PCR Primers with Enhanced Specificity for Nematode-Trapping Fungi ( Orbiliales )

• Published in: Microbial ecology Vol. 58; no. 1; pp. 117 - 128
• Main Authors: Smith, Matthew E., Jaffee, Bruce A.
• Format: Journal Article
• Language: English
• Published: New York Springer Science + Business Media, Inc 01.07.2009; Springer-Verlag; Springer; Springer Nature B.V
• Subjects: Animals; Ascomycota; Ascomycota - classification; Ascomycota - genetics; Ascomycota - isolation & purification; Biological and medical sciences; Biological taxonomies; Biomedical and Life Sciences; DNA Primers; DNA, Fungal - genetics; DNA, Ribosomal - genetics; Ecological studies; Ecology; ENVIRONMENTAL MICROBIOLOGY; Fundamental and applied biological sciences. Psychology; Fungi; Geoecology/Natural Processes; Leaf litter; Life history; Life Sciences; Microbial Ecology; Microbiology; Nature Conservation; Nematoda; Nematoda - microbiology; Nematodes; Nematophagous fungi; Original Article; Parasites; Phylogeny; Polymerase chain reaction; Polymerase Chain Reaction - methods; Product category rules; RNA, Ribosomal, 28S - genetics; Sensitivity and Specificity; Soil fungi; Soil Microbiology; Soil nematodes; Soil samples; Taxa; Trapping; Water Quality/Water Pollution; Restriction Fragment Length Polymorphism; Ascomycota; Clone Library; Restriction Fragment Length Polymorphism Pattern; Leaf Litter; Life history; Litter; Primer; Parasite; Method; Capture; Fungi; Polymerase chain reaction; Substrate; Trapping; Specificity; Pest; Helmintha; Nemathelminthia; Predator; Invertebrata; Detection; Nematoda; Culture
• ISSN: 0095-3628; 1432-184X
• DOI: 10.1007/s00248-008-9453-0

Nematode-trapping fungi, a monophyletic lineage within the Orbiliales (Ascomycota), use specialized structures to capture and consume nematodes in soil, leaf litter, and other substrates. These fungi have been studied both because of their unique predatory life history and because they are potential control agents of important plantand animal-parasitic nematodes. Ecological studies of nematode-trapping fungi have primarily used culture-based methods, but molecular detection techniques are now available and should be useful for studying this group. We developed Orbiliales-specific PCR primers for the ITS and 28s rDNA to directly detect nematode-trapping fungi without culturing and also to screen fungal isolates for phylogenetic placement in the Orbiliales. We used these primers to selectively amplify, clone, and sequence Orbiliales DNA extracted from soil, litter, and wood, and we compared the results of molecular detection with those obtained using a culture-based method. Of the eight species of nematode-trapping Orbiliales detected with the culturebased assay, only three were detected with PCR. The molecular assay, however, detected 18 species of uncultured Orbiliales, many of which are closely related to nematode-trapping fungi and fungal parasites of nematode eggs. Our results suggest that the combined use of Orbiliales-specific primers and culture-based techniques may benefit future studies of nematophagous fungi.