Simultaneous detection of Clostridium perfringens and Clostridium colinum by duplex-polymerase chain reaction

In this study, we provide a protocol for detection of Clostridium colinum and Clostridium perfringens by the single-tube duplex-PCR (dPCR) test for simultaneous and specific detection of both bacteria from pure cultures and fecal samples spiked with these pathogens. Specific primers for each pathoge...

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Veröffentlicht in:Poultry science 2012-12, Vol.91 (12), p.3080-3085
Hauptverfasser: Roussan, D A, Shaheen, Ibrahem A, Totanji, W S, Khawaldeh, G Y, Al Rifai, R H
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container_end_page 3085
container_issue 12
container_start_page 3080
container_title Poultry science
container_volume 91
creator Roussan, D A
Shaheen, Ibrahem A
Totanji, W S
Khawaldeh, G Y
Al Rifai, R H
description In this study, we provide a protocol for detection of Clostridium colinum and Clostridium perfringens by the single-tube duplex-PCR (dPCR) test for simultaneous and specific detection of both bacteria from pure cultures and fecal samples spiked with these pathogens. Specific primers for each pathogen were selected that amplified products of predicted sizes from bacteria in the dPCR as well as in the single-tube PCR (sPCR) assays. The sensitivity and specificity of dPCR assay were compared with those of the sPCR. No product amplification was obtained with DNA from reference strains belonging to the genus Clostridium (except C. colinum and C. perfringens) and isolates belonging to other genera using these primer sets. The dPCR assay was as sensitive as the sPCR assay because bacterial detection limits were similar in both assays. The detection limits of sPCR and dPCR in bacterial suspension were 20 and 25 cfu/mL for C. colinum and C. perfringens, respectively. Meanwhile, in the presence of feces the sensitivity of both assays decreased to a detection limit of 1.25 × 10(4) and 1.94 × 10(4) cfu/g of feces for C. colinum and C. perfringens, respectively. In summary, dPCR assay holds potential to be an economical and rapid diagnostic method for the simultaneous detection of C. colinum and C. perfringens in pure cultures and could be used to screen fecal samples for the presence of these pathogens.
doi_str_mv 10.3382/ps.2012-02368
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source MEDLINE; Elektronische Zeitschriftenbibliothek - Frei zugängliche E-Journals; Alma/SFX Local Collection
subjects Animals
Clostridium - classification
Clostridium - genetics
Clostridium - isolation & purification
DNA, Bacterial - genetics
Polymerase Chain Reaction - methods
Sensitivity and Specificity
Species Specificity
title Simultaneous detection of Clostridium perfringens and Clostridium colinum by duplex-polymerase chain reaction
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