Clinical and Immunopathological Aspects of Heart Damage in Dogs Infected with Trypanosoma brucei

In an attempt to increase the understanding of the mechanisms of cardiac damage in African trypanosomiasis, a group of beagle dogs were infected with Trypanosoma brucei. Cardiac involvement during the course of the disease was monitored using various non-invasive techniques, including auscultation, palpation, echocardiography and electrocardiography (ECG). Parasitological, haematological, endocrinological and biochemical studies were carried out. Two dogs per time point were euthanised at mid-infection on days 10 and 15, and in the terminal stages on days 21, 22 and 26, and tissue samples taken from the heart for histological, histochemical, transmission electron microscopical and immunofluorescence investigations. The dogs developed an acute disease syndrome, characterised by high parasitaemia, persistent fever, increased plasma levels of acute phase proteins, severe anaemia, lymph node and splenic enlargement, wasting and weight loss. If the dogs were not treated or the experiments terminated by humane euthanasia, death would have invariably occurred during week 4 of infection. Clinical evidence of cardiac damage was demonstrated from the end of week 1 by echocardiography and ECG. The clinical signs included tachycardia, incompetence of all the cardiac valves, and first degree heart block. With progress of the disease, second degree and in some cases complete heart block developed. Towards the end of week 3 and beginning of week 4, cardiac performance deteriorated, as demonstrated by a reduction in left ventricular function, bradycardia, and accumulation of pericardial effusion. Histological and ultrastructural studies confirmed a severe pancarditis involving the myocardium, the cardiac valves, and the vasculature. Deposition of fibrinogen and sparse quantities of IgG, IgM and C3 was demonstrated in perivascular and interstitial spaces. A terminal fall in plasma levels of the cardiac hormone atrial natriuretic factor (ANF) occurred, and was associated with decreased numbers, sizes and electron density of the atrial storage granules. The low plasma levels of ANF were accompanied by an inverse increase in plasma renin activity, an indication that the dogs at that time were incapable of controlling blood volume. From the end of week 2 of infection, a gradually developing hyperlipidaemia and intense deposition of lipids in the myocardium and infiltrating macrophages was demonstrated for the first time. It is possible that cachectin/tumour necrosis factor (TNF) was contributing to the hyperlipidaemic state, following the detection of increased cachectin/TNF activity in monocytes from infected dogs. The presence of large numbers of trypanosomes, infiltrating cells that consisted mainly of macrophages, neutrophils, plasma cells and a few lymphocytes, in areas of extensive myocardial damage indicated that tissue damage was initiated by excessive immunological responses by the host to this highly antigenic parasite. Tissue damage was probably exacerbated by accumulation of toxic and biologically active substances released from dead trypanosomes and autolysing inflammatory cells, following obstruction of the lymphatic vessels draining the heart. Anaemia, vascular damage and interstitial oedema reduced tissue perfusion and increased the incidence of myocardial ischaemia. The presence of lipid deposits in ischaemic myocardium indicated that they too could be playing a major role in the pathogenesis of tissue damage. Further, the presence of mesangial deposits of immune complexes in the kidneys confirmed their presence in the circulation, indicating that immune complexes played a role in the pathogenesis of general tissue damage. Intravenous treatment of terminally ill dogs with the trypanocidal drug suramin at 10 to 20 mg/kg was usually unsuccessful. In most cases, a post-treatment reaction, with increased severity of heart damage, was observed. Nevertheless, the survival period of the dogs was prolonged, precipitating a condition of chronic cardiac failure. The dogs that survived longest were those that were best able to control anaemia after treatment. When a group of infected dogs were treated with the non-steroidal anti-inflammatory drugs (NSAIDs) cyclosporin A and azathioprine, or a combination of azathioprine and prednisolone, the severity of cardiac damage was reduced, despite intense trypanosome infiltration in the myocardium. These results confirmed the immunological and inflammatory nature of the cardiac disease induced by T. brucei. It would appear that if such drugs were used as an adjunct to trypanocidal drug treatment, the severity of post-treatment myocardial damage, and possibly general tissue damage, might have been reduced. The present work has conclusively demonstrated that cardiac damage in canine trypanosomiasis caused by T. brucei is mainly immunologically mediated. The similarity of the disease to human African trypanosomiasis indicated that cardiac damage in humans could be mediated through similar mechanisms. The future of the dog as large monogastric animal model for studying cardiac damage in African trypanosomiasis is promising

The burden of congenital Chagas disease and implementation of molecular diagnostic tools in Latin America

It is estimated that between 8000 and 15 000 Trypanosoma cruzi infected babies are born every year to infected mothers in Chagas disease endemic countries. Currently, poor access to and performance of the current diagnostic algorithm, based on microscopy at birth and serology at 8-12 months after delivery, is one of the barriers to congenital Chagas disease (CCD) control. Detection of parasite DNA using molecular diagnostic tools could be an alternative or complement to current diagnostic methods, but its implementation in endemic regions remains limited. Prompt diagnosis and treatment of CCD cases would have a positive clinical and epidemiological impact. In this paper, we analysed the burden of CCD in Latin America, and the potential use of molecular tests to improve access to early diagnosis and treatment of T. cruzi infected newborns.Fil: Picado, Albert. Foundation for Innovative New Diagnostics; SuizaFil: Cruz, Israel. Foundation for Innovative New Diagnostics; SuizaFil: Redard Jacot, Maël. Foundation for Innovative New Diagnostics; SuizaFil: Schijman, Alejandro Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; ArgentinaFil: Torrico, Faustino. Universidad Mayor de San Simón; Bolivia. Fundación CEADES; BoliviaFil: Sosa-Estani, Sergio Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Centro de Investigaciones en Epidemiología y Salud Pública. Instituto de Efectividad Clínica y Sanitaria. Centro de Investigaciones en Epidemiología y Salud Pública; Argentina. Drugs for Neglected Diseases initiative; BrasilFil: Katz, Zachary. Foundation for Innovative New Diagnostics; SuizaFil: Ndung'u, Joseph Mathu. Foundation for Innovative New Diagnostics; Suiz

Trypanosome diversity in wildlife species from the Serengeti and Luangwa Valley ecosystems

<p>Background: The importance of wildlife as reservoirs of African trypanosomes pathogenic to man and livestock is well recognised. While new species of trypanosomes and their variants have been identified in tsetse populations, our knowledge of trypanosome species that are circulating in wildlife populations and their genetic diversity is limited.</p> <p>Methodology/Principal Findings: Molecular phylogenetic methods were used to examine the genetic diversity and species composition of trypanosomes circulating in wildlife from two ecosystems that exhibit high host species diversity: the Serengeti in Tanzania and the Luangwa Valley in Zambia. Phylogenetic relationships were assessed by alignment of partial 18S, 5.8S and 28S trypanosomal nuclear ribosomal DNA array sequences within the Trypanosomatidae and using ITS1, 5.8S and ITS2 for more detailed analysis of the T. vivax clade. In addition to Trypanosoma brucei, T. congolense, T. simiae, T. simiae (Tsavo), T. godfreyi and T. theileri, three variants of T. vivax were identified from three different wildlife species within one ecosystem, including sequences from trypanosomes from a giraffe and a waterbuck that differed from all published sequences and from each other, and did not amplify with conventional primers for T. vivax.</p> <p>Conclusions/Significance: Wildlife carries a wide range of trypanosome species. The failure of the diverse T. vivax in this study to amplify with conventional primers suggests that T. vivax may have been under-diagnosed in Tanzania. Since conventional species-specific primers may not amplify all trypanosomes of interest, the use of ITS PCR primers followed by sequencing is a valuable approach to investigate diversity of trypanosome infections in wildlife; amplification of sequences outside the T. brucei clade raises concerns regarding ITS primer specificity for wildlife samples if sequence confirmation is not also undertaken.</p&gt

Pathogenicity of bloodstream and cerebrospinal fluid forms of Trypanosoma brucei rhodesiense in Swiss White Mice

Trypanosoma brucei rhodesiense (T.b.r.), the causative agent of the East African form of human African trypanosomiasis (HAT), is capable of crossing the blood brain barrier and invade the central nervous system (CNS). However, it is not clear whether bloodstream forms (BSF) of T.b.rhodesiense differ in biological characteristics from the cerebrospinal fluid (CSF) forms. The present study was carried out to compare the pathogenicity of CSF and BSF of T.b. rhodesiense parasites in Swiss white mice following intraperitoneal inoculation with 106 trypanosomes. The parasites were tested for presence of the serum resistance associated (SRA) gene. Parasitaemia, body weight, packed cell volume (PCV) and survival of the mice was monitored daily until the experiment was terminated. Data was analyzed using general linear model. Both forms of parasite were positive for the SRA gene, and there was no significant difference in progression of parasitaemia, PCV values or survival of the mice. However, the weights of BSF infected mice initially dropped faster than those of CSF infected mice (P<0.001)

Economics of Insecticide use and Potential for Bt Maize Varieties in the Control of Stalkborer in Kenya.

Maize is the staple food crop and source of income for majority of the Kenyan population and many sub-Saharan African countries. The increasing Kenyan population demands an increase in maize production if intermittent food deficits have to be averted. Since the introduction of improved maize varieties in mid-1960, the start of Green Revolution period, maize yields increased drastically up to 1970s and started declining from 1980s to-date. The key contributory factors are nutrient mining, sub-optimal input use and insect pest damage. Of the insect pests, stalk borer is of economic importance. Currently, KARI and CIMMYT are developing maize varieties that are tolerant to stalk borer damage. In order to evaluate the potential impact of these interventions economics of stalk borer control at farm level was evaluated. Surveys complemented with on-farm trials were executed in six major maize growing zones of Kenya. Farmers were randomly selected and a sample-frame established after which a total of 1854 households were randomly selected using random sampling technique. Each household was interviewed using structured questionnaire. Data on method of stalk borer control and the type insecticides used was collected. Partial budget and economic surplus models were used. The results indicated that very few farmers control stalk borer in maize despite significant stalk borer losses of about 15%. Therefore if Bt maize is introduced in Kenya it is likely to reduce these losses. This will benefit many hungry and poor Kenyans with improved household food supply and on farm incomes, in line with Government policy of food security and poverty eradication.Crop Production/Industries,

Access to prompt diagnosis: The missing link in preventing mental health disorders associated with neglected tropical diseases

Globally, there are an estimated 1 billion people suffering from at least one of the 20 neglected tropical diseases (NTDs) prioritized by the World Health Organization (WHO). Prevalent in tropical and subtropical regions, this group of NTDs comprises diverse diseases, including vector-borne parasitic diseases (such as human African trypanosomiasis [HAT], Chagas disease, and leishmaniasis), skin diseases caused by environmental bacteria (such as Buruli ulcer [BU]), foodborne parasitic diseases (such as taeniasis/cysticercosis) or snake bite envenoming, which—together with scabies and other ectoparasites, mycetoma, and deep mycoses—were recently added to the list [1]. Despite their differences, NTDs are synonymous with poverty, life-long disability, stigma, and discrimination, not to mention the lack of effective control tools such as vaccines, diagnostics, and drugs.S

Isolation and propagation of Trypanosoma brucei gambiense from sleeping sickness patients in south Sudan

This study aimed at isolating Trypanosoma brucei gambiense from human African trypanosomiasis (HAT) patients from south Sudan. Fifty HAT patients identified during active screening surveys were recruited, most of whom (49/50) were in second-stage disease. Blood and cerebrospinal fluid samples collected from the patients were cryopreserved using Triladyl® as the cryomedium. The samples were stored at −150 °C in liquid nitrogen vapour in a dry shipper. Eighteen patient stabilates could be propagated in immunosuppressed Mastomys natalensis and/or SCID mice. Parasitaemia was highest in SCID mice. Further subpassages in M. natalensis increased the virulence of the trypanosomes and all 18 isolates recovered from M. natalensis or SCID mice became infective to other immunosuppressed mouse breeds. A comparison of immunosuppressed M. natalensis and Swiss White, C57/BL and BALB/c mice demonstrated that all rodent breeds were susceptible after the second subpassage and developed a parasitaemia >106/ml by Day 5 post infection. The highest parasitaemias were achieved in C57/BL and BALB/c mice. These results indicate that propagation of T. b. gambiense isolates after initial isolation in immunosuppressed M. natalensis or SCID mice can be done in a range of immunosuppressed rodent

Evaluation of different DNA extraction methods and loop-mediated isothermal amplification primers for the detection of Mycobacterium ulcerans in clinical specimens

Background: Early diagnosis and treatment of Buruli ulcer is critical in order to avoid the debilitating effects of the disease. In this regard, the development of new diagnostic and point of care tools is encouraged. The loop-mediated isothermal amplification for the detection of Mycobacterium ulcerans represents one of the new tools with a good potential of being developed into a point of care test. There is however the need to standardize the assays, reduce sample preparation times, improve the detection/visualization system and optimize them for high-throughput screening, adaptable to low resourced laboratories. Methods: In this study, we assessed two DNA extraction protocols (modified Boom and EasyNAT methods), three previously published LAMP primer sets (BURULI, MU 2404 and BU-LAMP), and compared the sensitivity and specificity of LAMP assays on three DNA amplification platforms. Results: Our results show that Buruli ulcer diagnosis using primers targeting IS2404 for the LAMP method is sensitive (73.75-91.49%), depending on the DNA extraction method used. Even though the modified Boom DNA extraction method provided the best results, its instrumentation requirement prevent it from being field applicable. The EasyNAT method on the other hand is simpler and may represent the best method for DNA extraction in less resourced settings. Conclusions: For further work on the development and use of LAMP tests for Buruli diagnosis, it is recommended that the BURULI sets of primers be used, as these yielded the best results in terms of sensitivity (87.50-91.49%) and specificity (89.23-100%), depending on the DNA extraction methods used.This study was supported by the Swiss Agency for Development and Cooperation and the UBS Optimus Foundation through FIND, Geneva-Switzerland. The funders had no role in the design of the study and collection, analysis, and interpretation of data and in writing the manuscript.S

Haematology of Experimental Trypanosoma Brucei Rhodesiense Infection in Vervet Monkeys

Haematological aberrations associated with human infective trypanosomes were investigated in the vervet monkey model of the Rhodesian sleeping sickness. Four monkeys were infected intravenously with 104 Trypanosoma brucei rhodesiense and monitored for changes in the blood profile using a haematological analyser. A chronic infection lasting between 48 and 112 days was observed. Microcytic hypochromic anaemia, which was characterized by a decline in packed cell volume (PCV), red blood cell (RBC) numbers, mean corpuscular volume (MCV) and mean corpuscular haemoglobin concentration (MCH) developed at an early stage, and persisted throughout the infection. The mean platelet counts declined significantly from 3 x 105/\u3bcl (day 0 post infection) to 6.8 x 104/\u3bcl (day 7 post infection) and remained low in all the animals. However, the mean platelets volume rose during the course of the infection. An initial decline in total white blood cell (WBC) counts occurred between day 0 and 7 (3.1 x 106/\u3bcl) and remained low up to day 35 post infection (3.5 x 106/\u3bcl). This was followed by an increase in WBC counts, principally associated with increased lymphocyte numbers. It is concluded that microcytic hypochromic anaemia, thrombocytopaenia and an initial leucocytopaenia are the most important haematological changes associated with a chronic infection of T.b. rhodesiense infection in vervet monkeys