Study of ribosomal DNA in filarial nematodes by PCR-RFLP

Abstract

The methods used for species differentiation of filarial nematodes are Giemsa stain and acid phosphatase activity. Although both methods are practical, they require fresh samples to perform. They also require an expert to examine. Polymerase chain reaction-linked restriction fragment length polymorphism (PCR-RFLP) may be an alternative method for closely-related species differentiation that limits in morphology identification. Thus, the application of PCR-RFLP was used to study the polymorphism among of ribosomal DNA regions (rDNA) filarial species. This study was covered four species of filarial nematodes: Wuchereria bancrofti, Brugia malayi, B. pahangi and Dirofilaria immitis. The first and second internal transcribed spacers (ITS1&ITS2) of rDNA were selected to amplified and digested with restriction endonucleases separately: Acc I, Ase I or Hinƒ I for ITS1 region; Ase I or Rsa I for ITS2 region, respectively. In this study, the restriction patterns of ITS1 with Ase I could be used to distinguish filarial nematodes, W. bancrofti, B. malayi, B. pahangi and D. immitis, from one another. There were no variation detected in ITS1 and ITS2 RFLP patterns within the same filarial species although examined from the different hosts. This approach could be a useful tool for species differentiation in filarial nematodes.