Variety is the spice: the role of morphological variation of Triatoma infestans (Hemiptera, Reduviidae) at a macro-scale§.

Chagas disease is caused by the protozoan parasite Trypanosoma cruzi (Chagas, 1909). One of the primary vectors of T. cruzi in South America is Triatoma infestans (Klug, 1834). This triatomine species is distributed across a huge latitudinal gradient, inhabiting domiciliary , peridomiciliary , and wild environments. Its wide geographic distribution provides an excellent opportunity to study the relationships between environmental gradients and intraspecific morphological variation. In this study, we investigated variations in wing size and shape in T. infestans across six ecoregions. We aimed to address the following questions: How do wing size and shape vary on a regional scale, does morphological variation follow specific patterns along an environmental or latitudinal gradient, and what environmental factors might contribute to wing variation? Geometric morphometric methods were applied to the wings of 162 females belonging to 21 T. infestans populations, 13 from Argentina (n = 105), 5 from Bolivia (n = 42), and 3 from Paraguay (n = 15). A comparison of wing centroid size across the 21 populations showed significant differences. Canonical Variate Analysis (CVA) revealed significant differences in wing shape between the populations from Argentina, Bolivia, and Paraguay, although there was a considerable overlap, especially among the Argentinian populations. Well-structured populations were observed for the Bolivian and Paraguayan groups. Two analyses were performed to assess the association between wing size and shape, geographic and climatic variables: multiple linear regression analysis (MRA) for size and Partial Least Squares (PLS) regression for shape. The MRA showed a significant general model fit. Six temperature-related variables, one precipitation-related variable, and the latitude showed significant associations with wing size. The PLS analysis revealed a significant correlation between wing shape with latitude, longitude, temperature-related, and rainfall-related variables. Wing size and shape in T. infestans populations varied across geographic distribution. Our findings demonstrate that geographic and climatic variables significantly influence T. infestans wing morphology.

Relationship between genetic diversity and morpho-functional characteristics of flight-related traits in Triatoma garciabesi (Hemiptera: Reduviidae).

BACKGROUND: Triatoma garciabesi, a potential vector of the parasitic protozoan Trypanosoma cruzi, which is the causative agent of Chagas disease, is common in peridomestic and wild environments and found throughout northwestern and central Argentina, western Paraguay and the Bolivian Chaco. Genetic differentiation of a species across its range can help to understand dispersal patterns and connectivity between habitats. Dispersal by flight is considered to be the main active dispersal strategy used by triatomines. In particular, the morphological structure of the hemelytra is associated with their function. The aim of this study was to understand how genetic diversity is structured, how morphological variation of dispersal-related traits varies with genetic diversity and how the morphological characteristics of dispersal-related traits may explain the current distribution of genetic lineages in this species. METHODS: Males from 24 populations of T. garciabesi across its distribution range were examined. The cytochrome c oxidase I gene (coI) was used for genetic diversity analyses. A geometric morphometric method based on landmarks was used for morpho-functional analysis of the hemelytra. Centroid size (CS) and shape of the forewing, and contour of both parts of the forewing, the head and the pronotum were characterised. Length and area of the forewing were measured to estimate the aspect ratio. RESULTS: The morphometric and phylogenetic analysis identified two distinct lineages, namely the Eastern and Western lineages, which coincide with different ecological regions. The Eastern lineage is found exclusively in the eastern region of Argentina (Chaco and Formosa provinces), whereas the Western lineage is prevalent in the rest of the geographical range of the species. CS, shape and aspect ratio of the hemelytra differed between lineages. The stiff portion of the forewing was more developed in the Eastern lineage. The shape of both portions of the hemelytra were significantly different between lineages, and the shape of the head and pronotum differed between lineages. CONCLUSIONS: The results provide preliminary insights into the evolution and diversification of T. garciabesi. Variation in the forewing, pronotum and head is congruent with genetic divergence. Consistent with genetic divergence, morphometry variation was clustered according to lineages, with congruent variation in the size and shape of the forewing, pronotum and head.

Population structure and migration in Triatoma infestans (Hemiptera: Reduviidae) from the Argentine Chaco: An integration of genetic and morphometric data.

Genetic and morphological structure of vector populations are useful to identify panmictic groups, reinfestation sources and minimal units for control interventions. Currently, no studies have integrated genetic and morphometric data in Triatoma infestans (Hemiptera: Reduviidae), one of the main vectors of Trypanosoma cruzi. We characterized the genetic and phenotypic structure of T. infestans at a small spatial scale (2-8 km), identified potential migrants and compared flight-related traits among genetic groups and between migrant and non-migrant insects in a well-defined area without insecticide spraying in the previous 12 years. We obtained microsatellite genotypes (N = 303), wing shape and size (N = 164) and body weight-to-length ratios (N = 188) in T. infestans from 11 houses in Pampa del Indio, Argentine Chaco. The uppermost level of genetic structuring partially agreed with the morphological groups, showing high degrees of substructuring. The genetic structure showed a clear spatial pattern around Route 3 and one genetic group overlapped with an area of persistent infestation and insecticide resistance. Females harboured more microsatellite alleles than males, which showed signs of isolation-by-distance. Wing shape discriminant analyses of genetic groups revealed low reclassification scores whereas wing size differed among genetic groups for both sexes. Potential migrants (8%) did not differ from non-migrants in sex, ecotope, wing shape and size. However, male migrants had lower W/L than non-migrants suggesting poorer nutritional state. Our findings may contribute to the understanding of population characteristics, dispersal dynamics and ongoing elimination efforts of T. infestans.

Phenotypic variability in traits related to flight dispersal in the wing dimorphic species Triatoma guasayana.

BACKGROUND: Triatoma guasayana is considered an emerging vector of Chagas disease in the Southern Cone of South America. The presence of a triatomine population with brachypterous individuals, in which both wings are reduced, has recently been reported for this species. The aim of the present study was to determine if flight-related traits varied across populations, if these traits could explain differences in flight capacity across populations and if flight-related traits are associated with geographic and/or climatic variation. METHODS: The study involved 66 male T. guasayana specimens from 10 triatomine populations. Digital images of wing, head and pronotum were used to estimate linear and geometric morphometric variables. Variations in size and shape were analysed using one-way analysis of variance and canonical variate analysis (CVA), respectively. Mantel tests were applied to analyse the relationship between morphometric and geographic distances, and the association between size measurements was analysed using Pearson's correlation. We explored covariation between size and shape variables using partial least square analyses (PLS). The association of geographic and climatic variables with size measurements was tested using linear regression analyses. We performed PLS analyses for shape measurements. RESULTS: Wing size differed significantly across triatomine populations. The CVA showed that wing shape of the brachypterous population is well discriminated from that of the other populations. The Mantel test showed a positive and significant association between wing shape and geographic distances. The heads of the brachypterous population were significantly larger than those of the other populations. Similar to wing shape, the head shape of the brachypterous population was well discriminated from those of the other populations. Pronotum width did not show significant differences across populations. Geographic and climatic factors were associated with size and shape of both the wing and head, but not with pronotum width. CONCLUSIONS: Most of the traits related to flight dispersal varied across populations. Wing shape and head shape were found to be better markers for differentiated morphological variation across populations. Head measurements also varied in accordance with this condition. Geographic and climatic variables were associated with most of the flight-related traits.

Genetic structure of deltamethrin-resistant populations of Triatoma infestans (Hemiptera: Reduviidae) in the Gran Chaco.

The genetic structure of natural populations offers insight into the complexities of their dynamics, information that can be relevant to vector control strategies. Microsatellites are useful neutral markers to investigate the genetic structure and gene flow in Triatoma infestans, one of the main vectors of Chagas disease in South America. Recently, a heterogeneous pyrethroid-resistant hotspot was found in the Argentine Gran Chaco, characterized by the highest levels of deltamethrin resistance found at the present time. We applied population genetics analyses to microsatellite and village data and search for associations between the genetic variability and the heterogeneous toxicological pattern previously found. We genotyped 10 microsatellite loci in 67 T. infestans from 6 villages with no, low, and high pyrethroid resistance. The most genetically diverse populations were those susceptible or with low values of resistance. In contrast, high-resistance populations had lower herozygosity and some monomorphic loci. A negative association was found between variability and resistant ratios. Global and pairwise FSTs indicated significant differentiation between populations. The only susceptible population was discriminated in all the performed studies. Low-resistance populations were also differentiated by a discriminant analysis of principal components (DAPC) and were composed mostly by the same two genetic clusters according to STRUCTURE Bayesian algorithm. Individuals from the high-resistance populations were overlapped in the DAPC and shared significant proportions of a genetic cluster. These observations suggest that the resistant populations might have a common origin, although more genetic markers and samples are required to test this hypothesis more rigorously.

A Microsatellite-Based Analysis of House Infestation With Triatoma Infestans (Hemiptera: Reduviidae) After Insecticide Spraying in the Argentine Chaco.

Prevention of vector-borne transmission of Chagas disease mainly relies on residual insecticide spraying. Despite significant success at a regional scale, house infestation with Triatoma infestans (Klug) (Hemiptera: Reduviidae) still persists in the Gran Chaco ecoregion. One key aspect is the identification of the sources of reinfestant triatomines. After detecting fine-scale genetic structure in two rural villages of Pampa del Indio, Argentine Chaco, we tested hypotheses on the putative origins of the triatomines collected at 4, 8, and 12 mo after insecticide house spraying. We genotyped 10 microsatellite loci in 262 baseline and 83 postspraying triatomines from different houses. Genetic variability was similar between baseline and postspraying populations, but 13 low-frequency alleles were not detected at postspraying. FSTs were not significant between insects collected before and after insecticide spraying at the same house in all but one case, and they clustered together in a neighbor-joining tree. A clustering algorithm detected seven genetic groups, four of them mainly composed of baseline and postspraying insects from the same house. Assignment tests suggested multiple putative sources (including the house of collection) for most postspraying insects but excluded a house located more than 9 km from the study area. The origin of three triatomines was attributed to immigration from other unaccounted sources. Our study is compatible with the hypothesis that house reinfestations in the Argentine Chaco are mostly related to residual foci (i.e., survival of insects within the same community), in agreement with field observations, spatial analysis, and morphometric studies previously published.

The role of sigmodontine rodents as sylvatic hosts of Trypanosoma cruzi in the Argentinean Chaco.

The role of rodents in the sylvatic transmission of Trypanosoma cruzi has seldom been investigated using parasitological and molecular methods. We assessed the occurrence of T. cruzi in wild small rodents from Pampa del Indio, in the Argentinean Chaco, and identified the taxonomic status of positive rodents by sequencing a fragment of cytochrome b gene (cytb) and performing BLAST searches and phylogenetic analyses. A total of 176 Sigmodontinae rodents was captured in six surveys using 5425 trap-nights in a wide range of sylvatic habitats between 2009 and 2011. Host infection was determined by xenodiagnosis and by polymerase chain reaction amplification of the hyper-variable region of kinetoplast DNA minicircles of T. cruzi (kDNA-PCR) from blood samples. None of the 176 rodents examined was xenodiagnosis-positive. The prevalence of infection determined by kDNA-PCR from blood samples was 16.2% (95% confidence interval, 10.1-21.9%). Half of the infections detected by kDNA-PCR were confirmed by nuclear satellite DNA-PCR or by kDNA-PCR of the rectal contents of xenodiagnostic bugs. The 24 positive specimens were assigned to eight species, providing the first records of T. cruzi in Akodon montensis, Akodon toba, Graomys chacoensis, and Oligoryzomys chacoensis. The occurrence of T. cruzi infection in Oligoryzomys nigripes, Calomys callosus, Necromys lasiurus and Oecomys sp. (most probably Oecomys mamorae) from the Gran Chaco is also reported for the first time. Although sigmodontine rodents were frequently infected, the intensity of bug rectal infection with T. cruzi was below the detection limit of xenodiagnosis (subpatent infectiousness to bugs), indicating they had a low reservoir host competence.

Hidden sylvatic foci of the main vector of Chagas disease Triatoma infestans: threats to the vector elimination campaign?

BACKGROUND: Establishing the sources of reinfestation after residual insecticide spraying is crucial for vector elimination programs. Triatoma infestans, traditionally considered to be limited to domestic or peridomestic (abbreviated as D/PD) habitats throughout most of its range, is the target of an elimination program that has achieved limited success in the Gran Chaco region in South America. METHODOLOGY/PRINCIPAL FINDINGS: During a two-year period we conducted semi-annual searches for triatomine bugs in every D/PD site and surrounding sylvatic habitats after full-coverage spraying of pyrethroid insecticides of all houses in a well-defined rural area in northwestern Argentina. We found six low-density sylvatic foci with 24 T. infestans in fallen or standing trees located 110-2,300 m from the nearest house or infested D/PD site detected after insecticide spraying, when house infestations were rare. Analysis of two mitochondrial gene fragments of 20 sylvatic specimens confirmed their species identity as T. infestans and showed that their composite haplotypes were the same as or closely related to D/PD haplotypes. Population studies with 10 polymorphic microsatellite loci and wing geometric morphometry consistently indicated the occurrence of unrestricted gene flow between local D/PD and sylvatic populations. Mitochondrial DNA and microsatellite sibship analyses in the most abundant sylvatic colony revealed descendents from five different females. Spatial analysis showed a significant association between two sylvatic foci and the nearest D/PD bug population found before insecticide spraying. CONCLUSIONS: Our study shows that, despite of its high degree of domesticity, T. infestans has sylvatic colonies with normal chromatic characters (not melanic morphs) highly connected to D/PD conspecifics in the Argentinean Chaco. Sylvatic habitats may provide a transient or permanent refuge after control interventions, and function as sources for D/PD reinfestation. The occurrence of sylvatic foci of T. infestans in the Gran Chaco may pose additional threats to ongoing vector elimination efforts.

Geographic host use variability and host range evolutionary dynamics in the phytophagous insect Apagomerella versicolor (Cerambycidae).

The high diversity of phytophagous insects has been explained by the tendency of the group towards specialization; however, generalism may be advantageous in some environments. The cerambycid Apagomerella versicolor exhibits intraspecific geographical variation in host use. In northern Argentina it is highly specialized on the herb Pluchea sagittalis (Asteraceae), while in central and southern areas it uses seven Asteraceae species. To study host species geographical variation from ecological and evolutionary perspectives, we investigated field host availability and use across a wide latitudinal range, and performed laboratory studies on insect oviposition preference and larval performance and mitochondrial DNA (mtDNA) variation in a phylogeographical framework. Geographic variation in host use was unrelated to host availability but was highly associated with laboratory oviposition preference, larval performance, and mtDNA variation. Genetic studies revealed three geographic races of A. versicolor with gene flow restriction and recent geographic expansion. Trophic generalism and oligophagy within A. versicolor seem to have evolved as adaptations to seasonal and spatial unavailability of the preferred host P. sagittalis in cooler areas of the species' geographic range. No single genotype is successful in all environments; specialization may be advantageous in environments with uniform temporal and spatial host availability, while being a trophic generalist may provide an adaptive advantage in host-constrained environments.

Sylvatic Triatoma infestans (Reduviidae, Triatominae) in the Andean valleys of Bolivia.

Triatoma infestans is the main vector of Chagas disease in the Southern Cone countries. Wild populations of T. infestans appear widespread throughout the Andean valleys of Bolivia. In Cotapachi (2750 m asl), all sorts of rocky outcrops, regardless of their size, provided good refuges for T. infestans. Of the 1120 ecotopes investigated, 330 (29.5%) contained triatomines and 92% of the collected insects were nymphal instars. In the cold season, triatomine densities were similar in small and large outcrops. During the hot season, bug densities were higher in the larger outcrops, particularly in those located in peridomestic sites. T. infestans populations apparently produced one generation per year. Over half the sampled bugs were positive for T. cruzi infection. At Mataral (1750 m asl), a site located in the inter-Andean Chaco, a new morph of T. infestans was detected in a sylvatic environment.