Environmental stress speeds up DNA replication in <i>Pseudomonas putida</i> in chemostat cultivations

Lieder, Sarah; Jahn, Michael; Koepff, Joachim; Müller, Susann

doi:10.1002/biot.201500059

Cited by 16 publications

(14 citation statements)

References 45 publications

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“…Although both bacteria were expected to be in stationary phase at all sampling points (Fig. S9), it is possible that under stress the bacteria adapted their cell cycle behavior and DNA concentration (38). On the other hand, the bacteria might have maintained a similar DNA concentration but a higher RNA concentration, indicating a shift in their gene expression.…”

Section: Discussionmentioning

confidence: 98%

Coculturing Bacteria Leads to Reduced Phenotypic Heterogeneities

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Buysschaert

Props

et al. 2019

Appl Environ Microbiol

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Isogenic bacterial populations are known to exhibit phenotypic heterogeneity at the single-cell level. Because of difficulties in assessing the phenotypic heterogeneity of a single taxon in a mixed community, the importance of this deeper level of organization remains relatively unknown for natural communities. In this study, we have used membrane-based microcosms that allow the probing of the phenotypic heterogeneity of a single taxon while interacting with a synthetic or natural community. Individual taxa were studied under axenic conditions, as members of a coculture with physical separation, and as a mixed culture. Phenotypic heterogeneity was assessed through both flow cytometry and Raman spectroscopy. Using this setup, we investigated the effect of microbial interactions on the individual phenotypic heterogeneities of two interacting drinking water isolates. Through flow cytometry we have demonstrated that interactions between these bacteria lead to a reduction of their individual phenotypic diversities and that this adjustment is conditional on the bacterial taxon. Single-cell Raman spectroscopy confirmed a taxondependent phenotypic shift due to the interaction. In conclusion, our data suggest that bacterial interactions may be a general driver of phenotypic heterogeneity in mixed microbial populations. IMPORTANCE Laboratory studies have shown the impact of phenotypic heterogeneity on the survival and functionality of isogenic populations. Because phenotypic heterogeneity plays an important role in pathogenicity and virulence, antibiotic resistance, biotechnological applications, and ecosystem properties, it is crucial to understand its influencing factors. An unanswered question is whether bacteria in mixed communities influence the phenotypic heterogeneity of their community partners. We found that coculturing bacteria leads to a reduction in their individual phenotypic heterogeneities, which led us to the hypothesis that the individual phenotypic diversity of a taxon is dependent on the community composition.

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Section: Discussionmentioning

confidence: 98%

Coculturing Bacteria Leads to Reduced Phenotypic Heterogeneities

Heyse

Buysschaert

Props

et al. 2019

Appl Environ Microbiol

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“…Although both bacteria were expected to be in stationary phase at all sampling points ( Fig. S1 ), it is possible that under stress, the bacteria adapted their cell cycle behaviour and DNA concentration (47). On the other hand, the bacteria might have maintained a similar DNA concentration but a higher RNA concentration, indicating a shift in their gene expression.…”

Section: Discussionmentioning

confidence: 99%

Coculturing bacteria leads to reduced phenotypic heterogeneities

Heyse

Buysschaert

Props

et al. 2018

Preprint

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19Isogenic bacterial populations are known to exhibit phenotypic heterogeneity at the 20 single cell level. Because of difficulties in assessing the phenotypic heterogeneity of a 21 65 5 difficult to assess the heterogeneity of a single taxon within a mixed community. 66 Recently, several experimental approaches that assess the metabolic diversity of a 67 single taxon in natural communities have been developed (19, 20). However, these 68 approaches rely on FISH-probes that bind to 16S rRNA gene sequences for 69 identification of the taxon of interest. Hence, they do not allow to exclude the possibility 70 that some of the observed phenotypic differences are caused by minor genetic 71 differences between bacteria with very similar 16S rRNA genes. 72Two laser-based methods that are suitable for assessing phenotypes are flow cytometry 73 and Raman spectroscopy (21)(22)(23). Two types of light can be detected by the flow 74 cytometer, that is scattered light and fluorescence. The scattered light provides 75 information about the basic characteristics of the cells (e.g. size, shape and surface 76 properties), while the fluorescence data provides additional information about the cell 77 properties for which it has been stained (e.g. nucleic acid content, metabolic activity, 78 etc.) (24). Flow cytometry thus gives information regarding morphological as well as 79 specific physiological properties of single-cells. The Raman spectrum of a single cell 80 consists of a combination of the individual spectra of all the compounds that make up 81 this cell (e.g. proteins, nucleic acids, fatty acids, etc.). This results in a complex spectrum, 82 which can be interpreted as a chemical fingerprint of the cell (25, 26). Hence, single-cell 83 Raman spectra offer an in depth view on the biochemical composition of each 84 phenotype. 85A tool that can help to answer questions that are difficult to study directly in natural 86 communities is a synthetic ecosystem. A synthetic ecosystem consists of a selected set 87 of species under specific conditions. They are controllable and have a reduced 88 complexity in comparison to natural communities (27). Hence, they provide a way to 89 6 test ecological theories in order to better understand the rules of nature (28). A specific 90 setup for these synthetic ecosystems are co-cultures. The principle of such a system is 91 that two or more bacterial populations are cultivated together with some degree of 92 contact between them, which allows to study their interactions (29). 93An unanswered question, and the focus of this study, is whether bacteria in mixed 94 communities influence the phenotypic heterogeneity of their community partners. Here, 95 we used a synthetic community setup where two isolates were used as model 96 organisms. Four synthetic communities were created. The isolates were grown in 97 axenic cultures as a reference for non-interacting genotypes. To be able to study the 98 individual community members separately after they have been interacting via their 99 joint medi...

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“…Continuous culture systems allow for the long-term, controlled growth of microorganisms or cultured cells. Consequently, they have been used for many scientific and industrial uses, including producing biologics like small molecules [1–3] and recombinant proteins [4,5]; assessing the growth rate [6–9] or metabolism [10,11] of microorganisms or cultured cells under defined conditions; and for studying evolution [12–14]. Continuous culture systems typically operate in one of two modes: a chemostat, where a limited amount of nutrients are constantly added to the culture, or a turbidostat, where the culture density is kept constant [15].…”

Section: Introductionmentioning

confidence: 99%

Yeast grown in continuous culture systems can detect mutagens with improved sensitivity relative to the Ames test

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Molik

et al. 2020

Preprint

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Continuous culture systems allow for the controlled growth of microorganisms over a long period of time. Here, we develop a novel test for mutagenicity that involves growing yeast in continuous culture systems exposed to low levels of mutagen for a period of weeks. In contrast, most microorganism-based tests for mutagenicity expose the potential mutagen to the biological reporter at a high concentration of mutagen for a short period of time. Our test improves upon the sensitivity of the well-established Ames test by at least 20-fold for each of two mutagens that act by different mechanisms (the intercalator ethidium bromide and alkylating agent methylmethane sulfonate). To conduct the tests, cultures were grown in small, inexpensive continuous culture systems in media containing (potential) mutagen, and the resulting mutagenicity of the added compound was assessed via two methods: a canavanine-based plate assay and whole genome sequencing. In the canavanine-based plate assay, we were able to detect a clear relationship between the amount of mutagen and the number of canavanine-resistant mutant colonies over a period of one to three weeks of exposure. Whole genome sequencing of yeast grown in continuous culture systems exposed to MMS demonstrated that quantification of mutations is possible by identifying the number of unique variants across each strain but with lower sensitivity than the plate-based assay. In conclusion, we propose yeast grown in continuous culture systems can provide an improved and more sensitive test for mutagenicity.

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Environmental stress speeds up DNA replication in Pseudomonas putida in chemostat cultivations

Cited by 16 publications

References 45 publications

Coculturing Bacteria Leads to Reduced Phenotypic Heterogeneities

Coculturing Bacteria Leads to Reduced Phenotypic Heterogeneities

Coculturing bacteria leads to reduced phenotypic heterogeneities

Yeast grown in continuous culture systems can detect mutagens with improved sensitivity relative to the Ames test

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