N-glycosylation modulates enzymatic activity of Trypanosoma congolense trans-sialidase

Trypanosomes cause the devastating disease trypanosomiasis, in which the action of trans-sialidase (TS) enzymes harbored on their surface is a key virulence factor. TS are highly N-glycosylated, but the biological functions of the glycans remain elusive. In this study, we investigated the influence of N-glycans on the enzymatic activity and structure stability of TconTS1, a recombinant TS from the African parasite Trypanosoma congolense. MALDI-TOF MS revealed that eight asparagine sites were glycosylated with high-mannose type N-glycans. Deglycosylation of TconTS1 led to a 5-fold decrease in substrate affinity but to the same conversion rate relative to the untreated enzyme. After deglycosylation, no changes in secondary structure elements were observed in circular dichroism experiments. Molecular dynamics simulations revealed interactions between the highly flexible N-glycans and some conserved amino acids belonging to the catalytic site. These interactions led to conformational changes, possibly enhancing substrate accessibility and promoting enzyme/substrate complex stability. The here-observed modulation of catalytic activity via the N-glycan shield may be a structure-function relationship intrinsic of several members of the TS family.

Infection of West African Dwarf Rams With Trypanosoma Brucei Brucei and Trypanosoma Congolense Significantly Alter Serum Electrolytes, Redox Balance, Sperm Parameters, And Gonadal Morphology

Trypanotolerance of the West African dwarf (WAD) breeds may not rule out significant pathophysiological changes that may affect productivity. In this study, the effects of infection of WAD rams with Trypanosoma brucei brucei (Tbb) and Trypanosoma congolense (Tc) on their serum levels of electrolytes [calcium, phosphorus, sodium, potassium]; oxidative stress markers [superoxide dismutase (SOD), malondialdehyde (MDA)]; and sperm parameters [sperm count, motility, vitality, and morphology] were investigated. Fifteen WAD rams, assigned to 3 groups (A, B & C) of 5 rams each, were used for the study. Group A rams were infected with Tbb, while Group B rams were infected with Tc, both intraperitoneally, at the dose of 106 trypanosomes/animal. Group C rams served as the uninfected control. The infections were monitored for 70 days. Serum calcium levels were significantly (p < 0.05) lower in Tbb and Tc infected rams compared to the control throughout the study. Serum sodium was significantly (p < 0.05) higher in the Tb infected rams compared to the Tc infected and control rams on days 14 and 28 PI. Serum SOD activity decreased while MDA levels increased in both infected groups of rams. Tbb infected rams were azoospermic, while Tc infected rams had lower sperm motility, vitality and concentration and higher number of abnormal sperm cells compared to the control. Necrotic and inflammatory lesions occurred in the testis and epididymis of both infected rams. These results suggest that in spite of trypanotolerance, trypanosome infections in the WAD rams significantly impact on health and reproduction.

Haematological changes in Wistar rats experimentally infected with Trypanosoma congolense and Trypanosoma brucei brucei obtained from North-west Nigeria

This study determined haematological changes in Wistar rats experimentally infected with local strains of Trypanosoma congolense and Trypanosoma brucei brucei. Forty-five Wistar rats between 10 – 12 weeks old weighing between 210 – 240 g were used. The Wistar rats were randomly divided into four groups (A, B, C and D), with the infected groups (B, C and D) having 10 rats each, while the uninfected control group (A) had 15 rats. Group A rats were not infected and served as the control, group B were infected with Trypanosoma congolense, group C were infected with Trypanosoma brucei brucei and group D were co-infected with Trypanosoma congolense and Trypanosoma brucei brucei. Infection was achieved using 0.1mL of blood containing approximately 1 × 103 trypanosomes intraperitoneally into each Wistar rat in the infected groups. Clinical signs were observed. The changes in the blood cells were assayed in the groups post-infection. Duncan’s Least Square Deviation showed significantly (p<0.05) higher parasitaemia in infected groups. However, group D showed a higher significant (p<0.05) difference in parasitaemia when compared to groups B and C. The pattern of mean parasitaemia for the infected groups, revealed a positive correlation with days of post-infection (p<0.05) before the decline. The packed cell volume, total red blood cell count and haemoglobin concentration were significantly (p<0.05) lower in infected groups B, C and D. The total white blood cell count, platelet counts and differential leucocyte count were significantly (p<0.05) lower in infected groups when compared to the uninfected group. These findings suggest that co-infection with Trypanosoma congolense and Trypanosoma brucei brucei obtained from Wurno and Ngaski in Sokoto and Kebbi States respectively produced a more damaging effect on haematological parameters.

Divergent metabolism between Trypanosoma congolense and Trypanosoma brucei results in differential sensitivity to metabolic inhibition

Animal African Trypanosomiasis (AAT) is a debilitating livestock disease prevalent across sub-Saharan Africa, a main cause of which is the protozoan parasite Trypanosoma congolense. In comparison to the well-studied T. brucei, there is a major paucity of knowledge regarding the biology of T. congolense. Here, we use a combination of omics technologies and novel genetic tools to characterise core metabolism in T. congolense mammalian-infective bloodstream-form parasites, and test whether metabolic differences compared to T. brucei impact upon sensitivity to metabolic inhibition. Like the bloodstream stage of T. brucei, glycolysis plays a major part in T. congolense energy metabolism. However, the rate of glucose uptake is significantly lower in bloodstream stage T. congolense, with cells remaining viable when cultured in concentrations as low as 2 mM. Instead of pyruvate, the primary glycolytic endpoints are succinate, malate and acetate. Transcriptomics analysis showed higher levels of transcripts associated with the mitochondrial pyruvate dehydrogenase complex, acetate generation, and the glycosomal succinate shunt in T. congolense, compared to T. brucei. Stable-isotope labelling of glucose enabled the comparison of carbon usage between T. brucei and T. congolense, highlighting differences in nucleotide and saturated fatty acid metabolism. To validate the metabolic similarities and differences, both species were treated with metabolic inhibitors, confirming that electron transport chain activity is not essential in T. congolense. However, the parasite exhibits increased sensitivity to inhibition of mitochondrial pyruvate import, compared to T. brucei. Strikingly, T. congolense exhibited significant resistance to inhibitors of fatty acid synthesis, including a 780-fold higher EC50 for the lipase and fatty acid synthase inhibitor Orlistat, compared to T. brucei. These data highlight that bloodstream form T. congolense diverges from T. brucei in key areas of metabolism, with several features that are intermediate between bloodstream- and insect-stage T. brucei. These results have implications for drug development, mechanisms of drug resistance and host-pathogen interactions.