High Trypanosoma cruzi prevalence in armadillo (Zaedyus pichiy; Xenarthra: Chlamyphoridae) populations from Mendoza, Argentina.

Armadillos are considered important reservoir hosts for Trypanosoma cruzi, the causative agent of Chagas disease. The first report of T. cruzi infection in pichis (Zaedyus pichiy), a small armadillo species endemic to central Argentina and Chile, dates back to 1935. However, more recent reports on T. cruzi in this species are scarce. The objective of this study was to assess T. cruzi infection and parasite load in Z. pichiy from Mendoza Province, an area endemic to human Chagas disease. Blood samples were obtained in 2014-2016 from pichis from Lavalle (low Monte), Malargüe (Patagonian steppe), and San Carlos (ecotone) departments, Mendoza Province, Argentina. The detection and quantification of T. cruzi was performed through qPCR amplification using satellite primers. Of the 265 analyzed samples, 201 (76%) were positive for T. cruzi. Parasite loads varied between < 0.1-55.8 parasite-equivalents/mL (par-eq/mL), with a median of 1.1 par-eq/mL in quantifiable samples. The prevalence was similar in Malargüe and Lavalle (85-94%), but significantly lower in pichis from San Carlos (50%). Animals from Lavalle captured after hibernation had significantly higher parasite loads (median 2.0 par-eq/mL). In Malargüe, T. cruzi infection and parasite loads were significantly lower before than after hibernation in 2016. The high prevalence and low median parasite load suggest a chronic and persistent infection of T. cruzi in pichis. Regional differences and a marked increase in precipitation during 2015-2016 could have influenced annual and seasonal infection rates of this vector-borne disease.

Co-infection patterns of vector-borne zoonotic pathogens in owned free-ranging dogs in central Chile.

We investigated the co-occurrence of the nine of the most relevant canine vector-borne pathogens (CVBP) using conventional and real-time PCR and evaluated risk factors and potential non-apparent haematological alterations associated with co-infection in 111 rural, owned, free-ranging dogs in the Metropolitan Region of Chile.At least one pathogen was detected in 75% of the dogs. DNA of Anaplasma platys (Ap; 36%), Candidatus Mycoplasma haematoparvum (CMhp; 31%), Mycoplasma haemocanis (Mhc; 28%), Trypanosoma cruzi (17%), Leishmania spp. (4.5%), and Acanthocheilonema reconditum (1%) was detected. All dogs were negative for Ehrlichia spp., Rickettsia spp., Bartonella spp., Piroplasmida, and Hepatozoon spp. Thirty-eight dogs (34%) were coinfected. CMhp was involved in 71%, Mhc in 58%, and Ap in 50% of the co-infections. The most common co-infection pattern was CMhp-Mhc (37% of the cases). The prevalence of Ap was higher in juvenile than in adult dogs, whereas the opposite was found for CMhp and Mhc. Adult dogs were four times more likely of being co-infected than juveniles. Co-infected animals showed higher white blood cell count, segmented neutrophil count, and GGT levels than non-co-infected dogs. Clinically healthy but infected dogs may act as reservoirs of CVBP, and their free-ranging behavior would facilitate the spread of these pathogens to other dogs as well as human beings or wild carnivores.

Discrete Typing Units of Trypanosoma cruzi Identified by Real-Time PCR in Peripheral Blood and Dejections of Triatoma infestans Used in Xenodiagnosis Descriptive Study.

Chagas disease (ChD) is a vector zoonosis native to the American continent caused by the protozoan parasite Trypanosoma cruzi; the biological vectors are multiple species of hematophagous insects of the family Triatominae. A relevant aspect in the host-parasite relationship is the identification of the various genotypes of T. cruzi called discrete typing units (DTU) that circulate in mammals and vectors. In Chile, it has been described that the DTUs TcI, TcII, TcV, and TcVI circulate in infected humans, vectors, and wild animals. Identifying DTUs has acquired clinical importance, since it has been suggested that different genotypes could cause distinct pathologies, circulate in different geographical areas, and present different sensitivities to trypanocidal drugs. In this study, circulating T. cruzi DTUs in peripheral blood and Triatoma infestans dejections used in xenodiagnosis (XD) were amplified by qPCR in 14 Chilean patients with chronic ChD from highly endemic areas. More positive samples were detected by XD compared to peripheral blood samples, and 64.28% of the cases were simple infections and 35.72% mixed, with a statistically significant difference in the frequency of TcV DTU. This study would suggest that T. infestans from Chile is more competent to amplify one DTU over others, probably due to a process of co-evolution.

The Parasite Load of Trypanosoma cruzi Modulates Feeding and Defecation Patterns of the Chagas Disease Vector Triatoma infestans.

Trypanosoma cruzi is the causal agent of Chagas disease, a parasitic zoonosis transmitted mainly through the feces of triatomine insects. Triatoma infestans is the main triatomine vector of this disease in South America. Previous research has shown that T. cruzi infection modifies the behavior of triatomines. We evaluated, for the first time, the effect of parasite load on feeding and defecation behavior, which we quantified by using real-time PCR. The detection time of the host was shorter in infected individuals, and the number of bites increased, while the dejection time was reduced when compared with the non-infected group. A significant correlation between the parasite load and the behavioral changes registered in the infected triatomines was found. These results would indicate that the intensity of T. cruzi infection modulates the feeding and defecation behavior of T. infestans, increasing the vector competence of this triatomine vector.

Trypanosoma cruzi infection in the wild Chagas disease vector, Mepraia spinolai: Parasitic load, discrete typing units, and blood meal sources.

BACKGROUND: Mepraia spinolai, a wild vector of Trypanosoma cruzi in Chile, is an abundant triatomine species that is frequently infected by the parasite that causes Chagas disease. The aim of this study was to determine if the parasitic load of T. cruzi in M. spinolai is related to its blood meal source and the infecting DTUs of T. cruzi. METHODS: The vector was captured in rural areas. In the laboratory, DNA was extracted from its abdomen and T. cruzi was quantified using qPCR. Real time PCR assays for four T. cruzi DTUs were performed. Blood meal sources were identified by real-time PCR amplification of vertebrate cytochrome b gene sequences coupled with high resolution melting (HRM). RESULTS: Trypanosoma cruzi was detected in 735 M. spinolai; in 484 we identified one blood meal source, corresponding to human, sylvatic, and domestic species. From these, in 224 we were able to discriminate the infecting DTU. When comparing the parasitic loads between the unique blood meal sources, no significant differences were found, but infections with more than one DTU showed higher parasitic loads than single infections. DTU TcI was detected in a high proportion of the samples. CONCLUSIONS: Higher parasitic loads are related to a greater number of T. cruzi DTUs infecting M. spinolai, and this triatomine seems to have a wide span of vertebrate species in its diet.

Trypanosoma cruzi Parasite Load Modulates the Circadian Activity Pattern of Triatoma infestans.

American trypanosomiasis is a disease caused by the flagellate protozoan Trypanosoma cruzi, which is transmitted mainly in endemic areas by blood-sucking triatomine vectors. Triatoma infestans is the most important vector in the southern cone of South America, exhibiting a nocturnal host-seeking behavior. It has been previously documented that the parasite produces changes in some triatomine species, but this is the first time that the behavior of a vector has been evaluated in relation to its parasite load. After comparing the movement events and distance traveled of infected and non-infected T. infestans, we evaluated the change produced by different T. cruzi parasite loads on its circadian locomotor activity. We observed differences between infected and non-infected triatomines, and a significant relation between the parasite load and the increase in locomotor activity of T. infestans, which was accentuated during the photophase. This could have direct implications on the transmission of T. cruzi, as the increased movement and distance traveled could enhance the contact of the vector with the host, while increasing the predation risk for the vector, which could both constitute a risk for vectorial and oral transmission to mammals.

Trypanosoma cruzi-infected triatomines and rodents co-occur in a coastal island of northern Chile.

Trypanosoma cruzi, the cause agent of Chagas disease, is transmitted mainly by blood-feeding insects of the subfamily Triatominae. The T. cruzi life cycle alternates between triatomines and mammalian hosts, excluding birds and reptiles. Triatomines of Mepraia genus are wild vectors of T. cruzi in Chile. Mepraia specimens infected with T. cruzi have been detected in Pan de Azúcar and Santa María islands. The most common vertebrates that inhabit these islands are birds and reptiles, and it is unknown whether small mammals are present. Consequently, it is relevant to know whether there are any T. cruzi-infected small mammals on those islands to elucidate the T. cruzi cycle. To clarify this crossroads, islands of northern Chile were explored to determine if T. cruzi-infected triatomines and rodents co-occur in islands of northern Chile. T. cruzi DNA was detected by conventional and real-time PCR in three islands: on Santa María and Pan de Azúcar islands T. cruzi was detected in Mepraia sp samples, while on Pan de Azúcar (6.1%) and Damas islands (15%) was detected in the rodent Abrothrix olivacea. We show for the first time in Chile the occurrence of insular rodents infected with T. cruzi, and a complete T. cruzi life cycle in a coastal island. Our results provide new insights to understand the T. cruzi infection in the wild cycle.

High rates of Trypanosoma cruzi infection in goats from Mendoza province, Argentina: Parasite loads in blood and seasonal variation.

Mendoza province, in central west Argentina, is considered among the high-risk provinces for vector transmission of Trypanosoma cruzi, the causative agent of Chagas disease. Extensive goat farming is common in large parts of rural Mendoza, and goats may act as a reservoir for this parasite. The objective of this study was to determine infection rates, parasite loads, and seasonal variation of these parameters in T. cruzi infection in goats from rural areas of three departments of Mendoza. A total of 349 peripheral blood samples with EDTA / guanidine were analyzed from goats on 11 farms (three in Lavalle, three in San Carlos, and five in Malargüe department) in spring of 2014, 2015, and 2016; and in fall of 2015 and 2016 (only Malargüe). DNA was extracted using a Phenol: Chloroform: Isoamyl protocol. The detection and quantification of T. cruzi was performed through qPCR amplification using satellite oligonucleotides. Of the 349 blood samples, 267 (77%) were positive, with parasite loads ranging between <0.10 and 10.90 par-eq/mL (median 0.10). In spring, frequencies of infection in the three departments ranged between 86% and 95%, but differences were not significant. Median parasite loads were higher in Lavalle than in the other departments, while those in goats from San Carlos were consistently low. The frequency of infection and parasite loads in Malargüe were significantly higher in spring than in fall. This seasonal variation may have been related to a reduced nutritional status and impaired immune response of goats in spring. In conclusion, the high proportion of positive goats confirms the persistence of T. cruzi in rural Mendoza.

Course of serological tests in treated subjects with chronic Trypanosoma cruzi infection: A systematic review and meta-analysis of individual participant data.

OBJECTIVE: To determine the course of serological tests in subjects with chronic Trypanosoma cruzi infection treated with anti-trypanosomal drugs. METHODS: A systematic review and meta-analysis was conducted using individual participant data. Survival analysis and the Cox proportional hazards regression model with random effects to adjust for covariates were applied. The protocol was registered in the PROSPERO database (http://www.crd.york.ac.uk/PROSPERO; CRD42012002162). RESULTS: A total of 27 studies (1296 subjects) conducted in eight countries were included. The risk of bias was low for all domains in 17 studies (63.0%). Nine hundred and thirteen subjects were assessed (149 seroreversion events, 83.7% censored data) for enzyme-linked immunosorbent assay (ELISA), 670 subjects (134 events, 80.0% censored) for indirect immunofluorescence assay (IIF), and 548 subjects (99 events, 82.0% censored) for indirect hemagglutination assay (IHA). A higher probability of seroreversion was observed within a shorter time span in subjects aged 1-19 years compared to adults. The chance of seroreversion also varied according to the country where the infection might have been acquired. For instance, the pooled adjusted hazard ratio between children/adolescents and adults for the IIF test was 1.54 (95% confidence interval 0.64-3.71) for certain countries of South America (Argentina, Bolivia, Chile, and Paraguay) and 9.37 (95% confidence interval 3.44-25.50) for Brazil. CONCLUSIONS: The disappearance of anti-T. cruzi antibodies was demonstrated along the course of follow-up. An interaction between age at treatment and country setting was found.

Discrete typing units of Trypanosoma cruzi detected by real-time PCR in Chilean patients with chronic Chagas cardiomyopathy.

Chagas disease is a major public health problem in Latin America and has spread to other countries due to immigration of infected persons. 10-30% of patients with chronic Chagas disease will develop cardiomyopathy. Chagas cardiomyopathy is the worst form of the disease, due to its high morbidity and mortality. Because of its prognostic value and adequate medical monitoring, it is very important to identify infected people who could develop Chagas cardiomyopathy. The aim of this study was to determine if discrete typing units (DTUs) of Trypanosoma cruzi are related to the presence of heart disease in patients with chronic Chagas disease. A total of 86 untreated patients, 41 with cardiomyopathy and 45 without heart involvement were submitted to clinical study. Electrocardiograms and echocardiograms were performed on the group of cardiopaths, in which all important known causes of cardiomyopathy were discarded. Sinus bradycardia and prolonged QTc interval were the most frequent electrocardiographic alterations and patients were classified in group I (46%) and group II (54%) of New York Hearth Association. In all cases real-time PCR genotyping assays were performed. In the group with cardiomyopathy, the most frequent DTU was TcI (56.1%), followed by TcII (19.5%). Mixed infections TcI + TcII were observed in 7.3% of the patients. In the group without cardiac pathologies, TcI and TcII were found at similar rates (28.9 and 31.1%, respectively) and mixed infections TcI + TcII in 17.8% of the cases. TcIII and TcIV were not detected in any sample. Taken together, our data indicate that chronic Chagas cardiomyopathy in Chile can be caused by strains belonging to TcI and TcII.

Trypanosoma cruzi load in synanthropic rodents from rural areas in Chile.

BACKGROUND: Trypanosoma cruzi is the agent of Chagas disease, a major public health problem in Latin America. Many wild and domestic animals are naturally infected with T. cruzi; rodents are one of the groups which have been consistently detected infected in different countries. The aim of this work was to characterize blood T. cruzi load in naturally infected rodents from a Chagas disease endemic region in Chile. METHODS: Baited traps were set in domestic and peridomestic areas of rural dwellings. The rodents were anesthetized and blood sampled; DNA was extracted and the parasite load was quantified by T. cruzi satellite DNA real-time PCR assays. RESULTS: Seventy-one rodents of four species, Rattus rattus, Mus musculus, Phyllotis darwini and Octodon degus, were captured; R. rattus was the most abundant species. Fifty-nine samples (83.1%) were T. cruzi-positive and the median value of the parasite load was 2.99 parasite equivalents (par-eq)/ml. The comparison of frequency of infection or parasite load by species showed no differences. However, one R. rattus presented very elevated parasitemia (1644 par-eq/ml). CONCLUSIONS: The overall levels of parasitemia were similar to those found in humans in Chile. The high infection levels in exotic and endemic rodents very near to rural settlements increases their relevance as T. cruzi hosts.

Real-time PCR strategy for the identification of Trypanosoma cruzi discrete typing units directly in chronically infected human blood.

The protozoan Trypanosoma cruzi is the causative agent of Chagas disease, a major public health problem in Latin America. This parasite has a complex population structure comprised by six or seven major evolutionary lineages (discrete typing units or DTUs) TcI-TcVI and TcBat, some of which have apparently resulted from ancient hybridization events. Because of the existence of significant biological differences between these lineages, strain characterization methods have been essential to study T. cruzi in its different vectors and hosts. However, available methods can be laborious and costly, limited in resolution or sensitivity. In this study, a new genotyping strategy by real-time PCR to identify each of the six DTUs in clinical blood samples have been developed and evaluated. Two nuclear (SL-IR and 18S rDNA) and two mitochondrial genes (COII and ND1) were selected to develop original primers. The method was evaluated with eight genomic DNA of T. cruzi populations belonging to the six DTUs, one genomic DNA of Trypanosoma rangeli, and 53 blood samples from individuals with chronic Chagas disease. The assays had an analytical sensitivity of 1-25fg of DNA per reaction tube depending on the DTU analyzed. The selectivity of trials with 20fg/µL of genomic DNA identified each DTU, excluding non-targets DTUs in every test. The method was able to characterize 67.9% of the chronically infected clinical samples with high detection of TcII followed by TcI. With the proposed original genotyping methodology, each DTU was established with high sensitivity after a single real-time PCR assay. This novel protocol reduces carryover contamination, enables detection of each DTU independently and in the future, the quantification of each DTU in clinical blood samples.