Significance: Rhodnius prolixus is a major vector of Chagas disease, an illness caused by Trypanosoma cruzi which affects approximately 7 million people worldwide. This report describes the first genome sequence of a nondipteran insect vector of an important human parasitic disease. This insect has a gene repertoire substantially distinct from dipteran disease vectors, including immune signaling pathways that display major departures from the canonical network. Large gene expansions related to chemoreception, feeding, and digestion have facilitated triatomine adaptation to a blood-feeding lifestyle. This study provides information about the physiology and evolution of an important disease vector that will boost understanding of transmission of a life-threatening parasite and may lead to the development of innovative control methods.

The bloodsucking hemipteran Rhodnius prolixus is a vector of Chagas' disease, which affects 7–8 million people today in Latin America. In contrast to other hematophagous insects, the triatomine gut is compartmentalized into three segments that perform different functions during blood digestion. Here we report analysis of transcriptomes for each of the segments using pyrosequencing technology. Comparison of transcript frequency in digestive libraries with a whole-body library was used to evaluate expression levels. All classes of digestive enzymes were highly expressed, with a predominance of cysteine and aspartic proteinases, the latter showing a significant expansion through gene duplication. Although no protein digestion is known to occur in the anterior midgut (AM), protease transcripts were found, suggesting secretion as pro-enzymes, being possibly activated in the posterior midgut (PM). As expected, genes related to cytoskeleton, protein synthesis apparatus, protein traffic, and secretion were abundantly transcribed. Despite the absence of a chitinous peritrophic membrane in hemipterans - which have instead a lipidic perimicrovillar membrane lining over midgut epithelia - several gut-specific peritrophin transcripts were found, suggesting that these proteins perform functions other than being a structural component of the peritrophic membrane. Among immunity-related transcripts, while lysozymes and lectins were the most highly expressed, several genes belonging to the Toll pathway - found at low levels in the gut of most insects - were identified, contrasting with a low abundance of transcripts from IMD and STAT pathways. Analysis of transcripts related to lipid metabolism indicates that lipids play multiple roles, being a major energy source, a substrate for perimicrovillar membrane formation, and a source for hydrocarbons possibly to produce the wax layer of the hindgut. Transcripts related to amino acid metabolism showed an unanticipated priority for degradation of tyrosine, phenylalanine, and tryptophan. Analysis of transcripts related to signaling pathways suggested a role for MAP kinases, GTPases, and LKBP1/AMP kinases related to control of cell shape and polarity, possibly in connection with regulation of cell survival, response of pathogens and nutrients. Together, our findings present a new view of the triatomine digestive apparatus and will help us understand trypanosome interaction and allow insights into hemipteran metabolic adaptations to a blood-based diet. The bloodsucking bug Rhodnius prolixus is a vector of Chagas' disease, which affects 7–8 million people in Latin America. In contrast to other insects, the digestive tract of Rhodnius has three segments that perform different functions during blood digestion. Here we report analysis of transcriptomes for each of these segments using pyrosequencing technology amounting to several million sequences. Comparison of transcript frequency in digestive libraries with a whole-body library was used to evaluate expression levels, leading to the discovery of several families of enzymes associated with the digestion of proteins, carbohydrates, and lipids, as well as proteins involved in immunity, signal transduction, amino-acid metabolism, and detoxification. Together, our findings present a new view of the triatomine digestive apparatus and will help us understand the mechanism of blood digestion by Rhodnius and its interaction with the agent of Chagas' disease, Trypanosoma cruzi, a parasite that grows within the insect's digestive system.

Chromosomal organization is by and large evolutionarily stable. In Drosophila over 95% of the genes have remained on the same chromosome arm in 12 species that diverged about 63 million years ago. A study of Y chromosomes, however, reveals that only a quarter of D. melanogaster Y-linked genes are also Y-linked in the other 11 sequenced species. And in contrast to the mammalian Y chromosome, a picture of degeneration and gene loss, gene gains on the Drosophila Y chromosome outnumber losses by more than ten to one. In Drosophila more than 95% of the genes have remained on the same chromosome arm in 12 species that diverged 63 Myr ago. This study finds that only a quarter of D. melanogaster Y linked genes are also Y-linked in these 12 species. Gene losses are known to play a major role in the evolution of Y chromosomes, but this study shows that in contrast with the mammalian Y, gene gains play an important role in the evolution of the Drosophila Y chromosome. Chromosomal organization is sufficiently evolutionarily stable that large syntenic blocks of genes can be recognized even between species as distantly related as mammals and puffer fish (450 million years (Myr) of divergence)1,2,3,4,5,6,7. In Diptera, the gene content of the X chromosome and the autosomes is well conserved: in Drosophila more than 95% of the genes have remained on the same chromosome arm in the 12 sequenced species (63 Myr of divergence, traversing 400 Myr of evolution)2,4,6, and the same linkage groups are clearly recognizable in mosquito genomes (260 Myr of divergence)3,5,7. Here we investigate the conservation of Y-linked gene content among the 12 sequenced Drosophila species. We found that only a quarter of the Drosophila melanogaster Y-linked genes (3 out of 12) are Y-linked in all sequenced species, and that most of them (7 out of 12) were acquired less than 63 Myr ago. Hence, whereas the organization of other Drosophila chromosomes traces back to the common ancestor with mosquitoes, the gene content of the D. melanogaster Y chromosome is much younger. Gene losses are known to have an important role in the evolution of Y chromosomes8,9,10, and we indeed found two such cases. However, the rate of gene gain in the Drosophila Y chromosomes investigated is 10.9 times higher than the rate of gene loss (95% confidence interval: 2.3–52.5), indicating a clear tendency of the Y chromosomes to increase in gene content. In contrast with the mammalian Y chromosome, gene gains have a prominent role in the evolution of the Drosophila Y chromosome.

The Y chromosome and other heterochromatic regions present special challenges for genome sequencing and for the annotation of genes. Here we describe two new genes (ARY and WDY) on the Drosophila melanogaster Y, bringing its number of known single-copy genes to 12. WDY may correspond to the fertility factor kl-1.

The heme molecule is the prosthetic group of many hemeproteins involved in essential physiological processes, such as electron transfer, transport of gases, signal transduction, and gene expression modulation. However, heme is a pro-oxidant molecule capable of propagating reactions leading to the generation of reactive oxygen species. The blood-feeding insect Rhodnius prolixus releases enormous amounts of heme during host blood digestion in the midgut lumen when it is exposed to a physiological oxidative challenge. Additionally, this organism produces a hemolymphatic heme-binding protein (RHBP) that transports heme to pericardial cells for detoxification and to growing oocytes for yolk granules and as a source of heme for embryo development. Here, we show that silencing of RHBP expression in female fat bodies reduced total RHBP circulating in the hemolymph, promoting oxidative damage to hemolymphatic proteins. Moreover, RHBP knockdown did not cause reduction in oviposition but led to the production of heme-depleted eggs (white eggs). A lack of RHBP did not alter oocyte fecundation. However, produced white eggs were nonviable. Embryo development cellularization and vitellin yolk protein degradation, processes that normally occur in early stages of embryogenesis, were compromised in white eggs. Total cytochrome c content, cytochrome c oxidase activity, citrate synthase activity, and oxygen consumption, parameters that indicate mitochondrial function, were significantly reduced in white eggs compared with normal dark red eggs. Our results showed that reduction of heme transport from females to growing oocytes by RHBP leads to embryonic mitochondrial dysfunction and impaired embryogenesis.