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Assessment of PCR-DGGE for the identification of diverse Helicobacter species, and application to faecal samples from zoo animals to determine Helicobacter prevalence.

Abu Al-Soud, Waleed LU ; Bennedsen, Mads; On, Stephen L. W.; Ouis, Ibn-Sina LU ; Vandamme, Peter; Nilsson, Hans-Olof LU ; Ljungh, Åsa LU and Wadström, Torkel LU (2003) In Journal of Medical Microbiology 52(9). p.765-771
Abstract
Helicobacter species are fastidious bacterial pathogens that are difficult to culture by standard methods. A PCR-denaturing gradient gel electrophoresis (PCR-DGGE) technique for detection and identification of different Helicobacter species was developed and evaluated. The method involves PCR detection of Helicobacter DNA by genus-specific primers that target 16S rDNA and subsequent differentiation of Helicobacter PCR products by use of DGGE. Strains are identified by comparing mobilities of unknown samples to those determined for reference strains; sequence analysis can also be performed on purified amplicons. Sixteen DGGE profiles were derived from 44 type and reference strains of 20 Helicobacter species, indicating the potential of this... (More)
Helicobacter species are fastidious bacterial pathogens that are difficult to culture by standard methods. A PCR-denaturing gradient gel electrophoresis (PCR-DGGE) technique for detection and identification of different Helicobacter species was developed and evaluated. The method involves PCR detection of Helicobacter DNA by genus-specific primers that target 16S rDNA and subsequent differentiation of Helicobacter PCR products by use of DGGE. Strains are identified by comparing mobilities of unknown samples to those determined for reference strains; sequence analysis can also be performed on purified amplicons. Sixteen DGGE profiles were derived from 44 type and reference strains of 20 Helicobacter species, indicating the potential of this approach for resolving infection of a single host by multiple Helicobacter species. Some more highly related species were not differentiated whereas in highly heterogeneous species, sequence divergence was observed and more than one PCR-DGGE profile was obtained. Application of the PCR-DGGE method to DNA extracted from faeces of zoo animals revealed the presence of Helicobacter DNA in 13 of 16 samples; a correlation was seen between the mobility of PCR products in DGGE analysis and DNA sequencing. In combination, this indicated that zoo animals are colonized by a wide range of different Helicobacter species; seven animals appeared to be colonized by multiple Helicobacter species. By this approach, presumptive identifications were made of Helicobacter bilis and Helicobacter hepaticus in a Nile crocodile, Helicobacter cinaedi in a baboon and a red panda, and Helicobacter felis in a wolf and a Taiwan beauty snake. All of these PCR products (400 bp) showed 100 % sequence similarity to 16S rDNA sequences of the mentioned species. These results demonstrate the potential of PCR-DGGE-based analysis for identification of Helicobacter species in complex ecosystems, such as the gastrointestinal tract, and could contribute to a better understanding of the ecology of helicobacters and other pathogens with a complex aetiology. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Journal of Medical Microbiology
volume
52
issue
9
pages
765 - 771
publisher
Lippincott Williams & Wilkins
external identifiers
  • pmid:12909652
  • wos:000185413800006
  • scopus:0141539345
ISSN
0022-2615
DOI
10.1099/jmm.0.05314-0
language
English
LU publication?
yes
id
ef9d31a8-600b-48bf-9fa6-d7b5f552e20a (old id 117433)
date added to LUP
2007-07-09 15:01:49
date last changed
2018-01-07 08:38:39
@article{ef9d31a8-600b-48bf-9fa6-d7b5f552e20a,
  abstract     = {Helicobacter species are fastidious bacterial pathogens that are difficult to culture by standard methods. A PCR-denaturing gradient gel electrophoresis (PCR-DGGE) technique for detection and identification of different Helicobacter species was developed and evaluated. The method involves PCR detection of Helicobacter DNA by genus-specific primers that target 16S rDNA and subsequent differentiation of Helicobacter PCR products by use of DGGE. Strains are identified by comparing mobilities of unknown samples to those determined for reference strains; sequence analysis can also be performed on purified amplicons. Sixteen DGGE profiles were derived from 44 type and reference strains of 20 Helicobacter species, indicating the potential of this approach for resolving infection of a single host by multiple Helicobacter species. Some more highly related species were not differentiated whereas in highly heterogeneous species, sequence divergence was observed and more than one PCR-DGGE profile was obtained. Application of the PCR-DGGE method to DNA extracted from faeces of zoo animals revealed the presence of Helicobacter DNA in 13 of 16 samples; a correlation was seen between the mobility of PCR products in DGGE analysis and DNA sequencing. In combination, this indicated that zoo animals are colonized by a wide range of different Helicobacter species; seven animals appeared to be colonized by multiple Helicobacter species. By this approach, presumptive identifications were made of Helicobacter bilis and Helicobacter hepaticus in a Nile crocodile, Helicobacter cinaedi in a baboon and a red panda, and Helicobacter felis in a wolf and a Taiwan beauty snake. All of these PCR products (400 bp) showed 100 % sequence similarity to 16S rDNA sequences of the mentioned species. These results demonstrate the potential of PCR-DGGE-based analysis for identification of Helicobacter species in complex ecosystems, such as the gastrointestinal tract, and could contribute to a better understanding of the ecology of helicobacters and other pathogens with a complex aetiology.},
  author       = {Abu Al-Soud, Waleed and Bennedsen, Mads and On, Stephen L. W. and Ouis, Ibn-Sina and Vandamme, Peter and Nilsson, Hans-Olof and Ljungh, Åsa and Wadström, Torkel},
  issn         = {0022-2615},
  language     = {eng},
  number       = {9},
  pages        = {765--771},
  publisher    = {Lippincott Williams & Wilkins},
  series       = {Journal of Medical Microbiology},
  title        = {Assessment of PCR-DGGE for the identification of diverse Helicobacter species, and application to faecal samples from zoo animals to determine Helicobacter prevalence.},
  url          = {http://dx.doi.org/10.1099/jmm.0.05314-0},
  volume       = {52},
  year         = {2003},
}