Wang Huan,Huang Qing,Shi Dachuan,Chuai Zhenran,Fu Weiling.Rapid detection of Pseudomonas aeruginosa by color loop-mediated isothermal amplification[J].J Amry Med Univ (J Third Mil Med Univ),2012,34(22):2264-2268.
Wang Huan,Huang Qing,Shi Dachuan,Chuai Zhenran,Fu Weiling.Rapid detection of Pseudomonas aeruginosa by color loop-mediated isothermal amplification[J].J Amry Med Univ (J Third Mil Med Univ),2012,34(22):2264-2268.DOI:
Rapid detection of Pseudomonas aeruginosa by color loop-mediated isothermal amplification
Objective To establish a rapid and sensitive method of color loop-mediated isothermal amplification (LAMP) for detecting Pseudomonas aeruginosa (P. aeruginosa). Methods LAMP method with hydroxy naphthol blue (HNB) and calcein was designed to amplify and identify the OprL gene of P. aeruginosa. Three pairs of LAMP primers (inner
outer and ring primers) were designed according to the eight zones of the OprL gene. A positive reaction was indicated by a color change before and after the reaction
and was verified by agarose gel electrophoresis. The reaction conditions and system were optimized. The sensitivity and specificity of the detection method were evaluated
and were compared with those of conventional PCR. Both LAMP and PCR were applied to detect clinical specimens. Results A LAMP method for detecting P. aeruginosa was successfully established. The LAMP method showed 100% specificity for P. aeruginosa without other bacteria amplification
and the sensitivity (with the minimum detection limit of 17.414 pg/μl) for the detection of P. aeruginosa was 10-fold higher than that of PCR (with the minimum detection limit of 174.14 pg/μl). The detection rates of LAMP and PCR in clinical specimens were the same (27/27). Conclusion LAMP method
in which the amplification can be completed within 60 min
is faster than PCR. The established LAMP method in this study enables rapid
sensitive and specific detection of P. aeruginosa
and can be applied for grass roots and small-scale laboratories as well as field surveillance.