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Detection of nematode antagonistic bacteria by fluorogenic molecular probes *
Author(s) -
Ciancio A.,
Leonetti P.,
Sialer M. M. Finetti
Publication year - 2000
Publication title -
eppo bulletin
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.327
H-Index - 36
eISSN - 1365-2338
pISSN - 0250-8052
DOI - 10.1111/j.1365-2338.2000.tb00948.x
Subject(s) - amplicon , biology , polymerase chain reaction , bacteria , dna , nematode , hybridization probe , 16s ribosomal rna , oligomer restriction , population , molecular probe , molecular diagnostics , oligonucleotide , microbiology and biotechnology , genetics , gene , ecology , demography , sociology
Last‐generation DNA probes include molecules yielding a fluorogenic emission through an intramolecular change occurring after hybridization to a complementary sequence. They display a high sequence specificity and may detect even single‐base mutations in polymerase chain reaction (PCR) amplification products. We applied Scorpion primers for the detection of an unculturable nematode‐parasitic bacterium, Pasteuria sp., with potential as a biological control agent. A 16S rDNA oligonucleotide sequence unique to Pasteuria spp. was used to detect the parasite in juveniles of Heterodera goettingiana or in soil. The parasitized nematodes came from a population with a Pasteuria prevalence of 40–80% and were individually checked for parasitism. Probes with 6‐carboxy‐fluorescein (FAM) at the 5’terminus, used for PCR with nematodes or soil, successfully detected the parasite from both samples. The amplification of the expected 139bp fragment was shown both by fluorescence observed under UV excitation in the eppendorfs and by gel electrophoresis of the corresponding amplicons. The potential of this detection method for the study of unculturable bacteria is discussed.