Rapid visual detection of Mycobacterium avium subsp. paratuberculosis by recombinase polymerase amplification combined with a lateral flow dipstick
Journal of Veterinary Science
;
: 242-250, 2018.
Article
in English
| WPRIM
| ID: wpr-758796
ABSTRACT
Paratuberculosis (Johne's disease) is a chronic debilitating disease of domestic and wild ruminants. However, widespread point-of-care testing is infrequent due to the lack of a robust method. The isothermal recombinase polymerase amplification (RPA) technique has applied for rapid diagnosis. Herein, RPA combined with a lateral flow dipstick (LFD) assay was developed to estimate DNA from Mycobacterium avium subsp. paratuberculosis. First, analytical specificity and sensitivity of the RPA-nfo primer and probe sets were assessed. The assay successfully detected M. paratuberculosis DNA in 30 min at 39℃ with a detection limit of up to eight copies per reaction, which was equivalent to that of the real-time quantitative polymerase chain reaction (qPCR) assay. The assay was specific, as it did not amplify genomes from five other Mycobacterium spp. or five pathogenic enteric bacteria. Six hundred-twelve clinical samples (320 fecal and 292 serum) were assessed by RPA-LFD, qPCR, and enzyme-linked immunosorbent assay, respectively. The RPA-LFD assay yielded 100% sensitivity, 97.63% specificity, and 98.44% concordance rate with the qPCR results. This is the first report utilizing an RPA-LFD assay to visualize and rapidly detect M. paratuberculosis. Our results show this assay should be a useful method for the diagnosis of paratuberculosis in resource-constrained settings.
Full text:
Available
Index:
WPRIM (Western Pacific)
Main subject:
Paratuberculosis
/
Ruminants
/
DNA
/
Enzyme-Linked Immunosorbent Assay
/
Polymerase Chain Reaction
/
Sensitivity and Specificity
/
Genome
/
Recombinases
/
Diagnosis
/
Enterobacteriaceae
Type of study:
Diagnostic study
Limits:
Animals
Language:
English
Journal:
Journal of Veterinary Science
Year:
2018
Type:
Article
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