Project description:With its 2.5 Mb DNA genome packed in amphora-shaped particles of bacterium-like dimension (1.2 µm in length, 0.5 µm in diameter), the Acanthamoeba-infecting Pandoravirus salinus remained the most spectacular and intriguing virus since its description in 2013. Following its isolation from shallow marine sediment off the coast of central Chile, that of its relative Pandoravirus dulcis from a fresh water pond near Melbourne, Australia, suggested that they were the first representatives of an emerging worldwide-distributed family of giant viruses. This was further suggested when P. inopinatum discovered in Germany, was sequenced in 2015. We now report the isolation and genome sequencing of three new strains (P. quercus, P.neocaledonia, P. macleodensis) from France, New Caledonia, and Australia. Using a combination of transcriptomic, proteomic, and bioinformatic analyses, we found that these six viruses share enough distinctive features to justify their classification in a new family, the Pandoraviridae, distinct from that of other large DNA viruses.
Project description:Proteome analysis of a novel type of virus. The virus was grown in A. castellanii amobea before being purified. Viral particules were enriched after centrifugation and proteins solubilised by SDS before being stacked in the top of a SDS-PGE gel. After in-gel digestion, resulting peptides were injected for a 120min nanoLC-MS/MS analysis using an Ultimate U3000 system and a LTQ-Orbitrap Velos pro hybrid mass spectrometer (Top 20).Data processing and bioinformatics: Data were processed automatically using Mascot Daemon software (version 2.3.2, Matrix Science). Concomitant searches against Pandoravirus and A. castellanii protein sequence databanks as well as classical contaminants database and the corresponding reversed databases were performed using Mascot (version 2.4). ESI-TRAP was chosen as the instrument, trypsin/P as the enzyme and 2 missed cleavage allowed. Precursor and fragment mass error tolerances were set respectively at 10 ppm and 0.6 Da. Peptide modifications allowed during the search were: carbamidomethyl (C, fixes) acetyl (N-ter, variable), oxidation (M, variable) and deamidation (NQ, variable). The IRMa software (Dupierris et al., Bioinformatics, 2009, 25:1980-1, version 1.30.4) was used to filter the results: selection of rank 1 peptides, peptide identification FDR < 1% (as calculated by employing the reverse database strategy), and minimum of 1 specific peptide per identified protein group.
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:Pandoravirus aubagnensis was isolated from freshwater collected from “Manon des Sources” in Marseille city,France. By co-culture on Acanthamoeba castellanii. Pandoravirus belohorizontensis was isolated from soil samples collected from Belo Horizonte city in Brazil, by co-culture on Acanthamoeba castellanii.