Molecular differentiation and phylogeny of entomopathogenic nematodes (Rhabditida : heterorhabditidae) based on ND4 gene sequences of mitochondrial DNA

We determined partial ND4 gene sequences of mitochondrial DNA from 15 heter orhabditid nematode isolates, representing 5 species collected from differe nt regions of the world, by using polymerase chain reaction (PCR) and direc t-sequencing of PCR products. Aligned nucleotide as well as amino acid sequ ences were used to differentiate nematode species by comparing sequence div ergence and to infer phylogeny of the nematodes by using maximum parsimony and likelihood methods. Robustness of our phylogenetic trees was checked by bootstrap tests. The 15 nematode isolates can be divided into 7 haplotypes based on DNA sequences. On a larger scale, the sequence divergence reveale d 4 distinct groups corresponding to 4 described species. No sequence diver gence was detected from 5 isolates of Heterorhabditis bacteriophora or betw een Heterorhabditis marelatus to Heterorhahditis hepialius. Our sequence da ta yielded phylogenetic trees with identical topologies when different tree -building methods were used. Most relationships were also confirmed by usin g amino acid sequences in maximum parsimony analysis. Our molecular, phylog eny of Heterorhabditis species support an existing taxonomy that is based l argely on morphology and the sequence divergence of the ND4 gene permits sp ecies identification.