Differentiation of Setaria digitata and Setaria labiatopapillosa using molecular markers.

5S rRNA intergenic regions of Setaria digitata and Setaria labiatopapillosa were PCR amplified with primers designed from the 5S rRNA gene of Brugia malayi. The ladder-like banding patterns obtained for the amplifications were distinctly different for the two species. Four amplified products were cloned into the pBS vector and completely sequenced. DNA clones from two individual samples of S. digitata, Sd4 and Sd6, showed 97% sequence homology to each other. All sequenced clones showed the presence of the spliced leader (SL) RNA gene with a 22 nucleotide spliced leader sequence. The phylogenetic tree constructed using these data and the 5S rRNA intergenic regions of several other filarial nematodes showed the Setaria species sharing a branch with Dirofilaria. RAPD-PCR analyses identified 107 bands of which 86 were polymorphic (80%). A dendrogram constructed for S. digitata and S. labiatopapillosa separated the two species into two distinct clusters. The polymorphic loci identified by the RAPD-PCR analyses can be studied further to develop species-specific probes/PCR primers for the identification of each species.