Project description:Primary objectives: The primary objective is to investigate circulating tumor DNA (ctDNA) via deep sequencing for mutation detection and by whole genome sequencing for copy number analyses before start (baseline) with regorafenib and at defined time points during administration of regorafenib for treatment efficacy in colorectal cancer patients in terms of overall survival (OS).
Primary endpoints: circulating tumor DNA (ctDNA) via deep sequencing for mutation detection and by whole genome sequencing for copy number analyses before start (baseline) with regorafenib and at defined time points during administration of regorafenib for treatment efficacy in colorectal cancer patients in terms of overall survival (OS).
Project description:The grape mealybug Pseudococcus maritimus (Ehrhorn, 1900) (Hemiptera: Pseudococcidae) is a significant pest of grapevines (Vitis spp.) and a vector of disease-causing grape viruses, linked to its feeding on phloem sap. The management of this pest is constrained by the lack of naturally occurring resistance traits in Vitis. Here, we obtained proof of concept that RNA interference (RNAi) using double-stranded RNA (dsRNA) molecules against essential genes for phloem sap feeding can depress insect survival. The genes of interest code for an aquaporin (AQP) and a sucrase (SUC) that are required for osmoregulation in related phloem sap-feeding hemipteran insects (aphids and whiteflies). In parallel, we investigated the grape mealybug genes coding non-specific nucleases (NUC), which reduce RNAi efficacy by degrading administered dsRNA. Homologs of AQP and SUC with experimentally validated function in aphids, together with NUC, were identified in the published transcriptome of the citrus mealybug Planococcus citri by phylogenetic analysis, and sequences of the candidate genes were obtained for Ps. maritimus by PCR with degenerate primers. Using this first sequence information for Ps. maritimus, dsRNA was prepared and administered to the insects via an artificial diet. The treatment comprising dsRNA against AQP, SUC and NUC significantly increased insect mortality over three days, relative to dsRNA-free controls. The dsRNA constructs for AQP and NUC were predicted, from sequence analysis to have some activity against other mealybugs, but none of the three dsRNA constructs have predicted activity against aphids. This study provides the basis to develop in planta RNAi strategies against Ps. maritimus and other mealybug pests of grapevines.
Project description:Pseudococcus comstocki (Hemiptera: Pseudococcidae) is a mealybug species native to Eastern Asia and present as an invasive pest in northern Italy and southern France since the start of the century. It infests apple and pear trees, grapevines and some ornamental trees. Biocontrol programmes against this pest proved successful in central Asia and North America in the second half of the 20th century. In this study, we investigated possible biocontrol agents against P. comstocki, with the aim of developing a biocontrol programme in France. We carried out systematic DNA-barcoding at each step in the search for a specialist parasitoid. First we characterised the French target populations of P. comstocki. We then identified the parasitoids attacking P. comstocki in France. Finally, we searched for foreign mealybug populations identified a priori as P. comstocki and surveyed their hymenopteran parasitoids. Three mealybug species (P. comstocki, P. viburni and P. cryptus) were identified during the survey, together with at least 16 different parasitoid taxa. We selected candidate biological control agent populations for use against P. comstocki in France, from the species Allotropa burrelli (Hymenoptera: Platygastridae) and Acerophagus malinus (Hymenoptera: Encyrtidae). The coupling of molecular and morphological characterisation for both pests and natural enemies facilitated the programme development and the rejection of unsuitable or generalist parasitoids.