Project description:We report small RNA sequencing of the entomopathogenic nematode Steinernema carpocapsae. The nematodes were grown in liquid culture in homogenates of pig kidney/fat and infective juveniles were gathered. Then Galleria mellonella insect haemolymph was added to simulate insect infection, control nematodes weren't added haemolymph. Nematodes were collected after two hours after haemolymph addition. infective juveniles S. carpocapsae were incubated with and without haemolymph, three replicates
Project description:We report small RNA sequencing of the entomopathogenic nematode Steinernema carpocapsae. The nematodes were grown in liquid culture in homogenates of pig kidney/fat and infective juveniles were gathered. Then Galleria mellonella insect haemolymph was added to simulate insect infection, control nematodes weren't added haemolymph. Nematodes were collected after two hours after haemolymph addition.
Project description:Entomopathogenic nematode infective juveniles activate when they infect a host and begin releasing excreted-secreted products. These are the excreted-secreted proteins detected by mass spec from non-activated/naive Steinernema feltiae infective juveniles(infective juveniles not exposed to host or host tissue).
Project description:Entomopathogenic nematode infective juveniles activate when they infect a host and begin releasing excreted-secreted products. These are the excreted-secreted proteins detected by mass spec from activated Steinernema feltiae infective juveniles. The infective juveniles were activated in vitro by exposure to insect-host tissue for 6 hours.
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.
Project description:We investigate the potential metabolic costs for Steinernema nematode in relation to the maintenance and vectoring of their Xenorhabdus endosymbionts. we performed a comparative dual RNA-seq analysis of infective juveniles (IJs) of two symbiotic partners: S. carpocapsae-X. nematophila and S. puntauvense-X. bovienii.
Project description:Entomopathogenic nematodes (Rhabditida; Steinernematidae and Heterorhabditidae) are insect parasites which are of economic importance due to their use as biocontrol agents. The third larval stage, infective juveniles (IJs) leave the natal host and move in the soil to locate a new insect host. Both families are associated with mutualistic bacteria, which are released into the haemolymph of the host insect, killing the insect and providing nutrition for nematode development and reproduction, and are thus categorised as entomopathogenic nematodes. Similarities between these two families are due to convergent evolution associated with this lifestyle, rather than common ancestry. Heterorhabditids are closely related to the vertebrate parasites Strongylida (Adams and Nguyen, 2002) and to Caenorhabditis elegans, whereas steinernematids are more closely related to Strongyloididae. The IJ stage of parasitic nematodes is analogous to the dauer stage of C. elegans; both are developmentally arrested, stress resistant stages that disperse to colonise new hosts or food resources, respectively. By investigating the molecular mechanisms and consequences of temperature acclimation and aging in EPN IJs, insights into survival and the changes induced by low temperature exposure may be gained. Label free quantitative (LFQ) massspectrometery-based proteomics facilitates the identification and quantification of thousands of proteins in a single run. Such data allows for the comparison of the proteomes of EPN IJs after conditioning via gene ontology mapping and functional enrichment analysis. Understanding how the IJ proteome is affected by temperature and aging can provide a molecular basis for the wide array of phenotypes and behaviours these organisms may adopt. This study aims to provide proteomic data which may elucidate the molecular mechanisms underlying the phenotypic plasticity which EPN IJs of two distantly related species exhibit.
Project description:Parasitism is a major ecological niche for a variety of nematodes. Multiple nematode lineages have specialized as pathogens, including deadly parasites of insects that are used in biological control. We have sequenced and analyzed the draft genomes and transcriptomes of the entomopathogenic nematode Steinernema carpocapsae and four congeners (S. scapterisci, S. monticolum, S. feltiae, S. glaseri) distantly related to Caenorhabditis elegans. We used these genomes to establish phylogenetic relationships, explore gene conservation across species, identify genes uniquely expanded in insect parasites, and to identify conserved non-coding regulatory motifs that influence similar biological processes. Protein domain analysis of these genomes reveals a striking expansion of numerous putative parasitism genes including certain protease and protease inhibitor families as well as fatty acid- and retinol-binding proteins. We identify rapid evolution and expansion of the important developmental Hox gene cluster and identify novel conserved non-coding regulatory motifs associated with orthologous genes in Steinernema and Caenorhabditis. The deep conservation of the network of non-coding DNA motifs between these two genera for a subset of orthologous genes involved in neurogenesis and embryonic development suggests that a kernel of protein-DNA relationships is conserved through nematode evolution. We analyzed the gene expression of a total of 24 RNA-seq samples from 3 nematode species( S. carpocapsae, S. feltiae, and C. elegans) for comparative analysis. We collected the RNA at four developmental time points (mixed embryo, L1, infective juvenile/dauer, young adult) for each species in replicates.
