Biology subjects

Fichot, E. B.

Publications and source records attributed to Fichot, E. B..

3 recordsLinked to original sources

bioRxiv2019

Comprehensive single-PCR 16S and 18S rRNA community analysis validated with mock communities and denoising algorithms

Universal SSU rRNA primers allow comprehensive quantitative profiling of natural communities by simultaneously amplifying templates from Bacteria, Archaea, and Eukaryota in a single PCR reaction. Despite the potential to show all rRNA gene relative gene abundances, they are rarely used due to concerns about length bias against 18S amplicons and bioinformatic challenges converting mixed 16S/18S sequences into amplicon sequence variants. We thus developed 16S and 18S rRNA mock communities and a bioinformatic pipeline to validate this three-domain approach. To test for length biases, we mixed eukaryotic and prokaryotic mocks before PCR, and found consistent two-fold underestimation of longer 18S sequences due to sequencing but not PCR bias. Using these mocks, we show universal V4-V5 primers (515Y/926R) outperformed eukaryote-specific V4 primers in observed vs. expected abundance correlations and sequences with single mismatches to the primer were strongly underestimated (3-8 fold). A year of monthly time-series data from a protist-enriched 1.2-80 m size fraction yielded an average of 9% 18S, 17% chloroplast 16S, and 74% prokaryote 16S rRNA gene amplicons. These data demonstrate the potential for universal primers to generate quantitative and comprehensive microbiome profiles, although gene copy and genome size variability should be considered - as for any quantitative genetic analysis.

microbiology↗

bioRxiv2017

Dynamics of finely resolved, abundant symbiotic marine plankton and other interacting microbes via automated high-frequency sampling

Short time-scale observations are valuable for understanding microbial ecological processes. We assessed dynamics in relative abundance and potential activities by sequencing the small sub-unit ribosomal RNA gene (rRNA gene) and rRNA molecules (rRNA) of Bacteria, Archaea, and Eukaryota once to twice-daily between March 2014 and May 2014 from the surface ocean off Catalina Island, California. Typically Ostreococcus, Braarudosphaera, Teleaulax, and Synechococcus dominated phytoplankton sequences (including chloroplasts) while SAR11, Sulfitobacter, and Fluviicola dominated non-phytoplankton Bacteria and Archaea. We observed short-lived increases of diatoms, mostly Pseudo-nitzschia and Chaetoceros, with quickly responding Bacteria and Archaea including Flavobacteriaceae (Polaribacter & Formosa), Roseovarius, and Euryarchaeota (MGII), notably the exact amplicon sequence variants we observed responding similarly to another diatom bloom nearby, three years prior. We observed correlations representing known interactions among abundant phytoplankton rRNA sequences, demonstrating the biogeochemical and ecological relevance of such interactions: 1) The kleptochloroplastidic ciliate Mesodinium 18S rRNA gene sequences and a single Teleaulax taxon (via 16S rRNA gene sequences) were correlated (Spearman r =0.83) yet uncorrelated to a Teleaulax 18S rRNA gene OTU, or any other taxon (consistent with a kleptochloroplastidic or karyoklepty relationship) and 2) the photosynthetic prymnesiophyte Braarudosphaera bigelowii and two strains of diazotrophic cyanobacterium UCYN-A were correlated and each taxon was also correlated to other taxa, including B. bigelowii to a verrucomicrobium and a dictyochophyte phytoplankter (all r > 0.8). We also report strong correlations (r > 0.7) between various ciliates, bacteria, and phytoplankton, suggesting interactions via currently unknown mechanisms. These data reiterate the utility of high-frequency time-series to show rapid microbial reactions to stimuli, and provide new information about in-situ dynamics of previously recognized and hypothesized interactions.

microbiology↗

bioRxiv2017

Dynamic marine viral infections and major contribution to photosynthetic processes shown by regional and seasonal picoplankton metatranscriptomes

Viruses are an important top-down control on microbial communities, yet their direct study in natural environments has been hindered by culture limitations1-3. The advance of sequencing and bioinformatics over the last decade enabled the cultivation independent study of viruses. Many studies focus on assembling new viral genomes4-6 and studying viral diversity using marker genes amplified from free viruses7,8. We used cellular metatranscriptomics to study community-wide viral infections at three coastal California sites throughout a year. Generation of and recruitment to viral contigs (> 5kbp, N=66) allowed tracking of infection dynamics over time and space. Here we show that while these assemblies represent viral populations, they are likely biased towards clonal or low diversity assemblages. Furthermore, we demonstrate that published T4-like cyanophages (N=50) and pelagiphages (N=4), having genomic continuity between close relatives, are better tracked using marker genes. Additionally, we demonstrate determination of potential hosts by matching infection dynamics with microbial community composition. Finally, we quantify the relative contribution of various cyanobacteria and viruses to photosystem-II psbA expression in our study sites. We show sometimes >50% of all cyanobacterial+viral psbA expression we observed is of viral origin, which highlights the proportion of infected cells and makes viruses a remarkable contributor to photosynthesis and oxygen production.

ecology↗