[Establishment of real-time PCR and loop-mediated isothermal amplification for detecting Cryptococcus neoformans CAP10 gene].
OBJECTIVE
To establish real-time PCR and loop-mediated isothermal amplification (LAMP) systems for detecting Cryptococcus neoformans CAP10 gene.


METHODS
Specific primers were designed targeting CAP10 gene of Cryptococcus neoformans, and the plasmid was constructed. After optimization of the reaction condition, the plasmid was quantitatively detected using real-time PCR and LAMP, and the detection sensitivity and specificity were evaluated. Clinical samples were also detected using the two methods.


RESULTS
The detection sensitivity of real-time PCR and LAMP was 6.8×10(1) and 6.8×10(3) copies, respectively. Real-time PCR yielded a higher positivity rate than LAMP. Both of the two methods showed a high detection specificity and produced negative results in the detection of Neisseria meningitidis, Candida albicans, Candida tropicalis, Aspergillus flavus, Aspergillus niger and Escherichia coli.


CONCLUSIONS
Real-time PCR is highly sensitive and specific for detecting Cryptococcus neoformans CAP10 gene but requires sophisticated equipment. LAMP, though with a relatively lower sensitivity, is simple to operate without the need of special equipment, and the result can be conveniently observed. Both of the two methods are suitable for detecting Cryptococcus neoformans and evaluating the treatment outcomes.
Southern Medical University, Guangzhou, China. hanhuisunday@163.com
Hui
H
Han
Ziyou
Z
Hu
Bingyi
B
Wu
D003455:Cryptococcus neoformans / Q000235:genetics*; D005656:Fungal Proteins / Q000235:genetics*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D010957:Plasmids; D012331:RNA, Fungal / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
C121601:CAP10 protein, Cryptococcus neoformans; D005656:Fungal Proteins; D012331:RNA, Fungal; D012333:RNA, Messenger