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REP-PCR analysis of Pasteurella multocida isolates that cause haemorrhagic septicaemia
K M Townsend1, H J Dawkins, J M Papadimitriou
1UWA Department of Pathology, Q.E. II Medical Centre, Nedlands, Australia. kirsty.townsend@mailbox.uq.edu.au
Abstract:
Amplification of multiple P multocida genomic DNA fragments by outwardly-directed primers based on the repetitive extragenic palindromic (REP) consensus sequence, generated complex profiles in a PCR-based fingerprinting method known as REP-PCR. Polymorphisms within REP-PCR profiles were used to characterise 38 isolates of P multocida. The high degree of homogeneity observed among haemorrhagic septicaemia (HS) strains of serotype B and E provided evidence of a disease-associated REP profile that may serve as a novel method for the identification of HS strains regardless of serotype. REP-PCR profiles of other P multocida serotypes were highly variable, illustrating the potential of this technique for the molecular fingerprinting of fowl cholera or atrophic rhinitis isolates. A specific amplified REP fragment was isolated and used to probe membrane-bound digested P multocida genomic DNA. Hybridisation patterns not only distinguished HS-causing isolates from non-HS P multocida, but also demonstrated a degree of relatedness between HS and HS-like strains.
Insights
Repetitive extragenic palindromic (REP)-PCR fingerprinting identified unique DNA profiles for Pasteurella multocida strains. This molecular method shows promise for identifying haemorrhagic septicaemia strains and differentiating other P. multocida isolates.
Area of Science:
- Microbiology
- Molecular Biology
- Genomics
Background:
- Pasteurella multocida is a significant bacterial pathogen causing various diseases in animals.
- Accurate identification and characterization of P. multocida strains are crucial for disease control and management.
- Existing typing methods may lack the discriminatory power or specificity needed for certain applications.
Purpose of the Study:
- To develop and evaluate a PCR-based fingerprinting method, REP-PCR, for molecular characterization of P. multocida isolates.
- To investigate the potential of REP-PCR for identifying specific P. multocida strains, such as those causing haemorrhagic septicaemia (HS).
- To assess the utility of REP-PCR for differentiating between various P. multocida serotypes and disease associations.
Main Methods:
- Genomic DNA fragments of P. multocida were amplified using primers targeting repetitive extragenic palindromic (REP) sequences.
- The resulting DNA profiles (fingerprints) generated by REP-PCR were analyzed for polymorphisms.
- A specific REP-PCR amplified fragment was isolated and used as a probe for DNA hybridization analysis.
Main Results:
- REP-PCR generated complex and polymorphic DNA profiles for P. multocida isolates.
- A high degree of homogeneity was observed in REP-PCR profiles of haemorrhagic septicaemia (HS) strains (serotypes B and E), suggesting a disease-associated profile.
- REP-PCR profiles varied significantly among other P. multocida serotypes, indicating potential for differentiating fowl cholera and atrophic rhinitis isolates.
- Hybridization patterns using an isolated REP fragment distinguished HS-causing strains from non-HS strains and revealed relatedness between HS and HS-like strains.
Conclusions:
- REP-PCR is a valuable molecular fingerprinting technique for P. multocida characterization.
- The identified REP-PCR profile associated with HS strains offers a potential method for their identification irrespective of serotype.
- REP-PCR has the potential for molecular typing of P. multocida isolates involved in fowl cholera and atrophic rhinitis.
- DNA hybridization with REP fragments can further aid in differentiating and classifying P. multocida strains based on disease association.