Trypanosome Challenge Research Articles

Genetic parameters and correlations between growth traits and packed cell volume of N'Dama cattle in the Gambia.

The evolution of body weight under the natural trypanosome challenge and its association with disease tolerance to trypanosomosis is of utmost economic importance in cattle. This study estimated heritability for growth traits and packed cell volume (PCV) and their genetic correlations in the N'Dama cattle in the Gambia. A total of 2,488, 2,442, 1,471, 1,934, and 1,452 bodyweight records at 12months (WT12), 16months (WT16), 18months (WT18), 24months (WT24), 36months (WT36) and 50months (WT50) and 1,782, 1,800, 1,844, 1,608, and 1,459 records for PCV at 12months (PCV12) 18months (PCV18), 24months (PCV24), 36months (PCV36), and 50months (PCV50), respectively, were analysed. Genetic parameters were estimated using univariate and multivariate animal models using the GIBBSF90 software. Least square means for WT12, WT18, WT24, WT36 and WT50 were 75.08 ± 0.37kg, 95.58 ± 0.52, 123.80 ± 0.52kg, 149.90 ± 0.73kg and 190.37 ± 0.68kg, respectively. The Least square means for PCV declined from 24.56 ± 0.18 at 12months of age to 23.18 ± 0.19 at 50months. Heritability estimates for growth ranged from 0.47 ± 0.05 at 12months to 0.31 ± 0.06 at 50months, while for PCV the estimates were 0.09 ± 0.01 to 0.15 ± 0.01. Genetic correlations between bodyweights were high and positive (0.73 ± 0.01 to 0.96 ± 0.01) while those between PCV traits were low to medium and positive (0.16 ± 0.04 to 0.56 ± 0.02). Genetic correlations between bodyweight and PCV range from -0.14 ± 0.02 to 0.59 ± 0.02. BW18 and PCV18 had high heritability estimates and as well as the highest genetic correlation and therefore could be used as selection criteria for body weight and trypanotolerance, respectively.

Evaluation of biochemical indices in indigenous cattle breeds and crossbred genotypes under natural trypanosome challenge

Breeding for trypanotolerance could be a more sustainable control strategy against trypanosomiasis in African livestock species. This study was aimed at examining the biochemical indices of adult cattle (≥ 36 months of age) under natural trypanosome challenge. The cattle herd comprised of purebred N’dama (ND), Muturu (MU), Sokoto Gudahli (GU), Red Bororo (BO), and White Fulani (WF); and various crossbred genotypes. Animals sampled were ND (n = 21), WF (n = 37), GUxND (n = 3), NDxWF (n = 3); 3/4ND1/4WF (n = 3), ND(WFxMU)) (n = 6), ND(GUxWF) (n = 3), ND(BOxWF) (n = 3), and ND(GUxWFxMU) (n = 12). Serum levels of total protein, albumin, globulin, aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), bilirubin, creatinine and urea were compared among cattle genotypes, anaemic and non-anaemic, and parasitaemic and aparasitaemic groups. Non-anaemic cattle had higher (p<0.05) total protein and albumin compared to anaemic cattle. Globulin, albumin-globulin ratio, ALT, ALP, and creatinine did not differ significantly between cattle genotypes but AST was highest in anaemic NDxWF (38.50 ± 6.19 u/L) while indirect bilirubin and urea were highest in non-anaemic 3/4ND1/4WF. Aparasitaemic cattle had higher total protein, albumin and ALP but lower creatinine than parasitaemic cattle. AST and total bilirubin were highest in ND(GUxWFxMU) (33.18 ± 1.78 u/L) and ND(GUxWF) (0.30 ± 0.03 mg/dL), respectively and least in parasitaemic WF (24.63 ± 3.57 u/L) and NDxWF (0.19 ± 0.03 mg/dL), respectively. Creatinine, indirect bilirubin and urea were highest in 3/4ND1/4WF (1.23 ± 0.26 mg/dL, 0.23 ± 0.03 mg/dL, and 39.68 ± 3.05 mg/dL, respectively). ND (0.28 ± 0.10 mg/dL), parasitaemic WF (0.11 ± 0.03mg/dL), and ND(BOxWF) (22.84 ± 3.05 mg/dL) had lowest values of creatinine, indirect bilirubin and urea, respectively. Crossbred cattle and ND had higher serum total protein, albumin, AST, total and indirect bilirubin, but lower creatinine compared to WF cattle indicating better body nutrient reserves, lower oxidative stress challenge and stronger trypanotolerance capacity. La sélection d’animaux trypanotolérantes pourrait constituer une stratégie de lutte plus durable contre la trypanosomose affectant le cheptel africain. Cette étude visait à analyser les indices biochimiques chez des bovins adultes (âgés d’au moins 36 mois) exposés naturellement aux trypanosomes. L’échantillon étudié comprenait des races bovines pures N’Dama (ND), Muturu (MU), Sokoto Gudahli (GU), Red Bororo (BO) et White Fulani (WF), ainsi que divers génotypes issus de croisements. Les animaux échantillonnés étaient répartis comme suit : ND (n = 21), WF (n = 37), GUxND (n = 3), NDxWF (n = 3) ; 3/4ND1/4WF (n = 3), ND(WFxMU)) (n = 6), ND(GUxWF) (n = 3), ND(BOxWF) (n = 3) et ND(GUxWFxMU) (n = 12). Les concentrations sériques de protéines totales, d’albumine, de globuline, d’aspartate aminotransférase (AST), d’alanine aminotransférase (ALT), de phosphatase alcaline (ALP), de bilirubine, de créatinine et d’urée ont été comparées entre les génotypes bovins, les groupes anémiques et non anémiques, et les groupes parasitémiques et aparasitémiques. Les bovins non anémiques présentaient des taux de protéines totales et d’albumine significativement plus élevés (p < 0,05) que les bovins anémiques. La globuline, le rapport albumine/globuline, l’ALT, la PAL et la créatinine ne présentaient pas de différences significatives entre les génotypes bovins. L’AST était toutefois plus élevée chez les NDxWF anémiques (38,50 ± 6,19 u/L), tandis que la bilirubine indirecte et l’urée étaient plus élevées chez les 3/4ND1/4WF non anémiques. Les bovins aparasitémiques affichaient des taux plus élevés de protéines totales, d’albumine et de PAL, mais une créatinine plus faible que les bovins parasitémiques. L’AST et la bilirubine totale étaient les plus élevées chez ND(GUxWFxMU) (33,18 ± 1,78 u/L) et ND(GUxWF) (0,30 ± 0,03 mg/dL), respectivement, et les plus faibles chez les WF parasitémiques (24,63 ± 3,57 u/L) et NDxWF (0,19 ± 0,03 mg/dL), respectivement. La créatinine, la bilirubine indirecte et l’urée étaient les plus élevées chez les 3/4ND1/4WF (1,23 ± 0,26 mg/dL, 0,23 ± 0,03 mg/dL et 39,68 ± 3,05 mg/dL, respectivement). ND (0,28 ± 0,10 mg/dL), les WF parasitémiques (0,11 ± 0,03 mg/dL) et ND(BOxWF) (22,84 ± 3,05 mg/dL) présentaient quant à eux les valeurs les plus faibles de créatinine, de bilirubine indirecte et d’urée. Les bovins croisés et les ND présentaient des taux sériques plus élevés de protéines totales, d’albumine, d’AST, de bilirubine totale et indirecte, mais une créatinine plus faible par rapport aux bovins WF, indiquant de meilleures réserves en nutriments corporels, un stress oxydatif plus faible et une capacité de trypanotolérance supérieure.

Detrimental Effect ofTrypanosoma brucei bruceiInfection on Memory B Cells and Host Ability to Recall Protective B-cell Responses

Trypanosoma brucei brucei evades host immune responses by multiple means, including the disruption of B-cell homeostasis. This hampers anti-trypanosome vaccine development. Because the cellular mechanism underlying this pathology has never been addressed, our study focuses on the fate of memory B cells (MBCs) in vaccinated mice upon trypanosome challenge. A trypanosome variant surface glycoprotein (VSG) and fluorescent phycoerythrin were used as immunization antigens. Functional and cellular characteristics of antigen-specific MBCs were studied after homologous and heterologous parasite challenge. Immunization with AnTat1.1 VSG triggers a specific antibody response and isotype-switched CD73+CD273+CD80+ MBCs, delivering 90% sterile protection against a homologous parasite challenge. As expected, AnTat1.1 VSG immunization does not protect against infection with heterologous VSG-switched parasites. After successful curative drug treatment, mice were shown to have completely lost their previously induced protective immunity against the homologous parasites, coinciding with the loss of vaccine-induced MBCs. A phycoerythrin immunization approach confirmed that trypanosome infections cause the general loss of antigen-specific splenic and bone marrow MBCs and a reduction in antigen-specific immunoglobulin G. Trypanosomosis induces general immunological memory loss. This benefits the parasites by reducing the stringency for antigenic variation requirements.

Epidemiology of Trypanosomiasis in Wildlife-Implications for Humans at the Wildlife Interface in Africa.

While both human and animal trypanosomiasis continue to present as major human and animal public health constraints globally, detailed analyses of trypanosome wildlife reservoir hosts remain sparse. African animal trypanosomiasis (AAT) affects both livestock and wildlife carrying a significant risk of spillover and cross-transmission of species and strains between populations. Increased human activity together with pressure on land resources is increasing wildlife–livestock–human infections. Increasing proximity between human settlements and grazing lands to wildlife reserves and game parks only serves to exacerbate zoonotic risk. Communities living and maintaining livestock on the fringes of wildlife-rich ecosystems require to have in place methods of vector control for prevention of AAT transmission and for the treatment of their livestock. Major Trypanosoma spp. include Trypanosoma brucei rhodesiense, Trypanosoma brucei gambiense, and Trypanosoma cruzi, pathogenic for humans, and Trypanosoma vivax, Trypanosoma congolense, Trypanosoma evansi, Trypanosoma brucei brucei, Trypanosoma dionisii, Trypanosoma thomasbancrofti, Trypanosma elephantis, Trypanosoma vegrandis, Trypanosoma copemani, Trypanosoma irwini, Trypanosoma copemani, Trypanosoma gilletti, Trypanosoma theileri, Trypanosoma godfreyi, Trypansoma simiae, and Trypanosoma (Megatrypanum) pestanai. Wildlife hosts for the trypansomatidae include subfamilies of Bovinae, Suidae, Pantherinae, Equidae, Alcephinae, Cercopithecinae, Crocodilinae, Pteropodidae, Peramelidae, Sigmodontidae, and Meliphagidae. Wildlife species are generally considered tolerant to trypanosome infection following centuries of coexistence of vectors and wildlife hosts. Tolerance is influenced by age, sex, species, and physiological condition and parasite challenge. Cyclic transmission through Glossina species occurs for T. congolense, T. simiae, T. vivax, T. brucei, and T. b. rhodesiense, T. b. gambiense, and within Reduviid bugs for T. cruzi. T. evansi is mechanically transmitted, and T. vixax is also commonly transmitted by biting flies including tsetse. Wildlife animal species serve as long-term reservoirs of infection, but the delicate acquired balance between trypanotolerance and trypanosome challenge can be disrupted by an increase in challenge and/or the introduction of new more virulent species into the ecosystem. There is a need to protect wildlife, animal, and human populations from the infectious consequences of encroachment to preserve and protect these populations. In this review, we explore the ecology and epidemiology of Trypanosoma spp. in wildlife.

Genomic scan of selective sweeps in Djallonk\xe9 (West African Dwarf) sheep shed light on adaptation to harsh environments

The Djallonké (West African Dwarf) sheep is a small-sized haired sheep resulting from a costly evolutionary process of natural adaptation to the harsh environment of West Africa including trypanosome challenge. However, genomic studies carried out in this sheep are scant. In this research, genomic data of 184 Djallonké sheep (and 12 Burkina-Sahel sheep as an outgroup) generated using medium-density SNP Chips were analyzed. Three different statistics (iHS, XP-EHH and nSL) were applied to identify candidate selection sweep regions spanning genes putatively associated with adaptation of sheep to the West African environment. A total of 207 candidate selection sweep regions were defined. Gene-annotation enrichment and functional annotation analyses allowed to identify three statistically significant functional clusters involving 12 candidate genes. Genes included in Functional Clusters associated to selection signatures were mainly related to metabolic response to stress, including regulation of oxidative and metabolic stress and thermotolerance. The bovine chromosomal areas carrying QTLs for cattle trypanotolerance were compared with the regions on which the orthologous functional candidate cattle genes were located. The importance of cattle BTA4 for trypanotolerant response might have been conserved between species. The current research provides new insights on the genomic basis for adaptation and highlights the importance of obtaining information from non-cosmopolite livestock populations managed in harsh environments.

BH3 domain-independent apolipoprotein L1 toxicity rescued by BCL2 prosurvival proteins

The potent trypanolytic properties of human apolipoprotein L1 (APOL1) can be neutralized by the trypanosome variant surface antigen gene product known as serum resistance-associated protein. However, two common APOL1 haplotypes present uniquely in individuals of West African ancestry each encode APOL1 variants resistant to serum resistance-associated protein, and each confers substantial resistance to human African sleeping sickness. In contrast to the dominantly inherited anti-trypanosomal activity of APOL1, recessive inheritance of these two trypanoprotective APOL1 alleles predisposes to kidney disease. Proposed mechanisms of APOL1 toxicity have included BH3 domain-dependent autophagy and/or ion channel activity. We probed these potential mechanisms by expressing APOL1 in Xenopus laevis oocytes. APOL1 expression in oocytes increased ion permeability and caused profound morphological deterioration (toxicity). Coexpression of BCL2 family members rescued APOL1-associated oocyte toxicity in the order MCL1 ∼ BCLW > BCLXL ∼ BCL2A1 ≫ BCL2. Deletion of nine nominal core BH3 domain residues abolished APOL1-associated toxicity, but missense substitution of the same residues abolished neither oocyte toxicity nor its rescue by coexpressed MCL1. The APOL1 BH3 domain was similarly dispensable for the ability of APOL1 to rescue intact mice from lethal trypanosome challenge. Replacement of most extracellular Na(+) by K(+) also reduced APOL1-associated oocyte toxicity, allowing demonstration of APOL1-associated increases in Ca(2+) and Cl(-) fluxes and oocyte ion currents, which were similarly reduced by MCL1 coexpression. Thus APOL1 toxicity in Xenopus oocytes is BH3-independent, but can nonetheless be rescued by some BCL2 family proteins.

Midgut expression of immune-related genes in Glossina palpalis gambiensis challenged with Trypanosoma brucei gambiense.

Tsetse flies from the subspecies Glossina morsitans morsitans and Glossina palpalis gambiensis, respectively, transmit Trypanosoma brucei rhodesiense and Trypanosoma brucei gambiense. The former causes the acute form of sleeping sickness, and the latter provokes the chronic form. Although several articles have reported G. m. morsitans gene expression following trypanosome infection, no comparable investigation has been performed for G. p. gambiensis. This report presents results on the differential expression of immune-related genes in G. p. gambiensis challenged with T. b. gambiense. The aim was to characterize transcriptomic events occurring in the tsetse gut during the parasite establishment step, which is the crucial first step in the parasite development cycle within its vector. The selected genes were chosen from those previously shown to be highly expressed in G. m. morsitans, to allow further comparison of gene expression in both Glossina species. Using quantitative PCR, genes were amplified from the dissected midguts of trypanosome-stimulated, infected, non-infected, and self-cleared flies at three sampling timepoints (3, 10, and 20 days) after a bloodmeal. At the 3-day sampling point, transferrin transcripts were significantly up-regulated in trypanosome-challenged flies versus flies fed on non-infected mice. In self-cleared flies, serpin-2 and thioredoxin peroxidase-3 transcripts were significantly up-regulated 10 days after trypanosome challenge, whereas nitric oxide synthase and chitin-binding protein transcripts were up-regulated after 20 days. Although the expression levels of the other genes were highly variable, the expression of immune-related genes in G. p. gambiensis appears to be a time-dependent process. The possible biological significance of these findings is discussed, and the results are compared with previous reports for G. m. morsitans.

An Alternative Model for the Role of RP2 Protein in Flagellum Assembly in the African Trypanosome

The tubulin cofactor C domain-containing protein TbRP2 is a basal body (centriolar) protein essential for axoneme formation in the flagellate protist Trypanosoma brucei, the causal agent of African sleeping sickness. Here, we show how TbRP2 is targeted and tethered at mature basal bodies and provide novel insight into TbRP2 function. Regarding targeting, understanding how several hundred proteins combine to build a microtubule axoneme is a fundamental challenge in eukaryotic cell biology. We show that basal body localization of TbRP2 is mediated by twinned, N-terminal TOF (TON1, OFD1, and FOP) and LisH motifs, motifs that otherwise facilitate localization of only a few conserved proteins at microtubule-organizing centers in animals, plants, and flagellate protists. Regarding TbRP2 function, there is a debate as to whether the flagellar assembly function of specialized, centriolar tubulin cofactor C domain-containing proteins is processing tubulin, the major component of axonemes, or general vesicular trafficking in a flagellum assembly context. Here we report that TbRP2 is required for the recruitment of T. brucei orthologs of MKS1 and MKS6, proteins that, in animal cells, are part of a complex that assembles at the base of the flagellum to regulate protein composition and cilium function. We also identify that TbRP2 is detected by YL1/2, an antibody classically used to detect α-tubulin. Together, these data suggest a general processing role for TbRP2 in trypanosome flagellum assembly and challenge the notion that TbRP2 functions solely in assessing tubulin "quality" prior to tubulin incorporation into the elongating axoneme.

Trypanotolerance in N’Dama x Boran crosses under natural trypanosome challenge: effect of test-year environment, gender, and breed composition

BackgroundTrypanosomosis, a protozoal disease affecting livestock, transmitted by Glossina (tsetse) flies is a major constraint to agricultural production in Sub-Saharan Africa. It is accepted that utilization of the native trypanotolerance exhibited in some of the African cattle breeds to improve trypanotolerance of more productive but susceptible breeds, will offer a cost effective and sustainable solution to the problem. The success of this approach is based on the premise that quantitative trait loci previously identified under relatively controlled situations confer useful trypanotolerance under natural field situations. As part of a study to authenticate this hypothesis, a population of 192 cattle, consisting of six batches of N’Dama and Kenya-Boran backcross animals [(N’Dama x Kenya-Boran) x Kenya-Boran] born over the period 2002 to 2006 was constructed. Some of the batches also included pure Kenya-Boran cattle, or N’Dama x Kenya- Boran F1 animals. Each batch was exposed as yearlings to natural field trypanosomosis challenge over a period of about one year; the entire challenge period extending from December 2003 to June 2007. Performance of the animals was evaluated by weekly or biweekly measurements of body weight, packed blood cell volume (PCV), parasitemia score, and number of trypanocide treatments. From these basic data, 49 phenotypes were constructed reflecting dynamics of body weight, packed cell volume (PCV) and parasitemia under challenge.ResultsFemales were distinctly more trypanotolerant than males. F1, backcross and pure Kenya- Boran animals ranked in that order with respect to trypanotolerance. Overall batch effects were highly significant (p<0.001) for most traits, and were generally more significant than the gender or genetic type effects. The superior trypanotolerance of the F1 animals was expressed in all three components of animal defense strategies against pathogens: Avoidance resistance, and tolerance.ConclusionsThe results show that trypanotolerance derived from the N’Dama is expressed under field conditions; and that the trait is primarily additive in nature, being expressed in heterozygous condition and in a three-quarters Boran genetic background. The results further, underscore the complexity of the trait in the field manifesting all three host disease-control strategies, and show the importance of gender and local environmental conditions in determining response to challenge.

Nutritional stress affects the tsetse fly's immune gene expression

Tsetse-transmitted trypanosomiasis poses a serious threat to human and animal health in sub-Saharan Africa. The majority of tsetse flies (Glossina spp.) in a natural population will not develop a mature infection of either Trypanosoma congolense or Trypanosoma brucei sp. because of refractoriness, a phenomenon that is affected by different factors, including the tsetse fly's immune defence. Starvation of tsetse flies significantly increases their susceptibility to the establishment of a trypanosome infection. This paper reports the effects of nutritional stress (starvation) on (a) uninduced baseline levels of gene expression of the antimicrobial peptides attacin, defensin and cecropin in the tsetse fly, and (b) levels of expression induced in response to bacterial (Escherichia coli) or trypanosomal challenge. In newly emerged, unfed tsetse flies, starvation significantly lowers baseline levels of antimicrobial peptide gene expression, especially for attacin and cecropin. In response to trypanosome challenge, only non-starved older flies showed a significant increase in antimicrobial peptide gene expression within 5 days of ingestion of a trypanosome-containing bloodmeal, especially with T. brucei bloodstream forms. These data suggest that a decreased expression of immune genes in newly hatched flies or a lack of immune responsiveness to trypanosomes in older flies, both occurring as a result of fly starvation, may be among the factors contributing to the increased susceptibility of nutritionally stressed tsetse flies to trypanosome infection.

Activity of Indenoisoquinolines against African Trypanosomes

African trypanosomiasis (sleeping sickness), caused by protozoan Trypanosoma brucei species, is a debilitating disease that is lethal if untreated. Available drugs are antiquated, toxic, and compromised by emerging resistance. The indenoisoquinolines are a class of noncamptothecin topoisomerase IB poisons that are under development as anticancer agents. We tested a variety of indenoisoquinolines for their ability to kill T. brucei. Indenoisoquinolines proved trypanocidal at submicromolar concentrations in vitro. Structure-activity analysis yielded motifs that enhanced potency, including alkylamino substitutions on N-6, methoxy groups on C-2 and C-3, and a methylenedioxy bridge between C-8 and C-9. Detailed analysis of eight water-soluble indenoisoquinolines demonstrated that in trypanosomes the compounds inhibited DNA synthesis and acted as topoisomerase poisons. Testing these compounds on L1210 mouse leukemia cells revealed that all eight were more effective against trypanosomes than against mammalian cells. In preliminary in vivo experiments one compound delayed parasitemia and extended survival in mice subjected to a lethal trypanosome challenge. The indenoisoquinolines provide a promising lead for the development of drugs against sleeping sickness.

A Glycosylphosphatidylinositol-Based Treatment Alleviates Trypanosomiasis-Associated Immunopathology

The GPI-anchored trypanosome variant surface glycoprotein (VSG) triggers macrophages to produce TNF, involved in trypanosomiasis-associated inflammation and the clinical manifestation of sleeping sickness. Aiming at inhibiting immunopathology during experimental Trypanosoma brucei infections, a VSG-derived GPI-based treatment approach was developed. To achieve this, mice were exposed to the GPI before an infectious trypanosome challenge. This GPI-based strategy resulted in a significant prolonged survival and a substantial protection against infection-associated weight loss, liver damage, acidosis, and anemia; the latter was shown to be Ab-independent and correlated with reduced macrophage-mediated RBC clearance. In addition, GPI-based treatment resulted in reduced circulating serum levels of the inflammatory cytokines TNF and IL-6, abrogation of infection-induced LPS hypersensitivity, and an increase in circulating IL-10. At the level of trypanosomiasis-associated macrophage activation, the GPI-based treatment resulted in an impaired secretion of TNF by VSG and LPS pulsed macrophages, a reduced expression of the inflammatory cytokine genes TNF, IL-6, and IL-12, and an increased expression of the anti-inflammatory cytokine gene IL-10. In addition, this change in cytokine pattern upon GPI-based treatment was associated with the expression of alternatively activated macrophage markers. Finally, the GPI-based treatment also reduced the infection-associated pathology in Trypanosoma congolense and Trypanosoma evansi model systems as well as in tsetse fly challenge experiments, indicating potential field applicability for this intervention strategy.

The pathogenesis of anaemia in goats experimentally infected with Trypanosoma congolense or Trypanosoma brucei: Use of the myeloid:erythroid ratio

The present study examined the development of anaemia in Small East African goats experimentally infected with Trypanosoma congolense or Trypanosoma brucei. Experimental goats received a primary trypanosome challenge on day 0, treated with diminazene aceturate on day 49 and received a secondary trypanosome challenge on day 77 of the 136-day experiment. Both primary and secondary challenges were characterised by reduced peripheral erythrocyte counts, fall in packed cell volume (PCV), hypohaemoglobinaemia and reductions in the myeloid:erythroid ratios (M:E) compared with the uninfected goats. The progressive reduction in the M:E ratios denoted increased erythrogenesis in response to increased destruction of erythrocytes in blood by infecting trypanosomes or their products. The more rapid fall in M:E ratio in T. congolense infections shows that this parasite causes more severe clinical pathological effects in goats than T. brucei

Adult midgut expressed sequence tags from the tsetse fly Glossina morsitans morsitans and expression analysis of putative immune response genes.

Tsetse flies transmit African trypanosomiasis leading to half a million cases annually. Trypanosomiasis in animals (nagana) remains a massive brake on African agricultural development. While trypanosome biology is widely studied, knowledge of tsetse flies is very limited, particularly at the molecular level. This is a serious impediment to investigations of tsetse-trypanosome interactions. We have undertaken an expressed sequence tag (EST) project on the adult tsetse midgut, the major organ system for establishment and early development of trypanosomes. A total of 21,427 ESTs were produced from the midgut of adult Glossina morsitans morsitans and grouped into 8,876 clusters or singletons potentially representing unique genes. Putative functions were ascribed to 4,035 of these by homology. Of these, a remarkable 3,884 had their most significant matches in the Drosophila protein database. We selected 68 genes with putative immune-related functions, macroarrayed them and determined their expression profiles following bacterial or trypanosome challenge. In both infections many genes are downregulated, suggesting a malaise response in the midgut. Trypanosome and bacterial challenge result in upregulation of different genes, suggesting that different recognition pathways are involved in the two responses. The most notable block of genes upregulated in response to trypanosome challenge are a series of Toll and Imd genes and a series of genes involved in oxidative stress responses. The project increases the number of known Glossina genes by two orders of magnitude. Identification of putative immunity genes and their preliminary characterization provides a resource for the experimental dissection of tsetse-trypanosome interactions.

Activity of human trypanosome lytic factor in mice

The inability of the cattle pathogen Trypanosoma brucei brucei to infect humans is due to an innate factor in human serum termed Trypanosome Lytic Factor (TLF). Human haptoglobin-related protein is the proposed toxin in TLF and can exist either as a component of a minor subclass of high-density lipoprotein (TLF-1) or as a lipid free, high molecular weight protein complex (TLF-2). The trypanolytic activity of both TLF-1 and TLF-2 has been studied in vitro but their relative contributions to protection against T. b. brucei infection in vivo has not been established. In the present studies we show that treatment of T. b. brucei infected mice with TLF-1 resulted in a dose dependent decrease in parasite numbers but did not affect parasite numbers in mice infected with Trypanosoma brucei rhodesiense, the causative agent of the human sleeping sickness. Similarly, pretreatment of mice with TLF-1 resulted in protection against a challenge by T. b. brucei but had no effect on T. b. rhodesiense challenge. Induction of the acute phase protein haptoglobin, a natural antagonist of TLF-1, diminished but did not abolish the protection against trypanosome challenge. In addition, haptoglobin knockout mice showed higher levels of TLF-1 mediated protection against a T. b. brucei challenge. These results suggest that while TLF-1 is active in vivo, even in the presence of elevated levels of haptoglobin, its activity is modulated in a dose dependent fashion by haptoglobin in the circulation.

Comparison of the isometamidium-based trypanocidal drugs Samorin® and Veridium® in cattle under field conditions at Nguruman, Kenya

The trypanocidal activity of two commercially available isometamidium-based products, Samorin® (Merial, USA) and Veridium® (Sanofi Santé Nutrition Animale, France), used at a dose rate of 0.5 mg kg −1 bodyweight, was compared in a field trial involving groups of approximately 30 zebu cattle in a trypanosomosis endemic part of south-western Kenya. The trial took place between April 1997 and March 1998 during a time of higher than normal rainfall that resulted in periods of high trypanosome challenge. The trial consisted of five consecutive prophylactic cycles, each of approximately 10 weeks duration. It was demonstrated that there was no significant difference in the prophylactic activity of the two isometamidium-based products, and no significant difference between the relative activity of three different batches of the product Veridium® used during the course of the trial. There was some evidence that drug-resistant strains of trypanosomes may have been present, but it was concluded that isometamidium is still an effective trypanocidal drug in this location.

Innate resistance to experimental Trypanosoma congolense infection: differences in IL-10 synthesis by macrophage cell lines from resistant and susceptible inbred mice.

BALB/c and C57Bl/6 mice differ in resistance to T. congolense infections. We investigated the production of various cytokines (IL-10, IL-6, TNF-alpha and TGF-beta) by macrophages from these mice. Macrophage cell lines (BALB.BM cells) of BALB/c mice but not (ANA-I cells)of C57BL/6 mice constitutively produced IL-10. Challenge of these cells with trypanosomes induced the production of 50-100 times more IL-10 in BALB.BM cells than in ANA-1 cells. Pre-incubation of the cell lines with IFN-gamma. prior to the trypanosome challenge, further upregulated this IL-10 production in BALB.BM but not in ANA-1 cells. Primary cultures of bone marrow-derived macrophages (BMDM) from BALB/c mice also produced more IL-10 following challenge with IFN-gamma and opsonized trypanosomes than did the C57Bl/6 BMDM. Similarly after challenge with trypanosomes, BALB.BM and BALB/c BMDM produced significantly more IL-6 than did the analogous cells from the C57Bl/6 mice following such challenges. Higher steady state levels of TNF-alpha mRNA accumulated in ANA-1 cells than in BALB.BM cells following challenge with IFN-gamma and opsonized trypanosomes. Findings of this study for the first time indicate a differential regulation of cytokines (IL-10, IL-6 and TNF-alpha) in macrophages of mice that significantly differ in their susceptibility to infections with T. congolense.

Health and productivity of traditionally managed Djallonke sheep and West African dwarf goats under high and moderate trypanosomosis risk

Trypanosome infections, packed red cell volume levels (PCV), body weight and nematode faecal egg counts of village-based small ruminants were monitored in two areas in The Gambia with either moderate or high trypanosomosis risk for 24 and 30 months respectively. Outflows from the flock and new-born animals were recorded and data on housing and management were compiled. Reported mortality rates were higher in goats than in sheep, but for both species highest in the moderate risk area. The peak of trypanosome infections lagged the peak of tsetse densities by 1–3 months in both areas. Trypanosoma vivax was the predominant species found in the infected animals, followed by T. congolense. Trypanosome prevalence was, in general, higher in sheep than in goats but only significantly higher during Year 1 in the moderate risk area. Trypanosome infection reduced the PCV level significantly and seasonal effects indicated significantly lower PCV levels during the rains. Trypanosome infection significantly depressed weight gain in both species at periods where infection rates were highest. In both species considerably lower weight gains were observed during the rainy season. Abortion rates were higher in goats than in sheep in both study sites, and highest in the high-risk site. Trypanosome infection in ewes in the high risk area increased lamb mortality significantly but had no effect on birth weights, nor on growth rates up to 4 months. Offspring mortality up to 4 months was generally high at both sites. Trypanosome infection in the dam between 3.5 to 7 months post parturition significantly increased parturition interval in both species. Peak faecal egg output occurred at the end of the rainy season and was highest for both species in the moderate risk site. Poor grazing management was found responsible for a seasonal nutritional constraint. Based on these results, these breeds of sheep and goats can be considered as trypanotolerant since they are able to remain productive under high and moderate levels of trypanosome challenge. Nevertheless, trypanosomosis affected their health and production level as shown by reduced PCV levels, depressed weight gains, longer parturition intervals and higher lamb mortality. In addition, during the rains, helminth infections and poor management leading to nutritional constraints had also a negative impact on health and production and therefore influenced the innate resilience to trypanosomosis in those indigenous breeds. Adaptations in management may have an equal impact as certain disease control measures to improve biological and economical returns from small ruminants in tsetse infested rural areas.

New epidemiological features on animal trypanosomiasis by molecular analysis in the pastoral zone of Sideradougou, Burkina Faso.

A multidisciplinary work was undertaken in the agropastoral zone of Sidéradougou, Burkina Faso to try to elucidate the key factors determining the presence of tsetse flies. In this study the PCR was used to characterize trypanosomes infecting the vector (Glossina tachinoides and Glossina palpalis gambiensis) and the host, i.e. cattle. A 2-year survey involved dissecting 2211 tsetse of the two Glossina species. A total of 298 parasitologically infected tsetse were analysed by PCR. Trypanosoma vivax was the most frequently identified trypanosome followed by the savannah type of T. congolense and, to a lesser extent, the riverine forest type of T. congolense, and by T brucei. No cases of T. simiae were found. From the 107 identified infections in cattle, the taxa were the same, but T. congolense savannah type was more frequent, whereas T. vivax and T. congolense riverine forest types were found less frequently. A correlation was found between midgut infection rates of tsetse, nonidentified infections and reptile bloodmeals. These rates were higher in G.p. gambiensis, and in the western part of the study area. T. vivax infections were related to cattle bloodmeals, and were more frequent in G. tachinoides and in the eastern study area. The PCR results combined with bloodmeal analysis helped us to establish the relationships between the vector and the host, to assess the trypanosome challenge in the two parts of the area, to elucidate the differences between the two types of T. congolense, and to suspect that most midgut infections were originating from reptilian trypanosomes.

Control of tsetse and animal trypanosomosis using a combination of tsetse-trapping, pour-on and chemotherapy along the Uganda-Kenya border.

A joint tsetse and trypanosomosis control program has been carried out along the Uganda-Kenya border since July 1991. A combination of tsetse trapping, pour-on and chemotherapy has been used. Different combinations of control strategies were tried in the project area divided into three zones (A, B and C). In zone A, large-scale applications of pour-on, tsetse trapping (8-10 traps/km2) and chemotherapy were used. In zone B, only tsetse trapping (8-10 traps/km2) and chemotherapy were used. In zone C, block treatment of cattle in the entire area with diminazene aceturate was carried out followed by less intensive tsetse trapping (4-5 traps/km2). During monitoring, 400 cattle in each zone were screened every three months and the tsetse apparent density determined every month. From July 1991 to March 1997 reductions in the prevalence of trypanosomosis and apparent tsetse density of 94 and 99.5% in zone A, 89 and 99.5% in zone B and 79 and 95% in zone C, respectively, were achieved and maintained. The predominant Trypanosoma species found in cattle during the control period were: T. vivax in zone A, T. vivax and T. congolense in zone B, and T. vivax, T. congolense and T. brucei in zone C. Glossina fuscipes fuscipes was the only tsetse fly species caught. The most effective control strategy was an initial large-scale application of pour-on, followed by trapping and regular chemotherapy. However, control effectiveness seemed to be influenced by the level of trypanosome challenge, speed of initial reduction in tsetse density and sustainability of tsetse and trypanosomosis control inputs during the campaign. (Resume d'auteur)