immunomagnetic separation/loop‐mediated isothermal amplification method for rapid direct detection of thermotolerant Campylobacter spp. during poultry production

AIMS: To develop a rapid test for thermotolerant Campylobacter in poultry faeces. METHODS AND RESULTS: The reported method is based on immunomagnetic separation and loop‐mediated isothermal DNA amplification (IMS/LAMP). This LAMP assay is specific (demonstrated using 10 Campylobacter strains and 13...

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Bibliographic Details
Published in:Journal of applied microbiology 2016-02, Vol.120 (2), p.469-477
Main Authors: Romero, M.R, D'Agostino, M, Arias, A. Pérez, Robles, S, Casado, C. Fernández, Iturbe, L. Orueta, Lerma, O. Gurrutxaga, Andreou, M, Cook, N
Format: Article
Language:English
Subjects:
Animals
Campylobacter
Campylobacter - classification
Campylobacter - genetics
Campylobacter - isolation & purification
Campylobacter Infections - diagnosis
Campylobacter Infections - microbiology
Campylobacter Infections - veterinary
Chickens - microbiology
detection
DNA
enrichment culture
farms
Feces
Feces - microbiology
flocks
food chain
food safety
genome
heat tolerance
human resources
immunomagnetic separation
Immunomagnetic Separation - methods
internal amplification control
LAMP
loop-mediated isothermal amplification
Microbiology
Nucleic Acid Amplification Techniques - methods
Poultry
Poultry Diseases - diagnosis
Poultry Diseases - microbiology
poultry production
probability
rapid methods
Sensitivity and Specificity
turkeys
Turkeys - microbiology
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Summary:AIMS: To develop a rapid test for thermotolerant Campylobacter in poultry faeces. METHODS AND RESULTS: The reported method is based on immunomagnetic separation and loop‐mediated isothermal DNA amplification (IMS/LAMP). This LAMP assay is specific (demonstrated using 10 Campylobacter strains and 13 non‐Campylobacter bacterial species) and sensitive (95% probability of detecting 22 genome copies). A competitive internal amplification control (IAC) has been incorporated to give unambiguous determination of negative results. Immunoseparation of Campylobacter allows direct LAMP detection from poultry boot swab samples in 90 min without enrichment or DNA purification (74% probability of detecting 10⁴ CFU ml⁻¹ of a boot swab suspension). The analysis of 17 samples from commercial turkey farms showed 100% correlation with parallel results obtained by standard microbiological methods. CONCLUSIONS: A rapid test has been developed for direct detection of thermotolerant Campylobacter spp. in boot swab samples, thus bypassing culture enrichment or DNA extraction. The test has potential to be carried out by farm personnel on site. SIGNIFICANCE AND IMPACT OF THE STUDY: The method offers an inexpensive approach to monitor poultry infection in near real time, assisting flock management and controls to prevent introduction of Campylobacter into the food chain.
ISSN:1364-5072
1365-2672
DOI:10.1111/jam.13008