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Changes in community composition during dilution cultures of marine bacterioplankton as assessed by flow cytometric and molecular biological techniques

Changes in community composition during dilution cultures of marine bacterioplankton as assessed by flow cytometric and molecular biological techniques
Changes in community composition during dilution cultures of marine bacterioplankton as assessed by flow cytometric and molecular biological techniques
Dilution cultures are a common technique for measuring the growth of bacterioplankton communities. In this study, the taxonomic composition of marine bacterioplankton dilution cultures was followed in water samples from Plymouth Sound and the English Channel (UK). Bacterial abundances as well as protein and DNA content were closely monitored by flow cytometry. Denaturing gradient gel electrophoresis (DGGE) of polymerase chain reaction (PCR)-amplified 16S rDNA fragments and fluorescence in situ hybridization (FISH) were applied directly to the water samples and to cells sorted from the dilution cultures based on their protein and DNA content. As expected, a rapid activation of bacteria occurred. However, molecular techniques showed that the community developed in the dilution culture within 1 day was significantly different from that in the original water samples. Whereas in the original samples, cells detectable by FISH were dominated by members of the Cytophaga/Flavobacterium (CF) cluster, in dilution cultures, gamma-proteobacteria accounted for the majority of cells detected, followed by alpha-proteobacteria. An actively growing and an apparently non-growing population with average cellular protein contents of 24 and 4.5 fg respectively, were sorted by flow cytometry. FISH indicated mostly gamma- (64%) and alpha-proteobacteria (33%) in the first active fraction and 78% members of the CF cluster in the second fraction. Sequencing of DGGE bands confirmed the FISH assignments of the latter two groups. The data presented clearly show that even relatively short-term dilution experiments do not measure in situ growth, but rather growth patterns of an enrichment. Furthermore, it was demonstrated that the combination of flow cytometric analysis and sorting combined with FISH and DGGE analysis presented a fairly rapid method of analysing the taxonomic composition of marine bacterioplankton.
1462-2920
191-201
Fuchs, B.M.
1edd27c8-6b61-47f1-9c65-788d3fef9fd3
Zubkov, M.V.
b1dfb3a0-bcff-430c-9031-358a22b50743
Sahm, K.
91a225bf-c3ab-48da-a775-317d660af6ed
Burkill, P.H.
91175019-8b55-4fb5-84ea-334c12de2557
Amann, R.
c56b595a-e2d2-43f4-b058-a34b5fcf05f8
Fuchs, B.M.
1edd27c8-6b61-47f1-9c65-788d3fef9fd3
Zubkov, M.V.
b1dfb3a0-bcff-430c-9031-358a22b50743
Sahm, K.
91a225bf-c3ab-48da-a775-317d660af6ed
Burkill, P.H.
91175019-8b55-4fb5-84ea-334c12de2557
Amann, R.
c56b595a-e2d2-43f4-b058-a34b5fcf05f8

Fuchs, B.M., Zubkov, M.V., Sahm, K., Burkill, P.H. and Amann, R. (2000) Changes in community composition during dilution cultures of marine bacterioplankton as assessed by flow cytometric and molecular biological techniques. Environmental Microbiology, 2 (2), 191-201. (doi:10.1046/j.1462-2920.2000.00092.x).

Record type: Article

Abstract

Dilution cultures are a common technique for measuring the growth of bacterioplankton communities. In this study, the taxonomic composition of marine bacterioplankton dilution cultures was followed in water samples from Plymouth Sound and the English Channel (UK). Bacterial abundances as well as protein and DNA content were closely monitored by flow cytometry. Denaturing gradient gel electrophoresis (DGGE) of polymerase chain reaction (PCR)-amplified 16S rDNA fragments and fluorescence in situ hybridization (FISH) were applied directly to the water samples and to cells sorted from the dilution cultures based on their protein and DNA content. As expected, a rapid activation of bacteria occurred. However, molecular techniques showed that the community developed in the dilution culture within 1 day was significantly different from that in the original water samples. Whereas in the original samples, cells detectable by FISH were dominated by members of the Cytophaga/Flavobacterium (CF) cluster, in dilution cultures, gamma-proteobacteria accounted for the majority of cells detected, followed by alpha-proteobacteria. An actively growing and an apparently non-growing population with average cellular protein contents of 24 and 4.5 fg respectively, were sorted by flow cytometry. FISH indicated mostly gamma- (64%) and alpha-proteobacteria (33%) in the first active fraction and 78% members of the CF cluster in the second fraction. Sequencing of DGGE bands confirmed the FISH assignments of the latter two groups. The data presented clearly show that even relatively short-term dilution experiments do not measure in situ growth, but rather growth patterns of an enrichment. Furthermore, it was demonstrated that the combination of flow cytometric analysis and sorting combined with FISH and DGGE analysis presented a fairly rapid method of analysing the taxonomic composition of marine bacterioplankton.

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Published date: 2000

Identifiers

Local EPrints ID: 58450
URI: http://eprints.soton.ac.uk/id/eprint/58450
ISSN: 1462-2920
PURE UUID: 8b1405a5-f1ee-4ead-9377-ab14ddafa1c4

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Date deposited: 13 Aug 2008
Last modified: 13 Mar 2019 20:32

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Contributors

Author: B.M. Fuchs
Author: M.V. Zubkov
Author: K. Sahm
Author: P.H. Burkill
Author: R. Amann

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