Brugia malayi: transient transfection by microinjection and particle bombardment

To develop a method for the introduction of DNA into filarial parasites, several methods that have proven successful in other organisms were evaluated for their ability to transform Brugia malayi. Luciferase activity was detectable in embryos bombarded with gold particles coated with a construct consisting of a luciferase reporter gene under the control of the 5S rRNA intergenic spacer (SL promoter). Similar results were seen in adult parasites and infective larvae bombarded with this construct, or in adult female parasites microinjected with the plasmid. In similar experiments employing the SL promoter driving a green fluorescent protein (GFP) reporter, expression of the reporter was detectable in the intrauterine embryos of the microinjected adult parasites, and in the sub-cuticular tissues of biolistically transfected adult female parasites. A similar pattern of GFP expression to that seen in the SL promoter construct transfected parasites was noted in parasites transfected with constructs consisting of the upstream domain derived from an aspartyl aminoacyl tRNA synthetase gene of B. malayi. The ability to transfect B. malayi embryos may provide a foundation for studies of the regulation of gene expression and function in these organisms. Index Descriptors and Abbreviations: filarial nematodes; spliced leader; particle bombardment; microinjection; bp, base pairs; EST, expressed sequence tag; GFP, green fluorescent protein; L3, Infective larvae; nt, nucleotides; PCR, polymerase chain reaction; rRNA, ribosomal RNA; SL, spliced leader