Comparing de novo and reference-based transcriptome assembly strategies by applying them to the blood-sucking bug Rhodnius prolixus.

High Throughput Sequencing capabilities have made the process of assembling a transcriptome easier, whether or not there is a reference genome. But the quality of a transcriptome assembly must be good enough to capture the most comprehensive catalog of transcripts and their variations, and to carry out further experiments on transcriptomics. There is currently no consensus on which of the many sequencing technologies and assembly tools are the most effective. Many non-model organisms lack a reference genome to guide the transcriptome assembly. One question, therefore, is whether or not a reference-based genome assembly gives better results than de novo assembly. The blood-sucking insect Rhodnius prolixus-a vector for Chagas disease-has a reference genome. It is therefore a good model on which to compare reference-based and de novo transcriptome assemblies. In this study, we compared de novo and reference-based genome assembly strategies using three datasets (454, Illumina, 454 combined with Illumina) and various assembly software. We developed criteria to compare the resulting assemblies: the size distribution and number of transcripts, the proportion of potentially chimeric transcripts, how complete the assembly was (completeness evaluated both through CEGMA software and R. prolixus proteome fraction retrieved). Moreover, we looked for the presence of two chemosensory gene families (Odorant-Binding Proteins and Chemosensory Proteins) to validate the assembly quality. The reference-based assemblies after genome annotation were clearly better than those generated using de novo strategies alone. Reference-based strategies revealed new transcripts, including new isoforms unpredicted by automatic genome annotation. However, a combination of both de novo and reference-based strategies gave the best result, and allowed us to assemble fragmented transcripts.

New perspectives for population genetics of Chagas'disease vectors in the Northeastern Brazil: isolation of polymorphic microsatellite markers in Triatoma brasiliensis.

Triatoma brasiliensis is the most important Chagas disease vector in semiarid areas of Northeastern Brazil. Although generally found in natural environment, it regularly colonizes or reinvades domiciliary ecotopes. In order to analyse gene flow between habitats, we identified and characterized six microsatellite loci using a microsatellite-enriched genomic library. To assess the usefulness of these microsatellites for genetic studies, we investigated their variability in two natural populations of the T. brasiliensis complex sampled in two Brazilian districts (Corone Jose Dia, South of Piaui state; Curaça, North of Bahia state). The taxonomic status of the samples was checked using cytb sequences. Phylogenetic analyses confirmed that the individuals from Bahia belonged to the T. juazeirensis species. Moreover, primers cross-amplification was tested in 5 Triatoma species and 4 loci successfully amplified in T. infestans and T. guasayana.