Lizards as Silent Hosts of Trypanosoma cruzi.

We assessed 4 lizard species in Chile for Trypanosoma cruzi, the causative agent of Chagas disease, and 1 species for its ability to transmit the protozoan to uninfected kissing bugs. All lizard species were infected, and the tested species was capable of transmitting the protozoan, highlighting their role as T. cruzi reservoirs.

Phylogenetic incongruence inferred with two mitochondrial genes in Mepraia spp. and Triatoma eratyrusiformis(Hemiptera, Reduviidae).

Mitochondrial DNA (mtDNA) is widely used to clarify phylogenetic relationships among and within species, and to determine population structure. Due to the linked nature of mtDNA genes it is expected that different genes will show similar results. Phylogenetic incongruence using mtDNA genes may result from processes such as heteroplasmy, nuclear integration of mitochondrial genes, polymerase errors, contamination, and recombination. In this study we used sequences from two mitochondrial genes (cytochrome b and cytochrome oxidase subunit I) from the wild vectors of Chagas disease, Triatoma eratyrusiformis and Mepraia species to test for topological congruence. The results showed some cases of phylogenetic incongruence due to misplacement of four haplotypes of four individuals. We discuss the possible causes of such incongruence and suggest that the explanation is an intra-individual variation likely due to heteroplasmy. This phenomenon is an independent evidence of common ancestry between these taxa.

Phylogenetic Analyses of Lizards from the Chilean Humboldt Archipelago Reveal a New Species for the Chañaral Island (Squamata: Liolaemidae).

The Humboldt Archipelago, situated on Chile's north-central coast, is renowned for its exceptional biodiversity. However, lizards of the Liolaemus genus are a particularly understudied group in this archipelago. Liolaemus genus is divided into two clades: chiliensis and nigromaculatus. Within the nigromaculatus clade the zapallarensis group is restricted to the semi-arid and arid coastal habitats of the Atacama Desert in north-central Chile. While it has been reported that lizards from the zapallarensis group inhabit various islands within the Humboldt Archipelago, there has been limited knowledge regarding their specific species identification. To identify the lizard species inhabiting these islands, we conducted phylogenetic analyses using a mitochondrial gene and examined morphological characteristics. Our findings reveal that lizards from the Damas, Choros, and Gaviota islands belong to Liolaemus silvai. In contrast, the lizards on Chañaral Island form a distinct and previously unrecognised group, clearly distinguishable from Liolaemus silvai. In conclusion, our study not only confirms the presence of L. silvai on the Damas, Choros, and Gaviota islands but also describes a new lizard species on Chañaral Island named Liolaemus carezzae sp. nov. These findings contribute valuable insights into the biodiversity of these islands and introduce a newly discovered endemic taxon to the region, enriching our understanding of Chile's unique island ecosystems.

Humans as blood-feeding sources in sylvatic triatomines of Chile unveiled by next-generation sequencing.

BACKGROUND: Triatomines are blood-sucking insects capable of transmitting Trypanosoma cruzi, the parasite that causes Chagas disease in humans. Vectorial transmission entails an infected triatomine feeding on a vertebrate host, release of triatomine infective dejections, and host infection by the entry of parasites through mucous membranes, skin abrasions, or the biting site; therefore, transmission to humans is related to the triatomine-human contact. In this cross-sectional study, we evaluated whether humans were detected in the diet of three sylvatic triatomine species (Mepraia parapatrica, Mepraia spinolai, and Triatoma infestans) present in the semiarid-Mediterranean ecosystem of Chile. METHODS: We used triatomines collected from 32 sites across 1100 km, with an overall T. cruzi infection frequency of 47.1% (N = 4287 total specimens) by conventional PCR or qPCR. First, we amplified the vertebrate cytochrome b gene (cytb) from all DNA samples obtained from triatomine intestinal contents. Then, we sequenced cytb-positive PCR products in pools of 10-20 triatomines each, grouped by site. The filtered sequences were grouped into amplicon sequence variants (ASVs) with a minimum abundance of 100 reads. ASVs were identified by selecting the best BLASTn match against the NCBI nucleotide database. RESULTS: Overall, 16 mammal (including human), 14 bird, and seven reptile species were identified in the diet of sylvatic triatomines. Humans were part of the diet of all analyzed triatomine species, and it was detected in 19 sites representing 12.19% of the sequences. CONCLUSIONS: Sylvatic triatomine species from Chile feed on a variety of vertebrate species; many of them are detected here for the first time in their diet. Our results highlight that the sylvatic triatomine-human contact is noteworthy. Education must be enforced for local inhabitants, workers, and tourists arriving in endemic areas to avoid or minimize the risk of exposure to Chagas disease vectors.

Testing Phylogeographic Hypotheses in Mepraia (Hemiptera: Reduviidae) Suggests a Complex Spatio-Temporal Colonization in the Coastal Atacama Desert.

Mepraia is a genus (Triatominae) endemic to Chile and a vector of Trypanosoma cruzi. Alternative phylogeographic hypotheses have been suggested for Mepraia. We tested different colonization routes hypothesized using mitochondrial sequences and phylogeographic approaches to select the best-supported hypothesis. Our results suggest that, after the split from the sister genus Triatoma at ~4.3 Mya, Mepraia formed two main clades at ~2.1 Mya. The northern clade diverged from Mepraia sp. ~1.7 Mya, giving rise to M. parapatrica and M. gajardoi about ~1.4 Mya. The southern clade originated M. spinolai ~1.68 Mya. We suggest that Mepraia had an origin in the north-central Andes along with orogenic processes, reinforced by hyperaridity during the Pliocene. The hyperarid cycle would have separated the southern and northern clades. Then, in the northern clade, dispersal occurred north and south from the centre through corridors during the Pleistocene Climatic Oscillations. Climate changes may have induced a major speciation process in the Atacama Desert, while the more homogeneous habitat colonized by the southern clade led to only one, but structured, species.

Blood-Meal Sources and Trypanosoma cruzi Infection in Coastal and Insular Triatomine Bugs from the Atacama Desert of Chile.

Mepraia parapatrica is one of the lesser known and less abundant sylvatic triatomine species naturally infected by the protozoan Trypanosoma cruzi, the etiological agent of Chagas disease. M. parapatrica lives in sympatry with T. cruzi-infected rodents, but only birds, reptiles, and marine mammals have been reported as blood-meal sources of this vector species by serology. The distribution range of this kissing bug overlaps with fishers' settlements and tourist areas, and therefore the study of the blood-meal sources of this triatomine species is relevant. Here, we determined the blood-meal sources of M. parapatrica by NGS or standard sequencing from a coastal mainland area and an island in northern Chile, and T. cruzi infection by real-time PCR. The blood-meals of. M parapatrica included 61.3% reptiles, 35.5% mammals (including humans) and 3.2% birds. Feeding on reptiles was more frequent on the mainland, while on the island feeding on mammals was more frequent. The presence of T. cruzi-infected triatomine bugs and humans as part of the diet of M. parapatrica in both areas represents an epidemiological threat and potential risk to the human population visiting or established in these areas. Currently there are no tools to control wild triatomines; these results highlight the potential risk of inhabiting these areas and the necessity of developing information campaigns for the community and surveillance actions.

Trypanosoma cruzi DNA in Desmodus rotundus (common vampire bat) and Histiotus montanus (small big-eared brown bat) from Chile.

The protozoan Trypanosoma cruzi, the causative agent of Chagas disease, is transmitted by infected feces or consumption of blood-sucking triatomine insects to several mammalian orders including Chiroptera. In Chile, the distribution of several insectivorous and one hematophagous bat species overlaps with those of triatomine vectors, but the T. cruzi infection status of local chiropterans is unknown. In 2018, we live-captured bats from two protected areas in Chile to collect plagiopatagium tissue, feces and perianal swab samples, in search for T. cruzi-DNA by real time PCR assays using species-specific primers. In Pan de Azúcar island (∼26°S), we examined a roost of Desmodus rotundus (common vampire bat) and sampled tissue from 17 individuals, detecting T. cruzi-DNA in five of them. In Las Chinchillas National Reserve (∼31°S), we examined two roosts of Histiotus montanus (small big-eared brown bat), collecting feces or perianal swab samples from eight individuals, detecting T. cruzi-DNA in four of them. This is the first report of T. cruzi-DNA evidence in bat species from Chile. Both vector-borne and oral transmission are potential infection routes that can explain our results. Further investigation is needed for a better understanding of the role of bats in the T. cruzi transmission cycle.

Biogeographic origin and phylogenetic relationships of Mepraia (Hemiptera, Reduviidae) on islands of northern Chile.

Chagas disease is one of the main zoonoses mediated by vectors in America. The etiological agent is the protozoan Trypanosoma cruzi, transmitted mainly by hematophagous insects of the subfamily Triatominae. Mepraia species are triatomines endemic to Chile that play an important role in T. cruzi transmission in the wild cycle and are potential vectors for humans. In addition to the continental distribution, populations of Mepraia genus have been reported inhabiting islands of northern Chile. The presence of individuals of Mepraia in insular areas might be explained through passive dispersion by marine birds or by vicariance of an ancestral widespread population. To clarify the biogeographic origin and phylogenetic relationships of island individuals of Mepraia, mitochondrial COI and cyt b genes were sequenced in individuals from island and continental areas. Gene sequences were used to estimate phylogenetic relationships, divergence dates and migration rates between insular and continental populations. The dates of divergence estimates are congruent with sea level and tectonic changes that originated the islands during Pleistocene. Migration rates suggest symmetric historical island-continent gene flow. We suggest that the origin of island triatomines can be explained by both vicariance and dispersion. Phylogenetic relationships show that individuals from Santa María Island and the continent clustered in a clade different from those previously reported, indicating a new lineage of Mepraia genus. This study will contribute to understand the origin of the T. cruzi infection in coastal islands of northern Chile.

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.

Trypanosoma cruzi over the ocean: Insular zones of Chile with presence of infected vector Mepraia species.

Chagas disease is one of the main zoonosis mediated by vectors in America. The etiologic agent Trypanosoma cruzi infects different mammals and is transmitted principally by the subfamily Triatominae. Mepraia is a genus endemic to Chile, responsible for transmitting T. cruzi in the sylvatic cycle. Mepraia includes three species: M. gajardoi and M. parapatrica inhabit coastal areas, while M. spinolai inhabits coastal and interior valleys. Previous studies reported the occurrence of Mepraia in Pan de Azucar Island, currently classified as M. parapatrica, but T. cruzi has not been reported in these insects. It is suggested that this could be due to infrequent insect feeding on mammalian hosts. In order to detect T. cruzi in insects from coastal islands, specimens from Pan de Azucar and Santa Maria Islands were examined. A region of kDNA of T. cruzi was amplified by PCR and hybridization assays were performed for T. cruzi genotyping of insect feces. The presence of infected insect and mixed T. cruzi infections was demonstrated. This is the first report of infected Triatominae in coastal islands in Chile. We discuss T. cruzi detection in insular zones, and the presumptive reservoirs that may participate in maintaining its transmission cycle in this habitat. Mixed and unidentified infections suggest that there are complex and unknown reservoir interactions in these habitats.

Experimental crosses between Mepraia gajardoi and M. spinolai and hybrid chromosome analyses reveal the occurrence of several isolation mechanisms.

Hematophagous insects of the subfamily Triatominae include several species with a large variety of shapes, behavior and distribution. They have great epidemiological importance since most of them transmit the flagellated protozoan Trypanosoma cruzi, the etiologic agent of Chagas disease. In this subfamily several cases of species hybridization have been reported under experimental and natural conditions. Mepraia is a genus of Triatominae endemic in Chile, responsible for transmitting T. cruzi in the sylvatic cycle. This genus includes three species, M. gajardoi, M. spinolai and M. parapatrica; however, the differentiation of M. parapatrica as a separate species remains controversial considering the possible occurrence of introgression/hybridization processes in some populations of this putative species. Mepraia species show conspicuous wing polymorphism, and it has been proposed that the genes related to wings are linked to the Y chromosome, thus wingless males could not engender winged progeny. In order to determine the degree of reproductive isolation and to assess the wing phenotype in the offspring, we performed experimental crosses between the two most divergent Mepraia species (M. gajardoi and M. spinolai) together with chromosome analyses of hybrid progenies. Although fertile F1 hybrids were obtained in only one direction of crossing, we verified the existence of different isolation mechanisms between parental species, including hybrid breakdown. The occurrence of winged males in the offspring of wingless parental males suggests that the wing character is not linked to the Y chromosome.

Interactions Between Trypanosoma cruzi the Chagas Disease Parasite and Naturally Infected Wild Mepraia Vectors of Chile.

Chagas disease, which ranks among the world's most neglected diseases, is a chronic, systemic, parasitic infection caused by the protozoan Trypanosoma cruzi. Mepraia species are the wild vectors of this parasite in Chile. Host-parasite interactions can occur at several levels, such as co-speciation and ecological host fitting, among others. Thus, we are exploring the interactions between T. cruzi circulating in naturally infected Mepraia species in all areas endemic of Chile. We evaluated T. cruzi infection rates of 27 different haplotypes of the wild Mepraia species and identified their parasite genotypes using minicircle PCR amplification and hybridization tests with genotype-specific DNA probes. Infection rates were lower in northern Chile where Mepraia gajardoi circulates (10-35%); in central Chile, Mepraia spinolai is most abundant, and infection rates varied in space and time (0-55%). T. cruzi discrete typing units (DTUs) TcI, TcII, TcV, and Tc VI were detected. Mixed infections with two or more DTUs are frequently found in highly infected insects. T. cruzi DTUs have distinct, but not exclusive, ecological and epidemiological associations with their hosts. T. cruzi infection rates of M. spinolai were higher than in M. gajardoi, but the presence of mixed infection with more than one T. cruzi DTU was the same. The same T. cruzi DTUs (TcI, TcII, TcV, and TcVI) were found circulating in both vector species, even though TcI was not equally distributed. These results suggest that T. cruzi DTUs are not associated with any of the two genetically related vector species nor with the geographic area. The T. cruzi vectors interactions are discussed in terms of old and recent events. By exploring T. cruzi DTUs present in Mepraia haplotypes and species from northern to central Chile, we open the analysis on these invertebrate host-parasite interactions.