Improvement and validation of RAPD in combination with PFGE analysis of Salmonella enterica ssp enterica serovar Senftenberg strains isolated from feed mills

Charlotta Löfström, John Eriksson, Anna Aspan, Per Haggblom, Anders Gunnarsson, Elisabeth Borch, Peter Radstrom

Research output: Contribution to journalJournal articleResearchpeer-review

Abstract

In 1995 and 1996 a Swedish feed mill had problems due to a persistent contamination of Salmonella enterica spp. enterica serovar Senftenberg that was difficult to eliminate. Forty-eight strains isolated from the feed mill, together with unrelated strains included to evaluate the discriminatory power and reproducibility, were analysed by pulsed-field gel electrophoresis (PFGE). The source of contamination in the feed mill was identified and preventative measures were taken, that led to a resolution of the problem. A previously developed randomly amplified polymorphic DNA (RAPD) protocol was used, to evaluate a rapid and low-cost alternative to PFGE typing. The use of the alternative thermostable DNA polymerase Tth was shown to increase the reproducibility of the RAPD analysis. The reproducibility, in terms of Pearson's and Dice's similarity coefficients for duplicate runs, increased from 72.0 +/- 16.9% and 72.3 +/- 12.9% for Taq to 91.6 +/- 7.5% and 90.9 +/- 5.3% for the fingerprints obtained for the RAPD method employing Tth DNA polymerase. Simpson's index of diversity was calculated and found to be 0.580 for RAPD and 0.896 for PFGE. All of the seven RAPD, types could be subdivided into one or more PFGE types, whereas none of the 22 PFGE types was divided into more than one RAPD type. RAPD provides a simple, rapid and powerful screening method that can be used to initially select isolates for further analysis by PFGE (c) 2005 Elsevier B.V. All rights reserved.
Original languageEnglish
JournalVeterinary Microbiology
Volume114
Issue number3-4
Pages (from-to)345-351
Number of pages7
ISSN0378-1135
DOIs
Publication statusPublished - 2006
Externally publishedYes

Keywords

  • SALMONELLA
  • Animal Feed
  • Animals
  • DNA, Bacterial
  • Electrophoresis, Gel, Pulsed-Field
  • Food Contamination
  • Food Microbiology
  • Phylogeny
  • Random Amplified Polymorphic DNA Technique
  • Salmonella enterica
  • Sweden
  • DNA polymerase
  • animal food
  • article
  • bacterial strain
  • bacterium isolation
  • controlled study
  • correlation coefficient
  • discriminant validity
  • food contamination
  • genotype
  • nonhuman
  • pulsed field gel electrophoresis
  • random amplified polymorphic DNA
  • reproducibility
  • serotype
  • thermostability
  • validation process
  • Animalia
  • Bacteria (microorganisms)
  • Salmonella
  • Animal feed
  • Genotyping
  • Pulsed-field gel electrophoresis
  • Randomly amplified polymorphic DNA
  • MICROBIOLOGY
  • VETERINARY
  • POLYMORPHIC DNA ANALYSIS
  • CONTAMINATED FEED
  • OUTBREAK
  • ENTERITIDIS
  • SEROTYPES
  • ANIMALS
  • animal feed
  • genotyping
  • pulsed-field gel electrophoresis
  • randomly amplified polymorphic DNA
  • feed mill
  • Facultatively Anaerobic Gram-Negative Rods Eubacteria Bacteria Microorganisms (Bacteria, Eubacteria, Microorganisms) - Enterobacteriaceae [06702] Salmonella enterica enterica subspecies strain, serovar-Seftenberg
  • Salmonella enterica enterica Tth gene [Enterobacteriaceae]
  • DNA
  • DNA polymerase 9012-90-2
  • 03502, Genetics - General
  • 10062, Biochemistry studies - Nucleic acids, purines and pyrimidines
  • 10802, Enzymes - General and comparative studies: coenzymes
  • 31000, Physiology and biochemistry of bacteria
  • 31500, Genetics of bacteria and viruses
  • Biochemistry and Molecular Biophysics
  • pulsed field gel electrophoresis PFGE electrophoretic techniques, laboratory techniques
  • RAPD laboratory techniques, genetic techniques
  • Methods and Techniques
  • Molecular Genetics

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