Detection of Borrelia burgdorferi in ticks by species-specific amplification of the flagellin gene.
We developed a polymerase chain reaction (PCR) that specifically amplifies a fragment of the flagellin gene (fla) of Borrelia burgdorferi, the causative agent of Lyme disease. This fla target, amplified with nested primers, was conserved among all 80 strains of B. burgdorferi tested. Strains examined included cultures from ticks, humans, and rodents from major B. burgdorferi-endemic regions of the United States and parts of Europe and Asia. Templates from B. hermsii, B. parkeri, B. turicatae, and B. coriaceae were not amplified, nor were eukaryotic DNAs from three tick genera. Several host DNAs potentially present in a tick blood meal also were not amplified. Approximately six B. burgdorferi per PCR reaction could be detected by ethidium bromide staining of amplified DNA. Colony-raised Ixodes dammini were used to evaluate the method. One infected nymph in a pool of 40 ticks was routinely detected. The specificity of the assay for detecting B. burgdorferi-infected ticks in pools was 94% (29 of 31). This protocol should prove useful for assessing infection rates in other putative arthropod vectors.
Johnson
B J
BJ
Division of Vector-Borne Infectious Diseases, National Center for Infectious Diseases, Centers for Disease Control, Fort Collins, Colorado.
Happ
C M
CM
Mayer
L W
LW
Piesman
J
J
D000818:Animals; D001096:Arachnid Vectors / Q000382:microbiology*; D001483:Base Sequence; D015748:Borrelia burgdorferi Group / Q000235:genetics / Q000302:isolation & purification*; D015342:DNA Probes / Q000737:chemistry; D004269:DNA, Bacterial / Q000032:analysis*; D005408:Flagellin / Q000235:genetics; D008969:Molecular Sequence Data; D009758:Nymph / Q000382:microbiology; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D016133:Polymerase Chain Reaction; D012335:RNA, Ribosomal / Q000737:chemistry; D015183:Restriction Mapping; D012680:Sensitivity and Specificity; D013045:Species Specificity; D013987:Ticks / Q000382:microbiology*
D015342:DNA Probes; D004269:DNA, Bacterial; D009838:Oligodeoxyribonucleotides; D012335:RNA, Ribosomal; D005408:Flagellin