Conducting active screening for human African trypanosomiasis with rapid diagnostic tests: The Guinean experience (2016-2021).

Strategies to detect Human African Trypanosomiasis (HAT) cases rely on serological screening of populations exposed to trypanosomes. In Guinea, mass medical screening surveys performed with the Card Agglutination Test for Trypanosomiasis have been progressively replaced by door-to-door approaches using Rapid Diagnostic Tests (RDTs) since 2016. However, RDTs availability represents a major concern and medical teams must often adapt, even in the absence of prior RDT performance evaluation. For the last 5 years, the Guinean HAT National Control Program had to combine three different RDTs according to their availability and price: the SD Bioline HAT (not available anymore), the HAT Sero-K-SeT (most expensive), and recently the Abbott Bioline HAT 2.0 (limited field evaluation). Here, we assess the performance of these RDTs, alone or in different combinations, through the analysis of both prospective and retrospective data. A parallel assessment showed a higher positivity rate of Abbott Bioline HAT 2.0 (6.0%, n = 2,250) as compared to HAT Sero-K-SeT (1.9%), with a combined positive predictive value (PPV) of 20.0%. However, an evaluation of Abbott Bioline HAT 2.0 alone revealed a low PPV of 3.9% (n = 6,930) which was surpassed when using Abbott Bioline HAT 2.0 in first line and HAT Sero-K-SeT as a secondary test before confirmation, with a combined PPV reaching 44.4%. A retrospective evaluation of all 3 RDTs was then conducted on 189 plasma samples from the HAT-NCP biobank, confirming the higher sensitivity (94.0% [85.6-97.7%]) and lower specificity (83.6% [76.0-89.1%]) of Abbott Bioline HAT 2.0 as compared to SD Bioline HAT (Se 64.2% [52.2-74.6%]-Sp 98.4% [94.2-99.5%]) and HAT Sero-K-SeT (Se 88.1% [78.2-93.8%]-Sp 98.4% [94.2-99.5%]). A comparison of Abbott Bioline HAT 2.0 and malaria-RDT positivity rates on 479 subjects living in HAT-free malaria-endemic areas further revealed that a significantly higher proportion of subjects positive in Abbott Bioline HAT 2.0 were also positive in malaria-RDT, suggesting a possible cross-reaction of Abbott Bioline HAT 2.0 with malaria-related biological factors in about 10% of malaria cases. This would explain, at least in part, the limited specificity of Abbott Bioline HAT 2.0. Overall, Abbott Bioline HAT 2.0 seems suitable as first line RDT in combination with a second HAT RDT to prevent confirmatory lab overload and loss of suspects during referral for confirmation. A state-of-the-art prospective comparative study is further required for comparing all current and future HAT RDTs to propose an optimal combination of RDTs for door-to-door active screening.

Analytical sensitivity of loopamp and quantitative real-time PCR on dried blood spots and their potential role in monitoring human African trypanosomiasis elimination.

The objective set by WHO to reach elimination of human African trypanosomiasis (HAT) as a public health problem by 2020 is being achieved. The next target is the interruption of gambiense-HAT transmission in humans by 2030. To monitor progress towards this target, in areas where specialized local HAT control capacities will disappear, is a major challenge. Test specimens should be easily collectable and safely transportable such as dried blood spots (DBS). Monitoring tests performed in regional reference centres should be reliable, cheap and allow analysis of large numbers of specimens in a high-throughput format. The aim of this study was to assess the analytical sensitivity of Loopamp, M18S quantitative real-time PCR (M18S qPCR) and TgsGP qPCR as molecular diagnostic tests for the presence of Trypanosoma brucei gambiense in DBS. The sensitivity of the Loopamp test, with a detection limit of 100 trypanosomes/mL, was in the range of parasitaemias commonly observed in HAT patients, while detection limits for M18S and TgsGP qPCR were respectively 1000 and 10,000 trypanosomes/mL. None of the tests was entirely suitable for high-throughput use and further development and implementation of sensitive high-throughput molecular tools for monitoring HAT elimination are needed.

High Levels of Genetic Diversity within Nilo-Saharan Populations: Implications for Human Adaptation.

Africa contains more human genetic variation than any other continent, but the majority of the population-scale analyses of the African peoples have focused on just two of the four major linguistic groups, the Niger-Congo and Afro-Asiatic, leaving the Nilo-Saharan and Khoisan populations under-represented. In order to assess genetic variation and signatures of selection within a Nilo-Saharan population and between the Nilo-Saharan and Niger-Congo and Afro-Asiatic, we sequenced 50 genomes from the Nilo-Saharan Lugbara population of North-West Uganda and 250 genomes from 6 previously unsequenced Niger-Congo populations. We compared these data to data from a further 16 Eurasian and African populations including the Gumuz, another putative Nilo-Saharan population from Ethiopia. Of the 21 million variants identified in the Nilo-Saharan population, 3.57 million (17%) were not represented in dbSNP and included predicted non-synonymous mutations with possible phenotypic effects. We found greater genetic differentiation between the Nilo-Saharan Lugbara and Gumuz populations than between any two Afro-Asiatic or Niger-Congo populations. F3 tests showed that Gumuz contributed a genetic component to most Niger-Congo B populations whereas Lugabara did not. We scanned the genomes of the Lugbara for evidence of selective sweeps. We found selective sweeps at four loci (SLC24A5, SNX13, TYRP1, and UVRAG) associated with skin pigmentation, three of which already have been reported to be under selection. These selective sweeps point toward adaptations to the intense UV radiation of the Sahel.

Copy number variation in human genomes from three major ethno-linguistic groups in Africa.

BACKGROUND: Copy number variation is an important class of genomic variation that has been reported in 75% of the human genome. However, it is underreported in African populations. Copy number variants (CNVs) could have important impacts on disease susceptibility and environmental adaptation. To describe CNVs and their possible impacts in Africans, we sequenced genomes of 232 individuals from three major African ethno-linguistic groups: (1) Niger Congo A from Guinea and Côte d'Ivoire, (2) Niger Congo B from Uganda and the Democratic Republic of Congo and (3) Nilo-Saharans from Uganda. We used GenomeSTRiP and cn.MOPS to identify copy number variant regions (CNVRs). RESULTS: We detected 7608 CNVRs, of which 2172 were only deletions, 2384 were only insertions and 3052 had both. We detected 224 previously un-described CNVRs. The majority of novel CNVRs were present at low frequency and were not shared between populations. We tested for evidence of selection associated with CNVs and also for population structure. Signatures of selection identified previously, using SNPs from the same populations, were overrepresented in CNVRs. When CNVs were tagged with SNP haplotypes to identify SNPs that could predict the presence of CNVs, we identified haplotypes tagging 3096 CNVRs, 372 CNVRs had SNPs with evidence of selection (iHS > 3) and 222 CNVRs had both. This was more than expected (p < 0.0001) and included loci where CNVs have previously been associated with HIV, Rhesus D and preeclampsia. When integrated with 1000 Genomes CNV data, we replicated their observation of population stratification by continent but no clustering by populations within Africa, despite inclusion of Nilo-Saharans and Niger-Congo populations within our dataset. CONCLUSIONS: Novel CNVRs in the current study increase representation of African diversity in the database of genomic variants. Over-representation of CNVRs in SNP signatures of selection and an excess of SNPs that both tag CNVs and are subject to selection show that CNVs may be the actual targets of selection at some loci. However, unlike SNPs, CNVs alone do not resolve African ethno-linguistic groups. Tag haplotypes for CNVs identified may be useful in predicting African CNVs in future studies where only SNP data is available.

Macrophage migrating inhibitory factor expression is associated with Trypanosoma brucei gambiense infection and is controlled by trans-acting expression quantitative trait loci in the Guinean population.

Infection by Trypanosoma brucei gambiense is characterized by a wide array of clinical outcomes, ranging from asymptomatic to acute disease and even spontaneous cure. In this study, we investigated the association between macrophage migrating inhibitory factor (MIF), an important pro-inflammatory cytokine that plays a central role in both innate and acquired immunity, and disease outcome during T. b. gambiense infection. A comparative expression analysis of patients, individuals with latent infection and controls found that MIF had significantly higher expression in patients (n = 141; 1.25 ±â€¯0.07; p < .0001) and latent infections (n = 25; 1.23 ±â€¯0.13; p = .0005) relative to controls (n = 46; 0.94 ±â€¯0.11). Furthermore, expression decreased significantly after treatment (patients before treatment n = 33; 1.40 ±â€¯0.18 versus patients after treatment n = 33; 0.99 ±â€¯0.10, p = .0001). We conducted a genome wide eQTL analysis on 29 controls, 128 cases and 15 latently infected individuals for whom expression and genotype data were both available. Four loci, including one containing the chemokine CXCL13, were found to associate with MIF expression. Genes at these loci are candidate regulators of increased expression of MIF after infection. Our study is the first data demonstrating that MIF expression is elevated in T. b. gambiense-infected human hosts but does not appear to contribute to pathology.

Description of the first sleeping sickness case diagnosed in Burkina Faso since two decades.

Burkina Faso belongs to a group of countries in which human African trypanosomiasis (HAT), caused by Trypanosoma brucei gambiense, is no longer considered to be a public health problem. Although no native cases have been detected since 1993, there is still the risk of HAT re-emergence due to significant population movements between Burkina Faso and active HAT foci in Côte d'Ivoire. Since 2014, Burkina Faso receives support from the WHO to implement a passive surveillance program. This resulted in the detection in 2015 of the first putative native HAT case since two decades. However, epidemiological entomological and molecular biology investigations have not been able to identify with certainty the origin of this infection or to confirm that it was due to T. b. gambiense. This case emphasises the need to strengthen passive surveillance of the disease for sustained elimination of HAT as a public health problem in Burkina Faso.

The study of trypanosome species circulating in domestic animals in two human African trypanosomiasis foci of Côte d'Ivoire identifies pigs and cattle as potential reservoirs of Trypanosoma brucei gambiense.

BACKGROUND: Important control efforts have led to a significant reduction of the prevalence of human African trypanosomiasis (HAT) in Côte d'Ivoire, but the disease is still present in several foci. The existence of an animal reservoir of Trypanosoma brucei gambiense may explain disease persistence in these foci where animal breeding is an important source of income but where the prevalence of animal African trypanosomiasis (AAT) is unknown. The aim of this study was to identify the trypanosome species circulating in domestic animals in both Bonon and Sinfra HAT endemic foci. METHODOLOGY/PRINCIPAL FINDINGS: 552 domestic animals (goats, pigs, cattle and sheep) were included. Blood samples were tested for trypanosomes by microscopic observation, species-specific PCR for T. brucei sl, T. congolense, T. vivax and subspecies-specific PCR for T. b. gambiense and T. b. gambiense immune trypanolysis (TL). Infection rates varied significantly between animal species and were by far the highest in pigs (30%). T. brucei s.l was the most prevalent trypanosome species (13.7%) followed by T. congolense. No T. b. gambiense was identified by PCR while high TL positivity rates were observed using T. b. gambiense specific variants (up to 27.6% for pigs in the Bonon focus). CONCLUSION: This study shows that domestic animals are highly infected by trypanosomes in the studied foci. This was particularly true for pigs, possibly due to a higher exposure of these animals to tsetse flies. Whereas T. brucei s.l. was the most prevalent species, discordant results were obtained between PCR and TL regarding T. b. gambiense identification. It is therefore crucial to develop better tools to study the epidemiological role of potential animal reservoir for T. b. gambiense. Our study illustrates the importance of "one health" approaches to reach HAT elimination and contribute to AAT control in the studied foci.

Candidate gene polymorphisms study between human African trypanosomiasis clinical phenotypes in Guinea.

BACKGROUND: Human African trypanosomiasis (HAT), a lethal disease induced by Trypanosoma brucei gambiense, has a range of clinical outcomes in its human host in West Africa: an acute form progressing rapidly to second stage, spontaneous self-cure and individuals able to regulate parasitaemia at very low levels, have all been reported from endemic foci. In order to test if this clinical diversity is influenced by host genetic determinants, the association between candidate gene polymorphisms and HAT outcome was investigated in populations from HAT active foci in Guinea. METHODOLOGY AND RESULTS: Samples were collected from 425 individuals; comprising of 232 HAT cases, 79 subjects with long lasting positive and specific serology but negative parasitology and 114 endemic controls. Genotypes of 28 SNPs in eight genes passed quality control and were used for an association analysis. IL6 rs1818879 allele A (p = 0.0001, OR = 0.39, CI95 = [0.24-0.63], BONF = 0.0034) was associated with a lower risk of progressing from latent infection to active disease. MIF rs36086171 allele G seemed to be associated with an increased risk (p = 0.0239, OR = 1.65, CI95 = [1.07-2.53], BONF = 0.6697) but did not remain significant after Bonferroni correction. Similarly MIF rs12483859 C allele seems be associated with latent infections (p = 0.0077, OR = 1.86, CI95 = [1.18-2.95], BONF = 0.2157). We confirmed earlier observations that APOL1 G2 allele (DEL) (p = 0.0011, OR = 2.70, CI95 = [1.49-4.91], BONF = 0.0301) is associated with a higher risk and APOL1 G1 polymorphism (p = 0.0005, OR = 0.45, CI95 = [0.29-0.70], BONF = 0.0129) with a lower risk of developing HAT. No associations were found with other candidate genes. CONCLUSION: Our data show that host genes are involved in modulating Trypanosoma brucei gambiense infection outcome in infected individuals from Guinea with IL6 rs1818879 being associated with a lower risk of progressing to active HAT. These results enhance our understanding of host-parasite interactions and, ultimately, may lead to the development of new control tools.