Publications | Amano Lab | Hokkaido University

Major contribution of anaplerosis to inorganic carbon fixation in the dark ocean

Mon, 15 Jun 2026 00:00:00 +0000

While CO 2 fixation by photo- and chemolithoautotrophs is a central process of the global carbon cycle, many organisms also incorporate inorganic carbon into organic compounds through anaplerotic carbon fixation, a process that replenishes intermediates of central metabolic pathways. However, the active drivers and quantitative importance of anaplerotic carbon fixation in the oceanic carbon cycling remain poorly understood. Here, through analysis of global ocean multi-omics datasets, we identified widespread expression of enzymes involved in this process, especially phosphoenolpyruvate carboxylase. The heterotrophic bacterial genus Alteromonas , a globally distributed marine taxon lacking genes for autotrophic carbon fixation pathways, exhibited particularly high transcriptional and proteomic activity for this enzyme. Laboratory incubations confirmed that Alteromonas assimilated dissolved inorganic carbon (DIC) into biomass, with rates regulated by temperature and organic matter availability. Single-cell tracer analyses of the deep ocean microbial communities quantified Alteromonas ’s contribution at about 17% of total dark DIC fixation (median; confidence interval, 10–28%), equivalent to a potential global flux of about 0.2 PgC yr −1 . These results reveal substantial DIC fixation via anaplerosis, indicating that dark carbon fixation is partly supported by heterotrophic metabolism and modulated by environmental conditions, with responses that may differ from those of canonical autotrophic processes.

Major contribution of particle‐associated microbes to deep‐sea organic carbon degradation

Thu, 01 Jan 2026 00:00:00 +0000

The biological carbon pump mediates the export of particulate organic carbon from the euphotic zone to the deep ocean, where it provides the base of the food web. Although deep‐sea microbial metabolism is considered to be largely associated with macroscopic particles, such as marine snow, the specific contribution of particle‐associated microorganisms to the utilization of bulk organic matter has rarely been directly quantified. We used in situ pumps to collect particles larger than 3 μ m from mesopelagic and bathypelagic waters along a latitudinal transect in the North Atlantic. Prokaryotic abundance, respiration, heterotrophic biomass production, and community composition were determined and compared to the bulk prokaryotic community collected by Niskin bottles. Although particle‐associated prokaryotes represented less than 1% of bulk prokaryotic abundance, they contributed on average 28% to bulk prokaryotic respiration and 12% to biomass production. The organic carbon turnover time of particles mediated by prokaryotes was 0.5–1.5 months, while it was up to 3 yr for the total organic carbon fraction. Thus, particles represent hotspots of organic carbon remineralization in the mesopelagic and bathypelagic ocean. Furthermore, metagenomic analyses revealed clear differences in taxonomy and diversity between the free‐living (0.2–0.8 μ m) and particle‐associated (> 3 μ m) prokaryotic communities. Our results emphasize the significant role of particle‐associated prokaryotes in driving organic matter utilization in the dark ocean.

Single-cell heterotrophic activity in deep-ocean prokaryotic communities quantified by BONCAT and microautoradiography

Thu, 01 Jan 2026 00:00:00 +0000

Prokaryotes play a central role in marine biogeochemical cycles, yet quantifying their activity requires sensitive methods, particularly in the deep ocean where their biomass and metabolic rates are low. One widely used method to determine single-cell activity of prokaryotes is bioorthogonal non-canonical amino acid tagging (BONCAT), which offers a non-radioactive approach to measure protein synthesis. However, direct comparisons between BONCAT and radioisotope-based techniques across ocean depth gradients remain limited, particularly for low-activity prokaryotic communities. To address this knowledge gap, we applied BONCAT to quantify single-cell heterotrophic activity in prokaryotic communities from surface to bathypelagic depths (1000–4000 m) in the Southern Ocean near the Kerguelen Islands. Employing picolyl azide-based copper-catalysed click chemistry, we compared BONCAT (L-homopropargylglycine [HPG] incorporation) with microautoradiography (3H-methionine uptake). BONCAT consistently detected active cells throughout the water column, with HPG-derived total fluorescence intensity closely correlating with both microautoradiography (R2 = 0.91, P < .001) and bulk methionine incorporation (R2 = 0.94, P < .001). This strong relationship between BONCAT and microautoradiography was maintained into the upper bathypelagic depths, where detecting single-cell activity becomes challenging. Our results demonstrate that BONCAT provides estimates of single-cell heterotrophic activity consistent with microautoradiography in deep-ocean samples, supporting its application as a non-radioactive alternative in low-activity environments.

Microbial diagenesis of dissolved organic matter from the ocean’s surface to abyssal depths: a case study in the Humboldt upwelling system

Tue, 16 Dec 2025 00:00:00 +0000

Marine dissolved organic matter (DOM) represents one of Earth’s largest dynamic carbon pools—comparable in scale to atmospheric CO₂. Primarily derived from phytoplankton production in the sunlit surface ocean, DOM serves as a key substrate for heterotrophic microbes that actively transform and recycle it. The portion remaining after microbial diagenesis contributes to the long-lived deep-sea reservoir of refractory dissolved organic carbon (RDOC) with turnover times up to millennia. DOC lability is an important trait determining microbial utilization as well as carbon storage time in the ocean and can be inferred from its chemical composition, particularly changes in individual amino acids (AAs). In this study, we examined dissolved (DOC) and particulate organic carbon (POC) distribution, composition and concentration of dissolved hydrolyzable AAs (DHAA), microbial community structure, and activity along depth profiles from the surface to the abyssopelagic zone (down to 5,000 m) in the Humboldt upwelling system off Chile—one of the ocean’s most productive regions. Our results show a pronounced decrease in DOC concentration and lability, and in viral and prokaryotic abundance with depth. Below the mesopelagic zone, DOC displayed characteristics of RDOC: <42 μmol C L −1 , [DHAA-C]:[DOC] ~ 0.6%, and a glycine fraction of ~75 mol% DHAA. Bacterial biomass production and extracellular enzyme activities (EEA), however, were detectable below the mesopelagic zone and even at abyssal depths, albeit at very low rates. Cell-specific EEA and the proportion of high nucleic acid (HNA) cells increased with depth suggesting adaptation to an extremely low-substrate environment. We discuss microbial carbon turnover under varying assumptions of bacterial growth efficiency and conclude that microbial life in the bathy- and abyssopelagic zones of the Humboldt Current is likely sustained by the flux of sinking particulate organic matter.

Comparison of picolyl azide-based BONCAT and microautoradiography for assessing the heterotrophic prokaryotic activity in the deep ocean

Mon, 20 Oct 2025 00:00:00 +0000

Prokaryotes play a central role in marine biogeochemical cycles, yet quantifying their activity requires sensitive methods due to low biomass and metabolic rates, particularly in the deep ocean. One recent method to determine single-cell activity of prokaryotes is bioorthogonal non-canonical amino acid tagging (BONCAT), which offers a non-radioactive approach to measure protein synthesis. However, direct comparisons between BONCAT and radioisotope-based techniques across ocean depth gradients remain limited, particularly for low-activity prokaryotic communities. To address this knowledge gap, we tested an optimised BONCAT protocol using picolyl azide fluorophores (BONCAT-pic) to assess single-cell heterotrophic activity in prokaryotic communities from surface to bathypelagic depths (1000–4000 m) in the Southern Ocean near the Kerguelen Islands. The method was first optimised using aged coastal and open-ocean seawater, and then compared to microautoradiography with 3 H-methionine uptake. Statistical analysis shows that BONCAT-pic significantly improved detection sensitivity compared to standard azide reagents. BONCAT-pic consistently detected active cells in profiles over the open ocean water column, with cell proportions and fluorescence signals closely correlating with both microautoradiography (R 2 = 0.9, p < 0.001) and bulk methionine incorporation (R 2 = 0.6, p < 0.001). Our results demonstrate that BONCAT-pic is a reliable, fluorescence-based method for quantifying heterotrophic activity at the single-cell level, extending its applicability to prokaryotic communities in the deep ocean.

The contribution of pelagic fungi to ocean biomass

Tue, 01 Jul 2025 00:00:00 +0000

Anaplerotic processes are key contributors to dark carbon fixation in the ocean

Wed, 25 Sep 2024 00:00:00 +0000

Abstract Anaplerotic carbon fixation is ubiquitous in heterotrophic organisms including those inhabiting the ocean1. Despite its prevalence, the drivers of this process and its significance in ocean carbon cycling remain poorly understood2,3. Here we combined global ocean metatranscriptomic analysis, laboratory experiments on a bacterial model strain, and microautoradiography combined with catalyzed reporter deposition fluorescence in situ hybridization (MICRO-CARD-FISH) on marine microbial communities, to uncover the global prevalence of anaplerotic processes in oceanic dark dissolved inorganic carbon (DIC) fixation. Metatranscriptomic analysis revealed high expression levels of key anaplerotic genes, especially in mesopelagic waters, comparable to those of photo- and chemolithoautotrophic DIC fixation genes. Alteromonas emerged as the main contributor to anaplerotic DIC fixation gene expression, highlighting its role in DIC assimilation in the global ocean. Laboratory incubations with a marine Alteromonas representative confirmed their capability to fix DIC, which varied with organic matter availability and temperature. MICRO-CARD-FISH on oceanic samples revealed that Alteromonas contributed 0–40% (14 ± 16%, mean ± s.d.) to the dark DIC fixation in the pelagic ocean. Considering that Alteromonas is an obligate heterotroph lacking chemoautotrophic DIC fixation genes, its contribution to DIC fixation should be attributed to anaplerotic processes. Based on these results, we estimated a contribution of anaplerotic processes to dark DIC fixation of 0–0.5 C Pg y-1 in the global dark ocean. Yet, since Alteromonas is not the only taxon performing anaplerotic DIC fixation, our results represent a baseline conservative estimate. Collectively, our findings place anaplerotic DIC fixation as a relevant processes in the oceanic carbon cycling.

Metaproteomic analysis decodes trophic interactions of microorganisms in the dark ocean

Tue, 30 Jul 2024 00:00:00 +0000

Proteins in the open ocean represent a significant source of organic matter, and their profiles reflect the metabolic activities of marine microorganisms. Here, by analyzing metaproteomic samples collected from the Pacific, Atlantic and Southern Ocean, we reveal size-fractionated patterns of the structure and function of the marine microbiota protein pool in the water column, particularly in the dark ocean (>200 m). Zooplankton proteins contributed three times more than algal proteins to the deep-sea community metaproteome. Gammaproteobacteria exhibited high metabolic activity in the deep-sea, contributing up to 30% of bacterial proteins. Close virus-host interactions of this taxon might explain the dominance of gammaproteobacterial proteins in the dissolved fraction. A high urease expression in nitrifiers suggested links between their dark carbon fixation and zooplankton urea production. In summary, our results uncover the taxonomic contribution of the microbiota to the oceanic protein pool, revealing protein fluxes from particles to the dissolved organic matter pool.

Substrate uptake patterns shape niche separation in marine prokaryotic microbiome

Fri, 17 May 2024 00:00:00 +0000

Marine heterotrophic prokaryotes primarily take up ambient substrates using transporters. The patterns of transporters targeting particular substrates shape the ecological role of heterotrophic prokaryotes in marine organic matter cycles. Here, we report a size-fractionated pattern in the expression of prokaryotic transporters throughout the oceanic water column due to taxonomic variations, revealed by a multi-“omics” approach targeting ATP-binding cassette (ABC) transporters and TonB-dependent transporters (TBDTs). Substrate specificity analyses showed that marine SAR11, Rhodobacterales, and Oceanospirillales use ABC transporters to take up organic nitrogenous compounds in the free-living fraction, while Alteromonadales, Bacteroidetes, and Sphingomonadales use TBDTs for carbon-rich organic matter and metal chelates on particles. The expression of transporter proteins also supports distinct lifestyles of deep-sea prokaryotes. Our results suggest that transporter divergency in organic matter assimilation reflects a pronounced niche separation in the prokaryote-mediated organic matter cycles.

Bacterial degradation of ctenophore Mnemiopsis leidyi organic matter

Tue, 20 Feb 2024 00:00:00 +0000

Jellyfish blooms are increasingly becoming a recurring seasonal event in marine ecosystems, characterized by a rapid build-up of gelatinous biomass that collapses rapidly. Although these blooms have the potential to cause major perturbations, their impact on marine microbial communities is largely unknown. We conducted an incubation experiment simulating a bloom of the ctenophore Mnemiopsis leidyi in the Northern Adriatic, where we investigated the bacterial response to the gelatinous biomass. We found that the bacterial communities actively degraded the gelatinous organic matter, and overall showed a striking similarity to the dynamics previously observed after a simulated bloom of the jellyfish Aurelia aurita s.l . In both cases, we found that a single bacterial species, Pseudoalteromonas phenolica , was responsible for most of the degradation activity. This suggests that blooms of different jellyfish are likely to trigger a consistent response from natural bacterial communities, with specific bacterial species driving the remineralization of gelatinous biomass.

A ubiquitous gammaproteobacterial clade dominates expression of sulfur oxidation genes across the mesopelagic ocean

Mon, 24 Apr 2023 00:00:00 +0000

A device for assessing microbial activity under ambient hydrostatic pressure: The in situ microbial incubator (ISMI)

Wed, 14 Dec 2022 00:00:00 +0000

Microbes in the dark ocean are exposed to hydrostatic pressure increasing with depth. Activity rate measurements and biomass production of dark ocean microbes are, however, almost exclusively performed under atmospheric pressure conditions due to technical constraints of sampling equipment maintaining in situ pressure conditions. To evaluate the microbial activity under in situ hydrostatic pressure, we designed and thoroughly tested an in situ microbial incubator (ISMI). The ISMI allows autonomously collecting and incubating seawater at depth, injection of substrate and fixation of the samples after a preprogramed incubation time. The performance of the ISMI was tested in a high‐pressure tank and in several field campaigns under ambient hydrostatic pressure by measuring prokaryotic bulk 3H‐leucine incorporation rates. Overall, prokaryotic leucine incorporation rates were lower at in situ pressure conditions than under to depressurized conditions reaching only about 50% of the heterotrophic microbial activity measured under depressurized conditions in bathypelagic waters in the North Atlantic Ocean off the northwestern Iberian Peninsula. Our results show that the ISMI is a valuable tool to reliably determine the metabolic activity of deep‐sea microbes at in situ hydrostatic pressure conditions. Hence, we advocate that deep‐sea biogeochemical and microbial rate measurements should be performed under in situ pressure conditions to obtain a more realistic view on deep‐sea biotic processes.

Limited carbon cycling due to high-pressure effects on the deep-sea microbiome

Mon, 28 Nov 2022 00:00:00 +0000

Deep-sea microbial communities are exposed to high-pressure conditions, which has a variable impact on prokaryotes depending on whether they are piezophilic (that is, pressure-loving), piezotolerant or piezosensitive. While it has been suggested that elevated pressures lead to higher community-level metabolic rates, the response of these deep-sea microbial communities to the high-pressure conditions of the deep sea is poorly understood. Based on microbial activity measurements in the major oceanic basins using an in situ microbial incubator, we show that the bulk heterotrophic activity of prokaryotic communities becomes increasingly inhibited at higher hydrostatic pressure. At 4,000 m depth, the bulk heterotrophic prokaryotic activity under in situ hydrostatic pressure was about one-third of that measured in the same community at atmospheric pressure conditions. In the bathypelagic zone—between 1,000 and 4,000 m depth—~85% of the prokaryotic community was piezotolerant and ~5% of the prokaryotic community was piezophilic. Despite piezosensitive-like prokaryotes comprising only ~10% (mainly members of Bacteroidetes, Alteromonas ) of the deep-sea prokaryotic community, the more than 100-fold metabolic activity increase of these piezosensitive prokaryotes upon depressurization leads to high apparent bulk metabolic activity. Overall, the heterotrophic prokaryotic activity in the deep sea is likely to be substantially lower than hitherto assumed, with major impacts on the oceanic carbon cycling.

Impact of hydrostatic pressure on organic carbon cycling of the deep-sea microbiome

Thu, 31 Mar 2022 00:00:00 +0000

Deep-sea microbial communities are exposed to high hydrostatic pressure. While some of these deep-sea prokaryotes are adapted to high-pressure conditions, the contribution of piezophilic (i.e., pressure-loving) and piezotolerant prokaryotes to the total deep-sea prokaryotic community remains unknown. Here we show that the metabolic activity of prokaryotic communities is increasingly inhibited with increasing hydrostatic pressure. At 4,000 m depth, the bulk heterotrophic prokaryotic activity under in sit u hydrostatic pressure was only about one-third of that measured on the same community at atmospheric pressure conditions. Only ∼5% of the bathypelagic prokaryotic community are piezophilic while ∼85% of the deep-sea prokaryotes are piezotolerant. A small fraction (∼10%) of the deep-sea prokaryotes is piezosensitive (mainly members of Bacteroidetes, Alteromonas) exhibiting specific survival strategies at meso- and bathypelagic depths. These piezosensitive bacteria elevated their activity by more than 100-fold upon depressurization. Hence, the consistently higher bulk metabolic activity of the deep-sea prokaryotic community measured upon depressurization is due to a rather small fraction of the prokaryotic community. Overall, the heterotrophic prokaryotic activity in the deep-sea is substantially lower than hitherto assumed with major impacts on the oceanic carbon cycling.

Dynamics of the prokaryotic and eukaryotic microbial community during a cyanobacterial bloom

Mon, 18 Oct 2021 00:00:00 +0000

Toxic cyanobacterial blooms frequently develop in eutrophic freshwater bodies worldwide. Microcystis species produce microcystins (MCs) as a cyanotoxin. Certain bacteria that harbor the mlr gene cluster, especially mlrA, are capable of degrading MCs. However, MC-degrading bacteria may possess or lack mlr genes (mlr+ and mlr− genotypes, respectively). In this study, we investigated the genotype that predominantly contributes to biodegradation and cyanobacterial predator community structure with change in total MC concentration in an aquatic environment. The 2 genotypes coexisted but mlr+ predominated, as indicated by the negative correlation between mlrA gene copy abundance and total MC concentration. At the highest MC concentrations, predation pressure by Phyllopoda, Copepoda, and Monogononta (rotifers) was reduced; thus, MCs may be toxic to cyanobacterial predators. The results suggest that cooperation between MC-degrading bacteria and predators may reduce Microcystis abundance and MC concentration.

Autofluorescence Is a Common Trait in Different Oceanic Fungi

Sun, 29 Aug 2021 00:00:00 +0000

Natural autofluorescence is a widespread phenomenon observed in different types of tissues and organisms. Depending on the origin of the autofluorescence, its intensity can provide insights on the physiological state of an organism. Fungal autofluorescence has been reported in terrestrial and human-derived fungal samples. Yet, despite the recently reported ubiquitous presence and importance of marine fungi in the ocean, the autofluorescence of pelagic fungi has never been examined. Here, we investigated the existence and intensity of autofluorescence in five different pelagic fungal isolates. Preliminary experiments of fungal autofluorescence at different growth stages and nutrient conditions were conducted, reflecting contrasting physiological states of the fungi. In addition, we analysed the effect of natural autofluorescence on co-staining with DAPI. We found that all the marine pelagic fungi that were studied exhibited autofluorescence. The intensity of fungal autofluorescence changed depending on the species and the excitation wavelength used. Furthermore, fungal autofluorescence varied depending on the growth stage and on the concentration of available nutrients. Collectively, our results indicate that marine fungi can be auto-fluorescent, although its intensity depends on the species and growth condition. Hence, oceanic fungal autofluorescence should be considered in future studies when fungal samples are stained with fluorescent probes (i.e., fluorescence in situ hybridization) since this could lead to misinterpretation of results.

Microbial Processing of Jellyfish Detritus in the Ocean

Fri, 30 Oct 2020 00:00:00 +0000

Mesozooplankton taurine production and prokaryotic uptake in the northern Adriatic Sea

Thu, 25 Jun 2020 00:00:00 +0000

Dissolved free taurine, an important osmolyte in phytoplankton and metazoans, has been shown to be a significant carbon and energy source for prokaryotes in the North Atlantic throughout the water column. However, the extent of the coupling between taurine production and consumption over a seasonal cycle has not been examined yet. We determined taurine production by abundant crustacean zooplankton and its role as a carbon and energy source for several prokaryotic taxa in the northern Adriatic Sea over a seasonal cycle. Taurine concentrations were generally in the low nanomolar range, reaching a maximum of 22 nmol L−1 in fall during a Pseudonitzschia bloom and coinciding with the highest zooplankton taurine release rates. Taurine accounted for up to 5% of the carbon, 11% of the nitrogen, and up to 71% of the sulfur requirements of heterotrophic prokaryotes. Members of the Roseobacter clade, Alteromonas, Thaumarchaeota, and Euryarchaeota exhibited higher cell‐specific taurine assimilation rates than SAR11 cells. However, cell‐specific taurine and leucine assimilation were highly variable in all taxa, suggesting species and/or ecotype specific utilization patterns of taurine and dissolved free amino acids. Copepods were able to cover the bulk taurine requirements of the prokaryotic communities in fall and winter and partly in the spring–summer period. Overall, our study emphasizes the significance of taurine as a carbon and energy source for the prokaryotic community in the northern Adriatic Sea and the importance of crustacean zooplankton as a significant source of taurine and other organic compounds for the heterotrophic prokaryotic community.

Response of microcystin biosynthesis and its biosynthesis gene cluster transcription in Microcystis aeruginosa on electrochemical oxidation

Fri, 01 Jun 2018 00:00:00 +0000

Archaeal distribution and abundance in water masses of the Arctic Ocean, Pacific sector

Tue, 30 Apr 2013 00:00:00 +0000

かび臭産生藻類の個体群動態と水環境因子の関係解析

Mon, 15 Oct 2012 00:00:00 +0000

筑波大学における「院生プレゼンバトル」の事例報告 : 学園祭における科学コミュニケーション

Fri, 01 Jun 2012 00:00:00 +0000

深海における海洋細菌の代謝活性と増殖速度を測る : 現場培養機の開発・評価

Sat, 01 Jan 2011 00:00:00 +0000

Musty-odor occurrence and their causative microorganisms in a water supply source

Tue, 01 Jan 2008 00:00:00 +0000