Molecular techniques for the identification of nematodes
In many cases nematodes species can be easily identified by morphological means. However, in the case of morphologically conserved species (such as those found in the entomopathogenic nematode families Steinernematidae and Heterorhabditidae) or, when a large number of samples have to be identified, it is desirable to use a rapid, simple and reliable identification method. DNA-based molecular biological techniques, such as restriction fragment length polymorphism analysis, satisfy such criteria with the added benefit that it is possible (even for a relative novice) to obtain an accurate identification from any life cycle stage with as little material as a single nematode. Therefore the use of molecular techniques are becoming an increasingly important 'extra tool' in the field of taxonomy.
An ideal choice for molecular taxonomic purposes is the ribosomal DNA repeat unit. It is present as a multi-copy tandem repeat and contains highly conserved regions, the rRNA genes, and potentially highly variable regions, the internal transcribed spacer (ITS) and non-transcribed spacer (NTS) regions which separate the genes. This arrangement means that these regions are ideal for amplification by PCR as the conserved rRNA genes allow for the construction of oligonucleotide primers which are 'universal' in nature and enable the spacer regions, which are rich in interspecific polymorphisms, to be amplified.
At IIP we currently use RFLP analysis of the ITS region for the identification of Meloidogyne spp., Steinernema spp. and Heterorhabditis spp. although the technique could be adapted for the identification of any nematode.
In addition to species identification we also use the PCR based techniques RAPD and AFLP for the differentiation of populations of the parasitic nematodes Radopholus similis (a parasite of bananas) and the canid parasite Toxocara canis.
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