Rapid identification of Acanthamoeba from contact lens case using loop-mediated isothermal amplification method.
A method employing loop-mediated isothermal amplification (LAMP) of 18S ribosomal RNA gene was developed to detect Acanthamoeba in contact lens cases. A prevalence of 7% (10/150) was detected, with 100% sensitivity and 100% specificity when compared with the standard culture technique. Using visual inspection of turbidity a minimum of 10pg of Acanthamoeba DNA could be detected, 10 times more sensitive than quantitative PCR employing two of the LAMP primers. The production of LAMP amplicons was confirmed by gel-electrophoresis and ethidium bromide staining. The LAMP procedure takes less than 2h to perform and will be useful for incorporation into a point-of-care screening of suspected Acanthamoeba infection.
Lek-Uthai
Usa
U
Department of Parasitology, Faculty of Public Health, Mahidol University, 420/1 Rajavithi Road, Rajathewee District, Bangkok 10400, Thailand. phulu@mahidol.ac.th
Passara
Rangsima
R
Roongruangchai
Kosol
K
Buddhirakkul
Prayute
P
Thammapalerd
Nitaya
N
D000048:Acanthamoeba / Q000235:genetics / Q000302:isolation & purification*; D000818:Animals; D003261:Contact Lenses / Q000469:parasitology*; D017931:DNA Primers / Q000737:chemistry; D016054:DNA, Protozoan / Q000032:analysis; D004275:DNA, Ribosomal / Q000032:analysis; D006801:Humans; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D016053:RNA, Protozoan / Q000235:genetics; D012337:RNA, Ribosomal, 18S / Q000235:genetics; D012680:Sensitivity and Specificity; D013997:Time Factors
D017931:DNA Primers; D016054:DNA, Protozoan; D004275:DNA, Ribosomal; D016053:RNA, Protozoan; D012337:RNA, Ribosomal, 18S