Efficacy of oral fluralaner (Bravecto) against Tunga penetrans in dogs: A negative control,randomized field study in an endemic community in Brazil

The sand flea Tunga penetrans is one of the zoonotic agents of tungiasis, a parasitic skin disease of humans and animals. The dog is one of its main reservoirs. This negatively controlled, randomized, double-masked clinical trial evaluated the therapeutic and residual efficacy of fluralaner for treatment of dogs naturally infested with T. penetrans. Sixty-two dogs from an endemically affected community in Brazil were randomly assigned to either receive oral fluralaner (Bravecto chewable tablets) at a dose of 25 to 56 mg fluralaner/kg body weight, or no treatment (31 dogs per group). Dogs were clinically examined using a severity score for acute canine tungiasis (SCADT), parasitological examinations as defined by the Fortaleza classification, and pictures of lesions on days 0 (inclusion and treatment), 7 ± 2, 14 ± 2, 21 ± 2, 28 ± 2, 60 ± 7, 90 ± 7, 120 ± 7 and 150 ± 7. The percentage of parasite-free dogs after treatment was >90% between days 14 and 90 post-treatment with 100% efficacy on study days 21, 28 and 60. Sand flea counts on fluralaner treated dogs were significantly lower (p<0.025) than control dogs on all counts from day 7 to 120. The number of live sand fleas on treated dogs was reduced by > 90% on day 7, > 95% on days 14 and 90, and 100% from day 21 to 60, and with a significant difference between groups from day 7 to 120. From day 7 to day 120, mean SCADT scores were significantly reduced in treated dogs with a mean of 0.10 compared to 1.54 on day 120 in untreated dogs. Therefore, a single oral fluralaner administration is effective for treating and achieving long lasting (> 12 weeks) prevention for tungiasis in dogs.     

Sulfated glycosaminoglycans in two hematophagous arthropod vectors of Chagas disease, Triatoma brasiliensis and Rhodnius prolixus (Hemiptera: Reduviidae)

The characterization of sulfated glycosaminoglycans (GAGs) in hematophagous arthropod vectors in general has been limited, with the exception of the studies in the triatomine Rhodnius prolixus. Heparan sulfate (HS) and chondroitin sulfate (CS) were previously identified and structurally characterized in extracts of whole bodies of fourth instar larvae of R. prolixus. Recently, we showed the expression of these two sulfated GAGs in specific body tissues of adult males and females and in embryos of R. prolixus. In the present work, we identified and compared the sulfated GAG composition in specific tissues of adult insects and in embryos of another triatomine species, Triatoma brasiliensis. Sulfated GAGs were isolated from the fat body, intestinal tract, and the reproductive tracts of adult insects and from embryos. Only HS and CS were found in the tissues analyzed. The present results extend the initial observations on the sulfated GAG composition in R. prolixus by showing that these molecules are widely distributed among internal organs of triatomines. These observations may be useful for future investigations aiming to evaluate the possible implication of these compounds in physiological events that take place in a specific organ(s) in these insects.     

The main sceneries of Chagas disease transmission. The vectors,blood and oral transmissions - A comprehensive review

This review deals with transmission of Trypanosoma cruzi by the most important domestic vectors, blood transfusion and oral intake. Among the vectors, Triatoma infestans, Panstrongylus megistus, Rhodnius prolixus, Triatoma dimidiata, Triatoma brasiliensis, Triatoma pseudomaculata, Triatoma sordida, Triatoma maculata, Panstrongylus geniculatus, Rhodnius ecuadoriensis and Rhodnius pallescens can be highlighted. Transmission of Chagas infection, which has been brought under control in some countries in South and Central America, remains a great challenge, particularly considering that many endemic countries do not have control over blood donors. Even more concerning is the case of non-endemic countries that receive thousands of migrants from endemic areas that carry Chagas disease, such as the United States of America, in North America, Spain, in Europe, Japan, in Asia, and Australia, in Oceania. In the Brazilian Amazon Region, since Shaw et al. (1969) described the first acute cases of the disease caused by oral transmission, hundreds of acute cases of the disease due to oral transmission have been described in that region, which is today considered to be endemic for oral transmission. Several other outbreaks of acute Chagas disease by oral transmission have been described in different states of Brazil and in other South American countries.     

Transovarial transmission of a core virome in the Chagas disease vector Rhodnius prolixus

Triatomine assassin bugs comprise hematophagous insect vectors of Trypanosoma cruzi, the causative agent of Chagas disease. Although the microbiome of these species has been investigated to some extent, only one virus infecting Triatoma infestans has been identified to date. Here, we describe for the first time seven (+) single-strand RNA viruses (RpV1-7) infecting Rhodnius prolixus, a primary vector of Chagas disease in Central and South America. We show that the RpVs belong to the Iflaviridae, Permutotetraviridae and Solemoviridae and are vertically transmitted from the mothers to the progeny via transovarial transmission. Consistent with this, all the RpVs, except RpV2 that is related to the entomopathogenic Slow bee paralysis virus, established persistent infections in our R. prolixus colony. Furthermore, we show that R. prolixus ovaries express 22-nucleotide viral siRNAs (vsiRNAs), but not viral piRNAs, that originate from the processing of dsRNA intermediates during viral replication of the RpVs. Interestingly, the permutotetraviruses and sobemoviruses display shared pools of vsiRNAs that might provide the basis for a cross-immunity system. The vsiRNAs are maternally deposited in the eggs, where they likely contribute to reduce the viral load and protect the developing embryos. Our results unveil for the first time a complex core virome in R. prolixus and begin to shed light on the RNAi-based antiviral defenses in triatomines.     

Amines from vertebrates guide triatomine bugs to resources

Most triatomine bugs (Heteroptera: Reduviidae) are nest-living insects that require vertebrate blood or invertebrate haemolymph to complete their life cycle. Vertebrates accumulate excretory products in or near their nesting sites and we hypothesize that triatomines use emanations from such host wastes when searching for resources. Here we recount how triatomine bugs increase upwind locomotion on a servosphere in response to volatile amine constituents of vertebrate excretions. Fresh chicken faeces is strongly attractive to Rhodnius prolixus nymphs. Ammonia induces attraction and an increase in both speed and total path length by R. prolixus on the servosphere. Whereas ethylamine and dimethylamine attract R. prolixus, Triatoma infestans and Panstrongylus geniculatus, other amine constituents of vertebrate excretions such as isobutylamine and hexylamine induce R. prolixus nymphs to walk faster and for a longer period. These amines are derived from generally occurring metabolites of vertebrates and from gut flora metabolism. We conclude that amines and other products associated with nesting hosts serve as signals for foraging triatomines.     

Machine-learning model led design to experimentally test species thermal limits: The case of kissing bugs (Triatominae)

Species Distribution Modelling (SDM) determines habitat suitability of a species across geographic areas using macro-climatic variables; however, micro-habitats can buffer or exacerbate the influence of macro-climatic variables, requiring links between physiology and species persistence. Experimental approaches linking species physiology to micro-climate are complex, time consuming and expensive. E.g., what combination of exposure time and temperature is important for a species thermal tolerance is difficult to judge a priori. We tackled this problem using an active learning approach that utilized machine learning methods to guide thermal tolerance experimental design for three kissing-bug species: Triatoma infestans, Rhodnius prolixus, and Panstrongylus megistus (Hemiptera: Reduviidae: Triatominae), vectors of the parasite causing Chagas disease. As with other pathogen vectors, triatomines are well known to utilize micro-habitats and the associated shift in microclimate to enhance survival. Using a limited literature-collected dataset, our approach showed that temperature followed by exposure time were the strongest predictors of mortality; species played a minor role, and life stage was the least important. Further, we identified complex but biologically plausible nonlinear interactions between temperature and exposure time in shaping mortality, together setting the potential thermal limits of triatomines. The results from this data led to the design of new experiments with laboratory results that produced novel insights of the effects of temperature and exposure for the triatomines. These results, in turn, can be used to better model micro-climatic envelope for the species. Here we demonstrate the power of an active learning approach to explore experimental space to design laboratory studies testing species thermal limits. Our analytical pipeline can be easily adapted to other systems and we provide code to allow practitioners to perform similar analyses. Not only does our approach have the potential to save time and money: it can also increase our understanding of the links between species physiology and climate, a topic of increasing ecological importance.     

Dislodgement effect of natural semiochemicals released by disturbed triatomines: a possible alternative monitoring tool

The quick detection of domestic and peridomestic triatomines in their environments becomes difficult without the use of dislodgement substances that flush them out from their shelters. At present, tetramethrin 0.2% is being widely used in control programs. Although it is an efficient dislodging agent, its toxicity might affect the health of captured triatomines, of other insects and, to a lesser extent, of other animals, including humans. Here, we tested if semiochemicals released by disturbed adults of Triatoma infestans and/or Rhodnius prolixus can make larvae of the same species exit from their refuges. In a walking olfactometer we found that: 1) larvae of T. infestans were repelled by the odors released by disturbed adults of their own species and of R. prolixus, 2) larvae of R. prolixus did not change their behavior in the presence of odors released by adults of both species, and 3) activity levels were not modulated by these odors in any of both species. Besides, in pseudo‐natural conditions we found an increased flushing‐out activity of larvae of T. infestans when their shelters were sprayed with isobutyric acid or 3‐pentanol, and of larvae of R. prolixus when sprayed with 3‐methyl1butanol. We succeeded in this work to dislodge larvae of triatomines from artificial shelters using natural volatile compounds, allowing the capture of live bugs for further investigations (e.g., xenodiagnosis or genetic studies) and favoring ecological aspects (e.g., minimizing environmental insecticide‐contamination and non‐targeted mortality).     

Chagas disease: control,elimination and eradication. Is it possible?

From an epidemiological point of view, Chagas disease and its reservoirs and vectors can present the following characteristics: (i) enzooty, maintained by wild animals and vectors, with broad occurrence from southern United States of America (USA) to southern Argentina and Chile (42ºN 49ºS), (ii) anthropozoonosis, when man invades the wild ecotope and becomes infected with Trypanosoma cruzi from wild animals or vectors or when the vectors and wild animals, especially marsupials, invade the human domicile and infect man, (iii) zoonosis-amphixenosis and exchanged infection between animals and humans by domestic vectors in endemic areas and (iv) zooanthroponosis, infection that is transmitted from man to animals, by means of domestic vectors, which is the rarest situation in areas endemic for Chagas disease. The characteristics of Chagas disease as an enzooty of wild animals and as an anthropozoonosis are seen most frequently in the Brazilian Amazon and in the Pan-Amazon region as a whole, where there are 33 species of six genera of wild animals: Marsupialia, Chiroptera, Rodentia, Edentata (Xenarthra), Carnivora and Primata and 27 species of triatomines, most of which infected with T. cruzi . These conditions place the resident populations of this area or its visitors - tourists, hunters, fishermen and especially the people whose livelihood involves plant extraction - at risk of being affected by Chagas disease. On the other hand, there has been an exponential increase in the acute cases of Chagas disease in that region through oral transmission of T. cruzi , causing outbreaks of the disease. In four seroepidemiological surveys that were carried out in areas of the microregion of the Negro River, state of Amazonas, in 1991, 1993, 1997 and 2010, we found large numbers of people who were serologically positive for T. cruzi infection. The majority of them and/or their relatives worked in piassava extraction and had come into contact with and were stung by wild triatomines in that area. Finally, a characteristic that is greatly in evidence currently is the migration of people with Chagas disease from endemic areas of Latin America to non-endemic countries. This has created a new dilemma for these countries: the risk of transmission through blood transfusion and the onus of controlling donors and treating migrants with the disease. As an enzooty of wild animals and vectors, and as an anthropozoonosis, Chagas disease cannot be eradicated, but it must be controlled by transmission elimination to man.     

Distribution and natural infection status of synantrophic triatomines (Hemiptera: Reduviidae), vectors of Trypanosoma cruzi,reveals new epidemiological scenarios for chagas disease in the Highlands of Colombia

IntroductionUpdating the distribution and natural infection status of triatomine bugs is critical for planning, prioritizing, and implementing strategies to control Chagas disease (CD), especially after vector reduction programs. After carrying out a control program, the Department of Boyaca contains the highest number of Colombian municipalities certified by PAHO to be free of intradomiciliary transmission of Trypanosoma cruzi by Rhodnius prolixus. The present work describes the spatial distribution, natural infection (NI), and molecular characterization of T. cruzi in synanthropic triatomines from the Department of Boyaca in 2017 and 2018.Materials and methodsAn entomological survey was conducted in 52 municipalities in Boyaca known to have had previous infestations of triatomine bugs. Insects were collected through active searches carried out by technical personnel from the Secretary of Health and community members using Triatomine Collection Stations (PITs-acronym in Spanish). For evaluation of natural infection, triatomines were identified morphologically and grouped in pools of one to five individuals of the same species collected in the same household. DNA derived from the feces of each pool of insects was analyzed by PCR for the presence of T. cruzi using primers flanking the satellite DNA of the parasite. SL-IR primers were used to differentiate TCI from the other DTUs and to identify different genotypes. The distribution of the collected triatomines was analyzed to determine any vector hotspots using spatial recreation.ResultsA total of 670 triatomine bugs was collected, belonging to five species: Triatoma dimidiata (73.2%), Triatoma venosa (16.7%), Panstrongylus geniculatus (5.7%), Rhodnius prolixus (4.4%), and Panstrongylus rufotuberculatus (0.4%), from 29 of the 52 municipalities. In total, 71.6% of the bugs were collected within houses (intradomiciliary) and the rest around the houses (peridomiciliary). Triatoma dimidiata was the most widely distributed species and had the highest natural infection index (37.8%), followed by T. venosa and P. geniculatus. TcI was the only DTU found, with the TcI Dom genotype identified in 80% of positive samples and TcI sylvatic in the other insects. Spatial analysis showed clusters of T. dimidiata and T. venosa in the northeast and southwest regions of Boyaca.ConclusionsAfter some municipalities were certified free of natural transmission within houses (intradomiciliary transmission) of T. cruzi by R. prolixus, T. dimidiata has become the most prevalent vector present, and represents a significant risk of resurgent CD transmission. However, T. venosa, P. geniculatus, and P. rufotuberculatus also contribute to the increased risk of transmission. The presence of residual R. prolixus may undo the successes achieved through vector elimination programs. The molecular and spatial analysis used here allows us to identify areas with an ongoing threat of parasite transmission and improve entomological surveillance strategies.     

Rhodnius prolixus supergene families of enzymes potentially associated with insecticide resistance

Chagas disease or American trypanosomiasis, is a potentially life-threatening illness caused by the protozoan parasite, Trypanosoma cruzi. Once known as an endemic health problem of poor rural populations in Latin American countries, it has now spread worldwide. The parasite is transmitted by triatomine bugs, of which Rhodnius prolixus (Hemiptera, Reduviidae, Triatominae) is one of the vectors and a model organism. This species occurs mainly in Central and South American countries where the disease is endemic. Disease prevention focuses on vector control programs that, in general, rely intensely on insecticide use. However, the massive use of chemical insecticides can lead to resistance. One of the major mechanisms is known as metabolic resistance that is associated with an increase in the expression or activity of detoxification genes. Three of the enzyme families that are involved in this process – carboxylesterases (CCE), glutathione s-transferases (GST) and cytochrome P450s (CYP) – are analyzed in the R. prolixus genome. A similar set of detoxification genes to those of the Hemipteran Acyrthosiphon pisum but smaller than in most dipteran species was found in R. prolixus genome. All major CCE classes (43 genes found) are present but the pheromone/hormone processing class had fewer genes than usual. One main expansion was detected on the detoxification/dietary class. The phosphotriesterase family, recently associated with insecticide resistance, was also represented with one gene. One microsomal GST gene was found and the cytosolic GST gene count (14 genes) is extremely low when compared to the other hemipteran species with sequenced genomes. However, this is similar to Apis mellifera, a species known for its deficit in detoxification genes. In R. prolixus 88 CYP genes were found, with representatives in the four clans (CYP2, CYP3, CYP4 and mitochondrial) usually found in insects. R. prolixus seems to have smaller species-specific expansions of CYP genes than mosquitoes and beetles, among others. The number of R. prolixus CYP genes is similar to the hemipteran Ac. pisum, although with a bigger expansion in CYP3 and CYP4 clans, along with several gene fragments, mostly in CYP4 clan. Eleven founding members of new families were detected, consisting of ten genes in the CYP3 clan and 1 gene in the CYP4 clan. Members of these clans were proposed to have important detoxification roles in insects. The identification of CCE, GST and CYP genes is of utmost importance for directing detoxification studies on triatomines that can help insecticide management strategies in control programs.     

A Model for Chagas Disease with Oral and Congenital Transmission

This work presents a new mathematical model for the domestic transmission of Chagas disease, a parasitic disease affecting humans and other mammals throughout Central and South America. The model takes into account congenital transmission in both humans and domestic mammals as well as oral transmission in domestic mammals. The model has time-dependent coefficients to account for seasonality and consists of four nonlinear differential equations, one of which has a delay, for the populations of vectors, infected vectors, infected humans, and infected mammals in the domestic setting. Computer simulations show that congenital transmission has a modest effect on infection while oral transmission in domestic mammals substantially contributes to the spread of the disease. In particular, oral transmission provides an alternative to vector biting as an infection route for the domestic mammals, who are key to the infection cycle. This may lead to high infection rates in domestic mammals even when the vectors have a low preference for biting them, and ultimately results in high infection levels in humans.     

Retrospective distribution of Trypanosoma cruzi I genotypes in Colombia

Trypanosoma cruzi is the aetiological agent of Chagas disease, which affects approximately eight million people in the Americas. This parasite exhibits genetic variability, with at least six discrete typing units broadly distributed in the American continent. T. cruzi I (TcI) shows remarkable genetic diversity; a genotype linked to human infections and a domestic cycle of transmission have recently been identified, hence, this strain was named TcIDom. The aim of this work was to describe the spatiotemporal distribution of TcI subpopulations across humans, insect vectors and mammalian reservoirs in Colombia by means of molecular typing targeting the spliced leader intergenic region of mini-exon gene. We analysed 101 TcI isolates and observed a distribution of sylvatic TcI in 70% and TcIDom in 30%. In humans, the ratio was sylvatic TcI in 60% and TcIDom in 40%. In mammal reservoirs, the distribution corresponded to sylvatic TcI in 96% and TcIDom in 4%. Among insect vectors, sylvatic TcI was observed in 48% and TcIDom in 52%. In conclusion, the circulation of TcIDom is emerging in Colombia and this genotype is still adapting to the domestic cycle of transmission. The epidemiological and clinical implications of these findings are discussed herein.     

Metabolic Signatures of Triatomine Vectors of Trypanosoma cruzi Unveiled by Metabolomics

Chagas disease is a trypanosomiasis whose causative agent is the protozoan parasite Trypanosoma cruzi, which is transmitted to humans by hematophagous insects known as triatomines and affects a large proportion of South America. The digestive tract of the insect vectors in which T. cruzi develops constitutes a dynamic environment that affects the development of the parasite. Thus, we set out to investigate the chemical composition of the triatomine intestinal tract through a metabolomics approach. We performed Direct Infusion Fourier Transform Ion Cyclotron Resonance Mass Spectrometry on fecal samples of three triatomine species (Rhodnius prolixus, Triatoma infestans, Panstrongylus megistus) fed with rabbit blood. We then identified groups of metabolites whose frequencies were either uniform in all species or enriched in each of them. By querying the Human Metabolome Database, we obtained putative identities of the metabolites of interest. We found that a core group of metabolites with uniform frequencies in all species represented approximately 80% of the molecules detected, whereas the other 20% varied among triatomine species. The uniform core was composed of metabolites of various categories, including fatty acids, steroids, glycerolipids, nucleotides, sugars, and others. Nevertheless, the metabolic fingerprint of triatomine feces differs depending on the species considered. The variable core was mainly composed of prenol lipids, amino acids, glycerolipids, steroids, phenols, fatty acids and derivatives, benzoic acid and derivatives, flavonoids, glycerophospholipids, benzopyrans, and quinolines. Triatomine feces constitute a rich and varied chemical medium whose constituents are likely to affect T. cruzi development and infectivity. The complexity of the fecal metabolome of triatomines suggests that it may affect triatomine vector competence for specific T. cruzi strains. Knowledge of the chemical environment of T. cruzi in its invertebrate host is likely to generate new ways to understand the factors influencing parasite proliferation as well as methods to control Chagas disease.     

Impact of a community-based bug-hunting campaign on Chagas disease control: a case study in the department of Jalapa,Guatemala

Chagas disease control requires an innovative approach to strengthen community participation in vector surveillance. This paper presents a case study of a community-based bug-hunting campaign in Guatemala. The campaign was implemented in 2007 in the following three stages: (i) a four week preparation stage to promote bug-hunting, (ii) a one week bug-hunting stage to capture and collect bugs and (iii) a 10 week follow-up stage to analyse the bugs and spray insecticide. A total of 2,845 bugs were reported, of which 7% were Triatominae vectors, such as Rhodnius prolixus and Triatoma dimidiata. The bug-hunting campaign detected a five-six-fold higher amount of vectors in one week than traditional community-based surveillance detects in one year. The bug-hunting campaign effectively detected vectors during a short period, provided information to update the vector infestation map and increased community and political awareness regarding Chagas disease. This approach could be recommended as an effective and feasible strategy to strengthen vector surveillance on a larger scale.     

Risk factors associated with Trypanosoma cruzi exposure in domestic dogs from a rural community in Panama

Chagas disease, caused by Trypanosoma cruzi infection, is a zoonosis of humans, wild and domestic mammals, including dogs. In Panama, the main T. cruzi vector is hodnius pallescens, a triatomine bug whose main natural habitat is the royal palm, Attalea butyracea. In this paper, we present results from three T. cruzi serological tests (immunochromatographic dipstick, indirect immunofluorescence and ELISA) performed in 51 dogs from 24 houses in Trinidad de Las Minas, western Panama. We found that nine dogs were seropositive (17.6% prevalence). Dogs were 1.6 times more likely to become T. cruzi seropositive with each year of age and 11.6 times if royal palms where present in the peridomiciliary area of the dog''s household or its two nearest neighbours. Mouse-baited-adhesive traps were employed to evaluate 12 peridomestic royal palms. All palms were found infested with R. pallescens with an average of 25.50 triatomines captured per palm. Of 35 adult bugs analysed, 88.6% showed protozoa flagellates in their intestinal contents. In addition, dogs were five times more likely to be infected by the presence of an additional domestic animal species in the dog''s peridomiciliary environment. Our results suggest that interventions focused on royal palms might reduce the exposure to T. cruzi infection.     

Cultivation-independent methods reveal differences among bacterial gut microbiota in triatomine vectors of Chagas disease

Background

Chagas disease is a trypanosomiasis whose agent is the protozoan parasite Trypanosoma cruzi, which is transmitted to humans by hematophagous bugs known as triatomines. Even though insecticide treatments allow effective control of these bugs in most Latin American countries where Chagas disease is endemic, the disease still affects a large proportion of the population of South America. The features of the disease in humans have been extensively studied, and the genome of the parasite has been sequenced, but no effective drug is yet available to treat Chagas disease. The digestive tract of the insect vectors in which T. cruzi develops has been much less well investigated than blood from its human hosts and constitutes a dynamic environment with very different conditions. Thus, we investigated the composition of the predominant bacterial species of the microbiota in insect vectors from Rhodnius, Triatoma, Panstrongylus and Dipetalogaster genera.

Methodology/Principal Findings

Microbiota of triatomine guts were investigated using cultivation-independent methods, i.e., phylogenetic analysis of 16s rDNA using denaturing gradient gel electrophoresis (DGGE) and cloned-based sequencing. The Chao index showed that the diversity of bacterial species in triatomine guts is low, comprising fewer than 20 predominant species, and that these species vary between insect species. The analyses showed that Serratia predominates in Rhodnius, Arsenophonus predominates in Triatoma and Panstrongylus, while Candidatus Rohrkolberia predominates in Dipetalogaster.

Conclusions/Significance

The microbiota of triatomine guts represents one of the factors that may interfere with T. cruzi transmission and virulence in humans. The knowledge of its composition according to insect species is important for designing measures of biological control for T. cruzi. We found that the predominant species of the bacterial microbiota in triatomines form a group of low complexity whose structure differs according to the vector genus.     

New scenarios of Trypanosoma cruzi transmission in the Orinoco region of Colombia

Rhodnius prolixus, a blood-sucking triatomine with domiciliary anthropophilic habits, is the main vector of Chagas disease. The current paradigm of Trypanosoma cruzi transmission in Columbia includes a sylvatic and domiciliary cycle co-existing with domestic and sylvatic populations of reservoirs. The aim of this study is to evaluate the population densities and relative abundance of triatomines and mammals that may be involved in the sylvatic cycle of Chagas disease to clarify the epidemiological scenario in an endemic area in the province of Casanare. Insect vectors on Attalea butyracea palms were captured using both manual searches and bait traps. The capture of mammals was performed using Sherman and Tomahawk traps. We report an infestation index of 88.5% in 148 palms and an index of T. cruzi natural infection of 60.2% in 269 dissected insects and 11.9% in 160 captured mammals. High population densities of triatomines were observed in the sylvatic environment and there was a high relative abundance of reservoirs in the area, suggesting a stable enzootic cycle. We found no evidence of insect domiciliation. Taken together, these observations suggest that eco-epidemiological factors shape the transmission dynamics of T. cruzi, creating diverse scenarios of disease transmission.     

Intravital microscopy and image analysis of Rhodnius prolixus (Hemiptera: Reduviidae) hematophagy: The challenge of blood intake from mouse skin

Hematophagous insects transmit many of the most dangerous parasitic diseases. The transmission usually occurs during hematophagy or just after as this is when the vector and the host are in contact. The contact time is determined by the feeding performance of the insect in each host. In triatomines, feeding performance interferes with both their life cycle and the vectorial competence to transmit the hemoflagellate Trypanosoma cruzi. Triatomine bugs are vessel feeders, obtaining their blood meals directly from the vessels (venules or arterioles) of their vertebrate hosts. The host blood intake rate is not constant during the feeding, and the sucking frequency of triatomines tends to be higher and to contain fewer interruptions in pigeons than in mice. To identify the difficulties encountered by triatomine bugs in obtaining blood meals from mouse skin, we used intravital microscopy techniques associated with electromyograms of the cibarial pump. To monitor the vibration of the cannulated vessels and the blood flow through the head of the insect during the engorgement phase, we introduced a novel method for image analysis. The mean number of vessels used during a Rhodnius prolixus blood meal was 3.4 ± 1.2, and the insects fed more in venules (63%) than in arterioles (37%). An important increase in vascular permeability was observed throughout the feeding. Platelet aggregation, rolling and leukocyte adherence were analyzed on the venular endothelium, showing remarkable increases for some time following the R. prolixus feeding. The reduction in sucking frequency that was observed during insect feeding was likely due to the increased cibarial pump filling time. The monitoring of the vessel wall pulsation also permitted the registration of regurgitation-like movements during blood pumping, with these movements being recorded mostly during the second half of the feeding. The evaluation of blood flow through the head of the insect suggested that the regurgitation-like movements were not true regurgitations and were caused by abrupt difficulties in the function of the cibarial pump. The role of the platelet plugs and the changes in blood viscosity at the R. prolixus feeding site are discussed. The method introduced in the present study to analyze the images brings new insights into the interaction between hematophagous vectors and their hosts, reinforcing the importance of insect saliva throughout the feeding process.     

Decoding the Ubiquitin-Mediated Pathway of Arthropod Disease Vectors

Protein regulation by ubiquitin has been extensively described in model organisms. However, characterization of the ubiquitin machinery in disease vectors remains mostly unknown. This fundamental gap in knowledge presents a concern because new therapeutics are needed to control vector-borne diseases, and targeting the ubiquitin machinery as a means for disease intervention has been already adopted in the clinic. In this study, we employed a bioinformatics approach to uncover the ubiquitin-mediated pathway in the genomes of Anopheles gambiae, Aedes aegypti, Culex quinquefasciatus, Ixodes scapularis, Pediculus humanus and Rhodnius prolixus. We observed that (1) disease vectors encode a lower percentage of ubiquitin-related genes when compared to Drosophila melanogaster, Mus musculus and Homo sapiens but not Saccharomyces cerevisiae; (2) overall, there are more proteins categorized as E3 ubiquitin ligases when compared to E2-conjugating or E1-activating enzymes; (3) the ubiquitin machinery within the three mosquito genomes is highly similar; (4) ubiquitin genes are more than doubled in the Chagas disease vector (R. prolixus) when compared to other arthropod vectors; (5) the deer tick I. scapularis and the body louse (P. humanus) genomes carry low numbers of E1-activating enzymes and HECT-type E3 ubiquitin ligases; (6) R. prolixus have low numbers of RING-type E3 ubiquitin ligases; and (7) C. quinquefasciatus present elevated numbers of predicted F-box E3 ubiquitin ligases, JAB and UCH deubiquitinases. Taken together, these findings provide novel opportunities to study the interaction between a pathogen and an arthropod vector.     

The chemical basis of mate recognition in two parasitoid wasp species of the genus Nasonia

Triatomines (Hemiptera: Reduviidae) are vectors of Trypanosoma cruzi Chagas, the etiological agent of Chagas's disease. They display pre‐adult development delay – that is, a development time much longer than on average – which usually has been considered as a maladaptive trait. However, this hypothesis has not been tested. We carried out an experiment under controlled laboratory conditions to (1) test whether a development delay exists in the fifth nymphal stage of Rhodnius prolixus Stål (Hemiptera: Reduviidae, Rhodniini), and (2) measure any fitness cost related to such delay by estimating the relationship between individual development time and other life‐history traits. We analyzed the development time with various continuous statistical distributions (normal, log‐normal, Weibull, gamma, Pareto, Burr, and log‐logistic). Using goodness‐of‐fit tests, the best fit was obtained with asymmetrical distributions, with the Burr distribution showing the best fit to the data. We concluded that a development delay exists in stage five of R. prolixus without fitness cost. The combination of our results and previous work suggests that such a delay could be viewed as an adaptive response to environmental stochasticity and/or density‐dependence rather than as a maladaptive trait. We propose further investigations to provide a conclusive test of adaptive delay in triatomines.