouds have been regarded as atmospheric oasis for microbes including psychrophilic bacteria (Delort et al., 2017; Péguilhan et al., 2023). However, adaptations of bacteria to the cloud environment and interactions with viruses are not fully understood. In this study, cloudwater was sampled with six compact Caltech active strand cloud water collectors (Demoz et al., 1996) on the Mount Verde, a mountain of 744 m height on the São Vicente island in the tropical Atlantic Ocean (van Pinxteren et al., 2020) and stored frozen. From iron-flocculated and filtered cloudwater samples, DNA was short-read sequenced for metagenomics, and 24 bacteria were additionally isolated from these samples on Luria-Bertani (LB) and Reasoner's 2A (R2A) agar. After purification, the bacterial DNA was subjected to whole-genome sequencing, revealing a diverse array of microbial taxa. The isolate genomes were identified as belonging to Gram-positive species, including Agrococcus sp., Alkalihalobacillus_A gibsonii_A, Arthrobacter sp., Bacillus spizizenii, Cytobacillus oceanisediminis, Curtobacterium spp., Deinococcus sp., Micrococcus luteus, and Rossellomorea spp., as well as Gram-negative species such as Paracoccus marcusii, and Sphingomonas sp. This microbial diversity highlights the presence of spore-forming, halotolerant, and marine-associated bacteria in cloudwater. The genomes had an average GC content of 58.3% (range 41% – 73%) and encoded for cold-shock genes probably supporting survival during sample freezing and in supercooled cloudwater. The presence of 24 prophages and a diverse arsenal of antiviral defense systems, including adaptive CRISPR immunity targeting viral operational taxonomic units (vOTUs), indicates ongoing bacterial-viral interactions in cloudwater. On average, bacterial strains encoded for five defense systems, with restriction-modification systems being the most common. Interestingly, the isolated strain Sphingomonas sp. MPC37 encoded for the highest number of defense systems (12), indicating its potential ecological significance in this unique environment. Metagenomic sequencing identified 458 vOTUs, with major bacterial hosts predicted as Sphingomonas spp. (75 vOTUs), Deinococcus spp. (15), Novosphingobium spp. (14), and Methylobacterium spp. (13). Analysis of air mass trajectories for the cloudwater suggests a marine origin for certain samples, which were associated with the highest counts of both unique and total vOTUs. We also find genetic variability within a population of closely related viruses (microdiversity). Viral variants arise sequentially during different cloud events and are shared among temporally proximate events. Our results reveal clouds as dynamic microbial and viral ecosystems with complex survival strategies and interactions.

References

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Péguilhan, R., Rossi, F., Joly, M., … & Amato, P. (2023). Clouds, oases for airborne microbes – Differential metagenomics/ metatranscriptomics analyses of cloudy and clear atmospheric situations. bioRxiv, 2023.2012.2014.571671. https://doi.org/10.1101/2023.12.14.571671

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