APOL1 expression is induced by Trypanosoma brucei gambiense infection but is not associated with differential susceptibility to sleeping sickness

Most African trypanosome species are sensitive to trypanolytic factors (TLFs) present in human serum. Trypanosome lysis was demonstrated to be associated with apolipoprotein L-I (APOL1). Trypanosoma brucei (T. b.) gambiense and Trypanosoma brucei rhodesiense, the two human infective trypanosome species, have both developed distinct resistance mechanisms to APOL1 mediated lysis. Whereas T. b. rhodesiense resistance is linked with the expression of the serum resistance associated (SRA) protein that interacts with APOL1 inside the parasite lysosome, inhibiting its lytic action; T. b. gambiense resistance is rather controlled by a reduced expression of the parasite HpHb receptor, limiting APOL1 absorption by trypanosomes. Based on this last observation we hypothesised that variation in the host APOL1 environment could significantly alter T. b. gambiense growth and thus resistance/susceptibility to sleeping sickness. To test this hypothesis, we have measured blood APOL1 relative expression in HAT patients, uninfected endemic controls and serologically positive subjects (SERO TL⁺) that are suspected to control infection to parasitological levels that are undetectable by the available test used in the field. All RNA samples were obtained from medical surveys led in the HAT mangrove foci of Coastal Guinea. Results indicate that APOL1 expression is a complex trait dependant on a variety of factors that need to be taken into account in the analysis. Nevertheless, multivariate analysis showed that APOL1 expression levels were significantly higher in both HAT and SERO TL⁺ subject as compared to endemic controls (p = 0.006). This result suggests that APOL1 expression is likely induced by T. b. gambiense, but is not related to resistance/susceptibility in its human host.     

In Vitro Trypanocidal Activity of Antibodies to Bacterially Expressed Trypanosoma brucei Tubulin

Background

There are only four drugs for treating African trypanosomiasis, a devastating disease in sub-Saharan Africa. With slow discovery of better drugs, vaccination is viewed as the best method of control. We previously showed that antibodies to native Trypanosoma brucei brucei tubulin inhibit the growth of trypanosomes in culture. Here, we aimed to determine the effect of antibodies to bacterially expressed trypanosome tubulin on T. brucei brucei growth.

Methods

T. brucei brucei alpha and beta tubulin genes were individually expressed in Escherichia coli under the tryptophan promoter. Monoclonal tubulin antibodies reacted specifically with the expressed tubulins with no cross-reaction with the opposite tubulin. Rabbits were immunized with 450µg each of the concentrated recombinant tubulin, and production of antibodies assessed by ELISA and Western blotting. The effect of polyclonal antibodies on trypanosome growth was determined by culturing bloodstream T. brucei brucei in up to 25% of antisera.

Results

Low antisera dilutions (25%) from the immunized rabbits inhibited trypanosome growth. The most cytotoxic antisera were from one rabbit immunized with a mixture of both alpha and beta tubulins. However, the result was not reproduced in other rabbits and there was no apparent effect on growth at higher antisera dilutions.

Conclusion

Antibodies to bacterially expressed trypanosome tubulin are not effective at killing cultured bloodstream trypanosomes.     

Molecular modeling study and synthesis of novel dicationic flexible triaryl guanidines and imidamides as antiprotozoal agents

A new series of fourteen dicationic flexible triaryl bis-guanidines 3a,b, bis-N-substituted guanidines 7a,b and 8a,b as well as bis-imidamides 9–12a,b having a 1,3- or 1,4-diphenoxybenzene scaffold backbone were synthesized. The in vitro activity of the novel dications as antiprotozoal agents against Trypanosoma brucei rhodesiense (T.b.r.) and Plasmodium falciparum (P.f.) was assessed. Interestingly, six of the newly synthesized dications viz3a,b, 7a,b and 8a,b were more active against P.f. than the reference drug pentamidine. Also, some of the dications showed moderate antitrypanosomal activity. Thermal melting analysis of the novel dications was performed to determine their ligand-DNA relative binding affinities. Finally, docking of the dications with an AT rich DNA oligonucleotide was executed to understand their binding mode with the minor groove.     

The susceptibility of chickens to Trypanosoma brucei subspecies

Chickens were susceptible to infection with three different stocks of the subgenus Trypanozoon: two of presumptive Trypanosoma b. brucei and one of T. b. rhodesiense. Two groups of chickens were used: the first hatched following inoculation with either T. b. brucei or T. b. rhodesiense during embryonic development, and the second were infected as adult birds. In both experimental groups, parasitaemia persisted for prolonged periods, but was mostly subpatent and detectable only by subinoculation of blood into mice. In chickens infected as embryos, parasitaemias were patent for five weeks after hatching, but subpatent thereafter (to weeks 13 to 17). Quantitative estimations of the parasitaemias of seven of the birds hatched from embryos inoculated with T. b. brucei revealed fluctuations in the number of circulating trypanosomes, with an initial peak between days 2 to 9 after hatching. Between weeks 13 to 17 after hatching the chickens appeared to have recovered spontaneously from the trypanosome infections. Homologous challenge at week 20 failed to produce a recrudescence of parasitaemia, indicative of a possible acquired immunity.The infections of ten chickens inoculated with either T. b. brucei or T. b. rhodesiense as adult birds were microscopically subpatent throughout the observation period of six weeks, but subinoculation of blood into mice showed the chickens were parasitaemic from week one and thereafter. Different aspects of infection of avian hosts by the Trypanozoon subspecies are discussed.     

Rapid assessment of Hib disease burden in Vietnam

Background  

Several countries have applied the Haemophilus influenzae type b (Hib) rapid assessment tool (RAT) to estimate the burden of Hib disease where resources for hospital- or population-based surveillance are limited. In Vietnam, we used the Hib RAT to estimate the burden of Hib pneumonia and meningitis prior to Hib vaccine introduction.     

Enzyme electrophoresis in characterizing the causative organism of Gambian trypanosomiasis

Trypanosoma (Trypanozoon) brucei includes three morphologically identical subspecies which are poorly defined by clinical behaviour; T. b. brucei does not infect man, whereas T. b. rhodesiense causes an acute, and T. b. gambiense a chronic, disease. Thirty-three isolates of the complex, each of which had previously been identified on clinical or other criteria, were compared by the electrophoretic patterns of two trypanosomal enzymes, alanine aminotransferase (ALAT) and aspartate aminotransferase (ASAT). One particular ALAT pattern clearly segregated a group of human pathogens of which all except one were labelled T. b. gambiense. The exception was labelled T. b. rhodesiense, and in addition three putative T. b. gambiense isolates did not have this pattern; it is suggested that only one presents a serious anomaly. The T. b. gambiense group could also be subdivided by three ASAT patterns which coincided with known groupings based on serological criteria.     

Clinical and serologic responses to human 'apathogenic' trypanosomes

We describe a female patient suffering from a benign self-healing febrile disease with strongly positive serology for Trypanosoma brucei. The patient showed a clinical picture with similarities to that of human African trypanosomiasis (HAT). HAT due to T. b. gambiense and T. b. rhodesiense were ruled out. We performed serologic tests because the patient was worried about HAT after receiving tsetse bites. The possibilities of an infection with human 'apathogenic' trypanosomes such as T. b. brucei, T. congolense or T. vivax are discussed.     

First evidence that parasite infecting apparent aparasitemic serological suspects in human African trypanosomiasis are Trypanosoma brucei gambiense and are similar to those found in patients

Thanks to its sensitivity and its ease of use in the field, the card agglutination test for trypanosomiasis (CATT) is widely used for serological screening of Trypanosoma brucei gambiense human African trypanosomiasis (HAT). Positive subjects are then examined by microscopy to confirm the disease. However, the CATT exhibits false-positive results raising the question of whether CATT-positive subjects who are not confirmed by microscopic detection of trypanosomes (SERO) are truly exposed to T.b. gambiense infection. For this purpose, we applied microsatellite genotyping on DNA extracted from blood of both HAT confirmed patients and SERO subjects in Guinea and Côte d’Ivoire since microsatellite genotyping has proved useful for the study of T.b. gambiense genetic diversity. Problems of amplification failures raise the question of the sensitivity of microsatellite markers when applied on biological samples especially from SERO subjects for who low blood parasitaemia are suspected. Nevertheless, we have shown that the trypanosomes from SERO individuals that have been genotyped belong to T.b. gambiense group 1 and were identical to those found in HAT patients. These results constitute the first evidences that at least some SERO are indeed infected by T.b. gambiense group 1 and that they may constitute a human reservoir of parasite in HAT foci. Whether these individuals should undergo treatment remains an open question as long as their role in HAT transmission is unknown. Our results strongly recommend the follow-up of such subjects to improve control strategies.     

African sleeping sickness: new evidence that mature tsetse flies (Glossina morsitans) can become potent vectors

Starved mature male tsetse flies (21 to 25 days old) are capable of developing salivary gland (SG) infections of Trypanosoma brucei rhodesiense at rates nearly comparable to teneral males < 24 hours old when given an infective meal containing parasites, horse red cells and culture medium. Although the over-all SG infection rate for mature males starved for three, four or five days before infection was about half that for teneral males < 48 hours old (8 · 0% v. 15 · 6%), males starved for four days developed infection rates (12 · 3%) that were comparable to those of teneral flies < 24 hours old (11 · 8%). It is suggested that the acquisition of infection by mature flies should be considered when evaluating factors contributing either to maintenance of endemic infections or perhaps even epidemic infections of human sleeping sickness.     

Prevalence of plasmodium falciparum in active conflict areas of eastern Burma: a summary of cross-sectional data

Background  

Burma records the highest number of malaria deaths in southeast Asia and may represent a reservoir of infection for its neighbors, but the burden of disease and magnitude of transmission among border populations of Burma remains unknown.     

The testing of proven Trypanosoma brucei and T. rhodesiense strains by the blood incubation infectivity test

The authors describe a simple test (the blood incubation infectivity test) by which Trypanosoma brucei (sensu stricto) may be differentiated from T. rhodesiense without recourse to human volunteers. The method consists in incubating the strain of trypanosome under test for 5 hours at 37°C in vitro in human blood, followed by observation of the effect of this procedure on the strain''s infectivity to rats.     

Persistently elevated T cell interferon-γ responses after treatment for latent tuberculosis infection among health care workers in India: a preliminary report

Background  

T cell-based interferon-γ (IFN-γ) release assays (IGRAs) are novel tests for latent tuberculosis infection (LTBI). It has been suggested that T cell responses may be correlated with bacterial burden and, therefore, IGRAs may have a role in monitoring treatment response. We investigated IFN-γ responses to specific TB antigens among Indian health care workers (HCWs) before, and after LTBI preventive therapy.     

Rubella seroprevalence among primary and pre- primary school pupils at Moi's Bridge location, Uasin Gishu District, Kenya

Background  

Rubella is an infectious and generally mild childhood viral disease. The disease is of public health importance because infection acquired during early pregnancy often results in foetal abnormalities that are classified as congenital rubella syndrome (CRS). The burden of rubella infection in most developing countries in not well documented because of limited epidemiological data. However, availability of an effective vaccine has made it necessary to have all the countries with no routine vaccination schedule to evaluate the burden of disease in order to make informed decisions on rubella vaccination and strategy. To address this gap we conducted a study to determine age-specific rubella seroprevalence rates and related risk factors among primary and pre-primary school children in Uasin Gishu district, Moi's Bridge location of Kenya.     

Health and economic impact of rotavirus vaccination in GAVI-eligible countries

Background  

Rotavirus infection is responsible for about 500,000 deaths annually, and the disease burden is disproportionately borne by children in low-income countries. Recently the World Health Organization (WHO) has released a global recommendation that all countries include infant rotavirus vaccination in their national immunization programs. Our objective was to provide information on the expected health, economic and financial consequences of rotavirus vaccines in the 72 GAVI support-eligible countries.     

Trypanosoma brucei gambiense Type 1 populations from human patients are clonal and display geographical genetic differentiation

We have rigorously tested the hypothesis that Trypanosoma brucei gambiense Type 1 is composed of genetically homogenous populations by examining the parasite population present in Human African Trypanosomiasis (HAT) patients from the Democratic Republic of Congo (DRC) and Cameroon (CAM). We amplified eight microsatellite markers by PCR directly from blood spots on FTA filters, thereby avoiding the significant parasite selection inherent in the traditional isolation techniques of rodent inoculation or in vitro culture. All microsatellite markers were polymorphic, although for four markers there was only polymorphism between the DRC and CAM populations, not within populations, suggesting very limited genetic exchange. Within the largest population from the DRC, Hardy–Weinberg equilibrium is not evident at any loci. This evidence suggests a clonal population. However, there was significant sub-structuring between the DRC and CAM samples (F_ST = 0.32), indicating that Trypanosoma brucei gambiense Type 1 has genetically distinct clades. The data combine to indicate that genetic exchange plays a very limited role. The finding of distinct clades in different places suggests the possibility that samples from humans with clinical signs represent clonal expansions from an underlying population that requires identifying and characterising.     

A statistical study of the sensitivity of the haematocrit centrifuge technique in the detection of trypanosomes in blood

The sensitivity of the haematocrit centrifuge technique in detecting trypanosomes was investigated using both old laboratory and freshly isolated strains.Two alternative methods of analysing the results are suggested, based on the binomial and Poisson distributions.The analysis shows that the H.C.T. can detect about 85% of the trypanosomes present in capillary samples of Trypanosoma brucei, T. rhodesiense and T. evansii, and 30% of T. congolense group trypanosomes. The technique is therefore much more sensitive than previous wet preparation or thick smear techniques.Graphs are provided showing the minimum number of tubes that would have to be examined for 95% and 99% detection of low infections (i.e., in the range up to 500 trypanosomes per ml.).Analysis of the results using the Poisson model includes details of a simple method that estimates the efficiency of the H.C.T. from the slope of a graph.The present results do not show any significant relationship between the probability that a trypanosome will be missed by the H.C.T. and trypanosome density.     

Elevated rates of HIV infection among young Aboriginal injection drug users in a Canadian setting

Objectives  

Recent reports have suggested that Aboriginal and American Indian people are at elevated risk of HIV infection. We undertook the present study to compare socio-demographic and risk variables between Aboriginal and non-Aboriginal young (aged 13 – 24 years) injection drug users (IDUs) and characterize the burden of HIV infection among young Aboriginal IDUs.