title: Diagnosis of brugian filariasis by loop-mediated isothermal amplification.
abstract: In this study we developed and evaluated a Brugia Hha I repeat loop-mediated isothermal amplification (LAMP) assay for the rapid detection of Brugia genomic DNA. Amplification was detected using turbidity or fluorescence as readouts. Reactions generated a turbidity threshold value or a clear visual positive within 30 minutes using purified genomic DNA equivalent to one microfilaria. Similar results were obtained using DNA isolated from blood samples containing B. malayi microfilariae. Amplification was specific to B. malayi and B. timori, as no turbidity was observed using DNA from the related filarial parasites Wuchereria bancrofti, Onchocerca volvulus or Dirofilaria immitis, or from human or mosquito. Furthermore, the assay was most robust using a new strand-displacing DNA polymerase termed Bst 2.0 compared to wild-type Bst DNA polymerase, large fragment. The results indicate that the Brugia Hha I repeat LAMP assay is rapid, sensitive and Brugia-specific with the potential to be developed further as a field tool for diagnosis and mapping of brugian filariasis.
authors.affiliation: New England Biolabs, Ipswich, Massachusetts, United States of America.
authors.forename: Catherine B
authors.initials: CB
authors.lastname: Poole
authors.forename: Nathan A
authors.initials: NA
authors.lastname: Tanner
authors.forename: Yinhua
authors.initials: Y
authors.lastname: Zhang
authors.forename: Thomas C
authors.initials: TC
authors.lastname: Evans
authors.forename: Clotilde K S
authors.initials: CK
authors.lastname: Carlow
mesh_terms: D000818:Animals; D017178:Brugia malayi / Q000235:genetics / Q000302:isolation & purification*; D018509:DNA, Helminth / Q000235:genetics / Q000302:isolation & purification; D004285:Dogs; D005368:Filariasis / Q000175:diagnosis* / Q000469:parasitology*; D005470:Fluorometry; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D009391:Nephelometry and Turbidimetry; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D010274:Parasitology / Q000379:methods*; D012680:Sensitivity and Specificity; D013696:Temperature; D013997:Time Factors
chemical_list: D018509:DNA, Helminth