Domestic Transmission Cycles Research Articles

North American import? Charting the origins of an enigmatic Trypanosoma cruzi domestic genotype

BackgroundTrypanosoma cruzi, the agent of Chagas disease, is currently recognized as a complex of six lineages or Discrete Typing Units (DTU): TcI-TcVI. Recent studies have identified a divergent group within TcI - TcIDOM. TcIDOM. is associated with a significant proportion of human TcI infections in South America, largely absent from local wild mammals and vectors, yet closely related to sylvatic strains in North/Central America. Our aim was to examine hypotheses describing the origin of the TcIDOM genotype. We propose two possible scenarios: an emergence of TcIDOM in northern South America as a sister group of North American strain progenitors and dispersal among domestic transmission cycles, or an origin in North America, prior to dispersal back into South American domestic cycles. To provide further insight we undertook high resolution nuclear and mitochondrial genotyping of multiple Central American strains (from areas of México and Guatemala) and included them in an analysis with other published data.FindingsMitochondrial sequence and nuclear microsatellite data revealed a cline in genetic diversity across isolates grouped into three populations: South America, North/Central America and TcIDOM. As such, greatest diversity was observed in South America (Ar = 4.851, π = 0.00712) and lowest in TcIDOM (Ar = 1.813, π = 0.00071). Nuclear genetic clustering (genetic distance based) analyses suggest that TcIDOM is nested within the North/Central American clade.ConclusionsDeclining genetic diversity across the populations, and corresponding hierarchical clustering suggest that emergence of this important human genotype most likely occurred in North/Central America before moving southwards. These data are consistent with early patterns of human dispersal into South America.

Genetic variation of Gongylonema pulchrum from wild animals and cattle in Japan based on ribosomal RNA and mitochondrial cytochrome c oxidase subunit I genes

The gullet worm (Gongylonema pulchrum) has been recorded from a variety of mammals worldwide, including monkeys and humans. Due to its wide host range, it has been suggested that the worm may be transmitted locally to any mammalian host by chance. To investigate this notion, the ribosomal RNA gene (rDNA), mainly regions of the internal transcribed spacers (ITS) 1 and 2, and a cytochrome c oxidase subunit I (COI) region of mitochondrial DNA of G. pulchrum were characterized using parasites from the following hosts located in Japan: cattle, sika deer, wild boars, Japanese macaques, a feral Reeves's muntjac and captive squirrel monkeys. The rDNA nucleotide sequences of G. pulchrum were generally well conserved regardless of their host origin. However, a few insertions/deletions of nucleotides along with a few base substitutions in the ITS1 and ITS2 regions were observed in G. pulchrum from sika deer, wild boars and Japanese macaques, and those differed from G. pulchrum in cattle, the feral Reeves's muntjac and captive squirrel monkeys. The COI sequences of G. pulchrum were further divided into multiple haplotypes and two groups of haplotypes, i.e. those from a majority of sika deer, wild boars and Japanese macaques and those from cattle and zoo animals, were clearly differentiated. Our findings indicate that domestic and sylvatic transmission cycles of the gullet worm are currently present, at least in Japan.

Environmental factors associated with American cutaneous leishmaniasis in a new Andean focus in Colombia

To evaluate the environmental and ecological factors associated with Leishmania transmission and vector abundance in Chaparral, Tolima-Colombia. First, we compared the ecological characteristics, abundance of phlebotomies and potential reservoir hosts in the peridomestic environment (100 m radius) of randomly selected houses, between two townships with high and low cutaneous leishmaniasis incidence. Second, we examined peridomestic correlates of phlebotomine abundance in all 43 houses in the higher risk township. The high transmission township had higher coverage of forest (23%vs. 8.4%) and shade coffee (30.7%vs. 11%), and less coffee monoculture (16.8%vs. 26.2%) and pasture (6.3%vs. 12.3%), compared to the low transmission township. Lutzomyia were more abundant in the high transmission township 2.5 vs. 0.2/trap/night. Lutzomyia longiflocosa was the most common species in both townships: 1021/1450 (70%) and 39/80 (49%). Numbers of potential wild mammal reservoirs were small, although four species were found to be infected with Leishmania (Viannia) spp. In the high transmission township, the overall peridomiciliary capture rate of L. longiflocosa was 1.5/trap/night, and the abundance was higher in houses located nearer to forest (ρ = -0.30, P = 0.05). The findings are consistent with a domestic transmission cycle with the phlebotomies dependent on dense vegetation near the house.

Recent, independent and anthropogenic origins of Trypanosoma cruzi hybrids.

The single celled eukaryote Trypanosoma cruzi, a parasite transmitted by numerous species of triatomine bug in the Americas, causes Chagas disease in humans. T. cruzi generally reproduces asexually and appears to have a clonal population structure. However, two of the six major circulating genetic lineages, TcV and TcVI, are TcII-TcIII inter-lineage hybrids that are frequently isolated from humans in regions where chronic Chagas disease is particularly severe. Nevertheless, a prevalent view is that hybridisation events in T. cruzi were evolutionarily ancient and that active recombination is of little epidemiological importance. We analysed genotypes of hybrid and non-hybrid T. cruzi strains for markers representing three distinct evolutionary rates: nuclear GPI sequences (n = 88), mitochondrial COII-ND1 sequences (n = 107) and 28 polymorphic microsatellite loci (n = 35). Using Maximum Likelihood and Bayesian phylogenetic approaches we dated key evolutionary events in the T. cruzi clade including the emergence of hybrid lineages TcV and TcVI, which we estimated to have occurred within the last 60,000 years. We also found evidence for recent genetic exchange between TcIII and TcIV and between TcI and TcIV. These findings show that evolution of novel recombinants remains a potential epidemiological risk. The clearly distinguishable microsatellite genotypes of TcV and TcVI were highly heterozygous and displayed minimal intra-lineage diversity indicative of even earlier origins than sequence-based estimates. Natural hybrid genotypes resembled typical meiotic F1 progeny, however, evidence for mitochondrial introgression, absence of haploid forms and previous experimental crosses indicate that sexual reproduction in T. cruzi may involve alternatives to canonical meiosis. Overall, the data support two independent hybridisation events between TcII and TcIII and a recent, rapid spread of the hybrid progeny in domestic transmission cycles concomitant with, or as a result of, disruption of natural transmission cycles by human activities.

Virulence in Trypanosoma congolense Savannah subgroup. A comparison between strains and transmission cycles

Trypanosoma congolense strains have been shown to differ in their virulence both between subgroups and within the Savannah subgroup between strains. This review revisits these findings and complements them with information on the virulence of T.congolense Savannah subgroup strains isolated from cattle (domestic transmission cycle) in different geographical areas and of strains isolated in protected areas where trypanotolerant wildlife species are the reservoir of the trypanosomes (sylvatic transmission cycle). The virulence of a total of 62 T.congolense Savannah subgroup strains (50 domestic and 12 sylvatic), determined using a standard protocol in mice, was compared. Virulence varied substantially between strains with, depending on the strain, the median survival time of infected mice varying from five to more than sixty days. The proportion of highly virulent strains (median survival time <10 days) was significantly (P = 0·005) higher in strains from the sylvatic transmission cycle. The analysis highlights repercussions of the domestication of the trypanosomiasis transmission cycle that may have to be taken in consideration in the development of trypanosomiasis control strategies.

Trypanosoma cruzi TcIII / Z3 genotype as agent of an outbreak of Chagas disease in the Brazilian Western Amazonia

Chagas' disease is an emerging and neglected disease in the Brazilian Amazon region, where T. cruzi I predominates among the acute cases of the disease; and T. cruzi III/Z3, a population cluster from sylvatic areas of the Amazon basin, is rarely associated with human infections. On 23rd April 2007, the Foundation for Health Surveillance of the State of Amazonas, Brazil reported an outbreak of acute Chagas disease in the municipality of Coari on the Solimões River banks. Fresh blood examination confirmed the infection in 25 patients. Parasite culture in LIT medium was successful for 18 isolates. Molecular characterization was performed by PCR of the non-transcribed spacer of the mini-exon and by sequencing of the mitochondrial cytochrome c oxidase subunit II (COII) gene. The T. cruzi isolates were all from genotype Z3, and sequencing revealed that all isolates had equal COII sequences compatible with TcIII type, suggesting a single source of infection. To our knowledge, this is the first outbreak of acute cases caused uniquely by the genotype TcIII/Z3. Wild vectors harbouring TcIII stocks contribute to transmission when the triatomine species reaches human food chain or when humans invade the forest environment, where sylvatic cycle constitutes a reservoir of parasites that might be associated with specific epidemiological and clinical traits of the emergent Chagas disease in the Amazon.

A changing environment and the epidemiology of tsetse-transmitted livestock trypanosomiasis

The distribution, prevalence and impact of vector-borne diseases are often affected by anthropogenic environmental changes that alter the interactions between the host, the parasite and the vector. In the case of tsetse-transmitted livestock trypanosomiasis these changes are a result of the encroachment of people and their livestock into tsetse-infected wild areas. This has created a sequence of new epidemiological settings that is changing the relative importance of the domestic or sylvatic trypanosome transmission cycles and is causing concomitant changes in the impact of the disease on livestock. These changes in the dynamics of the epidemiology have an important impact on the factors that need to be considered when developing area-specific strategies for the future management of tsetse-transmitted livestock trypanosomiasis.

Genetic analyses of Trypanosoma cruzi isolates from naturally infected triatomines and humans in northeastern Brazil

Trypanosoma cruzi genetic diversity was investigated in 25 isolates (vectors and humans) from the semiarid zone of the State of Rio Grande do Norte, Brazil. Molecular markers (3′ region of the 24Sα rRNA; mitochondrial cytochrome oxidase subunit 2 (COII) gene; spliced leader intergenic region (SL-IR) gene; allelic size microsatellite polymorphism) identified 56% TcIII (100% Panstrongylus lutzi; 50% Triatoma brasiliensis); 40% TcII (91.7% humans; 50% T. brasiliensis) and 4% TcI (human). Microsatellite analysis revealed monoclonal and heterozygous patterns on one or more microsatellite loci in 64% of T. cruzi isolates (92.3% triatomines; 33.3% humans) and 36% putative polyclonal populations (66.7% humans; 7.7% triatomines) by loci SCLE10, SCLE11, TcTAT20, TcAAAT6, all belonging to TcII. Identical T. cruzi polyclonal profiles (88.9%) were detected, mostly from humans. The adaptative natural plasticity of TcII and TcIII and their potential for maintaining human infection in T. brasiliensis were confirmed. Intraspecific and phylogenetic T. cruzi diversity in the sylvatic and domestic transmission cycles in this specific region will provide exclusive control strategies.

Transmission Dynamics of Trypanosoma cruzi Determined by Low-Stringency Single Primer Polymerase Chain Reaction and Southern Blot Analyses in Four Indigenous Communities of the Sierra Nevada de Santa Marta, Colombia

This study attempted to evaluate the transmission dynamics of Trypanosoma cruzi in four indigenous communities of Sierra Nevada de Santa Marta (SNSM), Colombia. Low-stringency single primer-polymerase chain reaction (LSSP-PCR) of the minicircles and Southern blot analyses were used to characterize samples from patients, vectors, and reservoirs in these communities. The LSSP-PCR profiles revealed a high genetic variability but with similarities among the parasites present in the samples of vectors, patients, and reservoirs of the same and different communities. Cluster and analysis of molecular variance (AMOVA) analyses of data derived from LSSP-PCR and Southern blot suggest a gene flux among populations of T. cruzi circulating in patients, vectors, and reservoirs. The results support the idea that the domestic and wild transmission cycles overlap in the SNSM, with Rhodnius prolixus as the main vector and Triatoma dimidiata playing an important role in the transmission of Chagas disease in this zone, making the vector control strategy by spraying unsuccessful.

ITS–RFLP- and RAPD-based genetic variability of Trypanosoma cruzi I, human and vector strains in Santander (Colombia)

Chagas disease is a severe public health problem in Latin-American countries. In Colombia, the predominance of Trypanosoma cruzi I has been described in the literature, with a broad heterogeneity between strains. However, most of the studies carried out centered on isoenzyme analysis, with a smaller number that focus on other molecular methods. In this report, we discuss the results of a molecular analysis of T. cruzi I strains, isolated from the domestic cycle, from the department of Santander, one of the territorial divisions where the prevalence of infection is highest. Internal transcribed spacer-restriction fragment length polymorphism and random amplification of polymorphic DNA were used to characterize 16 strains from human and vector (Triatominae) hosts. The data reveal a clustering based on the biological origin. Human and vector strains grouped separately; however, three vector strains clustered together with human strains. These results indicate that genetic differences exist in the strains that infect both hosts. We conclude that T. cruzi I populations in the domestic cycle of transmission of Chagas disease in Santander are heterogeneous and are composed of different clones. The epidemiological and biological implications are discussed.

Evolutionary history ofTrypanosoma cruziaccording to antigen genes

Trypanosoma cruzi, the agent of Chagas disease is associated with a very high clinical and epidemiological pleomorphism. This might be better understood through studies on the evolutionary history of the parasite. We explored here the value of antigen genes for the understanding of the evolution within T. cruzi. We selected 11 genes and 12 loci associated with different functions and considered to be involved in host-parasite interaction (cell adhesion, infection, molecular mimicry). The polymorphism of the respective genes in a sample representative of the diversity of T. cruzi was screened by PCR-RFLP and evolutionary relationships were inferred by phenetic analysis. Our results support the classification of T. cruzi in 2 major lineages and 6 discrete typing units (DTUs). The topology of the PCR-RFLP tree was the one that better fitted with the epidemiological features of the different DTUs: (i) lineage I, being encountered in sylvatic as well as domestic transmission cycles, (ii) IIa/c being associated with a sylvatic transmission cycle and (iii) IIb/d/e being associated with a domestic transmission cycle. Our study also supported the hypothesis that the evolutionary history of T. cruzi has been shaped by a series of hybridization events in the framework of a predominant clonal evolution pattern.

African Swine Fever Virus DNA in Soft Ticks, Senegal

African swine fever is a highly contagious disease of pigs in Africa. Although its persistence in Senegal may be caused by asymptomatic carriers involved in the domestic transmission cycle, we demonstrated that the soft tick Ornithodoros sonrai can be naturally infected with the causative agent.

Group discussion: epidemiological and social determinants of Chagas disease and its control in the Amazon countries

The emergence of Chagas disease transmission in theAmazon region is a new challenge for controlling thedisease. It takes on special importance, considering that:(i) vector transmission in homes in many other areas hasnow been brought under control, or at least that the tech-nology available for control presents proven efficiency;(ii) the transmission patterns are different; (iii) some ofits epidemiological and social determinants or risk con-ditions are unknown.The enzootic transmission cycle of Trypanosomacruzi has long been known in the Amazon region. CarlosChagas himself already recognized flagellates isolated frommonkeys of the species Saimiri scirius as T. cruzi, in theyear 1924 (Chagas 1924). Following this, many other ani-mal reservoirs were identified (Deane 1961, 1964). Thetriatomine fauna is also very diversified and natural infec-tion has been observed in several of the species present(Barret 1988, Coura et al. 1994, Schofield 2002).However, the domestic transmission cycle has notbecome established in an evident manner or on a largescale. With the progressive human occupation of the re-gion, introduction of non-autochthonous species or colo-nization by native species as a result of human predatoryaction on the natural environment were considered likely(Silveira et al. 1984, Barret 1988, Silveira 1997). So far, ithas not been possible to confirm this. Nonetheless, au-tochthonous cases started to be recognized in the Ama-zon region, including in countries without a history ofoccurrences of this disease (Shaw 1969, Rambajan 1984,Beauder 1985). Moreover, increasing numbers of suchcases have been found over recent years (Valente 2005).On the one hand, this may be ascribed to possible newevents or environmental changes. On the other hand, thisis certainly due to the greater attention that has been givento studying the disease and performing disease surveil-lance in this region, even if done non-systematically.The means of transmission in the Amazon region donot correspond to those that are recognized as charac-teristic of the endemic disease, with “installation andpermanence” of the vector in the home. The known au-tochthonous cases are produced by the following meansof transmission: (i) oral transmission, which indirectlyor passively may be understood as vectorial, providedthat it only occurs by means of contamination of foodsby the feces of infected triatomines; (ii) vectorial trans-mission in the home, without colonization, by meansof periodic or regular incursions by specimens that in-vade the home; (iii) vectorial transmission outside ofthe home, by means of frequent human entry into for-ests and contact with wild triatomines, as occurs withRhodnius brethesi in the case of extraction activitiesrelating to the piacaba palm tree.The peculiarities of Chagas disease in the Amazonregion require the adoption of a surveillance model dif-fering from what has been followed in historically en-demic areas, in which it is fundamentally centered onentomological surveillance.Likewise, in conceptualizing a surveillance and con-trol model for the disease in this region, the followingmust be considered: (i) the extent of the Amazon terri-tory and the operational difficulties implied; (ii) the factthat it is not recognized as a public health problem inthat region; (iii) the resources already installed in the re-gion that mat serve for preventing endemic Chagas disease.

Trypanosoma cruzi Chagas, 1909: genetic variability of isolates from chronic chagasic patients in the Paraná state, Brazil

The present work had as objective to verify the genetic diversity among strains of Trypanosoma cruzi isolated in chronic chagasic patients in the Paraná state. Fifty patients with compatible clinical symptoms were selected (cardiopathy, megacolon and/or megaesophagus) and that presented positive serological reaction to T. cruzi. Six strains of the protozoan were isolated in hemoculture and identified by electrophoresis in starch thick gel with aid of 14 isoenzymes (6PGD; G6PD; ME¹; ME²; ICD; PGM; GPI; GOT¹; GOT²; NP¹; NP²; DIA; MPI and FH). The statistical analysis were accomplished by the softwares NTSYs and UPGMA. Eighty-four electromorphs were individualized and four isoenzymes (ICD, GPI, 6PGD, e PGM) showed compatibility for heterozygosis. The phenetic analysis evidenced the hypothesis of constant evolution (genetic distances = 0.0536 to 0.1429, Hardy-Weinberg = 1.0000 to 0.0769 and differentiation for exact tests = 1.0000 to 0.0658). A great intra-specific genetic diversity in T. cruzi was verified in the isolates obtained in humans and it indicates that the clonet II is associated to the domestic cycle of transmission.

Trypanosoma cruzi isolates from Chile are heterogeneous and composed of mixed populations when characterized by schizodeme and Southern analyses.

In total, 61 Chilean isolates of Trypanosoma cruzi, were analysed using schizodeme and Southern analysis, using as probes the highly variable regions of minicircles from cloned parasites. Isolates were collected and amplified from domestic and wild triatomines, and from infected subjects in all the endemic areas of Chile. Three major parasite genotypes could be detected in the domestic transmission cycle, whilst 1 major T. cruzi genotype is circulating in the wild transmission cycle. Schizodeme analysis suggested that T. cruzi isolates are mixed populations, whereas the Southern analyses detected only 3 mixed isolates using 4 selected minicircle segments as probes.

Chagas disease in a domestic transmission cycle, southern Texas, USA.

After three dogs died from acute Chagas cardiomyopathy at one location, an investigation was conducted of the home, garage, and grounds of the owner. A serologic study was conducted on stray dogs, and an ecologic niche model was developed to predict areas where the vector Tryiatoma gerstaeckeri might be expected.

Trypanosoma cruzi: presence of the two major phylogenetic lineages and of several lesser discrete typing units (DTUs) in Chile and Paraguay

Multilocus enzyme electrophoresis (MLEE) of 99 Chilean and 11 Paraguayan stocks of Trypanosoma cruzi, the agent of Chagas disease, was performed for 22 variable genetic loci. As previously shown for this parasite in other geographic areas, a pattern of long-term clonal evolution of T. cruzi genotypes was inferred, both by strong departures of Hardy–Weinberg expectations and high linkage disequilibrium. The presence of the two major phylogenetic lineages that subdivide the species T. cruzi [Tibayrenc, M., 1995. Population genetics of parasitic protozoa and other microorganisms. In: Baker, J.R., Muller, R., Rollinson, D. (Eds.), Advances in Parasitology, vol. 36, Academic Press, New York, pp. 47–115; Souto, R.P., Fernandes, O., Macedo, A.M., Campbell, D.A., Zingales, B., 1996. DNA markers define two major phylogenetic lineages of Trypanosoma cruzi. Mol. Biochem. Parasitol. 83, 141–152], and of several lesser genetic subdivisions (‘discrete typing units’ or DTUs; Tibayrenc, M., 1998a. Genetic epidemiology of parasitic protozoa and other infectious agents: the need for an integrated approach. Int. J. Parasitol. 28 (1), 85–104; Tibayrenc, M., 1998b. Beyond strain typing and molecular epidemiology: integrated genetic epidemiology of infectious diseases. Parasitol. Today 14, 323–329; Tibayrenc, M., 1998c. Integrated genetic epidemiology of infectious diseases: the Chagas model. Mem. Inst. Oswaldo Cruz 93 (5), 577–580), was recorded in this region. Comparison between clonal populations in sylvatic and domestic transmission cycles of the disease in Chile strongly suggests that these two cycles are at least partially separated from one another.

The epidemiology and control of leishmaniasis in Andean countries.

This paper reviews the current knowledge of leishmaniasis epidemiology in Venezuela, Colombia, Ecuador, Peru, and Bolivia. In all 5 countries leishmaniasis is endemic in both the Andean highlands and the Amazon basin. The sandfly vectors belong to subgenera Helcocyrtomyia, Nyssomiya, Lutzomyia, and Psychodopygus, and the Verrucarum group. Most human infections are caused by Leishmania in the Viannia subgenus. Human Leishmania infections cause cutaneous lesions, with a minority of L. (Viannia) infections leading to mucocutaneous leishmaniasis. Visceral leishmaniasis and diffuse cutaneous leishmaniasis are both rare. In each country a significant proportion of Leishmania transmission is in or around houses, often close to coffee or cacao plantations. Reservoir hosts for domestic transmission cycles are uncertain. The paper first addresses the burden of disease caused by leishmaniasis, focusing on both incidence rates and on the variability in symptoms. Such information should provide a rational basis for prioritizing control resources, and for selecting therapy regimes. Secondly, we describe the variation in transmission ecology, outlining those variables which might affect the prevention strategies. Finally, we look at the current control strategies and review the recent studies on control.

Identification of six Trypanosoma cruzi phylogenetic lineages by random amplified polymorphic DNA and multilocus enzyme electrophoresis

Genetic characterisation of Trypanosoma cruzi variants is of foremost importance, due to considerable genetic and biological heterogeneity in the parasite populations. Two major phylogenetic lineages, each highly heterogeneous, have been previously described within this species. Here we characterised a geographically and ecologically diverse sample of stocks representative of the breadth of the known clonal diversity of each major lineage, using random amplified polymorphic DNA with 20 primers and multilocus enzyme electrophoresis at 22 loci. Molecular hybridisation experiments were performed to control the homology of randomly amplified DNA markers. Both sets of data were highly consistent and supported the existence of two major lineages. Additionally, we found that lineage 2 appeared further partitioned into five sharply delineated phylogenetic clusters, each comprising one of the following reference strains: CanIII cl1 (Z3 reference), M5631 cl5, Esmeraldo cl3 (Z2 reference), CL Brener, and MN cl2. The two first clusters were found mainly in sylvatic environments, whereas the three latter were restricted to domestic transmission cycles and were only collected South to the Amazon Basin. In contrast, lineage 1, which included Miles' Z1 reference strain X10 cl1, was not further subdivided and was encountered across the entire endemic area, in both domestic and sylvatic cycles. Thus, T. cruzi appeared to be subdivided into six discrete typing units, or DTUs, exhibiting distinct geographic and ecological ranges. Reliable diagnostic markers for the two major lineages and the five smaller DTUs of lineage 2 are described, and correspondence with previous classifications of T. cruzi genotypes is given in order to help communication on T. cruzi phylogenetic diversity.

Prevalence of Trypanosoma cruzi infection in opossum (Didelphis albiventris) in Santiago del Estero, Argentina

The opossum of the genus Didelphis is one of the principal wild reservoirs of Trypanosoma cruzi and is widely distributed in the Western Hemisphere. Didelphis albiventris is the most common marsupial in Amamá and Trinidad, two communities in the province of Santiago del Estero, Argentina. The D. albiventris population is replaced every year, and the opossum normally has two reproductive periods, one at the beginning of the spring and another at the beginning of the summer. The two litters are weaned, and they leave the mother's marsupial pouch to join the population, the first (G1) at the beginning of the summer and the second (G2) at the beginning of the fall. Between 1988 and 1991 409 D. albiventris opossums were studied, and xenodiagnoses showed that 35% of them were infected with T. cruzi. Annual cycles of renewed infection were observed, with prevalences that ranged between 22% and 43%. The acquisition of the parasite occurred over the entire year, from the summer through the spring. The prevalence of infection increased with age. The G1 individuals tended to present higher prevalences than the G2 individuals, probably from being exposed to transmission for a longer period of time. In the first two (younger) age categories for the opossums, G2 individuals showed higher prevalences than did the G1 individuals. This indicates a significant increase in transmission intensity during the fall. Opossums should be regarded as a potential source of T. cruzi entry to the domestic transmission cycle.