From molecules to ecosystems: Insights into a network of interactions for a Chagas disease outbreak using Triatoma brasiliensis as natural samplers.

Exploring the dynamics of disease transmission involves an understanding of complex interactions within the eco-epidemiologic framework. In the context of Chagas disease (CD), elements are mainly represented by the interactions among the pathogen, insect vector, host, humans and the environment. We performed quantitative and qualitative analyses on a dataset derived from 98 Triatoma brasiliensis infected by trypanosomatids, which were linked to a CD outbreak in the semi-arid region of northeastern Brazil. We extracted invertebrate-derived DNA (iDNA) from these insects, comprising 18 populations around the outbreak area, each indicative of various strata of anthropogenic influence. Food source (FS) diversity, representing potential parasite reservoirs, was determined through mitochondrial gene (cyt b) sequencing of vertebrates, and parasite genotyping was accessed using fluorescent amplified fragment barcodes (FFLB) of trypanosomatids. We also assessed the residents' awareness of breeding sites for CD vectors in the inspected houses. The quantification of Trypanosoma cruzi was estimated via real-time PCR and is denominated here as the average parasite load (PL) per insect (T. cruzi/intestinal unit). We aimed to address vector-parasite-host-environment interactions that were discussed based on their significance among the components. Notably, among the significant interactions, we observed that the PL in the insects was significantly influenced by FS. Infected insects that fed on the classic reservoir, Didelphis albiventris, and Galea spixii exhibited higher PLs, compared to those that fed on Kerodon rupestris (p < 0.04)-a primary host. While D. albiventris is already recognized as a synanthropic species, we propose that G. spixii may also be undergoing a synanthropic process. Conversely, domestic cats are frequently identified as FS in infected insects from the sylvatic environment, suggesting a possible change in their behavior towards a wild state. Therefore, we propose that neglected anthropogenic actions have facilitated the reciprocal (sylvatic-peridomestic) circulation of T. cruzi-especially noted for TcI because it was predominant in insects found in peridomestic environments. Residents are often unaware of the existence of insect breeding grounds near their homes, particularly when it involves the storage of materials without planning for use, such as piles of tiles, bricks and wood. Although indirect inferences about the interaction among vector-parasite-host-environment are still incipient, we highlight the potential use of vectors as natural samplers of biological and ecological components in transmitting the disease.

The risk of oral transmission in an area of a Chagas disease outbreak in the Brazilian northeast evaluated through entomological, socioeconomic and schooling indicators.

Chagas disease is a neglected tropical disease strongly associated with low socioeconomic status, affecting nearly 8 million people - mainly Latin Americans. The current infection risk is based on acute case reports, most of which are typically associated with oral transmissions. In the semi-arid region of Northeastern Brazil, serious outbreaks of this transmission type have surged in the last years. One of those occurred in 2016 in the state of Rio Grande do Norte. Rural residents of four municipalities surrounding Marcelino Vieira ingested sugar cane juice - which was probably ground with Trypanosoma cruzi-infected insects. Eighteen cases of Chagas disease were confirmed serologically, with two deaths reported. Socioeconomic information, schooling of residents and the structure of peridomestic and domestic environments in the rural area of Marcelino Vieira, along with entomological indicators, were investigated to understand better the factors related to the outbreaks in this region. We found triatomines (mainly Triatoma brasiliensis) in 35% (24/67) of domiciliary units and all rocky outcrops inspected (n = 7). Overall, 25% (91/357) of examined T. brasiliensis were infected by T. cruzi in artificial ecotopes, with almost the same prevalence in the sylvatic environment (22%; 35/154). Among all ecotopes investigated, wood/tile/brick piles were the ones linked to high insect infestations and triatomine T. cruzi infection prevalence. Ninety-five percent of people interviewed recognized the triatomines and knew the classic route of transmission of disease - triatomine bite-dependent. However, only 7.5% admitted knowledge that Chagas disease can also be acquired orally - which poses a risk this transmission route currently recognized. Here, we highlight the physical proximity between humans and triatomine populations with high T. cruzi infection prevalence as an additional risk factor to oral/vector contaminations. In sum, residents have low income, low level of education, and/or a willful disregard for the routes of Chagas disease transmission (specifically oral transmission), a combination of factors that may have favored the Chagas disease outbreak. We here provide recommendations to avoid further outbreaks.

Dynamics of food sources, ecotypic distribution and Trypanosoma cruzi infection in Triatoma brasiliensis from the northeast of Brazil.

Innovative approaches used to combat Chagas disease transmission tend to combine a set of comprehensive efforts to understand the ecology of local vectors. In this work we identified molecularly the blood meal of 181 Triatoma brasiliensis, distributed in 18 populations (8 sylvatic and 10 peridomestic), which were collected across a range of 240 km (East-West) and 95 km (North-South) in the semi-arid region of northeastern, Brazil. We used the vertebrate mitochondrial gene (cytochrome B) sequencing applied to DNA isolated from bug midgut to identify the insect blood meal sources via the BLAST procedure. The peridomestic populations were classified according to two main hypotheses of site-occupancy for T. brasiliensis: the first says that the infestation is mainly driven by structures that resemble its natural habitat (stony-like ecotopes) and the second assumes that it is associated with key-hosts (rodents and goats). Rodents of the Caviidae family (Galea spixii and Kerodon rupestris) were identified as the key-host of T. brasiliensis, but also the potential Trypanosoma cruzi reservoir-able to connect the sylvatic and domestic T. cruzi cycle. Cats also deserve to be studied better, as potential T. cruzi reservoirs. By modeling the food sources + site-occupancy + T. cruzi natural infection, we identified man-made ecotopes suitable for forming dense triatomine infestations with high rates of T. cruzi natural infection, which may be taken into account for vector control measures.

Drivers of molecular and morphometric variation in Triatoma brasiliensis (Hemiptera: Triatominae): the resolution of geometric morphometrics for populational structuring on a microgeographical scale.

BACKGROUND: The protozoan Trypanosoma cruzi circulates in semiarid areas of northeastern Brazil in distinct ecotopes (sylvatic, peridomestic and domestic) where Triatoma brasiliensis Neiva, 1911 is the most important Chagas disease vector. In this study, we analyzed microevolutionary and demographic aspects of T. brasiliensis populations at the ecotypic, micro and macro-geographic scales by combining morphometrics and molecular results. Additionally, we aimed to address the resolution of both markers for delimiting populations in distinct scales. METHODS: We sampled populations of T. brasiliensis from distinct ecotypic and geographic sites in the states Rio Grande do Norte (RN) and Paraíba (PB). The geometric morphometry was carried out with 13 landmarks on the right wings (n = 698) and the genetic structure was assessed by sequencing a region of cytochrome b mitochondrial gene (n = 221). Mahalanobis distance (MD) and coefficient of molecular differentiation (ΦST) were calculated among all pairs of populations. The results of comparisons generated MD and ΦST dendrograms, and graphics of canonical variate analysis (CVA). RESULTS: Little structure was observed for both markers for macro-geographic scales. Mantel tests comparing geographic, morphometric and genetic matrices showed low correlation (all R2 < 0.35). The factorial graphics built with the CVA evidenced population delimitation for the morphometric data at micro-geographic scales. CONCLUSIONS: We believe that T. brasiliensis carries in its genotype a source of information to allow the phenotypical plasticity across its whole distribution for shaping populations, which may have caused a lack of population delimitation for CVAs in morphometric analysis for macro-geographic scale analysis. On the other hand, the pattern of morphometric results in micro-geographic scales showed well-defined groups, highlighting the potential of this tool to inferences on the source for infestation.

Triatoma petrocchiae (Hemiptera, Reduviidae, Triatominae): A Chagas disease vector of T. brasiliensis species complex associated to reptiles.

In semi-arid areas of northeastern Brazil, Chagas disease vectors of Triatoma brasiliensis species complex comprise a monophyletic group of kissing bugs that inhabit rock outcrops. Most of them exhibit allopatric or parapatric distribution; the exception is T. petrocchiae, which is found in cohabitation with T. brasiliensis in rock outcrops. We used vertebrate mitochondrial gene sequencing applied to DNA isolated from bug midgut to identify the insect blood meal sources via BLAST procedure. Fourteen sylvatic insects from four geographic districts in the states of Rio Grande do Norte and Paraíba had their blood meal sources detected. While T. brasiliensis is recorded to be associated mainly (52-71%) with rodents, T. petrocchiae samples were strongly associated (86%) with reptiles of Tropidurus and Hemidactylus genera. We suggest that T. petrocchiae is the single member within this complex to be associated with reptiles, indicating a distinct niche occupation related to the trophic resources.

Molecular eco-epidemiology on the sympatric Chagas disease vectors Triatoma brasiliensis and Triatoma petrocchiae: Ecotopes, genetic variation, natural infection prevalence by trypanosomatids and parasite genotyping.

Triatoma petrocchiae is the newly member of the Triatoma brasiliensis species complex. This species overlaps with T. brasiliensis in geographic and ecotypic occupation in the sylvatic habitat because both inhabit rocky outcrops in the semi-arid portion of Brazilian northeast. In this region T. brasiliensis is the most important Chagas disease vector because it constantly colonizes domiciles. In contrast, T. petrocchiae is rarely found in peri or intradomiciliary habitats - reason why little is known about this species. Therefore, Here, we present information for the first time on. the T. petrocchiae ecotopes, genetic diversity, Trypanosoma cruzi prevalence/genotyping in comparison to T. brasiliensis. We found T. brasilensis (N = 223) and T. petrocchiae (N = 69) in co-habitation in rocky outcrops in three Districts of Paraíba and Rio Grande do Norte states. Forty-tree T. petrocchiae insects of eleven sampling spots (composing three geographic populations) were genotyped for the mitochondrial Cyt B gene and little geographic structure was observed. Tajima's D test suggested that species is evolving toward a mutation-drift equilibrium in our collection range. Sylvatic T. petrocchiae had 4% (3/68) of infected insects by T. cruzi, whereas T. brasiliensis had 26% (59/223). Fluorescent Fragment Length Barcoding demonstrated that all three T. petrocchiae harbored TcI whereas T. brasiliensis had TcI, but also TcIII, TcII/TcVI and T. rangeli genotype A, sometimes under mixed infections. None of infected T. petrocchiae were carrying mixed infections. However, this result should be confirmed using a larger pool of infected bugs. We here presented the first documentation of T. rangeli infecting T. brasiliensis. The finding of infected T. petrocchiae calls for constant vector monitoring because the epidemiologic scenario is dynamic and sylvatic vectors are progressively found in adaptation to anthropic environments.