Amplification of repetitive DNA for the specific detection of Naegleria fowleri.
By using hybridization at low C0t values, a genomic library on Naegleria fowleri was screened for clones containing repetitive DNA. Partial sequence information from a repetitive clone, Nf9, showed sequence homologies with the mitochondrial ATPase 6 subunit from yeasts and other organisms. Synthetic DNA primers were selected and tested in amplification reactions. Nonstringent hybridization conditions were defined which allowed amplification of N. fowleri DNA and reduced amplification of DNA from nonpathogenic Naegleria species. Stringent conditions were selected which allowed detection only of N. fowleri. Identity of the amplified DNA was confirmed by using internal restriction sites and an internal primer. In a blind study, tissue from mice experimentally infected with N. fowleri was specifically detected by using stringent hybridization conditions.
McLaughlin
G L
GL
Department of Veterinary Pathobiology, College of Veterinary Medicine, University of Illinois, Urbana 61801.
Vodkin
M H
MH
Huizinga
H W
HW
D000562:Amebiasis / Q000175:diagnosis; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D016054:DNA, Protozoan / Q000235:genetics*; D006801:Humans; D008590:Meningoencephalitis / Q000175:diagnosis; D008969:Molecular Sequence Data; D009253:Naegleria / Q000235:genetics* / Q000302:isolation & purification / Q000472:pathogenicity; D016133:Polymerase Chain Reaction; D012091:Repetitive Sequences, Nucleic Acid
D016054:DNA, Protozoan