Phylogenetic and morphological characterization of trypanosomes from Brazilian armoured catfishes and leeches reveal high species diversity, mixed infections and a new fish trypanosome species

Abstract\ud \ud Background\ud Several Trypanosoma species transmitted by leeches infect marine and freshwater fish worldwide. To date, all South American fish trypanosome species identified have been based on unreliable morphological parameters. We recently isolated and cultured trypanosomes from the Brazilian armoured catfishes Hypostomus luetkeni and H. affinis. Here, we report the first phylogenetic analyses of South American (Brazilian) trypanosomes isolated from fish, and from leeches removed from these fish. We also analysed morphologically and morphometrically the different forms of fish, leech and cultured trypanosomes.\ud \ud \ud Methods\ud V7V8 SSU rRNA and gGAPDH sequences were used for phylogenetic analysis of Brazilian fish and leech trypanosomes. Trypanosomes from cultures, fish blood and leech samples were also characterized morphologically and morphometrically by light and electron microscopy.\ud \ud \ud Results\ud In blood smears from fish high trypanosome prevalence (90–100 %) and parasitemia (0.9-1.0x102) were observed. Phylogenetic relationships using SSU rRNA and gGAPDH showed that, despite relevant sequence divergence, all Brazilian fish (and derived cultures) and leech trypanosomes clustered together into a single clade. The Brazilian clade clustered with European, North American and African fish trypanosomes. Based on sequence analysis, we uncovered a new species of Brazilian fish trypanosome, Trypanosoma abeli n. sp. Trypanosoma abeli cultures contained pleomorphic epimastigotes, small trypomastigotes and rare sphaeromastigotes. Ultrastructural features of T. abeli included a cytostome-cytopharynx complex in epi- and trypomastigotes, a compact rod-like kinetoplast, lysosome-related organelles (LROs) and multivesicular bodies. Trypanosomes found in fish blood smears and leech samples were highly pleomorphic, in agreement with sequence data suggesting that catfishes and leeches often have mixed trypanosome infections.\ud \ud \ud Conclusions\ud \ud Trypanosoma abeli n. sp. is the first trypanosome from South American fishes isolated in culture, positioned in phylogenetic trees and characterized at the ultrastructural level. Trypanosoma abeli n. sp. is highly prevalent in H. luetkeni and H. affinis armoured catfish from the Atlantic Forest biome, and in other catfish species from the Amazon and the Pantanal. Sequencing data suggested that Brazilian catfish often have mixed trypanosome infections, highlighting the importance of molecular characterization to identify trypanosome species in fishes and leeches.We are grateful to Mr. Alcir Pereira de Souza, Ms. Alessandra Simões de\ud Toledo Pereira and Mrs. Tereza da Silva Lemos for their valuable support\ud with the fieldwork, and to Dr. José Carlos Oliveira and Valter M. AzevedoSantos\ud for fish identification. DNA sequencing and phylogenetic analyses\ud done in USP were supported by grants from CNPq and CAPES to MMGT. ML\ud is a postdoctoral fellow funded by CNPq, BRF is a PhD student funded by\ud CAPES, CSR is PhD student funded by CNPq, and LH is postdoctoral fellow\ud funded by FAPERJ

Bidirectional intraflagellar transport is restricted to two sets of microtubule doublets in the trypanosome flagellum

Intraflagellar transport (IFT) is the rapid bidirectional movement of large protein complexes driven by kinesin and dynein motors along microtubule doublets of cilia and flagella. In this study, we used a combination of high-resolution electron and light microscopy to investigate how and where these IFT trains move within the flagellum of the protist Trypanosoma brucei. Focused ion beam scanning electron microscopy (FIB-SEM) analysis of trypanosomes showed that trains are found almost exclusively along two sets of doublets (3–4 and 7–8) and distribute in two categories according to their length. High-resolution live imaging of cells expressing mNeonGreen::IFT81 or GFP::IFT52 revealed for the first time IFT trafficking on two parallel lines within the flagellum. Anterograde and retrograde IFT occurs on each of these lines. At the distal end, a large individual anterograde IFT train is converted in several smaller retrograde trains in the space of 3–4 s while remaining on the same side of the axoneme

Caderno de pós-graduação em direito: direito do trabalho contemporâneo

Os trabalhos científicos ora apresentados são fruto da disciplina Direito do trabalho contemporâneo, ministrada no segundo bimestre de 2022 pela Professora Moara Silva Vaz de Lima. No bimestre, foram elaborados artigos que abordam as novas dinâmicas empregatícias no mundo contemporâneo, sob o prisma do direito do trabalho. Foram selecionados 2 (dois) artigos sobre os assuntos e textos trabalhados durante o bimestre. Os textos são de autoria dos discentes da disciplina, sendo estas: Carolina de Morais Arruda e Ianka Laryssa Santos de Paula

Contribuição ao estudo da biologia dos Tripanossomas de Anuros neotropicais

The present research had as its main goal, checking the infection through trypanosomes in Anuran around Mato Grosso and Rio de Janeiro States. For that purpose 20 species from Bufonidae, Microhylidae, Hylidae and Leptodactylidae families were collected during the fauna rescue activities at Guapore Power Station the border of Vale de São Domingos, Pontes Lacerda, MT and in Seropédia County, at the Rural Federal University Campus in Rio de Janeiro, RJ. Trypanosomes were found in blood smears and organ impressions in 36,36% of the Osteocephalus sp. studied hosts, 50% of Leptodactylus chaquensis, 100% of Leptodactylus fuscus, 100% of Leptodactylus Lineatus and 100% of Leptodactylus ocellatus. Polimorfism was verified on the founded species, splitting into Trypanosoma sp.1, Trypanosoma sp.2, Trypanosoma chattoni, Trypanosoma rotatorium (lato sensu) and Trypanosoma sp.3 that exhibit high polymorphism. Periferal blood samples were inoculated through diphasic blood Agar and LIBHIT-K, liquid fase. Isolates of Trypanosoma sp.3 were obtained from three out of four collected hosts. Trypanosomes were observed 4 days after the inoculation, kept in cultures for 8 months and samples were taken for ultrastructural analysis. Epimastigotes, sphaeromastigotes and trypomastigotes were the culture forms founded. Binary fission was noticed in epimastigotes and in sphaeromastigotes only, trypomastigotes were not observed in reproduction. The ultrastructural analysis of Trypanosoma sp.3 confirmed its singularity through morphological characteristic such as elongated glicosomes. This research accomplishes its first record of the infection through trypanosomes in the Anuran Osteocephalus genre and in the species of Leptodactylus fuscus, Leptodactylus lineatus and the presence of T. chattoni in the Leptodactyllus species in Brazil. The usage of LIBHIT-K as an effective mean for Trypanosoma sp.3 maintenance and the first records of ultrastructure anuran trypanosomes in Brazil.O presente estudo teve como objetivo verificar a infecção por tripanossomas em anuros em regiões dos estados do Mato Grosso e Rio de Janeiro. Para tal, 20 espécies das famílias Bufonidae, Microhylidae, Hylidae e Leptodactylidae foram coletadas durante as atividades de resgate de fauna da Usina hidrelétrica do Guaporé, na divisa dos municípios Vale de São Domingos, Pontes e Lacerda, MT e no Município de Seropédica, Campus da Universidade Federal Rural do Rio de Janeiro, RJ. Verificou-se a presença de tripanossomas em esfregaços sangüíneos e impressões de órgãos em 36,36% dos hospedeiros analisados de Osteocephalus sp., 50% de Leptodactylus chaquensis, 100% de Leptodactylus fuscus, 100% de Leptodactylus lineatus e 100% de Leptodactylus ocellatus. Foi verificado polimorfismo das espécies encontradas, separando-se Trypanosoma sp.1; Trypanosoma sp.2; Trypanosoma sp.3; Trypanosoma chattoni e Trypanosoma rotatorium (lato sensu). Destes o polimorfismo mais acentuado foi observado em Trypanosoma sp.3. Amostras de sangue da circulação periférica e visceral de Leptodactylus ocellatus foram incluídas em meio de bifásico constituído de Agar e sangue e fase líquida de LIBHIT-K. Foram obtidos isolados de Trypanosoma sp.3 de três dos quatro hospedeiros coletados. Tripanossomas foram observados 4 dias após inoculação, mantidos por 8 meses em cultura e retiradas amostras para análise da ultra-estrutura. Nas formas de cultura foram observadas epimastigotas, esferomastigotas e tripomastigotas. Divisão binária foi observada em epimastigotas e esferomastigotas; tripomastigotas não foram observadas em divisão. A análise da ultra-estrutura de Trypanosoma sp.3 confirmou sua singularidade por características morfológicas como glicosomos alongados. Este estudo faz o primeiro registro da infecção por tripanossomas em anuros do gênero Ostheocephalus e nas espécies Leptodactylus chaquensis, Leptodactylus fuscus, Leptodactylus lineatus e a ocorrência de T. chattoni em espécies do gênero Leptodactyllus e no Brasil. Registra-se ainda a utilização do meio LIBHIT-K como eficaz para manutenção de Trypanosoma sp.3 e o primeiro estudo sobre registro da ultra-estrutura de tripanossomas de anuros no Brasil.CAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível Superio

First record of Trypanosoma chattoni in Brazil and occurrence of other Trypanosoma species in Brazilian frogs (Anura, Leptodactylidae)

The present study provides the first record of Trypanosoma chattoni Mathis and Leger, 1911, in a new host, Leptodactylus fuscus Schneider, 1799 (Anura, Leptodactylidae), and the occurrence of Trypanosoma rotatorium-like species in Leptodactylus chaquensis Cei, 1950. The anurans were captured in the State of Mato Grosso, Brazil. Blood samples were obtained by cardiac puncture, and blood smears were examined for the presence of hemoparasites. The Trypanosoma rotatorium-like species in this study refers to a short-bodied trypomastigote that has a conspicuous undulating membrane but lacks a free flagellum; T. chattoni refers to a monomorphic parasite that has a rounded body, a kinetoplast adjacent to the nucleus, and a short flagellum. © American Society of Parasitologists 2008

Timing and original features of flagellum assembly in trypanosomes during development in the tsetse fly

International audienceBackground: Trypanosoma brucei exhibits a complex life-cycle alternating between tsetse flies and mammalian hosts. When parasites infect the fly, cells differentiate to adapt to life in various tissues, which is accompanied by drastic morphological and biochemical modifications especially in the proventriculus. This key step represents a bottleneck for salivary gland infection.Methods: Here, we monitored flagellum assembly in trypanosomes during differentiation from the trypomastigote to the epimastigote stage, i.e. when the nucleus migrates to the posterior end of the cell, by using three-dimensional electron microscopy (focused ion beam scanning electron microscopy, FIB-SEM) and immunofluorescence assays.Results: The combination of light and electron microscopy approaches provided structural and molecular evidence that the new flagellum is assembled while the nucleus migrates towards the posterior region of the body. Two major differences with well-known procyclic cells are reported. First, growth of the new flagellum begins when the associated basal body is found in a posterior position relative to the mature flagellum. Secondly, the new flagellum acquires its own flagellar pocket before rotating on the left side of the anterior-posterior axis. FIB-SEM revealed the presence of a structure connecting the new and mature flagellum and serial sectioning confirmed morphological similarities with the flagella connector of procyclic cells. We discuss the potential function of the flagella connector in trypanosomes from the proventriculus.Conclusions: These findings show that T. brucei finely modulates its cytoskeletal components to generate highly variable morphologies

Morphological and Morphometric Characterization of Trypanosomes in Leptodactylus lineatus and Osteocephalus sp. (Anura) from Brazilian Midwest

The objective of this study was to evaluate parasitism caused by trypanosomes in naturally infected anurans from Brazilian Midwest and characterize trypanosomes by morphology and morphometry. Anurans were captured from margins of the Guaporé River in transition areas between Cerrado and Rain Forest. Blood samples of 16 anurans species were obtained by cardiac puncture and analyzed for the presence of hemoparasites. Trypanosomes were found infecting two anuran species, Leptodactylus lineatus (Leptodactylidae) and Osteocephalus sp. (Hylidae). It was observed high prevalence of trypanosomes (100%; N = 3) in L. lineatus, with intensity of infection of 9.9 × 102 parasites/ml. In Osteocephalus sp. the prevalence of trypanosomes was comparatively lower (36%; N = 4 from 11), however, the intensity of infection was higher 2.16 × 103 parasites/ml. Morphological variation related to body length and width of bloodstream trypomastigotes were observed in different host species and between individual hosts of the same species. Polymorphic trypanosomes were separated into morphotypes and classified as elongated or short trypanosomes. Trypanosomes infecting L. lineatus are elongated with slender or broad body and trypanosomes infecting Osteocephalus sp. presented a short body with broad posterior extremity. The parasitological analyses and morphological characterizations presented in this study contribute to the knowledge of diversity of trypanosomes from neotropical anurans.The objective of this study was to evaluate parasitism caused by trypanosomes in naturally infected anurans from Brazilian Midwest and characterize trypanosomes by morphology and morphometry. Anurans were captured from margins of the Guaporé River in transition areas between Cerrado and Rain Forest. Blood samples of 16 anurans species were obtained by cardiac puncture and analyzed for the presence of hemoparasites. Trypanosomes were found infecting two anuran species, Leptodactylus lineatus (Leptodactylidae) and Osteocephalus sp. (Hylidae). It was observed high prevalence of trypanosomes (100%; N = 3) in L. lineatus, with intensity of infection of 9.9 × 102 parasites/ml. In Osteocephalus sp. the prevalence of trypanosomes was comparatively lower (36%; N = 4 from 11), however, the intensity of infection was higher 2.16 × 103 parasites/ml. Morphological variation related to body length and width of bloodstream trypomastigotes were observed in different host species and between individual hosts of the same species. Polymorphic trypanosomes were separated into morphotypes and classified as elongated or short trypanosomes. Trypanosomes infecting L. lineatus are elongated with slender or broad body and trypanosomes infecting Osteocephalus sp. presented a short body with broad posterior extremity. The parasitological analyses and morphological characterizations presented in this study contribute to the knowledge of diversity of trypanosomes from neotropical anurans

Progress in Research on African Trypanosomes: Highlights from an Exceptional Decade

International audienceIn the late nineteenth century, Trypanosoma brucei was discovered as the parasitic protist responsible for Human African Trypanosomiasis (HAT), also known as sleeping sickness. It is transmitted by the bite of the tsetse fly where trypanosomes undergo several steps of differentiation, proliferation and migration that ultimately lead to the production of parasites than can again be infective for a mammalian host. Here, we review four major areas of trypanosome research that saw spectacular progress in knowledge over the last decade. The cell biology of the parasite can now be studied at unprecedented level thanks to the development of 3D electron microscopy, live imaging and super-resolution microscopy, revealing the architecture of all organelles, such as the flagellum that performs multiple essential functions. The omics area has lifted the basic vision of the genome sequence to a highly sophisticated appreciation of gene expression and chromatin organisation, with the ability to interrogate gene function thanks to advanced reverse genetics both at the individual and the global level. These developments were translated in vivo especially via imaging of the infection in the insect and the mammalian host. This resulted in a reconsideration of the life cycle, revealing the critical role of extravascular parasites in mammalian hosts where the skin now appears as a central reservoir for transmission. These findings will have an impact on monitoring and treating the disease in the field, as well as on the programme for elimination of HAT