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Pasteurella multocida detection by 5' Taq nuclease assay: a new tool for use in diagnosing fowl cholera
B G Corney1, I S Diallo, L L Wright
1Animal Research Institute, Department of Primary Industries and Fisheries, Yeerongpilly, Queensland 4105, Australia. bruce.corney@dpi.qld.gov.au
Abstract:
A 5' Taq nuclease assay utilising minor groove binder technology and targeting the 16S rRNA gene was designed to detect Pasteurella multocida (the causative agent of fowl cholera) in swabs collected from poultry. The assay was first evaluated using pure cultures. The assay correctly identified four P. multocida taxonomic type strains, 18 P. multocida serovar reference strains and 40 Australian field isolates (17 from poultry, 11 from pigs and 12 from cattle). Representatives of nine other Pasteurella species, 26 other bacterial species (18 being members of the family Pasteurellaceae) and four poultry virus isolates did not react in the assay. The assay detected a minimum of approximately 10 cfu of P. multocida per reaction. Of 79 poultry swabs submitted to the laboratory for routine bacteriological culture, 17 were positive in the 5' Taq nuclease assay, but only 10 were positive by culture. The other 62 swabs were negative for P. multocida by both 5' Taq nuclease assay and culture. The assay is suitable for use in diagnosing fowl cholera, is more rapid than bacteriological culture, and may also have application in diagnosing P. multocida infections in cattle and pigs.
Insights
A new 5' Taq nuclease assay accurately detects Pasteurella multocida, the cause of fowl cholera, in poultry swabs. This rapid diagnostic tool offers improved sensitivity over traditional culture methods.
Area of Science:
- Veterinary Microbiology
- Molecular Diagnostics
- Avian Pathology
Background:
- Fowl cholera, caused by Pasteurella multocida, significantly impacts poultry health and industry.
- Accurate and rapid detection of P. multocida is crucial for effective disease management.
- Existing diagnostic methods, such as bacteriological culture, can be time-consuming and less sensitive.
Purpose of the Study:
- To develop and evaluate a novel 5' Taq nuclease assay for the specific detection of Pasteurella multocida.
- To assess the assay's performance against pure cultures and field samples from poultry.
- To compare the assay's diagnostic efficiency with conventional bacteriological culture.
Main Methods:
- Design of a 5' Taq nuclease assay targeting the 16S rRNA gene with minor groove binder technology.
- Validation using P. multocida type strains, reference serovars, and Australian field isolates from poultry, pigs, and cattle.
- Testing specificity against related bacterial species and unrelated pathogens.
- Determination of the assay's limit of detection.
- Application to poultry swabs submitted for routine diagnosis and comparison with culture results.
Main Results:
- The assay demonstrated high specificity, correctly identifying all P. multocida strains and isolates while showing no reactivity with other Pasteurella species or common bacterial/viral pathogens.
- A limit of detection of approximately 10 colony-forming units (cfu) per reaction was established.
- In testing 79 poultry swabs, the assay identified 17 positive cases, whereas culture methods detected only 10.
- 62 swabs were negative by both methods.
Conclusions:
- The 5' Taq nuclease assay is a highly specific and sensitive molecular tool for diagnosing fowl cholera.
- The assay provides a rapid alternative to traditional bacteriological culture, improving diagnostic turnaround time.
- Potential applications extend to the diagnosis of P. multocida infections in cattle and pigs.
