Journal of applied microbiology · 2018
One step for Legionella pneumophila detection in environmental samples by DNA-gold nanoparticle probe.
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Nuthong B, Wilailuckana C, Tavichakorntrakool R, Boonsiri P, Daduang S, Bunyaraksyotin G, Suphan O, Daduang J
Аннотация
AIMS: To develop and evaluate a DNA-gold nanoparticle (DNA-AuNP) probe assay to detect Legionella pneumophila, which causes Legionnaires' disease, compared with the gold standard culture method.
METHODS AND RESULTS: Gold nanoparticles (AuNPs) were conjugated with DNA probes to detect the mip gene of L. pneumophila. The DNA-AuNP probe assay was evaluated for its specificity, sensitivity and stability. The results showed that only L. pneumophila mixed with this probe resulted in a red solution that was easily detected by the naked eye, and the colour was stable when 10 mmol l-1 MgSO4 was added. The 100 Legionella isolates and 10 other bacteria led to 100% specificity. Compared with the culture method, our method showed a 100% negative predictive value, 100% sensitivity (kappa = 0·87), and a detection limit of 4·5 ng DNA μl-1 with a 6-min response time for the 124 colonies suspected of being Legionella. The DNA-AuNP probe reagents were stable for more than 6 months.
CONCLUSIONS: The developed DNA-AuNP probe assay has good negative predictive value, sensitivity, rapidity and ease of use, which is helpful for ruling out negative samples.
SIGNIFICANCE AND IMPACT OF THE STUDY: The DNA-AuNP probe assay can detect the mip gene of L. pneumophila. Therefore, it may be an alternative method for screening colonies suspected of being L. pneumophila.
METHODS AND RESULTS: Gold nanoparticles (AuNPs) were conjugated with DNA probes to detect the mip gene of L. pneumophila. The DNA-AuNP probe assay was evaluated for its specificity, sensitivity and stability. The results showed that only L. pneumophila mixed with this probe resulted in a red solution that was easily detected by the naked eye, and the colour was stable when 10 mmol l-1 MgSO4 was added. The 100 Legionella isolates and 10 other bacteria led to 100% specificity. Compared with the culture method, our method showed a 100% negative predictive value, 100% sensitivity (kappa = 0·87), and a detection limit of 4·5 ng DNA μl-1 with a 6-min response time for the 124 colonies suspected of being Legionella. The DNA-AuNP probe reagents were stable for more than 6 months.
CONCLUSIONS: The developed DNA-AuNP probe assay has good negative predictive value, sensitivity, rapidity and ease of use, which is helpful for ruling out negative samples.
SIGNIFICANCE AND IMPACT OF THE STUDY: The DNA-AuNP probe assay can detect the mip gene of L. pneumophila. Therefore, it may be an alternative method for screening colonies suspected of being L. pneumophila.