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Isolation and characterization of low nucleic acid (LNA)-content bacteria

Yingying Wang, Frederik Hammes, Nico Boon UGent, Mohamed Chami and Thomas Egli (2009) ISME JOURNAL. 3(8). p.889-902
abstract
Most planktonic bacteria are 'uncultivable' with conventional methods. Flow cytometry (FCM) is one approach that has been taken to study these bacteria. In natural aquatic environments, bacteria with high nucleic acid (HNA) and low nucleic acid (LNA) content are commonly observed with FCM after staining with fluorescent dyes. Although several studies have focused on the relative abundance and in situ activities of these two groups, knowledge on the growth of particularly LNA bacteria is largely limited. In this study, typical LNA bacteria were enriched from three different freshwater sources using extinction dilution (ED) and fluorescence-activated cell sorting (FACS). We have shown for the first time that LNA bacteria can be isolated and cultivated by using sterile freshwater as a growth medium. During growth, the typical LNA characteristics (that is, low-fluorescence intensity and sideward scatter (SSC)) remained distinct from those of typical HNA bacteria. Three LNA pure cultures that are closely affiliated to the Polynucleobacter cluster according to 16S rRNA sequencing results were isolated. Owing to their small size, cells of the isolates remained intact during cryo-transmission electronic microscopy examination and showed a Gram-negative cell-wall structure. The extremely small cell volume (0.05 mu m(3)) observed for all three isolates indicates that they are among the smallest free-living heterotrophic organisms known in culture. Their isolation and cultivation allow further detailed investigation of this group of organisms under defined laboratory conditions. The ISME Journal (2009) 3, 889-902; doi: 10.1038/ismej.2009.46; published online 7 May 2009
Please use this url to cite or link to this publication:
author
organization
year
type
journalArticle (original)
publication status
published
subject
keyword
FLOW-CYTOMETRY, ESCHERICHIA-COLI, FRESH-WATER BACTERIOPLANKTON, DOMINANT BACTERIOPLANKTON GROUPS, IN-SITU HYBRIDIZATION, extinction dilution, low nucleic acid (LNA) bacteria, FACS, DGGE, AOC, cryo-TEM, MARINE-BACTERIA, PLANKTONIC BACTERIA, ELECTRON-MICROSCOPY, DRINKING-WATER, CELL-SIZE
journal title
ISME JOURNAL
ISME J.
volume
3
issue
8
pages
889 - 902
Web of Science type
Article
Web of Science id
000268741300003
JCR category
ECOLOGY
JCR impact factor
6.397 (2009)
JCR rank
5/127 (2009)
JCR quartile
1 (2009)
ISSN
1751-7362
DOI
10.1038/ismej.2009.46
language
English
UGent publication?
yes
classification
A1
copyright statement
I have transferred the copyright for this publication to the publisher
id
751756
handle
http://hdl.handle.net/1854/LU-751756
date created
2009-09-22 13:44:26
date last changed
2009-11-25 09:01:18
@article{751756,
  abstract     = {Most planktonic bacteria are 'uncultivable' with conventional methods. Flow cytometry (FCM) is one approach that has been taken to study these bacteria. In natural aquatic environments, bacteria with high nucleic acid (HNA) and low nucleic acid (LNA) content are commonly observed with FCM after staining with fluorescent dyes. Although several studies have focused on the relative abundance and in situ activities of these two groups, knowledge on the growth of particularly LNA bacteria is largely limited. In this study, typical LNA bacteria were enriched from three different freshwater sources using extinction dilution (ED) and fluorescence-activated cell sorting (FACS). We have shown for the first time that LNA bacteria can be isolated and cultivated by using sterile freshwater as a growth medium. During growth, the typical LNA characteristics (that is, low-fluorescence intensity and sideward scatter (SSC)) remained distinct from those of typical HNA bacteria. Three LNA pure cultures that are closely affiliated to the Polynucleobacter cluster according to 16S rRNA sequencing results were isolated. Owing to their small size, cells of the isolates remained intact during cryo-transmission electronic microscopy examination and showed a Gram-negative cell-wall structure. The extremely small cell volume (0.05 mu m(3)) observed for all three isolates indicates that they are among the smallest free-living heterotrophic organisms known in culture. Their isolation and cultivation allow further detailed investigation of this group of organisms under defined laboratory conditions. The ISME Journal (2009) 3, 889-902; doi: 10.1038/ismej.2009.46; published online 7 May 2009},
  author       = {Wang, Yingying and Hammes, Frederik and Boon, Nico and Chami, Mohamed and Egli, Thomas},
  issn         = {1751-7362},
  journal      = {ISME JOURNAL},
  keyword      = {FLOW-CYTOMETRY,ESCHERICHIA-COLI,FRESH-WATER BACTERIOPLANKTON,DOMINANT BACTERIOPLANKTON GROUPS,IN-SITU HYBRIDIZATION,extinction dilution,low nucleic acid (LNA) bacteria,FACS,DGGE,AOC,cryo-TEM,MARINE-BACTERIA,PLANKTONIC BACTERIA,ELECTRON-MICROSCOPY,DRINKING-WATER,CELL-SIZE},
  language     = {eng},
  number       = {8},
  pages        = {889--902},
  title        = {Isolation and characterization of low nucleic acid (LNA)-content bacteria},
  url          = {http://dx.doi.org/10.1038/ismej.2009.46},
  volume       = {3},
  year         = {2009},
}

Chicago
Wang, Yingying, Frederik Hammes, Nico Boon, Mohamed Chami, and Thomas Egli. 2009. “Isolation and Characterization of Low Nucleic Acid (LNA)-content Bacteria.” Isme Journal 3 (8): 889–902.
APA
Wang, Yingying, Hammes, F., Boon, N., Chami, M., & Egli, T. (2009). Isolation and characterization of low nucleic acid (LNA)-content bacteria. ISME JOURNAL, 3(8), 889–902.
Vancouver
1.
Wang Y, Hammes F, Boon N, Chami M, Egli T. Isolation and characterization of low nucleic acid (LNA)-content bacteria. ISME JOURNAL. 2009;3(8):889–902.
MLA
Wang, Yingying, Frederik Hammes, Nico Boon, et al. “Isolation and Characterization of Low Nucleic Acid (LNA)-content Bacteria.” ISME JOURNAL 3.8 (2009): 889–902. Print.