To the Skin and Beyond: The Immune Response to African Trypanosomes as They Enter and Exit the Vertebrate Host.

African trypanosomes are single-celled extracellular protozoan parasites transmitted by tsetse fly vectors across sub-Saharan Africa, causing serious disease in both humans and animals. Mammalian infections begin when the tsetse fly penetrates the skin in order to take a blood meal, depositing trypanosomes into the dermal layer. Similarly, onward transmission occurs when differentiated and insect pre-adapted forms are ingested by the fly during a blood meal. Between these transmission steps, trypanosomes access the systemic circulation of the vertebrate host via the skin-draining lymph nodes, disseminating into multiple tissues and organs, and establishing chronic, and long-lasting infections. However, most studies of the immunobiology of African trypanosomes have been conducted under experimental conditions that bypass the skin as a route for systemic dissemination (typically via intraperitoneal or intravenous routes). Therefore, the importance of these initial interactions between trypanosomes and the skin at the site of initial infection, and the implications for these processes in infection establishment, have largely been overlooked. Recent studies have also demonstrated active and complex interactions between the mammalian host and trypanosomes in the skin during initial infection and revealed the skin as an overlooked anatomical reservoir for transmission. This highlights the importance of this organ when investigating the biology of trypanosome infections and the associated immune responses at the initial site of infection. Here, we review the mechanisms involved in establishing African trypanosome infections and potential of the skin as a reservoir, the role of innate immune cells in the skin during initial infection, and the subsequent immune interactions as the parasites migrate from the skin. We suggest that a thorough identification of the mechanisms involved in establishing African trypanosome infections in the skin and their progression through the host is essential for the development of novel approaches to interrupt disease transmission and control these important diseases.

Novel distamycin analogues that block the cell cycle of African trypanosomes with high selectivity and potency.

Polyamides-based compounds related to the Streptomycetal distamycin and netropsin are potent cytostatic molecules that bind to AT-rich regions of the minor groove of the DNA, hence interfering with DNA replication and transcription. Recently, derivatives belonging to this scaffold have been reported to halt the proliferation of deadly African trypanosomes by different and unrelated mechanisms. Here we describe the synthesis and preliminary characterization of the anti-trypanosomal mode of action of new potent and selective distamycin analogues. Two tri-heterocyclic derivatives containing a central N-methyl pyrrole ring (16 and 17) displayed high activity (EC50 < 20 nM) and selectivity (selectivity index >5000 with respect to mammalian macrophages) against the infective form of T. brucei. Both compounds caused cell cycle arrest by blocking the replication of the mitochondrial DNA but without affecting its integrity. This mode of action clearly differs from that reported for classical minor groove binder (MGB) drugs, which induce the degradation of the mitochondrial DNA. In line with this, in vitro assays suggest that 16 and 17 have a comparatively lower affinity for different template DNAs than the MGB drug diminazene. Therapeutic efficacy studies and stability assays suggest that the pharmacological properties of the hits should be optimized. The compounds can be rated as excellent scaffolds for the design of highly potent and selective anti-T. brucei agents.

Comparative genomic analysis of six Glossina genomes, vectors of African trypanosomes.

BACKGROUND: Tsetse flies (Glossina sp.) are the vectors of human and animal trypanosomiasis throughout sub-Saharan Africa. Tsetse flies are distinguished from other Diptera by unique adaptations, including lactation and the birthing of live young (obligate viviparity), a vertebrate blood-specific diet by both sexes, and obligate bacterial symbiosis. This work describes the comparative analysis of six Glossina genomes representing three sub-genera: Morsitans (G. morsitans morsitans, G. pallidipes, G. austeni), Palpalis (G. palpalis, G. fuscipes), and Fusca (G. brevipalpis) which represent different habitats, host preferences, and vectorial capacity. RESULTS: Genomic analyses validate established evolutionary relationships and sub-genera. Syntenic analysis of Glossina relative to Drosophila melanogaster shows reduced structural conservation across the sex-linked X chromosome. Sex-linked scaffolds show increased rates of female-specific gene expression and lower evolutionary rates relative to autosome associated genes. Tsetse-specific genes are enriched in protease, odorant-binding, and helicase activities. Lactation-associated genes are conserved across all Glossina species while male seminal proteins are rapidly evolving. Olfactory and gustatory genes are reduced across the genus relative to other insects. Vision-associated Rhodopsin genes show conservation of motion detection/tracking functions and variance in the Rhodopsin detecting colors in the blue wavelength ranges. CONCLUSIONS: Expanded genomic discoveries reveal the genetics underlying Glossina biology and provide a rich body of knowledge for basic science and disease control. They also provide insight into the evolutionary biology underlying novel adaptations and are relevant to applied aspects of vector control such as trap design and discovery of novel pest and disease control strategies.

Inhibitor-Induced Dimerization of an Essential Oxidoreductase from African Trypanosomes.

Trypanosomal and leishmanial infections claim tens of thousands of lives each year. The metabolism of these unicellular eukaryotic parasites differs from the human host and their enzymes thus constitute promising drug targets. Tryparedoxin (Tpx) from Trypanosoma brucei is the essential oxidoreductase in the parasite's hydroperoxide-clearance cascade. In vitro and in vivo functional assays show that a small, selective inhibitor efficiently inhibits Tpx. With X-ray crystallography, SAXS, analytical SEC, SEC-MALS, MD simulations, ITC, and NMR spectroscopy, we show how covalent binding of this monofunctional inhibitor leads to Tpx dimerization. Intra- and intermolecular inhibitor-inhibitor, protein-protein, and inhibitor-protein interactions stabilize the dimer. The behavior of this efficient antitrypanosomal molecule thus constitutes an exquisite example of chemically induced dimerization with a small, monovalent ligand that can be exploited for future drug design.

Lysine Deacetylase Inhibitors in Parasites: Past, Present, and Future Perspectives.

Current therapies for human parasite infections rely on a few drugs, most of which have severe side effects, and their helpfulness is being seriously compromised by the drug resistance problem. Globally, this is pushing discovery research of antiparasitic drugs toward new agents endowed with new mechanisms of action. By using a "drug repurposing" strategy, histone deacetylase inhibitors (HDACi), which are presently clinically approved for cancer use, are now under investigation for various parasite infections. Because parasitic Zn2+- and NAD+-dependent HDACs play crucial roles in the modulation of parasite gene expression and many of them are pro-survival for several parasites under various conditions, they are now emerging as novel potential antiparasitic targets. This Perspective summarizes the state of knowledge of HDACi (both class I/II HDACi and sirtuin inhibitors) targeted to the main human parasitic diseases (schistosomiasis, malaria, trypanosomiasis, leishmaniasis, and toxoplasmosis) and provides visions into the main issues that challenge their development as antiparasitic agents.

Estudo do controle autonômico do coração na doença de chagas / Study of control autonômicodo heart in Chagas' disease

O comprometimento do sistema nervoso autônomo do coração na enfermidade chagasica crônica:referência ao envolvimento do sistema nervoso na Doença de Chagas foi inicialmente feita por Carlos Chagas, em 1913 ; "Relativement à la frequence des formes nerveuses de la trypanos- somiase, nous possedons des observations nombreuses, qui nous au torisent à affirmer que cette maladie est celle qui? peut-être, provoque, en pathologic humaine, la plus grand nombre d'affec tions organiques du systéme nerveux central" (01). Embora Carlos Chagas, nesta época, se referisse apenas a manifestações decorren tes do comprometimento do sistema nervoso central, ja em 1921, su geriu a possibilidade de "lesões dos nervos sensitivos intracar- díacos", ou de um "augmento do tonus sympathico", para esclarecer este "capítulo novo da pathologia humana"A relevância emprestada pelos diversos autores à lesão neuronal, como fator patogênico, tem variado bastante. Koborle (02) postulou ser a moléstia de Chagas uma enfermidade exclusiva do sistema nervoso parassimpãtico ("cardiopatia parassimpatico- priva"), em qu9 as ’“patias" chagásicas —cardiopatia, bronqui- ectasia, megas etc— representariam tão-somente sequelas das lesões neuronais ocorridas na fase aguda. Por outro lado, Andrade e Andrade (0 3), embora admitindo a ocorrência de intensa destruição neuronal no coração de chagãsicos brasileiros, a qual comunicaria certas peculiaridades ã miocardiopatia em nosso meio, acreditam que a lesão autonômica não á fundamental na cardiopatia.

The bacterial flora of tsetse fly midgut and its effect on trypanosome transmission.

The tsetse fly, Glossina palpalis is a vector of the trypanosome that causes sleeping sickness in humans and nagana in cattle along with associated human health problems and massive economic losses. The insect is also known to carry a number of symbionts such as Sodalis, Wigglesworthia, Wolbachia whose effects on the physiology of the insect have been studied in depth. However, effects of other bacterial flora on the physiology of the host and vector competence have received little attention. Epidemiological studies on tsetse fly populations from different geographic sites revealed the presence of a variety of bacteria in the midgut. The most common of the flora belong to the genera Entrobacter (most common), Enterococcus, and Acinetobacter. It was a little surprising to find such diversity in the tsetse midgut since the insect is monophagous consuming vertebrate blood only. Diversity of bacteria is normally associated with polyphagous insects. In contrast to the symbionts, the role of resident midgut bacterial flora on the physiology of the fly and vector competence remains to be elucidated. With regard, Sodalis glossinidius, our data showed that flies harbouring this symbiont have three times greater probability of being infected by trypanosomes than flies without the symbiont. The data delineated in these studies under score the need to carry out detailed investigations on the role of resident bacteria on the physiology of the fly and vector competence.

Apicomplexa, trypanosoma and parasitic nematode protein kinases as antiparasitic therapeutic targets.

Parasitic infections caused by Plasmodium, Trypanosoma, Leishmania, Toxoplasma and parasitic nematodes affect hundreds of millions of individuals worldwide and are the cause of significant mortality and morbidity, particularly in developing countries. These diseases also have an impact on individuals from developed countries; for example, some US troops in Iraq and Afghanistan have been infected with Leishmania. The annual mortality associated with parasitic infections is estimated to be 1.5 million deaths. The socioeconomic impact of the morbidity associated with parasitic infections is significant, and the development of new drugs, aimed at novel targets, is urgently needed to develop effective treatments for these diseases. The small-molecule inhibitors discussed in this review constitute useful tools with which to explore the relevance of kinase inhibition in inducing antiparasitic activity. The aim of recent target-based approaches used in the development of parasite kinase inhibitors is to identify novel antiparasitic agents with therapeutic potential.

Proteinogramas séricos de ratos Wistar experimentalmente infectados com Trypanosoma evansi / Serum protein concentrations in Wistar rats experimentally infected with Trypanosoma evansi

Estabeleceu-se o perfil eletroforético de proteínas séricas de ratos Wistar experimentalmente infectados com Tripanosoma evansi, utilizando-se 40 ratos, distribuídos em oito grupos de cinco animais cada. Um grupo foi mantido como testemunho (G1), e os demais (G2 a G8) foram inoculados, via intraperitoneal, com cerca de 10³tripomastigota de T. evansi. Amostras de sangue para obtenção de soro foram coletadas no quinto (G2), 10º (G3), 15º (G4), 30º (G5), 45º (G6), 60º (G7) e 75º (G1 e G8) dia após as inoculações. O fracionamento das proteínas foi realizado pela técnica SDS-PAGE. Foram identificadas 31 proteínas, sendo sete de fase aguda: ceruloplasmina (101KD), hemopexina (83KD), transferrina (75KD), albumina (66KD), antitripsina (60KD), haptoglobina (44KD) e glicoproteína ácida (38KD). As proteínas com pesos moleculares 12KD; 22KD; 25KD; 28KD; 32,5KD; 35KD; 53,5KD; 63KD e 72KD apareceram apenas nos ratos inoculados com T. evansi.

This study established the electrophoretic profile of serum proteins of Wistar rats experimentally infected with Tripanosoma evansi. For such, 40 rats were allocated into eight groups of five animals. A group was kept as control (G1) and the others (G2 to G8) were intraperitoneally inoculated with 1.0 x 10³ tripomastigote of T. evansi. Blood samples were collected at 5th (G2), 10th (G3), 15th (G4), 30th (G5), 45th (G6), 60th (G7), and 75th (G1 and G8) days after inoculation (DAI). The serum protein concentrations were determined by means of sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Thirty-one distinct proteins were identified, seven of these were identified as acute phase proteins: ceruloplasmin (110KD), hemopexin (83KD), transferrin (75KD), albumin (66KD), antitrypsin (60KD), haptoglobin (44KD), and acid glycoprotein (38KD). The proteins with molecular weights 12KD; 22KD; 25KD; 28KD; 32,5KD; 35KD; 53,5KD; 63KD, and 72KD were found only in infected rats.

Death of a trypanosome: a selfish altruism.

African trypanosomes and some related parasitic protozoa are affected by a form of programmed cell death (PCD) that shows typical hallmarks of apoptosis. Although it has been speculated that PCD has a function in life-cycle progression and the struggle for survival of these parasites, no satisfactory model has yet been proposed for the molecular mechanism(s) of PCD in protozoa, raising questions about its physiological relevance in these organisms. As we discuss here, the most important point that needs to be addressed is whether a single-celled organism can undertake a process that is considered altruistic.

Crabs, leeches and trypanosomes: an unholy trinity?

The red king crab Paralithodes camtschaticus was deliberately introduced to the Barents Sea in the 1960s and 1970s from its native area in the North Pacific. The carapace of these crabs is a favoured substrate for the leech Johanssonia arctica to deposit its eggs, and the leech is a vector for a trypanosome blood parasite of marine fish, including cod. We examined cod for trypanosome infections during annual cruises along the coast of Finnmark in North Norway over three successive years from stations along a gradient of over 1000 km. In every year the level of trypanosome infection in cod was significantly highest in the area with the greatest density of king crabs. We propose the hypothesis that the burgeoning population of red king crabs in this area is indirectly responsible for increased transmission of trypanosomes to cod by promoting an increase in the population of the leech vector.

Polymorphism in trypomastigotes of Trypanosoma (Megatrypanum) minasense in the blood of experimentally infected squirrel monkey and marmosets.

Experimental infections by Trypanosoma (Megatrypanum) minasense were performed in primates - Saimiri sciureus and Callithrix penicillata - with the objective of searching for morphological variations of the blood trypomastigotes with respect to hosts and time of infection. We carried out morphological and morphometric analysis of blood trypomastigotes. Illustrations are given. Both the squirrel monkey and marmoset became infected after the injection of blood trypomastigotes of T. minasense, although the parasitaemia were briefer in the squirrel monkey. The parasites detected in the later host were narrower and shorter than those found in the inoculated marmoset. In the marmoset, the blood stream parasites derived from culture metacyclic trypomastigotes were considerably smaller than those derived from the inoculation of infected blood. Stronger evidence of polymorphism was found when, at the same time of infection, the blood trypomastigotes found in squirrel monkey had smaller length, body width and the distance from posterior end of the body to the kinetoplast almost four times smaller than the parasite found in the marmoset. Therefore, conflicting results on morphology and morphometry of T. minasense obtained by previous investigators could be due to polymorphism.

DNA topoisomerases: a new twist for antiparasitic chemotherapy?

The parasitic protozoa are notorious for their bizarre cellular structures and metabolic pathways, a characteristic also true for their nucleic acids. Despite these florid differences from mammalian cells, however, it has proven surprisingly difficult to devise novel chemotherapy against these pathogens. In recent years, the DNA topoisomerases from parasites have been the focus of considerable study, not only because they are intrinsically interesting, but also because they may provide a target for much-needed new antiparasitic chemotherapy.

The infection rates of trypanosomes in squirrel monkeys at two sites in the Brazilian Amazon

A study was conducted to determine the prevalence of natural infections by trypanosome species in squirrel monkeys: Samiri sciureus (Linnaeus) and Samiri ustus (Geoffroy) caught repectively near 2 hydroelectric plants: Balbina, in the State of Amazonas, and Samuel, in the State of Rondonia, Brazil. A total of 165 squirrel monkeys were examined by thick and thin smears (BS), haemocultures and xenodiagnosis: 112 monkeys, 67.9 per cent (being 52.7 per cent with mix infections) were positive to trypanosomes. Four species of trypanosomes were found in Monkeys from the 2 areas: Trypanosoma (Tejeraia) rangeli Tejera or T. rangeli-like parasites in 58 squirrel monkeys (35.2 per cent). Trypanosoma (Megatrypanum) minasense Chagas in 55 (33.3 per cent). Trypanosoma (Herpetosoma) saimirii Rodhain or T. saimirii-like parasites in 53 (32.1 per cent) and Trypanosoma (Schizotrypanum) cruzi Chagas in 17 (10.3 per cent). As T. saimirii resembles T, minasense in blood-stream trypomastigotes and T. rangeli in cultural forms and in this survey almost all monkeys presenting trypanosomes morphologically indistinguishable from T. saimirii and/or T. minasense in BS were found through xenodiagnosis and/or haemoculture to be infected by T. rangeli, we suggest that the validity of T. saimirii needs to be evaluated.

Evalucion de la prueba de aglutinacion direta en el diagnostico del mal de caderas en equinos / Evaluation of the direct agglutination test to diagnose \"Mal das Caderas\" disease

Se evaluo la utilidad de la prueba de aglutinacion directa (AD) para diagnosticar el Mal de Caderas. Se emplearon cuarenta y cuatro sueros provenientes de dos lotes de equinos naturalmente infectados con el Trypanosoma evansi (Lote 1 y Lote 2). La AD fue positiva (Aglutinacion >= 1:512) en 13 de 16 equinos (81.2 por ciento), de los que se aislaron los parasitos. En doce de estos animales (92 por ciento) se detectaron IgM anti T. evansi mediante la AD realizada con 2-mercaptoetanol (AD+2-ME). La AD fue positiva en 17 de los 28 equinos que resultaron negativos al diagnostico parasitologico....

Infeccion por T.cruci en donantes de sangre en Santa Cruz-Bolivia (Influencia de la migracion) / Infection for T.cruzi in donors of blood in Santa Cruz-Bolivia

Se realizo un estudio retrospectivo del ano 1993, de los registros del banco de sangre del hospital San Juan de Dios (HSJDD) para conocer la prevalencia de infeccion por Tripanosoma cruzi entre los donantes de sangre y sus relacion con el lugar de nacimiento. Se contabilizo el volumen total de tranfusiones en los bancos de sangre de cinco centros asistenciales. El total de transfusiones realizadas durante el ano 1993 en los centros estudiados fue de 7199. Se que en Santa Cruz se realizan por lo menos 10000 transfusiones por ano. La prevalencia de infeccion por el T.cruci entre los donantes del bancos sangre

Influencia de los cambios estacionales en la dinamica de la transmision del chagas agudo en Santa Cruz-Bolivia / Influency of thes change seasonal in the dinamic ohnthe transfusion the chagas acute in Santa Cruz-Bolivia

Este trabajo prtetende establecer, la relacion existente entre ocurrencia de casos de chagas agudo y periodos del ano. Es producto del analisis de 205 casos agudo (92 vectoriales, 50 congenitos, 24 transfusionales y 39 indeterminados), visto entre 1977 y 1991 en el Centro Nacional de Enfermedades Tropicales (CENETROP). para su analisis se excluyeron los casos indeterminados. Se observo que existe un relacionamiento significativo entre la frecuencia de casos indeterminados por la via vectorial y las estaciones y meses del ano, observandose, una mayor cantidad sobre todo en Enero y Febrero (inicio de verano), con un segundo repunte sobre todo en Octubre y algo menor en Noviembre (inicio de la primavera). Es marcada la escases de casos en junio (fines del otono). Esta observacion es mas llamativa en ciertas regiones rurales y poblaciones provinciales del Dpto. y no tanto en la ciudad de Santa Cruz de la Sierra