Neurotoxin gene profiling of Clostridium botulinum types C and D gathered from different countries within Europe

C. Woudstra, A. Skarin, F. Anniballi, F. Fenicia, L. Bano, I. Drigo, M.G.J. Koene, M.H. Bäyon-Auboyer, J.P. Buffereau, D. Medici, P. Fach

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Abstract

Clostridium botulinum types C and D, as well as their mosaic variants C-D and D-C, are associated with avian and mammalian botulism. This study reports on the development of low-density macroarrays based on the GeneDisc cycler platform (Pall-GeneDisc Technologies) applied to the simultaneous detection of the C. botulinum subtypes C, C-D, D, and D-C. The limit of detection of the PCR assays was 38 fg of total DNA, corresponding to 15 genome copies. Artificially contaminated samples of cecum showed a limit of detection below 50 spores/g. The tests were performed with a large variety of bacterial strains, including C. botulinum types C (n = 12), C-D (n = 29), D (n = 5), and D-C (n = 10), other botulinum neurotoxin (BoNT)-producing Clostridium strains (n = 20), non-BoNT-producing clostridia (n = 20), and other bacterial species (n = 23), and showed a high specificity. These PCR assays were compared to previously published real-time PCRs for the detection of C. botulinum in 292 samples collected from cases of botulism events in four European regions. The majority of the samples originated from wild birds (n = 108), poultry (n = 60), and bovines (n = 56). Among the 292 samples, 144 were positive for either the bont/C-D or the bont/D-C gene by using the GeneDisc arrays. The reliability of the results tallied to 97.94%. Interestingly, only BoNT mosaics, types C-D and D-C, were found in naturally contaminated samples whatever their animal origin and their geographical location. Further investigations should now be performed in order to check that mosaic types dominate in Europe and that acquisition of mosaic types helps in survival or adaptation to particular niche
Original languageEnglish
Pages (from-to)3120-3127
JournalApplied and Environmental Microbiology
Volume78
Issue number9
DOIs
Publication statusPublished - 2012

Fingerprint

Clostridium botulinum type D
Clostridium botulinum D
Clostridium botulinum type C
Clostridium botulinum C
Dilatation and Curettage
neurotoxins
Neurotoxins
Botulism
Clostridium
gene
botulinum toxin
botulism
Clostridium botulinum
Genes
Limit of Detection
genes
assay
detection limit
sampling
Polymerase Chain Reaction

Keywords

  • real-time pcr
  • polymerase-chain-reaction
  • wound botulism
  • quantitative detection
  • bovine samples
  • cattle herds
  • alpha-toxin
  • outbreak
  • identification
  • diagnostics

Cite this

Woudstra, C. ; Skarin, A. ; Anniballi, F. ; Fenicia, F. ; Bano, L. ; Drigo, I. ; Koene, M.G.J. ; Bäyon-Auboyer, M.H. ; Buffereau, J.P. ; Medici, D. ; Fach, P. / Neurotoxin gene profiling of Clostridium botulinum types C and D gathered from different countries within Europe. In: Applied and Environmental Microbiology. 2012 ; Vol. 78, No. 9. pp. 3120-3127.
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abstract = "Clostridium botulinum types C and D, as well as their mosaic variants C-D and D-C, are associated with avian and mammalian botulism. This study reports on the development of low-density macroarrays based on the GeneDisc cycler platform (Pall-GeneDisc Technologies) applied to the simultaneous detection of the C. botulinum subtypes C, C-D, D, and D-C. The limit of detection of the PCR assays was 38 fg of total DNA, corresponding to 15 genome copies. Artificially contaminated samples of cecum showed a limit of detection below 50 spores/g. The tests were performed with a large variety of bacterial strains, including C. botulinum types C (n = 12), C-D (n = 29), D (n = 5), and D-C (n = 10), other botulinum neurotoxin (BoNT)-producing Clostridium strains (n = 20), non-BoNT-producing clostridia (n = 20), and other bacterial species (n = 23), and showed a high specificity. These PCR assays were compared to previously published real-time PCRs for the detection of C. botulinum in 292 samples collected from cases of botulism events in four European regions. The majority of the samples originated from wild birds (n = 108), poultry (n = 60), and bovines (n = 56). Among the 292 samples, 144 were positive for either the bont/C-D or the bont/D-C gene by using the GeneDisc arrays. The reliability of the results tallied to 97.94{\%}. Interestingly, only BoNT mosaics, types C-D and D-C, were found in naturally contaminated samples whatever their animal origin and their geographical location. Further investigations should now be performed in order to check that mosaic types dominate in Europe and that acquisition of mosaic types helps in survival or adaptation to particular niche",
keywords = "real-time pcr, polymerase-chain-reaction, wound botulism, quantitative detection, bovine samples, cattle herds, alpha-toxin, outbreak, identification, diagnostics",
author = "C. Woudstra and A. Skarin and F. Anniballi and F. Fenicia and L. Bano and I. Drigo and M.G.J. Koene and M.H. B{\"a}yon-Auboyer and J.P. Buffereau and D. Medici and P. Fach",
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language = "English",
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Woudstra, C, Skarin, A, Anniballi, F, Fenicia, F, Bano, L, Drigo, I, Koene, MGJ, Bäyon-Auboyer, MH, Buffereau, JP, Medici, D & Fach, P 2012, 'Neurotoxin gene profiling of Clostridium botulinum types C and D gathered from different countries within Europe', Applied and Environmental Microbiology, vol. 78, no. 9, pp. 3120-3127. https://doi.org/10.1128/AEM.07568-11

Neurotoxin gene profiling of Clostridium botulinum types C and D gathered from different countries within Europe. / Woudstra, C.; Skarin, A.; Anniballi, F.; Fenicia, F.; Bano, L.; Drigo, I.; Koene, M.G.J.; Bäyon-Auboyer, M.H.; Buffereau, J.P.; Medici, D.; Fach, P.

In: Applied and Environmental Microbiology, Vol. 78, No. 9, 2012, p. 3120-3127.

Research output: Contribution to journalArticleAcademicpeer-review

TY - JOUR

T1 - Neurotoxin gene profiling of Clostridium botulinum types C and D gathered from different countries within Europe

AU - Woudstra, C.

AU - Skarin, A.

AU - Anniballi, F.

AU - Fenicia, F.

AU - Bano, L.

AU - Drigo, I.

AU - Koene, M.G.J.

AU - Bäyon-Auboyer, M.H.

AU - Buffereau, J.P.

AU - Medici, D.

AU - Fach, P.

PY - 2012

Y1 - 2012

N2 - Clostridium botulinum types C and D, as well as their mosaic variants C-D and D-C, are associated with avian and mammalian botulism. This study reports on the development of low-density macroarrays based on the GeneDisc cycler platform (Pall-GeneDisc Technologies) applied to the simultaneous detection of the C. botulinum subtypes C, C-D, D, and D-C. The limit of detection of the PCR assays was 38 fg of total DNA, corresponding to 15 genome copies. Artificially contaminated samples of cecum showed a limit of detection below 50 spores/g. The tests were performed with a large variety of bacterial strains, including C. botulinum types C (n = 12), C-D (n = 29), D (n = 5), and D-C (n = 10), other botulinum neurotoxin (BoNT)-producing Clostridium strains (n = 20), non-BoNT-producing clostridia (n = 20), and other bacterial species (n = 23), and showed a high specificity. These PCR assays were compared to previously published real-time PCRs for the detection of C. botulinum in 292 samples collected from cases of botulism events in four European regions. The majority of the samples originated from wild birds (n = 108), poultry (n = 60), and bovines (n = 56). Among the 292 samples, 144 were positive for either the bont/C-D or the bont/D-C gene by using the GeneDisc arrays. The reliability of the results tallied to 97.94%. Interestingly, only BoNT mosaics, types C-D and D-C, were found in naturally contaminated samples whatever their animal origin and their geographical location. Further investigations should now be performed in order to check that mosaic types dominate in Europe and that acquisition of mosaic types helps in survival or adaptation to particular niche

AB - Clostridium botulinum types C and D, as well as their mosaic variants C-D and D-C, are associated with avian and mammalian botulism. This study reports on the development of low-density macroarrays based on the GeneDisc cycler platform (Pall-GeneDisc Technologies) applied to the simultaneous detection of the C. botulinum subtypes C, C-D, D, and D-C. The limit of detection of the PCR assays was 38 fg of total DNA, corresponding to 15 genome copies. Artificially contaminated samples of cecum showed a limit of detection below 50 spores/g. The tests were performed with a large variety of bacterial strains, including C. botulinum types C (n = 12), C-D (n = 29), D (n = 5), and D-C (n = 10), other botulinum neurotoxin (BoNT)-producing Clostridium strains (n = 20), non-BoNT-producing clostridia (n = 20), and other bacterial species (n = 23), and showed a high specificity. These PCR assays were compared to previously published real-time PCRs for the detection of C. botulinum in 292 samples collected from cases of botulism events in four European regions. The majority of the samples originated from wild birds (n = 108), poultry (n = 60), and bovines (n = 56). Among the 292 samples, 144 were positive for either the bont/C-D or the bont/D-C gene by using the GeneDisc arrays. The reliability of the results tallied to 97.94%. Interestingly, only BoNT mosaics, types C-D and D-C, were found in naturally contaminated samples whatever their animal origin and their geographical location. Further investigations should now be performed in order to check that mosaic types dominate in Europe and that acquisition of mosaic types helps in survival or adaptation to particular niche

KW - real-time pcr

KW - polymerase-chain-reaction

KW - wound botulism

KW - quantitative detection

KW - bovine samples

KW - cattle herds

KW - alpha-toxin

KW - outbreak

KW - identification

KW - diagnostics

U2 - 10.1128/AEM.07568-11

DO - 10.1128/AEM.07568-11

M3 - Article

VL - 78

SP - 3120

EP - 3127

JO - Applied and Environmental Microbiology

JF - Applied and Environmental Microbiology

SN - 0099-2240

IS - 9

ER -