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Novel metabarcoding method reveals pronounced seasonality and high turnover rate of Imitervirales in coastal communities

Prodinger, Florian 京都大学 DOI:10.14989/doctor.k24083

2022.05.23

概要

Imitervirales, an order of giant viruses of the phylum Nucleocytoviricota, are abundant in the ocean. In recent studies Imitervirales were further shown to be diverse and able to influence biogeochemical cycles. These findings highlight that Imitervirales are an important part of the microbial communities of the ocean and that it is necessary to study their community dynamics to understand the complex interaction of oceanic microorganisms. Yet, studies targeting their community dynamics and interactions with other microbial communities are still rare.

A recent study established a holistic metabarcoding approach named MEGAPRIMER to analyze Imitervirales communities in several oceanic areas in Japan. However, this method employed 82 degenerate primer pairs targeting a conserved gene, and thus necessitated 82 separate polymerase chain reaction (PCR) amplifications per sample. Therefore, its applicability was limited to studies with a small number of samples.

To overcome this limitation, I tried mixing MEGAPRIMER to “primer–pair–cocktails” in the first part of my thesis. I also investigated the possible bias of MEGAPRIMER with real time polymerase chain reaction (i.e., qPCR). I found that “primer–pair–cocktails” were an alternative to using primers separately. This new Imitervirales metabarcoding workflow reduced the amount of necessary sample DNA and preparation time while reproducing previous results.

In the second part of this thesis, I employed metabarcoding of different microbes (i.e., eukaryotes, prokaryotes, and Imitervirales) to study their diversity and community dynamics in a coastal area in Shikoku, Japan, based on a set of 43 samples collected during 20 months. The analysis resulted in three major discoveries. I showed that (1) all three microbial groups displayed similar seasonal cycles in their community dynamics. While the seasonality of eukaryotic and prokaryotic microbes was known, the seasonality of Imitervirales community has not been observed before. (2) The taxonomic units represented by amplicon sequence variants (ASVs) exhibited systematic differences in their persistence and recurrence among microbial communities when compared weeks, months, or one year apart. (3) Abiotic factors (e.g., temperature, nutrients, or salinity) had low explanatory power over the community compositions of eukaryotes and Imitervirales.

From these observations I concluded that how quickly communities changed their members varies for different communities. Imitervirales showed the highest turnover rate of any microbial community. This may be due to the high selective pressure on viruses to adapt to the also rapidly changing host community. The differences in community dynamics observed here were interpreted by formulating the “community memory” hypothesis. This hypothesis states that current communities influence the formation of future communities, however this influence diminishes over time. How long a previous community can impact the composition of a future community is different for different microbes.

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