Gustavo Fabián Enriquez

Factors Affecting Infestation by Triatoma infestans in a Rural Area of the Humid Chaco in Argentina: A Multi-Model Inference Approach

Background Transmission of Trypanosoma cruzi by Triatoma infestans remains a major public health problem in the Gran Chaco ecoregion, where understanding of the determinants of house infestation is limited. We conducted a cross-sectional study to model factors affecting bug presence and abundance at sites within house compounds in a well-defined rural area in the humid Argentine Chaco. Methodology/Principal Findings Triatoma infestans bugs were found in 45.9% of 327 inhabited house compounds but only in 7.4% of the 2,584 sites inspected systematically on these compounds, even though the last insecticide spraying campaign was conducted 12 years before. Infested sites were significantly aggregated at distances of 0.8–2.5 km. The most frequently infested ecotopes were domiciles, kitchens, storerooms, chicken coops and nests; corrals were rarely infested. Domiciles with mud walls and roofs of thatch or corrugated tarred cardboard were more often infested (32.2%) than domiciles with brick-and-cement walls and corrugated metal-sheet roofs (15.1%). A multi-model inference approach using Akaikes information criterion was applied to assess the relative importance of each variable by running all possible (17,406) models resulting from all combinations of variables. Availability of refuges for bugs, construction with tarred cardboard, and host abundance (humans, dogs, cats, and poultry) per site were positively associated with infestation and abundance, whereas reported insecticide use showed a negative association. Ethnic background (Creole or Toba) adjusted for other factors showed little or no association. Conclusions/Significance Promotion and effective implementation of housing improvement (including key peridomestic structures) combined with appropriate insecticide use and host management practices are needed to eliminate infestations. Fewer refuges are likely to result in fewer residual foci after insecticide spraying, and will facilitate community-based vector surveillance. A more integrated perspective that considers simultaneously social, economic and biological processes at local and regional scales is needed to attain effective, sustainable vector and disease control.

Unexpected Failures to Control Chagas Disease Vectors with Pyrethroid Spraying in Northern Argentina

ABSTRACT Effectiveness of the elimination efforts against Triatoma infestans (Klug) in South America through residual application of pyrethroid insecticides has been highly variable in the Gran Chaco region. We investigated apparent vector control failures after a standard community-wide spraying with deltamethrin SC in a rural area of northeastern Argentina encompassing 353 houses. Insecticide spraying reduced house infestation less than expected: from 49.5% at baseline to 12.3 and 6.7% at 4 and 8 mo postspraying, respectively. Persistent infestations were detected in 28.4% of houses, and numerous colonies with late-stage bugs were recorded after the interventions. Laboratory bioassays showed reduced susceptibility to pyrethroids in the local bug populations. Eleven of 14 bug populations showed reduced mortality in diagnostic dose assays (range, 35 ± 5% to 97 ± 8%) whereas the remainder had 100% mortality. A fully enclosed residual bug population in a large chicken coop survived four pyrethroid sprays, including two double-dose applications, and was finally suppressed with malathion. The estimated resistance ratio of this bug population was 7.17 (range, 4.47–11.50). Our field data combined with laboratory bioassays and a residual foci experiment demonstrate that the initial failure to suppress T. infestans was mainly because of the unexpected occurrence of reduced susceptibility to deltamethrin in an area last treated with pyrethroid insecticides 12 yr earlier. Our results underline the need for close monitoring of the impact of insecticide spraying to provide early warning of possible problems because of enhanced resistance or tolerance and determine appropriate responses.

The sylvatic transmission cycle of Trypanosoma cruzi in a rural area in the humid Chaco of Argentina

Little is known about the sylvatic transmission cycle of Trypanosoma cruzi in the Gran Chaco ecoregion. We conducted surveys to identify the main sylvatic hosts of T. cruzi, parasite discrete typing units and vector species involved in Pampa del Indio, a rural area in the humid Argentinean Chaco. A total of 44 mammals from 14 species were captured and examined for infection by xenodiagnosis and polymerase chain reaction amplification of the hyper-variable region of kinetoplast DNA minicircles of T. cruzi (kDNA-PCR). Ten (22.7%) mammals were positive by xenodiagnosis or kDNA-PCR. Four of 11 (36%) Didelphis albiventris (white-eared opossums) and six of nine (67%) Dasypus novemcinctus (nine-banded armadillos) were positive by xenodiagnosis and or kDNA-PCR. Rodents, other armadillo species, felids, crab-eating raccoons, hares and rabbits were not infected. Positive animals were highly infectious to the bugs that fed upon them as determined by xenodiagnosis. All positive opossums were infected with T. cruzi I and all positive nine-banded armadillos with T. cruzi III. Extensive searches in sylvatic habitats using 718 Noireau trap-nights only yielded Triatoma sordida whereas no bug was collected in 26 light-trap nights. Four armadillos or opossums fitted with a spool-and-line device were successfully tracked to their refuges; only one Panstrongylus geniculatus was found in an armadillo burrow. No sylvatic triatomine was infected with T. cruzi by microscopical examination or kDNA-PCR. Our results indicate that two independent sylvatic transmission cycles of T. cruzi occur in the humid Chaco. The putative vectors of both cycles need to be identified conclusively.

New Sylvatic Hosts of Trypanosoma cruzi and Their Reservoir Competence in the Humid Chaco of Argentina: A Longitudinal Study

A four-year longitudinal study of the structure of sylvatic transmission cycles of Trypanosoma cruzi, reservoir host competence and parasite discrete typing units was conducted in a disturbed rural area of the humid Chaco in Argentina. Among 190 mammals examined by xenodiagnosis and polymerase chain reaction amplification, the composite prevalence of infection was substantially higher in Dasypus novemcinctus armadillos (57.7%) and Didelphis albiventris opossums (38.1%) than in Euphractus sexcinctus (20.0%), Tolypeutes matacus (12.5%), and Chaetophractus vellerosus (6.3%) armadillos. Trypanosoma cruzi was detected for the first time in Thylamys pusilla small opossums and in two unidentified small rodents. Infection was spatially aggregated only in armadillos. All Didelphis were infected with T. cruzi I and all armadillo species were infected with T. cruzi III, implying two distinct sylvatic cycles with no inputs from the domestic cycle. Dasypus armadillos and Didelphis opossums were much more infectious to vectors than other armadillos, small opossums, or rodents.

High levels of Trypanosoma cruzi DNA determined by qPCR and infectiousness to Triatoma infestans support dogs and cats are major sources of parasites for domestic transmission

The competence of reservoir hosts of vector-borne pathogens is directly linked to its capacity to infect the vector. Domestic dogs and cats are major domestic reservoir hosts of Trypanosoma cruzi, and exhibit a much higher infectiousness to triatomines than seropositive humans. We quantified the concentration of T. cruzi DNA in the peripheral blood of naturally-infected dogs and cats (a surrogate of intensity of parasitemia), and evaluated its association with infectiousness to the vector in a high-risk area of the Argentinean Chaco. To measure infectiousness, 44 infected dogs and 15 infected cats were each exposed to xenodiagnosis with 10-20 uninfected, laboratory-reared Triatoma infestans that blood-fed to repletion and were later individually examined for infection by optical microscopy. Parasite DNA concentration (expressed as equivalent amounts of parasite DNA per mL, Pe/mL) was estimated by real-time PCR amplification of the nuclear satellite DNA. Infectiousness increased steeply with parasite DNA concentration both in dogs and cats. Neither the median parasite load nor the mean infectiousness differed significantly between dogs (8.1Pe/mL and 48%) and cats (9.7Pe/mL and 44%), respectively. The infectiousness of dogs was positively and significantly associated with parasite load and an index of the hosts body condition, but not with dogs age, parasite discrete typing unit and exposure to infected bugs in a random-effects multiple logistic regression model. Real-time PCR was more sensitive and less time-consuming than xenodiagnosis, and in conjunction with the body condition index, may be used to identify highly infectious hosts and implement novel control strategies.

Discrete typing units of Trypanosoma cruzi identified in rural dogs and cats in the humid Argentinean Chaco

The discrete typing units (DTUs) of Trypanosoma cruzi that infect domestic dogs and cats have rarely been studied. With this purpose we conducted a cross-sectional xenodiagnostic survey of dog and cat populations residing in 2 infested rural villages in Pampa del Indio, in the humid Argentine Chaco. Parasites were isolated by culture from 44 dogs and 12 cats with a positive xenodiagnosis. DTUs were identified from parasite culture samples using a strategy based on multiple polymerase-chain reactions. TcVI was identified in 37 of 44 dogs and in 10 of 12 cats, whereas TcV was identified in 5 dogs and in 2 cats -a new finding for cats. No mixed infections were detected. The occurrence of 2 dogs infected with TcIII -classically found in armadillos- suggests a probable link with the local sylvatic transmission cycle involving Dasypus novemcinctus armadillos and a potential risk of human infection with TcIII. Our study reinforces the importance of dogs and cats as domestic reservoir hosts and sources of various DTUs infecting humans, and suggests a link between dogs and the sylvatic transmission cycle of TcIII.

Heterogeneities in the ecoepidemiology of Trypanosoma cruzi infection in rural communities of the Argentinean Chaco.

We conducted a cross-sectional survey of Trypanosoma cruzi infection of Triatoma infestans as well as dogs and cats in 327 households from a well-defined rural area in northeastern Argentina to test whether the household distribution of infection differed between local ethnic groups (Tobas and Creoles) and identify risk factors for host infection. Overall prevalence of infection of bugs (27.2%; 95% confidence interval = 25.3-29.3%), dogs (26.0%; 95% confidence interval = 23.3-30.1%), and cats examined (28.7%; 95% confidence interval = 20.2-39.0%) was similar. A multimodel inference approach showed that infection in dogs was associated strongly with the intensity and duration of local exposure to infected bugs and moderately with household ethnic background. Overall, Toba households were at a substantially greater risk of infection than Creole households. The strong heterogeneities in the distribution of bug, dog, and cat infections at household, village, and ethnic group levels may be used for targeted vector and disease control.

Detection of Trypanosoma cruzi infection in naturally infected dogs and cats using serological, parasitological and molecular methods

Domestic dogs and cats are major domestic reservoir hosts of Trypanosoma cruzi and a risk factor for parasite transmission. In this study we assessed the relative performance of a polymerase chain reaction assay targeted to minicircle DNA (kDNA-PCR) in reference to conventional serological tests, a rapid dipstick test and xenodiagnosis to detect T. cruzi infection in dogs and cats from an endemic rural area in northeastern Argentina. A total of 43 dogs and 13 cats seropositive for T. cruzi by an immunosorbent assay (ELISA) and an indirect hemagglutination assay (IHA), which had been examined by xenodiagnosis, were also tested by kDNA-PCR. kDNA-PCR was nearly as sensitive as xenodiagnosis for detecting T. cruzi-infectious dogs and cats. kDNA-PCR was slightly more sensitive than xenodiagnosis in seropositive dogs (91% versus 86%, respectively) and cats (77% against 54%, respectively), but failed to detect all of the seropositive individuals. ELISA and IHA detected all xenodiagnosis-positive dogs and both outcomes largely agreed (kappa coefficient, κ=0.92), whereas both assays failed to detect all of the xenodiagnosis-positive cats and their agreement was moderate (κ=0.68). In dogs, the sensitivity of the dipstick test was 95% and agreed closely with the outcome of conventional serological tests (κ=0.82). The high sensitivity of kDNA-PCR to detect T. cruzi infections in naturally infected dogs and cats supports its application as a diagnostic tool complementary to serology and may replace the use of xenodiagnosis or hemoculture.

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.

The peri-urban interface and house infestation with Triatoma infestans in the Argentine Chaco: an underreported process?

Peri-urban infestations with triatomine bugs, their sources and their dynamics have rarely been investigated. Here, we corroborated the reported occurrence of Triatoma infestans in a peri-urban area and in neighbouring rural houses in Pampa del Indio, in the Argentine Chaco, and identified its putative sources using spatial analysis and demographic questionnaires. Peri-urban householders reported that 10% of their premises had triatomines, whereas T. infestans was collected by timed manual searches or community-based surveillance in only nine (3%) houses. Trypanosoma cruzi-infected T. infestans and Triatoma sordida were collected indoors only in peri-urban houses and were infected with TcV and TcI, respectively. The triatomines fed on chickens, cats and humans. Peri-urban infestations were most frequent in a squatter settlement and particularly within the recently built mud houses of rural immigrants, with large-sized households, more dogs and cats and more crowding. Several of the observed infestations were most likely associated with passive bug transport from other sources and with active bug dispersal from neighbouring foci. Thus, the households in the squatter settlement were at a greater risk of bug invasion and colonisation. In sum, the incipient process of domestic colonisation and transmission, along with persistent rural-to-urban migratory flows and unplanned urbanisation, indicate the need for active vector surveillance and control actions at the peri-urban interface of the Gran Chaco.