title: Development of a loop-mediated isothermal amplification (LAMP) assay for the sensitive detection of Haemonchus contortus eggs in ovine faecal samples.
abstract: A major constraint on the effective control and management of helminth parasites in livestock is the lack of rapid and reliable diagnostic tests to identify the parasite species responsible for disease and to allow informed treatment decisions to be made. In the present study, we have developed a novel DNA-based assay for the detection of Haemonchus contortus eggs in ovine faecal samples, using loop-mediated isothermal amplification (or LAMP). LAMP allows for rapid detection of H. contortus DNA under isothermal incubation conditions. The robust nature of this assay negates the need for extensive DNA extraction, allowing amplification from relatively crude samples. Preliminary results suggest that LAMP is highly specific, and does not cross-react with DNA from other common co-infecting parasites. The Haemonchus LAMP assay is also highly sensitive, exhibiting a 10 times lower detection limit than the equivalent PCR; 10(-5) ng/μl and 10(-4) ng/μl DNA, respectively, allowing detection in a faecal samples containing two Haemonchus eggs per gram. Translation of this assay onto a real-time platform provided rapid results and highlighted its potential as a quantitative assay which could inform treatment decisions in the future.
authors.forename: Lynsey
authors.initials: L
authors.lastname: Melville
authors.forename: Fiona
authors.initials: F
authors.lastname: Kenyon
authors.forename: Sajid
authors.initials: S
authors.lastname: Javed
authors.forename: Iain
authors.initials: I
authors.lastname: McElarney
authors.forename: Janina
authors.initials: J
authors.lastname: Demeler
authors.forename: Philip
authors.initials: P
authors.lastname: Skuce
mesh_terms: D000818:Animals; D017931:DNA Primers / Q000235:genetics; D005243:Feces / Q000469:parasitology; D006188:Haemonchiasis / Q000175:diagnosis* / Q000469:parasitology; D006189:Haemonchus / Q000235:genetics / Q000302:isolation & purification*; D057230:Limit of Detection; D021141:Nucleic Acid Amplification Techniques / Q000662:veterinary; D010063:Ovum; D016133:Polymerase Chain Reaction / Q000662:veterinary; D012680:Sensitivity and Specificity; D012756:Sheep
chemical_list: D017931:DNA Primers