Morphological and molecular characteristics of a Trypanosoma sp. from triatomines (Triatoma rubrofasciata) in China.

BACKGROUND: Triatomines (kissing bugs) are natural vectors of trypanosomes, which are single-celled parasitic protozoans, such as Trypanosoma cruzi, T. conorhini and T. rangeli. The understanding of the transmission cycle of T. conorhini and Triatoma rubrofasciata in China is not fully known. METHODS: The parasites in the faeces and intestinal contents of the Tr. rubrofasciata were collected, and morphology indices were measured under a microscope to determine the species. DNA was extracted from the samples, and fragments of 18S rRNA, heat shock protein 70 (HSP70) and glycosomal glyceraldehyde-3-phosphate dehydrogenase (gGAPDH) were amplified and sequenced. The obtained sequences were then identified using the BLAST search engine, followed by several phylogenetic analyses. Finally, laboratory infections were conducted to test whether Tr. rubrofasciata transmit the parasite to rats (or mice) through bites. Moreover, 135 Tr. rubrofasciata samples were collected from the Guangxi region and were used in assays to investigate the prevalence of trypanosome infection. RESULTS: Trypanosoma sp. were found in the faeces and intestinal contents of Tr. rubrofasciata, which were collected in the Guangxi region of southern China and mostly exhibited characteristics typical of epimastigotes, such as the presence of a nucleus, a free flagellum and a kinetoplast. The body length ranged from 6.3 to 33.9 µm, the flagellum length ranged from 8.7 to 29.8 µm, the nucleus index was 0.6 and the kinetoplast length was -4.6. BLAST analysis revealed that the 18S rRNA, HSP70 and gGAPDH sequences of Trypanosoma sp. exhibited the highest degree of similarity with those of T. conorhini (99.7%, 99.0% and 99.0%, respectively) and formed a well-supported clade close to T. conorhini and T. vespertilionis but were distinct from those of T. rangeli and T. cruzi. Laboratory experiments revealed that both rats and mice developed low parasitaemia after inoculation with Trypanosoma sp. and laboratory-fed Tr. rubrofasciata became infected after feeding on trypanosome-positive rats and mice. However, the infected Tr. rubrofasciata did not transmit Trypanosoma sp. to their offspring. Moreover, our investigation revealed a high prevalence of Trypanosoma sp. infection in Tr. rubrofasciata, with up to 36.3% of specimens tested in the field being infected. CONCLUSIONS: Our study is the first to provide a solid record of T. conorhini from Tr. rubrofasciata in China with morphological and molecular evidence. This Chinese T. conorhini is unlikely to have spread through transovarial transmission in Tr. rubrofasciata, but instead, it is more likely that the parasite is transmitted between Tr. rubrofasciata and mice (or rats). However, there was a high prevalence of T. conorhini in the Tr. rubrofasciata from our collection sites and numerous human cases of Tr. rubrofasciata bites were recorded. Moreover, whether these T. conorhini strains are pathogenic to humans has not been investigated.

Detection of Trypanosoma lewisi DNA from Rattus norvegicus and Rattus rattus in Hanoi, Vietnam.

Trypanosoma lewisi is a worldwide nonpathogenic parasite that is exclusively found in rats. In general, T. lewisi infection in humans is an opportunistic infection from rats to humans through fleas. However, recently, infection with T. lewisi in humans, including a fatal case, has been reported. Notably, rats living close to a human settlement showed a higher prevalence of infection with T. lewisi than those living in other places. It is possible that the urbanization is associated with the prevalence of T. lewisi in rats and enhances the risk of T. lewisi transmission to humans through fleas. In this study, a total of 88 rats were captured from hospitals, markets, and a cargo station, of which 81 were identified as Rattus norvegicus and 7 as Rattus rattus in Hanoi, the urbanizing city of Vietnam. Of these, 55 rats (62.5%) harbored T. lewisi, of which 52 were R. norvegicus and 3 were R. rattus.

'A flying start': Wildlife trypanosomes in tissues of Australian tabanids (Diptera: Tabanidae).

Tabanids (syn. horse flies) are biting-flies of medical and veterinary significance because of their ability to transmit a range of pathogens including trypanosomes - some species of which carry a combined health and biosecurity risk. Invertebrate vectors responsible for transmitting species of Trypanosoma between Australian wildlife remains unknown, thus establishing the role of potential vector candidates such as tabanids is of utmost importance. The current study aimed to investigate the presence of indigenous trypanosomes in tabanids from an endemic area of south-west Australia. A total of 148 tabanids were collected, with morphological analysis revealing two subgenera: Scaptia (Pseudoscione) and S. (Scaptia) among collected flies. A parasitological survey using an HRM-qPCR and sequencing approach revealed a high (105/148; 71%) prevalence of trypanosomatid DNA within collected tabanids. Individual tissues - proboscis (labrum, labium and mandibles, hypopharynx), salivary glands, proventriculus, midgut, and hindgut and rectum - were also tested from a subset of 20 tabanids (n = 140 tissues), confirming the presence of Trypanosoma noyesi in 31% of screened tissues, accompanied by T. copemani (3%) and T. vegrandis/T.gilletti (5%). An unconfirmed trypanosomatid sp. was also detected (9%) within tissues. The difference between tissues infected with T. noyesi compared with tissues infected with other trypanosome species was statistically significant (p < 0.05), revealing T. noyesi as the more frequent species detected in the tabanids examined. Fluorescence in situ hybridisation (FISH) and scanning electron microscopy (SEM) confirmed intact parasites within salivary glands and the proboscis respectively, suggesting that both biological and mechanical modes of transmission could occur. This study reveals the presence of Australian Trypanosoma across tabanid tissues and confirms intact parasites within tabanid salivary glands and the proboscis for the first time. Further investigations are required to determine whether tabanids have the vectorial competence to transmit Australian trypanosomes between wildlife.

Effect of wing length on the prevalence of trypanosomes in Glossina morsitans morsitans in eastern Zambia.

BACKGROUND: Tsetse flies (Diptera: Glossinidae) transmit trypanosomiasis (sleeping sickness in humans and nagana in livestock). Several studies have indicated that age, sex, site of capture, starvation and microbiome symbionts, among others, are important factors that influence trypanosome infection in tsetse flies. However, reasons for a higher infection rate in females than in males still largely remain unknown. Considering that tsetse species and sexes of larger body size are the most mobile and the most available to stationary baits, it was hypothesized in this study that the higher trypanosome prevalence in female than in male tsetse flies was a consequence of females being larger than males. METHODS: Black screen fly rounds and Epsilon traps were used to collect tsetse flies in eastern Zambia. Measurement of wing vein length and examination for presence of trypanosomes in the flies were carried out by microscopy. Principal component method was carried out to assess the potential of wing vein length as a predictor variable. The multilevel binary logistic regression method was applied on whole data, one-method data and one-sex data sets to evaluate the hypothesis. RESULTS: Data derived from a total of 2195 Glossina morsitans morsitans were evaluated (1491 males and 704 females). The wing length variable contributed the highest variance percentage (39.2%) to the first principal component. The variable showed significant influence on prevalence of trypanosomes when the analysis was applied on the whole data set, with the log odds for the prevalence of trypanosomes significantly increasing by 0.1 (P = 0.032), per unit increase in wing length. Females had higher trypanosome prevalence rates than males, though not always significant. Furthermore, moving from females to males, wing length significantly reduced by 0.2 (P < 0.0001). CONCLUSIONS: We conclude that wing length is an important predictor variable for trypanosome prevalence in Glossina morsitans morsitans and could partially explain the higher prevalence of trypanosomes in females than in males. However, reasonably representative population data are required for analysis-a serious challenge with the current tsetse sampling methods. Thus, analysis combining data from mobile and stationary methods that include both sexes' data could be useful to verify this hypothesis.

Evaluation of the relative roles of the Tabanidae and Glossinidae in the transmission of trypanosomosis in drug resistance hotspots in Mozambique.

BACKGROUND: Tsetse flies (Diptera: Glossinidae) and tabanids (Diptera: Tabanidae) are haematophagous insects of medical and veterinary importance due to their respective role in the biological and mechanical transmission of trypanosomes. Few studies on the distribution and relative abundance of both families have been conducted in Mozambique since the country's independence. Despite Nicoadala, Mozambique, being a multiple trypanocidal drug resistance hotspot no information regarding the distribution, seasonality or infection rates of fly-vectors are available. This is, however, crucial to understanding the epidemiology of trypanosomosis and to refine vector management. METHODS: For 365 days, 55 traps (20 NGU traps, 20 horizontal traps and 15 Epsilon traps) were deployed in three grazing areas of Nicoadala District: Namitangurine (25 traps); Zalala (15 traps); and Botao (15 traps). Flies were collected weekly and preserved in 70% ethanol. Identification using morphological keys was followed by molecular confirmation using cytochrome c oxidase subunit 1 gene. Trap efficiency, species distribution and seasonal abundance were also assessed. To determine trypanosome infection rates, DNA was extracted from the captured flies, and submitted to 18S PCR-RFLP screening for the detection of Trypanosoma. RESULTS: In total, 4379 tabanids (of 10 species) and 24 tsetse flies (of 3 species), were caught. NGU traps were more effective in capturing both the Tabanidae and Glossinidae. Higher abundance and species diversity were observed in Namitangurine followed by Zalala and Botao. Tabanid abundance was approximately double during the rainy season compared to the dry season. Trypanosoma congolense and T. theileri were detected in the flies with overall infection rates of 75% for tsetse flies and 13% for tabanids. Atylotus agrestis had the highest infection rate of the tabanid species. The only pathogenic trypanosome detected was T. congolense. CONCLUSIONS: Despite the low numbers of tsetse flies captured, it can be assumed that they are still the cyclical vectors of trypanosomosis in the area. However, the high numbers of tabanids captured, associated to their demonstrated capacity of transmitting trypanosomes mechanically, suggest an important role in the epidemiology of trypanosomosis in the Nicoadala district. These results on the composition of tsetse and tabanid populations as well as the observed infection rates, should be considered when defining strategies to control the disease.

Development of Trypanosoma everetti in Culicoides biting midges.

Trypanosoma species (Trypanosomatida, Kinetoplastea) are almost exclusively heteroxenous flagellated parasites, which have been extensively studied as the causative agents of severe trypanosomiasis in humans and domestic animals. However, the biology of avian trypanosomes remains insufficiently known, particularly in wildlife, despite information that some species might be pathogenic and affect the fitness of intensively infected individuals. Avian trypanosomes are cosmopolitans. Due to regular bird seasonal migrations, this host-parasite system might provide new insight for better understanding mechanisms of transcontinental dispersal of pathogens, their ecological plasticity, specificity and speciation. Trypanosoma everetti parasitizes numerous bird species globally, but data on its biology are scarce and its vectors remain unknown. This study aimed to test experimentally whether widespread Culicoides (Diptera: Ceratopogonidae) biting midges are susceptible to infection with this parasite. Two common house martins Delichon urbicum and two sedge warblers Acrocephalus schoenobaenus naturally infected with T. everetti were caught in the wild after arrival from African wintering grounds. Laboratory reared Culicoides nubeculosus and wild-caught Culicoides impunctatus biting midges were exposed by allowing them to take infected blood meals. The experimentally infected and control insects were maintained in the laboratory and dissected at intervals to follow the development of the parasite. Infections were determined using microscopic examination and PCR-based testing. Four closely related haplotypes of T. everetti were found, and each was present in different individual parasite-donor birds. These parasites readily developed and produced metacyclic trypomastigotes in C. nubeculosus and C. impunctatus biting midges. Molecular characterisation of T. everetti was developed. According to Bayesian phylogenetic analysis using a DNA fragment encoding 18S rRNA, the five species of small avian trypanosomes were closely related. Wild caught Culicoides biting midges were also collected and screened for the presence of natural infections. In all, 6.8% of wild-caught biting midges belonging to five Culicoides species were PCR-positive for kinetoplastids, including Trypanosoma species. Culicoides biting midges are readily susceptible and likely naturally transmit avian trypanosomes and thus, should be targeted in epidemiology research of avian trypanosomiasis.

Infecting Triatomines with Trypanosomes.

The infection of triatomines with trypanosomes can be performed with different forms of the parasite, and the procedure is important not only for vector-parasite interaction studies but also for maintaining the infectivity of parasite strains, which guarantees more realistic biological and molecular investigations. Here, I describe how to infect the vector Rhodnius prolixus, a model species, with two different species of Trypanosoma.

Modelling triatomine bug population and Trypanosoma rangeli transmission dynamics: Co-feeding, pathogenic effect and linkage with chagas disease.

Trypanosoma rangeli (T. rangeli), a parasite, is not pathogenic to human but pathogenic to some vector species to induce the behavior changes of infected vectors and subsequently impact the transmission dynamics of other diseases such as Chagas disease which shares the same vector species. Here we develop a mathematical model and conduct qualitative analysis for the transmission dynamics of T. rangeli. We incorporate both systemic and co-feeding transmission routes, and account for the pathogenic effect using infection-induced fecundity and fertility change of the triatomine bugs. We derive two thresholds Rv (the triatomine bug basic reproduction number) and R0 (the T. rangeli basic reproduction number) to delineate the dynamical behaviors of the ecological and epidemiological systems. We show that when Rv>1 and R0>1, a unique parasite positive equilibrium E* appears. We find that E* can be unstable and periodic oscillations can be observed where the pathogenic effect plays a significant role. Implications of the qualitative analysis and numerical simulations suggest the need of an integrative vector-borne disease prevention and control strategy when multiple vector-borne diseases are transmitted by the same set of vector species.

Molecular detection of Megatrypanum trypanosomes in tabanid flies.

Trypanosomes of the subgenus Megatrypanum have been isolated from many mammalian hosts around the world. They are usually non-pathogenic, although they may confuse the parasitological diagnosis of trypanosomosis. Additionally, Trypanosoma theileri has been associated with disease in cattle. Megatrypanum trypanosomes are considered to be transmitted by different arthropods, including tabanids. However, little is known about the potential vectors of Megatrypanum trypanosomes in different parts of the world. The present study reports on the detection of Megatrypanum trypanosomes in Heamatopota pluvialis, Tabanus bromius, Tabanus maculicornis and Tabanus distinguendus in Poland. It also discusses the possible role of these tabanids in the transmission of Megatrypanum trypanosomes.

Monitoring the presence of trypanosomes' DNA - Including Trypanosoma brucei gambiense DNA - From the midguts of riverine Glossina trapped in the south east outskirts of Kinshasa City (Democratic Republic of Congo).

Even if the number of Human African Trypanosomiasis (HAT) cases from Kinshasa province in DRC is going towards elimination for the last decade, cases still occur in the periphery of the city. The diagnosis of 21 cases in the south periphery of Kinshasa, between 2015 and 2017 gives evidence of the existence of an active focus in this area. Here, we present the results of a punctual entomological survey that was realized in july 2014 in the outskirts of the southeast of Kinshasa. Using pyramidal traps, we caught tsetse flies during 2 days, dissecting the fresh ones for further molecular analysis. The average Apparent Density of flies per Trap and per Day was three with a maximum of 5.6 flies in Nganda PIO. Polymerase chain reaction analysis of the midguts provided evidence of a high prevalence (57.2%) of infected flies. Ninety three percent of the trypanosomes that were identified belonged to the Nanomonas species, but Trypanozoon trypanosomes were also present in 24% of the infected flies, including mixed infections with Nanomonas, including 3 flies carrying Trypanosoma brucei gambiense, the human pathogen of trypanosomiasis. These results show that at the time of the field's study there was an active reservoir of trypanosomes, closed to pigsties, knowing that pig is a potential animal reservoir. It also demonstrates that xenomonitoring using the entomological approach can be an efficient tool for monitoring sleeping sickness. Finally, results are discussed in the frame of WHO's HAT elimination project. Regarding Kinshasa, it points out the need of regular epidemiologic surveys.

Tsetse peritrophic matrix influences for trypanosome transmission.

Tsetse flies are important vectors of parasitic African trypanosomes, agents of human and animal trypanosomiasis. Easily administrable and effective tools for disease control in the mammalian host are still lacking but reduction of the tsetse vector populations can reduce disease. An alternative approach is to reduce the transmission of trypanosomes in the tsetse vector. The gut peritrophic matrix (PM) has emerged as an important regulator of parasite transmission success in tsetse. Tsetse has a Type II PM that is constitutively produced by cells in the cardia organ. Tsetse PM lines the entire gut and functions as an immunological barrier to prevent the gut epithelia from responding to commensal environmental microbes present in the gut lumen. Tsetse PM also functions as a physical barrier to trypanosome infections that enter into the gut lumen in an infective blood meal. For persistence in the gut, African trypanosomes have developed an adaptive manipulative process to transiently reduce PM efficacy. The process is mediated by mammalian trypanosome surface coat proteins, Variant Surface Glycoproteins (VSGs) which are shed in the gut lumen and taken up by cardia cells. The mechanism of PM reduction involves a tsetse microRNA (miR-275) which acts thru the Wnt signaling pathway. The PM efficacy is once again reduced later in the infection process to enable the gut established parasites to reenter into the gut lumen to colonize the salivary glands, an essential process for transmission. The ability to modulate PM integrity can lead to innovative approaches to reduce disease transmission.

The reservoir system for Trypanosoma (Kinetoplastida, Trypanosomatidae) species in large neotropical wetland.

Distinct species of Trypanosoma have been documented sharing the same hosts in different environments in intricate transmission networks. Knowing this, this study investigated the role of different hosts in the transmission cycles of Trypanosoma species in the Pantanal biome. The mammals were sampled from November 2015 to October 2016. We sampled a total of 272 wild mammals from 27 species belonging to six orders and 15 families, and three species of triatomines (n = 7). We found high parasitemias by Hemoculture test for Trypanosoma cruzi (TcI), Trypanosoma rangeli, Trypanosoma cruzi marinkellei and Trypanosoma dionisii, and high parasitemias by Microhematocrit Centrifuge Technique for Trypanosoma evansi. The carnivore Nasua nasua is a key host in the transmission cycles since it displayed high parasitemias for T. cruzi, T. evansi and T. rangeli. This is the first report of high parasitemias in Tamandua tetradactyla and cryptic infection in Dasypus novemcinctus by T. cruzi; cryptic infection by T. evansi in Eira barbara, Euphractus sexcinctus and Dasyprocta azarae. The collection of Panstrongylus geniculatus increased the geographic distribution of this vector species in the South America. Our results indicate that Trypanosoma species circulate in a complex reservoir system including different host species with different infective competences.

Odor coding in the antenna of the tsetse fly Glossina morsitans.

Tsetse flies transmit trypanosomiasis to humans and livestock across much of sub-Saharan Africa. Tsetse are attracted by olfactory cues emanating from their hosts. However, remarkably little is known about the cellular basis of olfaction in tsetse. We have carried out a systematic physiological analysis of the Glossina morsitans antenna. We identify 7 functional classes of olfactory sensilla that respond to human or animal odorants, CO2, sex and alarm pheromones, or other odorants known to attract or repel tsetse. Sensilla differ in their response spectra, show both excitatory and inhibitory responses, and exhibit different response dynamics to different odor stimuli. We find striking differences between the functional organization of the tsetse fly antenna and that of the fruit fly Drosophila melanogaster One morphological type of sensilla has a different function in the 2 species: Trichoid sensilla respond to pheromones in Drosophila but respond to a wide diversity of compounds in G. morsitans. In contrast to Drosophila, all tested G. morsitans sensilla that show excitatory responses are excited by one odorant, 1-octen-3-ol, which is contained in host emanations. The response profiles of some classes of sensilla are distinct but strongly correlated, unlike the organization described in the Drosophila antenna. Taken together, this study defines elements that likely mediate the attraction of tsetse to its hosts and that might be manipulated as a means of controlling the fly and the diseases it transmits.

'Hang on a Tick' - Are Ticks Really the Vectors for Australian Trypanosomes?

Trypanosomes are global blood parasites that infect a wide range of vertebrate hosts. Several species of Trypanosoma cause disease in humans and domesticated animals, and the majority are transmitted between hosts by haematophagous invertebrate vectors. Ticks have long been speculated as vectors for Australian trypanosomes. Recent studies using advanced molecular techniques have refocused attention on these arthropods, and whilst they have renewed discussions about Trypanosoma species and their vectors, these reports have simultaneously led to premature conclusions concerning the role of ticks as vectors. Here the controversy surrounding ticks as trypanosome vectors is discussed. We highlight the unanswered questions concerning the role played by ticks in trypanosome transmission and suggest future approaches to resolving these key knowledge gaps.

Message in a vesicle - trans-kingdom intercommunication at the vector-host interface.

Vector-borne diseases cause over 700,000 deaths annually and represent 17% of all infectious illnesses worldwide. This public health menace highlights the importance of understanding how arthropod vectors, microbes and their mammalian hosts interact. Currently, an emphasis of the scientific enterprise is at the vector-host interface where human pathogens are acquired and transmitted. At this spatial junction, arthropod effector molecules are secreted, enabling microbial pathogenesis and disease. Extracellular vesicles manipulate signaling networks by carrying proteins, lipids, carbohydrates and regulatory nucleic acids. Therefore, they are well positioned to aid in cell-to-cell communication and mediate molecular interactions. This Review briefly discusses exosome and microvesicle biogenesis, their cargo, and the role that nanovesicles play during pathogen spread, host colonization and disease pathogenesis. We then focus on the role of extracellular vesicles in dictating microbial pathogenesis and host immunity during transmission of vector-borne pathogens.

Molecular identification of bloodmeal sources and trypanosomes in Glossina spp., Tabanus spp. and Stomoxys spp. trapped on cattle farm settlements in southwest Nigeria.

The interactions of host, vector and parasite in bovine trypanosomiasis transmission cycles in southwest Nigeria are not yet well understood. Trypanosoma (Trypanosomatida: Trypanosomatidae) species infection prevalences and bloodmeal sources were determined in transmitting vectors of the genera Glossina (Diptera: Glossinidae), Tabanus (Diptera: Tabanidae) and Stomoxys (Diptera: Muscidae) collected using Nzi traps in cattle settlements in southwest Nigeria. Sequenced cytochrome B mitochondrial DNA segments obtained from vector digestive tracts identified bloodmeal sources from eight host species, namely human, cattle, hippopotamus, giraffe, gazelle, spotted hyena, long-tailed rat and one unidentified species. Overall, 71.1% [95% confidence interval (CI) 63.0-78.1], 33.3% (95% CI 21.9-47.0) and 22.2% (95% CI 16.2-29.9), respectively, of Glossina, Tabanus and Stomoxys flies were positive for trypanosomes. The observed trypanosome species were Trypanosoma vivax, Trypanosoma congolense, Trypanosoma brucei, Trypanosoma evansi, Trypanosoma simiae and Trypanosoma godfreyi. Trypanosome DNA was more prevalent in tsetse (34.8% Tr. vivax, 51.1% Tr. b. brucei, 5.2% Tr. congolense, 4.4% Tr. simiae and 24.4% mixed infections) than in other flies and the main determinants in all flies were seasonal factors and host availability. To the best of the present group's knowledge, this is the first report of Trypanosoma species in Tabanus and Stomoxys flies in Nigeria. It indicates that vector control programmes should always consider biting flies along with tsetse flies in the control of human and animal trypanosomiasis.

Salivarian Trypanosomosis: A Review of Parasites Involved, Their Global Distribution and Their Interaction With the Innate and Adaptive Mammalian Host Immune System.

Salivarian trypanosomes are single cell extracellular parasites that cause infections in a wide range of hosts. Most pathogenic infections worldwide are caused by one of four major species of trypanosomes including (i) Trypanosoma brucei and the human infective subspecies T. b. gambiense and T. b. rhodesiense, (ii) Trypanosoma evansi and T. equiperdum, (iii) Trypanosoma congolense and (iv) Trypanosoma vivax. Infections with these parasites are marked by excessive immune dysfunction and immunopathology, both related to prolonged inflammatory host immune responses. Here we review the classification and global distribution of these parasites, highlight the adaptation of human infective trypanosomes that allow them to survive innate defense molecules unique to man, gorilla, and baboon serum and refer to the discovery of sexual reproduction of trypanosomes in the tsetse vector. With respect to the immunology of mammalian host-parasite interactions, the review highlights recent findings with respect to the B cell destruction capacity of trypanosomes and the role of T cells in the governance of infection control. Understanding infection-associated dysfunction and regulation of both these immune compartments is crucial to explain the continued failures of anti-trypanosome vaccine developments as well as the lack of any field-applicable vaccine based anti-trypanosomosis intervention strategy. Finally, the link between infection-associated inflammation and trypanosomosis induced anemia is covered in the context of both livestock and human infections.

Trypanosoma cruzi transmission in the wild and its most important reservoir hosts in Brazil.

Trypanosoma cruzi (Kinetoplastea: Trypanosomatidae) infects all tissues of its hosts, which along with humans, include hundreds of mammalian species in the Americas. The epidemiology of T. cruzi has been changing in that currently the majority of the cases and/or outbreaks of Chagas disease occur by the ingestion of comestibles contaminated by T. cruzi metacyclic forms. These cases/outbreaks occur in distinct regional scenarios, mainly in the Amazon biome and are related to the local interaction mode of humans with their surroundings, as well as with the overall local ecological peculiarities. As trypanosomiasis caused by T. cruzi is primarily a zoonosis, understanding the variables that influences its transmission in the wild as well as the role played by the extant fauna in the maintenance of the parasite, is critical in establishing control measures. Here, we present the results of our studies of T. cruzi infection of free ranging wild mammalian fauna in the five biomes of Brazil, a country of continental dimensions. From 1992 up to 2017, we examined a total of 6587 free-ranging non-volant wild mammal specimens. Our studies found that 17% of mammals were seropositive and 8% of all animals displayed positive hemocultures indicative of high parasitemia and, consequently, of infectivity potential. We observed that opossums, mainly Philander spp. and Didelphis spp., the coati Nasua nasua, the capuchin monkey Sapajus libidinosus and the golden lion tamarin Leontopithecus rosalia, were mammal taxa that demonstrated higher rates of positive hemocultures. Additionally, Didelphis spp. demonstrated to be a competent bioaccumulator of TcI diversity. Chiroptera were distinguished for hosting the greatest diversity of species and genotypes of Trypanosoma spp. Additionally the observation of the higher host range of some Trypanosoma spp., shows the need to reassess the ecology of representatives of the taxon. Altogether, our results showed that each locality, may display distinct enzootiological and epidemiological scenarios that must be taken into account when it comes to establishing control and/or clarification campaigns of the local population.

Rabbit trypanosome detection in Phlebotomus perniciosus sand flies from the leishmaniasis outbreak in Madrid, Spain.

Phlebotomine sand flies are known vectors of several pathogens. In Spain, Phlebotomus perniciosus and Phlebotomus ariasi are the proven vectors of Leishmania infantum. Since 2010 a human leishmaniasis outbreak has been notified in Madrid region, central Spain. Studies have shown that P. perniciosus is the only vector confirmed in the focus area and that rabbits and hares are the wild reservoirs incriminated in the cycle of the parasite in the outbreak. Trypanosoma nabiasi is a trypanosomatid found in wild rabbits and its presence has been reported in wild rabbits from southern Spain. Moreover, co-infection with L. infantum was found in some of these animals. However, in Madrid region, there is no information about the transmission of this trypanosome in rabbits. Hence, in this study we investigate if T. nabiasi could be circulating in the aforementioned leishmaniasis focus. Wild P. perniciosus female sand flies were captured in the affected area and analyzed using molecular methods. T. nabiasi DNA was detected in 20 out of 155 female sand flies fed on rabbits by amplification and subsequent sequencing of ITS1 and SSU rRNA fragments. Therefore, we describe for the first time the presence of T. nabiasi and its co-infection with L. infantum in P. perniciosus female sand flies. More investigation is needed in order to elucidate the role of P. perniciosus in the transmission of T. nabiasi among rabbits and its potential consequences.