Dataset Information

Stable Isotope Probing Identifies Bacterioplankton Lineages Capable of Utilizing Dissolved Organic Matter Across a Range of Bioavailability.

ABSTRACT: Bacterioplankton consume about half of the dissolved organic matter (DOM) produced by phytoplankton. DOM released from phytoplankton consists of a myriad of compounds that span a range of biological reactivity from labile to recalcitrant. Linking specific bacterioplankton lineages to the incorporation of DOM compounds into biomass is important to understand microbial niche partitioning. We conducted a series of DNA-stable isotope probing (SIP) experiments using 13C-labeled substrates of varying lability including amino acids, cyanobacteria lysate, and DOM from diatom and cyanobacteria isolates concentrated on solid phase extraction PPL columns (SPE-DOM). Amendments of substrates into Sargasso Sea bacterioplankton communities were conducted to explore microbial response and DNA-SIP was used to determine which lineages of Bacteria and Archaea were responsible for uptake and incorporation. Greater increases in bacterioplankton abundance and DOC removal were observed in incubations amended with cyanobacteria-derived lysate and amino acids compared to the SPE-DOM, suggesting that the latter retained proportionally more recalcitrant DOM compounds. DOM across a range of bioavailability was utilized by diverse prokaryotic taxa with copiotrophs becoming the most abundant 13C-incorporating taxa in the amino acid treatment and oligotrophs becoming the most abundant 13C-incorporating taxa in SPE-DOM treatments. The lineages that responded to SPE-DOM amendments were also prevalent in the mesopelagic of the Sargasso Sea, suggesting that PPL extraction of phytoplankton-derived DOM isolates compounds of ecological relevance to oligotrophic heterotrophic bacterioplankton. Our study indicates that DOM quality is an important factor controlling the diversity of the microbial community response, providing insights into the roles of different bacterioplankton in resource exploitation and efficiency of marine carbon cycling.

SUBMITTER: Liu S

PROVIDER: S-EPMC7575717 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Stable Isotope Probing Identifies Bacterioplankton Lineages Capable of Utilizing Dissolved Organic Matter Across a Range of Bioavailability.

Liu Shuting S Baetge Nicholas N Comstock Jacqueline J Opalk Keri K Parsons Rachel R Halewood Elisa E English Chance J CJ Giovannoni Stephen S Bolaños Luis M LM Nelson Craig E CE Vergin Kevin K Carlson Craig A CA

Frontiers in microbiology 20201007

Bacterioplankton consume about half of the dissolved organic matter (DOM) produced by phytoplankton. DOM released from phytoplankton consists of a myriad of compounds that span a range of biological reactivity from labile to recalcitrant. Linking specific bacterioplankton lineages to the incorporation of DOM compounds into biomass is important to understand microbial niche partitioning. We conducted a series of DNA-stable isotope probing (SIP) experiments using <sup>13</sup>C-labeled substrates ...[more]

PMID: 33117322

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Project description:BackgroundMany human microbial infectious diseases including dental caries are polymicrobial in nature. How these complex multi-species communities evolve from a healthy to a diseased state is not well understood. Although many health- or disease-associated oral bacteria have been characterized in vitro, their physiology within the complex oral microbiome is difficult to determine with current approaches. In addition, about half of these species remain uncultivated to date with little known besides their 16S rRNA sequence. Lacking culture-based physiological analyses, the functional roles of uncultivated species will remain enigmatic despite their apparent disease correlation. To start addressing these knowledge gaps, we applied a combination of Magnetic Resonance Spectroscopy (MRS) with RNA and DNA based Stable Isotope Probing (SIP) to oral plaque communities from healthy children for in vitro temporal monitoring of metabolites and identification of metabolically active and inactive bacterial species.Methodology/principal findingsSupragingival plaque samples from caries-free children incubated with (13)C-substrates under imposed healthy (buffered, pH 7) and diseased states (pH 5.5 and pH 4.5) produced lactate as the dominant organic acid from glucose metabolism. Rapid lactate utilization upon glucose depletion was observed under pH 7 conditions. SIP analyses revealed a number of genera containing cultured and uncultivated taxa with metabolic capabilities at pH 5.5. The diversity of active species decreased significantly at pH 4.5 and was dominated by Lactobacillus and Propionibacterium species, both of which have been previously found within carious lesions from children.Conclusions/significanceOur approach allowed for identification of species that metabolize carbohydrates under different pH conditions and supports the importance of Lactobacilli and Propionibacterium in the development of childhood caries. Identification of species within healthy subjects that are active at low pH can lead to a better understanding of oral caries onset and generate appropriate targets for preventative measures in the early stages.

Project description:The response of natural microbial communities to environmental change can be assessed by determining DNA- or RNA-targeted changes in relative abundance of 16S rRNA gene sequences by using fingerprinting techniques such as denaturing gradient gel electrophoresis (DNA-DGGE and RNA-DGGE, respectively) or by stable isotope probing (SIP) of 16S rRNA genes following incubation with a (13)C-labeled substrate (DNA-SIP-DGGE). The sensitivities of these three approaches were compared during batch growth of communities containing two or three Nitrosospira pure or enriched cultures with different tolerances to a high ammonia concentration. Cultures were supplied with low, intermediate, or high initial ammonia concentrations and with (13)C-labeled carbon dioxide. DNA-SIP-DGGE provided the most direct evidence for growth and was the most sensitive, with changes in DGGE profiles evident before changes in DNA- and RNA-DGGE profiles and before detectable increases in nitrite and nitrate production. RNA-DGGE provided intermediate sensitivity. In addition, the three molecular methods were used to follow growth of individual strains within communities. In general, changes in relative activities of individual strains within communities could be predicted from monoculture growth characteristics. Ammonia-tolerant Nitrosospira cluster 3b strains dominated mixed communities at all ammonia concentrations, and ammonia-sensitive strains were outcompeted at an intermediate ammonia concentration. However, coexistence of ammonia-tolerant and ammonia-sensitive strains occurred at the lowest ammonia concentration, and, under some conditions, strains inhibited at high ammonia in monoculture were active at high ammonia in mixed cultures, where they coexisted with ammonia-tolerant strains. The results therefore demonstrate the sensitivity of SIP for detection of activity of organisms with relatively low yield and low activity and its ability to follow changes in the structure of interacting microbial communities.

Project description:To identify and obtain the indigenous degraders metabolizing phenanthrene (PHE) and biphenyl (BP) from the complex microbial community within industrial wastewater, DNA-based stable-isotope probing (DNA-SIP) and cultivation-based methods were applied in the present study. DNA-SIP results showed that two bacterial taxa (Vogesella and Alicyclobacillus) were considered the key biodegraders responsible for PHE biodegradation only, whereas Bacillus and Cupriavidus were involved in BP degradation. Vogesella and Alicyclobacillus have not been linked with PHE degradation previously. Additionally, DNA-SIP helped reveal the taxonomic identity of Ralstonia-like degraders involved in both PHE and BP degradation. To target the separation of functional Ralstonia-like degraders from the wastewater, we modified the traditional cultivation medium and culture conditions. Finally, an indigenous PHE- and BP-degrading strain, Ralstonia pickettii M1, was isolated via a cultivation-dependent method, and its role in PHE and BP degradation was confirmed by enrichment of the 16S rRNA gene and distinctive dioxygenase genes in the DNA-SIP experiment. Our study has successfully established a program for the application of DNA-SIP in the isolation of the active functional degraders from an environment. It also deepens our insight into the diversity of indigenous PHE- and BP-degrading communities.IMPORTANCE The comprehensive treatment of wastewater in industrial parks suffers from the presence of multiple persistent organic pollutants (POPs), such as polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs), which reduce the activity of activated sludge and are difficult to eliminate. Characterizing and applying active bacterial degraders metabolizing multiple POPs therefore helps to reveal the mechanisms of synergistic metabolism and to improve wastewater treatment efficiency in industrial parks. To date, SIP studies have successfully investigated the biodegradation of PAHs or PCBs in real-world habitats. DNA-SIP facilitates the isolation of target microorganisms that pose environmental concerns. Here, an indigenous phenanthrene (PHE)- and biphenyl (BP)-degrading strain in wastewater, Ralstonia pickettii M1, was isolated via a cultivation-dependent method, and its role in PHE and BP degradation was confirmed by DNA-SIP. Our study provides a routine protocol for the application of DNA-SIP in the isolation of the active functional degraders from an environment.

Dataset Information

Stable Isotope Probing Identifies Bacterioplankton Lineages Capable of Utilizing Dissolved Organic Matter Across a Range of Bioavailability.

Publications

Stable Isotope Probing Identifies Bacterioplankton Lineages Capable of Utilizing Dissolved Organic Matter Across a Range of Bioavailability.

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