3:30 PM - 3:45 PM
[AOS17-07] Diel Patterns of Marine Prokaryotes Investigated by High-Resolution Sampling at the Kuroshio Extension Observatory Station
★Invited Papers
Keywords:Diel patterns, Marine prokaryotes, High-Resolution Sampling, Kuroshio Extension Observatory station, Metagenomics, Metatranscriptomics
Introduction
The energy that drives marine ecosystems is primarily supplied from the surface in the form of organic matter. This organic matter is synthesized by photosynthetic microorganisms in the euphotic zone and subsequently consumed by heterotrophic organisms. In the ocean, photosynthetic autotrophs, such as phytoplankton, are responsible for primary production, providing organic matter to the ecosystem while adapting to diel cycles. Previous studies have demonstrated that many photosynthetic autotrophs exhibit pronounced diel transcriptional oscillations in genes involved in various metabolic processes such as photosynthesis and cell division. In addition, recent studies have shown that even heterotrophic prokaryotes, which do not directly utilize light, exhibit diel oscillations in gene expression. These findings suggest that marine microbial communities in the surface layer may regulate gene expression in synchrony with environmental changes and the metabolic activities of surrounding microorganisms. Advances in in situ robotic environmental sample processors have enabled high-frequency DNA/RNA collection, facilitating research in the diel variations of marine microbial activity. However, most studies have focused on the surface layer, and how diel microbial variations extend into deeper ocean layers remains poorly understood. This study aims to investigate the vertical extent of diel microbial activity in the euphotic zone of the ocean and the associated metabolic processes, based on high-frequency time-series sampling.
Materials & Methods
Seawater samples were collected at the Kuroshio Extension Observatory (KEO) station during the R/V Mirai MR24-01C cruise. Sampling was conducted at 12 depths (10, 30, 50, 70, 90, 110, 130, 150, 170, 200, 250, and 300 m) every three hours over 72 hours from 18:00 on February 21 to 18:00 on February 24, 2024. The seawater samples were filtered using polycarbonate filters with pore sizes of 3.0 µm and 0.2 µm, and the filtered samples were used for DNA/RNA extraction, separated into eukaryotic and prokaryotic fractions. Additional samples were analyzed for nutrient content and microbial abundance. Environmental parameters, including temperature, salinity, and dissolved oxygen, were measured using CTD sensors. To analyze the microbial community structure, 16S rRNA amplicon sequencing was performed on 100 samples from four depths (10, 70, 130, and 200 m) using 0.2 µm filters. Additionally, to investigate genomic characteristics and gene composition across depths, metagenomic sequencing was conducted on 24 samples from 12 depths at two time points (09:00 for daytime and 21:00 for nighttime).
Results & Discussion, Future Perspectives
During the second night of the sampling period, an atmospheric depression with rainfall passed over the site, deepening the mixed layer, as confirmed by CTD data. Despite minor shifts in community structure, no clear diel variations were detected at any of the four analyzed depths (10, 70, 130, and 200 m). Meanwhile, microbial α-diversity increased with depth, and at 200 m, the community structure was significantly distinct from that of the surface layers (10–130 m). Notably, metagenomic analysis of day and night samples revealed a substantial transition in genomic composition around 150–170 m, where the gradients of oxygen and salinity were most pronounced. Future work will focus on improving prokaryotic genome reconstruction from metagenomic data to better estimate gene functions, aiming to further elucidate microbial metabolism at different depths. Additionally, metatranscriptomic analysis will be performed to examine gene expression patterns and determine which metabolic activities of prokaryotes show diel variations and how deeply these variations extend.
The energy that drives marine ecosystems is primarily supplied from the surface in the form of organic matter. This organic matter is synthesized by photosynthetic microorganisms in the euphotic zone and subsequently consumed by heterotrophic organisms. In the ocean, photosynthetic autotrophs, such as phytoplankton, are responsible for primary production, providing organic matter to the ecosystem while adapting to diel cycles. Previous studies have demonstrated that many photosynthetic autotrophs exhibit pronounced diel transcriptional oscillations in genes involved in various metabolic processes such as photosynthesis and cell division. In addition, recent studies have shown that even heterotrophic prokaryotes, which do not directly utilize light, exhibit diel oscillations in gene expression. These findings suggest that marine microbial communities in the surface layer may regulate gene expression in synchrony with environmental changes and the metabolic activities of surrounding microorganisms. Advances in in situ robotic environmental sample processors have enabled high-frequency DNA/RNA collection, facilitating research in the diel variations of marine microbial activity. However, most studies have focused on the surface layer, and how diel microbial variations extend into deeper ocean layers remains poorly understood. This study aims to investigate the vertical extent of diel microbial activity in the euphotic zone of the ocean and the associated metabolic processes, based on high-frequency time-series sampling.
Materials & Methods
Seawater samples were collected at the Kuroshio Extension Observatory (KEO) station during the R/V Mirai MR24-01C cruise. Sampling was conducted at 12 depths (10, 30, 50, 70, 90, 110, 130, 150, 170, 200, 250, and 300 m) every three hours over 72 hours from 18:00 on February 21 to 18:00 on February 24, 2024. The seawater samples were filtered using polycarbonate filters with pore sizes of 3.0 µm and 0.2 µm, and the filtered samples were used for DNA/RNA extraction, separated into eukaryotic and prokaryotic fractions. Additional samples were analyzed for nutrient content and microbial abundance. Environmental parameters, including temperature, salinity, and dissolved oxygen, were measured using CTD sensors. To analyze the microbial community structure, 16S rRNA amplicon sequencing was performed on 100 samples from four depths (10, 70, 130, and 200 m) using 0.2 µm filters. Additionally, to investigate genomic characteristics and gene composition across depths, metagenomic sequencing was conducted on 24 samples from 12 depths at two time points (09:00 for daytime and 21:00 for nighttime).
Results & Discussion, Future Perspectives
During the second night of the sampling period, an atmospheric depression with rainfall passed over the site, deepening the mixed layer, as confirmed by CTD data. Despite minor shifts in community structure, no clear diel variations were detected at any of the four analyzed depths (10, 70, 130, and 200 m). Meanwhile, microbial α-diversity increased with depth, and at 200 m, the community structure was significantly distinct from that of the surface layers (10–130 m). Notably, metagenomic analysis of day and night samples revealed a substantial transition in genomic composition around 150–170 m, where the gradients of oxygen and salinity were most pronounced. Future work will focus on improving prokaryotic genome reconstruction from metagenomic data to better estimate gene functions, aiming to further elucidate microbial metabolism at different depths. Additionally, metatranscriptomic analysis will be performed to examine gene expression patterns and determine which metabolic activities of prokaryotes show diel variations and how deeply these variations extend.