The parasitic Nematomorph hairworm, Spinochordodes tellinii (Camerano) develops inside the terrestrial grasshopper, Meconema thalassinum (De Geer) (Orthoptera: Tettigoniidae), changing the insect's responses to water. The resulting aberrant behaviour makes infected insects more likely to jump into an aquatic environment where the adult parasite reproduces. We used proteomics tools (i.e. two-dimensional gel electrophoresis (2-DE), computer assisted comparative analysis of host and parasite protein spots and MALDI-TOF mass spectrometry) to identify these proteins and to explore the mechanisms underlying this subtle behavioural modification. We characterized simultaneously the host (brain) and the parasite proteomes at three stages of the manipulative process, i.e. before, during and after manipulation. For the host, there was a differential proteomic expression in relation to different effects such as the circadian cycle, the parasitic status, the manipulative period itself, and worm emergence. For the parasite, a differential proteomics expression allowed characterization of the parasitic and the free-living stages, the manipulative period and the emergence of the worm from the host. The findings suggest that the adult worm alters the normal functions of the grasshopper's central nervous system (CNS) by producing certain 'effective' molecules. In addition, in the brain of manipulated insects, there was found to be a differential expression of proteins specifically linked to neurotransmitter activities. The evidence obtained also suggested that the parasite produces molecules from the family Wnt acting directly on the development of the CNS. These proteins show important similarities with those known in other insects, suggesting a case of molecular mimicry. Finally, we found many proteins in the host's CNS as well as in the parasite for which the function(s) are still unknown in the published literature (www) protein databases. These results support the hypothesis that host behavioural changes are mediated by a mix of direct and indirect chemical manipulation.

Despite increasing evidence of host phenotypic manipulation by parasites, the underlying mechanisms causing infected hosts to act in ways that benefit the parasite remain enigmatic in most cases. Here, we used proteomics tools to identify the biochemical alterations that occur in the head of the cricket Nemobius sylvestris when it is driven to water by the hairworm Paragordius tricuspidatus. We characterized host and parasite proteomes during the expression of the water-seeking behaviour. We found that the parasite produces molecules from the Wnt family that may act directly on the development of the central nervous system (CNS). In the head of manipulated cricket, we found differential expression of proteins specifically linked to neurogenesis, circadian rhythm and neurotransmitter activities. We also detected proteins for which the function(s) are still unknown. This proteomics study on the biochemical pathways altered by hairworms has also allowed us to tackle questions of physiological and molecular convergence in the mechanism(s) causing the alteration of orthoptera behaviour. The two hairworm species produce effective molecules acting directly on the CNS of their orthoptera hosts.

Phloem Protein2 (PP2) is a component of the phloem protein bodies found in sieve elements. We describe here the lectin properties of the Arabidopsis (Arabidopsis thaliana) PP2-A1. Using a recombinant protein produced in Escherichia coli, we demonstrated binding to N-acetylglucosamine oligomers. Glycan array screening showed that PP2-A1 also bound to highmannose N-glycans and 9-acyl-N-acetylneuraminic sialic acid. Fluorescence spectroscopy-based titration experiments revealed that PP2-A1 had two classes of binding site for N,N#,N$-triacetylchitotriose, a low-affinity site and a high-affinity site, promoting the formation of protein dimers. A search for structural similarities revealed that PP2-A1 aligned with the Cbm4 and Cbm22-2 carbohydrate-binding modules, leading to the prediction of a b-strand structure for its conserved domain. We investigated whether PP2-A1 interacted with phloem sap glycoproteins by first characterizing abundant Arabidopsis phloem sap proteins by liquid chromatography-tandem mass spectrometry. Then we demonstrated that PP2-A1 bound to several phloem sap proteins and that this binding was not completely abolished by glycosidase treatment. As many plant lectins have insecticidal activity, we also assessed the effect of PP2-A1 on weight gain and survival in aphids. Unlike other mannosebinding lectins, when added to an artificial diet, recombinant PP2-A1 had no insecticidal properties against Acyrthosiphon pisum and Myzus persicae. However, at mid-range concentrations, the protein affected weight gain in insect nymphs. These results indicate the presence in PP2-A1 of several carbohydrate-binding sites, with potentially different functions in the trafficking of endogenous proteins or in interactions with phloem-feeding insects.