Project description:Third-stage larvae (L3s) of the canine hookworm, Ancylostoma caninum, endure a period of arrested development preceding transmission to a host. Many of the mRNAs up-regulated at this stage are likely to encode proteins that act at the host-parasite interface and facilitate the transition from a free-living to a parasitic larva. The initial phase of the infection of a mammalian host by A. caninum L3s (herein termed “activation”) can be mimicked in vitro by culturing L3s in serum-containing medium. The mRNAs differentially transcribed between activated and non-activated L3s were identified by suppression subtractive hybridisation (SSH). The analysis of these mRNAs on a custom oligonucleotide microarray printed with the SSH ESTs and publicly available A. caninum ESTs (non-subtracted) yielded a total of 602 differentially expressed mRNAs, of which the most highly represented sequences (27) encoded products belonging to the pathogenesis-related protein (PRP) superfamily and different mechanistic classes of proteases. Comparison of these A. caninum mRNAs with those of Caenorhabditis elegans larvae exiting from developmental (dauer) arrest demonstrated unexpectedly large differences with respect to gene ontology profiles. C. elegans L3 exiting developmental arrest up-regulated the expression of collagens and other (mostly intracellular) molecules involved in growth and development. Such mRNAs are virtually absent from activated hookworm larvae, and instead are represented by an inordinately large number of mRNAs encoding extracellular proteins, suggesting that many of the activation-associated hookworm mRNAs are involved in host-parasite interactions. The near absence of mRNAs associated with reproduction, growth and development among activated hookworm L3s probably reflects their ability to further arrest (i.e. undergo hypobiosis) in tissues of non-permissive hosts or in the external environment when conditions for transmission are unfavourable. Although this should not necessarily invalidate C. elegans dauer exit as a model for hookworm activation, it provides substantial information on the limitations of this free-living nematode as a model organism for the transition of nematode larvae from a free-living to a parasitic state. Keywords: Comparative transcriptomic hybridisation
Project description:Recent advances in high throughput sequencing methodologies allow the opportunity to probe in depth the transcriptomes of organisms including N. caninum. In this project, we are using Illumina sequencing technology to analyze the transcriptome (RNA-Seq) of experimentally accessible stages (e.g. tachyzoites at different times points) of N. caninum NCLiv. The aim is to make transcriptional landscape maps at different time points at different life cycle stages of N. caninum and compare it with equivalent datasets from the closely related parasite Toxoplasma gondii
