Tissue-Specific Transcriptomics and Proteomics of a Filarial Nematode and its Wolbachia Endosymbiont
ABSTRACT: We sequenced total RNA from Dirofilaria immitis in order to generate the first tissue-specific gene expression profile of a filarial nematode and its Wolbachia endosymbiont. Examination of transcript levels in 7 different Dirofilaria immitis tissues, in duplicate, using Illumina GAIIx.
Project description:We report the presence of circulating miRNAs released by the filarial nematode Dirofilaria immitis into the host (Canis familiaris) bloodstream. MiRNA deep-sequencing combined with bioinformatics revealed over 200 mature miRNA sequences of potential nematode origin in Dirofilaria immitis-infected dog plasma in two independent analyses
Project description:Female worms (Brugia malayi) were collected from infected jirds treated with 2.5 mg/ml tetracycline in drinking water for 7, 14, or 21 days to eliminate the worm's endosymbiont, Wolbachia.<br>Control age matched female worms were recovered from infected jirds given normal water for drinking.<br>The Filarial Nematode Oligonucleotide Array (version 2) was used in hybridization analyses on cDNA generated from extracted total RNA.<br>Each microarray was hybridized with a mixture of control and experimental cDNA differentially labeled with Cy3 and Cy5 in a flip-dye experiment.<br>Gridding and analysis of images were performed using ScanArray v3.0, each spot defined pixel-by-pixel using a modified Mann-Whitney test, and the resulting values processed with Gene-Spring 7.1 software.
Project description:We report the presence of Onchocerca ochengi and O. volvulus derived small RNAs in bovine nodule fluids and human serum and plasma, respectively. Further comparisons with other related filarial nematodes like Litomosoides sigmodontis and Dirofilaria immitis reveal common and distictive signatures associated to the Onchocerca species. Examination of small RNA content in bovine nodule fluids and human serum/plasma by Next Generation sequencing
Project description:To understand how microRNA are involved in the complex biology of this zoonotic parasite, we describe our initial attempts to characterize small RNA in adult Dirofilaria immitis by using Illumina/Solexa deep-sequencing technology. Examination of 4 different adult Dirofilaria immitis (pooled)
Project description:We characterized the miRNA composition of the nucleus and the cytoplasm of uninfected cells and compared it with the one of cells infected with the endosymbiotic bacterium Wolbachia strain wMelPop-CLA. We found an overall increase of small RNAs between 18 and 28 nucleotides in both cellular compartments in Wolbachia-infected cells and identified specific miRNAs induced and/or suppressed by the Wolbachia infection. We discuss the mechanisms that the cell may use to shuttle miRNAs between the cytoplasm and the nucleus. In addition, we identified piRNAs that changed their abundance in response to Wolbachia infection. The miRNAs and piRNAs identified in this study provide promising leads for investigations into the host-endosymbiont interactions and for better understanding of how Wolbachia manipulates the host miRNA machinery in order to facilitate its persistent replication in infected cells. Examination of small RNA profile in cytoplasmic and nuclear fractions of Aag 2 and Pop cells (Aedes aegypti)
Project description:We report analyses of the types, amounts and stage-dependence of microRNAsfound in D. immitis culture media recovered after incubating 800,000 microfilariae for 6 days, 500 third-stage larvae (L3) and 500 fourth-stage larvae (L4) for 7 days, as well as 40 adult females and 40 adult males for 48 hours, all separately. Overall design: Examination of excretory/secretory miRNAs in 5 different stages of Dirofilaria immitis released in culture media.
Project description:Wolbachia, an endosymbiotic bacterium, is being investigated as a vector control agent in several insect species. Along with the well known classical reproductive parasitism Wolbachia employs against its host to spread within the population, it is emerging that the bacteria can protect the host against pathogens and reduced pathogen transmission. Anopheles mosquitoes, which transmit malaria, have never been found to harbour Wolbachia in nature, and despite numerous transinfection attempts, no stable line has been developed. However recently, two strains of Wolbachia, wAlbB from Aedes albopictus, and wRi from Drosophila simulans were cultured in Anopheles gambiae Sua5B cells. These cell lines provides an amenable system to study Wolbachia-Anopheles interaction in the absence of a stable transinfected line. It has been proposed that the compromised vector competence of Wolbachia infected insects is due to an up regulation of the basal immune state. We therefore completed a genome wide expression profile of Wolbachia infected Anopheles, assessing both wAlbB and wRi infected cells in parallel against uninfected Sua5B cells. Overall design: Two strains of Wolbachia, wRi from Drosophila simulans and wAlbB from Aedes albopictus were transfered into Anopheles gambiae Sua5B cells via the shell vial technique. After over 30 passages, these Wolbachia infected cells lines were then compared, in parallel, to the original uninfected Sua5B cells using Affymetrix microarrays.