Rhodnius prolixus not only has served as a model organism for the study of insect physiology, but also is a major vector of Chagas disease, an illness that affects approximately seven million people worldwide. We sequenced the genome of R. prolixus, generated assembled sequences covering 95% of the genome (∼702 Mb), including 15,456 putative protein-coding genes, and completed comprehensive genomic analyses of this obligate blood-feeding insect. Although immune-deficiency (IMD)-mediated immune responses were observed, R. prolixus putatively lacks key components of the IMD pathway, suggesting a reorganization of the canonical immune signaling network. Although both Toll and IMD effectors controlled intestinal microbiota, neither affected Trypanosoma cruzi, the causal agent of Chagas disease, implying the existence of evasion or tolerance mechanisms. R. prolixus has experienced an extensive loss of selenoprotein genes, with its repertoire reduced to only two proteins, one of which is a selenocysteine-based glutathione peroxidase, the first found in insects. The genome contained actively transcribed, horizontally transferred genes from Wolbachia sp., which showed evidence of codon use evolution toward the insect use pattern. Comparative protein analyses revealed many lineage-specific expansions and putative gene absences in R. prolixus, including tandem expansions of genes related to chemoreception, feeding, and digestion that possibly contributed to the evolution of a blood-feeding lifestyle. The genome assembly and these associated analyses provide critical information on the physiology and evolution of this important vector species and should be instrumental for the development of innovative disease 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.

Tsetse transmit African trypanosomiasis, which is a disease fatal to both humans and animals. A vaccine to protect against this disease does not exist so transmission control relies on eliminating tsetse populations. Although neurotoxic insecticides are the gold standard for insect control, they negatively impact the environment and reduce populations of insect pollinator species. Here we present a promising, environment-friendly alternative to current insecticides that targets the insect tyrosine metabolism pathway. A bloodmeal contains high levels of tyrosine, which is toxic to haematophagous insects if it is not degraded and eliminated. RNA interference (RNAi) of either the first two enzymes in the tyrosine degradation pathway (tyrosine aminotransferase (TAT) and 4-hydroxyphenylpyruvate dioxygenase (HPPD)) was lethal to tsetse. Furthermore, nitisinone (NTBC), an FDA-approved tyrosine catabolism inhibitor, killed tsetse regardless if the drug was orally or topically applied. However, oral administration of NTBC to bumblebees did not affect their survival. Using a novel mathematical model, we show that NTBC could reduce the transmission of African trypanosomiasis in sub-Saharan Africa, thus accelerating current disease elimination programmes.

27 28 Contact during submission: alvaro.acosta-serrano@lstmed.ac.uk 29 30 Keywords: tyrosine catabolism, 4-hydroxyphenylpyruvate dioxygenase (HPPD), 31 hematophagous insects, tsetse, Glossina, African trypanosomiasis, endectocide, 32 nitisinone, Bombus terrestris, selective vector control. 33 Abstract 34 Tsetse transmit African trypanosomiasis, which is a disease fatal to both 35 humans and animals. A vaccine to protect against this disease does not exist so 36 transmission control relies on eliminating tsetse populations. Although 37 neurotoxic insecticides are the gold standard for insect control, they negatively 38 impact the environment and reduce insect pollinator species. Here we present a 39 promising, environment-friendly alternative that targets insect tyrosine 40 metabolism pathway. A bloodmeal contains high levels of tyrosine, which is toxic 41 to haematophagous insects if it is not degraded. RNAi silencing of either the first 42 two enzymes in the tyrosine degradation pathway (TAT and HPPD) was lethal to 43 tsetse. Furthermore, nitisinone (NTBC), an FDA-approved tyrosine catabolism 44 inhibitor, killed tsetse regardless if the drug was orally or topically applied. 45 However, it did not affect bumblebee survival. A mathematical model shows that 46 NTBC could reduce the transmission of African trypanosomiasis in sub-Saharan 47 Africa, thus accelerating current elimination programmes.48 49 50 51 52 53 54 55 56 57 58 Introduction 59 Human African trypanosomiasis (HAT), also known as sleeping sickness, is 60 a parasitic disease caused predominantly by the parasite Trypanosoma brucei 61 gambiense. These parasites are transmitted when infected tsetse flies (Glossina 62 spp.) blood feed. HAT currently affects 3,500 people/year; most patients live in the 63 Democratic Republic of the Congo and an estimated 70 million people remain at 64 risk of infection in sub-Saharan Africa 1 . Tsetse also spread animal African 65 trypanosomiasis (AAT), which causes high mortality rates in livestock and 66 consequently severely limits animal production 2 . As no vaccine for either HAT or 67 AAT exist, and efficacious treatments are often difficult to obtain, tsetse population 68 control remains essential to limit the spread of trypanosomiasis. In the last 69 decades, tsetse control tools such as aerial spraying of insecticides (pyrethroids), 70 visual and odour baited traps, insecticide-treated livestock, live traps, insecticide-71 impregnated traps and targets, and sterile male releases have been employed 3-7 .72 Despite such efforts, AAT and HAT persist and both economic development and 73 public health continue to be jeopardised 8 . Consequently, a novel complementary 74 strategy to control these parasitic diseases is highly desired. Tsetse, like other 75 blood-feeding arthropods, ingest large quantities of blood and often exceed twice 76 their body weight in a single meal 9 . Since more than 80% of blood dry weight 77 consists of proteins, large quantities of amino acids are released in the midgut 78 during bloodmeal di...