Trypanosoma cruzi from the Paraguayan Chaco: isoenzyme profiles of strains isolated at Makthlawaiya

At Makthlawaiya, in the Paraguayan Chaco, the prevalence of Trypanosoma (Schizotrypanum) cruzi infection among both domestic Triatoma infestans and domestic dogs was 38%, and IgG anti-T. cruzi antibody was detected by the quantitative enzyme-linked immunosorbent assay (ELISA) in 80% (105/133) of human sera. Ninety percent (25/28) of T. cruzi strains isolated from both T. infestans and dogs showed heterozygous isoenzyme profiles for glucose phosphate isomerase, phosphoglucomutase and 6-phosphogluconate dehydrogenase. These strains appeared to be closely related to Bolivian zymodeme 2. Three Paraguayan T. cruzi strains showed homozygous isoenzyme profiles, similar to those of major Brazilian zymodemes. It was concluded that T. cruzi strains with heterozygous isoenzyme profiles predominate in domestic transmission cycles in this highly endemic area of the Paraguayan Chaco.     

Mitochondrial DNA variation of Triatoma infestans populations and its implication on the specific status of T. melanosoma

DNA sequence comparison of 412 base-pairs fragments of the mitochondrial cytochrome B gene was used to infer the genetic structure of nine geographical Triatoma infestans populations and their phylogenetic relationship with T. melanosoma and T. brasiliensis. T. infestans and T. melanosoma were compared by morphometry, allozyme and cytogenetic analyses, as well as subjected to reciprocal crosses, in order to clarify the taxonomic status of the latter. No differences were found to distinguish the two species and the crosses between them yielded progeny. T. infestans populations presented four haplotypes that could be separated in two clusters: one formed by the samples from Bolivia (Andes and Chaco) and the other formed by samples from Argentina and Brazil. Silvatic and domestic T. infestans populations from Bolivia (Andes) were genetically identical.     

First report of colonies of sylvatic Triatoma infestans (Hemiptera: Reduviidae) in the Paraguayan Chaco, using a trained dog

In the Gran Chaco region, control of Triatoma infestans has been limited by persistent domestic infestations despite the efforts of the Vector Control Services. In Paraguay, this region is the highest endemic area in the country, showing high levels of indoor and outdoor infestation. Although sylvatic T. infestans have been found in the Bolivian and Argentine Chaco, similar searches for sylvatic populations of this species in Paraguay had been unsuccessful over the last 20 years. Here we present a new approach to detecting sylvatic Triatominae, using a trained dog, which has successfully confirmed sylvatic populations of T. infestans and other triatomine species in Paraguay. A total of 22 specimens corresponding to dark morph forms of T. infestans were collected, and 14 were confirmed as T. infestans by the mitochondrial cytochrome B gene analysis. Through this analysis, one of which were previously reported and a second that was a new haplotype. Triatomines were captured from amongst vegetation such as dry branches and hollows trees of different species such Aspidosperma quebracho-blanco, Bulnesia sarmientoi and Stetsonia coryne. The colonies found have been small and without apparent infection with Trypanosoma cruzi. During the study, Triatoma sordida and Triatoma guasayana have also been found in ecotopes close to those of T. infestans.     

Molecular Population Genetics and Evolution of the Chagas’ Disease Vector Triatoma infestans (Hemiptera: Reduviidae)

Triatoma infestans (Klug) is the main vector of Chagas’ disease in the Southern Cone of Latin America between the latitudes 10° S and 46° S. The long-term effectiveness of the control campaigns is greatly dependent upon the vector population structure. Mitochondrial DNA (mtDNA) genes have been used in a number of T. infestans population genetic analyses. However, the maternally inherited markers as well as nuclear ribosomal DNA analyzed until the present exhibited low or limited levels of variation. Analyses based on microsatellite markers strongly supported the existence of some type of stratification in T. infestans populations and supported the hypothesis of vector population recovery from survivors of the insecticide-treated areas, highlighting the value of population genetic analyses in assessing the effectiveness of Chagas’ disease vector control programmes. Although phylogeographic studies have generally suggested a Bolivian Andean origin of T. infestans, they recovered two reciprocal monophyletic groups of T. infestans and Bolivian populations who were not basal as expected for an ancestral group. In addition, a non-Andean origin could not be excluded by mtDNA genealogies that included sylvatic bugs from Gran Chaco. On the other side, mitochondrial and microsatellite markers supported the hypothesis of two independent migration events of colonization and secondary contacts in southern South America. Since the phylogenetic analyses remain inconclusive, more sequences, not only from mitochondrial genes but also from nuclear genes, need to be examined.     

Wing geometry in Triatoma infestans (Klug) and T. melanosoma Martinez, Olmedo & Carcavallo (Hemiptera: Reduviidae)

Abstract. Geometric morphometrics is a novel approach to biological shape analysis, and its application to medical entomology is just beginning. Here, we use it to examine geographical and interspecific variation in the Triatoma infestans complex, vectors of Chagas disease in southern Latin America. Using six landmarks defining the membranous part of the hemelytra, we analysed the geographical patterns of variation in several populations of T . infestans , including the recently described 'dark morph' of T . infestans from hollow trees in the Bolivian Chaco. As a potential outgroup, T. melanosoma was added to the sample. A consistent geographical differentiation was observed, but no evidence was found in either sex to consider either the dark morph or T. melanosoma as a species distinct from T. infestans . Triatoma melanosoma is relegated to synonymy with T. infestans (syn.n.).     

Residual effect of a micro-encapsulated formulation of organophosphates and piriproxifen on the mortality of deltamethrin resistant Triatoma infestans populations in rural houses of the Bolivian Chaco region

The Bolivian Chaco is part of the endemic region of Chagas disease and an area where pyrethroid resistant Triatoma infestans (Hemiptera: Reduviidae) populations has been reported. The World Health Organization identified these resistant populations as an important focus for research. The objective of this study was to evaluate the residual effect of a micro-encapsulated formulation containing organophosphate active ingredients and a juvenile hormone analogue (Inesfly 5A IGR) on the mortality of T. infestans. Studies took place in rural houses of the Bolivian Chaco that were treated up to 34 months before and evaluated the susceptibility to pyrethroids of the offspring of field collected insects. Thirty houses were randomly selected within three communities to carry out wall bio-assays with T. infestans nymphs. Mortality was recorded 24, 48 and 72 h after wall contact. Eggs laid by females collected in the area were used to obtain first-instar nymphs and carry out pyrethroid susceptibility tests. The wall bio-assays showed that the micro-encapsulated insecticide eliminates T. infestans populations and produces detectable mortality of insects exposed to walls treated 34 months prior to the tests. The discriminant dose of deltamethrin (0.01 mg/mL) showed 65% nymph survival, whereas at the highest tested dose (1.0 mg/mL) 14% of the nymphs survived. These results show that Inesfly 5A IGR is an appropriate tool for the elimination of intradomestic and peridomestic populations of T. infestans resistant to pyrethroids.     

Infestation of rural houses by Triatoma infestans in the region of Los Llanos (La Rioja, Argentina)

Vectorial transmission of Chagas disease has been strongly reduced in most parts of the Southern Cone countries of South America, except in the Gran Chaco region of Argentina, Bolivia, and Paraguay. Given periodical interruptions of the vector control programmes in the endemic region of the Gran Chaco of Argentina, the vectorial transmission of the disease has been increasing during the last years. From the beginning of 2004, the provincial Ministry of Health of La Rioja, Argentina, started a vector control programme to cover the rural houses of the Los Llanos area in the southwestern area of the Gran Chaco region. This article reports the result of a standardized entomological survey and insecticide application against Chagas disease vectors in the intra and peridomestic structures of the rural houses of Los Llanos. A total of 4062 houses were inspected, of which 46.8% were found to be infested by Triatoma infestans. Infestation by vector species other than T. infestans was less than 0.5%(T. eratyrusiformis and T. platensis). Intradomestic infestation was found in 27.2%, whereas peridomestic infestation was found in 39.3% of the houses. The lowest figure of intradomestic infestation was 6.6% (Department F Varela), and the highest value of intradomestic infestation was 45.1% (Department Independencia). In spite of the demonstrated success of vector control elsewhere, this study shows that the vector populations are susceptible to pyrethroid insecticides in the southern area of the Gran Chaco of Argentina, that there still are regions where rural houses show heavy infestation by T. infestans associated with big peridomestic structures and that the vectorial transmission of the Chagas disease will continue, unless a sustained and well organized vector control effort is installed in the region.     

Changes in the sexual dimorphism of triatominae in the transition from natural to artificial habitats.

A shift from large to small average sizes was observed in Triatoma infestans and Rhodnius domesticus between field and domestic (or laboratory) conditions of life. It was more pronounced in the female specimens, leading to a subsequent reduction of sexual size dimorphism. This feature is discussed in terms of genetic and populational changes occurring from natural to artificial habitats, in particular those related to population densities. Sexual size dimorphism is then recommended as a new character to be used in the study of species of Triatominae adapting to domestic ecotopes.     

Mitochondrial DNA haplogroups in Amerindian populations from the Gran Chaco.

Mitochondrial DNA from 141 individuals was typed for diagnostic restriction sites and the 9-bp region V deletion to examine the distribution of the founding mtDNA lineage haplotypes in three Amerindian populations (Mataco, Toba, and Pilagá) who currently inhabit the Argentinian part of the Gran Chaco. All four lineages were identified in the three tribes and four population samples studied. Disregarding ethnic or geographic origin, haplogroups B and D exhibit high incidence among the Gran Chaco inhabitants, whereas haplogroups A and C are present in a lower frequency. Three individuals possess none of the characteristic markers and, therefore, could not be assigned to one of those lineages. A neighbor-joining representation of F(ST) distances reflects the current geographic location of the populations, and this also corresponds to their historic distribution. After separating South America into four major regions (Tropical Forest, Andes, Gran Chaco, and Patagonia-Tierra del Fuego), the Gran Chaco populations present the highest average intragroup variability (Hs = 0.64) as well as the lowest intergroup diversity (G(')(ST) = 0.06). These findings suggest high levels of gene flow among the Chaco tribes, as well as with neighbor populations from outside the region.     

Mitochondrial 16S DNA variation in populations of Triatoma infestans from Argentina

Abstract.  Variation in the mtDNA 16S ribosomal RNA gene in populations of Triatoma infestans (Klug) was surveyed. DNA sequence comparisons yielded 18 haplotypes among 130 individuals from 16 localities that represent a large proportion of the range of T. infestans in Argentina. The most common genotype in all populations was found in 76.9% of individuals and two other haplotypes were shared among different populations. The remaining 15 haplotypes were present exclusively in one of the populations, suggesting currently low levels of genetic exchange. Analysis of mtDNA 16S sequences uncovered substantial genetic variation among T. infestans populations. Haplotype and nucleotide diversities varied among populations, from 0% to 0.84% and 0% to 0.29%, respectively. It appears that this locus has a low mutation rate. Uncorrected pairwise differences of T. infestans haplotypes ranged from 0% to 1.2%. The molecular phylogeny supported the monophyly of T. infestans haplotypes and clustered two different pairs of haplotypes with a moderate degree of bootstrap support (∼ 60%). Mitochondrial DNA phylogeographic differentiation was not evident, suggesting a recent rapid spread of the species. Analysis of molecular variance showed hierarchical structure in the data. Considerably less variation was found among T. infestans populations from the northwest and northeast regions than among those belonging to the central area. Such a lack of variation may be indicative of one or more past population bottlenecks.     

Is imidacloprid an effective alternative for controlling pyrethroid-resistant populations of Triatoma infestans (Hemiptera: Reduviidae) in the Gran Chaco ecoregion?

The prevention of Chagas disease is based primarily on the chemical control of Triatoma infestans (Klug) using pyrethroid insecticides. However, high resistance levels, correlated with control failures, have been detected in Argentina and Bolivia. A previous study at our laboratory found that imidacloprid could serve as an alternative to pyrethroid insecticides. We studied the delayed toxicity of imidacloprid and the influence of the blood feeding condition of the insect on the toxicity of this insecticide; we also studied the effectiveness of various commercial imidacloprid formulations against a pyrethroid-resistant T. infestans population from the Gran Chaco ecoregion. Variations in the toxic effects of imidacloprid were not observed up to 72 h after exposure and were not found to depend on the blood feeding condition of susceptible and resistant individuals. Of the three different studied formulations of imidacloprid on glass and filter paper, only the spot-on formulation was effective. This formulation was applied to pigeons at doses of 1, 5, 20 and 40 mg/bird. The nymphs that fed on pigeons treated with 20 mg or 40 mg of the formulation showed a higher mortality rate than the control group one day and seven days post-treatment (p < 0.01). A spot-on formulation of imidacloprid was effective against pyrethroid-resistant T. infestans populations at the laboratory level.     

Molecular phylogeography of the Chagas' disease vector Triatoma infestans in Argentina

Triatoma infestans is the main vector of Chagas' disease in South America between latitudes 10°S and 46°S. A multilocus microsatellite data set of 836 individuals from 27 populations of T. infestans, from all its range of distribution in Argentina, was analyzed. Our results favor the hypothesis of two independent migration events of colonization in Argentina and secondary contacts. The majority of the populations of the western provinces of Catamarca, La Rioja, San Juan and the west of Cordoba province, had almost no shared ancestry with the rest of the populations analyzed. Probably those populations, belonging to localities close to the Andean region, could have been established by the dispersal line of T. infestans that would have arrived to Argentina through the Andes, whereas most of the rest of the populations analyzed may have derived from the dispersal line of T. infestans in non-Andean lowlands. Among them, those from the provinces of Formosa, Chaco, Santiago del Estero and Santa Fe shared different percentages of ancestry and presented lower degree of genetic differentiation. The migratory movement linked to regional economies and possibly associated with passive dispersal, would allow a higher genetic exchange among these populations of T. infestans. This study, using microsatellite markers, provides a new approach for evaluating the validity of the different hypotheses concerning the evolutionary history of this species. Two major lineages of T. infestans, an Andean and non-Andean, are suggested.     

Trypanosoma cruzi from the Paraguayan Chaco: Isoenzyme Profiles of Strains Isolated at Makthlawaiya1

At Makthlawaiya, in the Paraguayan Chaco, the prevalence of Trypanosoma (Schizotrypanum) cruzi infection among both domestic Triatoma infestans and domestic dogs was 38%, and IgG anti-T. cruzi antibody was detected by the quantitative enzyme-linked immunosorbent assay (ELISA) in 80% (105/133) of human sera. Ninety percent (25/28) of T. cruzi strains isolated from both T. infestans and dogs showed heterozygous isoenzyme profiles for glucose phosphate isomerase, phosphoglucomutase and 6-phosphogluconate dehydrogenase. These strains appeared to be closely related to Bolivian zymodeme 2. Three Paraguayan T. cruzi strains showed homozygous isoenzyme profiles, similar to those of major Brazilian zymodemes. It was concluded that T. cruzi strains with heterozygous isoenzyme profiles predominate in domestic transmission cycles in this highly endemic area of the Paraguayan Chaco.     

Phytophthora infestans isolates from Northern China show high virulence diversity but low genotypic diversity

Phenotypic and genotypic characteristics of 48 Phytophthora infestans isolates , collected in five provinces in Northern China between 1997 and 2003, were determined and compared with reference isolates. Characterisation included mating type, virulence, mitochondrial DNA (mtDNA) haplotype and DNA fingerprinting patterns based on simple sequence repeats (SSR) and amplified fragment length polymorphisms (AFLP). All isolates had the A1 mating type, mtDNA haplotype IIa and an identical SSR genotype (designated as SG-01-01) that differed from SSR genotypes found in the reference isolates, including those representing the 'old' US-1 lineage that dominated the P. infestans population worldwide prior to 1980. In contrast, the virulence spectra were highly variable and virulence to all resistance genes present in the standard differential set ( R1 to R11 ) was found. AFLP analysis revealed some diversity; eight different AFLP genotypes were found that could be grouped into two major clusters. This study shows that there is very little genotypic diversity in the P. infestans population in Northern China. The occurrence of many different races within this rather uniform population is discussed in the framework of recent insights into the molecular determinants of avirulence in potato– P. infestans 'gene-for-gene' interactions.     

Determinants of the domiciliary density of Triatoma infestans, vector of Chagas disease

In two heavily infested rural villages of Santiago del Estero, Argentina, where no indoor-spraying with residual insecticides had ever been carried out by official control services, we studied the influence of roof and wall structure, domestic use of insecticide, family size and the number of domestic dogs, on the domiciliary density of Triatoma infestans (Klug). Bug density was significantly associated with (1) the interaction between insecticide use and type of roof, (2) the structure of indoor walls, (3) the number of dogs sharing sleeping areas of people (room-mate dogs), and (4) the number of people plus room-mate dogs, but not with just the number of people resident in the house. The interaction between insecticide use and a roof made of 'simbol', a locally available grass (Pennisetum sp.), also reflected a younger age structure of domestic bug populations. In infested houses, the density of bugs infected with Trypanosoma cruzi Chagas was significantly correlated with overall bug density. Our data suggest that the application of environmental management measures by the affected people, such as plastering of walls and modification of roofs, coupled with keeping dogs away from bedrooms and application of insecticides, should limit the domestic population density of T. infestans and thus reduce the transmission of T. cruzi to people.     

Microsatellite analysis of genetic structure in natural Triatoma infestans (Hemiptera: Reduviidae) populations from Argentina: its implication in assessing the effectiveness of Chagas' disease vector control programmes

The genetic structure in populations of the Chagas' disease vector Triatoma infestans was examined. Comparisons of the levels of genetic variability in populations of this species from areas with different periods since last insecticide treatment and from areas that never received treatment were also carried out. A total of 598 insects from 19 populations were typed for 10 polymorphic microsatellite loci. The average observed and expected heterozygosities ranged from 0.186 to 0.625 and from 0.173 to 0.787, respectively. Genetic drift and limited gene flow appear to have generated a substantial degree of genetic differentiation among the populations of T. infestans. Departures from Hardy-Weinberg expectations due to an excess of homozygotes suggested the presence of null alleles and population subdivision. Microgeographical analysis supports the existence of subdivision in T. infestans populations. Levels of genetic diversity in the majority of the populations of T. infestans from insecticide-treated localities were similar or higher than those detected in populations from areas without treatment. Since the populations of T. infestans are subdivided, a population bottleneck would result in independent genetic drift effects that could randomly preserve different combinations of alleles in each subpopulation. These events followed by a rapid population growth could have preserved high levels of genetic diversity. This study supports the hypothesis of vector population recovery from survivors of the insecticide-treated areas and therefore highlights the value of population genetic analyses in assessing the effectiveness of Chagas' disease vector control programmes.     

Genetic structure and gene flow in Gran Chaco populations of Argentina: evidence from Y-chromosome markers

The Gran Chaco region of central South America has been settled by humans for only the last 4,000-5,000 years. To investigate population structure and variation in this region's indigenous population, we scored males from tribes of the Argentinean part of the Gran Chaco (Pilagá, Wichí, and Toba, representing two major language groups, the Mataco and Guaycurú) for a number of Y-chromosome polymorphisms. The markers included eight microsatellites (DYS19, DYS390, DYS391, DYS392, DYS393, DYS437, DYS438, and DYS439) and the unique native American single nucleotide polymorphism, DYS199. Sixty males (77%) from the total sample carried the DYS199T chromosome, and these were the focus of the present analysis. Unlike most other native Americans, Gran Chaco males show a moderate level of diversity at the DYS19 locus but still less than that seen in non-native Americans. The FST value for Y-chromosome markers in Gran Chaco was 0.107, a value that is more than double that found for mtDNA haplogroups in the same tribes but is not particularly high compared with other Y-chromosome studies. Phylogenetic trees based on all eight microsatellites showed relatively poor correlation of the tribes with either geography or language, and this is possibly explained by their ecology. They are seasonal hunters living in small bands, and under such circumstances drift will be a powerful evolutionary force. An UPGMA tree based on five microsatellites (DYS19, DYS390, DYS391, DYS392, and DYS393), however, showed a more positive relationship, suggesting that DYS437, DYS438, and DYS439 may behave differently from the other microsatellites. No association was found between maternal and paternal lineage distributions. The time to the most recent common ancestor of the DYS199T chromosome is calculated to lie between 13,000 and 26,000 years. This range is consistent with estimates from other Y-chromosome studies as well as with estimates from mtDNA and the archeology of the colonization of South America. We conclude that the male lineages present in the contemporary Gran Chaco population reflect the level of diversity found in South America and that the region's male founders did not carry a restricted gene pool.     

Effects of chickens on the prevalence of infestation and population density of Triatoma infestans in rural houses of north-west Argentina

Abstract Effects of the presence of chickens on population density of the bug Triatoma infestans , principal vector of Chagas disease, were investigated by standardized sampling (indoors and peridomestically) from sixty-eight houses of three rural communities in north-west Argentina, during March 1992.
The domiciliary density of T. infestans increased linearly with the percentage of bugs that fed on chickens, as identified by agar double-diffusion tests. Bug density was significantly higher in houses where hens usually nested indoors than in those where they did not, as determined by concurrent direct observations and interviews of householders. Multiple linear regression analysis of domiciliary bug density on (a) the total number of people, dogs and cats per house; (b) the percentage of domiciliary bugs that fed on chickens, or (c) the indoor-brooding habit of hens, showed the two variables (b, c) related to chickens as significant predictors in each regression model. Inclusion both variables representing chickens increased the fit significantly. Addition of other potentially confounding factors (domestic insecticide use, type of roof and walls of house) did not affect the significant variables retained in the best-fitting regression model.
Peridomestic infestation was positively associated with the household number of fowls. Chickens were the main bloodmeal source of peridomestic T.infestans populations. Human-fed bugs were detected in peridomestic sites of sixteen houses, indicating active dispersal of adults and large nymphal instars from bedroom areas.
Exclusion of hens from domiciliary areas and promotion of chicken sheds, of an appropriate design that would not harbour bug populations, should limit the triatomine population growth rate and reduce the risk of infestation.     

Comparative developmental and susceptibility to insecticide of Bolivian and Brazilian populations of Triatoma infestans

The triatomine bug Triatoma infestans probably originated in Bolivia and dispersed passively over wide areas of South America, where it is the principal vector of Trypanosoma cruzi. In the region of its probable origin this species shows colonization in two different ecotopes, so that it may be encountered in sylvatic as well as in artificial habitats. The sylvatic colonization pattern is not observed in the rest of its range, where T. infestans is exclusive to man-made habitats. The objective of this study was to compare several aspects of two T. infestans populations, one from Minas Gerais (Brazil) and the other from the Cochabamba Valley (Bolivia), with a view to elucidate the factors associated with the different colonization patterns observed for this species. The differences between the developmental cycle, weight, capacity to ingest blood and mortality rate of first instar nymphs should indicate more fragility of Brazilian population that may be related to its elimination possibility.     

Ecogenetics of Triatoma sordida and Triatoma guasayana (Hemiptera: reduviidae) in the Bolivian chaco.

Triatoma guasayana and two putative cryptic species pertaining to T. sordida complex (named groups 1 and 2) occur in sympatry in the Bolivian Chaco. Using multilocus enzyme electrophoresis and subsequent genetic analysis, our work assesses their population distribution and dispersal capacity in domestic, peridomestic, and silvatic environments. Our collections by light trap in the silvatic environment indicated a predominance of T. guasayana and T. sordida group 2 and a lesser abundance of T. sordida group 1 ( pound 10% of the total of captures). Their similar distribution in two silvatic areas 80 km apart supports the hypothesis of their homogeneous dispersal through the Bolivian Chaco. The distribution of T. guasayana and T. sordida groups 1 and 2 was similar between silvatic environment and peridomestic ecotopes where 25% of positive places was occupied by two or three species. Bromeliads were confirmed as favorable shelter for T. guasayana but were free of T. sordida. T. sordida group 1 and to a lesser extent T. guasayana would be more invasive vectors for houses than T. sordida group 2. The spatial partition in the three species sampled in two distant sites suggested a reduced dispersive capacity.