Progress with schistosome transgenesis

Abstract

Genome sequences for Schistosoma japonicum and Schistosoma mansoni are now available. The schistosome genome encodes ~13,000 protein encoding genes for which the function of only a minority is understood. There is a valuable role for transgenesis in functional genomic investigations of these new schistosome gene sequences. In gain-of-function approaches, transgenesis can lead to integration of transgenes into the schistosome genome which can facilitate insertional mutagenesis screens. By contrast, transgene driven, vector-based RNA interference (RNAi) offers powerful loss-of-function manipulations. Our laboratory has focused on development of tools to facilitate schistosome transgenesis. We have investigated the utility of retroviruses and transposons to transduce schistosomes. Vesicular stomatitis virus glycoprotein (VSVG) pseudotyped murine leukemia virus (MLV) can transduce developmental stages of S. mansoni including eggs. We have also observed that the piggyBac transposon is transpositionally active in schistosomes. Approaches with both VSVG-MLV and piggyBac have resulted in somatic transgenesis and have lead to integration of active reporter transgenes into schistosome chromosomes. These findings provided the first reports of integration of reporter transgenes into schistosome chromosomes. Experience with these systems is reviewed herewith, along with findings with transgene mediated RNAi and germ line transgenesis, in addition to pioneering and earlier reports of gene manipulation for schistosomes.

Transfection of Platyhelminthes

“…Stable transfections have been achieved, allowing the approach to questions about the involvement of specific genes in disease pathogenesis or the identification of new target candidates for drug treatment [ 24 ]. Several reviews are available where the genomic history of schistosomes, including advances on transfection, is well organized [ 8 , 10 , 25 – 29 ]. Table 2 summarizes the progress in the transfection of S. mansoni and S. japonicum .…”

Gaining biological perspectives from schistosome genomes

“…These observations need a more stable intrinsic and inheritable source of knockdown, whereas alternatively over expression of a gene can present an interesting phenotype, both of which can be provided by transgenesis. There are a number of excellent recent papers and reviews on the many facets of achieving schistosome transgenesis [75][76][77][78][79][80], which is highly complex and very much a work in progress, but it is a critical area of future study. Of the available retrovirus-mediated transgenesis tools, murine leukemia virus (MLV) [76] is a vector that can introduce DNA into schistosomes through direct chromosomal integration, using reporter marker transgenes and sh-RNA constructs.…”

“…MLV also can be used for drug selection of successfully transduced parasites with the use of appropriate antibiotic markers [79]. Transposon piggBac [78] is another example of successful transduction in schistosomes, resulting in activation of proviral reporter transgenes and somatic transfer.…”

Genetic manipulation of schistosomes – progress with integration competent vectors

“…Advances with genetic manipulation of parasitic helminths including schistosomes have been reviewed (Grevelding, 2006; Brindley and Pearce, 2007; Kalinna and Brindley, 2007; Alrefaei et al 2011; Mann et al 2011). In brief, transgenesis of schistosomes was pioneered by Davis and co-workers who bombarded adult stages of S. mansoni with mRNA-encoding firefly luciferase and a luciferase-encoding plasmid (Davis et al 1999).…”

“…see Mates et al 2007; Damasceno et al 2010; Turan et al 2011). Several of these are now being actively investigated for utility in integrating transgenes into the schistosome genome (Alrefaei et al 2011). …”