Project description:Purpose: The goal of this study was to track fluctuations in gene expression across all defined life cycle stages of the cattle lungworm Dictyocaulus viviparus. Methods: Batches of worms from a Hannover, Germany strain were harvested in duplicate as eggs, L1, L2, L3, L4, hypobiotic L5, mixed-sex L5, male L5, female L5, mature males and mature females. RNA was isolated, reverse transcribed and sequenced on the Illumina platform. Finally, reads were mapped to the D. viviparus draft genome assembly and read counts associated with each feature were used to estimate gene expression levels and predict differential gene expression at various transitional points across the life cycle. Results: Principal component analyses indicated that major gene expression shifts occur upon hatching, infection of the bovid host, and sexual maturation.
Project description:Entomopathogenic nematodes (EPNs) are unique parasitic nematodes due to their symbiosis with entomopathogenic bacteria and their ability to kill insect hosts quickly after infection. Although it has been widely believed that EPNs rely on their bacterial partners for killing insect hosts, compelling evidence from previous studies challenges this model. We developed an improved method of activating millions of Steinernema carpocapsae infective juveniles (IJs) in vitro to harvest excreted/secreted (ES) proteins for bioactivity tests and proteomics analysis. We found that a low dose of the ES proteins from early activated nematodes is lethal to Drosophila melanogaster adults within 2-6 hours. We analyzed the protein composition of this venom using mass spectrometry and identified 472 proteins. Many of these venom proteins share high homology with those of vertebrate-parasitic nematodes. Among many different families of proteins found in the venom, proteases and protease inhibitors are especially abundant. Some toxin-related proteins such as Shk domain-containing proteins were also detected. We further analyzed the transcriptomes of individual non-activated IJs and nematodes that were activated in vitro and in vivo, which revealed a dramatic shift in gene expression during IJ activation. By comparing the whole transcriptomes and the genes encoding venom proteins between the in vitro and in vivo activated nematodes, we confirmed that the in vitro activation is a good approximation of the in vivo process. In summary, our findings strongly support a new model that S. carpocapsae and likely other Steinernema EPNs have a more active role in contributing to the pathogenicity of the nematode-bacterium complex than simply relying on their symbiotic bacteria. Furthermore, we propose that EPNs are a good model system for investigating vertebrate- and human-parasitic nematodes, especially regarding the function of ES products.
