2b-RAD genotyping for population genomic studies of Chagas disease vectors: Rhodnius ecuadoriensis in Ecuador

Background: Rhodnius ecuadoriensis is the main triatomine vector of Chagas disease, American trypanosomiasis, in Southern Ecuador and Northern Peru. Genomic approaches and next generation sequencing technologies have become powerful tools for investigating population diversity and structure which is a key consideration for vector control. Here we assess the effectiveness of three different 2b restriction site-associated DNA (2b-RAD) genotyping strategies in R. ecuadoriensis to provide sufficient genomic resolution to tease apart microevolutionary processes and undertake some pilot population genomic analyses. Methodology/Principal findings: The 2b-RAD protocol was carried out in-house at a non-specialized laboratory using 20 R. ecuadoriensis adults collected from the central coast and southern Andean region of Ecuador, from June 2006 to July 2013. 2b-RAD sequencing data was performed on an Illumina MiSeq instrument and analyzed with the STACKS de novo pipeline for loci assembly and Single Nucleotide Polymorphism (SNP) discovery. Preliminary population genomic analyses (global AMOVA and Bayesian clustering) were implemented. Our results showed that the 2b-RAD genotyping protocol is effective for R. ecuadoriensis and likely for other triatomine species. However, only BcgI and CspCI restriction enzymes provided a number of markers suitable for population genomic analysis at the read depth we generated. Our preliminary genomic analyses detected a signal of genetic structuring across the study area. Conclusions/Significance: Our findings suggest that 2b-RAD genotyping is both a cost effective and methodologically simple approach for generating high resolution genomic data for Chagas disease vectors with the power to distinguish between different vector populations at epidemiologically relevant scales. As such, 2b-RAD represents a powerful tool in the hands of medical entomologists with limited access to specialized molecular biological equipment. Author summary: Understanding Chagas disease vector (triatomine) population dispersal is key for the design of control measures tailored for the epidemiological situation of a particular region. In Ecuador, Rhodnius ecuadoriensis is a cause of concern for Chagas disease transmission, since it is widely distributed from the central coast to southern Ecuador. Here, a genome-wide sequencing (2b-RAD) approach was performed in 20 specimens from four communities from Manabí (central coast) and Loja (southern) provinces of Ecuador, and the effectiveness of three type IIB restriction enzymes was assessed. The findings of this study show that this genotyping methodology is cost effective in R. ecuadoriensis and likely in other triatomine species. In addition, preliminary population genomic analysis results detected a signal of population structure among geographically distinct communities and genetic variability within communities. As such, 2b-RAD shows significant promise as a relatively low-tech solution for determination of vector population genomics, dynamics, and spread

Bats, Trypanosomes, and Triatomines in Ecuador: New Insights into the Diversity, Transmission, and Origins of Trypanosoma cruzi and Chagas Disease

The generalist parasite Trypanosoma cruzi has two phylogenetic lineages associated almost exclusively with bats—Trypanosoma cruzi Tcbat and the subspecies T. c. marinkellei. We present new information on the genetic variation, geographic distribution, host associations, and potential vectors of these lineages. We conducted field surveys of bats and triatomines in southern Ecuador, a country endemic for Chagas disease, and screened for trypanosomes by microscopy and PCR. We identified parasites at species and genotype levels through phylogenetic approaches based on 18S ribosomal RNA (18S rRNA) and cytochrome b (cytb) genes and conducted a comparison of nucleotide diversity of the cytb gene. We document for the first time T. cruzi Tcbat and T. c. marinkellei in Ecuador, expanding their distribution in South America to the western side of the Andes. In addition, we found the triatomines Cavernicola pilosa and Triatoma dispar sharing shelters with bats. The comparisons of nucleotide diversity revealed a higher diversity for T. c. marinkellei than any of the T. c. cruzi genotypes associated with Chagas disease. Findings from this study increased both the number of host species and known geographical ranges of both parasites and suggest potential vectors for these two trypanosomes associated with bats in rural areas of southern Ecuador. The higher nucleotide diversity of T. c. marinkellei supports a long evolutionary relationship between T. cruzi and bats, implying that bats are the original hosts of this important parasite

Limitations of selective deltamethrin application for triatomine control in central coastal Ecuador

<p>Abstract</p> <p>Background</p> <p>This year-long study evaluated the effectiveness of a strategy involving selective deltamethrin spraying and community education for control of Chagas disease vectors in domestic units located in rural communities of coastal Ecuador.</p> <p>Results</p> <p>Surveys for triatomines revealed peridomestic infestation with <it>Rhodnius ecuadoriensis </it>and <it>Panstrongylus howardi</it>, with infestation indices remaining high during the study (13%, 17%, and 10%, at initial, 6-month, and 12-month visits, respectively), which indicates a limitation of this strategy for triatomine population control. Infestation was found 6 and 12 months after spraying with deltamethrin. In addition, a large number of previously vector-free domestic units also were found infested at the 6- and 12-month surveys, which indicates new infestations by sylvatic triatomines. The predominance of young nymphs and adults suggests new infestation events, likely from sylvatic foci. In addition, infection with <it>Trypanosoma cruzi </it>was found in 65%, 21% and 29% at initial, 6-month and 12-month visits, respectively. All parasites isolated (n = 20) were identified as TcI.</p> <p>Conclusion</p> <p>New vector control strategies need to be devised and evaluated for reduction of <it>T. cruzi </it>transmission in this region.</p

Population genomics and geographic dispersal in Chagas disease vectors: landscape drivers and evidence of possible adaptation to the domestic setting

Accurate prediction of vectors dispersal, as well as identification of adaptations that allow blood-feeding vectors to thrive in built environments, are a basis for effective disease control. Here we adopted a landscape genomics approach to assay gene flow, possible local adaptation, and drivers of population structure in Rhodnius ecuadoriensis, an important vector of Chagas disease. We used a reduced-representation sequencing technique (2b-RADseq) to obtain 2,552 SNP markers across 272 R. ecuadoriensis samples from 25 collection sites in southern Ecuador. Evidence of high and directional gene flow between seven wild and domestic population pairs across our study site indicates insecticide-based control will be hindered by repeated re-infestation of houses from the forest. Preliminary genome scans across multiple population pairs revealed shared outlier loci potentially consistent with local adaptation to the domestic setting, which we mapped to genes involved with embryogenesis and saliva production. Landscape genomic models showed elevation is a key barrier to R. ecuadoriensis dispersal. Together our results shed early light on the genomic adaptation in triatomine vectors and facilitate vector control by predicting that spatially-targeted, proactive interventions would be more efficacious than current, reactive approaches

Phenotypic Variability of Rhodnius ecuadoriensis Populations at the Ecuadorian Central and Southern Andean Region

Rhodnius ecuadoriensis is an important vector of Chagas disease in Ecuador. Whereas only sylvatic and peridomestic populations are common in Manab province, this species occupies domestic, peridomestic, and sylvatic habitats in Loja province where high reinfestation of houses was observed. To explore the existence of phenetic changes linked to the domiciliation of the species, this study set out to analyze the wing and antennal phenotypes of R. ecuadoriensis in these two provinces where the vector presents different affinity for domestic habitats. The antennal phenotype and the wing size and shape distinguish the two geographical populations of R. ecuadoriensis. In Manab, sylvatic and peridomestic specimens were very similar. In Loja, sylvatic and nonsylvatic (domestic and peridomestic) populations showed distinctive characteristics. Remarkable sexual dimorphism of wing and antenna, exclusive of domestic specimens, and high metric disparity in the wing shape of the domestic females point out the existence of a particular situation in this habitat. The results of this phenotypic analysis and previous evidence of behavioral differences support the hypothesis of disruptive selection acting upon R. ecuadoriensis populations.Fil: Villacís, Anita G.. Pontificia Universidad Católica del Ecuador; EcuadorFil: Grijalva, Mario J.. Pontificia Universidad Católica del Ecuador; EcuadorFil: Catala, Silvia Susana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Regional de Investigaciones Científicas y Transferencia Tecnológica de La Rioja. - Universidad Nacional de La Rioja. Centro Regional de Investigaciones Científicas y Transferencia Tecnológica de La Rioja. - Universidad Nacional de Catamarca. Centro Regional de Investigaciones Científicas y Transferencia Tecnológica de La Rioja. - Secretaría de Industria y Minería. Servicio Geológico Minero Argentino. Centro Regional de Investigaciones Científicas y Transferencia Tecnológica de La Rioja. - Provincia de La Rioja. Centro Regional de Investigaciones Científicas y Transferencia Tecnológica de La Rioja; Argentin

Determinants of intentions to prevent triatomine infestation based on the health belief model: An application in rural Southern Ecuador

Introduction Control of triatomine infestation is a key strategy for the prevention of Chagas disease (CD). To promote this strategy, it is important to know which antecedents to behavioral change are the best to emphasize when promoting prevention. Objective The aim of this study was to determine predictors for intention to prevent home infestation based on the Health Belief Model (HBM), a commonly used health intervention planning theory. Materials & methods A cross-sectional study was conducted with 112 heads of household in six communities with endemic and high rates of triatomine infestation in Loja province, Ecuador. The data was collected by a questionnaire including perceived severity, susceptibility, benefits to action, barriers to action, and self-efficacy. These data were also used to predict actual infestation of homes. Results Community members reported strong intentions to prevent home infestation. HBM constructs predicted about 14% of the observed variance in intentions. Perceived susceptibility and severity did not predict behavioral intention well; perceived barriers to small-scale action that reduce likelihood of infestation and self-efficacy in participating in surveillance systems did. Self-efficacy and perception of barriers were equally powerful predictors. The HBM constructs, however, did not predict well actual infestation. Conclusion The findings supported the HBM as a way to predict intentions to prevent infestation of the home by triatomine bugs. The findings highlight that messages emphasizing self-efficacy in participating in surveillance systems and overcoming barriers to small-scale action that reduce likelihood of infestation, rather than a focus on risk, should be central messages when designing and implementing educational interventions for CD. The gap between behavioral intention and actual infestation reveals the need to assess home practices and their actual efficacy to fully enact and apply the HBM

Absence of domestic triatomine colonies in an area of the coastal region of Ecuador where Chagas disease is endemic

Rhodnius ecuadoriensis is considered the second most important vector of Chagas disease in Ecuador. It is distributed across six of the 24 provinces and occupies intradomiciliary, peridomiciliary and sylvatic habitats. This study was conducted in six communities within the coastal province of Guayas. Triatomine searches were conducted in domestic and peridomestic habitats and bird nests using manual searches, live-bait traps and sensor boxes. Synantrhopic mammals were captured in the domestic and peridomestic habitats. Household searches (n = 429) and randomly placed sensor boxes (n = 360) produced no live triatomine adults or nymphs. In contrast, eight nymphs were found in two out of six searched Campylorhynchus fasciatus (Troglodytidae) nests. Finally, Trypanosoma cruzi DNA was amplified from the blood of 10% of the 115 examined mammals. Environmental changes in land use (intensive rice farming), mosquito control interventions and lack of intradomestic adaptation are suggested among the possible reasons for the lack of domestic triatomine colonies

Distribution of triatomine species in domestic and peridomestic environments in central coastal Ecuador.

Although the central coast of the Ecuador is considered endemic for Chagas disease, few studies have focused on determining the risk of transmission in this region. In this study we describe the triatomine household infestation in Manabí province (Central Coast region), determine the rate of Trypanosoma cruzi infection and study the risk factors associated with infestation by Rhodnius ecuadoriensis.An entomological survey found three triatomine species (Rhodnius ecuadoriensis, Panstrongylus rufotuberculatus and P. howardi) infesting domiciles in 47.4% of the 78 communities visited (total infestation rate of 4.5%). Four percent of domiciles were infested, and nymphs were observed in 77% of those domiciles. The three species were found in altitudes below 500 masl and in all ecological zones except cloud forest. Within the domicile, we found the three species mostly in bedrooms. Rhodnius ecuadoriensis and P. rufotuberculatus were abundant in bird nests, including chicken coops and P. howardi associated with rats in piles of bricks, in the peridomicile. Triatomine infestation was characterized by high rates of colonization, especially in peridomicile. Flagelates infection was detected in only 12% of the samples by microscopy and Trypanosoma cruzi infection in 42% of the examined triatomines by PCR (n = 372). The most important risk factors for house infestation by R. ecuadoriensis were ecological zone (w = 0.99) and presence of chickens (w = 0.96). Determinants of secondary importance were reporting no insecticide applications over the last twelve months (w = 0.86) and dirt floor (w = 0.70). On the other hand, wood as wall material was a protective factor (w = 0.85).According the results, approximately 571,000 people would be at high risk for T. cruzi infection in Manabí province. A multidisciplinary approximation and the adhesion to a periodic integrated vector management (IVM) program are essential to guarantee sustainable preventive and control strategies for Chagas disease in this region