title: Development and application of a PCR diagnostic assay for the accurate and rapid identification of the nematophagous fungus Duddingtonia flagrans.
abstract: The nematophagous fungus, Duddingtonia flagrans is a potential biocontrol agent against nematode parasites of ruminants. Improved methods for the rapid and accurate detection of D. flagrans would aid the evaluation of this fungus as a biocontrol agent and its suitability for environmental release. To date, detection and identification of D. flagrans is reliant on morphological methods, which can be laborious, time-consuming, and error prone. In this study, a PCR assay using species-specific primers located in the ITS regions was developed for the rapid and accurate identification of D. flagrans. The PCR assay was specific to five different isolates of D. flagrans and was capable of detecting a minimum concentration of 100 chlamydospores per gram of soil. In contrast to cultured-based detection and identification methods, this assay is amenable to high throughput screening of environmental samples. The assay detected D. flagrans in faecal, leaf litter, and soil samples collected from 80% of the Irish farms tested indicating that the fungus is abundant in Ireland.
authors.affiliation: Teagasc, Animal Production Research Centre, Mellows Campus, Athenry, Co. Galway, Ireland.
authors.forename: Paula
authors.initials: P
authors.lastname: Kelly
authors.forename: Barbara
authors.initials: B
authors.lastname: Good
authors.forename: Richard
authors.initials: R
authors.lastname: Fitzpatrick
authors.forename: J P
authors.initials: JP
authors.lastname: Hanrahan
authors.forename: Theo D T
authors.initials: TD
authors.lastname: de Waal
mesh_terms: D000818:Animals; D001203:Ascomycota / Q000145:classification / Q000235:genetics / Q000302:isolation & purification / Q000502:physiology; D017931:DNA Primers; D004271:DNA, Fungal / Q000032:analysis / Q000302:isolation & purification; D021903:DNA, Ribosomal Spacer / Q000032:analysis; D005243:Feces / Q000382:microbiology; D016533:Mycological Typing Techniques; D009348:Nematoda / Q000382:microbiology*; D018515:Plant Leaves / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D012988:Soil Microbiology; D013045:Species Specificity; D013172:Spores, Fungal / Q000302:isolation & purification / Q000502:physiology; D013997:Time Factors
chemical_list: D017931:DNA Primers; D004271:DNA, Fungal; D021903:DNA, Ribosomal Spacer