Corrigendum to "Trypanosoma spp. Neobats: Insights about those poorly known trypanosomatids" [Int. J. Parasitol. Parasites Wildl. 16 (2021) 145-152].

[This corrects the article DOI: 10.1016/j.ijppaw.2021.09.003.].

Trypanosomatid diversity in a bat community of an urban area in Campo Grande, Mato Grosso do Sul, Brazil.

Bats have a long evolutionary history with trypanosomatids, but the role of these flying mammals on parasite transmission cycles in urban areas, especially for Trypanosoma and Leishmania species, remains poorly known. The objective of this study was to evaluate the species richness of trypanosomatids parasitizing a bat community in Campo Grande (CG), a state capital within the Cerrado of the Brazilian Midwest. We evaluated 237 bats of 13 species by means of hemoculture and molecular detection in spleen samples. The bat community of CG appears to participate in the transmission cycles of various species of trypanosomatids. We report an overall trypanosomatid detection rate of 34.2% (n = 81), involving 11 out of 13 sampled bat species. We identified six species of trypanosomatids from 61 bats by analyzing SSU rRNA and/or kDNA: Trypanosoma cruzi DTU TcI, T. c. marinkellei, T. dionisii, Leishmania infantum, L. amazonensis, and T. janseni, with this latter being detected by hemoculture for the first time in a bat species. We also detected a Molecular Operational Taxonomic Unit, Trypanosoma sp. DID, in the phyllostomids Glossophaga soricina and Platyrrhinus lineatus. The highest trypanosomatid richness was observed for Sturnira lilium, which hosted three species: L. infantum, T. dionisii and T. janseni. Given that visceral leishmaniasis is endemic in CG, special focus should be placed on L. infantum. Moreover, L. amazonensis and T. cruzi warrant attention, since these are zoonotic parasites responsible for human cases of tegumentary leishmaniasis and Chagas disease, respectively. In this respect, we discuss how bat communities may influence the Leishmania spp. transmission in endemic areas.

Unraveling the diversity of Trypanosoma species from Central Mexico: Molecular confirmation on the presence of Trypanosoma dionisii and novel Neobat linages.

Bats are one of the groups of mammals with the highest number of associated Trypanosoma taxa. There are 50 Trypanosoma species and genotypes infecting more than 75 species of bats across five continents. However, in Mexico, the inventory of species of the genus Trypanosoma associated with bats is limited to only two species (Trypanosoma vespertilionis and Trypanosoma cruzi) even though 140 species of bats inhabit this country. Specifically, 91 bat species have been recorded in the state of Veracruz, but records of trypanosomatids associated with this mammalian group are absent. Due to the complex Trypanosoma-bat relationship, the high diversity of bat species in Veracruz, as well as the lack of records of trypanosomatids associated with bats for this state, the aim of this work was to analyze the diversity of species of the genus Trypanosoma and their presence from a bat community in the central area of the state of Veracruz, Mexico. During the period of January to August 2022 in the Tequecholapa Environmental Management Unit where bats were collected using mist nets and blood samples were obtained from their thumbs. We extracted genetic material and amplified a fragment of 800 bp of the 18S ribosomal gene of the genus Trypanosoma by conventional PCR. The positive amplicons were sequenced, and phylogenetic reconstruction was performed to identify the parasite species. A total of 285 bats (149♀, 136♂) belonging to 13 species from 10 genera and a single family (Phyllostomidae) were collected. Twenty-three specimens from six species tested positive for the presence of Trypanosoma dionisii, Trypanosoma sp. Neobat 4, and a potential novelty species provisionally named as Trypanosoma sp. Neobat 6. The results of the present work increase the number of species of the genus Trypanosoma infecting bats in Mexico and in the Neotropical region.

Corrigendum: Leishmania infantum infecting the carnivore Nasua nasua from urban forest fragments in an endemic area of visceral leishmaniasis in Brazilian Midwest.

[This corrects the article DOI: 10.3389/fcimb.2022.1050339.].

Old Methods, New Insights: Reviewing Concepts on the Ecology of Trypanosomatids and Bodo sp. by Improving Conventional Diagnostic Tools.

Mixed infections by different Trypanosoma species or genotypes are a common and puzzling phenomenon. Therefore, it is critical to refine the diagnostic techniques and to understand to what extent these methods detect trypanosomes. We aimed to develop an accessible strategy to enhance the sensitivity of the hemoculture, as well as to understand the limitations of the hemoculture and the blood clot as a source of parasitic DNA. We investigated trypanosomatid infections in 472 bats by molecular characterization (18S rDNA gene) of the DNA obtained from the blood clot and, innovatively, from three hemoculture sample types: the amplified flagellates ("isolate"), the pellet of the culture harvested in its very initial growth stage ("first aliquot"), and the pellet of non-grown cultures with failure of amplification ("sediment"). We compared (a) the characterization of the flagellates obtained by first aliquots and isolates; and (b) the performance of the hemoculture and blood clot for trypanosomatid detection. We observed: (i) a putative new species of Bodo in Artibeus lituratus; (ii) the potential of Trypanosoma cruzi selection in the hemoculture; (iii) that the first aliquots and sediments overcome the selective pressure of the hemoculture; and (iv) that the blood clot technique performs better than the hemoculture. However, combining these methods enhances the detection of single and mixed infections.

Kinetoplastid Species Maintained by a Small Mammal Community in the Pantanal Biome.

Kinetoplastids include species economically important in agriculture, livestock, and human health. We evaluated the richness of kinetoplastids that infect small mammals in patches of unflooded forests in the Pantanal biome, an area where we hypothesize that its diversity is higher than currently recognized. Hemocultures (HC) and Next Generation Sequencing (NGS) targeting the 18S rDNA gene were employed for the detection of kinetoplastids. We grouped the positive samples into pools for each small mammal species (Monodelphis domestica, Thylamys macrurus, Oecomys mamorae, Thrichomys fosteri, Clyomys laticeps, and Holochilus chacarius). Eight parasite species were identified: Leishmania amazonensis, L. infantum; Trypanosoma cascavelli (HC + NGS), T. cruzi, T. lainsoni, T. rangeli (HC + NGS), Trypanosoma sp. DID, and Neobodo sp. The use of a tool as sensitive as NGS has increased our awareness of the diversity of kinetoplastids, as well as their host range, with emphasis on the species O. mamorae (seven kinetoplastid species, excepting T. cascavelli in a pool of nine individuals) and T. macrurus (four kinetoplastid species in a single individual). Furthermore, L. infantum and L. amazonensis infections were described in small mammals from this region for the first time. These findings make it mandatory to revisit the kinetoplastids/host associations proposed so far.

Leishmania infantum infecting the carnivore Nasua nasua from urban forest fragments in an endemic area of visceral leishmaniasis in Brazilian Midwest.

Introduction: The aim of the present study was to investigate the occurrence of Leishmania infantum in South American coatis inhabiting two forest fragments in Campo Grande, Mato Grosso do Sul, Midwest region of Brazil, an endemic area of human and canine visceral leishmaniasis (VL). Material and methods: A total of 110 South American coatis were sampled in the conservation unit "Parque Estadual do Prosa" (PEP) and in the residential area "Vila da Base Aérea" (VBA) from March 2018 to April 2019. As a longitudinal study that include up to six recaptures of the same individual, a total of 190 capture events were obtained. Blood, bone marrow and skin samples were obtained for parasitological (axenic culture), serological (Enzyme Linked Immunosorbent Assay - ELISA and Dual-path Platform immunoassay - DPP® CVL) and molecular diagnostic assays (targeting kDNA for Leishmania spp. and L. infantum; and HSP70 followed by sequence analysis). Results: Seropositivity for L. infantum was found in 33 individuals, six in PEP and 27 in VBA. Furthermore, L. infantum was detected by molecular analysis in 16 individuals, seven from PEP and nine from VBA. We also isolated L. infantum from bone marrow of one individual and detected a single positive skin sample in molecular assay from other individual, both from VBA. Discussion: An overall infection rate of 36.4% (40/110) was observed, significantly higher in the VBA (49.1%) than in the PEP (21.6%), probably because VBA presents: (i) a large number of resident dogs and chickens that would be attracting sandflies; (ii) a denser population of this wild mammal species; and (iii) physical barriers and a lack of functional connectivity in the surroundings, preventing these animals to disperse out. We conclude that South American coati populations living in urban forest fragments of Campo Grande are affected by the epidemiological scenario of VL, known to involve dogs, vectors and humans. We highlight the importance of investigate the parasitism by L. infantum in this and other potential L. infantum reservoirs that inhabit urbanized regions endemic to VL.

Trypanosoma spp. Neobats: Insights about those poorly known trypanosomatids.

Bats are infected with several trypanosomatid species; however, assessing the diversity of this interaction remains challenging since there are species apparently unable to grow in conventional culture media. Accordingly, the ecology and biology of the Molecular Operational Taxonomic Units (MOTUs) Trypanosoma spp. Neobats are unknown. Therefore, we performed the molecular characterization targeting the 18S small subunit rDNA from the blood clot of 280 bats of three Brazilian regions (Paraíba, Rio de Janeiro and Acre states), bypassing the selective pressure of hemoculture. From 68 (24%) positive blood clot samples, we obtained 49 satisfactory sequences. Of these successfully sequenced results, T. spp. Neobats (1, 3 and 4) represented 67%, with the most abundant T. sp. Neobat 4 (53%). Our results show: (1) high abundance and wide geographic range of T. sp. Neobat 4, restricted to Carollia bats; (2) high infection rate of T. sp. Neobat 4 in Carollia perspicillata populations (mean 26%); (3) infection with the monoxenous Crithidia mellificae; and (4) a new MOTU (T. sp. Neobat 5) in Artibeus cinereus, positioning in the Trypanosoma wauwau clade. These data corroborate the importance of bats as hosts of many Trypanosoma species and C. mellificae. They also show that the diversity of the T. wauwau clade is underestimated and warn about the high magnitude of trypanosomes we overpass with the hemoculture. Our findings combined with previous data show that T. spp. Neobats include host-specific and host-generalist species, probably playing different ecological roles: T. sp. Neobat 1 shows broad host range; T. spp. Neobat 3 and 4 are restricted to Artibeus and Carollia, respectively. Finally, T. Neobat 4 seems to be a well-succeeded parasite, especially within C. perspicillata metapopulations across a wide geographical distribution. This work is a step forward to understand the biology and life history of T. spp. Neobats.

Complexity and multi-factoriality of Trypanosoma cruzi sylvatic cycle in coatis, Nasua nasua (Procyonidae), and triatomine bugs in the Brazilian Pantanal.

BACKGROUND: Trypanosoma cruzi is dispersed in nature through many transmission mechanisms among a high diversity of vectors and mammalian species, representing particular behaviors and habitats. Thus, each locality has a unique set of conditions underlying the maintenance of this parasite in the wild. The aim of the present study was to evaluate the life-cycle of T. cruzi in free-ranging coatis from the central region of the Brazilian Pantanal using a multi-factorial approach. METHODS: Three methodological blocks were used in the present study: (i) We evaluated T. cruzi infection using serological (ELISA) and parasitological (hemoculture) tests in free-ranging coatis captured from October 2010 to March 2012. In addition, we characterized T. cruzi isolates as DTUs (Discrete Typing Units); (ii) We evaluated Trypanosoma infection in species of Triatoma and Rhodnius inhabiting coati arboreal nests from May to September 2012 using parasitological and molecular assays; and (iii) We analyzed a set of longitudinal data (from 2005 to 2012) concerning the effects of T. cruzi infection on this coati population. Herein, we investigated whether seasonality, host sex, and host age influence T. cruzi prevalence and patterns of infection. RESULTS: The 2010-2012 period presented high seroprevalence on coatis (72.0 % ELISA) and a high percentage of individuals with infectivity competence (20.5 % positive hemoculture). All isolates presented TcI band patterns, occurring in single (n = 3) and mixed infections (1 TcI/T. rangeli; 4 with undefined characterization). Both male and female individuals presented the same transmission potential, expressed as positive hemoculture, which was only detected in the summer. However, overall, the data (2005-2012) highlighted the importance of females for T. cruzi maintenance in the winter. Moreover, twenty-three (67.6 %) bugs from five coati nests (71.4 %) were infected with flagellated forms. Seventeen samples were characterized as T. cruzi (TcI and TcIII genotypes). CONCLUSION: In the Pantanal region, T. cruzi is transmitted in a complex, multifactorial, dynamic and non-linear transmission web. The coati nests might be inserted in this web, acting as important transmission foci at the arboreal stratum to different mammal species with arboreal or scansorial behavior.

Infestation of arboreal nests of coatis by triatomine species, vectors of Trypanosoma cruzi, in a large Neotropical wetland.

The coati (Nasua nasua, Carnivora) is a medium-sized mammal common in the Pantanal of Brazil. Unlike most mammals, coatis construct arboreal nests used for resting and reproduction. In this region, the coati is an important host of Trypanosoma cruzi, the causative agent of Chagas disease. There are two possible routes through coatis can be infected by T. cruzi: the oral route or the vectorial route. However, the relative importance of each of these routes in the infection of coatis and its role in the sylvatic cycle of the parasite are unknown. Our objectives were to investigate: (i) whether coati nests were infested by triatomine bugs, (ii) what species were frequent in the nests, (iii) whether the triatomines in nests were infected by T. cruzi, and (iv) what were the food resources of these triatomines. Eight of the 24 nests sampled were infested with triatomines, a total of 37 specimens of at least two species (Rhodnius stali and Triatoma sordida). In one nest, R. stali and T. sordida co-occurred and both fed on multiple resources, including coatis. This is the first report of triatomines occurring in arboreal nests of coatis. The co-occurrence of two different genera of triatomine vectors and coatis within the limited space of the coati nests provide multiple opportunities for the exchange of the protozoan parasite through both the vectorial and oral transmission routes.

A complexidade, multifatoriedade e não linearidade da rede de transmissão do Trypanosoma cruzi (Trypanosomatida: Trypanosomatidae) em quatis (Carnivora: Procyonidae: Nasua nasua) de vida livre do Pantanal/MS, um estudo longitudinal / The complex, multifactorial and nonlinearity of the transmission of Trypanosoma cruzi (Trypanosomatida: Trypanosomatidae) in raccoons (Carnivora: Procyonidae: Nasua Nasua) free-living Pantanal / MS, a longitudinal study

O Trypanosoma cruzi é um parasito primariamente enzoótico capaz de infectar uma grande variedade de espécies de mamíferos e vetores triatomíneos (Reduviidae: Triatominae). Prévios trabalhos concernentes à infecção por T. cruzi em quatis (Carnívora: Procyonidae: Nasua nasua) de vida livre de Nhecolândia, Pantanal/MS, observaram (i) que essa espécie de hospedeiro é um competente reservatório das principais subpopulações do parasito [TcI, TCII (TcII/TcV/TcVI) e Z3 (TcIII/TcIV)], (ii) maior população de fêmeas com potencial de transmissão do parasito, expresso por hemocultivo (HC) positivo, e (ii) maior infectividade (HC positivo)pela população no período seco do Pantanal. Indo além, refinamos e ampliamos nessa dissertação as análises quanto à dinâmica de transmissão das principais linhagens do T. cruzi nessa população de quatis, considerando variáveis temporais, climáticas, idade e sexo do hospedeiro. Para tal, (l) foi padronizado e validado ao ensaio imunoenzimático (ELISA); (2) foi avaliada a infecção por T. cruzi em 35 triatomíneos (adultos das espécies Rhodnius stali e Triatoma sordida e ninfas do mesmo gênero) coletados em sete ninhos de quatis entre maio a setembro/2012; (3) foi dado seguimento ao estudo da infecção por T. cruzi em quatis entre outubro/2010 a março/2012 e (4) foi realizada uma análise conjunta dos dados obtidos em sete anos de acompanhamento da população de quatis (março 2005 a 2012). Os resultados encontrados foram: o diagnóstico por ELISA apresentou 92% de sensibilidade (considerando tanto imunofluorescência indireta – RIFI – quanto HC como padrão ouro e 60% de especificidade (RIFI) como padrão ouro). (2) De sete ninhos, cinco albergavam triatomíneos infectados por T. cruzi. Dos 23 (66%) espécimes positivos pelo exame a fresco, 17 apresentaram infecções simples e mistas por TcI e Z3. (3) A circulação das principais subpopulações (TcI, TCII e Z3) do T. cruzi se manteve, contudo não observamos o mesmo perfil de transmissão descrito anteriormente: foi observada maior infectividade pela população no período chuvoso, além de machos e fêmeas com maior potencial de transmissão (HC positivo). Por fim, (4) observamos (a) um ciclo bem estabelecido do T. cruzi na população de quatis, apresentando papel primário na dispersão e manutenção das principais subpopulações do parasito, (b) o genótipo TcI predominou em todos os anos; (c) o caráter multifatorial e a temporalidade da dinâmica de transmissão do T. cruzi: fêmeas apresentaram maior potencial de transmitir o T. cruzi, o que ocorreu principalmente na estação seca (inverno) (expresso por hemocultivo positivo). Concluímos que o ELISA é promissor como diagnóstico sorológico da infecção por T. cruzi em quatis; os ninhos são focos de transmissão para diferentes espécies de mamíferos que frequentam o estrato arbóreo; os quatis de infectam desde jovens nos ninhos; e na região do Pantanal, a transissão do T. cruzi ocorre através de uma complexa e dinâmica rede de transmissão não linear, de caráter multifatorial e imprevisível, o que demonstra a importância de estudos de metodologia longitudinal.

Modulating variables of Trypanosoma cruzi and Trypanosoma evansi transmission in free-ranging Coati (Nasua nasua) from the Brazilian Pantanal region.

This is a long-term follow-up of infection by Trypanosoma cruzi (TC) and Trypanosoma evansi (TE) in the free-ranging coatis (Procyonidae: Nasua nasua) from Pantanal region (Mato Grosso do Sul, Brazil). We evaluated TC and TE infection by immunofluorescence assay, hemoculture (HC), and microhematocrit centrifuge techniques (MHCT). We also examined coatis health by quantifying hematological parameters including packed cell volume (PCV), white blood cell (WBC) count, and differential leukocyte count. TC isolates thought HC were typed by miniexon gene. Mixed infections by both parasites and the two main lineages of TC (76% TCI, 3% TCII, and 14% TCI/TCII) were observed. Trypanosoma rangeli was also isolated (7%). Overall, seroprevalence of TC and TE infection were 53.5% and 42.0%, respectively. Positive HC (indicating high TC parasitemia) occurred in 34% of seropositive coatis for TC, and positive MHCT (high TE parasitemia) were observed in 36.4% of seropositive coatis for TE. We detected higher prevalence of positive HC in females (72%) than males (43%), and also during the dry season, indicating a seasonal potential of this host species on TC transmission. These features did not occur for TE infection. However, prevalence of TE based on serology and MHCT was higher among adults than subadults. Coatis with positive HC or MHCT displayed a slight decrease in their WBC. In contrast to the animals with positive HC, coatis with positive MHCT displayed a decrease on their PCV. Moreover, concurrent high TC and TE parasitemia caused a larger decrease of PCV values. This study corroborates the importance of coatis in the maintenance of TC and TE transmission cycles in the southern Pantanal and shows a seasonal character of TC transmissibility to its vector by the coati population from the study area.