Culturing the ubiquitous freshwater actinobacterial acI lineage by supplying a biochemical ‘helper’ catalase

Kim, Suhyun; Kang, Ilnam; Seo, Ji‐Hui; Cho, Jang-Cheon

doi:10.1101/343640

Cited by 16 publications

(21 citation statements)

References 53 publications

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“…Thus far, DTE culturing has led to isolation of many important marine and freshwater groups, such as marine SAR11 Alphaproteobacteria ( 28 , 29 , 34 – 36 ), the freshwater SAR11 LD12 clade ( 29 ), SUP05/Arctic96BD-19 Gammaproteobacteria ( 37 – 39 ), OM43 Betaproteobacteria ( 26 , 27 , 31 , 40 , 41 ), HIMB11-type Roseobacter spp. ( 35 , 42 ), numerous so-called oligotrophic marine Gammaproteobacteria ( 43 ), and acI Actinobacteria ( 44 ).…”

Section: Introductionmentioning

confidence: 99%

Expanding the Diversity of Bacterioplankton Isolates and Modeling Isolation Efficacy with Large-Scale Dilution-to-Extinction Cultivation

Henson

Lanclos

Pitre

et al. 2020

Appl Environ Microbiol

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Cultivated bacterioplankton representatives from diverse lineages and locations are essential for microbiology, but the large majority of taxa either remain uncultivated or lack isolates from diverse geographic locales. We paired large scale dilution-to-extinction (DTE) cultivation with microbial community analysis and modeling to expand the phylogenetic and geographic diversity of cultivated bacterioplankton and to evaluate DTE cultivation success. Here, we report results from 17 DTE experiments totaling 7,820 individual incubations over three years, yielding 328 repeatably transferable isolates. Comparison of isolates to microbial community data of source waters indicated that we successfully isolated 5% of the observed bacterioplankton community throughout the study. 43% and 26% of our isolates matched operational taxonomic units and amplicon single nucleotide variants, respectively, within the top 50 most abundant taxa. Isolates included those from previously uncultivated clades such as SAR11 LD12 and Actinobacteria acIV, as well as geographically novel members from other ecologically important groups like SAR11 subclade IIIa, SAR116, and others; providing isolates in eight putatively new genera and seven putatively new species. Using a newly developed DTE cultivation model, we evaluated taxon viability by comparing relative abundance with cultivation success. The model i) revealed the minimum attempts required for successful isolation of taxa amenable to growth on our media, and ii) identified possible subpopulation viability variation in abundant taxa such as SAR11 that likely impacts cultivation success. By incorporating viability in experimental design, we can now statistically constrain the effort necessary for successful cultivation of specific taxa on a defined medium. Importance Even before the coining of the term “great plate count anomaly” in the 1980s, scientists had noted the discrepancy between the number of microorganisms observed under the microscope and the number of colonies that grew on traditional agar media. New cultivation approaches have reduced this disparity, resulting in the isolation of some of the “most wanted” bacterial lineages. Nevertheless, the vast majority of microorganisms remain uncultured, hampering progress towards answering fundamental biological questions about many important microorganisms. Furthermore, few studies have evaluated the underlying factors influencing cultivation success, limiting our ability to improve cultivation efficacy. Our work details the use of dilution-to-extinction (DTE) cultivation to expand the phylogenetic and geographic diversity of available axenic cultures. We also provide a new model of the DTE approach that uses cultivation results and natural abundance information to predict taxon-specific viability and iteratively constrain DTE experimental design to improve cultivation success.

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Section: Introductionmentioning

confidence: 99%

Expanding the Diversity of Bacterioplankton Isolates and Modeling Isolation Efficacy with Large-Scale Dilution-to-Extinction Cultivation

Henson

Lanclos

Pitre

et al. 2020

Appl Environ Microbiol

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“…We (35) and others (30) have also adapted DTE culturing by incorporating artificial media in place of natural seawater media to successfully isolate abundant bacterioplankton. Thus far, DTE culturing has lead to isolation of many numerically abundant marine and freshwater groups such as marine SAR11 Alphaproteobacteria (28, 29, 34–36), the freshwater SAR11 LD12 clade (29), SUP05/Arctic96BD-19 Gammaproteobacteria (37–39), OM43 Betaproteobacteria (26, 27, 31, 40, 41), HIMB11-Type Roseobacter (35, 42), numerous so-called Oligotrophic Marine Gammaproteobacteria (43), and acI Actinobacteria (44).…”

Section: Introductionmentioning

confidence: 99%

Expanding the diversity of bacterioplankton isolates and modeling isolation efficacy with large scale dilution-to-extinction cultivation

Henson

Lanclos

Pitre

et al. 2020

Preprint

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48Cultivated representatives of bacterioplankton from diverse lineages and locations are 49 essential for experimentally determining the physiology and metabolism that influences their 50 distributions and ecological functions. Despite widespread efforts to bring more microorganisms 51 into culture, the large majority of taxa either remain uncultivated or lack isolates from diverse 52 geographic locales. Furthermore, we have little understanding of the factors that influence 53 relative cultivation success for any particular taxon. We paired large scale dilution-to-extinction 54 (DTE) cultivation with microbial community analysis and DTE modeling to expand the 55 phylogenetic and geographic diversity of cultivated bacterioplankton and to evaluate the success 56 of DTE cultivation for isolating abundant taxa. Here we report the results from 17 DTE 57 experiments totaling 7,820 individual incubations over three years that resulted in 328 repeatably 58 transferable cultures. Comparison of isolates to microbial community taxonomic profiles of 59 source waters indicated that we successfully isolated 5% of the observed bacterioplankton taxa 60 across all 17 sites. 43% and 26% of our isolates matched operational taxonomic units (OTUs) 61 and amplicon single nucleotide variants (ASVs), respectively, within the top 50 most abundant 62 taxa observed during the study. Isolates included those from previously uncultivated clades such 63 as SAR11 LD12 and Actinobacteria acIV, as well as geographically novel members from other 64 ecologically important groups like SAR11 subclade III, HIMB59 Alphaproteobacteria, SAR116, 65OM43, and HIMB11-type Roseobacter spp.. The effort resulted in the first isolates in eight 66 genera and seven species. We also developed a new model of DTE cultivation to evaluate 67 viability of taxa based on the relationship between relative abundance and cultivation success. 68The model i) revealed minimum levels of wells required for successful isolation of taxa 69 amenable to growth on our media, and ii) identified several important and abundant taxa such as 70 SAR11 with low viability that likely impacts cultivation success. By incorporating predicted, 71 measured, or assumed viability in an iterative manner, we can now provide a statistically 72 constrained number of attempts necessary for successful cultivation of a given taxon on a defined 73 medium based on relative abundance. 74 75 76 Importance 77Even before the coining of the term "great plate count anomaly" in the 1980s, scientists 78 had noted the discrepancy between the number of microorganisms observed under the 79 microscope and the number of colonies that grew on traditional agar media. New cultivation 80 approaches have reduced this disparity, resulting in the isolation of some of the "most wanted" 81 bacterial lineages. Nevertheless, the vast majority of microorganisms remain uncultured, 82 hampering progress towards answering fundamental biological questions about many important 83 microorganisms. Furthermore, few studies have evaluate...

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“…Several recent breakthroughs in axenic prokaryotic cultivation have occurred since the classic work targeting Pelagibacter in the SAR11 clade (Rappé et al ), including isolation of the freshwater SAR11 sister clade LD12 ( Candidatus Fonsibacter; Henson et al ), Methylophilales members of the LD28 group ( Candidatus Methylopumilus ; Salcher et al ) and members of the recalcitrant acI lineage of actinobacteria (Kim et al ). Combined cultivation and metagenomic work focused on freshwater polynucleobacter has also provided important insights into evolution and ecological principles of this ubiquitous genus (Hahn et al ; Hoetzinger and Hahn ) Further testament to the importance of pure cultures in aquatic microbiology is the insightful research on nitrification by marine thaumarchaeota (Santoro et al ).…”

Section: Future Outlooksmentioning

confidence: 99%

Linking metagenomics to aquatic microbial ecology and biogeochemical cycles

Grossart

Massana

McMahon

et al. 2019

Limnology & Oceanography

112

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Microbial communities are essential components of aquatic ecosystems through their contribution to food web dynamics and biogeochemical processes. Aquatic microbial diversity is immense and a general challenge is to understand how metabolism and interactions of single organisms shape microbial community dynamics and ecosystemscale biogeochemical transformations. Metagenomic approaches have developed rapidly, and proven to be powerful in linking microbial community dynamics to biogeochemical processes. In this review, we provide an overview of metagenomic approaches, followed by a discussion on some recent insights they have provided, including those in this special issue. These include the discovery of new taxa and metabolisms in aquatic microbiomes, insights into community assembly and functional ecology as well as evolutionary processes shaping microbial genomes and microbiomes, and the influence of human activities on aquatic microbiomes. Given that metagenomics can now be considered a mature technology where data generation and descriptive analyses are relatively routine and informative, we then discuss metagenomic-enabled research avenues to further link microbial dynamics to biogeochemical processes. These include the integration of metagenomics into well-designed ecological experiments, the use of metagenomics to inform and validate metabolic and biogeochemical models, and the pressing need for ecologically relevant model organisms and simple microbial systems to better interpret the taxonomic and functional information integrated in metagenomes. These research avenues will contribute to a more mechanistic and predictive understanding of links between microbial dynamics and biogeochemical cycles. Owing to rapid climate change and human impacts on aquatic ecosystems, the urgency of such an understanding has never been greater.

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Culturing the ubiquitous freshwater actinobacterial acI lineage by supplying a biochemical ‘helper’ catalase

Cited by 16 publications

References 53 publications

Expanding the Diversity of Bacterioplankton Isolates and Modeling Isolation Efficacy with Large-Scale Dilution-to-Extinction Cultivation

Expanding the Diversity of Bacterioplankton Isolates and Modeling Isolation Efficacy with Large-Scale Dilution-to-Extinction Cultivation

Expanding the diversity of bacterioplankton isolates and modeling isolation efficacy with large scale dilution-to-extinction cultivation

Linking metagenomics to aquatic microbial ecology and biogeochemical cycles

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