Breeding of African sheep reared under low-input/output smallholder production systems for trypanotolerance.

Trypanosomiasis is a disease caused by unicellular protozoan parasites. Small ruminants succumb to trypanosomiasis in areas of high tsetse fly challenge, resulting in serious economic loss often to farmers in low-input smallholder systems. At present, trypanosomiasis is treated with trypanocidal drugs, but access to these can be limited, and increasing parasite resistance raises questions about their efficacy. The development of trypanotolerance in small ruminant flocks through targeted breeding strategies is considered a sustainable and economical option for controlling African trypanosomiasis. Recently, quantitative trait loci (QTLs) associated with trypanotolerance traits in sheep have been reported. The results of these studies form the basis for more studies to identify QTLs associated with trypanosomiasis resistance, particularly in African livestock species. For example, signatures of positive selection for trypanotolerance have been identified using genome-wide single-nucleotide polymorphism data. However, there are several challenges in performing genetic analyses using data from low-input smallholder systems, including a lack of recorded pedigree and production records and the need for large sample sizes when flock sizes are often fewer than 50 animals. Breeding strategies to improve trypanotolerance should also preserve existing genetic diversity as well as minimize excessive genetic introgression by trypanosusceptible breeds. This review discusses the possibilities of breeding for trypanosome tolerance/resistance in low-input/low-output small ruminant production systems. Potential challenges are outlined, and potential available genetic resources are described as a foundation for future work.

Repurposing Carvedilol as a Novel Inhibitor of the Trypanosoma cruzi Autophagy Flux That Affects Parasite Replication and Survival.

T. cruzi, the causal agent of Chagas disease, is a parasite able to infect different types of host cells and to persist chronically in the tissues of human and animal hosts. These qualities and the lack of an effective treatment for the chronic stage of the disease have contributed to the durability and the spread of the disease around the world. There is an urgent necessity to find new therapies for Chagas disease. Drug repurposing is a promising and cost-saving strategy for finding new drugs for different illnesses. In this work we describe the effect of carvedilol on T. cruzi. This compound, selected by virtual screening, increased the accumulation of immature autophagosomes characterized by lower acidity and hydrolytic properties. As a consequence of this action, the survival of trypomastigotes and the replication of epimastigotes and amastigotes were impaired, resulting in a significant reduction of infection and parasite load. Furthermore, carvedilol reduced the whole-body parasite burden peak in infected mice. In summary, in this work we present a repurposed drug with a significant in vitro and in vivo activity against T. cruzi. These data in addition to other pharmacological properties make carvedilol an attractive lead for Chagas disease treatment.

Prevalence and associated risk factors of bovine trypanosomosis in tsetse suppression and non-suppression areas of South Omo Zone, Southwestern Ethiopia.

A cross-sectional study aimed to elucidate the prevalence of bovine trypanosomosis and its potential risk factors was conducted in tsetse suppression and non-suppression areas of South Omo Zone, Southern Ethiopia from November 2018- May 2019. A total of 1284 blood samples from local zebu cattle (642 each in dry and wet season) were examined by using buffy coat technique and thin blood smear method. The overall prevalence was 11.05 % with 14.33 % in dry and 7.78 % in wet season. According to multiple logistic regression analysis of tsetse suppression areas, higher prevalence in female than male (OR = 0.48, 95 % CI: 0.27, 0.83), in poor (OR = 3.25, 95 % CI: 1.26, 11.09) and medium (OR = 2.07, 95 % CI: 0.74, 7.37) than good body conditioned animals was recorded. Moreover, tethered animals (OR = 2.07, 95 % CI: 1.06, 3.92) were more likely to be infected than communal grazers and also higher prevalence in dry season than wet season (OR = 0.52, 95 % CI: 0.30, 0.87). Similarly, in tsetse non-suppression areas, higher prevalence in female than male (OR = 0.48, 95 % CI: 0.27, 0.85) and in wet season (OR = 0.41, 95 % CI: 0.23, 0.7) than dry season was recorded. Trypanosoma congolense and Trypanosoma vivax were found in cattle with the former more prevalent in both areas. Overall pooled mean packed cell volume (PCV) of parasitaemic animals (23.57 ± 3.13) was significantly lower than aparasitaemic animals (27.80 ± 4.95). Similarly, parasitaemic animals from tsetse suppression areas and tsetse non-suppression areas had significantly lower mean PCV than their aparasitaemic counterparts. Mean PCV of T. congolense (23.59 ± 3.22) infected animals was not different (P > 0.05) from T. vivax infected animals (23.26 ± 3.31). It was also indicated that the probability of anaemic animals to be parasitaemic was significantly higher (P < 0.05) than non-anaemic animals in both areas. In conclusion, the prevalence of trypanosomosis revealed its endemicity which bottlenecked the livestock production and productivity in the study area despite of tsetse suppression activities. Therefore, integrated parasite and vector control approach should be undertaken to curve the disease.

Identification of Trypanosoma cruzi Polyamine Transport Inhibitors by Computational Drug Repurposing.

Trypanosoma cruzi is the causative agent of Chagas disease, a parasitic infection endemic in Latin America. In T. cruzi the transport of polyamines is essential because this organism is unable to synthesize these compounds de novo. Therefore, the uptake of polyamines from the extracellular medium is critical for survival of the parasite. The anthracene-putrescine conjugate Ant4 was first designed as a polyamine transport probe in cancer cells. Ant4 was also found to inhibit the polyamine transport system and produced a strong trypanocidal effect in T. cruzi. Considering that Ant4 is not currently approved by the FDA, in this work we performed computer simulations to find trypanocidal drugs approved for use in humans that have structures and activities similar to Ant4. Through a similarity ligand-based virtual screening using Ant4 as reference molecule, four possible inhibitors of polyamine transport were found. Three of them, promazine, chlorpromazine, and clomipramine, showed to be effective inhibitors of putrescine uptake, and also revealed a high trypanocidal activity against T. cruzi amastigotes (IC50 values of 3.8, 1.9, and 2.9 µM, respectively) and trypomastigotes (IC50 values of 3.4, 2.7, and 1.3 µM, respectively) while in epimastigotes the IC50 were significantly higher (34.7, 41.4, and 39.7 µM, respectively). Finally, molecular docking simulations suggest that the interactions between the T. cruzi polyamine transporter TcPAT12 and all the identified inhibitors occur in the same region of the protein. However, this location is different from the site occupied by the natural substrates. The value of this effort is that repurposing known drugs in the treatment of other pathologies, especially neglected diseases such as Chagas disease, significantly decreases the time and economic cost of implementation.

Clinical and therapeutic aspects of an outbreak of canine trypanosomiasis / Aspectos clínicos e terapêuticos de um surto de tripanossomíase canina

Abstract Trypanosomiasis caused by Trypanosoma evansi can seriously affect both domestic and wild animals. This article reports on an outbreak of canine trypanosomiasis on a farm in the Pantanal region of Brazil. The farm had 38 dogs, 20 of which died before receiving veterinary care. The remaining 18 dogs were underwent anamnesisn, clinical examination, hematological and biochemical evaluations. Blood smears and PCR analysis were performed for the diagnosis. The treatment protocols used according to the clinical recovery or parasitological cure of the dogs, using diminazene diaceturate, isometamidium chloride or quinapyramine sulfate. Post-treatment parasitological evaluation was performed by the microhematocrit technique. 7/18 dogs were PCR positive for T. evansi (confirmed by sequencing). There was clinical findings, which were consistent with both the acute and chronic stages of the disease in dogs. The infected dogs all exhibited at least one clinical sign of the disease. The hematological findings were compatible with trypanosomiasis, highlighting the hypochromic microcytic anemia as the main outcome. No treatment protocol was fully effective and the prolonged use of diminazene diaceturate caused the death of an animal. The trypanosomiasis can cause high rates of morbidity and mortality in dogs and difficulty in establishment an effective and safe therapeutic protocol.

Resumo A tripanossomíase causada por Trypanosoma evansi pode acometer gravemente os animais domésticos e selvagens. Este artigo relata um surto de tripanossomíase canina em uma fazenda na região do Pantanal, Brasil. Na fazenda havia 38 cães, 20 dos quais morreram antes de receber cuidados veterinários. Os 18 cães restantes foram submetidos a anamnese, exame clínico, avaliação hematológica e bioquímica. Esfregaços de sangue e análise da PCR foram realizados para o diagnóstico. Os protocolos de tratamento foram utilizados de acordo com a recuperação clínica ou cura parasitológica dos cães, utilizando diaceturato de diminazeno, cloreto de isometamídio ou sulfato de quinapiramina. A avaliação parasitológica pós-tratamento foi realizada pela técnica de microhematócrito. 7/18 cães foram PCR positivos para T. evansi (confirmado por sequenciamento). Os achados clínicos encontrados, foram consistentes com os estágios agudo e crônico da doença em cães. Todos os cães infectados exibiram pelo menos um sinal clínico da doença. Os achados hematológicos foram compatíveis com a tripanossomíase, destacando a anemia microcítica hipocrômica como principal consequência. Nenhum protocolo de tratamento foi totalmente eficaz e o uso prolongado de diaceturato de diminazeno causou a morte de um animal. A tripanossomíase pode causar altas taxas de morbidade e mortalidade em cães e dificultar o estabelecimento de um protocolo terapêutico eficaz e seguro.

Autophagic-related cell death of Trypanosoma brucei induced by bacteriocin AS-48.

The parasitic protozoan Trypanosoma brucei is the causative agent of human African trypanosomiasis (sleeping sickness) and nagana. Current drug therapies have limited efficacy, high toxicity and/or are continually hampered by the appearance of resistance. Antimicrobial peptides have recently attracted attention as potential parasiticidal compounds. Here, we explore circular bacteriocin AS-48's ability to kill clinically relevant bloodstream forms of T. brucei gambiense, T. brucei rhodesiense and T. brucei brucei. AS-48 exhibited excellent anti-trypanosomal activity in vitro (EC50 = 1-3 nM) against the three T. brucei subspecies, but it was innocuous to human cells at 104-fold higher concentrations. In contrast to its antibacterial action, AS-48 does not kill the parasite through plasma membrane permeabilization but by targeting intracellular compartments. This was evidenced by the fact that vital dye internalization-prohibiting concentrations of AS-48 could kill the parasite at 37 °C but not at 4 °C. Furthermore, AS-48 interacted with the surface of the parasite, at least in part via VSG, its uptake was temperature-dependent and clathrin-depleted cells were less permissive to the action of AS-48. The bacteriocin also caused the appearance of myelin-like structures and double-membrane autophagic vacuoles. These changes in the parasite's ultrastructure were confirmed by fluorescence microscopy as AS-48 induced the production of EGFP-ATG8.2-labeled autophagosomes. Collectively, these results indicate AS-48 kills the parasite through a mechanism involving clathrin-mediated endocytosis of VSG-bound AS-48 and the induction of autophagic-like cell death. As AS-48 has greater in vitro activity than the drugs currently used to treat T. brucei infection and does not present any signs of toxicity in mammalian cells, it could be an attractive lead compound for the treatment of sleeping sickness and nagana.

Rapid test for the evaluation of the activity of the prodrug hydroxymethylnitrofurazone in the processing of Trypanosoma cruzi messenger RNAs

No fully effective treatment has been developed since the discovery of Chagas' disease by Carlos Chagas in 1909. Since drug-resistant Trypanosoma cruzi strains are occurring and the current therapy is effectiveness in the acute phase but with various adverse side effects, more studies are needed to characterize the susceptibility of T. cruzi to new drugs. Many natural and/or synthetic substances showing trypanocidal activity have been used, even though they are not likely to be turned into clinically approved drugs. Originally, drug screening was performed using natural products, with only limited knowledge of the molecular mechanism involved in the development of diseases. Trans-splicing, which is unusual RNA processing reaction and occurs in nematodes and trypanosomes, implies the processing of polycistronic transcription units into individual mRNAs; a short transcript spliced leader (SL RNA) is trans-spliced to the acceptor pre-mRNA, giving origin to the mature mRNA. In the present study, permeable cells of T. cruzi epimastigote forms (Y, BOL and NCS strains) were treated to evaluate the interference of two drugs (hydroxymethylnitrofurazone - NFOH-121 and nitrofurazone) in the trans-splicing reaction using silver-stained PAGE analysis. Both drugs induced a significant reduction in RNA processing at concentrations from 5 to 12.5 æM. These data agreed with the biological findings, since the number of parasites decreased, especially with NFOH-121. This proposed methodology allows a rapid and cost-effective screening strategy for detecting drug interference in the trans-splicing mechanism of T. cruzi.