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The present research aimed to document the incidence, clinical signs, haematological, and serum biochemical alterations, as well as electrocardiography and echocardiography findings in 62 buffaloes (selected from a total of 240) infected with Trypanosoma evansi. The study spanned one year, from January 2022 to December 2022. Morphological identification of Trypanosoma evansi was done by the presence of a centrally positioned nucleus with a small sub-terminal kinetoplast at the posterior position through microscopic examination of Giemsa stained peripheral blood smears. The incidence of trypanosomosis were determined to be 26% (62/240) using stained blood smear examination and 41% (98/240) through polymerase chain reaction assay. Clinical signs exhibited by buffaloes with trypanosomosis included the lack of rumination (94%; 58/62), anorexia (90%; 56/62), emaciation (87%; 54/62), loss of milk yield (84%; 52/62), ocular discharges (82%; 51/62), depressed demeanour (81%; 50/62), sunken eye balls (61%; 38/62), fever (60%; 37/62), scleral congestion (56%; 35/62) and intermittent fever (42%; 26/62). Cardiovascular clinical findings in affected buffaloes included tachycardia (44%; 27/62), cardiac arrhythmia (24%; 15/62), cardiac murmurs (19%; 12/62) and muffled heart sounds (18%; 11/62). In the present study, buffaloes with trypanosomosis exhibited significant reduction in haemoglobin (p = 0.008), packed cell volume (p = 0.004), total erythrocyte count (p = 0.003), mean corpuscular volume (p = 0.042), total leucocyte count (p = 0.048) and absolute neutrophil count (p = 0.012); a significant increase in absolute eosinophil count (p = 0.011) and absolute monocyte count (p = 0.008) compared to the apparently healthy buffaloes. Additionally significant decrease in albumin (p = 0.001), A/G ratio (p = 0.007), calcium (p = 0.008), glucose (p = 0.007), phosphorous (p = 0.048), sodium (p = 0.008), potassium (p = 0.041) and chloride (p = 0.046) were observed in buffaloes with trypanosomosis compared to healthy ones. Buffaloes with trypanosomosis also showed significant increase in globulin (p = 0.004), aspartate aminotransferase (p = 0.008), bilirubin (p = 0.034), blood urea nitrogen (p = 0.071), creatinine (p = 0.029), cholesterol (p = 0.046), lactate dehydrogenase (p = 0.009), gamma-glutamyl transferase (p = 0.004) and creatine kinase-myoglobin binding levels (p = 0.005). Electrocardiography explorations in buffaloes with trypanosomosis revealed sinus tachycardia, low voltage QRS complex, ST segment elevation, wide QRS complex, sinus arrhythmia, sinus bradycardia, wandering pace maker, first degree atrio ventricular block, biphasic T wave and tall T wave. Echocardiography examination unveiled cardiac chamber dilatation, ventricular wall thickening and indications of pericarditis/cardiac tamponade. Necropsy was carried on the dead buffaloes during the study period disclosed severely congested blood vessels on epicardial surface, endocardial haemorrhages, and presence of pericardial fluid. Histopathological examination of the heart revealed hyaline degeneration, haemorrhages in the cardiac muscles and varying degrees of degenerative changes. Additionally, the pericardium displayed increased thickness due to presence of more elastic fibres, fibroblast cells in the myocardium, discontinuity of muscle layers, vascular congestion, perivascular mono nuclear cell infiltration and augmented thickness of the endocardium with fibroblast cell proliferation. The study's conclusion highlights cardiac alterations as secondary complications in buffaloes infected with Trypanosoma evansi. Further investigations are recommended to elucidate therapeutic modifications and refine the treatment paradigm., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Many eukaryotic cell-surface proteins are post-translationally modified by a glycosylphosphatidylinositol (GPI) moiety that anchors them to the cell membrane. The biosynthesis of GPI anchors is initiated in the endoplasmic reticulum by transfer of GlcNAc from UDP-GlcNAc to phosphatidylinositol. This reaction is catalyzed by GPI GlcNAc transferase, a multisubunit complex comprising the catalytic subunit Gpi3/PIG-A as well as at least five other subunits, including the hydrophobic protein Gpi2, which is essential for the activity of the complex in yeast and mammals, but the function of which is not known. To investigate the role of Gpi2, we exploited Trypanosoma brucei (Tb), an early diverging eukaryote and important model organism that initially provided the first insights into GPI structure and biosynthesis. We generated insect-stage (procyclic) trypanosomes that lack TbGPI2 and found that in TbGPI2-null parasites, (i) GPI GlcNAc transferase activity is reduced, but not lost, in contrast with yeast and human cells, (ii) the GPI GlcNAc transferase complex persists, but its architecture is affected, with loss of at least the TbGPI1 subunit, and (iii) the GPI anchors of procyclins, the major surface proteins, are underglycosylated when compared with their WT counterparts, indicating the importance of TbGPI2 for reactions that occur in the Golgi apparatus. Immunofluorescence microscopy localized TbGPI2 not only to the endoplasmic reticulum but also to the Golgi apparatus, suggesting that in addition to its expected function as a subunit of the GPI GlcNAc transferase complex, TbGPI2 may have an enigmatic noncanonical role in Golgi-localized GPI anchor modification in trypanosomes., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

The parasite Trypanosoma brucei is the main cause of the sleeping sickness threatening millions of populations in many African countries. The parasitic infection is currently managed by some synthetic medications, most of them suffer limited activity spectrum and/or serious adverse effects. Some studies have pointed out the promising therapeutic potential of the plant extracts rich in polyphenols to curb down parasitic infections caused by T. brucei and other trypanosomes. In this work, the main components dominating Eugenia uniflora and Syzygium samarangense plant extracts were virtually screened, through docking, as inhibitors of seven T. brucei enzymes validated as potential drug targets. The in vitro and in vivo anti-T. brucei activities of the extracts in two treatment doses were evaluated. Moreover, the extract effects on the packed cell volume level, liver, and kidney functions were assessed. Five compounds showed strong docking and minimal binding energy to five target enzymes simultaneously and three other compounds were able to bind strongly to at least four of the target enzymes. These compounds represent lead hits to develop novel trypanocidal agents of natural origin. Both extracts showed moderate in vitro anti-trypanosomal activity. Infected animal groups treated over 5 days with the studied extracts showed an appreciable in vivo anti-trypanosomal activity and ameliorated in a dose dependent manner the anaemia, liver, and kidney damages induced by the infection. In conclusion, Eugenia uniflora and Syzygium samarangense could serve as appealing sources to treat trypanosomes infections., (Copyright © 2021 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

A 2.5-year-old male German Shepherd was presented to a private veterinary clinic in Hanoi, Vietnam showing anorexia, weakness, lethargy, reluctant to go for walks with a recent history of intermittent fever. Clinical examination of the dog showed pale mucous membrane, impaired eyesight, edema of the back legs. Complete blood count revealed severe anemia; red blood cell 3.8 × 10 12 /l, hemoglobin 8.7 g/dl, hematocrit 26.4%, associated with thrombocytopenia 145 × 10 9 /l. Biochemical analysis showed a moderate increase of alanine transaminase (150.7 UI/l) and alkaline phosphatase activities (266 UI/I) with mild hypoglycemia (71.46 mg/dl). Trypanosoma evansi was observed in Giemsa-stained blood smears under microscopic observation which was confirmed by PCR. This is the first report of canine trypanosomiasis caused by T. evansi in Vietnam., (Copyright © 2020 Elsevier B.V. All rights reserved.)

This study reports the first autochthonous "surra" outbreak in horses in the State of Santa Catarina, southern Brazil. Six horses with clinical suspicion of trypanosomosis had the natural infection by T. evansi confirmed by PCR and rapid serum agglutination test. Clinical, parasitological, and hematology evaluations were performed at initial observation (T0) and 90 days after (T1). At T0, all animals that tested positive for T. evansi in PCR presented with severe clinical signs and out of normal range hematological hematological (hematocrit, leukocytes, platelets, hemoglobin) and serum biochemical parameters (alanine aminotransferase (ALT), creatinine phosphokinase (CPK), lactate dehydrogenase (LDH), creatinine, bilirubin, and glucose). At T1, the progressive clinical recovery of animals, normalization of hemato-biochemical parameters, and negative PCR results for T. evansi were observed. It was not possible to identify the vector/mechanism of transmission through which animals were infected; therefore, the implementation of surveillance and control measures is essential to prevent the spread of this disease in horse herds, as well as to other animal species., Competing Interests: Declaration of Competing Interest The authors declared no conflict of interest., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Leishmania infantum is a trypanosomatid that causes parasitic dermatopathy in dogs. Trypanosoma caninum is another trypanosomatid, which infects the skin of dogs, although cutaneous abnormalities are absent. This study aimed to investigate the occurrence of T. caninum infection and its associated cutaneous and histological changes and compare it with the occurrence of L. infantum infection in dogs. The study included 150 dogs, of which T. caninum infection was identified in 3 (2%) and L. infantum infection in 15 (10%) of them, with no association (p>0.05) of these infections with the breed, gender, age, or cutaneous abnormalities. The cutaneous abnormalities were based on 1 (4.8%) and 12 (57.1%) dogs infected by T. caninum and L. infantum, respectively. The dermatohistopathological abnormalities in the dogs infected with T. caninum included mild perivascular lymphohistioplasmacytic infiltrates in the clinically asymptomatic ones, while in those with dermatological abnormalities, acanthosis, epidermal orthokeratotic hyperkeratosis, melanomacrophages, and co-infection with Microsporum sp. and Trichophyton sp. were observed. InL. infantum infected, the histopathological findings included chronic granulomatous inflammatory infiltrates and structures compatible with amastigotes. Despite the low frequency of T. caninum infection, our findings suggest that this trypanosomatid, unlike L. infantum, does not cause any macroscopic skin abnormalities.

The aim of this study was to evaluate the effect of trypanosomes on cultured largemouth bass (Micropterus salmoides) and describe the taxonomic identification of the parasite. The effects of the parasite on M. salmoides were examined based on clinical symptoms, hemograms, histopathology, and serum biochemistry. Diseased fish showed typical clinical symptoms of trypanosomiasis, which included lethargy, anorexia, and histopathological lesions in the liver, head kidney, and spleen. The serum of diseased fish had significantly lower concentrations of glucose, triglyceride, and low-density lipoprotein, and significantly higher alanine transaminase (ALT), aspartate transaminase (AST), and lactate dehydrogenase (LDH) activities. The morphology of the trypanosomes was also analyzed using light microscopy, and their 18S rDNA sequence was analyzed to establish genetic relationships with other known strains. We found that the trypomastigote form of the trypanosomes from M. salmoides was similar to those isolated from Pelteobagrus fulvidraco. The trypanosomes had a slender and narrow body with a relatively long free flagellum, not well-developed undulating membrane, and an oval kinetoplast located near the subterminal posterior end of the body. The 18S rDNA sequences of the trypanosome from M. salmoides had the highest similarity (99.8%) with that of P. fulvidraco, suggesting they are identical species. Based on the differences in morphological characteristics and 18S rDNA sequence compared to trypanosomes isolated from other freshwater fish, it is considered as a new species and we propose the name Trypanosoma micropteri n. sp.

2-Amino-benzo[ d]thiazole was identified as a new scaffold for the development of improved pteridine reductase-1 (PTR1) inhibitors and anti-trypanosomatidic agents. Molecular docking and crystallography guided the design and synthesis of 42 new benzothiazoles. The compounds were assessed for Trypanosoma brucei and Leishmania major PTR1 inhibition and in vitro activity against T. brucei and amastigote Leishmania infantum. We identified several 2-amino-benzo[ d]thiazoles with improved enzymatic activity ( TbPTR1 IC 50 = 0.35 μM; LmPTR1 IC 50 = 1.9 μM) and low μM antiparasitic activity against T. brucei. The ten most active compounds against TbPTR1 were able to potentiate the antiparasitic activity of methotrexate when evaluated in combination against T. brucei, with a potentiating index between 1.2 and 2.7. The compound library was profiled for early ADME toxicity, and 2-amino- N-benzylbenzo[ d]thiazole-6-carboxamide (4c) was finally identified as a novel potent, safe, and selective anti-trypanocydal agent (EC 50 = 7.0 μM). Formulation of 4c with hydroxypropyl-β-cyclodextrin yielded good oral bioavailability, encouraging progression to in vivo studies.

Trypanosoma carassii is a flagellated bloodstream parasite of cyprinid fish with pathogenesis manifesting primarily as anemia in experimentally infected fish. This anemia is characterized by decreases in the number of circulating red blood cells (RBCs) during peak parasitemia. We examined changes in the key blood metrics and expression of genes known to be important in the regulation of erythropoiesis. Increasing parasitemia was strongly correlated with an overall decrease in the total number of circulating RBCs. Gene expression of key erythropoiesis regulators (EPO, EPOR, GATA1, Lmo2, and HIFα) and proinflammatory cytokines (IFNγ and TNFα) were measured and their expressions differed from those in fish made anemic by injections of phenylhydrazine (PHZ). Significant upregulation of pro-erythropoietic genes was observed in PHZ-induced anemia, but not during peak parasitic infection. Previously, we reported on functional characterization of goldfish erythropoietin (rgEPO) and its ability to induce survival and differentiation of erythroid progenitor cells in vitro. Treatment of goldfish during the infection with rgEPO reduced the severity of anemia but failed to fully prevent the onset of the anemic state in infected fish. Proinflammatory cytokines have been implicated in the suppression of erythropoiesis during trypanosomiasis, specifically the cytokines TNFα, IFNγ, and IL-1β. Analysis of key proinflammatory cytokines revealed that mRNA levels of IFNγ and TNFα were upregulated in response to infection, but only TNFα increased in response to PHZ treatment. Synergistic activity of the proinflammatory cytokines may be required to sustain prolonged anemia. These findings provide insight into the relationship between T. carassii and host anemia and suggest that T. carassii may directly or indirectly suppress host erythropoiesis.

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.

Hemoparasitic diseases like trypanosomiasis have an adverse influence on the health and working capability of infected animals. Monitoring and identification of blood born parasitic infections in dairy animals are of vital importance to get the optimum production. In this study blood samples were collected from Nili Ravi buffaloes (n = 390) kept at different villages of district Lodhran, Punjab province of Pakistan. Blood samples were evaluated for red blood cell counts, total and differential leukocyte counts, hematocrit, hemoglobin, total proteins and different serum parameters such as alanine aminotransferase, aspartate aminotransferase, malondialdehyde, alkaline phosphatase, lactate dehydrogenase, phosphorous, copper, calcium and magnesium. Overall prevalence of Trypanosoma evansi was 4.61% based on microscopic smear examination, 11.02% with Formol Gel Test and 16.15% with PCR. Infected buffaloes showed different clinical signs, including high fever (105 ± 1.0 °F), edema of face and legs, hyperemic mucosa of eyes, lachrymation, bulging eyes, pale mucus membranes and frequent urination. Microscopic examination of blood films showed morphologically different parasites. Statistical analysis did not indicate an association of infection based on age and sex of buffaloes. Results revealed significantly (p < 0.05) lower values of red blood cell counts, hemoglobin, hematocrit, mean corpuscular hemoglobin concentration and total proteins, while increased values of mean corpuscular volume, total white blood cells, monocyte, neutrophils, basophils and eosinophils in infected animals. Infected buffaloes were suffering from macrocytic hypochromic anemia. A significant (p < 0.05) increase in serum lipid per oxidation product (malondialdehyde) level and serum enzymes while a decrease in macrominerals and trace mineral (copper) in trypanosomiasis positive buffaloes were recorded. It was concluded that Trypanosoma evansi is prevalent in Pakistan under tropical and subtropical climatic conditions. It causes clinical disease with macrocytic hypochromic anemia and oxidative stress in infected buffaloes., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Small populations of Virginia opossum (Didelphis virginiana) in western Mexico are endangered by hunting and natural predators as well as by different kinds of diseases. After two serological analyses using Serodia® latex particle agglutination and indirect haemagglutination (IHA) tests, 35 (53.03%) of 66 collected opossums in two small towns in western Mexico were positive for the presence of Trypanosoma cruzi. Twenty-eight of the 35 seropositive opossums had pathological lesions: 11 had changes in only one organ, 13 in two organs, and four had pathological changes in three organs. Splenomegaly was the most common finding in the examined opossums, followed by hepatomegaly. These potentially fatal pathological changes could contribute to the scarcity of the opossum population, even leading to the extinction of this species in western Mexico.

The blood protozoan Trypanosoma evansi, which is transmitted by biting flies, is frequently neglected due to subclinical infections. This report describes a case of trypanosomiasis due to T. evansi in a 9-yr-old male puma (Felis concolor) housed at the Lahore Zoo in Pakistan. Early in January 2015, this male puma presented with chronic lethargy, weight loss, incoordination, hyperthermia, anorexia, sunken eyes, and unthriftiness. Microscopic examination of Giemsa-stained blood smears showed numerous Trypanosoma parasites. The puma was treated with diminazene aceturate subcutaneously twice. A few days later, a blood smear examination showed absence of trypanosomes. Five months later the cat presented with acute epistaxis and died. Postmortem examination showed emaciation, pale liver and kidneys, and hemorrhages on the spleen. Examination of a blood smear taken at the time of death showed numerous Trypanosoma parasites. PCR testing confirmed the presence of Trypanosoma DNA. DNA sequencing of two amplicons confirmed the presence of Trypanosoma in the blood smears with a 98-99% identity with the previously identified GenBank sequences. A phylogenetic tree was then constructed. Further studies are needed to improve our knowledge about the epidemiology and pathogenesis of T. evansi infection in wild animal species.

The aim of this study was to evaluate whether the treatment with Achyrocline satureioides essential oil-loaded in nanocapsules (AS-NC) is able to protect the hepatic tissue against cytotoxic damage caused by Trypanosoma evansi. Thus, the rats were divided into four groups (n = 6 per group): uninfected/saline, uninfected/AS-NC, infected/saline, and infected/AS-NC. At day 4 post-infection (PI), the animals were euthanized and liver and sera samples were collected to perform the hepatic cell viability assay, and to determine seric levels of reactive oxygen species (ROS) and nitric oxide metabolites (NOx). Cell viability decreased (p < 0.05) in the infected/saline group compared to uninfected/saline group, while the treatment with AS-NC avoided this alteration in infected rats. Seric ROS and NOx levels increased (p < 0.05) in the infected/saline group compared to uninfected/saline group, while the treatment with AS-NC avoided this effect on ROS levels of infected rats. In summary, the treatment with AS-NC was able to protect the liver tissue against the cytotoxic effect caused by the parasite by avoiding exacerbated production of ROS., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Aim of the present study was to assess the cytokine gene expression in liver, kidney and spleen and histopathological changes in mice infected with buffalo and dog isolates of Trypanosoma evansi. Forty-four Swiss albino mice was divided into eleven groups of four mice each and injected subcutaneously with 1 × 10 5 trypanosomes of buffalo and dog isolate to twenty mice each, four mice served as control. Mice were examined for clinical signs, blood smear for trypanosome counts. Blood for PCR, liver, kidney, spleen, heart, lung, testis and abdominal muscle for histopathology and liver, kidney, spleen for cytokine gene expression studies, were collected. Mice showed dullness, lethargy, hunched back, sluggish movements on D4 and D5 in buffalo and dog isolate, respectively. Parasite count in blood varied between the two isolates of T. evansi. By PCR, trypanosome DNA was detected on D1 and D2 for buffalo and dog isolate, respectively. Splenomegaly was observed in mice infected with buffalo isolate but not with dog isolate. Histopathological changes were observed in liver, kidney, spleen and heart of mice but no changes in testis and abdominal muscles. Blood vessels of liver, heart, lung showed presence of trypanosomes in mice infected with buffalo isolate but not for dog isolate. Cytokine gene expression of IL-2, IL-4, IL-6, IL-12, TNF-α and IFN-γ increased in liver, kidney and spleen in both these isolates. However, the buffalo isolate exhibited pronounced increase in cytokine gene expression when compare to dog isolate of T. evansi. Anti-inflammatory cytokine gene IL-10 showed 50-60 and 10-20 folds increment in buffalo and dog isolates, respectively. This is the first report of IL-4, IL-6, IL-10 and IL-12 cytokine changes in mice infected with T. evansi. A variation in pathogenicity between buffalo and dog isolates was recorded indicating buffalo isolate of T. evansi remained more pathogenic in mice., (Copyright © 2016 Elsevier Inc. All rights reserved.)

During infection in mammals, the protozoan parasite Trypanosoma brucei transforms from a proliferative bloodstream form to a quiescent form that is pre-adapted to host transition. AMP analogs are known to induce quiescence and also inhibit TbTOR4. To examine the role of AMP-activated kinase (AMPK) in the regulation of this developmental transition, we characterized trypanosome TbAMPK complexes. Expression of a constitutively active AMPKα1 induces quiescence of the infective form, and TbAMPKα1 phosphorylation occurs during differentiation of wild-type pleomorphic trypanosomes to the quiescent stumpy form in vivo. Compound C, a well-known AMPK inhibitor, inhibits parasite differentiation in mice. We also provide evidence linking oxidative stress to TbAMPKα1 activation and quiescent differentiation, suggesting that TbAMPKα1 activation balances quiescence, proliferation, and differentiation., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

The aim of this study was to evaluate the oxidative stress variables, and enzymes of cholinergic (acetylcholinesterase (AChE) and butyrylcholinesterase (BChE)) and adenosinergic (adenosine deaminase (ADA)) systems in renal tissue, as well as the relationship between these systems and lipid peroxidation. The animals were divided into two groups with six animals each: uninfected (negative control) and infected (positive control). On day 4 post-infection (PI), animals were euthanized and the kidney was collected. Thiobarbituric acid reactive species (TBARS) and lipid peroxidation (FOX) levels increased, while the catalase (CAT), AChE, BChE and ADA activities decreased in kidney tissue on day 4 PI. A negative correlation between AChE and TBARS (r = -0.750), AChE and FOX (r = -0.650), as well as ADA and TBARS (r = -0.345) and ADA and FOX (r = -0.540) were observed (p < 0.05). In summary, the T. evansi infection cause lipid peroxidation in the renal tissue, altering the antioxidant-oxidant status, alterations compatible to oxidative stress and oxidative damage. Also, the T. evansi decrease the activities of AChE, BChE and ADA in order to reduce the oxidative damage increasing the levels of ACh, BCh and adenosine. These alterations in the kidney may be contribute on pathophysiology of T. evansi infection., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Trypanosoma evansi is a widely-distributed haemoflagellated parasite of veterinary importance that infects a variety of mammals including horses, mules, camels, buffalos, cattle and deer. It is the causal agent of a trypanosomiasis known as Surra which produces epidemics of great economic importance in Africa, Asia and South America. The main pathology includes an enlarged spleen with hypertrophy of lymphoid follicles, congested lungs, neuronal degeneration and meningoencephalitis, where migration of the parasites from the blood to the tissues is essential. Most cells, including pathogenic cells, use diverse strategies for tissue invasion, such as the expression of surface receptors to bind plasminogen or plasmin. In this work, we show that T. evansi is able to bind plasminogen and plasmin on its surface. The analysis of this binding revealed a high affinity dissociation constant (Kd of 0.080±0.009μM) and 1×10(5) plasminogen binding sites per cell. Also a second population of receptors with a Kd of 0.255±0.070μM and 3.2×10(4) plasminogen binding sites per cell was determined. Several proteins with molecular masses between ∼18 and ∼70kDa are responsible for this binding. This parasite-plasminogen interaction may be important in the establishment of the infection in the vertebrate host, where the physiological concentration of available plasminogen is around 2μM., (Copyright © 2016 Elsevier B.V. All rights reserved.)

This study was conducted to investigate the occurrence of oxidative stress in the heart tissue of rats infected with Trypanosoma evansi. Rats were divided into 2 groups (A and B) with 12 animals each, and further subdivided into 4 subgroups (A1 and A2, 6 animals/each; and B1 and B2, 6 animals/each). Animals in the groups B1 and B2 were subcutaneously inoculated with T. evansi. Thiobarbituric acid reactive substances (TBARS), superoxide dismutase activity (SOD), glutathione S-transferase activity (GST), reduced glutathione activity (GSH), and non-protein thiols (NPSH) in the heart tissue were evaluated. At day 5 and 15 post-infection (PI), an increase in the TBARS levels and a decrease in the SOD activity (P<0.05) were observed. GSH and GST activities were decreased in infected animals at day 15 PI (P<0.05). Considering the proper functioning of the heart, it is possible that the changes in the activity of these enzymes involved in the oxidative stress may be related, at least in part, in the pathophysiology of rats infected with T. evansi.

Twenty pregnant Yankasa ewes were assigned to three groups to determine the effect of Trypanosoma evansi infection on pregnancy. Groups A and B comprising seven ewes each were infected with approximately 1.0 × 10(6) cells of T evansi per ewe through venepuncture at the second and third trimesters of pregnancy, respectively. Group C comprising six ewes served as uninfected control. There was slight pyrexia in the infected groups (groups A and B) but was absent in group C. The mean body weight, glucose concentration, and packed cell volume of ewes in group A were not significantly different from those in group C throughout the study. There was also no significant difference in mean glucose concentration between groups B and C. However, in group B, mean body weight was significantly (P < 0.05) lower compared to group C at week 2 and from week 4 post infection (pi) till the end of the study; the packed cell volume also significantly (P < 0.05) decreased but at weeks 4 and 6 pi. The mean plasma protein of ewes in group A was significantly (P < 0.05) increased compared to those of group C at weeks 7, 11 pi and thereafter till the end of the study. On the contrary, the plasma protein of ewes in group B decreased significantly (P < 0.05) compared to those in group C at weeks 2 and 6 pi. There were no reproductive losses throughout the study. This was characterized by insignificant differences in the gestation length between ewes in the infected groups (groups A and B) compared with those in group C. However, there were significant (P < 0.05) decreases in lamb birth weights of ewes in group B compared with ewes in groups A and C. Mice inoculation with blood from infected ewes postpartum was parasitemic 18 to 25 days pi, for ewes in group B, whereas none of the mice in groups A and C were parasitemic. Lambs born from the infected groups (groups A and B) were also aparasitemic for 40 days postpartum. It was therefore concluded that the T evansi isolate used caused mild trypanosomosis when infected at third trimester, whereas ewes infected at second trimester were resistant., (Copyright © 2016 Elsevier Inc. All rights reserved.)

A study was made to determine the prevalence of camel trypanosomosis (surra) and its associated risk factors in Borena zone, southern Ethiopia during 2013-2014. A total of 2400 blood samples were collected and examined by the buffy coat and thin blood film laboratory methods, and data were analyzed using the SPSS statistical software. The overall prevalence of camel trypanosomosis in the area was found to be 2.33 %. Prevalence was significantly different among the surveyed districts (P = 0.000), the pastoral associations (F = 6.408, P = 0.000), altitudinal divisions (P = 0.000), age groups (P = 0.034), and between animals possessing packed cell volume (PCV) values greater than 25 % and less than 25 % (P = 0.000); whereas, prevalence of the disease was not statistically significantly different between the sexes (P = 0.311) and among the body condition score groups (P = 0.739). The PCV of trypanosome positive and trypanosome negative camels differ significantly (P = 0.001), and prevalence of trypanosomosis was seen to be negatively correlated with packed cell volume (r = -0.069, P = 0.000) revealing the effect of camel trypanosomosis on anemia state of parasitized animals. In conclusion, camel trypanosomosis is a serious and economically important disease hampering camel production and productivity in southern Ethiopia. Further studies involving more sensitive molecular techniques to reveal the precise magnitude of the disease and to identify the vector species of the parasite are recommended.

In the present study, a total of 14 (73.68%) cases of abortions and two (100%) cases of still births were detected positive for Trypanosoma evansi infection by wet and dry blood smear examination and fetal tissue PCR in camels of an organized farm. The abortions in infected dams were recorded from 8 to 11.5 month of gestation, however majority occurred during 9th to 10th month. The important laboratorial findings in infected dams were anemia, hypoglycemia, hyperproteinemia and leukocytosis. At necropsy the T. evansi infected aborted and still born fetuses showed subcutaneous edema, presence of moderate amount of dark red hemolysed blood in thoracic and abdominal cavity, bronchopneumonia, hepatic necrosis and acute congestion in all vital organs. Microscopically, there was severe congestion, thickening of bronchial and alveolar wall and mononuclear infiltration in the fetal lung, necrotic and degenerative changes in the liver, nephritis along with severe congestion and tubular necrosis in the kidneys and necrotic and degenerative changes and congestion of capillaries in the brain. The T. evansi DNA was detected by PCR from blood, lung, spleen, liver, kidney and brain of all the infected aborted and still born fetuses. The results of the study indicated that T. evansi can cross placental barrier and cause pathological events in the fetus resulting into abortion or still birth in pregnant camels.

Trypanosoma evansi is an important pathogen that causes changes in nitric oxide (NO) levels and antioxidant enzymes, as well as oxidative stress. The present study evaluated the in vivo effect of T. evansi infection on frequency and index of DNA damage in liver, heart, spleen and total blood of rats. Twenty rats were assigned into two groups with ten rats each, being subdivided into four subgroups (A1 and A2, 5 animals/group; and B1 and B2, 5 animals/group). Rats in the subgroups A1 and A2 were used as control (uninfected) and animals in the subgroups B1 and B2 were inoculated with T. evansi (infected). NO in serum and the comet assay were used to measure DNA damage index (DI) and damage frequency (DF) in liver, heart, spleen and total blood of infected rats. Increased NO levels on days 3 and 9 post-infection (PI) was observed (P < 0.001). Also, it was verified an increase on DI and DF in the evaluated organs on days 3 and 9 PI (P < 0.001). Our data show that T. evansi infection causes genotoxicity due to the production of NO, causing not only the death of the protozoan, but also inducing DNA damage in the host., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The aim of this study was to investigate the activities of important enzymes involved in the energetic metabolism in the liver of rats experimentally infected by Trypanosoma evansi. Adenylate kinase (AK), pyruvate kinase (PK), and lactate dehydrogenase (LDH) in liver homogenate, as well as aspartate aminotransferase (AST), alanine aminotransferase (ALT), and clotting time in plasma were evaluated at 5 and 15 days post-infection (PI). The activities of the respiratory chain complexes and of Na(+), K(+)-ATPase were also evaluated. This study demonstrates energetic metabolism impairment in rats infected by T. evansi. A reduced energy metabolism in the liver of rats infected by T. evansi was observed, demonstrated by AK decreased and PK increased activities at 5 days PI, a mechanism known as energetic compensation. However, at 15 days PI a decrease of AK and PK activities were observed. In addition, an increase in the activities of respiratory chain complexes II, II-III and IV in infected rats at 15 days PI, and a decrease of Na(+), K(+)-ATPase activities in infected rats on days 5 and 15 PI were verified. In the plasma, we observed an increase in ALT and AST activities on days 5 and 15 PI, and increase in clotting time in infected rats. The changes caused by T. evansi infection on the activity of enzymes of hepatic energy metabolism can corroborate to elucidate the mechanisms that lead to liver injury and inflammatory infiltration verified in T. evansi infected rats. Therefore, these alterations are directly related to disease pathogenesis., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Blood samples were collected from 05 clinically healthy and 10 adult female water buffaloes naturally infected with Trypanosoma evansi. Confirmation of disease free and infected status of buffaloes was made on clinical signs, observation of T. evansi parasites in the blood smear and duplex PCR based assay. Blood samples were evaluated for levels of haemoglobin (Hb), packed cell volume (PCV), differential leucocytes count (DLC), lipid peroxidation (LPO), calcium, phosphorous, magnesium sodium and potassium and activities of superoxide dismutase (SOD), catalase (CAT), aspartate transaminase (AST), lactate dehydogenase (LDH) and alkaline phosphatase (ALP). The results of the study revealed substantial decrease in levels of Hb, PCV and increase in LPO, SOD, CAT and AST in infected animals compared to healthy animals. However other haematological and biochemical indices did not show significant variations in infected and healthy buffaloes. The enhanced erythrocytic oxidation and reduction of hematological indices, suggests that the enhanced oxidation of the erythrocytes may be a contributory factor in erythrocytic destruction and progression of the anaemia in T. evansi infection in water buffaloes., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Arginine-Serine (RS) domain-containing proteins are RNA binding proteins with multiple functions in RNA metabolism. In mammalian cells this group of proteins is also implicated in regulation and coordination of cell cycle and apoptosis. In trypanosomes, an early branching group within the eukaryotic lineage, this group of proteins is represented by 3 members, two of them are SR proteins and have been recently shown to be involved in rRNA processing as well as in pre-mRNA splicing and stability. Here we report our findings on the 3rd member, the SR-related protein TbRRM1. In the present study, we showed that TbRRM1 ablation by RNA-interference in T. brucei procyclic cells leads to cell-cycle block, abnormal cell elongation compatible with the nozzle phenotype and cell death by an apoptosis-like mechanism. Our results expand the role of the trypanosomal RS-domain containing proteins in key cellular processes such as cell cycle and apoptosis-like death, roles also carried out by the mammalian SR proteins, and thus suggesting a conserved function in this phylogenetically conserved protein family.

CpG oligodeoxynucleotides (CpG-ODN) stimulate immune cells from a wide spectrum of mammalian species. Class C CpG-ODN is relatively stable and has the combined immune effects of both A and B classes of CpG-ODN. Trypanosoma evansi produces the state of immuno-suppression in the infected hosts. The current chemotherapeutic agents against this parasite are limited in number and usually associated with severe side effects. The present work aimed to determine the immunostimulatory effects of CpG-ODN class C in T. evansi infected rabbits. Rabbits inoculated with CpG C and challenged with T. evansi resulted in delayed onset of clinical signs with reduced severity in comparison to that of T. evansi infected rabbits. The treatment also enhanced humoral immune responses. Histopathological findings in liver and spleen revealed enhancement of mononuclear cell infiltration and secondary B cell follicles. These results demonstrate that CpG-ODN class C, has immunostimulatory properties in rabbit model of trypanosomosis. The use of booster doses or sustained delivery of CpG-ODN will further elucidate the prolonged CpG-ODN generated immune responses.

This study aimed to evaluate in vitro and in vivo trypanocidal activity of free and nanoencapsulated curcumin against Trypanosoma evansi. In vitro efficacy of free curcumin (CURC) and curcumin-loaded in lipid-core nanocapsules (C-LNCs) was evaluated to verify their lethal effect on T. evansi. To perform the in vivo tests, T. evansi-infected animals were treated with CURC (10 and 100 mg kg(-1), intraperitoneally [i.p.]) and C-LNCs (10 mg kg(-1), i.p.) during 6 days, with the results showing that these treatments significantly attenuated the parasitaemia. Infected untreated rats showed protein peroxidation and an increase of nitrites/nitrates, whereas animals treated with curcumin showed a reduction on these variables. As a result, the activity of antioxidant enzymes (superoxide dismutase and catalase) differs between groups (P<0.05). Infected animals and treated with CURC exhibited a reduction in the levels of alanine aminotransferase and creatinine, when compared with the positive control group. The use of curcumin in vitro resulted in a better parasitaemia control, an antioxidant activity and a protective effect on liver and kidney functions of T. evansi-infected adult male Wistar rats.

Electron spin resonance (ESR), called also electron paramagnetic resonance (EPR) together with the spin trapping technique, has allowed us to study and understand how free radicals are involved in various pathologies. In this review, the importance of spin trapping technique in the study of diseases such as cancer, diabetes, hypertension and parasitic diseases is discussed. In addition, advances in the use of this technique as therapeutic agents and other interesting applications as the immuno-spin trapping technique are reviewed.

Surra, caused by Trypanosoma evansi, is a neglected disease due to frequent subclinical evolution, especially in bovines in Asia. However, acute and chronic signs are regularly observed, with significant sanitary and economic impacts. In this study, we evaluated and applied an indirect-ELISA test for the detection of anti-T. evansi immunoglobulin G in buffaloes using antibovine conjugate. Based on buffalo reference sera from the Philippines, a two-graph receiver operating characteristics analysis (TG-ROC) was conducted to define an optimal cut-off value; sensitivity and specificity were estimated at 92.5% and 94.2%, respectively. A cross-sectional serological survey was carried out in the major buffalo breeding areas of Thailand; 892 buffaloes from 8 provinces were sampled in North, Northeastern, and Southern Thailand. Seropositive buffaloes were found in all 8 provinces, on 20.3% of farms for an overall prevalence of 12.2% (95% CI 10.2-14.5%). Nearly one-third of the sampled population was exposed to infection. Broader sampling would be necessary but is not possible in the southern half-wild breeding systems. According to our results, buffaloes may constitute a large and robust reservoir for T. evansi, which is a permanent threat to other livestock such as cattle and horses as well as wild animals such as elephants in Southest Asia.

The aim of this study was to evaluate the effect of treatment with liposomal (L-DMZ) and conventional (C-DMZ) diminazene aceturate formulations on hepatic and renal functions of rats, experimentally infected with Trypanosoma evansi. For this purpose, 72 Wistar rats (Rattus norvegicus) were divided into six groups (A, B, C, D, E, and F). Each group was subdivided into two other subgroups in order to assess the biochemical and histological results on days 7 and 40 post-treatment (PT). Treatments were carried out based on two different therapeutic protocols: L-DMZ and C-DMZ at 3.5mg/kg(-1), single dose (groups C and D), and five successive doses within intervals of 24h (groups E and F). Groups A and B corresponded to uninfected and infected (without treatment) animals, respectively. Sample collections were held on days 7 and 40 PT for the assessment of hepatic [alkaline phosphatase (AP), alanine transferase (ALT), albumin, gamma glutamil transferase (GGT) and renal functions (creatinine and urea). Additionally, the histology of fragments of liver, kidney, and spleen was performed. Animals in group B showed a significant increase in AP, GGT, ALT, and urea when compared with group A. On day 7 post-inoculation (PI), the biochemical analysis showed a reduction (P<0.05) of AP and GGT, while the levels of urea were increased in groups C, D, E, F. On day 40 PT, ALT was increased in these same groups when compared with group A. In histopathology, changes in liver samples were observed on day 7 PT in groups D and F, especially regarding the area and density of the hepatocytes. Renal analysis exhibited changes in glomerular space, glomerular, and corpuscular areas in group E. Therefore, these results allowed us to conclude that the treatment with L-DMZ and C-DMZ led to variable biochemical changes, which defined the functions of the liver and kidneys of treated animals, since the main histopathology alterations were observed in animals treated with liposomes, at their higher dosages. Thus, treatments with L-DMZ and C-DMZ in five consecutive doses were effective although being followed by liver toxicity., (Copyright © 2014 Elsevier GmbH. All rights reserved.)

The detection of Trypanosoma evansi in blood is intricate, primarily in chronic stage of infection, as the parasitaemia is often low and fluctuating. The climatic conditions of the target area of Punjab (a province of India with a total of 34,000 horses and ponies used for sports and transport) are conducive for the parasite propagation. The objective of present investigation was to assess the prevalence of T. evansi in central and western Punjab by PCR and card agglutination test (CATT/T. evansi) in relation to clinico-haematobiochemical alterations and risk factors associated with latent trypanosomosis. A total of 169 equine blood and serum samples tested by CATT/T. evansi revealed 16 cases positive, with 6.8% from central plain and 13.63% from western zone. To assess the specificity of serological test, PCR1 was performed using established primer pair TR3 5'-GCG CGG ATT CTT TGC AGA CGA-3' and TR4 5'-TGC AGA CAC TGG AAT GTT ACT-3' for T. evansi. PCR2 applied with primer pair RoTat1.2F: 5'-ATG TCA ACG ATG CCT GTT ACA TTA CGC AC-3' and RoTat1.2R: 5'-TAA ATA TCA CTG TCA AGA CCT GCT GCG G-3' to rule out the consensus between the finding of the two PCR assays and agglutination test for T. evansi, which displayed results in concordance with PCR1. PCR assays showed 1.92 and 1.51% positive samples from central plain and western zone, respectively. With respect to PCR assay, CATT/T. evansi showed 100% sensitivity and 92.1% specificity. Microscopy showed a very low prevalence rate of 0.59% with only one sample positive with teaming parasitaemia. Comparison between sexes revealed higher positivity in mares by the three tests (BSE: 0.95%, PCR: 2.88%, CATT/T. evansi: 14.42%). The haemato-biochemical factors were found to be altered in PCR positive cases, while the mean value of vital parameters lied in normal range in seropositive cases. The female horse (RR=0.0937, 95% CI=1.388-190.223%) population was found to be at the highest risk of seropositivity for T. evansi, particularly in the unorganized farms (RR=19.726, 95% CI=2.918-400.221%)., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The aim of this study was to evaluate the effect of zinc supplementation on the ecto-adenosine deaminase activity (E-ADA), zinc seric levels and cytokines (TNF-α, IL-1, IL-6, and IL -10) on rats experimentally infected by Trypanosoma evansi. Four groups with 10 rats each were used as negative controls (groups A and B), while the animals from the groups C and D were infected intraperitoneally with 0.1 mL of cryopreserved blood containing 1.4 × 10(4) of trypanosomes. Animals of groups B and D received two doses of Zinc (Zn) at 5 mg kg(-1), subcutaneously, on the 2nd and 7th day post-infection (PI). Blood samples were collected on days 5 (n = 5) and 15 PI (n = 5). Zn supplementation was able to increase the rat's longevity and to reduce their parasitemia. It was observed that seric Zn levels were increased on infected animals under Zn supplementation. Animals that were infected and supplemented with Zn showed changes in E-ADA activity and in cytokine levels (P < 0.05). Zn supplementation of healthy animals (Group B), increased the E-ADA activity, as well as reduced the concentration of cytokines. Infected animals from groups C and D showed increased levels of cytokines. Finally, we observed that Zn supplementation led to a modulation on cytokine's level in rats infected by T. evansi, as well as in E-ADA activity., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Trypanosomatid parasites infect over 21 million people worldwide, with a range of disease phenotypes. Trypanosoma cruzi causes American trypanosomiasis, wherein 30-40% of infected individuals develop disease manifestations, most commonly cardiomyopathy but also digestive megasyndromes. In the case of Trypanosoma brucei, the etiological agent of African trypanosomiasis, disease progression can be rapid or slow, with early or late central nervous system involvement. Finally, Leishmania species cause leishmaniasis, a disease that ranges from self-healing but scarring cutaneous lesions to fatal visceral leishmaniasis in which parasites disseminate to the liver, spleen, and bone marrow. This review highlights parasite factors involved in disease phenotype in all three trypanosomatid diseases, with a particular focus on recent advances using large-scale 'omics' techniques., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Background: The development of techniques that allow the imaging of animals infected with parasites expressing luciferase opens up new possibilities for following the fate of parasites in infected mammals., Methods: D-luciferin potassium salt stock solution was prepared in phosphate-buffered saline (PBS) at 15 mg/ml. To produce bioluminescence, infected and control mice received an intraperitoneal injection of luciferin stock solution (150 mg/kg). All mice were immediately anesthetized with 2% isofluorane, and after 10 minutes were imaged. Ex vivo evaluation of infected tissues and organs was evaluated in a 24-well plate in 150 μg/ml D-luciferin diluted in PBS. Images were captured using the IVIS Lumina image system (Xenogen). Dissected organs were also evaluated by microscopy of hematoxylin-eosin stained sections., Results: Here we describe the results obtained using a genetically modified Dm28c strain of T. cruzi expressing the firefly luciferase to keep track of infection by bioluminescence imaging. Progression of infection was observed in vivo in BALB/c mice at various intervals after infection with transgenic Dm28c-luc. The bioluminescent signal was immediately observed at the site of T. cruzi inoculation, and one day post infection (dpi) it was disseminated in the peritoneal cavity. A similar pattern in the cavity was observed on 7 dpi, but the bioluminescence was more intense in the terminal region of the large intestine, rectum, and gonads. On 14 and 21 dpi, bioluminescent parasites were also observed in the heart, snout, paws, hind limbs, and forelimbs. From 28 dpi to 180 dpi in chronically infected mice, bioluminescence declined in regions of the body but was concentrated in the gonad region. Ex vivo evaluation of dissected organs and tissues by bioluminescent imaging confirmed the in vivo bioluminescent foci. Histopathological analysis of dissected organs demonstrated parasite nests at the rectum and snout, in muscle fibers of mice infected with Dm28c-WT and with Dm28c-luc, corroborating the bioluminescent imaging., Conclusion: Bioluminescence imaging is accurate for tracking parasites in vivo, and this methodology is important to gain a better understanding of the infection, tissue inflammation, and parasite biology regarding host cell interaction, proliferation, and parasite clearance to subpatent levels.

Trypanosoma evansi infection typically produces wasting disease, but it can also develop into a neurological or meningoencephalitis form in equids. Trypanosomiasis in horses was treated with quinapyramine sulfate, and all the 14 infected animals were recovered clinically. After clinical recovery, four animals developed a neurological form of the disease at various intervals. Two of these animals treated with diminazene aceturate recovered temporarily. Repeated attempts failed to find the parasite in the blood or the cerebrospinal fluid (CSF), but all of the animals were positive in enzyme-linked immunosorbent assay. The calculation of the antibody index (AI) in the serum and the CSF and polymerase chain reaction (PCR) analysis of the CSF and brain tissue were carried out to confirm the neuro-infection. We found PCR and AI analyses of the CSF to be useful tools in the diagnosis of the neurological form of trypanosomiasis when the organism cannot be found in the blood or CSF. The increased albumin quotient is indicative of barrier leakage due to neuroinflammation. The biochemical changes in the CSF due to nervous system trypanosomiasis include increases in the albumin quotient, total protein, and urea nitrogen. It seems to be the first report on relapse of the nervous form of trypanosomiasis in equids even after quinapyramine treatment in endemic areas.

African trypanosomosis is a potentially fatal disease that is caused by extracellular parasitic protists known as African trypanosomes. These parasites inhabit the blood stream of their mammalian hosts and produce a number of pathological features, amongst which is anemia. Etiology of the anemia has been partly attributed to an autoimmunity-like mediated erythrophagocytosis of de-sialylated red blood cells (dsRBCs) by macrophages. Lactose infusion to infected animals has proven effective at delaying progression of the anemia. However, the mechanism of this anemia prevention is yet to be well characterized. Here, the hypothesis of a likely induced further modification of the dsRBCs was investigated. RBC membrane galactose (RBC m-GAL) and packed cell volume (PCV) were measured during the course of experimental trypanosomosis in mice infected with Trypanosoma congolense (stb 212). Intriguingly, while the membrane galactose on the RBCs of infected and lactose-treated mice (group D) decreased as a function of parasitemia, that of the lactose-untreated infected group (group C) remained relatively constant, as was recorded for the uninfected lactose-treated control (group B) animals. At the peak of infection, the respective cumulative percent decrease in PCV and membrane galactose were 30 and 185 for group D, and 84 and 13 for group C. From this observed inverse relationship between RBCs membrane galactose and PCV, it is logical to rationalize that the delay of anemia progression during trypanosomosis produced by lactose might have resulted from an induction of galactose depletion from dsRBCs, thereby preventing their recognition by the macrophages., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Trypanosoma evansi (T. evansi) is an endemic disease of camels and other domestic animals in Egypt. This study aimed to determine the prevalence of clinical and sub-clinical T. evansi infection among camels in Ismailia, Egypt, as well as survey their owners for T. evansi infection. The diagnostic sensitivity of three different PCR assays for detection of T. evansi in blood samples was evaluated. Blood samples were collected from 100 camels and 20 of their owners in the Ismailia governorate. Results revealed that the percentage of infected of camels with T. evansi vary with the detection method, ranging from 10% to 46% by PCR compared to 12% by microscopic examination of stained blood smears. Targeting the highly repeated sequence of mini-chromosome satellite DNA (TBR1/2 primer set) was more often seen in the PCR method (46% positive) compared to targeting ITS 1 (16% positive) or RoTat 1.2 VSG (10% positive) sequences. A partial sequence of RoTat 1.2 VSG gene was identical to the T. evansi sequences reported from India and Kenya, but varied similarity was seen when aligned with Egyptian T. evansi sequences. None of the camel owners were positive for T. evansi by microscopic examination of stained blood smears or PCR assays. PCR assay based on TBR sets is useful in the diagnosis and control disease and reducing economic losses., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The two classical forms of human trypanosomoses are sleeping sickness due to Trypanosoma brucei gambiense or T. brucei rhodesiense, and Chagas disease due to T. cruzi. However, a number of atypical human infections caused by other T. species (or sub-species) have been reported, namely due to T. brucei brucei, T. vivax, T. congolense, T. evansi, T. lewisi, and T. lewisi-like. These cases are reviewed here. Some infections were transient in nature, while others required treatments that were successful in most cases, although two cases were fatal. A recent case of infection due to T. evansi was related to a lack of apolipoprotein L-I, but T. lewisi infections were not related to immunosuppression or specific human genetic profiles. Out of 19 patients, eight were confirmed between 1974 and 2010, thanks to improved molecular techniques. However, the number of cases of atypical human trypanosomoses might be underestimated. Thus, improvement, evaluation of new diagnostic tests, and field investigations are required for detection and confirmation of these atypical cases.

HUMAN AFRICAN TRYPANOSOMIASIS (HAT) MANIFESTS IN TWO STAGES OF DISEASE: firstly, haemolymphatic, and secondly, an encephalitic phase involving the central nervous system (CNS). New drugs to treat the second-stage disease are urgently needed, yet testing of novel drug candidates is a slow process because the established animal model relies on detecting parasitemia in the blood as late as 180 days after treatment. To expedite compound screening, we have modified the GVR35 strain of Trypanosoma brucei brucei to express luciferase, and have monitored parasite distribution in infected mice following treatment with trypanocidal compounds using serial, non-invasive, bioluminescence imaging. Parasites were detected in the brains of infected mice following treatment with diminazene, a drug which cures stage 1 but not stage 2 disease. Intravital multi-photon microscopy revealed that trypanosomes enter the brain meninges as early as day 5 post-infection but can be killed by diminazene, whereas those that cross the blood-brain barrier and enter the parenchyma by day 21 survived treatment and later caused bloodstream recrudescence. In contrast, all bioluminescent parasites were permanently eliminated by treatment with melarsoprol and DB829, compounds known to cure stage 2 disease. We show that this use of imaging reduces by two thirds the time taken to assess drug efficacy and provides a dual-modal imaging platform for monitoring trypanosome infection in different areas of the brain.

The aim of this study was to evaluate the changes in hematological and biochemical parameters of blood during acute Trypanosoma evansi infection in Wistar rats. The end points studied were hematologic parameters, red blood cell fragility, iron content, and glutathione and lipid peroxidation levels. Forty-eight animals were infected with trypomastigotes and distributed into five groups according to the level of parasitemia. Twelve non-inoculated animals were used as control. Parasitemia increased progressively, reaching highest scores at 15 days post-inoculation. At this point, several deleterious effects were observed such as an increase in iron content, in osmotic fragility, and in lipid peroxidation index, while glutathione decreased drastically. These changes were highly correlated to parasitemia (p < 0.0001) and among each other (p ≤ 0.001). Hematological indices (Hb, packed cell volume (PCV), red blood cells (RBC), and mean corpuscular hemoglobin concentration) were also correlated to parasitemia (p ≤ 0.0003) but failed to correlate to the other variables. Along with increase in iron, RBC fragility produced a decrease in RBC, PCV, and Hb, but not in mean corpuscular volume. Decrease in glutathione was negatively correlated to the end products of lipid peroxidation, clearly indicating the establishment of a pro-oxidant condition. The results show that the infection causes hematological impairments, increases iron and osmotic fragility, along with marked oxidative stress in red blood cells of rats inoculated with T. evansi.

In recent years, the emergence of highly pathogenic Trypanosoma evansi strains in the Philippines has resulted in substantial losses in livestock production. In this study, we isolated T. evansi from infected-water buffaloes in the Philippines and analyzed their virulence using mice and cattle. A total of 10 strains of T. evansi were isolated. Evaluation of the virulence of each strain using mice depicted significant differences among the strains in the prepatent period, the level of parasitemia, and the survival time of the infected animals. In mice infected with the highly pathogenic T. evansi, signs of excessive inflammation such as marked splenomegaly and increase more than 6-fold in the number of leukocytes were observed at 8 days post-infection. To study the virulence of the parasite strains in cattle (which are the common T. evansi hosts in Philippines), cattle were infected with the T. evansi isolates that showed high and low virulence in mice. The rate of parasite growth and the length of the prepatent periods were found to be similar to those observed in mice for the respective strains. The cattle infected with the highly pathogenic strain developed anemia and a marked decrease in leukocyte counts. To determine the cause of the pathological changes, we analyzed the expression levels of inflammatory cytokines and observed up-regulation of tumor necrosis factor-α in anemic infected cattle. Our findings suggest that the epidemic of T. evansi in the Philippines is characterized by T. evansi strains with varying virulences from low to very high pathogenicity in cattle.

Trypanosoma (subgenus Megatrypanum) theileri was first identified over one hundred years ago, and is a widespread parasite in cattle. Its life cycle within the mammalian host has rarely been reported. Whether there is an intracellular stage in tissues is unknown and such a stage has not been demonstrated experimentally. Intriguingly, using Giemsa staining with light microscopy and transmission electron microscopy examination, we found that the parasite was able not only to attach to cells but also to invade several phagocytic and non-phagocytic mammalian cells. Based on these findings, we conducted further investigations using a special antibody in immunofluorescence confocal images. Moreover, we examined a series of possible events of cell invasion in T. theileri. The results revealed that GM1, a marker of membrane rafts, was implicated in the mechanism of entry by this parasite. After incubation with tissue culture trypomastigotes, the gelatinolytic activity was significantly increased and accumulated at the attachment sites. Using ultrastructural localization detection by CytoTracker live imaging and confocal immunofluorescence microscopy, we found that lysosome fusion and the autophagy pathway were engaged in invaginating processes. T. theileri amastigotes also invaded cells and were enclosed by the lysosomes. Furthermore, tissue-cultured trypomastigotes were found to be capable of triggering intracellular free Ca(2+) transients and TGF-β-signaling. Our findings that intracellular amastigote stages exist in mammalian cells infected with T. theileri and that the invasion processes involved various host cell components and cell signalings were extremely surprising and warrant further investigation., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Infection by Trypanosoma vivax and other African trypanosomes plays an important role in reproductive disorders in male and female livestock. Outbreaks of T. vivax in the semi-arid region of northeastern Brazil are characterized by wasting disease in cattle, sheep and goats with hematological, cardiac and nervous compromises in addition to reproductive failures. Similar to reports from Africa, we previously observed a reduction in fertility rates and severe testicular degeneration and epididymitis in male sheep infected with T. vivax from this region. Although anestrus is frequently reported in goats and sheep infected with T. vivax, the effects of this infection on the female reproductive organs need clarification. In this study, we addressed this issue through a histopathological evaluation of ovarian follicular morphology and classification in goats experimentally infected with a T. vivax isolate from the Brazilian semi-arid region. The infected animals presented typical clinical signs of trypanosomosis by T. vivax, including anemia, hyperthermia, pallor of the mucous membranes, enlarged lymph nodes, and progressive loss of weight. All the infected goats remained anestrus throughout the experimental period and exhibited important disturbances in the ovaries, evidenced by reduced size and a smooth surface without follicles or corpora lutea, and abnormal follicular development. In addition, through PCR, we detected T. vivax DNA in the ovarian tissues of the infected goats. Our findings contributed to understand the female reproductive failure associated with trypanosomosis caused by T. vivax., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Trypanosoma evansi, the agent of "surra," is a salivarian trypanosome, originating from Africa. It is thought to derive from Trypanosoma brucei by deletion of the maxicircle kinetoplastic DNA (genetic material required for cyclical development in tsetse flies). It is mostly mechanically transmitted by tabanids and stomoxes, initially to camels, in sub-Saharan area. The disease spread from North Africa towards the Middle East, Turkey, India, up to 53° North in Russia, across all South-East Asia, down to Indonesia and the Philippines, and it was also introduced by the conquistadores into Latin America. It can affect a very large range of domestic and wild hosts including camelids, equines, cattle, buffaloes, sheep, goats, pigs, dogs and other carnivores, deer, gazelles, and elephants. It found a new large range of wild and domestic hosts in Latin America, including reservoirs (capybaras) and biological vectors (vampire bats). Surra is a major disease in camels, equines, and dogs, in which it can often be fatal in the absence of treatment, and exhibits nonspecific clinical signs (anaemia, loss of weight, abortion, and death), which are variable from one host and one place to another; however, its immunosuppressive effects interfering with intercurrent diseases or vaccination campaigns might be its most significant and questionable aspect.

The virulence of three Trypanosoma evansi isolates in Luzon, Visayas and Mindanao water buffaloes was compared determining the mortality rate, parasitemia level, clinical signs, and lesions on mice. A total of 51 inbred Balb/c mice (5-6 weeks old) were used and divided into two sets. Set A had three groups corresponding to three trypanosomes isolates (Luzon, Visayas, and Mindanao) with seven mice each whose parasitemia level, clinical signs, and lesions were noted at necropsy. Set B had three groups corresponding to the three isolates with ten mice each whose mortality was monitored. Each infected mouse was inoculated with 0.2 ml of T. evansi intraperitoneally and blood was examined under high power magnification. Their parasitemia level was determined using "Rapid Matching Method". Dead mice were subjected to necropsy and the lungs, liver, spleen, brain and heart were subjected to histopathological processing. Results showed that the mortality rate was highest at Day 3 for the Visayas isolates (70%), while at Day 5 for Luzon (90%) and Mindanao (70%) isolates. The parasitemia level of Visayas isolates (1×10(8.7)) reached the earliest peak at Day 4 while Luzon isolates (1×10(9)) at Day 6 and Mindanao isolates (1×10(8.7)) at Day 8. Statistical analysis using Least significant difference (LSD) revealed significant difference among treatment means at Days 2 and 4. All of the affected mice showed rough hair coat, decreased body weight, and decreased packed cell volume. The most obvious gross lesions observed were pale liver with petechiations and pale muscles. Histopathological examination revealed depletion of the red pulp and extramedullary hematopoiesis in the spleen. Congestion, intralesional trypanosomes in blood vessel and extramedullary hematopoiesis were observed in the liver. In the lungs non-specific lesions observed were pulmonary edema, congestion and hemosiderosis., (Copyright © 2011 Elsevier Inc. All rights reserved.)

The aim of this study was to investigate whether rats infected with Trypanosoma evansi had neurological and locomotor signs, as well as histological lesions in the central nervous system (CNS) and pelvic muscles. To carry out this study, 52 rats were used and divided into two groups. The animals in Group A (n=40) were infected with T. evansi, and the rats in Group B (n=12) were used as negative controls (non-infected). Neurological examination was performed at Days 5, 15, 30 and 150 post-infection (PI) with eventual euthanasia of the rats. Samples of brain, spinal cord and skeletal muscle (biceps femoris and gastrocnemius muscles) were collected. The neurological tests evaluated motor capacity, balance and pain sensitivity. At Day 5 PI in Subgroup A1, the rats showed high parasitemia, became apathetic and presented with slow movements and signs of disorientation. After Day 15 PI in Subgroup A2 and Day 30 PI in Subgroup A3, no more clinical abnormalities were observed. Histologically, there was no damage to the CNS in these three subgroups, but within Subgroup A3, mononuclear infiltration of the muscle was observed. Rats chronically infected (Subgroup A4 - Day 150 PI) showed muscle atrophy, walking dysfunction and paralysis of the hind limbs. Mild mononuclear inflammatory infiltrates and perivascular cuffs were observed in the CNS of some of the animals in Subgroup A4. In these rats, severe muscle damage was observed in the skeletal muscle which included atrophy and loss of muscle fibers, multinucleated giant muscle cells, mononuclear myositis, Wallerian degeneration of the innervating fibers and mononuclear inflammatory infiltrate in the perineurium and adipose tissue. Based upon these findings, we conclude that infection by T. evansi in rats leads to muscle damage, which is probably the cause of the paralysis of hind limbs., (Copyright © 2011 Elsevier GmbH. All rights reserved.)

Trypanosoma evansi infections in domestic animals are characterized by anemia and thrombocytopenia. The cause of the platelets decrease is unknown, but researchers suggest that thrombocytopenia may result from damage of the bone marrow, reduced survival of platelets, auto-immune thrombocytopenia, disseminated intravascular coagulation and splenic sequestration. Some of these causes have already been tested by our research group and found to be unrelated. Therefore, this study has the objective of testing the hypothesis that splenic sequestration might be responsible for thrombocytopenia in T. evansi-infected rats. A total of 28 rats assigned to four groups were used in the experiment. Group A rats were splenectomized and infected with T. evansi, group B rats were infected with T. evansi, group C rats were splenectomized, but not infected and group D rats were normal controls. Five days post-infection all rats were anesthetized and blood was collected in order to measure the number of circulating platelets, fibrinogen levels, prothrombin time (PT) and activated partial thromboplastin time (aPTT). The spleens of groups B and D were weighed at necropsy. The infected animals (groups A and B) showed a significant reduction in platelets and increased PT and aPTT when compared to negative control groups (groups C and D). Animals from group A showed increased levels of fibrinogen. The mean weight of spleen differed between group B (2.62g) and group D (0.55g). It was concluded that there is no relationship between thrombocytopenia and splenic sequestration in infection by T. evansi., (Copyright © 2010 Elsevier Ltd. All rights reserved.)